 
REBASE version 501                                              pubrefsa.501
 
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    REBASE, The Restriction Enzyme Database   http://rebase.neb.com
    Copyright (c)  Dr. Richard J. Roberts, 2014.   All rights reserved.
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Rich Roberts                                                    Dec 30 2014
 

<1> <AUTHORS>
<2> <TITLE>
<3> <JOUR_BOOK/PUBLISHER, EDITOR, CITY/PATENT OFFICE>
<4> <VOLUME/PATENT NUMBER>
<5> <PAGE>
<6> <YEAR>
<7> <ABSTRACT>
<>

<1>Aagaard, C., Awayez, M.J., Garrett, R.A.
<2>Profile of the DNA recognition site of the archaeal homing endonuclease I-DmoI.
<3>Nucleic Acids Res.
<4>25
<5>1523-1530
<6>1997
<7>I-DmoI is a homing enzyme of the LAGLI-DADG type that recognizes up to 20 bp of DNA and is
encoded by an archaeal intron of the hyperthermophilic archaeon Desulfurococcus mobilis.  A
combined mutational and DNA footprinting approach was employed to investigate the specificity
of the I-DmoI-substrate interaction.  The results indicate that the enzyme binds primarily to
short base paired regions that border the sites of DNA cleavage and intron insertion.  The
minimal substrate spans no more than 15 bp and while sequence degeneracy is tolerated in the
DNA binding regions, the sequence and size of the cleavage region is highly conserved.  The
enzyme has a slow turnover rate and cuts the coding strand with a slight preference over the
non-coding strand.  Complex formation produces some distortion of the DNA double helix within
the cleavage region.  The data are compatible with the two DNA-binding domains of I-DmoI
bridging the minor groove, where cleavage occurs, and interacting within the major groove on
either side, thereby stabilizing a distorted DNA double helix.  This may provide a general
mode of DNA interaction at least for the LAGLIDADG-type homing enzymes.

<>

<1>Aagaard, C., Dalgaard, J.Z., Garrett, R.A.
<2>Intercellular mobility and homing of an archaeal rDNA intron confers a selective advantage over intron- cells of Sulfolobus acidocaldarius.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>92
<5>12285-12289
<6>1995
<7>Some intron-containing rRNA genes of archaea encode homing-type endonucleases, which
facilitate intron insertion at homologous sites in intron- alleles. These archaeal rRNA genes,
in contrast to their eukaryotic counterparts, are present in single copies per cell, which
precludes intron homing within one cell. However, given the highly conserved nature of the
sequences flanking the intron, homing may occur in intron- rRNA genes of other archaeal cells.
To test whether this occurs, the intron-containing 23S rRNA gene of the archaeal
hyperthermophile Desulfurococcus mobilis, carried on nonreplicating bacterial vectors, was
electroporated into an intron- culture of Sulfolobus acidocaldarius. PCR experiments
demonstrated that the intron underwent homing and spread through the culture. By using a
double drug-resistant mutant of S. acidocaldarius, it was shown that spreading resulted partly
from a selective advantage of intron+ cells and partly from intercellular mobility of the
intron and homing.

<>

<1>Aapola, U., Kawasaki, K., Scott, H.S., Ollila, J., Vihinen, M., Heino, M., Shintani, A., Kawasaki, K., Minoshima, S., Krohn, K., Antonarakis, S.E., Shimizu, N., Kudoh, J., Peterson, P.
<2>Isolation and initial characterization of a novel zinc finger gene, DNMT3L, on 21q22.3, related to the cytosine-5-methyltransferase 3 gene family.
<3>Genomics
<4>65
<5>293-298
<6>2000
<7>We have isolated the DNMT3L gene that is related to the cytosine-5-methyltransferase 3 (DNMT3)
family. The gene is located on chromosome 21q22.3 between the AIRE and the KIAA0653 genes and
spans approximately 16 kb of genomic sequence. The encoded protein of 387 amino acids has a
cysteine-rich region containing a novel-type zinc finger domain that is conserved in DNMT3A
and DNMT3B but also in ATRX, a member of the SNF2 protein family. The novel domain, called an
ADD (ATRX, DNMT3, DNMT3L)-type zinc finger, contains two subparts: a C2C2 and an imperfect PHD
zinc finger. Expression of the DNMT3L mRNA was not detectable by Northern blotting; however,
RT-PCR amplification revealed that it is expressed at low levels in several tissues including
testis, ovary, and thymus.

<>

<1>Aapola, U., Liiv, I., Peterson, P.
<2>Imprinting regulator DNMT3L is a transcriptional repressor associated with histone deacetylase activity.
<3>Nucleic Acids Res.
<4>30
<5>3602-3608
<6>2002
<7>DNMT3L is a regulator of imprint establishment of normally methylated maternal genomic
sequences. DNMT3L shows high similarity to the de novo DNA methyltransferases, DNMT3A and
DNMT3B, however, the amino acid residues needed for DNA cytosine methyltransferase activity
have been lost from the DNMT3L protein sequence.  Apart from methyltransferase activity,
Dnmt3a and Dnmt3b serve as transcriptional repressors associating with histone deacetylase
(HDAC) activity. Here we show that DNMT3L can also repress transcription by binding directly
to HDAC1 protein. We have identified the PHD-like zinc finger of the ATRX domain as a main
repression motif of DNMT3L, through which DNMT3L recruits the HDAC activity needed for
transcriptional silencing.  Furthermore, we show that DNMT3L protein contains an active
nuclear localisation signal at amino acids 156-159.  These results describe DNMT3L as a
co-repressor protein and suggest that a transcriptionally repressed chromatin organisation
through HDAC activity is needed for establishment of genomic imprints.

<>

<1>Aapola, U., Lyle, R., Krohn, K., Antonarakis, S.E., Peterson, P.
<2>Isolation and initial characterization of the mouse Dnmt3L gene.
<3>Cytogenet. Cell Genet.
<4>92
<5>122-126
<6>2001
<7>We have isolated and sequenced the mouse zinc finger gene, Dnmt3l (DNA
cytosine-5-methyltransferase 3-like), on mouse chromosome 10, showing similarity to members of
the DNMT3/Dnmt3 family. The Dnmt3l protein contains an ADD zinc finger, which Dnmt3l shares
with other Dnmt3 family members and Atrx. RT-PCR analysis showed Dnmt3l expression in testis,
thymus, ovary, and heart, as well as in 7-day, 15-day, and 17-day mouse embryos.

<>

<1>Aapola, U., Maenpaa, K., Kaipia, A., Peterson, P.
<2>Epigenetic modifications affect the Dnmt3L expression.
<3>Biochem. J.
<4>380
<5>705-713
<6>2004
<7>Imprinted genes are expressed from a single allele due to differential methylation of maternal
or paternal alleles during gametogenesis. Dnmt3L, a member of de novo methyltranferase Dnmt3
protein family, is a regulator of maternal imprinting. In the present study, we have
characterised the promoter region of the mouse Dnmt3L gene. Transient transfection assays
performed with 5'-deletion promoter constructs indicated a minimal promoter area within 440
bp upstream from the translational start. Longer promoter constructs showed decreased activity
suggesting the presence of repressor elements within the upstream sequences. According to
electrophoretic mobility shift assays and mutation analysis minimal promoter region contained
four functional binding sites for the Sp1 family of transcription factors, Sp1 and Sp3. In
vitro methylation of Dnmt3L promoter constructs decreased the transcriptional activity
dramatically demonstrating downregulation by cytosine methylation. This was supported by the
results from bisulfite sequencing and quantitative RT-PCR analysis of different mouse cell
lines and tissues. In testis and ES cells showing strong Dnmt3L expression, all studied CpG
sites were fully unmethylated whereas non-expressive cell-lines and tissues with lesser Dnmt3L
expression showed complete or diverse CpG methylation levels. Treatment of Dnmt3L
non-expressive cell lines with deacetylase inhibitor trichostatin A and methyltransferase
inhibitor 5-aza-2'-deoxycytidine induced the expression of Dnmt3L mRNA. Furthermore, we show
that repressional effect of longer promoter fragments was also relieved by these inhibitors,
altogether indicating an epigenetic control for Dnmt3L gene regulation.

<>

<1>Abadjieva, A., Patel, J., Webb, M., Zinkevich, V., Firman, K.
<2>A deletion mutant of the type IC restriction endonuclease EcoR124I expressing a novel DNA specificity.
<3>Nucleic Acids Res.
<4>21
<5>4435-4443
<6>1993
<7>We have developed a complementation assay which allows us to distinguish between mutations
affecting subunit assembly and mutations affecting DNA binding in the DNA recognition subunit
(HsdS) of the multimeric restriction endonuclease EcoR124I. A number of random point mutations
were constructed to test the validity of this assay. Two of the mutants produced were found to
be truncated polypeptides that were still capable of complementation with the EcoR124I Hsd
subunits to give an active restriction enzyme of novel DNA specificity. The N-terminal
variable domain (responsible for recognition of GAA from the EcoR124I recognition sequence
GaannnnnnRTCG) and the spacer region (central conserved region) is intact in both of these
mutants. One of these mutant genes (hsdS(del50)) has been cloned as an active Mtase.
Purification of the Mtase proved to be difficult because the complex is weak. However, Mtase
activity was obtained from a soluble cell extract, and this allowed us to determine the DNA
recognition sequence of the Mtase to be GAAnnnnnnnTTC. This recognition sequence is an
inverted repeat of the 5'-end of the EcoR124I recognition sequence. This suggests that the
mutant Mtase is assembled from two inverted HsdS(D50) subunits, possibly held together by the
HsdM subunits.

<>

<1>Abadjieva, A., Patel, J., Zinkevich, V., Weiserova, M., Firman, K.
<2>The domain structure of the DNA specificity subunit of type I restriction endonucleases.  II.  Mutations affecting subunit assembly.
<3>DNA Transfer and Gene Expression in Microorganisms, Intercept, Balla, E., Berencsi, G., Szentirmai, A., Andover
<4>0
<5>189-196
<6>1993
<7>Type I restriction endonucleases are complex multimeric enzymes comprising three subunits.
HsdM and HsdS are sufficient to produce an active methylase and have the stoichiometry M2S.
To produce an active endonuclease the HsdR subunit is also required along with the cofactors
ATP, Mg2+ and S-adenosylmethionine.  The stoichiometry of the endonuclease is less well
defined and may be influenced by the level of production of the individual subunits.
Classically mutations within the hsdR gene of type I R-M systems produced an R- M+ phenotype,
while those in the hsdM or the hsdS produced an R-M- phenotype.  However, recently we have
described temperature-sensitive mutations within the hsdS and hsdM genes of EcoK that are
altered in their restriction phenotype.  These mutations appear to affect the ability of the
HsdS or HsdM subunits to interact with the HsdR subunit.  In this paper we describe a number
of observations that suggest the assembly of these multi-subunit enzymes may be controlled by
the concentration of the individual subunits.  This type of control is shown to be correct and
it represents a novel method for genetic control of enzyme function, beyond that of the
control of transcription and translation.  We also propose a testable model of how this
control may function within type I R-M systems.

<>

<1>Abadjieva, A., Scarlett, G., Janscak, P., Dutta, C.F., Firman, K.
<2>Characterization of an EcoR1241 restriction-modification enzyme produced from a deleted form of the DNA-binding subunit, which results in a novel DNA specificity.
<3>Folia Microbiol. (Praha)
<4>48
<5>319-328
<6>2003
<7>We purified and characterized both the methyltransferase and the endonuclease containing the
HsdS delta 50 subunit (type I restriction
endonucleases are composed of three subunits--HsdR required for
restriction, HsdM required for methylation and HsdS responsible for DNA
recognition) produced from the deletion mutation hsdS delta 50 of the type
IC R-M system EcoR 124I; this mutant subunit lacks the C-terminal 163
residues of HsdS and produces a novel DNA specificity. Analysis of the
purified HsDs delta 50 subunit indicated that during purification it is
subject to partial proteolysis resulting in removal of approximately 1 kDa
of the polypeptide at the C-terminus. This proteolysis prevented the
purification of further deletion mutants, which were determined as having
a novel DNA specificity in vivo. After biochemical characterization of the
mutant DNA methyltransferase (MTase) and restriction endonuclease we found
only one difference comparing with the wild-type enzyme--a significantly
higher binding affinity of the MTase for the two substrates of
hemimethylated and fully methylated DNA. This indicates that MTase delta
50 is less able to discriminate the methylation status of the DNA during
its binding. However, the mutant MTase still preferred hemimethylated DNA
as the substrate for methylation. We fused the hsdM and hsdS delta 50
genes and showed that the HsdM-HsdS delta 50 fusion protein is capable of
dimerization confirming the model for assembly of this deletion mutant.

<>

<1>Abadjieva, A., Webb, M., Patel, J., Zinkevich, V., Firman, K.
<2>Deletions within the DNA recognition subunit of M.EcoR124I that identify a region involved in protein-protein interactions between HsdS and HsdM.
<3>J. Mol. Biol.
<4>241
<5>35-43
<6>1994
<7>The DNA recognition subunit (HsdS) of type I restriction endonucleases can be divided into
domains by means of amino acid identity between subunits from the same family. It has been
proposed that DNA-protein interactions occur within the variable domains of the subunit and
that protein-protein interactions involve the conserved domains. We have constructed a number
of deletion mutants of HsdS that have allowed us to investigate protein-protein interactions.
Using a combination of a "competitive" complementation assay and the ability of HsdM to
"solubilize" HsdS, we have defined a region within the central conserved domain of HsdS that
is responsible for HsdS-HsdM interaction. Computer analysis of amino acid identity between the
N-terminal half and the C-terminal half of HsdS identifies a region (repeated in both
conserved domains), one copy of which overlaps the region we have identified as essential for
HsdS-HsdM interactions, which may be responsible for such protein-protein interactions.

<>

<1>Abbasalizadeh, S., Salehi, J.G., Motamedi, J.M., Azarbaijani, R., Parsa, Y.L., Ahmad, R.M., Mardi, M., Salekdeh, G.H.
<2>Draft Genome Sequence of Ureibacillus thermosphaericus Strain Thermo-BF, Isolated from Ramsar Hot Springs in Iran.
<3>J. Bacteriol.
<4>194
<5>4431
<6>2012
<7>Ureibacillus thermosphaericus strain Thermo-BF is an aerobic, thermophilic bacillus which has
been characterized to biosynthesize gold nanoparticles. Here
we present the draft genome sequence of Ureibacillus thermosphaericus strain
Thermo-BF which consists of a 2,864,162-bp chromosome. This is the first report
of a shotgun sequenced draft genome of a species in the Ureibacillus genus.

<>

<1>Abbasifar, R., Kropinski, A.M., Sabour, P.M., Ackermann, H.W., Lingohr, E.J., Griffiths, M.W.
<2>Complete Genome Sequence of Cronobacter sakazakii Bacteriophage vB_CsaM_GAP161.
<3>J. Virol.
<4>86
<5>13806-13807
<6>2012
<7>Cronobacter sakazakii is an opportunistic pathogen that causes infant meningitis
and is often associated with milk-based infant formula. We have fully sequenced
the genome of a newly isolated lytic C. sakazakii myovirus, vB_CsaM_GAP161,
briefly named GAP161. It consists of 178,193 bp and has a G+C content of 44.5%. A
total of 277 genes, including 275 open reading frames and two tRNA-encoding
genes, were identified. This phage is closely related to coliphages RB16 and RB43
and Klebsiella pneumoniae phage KP15.

<>

<1>Abdallah, A.M., Rashid, M., Adroub, S.A., Arnoux, M., Ali, S., van Soolingen, D., Bitter, W., Pain, A.
<2>Complete Genome Sequence of Mycobacterium phlei Type Strain RIVM601174.
<3>J. Bacteriol.
<4>194
<5>3284-3285
<6>2012
<7>Mycobacterium phlei is a rapidly growing nontuberculous Mycobacterium species that is
typically nonpathogenic, with few reported cases of human disease. Here
we report the whole genome sequence of M. phlei type strain RIVM601174.

<>

<1>Abdallah, A.M., Rashid, M., Adroub, S.A., Elabdalaoui, H., Ali, S., van Soolingen, D., Bitter, W., Pain, A.
<2>Complete Genome Sequence of Mycobacterium xenopi Type Strain RIVM700367.
<3>J. Bacteriol.
<4>194
<5>3282-3283
<6>2012
<7>Mycobacterium xenopi is a slow-growing, thermophilic, water-related Mycobacterium species.
Like other nontuberculous mycobacteria, M. xenopi more commonly infects
humans with altered immune function, such as chronic obstructive pulmonary
disease patients. It is considered clinically relevant in a significant
proportion of the patients from whom it is isolated. We report here the whole
genome sequence of M. xenopi type strain RIVM700367.

<>

<1>Abdurashitov, M.A., Belichenko, O.A., Lebedeva, N.A., Degtyarev, S.K.
<2>PsiI, a novel restriction endonuclease recognizing the DNA sequence 5'-TTA^TAA-3'.
<3>Biokhimiia
<4>64
<5>574-576
<6>1999
<7>PsiI, a novel restriction endonuclease produced by the bacterial strain Pseudomonas sp.
SE-G49, has been isolated and characterized. The enzyme cleaves DNA in the middle of its
palindromic recognition sequence 5'-TTA^TAA-3'.  Thus, PsiI belongs to a rare group of type
II restriction endonucleases whose recognition sites consist of AT base pairs only.

<>

<1>Abdurashitov, M.A., Belichenko, O.A., Shevchenko, A.V., Dedkov, V.S., Degtyarev, S.K.
<2>BstAPI, an ApaBI isoschizomer, cleaves DNA at 5'-GCANNNN/NTGC-3'.
<3>Nucleic Acids Res.
<4>25
<5>2301-2302
<6>1997
<7>Cleavage positions of BstAPI, a new restriction endonuclease (Enase) that recognizes
palindromic interrupted DNA sequence, have been determined.  Recognition sequences and
cleavage sites comparison shows that BstAPI shares similarity with a number of type II
restriction enzymes.

<>

<1>Abdurashitov, M.A., Belichenko, O.A., Shevchenko, A.V., Degtyarev, S.K.
<2>SimI, a new restriction endonuclease that recognizes non-palindromic sequence 5'-GGGTC-3'(-3/0).
<3>Nucleic Acids Res.
<4>23
<5>2571-2572
<6>1995
<7>SimI, a type II restriction endonuclease, has been isolated from Staphylococcus intermedius 6H
using heparin and hydroxyapatite chromatographic steps. The crude extract contained
approximately 3000 U SimI per gram of cells. Three cleavage positions of SimI on pUC19 DNA
(approximately 999, 1480 and 1768) have been mapped by double digests with enzymes, BglI,
Acc1131I (isoschizomer of ScaI), SalI, PvuI, NruGI (isoschizomer of Eam11051) and Mly1131
(isoschizomer of NarI). A homology search has revealed that the pentanucleotide sequence
5'-GGGTC-3' was located at these positions. The recognition sequence has been confirmed by
comparison of cleavage patterns generated with SimI on commonly used DNAS with computer
predicted ones. The number of SimI sites that occur in lambda, T7, adenovirus-2, pBR322 and
pUC19 DNA, are 40, 94, 73, 8 and 3, respectively. The cleavage points of the restriction
endonuclease SimI have been determined by sequencing of alpha-32P-labelled HindIII-XbaI and
XbaI-EcoRI fragments of the pMVPRL plasmid by the modified Maxam-Gilbert method. The results
indicate the SimI cleaves the DNA sequence shown below: 5'-GG/GTC-3' 3'-CCCAG/-5'.

<>

<1>Abdurashitov, M.A., Belichenko, O.A., Shevchenko, A.V., Degtyarev, S.K.
<2>N.BstSE - site-specific nuclease from Bacillus stearothermophilus SE-589 - restriction endonuclease production.
<3>Mol. Biol. (Mosk)
<4>30
<5>1261-1267
<6>1996
<7>A site-specific nickase recognizing and cleaving
the DNA site 5'-GAGTCNNNN^N-3' was isolated from Bacillus
stearothermophilus SE-589 and named N.BstSE. Its properties
indicate probable relation with type II restriction
endonucleases.

<>

<1>Abdurashitov, M.A., Kileva, E.V., Myakisheva, T.V., Dedkov, V.S., Shevchenko, A.V., Degtyarev, S.K.
<2>AccBSI: A new restriction endonuclease from Acinetobacter calcoaceticus BS.
<3>Prikl. Biokhim. Mikrobiol.
<4>33
<5>556-558
<6>1997
<7>The recognition site of a new restriction endonuclease from Acinetobacter calcoaceticus BS was
determined.  This is a nonpalindromic sequence 5'-GAGCGG-3' 3'-CTCGCC-5.  AccBSI
restrictase cleaves DNA chains in the middle of the recognition sequence; therefore, ligation
of its digestion fragments restored AccBSI recognition sites and generated palindromic
sequences recognized by SacI and SacII restrictases.

<>

<1>Abdurashitov, M.A., Kileva, E.V., Shinkarenko, N.M., Shevchenko, A.V., Dedkov, V.S., Degtyarev, S.K.
<2>BstF5I, an unusual isoschizomer of FokI.
<3>Gene
<4>172
<5>49-51
<6>1996
<7>BstF5, a new restriction endonuclease (Enase) from Bacillus stearothermophilus
F5, has been discovered.  This enzyme recognizes 5'-GGATG-3' and cleaves DNA, generating a
2-base 3' extension: 5'-GGATG NN/-3' / 3'-CCTAC/NN-5'.  BstF5I is an isoschizomer of FokI
and seems to be evolutionarily close to other nonpalindromic-recognizing ENases from
thermophilic bacilli.

<>

<1>Abdurashitov, M.A., Netesova, N.A., Golikova, L.N., Gutorov, V.V., Belavin, P.A., Degtyarev, S.K.
<2>The second methyltransferase of the BstF5I restriction-modification system is homologous to the C-terminal domains of FokI and StsI methylases.
<3>Mol. Biol. (Mosk)
<4>34
<5>87-94
<6>2000
<7>Nucleotide sequence of DNA-methyltransferase gene M. BstF51-2 was determined. This
enzyme is a part of the restriction-modification system of Bacillus stearothermophilus strain
F. On bacterial chromosome, gene bstF51M2 is located behind methylase gene M.BStF51-1 and has
the same orientation. Protein encoded by gene bstF1M-2 contains conservative regions specific
for adenine DNA-methyltransferase of class D12. DNA-methylase M.BstF51-2 is homologous to
C-terminal domains of DNA methylases Fok1 enzymes M.Fok1 and M.Sts1 modifying the second DNA
strand in the recognition site and Sts1 possessing the same recognition sequence. It is shown
that the restriction-modification system BstF51 contains a unique enzyme M.BstF51-1 modifying
the upper strand of the recognized sequence and M.BstF51-2 homologous to C-terminal parts of
enzymes modifying the second DNA strand in the recognition site.

<>

<1>Abdurashitov, M.A., Okhapkina, S.S., Netesova, N.A., Golikova, L.N., Gonchar, D.A., Degtyarev, S.K.
<2>The unique FauI restriction-modification system: Cloning and protein sequence comparisons.
<3>Mol. Biol. (Mosk)
<4>37
<5>619-624
<6>2003
<7>The nucleotide sequence was established for the full-length Flavobacterium aquatile operon
coding for the FauI restriction-modification system. The
operon is unusual in structure and has the gene order control protein
gene-DNA methyltransferase A gene-restriction endonuclease gene-DNA
methyltransferase B gene, other than in the known analogs. The genes are
similarly oriented and overlap. On evidence of sequence analysis, both
methyltransferases are C5 enzymes, the control protein is similar to that
of other restriction-modification systems, and restriction endonuclease is
low-homologous to other enzymes cleaving the DNA upper strand in position
4 or 5 relative to the recognition site.

<>

<1>Abdurashitov, M.A., Tomilov, V.N., Chernukhin, V.A., Gonchar, D.A., Degtyarev, S.K.
<2>Comparative analysis of human chromosomal DNA digestion with restriction endonucleases in vitro and in silico.
<3>Medical Genetics
<4>6
<5>29-36
<6>2007
<7>Theoretical analysis of human chromosomal DNA cleavage at 15 nucleotide sequences, which are
the recognition sites of various restriction endonucleases, has been carried out.
Distribution diagrams of calculated DNA fragments have been constructed based on earlier
proposed method of mammalian genomes digestion in silico.  A similar study of human Alu- and
LINE1-repeats nucleotide sequences, which are present in informational databases, has been
performed and corresponding diagrams of DNA fragments distribution have been plotted.
Distribution diagrams of chromosomal DNA digestion, which results in formation of low
molecular weight DNA fragments, correspond to those for Alu-repeats; whereas the digestion,
which results in formation of large molecular weight DNA fragments - are similar to those for
LINE-repeats.  All theoretical data have been compared to experimental patterns of human
genomic DNA cleavages with respective restriction endonucleases and a good correspondence for
the most of DNA diagrams has been observed.

<>

<1>Abicht, H.K., Mancini, S., Karnachuk, O.V., Solioz, M.
<2>Genome Sequence of Desulfosporosinus sp. OT, an Acidophilic Sulfate-Reducing Bacterium from Copper Mining Waste in Norilsk, Northern  Siberia.
<3>J. Bacteriol.
<4>193
<5>6104-6105
<6>2011
<7>We have sequenced the genome of Desulfosporosinus sp. OT, a Gram-positive, acidophilic
sulfate-reducing Firmicute isolated from copper tailing
sediment in the Norilsk mining-smelting area in Northern Siberia, Russia.
This represents the first sequenced genome of a Desulfosporosinus species.
The genome has a size of 5.7 Mb and encodes 6,222 putative proteins.

<>

<1>Abrahante, J.E., Johnson, T.J., Hunter, S.S., Maheswaran, S.K., Hauglund, M.J., Bayles, D.O., Tatum, F.M., Briggs, R.E.
<2>Draft Genome Sequences of Two Virulent Serotypes of Avian Pasteurella multocida.
<3>Genome Announcements
<4>1
<5>e00058-12
<6>2013
<7>Here we report the draft genome sequences of two virulent avian strains of Pasteurella
multocida. Comparative analyses of these genomes were done with the
published genome sequence of avirulent P. multocida strain Pm70.

<>

<1>Abrahante, J.E., Veeregowda, B.M., Hogtapur, S.S., Briggs, R.E., Maheswaran, S.K., Sreevatsan, S.
<2>Draft Genome Sequences of Two Pasteurella multocida Strains Isolated from Buffaloes in India with Hemorrhagic Septicemia Disease.
<3>Genome Announcements
<4>2
<5>e00798-14
<6>2014
<7>Pasteurella multocida serotype B:2 is the causative agent of hemorrhagic septicemia in cattle
and buffaloes in Asia. It is an acute fatal disease and is
considered one of the most economically important diseases in this region of the
world. We present here the draft genome sequences of strains 2213 and 3213 of P.
multocida.

<>

<1>Abriouel, H., Benomar, N., Perez, P.R., Canamero, M.M., Galvez, A.
<2>Annotated genome sequence of Lactobacillus pentosus MP-10, which has probiotic potential, from naturally fermented Alorena green table olives.
<3>J. Bacteriol.
<4>193
<5>4559-4560
<6>2011
<7>Lactobacillus pentosus MP-10 was isolated from brines of naturally-fermented Alorena green
table olives. MP-10 has potential probiotic traits: inhibition of human pathogenic bacteria,
survival at low pH (1.5) and bile slat tolerance (3%). Here, we report for the first time the
annotated genome sequence of Lb. pentosus.

<>

<1>Abrol, S., Chaudhary, V.K.
<2>Excess PCR primers inhibit DNA cleavage by some restriction endonucleases.
<3>Biotechniques
<4>15
<5>630-632
<6>1993
<7>For cloning DNA segments into expression vectors, restriction endonuclease recognition sites
can be generated by polymerase chain reaction utilizing synthetic oligonucleotides as PCR
primers that contain the relevant sites.  Following PCR amplification, there usually remains a
10-fold to 100-fold molar excess of unused primers over the desired amplified product in the
reaction mixture.  These excess oligonucleotides may inhibit the cleavage of amplified
products by the restriction enzyme and thus interfere in the generation of DNA fragments with
compatible termini for cloning into expression vectors.  We have investigated the effect of
single-stranded oligonucleotides containing a restriction endonuclease recognition site on the
digestion of linearized double-stranded DNA substrates (mimicking PCR product) by the
respective restriction enzymes.  This information will be useful in deciding if the removal of
excess oligonucleotide primers is necessary or not.

<>

<1>Abrosimova, L., Monakhova, M., Schierling, B., Volkov, E., Romanova, E., Wende, W., Pingoud, A., Kubareva, E., Oretskaya, T.
<2>Light dependent activity of restriction endonucleases.
<3>FEBS J.
<4>280
<5>597
<6>2013
<7>Sequence-specific endonucleases with extended recognition sites can cleave a unique site in
complex genomes. Since the nucleases can show off-target cleavage activity, spatio-temporal
control of their activity is necessary for the precise genome engineering.  It would be
desirable to control the enzymatic activity by an external signal, e.g. by light.  Such
photoregulation is based on the azobenzene 'photoswitch'.  Azobenzene isomerizes between the
extended trans- and the cis-configuration by illumination with UV (trans-cis) or blue light
(cis-trans) as well as by thermal relaxation (cis-trans).  We are developing the 'molecular
gate's strategy based on the fact that most type II restriction endonucleases are homodimers.
The DNA-binding center is located in the interface between the two subunits.  It is possible
to modify the protein at the entrance of the DNA-binding site and block its activity.  To
create the obstacle for DNA penetration to the active center we suggest to use the ability of
oligonucleotides containing azobenzene insertion to form a duplex.  Azobenzene in
trans-configuration stabilizes the duplex and cis-configuration causes destabilization.  Thus
formation and dissociation of the duplex can be reversibly photo-regulated.  The strategy is
illustrated for the type II RE SsoII.  To choose the optimal length of the duplex 10-mer and
15-mer modified oligonucleotides were synthesized.  After attachment to the protein these
oligonucleoties are supposed to form 10-mer DNA duplexes.  The initial rates of DNA cleavage
at 37oC upon UV-illumination was two times higher than upon blue light illumination.  Our
results demonstrate the possibility of changing the enzymatic activity upon illumination. This
work was supported by the program 'International Research Training Groups' (grants RFBR-DFG
11-04-91338 and GRK 1384).

<>

<1>Abrosimova, L.A., Monakhova, M.V., Migur, A.Yu., Wolfgang, W., Pingoud, A., Kubareva, E.A., Oretskaya, T.S.
<2>Thermo-switchable Activity of the Restriction Endonuclease SsoII Achieved by Site-Directed Enzyme Modification.
<3>Life
<4>65
<5>1012-1016
<6>2013
<7>In this work, the possibility of constructing a thermo-switchable enzyme according to the
molecular gate strategy is demonstrated. The approach is based on the covalent attachment of
oligodeoxyribonucleotides to cysteine residues of an enzyme adjacent to its active center to
form a temporal barrier for enzyme-substrate complex formation. The activity of the modified
enzyme that had been studied herethe restriction endonuclease SsoII (R.SsoII)was diminished by
a factor of 180 at 25 degrees C that alm st abolished the enzymatic activity when compared
with the unmodified enzyme. However, heating of the modified enzyme to 45 degrees C resulted
in a 30-fold increase of activity. The activity of unmodified R.SsoII also increased on
heating from 25 to 45 degrees; however, the difference did not exceed a factor of 3-4. The
changes in enzymatic activity observed were shown to be reversible for both the unmodified and
the modified R.SsoII. Variation of the length and the sequence of the attached oligodeoxyribon
ucleotides might allow greater modulation of the activity of DNA-protein conjugates.

<>

<1>Abt, B. et al.
<2>Complete genome sequence of the termite hindgut bacterium Spirochaeta coccoides type strain (SPN1(T)), reclassification in the genus Sphaerochaeta as  Sphaerochaeta coccoides comb. nov. and emendations of the family Spirochaetaceae   and the genus Sphaer.
<3>Standards in Genomic Sciences
<4>6
<5>194-209
<6>2012
<7>Spirochaeta coccoides Droge et al. 2006 is a member of the genus Spirochaeta Ehrenberg 1835,
one of the oldest named genera within the Bacteria. S. coccoides
is an obligately anaerobic, Gram-negative, non-motile, spherical bacterium that
was isolated from the hindgut contents of the termite Neotermes castaneus. The
species is of interest because it may play an important role in the digestion of
breakdown products from cellulose and hemicellulose in the termite gut. Here we
provide a taxonomic re-evaluation for strain SPN1(T), and based on physiological
and genomic characteristics, we propose its reclassification as a novel species
in the genus Sphaerochaeta, a recently published sister group of the Spirochaeta.
The 2,227,296 bp long genome of strain SPN1(T) with its 1,866 protein-coding and
58 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Abt, B. et al.
<2>Complete genome sequence of Leadbetterella byssophila type strain (4M15).
<3>Standards in Genomic Sciences
<4>4
<5>2-12
<6>2011
<7>Leadbetterella byssophila Weon et al. 2005 is the type species of the genus Leadbetterella of
the family Cytophagaceae in the phylum Bacteroidetes. Members
of the phylum Bacteroidetes are widely distributed in nature, especially in
aquatic environments. They are of special interest for their ability to degrade
complex biopolymers. L. byssophila occupies a rather isolated position in the
tree of life and is characterized by its ability to hydrolyze starch and
gelatine, but not agar, cellulose or chitin. Here we describe the features of
this organism, together with the complete genome sequence, and annotation. L.
byssophila is already the 16(th) member of the family Cytophagaceae whose genome
has been sequenced. The 4,059,653 bp long single replicon genome with its 3,613
protein-coding and 53 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Abt, B. et al.
<2>Complete genome sequence of Cellulophaga algicola type strain (IC166).
<3>Standards in Genomic Sciences
<4>4
<5>72-80
<6>2011
<7>Cellulophaga algicola Bowman 2000 belongs to the family Flavobacteriaceae within  the phylum
'Bacteroidetes' and was isolated from Melosira collected from the
Eastern Antarctic coastal zone. The species is of interest because its members
produce a wide range of extracellular enzymes capable of degrading proteins and
polysaccharides with temperature optima of 20-30 degrees C. This is the first
completed genome sequence of a member of the genus Cellulophaga. The 4,888,353 bp
long genome with its 4,285 protein-coding and 62 RNA genes consists of one
circular chromosome and is a part of the Genomic Encyclopedia of Bacteria and
Archaea project.

<>

<1>Abt, B. et al.
<2>Complete genome sequence of Cellulomonas flavigena type strain (134).
<3>Standards in Genomic Sciences
<4>3
<5>15-25
<6>2010
<7>Cellulomonas flavigena (Kellerman and McBeth 1912) Bergey et al. 1923 is the type species of
the genus Cellulomonas of the actinobacterial family
Cellulomonadaceae. Members of the genus Cellulomonas are of special interest for
their ability to degrade cellulose and hemicellulose, particularly with regard to
the use of biomass as an alternative energy source. Here we describe the features
of this organism, together with the complete genome sequence, and annotation.
This is the first complete genome sequence of a member of the genus Cellulomonas,
and next to the human pathogen Tropheryma whipplei the second complete genome
sequence within the actinobacterial family Cellulomonadaceae. The 4,123,179 bp
long single replicon genome with its 3,735 protein-coding and 53 RNA genes is
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Abt, B. et al.
<2>Genome sequence of the thermophilic fresh-water bacterium Spirochaeta caldaria type strain (H1(T)), reclassification of Spirochaeta caldaria, Spirochaeta  stenostrepta, and Spirochaeta zuelzerae in the genus Treponema as Treponema  caldaria comb. nov., T.
<3>Standards in Genomic Sciences
<4>8
<5>88-105
<6>2013
<7>Spirochaeta caldaria Pohlschroeder et al. 1995 is an obligately anaerobic, spiral-shaped
bacterium that is motile via periplasmic flagella. The type strain,
H1(T), was isolated in 1990 from cyanobacterial mat samples collected at a
freshwater hot spring in Oregon, USA, and is of interest because it enhances the
degradation of cellulose when grown in co-culture with Clostridium thermocellum.
Here we provide a taxonomic re-evaluation for S. caldaria based on phylogenetic
analyses of 16S rRNA sequences and whole genomes, and propose the
reclassification of S. caldaria and two other Spirochaeta species as members of
the emended genus Treponema. Whereas genera such as Borrelia and Sphaerochaeta
possess well-distinguished genomic features related to their divergent
lifestyles, the physiological and functional genomic characteristics of
Spirochaeta and Treponema appear to be intermixed and are of little taxonomic
value. The 3,239,340 bp long genome of strain H1(T) with its 2,869 protein-coding
and 59 RNA genes is a part of the G enomic E ncyclopedia of Bacteria and Archaea
project.

<>

<1>Abulencia, C.B., Wyborski, D.L., Garcia, J.A., Podar, M., Chen, W., Chang, S.H., Chang, H.W., Watson, D., Brodie, E.L., Hazen, T.C., Keller, M.
<2>Environmental whole-genome amplification to access microbial populations in contaminated sediments.
<3>Appl. Environ. Microbiol.
<4>72
<5>3291-3301
<6>2006
<7>Low-biomass samples from nitrate and heavy metal contaminated soils yield DNA amounts that
have limited use for direct, native analysis and
screening. Multiple displacement amplification (MDA) using phi29 DNA
polymerase was used to amplify whole genomes from environmental,
contaminated, subsurface sediments. By first amplifying the genomic DNA
(gDNA), biodiversity analysis and gDNA library construction of microbes
found in contaminated soils were made possible. The MDA method was
validated by analyzing amplified genome coverage from approximately five
Escherichia coli cells, resulting in 99.2% genome coverage. The method was
further validated by confirming overall representative species coverage
and also an amplification bias when amplifying from a mix of eight known
bacterial strains. We extracted DNA from samples with extremely low cell
densities from a U.S. Department of Energy contaminated site. After
amplification, small-subunit rRNA analysis revealed relatively even
distribution of species across several major phyla. Clone libraries were
constructed from the amplified gDNA, and a small subset of clones was used
for shotgun sequencing. BLAST analysis of the library clone sequences
showed that 64.9% of the sequences had significant similarities to known
proteins, and "clusters of orthologous groups" (COG) analysis revealed
that more than half of the sequences from each library contained sequence
similarity to known proteins. The libraries can be readily screened for
native genes or any target of interest. Whole-genome amplification of
metagenomic DNA from very minute microbial sources, while introducing an
amplification bias, will allow access to genomic information that was not
previously accessible. The reported SSU rRNA sequences and library clone
end sequences are listed with their respective GenBank accession numbers,
DQ 404590 to DQ 404652, DQ 404654 to DQ 404938, and DX 385314 to DX
389173.

<>

<1>Accetto, T., Peterka, M., Avgustin, G.
<2>Type II restriction modification systems of Prevotella bryantii TC1-1 and Prevotella ruminicola 23 strains and their effect on the efficiency  of DNA introduction via electroporation.
<3>FEMS Microbiol. Lett.
<4>247
<5>177-183
<6>2005
<7>The restriction endonucleases PbrTI and Pru2I, isoschizomers of Sau3AI and HaeIII, were
partially purified and characterized from anaerobic
rumen bacteria Prevotella bryantii TCl-1 and Prevotella ruminicola 23,
respectively. These are the first type II restriction endonucleases
discovered in strains of the genus Prevotella, and they represent one
of the barriers hindering gene transfer in these microorganisms.
Heterologous DNA was protected against the action of PbrTI or Pru2I
by incubation in a cell-free extract of the respective strain which
contained 20 mM EDTA. This led to the development of a protocol
enabling successful electrotransformation of the P. bryantii TCl-1
strain with a pRH3 Bacteroides-Escherichia coli shuttle vector
containing up to 7-kb long DNA inserts. Plasmid DNA isolated from the
transformed strain facilitated the transfer with further increased
efficiency and made possible the introduction of ligation reaction
products directly to P. bryantii TCl-1 without passing them first
through E. coli.

<>

<1>Acharya, A.S., Roy, K.B.
<2>Reduced activity of BamHI variants C54I, C64W, and C54D/C64R is consistent with the substrate-assisted catalysis model.
<3>Biochem. Biophys. Res. Commun.
<4>287
<5>153-159
<6>2001
<7>Three specific mutants, C54I, C54W, and a double-mutant C54D:C64R of restriction endonuclease
BamHI, were generated and studied to investigate the role, if any, of the 54th and 64th
cysteine residues in the catalysis of BamHI. The mutation was achieved using the megaprimer
approach for PCR. The mutant genes were cloned and characterized by sequencing. The mutant and
the wild-type proteins were expressed and purified and their kinetic parameters were
determined using short synthetic oligonucleotides as substrates. All mutants had higher K(m)
values than that of the wild-type enzyme suggesting a decrease in the affinity of the enzyme
for its substrate. The mutant protein C54W showed significant changes in the CD spectra
vis-a-vis wild-type enzyme and had the lowest K(m)/K(cat) value among the mutants indicative
of changes in the secondary structure of the protein. The melting curves of the mutant
proteins overlapped that of the wild-type enzyme. Analysis of the K(cat) values in the context
of cocrystal structure suggests that the effect of Cys54 mutation is probably through the
perturbation of the local structure whereas reduced activity of the double mutant is
consistent with the substrate-assisted catalysis mechanism.

<>

<1>Acharya, A.S., Roy, K.B.
<2>An alternative approach for screening active Bam HI variants: Overexpression in T-7 RNA polymerase based system.
<3>Indian J. Biochem. Biophys.
<4>38
<5>303-308
<6>2001
<7>The type II restriction endonuclease, BamHI, has been overexpressed in E. coli by cloning the
BamHI gene in frame with an E. coli Ribosome
Binding Site (RBS) under the T7 promoter of an E. coli expression
vector pRSET A. The expression level of BamHI endonuclease using this
construct was found to be higher than that reported of the
overexpressing clone pAEK14. Our overexpressing clone, pAABRw in BL21
cells in presence of BamHI methylase in pMAP6 following induction with
IPTG yields about 9.2x10(6) units per gram wet cell paste. In vivo
activity of the recombinant endonuclease could be confirmed by the SOS
induction assay in JH139 cells even in the absence of T7 polymerase and
cognate BamHI methylase because of leaky expression in E.coli. This
provides an alternate way to screen the active endonuclease and its
variants.

<>

<1>Adam, Z., Tambong, J.T., Chen, Q., Lewis, C.T., Levesque, C.A., Xu, R.
<2>Draft Genome Sequence of Pseudomonas sp. Strain 2-92, a Biological Control Strain Isolated from a Field Plot Under Long-Term Mineral Fertilization.
<3>Genome Announcements
<4>2
<5>e01121-13
<6>2014
<7>Pseudomonas sp. strain 2-92, isolated from a Canadian field plot under long-term  mineral
fertilization, strongly inhibits the growth of Fusarium graminearum,
Rhizoctonia solani, and Gaeumannomyces graminis. Here, we report the draft genome
sequence of Pseudomonas sp. strain 2-92.

<>

<1>Adam, Z., Tambong, J.T., Lewis, C.T., Levesque, C.A., Chen, W., Bromfield, E.S., Khan, I.U., Xu, R.
<2>Draft Genome Sequence of Pantoea ananatis Strain LMG 2665T, a Bacterial Pathogen  of Pineapple Fruitlets.
<3>Genome Announcements
<4>2
<5>e00489-14
<6>2014
<7>We report the draft genome sequence of Pantoea ananatis LMG 2665(T), the bacterial causal
agent of pineapple fruitlet rot.

<>

<1>Adam, Z., Whiteduck-Leveillee, K., Cloutier, M., Chen, W., Lewis, C.T., Levesque, C.A., Topp, E., Lapen, D.R., Tambong, J.T., Talbot, G., Khan, I.U.
<2>Draft genome sequences of two arcobacter strains isolated from human feces.
<3>Genome Announcements
<4>2
<5>e00113-14
<6>2014
<7>Arcobacter species are members of the family Campylobacteraceae and are considered emerging
enteropathogens and potential zoonotic agents. Here, we report the draft genome sequences of
two Arcobacter strains isolated from human feces in an effort to provide further genetic
resources for understanding the pathogenic dynamics and diversity of this important genus.

<>

<1>Adam, Z., Whiteduck-Leveillee, K., Cloutier, M., Chen, W., Lewis, C.T., Levesque, C.A., Topp, E., Lapen, D.R., Tambong, J.T., Talbot, G., Khan, I.U.
<2>Draft Genome Sequence of Arcobacter cibarius Strain LMG21996T, Isolated from Broiler Carcasses.
<3>Genome Announcements
<4>2
<5>e00034-14
<6>2014
<7>
<>

<1>Adam, Z., Whiteduck-Leveillee, K., Cloutier, M., Tambong, J.T., Chen, W., Lewis, C.T., Levesque, C.A., Topp, E., Lapen, D.R., Talbot, G., Khan, I.U.
<2>Draft genome sequences of three arcobacter strains of pig and dairy cattle manure origin.
<3>Genome Announcements
<4>2
<5>e00377-14
<6>2014
<7>The genus Arcobacter has been associated with human illness and fecal contamination by humans
and animals. Here, we announce the draft genome sequences
of three strains of Arcobacter species cultured from pig and dairy cattle manure
tanks. This information will assist in the characterization of features related
to host specificities and identify potential pathogenic health risks to humans
and animals.

<>

<1>Adamczyk-Poplawska, M., Kondrzycka, A., Urbanek, K., Piekarowicz, A.
<2>Tetra-amino-acid tandem repeats are involved in HsdS complementation in type IC restriction-modification systems.
<3>Microbiology
<4>149
<5>3311-3319
<6>2003
<7>All known type I restriction and modification (R-M) systems of Escherichia coli and Salmonella
enterica belong to one of four discrete families: type
IA, IB, IC or ID. The classification of type I systems from a wide range
of other genera is mainly based on complementation and molecular evidence
derived from the comparison of the amino acid similarity of the
corresponding subunits. This affiliation was seldom based on the strictest
requirement for membership of a family, which depends on relatedness as
demonstrated by complementation tests. This paper presents data indicating
that the type I NgoAV R-M system from Neisseria gonorrhoeae, despite the
very high identity of HsdM and HsdR subunits with members of the type IC
family, does not show complementation with E. coli type IC R-M systems.
Sequence analysis of the HsdS subunit of several different potential type
IC R-M systems shows that the presence of different tetra-amino-acid
sequence repeats, e.g. TAEL, LEAT, SEAL, TSEL, is characteristic for type
IC R-M systems encoded by distantly related bacteria. The other regions of
the HsdS subunits potentially responsible for subunit interaction are also
different between a group of distantly related bacteria, but show high
similarity within these bacteria. Complementation between the NgoAV R-M
system and members of the EcoR124 R-M family can be restored by changing
the tetra-amino-acid repeat within the HsdS subunit. The authors propose
that the type IC family of R-M systems could consist of several
complementation subgroups whose specificity would depend on differences in
the conserved regions of the HsdS polypeptide.

<>

<1>Adamczyk-Poplawska, M., Lower, M., Piekarowicz, A.
<2>Characterization of the NgoAXP: phase-variable type III restriction-modification system in Neisseria gonorrhoeae.
<3>FEMS Microbiol. Lett.
<4>300
<5>25-35
<6>2009
<7>Methyltransferases associated with type III restriction-modification (RM) systems are
phase-variably expressed in a variety of pathogenic
bacteria. NgoAXP, the type III RM system encoded by Neisseria
gonorrhoeae, was characterized in this study. The cloned resngoAXP and
ngoAXPmod genes were expressed in Escherichia coli strains. The
restriction and modification activities of NgoAXP were confirmed in
vivo by the lambda phage restriction and modification test and in vitro
by the methylation of DNA substrates in the presence of
[methyl-3H]AdoMet. As in all known type III systems, the restriction
activity needed the presence of both genes, while the presence of the
ngoAXPmod gene was sufficient for DNA methylation. Following its
overexpression, the DNA methyltransferase M.NgoAXP was purified to
apparent homogeneity using metal affinity chromatography. The specific
sequence recognized by this enzyme was determined as a nonpalindromic
sequence: 5'-CCACC-3', in which the adenine residue is methylated. We
observed that in E. coli cells, the expression of the restriction
phenotype associated with NgoAXP switched randomly. This phase
variation was associated with the change in the number of
pentanucleotide repeats (5'-CCAAC/G-3') present at the 5'-end of the
coding region of the ngoAXPmod gene.

<>

<1>Adamczyk-Poplawska, M., Lower, M., Piekarowicz, A.
<2>Deletion of One Nucleotide within the Homonucleotide Tract Present in the hsdS Gene Alters the DNA Sequence Specificity of Type I  Restriction-Modification System NgoAV.
<3>J. Bacteriol.
<4>193
<5>6750-6759
<6>2011
<7>As a result of a frameshift mutation, the hsdS locus of the NgoAV type IC restriction and
modification (RM) system comprises two genes, hsdS(NgoAV1)
and hsdS(NgoAV2). The specificity subunit, HsdS(NgoAV), the product of the
hsdS(NgoAV1) gene, is a naturally truncated form of an archetypal
specificity subunit (208 N-terminal amino acids instead of 410). The
presence of a homonucleotide tract of seven guanines (poly[G]) at the 3'
end of the hsdS(NgoAV1) gene makes the NgoAV system a strong candidate for
phase variation, i.e., stochastic addition or reduction in the guanine
number. We have constructed mutants with 6 guanines instead of 7 and
demonstrated that the deletion of a single nucleotide within the 3' end of
the hsdS(NgoAV1) gene restored the fusion between the hsdS(NgoAV1) and
hsdS(NgoAV2) genes. We have demonstrated that such a contraction of the
homonucleotide tract may occur in vivo: in a Neisseria gonorrhoeae
population, a minor subpopulation of cells appeared to have only 6
guanines at the 3' end of the hsdS(NgoAV1) gene. Escherichia coli cells
carrying the fused gene and expressing the NgoAVDelta RM system were able
to restrict lambda phage at a level comparable to that for the wild-type
NgoAV system. NgoAV recognizes the quasipalindromic interrupted sequence
5'-GCA(N(8))TGC-3' and methylates both strands. NgoAVDelta recognizes DNA
sequences 5'-GCA(N(7))GTCA-3' and 5'-GCA(N(7))CTCA-3', although the latter
sequence is methylated only on the complementary strand within the
5'-CTCA-3' region of the second recognition target sequence.

<>

<1>Adams, G.M.
<2>Preventing autorestriction: A functional analysis of the pvuIIM and pvuIIW gene products.
<3>Diss. Abstr.
<4>57
<5>108
<6>1996
<7>Restriction-modification systems must be regulated to prevent lethal
autorestriction of the bacterial DNA.  Little is known about this regulation.  This
dissertation examines several ways in which the restriction is controlled.  To date, the
PvuII restriction-modification system had been understood to contain three genes coding
for a DNA methyltransferase, a restriction endonuclease and a protein required for
endonuclease expression.  There is a fourth open reading frame (ORF) within and opposite
to the methyltransferase gene.  This ORF is transcribed  and has also been found in the
SmaI restriction-modification system.  The sequence of the PvuII ORF resembles that of
the PvuII endonuclease dimer interface.  Cells carrying clones of the ORF along with the
intact restriction modification systems had decreased ability to restrict.  The synthetic
peptide corresponding to this ORF inhibited renaturation of urea-denatured endonuclease in
a concentration-dependent manner.  The ORF, named pvuIIW may delay appearance of
endonuclease activity by increasing the concentration needed for dimerization, giving the
methyltransferase additional time to protect the bacterial DNA.  A kinetic anlaysis of PvuII
methyltransferase is the first reported for an N4-methylcytosine methyltransferase.  It
revealed an ordered Bi Bi mechanism, with the preferred order of substrate binding as
DNA first followed by S-adenosyl-L-methionine (AdoMet).  The enzyme was found to
have a kcat of 0.07s-1, KmAdoMet was 4.2 uM and the KmDNA was 0.58 uM.  When the
methyltransferase was preincubated with [3H-CH3]-AdoMet, the expected burst of product
formation resulted when DNA was added.  However, preincubation of the
methyltransferase with DNA gave a 3-5 second lag in product formation after the addition
of AdoMet.  This suggests that the enzyme forms a catalytically-incompetent, dead-end
complex with DNA in the absence of AdoMet.  The methyltransferase was also found to
bind two molecules of the substrate AdoMet, each with a distinct affinity.  A kinetic model
was proposed to explain formation of the dead-end complex with DNA in the absence of
AdoMet, as well as a role for the second bound molecule of AdoMet.

<>

<1>Adams, G.M., Blumenthal, R.M.
<2>Gene pvuIIW: a possible modulator of PvuII endonuclease subunit association.
<3>Gene
<4>157
<5>193-199
<6>1995
<7>The PvuII restriction-modification system has been found to contain three genes which code for
a DNA methyltransferase (MTase), a restriction endonuclease (ENase) and a small protein
required for expression of the ENase-encoding gene.  In addition, there is a small open
reading frame (ORF) within and opposite to the MTase-encoding gene.  The region containing
this ORF is transcribed, and the ORF has an excellent Shine-Dalgarno sequence with an ATA
start codon.  A closely related ORF is present in the SmaI system.  The 28-amino-acid (aa)
predicted peptide from the PvuII ORF resembles a region of the PvuII ENase at the dimer
interface.  We have cloned this ORF, giving it an ATG start codon and putting it under the
control of an inducible promoter: induction leads to a slight but significant decrease in
restriction of bacteriophage lambda.  We also have obtained the 28-aa synthetic peptide, and
are exploring the possibility that it modulates ENase subunit association.  While this peptide
has no detectible effect on dimeric PvuII ENase, it inhibits renaturation of urea-denatured
ENase in a concentration-dependent manner.  The ORF may represent an additional safeguard
during establishment of the PvuII restriction-modification system in a new host cell, helping
to delay the appearance of active ENase dimers, while the MTase accumulates and protects the
host chromosome.

<>

<1>Adams, G.M., Blumenthal, R.M.
<2>The PvuII DNA (cytosine-N4)-methyltransferase comprises two trypsin-defined domains, each of which binds a molecule of S-adenosyl-L-methionine.
<3>Biochemistry
<4>36
<5>8284-8292
<6>1997
<7>Earlier studies have shown that PvuII methyltransferase is monomeric and transfers a methyl
group from S-adenosyl-L-methionine to cytosine, generating N4-methylcytosine in duplex
5'-CAGCTG-3' DNA.  This study examines the interactions between PvuII methyltransferase and
AdoMet.  Trypsin preferentially cleaved the protein into two large fragments, with initial
cleavages after Arg183 and Lys186.  UV-mediated photochemical labeling with [3H-CH3]AdoMet,
followed by trypsin digestion, revealed that both large fragments of the protein were labeled.
Rapid gel filtration confirmed that each molecule of the intact enzyme bound two molecules of
AdoMet (net Kd = 9.3 microM).  When PvuII methyltransferase was preincubated with a range of
[3H-CH3}AdoMet concentrations, bursts of product formation resulted upon DNA addition.  These
data indicate that PvuII methyltransferase is catalytically competent with one and with two
bound molecules of AdoMet.  These results, together with those from earlier studies, suggest
possible roles for the second molecule of AdoMet.

<>

<1>Adams, G.M., Blumenthal, R.M.
<2>What is the significance of the internal duplication in the PvuII DNA methyltransferase?
<3>FASEB J.
<4>9
<5>A1399
<6>1995
<7>The DNA methyltransferases (MTases) from type II restriction-modification systems are
suggested to have evolved via a gene duplication mechanism. The evidence for this varies in
quality and, in most MTases, any duplication which may have occurred has been obscured by
divergence. The MTase gene from the PvuII restriction-modification system was previously
cloned, sequenced and overexpressed in this laboratory. It contains an unusually clear
sequence duplication, consistent with earlier gene duplication. We have studied the
proteolytic susceptibility of PvuII MTase, and its interaction with the substrate methyl donor
S-adenosylmethionine (AdoMet), to investigate the possibility that this monomeric MTase is
functionally symmetrical. We have found that trypsin preferentially cleaves in the predicted
hinge region between the sequence repeats, and that both parts of the protein can be
crosslinked by UV irradiation to [3H-CH3]AdoMet.

<>

<1>Adams, K.L., Clements, M.J., Vaughn, J.C.
<2>The Peperomia mitochondrial coxI group I intron: Timing of horizontal transfer and subsequent evolution of the intron.
<3>J. Mol. Evol.
<4>46
<5>689-696
<6>1998
<7>The Peperomia polybotrya coxI gene intron is the only currently reported group I intron in a
vascular plant mitochondrial genome and it likely originated by horizontal transfer from a
fungal donor.  We provide a clearer picture of the horizontal transfer and a portrayal of the
evolution of the group I intron since it was gained by the Peperomia mitochondrial genome.
The intron was transferred recently in terms of plant evolution, being restricted to the
single genus Peperomia among the order Piperales.  Additional support is presented for the
suggestion that a recombination/repair mechanism was used by the intron for integration into
the Peperomia mitochondrial genome, as a perfect 1:1 correspondence exists between the
intron's presence in a species and the presence of divergent nucleotide markers flanking the
intron insertion site.  Sequencing of coxI introns from additional Peperomia species revealed
that several mutations have occurred in the intron since the horizontal transfer, but sequence
alterations have not caused frameshifts or created stop codons in the intronic open reading
frame.  In addition, two coxI pseudogenes in Peperomia cubensis were discovered that lack a
large region of coxI exon 2 and contain a truncated version of the group I intron that likely
cannot be spliced out.

<>

<1>Adams, M.D. et al.
<2>The genome sequence of Drosophila melanogaster.
<3>Science
<4>287
<5>2185-2195
<6>2000
<7>The fly Drosophila melanogaster is one of the most intensively studied organisms in biology
and serves as a model system for the investigation of many developmental and cellular
processes common to higher eukaryotes, including humans. We have determined the nucleotide
sequence of nearly all of the approximately 120-megabase euchromatic portion of the Drosophila
genome using a whole-genome shotgun sequencing strategy supported by extensive clone-based
sequence and a high-quality bacterial artificial chromosome physical map. Efforts are under
way to close the remaining gaps; however, the sequence is of sufficient accuracy and
contiguity to be declared substantially complete and to support an initial analysis of genome
structure and preliminary gene annotation and interpretation. The genome encodes approximately
13,600 genes, somewhat fewer than the smaller Caenorhabditis elegans genome, but with
comparable functional diversity.

<>

<1>Adams, M.D., Chan, E.R., Molyneaux, N.D., Bonomo, R.A.
<2>Genomewide analysis of divergence of antibiotic resistance determinants in closely related isolates of Acinetobacter baumannii.
<3>Antimicrob. Agents Chemother.
<4>54
<5>3569-3577
<6>2010
<7>Multidrug resistance has emerged as a significant concern with infections
caused by Acinetobacter baumannii. Ample evidence supports the involvement
of mobile genetic elements in the transfer of antibiotic resistance genes,
but the extent of variability and the rate of genetic change associated
with the acquisition of antibiotic resistance have not been studied in
detail. Whole-genome sequence analysis of six closely related clinical
isolates of A. baumannii, including four from the same hospital, revealed
extensive divergence of the resistance genotype that correlated with
observed differences in antimicrobial susceptibility. Resistance genes
associated with insertion sequences, plasmids, and a chromosomal
resistance gene island all showed variability. The highly dynamic
resistance gene repertoire suggests rapid evolution of drug resistance.

<>

<1>Adams, M.D., Goglin, K., Molyneaux, N., Hujer, K.M., Lavender, H., Jamison, J.J., MacDonald, I.J., Martin, K.M., Russo, T., Campagnari, A.A., Hujer, A.M., Bonomo, R.A., Gill, S.R.
<2>Comparative genome sequence analysis of multidrug-resistant Acinetobacter baumannii.
<3>J. Bacteriol.
<4>190
<5>8053-8064
<6>2008
<7>The recent emergence of multidrug resistance (MDR) in Acinetobacter
baumannii has raised concern in health care settings worldwide. In order
to understand the repertoire of resistance determinants and their
organization and origins, we compared the genome sequences of three MDR
and three drug-susceptible A. baumannii isolates. The entire MDR phenotype
can be explained by the acquisition of discrete resistance determinants
distributed throughout the genome. A comparison of closely related MDR and
drug-susceptible isolates suggests that drug efflux may be a less
significant contributor to resistance to certain classes of antibiotics
than inactivation enzymes are. A resistance island with a variable
composition of resistance determinants interspersed with transposons,
integrons, and other mobile genetic elements is a significant but not
universal contributor to the MDR phenotype. Four hundred seventy-five
genes are shared among all six clinical isolates but absent from the
related environmental species Acinetobacter baylyi ADP1. These genes are
enriched for transcription factors and transporters and suggest
physiological features of A. baumannii that are related to adaptation for
growth in association with humans.

<>

<1>Adams, R.L.P.
<2>Eukaryotic DNA methyltransferases - structure and function.
<3>Bioessays
<4>17
<5>139-145
<6>1995
<7>Methylation of DNA plays an important role in the control of gene expression in higher
eukaryotes. This is largely achieved by the packaging of methylated DNA into chromatin
structures that are inaccessible to transcription factors and other proteins. Methylation
involves the addition of a methyl group to the 5-position of the cytosine base in DNA, a
reaction catalysed by a DNA (cytosine-5) methyltransferase. This reaction occurs in nuclear
replication foci where the chromatin structure is loosened for replication, thereby allowing
access to methyltransferases. Partly as a result of their recognizing the presence of a
methylcytosine on the parental strand following replication, these large enzymes are able to
maintain the distribution of methyl groups along the DNA of somatic cells and, thereby,
maintain tissue-specific patterns of gene expression.

<>

<1>Adams, R.L.P., Pradhan, S., Johnson, C.A., Lindsay, H., Shek, E.W.L., Jenkins, G.I., Urwin, N.A.R.
<2>Plant methyltransferases and their targets in the plant genome.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, Russo, V.E.A., Martienssen, R.A., Riggs, A.D., Cold Spring Harbor
<4>0
<5>95-108
<6>1996
<7>In addition to the methylation of cytosine in some CG dinucleotides, plant genomes contain
5-methylcytosine (a cytosine with a methyl group in place of the hydrogen at position 5) in
the trinucleotide sequence mCNG, where N is reportedly any of the four common DNA bases.
5-Methylcytosine has also been found in nonsymmetrical sequences in transgenes, but there is
no evidence in plants for the N-4 methylcytosine that is found in some prokaryotes.  We have
shown that CG and CNG methylation are carried out by different methyltransferases, and one
possible consequence of this is that the two reactions could be independently regulated.  This
would allow the methylation of CG and CNG sequences to have unrelated functions in the plant
cell, although to date there are no data to support this suggestion.

<>

<1>Adelskov, J., Patel, B.K.
<2>Draft Genome Sequence of Bacillus subtilis Strain D7XPN1, Isolated from Commercial Bioreactor-Degrading Food Waste.
<3>Genome Announcements
<4>2
<5>e00989-14
<6>2014
<7>The analysis of the 4.1-Mb draft genome sequence of a moderately thermophilic, heterotrophic,
and facultatively anaerobic bacterium, Bacillus subtilis strain
D7XPN1, identified genes for a range of enzymes with potential in the
biodegradation of food waste, a property consistent with the ecological habitat
of the isolate.

<>

<1>Adimpong, D.B., Sorensen, K.I., Nielsen, D.S., Thorsen, L., Rasmussen, T.B., Derkx, P.M., Jespersen, L.
<2>Draft Whole-Genome Sequence of Bacillus sonorensis Strain L12, a Source of Nonribosomal Lipopeptides.
<3>Genome Announcements
<4>1
<5>e0009713
<6>2013
<7>The Bacillus sonorensis L12 draft genome sequence is approximately 4,647,754 bp in size with a
G+C content of 45.2%. Over 86% of the genome contains
protein-encoding genes, including several gene clusters for de novo biosynthesis
of the nonribosomal lipopeptides iturin, bacitracin, and fengycin, which could
mean that the strain exhibits antifungal effects.

<>

<1>Adlakha, N., Ritturaj, K.H., Rajagopal, R., Yazdani, S.S.
<2>Draft Genome Sequence of the Paenibacillus sp. Strain ICGEB2008 (MTCC 5639) Isolated from the Gut of Helicoverpa armigera.
<3>Genome Announcements
<4>1
<5>e00026-12
<6>2013
<7>Paenibacillus sp. strain ICGEB2008 (MTCC 5639) is a Gram-positive cellulolytic bacterium,
isolated from the gut of Helicoverpa armigera. Here, we report the draft genome sequence of
Paenibacillus sp. ICGEB2008. The annotation of the ~5.7-Mb sequence indicated a cluster of
genes related to the glycosyl hydrolase family and the butanediol biosynthesis pathway.

<>

<1>Adler, S.P., Nathans, D.
<2>Studies of SV40 DNA: V. Conversion of circular to linear SV40 DNA by restriction endonuclease from Escherichia coli B.
<3>Biochim. Biophys. Acta
<4>299
<5>177-188
<6>1973
<7>Restriction endonuclease preparations from Escherichia coli strains B, K(P1)
and B(RTF2) have been found to cleave SV40 DNA.  Purified E. coli B restriction
endonuclease converted covalently closed circular SV40 DNA to predominantly
full length linear molecules with intact single strands; open circular
molecules were intermediates in the reaction.  After denaturation and
renaturation, the linear products formed circular duplexes with high
efficiency, indicating that individual SV40 DNA molecules had been cleaved by
the B enzyme at different sites.  Consistent with this conclusion was the
finding that digestion of the linear product with restriction endonuclease from
Hemophilus influenzae yielded DNA fragments indistinguishable
electrophoretically from those found after digestion of circular SV40 DNA with
the same enzyme.

<>

<1>Adriaenssens, E.M., Guerrero, L.D., Makhalanyane, T.P., Aislabie, J.M., Cowan, D.A.
<2>Draft Genome Sequence of the Aromatic Hydrocarbon-Degrading Bacterium Sphingobium sp. Strain Ant17, Isolated from Antarctic Soil.
<3>Genome Announcements
<4>2
<5>e00212-14
<6>2014
<7>Here, we present the draft genome sequence of Sphingobium sp. strain Ant17, an aromatic
hydrocarbon-degrading bacterium that was isolated from Antarctic oil-contaminated soil. An
analysis of this genome can lead to insights into the mechanisms of xenobiotic degradation
processes at low temperatures and potentially aid in bioremediation applications.

<>

<1>Advani, S., Mishra, P., Dubey, S., Thakur, S.
<2>Categoric prediction of metal ion mechanisms in the active sites of 17 select type II restriction endonucleases.
<3>Biochem. Biophys. Res. Commun.
<4>402
<5>177-179
<6>2010
<7>Recently determined crystal structures of type II restriction endonucleases have produced a
plethora of information on the basis for
target site sequence selectivity. The positioning and role of metal
ions in DNA recognition sites might reflect important properties of
protein-DNA interaction. Although acidic and basic groups in the active
sites can be identified, and in some cases divalent-metal binding sites
delineated, a convincing picture clarifying the way in which the
attacking hydroxide ion is generated, and the leaving group stabilized,
has not been elucidated for any of the enzymes. We have examined the
interatomic distances between metal ions and proposed key catalytic
residues in the binding sites of seventeen type II restriction
endonucleases whose crystal structures are documented in literature.
The summary and critical evaluation of structural assignments and
predictions made earlier have been useful to group these enzymes. All
the enzymes used for this study have been categorized on the basis of
the number of metal ions identified in their crystal structures. Among
17 experimentally characterized (not putative) type II REases, whose
apparently full-length sequences are available in REBASE, we predict 8
(47%) to follow the single metal ion mechanism, 5 to follow the two
metal ion mechanism, 2, the three metal ion mechanism, 1, the four
metal ion mechanism and 1 the six metal ion mechanism.

<>

<1>Advani, S., Roy, K.B.
<2>BanI restriction endonuclease binds in the major groove of DNA: An inhibition kinetic study using substrates with mismatch basepairs.
<3>Biochem. Biophys. Res. Commun.
<4>269
<5>35-40
<6>2000
<7>Structural information on BanI-DNA interaction was obtained from simple inhibition kinetic
assays using altered substrates. Self-complementary 13-mer oligodeoxynucleotides with or
without mismatch basepairs in the BanI recognition sequence (GGPyPuCC) were synthesized. UV
melting curves and CD spectra indicated double-stranded B-DNA structure for all the oligomers.
Among the seven oligomers with recognition sequences, GGTACC, GGTGCC, GGTATC, GGCACC, GGAGCC,
GGTAAC, and GGATCC, only the first two were cleaved with BanI. Kinetics of BanI cleavage of
the native substrate was inhibited competitively by all of the other oligomers except the one
with sequence GGCACC. From inhibition kinetic analysis in presence of a fixed concentration of
the inhibitor, apparent K(m) and K(I) were determined. The data were analyzed in the context
of alterations made in the hydrogen bonding potential in the major and minor groove of DNA
within the recognition sequence due to basepair mismatches. Such analyses led to the
conclusion that BanI, like BamHI, binds in the major groove and the central thymines make
important contact with the protein.

<>

<1>Advani, S., Roy, K.B.
<2>Properties and secondary structure analysis of BanI endonuclease: identification of putative active site.
<3>Biochem. Biophys. Res. Commun.
<4>279
<5>11-16
<6>2000
<7>Biochemical properties of Type II restriction enzyme BanI were characterized. Kinetic
parameters were evaluated and an enhancement of rate was observed when the recognition site
was located in a more central position in the substrate, suggesting that BanI locates its
recognition site by a sliding mechanism. As BanI has three cysteine residues in its primary
sequence, the effect of thiol inhibitors on BanI activity was also studied. Partial inhibition
was observed only at a very high concentration of the inhibitor indicating that cysteine
residues are not directly involved in catalysis. The gel electrophoretic mobility shift assay
demonstrated specific complex formation between BanI and the DNA substrate in the presence of
poly dI-dC and Mg(2+). A secondary structure analysis and comparison with EcoRI and BamHI
crystal structure revealed a putative active site similar to that seen in BamHI but different
in the order in which the catalytic domain (central beta-sheet) and recognition domain
(adjacent alpha-helix) were arranged in the protein.

<>

<1>Aeling, K.A., Steffen, N.R., Johnson, M., Hatfield, G.W., Lathrop, R.H., Senear, D.F.
<2>DNA deformation energy as an indirect recognition mechanism in protein-DNA interactions.
<3>IEEE/ACM Trans. Comput. Biol. Bioinform.
<4>4
<5>117-125
<6>2007
<7>Proteins that bind to specific locations in genomic DNA control many basic cellular functions.
Proteins detect their binding sites using
both direct and indirect recognition mechanisms. Deformation energy,
which models the energy required to bend DNA from its native shape to
its shape when bound to a protein, has been shown to be an indirect
recognition mechanism for one particular protein, Integration Host
Factor (IHF). This work extends the analysis of deformation to two
other DNA-binding proteins, CRP and SRF, and two enclonucleases, I-Crel
and I-Ppol. Known binding sites for all five proteins showed
statistically significant differences in mean deformation energy as
compared to random sequences. Binding sites for the three DNA-binding
proteins and one of the enclonucleases had mean deformation energies
lower than random sequences. Binding sites for I-Ppol had mean
deformation energy higher than random sequences. Classifiers that were
trained using the deformation energy at each base pair step showed good
cross-validated accuracy when classifying unseen sequences as binders
or nonbinders. These results support DNA deformation energy as an
indirect recognition mechanism across a wider range of DNA-binding
proteins. Deformation energy may also have a predictive capacity for
the underlying catalytic mechanism of DNA-binding enzymes.

<>

<1>Aertsen, A., Mebrhatu, M.T., Michiels, C.W.
<2>Activation of the Salmonella Typhimurium Mrr protein.
<3>Biochem. Biophys. Res. Commun.
<4>367
<5>435-439
<6>2008
<7>The Mrr protein of Escherichia coli K12 is a cryptic type IV restriction endonuclease with
specificity for methylated DNA. Recently
it was discovered that endogenous activation of E. coli Mrr could be
triggered by high pressure stress, resulting in the generation of
double strand breaks in the host chromosome and concomitant induction
of the SOS response. In this report we focused on Mrr activity of
Salmonella Typhimurium LT2, and although we surprisingly found no
evidence of high pressure induced activation, a large number of
constitutively activated Mrr mutants could be isolated when the mrr
gene was routinely cloned in an expression vector. Analysis of several
spontaneous mutants revealed different single mutations that rendered
the Mrr protein constitutively active. Moreover, a spontaneous S.
Typhimurium mutant could be isolated that displayed an increased basal
SOS induction because of a point mutation in the chromosomal mrr gene.
Based on these findings the physiological role of Mrr in the cell is
discussed.

<>

<1>Aertsen, A., Michiels, C.W.
<2>Mrr instigates the SOS response after high pressure stress in Escherichia coli.
<3>Mol. Microbiol.
<4>58
<5>1381-1391
<6>2005
<7>The bacterial SOS response is not only a vital reply to DNA damage but also constitutes an
essential mechanism for the generation of genetic
variability that in turn fuels adaptation and resistance development in
bacterial populations. Despite the extensive depiction of the SOS regulon
itself, its activation by stresses different from typical DNA damaging
treatments remains poorly characterized. Recently, we reported the RecA-
and LexA-dependent induction of the SOS response in Escherichia coli
MG1655 after exposure to high hydrostatic pressure (HP, approximately 100
MPa), a physical stress of which the cellular effects are not well known.
We now found this HP mediated SOS response to depend on RecB and not on
RecF, which is a strong indication for the involvement of double strand
breaks. As the pressures used in this work are thermodynamically unable to
break covalent bonds in DNA, we hypothesized the involvement of a cellular
function or pathway in the formation of this lesion. A specialized
screening allowed us to identify the cryptic type IV restriction
endonuclease Mrr as the final effector of this pathway. The HP SOS
response and its corresponding phenotypes could be entirely attributed to
the HP triggered activation of Mrr restriction activity. Several
spontaneously occurring alleles of mrr, incapable of triggering the
HP-induced SOS response, were isolated and characterized. These results
provide evidence for a specific pathway that transmits the perception of
HP stress to induction of the SOS response and support a role for Mrr in
bacterial stress physiology.

<>

<1>Agarkova, I.V., Dunigan, D.D., Van Etten, J.L.
<2>Virion-associated restriction endonucleases of Chloroviruses.
<3>J. Virol.
<4>80
<5>8114-8123
<6>2006
<7>Chloroviruses are large, double-stranded-DNA, plaque-forming viruses that infect certain
eukaryotic chlorella-like green algae. The
prototype of the genus is Paramecium bursaria chlorella virus 1
(PBCV-1). Chlorovirus genomes contain various amounts of methylated
nucleotides due to virus-encoded DNA methyltransferases (NITases);
about 25% of the MTases are associated with companion DNA site-specific
(restriction) endonucleases (REases). These enzymes constitute virally
encoded restriction-modification (R/M) systems. Although several of the
chlorovirus R/M systems are characterized, their biological functions
are unknown. The PBCV-1 proteome reveals that two virus-encoded REases,
but not their companion MTases, are virion associated, suggesting that
viral REases might help degrade the host DNA early in infection. To
test this hypothesis, host chromosomal DNA from PBCV-1-infected cells
was examined by pulsed-field gel electrophoresis. Initiation of host
chromosomal DNA degradation occurred within 5 min postinfection (p.i.).
The DNA degradation was insensitive to protein synthesis inhibitors or
UV inactivation of virus particles, consistent with the agent being a
small protein associated with the virion. Nuclease activities,
including those of the two predicted REases and an uncharacterized
general nuclease(s), were detected in disrupted PBCV-1 particles. The
general nuclease(s) degraded both host and viral DNAs in vitro,
although the viral DNA was not degraded in vivo, suggesting
differential intracellular trafficking of the virion-associated
nucleases. Infection with chloroviruses lacking an R/M system(s)
resulted in either delayed host chromosomal DNA degradation or no
detectable host chromatin changes. These immediate-early events
associated with chlorovirus infections may facilitate rapid switching
of the host transcriptional apparatus to viral transcription, which
begins within 5 to 10 min p.i.

<>

<1>Agarwal, L., Purohit, H.J.
<2>Genome Sequence of Rhizobium lupini HPC(L) Isolated from Saline Desert Soil, Kutch (Gujarat).
<3>Genome Announcements
<4>1
<5>e00071-12
<6>2013
<7>The Rhizobium lupini strain HPC(L) was isolated from saline desert soil. It grows on minimal
media supplemented with CaCO(3) as a carbon source. It can also grow under both oligotrophic
and heteroptrophic conditions. We report the annotated genome sequence of this strain in a
5.27-Mb scaffold.

<>

<1>Agarwal, P.K., Bhattacharya, S.K.
<2>Construction of a multi RE module: Exploitation of mechanochemistry of restriction endonucleases.
<3>Biotechnol. Bioeng.
<4>65
<5>233-239
<6>1999
<7>We describe the construction of a multi-immobilized restriction endonuclease module (Multi RE
module). We demonstrate that the applied mechanical stress enables modulation of enzyme
activity and modulation of recognition site selectivity (in oligonucleotides of approximately
200 bp) of immobilized restriction endonucleases. The central module which consists of
different strips of immobilized restriction endonucleases allows limited digestion of a large
DNA sample in a controlled manner as a function of applied mechanical stress on strips. The
stress-activity relationship and the effect of repeated cycles of stress and relaxation on the
immobilized strips are presented here.

<>

<1>Aggarwal, A.K.
<2>Structure and function of restriction endonucleases.
<3>Curr. Opin. Struct. Biol.
<4>5
<5>11-19
<6>1995
<7>Structures of two restriction endonucleases, BamHI and PvuII, were reported in the past year.
This doubles the number of restriction endonuclease structures now known from two to four, and
enables a comparative analysis of their structures and modes of DNA recognition. Despite a
lack of sequence homology between the enzymes, BamHI turns out to resemble EcoRI, and PvuII
turns out to resemble EcoRV. The active-site regions are structurally similar in all four
enzymes, but their mechanisms of cleavage may differ.

<>

<1>Aggarwal, A.K.
<2>Crystallization of DNA binding proteins with oligodeoxynucleotides.
<3>Methods
<4>1
<5>83-90
<6>1990
<7>In contrast to oligodeoxynucleotides, protein:DNA complexes crystallize from a broad range of
precipitants and conditions, much as proteins by themselves.  There are, however, a number of
factors that should be considered, at least in the early stages of cocrystallization attempts.
These include the length and construction of the oligodeoxynucleotide itself, a pH near or
below neutrality, a stoichiometric excess of DNA, and di- and polyvalent cations.  By far the
more important of these factors are the length of the DNA fragment and the nature of the
terminal nucleotides.  Unfortunately, experience suggests that, in general, the length and
construction of the DNA fragment for optimal crystal growth cannot be predictd in advance.  A
systematic search of cocrystallization conditions with different DNA fragments will normally
be required.  It is important to avoid sequences that provide possible subsites within the
fragment or at junctions between fragments related by translations.  Sample purity, especially
that of the DNA, can also have important effects.  Detailed protocols for the purification of
chemically synthesized fragments are suggested.  Finally, a set of conditions for initial
cocrystallization trails with a new DNA binding protein is suggested.

<>

<1>Aggarwal, A.K., Wah, D.A.
<2>Novel site-specific DNA endonucleases.
<3>Curr. Opin. Struct. Biol.
<4>8
<5>19-25
<6>1998
<7>Site-specific hydrolysis of DNA is common to many biological processes.  Three new structures,
FokI, I-CreI and PI-SceI, were reported in the past year, providing the first view of type IIs
endonucleases and homing endonucleases.  Together, they reveal an extraordinary set of new
mechanisms by which endonucleases target the hydrolysis of specific DNA sequences.

<>

<1>Agren, J., Finn, M., Bengtsson, B., Segerman, B.
<2>Microevolution during an Anthrax Outbreak Leading to Clonal Heterogeneity and Penicillin Resistance.
<3>PLoS ONE
<4>9
<5>E89112
<6>2014
<7>Anthrax is a bacterial disease primarily affecting grazing animals but it can
also cause severe disease in humans. We have used genomic epidemiology to study
microevolution of the bacterium in a confined outbreak in cattle which involved
emergence of an antibiotic-resistant phenotype. At the time of death, the animals
contained a heterogeneous population of Single Nucleotide Variants (SNVs), some
being clonal but most being subclonal. We found that independent isolates from
the same carcass had similar levels of SNV differences as isolates from different
animals. Furthermore the relative levels of subclonal populations were different
in different locations in the same carcass. The heterogeneity appeared to be
derived in part from heterogeneity in the infectious dose. The resistance
phenotype was linked to clonal mutations in an anti-sigma factor gene and in one
case was preceded by an acquisition of a hypermutator phenotype. In another
animal, small subclonal populations were observed with counteracting mutations
that had turned off the resistance genes. In summary, this study shows the
importance of accounting for both acquired and inherited heterogeneity when doing
high-resolution infection tracing and when estimating the risks associated with
penicillin treatment.

<>

<1>Agron, P.G., Sobecky, P., Andersen, G.L.
<2>Establishment of uncharacterized plasmids in Escherichia coli by in vitro transposition.
<3>FEMS Microbiol. Lett.
<4>217
<5>249-254
<6>2002
<7>We present a simple approach that permits any circular plasmid, such as uncharacterized
plasmids from diverse prokaryotes, to be established in
Escherichia coli, thereby facilitating subsequent structural and
functional studies. An in vitro transposition reaction is used to
introduce a well-characterized replicon and selectable marker into
purified plasmids, which are then used to transform E. coli. The
approach was demonstrated using a small 3.4-kb archaeal plasmid and a
large 60-kb uncharacterized plasmid from a Gram-negative bacterium.
Replicon function in E coli was tested for each plasmid, and direct
sequencing of the large plasmid revealed similarity to
restriction-modification systems.

<>

<1>Aguado-Urda, M., Lopez-Campos, G.H., Blanco, M.M., Fernandez-Garayzabal, J.F., Cutuli, M.T., Aspiroz, C., Lopez-Alonso, V., Gibello, A.
<2>Genome sequence of Lactococcus garvieae 21881, isolated in a case of human septicemia.
<3>J. Bacteriol.
<4>193
<5>4033-4034
<6>2011
<7>Lactococcus garvieae is a Gram-positive bacterium considered as an important opportunistic
emerging human pathogen and also a well recognized
fish pathogen. Here, we present the draft genome sequence of Lactococcus
garvieae strain 21881 (2,164,557 bp, with a G+C content of 37.9%), which
represents the first report of a genome sequence on Lactococcus garvieae.

<>

<1>Aguado-Urda, M., Lopez-Campos, G.H., Gibello, A., Cutuli, M.T., Lopez-Alonso, V., Fernandez-Garayzabal, J.F., Blanco, M.M.
<2>Genome Sequence of Lactococcus garvieae 8831, Isolated from Rainbow Trout Lactococcosis Outbreaks in Spain.
<3>J. Bacteriol.
<4>193
<5>4263-4264
<6>2011
<7>Lactococcus garvieae is the etiological agent of lactococcosis, one of the most important
disease threats to the sustainability of the rainbow trout
farming industry. Here, we present the draft genome sequence of
Lactococcus garvieae strain 8831, isolated from diseased rainbow trout,
which is composed of 2,087,276 bp with a G+C content of 38%.

<>

<1>Aguilar-Bultet, L., Calderon-Copete, S.P., Frey, J., Falquet, L.
<2>Draft Genome Sequence of the Virulent Avibacterium paragallinarum Serotype A Strain JF4211 and Identification of Two Toxins.
<3>Genome Announcements
<4>1
<5>e00592-13
<6>2013
<7>Avibacterium paragallinarum is an important pathogen of chicken livestock causing infectious
coryza. Here, we report the draft genome sequence of the virulent A.
paragallinarum serotype A strain JF4211 (2.8 Mbp and G+C content of 41%) and the
two toxin operons discovered from the annotation of the genome.

<>

<1>Aguirre-Arteta, A.M., Grunewald, I., Cardoso, M.C., Leonhardt, H.
<2>Expression of an alternative Dnmt1 isoform during muscle differentiation.
<3>Cell Growth Differ.
<4>11
<5>551-559
<6>2000
<7>The methylation pattern of genomic DNA undergoes dramatic changes during
mammalian development, with extensive de novo methylation occurring during
gametogenesis and after implantation. We identified an alternative Dnmt1
transcript in skeletal muscle by Northern blot analysis and cloned the
corresponding cDNA by rapid amplification of cDNA ends and reverse
transcription-PCR. Using an in vitro skeletal muscle differentiation
system, we show that this alternative Dnmt1 isoform is specifically
expressed in differentiated myotubes, whereas the ubiquitously expressed
isoform is down-regulated during myogenesis. Sequence analysis showed that
this skeletal Dnmt1 isoform is identical to the one present in testis,
which had been described as untranslatable. Here we present evidence that
this alternative Dnmt1 transcript present in testis and skeletal muscle is
translated despite the presence of several out-of-frame upstream ATGs and
gives rise to a shorter Dnmt1 isoform, which could play an active role in
the change of DNA methylation patterns during gametogenesis and
myogenesis.

<>

<1>Aherfi, S., Pagnier, I., Fournous, G., Raoult, D., La Scola, B., Colson, P.
<2>Complete genome sequence of Cannes 8 virus, a new member of the proposed family 'Marseilleviridae'.
<3>Virus Genes
<4>47
<5>550-555
<6>2013
<7>Marseillevirus is a giant virus that was isolated in 2007 by culturing water
collected from a cooling tower in Paris, France, on Acanthamoeba polyphaga. Since
then, five other marseilleviruses have been detected in environmental or human
samples. The genomes of two of the six marseilleviruses have been described in
detail. We describe herein the genome of Cannes 8 virus, a new member of the
proposed family "Marseilleviridae." Cannes 8 virus was isolated from water
collected from a cooling tower in Cannes in southeastern France. Its genome is a
circular double-stranded DNA molecule with 374,041 base pairs, larger than the
Marseillevirus and Lausannevirus genomes. This genome harbors 484 open reading
frames predicted to encode proteins with sizes ranging from 50 to 1,537 amino
acids, among which 380 (79 %) and 272 (56 %) are bona fide orthologs of
Marseillevirus and Lausannevirus proteins, respectively. In addition, 407 and 336
predicted proteins have significant hits against Marseillevirus and Lausannevirus
proteins, respectively, and 294 proteins are shared by all three
marseilleviruses. The Cannes 8 virus genome has a high level of collinearity (for
96 % of orthologs) with the Marseillevirus genome. About two-thirds of the Cannes
8 virus gene repertoire is composed of family ORFans. The description and
annotation of the genomes of new marseilleviruses that will undoubtedly be
recovered from environmental or clinical samples will be helpful to increase our
knowledge of the pan-genome of the family "Marseilleviridae."

<>

<1>Ahir, V.B., Roy, A., Jhala, M.K., Bhanderi, B.B., Mathakiya, R.A., Bhatt, V.D., Padiya, K.B., Jakhesara, S.J., Koringa, P.G., Joshi, C.G.
<2>Genome Sequence of Pasteurella multocida subsp. gallicida Anand1_poultry.
<3>J. Bacteriol.
<4>193
<5>5604
<6>2011
<7>We report the finished and annotated genome sequence of Pasteurella multocida gallicida strain
Anand1_poultry, which was isolated from the
liver of a diseased adult female chicken. The strain causes a disease
called 'fowl cholera,' which is a contagious disease in birds. We compared
it with the published genome sequence of Pasteurella multocida Pm70.

<>

<1>Ahlert, D., Stegemann, S., Kahlau, S., Ruf, S., Bock, R.
<2>Insensitivity of chloroplast gene expression to DNA methylation.
<3>Mol. Genet. Genomics
<4>282
<5>17-24
<6>2009
<7>Presence and possible functions of DNA methylation in plastid genomes of higher plants have
been highly controversial. While a number of
studies presented evidence for the occurrence of both cytosine and
adenine methylation in plastid genomes and proposed a role of cytosine
methylation in the transcriptional regulation of plastid genes, several
recent studies suggested that at least cytosine methylation may be
absent from higher plant plastid genomes. To test if either adenine or
cytosine methylation can play a regulatory role in plastid gene
expression, we have introduced cyanobacterial genes for adenine and
cytosine DNA methyltransferases (methylases) into the tobacco plastid
genome by chloroplast transformation. Using DNA cleavage with
methylation-sensitive and methylation-dependent restriction
endonucleases, we show that the plastid genomes in the transplastomic
plants are efficiently methylated. All transplastomic lines are
phenotypically indistinguishable from wild-type plants and, moreover,
show no alterations in plastid gene expression. Our data indicate that
the expression of plastid genes is not sensitive to DNA methylation
and, hence, suggest that DNA methylation is unlikely to be involved in
the transcriptional regulation of plastid gene expression.

<>

<1>Ahmad, F., Huang, X., Lan, H.X., Huma, T., Bao, Y.M., Huang, J., Zhang, H.S.
<2>Comprehensive gene expression analysis of the DNA (cytosine-5) methyltransferase family in rice (Oryza sativa L.).
<3>Gen. Mol. Res.
<4>13
<5>5159-5172
<6>2014
<7>Cytosine DNA methylation is a conserved epigenetic regulatory mechanism in both plants and
animals. DNA methyltransferases (DNA MTases) not only initiate (de novo) but also maintain the
process of DNA methylation. Here, we characterized the genome-wide expression profiles of 10
cytosine DNA MTase genes belonging to 4 subfamilies, MET1, CMT, DNMT2, and DRM, in rice.
Tissue-specific gene expression analysis showed that all family members varied widely in their
expression and specificities and might be involved in some basic metabolic pathways.
Similarly, the expression of all rice cytosine DNA MTase genes was not regulated by plant
hormones except OsDRM1a and OsDRM1b, which were downregulated by jasmonic acid. The
transcription level of 10 genes in rice shoots and roots was also measured under salt and
osmotic stress. Meanwhile, quantitative polymerase chain reaction data of the japonica and
indica rice cultivars revealed that there is large variation in the expression activities of
all genes. The results provide a foundation to further explore the roles of DNA MTases and the
epigenetic regulation of abiotic stress responses in rice.

<>

<1>Ahmad, I., Krishnamurthy, V., Rao, D.N.
<2>DNA recognition by the EcoP15I and EcoPI modification methyltransferases.
<3>Gene
<4>157
<5>143-147
<6>1995
<7>The DNA-binding properties of the EcoP15I DNA methyltransferase (M.EcoP15I; MTase) were
studied using electrophoretic mobility shift assays.  We show by molecular size-exclusion
chromatography and dimethyl suberimidate crosslinking that M.EcoP15I is a dimer in solution.
While M.EcoP15I binds approximately three-fold more tightly to its recognition sequence,
5'-CAGCAG-3', than to non-specific sequences in the presence of AdoMet or its analogs, the
discrimination between specific and non-specific sequences significantly increases in the
presence of ATP.  These results suggest for the first time a role for ATP in DNA recognition
by type-III restriction-modification enzymes.  Furthermore, we show that although c2 EcoPI
mutant MTases are defective in AdoMet binding, they are still able to bind DNA in a
sequence-specific manner.

<>

<1>Ahmad, I., Rao, D.N.
<2>Interaction of EcoP15I DNA methyltransferase with oligonucleotides containing the asymmetric sequence 5'-CAGCAG-3'.
<3>J. Mol. Biol.
<4>242
<5>378-388
<6>1994
<7>EcoP15I DNA methyltransferase (Mtase) recognizes the asymmetric sequence CAGCAG and catalyzes
the transfer of a methyl group from S-adenosyl-L-methionine to the second adenine residue. We
have investigated the DNA binding properties of EcoP15I DNA Mtase using gel mobility shift
assays. EcoP15I DNA Mtase binds approximately threefold more tightly to DNA containing its
recognition sequence, CAGCAG, than to non-specific sequences in the absence or presence of
cofactors. Interestingly, in the presence of ATP the discrimination between specific and
non-specific sequences increases significantly. These results suggest for the first time a
role for ATP in DNA recognition by type III restriction-modification enzymes. In addition, we
have shown that bromodeoxyuridine-containing oligonucleotides form complexes with EcoP15I DNA
Mtase that are cross-linked upon irradiation. More importantly, we have shown that the
crosslink site is at the site of DNA binding, since it can be suppressed by an excess of
unmodified oligonucleotide. EcoP15I DNA Mtase exhibited Michaelis-Menten kinetics with both
unmodified and bromodeoxyuridine-substituted DNA, with a higher specificity constant for the
latter. Furthermore, gel mobility shift assays showed that proteolyzed EcoP15I DNA Mtase
formed a specific complex with DNA, which had similar mobility as the native protein-DNA
complex. Taken together these results form the basis for a detailed structure-function
analysis of EcoP15I DNA Mtase.

<>

<1>Ahmad, I., Rao, D.N.
<2>Functional analysis of conserved motifs in EcoP15I DNA methyltransferase.
<3>J. Mol. Biol.
<4>259
<5>229-240
<6>1996
<7>EcoP15I DNA methyltransferase recognizes the sequence 5'-CAGCAG-3' and transfers a methyl
group to N-6 of the second adenine residue in the recognition sequence.  All N-6 adenine
methyltransferases contain two highly conserved sequences, FxGxG (motif 1), postulated to form
part of the S-adenosyl-L-methionine binding site and (D/N/S)PP(Y/F) (motif IV) involved in
catalysis.  We have altered the second glycine residue in motif I to arginine and serine, and
substituted tyrosine in motif IV with tryptophan in EcoP15I DNA methyltransferase, using
site-directed mutagenesis.  The mutant enzymes were overexpressed, purified and characterized
by biochemical methods.  The mutations in motif I completely abolished AdoMet binding but left
target DNA recognition unaltered.  Although the mutation in motif IV resulted in loss of
enzyme activity, we observed enhanced crosslinking of S-adenosyl-L-methionine and DNA.  This
implies that DNA and AdoMet binding sites are close to motif IV.  Taken together, these
results reinforce the importance of motif I in AdoMet binding and motif IV in catalysis.
Additionally, limited proteolysis and UV crosslinking experiments with EcoP15I DNA
methyltransferase imply that DNA binds in a cleft formed by two domains in the protein.
Methylation protection analysis provides evidence for the fact that EcoP15I DNA Mtase makes
contacts in the major groove of its substrate DNA.  Interestingly, hypermethylation of the
guanine residue next to the target adenine residue indicates that the protein probably flips
out the target adenine residue.

<>

<1>Ahmad, I., Rao, D.N.
<2>Chemistry and biology of DNA methyltransferases.
<3>Crit. Rev. Biochem. Mol. Biol.
<4>31
<5>361-380
<6>1996
<7>Recognition of a specific DNA sequence by a protein is probably the best example of
macromolecular interactions leading to various events.  It is a prerequisite to understanding
the basis of protein-DNA interactions to obtain a better insight into fundamental processes
such as transcription, replication, repair, and recombination.  DNA methyltransferases with
varying sequence specificities provide an excellent model system for understanding the
molecular mechanism of specific DNA recognition.  Sequence comparison of cloned genes, along
with mutational analyses and recent crystallographic studies, have clearly defined the
functions of various conserved motifs.  These enzymes access their target base in an elegant
manner by flipping it out of the DNA double helix.  The drastic protein-induced DNA
distortion, first reported for HhaI DNA methyltransferase, appears to be a common mechanism
employed by vairous proteins that need to act on bases.  A remarkable feature of the catalytic
mechanism of DNA (cytosine-5) methyltransferases is the ability of these enzymes to induce
deamination of the target cytosine in the absence of S-adenosyl-L-methionine or its analogs.
The enzyme-catalyzed deamination reaction is postulated to be the major cause of mutational
hotspots at CpG islands responsible for various human genetic disorders.  Methylation of
adenine residues in Escherichia coli is known to regulate various processes such as
transcription, replication, repair, recombination, transposition, and phage packaging.

<>

<1>Ahmad, I., Rao, D.N.
<2>Photolabeling of the EcoP15 DNA methyltransferase with S-adenosyl-L-methionine.
<3>Gene
<4>142
<5>67-71
<6>1994
<7>Radioactivity from S-adenosyl-L[methyl-3H]methionine ([methyl-3H]AdoMet) was bound to the
EcoP15 DNA methyltransferase (M.EcoP15) following short-wave ultraviolet (UV) irradiation. The
labeled protein was subjected to polyacrylamide-gel electrophoresis in the presence of sodium
dodecyl sulfate (SDS-PAGE), and detected by fluorography and autoradiography. Labeling was
found to be dependent on the concentration of AdoMet and time of UV irradiation. The
photolabeling by [methyl-3H]AdoMet was specific and blocked by S-adenosyl-L-homocysteine
(AdoHcy) and sinefungin which are known to function as competitive inhibitors. Limited
digestion of the M.EcoP15-AdoMet adduct by Staphylococcus aureus protease V8 generated three
peptides of approx. 50, 32 and 30 kDa. Interestingly, only the 30-kDa peptide fragment
contained radioactivity, as detected by SDS-PAGE, followed by fluorography and
autoradiography. Further, sequencing of a few amino acids at the N-terminus of these peptides
showed that the 30-kDa fragment was the N-terminal portion of M.EcoP15. These results suggest
that photolabeling is at the AdoMet-binding site and that the N-terminal half of M.EcoP15 may
be involved in substrate binding.

<>

<1>Ahmed, A., Earl, J., Retchless, A., Hillier, S.L., Rabe, L.K., Cherpes, T.L., Powell, E., Janto, B., Eutsey, R., Hiller, N.L., Boissy, R., Dahlgren, M.E., Hall, B.G., Costerton, J.W., Post, J.C., Hu, F.Z., Ehrlich, G.D.
<2>Comparative Genomic Analyses of 17 Clinical Isolates of Gardnerella vaginalis Provide Evidence of Multiple Genetically Isolated Clades Consistent with  Subspeciation into Genovars.
<3>J. Bacteriol.
<4>194
<5>3922-3937
<6>2012
<7>Gardnerella vaginalis is associated with a spectrum of clinical conditions, suggesting high
degrees of genetic heterogeneity among stains. Seventeen G.
vaginalis isolates were subjected to a battery of comparative genomic analyses to
determine their level of relatedness. For each measure, the degree of difference
among the G. vaginalis strains was the highest observed among 23 pathogenic
bacterial species for which at least eight genomes are available. Genome sizes
ranged from 1.491 to 1.716 Mb; GC contents ranged from 41.18% to 43.40%; and the
core genome, consisting of only 746 genes, makes up only 51.6% of each strain's
genome on average and accounts for only 27% of the species supragenome.
Neighbor-grouping analyses, using both distributed gene possession data and core
gene allelic data, each identified two major sets of strains, each of which is
composed of two groups. Each of the four groups has its own characteristic genome
size, GC ratio, and greatly expanded core gene content, making the genomic
diversity of each group within the range for other bacterial species. To test
whether these 4 groups corresponded to genetically isolated clades, we inferred
the phylogeny of each distributed gene that was present in at least two strains
and absent in at least two strains; this analysis identified frequent homologous
recombination within groups but not between groups or sets. G. vaginalis appears
to include four nonrecombining groups/clades of organisms with distinct gene
pools and genomic properties, which may confer distinct ecological properties.
Consequently, it may be appropriate to treat these four groups as separate
species.

<>

<1>Ahmed, I., Oshima, K., Suda, W., Kitamura, K., Iida, T., Ohmori, Y., Fujiwara, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequence of the Boron-Tolerant and Moderately Halotolerant Bacterium Gracilibacillus boraciitolerans JCM 21714T.
<3>Genome Announcements
<4>2
<5>e00097-14
<6>2014
<7>Gracilibacillus boraciitolerans JCM 21714(T) has been characterized as a highly boron-tolerant
and moderately halotolerant bacterium. Here, we report the draft
genome sequence of this strain. The genome sequence facilitates an understanding
of the biochemical functions of boron and provides a base to identify the gene(s)
involved in the boron tolerance mechanism of the strain.

<>

<1>Ahmed, I.H., Manning, G., Wassenaar, T.M., Cawthraw, S., Newell, D.G.
<2>Identification of genetic differences between two Campylobacter jejuni strains with different colonization potentials.
<3>Microbiology
<4>148
<5>1203-1212
<6>2002
<7>The consumption of poultry meat contaminated with Campylobacter jejuni is considered to be a
risk factor for human campylobacteriosis. The
development of targeted strategies to control campylobacters in
broilers would benefit from knowledge of those bacterial factors
important in colonization of the avian gut. During preliminary studies
it was noted that C. jejuni NCTC 11168 was a poorer colonizer of
chickens than strain 81116. This poor colonization could not be fully
restored by in vivo passage, suggesting that it was a genetically
endowed property of strain 11168. As the genome sequence is available
for this strain, the technique of subtractive hybridization was used to
identify gene fragments of strain 81116 not present in strain 11168.
After two screening cycles, 24 out of 42 clones were identified as
having DNA inserts specific for strain 81116. Six of these 24 clones
contained gene fragment inserts with similarities to
restriction-modification enzymes found in other bacteria. Two inserts
had similarity to arsenic-resistance genes, whereas four others had
similarities to cytochrome c oxidase III, dTDP-glucose 4,6-dehydratase,
gamma-glutamyl transpeptidase and an abortive phage-resistance protein.
At least some of these genes may be involved with colonization. A
further six inserts had weak similarities to hypothetical proteins or
to proteins with assigned functions from strain 11168. The remaining
six clones had gene-fragment inserts with no database matches.
Southern-blot analysis confirmed that strain-dependent variation
existed for each of these DNA inserts. These results indicate that
subtractive hybridization can successfully identify genes that are
absent from the only C. jejuni strain for which the genome sequence is
currently available.

<>

<1>Ahmed, N., Saini, V., Raghuvanshi, S., Khurana, J.P., Tyagi, A.K., Tyagi, A.K., Hasnain, S.E.
<2>Molecular analysis of a leprosy immunotherapeutic bacillus provides insights into Mycobacterium evolution.
<3>PLoS ONE
<4>2
<5>E968
<6>2007
<7>BACKGROUND: Evolutionary dynamics plays a central role in facilitating the
mechanisms of species divergence among pathogenic and saprophytic mycobacteria.
The ability of mycobacteria to colonize hosts, to proliferate and to cause
diseases has evolved due to its predisposition to various evolutionary forces
acting over a period of time. Mycobacterium indicus pranii (MIP), a taxonomically
unknown 'generalist' mycobacterium, acts as an immunotherapeutic against leprosy
and is approved for use as a vaccine against it. The large-scale field trials of
this MIP based leprosy vaccine coupled with its demonstrated immunomodulatory and
adjuvant property has led to human clinical evaluations of MIP in interventions
against HIV-AIDS, psoriasis and bladder cancer. MIP, commercially available as
'Immuvac', is currently the focus of advanced phase III clinical trials for its
antituberculosis efficacy. Thus a comprehensive analysis of MIP vis-a-vis
evolutionary path, underpinning its immanent immunomodulating properties is of
the highest desiderata. PRINCIPAL FINDINGS: Genome wide comparisons together with
molecular phylogenetic analyses by fluorescent amplified fragment length
polymorphism (FAFLP), enterobacterial repetitive intergenic consensus (ERIC)
based genotyping and candidate orthologues sequencing revealed that MIP has been
the predecessor of highly pathogenic Mycobacterium avium intracellulare complex
(MAIC) that did not resort to parasitic adaptation by reductional gene evolution
and therefore, preferred a free living life-style. Further analysis suggested a
shared aquatic phase of MAIC bacilli with the early pathogenic forms of
Mycobacterium, well before the latter diverged as 'specialists'.
CONCLUSIONS/SIGNIFICANCE: This evolutionary paradigm possibly affirms to marshall
our understanding about the acquisition and optimization of virulence in
mycobacteria and determinants of boundaries therein.

<>

<1>Ahuja, Y.R.
<2>Bacterial infection restriction endonucleases, genetic damage and cancer.
<3>Biol. Zentralbl.
<4>110
<5>179-187
<6>1991
<7>Restriction endonucleases are known to cut naked DNA at specific sites by
recognizing their palindromes.  Similarly, in vitro in human and mammalian
cells, restriction endonucleases have been shown to cause nonrandom chromosome
damage, in addition to inducing gene amplification and mutations.  Bacteria,
which are the source of restriction endonucleases, have also been shown to be
clastogenic in vivo in human and mammalian cells.  Furthermore, it has been
demonstrated that the bacteria cause nonrandom chromosome damage.  Normally,
bacteria are phagocytized and their contents degraded by the cellular enzymes.
However, it is possible that for some reason (mutation or physiological) the
restriction endonucleases may cause specific chromosome damage, gene mutation
or gene amplification, which in turn may involve oncogene activation, leading
to cancer.

<>

<1>Ahuja, Y.R., Obe, G.
<2>Are rogue cells an indicator of cancer risk due to the action of bacterial restriction endonucleases?
<3>Mutat. Res.
<4>310
<5>103-112
<6>1994
<7>Cytogenetic surveys in normal individuals have occasionally shown the occurrence of cells with
multiple chromosome-type aberrations in some of the subjects.  These cells, which are rare,
have been termed as rogue cells.  Rogue cells, which have been observed worldwide, have a
mysterious nature.  It has been suggested that they may give rise to cancer.  Various
mechanisms have been considered for the causation of the rogue-cell phenomenon in the past but
none of them appears to be fully justified.  In this paper we propose their occurrence due to
the action of bacterial restriction endonucleases.

<>

<1>Ai, H., Zhang, J., Yang, M., Yu, P., Li, S., Zhu, M., Dong, H., Wang, S., Wang, J.
<2>Draft Genome Sequence of an Anaerobic, Thermophilic Bacterium, Thermoanaerobacterium aotearoense SCUT27, Isolated from a Hot Spring in China.
<3>Genome Announcements
<4>2
<5>e00041-14
<6>2014
<7>Thermoanaerobacterium aotearoense SCUT27, isolated from a hot spring in China, is a strictly
anaerobic, thermophilic bacterium capable of degrading xylan and
converting both pentose and hexose to ethanol with high yields. Here, we report
the draft genome sequence of SCUT27, which reveals insights into the mechanisms
of carbon source coutilization and xylan degradation in this thermophilic
microorganism.

<>

<1>Ai, L., Chen, C., Zhou, F., Wang, L., Zhang, H., Chen, W., Guo, B.
<2>Complete genome sequence of probiotic Lactobacillus casei BD-II.
<3>J. Bacteriol.
<4>193
<5>3160-3161
<6>2011
<7>Lactobacillus casei BD-II, a patented probiotic strain (U.S. patent 7,270,994 B2), was
isolated from homemade koumiss in China and has been
implemented in the industrial production as starter cultures. Here we
report the complete genome sequence of BD-II which shows high similarity
with the well studied probiotic BL23.

<>

<1>Aiba, Y., Sumaoka, J., Komiyama, M.
<2>Artificial DNA cutters for DNA manipulation and genome engineering.
<3>Chem. Soc. Rev.
<4>40
<5>5657-5668
<6>2011
<7>This tutorial review provides recent developments in artificial cutters for site-selective
scission of DNA with the focus on chemistry-based DNA cutters. They are useful tools for
molecular biology and biotechnology, since their site-selectivity of scission is much higher
than that of naturally occurring restriction enzymes and also their scission site is freely
chosen. In order to prepare these cutters, a DNA-cutting molecule is combined with a
sequence-recognizing molecule in a covalent or non-covalent way. At targeted sites in
single-stranded and double-stranded DNAs, the scission occurs via either oxidative cleavage of
nucleotides or hydrolysis of phosphodiester linkages. Among many successful examples, an
artificial restriction DNA cutter, prepared from Ce(IV)/EDTA and pseudo-complementary peptide
nucleic acid, hydrolyzed double-stranded DNA at the target site. The scission site and
scission specificity are determined simply in terms of the Watson-Crick rule so that even the
whole genome of human beings was selectively cut at one predetermined site. Consistently,
homologous recombination in human cells was successfully promoted by this tool. For the
purpose of comparison, protein-based DNA cutters (e.g., zinc finger nucleases) are also
briefly described. The potential applications of these cutters and their future aspects are
discussed.

<>

<1>Aikawa, C., Furukawa, N., Watanabe, T., Minegishi, K., Furukawa, A., Eishi, Y., Oshima, K., Kurokawa, K., Hattori, M., Nakano, K., Maruyama, F., Nakagawa, I., Ooshima, T.
<2>Complete Genome Sequence of the Serotype k Streptococcus mutans Strain LJ23.
<3>J. Bacteriol.
<4>194
<5>2754-2755
<6>2012
<7>Streptococcus mutans is the major pathogen of dental caries and occasionally causes infective
endocarditis. Here we report the complete genome sequence of
serotype k S. mutans strain LJ23, which was recently isolated from the oral
cavity of a Japanese patient.

<>

<1>Aiken, C., Gumport, R.I.
<2>Restriction endonuclease RsrI from Rhodobacter sphaeroides, an isoschizomer of EcoRI:  purification and properties.
<3>Nucleic Acids Res.
<4>16
<5>7901-7916
<6>1988
<7>We have purified RsrI endonuclease (R.RsrI), an isoschizomer of EcoRI, from Rhodobacter
sphaeroides strain 630. The enzyme is homogeneous as judged by polyacrylamide gel
electrophoresis and size-exclusion high-performance liquid chromatography. RsrI endonuclease
is a dimer over the concentration range of 0.05 to 1.4 mg/ml. The reduced and denatured
molecular weight of the enzyme is 30,000 Da. R.RsrI, like R.EcoRI, catalyzes the cleavage of
duplex DNA and oligodeoxyribonucleotides between the first two residues of the sequence
GAATTC. R.RsrI exhibits a kM of 14 nM and a kcat of 6.5 min-1 when reacting with pBR322 DNA at
25C. R.RsrI differs from R.EcoRI in its N-terminal amino acid sequence, susceptibility to
inhibition by antibodies, sensitivity to N-ethylmaleimide, isoelectric point, state of
aggregation at high concentrations, temperature lability, and conditions for optimal reaction.
R.RsrI displays a reduction of specificity (star activity) under conditions that also relax
the specificity of R.EcoRI.

<>

<1>Aiken, C., Gumport, R.I.
<2>Interaction of RsrI endonuclease, an isoschizomer of EcoRI, with oligodeoxyribonucleotides containing base analogues.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>79
<6>1989
<7>We have initiated a systematic kinetic study of the RsrI endonuclease, an
isoschizomer of EcoRI, using oligodeoxyribonucleotides containing base
analogues as substrates.  Several of these substrates were provided by Dr. L.
McLaughlin, Boston College, who tested them with EcoRI.  All of the base
substitutions tested affect the interaction of both enzymes with DNA.  In
general, the effects with RsrI are more pronounced than with EcoRI, as measured
by the greater reduction in the specificity constant (kcat/Km) for a given
substitution.  For two substrates, the RsrI-catalyzed reaction was very slow;
less than one-tenth the rate of the corresponding EcoRI reaction.  The lower
tolerance of RsrI for base analogues in its recognition sequence may reflect a
difference in the mechanisms by which these two enzymes recognize the same DNA
sequence and cut it at the same site.  We are presently determining whether
RsrI kinks and unwinds DNA to the same extent that EcoRI does.

<>

<1>Aiken, C., Milarski-Brown, K., Gumport, R.I.
<2>RsrI and EcoRI are two different restriction endonucleases that recognize the same DNA sequence.
<3>Fed. Proc.
<4>45
<5>1914
<6>1986
<7>In order to study the recognition of specific DNA sequences by proteins, we
have isolated RsrI endonuclease from Rhodopseudomonas sphaeroides strain 630.
The preparation is approximately 80% pure, with one major band which comigrates
on SDS gels with EcoRI endonuclease (M=30,000+-2000).  RsrI cleaves the duplex
octanucleotide pGGAATTCC to form pGG.  Plasmid and phage DNA digests confirm
that RsrI and EcoRI cleave the same sequence.  However, the enzymes appear to
be different.  Antiserum to EcoRI required 1000-fold increased concentrations
to effectively inhibit RsrI activity.  In addition, amino acid sequence
analysis of the first 40 amino acids of RsrI revealed little homology with
EcoRI.  Furthermore, the R. sphaeroides genome shows little homology with the
EcoRI gene when tested by Southern blotting and hybridization with washing at
low stringency.  A corresponding RsrI methylase activity has been identified
and separated from the endonuclease by chromatography on phosphocellulose.
This activity catalyzes the transfer of methyl groups from AdoMet to DNA,
rendering the DNA resistant to cleavage by both RsrI and EcoRI endonucleases.
It will be of interest to compare the mechanisms by which these enzymes
recognize their common DNA sequence.

<>

<1>Aiken, C.R.
<2>Sequence-specific recognition of DNA by RsrI endonuclease of Rhodobacter sphaeroides, an isoschizomer of EcoRI.
<3>Diss. Abstr.
<4>52
<5>1399
<6>1991
<7>In order to study the interaction of RsrI endonuclease with its DNA recognition sequence and
to determine whether RsrI recognizes DNA using a mechanism similar to that of EcoRI, I have
purified milligram amounts of the enzyme to near homogeneity from its natural source,
Rhodobacter sphaeroides strain 630. The enzyme comigrates with EcoRI during SDS-PAGE, with a
reduced and denatured molecular weight of 30,000+/- 2000 Da. RsrI endonuclease is a dimer at
concentrations of 0.05 to 1.4 mg/ml. In contrast to EcoRI, no tetrameric form was observed.
The isoelectric point of the endonuclease was determined to be 7.0-different than the value of
6.3 reported for EcoRI. The temperature and salt-dependence of the catalytic activity of RsrI
also differ from those of EcoRI. The reaction exhibits Michaelis-Menten kinetics, and values
of 14 nM and 6.5 min-1 were determined for Km and kcat, respectively using plasmid pBR322 as a
substrate. RsrI endonuclease, unlike EcoRI, is inactivated by preincubating the enzyme with 10
mM N-ethylmaleimide. This suggests that RsrI but not EcoRI has an essential sulfhydryl. RsrI
endonuclease cleaves noncanonical sequences in reactions containing 12.5 mM Tris-HCl, 2.5 mM
MgCl2, and 26% ethylene glycol. RsrI binding bends DNA by approximately 50 degrees and unwinds
the DNA helix by 25 +/- 5 degree - values similar to those reported for EcoRI endonuclease.
RsrI binding protects 12 nucleotides from cleavage by hydroxyl radical and MPE-Fe(II)
footprinting reagents. I have tested the RsrI endonuclease with a series of
decadeoxyribonucleotides containing base analogues as substrates. The kinetics of RsrI
cleavage are affected by each substitution, and the effects are generally more pronounced than
with EcoRI, as shown by the greater reduction in the specificity constant kcat/Km for a given
substitution. For several substrates, cleavage by RsrI was very slow - less than one-tenth the
rate of the corresponding EcoRI reaction. This lower tolerance of the RsrI endonuclease for
functional group changes in its recognition site may reflect differences in the mechanisms by
which RsrI and EcoRI recognize and cleave DNA.

<>

<1>Aiken, C.R., Fisher, E.W., Gumport, R.I.
<2>The specific binding, bending, and unwinding of DNA by RsrI endonuclease, an isoschizomer of EcoRI endonuclease.
<3>J. Biol. Chem.
<4>266
<5>19063-19069
<6>1991
<7>To determine whether RsrI endonuclease recognizes and cleaves the sequence
GAATTC in duplex DNA similarly to its isoschizomer EcoRI we initiated a
functional comparison of the two enzymes.  Equilibrium binding experiments
showed that at 20C RsrI endonuclease binds to specific and nonspecific
sequences in DNA with affinities similar to those of EcoRI.  At 0C the affinity
of RsrI for its specific recognition sequence is reduced 7-fold whereas the
affinity for noncanonical sequences remains relatively unchanged.  Unlike
EcoRI, incubation of RsrI endonuclease with N-ethylmaleimide inactivates the
enzyme; however, preincubation with DNA prevents the inactivation.  The
N-ethylmaleimide-treated enzyme fails to bind DNA as assayed by gel mobility
shift assays.  Comparison of the deduced amino acid sequences of RsrI and EcoRI
endonucleases suggests that modification of Cys245 is responsible for the
inactivation.  Fe(II).EDTA and methidiumpropyl-EDTA.Fe(II) footprinting results
indicate that RsrI, like EcoRI, protects 12 base pairs from cleavage when bound
to its specific recognition sequence in the absence of Mg2+.  RsrI bends DNA by
approximately 50 degrees, as determined by measuring the relative
electrophoretic mobilities of specific RsrI-DNA complexes with the binding site
in the center or near the end of the DNA fragment.  This value is similar to
that reported for EcoRI.  RsrI also unwinds the DNA helix by 25 degrees +/- 5
degrees, a value close to that reported for EcoRI endonuclease.  Collectively,
these results indicate that the overall structural changes induced in the DNA
by the binding of RsrI and EcoRI endonucleases to DNA in the absence of Mg2+
are similar.  In the accompanying paper (Aiken, C.R., McLaughlin, L.W., and
Gumport, R.I. (1991) J. Biol. Chem. 266, 19070-19078) we present results of
studies of RsrI endonuclease using oligonucleotide substrates containing base
analogues which suggest differences in the ways the two enzymes cleave DNA.

<>

<1>Aiken, C.R., Gumport, R.I.
<2>Base analogs in study of restriction enzyme-DNA interactions.
<3>Methods Enzymol.
<4>208
<5>433-457
<6>1991
<7>Enzymologists have long sought to discern the topography of the active sites of enzymes by
examining substrate analogs for their ability to serve as reactants. Such investigations aim
to contribute to our understanding of the kinetic and chemical mechanisms as well as the
stereochemistry and stereoselectivity of a reaction.

<>

<1>Aiken, C.R., McLaughlin, L.W., Gumport, R.I.
<2>The highly homologous isoschizomers RsrI endonuclease and EcoRI endonuclease do not recognize their target sequence identically.
<3>J. Biol. Chem.
<4>266
<5>19070-19078
<6>1991
<7>Using a series of decadeoxyribonucleotides containing base analogues as
substrates we measured the steady-state kinetic parameters for the reaction
catalyzed by RsrI endonuclease and compared the results to those with its
isoschizomer EcoRI.  The kinetics of RsrI cleavage are affected by each
substitution, with the effects being generally more deleterious than with
EcoRI, as shown by the greater reduction in the specificity constant kcat/Km.
The magnitudes of the effects of several substitutions are consistent with the
formation of direct enzyme-nucleobase contacts at the indicated positions.
With substrates containing 2-amino-purine or 2,6-diaminopurine at the central
adenine or uracil at the outermost thymine in the recognition sequence,
cleavage by RsrI was very slow, less than one-tenth the rate of the
corresponding EcoRI-catalyzed reaction.  The lower tolerance of RsrI
endonuclease for functional group changes in its recognition site may reflect
differences in the mechanisms of DNA recognition by the two enzymes.  Although
RsrI and EcoRI endonucleases bind with similar affinities to specific and
nonspecific DNA sequences and appear to introduce similar structural
distortions in DNA upon binding, the use of substrate analogues reveals
significant differences at the level of catalysis in the mechanisms by which
these two endonucleases recognize the duplex sequence GAATTC.

<>

<1>Ainala, S.K., Ashok, S., Ko, Y., Park, S.
<2>Glycerol assimilation and production of 1,3-propanediol by Citrobacter amalonaticus Y19.
<3>Appl. Microbiol. Biotechnol.
<4>97
<5>5001-5011
<6>2013
<7>Citrobacter amalonaticus Y19 (Y19) was isolated because of its ability for carbon
monoxide-dependent hydrogen production (water-gas shift reaction). This paper
reports the assimilation of glycerol and the production of 1,3-propanediol
(1,3-PDO) by Y19. Genome sequencing revealed that Y19 contained the genes for the
utilization of glycerol and 1,2-propanediol (pdu operon) along with those for the
synthesis of coenzyme B(12) (cob operon). On the other hand, it did not possess
the genes for the fermentative metabolism of glycerol of Klebsiella pneumoniae,
which consists of both the oxidative (dhaD and dhaK) and reductive (dhaB and
dhaT) pathways. In shake-flask cultivation under aerobic conditions, Y19 could
grow well with glycerol as the sole carbon source and produced 1,3-PDO. The level
of 1,3-PDO production was improved when vitamin B(12) was added to the culture
medium under aerobic conditions. Under anaerobic conditions, cell growth and
1,3-PDO production on glycerol was also possible, but only when an exogenous
electron acceptor, such as nitrate or fumarate, was added. This is the first
report of the glycerol metabolism and 1,3-PDO production by C. amalonaticus Y19.

<>

<1>Ainala, S.K., Somasundar, A., Park, S.
<2>Complete Genome Sequence of Pseudomonas denitrificans ATCC 13867.
<3>Genome Announcements
<4>1
<5>e00257-13
<6>2013
<7>Pseudomonas denitrificans ATCC 13867, a Gram-negative facultative anaerobic bacterium, is
known to produce vitamin B12 under aerobic conditions. This paper
reports the annotated whole-genome sequence of the circular chromosome of this
organism.

<>

<1>Ainsworth, S., Mahony, J., van Sinderen, D.
<2>The Plasmid Complement of Lactococcus lactis UC509.9 Encodes Multiple Bacteriophage Resistance Systems.
<3>Appl. Environ. Microbiol.
<4>80
<5>4341-4349
<6>2014
<7>Lactococcus lactis subsp. cremoris strains are used globally for the production of fermented
dairy products, particularly hard cheeses. Believed to be of plant origin, L. lactis strains
that are used as starter cultures have undergone extensive adaptation to the dairy
environment, partially through the acquisition of extrachromosomal DNA in the form of plasmids
that specify technologically important phenotypic traits. Here, we present a detailed analysis
of the eight plasmids of L. lactis UC509.9, an Irish dairy starter strain. Key industrial
phenotypes were mapped, and genes that are typically associated with lactococcal plasmids were
identified. Four distinct, plasmid-borne bacteriophage resistance systems were identified,
including two abortive infection systems, AbiB and AbiD1, thereby supporting the observed
phage resistance of L. lactis UC509.9. AbiB escape mutants were generated for phage sk1, which
were found to carry mutations in orf6, which encodes the major capsid protein of this phage.

<>

<1>Ainsworth, S., Zomer, A., de Jager, V., Bottacini, F., van Hijum, S.A., Mahony, J., van Sinderen, D.
<2>Complete Genome of Lactococcus lactis subsp. cremoris UC509.9, Host for a Model Lactococcal P335 Bacteriophage.
<3>Genome Announcements
<4>1
<5>e00119-12
<6>2013
<7>Here, we report the complete genome of Lactococcus lactis subsp. cremoris UC509.9, an Irish
dairy starter. The circular chromosome of L. lactis UC509.9
represents the smallest among those of the sequenced lactococcal strains, while
its large complement of eight plasmids appears to be a reflection of its
adaptation to the dairy environment.

<>

<1>Aizawa, Y., Xiang, Q., Pyle, A.M.
<2>Kinetic dissection of the multistep process in L1.ltrB intron mobility.
<3>Nucleic Acids Res. Suppl.
<4>1
<5>249-250
<6>2001
<7>A kinetic characterization is presented for intron insertion into a target duplex DNA site
during L1.ltrB intron mobility. This reaction is catalyzed by a ribonucleoprotein particle
(RNP), consisting of a lariat form of group II intron RNA and the intron-encoded LtrA protein.
In the first stage of intron mobility, the RNA component of the enzyme by itself inserts
directly into the target ds DNA site, and thus, the RNP enzyme does not carry out turnover.
Using single-turnover kinetics, we established an in vitro kinetic assay system and
investigated mechanism of the intron RNA insertion process.

<>

<1>Ajdic, D., McShan, W.M., McLaughlin, R.E., Savic, G., Chang, J., Carson, M.B., Primeaux, C., Tian, R., Kenton, S., Jia, H., Lin, S., Qian, Y., Li, S., Zhu, H., Najar, F., Lai, H., White, J., Roe, B.A., Jerretti, J.J.
<2>Genome sequence of Streptococcus mutans UA159, a cariogenic dental pathogen.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>14434-14439
<6>2002
<7>Streptococcus mutans is the leading cause of dental caries (tooth decay) worldwide and is
considered to be the most cariogenic of all of the oral streptococci. The genome of S. mutans
UA159, a serotype c strain, has been completely sequenced and is composed of 2,030,936 base
pairs. It contains 1,963 ORFs, 63% of which have been assigned putative functions. The genome
analysis provides further insight into how S. mutans has adapted to surviving the oral
environment through resource acquisition, defense against host factors, and use of gene
products that maintain its niche against microbial competitors. S. mutans metabolizes a wide
variety of carbohydrates via nonoxidative pathways, and all of these pathways have been
identified, along with the associated transport systems whose genes account for almost 15% of
the genome. Virulence genes associated with extracellular adherent glucan production,
adhesins, acid tolerance, proteases, and putative hemolysins have been identified. Strain
UA159 is naturally competent and contains all of the genes essential for competence and quorum
sensing. Mobile genetic elements in the form of IS elements and transposons are prominent in
the genome and include a previously uncharacterized conjugative transposon and a composite
transposon containing genes for the synthesis of antibiotics of the gramicidin/bacitracin
family; however, no bacteriophage genomes are present.

<>

<1>Ajinomoto, K.K.
<2>Microorganism variants derived from Brevibacterium spp. having reduced restriction enzyme activity, permitting production of strains giving high L-glutamic acid yields.
<3>Japanese Patent Office
<4>JP 58107172 A
<5>
<6>1983
<7>Microorganism variants are derived from Brevibacterium and show reduced restriction enzyme
activity. Though some of the bacteria of Brevibacterium can produce L-glutamic acid
efficiently, it has been difficult to improve them by DNA-recombination due to their high
restriction enzyme activity. By mutating the bacteria of Brevibacterium by the usual methods
and selecting the variants which are more easily infected with phage, that is, tolerant to
phage, variants showing reduced restriction enzyme activity, can be obtained. An example of a
variant is Brevibacterium lactofermentum AJ11736 (FERM-P6249) and they can be also derived
from B. ammoniagenes ATCC6871, B. debaricatum ATCC14020, B. flavum ATCC 13826, etc. The
variant showing restriction enzyme activity as low as possible is desirable.  Practically the
variety, of which restriction enzyme activity is below 1/100 as that of parent strain, can be
used for DNA-recombination. B. lactofermentum is infected by F2-phage and F3-phage which
originated from the analogous strains of B. flavum, with 1 million-10 million times more
easily than parent strain.

<>

<1>Akaha, S.
<2>Detecting presence of target DNA, involves amplifying nucleic acid signal generated by oligonucleotide, using base sequences complementary  to and non-complementary to base sequence of target DNA and probe for  target DNA detection - DNA probe, restricti.
<3>Japanese Patent Office
<4>JP 2005110664
<5>
<6>2005
<7>NOVELTY - Detecting (M1) presence of target DNA, involves using a base sequence complementary
to the base sequence of target DNA, an arbitrarily selectable base sequence which is not
complementary to the base sequence of the target DNA and a probe for target DNA detection, and
amplifying a nucleic acid signal generated by an oligonucleotide containing the complementary
and non-complementary base sequence with respect to target DNA and the probe. DETAILED
DESCRIPTION - INDEPENDENT CLAIMS are also included for the following: (1) kit for carrying out
(M1); (2) detecting or determining an immunological anti-ligand based on (M1); and (3)
detecting or determining an antigen based on (M1). BIOTECHNOLOGY - Preferred Method: In (M1),
the probe for detecting the target DNA comprises a base sequence which is complementary to the
base sequence of the target DNA at the 3' terminal and a base sequence which is not
complementary to base sequence of target DNA at the 5' terminal. The probe comprises the base
sequence, which hybridizes with the restriction enzyme recognition site of target DNA and a
base sequence, which is modified to prevent cleavage by restriction enzyme. (M1) further
involves hybridizing the probe with the target DNA, extending the 3' terminal of the probe,
reacting restriction enzyme on the restriction enzyme recognition site of the target DNA which
was hybridized, and producing double-stranded DNA containing the complementary strand and the
target DNA, cleaving the target DNA from the double-stranded DNA using restriction enzymes
extending the 3' terminal of target DNA by using DNA polymerase, and repeating the above
steps to amplify a nucleic acid signal generated by the oligonucleotide. USE - (M1) is useful
for detecting the presence of target DNA. (M1) is useful for detecting or determining an
immunological anti-ligand and for detecting or determining an antigen (claimed). (M1) is
useful for screening trace amounts of DNA in clinical and industrial fields. (M1) is also
useful for detecting antibody or antigen in immunological assays and for detecting the
presence of microorganisms causing food poisoning, pathogenic microorganisms in swimming
pools, etc. ADVANTAGE - (M1) enables to detect target DNA without amplifying the target DNA
and thus the detection is not controlled by the base sequence of target DNA. A number of
different target DNAs can be amplified using single probe. (M1) is simple to carry out and is
economical. EXAMPLE - No relevant example is given. (24 pages)

<>

<1>Akasaka, T., Matsuura, K., Emi, N., Kobayashi, K.
<2>Conjugation of plasmid DNAs with lactose via diazocoupling enhances resistance to restriction enzymes and acquires binding affinity to galactose-specific lectin.
<3>Biochem. Biophys. Res. Commun.
<4>260
<5>323-328
<6>1999
<7>Oligosaccharide-plasmid DNA conjugates were synthesized simply and effectively via the
diazocoupling method.  Plasmids (pUC19, pTRI-beta-actin, and pEGFP-C1) were treated with an
N-beta-lactoside-substituted diazonium salt to yield diazocoupling products with degree of
substitutions of 2.5-3.1 mol% of overall nucleobases. The lactose-pUC19 conjugate was found to
resist restriction enzymes more strongly than the nonconjugated plasmid DNA and to acquire a
strong binding affinity to galactose-specific lectin RCA(120). The diazocoupling modification
of pTRI-beta-actin plasmid DNA little influenced in vitro transcription with T7 RNA
polymerase. When lactose-pEGFP-C1 conjugate was transfected to baby hamster kidney (BHK) cells
by means of cationic lipids, transduced gene was expressed in BHK cells similarly with the
nonconjugated pEGFP-C1. The modification of plasmid DNA with carbohydrate enhanced the
resistance to restriction enzymes and developed a strong binding affinity to
galactose-specific lectin.

<>

<1>Akatov, A.K., Zueva, V.S., Dmitrenko, O.A.
<2>A new approach to establishing the set of phages for typing methicillin-resistant Staphylococcus aureus.
<3>J. Chemother.
<4>3
<5>275-278
<6>1991
<7>A new approach to using experimental phages for typing methicillin-resistant S.
aureus (MRSA) non-sensitive to the phages of International Basic Set (IBS) is
described.  The collection includes phage 85, modified on a culture of MRSA,
and 5 phages induced from MRSA strains isolated in clinics of Moscow in
1975-76.  Firstly, the modified phage selects cultures according to the
specific character of its restriction-modification system, then the induced
phages differentiate the selected strains into 5 groups (1,2,3,4,5) based on
the specificity of the prophages they contain.  Group 1 strains can further be
differentiated into 5 subgroups (A,B,C,D,E) by additional phages.  Forty-one
MRSA strains isolated in 1987-90 in various hospitals of Moscow showing no
sensitivity to IBS phages, were lysed by the modified phage, 15 of them
belonging to Group 2 and isolated in the traumatological hospital, 26 belonging
to Group 1 and were circulating in the burn center.  Twenty-three strains of
Group 1 appertain to subgroup 1B and were isolated over a 4-year period from
the burned surface of patients and from the throat of a medical staff carrier.

<>

<1>Akcelik, M.
<2>The conjugal plasmid pLL10236 encodes lactose fermentation ability, restriction/modification activity, bacteriocin production and immunity in Lactococcus lactis subsp. Lactis LL102.
<3>Food Microbiol.
<4>16
<5>487-494
<6>1999
<7>A 36 kb plasmid, designed as pLL 10236, was determined in Lactococcus lactis subsp. Lactis
LL102.  This plasmid mediates lactose fermentation ability (Lac+), bacteriocin production
(Bac+) and immunity (Imm+), and restriction/modification (R+M+) activity against bacteriophage
O1120 in strain LL102.  The conjugal ability of pLL 10236 and expression of pLL 10236 encoding
traits were determined by using a Lac-, Bac-, Imm- and (R-M-) recipient strain L. lactis
subsp. Lactis P81-1.

<>

<1>Akcelik, M., Sanlibaba, P., Tukel, C.
<2>Phage resistance in Lactococcus lactis subsp lactis strains isolated from traditional fermented milk products in Turkey.
<3>Int. J. Food Sci. Technol.
<4>35
<5>473-481
<6>2000
<7>Lactococcus lactis subsp. lactis strains isolated from traditional fermented milk products in
Turkey were used to determine their phage
resistance against three different lactic phages. The following modes
of action were examined: phage adsorption inhibition in five strains,
abortive infection (heat sensitive phage resistance) in three strains,
restriction/modification in four strains and blocking of phage DNA
injection in one strain. The genetic nature of the phage resistance
systems in these strains was determined by comparison of phage
proliferation parameters, e.g. adsorption (%), EOP, burst size, latent
period and production of major capsid protein, between wild-type
strains and their plasmid-cured derivatives.

<>

<1>Akhtar, M., Perehinec, T.M., Nazli, A., Hill, P.J., Rees, C.E.D.
<2>Partial DNA methylation in Listeria monocytogenes creates variable populations of cells with altered virulence.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>411
<6>2001
<7>We have recently identified variable patterns of DNA methylation sites in the major virulence
region of the food borne pathogen Listeria
monocytogenes. The degree of partial methylation is specific to the
particular isolate, but high level methylation was only found in
serotype 4b strains, which are associated with outbreaks of
listeriosis. The methylation is visualised as reproducible band
patterns when DNA from a culture of cells is subjected to restriction
digest and southern hybridisation using a prfA-hlyA gene probe.
Controls were carried out to ensure that this partial restriction was
due to specific methylation and not due to simple enzyme inhibition.
Use of isoschizomers with differing methylation sensitivities allowed
us to identify this as cytosine-specific methylation. The gene
responsible for this methylation was cloned and found to have homology
to other cytosine methyl-transferases. To create these reproducible
patterns of partial DNA methylation, the methylation event must be
controlled. To investigate this, single Listeria monocytogenes cells,
which can only be methylated at one of the susceptible sites, were used
to inoculate tubes of broth and allowed to grow to stationary phase.
When the DNA was recovered from each of these cultures, all samples
exhibited exactly the same mixed pattern of DNA methylation indicating
that these patterns are not inherited, but rather methylation at this
locus must occur in a controlled manner. Either activity of the
methylase is directly modulated or architecture of the chromosomal DNA
must control access of the methylase to susceptible sites. In invasion
assays using Caco-2 cell lines, a correlation could be found between
the degree of DNA methylation associated with an isolate and the
invasiveness of that strain. This suggests that the methylation may
have an influence on gene expression and that in these cultures with
highly variable methylation patterns, some cells may be pre-primed to
cause disease.

<>

<1>Aklujkar, M., Young, N.D., Holmes, D., Chavan, M., Risso, C., Kiss, H.E., Han, C.S., Land, M.L., Lovley, D.R.
<2>The genome of Geobacter bemidjiensis, exemplar for the subsurface clade of Geobacter species that predominate in Fe(III)-reducing subsurface environments.
<3>BMC Genomics
<4>11
<5>490
<6>2010
<7>BACKGROUND: Geobacter species in a phylogenetic cluster known as
subsurface clade 1 are often the predominant microorganisms in subsurface
environments in which Fe(III) reduction is the primary electron-accepting
process. Geobacter bemidjiensis, a member of this clade, was isolated from
hydrocarbon-contaminated subsurface sediments in Bemidji, Minnesota, and
is closely related to Geobacter species found to be abundant at other
subsurface sites. This study examines whether there are significant
differences in the metabolism and physiology of G. bemidjiensis compared
to non-subsurface Geobacter species. RESULTS: Annotation of the genome
sequence of G. bemidjiensis indicates several differences in metabolism
compared to previously sequenced non-subsurface Geobacteraceae, which will
be useful for in silico metabolic modeling of subsurface bioremediation
processes involving Geobacter species. Pathways can now be predicted for
the use of various carbon sources such as propionate by G. bemidjiensis.
Additional metabolic capabilities such as carbon dioxide fixation and
growth on glucose were predicted from the genome annotation. The presence
of different dicarboxylic acid transporters and two oxaloacetate
decarboxylases in G. bemidjiensis may explain its ability to grow by
disproportionation of fumarate. Although benzoate is the only aromatic
compound that G. bemidjiensis is known or predicted to utilize as an
electron donor and carbon source, the genome suggests that this species
may be able to detoxify other aromatic pollutants without degrading them.
Furthermore, G. bemidjiensis is auxotrophic for 4-aminobenzoate, which
makes it the first Geobacter species identified as having a vitamin
requirement. Several features of the genome indicated that G. bemidjiensis
has enhanced abilities to respire, detoxify and avoid oxygen. CONCLUSION:
Overall, the genome sequence of G. bemidjiensis offers surprising insights
into the metabolism and physiology of Geobacteraceae in subsurface
environments, compared to non-subsurface Geobacter species, such as the
ability to disproportionate fumarate, more efficient oxidation of
propionate, enhanced responses to oxygen stress, and dependence on the
environment for a vitamin requirement. Therefore, an understanding of the
activity of Geobacter species in the subsurface is more likely to benefit
from studies of subsurface isolates such as G. bemidjiensis than from the
non-subsurface model species studied so far.

<>

<1>Akman, L., Rio, R.V.M., Beard, C.B., Aksoy, S.
<2>Genome size determination and coding capacity of Sodalis glossinidius, an enteric symbiont of Tsetse flies, as revealed by hybridization to Escherichia coli gene arrays.
<3>J. Bacteriol.
<4>183
<5>4517-4525
<6>2001
<7>Recent molecular characterization of various microbial genomes has revealed differences in
genome size and coding capacity between obligate symbionts and intracellular pathogens versus
free-living organisms. Multiple symbiotic microorganisms have evolved with tsetse fly, the
vector of African trypanosomes, over long evolutionary times. Although these symbionts are
indispensable for tsetse fecundity, the biochemical and molecular basis of their functional
significance is unknown. Here, we report on the genomic aspects of the secondary symbiont
Sodalis glossinidius. The genome size of Sodalis is approximately 2 Mb. Its DNA is subject to
extensive methylation and based on some of its conserved gene sequences has an A+T content of
only 45%, compared to the typically AT-rich genomes of endosymbionts. Sodalis also harbors an
extrachromosomal plasmid about 134 kb in size. We used a novel approach to gain insight into
Sodalis genomic contents, i.e., hybridizing its DNA to macroarrays developed for Escherichia
coli, a closely related enteric bacterium. In this analysis we detected 1,800 orthologous
genes, corresponding to about 85% of the Sodalis genome. The Sodalis genome has apparently
retained its genes for DNA replication, transcription, translation, transport, and the
biosynthesis of amino acids, nucleic acids, vitamins, and cofactors. However, many genes
involved in energy metabolism and carbon compound assimilation are apparently missing, which
may indicate an adaptation to the energy sources available in the only nutrient of  the tsetse
host, blood. We present gene arrays as a rapid tool for comparative genomics in the absence of
whole genome sequence to advance our understanding of closely related bacteria.

<>

<1>Akman, L., Yamashita, A., Watanabe, H., Oshima, K., Shiba, T., Hattori, M., Aksoy, S.
<2>Genome sequence of the endocellular obligate symbiont of tsetse flies, Wigglesworthia glossinidia.
<3>Nat. Genet.
<4>32
<5>402-407
<6>2002
<7>Many insects that rely on a single food source throughout their developmental cycle harbor
beneficial microbes that provide nutrients
absent from their restricted diet. Tsetse flies, the vectors of African
trypanosomes, feed exclusively on blood and rely on one such intracellular
microbe for nutritional provisioning and fecundity. As a result of
co-evolution with hosts over millions of years, these mutualists have lost
the ability to survive outside the sheltered environment of their host
insect cells. We present the complete annotated genome of Wigglesworthia
glossinidia brevipalpis, which is composed of one chromosome of 697,724
base pairs (bp) and one small plasmid, called pWig1, of 5,200 bp. Genes
involved in the biosynthesis of vitamin metabolites, apparently essential
for host nutrition and fecundity, have been retained. Unexpectedly, this
obligate's genome bears hallmarks of both parasitic and free-living
microbes, and the gene encoding the important regulatory protein DnaA is
absent.

<>

<1>Akopyants, N.S., Clifton, S.W., Kersulyte, D., Crabtree, J.E., Youree, B.E., Reece, C.A., Bukanov, N.O., Drazek, E.S., Roe, B.A., Berg, D.E.
<2>Analyses of the cag pathogenicity island of Helicobacter pylori.
<3>Mol. Microbiol.
<4>28
<5>37-53
<6>1998
<7>Most strains of Helicobacter pylori from patients with peptic ulcer disease or intestinal-type
gastric cancer carry cagA, a gene that encodes
an immunodominant protein of unknown function, whereas many of the strains
from asymptomatically infected persons lack this gene. Recent studies
showed that the cagA gene lies near the right end of a approximately 37kb
DNA segment (a pathogenicity island, or PAI) that is unique to cagA+
strains and that the cag PAI was split in half by a transposable element
insertion in the reference strain NCTC11638. In complementary experiments
reported here, we also found the same cag PAI, and sequenced a 39 kb
cosmid clone containing the left 'cagII' half of this PAI. Encoded in
cagII were four proteins each with homology to four components of
multiprotein complexes of Bordetella pertussis ('Ptl'), Agrobacterium
tumefaciens ('Vir'), and conjugative plasmids ('Tra') that help deliver
pertussis toxin and T (tumour inducing) and plasmid DNA, respectively, to
target eukaryotic or prokaryotic cells, and also homologues of eukaryotic
proteins that are involved in cytoskeletal structure. To the left of cagII
in this cosmid were genes for homologues of HsIU (heat-shock protein) and
Era (essential GTPase); to the right of cagII were homologues of genes for
a type I restriction endonuclease and ion transport functions. Deletion of
the cag PAI had no effect on synthesis of the vacuolating cytotoxin, but
this deletion and several cag insertion mutations blocked induction of
synthesis of proinflammatory cytokine IL-8 in gastric epithelial cells.
Comparisons among H. pylori strains indicated that cag PAI gene content
and arrangement are rather well conserved. We also identified two genome
rearrangements with end-points in the cag PAI. One, in reference strain
NCTC11638, involved IS605, a recently described transposable element (as
also found by others). Another rearrangement, in 3 of 10 strains tested
(including type strain NCTC11637), separated the normally adjacent cagA
and picA genes and did not involve IS605. Our results are discussed in
terms of how cag-encoded proteins might help trigger the damaging
inflammatory responses in the gastric epithelium and possible
contributions of DNA rearrangements to genome evolution.

<>

<1>Akopyants, N.S., Fradkov, A., Diatchenko, L., Hill, J.E., Siebert, P.D., Lukyanov, S.A., Sverdlov, E.D., Berg, D.E.
<2>PCR-based subtractive hybridization and differences in gene content among strains of Helicobacter pylori.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>13108-13113
<6>1998
<7>Genes that are characteristic of only certain strains of a bacterial species can be of great
biologic interest.  Here we describe a PCR-based subtractive hybridization method for
efficiently detecting such DNAs and apply it to the gastric pathogen Helicobacter pylori.
Eighteen DNAs specific to a monkey-colonizing strain (J166) were obtained by subtractive
hybridization against an unrelated strain whose genome has been fully sequenced (26695).
Seven J166-specific clones had no DNA sequence match to the 26695 genome, and 11 other clones
were mixed, with adjacent patches that did and did not match any sequences in 26695.  At the
protein level, seven clones had homology to putative DNA restriction-modification enzymes, and
two had homology to putative metabolic enzymes.  Nine others had no database match with
proteins of assigned function.  PCR tests of 13 unrelated H. pylori strains by using primers
specific for 12 subtracted clones and complementary Southern bot hybridizations indicated that
these DNAs are highly polymorphic in the H. pylori population, with each strain yielding a
different pattern of gene-specific PCR amplification.  The search for polymorphic DNAs, as
described here, should help identify previously unknown virulence genes in pathogens and
provide new insights into microbial genetic diversity and evolution.

<>

<1>Akulenko, N.V., Ivashina, T.V., Shaloiko, L.A., Shlyapnikov, M.G., Ksenzenko, V.N.
<2>New site-specific endonucleases F-TflI, F-TflII, and F-TflIV encoded by bacteriophage T5.
<3>Mol. Biol. (Mosk)
<4>38
<5>530-537
<6>2004
<7>Site-specific endonucleases F-TflI, F-TflII, and F-TflIV have been revealed, which belong to
the H-N-H family and are encoded by ORFs
located in the tRNA gene region of bacteriophage T5. It has been shown
that endonuclease F-TflIV introduces a double-strand break in a 17-bp
pseudopalindromic DNA sequence to yield 1-nt 3'-protruding ends. Unlike
F-TflIV, F-TflI, and F-TflII introduce single-strand breaks in
asymmetrical, highly degenerate sequences, each cleaving only one
(template or coding) strand. Amino acid sequence analysis has revealed
a high homology of the enzymes in the region of the H-N-H motif and in
the putative C-terminal catalytic domain. The N-terminal region of
F-TflIV proved to be homologous to the HTH domain of LuxR-related
transcriptional regulators, which is responsible for DNA recognition
and binding. The N-terminal regions of F-TflI and F-TflII contain a
composite motif NUMOD4, which is characteristic of a putative
recognition domain of some H-N-H endonucleases. A two-domain structure,
with the N-terminal recognition and C-terminal catalytic domains, and
evolutionary origin via recombination of the catalytic and recognition
domain-coding regions are proposed for F-TflI, F-TflII, and F-TflIV.

<>

<1>Akulinin, G.E., Getko, G.A., Repin, V.E., Degtyarev, S.K.
<2>New restriction endonuclease from the soil strain Bacillus megaterium 12.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>14
<5>105-108
<6>1988
<7>The soil strain Bacillus megaterium 12 carrying a restriction-modification
system has been discovered.  A Type 2 restriction endonuclease has been
partially purified.  Bme12I has been shown to be an isoschizomer of Sau3AI.

<>

<1>Akulinin, G.E., Repin, V.E.
<2>Bacillus megaterium strain - is used as producer of restriction endonuclease Bme12I.
<3>Soviet Patent Office
<4>SU 1486516 A
<5>
<6>1989
<7>Restriction endonuclease Bme12I, which is an isoschizomer of the restrictase Sau3AI, is
produced by Bacillus megaterium strain B-3677(12). The strain is extracted from the soil and
separated by clonal analysis, and the biomass is grown with aeration in a medium containing a
hydrolysate of sprats and yeast extract, until the growth slows down. The cells are then
disrupted by ultrasonication and the ferment separated by chromatography on phosphocellulose
and heparin sepharose. The product splits the sequence 5'-GATC-3' in double-stranded DNA.

<>

<1>Al-Awadhi, S., Welch, S.G., Smith, K.E., Williams, R.A.D.
<2>BstB7SI (R/CCGGY), a thermostable isoschizomer of Cfr10I, from a strain of Bacillus stearothermophilus isolated from oil-contaminated soil in Kuwait.
<3>FEMS Microbiol. Lett.
<4>160
<5>205-208
<6>1998
<7>Isolates of thermophilic bacteria from desert soil in Kuwait, heavily
contaminated with crude
oil, have been screened for the presence of restriction endonuclease activity. One of the
isolates (B7S),
identified as Bacillus stearothermophilus, showed a high level of restriction endonuclease
activity when a cell-
free extract was incubated with lambda bacteriophage DNA at 65oC.  A type II restriction
endonuclease
(BstB7SI) has been partially purified from this isolate.  BstB7SI recognizes the six-base
sequence RCCGGY
(R=A or G; Y=T or C) and hydrolyses the phosphodiester bond in both strands of the
DNA substrate
between the first and second bases of the recognition sequence 5'-R/CCGGY-3' producing
four-base 5'
overhangs.  BstB7SI is therefore an isoschizomer of the mesophilic prototype restriction
endonuclease
Cfr10I.  BstB7SI has a pH optimum of 9.7, requires 10 mM MgCl2 and 75 mM NaCl for
maximum activity,
and retains full enzyme activity when incubated for 5 min at temperatures up to 70oC.

<>

<1>Al-Rashdi, A.S.A., Jadhav, B.L., Deshpande, T., Deshpande, U.
<2>Whole-Genome Sequencing and Annotation of a Clinical Isolate of Mycobacterium tuberculosis from Mumbai, India.
<3>Genome Announcements
<4>2
<5>e00154-14
<6>2014
<7>We report here the annotated genome sequence of a clinical isolate of Mycobacterium
tuberculosis from Mumbai, India.

<>

<1>Al-Saari, N., Meirelles, P.M., Mino, S., Suda, W., Oshima, K., Hattori, M., Ohkuma, M., Thompson, F.L., Gomez-Gil, B., Sawabe, T., Sawabe, T.
<2>Draft Genome Sequences of Two Vibrionaceae Species, Vibrio ponticus C121 and Photobacterium aphoticum C119, Isolated as Coral Reef Microbiota.
<3>Genome Announcements
<4>2
<5>e01095-14
<6>2014
<7>Here, the draft genome sequences of two Vibrionaceae, Vibrio ponticus C121 and Photobacterium
aphoticum C119, which were isolated from the coral reef vicinity
in Okinawa, Japan, are reported. The genome provides further insight into the
genomic plasticity, biocomplexity, and ecophysiology, including pathogenicity and
evolution, of these genera.

<>

<1>Alam, M., Roy, C., Pyne, P., Agarwal, A., George, A., Ghosh, W.
<2>Whole-Genome Shotgun Sequence of the Sulfur-Oxidizing Chemoautotroph Pseudaminobacter salicylatoxidans KCT001.
<3>J. Bacteriol.
<4>194
<5>4743-4744
<6>2012
<7>The facultatively sulfur-oxidizing chemolithoautotrophic alphaproteobacterium Pseudaminobacter
salicylatoxidans KCT001 (MTCC 7265) belongs to the family
Phyllobacteriaceae of the order Rhizobiales. Analysis of its genome offers
valuable insight into the adaptive specializations and evolution of free-living
soil bacteria that are phylogenetically closely related to symbiotic and invasive
rhizobacteria.

<>

<1>Alam, N., Sittman, D.B.
<2>Characterization of the cytotoxic effect of a chimeric restriction enzyme, H1 degrees-FokI.
<3>Gene Ther. Mol. Biol.
<4>10A
<5>147-159
<6>2006
<7>Our primary goal was to create an efficient cytotoxic agent. To do this, we created a gene
that expresses a chimeric hybrid of the linker
histone, H1(0) and the nuclease domain of the type IIs restriction
enzyme, FokI. The linkage of the FokI nuclease domain to a high
affinity but low DNA-sequence-specificity binding protein is unique. It
is highly cytotoxic. We demonstrate, by transiently transfecting 3T3
mouse fibroblasts, that 63% of the cells taking up the chimeric gene
are killed. The chimeric protein is localized to the nucleus. An
extract of the protein produced in E. coli degrades DNA, indicating
that it is nucleolytically active. The ultimate mechanism through which
the chimeric protein produces cell death is likely through the
induction of apoptosis.

<>

<1>Alatossava, T., Klaenhammer, T.R.
<2>Molecular characterization of three small isometric-headed bacteriophages which vary in their sensitivity to the lactococcal phage resistance plasmid pTR2030.
<3>Appl. Environ. Microbiol.
<4>57
<5>1346-1353
<6>1991
<7>Lactococcus lactis LMA12-4 is a pTR2030 transconjugant that has been used as an
industrial starter culture because of its resistance to phages predominant in
cheese plants.  Plasmid pTR2030 interferes with susceptible phages in this host
strain via two mechanisms, restriction and modification (R/M) and abortive
infection (Hsp).  After prolonged use of LMA12-4 transconjugants in the
industry, two different bacteriophages, designated nck202.Phi 48 (Phi 48) and
nck202.Phi 50 (Phi 50), were isolated which could produce plaques on LMA12-4
containing pTR2030.  In this study, these two phages were characterized and
compared with a third phage, nck202.Phi 31 (Phi 31), which is susceptible to
both the R/M and Hsp activities encoded by pTR2030.  Phage Phi-48 was not
susceptible to inhibition by Hsp, whereas Phi 50 was unaffected by either the
R/M or Hsp mechanisms.  All three were small isometric-headed phages, but small
differences were noted between the phages in the structural details of the tail
base plate, susceptibility to chloroform treatment, and requirements for
calcium infectivity.  The phage genomes were all between 29.9 and 31.9 kb in
length.  Phages Phi 31 and Phi 48 harbored cohesive ends, whereas the phage Phi
50 genome was circularly permuted, terminally redundant, and carried a putative
packaging initiation site.  DNA-DNA hybridization experiments conducted between
the phages revealed a common region in Phi 48 and Phi 50 that may correlate
with the resistance of the two phages to the Hsp-abortive infection induced by
pTR2030.  Phage Phi 50 also harbored DNA sequences that shared homology to
pTR2030 in the region where R/M activities have been localized on the plasmid.
Molecular characterization of the three phages localized regions within the
genomes of the pTR2030-resistant phages that may be responsible for
circumventing plasmid-encoded Hsp and R/M defense mechanisms in lactococci.

<>

<1>Albalat, R.
<2>Evolution of DNA-methylation machinery: DNA methyltransferases and methyl-DNA binding proteins in the amphioxus Branchiostoma floridae.
<3>Dev. Genes Evol.
<4>218
<5>691-701
<6>2008
<7>DNA methylation is an epigenetic mark associated with gene regulation and cell memory,
silencing of transposable elements, genomic
imprinting, and repression of spurious transcription of duplicated
sequences. These roles have varied widely during animal evolution and
current functions depend on the specific methylation pattern of the
species under consideration. The patterns of methylation are
established, maintained, and translated into appropriate functional
states by the DNA-methylation machinery, which includes three groups of
methyltransferase enzymes, Dnmt1, Dnmt2 and Dnmt3, and five methyl-DNA
binding proteins, Mbd1, Mbd2, Mbd3, Mbd4, and MeCP2. In this study, I
have identified the members of the Dnmt and the Mbd gene families in
the cephalochordate amphioxus (Branchiostoma floridae), the most basal
extant chordate and one of the closest sister groups of vertebrates.
Database searches, phylogenetic studies and protein domain analyses
revealed the presence of the three major groups of Dnmt enzymes in the
cephalochordate genome, whereas only two Mbd members, Mbd2/3 and Mbd4,
were found. Analysis of the amphioxus methylation machinery suggested
that the complexity and the structural organization of cephalochordate
methyltransferases do not differ substantially from those of current
vertebrate enzymes, while new Mbd proteins arose in vertebrates, which
perhaps minimized certain collateral effects associated with the major
genomic changes that occurred during the invertebrate-vertebrate
transition.

<>

<1>Albersmeier, A., Bomholt, C., Glaub, A., Ruckert, C., Soriano, F., Fernandez-Natal, I., Tauch, A.
<2>Draft Genome Sequence of the Multidrug-Resistant Clinical Isolate Dermabacter hominis 1368.
<3>Genome Announcements
<4>2
<5>e00728-14
<6>2014
<7>Dermabacter hominis is a common colonizer of the healthy human skin and is rarely detected as
an opportunistic human pathogen. The genome sequence of the
multidrug-resistant D. hominis strain 1368, isolated from blood cultures of a
pyelonephritis patient, provides insights into the repertoire of antibiotic
resistance genes.

<>

<1>Albertsen, H.M., Le Paslier, D., Abderrahim, H., Dausset, J., Cann, H., Cohen, D.
<2>Improved control of partial DNA restriction enzyme digest in agarose using limiting concentrations of Mg++.
<3>Nucleic Acids Res.
<4>17
<5>808
<6>1989
<7>Partial digests of DNA for cloning experiments are usually performed using
either the quantity of restriction enzyme or incubation time, or both, as
controlling factors.  However, we have found that more precise and reproducible
partial digests are obtained, when a limiting concentration of the required
cofactor Mg++ is used.  This method is especially useful for partial digests of
DNA contained in agarose.  In agarose, diffusion time of the much smaller Mg++
ion is shorter than that of a restriction enzyme.  The result is a more
homogenous digestion of the DNA throughout the agarose block.  Since the
concentration of Mg++ in this procedure is usually only 10% or less of that
normally used, the digestion is easily interrupted by excess EDTA.  We have
successfully applied this partial digestion technique with EcoRI to obtain
large human DNA fragments that have been used to construct artificial yeast
chromosomes (YAC) of several hundred kb in length.

<>

<1>Albertsen, M., Hugenholtz, P., Skarshewski, A., Nielsen, K.L., Tyson, G.W., Nielsen, P.H.
<2>Genome sequences of rare, uncultured bacteria obtained by differential coverage binning of multiple metagenomes.
<3>Nat. Biotechnol.
<4>31
<5>533-538
<6>2013
<7>Reference genomes are required to understand the diverse roles of microorganisms in ecology,
evolution, human and animal health, but most species remain uncultured. Here we present a
sequence composition-independent approach to recover high-quality microbial genomes from
deeply sequenced metagenomes. Multiple metagenomes of the same community, which differ in
relative population abundances, were used to assemble 31 bacterial genomes, including rare
(<1% relative abundance) species, from an activated sludge bioreactor. Twelve genomes were
assembled into complete or near-complete chromosomes. Four belong to the candidate bacterial
phylum TM7 and represent the most complete genomes for this phylum to date (relative
abundances, 0.06-1.58%). Reanalysis of published metagenomes reveals that differential
coverage binning facilitates recovery of more complete and higher fidelity genome bins than
other currently used methods, which are primarily based on sequence composition. This approach
will be an important addition to the standard metagenome toolbox and greatly improve access to
genomes of uncultured microorganisms.

<>

<1>Albu, R.F., Jurkowski, T.P., Jeltsch, A.
<2>The Caulobacter crescentus DNA-(adenine-N6)-methyltransferase CcrM methylates DNA in a distributive manner.
<3>Nucleic Acids Res.
<4>40
<5>1708-1716
<6>2012
<7>
<>

<1>Albu, R.F., Zacharias, M., Jurkowski, T.P., Jeltsch, A.
<2>DNA Interaction of the CcrM DNA Methyltransferase: A Mutational and Modeling Study.
<3>Chembiochem
<4>13
<5>1304-1311
<6>2012
<7>Caulobacter crescentus CcrM is a DNA-(adenine N6)-methyltransferase that methylates adenine in
the sequence GANTC with high specificity. To
investigate its mechanism of DNA recognition, we used the crystal
structure of a related methyltransferase (M1.MboII, which modifies
GAAGA) as a starting point, and docked into it a DNA substrate to
identify the protein regions that approach the DNA. After alignment of
CcrM and M1.MboII, we identified four candidate regions in CcrM to
contain residues involved in DNA recognition. We mutated 20 amino acid
residues within these regions, purified the CcrM variants, and
determined their DNA-binding and catalytic activity on a cognate GANTC
substrate and on nine near-cognate substrates, each of which contained
a single base-pair substitution in the recognition sequence.
Altogether, we identified four residues in two of the regions,
mutations of which resulted in a strong (>100-fold) reduction of
methylation activity. Our data show that DNA recognition by CcrM is a
cooperative process, because disruption of critical contacts led to
loss of catalytic activity but not to a relaxation in specificity. In
addition, we identified a change in the readout of the fifth base pair
in the GANTC sequence with two other CcrM variants that showed smaller
reductions in overall activity. Based on this and the sequence
alignment of CcrM with other DNA methyltransferases of same or related
recognition sequence, we propose roles for these two regions in DNA
recognition by CcrM.

<>

<1>Alcaraz, L.D. et al.
<2>The genome of Bacillus coahuilensis reveals adaptations essential for survival in the relic of an ancient marine environment.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>5803-5808
<6>2008
<7>The Cuatro Cienegas Basin (CCB) in the central part of the Chihuahan desert (Coahuila, Mexico)
hosts a wide diversity of microorganisms
contained within springs thought to be geomorphological relics of an
ancient sea. A major question remaining to be answered is whether bacteria
from CCB are ancient marine bacteria that adapted to an oligotrophic
system poor in NaCl, rich in sulfates, and with extremely low phosphorus
levels (<0.3 muM). Here, we report the complete genome sequence of
Bacillus coahuilensis, a sporulating bacterium isolated from the water
column of a desiccation lagoon in CCB. At 3.35 Megabases this is the
smallest genome sequenced to date of a Bacillus species and provides
insights into the origin, evolution, and adaptation of B. coahuilensis to
the CCB environment. We propose that the size and complexity of the B.
coahuilensis genome reflects the adaptation of an ancient marine bacterium
to a novel environment, providing support to a "marine isolation origin
hypothesis" that is consistent with the geology of CCB. This genomic
adaptation includes the acquisition through horizontal gene transfer of
genes involved in phosphorous utilization efficiency and adaptation to
high-light environments. The B. coahuilensis genome sequence also revealed
important ecological features of the bacterial community in CCB and offers
opportunities for a unique glimpse of a microbe-dominated world last seen
in the Precambrian.

<>

<1>Alcaraz, L.D., Moreno-Hagelsieb, G., Eguiarte, L.E., Souza, V., Herrera-Estrella, L., Olmedo-Alvarez, G.
<2>Understanding the evolutionary relationships and major traits of Bacillus through comparative genomics.
<3>BMC Genomics
<4>11
<5>332
<6>2010
<7>BACKGROUND: The presence of Bacillus in very diverse environments reflects
the versatile metabolic capabilities of a widely distributed genus.
Traditional phylogenetic analysis based on limited gene sampling is not
adequate for resolving the genus evolutionary relationships. By
distinguishing between core and pan-genome, we determined the evolutionary
and functional relationships of known Bacillus. RESULTS: Our analysis is
based upon twenty complete and draft Bacillus genomes, including a newly
sequenced Bacillus isolate from an aquatic environment that we report for
the first time here. Using a core genome, we were able to determine the
phylogeny of known Bacilli, including aquatic strains whose position in
the phylogenetic tree could not be unambiguously determined in the past.
Using the pan-genome from the sequenced Bacillus, we identified functional
differences, such as carbohydrate utilization and genes involved in signal
transduction, which distinguished the taxonomic groups. We also assessed
the genetic architecture of the defining traits of Bacillus, such as
sporulation and competence, and showed that less than one third of the B.
subtilis genes are conserved across other Bacilli. Most variation was
shown to occur in genes that are needed to respond to environmental cues,
suggesting that Bacilli have genetically specialized to allow for the
occupation of diverse habitats and niches. CONCLUSIONS: The aquatic
Bacilli are defined here for the first time as a group through the
phylogenetic analysis of 814 genes that comprise the core genome. Our data
distinguished between genomic components, especially core vs. pan-genome
to provide insight into phylogeny and function that would otherwise be
difficult to achieve. A phylogeny may mask the diversity of functions,
which we tried to uncover in our approach. The diversity of sporulation
and competence genes across the Bacilli was unexpected based on previous
studies of the B. subtilis model alone. The challenge of uncovering the
novelties and variations among genes of the non-subtilis groups still
remains. This task will be best accomplished by directing efforts toward
understanding phylogenetic groups with similar ecological niches.

<>

<1>Alegado, R.A., Ferriera, S., Nusbaum, C., Young, S.K., Zeng, Q., Imamovic, A., Fairclough, S.R., King, N.
<2>Complete Genome Sequence of Algoriphagus sp. PR1, Bacterial Prey of a Colony-Forming Choanoflagellate.
<3>J. Bacteriol.
<4>193
<5>1485-1486
<6>2011
<7>Bacteria are the primary food source of choanoflagellates, the closest known relatives of
animals. Studying signaling interactions between the
Gram-negative Bacteroidetes bacterium Algoriphagus sp. PR1 and its
predator, the choanoflagellate Salpingoeca rosetta, provides a promising
avenue for testing hypotheses regarding the involvement of bacteria in
animal evolution. Here we announce the complete genome sequence of
Algoriphagus sp. PR1 and initial findings from its annotation.

<>

<1>Alegre, M.T., Rodriguez, M.C., Mesas, J.M.
<2>Characterization of pRS5: A theta-type plasmid found in a strain of Pediococcus pentosaceus isolated from wine that can be used to generate cloning vectors for lactic acid bacteria.
<3>Plasmid
<4>61
<5>130-134
<6>2009
<7>The nucleotide sequence of pRS5 (10153 bp) is reported. Through sequence analysis, 9 open
reading frames (ORFs) were identified and the
following features observed: a region likely involved in replication
whose structural features indicate that pRS5 belongs to the pUCL287
group of theta-type replicons, and hypothetical proteins putatively
involved in plasmid copy number control, restriction-modification
system, toxin-antitoxin system and a putative integrase. Shuttle
vectors for Escherichia coli and lactic acid bacteria (LAB) as well as
a small cloning vector for direct use in LAB were constructed using the
replication region of pRS5. The ability of such vectors to accept and
express other genes was assessed. All pRS5-derivatives were maintained
at a high rate over 200 generations without selective pressure.

<>

<1>Aleksandrzak-Piekarczyk, T., Koryszewska-Baginska, A., Bardowski, J.
<2>Genome Sequence of the Probiotic Strain Lactobacillus rhamnosus (Formerly Lactobacillus casei) LOCK900.
<3>Genome Announcements
<4>1
<5>e00640-13
<6>2013
<7>Lactobacillus rhamnosus LOCK900 fulfills the criteria required for probiotic strains. In this
study, we report a whole-genome sequence of this isolate and
compare it with other L. rhamnosus complete genome sequences already published.

<>

<1>Alencar, V.C., Barbosa, D., Santos, D.S., Oliveira, A.C., de Oliveira, R.C., Nunes, L.R.
<2>Genomic Sequencing of Two Coffee-Infecting Strains of Xylella fastidiosa Isolated from Brazil.
<3>Genome Announcements
<4>2
<5>e01190-13
<6>2014
<7>Here, we describe the draft genome sequences of two Xylella fastidiosa strains: Xf6c and Xf32,
which have been obtained from infected coffee plants in Brazil,
and are associated with the disease known as coffee leaf scorch (CLS).

<>

<1>Aleshkin, G.I., Skavronskaya, A.G.
<2>R.M.EcoRI coding plasmids derived from recombinant R factor.
<3>Genetika
<4>14
<5>1466-1469
<6>1978
<7>Bacteriophages P1vir and Mu-1 have been used for transductional shortening of a recombinant R
factor coding for R.M.EcoRI isolated by Yoshimori et al.  P1 shortening made possible the
isolation of transmissive isogenic plasmids coding for R.M.EcoRI and differing in antibiotic
resistance, as well as isolation of plasmids differing only in their R.M.EcoRI genes.  Mu-1
mediated shortening favored the isolation of transmissive R plasmids having lost the
resistance to chloramphenicol but having all other markers of the recombinant R factor intact.
The data are interpreted in support of Yoshimori et al.'s supposition concerning the
existence of R.M.EcoRI coding recombinant R factor of Escherichia coli.

<>

<1>Aleshkin, G.I., Strikhanov, S.N., Skavronskaya, A.G.
<2>Plasmid recombination stimulated by restriction endonuclease EcoRI in vivo: formation of recombinant plasmids in recA+-cells of E. coli.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>4
<5>15-21
<6>1985
<7>The possible participation of restriction endonuclease EcoRI in recombination of compatible
nonhomologous plasmids in E. coli cells has been studied.  To study the process, plasmids RP4
and R245 have been transferred by conjugation into the recipient cells of E. coli harboring
one of isogenic plasmids, pSA14 and pSA25, different for the genes coding restriction
endonuclease EcoRI.  The genetic analysis of transconjugant phenotypes, coded by the plasmids,
has permitted to register the recombinant plasmids after compatibility of parent plasmids in
E. coli cells.  Recombination of plasmid RP4 with the plasmid pSA14, carrying EcoRI genes, has
been registered in E. coli cells, producing the restriction endonuclease, while plasmid
recombination has not been found in the cells harboring plasmid pSA25, isogenic for all genes,
except for EcoRI genes, with plasmid pSA14.  Restriction endonuclease EcoRI is concluded to
stimulate site specific recombination of nonhomologous compatible plasmids in vivo.
EcoRI-mediated recombination of plasmid R245 with plasmid pSA14 is discussed.

<>

<1>Alexeyev, M.F., Gunkovskaya, N.V., Romanovskaya, V.A., Malashenko, Y.R.
<2>Methylococcus whittenburyi - producer of MwhI, new isoschizomer of HpaI.
<3>Mikrobiol. Zh.
<4>54
<5>70-73
<6>1992
<7>Type II specific restriction endonucleases (restrictases) draw so much interest not only
because gene engineering is based on using them but also because they are convenient objects
to solve the problem of nucleic acid-protein recognition. The list of these enzymes is being
enlarged, though some of them are more preferable for practical use. HpaI (from Haemophilus
parainfluenzae) is one such enzyme. A restriction endonuclease has been detected in
Methylococcus whittenburyi recognizing the same nucleotide sequence as HpaI. The new
isoschizomer was named MwhI according to the existing nomenclature. As M. whittenburyi (as
compared to H. parainfluenze) grows on simple mineral medium, is not pathogenic, comprises a
single enzyme, it might be promising for enzyme production.

<>

<1>Alexiev, A., Coil, D.A., Badger, J.H., Enticknap, J., Ward, N., Robb, F.T., Eisen, J.A.
<2>Complete Genome Sequence of Coprothermobacter proteolyticus DSM 5265.
<3>Genome Announcements
<4>2
<5>e00470-14
<6>2014
<7>Here we present the complete 1,424,912-bp genome sequence of Coprothermobacter proteolyticus
DSM 5265, isolated from a thermophilic digester fermenting tannery
wastes and cattle manure.

<>

<1>Algire, M.A., Montague, M.G., Vashee, S., Lartigue, C., Merryman, C.
<2>A type III restriction-modification system in Mycoplasma mycoides subsp. capri.
<3>Open Biology
<4>2
<5>120115
<6>2012
<7>the sequenced genome of Mycoplasma mycoides subsp. capri revealed the presence of a Type III
restriction-modification system (MmyCI).  The methyltransferase (modification) subunit of
MmyCI (M.MmyCI) was shown to recognize the sequence 5'-TGAG-3' and methylate the adenine.
The coding region of the methyltransferase gene contains 12 consecutive AG dinucleotide
repeats that result in a translational termination at a TAA codon immediately beyond the
repeat region.  This strain does not have MmyCI activity.  A clone was found with 10 AG
repeats such that the gene is in frame, and this strain has MmyCI activity, suggesting that
the expression of the MmyCI methyltransferase may be phase variable.

<>

<1>Aliyu, H., De Maayer, P., Rees, J., Tuffin, M., Cowan, D.A.
<2>Draft Genome Sequence of the Antarctic Polyextremophile Nesterenkonia sp. Strain  AN1.
<3>Genome Announcements
<4>2
<5>e00197-14
<6>2014
<7>Nesterenkonia sp. strain AN1 was isolated from Antarctic soil and is a polyextremophile, being
tolerant of low temperatures, high salt concentrations,
and high alkalinity. Here we report the draft genome sequence of this strain.

<>

<1>Allan, B.W.
<2>The dynamic origin of sequence discrimination by the EcoRI DNA methyltransferase.
<3>Diss. Abstr.
<4>59
<5>3407B
<6>1999
<7>All living cells use DNA modifying enzymes to maintain or expand the information encoded
within the genome.  These enzymes catalyze the chemical transformation of sterically occluded
target DNA residues within a huge excess of nontarget DNA.  Enzyme-DNA complexes reveal
striking distortions of DNA structure such as DNA bending and base flipping; yet, the
mechanism whereby DNA distortion modulates sequence specificity is only now being elucidated.
Intriguing mechanistic questions remain unresolved.  Do DNA modifying enzymes capture
transient structural intermediates or actively induce the conformational changes required for
DNA binding and catalysis?  Determination of the temporal correlation between the individual
steps in the reaction pathway provides a means to discern the origin of sequence
discrimination.  For the EcoRI DNA methyltransferase nearly absolute synchrony between DNA
binding and base flipping transitions are revealed during both complex assembly and
disassembly.  Because the rate of product formation under presteady-state conditions is
determined not by the methylation reaction, but rather by the pre-catalytic isomerization of
the complex, the base flipping transition is the selection gate whereby the commitment to
catalysis is manifested.  Our characterization of a methyltransferase mutant which unlike the
wild type methyltransferase displays no detectable DNA bending, shows that slowing the rate of
an obligate pre-catalytic isomerization can result in enhanced discrimination for an
induced-fit enzyme.  The recent extension of the experimental methodologies developed in this
work to other base flipping enzymes is providing fundamental information concerning the
integration of DNA distortion and enzyme specificity for other crucial enzymes that modify or
repair the DNA.  Finally, the high signal to noise, solution-based nature, and millisecond
timing resolution of this enabling technology is compatible with the rapid throughput
screening of protein-DNA interactions.

<>

<1>Allan, B.W., Beechem, J.M., Lindstrom, W.M., Reich, N.O.
<2>Direct real time observation of base flipping by the EcoRI DNA methyltransferase.
<3>J. Biol. Chem.
<4>273
<5>2368-2373
<6>1998
<7>DNA methyltransferases are excellent prototypes for investigating DNA distortion and enzyme
specificity because catalysis requires the extrahelical stabilization of the target base
within the enzyme active site.  The energetics and kinetics of base flipping by the EcoRI DNA
methyltransferase were investigated by two methods.  First, equilibrium dissociation constants
(KD/DNA) were determined for the binding of the methyltransferase to DNA containing abasic
sites or base analogs incorporated at the target base.  Consistent with a base flipping
mechanism, tighter binding to oligonucleotides containing destabilized target base pairs was
observed.  Second, total intensity stopped flow fluorescence measurements of DNA containing
2-aminopurine allowed presteady-state real time observation of the base flipping transition.
Following the rapid formation of an enzyme-DNA collision complex, a biphasic increase in total
intensity was observed.  The fast phase dominated the total intensity increase with a rate
nearly identical to k(methylation) determined by rapid chemical quench-flow techniques.  The
restacking of the extrahelical base also revealed biphasic kinetics with the recovered
amplitudes from these off-rate experiments matching very closely to those observed during the
base unstacking process.  These results provide the first direct and continuous observation of
base flipping and show that at least two distinct conformation transitions occurred at the
flipped base subsequent to complex formation.  Furthermore, our results suggest that the
commitment to catalysis during the methylation of the target site is not determined at the
level of the chemistry step but rather is mediated by prior intramolecular isomerization
within the enzyme-DNA complex.

<>

<1>Allan, B.W., Garcia, R., Maegley, K., Mort, J., Wong, D., Lindstrom, W., Beechem, J.M., Reich, N.O.
<2>DNA bending by EcoRI DNA methyltransferase accelerates base flipping but compromises specificity.
<3>J. Biol. Chem.
<4>274
<5>19269-19275
<6>1999
<7>EcoRI DNA methyltransferase was previously shown to bend its cognate DNA sequence by 52
degrees and stabilize the target adenine in an extrahelical orientation. We describe the
characterization of an EcoRI DNA methyltransferase mutant in which histidine 235 was
selectively replaced with asparagine. Steady-state kinetic and thermodynamic parameters for
the H235N mutant revealed only minor functional consequences: DNA binding affinity (KDDNA) was
reduced 10-fold, and kcat was decreased 30%. However, in direct contrast to the wild type
enzyme, DNA bending within the mutant enzyme-DNA complexes was not observed by scanning force
microscopy. The bending-deficient mutant showed enhanced discrimination against the
methylation at nontarget sequence DNA. This enhancement of enzyme discrimination was
accompanied by a change in the rate-limiting catalytic step. No presteady-state burst of
product formation was observed, indicating that the chemistry step (or prior event) had become
rate-limiting for methylation. Direct observation of the base flipping transition showed that
the lack of burst kinetics was entirely due to slower base flipping. The combined data show
that DNA bending contributes to the correct assembly of the enzyme-DNA complex to accelerate
base flipping and that slowing the rate of this precatalytic isomerization can enhance
specificity.

<>

<1>Allan, B.W., Reich, N.O.
<2>Targeted base stacking disruption by the EcoRI DNA methyltransferase.
<3>Biochemistry
<4>35
<5>14757-14762
<6>1996
<7>We describe a novel fluorescence-based assay for detecting DNA conformational alterations
within enzyme--DNA complexes.  The target adenine for EcoRI DNA methyltransferase to the
duplex binding site results in a 14-fold increase in fluorescence intensity with a 10 nm blue
shift.  The fluorescence is ~50% of that observed with equimolar free nucleoside, consistent
with extrahelical stabilization of the target base in the enzyme--DNA complex.  The shift in
lambda/max further implies the base is placed into a low dielectric environment.  For
adenine-specific DNA methyltransferases, a hydrophobic pocket composed of highly conserved
amino acids lies proximal to the cofactor binding site.  Substitution of 2-aminopurine
adjacent to the target base also results in detectable changes in fluorescence emission
following complex formation with the methyltransferase.  Thus, other classes of enzymes
hypothesized to utilize base flipping can be investigated by this method.

<>

<1>Allan, B.W., Reich, N.O., Beechem, J.M.
<2>Simultaneous real-time measurement of binding base-flipping, and dissociation base unflipping in EcoRI DNA methyltransferase.
<3>Biophys. J.
<4>74
<5>A69
<6>1998
<7>Many DNA binding proteins generate large changes in the conformation of DNA upon interaction.
High signal-to-noise data of the exact correlation in-time between the binding event and DNA
conformational change, would allow a much deeper understanding of this process.  In this
study, "doubly-labeled" 2-color fluorescent DNA has been synthesized and utilized in
stopped-flow fluorescence experiments, allowing the simultaneous measurement of DNA binding
(via anisotropy changes of 5'-Rhodamine-X labeled 14mers; red-signal) and internal
base-flipping (via changes in the fluorescence intensity of 2-aminopurine at the target
methylation site; blue signal) (see figure for end-points; up arrow, down arrow indicates
protein addition).  The binding-rate (kon = 1.6 x 10^7M-1 s-1) and base-flipping rates in
EcoRI MTase were found to be almost absolutely correlated, whereas differences in off-rate and
"unflipping" were detected.

<>

<1>Allan, B.W., Reich, N.O., Beechem, J.M.
<2>Measurement of the absolute temporal coupling between DNA binding and base flipping.
<3>Biochemistry
<4>38
<5>5308-5314
<6>1999
<7>The absolute temporal couplings between DNA binding and base flipping were examined for the
EcoRI DNA methyltransferase.  The binding event (monitored using rhodamine-x fluorescence
anisotropy) was monophasic with a second-order on-rate of 1.1 x 10^7 M-1 s-1 <- kon <- 2.25 x
10^7 M-1 s-1.  Base-flipping kinetics (monitored using 2-aminopurine fluorescence intensity)
were essentially synchronous with the binding kinetics, with less than a 4 ms delay between
enzyme binding and target base flipping.  The 4 ms delay translates into a base-flipping rate
of at least 195 s-1, when the data are analyzed in terms of a sequential DNA binding and
base-flipping reaction mechanism.  Synchrony of binding and base flipping was only observed
during the first 80% of the reaction, and an additional 20% base-flipping signal occurred well
after DNA binding was complete.  This additional 2AP fluorescence change, with an effective
rate of 0.55 s-1, is an intramolecular isomerization reaction which greatly accelerates the
dissociation of the enzyme from DNA.  The correlation between the dissociation of the
enzyme-DNA complex and the restacking of the extrahelical base also revealed a very tight
coupling of these two events.  Both dissociation and base restacking were found to be
biphasic.  These data are consistent with the following mechanism.  The initial binding rate
and base-flipping rates map very closely with previously determined pre-steady-state
burst-rate kinetics for methyl transfer.  Hence, binding, flipping, and methylation appear to
occur in nearly a single concerted step.  The bound complex then slowly isomerizes (0.1 s-1)
to a distinct configuration that accelerates the product-release phase of the reaction.  The
product-release enzyme configuration dissociates from DNA approximately 8 times faster than
the initial bound complex (0.18 s-1 vs. 0.024 s-1).  When the enzyme dissociates from the DNA
along the product-release pathway, the target base remains in an extrahelical conformation and
restacks at a rate of only 0.6 s-1 .  This "multicolor" fluorescence kinetic approach directly
measures the absolute temporal correlation between DNA binding and base flipping, with
millisecond timing resolution.  The data reveal that even when the B-DNA structure is altered
in a radical manner (e.g., via base flipping), enzymes can perform this operation in a highly
efficient, if not completely concerted manner.

<>

<1>Allard, M.W., Luo, Y., Strain, E., Pettengill, J., Timme, R., Wang, C., Li, C., Keys, C.E., Zheng, J., Stones, R., Wilson, M.R., Musser, S.M., Brown, E.W.
<2>On the evolutionary history, population genetics and diversity among isolates of Salmonella Enteritidis PFGE pattern JEGX01.0004.
<3>PLoS ONE
<4>8
<5>E55254
<6>2013
<7>Facile laboratory tools are needed to augment identification in contamination
events to trace the contamination back to the source (traceback) of Salmonella
enterica subsp. enterica serovar Enteritidis (S. Enteritidis). Understanding the
evolution and diversity within and among outbreak strains is the first step
towards this goal. To this end, we collected 106 new S. Enteriditis isolates
within S. Enteriditis Pulsed-Field Gel Electrophoresis (PFGE) pattern JEGX01.0004
and close relatives, and determined their genome sequences. Sources for these
isolates spanned food, clinical and environmental farm sources collected during
the 2010 S. Enteritidis shell egg outbreak in the United States along with
closely related serovars, S. Dublin, S. Gallinarum biovar Pullorum and S.
Gallinarum. Despite the highly homogeneous structure of this population, S.
Enteritidis isolates examined in this study revealed thousands of SNP differences
and numerous variable genes (n = 366). Twenty-one of these genes from the
lineages leading to outbreak-associated samples had nonsynonymous (causing amino
acid changes) changes and five genes are putatively involved in known Salmonella
virulence pathways. While chromosome synteny and genome organization appeared to
be stable among these isolates, genome size differences were observed due to
variation in the presence or absence of several phages and plasmids, including
phage RE-2010, phage P125109, plasmid pSEEE3072_19 (similar to pSENV), plasmid
pOU1114 and two newly observed mobile plasmid elements pSEEE1729_15 and
pSEEE0956_35. These differences produced modifications to the assembled bases for
these draft genomes in the size range of approximately 4.6 to 4.8 mbp, with S.
Dublin being larger ( approximately 4.9 mbp) and S. Gallinarum smaller (4.55 mbp)
when compared to S. Enteritidis. Finally, we identified variable S. Enteritidis
genes associated with virulence pathways that may be useful markers for the
development of rapid surveillance and typing methods, potentially aiding in
traceback efforts during future outbreaks involving S. Enteritidis PFGE pattern
JEGX01.0004.

<>

<1>Allard, M.W., Muruvanda, T., Strain, E., Timme, R., Luo, Y., Wang, C., Keys, C.E., Payne, J., Cooper, T., Luong, K., Song, Y., Chin, C.S., Korlach, J., Roberts, R.J., Evans, P., Musser, S.M., Brown, E.W.
<2>Fully Assembled Genome Sequence for Salmonella enterica subsp. enterica Serovar Javiana CFSAN001992.
<3>Genome Announcements
<4>2
<5>e00293-14
<6>2014
<7>Volume 1, no. 2, e00081-13, 2013.  Page 1: The article byline should read as given above.
Page 2: The following should be added to the Acknowledgments.  "Research reported in this
publication was supported by the Small Business Innovation Research Program (NIGMS) of the
National Institutes of Health under award number R44GM105125 to R.J.R.  The content is solely
the responsibility of the authors and does not necessarily represent the official views of the
National Institutes of Health."

<>

<1>Allard, M.W., Muruvanda, T., Strain, E., Timme, R., Luo, Y., Wang, C., Keys, C.E., Payne, J., Cooper, T., Luong, K., Song, Y., Chin, C.S., Korlach, J., Roberts, R.J., Evans, P., Musser, S.M., Brown, E.W.
<2>Fully Assembled Genome Sequence for Salmonella enterica subsp. enterica Serovar Javiana CFSAN001992.
<3>Genome Announcements
<4>1
<5>e00081-13
<6>2013
<7>We report a closed genome of Salmonella enterica subsp. enterica serovar Javiana  (S.
Javiana). This serotype is a common food-borne pathogen and is often associated with fresh-cut
produce. Complete (finished) genome assemblies will support pilot studies testing the utility
of next-generation sequencing (NGS) technologies in public health laboratories.

<>

<1>Allen, E.E., Tyson, G.W., Whitaker, R.J., Detter, J.C., Richardson, P.M., Banfield, J.F.
<2>Genome dynamics in a natural archaeal population.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>1883-1888
<6>2007
<7>Evolutionary processes that give rise to, and limit, diversification within
strain populations can be deduced from the form and distribution of genomic
heterogeneity. The extent of genomic change that distinguishes the acidophilic
archaeon Ferroplasma acidarmanus fer1 from an environmental population of the
same species from the same site, fer1(env), was determined by comparing the
1.94-megabase (Mb) genome sequence of the isolate with that reconstructed from 8
Mb of environmental sequence data. The fer1(env) composite sequence sampled
approximately 92% of the isolate genome. Environmental sequence data were also
analyzed to reveal genomic heterogeneity within the coexisting, coevolving
fer1(env) population. Analyses revealed that transposase movement and the
insertion and loss of blocks of novel genes of probable phage origin occur
rapidly enough to give rise to heterogeneity in gene content within the local
population. Because the environmental DNA was derived from many closely related
individuals, it was possible to quantify gene sequence variability within the
population. All but a few gene variants show evidence of strong purifying
selection. Based on the small number of distinct sequence types and their
distribution, we infer that the population is undergoing frequent genetic
recombination, resulting in a mosaic genome pool that is shaped by selection. The
larger genetic potential of the population relative to individuals within it and
the combinatorial process that results in many closely related genome types may
provide the basis for adaptation to environmental fluctuations.

<>

<1>Allen, M.J., Tait, K., Muhling, M., Weynberg, K., Bradley, C., Trivedi, U., Gharbi, K., Nissimov, J., Mavromatis, K., Jensen, C.N., Grogan, G., Ali, S.T.
<2>Genome Sequence of Stenotrophomonas maltophilia PML168, Which Displays Baeyer-Villiger Monooxygenase Activity.
<3>J. Bacteriol.
<4>194
<5>4753-4754
<6>2012
<7>Stenotrophomonas maltophilia PML168 was isolated from Wembury Beach on the English Coast from
a rock pool following growth and selection on agar plates.
Here we present the permanent draft genome sequence, which has allowed prediction
of function for several genes encoding enzymes relevant to industrial
biotechnology, including a novel flavoprotein monooxygenase.

<>

<1>Allet, B.
<2>Fragments produced by cleavage of lambda deoxyribonucleic acid with the Hemophilus parainfluenzae restriction enzyme HpaII.
<3>Biochemistry
<4>12
<5>3972-3977
<6>1973
<7>One of the restricting enzymes extracted from Hemophilus parainfluenzae, HpaII,
is shown to cleave lambda DNA at more than 50 sites.  The resulting DNA
fragments have been prepared from a variety of deletion or
deletion-substitution mutants of lambda, and analyzed by polyacrylamide gel
electrophoresis.  The major DNA segments (larger than 0.3 Mdalton) produced by
digestion of lambda DNA have been mapped and many of the cleavage sites in the
immunity region, in the b2 region, and to the right of the immunity region have
been identified.

<>

<1>Allet, B., Jeppesen, P.G.N., Katagiri, K.J., Delius, H.
<2>Mapping the DNA fragments produced by cleavage of lambda DNA with endonuclease RI.
<3>Nature
<4>241
<5>120-123
<6>1973
<7>None

<>

<1>Allet, B., Rochaix, J.-D.
<2>Structure analysis at the ends of the intervening DNA sequences in the chloroplast 23S ribosomal genes of C. reinhardii.
<3>Cell
<4>18
<5>55-60
<6>1979
<7>All of the chloroplast 23S ribosomal genes of C. reinhardii are interrupted by a 0.87 kb
sequence.  We have sequenced the DNA across the two ends of this intervening element.  In
parallel, we have examined the nucleotide sequences in the corresponding part of the 23S
ribosomal RNA.  This allowed us to locate precisely the boundaries between the coding (that
is, transcribed into mature 23S rRNA) and the noncoding DNA.  The results show that the
intervening sequence is flanked by two identical sets of 3 bp (5'-CGT) oriented as direct
repeats.  In addition, a sequence of 5 bp (5'-CGTGA) lies exactly next to one end and is
found very close (16 bp) to the other end, in the coding part of the gene.  These two sets are
also oriented as direct repeats.  Finally, sequences near one end of the intervening element
are found with a few alterations near the other end, but in an inverted orientation.  Possible
interpretations of these results are discussed.

<>

<1>Allison, D.P., Kerper, P.S., Doktycz, M.J., Spain, J.A., Modrich, P., Larimer, F.W., Thundat, T.
<2>Direct atomic force microscope imaging of EcoRI endonuclease site specifically bound to plasmid DNA molecules.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>8826-8829
<6>1996
<7>Direct imaging with the atomic force microscope has been used to identify specific nucleotide
sequences in plasmid DNA molecules.  This was accomplished using EcoRI(Gln-111), a mutant of
the restriction enzyme that has a 1000-fold greater binding affinity than the wild-type enzyme
but with cleavage rate constants reduced by a factor of 10^4.  ScaI-linearized plasmids with
single (pBS+) and double (pGEM-luc and pSV-beta-galactosidase) EcoRI recognition sites were
imaged, and the bound enzyme was localized to a 50- to 100-nt resolution.  The high affinity
for the EcoRI binding site exhibited by this mutant endonuclease, coupled with an observed low
level of nonspecific binding, should prove valuable for physically mapping large DNA clones by
direct atomic force microscope imaging.

<>

<1>Allison, G.E., Angeles, D., Tran-Dinh, N., Verma, N.K.
<2>Complete genomic sequence of SfV, a serotype-converting temperate bacteriophage of Shigella flexneri.
<3>J. Bacteriol.
<4>184
<5>1974-1987
<6>2002
<7>Bacteriophage SfV is a temperate serotype-converting phage of Shigella flexneri. SfV encodes
the factors involved in type V O-antigen
modification, and the serotype conversion and integration-excision
modules of the phage have been isolated and characterized. We now
report on the complete sequence of the SfV genome (37,074 bp). A total
of 53 open reading frames were predicted from the nucleotide sequence,
and analysis of tile corresponding proteins was used to construct a
functional map. The general organization of the genes in the SfV genome
is similar to that of bacteriophage X, and numerous features of the
sequence are described. The superinfection immunity system of SfV
includes a lambda-like repression system and a P4-like transcription
termination mechanism. Sequence analysis also suggests that SfV encodes
multiple DNA methylases, and experiments confirmed that orf-41 encodes
a Dam methylase. Studies conducted to determine if the phage-encoded
methylase confers host DNA methylation showed that the two S. flexneri
strains analyzed encode their own Dam methylase. Restriction mapping
and sequence analysis revealed that the phage genome has cos sites at
the termini. The tail assembly and structural genes of SfV show
homology to those of phage Mu and Mu-like prophages in the genome of
Escherichia coli O157:H7 and Haemophilus influenzae. Significant
homology (30% of the genome in total) between sections of the early,
regulatory, and structural regions of the SfV genome and the e14 and
KpLE1 prophages in the E. coli K-12 genome were noted, suggesting that
these three phages have common evolutionary origins.

<>

<1>Allison, G.E., Klaenhammer, T.R.
<2>Phage resistance mechanisms in lactic acid bacteria.
<3>Int. Dairy Journal
<4>8
<5>207-226
<6>1998
<7>Dairy fermentations involving Lactococcus lactis and more recently Streptococcus thermophilus,
are commonly attacked by bacteriophages.  Efforts to protect these dairy starter cultures have
resulted in a significant body of knowledge about the bacteriophages, their interactions with
the host, and natural phage defense mechanisms that have evolved within bacteria operating
under the most dynamic and devastating phage environment faced by industrial fermentations.
This paper will overview this area and discuss the novel genetic approaches that are now being
investigated in an effort to provide long term phage protection to dairy starter cultures hat
are used extensively in the industry. A review.

<>

<1>Allison, L., Yee, C.N.
<2>Restriction site mapping is in separation theory.
<3>Comput. Appl. Biosci.
<4>4
<5>97-101
<6>1988
<7>A computer algorithm for restriction-site mapping consists of a generator of
partial maps and a consistency checker.  This paper examines consistency
checking and argues that a method based on separation theory extracts the
maximum amount of information from fragment lengths in digest data.  It results
in the minimum number of false maps being generated.

<>

<1>Alm, R.A., Ling, L.-S.L., Moir, D.T., King, B.L., Brown, E.D., Doig, P.C., Smith, D.R., Noonan, B., Guild, B.C., deJonge, B.L., Carmel, G., Tummino, P.J., Caruso, A., Uria-Nickelsen, M., Mills, D.M., Ives, C., Gibson, R., Merberg, D., Mills, S.
<2>Genomic-sequence comparison of two unrelated isolates of the human gastric pathogen Helicobacter pylori.
<3>Nature
<4>397
<5>176-180
<6>1999
<7>Helicobacter pylori, one of the most common bacterial pathogens of humans, colonizes the
gastric mucosa, where it appears to persist throughout the host's life unless the patient is
treated.  Colonization induces chronic gastric inflammation which can progress to a variety of
diseases, ranging in severity from superficial gastritis and peptic ulcer to gastric cancer
and mucosal-associated lymphoma.  Strain-specific genetic diversity has been proposed to be
involved in the organism's ability to cause different diseases or even be beneficial to the
infected host and to participate in the lifelong chronicity of infection.  Here we compare the
complete genomic sequences of two untreated H. pylori isolates.  This is, to our knowledge,
the first such genomic comparison.  H. pylori was believed to exhibit a large degree of
genomic and allelic diversity, but we find that the overall genomic organization, gene order
and predicted proteomes (sets of proteins encoded by the genomes) of the two strains are quite
similar.  Between 6 to 7% of the genes are specific to each strain, with almost half of these
genes being clustered in a single hypervariable region. Full author list: Alm, R.A., Ling,
L.-S.L., Moir, D.T., King, B.L., Brown, E.D., Doig, P.C., Smith, D.R., Noonan, B., Guild,
B.C., deJonge, B.L., Carmel, G., Tummino, P.J., Caruso, A., Uria-Nickelsen, M., Mills, D.M.,
Ives, C., Gibson, R., Merberg, D., Mills, S.

<>

<1>Alm, R.A., Trust, T.J.
<2>Analysis of the genetic diversity of Helicobacter pylori: the tale of two genomes.
<3>J. Mol. Med.
<4>77
<5>834-846
<6>1999
<7>Infection with Helicobacter pylori has been linked to numerous severe gastroduodenal diseases
including peptic ulcer and gastric cancer. Several techniques have
been used to measure the genetic heterogeneity of H. pylori at several different levels and to
determine whether there is any correlation with severity of disease. The
availability of two completed genome sequences from unrelated strains (J99 and 26,695) has
allowed an analysis of the level of diversity from a large-scale yet detailed
perspective. Although the two chromosomes are organized differently in a limited number of
discrete regions, the genome size and gene order of these two "high-virulence"
(cagA+ and vacA+) H. pylori isolates was found to be highly similar. The regions of
organizational difference are associated with insertion sequences, DNA
restriction/modification genes, repeat sequences, or a combination of the above. A significant
level of variation at the nucleotide level is seen across the genome, providing an
explanation for why the nucleotide-based typing techniques have such high discriminatory power
among independent H. pylori isolates. This nucleotide variation together
with the organizational rearrangements appears to have provided an over-estimation of the gene
order diversity of H. pylori as assessed by pulse-field gel electrophoresis.
Functional assignments are assigned to approximately only 60% of the gene products in each
strain, with one-half of the remaining gene products of unknown function having
homologues in other bacteria, while the remainder appear to be H. pylori-specific. Between 6%
and 7% of the coding capacity of each strain are genes that are absent from the
other strain, with almost one-half of these strain-specific genes located in a single
hypervariable region called the plasticity zone. The majority of the strain-specific genes in
each strain are also H. pylori-specific, with no homologues being identified in the public
databases. Significantly, over one-half of the functionally assigned strain-specific
genes in both H. pylori J99 and 26695 encode DNA restriction/modification enzymes. Analysis of
the level of conservation between orthologues from the two strains indicates
that the H. pylori specific genes have a lower level of conservation than those orthologues to
which a putative function can be assigned. The plasticity zone represents one of
several regions across each genome that is comprised of lower (G+C)% content DNA, some of
which has been detected in self-replicating plasmids, suggesting that both
horizontal transfer from other species and plasmid integration are responsible for the
strain-specific diversity at this locus. These analyses have yielded results with
important implications for understanding the genetic diversity of H. pylori and its associated
diseases, and imply a need to reassess the respective roles of bacterial and host
factors in H. pylori associated diseases.

<>

<1>Almeida, N.F., Yan, S., Lindeberg, M., Studholme, D.J., Schneider, D.J., Condon, B., Liu, H., Viana, C.J., Warren, A., Evans, C., Kemen, E., Maclean, D., Angot, A., Martin, G.B., Jones, J.D., Collmer, A., Setubal, J.C., Vinatzer, B.A.
<2>A draft genome sequence of Pseudomonas syringae pv. tomato T1 reveals a type III effector repertoire significantly divergent from that of Pseudomonas syringae pv. tomato DC3000.
<3>Mol. Plant Microbe Interact.
<4>22
<5>52-62
<6>2009
<7>Diverse gene products including phytotoxins, pathogen-associated molecular
patterns, and type III secreted effectors influence interactions between
Pseudomonas syringae strains and plants, with additional yet
uncharacterized factors likely contributing as well. Of particular
interest are those interactions governing pathogen-host specificity.
Comparative genomics of closely related pathogens with different host
specificity represents an excellent approach for identification of genes
contributing to host-range determination. A draft genome sequence of
Pseudomonas syringae pv. tomato T1, which is pathogenic on tomato but
nonpathogenic on Arabidopsis thaliana, was obtained for this purpose and
compared with the genome of the closely related A. thaliana and tomato
model pathogen P. syringae pv. tomato DC3000. Although the overall genetic
content of each of the two genomes appears to be highly similar, the
repertoire of effectors was found to diverge significantly. Several P.
syringae pv. tomato T1 effectors absent from strain DC3000 were confirmed
to be translocated into plants, with the well-studied effector AvrRpt2
representing a likely candidate for host-range determination. However, the
presence of avrRpt2 was not found sufficient to explain A. thaliana
resistance to P. syringae pv. tomato T1, suggesting that other effectors
and possibly type III secretion system-independent factors also play a
role in this interaction.

<>

<1>Alonso-Vega, P., Normand, P., Bacigalupe, R., Pujic, P., Lajus, A., Vallenet, D., Carro, L., Coll, P., Trujillo, M.E.
<2>Genome Sequence of Micromonospora lupini Lupac 08, Isolated from Root Nodules of  Lupinus angustifolius.
<3>J. Bacteriol.
<4>194
<5>4135
<6>2012
<7>Micromonospora strains have been isolated from diverse niches, including soil, water, and
marine sediments and root nodules of diverse symbiotic plants. In this
work, we report the genome sequence of Micromonospora lupini Lupac 08 isolated
from root nodules of the wild legume Lupinus angustifolious.

<>

<1>Aloui, A., Tagourti, J., El May, A., Petit, D.J., Landoulsi, A.
<2>The effect of methylation on some biological parameters in Salmonella enterica serovar Typhimurium.
<3>Pathol. Biol. (Paris)
<4>59
<5>192-198
<6>2011
<7>Cell growth is tightly coupled to DNA replication and its methylation [Proc Nail Acad Sci US A
93 (1996) 12206-12211]. In a culture medium,
growing of Salmonella Typhimurium (S. Typhimurium) mutant cells
(dam(-)) decreased (2.5 fold) relative to the wild type strain
(dam(+)). In this study, we show that the reason for this growth
deficiency is due to the DNA methylation. The absence of a Dam
methyltransferase protein results in poor growth efficiency and
disturbs the synchrony of replication initiation in vivo, as evaluated
by flow cytometry. On the other hand, we show that lack of methylation
could increase the DNA response to thermal stress (decreasing the DNA
melting temperature, T-m), and the reason for this effect is due to the
methylation status and not to the number of guanine and cytosine bases
(G + C) in the duplex DNA. Our results show that methylation is an
epigenetic factor that may play a key role in the cell growth, the
synchrony of DNA replication [C R Biologies 330 (2007) 576-580] and the
stress protection.

<>

<1>Aloyo, M.C., Campbell, D., Combates, N.J., Gonzalez, J., Kwok, Y., Sheardy, R.D., Winkle, S.A.
<2>Restriction enzymes have altered cleavage rates at sites near certain sequences.
<3>Biophys. J.
<4>64
<5>A280
<6>1993
<7>Under appropriate environmental conditions, e.g. supercoiled DNA, 50-100 uM cobalt hexamine,
the DNA sequence (CG)4 forms non-B or Z-type structures. Restriction enzymes, e.g. BglI,
EcoRV, HaeIII, MboI, NotI, PstI, TaqI, cutting at sites up to circa 50 base pairs from
inserted (CG)4 segments show enhanced cleavage rates (in the presence of cobalt hexamine)
relative to cleavage rates for DNA fragments not containing the (CG)4. We have previously
shown that the sequence TCTTG is a hot spot for the binding of the carcinogen
N-acetoxy-N-acetyl-2-aminofluorene. Restriction enzymes cleaving near an inserted TCTTG
segment have decreased reaction rates. Gel retardation binding studies and exonuclease
footprinting studies suggest that the observed alterations in restriction enzyme reaction
rates arise from changes in the binding of the restriction enzyme to DNA when either (CG)4 or
TCTTG segments are present. These studies suggest that certain sequences can modulate DNA
functions.

<>

<1>Alpert, C.A., Crutz-Le, C.A.M., Malleret, C., Zagorec, M.
<2>Characterization of a theta-type plasmid from Lactobacillus sakei: a potential basis for low-copy-number vectors in lactobacilli.
<3>Appl. Environ. Microbiol.
<4>69
<5>5574-5584
<6>2003
<7>The complete nucleotide sequence of the 13-kb plasmid pRV500, isolated from Lactobacillus
sakei RV332, was determined. Sequence analysis enabled
the identification of genes coding for a putative type I
restriction-modification system, two genes coding for putative
recombinases of the integrase family, and a region likely involved in
replication. The structural features of this region, comprising a putative
ori segment containing 11- and 22-bp repeats and a repA gene coding for a
putative initiator protein, indicated that pRV500 belongs to the pUCL287
subfamily of theta-type replicons. A 3.7-kb fragment encompassing this
region was fused to an Escherichia coli replicon to produce the shuttle
vector pRV566 and was observed to be functional in L. sakei for plasmid
replication. The L. sakei replicon alone could not support replication in
E. coli. Plasmid pRV500 and its derivative pRV566 were determined to be at
very low copy numbers in L. sakei. pRV566 was maintained at a reasonable
rate over 20 generations in several lactobacilli, such as Lactobacillus
curvatus, Lactobacillus casei, and Lactobacillus plantarum, in addition to
L. sakei, making it an interesting basis for developing vectors. Sequence
relationships with other plasmids are described and discussed.

<>

<1>Altermann, E., Attwood, G., Leahy, S., Kelly, W., Ronimus, R., Li, D., Kong, Z., Schofield, L., Dey, D., Tootill, C., Sang, C., Moon, C., Janssen, P.
<2>Phage omega-mru polynucleotides and polypeptides and uses thereof.
<3>International Patent Office
<4>WO 2009041831 A
<5>
<6>2009
<7>The invention encompasses phage omega-mru including phage induction, phage particles, and the
phage genome.  Also encompassed are phage polypeptides, as well as polynucleotides which
encode these polypeptides, expression vectors comprising these polynucleotides, and host cells
comprising these vectors.  The invention further encompasses compositions and methods for
detecting, targeting, permeabilising, and inhibiting microbial cells, especially methanogen
cells, using the disclosed phage, polypeptides, polynucleotides, expression vectors, or host
cells.

<>

<1>Altermann, E., Russell, W.M., Azcarate-Peril, M.A., Barrangou, R., Buck, B.L., McAuliffe, O., Souther, N., Dobson, A., Duong, T., Callanan, M., Lick, S., Hamrick, A., Cano, R., Klaenhammer, T.R.
<2>Inaugural Article: From the Cover: Complete genome sequence of the probiotic lactic acid bacterium Lactobacillus acidophilus NCFM.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>3906-3912
<6>2005
<7>Lactobacillus acidophilus NCFM is a probiotic bacterium that has been produced commercially
since 1972. The complete genome is 1,993,564 nt and
devoid of plasmids. The average GC content is 34.71% with 1,864 predicted
ORFs, of which 72.5% were functionally classified. Nine phage-related
integrases were predicted, but no complete prophages were found. However,
three unique regions designated as potential autonomous units (PAUs) were
identified. These units resemble a unique structure and bear
characteristics of both plasmids and phages. Analysis of the three PAUs
revealed the presence of two R/M systems and a prophage maintenance system
killer protein. A spacers interspersed direct repeat locus containing 32
nearly perfect 29-bp repeats was discovered and may provide a unique
molecular signature for this organism. In silico analyses predicted 17
transposase genes and a chromosomal locus for lactacin B, a class II
bacteriocin. Several mucus- and fibronectin-binding proteins, implicated
in adhesion to human intestinal cells, were also identified. Gene clusters
for transport of a diverse group of carbohydrates, including
fructooligosaccharides and raffinose, were present and often accompanied
by transcriptional regulators of the lacI family. For protein degradation
and peptide utilization, the organism encoded 20 putative peptidases,
homologs for PrtP and PrtM, and two complete oligopeptide transport
systems. Nine two-component regulatory systems were predicted, some
associated with determinants implicated in bacteriocin production and acid
tolerance. Collectively, these features within the genome sequence of L.
acidophilus are likely to contribute to the organisms' gastric survival
and promote interactions with the intestinal mucosa and microbiota.

<>

<1>Althabegoiti, M.J., Lozano, L., Torres-Tejerizo, G., Ormeno-Orrillo, E., Rogel, M.A., Gonzalez, V., Martinez-Romero, E.
<2>Genome Sequence of Rhizobium grahamii CCGE502, a Broad-Host-Range Symbiont with Low Nodulation Competitiveness in Phaseolus vulgaris.
<3>J. Bacteriol.
<4>194
<5>6651-6652
<6>2012
<7>Here we present the genome sequence of Rhizobium grahamii CCGE502. R. grahamii groups with
other newly described broad-host-range species, which are not very
efficient Phaseolus vulgaris symbionts, with a wide geographic distribution and
which constitutes a novel Rhizobium clade.

<>

<1>Altintas, M.M., Ozer, N., Ulgen, K.O.
<2>Purification of TaqI endonuclease from Thermus aquaticus.
<3>J. Chromatogr. A
<4>828
<5>373-381
<6>1998
<7>A purification procedure for the thermostable restriction enzyme TaqI was developed using
high-performance ion-exchange liquid chromatography.  The effects of various operating
conditions on the separation behavior of TaqI endonuclease from the cell extracts were
investigated for optimization and scaling up.  The separation of the enzyme by HPLC was found
to be strongly dependent on the sample volume, slope of linear gradient and order of the
ion-exchange columns.  The final yield of the enzyme is also dependent to a great extent upon
the number of fractionation steps employed to purify the enzyme.  In the present study, 4000 U
TaqI endonuclease per mg. Protein was recovered from 2g Thermus aquaticus cells with a
two-step purification protocol in one day.  The purification factor was 24.  Compared to other
classical methods of purification reported in literature with 4,000 or 32,000 U enzyme from
200 g of Thermus aquaticus cells, HPLC yielded 190,000 U enzyme from 200g cells using cation
and anion HPLC  columns sequentially and thus resulted in a higher efficiency.

<>

<1>Altintas, M.M., Ulgen, K.O.
<2>Growth of Thermus aquaticus and its TaqI endonuclease production.
<3>Acta Biotechnol.
<4>19
<5>45-56
<6>1999
<7>The culture behavior of Thermus aquaticus was characterized.  The response of the bacterium to
various carbon (tryptone, glucose, glycerol) and nitrogen sources (yeast extract, NaNO3,
(NH4)2SO4, leucine, thymine, thiamine, glutamic acid) was studied.  Amino acids did not
support growth, but Castenholz salt medium supplemented with yeast extract and glucose or
tryptone resulted in good growth and production.  A suitable medium composition giving the
highest biomass concentration and enzyme yield was developed.  The simple medium containing
TYE-NaCl resulted in the highest biomass concentration, whereas Castenholz mineral medium
supplemented with tryptone and yeast extract gave the highest specific activity and enzyme
yield.  The effect of inoculum age and size on growth was also investigated in order to
improve the yield and process consistency.  The use of shake flasks inoculated with
precultures at their early or late stationary phase resulted in the same biomass concentration
(0.560+- 0.015 g/l) and similar maximum specific growth rates (0.258 +- 0.003 h-1).  Inoculum
sizes between 1 and 2.5 percent were optimal for cell growth.  As the other papers on
thermophilic microorganisms, including the T. aquaticus YT-1 strain, gave qualitative
information on growth, the results presented here cannot be compared with others on a
quantitative basis.  TaqI endonuclease was purified using a 5 step protocol including cell
disruption, adsorption, precipitation, column chromatography and final dialysis.  The enriched
fraction had a specific activity of 33,600 U TaqI endonuclease per mg protein.

<>

<1>Altschmied, J., Volff, J., Winkler, C., Gutbrod, H., Korting, C., Pagany, M., Schartl, M.
<2>Primary structure and expression of the Xiphophorus DNA-(cytosine-5)-methyltransferase XDNMT-1.
<3>Gene
<4>249
<5>75-82
<6>2000
<7>Small aquarium fishes become increasingly important in the study of normal vertebrate
development and disease. Differential DNA methylation might play a role in these processes. In
the teleost Xiphophorus, a well-established animal model for melanoma formation,
tumour-specific hypomethylation of the melanoma-inducing gene ONC-Xmrk has been observed. We
have isolated a cDNA for the DNA-(cytosine-5)-methyltransferase XDNMT-1 from this organism,
which encodes the first full-length protein from a fish species. Linkage analysis showed that
Xdnmt-1 is different from the Xiphophorus tumour suppressor R, which is involved in the
transcriptional repression of the ONC-Xmrk melanoma oncogene in healthy fish. As methylation
has been implicated in the regulation of ONC-Xmrk expression, XDNMT-1 might play a role by
acting up- or downstream of R. Expression analysis demonstrated that the Xdnmt-1 transcript is
present in all adult tissues and cell lines tested. However, developing embryos show a
spatially and temporally regulated expression pattern suggesting that the enzyme might play a
role during development in fish.

<>

<1>Alvarez, C., Kukutla, P., Jiang, J., Yu, W., Xu, J.
<2>Draft Genome Sequence of Pseudomonas sp. Strain Ag1, Isolated from the Midgut of  the Malaria Mosquito Anopheles gambiae.
<3>J. Bacteriol.
<4>194
<5>5449
<6>2012
<7>A Pseudomonas sp. bacterium was isolated from the midguts of Anopheles gambiae mosquitoes.
Here we present the annotated Pseudomonas sp. draft genome sequence
as a contribution to the efforts of characterization of the mosquito gut
microbiome.

<>

<1>Alvarez, M.A., Chater, K.F., Rosario, M.R.
<2>Complex transcription of an operon encoding the Sall restriction-modification system of Streptomyces albus G.
<3>Mol. Microbiol.
<4>8
<5>243-252
<6>1993
<7>High-resolution S1 nuclease mapping of mRNA synthesized in vivo, in vitro run-off
transcription with RNA polymerase from Streptomyces lividans and gene fusions were used to
analyse the transcriptional organization of the Sall restriction-modification system of
Streptomyces albus G. The sallR and sallM genes that encode the restriction endonuclease and
its cognate methyltransferase constitute an operon which is mainly transcribed from sal-pR1, a
promoter located immediately upstream of sallR, with two possible minor promoters further
upstream. Another promoter, sal-pM, is within the 3' end of the sallR coding region, and
allows expression of the modification gene in the absence of sal-pR1. The sal-pM promoter
might be involved in the establishment of modification prior to restriction endonuclease
activity. Sequences upstream of the apparent transcriptional start sites for sal-pR1 and
sal-pM show similarity with the -10 region of typical vegetatively expressed eubacterial
promoters, but appropriately centered -35 regions are absent.

<>

<1>Alvarez, M.A., Gomez, A., Gomez, P., Brooks, J.E., Rodicio, M.R.
<2>Comparative analysis of expression of the SalI restriction-modification system in Escherichia coli and Streptomyces.
<3>Mol. Gen. Genet.
<4>253
<5>74-80
<6>1996
<7>The salIR and salIM genes encode the endonuclease and methyltransferase components of the SalI
restriction-modification system from Streptomyces albus G.  Expression of the salI genes in
Escherichia coli was investigated and major differences with Streptomyces were found.  In E.
coli there is no detectable expression of the salIR gene due to inactivity of the sal-pR
promoter region.  In the natural host of the system this region directs transcription of the
salI genes as a bicistronic message.  In contrast to salIR, salIM is transcribed in the
heterologous host from a promoter within the salI DNA.  Since sal-pR is not active, the gene
cannot be expressed as part of the salI operon.  It is probably transcribed from sal-pM, a
promoter internal to the operon which allows independent expression of the modification gene
in Streptomyces.  Replacement of sal-pR by the strong pLac promoter allows expression of salIR
in E. coli and enhances expression of salIM.  The resulting strain produces about 10 times
more endonuclease than a Streptomyces clone containing the SalI system under the control of
sal-pR.

<>

<1>Alvarez, M.A., Gomez, A., Gomez, P., Rodicio, M.R.
<2>Expression of the SalI restriction-modification system of Streptomyces albus G in Escherichia coli.
<3>Gene
<4>157
<5>231-232
<6>1995
<7>The salIR and salIM genes of Streptomyces albus G encode the restriction endonuclease (ENase)
and DNA methyltransferase (MTase) of the SalI restriction-modification (R-M) system.  In S.
albus G, the genes constitute an operon that is mainly transcribed from a promoter located
upstream from SalIR, the first gene of the operon.  In addition, a second promoter, at the 3'
end of SalIR, allows independent transcription of the MTase gene.  Expression of salIR and
salIM in Escherichia coli was investigated.  The ENase gene was not expressed in the
heterologous host, probably due to inactivity of the main promoter of the salI operon.  In
contrast to salIR, salIM was functional in E. coli.  Preliminary S1 nuclease mapping
experiments suggest that the alternative promoter of the MTase gene can initiate transcription
in the heterologous, as well as in the homologous host.

<>

<1>Alves, J., Kohler, E., Selent, U., Thielking, V., Wolfes, H., Pingoud, A.
<2>The recognition of DNA by the EcoRV restriction endonuclease: Accuracy and cation dependence.
<3>Biol. Chem. Hoppe Seyler
<4>372
<5>627
<6>1991
<7>In order to investigate the accuracy of the EcoRV restriction endonuclease we have synthesized
20 single stranded oligodeoxynucleotides similarly as described for EcoRI recently. These can
be hybridized to give a set of double stranded substrates comprising the canonical recognition
sequence, the 9 sequences deviating from it by one base pair (star substrates) or the 18
sequences with all possible mismatched base pairs within the recognition sequence (mismatch
substrates).

<>

<1>Alves, J., Pingoud, A., Haupt, W., Langowski, J., Peters, F., Maass, G., Wolff, C.
<2>The influence of sequences adjacent to the recognition site on the cleavage of oligodeoxynucleotides by the EcoRI endonuclease.
<3>Eur. J. Biochem.
<4>140
<5>83-92
<6>1984
<7>We have investigated the influence of the nucleotide sequence adjacent to the
recognition site on the rate of cleavage of DNA by the restriction endonuclease
EcoRI.  For this purpose two decadeoxynucleotides, d(G-G-G-A-A-T-T-C-t-T) (Ia)
and d(A-A-G-A-A-t-T-C-C-C) (Ib) were synthesized.  The duplex Ia Ib is cleaved
by EcoRI preferentially in the dA-rich strand (approximately 10 times over the
dG-rich strand).  The individual nucleotides Ia and Ib are also cleaved by
EcoRI, Ib at a higher rate than Ia and both at a lower rate than Ia Ib.  The
temperature dependence of the reaction rate shows that only double-stranded
oligodeoxynucleotides are substrates for the EcoRI endonuclease.  We have,
furthermore, synthesized oligomers of d(G-G-A-A-T-T-C-C), which contain two,
three and four EcoRI sites, respectively.  These oligodeoxynucleotides are
preferentially cleaved at the sites next to the 5' end, where the recognition
site is only flanked by one dG dC base pair, in contrast to the other sites
which are flanked by three such pairs.  These data indicate that sequences
adjacent to the recognition site influence the rate of cleavage: dA dT base
pairs enhance and dG dC base pairs slow down the hydrolytic activity of the
EcoRI endonuclease.

<>

<1>Alves, J., Pingoud, A., Langowski, J., Urbanke, C., Maass, G.
<2>Two identical subunits of the EcoRI restriction endonuclease co-operate in the binding and cleavage of the palindromic substrate.
<3>Eur. J. Biochem.
<4>124
<5>139-142
<6>1982
<7>The cleavage of radioactively labelled double-stranded d(G-G-A-A-T-T-C-C) was
studied in single turnover experiments with substrate and enzyme both being in
the micromolar range.  The reaction rate was found to increase with enzyme
concentration until a ratio of one tetrameric enzyme to two double-stranded
substrates was reached, further increase of the enzyme concentration then leads
to a sharp decline of the reaction rate.  These findings are interpreted in the
following manner.  (a) Two subunits of the EcoRI endonuclease co-operate in
binding and possibly also in cleaving the palindromic substrate.  (b) The
enzymatic action of the EcoRI endonuclease is inhibited by excess enzyme,
possibly due to unspecific binding of the enzyme-substrate complex.  The
self-inhibition of EcoRI endonuclease has also been observed with
macromolecular substrates.

<>

<1>Alves, J., Ruter, T., Geiger, R., Fliess, A., Maass, G., Pingoud, A.
<2>Changing the hydrogen-bonding potential in the DNA binding site of EcoRI by site-directed mutagenesis drastically reduces the enzymatic activity, not the preference of this restriction endonuclease for cleavage within the site GAATTC.
<3>Biochemistry
<4>28
<5>2678-2684
<6>1989
<7>According to the X-ray structure analysis of an EcoRI-oligodeoxynucleotide
complex [McClarin et al. (1986) Science 234, 1526], sequence specificity is
mediated by 12 hydrogen bonds, 6 from each of the two identical subunits of the
dimeric enzyme to the recognition site -GAATTC-: Arg200 forms two hydrogen
bonds with guanine, while Glu144 and Arg145 form four hydrogen bonds to
adjacent adenine residues.  Changing the hydrogen-bonding potential at the
recognition site without perturbing the rest of the interface should lead to
the recognition of degenerate sequences [Rosenberg et al. (1987) in Protein
Engineering (Oxender, D.L., & Fox, C.F., Eds.) pp 237-250, Liss, New York].  We
have shown previously that replacing Glu144 by Gln and Arg145 by Lys affects
the activity of the enzyme, not, however, its specificity [Wolfes et al. (1986)
Nucleic Acids Res. 14, 9063].  We show now that mutation of Arg200 to Lys, the
double mutation Glu144Arg145 to GlnLys, and the triple mutation
Glu144Arg145Arg200 to GlnLysLys do not lead to a detectable degeneracy of the
specificity of cleavage by EcoRI but significantly impair the catalytic
activity of this enzyme.  A detailed analysis of the steady-state kinetics of
cleavage of pUC8 DNA and a tridecadeoxynucleotide substrate demonstrates that
the reduction in activity for all DNA binding site mutants investigated so far
is mainly due to a decrease in Kcat, with the exception of the Arg200 to Lys
mutant, which is only impaired in its Km.  This observation is confirmed by
nitrocellulose filter experiments which show that this mutant has a lower
affinity toward oligodeoxynucleotide substrates than the other DNA binding site
mutants and a much lower affinity than wild-type EcoRI.  While these and
previous data clearly demonstrate that Glu144, Arg145, and Arg200 are essential
for efficient substrate binding and catalysis, their involvement in the
specific recognition is more intricate than proposed on the basis of the X-ray
structure analysis.  Our results also suggest that the specificity of EcoRI
cannot easily be relaxed, and in particular not without affecting its activity.

<>

<1>Alves, J., Selent, U., Wolfes, H.
<2>Accuracy of the EcoRV restriction endonuclease: binding and cleavage studies with oligodeoxynucleotide substrates containing degenerate recognition sequences.
<3>Biochemistry
<4>34
<5>11191-11197
<6>1995
<7>In order to investigate the accuracy of the EcoRV restriction endonuclease, we have
synthesized a set of double-stranded oligodeoxynucleotides comprising the canonical
recognition sequence, the 9 star sequences (i.e., sequences deviating by one base pair from
the canonical sequence), and the 18 mismatch sequences (i.e. sequences deviating by one base
from the canonical sequence).  For each individual single strand of all these 28 substrates we
have measured the rate of phosphodiester bond cleavage under normal buffer conditions.
Double-strand cleavage of star substrates is at least 5 orders of magnitude slower than
cleavage of the canonical substrate.  In contrast, most of the mismatch substrates are
accepted more readily.  In the absence of the essential cofactor Mg2+, EcoRV binds weakly but
equally to the canonical and degenerate substrates (i.e., KDiss is in the micromolar range).
However, the inactive catalytic site mutant D90A in the presence of Mg2+ binds the canonical
substrate 1-2 orders of magnitude better than degenerate substrates.  Therefore, the EcoRV
endonuclease needs the essential cofactor Mg2+ to create thermodynamic discrimination between
degenerate and canonical sites.  But the main discrimination is kinetically controlled and
takes place during cleavage.  While in the canonical substrate both single strands are cleaved
with an equal velocity, in all other substrates one single strand is cleaved faster than the
other one, resulting in a dissociation of the enzyme from the DNA between the two cuts.  In
vivo this may lead to repair of the erroneous cleavage site by DNA ligases.  The order of
single-strand nicking together with the division of base contacts on both subunits suggests
that correct recognition by one subunit triggers cleavage by the other one.

<>

<1>Alves, J., Thielking, V., Selent, U., Kohler, E., Wolfes, H., Pingoud, A.
<2>The recognition of DNA by the EcoRV restriction endonuclease:  accuracy and cation dependence.
<3>J. Biomol. Struct. Dyn.
<4>8
<5>a007
<6>1991
<7>None

<>

<1>Alves, J., Urbanke, C., Fliess, A., Maass, G., Pingoud, A.
<2>Fluorescence stopped-flow kinetics of the cleavage of synthetic oligodeoxynucleotides by the EcoRI restriction endonuclease.
<3>Biochemistry
<4>28
<5>7879-7888
<6>1989
<7>We have investigated in fluorescence stopped-flow and temperature-jump
experiments the EcoRI-catalyzed cleavage of synthetic palindromic
tridecadeoxynucleotides which contain the EcoRI site but differ in the flanking
sequences.  The overall reaction can be resolved in several reactions which
were analyzed by a nonlinear least-squares fitting procedure on the
experimental data.  The result of this analysis is a minimal scheme that
describes the overall reaction in terms of the rate constants of the individual
reactions.  According to this scheme EcoRI and the tridecadeoxynucleotide
substrates associate in the presence of Mg2+ in a nearly diffusion-controlled
process.  This is followed by a reaction which is or includes the cleavage of
the first phosphodiester bond.  There is no indication for a time-resolved
conformational transition prior to catalysis.  After cleavage of the first
strand, dissociation of the nicked double strand can occur, which then
rearranges to the original palindromic double-stranded substrate and is bound
again by the enzyme.  Alternatively, the nicked double strand can be cleaved in
the second strand.  This reaction is followed by product release from the
enzyme.  The magnitude of the individual rate constants depends on the
substrate used; the differences explain the preference of EcoRI for substrates
that contain AT as compared to GC base pairs next to the recognition site.

<>

<1>Alves, J., Vennekohl, P.
<2>Protein engineering of restriction enzymes.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>393-411
<6>2004
<7>Restriction endonucleases are among the most accurate enzymes known.  Consequently, changing
their very high sequence specificity is one of the scientific goals in studying these enzymes.
This review focuses on protein engineering regarding Type II restriction enzymes.  They
comprise the vast majority of the about 3600 entities listed in REBASE today, although the
number of those studied in detail is far smaller.  First, we will discuss different approaches
to changing the recognition of a given sequence starting with rational design using
site-directed mutagenesis and progressing to random mutagenesis combined with in vivo
selection.  Then, we will describe attempts to lengthen the recognition sequence by designing
additional enzyme contacts to DNA.  As the dimeric nature of all restriction enzymes doubles
the effect of each amino acid exchange, we will also discuss engineering of subunit
composition.  Finally, we will speculate on possible future directions of protein engineering
of restriction enzymes.

<>

<1>Alves, J.M., Serrano, M.G., Maia-da-Silva, F., Voegtly, L.J., Matveyev, A.V., Teixeira, M.M., Camargo, E.P., Buck, G.A.
<2>Genome evolution and phylogenomic analysis of candidatus kinetoplastibacterium, the betaproteobacterial endosymbionts of strigomonas and angomonas.
<3>Genome Biol. Evol.
<4>5
<5>338-350
<6>2013
<7>It has been long known that insect-infecting trypanosomatid flagellates from the genera
Angomonas and Strigomonas harbor bacterial endosymbionts (Candidatus Kinetoplastibacterium or
TPE [trypanosomatid proteobacterial endosymbiont]) that supplement the host metabolism. Based
on previous analyses of other bacterial endosymbiont genomes from other lineages, a
stereotypical path of genome evolution in such bacteria over the duration of their association
with the eukaryotic host has been characterized. In this work, we sequence and analyze the
genomes of five TPEs, perform their metabolic reconstruction, do an extensive phylogenomic
analyses with all available Betaproteobacteria, and compare the TPEs with their nearest
betaproteobacterial relatives. We also identify a number of housekeeping and central
metabolism genes that seem to have undergone positive selection. Our genome structure analyses
show total synteny among the five TPEs despite millions of years of divergence, and that this
lineage follows the common path of genome evolution observed in other endosymbionts of diverse
ancestries.
As previously suggested by cell biology and biochemistry experiments, Ca.
Kinetoplastibacterium spp. preferentially maintain those genes necessary for the biosynthesis
of compounds needed by their hosts. We have also shown that metabolic and informational genes
related to the cooperation with the host are overrepresented amongst genes shown to be under
positive selection. Finally, our phylogenomic analysis shows that, while being in the
Alcaligenaceae family of Betaproteobacteria, the closest relatives of these endosymbionts are
not in the genus Bordetella as previously reported, but more likely in the Taylorella genus.

<>

<1>Alzhanov, D.T., Alzhanova, D.V., Zheleznaya, L.A., Matvienko, N.I.
<2>Thermophilic strain Bacillus species AA contains several site-specific endonucleases.
<3>Biokhimiia
<4>63
<5>636-645
<6>1998
<7>Thermophilic strain Bacillus species AA contains several site-specific endonucleases and three
of them have been identified.  BspAAI recognizes the sequence 5'-C/TCGAG-3' and cleaves it
after the first "C" forming 4-nucleotide 5'-ends and is an isoschizomer of XhoI.  BspAAII
recognizes the sequence 5'-T/CTAGA-3' and cleaves it after the first "T" forming
4-nucleotide 5'-ends and is an isoschizomer of XbaI.  BspAAIII recognizes the sequence
5'-GGATCC-3' and is an isomer of BamHI.  The optimal temperature and pH values are 42-48 C
and 7.0-8.0, respectively.

<>

<1>Alzhanova, D.V., Alzhanov, D.T., Zheleznaya, L.A., Matvienko, N.I.
<2>Isolation and characterization of site-specific endonuclease Bsp123I from thermophilic strain Bacillus species 123.
<3>Biokhimiia
<4>63
<5>207-211
<6>1998
<7>The preparation of site-specific endonuclease Bsp123I was isolated and purified from the
thermophilic strain Bacillus species 123.  Endonuclease Bsp123I recognizes the sequence CGCG
and cleaves it in the middle between nucleotides G and C, producing blunt ends.  Thus, Bsp123I
is an isoschizomer of FnuDII.  The maximal activity of the enzyme is displayed at 42 C, pH
8.0, 100mM NaCl, 10mM MgCl2.

<>

<1>Amadio, A.F., Benintende, G.B., Zandomeni, R.O.
<2>Complete sequence of three plasmids from Bacillus thuringiensis INTA-FR7-4 environmental isolate and comparison with related plasmids from the Bacillus cereus group.
<3>Plasmid
<4>62
<5>172-182
<6>2009
<7>Bacillus thuringiensis is an insect pathogen used worldwide as a
bioinsecticide. It belongs to the Bacillus cereus sensu lato group as well
as Bacillus anthracis and B. cereus. Plasmids from this group of organisms
have been implicated in pathogenicity as they carry the genes responsible
for different types of diseases that affect mammals and insects. Some
plasmids, like pAW63 and pBT9727, encode a functional conjugation
machinery allowing them to be transferred to a recipient cell. They also
share extensive homology with the non-functional conjugation apparatus of
pXO2 from B. anthracis. In this study we report the complete sequence of
three plasmids from an environmental B. thuringiensis isolate from
Argentina, obtained by a shotgun sequencing method. We obtained the
complete nucleotide sequence of plasmids pFR12 (12,095bp), pFR12.5
(12,459bp) and pFR55 (55,712bp) from B. thuringiensis INTA-FR7-4. pFR12
and pFR12.5 were classified as cryptic as they do not code for any obvious
functions besides replication and mobilization. Both small plasmids were
classified as RCR plasmids due to similarities with the replicases they
encode. Plasmid pFR55 showed a structural organization similar to that
observed for plasmids pAW63, pBT9727 and pXO2. pFR55 also shares a tra
region with these plasmids, containing genes related to T4SS and
conjugation. A comparison between pFR55 and conjugative plasmids led to
the postulation that pFR55 is a conjugative plasmid. Genes related to
replication functions in pFR55 are different to those described for
plasmids with known complete sequences. pFR55 is the first completely
sequenced plasmid with a replication machinery related to that of ori44.
The analysis of the complete sequence of plasmids from an environmental
isolate of B. thuringiensis permitted the identification of a near
complete conjugation apparatus in pFR55, resembling those of plasmids
pAW63, pBT9727 and pXO2. The availability of this sequence is a step
forward in the study of the molecular basis of the conjugative process in
Gram positive bacteria, particularly due to the similarity with known
conjugation systems. It is also a contribution to the expansion of the
non-pathogenic B. cereus plasmid gene pool.

<>

<1>Amaral, G.R., Silva, B.S., Santos, E.O., Dias, G.M., Lopes, R.M., Edwards, R.A., Thompson, C.C., Thompson, F.L.
<2>Genome Sequence of the Bacterioplanktonic, Mixotrophic Vibrio campbellii Strain PEL22A, Isolated in the Abrolhos Bank.
<3>J. Bacteriol.
<4>194
<5>2759-2760
<6>2012
<7>Vibrio campbellii PEL22A was isolated from open ocean water in the Abrolhos Bank. The genome
of PEL22A consists of 6,788,038 bp (the GC content is 45%). The number
of coding sequences (CDS) is 6,359, as determined according to the Rapid
Annotation using Subsystem Technology (RAST) server. The number of ribosomal
genes is 80, of which 68 are tRNAs and 12 are rRNAs. V. campbellii PEL22A
contains genes related to virulence and fitness, including a complete
proteorhodopsin cluster, complete type II and III secretion systems, incomplete
type I, IV, and VI secretion systems, a hemolysin, and CTXPhi.

<>

<1>Amari, M., Laguerre, S., Vuillemin, M., Robert, H., Loux, V., Klopp, C., Morel, S., Gabriel, B., Remaud-Simeon, M., Gabriel, V., Moulis, C., Fontagne-Faucher, C.
<2>Genome Sequence of Weissella confusa LBAE C39-2, Isolated from a Wheat Sourdough.
<3>J. Bacteriol.
<4>194
<5>1608-1609
<6>2012
<7>Weissella confusa is a rod-shaped heterofermentative lactic acid bacterium from the family of
Leuconostocaceae. Here we report the draft genome sequence of the
strain W. confusa LBAE C39-2 isolated from a traditional French wheat sourdough.

<>

<1>Amaro, F., Gilbert, J.A., Owens, S., Trimble, W., Shuman, H.A.
<2>Whole-Genome Sequence of the Human Pathogen Legionella pneumophila Serogroup 12 Strain 570-CO-H.
<3>J. Bacteriol.
<4>194
<5>1613-1614
<6>2012
<7>We present the genomic sequence of the human pathogen Legionella pneumophila serogroup 12
strain 570-CO-H (ATCC 43290), a clinical isolate from the Colorado
Department of Health, Denver, CO. This is the first example of a genome sequence
of L. pneumophila from a serogroup other than serogroup 1. We highlight the
similarities and differences relative to six genome sequences that have been
reported for serogroup 1 strains.

<>

<1>Ambalam, P., Pithva, S., Kothari, C., Kothari, R., Parmar, N., Nathani, N.M., Koringa, P.G., Joshi, C.G., Dave, J.M., Vyas, B.R.
<2>Insight into the Draft Genome Sequence of Human Isolate Lactobacillus rhamnosus LR231, a Bacterium with Probiotic Potential.
<3>Genome Announcements
<4>2
<5>e00111-14
<6>2014
<7>Lactobacillus rhamnosus strain LR231 was isolated from the feces of healthy human subjects. It
is observed to be a potential probiotic strain, having a broad
spectrum of antimicrobial activity against a wide range of human pathogens and
food pathogens. Here, we provide the 2.59-Mb draft genome sequence of L.
rhamnosus LR231.

<>

<1>Amin, O., Fardeau, M.L., Valette, O., Hirschler-Rea, A., Barbe, V., Medigue, C., Vacherie, B., Ollivier, B., Bertin, P.N., Dolla, A.
<2>Genome Sequence of the Sulfate-Reducing Bacterium Desulfotomaculum hydrothermale  Lam5(T).
<3>Genome Announcements
<4>1
<5>e00114-12
<6>2013
<7>Here, we report the draft genome sequence of Desulfotomaculum hydrothermale, a
sulfate-reducing, spore-forming bacterium isolated from a Tunisian hot spring. The genome is
composed of 2.7 Mb, with a G+C content of 49.48%, and it contains 2,643 protein-coding
sequences.

<>

<1>Amitsur, M., Benjamin, S., Rosner, R., Chapman-Shimshoni, D., Meidler, R., Blanga, S., Kaufmann, G.
<2>Bacteriophage T4-encoded Stp can be replaced as activator of anticodon nuclease by a normal host cell metabolite.
<3>Mol. Microbiol.
<4>50
<5>129-143
<6>2003
<7>The bacterial tRNALys-specific anticodon nuclease is known as a phage T4 exclusion system. In
the uninfected host cell anticodon nuclease is kept
latent due to the association of its core protein PrrC with the DNA
restriction-modification endonuclease EcoprrI. Stp, the T4-encoded peptide
inhibitor of EcoprrI activates the latent enzyme. Previous in vitro work
indicated that the activation by Stp is sensitive to DNase and requires
added nucleotides. Biochemical and mutational data reported here suggest
that Stp activates the latent holoenzyme when its EcoprrI component is
tethered to a cognate DNA substrate. Moreover, the activation is driven by
GTP hydrolysis, possibly mediated by the NTPase domain of PrrC. The data
also reveal that Stp can be replaced as the activator of latent anticodon
nuclease by certain pyrimidine nucleotides, the most potent of which is
dTTP. The activation by dTTP likewise requires an EcoprrI DNA substrate
and GTP hydrolysis but involves a different form of the latent
holoenzyme/DNA complex. Moreover, whereas Stp relays its activating effect
through EcoprrI, dTTP targets PrrC. The activation of the latent enzyme by
a normal cell constituent hints that anticodon nuclease plays additional
roles, other than warding off phage T4 infection.

<>

<1>Amitsur, M., Morad, I., Chapman-Shimshoni, D., Kaufmann, G.
<2>HSD restriction-modification proteins partake in latent anticodon nuclease.
<3>EMBO J.
<4>11
<5>3129-3134
<6>1992
<7>Phage T4-induced anticodon nuclease triggers cleavage-ligation of the host tRNALys. The enzyme
is encoded in latent form by the optional Escherichia coli locus prr and is activated by the
product of the phage stp gene. Anticodon nuclease latency is attributed to the masking of the
core function prrC by flanking elements homologous with type I restriction -modification genes
(prrA-hsdM and prrD-hadR). Activation of anticodon nuclease in extracts of uninfected prr+
cells required synthetic Stp, ATP and GTP and appeared to depend on endogenous DNA. Stp could
be substituted by a small, heat-stable E. coli factor, hinting that anticodon nuclease may be
mobilized in cellular situations other thant T4 infection. Hsd antibodies recognized the
anticodon nuclease holoenzyme but not the prrC-encoded core. Taken together, these data
indicate that Hsd proteins partake in the latent ACNase complex where they mask the core
factor PrrC. Presumable, this making interaction is disrupted by Stp in conjunction with Hsd
ligands. The Hsd-PrrC interaction may signify coupling and mutal enhancement of two
prokaryotic restriction systems operating at the DNA and tRNA levels.

<>

<1>An, L., Li, Y., Chen, Z., Chang, D., Zhang, X., Guo, Y., Wang, J., Li, T., Dai, W., Liu, C.
<2>Genome Sequence of Enterococcus faecium Strain LCT-EF297, Which Has Specific Biochemical Features.
<3>Genome Announcements
<4>2
<5>e00102-14
<6>2014
<7>An Enterococcus faecium strain was sent into space on the Shenzhou-VIII craft. After space
flight, the strain E. faecium LCT-EF297 was selected based on its
metabolic properties.

<>

<1>Anand, S., Sangwan, N., Lata, P., Kaur, J., Dua, A., Singh, A.K., Verma, M., Kaur, J., Khurana, J.P., Khurana, P., Mathur, S., Lal, R.
<2>Genome Sequence of Sphingobium indicum B90A, a Hexachlorocyclohexane-Degrading Bacterium.
<3>J. Bacteriol.
<4>194
<5>4471-4472
<6>2012
<7>Sphingobium indicum B90A, an efficient degrader of hexachlorocyclohexane (HCH) isomers, was
isolated in 1990 from sugarcane rhizosphere soil in Cuttack, India.
Here we report the draft genome sequence of this bacterium, which has now become
a model system for understanding the genetics, biochemistry, and physiology of
HCH degradation.

<>

<1>Ancora, M., Marcacci, M., Orsini, M., Zilli, K., Di Giannatale, E., Garofolo, G., Camma, C.
<2>Complete Genome Sequence of a Brucella ceti ST26 Strain Isolated from a Striped Dolphin (Stenella coeruleoalba) on the Coast of Italy.
<3>Genome Announcements
<4>2
<5>e00068-14
<6>2014
<7>Brucella spp. are important pathogens affecting a wide range of terrestrial and aquatic
animals. We report the complete and annotated genome sequence of Brucella
ceti ST26 strain TE10759-12, isolated from a striped dolphin (Stenella
coeruleoalba) stranded along the Italian shoreline in March of 2012.

<>

<1>Andersen, J.T., Joergensen, S.T., Rasmussen, M.D., Olsen, P.B., Clausen, I.G.
<2>Improved bacillus host cell.
<3>European Patent Office
<4>EP 2213745 A
<5>
<6>2010
<7>A Bacillus licheniformis mutant host cell derived from a parent B. licheniformis host cell,
which mutant host cell is mutated in one or more gene(s) encoding one or more polypeptide(s)
having proteolytic activity which is at least 80% identical to one or more of the polypeptides
shown in SEQ ID NO's: 2 to 84, wherein the mutant host cell expresses at least 5% less of the
one or more polypeptide(s) having proteolytic activity than the parent host cell, when they
are cultivated under comparable conditions.

<>

<1>Andersen, J.T., Joergsen, S.T., Rasmussen, M.D., Olsen, P.B., Clausen, I.G.
<2>Improved bacillus host cell.
<3>European Patent Office
<4>EP 1696035 A
<5>
<6>2006
<7>A Bacillus licheniformis mutant host cell derived from a parent B. licheniformis host cell,
which mutant host cell is mutated in one or more gene(s) encoding one or more polypeptide(s)
having proteolytic activity which is at least 80% identical to one or more of the polypeptides
shown in SEO ID NO's: 2 to 84, wherein the mutant host cell expresses at least 5% less of the
one or more polypeptide(s) having proteolytic activity than the parent host cell, when they
are cultivated under comparable conditions.

<>

<1>Andersen, J.T., Jorgensen, S.T., Rasmussen, M.D., Olsen, P.B., Clausen, I.G.
<2>Improved bacillus host cell.
<3>International Patent Office
<4>WO 03087149 A
<5>
<6>2003
<7>A Bacillus licheniformis mutant host cell derived from a parent B. licheniformis host cell,
which mutant host cell is mutated in one or more gene(s) encoding one or more polypeptide(s)
having proteolytic activity which is at least 80% identical to one or more of the polypepides
shown in SEQ ID NO's: 2 to 84, wherein the mutant host cell expresses at least 5% less of the
one or more polypeptide(s) having proteolytic activity than the parent host cell, when they
are cultivated under comparable conditions.

<>

<1>Andersen, M.T., Liefting, L.W., Havukkala, I., Beever, R.E.
<2>Comparison of the complete genome sequence of two closely related isolates of 'Candidatus Phytoplasma australiense' reveals genome plasticity.
<3>BMC Genomics
<4>14
<5>529
<6>2013
<7>Background: 'Candidatus Phytoplasma australiense' is associated with at least nine diseases
in Australia and New Zealand. The impact of this
phytoplasma is considerable, both economically and environmentally. The
genome of a NZ isolate was sequenced in an effort to understand its
pathogenicity and ecology. Comparison with a closely related Australian
isolate enabled us to examine mechanisms of genomic
rearrangement.Results: The complete genome sequence of a strawberry
lethal yellows (SLY) isolate of 'Candidatus Phytoplasma australiense'
was determined. It is a circular genome of 959,779 base pairs with 1126
predicted open reading frames. Despite being 80 kbp larger than another
'Ca. Phytoplasma australiense' isolate PAa, the variation between
housekeeping genes was generally less than 1% at a nucleotide level.
The difference in size between the two isolates was largely due to the
number and size of potential mobile units (PMUs), which contributed to
some changes in gene order. Comparison of the genomes of the two
isolates revealed that the highly conserved 5' UTR of a putative
DNA-directed RNA polymerase seems to be associated with insertion and
rearrangement events. Two types of PMUs have been identified on the
basis of the order of three to four conserved genes, with both PMUs
appearing to have been present in the last common ancestor of 'Ca.
Phytoplasma asteris' and 'Ca. Phytoplasma australiense'. Comparison
with other phytoplasma genomes showed that modification methylases
were, in general, species-specific. A putative methylase (xorIIM) found
in 'Ca. Phytoplasma australiense' appeared to have no analogue in any
other firmicute, and we believe has been introduced by way of lateral
gene transfer. A putative retrostransposon (ltrA) analogous to that
found in OY-M was present in both isolates, although all examples in
PAa appear to be fragments. Comparative analysis identified highly
conserved 5' and 3' UTR regions of ltrA, which may indicate how the
gene is excised and inserted.Conclusions: Comparison of two assembled
'Ca. Phytoplasma australiense' genomes has shown they possess a high
level of plasticity. This comparative analysis has yielded clues as to
how rearrangements occur, and the identification of sets of genes that
appear to be associated with these events.

<>

<1>Andersen, P.S., Stegger, M., Aziz, M., Contente-Cuomo, T., Gibbons, H.S., Keim, P., Sokurenko, E.V., Johnson, J.R., Price, L.B.
<2>Complete Genome Sequence of the Epidemic and Highly Virulent CTX-M-15-Producing H30-Rx Subclone of Escherichia coli ST131.
<3>Genome Announcements
<4>1
<5>e00988-13
<6>2013
<7>We report the complete genome sequence, including five complete plasmid sequences, of
Escherichia coli ST131 isolate JJ1886. The isolate was obtained in
2007 in the United States from a patient with fatal urosepsis and belongs to the
virulent, CTX-M-15-producing H30-Rx sublineage.

<>

<1>Anderson, B., McDonald, G.
<2>Construction of DNA libraries of A-T rich organisms using EcoRI star activity.
<3>Anal. Biochem.
<4>211
<5>325-327
<6>1993
<7>Construction of genomic or expression libraries involves size fractionation of the DNA to be
cloned by one of several methods. Methods of fractionation include mechanical shearing,
digestion with DNAse, or digestion with restriction endonucleases that cleave frequently
within a genome (i.e., Sau3AI). The latter method is most frequently used for practical
reasons. Near-random cleavage is best obtained by partial digestion with an enzyme that most
frequently cleaves a given genome. The frequency of cleavage of each restriction endonuclease
within a given genome is dependent on the G + C content of each individual organism.
Therefore, a restriction endonuclease with an A-T rich recognition sequence would be preferred
when cloning DNA from organisms with A-T rich genomes. We describe the use of EcoRI star
activity to generate near-random DNA, from A-T rich genomes, for cloning. Expression/genomic
libraries for three A-T rich bacteria were easily and successfully constructed using this
method.

<>

<1>Anderson, E.S., Felix, A.
<2>Variation in Vi-Phage II of Salmonella typhi.
<3>Nature
<4>170
<5>492-494
<6>1952
<7>The recent description of phenotypic variation in bacteriophages by Luria and
Bertani has prompted us to publish the results of experiments carried out in
1947 which show analogous features.  The object of the work was primarily to
investigate the possibility of tracing a specific Vi-phage type of Salmonella
typhi in which "degraded" variants had arisen (see Craigie and Felix).  It was
hoped also to gain more information about the mechanism of "degradation" of
cultures and the processes concerned in the adaptation of Vi-phage II of
Craigie and Yen to the specific Vi-types of S. typhi.  Degradation in a
spsecific Vi-type can take all forms from the appeaerance of minor
cross-reactions with heterologous phages, typical of partly degraded strains,
to the full susceptibility to all adapted phages which characterizes Type A.

<>

<1>Anderson, E.S., Threlfall, E.J., Carr, J.M., Savoy, L.G.
<2>Bacteriophage restriction in Salmonella typhimurium by R factors and transfer factors.
<3>J. Hyg. (Lond)
<4>71
<5>619
<6>1973
<7>A total of 2716 R factors and transfer factors isolated from Escherichia coli
and Salmonellas of human and animal origin were studied for their
phage-restrictive effects in Salmonella typhimurium phage type 36.  All of 1402
wild fi+ factors were non-restricting.  The F factor of E. coli K12 was unique
among the F-like factors tested in that it inhibited lysis of type 36 by one
typing phage.  In contrast, eleven distinct changes in the phage type of 36
were produced by fi- I-like factors.  I-like plasmids can thus be subdivided by
this method.  I-like R factors and transfer factors from human and animal
enterobacteria were categorized by their phage-restrictive effects in type 36.
Factors resembling delta in this respect predominated among fi- I-like factor
from human E. coli and S. typhimurium and from porcine E. coli.  delta-like and
ColI-like fi-factors were equally distributed in bovine S. typhimurium.
ColI-like factors were commonest in bovine and avian E. coli.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of Halopiger xanaduensis type strain (SH-6T).
<3>Standards in Genomic Sciences
<4>6
<5>31-42
<6>2012
<7>Halopiger xanaduensis is the type species of the genus Halopiger and belongs to the
euryarchaeal family Halobacteriaceae. H. xanaduensis strain SH-6, which is designated as the
type strain, was isolated from the sediment of a salt lake in Inner Mongolia, Lake
Shangmatala. Like other members of the family Halobacteriaceae, it is an extreme halophile
requiring at least 2.5 M salt for growth. We report here the sequencing and annotation of the
4,355,268 bp genome, which includes one chromosome and three plasmids. This genome is part of
a Joint Genome Institute (JGI) Community Sequencing Program (CSP) project to sequence diverse
haloarchaeal genomes.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of Nitratifractor salsuginis type strain (E9I37-1).
<3>Standards in Genomic Sciences
<4>4
<5>322-330
<6>2011
<7>Nitratifractor salsuginis Nakagawa et al. 2005 is the type species of the genus
Nitratifractor, a member of the family Nautiliaceae. The species is of interest
because of its high capacity for nitrate reduction via conversion to N(2) through
respiration, which is a key compound in plant nutrition. The strain is also of
interest because it represents the first mesophilic and facultatively anaerobic
member of the Epsilonproteobacteria reported to grow on molecular hydrogen. This
is the first completed genome sequence of a member of the genus Nitratifractor
and the second sequence from the family Nautiliaceae. The 2,101,285 bp long
genome with its 2,121 protein-coding and 54 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of Ferroglobus placidus AEDII12DO.
<3>Standards in Genomic Sciences
<4>5
<5>50-60
<6>2011
<7>Ferroglobus placidus belongs to the order Archaeoglobales within the archaeal phylum
Euryarchaeota. Strain AEDII12DO is the type strain of the species and was
isolated from a shallow marine hydrothermal system at Vulcano, Italy. It is a
hyperthermophilic, anaerobic chemolithoautotroph, but it can also use a variety
of aromatic compounds as electron donors. Here we describe the features of this
organism together with the complete genome sequence and annotation. The 2,196,266
bp genome with its 2,567 protein-coding and 55 RNA genes was sequenced as part of
a DOE Joint Genome Institute Laboratory Sequencing Program (LSP) project.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of the hyperthermophilic chemolithoautotroph Pyrolobus fumarii type strain (1A).
<3>Standards in Genomic Sciences
<4>4
<5>381-392
<6>2011
<7>Pyrolobus fumarii Blochl et al. 1997 is the type species of the genus Pyrolobus,  which
belongs to the crenarchaeal family Pyrodictiaceae. The species is a
facultatively microaerophilic non-motile crenarchaeon. It is of interest because
of its isolated phylogenetic location in the tree of life and because it is a
hyperthermophilic chemolithoautotroph known as the primary producer of organic
matter at deep-sea hydrothermal vents. P. fumarii exhibits currently the highest
optimal growth temperature of all life forms on earth (106 degrees C). This is
the first completed genome sequence of a member of the genus Pyrolobus to be
published and only the second genome sequence from a member of the family
Pyrodictiaceae. Although Diversa Corporation announced the completion of
sequencing of the P. fumarii genome on September 25, 2001, this sequence was
never released to the public. The 1,843,267 bp long genome with its 1,986
protein-coding and 52 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of Staphylothermus hellenicus P8.
<3>Standards in Genomic Sciences
<4>5
<5>12-20
<6>2011
<7>Staphylothermus hellenicus belongs to the order Desulfurococcales within the archaeal phylum
Crenarchaeota. Strain P8(T) is the type strain of the species and
was isolated from a shallow hydrothermal vent system at Palaeochori Bay, Milos,
Greece. It is a hyperthermophilic, anaerobic heterotroph. Here we describe the
features of this organism together with the complete genome sequence and
annotation. The 1,580,347 bp genome with its 1,668 protein-coding and 48 RNA
genes was sequenced as part of a DOE Joint Genome Institute (JGI) Laboratory
Sequencing Program (LSP) project.

<>

<1>Anderson, I. et al.
<2>Genome sequence of the homoacetogenic bacterium Holophaga foetida type strain (TMBS4(T)).
<3>Standards in Genomic Sciences
<4>6
<5>174-184
<6>2012
<7>Holophaga foetida Liesack et al. 1995 is a member of the phylum Acidobacteria and is of
interest for its ability to anaerobically degrade aromatic compounds and
for its production of volatile sulfur compounds through a unique pathway. The
genome of H. foetida strain TMBS4(T) is the first to be sequenced for a
representative of the class Holophagae. Here we describe the features of this
organism, together with the complete genome sequence (improved high quality
draft), and annotation. The 4,127,237 bp long chromosome with its 3,615
protein-coding and 57 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of the thermophilic sulfate-reducing ocean bacterium Thermodesulfatator indicus type strain (CIR29812(T)).
<3>Standards in Genomic Sciences
<4>6
<5>155-164
<6>2012
<7>Thermodesulfatator indicus Moussard et al. 2004 is a member of the Thermodesulfobacteriaceae,
a family in the phylum Thermodesulfobacteria that is
currently poorly characterized at the genome level. Members of this phylum are of
interest because they represent a distinct, deep-branching, Gram-negative
lineage. T. indicus is an anaerobic, thermophilic, chemolithoautotrophic sulfate
reducer isolated from a deep-sea hydrothermal vent. Here we describe the features
of this organism, together with the complete genome sequence, and annotation. The
2,322,224 bp long chromosome with its 2,233 protein-coding and 58 RNA genes is a
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Anderson, I. et al.
<2>Genome sequence of the flexirubin-pigmented soil bacterium Niabella soli type strain (JS13-8(T)).
<3>Standards in Genomic Sciences
<4>7
<5>210-220
<6>2012
<7>Niabella soli Weon et al. 2008 is a member of the Chitinophagaceae, a family within the class
Sphingobacteriia that is poorly characterized at the genome
level, thus far. N. soli strain JS13-8(T) is of interest for its ability to
produce a variety of glycosyl hydrolases. The genome of N. soli strain JS13-8(T)
is only the second genome sequence of a type strain from the family
Chitinophagaceae to be published, and the first one from the genus Niabella. Here
we describe the features of this organism, together with the complete genome
sequence and annotation. The 4,697,343 bp long chromosome with its 3,931
protein-coding and 49 RNA genes is a part of the Genomic Encyclopedia ofBacteria
andArchaea project.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of the moderately thermophilic mineral-sulfide-oxidizing firmicute Sulfobacillus acidophilus type strain (NAL(T)).
<3>Standards in Genomic Sciences
<4>6
<5>1-13
<6>2012
<7>Sulfobacillus acidophilus Norris et al. 1996 is a member of the genus Sulfobacillus which
comprises five species of the order Clostridiales.
Sulfobacillus species are of interest for comparison to other sulfur and iron
oxidizers and also have biomining applications. This is the first completed
genome sequence of a type strain of the genus Sulfobacillus, and the second
published genome of a member of the species S. acidophilus. The genome, which
consists of one chromosome and one plasmid with a total size of 3,557,831 bp
harbors 3,626 protein-coding and 69 RNA genes, and is a part of the
GenomicEncyclopedia ofBacteria andArchaea project.

<>

<1>Anderson, I. et al.
<2>Genome sequence of Frateuria aurantia type strain (Kondo 67(T)), a xanthomonade isolated from Lilium auratium Lindl.
<3>Standards in Genomic Sciences
<4>9
<5>83-92
<6>2013
<7>Frateuria aurantia (ex Kondo and Ameyama 1958) Swings et al. 1980 is a member of  the
bispecific genus Frateuria in the family Xanthomonadaceae, which is already
heavily targeted for non-type strain genome sequencing. Strain Kondo 67(T) was
initially (1958) identified as a member of 'Acetobacter aurantius', a name that
was not considered for the approved list. Kondo 67(T) was therefore later
designated as the type strain of the newly proposed acetogenic species Frateuria
aurantia . The strain is of interest because of its triterpenoids (hopane
family). F. aurantia Kondo 67(T) is the first member of the genus Frateura whose
genome sequence has been deciphered, and here we describe the features of this
organism, together with the complete genome sequence and annotation. The
3,603,458-bp long chromosome with its 3,200 protein-coding and 88 RNA genes is a
part of the G enomic E ncyclopedia of Bacteria and Archaea project.

<>

<1>Anderson, I. et al.
<2>Complete genome sequence of Halorhabdus utahensis type strain (AX-2).
<3>Standards in Genomic Sciences
<4>1
<5>218-225
<6>2009
<7>Halorhabdus utahensis Waino et al. 2000 is the type species of the genus, which is of
phylogenetic interest because of its location on one of the deepest
branches within the very extensive euryarchaeal family Halobacteriaceae. H.
utahensis is a free-living, motile, rod shaped to pleomorphic, Gram-negative
archaeon, which was originally isolated from a sediment sample collected from the
southern arm of Great Salt Lake, Utah, USA. When grown on appropriate media, H.
utahensis can form polyhydroxybutyrate (PHB). Here we describe the features of
this organism, together with the complete genome sequence, and annotation. This
is the first complete genome sequence of the a member of halobacterial genus
Halorhabdus, and the 3,116,795 bp long single replicon genome with its 3027
protein-coding and 48 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Anderson, I.J. et al.
<2>The complete genome sequence of Staphylothermus marinus reveals differences in sulfur metabolism among heterotrophic Crenarchaeota.
<3>BMC Genomics
<4>10
<5>145
<6>2009
<7>BACKGROUND: Staphylothermus marinus is an anaerobic, sulfur-reducing peptide fermenter of the
archaeal phylum Crenarchaeota. It is the third heterotrophic, obligate sulfur reducing
crenarchaeote to be sequenced and provides an opportunity for comparative analysis of the
three genomes. RESULTS: The 1.57 Mbp genome of the hyperthermophilic crenarchaeote
Staphylothermus marinus has been completely sequenced. The main energy generating pathways
likely involve 2-oxoacid:ferredoxin oxidoreductases and ADP-forming acetyl-CoA synthases. S.
marinus possesses several enzymes not present in other crenarchaeotes including a sodium
ion-translocating decarboxylase likely to be involved in amino acid degradation. S. marinus
lacks sulfur-reducing enzymes present in the other two sulfur-reducing crenarchaeotes that
have been sequenced -- Thermofilum pendens and Hyperthermus butylicus. Instead it has three
operons similar to the mbh and mbx operons of Pyrococcus furiosus, which may play a role in
sulfur reduction and/or hydrogen production. The two marine organisms, S. marinus and H.
butylicus, possess more sodium-dependent transporters than T. pendens and use symporters for
potassium uptake while T. pendens uses an ATP-dependent potassium transporter. T. pendens has
adapted to a nutrient-rich environment while H. butylicus is adapted to a nutrient-poor
environment, and S. marinus lies between these two extremes. CONCLUSION: The three
heterotrophic sulfur-reducing crenarchaeotes have adapted to their habitats, terrestrial vs.
marine, via their transporter content, and they have also adapted to environments with
differing levels of nutrients. Despite the fact that they all use sulfur as an electron
acceptor, they are likely to have different pathways for sulfur reduction.

<>

<1>Anderson, I.J. et al.
<2>Complete genome sequence of Methanoculleus marisnigri Romesser et al. 1981 type strain JR1.
<3>Standards in Genomic Sciences
<4>1
<5>189-196
<6>2009
<7>Methanoculleus marisnigri Romesser et al. 1981 is a methanogen belonging to the order
Methanomicrobiales within the archaeal phylum Euryarchaeota. The type
strain, JR1, was isolated from anoxic sediments of the Black Sea. M. marisnigri
is of phylogenetic interest because at the time the sequencing project began only
one genome had previously been sequenced from the order Methanomicrobiales. We
report here the complete genome sequence of M. marisnigri type strain JR1 and its
annotation. This is part of a Joint Genome Institute 2006 Community Sequencing
Program to sequence genomes of diverse Archaea.

<>

<1>Anderson, I.J., Sieprawska-Lupa, M., Goltsman, E., Lapidus, A., Copeland, A., Glavina, D.Rio.T., Tice, H., Dalin, E., Barry, K., Pitluck, S., Hauser, L., Land, M., Lucas, S., Richardson, P., Whitman, W.B., Kyrpides, N.C.
<2>Complete genome sequence of Methanocorpusculum labreanum type strain Z.
<3>Standards in Genomic Sciences
<4>1
<5>197-203
<6>2009
<7>Methanocorpusculum labreanum is a methanogen belonging to the order Methanomicrobiales within
the archaeal kingdom Euryarchaeota. The type strain Z
was isolated from surface sediments of Tar Pit Lake in the La Brea Tar Pits in
Los Angeles, California. M. labreanum is of phylogenetic interest because at the
time the sequencing project began only one genome had previously been sequenced
from the order Methanomicrobiales. We report here the complete genome sequence of
M. labreanum type strain Z and its annotation. This is part of a 2006 Joint
Genome Institute Community Sequencing Program project to sequence genomes of
diverse Archaea.

<>

<1>Anderson, I.J., Sun, H., Lapidus, A., Copeland, A., Glavina, D.Rio.T., Tice, H., Dalin, E., Lucas, S., Barry, K., Land, M., Richardson, P., Huber, H., Kyrpides, N.C.
<2>Complete genome sequence of Staphylothermus marinus Stetter and Fiala 1986 type strain F1.
<3>Standards in Genomic Sciences
<4>1
<5>183-188
<6>2009
<7>Staphylothermus marinus Fiala and Stetter 1986 belongs to the order Desulfurococcales within
the archaeal phylum Crenarchaeota. S. marinus is a
hyperthermophilic, sulfur-dependent, anaerobic heterotroph. Strain F1 was
isolated from geothermally heated sediments at Vulcano, Italy, but S. marinus has
also been isolated from a hydrothermal vent on the East Pacific Rise. We report
the complete genome of S. marinus strain F1, the type strain of the species. This
is the fifth reported complete genome sequence from the order Desulfurococcales.

<>

<1>Anderson, J.E.
<2>Restriction endonucleases and modification methylases.
<3>Curr. Opin. Struct. Biol.
<4>3
<5>24-30
<6>1993
<7>Restriction endonucleases and modification methylases offer excellent systems for studying
protein-DNA interactions. The past year has seen the experimental verification of some aspects
of the catalytic mechanisms of both types of enzyme. The first structures of methylases were
determined and a number of endonuclease structures should become available soon.

<>

<1>Andersson, S.G.E., Zomorodipour, A., Andersson, J.O., Sicheritz-Ponten, T., Alsmark, U.C.M., Podowski, R.M., Naslund, A.K., Eriksson, A.-S., Winkler, H.H., Kurland, C.G.
<2>The genome sequence of Rickettsia prowazekii and the origin of mitochondria.
<3>Nature
<4>396
<5>133-140
<6>1998
<7>We describe here the complete genome sequence (1,111,523 base pairs) of the obligate
intracellular parasite Rickettsia prowazekii, the causative agent of epidemic typhus.  This
genome contains 834 protein-coding genes.  The functional profiles of these genes show
similarities to those of mitochondrial genes: no genes required for anaerobic glycolysis are
found in either R. prowazekii or mitochondrial genomes, but a complete set of genes encoding
components of the tricarboxylic acid cycle and the respiratory-chain complex is found in R.
prowazekii.  In effect, ATP production in Rickettsia is the same as that in mitochondria.
Many genes involved in the biosynthesis and regulation of biosynthesis of amino acids and
nucleosides in free-living bacteria are absent from R. prowazekii and mitochondria.  Such
genes seem to have been replaced by homologues in the nuclear (host) genome.  The R.
prowazekii genome contains the highest proportion of non-coding DNA (24%) detected so far in a
microbial genome.  Such non-coding sequences may be degraded remnants of 'neutralized' genes
that await elimination from the genome.  Phylogenetic analyses indicate that R. prowazekii is
more closely related to mitochondria than is any other microbe studied so far.

<>

<1>Ando, T.
<2>Novel DNA enzymes:  specificity and application.
<3>Reports Inst. Phys. Chem. Res.
<4>61
<5>239-255
<6>1985
<7>None

<>

<1>Ando, T., Aras, R., Kusugami, K., Peek, R.M., Blaser, M.J., Wassenaar, T.M.
<2>Presence of hrgA and hrgB (iceA2) differentially associated with Helicobacter pylori virulence.
<3>Gastroenterology
<4>124
<5>A272-A273
<6>2003
<7>In the hpyI and hpyIII loci of Helicobacter pylori, the restriction endonuclease-encoding gene
of these Restriction-Modification (R-M)
systems can be replaced by a specific non-related gene: hrgA can
replace hpyIIIR, and hrgB (formerly called iceA2) can replace hpyIR.
Thus, there are 2 genotypes for each locus, either bearing or lacking
the R-gene. Although both hrgB and hrgA have been associated with H.
pylori virulence, we now examined the correlation between the hpyI and
hpyIII genotypes with pathogenicity in greater detail. 174 H. pylori
strains were assessed for sensitivity to MboI (indicative of hpyIIIM
activity) and NlaIII (for hpyIM) digestion. Presence of hpyIIIR or
hrgA, of hpyIR (iceA1) or hrgB, and of cagA and vacA was determined by
PCR and/or LIPA. The induction of IL-8 in co-cultured AGS cells was
determined for representative strains. We assessed the correlation
between genotypes, clinical features, and IL-8 induction. In all
strains examined, hrgA was found to be mutually exclusive with hpyIIIR,
and iceA1 with hrgB; the presence of hrgA and hrgB were independent.
HpyIM and HpyIIIM activities were detected in 100% and 95%,
respectively, of the strains studied. Among 132 Asian strains, the
combination hpyIIIR/iceA1 is the most common genotype (52%) and
hrgA/hrgB is least (4%) common, whereas in 42 Western strains
hpyIIIR/hrgB (40%), and hrgA/hrgB (31%) are the most common. The 33
strains with iceA1 induced significantly more IL-8 (median 1483 pg/ml,
range 287-2898) (p = 0.03) than 30 strains with hrgB (median 955 pg/ml,
range 151-2452), whereas there was no difference in relation to the
hpyIII locus. In Asian strains from gastric cancer patients (n=43, all
cagA positive and vacA s1), 41% were hrgA and 20% were hrgB. This is an
overrepresentation of hrgA, but not of hrgB, compared with non-gastric
cancer diseases. Although gastric cancer strains from Western countries
were not available for analysis, in patients with duodenal or gastric
ulcer, hrgA but not hrgB was overrepresented (71%) compared to
non-ulcer dyspepsia (39%, p<0.05) isolates. Conclusion: In this study,
hrgA presence correlates with H. pylori clinical outcome, whereas iceA1
presence was associated with enhanced IL-8 induction.

<>

<1>Ando, T., Aras, R.A., Kusugami, K., Blaser, M.J., Wassenaar, T.M.
<2>Evolutionary history of hrgA, which replaces the restriction gene hpyIIIR in the hpyIII locus of Helicobacter pylori.
<3>J. Bacteriol.
<4>185
<5>295-301
<6>2003
<7>A recently identified Helicobacter pylori gene, hrgA, was previously reported to be present in
70 (33%) of 208 strains examined (T. Ando, T. M.
Wassenaar, R. M. Peek, R. A. Aras, A. I. Tschumi, L.-J. Van Doorn, K.
Kusugami, and M. J. Blaser, Cancer Res. 62:2385-2389, 2002). Sequence
analysis of nine such strains indicated that in each strain hrgA replaced
hpyIIIR, which encodes a restriction endonuclease and which, together with
the gene for its cognate methyltransferase, constitutes the hpyIII locus.
As a consequence of either the hrgA insertion or independent mutations,
hpyIIIM function was lost in 11 (5%) of the 208 strains examined,
rendering chromosomal DNA sensitive to MboI digestion. The evolutionary
history of the locus containing either hpyIII or hrgA was reconstructed.
By homologous recombination involving flanking sequences, hrgA and hpyIIIR
can replace one another in the hpyIII locus, and there is simultaneous
replacement of several flanking genes. These findings, combined with the
hpyIM/iceA2 locus discovered previously, suggest that the two most
strongly conserved methylase genes of H. pylori, hpyIIIM and hpyIM, are
both preceded by alternative genes that compete for presence at their
loci.

<>

<1>Ando, T., Hayase, E., Ikawa, S., Shibata, T.
<2>Site-specific restriction endodeoxyribonucleases in Bacilli.
<3>Microbiology-1982, American Society for Microbiology, Schlessinger, D., Washington
<4>0
<5>66-70
<6>1982
<7>Biochemical studies on the biological phenomena of restriction and modification
of phages have resulted in the discovery of site-specific endonucleases (type
II restriction endonucleases) that cleave double-stranded DNA at or near
nucleotide sequences specific to each enzyme.  Site-specific endonucleases have
been widely used as tools for research in molecular biology, particularly that
on recombinant DNA.  During restriction and modification of phage Phi105C, we
previously observed that Bacillus subtilis (including B. amyloliquefaciens) had
site-specific restriction endonucleases BamNI and BamNx, which recognize  5'
G^G A T C C 3' and 5' G^G A C C 3' 3' C C T A G^G 5'     3' C C T G^G5'
respectively, and cut the phosphodiester bonds as indicated by arrows.  We
have, since then, systematically screened strains of Bacilli for site-specific
endonucleases.  In this paper, we describe related biochemical studies.

<>

<1>Ando, T., Ishiguro, K., Watanabe, O., Miyake, N., Kato, T., Hibi, S., Mimura, S., Nakamura, M., Miyahara, R., Ohmiya, N., Niwa, Y., Goto, H.
<2>Restriction-modification systems may be associated with Helicobacter pylori virulence.
<3>J. Gastroenterol. Hepatol.
<4>25
<5>S95-S98
<6>2010
<7>Restriction-modification (R-M) systems are exclusive to unicellular organisms and ubiquitous
in the bacterial world. Bacteria use R-M
systems as a defense against invasion by foreign DNA. Analysis of the
genome sequences of Helicobacter pylori strains 26 695 and J99
identified an extraordinary number of genes with homology to R-M genes
in other bacterial species. All H. pylori strains possess their own
unique complement of active R-M systems. All of the methylases that
have been studied so far were present in all major human population
groupings, suggesting that their horizontal acquisition pre-dated the
separation of these populations. The two most strongly conserved
methylase genes of H. pylori, hpy IM and hpy IIIM, are both preceded by
alternative genes that compete for presence at their loci, and
furthermore these genes may be associated with H. pylori pathogenicity.
Further study should investigate the roles of H. pylori R-M systems.

<>

<1>Ando, T., Peek, R.M. Jr., Kusugami, K., Van Doorn, L.-J., Wassenaar, T., Kim, S.-K., Blaser, M.J.
<2>H. pylori strains with the novel gene MboX are correlated with gastric cancer in Asian patients.
<3>Gastroenterology
<4>120
<5>A652
<6>2001
<7>Background: H. pylori exhibits substantial genomic diversity that at least in part influences
clinical outcome.  Type II restriction-modification (R-M) systems are highly diverse between
strains, a characteristic unique for H. pylori.  HpyIII, a restriction enzyme found in H.
pylori, is highly homologous to MboI, which recognizes GATC.  We sought to determine the
variability of hpyIII R-M genotypes in H. pylori strains and whether differences are linked to
geographic origin or clinical features.  Method: We examined 228 strains (US 50, South America
12, Europe 15, Asia 147, Oceana 4), by assessing digestion of chromosomal DNA by MboI, and by
performing PCR for hpyIII.  We assessed the correlation between hpyIII status, other
genotypes, and clinical features among 182 strains (50 US, 85 Japan, 28 China, and 19 Korea).
cagA, vacA, and iceA status were determined by PCR and LIPA.  Results: Eleven (5%) of 228
strains tested were MboI-sensitive, and by PCR, 10 (91%) of these 11 did not have hpyIIIR.  In
these 10 strains, we identified a novel gene (that we termed MboX) in the hpyIII position,
followed by the cognate methylase, hpyIIIM.  MboX has substantial homology with C. jejuni
Cj1602, but no function is known for either.  Strain 88-29 which is Mbo-I-sensitive, possesses
hpyIII, but the upstream promoter is truncated.  We next examined the hpyIII locus in 217
MboI-resistant strains, and found that 148 (68%) have hpyIIIR, and 69 (32%) have MboX.  In
Western countries, MboX is more prevalent (55%) than in Asia (25%) (p<0.0001, c2).  In Asia,
MboX is more prevalent among gastric cancer patients (18/43; 42%) than non-cancer patients
(14/89; 16%) (p=0.001, c2).  Al 132 Asian strains tested were cagA+, but among US strains,
MboX is more prevalent in those that are cagA+ (19/29; 63%) than cagA (7/20; 35%) (p=0.04,
c2).  In vitro, MboX can be replaced by hpyIIIR, and vice versa, by homologous recombination.
Conclusion: The hpyIII locus may contain either hpyIIIR or MboX, followed by hpyIIIM.  MboX is
a novel strain-specific gene that is associated with gastric cancer among H. pylori isolates
from Asian patients.  If this observation is confirmed, MboX may be used in the future to
identify individuals of higher gastric cancer risk.

<>

<1>Ando, T., Shibata, T.
<2>Host-controlled modification and restriction and the relating enzymes in Bacillus subtilis and other Bacillus strains.
<3>Tanpakushitsu Kakusan Koso
<4>22
<5>1012-1019
<6>1977
<7>
<>

<1>Ando, T., Shibata, T., Kazami, J.
<2>Restriction endonucleases.
<3>Tanpakushitsu Kakusan Koso
<4>30
<5>1429-1444
<6>1985
<7>Review - in Japanese

<>

<1>Ando, T., Wassenaar, T.M., Peek, R.M. Jr., Aras, R.A., Tschumi, A.I., van Doorn, L.-Jan., Kusugami, K., Blaser, M.J.
<2>A Helicobacter pylori restriction endonuclease-replacing gene, hrgA, is associated with gastric cancer in Asian strains.
<3>Cancer Res.
<4>62
<5>2385-2389
<6>2002
<7>The sensitivity of Helicobacter pylori chromosomal DNA to MboI digestion was investigated in
208 strains from several continents. Only
11 (5%) of the strains were sensitive to MboI, and it was hypothesized that
HpyIII, a type II restriction/modification enzyme with sequence
homology to MboI, mediated the protection. This was confirmed by PCR
analysis of the gene locus of hpyIII, normally composed of hpyIIIR and
hpyIIIM. In all but one strain sensitive to MboI, no PCR product of
hpyIIIR was obtained. In contrast, all strains yielded a product for
hpyIIIM, independent of MboI phenotype. Further examination of the
hpyIII locus in strains lacking a hpyIIIR PCR product identified a
novel gene, hrgA, upstream of hpyIIIM. All 208 strains examined had
either hpyIIIR or hrgA, but not both, upstream of hpyIIIM. Although
hrgA has homology with a Campylobacter jejuni gene (Cj1602), its
function is not known. In Western countries, hrgA was more prevalent
(53%) than in Asia (25%; P<0.0001, chi2). In Asia, hrgA was more
prevalent among gastric cancer patients (18 of 43; 42%) than among
noncancer patients (16 of 95; 17%; P=0.001, chi2). All 143 Asian
strains tested were cagA+, but among Western strains, hrgA was more
prevalent in cagA+ strains (26 of 42; 62%) than in cagA- strains (9 of
23; 39%; P=0.04, chi2). In coculture with epithelial cells, hpyIIIR and
hrgA strains did not show any significant differences in interleukin-8
induction and apoptosis. Although a direct function for hrgA in
virulence could not be demonstrated, our data indicate that hrgA is a
strain-specific gene that might be associated with gastric cancer among
H. pylori isolates from Asian patients.

<>

<1>Ando, T., Wassenaar, T.M., Peek, R.M., Aras, R.A., Kusugami, K., van Doom, L., Blaser, M.J.
<2>hypX shares a genomic locus with the restriction endonuclease gene hpyIIIR in Helicobacter pylori.
<3>Int. J. Med. Microbiol.
<4>291S
<5>28
<6>2001
<7>A novel gene, hypX, with potential predictive value to differentiate virulence amongst
cagA-positive H. pylori strains has been identified.  This gene may replace hpyIIIR, a
restriction endonuclease that, together with its methyltransferase, constitutes the hpyIII
locus, which is homologous to the MboI R-M system.  hypX has substantial homology with Cj1602
in C. jejuni, but the function for both is unknown.  From 208 strains examined, originating
from several continents, hypX and hpyIIIR were found to be mutually exclusive in the hpyIII
locus.  Of the 138 strains bearing hpyIIIR, which we designated Type I, 137 (99%) are Mbo-I
resistant.  Of the 70 strains bearing hypX, or Type II, 14% are MboI-sensitive.  Type I and
Type II strains were compared for linkage to geographic origin or clinical features.  In
Western countries, hypX is more prevalent (53%) than in Asian (25%) (p is less than or equal
to 0.001, c2).  In Asian, hypX is more prevalent among gastric cancer patients (18/43; 42%)
than non-cancer patients (16/95; 17%) (p=0.001, X2).  All 143 Asian strains tested were cagA+
strains (24/39; 62%) than in cagA- strains (9/22; 41%) (p=0.04, X2).  Although a direct
function for phpX in virulence was not examined, we postulate that hypX is a strain-specific
marker that might be associated with gastric cancer among H. pylori isolates from Asian
patients.  Gene -specific assays for hypX may provide tools to identify individuals of higher
gastric cancer risk in populations in which cagA-positive strains predominate.

<>

<1>Ando, T., Xu, Q., Torres, M., Kusugami, K., Israel, D.A., Blaser, M.J.
<2>Restriction-modification system differences in Helicobacter pylori are a barrier to interstrain plasmid transfer.
<3>Mol. Microbiol.
<4>37
<5>1052-1065
<6>2000
<7>Helicobacter pylori cells are naturally competent for the uptake of both plasmid and
chromosomal DNA. However, we demonstrate that there are strong barriers to transformation of
H. pylori strains by plasmids derived from unrelated strains. We sought to determine the
molecular mechanisms underlying these barriers. Transformation efficiency was assessed using
pHP1, an Escherichia coli-H. pylori shuttle vector conferring kanamycin resistance.
Transformation of 33 H. pylori strains was attempted with pHP1 purified from either E. coli or
H. pylori, and was successfully introduced into only 11 strains. Digestion of H. pylori
chromosomes with different restriction endonucleases (REs) showed that DNA methylation
patterns vary substantially among strains. The strain most easily transformed, JP26, was found
to have extremely low endogenous RE activity and to lack a restriction-modification (R-M)
system, homologous to MboI, which is highly conserved among H. pylori strains. When we
introduced this system to JP26, pHP1 from MboI.M+ JP26, but not from wild-type JP26,
transformed MboI R-M+ JP26 and heterologous MboI R-M+ wild-type H. pylori strains. Parallel
studies with pHP1 from dam+ and dam- E. coli strains confirmed these findings. These data
indicate that the endogenous REs of H. pylori strains represent a critical barrier to
interstrain plasmid transfer among H. pylori.

<>

<1>Andreeva, I.S., Afinogenova, G.N., Lebedev, I.R., Nadolinnaya, I.G., Pustoshilova, N.M.
<2>Kpn378I - a type II restriction endonuclease from Klebsiella pneumoniae.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>2
<5>31-32
<6>1991
<7>A strain of Klebsiella pneumoniae isolated from the air showed the presence of
a type II restriction endonuclease. Selection for clones that would not form
capsula resulted in the isolation of a non-pathogenic subclone. The yield of
the highly purified enzyme is 40,000 units per g of cells. Judging from the
fragmentation pattern of lambda phage DNA, the sequence specificity of
R.Kpn378I is indistinguishable from that of SacII endonuclease. Optimal
reaction conditions are: 50-70mM NaCl/KCl, 10 mM MgCl2, pH 7.5-8.5, 37C. Unlike
SacII, Kpn378I is not inhibited by NaCl or KCl at concentrations up to 120mM.

<>

<1>Andreeva, I.S., Afinogenova, G.N., Lebedev, L.R., Pustoshilova, N.M., Gordienko, N.Y., Maistrenko, G.G.
<2>Site specific restriction endonuclease Rme21I from Rhizobium meliloti.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>30-31
<6>1991
<7>A new site specific restriction endonuclease Rme21I from Rhizobium meliloti has
been shown to recognize the following nucleotide sequence 5'-ATCGAT-3' in the
doublestranded DNA.  Thus, the enzyme is a true isoschizomer of the restriction
endonucleases Bsu15I and ClaI.

<>

<1>Andreeva, I.S., Atinogenova, G.N., Lebedev, L.R., Pustoshelova, N.M., Gordienko, N.Y., Maystrenko, G.G.
<2>The method of isolation of the restriction endonuclease recognizing the nucleotide sequence GTCGAC.
<3>Soviet Patent Office
<4>SU 1752769 A
<5>
<6>1992
<7>The present invention provides the method of producing of the restriction endonuclease
recognizing the GTCGAC nucleotide sequence from Rhizobium trifolii containing only one
restriction endonuclease by 1) extraction of a lysate in a two-phase mixture of 7%
polyethyleneglycol, 2% dextran and 0.7 M NaCl; 2) chromatography on phosphocellulose P-11 and
heparin-sepharose with a linear salt gradient of 0.5-0.7 M NaCl. The yield of enzyme was 6-8
fold higher.

<>

<1>Andreeva, I.S., Puchkova, L.I., Saranina, I.V., Lokhova, I.A., Repin, V.E.
<2>A strain of Bacillus stearothermophilus producing a restriction endonuclease cleaving the sequence GGTNACC.
<3>Russian Patent Office
<4>RU 2115728 C
<5>
<6>1998
<7>
<>

<1>Andres, S., Skoglund, A., Nilsson, C., Krabbe, M., Bjorkholm, B., Engstrand, L.
<2>Type I Restriction-Modification Loci Reveal High Allelic Diversity in Clinical Helicobacter pylori Isolates.
<3>Helicobacter
<4>15
<5>114-125
<6>2010
<7>Background: A remarkable variety of restriction-modification (R-M) systems is
found in Helicobacter pylori. Since they encompass a large portion of
the strain-specific H. pylori genes and therefore contribute to genetic
variability, they are suggested to have an impact on disease outcome.
Type I R-M systems comprise three different subunits and are the most
complex of the three types of R-M systems.
Aims:
We investigated the genetic diversity and distribution of type I
R-M systems in clinical isolates of H. pylori.
Material and methods:
Sixty-one H. pylori isolates from a Swedish hospital based
case-control study and 6 H. pylori isolates of a Swedish
population-based study were analyzed using polymerase chain reaction
for the presence of the three R-M systems' subunits. Representative
gene variants were sequenced.
Results:
Although the hsdM and hsdR genes appeared conserved in our clinical
H. pylori isolates, the sequences of the hsdS loci were highly
variable. Despite their sequence diversity, the genes per se were
present at high frequencies. We identified a number of novel allelic
hsdS variants, which are distinct from corresponding hsdS loci in the
sequenced H. pylori strains 26695, J99 and HPAG1. In analyses of paired
H. pylori isolates, obtained from the same individuals with a 4-year
interval, we observed genetic modifications of hsdS genes in patients
with atrophic gastric mucosa.
Discussion:
We propose that the genetic variability of hsdS genes in a
bacterial population will give rise to new specificities of these
enzymes, which might lead to adaptation to an ever-changing gastric
environment.

<>

<1>Andres-Barrao, C., Falquet, L., Calderon-Copete, S.P., Descombes, P., Ortega, P.R., Barja, F.
<2>Genome sequence of high acetic acid resistant bacteria: Gluconacetobacter europaeus LMG 18890T, Gluconacetobacter europaeus LMG 18494 (references  strains), Gluconacetobacter europaeus 5P3 and Gluconacetobacter oboediens  174Bp2 (isolated from vinegar).
<3>J. Bacteriol.
<4>193
<5>2670-2671
<6>2011
<7>Bacteria of the genus Gluconacetobacter are usually involved in the industrial production of
vinegars with high acetic acid concentration. We
describe here the genome sequence of three Gluconacetobacter europaeus
strains, a very common bacterial species from industrial fermenters, as
well as of a Gluconacetobacter oboediens strain.

<>

<1>Andrews, K.T., Patel, B.K.C., Clarke, F.M.
<2>FgoI, a type II restriction endonuclease from the thermoanaerobe Fervidobacterium gondwanense AB39T.
<3>Anaerobe
<4>4
<5>227-232
<6>1998
<7>Restriction endonuclease activity was detected in 11 out of 13 Fervidobacterium isolates,
including F. islandicum H21T, F. gondwanense AB39T, and nine other Fervidobacterium-like
strains isolated from the Great Artesian Basin of Australia.  The restriction endonuclease
from F. gondwanense AB39T was partially purified and designated FgoI.  FgoI recognized a 4
nucleotide sequence 5'-CTAG-3' and cleaved between nucleotides C and T to produce a 2 base
5' overhang (5'C/TAG-3').  As predicted from the enzyme recognition and cleavage
specificity, FgoI was found to cleave phage DNA 13 times, phiX174 three times, pBR322 five
times, pUC18 four times, and pSK six times.  FgoI exhibited a broad temperature optimum range
(between 60 to 70oC) and was active at pH 6.5 to 8.5, but not at pH 9.0.  Manganese could
replace magnesium as a cofactor for activity, but not cobalt chloride, calcium chloride,
cupric chloride, or zinc chloride.  The restriction endonuclease was completely inactivated by
phenol/chloroform extraction and was heat inactivated at 80 C for 60 min or at 100 C for 15
min.  FgoI has been identified as a heat stable isoschizomer of the Type II restriction
endonucleases, MaeI and BfaI.

<>

<1>Andriashvili, I.A., Kvachadze, L.I., Vashakidze, R.P., Adamiya, R.S., Chanishvili, T.G.
<2>Molecular mechanism of the protection of phage DNA from restriction endonuclease of Staphylococcus aureus cells.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>43-45
<6>1986
<7>A representative of the group of virulent staphylococcal bacteriophages Sb-I is
not limited by the systems of host specificity of DNA in reproduction in
Staphylococcus aureus cells.  It was also shown that the DNA of this phage is
not subjected to fragmentation by Sau3A and other restriction endonucleases
specific for the GATC site.  By restriction analysis of cloned fragments of
phage DNA in the plasmid pBR322, replicated in E. coli HB101, it was
established that virus-specific protection is determined by the absence of a
GATC site in the phage genome.  The evolutionary nature of the elimination of
the recognition site in the DNA of virulent staphylophages as a method of
protection from the main type of restriction of the phage DNA in S. aureus
cells is discussed.

<>

<1>Ang, M.Y., Dymock, D., Tan, J.L., Thong, M.H., Tan, Q.K., Wong, G.J., Paterson, I.C., Choo, S.W.
<2>Genome Sequence of Parvimonas micra Strain A293, Isolated from an Abdominal Abscess from a Patient in the United Kingdom.
<3>Genome Announcements
<4>1
<5>e01025-13
<6>2013
<7>Parvimonas micra is an important oral microbe that has the ability to grow and proliferate
within oral biofilms and is involved in periodontal disease, leading
to gingival bleeding, gingival recession, alveolar bone loss, and tooth mobility.
However, occasionally these normally oral pathogens can cause infections at other
sites in the body. We present the genome sequence of Parvimonas micra strain
A293, a smooth Parvimonas micra strain isolated from an abdominal abscess from a
patient at Barts Hospital, London, United Kingdom.

<>

<1>Ang, M.Y., Dymock, D., Tan, J.L., Thong, M.H., Tan, Q.K., Wong, G.J., Paterson, I.C., Choo, S.W.
<2>Genome Sequence of Fusobacterium nucleatum Strain W1481, a Possible New Subspecies Isolated from a Periodontal Pocket.
<3>Genome Announcements
<4>2
<5>e00009-14
<6>2014
<7>Fusobacterium nucleatum is a bacterial species commonly detected in dental plaque within the
human oral cavity, with some strains associated with periodontal
disease, one of the most common clinical bacterial infections in the human body.
The exact mechanisms of its pathogenesis are still not completely understood. In
this study, we present the genome sequence and annotation of F. nucleatum strain
W1481, isolated from a periodontal pocket of a dental patient at the University
of Bristol, United Kingdom, the 16S rRNA gene sequencing of which showed it to be
markedly different from the five previously named subspecies.

<>

<1>Angelov, A., Liebl, S., Ballschmiter, M., Bomeke, M., Lehmann, R., Liesegang, H., Daniel, R., Liebl, W.
<2>Genome Sequence of the Polysaccharide-Degrading, Thermophilic Anaerobe Spirochaeta thermophila DSM 6192.
<3>J. Bacteriol.
<4>192
<5>6492-6493
<6>2010
<7>Spirochaeta thermophila is a thermophilic, free-living anaerobe that is able to degrade
various alpha- and beta-linked sugar polymers, including
cellulose. We report here the complete genome sequence of S. thermophila
DSM 6192, which is the first genome sequence of a thermophilic,
free-living member of the Spirochaetes phylum. The genome data reveal a
high density of genes encoding enzymes from more than 30 glycoside
hydrolase families, a noncellulosomal enzyme system for (hemi)cellulose
degradation, and indicate the presence of a novel carbohydrate-binding
module.

<>

<1>Angelov, A., Voss, J., Liebl, W.
<2>Characterization of Plasmid pPO1 from the Hyperacidophile Picrophilus oshimae.
<3>Archaea
<4>0
<5>723604
<6>2011
<7>Picrophilus oshimae and Picrophilus torridus are free-living, moderately thermophilic and
acidophilic organisms from the lineage of
Euryarchaeota. With a pH optimum of growth at pH 0.7 and the ability to
even withstand molar concentrations of sulphuric acid, these organisms
represent the most extreme acidophiles known. So far, nothing is known
about plasmid biology in these hyperacidophiles. Also, there are no
genetic tools available for this genus. We have mobilized the 7.6 Kbp
plasmid from P. oshimae in E. coli by introducing origin-containing
transposons and described the plasmid in terms of its nucleotide
sequence, copy number in the native host, mode of replication, and
transcriptional start sites of the encoded ORFs. Plasmid pPO1 may
encode a restriction/modification system in addition to its replication
functions. The information gained from the pPO1 plasmid may prove
useful in developing a cloning system for this group of extreme
acidophiles.

<>

<1>Anglana, M., Bacchetti, S.
<2>Construction of a recombinant adenovirus for efficient delivery of the I-SceI yeast endonuclease to human cells and its application in the in vivo cleavage of chromosomes to expose new potential telomeres.
<3>Nucleic Acids Res.
<4>27
<5>4276-4281
<6>1999
<7>We have constructed a replication-defective adenovirus vector encoding the yeast I- SceI
endonuclease under the control of the murine cytomegalovirus immediate-early gene promoter
(AdM SceI) for efficient delivery of this enzyme to mammalian cells.  We present evidence of
AdM SceI-mediated I-Sce I protein expression and cleavage activity in replication-permissive
293 cells, and of cleavage of chromosomes in vivo in both 293 cells and in non-permissive
human cells. We have exploited this system for the generation of chromosomes capped by
artificial telomeric sequences in cells with integrated plasmids containing telomeric DNA
arrays adjacent to an I-SceI recognition site. The properties of the AdM SceI virus described
here make it a useful tool for studying biological processes involving induction of DNA
breaks, recombination and gene targeting in cells grown in culture and in vivo.

<>

<1>Aniello, F., Locascio, A., Fucci, L., Geraci, G., Branno, M.
<2>Isolation of cDNA clones encoding DNA methyltransferase of sea urchin P. lividus: expression during embryonic development.
<3>Gene
<4>178
<5>57-61
<6>1996
<7>A full-length cDNA, encoding a DNA (cytosine-5)-methyltransferase has been assembled from a
series of overlapping cDNA clones isolated from P. lividus sea urchin embryo cDNA libraries.
The cDNA contains 103 bp 5'-UTR, 4839 bp open reading frame corresponding to a 1612 amino
acids protein and 2240 bp 3'-UTR including a terminal 18-bp poly(A) tail.  Both the cDNA and
the encoded protein are the longest so far reported for DNA Mtases.  The protein shows five
distinct and sequential regions of identity with the other animal DNA Mtases, with values of
identity from zero to 80%.  Northern blot analyses reveal a single RNA band of about 7.5 kb in
length showing a highly regulated concentration pattern during development with peak value at
the four blastomere stage.

<>

<1>Aniello, F., Villano, G., Corrado, M., Locascio, A., Russo, M.T., D'Aniello, S.D., Francone, M., Fucci, L., Branno, M.
<2>Structural organization of the sea urchin DNA (cytosine-5)-methyltransferase gene and characterization of five alternative spliced transcripts.
<3>Gene
<4>302
<5>1-9
<6>2003
<7>Sea urchin DNA (cytosine-5)-methyltransferase (Dnmt1) that is responsible for maintenance of
DNA methylation patterns clearly shares similarity with
various Dnmt1s identified in vertebrates. In this study, we determined the
structure of the sea urchin Dnmt1 gene by screening a genomic library of
the sea urchin Paracentrotus lividus with the complementary DNA (cDNA) as
probe. Analysis of the positive clones demonstrated that the Dnmt1 gene
consists of 34 exons and 33 introns spanning a distance of 35 kb. All
exon-intron junction sequences agree with the GT/AG consensus with the
exception of the 3' acceptor site of intron 8 where CT replaces AG
consensus.The differences in the total number of exons between sea urchin
and mouse genes reside mainly in the N-terminal region of the protein
(exons 5-7 of the sea urchin, exons 5-12 of the mouse) where there is very
low similarity in the amino acid sequence.By reverse
transcription-polymerase chain reaction using oligonucleotides spanning
different regions of the cDNA we carried out a comprehensive analysis of
alternative splicing of the Dnmt1 messenger RNA (mRNA) in sea urchin
embryos at different stages of development. We demonstrated the presence
of at least five alternative spliced mRNAs that are regulated during
development and are translated in truncated or deleted proteins.

<>

<1>Ankenbauer, W., Penzberg, D.E., Jarsch, M., Kessler, C., Laue, F.
<2>Type II restriction endonuclease DsaV.
<3>German Patent Office
<4>DE 3924709 A
<5>
<6>1991
<7>The Type II restriction enzyme DsaV has the following recognition sequence and digests at the
marked positions: ^CCNGG, and is originating from a microorganism of the genus
Dactylococcopsis. It can be used for the recognition and digestion of double stranded DNA with
the palindromic sequence CCNGG.

<>

<1>Ankrah, N.Y., Lane, T., Budinoff, C.R., Hadden, M.K., Buchan, A.
<2>Draft Genome Sequence of Sulfitobacter sp. CB2047, a Member of the Roseobacter Clade of Marine Bacteria, Isolated from an Emiliania huxleyi Bloom.
<3>Genome Announcements
<4>2
<5>e01125-14
<6>2014
<7>We announce the draft genome sequence of Sulfitobacter sp. strain CB2047, a marine bacterium
of the Roseobacter clade, isolated from a phytoplankton bloom.
The genome encodes pathways for the catabolism of aromatic compounds as well as
transformations of carbon monoxide and sulfur species. The strain also encodes a
prophage as well as the gene transfer agent (GTA), both of which are prevalent
among members of the Rhodobacterales order.

<>

<1>Ankri, S., Reyes, O., Leblon, G.
<2>Improved electro-transformation of highly DNA-restrictive corynebacteria with DNA extracted from starved Escherichia coli.
<3>FEMS Microbiol. Lett.
<4>140
<5>247-251
<6>1996
<7>Differences of up to 33,000-fold in electro-transformability of highly DNA restrictive
corynebacteria are observed in the DNA of a shuttle plasmid extracted from Escherichia coli
hosts propagated in different nutritional conditions.  Growth of the host in minimal medium
increases plasmid transformability is reverted by supplementing with methionine, an obligate
S-adenosyl-methionine (SAM) precursor.  This suggests that an E. coli nutritionally modulated
SAM-dependent DNA-methyltransferase may be involved in this phenomenon.

<>

<1>Anton, B.P.
<2>Phylogenomic analysis of Escherichia coli methyltransferases and characterization of a novel subfamily of enzymes performing methyl transfer.
<3>Ph.D. Thesis, Boston University
<4>
<5>1-271
<6>2011
<7>Methyltransferases, which catalyze the transfer of a methyl group from the donor
S-adenosyl-L-methionine to a substrate molecule, are an ancient superfamily of enzymes
comprising significant sequence and structural diversity.  They are involved with nearly every
conceivable biological process, having evolved to perform this same biochemical reaction on a
wide variety of substrates including DNA, RNA, proteins, lipids, and small molecules.
Identifying sequence-structure-function relationships within this superfamily presents
considerable challenges, and this thesis comprises several computational and experimental
projects directed towards that end.  We examined different types of prokaryotic
methyltransferases by phylogenetic profiling and comparative genomic approaches, finding that
those involved with translation exhibit relatively low rates of gene loss and horizontal
exchange.  DNA methyltransferases, by contrast, undergo frequent horizontal exchange and are
not vertically inherited for long periods of time.  We found the model organism Escherichia
coli K-12 encodes a total of 48 known and 14 probable methyltransferases, and we mined the
literature to identify methylation activities for which the enzyme responsible was unknown.
Careful comparison of these two data sets enabled predictions of the activities of many of the
uncharacterized methyltransferases. We validated one of these predictions by mass
spectrometry, showing that the yliG gene product (renamed RimO) modifies a universally
conserved aspartic acid residue in ribosomal protein S12.  This modification (-SCH3 addition)
involves a radical-SAM mediated sulfur insertion as well as a methylation reaction.  RimO thus
belongs to a unique methyltransferase subfamily called methylthiotransferases (MTTases), and
we showed phylogenetically that MTTases comprise four subclades, of which RimO orthologs form
one.  We predicted substrates for the two remaining uncharacterized subclades and validated
one of these predictions, showing that YqeV of B. subtilis methylthiolates specific tRNA
adenosine residues. Despite acting on different substrates (protein vs. tRNA), all found
MTTase subclades share a surprisingly high degree of sequence similarity in the predicted
substrate-binding region.  We used this similarity to computationally identify candidate
residues involved with substrate specificity.  These proteins exhibit a novel protein fold
compared to other methyltransferases, and we present preliminary biochemical results that shed
light on the nature of the methylation reaction carried out by RimO.

<>

<1>Anton, B.P., Heiter, D.F., Benner, J.S., Hess, E.J., Greenough, L., Moran, L.S., Slatko, B.E., Brooks, J.E.
<2>Cloning and characterization of the BglII restriction-modification system reveals a possible evolutionary footprint.
<3>Gene
<4>187
<5>19-27
<6>1997
<7>BglII, a type II restriction-modification (R-M) system from Bacillus globigii, recognizes the
sequence 5'-AGATCT-3'.  The system has been cloned into E. coli in multiple steps: first the
methyltransferase (Mtase) gene, bglIIM, was cloned from B. globigii RUB561, a variant
containing an inactivated endonuclease (Enase) gene (bglIIR).  Next the ENase protein (R.
BglII) was purified to  homogeneity from RUB562, a strain expressing the complete R-M system.
Oligonucleotide probes specific for the 5' end of the gene were then synthesized and used to
locate bglIIR, and the gene was isolated and cloned in a subsequent step.  The nucleotide
sequence of the system has been determined, and several interesting features have been found.
The genes are tandemly arranged, with bglIIR preceding bglIIM.  The amino acid sequence of
M.BglII is compared to those of other known MTases.  A third gene encoding a protein with
sequence similarity to known C elements of other R-M sysems is found upstream of bglIIR.  This
is the first instance of a C gene being associated with an R-M system where the R and M genes
are collinear.  In addition, open reading frames (ORFs) resembling genes involved with DNA
mobility are found in close association with BglII.  These may shed light on the evolution of
the R-M system.

<>

<1>Anton, B.P., Raleigh, E.A.
<2>Transposon-mediated linker insertion scanning mutagenesis of the Escherichia coli McrA endonuclease.
<3>J. Bacteriol.
<4>186
<5>5699-5707
<6>2004
<7>McrA is one of three functions that restrict modified foreign DNA in Escherichia coli K-12,
affecting both methylated and hydroxymethylated
substrates. We present here the first systematic analysis of the
functional organization of McrA by using the GPS-LS insertion scanning
system. We collected in-frame insertions of five amino acids at 46
independent locations and C-terminal truncations at 20 independent
locations in the McrA protein. Each mutant was assayed for in vivo
restriction of both methylated and hydroxymethylated bacteriophage
(M.HpaII-modified lambda and T4gt, respectively) and for induction of the
E. coli SOS response in the presence of M.HpaII methylation, indicative of
DNA damage. Our findings suggest the presence of an N-terminal DNA-binding
domain and a C-terminal catalytic nuclease domain connected by a linker
region largely tolerant of amino acid insertions. DNA damage inflicted by
a functional C-terminal domain is required for restriction of phage T4gt.
Disruption of the N-terminal domain abolishes restriction of both
substrates. Surprisingly, truncation mutations that spare the N-terminal
domain do not mediate DNA damage, as measured by SOS induction, but
nevertheless partially restrict M.HpaII-modified lambda in vivo. We
suggest a common explanation for this "restriction without damage" and a
similar observation seen in vivo with McrB, a component of another of the
modified-DNA restriction functions. Briefly, we propose that unproductive
site-specific binding of the protein to a vulnerable position in the
lambda genome disrupts the phage development program at an early stage. We
also identified a single mutant, carrying an insertion in the N-terminal
domain, which could fully restrict lambda but did not restrict T4gt at
all. This mutant may have a selective impairment in substrate recognition,
distinguishing methylated from hydroxymethylated substrates. The study
shows that the technically easy insertion scanning method can provide a
rich source of functional information when coupled with effective
phenotype tests.

<>

<1>Anton, B.P., Raleigh, E.A.
<2>McrA variants and uses thereof.
<3>US Patent Office
<4>US 7833769 B
<5>
<6>2010
<7>Compositions and methods are provided in which the composition is a protein with at least 50%
but less than 100% amino acid sequence identity with McrA or is a variant McrA protein with at
least one amino acid sequence modification.  The variant or protein has the property of
cleaving DNA with methylated cytosine and not hydroxymethylated cytosine in a target DNA
sequence, or substantially lacks catalytic activity while maintaining binding activity.
Methods are provided in which the protein or McrA variant are used to identify methylation
sites either by cleavage or by binding to the methylation site in the presence of a marker or
by binding to an immobilized protein or McrA variant.

<>

<1>Antonenko, V., Pawlow, V., Heesemann, J., Rakin, A.
<2>Characterization of a novel unique restriction-modification system from Yersinia enterocolitica O:8 1B.
<3>FEMS Microbiol. Lett.
<4>219
<5>249-252
<6>2003
<7>Genetic manipulations with enteropathogenic Yersinia enterocolitica O:8 are complicated by the
presence of an efficient PstI-like YenI
restriction-modification (R-M) system. We have characterized the YenI R-M
system in Y. enterocolitica O:8, biotype 1B. A 5039 bp DNA fragment of the
pSAK2 recombinant plasmid carrying the yenI locus was used to determine
the nucleotide sequence. DNA sequence analysis identified a single 2481 bp
open reading frame (ORF) that encodes an 826 amino acid large polypeptide
having an apparent molecular mass of 93 kDa. The N-terminal part of the
YenI ORF has 45 and 40% identity to PstI and BsuI methyltransferases
(MTases), respectively; while the C-terminal part depicts 55 and 45%
identity to endonucleases (ENases) of both isoschyzomeric enzymes. The
yenI gene was cloned into pT7-5 plasmid and has been shown to encode a
single polypeptide of expected molecular mass. A specific recognition
sequence, typical to the type II R-M systems and single peptide
organization, typical to type IV R-M systems, make YenI unique among known
restriction-modification systems. We have constructed a truncated
recombinant variant of YenI enzyme, which conserved only MTase activity,
and that can be applied to YenI methylation of the DNA to be transformed
into Y. enterocolitica O:8 biotype 1B strains.

<>

<1>Antonopoulos, D.A., Nelson, K.E., Morrison, M., White, B.A.
<2>Strain-specific genomic regions of Ruminococcus flavefaciens FD-1 as revealed by combinatorial random-phase genome sequencing and suppressive subtractive hybridization.
<3>Environ. Microbiol.
<4>6
<5>335-346
<6>2004
<7>Two closely related strains of the Gram-positive, cellulolytic ruminal
bacterium Ruminococcus flavefaciens were compared at the genomic level by
suppressive subtractive hybridization. The two strains investigated in
this study differ by 1.94% in their respective 16S rDNA genes. Three
hundred and eighty-four PCR-amplified products were cloned and then
screened for their strain identity by dot blot hybridization. Based on
redundancy percentages of the clones sequenced, 9.5% of the genome of the
R. flavefaciens FD-1 strain is not present in the JM1 strain. The majority
of identities of individual cloned subtracted products (642 bp average
length) bore no relation to deposited sequences in GenBank (42% of the
subtracted library), whereas of those with putative assigned functions 7%
are loosely associated with fibre-degradation, 6% with insertion elements,
transposons and phage-like ORFs, 5% with cell membrane associated proteins
and 3% with signal transduction. Subtracted sequences were then
supplemented with the draft (2 x coverage) genome sequence of R.
flavefaciens FD-1 to indicate potential regions of rearrangement within
the genome, including a novel insertion sequence.

<>

<1>Antunes, A., Alam, I., Bajic, V.B., Stingl, U.
<2>Genome sequence of Salinisphaera shabanensis, a gammaproteobacterium from the harsh, variable environment of the brine-seawater interface of the  Shaban Deep in the Red Sea.
<3>J. Bacteriol.
<4>193
<5>4555-4556
<6>2011
<7>We present the genome of Salinisphaera shabanensis isolated from a brine-seawater interface
and representing a new order within the
Gammaproteobacteria. Adaptations to physicochemical and nutrient
availability fluctuation include six genes encoding heavy metal
translocating P-type ATPases, multiple genes involved in iron-uptake,
siderophore production and poly-beta-hydroxybutyrate synthesis.

<>

<1>Antunes, A., Alam, I., Bajic, V.B., Stingl, U.
<2>Genome sequence of Halorhabdus tiamatea, the first archaeon isolated from a deep-sea anoxic brine lake.
<3>J. Bacteriol.
<4>193
<5>4553-4554
<6>2011
<7>We present the draft genome of Halorhabdus tiamatea, the first member of the Archaea ever
isolated from a deep-sea anoxic brine. Genome comparison
with H. utahensis, revealed some striking differences, including marked
increase in genes associated with trans-membrane transport, and putative
genes for a trehalose synthase and a lactate-dehydrogenase.

<>

<1>Antunes, A., Alam, I., El Dorry, H., Siam, R., Robertson, A., Bajic, V.B., Stingl, U.
<2>Genome sequence of Haloplasma contractile, an unusual contractile bacterium from a deep-sea anoxic brine lake.
<3>J. Bacteriol.
<4>193
<5>4551-4552
<6>2011
<7>We present the draft genome of Haloplasma contractile, isolated from a deep-sea brine and
representing a new order between Firmicutes and
Mollicutes. Its complex morphology with contractile protrusions might be
strongly influenced by the presence of seven MreB/Mbl homologs, which
appear to be the highest copy number ever reported.

<>

<1>Antwerpen, M., Georgi, E., Zimmermann, P., Hoermansdorfer, S., Meyer, H., Grass, G.
<2>Draft Genome Sequence of Strain BF-4, a Lysinibacillus-Like Bacillus Isolated during an Anthrax Outbreak in Bavaria.
<3>Genome Announcements
<4>2
<5>e00918-14
<6>2014
<7>We report the draft genome sequence of Lysinibacillus sp. strain BF-4. Strain BF-4 has a
notably small genome for a free-living bacillus, with a size of 2.63
Mbp. In agreement with phenotypic observations, the genome lacks genes essential
for endospore formation.

<>

<1>Antwerpen, M., Proenca, D.N., Ruckert, C., Licht, K., Kalinowski, J., Hanczaruk, M., Tiemann, C., Grass, G.
<2>Draft Genome Sequence of Bacillus anthracis BF-1, Isolated from Bavarian Cattle.
<3>J. Bacteriol.
<4>194
<5>6360-6361
<6>2012
<7>Bacillus anthracis BF-1 was isolated from a cow in Bavaria (Germany) that had succumbed to
anthrax. Here, we report the draft genome sequence of this strain,
which belongs to the European B2 subclade of B. anthracis. The closest
phylogenetic neighbor of strain BF-1 is a strain isolated from cattle in France.

<>

<1>Antwerpen, M.H., Schacht, E., Kaysser, P., Splettstoesser, W.D.
<2>Complete Genome Sequence of a Francisella tularensis subsp. holarctica Strain from Germany Causing Lethal Infection in Common Marmosets.
<3>Genome Announcements
<4>1
<5>e00135-12
<6>2013
<7>Here, we describe the genome sequence of the Francisella tularensis subsp. holarctica strain
F92, belonging to the Franco-Iberian subgroup. This strain
represents the first-time isolate of this subgroup in Germany and was obtained
from naturally infected marmosets.

<>

<1>Anvar, S.Y., Frank, J., Pol, A., Schmitz, A., Kraaijeveld, K., den Dunnen, J.T., Op den Camp, H.J.
<2>The genomic landscape of the verrucomicrobial methanotroph Methylacidiphilum fumariolicum SolV.
<3>BMC Genomics
<4>15
<5>914
<6>2014
<7>Background: Aerobic methanotrophs can grow in hostile volcanic environments and use methane as
their sole source of energy. The discovery of three verrucomicrobial Methylacidiphilum strains
has revealed diverse metabolic pathways used by these methanotrophs, including mechanisms
through which methane is oxidized. The basis of a complete understanding of these processes
and of how these bacteria evolved and are able to thrive in such extreme environments
partially resides in the complete characterization of their genome and its
architecture.Results: In this study, we present the complete genome sequence of
Methylacidiphilum fumariolicum SolV, obtained using Pacific Biosciences single-molecule
real-time (SMRT) sequencing technology. The genome assembles to a single 2.5 Mbp chromosome
with an average GC content of 41.5%. The genome contains 2,741 annotated genes and 314
functional subsystems including all key metabolic pathways that are associated with
Methylacidiphilum strains, including the CBB pathway for CO2 fixation. However, it does not
encode the serine cycle and ribulose monophosphate pathways for carbon fixation. Phylogenetic
analysis of the particulate methane mono-oxygenase operon separates the Methylacidiphilum
strains from other verrucomicrobial methanotrophs. RNA-Seq analysis of cell cultures growing
in three different conditions revealed the deregulation of two out of three pmoCAB operons. In
addition, genes involved in nitrogen fixation were upregulated in cell cultures growing in
nitrogen fixing conditions, indicating the presence of active nitrogenase. Characterization of
the global methylation state of M. fumariolicum SolV revealed methylation of adenines and
cytosines mainly in the coding regions of the genome. Methylation of adenines was
predominantly associated with 5'-(m6)ACN(4)GT-3' and 5'-CC(m6)AN(5)CTC-3'
methyltransferase recognition motifs whereas methylated cytosines were not associated with any
specific motif.Conclusions: Our findings provide novel insights into the global methylation
state of verrucomicrobial methanotroph M. fumariolicum SolV. However, partial conservation of
methyltransferases between M. fumariolicum SolV and M. infernorum V4 indicates potential
differences in the global methylation state of Methylacidiphilum strains. Unravelling the M.
fumariolicum SolV genome and its epigenetic regulation allow for robust characterization of
biological processes that are involved in oxidizing methane. In turn, they offer a better
understanding of the evolution, the underlying physiological and ecological properties of SolV
and other Methylacidiphilum strains.

<>

<1>Aoki, A., Suetake, I., Miyagawa, J., Fujio, T., Chijiwa, T., Sasaki, H., Tajima, S.
<2>Enzymatic properties of de novo-type mouse DNA (cytosine-5) methyltransferases.
<3>Nucleic Acids Res.
<4>29
<5>3506-3512
<6>2001
<7>We have purified GST-fused recombinant mouse Dnmt3a and three isoforms of mouse Dnmt3b to near
homogeneity. Dnmt3b3, an isoform of Dnmt3b, did not have DNA methylation activity.  Dnmt3a,
Dnmt3b1 or Dnmt3b2 showed similar activity toward poly(dG-dC)-poly(dG-dC) for measuring de
novo methylation activity, and toward poly(dI-dC)-poly(dI-dC) for measuring total activity.
This indicates that the enzymes are de novo-type DNA methyltransferases. The enzyme activity
was inhibited by NaCl or KCl at concentrations >100 mM. The kinetic parameter, K(m)(AdoMet),
for Dnmt3a, Dnmt3b1 and Dnmt3b2 was 0.4, 1.2 and 0.9 microM when poly(dI-dC)-poly(dI-dC) was
used, and 0.3, 1.2 and 0.8 microM when poly(dG-dC)-poly(dG-dC) was used, respectively. The
K(m)(DNA) values for Dnmt3a, Dnmt3b1 and Dnmt3b2 were 2.7, 1.3 and 1.5 microM when
poly(dI-dC)-poly(dI-dC) was used, and 3.5, 1.0 and 0.9 microM when poly(dG-dC)-poly(dG-dC) was
used, respectively. For the methylation specificity, Dnmt3a significantly methylated CpG >>
CpA. On the other hand, Dnmt3b1 methylated CpG > CpT >/= CpA. Immuno-purified Dnmt3a,
Myc-tagged and overexpressed in HEK 293T cells, methylated CpG >> CpA > CpT. Neither Dnmt3a
nor Dnmt3b1 methylated the first cytosine of CpC.

<>

<1>Aonuma, M.
<2>DNA methyltransferase assay with immobilization of a ligand bound DNA substrate to a receptor bound solid support.
<3>Japanese Patent Office
<4>JP 2000232889 A
<5>8
<6>2000
<7>
<>

<1>Aparicio, J.F., Barbes, C., Hardisson, C., Sanchez, J.
<2>Sensitivity to phages of Streptomyces coelicolor strains harbouring type II restriction endonucleases.
<3>Microbiol. Sem.
<4>6
<5>71-75
<6>1990
<7>The role of type II restriction endonucleases in phage development in two
different strains of Streptomyces coelicolor has been analyzed.  Two of ten
phages tested (PhiA4 and R4c1) presented a low efficiency of plating (e.o.p.)
in the studied strains.  The isolation of host-range mutants of PhiA4 and R4c1,
with improved e.o.p. and higher adsorption capability in these two bacterial
strains, suggests that the presence of host endonucleases is not the main
barrier for these phages, but rather adsorption inability.

<>

<1>Arahal, D.R., Shao, Z., Lai, Q., Pujalte, M.J.
<2>Draft Genome Sequence of Actibacterium mucosum KCTC 23349, a Marine Alphaproteobacterium with Complex Ionic Requirements Isolated from Mediterranean   Seawater at Malvarrosa Beach, Valencia, Spain.
<3>Genome Announcements
<4>2
<5>e00486-14
<6>2014
<7>Strain R46 (CECT 7668; KCTC 23349), a nomenclatural type of Actibacterium mucosum, was
isolated from surface seawater collected at Malvarrosa Beach
(Valencia, Spain) in July 2008. The draft genome sequence of strain R46
(approximately 3.72 Mbp) contains 22 scaffolds and 3,619 protein-encoding genes, with a G+C
content of 60.8 mol%.

<>

<1>Arai, H., Kanbe, H., Ishii, M., Igarashi, Y.
<2>Complete genome sequence of the thermophilic, obligately chemolithoautotrophic hydrogen-oxidizing bacterium Hydrogenobacter  thermophilus TK-6.
<3>J. Bacteriol.
<4>192
<5>2651-2652
<6>2010
<7>Hydrogenobacter thermophilus is a thermophilic, obligately chemolithoautotrophic and aerobic
hydrogen-oxidizing bacterium. It is
unique in its ability to fix carbon dioxide via the reductive
tricarboxylic acid cycle under aerobic conditions. It utilizes molecular
hydrogen, elemental sulfur, or thiosulfate as the sole energy source.
Here, we report the complete genome sequence of H. thermophilus TK-6.

<>

<1>Arai, T., Aoki, T.
<2>New R plasmid-mediated restriction-modification system of deoxyribonucleic acid conferred by group E R plasmids.
<3>J. Bacteriol.
<4>130
<5>529-531
<6>1977
<7>A new R plasmid-mediated restriction-modification system of deoxyribonucleic
acid was identified.  This system is specific for group E plasmids which have
been detected in unidentified marine Vibrio fish pathogens.

<>

<1>Aranda, J., Roca, M., Lopez-Canut, V., Tunon, I.
<2>Theoretical Study of the Catalytic Mechanism of DNA-(N4-Cytosine)-Methyltransferase from the Bacterium Proteus vulgaris.
<3>J. Phys. Chem. B
<4>114
<5>8467-8473
<6>2010
<7>In this paper the reaction mechanism for methylation of cytosine at the exocyclic N4 position
catalyzed by M.PvuII has been explored by means of hybrid quantum mechanics/molecular
mechanics (QM/MM) methods.   A reaction model was prepared by placing a single cytosine base
in the active site of the enzyme. In this model the exocyclic amino group of the base
establishes hydrogen bond interactions with the hydroxyl oxygen atom of Ser53 and the carbonyl
oxygen atom of Pro54. The reaction mechanism involves a direct methyl transfer from AdoMet to
the N4 atom and a proton transfer from this atom to Ser53, which in turn transfers a proton to
Asp96. Different timings for the proton transfers and methylation steps have been explored at
the AM1/MM and B3LYP/MM levels including localization and characterization of stationary
structures. At our best estimate the reaction proceeds by means of a simultaneous but
asynchronous proton transfer from Ser53 to Asp96 and from N4 of cytosine to Ser53 followed by
a direct methyl transfer from AdoMet to the exocyclic N4 of cytosine.

<>

<1>Aranda, J., Roca, M., Tunon, I.
<2>Substrate promiscuity in DNA methyltransferase M.PvuII. A mechanistic insight.
<3>Org. Biomol. Chem.
<4>10
<5>5395-5400
<6>2012
<7>M. PvuII is a DNA methyltransferase from the bacterium Proteus vulgaris that catalyzes
methylation of cytosine at the N4 position. This enzyme
also displays promiscuous activity catalyzing methylation of adenine at
the N6 position. In this work we use QM/MM methods to investigate the
reaction mechanism of this promiscuous activity. We found that N6
methylation in M. PvuII takes place by means of a stepwise mechanism in
which deprotonation of the exocyclic amino group is followed by the
methyl transfer. Deprotonation involves two residues of the active
site, Ser53 and Asp96, while methylation takes place directly from the
AdoMet cofactor to the target nitrogen atom. The same reaction
mechanism was described for cytosine methylation in the same enzyme,
while the reversal timing, that is methylation followed by
deprotonation, has been described in M. TaqI, an enzyme that catalyzes
the N6-adenine DNA methylation from Thermus aquaticus. These
mechanistic findings can be useful to understand the evolutionary paths
followed by N-methyltransferases.

<>

<1>Aranda, J., Zinovjev, K., Roca, M., Tunon, I.
<2>Dynamics and Reactivity in Thermus aquaticus N6-Adenine Methyltransferase.
<3>J. Am. Chem. Soc.
<4>136
<5>16227-16239
<6>2014
<7>M.TaqI is a DNA methyltransferase from Thermus aquaticus that catalyzes the transfer of a
methyl group from S-adenosyl-L-methionine to the N6 position of an adenine, a process
described only in prokaryotes. We have used full atomistic classical molecular dynamics
simulations to explore the protein-SAM-DNA ternary complex where the target adenine is flipped
out into the active site. Key protein DNA interactions established by the target adenine in
the active site are described in detail. The relaxed structure was used for a combined quantum
mechanics/molecular mechanics exploration of the reaction mechanism using the string method.
According to our free energy calculations the reaction takes place through a stepwise
mechanism where the methyl transfer precedes the abstraction of the proton from the exocyclic
amino group. The methyl transfer is the rate-determining step, and the obtained free energy
barrier is in good agreement with the value derived from the experimental rate constant. Two
possible candidates to extract the leftover proton have been explored: a water molecule found
in the active site and Asn105, a residue activated by the hydrogen bonds formed through the
amide hydrogens. The barrier for the proton abstraction is smaller when Asn105 acts as a base.
The reaction mechanisms can be different in other N6-DNA-methyltransferases, as determined
from the exploration of the reaction mechanism in the Asn105Asp M.TaqI mutant.

<>

<1>Aranda, J., Zinovjev, K., Swiderek, K., Roca, M., Tunon, I.
<2>Molecular dynamics and QM/MM free energy profiles of cytosine C5-methyltransferase M.Hhai.
<3>FEBS J.
<4>280
<5>105
<6>2013
<7>The target of this work is the study of mechanism of the reaction catalyzed by M.Hhai, an
enzyme that belongs to the restriction-modification system of the bacterium Haemophilus
haemolyticus which catalyzes the methyl transfer from S-adenosil-L-metionine to C5 position of
a cytosine base of DNA, working at the 5'-GCGC-3' sequence generating C5-methyl-cytosine.
The X-Ray structure of the enzyme complexed with SAM and a DNA sequence was the starting point
of our simulations.  We performed all calculations considering the whole enzymatic and DNA
environment and the solvation effect by including the enzyme in a orthorhombic box of TIP3P
water molecules of 88 x 87 x 99 A of side, including sodium counterions to neutralize the
charge of the system.  Using the NAMD program we equilibrated the system by means of 10 ns of
classical molecular dynamics with the AMBER force field, employing periodic boundary
conditions, Ewald summations, a temperature of 300 K and a time step of 1 fs.  We then
performed 100 ns MD simulaton in order to analyze the most important interactions formed
between the enzyme and DNA, the interactions within the active site, how the unpaired base is
stabilized and how the DNA helix accommodates the great perturbation that a flipped out base
means for its structure.  To analyse the chemical reaction we performed 500 ps of quantum
mechanics/molecular mechanics (QM/MM) MD simulation at 300 K using Dynamo program.  Quantum
subsystem was treated using the AM1 semiempirical hamiltonian adding corrections at the
M062x/6-311 + G level.  By means of the on-the-fly string method the minimum free energy path
for each step of the reaction was obtained.  Then, the path collective variable was defined
along these paths, to obtain the potential of mean force using umbrella sampling.

<>

<1>Aras, R.A.
<2>Helicobacter pylori regulates hpyII restriction-modification function using gene deletion and horizontal reacquisition.
<3>Int. J. Med. Microbiol.
<4>291S
<5>95
<6>2001
<7>Helicobacter pylori possess strain-specific complements of functional restriction-modification
systems.  The large number of R-M's homologous to those in other bacterial species and their
strain-specificity suggests that H. pylori may have horizontally acquired these genes.  A type
IIs R-M system, HpyII, was active in 2 of the 6 H. pylori strains studied.  We now demonstrate
that in most strains lacking HpyII.M function there is complete absence of the R-M system.
Direct DNA repeats of 80-bp flanking the hpyII R-M system allow its deletion, resulting in an
"empty-site" genotype.  We show that strains possessing this empty-site genotype and strains
with a full but inactive hpyII R-M can reacquire the hpyII R-M cassette and functional
activity through natural transformation by DNA from the parental R+M+ strain.  Identical
isolates divergent for the presence of an active HpyII R-M pose different restriction barriers
to transformation by foreign DNA.  That H. pylori can regulate HpyII R-M function through
deletion or mutation, and subsequent horizontal reacquisition of the hpyII R-M cassette
providing an example of a novel mechanism for R-M regulation, supports the hypothesis that H.
pylori populations use mutation and transformation to regulate gene function.

<>

<1>Aras, R.A., Blaser, M.J.
<2>Conservation of a type IIs restriction-modification system in Helicobacter pylori with homology to the MboII R-M system in Moraxella bovis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>100
<5>290-291
<6>2000
<7>Bacteria use restriction and modification systems as a defense against invasion from foreign
DNA.  Comparison of the genomes from Helicobacter pylori strains 26695 and J99 identified a
number of strain-specific R-M systems.  Here we show the conservation of a type IIs R-M system
among 18 H. pylori strains and its possible involvement in horizontal gene transfer.  Strain
26695 has a potential type IIs R-M system that has homology to the Moraxella bovis MboII R-M
system, which recognizes the non-palindromic sequence GAAGA.  PCR analysis of 17 other strains
show that 10 contain a complete R-M system, 2 contain partial R-M systems and 5 contain no R-M
system.  Sequence analysis of the 5' end of the restriction endonuclease subunit from 7
different strains shows predominantly synonymous variations in codon usage; indicating the
locus is under substantial selective pressure.  For all strains except J188, which has a
nonfunctional RE subunit, presence of both methyltransferases (MTs) correlates with protection
of genomic DNA from MboII digestion, implying that the MTs are responsible for modification of
the sequence GAAGA.  G+C content and phylogenetic analysis indicates that the H. pylori type
IIs R-M system may have been acquired through horizontal gene transfer.

<>

<1>Aras, R.A., Small, A.J., Ando, T., Blaser, M.J.
<2>Horizontal transfer of the Helicobacter pylori restriction-modification system, HpyII, by a conjugation-like mechanism.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>102
<5>165-166
<6>2002
<7>Helicobacter pylori, gram-negative, curved bacteria that colonize the human gastric mucosa,
possess a large number of genes for
restriction-modification (R-M) systems, and essentially every strain
possesses a unique complement of functional and partial R-M systems.
Nearly half of the H. pylori strains studied possess an active, type
IIs R-M system, HpyII, with the recognition sequence GAAGA.
Recombination between direct repeats that flank the R-M allows for its
deletion whereas strains lacking hpyIIRM can acquire this cassette
through natural transformation. We now asked whether strains lacking
HpyII R-M activity can acquire an active 4.8 kb hpyIIRM cassette
(containing a kanamycin resistance (aphA) marker) through a
DNase-resistant mechanism. We found that hpyIIRM strains 6c and J166
acquired an hpyIIRM cassette from strain 6a, through a DNase-resistant
mechanism, if strains are isogenic (6a fwdarw 6c;
frequency=3.5e-7+-4.0e-7), but not if strains are non-isogenic (6a
fwdarw J166; frequency<1.8e-8). The frequency of conjugation-like
transfer of a point mutation conferring streptomycin resistance was not
significantly (p=0.47) different for isogenic
(frequency=2.7e-6+-2.3e-6) and non-isogenic (frequency=1.6e-6+-4.8e-7)
strains. A non-isogenic strain, J188, containing a full but inactive
hpyIIRM reactivated its HpyII R-M through natural transformation
(frequency=2.4e-7+-1.5e-7) but not through conjugation-like
(frequency<6.6e-8), horizontal acquisition of a functional hpyIIRM.
Strain J188 was transformed with the hpyIIRM cassette at a
significantly (p<0.05) lower frequency then was the isogenic 6a strain
(frequency=7.9e-6+-2.0e-6). These data indicate that a conjugation-like
mechanism contributes to the transfer of large (4.8 kb) fragments of
chromosomal DNA between H. pylori strains and that inactive or partial
R-M systems may act as contingency genes that can be reactivated upon
recombination with a functional allele. That functional R-M systems
appear to restrict acquisition of chromosomal DNA suggests that there
is a double-stranded DNA intermediate in the H. pylori uptake process.

<>

<1>Aras, R.A., Small, A.J., Ando, T., Blaser, M.J.
<2>Helicobacter pylori interstrain restriction-modification diversity prevents genome subversion by chromosomal DNA from competing strains.
<3>Nucleic Acids Res.
<4>30
<5>5391-5397
<6>2002
<7>Helicobacter pylori, bacteria that colonize the human gastric mucosa, possess a large number
of genes for restriction-modification (R-M) systems, and essentially, every strain possesses a
unique complement of functional and partial R-M systems. Nearly half of the H.pylori strains
studied possess an active type IIs R-M system, HpyII, with the recognition sequence GAAGA.
Recombination between direct repeats that flank the R-M cassette allows for its deletion
whereas strains lacking hpyIIRM can acquire this cassette through natural transformation. We
asked whether strains lacking HpyII R-M activity can acquire an active hpyIIRM cassette
[containing a 1.4 kb kanamycin resistance (aphA) marker], whether such acquisition is DNase
sensitive or resistant and whether restriction barriers limit acquisition of chromosomal DNA.
Our results indicate that natural transformation and conjugation-like mechanisms may
contribute to the transfer of large (4.8 kb) insertions of chromosomal DNA between H.pylori
strains, that inactive or partial R-M systems can be reactivated upon recombination with a
functional allele, consistent with their being contingency genes, and that H.pylori R-M
diversity limits acquisition of chromosomal DNA fragments of 1 kb.

<>

<1>Aras, R.A., Takata, T., Ando, T., van der Ende, A., Blaser, M.J.
<2>Regulation of the HpyII restriction-modification system of Helicobacter pylori by gene deletion and horizontal reconstitution.
<3>Mol. Microbiol.
<4>42
<5>369-382
<6>2001
<7>Helicobacter pylori, Gram-negative, curved bacteria colonizing the human stomach, possess
strain-specific complements of functional
restriction-modification (R-M) systems. Restriction-modification
systems have been identified in most bacterial species studied and are
believed to have evolved to protect the host genome from invasion by
foreign DNA. The large number of R-Ms homologous to those in other
bacterial species and their strain-specificity suggest that H. pylori
may have horizontally acquired these genes. A type IIs
restriction-modification system, hpyIIRM, was active in two out of the
six H. pylori strains studied. We demonstrate now that in most strains
lacking M.HpyII function, there is complete absence of the R-M system.
Direct DNA repeats of 80 bp flanking the hpyIIRM system allow its
deletion, resulting in an 'empty-site' genotype. We show that strains
possessing this empty-site genotype and strains with a full but
inactive hpyIIRM can reacquire the hpyIIRM cassette and functional
activity through natural transformation by DNA from the parental R-M+
strain. Identical isolates divergent for the presence of an active
HpyII R-M pose different restriction barriers to transformation by
foreign DNA. That H. pylori can lose HpyII R-M function through
deletion or mutation, and can horizontally reacquire the hpyIIRM
cassette, is, in composite, a novel mechanism for R-M regulation,
supporting the general hypothesis that H. pylori populations use
mutation and transformation to regulate gene function.

<>

<1>Aras, R.A., Takata, T., van der Ende, A., Blaser, M.J.
<2>Helicobacter pylori variants from a single host that differ in the presence of the hpyII restriction-modification system.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>296
<6>2001
<7>Helicobacter pylori are gram-negative, curved bacteria that reside in the human gastric
mucosa. Sequencing of two independent H. pylori
strains, 26695 and J99, identified an extraordinary number of
restriction-modification (R-M) genes. Many are strain-specific, have
deviant G+C content, and are associated with genomic rearrangements,
suggesting their horizontal acquisition from other bacteria. Strain
26695 possesses a type IIs R-M system, hpyII R-M, with homology to the
Moraxella bovis MboII R-M system, recognizing the sequence GAAGA. Since
genomic analysis identified 80-nucleotide direct repeats flanking the
26695 R-M system that could permit deletion of intervening regions, we
searched for isolates from a single host that differ in hpyII R-M
status. The resistance of an isolate's DNA to MboII digestion was used
to indicate the presence of the hpyII R-M system. From a Dutch patient,
we found two isolates that differed. Using primers specific for
different regions of the hpyII R-M, all expected PCR products were
present for the resistant isolate (2a), but were absent for the
sensitive isolate (2b). PCR using primers that flank the repeat
sequences amplified a 3.5 kb PCR product in strain 2a (consistent with
a full hpyII R-M system), and an "empty-site" (276bp) product in strain
2b (consistent with deletion of the R-M system and one repeat);
sequencing of the "empty-site" product confirmed the deletion. That the
strains had identical RAPD and RFLP profiles and were otherwise
identical in susceptibility to 13 other restriction enzymes indicate
that they are clonal variants rather than different strains. Parallel
results were found for paired isolates obtained from one of the
patient's daughters. The isolation from a single host of clonal
variants divergent in hpyII R-M status, coupled with the sequence data
suggest deletion in vivo, findings consistent with the hypothesis of
R-M system mobility within H. pylori.

<>

<1>Araujo, F.D., Croteau, S., Slack, A.D., Milutinovic, S., Bigey, P., Price, G.B., Zannis-Hajopoulos, M., Szyf, M.
<2>The dnmt1 target recognition domain resides in the n terminus.
<3>J. Biol. Chem.
<4>276
<5>6930-6936
<6>2001
<7>DNA-cytosine-5-methyltransferase 1 (DNMT1) is the enzyme believed to be responsible for
maintaining the epigenetic information encoded by DNA methylation patterns. The target
recognition domain of DNMT1, the domain responsible for recognizing hemimethylated CGs, is
unknown. However, based on homology with bacterial cytosine DNA methyltransferases it has been
postulated that the entire catalytic domain, including the target recognition domain, is
localized to 500 amino acids at the C terminus of the protein. The N-terminal domain has been
postulated to have a regulatory role, and it has been suggested that the mammalian DNMT1 is a
fusion of a prokaryotic methyltransferase and a mammalian DNA-binding protein. Using a
combination of in vitro translation of different DNMT1 deletion mutant peptides and a
solid-state hemimethylated substrate, we show that the target recognition domain of DNMT1
resides in the N terminus (amino acids 122-417) in proximity to the proliferating cell nuclear
antigen binding site. Hemimethylated CGs were not recognized specifically by the postulated
catalytic domain. We have previously shown that the hemimethylated substrates utilized here
act as DNMT1 antagonists and inhibit DNA replication. Our results now indicate that the
DNMT1-PCNA interaction can be disrupted by substrate binding to the DNMT1 N terminus. These
results point toward new directions in our understanding of the structure-function of DNMT1.

<>

<1>Araujo, F.D., Knox, J.D., Ramchandrani, S., Pelletier, R., Bigey, P., Price, G., Szyf, M., Zannis-Hadjopoulos, M.
<2>Identification of initiation sites for DNA replication in the human dnmt1 (DNA-methyltransferase) locus.
<3>J. Biol. Chem.
<4>274
<5>9335-9341
<6>1999
<7>Vertebrates have developed multiple mechanisms to coordinate the replication of epigenetic and
genetic information.  Dnmt1 encodes the maintenance enzyme DNA-methyltransferase, which is
responsible for propagating the DNA methylation pattern and the epigenetic information that it
encodes during replication.  Direct sequence analysis and bisulfite mapping of the 5' region
of DNA-methyltransferase 1 have indicated the presence of many sequence elements associated
with previously characterized origins of DNA replication.  This study tests the hypothesis
that the dnmt1 region containing these elements is an origin of replication in human cells.
First, we demonstrate that a vector containing this dnmt1 sequence is able to support
autonomous replication when transfected into HeLa cells.  Second, using a gel retardation
assay, we show that it contains a site for binding of origin-rich sequences binding activity,
a recently purified replication protein.  Finally, using competitive polymerase chain
reaction, we show that replication initiates in this region in vivo.  Based on these lines of
evidence, we propose that initiation sites for DNA replication are located between the first
intron and exon 7 of the human dnmt1 locus.

<>

<1>Aravind, L., Makarova, K.S., Koonin, E.V.
<2>Holliday junction resolvases and related nucleases: identification of new families, phyletic distribution and evolutionary trajectories.
<3>Nucleic Acids Res.
<4>28
<5>3417-3432
<6>2000
<7>Holliday junction resolvases (HJRs) are key enzymes of DNA recombination. A detailed computer
analysis of the structural and evolutionary relationships of HJRs and related nucleases
suggests that the HJR function has evolved independently from at least four distinct
structural folds, namely RNase H, endonuclease, endonuclease VII-colicin E and RusA. The
endonuclease fold, whose structural prototypes are the phage lambda exonuclease, the very
short patch repair nuclease (Vsr) and type II restriction enzymes, is shown to encompass by
far a greater diversity of nucleases than previously suspected. This fold unifies archaeal
HJRs, repair nucleases such as RecB and Vsr, restriction enzymes and a variety of predicted
nucleases whose specific activities remain to be determined. Within the RNase H fold a new
family of predicted HJRs, which is nearly ubiquitous in bacteria, was discovered, in addition
to the previously characterized RuvC family. The proteins of this family, typified by
Escherichia coli YqgF, are likely to function as an alternative to RuvC in most bacteria, but
could be the principal HJRs in low-GC Gram-positive bacteria and Aquifex. Endonuclease VII of
phage T4 is shown to serve as a structural template for many nucleases, including McrA and
other type II restriction enzymes. Together with colicin E7, endonuclease VII defines a
distinct metal-dependent nuclease fold. As a result of this analysis, the principal HJRs are
now known or confidently predicted for all bacteria and archaea whose genomes have been
completely sequenced, with many species encoding multiple potential HJRs. Horizontal gene
transfer, lineage-specific gene loss and gene family expansion, and non-orthologous gene
displacement seem to have been major forces in the evolution of HJRs and related nucleases. A
remarkable case of displacement is seen in the Lyme disease spirochete Borrelia burgdorferi,
which does not possess any of the typical HJRs, but instead encodes, in its chromosome and
each of the linear plasmids, members of the lambda exonuclease family predicted to function as
HJRs. The diversity of HJRs and related nucleases in bacteria and archaea contrasts with their
near absence in eukaryotes. The few detected eukaryotic representatives of the endonuclease
fold and the RNase H fold have probably been acquired from bacteria via horizontal gene
transfer. The identity of the principal HJR(s) involved in recombination in eukaryotes remains
uncertain; this function could be performed by topoisomerase IB or by a novel, so far
undetected, class of enzymes. Likely HJRs and related nucleases were identified in the genomes
of numerous bacterial and eukaryotic DNA viruses. Gene flow between viral and cellular genomes
has probably played a major role in the evolution of this class of enzymes. This analysis
resulted in the prediction of numerous previously unnoticed nucleases, some of which are
likely to be new restriction enzymes.

<>

<1>Arber, W.
<2>DNA Modification and Restriction.
<3>Prog. Nucleic Acid Res. Mol. Biol.
<4>14
<5>1-37
<6>1974
<7>The reader of the scientific literature may have had his attention attracted
recently to a growing number of highly interesting reports on research on DNA
restriction endonucleases and DNA modification methylases.  A striking
illustration is the November 1972 issue of the Proceedings of the National
Academy of Sciences of the United States of America:  it contains seven papers
on restriction endonucleases, and none of the 16 authors signed more than one
of these papers.

<>

<1>Arber, W.
<2>Host-controlled modification of bacteriophage.
<3>Annu. Rev. Microbiol.
<4>19
<5>365-378
<6>1965
<7>Host-controlled modification of viruses is a general term applied to those
cases in which passage through certain host strains imparts one or more new,
nonheritable properties to the virus without altering its genetic information
content.  The terms of host-induced modification, host-controlled variation, or
host-induced variation are sometimes used as synonyms to designate the same
phenomena.  However, we would like to recommend the use of "host-controlled
modification," at least for the cases discussed in this paper, since control
mechanisms are involved here rather than induction phenomena and since the term
"variation" may erroneously suggest some change in the genetic message.

<>

<1>Arber, W.
<2>Strain-specific restriction and modification of DNA.
<3>Bull. Schweiz. Akad. Med. Wiss.
<4>25
<5>99-100
<6>1970
<7>Experimental investigation of the molecular mechanism of host-controlled
modification of bacteriophages has led to the discovery of very interesting
bacterial enzyme systems.  Each of these systems has a specific restriction
enzyme (endonuclease) and a specific modification enzyme (DNA methylase).  Both
enzymes appear to work with high affinity on the same position of DNA.  This
point of affinity consists of a quite definite sequence of 6 to 9 DNA base
pairs.  The recognition of the point of affinity by the enzymes is taken over
by a gene product as the common factor for both enzyme activities.  The genetic
basis of the activities, the enzymes themselves and the substrate can be
experimentally investigated and offer the molecular biologist an interesting
model of the specific interaction of enzymes with nucleic acids.  The
biological significance of DNA restriction lies in an effective defence
mechanism against infection with foreign genetic material.  For a detailed
account of the present knownledge on this subject, the reader is referred to a
review by Arber and Linn (Ann. Rev. Biochem. 38, 467[1969].

<>

<1>Arber, W.
<2>Host-controlled modification of DNA.
<3>Naturwissenschaften
<4>56
<5>155-160
<6>1969
<7>None

<>

<1>Arber, W.
<2>Promotion and limitation of genetic exchange.
<3>Science
<4>205
<5>361-365
<6>1979
<7>None

<>

<1>Arber, W.
<2>Specificites biologiques de l'acide desoxyribonucleique.
<3>Pathol. Microbiol. (Basel)
<4>25
<5>668-681
<6>1962
<7>A new biological specificity of phage DNA molecules is described, which is
distinct from the genetic specificity of DNA residing in the base sequence.
Unlike the genetic specificity this newly discovered "host specificity" is not
multiplied as DNA replicates, but is nevertheless shown to be fixed firmly to
the DNA molecule and so to be transferred into progeny molecules jointly with
the parental DNA material.  Production of DNA host specificity is governed both
by the bacterial genome and by episomes which are present in the cell.  Host
specificity apparently exists not only on phage DNA but as well on the DNA of
the host itself.  It is thought to be the element changed in "host controlled
modification" of phage and also the element recognized by restricting hosts
which degrade nonadapted, infecting DNA.

<>

<1>Arber, W.
<2>Host specificity of DNA produced by Escherichia coli V. The role of Methionine in the Production of Host Specificity.
<3>J. Mol. Biol.
<4>11
<5>247-256
<6>1965
<7>Bacteriophage lambda grown in auxotrophic met-, pro- or arg- strains of
Escherichia coli K12 in the presence of the required amino acids show an
efficiency of plating of approximately 1 on E. coli strains K12 and C.
However, if met- cells are deprived of methionine during a portion of the
latent period, the efficiency of plating of the progeny phage is lower on the
host K12 than on strain C.  Similar results are obtained with met- auxotrophs
of strains B and K12 (P1); deprivation of methionine during the latent period
results in the production of phage with lower efficiency of plating on the host
strain than on strain C.  Such an effect is not observed following a similar
starvation for proline or arginine.  These results suggest that methionine is
specifically required for the production of host specificity of DNA.

<>

<1>Arber, W.
<2>Host-controlled restriction and modification of bacteriophage.
<3>Symp. Soc. Gen. Microbiol.
<4>18
<5>295-314
<6>1968
<7>Viral properties have frequently been reported to undergo non-heritable changes
upon passage through certain host strains.  But very little was known about the
molecular mechanisms forming the basis of such host-controlled modifications,
when we started in 1960 to investigate one particular virus-host system.  This
work, carried out with bacteriophage lambda and a few host strains all derived
from Escherichia coli, soon revealed the existence of a highly specific
recognition mechanism able to screen infecting and intracellular DNA molecules
for absence or presence on these molecules of a host-specific stamp.  This has
more recently been identified as nucleotide methylation.  It is the scope of my
contribution to this symposium to lay out the crucial experiments and arguments
that lead to the present understanding of this system, which may be interpreted
as serving the cell as a defense mechanism against infection with foreign
genetic material.

<>

<1>Arber, W.
<2>Host specificity of DNA produced by Escherichia coli.  III.  Effects on transduction mediated by lambda dg.
<3>Virology
<4>23
<5>173-182
<6>1964
<7>Transducing phage lambda dg carrying bacterial gal markers shows the same
effects of host-controlled modification as does normal lambda.  Phage grown on
Escherichia coli K12 is restricted (not accepted) on E. coli B and K12(P1), and
phage grown on B is restricted on K12 and K12(P1) independently of whether the
gal markers of lambda dg originated from the K12 or B chromosome.  Transduction
to a restricting host yields a very low proportion of gal+ transductants, which
are mostly heterogenotes.  Superinfecting restricted lambda phage does not
exert helper functions for lysogenization or reproduction of nonrestricted
lambda dg particles.  In cells infected with restricted lambda dg and
superinfected with nonrestricted lambda, a greatly increased transduction
frequency is observed, probably due to marker rescue.  A small helper effect is
observed in cells infected with restricted lambda dg and restricted lambda,
presumably caused by help in lysogenization in those few cells which accept the
infecting lambda dg.  For nonrestricting hosts it is known that ultraviolet
irradiated lambda dg no longer transduces by lysogenization but by stable
integration of the gal markers into the bacterial chromosome.  The same
behavior is found for restricting recipients.  Here, the maximum number of
stable transductants is not notably higher than the number of heterogenotic
transductants obtained on the same bacteria infected with nonirradiated lambda
dg.  This is in agreement with the notion that most of the restricted genomes
are degraded after their injection.  Particles giving rise to stable
transduction show a very low UV sensitivity.  The ease with which stable
transduction occurs probably reflects the degree of homology between endo- and
exogenotic gal regions.

<>

<1>Arber, W.
<2>Host-controlled variation.
<3>Bacteriophage Lambda, Cold Spring Harbor Laboratory Press, Hershey, A.D., Cold Spring Harbor, New York
<4>0
<5>83-96
<6>1971
<7>About twenty years ago a number of similar observations were made in work with various
bacteriophage species.  They showed that the host range of a given phage preparation depended
on the bacterial strain in which the phage had last propagated.  This effect was called
host-controlled variation to distinguish its host-dependent and thus genetically unstable
nature from persistent hereditary chcanges such as are found in host-range mutants.  Many
examples now are known to reflect the phenomena described in this chapter:  strain-specific
restriction and modification of DNA.

<>

<1>Arber, W.
<2>What is the function of DNA restriction enzymes?
<3>Trends Biochem. Sci.
<4>2
<5>N176-N178
<6>1977
<7>None

<>

<1>Arber, W.
<2>On the generation of genetic diversity in microorganisms.
<3>Proc. Indian Natn. Sci. Acad.
<4>B60
<5>357-364
<6>1994
<7>Bacterial genetics strongly influenced the development of molecular genetic strategies and
techniques now available to study gene structure and functions of practically any living
organisms.  It also revealed natural processes of horizontal gene transfer (transformation,
conjugation, phage-mediated transduction) as well as systems (e.g. restriction-modification
systems) to hold such gene transfer in tolerably low frequencies to ensure a certain degree of
genetic stability.  Work with bacterial and bacteriophage systems has helped to unravel both
homologous and non-homologous enzyme-mediated recombination processes at the molecular level.
The acquired knowledge now helps to understand molecular processes contributing to the
generation of genetic variation, especially DNA rearrangements resulting from transposition
and from site-specific recombination which sometimes occurs at secondary crossing-over sites.
Present knowledge on the genetic plasticity of haploid microorganisms offers insights into
the molecular basis for the natural interplay between mutagenesis and selection.  This
approach greatly profits from the short generation times and relatively small genome sizes of
haploid microorganisms which allows one to investigate population genetic questions and to
draw conclusions on the mechanisms of evolutionary processes.

<>

<1>Arber, W.
<2>Origin and properties of type A host specificity in Escherichia coli.
<3>Pathol. Microbiol. (Basel)
<4>34
<5>147
<6>1969
<7>
<>

<1>Arber, W.
<2>Genetic variation: molecular mechanisms and impact on microbial evolution.
<3>FEMS Microbiol. Rev.
<4>24
<5>1-7
<6>2000
<7>On the basis of established knowledge of microbial genetics one can distinguish three major
natural strategies in the spontaneous
generation of genetic variations in bacteria. These strategies are: (1)
small local changes in the nucleotide sequence of the genome, (2)
intragenomic reshuffling of segments of genomic sequences and (3) the
acquisition of DNA sequences from another organism. The three general
strategies differ in the quality of their contribution to microbial
evolution. Besides a number of non-genetic factors, various specific
gene products are involved in the generation of genetic variation and
in the modulation of the fequency of genetic variation. The underlying
genes are called evolution genes. They act for the benefit of the
biological evolution of populations as opposed to the action of
housekeeping genes and accessory genes which are for the benefit of
individuals. Examples of evolution genes acting as variation generators
are found in the transposition of mobile genetic elements and in
so-called site-specific recombination systems. DNA repair systems and
restriction-modification systems are examples of modulators of the
frequency of genetic variation. The involvement of bacterial viruses
and of plasmids in DNA reshuffling and in horizontal gene transfer is a
hint for their evolutionary functions. Evolution genes are thought to
undergo biological evolution themselves, but natural selection for
their functions is indirect, at the level of populations, and is called
second-order selection. In spite of an involvement of gene products in
the generation of genetic variations, evolution genes do not
programmatically direct evolution towards a specific goal. Rather, a
steady interplay between natural selection and mixed populations of
genetic variants gives microbial evolution its direction.

<>

<1>Arber, W.
<2>Roots, strategies and prospects of functional genomics - functional genomics, human genome, bioinformatic software and polymerase chain  reaction; a review.
<3>Curr. Sci.
<4>83
<5>826-828
<6>2002
<7>AUTHOR ABSTRACT - This essay traces,the historical development of classical and molecular
genetics from their early roots to the actual
research strategies and their prospects. Attention is also given to the
risk evaluation of genetic engineering by comparing designed genetic
alterations with the spontaneous genetic variation known to form the
substrate for biological evolution. DERWENT ABSTRACT: Roots, strategies
and prospects of functional genomics was discussed with respect to
historical development of classical molecular genetics from their early
roots to the actual research strategies and their prospects. Classical
genetics has its roots in the 19th century, when Gregor Mendel carried
out experiments with peas displaying phenotypically distinct traits
which got inherited to the progeny. However, efficient methods to
experimentally determine large extents of such sequences were still
missing. In the 1950s and 1960s it also became clear that bacteria
succeed by a number of different strategies to hold the frequency of
acquisition of foreign genetic information low. One of these strategies
is the widely encountered phenomenon of restriction and modification of
DNA. Restriction-modification systems allow bacteria to specifically
distinguish foreign DNA from the cell's own DNA. As a consequence,
foreign DNA becomes cleaved into fragments upon its entry into the
cell. In 1970 investigators succeeded in preparing recombinant DNA
molecules in vitro and to get them replicated after their transfer into
appropriate host cells. In these experiments plasmids and viral genomes
served as gene vectors into which fragments of DNA, often prepared by
restriction cleavage, were spliced. In 1990s, the gateway to the
determination of the nucleotide sequences of entire genomes was thus
open. More recently, the DNA sequences of several eukaryotic organisms
were published, including the human genome of about three billion base
pairs(3 pages)

<>

<1>Arber, W.
<2>Genetic Variation and Molecular Evolution.
<3>Encyclopedia of Molecular Cell Biology and Molecular Medicine., Wiley-VCH Verlag, Meyers, R.A., 
<4>5
<5>331-352
<6>2004
<7>The comparison of DNA sequences of genes and entire genomes offers interesting insights into
the possible evolutionary relatedness of genetic information of living organisms.  Together
with a relatively rich database from experimental microbial genetics, conclusions can be drawn
on the molecular mechanisms by which genetic variations are spontaneously generated.  A number
of different specific mechanisms contribute to the overall mutagenesis.  These mechanisms are
here grouped into three natural strategies of the spontaneous generation of genetic
variations: local changes of DNA sequences, intragenomic rearrangement of DNA segments, and
acquisition of foreign DNA by horizontal gene transfer.  These three strategies have different
qualities with regard to their contributions to the evolutionary process.  As a general rule,
none of the known mechanisms producing genetic variants is clearly directed.  Rather, the
resulting alterations in the inherited genomes are more random.  In addition, usually only a
minority of resulting variants provide a selective advantage.  Interestingly, in most of the
molecular mechanisms involved, the products of so-called evolution genes are involved as
generators of genetic variation and/or as modulators of the frequencies of genetic variation.
Products of evolution genes work in tight collaboration with nongenetic factors such as
structural flexibilities and chemical instabilities of molecules, chemical and physical
mutagens, and random encounter.  All of these aspects contributing to the spontaneous
generation of genetic variations together form the core of the theory of molecular evolution.
This theory brings neo-Darwinism to the molecular level.  In view of the increasing evidence
coming particularly from microbial genetics, knowledge of molecular evolution can be seen as a
confirmation of Darwinism at the level of biologically active molecules, in particular,
nucleic acids and proteins.  Philosophical and practical implications of this knowledge will
be briefly discussed.

<>

<1>Arber, W.
<2>Host specificity of DNA produced by Escherichia coli.   9. Host-controlled modification of bacteriophage fd.
<3>J. Mol. Biol.
<4>20
<5>483-496
<6>1966
<7>Host-controlled modification is shown to occur with four related male-specific
bacteriophage strains containing single-stranded DNA: fd,f1,M13 and F12.  All
four phages are restricted and modified in bacteria with B host specificity,
the first three also in P1-lysogenic cells.  None of the phages is restricted
in strains with K host specificity or carrying the episome RTF-2.  The
bacterial characters rB mB which control the B host specificity of lambda DNA,
are also responsible for restriction and modification of phage fd.  The
apparent difference in K restriction, which is encountered by lambda, but not
by fd, is thought to find its explanation in the small molecular size of fd
DNA, on which K specificity sites might be lacking.  Indeed, restriction and
modification act on the DNA of fd:  DNA from fd phages which infect restricting
host cells is partially broken down to acid-soluble products.  On the other
hand, one-cycle growth of fd.B on non-restricting and non-modifying Kr-m-
bacteria yields, among a majority of progeny of fd.Kr-m- phage, some phage
particles with parental B host specificity, and they also have parental DNA as
shown by density labelling of the infecting phage.  The efficiency of such
transfer of parental fd.B DNA was found to be 0.12 if measured after 18 minutes
incubation of the infected cells.  The implication of this transfer on the
mechanism of phage DNA replication is discussed.

<>

<1>Arber, W.
<2>Restriction Enzymes: From Their Discovery to Their Applications.
<3>Biovalley Monographs, Karger, Postfach, CH-4009, Piguet, P., Poindron, P., Basel, Switzerland
<4>3
<5>33-38
<6>2012
<7>As a contribution to the history of science, the discovery and the functions of bacterial
restriction endonucleases, as well as their
applications in molecular genetic analysis and in biotechnological
innovations, including genetic engineering, are described here from the
personal viewpoint of the author.

<>

<1>Arber, W.
<2>Elements in microbial evolution.
<3>J. Mol. Evol.
<4>33
<5>4-12
<6>1991
<7>Spontaneous mutation, selection, and isolation are key elements in biological evolution.
Molecular genetic approaches reveal a multitude of different mechanisms by which spontaneous
mutants arise. Many of these mechanisms depend on enzymes, which often do not act fully at
random on the DNA, although a large number of sites of action can be observed. Of particular
interest in this respect are DNA rearrangement processes, e.g., by transposition and by
site-specific recombination systems. The development of gene functions has thus to be seen as
the result of both DNA rearrangement processes and sequence alterations brought about by
nucleotide substitutions and small local deletions, insertions, and duplications. Prokaryotic
microorganisms are particularly appropriate for studying the effects of spontaneous mutation
and thus microbial evolution, as they have haploid genomes, so that genetic alterations become
rapidly apparent phenotypically. In addition, bacteria and their viruses and plasmids have
relatively small genomes and short generation times, which also facilitate research on
evolutionary processes. Besides the strategy of development of gene functions in the vertical
transmission of genomes from generation to generation, the acquisition of short DNA segments
from other organisms appears to be an important strategy in microbial evolution. In this
process of horizontal evolution natural vector DNA molecules are often involved. Because of
acquisition barriers, the acquisition strategy works best for relatively small DNA segments,
hence at the level of domains, single genes, or at most operons. Among the many enzymes and
functional systems involved in vertical and horizontal microbial evolution, some may serve
primarily for essential life functions in each individual and only secondarily contribute to
evolution.Others, however, might serve primarily for evolution and thus exert their biological
functions at the level of populations rather than at that of a single organism.

<>

<1>Arber, W.
<2>Genetically encoded generators of genetic variants.
<3>J. Proteomics
<4>72
<5>836-837
<6>2009
<7>Several specific molecular mechanisms contribute to the generation of genetic variants at low
rates. Some of these mechanisms involve the action of specific gene products as variation
generators. We discuss here known as well as still hypothetical ways by which natural reality
may succeed to keep the rates of genetic variation at low levels that insure a relatively high
genetic stability of the individual organisms.

<>

<1>Arber, W., Dussoix, D.
<2>Host Specificity of DNA Producted by Escherichia Coli:  I. Host controlled modification of bacteriophage lambda.
<3>J. Mol. Biol.
<4>5
<5>18-36
<6>1962
<7>Lambda bacteriophage particles carry a "host specificity" determined by the
bacterial strains on which they were produced.  Upon infection of a different
bacterial host (1) the phage DNA may be either accepted or rejected on the
basis of this specificity, (2) if accepted, the phage multiplies and progeny
phage are produced.  Those progeny to which the parental phage DNA molecule is
transferred, in either conserved or semi-conserved form, also receive the
parental phage host specificity.  All progeny containing only newly synthesized
DNA receive only the specificity of the new bacterial host.  It is concluded
that host specificity is carried on the bacteriophage DNA.  Phage P1, present
in a bacterial cell as either prophage or vegetative phage, imparts to lambda
DNA multiplying in the same cell a host specificity over and above that
determined by the host itself.  Such P1-induced specificity can be impressed
equally well onto replicating and non-replicating lambda DNA.

<>

<1>Arber, W., Hattman, S., Dussoix, D.
<2>On the host-controlled modification of bacteriophage lambda.
<3>Virology
<4>21
<5>30-35
<6>1963
<7>Phage lambda.C, grown on Escherichia coli strain C, is restricted on E. coli
strains K12, K12(P1) and B251, and its DNA is broken down after injection into
these bacteria strains.  Phage lambda.K(P1)-that is, grown on K12 or K12(P1)-is
accepted by C, but undergoes host controlled modification upon reproduction in
C (Weigle and Bertani, 1953).  In a one-cycle growth of lambda.K(P1) on E. coli
C, parental DNA and host specificity undergo linked transfer into the progeny.
Thus, host specificity is a property of the DNA molecule, as was previously
shown for phage lambda in another host system (Arber and Dussoix, 1962).  Host
specificity is serially transferable, and the probability of transfer to the
progeny is constant for any given DNA strand under given experimental
conditions.

<>

<1>Arber, W., Kuhnlein, U.
<2>Mutational loss of B-specific restriction of the bacteriophage fd.
<3>Pathol. Microbiol. (Basel)
<4>30
<5>946-952
<6>1967
<7>Bacteriophage fd undergoes host-controlled restriction and modification in
strains of E. coli B: non modified phage fd.O plates with a probability of only
7 x 10-4 on B strains 2027.  Phage mutants u1 were isolated showing an
efficiency of plating of 3 x 10-2 on B.  Starting from such intermediately
restricted fd genomes, completely unrestricted double mutants, u1, u2 were
found.  No notable changes in the physiological properties of the mutants were
observed.  The stepwise loss of restriction is taken as an evidence that the fd
DNA molecule carries two B-specific sites, which are most probably determined
by a specific base sequence, in confirmation of findings made before upon
methylation analysis of fd DNA.

<>

<1>Arber, W., Kuhnlein, U.
<2>Repair and modification of genetic material:  DNA modification and restriction.
<3>Int. Congr. Biochem.
<4>8
<5>180
<6>1970
<7>B-specific modification enzyme has been partly purified from extracts of
Escherichia coli strain B.  This enzyme specifically methylates unmodified DNA
and thereby renders the DNA resistant to cleavage by endonuclease R.B.
S-adenosylmethionine is the methyl donor, and the product of the reaction is
6-methylaminopurine.  These observations confirm older in vivo results on the
chemical nature of strain-specific modification.  Quantitatively, each fully
modified specificity site on the DNA contains two methylated bases, one on each
strand.  At the genetic level, two independent systems of host specificity have
been found to reside in strains of E. coli 15.  One of these systems, called A,
has its genetic determinants located on the bacterial chromosome, linked to the
thr region.  This observation suggests its relation to the K- and B-specificity
types.  The genetic basis for the 15-specific and modification, on the other
hand, resides on a plasmid which by a number of criteria is shown to be related
to phage P1.  In particular, P1 and the plasmid in question seem to possess
homologous replicators.  In vivo complementation data obtained with various
restriction- and modification-deficient mutants measure the functional
relatedness of the various host specificity systems.

<>

<1>Arber, W., Linn, S.
<2>DNA modification and restriction.
<3>Annu. Rev. Biochem.
<4>38
<5>467-500
<6>1969
<7>In this review, we have subdivided the discussion on DNA modification and
restriction into three convenient (if somewhat arbitrary) areas of study:  (a)
the enzymatic activities in vivo and in vitro; (b) the substrate, with the
characterization of "specificity sites" as particular regions on DNA molecules
showing affinity for modification and restriction activities; and (c) the
genetic determinants for the enzymatic activities.  Limitations of length
necessitate the consideration of only those experimental observations which
seem to us the most relevant for the understanding of the phenomena.
Previously published reviews may fill some of the gaps that will appear, but
for the remaining ones we must refer the reader to the original literature.  In
particular, we regret the omission of studies made with bacterial strains other
than E. coli, although many of these systems seem strongly related in their
fundamental properties to those discussed here.  Finaly, the thoroughly
explored restriction by E. coli of unglucosylated T-even phage must also be
omitted.

<>

<1>Arber, W., Morse, M.L.
<2>Host specificity of DNA produced by Escherichia coli.  VI.  Effects on bacterial conjugation.
<3>Genetics
<4>51
<5>137-148
<6>1965
<7>Bacterial host cells endow the DNA of bacteriophage lambda and that of the
transducing phage lambda with host specificity (Arber and Dussoix 1962; Arber
1964).  This label on the DNA plays an important role in phage infection.  In
the absence of the required host specificity, the bacteria degrade the DNA of
the infecting phage, which is then said to be restricted (Dussoix and Arber
1962).  Preliminary evidence has been given by Arber and Dussoix (1961) and
Arber (1962) that bacterial DNA is also subject to this same control mechanism.
The enzymatic system endowing lambda DNA with host specificity would thus act
on the host DNA as well, and the host specificity could play a role in the
decision of whether DNA transferred in conjugation from male to female bacteria
is accepted or rejected.  This last action can be checked by measuring the
frequency of formation of recombinants, provided other factors antagonistic to
genetic integration, such as nonhomology of the male and female DNA molecules,
are also considered.  The present paper gives an account of experiments carried
out to investigate restriction in bacterial conjugation involving Hfr, F+,
F-gal+, F-lac+ and (RTF)+ donor strains.  Since our first experiments the
results have been confirmed and extended by various other investigators (Boice
and Luria 1963; Hoekstra and De Haan 1963; Glover, Schell, Symonds and Stacey
1963; Pittard 1964; Boyer 1964).

<>

<1>Arber, W., Rifat, A., Wauters-Willems, D., Kuhnlein, U.
<2>Host specificity of DNA produced by Escherichia coli.
<3>Mol. Gen. Genet.
<4>115
<5>195-207
<6>1972
<7>Bacteria with A-specific restriction plate unmodified phage lambda with an
efficiency of 10-2.  One mutational event can produce restriction insensitive
(sAo) mutants of lambda.  These differ from the original sA form of lambda by
no other property than their response to A-host specificity.  Two-parental
phage crosses involving sA and sAo, respectively, as non-selective marker
allowed to map sA between genes cII and O.  These data indicate that sA is the
only site on lambda DNA with affinity for A-specific restriction.  Lambda DNA
is thus an interesting substrate in in vitro A-specific restriction and
modification.  Using an assay based on the infectivity of lambda DNA on
helper-infected bacteria, A-specific modification activity was found in
partially purified sonicates of bacteria with A-host specificity.  In parallel
to modification, 3H-methyl label from S-adenosylmethionine, the only cofactor
required for modification, was transferred to unmodified lambda DNA.  No
association of radioactivity was observed in control experiments with DNA from
either modified lambda.A or from a lambda sAo mutant.  These data suggest that
A-specific modification is brought about by DNA methylation and that the sAo
mutation not only abolished the affinity for A-specific restriction, but also
for A-specific modification.

<>

<1>Arber, W., Smith, J.D.
<2>Kinetics of reproduction of nucleic acids and their role in biosynthesis of proteins.
<3>Int. Congr. Microbiol.
<4>5
<5>5
<6>1966
<7>Various filamentous, male specific phages containing single stranded DNA
undergo host-controlled modification in E. coli B and in P1 lysogenic bacteria.
The B-specific modification is directed by those gene regions of E. coli which
also are responsible for host-controlled modification of DNA of the bacterial
host itself, and of other phages such as lambda.  Study of restriction and
modification with phage fd reveals:  (1) a correlation between restriction in
phage reproduction and appearance of acid soluble breakdown products from the
infecting DNA molecule; (2) a correlation between occurrence of modification
and a distinct increase in the level of 6-methylaminopurine carried by the
phage DNA.  E. coli K12 does not restrict phage fd, nor does fd-K show a higher
level of methylated bases than fd grown on mutants of K12 which are deficient
in providing bacterial DNA with K-specific modification.  Absence of K-specific
sites on the small DNA molecule (about 4500 nucleotides) of phage fd could
reasonably explain these facts.  Bacteria of strain K12 infected at very low
multiplicity with 2H and 15N density labelled fdB liberate, among early progeny
phage, particles which still have the parental B-specific modification and
carry heavy, parental DNA molecules wrapped into light, new phage proteins.
This finding, besides confirming that host specificity of phage fd is closely
associated with the DNA molecule, demonstrates that single stranded DNA
molecules of infecting fd phage have a fair chance to be transferred into phage
progeny particles.

<>

<1>Arber, W., Wauters-Willems, D.
<2>Host specificity of DNA produced by Escherichia coli. XII.  The two restriction and modification systems of strain 15T-.
<3>Mol. Gen. Genet.
<4>108
<5>203-217
<6>1970
<7>E. coli 15T- carries two distinct sets of DNA restriction and modification
activities.  The genetic information for system A is contained in the bacterial
chromosome and linked to the thr region.  This fact suggests host specificity A
to be related to those of strains K and B.  The genes controlling system 15 are
on a plasmid which is related to phage P1: it competes with P1 for stable
inheritance in the carried state and it genetically recombines with P1.  This
recombination may produce plasmid genomes with newly assorted characters.  One
of them is an active, P1-like prophage with the 15-specific instead of the
parental P1-specific restriction and modification characters.  Superinfection
of 15T- with P1 may also result in curing of the bacteria from the restriction
plasmid.  Both A- and 15-specific restrictions and modifications act on
bacterial DNA, on the DNA of various sex factors and on the DNA of certain
bacteriophages, e.g. of phage lambda.  Phage 82 DNA is sensitive only to
15-specific restriction, but not to A-specific restriction.  Independently of
the A- and 15-specific restrictions, the growth of phage lambdain E. coli 15T-
encounters another limitation of yet unknown nature.  No such limitation is
observed either with phage 82 or with mutants of lambda occurring at a
frequency of about 10-5.

<>

<1>Arber, W., Yuan, R., Bickle, T.A.
<2>Strain-specific modification and restriction of DNA in bacteria.
<3>Post Synthetic Modification of Macromolecules.  Symposium at 9th FEBS Meeting, Budapest. 1974, Elsevier, Antoni, F., Farago, A., New York
<4>34
<5>3-22
<6>1975
<7>Within the last few years bacterial restriction endonucleases have become an
important tool in genetic, structural and functional studies of DNA.  Many of
these enzymes reproducibly cleave large double-stranded DNA molecules at sites
determined by particular nucleotide sequences.  This yields unique populations
of smaller fragments.  Since the site of cleavage is usually unique for each
restriction endonclease, any genome can in principle be cleaved by application
of one or several properly chosen enzyme preparations into DNA fragments of any
desired length and containing any desired genetic message.  These possibilities
have attracted widespread attention to the molecular mechanism of these
enzymes, in particular their interaction with their DNA substrate.  Bacterial
strains producing restriction endonucleases must necessarily have developed a
mechanism to protect their own DNA from intracellular cleavage.  In those
strains studied up to date, this protection is brought about by the enzymatic
methylation of specific nucleotides and is generally known as strain-specific
modification of DNA.  This modification related methylation which usually
represents only a fraction of the methylated bases in DNA clearly defines one
of the roles of DNA methylation, that is to protect the nucleic acid from
specific endonucleolytic cleavage.  On the other hand, the biological role of
strain-specific restriction of DNA is generally believed to reside in a
primitive system of immunity of bacteria against invasion by foreign genetic
material.  Bacterial strains not possessing any known restriction-modification
system (R-M system) are perfectly viable.  Therefore, restriction appears to be
a dispensable function.  Nevertheless, many naturally occurring bacterial
strains do possess one or several independent R-M systems, which suggests that
restriction is not absolutely worthless to bacteria.  A closer look at several
R-M systems allows us to distinguish two general types of mechanisms.  These
were tentatively called type I and type II by Boyer (1971).  It is the object
of our contribution to this symposium to discuss the present state of knowledge
concerning these two types of restriction and modification mechanisms, and we
would like to do so by presenting selected examples rather than by strictly
defining the types.  It is quite likely that future studies may reveal other
mechanisms with distinctly different features.  Recently published reviews in
this field were written by Boyer (1971), Meselson et al. (1972) and Arber
(1974).  The nomenclature used follows the recommendations made by Arber and
Linn (1969), Smith and Nathans (1973) and Arber (1974).

<>

<1>Archer, C.T., Kim, J.F., Jeong, H., Park, J.H., Vickers, C.E., Lee, S.Y., Nielsen, L.K.
<2>The genome sequence of E. coli W (ATCC 9637): comparative genome analysis and an improved genome-scale reconstruction of E. coli.
<3>BMC Genomics
<4>12
<5>9
<6>2011
<7>biotechnology. E. coli W (ATCC 9637), one of four strains designated as safe for laboratory
purposes, has not been sequenced. E. coli W is a fast-growing strain and is the only safe
strain that can utilize sucrose as a carbon source.  Lifecycle analysis has demonstrated that
sucrose from sugarcane is a preferred carbon source for industrial
bioprocesses.  Results: We have sequenced and annotated the genome of E. coli W. The
chromosome is 4,900,968 bp and encodes 4,764 ORFs. Two plasmids, pRK1 (102,536 bp) and pRK2
(5,360 bp), are also present. W has unique features relative to other sequenced laboratory
strains (K-12, B and Crooks): it has a larger genome and belongs to phylogroup B1 rather than
A. W also grows on a much broader range of carbon sources than does K-12.
A genome-scale reconstruction was developed and validated in order to interrogate metabolic
properties.  Conclusions: The genome of W is more similar to commensal and pathogenic B1
strains than phylogroup A strains, and therefore has greater utility for comparative analyses
with these strains. W should therefore be the strain of choice, or 'type strain' for group
B1 comparative analyses. The genome annotation and tools created here
are expected to allow further utilization and development of E. coli W as an industrial
organism for sucrose-based bioprocesses. Refinements in our E. coli metabolic reconstruction
allow it to more accurately define E. coli metabolism relative to previous models.

<>

<1>Arden, S.B., Barksdale, L.
<2>Restriction and modification and the phage typing of true Corynebacteria.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>71
<5>181
<6>1971
<7>Corynebacterium ulcerans, strain 603, neither restricts nor modifies phages Z
and Zv.  However, C. diphtheriae, strain C7, C. ovis, strain 21, and C.
belfanti, strain 1030, each produces a distinct restricting endonuclease.
These we have designated r1, r2, r3, respectively.  Each is in turn linked to a
distinct host controlled modification which specifically and epigenetically
renders the phage DNA immune to the action of restricting enzyme.  These are
designated m1, m2 and m3.  Thus, whereas strain 603 is m0r0, nonrestricting
nonmodifying, strain C7 is m1r1.  Stocks of Zv phage produced in C7 have the m1
phenotype.  By preparing phage stocks of m0, m1, m2 and m3 phenotypes the
discriminatory powers of these phages have been extended when used in
conjunction with bar mutation and lysogenic immunity in a system of phage
typing.  This system allows for distinguishing between each of 21 bacteria,
some of which differ from one another by only one gene; some by one prophage.
Since the host strains require methionine, certain comparisons between the
basis for the modification in Zv and that reported for lambda can be made.

<>

<1>Argast, G.M., Stephens, K.M., Emond, M.J., Monnat, R.J. Jr.
<2>I-PpoI and I-CreI homing site sequence degeneracy determined by random mutagenesis and sequential in vitro enrichment.
<3>J. Mol. Biol.
<4>280
<5>345-353
<6>1998
<7>Plasmid libraries containing partially randomized cleavage sites for the eukaryotic homing
endonucleases I-PpoI and I-CreI were constructed, and sites that could be cleaved by I-PpoI or
I-CreI were selectively recovered by successive cycles of cleavage and gel separation followed
by religation and growth in Escherichia coli.  Twenty-one different I-PpoI-sensitive homing
sites, including the native homing site, were isolated.  These sites were identical at four
nucleotide positions within the 15 bp homing site, had a restricted pattern of base
substitutions at the remaining 11 positions and displayed a preference for purines flanking
the top strand of the homing site sequence.  Twenty-one different I-CreI-sensitive homing
sites, including the native site, were isolated.  Ten nucleotide positions were identical in
homing site variants that were I-CreI-sensitive and required the addition of SDS for efficient
cleavage product release.  Four of those ten positions were identical in homing sites that did
not require SDS for product release.  There was a preference for pyrimidines flanking the top
strand of the homing site sequence.  Three of the 24 I-CreI homing site nucleotide positions
apparently lacked informational content, i.e. were permissive of cleavage when occupied by any
nucleotide.  These results suggest that I-PpoI and I-CreI make a larger number of DNA-protein
contacts across their homing site sequences, and that different subsets of these contacts may
be sufficient to maintain a high degree of sequence-specific homing site recognition and
cleavage.  The sequential enrichment protocol we used should be useful for defining the
sequence degeneracy and informational content of other homing endonuclease target sites.

<>

<1>Argos, P.
<2>Evidence for a repeating domain in type I restriction enzymes.
<3>EMBO J.
<4>4
<5>1351-1355
<6>1985
<7>The primary structures of the recognition subunit (hsdS) in type I restriction
enzymes from three isolates of Escherichia coli were compared and aligned by
use of amino acid physical properties.  A repeating domain was found in each of
the subunits suggesting a pseudo-dimeric structure.  Secondary structure
predictions delineated two helical regions in each domain which suggested that
the recognition subunits may act in a fashion similar to that proposed for
repressor and activator molecules; namely, interaction with double-stranded DNA
through helices and in two successive major grooves on the same DNA side.  One
helical motif could provide the specific recognition site and the other, the
restriction site.

<>

<1>Arigoni, F., Delley, M., Mollet, B., Pridmore, R.D., Schell, M.A., Pohl, T., Zwahlen, M.-C.
<2>NCC2705--the genome of a bifidobacterium.
<3>US Patent Office
<4>US 7183101 A
<5>
<6>2007
<7>A novel microorganism of the genus Bifdobacterium longum, in particular to its genomic and
plasmid sequence and to a method of producing polypeptides of said Bifidobacterium,
respectively.  Also methods of detecting these nucleic acids or polypeptides, respectively.  A
data carrier is provided comprising nucleotide sequences and/or polypeptide sequences of
NCC2705.  In addition, the Bifidobacterium longum strain NCC2705 and also to food and
pharmaceutical compositions containing said Bifidobacterium or active components thereof for
the prevention and/or treatment of diarrhoea brought about by rotaviruses and pathogenic
bacteria are provided.

<>

<1>Arigoni, F., Delley, M., Mollet, B., Pridmore, R.D., Schell, M.A., Pohl, T.G., Zwahlen, M.C.
<2>The genome of a Bifidobacterium.
<3>International Patent Office
<4>WO 02074798 A
<5>
<6>2002
<7>The present invention pertains to a novel micro-organism of the genus Bifidobacterium longum,
in particular to its genomic and plasmid sequence and to a method of producing polypeptides of
said Bifidobacterium, respectively.  The invention also relates to methods of detecting these
nucleic acids or polypeptides, respectively.  A data carrier is provided comprising nucleotide
sequences and/or polypeptide sequences of NCC2705.  In addition, the present invention
pertains to the Bifidobacterium longum strain NCC2705 and also to food and pharmaceutical
compositions containing said Bifidobacterium or active components thereof for the prevention
and/or treatment of diarrhoea brought about by rotaviruses and pathogenic bacteria.

<>

<1>Arigoni, F., Delley, M., Mollet, B., Pridmore, R.D., Schell, M.A., Pohl, T.G., Zwahlen, M.C.
<2>The genome of a Bifidobacterium.
<3>European Patent Office
<4>EP 1227152
<5>
<6>2002
<7>
<>

<1>Arimondo, P.B.
<2>IDENTIFICATION AND DESIGN OF NEW C5-DNA METHYLTRANSFERASE INHIBITORS AND THEIR BIOLOGICAL ACTIVITY.
<3>Anticancer Res.
<4>34
<5>5813-5814
<6>2014
<7>DNA methylation is involved in the regulation of gene expression and plays an important role
in normal developmental processes and disease.  In particular, the epigenetic landscape is
altered in cancers where abnormal hypermethylation leads to silencing of certain genes such as
tumor suppressor genes.  In mammals, DNA methyltransferases are the enzymes responsible for
DNa methylation on the position 5 of cytidine in a CpG context.  Few direct enzyme inhibitors
are known and those have several drawbacks.  In order to identify novel inhibitors, we
developed three chemical strategies.  First a fluorescene High-Throughput Screening for the
inhibition of the murine catalytic Dnmt3a/3L complex on the chemical library of the Museum
Naturelle d'Historie Naturelle and found twelve hits with low micromolar activities.
Interestingly, they showed little cytotoxicity.  Dichlone, a small halogenated naphthoquinone,
classically used as pesticide and fungicide, showed the lowest Ec50 at 460 NM.  Two molecules
including Dichlone, efficiently reactivated YFP gene expression in a stable HEK293 cell line
by promoter demethylation.  Their efficacy was comparable to the DNMT inhibitor of reference
5-azecytidine.  Second, based on molecular modeling studies of quinolone inhibitor SGI1027 in
the crystal structure of M.HhaI C5 DNA methyltransferase, suggesting that the quinolone and
the aminopyridimine are important for the interaction with the substrates and the protein, we
synthesized twenty five new derivatives.

<>

<1>Armalyte, E., Bujnicki, J.M., Giedriene, J., Gasiunas, G., Kosinski, J., Lubys, A.
<2>Mva1269I: A monomeric type IIS restriction endonuclease from micrococcus varians with two EcoRI- and FokI-like catalytic domains.
<3>J. Biol. Chem.
<4>280
<5>41584-41594
<6>2005
<7>Type II restriction endonuclease Mva1269I recognizes an asymmetric DNa sequence 5'-GAATGCN*
-3'/5' -NG* CATTC-3' and cuts top and bottom DNA strands at positions, indicated by the "*"
symbol.  Most restriction endonucleases require dimerization to cleave both strands of DNA.
We found that Mva1269I is a monomer both in solution and upon binding of cognate DNA.  Protein
fold-recognition analysis revealed that Mva1269I comprises two "PD-(D/E)XK" domains.  The
N-terminal domain is related to the 5'-GAATTC-3'-specific restriction endonuclease EcoRI,
whereas the C-terminal one resembles the nonspecific nuclease domain is related to the
5'-GAATTC-3'-specific restriction endonuclease EcoRI, whereas the C-terminal one resembles
the nonspecific nuclease domain of restriction endonuclease FokI.  Inactivation of the
C-terminal catalytic site transformed Mva1269I into a very active bottom strand-nicking
enzyme, whereas mutants in the N-terminal domain nicked the top strand, but only at elevated
enzyme concentrations.  We found that the cleavage of the bottom strand is a prerequisite for
the cleavage of the top strand.  We suggest that Mva1269I evolved the ability to recognize and
to cleave its asymmetrical target by a fusion of an EcoRI-like domain, which incises the
bottom strand within the target, and a FokI-like domain that completes the cleavage within the
nonspecific region outside the target sequence.  Our results have implications for the
molecular evolution of restriction endonucleases, as well as for perspectives of engineering
new restriction and nicking enzymes with asymmetric target sites.

<>

<1>Armbrust, E.V. et al.
<2>The genome of the diatom Thalassiosira pseudonana: ecology, evolution, and metabolism.
<3>Science
<4>306
<5>79-86
<6>2004
<7>Diatoms are unicellular algae with plastids acquired by secondary endosymbiosis. They are
responsible for approximately 20% of global carbon
fixation. We report the 34 million-base pair draft nuclear genome of the
marine diatom Thalassiosira pseudonana and its 129 thousand-base pair
plastid and 44 thousand-base pair mitochondrial genomes. Sequence and
optical restriction mapping revealed 24 diploid nuclear chromosomes. We
identified novel genes for silicic acid transport and formation of
silica-based cell walls, high-affinity iron uptake, biosynthetic enzymes
for several types of polyunsaturated fatty acids, use of a range of
nitrogenous compounds, and a complete urea cycle, all attributes that
allow diatoms to prosper in aquatic environments.

<>

<1>Armitano, R.I., Zerbetto, De.P.G., Matteo, M.J., Revale, S., Romero, S., Traglia, G.M., Catalano, M.
<2>Draft Genome Sequences of Helicobacter pylori Strains HPARG63 and HPARG8G, Cultured from Patients with Chronic Gastritis and Gastric Ulcer Disease.
<3>Genome Announcements
<4>1
<5>e00700-13
<6>2013
<7>Helicobacter pylori colonizes the human gastric mucosa, leading to a spectrum of  gastric
diseases in susceptible populations. Here we announce the draft genome
sequences of strains HPARG8G and HPARG63. The data for both genome sequences
provide insights regarding the diversity in gene content and rearrangement of the
genomic islands commonly harbored by H. pylori.

<>

<1>Armstrong, K., Bauer, W.R.
<2>Preferential site-dependent cleavage by restriction endonuclease PstI.
<3>Nucleic Acids Res.
<4>10
<5>993-1007
<6>1982
<7>The four identical recognition sites for the restriction endonuclease PstI in
purified plasmid pSM1 DNA I are cleaved at markedly different rates.  The order
and relative frequencies of cleavage at these four PstI sites have been
determined from the order of appearance of partial cleavage products and from
an analysis of production of specific unit length linear molecules.  The same
pattern of preferential cleavage is also found when linear, nicked circular, or
relaxed closed circular forms of the same plasmid DNA are used as substrates
for PstI.  Inspection of the nucleotide sequences immediately adjoining each of
the PstI sites suggests that the presence of adjacent runs of G-C base pairs
confers significant resistance to cleavage.

<>

<1>Armstrong, K.A., Bauer, W.R.
<2>Site-dependent cleavage of pBR322 DNA by restriction endonuclease HinfI.
<3>Nucleic Acids Res.
<4>11
<5>4109-4126
<6>1983
<7>Cleavage of pBR322 DNA I by the restriction endonuclease HinfI is preferentially inhibited at
specific HinfI cleavage sites.  These sites in pBR322 DNA I have been identified and ordered
with respect to the frequency with which they are cleaved.  The HinfI site most resistant to
cleavage in pBR322 DNA I is unique in that runs of G-C base pairs are immediately adjacent on
both sides.  Two differently permuted linear (DNA III) species were produced by cleavage with
two different restriction endonucleases, PstI and AvaI.  Only one of these linear molecules,
that produced by PstI, exhibits the same preferential cleavage pattern as DNA I.  The second
linear species, that arising from AvaI digestion, shows pronounced relative inhibition of
cleavage at the HinfI sites nearest the ends of the molecule (100 and 120 base pairs away,
respectively).  This result suggests that proximity to the termini of a linear DNA molecule
might also influence preferential cleavage.  The possibility of formation of stem-loop
structures does not appear to influence preferential cleavage by HinfI.

<>

<1>Arnold, H.P., Ziese, U., Zillig, W.
<2>SNDV, a novel virus of the extremely thermophilic and acidophilic archaeon Sulfolobus.
<3>Virology
<4>272
<5>409-416
<6>2000
<7>We describe a novel virus, SNDV (Sulfolobus neozealandicus droplet-shaped virus), of the
crenarchaeotal archaeon Sulfolobus, which was found in a carrier state in a Sulfolobus strain
isolated from a field sample from New Zealand. SNDV particles are droplet-shaped and densely
covered by thin tail fibers at their pointed ends. The virion consists of a core and a coat.
The latter has the appearance of a beehive and has a surface that is either helically ribbed
or a stack of hoops. The genome is cccDNA of 20 kb, which is modified by dam-like methylation.
It is cleaved by only a few type II restriction enzymes e.g., DpnI but not MboI, demonstrating
an N(6)-methylation of the adenine residue in GATC sequences. The DNA-modifying system
differentiates between virus and host. We postulate a virus-encoded methylase that is active
on hemimethylated DNA. The host range of SNDV is confined to few Sulfolobus strains from New
Zealand.  The virus persists in an unstable carrier state rather than as a prophage. Due to
its uniqueness we propose to assign it to a novel virus family termed Guttaviridae.

<>

<1>Arnould, S., Bruneau, S., Cabaniols, J.-P., Chames, P., Choulika, A., Duchateau, P., Epinat, J.-C., Gouble, A., Lacroix, E., Paques, F., Perez-Michaut, C., Smith, J., Sourdive, D.
<2>Custom-made meganuclease and use thereof.
<3>International Patent Office
<4>WO 200467736 A
<5>124
<6>2004
<7>New rare-cutting endonucleases, also called custom-made meganucleases, which recognize and
cleave a specific nucleotide sequence, derived polynucleotide sequences, recombinant vector
cell, animal, or plant comprising said polynucleotide sequences, process for producing said
rare-cutting endonucleases and any use thereof, more particularly, for genetic engineering,
antiviral therapy and gene therapy.

<>

<1>Arnould, S., Chames, P., Choulika, A., Epinat, J.-C., Lacroix, E.
<2>Hybrid and single chain meganucleases and use thereof.
<3>European Patent Office
<4>EP 1485475 B
<5>
<6>2007
<7>This patent application relates to hybrid and/or single-chain rare-cutting endonucleases,
called meganucleases, which recognize and cleave a specific nucleotide sequence, to
polynucleotide sequences encoding for said rare-cutting endonucleases, to a vector comprising
one of said polynucleotide sequences, to a cell or animal non-human comprising one of said
polynucleotide sequences or said rare-cutting endonucleases, to a process for producing one of
said rare-cutting endonucleases and any use of the disclosed products and methods.  More
particularly, this invention contemplates any use of such rare-cutting endonuclease for
genetic engineering and gene therapy.

<>

<1>Arnould, S., Chames, P., Perez, C., Lacroix, E., Duclert, A., Epinat, J.C., Stricher, F., Petit, A.S., Patin, A., Guillier, S., Rolland, S., Prieto, J., Blanco, F.J., Bravo, J., Montoya, G., Serrano, L., Duchateau, P., Paques, F.
<2>Engineering of large numbers of highly specific homing endonucleases that induce recombination on novel DNA targets.
<3>J. Mol. Biol.
<4>355
<5>443-458
<6>2005
<7>The last decade has seen the emergence of a universal method for precise and efficient genome
engineering. This method relies on the use of sequence-specific endonucleases such as homing
endonucleases. The structures of several of these proteins are known, allowing for
site-directed mutagenesis of residues essential for DNA binding. Here, we show that a
semi-rational approach can be used to derive hundreds of novel proteins from I-CreI, a homing
endonuclease from the LAGLIDADG family. These novel endonucleases display a wide range of
cleavage patterns in yeast and mammalian cells that in most cases are highly specific and
distinct from I-CreI. Second, rules for protein/DNA interaction can be inferred from
statistical analysis. Third, novel endonucleases can be combined to create heterodimeric
protein species, thereby greatly enhancing the number of potential targets. These results
describe a straightforward approach for engineering novel endonucleases with tailored
specificities, while preserving the activity and specificity of natural homing endonucleases,
and thereby deliver new tools for genome engineering.

<>

<1>Arnould, S., Delenda, C., Grizot, S., Desseaux, C., Paques, F., Silva, G.H., Smith, J.
<2>The I-CreI meganuclease and its engineered derivatives: applications from cell modification to gene therapy.
<3>Protein Eng. Des. Sel.
<4>24
<5>27-31
<6>2011
<7>Meganucleases (MNs) are highly specific enzymes that can induce homologous recombination in
different types of cells, including
mammalian cells. Consequently, these enzymes are used as scaffolds for
the development of custom gene-targeting tools for gene therapy or
cell-line development. Over the past 15 years, the high resolution
X-ray structures of several MNs from the LAGLIDADG family have improved
our understanding of their protein-DNA interaction and mechanism of DNA
cleavage. By developing and utilizing high-throughput screening methods
to test a large number of variant-target combinations, we have been
able to re-engineer scores of I-CreI derivatives into custom enzymes
that target a specific DNA sequence of interest. Such customized MNs,
along with wild-type ones, have allowed for exploring a large range of
biotechnological applications, including protein-expression cell-line
development, genetically modified plants and animals and therapeutic
applications such as targeted gene therapy as well as a novel class of
antivirals.

<>

<1>Arnould, S., Perez, C., Cabaniols, J.-P., Smith, J., Gouble, A., Grizot, S., Epinat, J.-C., Duclert, A., Duchateau, P., Paques, F.
<2>Engineered I-Crel derivatives cleaving sequences from the human XPC gene can induce highly efficient gene correction in mammalian cells.
<3>J. Mol. Biol.
<4>371
<5>49-65
<6>2007
<7>Meganucleases are sequence-specific endonucleases which recognize large (> 12 bp) target sites
in living cells and can stimulate homologous
gene targeting by a 1000-fold factor at the cleaved locus. We have
recently described a combinatorial approach to redesign the I-Crel
meganuclease DNA-binding interface, in order to target chosen
sequences. However, engineering was limited to the protein regions
shown to directly interact with DNA in a base-specific manner. Here, we
take advantage of I-Crel natural degeneracy, and of additional
refinement steps to extend the number of sequences that can be
efficiently cleaved. We searched the sequence of the human XPC gene,
involved in the disease Xeroderma Pigmentosum (XP), for potential
targets, and chose three sequences that differed from the I-Crel
cleavage site over their entire length, including the central four
base-pairs, whose role in the DNA/protein recognition and cleavage
steps remains very elusive. Two out of these targets could be cleaved
by engineered I-Crel derivatives, and we could improve the activity of
weak novel meganucleases, to eventually match the activity of the
parental I-Crel scaffold. The novel proteins maintain a narrow cleavage
pattern for cognate targets, showing that the extensive redesign of the
I-Crel protein was not made at the expense of its specificity. Finally,
we used a chromosomal reporter system in CHO-K1 cells to compare the
gene targeting frequencies induced by natural and engineered
meganucleases. Tailored I-Crel derivatives cleaving sequences from the
XPC gene were found to induce high levels of gene targeting, similar to
the I-Crel scaffold or the I-SceI "gold standard". This is the first
time an engineered homing endonuclease has been used to modify a
chromosomal locus.

<>

<1>Arraj, J.A., Marinus, M.G.
<2>Phenotypic reversal in dam mutants of Escherichia coli K-12 by a recombinant plasmid containing the dam+ gene.
<3>J. Bacteriol.
<4>153
<5>562-565
<6>1983
<7>A recombinant plasmid, pMQ3, carrying the dam gene of Escherichia coli K-12, was constructed
and transformed into dam+ and dam- strains. Both dam- and dam+ strains containing pMQ3 showed
a wild phenotype for all traits, including mutation rate, except for a 10-fold increase in DNA
adenine methylase activity.

<>

<1>Arraj, J.A., Wu, T.-H., Marinus, M.G.
<2>Expression of a DNA methylation (dam) gene in Escherichia coli K-12.
<3>Curr. Microbiol.
<4>20
<5>133-136
<6>1990
<7>Plasmid pMQ3, carrying the dam gene of Escherichia coli on a 6.1 Kb fragment, shows a tenfold
increase in relative DNA adenine methylase activity, while plasmid pdam118, with a 1.14 Kb dam
insert, shows only a twofold increase, although both plasmids were derived from plasmid
pLC13-42. Since a copy number effect did not seem to be the cause of this difference, we have
subcloned pMQ3 in order to determine whether the additional chromosomal DNA present in this
plasmid is responsible for the enhancement of methylase activity. We show that the 346 base
pairs upstream of dam contain sequences necessary for expression. DNA sequence analysis has
revealed that in pdam 118 only the 118 bases 5' to the dam gene are present in other
constructs and that the additional upstream material is pBR322 DNA. This shows that pdam118
carries a DNA duplication.

<>

<1>Arrand, J.R., Myers, P.A., Roberts, R.J.
<2>A new restriction endonuclease from Streptomyces albus G.
<3>J. Mol. Biol.
<4>118
<5>127-135
<6>1978
<7>A restriction endonuclease, SalI, has been partially purified from Streptomyces
albus G.  This enzyme cleaves adenovirus-2 DNA at three sites, bacteriophage
lambda DNA at two sites, but does not cleave simian virus 40 DNA or PhiX174
DNA.  It recognizes the sequence 5'-G^T-C-G-A-C-3' 3'-C-A-G-C-T-^G-5' and cuts
at the siteds indicated by the arrows.  An endonuclease (XamI) with similar
specificity has also been isolated from Xanthomonas amaranthicola.

<>

<1>Arsene-Ploetze, F. et al.
<2>Structure, function, and evolution of the Thiomonas spp. genome.
<3>PLoS Genet.
<4>6
<5>e1000859
<6>2010
<7>Bacteria of the Thiomonas genus are ubiquitous in extreme environments, such as arsenic-rich
acid mine drainage (AMD).  The genome of one of these strains, Thiomonas sp. 3As, was
sequenced, annotated, and examined, revealing specific adaptations allowing this bacterium to
survive and grow in its highly toxic environment. In order to explore genomic diversity as
well as genetic evolution in Thiomonas spp., a comparative genomic hybridization (CGH)
approach was used on eight different strains of the Thiomonas genus, including five strains of
the same species. Our results suggest that the Thiomonas genome has evolved through the gain
or loss of genomic islands and that this evolution is influenced by the specific environmental
conditions in which the strains live.

<>

<1>Artyukhin, A.B., Woo, Y.H.
<2>DNA extraction method with improved efficiency and specificity using DNA methyltransferase and 'click' chemistry.
<3>Anal. Biochem.
<4>425
<5>169-174
<6>2012
<7>In an attempt to develop an alternative method to extract DNA from complex samples with much
improved sensitivity and efficiency, here we
report a proof-of-concept work for a new DNA extraction method using
DNA methyltransferase (Mtase) and 'click' chemistry. According to our
preliminary data, the method has improved the current methods by (i)
employing a DNA-specific enzyme, TaqI DNA Mtase, for improved
selectivity, and by (ii) capturing the DNA through covalent bond to the
functionalized surface, enabling a broad range of treatments yielding
the final sample DNA with minimal loss and higher purity such that it
will be highly compatible with downstream analyses. By employing Mtase,
a highly DNA specific and efficient enzyme, and click chemistry, we
demonstrated that as little as 0.1 fg of lambda-DNA (close to copy
number 1) was captured on silica (Si)-based beads by forming a covalent
bond between an azide group on the surface and the propargyl moiety on
the DNA. This method holds promise in versatile applications where
extraction of minute amounts of DNA plays critical roles such as basic
and applied molecular biology research, bioforensic and biosecurity
sciences, and state-of-the-art detection methods.

<>

<1>Arutyunyan, E.E., Gonchar, N.A., Gruber, I.M., Nikolskaya, N.I.
<2>Purification and characterization of DNA methyltransferase Sau6782 of Staphylococcus aureus.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>11-1
<5>26-29
<6>1992
<7>
<>

<1>Arutyunyan, E.E., Gonchar, N.A., Levchenko, I.Y., Gruber, I.M., Nikolskaya, I.I.
<2>Isoelectrofocusing of methylation and restriction enzymes of Staphylococcus aureus 6782.
<3>Biokhimiia
<4>56
<5>281-288
<6>1991
<7>The behaviour of methylation and restriction enzymes of Staphylococcus aureus
6782 during their isoelectrofocusing on ampholines was studied.  It was found
that the R.Sau6782I enzyme is represented by two isoforms, RI and RII, with
isoelectric points of 4.2 and 7.9, respectively.  Data from isoelectrofocusing
analysis suggest that RI and RII are devoid of the relaxed specificity found in
the original preparation.  It was shown that the relaxed specificity is also
inherent in the isoschizomeric enzyme, R.Sau3AI.  Isoelectrofocusing of the
original preparation of R.Sau3AI, as is the case for R.Sau6782I, allows the
identification of two peaks, RI and RII, and the separation of each peak from
the trace activity.  Multiple forms of DNA-methylase of the Sau6782I type are
represented by four isoenzymes possessing acidic properties.  The method allows
one to single out from the total methylase pool a modifying methylase with pI
(3.9) that is close to that of R.Sau6782I and thus the enzyme cannot serve for
correct separation of restriction and methylation enzymes of Sau6782I.

<>

<1>Arutyunyan, E.E., Gruber, I.M., Polyachenko, V.M., Kvachadze, L.J., Andriashvili, I.A., Chanishvili, T.G., Nikolskaya, I.I.
<2>Restricting Endonuclease SAU 6782.
<3>Vopr. Med. Khim.
<4>31
<5>127-132
<6>1985
<7>Data are described on identification, isolation and purification of restricting
endonuclease Sau 6782 as well as on estimation of the enzyme recognition site.
Conditions were developed for growing of Staphylococcus aureus 6782 strain,
which enabled to produce a maximal yield of the restricting activity containing
minimal level of nucleases.  The procedure for isolation and purification of
restrictase Sau 6782 involved affinity chromatography on Blue Sepharose and
cation exchange chromatography on phosphocellulose PII.  The enzyme preparation
obtained was free from impurities of unspecific nucleases.  The yield of the
Sau 6782 restrictase constituted 1,0 un from 1 g of the culture cells.
Restrictase Sau 6782 recognized the nucleotide sequence 5'...GATC...3' and was
the isoshizomere of the Sau 3A enzyme.

<>

<1>Arutyunyan, E.E., Gruber, I.M., Polyachenko, V.M., Nikolskaya, I.I., Debov, S.S.
<2>Isolation of methylases from Sau 6782 strain.
<3>Vopr. Med. Khim.
<4>36
<5>75-79
<6>1990
<7>A heterogenous profile of methylases was found in S. aureus 6782.  A procedure
was developed for isolation of individual methylases from Sau 6782, free of Sau
6782 restrictases and unspecific nucleases, by means of hydrophobic
chromatography on phenyl-Sepharose.  Ion exchange, affinity chromatographies
and gel filtration were also used for isolation of the Sau 6782 methylases.
But this technique was of limited suitability for isolation of individual
methylating enzymes of the Sau 6782 type because it did not allow the complete
separation of these methylases from contaminating enzymes of DNA degradation.
Effects of salts, glycerol and Triton X-100 on activity of total preparation of
methylases Sau 6782 were studied.  Na+ and K+ chlorides, ammonium sulfate at
concentrations 0.4 M and higher as well as Triton X-100 reversibly inhibited
the methylase activity followed by complete reduction up to initial level after
dialysis.  Glycerol at 60% concentration activated Sau 6782 methylases by 50%
and stabilized the enzyme.

<>

<1>Arwert, F., Rutberg, L.
<2>Restriction and modification in Bacillus subtilis.  Induction of a modifying activity in Bacillus subtilis 168.
<3>Mol. Gen. Genet.
<4>133
<5>175-177
<6>1974
<7>Bacillus subtilis strain 5GR will restrict and modify phage SPO2 previously grown in strain
168.  SPO2 grown in 168 pretreated with Mitomycin C is less restricted by 5GR.  Strain MB500
is temperature inducible for the defective PBSX prophage.  SPO2 grown in MB500 at inducing
temperature is less restricted by 5GR compared to phage grown in MB500 at non-inducing
temperature.  It is suggested that strain 168 carries genetic determinants for modification of
SPO2 DNA, and that those determinants may be associated with the defective phage PBSX.

<>

<1>Arya, G., Petronella, N., Crosthwait, J., Carrillo, C.D., Shwed, P.S.
<2>Draft Genome Sequence of Bacillus megaterium Type Strain ATCC 14581.
<3>Genome Announcements
<4>2
<5>e01124-14
<6>2014
<7>Bacillus megaterium is a Gram-positive, rod-shaped, spore-forming bacterium of
biotechnological importance. Here, we report a 5.7-Mbp draft genome sequence of
B. megaterium ATCC 14581, which is the type strain of the species.

<>

<1>Asahina, A.Y., Hadfield, M.G.
<2>Complete Genome Sequence of Cellulophaga lytica HI1 Using PacBio Single-Molecule  Real-Time Sequencing.
<3>Genome Announcements
<4>2
<5>e01148-14
<6>2014
<7>We report here the complete genome sequence of Cellulophaga lytica HI1 isolated from a
seawater table located at the Kewalo Marine Laboratory (Honolulu, HI).
This is the first complete de novo genome assembly of C. lytica HI1 using PacBio
single-molecule real-time (SMRT) sequencing, which resulted in a single scaffold
of 3.8 Mb.

<>

<1>Asai, A., Hirai, H., Bodell, W.J., Hoshino, T.
<2>Restriction endonuclease recognition and Southern hybridization of bromodeoxyuridine-substituted genomic DNA.
<3>Cell Prolif.
<4>26
<5>271-280
<6>1993
<7>Human glioma cell lines exposed to various concentrations of bromodeoxyuridine (BrdUrd) were
studied to determine the effect of BrdUrd substitution on restriction endonuclease recognition
and Southern hybridization of genomic DNA. BrdUrd substitution had no effect on the
recognition of restriction endonucleases. When the exposure to BrdUrd was 2 h or less and the
BrdUrd substitution rate was less than 40%, there was no difference in the density of
hybridized bands after Southern hybridization using human non-recombinant complementary DNA as
a probe. Hybridization was suppressed significantly by exposures longer than 24 h or BrdUrd
substitution rates greater than 40%. These results suggest that the Brdurd substitution rate
and the exposure time to BrdUrd influence the hybridization reaction by a DNA probe. Brief
exposure (up to 2 h) to BrdUrd does not influence restriction endonuclease recognition or
Southern hybridization of genomic DNA.

<>

<1>Asakura, Y., Kobayashi, I.
<2>From damaged genome to cell surface: transcriptome changes during bacterial cell death triggered by loss of a restriction-modification gene  complex.
<3>Nucleic Acids Res.
<4>37
<5>3021-3031
<6>2009
<7>Genetically programmed cell deaths play important roles in unicellular prokaryotes. In
postsegregational killing, loss of a gene complex from a
cell leads to its descendants' deaths. With type II
restriction-modification gene complexes, such death is triggered by
restriction endonuclease's attacks on under-methylated chromosomes. Here,
we examined how the Escherichia coli transcriptome changes after loss of
PaeR7I gene complex. At earlier time points, activation of SOS genes and
sigma(E)-regulon was noticeable. With time, more SOS genes,
stress-response genes (including sigma(S)-regulon, osmotic-, oxidative-
and periplasmic-stress genes), biofilm-related genes, and many hitherto
uncharacterized genes were induced, and genes for energy metabolism,
motility and outer membrane biogenesis were repressed. As expected from
the activation of sigma(E)-regulon, the death was accompanied by cell
lysis and release of cellular proteins. Expression of several
sigma(E)-regulon genes indeed led to cell lysis. We hypothesize that some
signal was transduced, among multiple genes involved, from the damaged
genome to the cell surface and led to its disintegration. These results
are discussed in comparison with other forms of programmed deaths in
bacteria and eukaryotes.

<>

<1>Asakura, Y., Kojima, H., Kobayashi, I.
<2>Evolutionary genome engineering using a restriction-modification system.
<3>Nucleic Acids Res.
<4>39
<5>9034-9046
<6>2011
<7>Modification of complex microbial cellular processes is often necessary to obtain organisms
with particularly favorable characteristics, but such
experiments can take many generations to achieve. In the present article,
we accelerated the experimental evolution of Escherichia coli populations
under selection for improved growth using one of the
restriction-modification systems, which have shaped bacterial genomes.
This resulted in faster evolutionary changes in both the genome and
bacterial growth. Transcriptome/genome analysis at various stages enabled
prompt identification of sequential genome rearrangements and dynamic
gene-expression changes associated with growth improvement. The changes
were related to cell-to-cell communication, the cell death program, as
well as mass production and energy consumption. These observed changes
imply that improvements in microorganism population growth can be achieved
by inactivating the cellular mechanisms regulating fraction of active
cells in a population. Some of the mutations were shown to have additive
effects on growth. These results open the way for the application of
evolutionary genome engineering to generate organisms with desirable
properties.

<>

<1>Asamizu, E., Sato, S., Kaneko, T., Nakamura, Y., Kotani, H., Miyajima, N., Tabata, S.
<2>Structural analysis of Arabidopsis thaliana chromosome 5. VIII. Sequence features of the regions of 1,081,958 bp covered by seventeen physically assigned P1 and TAC clones.
<3>DNA Res.
<4>5
<5>379-391
<6>1998
<7>A total of 17 Pl and TAC clones each representing an assigned region of
chromosome 5 were isolated from P1 and TAC genomic libraries of
Arabidopsis thaliana Columbia, and their nucleotide sequences were
determined. The length of the clones sequenced in this study summed up to
1,081,958 bp. As we have previously reported the sequence of 9,072,622 bp
by analysis of 125 P1 and TAC clones, the total length of the sequences of
chromosome 5 determined so far is now 10,154,580 bp. The sequences were
subjected to similarity search against protein and EST databases and
analysis with computer programs for gene modeling. As a consequence, a
total of 253 potential protein-coding genes with known or predicted
functions were identified. The positions of exons, which do not show
apparent similarity to known genes were also assigned using computer
programs for exon prediction. The average density of the genes identified
in this study was 1 gene per 4277 bp. Introns were observed in 74% of the
potential protein genes, and the average number per gene and the average
length of the introns were 4.3 and 168 bp, respectively. The sequence data
and gene information are available on the World Wide Web database KAOS
(Kazusa Arabidopsis data Opening Site) at http://www.kazusa.or.jp/arabi/.

<>

<1>Ashapkin, V.V., Kutueva, L.I., Vanyushin, B.F.
<2>The gene for domains rearranged methyltransferase (DRM2) in Arabidopsis thaliana plants is methylated at both cytosine and adenine residues.
<3>FEBS Lett.
<4>532
<5>367-372
<6>2002
<7>The methylation patterns of cytosine and adenine residues in the Arabidopsis thaliana gene for
domains rearranged methyltransferase (DRM2) were studied in wild-type and several transgene
plant lines containing antisense fragments of the cytosine DNA-methyltransferase gene METI
under the control of copper-inducible promoters. It was shown that the promoter region of the
DRM2 gene is mostly unmethylated at the internal cytosine residue in CCGG sites whereas the
3'-end proximal part of the gene coding region is highly methylated. The DRM2 gene was found
to be also methylated at adenine residues in some GATC sequences. Cytosine methylation in CCGG
sites and adenine methylation in GATC sites in the DRM2 gene are variable between wild-type
and different transgenic plants. The induction of antisense METI constructs with copper ions
in transgene plants in most cases leads to further alterations in the DRM2 gene methylation
patterns.

<>

<1>Ashapkin, V.V., Kutueva, L.I., Vanyushin, B.F.
<2>Is the cytosine DNA methyltransferase gene MET1 regulated by DNA methylation in Arabidopsis thaliana plants?
<3>Genetika
<4>47
<5>279-288
<6>2011
<7>The methylation patterns of the MET1 gene in organs of Arabidopsis thaliana were studied by
Southern blot hybridization of DNA samples
hydrolyzed with differentially methylation-sensitive restriction
endonucleases. A highly methylated on internal cytosine residue CCGG
site was found 1.5 kb upstream of the gene, whereas CCGG sites located
in more proximal parts of the 5'-flanking region and the gene itself
are essentially unmethylated. This methylation pattern was observed in
different organs of plants belonging to two different ecotypes as well
as in different transgenic plant lines. The methylation level of a CCGG
site in exon 3 (2.1 kb from the gene's 5'-end) occurred to be variable
between different transgenic plant lines and two ecotypes studied.
Transcription levels of the MET1 gene vary slightly in organs of
wild-type plants without any obvious correlation with its methylation.
The transgenic antisense MET1 constructs expressed in plant genome do
affect both MET1 methylation and its transcription but again without
any obvious correlation. The comparative investigation of transcription
levels of different genes of cytosine DNA methyltransferase family MET
(MET1, MET2a, MET2b, MET3) and their methylation patterns shows that
there may exist some mechanisms defending the most actively transcribed
gene MET1 of this family from methylation mediated silencing. In
contrast to DRM2 gene we could not find any adenine methylated GATC
sites in the MET1 gene.

<>

<1>Ashraf, M., Altschuler, M., Galasinski, S., Griffiths, T.D.
<2>Alteration of gene expression by restriction enzymes electroporated into plant cells.
<3>Mutat. Res.
<4>302
<5>75-82
<6>1993
<7>The alteration in the expression of a B-glucuronidase (GUS) reporter gene was used to monitor
the effect of restriction endonuclease electroporated into the tobacco (Nicotiana tabacum L.)
protoplasts. Restriction enzyme (RE) HindIII which does not have a recognition site within the
gene cassette, had little effect on enzyme activity. In contrast restriction endonucleases
HaeIII and Sau3AI which possess 8 and 16 recognition sites in the GUS cassette, were found to
reduce the enzyme activity by 89% and 94% respectively when compared to control
electroporations. Restriction-site mutation analysis (RSM) and Southern blot analysis
indicated the enzymatic degradation of GUS coding sequence by the REs HaeIII and Sau3AI.
Results of this study suggest that on electroporation, REs can enter into plant cells and
alter the expression of the GUS gene. The alteration of gene expression is thus correlated
with the digestion of GUS template DNA. Future applications of this technique could include
addressing fundamental questions with regard to DNA repair, site-specific recombination,
identifying mutations, insertional mutagenesis, enhancement of stable transformation and gene
tagging in plants.

<>

<1>Ashworth, J., Havranek, J.J., Duarte, C.M., Sussman, D., Monnat, R.J., Stoddard, B.L., Baker, D.
<2>Computational redesign of endonuclease DNA binding and cleavage specificity.
<3>Nature
<4>441
<5>656-659
<6>2006
<7>The reprogramming of DNA-binding specificity is an important challenge for computational
protein design that tests current understanding of
protein - DNA recognition, and has considerable practical relevance for
biotechnology and medicine(1-6). Here we describe the computational
redesign of the cleavage specificity of the intron-encoded homing
endonuclease I-MsoI(7) using a physically realistic atomic-level
forcefield(8,9). Using an in silico screen, we identified single
base-pair substitutions predicted to disrupt binding by the wild-type
enzyme, and then optimized the identities and conformations of clusters
of amino acids around each of these unfavourable substitutions using
Monte Carlo sampling(10). A redesigned enzyme that was predicted to
display altered target site specificity, while maintaining wild-type
binding affinity, was experimentally characterized. The redesigned
enzyme binds and cleaves the redesigned recognition site similar to
10,000 times more effectively than does the wild-type enzyme, with a
level of target discrimination comparable to the original endonuclease.
Determination of the structure of the redesigned nuclease-recognition
site complex by X-ray crystallography confirms the accuracy of the
computationally predicted interface. These results suggest that
computational protein design methods can have an important role in the
creation of novel highly specific endonucleases for gene therapy and
other applications.

<>

<1>Ashworth, J., Taylor, G.K., Havranek, J.J., Quadri, S.A., Stoddard, B.L., Baker, D.
<2>Computational reprogramming of homing endonuclease specificity at multiple adjacent base pairs.
<3>Nucleic Acids Res.
<4>38
<5>5601-5608
<6>2010
<7>Site-specific homing endonucleases are capable of inducing gene conversion via homologous
recombination. Reprogramming their cleavage specificities
allows the targeting of specific biological sites for gene correction or
conversion. We used computational protein design to alter the cleavage
specificity of I-MsoI for three contiguous base pair substitutions,
resulting in an endonuclease whose activity and specificity for its new
site rival that of wild-type I-MsoI for the original site. Concerted
design for all simultaneous substitutions was more successful than a
modular approach against individual substitutions, highlighting the
importance of context-dependent redesign and optimization of protein-DNA
interactions. We then used computational design based on the crystal
structure of the designed complex, which revealed significant
unanticipated shifts in DNA conformation, to create an endonuclease that
specifically cleaves a site with four contiguous base pair substitutions.
Our results demonstrate that specificity switches for multiple concerted
base pair substitutions can be computationally designed, and that
iteration between design and structure determination provides a route to
large scale reprogramming of specificity.

<>

<1>Assis-das-Gracas, D., Thiago-Juca-Ramos, R., Vieira-Araujo, A.C., Zahlouth, R., Ribeiro-Carneiro, A., Souza-Lopes, T., Azevedo-Barauna, R., Azevedo, V., Cruz-Schneider, M.P., Pellizari, V.H., Silva, A.
<2>Complete Genome of a Methanosarcina mazei Strain Isolated from Sediment Samples from an Amazonian Flooded Area.
<3>Genome Announcements
<4>1
<5>e00271-13
<6>2013
<7>Methanosarcina mazei is a strictly anaerobic methanogen from the Methanosarcinales order,
which is known for its broad catabolic range among
methanogens and is widespread throughout diverse environments. The draft genome of the strain
presented here was cultivated from sediment samples collected from the Tucurui hydroelectric
power station reservoir.

<>

<1>Atanasiu, C., Byron, O., McMiken, H., Sturrock, S.S., Dryden, D.T.F.
<2>Characterisation of the structure of ocr, the gene 0.3 protein of bacteriophage T7.
<3>Nucleic Acids Res.
<4>29
<5>3059-3068
<6>2001
<7>The product of gene 0.3 of bacterlophage T7, ocr, is a potent inhibitor of type I DNA
restriction and modification enzymes. We have used
biophysical methods to examine the mass, stability, shape and surface
charge distribution of ocr. Ocr is a dimeric protein with hydrodynamic
behaviour equivalent to a prolate ellipsoid of axial ratio 4.3 +/-
0.7:1 and mass of 27 kDa. The protein is resistant to denaturation but
removal of the C-terminal region reduces stability substantially. Six
amino acids, N4, D25, N43, D62, S68 and W94, are all located on the
surface of the protein and N4 and S68 are also located at the interface
between the two 116 amino acid monomers. Negatively charged amino acid
side chains surround W94 but these side chains are not part of the
highly acidic C-terminus after W94. Ocr is able to displace a short DNA
duplex from the binding site of a type I enzyme with a dissociation
constant of the order of 100 pM or better. These results suggest that
ocr is of a suitable size and shape to effectively block the DNA
binding site of a type I enzyme and has a large negatively charged
patch on its surface. This charge distribution may be complementary to
the charge distribution within the DNA binding site of type I DNA
restriction and modification enzymes.

<>

<1>Atanasiu, C., Su, T.-J., Sturrock, S.S., Dryden, D.T.F.
<2>Interaction of the ocr gene 0.3 protein of bacteriophage T7 with EcoKI restriction/modification enzyme.
<3>Nucleic Acids Res.
<4>30
<5>3936-3944
<6>2002
<7>The ocr protein, the product of gene 0.3 of bacteriophage T7, is a structural mimic of the
phosphate backbone of B-form DNA. In total it mimics 22 phosphate groups over 24 bp of DNA.
This mimicry allows it to block DNA binding by type I DNA restriction enzymes and to inhibit
these enzymes. We have determined that multiple ocr dimers can bind stoichiometrically to the
archetypal type I enzyme, EcoKI. One dimer binds to the core methyltransferase and two to the
complete bifunctional restriction and modification enzyme. Ocr can also bind to the component
subunits of EcoKI. Binding affinity to the methyltransferase core is extremely strong with a
large favourable enthalpy change and an unfavourable entropy change. This strong interaction
prevents the dissociation of the methyltransferase which occurs upon dilution of the enzyme.
This stabilisation arises because the interaction appears to involve virtually the entire
surface area of ocr and leads to the enzyme completely wrapping around ocr.

<>

<1>Athanasiadis, A., Gregoriu, M., Thanos, D., Kokkinidis, M., Papamatheakis, J.
<2>Complete nucleotide sequence of the PvuII restriction enzyme gene from Proteus vulgaris.
<3>Nucleic Acids Res.
<4>18
<5>6434
<6>1990
<7>None

<>

<1>Athanasiadis, A., Kokkinidis, M.
<2>Purification, crystallization and preliminary X-ray diffraction studies of the PvuII endonuclease.
<3>J. Mol. Biol.
<4>222
<5>451-453
<6>1991
<7>The PvuII endonuclease (PvuIIR) is a restriction enzyme from a type II
restriction-modification system of Proteus vulgaris coded on plasmid pPvu1.
The protein recognizes the DNA sequence 5' CAG'CTG 3' and shows no sequence
homology to other restriction enzymes.  This makes PvuIIR an interesting
subject for structural determination.  A purification procedure was developed
that yields milligram quantitites of the PvuIIR from plasmids expressed in the
Escherichia coli strain HB101.  The protein was crystallized using ammonium
sulphate as precipitant.  The crystals are orthorhombic, space group P2(1)2(1)2
with cell dimensions: a=84.2 Angstrom, b=106.2 Angstrom, c=46.9 Angstrom.  The
asymmetric unit contains one PvuIIR dimer.  Diffraction extends to 2.3
Angstrom, so the crystals may permit structural determination at atomic
resolution.

<>

<1>Athanasiadis, A., Papanikolau, Y., Rina, M., Papadovasilaki, M., Dauter, Z., Petratos, K., Bouriotis, V., Kokkindis, M.
<2>Purification, crystallization and preliminary x-ray analysis of the M.BseCI DNA methyltransferase from Bacillus stearothermophilus.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>53
<5>477-479
<6>1997
<7>The DNA methyltransferase M.BseCI from B. stearothermophilus methylates the N6 atom of the 3'
adenine in the sequence 5'-ATCGAT-3'.  The 579-residue protein has been isolated and
crystallized using seeding and microdialysis techniques.  The crystals are monoclinic, space
group P21 with cell dimensions a=53.7, b=85.7, c=151.8A and b=95.1o, two molecules in the
asymmetric unit and diffract to at least 2.5A resolution.

<>

<1>Athanasiadis, A., Vlassi, M., Kotsifaki, D., Tucker, P.A., Wilson, K.S., Kokkinidis, M.
<2>Crystal structure of PvuII endonuclease reveals extensive structural homologies to EcoRV.
<3>Nat. Struct. Biol.
<4>1
<5>469-475
<6>1994
<7>The crystal structure of the dimeric PvuII restriction endonuclease (R.PvuII) has been
determined at a resolution of 2.4 angstroms. The protein has a mixed a/b architecture and
consists of two subdomains. Despite a lack of sequence homology, extensive structural
similarities exist between one R.PvuII subdomain and the DNA-binding subdomain of EcoRV
endonuclease (R.EcoRV); the dimerization subdomains are unrelated. Within the similar domains,
flexible segments of R.PvuII are topologically equivalent to the DNA-binding turns of R.EcoRV;
potential catalytic residues can be deduced from the structural similarities to R.EcoRV.
Conformational flexibility is important for the interaction with DNA. A possible
classification of endonuclease structures on the basis of the positions of the scissile
phosphates is discussed.

<>

<1>Attie, O., Jayaprakash, A., Shah, H., Paulsen, I.T., Morino, M., Takahashi, Y., Narumi, I., Sachidanandam, R., Satoh, K., Ito, M., Krulwich, T.A.
<2>Draft Genome Sequence of Bacillus alcalophilus AV1934, a Classic Alkaliphile Isolated from Human Feces in 1934.
<3>Genome Announcements
<4>2
<5>e01175-14
<6>2014
<7>Bacillus alcalophilus AV1934, isolated from human feces, was described in 1934 before
microbiome studies and recent indications of novel potassium ion coupling
to motility in this extremophile. Here, we report draft sequences that will
facilitate an examination of whether that coupling is part of a larger cycle of
potassium ion-coupled transporters.

<>

<1>Attwood, G.T., Kelly, W.J., Altermann, E.H., Leahy, S.C.
<2>Analysis of the Methanobrevibacter ruminantium draft genome: understanding methanogen biology to inhibit their action in the rumen.
<3>Aust. J. Exp. Agr.
<4>48
<5>83-88
<6>2008
<7>Methane is produced in the foregut (rumen) of ruminants by methanogens, which act as terminal
reducers of carbon in the rumen system. The
multistep methanogenesis pathway is well elucidated, mainly from the
study of non-rumen methanogens, but the adaptations that allow
methanogens to grow and persist in the rumen are not well understood.
The Pastoral Greenhouse Gas Research Consortium is sequencing the
genome of Methanobrevibacter ruminantium, a prominent methanogen in New
Zealand ruminants, as part of a project to mitigate greenhouse gases.
The genome is similar to 3.0Mb in size with a guanine-cytosine (GC)
content of 33.68%. All of the components of the methanogenesis pathway
have been identified and comparison of these gene sequences with those
from Methanothermobacter thermoautotrophicus and Methanosphaera
stadtmanae indicates that methanogenesis gene organisation is conserved
within the Methanobacteriales. The genome of M. ruminantium contains a
prophage sequence (designated phi mru) with distinct functional modules
encoding phage integration, DNA replication and packaging, capsid
proteins and lysis functions. A low GC region found at the distal end
of the phage sequence harbours a putative DNA restriction/modification
system which might provide additional protection against foreign DNA.
The genome also contains many large surface proteins with
characteristics that indicate that they may mediate association with
other rumen microbes. Approximately half of the genes identified within
the genome have no known function. Determining the function of these
new genes will assist in de. ning the role of M. ruminantium in methane
formation in the rumen and help identify means to control methane
emissions from ruminant animals.

<>

<1>Atyame, C.M., Delsuc, F., Pasteur, N., Weill, M., Duron, O.
<2>Diversification of Wolbachia endosymbiont in the Culex pipiens mosquito.
<3>Mol. Biol. Evol.
<4>28
<5>2761-2772
<6>2011
<7>The alpha-proteobacteria Wolbachia are among the most common intracellular bacteria and have
recently emerged as important drivers of arthropod biology. Wolbachia commonly act as
reproductive parasites in arthropods by inducing cytoplasmic incompatibility (CI), a type of
conditional sterility between hosts harboring incompatible infections. In this study, we
examined the evolutionary histories of Wolbachia infections, known as wPip, in the common
house mosquito Culex pipiens, which exhibits the greatest variation in CI crossing patterns
observed in any insect. We first investigated a panel of twenty wPip strains for their genetic
diversity through a multi-locus scheme combining thirteen Wolbachia genes. Because Wolbachia
depend primarily on maternal transmission for spreading within arthropod populations, we also
studied the variability in the co-inherited Cx. pipiens mitochondria. In total, we identified
fourteen wPip haplotypes, which all share a monophyletic origin and clearly cluster into five
distinct wPip groups. The diversity of Cx. pipiens mitochondria was extremely reduced, which
is likely a consequence of cytoplasmic hitchhiking driven by a unique and recent Wolbachia
invasion. Phylogenetic evidence indicates that wPip infections and mitochondrial DNA have
co-diverged through stable co-transmission within the cytoplasm and shows that a rapid
diversification of wPip has occurred. The observed pattern demonstrates that a considerable
degree of Wolbachia diversity can evolve within a single host species over short evolutionary
periods. In addition, multiple signatures of recombination were found in most wPip genomic
regions, leading us to conclude that the mosaic nature of wPip genomes may play a key role in
their evolution.

<>

<1>Atyame, C.M., Duron, O., Tortosa, P., Pasteur, N., Fort, P., Weill, M.
<2>Multiple Wolbachia determinants control the evolution of cytoplasmic incompatibilities in Culex pipiens mosquito populations.
<3>Mol. Ecol.
<4>20
<5>286-298
<6>2011
<7>Wolbachia are maternally inherited endosymbionts that can invade arthropod
populations through manipulation of their reproduction. In mosquitoes,
Wolbachia induce embryonic death, known as cytoplasmic incompatibility
(CI), whenever infected males mate with females either uninfected or
infected with an incompatible strain. Although genetic determinants of CI
are unknown, a functional model involving the so-called mod and resc
factors has been proposed. Natural populations of Culex pipiens mosquito
display a complex CI relationship pattern associated with the highest
Wolbachia (wPip) genetic polymorphism reported so far. We show here that
C. pipiens populations from La Reunion, a geographically isolated island
in the southwest of the Indian Ocean, are infected with genetically
closely related wPip strains. Crossing experiments reveal that these
Wolbachia are all mutually compatible. However, crosses with genetically
more distant wPip strains indicate that Wolbachia strains from La Reunion
belong to at least five distinct incompatibility groups (or crossing
types). These incompatibility properties which are strictly independent
from the nuclear background, formally establish that in C. pipiens, CI is
controlled by several Wolbachia mod/resc factors.

<>

<1>Auad, L., Peril, M.A.A., de Ruiz Holgado, A.A.P., Raya, R.R.
<2>Evidence of a restriction/modification system in Lactobacillus delbrueckii subsp. lactis CNRZ 326.
<3>Curr. Microbiol.
<4>36
<5>271-273
<6>1998
<7>Lactobacillus delbrueckii subsp. lactis (Lb. lactis) CNRZ 326 is widely used in the
propagation of Lb. delbrueckii bacteriophages.  In this study, evidence is presented that this
strain possesses a restriction-modification system. The mitomycin C-induced temperate
bacteriophage lb539 has a reduced efficiency of plaquing (EOP) on CNRZ 326 cells (EOP=10^-3),
but after several passages on this strain, or on the indicator strain Lb. lactis LKT, the
recovered phages (phages lb539.326 and lb539.LKT) have an EOP equal to 1.  Restrictive
development on CNRZ 326 was also observed after phage lb539.326 was propagated on the strain
Lb. lactis CRL 934.  The R/M system was also active against the virulent Lb. delbrueckii phage
ll-h.  Plasmid DNA was not detected in CNRZ 326, which suggests that the R/M system described
is chromosomally encoded.

<>

<1>Aubert, E., Spurgeon, S., Ray, W., Davies, J.
<2>Purification and characterization of an isoschizomer of SphI from Bacillus circulans.
<3>Nucleic Acids Res.
<4>18
<5>6152
<6>1990
<7>None

<>

<1>Aubol, B.E., Reich, N.O.
<2>Exploring the N-terminal function of murine methyltransferase.
<3>ACS Abstracts
<4>217
<5>453
<6>1999
<7>The mammalian DNA methyltransferase methylates specific cytosine residues on the host genome.
DNA methylation is essential for development, gene regulation, and aberrant DNA methylation
leads to tumorgenesis.  We are using a highly homogeneous preparation of the murine DCMTase to
gain basic insights into its function, and the design of novel anti-cancer strategies.
DCMTase contains a large, poorly characterized N-terminal domain not found in bacterial
enzymes that have similar functions.  This domain binds zinc with characteristics of a zinc
finger.  We have probed the affect of zinc binding on numerous aspects of DCMTase functions.
Methylation spreading, activation by adjacent 5-methylcytosine residues with single stranded
DNA as a substrate, is one of these functions, and is of particular interest.  This activation
may lead to the process of "methylation spreading" and our approach will determine if the zinc
plays a role in this regulation of enzyme function.

<>

<1>Aubol, B.E., Reich, N.O.
<2>Murine DNA cytosine C5-methyltransferase: in vitro studies of de novo methylation spreading.
<3>Biochem. Biophys. Res. Commun.
<4>310
<5>209-214
<6>2003
<7>The preference of murine DNA (cytosine-5)-methyltransferase (Dnmt1) for single stranded DNA
substrates is increased up to 50-fold by the presence
of a proximal 5-methyl cytosine (5(me)C). This modulation is
distance-dependent and is due to an enhanced binding affinity and minor
changes in catalytic efficiency. No modulation was observed with double
stranded DNA. Modulation requires that the 5(me)C moiety be attached to
the DNA strand containing the CpG methylation target. Our results support
a model in which 5(me)C binding by the enzyme occurs to at least one site
outside the region involved in CpG recognition. No modulation in response
to 5(me)C is observed with the bacterial enzyme M.SssI, which lacks the
large N-terminal regulatory domain found in Dnmt1. We suggest that this
allosteric modulation involves the N-terminal domain of Dnmt1.

<>

<1>Audic, S., Lescot, M., Claverie, J.M., Cloeckaert, A., Zygmunt, M.S.
<2>The genome sequence of Brucella pinnipedialis B2/94 sheds light on the evolutionary history of the genus Brucella.
<3>BMC Evol. Biol.
<4>11
<5>200
<6>2011
<7>ABSTRACT: BACKGROUND: Since the discovery of the Malta fever agent,
Brucella melitensis, in the 19th century, six terrestrial
mammal-associated Brucella species were recognized over the next century.
More recently the number of novel Brucella species has increased and among
them, isolation of species B. pinnipedialis and B. ceti from marine
mammals raised many questions about their origin as well as on the
evolutionary history of the whole genus. RESULTS: We report here on the
first complete genome sequence of a Brucella strain isolated from marine
mammals, Brucella pinnipedialis strain B2/94. A whole gene-based
phylogenetic analysis shows that five main groups of host-associated
Brucella species rapidly diverged from a likely free-living ancestor close
to the recently isolated B. microti. However, this tree lacks the
resolution required to resolve the order of divergence of those groups.
Comparative analyses focusing on a) genome segments unshared between B.
microti and B. pinnipedialis, b) gene deletion/fusion events and c)
positions and numbers of Brucella specific IS711 elements in the available
Brucella genomes provided enough information to propose a branching order
for those five groups. CONCLUSIONS: In this study, it appears that the
closest relatives of marine mammal Brucella sp. are B. ovis and Brucella
sp. NVSL 07-0026 isolated from a baboon, followed by B. melitensis and B.
abortus strains, and finally the group consisting of B. suis strains,
including B. canis and the group consisting of the single B. neotomae
species. We were not able, however, to resolve the order of divergence of
the two latter groups.

<>

<1>Audic, S., Lescot, M., Claverie, J.M., Scholz, H.C.
<2>Brucella microti: the genome sequence of an emerging pathogen.
<3>BMC Genomics
<4>10
<5>352
<6>2009
<7>ABSTRACT: BACKGROUND: Using a combination of pyrosequencing and
conventional Sanger sequencing, the complete genome sequence of the
recently described novel Brucella species, Brucella microti, was
determined. B. microti is a member of the genus Brucella within the
Alphaproteobacteria, which consists of medically important highly
pathogenic facultative intracellular bacteria. In contrast to all other
Brucella species, B. microti is a fast growing and biochemically very
active microorganism with a phenotype more similar to that of
Ochrobactrum, a facultative human pathogen. The atypical phenotype of B.
microti prompted us to look for genomic differences compared to other
Brucella species and to look for similarities with Ochrobactrum. RESULTS:
The genome is composed of two circular chromosomes of 2,117,050 and
1,220,319 base pairs. Unexpectedly, we found that the genome sequence of
B. microti is almost identical to that of Brucella suis 1330 with an
overall sequence identity of 99.84% in aligned regions. The most
significant structural difference between the two genomes is a
bacteriophage-related 11,742 base pairs insert only present in B. microti.
However, this insert is unlikely to have any phenotypical consequence.
Only four protein coding genes are shared between B. microti and
Ochrobactrum anthropi but impaired in other sequenced Brucella. The most
noticeable difference between B. microti and other Brucella species was
found in the sequence of the 23S ribosomal RNA gene. This unusual
variation could have pleiotropic effects and explain the fast growth of B.
microti. CONCLUSIONS: Contrary to expectations from the phenotypic
analysis, the genome sequence of B. microti is highly similar to that of
known Brucella species, and is remotely related to the one of O. anthropi.
How the few differences in gene content between B. microti and B. suis
1330 could result in vastly different phenotypes remains to be elucidated.
This unexpected finding will complicate the task of identifying virulence
determinants in the Brucella genus. The genome sequence of B. microti will
serve as a model for differential expression analysis and complementation
studies. Our results also raise some concerns about the importance given
to phenotypical traits in the definition of bacterial species.

<>

<1>Auer, B., Schweiger, M.
<2>Evidence that Escherichia coli virus T1 induces a DNA methyltransferase.
<3>J. Virol.
<4>49
<5>588-590
<6>1984
<7>DNA of Escherichia coli virus T1 is resistant to MboI cleavage and appears
to be heavily methylated.  Analysis of methylation by the isoschizomeric restriction
enzymes Sau3AI and DpnI revealed that recognition sites for E. coli DNA adenine
methylase (dam methylase) are methylated.  The same methylation pattern was found for
virus T1 DNA grown on an E. coli dam host, indicating a T1-specific DNA
methyltransferase.

<>

<1>Auger, S., Galleron, N., Segurens, B., Dossat, C., Bolotin, A., Wincker, P., Sorokin, A.
<2>Complete Genome Sequence of the Highly Hemolytic Strain Bacillus cereus F837/76.
<3>J. Bacteriol.
<4>194
<5>1630
<6>2012
<7>Highly hemolytic strain Bacillus cereus F837/76 was isolated in 1976 from a contaminated
prostate wound. The complete nucleotide sequence of this strain
reported here counts nearly 36,500 single-nucleotide differences from the closest
sequenced strain, Bacillus thuringiensis Al Hakam. F827/76 also contains a 10-kb
plasmid that was not detected in the Al Hakam strain.

<>

<1>Aujame, L., Bouchardon, A., Renauld-Mongenie, G., Rokbi, B., Nassif, X., Tinsley, C., Perrin, A.
<2>Specific nucleic acids and polypeptide from pathogenic strains ofNeisseria.
<3>European Patent Office
<4>EP 1475441 A
<5>
<6>2004
<7>Cette invention concerne des acides nucleiques codant pour les polypeptides specifiques des
souches pathogenes du genre Neisseria, les polypeptides correspondants, et leurs applications
diagnostiques et therapeutiques.

<>

<1>Aujame, L., Bouchardon, A., Renauld-Mongenie, G., Rokbi, B., Nassif, X., Tinsley, C., Perrin, A.
<2>Specific nucleic acids and polypeptide from pathogenic strains ofNeisseria.
<3>European Patent Office
<4>EP 1724352 A
<5>
<6>2006
<7>
<>

<1>Aujame, L., Bouchardon, A., Renauld-Mongenie, G., Rokbi, B., Nassif, X., Tinsley, C., Perrin, A.
<2>Specific polypeptides and nucleic acids of pathogenic strains of the Neisseria genus.
<3>European Patent Office
<4>EP 2100960 A
<5>
<6>2009
<7>Cette invention concerne des acides nucleiques odant pour les polypeptides specifiques des
souches pathogenes du genre Neisseria, les polypeptides correspondants, et leurs applications
diagnostiques et therapeutiques.

<>

<1>Avasthi, T.S., Devi, S.H., Taylor, T.D., Kumar, N., Baddam, R., Kondo, S., Suzuki, Y., Lamouliatte, H., Megraud, F., Ahmed, N.
<2>Genomes of the two chronological isolates (Helicobacter pylori 2017 and 2018) of the West African Helicobacter pylori strain 908, obtained from a  single patient.
<3>J. Bacteriol.
<4>193
<5>3385-3386
<6>2011
<7>The diverse clinical outcomes of colonization by Helicobacter pylori reflect need to
understand genomic rearrangements enabling the bacterium
to adapt to host niches and exhibit varied colonization/virulence
potential. We describe genome sequences of the two serial isolates, H.
pylori 2017 and 2018 (the chronological sub-clones of H. pylori 908),
cultured in 2003 from corpus and antrum, respectively, of an African
patient who suffered from recrudescent duodenal ulcer disease. The genome
sequences when compared with the genome of parent strain, HP 908 (isolated
from antrum of the same patient in 1994) revealed genomic alterations
relevant in virulence optimization or host-specific adaptation.

<>

<1>Avasthi, T.S., Kumar, N., Baddam, R., Hussain, A., Nandanwar, N., Jadhav, S., Ahmed, N.
<2>Genome of Multidrug-Resistant Uropathogenic Escherichia coli Strain NA114 from India.
<3>J. Bacteriol.
<4>193
<5>4272-4273
<6>2011
<7>Uropathogenic Escherichia coli (UPEC) causes serious infections in people at risk and has a
significant environmental prevalence due to
contamination by human and animal excreta. In developing countries, UPEC
assumes importance in certain dwellings because of poor community/personal
hygiene and exposure to contaminated water or soil. We report the complete
genome sequence of E. coli strain NA114 from India, a UPEC strain with a
multidrug resistance phenotype and the capacity to produce
extended-spectrum beta-lactamase. The genome sequence and comparative
genomics emanating from it will be significant in under-standing the
genetic makeup of diverse UPEC strains and in boosting the development of
new diagnostics/vaccines.

<>

<1>Avendano-Herrera, R., Suarez, R., Lazo, E., Bravo, D., Llegues, K.O., Romalde, J.L., Godoy, M.G.
<2>Genome Sequence of Streptococcus phocae subsp. salmonis Strain C-4T, Isolated from Atlantic Salmon (Salmo salar).
<3>Genome Announcements
<4>2
<5>e01269-14
<6>2014
<7>Streptococcus phocae subsp. salmonis is a fish pathogen that has an important impact on the
Chilean salmon industry. Here, we report the genome sequence of the
type strain C-4(T) isolated from Atlantic salmon (Salmo salar), showing a number
of interesting features and genes related to its possible virulence factors.

<>

<1>Aw, Y.K., Ong, K.S., Yule, C.M., Gan, H.M., Lee, S.M.
<2>Draft Genome Sequence of Paenibacillus sp. Strain MSt1 with Broad Antimicrobial Activity, Isolated from Malaysian Tropical Peat Swamp Soil.
<3>Genome Announcements
<4>2
<5>e01024-14
<6>2014
<7>We report the draft genome sequence of Paenibacillus sp. strain MSt1, which has broad-range
antimicrobial activity, isolated from tropical peat swamp soil. Genes
involved in antimicrobial biosynthesis are found to be present in this genome.

<>

<1>Axelsson, L., Rud, I., Naterstad, K., Blom, H., Renckens, B., Boekhorst, J., Kleerebezem, M., van Hijum, S., Siezen, R.J.
<2>Genome Sequence of the Naturally Plasmid-Free Lactobacillus plantarum Strain NC8  (CCUG 61730).
<3>J. Bacteriol.
<4>194
<5>2391-2392
<6>2012
<7>Lactobacillus plantarum is a highly versatile lactic acid bacterium found in various
ecological niches, such as fermented vegetable, meat, and dairy products
and the gastrointestinal tract. We sequenced the genome of L. plantarum NC8, a
naturally plasmid-free strain, which has been used as a model strain in many
laboratories worldwide.

<>

<1>Ayano, H., Kuroda, M., Soda, S., Ike, M.
<2>Draft Genome Sequence of Pseudomonas aeruginosa Strain RB, a Bacterium Capable of Synthesizing Cadmium Selenide Nanoparticles.
<3>Genome Announcements
<4>2
<5>e00368-14
<6>2014
<7>Pseudomonas aeruginosa strain RB is a bacterium capable of synthesizing cadmium selenide
(CdSe) nanoparticles and was isolated from a soil sample. Here, we
present the draft genome sequence of P. aeruginosa strain RB. To the best of our
knowledge, this is the first report of a draft genome of a CdSe-synthesizing
bacterium.

<>

<1>Ayarza, J.M., Figuerola, E.L., Erijman, L.
<2>Draft Genome Sequences of Type Strain Sediminibacterium salmoneum NJ-44 and Sediminibacterium sp. Strain C3, a Novel Strain Isolated from Activated Sludge.
<3>Genome Announcements
<4>2
<5>e01073-13
<6>2014
<7>The genus Sediminibacterium comprises species present in diverse natural and engineered
environments. Here, we report for the first time the genome sequences
of the type strain Sediminibacterium salmoneum NJ-44 (NBRC 103935) and
Sediminibacterium sp. strain C3 (BNM541), isolated from activated sludge, a
valuable model for the study of substrate-dependent autoaggregation.

<>

<1>Aylward, F.O. et al.
<2>Complete Genome of Enterobacteriaceae Bacterium Strain FGI 57, a Strain Associated with Leaf-Cutter Ant Fungus Gardens.
<3>Genome Announcements
<4>1
<5>e00238-12
<6>2013
<7>The Enterobacteriaceae bacterium strain FGI 57 was isolated from a fungus garden  of the
leaf-cutter ant Atta colombica. Analysis of its single 4.76-Mbp chromosome
will shed light on community dynamics and plant biomass degradation in ant fungus
gardens.

<>

<1>Aylward, F.O. et al.
<2>Complete Genome of Serratia sp. Strain FGI 94, a Strain Associated with Leaf-Cutter Ant Fungus Gardens.
<3>Genome Announcements
<4>1
<5>e0023912
<6>2013
<7>Serratia sp. strain FGI 94 was isolated from a fungus garden of the leaf-cutter ant Atta
colombica. Analysis of its 4.86-Mbp chromosome will help advance our
knowledge of symbiotic interactions and plant biomass degradation in this ancient
ant-fungus mutualism.

<>

<1>Azarinskas, A., Maneliene, Z., Jakubauskas, A.
<2>Hin4II, a new prototype restriction endonuclease from Haemophilus influenzae RFL4: Discovery, cloning and expression in Escherichia coli.
<3>J. Biotechnol.
<4>123
<5>288-296
<6>2006
<7>The genes encoding restriction-modification system of unknown specificity Hin4II from
Haemophilus influenzae RFL4 were cloned in
Escherichia coli and sequenced. The Hin4II system comprises three
tandemly arranged genes coding for m6A DNA methyltransferase, m5C DNA
methyltransferase and restriction endonuclease, respectively.
Restriction endonuclease was expressed in E. coli and purified to
apparent homogeneity. The DNA recognition sequence and cleavage
positions were determined. R.Hin4II recognizes the novel
non-palindromic sequence 5'-CCTTC-3' and cleaves the DNA 6 and 5 nt
downstream in the top and bottom strand, respectively. The new
prototype restriction endonuclease Hin4II was classified as a potential
candidate of HNH nuclease family after comparison against SMART
database. An amino acid sequence motif 297H-X-14-N-X-8-H of Hin4II was
proposed as forming a putative catalytic center.

<>

<1>Azcarate-Peril, M.A., Altermann, E., Goh, Y.J., Tallon, R., Sanozky-Dawes, R.B., Pfeiler, E.A., O'Flaherty, S., Buck, B.L., Dobson, A., Duong, T., Miller, M.J., Barrangou, R., Klaenhammer, T.R.
<2>Analysis of the genome sequence of Lactobacillus gasseri ATCC 33323 reveals the molecular basis of an autochthonous intestinal organism.
<3>Appl. Environ. Microbiol.
<4>74
<5>4610-4625
<6>2008
<7>This study presents the complete genome sequence of Lactobacillus gasseri ATCC 33323, a
neotype strain of human origin and a native
species found commonly in the gastrointestinal tracts of neonates and
adults. The plasmid-free genome was 1,894,360 bp in size and predicted
to encode 1,810 genes. The GC content was 35.3%, similar to the GC
content of its closest relatives, L. johnsonii NCC 533 (34%) and L.
acidophilus NCFM (34%). Two identical copies of the prophage LgaI
(40,086 bp), of the Sfi11-like Siphoviridae phage family, were
integrated tandomly in the chromosome. A number of unique features were
identified in the genome of L. gasseri that were likely acquired by
horizontal gene transfer and may contribute to the survival of this
bacterium in its ecological niche. L. gasseri encodes two restriction
and modification systems, which may limit bacteriophage infection. L.
gasseri also encodes an operon for production of heteropolysaccharides
of high complexity. A unique alternative sigma factor was present
similar to that of B. caccae ATCC 43185, a bacterial species isolated
from human feces. In addition, L. gasseri encoded the highest number of
putative mucus-binding proteins (14) among lactobacilli sequenced to
date. Selected phenotypic characteristics that were compared between
ATCC 33323 and other human L. gasseri strains included carbohydrate
fermentation patterns, growth and survival in bile,,oxalate
degradation, and adhesion to intestinal epithelial cells, in vitro. The
results from this study indicated high intraspecies variability from a
genome encoding traits important for survival and retention in the
gastrointestinal tract.

<>

<1>Azeddoug, H., Hubert, J., Reysset, G.
<2>Characterization of a methyl-specific restriction system in Clostridium acetobutylicum strain N1-4081.
<3>FEMS Microbiol. Lett.
<4>65
<5>323-326
<6>1989
<7>A type II restriction endonuclease, named CacI, was detected in Clostridium acetobutylicum
strain N1-4081. CacI cleaved the tetranucleotide sequence [5'-GATC-3']. The modification
system consisted of the methylation of the adenine present in this sequence. CacI, an
isoschizomer of MboI, is inactive on dam methylated substrates.

<>

<1>Azeddoug, H., Hubert, J., Reysset, G.
<2>Restriction endonuclease in Clostridium acetobutylicum strain NI-4081.
<3>Clinical and Molecular Aspects of Anaerobes, Wrightson Biomedical Publishing Ltd., Borriello, S.P., Petersfield
<4>0
<5>249-250
<6>1990
<7>That a restriction-modification system may exist in Clostridium acetobutylicum
strain NI-4081 was suggested by the observation that the efficiency of plasmid
transformation was dependent upon the DNA source of the replicon, and by
difficulties encountered in gene cloning experiments with DNA of heterologous
Clostridia.  These findings led us to search for restriction endonuclease and
methylase activities in strain NI-4081.

<>

<1>Azeddoug, H., Reysset, G.
<2>Recognition sequence of a new methyl-specific restriction system from Clostridium acetobutylicum strain ABKn8.
<3>FEMS Microbiol. Lett.
<4>78
<5>153-156
<6>1991
<7>A new type II restriction endonuclease, named CacII was detected in Clostridium
acetobutylicum strain ABKn8.  CacII cleaved the hexanucleotide sequence
[5'-GCN^NGC-3'] and generated blunt ends.  Up to now no isoschizomer of CacII
has been described. Note that the name of this enzyme has been changed to
Cac8I.

<>

<1>Azeddoug, H., Reysset, G., Sebald, M.
<2>Characterization of restriction endonuclease BfrBI from Bacteroides fragilis strains BE3 and AIP 10006.
<3>FEMS Microbiol. Lett.
<4>95
<5>133-136
<6>1992
<7>A new type II restriction endonuclease, named BfrBI, was detected in two stains of Bacteroides
fragilis, BE3 and AIP 10006 (NCTC 9343T). The enzyme BfrBI, an isoschizomer of NsiI and Ava
III, recognized the hexanucleotide sequence [5'-ATG CAT-3'], with a cleavage site generating
blunt ends.

<>

<1>Azizbekyan, R.R., Rebentish, B.A., Netyksa, E.M.
<2>Bacillus thuringiensis restrictase sensitive to dam-methylation.
<3>Biotekhnologiya
<4>4
<5>197-198
<6>1988
<7>A new restriction enzyme from Bacillus thuringiensis is described. It is an isoschizomer of
ClaI and is sensitive to dam methylation.

<>

<1>Azizbekyan, R.R., Rebentish, B.A., Netyksa, E.M., Bychkova, M.A., Bolotin, A.P.
<2>Site-specific restriction endonuclease from Bacillus thuringiensis var. Kumantoenis.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>1-2
<5>13-15
<6>1992
<7>Site-specific restriction endonucleases are widely used in recombinant DNA technology for the
construction of various molecular vectors. Possessing different specificity toward definite
nucleotide sequences, they permit the construction of recombinant DNA molecules for their
subsequent introduction into cells and yet, the presence of restriction-modification systems
in the cells of the recipient bacteria limit the effectiveness of the introduction of foreign
DNA. Recent investigations on site-specific restriction endonucleases have been conducted
along two lines: the search for new restriction-modification systems and the search for
recipients lacking restriction endonucleases. The entoropathogenic bacteria Bacillus
thuringiensis, which produce a crystalline protein endotoxin, are finding wide use as
ecologically safe biological insecticides. Earlier we detected restriction endonucleases of
various specificity in the cells of strains of various B. thuringiensis serovariants. In the
course of genetic manipulations with B. thuringiensis (H18) strain 4W1, we found that this
strain limits the development of certain phages. In this work we studied the mechanism of the
limitation of phage development and isolated two site-specific endonucleases from cells of the
strain 4W1.

<>

<1>Azizbekyan, R.R., Rebentish, B.A., Stepanova, T.V., Netyksa, E.M., Bychkova, M.A.
<2>Site specific restriction endonuclease from a Bacillus thuringiensis strain.
<3>Dokl. Akad. Nauk.
<4>274
<5>742-744
<6>1984
<7>Site-specific restriction endonucleases are used in recombinant DNA methodology
for the construction of recombinant DNAs and the creation of strains that
produce biologically active compounds on the basis of them.  The second aspect
of the use of restriction endonucleases is due to the possibility of their use
for blocking the expression of foreign genetic information; in particular,
restriction systems can be introduced into strains that produce biologically
active compounds to increase their resistance to infection by bacteriophages.

<>

<1>Azorin, F., Hahn, R., Rich, A.
<2>Restriction endonucleases can be used to study B-Z junctions in supercoiled DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>81
<5>5714-5718
<6>1984
<7>Plasmids containing (C-G)n inserts have been used to study the inhibition of cleavage by
restriction endonucleases due to Z-DNA formation in negatively supercoiled plasmids.  The
enzyme BssHII, which recognizes G-C-G-C-G-C, is strongly inhibited when the insert forms
Z-DNA.  The BamHI recognition sequence (G-G-A-T-C-C) was placed in four different positions
near the B-Z junction and the inhibition of BamHI cleavage was determined as a function of
negative superhelical density.  Formation of Z-DNA in the (C-G)n insert inhibited cleavage by
BamHI when its recognition sequence was located immediately adjacent to the insert or four
base pairs away from it.  However, no inhibition was found when the BamHI recognition site was
eight base pairs away.  These experiments help to define the limits of the structural
perturbation associated with the B-Z junction.

<>

<1>Azuma, Y., Hosoyama, A., Matsutani, M., Furuya, N., Horikawa, H., Harada, T., Hirakawa, H., Kuhara, S., Matsushita, K., Fujita, N., Shirai, M.
<2>Whole-genome analyses reveal genetic instability of Acetobacter pasteurianus.
<3>Nucleic Acids Res.
<4>37
<5>5768-5783
<6>2009
<7>Acetobacter species have been used for brewing traditional vinegar and are
known to have genetic instability. To clarify the mutability, Acetobacter
pasteurianus NBRC 3283, which forms a multi-phenotype cell complex, was
subjected to genome DNA sequencing. The genome analysis revealed that
there are more than 280 transposons and five genes with hyper-mutable
tandem repeats as common features in the genome consisting of a 2.9-Mb
chromosome and six plasmids. There were three single nucleotide mutations
and five transposon insertions in 32 isolates from the cell complex. The
A. pasteurianus hyper-mutability was applied for breeding a
temperature-resistant strain grown at an unviable high-temperature (42
degrees C). The genomic DNA sequence of a heritable mutant showing
temperature resistance was analyzed by mutation mapping, illustrating that
a 92-kb deletion and three single nucleotide mutations occurred in the
genome during the adaptation. Alpha-proteobacteria including A.
pasteurianus consists of many intracellular symbionts and parasites, and
their genomes show increased evolution rates and intensive genome
reduction. However, A. pasteurianus is assumed to be a free-living
bacterium, it may have the potentiality to evolve to fit in natural niches
of seasonal fruits and flowers with other organisms, such as yeasts and
lactic acid bacteria.

<>

<1>Baar, C., Eppinger, M., Raddatz, G., Simon, J., Lanz, C., Klimmek, O., Nandakumar, R., Gross, R., Rosinus, A., Keller, H., Jagtap, P., Linke, B., Meyer, F., Lederer, H., Schuster, S.C.
<2>Complete genome sequence and analysis of Wolinella succinogenes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>11690-11695
<6>2003
<7>To understand the origin and emergence of pathogenic bacteria, knowledge of the genetic
inventory from their nonpathogenic relatives is a prerequisite.
Therefore, the 2.11-megabase genome sequence of Wolinella succinogenes, which is
closely related to the pathogenic bacteria Helicobacter pylori and Campylobacter
jejuni, was determined. Despite being considered nonpathogenic to its bovine
host, W. succinogenes holds an extensive repertoire of genes homologous to known
bacterial virulence factors. Many of these genes have been acquired by lateral
gene transfer, because part of the virulence plasmid pVir and an N-linked
glycosylation gene cluster were found to be syntenic between C. jejuni and
genomic islands of W. succinogenes. In contrast to other host-adapted bacteria,
W. succinogenes does harbor the highest density of bacterial sensor kinases found
in any bacterial genome to date, together with an elaborate signaling circuitry
of the GGDEF family of proteins. Because the analysis of the W. succinogenes
genome also revealed genes related to soil- and plant-associated bacteria such as
the nif genes, W. succinogenes may represent a member of the epsilon
proteobacteria with a life cycle outside its host.

<>

<1>Baba, T., Bae, T., Schneewind, O., Takeuchi, F., Hiramatsu, K.
<2>Genome sequence of Staphylococcus aureus strain Newman and comparative analysis of staphylococcal genomes: polymorphism and evolution of two major pathogenicity    islands.
<3>J. Bacteriol.
<4>190
<5>300-310
<6>2008
<7>Strains of Staphylococcus aureus, an important human pathogen, display up to 20%  variability
in their genome sequence, and most sequence information is available
for human clinical isolates that have not been subjected to genetic analysis of
virulence attributes. S. aureus strain Newman, which was also isolated from a
human infection, displays robust virulence properties in animal models of disease
and has already been extensively analyzed for its molecular traits of
staphylococcal pathogenesis. We report here the complete genome sequence of S.
aureus Newman, which carries four integrated prophages, as well as two large
pathogenicity islands. In agreement with the view that S. aureus Newman prophages
contribute important properties to pathogenesis, fewer virulence factors are
found outside of the prophages than for the highly virulent strain MW2. The
absence of drug resistance genes reflects the general antibiotic-susceptible
phenotype of S. aureus Newman. Phylogenetic analyses reveal clonal relationships
between the staphylococcal strains Newman, COL, NCTC8325, and USA300 and a
greater evolutionary distance to strains MRSA252, MW2, MSSA476, N315, Mu50, JH1,
JH9, and RF122. However, polymorphism analysis of two large pathogenicity islands
distributed among these strains shows that the two islands were acquired
independently from the evolutionary pathway of the chromosomal backbones of
staphylococcal genomes. Prophages and pathogenicity islands play central roles in
S. aureus virulence and evolution.

<>

<1>Baba, T., Kuwahara-Arai, K., Uchiyama, I., Takeuchi, F., Ito, T., Hiramatsu, K.
<2>Complete genome sequence of Macrococcus caseolyticus strain JSCS5402, reflecting the ancestral genome of the human-pathogenic staphylococci.
<3>J. Bacteriol.
<4>191
<5>1180-1190
<6>2009
<7>We isolated the methicillin-resistant Macrococcus caseolyticus strain JCSC5402 from animal
meat in a supermarket and determined its whole-genome
nucleotide sequence. This is the first report on the genome analysis of a
macrococcal species that is evolutionarily closely related to the human
pathogens Staphylococcus aureus and Bacillus anthracis. The essential
biological pathways of M. caseolyticus are similar to those of
staphylococci. However, the species has a small chromosome (2.1 MB) and
lacks many sugar and amino acid metabolism pathways and a plethora of
virulence genes that are present in S. aureus. On the other hand, M.
caseolyticus possesses a series of oxidative phosphorylation machineries
that are closely related to those in the family Bacillaceae. We also
discovered a probable primordial form of a Macrococcus methicillin
resistance gene complex, mecIRAm, on one of the eight plasmids harbored by
the M. caseolyticus strain. This is the first finding of a
plasmid-encoding methicillin resistance gene. Macrococcus is considered to
reflect the genome of ancestral bacteria before the speciation of
staphylococcal species and may be closely associated with the origin of
the methicillin resistance gene complex of the notorious human pathogen
methicillin-resistant S. aureus.

<>

<1>Baba, T., Takeuchi, F., Kuroda, M., Yuzawa, H., Aoki, K., Oguchi, A., Nagai, Y., Iwama, N., Asano, K., Naimi, T., Kuroda, H., Cui, L., Yamamoto, K., Hiramatsu, K.
<2>Genome and virulence determinants of high virulence community-acquired MRSA.
<3>Lancet
<4>359
<5>1819-1827
<6>2002
<7>BACKGROUND: A new type of meticillin-resistant Staphylococcus aureus (MRSA), designated
community-acquired MRSA, is becoming increasingly
noticeable in the community, some strains of which cause fatal infections
in otherwise healthy individuals. By contrast with hospital-acquired MRSA,
community-acquired MRSA is more susceptible to non beta-lactam antibiotics.
We investigated the high virulence potential of certain strains of this
bacterium. METHODS: We ascertained the whole genome sequence of MW2, a
strain of community-acquired MRSA, by shotgun cloning and sequencing. MW2
caused fatal septicaemia and septic arthritis in a 16-month-old girl in
North Dakota, USA, in 1998. The genome of this strain was compared with
those of hospital-acquired MRSA strains, including N315 and Mu50.
FINDINGS: Meticillin resistance gene (mecA) in MW2 was carried by a novel
allelic form (type IVa) of staphylococcal cassette chromosome mec
(SCCmec), by contrast with type II in N315 and Mu50. Type IVa SCCmec did
not carry any of the multiple antibiotic resistance genes reported in type
II SCCmec. By contrast, 19 additional virulence genes were recorded in the
MW2 genome. All but two of these virulence genes were noted in four of the
seven genomic islands of MW2. INTERPRETATION: MW2 carried a range of
virulence and resistance genes that was distinct from those displayed on
the chromosomes of extant S aureus strains. Most genes were carried by
specific allelic forms of genomic islands in the MW2 chromosome. The
combination of allelic forms of genomic islands is the genetic basis that
determines the pathogenicity of medically important phenotypes of S
aureus, including those of community-acquired MRSA strains.

<>

<1>Babcock, M.J., Schottel, J.L.
<2>SsbI, an isoschizomer of HindIII isolated from Streptomyces scabies.
<3>Nucleic Acids Res.
<4>19
<5>3457
<6>1991
<7>SsbI is a restriction endonuclease identified in cell lysates of the soil
bacterium Streptomyces scabies, isolate FL1.  SsbI was partially purified by
DEAE-Sephadex chromatography to remove contaminating nuclease activity.  The
partially purified SsbI fraction was used to determine the recognition sequence
and the cleavage site for the endonuclease.

<>

<1>Babic, A.C., Little, E.J., Manohar, V.M., Bitinaite, J., Horton, N.C.
<2>DNA Distortion and Specificity in a Sequence-Specific Endonuclease.
<3>J. Mol. Biol.
<4>383
<5>186-204
<6>2008
<7>Five new structures of the Q138F HincII enzyme bound to a total of three different DNA
sequences and three different metal ions (Ca(2+), Mg(2+),
and Mn(2+)) are presented. While previous structures were produced from
soaking Ca(2+) into preformed Q138F HincII/DNA crystals, the new
structures are derived from cocrystallization with Ca(2+), Mg(2+), or
Mn(2+). The Mn(2)(+)-bound structure provides the first view of a product
complex of Q138F HincII with cleaved DNA. Binding studies and a crystal
structure show how Ca(2+) allows trapping of a Q138F HincII complex with
noncognate DNA in a catalytically incompetent conformation. Many Q138F
HincII/DNA structures show asymmetry, despite the binding of a symmetric
substrate by a symmetric enzyme. The various complexes are fit into a
model describing the different conformations of the DNA-bound enzyme and
show how DNA conformational energetics determine DNA-cleavage rates by the
Q138F HincII enzyme.

<>

<1>Babinger, P., Volkl, R., Cakstina, I., Maftei, A., Schmitt, R.
<2>Maintenance DNA methyltransferase (Met1) and silencing of CpG-methylated foreign DNA in Volvox carteri.
<3>Plant Mol. Biol.
<4>63
<5>325-336
<6>2007
<7>DNA methylation plays an important role in the gene-silencing network of higher eukaryotes. We
have analyzed the 21.5-kb maintenance
methyltransferase (M-MTase) gene, met1, of the multicellular green alga
Volvox carteri. The met1 transcript was detected only during the period
when DNA replication and cell division are taking place. It encodes a
238 kDa protein containing eight C-terminal activity domains typical of
M-MTases, plus upstream DNA-binding domains including the ProDom domain
PD003757, which experimental analyses in animal systems have indicated
is required for targeting the enzyme to DNA-replication foci. Several
insertions of unknown function make Volvox Met1 the largest known
member of the Met1/Dnmt1 family. Here we also show that several
endogenous transposon families are CpG-methyated in Volvox, which we
think causes them to be inactive. This view is supported by the
observation that an in vitro CpG-methylated gene introduced into Volvox
was maintained in the methylated and silent state over > 100
generations. Thus, we believe that Met1 recognizes and perpetuates the
in vitro methylation signal, and that the silencing machinery is then
able to transduce such a methylation-only signal into a stable
heterochromatic (and silent) state.

<>

<1>Babkina, O.V., Chutko, C.A., Shashkov, A.A., Dzhidzhoev, M.S., Eritja, R., Gromova, E.S.
<2>Iodouracil-mediated photocrosslinking of DNA to EcoRII restriction endonuclease in catalytic conditions.
<3>Photochem. Photobiolog.
<4>1
<5>636-640
<6>2002
<7>We used a XeCl excimer laser with 50 ns pulses, a frequency of 0.3 Hz and a wavelength of 308
nm in appropriate conditions for the photocrosslinking
of EcoRII restriction endonuclease to a 14-mer DNA duplex, containing a
5-iodo-2'-deoxyuridine residue (IdU). IdU replaced the thymidine residue
within the EcoRII recognition sequence 5'-CCT/AGG. The binding of EcoRII
endonuclease to the IdU-containing DNA duplex was analyzed by gel
retardation assay in the presence of Ca2+ or Mg2+ ions. Photocrosslinking
of EcoRII to the IdU-containing DNA duplex occurred in a pre-reactive
complex formed in the presence of Ca2+ ions. Photocrosslinking yields as a
function of time and UV-laser light intensity were studied.

<>

<1>Babkina, O.V., Evstafieva, A.G., Chichkova, N.V., Vartapetian, A.B., Muller, S., Baskunov, V.B., Petrauskene, O.V., Kochetkov, S.N., Gromova, E.S.
<2>Recombinant components of the EcoRII restriction-modification system: Restriction endonuclease can interact with DNA-RNA duplexes.
<3>Mol. Biol. (Mosk)
<4>34
<5>1065-1073
<6>2000
<7>To obtain recombinant restriction endonuclease (R) and methylase (M) of the EcoRII
restriction-modification system, bacterial strains
overproducing their functional hexahistidine derivatives were
constructed. Active full-length EcoRII was produced only in cells
that also expressed M.EcoRII from a multicopy plasmid. Recombinant
EcoRII bound with hybrid DNA-RNA duplexes.

<>

<1>Babkina, O.V., Petrauskene, O.V., Gromova, E.S.
<2>Restriction endonuclease EcoRII hydrolyzes one of the two DNA restriction sites forming the catalytic complex.
<3>Mol. Biol. (Mosk)
<4>32
<5>793-796
<6>1998
<7>Hydrolysis of 71-mer substrates containing two EcoRII recognition sites was considered in
order to investigate the two-site mechanism of DNA recognition, which implies the presence in
the catalytic complex of four internucleotide bonds cleavable by the enzyme.  It is shown that
the enzyme hydrolyzes only two phosphodiester bonds in one enzyme-substrate complex.  To find
out whether both bonds belong to one and the same or different recognition sites, scissile
phosphodiester bonds in one of the recognition sites in the 71-mer duplexes were replaced with
nonscissile pyrophosphate bonds alternatively or in both strands simultaneously.  This allowed
one to preclude hydrolysis of the corresponding bonds.  It is shown that endonuclease EcoRII
cleaves simultaneously two internucleotide phosphodiester bonds belonging to the same
recognition site.

<>

<1>Baby, V., Matteau, D., Knight, T.F., Rodrigue, S.
<2>Complete Genome Sequence of the Mesoplasma florum W37 Strain.
<3>Genome Announcements
<4>1
<5>e00879-13
<6>2013
<7>Mesoplasma florum is a small-genome fast-growing mollicute that is an attractive  model for
systems and synthetic genomics studies. We report the complete
825,824-bp genome sequence of a second representative of this species, M. florum
strain W37, which contains 733 predicted open reading frames and 35 stable RNAs.

<>

<1>Bachi, B., Reiser, J., Pirrotta, V.
<2>Methylation and cleavage sequences of the EcoP1 restriction-modification enzyme.
<3>J. Mol. Biol.
<4>128
<5>143-163
<6>1979
<7>EcoP1 is a restriction modification enzyme encoded by bacteriophage P1.  It
requires ATP for cleavage and S-adenosyl methionine for methylation of DNA.  We
have mapped the sites of both cleavage and methylation in simian virus 40 DNA
and determined their sequences.  The enzyme methylates the sequence A-G-mA-C-C
and cuts the DNA 25 to 27 base-pairs form the site of methylation in the 3'
direction, with a two to four base-pair stagger between cuts.  Consistent with
the fact that the methylation sequence is asymmetric, the enzyme methylates
only one strand in vitro.  One variant of simian virus 40 has acquired an
additional EcoP1 methylation and cleavage site by changing a A-G-A-A-C sequence
to A-G-A-C-C.

<>

<1>Bachman, K.E., Rountree, M.R., Baylin, S.B.
<2>Dnmt3a and Dnmt3b are transcriptional repressors that exhibit unique localization properties to heterochromatin.
<3>J. Biol. Chem.
<4>276
<5>32282-32287
<6>2001
<7>We demonstrate that the recently identified DNA methyltransferases, Dnmt3a and Dnmt3b, like
DNMT1, repress transcription in a methylation-independent manner. Dnmt3a and Dnmt3b repress
transcription primarily through a plant homeodomain-like motif that is shared with the ATRX
protein but is not present in DNMT1. Unlike DNMT1, which localizes to replication foci during
S-phase in murine embryonic fibroblasts, Dnmt3a co-localizes with heterochromatin protein 1
alpha (HP1 alpha) and methyl-CpG binding proteins throughout the cell cycle to
late-replicating pericentromeric heterochromatin. In contrast to Dnmt3a, Dnmt3b remained
diffuse in the nucleus of embryonic fibroblasts at all cell cycle stages. However, Dnmt3a and
Dnmt3b co-localize to these pericentromeric heterochromatin regions in murine embryonic stem
cells. This finding is important to the fact that mutations in DNMT3B are found in the
developmental syndrome, ICF (immunodeficiency, centromeric heterochromatin instability, and
facial anomalies), which involves extensive loss of methylation from pericentromeric regions.
The localization of Dnmt3a and Dnmt3b was unaffected in Dnmt1 null embryonic stem cells, which
lose the majority of methylation at pericentromeric major satellite repeats, suggesting that
these enzymes are not dependent upon preexisting methylation for their targeting. DNMT1 is
then positioned to reestablish transcriptionally repressive chromatin as cells replicate,
while Dnmt3a and Dnmt3b may help to establish such chromatin in late S-phase and maintain this
repressive heterochromatin throughout the cell cycle in a developmentally and/or cell type
manner.

<>

<1>Backman, K.
<2>A cautionary note on the use of certain restriction endonucleases with methylated substrates.
<3>Gene
<4>11
<5>169-171
<6>1980
<7>Methylation of adenine and cytosine residues in DNA isolated from common strains of
Escherichia coli K-12 can render that DNA resistant to cleavage by certain restriction
endonucleases at those sites at which the recognition sequence for such an endonuclease
overlaps (but does not include) a sequence recognized by methylases specified by the dam or
dcm gene.

<>

<1>Bacolla, A., Pradhan, S., Larson, J.E., Roberts, R.J., Wells, R.D.
<2>Recombinant Human DNA (Cytosine-5) Methyltransferase. III. Allosteric control, reaction order, and influence of plasmid topology and triplet repeat length on methylation of the Fragile X CGG.CCG sequence.
<3>J. Biol. Chem.
<4>276
<5>18605-18613
<6>2001
<7>Steady-state kinetic analyses revealed that the methylation reaction of the human DNA
(cytosine-5) methyltransferase 1 (DNMT1) is repressed by the N-terminal domain comprising the
first 501 amino acids, and that repression is relieved when methylated DNA binds to this
region. DNMT1 lacking the first 501 amino acids retains its preference for hemimethylated DNA.
The methylation reaction proceeds by a sequential mechanism, and either substrate
(S-adenosyl-l-methionine and unmethylated DNA) may be the first to bind to the active site.
However, initial binding of S-adenosyl-l-methionine is preferred. The binding affinities of
DNA for both the regulatory and the catalytic sites increase in the presence of methylated CpG
dinucleotides and vary considerably (more than one hundred times) according to DNA sequence.
DNA topology strongly influences the reaction rates, which increased with increasing negative
superhelical tension. These kinetic data are consistent with the role of DNMT1 in maintaining
the methylation patterns throughout development and suggest that the enzyme may be involved in
the etiology of fragile X, a syndrome characterized by de novo methylation of a greatly
expanded CGG.CCG triplet repeat sequence.

<>

<1>Bacolla, A., Pradhan, S., Roberts, R.J., Wells, R.D.
<2>Recombinant human DNA (cytosine-5) methyltransferase. II. Steady-state kinetics reveal allosteric activation by methylated DNA.
<3>J. Biol. Chem.
<4>274
<5>33011-33019
<6>1999
<7>Initial velocity determinations were conducted with human DNA (cytosine-5) methyltransferase
(DNMT1) on unmethylated and hemimethylated DNA templates in order to assess the mechanism of
the reaction. Initial velocity data with DNA and S-adenosylmethionine (AdoMet) as variable
substrates and product inhibition studies with methylated DNA and S-adenosylhomocysteine
(AdoHcy) were obtained and evaluated as double-reciprocal plots. These relationships were
linear for plasmid DNA, exon-1 from the imprinted small nuclear ribonucleoprotein-associated
polypeptide N, (CGG.CCG)(12), (m(5)CGG.CCG)(12), and (CGG.CCG)(73) but were not linear for
(CGG.Cm(5)CG)(12). Inhibition by AdoHcy was apparently competitive versus AdoMet and
uncompetitive/noncompetitive versus DNA at </=20 microM AdoMet. Addition of the product
(methylated DNA) to unmethylated plasmid DNA increased V(max(app)) resulting in mixed
stimulation and inhibition. Velocity equations indicated a two-step mechanism as follows:
first, activation of DNMT1 by methylated DNA that bound to an allosteric site, and second, the
addition of AdoMet and DNA to the catalytic site. The preference of DNMT1 for hemimethylated
DNA may be the result of positive cooperativity of AdoMet binding mediated by allosteric
activation by the methylated CG steps. We propose that this activation plays a role in vivo in
the regulation of maintenance methylation.

<>

<1>Baddam, R., Kumar, N., Thong, K.L., Ngoi, S.T., Teh, C.S., Yap, K.P., Chai, L.C., Avasthi, T.S., Ahmed, N.
<2>Genetic Fine Structure of a Salmonella enterica Serovar Typhi Strain Associated with the 2005 Outbreak of Typhoid Fever in Kelantan, Malaysia.
<3>J. Bacteriol.
<4>194
<5>3565-3566
<6>2012
<7>Among enteric pathogens, Salmonella enterica serovar Typhi is responsible for the largest
number of food-borne outbreaks and fatalities. The ability of the
pathogen to cause systemic infection for extended durations leads to a high cost
of disease control. Chronic carriers play important roles in the evolution of
Salmonella Typhi; therefore, identification and in-depth characterization of
isolates from clinical cases and carriers, especially those from zones of
endemicity where the pathogen has not been extensively studied, are necessary.
Here, we describe the genome sequence of the highly virulent Salmonella Typhi
strain BL196/05 isolated during the outbreak of typhoid in Kelantan, Malaysia, in
2005. The whole-genome sequence and comparative genomics of this strain should
enable us to understand the virulence mechanisms and evolutionary dynamics of
this pathogen in Malaysia and elsewhere.

<>

<1>Baddam, R., Thong, K.L., Avasthi, T.S., Shaik, S., Yap, K.P., Teh, C.S., Chai, L.C., Kumar, N., Ahmed, N.
<2>Whole-Genome Sequences and Comparative Genomics of Salmonella enterica Serovar Typhi Isolates from Patients with Fatal and Nonfatal Typhoid Fever in Papua New  Guinea.
<3>J. Bacteriol.
<4>194
<5>5122-5123
<6>2012
<7>Many of the developing countries of the Southeast Asian region are significantly  affected by
endemic typhoid fever, possibly as a result of marginal living
standards. It is an important public health problem in countries such as Papua
New Guinea, which is geographically close to some of the foci of endemicity in
Asia. The severity of the disease varies in different regions, and this may be
attributable to genetic diversity among the native strains. Genome sequence data
on strains from different countries are needed to clearly understand their
genetic makeup and virulence potential. We describe the genomes of two Salmonella
Typhi isolates from patients with fatal and nonfatal cases of typhoid fever in
Papua New Guinea. We discuss in brief the underlying sequencing methodology,
assembly, genome statistics, and important features of the two draft genomes,
which form an essential step in our functional molecular infection epidemiology
program centering on typhoid fever. The comparative genomics of these and other
isolates would enable us to identify genetic rearrangements and mechanisms
responsible for endemicity and the differential severity of pathogenic
salmonellae in Papua New Guinea and elsewhere.

<>

<1>Badger, J.H. et al.
<2>Comparative genomic evidence for a close relationship between the dimorphic prosthecate bacteria Hyphomonas neptunium and Caulobacter  crescentus.
<3>J. Bacteriol.
<4>188
<5>6841-6850
<6>2006
<7>The dimorphic prosthecate bacteria (DPB) are alpha-proteobacteria that reproduce in an
asymmetric manner rather than by binary fission and are of
interest as simple models of development. Prior to this work, the only
member of this group for which genome sequence was available was the model
freshwater organism Caulobacter crescentus. Here we describe the genome
sequence of Hyphomonas neptunium, a marine member of the DPB that differs
from C. crescentus in that H. neptunium uses its stalk as a reproductive
structure. Genome analysis indicates that this organism shares more genes
with C. crescentus than it does with Silicibacter pomeroyi (a closer
relative according to 16S rRNA phylogeny), that it relies upon a
heterotrophic strategy utilizing a wide range of substrates, that its cell
cycle is likely to be regulated in a similar manner to that of C.
crescentus, and that the outer membrane complements of H. neptunium and C.
crescentus are remarkably similar. H. neptunium swarmer cells are highly
motile via a single polar flagellum. With the exception of cheY and cheR,
genes required for chemotaxis were absent in the H. neptunium genome.
Consistent with this observation, H. neptunium swarmer cells did not
respond to any chemotactic stimuli that were tested, which suggests that
H. neptunium motility is a random dispersal mechanism for swarmer cells
rather than a stimulus-controlled navigation system for locating specific
environments. In addition to providing insights into bacterial
development, the H. neptunium genome will provide an important resource
for the study of other interesting biological processes including
chromosome segregation, polar growth, and cell aging.

<>

<1>Badie, G.
<2>Role of DNA adenine methylase in regulation of bacterial gene expression, virulence, and the elicitation of immune responses.
<3>Ph.D. Thesis, Univ. of California, Santa Barbara
<4>
<5>1-149
<6>2005
<7>The threat of emerging infectious diseases and bio-warfare agents has underscored the need for
development of safe and effective anti-microbial therapies.  One viable approach to address
this issue is through protective vaccination, which second to water sanitation, is the
principal method for reduction of morbidity and mortality worldwide.  This dissertation
describes the role of the DNA adenine methylase as a global regulator of bacterial virulence
and the utility of Dam-based technology for the development of live-attenuated vaccines that
elicit potent states of immunity against a variety of infectious agents.  Dam is a highly
conserved enzyme found in many pathogens, and is involved in a variety of cellular processes
including the regulation of bacterial virulence and elicitation of protective immune
responses.  Work presented in this dissertation has further evaluated the role of Dam in
regulation of several virulence factors in Salmonella typhimurium and Yersinia
pseudotuberculosis.  Studies in Yersinia revealed that Dam overproduction alters the stringent
regulation of many virulence factors, including a principal Yersinia immunogen, LcrV, and an
actin cytotoxin, YopE.  Dysregulation of these virulence factors may be the mechanism by which
dam mutants elicit protective immunity against Yersinia infections in vaccinated hosts.  In
Salmonella, Dam regulates the expression of several virulence genes as well.  Moreover,
expression of these genes was differentially affected by altered Dam levels in closely-related
pathogenic and non-pathogenic strains of Salmonella.  These data suggest that differential
gene regulation, rather than gross genomic differences contributes to the virulence
disparities observed between these pathogenic and non-pathogenic isolates.  Finally, by
expressing reovirus antigens in Salmonella, the possibility of using Dam-based vaccines as
carriers of foreign antigens was investigated.  Effective live-attenuated carrier vaccines
could be used to protect hosts from a variety of diseases for which no therapy is currently
available.  Investigating the role of Dam in regulation of virulence factors in pathogens such
as Salmonella and Yersinia will not only help decipher its role in regulation of virulence in
other pathogens, but will also help design new vaccines that provide potent immunity against a
wide range of infectious diseases that threaten the safety of public health worldwide.

<>

<1>Badie, G., Heithoff, D.M., Mahan, M.J.
<2>LcrV synthesis is altered by DNA adenine methylase overproduction in Yersinia pseudotuberculosis and is required to confer immunity in  vaccinated hosts.
<3>Infect. Immun.
<4>72
<5>6707-6710
<6>2004
<7>Yersinia pseudotuberculosis mutants that overproduce the DNA adenine methylase (Dam(OP)
Yersinia) are attenuated, confer robust protective
immune responses, and synthesize or secrete several Yersinia outer
proteins (Yops) under conditions that are nonpermissive for synthesis
and secretion in wild-type strains. To understand the molecular basis
of immunity elicited by Dam(OP) Yersinia, we investigated the effects
of Dam overproduction on the synthesis and localization of a principal
Yersinia immunogen, LcrV, a low-calcium-responsive virulence factor
involved in Yop synthesis, localization, and suppression of host
inflammatory activities. Dam overproduction relaxed the stringent
temperature and calcium regulation of LcrV synthesis. Moreover, the
LcrV-dependent synthesis and localization of the actin cytotoxin, YopE,
were shown to be relaxed in Dam(OP) cells, suggesting that the
synthesis and localization of Yops can occur via both LcrV-dependent
and -independent mechanisms. Last, the immunity conferred by Dam(OP)
Yersinia was strictly dependent on the presence of LcrV, which may
result from its role (i) as an immunogen, (ii) as an immunomodullator
of host anti-inflammatory activities, or (iii) in the altered synthesis
and localization of Yops that could contribute to immunogen repertoire
expansion.

<>

<1>Badie, G., Heithoff, D.M., Sinsheimer, R.L., Mahan, M.J.
<2>Altered Levels of Salmonella DNA Adenine Methylase Are Associated with Defects in Gene Expression, Motility, Flagellar Synthesis, and Bile  Resistance in the Pathogenic Strain 14028 but Not in the Laboratory Strain  LT2.
<3>J. Bacteriol.
<4>189
<5>1556-1564
<6>2007
<7>Comparative genomic analysis has revealed limited strain diversity between Salmonella
pathogenic and nonpathogenic isolates. Thus, some of the
relative virulence and host-immune response disparities may be credited to
differential gene regulation rather than gross differences in genomic
content. Here we show that altered levels of Salmonella DNA adenine
methylase (Dam) resulted in acute defects in virulence-associated gene
expression, motility, flagellin synthesis, and bile resistance in the
Salmonella pathogenic strain 14028 but not in avirulent laboratory strain
LT2. The defects in motility exhibited by 14028 in response to altered Dam
levels was not dependent on the presence of the regulatory protein, RpoS.
The transitioning between flagellar types (phase variation) was also
differentially regulated in 14028 versus LT2 in response to dam levels,
resulting in distinct differences in flagellin expression states. These
data suggest that differential gene regulation may contribute to the
relative virulence disparities observed between Salmonella serovars that
are closely related at the DNA level.

<>

<1>Bae, H., Kim, K.P., Lee, J.I., Song, J.G., Kil, E.J., Kim, J.S., Kwon, S.T.
<2>Characterization of DNA polymerase from the hyperthermophilic archaeon Thermococcus marinus and its application to PCR.
<3>Extremophiles
<4>13
<5>657-667
<6>2009
<7>The family B DNA polymerase gene from the archaeon Thermococcus marinus (Tma) contains a long
open reading frame of 3,939 bp that encodes 1,312
amino acid residues. The gene is split by one intervening sequence that
forms a continuous open reading frame with the two polymerase exteins. In
this study, the Tma DNA polymerase gene both with (precursor form) and
without (mature form) its intein was expressed in Escherichia coli,
purified by heat treatment and HiTrap Heparin HP column chromatography and
characterized. Primary sequence analysis of the mature Tma polymerase
showed high sequence identity with DNA polymerases in the genus
Thermococcus. The expressed precursor form was easily spliced during
purification steps. The molecular mass of the purified Tma DNA polymerases
is about 90 kDa, as estimated by SDS-PAGE. Both Tma DNA polymerases showed
the same properties. PCR performed with this enzyme was found to be
optimal in the presence of 50 mM Tris-HCl (pH 8.4), 40 mM KCl, 12.5 mM
(NH(4))(2)SO(4,) 2 mM MgCl(2,) 0.05% Triton X-100 and 0.0075% BSA.
Furthermore, long-range PCR and time-saving PCR were performed using
various specific ratios of Taq and Tma DNA polymerases (Tma plus DNA
polymerase).

<>

<1>Bae, H., Kim, K.P., Song, J.M., Kim, J.H., Yang, J.S., Kwon, S.T.
<2>Characterization of intein homing endonuclease encoded in the DNA polymerase gene of Thermococcus marinus.
<3>FEMS Microbiol. Lett.
<4>297
<5>180-188
<6>2009
<7>The DNA polymerase gene of Thermococcus marinus (Tma) contains an intein inserted at the pol-b
site that possesses a 1611-bp ORF encoding
a 537-amino acid residue. The LAGLIDADG motif, often found in
site-specific DNA endonucleases, was detected within the amino acid
sequence of the intein. The intein endonuclease, denoted as PI-Tma, was
purified as a naturally spliced product from the expression of the
complete DNA polymerase gene in Escherichia coli. PI-Tma cleaved
intein-less DNA sequences, leaving four-base-long, 3'-hydroxyl
overhangs with 5'-phosphate. Nonpalindromic recognition sequences 19 bp
long were also identified using partially complementary oligonucleotide
pair sequences inserted into the plasmid pET-22b(+). Cleavage by PI-Tma
was optimal when present in 50 mM glycine-NaOH (pH 10.5), 150 mM KCl
and 12 mM MgCl2 at 70 degrees C.

<>

<1>Bae, M., Par, J.-O., Hwang, H.-Y., Yim, J.
<2>Purification and characterization of a restriction endonuclease from Streptomyces exfoliatus.
<3>Korean J. Microbiol.
<4>31
<5>544-549
<6>1993
<7>Thirty strains of Streptomyces sp. isolated from soil samples were screened for the presence
of site-specific endonuclease.  One strain among them showed endonuclease activity to cleave
lambda DNA.  This study describes the purification and characterization of a site-specific
restriction endonuclease SexIII from Streptomyces exfoliatus.  The purification procedure
includes streptomycin sulfate treatment, DEAE-cellulose, hydroxylapatite, phosphocellulose P11
column chromatography.  This enzyme required high Mg2+ concentration (20-40 mM), and showed
maximal activity at neutral pH(7-8) in the absence of NaCl.  The enzyme did not require salt
for its activity and was inhibited by over 75 mM NaCl.  SexIII cut lambda DNA and plasmid
pBluescript, recognized the hexanucleotide sequence 5'-CCGC/GG-3', and proved to be an
isoschizomer of SacII.

<>

<1>Bae, M., Song, E.-S.
<2>Purification of restriction endonuclease, SdiI, from Steptomyces diastatochromogenes.
<3>Korean J. Microbiol.
<4>32
<5>297-300
<6>1994
<7>About thirty bacterial strains of Actinomycetes, isolated from the soil, were examined for the
presence of restriction endonuclease activity.  Streptomyces diastatochromogenes, which was
identified previously, was found to contain restriction endonuclease activity.  The
purification of this enzyme, SdiI, was carried out via streptomycin sulfate precipitation and
ammonium sulfate fractionation followed by hydroxylapatite column chromatography.  Sephacryl
S-200 HR column chromatography and a second hydroxylapatite column chromatography.
SDS-polyacrylamide gel electrophoresis of the active protein (purified from various column
chromatography) resulted in 35,000 Da protein.

<>

<1>Bae, M., Song, E.-S., Hwang, H.-Y., Yim, J.
<2>Characterization of the restriction endonuclease, SdiI, from Streptomyces diastatochromogenes.
<3>Korean J. Microbiol.
<4>32
<5>301-305
<6>1994
<7>In catalytic properties of the restriction endonuclease, SdiI, which was purified from
Streptomyces diastatochromogenes, this enzyme was active over a wide pH range between 7.0 and
12.5 and up to 60oC and 500 mM NaCl.  It was stable between 20oC and 60oC, and MgCl2 is
essential for endonuclease activity.  The restriction map of lambda DNA which was obtained by
double digestion with various enzymes suggested that SdiI was an isoschizomer of XhoI.  From
the determination of the restriction site, based on DNA sequencing, the recognition and
cleavage specificity of SdiI was concluded to be
5'-C/TCGA G-3'
3'-G AGCT/C-5'.

<>

<1>Bae, M., Suh, W.-N., Park, J.-O., Kim, H.T., Lee, K.-J.
<2>Numerical identification of Streptomyces sp. 58 producing restriction endonuclease SexIII, a novel isoschizomer of SacII.
<3>Korean J. Microbiol.
<4>31
<5>465-470
<6>1993
<7>Numerical identification was carried out for an isolate of Streptomyces sp. 58 producing a new
restriction endonuclease SexIII.  Fifty taxonomic unit characters were tested and the data
were analyzed numerically using the TAXON program.  The isolate was identified to the major 5
of Streptomyces.  Therefore, it was concluded that the isolate was identified to be a member
of Streptomyces exfoliatus.

<>

<1>Bae, M., Suh, W.-N., Song, E.-S., Lee, K.-J.
<2>Numerical identification of a Streptomyces strain producing restriction endonuclease SdiI.
<3>Korean J. Appl. Microbiol. Biotechnol.
<4>22
<5>126-133
<6>1994
<7>Numerical identification was applied for Streptomyces sp. 264, an isolate producing a new
restriction endonuclease SdiI. The restriction enzyme would appear to be an isoschizomer of
XhoI. Fifty taxonomic unit characters were tested and the data obtained were analyzed
numerically by using the TAXON program. The isolate was identified to be the major cluster 19
of Streptomyces and best matched to S. diastatochromogenes. It was, therefore, concluded that
the isolate was identified to be a member of Streptomyces diastatochromogenes.

<>

<1>Bae, M., Yun, M.-S., Kim, H.-T., Lee, K.-J.
<2>Numerical identification of a Streptomyces strain producing a thermotolerable restriction endonuclease SviI.
<3>Korean J. Appl. Microbiol. Biotechnol.
<4>21
<5>299-305
<6>1993
<7>Numerical identification was carried out for an isolate of Streptomyces D2-5 producing a new
restriction endonuclease SviI. Fifty taxonomic unit characters were tested and the data were
analyzed numerically using the TAXON program. The isolate was best matched to Streptomyces
violochromogenes in the major cluster 18 of Streptomyces. Therefore, it was concluded that the
isolate was identified to be a member of Streptomyces violochromogenes.

<>

<1>Baev, A.A., Kravets, A.N., Solonin, A.S., Kuzmin, N.P., Moroz, A.F., Glatman, L.I., Tanyashin, V.I.
<2>Recombinant plasmid DNA PILRV8-delta determines synthesis of restriction endonuclease EcoRV.
<3>Soviet Patent Office
<4>SU 1130602 A
<5>
<6>1984
<7>Recombinant plasmid DNA pILRV8-delta has a size of 8,400 base pairs made up of fragments of
1.73 Kb of phage-lambda DNA and 4.0 Kb of pBR322 DNA; it contains the beta-lactamase gene and
rex phage-lambda gene; and the gene for the EcoRV restriction-modification system is under the
control of the righthand promoter of phage lambda, Pr. Also claimed is Escherichia coli
strain VKM R-4D which is a producer of restriction endonuclease EcoRV. In a preferred process,
recombinant plasmid DNA pILRV8-delta is constructed by genetic engineering by: splitting
recombinant plasmid DNA pILRV8 with HindIII; ligating the resulting fragments with T4
DNA-ligase; transforming E. coli cells lysogenic for phage lambda (C1+); and recovering the
desired recombinant plasmid DNA from ampicillin-stable clones.

<>

<1>Baev, A.A., Kravets, A.N., Solonin, A.S., Kuzmin, N.P., Moroz, A.F., Glatman, L.I., Tanyushin, V.I.
<2>Bacterial strain Escherichia coli VKM V-1455 D is useful in genetic engineering for producing restriction endonuclease EcoRV.
<3>Soviet Patent Office
<4>SU 1074139 A
<5>
<6>1984
<7>The strain is used to construct recombinant DNA plasmid pIL RV8. The method comprises:
preparing a medium of clones of the strain with DNA plasmid pILRV7; sampling the clones
containing basic DNA plasmid pILRV7-delta abbreviated to 500-600 pairs having the genes for
the EcoRV restriction-modification system and a unique endonuclease site for BglII; treating
the sampled DNA plasmid and DNA phage-lambda (C1875) with BglII; cross-linking the resulting
fragment with T4 DNA-ligase; transforming cells of Escherichia coli JC 5183 with the resulting
mixture of recombinant DNA molecules, and from the clones which are resistant to ampicillin
and have immunity against phage lambda, recovering the desired recombinant DNA plasmid.
Typically, the first stage of the process is carried out by culturing the above E.coli strain
containing plasmid pILRV7 in LB broth for 15 hrs. The culture is then sown into a selective
agar medium with ampicillin. Clones containing stable pILRV7 plasmid are sampled and the
plasmid DNA is separated by the Clewell-Helinski method.

<>

<1>Bahr, M.
<2>Biophysical characterization and biochemical direction of methyltransferase-DNA-interactions.
<3>Ph.D. Thesis, Germany
<4>
<5>
<6>2009
<7>
<>

<1>Bai, J., He, Z.
<2>Mammalian DNA methylase.
<3>Shengli Kexue Jinzhan
<4>28
<5>67-69
<6>1997
<7>
<>

<1>Baik, S.C., Lee, W.K., Song, J.Y., Choi, Y.J., Rye, B.D., Choi, S.H., Cho, M.J., Rhee, K.H.
<2>Purification and characterization of type II restriction endonuclease (HpyI) of Helicobacter pylori.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>0
<5>120
<6>1998
<7>This study describes the purification and characterization of type II restriction endonuclease
of Helicobacter pylori, a Gram-negative spiral bacterium associated with type B gastritis,
peptic ulcer, and gastric cancer in human.  Helicobacter pylori cytosol was subjected to
polyethyleneimine treatment, salt precipitation, heparin-sepharose column chromatography, and
fast protein liquid chromatography using Resource Q column and Mono Q column to purify the
type II restriction endonuclease.  HpyI was characterized to recognize the sequence
5'-GT(G/C)AC-3', yielding 5-base 5' protruding ends.  The restriction sequence was
identical to that of Tsp45I.  The molecular weight of native protein was estimated to be 22
kDa by size exclusion chromatography of FPLC.  The enzyme exhibited its maximal activity in
the presence of 10-20 mM NaCl, but was inhibited completely in the presence of NaCl more than
80 mM.  The enzyme showed its maximal activity in the presence of 1-10 mM MgCl2.  The optimal
pH and temperature for enzyme activity was pH 9.0 and 37E, respectively.  MnCl2 could not
substitute for MgCl2 in reaction mixture.  And addition of YA-mercaptoethanol and bovine serum
albumin in reaction mixture led to loss of enzyme activity of HpyI.  HpyI was confirmed to
digest genomic DNAs of enteric bacteria to less than 1 kb while it could not cut the genomic
DNAs of Helicobacter pylori isolates.  Eighty HpyI restriction sites were found in the whole
genome sequence of H. pylori reported by the Institute for Genomic Research, which are less
than 2.5% of estimated numbers of HpyI restriction sites in the whole genome.  The pBR322 DNA
that was treated with the crude extract of H. pylori cytosol in the presence of 0.3 mM
S-adenosylmethionine was partially protected from HpyI restriction.  Therefore, the enzymatic
activity in the crude extract is likely a DNA methyltransferase.  H. pylori DNA is supposed to
be protected from HpyI restriction by the rareness of HpyI restriction sites and methylation
of the site.  Taken together, HpyI might be the one of the barriers preventing the
introduction of foreign DNAs into H. pylori.

<>

<1>Bailey, A.C., Kellom, M., Poret-Peterson, A.T., Noonan, K., Hartnett, H.E., Raymond, J.
<2>Draft Genome Sequence of Massilia sp. Strain BSC265, Isolated from Biological Soil Crust of Moab, Utah.
<3>Genome Announcements
<4>2
<5>e01199-14
<6>2014
<7>Massilia sp. BSC265 was isolated from a biological soil crust near Moab, Utah. The strain
appears to be capable of chemotaxis and exopolysaccharide synthesis
for biofilm adhesion. The BSC265 genome contains a complete dissimilatory nitrate
reduction pathway as well as a TCA cycle, making it a facultative anaerobe.

<>

<1>Bailey, A.C., Kellom, M., Poret-Peterson, A.T., Noonan, K., Hartnett, H.E., Raymond, J.
<2>Draft Genome Sequence of Bacillus sp. Strain BSC154, Isolated from Biological Soil Crust of Moab, Utah.
<3>Genome Announcements
<4>2
<5>e01198-14
<6>2014
<7>Bacillus sp. BSC154 was isolated from a biological soil crust near Moab, Utah. The strain
appears to be capable of chemotaxis and biofilm production. The BSC154
genome contains iron siderophore production, nitrate reduction, mixed
acid-butanediol fermentation, and assimilatory and dissimilatory sulfate
metabolism pathways.

<>

<1>Bailey, A.C., Kellom, M., Poret-Peterson, A.T., Noonan, K., Hartnett, H.E., Raymond, J.
<2>Draft Genome Sequence of Microvirga sp. Strain BSC39, Isolated from Biological Soil Crust of Moab, Utah.
<3>Genome Announcements
<4>2
<5>e01197-14
<6>2014
<7>Microvirga sp. BSC39 was isolated from a biological soil crust near Moab, Utah. The strain
appears to be capable of chemotaxis and exopolysaccharide synthesis
for biofilm adhesion. The BSC39 genome contains iron siderophore uptake and
hydrolysis enzymes; however, it lacks siderophore synthesis pathways, suggesting
the uptake of siderophores produced by neighboring microbes.

<>

<1>Bailey, C.R., Winstanley, D.J.
<2>Inhibition of restriction in Streptomyces clavuligerus by heat treatment.
<3>J. Gen. Microbiol.
<4>132
<5>2945-2947
<6>1986
<7>Inefficient transformation of Streptomyces clavuligerus protoplasts by DNA from
the plasmid pIJ702, isolated from S. lividans, was attributed to restriction in
view of the observation that efficient transformation was observed using
modified pIJ702 (isolated from S. clavuligerus).  The restriction system could
be partially inhibited by treating protoplasts at 45C prior to transformation.
This treatment increased the transformation frequencies of pIJ702 DNA by
100-fold and was used to introduce other plasmids into S. clavuligerus.

<>

<1>Bair, C.L., Black, L.W.
<2>A Type IV Modification Dependent Restriction Nuclease that Targets Glucosylated Hydroxymethyl Cytosine Modified DNAs.
<3>J. Mol. Biol.
<4>366
<5>768-778
<6>2007
<7>The Escherichia coli CT596 prophage exclusion genes gmrS and gmrD were found to encode a novel
type IV modification-dependent restriction
nuclease that targets and digests glucosylated (glc)-hydroxymethylcytosine
(HMC) DNAs. The protein products GmrS (36 kDa) and GmrD (27 kDa) were
purified and found to be inactive separately, but together degraded
several different glc-HMC modified DNAs (T4, T2 and T6). The GMR enzyme is
able to degrade both alpha-glucosy-HMC T4 DNA and beta-glucosyl-HMC T4
DNA, whereas no activity was observed against non-modified DNAs including
unmodified T4 cytosine (C) DNA or non-glucosylated T4 HMC DNA. Enzyme
activity requires NTP, favors UTP, is stimulated by calcium, and initially
produces 4 kb DNA fragments that are further degraded to low molecular
mass products. The enzyme is inhibited by the T4 phage internal protein I*
(IPI*) to which it was found to bind. Overall activities of the purified
GmrSD enzyme are in good agreement with the properties of the cloned gmr
genes in vivo and suggest a restriction enzyme specific for sugar modified
HMC DNAs. IPI* thus represents a third generation bacteriophage defense
against restriction nucleases of the Gmr type.

<>

<1>Bair, C.L., Black, L.W.
<2>Setting the phage for evolution: The mechanism of host exclusion of bacteriophage T4 IPI.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>104
<5>392
<6>2004
<7>Interaction of bacteria and phages has led to the evolution of complex phage exclusion systems
and cognate exclusion avoidance mechanisms.  Chief among these are restriction-modification
system enzymes that attack the invading phage DNA and against which phages employ various
strategies including DNA modification.  The T-even phages of the myoviridae utilize extended
DNA modifications to host RMS's with base methylations, 100% hydroxymethylated-cytosine, and
alpha-, beta-, and other-glucosylated HmC derivaties.  The injected DNA-bound T4 IPI protein
(~350 molecules/DNA), nonessential in most hosts, has been shown to be necessary for
productive T4 infection of the pathogenic E. coli K isolate CT596.  This host has now been
shown to harbor a prophage that encodes the RMS genes gIBEGs (36.649 kb) and gIBEGd (26.854
kb), with 65% and 48% identity to H. pylori sequences of unknown function that have been
cloned and shown to be necessary and sufficient to restrict T4 DNA lacking IPI.
GpIBEGs/gpIBEGd encode a novel Type IV RMS that, unlike previously discovered E. coli
restriction systems, is able to digest the glucosylated-HmC T4 DNA, and is apparently
physically blocked from its recognition sequence by the presence of IPI on the DNA.  Active
tag derivatized forms of the two proteins have been purified to near homogeneity.  The
proteins are  inactive separately, but together degrade in apparently non-sequence-specific
manner glucosylated-HmC DNA substrates, with no activity against non-modified templates such
as T4 C-DNA.  The activity is stimulated by 1-2mM GTP and this effect is inhibited by excess
ATP.  It appears that this bacterial restriction system has evolved specifically to combat
infections by T-even phages.  Pursuing this struggle, the T-evens have evolved a highly
diverse and, in some cases, expanded family of capsid-targeted internal protein I (IPI) locus
gene products that may specifically shield the diverse modifications of their HmC residues
from IBEG family RMS's.

<>

<1>Bair, C.L., Rifat, D., Black, L.W.
<2>Exclusion of Glucosyl-Hydroxymethylcytosine DNA Containing Bacteriophages Is Overcome by the Injected Protein Inhibitor IPI*.
<3>J. Mol. Biol.
<4>366
<5>779-789
<6>2007
<7>The Escherichia coli isolate CT596 excludes infection by the Myoviridae T4 ip1(-) phage that
lacks the encapsidated IPI* protein normally injected into the host with the phage DNA.
Screening of a CT596 genomic library identified adjacent genes responsible for this exclusion,
gmrS (942 bp) and gmrD (708 bp) that are encoded by a cryptic prophage DNA. The two genes are
necessary and sufficient to confer upon a host the ability to exclude infection by T4 ip1(-)
phage and other glucosyl-hydroxymethylcytosine (glc-HMC) Tevens lacking the ip1 gene, yet
allow infection by phages with non-glucoslyated cytosine (C) DNA that lack the ip1 gene. A
plasmid expressing the ip1 gene product, IPI*, allows growth of Tevens lacking ip1 on E. coli
strains carrying the cloned gmrS/gmrD genes. Members of the Teven family carry a diverse and,
in some cases, expanded set of ip1 locus genes. In vivo analysis suggests a family of gmr
genes that specifically target sugar-HMC modified DNA have evolved to exclude Teven phages,
and these exclusion genes have in turn been countered by a family of injected exclusion
inhibitors that likely help determine the host range of different glc-HMC phages.

<>

<1>Bakaletz, L.O., Munson, R., Dyer, D.
<2>Genes of an otitis media isolate of nontypeable haemophilus influenzae.
<3>European Patent Office
<4>EP 2070945 A
<5>
<6>2009
<7>The invention relates to the polynucleotide sequence of a nontypeable strain of Haemophilus
influenzae (NTHi) and polypeptides encoded by the polynucleotides and uses thereof.  The
invention also relates to NTHi genes which are upregulated during or in response to NTHi
infection of the middle ear and/or the nasopharynx.

<>

<1>Bakaletz, L.O., Munson, R.S. Jr., Dyer, D.W.
<2>Polypeptide encoded by a nucleotide sequence of a nontypeable strain of Haemophilus influenzae genome.
<3>US Patent Office
<4>US 7241867 A
<5>
<6>2007
<7>The invention relates to the polynucleotide sequence of a nontypeable strain of Haemophilus
influenzae (NTHi) and polypeptides encoded by the polynucleotides and uses thereof.  The
invention also relates to NTHi genes which are upregulated during or in response to NTHi
infection of the middle ear and/or the nasopharynx.

<>

<1>Bakaletz, L.O., Munson, R.S. Jr., Dyer, D.W.
<2>GENES OF AN OTITIS MEDIA ISOLATE OF NONTYPEABLE HAEMOPHILUS.
<3>Japanese Patent Office
<4>JP 2006519605 A
<5>
<6>2006
<7>
<>

<1>Bakaletz, L.O., Munson, R.S., Dyer, D.W.
<2>Genes of an otitis media isolate of nontypeable haemophilus influenzae.
<3>International Patent Office
<4>WO 2004078949 A
<5>
<6>2004
<7>The invention relates to the polynucleotide sequence of a nontypeable strain of Haemophilus
influenzae (NTHi) and polypeptides encoded by the polynucleotides and uses thereof.  The
invention also relates to NTHi genes which are upregulated during or in response to NTHi
infection of the middle ear and/or the nasopharynx.

<>

<1>Baker, B.J., Comolli, L.R., Dick, G.J., Hauser, L.J., Hyatt, D., Dill, B.D., Land, M.L., Verberkmoes, N.C., Hettich, R.L., Banfield, J.F.
<2>Enigmatic, ultrasmall, uncultivated Archaea.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>8806-8811
<6>2010
<7>Metagenomics has provided access to genomes of as yet uncultivated microorganisms
in natural environments, yet there are gaps in our knowledge-particularly for
Archaea-that occur at relatively low abundance and in extreme environments.
Ultrasmall cells (<500 nm in diameter) from lineages without cultivated
representatives that branch near the crenarchaeal/euryarchaeal divide have been
detected in a variety of acidic ecosystems. We reconstructed composite,
near-complete approximately 1-Mb genomes for three lineages, referred to as ARMAN
(archaeal Richmond Mine acidophilic nanoorganisms), from environmental samples
and a biofilm filtrate. Genes of two lineages are among the smallest yet
described, enabling a 10% higher coding density than found genomes of the same
size, and there are noncontiguous genes. No biological function could be inferred
for up to 45% of genes and no more than 63% of the predicted proteins could be
assigned to a revised set of archaeal clusters of orthologous groups. Some core
metabolic genes are more common in Crenarchaeota than Euryarchaeota, up to 21% of
genes have the highest sequence identity to bacterial genes, and 12 belong to
clusters of orthologous groups that were previously exclusive to bacteria. A
small subset of 3D cryo-electron tomographic reconstructions clearly show
penetration of the ARMAN cell wall and cytoplasmic membranes by protuberances
extended from cells of the archaeal order Thermoplasmatales. Interspecies
interactions, the presence of a unique internal tubular organelle [Comolli, et
al. (2009) ISME J 3:159-167], and many genes previously only affiliated with
Crenarchaea or Bacteria indicate extensive unique physiology in organisms that
branched close to the time that Cren- and Euryarchaeotal lineages diverged.

<>

<1>Baker, D.
<2>Proteins by design.
<3>The Scientist
<4>20
<5>26-32
<6>2006
<7>Imagine having the power to create a brand new protein - biosensor for any small molecule,
say, or a novel enzyme - on demand.  It's not pure fantasy.  Computational structural biology
is poised to put this power into our hands.  Along with a team of research groups around the
world, we have begun designing novel proteins and folds from scratch, computing amino acid
sequences that will fold to create enzymatic activities never before seen in nature.  The
possibilities are limited only by our imaginations: Picture an endonuclease designed to thwart
malaria, molecular sensors for bioterror agents, or a vaccine that HIV is less likely to
evolve around.  The mechanics of these engineering feats are closely related, perhaps not
surprisingly, to their logical inverse: structure prediction.  Scientists have for years tried
to develop methods for predicting a protein's structure simply from its amino acid sequence.
Imagine that in the time it takes to sequence the genome of an organism, scientists could also
characterize the structure of each of its proteins.  This would unveil biomolecular
interactions, structural homologies, functional roles, and potential drug targets that might
never be found from gene sequence alone.  Although there is still a long way to go, with
improvements to algorithms and increases in computing power, exciting progress is being made
in both prediction and design.

<>

<1>Baker, D.J., Hardy, T.A., Smith, S.S.
<2>The influence of the dT.dG mispair on the activity of the human DNA (cytosine-5) methyltransferase.
<3>Biochem. Biophys. Res. Commun.
<4>146
<5>596-602
<6>1987
<7>Synthetic oligodeoxynucleotides containing a dT-dG mispair at a centrally located d(pCG) dimer
are methylated at a moderate rate by highly purified human DNA (cytosine-5) methyltransferase
(E.C.2.1.1.37). The presence of a mispaired dT in one strand induced the enzyme to
preferentially methylate the opposite strand.

<>

<1>Baker, D.J., Kan, J.L.C., Smith, S.S.
<2>Recognition of structural perturbations in DNA by human DNA(cytosine-5)methyltransferase.
<3>Gene
<4>74
<5>207-210
<6>1988
<7>We have used oligodeoxynucleotides to study DNA methyltransferase activity on several types of
DNA structure in which the cytosine is mispaired to test these predictions. These sequences,
30 nt in length, containing either C-C or A-C mismatches have been studied in detail. In both
cases mispaired cytosines are selectively enhanced as acceptors of DNA methylation. Our
findings with the C-C mispair are illustrative. A kinetic comparison showed that a duplex
containing a C-C mispair was methylated about seven-fold faster than a complementary
unmethylated duplex of otherwise identical sequence. Autoradiographic analysis of the
enzymatically methylated product showed that the methyl groups were applied to each of the
cytosines in the mismatch with about equal efficiency. MspI (but not HpaII) was able to
partially cleave the duplex structure containing the mismatch, suggesting that there is a
dynamic equilibrium in solution between structures like those shown in Fig. 1. The d(pCCGG)
recognition site is regenerated in structures (2) and (4).

<>

<1>Baker, D.J., Laayoun, A., Smith, S.S.
<2>Transition state analogs as affinity labels for human DNA methyltransferases.
<3>Biochem. Biophys. Res. Commun.
<4>196
<5>864-871
<6>1993
<7>A new class of affinity labels has been developed for human DNA (cytosine-5)
methyltransferases. These oligodeoxynucleotides contain 5-fluorodeoxycytidine at a mispair
within the recognition motif of the human enzyme. They were not effectively recognized by
bacterial methyltransferases. They can be viewed as analogs of the intermediates transiently
produced by methyltransferases during catalysis. Affinity labelling patterns suggest that both
the structurally induced activity and the methyl-directed activity of the human enzymes
operate by the same mechanism and reside on the same polypeptide chains.

<>

<1>Bakh, N.L., Semina, I.E.
<2>Obtaining the preparations of immobilized restrictases SalI and PvuII.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>2
<5>32-34
<6>1985
<7>Conditions for the immobilization of specific endonucleases SalI and PvuII on
BrCN-activated Sepharose 4B have been selected.  Some physico-chemical
properties of the preparations of immobilized restrictases SalI and PvuII have
been characterized.  The specific and general activity values of the
preparations thus obtained have been established.  The immobilized enzymes have
been used for the multiple restriction of the DNA of phage lambda and the DNA
of Neisseria meningitidis.

<>

<1>Bakh, N.L., Tsvetkova, N.V., Semina, I.E., Tarasov, A.P., Mileikovskaya, M.M., Gruber, I.M., Polyachenko, V.M., Romanenko, E.E.
<2>Isolation and Purification of Restriction Endonuclease PmiI from Pseudomonas Mirabilis 1667.
<3>Antibiot. Med. Biotekhnol.
<4>30
<5>342-344
<6>1985
<7>A new restriction endonuclease PmiI was detected in Proteus mirabilis 1667.
The enzyme hydrolyzes DNA of the phage lambda into 10 electrophoretically
separating fragments with molecular weights of 1.3-7.9 mD.  With the use of
two-stage chromatography on sepharose and phosphocellulose it is possible to
obtain restriction endonuclease PmiI free of the admixtures of ballast
proteins, nonspecific nucleases and phosphatases.

<>

<1>Bakhrat, A., Baranes, K., Krichevsky, O., Rom, I., Schlenstedt, G., Pietrokovski, S., Raveh, D.
<2>Nuclear import of Ho endonuclease utilizes two nuclear localization signals and four importins of the ribosomal import system.
<3>J. Biol. Chem.
<4>281
<5>12218-12226
<6>2006
<7>Activity of Ho, the yeast mating switch endonuclease, is restricted to a narrow time window of
the cell cycle. Ho is unstable and despite
being a nuclear protein is exported to the cytoplasm for proteasomal
degradation. We report here the molecular basis for the highly
efficient nuclear import of Ho and the relation between its short
half-life and passage through the nucleus. The Ho nuclear import
machinery is functionally redundant, being based on two bipartite
nuclear localization signals, recognized by four importins of the
ribosomal import system. Ho degradation is regulated by the DNA damage
response and Ho retained in the cytoplasm is stabilized, implying that
Ho acquires its crucial degradation signals in the nucleus. Ho arose by
domestication of a fungal VMA1 intein. A comparison of the primary
sequences of Ho and fungal VMA1 inteins shows that the Ho nuclear
localization signals are highly conserved in all Ho proteins, but are
absent from VMA1 inteins. Thus adoption of a highly efficient import
strategy occurred very early in the evolution of Ho. This may have been
a crucial factor in establishment of homothallism in yeast, and a key
event in the rise of the Saccharomyces sensu stricto.

<>

<1>Bakhrat, A., Jurica, M.S., Stoddard, B.L., Raveh, D.
<2>Homology modeling and mutational analysis of Ho endonuclease of yeast.
<3>Genetics
<4>166
<5>721-728
<6>2004
<7>Ho endonuclease is a LAGLIDADG homing endonuclease that initiates mating-type interconversion
in yeast. Ho is encoded by a free-standing
gene but shows 50% primary sequence similarity to the intein
(protein-intron encoded) PI-SceI. Ho is unique among LAGLIDADG
endonucleases in having a 120-residue C-terminal putative zinc finger
domain. The crystal structure of PI-SceI revealed a bipartite enzyme
with a protein-splicing domain (Hint) and intervening endonuclease
domain. We made a homology model for Ho on the basis of the PI-SceI
structure and performed mutational analysis of putative critical
residues, using a mating-type switch as a bioassay for activity and
GFP-fusion proteins to detect nuclear localization. We found that
residues of the N-terminal sequence of the Hint domain are important
for Ho activity, in particular the DNA recognition region. C-terminal
residues of the Hint domain are dispensable for Ho activity; however,
the C-terminal putative zinc finger domain is essential. Mutational
analysis indicated that residues in Ho that are conserved relative to
catalytic, active-site residues in PI-SceI and other related homing
endonucleases are essential for Ho activity. Our results indicate that
in addition to the conserved catalytic residues, Hint domain residues
and the zinc finger domain have evolved a critical role in Ho activity.

<>

<1>Bakshi, S., Cipps, T., Hanner, M., Pikalo, J., Satke, C., Nagy, E., Lundberg, U., Zierer, D., Meinke, A., Noiges, B., Stierschneider, U., Von Gabain, A.
<2>Klebsiella antigens.
<3>International Patent Office
<4>WO 2008135446 A
<5>
<6>2008
<7>The present invention relates to isolated nucleic acid molecules which encode an antigen, a
vector which comprises such nucleic acid molecule, and a host cell comprising such vector.
Furthermore, the invention provides antigens from Klebsiella species, as well as fragments and
variants thereof, a process for producing such antigens, and a process for producing a cell,
which expresses such antigen.  Moreover, the present invention provides antibodies binding to
such antigen, a hybridoma cell producing such antibodies, methods for producing such
antibodies, a pharmaceutical composition comprising such nucleic acid molecule, antigen,
vector or antibody, the use of such nucleic acid molecule, antigen, vector or antibody for the
preparation of a pharmaceutical composition, methods for identifying an antagonist capable of
binding such antigen or of reducing or inhibiting the interaction activity of such antigen,
methods for diagnosing an infection and methods for the treatment or prevention of an
infection.  More specifically such antigens are produced by or associated with bacterial
pathogens causing nocosomial infections or bacterial infections caused by Klebsiella
pneumonia.

<>

<1>Baksi, K., Rogerson, D.L., Rushizky, G.W.
<2>Rapid, single-step purification of restriction endonucleases on Cibacron blue F3GA-agarose.
<3>Biochemistry
<4>17
<5>4136-4139
<6>1978
<7>After sonication and high-speed centrifugation, crude extracts of B.
amyloliquefaciens, P. alcalifaciens, X. holicola, and B. globiggi were absorbed
on the dye Cibacron blue F3GA covalently cross-linked to agarose.  The
restriction endonucleases BamHI, PalI, XhoI, and BglI together with BglII were
isolated by elution of the dye column with linear gradients to 0.5 M NaCl.  The
enzymes so purified were free of contaminating nucleic acids and other
nucleases and were sufficiently concentrated for direct, specific DNA
hydrolysis.

<>

<1>Baksi, K., Rushizky, G.W.
<2>Rapid purification of restriction endonucleases on Cibacron Blue F3GA.
<3>Fed. Proc.
<4>37
<5>1414
<6>1978
<7>At present, purification of Type II restriction nucleases involves, after cell
breakage and high-speed centrifugation, at least two more steps: DNA removal
and column chromatography of the enzyme(s).  We show that the high-speed
supernatant may be directly adsorbed on Cibacron Blue F3GA covalently
cross-linked to agarose (CB) and restriction enzymes eluted with linear
gradients from 0-0.5 M NaCl.  Per gram of frozen cells, 700 or more units of
Bgl (I and II), Xho (I and II) and PalI were so isolated, free of contaminating
nuclease activity as judged by electrophoresis of lambda DNA digests on 1.4%
agarose gels.  The oligonucleotide band patterns observed agreed with those
reported by others for the purified enzymes.  While the content of restriction
nuclease activity varied between different batches of frozen cells, the
recovery of enzyme activity islated by CB chromatography is comparable to that
obtained by other procedures.  The extent of purification (Lowry protein) was
55-fold (PalI).  Purified AluI, BamHI, XhoI and HaeIII (the isoschizomer of
PalI) (N.E. Biolabs) were similarly bound on and eluted by serum albumin at 5
mg/ml or calf thymus nucleic acid at 125 microgram/ml.

<>

<1>Baksi, K., Rushizky, G.W.
<2>Purification of the restriction endonuclease PalI.
<3>Anal. Biochem.
<4>99
<5>207-212
<6>1979
<7>The restriction endonuclease PalI was purified from Providencia alcalifaciens 1650-fold with a
yield of 33%.  The purified protein moved as a single band upon polyacrylamide gel
electrophoresis.  When this was carried out in the presence of sodium dodecyl sulfate, a
molecular weight of 31,000 was obtained for PalI.  Gel filtration through Sephacryl S200 gave
molecular weights ranging from 44,000 to 53,000 when 58 to 1870 ng/ml enzyme were used,
respectively.  Other properties of the enzyme are described.

<>

<1>Bala, M., Kumar, S., Raghava, G.P., Mayilraj, S.
<2>Draft Genome Sequence of Rhodococcus ruber Strain BKS 20-38.
<3>Genome Announcements
<4>1
<5>e0013913
<6>2013
<7>We report the 6.1-Mb genome sequence of Rhodococcus ruber strain BKS 20-38, isolated from the
palm tree rhizosphere soil of Bhitarkanika National Park,
Odhisha, India. The draft genome sequence of strain BKS 20-38 consists of
6,126,900 bp, with a G+C content of 69.72%, 5,716 protein-coding genes, and 49
RNAs.

<>

<1>Bala, M., Kumar, S., Raghava, G.P., Mayilraj, S.
<2>Draft Genome Sequence of Rhodococcus qingshengii Strain BKS 20-40.
<3>Genome Announcements
<4>1
<5>e0012813
<6>2013
<7>We report the 5.8-Mb genome sequence of Rhodococcus qingshengii strain BKS 20-40, isolated
from a palm tree rhizosphere soil sample from Bhitarkanika National
Park, Odisha, India. The strain is capable of degrading cholesterol moiety. The
draft genome of strain BKS 20-40 consists of 6,601,618 bp, with 62.4% G+C
content.

<>

<1>Balabanov, V.P., Kotova, V.Yu., Kholodii, G.Y., Mindlin, S.Z., Zavilgelsky, G.B.
<2>A novel gene, ardD, determines antirestriction activity of the non-conjugative transposon Tn5053 and is located antisense within the tniA gene.
<3>FEMS Microbiol. Lett.
<4>337
<5>55-60
<6>2012
<7>The mercury-resistance transposon Tn5053 inhibits restriction activity of the type I
restriction-modification endonuclease EcoKI in
Escherichia coli K12 cells. This is the first report of antirestriction
activity of a non-conjugative transposon. The gene (ardD) coding for
the antirestriction protein has been cloned. The ardD gene is located
within the tniA gene, coding for transposase, on the complementary
strand. The direction of transcription is opposite to transcription of
the tniA gene.

<>

<1>Balabanov, V.P., Pustovoit, K.S., Zavilgelsky, G.B.
<2>Comparative analysis of antirestriction activity of the ArdA and ArdB proteins encoded by genes of the R64 transmissible plasmid (IncI1).
<3>Mol. Biol. (Mosk)
<4>46
<5>269-275
<6>2012
<7>Antirestriction proteins ArdA and ArdB are specific inhibitors of type I
restriction-modification enzymes. The ardA and yfeB (ardB) genes were cloned from the
transmissible plasmid R64 in the pUC18 and pZE21 vectors. The R64 ArdA and ArdB proteins were
shown to inhibit only restriction activity of the type I restriction-modification enzyme
(EcoKI) in Escherichia coli K12 cells. In contrast to ArdA, ArdB inhibited EcoKI restriction
activity only at a high intracellular concentration. Antirestriction activity of ArdB did not
depend on the ClpXP protease. The yfeB (ardB) gene of the R64 plasmid is transcribed from a
weak promoter located upstream of yfeA.

<>

<1>Balado, M., Lemos, M.L., Osorio, C.R.
<2>Integrating conjugative elements of the SXT/R391 family from fish-isolated Vibrios encode restriction-modification systems that confer resistance to bacteriophages.
<3>FEMS Microbiol. Ecol.
<4>83
<5>457-467
<6>2013
<7>Integrating conjugative elements (ICEs) of the SXT/R391 family have been described in Vibrios,
mainly Vibrio cholerae, and other bacteria
as carriers of variable gene content conferring adaptive advantages
upon their hosts, including antimicrobial resistance and motility
regulation. However, our knowledge on their host range and ecological
significance is still limited. Here, we report the identification and
characterization of ICEVspPor3 and ICEValSpa1, two novel ICEs of the
SXT/R391 family from fish-isolated Vibrio splendidus and Vibrio
alginolyticus, respectively. We found that ICEVspPor3 carries
tetracycline and HgCl2 resistance determinants and can be transferred
by conjugation to Escherichia coli and to several species of marine
bacteria including some of the major bacterial fish pathogens in marine
aquaculture, whereas ICEValSpa1 lacks resistance genes. Interestingly,
both ICEs harbor genes encoding distinct restrictionmodification (RM)
systems. We demonstrate here that these RM systems, when expressed in
E. coli, confer protection to infection by T1 bacteriophage and by
environmental water bacteriophages. Our results provide evidences that
the variable gene content of ICEs of the SXT/R391 family encodes
fitness functions beyond those related to antimicrobial resistance and
motility regulation and suggest that the host range of these elements
in the marine environment might be broader than expected.

<>

<1>Balcarova, A., Mrazek, J., Kleinwachter, W., Brabec, V.
<2>Cleavage by restriction enzymes of DNA modified with the antitumour drug cis-diamminedichloroplatinum (II).
<3>Gen. Physiol. Biophys.
<4>11
<5>579-588
<6>1992
<7>The effect of binding of an antitumour drug cis-diamminedichloroplatinum (II)
(cis-[Pt(NH3)2Cl2]) to DNA on cutting effectiveness of BamHI, EcoRI, and SalI restriction
endonucleases was quantitatively determined. The platinum complex inhibits the cleavage of
plasmid pHC624 DNA linearized by BglI restrictase. From the present results we conclude that
the yield of restriction endonuclease cleavage is also lowered if the platinum complex is
bound outside the recognition DNA sequence of these enzymes. We propose that the origin of
platinum adducts on DNA outside the recognition sequence can decrease the yield of restriction
enzyme cleavage via inducing a conformational perturbation in the DNA recognition sequence of
these enzymes and also via inhibition of the linear diffusion of these enzymes on DNA.

<>

<1>Baldauf, F., Kiss, A.
<2>Expression of cloned gene for methyltransferase from Bacillus subtilis bacteriophage SPbetaB.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>2
<5>26-28
<6>1985
<7>Expression of the methyltransferase gene from Bacillus subtilis lysogenizing phage SPbetaB was
studied by analyzing the sensitivity of the hybrid plasmid DNAs to restriction by the enzymes
BspRI, HpaII and MspI.  This gene produces the methylase M.BsuPbetaBI with specificity for
5'-GGCC.  The fragment carrying the SPbetaB derived gene also directs the synthesis in E.
coli of a second methylase activity (M.BsuPbetaBII) with 5'-CCGG specificity.  Indirect
evidence suggests that the two SPbetaB modification activities are encoded by the same gene.

<>

<1>Baldwin, D.N., Shepherd, B., Kraemer, P., Hall, M.K., Sycuro, L.K., Pinto-Santini, D.M., Salama, N.R.
<2>Identification of Helicobacter pylori genes that contribute to stomach colonization.
<3>Infect. Immun.
<4>75
<5>1005-1016
<6>2007
<7>Chronic infection of the human stomach by Helicobacter pylori leads to a variety of
pathological sequelae, including peptic ulcer and gastric
cancer, resulting in significant human morbidity and mortality. Several
genes have been implicated in disease related to H. pylori infection,
including the vacuolating cytotoxin and the cag pathogenicity island.
Other factors important for the establishment and maintenance of infection
include urease enzyme production, motility, iron uptake, and stress
response. We utilized a C57BL/6 mouse infection model to query a
collection of 2,400 transposon mutants in two different bacterial strain
backgrounds for H. pylori genetic loci contributing to colonization of the
stomach. Microarray-based tracking of transposon mutants allowed us to
monitor the behavior of transposon insertions in 758 different gene loci.
Of the loci measured, 223 (29%) had a predicted colonization defect. These
included previously described H. pylori virulence genes, genes implicated
in virulence in other pathogenic bacteria, and 81 hypothetical proteins.
We have retested 10 previously uncharacterized candidate colonization gene
loci by making independent null alleles and have confirmed their
colonization phenotypes by using competition experiments and by
determining the dose required for 50% infection. Of the genetic loci
retested, 60% have strain-specific colonization defects, while 40% have
phenotypes in both strain backgrounds for infection, highlighting the
profound effect of H. pylori strain variation on the pathogenic potential
of this organism.

<>

<1>Baldwin, G.S., Gormley, N.A., Halford, S.E.
<2>Manganese(II) as a probe for the mechanism and specificity of restriction endonucleases.
<3>Metal Ions in Biological Systems, Vol. 37: Manganese and its role in Biological Processes., Marcel Dekker, Sigel, A. & Sigel, H., New York
<4>37
<5>345-364
<6>2000
<7>A review of the role of metal ions in DNA cleavage by restriction enzymes and the experimental
possibilities offered by substituting manganese for magnesium.

<>

<1>Baldwin, G.S., Halford, S.E.
<2>Rapid reaction kinetics on the EcoRV restriction endonuclease.
<3>Biochem. Soc. Trans.
<4>22
<5>300S
<6>1994
<7>The EcoRV restriction endonuclease cleaves DNA between the central TpA step of its 6 bp
recognition sequence (GATATC) leaving blunt ends. It utilizes Mg2+ as its sole cofactor in the
hydrolysis of the phosphodiester backbone. Previous studies have provided a wealth of
information on the structure and function of the EcoRV restriction endonuclease. Perhaps the
most surprising aspect of EcoRV's properties is that it binds all DNA sequences with equal
affinity, yet it cleaves its target sequence with a 10^6 fold preference over the next best
site. The very high degree of specificity exhibited by EcoRV is thus derived from catalysis,
as opposed to sequence specific DNA binding. Nearly all of these previous studies were
performed under steady state conditions, and have consequently provided little insight into
the reaction pathway. We have used rapid reaction techniques under single turnover conditions
to resolve intermediates in the reaction pathway with the aim of determining the mechanism of
catalysis.

<>

<1>Baldwin, G.S., Kelly, S.M., Price, N.C., Wilson, G.W., Connolly, B.A., Artymiuk, P.J., Hornby, D.P.
<2>Ligand-induced conformational states of the cytosine-specific DNA methyltransferase M.HgaI-2.
<3>J. Mol. Biol.
<4>235
<5>545-553
<6>1994
<7>The interaction of one of the two DNA methyltransferases encoded by the HgaI restriction and
modification system, M.HgaI-2, with substrates and substrate analogues is described. Circular
dichroism spectroscopy has been used to demonstrate that addition of the methyl donor,
S-adenosyl-L-methionine and the inhibitory substrate analogue sinefungin, both induce
conformational transitions in the protein in the absence of DNA. Moreover, the addition of DNA
is shown to enhance the apparent secondary structure of M.HgaI-2 whilst addition of sinefungin
or S-adenosyl-L-methionine reduces apparent secondary structure. The circular dichroism
spectrum of the abortive complex between the enzyme, DNA and sinefungin is dominated by the
conformational properties of the binary complex of enzyme and sinefungin alone. Addition of a
specific oligodeoxynucleotide duplex in which the target cytosine is replaced by a
pyrimidinone, leads to a further ligand induced conformational transition as determined by
electrophoretic analysis. The addition of sinefungin, or S-adenosyl-L-methionine, to M.HgaI-2
bound to the reactive oligodeoxynucleotide duplex, leads to yet another conformational
transition in the protein as determined by the differential susceptibility of ternary and
binary complexes to proteolysis. These experiments identify at least six ligand-inducible
conformational states of M.HgaI-2 and, in view of the sequence similarity amongst this class
of enzymes, suggest that conformational flexibility is a general feature of C-5
cytosine-specific DNA methyltransferases. Moreover, the substitution of the target cytosine by
a pyrimidinone mimics the effect of 5-azacytosine incorporation into DNA.

<>

<1>Baldwin, G.S., Sessions, R.B., Erskine, S.G., Halford, S.E.
<2>DNA cleavage by the EcoRV restriction endonuclease: Roles in divalent metal ions in specificity and catalysis.
<3>J. Mol. Biol.
<4>288
<5>87-103
<6>1999
<7>The roles of divalent metal ions in DNA cleavage by the EcoRV endonuclease were studied by
using Co2+ or Mn2+ as substitutes for the natural cofactor Mg2+.  In steady-state experiments
with a 12 bp oligonucleotide substrate, Co2+ yielded a similar turnover rate to that with
Mg2+, but Mn2+ gave a slower rate.  Single turnovers of EcoRV on this substrate were analyzed
by stopped-flow and quench-flow methods, to determine the rates for the formation of the
ternary enzyme-DNA-metal complex, the hydrolysis of the phosphodiester bonds and the
dissociation of the cleaved DNA.  With Co2+, all three steps had similar rates to those with
Mg2+.  In contrast, Mn2+ gave a faster rate for phosphodiester hydrolysis than either Mg2+ or
Co2+, but a slower rate for product dissociation, thus accounting for its low turnover rate.
Single turnovers on plasmids also yielded faster rates for substrate hydrolysis with Mn2+
compared to Mg2+ and Co2+.  Since Mn2+ gave the most rapid rates for the hydrolytic step,
despite being less electronegative than Co2+, the function of the metal ion at the active site
of EcoRV cannot be just the polarization of the scissile phosphate.  Moreover, the minimal
scheme for the Co2+-catalysed reaction requires two metal ions for DNA cleavage.  The metal
ions seem to be involved in the precise positioning of both the substrate and the water that
acts as the attacking nucleophile and in activating that water molecule.  A model is presented
to account for how two metal ions might fulfil these functions.

<>

<1>Baldwin, G.S., Vipond, I.B., Halford, S.E.
<2>Rapid reaction analysis of the catalytic cycle of the EcoRV restriction endonuclease.
<3>Biochemistry
<4>34
<5>705-714
<6>1995
<7>We have used the intrinsic tryptophan fluorescence of the EcoRV restriction endonuclease to
monitor changes in protein conformation during binding and cleavage of a duplex
oligodeoxynucleotide substrate. Appropriate conditions for single-turnover reactions were
first determined by steady-state kinetics. When single turnovers were monitored by
stopped-flow fluorescence, the mixing together of EcoRV oligonucleotide and MgCl2 resulted in
a rapid increase in tryptophan fluorescence followed by a slow decrease. Further analysis by
order-of-mixing and quench experiments showed that the transient increase in fluorescence was
due to a conformational change coupled to DNA binding, while the subsequent decay was
concomitant with phosphodiester hydrolysis. The rate of the latter step varied with the
concentration of Mg2+ ions, but another Mg2+-dependent transition was observed upon the
addition of MgCl2 to a preformed enzyme--DNA complex. These results lead to a reaction scheme
in which one Mg2+ binds to the active site prior to phosphodiester hydrolysis but a second
Mg2+ is then needed to carry out the hydrolytic reaction. This scheme is correlated to the
crystal structures of the EcoRV endonuclease and its complexes with DNA and Mg2+ ions.

<>

<1>Bale, A., d'Alarcao, M., Marinus, M.G.
<2>Characterization of DNA adenine methylation mutants of Escherichia coli K12.
<3>Mutat. Res.
<4>59
<5>157-165
<6>1979
<7>The phenotypic traits of 7 independently isolated dam mutants of Escherichia coli have been
examined. The mutant strains differ from the wildtype in the following respects: (1) decreased
DNA adenine methylase activity in vivo and in vitro; (2) a 14--85-fold increase in spontaneous
mutability; (3) decreased survival after ultraviolet irradiation; (4) a 10--21-fold increase
in spontaneous induction of lambda phage from lysogens; (5) a 3--17-fold increase in the level
of recombination; and (6) inviability of double mutants containing dam- and recB- or recC-.
Unmethylated fd phage chromosomes are able to replicate normally in dam- mutants. A mutant
strain in which the dcm gene is deleted is viable, showing that the dcm gene product is
dispensible for growth.

<>

<1>Balendiran, K., Bonventre, J., Knott, R., Jack, W., Benner, J., Schildkraut, I., Anderson, J.E.
<2>Expression, purification, and crystallization of restriction endonuclease PvuII with DNA containing its recognition site.
<3>Proteins
<4>19
<5>77-79
<6>1994
<7>We have overexpressed the type II restriction endonuclease PvuII (R.PvuII) in E. coli,
prepared large amounts of the homogeneous enzyme, and crystallized it with an oligonucleotide
carrying a PvuII recognition site. The cocrystals are orthorhombic space group P212121 with
cell constants a = 95.8 A, b = 86.3 A, c = 48.5 A, and diffract X-rays to at least 2.7 A.
There is a complex of two protein subunits and one oligonucleotide duplex in the asymmetric
unit.

<>

<1>Balganesh, T.S., Reiners, L., Lauster, R., Noyer-Weidner, M., Wilke, K., Trautner, T.A.
<2>Construction and use of chimeric SPR/Phi3T DNA methyltransferases in the definition of sequence recognizing enzyme regions.
<3>EMBO J.
<4>6
<5>3543-3549
<6>1987
<7>Multispecific DNA methyltransferases (Mtases) of temperate Bacillus subtilis
phages SPR and Phi3T methylate the internal cytosine of the sequence GGCC.
They differ in their capacity to methylate additional sequences.  These are
CCGG anad CC(A/T)GG in SPR and GCNGC in Phi3T.  Introducing unique restriction
sites at equivalent locations within the two genes facilitated the construction
of chimeric genes.  These expressed Mtase activity at a level comparable to
that of the parental genes.  The methylation specificity of chimeric enzymes
was correlated with the location of chimeric fusions.  This analysis, which
also included the use of mutant genes, showed that domains involved in the
recognition of target sequences unique to each enezyme [CCGG, CC(A/T)GG or
GCNGC] are represented by the central non-conserved parts of the proteins,
whilst recognition of the sequence (GGCC), which is a target for both enzymes,
is determined by an adjacent conserved region.

<>

<1>Baliga, N.S., Bonneau, R., Facciotti, M.T., Pan, M., Glusman, G., Deutsch, E.W., Shannon, P., Chiu, Y., Weng, R.S., Gan, R.R., Hung, P., Date, S.V., Marcotte, E., Hood, L., Ng, W.V.
<2>Genome sequence of Haloarcula marismortui: A halophilic archaeon from the Dead Sea.
<3>Genome Res.
<4>14
<5>2221-2234
<6>2004
<7>We report the complete sequence of the 4,274,642-bp genome of Haloarcula marismortui, a
halophilic archaeal isolate from the Dead Sea. The genome
is organized into nine circular replicons of varying G+C compositions
ranging from 54% to 62%. Comparison of the genome architectures of
Halobacterium sp. NRC-1 and H. marismortui suggests a common ancestor for
the two organisms and a genome of significantly reduced size in the
former. Both of these halophilic archaea use the same strategy of high
surface negative charge of folded proteins as means to circumvent the
salting-out phenomenon in a hypersaline cytoplasm. A multitiered
annotation approach, including primary sequence similarities, protein
family signatures, structure prediction, and a protein function
association network, has assigned putative functions for at least 58% of
the 4242 predicted proteins, a far larger number than is usually achieved
in most newly sequenced microorganisms. Among these assigned functions
were genes encoding six opsins, 19 MCP and/or HAMP domain signal
transducers, and an unusually large number of environmental response
regulators-nearly five times as many as those encoded in Halobacterium sp.
NRC-1-suggesting H. marismortui is significantly more physiologically
capable of exploiting diverse environments. In comparing the physiologies
of the two halophilic archaea, in addition to the expected extensive
similarity, we discovered several differences in their metabolic
strategies and physiological responses such as distinct pathways for
arginine breakdown in each halophile. Finally, as expected from the larger
genome, H. marismortui encodes many more functions and seems to have fewer
nutritional requirements for survival than does Halobacterium sp. NRC-1.

<>

<1>Ball, C.A., Osuna, R., Ferguson, K.C., Johnson, R.C.
<2>Dramatic changes in Fis levels upon nutrient upshift in Escherichia coli.
<3>J. Bacteriol.
<4>174
<5>8043-8056
<6>1992
<7>Fis is a small basic DNA-binding protein from Escherichia coli that was identified because of
its role in site-specific DNA recombination
reactions. Recent evidence indicates that Fis also participates in
essential cell processes such as rRNA and tRNA transcription and
chromosomal DNA replication. In this report, we show that Fis levels vary
dramatically during the course of cell growth and in response to changing
environmental conditions. When stationary-phase cells are subcultured into
a rich medium, Fis levels increase from less than 100 to over 50,000
copies per cell prior to the first cell division. As cells enter
exponential growth, nascent synthesis is largely shut off, and
intracellular Fis levels decrease as a function of cell division. Fis
synthesis also transiently increases when exponentially growing cells are
shifted to a richer medium. The magnitude of the peak of Fis synthesis
appears to reflect the extent of the nutritional upshift. fis mRNA levels
closely resemble the protein expression pattern, suggesting that
regulation occurs largely at the transcriptional level. Two RNA
polymerase-binding sites and at least six high-affinity Fis-binding sites
are present in the fis promoter region. We show that expression of the fis
operon is negatively regulated by Fis in vivo and that purified Fis can
prevent stable complex formation by RNA polymerase at the fis promoter in
vitro. However, autoregulation only partially accounts for the expression
pattern of Fis. We suggest that the fluctuations in Fis levels may serve
as an early signal of a nutritional upshift and may be important in the
physiological roles Fis plays in the cell.

<>

<1>Ball, N., Streeter, S.D., Kneale, G.G., McGeehan, J.E.
<2>Structure of the restriction-modification controller protein C.Esp1396I.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>65
<5>900-905
<6>2009
<7>The controller protein of the Esp1396I restriction-modification (R-M) system binds
differentially to three distinct operator sequences
upstream of the methyltransferase (M) and endonuclease (R) genes to
regulate the timing of gene expression. The crystal structure of a
complex of the protein with two adjacent operator DNA sequences has
been reported; however, the structure of the free protein has not yet
been determined. Here, the crystal structure of the free protein is
reported, with seven dimers in the asymmetric unit. Two of the 14
monomers show an alternative conformation to the major conformer in
which the side chains of residues 43-46 in the loop region flanking the
DNA-recognition helix are displaced by up to 10 A. It is proposed that
the adoption of these two conformational states may play a role in
DNA-sequence promiscuity. The two alternative conformations are also
found in the R35A mutant structure, which is otherwise identical to the
native protein. Comparison of the free and bound protein structures
shows a 1.4 A displacement of the recognition helices when the dimer is
bound to its DNA target.

<>

<1>Ball, N.J., McGeehan, J.E., Streeter, S.D., Thresh, S.J., Kneale, G.G.
<2>The structural basis of differential DNA sequence recognition by restriction-modification controller proteins.
<3>Nucleic Acids Res.
<4>40
<5>10532-10542
<6>2012
<7>Controller (C) proteins regulate the expression of restriction-modification (RM)  genes in a
wide variety of RM systems. However, the RM system Esp1396I is of
particular interest as the C protein regulates both the restriction endonuclease
(R) gene and the methyltransferase (M) gene. The mechanism of this finely tuned
genetic switch depends on differential binding affinities for the promoters
controlling the R and M genes, which in turn depends on differential DNA sequence
recognition and the ability to recognize dual symmetries. We report here the
crystal structure of the C protein bound to the M promoter, and compare the
binding affinities for each operator sequence by surface plasmon resonance.
Comparison of the structure of the transcriptional repression complex at the M
promoter with that of the transcriptional activation complex at the R promoter
shows how subtle changes in protein-DNA interactions, underpinned by small
conformational changes in the protein, can explain the molecular basis of
differential regulation of gene expression.

<>

<1>Balmain, A.
<2>Exploring the bowels of DNA methylation.
<3>Curr. Biol.
<4>5
<5>1013-1016
<6>1995
<7>The role DNA methylation is thought to have in cancer has become increasingly complex.  In the
late 1970s and early 1980s, it was shown that methylation of CpG sites in DNA can exert a
negative influence on gene expression.  Treatment of fibroblastic 10T1/2 cells in culture with
demethylating agents, such as 5-azacytidine, facilitated the emergence of novel muscle or
brain-related cell types, presumably by removing methyl groups from developmentally important
genes, allowing their expression.  One of the genes responsible for the myogenesis programme,
MyoD, was subsequently found to be methylated in the original 10T1/2 cells, but became
demethylated (and expressed) in their muscle progeny.  Relevance to cancer development was
suggested by the observation that tumour cells tend to have hypomethylated DNA, and
consequently begin to express genes that could confer a growth advantage.  Obligingly, some
genes that are hypomethylated in cancers turned out to be oncogenes, completing a satisfying
hypothetical loop.  This concept gained further support with the observation that carcinogenic
alkylating agents can induce DNA hypomethylation, in addition to their known mutagenic
effects, possibly by preventing cytosine methylation at sites adjacent to O6-alkylguanosine
residues.  The conclusion to be drawn from these studies was that hypomethylation of tumour
DNA would allow uncontrolled or aberrant oncogene expression, with obvious advantages for
tumour cell growth.

<>

<1>Baltrus, D.A., Amieva, M.R., Covacci, A., Lowe, T.M., Merrell, D.S., Ottemann, K.M., Stein, M., Salama, N.R., Guillemin, K.
<2>The complete genome sequence of Helicobacter pylori strain G27.
<3>J. Bacteriol.
<4>191
<5>447-448
<6>2009
<7>Helicobacter pylori is a gram-negative pathogen that colonizes the stomachs of over half the
world's population and causes a spectrum of gastric diseases
including gastritis, ulcers, and gastric carcinoma. The H. pylori species
exhibits unusually high levels of genetic variation between strains. Here we
announce the complete genome sequence of H. pylori strain G27, which has been
used extensively in H. pylori research.

<>

<1>Baltrus, D.A., Yourstone, S., Lind, A., Guilbaud, C., Sands, D.C., Jones, C.D., Morris, C.E., Dangl, J.L.
<2>Draft Genome Sequences of a Phylogenetically Diverse Suite of Pseudomonas syringae Strains from Multiple Source Populations.
<3>Genome Announcements
<4>2
<5>e01195-13
<6>2014
<7>Here, we report the draft genome sequences for 7 phylogenetically diverse isolates of
Pseudomonas syringae, obtained from numerous environmental sources
and geographically proximate crop species. Overall, these sequences provide a
wealth of information about the differences (or lack thereof) between isolates
from disease outbreaks and those from other sources.

<>

<1>Baltz, R.H., McHenney, M.A.
<2>Transduction of plasmid DNA in Streptomyces spp.
<3>Genetics and Molecular Biology of Industrial Microorganisms, American Society for Microbiology, Hershberger, C.L., Queener, S.W., Megeman, G., Washington, DC
<4>0
<5>163-167
<6>1989
<7>None

<>

<1>Ban, C., Yang, W.
<2>Structural basis for MutH activation in E. coli mismatch repair and relationship of MutH to restriction endonucleases.
<3>EMBO J.
<4>17
<5>1526-1534
<6>1998
<7>MutS, MutL and MutH are the three essential proteins for initiation of methyl-directed DNA
mismatch repair to correct mistakes made during DNA replication in Escherichia coli.  MutH
cleaves a newly synthesized and unmethylated daughter strand 5' to the sequence d(GATC) in a
hemi-methylated duplex.  Activation of MutH requires the recognition of a DNA mismatch by MutS
and MutL.  We have crystallized MutH in two space groups and solved the structures at 1.7 and
2.3 Angstrom resolution, respectively.  The active site of MutH is located at an interface
between two subdomains that pivot relative to one another, as revealed by comparison of the
crystal structures, and this presumably regulates the nuclease activity.  The relative motion
of the two subdomains in MutH correlates with the position of a protruding C-terminal helix.
This helix appears to act as a molecular lever through which MutS and MutL may communicate the
detection of a DNA mismatch and activate MutH.  With sequence homology to Sau3AI and
structural similarity to PvuII endonuclease, MutH is clearly related to these enzymes by
divergent evolution, and this suggests that type II restriction endonucleases evolved from a
common ancestor.

<>

<1>Banas, J.A., Biswas, S., Zhu, M.
<2>Effects of DNA Methylation on Expression of Virulence Genes in Streptococcus mutans.
<3>Appl. Environ. Microbiol.
<4>77
<5>7236-7242
<6>2011
<7>Bacteria produce a variety of enzymes capable of methylating DNA. In many species, the
majority of adenine methylation is accomplished by
the DNA adenine methylase Dam. In Escherichia coli the Dam methylase
plays roles in the initiation of replication, mismatch repair, and gene
regulation. In a number of other bacterial species, mutation or
overexpression of Dam leads to attenuation of virulence. Homologues of
the dam gene exist in some members of the Firmicutes, including
Streptococcus mutans, a dental pathogen. An S. mutans strain
inactivated in the dam gene (SMU.504; here designated damA) was
engineered, and phenotypes linked to cariogenicity were examined. A
prominent observation was that the damA mutant produced greater amounts
of glucan than the parental strain. Real-time PCR confirmed
upregulation of gtfB. To determine whether other loci were affected by
the damA mutation, a microarray analysis was carried out. Seventy genes
were upregulated at least 2-fold in the damA mutant, and 33 genes were
downregulated at least 2-fold. In addition to gtfB (upregulated
2.6-fold; 1.7-fold when measured by real-time PCR), other upregulated
virulence factors included gbpC (upregulated 2.1-fold) and loci
predicted to encode bacteriocins (upregulated 2- to 7-fold). Various
sugar transport operons were also upregulated, the most extreme being
the cellobiose operon (upregulated nearly 40-fold). Expression of sacB,
encoding fructosyltransferase, was downregulated 2.4-fold. The sequence
5'-GATC-3' appeared to constitute the recognition sequence for
methylation. These results provide evidence that DNA methylation in S.
mutans has a global effect on gene expression, including that of genes
associated with cariogenic potential.

<>

<1>Banas, J.A., Ferretti, J.J., Progulske-Fox, A.
<2>Identification and sequence analysis of a methylase gene in Porphyromonas gingivalis.
<3>Nucleic Acids Res.
<4>19
<5>4189-4192
<6>1991
<7>A gene from the periodontal organism Porphyromonas gingivalis has been identified as encoding
a DNA methylase. The gene, referred to as pgiIM, has been sequenced and found to contain a
reading frame of 864 basepairs. The putative amino acid sequence of the encoded methylase was
288 amino acids, and shared 47% and 31% homology with the Streptococcus pneumoniae DpnII and
E. coli Dam methylases, respectively. The activity and specificity of the PgiI methylase
(M.PgiI) was confirmed by cloning the gene into a dam- strain of E. coli (JM110) and
performing a restriction analysis on the isolated DNA with enzymes whose activities depended
upon the methylation state of the DNA. The data indicated that M.PgiI, like DpnII and Dam,
methylated the adenine residue within the sequence 5'-GATC-3'.

<>

<1>Banas, J.A., Ferretti, J.J., Progulske-Fox, A.
<2>Identification of a methylase gene in Porphyromonas gingivalis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>174
<6>1991
<7>Sequence analysis of a portion of the P. gingivalis genome revealed an open reading frame
which encoded a protein whose putative amino acid sequence shared approximately fifty percent
and thirty percent identity with the Streptococcus pneumoniae Dpn methylase and Escherichia
coli Dam methylase, respectively. The P. gingivalis methylase was cloned into pUC18 and
transformed into E. coli JM110 (dam). Plasmid DNA isolated from transformants could be cut
with DpnI which only cuts at methylated 5'-GATC-3' sites, but not with NdeII which will not
cut at methylated 5'-GATC-3' sites. The methylase was expressed in both orientations. These
results suggest that the gene cloned from P. gingivalis specifies an adenine methylase that
recognizes the sequence 5'-GATC-3'.

<>

<1>Banavali, N.K., MacKerell, A.D. Jr.
<2>Free energy and structural pathways of base flipping in a DNA GCGC containing sequence.
<3>J. Mol. Biol.
<4>319
<5>141-160
<6>2002
<7>Structural distortions of DNA are essential for its biological function due to the genetic
information of DNA not being physically accessible in the duplex state. Base flipping is one
of the simplest structural distortions of DNA and may represent an initial event in strand
separation required to access the genetic code. Flipping is also utilized by DNA-modifying and
repair enzymes to access specific bases. It is typically thought that base flipping (or
base-pair opening) occurs via the major groove whereas minor groove flipping is only possible
when mediated by DNA-binding proteins. Here, umbrella sampling with a novel center-of-mass
pseudodihedral reaction coordinate was used to calculate the individual potentials of mean
force (PMF) for flipping of the Watson-Crick (WC) paired C and G bases in the CCATGCGCTGAC DNA
dodecamer. The novel reaction coordinate allowed explicit investigation of the complete
flipping process via both the minor and major groove pathways. The minor and major groove
barriers to flipping are similar for C base flipping while the major groove barrier is
slightly lower for G base flipping. Minor groove flipping requires distortion of the WC
partner while the flipping base pulls away from its partner during major groove flipping. The
flipped states are represented by relatively flat free energy surfaces, with a small, local
minimum observed for the flipped G base. Conserved patterns of phosphodiester backbone
dihedral distortions during flipping indicate their essential role in the flipping process.
During flipping, the target base tracks along the respective grooves, leading to
hydrogen-bonding interactions with neighboring base-pairs. Such hydrogen-bonding interactions
with the neighboring sequence suggest a novel mechanism of sequence dependence in DNA
dynamics.

<>

<1>Bancroft, I., Smith, R.J.
<2>An analysis of restriction endonuclease sites in cyanophages infecting the heterocystous cyanobacteria Anabaena and Nostoc.
<3>J. Gen. Virol.
<4>69
<5>739-743
<6>1988
<7>An analysis of restriction endonuclease cleavage of DNA isolated from cyanophages that infect
Anabaena and Nostoc species of cyanobacteria has provided evidence for counter-selection of
restriction endonuclease sites. These include sites containing subsequences which are
methylated by host (Anabaena PCC 7120) methylase(s) akin to the dam and dcm enzymes of
Escherichia coli. Other sites which are counter-selected have no common sequence structure.
The latter include those of the endogenous restriction endonucleases of the host, but other
absent sequences are not attributable to isoschizomers of any known Anabaena or Nostoc
restriction endonuclease. The cyanophages differ in their tolerance to DNA methylation.
Isolates A-4L, AN-13 and AN-23 do not tolerate adenosine methylation in the GATC sequence
whereas two cyanophages, A-1L and AN-10 (which are related) do tolerate dam-like methylation
of this sequence. In addition, A-1L allows cytosine methylation at GGCC sequences, but AN-10
has counter-selected these sequences and the remaining sites are not methylated. Analysis of
native and cloned A-4L DNA suggests that counter-selection has occurred against all sequences
which would be methylated by the host at either adenosine or cytosine nucleotides.

<>

<1>Bandaru, B.
<2>Study of cytosine to thymine mutations at the sites of dcm and M.HpaII methyltransferases.
<3>Diss. Abstr.
<4>57
<5>2535B-2536B
<6>1996
<7>It has been proposed that sites of methylation are "hotspots" for C to T mutations in
bacterial and human genomes.  I have developed a very sensitive genetic reversion assay that
quantifies the frequency of C to T mutations at various methylation sites and the repair of
resulting mismatches by DNA repair processes in Escherichia coli.  I have demonstrated that
the presence of HpaII methyltransferase (Mtase) in E. coli causes a substantial increase in C
to T mutations at CG sites.  This is similar to the known mutagenic effects of E. coli MTase
Dcm within its own recognition sequence.  Using this genetic system, I have tested a homolog
of an E. coli DNA repair gene in Haemophilus parainfluenzae for antimutagenic activity.
Unexpectedly, the homology was found to have little effect on the reversion frequency.  Using
this system, I have also shown that in vitro HpaII and SssI Mtases can convert cytosine to
uracil.  These studies define 5-methylcytosine as an intrinsic mutagen and further elaborate
the mutagenic potential of cytosine Mtases.  Multi-copy clones of E. coli DNA cytosine
methyltransferases (C5 Mtases) Dcm and M.EcoRII cause ~50 fold increase in C to T mutations at
their canonical site of methylation, 5'-CmeCAGG (meC is 5-methylcytosine).  I have observed
that these plasmids also cause transition mutations at the second cytosine in the sequences
CCGGG at ~10-fold lower frequency.  Similarly, M.HpaII was found to cause a significant
increase in C to T mutations at a CCAG site -- in addition to causing mutations at its
canonical site of methylation, CCGG.  Using a plasmid that substantially overprodces M.EcoRII,
I have shown that in vivo methylation at CCSGG (S is C or G) and other non-canonical sites
could be detected using a gel electrophoretic assay.  There is a direct correlation between
the level of M.EcoRII activity in cells, the extent of methylation at non-canonical sites and
frequency of mutations at these same sites.  Overproduction of M.EcoRII in cells also causes
degradation of DNA and induction of the SOS response.  I have shown that in vitro, M.EcoRII
methylates an oligonucleotide duplex containing a CCGGG site at a slow rate suggesting that
overproduction of the enzyme is essential for significant amounts of such methylation to
occur.  Together these results show that C5 Mtases occasionally methylate cellular DNA at
non-canonical sites and suggest that in E. coli methylation-specific restriction systems and
DNA mismatch correction systems may have evolved to accommodate this fact.  These results also
suggest that mutational effects of cytosine methyltransferases may be much broader than
previously imagined.

<>

<1>Bandaru, B., Gopal, J., Bhagwat, A.S.
<2>Overproduction of DNA cytosine methyltransferases causes methylation and C-T mutations at non-canonical sites.
<3>J. Biol. Chem.
<4>271
<5>7851-7859
<6>1996
<7>Multicopy clones of Escherichia coli cytosine methyltransferases Dcm and EcoRII methylase (M.
EcoRII) cause ~50-fold increase in C-T mutations at their canonical site of methylation.
5'-CmeCAGG (meC is 5-methylcytosine).  These plasmids also cause transition mutations at the
second cytosine in the sequences CCGGG at ~10-fold lower frequency.  Similarly, M.HpaII was
found to cause a significant increase in C-T mutations at a CCAG site, in addition to causing
mutations at its canonical site of methylation, CCGG.  Using a plasmid that substantially
overproduces M.EcoRII, in vivo methylation at CCSGG (S is C or G) and other noncanonical sites
could be detected using a gel electrophoretic assay.  There is a direct correlation between
the level of M.EcoRII activity in cells, the extent of methylation at non-canonical sites and
frequency of mutations at these same sites.  Overproduction of M.EcoRII in cells also causes
degradation of DNA and induction of the SOS response.  In vitro, M. EcoRII methylates an
oligonucleotide duplex containing a CCGGG site at a slow rate, suggesting that overproduction
of the enzyme is essential for significant amounts of such methylation to occur.  Together
these results show that cytosine methyltransferases occasionally methylate cellular DNA at
non-canonical sites and suggest that in E. coli, methylation-specific restriction systems and
sequence specificity of the DNA mismatch correction systems may have evolved to accommodate
this fact.  These results also suggest that mutational effects of cytosine methyltransferases
may be much broader than previously imagined.

<>

<1>Bandaru, B., Wyszynski, M., Bhagwat, A.S.
<2>HpaII methyltransferase is mutagenic in Escherichia coli.
<3>J. Bacteriol.
<4>177
<5>2950-2952
<6>1995
<7>A genetic reversion assay to study C-to-T mutations within CG sites in DNA is described. It
was used to demonstrate that the presence of HpaII methyltransferase (MTase) in Escherichia
coli causes a substantial increase in C-to-T mutations at CG sites. This is similar to the
known mutagenic effects of E. coli MTase Dcm within its own recognition sequence. With this
genetic system, a homolog of an E. coli DNA repair gene in Haemophilus parainfluenzae was
tested for antimutagenic activity. Unexpectedly, the homolog was found to have little effect
on the reversion frequency. The system was also used to show that HpaII and SssI MTases can
convert cytosine to uracil in vitro. These studies define 5-methylcytosine as an intrinsic
mutagen and further elaborate the mutagenic potential of cytosine MTases.

<>

<1>Bandyopadhyay, P.K., Studier, F.W., Hamilton, D.L., Yuan, R.
<2>Inhibition of the type I restriction-modification enzymes EcoB and EcoK by the gene 0.3 protein of bacteriophage T7.
<3>J. Mol. Biol.
<4>182
<5>567-578
<6>1985
<7>The gene 0.3 protein of bacteriophage T7 is a potent inhibitor of the
restriction-modification enzymes EcoB and EcoK, both in vivo and in vitro.  We have
analyzed the ability of purified 0.3 protein to inhibit different steps in the reactions of
EcoB
and EcoK with DNA.  Most of our experiments were done with EcoK, but selected tests
with EcoB indicate that the two enzymes are affected by 0.3 protein in the same way.
Purified 0.3 protein binds tightly to free enzyme, apparently to one of the small subunits,
and prevents it from binding to DNA.  If EcoK is allowed to form specific recognition
complexes with unmodified DNA before 0.3 protein is added, relatively low levels of 0.3
protein prevent the nuclease activity that would otherwise appear upon addition of ATP, but
considerably higher levels are needed to prevent formation of filter-binding complexes or
ATPase activity.  This, together with other results, suggests that the binding site for 0.3
protein is protected in recognition complexes and in the early stages of the ATP-stimulated
reactions, but that it becomes accessible again before cleavage of the DNA, perhaps after
the translocation step.  If added after the nuclease reaction is substantially over, 0.3
protein
has little effect on ATPase activity, and indeed, the subunit having the binding site for 0.3
protein apparently dissociates from the enzyme-DNA complex.  The methylase activity of
EcoK on hemi-methylated recognition sites is strongly inhibited by 0.3 protein added at any
stage of the reaction.

<>

<1>Bandyopadhyay, R., Das, J.
<2>The DNA adenine methyltransferase-encoding gene (dam) of Vibrio cholerae.
<3>Gene
<4>140
<5>67-71
<6>1994
<7>The DNA adenine methyltransferase (MTase)-encoding gene (dam) of Vibrio cholerae, an organism
belonging to the family Vibrionaceae, has been cloned and the complete nucleotide (nt)
sequence determined. V. cholerae dam encodes a 21.5-kDa protein and is directly involved in
methyl-directed DNA mismatch repair. It can substitute for the Escherichia coli enzyme and can
suppress the phenotypic traits associated with E. coli dam mutants. Overproduction of V.
cholerae Dam MTase does not result in hypermutability in either V. cholerae or E. coli cells.
Overproduction of V. cholerae Dam in a pUC plasmid, however, fails to suppress the
2-aminopurine (2-AP0-sensitive phenotype of E. coli dam mutants. Homology between the nt and
deduced amino acid (aa) sequences of the E. coli and V. cholerae dam genes is only 30-35%.

<>

<1>Bandyopadhyay, R., Sengupta, A., Das, J.
<2>A mutation in the dam gene of Vibrio cholerae: 2-aminopurine sensitivity with intact GATC methylase activity.
<3>Biochem. Biophys. Res. Commun.
<4>165
<5>561-567
<6>1989
<7>Vibrio cholerae mutants sensitive to 2-aminopurine (2AP) but with DNA adenine methylase
activity similar to parental cells have been isolated. The mutant strains were sensitive to
ultraviolet light (UV), methyl methane sulphonate (MMS) and 9-aminoacridine. The spontaneous
mutation frequency of the mutants were not significantly affected. Attempts to isolate dam V.
cholerae cells by screening 2AP sensitive cells have not been successful. All of the mutant
phenotypes could be suppressed by introducing the plasmid pRB103 carrying the dam gene of
Escherichia coli into the mutant cells.

<>

<1>Banerjee, A., Rao, D.N.
<2>Functional analysis of an acid adaptive DNA adenine methyltransferase from Helicobacter pylori 26695.
<3>PLoS ONE
<4>6
<5>e16810
<6>2011
<7>HP0593 DNA-(N(6)-adenine)-methyltransferase (HP0593 MTase) is a member of a Type  III
restriction-modification system in Helicobacter pylori strain 26695. HP0593
MTase has been cloned, overexpressed and purified heterologously in Escherichia
coli. The recognition sequence of the purified MTase was determined as
5'-GCAG-3'and the site of methylation was found to be adenine. The activity of
HP0593 MTase was found to be optimal at pH 5.5. This is a unique property in
context of natural adaptation of H. pylori in its acidic niche. Dot-blot assay
using antibodies that react specifically with DNA containing m6A modification
confirmed that HP0593 MTase is an adenine-specific MTase. HP0593 MTase occurred
as both monomer and dimer in solution as determined by gel-filtration
chromatography and chemical-crosslinking studies. The nonlinear dependence of
methylation activity on enzyme concentration indicated that more than one
molecule of enzyme was required for its activity. Analysis of initial velocity
with AdoMet as a substrate showed that two molecules of AdoMet bind to HP0593
MTase, which is the first example in case of Type III MTases. Interestingly,
metal ion cofactors such as Co(2+), Mn(2+), and also Mg(2+) stimulated the HP0593
MTase activity. Preincubation and isotope partitioning analyses clearly indicated
that HP0593 MTase-DNA complex is catalytically competent, and suggested that DNA
binds to the MTase first followed by AdoMet. HP0593 MTase shows a distributive
mechanism of methylation on DNA having more than one recognition site.
Considering the occurrence of GCAG sequence in the potential promoter regions of
physiologically important genes in H. pylori, our results provide impetus for
exploring the role of this DNA MTase in the cellular processes of H. pylori.

<>

<1>Banerjee, S., Chowdhury, R.
<2>An orphan DNA (cytosine-5-)-methyltransferase in Vibrio cholerae.
<3>Microbiology
<4>152
<5>1055-1062
<6>2006
<7>5-Methyl cytosine (m5C was detected in genomic DNA of the enteric pathogen Vibrio cholerae by
HPLC analysis and immunoblotting with
m5C-specific antibody. Although cleavage with the restriction
endonuclease EcoRII revealed the absence of a Dcm homologue in V.
cholerae, analysis of the genome sequence indicated the presence of a
gene, designated in this study as vchM, which encodes a DNA
(cytosine-5-)-methyltransferase (m5C-MTase) designated M.Vch. M.Vch is
not associated with a restriction endonuclease or a mismatch very short
patch repair (Vsr)-like endonuclease and is hence an 'orphan' or
solitary MTase, although analysis of a phylogenetic tree indicated that
related cytosine MTases are all components of restriction-modification
systems. M.Vch recognizes and methylates the first 5' C in the
degenerate sequence 5'-RCCGGY-3'. RT-PCR analysis suggested that vchM
gene expression is increased during the stationary phase of growth.
During stationary phase, the spontaneous mutation frequency in the V.
cholerae wild-type strain was significantly higher than in the
corresponding vchM mutant strain, suggesting that the presence of M.Vch
and the absence of a very short patch (VSP) repair-like system imposes
upon V. cholerae a mutator phenotype.

<>

<1>Banerjee, S., Fisher, O., Lohia, A., Ankri, S.
<2>Entamoeba histolytica DNA methyltransferase (Ehmeth) is a nuclear matrix protein that binds EhMRS2, a DNA that includes a scaffold/matrix  attachment region (S/MAR).
<3>Mol. Biochem. Parasitol.
<4>139
<5>91-97
<6>2005
<7>The protozoan parasite Entamoeba histolytica express a cytosine-5 DNA methyltransferase
(Ehmeth) that belongs to the DNMT2 protein family.
The biological function of members of this DNMT2 family is unknown. In
the present study, we have demonstrated that Ehmeth is a nuclear matrix
protein. Indeed, we showed by south-western analysis and yeast
one-hybrid system that Ehmeth binds to EhMRS2, a DNA element which
contains the eukaryotic consensus scaffold/inatrix attachment regions
(S/MAR) bipartite recognition sequences. S/MARs have been implicated in
a variety of important functions, such as genome organization and gene
expression. The methylation status of cytosine located within EhMRS2
was analyzed by bisulfite genomic sequencing. We observed the presence
of methylated cytosine within the 3'-end of EhMRS2. These data provide
the first evidence that a member of the DNMT2 family interacts with a
S/MAR containing DNA element.

<>

<1>Banfalvi, G., Sarkar, N.
<2>Effect of mercury substitution of DNA on its susceptibility to cleavage by restriction endonucleases.
<3>DNA Cell Biol.
<4>14
<5>445-450
<6>1995
<7>Mercurated DNA was synthesized in vitro by substituting Hg-dCTP or Hg-dUMP for dCTP in one
strand of M13mp8DNA in a DNA polymerase I reaction. Restriction enzymes, including SmaI, PstI,
BamHI, HindIII, and HincII, were completely inactive when their recognition sites were fully
substituted with Hg-dCMP, while Hg-dUMP containing DNA was hydrolyzed to some extent. Under
conditions favoring star activities, or when DNA was substituted with a low level of
mercury-nucleotide, DNA was cleaved by restriction enzymes. Susceptibility to degrading and
synthesizing enzymes and insensitivity to restriction endonucleases of fully mercurated DNA
makes mercuration an attractive molecular "tag" for in vitro manipulation and selective
isolation of Hg-DNA.

<>

<1>Banks, D.J., Porcella, S.F., Barbian, K.D., Beres, S.B., Philips, L.E., Voyich, J.M., DeLeo, F.R., Martin, J.M., Somerville, G.A., Musser, J.M.
<2>Progress toward Characterization of the Group A Streptococcus Metagenome: Complete Genome Sequence of a Macrolide-Resistant Serotype M6 Strain.
<3>J. Infect. Dis.
<4>190
<5>727-738
<6>2004
<7>We describe the genome sequence of a macrolide-resistant strain (MGAS10394) of serotype M6
group A Streptococcus (GAS). The genome is
1,900,156 bp in length, and 8 prophage-like elements or remnants compose
12.4% of the chromosome. A 8.3-kb prophage remnant encodes the SpeA4
variant of streptococcal pyrogenic exotoxin A. The genome of strain
MGAS10394 contains a chimeric genetic element composed of prophage genes
and a transposon encoding the mefA gene conferring macrolide resistance.
This chimeric element also has a gene encoding a novel surface-exposed
protein (designated "R6 protein"), with an LPKTG cell-anchor motif located
at the carboxyterminus. Surface expression of this protein was confirmed
by flow cytometry. Humans with GAS pharyngitis caused by serotype M6
strains had antibody against the R6 protein present in convalescent, but
not acute, serum samples. Our studies add to the theme that GAS
prophage-encoded extracellular proteins contribute to host-pathogen
interactions in a strain-specific fashion.

<>

<1>Banks, D.J., Porcella, S.F., Barbian, K.D., Martin, J.M., Musser, J.M.
<2>Structure and distribution of an unusual chimeric genetic element encoding macrolide resistance in phylogenetically diverse clones of group A Streptococcus.
<3>J. Infect. Dis.
<4>188
<5>1899-1909
<6>2003
<7>The resistance of group A Streptococcus (GAS) to macrolide antibiotics is now a worldwide
problem. Preliminary sequencing of the genome of an
erythromycin-resistant serotype M6 clone that was responsible for a
pharyngitis outbreak in Pittsburgh, Pennsylvania, was conducted to
determine the structure of the genetic element containing the mefA gene,
which encodes a macrolide efflux protein. The mefA gene is associated with
a 58.8-kb chimeric genetic element composed of a transposon inserted into
a prophage. This element also encodes a putative extracellular protein
with a cell-wall anchoring motif (LPKTG) located at the carboxyterminus.
The mefA element was present in phylogenetically diverse GAS strains
isolated throughout the United States. Culture supernatants, prepared
after mitomycin C treatment, of a strain representing the outbreak clone
contained mefA element DNA in a DNAse-resistant form. Together, these data
provide new information about the molecular genetic basis of macrolide
resistance and dissemination in GAS strains.

<>

<1>Bannam, T.L., Teng, W.L., Bulach, D., Lyras, D., Rood, J.I.
<2>Functional Identification of Conjugation and Replication Regions of the Tetracycline Resistance Plasmid pCW3 from Clostridium perfringens.
<3>J. Bacteriol.
<4>188
<5>4942-4951
<6>2006
<7>Clostridium perfringens causes fatal human infections, such as gas
gangrene, as well as gastrointestinal diseases in both humans and animals.
Detailed molecular analysis of the tetracycline resistance plasmid pCW3
from C. perfringens has shown that it represents the prototype of a unique
family of conjugative antibiotic resistance and virulence plasmids. We
have identified the pCW3 replication region by deletion and transposon
mutagenesis and showed that the essential rep gene encoded a basic protein
with no similarity to any known plasmid replication proteins. An 11-gene
conjugation locus containing 5 genes that encoded putative proteins with
similarity to proteins from the conjugative transposon Tn916 was
identified, although the genes' genetic arrangements were different.
Functional genetic studies demonstrated that two of the genes in this
transfer clostridial plasmid (tcp) locus, tcpF and tcpH, were essential
for the conjugative transfer of pCW3, and comparative analysis confirmed
that the tcp locus was not confined to pCW3. The conjugation region was
present on all known conjugative plasmids from C. perfringens, including
an enterotoxin plasmid and other toxin plasmids. These results have
significant implications for plasmid evolution, as they provide evidence
that a nonreplicating Tn916-like element can evolve to become the
conjugation locus of replicating plasmids that carry major virulence genes
or antibiotic resistance determinants.

<>

<1>Bannam, T.L., Yan, X.X., Harrison, P.F., Seemann, T., Keyburn, A.L., Stubenrauch, C., Weeramantri, L.H., Cheung, J.K., McClane, B.A., Boyce, J.D., Moore, R.J., Rood, J.I.
<2>Necrotic Enteritis-Derived Clostridium perfringens Strain with Three Closely Related Independently Conjugative Toxin and Antibiotic Resistance Plasmids.
<3>MBio
<4>2
<5>e00190-11
<6>2011
<7>The pathogenesis of avian necrotic enteritis involves NetB, a pore-forming
toxin produced by virulent avian isolates of Clostridium perfringens type
A. To determine the location and mobility of the netB structural gene, we
examined a derivative of the tetracycline-resistant necrotic enteritis
strain EHE-NE18, in which netB was insertionally inactivated by the
chloramphenicol and thiamphenicol resistance gene catP. Both tetracycline
and thiamphenicol resistance could be transferred either together or
separately to a recipient strain in plate matings. The separate
transconjugants could act as donors in subsequent matings, which
demonstrated that the tetracycline resistance determinant and the netB
gene were present on different conjugative elements. Large plasmids were
isolated from the transconjugants and analyzed by high-throughput
sequencing. Analysis of the resultant data indicated that there were
actually three large conjugative plasmids present in the original strain,
each with its own toxin or antibiotic resistance locus. Each plasmid
contained a highly conserved 40-kb region that included plasmid
replication and transfer regions that were closely related to the 47-kb
conjugative tetracycline resistance plasmid pCW3 from C. perfringens. The
plasmids were as follows: (i) a conjugative 49-kb tetracycline resistance
plasmid that was very similar to pCW3, (ii) a conjugative 82-kb plasmid
that contained the netB gene and other potential virulence genes, and
(iii) a 70-kb plasmid that carried the cpb2 gene, which encodes a
different pore-forming toxin, beta2 toxin. IMPORTANCE: The anaerobic
bacterium Clostridium perfringens can cause an avian gastrointestinal
disease known as necrotic enteritis. Disease pathogenesis is not well
understood, although the plasmid-encoded pore-forming toxin NetB, is an
important virulence factor. In this work, we have shown that the plasmid
that carries the netB gene is conjugative and has a 40-kb region that is
very similar to replication and transfer regions found within each of the
sequenced conjugative plasmids from C. perfringens. We also showed that
this strain contained two additional large plasmids that were also
conjugative and carried a similar 40-kb region. One of these plasmids
encoded beta2 toxin, and the other encoded tetracycline resistance. To our
knowledge, this is the first report of a bacterial strain that carries
three closely related but different independently conjugative plasmids.
These results have significant implications for our understanding of the
transmission of virulence and antibiotic resistance genes in pathogenic
bacteria.

<>

<1>Bannantine, J.P., Bayles, D.O., Robbe-Austerman, S., Burrell, A.M., Stabel, J.R.
<2>Draft Genome Sequence of a Mycobacterium avium Complex Isolate from a Broadbill Bird.
<3>Genome Announcements
<4>2
<5>e01268-13
<6>2014
<7>We report the draft genome sequence of a Mycobacterium avium complex isolate. This isolate has
an estimated genome size of 5.1 Mb with an average GC content of
68.9% and is predicted to carry 4,497 protein-encoding genes and 317 pseudogenes.

<>

<1>Bannantine, J.P., Li, L., Mwangi, M., Cote, R., Raygoza, G.J.A., Kapur, V.
<2>Complete Genome Sequence of Mycobacterium avium subsp. paratuberculosis, Isolated from Human Breast Milk.
<3>Genome Announcements
<4>2
<5>e01252-13
<6>2014
<7>Mycobacterium avium subsp. paratuberculosis is the etiologic agent of Johne's disease in
ruminants and has also been associated with human Crohn's disease. We
report the complete genome sequence of M. avium subsp. paratuberculosis, isolated
from the breast milk of a Crohn's disease patient. This sequence has high
identity with characterized strains recovered from cattle.

<>

<1>Bannister, D.
<2>Analysis of an R+ strain carrying two fi- sex factors.
<3>J. Gen. Microbiol.
<4>61
<5>273-281
<6>1970
<7>An Escherichia coli K12 strain, J5-3 (R313), was presumed to carry a fi-R factor, R313, which
determined resistance to the drugs tetracycline, streptomycin and sulphonamide, and also
determined a host specificity, hsII.  When this strain was used as a donor of drug resistance,
separation of R factor-carried determinants was observed in ex-conjugants.  Resistance to Tc
and hsII character were not separated, nor was resistance to Sm separated from resistance to
Su, but separation of these two pairs of characters was observed.  Tetracycline-sensitive
segregants, obtained by penicillin selection, were resistant to Sm and Su, but had lost the
hsII character.  Similarly, segregants for selected sensitivity to Su were sensitive to Sm,
but resistant to Tc and hsII+.  The same pattern of separation of characters was also observed
when drug resistance was transduced with phage PI, with the additional finding that the Sm and
Su resistant transductants lacked sex factor activity.  The resistance to Sm carried by these
transductants could be mobilized by an fi-R factor, R143.  Explanations of this behavior are
considered, including the possibility that the strain J5-3 (R313) had carried two fi-R
factors.  This explanation would also require that the transduction of an R factor by phage PI
is not always complete.

<>

<1>Bannister, D., Glover, S.W.
<2>Restriction and modification of bacteriophages by R+ strains of Escherichia coli K12.
<3>Biochem. Biophys. Res. Commun.
<4>30
<5>735-738
<6>1968
<7>None

<>

<1>Bannister, D., Glover, S.W.
<2>The isolation and properties of non-restricting mutants of two different host specificities associated with drug resistance factors.
<3>J. Gen. Microbiol.
<4>61
<5>63-71
<6>1970
<7>Non-restricting (r-) mutants of two different host specificities carried by
resistance transfer factors have been isolated.  As previously found with other
host specificities, the non-restricting mutants were of two phenotypes:  those
that retained the ability to modify DNA, and those which had lost the ability
to modify DNA.  These mutants were tested for complementation with wild-type
host specificities carried either on the Escherichia coli chromosome, on
resistance transfer factors, or on the phage PI.  No complementation was
observed and possible explanations for this finding are considered.

<>

<1>Bao, G., Wang, R., Zhu, Y., Dong, H., Mao, S., Zhang, Y., Chen, Z., Li, Y., Ma, Y.
<2>Complete genome sequence of Clostridium acetobutylicum DSM 1731, a solvent-producing strain with multireplicon genome architecture.
<3>J. Bacteriol.
<4>193
<5>5007-5008
<6>2011
<7>Clostridium acetobutylicum is an important microorganism for solvent production. We report the
complete genome sequence of C. acetobutylicum
DSM 1731, a genome with multi-replicon architecture. Comparison with the
sequenced type strain C. acetobutylicum ATCC 824, the genome of strain
DSM1731 harbors a 1.7 kb insertion and a novel 11.1 kb plasmid, which
might be acquired during evolution.

<>

<1>Bao, G., Zhang, Y., Du C, C.Z., Li, Y., Cao, Z., Ma, Y.
<2>Genome Sequence of Klebsiella oxytoca M5al, a Promising Strain for Nitrogen Fixation and Chemical Production.
<3>Genome Announcements
<4>1
<5>e00074-12
<6>2013
<7>Klebsiella oxytoca is an important microorganism for nitrogen fixation and chemical
production. Here, we report an annotated draft genome of K. oxytoca
strain M5al that contains 5,256 protein-coding genes and 95 structural RNAs,
which provides a genetic basis for a better understanding of the physiology of
this species.

<>

<1>Bao, H.X., Tang, L., Yu, L., Wang, X.Y., Li, Y., Deng, X., Li, Y.G., Li, A., Zhu, D.L., Johnston, R.N., Liu, G.R., Feng, Y., Liu, S.L.
<2>Differential efficiency in exogenous DNA acquisition among closely related Salmonella strains: implications in bacterial speciation.
<3>BMC Microbiol.
<4>14
<5>157
<6>2014
<7>BACKGROUND: Acquisition of exogenous genetic material is a key event in bacterial
speciation. It seems reasonable to assume that recombination of the incoming DNA
into genome would be more efficient with higher levels of relatedness between the
DNA donor and recipient. If so, bacterial speciation would be a smooth process,
leading to a continuous spectrum of genomic divergence of bacteria, which,
however, is not the case as shown by recent findings. The goal of this study was
todetermine if DNA transfer efficiency is correlated with the levels of sequence
identity. RESULTS: To compare the relative efficiency of exogenous DNA
acquisition among closely related bacteria, we carried out phage-mediated
transduction and plasmid-mediated transformation in representative Salmonella
strains with different levels of relatedness. We found that the efficiency was
remarkably variable even among genetically almost identical bacteria. Although
there was a general tendency that more closely related DNA donor-recipient pairs
had higher transduction efficiency, transformation efficiency exhibited over a
thousand times difference among the closely related Salmonella strains.
CONCLUSION: DNA acquisition efficiency is greatly variable among bacteria that
have as high as over 99% identical genetic background, suggesting that bacterial
speciation involves highly complex processes affected not only by whether
beneficial exogenous DNA may exist in the environment but also the "readiness" of
the bacteria to accept it.

<>

<1>Bao, P., Su, J.Q., Hu, Z.Y., Haggblom, M.M., Zhu, Y.G.
<2>Genome sequence of the anaerobic bacterium Bacillus sp. strain ZYK, a selenite and nitrate reducer from paddy soil.
<3>Standards in Genomic Sciences
<4>9
<5>646-654
<6>2014
<7>Bacillus sp. strain ZYK, a member of the phylum Firmicutes, is of interest for its ability to
reduce nitrate and selenite and for its resistance to arsenic
under anaerobic conditions. Here we describe some key features of this organism,
together with the complete genome sequence and annotation. The 3,575,797 bp long
chromosome with its 3,454 protein-coding and 70 RNA genes, and the information
gained from its sequence will be relevant to the elucidation of
microbially-mediated transformations of nitrogen, selenium and arsenic in paddy
soil.

<>

<1>Bao, Q., Tian, Y., Li, W., Xu, Z., Xuan, Z., Hu, S., Dong, W., Yang, J., Chen, Y., Xue, Y., Xu, Y., Lai, X., Huang, L., Dong, X., Ma, Y., Ling, L., Tan, H., Chen, R., Wang, J., Yu, J., Yang, H.
<2>A complete sequence of the T. tengcongensis genome.
<3>Genome Res.
<4>12
<5>689-700
<6>2002
<7>Thermoanaerobacter tengcongensis is a rod-shaped, gram-negative, anaerobic
eubacterium that was isolated from a freshwater hot spring in Tengchong,
China. Using a whole-genome-shotgun method, we sequenced its 2,689,445-bp
genome from an isolate, MB4(T) (Genbank accession no. AE008691). The
genome encodes 2588 predicted coding sequences (CDS). Among them, 1764
(68.2%) are classified according to homology to other documented proteins,
and the rest, 824 CDS (31.8%), are functionally unknown. One of the
interesting features of the T. tengcongensis genome is that 86.7% of its
genes are encoded on the leading strand of DNA replication. Based on
protein sequence similarity, the T. tengcongensis genome is most similar
to that of Bacillus halodurans, a mesophilic eubacterium, among all fully
sequenced prokaryotic genomes up to date. Computational analysis on genes
involved in basic metabolic pathways supports the experimental discovery
that T. tengcongensis metabolizes sugars as principal energy and carbon
source and utilizes thiosulfate and element sulfur, but not sulfate, as
electron acceptors. T. tengcongensis, as a gram-negative rod by empirical
definitions (such as staining), shares many genes that are characteristics
of gram-positive bacteria whereas it is missing molecular components
unique to gram-negative bacteria. A strong correlation between the G + C
content of tDNA and rDNA genes and the optimal growth temperature is found
among the sequenced thermophiles. It is concluded that thermophiles are a
biologically and phylogenetically divergent group of prokaryotes that have
converged to sustain extreme environmental conditions over evolutionary
timescale. Full author list: Bao, Q., Tian, Y., Li, W., Xu, Z., Xuan, Z., Hu, S., Dong, W.,
Yang, J., Chen, Y., Xue, Y., Xu, Y., Lai, X., Huang, L., Dong, X., Ma, Y., Ling, L., Tan, H.,
Chen, R., Wang, J., Yu, J., Yang, H.

<>

<1>Bao, Y., Higgins, L., Zhang, P., Chan, S.H., Laget, S., Sweeney, S., Lunnen, K., Xu, S.Y.
<2>Expression and purification of BmrI restriction endonuclease and its N-terminal cleavage domain variants.
<3>Protein Expr. Purif.
<4>58
<5>42-52
<6>2008
<7>BmrI (ACTGGG N5/N4) is one of the few metal-independent restriction endonucleases (REases)
found in bacteria. The BmrI
restriction-modification system was cloned by the methylase selection
method, inverse PCR, and PCR. BmrI REase shows significant amino acid
sequence identity to BfiI and a putative endonuclease MspBNCORF3798 from
the sequenced Mesorhizobium sp. BNC1 genome. The EDTA-resistant BmrI REase
was successfully over-expressed in a pre-modified E. coli strain from
pET21a or pBAC-expIQ vectors. The recombinant BmrI REase shows strong
promiscuous activity (star activity) in NEB buffers 1, 4, and an EDTA
buffer. Star activity was diminished in buffers with 100-150mM NaCl and
10mM MgCl(2). His-tagged BmrI192, the N-terminal cleavage domain of BmrI,
was expressed in E. coli and purified from inclusion bodies. The refolded
BmrI192 protein possesses non-specific endonuclease activity. BmrI192
variants with a single Ser to Cys substitution (S76C or S90C) and BmrI200
(T200C) with a single Cys at the C-terminal end were also constructed and
purified. BmrI200 digests both single-strand (ss) and double-strand (ds)
DNA and the nuclease activity on ss DNA is at least 5-fold higher than
that on ds DNA. The Cys-containing BmrI192 and BmrI200 nuclease variants
may be useful for coupling to other DNA binding elements such as synthetic
zinc fingers, thio-containing locked nucleic acids (LNA) or peptide
nucleic acids (PNA).

<>

<1>Bao, Y., Kwok, A.H., He, L., Jiang, J., Huang, Z., Leung, F.C., Sheng, X.
<2>Complete Genome Sequence of Dyella jiangningensis Strain SBZ3-12, Isolated from the Surfaces of Weathered Rock.
<3>Genome Announcements
<4>2
<5>e00416-14
<6>2014
<7>Dyella jiangningensis strain SBZ3-12 can weather biotite and release Al and Fe from biotite
under nutrient-poor conditions. Here, we report the first complete
genome sequence of D. jiangningensis strain SBZ3-12, which may facilitate a
better understanding of the molecular mechanism behind mineral weathering.

<>

<1>Bao, Y., Liang, Z., Booyjzsen, C., Mayfield, J.A., Li, Y., Lee, S.W., Ploplis, V.A., Song, H., Castellino, F.J.
<2>Unique Genomic Arrangements in an Invasive Serotype M23 strain of Streptococcus pyogenes Identify Genes that Induce Hypervirulence.
<3>J. Bacteriol.
<4>196
<5>4089-4102
<6>2014
<7>The first genome sequence of a Group A Streptococcus pyogenes M23 (emm23)
serotype (M23ND), isolated from an invasive human infection, has been completed.
This opacity factor-negative (SOF-) strain is composed of a circular chromosome
of 1,846,477 bp. Gene profiling showed that this strain contained six
phage-encoded and 24 chromosomally-inherited well-known virulence factors, as
well as 11 pseudogenes. The bacterium has acquired four large prophage elements,
PhiM23ND.1-4, harboring genes encoding streptococcal superantigen (ssa),
streptococcal pyrogenic exotoxins (spe) C, H, and I, and DNases spd1 and spd3,
with phage integrase genes present at one flank of each phage insertion,
suggesting that the phages were integrated by horizontal gene transfer.
Comparative analyses revealed unique large-scale genomic rearrangements that
differ from previously sequenced GAS strains. These rearrangements resulted in an
imbalanced genomic architecture and translocations of chromosome-encoded
virulence genes. The covS sensor in M23ND was identified as a pseudogene,
resulting in attenuation of speB function and increased expression of the
chromosomal virulence factors, mga, emm23, scpA, fibronectin-binding proteins
(sfb1 and fbp54), streptolysin O (slo), hyaluronic acid capsule (hasA),
streptokinase (ska), and a 2 DNases (spd and spd3), which were verified by PCR.
These genes are responsible for facilitating host epithelial cell binding and
and/or immune evasion, thus further contributing to the virulence of M23ND. In
conclusion, strain M23ND has become highly pathogenic as the result of a
combination of multiple genetic factors, particularly gene composition and
mutations, prophage integrations, unique genomic rearrangements, and regulated
expression of critical virulence factors.

<>

<1>Baranasic, D., Gacesa, R., Starcevic, A., Zucko, J., Blazic, M., Horvat, M., Gjuracic, K., Fujs, S., Hranueli, D., Kosec, G., Cullum, J., Petkovic, H.
<2>Draft Genome Sequence of Streptomyces rapamycinicus Strain NRRL 5491, the Producer of the Immunosuppressant Rapamycin.
<3>Genome Announcements
<4>1
<5>e00581-13
<6>2013
<7>Streptomyces rapamycinicus strain NRRL 5491 produces the important drug rapamycin. It has a
large genome of 12.7 Mb, of which over 3 Mb consists of 48
secondary metabolite biosynthesis clusters.

<>

<1>Baranasic, D., Zucko, J., Nair, M., Pain, A., Long, P.F., Hranueli, D., Cullum, J., Starcevic, A.
<2>Genome Sequences of the Oxytetracycline Production Strain Streptomyces rimosus R6-500 and Two Mutants with Chromosomal Rearrangements.
<3>Genome Announcements
<4>2
<5>e00517-14
<6>2014
<7>The genome sequence of Streptomyces rimosus R6-500, an industrially improved strain which
produces high titers of the important antibiotic oxytetracycline, is
reported, as well as the genome sequences of two derivatives arising due to the
genetic instability of the strain.

<>

<1>Barany, F.
<2>A genetic system for isolation and characterization of TaqI restriction endonuclease mutants.
<3>Gene
<4>56
<5>13-27
<6>1987
<7>The gene encoding TaqI restriction endonuclease has been subcloned downstream
from an inducible phoA promoter.  Certain strains of Escherichia coli remain
viable when endonuclease is expressed, even in the absence of (protective)
methylation.  Infecting lambda phage DNA is not restricted in vivo.  One E.
coli strain, MM294, exhibited a temperature-sensitive phenotype when TaqI
endonuclease was induced.  This allowed for design of an in vivo plate assay
for identification of specially constructed two-codon insertion mutants in the
endonuclease gene.  These mutants exhibited a wide range of in vitro
activities, including wild-type activity, greater activity in low-salt buffer,
and sequence-specific nicking activity.

<>

<1>Barany, F.
<2>The TaqI star reaction:  strand preferences reveal hydrogen-bond donor and acceptor sites in canonical sequence recognition.
<3>Gene
<4>65
<5>149-165
<6>1988
<7>TaqI endonuclease recognizes and cleaves its canonical sequence, TCGA, with complete fidelity
under standard conditions. In the presence of some organic solvents, TaqI endonuclease
introduced additional single-strand and double-strand cuts at sequences termed TaqI star
sites. Using middle-labeled DNA, the relative rates of cleavage of each strand were
simultaneously determined for several star sites. These star recognition sequences differed
from the canonical sequence by a single base, and all potential star sites were either nicked
or cleaved. Star sites within the middle labeled substrate represented ten of the twelve
possible star sequences for each strand. For each group of identical star sites, one strand
was consistently preferred for cleavage. Based on these preferences, a model for TaqI
recognition of the TCGA sequence is proposed. According to this model, sequence discrimination
is mediated by eight hydrogen bonds formed between TaqI and the cognate nucleotides within the
major groove.

<>

<1>Barany, F.
<2>Isolation and characterization of TaqI restriction endonuclease mutants.
<3>J. Cell Biochem. Suppl.
<4>13A
<5>82
<6>1989
<7>The gene encoding the TaqI restriction endonuclease (recognition sequence T^CGA) has been
subcloned downstream of an inducible phoA promoter, and overproduced to 30% of cellular
proteins. E. coli cells remain viable at 37C when endonuclease is expressed, even in the
absence of (protective) methylation. Analysis of wild type TaqI endonuclease star activity
(cleavage at closely related sequences) revealed that for a particular star site, one strand
of the DNA duplex was preferentially cleaved. Based on these preferences, a model is proposed
that sequence specific discrimination is mediated by 8 hydrogen bonds formed by TaqI and
specific base groups in the major groove. Using a strain that produces beta-galactosidase upon
DNA damage, and a temperature dependent in vivo plate assay as screens, over two dozen
two-codon insertion mutants have been isolated. These are being characterized with respect to
in vitro canonical site activity, DNA binding using gel mobility shifts, and star site
activity.

<>

<1>Barany, F.
<2>How TaqI endonuclease recognizes its cognate sequence.
<3>Gene
<4>74
<5>63-65
<6>1988
<7>Meeting Abstract

<>

<1>Barany, F.
<2>Two codon insertion mutagenesis of the TaqI restriction endonuclease.
<3>J. Cell Biochem. Suppl.
<4>11C
<5>240
<6>1987
<7>Recently, the thermophilic TaqI restriction endonuclease and corresponding
methylase (recognition sequence TCGA) have been cloned.  The endonuclease gene
has been subcloned under inducible control of an alkaline phosphatase promoter,
in a small high copy plasmid.  Surprisingly, E. coli cells overproducing TaqI
endonuclese are still viable at 37C, even in the absence of (protective)
methylation.  Indeed, neither transforming DNA nor infecting lambda phage are
restricted by cells harboring TaqI.  However, certain cells harboring this
plasmid are barely viable at 42C, and this phenotype has been exploited as a
plate assay for in vivo TaqI endonuclease activity.  Single stranded hexameric
linkers were inserted into the endonuclease gene using biological selection,
and the resultant two codon insertion mutants were assayed in vitro for TaqI
endonuclease activity.  A wide range of enzymatic activity was observed, from
wild type activity (3 mutants), slight temperature dependency (2 mutants),
moderate activity (2 mutants), greater activity in low salt buffers (2
mutants), nicking activity ( 1 mutant), to no activity (2 mutants, and 2 small
in-1. Slatko, B., Movan, L., Jager, T., Benner, J., Simcox, T., & Wilson, G.
Ms in preparation 1986.  2. Barany, F. (1985) Proc. Natl. Acad. Sci. USA
82:4202-4206.

<>

<1>Barany, F.
<2>Overproduction, purification and crystallization of TaqI restriction endonuclease.
<3>Gene
<4>65
<5>167-177
<6>1988
<7>Under phoA promoter control, TaqI endonuclease was overproduced to 5% of
Escherichia coli cellular proteins.  This was achieved by fusing the
endonuclease gene to the first four codons of the alkaline phosphatase signal
sequence.  For maximal overproduction (30% of cellular proteins), a putative
14-bp hairpin within the endonuclease coding sequence was replaced with
degenerate codons  In addition, TaqI methylase was required to protect host
DNA.  The endonuclease was purified in sufficient amounts for crystallization.

<>

<1>Barany, F., Danzitz, M., Zebala, J., Mayer, A.
<2>Cloning and sequencing of genes encoding the TthHB8I restriction and modification enzymes: comparison with the isoschizomeric TaqI enzymes.
<3>Gene
<4>112
<5>3-12
<6>1992
<7>Genes encoding the TthHB8I restriction and modification (R-M) system from
Thermus thermophilus HB8 (recognition sequence T^CGA) were cloned in
Escherichia coli.  The genes have the same transcriptional orientation, with
the last 13 codons of the methyltransferase (MTase) overlapping the first 13
codons of the endonuclease (ENase).  Nucleotide sequence analysis of the TthB8I
ENase revealed a single chain of 263 amino acid (aa) residues that share a 77%
identity with the corrected isoschizomeric TaqI ENase.  Likewise, the Tth MTase
(428 aa) shares a 79% identity with the corrected sequence of the TaqI MTase.
This high degree of aa conservation suggests a common origin between the Taq
and Tth R-M systems.  However, codon usage and G + C content for the R-M genes
differed markedly from that of other cloned Thermus genes.  This suggests that
these R-M genes were only recently introduced into the genus Thermus.

<>

<1>Barany, F., Slatko, B., Danzitz, M., Cowburn, D., Schildkraut, I., Wilson, G.G.
<2>The corrected nucleotide sequences of the TaqI restriction and modification enzymes reveal a thirteen-codon overlap.
<3>Gene
<4>112
<5>91-95
<6>1992
<7>The nucleotide sequence of the genes encoding methyltransferase TaqI (M.TaqI)
and restriction endonuclease TaqI (R.TaqI) with the recognition sequence, TCGA,
were analyzed in clones isolated from independent libraries.  The genes,
originally reported as 363 and 236 codons long [Slatko et al., Nucleic Acids
Res. 15 (1987) 9781-9796] were determined as 421 and 263 codons long,
respectively.  The C terminus of the taqIM gene overlaps the N terminus of the
taqIR gene by 13 codons, as observed with the isoschizomeric TthHB8I
restriction-modification system [Barany et al., Gene 112 (1992) 13-20].
Removal of the overlapping codons did not interfere with in vivo M.TaqI
activity.  We postulate the overlap plays a role in regulating taqIR
expression.

<>

<1>Barany, F., Zebala, J.
<2>Correlation between insertion mutant activities and amino acid sequence identities of the TaqI and TthHB8 restriction endonucleases.
<3>Gene
<4>112
<5>13-20
<6>1992
<7>A two-codon insertion mutagenesis method has been generalized.  Over two dozen
insertion mutants throughout the gene encoding TaqI restriction endonuclease
were constructed and activity was characterized.  All mutants with activity
either cleaved or nicked the canonical T^CGA recognition sequence.  Some
insertion mutants created duplication of gene regions, termed Gemini proteins,
which still retained activity.  The correlation between mutants with poor
activity and the regions of shared amino identity between the isoschizomeric
Taq I and TthHB8I suggests these regions are involved in DNA recognition and/or
catalysis.

<>

<1>Baranyi, U., Klein, R., Lubitz, W., Kruger, D.H., Witte, A.
<2>The archaeal halophilic virus-encoded Dam-like methyltransferase M.Phi Ch1-I methylates adenine residues and complements dam mutants in the low salt environment of Escherichia coli.
<3>Mol. Microbiol.
<4>35
<5>1168-1179
<6>2000
<7>The genome of the archaeal virus phi Ch1, infecting Natrialba magadii (formerly
Natronobacterium magadii), is composed of 58.5 kbp linear ds
DNA. Virus particles contain several RNA species in sizes of 100-800
nucleotides. A fraction of phi Ch1 genomes is modified within
5'-GATC-3' and related sequences, as determined by various restriction
enzyme digestion analyses. High performance liquid chromatography
revealed a fifth base, in addition to the four nucleosides, which was
identified as N-6-methyladenosine. Genetic analyses and subsequent
sequencing led to the identification of a DNA (N-6-adenine)
methyltransferase (mtase) gene. The protein product was designated
M.phi Ch1-I. By the localization of the most conserved motifs (a DPPY
motif occurring before FxGxG), the enzyme was placed within the
beta-subgroup of the (N-6-adenine) methyltransferase class. The mtase
gene of phi Ch1 was classified as a 'late' gene, as determined by
measuring the kinetics of mRNA and protein expression in N. magadii
during the lytic cycle of phi Ch1. After infection of cells, M.phi
Ch1-I mRNA and protein could be detected in lower amounts than in the
situation of virus induction from lysogenic cells. Consequently, only
about 5% of the phi Ch1 progeny genomes after infection of N. magadii
carry the M.phi Ch1-I methylation in contrast to 50% of virus genomes
generated by induction of phi Ch1-lysogenic N. magadii cells.
Heterologous expression of the mtase from a halophile with 3 M
cytoplasmic salt concentration showed an unexpected feature: the
protein was active in the low environment of Escherichia coli and was
able to methylate DNA in vivo. Interestingly, it seemed to exhibit a
higher sequence specificity in E. coli that resulted in adenine
methylation exclusively in the sequence 5'-GATC-3'. Additionally,
expression of M.phi Ch1-I in dam(-) E. coli cells led to a complete
substitution of the function of M.Dam in DNA mismatch repair.

<>

<1>Barauna, R.A., Guimaraes, L.C., Veras, A.A., de Sa, P.H., Gracas, D.A., Pinheiro, K.C., Silva, A.S., Folador, E.L., Benevides, L.J., Viana, M.V., Carneiro, A.R., Schneider, M.P., Spier, S.J., Edman, J.M., Ramos, R.T., Azevedo, V., Silva, A.
<2>Genome Sequence of Corynebacterium pseudotuberculosis MB20 bv. equi Isolated from a Pectoral Abscess of an Oldenburg Horse in California.
<3>Genome Announcements
<4>2
<5>e00977-14
<6>2014
<7>The genome of Corynebacterium pseudotuberculosis MB20 bv. equi was sequenced using the Ion
Personal Genome Machine (PGM) platform, and showed a size of
2,363,089 bp, with 2,365 coding sequences and a GC content of 52.1%. These
results will serve as a basis for further studies on the pathogenicity of C.
pseudotuberculosis bv. equi.

<>

<1>Barbau-Piednoir, E., Bertrand, S., Roosens, N.H., De Keersmaecker, S.C.
<2>Genome Sequence of the Salmonella enterica subsp. enterica Serovar Namur Strain 05-2929, Lacking the Salmonella Atypical Fimbrial Operon.
<3>Genome Announcements
<4>2
<5>e00299-14
<6>2014
<7>This paper announces the genome sequence and annotation of Salmonella enterica subsp. enterica
serovar Namur strain 05-2929. S. Namur is a new serovar (39:z4,z23:-) that was isolated from a
patient with salmonellosis in 2005 in Namur, Belgium, and has been identified as lacking the
Salmonella atypical fimbrial (saf) operon.

<>

<1>Barbe, V., Bouzon, M., Mangenot, S., Badet, B., Poulain, J., Segurens, B., Vallenet, D., Marliere, P., Weissenbach, J.
<2>Complete Genome Sequence of Streptomyces cattleya NRRL 8057, a Producer of Antibiotics and Fluorometabolites.
<3>J. Bacteriol.
<4>193
<5>5055-5056
<6>2011
<7>Streptomyces cattleya, a producer of the antibiotics thienamycin and cephamycin C, is one of
the rare bacteria known to synthesize fluorinated
metabolites. The genome consists of two linear replicons. The genes
involved in fluorine metabolism and in the biosynthesis of the antibiotic
thienamycin were mapped on both replicons.

<>

<1>Barbe, V., Vallenet, D., Fonknechten, N., Kreimeyer, A., Oztas, S., Labarre, L., Cruveiller, S., Robert, C., Duprat, S., Wincker, P., Ornston, L.N., Weissenbach, J., Marliere, P., Cohen, G.N., Medigue, C.
<2>Unique features revealed by the genome sequence of Acinetobacter sp. ADP1, a versatile and naturally transformation competent bacterium.
<3>Nucleic Acids Res.
<4>32
<5>5766-5779
<6>2004
<7>Acinetobacter sp. strain ADP1 is a nutritionally versatile soil bacterium closely related to
representatives of the well-characterized Pseudomonas
aeruginosa and Pseudomonas putida. Unlike these bacteria, the
Acinetobacter ADP1 is highly competent for natural transformation which
affords extraordinary convenience for genetic manipulation. The circular
chromosome of the Acinetobacter ADP1, presented here, encodes 3325
predicted coding sequences, of which 60% have been classified based on
sequence similarity to other documented proteins. The close evolutionary
proximity of Acinetobacter and Pseudomonas species, as judged by the
sequences of their 16S RNA genes and by the highest level of bidirectional
best hits, contrasts with the extensive divergence in the GC content of
their DNA (40 versus 62%). The chromosomes also differ significantly in
size, with the Acinetobacter ADP1 chromosome <60% of the length of the
Pseudomonas counterparts. Genome analysis of the Acinetobacter ADP1
revealed genes for metabolic pathways involved in utilization of a large
variety of compounds. Almost all of these genes, with orthologs that are
scattered in other species, are located in five major 'islands of
catabolic diversity', now an apparent 'archipelago of catabolic
diversity', within one-quarter of the overall genome. Acinetobacter ADP1
displays many features of other aerobic soil bacteria with metabolism
oriented toward the degradation of organic compounds found in their
natural habitat. A distinguishing feature of this genome is the absence of
a gene corresponding to pyruvate kinase, the enzyme that generally
catalyzes the terminal step in conversion of carbohydrates to pyruvate for
respiration by the citric acid cycle. This finding supports the view that
the cycle itself is centrally geared to the catabolic capabilities of this
exceptionally versatile organism.

<>

<1>Barber, R.D., Zhang, L., Harnack, M., Olson, M.V., Kaul, R., Ingram-Smith, C., Smith, K.S.
<2>Complete genome sequence of Methanosaeta concilii, a specialist in aceticlastic methanogenesis.
<3>J. Bacteriol.
<4>193
<5>3668-3669
<6>2011
<7>Complete genome sequence of Methanosaeta concilii, a specialist in Aceticlastic
Methanogenesis.
episome, has been determined and exhibits considerable differences in gene
content from that of Methanosaeta thermophila.

<>

<1>Barbes, C., Hardisson, C., Novella, I.S., Yebra, M.J., Sanchez, J.
<2>DNA-methyltransferase activities in Streptomyces antibioticus.
<3>FEMS Microbiol. Lett.
<4>55
<5>59-64
<6>1988
<7>Several DNA methyltransferases have been detected in Streptomyces antibioticus ETHZ 7451.  One
of these enzymes was purified and partially characterized from crude extracts of Streptomyces
antibioticus ETHZ 7451.  This enzyme only methylates the adenine residues of DNA.  However,
lambda and pBR322 DNAs were not protected in vitro with the methylase, indicating that the
enzyme does not seem to form part of a restriction-modification system.  Remarkably, the
efficiency of the enzyme is significantly higher on the DNA isolated from aerial mycelium than
from substrate mycelium in S. antibioticus.

<>

<1>Barbes, C., Sanchez, J., Yebra, M.J., Robert-Gero, M., Hardisson, C.
<2>Effects of sinefungin and S-adenosylhomocysteine on DNA and protein methyltransferases from Streptomyces and other bacteria.
<3>FEMS Microbiol. Lett.
<4>69
<5>239-244
<6>1990
<7>Sinefungin is a naturally occurring nucleoside isolated from cultures of
Streptomyces griseolus and S. incarnatus.  It is structurally related to
S-adenosyl-methionine (SAM) and S-adenosyl-L-homocysteine (SAH).  Its effect
and level of action on prokaryotes has not been studied with the same detail as
with eukaryotic cells.  In this report we describe the effect of sinefungin and
SAH on several Streptomyces methyltransferases (DNA and protein MTases) and on
other bacterial DNA-MTases.  Protein MTases are resistant to sinefungin,
whereas DNA-MTases are inhibited.  Adenine MTases however, seem more sensitive
to this analogue than cytosine MTases.

<>

<1>Barbeyron, T., Kean, K., Forterre, P.
<2>DNA adenine methylation of GATC sequences appeared recently in the Escherichia coli lineage.
<3>J. Bacteriol.
<4>160
<5>586-590
<6>1984
<7>We have examined the presence of methylated adenine at GATC sequences (Dam phenotype) in the
DNA of 23 eubacteria and 13 archaebacteria by using isoschizomer restriction enzymes. We have
found a completely Dam+ phenotype in bacteria of nine genera related to the families
Enterobacteriaceae, Parvobacteriaceae, and Vibrionaceae, and in the five cyanobacteria tested.
We have found a partial Dam+ phenotype in the two archaebacteria Halobacterium saccharovorum
and Methanobacterium sp. strain Ivanov. All of the other archaebacteria (three genera) and
eubacteria (nine genera) tested were Dam-. Phylogenetic analysis, based on the evolutionary
tree of Fox et al., indicates that dam methylation in the Escherichia coli lineage appeared
recently in bacterial evolution and is restricted to a small range of closely related
bacteria.

<>

<1>Barbier, F., Ruppe, E., Hernandez, D., Lebeaux, D., Francois, P., Felix, B., Desprez, A., Maiga, A., Woerther, P.L., Gaillard, K., Jeanrot, C., Wolff, M., Schrenzel, J., Andremont, A., Ruimy, R.
<2>Methicillin-resistant coagulase-negative staphylococci in the community: high homology of SCCmec IVa between Staphylococcus epidermidis and major clones of methicillin-resistant Staphylococcus aureus.
<3>J. Infect. Dis.
<4>202
<5>270-281
<6>2010
<7>BACKGROUND: Data on community spread of methicillin-resistant
coagulase-negative staphylococci (MR-CoNS) are scarce. We assessed their
potential role as a reservoir of staphylococcal cassette chromosome mec
(SCCmec) IVa, the leading SCCmec subtype in community-acquired
methicillin-resistant Staphylococcus aureus (CA-MRSA). METHODS: Nasal
carriage of MR-CoNS was prospectively investigated in 291 adults at
hospital admission. MR-CoNS were characterized by SCCmec typing,
long-range polymerase chain reaction (PCR) for SCCmec IV, and
multiple-locus variable-number tandem repeat analysis (MLVA) for
Staphylococcus epidermidis (MRSE) strains. Three SCCmec IVa elements were
fully sequenced. RESULTS: The carriage rate of MR-CoNS was 19.2% (25.9%
and 16.5% in patients with and patients without previous exposure to the
health care system, respectively; P = .09). MR-CoNS strains (n = 83,
including 58 MRSE strains with highly heterogeneous MLVA patterns) carried
SCCmec type IVa (n = 9, all MRSE), other SCCmec IV subtypes (n = 9,
including 7 MRSE), other SCCmec types (n = 15), and nontypeable SCCmec (n
= 50). Long-range PCR indicated structural homology between SCCmec IV in
MRSE and that in MRSA. Complete sequences of SCCmec IVa from 3 MRSE
strains were highly homologous to those available for CA-MRSA, including
major clones USA300 and USA400. CONCLUSIONS: MR-CoNS are probably
disseminated in the community, notably in subjects without previous
exposure to the health care system. MRSE, the most prevalent species, may
act as a reservoir of SCCmec IVa for CA-MRSA.

<>

<1>Barbier, P., Houel, A., Loux, V., Poulain, J., Bernardet, J.F., Touchon, M., Duchaud, E.
<2>Complete Genome Sequence of Flavobacterium indicum GPSTA100-9T, Isolated from Warm Spring Water.
<3>J. Bacteriol.
<4>194
<5>3024-3025
<6>2012
<7>We report here the complete annotated genome sequence of Flavobacterium indicum CIP 109464(T)
(= GPTSA100-9(T)), isolated from warm spring water in Assam, India.  The genome sequence of F.
indicum revealed a number of interesting features and genes in relation to its environmental
lifestyle.

<>

<1>Barbour, A.G., Campeau, M.S.
<2>Genome Sequence of Borrelia parkeri, an Agent of Enzootic Relapsing Fever in Western North America.
<3>Genome Announcements
<4>2
<5>e00018-14
<6>2014
<7>Borrelia parkeri is a relapsing fever agent that rarely causes human infection, unlike other
North American species. B. parkeri strain HR1 was isolated from
Ornithodoros parkeri ticks. The sequences of its linear chromosome and large
plasmid were determined by next-generation sequencing. These confirmed its closer
relatedness to Borrelia turicatae than to Borrelia hermsii.

<>

<1>Barcak, G.J., Kruger, D.H., Pein, C.D., Reuter, M.
<2>Restriction endonuclease cleavage of resistant DNA - in the presence of a second, sensitive DNA species.
<3>East German Patent Office
<4>DD 264231 A
<5>
<6>1989
<7>In a new procedure for cleaving a resistant DNA with restriction endonucleases, the resistant
DNA (DNA-I) is incubated with a second sensitive DNA species (DNA-II) in the presence of a
restriction endonuclease under conventional reaction conditions. DNA-II is preferably a
biologically replicated DNA (pref. plasmid pBR322 or phage lambda) or a synthetic
oligonucleotide duplex. The ratio of DNA-II to DNA-I should be such that the total number of
recognition sites in DNA-II is at least as high as the number of DNA-I.

<>

<1>Barciszewski, J., Markiewicz, W.T., Adamska, E., Plitta, B., Giel-Pietraszuk, M., Wyszko, E., Markiewicz, M., Fedoruk-Wyszomirska, A., Kulinski, T., Chmielewski, M.
<2>Preparation of cytosine analogs and nucleosides as DNA methyltransferase 1 inhibitors and antitumor agents.
<3>International Patent Office
<4>WO 2011115513 A
<5>
<6>2011
<7>
<>

<1>Barcus, V.A., Murray, N.E.
<2>Barriers to recombination: restriction.
<3>Population Genetics of Bacteria, Cambridge University Press, Baumberg, S., Young, J.P.W., Saunders, S.R., Wellington, E.M.H., Cambridge
<4>0
<5>31-58
<6>1995
<7>The biological barrier provided by restriction is far from complete, and it can be quantified
by determining the titre of unmodified phages on the restricting strain relative to the titre
on a non-restricting strain.  For phage genomes with small numbers of targets, the barrier
increases with target number, as witnessed by a decrease in the efficiency of plating (eop).
Foreign DNA can be recognized and attacked by restriction endonucleases irrespective of its
mode of entry into the cell and nature has many tricks for combating restriction.
Nevertheless, a recipient strain with a restriction system is endowed with a potential barrier
to the intrusion of large DNA molecules should these molecules include the relevant, but
unmodified, target sequences.  Whether or not the cutting of larger DNA molecules into smaller
ones is necessarily a barrier to genetic recombination is of critical concern, since DNA
breaks can be substrates for recombination.  The restriction of foreign DNA is dependent on
the donor bacterium lacking a modification system that protects its DNA against attack by a
restriction system present within a recipient cell.  Any evaluation of the importance of
restriction to the transfer of genetic information requires an awareness of the diversity of
restriction specificities within a bacterial species.  This review considers what is known
about the diversity of restriction systems within the Enterobacteriaceae, but most
particularly within the best studied species, Escherichia coli.  In the absence of any truly
systematic analyses, the focus will be primarily on screens for chromosomally encoded
restriction and modification (R-M) systems.

<>

<1>Barcus, V.A., Titheradge, A.J.B., Murray, N.E.
<2>The diversity of alleles at the hsd locus in natural populations of Escherichia coli.
<3>Genetics
<4>140
<5>1187-1197
<6>1995
<7>In enteric bacteria three discrete families of type I restriction and modification systems
(IA, IB and ID) are encoded by alleles of the serB-linked hsd locus.  Probes specific for each
of the three families were used to monitor the distribution of related systems in 37 of the
72 wild-type Escherichia coli strains comprising the ECOR collection.  All 25 members of group
A in this collection were screened; 12 were probe-positive, nine have hsd genes in the IA
family, two in the IB and one in the ID.  Twelve strains, representing all groups other than
A, were screened; five were probe-positive, one has hsd genes in the IA family, one in the IB
and three in the ID.  The type ID genes are the first representatives of this family in E.
coli, the probe-negative strains could have alternative families of hsd genes.  The type IA
and IB systems added at least five new specificities to the five already identified in natural
isolates of E. coli.  The distribution of alleles is inconsistant with the dendrogram of the
bacterial strains derived from other criteria.  This discrepancy and the dissimilar coding
sequences of allelic hsd genes both imply lateral transfer of hsd genes.

<>

<1>Barke, J., Seipke, R.F., Gruschow, S., Heavens, D., Drou, N., Bibb, M.J., Goss, R.J., Yu, D.W., Hutchings, M.I.
<2>A mixed community of actinomycetes produce multiple antibiotics for the fungus farming ant Acromyrmex octospinosus.
<3>BMC Biol.
<4>8
<5>109
<6>2010
<7>BACKGROUND: Attine ants live in an intensely studied tripartite mutualism
with the fungus Leucoagaricus gongylophorus, which provides food to the
ants, and with antibiotic-producing actinomycete bacteria. One hypothesis
suggests that bacteria from the genus Pseudonocardia are the sole,
co-evolved mutualists of attine ants and are transmitted vertically by the
queens. A recent study identified a Pseudonocardia-produced antifungal,
named dentigerumycin, associated with the lower attine Apterostigma
dentigerum consistent with the idea that co-evolved Pseudonocardia make
novel antibiotics. An alternative possibility is that attine ants sample
actinomycete bacteria from the soil, selecting and maintaining those
species that make useful antibiotics. Consistent with this idea, a
Streptomyces species associated with the higher attine Acromyrmex
octospinosus was recently shown to produce the well-known antifungal
candicidin. Candicidin production is widespread in environmental isolates
of Streptomyces, so this could either be an environmental contaminant or
evidence of recruitment of useful actinomycetes from the environment. It
should be noted that the two possibilities for actinomycete acquisition
are not necessarily mutually exclusive. RESULTS: In order to test these
possibilities we isolated bacteria from a geographically distinct
population of A. octospinosus and identified a candicidin-producing
Streptomyces species, which suggests that they are common mutualists of
attine ants, most probably recruited from the environment. We also
identified a Pseudonocardia species in the same ant colony that produces
an unusual polyene antifungal, providing evidence for co-evolution of
Pseudonocardia with A. octospinosus. CONCLUSIONS: Our results show that a
combination of co-evolution and environmental sampling results in the
diversity of actinomycete symbionts and antibiotics associated with attine
ants.

<>

<1>Barker, D., Hoff, M., Oliphant, A., White, R.
<2>A second type II restriction endonuclease from Thermus aquaticus with an unusual sequence specificity.
<3>Nucleic Acids Res.
<4>12
<5>5567-5581
<6>1984
<7>A type II restriction endonuclease activity free of TaqI was prepared from Thermus aquaticus
YT.  The fraction contains two endonucleolytic components with apparently different
specificities, however the major activity is sufficiently dominant to allow partial digestion
analysis of the position of recognition sites.  A precise determination of the location of
cleavage sites in pBR322 DNA and a computer-aided search for regions of homology in the
vicinity of the cut sites indicate that this enzyme recognizes the nonpalindromic sequences
GACCGA or CACCCA.  Other related sequences are not cleaved, in particular, GACCCA and CACCGA,
indicating that the enzyme requires the identity of nucleotides in the first and fifth
positions, a type of specificity that has not been previously reported.  The position of
cleavage is located outside of the site and is represented as:
5'-GACCGANNNNNNNNN^NN-3'
3'-CTGGCTNNNNNNNNN^NN-5'.

<>

<1>Barker, E.N., Helps, C.R., Peters, I.R., Darby, A.C., Radford, A.D., Tasker, S.
<2>Complete Genome Sequence of Mycoplasma haemofelis, a Hemotropic Mycoplasma.
<3>J. Bacteriol.
<4>193
<5>2060-2061
<6>2011
<7>Here, we present the genome sequence of Mycoplasma haemofelis strain Langford 1, representing
the first hemotropic mycoplasma (hemoplasma)
species to be completely sequenced and annotated. Originally isolated from
a cat with hemolytic anemia, this strain induces severe hemolytic anemia
when inoculated into specific-pathogen-free-derived cats. The genome
sequence has provided insights into the biology of this uncultivatable
hemoplasma and has identified potential molecular mechanisms underlying
its pathogenicity.

<>

<1>Barlow, D.P.
<2>Methylation and imprinting: from host defense to gene regulation?
<3>Science
<4>260
<5>309-310
<6>1993
<7>
<>

<1>Barlow, R.S., Gobius, K.S.
<2>Diverse class 2 integrons in bacteria from beef cattle sources.
<3>J. Antimicrob. Chemother.
<4>58
<5>1133-1138
<6>2006
<7>OBJECTIVES: The purpose of this study was to determine the diversity of class 2 integrons in
bacteria isolated from beef cattle sources. METHODS:
The variable regions of a subset of 11 class 2 integron-containing
bacteria were analysed by PCR and DNA sequencing for the presence of novel
rearrangements. RESULTS: A total of six different class 2 integron arrays
were identified and four of these were fully characterized. Three of the
four arrays characterized have been previously described; however the
remaining array is unlike previously described class 2 integrons. The
novel class 2 integron was found in Providencia stuartii and contains an
apparently functional class 2 integrase. Examination of the variable
region of the P. stuartii integron identified nine open reading frames,
mostly of unknown function, and represents the first report of a class 2
integron without inserted antibiotic resistance gene cassettes.
CONCLUSIONS: This study has identified a novel class 2 integron found in
P. stuartii that contains an apparently functional naturally occurring
class 2 integrase. Further investigation of this novel class 2 integron is
required to determine the impact of a functional class 2 integrase upon
the evolution of class 2 integrons.

<>

<1>Barnes, G., Rine, J.
<2>Regulated expression of endonuclease EcoRI in Saccharomyces cerevisiae: Nuclear entry and biological consequences.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>82
<5>1354-1358
<6>1985
<7>In an investigation to determine how proteins are localized within the nucleus
of a cell, we demonstrate that the restriction endonuclease EcoRI is able to
enter and function within the nucleus of Saccharomyces cerevisiae when this
prokaryotic protein is synthesized in vivo.  The EcoRI endonuclease was
produced in yeast under the transcriptional control of a regulated yeast
promoter by ligating a DNA fragment containing only coding sequences for the
endonuclease to the promoter element of the yeast GAL1 gene (the structural
gene for galactokinase, EC 2.7.1.6).  Yeast cells harboring a plasmid
containing this promoter-gene fusion are able to grow under conditions that
repress transcription from the Gal1 promoter.  However, under inducing
conditions, these yeast cells are unable to grow.  Moreover, rad52 mutants,
which are deficient in the repair of double-strand breaks, are more sensitive
to the presence of the promoter-gene fusion plasmid than are wild-type cells.
We demonstrate that the EcoRI endonuclease activity is present in lysates
prepared from yeast transformants grown under conditions that induce
transcription of GAL1, but this activity is not detectable in cells grown under
conditions that repress transcription from the promoter.  Furthermore, analysis
of yeast chromosomal DNA shows that the endonuclease enters the yeast nucleus
and cleaves DNA specifically at EcoRI recognition sites.

<>

<1>Barnes, H.E., Liu, G., Weston, C.Q., King, P., Pham, L.K., Waltz, S., Helzer, K.T., Day, L., Sphar, D., Yamamoto, R.T., Forsyth, R.A.
<2>Selective Microbial Genomic DNA Isolation Using Restriction Endonucleases.
<3>PLoS ONE
<4>9
<5>e109061
<6>2014
<7>To improve the metagenomic analysis of complex microbiomes, we have repurposed restriction
endonucleases as methyl specific DNA binding proteins. As an example, we use DpnI immobilized
on magnetic beads. The ten minute extraction technique allows specific binding of genomes
containing the DpnI G(m6) ATC motif common in the genomic DNA of many bacteria including
gamma-proteobacteria. Using synthetic genome mixtures, we demonstrate 80% recovery of
Escherichia coli genomic DNA even when only femtogram quantities are spiked into 10 mu g of
human DNA background. Binding is very specific with less than 0.5% of human DNA bound. Next
Generation Sequencing of input and enriched synthetic mixtures results in over 100-fold
enrichment of target genomes relative to human and plant DNA. We also show comparable
enrichment when sequencing complex microbiomes such as those from creek water and human
saliva. The technique can be broadened to other restriction enzymes allowing for the selective
enrichment of trace and unculturable organisms from complex microbiomes and the stratification
of organisms according to restriction enzyme enrichment.

<>

<1>Barra, R., Chiong, M., Gonzalez, E., Vasquez, C.
<2>A DNA-modification methylase from Bacillus stearothermophilus V.
<3>Biochem. J.
<4>255
<5>699-703
<6>1988
<7>A type II modification methylase (M.BstVI) was partially purified from the
thermophilic bacterium Bacillus stearothermophilus V.  The methylase catalyses
the transfer of methyl groups from S-adenosyl-L-methionine to unmodified
double-stranded DNA.  The product of methylation was identified by paper
chromatography as N6-methyladenine.  Since M.BstVI protects DNA against
cleavage by BstVI and XhoI restriction endonucleases, it follows that it
methylates the adenine residue in the sequence 5'-C-T-C-G-A-G-3'.

<>

<1>Barrado, L., Viedma, E., Vindel, A., Otero, J.R., Chaves, F.
<2>Draft Genome Sequence of Strain SA_ST125_MupR of Methicillin-Resistant Staphylococcus aureus ST125, a Major Clone in Spain.
<3>Genome Announcements
<4>1
<5>e00588-13
<6>2013
<7>Here, we report the draft genome sequence of a methicillin-resistant Staphylococcus aureus
(MRSA) strain with high-level mupirocin resistance
(SA_ST125_MupR), isolated from a patient with recurrent bacteremia. This strain
belonged to sequence type ST125, which was responsible for more than 50% of the
health care-associated infections caused by MRSA in Spain.

<>

<1>Barrangou, R., Briczinski, E.P., Traeger, L.L., Loquasto, J.R., Richards, M., Horvath, P., Coute-Monvoisin, A.C., Leyer, G., Rendulic, S., Steele, J.L., Broadbent, J.R., Oberg, T., Dudley, E.G., Schuster, S., Romero, D.A., Roberts, R.F.
<2>Comparison of the complete genome sequences of Bifidobacterium animalis subsp. lactis DSM 10140 and Bl-04.
<3>J. Bacteriol.
<4>191
<5>4144-4151
<6>2009
<7>Bifidobacteria are important members of the human gut flora, especially in infants.
Comparative genomic analysis of two Bifidobacterium animalis
subsp. lactis strains revealed evolution by internal deletion of
consecutive spacer-repeat units within a novel clustered regularly
interspaced short palindromic repeat locus, which represented the largest
differential content between the two genomes. Additionally, 47 single
nucleotide polymorphisms were identified, consisting primarily of
nonsynonymous mutations, indicating positive selection and/or recent
divergence. A particular nonsynonymous mutation in a putative glucose
transporter was linked to a negative phenotypic effect on the ability of
the variant to catabolize glucose, consistent with a modification in the
predicted protein transmembrane topology. Comparative genome sequence
analysis of three Bifidobacterium species provided a core genome set of
1,117 orthologs complemented by a pan-genome of 2,445 genes. The genome
sequences of the intestinal bacterium B. animalis subsp. lactis provide
insights into rapid genome evolution and the genetic basis for adaptation
to the human gut environment, notably with regard to catabolism of dietary
carbohydrates, resistance to bile and acid, and interaction with the
intestinal epithelium. The high degree of genome conservation observed
between the two strains in terms of size, organization, and sequence is
indicative of a genomically monomorphic subspecies and explains the
inability to differentiate the strains by standard techniques such as
pulsed-field gel electrophoresis.

<>

<1>Barras, F., Marinus, M.G.
<2>The great GATC:  DNA methylation in E. coli.
<3>Trends Genet.
<4>5
<5>139-143
<6>1989
<7>In Escherichia coli the methylation of the adenine in the sequence 5'-GATC-3' is catalysed
by the dam gene product, a DNA adenine methylase. We review the proposed roles for this
methylation, and the sequence it modifies, in mismatch repair, DNA-protein interaction, gene
expression, the initiation of chromosome replication, chromosome segregation, chromosome
structure and the occurrence of mutational hotspots.

<>

<1>Barreau, G., Tompkins, T.A., de Carvalho, V.G.
<2>Draft Genome Sequence of Probiotic Strain Pediococcus acidilactici MA18/5M.
<3>J. Bacteriol.
<4>194
<5>901
<6>2012
<7>Pediococcus acidilactici MA18/5M is a commercially available probiotic that is widely used in
swine, poultry, aquaculture feeds, and human
dietary supplements. We prepared a genome sequence for this strain
consisting of 2 scaffolds totaling 1,992,928 bases including gaps for a
total of 3,346 bases and a G+C content of 42%.

<>

<1>Barreiro, C. et al.
<2>Draft Genome of Streptomyces tsukubaensis NRRL 18488, the Producer of the Clinically Important Immunosuppressant Tacrolimus (FK506).
<3>J. Bacteriol.
<4>194
<5>3756-3757
<6>2012
<7>The macrocyclic polyketide tacrolimus (FK506) is a potent immunosuppressant that  prevents
T-cell proliferation produced solely by Streptomyces species. We report
here the first draft genome sequence of a true FK506 producer, Streptomyces
tsukubaensis NRRL 18488, the first tacrolimus-producing strain that was isolated
and that contains the full tacrolimus biosynthesis gene cluster.

<>

<1>Barrera, C., Valot, B., Rognon, B., Zaugg, C., Monod, M., Millon, L.
<2>Draft Genome Sequence of the Principal Etiological Agent of Farmer's Lung Disease, Saccharopolyspora rectivirgula.
<3>Genome Announcements
<4>2
<5>e01340-14
<6>2014
<7>Saccharopolyspora rectivirgula is the main cause of farmer's lung disease. The development of
recombinant antigens to standardize the serodiagnosis of the
disease requires knowledge of the S. rectivirgula genome. We sequenced the genome
of an environmental strain, S. rectivirgula DSM 43113. A total of 3,221 proteins
were found to be encoded in a short 3.9-Mb genome.

<>

<1>Barrero, R.A., Moolhuijzen, P., Indjein, L., Venus, B., Keeble-Gagnere, G., Power, J., Bellgard, M.I., Lew-Tabor, A.E.
<2>Draft Genome Sequences of Campylobacter fetus subsp. venerealis bv. venerealis Strain B6 and bv. intermedius Strain 642-21.
<3>Genome Announcements
<4>2
<5>e00943-14
<6>2014
<7>Campylobacter fetus subsp. venerealis is an important venereal pathogen. We sequenced the
genomes of Campylobacter fetus subsp. venerealis bv. venerealis
strain B6 and bv. intermedius strain 642-21. The genetic variability of these
Australian strains will facilitate the study of mechanisms of geographical
adaptation of these pathogens that impact livestock.

<>

<1>Barreto-Hernandez, E., Falquet, L., Reguero, M.T., Mantilla, J.R., Valenzuela, E.M., Gonzalez, E., Cepeda, A., Escalante, A.
<2>Draft Genome Sequences of Multidrug-Resistant Acinetobacter sp. Strains from Colombian Hospitals.
<3>Genome Announcements
<4>1
<5>e00868-13
<6>2013
<7>The draft genome sequences of the strains Acinetobacter baumannii 107m, Acinetobacter
nosocomialis 28F, and Acinetobacter pittii 42F, isolated from
Colombian hospitals, are reported here. These isolates are causative of
nosocomial infections and are classified as multidrug resistant, as they showed
resistance to four different antibiotic groups.

<>

<1>Barretto, C., Alvarez-Martin, P., Foata, F., Renault, P., Berger, B.
<2>Genome Sequence of the Lantibiotic Bacteriocin Producer Streptococcus salivarius  Strain K12.
<3>J. Bacteriol.
<4>194
<5>5959-5960
<6>2012
<7>Streptococcus salivarius is a prevalent commensal species of the oropharyngeal tract. S.
salivarius strain K12 is an isolate from the saliva of a healthy child,
used as an oral probiotic. Here, we report its genome sequence, i.e., the full
sequence of the 190-kb megaplasmid pSsal-K12 and a high-quality draft 2.2-Gb
chromosomal sequence.

<>

<1>Barshevsky, T., Benner, J.S.
<2>Site-directed mutagenesis of the EcoRV methylase for the study of the functional role of the conserved sequences in N6-adenine methylases.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>211
<6>1989
<7>EcoRV methylase has been used as a model system to investigate the function of the conserved
sequences in N6-adenine methylases. The EcoRV methylase gene was subcloned and the methylase
protein was purified by multiple chromatographic steps. The highly conserved DPPY potein
sequence in the EcoRV methylase was changed using site-directed oligonucleotide mutagenesis in
both conservative and radical ways. We produced protein sequence changes at the D, the first P
and the Y so that the predicted secondary and unknown tertiary structures of this region were
not only gently perturbed but also disrupted. The EcoRV methylase with the first change (DPPY
to EPPY) was subcloned back into a modified pUC19 vector. This change produced a methylase
which unlike the wild-type enzyme does not fully methylate in vivo the genomic or vector DNA.
We are examining the catalytic properties of the wild-type EcoRV methylase and some
mutagenized methylase proteins. Also efforts will be made to determine whether SAM or DNA
binding, if any, remains in mutant methylases.

<>

<1>Barsomian, J., Wilson, G.
<2>Method for producing the HhaI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0321270 A
<5>
<6>1988
<7>The present invention is directed to a method for cloning and producing the HhaI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Haemophilus haemolyticus
ATCC 10014 into a host whereby the restriction gene is expressed; 2) fermenting the host which
contains the vector encoding and expressing the HhaI restriction endonuclease, and 3)
purifying the HhaI restriction enodnuclease from the fermented host which contains the vector
encoding and expressing the HhaI restriction endonuclease activity.

<>

<1>Barsomian, J., Wilson, G.
<2>Method for producing the HinPI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0321269 B
<5>
<6>1994
<7>The present invention relates to clones for the HinPI restriction endonuclease and
modification methylase, and to the production of these enzymes from the clones.

<>

<1>Barsomian, J.M., Card, C.O., Wilson, G.G.
<2>Cloning of the HhaI and HinPI restriction-modification systems.
<3>Gene
<4>74
<5>5-7
<6>1988
<7>The genes for the HhaI (Roberts et al., 1976) and HinPI (Roberts, 1987)
restriction-modification (R-M) systems have been cloned in pBR322.  The HhaI
system was isolated on a 9-kb PstI fragment, and the HinPI system was isolated
on two PstI fragments of 1.5 and 4.6 kb in length.  The clones were isolated by
selecting for recombinant molecules that had protectively modified themselves.
The HhaI and HinPI R-M systems recognize the same sequence, GCGC, but
hybridization between the DNA fragments encoding them does not take place.
Note:  this should be HinP1I.

<>

<1>Barsomian, J.M., Wilson, G.G.
<2>Method for producing the HinPI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 4983522
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the HinPI restriction
endonuclease by (1) introducing the restriction endonuclease gene from Haemophilus influenza
P1 into a host whereby the restriction gene is expressed; (2) fermenting the host which
contains the vector encoding and expressing the HinPI restriction endonuclease, and (3)
purifying the HinPI restriction endonuclease from the fermented host which contains the vector
encoding and expressing the HinPI restriction endonuclease activity.

<>

<1>Barsomian, J.M., Wilson, G.G.
<2>Method for producing the HhaI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 4999293
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the HhaI restriction
endonuclease by (1) introducing the restriction endonuclease gene from Haemophilus
haemolyticus ATCC 10014 into a host whereby the restriction gene is expressed (2) fermenting
the host which contains the vector encoding and expressing the HhaI restriction endonuclease,
and (3) purifying the HhaI restriction endonuclease from the fermented host which contains the
vector encoding and expressing the Hha I restriction endonuclease activity.

<>

<1>Barsomian, J.M., Wilson, G.G.
<2>Method for producing the HinPI restriction endonuclease and methylase.
<3>Japanese Patent Office
<4>JP 2761227
<5>
<6>1998
<7>
<>

<1>Bart, A.
<2>Genetic variation in Neisseria meningitidis.
<3>Ph.D. Thesis, Univ. Amsterdam, Netherlands, Ponsen & Looijen, BV, , Wageningen
<4>
<5>1-115
<6>1999
<7>Neisseria meningitidis or meningococcus is a Gram-negative diplococcus that is a member of the
family of the Neisseriaceae, belonging to the beta subdivision of the proteobacteria.  The
genus Neisseria includes closely related species.  These diplococci are primarily commensals
of the mucous membranes of mammals.  N. meningitidis and N. gonorrhoeae, are well-known
pathogens of man, although various Neisseria species are opportunistic pathogens (e.g. N.
flavescens and N. lactamica).  The meningococcus, usually carried in the nasopharynx and
oroparynx of humans, is distinguished from other Neisseria species on the basis of its sugar
metabolism.  The Neisseria are usually described as being immobile, though a recent report
suggests that they may display "twitching motility".

<>

<1>Bart, A., Dankert, J., van der Ende, A.
<2>Operator sequences for the regulatory proteins of restriction modification systems.
<3>Mol. Microbiol.
<4>31
<5>1275-1281
<6>1999
<7>For some type II restriction modification systems, it has been shown that transcription of the
methylase gene and the restriction endonuclease gene is regulated by the control gene product.
In these systems, C is located directly upstream of R, and in most systems M is located
divergently from CR.  The control element is a short (~80 amino acids) protein containing a
helix-turn-helix DNA-binding motif, distantly related to the well-known phage lambda cl
regulator.

<>

<1>Bart, A., Dankert, J., van der Ende, A.
<2>Representational difference analysis of Neisseria meningitidis identifies sequences that are specific for the hyper-virulent lineage III clone.
<3>FEMS Microbiol. Lett.
<4>188
<5>111-114
<6>2000
<7>Neisseria meningitidis may cause meningitis and septicemia.  Since the early 1980s, an
increased incidence of meningococcal disease has been caused by the lineage III clone in many
countries in Europe and in New Zealand.  We hypothesized that lineage III meningococci have
specific DNA sequences, providing an opportunity to facilitate epidemiological studies by
detecting lineage III isolates rapidly.  Applying representational difference analysis on one
lineage III tester strain and two non-lineage III driver strains, we identified three lineage
III-specific sequences, probably part of a single locus encoding a restriction modification
system.  A PCR based on one of these sequences identified lineage III meningococcal isolates
with a sensitivity of 100% and a specificity of 93%, which is superior to the serological
identification of lineage III isolates.

<>

<1>Bart, A., Pannekoek, Y., Dankert, J., van der Ende, A.
<2>NmeSI restriction-modification system identified by representational difference analysis of a hypervirulent Neisseria meningitidis strain.
<3>Infect. Immun.
<4>69
<5>1816-1820
<6>2001
<7>Neisseria meningitidis is a gram-negative bacterium that may cause meningitis, sepsis, or
both. The increase in the incidence of
meningococcal disease in various countries in the past 2 decades is
mainly due the genotypically related lineage III meningococci. The
chromosomal DNA differences between lineage III strains and non-lineage
III strains were identified using representational difference analysis.
Thus, a 1.8-kb locus that is specific for lineage III meningococci was
identified. The locus contains three open reading frames encoding the
NmeSI restriction-modification system. The methyltransferase gene was
cloned and expressed in Escherichia coli. Site AGTACT was found to be
modified by the enzyme. In conclusion, lineage III strains differ from
endemic strains by the presence of a specific restriction-modification
system. This restriction-modification system may contribute to the
clonal and hypervirulent character of lineage III strains by
influencing horizontal gene transfer and transcription.

<>

<1>Bart, A., Pannekoek, Y., Dankert, J., van der Ende, A.
<2>The NmeSI restriction-modification system identified by representational difference analysis of a hypervirulent Neisseria  meningitidis strain.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>303
<6>2001
<7>Neisseria meningitidis is a Gram-negative bacterium that may cause meningitis, sepsis or both.
In the past two decades, an increase in the
incidence of meningococcal disease in the Netherlands and various other
countries has been observed. This increase is mainly due to the
genotypically related lineage III meningococci. We hypothesize that a
genetic trait is responsible for the observed clonal and hypervirulent
phenotype of lineage III strains. Chromosomal DNA differences between
lineage III strains and non-lineage III strains were identified using
representational difference analysis. Thus, a 1.8 kb locus that is
specific for lineage III meningococci was identified. The locus
contains three open reading frames encoding the NmeSI restriction
modification system. The methyltransferase gene was cloned and
expressed in Escherichia coli. The site AGTACT was found to be
N4-cytosine modified by the recombinant enzyme. Chromosomal DNA of
lineage III strains was shown to be methylated at AGTACT sequences, in
contrast to DNA of non-lineage III strains. In conclusion, lineage III
strains differ from endemic strains by the presence of a specific
restriction modification system, causing differential methylation of
chromosomal DNA. The restriction modification system may contribute to
the clonal and hypervirulent character of lineage III strains by
influencing horizontal gene transfer and transcription.

<>

<1>Bart, A., van Passel, M.W., van Amsterdam, K., van der Ende, A.
<2>Direct detection of methylation in genomic DNA.
<3>Nucleic Acids Res.
<4>33
<5>e124
<6>2005
<7>The identification of methylated sites on bacterial genomic DNA would be a useful tool to
study the major roles of DNA methylation in prokaryotes: distinction of
self and nonself DNA, direction of post-replicative mismatch repair, control of
DNA replication and cell cycle, and regulation of gene expression. Three types of
methylated nucleobases are known: N6-methyladenine, 5-methylcytosine and
N4-methylcytosine. The aim of this study was to develop a method to detect all
three types of DNA methylation in complete genomic DNA. It was previously shown
that N6-methyladenine and 5-methylcytosine in plasmid and viral DNA can be
detected by intersequence trace comparison of methylated and unmethylated DNA. We
extended this method to include N4-methylcytosine detection in both in vitro and
in vivo methylated DNA. Furthermore, application of intersequence trace
comparison was extended to bacterial genomic DNA. Finally, we present evidence
that intrasequence comparison suffices to detect methylated sites in genomic DNA.
In conclusion, we present a method to detect all three natural types of DNA
methylation in bacterial genomic DNA. This provides the possibility to define the
complete methylome of any prokaryote.

<>

<1>Bart, M.J., van Gent, M., van der Heide, H.G., Boekhorst, J., Hermans, P., Parkhill, J., Mooi, F.R.
<2>Comparative genomics of prevaccination and modern Bordetella pertussis strains.
<3>BMC Genomics
<4>11
<5>627
<6>2010
<7>BACKGROUND: Despite vaccination since the 1950s, pertussis has persisted and
resurged. It remains a major cause of infant death worldwide and is the most
prevalent vaccine-preventable disease in developed countries. The resurgence of
pertussis has been associated with the expansion of Bordetella pertussis strains
with a novel allele for the pertussis toxin (Ptx) promoter, ptxP3, which have
replaced resident ptxP1 strains. Compared to ptxP1 strains, ptxP3 produce more
Ptx resulting in increased virulence and immune suppression. To elucidate how B.
pertussis has adapted to vaccination, we compared genome sequences of two ptxP3
strains with four strains isolated before and after the introduction vaccination.
RESULTS: The distribution of SNPs in regions involved in transcription and
translation suggested that changes in gene regulation play an important role in
adaptation. No evidence was found for acquisition of novel genes. Modern strains
differed significantly from prevaccination strains, both phylogenetically and
with respect to particular alleles. The ptxP3 strains were found to have diverged
recently from modern ptxP1 strains. Differences between ptxP3 and modern ptxP1
strains included SNPs in a number of pathogenicity-associated genes. Further,
both gene inactivation and reactivation was observed in ptxP3 strains relative to
modern ptxP1 strains. CONCLUSIONS: Our work suggests that B. pertussis adapted by
successive accumulation of SNPs and by gene (in)activation. In particular changes
in gene regulation may have played a role in adaptation.

<>

<1>Bartee, L., Bender, J.
<2>Two Arabidopsis methylation-deficiency mutations confer only partial effects on a methylated endogenous gene family.
<3>Nucleic Acids Res.
<4>29
<5>2127-2134
<6>2001
<7>In Arabidopsis a SWI2/SNF2 chromatin remodeling factor-related protein DDM1 and a cytosine
methyltransferase MET1 are required for maintenance of genomic cytosine methylation. Mutations
in either gene cause global demethylation. In this work we have assessed the effects of these
mutations on the PAI tryptophan biosynthetic gene family, which consists of four densely
methylated genes arranged as a tail-to-tail inverted repeat plus two unlinked singlet genes.
The methylation mutations caused only partial demethylation of the PAI loci: ddm1 had a strong
effect on the singlet genes but a weaker effect on the inverted repeat, whereas met1 had a
stronger effect on the inverted repeat than on the singlet genes. The double ddm1 met1 mutant
also displayed partial demethylation of the PAI genes, with a pattern similar to the ddm1
single mutant. To determine the relationship between partial methylation and expression for
the singlet PAI2 gene we constructed a novel reporter strain of Arabidopsis in which PAI2
silencing could be monitored by a blue fluorescent plant phenotype diagnostic of tryptophan
pathway defects. This reporter strain revealed that intermediate levels of methylation
correlate with intermediate suppression of the fluorescent phenotype.

<>

<1>Bartee, L., Malagnac, F., Bender, J.
<2>Arabidopsis cmt3 chromomethylase mutations block non-CG methylation and silencing of an endogenous gene.
<3>Genes Dev.
<4>15
<5>1753-1758
<6>2001
<7>Plants maintain cytosine methylation at CG and non-CG residues to control gene expression and
genome stability. In a screen for Arabidopsis mutants that alter methylation and silencing of
a densely methylated endogenous reporter gene, we recovered 11 loss-of-function alleles in the
CMT3 chromomethylase gene. The cmt3 mutants displayed enhanced expression and reduced
methylation of the reporter, particularly at non-CG cytosines. CNG methylation was also
reduced at repetitive centromeric sequences. Thus, CMT3 is a key determinant for non-CG
methylation. The lack of CMT homologs in animal genomes could account for the observation that
in contrast to plants, animals maintain primarily CG methylation.

<>

<1>Bartels, M.D., Hansen, L.H., Boye, K., Sorensen, S.J., Westh, H.
<2>An Unexpected Location of the Arginine Catabolic Mobile Element (ACME) in a USA300-Related MRSA StrainJ3 region.
<3>PLoS ONE
<4>6
<5>e16193
<6>2011
<7>In methicillin resistant Staphylococcus aureus (MRSA), the arginine catabolic mobile element
(ACME) was initially described in USA300 (t008-ST8) where it is located downstream of the
staphylococcal cassette chromosome mec (SCCmec). A common health-care associated MRSA in
Copenhagen, Denmark (t024-ST8) is clonally related to USA300 and is frequently PCR
positive for the ACME specific arcA-gene. This study is the first to describe an ACME element
upstream of the SCCmec in MRSA. By traditional SCCmec typing schemes, the SCCmec of t024-ST8
strain M1 carries SCCmec IVa, but full sequencing of the cassette revealed that the entire J3
region had no homology to published SCCmec IVa. Within the J3 region of M1 was a 1705 bp
sequence only similar to a sequence in S. haemolyticus strain JCSC1435 and 2941 bps with no
homology found in GenBank. In addition to the usual direct repeats (DR) at each extremity of
SCCmec, M1 had two new DR between the orfX gene and the J3 region of the SCCmec. The region
between the orfX DR (DR1) and DR2 contained the ccrAB4 genes. An ACME II-like element was
located between DR2 and DR3. The entire 26,468 bp sequence between DR1 and DR3 was highly
similar to parts of the ACME composite island of S. epidermidis strain ATCC12228. Sequencing
of an ACME negative t024-ST8 strain (M299) showed that DR1 and the sequence between DR1 and
DR3 was missing. The finding of a mobile ACME II-like element inserted downstream of orfX and
upstream of SCCmec indicates a novel recombination between staphylococcal species.

<>

<1>Barton, J.K.
<2>Transition metal complexes in the design of synthetic restriction enzymes.
<3>ACS Abstracts
<4>195
<5>44
<6>1988
<7>Restriction endonucleases recognize specific double-stranded DNA sequences and
cleave both strands to yield 5'phosphate and 3'hydroxyl termini.  Coordination
complexes have now been designed which simulate specific binding and catalytic
features of restriction endonucleases.  Based upon a matching of shapes and
symmetries, phenanthroline derivatives of ruthenium complexes have been
prepared which recognize a variety of distinct conformations along the strand.
Transition metal complexes may be converted to novel DNA cleaving molecules by
tethering of polyamine arms onto the DNA binding moiety.  These polyamine arms,
directed to the sugar-phosphate backbone provide chelating centers for divalent
metal ions such as Zn2+, Cd2+, and Pb2+, and upon activation with these metal
ions, double-stranded hydrolytic cleavage of the phosphodiester linkages is
observed.  Aspects of both binding specificity and reactivity of the metal
complexes with DNA will be described.

<>

<1>Barton, J.K., Basile, L.A., Paranawithana, S.R.
<2>The presence of zinc in the restriction enzyme EcoRI*.
<3>J. Biol. Chem.
<4>257
<5>7911-7914
<6>1982
<7>We have determined that the restriction endonuclease EcoRI contains 1.0 +- 0.1
eq of zinc/monomeric enzyme.  DNA cleavage by EcoRI is inhibited by
orthophenanthroline after preincubation of the enzyme with the chelating agent.
A similar inhibition by the nonchelating meta-phenanthroline is not seen.  The
sensitivity of the inhibition by the neutral ligand ortho-phenanthroline to
preincubation is consistent with the tightly bound and inaccessible nature of
the metal site.  Extensive dialysis against the ortho-phenanthroline inhibitor
leads to the release of the bound metal with the concomitant loss of enzyme
activity.  The tightly bound Zn2+ cation, then, appears to be necessary for
enzyme function.  The finding of zinc in EcoRI further illustrates the ubiquity
of Zn2+ to DNA-protein complexes.

<>

<1>Barton, J.K., Paranawithana, S.R.
<2>Restriction endonuclease EcoRI alters the Enantiomeric Preference of Chiral Metallointercalators for DNA:  an illustration of a protein-induced DNA conformational change.
<3>Biochemistry
<4>25
<5>2205-2211
<6>1986
<7>A conformational change in the DNA plasmid ColE1 appears to occur upon specific
binding of the restriction endonuclese EcoRI.  Enzyme association alters the
chiral discrimination found in binding metallointercalators to DNA sites.  The
complexes tris(1,10-phenanthroline) ruthenium(II), Ru(phen) 32+, and
tris(4,7-diphenyl-1,10-phenanthroline)cobalt(III), Co(DIP)33+, in general, bind
stereoselectively to DNA helices, with enantiomers processing the D
configuration bound preferentially by right-handed B-DNA.  In the presence of
EcoRI, however, this enantioselectivity is altered.  The chiral intercalators,
at micromolar concentrations, inhibit the reaction of EcoRI, but for each
enantiomeric pair it is the K enantiomer, which binds only poorly to a B-DNA
helix, that inhibits EcoRI preferentially.  Kinetic studies in the presence of
K in the presence of K-Ru(DIP)32+ indicate that the enzyme inhibition occurs as
a result of the K enantiomer binding to the enzyme-DNA complex as well as to
the free enzyme.  Furthermore, photolytic strand cleavage experiments using
Co(DIP)33+ indicate that the metal complex interacts directly at the
protein-bound DNA site.  Increasing concentrations of bound EcoRI stimulate
photoactivated cleavage of the DNA helix by K-Co(DIP)33+, until a protein
concentration is reached where specific DNA recognition sites are saturated
with enzyme.  Thus, although K-Co(DIP)33+ does not bind closely to the DNA in
the absence of enzyme, specific binding of EcoRI appears to alter the DNA
structure so as to permit the close association of the K isomer to the DNA
helix.  Mapping experiments demonstrate that this association leads to
photocleavage of DNA by the cobalt complex at or very close to the EcoRI
recognition site.  This study provides evidence that in solution, under
enzymatic conditions, a DNA-binding protein may distort the DNA helical
structure and further illustrates how small molecular probes of DNA
conformation might be used in examining the structure of protein-bound DNA
sites.

<>

<1>Baryshev, M.M., Buryanov, Y.I., Kosykh, V.G., Bayev, A.A.
<2>Isolation, purification and some properties of restrictase and methylase BstNI from Bacillus stearothermophilus.
<3>Biokhimiia
<4>54
<5>1894-1903
<6>1989
<7>Restriction-methylation enzymes BstNI from Bacillus stearothermophilus were
isolated and purified.  These enzymes are related to a new class of
restriction-methylation enzymes of the second type, whose modifying component
is N4-cytosine-DNA-methylase.  Both enzymes recognize the DNA sequence
CC(A/T)GG.  Restrictase BstNI is a protein made up of one subunit with a
molecular mass of 25 kDa.  The temperature optima for restrictase and methylase
BstNI are around 60C.  Possible uses of BstNI restriction-methylation enzymes
for the analysis of cytosine methylation in bacterial and higher plant DNA are
discussed.

<>

<1>Barzel, A., Privman, E., Burstein, D., Gophna, U., Kupiec, M., Pupko, T.
<2>Method for searching for homing endonucleases, their genes and their targets.
<3>International Patent Office
<4>WO 2009101625 A
<5>
<6>2009
<7>A computer implemented method for generating nucleotide sequences containing candidate homing
endonuclease genes.  A search is performed in a database stored on a storage medium of
nucleotide sequences for amino acid sequences having a subsequence having a homology level
with the translation of asubsequence of eone or more predetermined HEGs.  For each amino acid
sequence generated by the search, one or more nucleotide sequences are retrieved encoding the
amino acid sequence.  The results of this search used in a second search of a database stored
on a storage medium to generate the HEG containing sequences.

<>

<1>Barzel, A., Privman, E., Peeri, M., Naor, A., Shachar, E., Burstein, D., Lazary, R., Gophna, U., Pupko, T., Kupiec, M.
<2>Native homing endonucleases can target conserved genes in humans and in animal models.
<3>Nucleic Acids Res.
<4>39
<5>6646-6659
<6>2011
<7>In recent years, both homing endonucleases (HEases) and zinc-finger nucleases (ZFNs) have been
engineered and selected for the targeting of
desired human loci for gene therapy. However, enzyme engineering is
lengthy and expensive and the off-target effect of the manufactured
endonucleases is difficult to predict. Moreover, enzymes selected to
cleave a human DNA locus may not cleave the homologous locus in the
genome of animal models because of sequence divergence, thus hampering
attempts to assess the in vivo efficacy and safety of any engineered
enzyme prior to its application in human trials. Here, we show that
naturally occurring HEases can be found, that cleave desirable human
targets. Some of these enzymes are also shown to cleave the homologous
sequence in the genome of animal models. In addition, the distribution
of off-target effects may be more predictable for native HEases. Based
on our experimental observations, we present the HomeBase algorithm,
database and web server that allow a high-throughput computational
search and assignment of HEases for the targeting of specific loci in
the human and other genomes. We validate experimentally the predicted
target specificity of candidate fungal, bacterial and archaeal HEases
using cell free, yeast and archaeal assays.

<>

<1>Barzilai, R.
<2>SV40 DNA:  Quantitative conversion of closed circular to open circular form by an ethidium bromide-restricted endonuclease.
<3>J. Mol. Biol.
<4>74
<5>739-742
<6>1973
<7>Closed circular molecules of SV40 DNA can be quantitatively converted to the
open circular form by digestion with endonucleases in the presence of ethidium
bromide.  Under these conditions almost no molecules were digested beyond the
open circular form, whereas in the absence of the dye the DNA was completely
fragmented.

<>

<1>Basavanna, U., Gonzalez-Escalona, N., Timme, R., Datta, S., Schoen, B., Brown, E.W., Zink, D., Sharma, S.K.
<2>Draft Genome Sequence of a Clostridium botulinum Isolate from Water Used for Cooling at a Plant Producing Low-Acid Canned Foods.
<3>Genome Announcements
<4>1
<5>e00200-12
<6>2013
<7>Clostridium botulinum is a pathogen of concern for low-acid canned foods. Here we report draft
genomes of a neurotoxin-producing C. botulinum strain isolated from
water samples used for cooling low-acid canned foods at a canning facility. The
genome sequence confirmed that this strain belonged to C. botulinum serotype B1,
albeit with major differences, including thousands of unique single nucleotide
polymorphisms (SNPs) compared to other genomes of the same serotype.

<>

<1>Bashtrykov, P., Jankevicius, G., Jurkowska, R.Z., Ragozin, S., Jeltsch, A.
<2>The UHRF1 Protein Stimulates the Activity and Specificity of the Maintenance DNA Methyltransferase DNMT1 by an Allosteric Mechanism.
<3>J. Biol. Chem.
<4>289
<5>4106-4115
<6>2014
<7>The ubiquitin-like, containing PHD and RING finger domains protein 1 (UHRF1) is essential for
maintenance DNA methylation by DNA methyltransferase 1 (DNMT1). UHRF1 has been shown to
recruit DNMT1 to replicated DNA by the ability of its SET and RING-associated (SRA) domain to
bind to hemimethylated DNA. Here, we demonstrate that UHRF1 also increases the activity of
DNMT1 by almost 5-fold. This stimulation is mediated by a direct interaction of both proteins
through the SRA domain of UHRF1 and the replication focus targeting sequence domain of DNMT1,
and it does not require DNA binding by the SRA domain. Disruption of the interaction between
DNMT1 and UHRF1 by replacement of key residues in the replication focus targeting sequence
domain led to a strong reduction of DNMT1 stimulation. Additionally, the interaction with
UHRF1 increased the specificity of DNMT1 for methylation of hemimethylated CpG sites. These
findings show that apart from the targeting of DNMT1 to the replicated DNA UHRF1 increases the
activity and specificity of DNMT1, thus exerting a multifaceted influence on the maintenance
of DNA methylation.

<>

<1>Bashtrykov, P., Ragozin, S., Jeltsch, A.
<2>Mechanistic details of the DNA recognition by the Dnmt1 DNA methyltransferase.
<3>FEBS Lett.
<4>586
<5>1821-1823
<6>2012
<7>A recently solved Dnmt1-DNA crystal structure revealed several enzyme-DNA contacts and large
structural rearrangements of the DNA at
the target site, including the flipping of the non-target strand base
of the base pair flanking the CpG site and formation of a non-canonical
base pair between the non-target strand Gua and the flanking base pair.
Here, we show that the contacts of the enzyme to the target base and
the Gua:5mC base pair that are observed in the structure are very
important for catalytic activity. The contacts to the non-target strand
Gua are not important since its exchange by Ade stimulated activity.
Except target base flipping, we could not find evidence that the DNA
rearrangements have a functional role.

<>

<1>Baskunov, V.B., Subach, F.V., Kolbanovskiy, A., Kolbanovskiy, M., Eremin, S.A., Johnson, F., Bonala, R., Geacintov, N.E., Gromova, E.S.
<2>Effects of benzo[a]pyrene-deoxyguanosine lesions on DNA methylation catalyzed by EcoRII DNA methyltransferase and on DNA cleavage effected  by EcoRII restriction endonuclease.
<3>Biochemistry
<4>44
<5>1054-1066
<6>2005
<7>DNA methylation is an important cellular mechanism for controlling gene expression. Whereas
the mutagenic properties of many DNA adducts, e.g.,
those arising from polycyclic aromatic hydrocarbons, have been widely
studied, little is known about their influence on DNA methylation. We
have constructed site-specifically modified 18-mer oligodeoxynucleotide
duplexes containing a pair of stereoisomeric adducts derived from a
benzo[a]pyrene-derived diol epoxide [(+)- and
(-)-r7,t8-dihydroxy-t9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene, or
B[a]PDE] bound to the exocyclic amino group of guanine. The adducts,
either (+)- or (-)-trans-anti-B[a]P-N-2-dG (G*), positioned either at
the 5'-side or the 3'-side deoxyguanosine residue in the recognition
sequence of EcoRII restriction-modification enzymes (5'-...CCA/TGG...)
were incorporated into 18-mer oligodeoxynucleotide duplexes. The
effects of these lesions on complex formation and the catalytic
activity of the EcoRII DNA methyltransferase (M.EcoRII) and EcoRII
restriction endonuclease (R.EcoRII) were investigated. The M.EcoRII
catalyzes the transfer of a methyl group to the C5 position of the
3'-side cytosine of each strand of the recognition sequence, whereas
R.EcoRII catalyzes cleavage of both strands. The binding of R.EcoRII to
the oligodeoxynucleotide duplexes and the catalytic cleavage were
completely abolished when G* was positioned at the 3'-side dG position
(5'-...CCTGG*...). When G* was at the 5'-side dG position, binding was
moderately diminished, but cleavage was completely blocked. In the case
of M.EcoRII, binding is diminished by factors of 5-30 but the catalytic
activity was either abolished or reduced 4-80-fold when the adducts
were located at either position. Somewhat smaller effects were observed
with hemimethylated oligodeoxynucleotide duplexes. These findings
suggest that epigenetic effects, in addition to genotoxic effects, need
to be considered in chemical carcinogenesis initiated by B[a]PDE, since
the inhibition of methylation may allow the expression of genes that
promote tumor development.

<>

<1>Basnakyan, A.G., Votrin, I.I.
<2>Restriction endonucleases as a possible factor in the evolution of DNA nucleotide composition.
<3>Zh. Evol. Biokhim. Fiziol.
<4>19
<5>20-24
<6>1983
<7>On the basis of a statistical analysis of the nucleotide and codogenic
composition of the sites for restriction endonucleases of type II it was
concluded that restriction endoncleases may perform an evolutionary function as
catalysts of the site-specific accumulation of the bacterial pre-genome in
parallel with a change in its nucleotide composition in the direction of its
enrichment with AT pairs and the amino acid composition of the proteins in the
direction of a decrease in the content of alanine, arginine, proline and
glycine.

<>

<1>Bassi, D., Fontana, C., Gazzola, S., Pietta, E., Puglisi, E., Cappa, F., Cocconcelli, P.S.
<2>Draft Genome Sequence of Clostridium tyrobutyricum Strain UC7086, Isolated from Grana Padano Cheese with Late-Blowing Defect.
<3>Genome Announcements
<4>1
<5>e00614-13
<6>2013
<7>Clostridium tyrobutyricum is considered the main agent of late-blowing defect in  the
production of hard cheese. Here, we described the draft genome sequences and
annotation of C. tyrobutyricum strain UC7086, which was isolated from Grana
Padano cheese with blowing defect, and C. tyrobutyricum DSM 2637 type strain in a
comparative study.

<>

<1>Bassing, C.H., Kim, Y.-G., Li, L., Chandrasegaran, S.
<2>Overproduction, purification and characterization of M.HinfI methyltransferase and its deletion mutant.
<3>Gene
<4>113
<5>83-88
<6>1992
<7>We have used the polymerase chain reaction to alter transcriptional and translational signals
surrounding the hinfIM gene [encoding M.HinfI methyltransferase (MTase)] so as to achieve
overexpression in Escherichia coli. The PCR-generated hinfIM gene was subcloned in a
high-expression vector under control of the hybrid trp-lac promoter. In addition, the positive
retroregulator stem-loop sequence derived from the crystal protein-encoding gene of Bacillus
thuringiensis was inserted downstream from hinfIM. Using a similar approach, we have also
constructed overproducer clones of a deletion mutant of M.HinfI MTase that has 97 amino acids
from the C terminus deleted. The plasmid from the mutant clones is fully protected from HinfI
restriction endonuclease digestion. It appears that the functional properties (recognition and
catalytic functions) are encoded within this mutant gene. The overproducer clones yield the
wild type (wt) and the mutant enzymes to about 10% of total cellular protein upon induction
with 1 mM IPTG. The wt M.HinfI and the mutant MTase were purified to near electrophoretic
homogeneity by phosphocellulose, DEAE and gel chromatography. Their monomer sizes by
SDS/polyacrylamide-gel electrophoresis are 43 kDa and 31 kDa, respectively, in good agreement
with that predicted from the nucleotide sequence. DNA methylation experiments with purified
enzymes using single-strand and double-strand M13mp18 DNA substrates indicate that while wt
enzyme methylates both forms of DNA substrates, the mutant enzyme appears to preferentially
methylate ss DNA substrate.

<>

<1>Bates, P.J., Hammond, G.B., Xu, B., Aponte, J.C., Malik, M.T., Martin, A.N., Choi, E.W., Thomas, S.D., Casson, L.K., Trent, J.O.
<2>Discovery of XB05, a potent antiproliferative agent and novel non-nucleoside inhibitor of DNA methyltransferase 1.
<3>Proc. Amer. Assoc. Cancer Res.
<4>49
<5>783
<6>2008
<7>While investigating the biological properties of fluorine-containing organic molecules, we
recently identified a novel small molecule, named XB05, which has tumor-selective
antiproliferative activity. The molecule was submitted to the Developmental Therapeutics
Program (DTP) at the National Cancer Institute (NCI) for testing in the 60 human tumor cell
line screen. The results confirmed that XB05 has potent activity against a diverse range of
cancer cell lines and also revealed an unusual activity profile, with a more than 1000-fold
difference in GI50 values between the most and least sensitive lines. For example, colon,
brain, renal, prostate and breast tumor cells were particularly sensitive to XB05, whereas
other tumor types were substantially less responsive. The GI50 values for the sensitive cell
lines were in the 10 - 100 nM range.
We analyzed the DTP data using the on-line COMPARE program, which allows comparison of the
activity profile with the 43,000 compounds in the public database and which has been validated
as grouping together agents with similar mechanisms of action, irrespective of their
structure. When we used XB05 as a COMPARE seed, we found a strong correlation (Pearson
coefficient 0.82) with a compound known as halomon.
Halomon is a marine-derived natural product, which has promising anticancer activity. However,
its development has been severely limited because attempts to re-isolate it from its natural
source or to develop an efficient synthesis have not enabled production of sufficient compound
for testing. Halomon was recently reported to be an inhibitor of human DNA methyltransferase 1
(DNMT1), the enzyme responsible for maintaining DNA methylation after each cell division.
DNMT1 is considered an attractive target for epigenetic therapy of cancer because
methylation-induced silencing of tumor suppressor genes occurs frequently in cancer cells.
XB05 has now been tested in various cell-free and cell-based assays of DNA methylation. We
find that it is a potent inhibitor of recombinant DNMT1 activity in vitro, with an IC50 of 10
nM. Moreover, XB05 could reactivate expression of several silenced tumor suppressor genes in
cancer cells at concentrations of 100 nM or less.
In conclusion, XB05 is a novel agent with potent antiproliferative activity against cell lines
derived from common cancers and a new type of DNMT1 inhibitor. XB05 is a mechanistic mimic of
a natural product named halomon, but, in contrast to halomon, XB05 can be easily and
inexpensively synthesized in large quantities. XB05 may also have substantial advantages over
other existing DNMT inhibitors such as 5-azacytidine and decitabine. XB05 is more active than
these agents in cell-free or cell-based DNMT assays and will likely be less toxic because it
cannot be incorporated into DNA. Also, whereas 5-azacytidine and decitabine are rapidly
degraded in aqueous solutions, XB05 appears to be completely stable.

<>

<1>Bath, A.J., Milsom, S.E., Gormley, N.A., Halford, S.E.
<2>Many type IIs restriction endonucleases interact with two recognition sites before cleaving DNA.
<3>J. Biol. Chem.
<4>277
<5>4024-4033
<6>2002
<7>Type IIs restriction endonucleases recognize asymmetric DNA sequences and cleave both DNA
strands at fixed positions, typically several base pairs away from the recognition site.
These enzymes are generally monomers that transiently associate to form dimmers to cleave both
strands.  Their reactions could involve bridging interactions between two copies of their
recognition sequence.  To examine this possibility, several type IIs enzymes were tested
against substrates with either one or two target sites.  Some of the enzymes cleaved the DNA
with two target sites at the same rate as that with one site, but most cut their two-site
substrate more rapidly than the one-site DNA.  In some cases, the two sites were cut
sequentially, at rates that were equal to each other but that exceeded the rate on the
one-site DNA.  In another case, the DNA with two sites was cleaved rapidly at one site, but
the residual site was cleaved at a much slower rate.  In a further example, the two sites were
cleaved concertedly to give directly the final products cut at both sites.  Many type IIs
enzymes thus interact with two copies of their recognition sequence before cleaving DNA,
although via several different mechanisms.

<>

<1>Batista, D.F., Freitas, N.O.C., Leite, L.R., Varani, A.M., Araujo, F.M., Salim, A., Almeida, A.M., Ribeiro, S.A., Oliveira, G.C., Barrow, P.A., Berchieri, J.A.
<2>Draft Genome Sequence of Salmonella enterica subsp. enterica Serovar Gallinarum Biovar Pullorum Strain FCAV198, a Brazilian Chicken Pathogen.
<3>Genome Announcements
<4>2
<5>e00028-14
<6>2014
<7>Salmonella enterica subsp. enterica serovar Gallinarum biovar Pullorum is a bird-restricted
pathogen which causes pullorum disease. The strain FCAV198 was
isolated from a pool of chicken ovaries in Brazil, and its genome may be helpful
for studies involving molecular mechanisms related to pathogenesis and other
related applications.

<>

<1>Battisti, J.M., Minnick, M.F.
<2>Development of a system for genetic manipulation of Bartonella bacilliformis.
<3>Appl. Environ. Microbiol.
<4>65
<5>3441-3448
<6>1999
<7>Lack of a system for site-specific genetic manipulation has severely hindered studies on the
molecular biology of all Bartonella species. We report the first site-specific mutagenesis and
complementation for a Bartonella species. A highly transformable strain of B. bacilliformis,
termed JB584, was isolated and found to exhibit a significant increase in transformation
efficiency with the broad-host-range plasmid pBBR1MCS-2, relative to wild-type strains.
Restriction analyses of genomic preparations with the methylation-sensitive restriction
enzymes ClaI and StuI suggest that strain JB584 possesses a dcm methylase mutation that
contributes to its enhanced transformability. A suicide plasmid, pUB1, which contains a
polylinker, a pMB1 replicon, and a nptI kanamycin resistance cassette, was constructed. An
internal 508-bp fragment of the B. bacilliformis flagellin gene (fla) was cloned into pUB1 to
generate pUB508, a fla-targeting suicide vector. Introduction of pUB508 into JB584 by
electroporation generated eight Kan(r) clones of B. bacilliformis.  Characterization of one of
these strains, termed JB585, indicated that allelic exchange between pUB508 and fla had
occurred. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis,
immunoblotting, and electron microscopy showed that synthesis of flagellin encoded by fla and
secretion/assembly of flagella were abolished.  Complementation of fla in trans was
accomplished with a pBBR1MCS recombinant containing the entire wild-type fla gene (pBBRFLAG).
These data conclusively show that inactivation of fla results in a bald, nonmotile phenotype
and that pMB1 and REP replicons make suitable B. bacilliformis suicide and shuttle vectors,
respectively. When used in conjunction with the highly transformable strain JB584, this system
for site-specific genetic manipulation and complementation provides a new venue for studying
the molecular biology of B. bacilliformis.

<>

<1>Batzilla, J., Hoper, D., Antonenka, U., Heesemann, J., Rakin, A.
<2>Complete Genome Sequence of Yersinia enterocolitica subsp. palearctica Serogroup O:3.
<3>J. Bacteriol.
<4>193
<5>2067
<6>2011
<7>We report here the first finished and annotated genome sequence of a representative of the
most epidemiologically successful Yersinia group, Y.
enterocolitica subsp. palearctica strain Y11, serotype O:3, biotype 4.
This strain is a certified type strain of the German DSMZ collection (DSM
no. 13030; Yersinia enterocolitica subsp. palearctica) that was isolated
from the stool of a human patient (H. Neubauer, S. Aleksic, A. Hensel, E.
J. Finke, and H. Meyer. Int. J. Med. Microbiol. 290:61-64, 2000).

<>

<1>Baumgart, M., Unthan, S., Ruckert, C., Sivalingam, J., Grunberger, A., Kalinowski, J., Bott, M., Noack, S., Frunzke, J.
<2>Construction of a prophage-free variant of Corynebacterium glutamicum ATCC 13032 - a platform strain for basic research and industrial biotechnology.
<3>Appl. Environ. Microbiol.
<4>79
<5>6006-6015
<6>2013
<7>The activity of bacteriophages and phage-related mobile elements is a major
source for genome rearrangements and genetic instability of their bacterial
hosts. The genome of the industrial amino acid producer Corynebacterium
glutamicum ATCC 13032 contains three prophages (CGP1-3) of so far unknown
functionality. Several phage genes are regularly expressed and the large prophage
CGP3 ( approximately 190 kbp) has recently been shown to be induced under certain
stress conditions. Here, we present the construction of MB001, a prophage-free
variant of C. glutamicum ATCC 13032 with a 6 % reduced genome. This strain does
not show any unfavorable properties during extensive phenotypic characterization
under various standard and stress conditions. As expected, we observed an
improved growth and fitness of MB001 under SOS-response inducing conditions that
trigger CGP3 induction in the wild type strain. Further studies revealed that
MB001 has a significantly increased transformation efficiency and produced about
30 % more of the heterologous model protein eYFP, presumably as a consequence of
an increased plasmid copy number. These effects were attributed to the loss of
the restriction-modification system (cg1996-98) located within CGP3. The deletion
of the prophages without any negative effect results in a novel platform strain
for metabolic engineering and represents a useful step towards the construction
of a C. glutamicum chassis genome of strain ATCC 13032 for biotechnological
applications and synthetic biology.

<>

<1>Baumstark, B.R., Roberts, R.J., RajBhandary, U.L.
<2>Use of short synthetic DNA duplexes as substrates for the restriction endonucleases HpaII and MnoI.
<3>J. Biol. Chem.
<4>254
<5>8943-8950
<6>1979
<7>In an attempt to determine whether cleavage by restriction enzymes is specified solely by the
nucleotide sequence at the recognition site or whether additional factors (e.g. the size of
the DNA or partial disruption of the DNA helix to form cruciform structures) are involved in
recognition, two restriction enzymes, HpaII and MnoI, were tested for their ability to cleave
synthetic DNA duplexes of limited size and well-defined sequence.  DNA duplexes ranging from 6
to 13 base pairs in length were shown to be recognized with high efficiency by HpaII and were
cleaved specifically within the HpaII recognition sequence.  5' d-C^C-G-G 3'  3' G-G-C^C-d
5'.  HpaII was totally inactive against single-stranded DNA; therefore, both strands of the
DNA duplex are necessary for cleavage to occur.  In two of the DNA duplexes, the recognition
sequences were located at the end of a base paired region and were thus almost certainly
present in the form of a linear duplex.  The fact that HpaII could cleave these duplexes
suggests that the information for HpaII recognition resides in the nucleotide sequence alone.
By incubation of these synthetic duplexes with restriction nuclease from MnoI we have shown
that the cleavage site for MnoI is identical with that of HpaII.  Although HpaII and MnoI
nucleases are isoschizomers, they do difer somewhat in their mode of action on the short
synthetic DNA duplexes.  With the duplexes used as substrates, it was found that whereas the
HpaII nuclease preferentially cleaved the pyrimidine-rich strand (3 to 4 times over the
purine-rich strand), the MnoI nuclease cleaved both strands at an equal rate.

<>

<1>Baxter, B.K., Topal, M.D.
<2>Formation of a cleavasome: enhancer DNA-2 stabilizes an active conformation of NaeI dimer.
<3>Biochemistry
<4>32
<5>8291-8298
<6>1993
<7>Cleavage of DNA by NaeI-type restriction enzymes is stimulated by a DNA element with affinity
for the activator site of the enzyme: a cleavage-enhancer DNA element. Measurements of the
mobility of NaeI activity in comparison with protein standards on gel permeation columns and
glycerol gradients demonstrated that NaeI, without enhancer, can form a 70,000 MW dimer. The
dimer, however is inactive: it could not cleave the resistant NaeI site in M13mp18 DNA in the
absence of enhancer. In cleavage assays, enhancer stimulated either DNA nicking or DNA
cleavage, depending upon NaeI concentration, and reduced the NaeI concentration required for
the transition from nicking to cleavage activity. A gel mobility-shift assay of the
interaction of NaeI with enhancer showed the formation of two complexes. Results using
different sized DNAs and different percentage acrylamide gels for gel mobility-shift analysis
implied that the two complexes were caused by NaeI monomer and dimer structures rather than
one and two DNA binding. Dimer formation increased with the affinity of enhancer for NaeI. UV
cross-linking captured the NaeI-enhancer complex; electrophoretic analysis of the cross-linked
products showed NaeI dimer bound to enhancer. These results imply a model for cleavage
enhancement in which enhancer binding stabilizes an active NaeI dimer conformation
(cleavasome) that cleaves both DNA strands before dissociating.

<>

<1>Baxter, S., Lambert, A.R., Kuhar, R., Jarjour, J., Kulshina, N., Parmeggiani, F., Danaher, P., Gano, J., Baker, D., Stoddard, B.L., Scharenberg, A.M.
<2>Engineering domain fusion chimeras from I-OnuI family LAGLIDADG homing endonucleases.
<3>Nucleic Acids Res.
<4>40
<5>7985-8000
<6>2012
<7>Although engineered LAGLIDADG homing endonucleases (LHEs) are finding increasing  applications
in biotechnology, their generation remains a challenging,
industrial-scale process. As new single-chain LAGLIDADG nuclease scaffolds are
identified, however, an alternative paradigm is emerging: identification of an
LHE scaffold whose native cleavage site is a close match to a desired target
sequence, followed by small-scale engineering to modestly refine recognition
specificity. The application of this paradigm could be accelerated if methods
were available for fusing N- and C-terminal domains from newly identified LHEs
into chimeric enzymes with hybrid cleavage sites. Here we have analyzed the
structural requirements for fusion of domains extracted from six single-chain
I-OnuI family LHEs, spanning 40-70% amino acid identity. Our analyses demonstrate
that both the LAGLIDADG helical interface residues and the linker peptide
composition have important effects on the stability and activity of chimeric
enzymes. Using a simple domain fusion method in which linker peptide residues
predicted to contact their respective domains are retained, and in which limited
variation is introduced into the LAGLIDADG helix and nearby interface residues,
catalytically active enzymes were recoverable for approximately 70% of domain
chimeras. This method will be useful for creating large numbers of chimeric LHEs
for genome engineering applications.

<>

<1>Bayliss, C.D., Bidmos, F.A., Anjum, A., Manchev, V.T., Richards, R.L., Grossier, J.P., Wooldridge, K.G., Ketley, J.M., Barrow, P.A., Jones, M.A., Tretyakov, M.V.
<2>Phase variable genes of Campylobacter jejuni exhibit high mutation rates and specific mutational patterns but mutability is not the major determinant of  population structure during host colonization.
<3>Nucleic Acids Res.
<4>40
<5>5876-5889
<6>2012
<7>Phase variation of surface structures occurs in diverse bacterial species due to  stochastic,
high frequency, reversible mutations. Multiple genes of Campylobacter
jejuni are subject to phase variable gene expression due to mutations in polyC/G
tracts. A modal length of nine repeats was detected for polyC/G tracts within C.
jejuni genomes. Switching rates for these tracts were measured using
chromosomally-located reporter constructs and high rates were observed for cj1139
(G8) and cj0031 (G9). Alteration of the cj1139 tract from G8 to G11 increased
mutability 10-fold and changed the mutational pattern from predominantly
insertions to mainly deletions. Using a multiplex PCR, major changes were
detected in 'on/off' status for some phase variable genes during passage of C.
jejuni in chickens. Utilization of observed switching rates in a stochastic,
theoretical model of phase variation demonstrated links between mutability and
genetic diversity but could not replicate observed population diversity. We
propose that modal repeat numbers have evolved in C. jejuni genomes due to
molecular drivers associated with the mutational patterns of these polyC/G
repeats, rather than by selection for particular switching rates, and that
factors other than mutational drift are responsible for generating genetic
diversity during host colonization by this bacterial pathogen.

<>

<1>Bayliss, C.D., Callaghan, M.J., Moxon, E.R.
<2>High allelic diversity in the methyltransferase gene of a phase variable type III restriction-modification system has implications for the fitness  of Haemophilus influenzae.
<3>Nucleic Acids Res.
<4>34
<5>4046-4059
<6>2006
<7>Phase variable restriction-modification (R-M) systems are widespread in Eubacteria.
Haemophilus influenzae encodes a phase variable homolog of
Type III R-M systems. Sequence analysis of this system in 22 non-typeable
H.influenzae isolates revealed a hypervariable region in the central
portion of the mod gene whereas the res gene was conserved. Maximum
likelihood (ML) analysis indicated that most sites outside this
hypervariable region experienced strong negative selection but evidence of
positive selection for a few sites in adjacent regions. A phylogenetic
analysis of 61 Type III mod genes revealed clustering of these
H.influenzae mod alleles with mod genes from pathogenic Neisseriae and,
based on sequence analysis, horizontal transfer of the mod-res complex
between these species. Neisserial mod alleles also contained a
hypervariable region and all mod alleles exhibited variability in the
repeat tract. We propose that this hypervariable region encodes the target
recognition domain (TRD) of the Mod protein and that variability results
in alterations to the recognition sequence of this R-M system. We argue
that the high allelic diversity and phase variable nature of this R-M
system have arisen due to selective pressures exerted by diversity in
bacteriophage populations but also have implications for other fitness
attributes of these bacterial species.

<>

<1>Bayona-Bafaluy, M.P., Blits, B., Battersby, B.J., Shoubridge, E.A., Moraes, C.T.
<2>Rapid directional shift of mitochondrial DNA heteroplasmy in animal tissues by a mitochondrially targeted restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>14392-14397
<6>2005
<7>Frequently, mtDNA with pathogenic mutations coexist with wild-type genomes (mtDNA
heteroplasmy). Mitochondrial dysfunction and disease ensue only
when the proportion of mutated mtDNAs is high, thus a reduction in this
proportion should provide an effective therapy for these disorders. We
developed a system to decrease specific mtDNA haplotypes by expressing a
mitochondrially targeted restriction endonuclease, ApaLI, in cells of
heteroplasmic mice. These mice have two mtDNA haplotypes, of which only
one contains an ApaLI site. After transfection of cultured hepatocytes
with mitochondrially targeted ApaLI, we found a rapid, directional, and
complete shift in mtDNA heteroplasmy (2-6 h). We tested the efficacy of
this approach in vivo, by using recombinant viral vectors expressing the
mitochondrially targeted ApaLI. We observed a significant shift in mtDNA
heteroplasmy in muscle and brain transduced with recombinant viruses. This
strategy could prevent disease onset or reverse clinical symptoms in
patients harboring certain heteroplasmic pathogenic mutations in mtDNA.

<>

<1>Bayot-Custodio, A.N., Alcantara, E.P., Zulaybar, T.O.
<2>Draft Genome Sequence of Insecticidal Streptomyces sp. Strain PCS3-D2, Isolated from Mangrove Soil in Philippines.
<3>Genome Announcements
<4>2
<5>e00448-14
<6>2014
<7>A draft genome sequence of a Streptomyces sp. isolated from mangrove soil in Cebu,
Philippines, is described here. This isolate produced compounds with
contact insecticidal activity against important corn pests. The genome contains
7,479,793 bp (in 27 scaffolds), 6,297 predicted genes, and 29 secondary
metabolite biosynthetic gene clusters.

<>

<1>Beaber, J.W., Hochhut, B., Waldor, M.K.
<2>Genomic and Functional Analyses of SXT, an Integrating Antibiotic Resistance Gene Transfer Element Derived from Vibrio cholerae.
<3>J. Bacteriol.
<4>184
<5>4259-4269
<6>2002
<7>SXT is representative of a family of conjugative-transposon-like mobile
genetic elements that encode multiple antibiotic resistance genes. In
recent years, SXT-related conjugative, self-transmissible integrating
elements have become widespread in Asian Vibrio cholerae. We have
determined the 100-kb DNA sequence of SXT. This element appears to be a
chimera composed of transposon-associated antibiotic resistance genes
linked to a variety of plasmid- and phage-related genes, as well as to
many genes from unknown sources. We constructed a nearly comprehensive set
of deletions through the use of the one-step chromosomal gene inactivation
technique to identify SXT genes involved in conjugative transfer and
chromosomal excision. SXT, unlike other conjugative transposons, utilizes
a conjugation system related to that encoded by the F plasmid. More than
half of the SXT genome, including the composite transposon-like structure
that contains its antibiotic resistance genes, was not required for its
mobility. Two SXT loci, designated setC and setD, whose predicted amino
acid sequences were similar to those of the flagellar regulators FlhC and
FlhD, were found to encode regulators that activate the transcription of
genes required for SXT excision and transfer. Another locus, designated
setR, whose gene product bears similarity to lambdoid phage CI repressors,
also appears to regulate SXT gene expression.

<>

<1>Beary, T.P., Braymer, H.D., Achberger, E.C.
<2>Evidence of participation of McrBs in McrBC restriction in Escherichia coli K-12.
<3>J. Bacteriol.
<4>179
<5>7768-7775
<6>1997
<7>The McrBC restriction system has the ability to restrict DNA containing
5-hydroxymethylcytosine, N4-methylcytosine, and 5-methylcytosine at specific sequences.  The
mcrB gene produces two gene products.  The complete mcrB open reading frame produces a 51-kDa
protein (McrBL) and a 33-kDa protein (McrBS).  The smaller McrB polypeptide is produced from
an in-frame, internal translational start site in the mcrB gene.  The McrBs is unknown,
although there has been speculation that it plays some role in the modulation of McrBC
restriction.  Studies of the function of McrBs have been challenging since it is produced in
frame with McrBL.  In this study, we tested the effects of underproduction (via antisense RNA)
and overproduction (via gene dosage) of mcrBC gene products on restriction levels of the
mcrBC+ strain JM107.  Among the parameters monitoredwas the induction of SOS responses, which
indicate DNA damage.  Evidence from this study suggests that McrBS is necessary for
stabilization of the McrBC restriction complex in vivo.

<>

<1>Beary, T.P., Ross, T.K., Braymer, H.D., Achberger, E.C.
<2>Characterization of genes involved in the McrB restriction system of Escherichia coli K12.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>89
<5>180
<6>1989
<7>The McrB system of E. coli K12 restricts DNA modified at the sequence 'GmC'.
Two of the genes involved in the restriction of 5-methylcytosine
(5mC)-containing DNA are mcrB and mcrC.  The nucleic acid sequences of these
genes were used to predict open reading frames encoding peptides consistent
with the 51-kilodalton (kDa) mcrB gene product and the 39-kDa mcrC gene
product.  Based on the nucleotide sequence and the analysis of frameshift
mutations within the mcrB gene, an internal, in phase translational start was
identified specifying a peptide of 33 kDa.  It has been shown that the genetic
components required for the restriction of DNA containing 5mC are different
than those for the restriction of nonglucosylated hydroxymethylcytosine
(hmC)-containing T4 phage DNA.  The possible involvement of the 33-kDa peptide
in the control of the 5mC-specific and hmC-specific activities was examined.
We found evidence for genetic elements downstream of the mcrC gene that
influence the restriction of DNA containing modified cytosine residues.

<>

<1>Beatson, S.A., Das-Gracas-de-Luna, M., Bachmann, N.L., Alikhanm, N.F., Hanksm, K.R., Sullivanm, M.J., Weem, B.A., Freitas-Almeida, A.C., Dos Santos, P.A., de Melo, J.T., Squire, D.J., Cunningham, A.F., Fitzgerald, J.R., Henderson, I.R.
<2>Genome sequence of the emerging pathogen Aeromonas caviae.
<3>J. Bacteriol.
<4>193
<5>1286-1287
<6>2010
<7>Aeromonas caviae is a Gram-negative, motile and rod-shaped facultative anaerobe that is
increasingly being recognised as a cause of diarrhea in  children. Here we present the first
genome sequence of an A. caviae strain  which was isolated as the sole pathogen from a child
with profuse diarrhea.

<>

<1>Beattie, K.L., Wakil, A.E., Driggers, P.H.
<2>Action of restriction endonucleases on transforming DNA of Haemophilus influenzae.
<3>J. Bacteriol.
<4>152
<5>332-337
<6>1982
<7>Cleavage of DNA from Haemophilus influenzae with restriction endonucleases
caused inactivation of transforming ability to an extent that depended on the
genetic marker and the enzyme.  The rate of inactivation, but not the final
level of survival, depended on the concentration of enzyme in the restriction
digest.  In general, the greatest extent of inactivation of transforming
activity was obtained with endonucleases that are known to produce the shortest
fragments.  We electrophoresed restriction digests of H. influenzae DNA in
agarose gels and assayed transforming activity of DNA extracted from gel
slices.  In this way, we determined the lengths of restriction fragments that
contain genetic markers of H. influenzae.  For the marker that we studied most
thoroughly (nov), the shortest restriction fragment that possessed detectable
transforming activity was a 0.9-kilobase pair fragment produced by endonuclease
R - PstI.  The shortest marker-bearing restriction fragment that retained
substantial transforming activity (50% of value for undigested DNA) was a
2.1-kilobase pair EcoRI fragment bearing the kan marker.  Among marker-bearing
restriction fragments 1 to 4 kilobase pairs in length, survival of transforming
activity varied 10,000-fold.  We relate these observations to the recent
finding by Sisco and Smith (Proc. Natl. Acad. Sci. USA 76: 972-976, 1979) that
efficient entry of DNA into competent H. influenzae cells appears to require
the presence of a recognition sequence that is scattered throughout the
Haemophilus genome in many more copies than in unrelated genomes.

<>

<1>Beaty, J.S., Mclean-Bowen, C.A., Brown, L.R.
<2>BvuI: a site-specific endonuclease from Bacillus vulgatis.
<3>Gene
<4>18
<5>61-67
<6>1982
<7>A site-specific endodeoxyribonuclease was partially purified from an extract of
osmotically shocked spheroplasts of a Bacillus vulgatis strain; the enzyme has
been designated BvuI and its activity was characterized.  The locations of
BvuI-generated cleavages on bacteriophage lambda and M13 derivatives mp7, mp8
and mp9, SV40 and pBR322 DNA molecules were determined.  BvuI was shown to
recognize the DNA sequence	5'-G-Pu-G-C-Py-^C-3'	3'-C-^Py-C-G-Pu-G-5'and cleaves
it at the positions indicated by arrows.  Two identical BvuI recognition sites
separated by fourteen nucleotide pairs were shown to occur within the
tetracycline resistance gene of pBR322; BvuI should be used for molecular
cloning in that plasmid vector.

<>

<1>Beaulieu, N., Morin, S., Chute, I.C., Robert, M.-F., Nguyen, H., MacLeod, A.R.
<2>An essential role for DNA methyltransferase DNMT3B in cancer cell survival.
<3>J. Biol. Chem.
<4>277
<5>28176-28181
<6>2002
<7>Abnormal methylation and associated silencing of tumor suppressor genes is a common feature of
many types of cancers. The observation of persistent methylation in human cancer cells lacking
the maintenance methyltransferase DNMT1 suggests the involvement of other DNA
methyltransferases in gene silencing in cancer. To test this hypothesis, we have evaluated
methylation and gene expression in cancer cells specifically depleted of DNMT3A or DNMT3B, de
novo methyltransferases that are expressed in adult tissues. Here we have shown that depletion
of DNMT3B, but not DNMT3A, induced apoptosis of human cancer cells but not normal cells.
DNMT3B depletion reactivated methylation-silenced gene expression but did not induce global or
juxtacentromeric satellite demethylation as did specific depletion of DNMT1. Furthermore, the
effect of DNMT3B depletion was rescued by exogenous expression of either of the splice
variants DNMT3B2 or DNMT3B3 but not DNMT1. These results indicate that DNMT3B has significant
site selectivity that is distinct from DNMT1, regulates aberrant gene silencing, and is
essential for cancer cell survival.

<>

<1>Beck, C., Cranz, S., Solmaz, M., Roth, M., Jeltsch, A.
<2>How does a DNA interacting enzyme change its specificity during molecular evolution? A site-directed mutagenesis study at the DNA binding site of the DNA-(adenine-N(6))-methyltransferase EcoRV.
<3>Biochemistry
<4>40
<5>10956-10965
<6>2001
<7>The EcoRV DNA-(adenine-N6)-methyltransferase (MTase) recognizes GATATC sequences and modifies
the first adenine residue within this site. Parts of its DNA interface show high sequence
homology to DNA MTases of the dam family which recognize and modify GATC sequences. A
phylogenetic analysis of M.EcoRV and dam-MTases suggests that EcoRV arose in evolution from a
primordial dam-MTase in agreement with the finding that M.EcoRV also methylates GATC sites
albeit at a strongly reduced rate. GATCTC sites that deviate in only one position from the
EcoRV sequence are preferred over general dam sites. We have investigated by site-directed
mutagenesis the function of 17 conserved and nonconserved residues within three loops flanking
the DNA binding cleft of M.EcoRV. M.EcoRV contacts the GATATC sequence with two highly
cooperative recognition modules. The contacts to the GAT-part of the recognition sequence are
formed by residues conserved between dam MTases and M.EcoRV. Mutations at these positions lead
to an increase in the discrimination between GATATC and GATC substrates. Our data show that
the change in sequence specificity from dam (GATC) to EcoRV (GATATC) was accompanied by the
generation of a second recognition module that contacts the second half of the target
sequence. The new DNA contacts are formed by residues from all three loops that are not
conserved between M.EcoRV and dam MTases. Mutagenesis at important residues within this module
leads to variants that show a decreased ability to recognize the TC-part of the GATATC
sequence.

<>

<1>Beck, C., Jeltsch, A.
<2>Probing the DNA interface of the EcoRV DNA-(adenine-N6)-methyltransferase by site-directed mutagenesis, fluorescence spectroscopy, and UV cross-linking.
<3>Biochemistry
<4>41
<5>14103-14110
<6>2002
<7>The EcoRV DNA-(adenine-N6)-methyltransferase recognizes GATATC sites and methylates the DNA as
indicated. It is related to the large family of dam methyltransferases which modify GATC
sites. We have studied the interaction of DNA with M.EcoRV and 12 M.EcoRV variants using
oligonucleotides containing 2-aminopurine as a fluorescence probe in equilibrium and
stopped-flow DNA binding studies and 5-iododeoxyuracil for UV cross-linking. M.EcoRV binds to
DNA in a multistep binding reaction, including two different conformations of the specific
enzyme-DNA complex, and induces a strong conformational change of the DNA at the fourth
position of the recognition site. Mutations at residues forming contacts to the GAT part of
the recognition site reduce the stability of both specific enzyme-DNA complexes. Two enzyme
variants which fail to recognize the ATC part do not induce the deformation of the DNA which
explains why they cannot interact properly with the recognition site. Other mutations at
residues which interact with the ATC part selectively reduce the stability of the second
enzyme-DNA complex. These results show that when approaching the DNA M.EcoRV first contacts
the GAT part of the target site. Since the residues mediating these contacts are conserved
among M.EcoRV and dam MTases, the kinetics of formation of the enzyme-DNA complex correspond
to the evolutionary history of the protein. Whether the observation that evolutionarily
conserved contacts are formed early during complex formation is a general rule for DNA
interacting enzymes or proteins that change their specificity during evolution remains to be
seen.

<>

<1>Beck, R., Burtscher, H.
<2>Introduction of arbitrary sequences into genes by use of class IIs restriction enzymes.
<3>Nucleic Acids Res.
<4>22
<5>886-887
<6>1994
<7>Introduction of additional nucleotides or modification of sequences at locations where no
convenient restriction endonuclease recognition site is available can be rather difficult,
especially if one has to conserve a particular reading frame and new cleavage sites cannot
simply be generated by taking advantage of the degeneracy of the genetic code. Overlap
extension can be very useful, however it has some limitations in fragment length and it can
take some time to find optimal conditions for a particular reaction. Using inverse primers to
amplify small plasmids followed by blunt-end ligation also has some restrictions (e.g.
amplification of a longer DNA sequence than necessary; use of polymerases with proof-reading
activity). We wanted to insert a DNA sequence coding for four histidine residues and an
enterokinase cleavage site between the signal sequence and the mature sequence of human
placental alkaline phosphatase for purification. We added the enterokinase recognition site in
order to allow regeneration of the authentic N-terminus of the enzyme. Here we describe a
convenient way to achieve this, using class IIs restriction endonucleases. Class IIs
restriction endonucleases cut DNA several nucleotides away from their recognition site
irrespective of the interventing sequence. This can be exploited to generate arbitrary sticky
ends for in-frame fusion of DNA sequences, combining two PCR reactions with a simple cloning
step.

<>

<1>Becker, M.M., Lesser, D., Kurpiewski, M., Baranger, A., Jen-Jacobson, L.
<2>Ultraviolet footprinting accurately maps sequence-specific contacts and DNA kinking in the EcoRI endonuclease-DNA complex.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>85
<5>6247-6251
<6>1988
<7>The UV footprinting technique has been used to detect contacts between EcoRI
endonuclease and its recognition sequence at single nucleotide resolution.
Comparison of the UV-footprinting results to the published crystal structure of
the EcoRI endonuclease-DNA complex allows us to determine how UV light detects
protein-DNA contacts.  We find that kinking of the DNA helix in the complex
greatly enhances the UV photoreactivity of DNA at the site of the kink.  In
contrast to kinking, contacts between the endonuclease and the DNA bases
inhibit the UV photoreactivity of DNA.  Similar analysis of a proteolytically
modified endonuclease that exhibits the same sequence specificity as wild-type
enzyme but that does not cleave DNA supports these conclusions.  Furthermore,
detection of enhanced photoreactivity at the same kink in the modified
enzyme-DNA complex allows us to conclude that the loss of cleavage activity by
the modified endonuclease is not due to its failure to kink DNA.

<>

<1>Beer, H.D., Maschke, H.E., Schugerl, K.
<2>Continuous production of restriction endonucleases: continuous two-stage cultivation with E. coli JM103; continuous cell disintegration and purification by affinity chromatography.
<3>Appl. Microbiol. Biotechnol.
<4>38
<5>220-225
<6>1992
<7>The optimization of the production of recombinant DNA-derived proteins in Escherichia coli was
investigated. We chose restriction endonucleases EcoRI and EcoRV from E. coli as model
proteins, despite the observation that overproduction can result in a toxic effect to the
cells. The enzymes were expressed as fusion proteins consisting of protein A from
Staphylococcus aureus and the desired enzymes in order to facilitate purification. The
expression of the fusion protein was induced by a temperature shift using the PR promoter of
phage lambda regulated by the repressor plasmid pRK248cI. Data from batch fermentations
provided the basis for planning a continuous two-stage fermentation. The EcoRI enzyme activity
was investigated as a function of the induction time after cell disintegration and allowed an
estimation of yield of the continuous culture. Plasmid instability, which was only observed
under continuous conditions, could be prevented by adding tetracycline (resistance of the
repressor plasmid) to the medium. We established a continuous cell disintegration system and
purified the fusion protein semicontinuously by affinity chromatography. The biological
activity of the fusion protein was the same as the native endonuclease so there was no need
for cleavage of the fusion protein and the product could be used without further processing.

<>

<1>Begley, T.J., Cunningham, R.P.
<2>Methanobacterium thermoformicicum thymine DNA mismatch glycosylase: conversion of an N-glycosylase to an AP lyase.
<3>Protein Eng.
<4>12
<5>333-340
<6>1999
<7>The thymine DNA mismatch glycosylase from Methanobacterium thermoformicicum, a member of the
endonuclease III family of repair proteins, excises the pyrimidine base from T-G and U-G
mismatches. Unlike endonuclease III, it does not cleave the phosphodiester backbone by a
beta-elimination reaction. This cleavage event has been attributed to a nucleophilic attack by
the conserved Lys120 of endonuclease III on the aldehyde group at C1' of the deoxyribose and
subsequent Schiff base formation. The inability of TDG to perform this beta-elimination event
appears to be due to the presence of a tyrosine residue at the position equivalent to Lys120
in endonuclease III. The purpose of this work was to investigate the requirements for AP lyase
activity. We replaced Tyr126 in TDG with a lysine residue to determine if this replacement
would yield an enzyme with an associated AP lyase activity capable of removing a mismatched
pyrimidine. We observed that this replacement abolishes the glycosylase activity of TDG but
does not affect substrate recognition. It does, however, convert the enzyme into an AP lyase.
Chemical trapping assays show that this cleavage proceeds through a Schiff base intermediate
and suggest that the amino acid at position 126 interacts with C1' on the deoxyribose sugar.

<>

<1>Begley, T.J., Haas, B.J., Morales, J.C., Kool, E.T., Cunningham, R.P.
<2>Kinetics and binding of the tymine-DNA mismatch glycosylase, Mig-Mth, with mismatch-containing DNA substrates.
<3>DNA Repair
<4>2
<5>107-120
<6>2003
<7>We have examined the removal of thymine residues from T-G mismatches in DNA by the thymine-DNA
mismatch glycosylase from Methanobacterium thermoautrophicum (Mig-Mth), within the context of
the base excision repair (BER) pathway, to investigate why this glycosylase has such low
activity in vitro. Using single-turnover kinetics and steady-state kinetics, we calculated the
catalytic and product dissociation rate constants for Mig-Mth, and determined that Mig-Mth is
inhibited by product apyrimidinic (AP) sites in DNA. Electrophoretic mobility shift assays
(EMSA) provide evidence that the specificity of product binding is dependent upon the base
opposite the AP site. The binding of Mig-Mth to DNA containing the non-cleavable substrate
analogue difluorotoluene (F) was also analyzed to determine the effect of the opposite base on
Mig-Mth binding specificity for substrate-like duplex DNA. The results of these experiments
support the idea that opposite strand interactions play roles in determining substrate
specificity. Endonuclease IV, which cleaves AP sites in the next step of the BER pathway, was
used to analyze the effect of product removal on the overall rate of thymine hydrolysis by
Mig-Mth. Our results support the hypothesis that endonuclease IV increases the apparent
activity of Mig-Mth significantly under steady-state conditions by preventing reassociation of
enzyme to product.

<>

<1>Behar, A., McCormick, L.J., Perlman, S.J.
<2>Rickettsia felis infection in a common household insect pest, Liposcelis bostrychophila (Psocoptera: Liposcelidae).
<3>Appl. Environ. Microbiol.
<4>76
<5>2280-2285
<6>2010
<7>Many species of Rickettsia are well-known mammalian pathogens transmitted
by blood-feeding arthropods. However, molecular surveys are continually
uncovering novel Rickettsia species, often in unexpected hosts, including
many arthropods that do not feed on blood. This study reports a systematic
molecular characterization of a Rickettsia infecting the psocid Liposcelis
bostrychophila (Psocoptera: Liposcelidae), a common and cosmopolitan
household pest. Surprisingly, the psocid Rickettsia is shown to be
Rickettsia felis, a human pathogen transmitted by fleas that causes
serious morbidity and occasional mortality. The plasmid from the psocid R.
felis was sequenced and was found to be virtually identical to the one in
R. felis from fleas. As Liposcelis insects are often intimately associated
with humans and other vertebrates, it is speculated that they acquired R.
felis from fleas. Whether the R. felis in psocids causes disease in
vertebrates is not known and warrants further study.

<>

<1>Behrens, B., Noyer-Weidner, M., Pawlek, B., Lauster, R., Balganesh, T.S., Trautner, T.A.
<2>Organization of multispecific DNA methyltransferases encoded by temperate Bacillus subtilis phages.
<3>EMBO J.
<4>6
<5>1137-1142
<6>1987
<7>B. subtilis phage Rho11s codes for a multispecific DNA methyltransferase
(Mtase) which methylates cytosine within the sequences GGCC and GAGCTC.  The
Mtase gene of Rho11s was isolated and sequenced.  It has 1509 bp, corresponding
to 503 amino acids (aa).  The enzyme's Mr of 57.2 kd predicted from the
nucleotide sequence was verified by direct Mr determinations of the Mtase.  A
comparison of the aa sequence of the Rho11s Mtase with those of related phages
SPR and Phi3T, which differ in their methylation potential, revealed
generalities in the building plan of such enzymes.  At least 70% of the aa of
each enzyme are contained in two regions of 243 and 109 aa at the N and C
termini respectively, which are highly conserved among the three enzymes.  In
each enzyme, variable sequences separate the conserved regions.  Variability is
generated through the single or multiple use of related and unrelated sequence
motifs.  We propose that the recognition of those DNA target sequences, which
are unique for each of the three enzymes, is determined by these variable
regions.  Evolutionary relationships between the three enzymes are discussed.

<>

<1>Behrens, B., Pawlek, B., Morelli, G., Trautner, T.A.
<2>Restriction and modification in Bacillus subtilis:  construction of hybrid lambda and SPP1 phages containing a DNA methyltransferase gene from B. subtilis phage SPR.
<3>Mol. Gen. Genet.
<4>189
<5>10-16
<6>1983
<7>The gene of B. subtilis phage SPR, specifying DNA methyltransferase activity
was identified within a 3.1 Md EcoRI fragment of SPR DNA.  This fragment was
cloned in E. coli using a lambda insertion vector.  A 1.25 Md PstI fragment
with this gene was subsequently derived from the lambda/SPR hybrid phage and
subcloned in B. subtilis using a new SPP1 vector, whose construction is
described.

<>

<1>Behrens, W., Bonig, T., Suerbaum, S., Josenhans, C.
<2>Genome Sequence of Helicobacter pylori hpEurope Strain N6.
<3>J. Bacteriol.
<4>194
<5>3725-3726
<6>2012
<7>Helicobacter pylori colonizes about half of the world's population. It is a causative agent
of stomach diseases, including malignant tumors. We report the
genome sequence of strain N6, which is widely used in H. pylori research and
appreciated for its large cell size and high transformation efficiency.

<>

<1>Beja, O., Aravind, L., Koonin, E.V., Suzuki, M.T., Hadd, A., Nguyen, L.P., Jovanovich, S.B., Gates, C.M., Feldman, R.A., Spudich, J.L., Spudich, E.N., DeLong, E.F.
<2>Bacterial rhodopsin: evidence for a new type of phototrophy in the sea.
<3>Science
<4>289
<5>1902-1906
<6>2000
<7>Extremely halophilic archaea contain retinal-binding integral membrane
proteins called bacteriorhodopsins that function as light-driven proton
pumps. So far, bacteriorhodopsins capable of generating a chemiosmotic
membrane potential in response to light have been demonstrated only in
halophilic archaea. We describe here a type of rhodopsin derived from
bacteria that was discovered through genomic analyses of naturally
occuring marine bacterioplankton. The bacterial rhodopsin was encoded in
the genome of an uncultivated gamma-proteobacterium and shared highest
amino acid sequence similarity with archaeal rhodopsins. The protein was
functionally expressed in Escherichia coli and bound retinal to form an
active, light-driven proton pump. The new rhodopsin exhibited a
photochemical reaction cycle with intermediates and kinetics
characteristic of archaeal proton-pumping rhodopsins. Our results
demonstrate that archaeal-like rhodopsins are broadly distributed among
different taxa, including members of the domain Bacteria. Our data also
indicate that a previously unsuspected mode of bacterially mediated
light-driven energy generation may commonly occur in oceanic surface
waters worldwide.

<>

<1>Bekker, O.B., Klimina, K.M., Vatlin, A.A., Zakharevich, N.V., Kasianov, A.S., Danilenko, V.N.
<2>Draft Genome Sequence of Streptomyces fradiae ATCC 19609, a Strain Highly Sensitive to Antibiotics.
<3>Genome Announcements
<4>2
<5>e01247-14
<6>2014
<7>We report here a sequence of the genome of the Streptomyces fradiae ATCC 19609 strain,
initially isolated from the soil, which produces tylosin. S. fradiae is
highly sensitive to different classes of antibiotics, compared to the
sensitivities of other bacteria. We have identified 9 groups of genes directly or
indirectly involved in the resistome formation.

<>

<1>Belavin, P.A., Dedkov, V.S., Degtyarev, S.K.
<2>A simple technique for detection of restriction endonucleases in bacterial colonies.
<3>Prikl. Biokhim. Mikrobiol.
<4>24
<5>121-124
<6>1988
<7>A simple technique is proposed for detection of bacterial restriction
endonucleases.  Analysis is performed directly in the cells from colonies
cultivated on Petri dishes.  The cells collected with an inoculation loop are
treated with lysozyme and Triton X-100.  After centrifugation the supernatant
is tested for endonuclease activity.  The technique enables up to 100 colonies
to be tested in 3-4 hr.

<>

<1>Belavin, P.A., Gorbunov, Y.A., Malygin, E.G., Rechkunova, N.I., Degtyarev, S.K.
<2>Substrate specificity of DNA methylase from Flavobacterium okeanokoites.
<3>Bioorg. Khim.
<4>15
<5>417-418
<6>1989
<7>The specificity of DNA methylase M.FokI towards oligonucleotides containing the
sequence 5' GGATG was investigated, and N6-methyladenine in the GGATG chain was
shown to be the only product of the modification.

<>

<1>Beletskaya, I.V., Zakharova, M.V., Shlyapnikov, M.G., Semenova, L.M., Solonin, A.S.
<2>DNA methylation at the CfrBI site is involved in expression control in the CfrBI restriction-modification system.
<3>Nucleic Acids Res.
<4>28
<5>3817-3822
<6>2000
<7>We have previously found that genes of the CfrBI restriction-modification (R-M) system from
Citrobacter freundii are oriented divergently and that their promoter regions overlap. The
overlapping promoters suggest regulation of gene expression at the transcriptional level. In
this study the transcription regulation of CfrBI R-M genes was analyzed in vivo and in vitro
in Escherichia coli. It was shown that in the presence of CfrBI methyltransferase (M.CfrBI),
cell galactokinase activity decreases 10-fold when the galactokinase gene (galK) is under the
control of the cfrBIM promoter and increases 20-fold when galK is under the control of the
cfrBIR promoter. The CfrBI site, proven to be unique for the entire CrfBI R-M gene sequence,
is located in the -35 cfrBIM promoter region and is in close vicinity of the -10 cfrBIR
promoter region. A comparison of the cfrBIM and the cfrBIR promoter activities in the in vitro
transcription system using methylated and unmethylated DNA fragments as templates demonstrated
that the efficiency of CfrBI R-M gene transcription is regulated by enzymatic modification at
the N-4-position of cytosine bases of the CfrBI site by M.CfrBI.  From the results of the in
vivo and in vitro experiments we suggest a new model of gene expression regulation in type II
R-M systems.

<>

<1>Beletskii, A., Bhagwat, A.S.
<2>Transcription-induced mutations: increase in C to T mutations in the nontranscribed strand during transcription in Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>13919-13924
<6>1996
<7>Cytosines in single-stranded DNA deaminate to uracils at 140 times the rate for cytosines in
double-stranded DNA.  If resulting uracils are not replaced with cytosine, C to T mutations
occur.  These facts suggest that cellular processes such as transcription that create
single-stranded DNA should promote C to T mutations.  We tested this hypothesis with the
Escherichia coli tac promoter and found that induction of transcription causes ~4-fold
increase in the frequency of C to U or 5-methylcytosine to T deaminations in the
nontranscribed strand.  Excess mutations caused by C to U deaminations were reduced, but not
eliminated, by uracil-DNA glycosylase.  Similarly, mutations caused by 5-methylcytosine to T
deaminations were only partially reduced by the very short-patch repair process in E. coli.
These effects are unlikely to be caused by differential repair of the two strands, and our
results suggest that all actively transcribed genes in E. coli should acquire more C to T
mutations in the nontranscribed strand.

<>

<1>Belfort, M.
<2>Two for the price of one: a bifunctional intron-encoded DNA endonuclease-RNA maturase.
<3>Genes Dev.
<4>17
<5>2860-2863
<6>2003
<7>Homing endonucleases are enzymes that are typically encoded by intervening sequences of
various kinds -- introns that splice at the RNA level, or inteins that splice at the protein
level.  The endonucleases function primarily to mobilize these elements, which are often
self-splicing, by facilitating their integration to new sites in DNA.  Because these sites are
most often allelic intronless or inteinless sites, this form of mobility is termed homing, and
the target sequences are termed homing sites.  Homing endonucleases sometimes do double duty,
also functioning as maturases, which promote RNA splicing.  Maturases are usually intron
specific, and act as cofactors that bind their intron-containing precursor RNA to facilitate
splicing.  Nevertheless, catalysis required for splicing remains a property of the RNA.

<>

<1>Belfort, M.
<2>Back to Basics: Structure, function, evolution and application of homing endonucleases and inteins.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Berlin Heidelberg
<4>16
<5>1-10
<6>2005
<7>"It is a profound and necessary truth that the deep things in science are not found because
they are useful; they are found because it was possible to find them." (Robert Oppenheimer,
1904-1967).  Oppenheimer's words resonate with this book's theme, which is how both applied
and theoretical science can emanate from answers to basic questions - whether they are being
asked to model organisms or in test tubes - about molecular structure and mechanism.  Thus,
fundamental work on DNA, RNA and proteins, which encode or constitute homing endonucleases and
inteins, is leading to refined theories of evolution in prokaryotes and eukaryotes on the one
hand, and the development of laboratory tools and health-care reagents on the other.  Homing
endonucleases and inteins, sometimes referred to as "protein introns", are linked at many
levels.  First, homing endonucleases are frequently encoded by introns that self-splice at the
RNA level, in analogy to inteins that self-splice at the protein level. Second, homing
endonucleases similar to those encoded by introns are often found embedded within and
co-translated with inteins.  Third, both types of intervening sequence are mobile elements,
capable of movement from genome to genome.  Fourth, the endonuclease component of both introns
and inteins imparts their mobility.  Fifth, each of these mobile intervening sequences is
thought to have originated from invasion of the gene encoding the self-splicing element, the
intron or intein, by an endonuclease gene, the primordial mobile element.  The final unifying
theme is the exploitation of introns, inteins and homing endonucleases by chemists,
geneticists, structural biologists and engineers, to generate reagents and tools that are
useful in basic research, biotechnology and medicine. This chapter serves as an introduction
to the volume entitled Homing Endonucleases and Inteins, which provides a wonderful
illustration of the point that fundamental studies of structure and mechanism fuel
evolutionary theory and technology development alike.

<>

<1>Belfort, M., Bonocora, R.P.
<2>Homing Endonucleases: From Genetic Anomalies to Programmable Genomic Clippers.
<3>Methods Mol. Biol.
<4>1123
<5>1-26
<6>2014
<7>Homing endonucleases are strong drivers of genetic exchange and horizontal transfer of both
their own genes and their local genetic environment. The mechanisms that govern the function
and evolution of these genetic oddities have been well documented over the past few decades at
the genetic, biochemical, and structural levels. This wealth of information has led to the
manipulation and reprogramming of the endonucleases and to their exploitation in genome
editing for use as therapeutic agents, for insect vector control and in agriculture. In this
chapter we summarize the molecular properties of homing endonucleases and discuss their
strengths and weaknesses in genome editing as compared to other site-specific nucleases such
as zinc finger endonucleases, TALEN, and CRISPR-derived endonucleases.

<>

<1>Belfort, M., Edgell, D., Shub, D., Van Roey, P., Derbyshire, V.
<2>A multicomponent, multifunctional intron endonuclease, I-TevI.
<3>J. Biomol. Struct. Dyn.
<4>20
<5>834-835
<6>2003
<7>I-TevI is a homing endonuclease that is encoded by and promotes the mobility of the td intron
of phage T4.  This 28-kD protein is remarkable in that it is site specific, yet recognizes a
target site of ~37 bp in a sequence-tolerant fashion.  The enzyme consists of two functionally
distinct domains, an N-terminal catalytic domain connected by a 75-amino acid linker.  What
has previously been defined as the DNA-binding domain has an extended structure consisting of
three distinct modules which contact ~20 bp of the homing site: A Zn finger, an alpha-helix
and a small helix-turn-helix subdomain.  It is now known that the zinc finger is not required
for DNA binding or catalysis, but rather is a spring-like component of the linker that directs
the catalytic domain to cleave the homing site at a fixed distance from the intron insertion
site.  In contrast, the catalytic domain, which contains the conserved GIY-YIG motif
characteristic of this family of homing endonucleases, is a compactly folded structure.  It is
provocative that there are similarities in the three-dimensional arrangement of the
catalytically important residues and the cation-binding site with those of I-PpoI, a member of
this His-Cys-box family of homing endonucleases.  These results suggest the possibility of
mechanistic relationships among these different families of enzymes, but whether this
represents convergent or divergent evolution is not known.  Regardless, the picture that has
built is one of a catalytic GIY-YIG cartridge that has become associated with modules that
direct DNA binding and distance measurement.  Surprisingly, the DNA-binding domain has
recently been shown to interact with regulatory sequences upstream of the I-TevI gene, such
that I-TevI acts as a repressor of its own synthesis.  Thus, I-TevI is a multicomponent,
multifunctional molecule that has evolved site-specific cleavage and autorepression functions
to maximize the invasiveness and persistence of its host intron.

<>

<1>Belfort, M., Perlman, P.S.
<2>Mechanisms of intron mobility.
<3>J. Biol. Chem.
<4>270
<5>30237-30240
<6>1995
<7>Group I and group II introns, which splice via RNA-catalyzed pathways, can invade DNA
sequences by virtue of proteins expressed from open reading frames (ORFs) contained within
them.  These intron products are endonucleases in the case of group I introns and reverse
transcriptases (RTs) with associated endonuclease activity in the case of the group II
introns.  Two kinds of mobility reactions will be considered for each intron type: homing into
cognate intronless alleles and transposition to non-allelic sites.

<>

<1>Belfort, M., Reaban, M.E., Coetzee, T., Dalgaard, J.Z.
<2>Prokaryotic introns and inteins: a panoply of form and function.
<3>J. Bacteriol.
<4>177
<5>3897-3903
<6>1995
<7>Following their initial discovery, introns and RNA splicing were considered, along with the
nuclear envelope, as characteristics that distinguish eukaryotes from prokaryotes (the
eubacteria, referred to simply as bacteria, and the archaebacteria, referred to as archaea).
This dogma became shaky with the identification of putative introns in tRNA genes of archaea
and finally crumbled with the discovery of self-splicing group I introns in the phages of
purple bacteria, which was followed by the discovery of group I and group II introns in
bacterial cells. Another breakthrough was the recent discovery of a nuclear pre-mRNA-like
intron in a bacterial plasmid. The variety of intervening sequences (IVSs) that span the
phylogenetic spectrum was further extended by the discovery of inteins, elements that can
splice at the protein level, in eukaryotes, archaea, and bacteria. This minireview will
illustrate that interrupted genes can no longer be considered the province of eukaryotes, but
rather than many forms of IVSs are phylogenetically diverse, occurring also in bacterial and
archaeal genomes.

<>

<1>Belfort, M., Roberts, R.J.
<2>Homing endonucleases: keeping the house in order.
<3>Nucleic Acids Res.
<4>25
<5>3379-3388
<6>1997
<7>Homing endonucleases are rare-cutting enzymes encoded by introns and inteins.  They have
striking structural and functional properties that distinguish them from restriction enzymes.
Nomenclature conventions analogous to those for restriction enzymes have been developed for
the homing endonucleases.  Recent progress in understanding the structure and function of the
four families of homing enzymes is reviewed.  Of particular interest are the first reported
structures of homing endonucleases of the LAGLIDADG family.  The exploitation of the homing
enzymes in genome analysis and recombination research is also summarized.  Finally, the
evolution of homing endonucleases is considered, both at the structure-function level and in
terms of their persistence in widely divergent biological systems.

<>

<1>Belfort, M., Van Roey, P., Derbyshire, V.
<2>Modular assembly of a multi-functional homing endonuclease.
<3>J. Biomol. Struct. Dyn.
<4>22
<5>809
<6>2005
<7>Homing endonucleases are usually encoded within self-splicing introns or inteins, and are
found in all three biological kingdoms.  The endonuclease genes and their host intron or
intein are composite selfish genetic elements that spread between related genomes by a process
called homing.  The homing pathway is initiated by the endonuclease binding to a sequence, the
homing site, centered on the intron insertion site of intronless alleles.  The role of the
homing endonuclease is limited to introduction of a double-strand break close to the intron
IS.  I-TevI, a member of the GIY-YIG family of homing endonucleases, is a site-specific yet
sequence tolerant enzyme.  It is remarkably extended molecule consisting of a well-folded
catalytic domain connected to a series of sub-domains by extended regions.  These include a
Zn-finger, a minor-goove binding helix and a helix-turn-helix subdomain.  The modular nature
of the enzyme is pertinent to the evolution of homing endonucleases.  We have begun to assess
the role of each subdomain in endonuclease function.  Most interestingly, we have found that
the Zn-finger subdomain does not play a role in substrate DNA binding.  Rather it is required
for positioning the catalytic domain at a set distance from the primary binding site for
cleavage to occur.  The next challenge is trying to understand the nature of the linker that
connects the catalytic and DNA-binding domains and how it interacts with the Zn-finger to
mediate distance determination.  In addition, recent work has shown that the enzyme is
bi-functional and can act as an auto-repressor.  It can bind to a site present in its promoter
region that exhibits sequence similarity to its homing site, but does not cleave the DNA.  It
is the very modular nature of the enzyme, with distinct sequence requirements for binding and
cleavage, that facilitates its bifunctionality by binding in the absence of cleavage, leading
to transcriptional repression.

<>

<1>Belinsky, S.A., Nikula, K.J., Baylin, S.B., Issa, J.-P.
<2>A microassay for measuring cytosine DNA methyltransferase activity during tumor progression.
<3>Toxicol. Lett.
<4>82
<5>335-340
<6>1995
<7>The cytosine DNA methyltransferase (MT) enzyme, which catalyzes DNA
methylation at CpG sites, is overexpressed at the mRNA level during the progressive stages of
colon cancer.  This paper describes the adaptation of a sensitive microassay for determining
MT
enzyme activity during tumor progression in human colon and murine lung.  MT activity was
progressively elevated in mucosa from familial adenomatosis polyposis patients, mucosa
adjacent
to cancers, and in colonic adenocarcinomas when compared to colonic mucosa from control
patients.  In addition, the activity of this enzyme was increased in alveolar type II but not
Clara
cells isolated from A/J mice following carcinogen exposure and continued to increase during
tumor
progression.  The use of a microassay for measuring MT activity indicates that changes in
enzyme
activity were in general agreement with previous findings of increased MT mRNA levels during
colon cancer progression and also implicates the involvement of this pathway in lung cancer
development.

<>

<1>Belinsky, S.A., Nikula, K.J., Baylin, S.B., Issa, J.-P.J.
<2>Increased cytosine DNA-methyltransferase activity is target-cell-specific and an early event in lung cancer.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>4045-4050
<6>1996
<7>The association between increased DNA-methyltransferase (DNA-MTase) activity
and tumor development suggests a fundamental role for this enzyme in the initiation and
progression of cancer.  A true functional role for DNA-MTase in the neoplastic process would
be
further substantiated if the target cells affected by the initiating carcinogen exhibit
changes in
enzyme activity.  This hypothesis was addressed by examining DNA-MTase activity in alveolar
type II (target) and Clara (nontarget) cells from A/J and C3H mice that exhibit high and low
susceptibility, respectively, for lung tumor formation.  Increased DNA-MTase activity was
found
only in the target alveolar type II cells of the susceptible A.J mouse and caused a marked
increase
in overall DNA methylation in these cells.  Both DNA-MTase and DNA methylation changes were
detected 7 days after carcinogen exposure and, thus, were early events in neoplastic
evolution.
Increased gene expression was also detected by RNA in situ hybridization in hypertrophic
alveolar
type II cells of carcinogen-treated A/J mice, indicating that elevated levels of expression
may be a
biomarker for premalignancy.  Enzyme activity increased incrementally during lung cancer
progression and coincided with increased expression of the DNA-MTase gene in hyperplasias,
adenomas, and carcinomas.  Thus, these results indicate that early increases in DNA-MTase
activity are strongly associated with neoplastic development and constitute a key step in
carcinogenesis.  The detection of premalignant lung disease through increased DNA-MTase
expression and the possibility of blocking the deleterious effects of this change with
specific
inhibitors will offer new intervention strategies for lung cancer.

<>

<1>Belkebir, A., Azeddoug, H.
<2>Purification and characterization of SepII a new restriction endonuclease from Staphylococcus epidermidis.
<3>Microbiol. Res.
<4>167
<5>90-94
<6>2012
<7>A Type II restriction enzyme SepII has been purified to apparent homogeneity from the
gram-positive coccus,Staphylococcus epidermidis.
The purification included an ammonium sulfate precipitation followed by
Q-sepharose, heparin-sepharose and MonoQ column chromatography on an
FPLC system. SDS-PAGE analysis showed a denatured molecular weight of
29 kDa. The effects of temperature, pH, NaCl, Mn2+,Ca2+, and Mg2+ ion
concentrations were studied to determine the optimal reaction
conditions. The enzyme exhibits near maximal levels of activity between
pH 8-10, at 10-20 mM MgCl2, 100-150 mM NaCl and 1 mM DTT. The results
also show that in NEB Buffer 3 the enzyme is active over a broad
temperature range from 0 to 70 degrees C, and in the absence of DNA,
enzyme thermostability is observed up to 50 degrees C for 20 min, while
most of the original activity is conserved in 50% glycerol for weeks at
room temperature. Single and double digestion in presence of commercial
restriction enzymes of known DNA substrates (lambda, pBR322, pET21,
pTrcHisB, pPB67) showed that the purified SepII recognized and cleaved
the same site as EcoRV. Genomic DNA modification status was also
determined.

<>

<1>Belkebir, A., Azeddoug, H.
<2>Metal ion dependence of DNA cleavage by SepMI and EhoI restriction endonucleases.
<3>Microbiol. Res.
<4>168
<5>99-105
<6>2013
<7>Most of type II restriction endonucleases show an absolute requirement for divalent metal ions
as cofactors for DNA cleavage. While Mg2+ is
the natural cofactor other metal ions can substitute it and mediate the
catalysis, however Ca2+ (alone) only supports DNA binding. To
investigate the role of Mg2+ in DNA cleavage by restriction
endonucleases, we have studied the Mg2+ and Mn2+ concentration
dependence of DNA cleavage by SepMI and EhoI. Digestion reactions were
carried out at different Mg2+ and Mn2+ concentrations at constant ionic
strength. These enzymes showed different behavior regarding the ions
requirement, SepMI reached near maximal level of activity between 10
and 20 mM while no activity was detected in the presence of Mn2+ and in
the presence of Ca2+ cleavage activity was significantly decreased.
However, EhoI was more highly active in the presence of Mn2+ than in
the presence of Mg2+ and can be activated by Ca2+. Our results propose
the two-metal ion mechanism for EhoI and the one-metal ion mechanism
for SepMI restriction endonuclease. The analysis of the kinetic
parameters under steady state conditions showed that SepMI had a K-m
value for pTrcHisB DNA of 6.15 nM and a V-max of 1.79 x 10(-2) nM
min(-1), while EhoI had a K-m for pUC19 plasmid of 8.66 nM and a V-max
of 2 x 10(-2) nM min(-1). (C) 2012 Elsevier GmbH. All rights reserved.

<>

<1>Bell, D.C., Cupples, C.G.
<2>Very-Short-Patch Repair in Escherichia coli Requires the dam Adenine Methylase.
<3>J. Bacteriol.
<4>183
<5>3631-3635
<6>2001
<7>Strains of Escherichia coli which lack the dam-encoded adenine methylase are mutators due to a
reduction in the efficiency of postreplication mismatch repair. In this study, we show that
Dam(-) strains are also defective in very-short-patch repair, the system which corrects T/G
mismatches arising from the deamination of 5-methylcytosine. This defect is associated with
decreased levels of Vsr, the endonuclease which initiates short-patch repair. We also show
that production of the dcm-encoded cytosine methylase is unaffected in Dam(-) strains. Since
the dcm and vsr genes are cotranscribed, the regulation of Vsr by Dam is probably
posttranscriptional.

<>

<1>Bell, K.S. et al.
<2>Genome sequence of the enterobacterial phytopathogen Erwinia carotovora subsp. atroseptica and characterization of virulence factors.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>11105-11110
<6>2004
<7>The bacterial family Enterobacteriaceae is notable for its well studied human pathogens,
including Salmonella, Yersinia, Shigella, and Escherichia spp. However, it also contains
several plant pathogens. We report the genome sequence of a plant pathogenic enterobacterium,
Erwinia carotovora subsp. atroseptica (Eca) strain SCRI1043, the causative agent of soft rot
and blackleg potato diseases. Approximately 33% of Eca genes are not shared with sequenced
enterobacterial human pathogens, including some predicted to facilitate unexpected metabolic
traits, such as nitrogen fixation and opine catabolism. This proportion of genes also contains
an overrepresentation of pathogenicity determinants, including possible horizontally acquired
gene clusters for putative type IV secretion and polyketide phytotoxin synthesis. To
investigate whether these gene clusters play a role in the disease process, an arrayed set of
insertional mutants was generated, and mutations were identified. Plant bioassays showed that
these mutants were significantly reduced in virulence, demonstrating both the presence of
novel pathogenicity determinants in Eca, and the impact of functional genomics in expanding
our understanding of phytopathogenicity in the Enterobacteriaceae.

<>

<1>Bell-Pedersen, D., Quirk, S., Clyman, J., Belfort, M.
<2>Intron mobility in phage T4 is dependent upon a distinctive class of endonucleases and independent of DNA sequences encoding the intron core: mechanistic and evolutionary implications.
<3>Nucleic Acids Res.
<4>18
<5>3763-3770
<6>1990
<7>Although mobility of the phylogenetically widespread group I introns appears to be
mechanistically similar, the phage T4 intron-encoded endonucleases that promote mobility of
the td and sunY introns are different from their eukaryotic counterparts. Most notably, they
cleave at a distance from the intron insertion sites. The td enzyme was shown to cleave 23-26
nt 5' and the sunY endonuclease 13-15 nt 3' to the intron insertion site to generate 3-nt or
2-nt 3'-OH extensions, respectively. The absolute coconversion of exon markers between the
distant cleavage and insertion sites is consistent with the double-strand-break repair model
for intron mobility. As a further critical test of the model we have demonstrated that the
mobility event is independent of DNA sequences that encode the catalytic intron core
structure. Thus, in derivatives in which the lacZ or kanR coding sequences replace the intron,
these marker genes are efficiently inserted into intron-minus alleles when the cognate
endonuclease is provided in trans. The process is therefore endonuclease-dependent, rather
than dependent on the intron per se. These findings, which imply that the endonucleases rather
than the introns themselves were the primordial mobile elements, are incorporated into a model
for the evolution of mobile introns.

<>

<1>Bell-Pedersen, D., Quirk, S.M., Aubrey, M., Belfort, M.
<2>A site-secific endonuclease and co-conversion of flanking exons associated with the mobile td intron of phage T4.
<3>Gene
<4>82
<5>119-126
<6>1989
<7>The product of the td intron open reading frame (ORF) of phage T4 is required for
high-frequency transfer of the intervening sequence from intron-plus (In+) to intron-minus
(In-) alleles. In vivo studies have demonstrated that the td ORF product targets cleavage of
td In- DNA, and that cleavage is correlated with intron inheritance [Quirk et al., Cell 56
(1989) 455-465]. In the present study we show by in vitro synthesis of the td intron ORF
product, that the protein possesses endonuclease activity and efficiently cleaves
double-stranded DNA at or near the site of intron integration. In addition, we demonstrate
that intron insertion is accompanied by co-conversion of the flanking exon sequences.
Co-conversion of markers within 50 nt surrounding the site of intron insertion occurred at a
high frequency (80-100%), and decreased at greater distance from the intervening sequence.
Co-conversion may provide a mechanism for maintaining exon-intron RNA contacts required for
accurate splicing of the relocated intron. Cleavage of target DNA by an intron endonuclease
and co-conversion of flanking exon sequences are both features associated with mobile introns
of eukaryotes, indicating a common mechanism for intron transfer in the eukaryotic kingdoms.

<>

<1>Bell-Pedersen, D., Quirk, S.M., Bryk, M., Belfort, M.
<2>I-TevI, the endonuclease encoded by the mobile td intron, recognizes binding and cleavage domains on its DNA target.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>88
<5>7719-7723
<6>1991
<7>Mobility of the phage T4 td intron depends on activity of an intron-encoded endonuclease
(I-TevI), which cleaves a homologous intronless (delta-In) target gene. The double-strand
break initiates a recombination event that leads to intron transfer. We found previously that
I-TevI cleaves td-delta-IN target DNA 23-26 nucleotides upstream of the intron insertion site.
DNase I-footprinting experiments and gel-shift assays indicate that I-TevI makes primary
contacts around the intron insertion site. A synthetic DNA duplex spanning the insertion site
but lacking the cleavage site was shown to bind I-TevI specifically, and when cloned, to
direct cleavage into vector sequences. The behavior of the cloned duplex and that of deletion
and insertion mutants support a primary role for sequences surrounding the insertion site in
directing I-TevI binding, conferring cleavage ability, and determining cleavage polarity. On
the other hand, sequences around the cleavage site were shown to influence cleavage efficiency
and cut-site selection. The role of cleavage-site sequences in determining cleavage distance
argues against a strict "ruler" mechanism for cleavage by I-TevI. The complex nature of the
homing site recognized by this unusual type of endonuclease is considered in the context of
intron spread.

<>

<1>Bellaiche, Y., Mogila, V., Perrimon, N.
<2>I-SceI endonuclease, a new tool for studying DNA double-strand break repair mechanisms in Drosophila.
<3>Genetics
<4>152
<5>1037-1044
<6>1999
<7>As a step toward the development of a homologous recombination system in Drosophila, we have
developed a methodology to target double-strand breaks (DSBs) to a specific position in the
Drosophila genome. This method uses the mitochondrial endonuclease I-SceI that recognizes and
cuts an 18-bp restriction site. We find that >6% of the progeny derived from males that carry
a marker gene bordered by two I-SceI sites and that express I-SceI in their germ line lose the
marker gene. Southern blot analysis and sequencing of the regions surrounding the I-SceI sites
revealed that in the majority of the cases, the introduction of DSBs at the I-SceI sites
resulted in the complete deletion of the marker gene; the other events were associated with
partial deletion of the marker gene. We discuss a number of applications for this novel
technique, in particular its use to study DSB repair mechanisms.

<>

<1>Bellamy, S.R., Kovacheva, Y.S., Zulkipli, I.H., Halford, S.E.
<2>Differences between Ca2+ and Mg2+ in DNA binding and release by the SfiI restriction endonuclease: implications for DNA looping.
<3>Nucleic Acids Res.
<4>37
<5>5443-5453
<6>2009
<7>Many enzymes acting on DNA require Mg(2+) ions not only for catalysis but also to bind DNA.
Binding studies often employ Ca(2+) as a substitute for
Mg(2+), to promote DNA binding whilst disallowing catalysis. The SfiI
endonuclease requires divalent metal ions to bind DNA but, in contrast to
many systems where Ca(2+) mimics Mg(2+), Ca(2+) causes SfiI to bind DNA
almost irreversibly. Equilibrium binding by wild-type SfiI cannot be
conducted with Mg(2+) present as the DNA is cleaved so, to study the
effect of Mg(2+) on DNA binding, two catalytically-inactive mutants were
constructed. The mutants bound DNA in the presence of either Ca(2+) or
Mg(2+) but, unlike wild-type SfiI with Ca(2+), the binding was reversible.
With both mutants, dissociation was slow with Ca(2+) but was in one case
much faster with Mg(2+). Hence, Ca(2+) can affect DNA binding differently
from Mg(2+). Moreover, SfiI is an archetypal system for DNA looping; on
DNA with two recognition sites, it binds to both sites and loops out the
intervening DNA. While the dynamics of looping cannot be measured with
wild-type SfiI and Ca(2+), it becomes accessible with the mutant and
Mg(2+).

<>

<1>Bellamy, S.R., Milsom, S.E., Kovacheva, Y.S., Sessions, R.B., Halford, S.E.
<2>A Switch in the Mechanism of Communication between the Two DNA-Binding Sites in the SfiI Restriction Endonuclease.
<3>J. Mol. Biol.
<4>373
<5>1169-1183
<6>2007
<7>While many Type II restriction enzymes are dimers with a single DNA-binding cleft between the
subunits, SfiI is a tetramer of identical
subunits. Two of its subunits (a dimeric unit) create one DNA-binding
cleft, and the other two create a second cleft on the opposite side of the
protein. The two clefts bind specific DNA cooperatively to give a complex
of SfiI with two recognition sites. This complex is responsible for
essentially all of the DNA-cleavage reactions by SfiI: virtually none is
due to the complex with one site. The communication between the
DNA-binding clefts was examined by disrupting one of the very few polar
interactions in the otherwise hydrophobic interface between the dimeric
units: a tyrosine hydroxyl was removed by mutation to phenylalanine. The
mutant protein remained tetrameric in solution and could bind two DNA
sites. But instead of being activated by binding two sites, like wild-type
SfiI, it showed maximal activity when bound to a single site and had a
lower activity when bound to two sites. This interaction across the dimer
interface thus enforces in wild-type SfiI a cooperative transition between
inactive and active states in both dimers, but without this interaction as
in the mutant protein, a single dimer can undergo the transition to give a
stable intermediate with one inactive dimer and one active dimer.

<>

<1>Bellamy, S.R., Mina, P., Retter, S.E., Halford, S.E.
<2>Fidelity of DNA Sequence Recognition by the SfiI Restriction Endonuclease Is Determined by Communications between Its Two DNA-Binding Sites.
<3>J. Mol. Biol.
<4>384
<5>557-563
<6>2008
<7>The SfiI restriction endonuclease is a tetramer in which two subunits form a dimeric unit that
contains one DNA binding cleft and the other two
subunits contain a second cleft on the opposite side of the protein. Full
activity requires both clefts to be filled with its recognition sequence:
SfiI has low activity when bound to one site. The ability of SfiI to
cleave non-cognate sites, one base pair different from the true site, was
initially tested on substrates that lacked specific sites but which
contained either one or multiple non-cognate sites. No cleavage of the DNA
with one non-cognate site was detected, while a small fraction of the DNA
with multiple sites was nicked. The alternative sequences were, however,
cleaved in both strands, albeit at low levels, when the DNA also carried
either a recognition site for SfiI or the termini generated by SfiI.
Further tests employed a mutant of SfiI, altered at the dimer interface,
which was known to be more active than wild-type SfiI when bound to a
single site. This mutant similarly failed to cleave DNA with one
non-cognate site, but cleaved the substrates with multiple non-cognate
sites more readily than did the native enzyme. To cleave additional sites,
SfiI thus needs to interact concurrently with either two non-cognate sites
or one non-cognate and one cognate site (or the termini thereof), yet this
arrangement is still restrained from cleaving the alternative site unless
the communication pathway between the two DNA-binding clefts is disrupted.

<>

<1>Bellamy, S.R.W., Milsom, S.E., Scott, D.J., Daniels, L.E., Wilson, G.G., Halford, S.E.
<2>Cleavage of individual DNA strands by the different subunits of the heterodimeric restriction endonuclease BbvCI.
<3>J. Mol. Biol.
<4>348
<5>641-653
<6>2005
<7>BbvCI cleaves an asymmetric DNA sequence, 5'-CC^TCAGC-3'/5'-GC^TGAGG-3', as indicated.
While many Type II
restriction enzymes consist of identical subunits, BbvCI has two
different subunits: R-1, which acts at GC^TGAGG; and R-2,
which acts at CC^TCAGC. Some mutants of BbvCI with defects
in one subunit either R1-R2+ or R1+R2-, cleave only one strand, that
attacked by the native subunit. In analytical ultracentrifugation at
various concentrations of protein, wild-type and mutant BbvCI enzymes
aggregated extensively, but are R1R2 heterodimers at the concentrations
used in DNA cleavage reactions. On a plasmid with one recognition site,
wild-type BbvCI cleaved both strands before dissociating from the DNA,
while the R1-R2+ and R1+R2- mutants acted almost exclusively on their
specified strands, albeit at relatively slow rates. During the
wild-type reaction, the DNA is cleaved initially in one strand, mainly
that targeted by the R, subunit. The other strand is then cleaved
slowly by R-2 before the enzyme dissociates from the DNA. Hence, the
nicked form accumulates as a transient intermediate. This behaviour
differs from that of many other restriction enzymes, which cut both
strands at equal rates. However, the activities of the R-1(+) and
R-2(+) subunits in the wild-type enzyme can differ from their
activities in the R1+R2- and R1-R2+ mutants. Each active site in BbvCI
therefore influences the other.

<>

<1>Belland, R.J., Morrison, S.G., Hogan, D.
<2>A phase-variable type III restriction-modification system in Neisseria gonorrhoeae.
<3>Abs. Tenth Int. Conf. Pathogenegic Neisseria, , Zollinger, W.D., Frasch, C.E., Deal, C.D., Baltimore, MD
<4>0
<5>360-361
<6>1996
<7>Neisseria gonorrhoeae possesses a number of type II restriction/modification systems but
characteristically expresses fewer restriction enzyme activities than corresponding
methyltransferases.  Methyltransferase specificities have been identified for S.NgoI
(PuGCGCPy), S.NgoII (GGCC), S.NgoIII (CCGCGG), S.NgoIV (GCCGGC), S.NgoV (GGNNCC), S.NgoVI
(GATC), S.NgoVII (GC[C/G]GC), S.NgoVIII (TCACC) of purified methyltransferase which recognizes
the sequence GTAN5CTC (S.NgoIX).  Restriction enzymes have been detected which correspond to
the sequences for NgoI, II, III, IV, and IX in various strains.  We have identified the genes
encoding a type III restriction modification system present in all strains of N. gonorrhoeae
tested.  Sequence analysis of the ngoX locus from strain MS11 indicate the predicted protein
for the restriction enzyme (NgoX.R) is 59% identical to the restriction enzyme of the P1
system.  The modification enzyme (NgoX.M) is predicted to be 38% identical to the modification
enzyme of the P1 system. The postulated 5' region of the ngoX.M gene is unusual in that the
predicted start codon is followed by a series of direct pentameric repeats reminiscent of the
signal-peptide encoding region of neisserial opa genes, responsible for regulating expression
of opa genes by "phase variation".  The ngoX.M gene of strain MS11 contains eight repeat
elements (an out-of-frame configuration) while strain FA228 contains twelve repeat elements
(an in-frame configuration).  Mutations constructed in Escherichia coli and returned to the
gonococcal chromosome by allelic replacement indicate that the ngoX.M gene is expressed in
strain FA228 but not in strain MS11 but the gene is phase-variable from both the on and off
configurations.  Phase variation cannot be detected in the viable cell population in wild-type
strains.  Purified heterodimeric NgoX enzyme is active against DNA purified from strains MS11
mk (no expression of NgoX.M) and FA228 ngoX.M::lacZ but not strain FA228 (expresses NgoX.M).

<>

<1>Bellemare, G., Potvin, C.
<2>Classification of type-II restriction endonucleases and cloning of non-identical cohesive-end fragments without self-polymerization using nonpalindromic oligodeoxyribonucleotide adapters.
<3>Gene
<4>101
<5>67-74
<6>1991
<7>Enzymatic partial filling-in of recessed 3'-end sequences, left after digestion
of DNA by the restriction endonucleases (ENases) Sau3AI and SalI, with the
Klenow fragment of E. coli DNA polymerase I allows the forced ligation of the
resulting fragments; this technology is already used for subcloning and for
genomic bank construction.  To simplify and generalize its utilization,
class-II ENases have been arranged into 16 different families according to the
composition of the 5'-protruding sequences present after cleavage.  Moreover,
this system was extended to allow the joining of noncompatible ends by the use
of nonpalindromic complementary oligodeoxyribonucleotides (NPCOs) containing
two nucleotides protruding at each 5' end.  The use of these synthetic adapters
maintains all the advantages of the initial gap-filling cloning technique: only
one insert can be cloned per vector molecule and no self-ligation or
-polymerization can occur with any of the DNA molecules involved.  Only 22 such
oligodeoxyribonucleotides are needed to generate the 60 NPCO pairs necessary to
ligate to each other any member of twelve ENase families when the regeneration
of ENase recognition sites is not required.

<>

<1>Beller, H.R., Chain, P.S., Letain, T.E., Chakicherla, A., Larimer, F.W., Richardson, P.M., Coleman, M.A., Wood, A.P., Kelly, D.P.
<2>The genome sequence of the obligately chemolithoautotrophic, facultatively anaerobic bacterium Thiobacillus denitrificans.
<3>J. Bacteriol.
<4>188
<5>1473-1488
<6>2006
<7>The complete genome sequence of Thiobacillus denitrificans ATCC 25259 is the first to become
available for an obligately chemolithoautotrophic,
sulfur-compound-oxidizing, beta-proteobacterium. Analysis of the
2,909,809-bp genome will facilitate our molecular and biochemical
understanding of the unusual metabolic repertoire of this bacterium,
including its ability to couple denitrification to sulfur-compound
oxidation, to catalyze anaerobic, nitrate-dependent oxidation of Fe(II)
and U(IV), and to oxidize mineral electron donors. Notable genomic
features include (i) genes encoding c-type cytochromes totaling 1 to 2
percent of the genome, which is a proportion greater than for almost all
bacterial and archaeal species sequenced to date, (ii) genes encoding two
[NiFe]hydrogenases, which is particularly significant because no
information on hydrogenases has previously been reported for T.
denitrificans and hydrogen oxidation appears to be critical for anaerobic
U(IV) oxidation by this species, (iii) a diverse complement of more than
50 genes associated with sulfur-compound oxidation (including sox genes,
dsr genes, and genes associated with the AMP-dependent oxidation of
sulfite to sulfate), some of which occur in multiple (up to eight) copies,
(iv) a relatively large number of genes associated with inorganic ion
transport and heavy metal resistance, and (v) a paucity of genes encoding
organic-compound transporters, commensurate with obligate
chemolithoautotrophy. Ultimately, the genome sequence of T. denitrificans
will enable elucidation of the mechanisms of aerobic and anaerobic
sulfur-compound oxidation by beta-proteobacteria and will help reveal the
molecular basis of this organism's role in major biogeochemical cycles
(i.e., those involving sulfur, nitrogen, and carbon) and groundwater
restoration.

<>

<1>Bellgard, M.I., Wanchanthuek, P., La, T., Ryan, K., Moolhuijzen, P., Albertyn, Z., Shaban, B., Motro, Y., Dunn, D.S., Schibeci, D., Hunter, A., Barrero, R., Phillips, N.D., Hampson, D.J.
<2>Genome sequence of the pathogenic intestinal spirochete brachyspira hyodysenteriae reveals adaptations to its lifestyle in the porcine large intestine.
<3>PLoS ONE
<4>4
<5>E4641
<6>2009
<7>Brachyspira hyodysenteriae is an anaerobic intestinal spirochete that
colonizes the large intestine of pigs and causes swine dysentery, a
disease of significant economic importance. The genome sequence of B.
hyodysenteriae strain WA1 was determined, making it the first
representative of the genus Brachyspira to be sequenced, and the
seventeenth spirochete genome to be reported. The genome consisted of a
circular 3,000,694 base pair (bp) chromosome, and a 35,940 bp circular
plasmid that has not previously been described. The spirochete had 2,122
protein-coding sequences. Of the predicted proteins, more had similarities
to proteins of the enteric Escherichia coli and Clostridium species than
they did to proteins of other spirochetes. Many of these genes were
associated with transport and metabolism, and they may have been gradually
acquired through horizontal gene transfer in the environment of the large
intestine. A reconstruction of central metabolic pathways identified a
complete set of coding sequences for glycolysis, gluconeogenesis, a
non-oxidative pentose phosphate pathway, nucleotide metabolism,
lipooligosaccharide biosynthesis, and a respiratory electron transport
chain. A notable finding was the presence on the plasmid of the genes
involved in rhamnose biosynthesis. Potential virulence genes included
those for 15 proteases and six hemolysins. Other adaptations to an enteric
lifestyle included the presence of large numbers of genes associated with
chemotaxis and motility. B. hyodysenteriae has diverged from other
spirochetes in the process of accommodating to its habitat in the porcine
large intestine.

<>

<1>Bellon, B.
<2>Construction of restriction maps.
<3>Comput. Appl. Biosci.
<4>4
<5>111-116
<6>1988
<7>A computer program is described, which constructs maps of restriction
endonuclease cleavage sites in linear or circular DNA molecules, given the
fragment lengths in single and double digestions with two enzymes.  The
algorithm is based upon a partition method and a very simple rule to chain
fragments.  The program is written in Prolog II.

<>

<1>Bellstein, F., Dreiseikelmann, B.
<2>Temperate bacteriophage Phi O18P from an Aeromonas media isolate: Characterization and complete genome sequence.
<3>Virology
<4>373
<5>25-29
<6>2008
<7>group of 74 Aeromonas isolates from surface water of three ponds in Bielefeld, Germany was
screened for prophage induction after UV
irradiation. The phage Phi O18P was induced from the Aeromonas media
isolate O18. Phi O18P belongs to the Myoviridae phage family. The
complete nucleotide sequence of the double stranded DNA genome
ofbacteriophage Phi O18P consists of 33,985 bp. The genome has 5'
protruding cohesive ends of 16 bases. On the Phi O18P genome 46 open
reading frames (orfs) were identified which are organized in the
modules integration and regulation, replication, head, packaging, tail
and lysis. Additionally the phage DNA includes a methylase gene.
Comparison of the genome architecture with those of other
bacteriophages revealed significant similarities to the P2 phage family
and especially to the prophages of Aeromonas salmonicida and the Vibrio
cholerae phage K139.

<>

<1>Belogurov, A.A., Delver, E.P.
<2>A motif conserved among the type I restriction-modification enzymes and antirestriction proteins: a possible basis for mechanism of action of plasmid-encoded antirestriction functions.
<3>Nucleic Acids Res.
<4>23
<5>785-787
<6>1995
<7>Antirestriction proteins Ard encoded by some self-transmissible plasmids specifically inhibit
restriction by members of all three families of type I restriction-modification (R-M) systems
in E. coli. Recently, we have identified the amino acid region, 'antirestriction' domain,
that is conserved within different plasmid and phage T7-encoded antirestriction proteins and
may be involved in interaction with the type I R-M systems. In this paper we demonstrate that
this amino acid sequence shares considerable similarity with a well-known conserved sequence
(the Argos repeat) found in the DNA sequence specificity (S) polypeptides of type I systems.
We suggest that the presence of these similar motifs in restriction and antirestriction
proteins may give a structural basis for their interaction and that the antirestriction action
of Ard proteins may be a result of the competition between the 'antirestriction' domains of
Ard proteins and the similar conserved domains of the S subunits that are believed to play a
role in the subunit assembly of type I R-M systems.

<>

<1>Belogurov, A.A., Delver, E.P., Agafonova, O.V., Belogurova, N.G., Lee, L.Y., Kado, C.I.
<2>Antirestriction protein Ard (Type C) encoded by IncW plasmid pSa has a high similarity to the "Protein Transport" domain of TraC1 primase of promiscuous plasmid RP4.
<3>J. Mol. Biol.
<4>296
<5>969-977
<6>2000
<7>The IncW plasmid pSa contains the gene ard encoding an antirestriction function that is
specific for type I restriction and modification systems. The nucleotide
sequence of ard was determined and an appropriate polypeptide of about 33 kDa was identified
in Escherichia coli T7 expression system. Analysis of deduced amino acid
sequence of Ard encoded by pSa revealed that this protein has no significant similarities with
the known Ard proteins (ArdA and ArdB types) except the "antirestriction"
motif (14 amino acid residues in length) conserved for all known Ard proteins. This finding
suggests that pSa Ard may be classified as a new type of Ard proteins which we
designated ArdC. The remarkable feature of ArdC is that it has a high degree of similarity
(about 38 % identity) to the N-terminal region of RP4 TraC1 primase which
includes about 300 amino acid residues and seems to be essential for binding to the
single-stranded DNA and TraC1 protein transport to the recipient cells during the
conjugal transfer of plasmid DNA. ArdC also binds to single-stranded DNA. In addition, this
protein is able in vitro to protect the single-stranded but not double-stranded
plasmid DNA against the activity of type II restriction endonuclease HhaI that cleaves both
single and double-stranded DNA. We suggest that like TraC1, ArdC would be
transported as a result of their interaction with the single-stranded DNA of transferred
plasmid strand during conjugative passage through the cell envelope to the recipient
bacterium. Such properties of ArdC protein might be useful to protect immediately the incoming
single-stranded DNA from the host endonucleases.

<>

<1>Belogurov, A.A., Delver, E.P., Rodzevich, O.V.
<2>IncN plasmid pKM101 and IncI1 Plasmid ColIb-P9 encode homologous antirestriction proteins in their leading regions.
<3>J. Bacteriol.
<4>174
<5>5079-5085
<6>1992
<7>The IncN plasmid pKM101 (a derivative of R46), like the the IncI1 plasmid ColIb-P9, carries a
gene (ardA, for alleviation of restriction of DNA) encoding an antirestriction function, ardA
was located about 4 kb from the origin of transfer, in the region transferred early during
bacterial conjugation. The nucleotide sequence of ardA was determined, and an appropriate
polypeptide with the predicted molecular weight of about 19,500 was identified in maxicells of
Escherichia coli. Comparison of the deduced amino acid sequences of the antirestriction
proteins of the unrelated plasmids pKM101 and ColIb (ArdA and Ard, respectively) revealed that
these proteins have about 60% identify. Like ColIb ard, pMK101 ArdA specifically inhibits both
the restriction and modification activities of five type I systems of E. coli tested and does
not influence type III (EcoPI) restriction or the 5-methylcytosine-specific restriction
systems McrA and McrBC. However, in contrast to ColIb ard, pKM101 ArdA is effective against
the type II enzyme EcoRI. The Ard proteins are believed to overcome the host restriction
barrier during bacterial conjugation. We have also identified two other genes of pKM101, ardR
and ardK, which seem to control ardA activity and ardA-medicated lethality, respectively. Our
findings suggest that ardR may serve as a genetic switch that determines whether the ard-A
encoded antirestriction function is induced during mating.

<>

<1>Belogurov, A.A., Delver, E.P., Rodzevich, O.V.
<2>Plasmid pKM101 encodes two nonhomologous antirestriction proteins (ArdA and ArdB) whose expression is controlled by homologous regulatory sequences.
<3>J. Bacteriol.
<4>175
<5>4843-4850
<6>1993
<7>The Inc plasmid pKM101 (a derivative of R46) encodes the antirestriction protein ArdB
(alleviation of restriction of DNA) in addition to another antirestriction protein, ArdA,
described previously. The relevant gene, ardB was located in the leading region of pKM101,
about 7 kb from oriT. The nucleotide sequence of ardB was determined, and an appropriate
polypeptide was identified in maxicells of Escherichia coli. Like ArdA, ArdB efficiently
inhibits restriction by members of the three known families of type I systems of E. coli and
only slightly affects the type II enzymes, EcoRI. However, in contrast to ArdA, ArdB is
ineffective against the modification activity of the type I (EcoK) system. Comparison of
deduced amino acid sequences of ArdA and ArdB revealed only one small region of similarity
(nine residues), suggesting that this region may be somehow involved in the interaction with
the type I restriction systems. We also found that the expression of both ardA and ardB genes
is controlled jointly by two pKM101-encoded proteins, ArdK and ArdR, with molecular weights of
about 15,000 and 20,000 respectively. The finding that the sequences immediately upstream of
ardA and ardB share about 94% identity over 218 bp suggests that their expression may be
controlled by ArdK and ArdR at the transcriptional level. Deletion studies and promoter probe
analysis of these sequences revealed the regions responsible for the action of ArdK and ArdR
as regulatory proteins. We propose that both types of antirestriction proteins may play a
pivotal role in overcoming the host restriction barrier by self-transmissible broad-host-range
plasmids. It seems likely that the ardKR-dependent regulatory system serves in this case as a
genetic switch that controls the expression of plasmid-encoded antirestriction functions
during mating.

<>

<1>Belogurov, A.A., Efimova, E.P., Delver, E.P., Zavilgelsky, G.B.
<2>Alleviation of I type restriction in E. coli: Effect of a dam mutation.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>9
<5>10-16
<6>1987
<7>The host-controlled EcoK-restriction of unmodified phages lambda.0 and T7 ocr, is 100-fold
alleviated in dam- mutants of E. coli.  In addition the EcoK modification activity is
considerably decreased in dam- strains.  Type I and III restriction (EcoB, EcoD, EcoK and
EcoPI) were relieved in dam- mutants, but no alleviation of EcoRI restriction occurred in dam-
strains.  We interpret the alleviation of the type I restriction in dam- mutants as a
consequence of induction of the function, which interferes with the type I restriction
systems.

<>

<1>Belogurov, A.A., Efimova, E.P., Delver, E.P., Zavilgelsky, G.B.
<2>Alleviation of type I restriction in Escherichia coli: Effect of 2-amino-purine and 5-bromouracil.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>12
<5>34-40
<6>1987
<7>2-aminopurine and 5-bromouracil, strong mutagens of the base analog type, were found to induce
efficiently the alleviation of type I restriction in Escherichia coli.  2-AP induced
restriction alleviation occurs in recA, lexA and mut- mutants, but no additional relief of
restriction is registered in dam- bacteria in the presence of sublethal 2-AP concentrations.
2-AP specifically alleviates type I restriction in Escherichia coli (EcoA, EcoB, EcoD, and
EcoK) and does not affect restriction system of types II (EcoRI) and III (EcoPI).  We suggest
that 2-AP-induced mismatches and other replication errors may be signals inducing restriction
alleviation in Escherichia coli.

<>

<1>Belogurov, A.A., Yussifov, T.N., Kotova, V.U., Zavilgelsky, G.B.
<2>The novel gene(s) ARD of plasmid pKM101: alleviation of EcoK restriction.
<3>Mol. Gen. Genet.
<4>198
<5>509-513
<6>1985
<7>The host-controlled K restriction of unmodified phage lambda was 10-100 fold
alleviated in the wild-type strain E.coli K12, carrying plasmid pKM101 of
incompability group N.  pKM101-mediated release of K restriction was also
observed in lexA-, recA-, and recB- strains of E.coli K12.  By restriction
mapping Tn5 insertions in pKM101, which reduced pKM101-mediated alleviation of
restriction, were shown to be located within the BglIIB fragment approximately
11kb anticlockwise from the RI site of pKM101.  We have termed the gene(s)
promoting the alleviation of K restriction of phage lambda ard (alleviation of
restriction of DNA).  It was shown (1) that ard function affected only the EcoK
restriction system and not the EcoB, EcoRI, EcoRIII, or EcoPI systems, (2) ard
gene(s) did not mediate EcoK type modification of lambda DNA and did not
increase the modification activity of the EcoK system in a way similar to that
observed with gene ral of bacteriophage lambda.

<>

<1>Belogurov, A.A., Zavilgelskii, G.B.
<2>Effect of plasmid pKM101 on the K-specific restriction-modification of bacteriophage lambda DNA.
<3>Dokl. Akad. Nauk.
<4>269
<5>738-741
<6>1983
<7>
<>

<1>Belogurov, A.A., Zavilgelsky, G.B.
<2>The repair of the interstrand cross links in DNA: Role of restriction system K.
<3>Mol. Biol. (Mosk)
<4>13
<5>160-168
<6>1979
<7>The interstrand cross links formed in the DNA after psoralen plus light treatment of bacteria
E. coli K12 and bacteriophage lambda are repaired by the restriction and modification system
K.  Strain E. coli C, which does not have the restriction and modification system is 2 times
more sensitive to psoralen plus light treatment, than the wild type E. coli K12.  But these
strains are equally sensitive to inactivation by UV-light (254 nm).  By studying the
sedimentation of bacterial DNA in alkaline sucrose it was found that cells of E. coli C are
deficieint in filling of gaps, appearing in parental DNA after enzymatic excision of the arms
of cross links.  In experiments with bacteriophage lambda (W-reactivation,
prophage-reactivation), treated by psoralen plus light, it was shown that E. coli C and
mutants of E. coli K12 rR-mR+/- are deficient in non-recombination lexA-dependent repair of
cross links.  The function of restriction and recognition of restriction and modification
system are essential for repair of cross links.  It is suggested that the restriction and
modification system K acts either as a regulatory protein or as a protector of single-stranded
regions of DNA against nuclease attack.

<>

<1>Belova, T.S., Surikov, N.N., Prozorov, A.A.
<2>Transformation and transduction of Bacillus subtilis strains possessing the BsuR system of restriction-modification by nonmodified and modified plasmid pUB110 DNA.
<3>Genetika
<4>17
<5>794-800
<6>1981
<7>Plasmid pUB110 DNA is restricted and modified during transformation of Bacillus subtilis cells
possessing the BsuR system of restriction-modification.  Restriction has comparatively little
influence on the frequency of plasmid transformation only reducing it 20 times.  The frequency
of transduction of nonmodified plasmid DNA into modifying recipient cells, using phage Ar9 is
also reduced a little.  The frequency of transduction of chromosomal markers is much more
lowered under the same experimental conditions.

<>

<1>Beltramo, C., Oraby, M., Bourel, G., Garmyn, D., Guzzo, J.
<2>A new vector, pGID052, for genetic transfer in Oenococcus oeni.
<3>FEMS Microbiol. Lett.
<4>236
<5>53-60
<6>2004
<7>Despite the large number of techniques available for the transformation of
bacteria, several species are still resistant to the introduction of
foreign DNA. Oenococcus oeni are among the organisms that are particularly
refractory to transformation. However, conjugal experiments from
Lactococcus lactis to O. oeni with a new plasmid, pGID052, were performed
via mobilization with success. This plasmid, a derivative of pORI19,
encompasses: (i) the oriT of pIP501, which permitted the transfer to O.
oeni, (ii) the replication genes of a native Leuconostoc citreum plasmid.
Frequencies of 10(-7) conjugants per recipient were found. The transfer
did not affect the structure of this low-copy-number plasmid. Moreover,
pGID052 seems segregationally stable and could be used in the future as an
expression vector.

<>

<1>Ben Zakour, N.L., Bannoehr, J., van den Broek, A.H., Thoday, K.L., Fitzgerald, J.R.
<2>Complete genome sequence of the canine pathogen Staphylococcus pseudintermedius.
<3>J. Bacteriol.
<4>193
<5>2363-2364
<6>2011
<7>We report the first whole genome sequence for a clinical isolate of Staphylococcus
pseudintermedius (ED99), the major pathogen responsible for canine bacterial pyoderma. S.
pseudintermedius contains numerous mobile genetic elements, and encodes an array of putative
virulence factors including superantigenic, cytolytic and exfoliative toxins, and cell-wall
associated surface proteins.

<>

<1>Ben-Hattar, J., Jiricny, J.
<2>Effect of cytosine methylation on the cleavage of oligonucleotide duplexes with restriction endonucleases HpaII and MspI.
<3>Nucleic Acids Res.
<4>16
<5>4160
<6>1988
<7>the pattern of eucaryotic DNA methylation is commonly determined by restriction
analysis with methylation-sensitive enzymes.  Due to the likely biological
significance of hemimethylated CpG dinucleotides, we investigated the MspI and
HpaII hydrolysis of synthetic 29-mer oligos (Fig. 1a), unmethylated,
hemimethylated or fully-methylated at the internal C residue of their
recognition site CCGG.  As shown in Fig. 1b, MspI cleaved all the four
substrates, irrespective of their state of methylation.  HpaII failed to digest
the symmetrically-methylated duplex (mC29/mC33), while the unmethylated duplex
(C29/C33) was completely and efficiently digested.  Surprisingly, neither
cleavage nor nicking of the unmethylated strand were observed with the
hemimethylated substrates.  These results demonstrate that the use of HpaII in
the search for regions of active, unmethylated chromatin would fail to identify
hemimethylated CpG sites, recently shown to be involved in the process of
transcriptional gene activation.

<>

<1>Benachour, A., Frere, J., Novel, G.
<2>pUCL287 plasmid from Tetragenococcus halophila (Pediococcus halophilus) ATCC 33315 represents a new theta-type replicon family of lactic acid bacteria.
<3>FEMS Microbiol. Lett.
<4>128
<5>167-175
<6>1995
<7>A cryptic plasmid, pUCL287, was isolated from Tetragenococcus halophila
(Pediococcus halophilus) ATCC 33315. It had a theta-type mechanism of
replication in its natural host. Its minimal replicon, Rep287, was
isolated on a 1.6-kb EcoRI fragment. The Rep287 host range included the
genera Pediococcus, Enterococcus, Lactobacillus and Leuconostoc but not
genus Lactococcus. Plasmids hybridizing to pUCL287 are rare among lactic
acid bacteria. As assessed by hybridization, Rep287 is dissimilar to pAM
beta 1, pIP501 and pUCL22, representatives of the most common theta-type
replicon groups in Gram-positive bacteria. Therefore, pUCL287 appears to
represent a new theta-type replicon family from lactic acid bacteria.

<>

<1>Benahmed, F.H., Gopinath, G.R., Harbottle, H., Cotta, M.A., Luo, Y., Henderson, C., Teri, P., Soppet, D., Rasmussen, M., Whitehead, T.R., Davidson, M.
<2>Draft Genome Sequences of Streptococcus bovis Strains ATCC 33317 and JB1.
<3>Genome Announcements
<4>2
<5>e01012-14
<6>2014
<7>We report the draft genome sequences of Streptococcus bovis strain ATCC 33317 (CVM42251)
isolated from cow dung and strain JB1 (CVM42252) isolated from a cow
rumen in 1977. The strains were sequenced using the Genome Sequencer FLX 454
system. The genome sizes are approximately 2 Mb and 2.2 Mb, respectively.

<>

<1>Benahmed, F.H., Gopinath, G.R., Wang, H., Jean-Gilles, B.J., Grim, C., Cheng, C.M., McClelland, M., Ayers, S., Abbott, J., Desai, P., Frye, J.G., Weinstock, G., Hammack, T.S., Hanes, D.E., Rasmussen, M.A., Davidson, M.K.
<2>Whole-Genome Sequencing of Salmonella enterica subsp. enterica Serovar Cubana Strains Isolated from Agricultural Sources.
<3>Genome Announcements
<4>2
<5>e01184-13
<6>2014
<7>We report the draft genomes of Salmonella enterica subsp. enterica serovar Cubana strain
CVM42234, isolated from chick feed in 2012, and S. Cubana strain 76814,
isolated from swine in 2004. The genome sizes are 4,975,046 and 4,936,251 bp,
respectively.

<>

<1>Benbadis, L., Faelen, M., Slos, P., Fazel, A., Mercenier, A.
<2>Characterization and comparison of virulent bacteriophages of Streptococcus thermophilus isolated from yogurt.
<3>Biochimie
<4>72
<5>855-862
<6>1990
<7>Seven virulent bacteriophages of Streptococcus thermophilus were characterized at the
molecular level and classified into 2 subgroups (A and B) by DNA/DNA hybridization experiments
and analysis of their structural proteins. Two representatives of subgroups A and B were
compared to 3 representatives of Neve's subgroups I, II and III (Neve et al, 1989) by
Southern blot experiments. These isometric-headed phages possess a double-stranded DNA genome
varying between 30-44 kilobase (kb) pairs. Subgroup A is composed of 3 phages (phi 57 as
representative) with similar structural proteins as determined by sodium dodecyl
sulfate-polyacrylamide gel (SDS-PAGE) electrophoresis (estimated molecular weight of 31,000
and 27,500 for phage phi 57 and 32,000 and 27,000 for the 2 others). A common structural
protein of 43,000 was found for phages of subgroup B. Phages phi 57(subgroup D) and a10/J9 or
PO (Neve's subgroups I or II, respectively) belonged to the same subgroup as determined by
DNA/DNA hybridization experiments. Partial DNA homology was detected among all the phages
tested except for phage phi ST27 of AW Jarvis. Phage-host interactions were also investigated
by cross-propagation of the 7 studied phages on different indicator strains. A complete lack
of correlation existed between the DNA homology grouping of the phages and their host range.
Various restriction-modification systems were detected in some of the Streptococcus
thermophilus strains.

<>

<1>Benbadis, L., Garel, J.R., Hartley, D.L.
<2>Purification, properties, and sequence specificity of SslI, a new type II restriction endonuclease from Streptococcus salivarius subsp. thermophilus.
<3>Appl. Environ. Microbiol.
<4>57
<5>3677-3678
<6>1991
<7>SslI, a type II restriction endonuclease, was purified from Streptococcus
salivarius subsp. thermophilus strain BSN 45.  SslI is an isoschizomer of
BstNI.  SslI activity was maximum at pH 8.8, 0 to 50 mM NaCl, 2 to 8 mM Mg2+,
and 42C.  Activity against phage DNA in vitro was demonstrated.

<>

<1>Bench, S.R., Hanson, T.E., Williamson, K.E., Ghosh, D., Radosovich, M., Wang, K., Wommack, K.E.
<2>Metagenomic characterization of Chesapeake Bay virioplankton.
<3>Appl. Environ. Microbiol.
<4>73
<5>7629-7641
<6>2007
<7>Viruses are ubiquitous and abundant throughout the biosphere. In marine systems,
virus-mediated processes can have significant impacts on microbial diversity and on global
biogeocehmical cycling. However, viral genetic diversity remains poorly characterized. To
address this shortcoming, a metagenomic library was constructed from Chesapeake Bay
virioplankton. The resulting sequences constitute the largest collection of long-read
double-stranded DNA (dsDNA) viral metagenome data reported to date. BLAST homology comparisons
showed that Chesapeake Bay virioplankton contained a high proportion of unknown (homologous
only to environmental sequences) and novel (no significant homolog) sequences. This analysis
suggests that dsDNA viruses are likely one of the largest reservoirs of unknown genetic
diversity in the biosphere. The taxonomic origin of BLAST homologs to viral library sequences
agreed well with reported abundances of cooccurring bacterial subphyla within the estuary and
indicated that cyanophages were abundant. However, the low proportion of Siphophage homologs
contradicts a previous assertion that this family comprises most bacteriophage diversity.
Identification and analyses of cyanobacterial homologs of the psbA gene illustrated the value
of metagenomic studies of virioplankton. The phylogeny of inferred PsbA protein sequences
suggested that Chesapeake Bay cyanophage strains are endemic in that environment.  The ratio
of psbA homologous sequences to total cyanophage sequences in the metagenome indicated that
the psbA gene may be nearly universal in Chesapeake Bay cyanophage genomes. Furthermore, the
low frequency ofpsbD homologs in the library supports the prediction that Chesapeake Bay
cyanophage populations are dominated by Podoviridae.

<>

<1>Bendall, M.L., Luong, K., Wetmore, K.M., Blow, M., Korlach, J., Deutschbauer, A., Malmstrom, R.R.
<2>Exploring the roles of DNA methylation in the metal-reducing bacterium Shewanella oneidensis MR-1.
<3>J. Bacteriol.
<4>195
<5>4966-4974
<6>2013
<7>We performed whole genome analyses of DNA methylation in Shewanella oneidensis MR-1 to examine
its possible role in regulating gene expression and other
cellular processes. Single-Molecule Real Time (SMRT) sequencing revealed
extensive methylation of adenine (N6mA) throughout the genome. These methylated
bases were located in five sequence motifs, including three novel targets for
Type I restriction/modification enzymes. The sequence motifs targeted by putative
methyltranferases were determined via SMRT sequencing of gene knockout mutants.
In addition, we found S. oneidensis MR-1 cultures grown under various culture
conditions displayed different DNA methylation patterns. However, the small
number of differentially methylated sites could not be directly linked to the
much larger number of differentially expressed genes in these conditions,
suggesting DNA methylation is not a major regulator of gene expression in S.
oneidensis MR-1. The enrichment of methylated GATC motifs in the origin of
replication indicate DNA methylation may regulate genome replication in a manner
similar to that seen in Escherichia coli. Furthermore, comparative analyses
suggest that many Gammaproteobacteria, including all members of the
Shewanellaceae family, may also utilize DNA methylation to regulate genome
replication.

<>

<1>Bender, J.
<2>A vicious cycle: RNA silencing and DNA methylation in plants.
<3>Cell
<4>106
<5>129-132
<6>2001
<7>Several new studies have stimulated intense interest in understanding the mechanism and
evolutionary significance of RNA silencing, the targeted degradation of RNA.  RNA silencing is
typically triggered by double-stranded RNA (dsRNA).  The dsRNA trigger is diced into very
small species of 21-25 nt (siRNAs), which then guide sequence-specific cleavage of other
homologous RNAs, such as messenger RNAs.  Thus, both the trigger and target RNAs are
ultimately destroyed.  This mechanism occurs in eukaryotic organisms as diverse as the
laboratory plant Arabidopsis thaliana, nematode worms, and mice.  It is speculated that RNA
silencing exists as a defense against invasive dsRNA species such as RNA viruses.  In fact,
plants can recover from infection by RNA viruses via RNA silencing.  RNA silencing might also
have a role in developmental regulation.  Furthermore, RNA silencing induced by injected dsRNA
or dsRNA expressed from a transgene provides a powerful tool for reverse genetics in animals
and plants.

<>

<1>Bendiks, Z.A., Lang, J.M., Darling, A.E., Eisen, J.A., Coil, D.A.
<2>Draft Genome Sequence of Microbacterium sp. Strain UCD-TDU (Phylum Actinobacteria).
<3>Genome Announcements
<4>1
<5>e0012013
<6>2013
<7>Here, we present the draft genome sequence of Microbacterium sp. strain UCD-TDU,  a member of
the phylum Actinobacteria. The assembly contains 3,746,321 bp (in 8
scaffolds). This strain was isolated from a residential toilet as part of an
undergraduate student research project to sequence reference genomes of microbes
from the built environment.

<>

<1>Bendjama, E., Loucif, L., Diene, S.M., Michelle, C., Gacemi-Kirane, D., Rolain, J.M.
<2>Non-contiguous finished genome sequence and description of Paucisalibacillus algeriensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1352-1365
<6>2014
<7>Paucisalibacillus algeriensis strain EB02(T) is the type strain of Paucisalibacillus
algeriensis sp. nov., a new species within the genus
Paucisalibacillus. This strain, whose genome is described here, was isolated from
soil sample from the hypersaline lake Ezzemoul Sabkha in northeastern Algeria.
Paucisalibacillus algeriensis is a Gram-positive and strictly aerobic bacterium.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. The 4,006,766 bp long genome (1 chromosome but no
plasmid) exhibits a low G+C content of 36% and contains 3,956 protein-coding and
82 RNA genes, including 9 rRNA genes.

<>

<1>Bendjama, E., Loucif, L., Diene, S.M., Michelle, C., Gacemi-Kirane, D., Rolain, J.M.
<2>Non-contiguous finished genome sequence and description of Bacillus massilioalgeriensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1046-1061
<6>2014
<7>Strain EB01(T) sp. nov. is the type strain of Bacillus massilioalgeriensis, a new species
within the genus Bacillus. This strain, whose genome is described here,
was isolated from sediment sample of the hypersaline lake Ezzemoul sabkha in
northeastern Algeria. B. massilioalgeriensis is a facultative anaerobic
Gram-positive bacillus. Here we describe the features of this organism, together
with the complete genome sequence and annotation. The 5,269,577 bp long genome
contains 5,098 protein-coding and 95 RNA genes, including 12 rRNA genes.

<>

<1>Beneduzi, A., Campos, S., Ambrosini, A., de Souza, R., Granada, C., Costa, P., Arruda, L., Moreira, F., Vargas, L.K., Weiss, V., Tieppo, E., Faoro, H., de Souza, E.M., Pedrosa, F.O., Passaglia, L.M.
<2>Genome Sequence of the Diazotrophic Gram-Positive Rhizobacterium Paenibacillus riograndensis SBR5T.
<3>J. Bacteriol.
<4>193
<5>6391-6392
<6>2011
<7>Paenibacillus riograndensis SBR5(T), a nitrogen-fixing Gram-positive rhizobacterium isolated
from a wheat field in the south of Brazil, has a
great potential for agricultural applications due to its plant growth
promotion effects. Here we present the draft genome sequence of P.
riograndensis SBR5(T). Its 7.37-Mb genome encodes determinants of the
diazotrophic lifestyle and plant growth promotion, such as nitrogen
fixation, antibiotic resistance, nitrate utilization, and iron uptake.

<>

<1>Benetti, R., Gonzalo, S., Jaco, I., Munoz, P., Gonzalez, S., Schoeftner, S., Murchison, E., Andl, T., Chen, T., Klatt, P., Li, E., Serrano, M., Millar, S., Hannon, G., Blasco, M.A.
<2>A mammalian microRNA cluster controls DNA methylation and telomere recombination via Rbl2-dependent regulation of DNA methyltransferases (vol 15, pg 268, 2008).
<3>Nat. Struct. Mol. Biol.
<4>15
<5>998
<6>2008
<7>Dicer initiates RNA interference by generating small RNAs involved in various silencing
pathways.  Dicer participates in centromeric silencing, but its role in the epigenetic
regulation of other chromatin domains has not been explored.  Here we show that Dicer1
deficiency in Mus musculus leads to decreased DNA methylation, concomitant with increased
telomere recombination and telomere elongation.  These DNA-methylation defects correlate with
decreased expression of Dnmt1, Dnmt3a and Dnmt3b DNA methyltransferases, and methylation
levels can be recoverd by their overexpression.  We identify the retinoblastoma-like 2 protein
as responsible for decreased Dnmt expression in Dicer1-null cells, suggesting the existence of
Dicer-dependent small RNAs that target Rbl2.  We identify the miR-290 cluster as being
downregulated in Dicer1-deficient cells and show that it silences Rbl2, thereby controlling
Dnmt expression.  These results identify a pathway by which miR-290 directly regulates
Rbl2-dependent Dnmt expression, indirectly affecting telomere-length homeostasis.

<>

<1>Benight, A.S., Gallo, F.J., Paner, T.M., Bishop, K.D., Faldasz, B.D., Lane, M.J.
<2>Sequence context and DNA reactivity: Application to sequence-specific cleavage of DNA.
<3>Adv. Biophys. Chem., JAI Press, Inc., Bush, C.A., 
<4>5
<5>1-55
<6>1995
<7>Reactions between duplex DNA and ligands are largely dictated and mediated by the interplay of
the structural, thermodynamic, and dynamic characteristics of DNA and the recognition
mechanisms of reacting ligands.  Ligands that bind to DNA span a broad range of sizes from
small cations to large proteins and assembled protein aggregates.  As might be expected, a
wide variety of experimental strategies have been exploited to examine the sequence
specificity, or lack thereof, exhibited by ligands that interact with DNA.  Sequence-dependent
variations in local conformation and charge configuration along DNA are thought to be the
principal means by which ligands discriminate between various DNA sequences.  In efforts to
define the thermodynamic basis of such sequence-specific discrimination, a variety of
parameters have been evaluated from studies of DNA alone and ligand/DNA complexes.

<>

<1>Benkovic, S.J., Baker, S.J., Alley, M.R.K., Woo, Y.H., Zhang, Y.K., Akama, T., Mao, W.M., Baboval, J., Rajagopalan, P.T.R., Wall, M., Kahng, L.S., Tavassoli, A., Shapiro, L.
<2>Identification of borinic esters as inhibitors of bacterial cell growth and bacterial methyltransferases, CcrM and MenH.
<3>J. Med. Chem.
<4>48
<5>7468-7476
<6>2005
<7>As bacteria continue to develop resistance toward current antibiotics, we find ourselves in a
continual battle to identify new antibacterial agents and targets. We report herein a class of
boron-containing compounds termed borinic esters that have broad spectrum antibacterial
activity with minimum inhibitory concentrations (MIC) in the low microgram/mL range. These
compounds were identified by screening for inhibitors against Caulobacter crescentus CcrM, an
essential DNA methyltransferase from gram negative alpha-proteobacteria. In addition, we
demonstrate that borinic esters inhibit menaquinone methyltransferase in gram positive
bacteria using a new biochemical assay for MenH from Bacillus subtilis. Our data demonstrate
the potential for further development of borinic esters as antibacterial agents as well as
leads to explore more specific inhibitors against two essential bacterial enzymes.

<>

<1>Benkovic, S.J., Berdis, A., Lee, I., Shapiro, L., Wright, R., Stephens, C., Kahng, L.S.
<2>DNA adenine methyltransferases and uses thereof.
<3>US Patent Office
<4>US 6413751
<5>
<6>2002
<7>The present invention relates to the isolation and sequencing of a novel class of
methyltransferase genes, including the methyltransferase
gene from Rhizobium meliloti, Agrobacterium tumefaciens, Brucella
abortus, and Helicobacter pylori. The invention further comprises
efficient methods of assaying methyltransferase activity.

<>

<1>Benkovic, S.J., Shapiro, L., Baker, S.J., Wahnon, D.C., Wall, M.
<2>Preparation of boronic acid-containing purines, nucleosides, and amino acids as inhibitors of bacterial adenine DNA methyltransferases.
<3>International Patent Office
<4>WO 200075142 A
<5>55
<6>2000
<7>
<>

<1>Benkovic, S.J., Shapiro, L., Wright, R., Stephens, C., Kahng, L.S., Berdis, A., Lee, I.
<2>Purine nucleoside analogs as antibiotic inhibitors of bacterial DNA methyltransferases.
<3>US Patent Office
<4>US 20050227933 A
<5>
<6>2005
<7>Methods for treating and/or preventing disease conditions caused or induced or aggravated by
microbes, especially bacteria, by inhibiting DNA methyltransferase activity, such as by
administering to an animal a DNA methyltransferase inhibitor, are disclosed, along with
methods of reducing or ablating virulence in bacteria by inhibiting DNA methyltransferase
activity.

<>

<1>Benner, J., Dalton, M.A., Xu, S., Dore, A.
<2>Method for Cloning And Expression Of TspRI Restriction Endonuclease And TspRI Methylase In E. coli.
<3>Japanese Patent Office
<4>JP 2005506843 A
<5>
<6>2005
<7>
<>

<1>Benner, J.S.
<2>Method for producing the NdeI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5139942
<5>
<6>1992
<7>The present invention is directed to a method for cloning and producing the NdeI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Neisseria denitrificans
into a host whereby the restriction gene is expressed; 2) fermenting the host which contains
the vector encoding and expressing the NdeI restriction endonuclease, and 3) purifying the
NdeI restriction endonuclease from the fermented host which contains the vector encoding and
expressing the NdeI restriction endonuclease activity.

<>

<1>Benner, J.S.
<2>Method for producing the NdeI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0343009 B
<5>
<6>1994
<7>The present invention relates to clones for the NdeI restriction endonuclease and modification
methylase, and to the production of these enzymes from the clones.

<>

<1>Benner, J.S., Coe, L.H.
<2>Method for producing the SspI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0707066 A
<5>
<6>1996
<7>The present invention is directed to a method for cloning and producing the SspI
restriction endonuclease by 1) introducing the restriction endonuclease gene from Sphaerotilus
species into a host whereby the restriction gene is expressed; 2) fementing the host which
contains
the vector encoding and expressing the SspI restriction endonuclease, and 3) purifying the
SspI
restriction endonuclease from the fermented host which contains the vector encoding and
expressing the SspI restriction endonuclease activity.

<>

<1>Benner, J.S., Coe, L.H.
<2>Method for producing the SspI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5516678
<5>
<6>1996
<7>The present invention is directed to a method for cloning and producing the SspI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Sphaerotilus species
into a host whereby the restriction gene is expressed; 2) fermenting the host which contains
the vector encoding and expressing the SspI restriction endonuclease from the fermented host
which contains the vector encoding and expressing the SspI restriction endonuclease activity.

<>

<1>Benner, J.S., Rees, P.
<2>Method for producing the HpaI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0422951 A
<5>
<6>1995
<7>The present invention relates to recombinant DNA which encodes the HpaI restriction
endonuclease and modification methylase, as well as to the enzymes produced from such a
recombinant DNA.  A method is described by which the HpaI restriction gene and methylase gene
are cloned and expressed.

<>

<1>Benner, J.S., Rees, P.
<2>Method for producing the HpaI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5298404
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the HpaI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Haemophilus
parainfluenzae into a host whereby the restriction gene is expressed; 2) fermenting the host
which contains the vector encoding and expressing the HpaI restriction endonuclease, and 3)
purifying the HpaI restriction endonuclease from the fermented host which contains the vector
encoding and expressing the HpaI restriction endonuclease activity.

<>

<1>Benner, J.S., Rees, P.A.
<2>Method of producing the HincII restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5015581
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the HincII restriction
endonuclease by (1) introducing the restriction endonuclease gene from Haemophilus influenzae
Re into a host whereby the restriction gene is expressed ; (2) fermenting the host which
contains the vector encoding and expressing the HincII restriction endonuclease, and (3)
purifying HincII restriction endonuclease from the fermented host which contains the vector
encoding and expressing the HincII restriction endonuclease activity.

<>

<1>Benner, J.S., Rees, P.A.
<2>Method for producing the HincII restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0388212 A
<5>
<6>1995
<7>The present invention is directed to a method for cloning and producing the HincII restriction
endonuclease by 1) introducing the restriction endonuclease gene from Haemophilus influenzae
Rc into a host whereby the restriction gene is expressed; 2) fermenting the host which
contains the vector encoding and expressing the HincII restriction endonuclease, and 3)
purifying the HincII restriction endonuclease from the fermented host which contains the
vector encoding and expressing the HincII restriction endonuclease activity.

<>

<1>Benner, J.S., Rees, P.A.
<2>Method for producing the HincII restriction endonuclease and methylase.
<3>Japanese Patent Office
<4>JP 2672683
<5>
<6>1997
<7>
<>

<1>Bennett, S.P.
<2>Restriction enzymes with asymmetrical recognition sites.
<3>Diss. Abstr.
<4>49
<5>722B
<6>1988
<7>The class II restriction endonucleases, CauI and CauII, recognize the DNA
sequences, 5'-GG(A/T)CC-3' and 5' -CC(C/G)GG-3', respectively.  Both of these
enzymes have been purified to homogeneity.  Molecular weight measurements
indicate that the active form of the CauI enzyme is a dimer of identical
protein subunits (of Mr 2x27,500).  In contrast, CauII appears to be a
heterologous dimer made up of different protein subunits (one of Mr 31,500 and
one of Mr 29,000).  Kinetic studies using the replicative DNA (RFI) of phage
PhiX174 as substrate, suggest that CauI and CauII cleave duplex DNA by a
concerted mechanism, in which both strands of the DNA are cut within the
lifetimes of a single enzyme-DNA complex.  Each endonuclease shows the same
activity towards its recognition site on both the supercoiled and relaxed forms
of PhiX174 RFDNA.  This implies that the unwinding of the DNA plays no
significant role in either the CauI or CauII cleavage reaction.  In standard
reactions, CauI and CauII display high specificities towards their recognition
sequences.  However, in the presence of dimethyl sulphoxide and at high pH,
each enzyme cleaves DNA at sites that differ from its recognition sequence by
one nucleotide.  The CauII enzyme cleaves DNA to generate fragments of DNA that
have at their 5' terminal, single nucleotide extensions of either 5' C or 5'G.
Ligation of these fragments generates cytosine:cytosine in duplex DNA.

<>

<1>Bennett, S.P.
<2>Restriction enzymes with asymmetrical recognition sites.
<3>Ph.D. Thesis, University of Bristol, UK
<4>
<5>1-196
<6>1987
<7>The class-II restriction endonucleases, CauI and CauII, recognize the DNA
sequences, 5'-GG(A/T)CC-3' and 5'-CC(C/G)GG-3', respectively.  Both of these
enzymes have been purified to homogeneity.  Molecular weight measurements
indicate that the active form of the CauI enzyme is a dimer of identical
protein subunits (of Mr 2 x 27500).  In contrast, CauII appears to be a
heterologous dimer made up of different protein subunits (one of Mr 31500 and
one of Mr 29000).  Kinetic studies using the replicative DNA (RFI) of phage
PhiX174 as substrate, suggest that CauI and CauII cleave duplex DNA by a
concerted mechanism, in which both strands of the DNA are cut within the
lifetime of a single enzyme-DNA complex.  Each endonuclease shows the same
activity towards its recognition site on both the supercoiled and relaxed forms
of PhiX174 RFDNA.  This implies that the unwinding of the DNA plays no
significant role in either the CauI or CauII cleavage reaction.  In standard
reactions, CauI and CauII display high specificities towards their recognition
sequences.  However, in the presence of dimethyl sulphoxide and at high pH,
each enzyme cleaves DNA at sites that differ from its recognition sequence by
one nucleotide.  The CauII enzyme cleaves DNA to generate fragments that have
at their 5' termini, single nucleotide extensions of either 5'C or 5'G.
Ligation of these fragments generates cytosine:cytosine in duplex DNA.

<>

<1>Bennett, S.P., Halford, S.E.
<2>Recognition of DNA by Type II restriction enzymes.
<3>Curr. Top. Cell. Regul.
<4>30
<5>57-104
<6>1989
<7>A review

<>

<1>Bennett, S.P., Halford, S.E.
<2>Mechanism and specificity of two restriction enzymes, CauI and CauII, that recognize asymmetrical DNA sequences.
<3>DNA-ligand interactions: from drugs to proteins., Plenum Press, Guschlbauer, W., Saenger, W., New York
<4>
<5>239-250
<6>1987
<7>Type II restriction endonucleases are enzymes that recognize specific nucleotide sequences on
duplex DNA and cleave both strands of the duplex at fixed locations relative to their
recognition sites.  The only co-factor that they need is Mg2+, while type I and type III
restriction enzymes also need ATP and S-adenosyl methionine for maximal activity.  Type I and
type III enzymes also differ from type II by cleaving DNA at variable distances from their
recognition sites.  Hence, the study of type II restriction enzymes was initially motivated by
their unique applications in the analysis of DNA and in the construction of recombinant DNA
molecules.  However, these enzymes also provide examples of DNA-protein interactions whose
mechanisms are amenable to molecular analysis.

<>

<1>Bennett, S.P., Halford, S.E.
<2>Cytosine: cytosine formed in duplex DNA by the ligation of NciI- or CauII-cleaved DNA.
<3>Biochem. Soc. Trans.
<4>14
<5>259
<6>1986
<7>The type II restriction endonucleases, CauII and its isoschizomer NciI, recognize the sequence
5'-C-C/-G-G-G-3' 3'-G-G-C/-C-C-5' on duplex DNA and cleave each strand at the sites marked
by arrows.  Hence these enzymes generate fragments of double-stranded DNA that have single
nucleotide 5' extensions of either G or C.  The DNA ligase encoded by bacteriophage T4 can
ligate DNA molecules that have either cohesive termini (due to single-strand extensions) or
blunt ends.  The efficiency of blunt-end ligations is usually low but can be increased by
adding polyethylene glycol to the reactions.

<>

<1>Benomar, N., Abriouel, H., Lee, H., Cho, G.S., Huch, M., Pulido, R.P., Holzapfel, W.H., Galvez, A., Franz, C.M.
<2>Genome Sequence of Weissella thailandensis fsh4-2.
<3>J. Bacteriol.
<4>193
<5>5868
<6>2011
<7>Weissella thailandensis fsh4-2 is a heterofermentative lactic acid bacterium isolated from the
Korean fermented seafood condiment jeotkal.
Here we report the draft genome sequence of W. thailandensis fsh4-2 (1,651
genes, 1,436 encoding known proteins, 183 encoding unknown proteins, 32
RNA genes), which consists of 50 large contigs of >100 bp.

<>

<1>Bens, C.C., Voss, A., Klaassen, C.H.
<2>Presence of a novel DNA methylation enzyme in methicillin-resistant Staphylococcus aureus isolates associated with pig farming leads to  uninterpretable results in standard pulsed-field gel electrophoresis  analysis.
<3>J. Clin. Microbiol.
<4>44
<5>1875-1876
<6>2006
<7>Genomic DNA from methicillin-resistant Staphylococcus aureus isolates recovered from pigs and
their caretakers proved resistant to SmaI
digestion, leading to uninterpretable results in standard pulsed-field gel
electrophoresis. This is the result of a yet unknown
restriction/methylation system in the genus Staphylococcus with the
recognition sequence CCNGG.

<>

<1>Benson, M.A., Ohneck, E.A., Ryan, C., Alonzo, F. III, Smith, H., Narechania, A., Kolokotronis, S.O., Satola, S.W., Uhlemann, A.C., Sebra, R., Deikus, G., Shopsin, B., Planet, P.J., Torres, V.J.
<2>Evolution of hypervirulence by a MRSA clone through acquisition of a transposable element.
<3>Mol. Microbiol.
<4>93
<5>664-681
<6>2014
<7>Staphylococcus aureus has evolved as a pathogen that causes a range of diseases
in humans. There are two dominant modes of evolution thought to explain most of
the virulence differences between strains. First, virulence genes may be acquired
from other organisms. Second, mutations may cause changes in the regulation and
expression of genes. Here we describe an evolutionary event in which
transposition of an IS element has a direct impact on virulence gene regulation
resulting in hypervirulence. Whole-genome analysis of a methicillin-resistant S.
aureus (MRSA) strain USA500 revealed acquisition of a transposable element
(IS256) that is absent from close relatives of this strain. Of the multiple
copies of IS256 found in the USA500 genome, one was inserted in the promoter
sequence of repressor of toxins (Rot), a master transcriptional regulator
responsible for the expression of virulence factors in S. aureus. We show that
insertion into the rot promoter by IS256 results in the derepression of cytotoxin
expression and increased virulence. Taken together, this work provides new
insight into evolutionary strategies by which S. aureus is able to modify its
virulence properties and demonstrates a novel mechanism by which horizontal gene
transfer directly impacts virulence through altering toxin regulation.

<>

<1>Bentley, S.D. et al.
<2>Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2).
<3>Nature
<4>417
<5>141-147
<6>2002
<7>Streptomyces coelicolor is a representative of the group of soil-dwelling, filamentous
bacteria responsible for producing most natural antibiotics used in human and veterinary
medicine.  Here we report the 8,667,507 base pair linear chromosome of this organism,
containing the largest number of genes so far discovered in a bacterium.  The 7,825 predicted
genes include more than 20 clusters coding for known or predicted secondary metabolites.  The
genome contains an unprecedented proportion of regulatory genes, predominantly those likely to
be involved in responses to external stimuli and stresses, and many duplicated gene sets that
may represent 'tissue-specific' isoforms operating in different phases of colonial
development, a unique situation for a bacterium.  An ancient synteny was revealed between the
central 'core' of the chromosome and the whole chromosome of pathogens Mycobacterium
tuberculosis and Corynebacterium diphtheriae.  The genome sequence will greatly increase our
understanding of microbial life in the soil as well as aiding the generation of new drug
candidates by genetic engineering.

<>

<1>Bentley, S.D. et al.
<2>Sequencing and analysis of the genome of the Whipple's disease bacterium Tropheryma whipplei.
<3>Lancet
<4>361
<5>637-644
<6>2003
<7>BACKGROUND: Whipple's disease is a rare multisystem chronic infection, involving the
intestinal tract as well as various other organs. The
causative agent, Tropheryma whipplei, is a Gram-positive bacterium about
which little is known. Our aim was to investigate the biology of this
organism by generating and analysing the complete DNA sequence of its
genome. METHODS: We isolated and propagated T whipplei strain TW08/27 from
the cerebrospinal fluid of a patient diagnosed with Whipple's disease. We
generated the complete sequence of the genome by the whole genome shotgun
method, and analysed it with a combination of automatic and manual
bioinformatic techniques. FINDINGS: Sequencing revealed a condensed 925938
bp genome with a lack of key biosynthetic pathways and a reduced capacity
for energy metabolism. A family of large surface proteins was identified,
some associated with large amounts of non-coding repetitive DNA, and an
unexpected degree of sequence variation. INTERPRETATION: The genome
reduction and lack of metabolic capabilities point to a host-restricted
lifestyle for the organism. The sequence variation indicates both known
and novel mechanisms for the elaboration and variation of surface
structures, and suggests that immune evasion and host interaction play an
important part in the lifestyle of this persistent bacterial pathogen.

<>

<1>Bentzon-Tilia, M., Riemann, L., Gram, L.
<2>Draft Genome Sequence of Hoeflea sp. Strain BAL378, a Potential Producer of Bioactive Compounds.
<3>Genome Announcements
<4>2
<5>e01213-14
<6>2014
<7>Some phytoplankton-associated marine bacteria produce bioactive compounds. Members of the
genus Hoeflea may be examples of such bacteria; however, data
describing their metabolisms are scarce. Here, we report the draft genome
sequence of Hoeflea sp. strain BAL378, a putative producer of bacteriocins,
polyketides, and auxins, as demonstrated by genome mining.

<>

<1>Benzinger, R.
<2>Restriction of infectious bacteriophage fd DNA's and an assay for in vitro host-controlled restriction and modification.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>59
<5>1294-1299
<6>1968
<7>Arber has shown that bacteriophage fd, which contains single-stranded circular
DNA (Marvin and Schaller) is restricted by E. coli B and E. coli (P1)+.  This
report presents detailed data on the restriction of both the infectious,
single-stranded fd-phage DNA and the infectious, double-stranded fd-replicative
form (RF) DNA.  It also describes an infectivity assay for in vitro
host-controlled restriction and modification.  This assay has been used to find
restriction enzymes in fractionated extracts of E. coli.

<>

<1>Bera, A.K., Das, B., Chattopadhyay, S., Dasgupta, C.
<2>Refolding of denatured restriction endonucleases with ribosomal preparations from Methanosarcina barkeri.
<3>Biochem. Mol. Biol. Int.
<4>32
<5>315-323
<6>1994
<7>Two restriction endonucleases, EcoRI and HindIII were denatured by guanidine HCl or by storing
at room temperature (28oC) for several days. The activity of these enzymes could be restored
by incubating with ribosomal preparations from an archaebacterium Methanosarcina barkeri.
These results hint at a possible role of ribosomal preparations in folding polypeptides and
could be useful in working with restriction enzymes in experiments on genetic engineering and
molecular biology.

<>

<1>Berdis, A.J., Lee, I., Coward, J.K., Stephens, C., Wright, R., Shapiro, L., Benkovic, S.J.
<2>A cell cycle-regulated adenine DNA methyltransferase from Caulobacter crescentus processively methylates GANTC sites on hemimethylated DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>2874-2879
<6>1998
<7>The kinetic properties of an adenine DNA methyltransferase involved in cell cycle regulation
of Caulobacter crescentus have been elucidated by using defined unmethylated or hemimethylated
DNA substrates.  Catalytic efficiency is significantly enhanced with a DNAHM substrate.
Biphasic kinetic behavior during methyl incorporation is observed when unmethylated or DNAHM
substrates are used, indicating that a step after chemistry limits enzyme turnover and is most
likely the release of enzyme from methylated DNA product.  The enzyme is thermally inactivated
at 30oC within 20 min; this process substantially decreased in the presence of saturating
concentrations of DNAHM, suggesting that the enzyme preferentially binds DNA before
S-adenosylmethionine.  The activity of the enzyme shows an unusual sensitivity to salt levels,
apparently dissociating more rapidly from methylated DNA product as the salt level is
decreased.  The enzyme acts processively during methylation of specific DNA sequences,
indicating a preferred order of product release in which S-adenosylhomocysteine is released
from enzyme before fully methylated DNA.  The kinetic behavior and activity of the enzyme are
consistent with the temporal constraints during the cell cycle-regulated methylation of newly
replicated chromosomal DNA.

<>

<1>Berent, L.M., Messick, J.B.
<2>Physical Map and Genome Sequencing Survey of Mycoplasma haemofelis (Haemobartonella felis).
<3>Infect. Immun.
<4>71
<5>3657-3662
<6>2003
<7>Mycoplasma haemofelis is an uncultivable red-cell pathogen of cats.
Isolated M. haemofelis DNA was used to create a bacterial artificial
chromosome library and physical map. Random sequencing of this material
revealed 75 genes that had not been previously reported for M. haemofelis
or any other hemotrophic mycoplasma.

<>

<1>Beres, S.B., Richter, E.W., Nagiec, M.J., Sumby, P., Porcella, S.F., DeLeo, F.R., Musser, J.M.
<2>Molecular genetic anatomy of inter- and intraserotype variation in the human bacterial pathogen group A Streptococcus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>7059-7064
<6>2006
<7>In recent years we have studied the relationship between strain genotypes and patient
phenotypes in group A Streptococcus (GAS), a model human
bacterial pathogen that causes extensive morbidity and mortality
worldwide. We have concentrated our efforts on serotype M3 organisms
because these strains are common causes of pharyngeal and invasive
infections, produce unusually severe invasive infections, and can exhibit
epidemic behavior. Our studies have been hindered by the lack of
genome-scale phylogenies of multiple GAS strains and whole-genome
sequences of multiple serotype M3 strains recovered from individuals with
defined clinical phenotypes. To remove some of these impediments, we
sequenced to closure the genome of four additional GAS strains and
conducted comparative genomic resequencing of 12 contemporary serotype M3
strains representing distinct genotypes and phenotypes. Serotype M3
strains are a single phylogenetic lineage. Strains from asymptomatic
throat carriers were significantly less virulent for mice than
sterile-site isolates and evolved to a less virulent phenotype by multiple
genetic pathways. Strain persistence or extinction between epidemics was
strongly associated with presence or absence, respectively, of the
prophage encoding streptococcal pyrogenic exotoxin A. A serotype M3 clone
significantly underrepresented among necrotizing fasciitis cases has a
unique frameshift mutation that truncates MtsR, a transcriptional
regulator controlling expression of genes encoding iron-acquisition
proteins. Expression microarray analysis of this clone confirmed
significant alteration in expression of genes encoding iron metabolism
proteins. Our analysis provided unprecedented detail about the molecular
anatomy of bacterial strain genotype-patient phenotype relationships.

<>

<1>Beres, S.B., Sylva, G.L., Barbian, K.D., Lei, B., Hoff, J.S., Mammarella, N.D., Liu, M.-Y., Smoot, J.C., Porcella, S.F., Parkins, L.D., Campbell, D.S., Smith, T.M., McCormick, J.K., Leung, D.Y.M., Schlievert, P.M., Musser, J.M.
<2>Genome sequence of a serotype M3 strain of group A Streptococcus: Phage-encoded toxins, the high-virulence phenotype, and clone emergence.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>10078-10083
<6>2002
<7>Genome sequences are available for many bacterial strains, but there has been little progress
in using these data to understand the molecular basis of pathogen emergence and differences in
strain virulence. Serotype M3 strains of group A Streptococcus (GAS) are a common cause of
severe invasive infections with unusually high rates of morbidity and mortality. To gain
insight into the molecular basis of this high-virulence phenotype, we sequenced the genome of
strain MGAS315, an organism isolated from a patient with streptococcal toxic shock syndrome.
The genome is composed of 1,900,521 bp, and it shares approximately 1.7 Mb of related genetic
material with genomes of serotype M1 and M18 strains. Phage-like elements account for the
great majority of variation in gene content relative to the sequenced M1 and M18 strains.
Recombination produces chimeric phages and strains with previously uncharacterized arrays of
virulence factor genes. Strain MGAS315 has phage genes that encode proteins likely to
contribute to pathogenesis, such as streptococcal pyrogenic exotoxin A (SpeA) and SpeK,
streptococcal superantigen (SSA), and a previously uncharacterized phospholipase A(2)
(designated Sla).  Infected humans had anti-SpeK, -SSA, and -Sla antibodies, indicating that
these GAS proteins are made in vivo.  SpeK and SSA were pyrogenic and toxic for rabbits.
Serotype M3 strains with the phage-encoded speK and sla genes increased dramatically in
frequency late in the 20th century, commensurate with the rise in invasive disease caused by
M3 organisms. Taken together, the results show that phage-mediated recombination has played a
critical role in the emergence of a new, unusually virulent clone of serotype M3 GAS.

<>

<1>Berg, D.E., Hoffman, P.S., Appelmelk, B.J., Kusters, J.G.
<2>The Helicobacter pylori genome sequence: genetic factors for long life in the gastric mucosa.
<3>Trends Microbiol.
<4>5
<5>468-474
<6>1997
<7>The complete DNA sequence of the chromosome of a strain of Helicobacter pylori was released in
mid-1997 by The Institute for Genomic Research, a long-awaited and warmly welcomed event that
should greatly stimulate research on this pathogen and benefit public health.  Infection by H,
pylori occurs in more than half of all people worldwide and is a major cause of severe
gastritis and peptic ulcer disease and an early risk factor for gastric cancer, although most
infections are asymptomatic and some might even be beneficial.  H. pylori is Gram-negative,
fastidious in its nutritional requirements and microaerophilic: it chronically infects human
gastric epithelial cell surfaces and the overlying mucin layer.  Although some infections are
cleared spontaneously after a brief acute phase, many last for years or decades, and it is
these infections that are most often implicated in overt disease.

<>

<1>Berg-Miller, M.E., Antonopoulos, D.A., Rincon, M.T., Band, M., Bari, A., Akraiko, T., Hernandez, A., Thimmapuram, J., Henrissat, B., Coutinho, P.M., Borovok, I., Jindou, S., Lamed, R., Flint, H.J., Bayer, E.A., White, B.A.
<2>Diversity and strain specificity of plant cell wall degrading enzymes revealed by the draft genome of Ruminococcus flavefaciens FD-1.
<3>PLoS ONE
<4>4
<5>E6650
<6>2009
<7>BACKGROUND: Ruminococcus flavefaciens is a predominant cellulolytic rumen bacterium, which
forms a multi-enzyme cellulosome complex that could play an integral role in the ability of
this bacterium to degrade plant cell wall polysaccharides. Identifying the major enzyme types
involved in plant cell wall degradation is essential for gaining a better understanding of the
cellulolytic capabilities of this organism as well as highlighting potential enzymes for
application in improvement of livestock nutrition and for conversion of cellulosic biomass to
liquid fuels. METHODOLOGY/PRINCIPAL FINDINGS: The R. flavefaciens FD-1 genome was sequenced to
29x-coverage, based on pulsed-field gel electrophoresis estimates (4.4 Mb), and assembled into
119 contigs providing 4,576,399 bp of unique sequence. As much as 87.1% of the genome encodes
ORFs, tRNA, rRNAs, or repeats. The GC content was calculated at 45%. A total of 4,339 ORFs was
detected with an average gene length of 918 bp. The cellulosome model for R. flavefaciens was
further refined by sequence analysis, with at least 225 dockerin-containing ORFs, including
previously characterized cohesin-containing scaffoldin molecules. These dockerin-containing
ORFs encode a variety of catalytic modules including glycoside hydrolases (GHs),
polysaccharide lyases, and carbohydrate esterases. Additionally, 56 ORFs encode proteins that
contain carbohydrate-binding modules (CBMs). Functional microarray analysis of the genome
revealed that 56 of the cellulosome-associated ORFs were up-regulated, 14 were down-regulated,
135 were unaffected, when R. flavefaciens FD-1 was grown on cellulose versus cellobiose. Three
multi-modular xylanases (ORF01222, ORF03896, and ORF01315) exhibited the highest levels of
up-regulation. CONCLUSIONS/SIGNIFICANCE: The genomic evidence indicates that R. flavefaciens
FD-1 has the largest known number of fiber-degrading enzymes likely to be arranged in a
cellulosome architecture. Functional analysis of the genome has revealed that the growth
substrate drives expression of enzymes predicted to be involved in carbohydrate metabolism as
well as expression and assembly of key cellulosomal enzyme components.

<>

<1>Berg-Miller, M.E., Yeoman, C.J., Chia, N., Tringe, S.G., Angly, F.E., Edwards, R.A., Flint, H.J., Lamed, R., Bayer, E.A., White, B.A.
<2>Phage-bacteria relationships and CRISPR elements revealed by a metagenomic survey of the rumen microbiome.
<3>Environ. Microbiol.
<4>14
<5>207-227
<6>2012
<7>Viruses are the most abundant biological entities on the planet and play an
important role in balancing microbes within an ecosystem and facilitating
horizontal gene transfer. Although bacteriophages are abundant in rumen
environments, little is known about the types of viruses present or their
interaction with the rumen microbiome. We undertook random pyrosequencing of
virus-enriched metagenomes (viromes) isolated from bovine rumen fluid and
analysed the resulting data using comparative metagenomics. A high level of
diversity was observed with up to 28,000 different viral genotypes obtained from
each environment. The majority (~78%) of sequences did not match any previously
described virus. Prophages outnumbered lytic phages approximately 2:1 with the
most abundant bacteriophage and prophage types being associated with members of
the dominant rumen phyla (Firmicutes and Proteobacteria). Metabolic profiling
based on SEED subsystems revealed an enrichment of sequences with putative
functional roles in DNA and protein metabolism, but a surprisingly low proportion
of sequences assigned to carbohydrate and amino acid metabolism. We expanded our
analysis to include previously described metagenomic data and 14 reference
genomes. Clustered regularly interspaced short palindromic repeats (CRISPR) were
detected in most of the microbial genomes, suggesting previous interactions
between viral and microbial communities.

<>

<1>Berge, T., Ellis, D.J., Dryden, D.T.F., Edwardson, J.M., Henderson, R.M.
<2>Translocation-independent dimerization of the EcoKI endonuclease visualized by atomic force microscopy.
<3>Biophys. J.
<4>79
<5>479-484
<6>2000
<7>Bacterial type I restriction/modification systems are capable of performing multiple actions
in response to the methylation pattern on
their DNA recognition sequences. The enzymes making up these systems
serve to protect the bacterial cells against viral infection by binding
to their recognition sequences on the invading DNA and degrading it
after extensive ATP-driven translocation, DNA cleavage has been thought
to occur as the result of a collision between two translocating enzyme
complexes. Using atomic force microscopy (AFM), we show here that EcoKI
dimerizes rapidly when bound to a plasmid containing two recognition
sites for the enzyme. Dimerization proceeds in the absence of ATP and
is also seen with an EcoKI mutant (K477R) that is unable to translocate
DNA. Only monomers are seen when the enzyme complex binds to a plasmid
containing a single recognition site. Based on our results, we propose
that the binding of EcoKI to specific DNA target sequences is
accompanied by a conformational change that leads rapidly to
dimerization. This event is followed by ATP-dependent translocation and
cleavage of the DNA.

<>

<1>Bergemann, A.D., Whitley, J.C., Finch, L.R.
<2>Taxonomic significance of differences in DNA methylation within the Mycoplasma mycoides cluster' detected with restriction endonucleases MboI and DpnI.
<3>Lett. Appl. Microbiol.
<4>11
<5>48-51
<6>1990
<7>DNA from 22 strains belonging to the Mycoplasma mycoides cluster was tested for
methylation of adenine in GATC sequences by its resistance or susceptibility to
digestion by the restriction endonucleases MboI, which is inhibited by the
methylation, or DpnI, which requires the methylation.  Strains of bovine
serogroup 7, M. mycoides subsp. mycoides SC, and some M. mycoides subsp. capri
strains plus M. capricolum strain Cal Kid lacked the methylation, whereas other
strains of M. mycoides subsp. capri and of M. capricolum, plus the F38-like and
M. mycoides subsp. mycoides LC strains, possessed it.  We conclude that this
simple test could provide a valuable criterion in identifying these
mycoplasmas.

<>

<1>Berger, S., Alauzet, C., Aissa, N., Henard, S., Rabaud, C., Bonnet, R., Lozniewski, A.
<2>Characterization of a new blaOXA-48-carrying Plasmid in Enterobacteriaceae.
<3>Antimicrob. Agents Chemother.
<4>57
<5>4064-4067
<6>2013
<7>In this work we characterized a new 160-kb blaOXA-48-harboring IncL/M-type
plasmid, isolated from a Klebsiella pneumoniae strain from France. Moreover, we
report the transfer of a 60-kb OXA-48 encoding plasmid from Klebsiella pneumoniae
to other Enterobacteriaceae in two patients.

<>

<1>Berger, S.L.
<2>Expanding the potential of restriction endonucleases: Use of hapaxoterministic enzymes.
<3>Anal. Biochem.
<4>222
<5>1-8
<6>1994
<7>A new class of restriction endonucleases called hapoxoterministic enzymes, hapaxomers for
short, has been defined. A hapaxomer cleaves DNA outside the recognition site or within an
interrupted "palindrome" at bases which are not specified producing fragments with asymmetric,
staggered ends. Such termini are unique; in the general case, the protrusions of a fragment
obtained with the aid of a hapaxomer cannot self-hybridize, nor can the fragment be ligated to
the vast majority of other fragments produced by the same enzyme. When the fragments generated
by a hapaxoterministic enzyme are mixed, they can reassemble in only one configuration--that
of the original structure from which they were derived. Hapaxomers, then, are characterized
not by their recognition sites, which may be symmetric or asymmetric, but by their cleavage
sites. The ability to reunite once-contiguous fragments efficiently means that hapaxomers
cleaving DNA at many locations are virtually equivalent to restriction enzymes cutting at
unique sites. These properties can be exploited for applications such as site-specific
mutagenesis or the isolation of large intact DNA fragments.

<>

<1>Berger, S.L.
<2>Gene modification with hapaxoterministic restriction enzymes.
<3>Methods Mol. Biol.
<4>160
<5>443-458
<6>2001
<7>Subcloning can be extraordinarily easy or extremely difficult.  The simplest case requires
cleavage of the desired fragment from the DNA in which it is located with one or two
restriction enzymes, cleavage of the plasmid that will serve as the ultimate recipient with
the same enzymes, and ligation of the fragment to the plasmid.  In a somewhat more complicated
version of simple cloning, the investigator uses the polymerase chain reaction to amplify the
DNA to be subcloned.  In this case, restriction sites can be incorporated into the primers
used for PCR, generating products with the desired restriction sites located near the ends of
the newly synthesized DNA.  After cleavage of those sites, the fragment is ready for insertion
into an appropriately cut vector.  The major disadvantage of the method is the need for
sequencing the amplified material, because even the best of the thermostable polymerases makes
mistakes that might render the DNA useless for the anticipated application.

<>

<1>Bergerat, A., Guschlbauer, W.
<2>The double role of methyl donor and allosteric effector of S-adenosyl-methionine for Dam methylase of E. coli.
<3>Nucleic Acids Res.
<4>18
<5>4369-4375
<6>1990
<7>The turnover of DNA-adenine-methylase of E. coli strongly decreases when the temperature is
lowered. This has allowed us to study the binding of Dam methylase on 14 bp DNA fragments at
0C by gel retardation in the presence of Ado-Met, but without methylation taking place. The
enzyme can bind nonspecific DNA with low affinity. Binding to the specific sequence occurs in
the absence of S-adenosylmethionine (Ado-Met), but is activated by the presence of the methyl
donor. The two competitive inhibitors of Ado-Met, sinefungin and S-adenosyl-homocysteine, can
neither activate this binding to DNA by themselves, nor inhibit this activation by Ado-Met.
This suggests that Ado-Met could bind to Dam methylase in two different environments. In one
of them, it could play the role of an allosteric effector which would reinforce the affinity
of the enzyme for the GATC site. The analogues can not compete for such binding. In the other
environment Ado-Met would be in the catalytic site and could be exchanged by its analogues. We
have also visualized conformational changes in Dam methylases induced by the simultaneous
binding of Ado-Met and the specific target sequence of the enzyme, by an anomaly of migration
and partial resistance to proteolytic treatment of the ternary complex Ado-Met/Dam
methylase/GATC.

<>

<1>Bergerat, A., Guschlbauer, W., Fazakerley, G.V.
<2>Allosteric and catalytic binding of S-adenosylmethionine to Escherichia coli DNA adenine methyltransferase monitored by 3H NMR.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>88
<5>6394-6397
<6>1991
<7>Adenine methylation of GATC sequences in DNA is carried out by the DNA adenine
methyltransferase with the methyl group source being the cofactor
S-adenosylmethionine.  We report 3H NMR studies on the interaction of DNA
adenine methyltransferase with S-adenosylmethionine and the reaction when the
ternary complex is formed with an oligonucleotide containing a GATC site.  The
methylation reaction was also studied in the presence of a competitive
inhibitor and this showed two successive stages involved in the methylation and
tw sites of binding for S-adenosylmethionine.

<>

<1>Bergerat, A., Kriebardis, A., Guschlbauer, W.
<2>Preferential site-specific hemimethylation of GATC sites in pBR322 DNA by dam methyltransferase from Escherichia coli.
<3>J. Biol. Chem.
<4>264
<5>4064-4070
<6>1989
<7>The methylation pattern of the 22 GATC sites of pBR322 (dam-) by Dam methyltransferase from
Escherichia coli has been studied. Preferential hemimethylation took place at positions 3042
and 349. It was found that these preferential methylations were the same in supercoiled
circular and linear DNAs. The flanking regions of these preferentially methylated sites
contain three GC pairs on one side and two AT pairs and one GC pair on the other. This
preferential methylation was confirmed on a 126-base pair oligonucleotide containing two GATC
sites with different flanking sequences. The next sites methylated were, in both cases, the
first GATC site on the AT-rich side, although the orientation was different. The rapid
methylation of a second and third neighboring GATC site on the same plasmid suggests a
processive mechanism. The implications of the orientation of hemimethylation are discussed in
the context of the recognition of a palindromic target site by a monomeric DNA-binding
protein.

<>

<1>Bergez, P., Doignon, F., Crouzet, M.
<2>The sequence of a 44,420 bp fragment located on the left arm of chromosome XIV from Saccharomyces cerevisiae.
<3>Yeast
<4>11
<5>967-974
<6>1995
<7>We have determined the complete nucleotide sequence of a 44 420 bp DNA fragment from
chromosome XIV of Saccharomyces cerevisiae. The sequence data revealed 23 open reading frames
(ORFs) larger than 300 bp, covering 73.5% of the sequence. The ORFs N2418, N2428, N2441, N2474
and N2480 correspond to previously sequenced S. cerevisiae genes coding respectively for the
mitochondrial import protein Mas5, the nucleolar protein Nop2, the outer mitochondrial
membrane porin Por1, the cytochrome c oxidase polypeptide VA precursor CoxA and the yeast
protein tyrosine phosphatase Msg5. Translation products of three other ORFs N2406, N2411 and
N2430 exhibit similarity to previously known S. cerevisiae proteins: the ribosomal protein
YL9A, the protein Nca3 involved in the mitochondrial expression of subunits 6 and 8 of the ATP
synthase and actin; in addition N2505 presents strong similarity to an ORF of chromosome IX.
The predicted protein products of ORFs N2417 and N2403 present similarities with domains from
proteins of other organisms: the Candida maltosa cycloheximide-resistance protein, the human
interleukin enhancer-binding factor (ILF-2). The 12 remaining ORFs show no significant
similarity to known proteins. In addition, we have detected a DNA region very similar to the
yeast transposon Ty 1-15 of which insertion has disrupted a tRNA(Asp) gene.

<>

<1>Berglund, E.C., Frank, A.C., Calteau, A., Vinnere, P.O., Granberg, F., Eriksson, A.S., Naslund, K., Holmberg, M., Lindroos, H., Andersson, S.G.
<2>Run-off Replication of Host-Adaptability Genes is Associated with Gene Transfer Agents in the Genome of Mouse-Infecting Bartonella grahamii.
<3>PLoS Genet.
<4>5
<5>e1000546
<6>2009
<7>The genus Bartonella comprises facultative intracellular bacteria adapted to mammals,
including previously recognized and emerging human pathogens. We report the 2,341,328 bp
genome sequence of Bartonella grahamii, one of the most prevalent Bartonella species in wild
rodents. Comparative genomics revealed that rodent-associated Bartonella species have higher
copy numbers of genes for putative host-adaptability factors than the related human-specific
pathogens. Many of these gene clusters are located in a highly dynamic region of 461 kb. Using
hybridization to a microarray designed for the B. grahamii genome, we observed a massive,
putatively phage-derived run-off replication of this region. We also identified a novel gene
transfer agent, which packages the bacterial genome, with an over-representation of the
amplified DNA, in 14 kb pieces. This is the first observation associating the products of
run-off replication with a gene transfer agent. Because of the high concentration of gene
clusters for host-adaptation proteins in the amplified region, and since the genes encoding
the gene transfer agent and the phage origin are well conserved in Bartonella, we hypothesize
that these systems are driven by selection. We propose that the coupling of run-off
replication with gene transfer agents promotes diversification and rapid spread of
host-adaptability factors, facilitating host shifts in Bartonella.

<>

<1>Bergman, Y., Mostoslavsky, R.
<2>DNA demethylation: Turning genes on.
<3>Biol. Chem.
<4>379
<5>401-407
<6>1998
<7>The regulation of eukaryotic gene expression is a complicated process involving the
interaction of a large number of transacting factors with specific cis-regulatory elements.
DNA methylation plays a role in this scheme by acting in cis to modulate protein-DNA
interactions.  Several lines of evidence indicate that methylation serves to silence
transcription, mainly through indirect mechanisms involving the assembly of repressive
nucleoprotein complexes.  DNA demethylation is mostly an active enzymatic process, controlled
by cis regulatory elements which provide binding sites for trans demethylation factors.  In
the immune system DNA methylation plays multiple roles, such as regulating both gene
expression and gene rearrangement.

<>

<1>Berka, R., Clausen, I.G.
<2>Methods for monitoring multiple gene expression.
<3>International Patent Office
<4>WO 0229113 A
<5>
<6>2002
<7>The present invention relates to methods for monitoring differential expression of a plurality
of genes in a first Bacillus cell relative to expression of the same genes in one or more
second Bacillus cells using microarrays containing Bacillus genomic sequenced tags.  The
present invention also relates to computer readable media and computer-based systems.  The
present invention further relates to substrates containing an array of Bacillus licheniformis
or Bacillus clausii GSTs.

<>

<1>Berka, R., Clausen, I.G., Bolotine, A., Sorokine, A., Lapidus, A., Joergensen, S.T.
<2>Methods for disrupting a gene involved in PGA synthesis in Bacillus licheniformis.
<3>European Patent Office
<4>EP 2072526 A
<5>
<6>2009
<7>The present invention relates to methods for monitoring differential expression of a plurality
of genes in a first Bacillus cell relative to expression of the same genes in one or more
second Bacillus cells using microarrays containing Bacillus genomic sequenced tags.  The
present invention also relates to computer readable media and computer-based systems.  The
present invention further relates to substrates containing an array of Bacillus licheniformis
or Bacillus clausii GSTs.

<>

<1>Berka, R., Rey, M., Ramaiya, P.
<2>Bacillus licheniformis chromosome.
<3>International Patent Office
<4>WO 2008066931 A
<5>
<6>2008
<7>The present invention relates to an isolated polynucleotide of the complete chromosome of
Bacillus licheniformis SJ1904 (ATCC PTA-7992).  The present invention also relates to isolated
polynucleotides of the chromosome of Bacillus licheniformis SJ1904 that encode biologically
active substances and to nucleic acid constructs, vectors, and host cells comprising the
polynucleotides as well as methods for producing biologically active substances encoded by the
polynucleotides and to methods of using the iosolated polynucleotides of the complete
chromosome of Bacillus licheniformis SJ1904.

<>

<1>Berka, R., Rey, M., Ramaiya, P.
<2>Bacillus Licheniformis chromosome.
<3>European Patent Office
<4>EP 2210898 A
<5>
<6>2010
<7>The present invention relates to an isolated polynucleotide of Bacillus licheniformis SJ1904
(ATCC OPTA-7992) that encodes a biologically active substance and to nucleic acid constructs,
vectors, and host cells comprising the polynucleotides as well as methods for producing
biologically active substances encoded by the polynucleotides and to methods of using the
isolated polynucleotide.

<>

<1>Berka, R., Rey, M., Ramaiya, P., Andersen, J., Rasmussen, M., Olsen, P.
<2>Bacillus YvmA inactivation.
<3>European Patent Office
<4>EP 2284184 A
<5>
<6>2011
<7>The present invention relates to an isolated polynucleotide of the complete chromosome of
Bacillus licheniformis.  The present invention also relates to isolated genes of the
chromosome of Bacillus licheniformis which encode biologically active substances and to
nucleic acid constructs, vectors, and host cells comprising the genes as well as methods for
producing biologically active substances encoded by the genes and to methods of using the
isolated genes of the complete chromosome of Bacillus licheniformis.

<>

<1>Berka, R., Rey, M., Ramaiya, P., Andersen, J., Rasmussen, M., Olsen, P.
<2>Bacillus protein inactivation.
<3>European Patent Office
<4>EP 2284185 A
<5>
<6>2011
<7>The present invention relates to an isolated polynucleotide of the complete chromosome of
Bacillus licheniformis.  The present invention also relates to isolated genes of the
chromosome of Bacillus licheniformis which encode biologically active substances and to
nucleic acid constructs, vectors, and host cells comprising the genes as well as methods for
producing biologically active substances encoded by the genes and to methods of using the
isolated genes of the complete chromosome of Bacillus licheiniformis.

<>

<1>Berka, R., Rey, M., Ramaiya, P., Andersen, J., Rasmussen, M., Olsen, P.
<2>Bacillus licheniformis YvmC inactivation.
<3>European Patent Office
<4>EP 2287178 A
<5>
<6>2011
<7>
<>

<1>Berka, R., Rey, M., Ramaiya, P., Andersen, J., Rasmussen, M., Olsen, P.
<2>Increased bacillus YweA expression.
<3>European Patent Office
<4>EP 2287179 A
<5>
<6>2011
<7>
<>

<1>Berka, R., Rey, M., Ramaiya, P., Andersen, J., Rasmussen, M., Olsen, P.
<2>Increased bacillus YweA expression.
<3>European Patent Office
<4>EP 2298797 A
<5>
<6>2011
<7>A Bacillus host cell having increased expression of a polynucleotide encoding an YweA
polypeptide, a method for producing the host cell and methods of using the host cell.

<>

<1>Berka, R., Rey, M., Ramaiya, P., Andersen, J.T., Rasmussen, M.D., Olsen, P.B.
<2>Bacillus licheniformis chromosome.
<3>European Patent Office
<4>EP 2216640 A
<5>
<6>2010
<7>The present invention relates to an isolated polynucleotide of the complete chromosome of
Bacillus licheniformis.  The present invention also relates to isolated genes of the
chromosome of Bacillus licheniformis which encode biologically active substances and to
nucleic acid constructs, vectors, and host cells comprising the genes as well as methods for
producing biologically active substances encoded by the genes and to methods of using the
isolated genes of the complete chromosome of Bacillus licheniformis.

<>

<1>Berka, R.M., Clausen, I.G., Bolotine, A., Sorokine, A., Lapidus, A.
<2>Methods for monitoring multiple gene expression.
<3>US Patent Office
<4>US 7018794 A
<5>
<6>2006
<7>The present invention relates to methods for monitoring differential expression of a plurality
of genes in a first Bacillus cell relative to expression of the same genes in one or more
second Bacillus cells using microarrays containing Bacillus genomic sequenced tags.  The
present invention also relates to computer readable media and computer-based systems.  The
present invention further relates to substrates containing an array of Bacillus licheniformis
or Bacillus clausii GSTs.

<>

<1>Berkhout, B., van Wamel, J.
<2>Accurate scanning of the BssHII endonuclease in search for its DNA cleavage site.
<3>J. Biol. Chem.
<4>271
<5>1837-1840
<6>1996
<7>A facilitated diffusion mechanism has been proposed to account for the kinetic efficiency with
which restriction endonucleases are able to locate DNA recognition sites.  Such a mechanism
involves the initial formation of the DNA, with the subsequent diffusion of the protein along
the DNA helix until either a recognition site is located or the protein dissociates into
solution.  Protein translocation may be facilitated by either sliding along the DNA, hopping
to nearby sites, or intersegment transfer over larger distances.  Previous analyses of the
manner in which restriction enzymes cleave DNA substrates did rule out the latter mechanism.
To discriminate between protein sliding or scanning and protein hopping, we designed a unique
DNA template with three overlapping, mutually exclusive recognition sites for the BssHII
endonuclease.  Analysis of the cleavage pattern demonstrated efficient usage of both external
sites, whereas the centrally located site was not efficiently cleaved.  These results confirm
that linear diffusion of the BssHII enzyme occurs by scanning along the DNA.  Furthermore, the
scanning enzyme was found to stop and cleave at the first site encountered.  Thus, a sliding
restriction endonuclease recognizes cleavage sites with high fidelity, without skipping of
potential sites.

<>

<1>Berkner, K.L., Folk, W.R.
<2>The effects of substituted pyrimidines in DNAs on cleavage by sequence-specific endonucleases.
<3>J. Biol. Chem.
<4>254
<5>2551-2560
<6>1979
<7>The rates of cleavage of DNAs containing substituents at position 5 of thymine
or cytosine have been measured for a variety of sequence-specific
endonucleases, so as to determine which features in the DNA sequence are being
probed.  Phage PhiE DNA fully substituted with 5-hydroxymethyluracil is cleaved
more slowly by enzymes whose recognition sequences contain A-T base pairs than
are DNAs containing thymine, but both types of DNA are cleaved at similar rates
by enzymes recognizing sequences composed only of G-C base pairs.  Phage PBS2
DNA with uracil completely substituted for thymine is cleaved slowly by several
enzymes which recognize sequences containing A-T base pairs (endonucleases
HpaI, HindII, and HindIII), while the rates of cleavage by other enzymes
(endonucleases EcoRI and BamHI) are not affected.  Phage lambda- and P22 DNAs
containing 5-bromouracil are cleaved more slowly by several enzymes
(endonucleases HindIII, HpaI, BamHI) than are thymine-containing DNAs.  Enzymes
that recognize sequence isomers with the composition G:C:2A:2T (endonucleases
EcoRI, HpaI, HindIII) are not equally affected by substitution at position 5 of
thymine, suggesting that they differ in their contacts with A-T base pairs.
DNA containing glucosylated 5-hydroxymethylcytosine in place of cytosine is
resistant to cleavage by all the endonucleases examined.

<>

<1>Berkner, K.L., Folk, W.R.
<2>Overmethylation of DNAs by the EcoRI methylase.
<3>Nucleic Acids Res.
<4>5
<5>435-450
<6>1978
<7>EcoRI methylase is able to catalyze methyl incorporation into DNA at sequences
other than the canonical EcoRI site.  At high enzyme concentrations and over a
wide range of pH and ionic strengths, EcoRI methylase modifies polyoma DNA
(which contains one EcoRI site) at a number of sites.  This modification
prevents EcoRI endonuclease activity, and thus is presumably at or near the
EcoRI sequences (5')NAATTN.

<>

<1>Berkner, K.L., Folk, W.R.
<2>EcoRI cleavage and methylation of DNAs containing modified pyrimidines in the recognition sequence.
<3>J. Biol. Chem.
<4>252
<5>3185-3193
<6>1977
<7>The effects of substituents at position 5 in the pyrimidine ring of a variety
of phage DNAs upon EcoRI endonuclease and methylase activities have been
examined.  The replacement of cytidine in DNA with glucosylated
hydroxymethylcytidine confers resistance to cleavage by the EcoRI endonuclease.
Substitution of thymidine in DNA by hydroxymethyluridine (a change in the
methyl at position 5 of thymidine for a hydroxymethyl) lowers the maximal
velocity of endonucleolytic cleavage 20-fold, but has no detectble effect upon
the Km.  Substitution of thymidine in DNA by uridine (a change in the methyl at
position 5 of thymidine for a hydrogen atom) has no effect upon either the
maximal velocity or the Km.  The effect of these modifications upon EcoRI
methylase activity was markedly different.  DNA containing glucosylated
hydroxymethylcytidine is methylated as well as normal DNA.  DNA containing
uridine or hydroxymethyluridine, in place of thymidine, is much more poorly
methylated than normal DNA.  These different sensitivities of the EcoRI
endonuclease and methylase to modifications in the pyrimidine rings of DNA
suggest there are significant differences in the manner by which these enzymes
recognize and bind to the canonical EcoRI sequence.

<>

<1>Berkner, K.L., Folk, W.R.
<2>Quantitation of the various termini generated by Type II restriction endonucleases using the polynucleotide kinase exchange reaction.
<3>J. Biol. Chem.
<4>254
<5>2561-2564
<6>1979
<7>Parameters of the polynucleotide kinase-catalyzed exchange reaction between [gamma32P]ATP and
5'-phosphoryl DNAs have been measured with the termini generated by the following
endonucleases: EcoRI (Berkner, K.L. and Folk, W.R. (1977) J. Biol. Chem. 252:3176-3184),
HpaII, BamHI, and HindIII (external termini); HindII and HpaI (blunt termini); HaeII and HhaI
(internal termini). In every case, exchange is reproducible and proportional to the number of
termini. However, in most cases, the exchange reaction does not proceed to the theoretical
maximum. External termini and single-stranded DNAs are labeled more rapidly and to
approximately 5-fold higher levels than blunt or internal termini. Concentrations of 100 to
200 micromolar ADP and 12 micromolar ATP are optimal for labeling all types of termini with
the exchange reaction.

<>

<1>Berkner, K.L., Folk, W.R.
<2>An assay for the rates of cleavage of specific sites in DNA by restriction endonucleases:  Its use to study the cleavage of phage lambda DNA by EcoRI and phage P22 DNA containing thymine or 5-bromouracil by HindIII.
<3>Anal. Biochem.
<4>129
<5>446-456
<6>1983
<7>A method to measure the rates of cleavage of specific sites in DNAs by restriction
endonucleases is described.  Partial digests are prepared by incubating DNAs with limiting
amounts of endonuclease.  The termini generated by cleavage are labeled with 32P by the
polynucleotide kinase-exchange reaction.  The labeled termini are then identified by
completing the digestion with the same endonuclease and separating the products by gel
electrophoresis. As the products of complete digestion of DNA are often easily separated and
can be unequivocally identified, this procedure permits comparison of the rates of cleavage of
specific sites in DNAs:  furthermore, because detection of the products of cleavage utilizes
radioautography and does not depend upon their size, or amount, only small amounts of DNA need
to be utilized.  This method has been used to examine the cleavage of phgae lambda DNA by
EcoRI endonuclease, and to demonstrate that 5-bromouracil substitution in phage P22 DNA
reduces the rate of cleavage of most sites by HindIII endonuclease approximately threefold and
the rate of cleavage of one site approximately tenfold.

<>

<1>Berkyurek, A.Can., Suetake, I., Arita, K., Takeshita, K., Nakagawa, A., Shirakawa, M., Tajima, S.
<2>The DNA Methyltransferase Dnmt1 Directly Interacts with the SET and RING Finger-associated (SRA) Domain of the Multifunctional Protein Uhrf1 to Facilitate Accession of the Catalytic Center to Hemi-methylated DNA.
<3>J. Biol. Chem.
<4>289
<5>379-386
<6>2014
<7>Dnmt1 is responsible for the maintenance DNA methylation during replication to propagate
methylation patterns to the next generation. The replication foci targeting sequence (RFTS),
which plugs the catalytic pocket, is necessary for recruitment of Dnmt1 to the replication
site. In the present study we found that the DNA methylation activity of Dnmt1 was DNA
length-dependent and scarcely methylated 12-bp short hemi-methylated DNA. Contrarily, the
RFTS-deleted Dnmt1 and Dnmt1 mutants that destroyed th hydrogen bonds between the RFTS and
catalytic domain showed significant DNA methylation activity even toward 12-bp hemi-methylated
DNA. The DNA methylation activity of the RFTS-deleted Dnmt1 toward 12-bp hemi-methylated DNA
was strongly inhibited on the addition of RFTS, but to a lesser extent by Dnmt1 harboring the
mutations that impair the hydrogen bond formation. The SRA domain of Uhrf1, which is a
prerequisite factor for maintenance methylation and selectively binds to hemi-methylated DNA,
stimulated  the DNA methylation activity of Dnmt1. The SRA to Dnmt1 concentration ratio was
the determinant for the maximum stimulation. In addition, a mutant SRA, which had lost the DNA
binding activity but was able to bind to Dnmt1, stimulated the DNA methylation activity of
Dnmt1. The results indicate that the DNA methylation activity of Dnmt1 was stimulated on the
direct interaction of the SRA and Dnmt1. The SRA facilitated acceptance of the 12-bp
fluorocytosine-containing DNA by the catalytic center. We propose t hat the SRA removes the
RFTS plug from the catalytic pocket to facilitate DNA acceptance by the catalytic center.

<>

<1>Berland, J.L., de Carvalho, F.M., de Almeida, L.G., Bablishvili, N., Gauthier, M., Paranhos-Baccala, G., de Vasconcelos, A.T.
<2>Draft Genome Sequence of Mycobacterium tuberculosis Clinical Strain G-12-005.
<3>Genome Announcements
<4>2
<5>e00385-14
<6>2014
<7>Infection caused by drug-resistant Mycobacterium tuberculosis is a growing concern, especially
in eastern Europe. We report an annotated draft genome
sequence of M. tuberculosis strain G-12-005 obtained from a patient in Georgia.

<>

<1>Berlin, K.
<2>Amplifying genomic DNA so that the cytosine methylation pattern is retained in the amplification product comprises treatment of newly  synthesized products with a methyltransferase and a methyl donor -  genomic DNA amplification for cancer diagnosis.
<3>International Patent Office
<4>WO 200380862
<5>
<6>2003
<7>NOVELTY - Amplifying (M1) genomic DNA, where the cytosine methylation pattern of the genomic
DNA is retained in the
amplification products, is new. DETAILED DESCRIPTION - Amplifying (M1)
genomic DNA, where the cytosine methylation pattern of the genomic DNA
is retained in the amplification products comprises: (a) denaturing the
genomic DNA by heating; (b) cooling the denatured DNA in the presence
of single stranded oligonucleotide primers such that the primers anneal
to the DNA; (c) heating the mixture in the presence of a polymerase and
nucleotides to a temperature such that the primers are extended; (d)
contacting the double stranded nucleic acid with a methyltransferase
and a methyl donor molecule under conditions conducive to the
methylation of the newly synthesized strand such that the CpG
dinucleotides within the synthesized strand are methylated according to
the methylation status of the corresponding CpG dinucleotide on the
template strand thus preserving the genomic methylation pattern; and
(e) repeating the above steps a desired number of times to reach a
desired number of nucleic acids. INDEPENDENT CLAIMS are also included
for: (1) a device for performing M1 comprising two or more reaction
chambers, a channel providing fluid connections between chambers, and
means for regulating the temperature of each chamber (2) a nucleic acid
obtained by M1; and (3) manufacturing a methylated nucleic acid using
M1. BIOTECHNOLOGY - Preferred Method: In M1, the methyl-transferase is
a maintenance methyltransferase. The methlytransferase is DNA
(cytosine-5) Methyl-transferase (DNMT 1). The methyl donor molecule is
S-adenosylmethionine. The methyl group carries a detectable label which
is incorporated into the synthesized nucleic acid strand. The primer
oligonucleotides, methyltransferase and/or the polymerase may be
immobilized on a solid surface. M1 further comprises (as step (f))
treatment with an agent capable of distinguishing between methylated
and unmethylated cytosine bases. The agent is a methylation sensitive
restriction enzyme or a bisulfite solution. USE - M1 is useful for
amplifying genomic DNA, where the cytosine methylation pattern of the
genomic DNA is retained in the amplification product (claimed). The
method is potentially useful for research into, or diagnosis of,
conditions such as cancer.

<>

<1>Berlin, Y.A., Butkus, V.V.
<2>Synthesis of oligodeoxynucleotides containing the sequence GGTACC and their interaction with KpnI restriction endonuclease.
<3>Bioorg. Khim.
<4>7
<5>1224-1232
<6>1981
<7>It has been shown that in the phosphotriester synthesis of
oligodeoxynucleotides the simultaneous use of a dimethoxytrityl and a levulinyl
residue to protect the 5'-OH and 3'-OH, respectively, permits the growth of the
oligonucleotide chain in both directions: 3' 5 5' and 5' 5 3'.  Using this
approach, the oligodeoxynucleotides GGTACC, GGTACCGG, CCGGTACC, and CCGGTACCGG
containing the recognition site for KpnI restriction endonuclease have been
synthesized.  In a study of their interaction with this restrictase it has been
found that for the normal functioning of the enzyme the GGTACC site in its
substrate must be flanked in both chains from the 5'-end and in at least one
chain from the 3'-end.

<>

<1>Berna, L., Iraola, G., Greif, G., Coitinho, C., Rivas, C.M., Naya, H., Robello, C.
<2>Whole-Genome Sequencing of an Isoniazid-Resistant Clinical Isolate of Mycobacterium tuberculosis Strain MtURU-002 from Uruguay.
<3>Genome Announcements
<4>2
<5>e00655-14
<6>2014
<7>The incidence of tuberculosis in Uruguay has been effectively reduced to <30 per  100,000
population, although an increase in nonrisk populations in the last few
years is evident. Here, we present the genome sequence of Mycobacterium
tuberculosis strain MtURU-002 isolated from a patient showing bilateral pulmonary
tuberculosis that was resistant to isoniazid.

<>

<1>Bernacchia, G., Para, A., Pedrali-Noy, G., Cella, R.
<2>Isolation of a cDNA coding for DNA (cytosine-5)-methyltransferase (Accession No. AJ002140) from Lycopersicon esculentum.
<3>Plant Physiol.
<4>116
<5>446
<6>1998
<7>
<>

<1>Bernacchia, G., Primo, A., Giorgetti, L., Pitto, L., Cella, R.
<2>Carrot DNA-methyltransferase is encoded by two classes of genes with differing patterns of expression.
<3>Plant J.
<4>13
<5>317-329
<6>1998
<7>In the present study, the isolation and characterization of two distinct cDNAs that code for
carrot DNA (cytosine-5)-methyltransferase (DNA-METase) are reported.  The screening of a cDNA
library with a carrot genomic DNA fragment, previously obtained by PCR using degenerate
primers, has led to the isolation of clones that belong to two distinct classes of genes (Met1
and Met2) which differ in sequence and size.  Met1-5 and Met2-21 derived amino acid sequences
are more than 85% identical for most of the polypeptide and completely diverge at the
N-terminus.  The larger size of the Met2-21 cDNA is due to the presence of nearly perfect
fivefold repeat of a 171 bp sequence present only once in the Met1-5 cDNA.  Northern  and in
situ hybridization analyses with young carrot plants and somatic embryos indicate that both
genes are maximally expressed in proliferating cells (suspension cells, meri-stems and leaf
primordia), but differ quantitatively and spatially in their mode of expression.  Polyclonal
antibodies were raised in rabbit using fusion proteins corresponding to the regulatory and
catalytic regions of the most highly expressed gene (Met1-5).  In nuclear carrot extracts,
both antibodies were found to recognize a band of about 200 kDa along with some additional
bands of lower size.  These results provide the first direct demonstration that DNA-METases of
a higher eukaryote are encoded by a gene family.

<>

<1>Bernal, W.M., Raven, N.D.H., Williams, R.A.D.
<2>Restriction endonuclease from Thermus ruber.
<3>Proceedings 14th Int. Congress Microbiol.
<4>0
<5>204
<6>1986
<7>Chromosomal DNA purified from some strains ascribed to the genus Thermus were
digestable by restriction endonuclease TaqI, indicating that these strains do
not possess the TaqI restriction-modification system.  These strains were
tested for the possession of restriction enzymes with potentially novel sites
of recognition and cleavage.  Thermus ruber BKMB-1258 has a restriction enzyme
designated TruI, which was separated from other nucleases by phosphocellulose
column chromatography.  From digestion patterns of DNA from bacteriophage
lambda and phiX174 RF, and plasmids pBR322 and pAT153, the recognition sequence
of TruI was identified as GG(A/T)CC, which is the same as for AvaII.  This was
confirmed by TruI-AvaII double digests.

<>

<1>Bernan, V., Sznyter, L.A., Vaccaro, C.M., Jager-Quinton, T., Wilson, G., Brooks, J.E.
<2>Cloning and characterization of the SphI restriction-modification system from S. phaeochromogenes.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>89
<5>206
<6>1989
<7>SphI, a Type II restriction-modification system from the actinomycete
Streptomyces phaeochromogenes, recognizes the sequence GCATGC.  The
endonuclease cleaves at GCATG^C leaving a 3'four base overhang.  A 5.4kb insert
carrying the methylase gene has been cloned into the PstI site of pBR322 and
expressed in E. coli at a low level as evidenced by incomplete modification of
chromosomal DNA and lack of modification of phage DNA in vivo.  The clone has
no endonuclease activity.  The plasmid has been extensively mapped; deletion
subcloning has been used to locate the methylase gene.  The entire 5.4kb
methylase fragment has also been cloned into the Streptomyces promoter-probe
plasmids, pIJ486 and pIJ487 and the low copy Streptomyces vector, pIJ922.
Expression of the SphI methylase gene will be discussed.

<>

<1>Berndt, C., Meier, P., Wackernagel, W.
<2>DNA restriction is a barrier to natural transformation in Pseudomonas stutzeri JM300.
<3>Microbiology
<4>149
<5>895-901
<6>2003
<7>Natural transformation is a mechanism for intra- and interspecific transfer of chromosomal DNA
in Pseudomonas stutzeri. During this process a
single strand derived from duplex DNA is transported into the cytoplasm
and recombined with resident DNA. By electroporation, which introduces
duplex DNA into cells, 100-fold lower transformation frequencies of P.
stutzeri JM300 were observed with shuttle vector or broad-host-range
plasmid DNA when the plasmids had replicated in Escherichia coli and not
in P. stutzeri JM300. Moreover, the natural transformation with cloned
chromosomal P. stutzeri JM300 DNA was reduced about 40-fold when the DNA
had not been propagated in P. stutzeri JM300 but in E. coli. Restriction
was also active during natural transformation by single-stranded DNA.
Restriction during natural transformation and electroporation was
abolished in mutants isolated from mutagenized JM300 cells after applying
a multiple plasmid electroporation strategy for the enrichment of
restriction-defective strains. The mutants had retained the ability for
DNA modification. The P. stutzeri strain ATCC 17587 was found to have no
restriction-modification system as seen in JM300. It is discussed whether
restriction during natural transformation acts at presynaptic or
postsynaptic stages of transforming DNA. Restriction as a barrier to
transformation apparently contributes to sexual isolation and therefore
may promote speciation in the highly diverse species P. stutzeri.

<>

<1>Bernick, D.L., Karplus, K., Lui, L.M., Coker, J.K., Murphy, J.N., Chan, P.P., Cozen, A.E., Lowe, T.M.
<2>Complete genome sequence of Pyrobaculum oguniense.
<3>Standards in Genomic Sciences
<4>6
<5>336-345
<6>2012
<7>Pyrobaculum oguniense TE7 is an aerobic hyperthermophilic crenarchaeon isolated from a hot
spring in Japan. Here we describe its main chromosome of 2,436,033 bp,
with three large-scale inversions and an extra-chromosomal element of 16,887 bp.
We have annotated 2,800 protein-coding genes and 145 RNA genes in this genome,
including nine H/ACA-like small RNA, 83 predicted C/D box small RNA, and 47
transfer RNA genes. Comparative analyses with the closest known relative, the
anaerobe Pyrobaculum arsenaticum from Italy, reveals unexpectedly high synteny
and nucleotide identity between these two geographically distant species. Deep
sequencing of a mixture of genomic DNA from multiple cells has illuminated some
of the genome dynamics potentially shared with other species in this genus.

<>

<1>Bernstein, R.L., Dharmgrongartama, B., Srinivasan, P.R.
<2>Altered P2 bacteriophage restriction in E. coli B.
<3>Fed. Proc.
<4>29
<5>860
<6>1970
<7>P2 phage grown on E. coli C (P2.C) grows very well in E. coli C but is highly
restricted in E. coli B.  The plating efficiences of P2.C with E. coli C and E.
coli B grown in nutrient broth are of the order of 1 and 10-7, respectively.
This retriction can be altered by changing the growth conditions of E. coli B.
Thus growth on salts-glucose medium gives a 100-fold increase in plating
efficiency compared to cultures grown in enriched broth.  When E. coli B is
grown in salts-glucose medium supplemented with amino acids, purines,
pyrimidines, and vitamins, the higher P2.C plating efficiencies are found only
with those cultures not supplemented with methionine.  Conditions which lead to
the depletion of intracellular methionine or S-adenosylmethionine in the
prototrophic E. coli B further increase the P2.C plating efficiencies.  Plating
efficiency can also be raised by infection with T3 phage.  However, infection
with T3 mutants unable to produce the T3-specific early enzyme which cleaves
S-adenosylmethionine does not alter the plating efficiency.  These results
suggest that S-adenosylmethionine is involved in the mechanism of restriction
of P2 phage.

<>

<1>Bertalan, M. et al.
<2>Complete genome sequence of the sugarcane nitrogen-fixing endophyte Gluconacetobacter diazotrophicus Pal5.
<3>BMC Genomics
<4>10
<5>450
<6>2009
<7>BACKGROUND: Gluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that
lives in association with sugarcane plants. It has important
biotechnological features such as nitrogen fixation, plant growth promotion,
sugar metabolism pathways, secretion of organic acids, synthesis of auxin and the
occurrence of bacteriocins. RESULTS: Gluconacetobacter diazotrophicus Pal5 is the
third diazotrophic endophytic bacterium to be completely sequenced. Its genome is
composed of a 3.9 Mb chromosome and 2 plasmids of 16.6 and 38.8 kb, respectively.
We annotated 3,938 coding sequences which reveal several characteristics related
to the endophytic lifestyle such as nitrogen fixation, plant growth promotion,
sugar metabolism, transport systems, synthesis of auxin and the occurrence of
bacteriocins. Genomic analysis identified a core component of 894 genes shared
with phylogenetically related bacteria. Gene clusters for gum-like polysaccharide
biosynthesis, tad pilus, quorum sensing, for modulation of plant growth by indole
acetic acid and mechanisms involved in tolerance to acidic conditions were
identified and may be related to the sugarcane endophytic and plant-growth
promoting traits of G. diazotrophicus. An accessory component of at least 851
genes distributed in genome islands was identified, and was most likely acquired
by horizontal gene transfer. This portion of the genome has likely contributed to
adaptation to the plant habitat. CONCLUSION: The genome data offer an important
resource of information that can be used to manipulate plant/bacterium
interactions with the aim of improving sugarcane crop production and other
biotechnological applications.

<>

<1>Bertani, G., Weigle, J.J.
<2>Host controlled variation in bacterial viruses.
<3>J. Bacteriol.
<4>65
<5>113-121
<6>1953
<7>Passage through new hosts or new tissues is a widely used method for altering
the properties of viruses.  In some instances selection of spontaneous mutants
has been demonstrated to be the mechanism causing the variation (Luria, 1945).
Nonhereditary mechanisms sometimes have been postulated, but since no such case
has been analyzed sufficiently, it is often assumed that selection of mutants
is the only possible mechanism.  A detailed analysis of two cases of variation
in two different bacterial viruses is reported in this paper.  In both these
cases we are dealing with nonheritable alterations stemming directly from
passage through a new host and not with mutations.  A somewhat similar case of
host controlled variation involving other bacterial viruses has been reported
recently by Luria and Human (1952).

<>

<1>Bertani, I., Passos-da-Silva, D., Abbruscato, P., Piffanelli, P., Venturi, V.
<2>Draft Genome Sequence of the Plant Pathogen Dickeya zeae DZ2Q, Isolated from Rice in Italy.
<3>Genome Announcements
<4>1
<5>e00905-13
<6>2013
<7>Dickeya zeae is an emerging rice (Oryza sativa) pathogen causing bacterial foot rot. Related
pathogens affect maize (Zea mays) and potato (Solanum tuberosum) and
a variety of important ornamental and floral plants. Here, we present the draft
genome sequence of D. zeae DZ2Q, an isolate obtained from rice grown in Italy.

<>

<1>Bertelli, C., Collyn, F., Croxatto, A., Ruckert, C., Polkinghorne, A., Kebbi-Beghdadi, C., Goesmann, A., Vaughan, L., Greub, G.
<2>The Waddlia Genome: A Window into Chlamydial Biology.
<3>PLoS ONE
<4>5
<5>e10890
<6>2010
<7>Growing evidence suggests that a novel member of the Chlamydiales order, Waddlia chondrophila,
is a potential agent of miscarriage in humans and abortion in ruminants. Due to the lack of
genetic tools to manipulate chlamydia, genomic analysis is proving to be the most incisive
tool in stimulating investigations into the biology of these obligate intracellular
bacteria. 454/Roche and Solexa/Illumina technologies were thus used to sequence and assemble
de novo the full genome of the first representative of the Waddliaceae family, W.
chondrophila. The bacteria possesses a 291169312bp chromosome and a 159593 bp low-copy number
plasmid that might integrate into the bacterial chromosome. The Waddlia genome displays
numerous repeated sequences indicating different genome dynamics from classical chlamydia
which almost completely lack repetitive elements. Moreover, W. chondrophila exhibits many
virulence factors also present in classical chlamydia, including a functional type III
secretion system, but also a large complement of specific factors for resistance to host or
environmental stresses. Large families of outer membrane proteins were identified indicating
that these highly immunogenic proteins are not Chlamydiaceae specific and might have been
present in their last common ancestor.  Enhanced metabolic capability for the synthesis of
nucleotides, amino acids, lipids and other co-factors suggests that the common ancestor of the
modern Chlamydiales may have been less dependent on their eukaryotic host. The fine-detailed
analysis of biosynthetic pathways brings us closer to possibly developing a synthetic medium
to grow W. chondrophila, a critical step in the development of genetic tools. As a whole, the
availability of the W. chondrophila genome opens new possibilities in Chlamydiales research,
providing new insights into the evolution of members of the order Chlamydiales and the biology
of the Waddliaceae.

<>

<1>Bertelli, C., Goesmann, A., Greub, G.
<2>Criblamydia sequanensis Harbors a Megaplasmid Encoding Arsenite Resistance.
<3>Genome Announcements
<4>2
<5>e00949-14
<6>2014
<7>Criblamydia sequanensis is an amoeba-resisting bacterium recently isolated from the Seine
River. This Chlamydia-related bacterium harbors a genome of
approximately 3 Mbp and a megaplasmid of 89,525 bp. The plasmid encodes several
efflux systems and an operon for arsenite resistance. This first genome sequence
within the Criblamydiaceae family enlarges our view on the evolution and the
ecology of this important bacterial clade largely understudied so far.

<>

<1>Bertinuson, A., Illingworth, C.A., Otto, C.J., Hornemann, U.
<2>An apparently restriction deficient mutant of Streptomyces achromogenes var streptozoticus can be transformed by plasmid pIJ350.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>84
<5>H74
<6>1984
<7>We have isolated a mutant of S. achromogenes var streptozoticus (SAS) which
differed from wild type in respect to digestion of chromosomal DNA by NaeI and
susceptibility to phage infection.  While DNA of SAS was resistant to digestion
by NaeI, the DNA of mutant AB301 was NaeI-sensitive.  We found that AB301 could
be infected by 4 phages to which wild type SAS was resistant.  We were able to
transform AB301 with plasmid pIJ350 (confirmed by mini-prepping transformants),
although we had never successfully transformed wild type SAS with this or any
other plasmid.  The DNA of pIJ 350 and phages PhiCC 53, PhiCC 55, and PhiCC 57
was cut by NaeI, but that of phage PhiCC 26 was not.  We suggest that
restriction by NaeI is one barrier to transformation in SAS and that loss of
this barrier (and additional possible restriction barriers) contributed to the
successful transformation of this previously recalcitrant species.

<>

<1>Bertolini, C., van Aerle, R., Lampis, S., Moore, K.A., Paszkiewicz, K., Butler, C.S., Vallini, G., van der Giezen, M.
<2>Draft Genome Sequence of Stenotrophomonas maltophilia SeITE02, a Gammaproteobacterium Isolated from Selenite-Contaminated Mining Soil.
<3>Genome Announcements
<4>2
<5>e00331-14
<6>2014
<7>Stenotrophomonas maltophilia strain SeITE02 was isolated from the rhizosphere of  the
selenium-hyperaccumulating legume Astragalus bisculcatus. In this report, we
provide the 4.56-Mb draft genome sequence of S. maltophilia SeITE02, a
gammaproteobacterium that can withstand high concentrations of selenite and
reduce these to elemental selenium.

<>

<1>Bertschinger, J., Neri, D.
<2>Covalent DNA display as a novel tool for directed evolution of proteins in vitro.
<3>Protein Eng. Des. Sel.
<4>17
<5>699-707
<6>2004
<7>We present a novel method for the directed evolution of polypeptides, which combines in vitro
compartmentalization and covalent DNA display. A
library of linear DNA fragments is co-packaged with an in vitro
transcription/translation mixture in the compartments of a water-in-oil
emulsion. Experimental conditions are adjusted so that, in most cases, one
compartment contains one DNA molecule. The DNA fragments encode fusion
proteins containing a DNA-methyltransferase (M.Hae III), which can form a
covalent bond with a 5-fluorodeoxycytidine base at the extremity of the
DNA fragment. The resulting library of DNA-protein fusions is extracted
from the emulsion and DNA molecules displaying a protein with desired
binding properties are selected from the pool of DNA-protein fusions by
affinity panning on target antigens. We applied this methodology in model
selection experiments, using specific ligands for the capture of peptides
and globular proteins bound to DNA. We observed enrichment factors
>1000-fold for selections performed in separate emulsions and up to
150-fold for selections performed using mixtures of DNA molecules. M.Hae
III could be fused to small globular proteins (such as calmodulin and
fibronectin domains), which are ideally suited for the generation of
combinatorial libraries and for the isolation of novel binding
specificities.

<>

<1>Besnier, C.E., Kong, H.
<2>Converting MlyI endonuclease into a nicking enzyme by changing its oligomerization state.
<3>EMBO Rep.
<4>2
<5>782-786
<6>2001
<7>N.Bst NBI is a nicking endonuclease that recognizes the sequence GAGTC and nicks one DNA
strand specifically. The Type IIs endonuclease, Mly I, also recognizes GAGTC, but cleaves both
DNA strands. Sequence comparisons revealed significant similarities between N.Bst NBI and Mly
I. Previous studies showed that Mly I dimerizes in the presence of a cognate DNA, whereas
N.Bst NBI remains a monomer. This suggests that dimerization may be required for
double-stranded cleavage. To test this hypothesis, we used a multiple alignment to design
mutations to disrupt the dimerization function of Mly I. When Tyr491 and Lys494 were both
changed to alanine, the mutated endonuclease, N.Mly I, no longer formed a dimer and cleaved
only one DNA strand specifically. Thus, we have shown that changing the oligomerization state
of an enzyme changes its enzymatic function. This experiment also established a protocol that
could be applied to other Type IIs endonucleases in order to generate more novel nicking
endonucleases.

<>

<1>Bessen, D.E., Kumar, N., Hall, G.S., Riley, D.R., Luo, F., Lizano, S., Ford, C.N., McShan, W.M., Nguyen, S.V., Dunning-Hotopp, J.C., Tettelin, H.
<2>Whole Genome Association Study on Tissue Tropism Phenotypes in Group A Streptococcus.
<3>J. Bacteriol.
<4>193
<5>6651-6663
<6>2011
<7>Group A Streptococcus (GAS) has a rich evolutionary history of horizontal transfer among its
core genes. Yet, despite extensive genetic mixing, GAS strains have discrete ecological
phenotypes. To further our understanding of the molecular basis for ecological phenotypes,
comparative genomic hybridization of a set of 97 diverse strains to a GAS pan-genome
microarray was undertaken, and the association of accessory genes with emm genotypes that
define tissue tropisms for infection was determined. Of the 22 non-prophage, accessory gene
regions (AGRs) identified, only three AGRs account for all statistically significant linkage
disequilibrium among strains having the genotypic biomarkers for throat versus skin infection
specialist. Networked evolution and population structure analysis of loci representing each of
the AGRs reveals that most strains with the skin specialist and generalist biomarkers form
discrete clusters, whereas strains with the throat specialist biomarker are highly diverse. To
identify co-inherited and co-selected accessory genes, the strength of genetic associations
was determined for all possible pair wise combinations of accessory genes among the 97 GAS
strains. Accessory genes showing very strong associations provide the basis for an
evolutionary model, which reveals that a major transition between many throat and skin
specialist haplotypes correlates with the gain or loss of genes encoding fibronectin-binding
proteins. This study employs a novel synthesis of tools to help delineate the major genetic
changes associated with key adaptive shifts in an extensively recombined bacterial species.

<>

<1>Bessho, Y., Shibata, R., Shiramizu, M., Yokoyama, S., Ohama, T., Kurokawa, S.
<2>Preparation of homing endonuclease I-CsmI from Chlamydomonas smithii and its use for genetic mapping.
<3>Japanese Patent Office
<4>JP 2005341803 A
<5>
<6>2005
<7>
<>

<1>Bestor, T.
<2>Structure of mammalian DNA methyltransferase as deduced from the inferred amino acid sequence and direct studies of the protein.
<3>Biochem. Soc. Trans.
<4>16
<5>944-947
<6>1988
<7>DNA (cytosine-5)-methyltransferase establishes and maintains methylation patterns in the
genome of higher eukaryotes.  This enzyme has been purified, and the cDNA which encodes it has
been cloned and sequenced.  DNA MeTase appears to contain a large (1000 amino acid) N-terminal
domain that contains potential metal-binding sites.  This domain appears to contain a series
of five to seven structural units of Mr about 20,000, since post-translational processing in
vivo or partial proteolysis of the purified protein in vitro leads to the production of a
series of catalytically active species differing in Mr by units of 20,000.  The N-terminal
domain is fused to a small (570 amino acid) C-terminal domain that is related to bacterial
type II cytosine methyltransferases.  The relevance of these findings for the biological
function of DNA MeTase is discussed.

<>

<1>Bestor, T.
<2>Supercoiling-dependent sequence specificity of mammalian DNA mthyltransferase.
<3>Nucleic Acids Res.
<4>15
<5>3835-3843
<6>1987
<7>Negative supercoiling of substrate DNA dramatically alters the in vitro sequence specificity
of mammalian DNA methyltransferase.  This result suggests in vivo site selection by DNA MeTase
could be regulated by conformational information in the form of alternative secondary
structures indicated in DNA by local supercoiling or by the binding of specific nuclear
proteins.  DNA in the left-handed Z-form is shown not to be a substrate for mammalian DNA
MeTase.  The sensitivity of DNA MeTase to DNA structure may also make it useful as a probe for
sequences which undergo supercoiling-dependent structural transitions in vitro.

<>

<1>Bestor, T., Laudano, A., Mattaliano, R., Ingram, V.
<2>Cloning and sequencing of a cDNA encoding DNA methyltransferase of mouse cells.
<3>J. Mol. Biol.
<4>203
<5>971-983
<6>1988
<7>A cDNA encoding DNA (cytosine-5)-methyltransferase (DNA MTase) of mouse cells has been cloned
and sequenced.  The nucleotide sequence contains an open reading frame sufficient to encode a
polypeptide of 1573 amino acid residues, which is close to the apparent size of the largest
species of DNA MTase found in mouse cells.  The carboxylterminal 570 amino acid residues of
the inferred protein sequence shows striking similarities to bacterial type II DNA cytosine
methyltransferases and appears to represent a catalytic methyltransferase domain.  The
amino-terminal portion of the molecule may be involved in regulating the activity of the
carboxyl-terminal methyltransferase domain, since antibodies directed against a peptide
sequence located within this region inhibits transmethylase activity in vitro.  A 5200 base
DNA MTase-specific mRNA was found to be expressed in all mouse cell types tested, and cell
lines known to have different genomic methylation patterns were found to contain DNA MTase
proteins of similar or identical sizes and de novo sequence specificities.  The implications
of these findings for an understanding of the mechanisms involved in the establishment and
maintenance of methylation patterns are discussed.

<>

<1>Bestor, T.H.
<2>Cloning of a mammalian DNA methyltransferase.
<3>Gene
<4>74
<5>9-12
<6>1988
<7>Cloning and sequencing of cDNA clones has shown that mammalian DNA
(cytosine-5)-methyltransferase comprises a 1000-amino acid (aa) N-terminal
region of unknown function and a 570-aa C-terminal region that is clearly
related to bacterial Type II cytosine restriction methyltransferases.  These
findings indicate that the mammalian enzyme contains at least two structural
domains and suggest a common evolutionary origin for mammalian and prokaryotic
DNA (cytosine-5)-methyltransferases.

<>

<1>Bestor, T.H.
<2>Activation of mammalian DNA methyltransferase by cleavage of a Zn binding regulatory domain.
<3>EMBO J.
<4>11
<5>2611-2617
<6>1992
<7>Mammalian DNA (cytosine-5) methyltransferase contains a C-terminal domain that is closely
related to bacterial cytosine-5 restriction methyltransferases. This methyltransferase domain
is linked to a large N-terminal domain. It is shown here that the N-terminal domain contains a
Zn binding site and that the N- and C-terminal domains can be separated by cleavage with
trypsin or Staphylococcus aureus protease V8; the protease V8 cleavage site was determined by
Edman degradation to lie 10 residues C-terminal of the run of alternating lysyl and glycyl
residues which joins the two domains and six residues N-terminal of the first sequence motif
conserved between the mammalian and bacterial cytosine methyltransferases. While the intact
enzyme had little activity on unmethylated DNA substrates, cleavage between the domains caused
a large stimulation of the initial velocity of methylation of unmethylated DNA without
substantial change in the rate of methylation of hemimethylated DNA. These findings indicate
that the N-terminal domain of DNA methyltransferase ensures the clonal propagation of
methylation patterns through inhibition of the de novo activity of the C-terminal domain.
Mammalian DNA methyltransferase is likely to have arisen via fusion of a prokaryotic-like
restriction methyltransferase and an unrelated DNA binding protein. Stimulation of the de novo
activity of DNA methyltransferase by proteolytic cleavage in vivo may contribute to the
process of ectopic methylation observed in the DNA of aging animals, tumors and in lines of
cultured cells.

<>

<1>Bestor, T.H.
<2>DNA methylation: evolution of a bacterial immune function into a regulator of gene expression and genome structure in higher eukaryotes.
<3>Philos. Trans. R. Soc. Lond. B. Biol. Sci.
<4>326
<5>179-187
<6>1990
<7>The amino acid sequence of mammalian DNA methyltransferase has been deduced from the
nucleotide sequence of a cloned cDNA. It appears that the mammalian enzyme arose during
evolution via fusion of a prokaryotic restriction methyltransferase gene and a second gene of
unknown function. Mammalian DNA methyltransferase currently comprises an N-terminal domain of
about 1000 amino acids that may have a regulatory role and a C-terminal 570 amino acid domain
that retains similarities to bacterial restriction methyltransferases. The sequence
similarities among mammalian and bacterial DNA cytosine methyltransferases suggest a common
evolutionary origin. DNA methylation is uncommon among those eukaryotes having genomes of less
than 108 base pairs, but nearly universal among large-genome eukaryotes. This and other
considerations make it likely that sequence inactivation by DNA methylation has evolved to
compensate for the expansion of the genome that has accompanied the development of higher
plants and animals. As methylated sequences are usually propagated in the repressed,
nuclease-insensitive state, it is likely that DNA methylation compartmentalizes the genome to
facilitate gene regulation by reducing the total amount of DNA sequence that must be scanned
by DNA-binding regulatory proteins. DNA methylation is involved in immune recognition in
bacteria but appears to regulate the structure and expression of the genome in complex higher
eukaryotes. I suggest that the DNA-methylating system of mammals was derived from that of
bacteria by way of a hypothetical intermediate that carried out selective de novo methylation
of exogenous DNA and propagated the methylated DNA in the repressed state within its own
genome. During the evolution of complex plants and animals the inactivating effects of DNA
methylation spread to extraneous cellular sequences, such as highly repetitive DNA and
transposable elements, and later to tissue-specific genes. Modern large-genome eukaryotes have
adapted a primitive prokaryotic immune system to enable them to manage a genome that has
expanded more than 1000-fold as a result of accumulation of extraneous sequences and
tissue-specific genes.

<>

<1>Bestor, T.H.
<2>Methylation patterns in the vertebrate genome.
<3>J. NIH Res.
<4>5
<5>57-60
<6>1993
<7>The estimated amount of DNA in a mammalian cell is 6x109 base pairs, the equivalent of more
than 2 m of linear DNA. Cells of some amphibians and vascular plants contain even larger
amounts. However, only a small fraction of the genome is available for interaction with
transcription factors that regulate gene expression, and the accessibility of specific
sequences to these factors appears to be controlled in large part by methylation at the 5
position of cytosine residues in 5'-CpG-3' dinucleotides. Most DNA in large-genome
eukaryotes is methylated and inaccessible to the transcription apparatus, whereas promoter
regions are normally unmethylated and accessible to regulatory factors. Methylation of
promoter regions prevents transcription, as in the case of genes inactivated by genomic
imprinting and those on the inactive X chromosome. Programmed changes in the methylation
status of DNA may also be involved in the regulation of tissue-specific genes during
development, although this hypothesis remains controversial. Another unsolved question concern
the ways in which sex-and tissue-specific methylation patterns are established during
gametogenesis and early development; this is an important issue for human health because
certain human diseases are likely to involve defective establishment or maintenance of genomic
methylation patterns.

<>

<1>Bestor, T.H.
<2>DNA methyltransferases in mammalian development and genome defense.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, Russo, V.E.A., Martienssen, R.A., Riggs, A.D., Cold Spring Harbor
<4>0
<5>61-76
<6>1996
<7>The mammalian genome is modified by the addition of about 3 x 10^7 methyl groups, all at the 5
position of cytosine and most at 5'-CpG-3' dinucleotides.  Methylation patterns are
transmitted by clonal inheritance and increase the information content of the genome;
transcription is repressed when CpG sites within promoters are methylated.  Cytosine
methylation is dangerous: 5-methylcytosine (m5C) is the major endogenous mutagen (deamination
results in C-T transition mutations at CpG sites, which account for about one-third of all
mutations in humans, and tumor suppressor genes are frequently inactivated by ectopic de novo
methylation of promoter regions.  However, there must be benefits that yield a net selective
advantage.  This is shown by the retention of cytosine methylation by virtually all organisms
with genomes of more than 5 x 10^8 bp and by the fact that perturbations of methylation
patterns are lethal to mouse embryos and to differentiated cells.  Severe developmental
abnormalities are seen when m5C levels are reduced in Arabidopsis as the result of mutations
at uncharacterized loci or by expression of a DNA methyltransferase antisense construct.
Although methylation patterns clearly play an essential role in large-genome eukaryotes, the
nature of that role is not understood.  It is argued here that an understanding of the
regulation of de novo methylation will reveal the biological function of methylation patterns.

<>

<1>Bestor, T.H.
<2>Chimeric DNA-binding/DNA methyltransferase nucleic acid and polypeptide and uses thereof.
<3>International Patent Office
<4>WO 9711972
<5>
<6>1997
<7>The present invention provides a chimeric protein which comprises a mutated DNA
methyltransferase portion and a DNA binding protein portion that binds sufficiently close to a
promoter sequence of a target gene which promoter sequence contains a methylation site, to
specifically methylate the site and inhibit activity of the promoter and thus inhibit
expression of the target gene.  This invention also provides for a method for inhibiting the
expression of a target gene which includes contacting a promoter of the target gene with the
chimeric protein, so as to specifically methylate the promoter sequence of the target gene
thus inhibiting expression of the target gene.

<>

<1>Bestor, T.H.
<2>The DNA methyltransferases of mammals.
<3>Hum. Mol. Genet.
<4>9
<5>2395-2402
<6>2000
<7>The biological significance of 5-methylcytosine was in doubt for many years, but is no longer,
Through targeted mutagenesis in mice it has
been learnt that every protein shown by biochemical tests to be
involved in the establishment, maintenance or interpretation of genomic
methylation patterns is encoded by an essential gene. A human genetic
disorder (ICF syndrome) has recently been shown to be caused by
mutations in the DNA methyltransferase 3B (DNMT3B) gene, A second human
disorder (Rett syndrome) has been found to result from mutations in the
MECP2 gene, which encodes a protein that binds to methylated DNA.
Global genome demethylation caused by targeted mutations in the DNA
methyltransferase-l (Dnmt1) gene has shown that cytosine methylation
plays essential roles in X-inactivation, genomic imprinting and genome.
stabilization. The majority of genomic 5-methylcytosine is now known to
enforce the transcriptional silence of the enormous burden of
transposons and retroviruses that have accumulated in the mammalian
genome. It has also become clear that programmed changes in methylation
patterns are less important in the regulation of mammalian development
than was previously believed. Although a number of outstanding
questions have yet to be answered (one of these questions involves the
nature of the cues that designate sites for methylation at particular
stages of gametogenesis and early development), studies of DNA
methyltransferases are likely to provide further insights into the
biological functions of genomic methylation patterns.

<>

<1>Bestor, T.H., Ingram, V.M.
<2>Growth-dependent expression of multiple species of DNA methyltransferase in murine erythroleukemia cells.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>82
<5>2674-2678
<6>1985
<7>Friend murine erythroleukemia cells were found to contain three distinct species of DNA
(cytosine-5-)-methyltransferase whose relative proportions were a characteristic function of
the proliferative state of the cells.  Rapidly proliferating cells contained a Mr 190,000
species of DNA MeTase, whereas cells in the late logarithmic/early plateau phase of cellular
growth contained two species of Mr 150,000 and 175,000 (DNA MeTases I and II); stationary
phase cells contained primarily DNA MeTase I.  The three species of DNA MeTase displayed
structural similarities, as determined by analysis of partial proteolysis products, and have
similar de novo sequence specificites in transmethylation reactions involving purified enzyme
and prokaryotic DNA.  The different relative proportions of the enzymes in cells under
different growth conditions suggest that the three species of DNA MeTase fulfill different
roles in processes leading to the perpetuation of DNA methylation patterns.

<>

<1>Bestor, T.H., Ingram, V.M.
<2>Two DNA methyltransferases from murine erythroleukemia cells: purification, sequence specificity, and mode of interaction with DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>80
<5>5559-5563
<6>1983
<7>Dye-ligand chromatography on Cibacron blue F3GA-agarose has been used to resolve two species
of DNA (cytosine-5-)-methyltransferase from nuclear
extracts of uninduced Friend murine erythroleukemia cells. Each species
has been highly purified; the activities in the first and second peaks
were associated with polypeptides of Mr 150,000 and 175,000, respectively.
Analysis of substrate specificity with synthetic DNAs and restriction
fragments of phi X174 replicative form DNA and pBR322 DNA showed that
neither enzyme had dependence on the sequence context of CpG
dinucleotides; poly(dG-dC) had the greatest methyl-accepting activity of
any unmethylated DNA substrate tested. De novo methylation by both enzymes
was inefficient relative to methylation of hemimethylated sites.
Methyl-accepting activity was strongly dependent on DNA chain length. This
observation suggests that binding to DNA, followed by one-dimensional
diffusion of enzyme along the DNA molecule, is important in the mechanism
by which DNA methyltransferase locates its recognition sites.

<>

<1>Bestor, T.H., Verdine, G.L.
<2>DNA methyltransferases.
<3>Curr. Opin. Cell Biol.
<4>6
<5>380-389
<6>1994
<7>Mammals have long been known to tag their DNA by the addition of methyl groups to cytosine
residues. Only quite recently, however, has the functional significance of DNA methylation
established a firm footing. Evidence now indicates that DNA methylation is essential for
development, and is involved in both programmed and ectopic gene inactivation. Recent
structural and mechanistic work on bacterial cytosine-5-methyltransferases has provided much
insight into the function of the carboxy-terminal catalytic domain of eukaryotic
cytosine-5-methyltransferases; evidence is emerging that the amino-terminal domain targets the
enzyme to the replication machinery and may be involved in sensing the pre-existing
methylation state of the DNA.

<>

<1>Bettegowda, C., Huang, X., Lin, J., Cheong, I., Kohli, M., Szabo, S.A., Zhang, X., Diaz, L.A. Jr., Velculescu, V.E., Parmigiani, G., Kinzler, K.W., Vogelstein, B., Zhou, S.
<2>The genome and transcriptomes of the anti-tumor agent Clostridium novyi-NT.
<3>Nat. Biotechnol.
<4>24
<5>1573-1580
<6>2006
<7>Bacteriolytic anti-cancer therapies employ attenuated bacterial strains that selectively
proliferate within tumors. Clostridium novyi-NT spores
represent one of the most promising of these agents, as they generate
potent anti-tumor effects in experimental animals. We have determined the
2.55-Mb genomic sequence of C. novyi-NT, identifying a new type of
transposition and 139 genes that do not have homologs in other bacteria.
The genomic sequence was used to facilitate the detection of transcripts
expressed at various stages of the life cycle of this bacterium in vitro
as well as in infections of tumors in vivo. Through this analysis, we
found that C. novyi-NT spores contained mRNA and that the spore
transcripts were distinct from those in vegetative forms of the bacterium.

<>

<1>Beuck, C., Singh, I., Bhattacharya, A., Hecker, W., Parmar, V.S., Seitz, O., Weinhold, E.
<2>Polycyclic Aromatic DNA-base surrogates: High-affinity binding to an adenine-specific base-flipping DNA methyltransferase.
<3>Angew. Chem. Int. Ed. Engl.
<4>42
<5>3958-3960
<6>2003
<7>DNA methylation is an important biological event that serves diverse cellular functions such
as protection against endogenous restriction endonucleases, direction of DNA-mismatch repair,
as well as regulation of gene expression and DNA replication.  DNA methylation is catalyzed by
DNA methyltransferases (MTases), which bind to specific DNA sequences and transfer a methyl
group from S-adenosyl-L-methionine to the exocyclic amino groups of adenine or cytosine or to
C5 of cytosine.  Interestingly, methylation is commenced by flipping the target nucleotide
completely out of the helix.  The energy cost for disrupting Watson-Crick hydrogen bonds and
base-stacking interactions is mainly compensated by specific binding of the target bases
within the active sites of the enzymes and by stabilizing the formed apparent abasic site.
Three different mechanisms of abasic-site stabilization have been observed in crystal
structures of DNA MTases complexed with DNA.  The cytosine-specific DNA MTases M.HhaI and
M.HaeIII either only insert an amino acid side chain into the opened space or insert an amino
acid side chain in addition to rearranging the base pairing.  This is in contrast to the
adenine-specific DNA MTase M.TaqI.  In this case, the formed unpaired partner base is inserted
into the opened space, resulting in interstrand stacking.

<>

<1>Beutin, L., Hammerl, J.A., Strauch, E., Reetz, J., Dieckmann, R., Kelner-Burgos, Y., Martin, A., Miko, A., Strockbine, N.A., Lindstedt, B.A., Horn, D., Monse, H., Huettel, B., Muller, I., Stuber, K., Reinhardt, R.
<2>Spread of a Distinct Stx2-Encoding Phage Prototype among Escherichia coli O104:H4 Strains from Outbreaks in Germany, Norway, and Georgia.
<3>J. Virol.
<4>86
<5>10444-10455
<6>2012
<7>Shiga toxin 2 (Stx2)-producing Escherichia coli (STEC) O104:H4 caused one of the
world's largest outbreaks of hemorrhagic colitis and hemolytic uremic syndrome in
Germany in 2011. These strains have evolved from enteroaggregative E. coli (EAEC)
by the acquisition of the Stx2 genes and have been designated enteroaggregative
hemorrhagic E. coli. Nucleotide sequencing has shown that the Stx2 gene is
carried by prophages integrated into the chromosome of STEC O104:H4. We studied
the properties of Stx2-encoding bacteriophages which are responsible for the
emergence of this new type of E. coli pathogen. For this, we analyzed Stx
bacteriophages from STEC O104:H4 strains from Germany (in 2001 and 2011), Norway
(2006), and the Republic of Georgia (2009). Viable Stx2-encoding bacteriophages
could be isolated from all STEC strains except for the Norwegian strain. The Stx2
phages formed lysogens on E. coli K-12 by integration into the wrbA locus,
resulting in Stx2 production. The nucleotide sequence of the Stx2 phage P13374 of
a German STEC O104:H4 outbreak was determined. From the bioinformatic analyses of
the prophage sequence of 60,894 bp, 79 open reading frames were inferred.
Interestingly, the Stx2 phages from the German 2001 and 2011 outbreak strains
were found to be identical and closely related to the Stx2 phages from the
Georgian 2009 isolates. Major proteins of the virion particles were analyzed by
mass spectrometry. Stx2 production in STEC O104:H4 strains was inducible by
mitomycin C and was compared to Stx2 production of E. coli K-12 lysogens.

<>

<1>Beutin, L., Kruger, U., Krause, G., Miko, A., Martin, A., Strauch, E.
<2>Evaluation of major types of Shiga toxin 2E-producing Escherichia coli bacteria present in food, pigs, and the environment as potential pathogens for humans.
<3>Appl. Environ. Microbiol.
<4>74
<5>4806-4816
<6>2008
<7>Shiga toxin 2e (Stx2e)-producing strains from food (n = 36), slaughtered
pigs (n = 25), the environment (n = 21), diseased pigs (n = 19), and
humans (n = 9) were investigated for production of Stx2e by enzyme-linked
immunosorbent assay, for virulence markers by PCR, and for their serotypes
to evaluate their role as potential human pathogens. Stx2e production was
low in 64% of all 110 strains. Stx2e production was inducible by mitomycin
C but differed considerably between strains. Analysis by nucleotide
sequencing and transcription of stx(2e) genes in high- and
low-Stx2e-producing strains showed that toxin production correlated with
transcription rates of stx(2e) genes. DNA sequences specific for the int,
Q, dam, and S genes of the stx(2e) bacteriophage P27 were found in 109
strains, indicating cryptic P27-like prophages, although 102 of these were
not complete for all genes tested. Genes encoding intimin (eae),
enterohemorrhagic Escherichia coli hemolysin (ehx), or other stx(1) or
stx(2) variants were not found, whereas genes for heat-stable enterotoxins
STI, STII, or EAST1 were present in 54.5% of the strains. Seven major
serotypes that were associated with diseased pigs (O138:H14, O139:H1, and
O141:H4) or with slaughter pigs, food, and the environment (O8:H4, O8:H9,
O100:H30, and O101:H9) accounted for 60% of all Stx2e strains. The human
Stx2e isolates did not belong to these major serotypes of Stx2e strains,
and high production of Stx2e in human strains was not related to diarrheal
disease. The results from this study and other studies do not point to
Stx2e as a pathogenicity factor for diarrhea and hemolytic uremic syndrome
in humans.

<>

<1>Bevan, M. et al.
<2>Analysis of 1.9 Mb of contiguous sequence from chromosome 4 of Arabidopsis thaliana.
<3>Nature
<4>391
<5>485-488
<6>1998
<7>The plant Arabidopsis thaliana has become an important model species for the study of many
aspects of plant biology.  The relatively small size of the nuclear genome and the
availability of extensive physical maps of the five chromosomes provide a feasible basis for
initiating sequencing of the five chromosomes.  The YAC-based physical map of chromosome 4 was
used to construct a sequence-ready map of cosmid and BAC clones covering a 1.9-megabase
contiguous region, and the sequence of this region is reported here.  Analysis of the sequence
revealed an average gene density of one gene every 4.8 kilobases, and 54% of the predicted
genes had significant similarity to known genes.  Other interesting features were found, such
as the sequence of a disease-resistance gene locus, the distribution of retroelements, the
frequent occurrence of clustered gene families, and the sequence of several classes of genes
not previously encountered in plants.

<>

<1>Bey, S.J., Tsou, M.F., Huang, C.H., Yang, C.C., Chen, C.W.
<2>The homologous terminal sequence of the Streptomyces lividans chromosome and SLP2 plasmid.
<3>Microbiology
<4>146
<5>911-922
<6>2000
<7>The chromosome of Streptomyces lividans shares 15.4 kb homology with one end of the linear
plasmid SLP2, consisting of a 10.1 kb terminal sequence
followed by the 5.3 kb transposable element Tn4811. The 10.1 kb terminal
sequence was determined. The mean G+C content of this sequence is 67.9
mol% with a striking G vs C bias in the last kb. The terminal 232 nt
contained 10 palindromic sequences with potential to form complex
secondary structures. One typical Streptomyces coding sequence (designated
ORF1) of 2643 bp was predicted in the determined sequence. The amino acid
sequence of the ORF1 product contained a DEAH helicase motif, and
exhibited similarity to type I restriction enzyme HsdR subunits in the
database, suggesting a possible role in replication of the telomeres.
However, all the ORF1 sequences on the chromosome and SLP2 could be
simultaneously knocked out by targeted recombination without affecting the
viability of the cells and the linearity of the chromosome and SLP2. This
ruled out ORF1 as an essential component in the maintenance of the linear
chromosome and plasmids.

<>

<1>Beyersmann, P.G., Chertkov, O., Petersen, J., Fiebig, A., Chen, A., Pati, A., Ivanova, N., Lapidus, A., Goodwin, L.A., Chain, P., Detter, J.C., Rohde, M., Gronow, S., Kyrpides, N.C., Woyke, T., Simon, M., Goker, M., Klenk, H.P., Brinkhoff, T.
<2>Genome sequence of Phaeobacter caeruleus type strain (DSM 24564(T)), a surface-associated member of the marine Roseobacter clade.
<3>Standards in Genomic Sciences
<4>8
<5>403-419
<6>2013
<7>In 2009 Phaeobacter caeruleus was described as a novel species affiliated with the marine
Roseobacter clade, which, in turn, belongs to the class
Alphaproteobacteria. The genus Phaeobacter is well known for members that produce
various secondary metabolites. Here we report of putative quorum sensing systems,
based on the finding of six N-acyl-homoserine lactone synthetases, and show that
the blue color of P. caeruleus is probably due to the production of the secondary
metabolite indigoidine. Therefore, P. caeruleus might have inhibitory effects on
other bacteria. In this study the genome of the type strain DSM 24564(T) was
sequenced, annotated and characterized. The 5,344,419 bp long genome with its
seven plasmids contains 5,227 protein-coding genes (3,904 with a predicted
function) and 108 RNA genes.

<>

<1>Beylot, B., Spassky, A.
<2>Chemical probing shows that the intron-encoded endonuclease I-SceI distorts DNA through binding in monomeric form to its homing site.
<3>J. Biol. Chem.
<4>276
<5>25243-25253
<6>2001
<7>Despite its small size (27.6 kDa), the group I intron-encoded I-SceI endonuclease initiates
intron homing by recognizing and specifically cleaving a large intronless DNA sequence. Here,
we used gel shift assays and footprinting experiments to analyze the interaction between
I-SceI and its target. I-SceI was found to bind to its substrate in monomeric form.
Footprinting using DNase I, hydroxyl radical, phenanthroline copper complexes, UV/DH-MePyPs
photosensitizer, and base-modifying reagents revealed the asymmetric nature of the interaction
and provided a first glimpse into the architecture of the complex. The protein interacts in
the minor and major grooves and distorts DNA at three distinct sites: one at the intron
insertion site and the other two, respectively, downstream (-8, -9) and upstream (+9, +10)
from this site. The protein appears to stabilize the DNA curved around it by bridging the
minor groove on one face of the helix. The scissile phosphates would lie on the outside of the
bend, facing in the same direction relative to the DNA helical axis, as expected for an
endonuclease that generates 3' overhangs. An internally consistent model is proposed in which
the protein would take advantage of the concerted flexibility of the DNA sequence to induce a
synergistic binding/kinking process, resulting in the correct positioning of the enzyme active
site.

<>

<1>Bhagwat, A.S.
<2>Restriction enzymes: Properties and Use.
<3>Methods Enzymol.
<4>216
<5>199-224
<6>1992
<7>Most experiments in molecular biology involve the use of restriction enzymes (REs) at some
stage of the experiment. The ability of these enzymes to cut DNA at a specific sequence of
bases has greatly stimulated the growth of recombinant DNA technology. The popularity of these
enzymes is, in part, due to their wide commercial availability and the simplicity of their
use. Over 1900 REs are known and of these 275 are available from companies based around the
world. The purpose of this chapter is to list all commercially available enzymes, to describe
their general properties, the basic procedures for their use, and ways of troubleshooting
problems encountered in their use. Although the number of enzymes being discussed is large, a
few simple rules apply to virtually all the enzymes. One of the aims of this chapter is to
show that most enzymes can be used successfully without the help of pre-made kits, colorful
charts, complex buffers, or specialized equipment. Some of this ground has been covered in
previous reviews by Fuchs and Blakesley, Brooks, and Roberts and Macelis.

<>

<1>Bhagwat, A.S., Beletskii, A.
<2>Transcription is intrinsically mutagenic: Direct correlation between transcription and C to U or 5meC to T deaminations in the non-transcribed strand.
<3>FASEB J.
<4>10
<5>A962
<6>1996
<7>Among mutations induced by external agents, more mutations occur in the
non-transcribed than the transcribed strand.  This strand bias occurs because of preferential
repair of the transcribed strand.  We have now shown that there is also a strand bias in the
spontaneous process of hydrolytic deamination of cytosine and 5-methylcytosine (5meC)
and it causes accumulation of C to T mutations in the non-transcribed strand.  To
demonstrate this, we used a genetic reversion system involving a transcriptionally regulated
kanamycin-resistance gene.  Induction of transcription increased the frequency of
conversion of target C to T only when it was present in the non-transcribed strand.  We
conclude that the mutations occur as a result of deamination of C or 5meC because
mismatch correction processes that repair U:G and T:G mismatches partially suppress the
mutations.  Further, methylation of target cytosine increases frequency of mutations -
consistent with the higher rate of hydrolytic deamination of 5meC compared to C.  The
non-transcribed strand suffers more deaminations probably because it is in single-stranded
state during transcription, and deamination of cytosines in single-stranded DNA is ~1000
times the rate in double-stranded DNA.  Because transcription requires separation of the
two DNA strands, we suggest that similar strand bias in hydrolytic deamination of C and
5meC must exist in all transcription events.

<>

<1>Bhagwat, A.S., Gabbara, S.
<2>The mechanism of activation of EcoRII endonuclease.
<3>FASEB J.
<4>6
<5>A281
<6>1992
<7>EcoRII is unusual among restriction enzymes in that it requires a high density
of its sites [sequence- 5'-CC(A/T)GG-3'] in a DNA for efficient cleavage.
Substrates such as phage T3 and T7 that contain only a few well separated sites
for the enzyme are cut poorly, while DNAs that contain several sites for the
enzyme are cut efficiently.  Interestingly, pBR322, pUC119 or short
oligonucleotide duplexes containing a single site for the enzyme can activate
the enzyme to cut poor substrates.  We are investigating the mechanism of this
activation using short oligonucleotide duplexes.  We have shown that activator
short duplexes are themselves cleaved poorly by the enzyme despite the fact
that the enzyme binds to the duplexes efficiently.  Thus, binding of the enzyme
to the substrate is not sufficient for cleavage.  As expected, pUC119 DNA can
activate EcoRII to cut these duplexes.  We have also shown that the inefficient
step in the reaction is not the release of the product after catalysis.
Interestingly, the efficiency of cleavage of a duplex increases with its
concentration in the reaction, ie. a duplex can act as its own activator.
Further, at high concentrations, a duplex can activate the enzyme to cut
another duplex.  Our results show that EcoRII endonuclease resembles
recombinational enzymes such as gamma/delta resolvase and phage lambda Int
protein in that it requires the simultaneous binding of two DNA molecules for
catalysis.

<>

<1>Bhagwat, A.S., Johnson, B., Weule, K., Roberts, R.J.
<2>Primary sequence of the EcoRII endonuclease and properties of its fusions with beta-galactosidase.
<3>J. Biol. Chem.
<4>265
<5>767-773
<6>1990
<7>The EcoRII endonuclease cleaves DNA containing the sequence CC(A/T)GG before the first
cytosine. The methylation of the second cytosine in the sequence by either the EcoRII
methylase or Dcm, a chromosomally coded protein in Escherichia coli, inhibits the cleavage.
The gene for the EcoRII endonuclease was mapped by analysis of derivatives containing linker
insertions, transposon insertions, and restriction fragment deletions. Surprisingly, plasmids
carrying the wild-type endonuclease gene and the EcoRII methylase gene interrupted by
transposon insertions appeared to be lethal to dcm+ strains of E. coli. We conclude that not
all the EcoRII/Dcm recognition sites in the cellular DNA are methylated in dcm+ strains. The
DNA sequence of a 1650-base pair fragment containing the endonuclease gene was determined. It
revealed an open reading frame that could code for a 45.6-kDa protein. This predicted size is
consistent with the known size of the endonuclease monomer (44 kDa). The endonuclease and
methylase genes appear to be transcribed convergently from separate promoters. The reading
frame of the endonuclease gene was confirmed at three points by generating random protein
fusions between the endonuclease and beta-galactosidase, followed by an analysis of the
sequence at the junctions. One of these fusions is missing 18 COOH-terminal amino acids of the
endonuclease but still displays significant ability to restrict incoming phage in addition to
beta-galactosidase activity. No striking similarity between the sequence of the endonuclease
and any other protein in the PIR data base was found. The knowledge of the primary sequence of
the endonuclease and the availability of the various constructs involving its gene should be
helpful in the study of the interaction of the enzyme with its substrate DNA.

<>

<1>Bhagwat, A.S., McClelland, M.
<2>DNA mismatch correction by very short patch repair may have altered the abundance of oligonucleotides in the E. coli genome.
<3>Nucleic Acids Res.
<4>20
<5>1663-1668
<6>1992
<7>A base mismatch correction process in E. coli K-12 called Very Short Patch (VSP) repair
corrects T:G mismatches to C:G when found in certain sequence contexts. Two of the substrate
mismatches (5'-CTWGG/3'-GGW'CC; W = A or T) occur in the context of cytosine methylation in
DNA and reduce the mutagenic effects of 5-methylcytosine deamination to thymine. However, VSP
repair is also known to repair T:G mismatches that are not expected to arise from
5-methylcytosine deamination (example--CTAG/GGT-C). In these cases, if the original base pair
were a T:A, VSP repair would cause a T to C transition. We have carried out Markov chain
analysis of an E. coli sequence database to determine if repair at the latter class of sites
has altered the abundance of the relevant tetranucleotides. The results are consistent with
the prediction that VSP repair would tend to deplete the genome of the 'T' containing
sequences (example--CTAG), while enriching it for the corresponding 'C' containing sequences
(CCAG). Further, they provide an explanation for the known scarcity of CTAG containing
restriction enzyme sites among the genomes of enteric bacteria and identify VSP repair as a
force in shaping the sequence composition of bacterial genomes.

<>

<1>Bhagwat, A.S., Person, S.
<2>Structure and properties of the region of homology between plasmids pMB1 and ColE1.
<3>Mol. Gen. Genet.
<4>182
<5>505-507
<6>1981
<7>Physical maps of the two independently isolated Escherichia coli plasmids, pMB1
and Co1E1, were prepared with 13 restriction endonucleases and compared.  A 5.1
kilobase continuous region covering 55% of pMB1 and 75% of Co1E1 was found to
have similar, but non-identical, restriction maps.  The differences in the maps
of this region probably arose by localized mutational events rather than by
major sequence rearrangements.  The F-factor was found to mobilize pMB1
efficiently for conjugal transfer.  A region on pMB1 required for its
F-mediated transfer was mapped.  Results of our study combined with results of
other investigators suggest that pMB1 and Co1E1 share functional properties
such as colicin production, colicin immunity, mode of replication, and
mobilization by the F-factor, and that the sequences required to code these
functions are contained within the 5.1 kilobase homologous region.  A
nomenclature for the genes encoding restriction and modification enzymes is
proposed.

<>

<1>Bhagwat, A.S., Roberts, R.J.
<2>Genetic analysis of the 5-azacytidine sensitivity of Escherichia coli K-12.
<3>J. Bacteriol.
<4>169
<5>1537-1546
<6>1987
<7>DNA containing 5-azacytidine (5-azaC) has been shown to form stable detergent-resistent
complexes with cytosine methylases. We reasoned that if 5-azaC treatment causes protein-DNA
cross-links in vivo, then mutations in DNA repair and recombination genes may increase the
sensitivity of a cell to 5-azaC. We found that although recA (defective) and lexA
(induction-negative) mutants of Escherichia coli were very sensitive to the drug, mutations in
uvrA and ung genes had little effect on cell lethality. The sensitivity of recA strains to
5-azaC was dose dependent and was enhanced by the overproduction of a DNA cytosine methylase
in the cell. Unexpectedly, a strain of E. coli carrying a recA mutation and a deletion of the
DNA cystosine methylase gene (dcm) was found to be significantly sensitive to 5-azaC. Study of
mutations in the pyrimidine salvage pathway of E. coli suggests that direct phosphorylation of
5-azaC, rather than phosphorylation of its degradation products, is largely responsible for
the lethal effects of the drug. The addition of uracil to the growth medium had little effect
on cell lethality of recA mutants, but it partially reversed the inhibition of cell growth
caused by 5-azaC. This reversal of the bacteriostatic effects of the drug could not be
achieved by adding cytosine or orotic acid to the growth medium and required the presence of
functional UMP-pyrophosphorylase (gene upp) in the cell.

<>

<1>Bhagwat, A.S., Sohail, A., Lieb, M.
<2>The ability of DCM and EcoRII methylases to methylate CC(A/T)GG sequences is not sufficient for their participation in very short patch repair at the same sites.
<3>J. Cell Biochem. Suppl.
<4>12A
<5>282
<6>1988
<7>E. coli has the ability to correct mismatches such as -CTAGG- -CTTGG- -GGTCC- -GGACC- to
restore CC(A/T)GG sites by a mechanism called very short patch (VSP) repair. It also codes for
a DNA cytosine methylase, Dcm, that methylates the second cytosine within CC(A/T)GG sequence
at its C5 position. Several mutations in dcm that lack the ability to methylate are defective
in VSP repair as well, suggesting a role for dcm in VSP repair. We have isolated a deletion
mutant of dcm that is active in methylation, but inactive in VSP repair. Another strain of E.
coli carries the restriction-modification system EcoRII that includes a DNA methylase with the
same sequence specificity as Dcm. The two enzymes methylate the same cytosine in the sequence
at the identical position, but EcoRII methylase cannot complement dcm mutations defective in
VSP repair. This is despite the fact that the two methylases share signficant sequence
homology. It appears that the two methylases have evolved to perform separate functions in the
cell.

<>

<1>Bhagwat, A.S., Sohail, A., Lieb, M.
<2>Methylation abilities of EcoRII and DCM methylases are not sufficient for their involvement in very short patch repair.
<3>UCLA Symp. Mol. Cell. Biol., Alan R. Liss, Inc., , New York
<4>83
<5>173-178
<6>1988
<7>Dcm, a methylase coded by a chromosomal gene in E. coli K-12, methylates the second cytosine
in the sequence CC(A/T)GG. E. coli can repair mismatches in DNA such as -CTAGG- or -CTTGG-
-GGTCC- -GGACC- in favor of the lower strand, creating CC(A/T)GG sites. This is called very
short patch (VSP) repair. Mutation dcm6 destroys the ability of the cell to methylate DNA and
to repair mismatches of the kind mentioned above. The methylase in the
restriction-modification system EcoRII recognizes and methylates the same sequence as Dcm. We
have found that the gene for the EcoRII methylase is closely related to dcm. Despite these
similarities, EcoRII methylase cannot complement the dcm6 mutation for VSP repair. We have
also studied the ability of certain deletion derivatives of the cloned dcm gene to complement
the dcm6 mutation. At least two deletions complement the mutation for methylation but not for
repair. A mutated methylase may lose a role in DNA repair without losing its methylating
ability.

<>

<1>Bhagwat, A.S., Sohail, A., Roberts, R.J.
<2>Cloning and characterization of the dcm locus of Escherichia coli K-12.
<3>J. Bacteriol.
<4>166
<5>751-755
<6>1986
<7>The dcm locus of Escherichia coli K-12 has been shown to code for a methylase that methylates
the second cytosine within the sequence 5'-CC(A/T)GG-3'. This sequence is also recognized by
the EcoRII restriction-modification system coded by the E. coli plasmid N3. The methylase
within the EcoRII system methylates the same cytosine as the dcm protein. We have isolated,
from a library of E. coli K-12 DNA, two overlapping clones that carry the dcm locus. We show
that the two clones carry overlapping sequences that are present in a dcm+ strain, but are
absent in a dcm deletion strain. We also show that the cloned gene codes for a methylase, that
it complements mutations in the EcoRII methylase, and that it protects EcoRII recognition
sites from cleavage by the EcoRII endonuclease. We found no phage restriction activity
associated with the dcm clones.

<>

<1>Bhalla, A., Kainth, A.S., Sani, R.K.
<2>Draft Genome Sequence of Lignocellulose-Degrading Thermophilic Bacterium Geobacillus sp. Strain WSUCF1.
<3>Genome Announcements
<4>1
<5>e00595-13
<6>2013
<7>Geobacillus sp. strain WSUCF1 is a thermophilic spore-forming member of the phylum Firmicutes,
isolated from a soil sample collected from the compost
facility. We report the draft genome sequence of this isolate with an estimated
genome size of 3.4 Mb. The genome sequence of this isolate revealed several genes
encoding glycoside hydrolases, making it a potential candidate for plant biomass
degradation.

<>

<1>Bhat, A., Tamuli, R., Kasbekar, D.P.
<2>Genetic Transformation of Neurospora tetrasperma, Demonstration of Repeat-Induced Point Mutation (RIP) in Self-Crosses and a Screen for Recessive RIP-Defective Mutants.
<3>Genetics
<4>167
<5>1155-1164
<6>2004
<7>The pseudohomothallic fungus Neurospora tetrasperma is naturally resistant
to the antibiotic hygromycin. We discovered that mutation of its erg-3
(sterol C-14 reductase) gene confers a hygromycin-sensitive phenotype that
can be used to select transformants on hygromycin medium by
complementation with the N. crassa erg-3+ and bacterial hph genes.
Cotransformation of hph with PCR-amplified DNA of other genes enabled us
to construct strains duplicated for the amplified DNA. Using
transformation we constructed self-fertile strains that were homoallelic
for an ectopic erg-3+ transgene and a mutant erg-3 allele at the
endogenous locus. Self-crosses of these strains yielded erg-3 mutant
ascospores that produced colonies with the characteristic morphology on
Vogel's sorbose agar described previously for erg-3 mutants of N. crassa.
The mutants were generated by repeat-induced point mutation (RIP), a
genome defense process that causes numerous G:C to A:T mutations in
duplicated DNA sequences. Homozygosity for novel recessive RIP-deficient
mutations was signaled by self-crosses of erg-3-duplication strains that
fail to produce erg-3 mutant progeny. Using this assay we isolated a
UV-induced mutant with a putative partial RIP defect. RIP-induced mutants
were isolated in rid-1 and sad-1, which are essential genes, respectively,
for RIP and another genome defense mechanism called meiotic silencing by
unpaired DNA.

<>

<1>Bhatia, U., Robison, K., Gilbert, W.
<2>Dealing with database explosion: A cautionary note.
<3>Science
<4>276
<5>1724-1725
<6>1997
<7>Carol J. Bult et al. Report the first entire archaea genome sequence of Methanococcus
jannashii (Mja).  Because the initial gene assignments were conservative, we anticipated that
much interesting biological information would be missing.  We searched the database for
additional open reading frames, and found 15 ORFs: four within intergenic regions (1 through
M4, Table 1); five overlapping with previously identified ORFs but that read off in a
different frame (M5 through M9, Table 1); and six that are extended or truncated as a result
of potential frameshifts (M10 through M15, Table 2).  Although the potential frameshifts we
describe might be bona fide, it cannot be ruled out that they represent actual sequencing
artifacts.  Erroneous sequences in public databases are a substantial problem and have been
estimated to be in the range of 0.37 to 2.9 errors per 1000 nucleotides, making data
interpretation sometimes difficult.  This is especially true, for example, in studies that
utilize protein and DNA sequence information to estimate evolutionary distances.  It is not
known how the error rate in this study compares with error rates in the database, but a
previous study suggests that error rates generally vary between 1 in 5000 to 1 in 10,000
nucleotides.

<>

<1>Bhattacharjee, S.K., Mahajan, S.K.
<2>The origin of adaptive mutations: Explaining the mutational spectra of Lac+ revertants of the Escherichia coli strain FC40.
<3>Curr. Sci.
<4>74
<5>583-590
<6>1998
<7>Reversion of lacI33-lacZ frameshift mutation to produce Lac+ colonies takes place by
demonstrably different mechanisms in growing and starving cultures of strains (such as FC40)
harboring this mutation.  The revertants appear to arise in starving cells only under
conditions where they can immediately promote growth.  This is reminiscent of Lamarckism.
Here we propose a mutagenic mechanism which is Darwinian inasmuch as it is blind to the
adaptive fitness of the mutants but can explain the mutational spectra seen in C40 and certain
other strains.  This mechanism can produce mutations only in the neighborhood of a
methylatable cytosine.

<>

<1>Bhattacharya, S., Dubey, A.K.
<2>Metabolic burden as reflected by maintenance coefficient of recombinant Escherichia coli overexpressing target gene.
<3>Biotechnol. Lett.
<4>17
<5>1155-1160
<6>1995
<7>Metabolic burden as a consequence of the overexpression of a target gene in a recombinant
strain of E. coli 1727 has been analyzed with respect to the maintenance energy coefficient
(m).  The values of 'm' for the host, uninduced recombinant and IPTG induced recombinant
were determined to be 0.12, 0.17 and 0.32 g.g-1.h-1 respectively.  Transient plasmid
instability and a nearly 33% fall in maximum specific growth rate were observed under
conditions of enhanced requirements for maintenance energy.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>Preparation and properties of restriction endonuclease SphI coupled to a nylon fabric filter.
<3>Biotechnol. Appl. Biochem.
<4>20
<5>141-146
<6>1994
<7>A procedure has been devised for covalent coupling of SphI, a type II restriction endonuclease
(EC 3.1.2.1.4) from Streptomyces phaeochromogenes, to a nylon-fabric filter.  Immobilized SphI
preparations were characterized with respect to operational parameters, re-usability and
stability during storage.  It is shown that immobilization of SphI on nylon filter discs did
not affect the enzymic behavior of the enzyme, but did enhance its thermal stability.  The
rates of inactivation of free and bound SphI at 45oC were determined to be 5.0 h-1 and 3.0 h-1
respectively.  The membrane-bound preparations could be stored for over 2 months at 4oC
without significant loss of activity, compared with free enzyme, which had lost most of its
activity during this period.  The insolubilized R. SphI could be used up to three times.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>Expression parameters for target gene cloned in Escherichia coli in response to phosphate supply.
<3>Biotechnol. Lett.
<4>18
<5>1145-1148
<6>1996
<7>A recombinant strain of E. coli 1727 overexpressed a target gene at enhanced levels when
supplied with an excess of inorganic phosphate.  The rate of target gene expression, the yield
coefficient for the target gene product and the plasmid copy number increased significantly:
50%, 100% and 40% respectively at 125 mM excess phosphate.  This was, however, accompanied by
a 26% decrease in the specific growth rate and 30% in the cellular growth yield coefficient.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>Effects of dissolved oxygen and oxygen mass transfer on overexpression of target gene in recombinant E. coli.
<3>Enzyme Microb. Technol.
<4>20
<5>355-360
<6>1997
<7>Overexpression of target gene (MspI methylase) in recombinant Escherichia coli has been
studied in response to dissolved oxygen mass transfer.  The rate of target gene expression
attained its maximum value (2.67 U mg^-1p^-1h^-1) when the culture was induced with 1 mM
isopropyl-beta-D-thiogalactopyranoside (IPTG) in the mid-exponential phase of growth.  The
maximum value of 6.7 x 10^2 U mg^-1p^-1 for the target gene expression was attained in about 3
h of postinduction cultivation.  An oxygen-sufficient condition (0.22 mmoles l^-1) was
necessary to obtain optimal expression and cell productivity.  Expression of the target gene
upon induction was accompanied by an increase in oxygen uptake rate (OUR) of the cells and
decrease in dissolved oxygen (DO) concentration in the cultivation broth.  Volumetric oxygen
transfer coefficient (Kla) of 61 h^-1 was optimum for cell productivity of uninduced culture
while that for the induced culture was 84 h^-1 which was optimum for foreign protein synthesis
also.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>High-level expression of a heterologous gene in Escherichia coli in response to carbon-nitrogen source and C/N ration in a batch bioreactor.
<3>Biotechnol. Prog.
<4>13
<5>151-155
<6>1997
<7>Overexpression of the target gene in a recombinant strain of Escherichia coli has been
analyzed in view of changes in the carbon-nitrogen source and as a function of C/N ratio.  The
rate of target gene expression varied over a range of 200%, and the yield coefficient for the
target gene product changed over 400% with change in carbon source at 10 gL^-1 in M9 medium.
With variation in nitrogen, source, however, only marginal changes (10-15%) occurred in these
parameters of overexpression.  Higher concentration, for example 2.5 g L^-1, of any nitrogen
source adversely affected heterologous expression as rate of target gene expression declined
in the range 35%-50% and the yield coefficient of foreign protein decreased between 45% and
60%.  A C/N ratio of 15 was found to be optimum for both parameters for overexpression and
host specific parameters such as specific growth rate and observed yield coefficient for
cellular growth.  An analysis with respect to temperature and pH revealed that host and
expression parameters were quite susceptible to changes in these factors.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>Kinetic mechanism of cytosine DNA methyltransferase MspI.
<3>J. Biol. Chem.
<4>274
<5>14743-14749
<6>1999
<7>A kinetic analysis of MspI DNA methyltransferase (M.MspI) is presented. The enzyme catalyzes
methylation of lambda-DNA, a 50-kilobase pair linear molecule with multiple M.MspI-specific
sites, with a specificity constant (kcat/KM) of 0.9 x 10^8 M-1 s-1. But the values of the
specificity constants for the smaller DNA substrates (121 and 1459 base pairs (bp)) with
single methylation target or with multiple targets (sonicated lambda-DNA) were less by an
order of magnitude. Product inhibition of the M.MspI-catalyzed methylation reaction by
methylated DNA is competitive with respect to DNA and noncompetitive with respect to
S-adenosylmethionine (AdoMet). The S-adenosylhomocysteine inhibition of the methylation
reaction is competitive with respect to AdoMet and uncompetitive with respect to DNA. The
presteady state kinetic analysis showed a burst of product formation when AdoMet was added to
the enzyme preincubated with the substrate DNA. The burst is followed by a constant rate of
product formation (0.06 mol per mol of enzyme s-1) which is similar to catalytic constants
(kcat = approximately 0.056 s-1) measured under steady state conditions. The isotope exchange
in chasing the labeled methyltransferase-DNA complex with unlabeled DNA and AdoMet leads to a
reduced burst as compared with the one involving chase with labeled DNA and AdoMet. The enzyme
is capable of exchanging tritium at C-5 of target cytosine in the substrate DNA in the absence
of cofactor AdoMet. The kinetic data are consistent with an ordered Bi Bi mechanism for the
M.MspI-catalyzed DNA methylation where DNA binds first.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>The N-terminus of m5C-DNA methyltransferase MspI is involved in its topoisomerase activity.
<3>Eur. J. Biochem.
<4>269
<5>2491-2497
<6>2002
<7>DNA cytosine methyltransferase MspI (M.MspI) must require a different type of interaction of
protein with DNA from other bacterial DNA
cytosine methyltransferases (m5C-MTases) to evoke the topoisomerase
activity that it possesses in addition to DNA-methylation ability. This
may require a different structural organization in the solution phase
from the reported consensus structural arrangement for m5C-MTases.
Limited proteolysis of M.MspI, however, generates two peptide
fragments, a large one (p26) and a small one (p18), consistent with
reported m5C-MTase structures. Examination of the amino-acid sequence
of M.MspI revealed similarity to human topoisomerase I at the
N-terminus. Alignment of the amino-acid sequence of M.MspI also
uncovered similarity (residues 245-287) to the active site of human DNA
ligase I. To evaluate the role of the N-terminus of M.MspI,
2-hydroxy-5-nitrobenzyl bromide (HNBB) was used to truncate M.MspI
between residues 34 and 35. The purified HNBB-truncated protein has a
molecular mass of approximately 45 kDa, retains DNA binding and
methyltransferase activity, but does not possess topoisomerase
activity. These findings were substantiated using a purified
recombinant MspI protein with the N-terminal 34 amino acids deleted.
Changing the N-terminal residues Trp34 and Tyr74 to alanine results in
abolition of the topoisomerase I activity while the methyltransferase
activity remains intact.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>Transient DNA binding by a proteolytic peptide from m5C-DNA methyltransferase MspI.
<3>J. Biochem. Mol. Biol. Biophys.
<4>6
<5>357-364
<6>2002
<7>A peptide fragment (p26) generated as a result of limited tryptic proteolysis of
methyltransferase MspI retains transient but non-specific DNA binding capability. The
transient DNA binding by p26 was characterized with respect to physicochemical factors.
Limited proteolysis was performed to probe gross structural deviation from the reported
two-domain organization for m5C-MTases, in light of topoisomerase activity shown by MspI,
resulted in two peptide fragments; a large fragment p26 and a small fragment p18, consistent
with the other reported m5C-MTase structures. The purified large peptide fragment p26, spans
between 6 and 251 in the amino acid sequence of M.MspI. The peptide p26 does not bind
S-adenosylmethionine, although in the intact protein the AdoMet binding region can be mapped
to a region in the protein that is present in this peptide. Such transient DNA binding has not
been reported for other protolytic product of any other m5C-DNA methyltransferase.

<>

<1>Bhattacharya, S.K., Dubey, A.K.
<2>Amino acid residues involved in catalysis and DNA binding by cytosine DNA methyltransferase MspI.
<3>J. Biochem. Mol. Biol. Biophys.
<4>4
<5>109-120
<6>2000
<7>Chemical modifications of chosen amino acid residues: arginine, histidine, cysteine, glycine
and tryptophan have been attempted to investigate their role in catalysis and DNA binding by
MspI DNA methyltransferase (M.MspI).  The catalytic inactivation following modifications of
two arginine and three histidine moieties was a consequence of loss of DNA binding property of
the methyltransferase.  Modification of a single cysteine residue completely abolished the
ability of M.MspI to transfer a methyl group but there was no loss of DNA binding activity.
While tryptophan had no apparent functional role, glycine was critical for AdoMet binding and
target base methylation by the M.MspI.

<>

<1>Bhattacharya, S.K., Ramchandani, S., Cervoni, N., Szyf, M.
<2>A mammalian protein with specific demethylase activity for mCpG DNA.
<3>Nature
<4>397
<5>579-583
<6>1999
<7>DNA-methylation patterns are important for regulating genome functions, and are determined by
the enzymatic processes of methylation and demethylation.  The demethylating enzyme has now
been identified; a mammalian complementary DNA encodes a methyl-CpG-binding domain, bears a
demethylase activity that transforms methylated cytosine bases to cytosine, and demethylates a
plasmid when the cDNA is translated or transiently transfected into human embryonal kidney
cells in vitro.  The discovery of this DNA demethylase should provide a basis for the
molecular and developmental analysis of the role of DNA methylation and demethylation.

<>

<1>Bhave, N., Woodhead, J.L., Malcolm, A.D.B.
<2>Cation-dependence of the restriction endodeoxyribonuclease EcoRI.
<3>Biochem. Soc. Trans.
<4>8
<5>634
<6>1980
<7>The restriction endodeoxyribonucleases and DNA methylases provide excellent examples for the
enzymologist of highly specific protein-nucleic acid interactions.  The endodeoxyribonuclease
EcoRI is comparatively easy to prepare, and is therefore among the best studied of these
restriction enzymes.  It is unusual (although not unique) in that its recognition sequence
depends on the exact assay conditions.  At low pH and high ionic strength the enzyme
recognizes the DNA sequence.

<>

<1>Bheemanaik, S., Bujnicki, J.M., Nagaraja, V., Rao, D.N.
<2>Functional analysis of amino acid residues at the dimerisation interface of Kpnl DNA methyltransferase.
<3>Biol. Chem.
<4>387
<5>515-523
<6>2006
<7>KpnI DNA-(N-6-adenine) methyltransferase (M.Kpnl) recognises the sequence 5'-GGTACC-3' and
transfers the methyl group from
S-adenosyl-L-methionine (AdoMet) to the N6 position of the adenine
residue in each strand. Earlier studies have shown that M.Kpnl exists
as a dimer in solution, unlike most other MTases. To address the
importance of dimerisation for enzyme function, a three-dimensional
model of M.Kpnl was obtained based on protein fold-recognition
analysis, using the crystal structures of M.Rsrl and M.MbollA as
templates. Residues l146, l161 and Y167, the side chains of which are
present in the putative dimerisation interface-in the model, were
targeted for site-directed mutagenesis. Methylation and in vitro
restriction assays showed that the mutant MTases are catalytically
inactive. Mutation at the l146 position resulted in complete disruption
of the dimer. The replacement of l146 led to drastically reduced DNA
and cofactor binding. Substitution of l161 resulted in weakening of the
interaction between monomers, leading to both monomeric and dimeric
species. Steady-state fluorescence measurements showed that the
wild-type KpnI MTase induces structural distortion in bound DNA, while
the mutant MTases do not. The results establish that monomeric MTase is
catalytically inactive and that dimerisation is an essential event for
M.Kpnl to catalyse the methyl transfer reaction.

<>

<1>Bheemanaik, S., Chandrashekaran, S., Nagaraja, V., Rao, D.N.
<2>Kinetic and catalytic properties of dimeric KpnI DNA methyltransferase.
<3>J. Biol. Chem.
<4>278
<5>7863-7874
<6>2003
<7>KpnI DNA-(N(6)-adenine)-methyltransferase (KpnI MTase) is a member of a
restriction-modification (R-M) system in Klebsiella pneumoniae and
recognizes the sequence 5'-GGTACC-3'. It modifies the recognition sequence
by transferring the methyl group from S-adenosyl-l-methionine (AdoMet) to
the N(6) position of adenine residue. KpnI MTase occurs as a dimer in
solution as shown by gel filtration and chemical cross-linking analysis.
The nonlinear dependence of methylation activity on enzyme concentration
indicates that the functionally active form of the enzyme is also a dimer.
Product inhibition studies with KpnI MTase showed that
S-adenosyl-l-homocysteine is a competitive inhibitor with respect to
AdoMet and noncompetitive inhibitor with respect to DNA. The methylated
DNA showed noncompetitive inhibition with respect to both DNA and AdoMet.
A reduction in the rate of methylation was observed at high concentrations
of duplex DNA. The kinetic analysis where AdoMet binds first followed by
DNA, supports an ordered bi bi mechanism. After methyl transfer,
methylated DNA dissociates followed by S-adenosyl-l-homocysteine.
Isotope-partitioning analysis showed that KpnI MTase-AdoMet complex is
catalytically active.

<>

<1>Bheemanaik, S., Reddy, Y.V.R., Rao, D.N.
<2>Structure, function and mechanism of exocyclic DNA methyltransferases.
<3>Biochem. J.
<4>399
<5>177-190
<6>2006
<7>DNA MTases (methyltransferases) catalyse the transfer of methyl groups to DNA from AdoMet
(S-adenosyl-L-methionine) producing AdoHcy
(S-adenosyl-L-homocysteine) and methylated DNA. The C-5 and N-4
positions of cytosine and N-6 position of adenine are the target sites
for methylation. All three methylation patterns are found in
prokaryotes, whereas cytosine at the C-5 position is the only
methylation reaction that is known to occur in eukaryotes. In general,
MTases are two-domain proteins comprising one large and one small
domain with the DNA-binding cleft located at the domain interface. The
striking feature of all, the structurally characterized DNA MTases is
that they share a common core structure referred to as an
'AdoMet-dependent MTase fold'. DNA methylation has been reported to be
essential for bacterial virulence, and it has been suggested that DNA
adenine MTases (Dams) could be potential targets for both vaccines and
antimicrobials. Drugs that block Dam could slow down bacterial growth
and therefore drug-design initiatives could result in a whole new
generation of antibiotics. The transfer of larger chemical entities in
A MTase-catalysed reaction has been reported and this represents an
interesting challenge for bio-organic chemists. In general, amino
MTases could therefore be used as delivery systems for fluorescent or
other reporter groups on to DNA. This is one of the potential
applications of DNA MTases towards developing non-radioactive DNA
probes and these could have interesting applications in molecular
biology. Being nucleotide-sequence-specific, DNA MTases provide
excellent model systems for studies on protein-DNA interactions. The
focus of this review is on the chemistry, enzymology and structural
aspects of exocyclic amino MTases.

<>

<1>Bheemanaik, S., Reddy, Y.V.R., Rao, D.N.
<2>Structure, function and mechanism of exocyclic DNA methyltransferases (vol 399, pg 177, 2006).
<3>Biochem. J.
<4>399
<5>543
<6>2006
<7>In Table 2, the Kd(DNA) values for the enzyme M.EcoRI were incorrect.  The correct table
appears here.

<>

<1>Bheemanaik, S., Sistla, S., Krishnamurthy, V., Sampath, A., Desirazu, N.R.
<2>Kinetics of methylation by EcoP1I DNA methyltransferase.
<3>Enzyme Res.
<4>2010
<5>302731
<6>2010
<7>EcoP1I DNA MTase (M.EcoP1I), an N6-adenine MTase from bacteriophage P1, is a part of the
EcoP1I restriction-modification
(R-M) system which belongs to the Type III R-M system. It recognizes the sequence
5'-AGACC-3' and methylates the internal adenine. M.EcoP1I requires Mg2+ for the transfer of
methyl groups to DNA. M.EcoP1I is shown to exist as dimer in solution, and even at high salt
concentrations (0.5M) the dimeric M.EcoP1I does not dissociate into monomers suggesting a
strong interaction
between the monomer subunits. Preincubation and isotope partitioning studies with M.EcoP1I
indicate a kinetic mechanism where the duplex DNA binds first followed by AdoMet.
Interestingly, M.EcoP1I methylates DNA substrates in the presence of Mn2+ and Ca2+ other than
Mg2+ with varying affinities. Amino acid analysis and methylation assays in the presence of
metal ions suggest that M.EcoP1I has indeed two metal ion-binding sites [358ID(x)n . . .
ExK401 and 600DxDxD604 motif]. EcoP1I DNA MTase catalyzes the transfer of methyl groups using
a distributive mode of methylation on DNA containing more than one recognition site. A
chemical modification of EcoP1I DNA MTase using N-ethylmaleimide resulted in an irreversible
inactivation of enzyme activity suggesting the possible role of cysteine residues in
catalysis.

<>

<1>Bhugra, B., Voelker, L.L., Zou, N., Yu, H., Dybvig, K.
<2>Mechanism of antigenic variation in Mycoplasma pulmonis: interwoven, site-specific DNA inversions.
<3>Mol. Microbiol.
<4>18
<5>703-714
<6>1995
<7>The chromosome of the murine pathogen Mycoplasma pulmonis undergoes rearrangements at a high
frequency.  We show that some of these rearrangements regulate the phase-variable expression
of a cluster of genes (the vsa locus) that encode the variable V-1 surface antigens.  Only one
vsa gene is associated with an expression site; the other vsa genes are transcriptionally
silent.  The silent genes lack the 5' end region (promoter and ribosome-binding site) that is
present in the expressed gene, and DNA rearrangements regulate gene expression by reassorting
the 5' end region from an expressed gene with the 3' end region from a previously silent
gene.  All vsa rearrangements identified so far are site-specific DNA inversions that occur
between copies of a specific 34bp sequence that is conserved in each vsa gene.  Interestingly,
DNA inversions within the vsa locus apparently occur in concert with inversion of the hsd1
element, which regulates restriction and modification activity in M. pulmonis.

<>

<1>Bian, F., Qin, Q.L., Xie, B.B., Shu, Y.L., Zhang, X.Y., Yu, Y., Chen, B., Chen, X.L., Zhou, B.C., Zhang, Y.Z.
<2>Complete Genome Sequence of Seawater Bacterium Glaciecola nitratireducens FR1064T.
<3>J. Bacteriol.
<4>193
<5>7006-7007
<6>2011
<7>Glaciecola nitratireducens strain FR1064(T) was isolated from seawater and described as a new
species by Baik et al. in 2006. The genome size is
about 1.01 to 1.26 Mb smaller than two reported Glaciecola genomes,
indicating the gain or loss of large genome segments in the evolution of
Glaciecola strains.

<>

<1>Bian, F., Xie, B.B., Qin, Q.L., Shu, Y.L., Zhang, X.Y., Yu, Y., Chen, B., Chen, X.L., Zhou, B.C., Zhang, Y.Z.
<2>Genome sequences of six pseudoalteromonas strains isolated from arctic sea ice.
<3>J. Bacteriol.
<4>194
<5>908-909
<6>2012
<7>Yu et al. (Polar Biol. 32:1539-1547, 2009) isolated 199 Pseudoalteromonas strains from Arctic
sea ice. We sequenced the genomes of six of these
strains, which are affiliated to different Pseudoalteromonas species based
on 16S rRNA gene sequences, facilitating the study of physiology and
adaptation of Arctic sea ice Pseudoalteromonas strains.

<>

<1>Bian, J., Li, C.
<2>Disruption of a Type II Endonuclease (TDE0911) Enables Treponema denticola ATCC 35405 To Accept an Unmethylated Shuttle Vector.
<3>Appl. Environ. Microbiol.
<4>77
<5>4573-4578
<6>2011
<7>The oral spirochete Treponema denticola is associated with human periodontal disease. T.
denticola ATCC 35405 and ATCC 33520 are two
routinely used laboratory strains. Compared to T. denticola ATCC 33520,
ATCC 35405 is more virulent but less accessible to genetic
manipulations. For instance, the shuttle vectors of ATCC 33520 cannot
be transformed into strain ATCC 35405. The lack of a shuttle vector has
been a barrier to study the biology and virulence of T. denticola ATCC
35405. In this report, we hypothesize that T. denticola ATCC 35405 may
have a unique DNA restriction-modification (R-M) system that prevents
it from accepting the shuttle vectors of ATCC 33520 (e. g., the shuttle
plasmid pBFC). To test this hypothesis, DNA restriction digestion, PCR,
and Southern blot analyses were conducted to identify the differences
between the R-M systems of these two strains. DNA restriction digestion
analysis of these strains showed that only the cell extract from ATCC
35405 was able to digest pBFC. Consistently, PCR and Southern blot
analyses revealed that the genome of T. denticola ATCC 35405 encodes
three type II endonucleases that are absent in ATCC 33520. Among these
three endonucleases, TDE0911 was predicted to cleave unmethylated
double-stranded DNA and to be most likely responsible for the cleavage
of unmethylated pBFC. In agreement with this prediction, the mutant of
TDE0911 failed to cleave unmethylated pBFC plasmid, and it could accept
the unmethylated shuttle vector. The study described here provides us
with a new tool and strategy to genetically manipulate T. denticola, in
particular ATCC 35405, and other strains that may carry similar
endonucleases.

<>

<1>Bianco, P.R., Hurley, E.M.
<2>The type I restriction endonuclease EcoR124I, couples ATP hydrolysis to bidirectional DNA translocation.
<3>J. Mol. Biol.
<4>352
<5>837-859
<6>2005
<7>Type I restriction endonuclease holoenzymes contain methylase (M), restriction (R) and
specificity (S) subunits, present in an M-2:R-2:S-1
stoichiometry. These enzymes bind to specific DNA sequences and
translocate dsDNA in an ATP-dependent manner toward the holoenzyme
anchored at the recognition sequence. Once translocation is impeded,
DNA restriction, which functions to protect the host cell from invading
DNA, takes place. Translocation and DNA cleavage are afforded by the
two diametrically opposed R-subunits. To gain insight into the
mechanism of translocation, a detailed characterization of the ATPase
activity of EcoR124I was done. Results show that following recognition
sequence binding, ATP hydrolysis-coupled, bidirectional DNA
translocation by EcoR124I ensues, with the R-subunits transiently
disengaging, on average, every 515 bp. Macroscopic processivity of
2031( 184) bp is maintained, as the R-subunits remain in close
proximity to the DNA through association with the methyltransferase.
Transient uncoupling of ATP hydrolysis from translocation results in
3.1( 0.4) ATP molecules being hydrolyzed per base-pair translocated
per R-subunit. This is the first clear demonstration of the coupling of
ATP hydrolysis to dsDNA translocation, albeit inefficient. Once
translocation is impeded on supercoiled DNA, the DNA is cleaved. DNA
cleavage inactivates the EcoR124I holoenzyme partially and reversibly,
which explains the stoichiometric behaviour of type I restriction
enzymes. Inactivated holoenzyme remains bound to the DNA at the
recognition sequence and immediately releases the nascent ends. The
release of nascent ends was demonstrated using a novel,
fluorescence-based, real-time assay that takes advantage of the ability
of the Escherichia coli RecBCD enzyme to unwind restricted dsDNA. The
resulting unwinding of EcoR124I-restricted DNA by RecBCD reveals
coordination between the restriction-modification and recombination
systems that functions to destroy invading DNA efficiently. In
addition, we demonstrate the displacement of EcoR124I following DNA
cleavage by the translocating RecBCD enzyme, resulting in the
restoration of catalytic function to EcoR124I.

<>

<1>Bianco, P.R., Xu, C., Chi, M.
<2>Type I restriction endonucleases are true catalytic enzymes.
<3>Nucleic Acids Res.
<4>37
<5>3377-3390
<6>2009
<7>Type I restriction endonucleases are intriguing, multifunctional complexes that restrict DNA
randomly, at sites distant from the target sequence.
Restriction at distant sites is facilitated by ATP hydrolysis-dependent,
translocation of double-stranded DNA towards the stationary enzyme bound
at the recognition sequence. Following restriction, the enzymes are
thought to remain associated with the DNA at the target site, hydrolyzing
copious amounts of ATP. As a result, for the past 35 years type I
restriction endonucleases could only be loosely classified as enzymes
since they functioned stoichiometrically relative to DNA. To further
understand enzyme mechanism, a detailed analysis of DNA cleavage by the
EcoR124I holoenzyme was done. We demonstrate for the first time that type
I restriction endonucleases are not stoichiometric but are instead
catalytic with respect to DNA. Further, the mechanism involves formation
of a dimer of holoenzymes, with each monomer bound to a target sequence
and, following cleavage, each dissociates in an intact form to bind and
restrict subsequent DNA molecules. Therefore, type I restriction
endonucleases, like their type II counterparts, are true enzymes. The
conclusion that type I restriction enzymes are catalytic relative to DNA
has important implications for the in vivo function of these previously
enigmatic enzymes.

<>

<1>Bickle, T., Arber, W.
<2>Host-controlled restriction and modification of filamentous I- and F-specific bacteriophages.
<3>Virology
<4>39
<5>605-607
<6>1969
<7>In the study of strain-specific DNA restriction and modification, the DNA
molecules of small bacteriophages have proved very useful, particularly those
possessing only one sitie of affinity for the enzymes of a given restriction
and modification system.  The male specific phage fd and other small phages
closely related to fd undergo host-controlled modification in E. coli B, and
some are also modified in P1-lysogenic strains.  However, they are not
restricted in E. coli K12 or in strains carrying two other specificity systems,
as shown here, probably because their DNA lacks the appropriate specificity
sites.  With the aim of finding small phages restricted by host specificity
systems to which fd does not respond, we have screened other filamentous
bacteriophages for their responses to these sytems, including the two
I-specific phages If1 and If2 and two additional F-specific phages, Ec9 and HR.

<>

<1>Bickle, T.A.
<2>The ATP-dependent restriction endonucleases.
<3>Nucleases, Cold Spring Harbor Laboratory Press, Linn, S.M., Roberts, R.J., New York
<4>0
<5>85-108
<6>1982
<7>None

<>

<1>Bickle, T.A.
<2>Adenosine triphosphate-requiring restriction enzymes.
<3>Biochem. Soc. Trans.
<4>8
<5>396-397
<6>1980
<7>None

<>

<1>Bickle, T.A.
<2>DNA restriction and modification systems.
<3>Escherichia coli and Salmonella typhimurium: Cellular and molecular biology., Amer. Soc. Microbiology, Neidhardt, F.C., Washington
<4>1
<5>692-696
<6>1987
<7>The phenomenon of DNA restriction and modification was discovered in
Escherichia coli more than three decades ago by Bertani and Weigle in the
course of experiments with the phages P2 and lambda.  They found that phages
grown on E. coli K-12 plated with equal efficiency on strains K-12 and C, but
the converse was not true:  phage grown on E. coli C plated with high
efficiency on E. coli C but poorly on E. coli K-12.  Moreover, a single cycle
of growth in E. coli C was sufficient to remove the ability to grow well in E.
coli K-12.  The molecular explanation for this Lamarckian effect came from a
series of experiments conducted in Arber's laboratory in the 1960s.  The DNA of
phage grown in E. coli C is degraded by a DNA sequence-specific endonuclese
present in E. coli K-12.  When the phage are grown in E. coli K-12, the DNA is
protected from the endonuclease because a DNA methylase present in E. coli K-12
and absent from E. coli C methylates the sequences recognized by the
endonuclease, rendering them resistant to digestion.  Arber coined the term
host-controlled restriction and modification of DNA to describe this
phenomenon.  The endonuclease involved is a restriction endonuclease, and the
methylase is a modification methylase.  The primary function of the
modification methylase is to protect the cell's own genome from restriction.
Restriction and modification systems (R-M systems) are common throughout the
procaryotic world, and this chapter will focus on those systems that have been
found in E. coli, Salmonella typhimurium, and their close relatives.  Several
comprehensive reviews on restriction and modification have been published
recently.  These reviews should be consulted for details of the enzymology of
R-M systems and for references to the early literature.

<>

<1>Bickle, T.A.
<2>The ATP-dependent restriction enzymes.
<3>Nucleases, Cold Spring Harbor Laboratory Press, Linn, S.M., Lloyd, R.S., Roberts, R.J., Cold Spring Harbor
<4>0
<5>89-109
<6>1993
<7>*
  I. Introduction

 II. Type I restriction enzymes
        A. Occurrences
        B. Structural genes
        C. Family relationships among type I restriction enzymes
            1. DNA and protein sequence homologies within families
            2. DNA and protein sequence homologies between families
        D. Evolution of DNA sequence specificity in type I enzymes
            1. By homologous recombination
            2. By unequal crossing over
            3. By transposition
        E. Enzyme structure and mechanisms
            1. Enzyme structures
            2. Enzyme mechanisms

III. Type III restriction enzymes
        A. Occurrence
        B. Genetics
        C. Enzyme mechanism
        D. DNA cleavage

 IV. Concluding remarks


<>

<1>Bickle, T.A.
<2>Phage introns on the hop: an alternative view.
<3>Curr. Biol.
<4>2
<5>150-151
<6>1992
<7>The nuclease encoded by a rare bacteriophage intron is very similar to one class of bacterial
restriction enzyme, and may provide clues as to the co-evolution of host and phage.

<>

<1>Bickle, T.A.
<2>Restricting restriction.
<3>Mol. Microbiol.
<4>51
<5>3-5
<6>2003
<7>Systems biology is a new, fashionable and well-funded discipline, which to quote from a recent
review aims to 'examine the structure and dynamics of cellular and organismal function, raher
than the characteristics of isolated parts of a cell or organism . . . '.  Systems biology
will do this by profiting from the vast amounts of biological information that are available
in the genomics era and make extensive use of computer modelling.  But: 'many breakthroughs
in experimental devices, advanced software and analytical methods are required before the
achievements of system biology can live up to their much-touted potential'.  This edition of
Molecular Microbiology contains a paper that is the product of traditional experimental
biology but which could serve as a test case for systems biology.  The paper shows how
bacteria integrate such disparate subsystems as DNA restriction, homologous recombination and
regulated proteolysis to protect their chromosomes from degradation.  When systems biology can
predict this level of choreography, it will be a mature discipline.

<>

<1>Bickle, T.A., Brack, C., Yuan, R.
<2>ATP-induced conformational changes in the restriction endonuclease from Escherichia coli K-12.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>75
<5>3099-3103
<6>1978
<7>ATP induces a conformational change in the Escherichia coli K-12 restriction
enzyme that allows it to discriminate between unmodified and modified DNA
recognition sequences.  This conformational change does not require ATP
hydrolysis.  However, ATP hydrolysis is a requirement for DNA cleavage.

<>

<1>Bickle, T.A., Ineichen, K.
<2>The DNA sequence recognised by BglI.
<3>Gene
<4>9
<5>205-212
<6>1980
<7>The restriction enzyme BglI recognizes the DNA sequence: 5'-GCCNNNN^NGGC-3'
3'-CGGN^NNNNCCG-5' and cleaves it in the position shown by the arrows to leave
3' single-stranded protrusions three bases long.

<>

<1>Bickle, T.A., Kruger, D.H.
<2>Biology of DNA Restriction.
<3>Microbiol. Rev.
<4>57
<5>434-450
<6>1993
<7>*
Introduction
Type I R-M Systems
Type I Systems Form Families of Related Enzymes
Structure of hsd Genes
Evolution of DNA Sequence Recognition by Recombination between hsdS Genes
Mutations Affecting Modification Activity
Type II R-M Systems
Evolutionary Aspects
Control of Expression of Type II RM Genes
Cytosine Can Be Methylated on Either C-5 or N4: Consequences for Mutagenesis
Type II Restriction Endonuclease That Require Two Recognition Sites for Cleavage
What is the Function of Type IIS Enzymes?
Type III R-M Systems
Genetics of Type III Systems
DNA Recognition by Type III Enzymes:
   Different Sequence Requirements for Restriction and Modification
Sty LTI System
Restriction Systems Specific and Modified DNA
DpnI and DpnII
Rediscovery of Methyl-Directed Restriction in E. coli
McrBC
McrA
Mrr
Phage Antirestriction
Recent Developments
"Nature Red in Tooth and Claw": The case of Phage T4
Concluding Remarks
Acknowledgments
References


<>

<1>Bickle, T.A., Pirrotta, V., Imber, R.
<2>Purification and properties of the BglI and II endonucleases.
<3>Methods Enzymol.
<4>65
<5>132-138
<6>1980
<7>The presence of two distinct restriction endonucleases in Bacillus globigii,
BglI and II, was first reported by Wilson and Young.

<>

<1>Bickle, T.A., Pirrotta, V., Imber, R.
<2>A simple, general procedure for purifying restriction endonucleases.
<3>Nucleic Acids Res.
<4>4
<5>2561-2572
<6>1977
<7>A simple, general method for purifying restriction endonucleases is described.
The method employs precipitation of nucleic acids from crude extracts with
polyethyleneimine followed by affinity chromatography on columns of heparin
covalently linked to agarose.  Most of the sixteen enzymes tested could be
purified to a degree sufficient for DNA sequencing work by this method
sometimes supplemented by at most one step of ion exchange chromatography.

<>

<1>Bickle, T.A., Streiff, M.
<2>Methods for purification of restriction enzymes.
<3>Nucleic Acid Biochem.
<4>B510
<5>1-8
<6>1983
<7>None

<>

<1>Biddle, A.S. et al.
<2>The complete genome sequence of Clostridium indolis DSM 755(T.).
<3>Standards in Genomic Sciences
<4>9
<5>1089-1104
<6>2014
<7>Clostridium indolis DSM 755(T) is a bacterium commonly found in soils and the feces of birds
and mammals. Despite its prevalence, little is known about the
ecology or physiology of this species. However, close relatives, C.
saccharolyticum and C. hathewayi, have demonstrated interesting metabolic
potentials related to plant degradation and human health. The genome of C.
indolis DSM 755(T) reveals an abundance of genes in functional groups associated
with the transport and utilization of carbohydrates, as well as citrate, lactate,
and aromatics. Ecologically relevant gene clusters related to nitrogen fixation
and a unique type of bacterial microcompartment, the CoAT BMC, are also detected.
Our genome analysis suggests hypotheses to be tested in future culture based work
to better understand the physiology of this poorly described species.

<>

<1>Bidle, K.D., Lee, S., Marchant, D.R., Falkowski, P.G.
<2>Fossil genes and microbes in the oldest ice on earth.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>13455-13460
<6>2007
<7>Although the vast majority of ice that formed on the Antarctic continent over the past 34
million years has been lost to the oceans, pockets of
ancient ice persist in the Dry Valleys of the Transantarctic Mountains.
Here we report on the potential metabolic activity of microbes and the
state of community DNA in ice derived from Mullins and upper Beacon
Valleys. The minimum age of the former is 100 ka, whereas that of the
latter is approximately 8 Ma, making it the oldest known ice on Earth. In
both samples, radiolabeled substrates were incorporated into
macromolecules, and microbes grew in nutrient-enriched meltwaters, but
metabolic activity and cell viability were critically compromised with
age. Although a 16S rDNA-based community reconstruction suggested
relatively low bacterial sequence diversity in both ice samples,
metagenomic analyses of community DNA revealed many diverse orthologs to
extant metabolic genes. Analyses of five ice samples, spanning the last 8
million years in this region, demonstrated an exponential decline in the
average community DNA size with a half-life of approximately 1.1 million
years, thereby constraining the geological preservation of microbes in icy
environments and the possible exchange of genetic material to the oceans.

<>

<1>Bieber, J., Schultz, J.
<2>High yield production of restriction endonuclease by culturing bacteria on medium containing yeast extract and pancreatic peptone.
<3>East German Patent Office
<4>DD 232722 A
<5>
<6>1986
<7>Production of restriction endonucleases (I) comprises growing bacteria in a liquid nutrient
medium consisting of yeast extract, pancreatic peptone and NaCl. The cells are then harvested,
lysed and the crude extract purified by usual methods. The bacteria are cultivated under
usual aerobic conditions, esp. on a medium containing, per l, 4-6g yeast extract, 8-12g
peptone and 4-6g NaCl in distilled water, adjusted to pH 7.3 to 7.5.

<>

<1>Biedrzycka, I., Sektas, M., Kaczorowski, T.
<2>Characterization of pEC156, a ColE1-type plasmid from Escherichia coli E1585-68 that carries genes of the EcoVIII restriction-modification system.
<3>Plasmid
<4>45
<5>161
<6>2001
<7>Type II restriction-modification systems are common in prokaryotes.  It is believed that they
evolved to protect these organisms against viral invasion.  Genes encoding R-M systems are
often present on natural plasmids which may facilitate the spread of these systems among
bacteria by horizontal gene transfer.  It has been shown that the genes coding for R-M systems
can increase the stability of the plasmids which carry them.  Bacterial strains isolated from
clinical specimens are especially abundant in plasmids.  Some of them possess Hsd (host
specificity for DNA) plasmids carrying R-M systems.  The strain of E. coli E1585-68 used in
our study was isolated from a patient in England in 1973, and since then it has often been
used as a reference strain for the E. coli O156 serotype.  Later it was discovered that this
strain possesses a natural plasmid (pEC156), carrying genes for the EcoVIII R-M system, a
HindIII isoschizomer.  The complete 4312-bp sequence of the plasmid pEC156 has been
determined.  The plasmid, numbering 20 copies per cell, has been sequenced on both strands and
the sequence has been deposited in GenBank (Accession No. AF158026).  The pEC156 plasmid
carries genes encoding the EcoVIII R-M system - an isoschizomer of HindIII from Haemophilus
influenzae.  Nucleotide sequence analysis of pEC156 suggests that this plasmid possesses a
ColE1 replicon.  It consists of an origin of replication and two untranslated genes encoding
RNAI and RNAII, both involved in the regulation of plasmid DNA replication.  The replication
region also contains a gene, encoding a 64-amino-acid Rom-like (RNA one modulator) peptide,
which presumably is involved in the negative regulation of plasmid pEC156 replication by
stabilizing the NRAI-RNAII complex.  The DNA sequence homologous to the rom region of ColE1
plasmids was also identified.  The stable maintenance of ColE1 plasmids in bacteria depends on
the existence of cis-acting cer locus, which ensures that at cell division each daughter cell
receives at least one copy of the plasmid.  Computational analysis of pEC156 nucleotide
sequence revealed the presence of a DNA region that has high homology to the cer locus of
ColE1 plasmid.  The replication of all ColE1-type plasmids is dependent on the activity of E.
coli DNA polymerase I.  It was shown that pEC156 derivatives carrying antibiotic resistance
genes failed to replicate in an E. coli polA12(ts) mutant at 43oC.  It was also shown that the
pEC156 plasmid copy number increase after treating bacteria with chloramphenicol and decrease
in E. coli pcnB80 mutant.  The pEC156 plasmid carries the genes encoding the EcoVIII R-M
system belonging to type II family.  This system consists of two separate enzymes: an
endonuclease and cognate methyltransferase.  The first of them is the EcoVIII endonuclease
which recognizes specific nucleotide sequence and is capable of cleaving two strands of DNA at
the recognition site, between two adenine residues in the recognition site:
A'A/AGCT-3'/3'TTCGA/A.  the second component is the methyltransferase, which protects
bacterial DNA against cleavage by cognate endonuclease.  Both proteins have been purified and
characterized.

<>

<1>Bielas, J.H., Loeb, L.A.
<2>Quantification of random genomic mutations.
<3>Nat. Methods
<4>2
<5>285-290
<6>2005
<7>Cancer cells contain numerous clonal mutations. It has been theorized that malignant cells
sustain an elevated mutation rate and, as a consequence, harbor yet larger numbers of random
point mutations. Testing this hypothesis has been precluded by lack of an assay to measure
random mutations-that is, mutations that occur in only one or a few cells of a population. We
have established a method that has permitted us to detect and identify rare random mutations
in human cells, at a frequency of 1 per 10(8) base pairs. The assay is based on gene capture,
by hybridization with a uracil-containing probe, followed by magnetic separation. Mutations
that render the mutational target sequence non-cleavable by a restriction enzyme are
quantified by dilution to single molecules and real-time quantitative PCR amplification. The
assay can be extended to quantify mutation in any DNA-based organism, at different sites in
the genome, in introns and exons, in unselected and selected genes, and in proliferating and
quiescent cells.

<>

<1>Bier, F.F., Kleinjung, F., Schmidt, P.M., Scheller, F.W.
<2>Determination of the turnover number of the restriction endonuclease EcoRI using evanescent wave technology.
<3>Anal. Bioanal. Chem.
<4>372
<5>308-313
<6>2002
<7>Binding and catalytic activity of the type II restriction endonuclease EcoRI on immobilized
DNA has been observed in real time using three different evanescent wave biosensors and two
different immobilization techniques.  The method gives direct access to the turnover number
(kcat) without the necessity for the determination of any concentration or activity.  The
combination of different evanescent wave techniques gives access to the catalytic mechanism
and allows the determination of the rate limiting step.

<>

<1>Bigey, P., Knox, J.D., Croteau, S., Bhattacharya, S.K., Theberge, J., Szyf, M.
<2>Modified oligonucleotides as bona fide antagonists of proteins interacting with DNA.
<3>J. Biol. Chem.
<4>274
<5>4594-4606
<6>1999
<7>The study of the biological role of DNA methyltransferase has been impeded by the lack of
direct and specific inhibitors.  This report describes the design of potent DNA based
antagonists of DNA MeTase and their utilization to define the interactions of DNA MeTase with
its substrate and to study its biological role.  We demonstrate that the size, secondary
structure, hemimethylation, and phosphorothioate modification strongly affect the antagonists
interaction with DNA MeTase whereas base substitutions do not have a significant effect.  To
study whether DNA MeTase is critical for cellular transformation, human lung non-small
carcinoma cells were treated with the DNA MeTase antagonists.  Ex vivo, hairpin inhibitors of
DNA MeTase are localized to the cell nucleus in lung cancer cells.  They inhibit DNA MeTase,
cell growth, and anchorage independent growth (an indicator of tumorigenesis in cell culture)
in a dose-dependent manner.  The inhibitors developed in this study are the first documented
example of direct inhibitors of DNA MeTase in living cells and of modified oligonucleotides as
bona fide antagonists of critical cellular proteins.

<>

<1>Bigey, P., Ramchandani, S., Theberge, J., Araujo, F.D., Szyf, M.
<2>Transcriptional regulation of the human DNA methyltransferase (dnmt1) gene.
<3>Gene
<4>242
<5>407-418
<6>2000
<7>DNA methylation is an important component of the epigenetic control of genome functions.
Understanding the regulation of the DNA Methyltransferase (dnmt1) gene expression is critical
for comprehending how DNA methylation is coordinated with other critical biological
processes. In this paper, we investigate the transcriptional regulatory
region of the human dnmt1 gene using a combination of RACE, RNase
protection analysis and CAT assays. We identified one major and three
minor transcription initiation sites in vivo (P1-P4), which are
regulated by independent enhancers and promoter sequences. The minimal
promoter elements of P1, P2 and P4 are mapped within 256 bp upstream of
their respective transcription initiation sites. P1 is nested within a
CC-rich area, similar to other housekeeping genes, whereas P2-P4 are
found in CG-poor areas. Three c-Jun-dependent enhancers are located
downstream to P1 and upstream to P2-P4, thus providing a molecular
explanation for the responsiveness of dnmt1 to oncogenic signals that
are mediated by the Ras-c-Jun oncogenic signaling pathway.

<>

<1>Bigger, C.H., Murray, K., Murray, N.E.
<2>Recognition sequence of a restriction enzyme.
<3>Nature New Biol.
<4>244
<5>7-10
<6>1973
<7>Restriction endonuclease EcoRII makes about twenty double-stranded breaks per
molecule of lambda h80 DNA.  The 5'- terminal sequences are pC-C-A-G-G and
pC-C-T-G-G.  These are complementary and rotationally symmetrical, showing how
the enzyme may produce DNA fragments with short cohesive ends.

<>

<1>Bikandi, J., San, M.R., Rementeria, A., Garaizar, J.
<2>In silico analysis of complete bacterial genomes: PCR, AFLP-PCR and endonuclease restriction.
<3>Bioinformatics
<4>20
<5>798-799
<6>2004
<7>We have developed a website, www.in-silico.com, which runs a software program that performs
three basic tasks in completely sequenced bacterial genomes by in silico analysis: PCR
amplification, amplified fragment length polymorphism (AFLP-PCR) and endonuclease restriction.
For PCR, after selection of the genome and introduction of primers, fragment size, DNA
sequence and corresponding open reading frame (ORF) identity of the resulting PCR product is
computed. Plasmids of sequenced species may be included in the analysis. Theoretical AFLP-PCR
analyzes similar parameters, and includes a suggestion tool providing a list of commercial
restriction enzyme pairs yielding up to 50 amplicons in the selected genome. Endonuclease
restriction analysis of complete genomes and plasmids calculates the number of restriction
sites for endonucleases in a given genome. If the number of fragments is 50 or fewer, pulsed
field gel electrophoresis image and restriction maps are illustrated. Other tools that have
been included in this site are ORF search by name and DNA to protein translation as well as
restriction digestion of user-defined DNA sequences. AVAILABILITY: This is a new molecular
biology resource freely available over the Internet at http://www.in-silico.com

<>

<1>Bilcock, D.T., Daniels, L.E., Bath, A.J., Halford, S.E.
<2>Reactions of type II restriction endonucleases with 8-base pair recognition sites.
<3>J. Biol. Chem.
<4>274
<5>36379-36386
<6>1999
<7>Type II restriction endonucleases usually recognize 4-6-base pair (bp) sites on DNA and cleave
each site in a separate reaction.  A few type II endonucleases have 8-bp recognition sites,
but these seem unsuited for restriction, since their sites are rare on most DNA. Moreover,
only one endonuclease that recognizes a target containing 8 bp has been examined to date, and
this enzyme, SfiI, needs two copies of this site for its DNA cleavage reaction. In this study,
several endonucleases with 8-bp sites were tested on plasmids that have either one or two
copies of the relevant sequence to determine if they also need two sites. SgfI, SrfI, FseI,
PacI, PmeI, Sse8781I, and SdaI all acted through equal and independent reactions at each site.
AscI cleaved the DNA with one site at the same rate as that with two sites but acted
processively on the latter. In contrast, SgrAI showed a marked preference for the plasmid with
two sites and cleaved both sites on this DNA in a concerted manner, like SfiI. Endonucleases
that require two copies of an 8-bp sequence may be widespread in nature, where, despite this
seemingly inappropriate requirement, they may function in DNA restriction.

<>

<1>Bilcock, D.T., Halford, S.E.
<2>DNA restriction dependent on two recognition sites: activities of the SfiI restriction-modification system in Escherichia coli.
<3>Mol. Microbiol.
<4>31
<5>1243-1254
<6>1999
<7>In contrast to many type II restriction enzymes, dimeric proteins that cleave DNA at
individual recognition sites 4-6 bp long, the SfiI endonuclease is a tetrameric protein that
binds to two copies of an elongated sequence before cutting the DNA at both sites.  The mode
of action of the SfiI endonuclease thus seems more appropriate for DNA rearrangements than for
restriction. To elucidate its biological function, strains of Escherichia coli expressing the
SfiI restriction-modification system were transformed with plasmids carrying SfiI sites. The
SfiI system often failed to restrict the survival of a plasmid with one SfiI site, but
plasmids with two or more sites were restricted efficiently.  Plasmids containing methylated
SfiI sites were not restricted.  No rearrangements of the plasmids carrying SfiI sites were
detected among the transformants.  Hence, provided the target DNA contains at least two
recognition sites, SfiI displays all of the hallmarks of a restriction-modification system as
opposed to a recombination system in E. coli cells.,  The properties of the system in vivo
match those of the enzyme in vitro.  For both restriction in vivo and DNA cleavage in vitro,
SfiI operates best with two recognition sites on the same DNA.

<>

<1>Billington, S.J., Huggins, A.S., Johanesen, P.A., Crellin, P.K., Cheung, J.K., Katz, M.E., Wright, C.L., Haring, V., Rood, J.I.
<2>Complete nucleotide sequence of the 27-kilobase virulence related locus (vrl) of Dichelobacter nodosus: evidence for extrachromosomal origin.
<3>Infect. Immun.
<4>67
<5>1277-1286
<6>1999
<7>The vrl locus is preferentially associated with virulent isolates
of the ovine footrot pathogen, Dichelobacter nodosus. The complete
nucleotide sequence of this 27.1-kb region has now been determined. The
data reveal that the locus has a G+C content much higher than the rest
of the D. nodosus chromosome and contains 22 open reading frames (ORFs)
encoding products including a putative adenine-specific methylase, two
potential DEAH ATP-dependent helicases, and two products with sequence
similarity to a bacteriophage resistance system. These ORFs are all in
the same orientation, and most are either overlapping or separated by
only a few nucleotides, suggesting that they comprise an operon and are
translationally coupled. Expression vector studies have led to the
identification of proteins that correspond to many of these ORFs. These
data, in combination with evidence of insertion of vrl into the 3' end
of an ssrA gene, are consistent with the hypothesis that the vrl locus
was derived from the insertion of a bacteriophage or plasmid into the
D. nodosus genome.

<>

<1>Billington, S.J., Jost, B.H.
<2>Multiple genetic elements carry the tetracycline resistance gene, tet(W) in the animal pathogen, Arcanobacterium pyogenes.
<3>Antimicrob. Agents Chemother.
<4>50
<5>3580-3587
<6>2006
<7>The tet(W) gene is associated with tetracycline resistance in a wide range
of bacterial species, including obligately anaerobic rumen bacteria and
isolates from the human gut and oral mucosa. However, little is known
about how this gene is disseminated and the types of genetic elements it
is carried on. We examined tetracycline-resistant isolates of the animal
commensal and opportunistic pathogen Arcanobacterium pyogenes, all of
which carried tet(W), and identified three genetic elements designated
ATE-1, ATE-2, and ATE-3. These elements were found in 25%, 35%, and 60% of
tetracycline-resistant isolates, respectively, with some strains carrying
both ATE-2 and ATE-3. ATE-1 shows characteristics of a mobilizable
transposon, and the tet(W) genes from strains carrying this element can be
transferred at low frequencies between A. pyogenes strains. ATE-2 has
characteristics of a simple transposon, carrying only the resistance gene
and a transposase, while in ATE-3, the tet(W) gene is associated with a
streptomycin resistance gene that is 100% identical at the DNA level with
the aadE gene from the Campylobacter jejuni plasmid pCG8245. Both ATE-2
and ATE-3 show evidence of being carried on larger genetic elements, but
conjugation to other strains was not observed under the conditions tested.
ATE-1 was preferentially associated with A. pyogenes strains of bovine
origin, while ATE-2 and ATE-3 elements were primarily found in porcine
isolates, suggesting that these elements may circulate in different
environments. In addition, four alleles of the tet(W) gene, primarily
associated with different elements, were detected among A. pyogenes
isolates.

<>

<1>Bina, J.E., Nano, F., Hancock, R.E.W.
<2>Utilization of alkaline phosphatase fusions to identify secreted proteins, including potential efflux proteins and virulence factors from Helicobacter pylori.
<3>FEMS Microbiol. Lett.
<4>148
<5>63-68
<6>1997
<7>The targeted genomic strategy of random fusions to a partial gene encoding
a signal sequence-deficient fragment of bacterial alkaline phosphatase was
utilized to screen for secreted proteins in Helicobacter pylori. The
rationale for targeting extracytoplasmic proteins was based on the
hypothesis that most virulence factors and vaccine candidates are secreted
or exported proteins. In addition, extracytosolic proteins represent good
potential targets for drug intervention since they are in general more
accessible to drugs than are cytoplasmically localized proteins. The
application of this strategy to H. pylori allowed the identification of
putative virulence factors and novel targets for drug intervention
including four putative antibiotic efflux genes. The strategy used here is
rapid and technically simple, relatively inexpensive, adaptable to a wide
variety of microbes and genetic systems, and selects for expressed and
accessible proteins.

<>

<1>Binetti, A.G., Suarez, V.B., Tailliez, P., Reinheimer, J.A.
<2>Characterization of spontaneous phage-resistant variants of Streptococcus thermophilus by randomly amplified polymorphic DNA analysis and identification of phage-resistance mechanisms.
<3>Int. Dairy Journal
<4>17
<5>1115-1122
<6>2007
<7>A total of 100 spontaneous phage-resistant mutants isolated from nine commercial Streptococcus
thermophilus strains were characterized
preliminarily by randomly amplified polymorphic DNA (RAPD) and the
nature of their phage-resistance mechanisms was investigated. Only for
mutants isolated from one strain, free phages were detected in their
culture supernatants when these were titrated on the sensitive strain,
suggesting that the mutants could have acquired the resistance
phenotype by integrating the phage in their genomes (lysogeny).
Adsorption interference was observed in the derivatives isolated from
two strains. For mutants isolated from two other strains.
restriction-modification (R-M) type systems were detected. In one of
these cases, R-M was probably combined with another intracellular
anti-phage system. In most cases, the molecular profiles (RAPD
fingerprints) obtained with four arbitrary primers showed a high
similarity among parent strains and their respective phage-resistant
mutants. Some of these mutants were identified as potentially improved
strains for industrial use.

<>

<1>Bingham, A.H.A., Atkinson, T.
<2>Restriction endonucleases and modification methylases in bacteria.
<3>Biochem. Soc. Trans.
<4>6
<5>315-324
<6>1978
<7>There are three mechanisms by which foreign DNA may enter a prokaryotic organism: conjugation
between Gram-negative bacteria, uptake of exogenous DNA (transformation) and infection by
viral (bacteriophage) nucleic acid.  The interaction between a bacteriophage and its host
bacterial cell has been extensively investigated.  A bacterium can restrict phage infection
through its ability to degrade the infectious DNA on entry into the cell.  This is achieved by
site-specific deoxyriboendonucleases, called restriction endonucleases, that cleave DNA into a
limited, but defined, number of fragments, which are then susceptible to exonuclease attack.
The bacterium can prevent cleavage of its chromosomal DNA by site-specific methylation of the
nucleotide bases, for which another activity, the DNA-modification methylase, is responsible.
The modification methylase can also modify bacteriophage DNA on entry into the cell, thus
making the modified phage DNA resistant to the action of the restriction endonuclease.
Although the modification methylase protects the host bacterial chromosome, it can also afford
protection to the invading phage DNA; as a result, phages grown in a given bacterial strain
are insensitive to restriction by that strain. A review.

<>

<1>Bingham, A.H.A., Atkinson, T., Sciaky, D., Roberts, R.J.
<2>A specific endonuclease from Bacillus caldolyticus.
<3>Nucleic Acids Res.
<4>5
<5>3457-3467
<6>1978
<7>The purification and characterization of a new restriction endonuclease, BclI,
from the extreme thermophile Bacillus caldolyticus is reported.  This enzyme
recognizes the sequence 5'-T^-G-A-T-C-A-3' 3'-A-C-T-A-G^-T-5' and cleaves at
the positions indicated by the arrows.

<>

<1>Bingham, A.H.A., Darbyshire, J.
<2>Isolation of two restriction endonucleases from Chloroflexus aurantiacus (CauI, CauII).
<3>Gene
<4>18
<5>87-91
<6>1982
<7>The purification and characterization of two restriction endonucleases from the
photosynthetic gliding bacterium Chloroflexus aurantiacus are described.

<>

<1>Bingham, A.H.A., Sharman, A.F., Atkinson, T.
<2>The purification of restriction endonuclease EcoRI by precipitation involving polyethyleneimine.
<3>FEBS Lett.
<4>76
<5>250-256
<6>1977
<7>The restriction endonuclease EcoRI from an Escherichia coli containing the
plasmid pMB 1,3 or 4 is widely used in the analysis and manipulation of DNA
molecules.  The enzyme has been used in the physical mapping of viral genomes
and mitochondrial DNA, the preliminary analysis of DNA molecules and since
cleavage with EcoRI yields cohesive terminii, in the in vitro construction of
recombinant DNA molecules.  Homogeneous enzyme preparations are not essential
for the in vitro analysis and manipulation of DNA molecules; a preparation
completely free of contaminating non-specific nucleases is adequate for most
experiments.  There are several published procedures for the puruification of
EcoRI, however they all suffer from two major disadvantages, the large number
of steps involved in the preparation of exonuclease-free material and the poor
yields of the enzyme finally obtained.  The use of polyethyleneimine (PEI) for
the removal of nucleic acids from microbial extracts was demonstrated by
Atkinson and Jack, and it was found that if this procedure was carried out
under conditions of low ionic strength with an extract of E. coli RY13 the
EcoRI activity precipitated with the PEI-DNA complex.  This paper describes a
rapid, two step method for the purification of exonuclease-free EcoRI
restriction endonuclease in high yield, based on elution of the enzyme from a
PEI precipitated DNA-enzyme complex.  The preparation of other restriction
endonucleases using a similar technique is also discussed.

<>

<1>Binh, T.T., Suzuki, R., Shiota, S., Kwon, D.H., Yamaoka, Y.
<2>Complete Genome Sequences of Helicobacter pylori Clarithromycin-Resistant Strains.
<3>Genome Announcements
<4>1
<5>e00912-13
<6>2013
<7>We report the complete genome sequences of two Helicobacter pylori clarithromycin-resistant
strains. Clarithromycin (CLR)-resistant strains were
obtained under the exposure of H. pylori strain 26695 on agar plates with low
clarithromycin concentrations. The genome data provide insights into the genomic
changes of H. pylori under selection by clarithromycin in vitro.

<>

<1>Bini, E. et al.
<2>Complete genome sequence of Desulfurispirillum indicum stain S5.
<3>Standards in Genomic Sciences
<4>5
<5>371-378
<6>2011
<7>Desulfurispirillum indicum strain S5T is a strictly anaerobic bacterium isolated from river
se-diment in Chennai, India. D. indicum belongs to the deep branching phylum of
Chrysioge-netes, which currently only includes three other cultured species. Strain S5T is the
type strain of the species and it is capable of growth using selenate, selenite, arsenate,
nitrate or nitrite as terminal electron acceptors. The 2,928,377 bp genome encodes 2,619
proteins and 49 RNA genes, and the information gained from its sequence will be relevant to
the elucidation of mi-crobially-mediated transformations of arsenic and selenium, in addition
to deepening our knowledge of the underrepresented phylum of Chrysiogenetes.

<>

<1>Biniszkiewicz, D., Cesnaviciene, E., Shub, D.A.
<2>Self-splicing group I intron in cyanobacterial initiator methionine tRNA: evidence for lateral transfer of introns in bacteria.
<3>EMBO J.
<4>13
<5>4629-4635
<6>1994
<7>A group I self-splicing intron has been found in the anticodon loop of
tRNA(fMet) genes in three cyanobacterial genera: Dermocarpa, Scytonema and
Synechocystis; it is absent in nine others. The Synechocystis intron is
also interrupted by an open reading frame (ORF) of 150 codons. Of these
three bacteria, only Scytonema also contains the group I intron that has
previously been reported in tRNA(Leu) (UAA) genes in both cyanobacteria
and chloroplasts. The presence of an ORF in the tRNA(fMet) intron, the
sporadic distribution of the intron among cyanobacteria and the lack of
correlation between relatedness of the intron sequences and the bacteria
in which they reside, are all consistent with recent introduction of this
intron by lateral transfer.

<>

<1>Bircakova, M., Truksa, M., Scouten, W.H.
<2>Oriented immobilization of restriction endonuclease EcoRI.
<3>J. Mol. Recognit.
<4>9
<5>683-690
<6>1996
<7>Two activated matrices have been developed to determine whether immobilization chemistry can
be used to orient proteins on a support.  Restriction endonuclease EcoRI from Escherichia coli
RY13 (E.C.3.1.23.13) was used as a model in these studies.  Thiol-activated Sephadex G-10 was
used to couple the EcoRI endonuclease through its free sulfhydryl, while amino-activated
Sephadex G-10 was used to couple it randomly through its free carboxyl groups.  To determine
whether the enzyme was immobilized randomly or specifically, both lower and higher molecular
weight substrates were used.  The polymerase chain reaction amplified multiple cloning site
region of pBluescript KS obtained using T3 and T7 primers was considered as the small
substrate.  The plasmid Sp64 containing firefly luciferase gene was the large substrate.
Immobilized EcoRI preparations were characterized with respect to repeated usage and storage
stability.  The EcoRI immobilized on thiopropyl-Sepharose 4B could be stored for over 14 days
at 4oC without observable loss of activity.  In an independent experiment the same gel was
used thrice repeatedly without any discernible loss of activity.

<>

<1>Bird, A.
<2>The essentials of DNA methylation.
<3>Cell
<4>70
<5>5-8
<6>1992
<7>An obstacle to progress in understanding eukaryotic DNA methylation has been the lack of a
good genetic system - the most favorable eukaryotes for genetic analysis (yeasts,
Caenorhabditis, Drosophila) have no detectable methylation in their genomes. Not only has this
hampered progress, but the clear evidence for life without methylation that these organisms
provide has raised questions about the relevance of the whole phenomenon. The current status
of methylation is reviewed.

<>

<1>Bird, A.
<2>DNA methylation de novo.
<3>Science
<4>286
<5>2287-2288
<6>1999
<7>In an ideal world, biological processes would be understood at the molecular level by first
identifying all the participating components and then by deducing their roles in the system
through experiment.  In reality, knowledge of each component is hard-won, and the temptation
to assume that the key players are those that are currently known, ever-present.  Fortunately,
as human cDNA sequencing approaches saturation, candidate components in mammalian systems are
becoming easier to find.  One beneficiary is the field of DNA methylation in which researchers
study the shutting down of gene expression through the addition of methyl groups to cytosine
bases in the DNA.  Few players are more important in this arena than DNA methyltransferases,
the enzymes responsible for methylating DNA.  Thanks to DNA sequence databases, a clutch of
new Dnmts as well as proteins that bind methylated C bases have recently been uncovered.
Three recent papers make clear that several of these newly recognized Dnmts are capable of de
novo DNA methylation (that is, addition of methyl groups to DNA that has not been methylated
before).  The new work demonstrates the crucial importance of an appropriately methylated
genome for successful embryonic development.

<>

<1>Bird, A., Taggart, M., Frommer, M., Miller, O.J., Macleod, D.
<2>A fraction of the mouse genome that is derived from islands of nonmethylated, CpG-rich DNA.
<3>Cell
<4>40
<5>91-99
<6>1985
<7>About 1% of the mouse genome is cleaved by HpaII to give a discrete fraction on
gels.  The nonmethylated fraction is present in all tested tissues, including
sperm, and contains HpaII sites at about 15 times their frequency in bulk DNA.
About 80% of the fraction is composed of sequences that occur once or a few
times per genome; the remainder is largely rDNA.  Unlike bulk DNA, the fraction
is not deficient in CpG, and this may be directly due to the lack of
methylation.  Genomic mapping of three nonribosomal fragments showed that they
are part of islands of DNA within which nonmethylated HpaII and HhaI sites are
highly concentrated.  We estimate about 30,000 islands per haploid genome and
discuss evidence that many may be associated with genes.

<>

<1>Bird, A.P.
<2>Gene number, noise reduction and biological complexity.
<3>Trends Genet.
<4>11
<5>94-100
<6>1995
<7>Preliminary estimates suggest that gene number, and hence biological complexity, increased
suddenly at two periods of macroevolutionary change (the origin of eukaryotes and the origin
of vertebrates), but otherwise remained relatively constant. As the genome is in constant
flux, what normally constrains the number of different genes that an organism can retain?
Here, I suggest that an important limitation on gene number is the efficiency of mechanisms
that reduce transcriptional background noise. The appearance of both eukaryotes and
vertebrates coincided with novel mechanisms of noise reduction.

<>

<1>Bird, A.P., Taggart, M.H., Smith, B.A.
<2>Methylated and unmethylated DNA compartments in the sea urchin genome.
<3>Cell
<4>17
<5>889-901
<6>1979
<7>Sea urchin (Echinus esculentus) DNA has been separated into high and low molecular weight
fractions by digestion with the mCpG-sensitive restriction endonucleases HpaII, HhaI and AvaI.
The separation was due to differences in methylation at the recognition sequences for these
enzymes because an mCpG-insensitive isoschizomer of HpaII (MspI digested HpaII-resistant DNA
to low molecular weight, showing that many HapII sites were in fact present in this fraction;
and because 3H-methyl methionine administered to embryos was incorporated into the high
molecular weight HpaII-, HhaI- and AvaI-resistant fraction, but not significantly into the low
molecular weight fraction. The fraction resistant to HpaII, HhaI and AvaI amounted to about
401f the total DNA. It consisted of long sequence tracts between 15 and well over 50 kb in
length, in which many sites for each of these enzymes were methylated consecutively. The
remaining 600f the genome, (m-), was not significantly methylated. Methylated and unmethylated
fractions were considered to be subfractions of the genome because enriched unique sequences
from one fraction cross-reassociated poorly with the other fraction and specific sequences
were found in either (m+) or (m-) but not in both (see below). Similar (m+) and (m-)
compartments were found in embryos, germ cells and adult somatic tissues. Furthermore, we
found no evidence for changes in the sequence composition of (m+) or (m-) between sperm,
embryo or intestine DNAs, although low levels of exchange would not have been detected. Using
cloned Echinus histone DNA, heterologous 5S DNA and ribosomal DNA probes, we have found that
each of these gene families belongs to the unmethylated DNA compartment in all the tissues
examined. In particular, there was no detectable methylation of histone DNA either in early
embryos, which are thought to be actively transcribing the bulk of histone genes, or in sperm
and gastrulae, in which most histone genes are not being transcribed. In contrast to these
gene families, sequences complementary to an internally repetitious Echinus DNA clone were
found primarily in the methylated DNA compartment. nt.

<>

<1>Bird, A.P., Wolffe, A.P.
<2>Methylation-induced repression - belts, braces, and chromatin.
<3>Cell
<4>99
<5>451-454
<6>1999
<7>DNA methylation is essential for development in the mouse and plays an important role in
inactivation of the X-chromosome and genomic imprinting.  It may also contribute to
immobilization of mammalian transposons, suppression of transcriptional noise, and the control
of tissue-specific gene expression, but decisive evidence on these points is lacking.  The
theme that is common to all these phenomena is transcriptional repression.  Work on animals,
plants, and fungi now leaves little doubt that gene silencing is a major biological
consequence of DNA methylation.

<>

<1>Birdsell, D.N., Antwerpen, M., Keim, P., Hanczaruk, M., Foster, J.T., Sahl, J.W., Wagner, D.M., Grass, G.
<2>Draft Genome Sequences of Two Bulgarian Bacillus anthracis Strains.
<3>Genome Announcements
<4>1
<5>e00152-13
<6>2013
<7>Bacillus anthracis strains previously isolated from Bulgaria form a unique subcluster within
the A1.a cluster that is typical for isolates from southeastern
Europe. Here, we report the draft genome sequences of two Bulgarian B. anthracis
strains belonging to the A branch (A.Br.)008/009 canonical single nucleotide
polymorphism (SNP) group of the major A branch.

<>

<1>Biro, J.C., Biro, J.M.
<2>Frequent occurrence of recognition site-like sequences in the restriction endonucleases.
<3>BMC Bioinformatics
<4>5
<5>30
<6>2004
<7>BACKGROUND: There are two different theories about the development of the genetic code. Woese
suggested that it was developed in connection with the
amino acid repertoire, while Crick argued that any connection between
codons and amino acids is only the result of an "accident". This question
is fundamental to understand the nature of specific protein-nucleic acid
interactions. RESULTS: The nature of specific protein-nucleic acid
interaction between restriction endonucleases (RE) and their recognition
sequences (RS) was studied by bioinformatics methods. It was found that
the frequency of 5-6 residue long RS-like oligonucleotides is unexpectedly
high in the nucleic acid sequence of the corresponding RE (p < 0.05 and p
< 0.001 respectively, n = 7). There is an extensive conservation of these
RS-like sequences in RE isoschizomers. A review of the seven available
crystallographic studies showed that the amino acids coded by codons that
are subsets of recognition sequences were often closely located to the RS
itself and they were in many cases directly adjacent to the codon-like
triplets in the RS.  Fifty-five examples of this codon-amino acid
co-localization are found and analyzed, which represents 41.5% of total
132 amino acids which are localized within 8 angstroms distance to the C1' atoms
in the DNA. The average distance between the closest atoms in the codons
and amino acids is 5.5 +/- 0.2 A (mean +/- S.E.M, n = 55), while the
distance between the nitrogen and oxygen atoms of the co-localized
molecules is significantly shorter, (3.4 +/- 0.2 A, p < 0.001, n = 15),
when positively charged amino acids are involved. This is indicating that
an interaction between the nucleic- and amino acids might occur.
CONCLUSION: We interpret these results in favor of Woese and suggest that
the genetic code is "rational" and there is a stereospecific relationship
between the codes and the amino acids.

<>

<1>Bischofberger, N., Ng, P.G., Webb, T.R., Matteucci, M.D.
<2>Cleavage of single stranded oligonucleotides by EcoRI restriction endonuclease.
<3>Nucleic Acids Res.
<4>15
<5>709-716
<6>1987
<7>The 31mer 5'-TCA ACG CTA GAA TTC GGA TCC ATC GCT TGG T, the complementary 33mer
5'-CCA AGC GAT GGA TCC GAA TTC TAG CGT TGA GAT, the 40 mer 5'-GGC CAG GAT GGT
GAA GAA TTC GAT CCG GTA CGT AGC TAA G, and the complementary 42mer 5'-TAC TTA
GCT ACG TAC CGG ATC GAA TTC TTC ACC ATC CTG GCC were synthesized and their
reactivity towards EcoRI was studied.  It was found that the 31mer and the
40mer were cleaved at a comparable rate to the 31mer-33mer hybrid and the
40mer-42mer hybrid, respectively.  The rate of cleavage of the 33mer and the
42mer was an order of magnitude lower.  To rule out possible intermolecular
duplex formation, the 33mer was immobilized on cellulose by ligation and
labelled with a 32P-dCTP using Klenow fragment of E. coli DNA polymerase.
EcoRI cleaved this immobilized oligomer into specific fragments.

<>

<1>Bishop, J.O.
<2>A DNA sequence cleaved by restriction endonuclease R.EcoRI in only one strand.
<3>J. Mol. Biol.
<4>128
<5>545-559
<6>1979
<7>Restriction endonuclease EcoRI cuts both strands of the DNA sequence GAATTC
generating two separate frayed ends (Hedgpeth et al., 1972).  Here it is shown
that under standard digestion conditions, the enzyme also attacks the sequence
GAATTA, but cuts only one strand.  The resulting nick is an efficient
initiation point for DNA synthesis by Escherichia coli DNA polymerase I,
allowing the selective labelling of one strand of the DNA duplex.  In buffers
of low molarity and high pH (8.5), EcoRI cleaves sequences with the form NAATTN
(Polisky et al., 1975).  Thus it seems that under both sets of conditions the
enzyme recognizes the four-base-pair core sequence AATT, and that its ability
to cleave different adjacent phosphodiester bonds varies with pH and ionic
strength.

<>

<1>Bishop-Lilly, K.A., Frey, K.G., Daligault, H.E., Davenport, K.W., Bruce, D.C., Chain, P.S., Coyne, S.R., Chertkov, O., Freitas, T., Jaissle, J., Koroleva, G.I., Ladner, J.T., Minogue, T.D., Palacios, G.F., Redden, C.L., Xu, Y., Johnson, S.L.
<2>Complete Genome Sequence of Salmonella enterica subsp. enterica Serovar Enteritidis Strain SEJ.
<3>Genome Announcements
<4>2
<5>e01084-14
<6>2014
<7>Salmonella enterica constitutes a group of enteric pathogens with a broad host range,
including humans, reptiles, and birds. S. enterica subsp. enterica is a
common cause of inflammatory diarrhea in humans. We present the draft genome of
S. enterica subsp. enterica serovar Enteritidis strain SEJ, including a 59-kbp
plasmid.

<>

<1>Bishop-Lilly, K.A., Johnson, S.L., Verratti, K., Luu, T., Khiani, A., Awosika, J., Mokashi, V.P., Chain, P.S., Sozhamannan, S.
<2>Genome sequencing of 15 clinical Vibrio isolates, including 13 non-o1/non-o139 serogroup strains.
<3>Genome Announcements
<4>2
<5>e00893-14
<6>2014
<7>We present draft genome sequences of 15 clinical Vibrio isolates of various serogroups. These
are valuable data for use in studying Vibrio cholerae genetic
diversity, epidemic potential, and strain attribution.

<>

<1>Bist, P., Madhusoodanan, U.K., Rao, D.N.
<2>A mutation in the Mod subunit of EcoP15I restriction enzyme converts the DNA methyltransferase to a site-specific endonuclease.
<3>J. Biol. Chem.
<4>282
<5>3520-3530
<6>2007
<7>A closer inspection of the amino acid sequence of EcoP15I DNA methyltransferase revealed a
region of similarity to the PDXn(D/E)XK
catalytic site of type II restriction endonucleases, except for methionine
in EcoP15I DNA methyltransferase instead of proline. Substitution of
methionine at position 357 by proline converts EcoP15I DNA
methyltransferase to a site-specific endonuclease. EcoP15I-M357P DNA
methyltransferase specifically binds to the recognition sequence
5'-CAGCAG-3' and cleaves DNA asymmetrically EcoP151-M357P.DNA
methyltransferase specifically binds to the recognition sequence
5'-CAGCAG-3' and cleaves DNA asymmetrically, 5'-CAGCAG(N)(10)-3', as
indicated by the arrows, in presence of magnesium ions.

<>

<1>Bist, P., Rao, D.N.
<2>Identification and mutational analysis of Mg2+ binding site in EcoP15I DNA methyltransferase: involvement in target base eversion.
<3>J. Biol. Chem.
<4>278
<5>41837-41848
<6>2003
<7>EcoP15I DNA methyltransferase catalyzes the transfer of the methyl group of
S-adenosyl-l-methionine to the N6 position of the second adenine within
the double-stranded DNA sequence 5'-CAGCAG-3'. To achieve catalysis, the
enzyme requires a magnesium ion. Binding of magnesium to the enzyme
induces significant conformational changes as monitored by circular
dichroism spectroscopy. EcoP15I DNA methyltransferase was rapidly
inactivated by micromolar concentrations of ferrous sulfate in the
presence of ascorbate at pH 8.0. The inactivated enzyme was cleaved into
two fragments with molecular masses of 36 and 35 kDa. Using this affinity
cleavage assay, we have located the magnesium binding-like motif to amino
acids 355-377 of EcoP15I DNA methyltransferase. Sequence homology
comparisons between EcoP15I DNA methyltransferase and other restriction
endonucleases allowed us to identify a PD(X)n(D/E)XK-like sequence as the
putative magnesium ion binding site. Point mutations generated in this
region were analyzed for their role in methyltransferase activity, metal
coordination, and substrate binding. Although the mutant
methyltransferases bind DNA and S-adenosyl-l-methionine as well as the
wild-type enzyme does, they are inactive primarily because of their
inability to flip the target base. Collectively, these data are consistent
with the fact that acidic amino acid residues of the region 355-377 in
EcoP15I DNA methyltransferase are important for the critical positioning
of magnesium ions for catalysis. This is the first example of
metal-dependent function of a DNA methyltransferase. These findings
provide impetus for exploring the role(s) of metal ions in the structure
and function of DNA methyltransferases.

<>

<1>Bist, P., Sistla, S., Krishnamurthy, V., Acharya, A., Chandrakala, B., Rao, D.N.
<2>S-adenosyl-L-methionine is required for DNA cleavage by type III restriction enzymes.
<3>J. Mol. Biol.
<4>310
<5>93-109
<6>2001
<7>The requirement of S-adenosyl-L-methionine (AdoMet) in the cleavage reaction carried out by
type III restriction-modification enzymes has
been investigated. We show that DNA restriction by EcoPI restriction
enzyme does not take place in the absence of exogenously added AdoMet.
Interestingly, the closely related EcoP15I enzyme has endogenously
bound AdoMet and therefore does not require the addition of the
cofactor for DNA cleavage. By employing a variety of AdoMet analogs,
which differ structurally from AdoMet, this study demonstrates that the
carboxyl group and any substitution at the epsilon carbon of methionine
is absolutely essential for DNA cleavage. Such analogs could bring
about the necessary conformational change(s) in the enzyme, which make
the enzyme proficient in DNA cleavage. Our studies, which include
native polyacrylamide gel electrophoresis, molecular size exclusion
chromatography, UV, fluorescence and circular dichroism spectroscopy,
clearly demonstrate that the holoenzyme and apoenzyme forms of EcoP15I
restriction enzyme have different conformations. Furthermore, the Res
and Mod subunits of the EcoP15I restriction enzyme can be separated by
gel filtration chromatography in the presence of 2 M NaCl.
Reconstitution experiments, which involve mixing of the isolated
subunits, result in an apoenzyme form, which is restriction proficient
in the presence of AdoMet. However, mixing the Res subunit with Mod
subunit deficient in AdoMet binding does not result in a functional
restriction enzyme. These observations are consistent with the fact
that AdoMet is required for DNA cleavage. In vivo complementation of
the defective mod allele with a wild-type mod allele showed that an
active restriction enzyme could be formed. Furthermore, we show that
while the purified c2-134 mutant restriction enzyme is unable to cleave
DNA, the c2-440 mutant enzyme is able to cleave DNA albeit poorly.
Taken together, these results suggest that AdoMet binding causes
conformational changes in the restriction enzyme and is necessary to
bring about DNA cleavage.

<>

<1>Biswas, S., Biswas, I.
<2>Complete Genome Sequence of Streptococcus mutans GS-5, a Serotype c Strain.
<3>J. Bacteriol.
<4>194
<5>4787-4788
<6>2012
<7>Streptococcus mutans, a principal causative agent of dental caries, is considered to be the
most cariogenic among all oral streptococci. Of the four S. mutans
serotypes (c, e, f, and k), serotype c strains predominate in the oral cavity.
Here, we present the complete genome sequence of S. mutans GS-5, a serotype c
strain originally isolated from human carious lesions, which is extensively used
as a laboratory strain worldwide.

<>

<1>Bitinaite, J., Grigaite, R., Maneliene, Z., Butkus, V., Janulaitis, A.
<2>Esp3I - a novel type IIs restriction endonuclease from Hafnia alvei that recognizes the sequence 5'-CGTCTC(N)1/5-3' .
<3>Nucleic Acids Res.
<4>19
<5>5076
<6>1991
<7>A new type IIs restriction endonuclease, Esp3I, has been isolated from Hafnia
alvei RFL3*.  Esp3I recognizes the non-palindromic sequence 5'-CGTCTC-3' and
cleaves DNA one nucleotide to the right from the noted recognition strand.

<>

<1>Bitinaite, J., Maneliene, Z., Menkevicius, S., Klimasauskas, S., Butkus, V., Janulaitis, A.
<2>Alw26I, Eco31I and Esp3I - type IIs methyltransferases modifying cytosine and adenine in complementary strands of the target DNA.
<3>Nucleic Acids Res.
<4>20
<5>4981-4985
<6>1992
<7>*The specificity of three DNA methyltransferases M.Alw261, M.Eco31I and M.Esp3I,
isolated from Acinetobacter lwoffi RFL26, Escherichia coli RFL31 and Hafnia
alvei RFL3+, respectively, was determined. All the enzymes methylate both
strands of asymmetric recognition sites yielding m5C in the top-strand and
m6A in the bottom-strand, as below: 

   5'-GTm5CTC      5'-GGTm5CTC     5'-CGTm5CTC 
   3'-Cm6AGAG      3'-CCm6AGAG     3'-GCm6AGAG
   (M.Alw26I)      (M.Eco31I)      (M.Esp3I)
are the first members of type IIs methyltransferases that modify different
types of nucleotides in the recognition sequence.


<>

<1>Bitinaite, J., Mitkaite, G., Dauksaite, V., Jakubauskas, A., Timinskas, A., Vaisvila, R., Lubys, A., Janulaitis, A.
<2>Evolutionary relationship of Alw261, Eco31I and Esp3I, restriction endonucleases that recognise overlapping sequences.
<3>Mol. Genet. Genomics
<4>267
<5>664-672
<6>2002
<7>Type II restriction endonucleases (ENases) have served as models for understanding the
enzyme-based site-specific cleavage of DNA. Using the
knowledge gained from the available crystal structures, a number of
attempts have been made to alter the specificity of ENases by
mutagenesis. The negative results of these experiments argue that the
three-dimensional structure of DNA-ENase complexes does not provide
enough information to enable us to understand the interactions between
DNA and ENases in detail. This conclusion calls for alternative
approaches to the study of structure-function relationships related to
the specificity of ENases. Comparative analysis of ENases that manifest
divergent substrate specificities, but at the same time are
evolutionarily related to each other, may be helpful in this respect.
The success of such studies depends to a great extent on the
availability of related ENases that recognise partially overlapping
nucleotide sequences (e.g. sets of enzymes that bind to recognition
sites of increasing length). In this study we report the cloning and
sequence analysis of genes for three Type I IS restriction-modification
(RM) systems. The genes encoding the ENases Alw26I, Eco31I and Esp3I
(whose recognition sequences are 5'-GTCTC-3', 5'-GGTCTC-3' and
5'-CGTCTC-3'. respectively) and their accompanying methyltransferases
(MTases) have been cloned and the deduced amino acid sequences of their
products have been compared. In pairwise comparisons, the degree of
sequence identity between Alw261, Eco31I and Esp3I ENases is higher
than that observed hitherto among ENases that recognise partially
overlapping nucleotide sequences. The sequences of Alw26I, Eco31I and
Esp3I also reveal identical mosaic patterns of sequence conservation,
which supports the idea that they are evolutionarily related and
suggests that they should show a high level of structural similarity.
Thus these ENases represent very attractive models for the study of the
molecular basis of variation in the specific recognition of DNA
targets. The corresponding MTases are represented by proteins of
unusual structural and functional organisation. Both M.Alw26I and
M.Esp3I are represented by a single bifunctional protein, which is
composed of an m6A-MTase domain fused to aN m5C-MTase domain. In
contrast, two separate genes encode the m6A-MTase and m5C-MTase in
the Eco31I RM system. Among the known bacterial m5C-MTases, the m5C-MTases of M.Alw26I,
M.Eco31I and M.Esp3I represent unique examples of
the circular permutation of their putative target recognition domains
together with the conserved motifs IX and X.

<>

<1>Bitinaite, J., Schildkraut, I.
<2>Self-generated DNA termini relax the specificity of SgrAI restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>1164-1169
<6>2002
<7>The primary target of SgrAI restriction endonuclease is a multiple sequence of the form
5'-Cpu^CCGGPyG. Previous work had indicated that SgrAI must bind two recognition sites
simultaneously for catalysis [Bilcock, D. T., Daniels, L. E., Bath, A. J. & Halford, S. E.
(1999) J. Biol. Chem. 274, 36379-36386]. In the present study, SgrAI is shown to cleave not
only its canonical sequences, but also the sequences 5'-CPuCCGGPy(A,T,C) and 5'-CPuCCGGGG,
both referred to as secondary sequences. On plasmid pSK7, SgrAI cleaves secondary sites
26-fold slower than the canonical site. However, the same plasmid, but without the canonical
site, is cleaved 200-fold slower. We show that DNA termini generated by cleaving the canonical
site for SgrAI assist in the cleavage of secondary sites. The SgrAI-termini in cis with
respect to secondary site are markedly preferred over those in trans. The SgrAI-termini
provided in a form of oligonucleotide duplex are also shown to stimulate canonical site
cleavage. At a 40-fold molar excess of the SgrAI-termini over substrate, the SgrAI specificity
is shown to improve by two orders of magnitude, because of concurrent 10-fold increase in the
cleavage of canonical site and 50-fold decrease in the cleavage of secondary sites. The
unconventional reaction pathway by which SgrAI utilizes the self-generated DNA termini to
cleave its DNA targets has not been observed hitherto among type II restriction endonucleases.
Based on our work and previous reports, a pathway of DNA binding and cleavage by the SgrAI
restriction endonuclease is proposed.

<>

<1>Bitinaite, J., Wah, D.A., Aggarwal, A.K., Schildkraut, I.
<2>FokI dimerization is required for DNA cleavage.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>10570-10575
<6>1998
<7>FokI is a type IIs restriction endonuclease comprised of a DNA recognition domain and a
catalytic domain.  The structural similarity of the FokI catalytic domain to the type II
restriction endonuclease BamHI monomer suggested that the FokI catalytic domains may dimerize.
In addition, the FokI structure, presented in an accompanying paper in this issue of
Proceedings, reveals a dimerization interface between catalytic domains.  We provide evidence
here that FokI catalytic domain must dimerize for DNA cleavage to occur.  First, we show that
the rate of DNA cleavage catalyzed by various concentrations of FokI are not directly
proportional to the protein concentration, suggesting a cooperative effect for DNA cleavage.
Second, we constructed a FokI variant, FokN13Y, which is unable to bind the FokI recognition
sequence but when mixed with wild-type FokI increases the rate of DNA cleavage.  Additionally,
the FokI catalytic domain that lacks the DNA binding domain was shown to increase the rate of
wild-type FokI cleavage of DNA.  We also constructed an FokI variant, FokD483A, R487A, which
should be defective for dimerization because the altered residues reside at the putative
dimerization interface.  Consistent with the FokI dimerization model, the variant FokD483A,
R487A revealed greatly impaired DNA cleavage.  Based on our work and previous reports, we
discuss a pathway of DNA binding, dimerization, and cleavage by FokI endonuclease.

<>

<1>Bitinaite, J.B., Klimasauskas, S.J., Butkus, V.V., Janulaitis, A.
<2>Characterization of restriction-modification enzymes Cfr13I from Citrobacter freundii RFL13.
<3>FEBS Lett.
<4>182
<5>509-513
<6>1985
<7>This communication describes some properties of R.Cfr13I and M.Cfr13I, isolated from
Citrobacter freundii RFL13. R.Cfr13I restriction enzyme recognizes the 5'-G^GNCC sequence and
cleaves, as indicated by the arrow. M.Cfr13I methylase modifies the internal cytosine
producing m5C (5'-GGNm5CC). R.Cfr13I is sensitive not only to this type of substrate
modification but also to hemimethylation in overlapping sites by M.Cfr10I (internal cytosine
of R.Cfr13I recognition is methylated) and M.HpaII (external cytosine is methylated). From
these results the sensitivity of R.Cfr13I to methylation by dcm methylase of E.coli in
overlapping sites is deduced.

<>

<1>Bitner, R.M., Chan-Wha, K., Williams, M.G.
<2>Deproteinization with azlactone-functional supports.
<3>International Patent Office
<4>WO 9400464
<5>
<6>1994
<7>A method of using azlactone-functional supports to separate proteinaceous materials from
non-proteinaceous materials is disclosed. Deproteinization with azlactone-functional supports
can retain biological activity of the non-proteinaceous material and can bind both
net-positive and net-negative proteinaceous materials from biological fluids. A method for
controlling restriction enzyme activities in a biological fluid is also disclosed.

<>

<1>Bitzer, A.S., Garbeva, P., Silby, M.W.
<2>Draft Genome Sequence of Pedobacter sp. Strain V48, Isolated from a Coastal Sand  Dune in the Netherlands.
<3>Genome Announcements
<4>2
<5>e00094-14
<6>2014
<7>Pedobacter sp. strain V48 participates in an interaction with Pseudomonas fluorescens which
elicits interaction-induced phenotypes. We report the draft
genome sequence of Pedobacter sp. V48, consisting of 6.46 Mbp. The sequence will
contribute to improved understanding of the genus and facilitate genomic analysis
of the model interspecies interaction with P. fluorescens.

<>

<1>Bjerga, G.E., Hjerde, E., De Santi, C., Williamson, A.K., Smalas, A.O., Willassen, N.P., Altermark, B.
<2>High quality draft genome sequence of Streptomyces sp. strain AW19M42 isolated from a sea squirt in Northern Norway.
<3>Standards in Genomic Sciences
<4>9
<5>676-686
<6>2014
<7>Here we report the 8 Mb high quality draft genome of Streptomyces sp. strain AW19M42, together
with specific properties of the organism and the generation,
annotation and analysis of its genome sequence. The genome encodes 7,727 putative
open reading frames, of which 6,400 could be assigned with COG categories. Also,
62 tRNA genes and 8 rRNA operons were identified. The genome harbors several gene
clusters involved in the production of secondary metabolites. Functional
screening of the isolate was positive for several enzymatic activities, and some
candidate genes coding for those activities are listed in this report. We find
that this isolate shows biotechnological potential and is an interesting target
for bioprospecting.

<>

<1>Bjorkholm, B.M., Guruge, J.L., Oh, J.D., Syder, A.J., Salama, N., Guillemin, K., Falkow, S., Nilsson, C., Falk, P.G., Engstrand, L., Gordon, J.I.
<2>Colonization of germ-free transgenic mice with genotyped Helicobacter pylori strains from a case-control study of gastric cancer reveals a  correlation between host responses and HsdS components of type I  restriction-modification systems.
<3>J. Biol. Chem.
<4>277
<5>34191-34197
<6>2002
<7>Helicobacter pylori infects the stomachs of half of all humans. It has a relatively benign
relationship with most hosts but produces severe
pathology, including gastric cancer, in others. Identifying the relative
contributions of host, microbial, and environmental factors to the outcome
of infection has been challenging. Here we describe one approach for
identifying microbial genes that affect the magnitude of host responses to
infection. Single colony purified H. pylori isolates were obtained from 25
cases and 71 controls in a Swedish case-control study of gastric cancer.
Strains were first phenotyped based on their ability to produce adhesins
that recognize two classes of human gastric epithelial receptors. Thirteen
binding strains and two non-binding controls were then subjected to whole
genome genotyping using H. pylori DNA microarrays. A cohort of "variable"
genes was identified based on a microarray-determined call of "absent" in
at least one member of the strain panel. Each strain was subsequently
introduced into two types of germ-free transgenic mice, each programmed to
express a different host factor postulated to pose increased risk for
development of severe pathology. Expression of biomarkers of host defense
was quantitated 4 weeks after inoculation, and the magnitude of the
response correlated with bacterial genotype. The proportion of genes
encoding HsdS homologs (specificity subunit of heterooligomeric type I
restriction-modification systems) was significantly higher in the pool of
18 variable genes whose presence directly correlated with a robust host
response than their proportion in the remaining 352 members of the
variable gene pool. This suggests that the functions of these HsdS
homologs may include control of expression of microbial determinants that
affect the extent of gastric responses to this potentially virulent
pathogen.

<>

<1>Black, M.T., Hodgson, J.E., Knowles, D.J.C., Nicholas, R.O., Stodola, R.K.
<2>Compounds.
<3>US Patent Office
<4>US 6348328 B
<5>
<6>2002
<7>This invention relates to newly identified polynucleotides, polypeptides encoded by such
polynucleotides, the uses of such polynucleotides and polypeptides, as well as the production
of such polynucleotides and polypeptides and recombinant host cells transformed with the
polynucleotides.  This invention also relates to inhibiting the biosynthesis or action of such
polynucleotides or polypeptides and to the use of such inhibitors in therapy.

<>

<1>Black, M.T., Hodgson, J.E., Knowles, D.J.C., Nicholas, R.O., Stodola, R.K.
<2>Novel compound.
<3>Japanese Patent Office
<4>JP 2000508178 A
<5>
<6>2000
<7>
<>

<1>Black, M.T., Hodgson, J.E., Nouruzu, D.J.C., Lonetto, M.A., Nicholas, R.O., Stodola, R.K.
<2>Novel prokaryotic polynucleotides, polypeptides and their uses.
<3>Japanese Patent Office
<4>JP 2000514308 A
<5>
<6>2000
<7>
<>

<1>Blakely, G.W., Murray, N.E.
<2>Control of the endonuclease activity of type I restriction-modification systems is required to maintain chromosome integrity following  homologous recombination.
<3>Mol. Microbiol.
<4>60
<5>883-893
<6>2006
<7>A type I restriction-modification enzyme will bind to an unmethylated target sequence in DNA
and, while still bound to the target,
translocate DNA through the protein complex in both directions. DNA
breakage occurs when two translocating complexes collide. However, if
type I restriction-modification systems bind to unmodified target
sequences within the resident bacterial chromosome, as opposed to
incoming 'foreign' DNA, their activity is curtailed; a process known as
restriction alleviation (RA). We have identified two genes in
Escherichia coli, rnhA and recG, mutations in which lead to the
alleviation of restriction. Induction of RA in response to these
mutations is consistent with the production of unmodified target
sequences following DNA synthesis associated with both homologous
recombination and R-loop formation. This implies that a normal function
of RA is to protect the bacterial chromosome when recombination
generates unmodified products. For EcoKI, our experiments demonstrate
the contribution of two pathways that serve to protect unmodified DNA
in the bacterial chromosome: the primary pathway in which ClpXP
degrades the restriction endonucleas and a mechanism dependent on the
lar gene within Rac, a resident, defective prophage of E. coli K-12.
Previously, the potential of the second pathway has only been
demonstrated when expression of lar has been elevated. Our data
identify the effect of lar from the repressed prophage.

<>

<1>Blakesley, R.W.
<2>Restriction endonuclease:  cleavage, ligation, and sensitivity.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>51-102
<6>1987
<7>Restriction endonucleases are important tools for the molecular biologist.
These enzymes are routinely used to subdivide DNA molecules in a very specific,
predictable fashion, allowing one to isolate certain regions for study.  DNA
sequencing, cloning, mapping, hybridization, and genome characterization are
some of the more common procedures incorporating restriction endonuclease
treated DNA.  The characteristics of the restriction endonucleases and their
reactions have been reviewed.  A comprehensive listing of the enzymes and the
diversity of microbiological sources appears in this volume.  Informative
tables on some characteristics of restriction endonucleases have appeared
elsewhere.  Tabulated in this chapter are a number of facts frequently used
when planning experiments that utilize restriction endonucleases.  The final
table summarizes a number of general characteristics of the enzyme which should
be valuable when designing complex strategies or trouble-shooting unexpected
results.

<>

<1>Blakesley, R.W.
<2>Restriction endonuclease: specificities, diversities and computer analysis.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>1-43
<6>1981
<7>I. Introduction II. Table I: DNA cleavage frequency for restriction
endonucleases with known recognition sequences III. Table II: DNA cleavage
frequency for restriction endonucleases with undetermined recognition sequences
IV. Table III: Microorganisms as sources for restriction endonucleases V. Table
IV: Reaction conditions for certain restriction endonucleases VI. Table V:
Restriction endonuclease cross reference: Tables I-IV, references VII. Table
VI: Restriction endonuclease site combinations for ligation.

<>

<1>Blakesley, R.W., Dodgson, J.B., Nes, I.F., Wells, R.D.
<2>Duplex regions in "single-stranded" PhiX174 DNA are cleaved by a restriction endonuclease from Haemophilus aegyptius.
<3>J. Biol. Chem.
<4>252
<5>7300-7306
<6>1977
<7>DNA from bacteriophage PhiX174 is cleaved by several DNA restriction
endonucleases.  We wish to determine whether or not the recognition sites are
within duplex regions in this single-stranded DNA.  We report here the
influence of two perturbants of duplex DNA structure, temperature and
actinomycin, on the cleavage of PhiX174 (single-stranded, circular) DNA and its
double-stranded, replicative form (RF DNA) by the restriction endonuclease,
HaeIII.  The conditions for optimal rates of cleavage for the (+)-strand and RF
DNA's by HaeIII were virtually identical (25 mM Tris/HCl, pH 7.5; 5 mM MgCl2;
30 mM NaCl).  At 37C, RF DNA was cleaved 16 times faster than (+)-strand DNA.
When the initial rates of reaction for both DNA's were measured as a function
of temperature of incubation, the maximum rates occurred at 72C and 47C for the
RF and (+)-strand DNA's, respectively.  At 10-15C above the respective maxima,
the rates fell to zero.  Near the temperature optima, a preferential
disappearance of certain fragments with temperature occurred with (+)-strand,
but not with RF DNA.  These results indicate that the HaeIII restriction sites
in PhiX174 (+)-strand DNA are located in several different, noncontiguous,
duplex regions with different sequence-dependent tM values.  The rate of
cleavage of each DNA was also inhibited by actinomycin; 50% inhibition occurred
at a molar ratio (actinomycin/nucleotide) of 1.2 for RF and 0.04 for (+)-strand
DNA.  Netropson, which, like actinomycin binds only to duplex DNA, inhibited
the cleavage of both DNA's.  The restriction endonuclease HhaI, gave results
similar to those found for HaeIII.  RF DNA was cleaved once by the restriction
endonuclease, PstI; (+)-strand DNA was not cleaved.  This result for (+)-strand
would be expected if intramolecular base-pairing were required for cleavage.
RF and (+)-strand DNA's were also cleaved by MboI and HinfI.  Neither DNA was
cleaved by BamHI, SmaI, or SalI.  These results indicate that certain DNA
restriction endonucleases cleave "single-stranded" viral DNA's at duplex
regions.

<>

<1>Blakesley, R.W., Wells, R.D.
<2>Single-stranded DNA from PhiX174 and M13 is cleaved by certain restriction endonucleases.
<3>Nature
<4>257
<5>421-422
<6>1975
<7>During a systematic survey of the substrate specificities of a variety of
restriction endonucleases, it was discovered that the Haemophilus aegyptius
restriction endonuclease III (HaeIII) specifically cleaved 'single-stranded'
DNA from PhiX174 and M13.  Since these DNAs are not doubled-stranded and
restriction enzymes recognize duplex DNA with twofold sequence symmetry, this
result was unexpected.  This finding is significant with regard to the
conformation of single-stranded viral DNA as well as the biochemical mechanism
and physiological role of restriction enzymes.

<>

<1>Blakeway, L.V., Power, P.M., Jen, F.E., Worboys, S.R., Boitano, M., Clark, T.A., Korlach, J., Bakaletz, L.O., Jennings, M.P., Peak, I.R., Seib, K.L.
<2>ModM DNA methyltransferase methylome analysis reveals a potential role for Moraxella catarrhalis phasevarions in otitis media.
<3>FASEB J.
<4>28
<5>5197-5207
<6>2014
<7>Moraxella catarrhalis is a significant cause of otitis media and exacerbations of
chronic obstructive pulmonary disease. Here, we characterize a phase-variable DNA
methyltransferase (ModM), which contains 5'-CAAC-3' repeats in its open reading
frame that mediate high-frequency mutation resulting in reversible on/off
switching of ModM expression. Three modM alleles have been identified (modM1-3),
with modM2 being the most commonly found allele. Using single-molecule, real-time
(SMRT) genome sequencing and methylome analysis, we have determined that the
ModM2 methylation target is 5'-GARm6AC-3', and 100% of these sites are methylated
in the genome of the M. catarrhalis 25239 ModM2 on strain. Proteomic analysis of
ModM2 on and off variants revealed that ModM2 regulates expression of multiple
genes that have potential roles in colonization, infection, and protection
against host defenses. Investigation of the distribution of modM alleles in a
panel of M. catarrhalis strains, isolated from the nasopharynx of healthy
children or middle ear effusions from patients with otitis media, revealed a
statistically significant association of modM3 with otitis media isolates. The
modulation of gene expression via the ModM phase-variable regulon (phasevarion),
and the significant association of the modM3 allele with otitis media, suggests a
key role for ModM phasevarions in the pathogenesis of this organism.-Blakeway, L.
V., Power, P. M., Jen, F. E.-C., Worboys, S. R., Boitano, M., Clark, T. A.,
Korlach, J., Bakaletz, L. O., Jennings, M. P., Peak, I. R., Seib, K. L. ModM DNA
methyltransferase methylome analysis reveals a potential role for Moraxella
catarrhalis phasevarions in otitis media.

<>

<1>Blanc, V., Gil, P., Bamas-Jacques, N., Lorenzon, S., Zagorec, M., Schleuniger, J., Bisch, D., Blanche, F., Debussche, L., Crouzet, J., Thibaut, D.
<2>Identification and analysis of genes from Streptomyces pristinaespiralis encoding enzymes involved in the biosynthesis of the 4-dimethylamino-L-phenylalanine precursor of pristinamycin I.
<3>Mol. Microbiol.
<4>23
<5>191-202
<6>1997
<7>Four pap genes (papA, papB, papC, papM) were found by sequencing near to snbA, a Streptomyces
pristinaespiralis gene which was previously shown to encode one of the pristinamycin I (PI)
synthetases. Analysis of the homologies observed from the deduced amino acid sequences
suggested that these four genes could be involved in the biosynthesis of the PI precursor
4-dimethylamino-L-phenylalanine (DMPAPA). This was first verified when disruption of papA in
S. pristinaespiralis led to a PI- phenotype, which was reversed by the addition of DMPAPA into
the culture medium. Further confirmation was obtained when papM was overexpressed in
Escherichia coli and the corresponding protein purified to homogeneity. It catalysed the two
successive N-methylation steps of 4-amino-L-phenylalanine leading to DMPAPA via
4-methylamino-L-phenylalanine. These results allowed us to assign a function to each of the
four pap genes and to propose a biosynthetic pathway for DMPAPA.

<>

<1>Blanck, A., Gluck, B., Wartbichler, R., Bender, S., Poll, M., Brandl, A.
<2>Activity of restriction enzymes in a PCR mix.
<3>Biochemica, Boehringer Mannheim GmbH, , 
<4>2
<5>14
<6>1995
<7>By cutting PCR products with restriction enzymes, the resulting DNA fragments can be cloned
directionally into vector DNA.  Alternatively, researchers can use this cleavage procedure to
verify that the DNA of interest has been successfully amplified or even to show a Restriction
Fragment Length Polymorphism (RFLP) in the amplified fragments.  For maximum convenience, the
restriction enzyme digest would be performed directly in the PCR mix, without prior
purification of the amplified fragment.  To investigate which restriction enzymes show
sufficient activity to be used directly in the PCR mix, we performed an activity test with 50
selected restriction enzymes.

<>

<1>Blanga-Kanfi, S., Amitsur, M., Azem, A., Kaufmann, G.
<2>PrrC-anticodon nuclease: functional organization of a prototypical bacterial restriction RNase.
<3>Nucleic Acids Res.
<4>34
<5>3209-3219
<6>2006
<7>The tRNA(Lys) anticodon nuclease PrrC is associated in latent form with the type Ic DNA
restriction endonuclease EcoprrI and activated by a phage
T4-encoded inhibitor of EcoprrI. The activation also requires the
hydrolysis of GTP and presence of dTTP and is inhibited by ATP. The
N-proximal NTPase domain of PrrC has been implicated in relaying the
activating signal to a C-proximal anticodon nuclease site by interacting
with the requisite nucleotide cofactors [Amitsur et al. (2003) Mol.
Microbiol., 50, 129-143]. Means described here to bypass PrrC's
self-limiting translation and thermal instability allowed purifying an
active mutant form of the protein, demonstrating its oligomeric structure
and confirming its anticipated interactions with the nucleotide cofactors
of the activation reaction. Mutagenesis and chemical rescue data shown
implicate the C-proximal Arg320, Glu324 and, possibly, His356 in anticodon
nuclease catalysis. This triad exists in all the known PrrC homologs but
only some of them feature residues needed for tRNA(Lys) recognition by the
Escherichia coli prototype. The differential conservation and consistent
genetic linkage of the PrrC proteins with EcoprrI homologs portray them as
a family of restriction RNases of diverse substrate specificities that are
mobilized when an associated DNA restriction nuclease is compromised.

<>

<1>Blanks, R., McLaughlin, L.W.
<2>An oligodeoxynucleotide affinity column for the isolation of sequence specific DNA binding proteins.
<3>Nucleic Acids Res.
<4>16
<5>10283-10299
<6>1988
<7>A nucleic acid affinity matrix containing a short oligodeoxynucleotide ligand
has been prepared as an example of a material which can be used for the rapid
and effective isolation of sequence specific DNA binding proteins.  Two
complementary oligodeoxynucleotides have been employed, one of which contains a
small 5'-spacer arm with a terminal thiol group.  Using this terminal thiol
group, the ligand can be covalently coupled to Tresyl-activated Sepharose 4B or
Epoxy-activated Sepharose 6B via a thioether linkage.  This approach allows the
specific attachment of the nucleic acid ligand via its 5'-terminus to the
insoluble matrix.  The double stranded affinity material was obtained by
annealing of the complementary DNA fragment.  As an example, we have used an
eicosomer affinity column containing the sequence d(GAATTC) for the isolation
of the EcoRI restriction endonuclease.  Using a single column, the enzyme could
be isolated by eluting the column with a single step or multistep gradient of
increasing salt concentration.  The enzyme was purified to 75%-85% homogeneity
with yields of 0.1 mg to 0.2 mg from 0.5 g of cell paste.

<>

<1>Blasiak, J., Kowalik, J.
<2>Interaction between organophosphorus compounds and DNA assayed by the restriction endonuclease EcoRI.
<3>Acta Univ. Lodz. Folia Biochim. Biophys.
<4>13
<5>61-67
<6>1998
<7>Restriction endonucleases due to the nature of their action may provide information on the
location or the sequence specificity of a compound that binds to DNA.  On the other hand, the
action of the enzymes may be disturbed by compounds that have an ability to methylate DNA
bases.  The latter feature can be considered as a simple method for primary selection of
potentially genotoxic compounds.  In the present work we investigated the action of
restriction endonuclease EcoRI on DNA which had been incubated with some organophosphorus
agents.  PUC19 plasmid DNA at a concentration of 78 microgram/ml was incubated for 72 h with
organophosphorus insecticides parathion, methylparathion and their main metabolites: paraoxon
and methylparaoxon, respectively, at a concentration of 300 microM.  After incubation
non-bound insecticides were removed and DNA was subjected to 1 h incubation with the
restriction endonuclease EcoRI and electrophoresed in 0.8% agarose gel.  The organophosphorus
compound methylparaoxon evoked unwinding of supercoiled DNA and the action of EcoRI on the DNA
was disturbed that was displayed in changes in restriction pattern.

<>

<1>Blattler, M.O., Wenz, C., Pingoud, A., Benner, S.A.
<2>Distorting duplex DNA by dimethylenesulfone substitution: A new class of "transition state analog" inhibitors for restriction enzymes.
<3>J. Am. Chem. Soc.
<4>120
<5>2674-2675
<6>1998
<7>After they bind (but before they cleave) duplex DNA, some restriction enzymes (such as EcoRV
and EcoRI) distort the duplex.  The distorted duplex is not, of course, in its ground-state
conformation; it requires "binding energy" to bend DNA.  Thus, an analogue of DNA that
generates this distortion in the unbound state (without altering other features of the
substrate that are recognized by the enzyme) should bind to these restriction enzymes with a
higher affinity than the DNA substrate itself.  This is, of course, the principle underlying
transition-state analogues generally, which approximate in structure the "distorted"
transition state (or a distorted high-energy intermediate) for an enzymatic reaction.

<>

<1>Blattner, F.R., Burland, V., Perna, N.T., Plunkett, G., Welch, R.
<2>Sequences of E. coli O157.
<3>US Patent Office
<4>US 6365723 A
<5>
<6>2002
<7>The entire genome of pathogenic E. coli strain O157:H7 has been sequenced.  All of the genomic
DNA sequences present in O157 and absent in the previously sequenced laboratory strain K12 are
presented here.

<>

<1>Blattner, F.R., Burland, V., Perna, N.T., Plunkett, G., Welch, R.
<2>Sequences of E. coli O157.
<3>US Patent Office
<4>US 6855814 A
<5>
<6>2005
<7>The entire genome of pathogenic E. coli strain 0157:H7 has been sequenced.  All of the genomic
DNA sequences present in 0157 and absent in the previously sequenced laboratory strain K12 are
presented here.

<>

<1>Blattner, F.R., Burland, V., Rose, D.J., Mayhew, G.F., Perna, N., Perry, R.D., Straley, S.C., Fetherston, J.D., Lindler, L.E., Plano, G.V.
<2>Plasmid DNA from Yersinia pestis.
<3>US Patent Office
<4>US 6706522 B
<5>
<6>2004
<7>The complete DNA sequence of three plasmids from the bacterium Yersinia pestis, the causative
agent for bubonic plague, have been determined and are set forth.  The open reading frames, or
protein coding regions, of the plasmids have been determined.  The DNA sequence and ORF
information is useful for the creation of diagnostic, prophylactic and therapeutic tools for
combating the disease caused by this agent.

<>

<1>Blattner, F.R., Plunkett, G. III, Bloch, C.A., Perna, N.T., Burland, V., Riley, M., Collado-Vides, J., Glasner, J.D., Rode, C.K., Mayhew, G.F., Gregor, J., Davis, N.W., Kirkpatrick, H.A., Goeden, M.A., Rose, D.J., Mau, B., Shao, Y.
<2>The complete genome sequence of Escherichia coli K-12.
<3>Science
<4>277
<5>1453-1462
<6>1997
<7>The 4,639,221-base pair sequence of Escherichia coli K-12 is presented. Of 4288 protein-coding
genes annotated, 38 percent have no attributed function.  Comparison with five other sequenced
microbes reveals ubiquitous as well as narrowly distributed gene families; many families of
similar genes within E. coli are also evident.  The largest family of paralogous proteins
contains 80 ABC transporters.  The genome as a whole is strikingly organized with respect to
the local direction of replication; guanines, oligonucleotides possibly related to replication
and recombination, and most genes are so oriented.  The genome also contains insertion
sequence elements, phage remnants, and many other patches of unusual composition indicating
genome plasticity through horizontal transfer.

<>

<1>Blattner, F.R., Welch, R.A., Burland, V.D.
<2>DNA sequences of Escherichia coli CFT073.
<3>International Patent Office
<4>WO 02059320 A
<5>
<6>2002
<7>The entire genome of pathogenic E. coli strain CFT073 has been sequenced.  Nearly all of the
genomic DNA sequences present in CFT073 and absent in the previously sequenced laboratory
strain K-12 are presented here.

<>

<1>Bleckwedel, J., Teran, L.C., Bonacina, J., Saavedra, L., Mozzi, F., Raya, R.R.
<2>Draft Genome Sequence of the Mannitol-Producing Strain Lactobacillus mucosae CRL573.
<3>Genome Announcements
<4>2
<5>e01292-14
<6>2014
<7>Lactobacillus mucosae CRL573, isolated from child fecal samples, efficiently converts fructose
and/or sucrose into the low-calorie sugar mannitol when
cultured in modified MRS medium at pH 5.0. Also, the strain is capable of
producing bacteriocin. The draft genome sequence of this strain with potential
industrial applications is presented here.

<>

<1>Blom, J., Rueckert, C., Niu, B., Wang, Q., Borriss, R.
<2>The Complete Genome of Bacillus amyloliquefaciens subsp. plantarum CAU B946 Contains a Gene Cluster for Nonribosomal Synthesis of Iturin A.
<3>J. Bacteriol.
<4>194
<5>1845-1846
<6>2012
<7>The genome of the rhizobacterium Bacillus amyloliquefaciens subsp. plantarum CAU  B946 was
4.02 Mb in size and harbored 3,823 genes (coding sequences [CDS]). Nine
giant gene clusters were dedicated to nonribosomal synthesis of antimicrobial
compounds. Remarkably, strain CAU B946 possessed a gene cluster involved in
synthesis of iturin A.

<>

<1>Blouin, Y., Platonov, M.E., Pourcel, C., Evseeva, V.V., Afanas'ev, M.V., Balakhonov, S.V., Anisimov, A.P., Vergnaud, G.
<2>Draft Genome Sequences of Two Yersinia pseudotuberculosis ST43 (O:1b) Strains, B-7194 and B-7195.
<3>Genome Announcements
<4>1
<5>e00510-13
<6>2013
<7>We report the first draft genome sequences of two Yersinia pseudotuberculosis sequence type 43
(ST43) (O:1b) strains, B-7194 and B-7195, isolated in Russia.
The total lengths of the assemblies are 4,427,121 bp and 4,608,472 bp, and 3,819
and 4,018 coding sequences, respectively, were predicted within the genomes.

<>

<1>Blum, E., Horst, G., Kroger, M.
<2>PCR directed preparation and single step purification of highly active histidine-tagged restriction endonuclease HgiBI (GGWCC).
<3>J. Biochem. Biophys. Methods
<4>29
<5>113-121
<6>1994
<7>The polymerase-chain-reaction technique is used to produce fusion proteins via deletion of any
intervening piece of DNA. Here a stretch of six histidine codons is fused to the 3'-terminus
of any defined gene using a standard plasmid vector or a derivative thereof. The advantage
over existing methods is that no other amino acids besides the six histidines are added to the
protein terminus and only one oligonucleotide needs to be synthesized as special primer. Genes
of interest must only be cloned in the correct orientation into a universal multilinker. Using
just one specific primer derived from the 3'-terminus of the gene and one standard primer
derived from the six histidine codons the fusion is performed by amplifying the entire vector
system as described for inverse PCR. As an example, we report on the modification and
purification of the restriction endonuclease HgiBI (GGWCC). Enzymatically active protein was
obtained in a single step purification under nondenaturing conditions with a purity greater
than 95% according to polyacrylamide gel electrophoresis.

<>

<1>Blum, E., Horst, G., Kroger, M.
<2>PCR-directed preparation and single-step purification of highly active histidine-tagged restriction endonuclease HgiBI (GGWCC).
<3>Gene
<4>157
<5>107-108
<6>1995
<7>The polymerase chain reaction was used to produce His6 fusion proteins via deletion of an
intervening piece of DNA.  The generally applicable method was performed using a standard
primer with the advantage that the fusion does not produce additional amino acids.  In a
single-step purification highly purified, enzymatically active restriction endonuclease was
obtained.

<>

<1>Blum, S., Sela, N., Heller, E.D., Sela, S., Leitner, G.
<2>Genome Analysis of Bovine-Mastitis-Associated Escherichia coli O32:H37 Strain P4.
<3>J. Bacteriol.
<4>194
<5>3732
<6>2012
<7>Escherichia coli is a major pathogen of bovine intramammary infections. Here we report the
first draft of the genome sequence of the E. coli O32:H37 P4 strain,
which is widely used in experimental bovine mastitis studies.

<>

<1>Blumenthal, R.M.
<2>The PvuII restriction-modification system:  cloning, characterization and use in revealing in E. coli barrier to certain methylases or methylated DNAs.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>227-245
<6>1987
<7>The physiology of the type II restriction-modification systems (RMS2s) is not
yet understood in detail.  Relatively few RMS2s have actually been demonstrated
to carry out the role for which they were named:  restriction of DNA entry into
the cell through selective endonuclease action.  On the other hand, there are
numerous examples of DNA methylases, biochemically identical to RMS2 enzymes
but apparently lacking a cognate endonuclease activity, for which no role has
yet been discovered.  Our limited knowledge in this area has hindered attempts
to understand why the genes for certain RMS2s have been difficult to clone and
express.  Several basic questions remain to be answered regarding the
regulation of and sequence recognition by RMS2 enzymes.  Also, does the
methylase of a newly introduced RMS2 have to fully protect the DNA of its new
host before the endonuclease gene is expressed, or can the cell repair a
significant amount of endonucleolytic damage?  And what are the effects of a
foreign DNA methylase on the cell?  We have cloned and characterized the genes
for the PvuII RMS2 from Proteus vulgaris and, in the course of studies designed
to examine their regulation, found that the majority of E. coli strains tested
are poorly transformed by these genes.  We found that this effect is due to the
PvuII methylase, not to the endonuclease, and that DNAs methyolated by the
PvuII methylase also transform most tested strains poorly.  Others have found
this effect with several other DNA methylases, and it has raised new questions
about gene cloning procedures and about E. coli physiology.

<>

<1>Blumenthal, R.M.
<2>E. coli can restrict methylated DNA and may skew genomic libraries.
<3>Trends Biotechnol.
<4>4
<5>302-305
<6>1986
<7>None

<>

<1>Blumenthal, R.M.
<2>Cloning and restriction of methylated DNA in Escherichia coli.
<3>BRL Focus
<4>11
<5>41-46
<6>1989
<7>A review of mcr systems.

<>

<1>Blumenthal, R.M., Cheng, X.
<2>A Taq attack displaces bases.
<3>Nat. Struct. Biol.
<4>8
<5>101-103
<6>2001
<7>The cocrystal structure of TaqI DNA methyltransferase in complex with its specific DNA
substrate adds to the growing list of structurally confirmed DNA base-flipping enzymes, and
provides a basis for the general mechanism of AdoMet-dependent DNA N-methylation.

<>

<1>Blumenthal, R.M., Cotterman, M.M.
<2>Isolation of mutants in a DNA methyltransferase through mcr-B-mediated restriction.
<3>Gene
<4>74
<5>271-273
<6>1988
<7>A procedure has been developed that permits the positive selection of mutants
in a DNA methyltransferase (MTase) gene.  The stringency of this selection can
be varied so as to yield null mutants only, or a mixture of null and partially
defective mutants.  The procedure was developed with the PvuII MTase gene
(pvuIIM), which was subcloned into a bacteriophage Lambda vector.  Growth of
this lambda pvuIIM construct on an mcrB+ host selected for non-methylating
mutants, and the stringency of selection was proportional to the number of
consecutive lytic cycles.  Many cytosine MTases have been found to generate
substrates for mcrB-mediated restriction, and this procedure should be
applicable to a number of cytosine MTase genes.

<>

<1>Blumenthal, R.M., Gregory, S.A., Cooperider, J.S.
<2>Cloning of a restriction-modification system from Proteus vulgaris and its use in analyzing a methylase-sensitive phenotype in Escherichia coli.
<3>J. Bacteriol.
<4>164
<5>501-509
<6>1985
<7>A 4.84 kilobasepair plasmid was isolated from Proteus vulgaris (ATCC 13315) and
cloned into the plasmid vector pBR322.  Plasmid pBR322 contains substrate sites
for the restriction endonucleases PvuI and PvuII.  The recombinant plasmids
were resistant to in vitro cleavage by PvuII but not PvuI endonuclease and were
found to cause production of PvuII endonuclease or methylase activity or both
in Escherichia coli strain HB101.  The approximate endonuclease and methylase
gene boundaries were determined through subcloning, Bal31 resection,
insertional inactivation, DNA-dependent translation, and partial DNA
sequencing.  The two genes are adjacent and appear to be divergently
transcribed.  Most E. coli strains tested were poorly transformed by the
recombinant plasmids, and this was shown by subcloning and insertional
inactivation to be due to the dPvuII methylase gene.  At a low freqency, stable
methylase-producing transformants of a "methylase-sensitive" strain were
obtained, and efficiently-transformed cell mutants were isolated from them.

<>

<1>Blumenthal, R.M., Vijesurier, R.M., Carlock, L., Dunbar, J.C.
<2>C.PvuII, an unusual transcriptional activator conserved among restriction-modification systems.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>99
<5>354
<6>1999
<7>The pvuII restriction-modification system is a type II system, which means that its
restriction endonuclease and modification methyltransferase are independently active proteins.
Because the PvuII genes are carried on a plasmid their expression must be regulated such that
movement into a new host cell is initially followed by expression of the methyltransferase
gene alone, so that the new host's DNA is protected before endonuclease activity appears.
Previous studies have identified a regulatory gene (pvuIIC) between the divergently-oriented
genes for the restriction endonuclease (pvuIIM), with pvuIIC in the same orientation as and
partially overlapping pvuIIR.  The product of pvuIIC, C.PvuII, was found to act in trans and
to be required for expression of pvuIIR.  We have demonstrated that premature expression of
pvuIIC prevents establishment of the PvuII genes, consistent with the model that requiring
C.PvuII for pvuIIR expression provides a timing delay essential for protection of the new
host's DNA.  We find that the opposing pvuIIC and pvuIIM transcripts overlap by over 60
nucleotides at their 5' ends, raising the possibility that their hybridization might play a
regulatory role.  We furthermore characterize the action of C.PvuII, demonstrating that it is
a sequence-specific DNA-binding protein that binds the DNA just upstream of the pvuIIC
promoter and stimulates transcription of both pvuIIC and pvuIIR into a polycistronic mRNA. The
location of CoPvuII binding immediately adjacent to the pvuIIC transcriptional stating points
is highly unusual among transcriptional activators.  Another surprise is that we found no
evidence for a pvuIIR-specific promoter, leaving open the question of whether the endonuclease
gene is actually expressed (if at low levels) as early in establishment as the
methyltransferase gene.  We are exploring the role of predicted hairpins in the mRNA just
upstream of pvuIIR, one of which would obstruct the Shine-Dalgarno sequence and might be used
to delay the appearance of endonuclease activity.

<>

<1>Blumer-Schuette, S.E. et al.
<2>Complete Genome Sequences for the Anaerobic, Extremely Thermophilic Plant Biomass-Degrading Bacteria Caldicellulosiruptor hydrothermalis,  Caldicellulosiruptor kristjanssonii, Caldicellulosiruptor kronotskyensis,  Caldicellulosiruptor owensensis, and Cal.
<3>J. Bacteriol.
<4>193
<5>1483-1484
<6>2011
<7>The genus Caldicellulosiruptor contains the most thermophilic, plant biomass-degrading
bacteria isolated to date. Previously, genome sequences
from three cellulolytic members of this genus were reported (C.
saccharolyticus, C. bescii, and C. obsidiansis). To further explore the
physiological and biochemical basis for polysaccharide degradation within
this genus, five additional genomes were sequenced: C. hydrothermalis, C.
kristjanssonii, C. kronotskyensis, C. lactoaceticus, and C. owensensis.
Taken together, the seven completed and one draft-phase
Caldicellulosiruptor genomes suggest that, while central metabolism is
highly conserved, significant differences in glycoside hydrolase
inventories and numbers of carbohydrate transporters exist, a finding
which likely relates to variability observed in plant biomass degradation
capacity.

<>

<1>Bobst, A.M., Pauly, G.T., Keyes, R.S., Bobst, E.V.
<2>Enzymatic sequence-specific spin labeling of a DNA fragment containing the recognition sequence of EcoRI endonuclease.
<3>FEBS Lett.
<4>228
<5>33-36
<6>1988
<7>Deoxyuridine analogs spin labeled in position 5 have been enzymatically
incorporated sequence specifically into an oligodeoxyribonucleotide to form a
spin-labeled 26-mer.  The 26-mer contains the EcoRI-binding site and two labels
which are located symmetrically close to the binding site.  The labels are
separated from one another far beyond the Heisenberg spin-exchange distance.
The local base motion as determined by ESR spectroscopy is of the order of 4 ns
in the oligonucleotide duplex.  This is the same value as reported earlier for
local T motions in polynucleotide duplexes, thereby providing direct
experimental evidence that the ESR line shape of spin levels covalently
attached to nucleic acids depends primarily on the local dynamics of the
nucleic acid building blocks.

<>

<1>Boch, J., Bonas, U.
<2>Xanthomonas AvrBs3 family-type III effectors: discovery and function.
<3>Annu. Rev. Phytopathol.
<4>48
<5>419-436
<6>2010
<7>Xanthomonads are bacterial plant pathogens that cause diseases on many plant species,
including important crops. Key to pathogenicity of most Xanthomonas
pathovars is a Hrp-type III secretion (T3S) system that translocates effector
proteins into plant cells. Within the eukaryotic cell, the effectors are thought
to perform a variety of tasks to support bacterial virulence, proliferation, and
dissemination. We are only beginning to understand the host targets of different
effectors. The largest effector family found in Xanthomonas spp. is the
AvrBs3/PthA or TAL (transcription activator-like) family. TAL effectors act as
transcriptional activators in the plant cell nucleus. Specificity of TAL
effectors is determined by a novel modular DNA-binding domain. Here, we describe
the discovery of TAL effectors and their structure, activity, and host targets.

<>

<1>Bochow, S., Elliman, J., Owens, L.
<2>Bacteriophage adenine methyltransferase: a life cycle regulator? Modelled using Vibrio harveyi myovirus like.
<3>J. Appl. Microbiol.
<4>113
<5>1001-1013
<6>2012
<7>The adenine methyltransferase (DAM) gene methylates GATC sequences that have been demonstrated
in various bacteria to be a powerful gene
regulator functioning as an epigenetic switch, particularly with
virulence gene regulation. However, overproduction of DAM can lead to
mutations, giving rise to variability that may be important for
adaptation to environmental change. While most bacterial hosts carry a
DAM gene, not all bacteriophage carry this gene. Currently, there is no
literature regarding the role DAM plays in life cycle regulation of
bacteriophage. Vibriocampbellii strain 642 carries the bacteriophage
Vibrioharveyi myovirus like (VHML) that has been proven to increase
virulence. The complete genome sequence of VHML bacteriophage revealed
a putative adenine methyltransferase gene. Using VHML, a new model of
phage life cycle regulation, where DAM plays a central role between the
lysogenic and lytic states, will be hypothesized. In short, DAM
methylates the rha antirepressor gene and once methylation is removed,
homologous CI repressor protein becomes repressed and non-functional
leading to the switching to the lytic cycle. Greater understanding of
life cycle regulation at the genetic level can, in the future, lead to
the genesis of chimeric bacteriophage with greater control over their
life cycle for their safe use as probiotics within the aquaculture
industry.

<>

<1>Bochtler, M., Szczepanowski, R.H., Tamulaitis, G., Grazulis, S., Czapinska, H., Manakova, E., Siksnys, V.
<2>Nucleotide flips determine the specificity of the Ecl18kI restriction endonuclease.
<3>EMBO J.
<4>25
<5>2219-2229
<6>2006
<7>Restricion endonuclease Ecl18kI is specific for the sequence /CCNGG and cleaves it before the
outer C to generate 5 nt 5'-overhangs. It has been suggested that Ecl18kI is evolutionarily
related to NgoMIV, a 6-bp cutter that cleaves the sequence G/CCGGC and leaves 4 nt
5'-overhangs. Here, we report the crystal structure of the Ecl18kI-DNA complex at 1.7 A
resolution and compare it with the known structure of the NgoMIV-DNA complex. We find that
Ecl18kI flips both central nucleotides within the CCNGG sequence and buries the extruded bases
in pockets within the protein. Nucleotide flipping disrupts Watson-Crick base pairing, induces
a kink in the DNA and shifts the DNA register by 1 bp, making the distances between scissile
phosphates in the Ecl18kI and NgoMIV cocrystal structures nearly identical. Therefore, the two
enzymes can use a conserved DNA recognition module, yet recognize different sequences, and
form superimposable dimers, yet generate different cleavage patterns. Hence, Ecl18kI is the
first example of a restriction endonuclease that flips nucleotides to achieve specificity for
its recognition site.

<>

<1>Bocklage, H., Heeger, K., Muller-Hill, B.
<2>Cloning and characterization of the MboII restriction-modification system.
<3>Nucleic Acids Res.
<4>19
<5>1007-1013
<6>1991
<7>The two genes encoding the class IIS restriction-modification system MboII from
Moraxella bovis were cloned separately in two compatible plasmids and expressed
in E. coli RR1DeltaM15.  The nucleotide sequences of the MboII endonuclease
(R.MboII) and methylase (M.MboII) genes were determined and the putative start
codon of R.MboII was confirmed by amino acid sequence analysis.  The mboIIR
gene specifies a protein of 110 amino acids (MW:48,617) while the mboIIM gene
codes for a putative 260-residue polypeptide (MW:30,077).  Both genes are
aligned in the same orientation.  The coding region of the methylase gene ends
11 bp upstream of the start codon of the restrictase gene.  Comparing the amino
acid sequence of M.MboII with sequences of other N6 adenine methyltransferases
reveals a significant homology to M.RsrI, M.HinfI and M.DpnA.  Furthermore,
M.MboII shows homology to the N4-cytosine methyltransferase BamHI.

<>

<1>Bocsanczy, A.M., Huguet-Tapia, J.C., Norman, D.J.
<2>Whole-Genome Sequence of Ralstonia solanacearum P673, a Strain Capable of Infecting Tomato Plants at Low Temperatures.
<3>Genome Announcements
<4>2
<5>e00106-14
<6>2014
<7>Ralstonia solanacearum is the causal agent of bacterial wilt, one of the most destructive
bacterial plant diseases. We present the whole-genome sequence of the
strain P673 (phylotype IIB, sequevar 4). This strain is capable of producing
disease in tomato plants at low temperatures. P673 has 311 unique genes.

<>

<1>Bodaker, I., Sharon, I., Suzuki, M.T., Feingersch, R., Shmoish, M., Andreishcheva, E., Sogin, M.L., Rosenberg, M., Maguire, M.E., Belkin, S., Oren, A., Beja, O.
<2>Comparative community genomics in the Dead Sea: an increasingly extreme environment.
<3>ISME J.
<4>4
<5>399-407
<6>2010
<7>Owing to the extreme salinity ( approximately 10 times saltier than the
oceans), near toxic magnesium levels (approximately 2.0 M Mg(2+)), the
dominance of divalent cations, acidic pH (6.0) and high-absorbed radiation
flux rates, the Dead Sea represents a unique and harsh ecosystem. Measures
of microbial presence (microscopy, pigments and lipids) indicate that
during rare bloom events after exceptionally rainy seasons, the microbial
communities can reach high densities. However, most of the time, when the
Dead Sea level is declining and halite is precipitating from the water
column, it is difficult to reliably measure the presence of microorganisms
and their activities. Although a number of halophilic Archaea have been
previously isolated from the Dead Sea, polar lipid analyses of biomass
collected during Dead Sea blooms suggested that these isolates were not
the major components of the microbial community of these blooms. In this
study, in an effort to characterize the perennial microbial community of
the Dead Sea and compare it with bloom assemblages, we performed
metagenomic analyses of concentrated biomass from hundreds of liters of
brine and of microbial material from the last massive Dead Sea bloom. The
difference between the two conditions was reflected in community
composition and diversity, in which the bloom was different and less
diverse from the residual brine population. The distributional patterns of
microbial genes suggested Dead Sea community trends in mono- and divalent
cation metabolisms as well as in transposable elements. This may indicate
possible mechanisms and pathways enabling these microbes to survive in
such a harsh environment.

<>

<1>Boden, R. et al.
<2>Complete Genome Sequence of the Aerobic Marine Methanotroph Methylomonas methanica MC09.
<3>J. Bacteriol.
<4>193
<5>7001-7002
<6>2011
<7>Methylomonas methanica MC09 is a mesophilic, halotolerant, aerobic, methanotrophic member of
the Gammaproteobacteria, isolated from coastal
seawater. Here we present the complete genome sequence of this strain, the
first available from an aerobic marine methanotroph.

<>

<1>Boden, R., Ferriera, S., Johnson, J., Kelly, D.P., Murrell, J.C., Schafer, H.
<2>Draft genome sequence of the chemolithoheterotrophic halophilic methylotroph Methylophaga thiooxydans DMS010.
<3>J. Bacteriol.
<4>193
<5>3154-3155
<6>2011
<7>Methylophaga thiooxydans is a mesophilic, obligately halophilic bacterium that is capable of
methylotrophic growth on a range of one-carbon
compounds as well as chemolithoheterotrophic growth at the expense of
thiosulfate. Here we present the draft genome sequence of Methylophaga
thiooxydans DMS010 (DSM 22068(T), VKM B2586(T)), the type strain of the
species, which has allowed prediction of the genes involved in one-carbon
metabolism, nitrogen metabolism and other aspects of central metabolism.

<>

<1>Bodnar, J.W., Zempsky, W., Warder, D., Bergson, C., Ward, D.C.
<2>Effect of nucleotide analogs on the cleavage of DNA by the restriction enzymes AluI, DdeI, HinfI, RsaI, and TaqI.
<3>J. Biol. Chem.
<4>258
<5>15206-15213
<6>1983
<7>The cleavage of specific DNA sequences by the restriction endonucleases AluI,
DdeI, HinfI, RsaI, and TaqI has been studied by monitoring the effect of
various nucleotide modifications on the rate of DNA digestion.  Bacteriophage
fd DNA was completely substituted in one strand with a single nucleotide
analog, using an in vitro primed DNA synthesis reaction on a single-stranded
viral DNA template.  Twelve deoxynucleotide analogs were incorporated into
these DNA substrates:  2-aminopurine, 2,6-diaminopurine, deoxytubercidin,
deoxyuridine, 5-bromodeoxyuridine, 5-allylamine deoxyuridine, 5-biotinyl
deoxyuridine, deoxypseudouridine, deoxyinosine, 8-azadeoxyguanosine,
5-iododeoxycytidine, and 5-bromodeoxycytidine.  The restriction enzymes tested
varied considerably in their ability to digest hemi-substituted DNAs containing
these modified nucleotides.  Structural alterations in the base pairs
immediately adjacent to the phosphodiester bonds cleaved by the enzyme reduced
the rate of enzyme activity most dramatically, and in most cases more than a
single determinant on each base pair altered activity.  Interactions with
nucleotides outside the recognition site seem to have little importance in the
binding or catalytic activity of these enzymes.

<>

<1>Bogdanova, E., Djordjevic, M., Papapanagiotou, I., Heyduk, T., Kneale, G., Severinov, K.
<2>Transcription regulation of the type II restriction-modification system AhdI.
<3>Nucleic Acids Res.
<4>36
<5>1429-1442
<6>2008
<7>The Restriction-modification system AhdI contains two convergent transcription units, one with
genes encoding methyltransferase subunits M and S and another with genes encoding the
controller (C) protein and the restriction endonuclease (R). We show that AhdI transcription
is controlled by two independent regulatory loops that are well-optimized to ensure successful
establishment in a naive bacterial host. Transcription from the strong MS promoter is
attenuated by methylation of an AhdI site overlapping the -10 element of the promoter.
Transcription from the weak CR promoter is regulated by the C protein interaction with two
DNA-binding sites. The interaction with the promoter-distal high-affinity site activates
transcription, while interaction with the weaker promoter-proximal site represses it. Because
of high levels of cooperativity, both C protein-binding sites are always occupied in the
absence of RNA polymerase, raising a question how activated transcription is achieved. We
develop a mathematical model that is in quantitative agreement with the experiment and
indicates that RNA polymerase outcompetes C protein from the promoter-proximal-binding site.
Such an unusual mechanism leads to a very inefficient activation of the R gene transcription,
which presumably helps control the level of the endonuclease in the cell.

<>

<1>Bogdanova, E., Zakharova, M., Streeter, S., Taylor, J., Heyduk, T., Kneale, G., Severinov, K.
<2>Transcription regulation of restriction-modification system Esp1396I.
<3>Nucleic Acids Res.
<4>37
<5>3354-3366
<6>2009
<7>The convergently transcribed restriction (R) and methylase (M) genes of the
Restriction-Modification system Esp1396I are tightly regulated by a
controller (C) protein that forms part of the CR operon. We have mapped
the transcriptional start sites from each promoter and examined the
regulatory role of C.Esp1396I in vivo and in vitro. C-protein binding at
the CR and M promoters was analyzed by DNA footprinting and a range of
biophysical techniques. The distal and proximal C-protein binding sites at
the CR promoter are responsible for activation and repression,
respectively. In contrast, a C-protein dimer binds to a single site at the
M-promoter to repress the gene, with an affinity much greater than for the
CR promoter. Thus, during establishment of the system in a naive host, the
activity of the M promoter is turned off early, preventing excessive
synthesis of methylase. Mutational analysis of promoter binding sites
reveals that the tetranucleotide inverted repeats long believed to be
important for C-protein binding to DNA are less significant than
previously thought. Instead, symmetry-related elements outside of these
repeats appear to be critical for the interaction and are discussed in
terms of the recent crystal structure of C.Esp139I bound to the CR
promoter.

<>

<1>Bogdanove, A.J. et al.
<2>Two New Complete Genome Sequences Offer Insight into Host and Tissue Specificity of Plant Pathogenic Xanthomonas spp.
<3>J. Bacteriol.
<4>193
<5>5450-5464
<6>2011
<7>Xanthomonas is a large genus of bacteria that collectively cause disease on more than 300
plant species. The broad host range of the genus
contrasts with stringent host and tissue specificity for individual
species and pathovars. Whole-genome sequences of Xanthomonas campestris
pv. raphani strain 756C and X. oryzae pv. oryzicola strain BLS256,
pathogens that infect the mesophyll tissue of the leading models for plant
biology, Arabidopsis thaliana and rice, respectively, were determined and
provided insight into the genetic determinants of host and tissue
specificity. Comparisons were made with genomes of closely related strains
that infect the vascular tissue of the same hosts and across a larger
collection of complete Xanthomonas genomes. The results suggest a model in
which complex sets of adaptations at the level of gene content account for
host specificity and subtler adaptations at the level of amino acid or
noncoding regulatory nucleotide sequence determine tissue specificity.

<>

<1>Bogdarina, I.G., Burianov, I.I., Baev, A.A.
<2>DNA modification in vitro by E. coli C and E. coli MRE 600 DNA-cytosine-methyltransferases: increased resistance of bacteriophage lambda DNA to the RII restriction system.
<3>Dokl. Akad. Nauk.
<4>233
<5>498-501
<6>1977
<7>
<>

<1>Bogdarina, I.G., Buryanov, Y.I., Bayev, A.A.
<2>Isolation and properties of DNA-cytosine-methyltransferase EcoRII and E. coli K12.
<3>Biokhimiia
<4>44
<5>440-452
<6>1979
<7>The methods of isolation and partial purification of two DNA-cytosine-methylases EcoRII and E.
coli K12 are described.  After chromatography on phosphocellulose the enzymes were purified
100-fold, the yield being 30%.  Further purification of the enzymes was performed by
sedimentation in a sucrose concentration gradient.  Both enzymes have native molecular weights
of 50,000; DC-methylase from E. coli K12 may simultaneously occur in the forms with molecular
weights of 70,000, 90,000 and 110,000.  Both DC-methylases modify identical nucleotide
sequences of DNA, have equal numbers of methylation sites in phage lambda DNA and provide in
vitro a complete protection of phage lambda DNA against restriction endonuclease EcoRII.
DC-methylases E. coli K12 and EcoRII differ in their chromatographic behavior on
phosphocellulose and capacity to form complexes with the cell DNA-adenine-methylase.

<>

<1>Bogdarina, I.G., Buryanov, Y.I., Bayev, A.A.
<2>Isolation and properties of DNA-cytosine-methyl-transferase from Escherichia coli C.
<3>Mol. Biol. (Mosk)
<4>13
<5>281-291
<6>1979
<7>The method of isolation and partial purification of DNA-cytosine-methyltransferase from E.
coli C is described.  The enzyme underwent approximately 100-fold purification.  The obtained
preparation of DC-methylase can be additionally considerably purified by sedimentation in
sucrose gradient.  Native molecular weight of DC-methylase from E. coli C is 70,000.  The
activity of the enzyme does not depend on Mg2+ ions.  The DC-methylase from E. coli C provides
DNA of lambda phage in vitro with full resistance against restriction endonuclease EcoRII.  In
DNA methylated by DC-methylase the modified cytosine, mainly in C-MC and C-MC-T sequences,
corresponds to the pyrimidine sequences of specific site EcoRII.  DNA of lambda.B phage
contains approximately 80 sites for modification by DC-methylase E. coli C.  The results
obtained point to the same specificity in vitro of DNA-cytosine-methylase E. coli C and
DNA-methylase EcoRII.

<>

<1>Bogdarina, I.G., Nadirova, I.M., Buryanov, Y.I.
<2>A technique for the detection of new strains producing enzymes involved in DNA modification and restriction - isoschizomers and isomethylomers of the known restrictases and methylases.
<3>Mikrobiologiia
<4>55
<5>699-700
<6>1986
<7>The paper describes a technique for the detection of new strains producing enzymes which
mediate DNA modification and restriction, and isoschizomers and isomethylomers of the known
restriction endonucleases and methylases.  Three Bacillus subtilis strains whose DNA carries a
BamHI modification have been found.  Two of these strains exert the restrictase activity with
an R.BamHI specificity.

<>

<1>Bogdarina, I.G., Reuter, M., Kruger, D.H., Buryanov, Y.I., Baev, A.A.A.
<2>Methylation of DNA of phages T3 and T7 by various types of DNA-adenine methylases and inhibition of the EcoK methylase by the ocR+ protein.
<3>Dokl. Akad. Nauk.
<4>273
<5>234-237
<6>1983
<7>In recent years interest in the study of the role of site-specific DNA methylation in cells of
prokaryotes, eukaryotes, and their viruses in the regulation of gene activity has increased
substantially. It is known that the manifestation of the activity of the ocR+ gene of
bacteriophages T3 and T7 in infected Escherichia coli cells suppresses the restriction and
modification of the virus DNA by Type I enzymes, and, moreover, the methylation of
bacteriophage T3 DNA does not occur at all, as a result of breakdown of the intracellular
S-adenosyl-L-methionine by SAMase (S-adenosyl-L-methionine hydrolase), which is also coded by
the gene 0.3 of phage T3. In view of this it is of interest to study the methylation of DNA of
phages T3 and T7 by DNA-adenine methylases EcoK and EcoDam in vivo and to determine the
influence of the ocR gene on the activity of these enzymes. For this purpose we determined the
number of CH3 groups incorporated into the phage genome by various DNA methylases in vitro to
determine the number of recognition sites in their DNA that are not methylated in vivo.

<>

<1>Bogdarina, I.G., Vagabova, L.M., Buryanov, Y.I.
<2>DNA-cytosine methylation in E. coli MRE 600 cells.
<3>FEBS Lett.
<4>68
<5>177-180
<6>1976
<7>Two DNA-cytosine methyltransferases (Dcm I and II) modifying cytosine in different nucleotide
sequences have been isolated from E. coli MRE 600 cells. Physiological role of these
methylases is still unknown but it is possible that one of them is connected with the DNA
modification and restriction system. This supposition is supported by the fact that the main
targets in DNA fro Dcm II are the pyrimidine sequences C-m5C and C-m5C-T (m5C:
5-methylcytosine) identical to pyrimidine fragments in DNA sequences, which are methylated by
DNA methylase of resistance factor RII. The work presents the data on in vivo methylation of
cytosine in E. coli MRE 600 DNA and we describe a new method for separate determination of Dcm
I and Dcm II action in the cell. Structural peculiarities of nucleotide sequences recognized
by two enzymes constitute the basis of this method. More that 90% of cytosine modified by Dcm
I appears in the sequence Pu-m5C-C-Pu and about 30% of cytosine methylated by Dcm II is
detected in the sequence Pu-C-m5C-Pu. So, separate determination of Dcm I and Dcm II activity
in the cell is based on quantitative estimation of 5-methylcytosine content at 5'- and
3'-termini of Pu-C-C-Pu- sequences of E. coli MRE 600 DNA.

<>

<1>Bohle, H., Henriquez, P., Grothusen, H., Navas, E., Sandoval, A., Bustamante, F., Bustos, P., Mancilla, M.
<2>Comparative Genome Analysis of Two Isolates of the Fish Pathogen Piscirickettsia  salmonis from Different Hosts Reveals Major Differences in Virulence-Associated  Secretion Systems.
<3>Genome Announcements
<4>2
<5>e01219-14
<6>2014
<7>Outbreaks caused by Piscirickettsia salmonis are one of the major threats to the
sustainability of the Chilean salmon industry. We report here the annotated draft
genomes of two P. salmonis isolates recovered from different salmonid species. A
comparative analysis showed that the number of virulence-associated secretion
systems constitutes a main genomic difference.

<>

<1>Bokor, A.A., van Kan, J.A., Poulter, R.T.
<2>Sexual mating of Botrytis cinerea illustrates PRP8 intein HEG activity.
<3>Fungal. Genet. Biol.
<4>47
<5>392-398
<6>2010
<7>Strains of Botrytis cinerea are polymorphic for the presence of an intein in the Prp8 gene
(intein +/-). The intein encodes a homing endonuclease
(HEG). During meiosis in an intein +/- heterozygote, the homing
endonuclease initiates intein 'homing' by inducing gene conversion. In
such meioses, the homing endonuclease triggers gene conversion of the
intein together with its flanking sequences into the empty allele. The
efficiency of gene conversion of the intein was found to be 100%. The
extent of flanking sequence affected by the gene conversion varied in
different meioses. A survey of the inteins and flanking sequences of a
group B. cinerea isolates indicates that there are two distinct variants
of the intein both of which have active HEGs. The survey also suggests
that the intein has been actively homing during the evolution of the
species and that the PRP8 intein may have entered the species by
horizontal transfer.

<>

<1>Boland, M.J.
<2>Identification and characterization of interactions between de novo DNA methyltransferase 3b and thymine-DNA glycosylase.
<3>Ph.D. Thesis, University of Nebraska, USA
<4>
<5>
<6>2007
<7>The majority of methylation found in the vertebrate genome occurs at carbon 5 of the cytosine
pyrimidine ring within CpG dinucleotides.  DNA methylation patterns are established during
early embryogenesis by the de novo methyltransferases, Dnmt3a and Dnmt3b.  These patterns play
a large role in genetic commitment and cellular differentiation via the heritable repression
of certain developmental programs.  An important in vivo function of Dnmt3b is methylation of
the centromeric and pericentromeric satellite repeats, regions of heterochromatin that are
rich in methylated CpG sites.  Methylation of these repeats contributes to the higher-order
heterochromatin structure that is essential to maintain genomic stability, repression of
aberrant mitotic recombination and assurance of proper chromosome segregation.  Mutations that
ablate Dnmt3b activity, exemplified by those found in the autosomal recessive genetic
disorder, ICF syndrome, lead to hypomethylation of the pericentromeric satellite repeats
resulting in chromatin decondensationa nd enhanced chromosomal rearrangements.  A significant
source of endogenous DNa damage is the spontaneous hydrolytic deamination of cytosine and
5-methylcytosine to uracil and thymidine, thereby generating U.G and T.C mismatches,
respectively.  If these promutagenic lesions are not repaired, C - T transition mutations
occur during replication.  Mechanisms to repair such damage have evolved in the form of
mismatch-specific DNA glycosylases.  G/T mismatch-specific thymine-DNA glycosylase initiates
post-replicative base excision repair at G.G and U.G mismatches ultimately leading to
reformation of the original C.G base pair.  Tdg was isolated as a positive protein interactor
with Dnmt3b in a yeast two-hybrid screen.  Subsequently, interaction between the endogenous
proteins was demonstrated.  It was determined that Dnmt3b and Tdg are targeted to genomic
regions of heterochromatin.  the regions/domains of Dnmt3b that mediate the interaction with
Tdg were determined to be regions that have been previously identified as necessary for
targeting Dnmt3b to pericentromeric heterochromatin.  A t.G mismatch repair assay utilizing ES
cells nullizygous for different DNA methyltransferases was developed to study the effect of
DNA methyltransferases on base excision repair.  Results of these experiments suggest that DNA
methyltransferases potentiate T.G mismatch repair.  During this course of investigation, it
was determined that a putative RNA component is essential for proper T.G mismatch repair.

<>

<1>Boland, M.J., Christman, J.K.
<2>A novel interaction between thymine DNA-glycosylase and the de novo methyltransferase, Dnmt3b.
<3>Proc. Amer. Assoc. Cancer Res.
<4>46
<5>645
<6>2005
<7>DNA methylation patterns are established shortly before gastrulation by the de novo
methyltransferases, Dnmt3a and Dnmt3b.  The majority of methylation found in the vertebrate
genome occurs at carbon 5 of cytosine (m5C) within CpG dinucleotides.  These patterns are
thought to set the stage for genetic commitment and cellular differentiation.  Aberrant DNA
methylation has been implicated in cancer and certain genetic diseases.  An important in vivo
function of Dnmt3b is methylation of the pericentric heterochromatin major satellite repeats.
DNA methylation of these repeats contributes to the higher-order chromatin structure necessary
for genome stability and proper sister chromatid segregation during mitosis.  Mutations that
ablate Dnmt3b activity such as those found in the rare recessive genetic disorder, ICF
syndrome lead to hypomethylation of the pericentric satellite repeats.  This results in
chromatin decondensation and enhanced chromosomal rearrangements.  Analogous chromosomal
abnormalities have been observed in breast carcinoma, ovarian epithelial carcinoma, and
pediatric Wilms tumors although they have not been directly attributed to Dnmt3b malfunction.
Dnmt3b is composed of multiple domains, suggesting it can form interactions with many
proteins.  In order to further elucidate how de novo methylation is regulated, we initiated a
search for proteins that interact with Dnmt3b.  Using Dnmt3b as bait in a yeast two-hybrid
screen of a murine embryonic cDNA library, we isolated thymine DNA-glycosylase (Tdg) as a
positive protein interactor.  This interaction has been confirmed by coimmunoprecipitation. We
show that both the catalytic domain and the PWWP domain of Dnmt3b are able to interact with
Tdg.  In addition, we demonstrate an in vivo nuclear interaction between Tdg and Dnmt3b in HEK
293T cells using confocal microscopy.  The interaction between Dnmt3b and Tdg is enhanced in
mitotic cells and can be induced by arresting cells in mitosis with the cell cycle inhibitors
vinblastine and demecolcine.  Spontaneous hydrolytic deamination of C or m5C residues within a
CpG dinucleotide results in U:G or T:G mismatches, respectively.  Mechanisms to repair such
damage have evolved in the form of mismatch-specific DNA glycosylases.  Tdg initiates the base
excision repair of T:G and U:G mismatches leading to reformation of a C:G base pair.  The
evidence presented here supports the hypothesis that the association of Dnmt3b and Tdg
promotes genomic stability and reduces the incidence of pericentromeric heterochromatin
rearrangement through repair of T:G and/or U:G mismatches followed by or simultaneous with
methylation of the repaired product, thereby restoring the original methylation status at that
locus.  Supported by NIH R21-CA91315 to JKC.
Note: Our subsequent studies of endogenous interactions between Dnmt3b and Tdg in murine P19
cells did not confirm the enhancement of interaction between these enzymes during mitosis.
This suggests that the result reported above may be the result of an elevated,
non-physiological level of exogenously expressed Dnmt3b and Tdg.

<>

<1>Boland, M.J., Christman, J.K.
<2>Mammalian DNA methyltransferases Catalytic mechanism, structure, and functions.
<3>Nutrients and Epigenetics, CRC Press-Taylor and Francis Group, Choi, S.W., Friso, S., 
<4>0
<5>37-65
<6>2009
<7>5-Methylcytosine was first detected in mammalian DNA 60 years ago and within six years, it was
demonstrated that the only dinucleotide with significant 5mC content was 5mC,G.  It took
almost another decade to determine that cytosine residues were enzymatically methylated after
incorporation into DNA, establishing the basis for epigenetic modulation of gene expression.
The first demonstrations that inhibition of DNA methylation could induce differentiation of
cultured cells were published in the late 1970s.  Since then, it has become increasingly clear
that DNA methylation plays a number of important roles in cellular homeostasis and regulation
of normal mammalian development.  Methylation of DNA promotes genomic stability through
repression of mitotic recombination and transposition, assuring proper chromatid segregation
and maintenance of higher-order heterochromatin structure.  Genomic methylation patterns also
play a crucial role during embryogenesis, leading to temporal transcriptional repression of
critical developmental programs during cellular differentiation through its ability to
regulate chromatin structure.  Additionally, DNA methylation is integral to the processes of
genomic imprinting and gene dosage compensation in females through inactivation of one X
chromosome.  A variety of tumors exhibit aberrant DNA methylation patterns.  The most common
change is an early global loss or reduction in DNA methylation, that is hypomethylation.  This
is followed by localized promoter hypermethylation of tumor-suppressor genes.

<>

<1>Boland, M.J., Christman, J.K.
<2>Characterization of Dnmt3b:Thymine-DNA Glycosylase Interaction and Stimulation of Thymine Glycosylase-Mediated Repair by DNA  Methyltransferase(s) and RNA.
<3>J. Mol. Biol.
<4>379
<5>492-504
<6>2008
<7>Methylation of cytosine residues in CpG dinucleotides plays an important role in epigenetic
regulation of gene expression and chromatin
structure/stability in higher eukaryotes. DNA methylation patterns are
established and maintained at CpG dinucleotides by DNA methyltransferases
(Dnmt1, Dnmt3a, and Dnmt3b). In mammals and many other eukaryotes, the CpG
dinucleotide is underrepresented in the genome. This loss is postulated to
be the result of unrepaired deamination of cytosine and 5-methylcytosine
to uracil and thymine, respectively. Two thymine glycosylases are believed
to reduce the impact of 5-methylcytosine deamination. G/T
mismatch-specific thymine-DNA glycosylase (Tdg) and methyl-CpG binding
domain protein 4 can both excise uracil or thymine at U.G and T.G
mismatches to initiate base excision repair. Here, we report the
characterization of interactions between Dnmt3b and both Tdg and
methyl-CpG binding domain protein 4. Our results demonstrate (1) that both
Tdg and Dnmt3b are colocalized to heterochromatin and (2) reduction of T.G
mismatch repair efficiency upon loss of DNA methyltransferase expression,
as well as a requirement for an RNA component for correct T.G mismatch
repair.

<>

<1>Bolduc, J.M., Spiegel, P.C., Chatterjee, P., Brady, K.L., Downing, M.E., Caprara, M.G., Waring, R.B., Stoddard, B.L.
<2>Structural and biochemical analyses of DNA and RNA binding by a bifunctional homing endonuclease and group I intron splicing factor.
<3>Genes Dev.
<4>17
<5>2875-2888
<6>2003
<7>We determined the crystal structure of a bifunctional group I intron splicing factor and
homing endonuclease, termed the I-AniI maturase, in
complex with its DNA target at 2.6 A resolution. The structure
demonstrates the remarkable structural conservation of the beta-sheet
DNA-binding motif between highly divergent enzyme subfamilies. DNA
recognition by I-AniI was further studied using nucleoside deletion and
DMS modification interference analyses. Correlation of these results with
the crystal structure provides information on the relative importance of
individual nucleotide contacts for DNA recognition. Alignment and modeling
of two homologous maturases reveals conserved basic surface residues,
distant from the DNA-binding surface, that might be involved in RNA
binding. A point mutation that introduces a single negative charge in this
region uncouples the maturase and endonuclease functions of the protein,
inhibiting RNA binding and splicing while maintaining DNA binding and
cleavage.

<>

<1>Bolhuis, H.H., Palm, P.P., Wende, A.A., Falb, M.M., Rampp, M.M., Rodriguez-Valera, F.F., Pfeiffer, F.F., Oesterhelt, D.D.
<2>The genome of the square archaeon Haloquadratum walsbyi: life at the limits of water activity.
<3>BMC Genomics
<4>7
<5>169
<6>2006
<7>ABSTRACT: BACKGROUND: The square halophilic archaeon Haloquadratum walsbyi dominates
NaCl-saturated and MgCl2 enriched aquatic ecosystems, which
imposes a serious desiccation stress, caused by the extremely low water
activity. The genome sequence was analyzed and physiological and physical
experiments were carried out in order to reveal how H. walsbyi has
specialized into its narrow and hostile ecological niche and found ways to
cope with the desiccation stress. RESULTS: A rich repertoire of proteins
involved in phosphate metabolism, phototrophic growth and extracellular
protective polymers, including the largest archaeal protein (9159 amino
acids), a homolog to eukaryotic mucins, are amongst the most outstanding
features. A relatively low GC content (47.9%), 15-20% less than in other
halophilic archaea, and one of the lowest coding densities (76.5%) known
for prokaryotes might be an indication for the specialization in its
unique environment CONCLUSIONS: Although no direct genetic indication was
found that can explain how this peculiar organism retains its square
shape, the genome revealed several unique adaptive traits that allow this
organism to thrive in its specific and extreme niche.

<>

<1>Bolker, M., Bohnert, H.U., Braun, K.H., Gorl, J., Kahmann, R.
<2>Tagging pathogenicity genes in Ustilago maydis by restriction enzyme-mediated integration (REMI).
<3>Mol. Gen. Genet.
<4>248
<5>547-552
<6>1995
<7>In the maize pathogenic fungus Ustilago maydis integration of transforming DNA at homologous
or heterologous sites is often accompanied by duplications of the DNA.  We show that it is
possible to generate single-copy integration events with high efficiency by restriction
enzyme-mediated integration (REMI).  In about 50% of cases, a plasmid that contains a single
BamHI site is integrated at chromosomal BamHI sites, if BamHI is added to the transformation
mixtures.  In the other cases it appears that integration events have also occurred
preferentially at BamHI sites, but without restoration of the recognition sites.  Using REMI
we have generated approximately 1000 insertion mutants.  Pathogenicity tests demonstrated that
about 1-2% of these mutants were unable to induce symptoms when tested in planta.  For two of
the mutants we have shown that the phenotype is linked to the insertion event.

<>

<1>Bollmann, A. et al.
<2>Complete genome sequence of Nitrosomonas sp. Is79, an ammonia oxidizing bacterium adapted to low ammonium concentrations.
<3>Standards in Genomic Sciences
<4>7
<5>469-482
<6>2013
<7>Nitrosomonas sp. Is79 is a chemolithoautotrophic ammonia-oxidizing bacterium that belongs to
the family Nitrosomonadaceae within the phylum Proteobacteria. Ammonia
oxidation is the first step of nitrification, an important process in the global
nitrogen cycle ultimately resulting in the production of nitrate. Nitrosomonas
sp. Is79 is an ammonia oxidizer of high interest because it is adapted to low
ammonium and can be found in freshwater environments around the world. The
3,783,444-bp chromosome with a total of 3,553 protein coding genes and 44 RNA
genes was sequenced by the DOE-Joint Genome Institute Program CSP 2006.

<>

<1>Bolot, S., Guy, E., Carrere, S., Barbe, V., Arlat, M., Noel, L.D.
<2>Genome Sequence of Xanthomonas campestris pv. campestris Strain Xca5.
<3>Genome Announcements
<4>1
<5>e00032-12
<6>2013
<7>An annotated high-quality draft genome sequence for Xanthomonas campestris pv. campestris race
1 strain Xca5 (originally described as X. campestris pv.
armoraciae), the causal agent of black rot on Brassicaceae plants, has been
determined. This genome sequence is a valuable resource for comparative genomics
within the campestris pathovar.

<>

<1>Bolot, S., Munoz, B.A., Cunnac, S., Ortiz, E., Szurek, B., Noel, L.D., Arlat, M., Jacques, M.A., Gagnevin, L., Portier, P., Fischer-Le, S.M., Carrere, S., Koebnik, R.
<2>Draft Genome Sequence of the Xanthomonas cassavae Type Strain CFBP 4642.
<3>Genome Announcements
<4>1
<5>e00679-13
<6>2013
<7>We report the draft genome sequence of the Xanthomonas cassavae type strain CFBP  4642, the
causal agent of bacterial necrosis on cassava plants. These data will
allow the comparison of this nonvascular pathogen with the vascular pathogen
Xanthomonas axonopodis pv. manihotis, both infecting the same host, which will
facilitate the development of diagnostic tools.

<>

<1>Bolot, S., Roux, B., Carrere, S., Jiang, B.L., Tang, J.L., Arlat, M., Noel, L.D.
<2>Genome Sequences of Three Atypical Xanthomonas campestris pv. campestris Strains, CN14, CN15, and CN16.
<3>Genome Announcements
<4>1
<5>e00465-13
<6>2013
<7>Xanthomonas campestris pv. campestris is the causal agent of black rot on Brassicaceae. The
draft genome sequences of three strains (CN14, CN15, and CN16)
that are highly aggressive on Arabidopsis have been determined. These genome
sequences present an unexpected genomic diversity in X. campestris pv.
campestris, which will be valuable for comparative analyses.

<>

<1>Bolotin, A., de Wouters, T., Schnupf, P., Bouchier, C., Loux, V., Rhimi, M., Jamet, A., Dervyn, R., Boudebbouze, S., Blottiere, H.M., Sorokin, A., Snel, J., Cerf-Bensussan, N., Gaboriau-Routhiau, V., van de Guchte, M., Maguin, E.
<2>Genome Sequence of 'Candidatus Arthromitus' sp. Strain SFB-Mouse-NL, a Commensal  Bacterium with a Key Role in Postnatal Maturation of Gut Immune Functions.
<3>Genome Announcements
<4>2
<5>e00705-14
<6>2014
<7>'Candidatus Arthromitus' sp. strain SFB-mouse-NL (SFB, segmented filamentous bacteria) is a
commensal bacterium necessary for inducing the postnatal
maturation of homeostatic innate and adaptive immune responses in the mouse gut.
Here, we report the genome sequence of this bacterium, which sets it apart from
earlier sequenced mouse SFB isolates.

<>

<1>Bolotin, A., Quinquis, B., Ehrlich, S.D., Sorokin, A.
<2>Complete Genome Sequence of Lactococcus lactis subsp. cremoris A76.
<3>J. Bacteriol.
<4>194
<5>1241-1242
<6>2012
<7>We report the complete genome sequence of Lactococcus lactis subsp. cremoris A76, a dairy
strain isolated from a cheese production outfit. Genome analysis detected
two contiguous islands fitting to the L. lactis subsp. lactis rather than to the
L. lactis subsp. cremoris lineage. This indicates the existence of genetic
exchange between the diverse subspecies, presumably related to the technological
process.

<>

<1>Bolotin, A., Wincker, P., Mauger, S., Jaillon, O., Malarme, K., Weissenbach, J., Ehrlich, S.D., Sorokin, A.
<2>The complete genome sequence of the lactic acid bacterium Lactococcus lactis ssp. lactis IL1403.
<3>Genome Res.
<4>11
<5>731-753
<6>2001
<7>Lactococcus lactis is a nonpathogenic AT-rich gram-positive bacterium closely related to the
genus Streptococcus and is the most commonly used cheese starter. It is also the
best-characterized lactic acid bacterium. We sequenced the genome of the laboratory strain
IL1403, using a novel two-step strategy that comprises diagnostic sequencing of the entire
genome and a shotgun polishing step. The genome contains 2,365,589 base pairs and encodes 2310
proteins, including 293 protein-coding genes belonging to six prophages and 43 insertion
sequence (IS) elements. Nonrandom distribution of IS elements indicates that the chromosome of
the sequenced strain may be a product of recent recombination between two closely related
genomes. A complete set of late competence genes is present, indicating the ability of L.
lactis to undergo DNA transformation. Genomic sequence revealed new possibilities for
fermentation pathways and for aerobic respiration. It also indicated a horizontal transfer of
genetic information from Lactococcus to gram-negative enteric bacteria of
Salmonella-Escherichia group.

<>

<1>Boltner, D., MacMahon, C., Pembroke, J.T., Strike, P., Osborn, A.M.
<2>R391: a Conjugative Integrating Mosaic Comprised of Phage, Plasmid, and Transposon Elements.
<3>J. Bacteriol.
<4>184
<5>5158-5169
<6>2002
<7>The conjugative, chromosomally integrating element R391 is the archetype
of the IncJ class of mobile genetic elements. Originally found in a South
African Providencia rettgeri strain, R391 carries antibiotic and mercury
resistance traits, as well as genes involved in mutagenic DNA repair.
While initially described as a plasmid, R391 has subsequently been shown
to be integrated into the bacterial chromosome, employing a phage-like
integration mechanism closely related to that of the SXT element from
Vibrio cholerae O139. Analysis of the complete 89-kb nucleotide sequence
of R391 has revealed a mosaic structure consisting of elements originating
in bacteriophages and plasmids and of transposable elements. A total of 96
open reading frames were identified; of these, 30 could not be assigned a
function. Sequence similarity suggests a relationship of large sections of
R391 to sequences from Salmonella, in particular those corresponding to
the putative conjugative transfer proteins, which are related to the
IncHI1 plasmid R27. A composite transposon carrying the kanamycin
resistance gene and a novel insertion element were identified. Challenging
the previous assumption that IncJ elements are plasmids, no plasmid
replicon was identified on R391, suggesting that they cannot replicate
autonomously.

<>

<1>Bolton, B., Comer, M.J., Kessler, C., Nesch, G.
<2>New type II restriction endonuclease Asp718I isolated from cells of Achromobacter sp. 718 DSM 2969.
<3>US Patent Office
<4>US 4746609
<5>
<6>1988
<7>Restriction endonuclease Asp 718, having the palindromic recognition sequence G^GTACC and the
cleavage site defined by the arrows, is new. The enzyme has a temperature option between 35-39
deg.C and a pH optimum at 8.5/37 deg. in tris/HCl buffer. The new enzyme is produced by
cultivating Achromobacter sp. 718, DSM 2969, and recovering the enzyme from the cells.
Preferably the enzyme is recovered by disintegrating the cells, treating the extract with
streptomycin sulphate until pptn. is complete, and recovering the supernatant. Fine
purification of the supernatant is preferably effected by affinity chromatography (preferably
on immobilised heparin), molecular sieve fractionation, and chromatography on weakly basic
anion-exchangers.

<>

<1>Bolton, B., Comer, M.J., Kessler, C., Nesch, G.
<2>New type II restriction endonuclease Asp718I isolated from cells of Achromobacter sp. 718 DSM 2969.
<3>German Patent Office
<4>DE 3512435
<5>
<6>1988
<7>Restriction endonuclease Asp 718, having the palindromic recognition sequence G^GTACC and the
cleavage site defined by the arrows, is new. The enzyme has a temperature option between 35-39
deg.C and a pH optimum at 8.5/37 deg. in tris/HCl buffer. The new enzyme is produced by
cultivating Achromobacter sp. 718, DSM 2969, and recovering the enzyme from the cells.
Preferably the enzyme is recovered by disintegrating the cells, treating the extract with
streptomycin sulphate until pptn. is complete, and recovering the supernatant. Fine
purification of the supernatant is preferably effected by affinity chromatography (preferably
on immobilised heparin), molecular sieve fractionation, and chromatography on weakly basic
anion-exchangers.

<>

<1>Bolton, B.J., Comer, M., Kessler, C.
<2>Asp700I, a novel isoschizomer of XmnI from Achromobacter species 700 recognizing 5'-GAANN/NNTTC-3'.
<3>Nucleic Acids Res.
<4>17
<5>8879
<6>1989
<7>None

<>

<1>Bolton, B.J., Holtke, H.-J., Schmitz, G.G., Jarsch, M., Kessler, C.
<2>AspHI, a novel isoschizomer of HgiAI from Achromobacter species H recognizing 5'-GWGCW/C-3'.
<3>Nucleic Acids Res.
<4>17
<5>9500
<6>1989
<7>None

<>

<1>Bolton, B.J., Jarsch, M., Gudrun, S., Kessler, C.
<2>New restriction endonuclease Ksp632I - derived from Kluyvera sp., useful for DNA analysis.
<3>European Patent Office
<4>EP 0306847 B
<5>
<6>1988
<7>The new type-II restriction endonuclease Ksp632I is characterised by the following recognition
sequence, 5' CTCTTCN^NNN 3' GAGAAGN NNN^ the arrows indicating the cleavage site. Ksp632I is
obtained by culturing Kluyvera sp. 632 (DSM 4196) and isolating the enzyme from the cells,
preferably by disrupting the cells and purifying the cell supernatant by sequential affinity
chromatography (esp. on immobilised heparin), molecular sieve chromatography and
cation-exchange chromatography.

<>

<1>Bolton, B.J., Jarsch, M., Schmitz, G., Kessler, C.
<2>Class II restriction endonuclease KspI and a process for obtaining it.
<3>US Patent Office
<4>US 4975376
<5>
<6>1990
<7>*
The present invention provides a restriction endonuclease which recognizes palindromic
sequences:

5' CCGC^GG 3' and 5' CCGC^GG 3'
3' GG^CGCC 5'     3' GG^CGCC 5'

where C* is methylated, and cleaves these sequences at the position indicated by the arrows.
This endonuclease is preferably from a microorganism of the genus Kluyvera. The present
invention also provides a process for obtaining this new restriction endonuclease and a
method for using the endonuclease.


<>

<1>Bolton, B.J., Jarsch, M., Schmitz, G., Kessler, C.
<2>New restriction endonuclease EclXI - derived from Enterobacter species.
<3>European Patent Office
<4>EP 336284 A
<5>
<6>1989
<7>EclXI is a new type-II restriction endonuclease derived from a microorganism of the genus
Enterobacter. It has the recognition sequence and cleavage site shown G^GGCCG and has a pH
optimum of 7.2-7.8. EclXI is produced by culturing Enterobacter cloacae DSM 4319; disrupting
the cells; and isolating EclXI from the cell supernatant, preferably by affinity
chromatography on immobilised heparin.

<>

<1>Bolton, B.J., Jarsch, M., Schmitz, G., Kessler, C.
<2>New restriction endonuclease KspI - derived from Kluyvera sp. for DNA analysis.
<3>European Patent Office
<4>EP 336286 A
<5>
<6>1988
<7>KspI is a new type-II restriction endonuclease derived from a microorganism of the genus
Kluyvera.  It has the recognition sequence and cleavage site shown in formula (I) and is
insensitive to methylation at the first cytosine residue.  KspI is produced by culturing
Kluyvera sp. DSM 4496; disrupting the cells; and isolating KspI from the cell supernatant,
pref. by affinity chromatography on immobilized heparin.

<>

<1>Bolton, B.J., Jarsch, M.D., Schmitz, G., Kessler, C.
<2>New restriction endonuclease AspI - derived from Achromobacter sp.
<3>European Patent Office
<4>EP 336285 A
<5>
<6>1988
<7>AspI is a new type-II restriction endonuclease derived from a microorganism of the genus
Achromobacter.  It has the recognition sequence and cleavage site shown in formula (I) and has
a temp. optimum of ca. 37 deg. C.  AspI is produced by culturing Achromobacter sp. DSM 4318;
disrupting the cells; and isolating AspI from the cell supernatant, pref. by affinity
chromatography on immobilized heparin.

<>

<1>Bolton, B.J., Kaluza, K., Herz, M.J., Berger, G., Kessler, C., Schmitz, G.
<2>Screening for novel type II restriction endonucleases.
<3>Fresenius Z. Anal. Chem.
<4>330
<5>376
<6>1988
<7>Site specific endonucleases have become indispensable tools for recombinant DNA
techniques.  Although to date the recognition sequences for 115 different class
II restriction endonucleases have been reported, additional enzymes with novel
specificities are required for the analysis and manipulation of DNA.  We have
examined a wide variety of bacteria for the presence of class II restriction
endonucleases.  Our aim was to find novel specific activities, to purity and
characterize the enzymes and to determine the specificities in respect to their
recognition sequence and cleavage position.  Hewre we give an overview on the
occurance of class II enzymes in various microorganisms tested and describe a
new class IIS enzyme discovered in Kluyvera species 632.

<>

<1>Bolton, B.J., Nesch, G., Comer, M.J., Wolf, W., Kessler, C.
<2>Asp718 from a non-pathogenic species of the genus Achromobacter: a KpnI isoschizomer generating DNA-fragments with 5'-protruding ends.
<3>FEBS Lett.
<4>182
<5>130-134
<6>1985
<7>A new type II restriction endonuclease Asp718 has been isolated from the
non-pathogenic species of Achromobacter 718.  This novel enzyme, an
isoschizomer of KpnI, recognizes and cleaves specifically within the nucleotide
sequence:5'-G^GTAC-C-3'3'-C-CATG^G-5'In contrast to KpnI, Asp718 generates
fragments with 5'-protruding single-stranded ends.  These 5'-terminal
extensions of the Asp718 fragments may be efficiently labeled with T4
polynucleotide kinase, whereas the recessed 3'-ends are suitable substrates for
the terminal labeling reaction applying Klenow enzyme.  The presence of only
one restriction activity in this Achromobacter strain facilitates the
preparation of Asp718 free of other contaminating site-specific nucleases which
could interfere with the in vitro digestion of DNA.

<>

<1>Bolton, B.J., Reischl, U., Holtke, H.-J., Schmitz, G.G., Jarsch, M., Kessler, C.
<2>EclXI, a novel isoschizomer of XmaIII from Enterobacter cloacae 590 recognizing 5'-C/GGCCG-3' .
<3>Nucleic Acids Res.
<4>18
<5>381
<6>1990
<7>None

<>

<1>Bolton, B.J., Schmitz, G.G., Jarsch, M., Comer, M.J., Kessler, C.
<2>Ksp632I: a novel class IIS restriction endonuclease from Kluyvera species 632 with the asymmetric hexanucleotide recognition sequence: 5'-CTCTTCN^-3' 3'-GAGAAGNNNN^-5' .
<3>Gene
<4>66
<5>31-43
<6>1988
<7>A new class-IIS restriction endonuclease, Ksp632I, with novel sequence
specificity has been discovered in a non-pathogenic species of Kluyvera.  The
presence of only a single site-specific activity in this Kluyvera sp. strain
632 enables Ksp632I to be isolated in highly purified form free of
contaminating nucleases.  Ksp632I recognition sites and cleavage positions were
deduced using experimental and computer-assisted mapping and sequencing.  The
cleavage specificity corresponds to the sequence 5'-CTCTTCN^NNN-N-3'
3'-GAGAAGN-NNN^N-5' The enzyme recognizes an asymmetric hexanucleotide sequence
and cleaves in the presence of Mg2+ ions specific phosphodiester bonds in both
DNA strands, 1 and 4 nucleotides distal to the recognition sequence.  The
staggered cuts generate 5'-protruding ends with single-stranded
5'-phosphorylated trinucleotides.  Several slow cleavage sites for Ksp632I were
observed on lambda cI857 Sam7 DNA.  Ksp632I may complement other class-IIS
enzymes in the universal restriction approach and may serve as a tool for
generating defined unidirectional deletions or insertions.

<>

<1>Bolton, B.J., Schmitz, G.G., Jarsch, M., Kessler, C.
<2>KspI, a novel isoschizomer of SacII from Kluyvera species recognizing 5'-CCGC/GG-3'.
<3>Nucleic Acids Res.
<4>17
<5>9476
<6>1989
<7>None

<>

<1>Bolton, B.J., Schmitz, G.G., Jarsch, M., Kessler, C.
<2>AspI, a novel isoschizomer of Tht111l from Achromobacter species 699 recognizing 5'-GACN/NNGTC-3'.
<3>Nucleic Acids Res.
<4>18
<5>3422
<6>1990
<7>None

<>

<1>Bomar, L., Stephens, W.Z., Nelson, M.C., Velle, K., Guillemin, K., Graf, J.
<2>Draft Genome Sequence of Aeromonas veronii Hm21, a Symbiotic Isolate from the Medicinal Leech Digestive Tract.
<3>Genome Announcements
<4>1
<5>e00800-13
<6>2013
<7>Aeromonas veronii strain Hm21 was isolated from the digestive tract of the medicinal leech
Hirudo verbana and has been used to identify genes that are
important for host colonization. This species is also a symbiont in the gut of
zebrafish and is a pathogen of mammals and fish. We present here a 4.68-Mbp draft
genome sequence for Hm21.

<>

<1>Bomholt, C., Glaub, A., Gravermann, K., Albersmeier, A., Brinkrolf, K., Ruckert, C., Tauch, A.
<2>Whole-Genome Sequence of the Clinical Strain Corynebacterium argentoratense DSM 44202, Isolated from a Human Throat Specimen.
<3>Genome Announcements
<4>1
<5>e00793-13
<6>2013
<7>Corynebacterium argentoratense is part of the human skin microbiota and is occasionally
detected in the upper respiratory tract of patients suffering from
tonsillitis. The complete DNA sequence of the type strain DSM 44202 comprises
2,031,902 bp, yielding the smallest genome sequenced thus far for a
corynebacterium associated with humans.

<>

<1>Bonacina, J., Saavedra, L., Suarez, N.E., Sesma, F.
<2>Draft Genome Sequence of the Nonstarter Bacteriocin-Producing Strain Enterococcus mundtii CRL35.
<3>Genome Announcements
<4>2
<5>e00444-14
<6>2014
<7>Enterococcus mundtii CRL35 is a bacteriocinogenic strain isolated from an artisanal cheese of
northwestern Argentina. Here we report its draft genome
sequence, consisting of 82 contigs. In silico genomic analysis of
biotechnological properties was performed to determine the potential of this
microorganism to be used in a food model system.

<>

<1>Bonaldo, M.F., Lennon, G., Soares, M.B.
<2>Normalization and subtraction: two approaches to facilitate gene discovery.
<3>Genome Res.
<4>6
<5>791-806
<6>1996
<7>Large-scale sequencing of cDNAs randomly picked from libraries has proven
to be a very powerful approach to discover (putatively) expressed
sequences that, in turn, once mapped, may greatly expedite the process
involved in the identification and cloning of human disease genes.
However, the integrity of the data and the pace at which novel sequences
can be identified depends to a great extent on the cDNA libraries that are
used. Because altogether, in a typical cell, the mRNAs of the prevalent
and intermediate frequency classes comprise as much as 50-65% of the total
mRNA mass, but represent no more than 1000-2000 different mRNAs, redundant
identification of mRNAs of these two frequency classes is destined to
become overwhelming relatively early in any such random gene discovery
programs, thus seriously compromising their cost-effectiveness. With the
goal of facilitating such efforts, previously we developed a method to
construct directionally cloned normalized cDNA libraries and applied it to
generate infant brain (INIB) and fetal liver/spleen (INFLS) libraries,
from which a total of 45,192 and 86,088 expressed sequence tags,
respectively, have been derived. While improving the representation of the
longest cDNAs in our libraries, we developed three additional methods to
normalize cDNA libraries and generated over 35 libraries, most of which
have been contributed to our integrated Molecular Analysis of Genomes and
Their Expression (IMAGE) Consortium and thus distributed widely and used
for sequencing and mapping. In an attempt to facilitate the process of
gene discovery further, we have also developed a subtractive hybridization
approach designed specifically to eliminate (or reduce significantly the
representation of) large pools of arrayed and (mostly) sequenced clones
from normalized libraries yet to be (or just partly) surveyed. Here we
present a detailed description and a comparative analysis of four methods
that we developed and used to generate normalize cDNA libraries from human
(15), mouse (3), rat (2), as well as the parasite Schistosoma mansoni (1).
In addition, we describe the construction and preliminary characterization
of a subtracted liver/spleen library (INFLS-SI) that resulted from the
elimination (or reduction of representation) of -5000 INFLS-IMAGE clones
from the INFLS library.

<>

<1>Bonamy, C., Guyonvarch, A., Ryes, O., David, F., Leblon, G.
<2>Interspecies electro-transformation in Corynebacteria.
<3>FEMS Microbiol. Lett.
<4>66
<5>263-270
<6>1990
<7>Plasmid DNA was efficiently electro-transformed into intact cells of nine Corynebacteria
strains belonging to Brevibacterium lactofermentum, Brevibacterium flavum, Corynebacterium
glutamicum and Corynebacterium melassecola.  Relationships were explored between
transformation efficiency and parameters such as electric field strength and pulse length, DNA
concentration, physiological state and concentration of the cells.  In optimal conditions,
more than 10^7 transformants per microgram of DNA could be obtained.  Electrotransformation
with plasmid DNA isolated from different sources indicates that DNA modification may play a
role in transformation efficiency.

<>

<1>Bonfils, C., Beaulieu, N., Chan, E., Cotton-Montpetit, J., MacLeod, A.R.
<2>Characterization of the human DNA methyltransferase splice variant dnmt1b.
<3>J. Biol. Chem.
<4>275
<5>10754-10760
<6>2000
<7>Tissue- and gene-specific patterns of cytosine-DNA methylation are characteristic features of
vertebrate genomes. The generation and proper maintenance of DNA methylation patterns are
essential for embryonic development, as demonstrated by the lethal phenotypes of mice with
either a targeted disruption of Dnmt1, the gene responsible for the maintenance of DNA
methylation, or targeted disruption of Dnmt3a or Dnmt3b, the genes involved in generation of
newly formed methylation patterns. Recently, a novel mRNA, Dnmt1b, resulting from alternative
splicing of Dnmt1 was identified (Hsu, D. W., Lin, M. J., Lee, T. L., Wen, S. C., Chen, X.,
and Shen, C. K., (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 9751-9756). The abundance of
Dnmt1b mRNA was estimated by semiquantitative reverse transcription polymerase chain reaction
and was suggested to encode a major C-5 DNA methyltransferase isoform. Here we report
characterization of this novel DNA methyltransferase transcript, Dnmt1b, and its protein
product in human cell lines and in freshly isolated human peripheral blood mononuclear cells.
The abundance of Dnmt1b transcript, as determined by quantitative RNase protection analysis,
was determined to range from 6% to 25% of Dnmt1 in human cells. Second generation antisense
inhibitors targeted to the 5'- and 3'-ends of Dnmt1 inhibited the accumulation of both Dnmt1
and Dnmt1b in cells. Dnmt1b protein purified from a baculovirus expression system was
demonstrated to be a functional DNA methyltransferase, and to have Michaelis constants for
both DNA and S-adenosyl-L-methionine similar to baculovirus-expressed Dnmt1. However,
antibodies raised against Dnmt1b epitopes demonstrated that Dnmt1b protein was present at
approximately 2-5% of the level of Dnmt1 and therefore represents only a minor DNA
methyltransferase isoform in human cells.

<>

<1>Bonitz, S.G., Coruzzi, G., Thalenfeld, B.E., Tzagoloff, A.
<2>Assembly of the mitochondrial membrane system.
<3>J. Biol. Chem.
<4>255
<5>11927-11941
<6>1980
<7>The oxi3 locus of yeast mitochondrial DNA has been sequenced in Saccharomyces cerevisiae
D273-10B. The sequence was obtained from the mitochondrial genomes of a series of cytoplasmic
"petite" mutants selected for the retention of genetic markers in the oxi3 locus. The oxi3
locus has been ascertained to code for Subunit 1 of cytochrome oxidase. The Subunit 1 gene is
9,979 nucleotides long, consisting of seven to eight exons that account for only 16% of the
gene sequence. The coding sequences have been identified on the basis of protein sequence
homology with Subunit 1 of human cytochrome oxidase. The yeast Subunit 1 is 510 amino acid
residues long and has a molecular weight of 56,000. In addition to the exon sequences, the
Subunit I gene contains six to seven introns. The first four introns have long reading frames
that are continuous with the exon coding sequences. These reading frames are potentially
capable for coding for basic proteins with molecular weights ranging from 30,000 to 80,000.
The first two introns of the gene have a sequence homology of 50%, while the reading frame of
the fourth intron is 70% homologous with an intron of the apocytochrome b gene. At least five
stable transcripts have been found by Northern blot hybridizations with single-stranded DNA
probes containing either exon or intron sequences. A 1.9-kilobase transcript hybridizes only
with probes from the exon regions of the gene. This RNA species has been tentatively
identified as the fully processed messenger of Subunit 1. Other transcripts are detected with
intron probes. Three transcripts with sizes of 2.5, 2.4, and 0.85 kilobases appear to be
stable excision products from the first, second, and fifth introns.

<>

<1>Bonnassie, S., Burini, J.-F., Oreglia, J., Trautwetter, A., Patte, J.-C., Sicard, A.M.
<2>Transfer of plasmid DNA to Brevibacterium lactofermentum by electrotransformation.
<3>J. Gen. Microbiol.
<4>136
<5>2107-2112
<6>1990
<7>The Escherichia coli-Brevibacterium lactofermentum shuttle vector pBLA was introduced into
intact cells of B. lactofermentum by electrotransformation.  Several parameters of this
procedure such as voltage and cell concentration were analysed.  Optimal conditions gave an
efficiency of 10^6 transformants per microgram of DNA.  Two recalcitrant strains could be
electrotransformed when an ampicillin pretreatment step was used.  Electrotransformation
experiments using DNAse or different structural forms of plasmid DNA showed that the
electrotransformation process is quite different from natural transformation involving
competence development.  Restriction-modification-proficient B. lactofermentum could be
efficiently electrotransformed with pBLA DNA isolated from E. coli.  This
restriction-modification system therefore seems to be overcome by electrotransformation.  Thus
electrotransformation may efficiently replace the protoplast bacterial transformation method.

<>

<1>Bonnassie, S., Oreglia, J., Trautwetter, A., Sicard, A.M.
<2>Isolation and characterization of a restriction and modification deficient mutant of Brevibacterium lactofermentum.
<3>FEMS Microbiol. Lett.
<4>72
<5>143-146
<6>1990
<7>In order to facilitate genetic engineering in amino-acid producing bacteria we
have isolated two restriction-deficient Brevibacterium lactofermentum strains.
They have been selected for their ability to obtain a high yield of plaques
from CL31 phage which was grown on Corynebacterium lilium.  These mutant
strains do not restrict either phage DNA by transfection or DNA from the
shuttle vector pBLA extracted from Escherichia coli by protoplast
transformation.  These mutants have also lost modification activity.  We also
report the presence of a restriction modification system in C. lilium ATCC
15990.

<>

<1>Bonnet, I., Biebricher, A., Porte, P.L., Loverdo, C., Benichou, O., Voituriez, R., Escude, C., Wende, W., Pingoud, A., Desbiolles, P.
<2>Sliding and jumping of single EcoRV restriction enzymes on non-cognate DNA.
<3>Nucleic Acids Res.
<4>36
<5>4118-4127
<6>2008
<7>The restriction endonuclease EcoRV can rapidly locate a short recognition site within long
non-cognate DNA using 'facilitated diffusion'. This
process has long been attributed to a sliding mechanism, in which the
enzyme first binds to the DNA via nonspecific interaction and then moves
along the DNA by 1D diffusion. Recent studies, however, provided evidence
that 3D translocations (hopping/jumping) also help EcoRV to locate its
target site. Here we report the first direct observation of sliding and
jumping of individual EcoRV molecules along nonspecific DNA. Using
fluorescence microscopy, we could distinguish between a slow 1D diffusion
of the enzyme and a fast translocation mechanism that was demonstrated to
stem from 3D jumps. Salt effects on both sliding and jumping were
investigated, and we developed numerical simulations to account for both
the jump frequency and the jump length distribution. We deduced from our
study the 1D diffusion coefficient of EcoRV, and we estimated the number
of jumps occurring during an interaction event with nonspecific DNA. Our
results substantiate that sliding alternates with hopping/jumping during
the facilitated diffusion of EcoRV and, furthermore, set up a framework
for the investigation of target site location by other DNA-binding
proteins.

<>

<1>Bonnin, R.A., Nordmann, P., Carattoli, A., Poirel, L.
<2>Comparative genomics of IncL/M-type plasmids; evolution by acquisition of resistance genes and insertion sequences.
<3>Antimicrob. Agents Chemother.
<4>57
<5>674-676
<6>2013
<7>IncL/M-type plasmids R446b and R471a were originally isolated from Morganella
morganii (formerly Proteus morganii) as the first members of the IncL/M group of
multidrug resistance (MDR) plasmids (6, 7)....

<>

<1>Bonnin, R.A., Poirel, L., Carattoli, A., Nordmann, P.
<2>Characterization of an IncFII Plasmid Encoding NDM-1 from Escherichia coli ST131.
<3>PLoS ONE
<4>7
<5>E34752
<6>2012
<7>BACKGROUND: The current spread of the gene encoding the metallo-ss-lactamase
NDM-1 in Enterobacteriaceae is linked to a variety of surrounding genetic
structures and plasmid scaffolds. METHODOLOGY: The whole sequence of plasmid
pGUE-NDM carrying the bla(NDM-1) gene was determined by high-density
pyrosequencing and a genomic comparative analysis with other bla(NDM-1)-negative
IncFII was performed. PRINCIPAL FINDINGS: Plasmid pGUE-NDM replicating in
Escherichia coli confers resistance to many antibiotic molecules including
beta-lactams, aminoglycosides, trimethoprim, and sulfonamides. It is 87,022 bp
in-size and carries the two beta-lactamase genes bla(NDM-1) and bla(OXA-1),
together with three aminoglycoside resistance genes aacA4, aadA2, and aacC2.
Comparative analysis of the multidrug resistance locus contained a module
encompassing the bla(NDM-1) gene that is actually conserved among different
structures identified in other enterobacterial isolates. This module was
constituted by the bla(NDM-1) gene, a fragment of insertion sequence ISAba125 and
a bleomycin resistance encoding gene. SIGNIFICANCE: This is the first
characterized bla(NDM-1)-carrying IncFII-type plasmid. Such association between
the bla(NDM-1) gene and an IncFII-type plasmid backbone is extremely worrisome
considering that this plasmid type is known to spread efficiently, as examplified
with the worldwide dissemination of bla(CTX-M-15)-borne IncFII plasmids.

<>

<1>Bonnin, R.A., Poirel, L., Nordmann, P., Eikmeyer, F.G., Wibberg, D., Puhler, A., Schluter, A.
<2>Complete sequence of broad-host-range plasmid pNOR-2000 harbouring the metallo-beta-lactamase gene blaVIM-2 from Pseudomonas aeruginosa.
<3>J. Antimicrob. Chemother.
<4>68
<5>1060-1065
<6>2013
<7>OBJECTIVES: Metallo-beta-lactamases (MBLs) are increasingly reported not only in
Enterobacteriaceae but also in Pseudomonas spp. These enzymes hydrolyse all
beta-lactams, including carbapenems, and are not inhibited by beta-lactamase
inhibitors. The aim of this study was to fully characterize a plasmid bearing the
bla(VIM-2) MBL gene identified in a Pseudomonas aeruginosa isolate. METHODS: This
plasmid was fully sequenced by high-density pyrosequencing and annotated using
the GenDB version 2.0 annotation tool. The evaluation of the broad-host-range
replication of the pNOR-2000 replication initiation gene was assessed using
electro-transformation and conjugation assays and the distribution of this
replicase gene was evaluated using an international collection of VIM-producing
Pseudomonas spp. RESULTS: Analysis of the 21 880 bp sequence of pNOR-2000
revealed a truncated and non-functional transfer operon, in addition to novel
genes encoding a serine protease and toxin/antitoxin addiction systems. This
broad-host-range plasmid shares high gene synteny with part of the mobile genomic
island pKLC102 identified in P. aeruginosa strain C. CONCLUSIONS: We report here
the complete nucleotide sequence of plasmid pNOR-2000 from a P. aeruginosa
clinical isolate harbouring the integron-located MBL gene bla(VIM-2).

<>

<1>Bonnist, E.Y.M., Liebert, K., Dryden, D.T.F., Jeltsch, A., Jones, A.C.
<2>Using the fluorescence decay of 2-aminopurine to investigate conformational change in the recognition sequence of the EcoRV DNA-(adenine-N6)-methyltransferase on enzyme binding.
<3>Biophys. Chem.
<4>160
<5>28-34
<6>2012
<7>The EcoRV DNA methyltransferase methylates the first adenine in the GATATC recognition
sequence. It is presumed that methylation proceeds
by a nucleotide flipping mechanism but no crystal structure is
available to confirm this. A popular solution-phase assay for
nucleotide flipping employs the fluorescent adenine analogue,
2-aminopurine (2AP), substituted at the methylation target site; a
substantial increase in fluorescence intensity on enzyme binding
indicates flipping. However, this appeared to fail for M.EcoRV, since
2AP substituted for the non-target adenine in the recognition sequence
showed a much greater intensity increase than 2AP at the target site.
This anomaly is resolved by recording the fluorescence decay of 2AP
which shows that the target 2AP is indeed flipped by the enzyme, but
its fluorescence is quenched by interaction with aromatic residues in
the catalytic site, whereas bending of the duplex at the non-target
site alleviates inter-base quenching and exposes the 2AP to solvent.

<>

<1>Bonocora, R.P., Belfort, M.
<2>Mapping Homing Endonuclease Cleavage Sites Using In Vitro Generated Protein.
<3>Methods Mol. Biol.
<4>1123
<5>55-67
<6>2014
<7>Mapping the precise position of endonucleolytic cleavage sites is a fundamental experimental
technique used to describe the function of a homing endonuclease. However, these proteins are
often recalcitrant to cloning and over-expression in biological systems because of toxicity
induced by spurious DNA cleavage events. In this chapter we outline the steps to successfully
express a homing endonuclease in vitro and use this product in nucleotide-resolution cleavage
assays.

<>

<1>Bonocora, R.P., Shub, D.A.
<2>A novel group I intron-encoded endonuclease specific for the anticodon region of tRNA(fMet) genes.
<3>Mol. Microbiol.
<4>39
<5>1299-1306
<6>2001
<7>Open reading frames (ORFs) are frequently inserted into group I self-splicing introns. These
ORFs encode either maturases that are
required for splicing of the intron or DNA endonucleases that promote
intron mobility. A self-splicing intron in the tRNA(fMet) gene of
Synechocystis PCC 6803, which has been proposed to have moved laterally
within the cyanobacteria, contains an ORF that is unrelated to known
intron-encoded endonucleases or maturases. Here, using an in vitro
transcription-translation system, we show that this intronic ORF
encodes a double-strand DNA endonuclease, I-Ssp6803I. I-Ssp6803I
cleaves each strand of the intronless tRNA(fMet) gene adjacent to the
anticodon triplet leaving 3 bp 3' extensions and has no activity at
intron-exon boundaries. Using an in vitro cleavage assay and scanning
deletion mutants of the intronless target site, the minimal recognition
site was determined to be a partially palindromic 20 bp region
encompassing the entire anticodon stem and loop of the tRNA(fMet) gene.
I-Ssp6803I represents a novel intron-encoded DNA endonuclease and is
the first example of a chromosomally encoded group I intron
endonuclease in bacteria.

<>

<1>Bonocora, R.P., Shub, D.A.
<2>A self-splicing group I intron in DNA polymerase genes of T7-like bacteriophages.
<3>J. Bacteriol.
<4>186
<5>8153-8155
<6>2004
<7>Group I introns are inserted into genes of a wide variety of bacteriophages of gram-positive
bacteria. However, among the phages of
enteric and other gram-negative proteobacteria, introns have been
encountered only in phage T4 and several of its close relatives. Here we
report the insertion of a self-splicing group I intron in the coding
sequence of the DNA polymerase genes of PhiI and W31, phages that are
closely related to T7. The introns belong to subgroup IA2 and both contain
an open reading frame, inserted into structural element P6a, encoding a
protein belonging to the HNH family of homing endonucleases. The introns
splice efficiently in vivo and self-splice in vitro under mild conditions
of ionic strength and temperature. We conclude that there is no barrier
for maintenance of group I introns in phages of proteobacteria.

<>

<1>Bonocora, R.P., Shub, D.A.
<2>A likely pathway for formation of mobile group I introns.
<3>Curr. Biol.
<4>19
<5>223-228
<6>2009
<7>Mobile group I introns are RNA splicing elements that have been invaded by endonuclease genes.
These endonucleases facilitate intron mobility by a
unidirectional, duplicative gene-conversion process known as homing [1].
Survival of the invading endonuclease depends upon its ability to promote
intron mobility. Therefore, the endonuclease must either quickly change
its cleavage specificity to match the site of intron insertion, or it must
already be preadapted to cleave this sequence. Here we show that the group
I intron in the DNA polymerase gene of T7-like bacteriophage PhiI is
mobile, dependent upon its intronic HNH homing endonuclease gene, I-TslI.
We also show that gene 5.3 of phage T3, located adjacent to its intronless
DNA polymerase gene, is a homologous homing endonuclease gene whose
protein product initiates efficient spread of gene 5.3 into empty sites in
related phages. Both of these endonucleases cleave intronless DNA
polymerase genes at identical positions. This shared feature between an
intronic and free-standing endonuclease is unprecedented. Based on this
evidence, we propose that introns and their homing endonucleases evolve
separately to target the same highly conserved sequences, uniting
afterwards to create a composite mobile element.

<>

<1>Boon, E.M., Salas, J.E., Barton, J.K.
<2>An electrical probe of protein-DNA interactions on DNA-modified surfaces.
<3>Nat. Biotechnol.
<4>20
<5>282-286
<6>2002
<7>DNA charge transport chemistry is found to provide a sensitive method for probing
protein-dependent changes in DNA structure and enzymatic reactions. Here we describe the
development of an electrochemical assay of protein binding to DNA-modified electrodes based
upon the detection of associated perturbations in DNA base stacking. Gold electrode surfaces
that were modified with loosely packed DNA duplexes, covalently crosslinked to a redox-active
intercalator and containing the binding site of the test protein, were constructed. Charge
transport through DNA as a function of protein binding was then assayed. Substantial
attenuation in current is seen in the presence of the base-flipping enzymes HhaI methylase and
uracil DNA glycosylase, as well as with TATA-binding protein. When restriction endonuclease
PvuII (R.PvuII) binds to its methylated target, little base-stacking perturbation occurs and
little diminution in current flow is observed. Importantly, the kinetics of restriction by
R.PvuII of its nonmethylated target is also easily monitored electrochemically. This approach
should be generally applicable to assaying protein--DNA interactions and reactions on
surfaces.

<>

<1>Boonma, P., Spinler, J.K., Qin, X., Jittaprasatsin, C., Muzny, D.M., Doddapaneni, H., Gibbs, R., Petrosino, J., Tumwasorn, S., Versalovic, J.
<2>Draft genome sequences and description of Lactobacillus rhamnosus strains L31, L34, and L35.
<3>Standards in Genomic Sciences
<4>9
<5>744-754
<6>2014
<7>Lactobacillus rhamnosus is a facultative, lactic acid bacterium in the phylum Firmicutes.
Lactobacillus spp. are generally considered beneficial, and specific
strains of L. rhamnosus are validated probiotics. We describe the draft genomes
of three L. rhamnosus strains (L31, L34, and L35) isolated from the feces of Thai
breastfed infants, which exhibit anti-inflammatory properties in vitro. The three
genomes range between 2.8 - 2.9 Mb, and contain approximately 2,700 protein
coding genes.

<>

<1>Boonmak, C., Takahasi, Y., Morikawa, M.
<2>Draft Genome Sequence of Geobacillus thermoleovorans Strain B23.
<3>Genome Announcements
<4>1
<5>e00944-13
<6>2013
<7>Here, we report the draft genome sequence of Geobacillus thermoleovorans strain B23, which was
isolated from a deep subterranean petroleum reservoir in Japan. An
array of genes related to unique long-chain alkane degradation pathways in G.
thermoleovorans B23 has been identified by whole-genome analyses of this strain.

<>

<1>Borase, H.P., Patil, C.D., Salunkhe, R.B., Suryawanshi, R.K., Salunke, B.K., Patil, S.V.
<2>Inhibition of restriction endonucleases by biofunctionalized silver nanoparticles: An in vitro study.
<3>Mater. Lett.
<4>134
<5>24-26
<6>2014
<7>Ecofriendly synthesis of metal nanoparticles and their unique properties hold promise for
innovative therapeutic applications. In the present study, rapid, novel, low cost method for
synthesis of silver nanoparticles (AgNPs) using aqueous leaves extract of medicinal plant
Euphorbia heterophylla (E. heterophylla) is reported. AgNPs synthesized by plant extract
exhibited absorption peak at 422 nm in UV-vis spectroscopy. Transmission electron microscopy
micrographs revealed presence of AgNPs with average size of 13 nm with grain and triangular
shapes. Fourier Transform Infrared Spectroscopy analysis revealed role of leaves metabolites
as reducing and capping agent in synthesis of AgNPs. X ray diffraction pattern and selected
area electron diffraction study confirmed crystalline nature of AgNPs. Inhibition of DNA
cutting activity of restriction endonucleases (EcoRI, HindIII and BamHI) by biofunctionalized
AgNPs in agarose gel electrophoresis is first time reported in the present study. This
potential of AgNPs can be useful for enhancing effectiveness of phage therapy by acting as
synergistic cocktail with bacteriophages to kill bacteria via inhibition of their restriction
endonucleases.

<>

<1>Borck, K., Beggs, J.D., Brammar, W.J., Hopkins, A.S., Murray, N.E.
<2>The construction in vitro of transducing derivatives of phage lambda.
<3>Mol. Gen. Genet.
<4>146
<5>199-207
<6>1976
<7>Methods are described for the construction of plaque-forming, transducing
derivatives of phage lambda, using appropriate receptor genomes and fragments
of DNA generated by the restriction enzymes endo R.EcoRI and endo R.HindIII.
The general properties of the transducing derivatives are described and
discussed.  Plaque-forming phages carrying the E. coli trp, his, cysB, thyA,
supD, supE, supF, hsd, tna and lig genes have been isolated.

<>

<1>Borek, E., Srinivasan, P.R.
<2>The methylation of nucleic acids.
<3>Annu. Rev. Biochem.
<4>35
<5>275-298
<6>1966
<7>The evidence for the methylation of nucleic acids at the macromolecular level
has been presented in several review articles and, therefore, will not be
repeated here.  The biochemistry of the phenomenon will be emphasized in this
article and the biological implications in a companion article in which all
alterations of macromolecules effected by enzymes will be reviewed.

<>

<1>Boreskov, Y.G., Titeeva, G.R., Berlin, Y.A.
<2>Synthetic oligodeoxynucleotides in studying MspI restriction endonuclease.
<3>Bioorg. Khim.
<4>14
<5>333-339
<6>1988
<7>Interaction of MspI restriction endonuclease with a series of
oligodeoxynucleotides varying in stability of secondary structure and in
location of the restriction site, has been studied.  It is shown that a
functionally active MspI site must be double-stranded and flanked from both
sides.  Separate MspI cleavage of dodecanucleotides dCGACCCGGGATC and
dGATCCCGGGTCG is inhibited by the reaction products as well as by
non-homological hexanucleotides dGGTACC and dGGATCC (but not by dCGGCGC).
Polyethylene glycol in low concentrations (1-3%) promotes and in higher
concentrations (7-14%) inhibits the cleavage.  A scheme of MspI functioning is
suggested including the enzyme's step-by-step recognition of the restriction
site and its nonspecific interaction with flanking segments of DNA, which leads
to formation of the productive complex.

<>

<1>Borgaro, J.G., Benner, N., Zhu, Z.Y.
<2>Fidelity Index Determination of DNA Methyltransferases.
<3>PLoS ONE
<4>8
<5>e63866
<6>2013
<7>DNA methylation is the most frequent form of epigenetic modification in the cell, which
involves gene regulation in eukaryotes and protection
against restriction enzymes in prokaryotes. Even though many
methyltransferases exclusively modify their cognate sites, there have
been reports of those that exhibit promiscuity. Previous experimental
approaches used to characterize these methyltransferases do not provide
the exact concentration at which off-target methylation occurs. Here,
we present the first reported fidelity index (FI) for a number of DNA
methyltransferases. We define the FI as the ratio of the highest amount
of methyltransferase that exhibits no star activity (off-target
effects) to the lowest amount that exhibits complete modification of
the cognate site. Of the methyltransferases assayed, M. MspI and M.
AluI exhibited the highest fidelity of >= 250 and >= 500, respectively,
and do not show star activity even at very high concentrations. In
contrast, M. HaeIII, M.EcoKDam and M. BamHI have the lowest fidelity of
4, 4 and 2, respectively, and exhibit star activity at concentrations
close to complete methylation of the cognate site. The fidelity indexes
provide vital information on the usage of methyltransferases and are
especially important in applications where site specific methylation is
required.

<>

<1>Borgaro, J.G., Zhu, Z.
<2>Characterization of the 5-hydroxymethylcytosine-specific DNA restriction endonucleases.
<3>Nucleic Acids Res.
<4>41
<5>4198-4206
<6>2013
<7>In T4 bacteriophage, 5-hydroxymethylcytosine (5hmC) is incorporated into DNA during
replication. In response, bacteria may have developed
modification-dependent type IV restriction enzymes to defend the cell from
T4-like infection. PvuRts1I was the first identified restriction enzyme to
exhibit specificity toward hmC over 5-methylcytosine (5mC) and cytosine. By using
PvuRts1I as the original member, we identified and characterized a number of
homologous proteins. Most enzymes exhibited similar cutting properties to
PvuRts1I, creating a double-stranded cleavage on the 3' side of the modified
cytosine. In addition, for efficient cutting, the enzymes require two cytosines
21-22-nt apart and on opposite strands where one cytosine must be modified.
Interestingly, the specificity determination unveiled a new layer of complexity
where the enzymes not only have specificity for 5-beta-glucosylated hmC
(5betaghmC) but also 5-alpha-glucosylated hmC (5alphaghmC). In some cases, the
enzymes are inhibited by 5betaghmC, whereas in others they are inhibited by
5alphaghmC. These observations indicate that the position of the sugar ring
relative to the base is a determining factor in the substrate specificity of the
PvuRts1I homologues. Lastly, we envision that the unique properties of select
PvuRts1I homologues will permit their use as an additive or alternative tool to
map the hydroxymethylome.

<>

<1>Borges, K.M., Wetterhahn, K.E.
<2>Chromium Bound to DNA alters cleavage by restriction endonucleases.
<3>Chem. Res. Toxicol.
<4>4
<5>638-641
<6>1991
<7>None

<>

<1>Bork, P., Ouzounis, C., Casari, G., Schneider, R., Sander, C., Dolan, M., Gilbert, W., Gillevet, P.M.
<2>Exploring the Mycoplasma capricolum genome: a minimal cell reveals its physiology.
<3>Mol. Microbiol.
<4>16
<5>955-967
<6>1995
<7>We report on the analysis of 214kb of the parasitic eubacterium Mycoplasma capricolum
sequenced by genomic walking techniques. The 287 putative proteins detected to date represent
about half of the estimated total number of 500 predicted for this organism. A large fraction
of these (75%) can be assigned a likely function as a result of similarity searches. Several
important features of the functional organization of this small genome are already apparent.
Among these are (i) the expected relatively large number of enzymes involved in metabolic
transport and activation, for efficient use of host cell nutrients; (ii) the presence of
anabolic enzymes; (iii) the unexpected diversity of enzymes involved in DNA replication and
repair; and (iv) a sizeable number of orthologues (82 so far) in Escherichia coli. This survey
is beginning to provide a detailed view of how M. capricolum manages to maintain essential
cellular processes with a genome much smaller than that of its bacterial relatives.

<>

<1>Borneman, A.R., Bartowsky, E.J., McCarthy, J., Chambers, P.J.
<2>Genotypic diversity in Oenococcus oeni by high-density microarray comparative genome hybridization and whole genome sequencing.
<3>Appl. Microbiol. Biotechnol.
<4>86
<5>681-691
<6>2010
<7>Many bacteria display substantial intra-specific genomic diversity that
produces significant phenotypic variation between strains of the same
species. Understanding the genetic basis of these strain-specific
phenotypes is especially important for industrial microorganisms where
these characters match individual strains to specific industrial
processes. Oenococcus oeni, a bacterium used during winemaking, is one
such industrial species where large numbers of strains show significant
differences in commercially important industrial phenotypes. To ascertain
the basis of these phenotypic differences, the genomic content of ten wine
strains of O. oeni were mapped by array-based comparative genome
hybridization (aCGH). These strains comprised a genomically diverse group
in which large sections of the reference genome were often absent from
individual strains. To place the aCGH results in context, whole genome
sequence was obtained for one of these strains and compared with two
previously sequenced, unrelated strains. While the three strains shared a
core group of conserved ORFs, up to 10% of the coding potential of any one
strain was specific to that isolate. The genome of O. oeni is therefore
likely to be much larger than that present in any single strain and it is
these strain-specific regions that are likely to be responsible for
differences in industrial phenotypes.

<>

<1>Boronin, A.M., Kochetkov, V.V., Kulakov, L.A., Tusupbekova, G.A., Alieva, R.M.
<2>PaeD253 restriction-modification system determined by pBS253 biodegradation plasmid of Pseudomonas aeruginosa.
<3>Dokl. Akad. Nauk.
<4>304
<5>1472-1474
<6>1989
<7>None

<>

<1>Borowczyk, E., Mohan, K.N., D'Aiuto, L., Cirio, M.C., Chaillet, J.R.
<2>Identification of a region of the DNMT1 methyltransferase that regulates the maintenance of genomic imprints.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>20806-20811
<6>2009
<7>Reprogramming of DNA methylation patterns during mammalian preimplantation development
involves the concurrent maintenance of
methylation on differentially methylated domains (DMDs) of imprinted
genes and a marked reduction of global (non-DMD) genomic methylation.
In the developing mammalian embryo, one allele of a DMD is
unmethylated, and the opposite parental allele is methylated, having
inherited this methylation from the parental gamete. The maintenance of
DMDs is important for monoallelic imprinted gene expression and normal
development of the embryo. Because the DNMT1 cytosine methyltransferase
governs maintenance methylation in mammals, rearrangements of non-DMD,
but not DMD methylation in preimplantation embryos suggest that the
preimplantation DNMT1-dependent maintenance mechanism specifically
targets DMD sequences. We explored this possibility using an engineered
mouse ES cell line to screen for mutant DNMT1 proteins that protect
against the loss of DMD and/or global (non-DMD) methylation in the
absence of the wild-type endogenous DNMT1 methyltransferase. We
identified DNMT1 mutants that were defective in maintenance of
either-DMD and/or non-DMD methylation. Among these, one mutant
maintained non-DMD methylation but not imprinted DMD methylation and
another mutant maintained just DMD methylation. The mutated amino acids
of these mutants reside in a mammal-specific, disordered region near
the amino terminus of DNMT1. These findings suggest that DNMT1
participates in epigenetic reprogramming through its ability to
distinguish different categories of methylated sequences.

<>

<1>Borrel, G., Harris, H.M., Parisot, N., Gaci, N., Tottey, W., Mihajlovski, A., Deane, J., Gribaldo, S., Bardot, O., Peyretaillade, E., Peyret, P., O'Toole, P.W., Brugere, J.F.
<2>Genome Sequence of 'Candidatus Methanomassiliicoccus intestinalis' Issoire-Mx1, a Third Thermoplasmatales-Related Methanogenic Archaeon from Human Feces.
<3>Genome Announcements
<4>1
<5>e00453-13
<6>2013
<7>'Candidatus Methanomassiliicoccus intestinalis' Issoire-Mx1 is a methanogenic archaeon found
in the human gut and is a representative of the novel order of
methanogens related to Thermoplasmatales. Its complete genome sequence is
presented here.

<>

<1>Borrel, G., Harris, H.M., Tottey, W., Mihajlovski, A., Parisot, N., Peyretaillade, E., Peyret, P., Gribaldo, S., O'Toole, P.W., Brugere, J.F.
<2>Genome Sequence of 'Candidatus Methanomethylophilus alvus' Mx1201, a Methanogenic Archaeon from the Human Gut Belonging to a Seventh Order of Methanogens.
<3>J. Bacteriol.
<4>194
<5>6944-6945
<6>2012
<7>We report the draft genome sequence of 'Candidatus Methanomethylophilus alvus' Mx1201, a
methanogen present in the human gut. It was enriched from human feces
under anaerobic conditions with methanol as the substrate. Its circular genome,
of around 1.7 Mb, contains genes needed for methylotrophic methanogenesis from
methanol and tri-, di-, and monomethylamine.

<>

<1>Borriss, M., Lombardot, T., Glockner, F.O., Becher, D., Albrecht, D., Schweder, T.
<2>Genome and proteome characterization of the psychrophilic Flavobacterium bacteriophage 11b.
<3>Extremophiles
<4>11
<5>95-104
<6>2007
<7>Virion DNA of bacteriophage 11b (phi 11b), which infects a psychrophilic Flavobacterium
isolate from Arctic sea-ice, was
determined to consist of 36,012 bp. With 30.6% its GC content
corresponds to that of host-genus species and is the lowest of all
phages of Gram-negative bacteria sequenced so far. Similarities of
several of 65 predicted ORFs, genome organization and phylogeny suggest
an affiliation to 'mesophilic' nonmarine siphoviruses, e.g. to
bacteriophages SPP1 and HK97. Early genes presumably encode an
essential recombination factor (ERF), a single strand binding (SSB)
protein, an endonuclease, and a DNA methylase. The late gene segment is
likely to contain a terminase, portal, minor head, protease and a major
capsid gene. Five ORFs exhibited similarities to Bacteroidetes species
and seem to reflect the host specificity of the phage. Among
PAGE-separated virion proteins that were identified by MALDI-ToF mass
spectrometry are the portal, the major capsid, and a putative conserved
tail protein. The phi 11b genome is the first to be described of a
cultivated virus infecting a psychrophilic host as well as a
Bacteroidetes bacterium.

<>

<1>Borriss, R., Chen, X., Rueckert, C., Blom, J., Becker, A., Baumgarth, B., Fan, B., Pukall, R., Schumann, P., Sproer, C., Junge, H., Vater, J., Puhler, A., Klenk, H.P.
<2>Relationship of Bacillus amyloliquefaciens clades associated with strains DSM7T and FZB42: a proposal for Bacillus amyloliquefaciens subsp. amyloliquefaciens subsp. nov. and plantarum subsp. nov. based on their discriminating complete genome sequences.
<3>Int. J. Syst. Evol. Microbiol.
<4>61
<5>1786-1801
<6>2011
<7>The whole genome sequenced rhizobacterium FZB42 (Chen et al., 2007) and other plant-associated
Bacillus strains either designated as Bacillus amyloliquefaciens or Bacillus subtilis are used
commercially to promote growth and health of crop plants. Previous investigations revealed
that the strains represent an own ecotype related to B. amyloliquefaciens, however its exact
taxonomic position remains elusive (Reva et al., 2004). Here we have demonstrated ability to
colonize Arabidopsis roots for a group of Bacillus strains, closely related to FZB42.
According to their phenotypic traits the strains were similar to Bacillus amyloliquefaciens
DSM 7T, but differed considerably in DNA sequences of the genes encoding 16S rRNA, gyrase
subunit A (gyrA), and histidine kinase (cheA) from the type strain. Phylogenetic analysis
performed with partial 16S rRNA, gyrA, and cheA sequences revealed that plant-associated
Bacillus strains including FZB42 form a lineage, which can be discriminated from the cluster
of strains closely related to B. amyloliquefaciens DSM 7T. DNA-DNA hybridization (DDH)
performed with genomic DNAs from DSM 7T and FZB42 yielded 63.7 to 71.2 % homology. As
complementary approach, we used several genomic methods, as direct whole genome comparison,
digital DDH, and microarray-based comparative genomic hybridization (M-CGH). Plant-associated
strains were discriminated from DSM 7T and B. subtilis type strain by their different
potential to synthesize non-ribosomally lipopeptides and polyketides. According to the
differences found in marker gene sequences and the whole genomes, we propose the two B.
amyloliquefaciens subspecies, 'plantarum' for their plant-associated, and
'amyloliquefaciens', for their non-plant associated representatives. This is in line with
results of DDH, MCGH, and the MALDI TOF mass spectrometry of cellular components which are
justifying that both ecovars represent two different subspecies.

<>

<1>Bottacini, F., Dal Bello, F., Turroni, F., Milani, C., Duranti, S., Foroni, E., Viappiani, A., Strati, F., Mora, D., van Sinderen, D., Ventura, M.
<2>Complete Genome Sequence of Bifidobacterium animalis subsp. lactis BLC1.
<3>J. Bacteriol.
<4>193
<5>6387-6388
<6>2011
<7>Bifidobacterium animalis subsp. lactis BLC1 is a probiotic bacterium that is widely exploited
by food industries as the active ingredient of various
functional foods. Here we report the complete genome sequence of B.
animalis subsp. lactis BLC1, which is expected to provide insights into
the biology of this health-promoting microorganism and improve our
understanding of its phylogenetic relatedness with other members of the B.
animalis subsp. lactis taxon.

<>

<1>Bottacini, F., Milani, C., Turroni, F., Sanchez, B., Foroni, E., Duranti, S., Serafini, F., Viappiani, A., Strati, F., Ferrarini, A., Delledonne, M., Henrissat, B., Coutinho, P., Fitzgerald, G.F., Margolles, A., van Sinderen, D., Ventura, M.
<2>Bifidobacterium asteroides PRL2011 Genome Analysis Reveals Clues for Colonization of the Insect Gut.
<3>PLoS ONE
<4>7
<5>E44229
<6>2012
<7>Bifidobacteria are known as anaerobic/microaerophilic and fermentative
microorganisms, which commonly inhabit the gastrointestinal tract of various
animals and insects. Analysis of the 2,167,301 bp genome of Bifidobacterium
asteroides PRL2011, a strain isolated from the hindgut of Apis mellifera var.
ligustica, commonly known as the honey bee, revealed its predicted capability for
respiratory metabolism. Conservation of the latter gene clusters in various B.
asteroides strains enforces the notion that respiration is a common metabolic
feature of this ancient bifidobacterial species, which has been lost in currently
known mammal-derived Bifidobacterium species. In fact, phylogenomic based
analyses suggested an ancient origin of B. asteroides and indicates it as an
ancestor of the genus Bifidobacterium. Furthermore, the B. asteroides PRL2011
genome encodes various enzymes for coping with toxic products that arise as a
result of oxygen-mediated respiration.

<>

<1>Bottacini, F., O'Connell-Motherway, M., Casey, E., McDonnell, B., Mahony, J., Ventura, M., van Sinderen, D.
<2>Discovery of a conjugative megaplasmid in Bifidobacterium breve.
<3>Appl. Environ. Microbiol.
<4>81
<5>166-176
<6>2014
<7>Bifidobacterium breve is a common and sometimes very abundant inhabitant of the human gut.
Genome sequencing of B. breve JCM 7017 revealed the presence of an extrachromosomal element,
designated pMP7017 consisting of more than 190 kb, thus representing the first reported
bifidobacterial megaplasmid. In silico characterization of this element revealed several
genomic features supporting a stable establishment of the megaplasmid in its host, illustrated
by predicted CRISPR-Cas functions that are known to protect the host against intrusion of
foreign DNA. Interestingly, pMP7017 is also predicted to encode a conjugative DNA transfer
apparatus and consistent with this notion we demonstrate conjugal transfer of pMP7017 to
representative strains of B. breve and B. longum subsp.
longum. We furthermore demonstrate the presence of a megaplasmid with homology to
pMP7017 in three B. longum subsp. longum strains.

<>

<1>Bottacini, F., O'Connell-Motherway, M., Kuczynski, J., O'Connell, K.J., Serafini, F., Duranti, S., Milani, C., Turroni, F., Lugli, G.A., Zomer, A., Zhurina, D., Riedel, C., Ventura, M., van Sinderen, D.
<2>Comparative genomics of the Bifidobacterium breve taxon.
<3>BMC Genomics
<4>15
<5>170
<6>2014
<7>BACKGROUND: Bifidobacteria are commonly found as part of the microbiota of the
gastrointestinal tract (GIT) of a broad range of hosts, where their presence is
positively correlated with the host's health status. In this study, we assessed
the genomes of thirteen representatives of Bifidobacterium breve, which is not
only a frequently encountered component of the (adult and infant) human gut
microbiota, but can also be isolated from human milk and vagina. RESULTS: In
silico analysis of genome sequences from thirteen B. breve strains isolated from
different environments (infant and adult faeces, human milk, human vagina) shows
that the genetic variability of this species principally consists of hypothetical
genes and mobile elements, but, interestingly, also genes correlated with the
adaptation to host environment and gut colonization. These latter genes specify
the biosynthetic machinery for sortase-dependent pili and exopolysaccharide
production, as well as genes that provide protection against invasion of foreign
DNA (i.e. CRISPR loci and restriction/modification systems), and genes that
encode enzymes responsible for carbohydrate fermentation. Gene-trait matching
analysis showed clear correlations between known metabolic capabilities and
characterized genes, and it also allowed the identification of a gene cluster
involved in the utilization of the alcohol-sugar sorbitol. CONCLUSIONS: Genome
analysis of thirteen representatives of the B. breve species revealed that the
deduced pan-genome exhibits an essentially close trend. For this reason our
analyses suggest that this number of B. breve representatives is sufficient to
fully describe the pan-genome of this species. Comparative genomics also
facilitated the genetic explanation for differential carbon source utilization
phenotypes previously observed in different strains of B. breve.

<>

<1>Botterman, J., Zabeau, M.
<2>High-level production of the EcoRI endonuclease under the control of the pL promoter of bacteriophage lambda.
<3>Gene
<4>37
<5>229-239
<6>1985
<7>Escherichia coli strains overproducing the EcoRI restriction endonuclease have
been constructed, using lambda pL promoter expression vectors.  In a first step
we constructed endRI::lacZ gene fusions by fusing the N-terminal part of the
endRI gene with a lacZ gene fragment, whereafter the hybrid gene was positioned
randomly under the control of the pL promoter to optimize the level of
expression.  These plasmids direct the synthesis of large amounts of fusion
protein approaching 30% of the total cellular protein content.  In most cases
the overproduced protein forms enzymatically inactive intracellular aggregates.
The position of the promoter in front of the hybrid gene had little effect on
the level of expression, except in fusions directly affecting the
ribosome-binding site (RBS).  In a second step, several of these promoter-gene
configurations were used to reconstruct the intact endRI gene in appropriate
hosts producing EcoRI methylase and cI-coded repressor.  The levels of EcoRI
endonuclease overproduction were similar to that obtained for the corresponding
fusion protein, despite the fourfold difference in protein size.  Intracellular
precipitation was also observed with the overproduced EcoRI endonuclease.

<>

<1>Botterman, J., Zabeau, M.
<2>High-level production of the restriction endonucleases EcoRI and EcoRV in Escherichia coli.
<3>Arch. Int. Physiol. Biochim.
<4>93
<5>S38
<6>1985
<7>Type II restriction and modification systems constitute an attractive system for analysis of
sequence-specific DNA-protein interaction.  These systems comprise a site-specific
endonuclease, which recognizes short target sequences in DNA at which they produce double
stranded cleavages, and a companion methylase, which specifically modifies this target
sequence to protect the host's DNA against degradation by the endonuclease.  However, the
limited available quantitites and elaborate purification procedures of these proteins hinder
biochemical and crystallographic studies.  The EcoRI and EcoRV restriction and modification
system have been cloned and characterized.  The development of a versatile plasmid expression
vector system based on the strong pL and pR promoters which are negatively regulated by the cI
repressor of phage lambda, made it possible to construct strains overproducing the enzymes.
The expression can be experimentally controlled in strains producing a temperature-sensitive
repressor (cIts), which is readily switched off and on at respectively low (28C) and high
temperatures (42C).  Since restriction endonucleases are potentially lethal to the cell, a
strategy was developed in which the expression of the EcoRI endonuclease (endRI) was first
optimized in an inactive form: an endRI-lacZ hybrid gene was randomly positioned downstream
from the pL promoter.  Optimal promoter-gene configurations were used to reconstruct the
intact gene under pL promoter control in appropriate hosts producing EcoRI methylase and cI
repressor.  Production levels of the EcoRI endonuclease reached 30% total cellular protein
content upon temperature induction, yielding after purification 5mg/g cells pure active
enzyme. Similar optimization of the expression of the EcoRV endonuclease yielded only moderate
levels of synthesis (2-5% total cellular protein content), which was nonetheless useful for
purification (1mg/g cells).  Further improvements of the ribosome binding site gave better
production.  Moreover, deletion of a putative regulatory signal downstream of the gene led to
increased levels of enzyme synthesis and indicated a possible mechanism for regulation of
prokaryotic gene expression, in which protein synthesis is modulated at the level of
translation initiation. (1) J. Botterman and M. Zabeau, Gene, in press. (2) J. Botterman, D.
DeBuyser, J. Spriet, M. Zabeau and G. Vansteenkiste, Biotechnology and Bioengineering, in
press. (3) L. Bougueleret, M. Tenchini, J. Botterman and M. Zabeau, Nucl. Acids Res., in
press. (4) J. Botterman, E. DeAlmeida, L. Bougueleret and M. Zabeau, UCLA Symposia on
Molecular and Cellular Biology 1985.

<>

<1>Bouchard, D., Peton, V., Almeida, S., Le Marechal, C., Miyoshi, A., Azevedo, V., Berkova, N., Rault, L., Francois, P., Schrenzel, J., Even, S., Hernandez, D., Le Loir, Y.
<2>Genome Sequence of Staphylococcus aureus Newbould 305, a Strain Associated with Mild Bovine Mastitis.
<3>J. Bacteriol.
<4>194
<5>6292-6293
<6>2012
<7>Staphylococcus aureus is a major etiological agent of mastitis in ruminants. We report here
the genome sequence of bovine strain Newbould 305, isolated in the
1950s in a case of bovine mastitis and now used as a model strain able to
reproducibly induce chronic mastitis in cows.

<>

<1>Bouche, J.P.C.
<2>Isolation and in vitro restriction of unmethylated DNA from Escherichia coli K.
<3>Biochim. Biophys. Acta
<4>366
<5>135-142
<6>1974
<7>A previous report (Bouche, J.P.C. (1973) J. Bacteriol. 115, 752-761) has
presented evidence that a part of the DNA synthesized in a methionine starved
dnaB mutant after return to the permissive temperature is completely or almost
completely unmethylated.  We describe here the isolation of this DNA, and
either its methylation in vitro by Escherichia coli K methylases, or its
restriction by endonucleases K and RI from E. coli, and dII from Haemophilus
influenzae.  These experiments show that this DNA not only lacks methylation
due to the general (non-modifying) methylases, but also due to the K
modifiction methylase.  Furthermore, the levels of restriction of this DNA were
found to be equivalent to those of an E. coli K DNA deficient only in
modification.  We conclude that there is no significant overlap in vitro
between the sequence specificities of major non-modifying methylases and of the
endonucleases tested.

<>

<1>Boucher, C., Elbaz, M., Genin, S., Guidot, A., Prior, P.
<2>Method for detecting ralstonia solanacearum race 3 biovar 2.
<3>European Patent Office
<4>EP 2045333 A
<5>
<6>2009
<7>The invention concerns a method for the detection of Ralstonia solanacearum race 3 biovar 2
strains in a medium, comprising the determination of the presence or the absence in a sample
of the medium, of: (1) at least one first nucleic acid target having a sequence selected from
the group constituted of SEQ ID NO: 1-49, complementary sequences thereof, and homologous
sequences thereof, or (ii) at least one fragment of said first target nucleic acid, wherein
said fragment is not constituted of or comprised in a sequence selected from the group
constituted of SEQ ID NO: 111-140; whereby, if said first nucleic acid target or fragment
thereof is present in the sample, it is determined that Ralstonia solanacearum race 3 biovar 2
strain is present in the medium.

<>

<1>Boucher, I., Emond, E., Parrot, M., Moineau, S.
<2>DNA sequence analysis of three Lactococcus lactis plasmids encoding phage resistance mechanisms.
<3>J. Dairy Sci.
<4>84
<5>1610-1620
<6>2001
<7>The three Lactococcus lactis plasmids pSRQ700, pSRQ800, and pSRQ900 encode the previously
described anti-phage resistance mechanisms
LlaDCHI, AbiK, and AbiQ, respectively. Since these plasmids are likely
to be introduced into industrial Lactococcus lactis strains used to
manufacture commercial fermented dairy products, their complete DNA
sequences were determined and analyzed. The plasmids pSRQ700 (7784 bp),
pSRQ800 (7858 bp), and pSRQ900 (10,836 bp) showed a similar genetic
organization including a common lactococcal theta-type replicon. A
second replication module showing features of the pMV158 family of
rolling circle replicons was also found on pSRQ700. The theta
replication regions of the three plasmids were associated with two
additional coding regions, one of which encodes for HsdS, the
specificity subunit of the type I restriction/modification system. When
introduced into L. lactis IL1403, the HsdS of pSRQ800 and pSRQ900
conferred a weak resistance against phage P008 (936 species). These
results indicated that both HsdS subunits can complement the
chromosomally encoded type I restriction/modification system in IL1403.
The genes involved in the phage resistance systems LlaDCHI, AbiK, and
AbiQ were found in close proximity to and downstream of the replication
modules. In pSRQ800 and pSRQ900, transfer origins and putative tyrosine
recombinases were found upstream of the theta replicons. Genes encoding
recombination proteins were also found on pSRQ700. Finally, open
reading frames associated with bacteriocin production were found on
pSRQ900, but no anti-lactococcal activity was detected. Based on our
current knowledge, these three plasmids are safe and suitable for
food-grade applications.

<>

<1>Boucher, Y., Nesbo, C.L., Joss, M.J., Robinson, A., Mabbutt, B.C., Gillings, M.R., Doolittle, W.F., Stokes, H.W.
<2>Recovery and evolutionary analysis of complete integron gene cassette arrays from Vibrio.
<3>BMC Evol. Biol.
<4>6
<5>3
<6>2006
<7>ABSTRACT: BACKGROUND: Integrons are genetic elements capable of the
acquisition, rearrangement and expression of genes contained in gene
cassettes. Gene cassettes generally consist of a promoterless gene
associated with a recombination site known as a 59-base element (59-be).
Multiple insertion events can lead to the assembly of large
integron-associated cassette arrays. The most striking examples are found
in Vibrio, where such cassette arrays are widespread and can range from 30
kb to 150 kb. Besides those found in completely sequenced genomes, no such
array has yet been recovered in its entirety. We describe an approach to
systematically isolate, sequence and annotate large integron gene cassette
arrays from bacterial strains. RESULTS: The complete Vibrio sp. DAT722
integron cassette array was determined through the streamlined approach
described here. To place it in an evolutionary context, we compare the
DAT722 array to known vibrio arrays and performed phylogenetic analyses
for all of its components (integrase, 59-be sites, gene cassette encoded
genes). It differs extensively in terms of genomic context as well as gene
cassette content and organization. The phylogenetic tree of the 59-be
sites collectively found in the Vibrio gene cassette pool suggests
frequent transfer of cassettes within and between Vibrio species, with
slower transfer rates between more phylogenetically distant relatives. We
also identify multiple cases where non-integron chromosomal genes seem to
have been assembled into gene cassettes and others where cassettes have
been inserted into chromosomal locations outside integrons. CONCLUSIONS:
Our systematic approach greatly facilitates the isolation and annotation
of large integrons gene cassette arrays. Comparative analysis of the
Vibrio sp. DAT722 integron obtained through this approach to those found
in other vibrios confirms the role of this genetic element in promoting
lateral gene transfer and suggests a high rate of gene gain/loss relative
to most other loci on vibrio chromosomes. We identify a relation between
the phylogenetic distance separating two species and the rate at which
they exchange gene cassettes, interactions between the non-mobile portion
of bacterial genomes and the vibrio gene cassette pool as well as
intragenomic translocation events of integrons in vibrios.

<>

<1>Bougueleret, L., Schwarzstein, M., Tsugita, A., Zabeau, M.
<2>Characterization of the genes coding for the EcoRV restriction and modification system of Escherichia coli.
<3>Nucleic Acids Res.
<4>12
<5>3659-3677
<6>1984
<7>A plasmid encoding the recently described Eco RV restriction and modification system has been
isolated and characterized.  This plasmid, pLB1, is 6.2 kb long and carries only the EcoRV
genes.  A subclone of 3 kb has been inserted in pBR322.  The relative positions of the
endonuclease and the methylase genes were determined by the construction of a set of
overlapping deletions generated by Bal31 resection.  The DNA sequence of a 2.2 kb fragment
containing the two genes was determined.  The two genes are transcribed divergently from a 310
bp region and the assignment of the coding region has been confirmed by direct aminoacid
sequence analysis.  Possible mechanisms of the regulation of the endonuclease gene expression
at the translational level are proposed and discussed.

<>

<1>Bougueleret, L., Tenchini, M.L., Botterman, J., Zabeau, M.
<2>Overproduction of the EcoRV endonuclease and methylase.
<3>Nucleic Acids Res.
<4>13
<5>3823-3839
<6>1985
<7>Strains overproducing the EcoRV endonuclease and methylase have been obtained by inserting
each of the two genes in expression vectors containing the lambda PL promoter.  The methylase
is overproduced to a level reaching 5-10% of the total cellular proteins, which represents a
50-100 fold increase.  A 30 fold overproduction of endonuclease was achieved by randomly
positioning the EcoRV gene downstream of the lambda PL promoter.  The situation in the
endonuclease overproducing clone resembles that encountered in maxi-cells.  The strains
described here allowed a quick purification of both enzymes in sufficient amounts for
crystallisation attempts.

<>

<1>Boujelben, I., Yarza, P., Almansa, C., Villamor, J., Maalej, S., Anton, J., Santos, F.
<2>Virioplankton community structure in Tunisian solar salterns.
<3>Appl. Environ. Microbiol.
<4>78
<5>7429-7437
<6>2012
<7>The microbial community inhabiting Sfax solar salterns on the east coast of
Tunisia has been studied by means of different molecular and culture-dependent
tools that have unveiled the presence of novel microbial groups as well as a
community structure different from that of other coastal hypersaline
environments. We have focused on the study of the viral assemblages of these
salterns and their changes along the salinity gradient and over time. Viruses
from three ponds (C4, M1, and TS) encompassing salinities from moderately
hypersaline to saturated (around 14, 19, and 35%, respectively) were sampled in
May and October 2009 and analyzed by transmission electron microscopy (TEM) and
pulsed-field gel electrophoresis (PFGE). Additionally, for all three October
samples and the May TS sample, viral metagenomic DNA was cloned in fosmids, end
sequenced, and analyzed. Viral concentration, as well as virus-to-cell ratios,
increased along the salinity gradient, with around 10(10) virus-like particles
(VLPs)/ml in close-to-saturation ponds, which represents the highest viral
concentration reported so far for aquatic systems. Four distinct morphologies
could be observed with TEM (spherical, tailed, spindled, and filamentous) but
with various proportions in the different samples. Metagenomic analyses indicated
that every pond harbored a distinct viral assemblage whose G+C content could be
roughly correlated with that of the active part of the microbial community that
may have constituted the putative hosts. As previously reported for hypersaline
metaviromes, most sequences did not have matches in the databases, although some
were conserved among the Sfax metaviromes. BLASTx, BLASTp, and dinucleotide
frequency analyses indicated that (i) factors additional to salinity could be
structuring viral communities and (ii) every metavirome had unique gene contents
and dinucleotide frequencies. Comparison with hypersaline metaviromes available
in the databases indicated that the viral assemblages present in
close-to-saturation environments located thousands of kilometers apart presented
some common traits among them in spite of their differences regarding the
putative hosts. A small core metavirome for close-to-saturation systems was found
that contained 7 sequences of around 100 nucleotides (nt) whose function was not
hinted at by in silico search results, although it most likely represents
properties essential for hyperhalophilic viruses.

<>

<1>Boukharov, A.A., Cao, Y., Dotson, S.B., Koshi, J.M., Kopvalic, D.K., Liu, J., McIninch, J.D., Wu, W.
<2>Genomic plant sequences and uses thereof.
<3>US Patent Office
<4>US 7365185 B
<5>
<6>2008
<7>The present invention discloses rice genomic promoter sequences.  The promoters are
particularly suited for use in rice and other cereal crops.  Methods of modifying, producing,
and using the promoters are also disclosed.  The invention further discloses compositions,
transformed host cells, transgenic plants, and seeds containing the rice genomic promoter
sequences, and methods for preparing and using the same.

<>

<1>Bourbonniere, M., Nalbantoglu, J.
<2>The restriction enzyme AlwNI is blocked by overlapping methylation.
<3>Nucleic Acids Res.
<4>19
<5>4774
<6>1991
<7>Most strains of E. coli contain two sequence specific methylases, Dam and Dcm. The adenine
residue of the sequence GATC is methylated at the N6 position by the Dam methylase whereas the
Dcm methylase recognizes the sequences CC(A/T)GG methylating the internal cytosine at the C5
position. The commercially available restriction enzyme AlwNI (New England Biolabs) has a
recognition sequence CAGNNN^CTG and is a useful enzyme for creating deletions. One site
AGCCCCTGgcaa is a potential dcm site and AlwNI cleavage at this site is blocked by dcm
methylation.

<>

<1>Bourchis, D., Bestor, T.H.
<2>Meiotic catastrophe and retrotransposon reactivation in male germ cells lacking Dnmt3L.
<3>Nature
<4>431
<5>96-99
<6>2004
<7>Mammalian genomes employ heritable cytosine methylation in the long-term silencing of
retrotransposons and genes subject to genomic imprinting and X chromosome inactivation. Little
is known of the mechanisms that direct cytosine methylation to specific sequences. Here we
show that DNA methyltransferase 3-like (Dnmt3L (ref. 1)) is expressed in testes during a brief
perinatal period in the non-dividing precursors of spermatogonial stem cells at a stage where
retrotransposons undergo de novo methylation. Deletion of the Dnmt3L gene prevented the de
novo methylation of both long-terminal-repeat (LTR) and non-LTR retrotransposons, which were
transcribed at high levels in spermatogonia and spermatocytes. Loss of Dnmt3L from early germ
cells also caused meiotic failure in spermatocytes, which do not express Dnmt3L. Whereas
dispersed repeated sequences were demethylated in mutant germ cells, tandem repeats in
pericentric regions were methylated normally. This result indicates that the Dnmt3L protein
might have a function in the de novo methylation of dispersed repeated sequences in a
premeiotic genome scanning process that occurs in male germ cells at about the time of birth.

<>

<1>Bourchis, D., Xu, G.-L., Lin, C.-S., Bollman, B., Bestor, T.H.
<2>Dnmt3L and the establishment of maternal genomic imprints.
<3>Science
<4>294
<5>2536-2539
<6>2001
<7>Complementary sets of genes are epigenetically silenced in male
and female gametes in a process termed genomic imprinting. The Dnmt3L gene
is expressed during gametogenesis at stages where genomic imprints are
established. Targeted disruption of Dnmt3L caused azoospermia in homozygous
males, and heterozygous progeny of homozygous females died before
midgestation. Bisulfite genomic sequencing of DNA from oocytes and embryos
showed that removal of Dnmt3L prevented methylation of sequences that are
normally maternally methylated. The defect was specific to imprinted
regions, and global genome methylation levels were not affected. Lack of
maternal methylation imprints in heterozygous embryos derived from
homozygous mutant oocytes caused biallelic expression of genes that are
normally expressed only from the allele of paternal origin. The key
catalytic motifs characteristic of DNA cytosine methyltransferases have
been lost from Dnmt3L, and the protein is more likely to act as a regulator
of imprint establishment than as a DNA methyltransferase.

<>

<1>Bourgogne, A. et al.
<2>Large scale variation in Enterococcus faecalis illustrated by the genome analysis of strain OG1RF.
<3>Genome Biology
<4>9
<5>R110
<6>2008
<7>BACKGROUND: Enterococcus faecalis has emerged as a major hospital
pathogen. To explore its diversity, we sequenced E. faecalis strain OG1RF,
which is commonly used for molecular manipulation and virulence studies.
RESULTS: The 2,739,625 base pair chromosome of OG1RF was found to contain
approximately 232 kilobases unique to this strain compared to V583, the
only publicly available sequenced strain. Almost no mobile genetic
elements were found in OG1RF. The 64 areas of divergence were classified
into three categories. First, OG1RF carries 39 unique regions, including 2
CRISPR loci and a new WxL locus. Second, we found nine replacements where
a sequence specific to V583 was substituted by a sequence specific to
OG1RF. For example, the iol operon of OG1RF replaces a possible prophage
and the vanB transposon in V583. Finally, we found 16 regions that were
present in V583 but missing from OG1RF, including the proposed
pathogenicity island, several probable prophages, and the cpsCDEFGHIJK
capsular polysaccharide operon. OG1RF was more rapidly but less frequently
lethal than V583 in the mouse peritonitis model and considerably
outcompeted V583 in a murine model of urinary tract infections.
CONCLUSION: E. faecalis OG1RF carries a number of unique loci compared to
V583, but the almost complete lack of mobile genetic elements demonstrates
that this is not a defining feature of the species. Additionally, OG1RF's
effects in experimental models suggest that mediators of virulence may be
diverse between different E. faecalis strains and that virulence is not
dependent on the presence of mobile genetic elements.

<>

<1>Bouriotis, V., Zafeiropoulos, A., Clonis, Y.D.
<2>High-performance liquid chromatography for the purification of restriction endonucleases, application to BanII, SacI, and SphI.
<3>Anal. Biochem.
<4>160
<5>127-134
<6>1987
<7>Conventional fractionation methods are time consuming, thus they prolong the
time required to process low-stability restriction enzymes.  We now report a
rapid and effective two-step chromatographic method that affords high purity
endonucleases in a short time.  Accordingly, an inexpensive chromatographic
adsorbent such as phosphocellulose or dyed agarose in the first step is coupled
to a high-performance ion exchanger, namely, MonoQ, in the second step.  The
purification schemes reported here are now in routine use to prepare
high-purity BanII, SacI, and SphI as judged by the "overdigestion" and
"cut-ligate-recut" stringent quality tests.

<>

<1>Bourniquel, A.A., Bickle, T.A.
<2>Complex restriction enzymes: NTP-driven molecular motors.
<3>Biochimie
<4>84
<5>1047-1059
<6>2002
<7>Survival is assuredly the prime directive for all living organisms either as individuals or as
a species. One of the main challenges encountered by bacterial populations is the danger of
bacteriophage attacks, since infection of a single bacterium may rapidly propagate, decimating
the entire population. In order to protect themselves against this acute threat, bacteria have
developed an array of defence mechanisms, which range from preventing the infection itself via
interference with bacteriophage adsorption to the cell surface and prevention of phage DNA
injection, to degradation of the injected phage DNA. This last defence mechanism is catalysed
by the bacterial restriction-modification (R-M) systems, and in particular, by nucleoside
5'-triphosphate (NTP)-dependent restriction enzymes, e.g. type I and type III R-M systems or
the modification-dependent endonucleases. Type I and type III restriction systems have dual
properties. They may either act as methylases and protect the host's own DNA against
restriction by methylating specific residues, or they catalyse ATP-dependent endonuclease
activity so that invading foreign DNA lacking the host-specific methylation is degraded. These
defence mechanism systems are further complemented by the presence of methylation-dependent,
GTP-dependent endonucleases that restricts specifically methylated DNA. Although all three
types of endonucleases are structurally very different, they share a common functional
mechanism. They recognise and bind to specific DNA sequences but do not cleave DNA within
those target sites. They belong to the general class of DNA motor proteins, which use the free
energy associated with nucleoside 5'-triphosphate hydrolysis to translocate DNA so that the
subsequent DNA cleavage event occurs at a distance from the endonuclease recognition site.
Moreover, DNA cleavage appears to be a random process triggered upon stalling of the DNA
translocation process and requiring dimerisation of the bound endonucleases for a concerted
break of both DNA strands. In this review, we present a detailed description and analysis of
the functional mechanism of the three known NTP-dependent restriction systems: type I and type
III restriction-modification enzymes, as well as the methylation-dependent McrBC endonuclease.

<>

<1>Bourniquel, A.A., Casey, M.G., Mollet, B., Pridmore, R.D.
<2>DNA sequence and functional analysis of Lactobacillus delbrueckii subsp. lactis plasmids pN42 and pJBL2.
<3>Plasmid
<4>47
<5>153-157
<6>2002
<7>The plasmids pN42 and pJBL2 were isolated from the Lactobacillus delbrueckii subsp. lactis
strains NCC88 and JCL414. DNA sequence determination and bioinformatic analysis revealed a
strikingly conserved genetic organization containing five major, highly conserved open reading
frames (ORFs). Transformation studies indicated that ORF2 (consisting of a primase fused to a
replicative DNA helicase), ori, and ORF3 constitute the minimal requirements for replication
of pN42 in the heterologous host Lactococcus lactis. The ORF1's are predicted to encode type
I restriction-modification (R-M) system HsdS subunits with different specificities on either
plasmid, suggesting that these plasmids may be involved in host defense by expanding their
host R-M system repertoire. These plasmids constitute the basis for the construction of novel
L. delbrueckii vectors.

<>

<1>Bourniquel, A.A., Mollet, B., Bickle, T.A.
<2>Type I restriction-modification systems from Lactobacillus delbrueckii subsp. lactis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>102
<5>238
<6>2002
<7>Background: Lactobacillus delbrueckii subsp. lactis is a lactic acid bacterium used worldwide
for the production of Swiss-type hard cheeses.
Whereas other dairy starters, e.g. Lactococcus lactis or Streptococcus
thermophilus, are highly susceptible to bacteriophage infections, very
few phages are known to target L. delbrueckii ssp. suggesting these
bacteria possess a very active and reliable/versatile endogenous
defense mechanism. Type I restriction-modification (R-M) systems are
acknowledged defense mechanisms that depend on the generation of novel
specificities for adaptability. Methods: Type I R-M hsd (host
specificity for DNA) gene clusters were isolated from two strains of L.
delbrueckii subsp. lactis, sequenced and characterized. In vivo
transcription of the identified genes was verified by Northern
blotting. The hsdR (restriction), hsdM (modification) and hsdS
(specificity of DNA binding) genes were cloned into expression vectors
for overexpression in Escherichia coli. The expressed proteins were
purified, tested for activity and their recognition sites determined.
Results: Both L. delbrueckii subsp. lactis strains examined possess
type I R-M systems. The two hsd clusters (apprx8 kb) share a similar
genetic organization consisting of: (i) the hsdR, hsdM and hsdS genes
coding for a type I restriction enzyme, (ii) a second hsdS gene with a
truncated 5'-end, and (iii) a gene similar to phage integrases. The
HsdR and HsdM snbunits on both strains are highly conserved (98%
identity), whereas the hsds genes code for subunits with different
specificities i.e. the two type I restriction enzymes have different
recognition sites. Conclusion: Until recently, research on type I R-M
systems focused on the enterobacteriaceae group in which hsd genes are
but slightly conserved in-between strains. In the gram-positive
bacterium L. delbrueckii subsp. lactis type I R-M genes are
well-conserved, genetic variability being limited to the DNA binding
domains of hsdS determining enzyme specificity. The presence of
truncated hsdS and integrases genes in the hsd clusters suggests that
novel specificities might be generated by domain shuffling.

<>

<1>Boussemaer, J.P., Schrauwen, P.P., Sourrouille, J.L., Guy, P.
<2>Multiple modification/restriction systems in lactic streptococci and their significance in defining a phage-typing system.
<3>J. Dairy Res.
<4>47
<5>401-409
<6>1980
<7>The reactions between 6 strains of mesophilic lactic steptococci and their respective phages
were studied quantitatively. Of 30 nonhomologous reactions, the bacteria were fully sensitive
in 4 and restricted the phages in 23. A mathematical model was developed that was used to
identify at least 4 and probably 5 modification restriction (M/R) systems of which up to 3
were found in the same strain. The model was based on 24 measued values and correctly
predicted the values of 5 others. One of the 3 negative reactions was shown to be due to a
restriction beyond the limit of detection, a second was due to lysogeny or lack of adsorption,
but was shown to have the predicted value when the homologous phage was modified on the host
of the challenging phage. In the last reaction a measurable restriction was predicted, but
could not be proven by means of a modified phage. These results suggest M/R to be one of the
main defenses of the lactic streptococci against their phages. They explaim why host range is
not a useful criterion in the classification of phages and suggest a rational approach to the
definition of a starter rotation.

<>

<1>Boutibonnes, P.
<2>Cell death in bacterial populations: caused or programmed?  Accepted or deliberate?
<3>Med. Sci. (Paris)
<4>13
<5>73-80
<6>1997
<7>In bacterial cells, plasmids are stabilized by numerous different mechanisms.  Here we
describe three different ways which mediate plasmid maintenance by selectively killing plasmid
free cells.  In the first system, plasmid genes encode a stable toxic protein (or the stable
corresponding mRNA) and an unstable antidote (or a small unstable antisense RNA which inhibits
the translation of the messenger).  The expression of the lethality is regulated
post-translationally (e.g. ccd system from plasmid F in Escherichia coli) or
post-transcriptionally (e.g. hok system from plasmid R1 in E. coli).  In the second system
(e.g. rm genes of Pseudomonas aeruginosa R7) plasmid genes encode a stable type II restriction
enzyme and the unstable cognate methylase which offers protection from endonucleolytic attack
by the restriction enzyme.  The third system is composed of a complex operon (e.g. MccB17
system in E. coli) whose genes encode the synthesis and maturation of a cytotoxic protein and
for a polypeptide which confers resistance or immunity to the killer cell by an unknown
mechanism.  The plasmid-free cells are killed by the cytotoxic protein excreted by bacteria
carrying the plasmid.  The set of two or more genes carried by a plasmid responsible for the
lethal consequences of plasmid loss can be viewed as an addiction module or a selfish symbiote
according to Yarmolinsky and Naito, respectively; the loss of these genomic units lead the
cells to an unavoidable death.

<>

<1>Bowen, L.M., Dupureur, C.M.
<2>Investigation of restriction enzyme cofactor requirements: A relationship between metal ion properties and sequence specificity.
<3>Biochemistry
<4>42
<5>12643-12653
<6>2003
<7>Restriction enzymes are important model systems for understanding the mechanistic
contributions of metal ions to nuclease activity. These
systems are unique in that they combine distinct functions which have been
shown to depend on metal ions: high-affinity DNA binding,
sequence-specific recognition of DNA, and Mg(II)-dependent phosphodiester
cleavage. While Ca(II) and Mn(II) are commonly used to promote DNA binding
and cleavage, respectively, the metal ion properties that are critical to
the support of these functions are not clear. To address this question, we
assessed the abilities of a series of metal ions to promote DNA binding,
sequence specificity, and cleavage in the representative PvuII
endonuclease. Among the metal ions tested [Ca(II), Sr(II), Ba(II),
Eu(III), Tb(III), Cd(II), Mn(II), Co(II), and Zn(II)], only Mn(II) and
Co(II) were similar enough to Mg(II) to support detectable cleavage
activity. Interestingly, cofactor requirements for the support of DNA
binding are much more permissive; the survey of DNA binding cofactors
indicated that Cd(II) and the heavier and larger alkaline earth metal ions
Sr(II) and Ba(II) were effective cofactors, stimulating DNA binding
affinity 20-200-fold. Impressively, the trivalent lanthanides Tb(III) and
Eu(III) promoted DNA binding as efficiently as Ca(II), corresponding to an
increase in affinity over 1000-fold higher than that observed under
metal-free conditions. The trend for DNA binding affinity supported by
these ions suggests that ionic radius and charge are not critical to the
promotion of DNA binding. To examine the role of metal ions in sequence
discrimination, we determined specificity factors
[K(a)(specific)/K(a)(nonspecific)] in the presence of Cd(II), Ba(II), and
Tb(III). Most interestingly, all of these ions compromised sequence
specificity to some degree compared to Ca(II), by either increased
affinity for a noncognate sequence, decreased affinity for the cognate
sequence, or both. These results suggest that while amino acid-base
contacts are important for specificity, the properties of metal ion
cofactors at the catalytic site are also critical for sequence
discrimination. This insight is invaluable to our efforts to understand
and subsequently design sequence-specific nucleases.

<>

<1>Bowen, L.M., Muller, G., Riehl, J.P., Dupureur, C.M.
<2>Lanthanide spectroscopic studies of the dinuclear and Mg(II)-dependent PvuII restriction endonuclease.
<3>Biochemistry
<4>43
<5>15286-15295
<6>2004
<7>Type II restriction enzymes are homodimeric systems that bind four to eight base pair
palindromic recognition sequences of DNA and catalyze
metal ion-dependent phosphodiester cleavage. While Mg(II) is required
for cleavage in these enzymes, in some systems Ca(II) promotes avid
substrate binding and sequence discrimination. These properties make
them useful model systems for understanding the roles of alkaline earth
metal ions in nucleic acid processing. We have previously shown that
two Ca(II) ions stimulate DNA binding by Pvull endonuclease and that
the trivalent lanthanide ions Tb(III) and Eu(III) support subnanomolar
DNA binding in this system. Here we capitalize on this behavior,
employing a unique combination of luminescence spectroscopy and DNA
binding assays to characterize Ln(III) binding behavior by this enzyme.
Upon excitation of tyrosine residues, the emissions of both Tb(III) and
Eu(III) are enhanced severalfold. This enhancement is reduced by the
addition of a large excess of Ca(II), indicating that these ions bind
in the active site. Poor enhancements and affinities in the presence of
the active site variant E68A indicate that Glu68 is an important
Ln(III) ligand, similar to that observed with Ca(II), Mg(II), and
Mn(II). At low micromolar Eu(III) concentrations in the presence of
enzyme (10-20 muM), Eu(III) excitation F-7(0) --> D-5(0) spectra yield
one dominant peak at 579.2 nm. A second, smaller peak at 579.4 nm is
apparent at high Eu(III) concentrations (150 muM). Titration data for
both Tb(III) and Eu(III) fit well to a two-site model featuring a
strong site (K-d = 1-3 muM) and a much weaker site (K-d approximate to
100-200 muM). Experiments with the E68A variant indicate that the Glu68
side chain is not required for the binding of this second Ln(III)
equivalent; however, the dramatic increase in DNA binding affinity
around 100 muM Ln(III) for the wild-type enzyme and metal-enhanced
substrate affinity for E68A are consistent with functional relevance
for this weaker site. This discrimination of sites should make it
possible to use lanthanide substitution and lanthanide spectroscopy to
probe individual metal ion binding sites, thus adding an important tool
to the study of restriction enzyme structure and function.

<>

<1>Boybek, A., Ray, T.D., Evans, M.C., Dyson, P.J.
<2>Novel site-specific DNA modification in Streptomyces: analysis of preferred intragenic modification sites present in a 5.7 kb amplified DNA sequence.
<3>Nucleic Acids Res.
<4>26
<5>3364-3371
<6>1998
<7>Both Streptomyces lividans and Streptomyces avermitilis encode similar systems of
post-replicative DNA modification which act site-specifically on closely opposed
guanines on either strand. The modifications can be detected since they react in
vitro with an oxidative derivative of Tris, resulting in strand cleavage.
Previous analysis of the preferred modification site of plasmid pIJ101 indicated
that extensive amounts of flanking sequence, including direct and inverted repeat
structures, are required to direct modification in vivo within a central 6 bp
palindrome. We have now examined the preferred modification sites of a
chromosomal element, the 5.7 kb amplified DNA sequence (ADS5.7) found in certain
S. lividans mutants. In contrast to the pIJ101 site, each of the ADS5. 7sites is
intragenic and modified with a 10-fold reduced frequency. However, similar
extents of flanking sequence are required for authentic double-strand
modification; deletion mutants exhibited different modification profiles,
including displaced double-stranded or single-stranded modi-fication. Comparison
of different modification sites reveals conservation of the central core
sequence, but no significant similarities between flanking sequences. Enhanced
modification was detected in a cloned region of the ADS5.7, suggesting that local
DNA topology, probably influenced by both DNA supercoiling and the nature of
flanking sequences, can influence the modifying activity.

<>

<1>Boyd, A.C., Charles, I.G., Keyte, J.W., Brammar, W.J.
<2>Isolation and computer-aided characterization of MmeI, a Type II restriction endonuclease from Methylophilus methylotrophus.
<3>Nucleic Acids Res.
<4>14
<5>5255-5274
<6>1986
<7>A Type II restriction endonuclease, MmeI, has been purified from the obligate
methylotroph, Methylophilus methylotrophus.  The enzyme was shown to have the
non-palindromic recognition sequence 5' - T C C Pu A C (N) 20- 3' 3' - A G G Py
T G (N) 18 - 5' and to cleave (as indicated) on the 3' side, generating a two
nucleotide 3' projection.  Determination of the recognition sequence was
achieved using two new computer programs; RECOG, which predicts recognition
sequences from the pattern of restriction fragments obtained from DNAs of known
sequence, and GELSIM, which generates graphical simulations of DNA band
patterns obtained by gel electrophoresis of restriction digests of sequenced
DNA molecules.

<>

<1>Boye, E., Lobner-Olesen, A.
<2>The role of dam methyltransferase in the control of DNA replication in E. coli.
<3>Cell
<4>62
<5>981-989
<6>1990
<7>The timing and control of initiation of DNA replication in E. coli was studied under
conditions where the cellular level of dam methyltransferase was controlled by a
temperature-inducible promoter. Flow cytometry was used to demonstrate that the synchrony of
initiation at the several origins within each cell was critically dependent on the level of
dam methyltransferase. Initiations were shown to be synchronous only in a narrow temperature
range. The data are explained by a model where a newly replicated and therefore hemimethylated
oriC is inert for reinitiation. Such a model may be applicable to eukaryotic cells, where
classes of origins are initiated in synchrony and only once per cell cycle.

<>

<1>Boye, E., Marinus, M.G., Lobner-Olesen, A.
<2>Quantitation of Dam methyltransferase in Escherichia coli.
<3>J. Bacteriol.
<4>174
<5>1682-1685
<6>1992
<7>An antiserum against Escherichia coli Dam methyltransferase has been developed in rabbits and
employed to detect and quantitate the enzyme in immunoblots. A wild-type, rapidly growing E.
coli cell (doubling time = 30 min) was found to contain about 130 molecules of Dam
methyltransferase.

<>

<1>Boyer, H.
<2>Genetic control of restriction and modification in Escherichia coli.
<3>J. Bacteriol.
<4>88
<5>1652-1660
<6>1964
<7>Bacterial crosses with K-12 strains of Escherichia coli as Hfr donors (Hfr
Hayes, Hfr Cavalli, and Hfr P4X-6) and B/r strains of E. coli as F- recipients
were found to differ from crosses between K-12 Hfr donors and K-12 F-
recipients in two ways:  (i) recombinants (leu,pro, lac, and gal) did not
appear at discrete time intervals but did appear simultaneously 30 min after
matings were initiated, and (ii) the linkage of unselected markers to selected
markers was reduced.  Integration of a genetic region linked to the threonine
locus of K-12 into the B/r genome resulted in a hybrid which no longer gave
anomalous results in conjugation experiments.  A similar region of the B strain
was introduced into the K-12 strain, which then behaved as a typical B F-
recipient.  These observations are interpreted as the manifestation of
host-controlled modification and restriction on the E. coli chromosome.  This
was verified by experiments on the restriction and modification of the
bacteriophage lambda, F-lac, F-gal, and sex-factor, F1.  It was found that the
genetic region that controlled the mating responses of the K-12 and B/r strains
also controlled the modification and restriction properties of these two
strains.  The genes responsible for the restricting and modifying properties of
the K-12 and B strains of E. coli were found to be allelic, linked to each
other, and linked to the threonine locus.

<>

<1>Boyer, H., Scibienski, E., Slocum, H., Roulland-Dussoix, D.
<2>The in vitro restriction of the replicative form of W.T. and mutant fd phage DNA.
<3>Virology
<4>46
<5>703-710
<6>1971
<7>Tritium- and 32P-labeled replicative form DNA of wild-type and restriction-site
mutants of the male specific phage fd were used as substrates for purified
Escherichia coli B restriction endonuclease and the products of the reaction
were analyzed by zonal centrifugation.  The products of the
endonuclease-treated unmodified wild-type RF DNA were found to be two fragments
about one-third and two-thirds the size of the genome.  The product of the
endonuclease-treated unmodified RF DNA of mutants partially resistant to in
vivo restriction was a single linear molecule.  The unmodified RF of phage
mutants totally resistant to in vivo restriction was undegraded by the E. coli
B restriction endonuclease.  These observations confirm the hypothesis of Arber
and Kuhnlein (1967) that the wild-type phage genome has two sites (each of
which consists of a limited sequence of base pairs), sensitive to the E. coli B
restriction endonuclease and mutational alteration of one or both sites results
in mutants either partially or completely resistant to E. coli B restriction.
All of the seven independently altered sites were found to be insensitive to
both steps of the restriction endonuclease attack; i.e., no single strand
phosphodiester bond cleavages were detected.  However, by using nitrocellulose
membrane filters to measure the interaction of the B restriction endonuclease
with fd RF DNA, we found that one of the mutant RF formed measurable amounts of
enzyme-substrate complexes.  In one other case no measurable complexes were
formed between the endonuclease and a mutant restriction site.  This suggests
that the restriction endonuclease relies on the proper sequence of base pairs
for recognition (formation of enzyme-substrate complex) and for the enzymatic
cleavage of phosphodiester bonds.

<>

<1>Boyer, H.W.
<2>DNA restriction and modification mechanisms in bacteria.
<3>Annu. Rev. Microbiol.
<4>25
<5>153-176
<6>1971
<7>The restriction and modification of DNA as currently understood was first
recognized as a unique biological mechanism because of the effect it has on the
ability of a bacteriophage to propagate on related bacterial strains.  Similar
observations were made prior to these reports, but they were never clearly
defined or pursued on an experimental basis.  The restriction and modification
of phage lambda by several host specificities found in E. coli has been
extensively exploited and has led to the general concept that two enzymes, an
endonuclease and a methylase, are responsible for the restriction and
modification of DNA.  The restriction endonuclease makes a double-strand
scission at a specific sequence of base pairs if it is unmodified, and the
modification methylase modifies this sequence of base pairs to render it
insensitive to the restriction endonuclease.  This mechanism (hs) appears to be
quite general in bacteria, but it should be pointed out that some restriction
and modification systems (e.g., the restriction and modification of T-even
phages, 87) could have a different molecular basis.  The plan of this review is
to treat the restriction and modification of DNA as a general bacterial
mechanism.  Emphasis will be placed on the two types of hs mechanisms
associated with E. coli, and two models of protein-polynucleotide interaction
are presented to account for the two types of restriction and modification
mechanisms.

<>

<1>Boyer, H.W.
<2>Restriction and modification of DNA: enzymes and substrates.
<3>Fed. Proc.
<4>33
<5>1125-1127
<6>1974
<7>A great deal of interest in the restriction and modification of DNA has been
generated in the past few years and a symposium devoted to some aspects of this
biological mechanism was part of the 1973 Federation Meeting.

<>

<1>Boyer, H.W., Chow, L.T., Dugaiczyk, A., Hedgpeth, J., Goodman, H.M.
<2>DNA substrate site for the EcoRII restriction endonuclease and modification methylase.
<3>Nature New Biol.
<4>244
<5>40-43
<6>1973
<7>The same nucleotide sequence in double stranded DNA restricted in double
stranded DNA restricted by the EcoRII restriction endonuclease is methylated by
the EcoRII modification methylase.  The EcoRII endonucleolytic cleavages
generate DNA fragments with cohesive termini.

<>

<1>Boyer, H.W., Greene, P.J., Meagher, R.B., Betlach, M.C., Russel, D., Goodman, H.M.
<2>The methylation of DNA as the biochemical basis of host controlled modification of DNA in bacteria.
<3>FEBS J., Elsevier, Antoni, F., Farago, A., New York
<4>34
<5>23-37
<6>1975
<7>In 1962 the work of Arber and Dussoix (Arber and Dussoix, 1962; Dussoix and
Arber, 1962) provided the conceptual framework for the study of the enzymatic
basis of the host controlled modification and restriction of bacteriophage
lambda.  Restriction and modification of phage lambda had received scant
attention until this time, probably because of its peripheral role to the ideas
occupying the attention of most molecular biologists of the time.  It had been
a long established fact that when a bacterial virus had been propagated on one
bacterial host, it would have a low efficiency of plating after infection of a
related host strain (for reviews see Arber and Linn, 1969; Boyer, 1971;
Meselson et al., 1972).  However, after one round of replication on the new
host, the few virus particles which were propagated would be modified in some
way to have the capacity to subsequently plate on that strain with a high
efficiency of plating (see Table 1).

<>

<1>Boyer, H.W., Roulland-Dussoix, D.
<2>A complementation analysis of the restriction and modification of DNA in Escherichia coli.
<3>J. Mol. Biol.
<4>41
<5>459-472
<6>1969
<7>A complementation analysis of host-controlled modification and restriction of
DNA by Escherichia coli has been carried out by examining the restriction and
modification phenotypes of partial, permanent diploids containing various
arrangements of wild type and mutant restriction and modification alleles.
Intercistronic complementation was observed between three classes of
restriction and modification mutants of E. coli B, indicating that at least
three cistrons (the ram cistrons) are involved in the genetic control of the
restriction and modification of DNA.  Mutations in one cistron (ramA) result in
a loss of restriction activity but not in modification activity (r-m+).
Mutations in a second cistron (ramC) result in a loss of restriction and
modification activities (r-m-).  Mutations in  third cistron result in a loss
of modification activity and appear to be lethal unless accompanied by a
mutation in the ramA or ramC cistrons.  A fourth class of mutations, which are
linked to the other ram cistrons and are expressed phenotypically as r-m-
mutants, are trans dominant to the wild-type ram alleles.  It is not known if
this latter class of mutants represents a fourth cistron of the ram locus.
Complementation was observed between E. coli K12 and B ramA and ramC mutations
and the host specificity of the restored restriction activity was dependent on
an intact ramC cistron.  However, complementation was not detected between the
P1 and K12 or P1 and B ram alleles.  A general model for the genetic control of
the restriction and modification properties of E. coli strains and their
episomes is presented.

<>

<1>Boyer, M., Yutin, N., Pagnier, I., Barrassi, L., Fournous, G., Espinosa, L., Robert, C., Azza, S., Sun, S., Rossmann, M.G., Suzan-Monti, M., La Scola, B., Koonin, E.V., Raoult, D.
<2>Giant Marseillevirus highlights the role of amoebae as a melting pot in emergence of chimeric microorganisms.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>21848-21853
<6>2009
<7>Giant viruses such as Mimivirus isolated from amoeba found in aquatic habitats show biological
sophistication comparable to that of simple cellular life forms and seem to evolve by similar
mechanisms, including extensive gene duplication and horizontal gene transfer (HGT), possibly
in part through a viral parasite, the virophage. We report here the isolation of "Marseille"
virus, a previously uncharacterized giant virus of amoeba. The virions of Marseillevirus
encompass a 368-kb genome, a minimum of 49 proteins, and some messenger RNAs. Phylogenetic
analysis of core genes indicates that Marseillevirus is the prototype of a family of
nucleocytoplasmic large DNA viruses (NCLDV) of eukaryotes. The genome repertoire of the virus
is composed of typical NCLDV core genes and genes apparently obtained from eukaryotic hosts
and their parasites or symbionts, both bacterial and viral. We propose that amoebae are
"melting pots" of microbial evolution where diverse forms emerge, including giant viruses with
complex gene repertoires of various origins.

<>

<1>Boyle, B., Fernandez, L., Laroche, J., Kukavica-Ibrulj, I., Mendes, C.M., Hancock, R.W., Levesque, R.C.
<2>Complete Genome Sequences of Three Pseudomonas aeruginosa Isolates with Phenotypes of Polymyxin B Adaptation and Inducible Resistance.
<3>J. Bacteriol.
<4>194
<5>529-530
<6>2012
<7>Clinical 'superbug' isolates of Pseudomonas aeruginosa were previously observed to be
resistant to several antibiotics, including polymyxin B,
and/or to have a distinct, reproducible adaptive polymyxin resistance
phenotype, identified by observing 'skipped' wells (appearance of extra
turbid wells) during broth microdilution testing. Here we report the
complete assembled draft genome sequences of three such polymyxin
resistant P. aeruginosa strains (9BR, 19BR, and 213BR).

<>

<1>Boyle, B., Vaillancourt, K., Bonifait, L., Charette, S.J., Gottschalk, M., Grenier, D.
<2>Genome Sequence of the Swine Pathogen Streptococcus suis Serotype 2 Strain S735.
<3>J. Bacteriol.
<4>194
<5>6343-6344
<6>2012
<7>Streptococcus suis is a major swine pathogen responsible for significant, worldwide economic
losses in the swine industry, in addition to being an emerging
zoonotic agent. Strains of serotype 2 are the most commonly associated with
infections causing meningitis, endocarditis, and septicemia. Here we present the
genome sequence of S. suis serotype 2 strain S735.

<>

<1>Bozic, D., Grazulis, S., Siksnys, V., Huber, R.
<2>Crystal structure of Citrobacter freundii restriction endonuclease Cfr10I at 2.15 A resolution.
<3>J. Mol. Biol.
<4>255
<5>176-186
<6>1996
<7>The X-ray crystal structure of Citrobacter freundii restriction endonuclease Cfr10I has been
determined at a resolution of 2.15 Angstroms by multiple isomorphous replacement methods and
refined to an R-factor of 19.64%.  The structure of Cfr10I represents the first structure of a
restriction endonuclease recognizing a degenerate nucleotide sequence.  Structural comparison
of Cfr10I with previously solved structures of other restriction enzymes suggests that
recognition of specific sequence occurs through contacts in the major and the minor grooves of
DNA.  The arrangement of the putative active site residues shows some striking differences
from previously described restriction endonucleases and supports a two-metal-ion mechanism of
catalysis.

<>

<1>Brabec, V., Balcarova, Z.
<2>Restriction-enzyme cleavage of DNA modified by platinum (II) complexes.
<3>Eur. J. Biochem.
<4>216
<5>183-187
<6>1993
<7>The effect of binding of cis-diamminedichloroplatinum(II), its trans isomer and
diethylenetriaminechloroplatinum(II) chloride to DNA on the splicing effectiveness of BamHI,
EcoRI and SalI restriction endonucleases has been determined by means of gel electrophoresis.
All three platinum complexes inhibit the cleavage of linearized plasmid DNA. In addition, the
three platinum complexes bound to DNA constitute a barrier across which the linear diffusion
of EcoRI on DNA is difficult. We interpret these findings to mean that the splicing
effectiveness of restriction enzymes is influenced by bifunctional and monofunctional DNA
adducts of platinum via both steric interference and DNA conformational distortions. Whereas
the platinum adducts in the restriction sites or in their very closed proximity inhibit the
cleavage, the lesions occurring a greater distance from the restriction site can slow down the
process of the localization of recognition sequences.

<>

<1>Brack, C., Eberle, H., Bickle, T.A., Yuan, R.
<2>Mapping of recognition sites for the restriction endonuclease from Escherichia coli K12 on bacteriophage PM2 DNA.
<3>J. Mol. Biol.
<4>108
<5>583-593
<6>1976
<7>Two different methods have been used to locate the recognition sites for the
restriction endonuclease from Escherichia coli K12 on the genome of
bacteriophage PM2.  The first method employs purified fragments of PM2 DNA
produced by digestion with HindIII.  The fragments are tested for their ability
to support the ATPase activity of the enzyme or, alternatively, to inhibit it
in the presence of intact substrate DNA.  In this way, recognition sites were
assigned to the HindIII fragments 2, 6 and 7.  An independent approach involved
the study of complexes between the enzyme and either the HindIII fragments or
linear DNA produced by cleavage of PM2 DNA at the unique site for HpaII.
Electron microscopic analysis of such preparations allowed mapping of the three
recognition sites to 40, 60 and 62 map units from the HpaII cleavage site.

<>

<1>Bradbury, M., Greenfield, P., Midgley, D., Li, D., Tran-Dinh, N., Vriesekoop, F., Brown, J.L.
<2>Draft Genome Sequence of Clostridium sporogenes PA 3679, the Common Nontoxigenic  Surrogate for Proteolytic Clostridium botulinum.
<3>J. Bacteriol.
<4>194
<5>1631-1632
<6>2012
<7>Clostridium sporogenes PA 3679 is widely used as a nontoxigenic surrogate for proteolytic
strains of Clostridium botulinum in the derivation and validation of
thermal processes in food. Here we report the draft assembly and annotation of
the C. sporogenes PA 3679 genome. Preliminary analysis demonstrates a high degree
of relatedness between C. sporogenes PA 3679 and sequenced strains of proteolytic
C. botulinum.

<>

<1>Braga, E.A., Nosikov, V.V., Polyanovsky, O.L.
<2>The use of antibiotics actinomycin D and distamycin A for mapping of phage lambda HindIII fragments.
<3>FEBS Lett.
<4>70
<5>91-95
<6>1976
<7>The antibiotics distamycin A and actinomycin D interact with double-stranded
DNA forming noncovalently bound complexes.  This interaction is rather
selective: distamycin forms complexes preferentially with d(A/T) rich regions
whereas actinomycin binds d(G/C) rich regions of DNA.  This property has
offered a possibility to use these antibiotics for effective and highly
selective protection of the cleavage sites recognized by restriction
endonucleases.  The recognition site of endo R. HindIII 5'AAGCTT 3'TTCGAA may
be protected by both actinomycin D and distamycin A as long as it contains TT
and GC sequences favourable for interaction of distamycin and actinomycin with
template DNA.  It has been demonstrated that different EcoRI recognition sites
on lambda DNA may be effectively blocked by different antibiotic concentrations
due to the peculiarities of the environment of different recognition sites.  In
the present paper it is shown that certain endo R. HindIII sites of phage
lambda DNA may be effectively protected with actinomycin whereas some other
sites are protected with distamycin.  This allows us to obtain a set of larger
overlapping fragments of DNA (hereafter we shall term them as distamycin and
actinomycin protected fragments) and to establish the location of all HindIII
fragments on the physical map of phage lambda DNA.  The data reported here and
earlier lead to a conclusion that the antibiotics may be useful for DNA mapping
and for obtaining new vector molecules.

<>

<1>Braga, E.A., Nosikov, V.V., Tanyashin, V.I., Zhuze, A.L., Polyanovskii, O.L.
<2>Limitation of the action of restriction endonucleases by antibiotics bound to DNA.
<3>Dokl. Akad. Nauk.
<4>225
<5>707-710
<6>1975
<7>The effect of the antibiotics distamycin A and actinomycin D on the splitting
of DNA by restriction endonucleases was studied.  Electrophoresis showed that
antibiotics binding with DNA can compete with the corresponding enzymes.  Their
selectivity was determined by the nucleotide sequence of the recognition
section for the enzyme and also the immediate neighbourhood of these DNA
sections.  With EcoRI and phage lambda DNA it was shown that distamycin A
limited the number of sections split by restriction endonuclease.

<>

<1>Bragin, E.Y., Shtratnikova, V.Y., Dovbnya, D.V., Schelkunov, M.I., Pekov, Y.A., Malakho, S.G., Egorova, O.V., Ivashina, T.V., Sokolov, S.L., Ashapkin, V.V., Donova, M.V.
<2>Comparative analysis of genes encoding key steroid core oxidation enzymes in fast-growing Mycobacterium spp. strains.
<3>J. Steroid Biochem. Mol. Biol.
<4>138
<5>41-53
<6>2013
<7>A comparative genome analysis of Mycobacterium spp. VKM Ac-1815D, 1816D and 1817D
strains used for efficient production of key steroid intermediates
(androst-4-ene-3,17-dione, AD, androsta-1,4-diene-3,17-dione, ADD, 9alpha-hydroxy
androst-4-ene-3,17-dione, 9-OH-AD) from phytosterol has been carried out by deep
sequencing. The assembled contig sequences were analyzed for the presence
putative genes of steroid catabolism pathways. Since
3-ketosteroid-9alpha-hydroxylases (KSH) and 3-ketosteroid-Delta(1)-dehydrogenase
(Delta(1) KSTD) play key role in steroid core oxidation, special attention was
paid to the genes encoding these enzymes. At least three genes of Delta(1) KSTD
(kstD), five genes of KSH subunit A (kshA), and one gene of KSH subunit B of
3-ketosteroid-9alpha-hydroxylases (kshB) have been found in Mycobacterium sp. VKM
Ac-1817D. Strains of Mycobacterium spp. VKM Ac-1815D and 1816D were found to
possess at least one kstD, one kshB and two kshA genes. The assembled genome
sequence of Mycobacterium sp. VKM Ac-1817D differs from those of 1815D and 1816D
strains, whereas these last two are nearly identical, differing by 13 single
nucleotide substitutions (SNPs). One of these SNPs is located in the coding
region of a kstD gene and corresponds to an amino acid substitution Lys (135) in
1816D for Ser (135) in 1815D. The findings may be useful for targeted genetic
engineering of the biocatalysts for biotechnological application.

<>

<1>Brahmachari, S.K., Dash, D., Sharma, R., Maheshwari, J.K.
<2>A computer based versatile method for identifying protein coding DNA sequences useful as drug targets.
<3>International Patent Office
<4>WO 2005057464 A
<5>
<6>2005
<7>The present invention relates to a versatile method for identifying protein coding DNA
sequences (genes) useful as drug targets in a genome using specially developed software
GeneDecipher, said method comprising steps of generating peptide libraries from the known
genomes with peptide of length 'N' computationally arranged in an alphabetical order,
artificially translating the test genome to obtain a polypeptide corresponding to each reading
frame, converting each polypeptide sequence into an alphanumeric sequence one corresponding to
each reading frame on the basis of overlappings with the peptide libraries, training
Artificial Neural Network (ANN) with sigmoidal learning function to the alphanumeric sequence,
deciphering the protein coding regions in the test genome, thus, identifying longer stretches
of peptides mapping to large number of known genes and their corresponding proteins and
lastly, a method of the management of the diseases caused by the pathogenic organisms
comprising a step of evaluation of the proposed drug candidate by inhibiting the functioning
of one or more proteins identified by the steps of the invention.

<>

<1>Brahmachari, S.K., Shouche, Y.S., Cantor, C.R., McClelland, M.
<2>Sequences that adopt non-B-DNA conformation in form V DNA as probed by enzymic methylation.
<3>J. Mol. Biol.
<4>193
<5>201-211
<6>1987
<7>pBR322 form V DNA is a highly torsionally strained molecule with a linking
number of zero.  We have used sequence-specific DNA methylases as probes for
B-DNA in this molecule, exploiting the inability of methylases to methylate
single-stranded DNA and Z-DNA, both of which are known to occur in form V DNA.
Some sequences in form V DNA were shown to be totally in the B-form, others
were totally in an altered, unmethylatable conformation, while still other
sites appeared to exist partly in altered and partly in normal B-conformation.
Some potential Z-forming sequences (alternating pyrimidine/purine) of less than
seven base-pairs were not in the Z conformation in form V DNA, whereas others
did adopt an altered structure, indicating a modulating influence of flanking
sequences.  Furthermore, regions of imperfect alternating pyrimidine/purine
structure were sometimes capable of adopting an altered structure.  In
addition, some regions of altered structure had no apparent Z-forming
sequences, nor were they in polypurine stretches, which have also been proposed
to form left-handed DNA.  These non-B-DNA conformations may represent novel
left-handed helical structures or sequences that become single stranded under
torsional strain.  Long regions of either altered (unmethylatable) DNA or B-DNA
were not always observed.  In fact, one region showed three transitions between
B-like DNA and altered structure within 26 base-pairs.

<>

<1>Braid, M.D., Silhavy, J.L., Kitts, C.L., Cano, R.J., Howe, M.M.
<2>Complete Genomic Sequence of Bacteriophage B3, a Mu-Like Phage of Pseudomonas aeruginosa.
<3>J. Bacteriol.
<4>186
<5>6560-6574
<6>2004
<7>Bacteriophage B3 is a transposable phage of Pseudomonas aeruginosa. In
this report, we present the complete DNA sequence and annotation of the B3
genome. DNA sequence analysis revealed that the B3 genome is 38,439 bp
long with a G+C content of 63.3%. The genome contains 59 proposed open
reading frames (ORFs) organized into at least three operons. Of these
ORFs, the predicted proteins from 41 ORFs (68%) display significant
similarity to other phage or bacterial proteins. Many of the predicted B3
proteins are homologous to those encoded by the early genes and head genes
of Mu and Mu-like prophages found in sequenced bacterial genomes. Only two
of the predicted B3 tail proteins are homologous to other
well-characterized phage tail proteins; however, several Mu-like prophages
and transposable phage D3112 encode approximately 10 highly similar
proteins in their predicted tail gene regions. Comparison of the B3
genomic organization with that of Mu revealed evidence of multiple genetic
rearrangements, the most notable being the inversion of the proposed B3
immunity/early gene region, the loss of Mu-like tail genes, and an extreme
leftward shift of the B3 DNA modification gene cluster. These differences
illustrate and support the widely held view that tailed phages are genetic
mosaics arising by the exchange of functional modules within a diverse
genetic pool.

<>

<1>Brambilla, E. et al.
<2>Complete genome sequence of Methanoplanus petrolearius type strain (SEBR 4847).
<3>Standards in Genomic Sciences
<4>3
<5>203-211
<6>2010
<7>Methanoplanus petrolearius Ollivier et al. 1998 is the type strain of the genus Methanoplanus.
The strain was originally isolated from an offshore oil field from
the Gulf of Guinea. Members of the genus Methanoplanus are of interest because
they play an important role in the carbon cycle and also because of their
significant contribution to the global warming by methane emission in the
atmosphere. Like other archaea of the family Methanomicrobiales, the members of
the genus Methanoplanus are able to use CO(2) and H(2) as a source of carbon and
energy; acetate is required for growth and probably also serves as carbon source.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. This is the first complete genome sequence of a member
of the family Methanomicrobiaceae and the sixth complete genome sequence from the
order Methanomicrobiales. The 2,843,290 bp long genome with its 2,824
protein-coding and 57 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Brammar, W.J., Murray, N.E., Winton, S.
<2>Restriction of lambda trp bacteriophages by Escherichia coli K.
<3>J. Mol. Biol.
<4>90
<5>633-647
<6>1974
<7>trp-transducing derivatives of phage lambda have been used to study Escherichia
coli K specific restriction in vivo.  The expression of the trp genes from
unmodified phages during infection of a rec+, restricting host is eliminated by
restriction.  In a K-restricting recB,C host, where degradation of restricted
phage DNA is prevented, expression of the trp genes is little affected by the
presence of a single unmodified, K-restriction recognition site, even when that
site is within the trpE gene.  RI restriction, in contrast to K restriction,
prevents trp gene expression in a recB,C host when the restriction target is
between the trp genes and the relevant promoter.  The presence of two
K-restriction recognition sites in a lambda trp phage can have a marked effect
on trp gene expression.  This effect can be interpreted as the result of
preferential breakage between the two restriction recognition sites.  We
conclude that K restriction does not break susceptible DNA at, or even
preferentially near, a restriction recognition sequence.

<>

<1>Brancaccio, V.F., Zhurina, D.S., Riedel, C.U.
<2>Tough nuts to crack Site-directed mutagenesis of bifidobacteria remains a challenge.
<3>Bioengineered
<4>4
<5>197-202
<6>2013
<7>Most members of the genus Bifidobacterium are commensals of the human gastrointestinal tract
and some strains were shown to exert beneficial effects on their host. Based on these effects
and due to their status as GRAS (generally recognized as safe) microorganisms, specific
strains of bifidobacteria are marketed as probiotics. Despite their important role in food and
dairy industries, the mechanisms responsible for the probiotic effects of bifidobacteria are
mostly unknown. Over the last decade, the genomes of a large number of bifidobacteria have
been sequenced and analyzed. This has yielded a number of genes and their products that are
speculated to contribute to the probiotic effects of bifidobacteria. The gold standard to
demonstrate a role for specific genes is the analysis of mutants. At present, only a small
number of mutants of bifidobacteria have been generated by targeted mutagenesis. This is owed
to the genetic inaccessibility of most strains and a lack of appropriate molecular tools.
Successful generation of mutants of bifidobacteria was achieved by various methods including
classical suicide vector strategies, increase of transformation efficiencies by methylation of
plasmids and the use of temperature-sensitive vectors. In this commentary, we will describe
the methods successfully used for mutagenesis of bifidobacteria and discuss their advantages
and limitations.

<>

<1>Brank, A.S.
<2>Development and characterization of DNA methyltransferase inhibitors.
<3>Diss. Abstr.
<4>61
<5>5291-B
<6>2001
<7>Enzymatic methylation of carbon 5 of cytosine bases is an epigenetic modification that is
implicated in the silencing of tumor suppressor genes in various forms of cancer.  As a
potential therapeutic approach for the treatment of such tumors, the goal of this project was
to develop direct, mechanism-based inhibitors of mammalian DNA (cytosine-C5)-methyltransferase
(DNMT1), which is the enzyme responsible for the maintenance of established methylation
patterns in the genome.  The inhibition of the bacterial DNA (Cytosine-C5)-methyltransferase,
HhaI methyltransferase (M.HhaI), was initially examined because of the large volume of
structural information reported for this enzyme and because of the high degree of homology
between catalytic domains of M.HhaI and DNMT1.  The oligodeoxyribonucleotide (ODN) inhibitor
containing an abasic site in the target position (normally occupied by cytosine) was the most
potent inhibitor of M.HhaI in comparison to a variety of other ODN inhibitors containing other
modified target bases.  In addition to this potent inhibition, correlations were found between
binding of M.HhaI to the most potent ODN inhibitors and the ability of these ODNs to induce a
major M.HhaI conformational change.  Analysis of the catalytic efficiency of purified,
recombinant DNMTI (rD-NMT1) with a variety of substrates revealed the preference of rDNMT1 for
single-stranded substrates that formed stem-loop structures containing DNMT1 recognition
sites.  These ODNs were substituted with various potential inhibitory target bases and were
tested for the ability to inhibit rDNMT1 activity.  Of the inhibitors tested, ODNs containing
abasic target sites were again found to be the most potent inhibitors and were also found to
induce a dramatic conformational change of DNMT1.  In conclusion, the results show that ODNs
containing an abasic target site are potent inhibitors of DNMT1 activity, which may be
effective in vivo inhibitors of DNMT1 and potential therapeutic agents for treatment of
cancer.

<>

<1>Brank, A.S., Eritja, R., Garcia, R., Marquez, V., Christman, J.
<2>Inhibition of HhaI DNA (cytosine-C5) methyltransferase by oligodeoxyribonucleotides containing 5-Aza-2'-deoxycytidine: Examination of the intertwined roles of co-factor, target, transition state structure and enzyme conformation.
<3>J. Mol. Biol.
<4>323
<5>53-67
<6>2002
<7>The presence of 5-azacytosine (ZCyt) residues in DNA leads to potent inhibition of DNA
(cytosine-C5) methyltranferases (C5-MTases) in vivo and in vitro. Enzymatic methylation of
cytosine in mammalian DNA is an epigenetic modification that can alter gene activity and
chromosomal stability, influencing both differentiation and tumorigenesis. Thus, it is
important to understand the critical mechanistic determinants of ZCyt's inhibitory action.
Although several DNA C5-MTases have been reported to undergo essentially irreversible binding
to ZCyt in DNA, there is little agreement as to the role of AdoMet and/or methyl transfer in
stabilizing enzyme interactions with ZCyt. Our results demonstrate that formation of stable
complexes between HhaI methyltransferase (M.HhaI) and oligodeoxyribonucleotides containing
ZCyt at the target position for methylation (ZCyt-ODNs) occurs in both the absence and
presence of co-factors, AdoMet and AdoHcy. Both binary and ternary complexes survive SDS-PAGE
under reducing conditions and take on a compact conformation that increases their
electrophoretic mobility in comparison to free M.HhaI. Since methyl transfer can occur only in
the presence of AdoMet, these results suggest (1) that the inhibitory capacity of ZCyt in DNA
is based on its ability to induce a stable, tightly closed conformation of M.HhaI that
prevents DNA and co-factor release and (2) that methylation of ZCyt in DNA is not required for
inhibition of M.HhaI.

<>

<1>Brank, A.S., Van Bemmel, D.M., Christman, J.K.
<2>Optimization of baculovirus-mediated expression and purification of  hexahistidine-tagged murine DNA (cytosine-C5)-methyltransferase-1 in   Spodoptera frugiperda 9 cells.
<3>Protein Expr. Purif.
<4>25
<5>31-40
<6>2002
<7>Enzymatic DNA methylation of carbon 5 of cytosines is an epigenetic  modification that plays a
role in regulating gene expression,
differentiation, and tumorigenesis. DNA (cytosine-C5)-methyltransferase-1
is the enzyme responsible for maintaining established methylation patterns
during replication in mammalian cells. It is composed of a large (approximately 1100 amino
acids (a.a.)) amino-terminal region containing many putative
regulatory domains and a smaller (approximately 500 a.a.) carboxy-terminal
region containing conserved, catalytic domains. In this study, murine DNA
(cytosine C5)-methyltransferase-1, fused to an amino-terminal
hexahistidine tag, was expressed by infecting Spodoptera frugiperda cells
for 46 h with a recombinant baculovirus carrying the DNA
(cytosine-C5)-methyltransferase-1 cDNA. A total of 3 X 10^8 infected S.
frugiperda cells yielded approximately 1 mg of full-length,
hexahistidine-tagged DNA (cytosine-C5)-methyltransferase-1, which was
purified approximately 450-fold from RNase-treated S. frugiperda cell extracts
by nickel affinity chromatography. The characterization of
hexahistidine-tagged DNA (cytosine-C5)-methyltransferase-1 through DNA
methylation and inhibitor-binding assays indicated that the purified
enzyme had at least a 30-fold higher catalytic efficiency with
hemimethylated double-stranded oligodeoxyribonucleotide substrates than
unmethylated substrates and was most active with small
oligodeoxyribonucleotide substrates with a capacity for forming stem-loop
structures. The expression and purification procedures reported here
differ significantly from the original reports of baculovirus-mediated
hexahistidine-tagged DNA (cytosine-C5)-methyltransferase-1 expression and
purification by nickel affinity chromatography and provide a consistent
yield of active enzyme.

<>

<1>Brankatschk, K., Blom, J., Goesmann, A., Smits, T.H., Duffy, B.
<2>Genome of a European Fresh-Vegetable Food Safety Outbreak Strain of Salmonella enterica subsp. enterica Serovar Weltevreden.
<3>J. Bacteriol.
<4>193
<5>2066
<6>2011
<7>The genome of Salmonella enterica subsp. enterica serovar Weltevreden strain 2007-60-3289-1
was sequenced. The genome sequence of this
fresh-vegetable isolate from Scandinavia will be useful for the
elucidation of plant host factors in comparison to other serovars of S.
enterica subsp. enterica.

<>

<1>Brassard, S., Paquet, H., Roy, P.H.
<2>A transposon-like sequence adjacent to the AccI restriction-modification operon.
<3>Gene
<4>157
<5>69-72
<6>1995
<7>We have cloned and sequenced the accIRM genes from Weeksella zoohelcum (the original
identification of this strain as Acinetobacter calcoaceticus was incorrect).  Our sequence
differs in the coding regions from a previously published sequence by the addition of three
nucleotides near the 3' end of the DNA methyltransferase-encoding gene (accIM).  We have
sequenced approx. 3 kb beyond this operon.  Two genes were found, convergently transcribed
with the R-M operon.  The first of these genes encodes a protein which shows significant
similarity to the recombinases of the phage integrase family.  The W. zoohelcum recombinase
may function as a transposon resolvase, as in Tn4430.  The recombinase-encoding gene is
followed by a putative transposase (Tnp), which is in turn followed by a terminator which is
predicted to be Rho-dependent for the recombinase-Tnp operon and Rho-independent for the
convergent R-M operon.  Since the G+C content of the two operons is notably different, it is
possible that the terminator is at the extremity of the mobile element and serves to protect
it from incoming transcription.

<>

<1>Brathwaite, K.J., Siringan, P., Moreton, J., Wilson, R., Connerton, I.F.
<2>Complete Genome Sequence of Universal Bacteriophage Host Strain Campylobacter jejuni subsp. jejuni PT14.
<3>Genome Announcements
<4>1
<5>e00969-13
<6>2013
<7>Campylobacter jejuni strain PT14 is a clinical isolate previously used to propagate
bacteriophages in the United Kingdom phage typing scheme. The strain
has proven useful in the isolation of Campylobacter bacteriophages from several
sources, and it functions as a model host in phage therapy experiments with
poultry and poultry meat.

<>

<1>Braun, J.
<2>Structural study of the inhibition of DNA (cytosine-5) methyltransferases by original non-nucleoside inhibitors.
<3>Chim. Nouv.
<4>28
<5>20-22
<6>2010
<7>
<>

<1>Brawer, R., Batista, F.D., Burrone, O., Sordelli, D.O., Cerquetti, M.C.
<2>The dam methyltransferase of a Salmonella typhimurium insertion mutant.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>96
<5>500
<6>1996
<7>A Salmonella typhimurium temperature-sensitive (ts) mutant was obtained after
transposition mutagenesis with a miniTn10dTc element.  This mutant replicated well at 28oC but
showed impaired growth at 37oC on LB agar.  Microscopic analysis revealed that the mutant
produced filaments when cultured at 37oC.  Sequences flanking the insertion site were cloned
by a
"jumping PCR" technique.  Sequencing data demonstrated that the miniTn10d Tc was inserted at
position 804 of the dam gene.  The Dam protein, coded by dam, methylates adenines in 5'-
GATC-3' sequences.  The miniTn10d Tc insertion created a stop codon truncating the
translation
of the Dam last 10 amino acids.  The insertion site GGCTCTGTA was duplicated due to the
transposition.  DNA digestion with MboI, DpnI and Sau3AI showed that the mutant genomic DNA
contained methylated and unmethylated GATC sites.  The S. typhimurium wt dam gene showed
82% and 92% homology in their nucleotide and deduced amino acid sequences, respectively, with
the E. coli dam gene.  Mutant transformation with pTP166, which overproduces E. coli Dam,
complemented the mutant phenotype.  We also constructed a recombinant plasmid (pDeldam)
which mimics the deletion produced by the miniTn10d Tc insertion.  The pDeldam was used to
transform an E. coli dam mutant.  The transformants showed a ts phenotype identical to that of
the
S. typhimurium insertion mutant.  We conclude that deletion of the last 10 amino acids from
protein Dam diminished the methylase activity and conferred a ts phenotype.

<>

<1>Brawer, R., Batista, F.D., Burrone, O.R., Sordelli, D.O., Cerquetti, M.C.
<2>A temperature-sensitive DNA adenine methyltransferase mutant of Salmonella typhimurium.
<3>Arch. Microbiol.
<4>169
<5>530-533
<6>1998
<7>A temperature-sensitive mutant of Salmonella typhimurium was isolated earlier after transposon
mutagenesis with Tn10dTet.  The mutant D220 grows well at 28 C but has a lower growth rate and
forms filaments at 37 C.  Transposon-flanking fragments of mutant D220 DNA were cloned and
sequenced.  The transposon was inserted in the dam gene between positions 803 and 804
(assigned allele number: dam-231::Tn10dTet) and resulted in a predicted ten-amino-acid-shorter
Dam protein.  The insertion created a stop codon that led to a truncated Dam protein with a
temperature-sensitive phenotype.  The insertion dam-231::Tn10dTet resulted in a dam "leaky"
phenotype since methylated and unmethylated adenines in GATC sequences were present.  In
addition, the dam-231::Tn10dTet insertion rendered dam mutants temperature-sensitive for
growth depending upon the genetic background of the S. typhimurium strain.  The wild-type dam
gene of S. typhimurium exhibited 82% identity with the Escherichia coli dam gene.

<>

<1>Breakwell, D.P., Barrus, E.Z., Benedict, A.B., Brighton, A.K., Fisher, J.N., Gardner, A.V., Kartchner, B.J., Ladle, K.C., Lunt, B.L., Merrill, B.D., Morrell, J.D., Burnett, S.H., Grose, J.H.
<2>Genome Sequences of Five B1 Subcluster Mycobacteriophages.
<3>Genome Announcements
<4>1
<5>e00968-13
<6>2013
<7>Mycobacteriophages infect members of the Mycobacterium genus in the phylum Actinobacteria and
exhibit remarkable diversity. Genome analysis groups the
thousands of known mycobacteriophages into clusters, of which the B1 subcluster
is currently the third most populous. We report the complete genome sequences of
five additional members of the B1 subcluster.

<>

<1>Brede, D.A., Snipen, L.G., Ussery, D.W., Nederbragt, A.J., Nes, I.F.
<2>Complete genome sequence of the commensal Enterococcus faecalis 62 isolated from a healthy Norwegian infant.
<3>J. Bacteriol.
<4>193
<5>2377-2378
<6>2011
<7>The genome of Enterococcus faecalis 62, a commensal isolate from a healthy Norwegian infant,
revealed multiple adaptive traits to the gastro intestinal tract (GIT) environment and the
milk containing diet of a breast fed infant. Adaptation to a commensal existence was
emphasized by lactose and other carbohydrate metabolism genes within genomic islands
accompanied by the absence of virulence traits.

<>

<1>Breider, S. et al.
<2>Genome sequence and emended description of Leisingera nanhaiensis strain DSM 24252(T) isolated from marine sediment.
<3>Standards in Genomic Sciences
<4>9
<5>687-703
<6>2014
<7>Leisingera nanhaiensis DSM 24252(T) is a Gram-negative, motile, rod-shaped marine
Alphaproteobacterium, isolated from sandy marine sediments. Here we present the
non-contiguous genome sequence and annotation together with a summary of the
organism's phenotypic features. The 4,948,550 bp long genome with its 4,832
protein-coding and 64 RNA genes consists of one chromosome and six
extrachromosomal elements with lengths of 236 kb, 92 kb, 61 kb, 58 kb, 56 kb, and
35 kb, respectively. The analysis of the genome showed that DSM 24252(T)
possesses all genes necessary for dissimilatory nitrite reduction, and the strain
was shown to be facultatively anaerobic, a deviation from the original
description that calls for an emendation of the species. Also present in the
genome are genes coding for a putative prophage, for gene-transfer agents and for
the utilization of methylated amines. Phylogenetic analysis and intergenomic
distances indicate that L. nanhaiensis might not belong to the genus Leisingera.

<>

<1>Breitbart, M., Haynes, M., Kelley, S., Angly, F., Edwards, R.A., Felts, B., Mahaffy, J.M., Mueller, J., Nulton, J., Rayhawk, S., Rodriguez-Brito, B., Salamon, P., Rohwer, F.
<2>Viral diversity and dynamics in an infant gut.
<3>Res. Microbiol.
<4>159
<5>367-373
<6>2008
<7>Metagenomic sequencing of DNA viruses from the feces of a healthy week-old
infant revealed a viral community with extremely low diversity. The
identifiable sequences were dominated by phages, which likely influence
the diversity and abundance of co-occurring microbes. The most abundant
fecal viral sequences did not originate from breast milk or formula,
suggesting a non-dietary initial source of viruses. Certain sequences were
stable in the infant's gut over the first 3 months of life, but microarray
experiments demonstrated that the overall viral community composition
changed dramatically between 1 and 2 weeks of age.

<>

<1>Breitbart, M., Hewson, I., Felts, B., Mahaffy, J.M., Nulton, J., Salamon, P., Rohwer, F.
<2>Metagenomic analyses of an uncultured viral community from human feces.
<3>J. Bacteriol.
<4>185
<5>6220-6223
<6>2003
<7>Here we present the first metagenomic analyses of an uncultured viral community from human
feces, using partial shotgun sequencing. Most of the
sequences were unrelated to anything previously reported. The recognizable
viruses were mostly siphophages, and the community contained an estimated
1,200 viral genotypes.

<>

<1>Breitbart, M., Salamon, P., Andresen, B., Mahaffy, J.M., Segall, A.M., Mead, D., Azam, F., Rohwer, F.
<2>Genomic analysis of uncultured marine viral communities.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>14250-14255
<6>2002
<7>Viruses are the most common biological entities in the oceans by an order of magnitude.
However, very little is known about their diversity. Here we report a genomic analysis of two
uncultured marine viral communities. Over 65% of the sequences were not significantly similar
to previously reported sequences, suggesting that much of the diversity is previously
uncharacterized. The most common significant hits among the known sequences were to viruses.
The viral hits included sequences from all of the major families of dsDNA tailed phages, as
well as some algal viruses. Several independent mathematical models based on the observed
number of contigs predicted that the most abundant viral genome comprised 2-3% of the total
population in both communities, which was estimated to contain between 374 and 7,114 viral
types. Overall, diversity of the viral communities was extremely high. The results also showed
that it would be possible to sequence the entire genome of an uncultured marine viral
community.

<>

<1>Bremer, M.C.D., Gimble, F.S., Thorner, J., Smith, C.L.
<2>VDE endonuclease cleaves Saccharomyces cerevisiae genomic DNA at a single site: physical mapping of the VMA1 gene.
<3>Nucleic Acids Res.
<4>20
<5>5484
<6>1992
<7>A DNA endonuclease, VDE, is derived from the VMA1 gene product of the yeast Saccharomyces
cerevisiae and is related to other nucleases involved in nucleic acid rearrangements. Analysis
of two cleavages sites showed that VDE recognizes an extended sequence,
5'=TATSYATGYYGGGTGY^GGRG-AARKMGKKAAWGAAAWG-3', and leaves a staggered double-strand break
with 4-bp 3'-hydroxyl overhangs. Cleavage of one site in the VMA1(delta)vde allele
(previously deleted for the segment encoding VDE) during meiosis of a heterozygous diploid
initiates a gene conversion event that transforms the mutant allele into a full-length VMA1
gene. Here VDE cleavage of this same site in vitro was used to physically map the VMA1 locus.

<>

<1>Brendler, T., Austin, S.
<2>Binding of SeqA protein to DNA requires interaction between two or more complexes bound to separate hemimethylated GATC sequences.
<3>EMBO J.
<4>18
<5>2304-2310
<6>1999
<7>The SeqA protein binds to the post-replicative forms of the origins of replication of the
Escherichia coli chromosome (oriC) and the P1 plasmid (P1oriR) at hemimethylated GATC adenine
methylation sites.  It appears to regulate replication by preventing premature reinitiation.
However, SeqA binding is not exclusive to replication origins: different fragments with
hemimethylated GATC sites can bind SeqA in vitro when certain rules apply.  Most notably, more
than one such site must be present on a bound fragment.  The protein appears to recognize
individual hemimethylated sites, but must undergo an obligate cooperative interaction with a
nearby found protein for stable binding.  SeqA contacts both DNA strands in a discrete patch
at each hemimethylated GATC sequence.  All four GATC bases are contacted and are essential for
binding.  Although the recognized sequence is symmetrical, the footprint on the methylated
strand is always broader, suggesting that the bound protein is positioned asymmetrically with
its orientation dictated by the position of the unique methyl group.  Studies of alternative
spacings and relative orientations of adjacent sites suggest that each site may be recognized
by a symmetrical dimer with an induced asymmetry in one of the subunits similar to that seen
with certain type II restriction endonucleases.

<>

<1>Brennan, C.A., Gumport, R.I.
<2>T4 RNA ligase catalyzed synthesis of base analogue-containing oligodeoxyribonucleotides and a characterization of their thermal stabilities.
<3>Nucleic Acids Res.
<4>13
<5>8665-8684
<6>1985
<7>Self-complementary oligodeoxyribonucleotides containing the base analogues
2-aminopurine, 2,6-diaminopurine, N6-methyladenine, uracil, and 5-bromouracil
were synthesized by a general method that allows incorporation of the analogues
at specific positions.  The method uses chemically synthesized partial
sequences but circumvents the need for protected base analogues by
incorporating their unprotected 3',5'-bisphosphate derivatives enzymatically.
T4 RNA ligase was used to add the analogues to the oligodeoxyribonucleotides
with yields from 54 to greater than 95 percent.  Oligodeoxyribonucleotides were
joined to the oligodeoxyribonucleotides containing the analogues at their
3'-termini in yields from 22 to 81 percent.  The high yields obtained in these
joinings suggest that RNA ligase should be of general use for the specific
incorporation of other deoxyribonucleoside analogues into
oligodeoxyribonucleotides.  The oligodeoxyribonucleotides containing the base
analogues were characterized by their mobilities during HPLC, nucleoside
compositions, sequences, and thermal stabilities.

<>

<1>Brennan, C.A., Van Cleve, M.D., Gumport, R.I.
<2>The effects of base analogue substitutions on the cleavage by the EcoRI restriction endonuclease of octadeoxyribonucleotides containing modified EcoRI recognition sequences.
<3>J. Biol. Chem.
<4>261
<5>7270-7278
<6>1986
<7>It has been proposed that recognition of specific DNA sequences by proteins is
accomplished by hydrogen bond formation between the protein and particular
groups that are accessible in the major and minor grooves of the DNA.  We have
examined the DNA-protein interactions involved in the recognition of the
hexameric DNA sequence, GAATTC, by the EcoRI restriction endonuclease by using
derivatives of an oligodeoxyribonucleotide that contain a variety of base
analogues.  The base analogues hypoxanthine, 2-aminopurine, 2,6-diaminopurine,
N6-methyladenine, 5-bromouracil, uracil, 5-bromocytosine, and 5-methylcytosine
were incorporated as single substitutions into the octadeoxyribonucleotide
d(pG-G-A-A-T-T-C-C).  The effects of the substitutions on the interactions
between the EcoRI endonuclease and its recognition sequence were monitored by
determining the steady state kinetic values of the hydrolysis reaction.  The
substitutions resulted in effects that varied from complete inactivity to
enhanced reactivity.  The enzyme exhibited Michaelis-Menten kinetics with those
substrates that were reactive, whereas octanucleotide analoguues containing
N6-methyladenine at either adenine position, uracil at the second thymine
position, of 5-bromocytosine or 5-methylcytosine at the cytosine position were
unreactive.  The results are discussed in terms of possible effects on the
interactions between the enzyme and its recognition site during the reaction.
An accompanying paper presents the results of a similar study using these
oligonucleotides with the EcoRI modification methylase.

<>

<1>Brennan, C.A., Van Cleve, M.D., Gumport, R.I.
<2>The effects of base analogue substitutions on the methylation by the EcoRI modification methylase of octadeoxyribonucleotides containing modified EcoRI recognition sequences.
<3>J. Biol. Chem.
<4>261
<5>7279-7286
<6>1986
<7>We have examined the DNA-protein interactions involved in the recognition of a
specific hexameric sequence, GAATTC, by the EcoRI modification methylase by
using derivatives of an oligodeoxyribonucleotide that contain a variety of base
analogues.  The base analogues 2-aminopurine, 5-bromocytosine, 5-bromouracil,
2,6-diaminopurine, hypoxanthine, 5-methylcytosine, N6-methyladenine, and uracil
were incorporated as single substitutions into the octadeoxyribonucleotide
d(pG-G-A-A-T-T-C-C).  The effects of the substitutions on the ability of the
enzyme to methylate the modified substrates were monitored by determining the
steady state kinetic values of the reaction in the presence of the cosubstrate,
S-adenosylmethionine.  The substitutions resulted in effects ranging from
complete inactivity to enhanced reactivity.  The enzyme exhibited
Michaelis-Menten kinetics with those analogues that were active, whereas the
octanucleotides containing hypoxanthine at the guanine site, N6-methyladenine
at the first or 2-aminopurine at the second adenine site, or uracil at the
second thymine site were completely inactive.  The results are discussed in
terms of the possible interactions between the methylase and its recognition
sequence.  In addition, the interactions are compared to those of the EcoRI
restriction endonuclease, which has been similarly tested with the same
analogue oligonucleotides.  The results of that study are reported in an
accompanying paper.  Although both enzymes recognize the same sequence, they do
so in different ways.

<>

<1>Brenneman, M., Gimble, F.S., Wilson, J.H.
<2>Stimulation of intrachromosomal homologous recombination in human cells by electroporation with site-specific endonucleases.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>3608-3612
<6>1996
<7>In somatic mammalian cells, homologous recombination is a rare event.  To study
the effects of chromosomal breaks on frequency of homologous recombination, site-specific
endonucleases were introduced into human cells by electroporation.  Cell lines with a partial
duplication within the HPRT (hypoxanthine phosphoribosyltransferase) gene were created through
gene targeting.  Homologous, intrachromosomal recombination between the repeated regions of
the gene can reconstruct a functioning, wild-type gene.  Treatment of thse cells with the
restriction
endonuclease XbaI, which has a recognition site within the repeated region of HPRT homology,
increased the frequency of homologous recombination by more than 10-fold.  Recombination
frequency was similarly increased by treatment with the rare-cutting yeast endonuclease
PI-SceI
when a cleavage site was placed within the repeated region of HPRT.  In contrast, four
restriction
enzymes that cut at positions either outside of the repeated regions or between them produced
no
change in recombination frequency.  The results suggest that homologous recombination between
intrachromosomal repeats can be specificially initiated by a double-strand break occurring
within
regions of homology, consistent with the predictions of a double-strand-break-repair model.

<>

<1>Brenner, C., Fuks, F.
<2>DNA methyltransferases: Facts, clues, mysteries.
<3>Curr. Top. Microbiol. Immunol.
<4>301
<5>45-66
<6>2006
<7>DNA methylation plays a pivotal role during development in mammals and is central to
transcriptional silencing. The DNA methyltransferases
(DNMTs) are responsible for the generation of genomic methylation
patterns leading to gene silencing, but the underlying molecular basis
remains largely shrouded in mystery. Here we review our current
understanding of the mechanisms by which DNMTs repress transcription
and how they are targeted to preferred DNA sequences. Emerging evidence
points to an essential and intricate web of interactions between DNMTs
and the chromatin environment in which they function. The recent
identification of novel transcription factors recruiting the DNMTs may
open new avenues of research into the origin of DNA methylation
patterns. Thanks to these emerging clues, researchers have begun to
lift the veil on the multi-faceted DNMTs, but there remains fascinating
work ahead for whoever wants to fully understand DNMTs and their role
in the mammalian cell.

<>

<1>Brenner, E.V., Brouchkov, A.V., Kurilshikov, A.M., Griva, G.I., Kashuba, E., Kashuba, V.I., Melefors, O., Repin, V.E., Melnikov, V.P., Vlassov, V.V.
<2>Draft Genome Sequence of Bacillus cereus Strain F, Isolated from Ancient Permafrost.
<3>Genome Announcements
<4>1
<5>e00561-13
<6>2013
<7>Bacillus cereus strain F was isolated and cultured from a sample of permafrost, aged
presumably about 3 million years, on the Mammoth Mountain (62 degrees 56'N,
133 degrees 59'E). These genome data provide the basis to investigate Bacillus
cereus F, identified as a long-term survivor of the extremely cold and close
environment.

<>

<1>Brenner, V., Focht, D.D.
<2>Cloning of the Type II methyltransferase from chlorobenzoate degrader Pseudomonas putida P111.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>459
<6>1994
<7>Pseudomonas putida P111 is able to utilize a broad range of mono-, di-, and trichlorinated
benzoates for growth. Phenotypical varient of this strain, designated P111B, was isolated to
enable the cloning of the gene complex specifying degradation of ortho-chlorinated benzoates
which was found on the indigenous plasmid pPB111. However, the cloning of this gene(s) was
hampered by the presence of the Type II restriction and modification (R-M) system, designed
PpuI, detected on the same plasmid. The R-M system has the same specificity as the R-M system
EcoRI, e.g. the restriction endonuclease and the methylase both recognize the sequence
G/AATTC. The gene for the methyltransferase PpuI was cloned into a broad host range cosmid
pLAFR3, in selection step including the transformation of DNA isolated from all recombinant
clones, and predigested thoroughly with EcoRI restriction endonuclease. Thus, only the DNA
from recombinants protected by the methyltransferase PpuI was not linearized and was able to
transform the recipient strain E. coli HB101. The region containing the gene for the
methyltransferase PpuI was further subcloned and reintroduced into the cosmid pLAFR3. The
isoschizomer enzymes recognizing the sequence G/AATTC were found in two reports on conjugative
plasmids (EcoRI, RrsII), therefore the comparison of their sequences may be an important
evolutionary development.

<>

<1>Brenner, V., Kiss, A., Venetianer, P.
<2>A method for construction of a new plasmid vector and use of this plasmid vector for cloning and expression of the EcaI methylase gene.
<3>Hungarian Patent Office
<4>HU T58808
<5>
<6>1992
<7>Plasmid vectors with EcaI detection sites capable of cloning genes of EcaI-DNA-methylase are
prepared.  A novel cloning process leads to genes for modified methylase-enzyme (M.EcaI)
neither described, nor isolated previously, and gives rise to a gp. producing EcaI-methylase
on large scale.  The insertion of a DNA sequence, containing four EcaI detection sites
(GGTNACC) into a known, vector renders it suitable for the cloning task.  DNA is isolated
subsequently from Enterobacter-cloacae and is partially digested.  The preceding vector and
the partially digested DNA are ligated.  Escherichia coli cells are transformed then with the
previous recombinant DNA and recombinant plasmids are separated  from transformed cells.
Exhaustive digestion of previously isolated DNA follows, with EcaI restriction endonuclease.
The determining feature of this method is the destruction of the overwhelming majority of
recombinant plasmids, leaving only those which contained active (intact) genes capable of
coding M.EcaI enzymes.  Repeated transformations of the Escherichia coli host follows with
DNA, exhaustively digested previously.  The structure of cloned genes is established by
characterizing transforming clones.  Coding sequences are isolated from plasmids, having
obtained and characterizing them earlier and are transplanted into vector plasmids being
constructed by them and ensuring high levels of gene development.  Thus, good yields of EcaI
methylase enzyme are obtained.

<>

<1>Brenner, V., Venetianer, P., Kiss, A.
<2>Cloning and nucleotide sequence of the gene encoding the EcaI DNA methyltransferase.
<3>Nucleic Acids Res.
<4>18
<5>355-359
<6>1990
<7>The gene coding for the GGTNACC specific EcaI DNA methyltransferase (M.EcaI)
has been cloned in E. coli from Enterobacter cloacae and its nucleotide
sequence has been determined.  The ecaIM gene codes for a protein of 452 amino
acids (Mr:51,111).  It was determined that M.EcaI is an adenine
methyltransferase.  M.EcaI shows limited amino acid sequence similarity to
other adenine methyltransferases.  A clone that expresses EcaI
methyltransferase at high level was constructed.

<>

<1>Brenot, A., Werts, C., Ottone, C., Sertour, N., Charon, N.W., Postic, D., Baranton, G., Saint, G.I.
<2>First evidence for a restriction-modification system in Leptospira sp.
<3>FEMS Microbiol. Lett.
<4>201
<5>139-143
<6>2001
<7>The LE1 leptophage exhibited a host range restricted to the saprophytic Leptospira hiflexa
[Saint Girons et al., Res. Microbiol. 141 (1990)
1131-1133] and mainly to the Patoc 1 strain (hereafter called PFRA)
kept in the Paris, France collection. Results of titration of LEI
lysates indicated the presence of a host-controlled modification and
restriction system within PUSA (Patoc 1 strain maintained in the
Morgantown, WV, USA collection) that was absent in PFRA. Because
genomic DNA of PITAL (Patoc 1 strain maintained in Trieste, Italy)
appeared smeared in pulsed field gel electrophoresis (PFGE), this
strain is likely to contain nucleases that are activated upon DNA
isolation. Moreover, comparative Nod digestions of PUSA and PFRA DNAs,
as visualized by PFGE, indicated that PUSA belonged to a different
serovar than PFRA. Finally, 16S ribosomal sequence analysis indicated
that PUSA belonged to the saprophytic Leptospira meyeri species, while
PITAL and PFRA appertained to L. biflexa. The evolutionary significance
and the importance of the restriction and modification enzymes or
non-specific nucleases within strains for genetic experiments are
discussed.

<>

<1>Brensing-Kuppers, J., Reischl, U., Schmitz, G.S., Kaluza, K., Jarsch, M., Kessler, C.
<2>McrI:  a novel class-II restriction endonuclease from Micrococcus cryophilus recognizing 5'-CGRY/CG-3'.
<3>FEBS Lett.
<4>264
<5>218-222
<6>1990
<7>A new class-II restriction endonuclease, McrI, with a novel sequence
specificity as isolated from the Gram-positive eubacterium Micrococcus
cryophilus.  McrI recognizes the palindromic hexanucleotide sequence
5'-CGRY^CG-3'
3'-GC^YRGC-5'.  The novel enzyme in the presence of Mg2+-ions cleaves
specifically both strands as indicated by the arrows.  The staggered cuts
generate 3'-protruding ends with single-stranded 5'-RY-3' dinucleotide
extensions.  The McrI recognition sequence was deduced from mapping data on
DNAs of bacteriophages PhiX174 RF and M13mp18 RF characterized by one and four
cleavage sites, respectively.  The cut positions within both strands of the
recognition sequence were determined in sequencing experiments by analyzing
hydrolysis of phosphodiester bonds within a polylinker region of M13mp18 RF DNA
containing an additional McrI recognition site including treatment with T4 DNA
polymerase.  The novel enzyme may be a useful tool for cloning experiments by
completion of the enzymes EclXI (5'-C/GGCCG-3'), NotI (5'-GC/GGCCGC-3'), PvuI
(5'-CGAT/CG-3') as well as EaeI (5'-Y/GGCCR-3') and XhoII (5'-Y/GATCR-3')
characterized by partly identical sequence specificities.

<>

<1>Breton, G., Bush, D.
<2>Nucleic acid and amino acid sequences relating to Acinetobacter baumannii for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6562958 A
<5>
<6>2003
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Acinetobacter baumannii that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Breton, G.L.
<2>Nucleic acid and amino acid sequences relating to Proteus mirabilis for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6605709 B
<5>
<6>2003
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from Proteus
fragilis that are useful in diagnosis and therapy of pathological conditions; antibodies
against the polypeptides; and methods for the production of the polypeptides.  The invention
also provides methods for the detection, prevention and treatment of pathological conditions
resulting from bacterial infection.

<>

<1>Breton, G.L.
<2>Nucleic acid and amino acid sequences relating to M. catarrhalis for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6673910 B
<5>
<6>2004
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from Moraxella
catarrhalis that are useful in diagnosis and therapy of pathological conditions; antibodies
against the polypeptides; and methods for the production of the polypeptides.  The invention
also provides methods for the detection, prevention and treatment of pathological conditions
resulting from bacterial infection.

<>

<1>Breton, G.L.
<2>Nucleic acid sequences relating to Bacteroides fragilis for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7090973 B
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Bacteroides fragilis that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Breton, G.L., Osborne, M.
<2>Nucleic acid amino sequences relating to Klebsiella pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6610836 B
<5>
<6>2003
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from Klebsiella
pneumoniae that are useful in diagnosis and therapy of pathological conditions; antibodies
against the polypeptides; and methods for the production of the polypeptides.  The invention
also provides methods for the detection, prevention and treatment of pathological conditions
resulting from bacterial infection.

<>

<1>Brevnov, M.G., Gritsenko, O.M., Mikhailov, S.N., Efimtseva, E.V., Ermolinsky, B.S., Van Aeroschot, A., Herdewijn, P., Repyk, A.V., Gromova, E.S.
<2>DNA duplexes with reactive dialdehyde groups as novel reagents for cross-linking to restriction-modification enzymes.
<3>Nucleic Acids Res.
<4>25
<5>3302-3309
<6>1997
<7>To create new, effective reagents for affinity modification of restriction-modification
enzymes, a regio-selective method for reactive dialdehyde group incorporation into
oligonucleotides, based on insertion of a 1-beta-D-galactopyranosylthymine residue, has been
developed.  We synthesized DNA duplex analogs of the substrates of the EcoRII and MvaI R-M
enzymes that contained a galactose or periodate-oxidized galactose residue as single
substituents either in the center of the EcoRII (MvaI) recognition site or in the flanking
nucleotide sequence.  The dependence of binding, cleavage and methylation of these substrate
analogs on the modified sugar location in the duplex was determined.  Cross-linking of the
reagents to the enzymes under different conditions was examined.  M.EcoRII covalent attachment
to periodate-oxidized substrate analogs proceeded in a specific way and to a large extent
depended on the location of the reactive dialdehyde group in the substrate.  The yield of
covalent attachment to a DNA duplex with a dialdehyde group in the flanking sequence with
EcoRII or MvaI methylases was 9-20% and did not exceed 4% for R.EcoRII.

<>

<1>Brevnov, M.G., Kubareva, E.A., Romanova, E.A., Volkov, E.M., Karyagina, A.S., Nikolskaya, I.I., Gromova, E.S.
<2>Interaction of the MvaI and SsoII methyltransferases with DNAs altered at the central base pair of the recognition sequence.
<3>Gene
<4>157
<5>149-152
<6>1995
<7>The interaction of the MvaI and SsoII DNA methyltransferases (MTases; M.MvaI and M.SsoII,
respectively) with a set of synthetic DNA duplexes, containing an M.MvaI and an M.SsoII
recognition site (CCWGG), was investigated.  In these DNA duplexes dA or dT of the recognition
site was replaced by nucleoside analogs with modified sugar moieties and heterocyclic bases
(2'-deoxy-2'-fluorouridine (flU), 1-(b-D-2'-deoxy-threo-pentofuranosyl) thymine (xT),
1-(b-D-3'-deoxy-threo-pentofuranosyl)uracil (tU)), or by 1,3-propanediol (Prd).  A new
approach for monitoring methylation of each strand of DNA duplexes by MTases was developed.
It allowed the determination of the influence of the modification in one DNA strand on the
methylation of the other.  In most cases, for both M.MvaI and M.SsoII, sugar analog-containing
duplexes showed inhibition of methylation of only the modified strand.  Prd-containing DNA
duplexes were not substrates for M.MvaI.  M.SsoII did not methylate DNA duplexes in which the
dT residue was replaced by Prd.

<>

<1>Brevnov, M.G., Kubareva, E.A., Volkov, E.M., Romanova, E.A., Oretskaya, T.S., Gromova, E.S., Shabarova, Z.A.
<2>Influence of a single modification of the DNA sugar-phosphate backbone on the functioning of restriction endonucleases EcoRII, MvaI, and BstNI.
<3>Mol. Biol. (Mosk)
<4>29
<5>1294-1300
<6>1995
<7>Oligonucleotide duplexes containing modified nucleoside residues
1-(2'-deoxy-betaD-threo-pentofuranosyl)thymine (xT) or
1-(3'-deoxy-betaD-threo-pentofuranosyl)uracil (tU) in the center of the recognition site for
EcoRII, MvaI, and BstNI (CCWGG) were constructed and examined in respect of their
physicochemical and substrate properties.  The restriction endonucleases proved to differ in
their sensitivity to the resulting conformational distortions.  For the first time with
EcoRII, the xT-containing analog turned out to be a much better substrate and, moreover, an
activator of the enzyme.

<>

<1>Brezellec, P., Hoebeke, M., Hiet, M.S., Pasek, S., Ferat, J.L.
<2>DomainSieve: a protein domain-based screen that led to the identification of dam-associated genes with potential link to DNA maintenance.
<3>Bioinformatics
<4>22
<5>1935-1941
<6>2006
<7>Motivation: The Dam methyltransferase (DamMT) activity, broadly distributed in association
with restriction endonucleases, as part of
the restriction-modification defense systems, has evolved to become
intimately associated with essential biological functions in a few
organisms. In Escherichia coli, DamMT is involved in multiple aspects
of DNA maintenance, replication initiation, daughter chromosome
segregation, DNA mismatch repair, gene expression control, etc.
The participation of DamMT in such a diverse set of functions
required that other genes adapted, or emerged through evolution, in
response to the DamMT-induced modification of the genomic environment.
One example is SeqA, a protein that senses the methylation status of
the origin of replication of the chromosome to control the timing of
replication initiation.
Interestingly, seqA is only present in a few DamMT-specifying
proteobacteria. This observation led us to hypothesize that other
genes, specifying related functions, might also be found in these
organisms. To test this hypothesis, we implemented a large-scale
comparative genomic screen meant to identify genes specifying DNA
methylation sensing domains, probably involved in DNA maintenance
functions.
Results: We carried out a phylogenetic analysis of DamMT,
identifying two contrasting behaviors of the protein. Based on this
phylogeny, we defined precisely a set of genomes, in which the protein
activity is likely to be involved in DNA maintenance functions, the
'resident' dam genomes. We defined a second set of genomes, in which
DamMT is not resident. We developed a new tool, 'DomainSieve', in
order to screen these two sets for protein domains that are strictly
associated with 'resident' dam genomes.
This approach was rewarding and generated a list of genes, among
which some, at least, specify activities with clear linkage to
DamMT-dependent DNA methylation and DNA maintenance.

<>

<1>Brickner, M., Chmielewski, J.
<2>Inhibiting the dimeric restriction endonuclease EcoRI using interfacial helical peptides.
<3>Chem. Biol.
<4>5
<5>339-343
<6>1998
<7>Many enzymes are active only in a dimeric form, including a variety of type II restriction
endonucleases.  Disruption of subunit interactions is therefore a potential method for
multimeric enzyme inhibition.  EcoRI is a homodimeric restriction endonuclease, the dimeric
interface of which consists of a four-helix bundle.  We set out to design helical peptides to
interact with this interface and block dimer formation, thus rendering EcoRI inactive.  Here
we describe two synthetic, helical peptides based on the interfacial region of EcoRI.  Both
peptides inhibit the enzyme, but the peptide derived from the alpha4 helix of EcoRI had both a
higher helical content and better efficacy than a variant peptide, alpha4(Leu), that has three
lle-Leu mutations (IC50 values of 27 microM and 90 microM, and helical contents of 29% and
10%, respectively).  Size-exclusion chromatography confirmed that the alpha4 peptide disrupted
dimerization of EcoRI, and circular dichroism indicated that EcoRI remained folded upon
binding to alpha4.  Inhibition with alpha4 and alpha4(Leu) was shown to be specific for EcoRI,
as the dimeric restriction enzyme PvuII was not affected by the peptides.

<>

<1>Bridger, S.L., Lancaster, W.A., Poole, F.L.I.I., Schut, G.J., Adams, M.W.
<2>Genome Sequencing of a Genetically Tractable Pyrococcus furiosus Strain Reveals a Highly Dynamic Genome.
<3>J. Bacteriol.
<4>194
<5>4097-4106
<6>2012
<7>The model archaeon Pyrococcus furiosus grows optimally near 100 degrees C on carbohydrates and
peptides. Its genome sequence (NCBI) was determined 12 years
ago. A genetically tractable strain, COM1, was very recently reported, and here
we describe its genome sequence. Of 1,909,827 bp in size, it is 1,571 bp longer
(0.1%) than the reference NCBI sequence. The COM1 genome contains numerous
chromosomal rearrangements, deletions, and single base changes. COM1 also has 45
full or partial insertion sequences (ISs) compared to 35 in the reference NCBI
strain, and these have resulted in the direct deletion or insertional
inactivation of 13 genes. Another seven genes were affected by chromosomal
deletions and are predicted to be nonfunctional. In addition, the amino acid
sequences of another 102 of the 2,134 predicted gene products are different in
COM1. These changes potentially impact various cellular functions, including
carbohydrate, peptide, and nucleotide metabolism; DNA repair; CRISPR-associated
defense; transcriptional regulation; membrane transport; and growth at 72 degrees
C. For example, the IS-mediated inactivation of riboflavin synthase in COM1
resulted in a riboflavin requirement for growth. Nevertheless, COM1 grew on
cellobiose, malto-oligosaccharides, and peptides in complex and minimal media at
98 and 72 degrees C to the same extent as did both its parent strain and a new
culture collection strain (DSMZ 3638). This was in spite of COM1 lacking several
metabolic enzymes, including nonphosphorylating glyceraldehyde-3-phosphate
dehydrogenase and beta-glucosidase. The P. furiosus genome is therefore of high
plasticity, and the availability of the COM1 sequence will be critical for the
future studies of this model hyperthermophile.

<>

<1>Briers, Y., Klumpp, J., Schuppler, M., Loessner, M.J.
<2>Genome Sequence of Listeria monocytogenes Scott A, a Clinical Isolate from a Food-Borne Listeriosis Outbreak.
<3>J. Bacteriol.
<4>193
<5>4284-4285
<6>2011
<7>Listeria monocytogenes is an opportunistic food-borne pathogen and the causative agent of
listeriosis in animals and humans. We present the
genome sequence of Listeria monocytogenes Scott A, a widely distributed
and frequently used serovar 4b clinical isolate from the 1983 listeriosis
outbreak in Massachusetts.

<>

<1>Briggs, R.E., Tatum, F.M.
<2>Characterization of a restriction endonuclease from Pasteurella haemolytica serotype A1 and construction of a gene-replacement Aroa mutant.
<3>Haemophilus, Actinobacillus and Pasteurella, Plenum Press, Donachie, W., Lainson, F.A., Hodgson, J.C., New York
<4>0
<5>221-222
<6>1995
<7>A new restriction endonuclease, PhaI, was isolated from P. haemolytica
serotype
1, strain NADC D60, obtained from pneumonic bovine lung.  PhaI recognizes the 5 base
non-
palindromic sequences 5'-GCATC-3' and 5'-GATGC-3'.  Cleavage occurs 5 bases 3'
from the
former recognition site and 9 bases 5' from the latter.  A gene encoding for a
methyltransferase
which protects against PhaI cleavage was cloned from P. haemolytica into E. coli.
Whereas
unmethylated plasmid DNA containing a P. haemolytica origin of replication was unable to
transform P. haemolytica when introduced by electroporation, the same plasmid DNA
obtained
from E. coli which contained cloned PhaI methyltransferase could do so.  The aroA gene of
P.
haemolytica serotype A1 was cloned and sequenced.  A P. haemolytica ampicillin-
resistance
fragment was cloned into the unique NdeI site of aroA.  A hybrid plasmid was constructed
by
joining the aroA replacement plasmid with the 4.2 kb P. haemolytica plasmid which
encodes
streptomycin resistance.  Following PhaI methylation, the hybrid plasmid was introduced
into P.
haemolytica by electroporation.  Allelic exchange between the replacement plasmid and
chromosome of P. haemolytica gave rise to an ampicillin-resistant mutant which was
unable to
grow on medium deficient in tryptophan.  Although transformation efficiency with
methylated
hybrid plasmid was <10^3/ug, the hybrid was capable of unstable replication in P.
haemolytica, so
this system may be suitable for construction of additional gene-replacement mutants.

<>

<1>Briggs, R.E., Tatum, F.M.
<2>Molecular genetic construction of vaccine strains of Pasteurellaceae.
<3>International Patent Office
<4>WO 9741823
<5>
<6>1997
<7>Tools for genetically engineering Pasteurellaceae are provided.  Replication-conditional
plasmids which are useful for the Pasteurellaceae have been isolated and characterized.  The
plasmids can be utilized for delivery of DNA segments into the Pasteurellaceae in situations
where control of extrachromosomal replication is desired, such as in achieving allelic
exchange or site-directed mutagenesis.  A restriction endonuclease, HsoI, was isolated from a
bovine lung isolate of Haemophilus somnus.  The enzyme was found to be a true isoschizomer of
HinPI, a commercially available enzyme originally isolated from haemophilus influenzae P1.
Commercially available HhaI methyltransferase was found to protect against cleavage by both
enzymes.  Methylation of foreign plasmid DNA was found to enhance transformation of
Haemophilus somnus in excess of four orders of magnitude.

<>

<1>Briggs, R.E., Tatum, F.M.
<2>DNA encoding Pasteurella haemolytica PhaI restriction endonuclease and methyltransferase.
<3>US Patent Office
<4>US 5693777
<5>
<6>1997
<7>Methylation of DNA can be a critical step in the introduction of DNA into P. haemolytica.  A
methyltransferase has been isolated and molecularly cloned for this purpose.  Use of the
methyltransferase has allowed construction of defined, attenuated mutants for use as vaccines
to protect cattle.

<>

<1>Briggs, R.E., Tatum, F.M.
<2>Pasteurella haemolytica PhaI restriction endonuclease and methyltransferase.
<3>US Patent Office
<4>US 5683900
<5>
<6>1997
<7>Methylation of DNA can be a critical step in the introduction of DNA into P. haemolytica.  A
methyltransferase has been isolated and molecularly cloned for this purpose.  Use of the
methyltransferase has allowed construction of defined, attenuated mutants for use as vaccines
to protect cattle.

<>

<1>Briggs, R.E., Tatum, F.M.
<2>Generation and molecular characterization of new temperature-sensitive plasmids intended for genetic engineering of Pasteurellaceae.
<3>Appl. Environ. Microbiol.
<4>71
<5>7187-7195
<6>2005
<7>Temperature-sensitive (TS) plasmids were generated through chemical mutagenesis of a
derivative of the streptomycin resistance parent plasmid pD70, isolated from Mannheimia
hemolytica serotype 1. Three TS plasmids which failed to replicate at or above 42 degrees C in
M. hemolytica but which were fully functional below 31 degrees C were selected for further
analysis. Two of the TS plasmids were shown by sequencing to possess unique single-base-pair
mutations. The third TS plasmid contained a unique base pair substitution and a second
mutation that had been previously identified. These mutations were clustered within a 200-bp
region of the presumed plasmid origin of replication. Site-directed single-nucleotide
substitutions were introduced into the wild-type pD70 origin of replication to confirm that
mutations identified by sequencing had conferred thermoregulated replication. Deletion
analysis on the wild-type pD70 plasmid replicon revealed that approximately 720 bp are
necessary for plasmid maintenance. Replication of the TS plasmids was thermoregulated in
Pasteurella multocida and Haemophilus somnus as well. To consistently transform H. somnus with
TS plasmid, in vitro DNA methylation with commercially available HhaI methyltransferase was
necessary to protect against the organism's restriction enzyme HsoI (recognition sequence
5'-GCGC-3') characterized herein.

<>

<1>Briggs, R.E., Tatum, F.M., Casey, T.A., Frank, G.H.
<2>Characterization of a restriction endonuclease, PhaI, from Pasteurella haemolytica Serotype A1 and protection of heterologous DNA by a cloned PhaI methyltransferase gene.
<3>Appl. Environ. Microbiol.
<4>60
<5>2006-2010
<6>1994
<7>Pasteurella haemolytica is the leading cause of economic loss to the beef cattle industry in
the United States and an important etiologic agent worldwide. Study of P. haemolytica is
hindered by researchers' inability to genetically manipulate the organism. A new restriction
endonuclease, PhaI, an isoschizomer of SfaNI (R.J. Roberts, Methods Enzymol. 65:19-36, 1980),
was isolated from P. haemolytica serotype I, strain NADC-D60, obtained from pneumonic bovine
lung, PhaI recognizes the 5-base nonpalindromic sequences 5'-GCATC-3' and 5'-GATGC-3'.
Cleavage occurs 5 bases 3' from the former recognition site and 9 bases 5' from the latter
recognition site. A gene encoding a methyltransferase which protects against PhaI cleavage was
cloned from P. haemolytica NADC-D60 into Escherichia coli. Whereas unmethylated plasmid DNA
containing a P. haemolytica origin of replication was unable to transform P. haemolytica when
introduced by electroporation, the same plasmid DNA obtained from E. coli which contained a
cloned PhaI methyltransferase gene could do so. The data indicate that PhaI is an effective
barrier to the introduction and establishment of exogenous DNA in P. haemolytica.

<>

<1>Briggs, R.E., Tatum, F.M., Casey, T.A., Frank, G.H.
<2>Characterization of a restriction endonuclease, PhaI, from Pasteurella haemolytica serotype A1 and protection of heterologous DNA by cloned PhaI methyltransferase.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>132
<6>1994
<7>Pasteurella haemolytica is the leading cause of economic loss to the beef cattle industry in
the US, and an important etiologic agent worldwide. Study of P. haemolytica is hindered by
researchers' inability to genetically manipulate the organism. A new restriction
endonuclease, PhaI, an isoschizomer of SfaNI, was isolated from Pasteurella haemolytica
serotype 1, strain NADC-D60, obtained from pneumonic bovine lung. PhaI recognizes the 5 base
non-palindromic sequences 5'-GCATC-3' and 5'-GATGC-3'. Cleavage occurs 5 bases 3' from
the former recognition site and 9 bases 5' from the latter recognition site. A gene encoding
for a methyltransferase which protects against PhaI cleavage was cloned from P. haemolytica
NADC-D60 into E. coli. Whereas unmethylated plasmid DNA containing a P. haemolytica origin of
replication was unable to transform P. haemolytica when introduced by electroporation, the
same plasmid DNA obtained from E. coli which contained cloned PhaI methyltransferase could do
so. The data indicate that PhaI is an effective barrier to the introduction and establishment
of exogenous DNA in P. haemolytica.

<>

<1>Brinkley, C., Burland, V., Keller, R., Rose, D.J., Boutin, A.T., Klink, S.A., Blattner, F.R., Kaper, J.B.
<2>Nucleotide Sequence Analysis of the Enteropathogenic Escherichia coli Adherence Factor Plasmid pMAR7.
<3>Infect. Immun.
<4>74
<5>5408-5413
<6>2006
<7>The complete nucleotide sequence was determined for pMAR7, an
enteropathogenic Escherichia coli (EPEC) adherence factor (EAF) plasmid
that contains genes encoding a type IV attachment pilus (Bfp) and the
global virulence regulator per. Prototypic EAF plasmid pMAR7 is
self-transmissible, unlike the smaller EAF plasmid pB171, which has no
genes encoding conjugative functions. The tra locus, a highly conserved
33-kb segment found in pMAR7, is similar to the tra (conjugation) region
of the F plasmid. ISEc13 copies flanking the pMAR7 tra region could
potentially mobilize or delete the tra genes. Hybridization of 134 EPEC
strains showed that a complete tra region is present only in strains of
the EPEC1 clonal group. This study confirms EPEC's potential for
dissemination of virulence attributes by horizontal transfer of the EAF
plasmid.

<>

<1>Brinkley, P., Bautista, D.S., Graham, F.L.
<2>The cleavage site of restriction endonuclease MnlI.
<3>Gene
<4>100
<5>267-268
<6>1991
<7>The cleavage site generated by restriction endonuclease MnlI has the structure:
5'-CCTC(N)7-3'
3'-GGAG(N)6-5'  with one-nucleotide 3' overhang.

<>

<1>Brinkrolf, K., Schneider, J., Knecht, M., Ruckert, C., Tauch, A.
<2>Draft Genome Sequence of Turicella otitidis ATCC 51513, Isolated from Middle Ear  Fluid from a Child with Otitis Media.
<3>J. Bacteriol.
<4>194
<5>5968-5969
<6>2012
<7>Turicella otitidis is an unusual corynebacterium with a controversial role in otitis media in
children. Metabolic capabilities deduced from the draft genome
indicate its adaptation to habitats on the human skin and in the intestine. The
lack of candidate virulence factors implies that T. otitidis has a low pathogenic
potential.

<>

<1>Broadbent, J.R., Hughes, J.E., Welker, D.L., Tompkins, T.A., Steele, J.L.
<2>Complete Genome Sequence for Lactobacillus helveticus CNRZ 32, an Industrial Cheese Starter and Cheese Flavor Adjunct.
<3>Genome Announcements
<4>1
<5>e00590-13
<6>2013
<7>Lactobacillus helveticus is a lactic acid bacterium widely used in the manufacture of cheese
and for production of bioactive peptides from milk
proteins. We present the complete genome sequence for L. helveticus CNRZ 32, a
strain particularly recognized for its ability to reduce bitterness and
accelerate flavor development in cheese.

<>

<1>Broadbent, J.R., Neeno-Eckwall, E.C., Stahl, B., Tandee, K., Cai, H., Morovic, W., Horvath, P., Heidenreich, J., Perna, N.T., Barrangou, R., Steele, J.L.
<2>Analysis of the Lactobacillus casei supragenome and its influence in species evolution and lifestyle adaptation.
<3>BMC Genomics
<4>13
<5>533
<6>2012
<7>BACKGROUND: The broad ecological distribution of L. casei makes it an insightful
subject for research on genome evolution and lifestyle adaptation. To explore
evolutionary mechanisms that determine genomic diversity of L. casei, we
performed comparative analysis of 17 L. casei genomes representing strains
collected from dairy, plant, and human sources. RESULTS: Differences in L. casei
genome inventory revealed an open pan-genome comprised of 1,715 core and 4,220
accessory genes. Extrapolation of pan-genome data indicates L. casei has a
supragenome approximately 3.2 times larger than the average genome of individual
strains. Evidence suggests horizontal gene transfer from other bacterial species,
particularly lactobacilli, has been important in adaptation of L. casei to new
habitats and lifestyles, but evolution of dairy niche specialists also appears to
involve gene decay. CONCLUSIONS: Genome diversity in L. casei has evolved through
gene acquisition and decay. Acquisition of foreign genomic islands likely confers
a fitness benefit in specific habitats, notably plant-associated niches. Loss of
unnecessary ancestral traits in strains collected from bacterial-ripened cheeses
supports the hypothesis that gene decay contributes to enhanced fitness in that
niche. This study gives the first evidence for a L. casei supragenome and
provides valuable insights into mechanisms for genome evolution and lifestyle
adaptation of this ecologically flexible and industrially important lactic acid
bacterium. Additionally, our data confirm the Distributed Genome Hypothesis
extends to non-pathogenic, ecologically flexible species like L. casei.

<>

<1>Broadbent, S.E., Balbontin, R., Casadesus, J., Marinus, M.G., van der Woude, M.
<2>YhdJ, a Nonessential CcrM-Like DNA Methyltransferase of Escherichia coli and Salmonella enterica.
<3>J. Bacteriol.
<4>189
<5>4325-4327
<6>2007
<7>The Caulobacter crescentus DNA adenine methyltransferase CcrM and its homologs in the
alpha-Proteobacteria are essential for viability. CcrM is
34% identical to the yhdJ gene products of Escherichia coli and Salmonella
enterica. This study provides evidence that the E. coli yhdJ gene encodes
a DNA adenine methyltransferase. In contrast to an earlier report,
however, we show that yhdJ is not an essential gene in either E. coli or
S. enterica.

<>

<1>Broberg, C.A., Palacios, M., Miller, V.L.
<2>Whole-Genome Draft Sequences of Three Multidrug-Resistant Klebsiella pneumoniae Strains Available from the American Type Culture Collection.
<3>Genome Announcements
<4>1
<5>e00312-13
<6>2013
<7>Infection with multidrug-resistant Klebsiella pneumoniae is a significant problem worldwide,
requiring a better understanding of how various strains are able to
defeat current antibiotic therapies and cause disease. Here, we report the draft
genome sequences of three K. pneumoniae strains harboring the SHV-18, KPC-2, or
NDM-1 beta-lactamases.

<>

<1>Broberg, C.A., Wu, W., Cavalcoli, J.D., Miller, V.L., Bachman, M.A.
<2>Complete Genome Sequence of Klebsiella pneumoniae Strain ATCC 43816 KPPR1, a Rifampin-Resistant Mutant Commonly Used in Animal, Genetic, and Molecular Biology  Studies.
<3>Genome Announcements
<4>2
<5>e00924-14
<6>2014
<7>Klebsiella pneumoniae is an urgent public health threat due to the spread of
carbapenem-resistant strains causing serious, and frequently fatal, infections.
To facilitate genetic, molecular, and immunological studies of this pathogen, we
report the complete chromosomal sequence of a genetically tractable, prototypical
strain used in animal models.

<>

<1>Brocchi, M., de Vasconcelos, A.T.R., Zaha, A.
<2>Restriction-modification systems in Mycoplasma spp.
<3>Genet. Mol. Biol.
<4>30
<5>236-244
<6>2007
<7>Restriction and Modification (R-M) systems are present in all Mycoplasma species sequenced so
far. The presence of these genes poses
barriers to gene transfer and could protect the cell against phage
infections. The number and types of R-M genes between different
Mycoplasma species are variable, which is characteristic of a
polymorphism. The majority of the CDSs code for Type III R-M systems
and particularly for methyltransferase enzymes, which suggests that
functions other than the protection against the invasion of
heterologous DNA may exist. A possible function of these enzymes could
be the protection against the invasion of other but similar R-M
systems. In Mycoplasma hyopneumoniae strain J, three of the putative
methyltransferase genes were clustered in a region forming a genomic
island. Many R-M CDSs were mapped in the vicinity of transposable
elements suggesting an association between these genes and reinforcing
the idea of R-M systems as mobile selfish DNA. Also, many R-M genes
present repeats within their coding sequences, indicating that their
expression is under the control of phase variation mechanisms.
Altogether, these data suggest that R-M systems are a remarkable
characteristic of Mycoplasma species and are probably involved in the
adaptation of these bacteria to different environmental conditions.

<>

<1>Brockes, J.P.
<2>Nucleotide sequences at the sites of action of the deoxyribonucleic acid modification enzyme of bacteriophage P1.
<3>J. Mol. Biol.
<4>88
<5>437-443
<6>1974
<7>Labelled oligonucleotides have been fractionated from DNAase digests of phage
lambda DNA that had been methylated with the phage P1 modification enzyme and
S-[methyl-14C]adenosyl-L-methionine.  The longest sequences established are the
tetranucleotides pG-A*-T-C and pA*-T-C-T, which, together with the other
sequences determined, particularly pA*-G-A, show that the modification enzyme,
M.EcoP1, methylates adenine residues within defined sequences and suggest that
the oligonucleotide sequence recognized by this enzyme is the hexanucleotide
pA-G-A-T-C-T.  The duplex formed by base-pairing this hexanucleotide with its
complementary sequence resembles the recognition sequence for several
restriction enzymes in being bisected by an axis of 2-fold rotational symmetry.

<>

<1>Brockes, J.P.
<2>The deoxyribonucleic acid-modification enzyme of bacteriophage P1. Subunit structure.
<3>Biochem. J.
<4>133
<5>629-633
<6>1973
<7>The bacteriophage P1 modification enzyme was purified 1400-fold from induced
lysogens of a thermoinducible mutant of bacteriophage P1.  The most purified
fraction, when analysed by polyacrylamide-gel electropohoresis in sodium
dodecyl sulphate, showed two principal stained bands.  The two bands
co-sedimented in a glycerol gradient with the modification activity, at a rate
which, when compared with the rate of sedimentation of marker proteins,
corresponds to a sedimentation coefficient in water of 6S.  The mobilities of
the bands on sodium dodecyl sulphate-polyacrylamide-gel electrophoresis
corresponded to polypeptides of molecular weight 70000 and 45000 and they were
present in equimolar amounts.  It was concluded that the 6S species of the
enzyme is a dimer of unlike subunits.

<>

<1>Brockes, J.P., Brown, P.R., Murray, K.
<2>The deoxyribonucleic acid modification enzyme of bacteriophage P1: Purification and properties.
<3>Biochem. J.
<4>127
<5>1-10
<6>1972
<7>The bacteriophage P1 modification enzyme, assayed by the specific methylation
of unmodified bacteriophage 82 DNA, has been purified 500-fold from a
bacteriophage P1 lysogen of Escherichia coli.  The enzyme catalyses the
incorporation of approximately 20-24 methyl groups per bacteriophage 82 DNA
molecule.  The sole product of methylation is 6-methylaminopurine.  Methylation
of unmodified bacteriophage DNA confers protection against a challenge by
purified bacteriophage P1 restriction enzyme.  The pH optimum is 6.0-6.25: the
apparent Km for S-adenosyl-L-methionine is 5 microM.

<>

<1>Brok-Volchanskaya, V.S., Kadyrov, F.A., Sivogrivov, D.E., Kolosov, P.M., Sokolov, A.S., Shlyapnikov, M.G., Kryukov, V.M., Granovsky, I.E.
<2>Phage T4 SegB protein is a homing endonuclease required for the preferred inheritance of T4 tRNA gene region occurring in co-infection with a related phage.
<3>Nucleic Acids Res.
<4>36
<5>2094-2105
<6>2008
<7>Homing endonucleases initiate nonreciprocal transfer of DNA segments containing their own
genes and the flanking sequences by cleaving the
recipient DNA. Bacteriophage T4 segB gene, which is located in a
cluster of tRNA genes, encodes a protein of unknown function,
homologous to homing endonucleases of the GIY-YIG family. We
demonstrate that SegB protein is a site-specific endonuclease, which
produces mostly 3 2-nt protruding ends at its DNA cleavage site.
Analysis of SegB cleavage sites suggests that SegB recognizes a 27-bp
sequence. It contains 11-bp conserved sequence, which corresponds to a
conserved motif of tRNA TC stem-loop, whereas the remainder of the
recognition site is rather degenerate. T4-related phages T2L, RB1 and
RB3 contain tRNA gene regions that are homologous to that of phage T4
but lack segB gene and several tRNA genes. In co-infections of phages
T4 and T2L, segB gene is inherited with nearly 100 of efficiency. The
preferred inheritance depends absolutely on the segB gene integrity and
is accompanied by the loss of the T2L tRNA gene region markers. We
suggest that SegB is a homing endonuclease that functions to ensure
spreading of its own gene and the surrounding tRNA genes among
T4-related phages.

<>

<1>Brolund, A., Franzen, O., Melefors, O., Tegmark-Wisell, K., Sandegren, L.
<2>Plasmidome-Analysis of ESBL-Producing Escherichia coli Using Conventional Typing and High-Throughput Sequencing.
<3>PLoS ONE
<4>8
<5>E65793
<6>2013
<7>Infections caused by Extended spectrum beta-lactamase (ESBL)-producing E. coli
are an emerging global problem, threatening the effectiveness of the extensively
used beta-lactam antibiotics. ESBL dissemination is facilitated by plasmids,
transposons, and other mobile elements. We have characterized the plasmid content
of ESBL-producing E. coli from human urinary tract infections. Ten diverse
isolates were selected; they had unrelated pulsed-field gel electrophoresis
(PFGE) types (<90% similarity), were from geographically dispersed locations and
had diverging antibiotic resistance profiles. Three isolates belonged to the
globally disseminated sequence type ST131. ESBL-genes of the CTX-M-1 and CTX-M-9
phylogroups were identified in all ten isolates. The plasmid content (plasmidome)
of each strain was analyzed using a combination of molecular methods and
high-throughput sequencing. Hidden Markov Model-based analysis of unassembled
sequencing reads was used to analyze the genetic diversity of the plasmid samples
and to detect resistance genes. Each isolate contained between two and eight
distinct plasmids, and at least 22 large plasmids were identified overall. The
plasmids were variants of pUTI89, pKF3-70, pEK499, pKF3-140, pKF3-70, p1ESCUM,
pEK204, pHK17a, p083CORR, R64, pLF82, pSFO157, and R721. In addition, small
cryptic high copy-number plasmids were frequent, containing one to seven open
reading frames per plasmid. Three clustered groups of such small cryptic plasmids
could be distinguished based on sequence similarity. Extrachromosomal prophages
were found in three isolates. Two of them resembled the E. coli P1 phage and one
was previously unknown. The present study confirms plasmid multiplicity in
multi-resistant E. coli. We conclude that high-throughput sequencing successfully
provides information on the extrachromosomal gene content and can be used to
generate a genetic fingerprint of possible use in epidemiology. This could be a
valuable tool for tracing plasmids in outbreaks.

<>

<1>Bromberg, S., Pratt, K., Hattman, S.
<2>Sequence specificity of DNA adenine methylase in the protozoan Tetrahymena thermophila.
<3>J. Bacteriol.
<4>150
<5>993-996
<6>1982
<7>The sequence specificity of the Tetrahymena DNA-adenine methylase was determined by
nearest-neighbor analyses of in vivo and in vitro methylated DNA.  In vivo all four common
bases were found to the 5' side of N6-methyladenine, but only thymidine was 3'.  Homologous
DNA already methylated in vivo and heterologous Micrococcus luteus DNA were methylated in
vitro by a partially purified DNA-adenine methylase activity isolated from Terrahymena
macronuclei.  The in vitro-methylated sequence differed from the in vivo sequence in that both
thymidine and cytosine were 3' nearest neighbors of N6-methyladenine.

<>

<1>Bron, S., Horz, W.
<2>Purification and properties of the Bsu endonuclease.
<3>Methods Enzymol.
<4>65
<5>112-132
<6>1980
<7>For various reasons Bacillus subtilis is an attractive organism to study
restriction and modification.  It is easy to grow and to manipulate, has a
fairly detailed genetic map, and, most important, possesses a simple
transformation/transfection system.  This enables the introduction of
biologically active DNA into cells of defined R-M background, and the analysis
of its fate.  In addition, the effects of in vitro treatments of biologically
active DNA with restriction and modification enzymes can conveniently be
studied by means of transformation and transfection.  Restriction and
modification in B. subtilis were demonstrated some years ago.  A site-specific
type II restriction endonuclease, Bsu, has been purified and characterized from
B. subtilis strain R.  The enzyme cleaves susceptible DNAs in the middle of the
tetranucleotide sequence 5'GGCC3'.  Resistance to Bsu is conferred to DNA by
the modification methylase from the same strain, which methylates the internal
C residues of the recognition sequence.  We will describe the purification and
properties of Bsu.

<>

<1>Bron, S., Janniere, L., Ehrlich, S.D.
<2>Restriction and modification in Bacillus subtilis Marburg 168:  Target sites and effects on plasmid transformation.
<3>Mol. Gen. Genet.
<4>211
<5>186-189
<6>1988
<7>The effects of the restriction system of Bacillus subtilis strain M on plasmid
transformation were studied.  Plasmid pHV1401 DNA prepared from B. subtilis
transformed the restriction-proficient M strain 100 times more efficiently than
the DNA prepared from Escherichia coli, while the two DNA preparations
transformed restriction-deficient derivatives of that strain with similar
efficiencies.  This indicates that transformation with pHV1401 is sensitive to
the M restriction system.  pHV1401 contains three CTCGAG (XhoI sites).
Successive removal of these abolished the effect of restriction.  This
indicates that the XhoI sites are the targets for the M restriction system.

<>

<1>Bron, S., Luxen, E., Trautner, T.A.
<2>Restriction and modification in B. subtilis:  the role of homology between donor and recipient DNA in transformation and transfection.
<3>Mol. Gen. Genet.
<4>179
<5>111-117
<6>1980
<7>Non-modified DNAs from phages SPO2 and Phi105, and prophage DNAs extracted from
lysogens carrying these phages, were used to transfect isogenic r-m- B.
subtilis recipients which were either non-lysogenic, or had been lysogenized
with a homologous or a non-homologous phage.  Restriction of transfecting phage
and prophage DNA occurred in non-lysogenic recipients and in recipients
lysogenic for a non-homologous phage.  No effect of restriction was observed
when phage or prophage DNA was used to transfect recipients carrying a
homologous prophage.  This is analogous to the absence of restriction in
transformation and indicates that in B. subtilis the distinction between
transforming and transfecting DNA is not made at the initial stages of DNA
uptake and processing, but rather at later stages, where recognition of
homologous regions in donor and recipient DNA plays an important role.

<>

<1>Bron, S., Luxen, E., Venema, G.
<2>Resistance of bacteriophage H1 to restriction and modification by Bacillus subtilis R.
<3>J. Virol.
<4>46
<5>703-708
<6>1983
<7>H1, a 5-hydroxymethyluracil (HMU)-containing Bacillus subtilis bacteriophage,
was neither restricted nor modified upon infection of B. subtilis R cells.  In
vitro, H1 DNA was not restricted by BsuR under standard conditions (200 mM
salt), although the expected frequency of -GGCC- cleavage sites was
approximately 250.  However, four specific sites were cleaved under nonstandard
conditions (low salt or high pH) or in the presence of organic solvents, like
dimethyl sulfoxide and glycerol.  After the substitution of thymine for HMU by
DNA cloning in B. subtilis, a BsuR cleavage site was restricted and modified
under standard conditions.  No additional sites were detected after
shotgun-cloning of about 11% of the chromosome.  The nucleotide sequence of a
cleavage site was found to be
5'..C-A-Hmu-A-A-C-Hmu-Hmu-Hmu-G-G-C-C-Hmu-A-G-..3', which shows the presence of
a bona fide BsuR (GGCC) recognition sequence, flanked by (Hmu-A)-rich
sequences.  The results suggested that the resistance of H1 to restriction and
modification by B. subtilis R was due to (i) a strong bias against the
GGCC-recognition sequence and (ii) protection of the four remaining GGCC sites
as a consequence of HMU-A base pairs flanking the sites.

<>

<1>Bron, S., Luxen, E., Venema, G.
<2>Restriction and modification in Bacillus subtilis: Effects on transfection under marker rescue conditions.
<3>J. Virol.
<4>42
<5>357-364
<6>1982
<7>The role of homology between donor and recipient DNAs in the protection of transfecting DNA
against restriction by competent Bacillus subtilis R cells was studied under marker rescue
conditions with modified helper phage.  By comparing restriction under conditions of
preinfection marker rescue and superinfection marker rescue, the significance of DNA homology
during the initial stages of DNA processing by competent cells could be studied.  The results
showed that in both preinfection and superinfection, complete protection against restriction
of transfectants produced via rescue by the modified homologous helper chromosome occurred.
Even up to 90 min after entry, DNA entering the helper-mediated pathway of transfection was
not affected by restriction.  The significance of these findings is discussed in the general
context of the role of DNA homology between donor and recipient on the fate of donor DNA in
competent B. subtilis, in particular in relation to the effects on restriction.

<>

<1>Bron, S., Luxen, E., Venema, G.
<2>Restriction of hemimethylated DNA by the Bacillus subtilis R system.
<3>Mol. Gen. Genet.
<4>195
<5>370-373
<6>1984
<7>The effects of restriction by the BsuR system on hemimethylated SPP1 DNA were investigated.
In vitro, single-stranded nicks were introduced in the nonmodified strand of the
hemimethylated DNA at the same sites as recognized in nonmodified homoduplex DNA.
Transfection with BsuR-treated hemimethylated DNA was severely reduced.  In vivo, transfection
with hemimethylated DNA was also severely reduced in competent B. subtilis R cells.  In
contrast, transfection of protoplasts of the R strain with this DNA was not affected.  The
apparent restriction by competent cells was attributed to the special mode of processing of
transfecting DNA.

<>

<1>Bron, S., Luxen, E., Venema, G., Trautner, T.
<2>Restriction and modification in B. subtilis:  Effects on transformation and transfection with native and single-stranded DNA.
<3>Mol. Gen. Genet.
<4>179
<5>103-110
<6>1980
<7>The effects of restriction in vivo by competent B. subtilis R cells and in
vitro by purified endonuclease BsuR on transformation and transfection with
native and denatured DNA were investigated.  The results show that
transformation by either native, or denatured DNA is not affected by
restriction, whereas transfection both with native and denatured SPP1 DNA is
severely restricted.  In contrast to the results obtained in vivo, the
biological activity of native and denatured transforming DNA is destroyed by
BsuR in vitro, as is the transfecting activity of native and denatured SPP1
DNA.  The sensitivity of denatured DNA, either with mixtures of the
complementary strands or with separated single strands alone, is significantly
lower than that of native DNA.  The results are discussed in the context of
possible mechanisms underlying the different responses of transforming and
transfecting DNA to in vivo restriction by B. subtilis R cells.

<>

<1>Bron, S., Murray, K.
<2>Restriction and modification in B. subtilis: Nucleotide sequence recognized by restriction endonuclease R.BsuR from strain R.
<3>Mol. Gen. Genet.
<4>143
<5>25-33
<6>1975
<7>Restriction endonuclease R from Bacillus subtilis strain R cleaves nonmodified
SPP1 DNA in approximately 80, and lambda DNA in about 200 different sites.  DNA
digests with this endonuclease and with endonuclease HaeIII from Haemophilus
aegyptius show identical fragmentation patterns on gel electrophoresis,
indicating that the two enzymes recognize the same nucleotide sequence.  The
polynucleotide kinase reaction was used in conjunction with two-dimensional
ionophoretic nucleotide mapping methods to identify the 5'-nucleotide sequences
at the sites of cleavage by the B. subtilis restriction endonuclease.  The
results show that the recognition sequence is 5' N-G-G^C-C-N- 3' 3'
N-C-C^G-G-N- 5' where arrows indicate the point of strand scission.  Each of
the four possible nucleotides can occur in the positions flanking the
recognition site.

<>

<1>Bron, S., Murray, K., Trautner, T.A.
<2>Restriction and Modification in B. subtilis:  Purification and General Properties of a Restriction Endonuclease from Strain R.
<3>Mol. Gen. Genet.
<4>143
<5>13-23
<6>1975
<7>All Bacillus subtilis R-type strains showing the phenomena of restriction and
modification contain an endonuclease that inactiviates in vitro the biological
activity of a variety of DNAs lacking R-specific modification, such as
transfecting SPP1, SPO2 and Phi-105 DNA, and transforming B. subtilis 168-type
DNA.  The corresponding DNAs carrying R-specific modification are resistant to
the enzyme.  The enzyme has been purified approximately 400-fold and is
essentially free from contaminating double strand-directed unspecific exo- or
endonuclease activity.  Only Mg2+ is required as cofactor.  The substrate DNAs
are cleaved at specific sites.  The double-stranded fragments produced from
SPP1 DNA (molecular weight 2.5 x 107) have an average molecular weight of about
300,000.

<>

<1>Brooks, J., Hattman, S.
<2>Location of the DNA-adenine methylase gene on the genetic map of phage T2.
<3>Virology
<4>55
<5>285-288
<6>1973
<7>The location of the gene controlling DNA-adenine methylase activity is reported to be in the
region between genes 49 and rI on the T2 genetic map. A new nomenclature is proposed to
describe the various phage types affected by mutations in the methylase gene.

<>

<1>Brooks, J., Masurekar, M., Hattman, S.
<2>In vitro methylation of phage lambda.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>75
<5>225
<6>1975
<7>Phage T2 produces an adenine-specific DNA methylase. Certain mutants (designated damh) produce
an altered enzyme which methylates various forms of T2 DNA to higher extents than does the
wild-type (dam+) enzyme. The higher methylation by damh protects nonglucosylated T2 DNA
against P1-restriction. We have studied in vitro methylation of phage lambda DNA by the dam+
and damh enzymes (using both crude extracts and highly purified enzyme preparations). DNA from
lambda grown in Escherichia coli strain K (=lambda.K DNA) is methylated to a two-fold greater
extent by the damh enzyme; ca.150 and ca.300 MeAde residues per lambda DNA are produced by
dam+ and damh, respectively (the same stoichiometry is observed with lambda.K(P1)DNA).
Following methylation by dam+ enzyme there is no change in the ability of lambda.K DNA to
transfect spheroplasts derived from E. coli K or K(P1); the relative transfecting titer
remains ca.10000 fold higher on K than K(P1). In contrast, methylation by the damh enzyme
resulted in a 1000 to 10000 fold increase in transfection ability on K(P1) spheroplasts; the
transfection efficiency on K(P1) was 20 to 40% that observed on K (under similar conditions
lambda.K(P1) DNA transfected both spheroplast preparations equally well). Thus, the damh
methylase recognizes more sites on lambda DNA than the dam+ enzyme; the additional methylation
protects against P1-restriction.

<>

<1>Brooks, J.E.
<2>Properties and uses of restriction endonucleases.
<3>Methods Enzymol.
<4>152
<5>113-129
<6>1987
<7>This chapter focuses on the properties and uses of restriction endonucleases.
The large battery of endonucleases now commercially available will first be
described in terms of nomenclature and properties.  The next section will cover
basic methods for their use in mapping and genetic engineering experiments, and
a final section will focus on the more detailed aspects of selecting
endonucleases for generating ends compatible with subsequent steps of
construction.

<>

<1>Brooks, J.E.
<2>Method for producing the BamHI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5137823
<5>
<6>1992
<7>A two-step method for cloning the BamHI restriction modification system is provided which
comprises introducing the BamHI modification or methylase gene into a host cell and expressing
the gene to protect the host cell followed by introducing the BamHI restriction or
endonuclease gene into the host.

<>

<1>Brooks, J.E.
<2>A comparative study of the wild type and mutant forms of phage T2 DNA-adenine methylase.
<3>Ph.D. Thesis, University of Rochester
<4>
<5>1-188
<6>1977
<7>Bacteriophage T2 produces an adenine-specific DNA methylase.  Certain mutants (designated
damh) produce an altered enzyme which methylates T2 DNA to a higher extent than does the wild
type (dam+) enzyme.  The higher methylation by the damh enzyme protects nonglucosylated phage
against P1 restriction.  The position of the T2 dam gene on the phage genetic map was
determined by a series of three-factor crosses.  The gene was located between the 49 and rI
loci on the map.  The kinetics of enzyme production were also studied: under the conditions
used, overproduction of methylase activity could not be achieved by blocking DNA synthesis.

<>

<1>Brooks, J.E., Benner, J.S., Heiter, D.F., Silber, K.R., Sznyter, L.A., Jager-Quinton, T., Moran, L.S., Slatko, B.E., Wilson, G.G., Nwankwo, D.O.
<2>Cloning the BamHI restriction modification system.
<3>Nucleic Acids Res.
<4>17
<5>979-997
<6>1989
<7>BamHI, a Type II restriction modification system from Bacillus amyloliquefaciens H recognizes
the sequence GGATCC. The methylase and endonuclease genes have been cloned into E. coli in
separate steps; the clone is able to restrict unmodified phage. Although within the clone the
methylase and endonuclease genes are present on the same pACYC184 vector, the system can be
maintained in E. coli only with an additional copy of the methylase gene present on a separate
vector. The initial selection for BamHI methylase activity also yielded a second BamHI
methylase gene which is not homologous in DNA sequence and hybridizes to different genomic
restriction fragments than does the endonuclease-linked methylase gene. Finally, the
interaction of the BamHI system with the E. coli Dam and the Mcr A and B functions, have been
studied and are reported here.

<>

<1>Brooks, J.E., Benner, J.S., Silber, K.R., Heiter, D.F., Sznyter, L.A., Jager-Quinton, T., Wilson, G.G., Moran, L.S., Slatko, B.E., Nwankwo, D.O.
<2>Cloning and characterization of the BamHI restriction modification system.
<3>Gene
<4>74
<5>13
<6>1988
<7>Meeting Abstract

<>

<1>Brooks, J.E., Blumenthal, R.M., Gingeras, T.R.
<2>The isolation and characterization of the Escherichia coli DNA adenine methylase (dam) gene.
<3>Nucleic Acids Res.
<4>11
<5>837-851
<6>1983
<7>The E. coli dam (DNA adenine methylase) enzyme is known to methylate the sequence GATC. A
general method for cloning sequence-specific DNA methylase genes was used to isolate the dam
gene on a 1.14 kb fragment, inserted in the plasmid vector pBR322. Subsequent restriction
mapping and subcloning experiments established a set of approximate boundaries of the gene.
The nucleotide sequence of the dam gene was determined, and analysis of that sequence revealed
a unique open reading frame which corresponded in length to that necessary to code for a
protein the size of dam. Amino acid composition derived from this sequence corresponds closely
to the amino acid composition of the purified dam protein. Enzymatic and DNA:DNA hybridization
methods were used to investigate the possible presence of dam genes in a variety of
prokaryotic organisms.

<>

<1>Brooks, J.E., Hattman, S.
<2>In vitro methylation of bacteriophage lambda DNA by wild type (dam+) mutant (damh) forms of the phage T2 DNA adenine methylase.
<3>J. Mol. Biol.
<4>126
<5>381-394
<6>1978
<7>The wild-type (dam+) and mutant (damh) forms of the bacteriophage T2 DNA adenine methylase
have been partially purified; these enzymes methylate the sequence, 5'...G-A-Py...3'
(Hattman et al., 1978a). However, in vitro methylation studies using phage lambda DNA revealed
the following: (1) T2 dam+ and damh enzymes differ in their ability to methylate lambda DNA;
under identical reaction conditions the T2 damh enzyme methylated lambda DNA to a higher level
than did the dam+ enzyme. However, the respective methylation sites are equally distributed on
the l and r strands. (2) Methylation with T2 damh, but not T2 dam+ protected lambda against P1
restriction. This was demonstrated by transfection of Escherichia coli (P1) spheroplasts and
by cleavage with R.EcoP1. (3) T2 dam+ and damh were similarly capable of methylating G-A-T-C
sequences on lambda DNA; e.g. k-dam3 DNA (contains no N6-methyladenine) methylated with either
enzyme was made resistant to cleavage by R.DpnII. In contrast, only the T2 damh modified DNA
was resistant to further methylation by M.EcoP1 (which methylates the sequence
5'...A-G-A-C-Py...3'; Hattman et al., 1978b). (4) lambda.dam3 DNA was partially methylated
to the same level with T2 dam+ or T2 damh; the two enzymes producted different patterns of
G-A-C versus G-A-T methylation. We propose that the T2 dam+ enzyme methylates G-A-C sequences
less efficiently than the T2 damh methylase; this property does not entirely account for the
large difference in methylation levels produced by the two enzymes.

<>

<1>Brooks, J.E., Heiter, D.F., Anton, B.P.
<2>Method for cloning and producing the BglII restriction endonuclease and modification methylase.
<3>European Patent Office
<4>EP 0619373 A
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the BglII endonuclease
by: 1) introducing the restriction endonuclease gene from Bacillus globigii into a host
whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid
encoding and expressing the BglII restriction endonuclease activity; and 3) purifying the
BglII restriction endonuclease from the fermented host which contains the plasmid encoding and
expressing the BglII restriction endonuclease.

<>

<1>Brooks, J.E., Heiter, D.F., Anton, B.P.
<2>Method for cloning and producing the BglII restriction endonuclease and modification methylase.
<3>European Patent Office
<4>EP 0619373 B
<5>
<6>2000
<7>
<>

<1>Brooks, J.E., Heiter, D.F., Anton, B.P.
<2>Method for cloning and producing the BglII restriction endonuclease and modification methylase.
<3>Japanese Patent Office
<4>JP 2005237393 A
<5>
<6>2005
<7>
<>

<1>Brooks, J.E., Heiter, D.F., Anton, B.P.
<2>Method for cloning and producing the BglII restriction endonuclease and modification methylase.
<3>US Patent Office
<4>US 5434068
<5>
<6>1995
<7>The present invention is directed to a method for cloning and producing the BglII endonuclease
by: 1) introducing the restriction endonuclease gene from Bacillus globigii into a host
whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid
encoding and expressing the BglII restriction endonuclease activity; and 3) purifying the
BglII restriction endonuclease from the fermented host which contains the plasmid encoding and
expressing the BglII restriction endonuclease.

<>

<1>Brooks, J.E., Howard, K.A.
<2>Method for cloning restriction modification system.
<3>US Patent Office
<4>US 5320957
<5>
<6>1994
<7>A two-step method for cloning restriction modification systems is provided which comprises
introducing the modification or methylase gene into a host cell and expressing the gene
therein to protect the host cell followed by introducing the restriction or endonuclease gene
into the host whereby the restriction gene is expressed at lower levels relative to the
modification gene. Related methods for producing restriction enzymes are also disclosed as
well as clones containing the BamHI and DdeI restriction genes.

<>

<1>Brooks, J.E., Howard, K.A.
<2>Method for cloning restriction modification systems.
<3>European Patent Office
<4>EP 0248678
<5>
<6>1994
<7>A two-step method for cloning restriction modification systems is provided which comprises
introducing the modification or methylase gene into a host cell and expressing the gene
therein to protect the host cell followed by introducing the restriction or endonuclease gene
into the host whereby the restriction gene is expressed at lower levels relative to the
modification gene. Related methods for producing restriction enzymes is also disclosed as well
as clones containing the BamI and DdeI restriction genes.

<>

<1>Brooks, J.E., Howard, K.A.
<2>Method for cloning the DdeI restriction modification system.
<3>European Patent Office
<4>EP 0618295 B
<5>
<6>1996
<7>This invention relates to clones for the DdeI restriction endonuclease and modification
methylase, and related methods for the production of these clones and enzymes.

<>

<1>Brooks, J.E., Howard, K.A.
<2>Method for cloning restriction modification systems.
<3>European Patent Office
<4>EP 0618296
<5>
<6>1996
<7>A two-step method for cloning restriction modification systems is provided which comprises
introducing the modification or methylase gene into a host cell and expressing the gene
therein to protect the host cell followed by introducing the restriction or endonuclease gene
into the host whereby the restriction gene is expressed at lower levels relative to the
modification gene.

<>

<1>Brooks, J.E., Nathan, P.D., Landry, D., Sznyter, L.A., Waite-Rees, P., Ives, C.L., Moran, L.S., Slatko, B.E., Benner, J.S.
<2>Characterization of the cloned BamHI restriction modification system:  its nucleotide sequence, properties of the methylase, and expression in heterologous hosts.
<3>Nucleic Acids Res.
<4>19
<5>841-850
<6>1991
<7>The BamHI restriction modification system was previously cloned into E. coli and maintained
with an extra copy of the methylase gene on a high copy vector (Brooks et al, (1989) Nucl.
Acids Res. 17, 979-997). The nucleotide sequence of a 3014 bp region containing the
endonuclease (R) and methylase (M) genes has now been determined. The sequence predicts a
methylase protein of 423 amino acids, Mr 49,527, and an endonuclease protein of 213 amino
acids, Mr 24,570. Between the two genes is a small open reading frame capable of encoding a
102 amino acid protein, Mr 13,351. The M.BamHI enzyme has been purified from a high expression
clone, its amino terminal sequence determined, and the nature of its substrate modification
studied. The BamHI methylase modifies the internal C within its recognition sequence at the N4
position. Comparisons of the deduced amino acid sequence of M.BamHI have been made with those
available for other DNA methylases: among them, several contain five distinct regions, 12 to
22 amino acids in length, of pronounced sequence similarity. Finally, stability and expression
of the BamHI system in both E. coli and B. subtilis have been studied. The results suggest R
and M expression are carefully regulated in a natural host like B. subtilis.

<>

<1>Brooks, J.E., Nwankwo, D., Sznyter, L., Jager, T., Wilson, G., Heiter, D., Slatko, B., Benner, J.
<2>Cloning of the BamHI restriction-modification system.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>87
<5>154
<6>1987
<7>BamHI, a TypeII restriction-modification system from Bacillus
amyloliquefaciens, recognizes the sequence GGATCC.  The system has been cloned
into E. coli in two steps.  First the methylase gene was cloned into pBR322 and
a derivative expressing higher levels was constructed using a pUC vector.  The
endonuclease gene was then located using Southern blot analyses; HindIII
fragments large enough to contain the endonuclease gene were cloned into
pACYC184, introduced into a host containing the methylase gene and screened for
endonuclease activity.  Both genes are stably maintained in E. coli on separate
but compatible plasmids.

<>

<1>Brooks, J.E., O'Donnell, K.H.
<2>Method for producing the DdeI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5354680
<5>
<6>1994
<7>A two-step method for cloning the DdeI restriction modification system is provided which
comprises introducing the DdeI modification or methylase gene into a host cell and expressing
the gene to protect the DdeI host cell followed by introducing the DdeI restriction or
endonuclease gene into the host.

<>

<1>Brooks, J.E., Roberts, R.J.
<2>Modification profiles of bacterial genomes.
<3>Nucleic Acids Res.
<4>10
<5>913-934
<6>1982
<7>DNAs were prepared from twenty-six bacterial species and digested with a
variety of restriction endonucleases to determine what modifications the DNAs
carry.  Several general conclusions could be made: 1) First, in no instance was
the DNA of a restriction enzyme strain cleaved by its own restriction enzyme.
2) The specificity of the DNA modification was the same as that of its
restriction counterpart; there were no cases of the DNAs being modified against
a less specific class of restriction enzymes. 3) In most (but not all) cases,
the resistance of a bacterium's DNA to its own restriction enzyme could be
generalized to include resistance to all other restriction enzymes with the
same specificity (isoschizomers).  4) DNA modified within the central tetramer
of a recognition sequence is usually protected against cleavage by all related
hexameric enzymes possessing that central tetramer.  Only three families of DNA
presented in this study disobey this rule.  5) Finally, a significant number of
cases emerge where bacterial DNA carries a modification but no corresponding
restriction endonuclease activity.

<>

<1>Brooks, J.E., Sznyter, L., Vaccaro, C., Arnaud, M., Jager-Quinton, T., Wilson, G., Moran, L., Slatko, B., Bernan, V.
<2>Cloning and characterization of the SphI restriction modification system from Streptomyces phaeochromogenes.
<3>J. Cell Biochem. Suppl.
<4>14A
<5>106
<6>1990
<7>SphI, a Type II restriction modification from Streptomyces phaeochromogenes,
recognizes the sequence GCATGC.  The endonuclease cleaves at GCATG^C to
generate a 3' four base overhang.  A 5.4 kb PstI fragment from the S.
phaeochromogenes genome has been cloned into pBR322 and shown to express the
SphI methylase at low level in E. coli.  Clones carrying the fragment have no
endonuclease activity.  Extensive mapping and subcloning have been used to
determine the position and orientation of the m gene.  The nucleotide sequence
of the region is now being determined.  The entire 5.4 kb fragment and also
various restriction fragments have been transferred into S. lividans on pIJ486,
pIJ487 and also the low copy vector pIJ922.  Their expression in Streptomyces
will be discussed.

<>

<1>Brooks, J.E., Sznyter, L.A.
<2>Method for producing the EagI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 4996151
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the EagI restriction
endonuclease by (1) introducing the restriction endonuclease gene from E. agglomerans ATCC
into a host whereby the restriction gene is expressed; (2) fermenting the host which contains
the vector encoding and expressing the EagI restriction endonuclease and (3) purifying the
EagI restriction endonuclease from the fermented host which contains the vector encoding and
expressing the EagI restriction endonuclease activity.

<>

<1>Brooks, J.E., Sznyter, L.A.
<2>Method for producing the EagI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0343010 B
<5>
<6>1995
<7>The present invention relates to clones for the EagI restriction endonuclease and modification
methylase, and to the production of these enzymes from the clones.

<>

<1>Brooks, S.L., Van Hamme, J.D.
<2>Whole-Genome Shotgun Sequence of Rhodococcus Species Strain JVH1.
<3>J. Bacteriol.
<4>194
<5>5492-5493
<6>2012
<7>Here we present a whole-genome shotgun sequence of Rhodococcus species strain JVH1, an
organism capable of degrading a variety of organosulfur compounds. In
particular, JVH1 is able to selectively cleave carbon-sulfur bonds within alkyl
chains. A large number of oxygenases were identified, consistent with other
members of the genus.

<>

<1>Brouwer, M.S., Allan, E., Mullany, P., Roberts, A.P.
<2>Draft Genome Sequence of the Nontoxigenic Clostridium difficile Strain CD37.
<3>J. Bacteriol.
<4>194
<5>2125-2126
<6>2012
<7>Here we report the draft genome sequence of Clostridium difficile strain CD37, the first
nontoxigenic strain sequenced. Every sequenced strain of Clostridium
difficile has been shown to contain multiple different mobile genetic elements.
The draft genome sequence of strain CD37 reveals the presence of two putative
conjugative transposons.

<>

<1>Brown, C.J., Sen, D., Yano, H., Bauer, M.L., Rogers, L.M., Van der Auwera, G.A., Top, E.M.
<2>Diverse broad-host-range plasmids from freshwater carry few accessory genes.
<3>Appl. Environ. Microbiol.
<4>79
<5>7684-7695
<6>2013
<7>Broad-host-range self-transferable plasmids are known to facilitate bacterial
adaptation by spreading genes between phylogenetically distinct hosts. These
plasmids typically have a conserved backbone region and a variable accessory
region that encodes host-beneficial traits. We do not know, however, how well
plasmids that do not encode accessory functions can survive in nature. The goal
of this study was to characterize the backbone and accessory gene content of
plasmids that were captured from freshwater sources without selecting for a
particular phenotype or cultivating their host. To do this, triparental matings
were used such that the only required phenotype was the plasmid's ability to
mobilize a nonconjugative plasmid. Based on complete genome sequences of 10
plasmids, only 5 carried identifiable accessory gene regions, and none carried
antibiotic resistance genes. The plasmids belong to four known incompatibility
groups (IncN, IncP-1, IncU, and IncW) and two potentially new groups. Eight of
the plasmids were shown to have a broad host range, being able to transfer into
alpha-, beta-, and gammaproteobacteria. Because of the absence of antibiotic
resistance genes, we resampled one of the sites and compared the proportion of
captured plasmids that conferred antibiotic resistance to their hosts with the
proportion of such plasmids captured from the effluent of a local wastewater
treatment plant. Few of the captured plasmids from either site encoded antibiotic
resistance. A high diversity of plasmids that encode no or unknown accessory
functions is thus readily found in freshwater habitats. The question remains how
the plasmids persist in these microbial communities.

<>

<1>Brown, D.R., May, M., Michaels, D.L., Barbet, A.F.
<2>Genome Annotation of Five Mycoplasma canis Strains.
<3>J. Bacteriol.
<4>194
<5>4138-4139
<6>2012
<7>To understand its potential to cause invasive disease, the genome of Mycoplasma canis strain
PG14(T) from a dog's throat was compared to those of isolates from
the genital tract or brain of dogs. The average nucleotide identity between
strain pairs is 98%, and their genome annotations are similar.

<>

<1>Brown, F.L., Musich, P.R., Maio, J.
<2>Cae I: an endonuclease isolated from the African green monkey with properties indicating site-specific cleavage of homologous and heterologous mammalian DNA.
<3>Nucleic Acids Res.
<4>5
<5>1093-1107
<6>1978
<7>Component a DNA is a highly repetitive sequence that comprises nearly a quarter of the African
green monkey (Ceropithecus aethiops) genome.  A previous microbial restriction enzyme analysis
showed that the repeat structure of component a DNA is based upon a nonomeric unit of 176  - 4
base-pairs.  An endonuclease, provisionally termed CaeI, has been isolated from African green
monkey teste that cleaves component a DNA into multimeric segments based upon the same repeat
periodicity as that revealed by microbial restriction enzymes. The primary sites of CaeI
cleavage in the component a sequence appear to be 120 +- 6 base-pairs distant from the HindIII
sites and 73 +- 6 base-pairs distant from the EcoRI* sites.  CaeI has been partially
characterized with special reference to the effects of ATP and S-adenosylmethionine on the
cleavage of component a DNA.  CaeI may be a member of a class of similar site-specific
nucleases present in mammalian cells. CaeI also cleave mouse satellite DNA into a multimeric
series of discrete segments: the periodicty of this series is shorter than that revealed by
EcoRII restriction analysis of mouse satellite DNA.

<>

<1>Brown, L.M., Gunasekera, T.S., Ruiz, O.N.
<2>Draft Genome Sequence of Pseudomonas aeruginosa ATCC 33988, a Bacterium Highly Adapted to Fuel-Polluted Environments.
<3>Genome Announcements
<4>2
<5>e01113-14
<6>2014
<7>Pseudomonas aeruginosa ATCC 33988 is highly adapted to grow in jet and diesel fuel, with a
defined regulation of adaptive genes and metabolization of
n-alkanes. The draft genome of strain ATCC 33988 is 6.4 Mb in size, with 5,975
coding sequences and 66.3% G+C content, and it is highly similar to that of the
clinical strain P. aeruginosa PAO1.

<>

<1>Brown, N.L.
<2>Sequence determination of restriction-endonuclease recognition sites.
<3>Biochem. Soc. Trans.
<4>8
<5>398-399
<6>1980
<7>Restriction endonucleases are widely used in the study of the physical structure of DNA
molecules, and in the genetic manipulation of DNA molecules in vitro.  They also provided
interesting and useful model systems for the study of protein-DNA interactions.  A knowledge
of the DNA sequence that is recognized by a restriction endonuclease, and of the structure of
the termini of the DNA fragments, is essential to the prediction of the uses to which the
enzyme may be put.

<>

<1>Brown, N.L., Hutchison, C.A. III, Smith, M.
<2>The specific non-symmetrical sequence recognized by restriction endonuclease MboII.
<3>J. Mol. Biol.
<4>140
<5>143-148
<6>1980
<7>The restriction endonuclease MboII, isolated from Moraxella bovis (ATCC 10900),
cleaves bacteriophage PhiX174am3 replicative form I DNA into ten fragments.
The physical map of these fragments has been aligned with the sequence of
PhiX174 DNA.  There is no sequence with 2-fold rotational symmetry common to
the region of all ten cleavage sites.  However, the non-symmetrical sequence
5'-G-A-A-G-A-3' 3'-C-T-T-C-T-5' occurs near to each cleavage site.  Precise
mapping of the cleavages in both DNA strands at several sites places the cuts
eight nucleotides to the right of the upper sequence and seven nucleotides to
the right of the lower sequence.

<>

<1>Brown, N.L., McClelland, M., Whitehead, P.R.
<2>HgiAI: A restriction endonuclease from Herpetosiphon giganteus HP1023.
<3>Gene
<4>9
<5>49-68
<6>1980
<7>A new class II restriction endonuclease, HgiAI has been partially purified from
Herpetosiphon giganteus HP1023.  The enzyme activity has been characterized and
shown to recognize the family of related hexanucleotide sequences 5'-G
(A/T)-G-C-(A/T)^C-3' 3'-C^(A/T)-C-G-(A/T)-G-5' where the second and fifth
nucleotide pairs are A:T pairs in either orientation.  Cleavage occurs as
shown, to give DNA fragments with 3'-terminal tetranucleotide extensions.  The
recognition sites of the enzymes SacI and SStI (GAGCT^C) form a subset of the
recognition site of HgiAI.  One of the four possible tetranucleotide
3'-extenions (cohesive ends), generated by HgiAI is identical with those
generated by SacI and SstI, another is identical with that of PstI.  HgiAI
should be useful for molecular cloning.

<>

<1>Brown, N.L., Smith, M.
<2>A general method for defining restriction enzyme cleavage and recognition sites.
<3>Methods Enzymol.
<4>65
<5>391-404
<6>1980
<7>Class II restriction endonucleases cleave double-stranded DNA into specific
fragments.  These enzymes are powerful tools for the dissection of DNA, and
they are essential for the construction of recombinant DNA molecules and for
DNA sequence determination.

<>

<1>Brown, N.L., Smith, M.
<2>The mapping and sequence determination of the single site in PhiX174am3 replicative form DNA cleaved by restriction endonuclease PstI.
<3>FEBS Lett.
<4>65
<5>284-287
<6>1976
<7>A large number of restriction endonucleases have now been isolated from
prokaryotes.  We have tested some of these enzymes on the covalently-closed
replicative form (RFI) of PhiX174am3 DNA.  The restriction endonuclease from
Providencia stuarti 164 (PstI) was found to convert RFI DNA to a linear form of
apparent unit length. In this paper we describe the mapping of the single PstI
site in PhiX174am3 RFI DNA and the determination of the sequence cleaved.  This
is the symmetrical hexanucleotide sequence: 5'-C-T-G-C-A-^G-3'
3'-G-^A-C-G-T-C-5' The methods used to determine the sequence recognised by
PstI and the site of cleavage within that sequence are novel, and should be of
general use.

<>

<1>Brown, N.L., Smith, M.
<2>Cleavage specificity of the restriction endonuclease isolated from Haemophilus gallinarum (HgaI).
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>74
<5>3213-3216
<6>1977
<7>The nucleotide sequences in the replicative form (duplex) of PhiX174 DNA around
six sites cut by HgaI, a restriction endonuclease from Haemophilus gallinarum,
have been compared.  The enzyme produces a staggered cleavage resulting in a
pentanucleotide 5'-terminal extension.  The sequences within and immediately
surrounding the pentanucleotide cleavage site have no obvious relationship.
However, the sequence 5'-G-A-C-G-C-3' 3'-C-T-G-C-G-5' occurs five nucleotide
pairs to the left of the cut in the upper strand and 10 nucleotide pairs to the
left of the cut in the lower strand and, therefore, is believed to constitute
the recognition in which recognition and cleavage sites lack 2-fold rotational
symmetry.  The method used to define the cleavage site is of general
applicability.

<>

<1>Brown, N.L., Walsby, A.E.
<2>Type II restriction endonuclease DsaI with process for obtaining it and the use thereof.
<3>US Patent Office
<4>US 4871664
<5>
<6>1989
<7>The present invention provides a type II restriction endonuclease isolated from
Dactylococcopsis salina. The endonuclease is known as DsaI, and recognizes the following
nucleotide sequence and has a cleavage point indicated by the slash: 5' C/CPuPyGG 3' 3'
GGPyPuC/C 5'. Also described is a process for obtaining DsaI from D. salina.

<>

<1>Brown, N.P., Sander, C., Bork, P.
<2>Frame: detection of genomic sequencing errors.
<3>Bioinformatics
<4>14
<5>367-371
<6>1998
<7>Motivation: The underlying error rate for genomic sequencing sometimes results in the
introduction of artificial frameshifts and in-frame stop codons into putative protein encoding
genes.  Severe errors are then introduced into the inferred transcripts through
mis-translation or premature termination.  Results: We describe a system for screening
segments of DNA for frameshift and in-frame stop errors in coding regions.  The method is
based on homology matching using blastx to compare all six reading frames of the query
nucleotide sequence against selected protein sequence databases.  Fragments of protein
matching neighboring regions of the query DNA are united and extended laterally to define
candidate open reading frames, within which, frameshifts and stops are identified.  Suitable
targets include prokaryotic or other intron-free genomic sequence and complementary DNA's.
As an example of its use, we report here two frameshifted ORFs that deviate from the original
TIGR sequence annotations for the recently released Helicobacter pylori genome.  Availability:
The tool is accessible via the URL http://www.sander.ebi.ac.uk/frame/.  Contact:
brown@ebi.ac.uk.

<>

<1>Brown, S.D. et al.
<2>Genome Sequence of the Mercury-Methylating Strain Desulfovibrio desulfuricans ND132.
<3>J. Bacteriol.
<4>193
<5>2078-2079
<6>2011
<7>Desulfovibrio desulfuricans strain ND132 is an anaerobic sulfate-reducing bacterium (SRB)
capable of producing methylmercury (MeHg), a potent human
neurotoxin. The mechanism of methylation by this and other organisms is
unknown. We present the 3.8-Mb genome sequence to provide further insight
into microbial mercury methylation.

<>

<1>Brown, S.D., Begemann, M.B., Mormile, M.R., Wall, J.D., Han, C.S., Goodwin, L.A., Pitluck, S., Land, M.L., Hauser, L.J., Elias, D.A.
<2>Complete Genome Sequence of the Haloalkaliphilic, Hydrogen-Producing bacterium Halanaerobium hydrogenoformans.
<3>J. Bacteriol.
<4>193
<5>3682-3683
<6>2011
<7>Halanaerobium hydrogenoformans is an alkaliphilic bacterium capable of biohydrogen production
at pH 11 and 7% (w/v) salt. We present the 2.6 Mb
genome sequence to provide insights into its physiology and potential for
bioenergy applications.

<>

<1>Brown, S.D., Hurt, R.A.Jr., Gilmour, C.C., Elias, D.A.
<2>Draft genome sequences for three mercury-methylating, sulfate-reducing bacteria.
<3>Genome Announcements
<4>1
<5>e00618-13
<6>2013
<7>The genetic basis for bacterial mercury methylation has been described recently.  For insights
into the physiology of mercury-methylating bacteria, we present
genome sequences for Desulfococcus multivorans strain DSM 2059, Desulfovibrio
alkalitolerans strain DSM 16529, and Desulfovibrio species strain X2.

<>

<1>Brown, S.D., Klingeman, D.M., Lu, T.Y., Johnson, C.M., Utturkar, S.M., Land, M.L., Schadt, C.W., Doktycz, M.J., Pelletier, D.A.
<2>Draft Genome Sequence of Rhizobium sp. Strain PDO1-076, a Bacterium Isolated from Populus deltoides.
<3>J. Bacteriol.
<4>194
<5>2383-2384
<6>2012
<7>Rhizobium sp. strain PDO1-076 is a plant-associated bacterium isolated from Populus deltoides,
and its draft genome sequence is reported.

<>

<1>Brown, S.D., Lamed, R., Morag, E., Borovok, I., Shoham, Y., Klingeman, D.M., Johnson, C.M., Yang, Z., Land, M.L., Utturkar, S.M., Keller, M., Bayer, E.A.
<2>Draft Genome Sequences for Clostridium thermocellum Wild-Type Strain YS and Derived Cellulose Adhesion-Defective Mutant Strain AD2.
<3>J. Bacteriol.
<4>194
<5>3290-3291
<6>2012
<7>Clostridium thermocellum wild-type strain YS is an anaerobic, thermophilic, cellulolytic
bacterium capable of directly converting cellulosic substrates into
ethanol. Strain YS and a derived cellulose adhesion-defective mutant strain, AD2,
played pivotal roles in describing the original cellulosome concept. We present
their draft genome sequences.

<>

<1>Brown, S.D., Palumbo, A.V., Panikov, N., Ariyawansa, T., Klingeman, D.M., Johnson, C.M., Land, M.L., Utturkar, S.M., Epstein, S.S.
<2>Draft Genome Sequence for Microbacterium laevaniformans Strain OR221, a Bacterium Tolerant to Metals, Nitrate, and Low pH.
<3>J. Bacteriol.
<4>194
<5>3279-3280
<6>2012
<7>Microbacterium laevaniformans strain OR221 was isolated from subsurface sediments obtained
from the Field Research Center (FRC) in Oak Ridge, TN. It was
characterized as a bacterium tolerant to heavy metals, such as uranium, nickel,
cobalt, and cadmium, as well as nitrate and low pH. We present its draft genome
sequence.

<>

<1>Brown, S.D., Podar, M., Klingeman, D.M., Johnson, C.M., Yang, Z.K., Utturkar, S.M., Land, M.L., Mosher, J.J., Hurt, R.A.Jr., Phelps, T.J., Palumbo, A.V., Arkin, A.P., Hazen, T.C., Elias, D.A.
<2>Draft Genome Sequences for Two Metal-Reducing Pelosinus fermentans Strains Isolated from a Cr(VI)-Contaminated Site and for Type Strain R7.
<3>J. Bacteriol.
<4>194
<5>5147-5148
<6>2012
<7>Pelosinus fermentans 16S rRNA gene sequences have been reported from diverse geographical
sites since the recent isolation of the type strain. We present the
genome sequence of the P. fermentans type strain R7 (DSM 17108) and genome
sequences for two new strains with different abilities to reduce iron, chromate,
and uranium.

<>

<1>Brown, S.D., Utturkar, S.M., Arkin, A.P., Deutschbauer, A.M., Elias, D.A., Hazen, T.C., Chakraborty, R.
<2>Draft Genome Sequence for Desulfovibrio africanus Strain PCS.
<3>Genome Announcements
<4>1
<5>e00144-13
<6>2013
<7>Desulfovibrio africanus strain PCS is an anaerobic sulfate-reducing bacterium (SRB) isolated
from sediment from Paleta Creek, San Diego, CA. Strain PCS is
capable of reducing metals such as Fe(III) and Cr(VI), has a cell cycle, and is
predicted to produce methylmercury. We present the D. africanus PCS genome
sequence.

<>

<1>Brown, S.D., Utturkar, S.M., Klingeman, D.M., Johnson, C.M., Martin, S.L., Land, M.L., Lu, T.Y., Schadt, C.W., Doktycz, M.J., Pelletier, D.A.
<2>Twenty-One Genome Sequences from Pseudomonas Species and 19 Genome Sequences from Diverse Bacteria Isolated from the Rhizosphere and Endosphere of Populus  deltoides.
<3>J. Bacteriol.
<4>194
<5>5991-5993
<6>2012
<7>To aid in the investigation of the Populus deltoides microbiome, we generated draft genome
sequences for 21 Pseudomonas strains and 19 other diverse bacteria
isolated from Populus deltoides roots. Genome sequences for isolates similar to
Acidovorax, Bradyrhizobium, Brevibacillus, Caulobacter, Chryseobacterium,
Flavobacterium, Herbaspirillum, Novosphingobium, Pantoea, Phyllobacterium,
Polaromonas, Rhizobium, Sphingobium, and Variovorax were generated.

<>

<1>Brown, S.D., Utturkar, S.M., Magnuson, T.S., Ray, A.E., Poole, F.L., Lancaster, W.A., Thorgersen, M.P., Adams, M.W., Elias, D.A.
<2>Complete Genome Sequence of Pelosinus sp. Strain UFO1 Assembled Using Single-Molecule Real-Time DNA Sequencing Technology.
<3>Genome Announcements
<4>2
<5>e00881-14
<6>2014
<7>Pelosinus species can reduce metals such as Fe(III), U(VI), and Cr(VI) and have been isolated
from diverse geographical regions. Five draft genome sequences have
been published. We report the complete genome sequence for Pelosinus sp. strain
UFO1 using only PacBio DNA sequence data and without manual finishing.

<>

<1>Brown, S.D., Wall, J.D., Kucken, A.M., Gilmour, C.C., Podar, M., Brandt, C.C., Teshima, H., Detter, J.C., Han, C.S., Land, M.L., Lucas, S., Han, J., Pennacchio, L., Nolan, M., Pitluck, S., Woyke, T., Goodwin, L., Palumbo, A.V., Elias, D.A.
<2>Genome Sequence of the Mercury-Methylating and Pleomorphic Desulfovibrio africanus strain Walvis Bay.
<3>J. Bacteriol.
<4>193
<5>4037-4038
<6>2011
<7>Desulfovibrio africanus strain Walvis Bay is an anaerobic sulfate-reducing bacterium (SRB)
capable of producing methylmercury (MeHg), a potent human
neurotoxin. The mechanism of methylation by this and other organisms is
unknown. We present the 4.2 Mb genome sequence to provide further insight
into microbial mercury methylation and sulfate-reducing bacteria.

<>

<1>Brown, T.A.
<2>Restriction and modification.
<3>Essential Molecular Biology
<4>2
<5>260-284
<6>1991
<7>*
1. Restriction endonucleases. Almost 1500 restriction endonucleases are now known and over 150
are commercially-available. Complete lists plus details of restriction sites and reaction
conditions have been published.

2. Site-specific methylases. At the last count 117 site-specific methylases had been
characterized. A full list including recognition sequences and other relevant information is
available.

3. Restriction fragment patterns. Restriction fragments are frequently used as size markers
for agarose and polyacrylamide gel electrophoresis. Figures 1-6 are restriction endonuclease
digests that have been computer-generated assuming that all digests are run in 1.4% agarose.
The scale on the left is in base-pairs.


<>

<1>Brown, W.M., Vinograd, J.
<2>Restriction endonuclease cleavage maps of animal mitochondrial DNAs.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>71
<5>4617-4621
<6>1974
<7>The restriction endonuclease, HindIII, gives rise to three fragments in each of
the three mitochondrial DNAs isolated from the established mammalian cell lines
LA9 (mouse), HeLa (human), and BSC-1 (African green monkey).  The restriction
endonuclease, EcoRI gives rise to three fragments in mitochondrial DNA from
HeLa and to two in DNAs form LA9 and BSC-1.  The sizes and the orders of the
fragments in the respective genomes have been determined with data obtained
from the electron microscope.  The origin and the direction of replication have
been designated in each of the cleavage maps.  Polyacrylamide gel
electrophoretic analyses demonstrated that additional fragments not detectable
in the electron microscope and larger than 50 nucleotide pairs were not
present.

<>

<1>Brownlee, C.
<2>Danna and Nathans: Restriction enzymes and the boon to modern molecular biology.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>5909
<6>2005
<7>In 1971, a paper published in PNAS helped jump-start the era of modern molecular biology and
biotechnology, eventually giving rise to many of the genetic advances that seem so commonplace
today.  The article, written by Academy member Daniel Nathans and his then graduate student,
Kathleen Danna, exposed the marvelous utility of restriction enzymes.  In the accompanying
Perspective highlighting this classic work of scientific literature, Rich Roberts provides a
historial account of the scientific discoveries leading up to the PNAS paper and the
unparalleled scientific advances made after its publication.

<>

<1>Bruce, T., Leite, F.G., Tschoeke, D.A., Miranda, M., Pereira, N.Jr., Valle, R., Thompson, C.C., Thompson, F.L.
<2>Exploring the Genome of a Butyric Acid Producer, Clostridium butyricum INCQS635.
<3>Genome Announcements
<4>2
<5>e01169-14
<6>2014
<7>The draft genome sequence of Clostridium butyricum INCQS635 was obtained by means of ion
sequencing. The genome provides further insight into the genetic
repertoire involved with metabolic pathways related to the fermentation of
different compounds and organic solvents synthesis (i.e., butyric acid) with
biofuel applications.

<>

<1>Brueckner, B., Kuck, D., Lyko, F.
<2>DNA methyltransferase inhibitors for cancer therapy.
<3>Cancer J.
<4>13
<5>17-22
<6>2007
<7>Aberrant DNA methylation patterns, including hypermethylation of tumor suppressor genes, have
been described in many human cancers. These epigenetic mutations can be reversed by DNA
methyltransferase inhibitors, which provide novel opportunities for cancer therapy. Clinical
concepts for epigenetic therapies are currently being developed by using azanucleosides for
the treatment of leukemias and other tumors. These trials will greatly benefit from the
inclusion of molecular markers for monitoring epigenetic changes in patients and for
maximizing biologic responses. In addition, novel inhibitors need to be developed that result
in a direct and specific inhibition of DNA methyltransferase activity. Several recent
developments indicate that rational design of small molecule DNA methyltransferase inhibitors
is feasible and that this approach can result in the establishment of novel drug candidates.
The use of novel DNA methyltransferase inhibitors in clinical trials that allow monitoring of
drug-induced DNA methylation changes should provide the foundation for improved epigenetic
cancer therapies.

<>

<1>Bruggemann, H., Baumer, S., Fricke, W.F., Wiezer, A., Liesegang, H., Decker, I., Herzberg, C., Martinez-Arias, R., Merkl, R., Henne, A., Gottschalk, G.
<2>The genome sequence of Clostridium tetani, the causative agent of tetanus disease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>1316-1321
<6>2003
<7>Tetanus disease is one of the most dramatic and globally prevalent diseases of humans and
vertebrate animals, and has been reported for over
24 centuries. The manifestation of the disease, spastic paralysis, is
caused by the second most poisonous substance known, the tetanus toxin,
with a human lethal dose of approximately 1 ng/kg. Fortunately, this
disease is successfully controlled through immunization with tetanus
toxoid; nevertheless, according to the World Health Organization, an
estimated 400,000 cases still occur each year, mainly of neonatal tetanus.
The causative agent of tetanus disease is Clostridium tetani, an anaerobic
spore-forming bacterium, whose natural habitat is soil, dust, and
intestinal tracts of various animals. Here we report the complete genome
sequence of toxigenic C. tetani E88, a variant of strain Massachusetts.
The genome consists of a 2,799,250-bp chromosome encoding 2,372 ORFs. The
tetanus toxin and a collagenase are encoded on a 74,082-bp plasmid,
containing 61 ORFs. Additional virulence-related factors could be
identified, such as an array of surface-layer and adhesion proteins (35
ORFs), some of them unique to C. tetani. Comparative genomics with the
genomes of Clostridium perfringens, the causative agent of gas gangrene,
and Clostridium acetobutylicum, a nonpathogenic solvent producer, revealed
a remarkable capacity of C. tetani: The organism can rely on an extensive
sodium ion bioenergetics. Additional candidate genes involved in the
establishment and maintenance of a pathogenic lifestyle of C. tetani are
presented.

<>

<1>Bruggemann, H., Henne, A., Hoster, F., Liesegang, H., Wiezer, A., Strittmatter, A., Hujer, S., Durre, P., Gottschalk, G.
<2>The Complete Genome Sequence of Propionibacterium Acnes, a Commensal of Human Skin.
<3>Science
<4>305
<5>671-673
<6>2004
<7>Propionibacterium acnes is a major inhabitant of adult human skin, where it resides within
sebaceous follicles, usually as a harmless commensal although it has been implicated in acne
vulgaris formation. The entire genome sequence of this Gram positive bacterium encodes 2333
putative genes and revealed numerous gene products involved in degrading host molecules,
including sialidases, neuraminidases, endoglycoceramidases, lipases, and pore forming factors.
Surface associated and other immunogenic factors have been identified, which might be involved
in triggering acne inflammation and other P. acnes associated diseases.

<>

<1>Brummet, S.R., Robb, F.T., Borges, K.M., Hujer, K.M., Domke, S.T.
<2>Proteins from Pyrococcus furiosus.
<3>US Patent Office
<4>US 5719056
<5>
<6>1998
<7>Purified nucleic acid comprising a nucleotide base sequence selected from sequence ID NOS.:
1-9.

<>

<1>Brunder, W., Karch, H., Schmidt, H.
<2>Complete sequence of the large virulence plasmid pSFO157 of the sorbitol-fermenting enterohemorrhagic Escherichia coli O157:H(-) strain 3072/96.
<3>Int. J. Med. Microbiol.
<4>296
<5>467-474
<6>2006
<7>The large virulence plasmid pSFO157 of sorbitol-fermenting E. coli
O157:H(-) strain 3072/96 has a size of 121,239bp and contains 96 open
reading frames >50bp. It is therefore 29,162bp larger (ca. 32%) than
plasmid pO157 of E. coli O157:H7 strain EDL933. Major differences between
the plasmids are the absence of katP, espP, and toxB in pSFO157 and,
instead of these, the presence of the sfp fimbriae gene cluster and a
large part of an F-plasmid transfer region, the latter accounting for most
of the additional DNA. The differences in the order of the genes and their
composition, as well as the presence of a number of replication-associated
genes and mobile genetic elements suggests that the large E. coli O157
virulence plasmids have a complex evolutionary origin.

<>

<1>Brunet-Galmes, I., Busquets, A., Pena, A., Gomila, M., Nogales, B., Garcia-Valdes, E., Lalucat, J., Bennasar, A., Bosch, R.
<2>Complete Genome Sequence of the Naphthalene-Degrading Bacterium Pseudomonas stutzeri AN10 (CCUG 29243).
<3>J. Bacteriol.
<4>194
<5>6642-6643
<6>2012
<7>Pseudomonas stutzeri AN10 (CCUG 29243) can be considered a model strain for aerobic
naphthalene degradation. We report the complete genome sequence of this
bacterium. Its 4.71-Mb chromosome provides insights into other biodegradative
capabilities of strain AN10 (i.e., benzoate catabolism) and suggests a high
number of horizontal gene transfer events.

<>

<1>Bruno-Barcena, J.M., Chinn, M.S., Grunden, A.M.
<2>Genome Sequence of the Autotrophic Acetogen Clostridium autoethanogenum JA1-1 Strain DSM 10061, a Producer of Ethanol from Carbon Monoxide.
<3>Genome Announcements
<4>1
<5>e00628-13
<6>2013
<7>Clostridium autoethanogenum is an anaerobic, autotrophic acetogen that is capable of
converting CO and CO2 into ethanol and acetate. Here we report the draft
genome sequence of C. autoethanogenum JA1-1 strain DSM 10061 (4.5 Mbp; G+C
content, 37.5%) and the findings obtained from annotation of the genome sequence.

<>

<1>Bryanskaya, A.V., Rozanov, A.S., Logacheva, M.D., Kotenko, A.V., Peltek, S.E.
<2>Draft Genome Sequence of Geobacillus icigianus Strain G1w1T Isolated from Hot Springs in the Valley of Geysers, Kamchatka (Russian Federation).
<3>Genome Announcements
<4>2
<5>e01098-14
<6>2014
<7>The Geobacillus icigianus G1w1(T) strain was isolated from sludge samples of unnamed vaporing
hydrothermal (97 degrees capital ES, Cyrillic) outlets situated
in a geyser in the Troinoy region (Valley of Geysers, Kronotsky Nature Reserve,
Kamchatka, Russian Federation; 54 degrees 25'51.40'N, 160 degrees 7'41.40'E).
The sequenced and annotated genome is 3,457,810 bp and encodes 3,342 genes.

<>

<1>Bryant, D.A., Costas, A.M., Maresca, J.A., Chew, A.G., Klatt, C.G., Bateson, M.M., Tallon, L.J., Hostetler, J., Nelson, W.C., Heidelberg, J.F., Ward, D.M.
<2>Candidatus Chloracidobacterium thermophilum: an aerobic phototrophic Acidobacterium.
<3>Science
<4>317
<5>523-526
<6>2007
<7>Only five bacterial phyla with members capable of chlorophyll (Chl)-based
phototrophy are presently known. Metagenomic data from the phototrophic
microbial mats of alkaline siliceous hot springs in Yellowstone National
Park revealed the existence of a distinctive bacteriochlorophyll
(BChl)-synthesizing, phototrophic bacterium. A highly enriched culture of
this bacterium grew photoheterotrophically, synthesized BChls a and c
under oxic conditions, and had chlorosomes and type 1 reaction centers.
"Candidatus Chloracidobacterium thermophilum" is a BChl-producing member
of the poorly characterized phylum Acidobacteria.

<>

<1>Bryk, M., Belisle, M., Mueller, J.E., Belfort, M.
<2>Selection of a remote cleavage site by I-TevI, the td intron-encoded endonuclease.
<3>J. Mol. Biol.
<4>247
<5>197-210
<6>1995
<7>I-TevI, a double-strand DNA endonuclease involved in the mobility of the td intron of phage
T4, is highly unusual in that it binds and cleaves intronless td alleles (td homing sites) in
a site-specific but sequence-tolerant manner. The endonuclease binds to sequences flanking the
intron insertion site and near the remote cleavage site, located 23 and 25 nucleotides away on
the top and bottom strands, respectively. Mapping studies indicate that I-TevI has both
sequence and distance sensors that function during cut-site selection. Although I-TevI
cleavage of many insertion and deletion variants of the homing site is impaired, double-strand
breaks are generated at positions that collectively span two turns of the helix, indicating
that the interaction is extraordinarily flexible. However, the endonuclease does exhibit
spacing preferences between its binding domains, and sequence preferences near the cleavage
site, with the G:C pair at -23 implicated as a cleavage determinant. Furthermore, I-TevI
appears to function through interactions across the minor groove at the cleavage site, as it
does at the intron insertion site, and to be capable of cleaving sequentially, first on the
bottom and then on the top strand. These properties of I-TevI are incorporated in a model
wherein the endonuclease effects distant cleavage via a flexible hinge.

<>

<1>Bryk, M., Quirk, S.M., Mueller, J.E., Loizos, N., Lawrence, C., Belfort, M.
<2>The td intron endonuclease I-TevI makes extensive sequence-tolerant contacts across the minor groove of its DNA target.
<3>EMBO J.
<4>12
<5>2141-2149
<6>1993
<7>I-TevI, a double-strand DNA endonuclease encoded by the mobile td intron of phage T4, has
specificity for the intronless td allele. Genetic and physical studies indicate that the
enzyme makes extensive contact with its DNA substrate over at least three helical turns and
around the circumference of the helix. Remarkably, no single nucleotide within a 48 bp region
encompassing this interaction domain is essential for cleavage. Although two subdomains (DI
and DII) contain preferred sequences, a third domain (DIII), a primary region of contact with
the enzyme, displays much lower sequence preference. While DII and DIII suffice for
recognition and binding of I-TevI, all three domains are important for formation of a
cleavage-competent complex. Mutational, footprinting and interference studies indicate
predominant interactions of I-TevI across the minor groove and phosphate backbone of the DNA.
Contacts appear not to be at the single nucleotide level; rather redundant interactions and/or
structural recognition are implied. These unusual properties provide a basis for understanding
how I-TevI recognizes T-even phage DNA, which is heavily modified in the major groove. These
recognition characteristics may increase the range of natural substrates available to the
endonuclease, thereby extending the invasive potential of the mobile intron.

<>

<1>Brzezinski, R., Piekarowicz, A.
<2>Steps in the reaction mechanism of the Haemophilus influenzae Rf restriction endonuclease.
<3>J. Mol. Biol.
<4>154
<5>615-627
<6>1982
<7>HinfIII is a restriction modification enzyme isolated from Haemophilus
influenzae strain Rf. It requires ATP for cleavage and S-adenosyl-L-methionine
for methylation of DNA.  S-Adenosyl-L-methionine acts as an allosteric effector
in the endonuclease reaction.  The enzyme forms a complex with unmodified DNA
in the absence of ATP and S-adenosyl-L-methionine.  This complex is sensitive
to inhibition by heparin.  S-Adenosyl-L-methionine is required for the
formation of a complex that is insensitive to such inhibition.  ATP acts as an
allosteric effector of HinfIII, and induces DNA cleavage followed probably by
the release of the enzyme from the DNA.

<>

<1>Brzuszkiewicz, E., Bruggemann, H., Liesegang, H., Emmerth, M., Olschlager, T., Nagy, G., Albermann, K., Wagner, C., Buchrieser, C., Emody, L., Gottschalk, G., Hacker, J., Dobrindt, U.
<2>How to become a uropathogen: Comparative genomic analysis of extraintestinal pathogenic Escherichia coli strains.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>12879-12884
<6>2006
<7>Uropathogenic Escherichia coli (UPEC) strain 536 (O6:K15:H31) is one of the model organisms of
extraintestinal pathogenic E. coli (ExPEC). To
analyze this strain's genetic basis of urovirulence, we sequenced the
entire genome and compared the data with the genome sequence of UPEC
strain CFT073 (O6:K2:H1) and to the available genomes of nonpathogenic E.
coli strain MG1655 (K-12) and enterohemorrhagic E. coli. The genome of
strain 536 is approximately 292 kb smaller than that of strain CFT073.
Genomic differences between both UPEC are mainly restricted to large
pathogenicity islands, parts of which are unique to strain 536 or CFT073.
Genome comparison underlines that repeated insertions and deletions in
certain parts of the genome contribute to genome evolution. Furthermore,
427 and 432 genes are only present in strain 536 or in both UPEC,
respectively. The majority of the latter genes is encoded within smaller
horizontally acquired DNA regions scattered all over the genome. Several
of these genes are involved in increasing the pathogens' fitness and
adaptability. Analysis of virulence-associated traits expressed in the two
UPEC O6 strains, together with genome comparison, demonstrate the marked
genetic and phenotypic variability among UPEC. The ability to accumulate
and express a variety of virulence-associated genes distinguishes ExPEC
from many commensals and forms the basis for the individual virulence
potential of ExPEC. Accordingly, instead of a common virulence mechanism,
different ways exist among ExPEC to cause disease.

<>

<1>Brzuszkiewicz, E., Schulz, T., Rydzewski, K., Daniel, R., Gillmaier, N., Dittmann, C., Holland, G., Schunder, E., Lautner, M., Eisenreich, W., Luck, C., Heuner, K.
<2>Legionella oakridgensis ATCC 33761 genome sequence and phenotypic characterization reveals its replication capacity in amoebae.
<3>Int. J. Med. Microbiol.
<4>303
<5>514-528
<6>2013
<7>Legionella oakridgensis is able to cause Legionnaires' disease, but is less
virulent compared to L. pneumophila strains and very rarely associated with human
disease. L. oakridgensis is the only species of the family legionellae which is
able to grow on media without additional cysteine. In contrast to earlier
publications, we found that L. oakridgensis is able to multiply in amoebae. We
sequenced the genome of L. oakridgensis type strain OR-10 (ATCC 33761). The
genome is smaller than the other yet sequenced Legionella genomes and has a
higher G+C-content of 40.9%. L. oakridgensis lacks a flagellum and it also lacks
all genes of the flagellar regulon except of the alternative sigma-28 factor FliA
and the anti-sigma-28 factor FlgM. Genes encoding structural components of type
I, type II, type IV Lvh and type IV Dot/Icm, Sec- and Tat-secretion systems could
be identified. Only a limited set of Dot/Icm effector proteins have been
recognized within the genome sequence of L. oakridgensis. Like in L. pneumophila
strains, various proteins with eukaryotic motifs and eukaryote-like proteins were
detected. We could demonstrate that the Dot/Icm system is essential for
intracellular replication of L. oakridgensis. Furthermore, we identified new
putative virulence factors of Legionella.

<>

<1>Brzuszkiewicz, E., Waschkowitz, T., Wiezer, A., Daniel, R.
<2>Complete Genome Sequence of the B12-Producing Shimwellia blattae Strain DSM 4481, Isolated from a Cockroach.
<3>J. Bacteriol.
<4>194
<5>4436
<6>2012
<7>Here we announce the complete genome sequence of the coenzyme B(12)-producing enteric
bacterium Shimwellia blattae (formerly Escherichia blattae). The genome
consists of a single chromosome (4,158,636 bp). The genome size is smaller than
that of most other enteric bacteria. Genome comparison revealed significant
differences from the Escherichia coli genome.

<>

<1>Bucci, C., Lavitola, A., Salvatore, P., Del Giudice, L., Massardo, D.R., Bruni, C.B., Alifano, P.
<2>Hypermutation in pathogenic bacteria: frequent phase variation in meningococci is a phenotypic trait of a specialized mutator biotype.
<3>Mol. Cell
<4>3
<5>435-445
<6>1999
<7>Expression of serogroup B meningococcal capsular polysaccharide undergoes frequent phase
variation involving reversible frameshift mutations within a homopolymeric repeat in the siaD
gene. A high rate of phase variation is the consequence of a biochemical defect in
methyl-directed mismatch repair. The mutator phenotype is associated to the absence of DNA
adenine methyltransferase (Dam) activity in all pathogenic isolates and in 50% of commensal
strains. Analysis of the meningococcal dam gene region revealed that in all Dam- strains a
gene encoding a putative restriction endonuclease (drg) that cleaves only the methylated DNA
sequence 5'-GmeATC-3' replaced the dam gene. Insertional inactivation of the dam and/or drg
genes indicated that high rates of phase variation and hypermutator phenotype are caused by
absence of a functional dam gene.

<>

<1>Buchrieser, C., Glaser, P., Rusniok, C., Nedjari, H., D'Hauteville, H., Kunst, F., Sansonetti, P., Parsot, C.
<2>The virulence plasmid pWR100 and the repertoire of proteins secreted by the type III secretion apparatus of Shigella flexneri.
<3>Mol. Microbiol.
<4>38
<5>760-771
<6>2000
<7>Bacteria of Shigella spp. are the causative agents of shigellosis. The
virulence traits of these pathogens include their ability to enter into
epithelial cells and induce apoptosis in macrophages. Expression of these
functions requires the Mxi-Spa type III secretion apparatus and the
secreted IpaA-D proteins, all of which are encoded by a virulence plasmid.
In wild-type strains, the activity of the secretion apparatus is tightly
regulated and induced upon contact of bacteria with epithelial cells. To
investigate the repertoire of proteins secreted by Shigella flexneri in
conditions of active secretion, we determined the N-terminal sequence of
14 proteins that are secreted by a mutant in which secretion was
deregulated. Sequencing of the virulence plasmid pWR100 of the S. flexneri
strain M90T (serotype 5) has allowed us to identify the genes encoding
these secreted proteins and suggests that approximately 25 proteins are
secreted by the type III secretion apparatus. Analysis of the G+C content
and the relative positions of genes and open reading frames carried by the
plasmid, together with information concerning the localization and
function of encoded proteins, suggests that pWR100 contains blocks of
genes of various origins, some of which were initially carried by four
different plasmids.

<>

<1>Buddruhs, N., Chertkov, O., Petersen, J., Fiebig, A., Chen, A., Pati, A., Ivanova, N., Lapidus, A., Goodwin, L.A., Chain, P., Detter, J.C., Gronow, S., Kyrpides, N.C., Woyke, T., Goker, M., Brinkhoff, T., Klenk, H.P.
<2>Complete genome sequence of the marine methyl-halide oxidizing Leisingera methylohalidivorans type strain (DSM 14336(T)), a representative of the  Roseobacter clade.
<3>Standards in Genomic Sciences
<4>9
<5>128-141
<6>2013
<7>Leisingera methylohalidivorans Schaefer et al. 2002 emend. Vandecandelaere et al. 2008 is the
type species of the genus Leisingera. The genus belongs to the
Roseobacter clade (Rhodobacteraceae, Alphaproteobacteria), a widely distributed
lineage in marine environments. Leisingera and particularly L.
methylohalidivorans strain MB2(T) is of special interest due to its
methylotrophy. Here we describe the complete genome sequence and annotation of
this bacterium together with previously unreported aspects of its phenotype. The
4,650,996 bp long genome with its 4,515 protein-coding and 81 RNA genes consists
of three replicons, a single chromosome and two extrachromosomal elements with
sizes of 221 kb and 285 kb.

<>

<1>Budker, V.G., Degtyarev, S.K., Sokolov, A.V.
<2>Cleavage of DNA adsorbed on the surface of phospholipid membranes by Type II restriction endonucleases.
<3>Biokhimiia
<4>51
<5>1496-1498
<6>1986
<7>The hydrolysis of phage lambda DNA by restriction endonucleases of type II in the presence of
model membranes from egg phosphatidylcholine was studied.  It was shown that the degree of
hydrolysis of DNA by the enzymes BspI, PstI, and BamHI under these conditions is sharply
reduced.  This is not associated with irreversible inactivation of the enzymes in their
interaction with the membrane.  The most probable explanation for the inhibition of DNA
hydrolysis in the presence of phospholipid vesicles is a change in the substrate properties of
the DNA as a result of its adsorption on the surface of the phospholipid membrane with the
participation of Mg2+ ions.

<>

<1>Budroni, S., Siena, E., Dunning, H.J.C., Seib, K.L., Serruto, D., Nofroni, C., Comanducci, M., Riley, D.R., Daugherty, S.C., Angiuoli, S.V., Covacci, A., Pizza, M., Rappuoli, R., Moxon, E.R., Tettelin, H., Medini, D.
<2>Neisseria meningitidis is structured in clades associated with restriction modification systems that modulate homologous recombination.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>4494-4499
<6>2011
<7>Molecular data on a limited number of chromosomal loci have shown that the population of
Neisseria meningitidis (Nm), a deadly human pathogen, is
structured in distinct lineages. Given that the Nm population undergoes
substantial recombination, the mechanisms resulting in the evolution of
these lineages, their persistence in time, and the implications for the
pathogenicity of the bacterium are not yet completely understood. Based on
whole-genome sequencing, we show that Nm is structured in phylogenetic
clades. Through acquisition of specific genes and through insertions and
rearrangements, each clade has acquired and remodeled specific genomic
tracts, with the potential to impact on the commensal and virulence
behavior of Nm. Despite this clear evidence of a structured population, we
confirm high rates of detectable recombination throughout the whole Nm
chromosome. However, gene conversion events were found to be longer within
clades than between clades, suggesting a DNA cleavage mechanism associated
with the phylogeny of the species. We identify 22 restriction modification
systems, probably acquired by horizontal gene transfer from outside of the
species/genus, whose distribution in the different strains coincides with
the phylogenetic clade structure. We provide evidence that these
clade-associated restriction modification systems generate a differential
barrier to DNA exchange consistent with the observed population structure.
These findings have general implications for the emergence of lineage
structure and virulence in recombining bacterial populations, and they
could provide an evolutionary framework for the population biology of a
number of other bacterial species that show contradictory population
structure and dynamics.

<>

<1>Buell, C.R. et al.
<2>The complete genome sequence of the Arabidopsis and tomato pathogen Pseudomonas syringae pv. tomato DC3000.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>10181-10186
<6>2003
<7>We report the complete genome sequence of the model bacterial pathogen Pseudomonas syringae
pathovar tomato DC3000 (DC3000), which is pathogenic
on tomato and Arabidopsis thaliana. The DC3000 genome (6.5 megabases)
contains a circular chromosome and two plasmids, which collectively encode
5,763 ORFs. We identified 298 established and putative virulence genes,
including several clusters of genes encoding 31 confirmed and 19 predicted
type III secretion system effector proteins. Many of the virulence genes
were members of paralogous families and also were proximal to mobile
elements, which collectively comprise 7% of the DC3000 genome. The
bacterium possesses a large repertoire of transporters for the acquisition
of nutrients, particularly sugars, as well as genes implicated in
attachment to plant surfaces. Over 12% of the genes are dedicated to
regulation, which may reflect the need for rapid adaptation to the diverse
environments encountered during epiphytic growth and pathogenesis.
Comparative analyses confirmed a high degree of similarity with two
sequenced pseudomonads, Pseudomonas putida and Pseudomonas aeruginosa, yet
revealed 1,159 genes unique to DC3000, of which 811 lack a known function.

<>

<1>Buera, M.P., Rossi, S., Moreno, S., Chirife, J.
<2>DSC confirmation that vitrification is not necessary for stabilization of the restriction enzyme EcoRI dried with saccharides.
<3>Biotechnol. Prog.
<4>15
<5>577-579
<6>1999
<7>The glass transition temperature (Tg) of preparations of the restriction enzyme EcoRI,
vacuum-dried in the presence of sucrose, trehalose, or raffinose, was determined using
differential scanning calorimetry.  Tg values were well below those expected for low-moisture
sucrose, trehalose, or raffinose, and this was attributed to the presence of glycerol (a
plasticizer), which was a main component of the restriction enzyme preparation.  This was
verified by determining the glass transition temperature of glycerol, which was found to be
(onset value) -77 C.  Present results confirmed that vitrification (i.e., glass formation) was
not necessary for enzyme protection in present low-moisture saccharide systems.  As shown in
previous work, the enzyme EcoRI was very stable when stored at 37/45 C in spite of the fact
that sugar matrices were completely rubbery, as unequivocally demonstrated in the present
work.

<>

<1>Bugreev, D.V., Nevinsky, G.A.
<2>Possibilities of the method of step-by-step complication of ligand structure in studies of protein-nucleic acid interactions: mechanisms of functioning of some replication, repair, topoisomerization, and restriction enzymes.
<3>Biokhimiia
<4>64
<5>291-305
<6>1999
<7>X-Ray structure analysis is one of the most informative methods for investigation of enzymes.
However, it does not provide quantitative estimation of the relative efficiency of formation
of contacts revealed by this method, and when interpreting the data this does not allow taking
into account the relative contribution of some specific and nonspecific interactions to the
total affinity of nucleic acids (NA) to enzymes. This often results in unjustified
overestimation of the role of specific enzyme--NA contacts in affinity and specificity of
enzyme action. In recent years we have developed new approaches to analysis of the mechanisms
of protein--nucleic acid interactions allowing quantitative estimation of the relative
contribution of virtually every nucleotide unit (including individual structural elements) to
the total affinity of enzymes to long DNA and RNA molecules. It is shown that the interaction
between enzymes and NA on the molecular level can be successfully analyzed by the methods of
synthesis and analysis, that is, step-by-step simplification or complication of the structure
of a long NA-ligand. This approach allows the demonstration that complex formation including
formation of contacts between enzymes and specific NA units can provide neither high affinity
of the enzymes to NA nor the specificity of their action. Using a number of
sequence-independent replication and repair enzymes specifically recognizing a modified unit
in DNA and also some sequence-dependent topoisomerization and restriction enzymes as examples,
it was shown that virtually all nucleotide units within the DNA binding cleft interact with
the enzyme, and high affinity mainly (up to 5-7 of 7-10 orders of magnitude) is provided by
many weak additive interactions between these enzymes and various structural elements of the
individual NA nucleotide units. At the same time, the relative contribution of specific
interactions to the total affinity of NA is rather small and does not exceed 1-2 orders of
magnitude. Specificity of enzyme action is provided by the stages of the enzyme-dependent NA
adaptation to the optimal conformation and directly of catalysis: kcat increases by 3-7 orders
of magnitude when changing from nonspecific to specific NA. In the present work we summarized
our experience in studies of enzymes by the method of step-by-step complication of the ligand
structure and performed a detailed analysis of the features of this approach and its
possibilities for the study of protein--nucleic acid interactions on the molecular level.

<>

<1>Buhk, H.-J., Behrens, B., Tailor, R., Wilke, K., Prada, J.J., Gunthert, U., Noyer-Weidner, M., Jentsch, S., Trautner, T.A.
<2>Restriction and modification in Bacillus subtilis: nucleotide sequence, functional organization and product of the DNA methyltransferase gene of bacteriophage SPR.
<3>Gene
<4>29
<5>51-61
<6>1984
<7>Bacillus subtilis phage SPR codes for a DNA methyltransferase (Mtase) which
methylates the 5' cytosine in the sequence GGCC and both cytosies in the
sequence CCGG.  A 2126-bp fragment of SPR DNA containing the Mtase gene has
been sequenced.  This fragment has only one significant open reading frame of
1347 bp, which corresponds to the Mtase gene.  Within the sequence the Mtase
promoter has been defined by S1 mapping.  The size of the SPR Mtase predicted
from the deduced amino acid composition is 49.9 kDal.  This is in agreement
with both the Mr of the purified enzyme and with that of the SPR Mtase gene
product identified here by minicell technique.  Base changes leading to mutants
affected in Mtase activity were localized within the Mtase gene.

<>

<1>Buhler, R., Yuan, R.
<2>Characterization of a restriction enzyme from Escherichia coli K carrying a mutation in the modification subunit.
<3>J. Biol. Chem.
<4>253
<5>6756-6760
<6>1978
<7>The restriction enzyme from a restriction and modification-deficient strain of
Escherichia coli K mutated in the modification gene (hsdM) has been purified
using an in vitro complementation assay with a mutant restriction enzyme from a
strain lacking only restriction.  The restriction enzyme from the hsdM mutant
lacks all of the activities that are associated with the wild type enzyme:
binding of unmodified DNA to filters, cleavage, or methylation of unmodified
DNA and ATP hydrolysis.  It is shown that the enzyme from this hsdM mutant
cannot bind S-adenosylmethionine, an allosteric effector in the restriction
reaction.  In the absence of enzyme activation by S-adenosylmethionine, no
binding to unmodified DNA takes place.  A comparison with other mutant
restriction enzymes allows us to outline the biochemical role of the subunits
of the E. coli K restriction endonuclease.

<>

<1>Bujnicki, J.M.
<2>Comparison of protein structures reveals monophyletic origin of AdoMet-dependent methyltransferase family and mechanistic convergence rather than recent differentiation of N4-cytosine and N6-adenine DNA methylation.
<3>In Silico Biology
<4>1
<5>175-182
<6>1999
<7>Phylogenetic analysis of the S-adenosyl-L-methionine-dependent methyltransferases was
performed based on similarity of positions of main chain alpha-carbon atoms in published
structures of members of this superfamily.  The evolutionary tree was inferred and the problem
of mono/polyphyletic origin of DNA methyltransferases from the Rossmann-fold enzymes was
solved, bridging two seemingly antithetical hypotheses.  The comparison of protein structures
provides evidence for an evolutionary link between widely diverged subfamilies of RNA and DNA
N6-adenine methyltransferases and argues against the close homology of N6-adenine and
N4-cytosine methyltransferases, apparent from biochemical data and comparison of fragments of
sequences.  Such evolutionary analysis of methyltransferases has never been published yet in
the literature and will guide further phylogenetical studies based on both sequence and
structure comparison.

<>

<1>Bujnicki, J.M.
<2>Phylogeny of the restriction endonuclease-like superfamily inferred from comparison of protein structures.
<3>J. Mol. Evol.
<4>50
<5>39-44
<6>2000
<7>To date all attempts to derive a phyletic relationship among restriction endonucleases
(ENases) from multiple sequence alignments have been limited by extreme divergence of these
enzymes.  Based on the approach of Johnson et al. (1990), I report for the first time the
evolutionary tree of the ENase-like protein superfamily inferred from quantitative comparison
of atomic coordinates of structurally characterized enzymes. The results presented are in
harmony with previous comparisons obtained by crystallographic analyses. It is shown that
lambda-exonuclease initially diverged from the common ancestor and then two "endonucleolytic"
families branched out, separating "blunt end cutters" from "5' four-base overhand cutters."
These data may contribute to a better understanding of ENases and encourage the use of
structure-based methods for inference of phylogenetic relationship among extremely divergent
proteins. In addition, the comparison of three-dimensional structures of ENase-like domains
provides a platform for further clustering analyses of sequence similarities among different
branches of this large protein family, rational choice of homology modeling templates, and
targets for protein engineering.

<>

<1>Bujnicki, J.M.
<2>Homology modelling of the DNA 5mC methyltransferase M.BssHII. Is permutation of functional subdomains common to all subfamilies of DNA methyltransferases?
<3>Int. J. Biol. Macromol.
<4>27
<5>195-204
<6>2000
<7>This work presents a full tertiary model of the M.BssHII methyltransferase (MTase) complexed
with substrate DNA and cofactor
S-adenosyl-L-methionine, built by homology modelling based on
previously solved complete structures of DNA MTases M.HaeIII and
M.HhaI. M.BssHII and the template proteins show high sequence
similarity, which indicates that they are evolutionary related.
However, they are topologically different: M.BssHII is a circularly
permuted variant of template MTases (Xu et al. Nucleic Acids Res.
1997,25:3991). The model developed in this work will be a good starting
point and valuable help in designing mutagenesis experiments to better
understand the biological function of methyltransferases and the
process of domain swapping.

<>

<1>Bujnicki, J.M.
<2>Understanding the evolution of restriction-modification systems: Clues from sequence and structure comparisons.
<3>Acta Biochim. Pol.
<4>48
<5>935-967
<6>2001
<7>Restriction-modification (RM) systems comprise two opposing enzymatic activities: a
restriction endonuclease, that targets specific DNA
sequences and performs endonucleolytic cleavage, and a modification
methyltransferase that renders these sequences resistant to cleavage.
Studies on molecular genetics and biochemistry of RM systems have been
carried out over the past four decades, laying foundations for modern
molecular biology and providing important models for mechanisms of
highly specific protein-DNA interactions. Although the number of known,
relevant sequences 3D structures of RM proteins is growing steadily, we
do not fully understand their functional diversities from an
evolutionary perspective and we are not yet able to engineer new
sequence specificities based on rational approaches. Recent findings on
the evolution of RM systems and on their structures and mechanisms of
action have led to a picture in which conserved modules with defined
function are shared between different RM proteins and other enzymes
involved in nucleic acid biochemistry. On the other hand, it has been
realized that some of the modules have been replaced in the evolution
by unrelated domains exerting similar function. The aim of this review
is to give a survey on the recent progress in the field of structural
phylogeny of RM enzymes with special emphasis on studies of
sequence-structure-function relationships and emerging potential
applications in biotechnology.

<>

<1>Bujnicki, J.M.
<2>A model of structure and action of Sau3AI restriction endonuclease that comprises two MutH-like endonuclease domains within a single polypeptide.
<3>Acta Microbiol. Pol.
<4>50
<5>219-231
<6>2001
<7>Sau3AI is a type II endonuclease that cleaves GATC sequences, producing sticky ends with
4-nucleotide 5'-overhangs.  Its activity is inhibited by cytosine C5-methylation within the
target sequence.  In the N-terminus, Sau3AI exhibits sequence similarity to the GATC-specific
single-strand nicking endonuclease MutH implicated in mismatch repair.  Sequence analysis of
Sau3AI and its homologs reveals that Sau3AI possesses an additional MutH-like domain in the
C-terminus.  Structure prediction suggests that the C-terminal domain lacks the endonuclease
active site but retains all putative DNA-binding elements.  As an illustration of these
findings, a model of quaternary structure of Sau3AI complexed with the target DNA is
presented.  These predictions have implications for evolution, structure and function of
bacterial DNA repair enzymes and restriction endonucleases.

<>

<1>Bujnicki, J.M.
<2>Sequence permutations in the molecular evolution of DNA methyltransferases.
<3>BMC Evol. Biol.
<4>2
<5>3
<6>2002
<7>BACKGROUND: DNA methyltransferases (MTases), unlike MTases acting on other substrates, exhibit
sequence permutation. Based on the sequential order of the cofactor-binding subdomain, the
catalytic subdomain, and the target recognition domain (TRD), several classes of permutants
have been proposed. The majority of known DNA MTases fall into the alpha, beta, and gamma
classes. There is only one member of the zeta class known and no members of the delta and
epsilon classes have been identified to date. Two mechanisms of permutation have been
proposed: one involving gene duplication and in-frame fusion, and the other involving inter-
and intragenic shuffling of gene segments. RESULTS: Two novel cases of sequence permutation in
DNA MTases implicated in restriction-modification systems have been identified, which suggest
that members of the delta and zeta classes (M.MwoI and M.TvoORF1413P, respectively) evolved
from beta-class MTases. This is the first identification of the delta-class MTase and the
second known zeta-class MTase (the first zeta-class member among DNA:m4C and m6A-MTases).
CONCLUSIONS: Fragmentation of a DNA MTase gene may result from attack of nucleases, for
instance when the RM system invades a new cell. Its reassembly into a functional form, the
order of motifs notwithstanding, may be strongly selected for, if the cognate ENase gene
remains active and poses a threat to the host's chromosome. The "cut-and-paste" mechanism is
proposed for beta-delta permutation, which is non-circular and involves relocation of one
segment of a gene. The circular beta-zeta permutation may be explained both by gene
duplication or shuffling of gene fragments. These two mechanisms are not mutually exclusive
and probably both played a role in the evolution of permuted DNA MTases.

<>

<1>Bujnicki, J.M.
<2>Crystallographic and bioinformatic studies on restriction endonucleases: Inference of evolutionary relationships in the 'midnight zone' of homology.
<3>Curr. Protein Pept. Sci.
<4>4
<5>327-337
<6>2003
<7>Type II restriction endonucleases (ENases) cleave DNA with remarkable sequence specificity.
Their discovery in 1970 and studies on molecular
genetics and biochemistry carried out over the past four decades laid
foundations for recombinant DNA techniques. Today, restriction enzymes are
indispensable tools in molecular biology and molecular medicine and a
paradigm for proteins that specifically interact with DNA as well as a
challenging target for protein engineering. The
sequence-structure-function relationships for these proteins are therefore
of central interest in biotechnology. However, among numerous ENase
sequences, only a few exhibit statistically significant similarity in
pairwise comparisons, which was initially interpreted as evidence for the
lack of common origin. Nevertheless, X-ray crystallographic studies of
seemingly dissimilar type II ENases demonstrated that they share a common
structural core and metal-binding/catalytic site, arguing for extreme
divergence rather than independent evolution. A similar nuclease domain
has been also identified in various enzymes implicated in DNA repair and
recombination. Ironically, following the series of crystallographic
studies suggesting homology of all type II ENases, bioinformatic studies
provided evidence that some restriction enzymes are in fact diverged
members of unrelated nuclease superfamilies: Nuc, HNH and GIY-YIG. Hence,
the restriction enzymes as a whole, represent a group of functionally
similar proteins, which evolved on multiple occasions and subsequently
diverged into the "midnight zone" of homology, where common origins within
particular groups can be inferred only from structure-guided comparisons.
The structure-guided approaches used for this purpose include:
identification of functionally important residues using superposition of
atomic coordinates, alignment of sequence profiles enhanced by secondary
structures, fold recognition, and homology modeling. This review covers
recent results of comparative analyses of restriction enzymes from the
four currently known superfamilies of nucleases with distinct folds, using
crystallographic and bioinformatic methods, with the emphasis on
theoretical predictions and their experimental validation by site-directed
mutagenesis and biochemical analyses of the mutants.

<>

<1>Bujnicki, J.M.
<2>Molecular phylogenetics of restriction endonucleaseses.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>63-93
<6>2004
<7>1. Discovery and classification of restriction enzymes.  The phenomenon of restriction and
modification was first discovered in the early 1950s.  It was observed that certain strains of
bacteria inhibited (restricted) the growth of bacteriophages previously propagated on a
different strain.  In the early 1960s, it was found that the restriction is due to the
enzymatic cleavage of the phage DNA by sequence-specific endonucleases (REases), which are
sensitive to covalent modification of bases in the target sequence.  Some of the REases
produced discrete DNA fragments upon cleavage.  This property proved very useful for analyzing
and rearranging DNA, which soon prompted the rapid development of genetic engineering
techniques as well as the search for more REases with novel recognition sequences.  It was in
the mid-1970s when cloning of R-M enzymes themselves began.  It was found that most of
restriction enzymes are genetically linked with modification enzymes of cognate specificity,
forming R-M systems, but a few solitary enzymes were also characterized.

<>

<1>Bujnicki, J.M., Feder, M., Ayres, C.L., Redman, K.L.
<2>Sequence-structure-function studies of tRNA:m5C methyltransferase Trm4p and its relationship to DNA:m5C and RNA:m5U methyltransferases.
<3>Nucleic Acids Res.
<4>32
<5>2453-2463
<6>2004
<7>Three types of methyltransferases (MTases) generate 5-methylpyrimidine in nucleic acids,
forming m5U in RNA, m5C in RNA and m5C in DNA. The DNA:m5C
MTases have been extensively studied by crystallographic, biophysical,
biochemical and computational methods. On the other hand, the
sequence-structure-function relationships of RNA:m5C MTases remain
obscure, as do the potential evolutionary relationships between the three
types of 5-methylpyrimidine-generating enzymes. Sequence analyses and
homology modeling of the yeast tRNA:m5C MTase Trm4p (also called Ncl1p)
provided a structural and evolutionary platform for identification of
catalytic residues and modeling of the architecture of the RNA:m5C MTase
active site. The analysis led to the identification of two invariant
residues that are important for Trm4p activity in addition to the
conserved Cys residues in motif IV and motif VI that were previously found
to be critical. The newly identified residues include a Lys residue in
motif I and an Asp in motif IV. A conserved Gln found in motif X was found
to be dispensable for MTase activity. Locations of essential residues in
the model of Trm4p are in very good agreement with the X-ray structure of
an RNA:m5C MTase homolog PH1374. Theoretical and experimental analyses
revealed that RNA:m5C MTases share a number of features with either
RNA:m5U MTases or DNA:m5C MTases, which suggested a tentative phylogenetic
model of relationships between these three classes of 5-methylpyrimidine
MTases. We infer that RNA:m5C MTases evolved from RNA:m5U MTases by
acquiring an additional Cys residue in motif IV, which was adapted to
function as the nucleophilic catalyst only later in DNA:m5C MTases,
accompanied by loss of the original Cys from motif VI, transfer of a
conserved carboxylate from motif IV to motif VI and sequence permutation.

<>

<1>Bujnicki, J.M., Feder, M., Radlinska, M., Blumenthal, R.M.
<2>Structure prediction and phylogenetic analysis of a functionally diverse family of proteins homologous to the MT-A70 subunit of the  human mRNA:m6A methyltransferase.
<3>J. Mol. Evol.
<4>55
<5>431-444
<6>2002
<7>MT-A70 is the S-adenosylmethionine-binding subunit of human mRNA:m6A methyltransferase
(MTase), an enzyme that sequence-specifically
methylates adenines in pre-mRNAs. The physiological importance yet
limited understanding of MT-A70 and its apparent lack of similarity to
other known RNA MTases combined to make this protein an attractive
target for bioinformatic analysis. The sequence of MT-A70 was subjected
to extensive in silico analysis to identify orthologous and paralogous
polypeptides. This analysis revealed that the MT-A70 family comprises
four subfamilies with varying degrees of interrelatedness. One
subfamily is a small group of bacterial DNA:m6A MTases. The other three
subfamilies are paralogous eukaryotic lineages, two of which have not
been associated with MTase activity but include proteins having
substantial regulatory effects. Multiple sequence alignments and
structure prediction for members of all four subfamilies indicated a
high probability that a consensus MTase fold domain is present.
Significantly, this consensus fold shows the permuted topology
characteristic of the beta class of MTases, which to date has only been
known to include DNA MTases.

<>

<1>Bujnicki, J.M., Radlinska, M.
<2>Molecular phylogenetics of DNA 5mC-methyltransferases.
<3>Acta Microbiol. Pol.
<4>48
<5>19-30
<6>1999
<7>We have identified a total of 88 members of the DNA-(cytosine-5) methyltransferase (5mC MTase)
family whose sequences have been deposited in the databases. The results of a comparison of
these sequences is presented in the form of an alignment-based phylogenetic tree and sequence
logos for 10 conserved motifs. Phylogenetic analysis showed that members of the family
aggregate into subfamilies which are usually consistent with their target specificity.
However, it was also shown that similar target specificity does not necessarily imply close
homology of the catalytic domain of MTases, which strongly supports the hypothesis that target
recognition evolved independently of catalytic properties. This analysis also indicate that
the 5mC MTase was present in the cenancestor (last common ancestor) of eubacteria,
archaebacteria, and eukaryotes. The phylogeny of the 5mC MTases catalytic domain provides the
basis for establishing the patterns of evolutionary change that characterize this family of
proteins with conserved structural core and variable and mobile modules not directly involved
in formation of the active site.

<>

<1>Bujnicki, J.M., Radlinska, M.
<2>Molecular evolution of DNA-(cytosine-N4) methyltransferases: evidence for their polyphyletic origin.
<3>Nucleic Acids Res.
<4>27
<5>4501-4509
<6>1999
<7>DNA N4-cytosine methyltransferases (N4mC MTases) are a family of S -adenosyl-L-methionine
(AdoMet)-dependent MTases. Members of this family were previously found to share nine
conserved sequence motifs, but the evolutionary basis of these similarities has never been
studied in detail. We performed phylogenetic analysis of 37 known and potential new family
members from the multiple sequence alignment using distance matrix, parsimony and maximum
likelihood approaches to infer the evolutionary relationship among the N4mC MTases and
classify them into groups of orthologs. All the treeing algorithms employed as well as results
of exhaustive sequence database searching support a scenario, in which the majority of N4mC
MTases, except for M.BalI and M.BamHI, arose by divergence from a common ancestor.
Interestingly, MTases M.BalI and M.BamHI apparently originated from N6-adenine MTases and
represent the most recent addendum to the N4mC MTase family. In addition to the previously
reported nine sequence motifs, two more conserved sequence patches were detected. Phylogenetic
analysis also provided the evidence for massive horizontal transfer of MTase genes, presumably
with the whole restriction-modification systems, between Bacteria and Archaea.

<>

<1>Bujnicki, J.M., Radlinska, M.
<2>Is the HemK family of putative S-adenosylmethionine-dependent methyltransferases a "missing" zeta subfamily of adenine methyltransferases?  A hypothesis.
<3>Life
<4>48
<5>247-249
<6>1999
<7>Previous comparative studies revealed close similarity among various groups of
S-adenosyl-L-methionine-dependent methyltransferases, indicating their common evolutionary
origin.  We present evidence for a remarkable similarity between the sequence and predicted
structure of HemK (a widespread family of putative proteins encoded in genomes from bacteria
to humans) and the catalytic domain of the gamma-subfamily of adenine-specific DNA MTases
(N6mA MTases).  We predict the structure and function of the putative catalytic domain of HemK
proteins and speculate that the target-recognizing function may be conferred by the N-terminal
variable region.

<>

<1>Bujnicki, J.M., Radlinska, M.
<2>Cloning and characterization of M.LmoA118I, a novel DNA:m4C methyltransferase from the Listeria  monocytogenes Phage A118, a close homolog of M.NgoMXV.
<3>Acta Microbiol. Pol.
<4>50
<5>155-160
<6>2001
<7>A homolog of M.NgoMXV DNA:M4C methyltransferase has been identified among the open reading
frames deduced from the genomic sequence of Listeria monocytogenes phage A118.  The gene
coding for this putative protein has been cloned in Escherichia coli and its enzymatic
activity in vivo in this host have been analyzed.  Remarkably, although M.NgoMXV and
M.LmoA118I exhibit high sequence similarity (58% identical and 19% conservatively substituted
residues), their target preferences differ: both proteins exhibit "relaxed" sequence
specificity, but while M.LmoA118I more efficiently methylates GGCC sites, it seems to target
only a subset of CCWGG sites methylated by M.NgoMXV.

<>

<1>Bujnicki, J.M., Radlinska, M., Rychlewski, L.
<2>Atomic model of the 5-methylcytosine-specific restriction enzyme McrA reveals an atypical zinc finger and structural similarity to beta beta alpha-Me endonucleases.
<3>Mol. Microbiol.
<4>37
<5>1280-1281
<6>2000
<7>Using a novel FFAS algorithm for supersensitive sequence comparison and fold recognition, we
have identified a domain in the 5-methylcytosine-specific restriction enzyme (ENase) McrA from
Escherichia coli that is common to the beta beta alpha-Me superfamily of nucleases.  We
carried out extensive threading of the McrA sequence onto the known protein structures and
detected a high compatibility with conformation assumed by the DNase domain of colicins E7 and
E9, although it displays no significant similarity to the sequences of these proteins.  The
model presented reveals that McrA shares a putative zinc finger with homologous group II
intron maturases and the more remotely related coliphage T4 endonuclease VII, despite the fact
that this structure is absent from colicins.  This is the first robust structure prediction of
an ENase ever performed.  It strongly suggests that ENases do not all share the same
three-dimensional fold and, as a functional class of proteins, they should be regarded as
products of convergent evolution.

<>

<1>Bujnicki, J.M., Radlinska, M., Zaleski, P., Piekarowicz, A.
<2>Cloning of the Haemophilus influenzae Dam methyltransferase and analysis of its relationship to the Dam methyltransferase encoded by the HP1 phage.
<3>Acta Biochim. Pol.
<4>48
<5>969-983
<6>2001
<7>In this paper we report cloning and experimental characterization of the DNA adenine
methyltransferase (dam) gene from Haemophilus
influenzae and comparison of its product with the Dam protein from the
lysogenic phage of H. influenzae, HP1. Molecular modeling of M.HinDam
and M.HP1Dam was carried out, providing a framework for a comparative
analysis of these enzymes and their close homologs in the structural
context. Both proteins share the common fold and essential
cofactor-binding and catalytic residues despite overall divergence.
However, subtle but significant differences in the cofactor-binding
pocket have been identified. Moreover, while M.HinDam seems to contact
its target DNA sequence using a number of loops, most of them are
missing from M.HP1Dam. Analysis of both MTases suggests that their
catalytic activity was derived from a common ancestor, but similar
sequence specificities arose by convergence.

<>

<1>Bujnicki, J.M., Rotkiewicz, P., Kolinski, A., Rychlewski, L.
<2>Three-dimensional modeling of the I-TevI homing endonuclease catalytic domain, a GIY-YIG superfamily member, using NMR restraints and Monte Carlo dynamics.
<3>Protein Eng.
<4>14
<5>717-721
<6>2001
<7>Using a recent version of the SICHO algorithm for in silico protein folding, we made a blind
prediction of the tertiary structure of the N-terminal, independently folded, catalytic domain
(CD) of the I-TevI homing endonuclease, a representative of the GIY-YIG superfamily of homing
endonucleases. The secondary structure of the I-TevI CD has been determined using NMR
spectroscopy, but computational sequence analysis failed to detect any protein of known
tertiary structure related to the GIY-YIG nucleases (Kowalski et al., Nucleic Acids Res.,
1999, 27, 2115-2125). To provide further insight into the structure-function relationships of
all GIY-YIG superfamily members, including the recently described subfamily of type II
restriction enzymes (Bujnicki et al., Trends Biochem. Sci., 2000, 26, 9-11), we incorporated
the experimentally determined and predicted secondary and tertiary restraints in a reduced
(side chain only) protein model, which was minimized by Monte Carlo dynamics and simulated
annealing. The subsequently elaborated full atomic model of the I-TevI CD allows the available
experimental data to be put into a structural context and suggests that the GIY-YIG domain may
dimerize in order to bring together the conserved residues of the active site.

<>

<1>Bujnicki, J.M., Rychlewski, L.
<2>Identification of a PD-(D/E)XK-like domain with a novel configuration of the endonuclease active site in the methyl-directed restriction enzyme Mrr and its homologs.
<3>Gene
<4>267
<5>183-191
<6>2001
<7>The Escherichia coli K-12 restriction enzyme Mrr recognizes and cleaves N6-methyladenine- and
5-methylcytosine-containing DNA. Its amino acid sequence has been subjected to structure
prediction and comparison with other sequences from publicly available sources. The results
obtained suggest that Mrr and related putative endonucleases possess a cleavage domain typical
for all the so far structurally characterized type II restriction enzymes, however with an
unusual glutamine residue at the central position of the (D/E)-(D/E)XK hallmark of the active
site. The 'missing' acidic side chain was instead found anchored in a different, unusual
position, suggesting that Mrr represents a third topological variant of the endonuclease
active site in addition to the two alternatives determined previously (Skirgaila et al., 1998.
J. Mol. Biol. 279, 473-481). One of the newly identified putative endonucleases from the Mrr
family is composed of two diverged cleavage domains, which possess both the 'typical' D-EXK
and the 'Mrr-like' D-QXK variants of the active site. Among the Mrr homologs there are also
proteins from yeast, in which restriction phenotype has not been observed, suggesting that the
free-standing Eukaryotic PD-(D/E)XK superfamily members might be implicated in other cellular
processes involving enzymatic DNA cleavage.

<>

<1>Bujnicki, J.M., Rychlewski, L.
<2>Unusual evolutionary history of the tRNA splicing endonuclease EndA: Relationship to the LAGLIDADG and PD-(D/E)XK deoxyribonucleases.
<3>Protein Sci.
<4>10
<5>656-660
<6>2001
<7>The tRNA splicing endoribonuclease EndA from Methanococcus jannaschii is a homotetramer formed
via heterologous interaction between the two pairs of homodimers. Each monomer consists of two
alpha/beta domains, the N-terminal domain (NTD) and the C-terminal domain (CTD) containing the
RNase A-like active site. Comparison of the EndA coordinates with the publicly available
protein structure database revealed the similarity of both domains to site-specific
deoxyribonucleases: the NTD to the LAGLIDADG family and the CTD to the PD-(D/E)XK family.
Superposition of the NTD on the catalytic domain of LAGLIDADG homing endonucleases allowed a
suggestion to be made about which amino acid residues of the tRNA splicing nuclease might
participate in formation of a presumptive cryptic deoxyribonuclease active site. On the other
hand, the CTD and PD-(D/E)XK endonucleases, represented by restriction enzymes and a phage
lambda exonuclease, were shown to share extensive similarities of the structural framework, to
which entirely different active sites might be attached in two alternative locations. These
findings suggest that EndA evolved from a fusion protein with at least two distinct
endonuclease activities: the ribonuclease, which made it an essential "antitoxin" for the
cells whose RNA genes were interrupted by introns, and the deoxyribonuclease, which provided
the means for homing-like mobility. The residues of the noncatalytic CTDs from the positions
corresponding to the catalytic side chains in PD-(D/E)XK deoxyribonucleases map to the surface
at the opposite side to the tRNA binding site, for which no function has been implicated. Many
restriction enzymes from the PD-(D/E)XK superfamily might have the potential to maintain an
additional active or binding site at the face opposite the deoxyribonuclease active site, a
property that can be utilized in protein engineering.

<>

<1>Bujnicki, J.M., Rychlewski, L.
<2>The herpesvirus alkaline exonuclease belongs to the restriction endonuclease PD-(D/E)XK superfamily: Insight from molecular modeling and phylogenetic analysis.
<3>Virus Genes
<4>22
<5>219-230
<6>2001
<7>The PD-(D/E)XK superfamily of deoxyribonucleases (Enases) comprises restriction endonucleases,
exonucleases and nicking enzymes, which share a common fold and the architecture of the active
site.  Their extreme divergence generally hampers identification of novel members based solely
on sequence comparisons.  Here we report a remote similarity between the phage lambda
exonuclease (lambda-exo), branching out early in the evolutionary history of Enases, with the
family of alkaline exonucleases encoded by various viruses infecting higher Eukaryota.  The
predicted structural compatibility and the conservation of the functionally important residues
between AE and Enases strongly suggest a distant evolutionary relationship between these
proteins.  According to the results of extensive sequence database mining, sequence/structure
threading and molecular modeling it is plausible that the AE proteins with lambda-exo and some
other putative phage-encoded exonucleases form a distinct subfamily of PD-(D/E)XK Enases.  The
phylogenetic history of this subfamily is inferred using sequence alignment and distance
matrix methods.

<>

<1>Bujnicki, J.M., Rychlewski, L., Radlinska, M.
<2>Polyphyletic evolution of type II restriction enzymes revisited: two independent sources of second-hand folds revealed.
<3>Trends Biochem. Sci.
<4>26
<5>9-11
<6>2001
<7>Using algorithms for protein sequence analysis we predict that some of the canonical type II
and type IIS restriction enzymes have an active site with a substantially different
architecture and fold from the "typical" PD-(D/E)xK superfamily.  These results suggest that
they are related to nucleases from the HNH and GIY-YIG superfamilies.

<>

<1>Bulach, D.M., Zuerner, R.L., Wilson, P., Seemann, T., McGrath, A., Cullen, P.A., Davis, J., Johnson, M., Kuczek, E., Alt, D.P., Peterson-Burch, B., Coppel, R.L., Rood, J.I., Davies, J.K., Adler, B.
<2>Genome reduction in Leptospira borgpetersenii reflects limited transmission potential.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>14560-14565
<6>2006
<7>Leptospirosis is one of the most common zoonotic diseases in the world, resulting in high
morbidity and mortality in humans and affecting global
livestock production. Most infections are caused by either Leptospira
borgpetersenii or Leptospira interrogans, bacteria that vary in their
distribution in nature and rely on different modes of transmission. We
report the complete genomic sequences of two strains of L. borgpetersenii
serovar Hardjo that have distinct phenotypes and virulence. These two
strains have nearly identical genetic content, with subtle frameshift and
point mutations being a common form of genetic variation. Starkly limited
regions of synteny are shared between the large chromosomes of L.
borgpetersenii and L. interrogans, probably the result of frequent
recombination events between insertion sequences. The L. borgpetersenii
genome is approximately 700 kb smaller and has a lower coding density than
L. interrogans, indicating it is decaying through a process of insertion
sequence-mediated genome reduction. Loss of gene function is not random
but is centered on impairment of environmental sensing and metabolite
transport and utilization. These features distinguish L. borgpetersenii
from L. interrogans, a species with minimal genetic decay and that
survives extended passage in aquatic environments encountering a mammalian
host. We conclude that L. borgpetersenii is evolving toward dependence on
a strict host-to-host transmission cycle.

<>

<1>Bulanenkova, S., Kotova, E., Snezhkov, E., Akopov, S., Nikolaev, L., Sverdlov, E.
<2>Dam methylase accessibility as an instrument for analysis of mammalian chromatin structure.
<3>FEBS J.
<4>279
<5>493
<6>2012
<7>The study of regulatory mechanisms operating at the level of the chromatin fiber is one of the
rapidly developing fields of modern molecular biology.  Changes in chromatin condensation
state is one of the main regulatory mechanisms of cell functions, such as transcription,
replication, DNA repair, and other processes. The lack of information on dynamic behavior of
extended chromatin regions leads to a gap in our understanding of mechanisms of coordinated
expression regulation of gene ensembles.  Chromatin structure analysis is in many cases based
on the accessibility of chromatin DNA to modifying agents.  The main benefits are granted by
in vivo detecting agents such as Dam methylase, for example.  For a 140 kb human genome locus,
an analysis of the distribution of Dam methylase accessible sites, DNase I sensitive and
resistant regions, and acetylated histone H3 molecules was performed and compared with
transcriptional activity of the genes within the locus.  It was demonstrated that promoter
regions of all highly and moderately transcribed genes are highly accessible to methylation by
Dam methylase.  In contrast, promoters of non-transcribed genes showed a very low extent of
Dam methylation.  Some highly Dam methylase accessible regions are present in the intergenic
regions of the locus suggesting that the latter contain either unidentified non-coding
transcripts or extended regulatory elements like locus control regions.

<>

<1>Bulgari, D., Minio, A., Casati, P., Quaglino, F., Delledonne, M., Bianco, P.A.
<2>Curtobacterium sp. Genome Sequencing Underlines Plant Growth Promotion-Related Traits.
<3>Genome Announcements
<4>2
<5>e00592-14
<6>2014
<7>Endophytic bacteria are microorganisms residing in plant tissues without causing  disease
symptoms. Here, we provide the high-quality genome sequence of
Curtobacterium sp. strain S6, isolated from grapevine plant. The genome assembly
contains 2,759,404 bp in 13 contigs and 2,456 predicted genes.

<>

<1>Bulkacz, J., Roulland-Dussoix, D., Boyer, H.W.
<2>Deoxyribonucleic acid modification by intermediate-type modification mutants of Escherichia coli K-12 and B.
<3>J. Bacteriol.
<4>124
<5>1395-1402
<6>1975
<7>The modification of bacteriophages grown on r-m+- restriction and modification
mutants of Escherichia coli K-12 or B appears to be related to the number of
restriction-specific sites in the viral genome.  Bacteriophage fd and its
mutant U1 fd, which carry two and one B-specific sites, respectively, are not
modified in vivo by rB-mB+- mutant strains.  In vitro treatment of fd RF.B+-
deoxyribonucleic acid (DNA) or U1 fd RF.B+- DNA by endo R.EcoB results in
cleavage of the substrate DNA.  Lambda bacteriophge, after growth in r-m+-
mutant host strains (k.K+- or k.B+-), is partially protected from in vivo
degradation by wild-type homospecific strains.  Its efficiency of plating on
these strains is approximately 10-2.  However, a hybrid Phi80-lambda phage
which carries only one K-specific site (skk-1) is not modified by rK-mK+-
strains.  Labeled DNAs from k.B+- and k.K+- phages were used as substrates for
endo R.Eco B and endo R.Eco K nucleases.  Zonal centrifugation analysis of the
products of the reactions indicate that rK-mK+- mutants do not protect lambda
DNA from in vitro degradation by endo R.EcoK.  In contrast, rB-mB+- mutants
appear to partially protect lambda DNA from attack by endo R.Eco B.

<>

<1>Bulkowska, U., Ishikawa, T., Kurlandzka, A., Trzcinska-Danielewicz, J., Derlacz, R., Fronk, J.
<2>Expression and of murine DNA methyltransferases Dnmt1 and Dnmt3a in the yeast Saccharomyces cerevisiae.
<3>Yeast
<4>24
<5>871-882
<6>2007
<7>Murine DNA methyltransferases Dnmt1 and Dnmt3a were expressed in the yeast Saccharomyces
cerevisiae. Adjustment to yeast preferences of the
nucleotide sequences upstream and downstream of the translation
initiation sites of both cDNAs was needed to obtain significant levels
of the methyltransferases. Both proteins were correctly localized to
the nucleus and their presence had no measurable influence on the
functioning of yeast cells. Both Dnmt1 and Dnmt3a expressed in yeast
cells were enzymatically active in vitro, and in vivo in the genomic
DNA of the transgenic S. cerevisiae ca. 0.06% and 0.4%, respectively,
of cytosines became methylated. This level of DNA methylation is about
100- to 10-fold less than that observed in mammalian cells. The
constructed system may be used to investigate the in vivo specificity
of individual mammalian DNA methyltransferases and to search for
additional factors needed to allow more efficient in vivo methylation
of chromatincontained DNA and to study their mechanism of action.

<>

<1>Bull, J.J., Vimr, E.R., Molineux, I.J.
<2>A tale of tails: Sialidase is key to success in a model of phage therapy against K1-capsulated Escherichia coli.
<3>Virology
<4>398
<5>79-86
<6>2010
<7>Prior studies treating mice infected with Escherichia coli O18:K1:H7 observed that phages
requiring the K1 capsule for infection (K1-dep) were superior to capsule-independent (K1-ind)
phages. We show that three K1-ind phages all have low fitness when grown on cells in serum
whereas fitnesses of four K1-dep phages were high. The difference is serum-specific, as
fitnesses in broth overlapped. Sialidase activity was associated with all K1-dep virions
tested but no K1-ind virions, a phenotype supported by sequence analyses. Adding endosialidase
to cells infected with K1-ind phage increased fitness in serum by enhancing productive
infection after adsorption. We propose that virion sialidase activity is the primary
determinant of high fitness on cells grown in serum, and thus in a mammalian host. Although
the benefit of sialidase is specific to K1-capsulated bacteria, this study may provide a
scientific rationale for selecting phages for therapeutic use in many systemic infections.

<>

<1>Bull, L.N., Hewitt, J.E., Cox, D.R., Myers, R.M.
<2>Sensitivity of HincII to CpG methylation.
<3>Nucleic Acids Res.
<4>21
<5>2021
<6>1993
<7>Restriction fragment length polymorphism (RFLP) analysis in human genetics can be confounded
by the sensitivity of restriction enzymes to cytosine methylation that occurs at many
cytosine-guanine (CpG) dinucleotides. Previous reports provided conflicting information on the
sensitivity of HincII to C methylation (1-3). We identified alleles at a chromosome 4p16 locus
that were inherited in a non-Mendelian manner due to inhibition of HincII cleavage by CpG
methylation. We confirmed that cleavage by HincII, whose recognition sequence is
5'GTPyPuAC3', is inhibited by the presence of 5-methyl-cytosine at the terminal base of its
recognition site.

<>

<1>Bullas, L.R.
<2>Cloning of the SA system for the restriction and modification of DNA from Salmonella typhimurium into phage lambda.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>224
<6>1994
<7>The Salmonella typhimurium SA system of genes for the restriction and modification of DNA (R-M
system), the first such system recognized in this organism, has been cloned. Selection for the
SA genes has been difficult since all SA E. coli/Salmonella hybrids derived years ago failed
to express either SA restriction or modification when tested at this time. However, such a
hybrid between E. coli and Salmonella typhi, designated LB4019, was found still to express SA
restriction and modification. Several libraries consisting of partial SauIIIA digests of
genomic S. typhimurium DNA ligated to phageEMBL4 and amplified on the non-restriction,
non-modifying strain of E. coli NM477, were plated out on a lawn of LB4019. All plaques which
were produced on LB4019 were picked, plated out on NM477 and the efficiencies of plating (eop)
on LB4019 determined. From one library, a number of plaques plated out at eop of 1. In
addition, plaques that gave significantly higher eop than unmodified phageEMBL on LB4019 (1 x
10-3) were isolated. The phages fell into four groups relative to their eop on LB4019. Group 1
clones consistently plated with an eop of 1.0, group 2 an eop of about 0.5, group 3 an eop of
about 0.2 and group 4 an eop of about 0.02. The cloning of these apparently partially
SA-modified clones in addition to the completely SA-modified clones was unexpected and is
unique to this system. Restriction analysis indicates that the sizes of the fragments cloned
in the four groups are similar. A molecular and genetic analysis of representative clones is
in progress.

<>

<1>Bullas, L.R., Colson, C.
<2>DNA restriction and modification systems in Salmonella.  III.  SP, a Salmonella potsdam system allelic to the SB system in Salmonella typhimurium.
<3>Mol. Gen. Genet.
<4>139
<5>177-188
<6>1975
<7>By screening 42 Salmonella strains with P3, a temperate bacteriophage with an
unusally wide host range, five new DNA restriction and modification systems
(R-M systems) were identified in five different serotypes in Kauffmann-White
group C.  One of these systems, SP, in a Pl-sensitive strain of S. potsdam, was
analyzed genetically by Pl transduction methods in which SP was transferred
into S. typhimurium and E. coli/S. typhimurium hybrids.  It was found that the
genes of the SP system were allelic and functionally homologous to the genes of
the SB system of S. typhimurium.

<>

<1>Bullas, L.R., Colson, C., Neufeld, B.
<2>Deoxyribonucleic acid restriction and modification systems in Salmonella:  chromosomally located systems of different serotypes.
<3>J. Bacteriol.
<4>141
<5>275-292
<6>1980
<7>With the use of four different phages, Salmonella strains representing 85 different serotypes
were examined to determine their restriction-modification phenotype.  They fell into one of
three groups on this basis:  group 1, those which lacked the common LT system; group 2, those
in which only the LT system could be recognized; and group 3, those which possessed the LT
system and at least one other system shown with some serotypes to be closely linked to serB.
The specificity of the serB-linked restricted-modification system was unique for each
serotype, but different strains of the same serotype expressed the same specificity.  Two of
the systems were shown to behave in genetic crosses as functional alleles of the S.
typhimurium SB system.  It is possible that these serB-linked restriction-modification systems
constitute a large multiallelic series of genes extending throughout the Salmonella genus and
Escherichia coli.  We suggest that the division of the Salmonella into the three
restriction-modification groups may be significant in defining a biological grouping of the
different serotypes within the genus which may ultimately be useful in describing the
Salmonella species.  From the genetic relatedness between the genes of some of the Salmonella
restriction-modification systems with those of the E. coli systems, we deduce that the
restriction endonucleases produced by the Salmonella serB-linked systems are of type I.
Determination of the nucleotide sequences of the recognition sites of the restriction
endonucleases of selected Salmonella systems should further our understanding of specificity
with these enzymes.

<>

<1>Bullas, L.R., Colson, C., van Pel, A.
<2>DNA restriction and modification systems in Salmonella. SQ, a new system derived by recombination between the SB system of Salmonella typhimurium and the SP system of Salmonella potsdam.
<3>J. Gen. Microbiol.
<4>95
<5>166-172
<6>1976
<7>As a result of PI-mediated cotransduction with serB from Salmonella potsdam to
the Escherichia coli/Salmonella typhimurium hybrid 4617, one recombinant,
L4004, was isolated which had a restriction-modification (R-M) system different
from the SB and SP systems of its parents, and was designated SQ.  The genes of
complementation experiments to be functionally related to those of the K system
of E. coli.  Evidence that the SQ system in L4004 arose as the result of a
recombination event within the hsdS genes of SB and SP is discussed.

<>

<1>Bullas, L.R., Harding, G.
<2>Cryptic genes for the restriction and modification of DNA in Salmonella typhi and Salmonella gallinarum.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>96
<5>258
<6>1996
<7>Salmonella typhi and Salmonella gallinarum fail to express the StyLTI and StyII
genes for the restriction and modification of DNA which are expressed by other Salmonella
serotypes.  S. typhi transformed by either pRUCL531 with the S. typhimurium StyLTI restriction
and modification genes or pXC1 with the StyLTIII modification gene, has a reduced ability to
propagate within human Mac6 macrophage-like cells.  In P1 transductions from S. typhi to an E.
coli/Salmonella StyLTIII+ recipient, the "lack of expression" of StyLTIII is co-transduced
with
serB to 17% of recombinants.  These observations prompted us to investigate whether S. typhi
and
S. gallinarum might possess cryptic StyLTI or StyLTIII systems of genes.  We isolated a 1.8kb
DNA sequence which included the end of the modification gene and the beginning of the
restriction
gene of StyLTI, and a 2.6 kb sequence which included parts of the StyLTIII modification and
specificity genes, and used them as probes in Southern hybridizations to genomic DNA of S.
typhi
and S. gallinarum.  The DNA of both serotypes hybridized strongly to both probes.  Thus, S.
typhi and S. gallinarum both possess cryptic StyLTI and StyLTIII R-M genes.  Since pRUCL531-
and pXC1-transformed S. typhi have reduced growth parameters in Mac6, it is likely that the S.
typhi cryptic R-M genes are concerned with the ability of this serotype to propagate within
human
macrophages.

<>

<1>Bullas, L.R., Nutter, R.L.
<2>Host-induced modification of Salmonella potsdam bacteriophage P3 by growth in E. coli.
<3>Bacteriol. Proc.
<4>69
<5>157
<6>1969
<7>The temperate Salmonella potsdam bacteriophage P3 lyses E. coli K and E. coli C
with high efficiency.  Growth of P3 in either E. coli K (producing phage P3.K)
or E. coli C (phage P3-C) modified the phage so that its original Salmonella
host is now insensitive, but the phage retains the ability to grow on either
strain of E. coli.  P3, therefore, propagated on S. potsdam (phage P3.S) has an
unrestricted host range, while P3.K and P3.C are restricted.  Rare P3.K and
P3.C phages (10-6) will grow on the original Salmonella host producing
unrestricted phages identical to P3.S.  E. coli is readily lysogenized by P3.S,
in which it is also inducible by UV light.  Phages grown either on Salmonella
or E. coli appear to be serologically identical, although comparison of the
rates of neutralization suggest that slight structural changes of the phage
particle may have occurred in E. coli, rendering P3.K and P3.C more susceptible
than P3.S to neutralization by antiserum produced against P3.S.  These and
other results indicate that a host-induced modification of the Salmonella phage
P3.S occurs by growth in E. coli K or E. coli C.  Similar results have been
obtained for the related Salmonella phage P9a.

<>

<1>Bullas, L.R., Rajadas, P.T.
<2>Evidence of additional families of TypeI hsd genes concerned with the restriction and modification of DNA in Salmonella.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>87
<5>154
<6>1987
<7>Hybridization and complementation experiments have shown that serB-linked genes
for the restriction and modification of DNA in E. coli and Salmonella comprise
at least two "families".  A third "family" consists of genes on a plasmid.  We
have prepared DNA probes from the cloned genes of the two chromosomal
families--hsdSB and hsdSP from the first family and hsdA from the second
family.  In addition we prepared a probe from the hsdSEN genes (from S.
enteritidis) which we had earlier cloned in phage lambda and subcloned into
pUC18 and whose family relationship was unknown.  With each of these probes
radioactively labelled with 32P and Southern gel transfer, we have performed a
series of hybridization experiments with the DNA from six E. coli/Salmonella
hybrids possessing the serB-linked hsd genes from a Salmonella serotype.
Neither of the two probes from the first family nor the A probe of the second
family hybridized with the DNA from any of the hybrids.  The SEN probe
hybridized with DNA from only two of the six hybrids.  Thus additional families
of the Type1 hsd genes must exist in Salmonella.

<>

<1>Bullas, L.R., Ryu, J.-I.
<2>Salmonella typhimurium LT2 strains which are r- m+ for all three chromosomally located systems of DNA restriction and modification.
<3>J. Bacteriol.
<4>156
<5>471-474
<6>1983
<7>We describe the derivation of two strains of Salmonella typhimurium LT2 which are r- m+ for
all three of the known chromosomal genes for the restriction and modification of DNA, hsdLT,
hsdSA, and hsdSB; the strains were designated LB5000 and LB5010.  LB5000 is a smooth
derivative sensitive to phage P22; LB5010 is a galE strain sensitive to phage P1.

<>

<1>Bult, C.J. et al.
<2>Complete genome sequence of the Methanogenic archaeon, Methanococcus jannaschii.
<3>Science
<4>273
<5>1058-1073
<6>1996
<7>The complete 1.66-megabase pair genome sequence of an autotrophic archaeon, Methanococcus
jannaschii, and its 58- and 16-kilobase pair extrachromosomal elements have been determined by
whole-genome random sequencing.  A total of 1738 predicted protein coding genes were
identified; however, only a minority of those (38 percent) could be assigned a putative
cellular role with high confidence.  Although the majority of genes related to energy
production, cell division, and metabolism in M. jannaschii are most similar to those found in
Bacteria, most of the genes involved in transcription, translation, and replication in M.
jannaschii are more similar to those found in Eukaryotes.

<>

<1>Bult, C.J., White, O.R., Smith, H.O., Woese, C.R., Venter, J.C.
<2>Selected polynucleotide and polypeptide sequences of the methanogenic archaeon, methanococcus jannashii.
<3>US Patent Office
<4>US 6503729 A
<5>
<6>2003
<7>The present application describes selected polynucleotide sequences from the 1.66-megabase
pair
genome sequence of an autotrophic archaeon, Methanococcus jannaschii, and its 58- and
16-kilobase pair extrachromosomal elements.

<>

<1>Buluwela, L., Malcolm, A.D.B., Coleman, D.V., Gardner, S.D.
<2>The use of the restriction endonuclease DdeI for mapping related DNAs.
<3>Biosci. Rep.
<4>1
<5>223-228
<6>1981
<7>Restriction endonuclease mapping of BK virus strain GS using DdeI has allowed
us to correct and extend the previously published map.

<>

<1>Bulyk, M.L., Gentalen, E., Lockhart, D.J., Church, G.M.
<2>Quantifying DNA-protein interactions by double-stranded DNA arrays.
<3>Nat. Biotechnol.
<4>17
<5>573-577
<6>1999
<7>We have created double-stranded oligonucleotide arrays to perform highly parallel
investigations of DNA-protein interactions.  Arrays of single-stranded DNA oligonucleotides,
synthesized by a combination of photolithography and solid-state chemistry, have been used for
a variety of applications, including large-scale mRNA expression monitoring, genotyping, and
sequence-variation analysis. We converted a single-stranded to a double-stranded array by
synthesizing a constant sequence at every position on an array and then annealing and
enzymatically extending a complementary primer. The efficiency of second-strand synthesis was
demonstrated by incorporation of fluorescently labeled dNTPs (2'-deoxyribonucleoside
5'-triphosphates) and by terminal transferase addition of a fluorescently labeled ddNTP. The
accuracy of second-strand synthesis was demonstrated by digestion of the arrayed
double-stranded DNA (dsDNA) on the array with sequence-specific restriction enzymes. We showed
dam methylation of dsDNA arrays by digestion with DpnI, which cleaves when its recognition
site is methylated. This digestion demonstrated that the dsDNA arrays can be further
biochemically modified and that the DNA is accessible for interaction with DNA-binding
proteins. This dsDNA array approach could be extended to explore the spectrum of
sequence-specific protein binding sites in genomes.

<>

<1>Bumgardner, E., Bey, R.F., Kittichotirat, W., Bumgarner, R.E., Lawrence, P.K.
<2>Genome Sequences of Seven Mycoplasma hyosynoviae Strains Isolated from the Joint  Tissue of Infected Swine (Sus scrofa).
<3>Genome Announcements
<4>2
<5>e00552-14
<6>2014
<7>Mycoplasma hyosynoviae is a Gram-negative bacterium that can cause debilitating arthritis in
swine. Currently, there are no M. hyosynoviae genome sequences in
the GenBank database, which makes it impossible to understand its pathogenesis,
nutrition, or colonization characteristics, or to devise an effective strategy
for its control. Here, we report the genome sequences of seven strains of M.
hyosynoviae. Within each genome, several virulence factors were identified that
may prove important in the pathogenesis of M. hyosynoviae-mediated arthritis and
serve as potential virulence markers that may be critical in vaccine development.

<>

<1>Bunina, Z.F., Kramarov, F.M., Matvienko, N.I.
<2>Bacillus brevis-80 strain used for production of site specific endonuclease employed in preparation of new DNA recombinant molecules.
<3>Soviet Patent Office
<4>SU 1190642 A
<5>
<6>1986
<7>
<>

<1>Bunina, Z.F., Kramarov, V.M., Mazanov, A.L., Pachkunov, D.M., Smolianinov, V.V., Matvienko, N.N.
<2>BbvII:  A new site-specific endonuclease from Bacillus brevis 80.
<3>Bioorg. Khim.
<4>9
<5>1578-1580
<6>1983
<7>BbvII, a new site-specific endonuclease, has been isolated from Bacillus brevis
80 by gel-filtration and chromatography on heparin-Sepharose.  The endonuclease
recognizes a non-symmetrical sequence 5'-GTGTTC-3'/3'-CAGAAG-5' in
double-stranded DNA and cleaves DNA in both strands outside the recognition
sequence.

<>

<1>Bunina, Z.F., Kramarov, V.M., Smolyaninov, V.V., Tolstova, L.A.
<2>XphI - A new sequence-specific endonuclease from Xanthomonas phaseoli.
<3>Bioorg. Khim.
<4>10
<5>1333-1335
<6>1984
<7>A sequence-specific endonuclease XphI was purified by chromatography on
Ultrogel AcA-44 and phosphocellulose.  The final preparation is free of
admixtures of non-specific nucleases.  It is demonstrated that endonuclease
XphI recognized 5'-CTGCAG-3' sequence like endonuclease PstI.  According to gel
filtration data, the molecular mass of endonuclease is 47000+2000.  DNA
methylated by methylases which block its hydrolysis by endonuclease PstI, is
also resistant to the cleavage by endonuclease XphI.

<>

<1>Bunina, Z.F., Smolyaninov, V.V.
<2>Dynamics of site-specific endonuclease accumulation in Caryophanon latum L. cells.
<3>Biotekhnologiya
<4>4
<5>621-623
<6>1988
<7>None

<>

<1>Bunting, K.A., Roe, S.M., Headley, A., Brown, T., Savva, R., Pearl, L.H.
<2>Crystal structure of the Escherichia coli dcm very-short-patch DNA repair endonuclease bound to its reaction product-site in a DNA superhelix.
<3>Nucleic Acids Res.
<4>31
<5>1633-1639
<6>2003
<7>Very-short-patch repair (Vsr) enzymes occur in a variety of bacteria, where they initiate
nucleotide excision repair of G:T mismatches arising
by deamination of 5-methyl-cytosines in specific regulatory sequences. We
have now determined the structure of the archetypal dcm-Vsr endonuclease
from Escherichia coli bound to the cleaved authentic
hemi-deaminated/hemi-methylated dcm sequence 5'-C-OH-3'
5'-p-T-p-A-p-G-p-G-3'/3'-G-p-G-p-T-p(Me5)C-p-C formed by self-assembly of
a 12mer oligonucleotide into a continuous nicked DNA superhelix. The
structure reveals the presence of a Hoogsteen base pair within the
deaminated recognition sequence and the substantial distortions of the DNA
that accompany Vsr binding to product sites.

<>

<1>Burbank, D.E., Shields, S.L., Schuster, A.M., Van Etten, J.L.
<2>5-Azacytidine-resistant mutants of Chlorella Virus IL-3A.
<3>Virology
<4>176
<5>311-315
<6>1990
<7>Many dsDNA-containing viruses which infect the unicellular, eukaryotic
Chlorella-like green alga strain NC64A encode for DNA methyltransferases and
DNA site-specific (restriction) endonucleases.  We have hypothesized that these
endonucleases help degrade host DNA permitting deoxynucleotides to recycle into
virus DNA.  This hypothesis was tested by isolating deletion mutants of
Chlorella virus IL-3A lacking functional genes for the cytosine
methyltransferase M.CviJI and the cognate site-specific endonuclease CviJI.
The growth and burst sizes of the mutants and parent virus were identical.
Also host nuclear and chloroplast DNAs disappeared from infected cells at the
same rates.  Thus M.CviJI and CviJI activities are not required for IL-3A
replication and CviJI activity is not essential for host DNA degradation.

<>

<1>Burckhardt, J., Weisemann, J., Hamilton, D.L., Yuan, R.
<2>Complexes formed between the restriction endonuclease EcoK and heteroduplex DNA.
<3>J. Mol. Biol.
<4>153
<5>425-440
<6>1981
<7>The complexes between the Escherichia coli K restriction endonuclease and
heteroduplex DNA (one strand methylated and one unmethylated) have been
characterized and shown to have different properties from those formed with
unmodified DNA.  The nature of the heteroduplex complex appears to commit the
enzyme to its methylase mode.

<>

<1>Burckhardt, J., Weisemann, J., Yuan, R.
<2>Characterization of the DNA methylase activity of the restriction enzyme from Escherichia coli K.
<3>J. Biol. Chem.
<4>256
<5>4024-4032
<6>1981
<7>The restriction endonuclease from Escherichia coli K is a multifunctional
protein which efficiently methylates heteroduplex DNA (one strand modified and
one strand unmodified) in the presence of S-adenosylmethionine (AdoMet), ATP,
and Mg2+.  The methylase activity is catalytic, and seems to modify different
heteroduplex host specificity sites for E. coli K with equal efficiency.

<>

<1>Burenina, O.Y., Fedotova, E.A., Ryazanova, A.Yu., Protsenko, A.S., Zakharova, M.V., Karyagina, A.S., Solonin, A.S., Oretskaya, T.S., Kubareva, E.A.
<2>Peculiarities of the Regulation of Gene Expression in the Ecl18kI RestrictionModification System.
<3>Acta Naturae
<4>5
<5>70-80
<6>2013
<7>Transcription regulation in bacterial restriction-modification (R-M) systems is an important
process, which provides coordinated expression levels of tandem enzymes, DNA methyltransferase
(MTase) and restriction endonuclease (RE) protecting cells against penetration of alien DNA.
The present study focuses on (cytosine-5)-DNA methyltransferase Ecl18kI (M. Ecl18kI), which is
almost identical to DNA methyltransferase SsoII (M. SsoII) in terms of its structure and
properties. Each of these enzymes inhibits expression of the intrinsic gene and activates
expression of the corresponding RE gene via binding to the regulatory site in the promoter
region of these genes. In the present work, complex formation of M. Ecl18kI and RNA polymerase
from Escherichia.oli with the promoter regions of the MTase and RE genes is studied. The
mechanism of regulation of gene expression in the Ecl18kI R-M system is thoroughly
investigated. M. Ecl18kI and RNA polymerase are shown to compete for binding to the promoter
region. However, no direct contacts between M. Ecl18kI and RNA polymerase are detected. The
properties of M. Ecl18kI and M. SsoII mutants are studied. Amino acid substitutions in the
N-terminal region of M. Ecl18kI, which performs the regulatory function, are shown to
influence not only M. Ecl18kI capability to interact with the regulatory site and to act as a
transcription factor, but also its ability to bind and methylate the substrate DNA. The loss
of methylation activity does not prevent MTase from performing its regulatory function and
even increases its affinity to the regulatory site. However, the presence of the domain
responsible for methylation in the M. Ecl18kI molecule is necessary for M. Ecl18kI to perform
its regulatory function.

<>

<1>Burger, K.J., Schinzel, R.
<2>Restriction endonuclease BglI as a tool for in vitro reconstruction and recombination of plasmid and bacteriophage genomes.
<3>Mol. Gen. Genet.
<4>189
<5>269-274
<6>1983
<7>The restriction endonuclease BglI produces different individual fragment ends from different
cut sites.  This property has allowed us to reconstruct efficiently several commonly used
plasmid and bacteriophage genomes and a number of recombinant plasmids containing up to seven
BglI restriction sites from their constituent BglI fragments.  It is demonstrated that in
vitro reconstitution from BglI fragments can be used to create, in a simple way, recombinant
DNA molecules by recombining in vitro BglI fragments from different mutated or otherwise
related genomes.  Further applications of the method are discussed.

<>

<1>Burgers, W.A., Blanchon, L., Pradhan, S., de Launoit, Y., Kouzarides, T., Fuks, F.
<2>Viral oncoproteins target the DNA methyltransferases.
<3>Oncogene
<4>26
<5>1650-1655
<6>2007
<7>Small DNA tumour viruses have evolved a number of mechanisms to drive nondividing cells into S
phase. Virally encoded oncoproteins such as
adenovirus E1A and human papillomavirus (HPV) E7 can bind an array of
cellular proteins to override proliferation arrest. The DNA
methyltransferase Dnmt1 is the major mammalian enzyme responsible for
maintaining CpG methylation patterns in the cell following replication.
One of the hallmarks of tumour cells is disrupted DNA methylation
patterns, highlighting the importance of the proper regulation of DNA
methyltransferases in normal cell proliferation. Here, we show that
adenovirus 5 E1A and HPV-16 E7 associate in vitro and in vivo with the
DNA methyltransferase Dnmt1. Consistent with this interaction, we find
that E1A and E7 can purify DNA methyltransferase activity from nuclear
extracts. These associations are direct and mediated by the extreme
N-terminus of E1A and the CR3 zinc-finger domain of E7. Furthermore, we
find that a point mutant at leucine 20 of E1A, a residue known to be
critical for its transformation functions, is unable to bind Dnmt1 and
DNA methyltransferase activity. Finally, both E1A and E7 can stimulate
the methyltransferase activity of Dnmt1 in vitro. Our results provide
the first indication that viral oncoproteins bind and regulate Dnmt1
enzymatic activity. These observations open up the possibility that
this association may be used to control cellular proliferation pathways
and suggest a new mechanism by which small DNA tumour viruses can steer
cells through the cell cycle.

<>

<1>Burguener, G.F., Maldonado, M.J., Revale, S., Fernandez, Do.P.D., Rascovan, N., Vazquez, M., Farias, M.E., Marti, M.A., Turjanski, A.G.
<2>Draft Genome Sequence of the Polyextremophilic Halorubrum sp. Strain AJ67, Isolated from Hyperarsenic Lakes in the Argentinian Puna.
<3>Genome Announcements
<4>2
<5>e01096-13
<6>2014
<7>Halorubrum sp. strain AJ67, an extreme halophilic UV-resistant archaeon, was isolated from
Laguna Antofalla in the Argentinian Puna. The draft genome sequence
suggests the presence of potent enzyme candidates that are essential for survival
under multiple environmental extreme conditions, such as high UV radiation,
elevated salinity, and the presence of critical arsenic concentrations.

<>

<1>Burkhart, J.G., Burkhart, B.A., Sampson, K., Malling, H.V.
<2>Evidence for a previously undetected CpG methyl-directed restriction system in E.coli.
<3>Nucleic Acids Res.
<4>20
<5>4368
<6>1992
<7>
<>

<1>Burks, J.N., Wujick, C.T., Marshall, L.M., Kozarov, E.V., Progulske-Fox, A.
<2>Function of the porphyromonas gingivalis DNA adenine methylase in the expression of virulence.
<3>J. Dent. Res.
<4>81
<5>A-497
<6>2002
<7>Objectives: DNA adenine methylases (dam) modify specific sequences of DNA and thus regulate
the expression of certain genes in many species of bacteria.  Therefore, the dam gene is a
prime target for the development of broad-spectrum antibiotics and vaccines.  Our lab has
previously identified pgiM, a Porphyromonas gingivalis 381 (P. gingivalis) dam gene that
restores wild-type function to an Escherichia coli dam- mutant.  Recently, it was shown that
the dam gene of Salmonella typhimurium (S. typhimurium) regulates the expression of virulence
genes activated by the global regulator, PhoP.  dam-, avirulent strains of S. typhimurium
showed complete restoration of virulence when complemented with the S. typhimurium dam gene.
We hypothesized that pgiM may play a similar role in the regulation of virulence in P.
gingivalis.  Methods: To test this theory, we constructed the dam complementation plasmid,
pDC1. pDC1 is a derivative of pWKS30, a modified version of pBluescriptIIks, and contains a
1.1 kb region of P. gingivalis DNA harboring pgiM.  Subsequently, we transformed the non-polar
dam- mutant S. typhimurium MT2188D232 with pDC1.  Transformed cells were plated onto Luria
Bertani agar plates containing 50 ug/ml of ampicillin.  Analysis of plasmid preps performed on
five of the resulting clones confirmed the presence of pDC1.  Next, the complemented S.
typhimurium mutant clones were plated onto Luria Bertani agar plates containing 0.6 mg/ml of
2-aminopurine.  In addition, we used a 5' truncated form of pgiM in a single crossover
through homologous recombination, to construct a P. gingivalis dam- mutant.  Results: The
complemented clones grew on 2-aminopurine, indicating a restoration of the dam gene adenine
methylase in P. gingivalis irulence.  Conclusion: This data demonstrates that the P. ginivalis
dam gene can functionally complement the S. typhimurium dam gene.

<>

<1>Burland, V., Plunkett, G. III, Sofia, H.J., Daniels, D.L., Blattner, F.R.
<2>Analysis of the Escherichia coli genome VI: DNA sequence of the region from 92.8 through 100 minutes.
<3>Nucleic Acids Res.
<4>23
<5>2105-2119
<6>1995
<7>The 338.5 kb of the Escherichia coli genome described here together with previously described
segments bring the total of contiguous finished sequence of this genome to >1 Mb.  Of 319 open
reading frames found in this 338.5 kb segment, 147 (46%) are potential new genes.  The
positions of several genes which had been previously located here by mapping or partial
sequencing have been confirmed.  Several ORFs have functions suggested by similarities to
other characterized genes but cannot be assigned with certainty.  Fifteen of the ORFs of
unknown function had been previously sequenced.  Eight transfer RNAs are encoded in the region
and there are two grey holes in which no features were found.  The attachment site for phage
P4 and three insertion sequences were located.  The region was also analyzed for chi sites,
bend sites, REP elements and other repeats.  A computer search identified potential promoters
and tentative transcription units were assigned.  The occurrence of the rare tetramer CTAG was
analyzed in 1.6 Mb of contiguous E. coli sequence.  Hypotheses addressing the rarity and
distribution of CTAG are discussed.

<>

<1>Burland, V., Shao, Y., Perna, N.T., Plunkett, G., Sofia, H.J., Blattner, F.R.
<2>The complete DNA sequence and analysis of the large virulence plasmid of Escherichia coli O157:H7.
<3>Nucleic Acids Res.
<4>26
<5>4196-4204
<6>1998
<7>The complete DNA sequence of pO157, the large virulence plasmid of EHEC strain O157:H7 EDL
933, is presented.  The 92 kb F-like plasmid is composed of segments of putative virulence
genes in a framework of replication and maintenance regions, with seven insertion sequence
elements, located mostly at the boundaries of the virulence segments.  One hundred open
reading frames were identified, of which 19 were previously sequenced potential virulence
genes.  Forty-two ORFs were sufficiently similar to known proteins for suggested functions to
be assigned, and 22 had no convincing similarity with any known proteins.  Of the newly
identified genes, an unusually large ORF of 3169 amino acids has a putative cytotoxin active
site shared with the large clostridial toxin (LCT) family and proteins such as ToxA and B of
Clostridium difficile.  A conserved motif was detected that links the large ORF and the LCT
proteins with the OCH1 family of glycosyltransferases.  In the complete sequence, the mosaic
form can be observed at the levels of base composition, codon usage and gene organization.
Insights were obtained from patterns of DNA composition as well as the pathogenic and
'housekeeping' gene segments.  Evolutionary trees built from shared plasmid maintenance
genes show that even these genes have heterogeneous origins.

<>

<1>Burns, B.P., Gudhka, R.K., Neilan, B.A.
<2>Genome Sequence of the Halophilic Archaeon Halococcus hamelinensis.
<3>J. Bacteriol.
<4>194
<5>2100-2101
<6>2012
<7>Halococcus hamelinensis was isolated from hypersaline stromatolites in Shark Bay, Australia.
Here we report the genome sequence (3,133,046 bp) of H. hamelinensis,
which provides insights into the ecology, evolution, and adaptation of this novel
microorganism.

<>

<1>Burrus, V., Bontemps, C., Decaris, B., Guedon, G.
<2>Characterization of a novel type II restriction-modification system, Sth368I, encoded by the integrative element ICESt1 of Streptococcus thermophilus CNRZ368.
<3>Appl. Environ. Microbiol.
<4>67
<5>1522-1528
<6>2001
<7>A novel type II restriction and modification (R-M) system, Sth368I, which confers resistance
to ST84, was found in Streptococcus thermophilus CNRZ368 but not in the very closely related
strain A054. Partial sequencing of the integrative conjugative element ICESt1, carried by S.
thermophilus CNRZ368 but not by A054, revealed a divergent cluster of two genes, sth368IR and
sth368IM. The protein sequence encoded by sth368IR is related to the type II endonucleases
R.LlaKR2I and R.Sau3AI, which recognize and cleave the sequence 5'-GATC-3'. The protein
sequence encoded by sth368IM is very similar to numerous type II 5-methylcytosine
methyltransferases, including M.LlaKR2I and M.Sau3AI. Cell extracts of CNRZ368 but not A054
were found to cleave at the GATC site. Furthermore, the C residue of the sequence 5'-GATC-3'
was found to be methylated in CNRZ368 but not in A054. Cloning and integration of a copy of
sth368IR and sth368IM in the A054 chromosome confers on this strain phenotypes similar to
those of CNRZ368, i.e., phage resistance, endonuclease activity of cell extracts, and
methylation of the sequence 5'-GATC-3'. Disruption of sth368IR removes resistance and
restriction activity. We conclude that ICESt1 encodes an R-M system, Sth368I, which recognizes
the sequence 5'-GATC-3' and is related to the Sau3AI and LlaKR2I restriction systems.

<>

<1>Burrus, V., Quezada-Calvillo, R., Marrero, J., Waldor, M.K.
<2>SXT-Related Integrating Conjugative Element in New World Vibrio cholerae.
<3>Appl. Environ. Microbiol.
<4>72
<5>3054-3057
<6>2006
<7>SXT-related integrating conjugative elements (ICEs) became prevalent in
Asian Vibrio cholerae populations after V. cholerae O139 emerged. Here, we
describe an SXT-related ICE, ICEVchMex1, in a Mexican environmental V.
cholerae isolate. Identification of ICEVchMex1 represents the first
description of an SXT-related ICE in the Western Hemisphere. The
significant differences between the SXT and ICEVchMex1 genomes suggest
that these ICEs have evolved independently.

<>

<1>Burt, A., Koufopanou, V.
<2>Homing endonuclease genes: the rise and fall and rise again of a selfish element.
<3>Curr. Opin. Genet. Dev.
<4>14
<5>609-615
<6>2004
<7>Homing endonuclease genes (HEGs) are selfish genetic elements that spread by first cleaving
chromosomes that do not contain them and then getting copied across to the broken chromosome
as a byproduct of the repair process. The success of this strategy will depend on the
opportunities for homing - in other words, the frequency with which HEG(+) and HEG(-)
chromosomes come into contact - which varies widely among host taxa. HEGs are also unusual in
that the selection pressure for endonuclease function disappears if they become fixed in a
population, which makes them susceptible to degeneration and imposes a need for regular
horizontal transmission between species. HEGs will be selected to reduce the harm done to the
host organism, and this is expected to influence the evolution of their sequence specificity
and maturase functions. HEGs may also be domesticated by their hosts, and are currently being
put to human uses.

<>

<1>Buryanov, Y., Shevchuk, T.
<2>The use of prokaryotic DNA methyltransferases as experimental and analytical tools in modern biology.
<3>Anal. Biochem.
<4>338
<5>1-11
<6>2005
<7>Prokaryotic DNA methyltransferases (MTases) are used as experimental and research tools in
molecular biology and molecular genetics due to
their ability to recognize and transfer methyl groups to target bases
in specific DNA sequences. As a practical tool, prokaryotic DNA MTases
can be used in recombinant DNA technology for in vitro alteration and
enhancing of cleavage specificity of restriction endonucleases. The
ability of prokaryotic DNA MTases to methylate cytosine residues in
specific sequences, which are also methylated in eukaryotic DNA, makes
it possible to use them as analytical reagent for determination of the
site-specific level of methylation in eukaryotic DNA. In vivo DNA
methylation by prokaryotic DNA MTases is used in different techniques
for probing chromatin structure and protein-DNA interactions.
Additional prospects are opened by development of the methods of DNA
methylation targeted to predetermined DNA sequences by fusion of DNA
MTases to DNA binding proteins. This review will discuss the
application of prokaryotic DNA MTases of Type II in the methods and
approaches mentioned above.

<>

<1>Buryanov, Y.I., Baryshev, M.M., Kosykh, V.G., Bayev, A.A.
<2>The purification and characterization of BstNI modification methylase.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>211
<6>1989
<7>We have isolated and purified the BstNI modification methylase (M.BstNI) from
Bacillus stearothermophilus.  This enzyme catalyses the transfer of methyl
groups from AdoMet to CC(A/T)GG DNA sequences yielding N4-methylcytosine.  This
modification renders the DNA resistant to cleavage by both BstNI and EcoRII
restriction endonucleases.  At the same time M.BstNI is able to modify the DNA
previously methylated by EcoRII methylase while the DNA methylated by M.BstNI
is resistant to EcoRII methylation.  This property of two methylases can be
used for analysis of the 5-methylcytosine presence in plant DNA in the
CC(A/T)GG sequence.  We have partially purified M.BstNI by DEAE-cellulose,
phosphocellulose and hydroxylapatite column chromatography.  The enzyme has a
molecular weight of 60 kD as determined by gel-filtration and
SDS-polyacrylamide gel electrophoresis.  E. coli clones were isolated from
libraries of B. stearothermophilus DNA fragments in pUC19 by selecting for
self-modified molecules that were resistant to BstNI endonuclease digestion.

<>

<1>Buryanov, Y.I., Bogdarina, I.G., Bayev, A.A.
<2>Site specificity and chromatographic properties of E. coli K12 and EcoRII DNA-cytosine methylases.
<3>FEBS Lett.
<4>88
<5>251-254
<6>1978
<7>It has been shown that DNA methylation in vitro by DNA-cytosine methylase from E. coli K12
provides phage lambda DNA with complete resistance against EcoRII restriction endonuclease.
E. coli C DNA-cytosine methylase and E. coli MRE 600 DNA-cytosine methylase II also provide
lambda DNA with resistance against RII restriction in transfection experiments.  It is likely
that all these DNA-cytosine methylases can display the same or overlapping site specificity as
EcoRII DNA methylase.  For E. coli MRE 600 DNA-cytosine methylase II and E. coli C
DNA-cytosine methylation in vivo the major targets in DNA are the dinucleotide C-m5C and
trinucleotide C-m5C-T (m5C:5-methylcytosine).  These pyrimidine fragments of DNA are identical
to pyrimidine sequences of the DNA site modified by EcoRII DNA methylase:
5'...C-m5C-A-G-G...3'  3'...G-G-T-m5C-C...5' However, the pattern of DNA modification in
vitro by E. coli K12 DNA-cytosine methylase and RII DNA methylase and the only major target
sequences C-m5C-T for these two enzymes do not correspond to the in vivo methylation pattern
of E. coli K12 DNA and to the mode of RII DNA methylase action.  The aim of this communication
is the additional analysis of DNA sequences methylated in vitro by RII and E. coli K12
DNA-cytosine methylases.  The results reported here show that E. coli K12 DNA-cytosine
methylase and RII DNA methylase display the same site specificity which is identical to that
of E. coli C and E. coli MRE 600 DNA-cytosine methylases.  Our data on chromatographic
behavior of RII DNA methylase on phosphocellulose differ from those in (1).

<>

<1>Buryanov, Y.I., Bogdarina, I.G., Vagabova, L.M.
<2>DNA-cytosine methylation in Escherichia coli MRE 600 cells.
<3>Dokl. Akad. Nauk.
<4>230
<5>976-978
<6>1976
<7>
<>

<1>Buryanov, Y.I., Bogdarina, I.T., Baev, A.A.
<2>Isolation of DNA-cytosine methyltransferases from E. coli B (RII), E. coli K12 and E. coli K12 (RII).  Analysis of nucleotide sequences anlayzed in vitro.
<3>Dokl. Akad. Nauk.
<4>237
<5>465-468
<6>1977
<7>
<>

<1>Buryanov, Y.I., Kholodkov, O.A., Bogdarina, I.G., Neserenko, V.F., Zakharchenko, V.N., Chernov, A.P.
<2>On the ability of E. coli DNA methylases to modify denatured DNA's.
<3>Biokhimiia
<4>48
<5>1752-1754
<6>1983
<7>Adenine and cytosine DNA methylases from different strains of E. coli are able to methylate
denaturated and single-stranded DNA's.

<>

<1>Buryanov, Y.I., Nesterenko, V.F., Dyacheniko, G.P., Korunskii, O.F., Skryabin, G.K.
<2>Specificity of different fractions of DNA-cytosine-methylase from Escherichia coli MRE 600.
<3>Dokl. Akad. Nauk.
<4>227
<5>228-231
<6>1976
<7>
<>

<1>Buryanov, Y.I., Nesterenko, V.F., Kosykh, V.G., Baev, A.A.
<2>Different molecular structure of DNA methylases EcoRII and Eco MRE600 dcmII.
<3>Dokl. Akad. Nauk.
<4>257
<5>495-497
<6>1981
<7>The cells of most strains of Escherichia coli contain DNA methylases that exhibit site
specificity of the methylase EcoRII. DNA methylases of the EcoRII type provide the host
chromosome with complete protection against the restriction endonuclease EcoRII and the
rapidly amplified DNAs of phages and plasmids with partial protection against this
endonuclease. In contrast to the plasmid nature of the enzymes of modification and restriction
of EcoRII, DNA methylases of the EcoRII type are determined by a chromosomal gene of E. coli,
and there is no corresponding restriction endonuclease in E. coli cells.

<>

<1>Buryanov, Y.I., Nesterenko, V.F., Vagabova, L.M.
<2>The presence of two DNA-cytosine methylases in cells of Escherichia coli MRE600.
<3>Dokl. Akad. Nauk.
<4>227
<5>1472-1475
<6>1976
<7>None

<>

<1>Buryanov, Y.I., Shevchuk, T.V.
<2>DNA methyltransferases and structural-functional specificity of eukaryotic DNA modification.
<3>Biochemistry
<4>70
<5>730-742
<6>2005
<7>Properties of the main families of mammalian, plant, and fungal DNA methyltransferases are
considered. Structural-functional specificity of
eukaryotic genome sequences methylated by DNA methyltransferases is
characterized. The total methylation of cytosine in DNA sequences is
described, as well as its relation with RNA interference. Mechanisms of
regulation of expression and modulation of DNA methyltransferase
activity in the eukaryotic cell are discussed.

<>

<1>Buryanov, Y.I., Zakharchenko, N.S., Shevchuk, T.V., Bogdarina, I.G.
<2>Effect of the M.EcoRII methyltransferase-encoding gene on the phenotype of Nicotiana tabacum transgenic cells.
<3>Gene
<4>157
<5>283-287
<6>1995
<7>The EcoRII DNA methyltransferase (M.EcoRII; MTase) modifies a cytosine in the DNA sequence
CCWGG which contains a CNG methylation motif characteristic of plant DNA.  The gene (ecoRIIM)
encoding this MTase has been cloned into the T-DNA of the wild-type Agrobacterium Ti-plasmid
pTiC58 downstream from the plant expression nopaline synthase-encoding gene promoter.
Nicotiana tabacum cells have been transformed with Agrobacterium tumefaciens harboring this
recombinant Ti-plasmid.  The primary transformed tabacco tissue line has given rise to novel
stable lines which are morphologically distinctive.  Southern hybridization analysis of all
transformed tissue lines has shown the presence, in each of them, of ecoRIIM.  The tissue
studied differed in morphology in callus culture, dependence on phytohormones and the ability
to synthesize nopaline.

<>

<1>Buryanov, Y.I., Zakharenko, V.N., Baev, A.A.
<2>Isolation, purification and properties of adenine DNA methylase Eco dam.
<3>Dokl. Akad. Nauk.
<4>259
<5>1492-1495
<6>1981
<7>
<>

<1>Buryanov, Y.I., Zinovev, V.V., Gorbunov, Y.A., Tuzikov, F.V., Rechkunova, N.I., Malygin, E.G., Bayev, A.A.
<2>Interaction of the Eco Dam methyltransferase with synthetic oligodeoxyribonucleotides.
<3>Gene
<4>74
<5>67-69
<6>1988
<7>Meeting Abstract

<>

<1>Buryanov, Y.I., Zinoviev, V.V., Vienozhinskis, M.T., Malygin, E.G., Nesterenko, V.F., Popov, S.G., Gorbunov, Y.A.
<2>Does the DNA methylase Eco dam pair nucleotide sequences to form site-specific duplexes?
<3>FEBS Lett.
<4>168
<5>166-168
<6>1984
<7>The Eco dam methylase is active on denatured DNA and single-stranded synthetic
oligonucleotides containing GATC sites.  The results suggest that on interaction with
single-stranded oligonucleotides the Eco dam methylase is able to form a duplex structure
within the GATC site, and that this duplex site is a substrate for enzyme.

<>

<1>Busquets, A. et al.
<2>Draft Genome Sequence of Pseudomonas azotifigens Strain DSM 17556T (6H33bT), a Nitrogen Fixer Strain Isolated from a Compost Pile.
<3>Genome Announcements
<4>1
<5>e00893-13
<6>2013
<7>Pseudomonas azotifigens strain 6H33b(T) is a nitrogen fixer isolated from a hyperthermal
compost pile in 2005 by Hatayama and collaborators. Here we report
the draft genome, which has an estimated size of 5.0 Mb, exhibits an average G+C
content of 66.73%, and is predicted to encode 4,536 protein-coding genes and 100
RNA genes.

<>

<1>Busquets, A., Pena, A., Gomila, M., Bosch, R., Nogales, B., Garcia-Valdes, E., Lalucat, J., Bennasar, A.
<2>Genome Sequence of Pseudomonas stutzeri Strain JM300 (DSM 10701), a Soil Isolate  and Model Organism for Natural Transformation.
<3>J. Bacteriol.
<4>194
<5>5477-5478
<6>2012
<7>Pseudomonas stutzeri strain JM300 (DSM 10701) is a denitrifying soil isolate and  a model
organism for natural transformation in bacteria. Here we report the first
complete genome sequence of JM300, the reference strain of genomovar 8 for the
species.

<>

<1>Busquets, A., Pena, A., Gomila, M., Mayol, J., Bosch, R., Nogales, B., Garcia-Valdes, E., Bennasar, A., Lalucat, J.
<2>Draft Genome Sequence of Pseudomonas stutzeri Strain B1SMN1, a Nitrogen-Fixing and Naphthalene-Degrading Strain Isolated from Wastewater.
<3>Genome Announcements
<4>1
<5>e00584-13
<6>2013
<7>Pseudomonas stutzeri strain B1SMN1 is a naphthalene-degrading and simultaneously
nitrogen-fixing strain isolated from a wastewater sample taken at a lagooning
treatment plant in Menorca (Balearic Islands, Spain). Here we report the draft
genome sequence of P. stutzeri B1SMN1. It is composed of a chromosome of an
estimated size of 5.2 Mb and two plasmids of 44,324 bp and 56,118 bp.

<>

<1>Busslinger, M., deBoer, E., Wright, S., Grosveld, F.G., Flavell, R.A.
<2>The sequence GGCmCGG is resistant to MspI cleavage.
<3>Nucleic Acids Res.
<4>11
<5>3559-3569
<6>1983
<7>MspI essentially fails to cut the sequence GGCmCGG at enzyme concentrations
which give total digestion of CCGG, CmCGG and GGCCGG sites.  This result
explains why certain sites in mammalian DNA are resistant to both MspI and HhaI
and shows that this results from an idiosyncracy of MspI rather than a novel
form of DNA methylation at this site in mammalian cells.

<>

<1>Butkus, V., Bitinaite, J., Kersulyte, D., Janulaitis, A.
<2>A new restriction endonuclease Eco31I recognizing a non-palindromic sequence.
<3>Biochim. Biophys. Acta
<4>826
<5>208-212
<6>1985
<7>A restriction endonuclease with a novel site-specificity has been isolated from the
Escherichia coli strain RFL31.  The nucleotide sequences around a single Eco311 cut on pBR322
DNA and two cuts of lambda DNA have been compared.  A common 5'GAGACC/3'CTCTGG sequence
occurs near each cleavage site.  Precise mapping of the cleavages in both DNA strands places
the cuts five nucleotides to the left of the upper sequence and one nucleotide to the left of
the lower sequence.  This enabled us to deduce the following recognition and cleavage
specificity of Eco31I:
5' GGTCTCN/
3' CCAGAGN NNNN/.

<>

<1>Butkus, V., Kazlauskiene, R., Gilvonauskaite, R., Petrusyte, M., Janulaitis, A.
<2>Determination of substrate specificity of restriction endonuclease Eco78I.
<3>Bioorg. Khim.
<4>11
<5>1572-1573
<6>1985
<7>The recognition sequence and cleavage point of restriction endonuclease Eco78I
have been determined at 5'-GGCG^CC-.  There are several known enzymes
recognizing the same sequence, although the prototype NarI and isoschizomers
NdaI and NunII cleave the substrate to produce 5'-protruding ends, whereas
cleavage with isoschizomer BbeI results in 3'-protruding ends.  Therefore,
restrictase Eco78I, generating flush ends, may be regarded as an enzyme with
new specificity among the restriction endonucleases recognizing the
5'-GGCGCC-sequence.

<>

<1>Butkus, V., Klimasauskas, S., Kersulyte, D., Vaitkevicius, D., Lebionka, A., Janulaitis, A.
<2>Investigation of restriction-modification enzymes from M. varians RFL19 with a new type of specificity toward modification of substrate.
<3>Nucleic Acids Res.
<4>13
<5>5727-5746
<6>1985
<7>The characterization of MvaI restriction-modification enzymes, isolated from Micrococcus
varians RFL19, is reported. Both enzymes recognize the 5'CC^(A/T)GG nucleotide sequence. The
endonuclease cleaves the sequence at the position indicated by the arrow, whereas the
methylase modifies the internal cytosine, yielding N4-methylcytosine. This type of
modification protects the substrate from R.MvaI cleavage. 5-methylcytosine in the same
position of the recognition sequence does not protect the substrate from R.MvaI cleavage.
R.MvaI proved to be the first example of a restriction endonuclease differentiating the
position of the methyl group in the heterocyclic ring of cytosine, located in the same site of
the recognition sequence. M.MvaI modifies DNA dcm+ in vitro yielding N4, 5-dimethylcytosine.
N4-methylcytosine cannot be differentiated from cytosine using the Maxam-Gilbert DNA
sequencing procedure.

<>

<1>Butkus, V., Klimasauskas, S., Petrauskiene, L., Maneliene, Z.
<2>The effects of N4- and C5-methylation of cytosine on DNA physical properties and restriction endonuclease action.
<3>Metabolism and Enzymology of Nucleic Acids, Publishing House of the Slovak Akademy of Science, Zelinka, J., Balan, J., Bratislava
<4>6
<5>119-127
<6>1987
<7>Not long ago a new type of methylase, generating N4-methylcytosine (4mC) in DNA
was isolated in our laboratory from Bacillus centrosporus (Janulaitis et al.,
1983).  At present a number of the 4mC type methylases are isolated (Janulaitis
et al., 1984; Butkus et al., 1985,1987) and a wide spread of N4-methylcytosine
in DNA of thermophilic and mesophilic bacteria is well documented (Ehrlich et
al., 1985,1987).  Since the two methylated cytosine bases occurred in native
DNA, a comparison of the effects of N4- and C5-methylation on DNA
physico-chemical properties and enzyme action seems to be of great interest.

<>

<1>Butkus, V., Klimasauskas, S., Petrauskiene, L., Maneliene, Z., Janulaitis, A., Minchenkova, L.E., Schyolkina, A.K.
<2>Synthesis and physical characterization of DNA fragments containing N4-methylcytosine and 5-methylcytosine.
<3>Nucleic Acids Res.
<4>15
<5>8467-8478
<6>1987
<7>The synthesis of N4-methyl-2'-deoxycytidine and its fully protected
mononucleotide, suitable for the oligonucleotide synthesis by phosphotriester
method is described.  A set of octanucleotides -d(CGCGCGCG), d(CG5mCGCGCG),
d(CG4mCGCGCG) and dodecanucleotides -d(GGACCCGGGTCC), d(GGA5mCCCGGGTCC),
d(GGA4mCCCGGGTCC) has been synthesized in a solution.  Physical
characterization of the oligonucleotide duplexes by means of UV and CD
spectrometry provides the evidence that 4mC similarly to 5mC favours the B-->Z
transition, although both of these methylated cytosines inhibit the B-->A
conformational change.  N4-Methylcytosine in contrast to 5-methylcytosine
reduces the DNA double helix thermal stability.

<>

<1>Butkus, V., Klimasauskas, S., Petrauskiene, L., Maneliene, Z., Lebionka, A., Janulaitis, A.
<2>Interaction of AluI, Cfr6I and PvuII restriction-modification enzymes with substrates containing either N4-methylcytosine or 5-methylcytosine.
<3>Biochim. Biophys. Acta
<4>909
<5>201-207
<6>1987
<7>The cleavage specificity of R.Cfr6I, an isoschizomer of PvuII restriction
endonuclease was determined to be 5'CAG^CTG and the methylation specificity of
Cfr6I and PvuII methylases, 5'CAG4mCTG.  Thus, M.Cfr6I and M.PvuII are new
additions to the list of methylases with N4-methylcytosine specificity.
Neither of the above RM enzymes acts on the substrates containing either
N4-methylcytosine or 5-methylcytosine in a cognate methylation position.

<>

<1>Butkus, V., Klimasauskas, S., Petrauskiene, L., Maneliene, Z., Minchenkova, L.E., Schyolkina, A.K., Janulaitis, A.
<2>The effects of N4- and C5-methylation of cytosine on DNA physical properties and restriction endonuclease action.
<3>Metabolism and Enzymology of Nucleic Acids, Plenum, Zelinka, J., Balan, J., New York
<4>7
<5>73-78
<6>1988
<7>Not long ago a new type of methylase, generating N4-methylcytosine (4mC) in DNA
was isolated in our laboratory from Bacillus centrosporus (Janulaitis et al.,
1983).  At present a number of the 4mC type methylases are isolated (Janulaitis
et al., 1984; Butkus et al., 1985; 1987) and the widespread occurrence of
N4-methylcytosine in DNA of thermophilic and mesophilic bacteria is documented
(Ehrlich et al., 1985, 1987).  Since the two methylated cytosine bases occur in
native DNA, a comparison of the effects of N4- and C5-methylation on DNA
physico-chemical properties and enzyme action seems to be of great interest.

<>

<1>Butkus, V., Petrauskiene, L., Maneliene, Z., Klimasauskas, S., Laucys, V., Janulaitis, A.
<2>Cleavage of methylated CCCGGG sequences containing either N4-methylcytosine or 5-methylcytosine with MspI, HpaII, SmaI, XmaI and Cfr9I restriction endonucleases.
<3>Nucleic Acids Res.
<4>15
<5>7091-7102
<6>1987
<7>The cleavage specificity of R.Cfr9I was determined to be C/CCGGG whereas the
methylation specificity of M.Cfr9I was C4mCCGGG.  The action of MspI, HpaII,
SmaI, XmaI and Cfr9I restriction endonucleases on an unmethylated parent
d(GGACCCGGGTCC) dodecanucleotide duplex and a set of oligonucleotide duplexes,
containing all possible substitutions of either 4mC or 5mC for C in the CCCGGG
sequence, was investigated.  It was found that 4mC methylation, in contrast to
5mC, renders the CCCGGG site resistant to practically all the investigated
endonucleases.  The cleavage of methylated substrates with restriction
endonucleases is discussed.

<>

<1>Butkus, V.V., Klimasauskas, S.J., Janulaitis, A.
<2>Analysis of products of DNA modification by methylases:  A procedure for the determination of 5- and N4-methylcytosines in DNA.
<3>Anal. Biochem.
<4>148
<5>194-198
<6>1985
<7>Although many different methods are used for the identification of methylated heterocyclic
bases in DNA not all of them possess the ability to discriminate N4-methylcytosine (m4C) and
5-methylcytosine (m5C).  Therefore, some of the methods need additional reexamination.  This
paper reinvestigates some chromatographic systems (thin-layer chromatography, paper
chromatography, electrophoresis) most widely used in the analysis of minor bases occuring in
nucleic acids according to their ability to separate m4C and m5C.  A simple procedure for the
preparation of the sample and a chromatographic system for its analysis was developed.  The
recommended chromatographic systems may be used for the simultaneous separation of not only
m4C and m5C but also both methylated cytosines together with N6-methyladenine and
7-methylguanine.

<>

<1>Butkus, V.V., Petrusyte, M.P., Janulaitis, A.
<2>Determination of substrate specificity of Eco47I and Eco52I restriction endonucleases.
<3>Bioorg. Khim.
<4>11
<5>987-988
<6>1985
<7>Cleavage sites for Eco47I and Eco52I restriction endonucleases, which are
isoschizomers of AvaII and XmaIII, respectively, have been structurally
elucidated.

<>

<1>Butler, C.A., Gotschlich, E.C.
<2>High-frequency mobilization of broad-host-range plasmids into Neisseria gonorrhoeae requires methylation in the donor.
<3>J. Bacteriol.
<4>173
<5>5793-5799
<6>1991
<7>Antibiotic resistance in Neisseria gonorrhoeae has been associated with the
acquisition of R plasmids from heterologous organisms.  The broad-host-range
plasmids of incompatibility groups P (IncP) and Q (IncQ) have played a role in
this genetic exchange in nature.  We have utilized derivatives of RSF1010
(IncQ) and RP1 (IncP) to demonstrate that the plethora of restriction barriers
associated with the gonococci markedly reduces mobilization of plasmids from
Escherichia coli into strains F62 and PGH 3-2.  Partially purified restriction
endonucleases from these gonococcal strains can digest RSF1010 in vitro.
Protection of RSF1010-km from digestion by gonococcal enzymes purified from
strain F62 is observed when the plasmid is isolated from E. coli containing a
coresident plasmid, pCAL7.  Plasmid pCAL7 produces a 5'-mCG-3' cytosine
methylase (M.SssI).  The M.SssI methylase only partially protects RSF1010-km
from digestion by restriction enzymes from strain PGH 3-2.  Total protection of
RSF1010-km from PGH 3-2 restriction requires both pCAL7 and a second coresident
plasmid, pFnuDI, which produces a 5'-GGmCC-3' cytosine methylase.  When both
F62 and PGH 3-2 are utilized as recipients in heterospecific matings with E.
coli, mobilization of RSF1010 from strains containing the appropriate
methylases into the gonococci occurs at frequencies 4 orders of magnitude
higher than from strains without the methylases.  Thus, protection of RSF1010
from gonococcal restriction enzymes in vitro correlates with an increase in the
conjugal frequency.  These data indicate that restriction is a major barrier
against efficient conjugal transfer between N. gonorrhoeae and heterologous
hosts.

<>

<1>Butler, D., Fitzgerald, G.F.
<2>Transcriptional analysis and regulation of expression of the ScrFI restriction-modification system of Lactococcus lactis subsp. cremoris UC503.
<3>J. Bacteriol.
<4>183
<5>4668-4673
<6>2001
<7>ScrFI is a type II restriction-modification system from Lactococcus lactis which recognizes
the nucleotide sequence 5'-CC^NGG-3', cleaving at the point indicated by the arrow, and it
comprises an endonuclease gene that is flanked on either side by genes encoding two
5-methylcytosine methylases. An open reading frame (orfX) of unknown function is located
immediately upstream of these genes. In this study Northern analysis was performed, and it
revealed that orfX, scrFIBM, and scrFIR are cotranscribed as a single polygenic mRNA molecule,
while scrFIAM is transcribed independently. 5' extension analysis indicated that the start
site for the scrFIAM promoter was a thymine located 4 bp downstream of the -10 motif. The
transcriptional start site for the orfX promoter was also found to be a thymine which is more
atypically located 24 bp downstream of the -10 motif proximal to the start codon. A
helix-turn-helix motif was identified at the N-terminal end of one of the methylases
(M.ScrFIA). In order to determine if this motif played a role in regulation of the ScrFI
locus, M.ScrFIA was purified. It was then employed in gel retardation assays using fragments
containing the two promoters found on the ScrFI operon, one located upstream of orfX and the
other located just upstream of scrFIAM. M.ScrFIA was found to bind to the promoter region
upstream of the gene encoding it, indicating that it may have a regulatory role. In further
studies the two putative promoters were introduced into a vector (pAK80) upstream of a
promoterless lacZ gene, and cloned fragments of the ScrFI locus were introduced in trans with
each of these promoter constructs to investigate the effect on promoter activity. These
results implicated M.ScrFIA in regulation of both promoters on the ScrFI locus.

<>

<1>Butler, M.I., Stockwell, P.A., Black, M.A., Day, R.C., Lamont, I.L., Poulter, R.T.M.
<2>Pseudomonas syringae pv. actinidiae from Recent Outbreaks of Kiwifruit Bacterial Canker Belong to Different Clones that Originated in China.
<3>PLoS ONE
<4>8
<5>e57464
<6>2013
<7>A recently emerged plant disease, bacterial canker of kiwifruit (Actinidia deliciosa and A.
chinensis), is caused by Pseudomonas syringae pv. actinidiae (PSA).  The disease was first
reported in China nad Japan in the 1980s.  A severe breakout of PSA began in Italy in 2008 and
has spread to other European countries.  PSA was found in both New Zealand and Chile in 2010.
To study the evolution of the pathogen and analyse the transmission of PSA between countries,
genomes of strands from China and Japan (where the genus Actinidia is endemic)m Italy, New
Zealand and Chile were sequenced.  The genomes of PSA strains are very similar.  However, all
strains from New Zealand share several single nucleotide polymorphisms (SNPs) that distinguish
them from all other PSA strains.  Similarly, all the PSA strains from the 2008 Italian
outbreak form a distinct clonal group and those from Chile form a third group.  In addition to
the rare SNPs present in the core genones, there is abundant genetic diversity in a genomic
island that is part of the accessory genome.  The island from several Chinese strains is
almost identical to the island present in the New Zealand strains. The island from a different
Chinese strain is identical to the island prsent in the strains from the recent Italian
outbreak.  The Chilean strains of PSA carry a third variant of this island.  These genome
islands are integrative conjugative elements (ICEs).  Sequencing of these ICEs provides
evidence of three recent horizontal transmissions of ICE from other strains of Pseudomonas
syringae to PSA.  The analyses of the core genome SNPs and the ICEs, combined with disease
history, all support the hypothesis of an independent Chinese origin for both the Italian and
the New Zealand outbreaks and suggest the Chilean strains also originate from China.

<>

<1>Butler-Wu, S.M., Sengupta, D.J., Kittichotirat, W., Matsen, F.A., Bumgarner, R.E.
<2>Genome sequence of a novel species, Propionibacterium humerusii.
<3>J. Bacteriol.
<4>193
<5>3678
<6>2011
<7>As part of a larger project to sequence multiple clinical isolates of Propionibacterium acnes,
we have produced a draft genome sequence of a
novel Propionibacterium species that is closely related to, yet distinct
(by sequence) from P. acnes. We have tentatively named this new species
Propionibacterium humerusii.

<>

<1>Butterer, A., Pernstich, C., Smith, R.M., Sobott, F., Szczelkun, M.D., Toth, J.
<2>Type III restriction endonucleases are heterotrimeric: comprising one helicase-nuclease subunit and a dimeric methyltransferase that binds only one  specific DNA.
<3>Nucleic Acids Res.
<4>42
<5>5139-5150
<6>2014
<7>Fundamental aspects of the biochemistry of Type III restriction endonucleases remain
unresolved despite being characterized by numerous research groups in the
past decades. One such feature is the subunit stoichiometry of these
hetero-oligomeric enzyme complexes, which has important implications for the
reaction mechanism. In this study, we present a series of results obtained by
native mass spectrometry and size exclusion chromatography with multi-angle light
scattering consistent with a 1:2 ratio of Res to Mod subunits in the EcoP15I,
EcoPI and PstII complexes as the main holoenzyme species and a 1:1 stoichiometry
of specific DNA (sDNA) binding by EcoP15I and EcoPI. Our data are also consistent
with a model where ATP hydrolysis activated by recognition site binding leads to
release of the enzyme from the site, dissociation from the substrate via a free
DNA end and cleavage of the DNA. These results are discussed critically in the
light of the published literature, aiming to resolve controversies and discuss
consequences in terms of the reaction mechanism.

<>

<1>Byrne-Bailey, K.G., Coates, J.D.
<2>Complete Genome Sequence of the Anaerobic Perchlorate-Reducing Bacterium Azospira suillum Strain PS.
<3>J. Bacteriol.
<4>194
<5>2767-2768
<6>2012
<7>Azospira suillum strain PS (formally Dechlorosoma suillum strain PS) is a metabolically
versatile betaproteobacterium first identified for its ability to
grow by dissimilatory reduction of perchlorate and chlorate [denoted
(per)chlorate]. Together with Dechloromonas species, these two genera represent
the dominant (per)chlorate-reducing bacteria in mesophilic freshwater
environments. In addition to (per)chlorate reduction, A. suillum is capable of
the anaerobic oxidation of humic substances and is the first anaerobic
nitrate-dependent Fe(II) oxidizer outside the Diaphorobacter and Acidovorax
genera for which there is a completed genome sequence.

<>

<1>Byrne-Bailey, K.G., Weber, K.A., Chair, A.H., Bose, S., Knox, T., Spanbauer, T.L., Chertkov, O., Coates, J.D.
<2>Completed genome sequence of the anaerobic iron oxidizing bacterium Acidovorax ebreus strain TPSY.
<3>J. Bacteriol.
<4>192
<5>1475-1476
<6>2009
<7>Acidovorax ebreus strain TPSY is the first anaerobic nitrate-dependent Fe(II) oxidizer for
which there is a completed genome sequence. Preliminary protein annotation revealed an
organism optimized for survival in a complex environmental system. Here, we briefly report the
completed and annotated genome sequence of strain TPSY.

<>

<1>Byrne-Bailey, K.G., Weber, K.A., Coates, J.D.
<2>Draft Genome Sequence of the Anaerobic, Nitrate-Dependent, Fe(II)-Oxidizing Bacterium Pseudogulbenkiania ferrooxidans Strain 2002.
<3>J. Bacteriol.
<4>194
<5>2400-2401
<6>2012
<7>Pseudogulbenkiania ferrooxidans strain 2002 was isolated as a lithoautotrophic,
Fe(II)-oxidizing, nitrate-reducing bacterium. Phylogenetically, it is in a clade
within the family Neisseriaceae in the order Nessieriales of the class
Betaproteobacteria. It is anticipated that comparative genomic analysis of this
strain with other nitrate-dependent, Fe(II)-oxidizing bacteria will aid in the
elucidation of the genetics and biochemistry underlying this critically important
geochemical metabolism.

<>

<1>Byrne-Bailey, K.G., Wrighton, K.C., Melnyk, R.A., Agbo, P., Hazen, T.C., Coates, J.D.
<2>Complete genome sequence of the electricity-producing Thermincola potens strain JR.
<3>J. Bacteriol.
<4>192
<5>4078-4079
<6>2010
<7>Thermincola potens strain JR is one of the first Gram-positive dissimilatory metal reducing
bacteria (DMRB) for which there is a draft
genome sequence. Consistent with the physiology of this organism,
preliminary annotation revealed an abundance of multi-heme c-type
cytochromes that are putatively associated with the periplasm and cell
surface in a Gram-positive bacterium. Here we report the draft genome
sequence of strain JR.

<>

<1>Cab-Barrera, E.L., Barrera-Saldana, H.A.
<2>A general method to optimize the amount of enzyme in restriction and DNA modification reactions using the beta galactosidase blue-white plaques assay.
<3>Biotechniques
<4>7
<5>132-135
<6>1989
<7>We propose a simple and economical method for assaying the activity of
restriction and other modifying enzymes.  The method involves assaying the use
of the blue and white colored phenotypes of bacterial colonies obtained by
digesting the polylinker sequence of M13 bacteriophage vectors followed by
transformation in appropriate strains on X-gal/IPTG plates.  In conjunction
with restriction enzymes and DNA ligases, the method can evaluate polymerase
activity and can be applied to test 3'-5' exonuclease activities such as that
of T4 DNA polymerase, without having to use expensive radioisotopes.  We
describe its application in the assessment of restriction enzymes, DNA ligase
and DNA polymerase activities.

<>

<1>Caballero, J.I., Zerillo, M.M., Snelling, J., Boucher, C., Tisserat, N.
<2>Genome Sequence of Xanthomonas arboricola pv. Corylina, Isolated from Turkish Filbert in Colorado.
<3>Genome Announcements
<4>1
<5>e00246-13
<6>2013
<7>Previously, we reported the isolation of a bacterium producing leaf spots in Turkish filbert.
Here, we present the draft genome assembly of the bacterium identified as Xanthomonas
arboricola pv. corylina. To our knowledge, this is the first published genome of this pathovar
of X. arboricola.

<>

<1>Cabria, G.L., Argayosa, V.B., Lazaro, J.E., Argayosa, A.M., Arcilla, C.A.
<2>Draft Genome Sequence of Haloalkaliphilic Exiguobacterium sp. Strain AB2 from Manleluag Ophiolitic Spring, Philippines.
<3>Genome Announcements
<4>2
<5>e00840-14
<6>2014
<7>Exiguobacterium sp. AB2 is a haloalkaliphilic bacterium isolated from a hyperalkaline spring
in Manleluag, Pangasinan, Philippines. Sequencing of
bacterial DNA assembled a 2.85 MB draft genome. Analysis suggests the presence of
genes for tolerance to stresses such as elevated pH and salt concentrations and
toxic metals.

<>

<1>Caceres, O., Montenegro, J., Padilla, C., Tarazona, D., Bailon, H., Garcia, P., Cespedes, M., Valencia, P., Guio, H.
<2>Whole-Genome Sequencing and Comparative Analysis of Yersinia pestis, the Causative Agent of a Plague Outbreak in Northern Peru.
<3>Genome Announcements
<4>1
<5>e00249-12
<6>2013
<7>The plague is a zoonotic disease caused by the bacterium . Here, we report the complete genome
sequence of the strain INS, which was isolated from swollen lymph
gland aspirate (bubo aspirate) of an infected patient from a pneumonic outbreak
in 2010 in northern Peru.

<>

<1>Cadillo-Quiroz, H., Didelot, X., Held, N.L., Herrera, A., Darling, A., Reno, M.L., Krause, D.J., Whitaker, R.J.
<2>Patterns of gene flow define species of thermophilic archaea.
<3>PLoS Biology
<4>10
<5>E1001265
<6>2012
<7>Despite a growing appreciation of their vast diversity in nature, mechanisms of
speciation are poorly understood in Bacteria and Archaea. Here we use
high-throughput genome sequencing to identify ongoing speciation in the
thermoacidophilic Archaeon Sulfolobus islandicus. Patterns of homologous gene
flow among genomes of 12 strains from a single hot spring in Kamchatka, Russia,
demonstrate higher levels of gene flow within than between two persistent,
coexisting groups, demonstrating that these microorganisms fit the biological
species concept. Furthermore, rates of gene flow between two species are
decreasing over time in a manner consistent with incipient speciation. Unlike
other microorganisms investigated, we do not observe a relationship between
genetic divergence and frequency of recombination along a chromosome, or other
physical mechanisms that would reduce gene flow between lineages. Each species
has its own genetic island encoding unique physiological functions and a unique
growth phenotype that may be indicative of ecological specialization. Genetic
differentiation between these coexisting groups occurs in large genomic
"continents," indicating the topology of genomic divergence during speciation is
not uniform and is not associated with a single locus under strong diversifying
selection. These data support a model where species do not require physical
barriers to gene flow but are maintained by ecological differentiation.

<>

<1>Cai, L., Shao, M.F., Zhang, T.
<2>Non-contiguous finished genome sequence and description of Sulfurimonas hongkongensis sp. nov., a strictly anaerobic denitrifying, hydrogen- and  sulfur-oxidizing chemolithoautotroph isolated from marine sediment.
<3>Standards in Genomic Sciences
<4>9
<5>1302-1310
<6>2014
<7>Here, we report a type strain AST-10 representing a novel species Sulfurimonas hongkongensis
within Epsilonproteobacteria, which is involved in marine
sedimentary sulfur oxidation and denitrification. Strain AST-10(T) (= DSM
22096(T) = JCM 18418(T)) was isolated from the coastal sediment at the Kai Tak
Approach Channel connected to Victoria Harbour in Hong Kong. It grew
chemolithoautotrophically using thiosulfate, sulfide or hydrogen as the sole
electron donor and nitrate as the electron acceptor under anoxic conditions. It
was rod-shaped and grew at 15-35 degrees C (optimum at 30 degrees C), pH 6.5-8.5
(optimum at 7.0-7.5), and 10-60 g L(-1) NaCl (optimum at 30 g L(-1)). Genome
sequencing and annotation of strain AST-10(T) showed a 2,302,023 bp genome size,
with 34.9% GC content, 2,290 protein-coding genes, and 42 RNA genes, including 3
rRNA genes.

<>

<1>Cai, L., Tan, D., Aibaidula, G., Dong, X.R., Chen, J.C., Tian, W.D., Chen, G.Q.
<2>Comparative genomics study of polyhydroxyalkanoates (PHA) and ectoine relevant genes from Halomonas sp. TD01 revealed extensive horizontal gene transfer events and co-evolutionary relationships.
<3>Microb. Cell Fact.
<4>10
<5>88
<6>2011
<7>BACKGROUND: Halophilic bacteria have shown their significance in industrial
production of polyhydroxyalkanoates (PHA) and are gaining more attention for
genetic engineering modification. Yet, little information on the genomics and PHA
related genes from halophilic bacteria have been disclosed so far. RESULTS: The
draft genome of moderately halophilic bacterium, Halomonas sp. TD01, a strain of
great potential for industrial production of short-chain-length
polyhydroxyalkanoates (PHA), was analyzed through computational methods to reveal
the osmoregulation mechanism and the evolutionary relationship of the enzymes
relevant to PHA and ectoine syntheses. Genes involved in the metabolism of PHA
and osmolytes were annotated and studied in silico. Although PHA synthase,
depolymerase, regulator/repressor and phasin were all involved in PHA metabolic
pathways, they demonstrated different horizontal gene transfer (HGT) events
between the genomes of different strains. In contrast, co-occurrence of ectoine
genes in the same genome was more frequently observed, and ectoine genes were
more likely under coincidental horizontal gene transfer than PHA related genes.
In addition, the adjacent organization of the homologues of PHA synthase phaC1
and PHA granule binding protein phaP was conserved in the strain TD01, which was
also observed in some halophiles and non-halophiles exclusively from
gamma-proteobacteria. In contrast to haloarchaea, the proteome of Halomonas sp.
TD01 did not show obvious inclination towards acidity relative to non-halophilic
Escherichia coli MG1655, which signified that Halomonas sp. TD01 preferred the
accumulation of organic osmolytes to ions in order to balance the intracellular
osmotic pressure with the environment. CONCLUSIONS: The accessibility of genome
information would facilitate research on the genetic engineering of halophilic
bacteria including Halomonas sp. TD01.

<>

<1>Cai, L., Zhang, T.
<2>Genome of Bacillus macauensis ZFHKF-1, a Long-Chain-Forming Bacterium.
<3>J. Bacteriol.
<4>194
<5>4780
<6>2012
<7>Here, we report the draft genome sequence of Bacillus macauensis ZFHKF-1, a novel long-chain
bacterium previously isolated and identified by us (Zhang T, Fan XJ,
Hanada S, Kamagata Y, Fang HHP, J. Syst. Evol. Microbiol. 56:349-353, 2006). The
genome provides basic genetic information to understand this particular species
and explore the potential mechanism of long-chain formation. The type strain is
ZFHKF-1 (= JCM 13285 = DSM 17262).

<>

<1>Cai, M., Chen, W.M., Nie, Y., Chi, C.Q., Wang, Y.N., Tang, Y.Q., Li, G.Y., Wu, X.L.
<2>Complete Genome Sequence of Amycolicicoccus subflavus DQS3-9A1T, an actinomycete isolated from crude oil-polluted soil.
<3>J. Bacteriol.
<4>193
<5>4538-4539
<6>2011
<7>Amycolicicoccus subflavus DQS3-9A1(T) , isolated from crude oil polluted soil in the Daqing
Oilfield in China , is a type strain of a newly
published novel species in the novel genus Amycolicicoccus. Here we report
the complete genome of DQS3-9A1(T) and genes associated with oil-polluted
environment.

<>

<1>Cailliez-Grimal, C., Chaillou, S., Anba-Mondoloni, J., Loux, V., Afzal, M.I., Rahman, A., Kergourlay, G., Champomier-Verges, M.C., Zagorec, M., Dalgaard, P., Leisner, J.J., Prevost, H., Revol-Junelles, A.M., Borges, F.
<2>Complete Chromosome Sequence of Carnobacterium maltaromaticum LMA 28.
<3>Genome Announcements
<4>1
<5>e00115-12
<6>2013
<7>Within the lactic acid bacterium genus Carnobacterium, Carnobacterium maltaromaticum is one of
the most frequently isolated species from natural
environments and food. It potentially plays a major role in food product
biopreservation. We report here on the 3.649-Mb chromosome sequence of C.
maltaromaticum LMA 28, which was isolated from ripened soft cheese.

<>

<1>Cajo, G.C., Brcic-Kostic, K., Ivancic, I., Trgovcevic, Z., Salaj-Smic, E.
<2>Inactivation of the EcoKI restriction in UV-irradiated Escherichia coli: The role of RecBCD enzyme.
<3>Periodicum Biologorum
<4>103
<5>157-161
<6>2001
<7>Background and Purpose: UV-induced restriction alleviation (UV-induced RA) has been known for
a long time, but the mechanism underlying this
phenomenon is not known. UV-induced RA depends on the induction of the
SOS response and on the functional recBCD genes. The experiments
described in this study attempt to clarify the role of the RecBCD
enzyme in UV-induced RA
Materials and Methods: Restriction alleviation (RA) is expressed as
the efficiency of plating unmodified lambda virC.0 on UV-irradiated
bacteria relative to the phage titer on E. coli C00r(k)(-)m(k)(-),
lacking the EcoKI restriction-modification system.
Results and Conclusions: Alleviation of EcoKI restriction following
induction of the SOS response and its dependence on RecBDC enzyme after
UV irradiation (UV induced RA) was further characterized. We examined
the efficiency of plating unmodified lambda (RA) in bacteria with
modified activities of RecBCD enzyme, i.e. in a recD mutant, in
bacteria producing the truncated RecB(1.929) polypeptide instead of
wild-type RecB and in a Gam(+)- producing strain in which the RecBCD
enzyme interacts with the Gam protein of phage lambda It follows from
the results presented here that the helicase activity of the RecBCD
enzyme is involved in UV-induced RA.

<>

<1>Cajthamlova, K., Sisakova, E., Weiser, J., Weiserova, M.
<2>Phosphorylation of type IA restriction-modification complex enzyme EcoKI on the HsdR subunit.
<3>FEMS Microbiol. Lett.
<4>270
<5>171-177
<6>2007
<7>Phosphorylation of Type I restriction-modification (R-M) enzymes EcoKI, EcoAI, and EcoR124I -
representatives of IA, IB, and IC families,
respectively - was analysed in vivo by immunoblotting of endogenous
phosphoproteins isolated from Escherichia coli strains harbouring the
corresponding hsd genes, and in vitro by a phosphorylation assay using
protein kinase present in subcellular fractions of E. coli. From all
three R-M enzymes, the HsdR subunit of EcoKI system was the only
subunit that was phosphorylated. Further, evidence is presented that
HsdR is phosphorylated in vivo only when coproduced with HsdM and HsdS
subunits - as part of assembled EcoKI restriction endonuclease, while
the individually produced HsdR subunit is not phosphorylated. In vitro
phosphorylation of the HsdR subunit of purified EcoKI endonuclease
occurs on Thr, and is strictly dependent on the addition of a catalytic
amount of cytoplasmic fraction isolated from E. coli. So far this is
the first case of phosphorylation of a Type I R-M enzyme reported.

<>

<1>Cal, S., Connolly, B.A.
<2>The EcoRV modification methylase causes considerable bending of DNA upon binding to its recognition sequence GATATC.
<3>J. Biol. Chem.
<4>271
<5>1008-1015
<6>1996
<7>The EcoRV methyltransferase modifies DNA by the introduction of a methyl group at the 6-NH2
position of the first deoxyadenosine in GATATC sequences.  The enzyme forms a stable and
specific complex with GATATC sequences in the presence of a nonreactive analogue, such as
sinefungin, of its natural cofactor S-adenosyl-L-methionine.  Using circular permutation band
mobility shift analysis (in which the distance between the GATATC sequence and the end of the
DNA is varied) of protein-DNA-cofactor complexes we have shown the methylase induces a bend of
just over 60o in the bound DNA.  This was confirmed by phasing analysis, in which the spacing
between the GATATC site and a poly(dA) tract is varied through a helical turn, which showed
that the orientation of the induced curve was toward the major groove.  There was no
significant difference in the bend angle measured using unmethylated GATATC sequences and
hemimethylated sequences which contain G6-MeATATC in one strand only.  These are the natural
substrates for the enzyme.  The EcoRV endonuclease, a very well characterized protein, served
as a positive control.  DNA bending by this protein has been previously determined both by
crystallographic and solution methods.  The two proteins bend DNA toward the major groove but
the bend angle produced by the methylase, slightly greater than 60o, is a little larger than
that observed with the endonuclease, which is approximately 44o.

<>

<1>Cal, S., Connolly, B.A.
<2>DNA distortion and base flipping by the EcoRV DNA methyltransferase.
<3>J. Biol. Chem.
<4>272
<5>490-496
<6>1997
<7>The EcoRV DNA methyltransferase introduces a CH3 group at the 6-amino position of the first dA
in the duplex sequence d(GATATC).  It has previously been reported that the methylase contacts
the four phosphates (pNpNpGpA) at, and preceding, the 5'-end of the recognition sequence as
well as the single dG in this sequence.  To study the possible role of the dA and T bases
within the ATAT sequence, interference studies have been carried out using
diethylpyrocarbonate and osmium tetroxide.  The methylase bound very strongly to
hemimethylated oligonucleotides modified at the second AT, of the ATAT sequence, in the
unmethylated strand of the duplex.  This probably arises because these modifications
facilitate DNA distortion that follows the binding of the nucleic acid to the protein.
Oligonucleotides containing modified bases at both the target dA base and its complementary T
were used to determine whether this dA methylase flips out its target base in a similar manner
to that observed for dC DNA methylases.  In binary EcoRV methylase-DNA complexes, analogues
that weakened the base pair caused an increase in affinity between the protein and the nucleic
acid.  In contrast, in ternary EcoRV methylase-DNA-sinefungin (an analogue of the natural
co-factor, S-adenosyl-L-methionine (AdoMet)) complexes, only small differences in affinity
were observed between the normal dA-T base pair and the analogues.  These results are almost
identical to those seen with DNA dC methylases and support a base-flipping mechanism for DNA
dA methylases.

<>

<1>Calarco, J.P., Martienssen, R.A.
<2>Imprinting: DNA Methyltransferases Illuminate Reprogramming.
<3>Curr. Biol.
<4>22
<5>R929-R931
<6>2012
<7>
<>

<1>Calcutt, M.J., Foecking, M.F.
<2>Genome Sequence of Mycoplasma putrefaciens Type Strain KS1.
<3>J. Bacteriol.
<4>193
<5>6094
<6>2011
<7>Mycoplasma putrefaciens is a causative agent of contagious agalactia in goats. Reported herein
is the complete genome sequence of the M.
putrefaciens type strain KS1.

<>

<1>Calcutt, M.J., Foecking, M.F.
<2>Complete Genome Sequence of Mycoplasma bovoculi Strain M165/69T (ATCC 29104).
<3>Genome Announcements
<4>2
<5>e00115-14
<6>2014
<7>Bovine ocular infections compromise animal health and result in significant economic losses.
Mycoplasma bovoculi is an etiological agent of conjunctivitis.
Presented here is the 760,240-bp complete genome sequence of the M. bovoculi type
strain M165/69(T). An analysis of the deduced proteome provides insights into the
adherence and antigenic variation mechanisms of the strain.

<>

<1>Calcutt, M.J., Foecking, M.F., Fox, L.K.
<2>Complete Genome Sequence of the Bovine Mastitis Pathogen Mycoplasma californicum  Strain ST-6T (ATCC 33461T).
<3>Genome Announcements
<4>2
<5>e00648-14
<6>2014
<7>Mycoplasma californicum is one of several mycoplasmal species associated with bovine mastitis.
The complete genome sequence of 793,841 bp has been determined
and annotated for the M. californicum ST-6 type strain, providing a resource for
the identification of surface antigens and putative pathoadaptive features.

<>

<1>Calcutt, M.J., Foecking, M.F., Fry, P.R., Hsieh, H.Y., Perry, J., Stewart, G.C., Scholl, D.T., Messier, S., Middleton, J.R.
<2>Draft Genome Sequence of Bovine Mastitis Isolate Staphylococcus agnetis CBMRN 20813338.
<3>Genome Announcements
<4>2
<5>e00883-14
<6>2014
<7>Presented here is a draft genome sequence for Staphylococcus agnetis CBMRN 20813338, isolated
from a lactating dairy cow with subclinical mastitis. The
genome is approximately 2,416 kb and has 35.79% G+C content. Analysis of the
deduced open reading frame (ORF) set identified candidate virulence attributes in
addition to potential molecular targets for species identification.

<>

<1>Calcutt, M.J., Foecking, M.F., Hsieh, H.Y., Perry, J., Stewart, G.C., Middleton, J.R.
<2>Genome Sequence Analysis of Staphylococcus equorum Bovine Mastitis Isolate UMC-CNS-924.
<3>Genome Announcements
<4>1
<5>e00840-13
<6>2013
<7>Intramammary infections in dairy cattle are frequently caused by staphylococci, resulting in
mastitis and associated economic losses. A draft genome sequence was
determined for Staphylococcus equorum UMC-CNS-924, isolated from the milk of a
Holstein cow, to better understand the genetic basis of its pathogenesis and
adaptation to the bovine mammary gland.

<>

<1>Calcutt, M.J., Foecking, M.F., Hsieh, H.Y., Perry, J., Stewart, G.C., Middleton, J.R.
<2>Draft Genome Sequence of Staphylococcus simulans UMC-CNS-990, Isolated from a Case of Chronic Bovine Mastitis.
<3>Genome Announcements
<4>1
<5>e01037-13
<6>2013
<7>Coagulase-negative staphylococci are frequently isolated from cases of subclinical bovine
mastitis. Reported here is a draft genome sequence of
Staphylococcus simulans UMC-CNS-990, an isolate recovered from a chronic
intramammary infection of a Holstein cow. Unexpectedly, a cluster of genes
encoding gas vesicle proteins was found within the 2,755-kb genome.

<>

<1>Calcutt, M.J., Foecking, M.F., Martin, N.T., Mhlanga-Mutangadura, T., Reilly, T.J.
<2>Draft Genome Sequence of Moraxella bovoculi Strain 237T (ATCC BAA-1259T) Isolated from a Calf with Infectious Bovine Keratoconjunctivitis.
<3>Genome Announcements
<4>2
<5>e00612-14
<6>2014
<7>Moraxella bovoculi is a recently identified species, recovered from the bovine eye, which is
under investigation as an etiological agent of infectious bovine
keratoconjunctivitis. A draft genome sequence of the Moraxella bovoculi type
strain 237(T) has been determined to identify features that may be important
during host colonization.

<>

<1>Calcutt, M.J., Foecking, M.F., Mhlanga-Mutangadura, T., Reilly, T.J.
<2>Genome Sequence of Actinobacillus suis Type Strain ATCC 33415T.
<3>Genome Announcements
<4>2
<5>e00926-14
<6>2014
<7>The assembled and annotated genome of Actinobacillus suis ATCC 33415(T) is reported here. The
2,501,598-bp genome encodes 2,246 open reading frames (ORFs)
with strain variable incursion of an integrative conjugative element into a tRNA
locus. Comparative analysis of the deduced gene set should inform our
understanding of pathogenesis, genomic plasticity, and serotype variation.

<>

<1>Calcutt, M.J., Foecking, M.F., Nagaraja, T.G., Stewart, G.C.
<2>Draft Genome Sequence of Fusobacterium necrophorum subsp. funduliforme Bovine Liver Abscess Isolate B35.
<3>Genome Announcements
<4>2
<5>e00412-14
<6>2014
<7>Fusobacterium necrophorum is a Gram-negative anaerobic bacterium that causes foot rot and
liver abscesses in cattle. F. necrophorum subsp. necrophorum and the less
virulent organism F. necrophorum subsp. funduliforme are recognized. We present
here a draft genome sequence of the bovine liver abscess isolate F. necrophorum
subsp. funduliforme strain B35, which affords a genomic perspective of virulence
and bovine adaptation.

<>

<1>Calcutt, M.J., Foecking, M.F., Rosales, R.S., Ellis, R.J., Nicholas, R.A.
<2>Genome Sequence of Mycoplasma hyorhinis Strain GDL-1.
<3>J. Bacteriol.
<4>194
<5>1848
<6>2012
<7>Mycoplasma hyorhinis impacts swine health and production in many countries, either as a
primary pathogen or as a component of a polymicrobial infection.
Isolates of this species are also common contaminants of tissue culture lines.
The genome sequence of the cell culture isolate M. hyorhinis GDL-1 is presented
herein.

<>

<1>Calisto, R.M., Pich, O.Q., Pinol, J., Fita, I., Querol, E., Carpena, X.
<2>Crystal structure of a putative type I restriction-modification S subunit from Mycoplasma genitalium.
<3>J. Mol. Biol.
<4>351
<5>749-762
<6>2005
<7>The crystal structure of the eubacteria Mycoplasma genitalium ORF MG438 peptide, determined by
multiple anomalous dispersion and refined at
poly 2.3 angstrom resolution, reveals the organization of S subunits
from the Type I restriction and modification system. The structure
consists of two globular domains, with about 150 residues each,
separated by a pair of 40 residue long antiparallel alpha-helices. The
globular domains correspond to the variable target recognition domains
(TRDs), as previously defined for S subunits on sequence analysis,
while the two helices correspond to the central (CRI) and C-terminal
(CR2) conserved regions, respectively. The structure of the MG438
subunit presents an overall cyclic topology with an intramolecular
2-fold axis that superimposes the N and the C-half parts, each half
containing a globular domain and a conserved helix. TRDs are found to
be structurally related with the small domain of the Type II N6-adenine
DNA MTase TaqI. These relationships together with the structural
peculiarities of MG438, in particular the presence of the
intramolecular quasi-symmetry, allow the proposal of a model for S
subunits recognition of their DNA targets in agreement with previous
experimental results. In the crystal, two subunits of MG438 related by
a crystallographic 2-fold axis present a large contact area mainly
involving the symmetric interactions of a cluster of exposed
hydrophobic residues. Comparison with the recently reported structure
of an S subunit from the archaea Methanococcus jannaschii highlights
the structural features preserved despite a sequence identity below
20%, but also reveals important differences in the globular domains and
in their disposition with respect to the conserved regions.

<>

<1>Callahan, S.J., Morgan, R.D., Jain, R., Townson, S.A., Wilson, G.G., Roberts, R.J., Aggarwal, A.K.
<2>Crystallization and preliminary crystallographic analysis of the type IIL restriction enzyme MmeI in complex with DNA.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>67
<5>1262-1265
<6>2011
<7>Type IIL restriction enzymes have rejuvenated the search for user-specified DNA binding and
cutting. By aligning and contrasting the
highly comparable amino-acid sequences yet diverse recognition
specificities across the family of enzymes, amino acids involved in DNA
binding have been identified and mutated to produce alternative binding
specificities. To date, the specificity of MmeI (a type IIL restriction
enzyme) has successfully been altered at positions 3, 4 and 6 of the
asymmetric TCCRAC (where R is a purine) DNA-recognition sequence. To
further understand the structural basis of MmeI DNA-binding
specificity, the enzyme has been crystallized in complex with its DNA
substrate. The crystal belonged to space group P1, with unit-cell
parameters a = 61.73, b = 94.96, c = 161.24 angstrom, alpha = 72.79,
beta = 89.12, gamma = 71.68 degrees, and diffracted to 2.6 angstrom
resolution when exposed to synchrotron radiation. The structure
promises to reveal the basis of MmeI DNA-binding specificity and will
complement efforts to create enzymes with novel specificities.

<>

<1>Callanan, M., Kaleta, P., O'Callaghan, J., O'Sullivan, O., Jordan, K., McAuliffe, O., Sangrador-Vegas, A., Slattery, L., Fitzgerald, G.F., Beresford, T., Ross, R.P.
<2>Genome sequence of Lactobacillus helveticus: an organism distinguished by selective gene loss and IS element expansion.
<3>J. Bacteriol.
<4>190
<5>727-735
<6>2008
<7>Mobile genetic elements are major contributing factors to the generation of genetic diversity
in prokaryotic organisms. For example insertion
sequence (IS) elements have been shown to specifically contribute to niche
adaptation by promoting a variety of genetic rearrangements. The complete
genome sequence of the cheese culture Lactobacillus helveticus DPC 4571
was determined and revealed significant conservation when compared to
three non-dairy gut lactobacilli. Despite originating from significantly
different environments, 65-75% of genes were conserved between the
commensal and dairy lactobacilli which allowed key niche-specific gene
sets to be described. However, the primary distinguishing feature was two
hundred and thirteen IS elements in the DPC 4571 genome, ten times more
than the other lactobacilli. Moreover, genome alignments revealed an
unprecedented level of genome stability between these four Lactobacillus
species considering the number of IS elements in the L. helveticus genome.
Comparative analysis also indicated that the IS elements were not the
primary agents of niche adaptation for the L. helveticus genome. A clear
bias towards the loss of genes reported to be important for gut
colonization was observed for the cheese culture but there was no clear
evidence of IS-associated gene deletion and decay for the majority of
genes lost. Furthermore, an extraordinary level of sequence diversity
exists between copies of certain IS elements in the DPC 4571 genome
indicating they may represent an ancient component of the L. helveticus
genome. These data suggests a special unobtrusive relationship between the
DPC 4571 genome and its mobile DNA complement.

<>

<1>Calleja, F., Dekker, B.M.M., Coursin, T., de Waard, A.
<2>A new sequence specific endonuclease EspI, of cyanobacterial origin.
<3>FEBS Lett.
<4>178
<5>69-72
<6>1984
<7>The isolation of a new sequence-specific endonuclease from a unicellular
cyanobacterium is described.  This enzyme specifically cleaves the nucleotide
sequence GC^TNAGC.

<>

<1>Calleja, F., Tandeau de Marsac, N., Coursin, T., van Ormondt, H., de Waard, A.
<2>A new endonuclease recognizing the deoxynucleotide sequence C^CNNGG from the cyanobacterium Synechocystis 6701.
<3>Nucleic Acids Res.
<4>13
<5>6745-6751
<6>1985
<7>A new sequence-specific endonuclease from the cyanobacterium Synechocystis
species PCC 6701 has been purified and characterized.  This enzyme, SecI, is
unique in recognizing the nucleotide sequence: 5' -C^C N N G G-3'3' -G G N N
C^C-5'	and cleaves it at the position indicated by the ^ symbol.  Two other
restriction endonucleases, SecII and SecIII, found in this organism are
isoschizomers of MspI and MstII, respectively.

<>

<1>Callow, P., Sukhodub, A., Taylor, J.E., Kneale, G.G.
<2>Shape and Subunit Organisation of the DNA Methyltransferase M.AhdI by Small-angle Neutron Scattering.
<3>J. Mol. Biol.
<4>369
<5>177-185
<6>2007
<7>Type I restriction-modification (R-M) systems encode multisubunit/multidomain enzymes. Two
genes (M and S) are required to form
the methyltransferase (MTase) that methylates a specific base within the
recognition sequence and protects DNA from cleavage by the endonuclease.
The DNA methyltransferase M.AhdI is a 170 kDa tetramer with the
stoichiometry M(2)S(2) and has properties typical of a type I MTase. The
M.AhdI enzyme has been prepared with deuterated S subunits, to allow
contrast variation using small-angle neutron scattering (SANS) methods.
The SANS data were collected in a number of (1)H:(2)H solvent contrasts to
allow matching of one or other of the subunits in the multisubunit enzyme.
The radius of gyration (R(g)) and maximum dimensions (D(max)) of the M
subunits in situ in the multisubunit enzyme (50 A and 190 A, respectively)
are close of those of the entire MTase (51 A and 190 A). In contrast, the
S subunits in situ have experimentally determined values of R(g)=35 A and
D(max)=110 A, indicating their more central location in the enzyme. Ab
initio reconstruction methods yield a low-resolution structural model of
the shape and subunit organization of M.AhdI, in which the Z-shaped
structure of the S subunit dimer can be discerned. In contrast, the M
subunits form a much more elongated and extended structure. The core of
the MTase comprises the two S subunits and the globular regions of the two
M subunits, with the extended portion of the M subunits most probably
forming highly mobile regions at the outer extremities, which collapse
around the DNA when the MTase binds.

<>

<1>Callow, P., Timmins, P., Kneale, G.
<2>Preliminary neutron scattering studies of the Type I restriction-modification enzyme M.Ahdl.
<3>Phys. Rev., B Condens. Matter
<4>385-386
<5>853-855
<6>2006
<7>Type I restriction-modification (R-M) systems encode multisubunit/multidomain enzymes. Two
genes (M and S) are required to
form the 160kDa methyltransferase that methylates a specific base
within the recognition sequence and protects DNA from cleavage by the
endonuclease. Small angle neutron scattering (SANS) revealed an
unusually large structural change in the EcoR124l methyltransferase
following DNA binding; this involves a major repositioning of the
subunits of the enzyme, resulting in a 60 angstrom reduction in the
dimensions of the enzyme on forming a complex with DNA. The related
methyltransferase M.Ahdl, with stoichiometry M2S2 has been prepared in
two protonated/deuterated states (S and M subunits protonated, S
deuterated and M protonated) for which SANS data have been collected in
a number of H:D solvent contrasts. The contrast match point of the
selectively deuterated enzyme confirms the successful reconstitution of
the enzyme with deuterated S subunits. Ab initio shape determination
using this contrast matched data is in progress to determine the
subunit organization of M.Ahdl and the large change in structure that
occurs on DNA binding.

<>

<1>Calmann, M.A., Marinus, M.G.
<2>Regulated expression of the Escherichia coli dam gene.
<3>J. Bacteriol.
<4>185
<5>5012-5014
<6>2003
<7>Regulated expression of the Escherichia coli dam gene has been achieved with the araBAD
promoter lacking a ribosome binding site. Cultures of dam
mutants containing plasmid pMQ430 show no detectable methylation in the
absence of arabinose and complete methylation in its presence. Dam
methyltransferase is a substrate for the Lon protease.

<>

<1>Calvin-Koons, M.D., Blumenthal, R.M.
<2>Characterization of pPvu1, the autonomous plasmid from Proteus vulgaris that carries the genes of the PvuII restriction-modification system.
<3>Gene
<4>157
<5>73-79
<6>1995
<7>Plasmid pPvu1 from Proteus vulgaris carries the genes of the PvuII restriction-modification
system.  This report focuses on physical and functional features of the 4.84-kb plasmid, which
shows a composite genetic architecture.  Plasmid pPvu1 has a replication origin and an
incompatibility locus that each function in Escherichia coli, and an apparent cer
recombination site.  The replication origin includes a possible RNAI gene, and the
incompatibility locus closely resembles a rom gene.  These loci show substantial sequence
similarity to corresponding loci from the E. coli plasmids P15A, ColEI and pSC101, and closely
flank the PvuII genes.  The close association between a recombinational locus and the PvuII
genes has implications for their mobility.

<>

<1>Cameron, B., Crouzet, J.
<2>Production of methylated plasmid DNA for gene therapy - plasmid-mediated DNA-methyltransferase expression in bacterium.
<3>French Patent Office
<4>FR 2750704 A
<5>
<6>1998
<7>
<>

<1>Camp, R., Wilson, G.
<2>Method for producing the BanI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0321268 A
<5>
<6>1988
<7>The present invention is directed to a method for cloning and producing the BanI restriction
endonuclease by 1) introducing the restriction endonuclease gene from B. aneurinolyticus IAM
1077 into a host whereby the restriction gene is expressed; 2) fermenting the host which
contains the vector encoding and expressing the BanI restriction endonuclease, and 3)
purifying the BanI restriction endonuclease from the fermented host which contains the vector
encoding and expressing the BanI restriction endonuclease activity.

<>

<1>Campbell, B.J., Smith, J.L., Hanson, T.E., Klotz, M.G., Stein, L.Y., Lee, C.K., Wu, D., Robinson, J.M., Khouri, H.M., Eisen, J.A., Cary, S.C.
<2>Adaptations to submarine hydrothermal environments exemplified by the genome of Nautilia profundicola.
<3>PLoS Genet.
<4>5
<5>e1000362
<6>2009
<7>Submarine hydrothermal vents are model systems for the Archaean Earth environment, and some
sites maintain conditions that may have favored the
formation and evolution of cellular life. Vents are typified by rapid
fluctuations in temperature and redox potential that impose a strong
selective pressure on resident microbial communities. Nautilia
profundicola strain Am-H is a moderately thermophilic, deeply-branching
Epsilonproteobacterium found free-living at hydrothermal vents and is a
member of the microbial mass on the dorsal surface of vent polychaete,
Alvinella pompejana. Analysis of the 1.7-Mbp genome of N. profundicola
uncovered adaptations to the vent environment--some unique and some shared
with other Epsilonproteobacterial genomes. The major findings included:
(1) a diverse suite of hydrogenases coupled to a relatively simple
electron transport chain, (2) numerous stress response systems, (3) a
novel predicted nitrate assimilation pathway with hydroxylamine as a key
intermediate, and (4) a gene (rgy) encoding the hallmark protein for
hyperthermophilic growth, reverse gyrase. Additional experiments indicated
that expression of rgy in strain Am-H was induced over 100-fold with a 20
degrees C increase above the optimal growth temperature of this bacterium
and that closely related rgy genes are present and expressed in bacterial
communities residing in geographically distinct thermophilic environments.
N. profundicola, therefore, is a model Epsilonproteobacterium that
contains all the genes necessary for life in the extreme conditions widely
believed to reflect those in the Archaean biosphere--anaerobic, sulfur,
H2- and CO2-rich, with fluctuating redox potentials and temperatures. In
addition, reverse gyrase appears to be an important and common adaptation
for mesophiles and moderate thermophiles that inhabit ecological niches
characterized by rapid and frequent temperature fluctuations and, as such,
can no longer be considered a unique feature of hyperthermophiles.

<>

<1>Campellone, K.G., Roe, A.J., Lobner-Olesen, A., Murphy, K.C., Magoun, L., Brady, M.J., Donohue-Rolfe, A., Tzipori, S., Gally, D.L., Leong, J.M., Marinus, M.G.
<2>Increased adherence and actin pedestal formation by dam-deficient enterohaemorrhagic Escherichia coli O157 : H7.
<3>Mol. Microbiol.
<4>63
<5>1468-1481
<6>2007
<7>Enterohaemorrhagic Escherichia coli (EHEC) are highly infectious pathogens capable of causing
severe diarrhoeal illnesses. As a critical
step during their colonization, EHEC adhere intimately to intestinal
epithelial cells and generate F-actin 'pedestal' structures that
elevate them above surrounding cell surfaces. Intimate adhesion and
pedestal formation result from delivery of the EHEC type III secretion
system (TTSS) effector proteins Tir and EspF(U) into the host cell and
expression of the bacterial outer membrane adhesin, intimin. To
investigate a role for DNA methylation during the regulation of
adhesion and pedestal formation in EHEC, we deleted the dam (DNA
adenine methyltransferase) gene from EHEC O157:H7 and demonstrate that
this mutation results in increased interactions with cultured host
cells. EHEC Delta dam exhibits dramatically elevated levels of
adherence and pedestal formation when compared with wild-type EHEC, and
expresses significantly higher protein levels of intimin, Tir and
EspF(U). Analyses of GFP fusions, Northern blotting, reverse
transcription polymerase chain reaction, and microarray experiments
indicate that the abundance of Tir in the dam mutant is not due to
increased transcription levels, raising the possibility that Dam
methylation can indirectly control protein expression by a
post-transcriptional mechanism. In contrast to other dam-deficient
pathogens, EHEC Delta dam is capable of robust intestinal colonization
of experimentally infected animals.

<>

<1>Campione-Piccardo, J., Moro, R.
<2>Modification of DNA ends and detection of restriction enzyme recognition sequences in their ligated junctions.
<3>Comput. Appl. Biosci.
<4>4
<5>215-216
<6>1988
<7>A program has been developed for the modelling of modifications in DNA ends,
for the construction of ligated junctions, and for the analysis in these
junctions of new restriction enzyme recognition sequences.  This program allows
the analysis of restriction enzyme specificities in ligated junctions of
cohesive or blunt DNA ends.  Cohesive ends are considered in their natural
configuration or after modification by possible blunt-ending procedures.  The
program also allows the modelling of partial filling-in for 5'-single-stranded
ends.   This program has proven useful for the design of sequences with new
restriction sites or to predict or confirm the sequences of junctions created
by the ligation of modified ends.

<>

<1>Campos-Guillen, J., Caballero, P.J., Cruz, M.J.A., Molina, V.C., Salas, R.L.M., Limpens, G.C., Garcia, S.I., Hernandez, R.M.R., Soto, A.G., Cruz, H.A., Saldana, G.C., Romero, G.S., Pastrana, M.X., Alvarez, H.E., Gosar, M., Dizdarevic, T.
<2>Draft Genome Sequence of the Mercury-Resistant Bacterium Acinetobacter idrijaensis Strain MII, Isolated from a Mine-Impacted Area, Idrija, Slovenia.
<3>Genome Announcements
<4>2
<5>e01177-14
<6>2014
<7>We report here the first draft assembly for the genome of Acinetobacter idrijaensis strain
MII, isolated from the Idrija mercury mine area (Slovenia).
This strain shows a strikingly high tolerance to mercury, and the genome sequence
shows genes involved in the mechanisms for heavy metal tolerance pathways and
multidrug efflux pumps.

<>

<1>Campoy, S., Aranda, J., Alvarez, G., Barbe, J., Llagostera, M.
<2>Isolation and Sequencing of a Temperate Transducing Phage for Pasteurella multocida.
<3>Appl. Environ. Microbiol.
<4>72
<5>3154-3160
<6>2006
<7>A temperate bacteriophage (F108) has been isolated through mitomycin C
induction of a Pasteurella multocida serogroup A strain. F108 has a
typical morphology of the family Myoviridae, presenting a hexagonal head
and a long contractile tail. F108 is able to infect all P. multocida
serogroup A strains tested but not those belonging to other serotypes.
Bacteriophage F108, the first P. multocida phage sequenced so far,
presents a 30,505-bp double-stranded DNA genome with cohesive ends
(CTTCCTCCCC cos site). The F108 genome shows the highest homology with
those of Haemophilus influenzae HP1 and HP2 phages. Furthermore, an F108
prophage attachment site in the P. multocida chromosome has been
established to be inside a gene encoding tRNA(Leu). By using several
chromosomal markers that are spread along the P. multocida chromosome, it has been
demonstrated that F108 is able to perform generalized transduction. This fact, together with
the absence of pathogenic genes in the F108 genome, makes this bacteriophage a valuable tool
for P.multocida genetic manipulation.

<>

<1>Campoy, S., Mazon, G., de Henestrosa, A.R.F., Llagostera, M., Monteiro, P.B., Barbe, J.
<2>A new regulatory DNA motif of the gamma subclass proteobacteria: identification of the LexA protein binding site of the plant pathogen  Xylella fastidiiosa.
<3>Microbiology
<4>148
<5>3583-3597
<6>2002
<7>Escherichia coli LexA protein is the repressor of a gene network whose members are directly
involved in the repair of damaged DNA and in the
survival of bacterial cells until DNA lesions have been eliminated. The
lexA gene is widely present in bacteria, although the sequences of only
three LexA-binding sites are known: Gram-positive, alpha Proteobacteria
and some members of gamma Proteobacteria represented by E. coli. Taking
advantage of the fact that the genome sequence of the plant-pathogenic
bacterium Xylella fastidiosa has been determined, its lexA gene has
been cloned and overexpressed in E. coli to purify its product. After
demonstration that X. fastidiosa lexA and recA genes are
co-transcribed, gel mobility shift assays and directed mutagenesis
experiments using the promoter of the lexA-recA transcriptional unit
demonstrated that the X. fastidiosa LexA protein specifically binds the
imperfect palindrome TTAGN(6)TACTA. This is the first LexA binding
sequence identified in the gamma Proteobacteria differing from the E.
coli-like LexA box. Although a computational search has revealed the
presence of TTAGN(6)TACTA-like motifs upstream of X. fastidiosa genes
other than lxA, X. fastidiosa LexA only binds the promoter of one of
them, XF2313, encoding a putative DNA-modification methylase. Moreover,
X. fastidiosa LexA protein does not bind any of the other genes whose
homologues are regulated by the LexA repressor in E. coli (uvrA, uvrB,
ssb, ruvAB, ftsK, dinG, recN and ybfE). RTPCR quantitative analysis has
also demonstrated that lexA-recA and XF2313 genes, as well as the X.
fastidiosa genes which are homologues to those of E. coli belonging to
the LexA regulon, with the exception of ssb, are DNA damage-inducible
in X. fastidiosa.

<>

<1>Camus, J.C., Pryor, M.J., Medigue, C., Cole, S.T.
<2>Re-annotation of the genome sequence of Mycobacterium tuberculosis H37Rv.
<3>Microbiology
<4>148
<5>2967-2973
<6>2002
<7>Original genome annotations need to be regularly updated if the information they contain is to
remain accurate and relevant. Here the
complete re-annotation of the genome sequence of Mycobacterium
tuberculosis strain H37Rv is presented almost 4 years after the first
submission. Eighty-two new protein-coding sequences (CDS) have been
included and 22 of these have a predicted function. The majority were
identified by manual or automated re-analysis of the genome and most of
them were shorter than the 100 codon cut-off used in the initial genome
analysis. The functional classification of 643 CDS has been changed based
principally on recent sequence comparisons and new experimental data from
the literature. More than 300 gene names and over 1000 targeted citations
have been added and the lengths of 60 genes have been modified. Presently,
it is possible to assign a function to 2058 proteins (52% of the 3995
proteins predicted) and only 376 putative proteins share no homology with
known proteins and thus could be unique to M. tuberculosis.

<>

<1>Canevari, C.A.B., Nishibe, C., Moura, A., de Alencar, A.P., de Azevedo, I.M., Hodon, M.A., Mota, P.M., Sales, E.B., Fonseca, J.A.A., Almeida, N.F., Araujo, F.R.
<2>Draft Genome Sequence of Mycobacterium bovis Strain AN5, Used for Production of Purified Protein Derivative.
<3>Genome Announcements
<4>2
<5>e00277-14
<6>2014
<7>Mycobacterium bovis strain AN5 has been used to produce purified protein derivative (PPD) for
the intradermal test for bovine tuberculosis since it was introduced in 1948. This work
reports the draft genome sequence of M. bovis AN5, which is used for the production of bovine
PPD in Brazil, as well as comparisons to other strains of M. bovis and Mycobacterium
tuberculosis.

<>

<1>Canosi, U., Luder, G., Trautner, T.A., Bron, S.
<2>Restriction and modification in B. subtilis:  effects on plasmid transformation.
<3>Transformation 1980:  Proceedings of the Fifth European Meeting on Bacterial Transformation and Transfection, Cotswold Press, , Oxford
<4>0
<5>179-187
<6>1981
<7>In previous communications in this series we have demonstrated that bacterial
transformation is resistent to restriction, whereas transfection is susceptible
to restriction.  To explain this difference in the behaviour of the two kinds
of DNA we have postulated that both transforming and transfecting DNA are first
converted to single stranded DNA molecules in the course of DNA processing.
Such single stranded transforming DNA would then hybridize to modified host DNA
of complementary polarity to form donor/recipient complex.  This
donor/recipient complex represents DNA heterozygous with respect to
methylation.  It would be highly reactive to become totally modified and hence
resistant to restriction.  In transfection, DNA/DNA hybridization would be
between complementary strands of opposite polarity derived from different
transfecting phage DNA molecules.  These molecules would be double stranded,
nonmodified in the pairing regions and would hence be substrates for the
restriction enzyme.  Major support for this interpretation came from
experiments in which we had demonstrated that transfection of lysogenic cells
with homologus phage DNA was insensitive to restriction.  This result pointed
to the important role played by homology between donor and recipient DNA in the
establishment of DNA taken up by restricting cells.

<>

<1>Cantalupo, G., Bucci, C., Salvatore, P., Pagliarulo, C., Roberti, V., Lavitola, A., Bruni, C.B., Alifano, P.
<2>Evolution and function of the Neisserial dam-replacing gene.
<3>FEBS Lett.
<4>495
<5>178-183
<6>2001
<7>Phase variation through slippage-like mechanisms involving homopolymeric tracts depends in
part on the absence of Dam-methylase in
several pathogenic isolates of Neisseria meningitidis, In Dam-defective
strains drg (dam-replacing gene), flanked by pseudo-transposable small
repeated elements (SREs), replaced dam. We demonstrate that drg encodes
a restriction endonuclease (NmeBII) that cleaves 5'-GmeATC-3'. drg is
also present in 50% of Neisseria lactamica strains, but in most of them
it is inactive because of the absence of an SRE-providing, promoter.
This is associated with the presence of GATmeC suggesting an
alternative restriction-modification system (RM) specific for
5'-GATC-3', similar to Sau3AI-RM of Staphylococcus aureus 3A.
Lactococcus lactis KR2 and Listeria monocytogenes.

<>

<1>Cao, B., Chen, C., DeMott, M.S., Cheng, Q., Clark, T.A., Xiong, X., Zheng, X., Butty, V., Levine, S.S., Yuan, G., Boitano, M., Khai, L., Song, Yi., Zhou, X., Deng, Z., Turner, S.W., Korlach, J., You, D., Wang, L., Chen, S., Dedon, P.C.
<2>Genomic mapping of phosphorothioates reveals partial modification of short consensus sequences.
<3>Nat. Commun.
<4>5
<5>3951
<6>2014
<7>Bacterial phosphorothioate (PT) DNA modifications are incorporated by Dnd proteins A-E and
often function with DndF-H as a restriction-modification (R-M) system, as in Escherichia coli
B7A. However, bacteria such as Vibrio cyclitrophicus FF75 lack dndF-H, which points to other
PT functions. Here we report two novel, orthogonal technologies to map PTs across the genomes
of B7A and FF75 with > 90% agreement: single molecule, real-time sequencing and deep
sequencing of iodine-induced cleavage at PT (ICDS). In B7A, we detect PT on both strands of
G(ps)AAC/G(ps)TTC motifs, but with only 12% of 40,701 possible sites modified. In contrast, PT
in FF75 occurs as a single-strand modification at C(ps)CA, again with only 14% of 160,541
sites modified. Single-molecule analysis indicates that modification could be partial at any
particular genomic site even with active restriction by DndF-H, with direct interaction of
modification proteins with GAAC/GTTC sites demonstrated with oligonucleotides. These results
point to highly unusual target selection by PT-modification proteins and rule out known R-M
mechanisms.

<>

<1>Cao, B., Ma, T., Ren, Y., Ren, Y., Li, G., Li, P., Guo, X., Ding, P., Feng, L.
<2>Complete Genome Sequence of Pusillimonas sp. T7-7, a cold-tolerate diesel oil-degrading bacterium isolated from the Bohai Sea in China.
<3>J. Bacteriol.
<4>193
<5>4021-4022
<6>2011
<7>Pusillimonas sp. T7-7 is a diesel oil-degrading cold-tolerate bacterium isolated from the
benthal mud of petroleum-contaminated site in Bohai Sea,
China. We present here the complete genome sequence of T7-7. Genome
analysis revealed many features of typical marine bacteria including the
absence of intact sugar metabolic pathways, the presence of glyoxylate and
gluconeogenesis pathways, and the abilities for nitrate assimilation and
denitrification, as well as sulfate reduction and sulfite oxidation.
Presence of novel genes for the degradation of diesel oils was suggested.

<>

<1>Cao, Bo., Cheng, Q., Gu, C., Yao, F., DeMott, M.S., Zheng, X., Deng, Z., Dedon, P.C., You, D.
<2>Pathological phenotypes and in vivo DNA cleavage by unrestrained activity of a phosphorothioate- based restriction system in Salmonella.
<3>Mol. Microbiol.
<4>93
<5>776-785
<6>2014
<7>Prokaryotes protect their genomes from foreign DNA with a diversity of defence mechanisms,
including a widespread restriction-modification (R-M) system involving phosphorothioate (PT)
modification of the DNA backbone. Unlike classical R-M systems, highly partial PT modification
of consensus motifs in bacterial genomes suggests an unusual mechanism of PT-dependent
restriction. In Salmonella enterica, PT modification is mediated by four genes dptB-E, while
restriction involves additional three genes dptF-H. Here, we performed a series of studies to
characterize the PT-dependent restriction, and found that it presented several features
distinct with traditional R-M systems. The presence of restriction genes in a PT-deficient
mutant was not lethal, but instead resulted in several pathological phenotypes. Subsequent
transcriptional profiling revealed the expression of > 600 genes was affected by restriction
enzymes in cells lacking PT, including induction of bacteriophage, SOS response and DNA
repair-related genes. These transcriptional responses are consistent with the observation that
restriction enzymes caused extensive DNA cleavage in the absence of PT modifications in vivo.
However, overexpression of restriction genes was lethal to the host in spite of the presence
PT modifications. These results point to an unusual mechanism of PT-dependent DNA cleavage by
restriction enzymes in the face of partial PT modification.

<>

<1>Cao, D.
<2>Cosolute effects as probes of the role of water in DNA binding and catalysis by three restriction endonucleases.
<3>Ph.D. Thesis, University of Pittsburgh
<4>
<5>283
<6>2002
<7>Site-specific protein-DNA interactions play important roles in many cellular processes and are
regulated by many factors, including the amount and type of cytoplasmic solutes.  In this work
I use restriction endonucleases EcoRI, BamHI and EcoRV endonucleases as models to probe the
role of water in formation of protein-DNA complexes and to understand how osmolytes
(cosolutes) promote protein-DNA association.  We observe that many cosolutes facilitate
specific binding of protein to DNA; however the extent of the effect depends on the
physico-chemical properties of the cosolute.  Our data show conclusively that cosolute effects
are best understood from the perspective of 'preferential interaction' rather than those of
'osmotic stress' or 'excluded volume effects.  Because we find that triethylene glycol is
nearly completely excluded from the macromolecular surfaces, we use the dependence of the
equilibrium association of protein and DNA on TEG concentration to estimate that 430, 350 and
450 molecules of H2O are released upon formation of specific EcoRI, BamHI and EcoRV complexes,
respectively.  The flanking sequence surrounding the specific site affects binding free energy
but does not affect water stoichiometry.  We show that binding to nonspecific sites involves
little water release whereas binding to miscognate sites (one incorrect base pair) entails
intermediate water release.  Thus a massive amount of water release is a thermodynamic
signature for the formation of the specific complex.  In addition, the experimental
determination of water release permits us to establish for the first time that the hydrophobic
effect can provide as much as -80 Kcal/mol, more than half of the driving force for specific
binding.  There is no dependence on cosolute concentration for cleavage of specific sites.
This implies that there is no additional release of water in achieving the specific transition
state complex; thus the specific recognition complex closely resembles the transition-state
complex.  By contrast, cosolutes enhance the cleavage of miscognate sites; we infer that the
rearrangements required of the adaptive miscognate complex to achieve the transition state are
accompanied by further release of water.

<>

<1>Cao, D., Ju, Z., Gao, C., Mei, X., Fu, D., Zhu, H., Luo, Y., Zhu, B.
<2>Genome-wide identification of cytosine-5 DNA methyltransferases and demethylases in Solanum lycopersicum.
<3>Gene
<4>550
<5>230-237
<6>2014
<7>Recent studies have reported that decreased level of DNA cytosine methylation in the global
genome was closely related to the initiation of tomato (Solanum lycopersicum) fruit ripening.
However, genome-scale analysis of cytosine-5 DNA methyltransferases (C5-MTases) and
demethylases in tomato has not been engaged. In this study, 7 C5-MTases and 3 demethylases
were identified in tomato genome, which probably contributed to DNA cytosine methylation level
in tomato. The 7 C5-MTases were categorized into 4 subgroups, and the 3 demethylases were
classified into 2 subgroups based on phylogenetic analyses. Comprehensive analysis of their
structure and genomic localization was also performed in this paper. According to online
RNA-seq data, 4 S. lycopersicum C5-MTase (SlC5-MTase) genes (SlMET, SlDRM1L1, SlDRM5, SlMET3L)
were expressed higher than others, and one DNA demethylase gene (SlDML) was significantly
changed during tomato fruit development and ripening. Furthermore, all these five gene
expressions at breaker (BK) stage changed with 1-methylcyclopropene (1-MCP) treatment,
indicating that they were regulated by ethylene directly or indirectly in tomato fruit. In
addition, subcellular localization analysis indicated that SlDRM1L1 and SlDRM5 located in the
nucleus might have responsibility for RNA-directed DNA methylation (RdDM). Collectively, this
paper provided a framework for gene discovery and functional characterization of C5-MTases and
DNA demethylases in other Solanaceae species.

<>

<1>Cao, G., Ju, W., Rump, L., Zhao, S., Zou, L., Wang, C., Strain, E., Luo, Y., Timme, R., Allard, M., Brown, E., Meng, J.
<2>Genome Sequences of Two Emerging Non-O157 Shiga Toxin-Producing Escherichia coli  Strains.
<3>Genome Announcements
<4>1
<5>e00200-13
<6>2013
<7>Shiga toxin-producing Escherichia coli (STEC) causes severe illness in humans, including
hemorrhagic colitis and hemolytic uremic syndrome. A parallel
evolutionary model was proposed in which E. coli strains of distinct phylogenies
independently integrate Shiga toxin-encoding genes and evolve into STEC. We
report the draft genomes of two emerging non-O157 STEC strains.

<>

<1>Cao, G., Liu, Y., Liu, G., Wang, J., Wang, G.
<2>Draft genome sequence of pseudomonas strain p818, isolated from glyphosate-polluted soil.
<3>Genome Announcements
<4>1
<5>e01079-13
<6>2013
<7>Pseudomonas strain P818 was isolated from glyphosate-polluted soil in China. This bacterium
presents a capacity for high glyphosate tolerance. We present the draft
genome sequence of the strain Pseudomonas P818. The genes involved in the
glyphosate tolerance were identified. This genomic information will facilitate
the study of glyphosate tolerance mechanisms.

<>

<1>Cao, G., Zhao, S., Strain, E., Luo, Y., Timme, R., Wang, C., Brown, E., Meng, J., Allard, M.
<2>Draft Genome Sequences of Eight Salmonella enterica Serotype Newport Strains from Diverse Hosts and Locations.
<3>J. Bacteriol.
<4>194
<5>5146
<6>2012
<7>Salmonellosis is a major contributor to the global public health burden. Salmonella enterica
serotype Newport has ranked among three Salmonella serotypes
most commonly associated with food-borne outbreaks in the United States. It was
thought to be polyphyletic and composed of independent lineages. Here we report
draft genomes of eight strains of S. Newport from diverse hosts and locations.

<>

<1>Cao, H., Shimura, Y., Masanobu, K., Yin, Y.
<2>Draft Genome Sequence of the Toxic Bloom-Forming Cyanobacterium Aphanizomenon flos-aquae NIES-81.
<3>Genome Announcements
<4>2
<5>e00044-14
<6>2014
<7>Aphanizomenon flos-aquae is a toxic filamentous cyanobacterium that causes water  blooms in
freshwaters across the globe. We present the draft genome sequence of
the A. flos-aquae strain NIES-81, which was determined by 454 pyrosequencing
technology. The draft genome is ~5.7 Mb, containing 5,802 predicted
protein-coding genes and 58 RNA genes, with a G+C content of 38.5%.

<>

<1>Cao, W.
<2>Binding kinetics and footprinting of TaqI endonuclease: Effects of metal cofactors on sequence-specific interactions.
<3>Biochemistry
<4>38
<5>8080-8087
<6>1999
<7>Restriction endonucleases achieve sequence-specific recognition and strand cleavage through
the interplay of base, phosphate backbone, and metal cofactor interactions. In this study, we
investigate the binding kinetics of TaqI endonuclease using the wild-type enzyme and a binding
proficient, catalysis deficient mutant TaqI-D137A both in the absence of a metal cofactor and
in the presence of Mg2+ or Ca2+. As demonstrated by gel mobility shift analyses, TaqI
endonuclease requires a metal cofactor for achieving high-affinity specific binding to its
cognate sequence, TCGA. In the absence of a metal cofactor, the enzyme binds all DNA sequences
(TaqI cognate site, star site, and nonspecific site) with essentially equal affinity, thereby
exhibiting little discrimination. The dissociation constant of the cognate sequence in the
presence of Mg2+ at 60 degrees C is 0.26 nM, a value comparable to our previously reported Km
of 0.5 nM measured under steady-state conditions. The TaqI-TCGA-Mg2+ complex is stable, with a
half-life of 21 min at 60 degrees C. The boundary of the protein-DNA interface is approximated
to be about 18 bp as determined by DNase I footprinting. Data from this study support the
notion that a metal cofactor plays a critical role for achieving sequence-specific
discrimination in a subset of nucleases, including TaqI, EcoRV, and others.

<>

<1>Cao, W., Barany, F.
<2>Identification of TaqI endonuclease active site residues by Fe2+-mediated oxidative cleavage.
<3>J. Biol. Chem.
<4>273
<5>33002-33010
<6>1998
<7>Metal cofactors (Mg2+ and Mn2+) modulate both specific DNA binding and strand cleavage in the
TaqI endonuclease.  This work attempts to establish the structural basis of TaqI-DNA-metal2+
interactions using an affinity cleavage technique.  The protein was cleaved by localized
hydroxyl radicals generated by oxidizing Fe2+ within the metal binding sites.  Cleavage
fragments were separated by SDS-polyacrylamide gel electrophoresis, and cleavage sites were
determined using micropeptide sequencing.  Eleven amino acid residues in the vicinity of
cleavage sites were selected for site-directed mutagenesis.  The negative charge at Asp137 is
essential for DNA cleavage but not required for sequence specific binding.  Mutations at
Asp142 abolish both specific binding and catalysis, except for D142E, which converts TaqI into
a completely Mn2+-dependent endonuclease.  The positive charge at Lys158 appears to be
important for both specific binding and catalysis.  Mutations at other sites affect binding
and/or catalysis to different degrees, except Trp113 and Glu135, which appear to be
nonessential for the TaqI enzyme activity.  The critical residues for TaqI function are
distinct from the PDX14-20(E/D)XK catalytic motif elucidated from other endonucleases.

<>

<1>Cao, W., Lu, J.
<2>Exploring the catalytic center of TaqI endonuclease: rescuing catalytic activity by double mutations and Mn(2+).
<3>Biochim. Biophys. Acta
<4>1546
<5>253-260
<6>2001
<7>TaqI is a metal-dependent endonuclease that recognizes T^CGA, with the arrow
indicating the cleavage site. Mutations at K158 render the enzyme inactive and mutations at
K157 significantly reduce DNA cleavage activity (W. Cao and F. Barany (1998) J. Biol. Chem.
273, 33002-33010). Aspartate, glutamate, and histidine substitutions were made at K158 in the
wild-type and K157S mutant TaqI endonuclease to understand the functional organization of the
active site. None of the mutants was active with Mg(2+), but the DNA cleavage activities were
partly rescued by Mn(2+) for K157S-K158E and K157S-K158H mutants. The rescuing effects were
observed with Mn(2+) but not with other divalent cations. K157S-K158E required higher Mn(2+)
concentrations than the wild-type enzyme for DNA cleavage activity, suggesting that a Mn(2+)
ion is weakly bound at the 158 position. The need to neutralize K157 to recover the catalytic
activity of K158E and K158H indicates that K158 and K157 may interact functionally. In analogy
with EcoRV, Ca(2+) stimulated Mn(2+)-mediated cleavage for the wild-type TaqI, suggesting the
existence of at least two metal ions at the catalytic center. A catalytic mechanism involving
two metal ions and the K157-K158 pair is proposed for TaqI endonuclease.

<>

<1>Cao, W., Lu, J., Barany, F.
<2>Nucleotide sequences and gene organization of TaqI endonuclease isoschizomers from Thermus sp. SM32 and Thermus filiformis Tok6AI.
<3>Gene
<4>197
<5>205-214
<6>1997
<7>Eight TaqI isoschizomer genes, two from Yellowstone National Park, one from Japan, two from
New Zealand, two from Portugal, and one from the Azores (1000 miles west of Portugal), were
PCR-amplified and sequenced.  Sequence alignment of isoschizomers isolated from close
geographical locations shows identical or almost identical protein sequences, while
isoschizomers from distant sites demonstrate considerable diversity, ranging from 54 to 75% in
amino acid identity.  Accordingly, these isoschizomers were arranged into four geographical
groups, i.e., USA as represented by Thermus aquaticus YT1, Japan by Thermus thermophilus HB8,
New Zealand by thermus filiformis Tok5AI, Portugal by Thermus sp. SM32.  The complete ORFs of
two new representative genes, tfiTok6AII and tsp32IR, were obtained by bubble PCR.  Unlike
M.TaqI-R.TaqI and M.TthHB81-R.TthHB8I which exhibit an unusual 13-codon overlap, the methylase
and endonuclease genes are each separated by 15 nucleotides in the TfiTok6AII and Tsp32IR
restriction-modification systems.  Phylogenetic analysis suggests that initially TfiTok6AII
diverged from a common ancestor, then Tsp32IR branched out, and finally TaqI and TthHB8I
diverged from each other during evolution.

<>

<1>Cao, W., Lu, J., Welch, S.G., Williams, R.A.D., Barany, F.
<2>Cloning and thermostability of TaqI endonuclease isoschizomers from Thermus species SM32 and Thermus filiformis Tok6A1.
<3>Biochem. J.
<4>333
<5>425-431
<6>1998
<7>Two TaqI endonuclease (hereafter referred to as TaqI) isoschizomer genes, tsp32IR from Thermus
species SM32 of Azores and tfiTok6A1I from T. filiformis Tok6A1 of New Zealand, were cloned in
Escherichia coli.  The overexpressed enzymes were partly purified and their thermostability
was determined.  In the medium-salt buffer, Tsp32IR, TfiTok6A1I and one previously cloned TaqI
isoschizomer (TthHB8I) were more thermostable than TaqI.  Tsp32IR remained partly active up to
90 C in the low-salt buffer.  Six amino acid residues that are identical in the three high
thermostability isoschizomers (Tsp32IR, TfiTok6A1I and TthHB8I) but differ in TaqI might
provide added rigidity for thermostabilization.  These include four proline residues located
in or near loop regions, and one alanine and one arginine located at helix regions in the
predicted TaqI endonuclease secondary structure.  The possible role of these residues in
thermostabilization was evaluated by mutagenizing the TaqI enzyme.  Mutants generated at these
six positions were less thermostable than wild-type TaqI.  The results suggest that the
surrounding sequence or structural context might be as important as the mutation itself.

<>

<1>Cao, W., Mayer, A.N., Barany, F.
<2>Stringent and relaxed specificities of TaqI endonuclease: interactions with metal cofactors and DNA sequences.
<3>Biochemistry
<4>34
<5>2276-2283
<6>1995
<7>We have studied the roles of metal cofactors Mg2+ and Mn2+ in modulating substrate
specificities during the enzymatic cycle of TaqI endonuclease using steady state and
single-turnover kinetics. In the presence of Mg2+, stringent discrimination of TaqI against
single base-pair changes (star sites) is manifested by the loss of tight, specific binding in
the early stage of the enzymatic cycle. In the presence of Mn2+, relaxed specificity for a
star site sequence is attributed to formation of three distinct classes of the ternary
complexes: the highly activated TaqI-cognate-Mn2+ complex; the partially activated
TaqI-star-Mn2+-complex; and the ground state, inactive TaqI-nonspecific-Mn2+ complex. In
addition to a high affinity for a TaqI-DNA complex, Mn2+ also binds to TaqI in a
DNA-independent fashion. This may facilitate enzyme activation, which could account for the
observed relaxation in substrate specificity. Thus, the TaqI-DNA-Mn2+ could be formed by
either of two pathways: TaqI binding to DNA followed by the binding of Mn2+ or TaqI first
binding to Mn2+ followed by the addition of DNA. The inactive, nonspecific TaqI-star-Mg2+
complex virtually prohibits transition state interactions, but a TaqI-star-Mn2+ complex
attains a measurable single-turnover rate. In the late stages of the enzymatic cycle, high
affinity of Mn2+ to a TaqI-DNA complex and to the TaqI enzyme may also account for a slower
rate of product release.

<>

<1>Cao, X., Springer, N.M., Muszynski, M.G., Phillips, R.L., Kaeppler, S., Jacobsen, S.E.
<2>Conserved plant genes with similarity to mammalian de novo DNA methyltransferases.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>4979-4984
<6>2000
<7>DNA methylation plays a critical role in controlling states of gene activity in most
eukaryotic organisms, and it is essential for proper growth and development. Patterns of
methylation are established by de novo methyltransferases and maintained by maintenance
methyltransferase activities. The Dnmt3 family of de novo DNA methyltransferases has recently
been characterized in animals. Here we describe DNA methyltransferase genes from both
Arabidopsis and maize that show a high level of sequence similarity to Dnmt3, suggesting that
they encode plant de novo methyltransferases. Relative to all known eukaryotic
methyltransferases, these plant proteins contain a novel arrangement of the motifs required
for DNA methyltransferase catalytic activity. The N termini of these methyltransferases
contain a series of ubiquitin-associated (UBA) domains. UBA domains are found in several
ubiquitin pathway proteins and in DNA repair enzymes such as Rad23, and they may be involved
in ubiquitin binding. The presence of UBA domains provides a possible link between DNA
methylation and ubiquitin/proteasome pathways.

<>

<1>Cao, X.F., Aufsatz, W., Zilberman, D., Mette, M.F., Huang, M.S., Matzke, M., Jacobsen, S.E.
<2>Role of the DRM and CMT3 Methyltransferases in RNA-directed DNA methylation.
<3>Curr. Biol.
<4>13
<5>2212-2217
<6>2003
<7>RNA interference is a conserved process in which double-stranded RNA is processed into 21-25
nucleotide siRNAs that trigger posttranscriptional
gene silencing. In addition, plants display a phenomenon termed
RNA-directed DNA methylation (RdDM) in which DNA with sequence identity to
silenced RNA is de novo methylated at its cytosine residues. This
methylation is not only at canonical CpG sites but also at cytosines in
CpNpG and asymmetric sequence contexts. In this report, we study the role
of the DRM and CMT3 DNA methyltransferase genes in the initiation and
maintenance of RdDM. Neither drm nor cmt3 mutants affected the maintenance
of preestablished RNA-directed CpG methylation. However, drm mutants
showed a nearly complete loss of asymmetric methylation and a partial loss
of CpNpG methylation. The remaining asymmetric and CpNpG methylation was
dependent on the activity of CMT3, showing that DRM and CMT3 act
redundantly to maintain non-CpG methylation. These DNA methyltransferases
appear to act downstream of siRNAs, since drm1 drm2 cmt3 triple mutants
show a lack of non-CpG methylation but elevated levels of siRNAs. Finally,
we demonstrate that DRM activity is required for the initial establishment
of RdDM in all sequence contexts including CpG, CpNpG, and asymmetric
sites.

<>

<1>Cao, X.F., Jacobsen, S.E.
<2>Role of the Arabidopsis DRM methyltransferases in de novo DNA methylation and gene silencing.
<3>Curr. Biol.
<4>12
<5>1138-1144
<6>2002
<7>Proper DNA methylation patterning requires the complementary processes of de novo methylation
(the initial methylation of unmethylated DNA
sequences) and maintenance methylation (the faithful replication of
preexisting methylation). Arabidopsis has two types of
methyltransferases with demonstrated maintenance activity: MET1, which
maintains CpG methylation [1-3] and is homologous to mammalian DNMT1,
and CHROMOMETHYLASE 3 (CMT3), which maintains CpNpG (N = A, T, C, or G)
methylation [3,4] and is unique to the plant kingdom. Here we describe
loss-of-function mutations in the Arabidopsis DOMAINS REARRANGED
METHYLASE (DRM) genes [5] and provide evidence that they encode de novo
methyltransferases. drm1 drm2 double mutants retained preexisting CpG
methylation at the endogenous FWA locus but blocked de novo CpG
methylation that is normally associated with FWA transgene silencing.
Furthermore, drm1 drm2 double mutants blocked de novo CpNpG and
asymmetric methylation and gene silencing of the endogenous SUPERMAN
(SUP) gene, which is normally triggered by an inverted SUP repeat.
However, drm1 drm2 double mutants did not show reactivation of
previously established SUPERMAN epigenetic silenced alleles. Thus, drm
mutants prevent the establishment but not the maintenance of gene
silencing at FWA and SUP, suggesting that the DRMs encode the major de
novo methylation enzymes affecting these genes.

<>

<1>Cao, X.F., Jacobsen, S.E.
<2>Locus-specific control of asymmetric and CpNpG methylation by the DRM and CMT3 methyltransferase genes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>16491-16498
<6>2002
<7>Many plant, animal, and fungal genomes contain cytosine DNA methylation in asymmetric sequence
contexts (CpHpH, H = A, T, C). Although the enzymes responsible for this methylation are
unknown, it has been assumed that asymmetric methylation is maintained by the persistent
activity of de novo methyltransferases (enzymes capable of methylating previously unmodified
DNA). We recently reported that the domains rearranged methylase (DRM) genes are required for
de novo DNA methylation in Arabidopsis thaliana because drm1 drm2 double mutants lack the de
novo methylation normally associated with transgene silencing. In this study, we have used
bisulfite sequencing and Southern blot analysis to examine the role of the DRM loci in the
maintenance of asymmetric methylation. At some loci, drm1 drm2 double mutants eliminated all
asymmetric methylation. However, at the SUPERMAN locus, asymmetric methylation was only
completely abolished in drm1 drm2 chromomethylase 3 (cmt3) triple mutant plants. drm1 drm2
double mutants also showed a strong reduction of CpNpG (n = A, T, C, or G) methylation at some
loci, but not at others. The drm1 drm2 cmt3 triple mutant plants did not affect CpG
methylation at any locus tested, suggesting that the primary CpG methylases are encoded by the
MET1 class of genes. Although neither the drm1 drm2 double mutants nor the cmt3 single mutants
show morphological defects, drm1 drm2 cmt3 triple mutant plants show pleiotropic effects on
plant development. Our results suggest that the DRM and CMT3 genes act in a partially
redundant and locus-specific manner to control asymmetric and CpNpG methylation.

<>

<1>Cao, X.M., Chen, T., Xu, L.Z., Yao, L.S., Qi, J., Zhang, X.L., Yan, Q.L., Deng, Y.H., Guo, T.Y., Wang, J., Hu, K.X., Xu, B.L.
<2>Complete Genome Sequence of Wohlfahrtiimonas chitiniclastica Strain SH04, Isolated from Chrysomya megacephala Collected from Pudong International Airport  in China.
<3>Genome Announcements
<4>1
<5>e0011913
<6>2013
<7>Wohlfahrtiimonas chitiniclastica bacilli that live in the larvae of a parasitic fly were
recently isolated and are speculated to be the cause of fulminant
sepsis. Here we report and analyze the complete genome sequence of
Wohlfahrtiimonas chitiniclastica strain SH04. No complete genome sequence of a
Wohlfahrtiimonas chitiniclastica isolate has been documented previously.

<>

<1>Cao, Y., Liu, X.F., Zhang, H.L., Chen, Y.J., Hu, C.J.
<2>Draft Genome Sequence of the Human-Pathogenic Bacterium Vibrio alginolyticus E0666.
<3>Genome Announcements
<4>1
<5>e00686-13
<6>2013
<7>Vibrio alginolyticus is a Gram-negative halophilic bacterium with worldwide distribution. In
this work, we report the draft genome sequence of a V.
alginolyticus strain (E0666) isolated from Epinephelus coioides ascites in the
Shantou city of Guangdong Province, China.

<>

<1>Cao, Y., Tian, B., Liu, Y., Cai, L., Wang, H., Lu, N., Wang, M., Shang, S., Luo, Z., Shi, J.
<2>Genome Sequencing of Ralstonia solanacearum FQY_4, Isolated from a Bacterial Wilt Nursery Used for Breeding Crop Resistance.
<3>Genome Announcements
<4>1
<5>e00125-13
<6>2013
<7>Ralstonia solanacearum strain FQY_4 was isolated from a bacterial wilt nursery, which is used
for breeding crops for Ralstonia resistance in China. Here, we
report the complete genome sequence of FQY_4 and its comparison with other
published R. solanacearum genomes, especially with the strains GMI1000 and Y45 in
the same group.

<>

<1>Capowski, E.E., Wells, J.M., Karrer, K.M.
<2>Maintenance of methylation patterns in Tetrahymena thermophila.
<3>Gene
<4>74
<5>103-104
<6>1988
<7>Historically, the maintenance of methylation paterns in differentiated cells has been
attributed to the action of a methyltransferase that functions after DNA replication and whose
target is palindromic, hemimethylated sites. This maintenance methyltransferase is thought to
methylate the daughter strand of newly replicated DNA, thus retaining the pattern of the
parent through successive divisions. We have begun a molecular anlaysis of the maintenance of
methylation patterns during somatic proliferation in the ciliated protozoan Tetrahymena
thermophila. The data we have accumulated indicate that this process is more complex than a
semiconservative copying model.

<>

<1>Capozzi, V., Russo, P., Lamontanara, A., Orru, L., Cattivelli, L., Spano, G.
<2>Genome Sequences of Five Oenococcus oeni Strains Isolated from Nero Di Troia Wine from the Same Terroir in Apulia, Southern Italy.
<3>Genome Announcements
<4>2
<5>e01077-14
<6>2014
<7>Oenococcus oeni is the principal lactic acid bacterium responsible for malolactic fermentation
in wine. Here, we announce the genome sequences of five O. oeni
strains isolated from Nero di Troia wine undergoing spontaneous malolactic
fermentation, and we report, for the first time, several genome sequences of
strains isolated from the same terroir.

<>

<1>Cappelletti, M., Di Gennaro, P., D'Ursi, P., Orro, A., Mezzelani, A., Landini, M., Fedi, S., Frascari, D., Presentato, A., Zannoni, D., Milanesi, L.
<2>Genome Sequence of Rhodococcus sp. Strain BCP1, a Biodegrader of Alkanes and Chlorinated Compounds.
<3>Genome Announcements
<4>1
<5>e00657-13
<6>2013
<7>Rhodococcus sp. strain BCP1 cometabolizes chlorinated compounds and mineralizes a broad range
of alkanes, as it is highly tolerant to them. The high-quality draft
genome sequence of Rhodococcus sp. strain BCP1, consisting of 6,231,823 bp, with
a G+C content of 70.4%, 5,902 protein-coding genes, and 58 RNA genes, is
presented here.

<>

<1>Carattoli, A., Villa, L., Poirel, L., Bonnin, R.A., Nordmann, P.
<2>Evolution of IncA/C blaCMY-2-Carrying Plasmids by Acquisition of the blaNDM-1 Carbapenemase Gene.
<3>Antimicrob. Agents Chemother.
<4>56
<5>783-786
<6>2012
<7>The bla(NDM-1) gene has been reported to be often located on broad-host-range
plasmids of the IncA/C type in clinical but also environmental bacteria recovered
from the New Delhi, India, area. IncA/C-type plasmids are the main vehicles for
the spread of the cephalosporinase gene bla(CMY-2), frequently identified in the
United States, Canada, and Europe. In this study, we completed the sequence of
IncA/C plasmid pNDM-KN carrying the bla(NDM-1) gene, recovered from a Klebsiella
pneumoniae isolate from Kenya. This sequence was compared with those of three
IncA/C-type reference plasmids from Escherichia coli, Yersinia ruckeri, and
Photobacterium damselae. Comparative analysis showed that the bla(NDM-1) gene was
located on a widely diffused plasmid scaffold known to be responsible for the
spread of bla(CMY-2)-like genes and consequently for resistance to broad-spectrum
cephalosporins. Considering that IncA/C plasmids possess a broad host range, this
scaffold might support a large-scale diffusion of the bla(NDM-1) gene among
Gram-negative rods.

<>

<1>Carballeira, N., Jorge, Y., Guerra, M., Madrazo, J.J., Lleonart, R., Silva, A., Garcia, O., De La Fuente, J., Herrera, L.
<2>Cloning and characterization of the PvuII restriction-modification system in Proteus vulgaris.  Purification of recombinant restrictase.
<3>Biotecnol. Apl.
<4>7
<5>167-175
<6>1990
<7>The PvuII restriction-modification system (RMS) from P. vulgaris was cloned and
expressed restrictase and methylase were studied in different E. coli strains.
Only the E. coli strain HB101 (mcrB-) was efficiently transformed by plasmids
carrying the RMSII PvuII.  A new procedure for the purification of recombinant
PvuII restrictase was applied, using ion exchange chromatography.  The methods
described in this paper allow the obtention of high amounts of PvuII
restrictase free of contaminants.

<>

<1>Carbone, I., Anderson, J.B., Kohn, L.M.
<2>A group-I intron in the mitochondrial small subunit ribosomal RNA gene of Sclerotinia sclerotiorum.
<3>Curr. Genet.
<4>27
<5>166-176
<6>1995
<7>A 1,380-bp intervening sequence within the mitochondrial small subunit
ribosomal RNA (mt SSU rRNA) gene of the fungus Sclerotinia sclerotiorum
has been sequenced and identified as a group-I intron. This is the first
report of an intron in the mt SSU rRNA gene. The intron shows close
similarity in secondary structure to the subgroup-IC2 introns from
Podospora (ND3i1, ND5i2, and COIi5) and Neurospora (ND5i1). The intron has
an open reading frame (ORF) that encodes a putative protein of 420 amino
acids which contains two copies of the LAGLI-DADG motif. The ORF belongs
to a family of ORFs identified in Podospora (ND3i1, ND4Li1, ND4Li2, ND5i2,
and COIi5) and Neurospora (ND5i1). The putative 420-aa polypeptide is also
similar to a site-specific endonuclease in the chloroplast large subunit
ribosomal RNA (LSU rRNA) gene of the green alga Chlamydomonas eugametos.
In each clone of S. sclerotiorum examined, including several clones which
were sampled over a 3-year period from geographically separated sites, all
isolates either had the intron or lacked the intron within the mt SSU rRNA
gene. Screening by means of Southern hybridization and PCR amplification
detected the intron in the mt SSU rRNA genes of S. minor, S. trifoliorum
and Sclerotium cepivorum, but not in other members of the Sclerotiniaceae,
such as Botrytis anamorphs of Botryotinia spp., or in other ascomycetous
and basidiomycetous fungi.

<>

<1>Card, C.O., Wilson, G.G., Weule, K., Hasapes, J., Kiss, A., Roberts, R.J.
<2>Cloning and characterization of the HpaII methylase gene.
<3>Nucleic Acids Res.
<4>18
<5>1377-1383
<6>1990
<7>The HpaII restriction-modification system from Haemophilus parainfluenzae
recognizes the DNA sequence CCGG.  The gene for the HpaII methylase has been
cloned into E. coli and its nucleotide sequence has been determined.  The DNA
of the clones is fully protected against cleavage by the HpaII restriction
enzyme in vitro, indicating that the methylase gene is active in E. coli.  The
clones were isolated in an McrA- strain of E. coli; attempts to isolate them in
an McrA+ strain were unsuccessful.  The clones do not express detectable HpaII
restriction endonuclease activity, suggesting that either the endonuclease gene
is not expressed well in E. coli, or that it is not present in its entirety in
any of the clones that we have isolated.  The derived amino acid sequence of
the HpaII methylase shows overall similarity to other cytosine methylases.  It
bears a particularly close resemblance to the sequences of the HhaI, BsuFI and
MspI methylases.  When compared with three other methylases that recognize
CCGG, the variable region of the HpaII methylase, which is believed to be
responsible for sequence specific recognition, shows some similarity to the
corresponding regions of the BsuFI and MspI methylases, but is rather
dissimilar to that of the SPR methylase.

<>

<1>Cardenas, J.P., Lazcano, M., Ossandon, F.J., Corbett, M., Holmes, D.S., Watkin, E.
<2>Draft Genome Sequence of the Iron-Oxidizing Acidophile Leptospirillum ferriphilum Type Strain DSM 14647.
<3>Genome Announcements
<4>2
<5>e01153-14
<6>2014
<7>The genomic features of the Leptospirillum ferriphilum type strain DSM 14647 are  described
here. An analysis of the predicted genes enriches our knowledge of the
molecular basis of iron oxidation, improves our understanding of its role in
industrial bioleaching, and suggests how it is adapted to live at extremely low
pH.

<>

<1>Cardoso, M.C., Leonhardt, H.
<2>DNA methyltransferase is actively retained in the cytoplasm during early development.
<3>J. Cell Biol.
<4>147
<5>25-32
<6>1999
<7>The overall DNA methylation level sharply decreases from the zygote to the blastocyst stage
despite the presence of high levels of DNA methyltransferase (Dnmt1). Surprisingly, the enzyme
is localized in the cytoplasm of early embryos despite the presence of several functional
nuclear localization signals. We mapped a region in the NH(2)-terminal, regulatory domain of
Dnmt1 that is necessary and sufficient for cytoplasmic retention during early development.
Altogether, our results suggest that Dnmt1 is actively retained in the cytoplasm, which
prevents binding to its DNA substrate in the nucleus and thereby contributes to the erasure of
gamete-specific epigenetic information during early mammalian development.

<>

<1>Carignani, G., Groudinsky, O., Frezza, D., Schiavon, E., Bergantino, E., Slonimski, P.P.
<2>An mRNA maturase is encoded by the first intron of the mitochondrial gene for the subunit I of cytochrome oxidase in S. cerevisiae.
<3>Cell
<4>35
<5>733-742
<6>1983
<7>We have localized ten oxi3- mutations in the first, ai1, intron of the coxI gene.  All are
splicing deficient, being unable to excise the intron.  Complementation experiments disclose
several domains in the intron ai1: the 5'-proximal and 3'-proximal domains harbor
cis-dominant mutations, while trans-recessive ones are located in the intron's open reading
frame.  Comprehensive analyses of allele-specific polypeptides accumulating in mutants show
that they result from the translation of the intron's ORF;  We conclude that a specific mRNA
maturase involved in splicing of oxidase mRNA is encoded by the intron ai1 in a manner similar
to the cytochrome b mRNA maturase.

<>

<1>Carle, P., Saillard, C., Carrere, N., Carrere, S., Duret, S., Eveillard, S., Gaurivaud, P., Gourgues, G., Gouzy, J., Salar, P., Verdin, E., Breton, M., Blanchard, A., Laigret, F., Bove, J.M., Renaudin, J., Foissac, X.
<2>Partial chromosome sequence of Spiroplasma citri reveals extensive viral invasion and important gene decay.
<3>Appl. Environ. Microbiol.
<4>76
<5>3420-3426
<6>2010
<7>The assembly of 20,000 sequencing reads obtained from shotgun and chromosome-specific
libraries of the Spiroplasma citri genome yielded 77
chromosomal contigs totaling 1,674 kbp (92%) of the 1,820-kbp chromosome.
The largest chromosomal contigs were positioned on the physical and
genetic maps constructed from pulsed-field gel electrophoresis and
Southern blot hybridizations. Thirty-eight contigs were annotated,
resulting in 1,908 predicted coding sequences (CDS) representing an
overall coding density of only 74%. Cellular processes, cell metabolism,
and structural-element CDS account for 29% of the coding capacity, CDS of
external origin such as viruses and mobile elements account for 24% of the
coding capacity, and CDS of unknown function account for 47% of the coding
capacity. Among these, 21% of the CDS group into 63 paralog families. The
organization of these paralogs into conserved blocks suggests that they
represent potential mobile units. Phage-related sequences were
particularly abundant and include plectrovirus SpV1 and SVGII3 and
lambda-like SpV2 sequences. Sixty-nine copies of transposases belonging to
four insertion sequence (IS) families (IS30, IS481, IS3, and ISNCY) were
detected. Similarity analyses showed that 21% of chromosomal CDS were
truncated compared to their bacterial orthologs. Transmembrane domains,
including signal peptides, were predicted for 599 CDS, of which 58 were
putative lipoproteins. S. citri has a Sec-dependent protein export
pathway. Eighty-four CDS were assigned to transport, such as
phosphoenolpyruvate phosphotransferase systems (PTS), the ATP binding
cassette (ABC), and other transporters. Besides glycolytic and ATP
synthesis pathways, it is noteworthy that S. citri possesses a nearly
complete pathway for the biosynthesis of a terpenoid.

<>

<1>Carlier, A., Agnoli, K., Pessi, G., Suppiger, A., Jenul, C., Schmid, N., Tummler, B., Pinto-Carbo, M., Eberl, L.
<2>Genome Sequence of Burkholderia cenocepacia H111, a Cystic Fibrosis Airway Isolate.
<3>Genome Announcements
<4>2
<5>e00298-14
<6>2014
<7>The Burkholderia cepacia complex (BCC) is a group of related bacterial species that are
commonly isolated from environmental samples. Members of the BCC can cause respiratory
infections in cystic fibrosis patients and immunocompromised individuals. We report here the
genome sequence of Burkholderia cenocepacia H111, a well-studied model strain of the BCC.

<>

<1>Carlson, K., Kosturko, L.D.
<2>Endonuclease II of coliphage T4: a recombinase disguised as a restriction endonuclease?
<3>Mol. Microbiol.
<4>27
<5>671-676
<6>1998
<7>Endo ll shares with restriction endonucleases the property of cleaving foreign DNA while
leaving the endonuclease-encoding genome intact, ensuring the survival of one DNA species in
the cell.  In addition, in vivo Endo ll cleaves a specific DNA sequence and cleavage is
context dependent.  These context effects extend over at least 1000 bp, largely limiting
cleavage to once within this distance.  Like homing endonucleases, in vivo Endo ll recognizes
a long, asymmetric and degenerate consensus sequence which has two distinct parts.
Recognition of one part of the consensus sequence involves base-specific bonds, and
recognition of the other involves sequence-dependent helical structure.  Endo ll fulfils an
obvious short-term survival role in ensuring the dominance of phase DNA in an infected cell,
but may also have a long-term evolutionary role, producing gene-size fragments of foreign DNA
to be enrolled in the phage genetic repertoire.

<>

<1>Carlson, K., Krabbe, M., Nystrom, A.C., Kosturko, L.D.
<2>DNA determinants of restriction.
<3>J. Biol. Chem.
<4>268
<5>8908-8918
<6>1993
<7>Endonuclease II of coliphage T4 is necessary for the in vivo restriction of plasmid DNA in
phage-infected cells. Double-stranded restriction cleavage at 12 sites in pBR322 commenced
before 10-min postinfection with T4 at 37C and proceeded more slowly in the presence of
competing phage DNA than in its absence, utilizing the same sites in both cases; in a 200-base
pair segment of the plasmid, single stranded nicks also were frequent. The plasmid sites were
cleaved with a speed that varied with the site, yielding frequencies of cleavage at different
sites varying between 10 and 90%, at 50-min postinfection. All sites contained good matches to
a consensus, 5'-GRCCGCNTYGC-3', most frequently cleaved around the variable central base
pair, generating fragments with blunt ends or 1-2-base 5' overhangs. Using the frequency of
cleavage to determine a weighted consensus, a larger sequence, 5'-CGRCCGCNTTGSYNGC-3' was
identified. Thus, DNA sequence elements 3' to the cut site appear important for rapid
cleavage. Several models describing the sequence-dependent structure of DNA suggest structural
anomalies around the cleavage sites. The endonuclease II restriction system is most similar to
type II systems, although it differs from known type II systems in several respects.

<>

<1>Carlson, K., Raleigh, E.A., Hattman, S.
<2>Restriction and Modification.
<3>Molecular Biology of Bacteriophage T4, American Society for Microbiology, Karam, J.D., Karam, J.D., Washington, D.C.
<4>
<5>369-381
<6>1994
<7>A host-parasite interaction is an intricate balance of mutual attack and defense.  Several
nucleases and other proteins encoded by Escherichia coli and T4 are involved in restriction
and modification, processes in which an organism causes degradation of "foreign" DNA while
modifying its own DNA to provide "immunity" to the restriction endonuclease(s).  Restriction
endonucleases encoded by the host (E. coli, EcoK or EcoB, Mcr {Rgl}, and Mrr) and its prophage
symbiont phage P1 (EcoP1) may restrict infecting phage DNA; in addition, E. coli exonuclease V
(ExoV; the RecBCD enzyme) and possibly endonuclease I may degrade phage DNA as it enters the
cell.  Phage-encoded nucleases, on the other hand, with endonuclease II (EndoII) as a crucial
participant, cause the restriction cleavage and ultimate breakdown of host DNA and other
cytosine-containing DNA in the cell.  The reutilization of host-derived nucleotides via a
"nucleotide scavenge pathway" permits the production of about 20 phage equivalents of phage
DNA per host genome scavenged.  Scavenging host DNA breakdown products this way undoubtedly is
evolutionarily advantageous to the phage when nutrients are limited.

<>

<1>Carlson, K., Wiberg, J.S.
<2>In vivo cleavage of cytosine-containing bacteriophage T4 DNA to genetically distinct, discretely sized fragments.
<3>J. Virol.
<4>48
<5>18-30
<6>1983
<7>Mutants of bacteriophage T4D that are defective in genes 42 (dCMP hydroxymethylase), 46 (DNA
exonuclease), and 56 (dCTPase) produce limited amounts of phage DNA in Escherichia coli B. In
this DNA, glucosylated 5-hydroxymethylcytosine is completely replaced by cytosine. We found
that this DNA rapidly becomes fragmented in vivo to at least 16 discrete bands as visualized
on agarose gels subjected to electrophoresis. The sizes of the fragments ranged from more than
20 to less than 2 kilobase pairs. When DNAs from two of these bands were radioactively labeled
in vitro by nick translation and hybridized to XbaI restriction fragments of
cytosine-containing T4 DNA, evidence was obtained that the two bands are genetically distinct,
i.e., they contain DNA from different parts of the T4 genome. Mutational inactivation of T4
endonuclease II (gene denA) prevented the fragmentation. Three different mutations in T4
endonuclease IV (gene denB) caused the same minor changes in the pattern of fragments. We
conclude that T4 endonuclease II is required, and endonuclease IV is involved to a minor
extent, in the in vivo production of these cytosine-containing T4 DNA fragments. We view these
DNA fragments as "restriction fragments" since they represent degradation products of DNA
"foreign" to T4, they are of discrete size, and they are genetically distinct. Thus, this
report may represent the first, direct in vivo demonstration of discretely sized genetically
distinct DNA restriction fragments.

<>

<1>Carlson, L.L., Page, A.W., Bestor, T.H.
<2>Properties and localization of DNA methyltransferase in preimplantation mouse embryos: implications for genomic imprinting.
<3>Genes Dev.
<4>6
<5>2536-2541
<6>1992
<7>Preimplantation mouse embryos contain very high levels of DNA methyltransferase activity. We
show here that the form of DNA methyltransferase (DNA MTase) in early embryos differs from the
form found in other cells and tissues by a slightly higher mobility on gel electrophoresis.
Levels of DNA MTase were found to be very high throughout preimplantation development even
though levels of 5-methylcytosine (m5C)in nuclear DNA are known to undergo a substantial
decline in the same period. Confocal laser scanning microscopy of mouse embryos stained with
DNA MTase-specific antibodies showed striking developmentally regulated changes in the
distribution of DNA MTase. From the oocyte stage to the four-cell-stage, most DNA MTase was
concentrated in peripheral cytoplasm, and nuclei did not contain detectable DNA MTase. In
four-and eight-cell embryos, DNA MTase was seen in cytoplasmic granules; and in eight-cell
embryos, DNA MTase was also present in large amounts in nuclei. Nuclei of blastocysts stained
only faintly, whereas the cytoplasmic granules remained prominent. Paradoxically, DNA MTase
was found to be at its highest levels in nuclei at a developmental stage where levels of m5C
in DNA are decreasing most rapidly. Changes in methylation patterns in preimplanation embryos
are therefore proposed to be under the control of unidentified regulatory factors rather than
DNA MTase itself; these regulatory factors could be members of the group that contains the
products of the Ssm-1 and Imp-1 genes, which are involved in the regulation of genomic
imprinting.

<>

<1>Carneiro, A.R. et al.
<2>Genome Sequence of Exiguobacterium antarcticum B7, Isolated from a Biofilm in Ginger Lake, King George Island, Antarctica.
<3>J. Bacteriol.
<4>194
<5>6689-6690
<6>2012
<7>Exiguobacterium antarcticum is a psychotropic bacterium isolated for the first time from
microbial mats of Lake Fryxell in Antarctica. Many organisms of the
genus Exiguobacterium are extremophiles and have properties of biotechnological
interest, e.g., the capacity to adapt to cold, which make this genus a target for
discovering new enzymes, such as lipases and proteases, in addition to improving
our understanding of the mechanisms of adaptation and survival at low
temperatures. This study presents the genome of E. antarcticum B7, isolated from
a biofilm sample of Ginger Lake on King George Island, Antarctic peninsula.

<>

<1>Carneiro, A.R., Juca, R.R.T., Barauna, R.A., de Sa, P.H., Marinho, A.D., Barbosa, S., Pereira, A., Alves, A., Egas, C., Correia, A., Henriques, I., Silva, A.
<2>Draft Genome Sequence of Serratia fonticola LMG 7882T Isolated from Freshwater.
<3>Genome Announcements
<4>1
<5>e00971-13
<6>2013
<7>Serratia fonticola is a Gram-negative bacterium with a wide distribution in aquatic
environments. On some occasions, it has also been regarded as a
significant human pathogen. In this work, we report the first draft genome
sequence of an S. fonticola strain (LMG 7882T), which was isolated from
freshwater.

<>

<1>Carnilli, R., Del Grosso, M., Lannelli, F., Pantostil, A.
<2>New genetic element carrying the erythromycin resistance determinant erm (TR) in Streptococcus pneumoniae.
<3>Antimicrob. Agents Chemother.
<4>52
<5>619-625
<6>2008
<7>erm(A) subclass erm(TR), a common macrolide resistance determinant in Streptococcus pyogenes
but quite rare in Streptococcus pneumoniae, was
found in a clinical S. pneumoniae isolate (AP200) from Italy. In this
isolate, erm(TR) was found included in a genetic element approximately
56 kb in size that did not appear to be conjugative but could be
transferred by transformation. An erm (TR)-containing DNA fragment of
approximately 10 kb was sequenced and 12 open reading frames (ORFs)
were identified. Upstream of erm(TR), a regulatory protein of the TOR
family and the two components of an efflux pump of the ABC type were
found. Downstream of erm(TR), there were ORFs homologous to a
spectinomycin phosphotransferase, transposases, and a relaxase. Since
the genomic sequence of S. pyogenes MGAS10750 carrying erm(TR) became
available, comparison between the erm (TR) -containing genetic elements
in AP200 and in MGAS10750 was performed. The region flanking erm(TR) in
MGAS10750 showed identity with AP200 for 10 ORFs out of 12. PCR mapping
using primers designed on the sequence of MGAS10750 confirmed that
AP200 carries a genetic element similar to that of MGAS10750. In AP200
the genetic element was inserted inside an ORF homologous to spr0790 of
S. pneumoniae R6, coding for a type I restriction modification system.
Homologies between the insertion sites in AP200 and MGAS10750 consisted
of eight conserved nucleotides, of which three were duplicated, likely
representing target site duplication. The structure of the erm
(TR)-carrying genetic element shows characteristics of a
transposon/prophage remnant chimera. In AP200 this genetic element was
designated Tn1806.

<>

<1>Caro-Quintero, A., Auchtung, J., Deng, J., Brettar, I., Hofle, M., Tiedje, J.M., Konstantinidis, K.T.
<2>Genome Sequencing of Five Shewanella baltica Strains Recovered from the Oxic-Anoxic Interface of the Baltic Sea.
<3>J. Bacteriol.
<4>194
<5>1236
<6>2012
<7>Here we describe five Shewanella baltica genomes recovered from the same sample,  as well as
12 years apart from the same sampling station. These genomes expand
the collection of previously sequenced S. baltica strains and represent a
valuable resource for assessing the role of environmental settings on genome
adaptation.

<>

<1>Carotti, D., Funiciello, S., Lavia, P., Caiafa, P., Strom, R.
<2>Different effects of histone H1 on de novo DNA methylation in vitro depend on both the DNA base composition and the DNA methyltransferase.
<3>Biochemistry
<4>35
<5>11660-11667
<6>1996
<7>We have characterized the inhibition exerted by histone H1 on the activity of human placenta
DNA (cytosine-5-)-methyltransferase.  Our experiments demonstrate that the extent of
inhibition depends on the DNA base composition, AT-rich substrates being more severely
affected than GC-rich substrates and CpG-rich islands.  With bacterial SssI methylase, the
effect is completely reversed since its activity on AT-rich substrates undergoes a 4-5-fold
stimulation upon the addition of H1.  Poly(L-lysine) mimicks H1 effects, suggesting an
essential role of lysine residues in both the inhibitory and stimulatory effects of H1.  By
comparison of the different behaviors of the two enzymes, the inhibitory effect over the
eukaryotic enzyme might be accounted for by hypothesizing a competition between minor
groove-binding motifs (SPKK-like) present in placenta methylase as well as in histone H1.

<>

<1>Carotti, D., Funiciello, S., Palitti, F., Strom, R.
<2>Influence of pre-existing methylation on the de novo activity of eukaryotic DNA methyltransferase.
<3>Biochemistry
<4>37
<5>1101-1108
<6>1998
<7>Aberrant de novo methylation of CpG island DNA sequences has been observed in cultured cell
lines or upon malignant transformation but the mechanisms underlying this phenomenon are
poorly understood.  Using eukaryotic DNA (cytosine-5)-methyltransferase (of both human and
murine origin), we have studied the in vitro methylation pattern of three CpG islands.  Such
sequences are intrinsically poor substrates of the enzyme, yet are efficiently methylated when
a small amount of 5-methylcytosine is randomly introduced by the M.SssI prokaryotic DNA
(cytosine-5)-methyltransferase prior to in vitro methylation by the eukaryotic enzyme.  A
stimulation was also found with several other double-stranded DNA substrates, either natural
or of synthetic origin, such as poly(dG-dC).poly(dG-dC).  An A+T-rich plasmid, pHbbeta1S,
showed an initial stimulation, followed by a severe inhibition of the activity of DNA
(cytosine-5)-methyltransferase.  Methylation of poly(dI-dC).poly(dI-dC) was instead inhibited
by pre-existing 5-methylcytosines.  The extent of stimulation observed with
poly(dG-dC).poly(dG-dC) depends on both the number and the distribution of the
5-methylcytosine residues, which probably must not be too closely spaced for the stimulatory
effect to be exerted.  The activity of the M.SssI prokaryotic DNA methyltransferase was not
stimulated, but was inhibited by pre-methylation on either poly(dG-dC).poly(dG-dC) or
poly(dI-dC).poly(dI-dC).  The prokaryotic and eukaryotic DNA methyltransferases also differed
in sensitivity to poly(dG-m5dC).poly(dG-m5dC), which is highly inhibitory for eukaryotic
enzymes and almost ineffective on prokaryotic enzymes.

<>

<1>Carpenter, M., Divvela, P., Pingoud, V., Bujnicki, J., Bhagwat, A.S.
<2>Sequence-dependent enhancement of hydrolytic deamination of cytosines in DNA by the restriction enzyme PspGI.
<3>Nucleic Acids Res.
<4>34
<5>3762-3770
<6>2006
<7>Hydrolytic deamination of cytosines in DNA creates uracil and, if unrepaired, these lesions
result in C to T mutations. We have suggested
previously that a possible way in which cells may prevent or reduce this
chemical reaction is through the binding of proteins to DNA. We use a
genetic reversion assay to show that a restriction enzyme, PspGI, protects
cytosines within its cognate site, 5'-CCWGG (W is A or T), against
deamination under conditions where no DNA cleavage can occur. It decreases
the rate of cytosine deamination to uracil by 7-fold. However, the same
protein dramatically increases the rate of deaminations within the site
5'-CCSGG (S is C or G) by approximately 15-fold. Furthermore, a similar
increase in cytosine deaminations is also seen with a catalytically
inactive mutant of the enzyme showing that endonucleolytic ability of the
protein is dispensable for its mutagenic action. The sequences of the
mutants generated in the presence of PspGI show that only one of the
cytosines in CCSGG is predominantly converted to thymine. Our results are
consistent with PspGI 'sensitizing' the cytosine in the central base pair
in CCSGG for deamination. Remarkably, PspGI sensitizes this base for
damage despite its inability to form stable complexes at CCSGG sites.
These results can be explained if the enzyme has a transient interaction
with this sequence during which it flips the central cytosine out of the
helix. This prediction was validated by modeling the structure of
PspGI-DNA complex based on the structure of the related enzyme Ecl18kI
which is known to cause base-flipping.

<>

<1>Carpenter, M.A.
<2>Studies of base flipping by the Pyrococcus species GI-H restriction-modification system.
<3>Ph.D. Thesis, Wayne State Univ., Detroit, MI, USA
<4>
<5>
<6>2008
<7>Water is the single most DNA damaging agent that cells regularly encounter.  Water induces DNA
damage by a variety of mechanisms including depurination, depyrimidination, and deamination.
Figure 1 shows some of the most frequently observed types of DNA damage.  Depurination and
depyrimidination occur when a water molecular attacks at the C1' of deoxyribose kicking the
DNA base off in an Sn2 type substitution.  Depurination occurs at a very high rate, inducing
an estimated 18,000 abasic sites per cell, per day in a typical human cell.  Depyrimidination
occurs about 300 fold less frequently than depurination.  Repair of abasic sites in
Escherichia coli is similar to Base Excision Repair except that the abasic site is caused by
hydrolytic attack of water and not by a DNA glycosylase.

<>

<1>Carpenter, M.A., Bhagwat, A.S.
<2>DNA base flipping by both members of the PspGI restriction-modification system.
<3>Nucleic Acids Res.
<4>36
<5>5417-5425
<6>2008
<7>The PspGI restriction-modification system recognizes the sequence CCWGG. R.PspGI cuts DNA
before the first C in the cognate sequence and M.PspGI is
thought to methylate N4 of one of the cytosines in the sequence. M.PspGI
enhances fluorescence of 2-aminopurine in DNA if it replaces the second C
in the sequence, while R.PspGI enhances fluorescence when the fluorophore
replaces adenine in the central base pair. This strongly suggests that the
methyltransferase flips the second C in the recognition sequence, while
the endonuclease flips both bases in the central base pair out of the
duplex. M.PspGI is the first N4-cytosine MTase for which biochemical
evidence for base flipping has been presented. It is also the first type
IIP methyltransferase whose catalytic activity is strongly stimulated by
divalent metal ions. However, divalent metal ions are not required for its
base-flipping activity. In contrast, these ions are required for both base
flipping and catalysis by the endonuclease. The two enzymes have similar
temperature profiles for base flipping and optimal flipping occurs at
temperatures substantially below the growth temperature of the source
organism for PspGI and for the catalytic activity of endonuclease. We
discuss the implications of these results for DNA binding by these enzymes
and their evolutionary origin.

<>

<1>Carraway, M., Youderian, P., Marinus, M.G.
<2>Spontaneous mutations occur near dam recognition sites in a dam- Escherichia coli host.
<3>Genetics
<4>116
<5>343-347
<6>1987
<7>The mismatch repair system of Escherichia coli K12 removes mispaired bases from DNA. Mismatch
repair can occur on either strand of DNA if it lacks N6-methyladenines within
5'-GATC-3'sequences. In hemimethylated heteroduplexes, repair occurs preferentially on the
unmethylated strand. If both strands are fully methylated, repair is inhibited. Mutant (dam-)
strains of E. coli defective in the adenine methylase that recognizes 5'-GATC-3' sequences
(Dam), and therefore defective in mismatch repair, show increased spontaneous mutation rates
compared to otherwise isogenic dam+ hosts. We have isolated and characterized 91 independent
mutations that arise as a consequence of the Dam- defect in a plasmid-borne phage P22
repressor gene, mnt. The majority of these mutations are A:T----G:C transitions that occur
within six base pairs of the two 5'-GATC-3' sequences in the mnt gene. In contrast, the
spectrum of mnt- mutations in a dam+ host is comprised of a majority of insertions of IS
elements and deletions that do not cluster near Dam recognition sites. These results show that
Dam-directed post-replicative mismatch repair plays a significant role in the rectification of
potential transition mutations in vivo, and suggest that sequences associated with Dam
recognition sites are particularly prone to replication or repair errors.

<>

<1>Carrias, A., Welch, T.J., Waldbieser, G.C., Mead, D.A., Terhune, J.S., Liles, M.R.
<2>Comparative genomic analysis of bacteriophages specific to the channel catfish pathogen Edwardsiella ictaluri.
<3>Virol. J.
<4>8
<5>6
<6>2011
<7>BACKGROUND: The bacterial pathogen Edwardsiella ictaluri is a primary
cause of mortality in channel catfish raised commercially in aquaculture
farms. Additional treatment and diagnostic regimes are needed for this
enteric pathogen, motivating the discovery and characterization of
bacteriophages specific to E. ictaluri. RESULTS: The genomes of three
Edwardsiella ictaluri-specific bacteriophages isolated from geographically
distant aquaculture ponds, at different times, were sequenced and
analyzed. The genomes for phages eiAU, eiDWF, and eiMSLS are 42.80 kbp,
42.12 kbp, and 42.69 kbp, respectively, and are greater than 95% identical
to each other at the nucleotide level. Nucleotide differences were mostly
observed in non-coding regions and in structural proteins, with
significant variability in the sequences of putative tail fiber proteins.
The genome organization of these phages exhibit a pattern shared by other
Siphoviridae. CONCLUSIONS: These E. ictaluri-specific phage genomes reveal
considerable conservation of genomic architecture and sequence identity,
even with considerable temporal and spatial divergence in their isolation.
Their genomic homogeneity is similarly observed among E. ictaluri
bacterial isolates. The genomic analysis of these phages supports the
conclusion that these are virulent phages, lacking the capacity for
lysogeny or expression of virulence genes. This study contributes to our
knowledge of phage genomic diversity and facilitates studies on the
diagnostic and therapeutic applications of these phages.

<>

<1>Carrick, K.L., Topal, M.D.
<2>Amino Acid Substitutions at Position 43 of NaeI Endonuclease - Evidence for Changes in NaeI Structure.
<3>J. Biol. Chem.
<4>278
<5>9733-9739
<6>2003
<7>NaeI endonuclease contains a 10-amino acid region with sequence similarity to the active site
KXDG motif of DNA ligase except for leucine (Leu-43) in
NaeI ((43)LXDG(46)). Changing Leu-43 to lysine abolishes the NaeI
endonuclease activity and replaces it with topoisomerase and recombinase
activities. Here we report the results of substituting Leu-43 with
alanine, arginine, asparagine, glutamate, and histidine. Quantitating
specific activities and DNA binding values for the mutant proteins
determined the range of amino acids at position 43 that alter NaeI
mechanism. Substituting alanine, asparagine, glutamate, and histidine for
Leu-43 maintained endonuclease activity, but at a lower level. On the
other hand, substituting positively charged arginine, like lysine at
position 43, converted NaeI to a topoisomerase with no observable
double-strand cleavage activity. The specific activities of NaeI-43K and
NaeI-43R and their relative sensitivities to salt, the
topoisomerase-inhibiting drug
N-[4-(9-acridinylamino)-3-methoxyphenyl]methane-sulfonamide (amsacrine)
and single-stranded DNA showed that the two activities are similar. The
effect of placing a positive charge at position 43 on NaeI structure was
determined by measuring (for NaeI and NaeI-43K) relative susceptibilities
to proteolysis, UV, circular dichroism spectra, and temperature melting
transitions. The results provide evidence that a positive charge at
position 43 induces dramatic changes in NaeI structure that affect both
the Endo and Topo domains of NaeI. The identification of four putative DNA
ligase motifs in NaeI leads us to speculate that structural changes that
superimpose these motifs on the ligase structure may account for the
changes in activity.

<>

<1>Carroll, R.K., Burda, W.N., Roberts, J.C., Peak, K.K., Cannons, A.C., Shaw, L.N.
<2>Draft Genome Sequence of Strain CBD-635, a Methicillin-Resistant Staphylococcus aureus USA100 Isolate.
<3>Genome Announcements
<4>1
<5>e00491-13
<6>2013
<7>We present the draft genome sequence of methicillin-resistant Staphylococcus aureus strain
CBD-635, from the USA100 lineage. This is a sepsis isolate obtained
from Tampa General Hospital. This strain is spa type t003 and multilocus sequence
typing (MLST) type ST5, and it has been used by our group in the study of novel
antimicrobial chemotherapeutics.

<>

<1>Carruthers, M.D., Harding, C.M., Baker, B.D., Bonomo, R.A., Hujer, K.M., Rather, P.N., Munson, R.S.Jr.
<2>Draft Genome Sequence of the Clinical Isolate Acinetobacter nosocomialis Strain M2.
<3>Genome Announcements
<4>1
<5>e00906-13
<6>2013
<7>We report the 3.78-Mbp high-quality draft assembly of the genome from a clinical  isolate of
Acinetobacter nosocomialis called strain M2 (previously known as
Acinetobacter baumannii strain M2).

<>

<1>Carter, A.T., Pearson, B.M., Crossman, L.C., Drou, N., Heavens, D., Baker, D., Febrer, M., Caccamo, M., Grant, K.A., Peck, M.W.
<2>Complete Genome Sequence of proteolytic Clostridium botulinum type A5 (B3') Strain H04402 065.
<3>J. Bacteriol.
<4>193
<5>2351-2352
<6>2011
<7>H04402 065 is one of a very small group of strains of proteolytic Clostridium botulinum that
form type A5 neurotoxin. Here, we report the complete 3.9 Mb genome sequence and annotation of
strain H04402 065, which was isolated from a botulism patient in the UK in 2004.

<>

<1>Carter, D., Jen, S., Zhang, Y., Bhatai, A., Maisonneuve, J.L., Wang, S.S., Mitcham, J.L., Persing, D.H., Skeiky, Y.A.W.
<2>Compositions and Methods for the Therapy and Diagnosis of Acnes.
<3>Japanese Patent Office
<4>JP 2004520803 A
<5>
<6>2004
<7>
<>

<1>Carter, J.A., Chater, K.F., Bruton, C.J., Brown, N.L.
<2>A comparison of DNA cleavage by the restriction enzymes SalPI and PstI.
<3>Nucleic Acids Res.
<4>8
<5>4943-4954
<6>1980
<7>Methods for obtaining highly active, exonuclease-free, stable preparations of
the Streptomyces albus P restriction enzyme SalPI are described.  SalPI and its
isoschizomer PstI (from the taxonomically distant Providencia stuartii 164)
both cleave their recognition sequence (5'-CTGCAG-3') to generate fragments
terminating in tetranucleotide 3' extensions whose sequence is 5'-TGCA-3'.
Bacteriophage R4G2 DNA, protected against SalPI cleavage by pregrowth on S.
albus P, is also protected against PstI cleavage; and total DNA of both S.
albus P and P. stuartii 164 is resistant to cleavage by both enzymes.

<>

<1>Carter, J.M., Friedrich, N.C., Kleinstiver, B., Edgell, D.R.
<2>Strand-specific Contacts and Divalent Metal Ion Regulate Double-strand Break Formation by the GIY-YIG Homing Endonuclease I-BmoI.
<3>J. Mol. Biol.
<4>374
<5>306-321
<6>2007
<7>GIY-YIG homing endonucleases are modular enzymes consisting of a well-defined N-terminal
catalytic domain connected to a variable
C-terminal DNA-binding domain. Previous studies have revealed that the
role of the DNA-binding domain is to recognize and bind intronless DNA
substrate, positioning the N-terminal catalytic domain such that it is
poised to generate a staggered double-strand break by an unknown
mechanism. Interactions of the N-terminal catalytic domain with intronless
substrate are therefore a critical step in the reaction pathway but have
been difficult to define. Here, we have taken advantage of the reduced
activity of I-BmoI, an isoschizomer of the well-studied bacteriophage T4
homing endonuclease I-TevI, to examine double-strand break formation by
I-BmoI. We present evidence demonstrating that I-BmoI generates a
double-strand break by two sequential but chemically independent nicking
reactions where divalent metal ion is a limiting factor in top-strand
nicking. We also show by in-gel footprinting that contacts by the I-BmoI
catalytic domain induce significant minor groove DNA distortions that
occur independently of bottom-strand nicking. Bottom-strand contacts are
critical for accurate top-strand nicking, whereas top-strand contacts have
little influence on the accuracy of bottom-strand nicking. We discuss our
results in the context of current models of GIY-YIG endonuclease function,
with emphasis on the role of divalent metal ion and strand-specific
contacts in regulating the activity of a single active site to generate a
staggered double-strand break.

<>

<1>Carvin, C.D., Dhasarathy, A., Friesenhahn, L.B., Jessen, W.J., Kladde, M.P.
<2>Targeted cytosine methylation for in vivo detection of protein-DNA interactions.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>7743-7748
<6>2003
<7>We report a technique, named targeted gene methylation (TAGM), for identifying in vivo
protein-binding sites in chromatin. M.CviPI, a
cytosine-5 DNA methyltransferase recognizing GC sites, is fused to a
DNA-binding factor enabling simultaneous detection of targeted
methylation, factor footprints, and chromatin structural changes by
bisulfite genomic sequencing. Using TAGM with the yeast transactivator
Pho4, methylation enrichments of up to 34- fold occur proximal to native
Pho4-binding sites. Additionally, significant selective targeting of
methylation is observed several hundred nucleotides away, suggesting the
detection of long-range interactions due to higher-order chromatin
structure. In contrast, at an extragenic locus lacking Pho4-binding sites,
methylation levels are at the detection limit at early times after Pho4
transactivation. Notably, substantial amounts of methylation are targeted
by Pho4-M.CviPI under repressive conditions when most of the
transactivator is excluded from the nucleus. Thus, TAGM enables rapid
detection of DNA-protein interactions even at low occupancies and has
potential for identifying factor targets at the genome-wide level.
Extension of TAGM from yeast to vertebrates, which use methylation to
initiate and propagate repressed chromatin, could also provide a valuable
strategy for heritable inactivation of gene expression.

<>

<1>Carvin, C.D., Parr, R.D., Kladde, M.P.
<2>Site-selective in vivo targeting of cytosine-5 DNA methylation by zinc-finger proteins.
<3>Nucleic Acids Res.
<4>31
<5>6493-6501
<6>2003
<7>Cytosine-5 DNA methylation is a critical signal defining heritable epigenetic states of
transcription. As aberrant methylation patterns often
accompany disease states, the ability to target cytosine methylation to
preselected regions could prove valuable in re-establishing proper gene
regulation. We employ the strategy of targeted gene methylation in yeast,
which has a naturally unmethylated genome, selectively directing de novo
DNA methylation via the fusion of C5 DNA methyltransferases to
heterologous DNA-binding proteins. The zinc-finger proteins Zif268 and
Zip53 can target DNA methylation by M.CviPI or M.SssI 5-52 nt from single
zinc-factor binding sites. Modification at specific GC (M.CviPI) or CG
(M.SssI) sites is enhanced as much as 20-fold compared with strains
expressing either the free enzyme or a fusion protein with the zinc-finger
protein moiety unable to bind to DNA. Interestingly, methylation is also
selectively targeted as far as 353 nt from the zinc-finger protein binding
sites, possibly indicative of looping, nucleosomes or higher-order
chromatin structure. These data demonstrate that methylation can be
targeted in vivo to a potentially broad range of sequences using
specifically engineered zinc-finger proteins. Further more, the selective
targeting of methylation by zinc-finger proteins demonstrates that binding
of distinct classes of factors can be monitored in living cells.

<>

<1>Casadesus, J., Low, D.
<2>Epigenetic gene regulation in the bacterial world.
<3>Microbiol. Mol. Biol. Rev.
<4>70
<5>830-856
<6>2006
<7>Like many eukaryotes, bacteria make widespread use of postreplicative DNA methylation for the
epigenetic control of DNA-protein interactions. Unlike eukaryotes, however, bacteria use DNA
adenine methylation (rather than DNA cytosine methylation) as an epigenetic signal. DNA
adenine methylation plays roles in the virulence of diverse pathogens of humans and livestock
animals, including pathogenic Escherichia coh, Salmonella, Vibrio, Yersinia, Haemophilus, and
Brucella. In Alphaproteobactetia, methylation of adenine at GANTC sites by the CcrM methylase
regulates the cell cycle and couples gene transcription to DNA replication. In
Gammaproteobacteria, adenine methylation at GATC sites by the Dam methylase provides signals
for DNA replication, chromosome segregation, mismatch repair, packaging of bacteriophage
genomes, transposase activity, and regulation of gene expression. Transcriptional repression
by Dam methylation appears to be more common than transcriptional activation. Certain
promoters are active only during the hemimethylation interval that follows DNA replication;
repression is restored when the newly synthesized DNA strand is methylated In the E. coli
genome, however, methylation of specific GATC sites can be blocked by cognate DNA binding
proteins. Blockage of GATC methylation beyond cell division permits transmission of DNA
methylation patterns to daughter cells and can give rise to distinct epigenetic states, each
propagated by a positive feedback loop. Switching between alternative DNA methylation patterns
can split clonal bacterial populations into epigenetic lineages in a manner reminiscent of
eukaryotic cell differentiation. Inheritance of self-propagating DNA methylation patterns
governs phase variation in the E. coli pap operon, the agn43 gene, and other loci encoding
virulence-related cell surface functions.

<>

<1>Casadesus, J., Torreblanca, J.
<2>Methylation-related epigenetic signals in bacterial DNA.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, V.E.A. Russo, R.A. Martiensson, A.D. Riggs, Cold Spring Harbor, NY
<4>
<5>141-153
<6>1996
<7>The term "epigenetic signals"  may seem extraneous to the life-style of bacteria, because the
classic concept of epigenetics has been applied to the changes in gene expression that govern
differentiation and development.  However, because differentiation is often associated with
changes in DNA or chromatin structure, DNA modification has provided an attractive model for
the study of epigenetic regulation.

<>

<1>Caserta, M., Zacharias, W., Nwankwo, D., Wilson, G.G., Wells, R.D.
<2>Cloning, sequencing, in vivo promoter mapping, and expression in Escherichia coli of the gene for the HhaI methyltransferase.
<3>J. Biol. Chem.
<4>262
<5>4770-4777
<6>1987
<7>A 1476-base pair DNA fragment from Haemophilus haemolyticus containing the HhaI
methyltransferase gene was isolated from a cell library and cloned into pBR322.
The nucleotide sequence of this fragment was determined.  The structural gene
is 981 nucleotides in length coding for a protein of 327 amino acids (Mr
37,000).  The translational start signal (ATG) is preceded by the putative
ribosome-binding site (TAAG).  Recombinant plasmids containing the
1476-basepair fragment are completely methylated when isolated from Escherichia
coli, as judged by their insusceptibility to the HhaI restriction endonuclease.
However, the presence of an active HhaI methylase gene in certain E. coli
stsrains results in a very poor yield of transformants and/or in
vivo-originated deletions due to the Rgl functions of these hosts.  The in vivo
transcription and initiation sites have been identified by S1 protection and
primer-extension experiments using specific probes with total RNA prepared from
E. coli cells (HB101 or RR1) which tolerate the expression of MHhaI.

<>

<1>Casjens, S., Hayden, M., Jackson, E., Deans, R.
<2>Additional restriction endonuclease cleavage sites on the bacteriophage P22 genome.
<3>J. Virol.
<4>45
<5>864-867
<6>1983
<7>We present complete restriction endonuclease cleavage site maps of the
bacteriophage P22 chromosome for 16 enzymes with six base recognition
sequences, thereby positioning 116 new sites on the chromosome.  Twenty-four
such restriction maps for P22 DNA, containing 162 sites, have now been
completed, and three enzymes were found that did not cut P22 DNA.  Our results
are consistent with the ideas that ClaI does not cleave the methylated
recognition sequence ATCGMeAT or MeATCGAT and StuI does not cleave the
methylated recognition sequence AGGCMeCT.

<>

<1>Casjens, S., Palmer, N., van Vugt, R., Huang, W.M., Stevenson, B., Rosa, P., Lathigra, R., Sutton, G., Peterson, J., Dodson, R.J., Haft, D., Hickey, E., Gwinn, M., White, O., Fraser, C.M.
<2>A bacterial genome in flux: the twelve linear and nine circular extrachromosomal DNAs in an infectious isolate of the Lyme disease spirochete Borrelia burgdorferi.
<3>Mol. Microbiol.
<4>35
<5>490-516
<6>2000
<7>We have determined that Borrelia burgdorferi strain B31 MI carries 21
extrachromosomal DNA elements, the largest number known for any bacterium.
Among these are 12 linear and nine circular plasmids, whose sequences
total 610 694 bp. We report here the nucleotide sequence of three linear
and seven circular plasmids (comprising 290 546 bp) in this infectious
isolate. This completes the genome sequencing project for this organism;
its genome size is 1 521 419 bp (plus about 2000 bp of undetermined
telomeric sequences). Analysis of the sequence implies that there has been
extensive and sometimes rather recent DNA rearrangement among a number of
the linear plasmids. Many of these events appear to have been mediated by
recombinational processes that formed duplications. These many regions of
similarity are reflected in the fact that most plasmid genes are members
of one of the genome's 161 paralogous gene families; 107 of these gene
families, which vary in size from two to 41 members, contain at least one
plasmid gene. These rearrangements appear to have contributed to a
surprisingly large number of apparently non-functional pseudogenes, a very
unusual feature for a prokaryotic genome. The presence of these damaged
genes suggests that some of the plasmids may be in a period of rapid
evolution. The sequence predicts 535 plasmid genes >/=300 bp in length
that may be intact and 167 apparently mutationally damaged and/or
unexpressed genes (pseudogenes). The large majority, over 90%, of genes on
these plasmids have no convincing similarity to genes outside Borrelia,
suggesting that they perform specialized functions.

<>

<1>Casjens, S., Winn-Stapley, D.A., Gilcrease, E.B., Morona, R., Kuhlewein, C., Chua, J.E., Manning, P.A., Inwood, W., Clark, A.J.
<2>The chromosome of Shigella flexneri bacteriophage Sf6: complete nucleotide sequence, genetic mosaicism, and DNA packaging.
<3>J. Mol. Biol.
<4>339
<5>379-394
<6>2004
<7>Shigella flexneri temperate bacteriophage Sf6 is of interest in part because its
prophageexpresses the oac gene that alters the antigenic
properties of the surface O-antigen polysaccharide of its host bacterium.
We have determined the complete sequence of its 39,044 bp genome. The
sequence shows that Sf6 is a member of the canonical lambdoid phage group,
and like other phages of this type has a highly mosaic genome. It has
chromosomal regions that encode proteins >80% identical with at least 15
different previously characterized lambdoid phages and prophages, but 43%
of the genome, including the virion assembly genes, is homologous to the
genome of one phage, HK620. An analysis of the nucleotide differences
between Sf6 and HK620 indicates that even these similar regions are highly
mosaic. This mosaicism suggests ways in which the virion structural
proteins might interact with each other. The Sf6 early operons are
arranged like a typical lambdoid phage, with "boundary sequences" often
found between functional modules in the "metabolic" genome domain. By
virtue of high degree of similarity in the encoding genes and their DNA
target sites, we predict that the integrase, early transcription
anti-terminator, CI and Cro repressors, and CII protein of Sf6 have DNA
binding specificities very similar to the homologous proteins encoded byphages HK620, lambda,
434 and P22, respectively. The late operon contains
two tRNA genes. The Sf6 terminase genes are unusual. Analysis of in vivo
initiation of the DNA packaging series showed that the Sf6 apparatus that
recognizes DNA for packaging appears to cleave DNA for initiation of
packaging series at many sites within a large region of about 1800 bp that
includes a possible pac site. This is unlike previously characterized
phage packaging mechanisms.

<>

<1>Casjens, S.R., Fraser-Liggett, C.M., Mongodin, E.F., Qiu, W.G., Dunn, J.J., Luft, B.J., Schutzer, S.E.
<2>Whole Genome Sequence of an Unusual Borrelia burgdorferi Sensu Lato Isolate.
<3>J. Bacteriol.
<4>193
<5>1489-1490
<6>2011
<7>Human Lyme disease is caused by a number of related Borrelia burgdorferi sensu lato species.
We report here the complete genome sequence of
Borrelia sp. isolate SV1 from Finland. This isolate is to date the closest
known relative of B. burgdorferi sensu stricto, but it is sufficiently
genetically distinct from that species that it and its close relatives
warrant its candidacy for new-species status. We suggest that this isolate
should be named 'Borrelia finlandensis.'

<>

<1>Casjens, S.R., Gilcrease, E.B., Huang, W.M., Bunny, K.L., Pedulla, M.L., Ford, M.E., Houtz, J.M., Hatfull, G.F., Hendrix, R.W.
<2>The pKO2 Linear Plasmid Prophage of Klebsiella oxytoca.
<3>J. Bacteriol.
<4>186
<5>1818-1832
<6>2004
<7>Temperate bacteriophages with plasmid prophages are uncommon in nature, and of these only
phages N15 and PY54 are known to have a linear plasmid prophage with closed hairpin telomeres.
We report here the complete nucleotide sequence of the 51,601-bp Klebsiella oxytoca linear
plasmid pKO2, and we demonstrate experimentally that it is also a prophage. We call this
bacteriophage phiKO2. An analysis of the 64 predicted phiKO2 genes indicate that it is a
fairly close relative of phage N15; they share a mosaic relationship that is typical of
different members of double-stranded DNA tailed-phage groups. Although the head, tail shaft,
and lysis genes are not recognizably homologous between these phages, other genes such as the
plasmid partitioning, replicase, prophage repressor, and protelomerase genes (and their
putative targets) are so similar that we predict that they must have nearly identical DNA
binding specificities. The phiKO2 virion is unusual in that its phage lambda-like tails have
an exceptionally long (3,433 amino acids) central tip tail fiber protein. The phiKO2 genome
also carries putative homologues of bacterial dinI and umuD genes, both of which are involved
in the host SOS response. We show that these divergently transcribed genes are regulated by
LexA protein binding to a single target site that overlaps both promoters.

<>

<1>Casjens, S.R., Mongodin, E.F., Qiu, W.G., Dunn, J.J., Luft, B.J., Fraser-Liggett, C.M., Schutzer, S.E.
<2>Whole-Genome Sequences of Two Borrelia afzelii and Two Borrelia garinii Lyme Disease Agent Isolates.
<3>J. Bacteriol.
<4>193
<5>6995-6996
<6>2011
<7>Human Lyme disease is commonly caused by several species of spirochetes in the Borrelia genus.
In Eurasia these species are largely Borrelia afzelii,
B. garinii, B. burgdorferi, and B. bavariensis sp. nov. Whole-genome
sequencing is an excellent tool for investigating and understanding the
influence of bacterial diversity on the pathogenesis and etiology of Lyme
disease. We report here the whole-genome sequences of four isolates from
two of the Borrelia species that cause human Lyme disease, B. afzelii
isolates ACA-1 and PKo and B. garinii isolates PBr and Far04.

<>

<1>Caspers, M.P., Boekhorst, J., Abee, T., Siezen, R.J., Kort, R.
<2>Complete Genome Sequence of Anoxybacillus flavithermus TNO-09.006, a Thermophilic Sporeformer Associated with a Dairy-Processing Environment.
<3>Genome Announcements
<4>1
<5>e00010-13
<6>2013
<7>Spores of thermophilic spore-forming bacteria are a common cause of contamination in dairy
products. We isolated the thermophilic strain TNO-09.006 from a
milk-processing plant, and we report the complete genome of this isolate
consisting of a single chromosome of 2.65 Mb.

<>

<1>Cassir, N., Croce, O., Pagnier, I., Benamar, S., Couderc, C., Robert, C., Raoult, D., La Scola, B.
<2>Non-contiguous finished genome sequence and description of Bacteroides neonati sp. nov., a new species of anaerobic bacterium.
<3>Standards in Genomic Sciences
<4>9
<5>794-806
<6>2014
<7>Bacteroides neonati strain MS4(T), is the type strain of Bacteroides neonati sp.  nov., a new
species within the genus Bacteroides. This strain, whose genome is
described here, was isolated from a premature neonate stool sample. B. neonati
strain MS4(T) is an obligate anaerobic Gram-negative bacillus. Here we describe
the features of this organism, together with the complete genome sequence and
annotation. The 5.03 Mbp long genome exhibits a G+C content of 43.53% and
contains 4,415 protein-coding and 91 RNA genes, including 9 rRNA genes.

<>

<1>Castado, C., Thonnard, J.
<2>Novel compounds.
<3>International Patent Office
<4>WO 03055905 A
<5>
<6>2003
<7>The invention provides BASB231 polypeptides and polynucleotides encoding BASB231 polypeptides
and methods for producing such polypeptides by recombinant techniques.  Also provided are
diagnostic, prophylactic and therapeutic uses.

<>

<1>Castel, A.L., Nakamori, M., Thornton, C.A., Pearson, C.E.
<2>Identification of restriction endonucleases sensitive to 5-cytosine methylation at non-CpG sites, including expanded (CAG)n/(CTG)n repeats.
<3>EPIGENETICS
<4>6
<5>417-421
<6>2011
<7>Most epigenetic studies assess methylation of 5'-CpG-3' sites but recent evidence indicates
that non-CpG cytosine methylation occurs at
high levels in humans and other species. This is most prevalent at
5'-CHG-3', where H = A, C or T, and it preferentially occurs at
5'-CpA-3' and 5'-CpT-3' sites. With the goal of facilitating the
detection of non-CpG methylation, the restriction endonucleases ApeKI,
BbvI, EcoP15I, Fnu4HI, MwoI and TseI were assessed for their
sensitivity to 5-methylcytosine at GpCpA, GpCpT, GpCpC or GpCpG sites,
where methylation is catalyzed by the DNA 5-cytosine 5'-GpC-3'
methyltransferase M.CviPI. We tested a variety of sequences including
various plasmid-based sites, a cloned disease-associated (CAG)83 center
dot(CTG)83 repeat and in vitro synthesized tracts of only (CAG)500
center dot(CTG) 500 or (CAG)800 center dot(CTG)800. The repeat tracts
are enriched for the preferred CpA and CpT motifs. We found that none
of the tested enzymes can cleave their recognition sequences when they
are 5'-GpC-3' methylated. A genomic site known to convert its non-CpG
methylation levels upon C2C12 differentiation was confirmed through the
use of these enzymes. These enzymes can be useful in rapidly and easily
determining the most common non-CpG methylation status in various
sequence contexts, as well as at expansions of (CAG) n.(CTG) n repeat
tracts associated with diseases like myotonic dystrophy and Huntington
disease.

<>

<1>Castelan-Vega, J.A., Jimenez-Alberto, A., Ribas-Aparicio, R.M.
<2>Homology modeling and molecular dynamics simulations of HgiDII methyltransferase in complex with DNA and S-adenosyl-methionine: Catalytic mechanism and interactions with DNA.
<3>J. Mol. Model.
<4>16
<5>1213-1222
<6>2010
<7>M.HgiDII is a methyltransferase (MTase) from Herpetosiphon giganteus that recognizes the
sequence GTCGAC. This enzyme belongs to a group of
MTases that share a high degree of amino acid similarity, albeit none
of them has been thoroughly characterized. To study the catalytic
mechanism of M.HgiDII and its interactions with DNA, we performed
molecular dynamics simulations with a homology model of M.HgiDII
complexed with DNA and S-adenosyl-methionine. Our results indicate that
M.HgiDII may not rely only on Glu119 to activate the cytosine ring,
which is an early step in the catalysis of cytosine methylation;
apparently, Arg160 and Arg162 may also participate in the activation by
interacting with cytosine O2. Another residue from the catalytic site,
Val118, also played a relevant role in the catalysis of M.HgiDII.
Val118 interacted with the target cytosine and kept water molecules
from accessing the region of the catalytic pocket where Cys79 interacts
with cytosine, thus preventing water-mediated disruption of
interactions in the catalytic site. Specific recognition of DNA was
mediated mainly by amino acids of the target recognition domain,
although some amino acids (loop 80-88) of the catalytic domain may also
contribute to DNA recognition. These interactions involved direct
contacts between M.HgiDII and DNA, as well as indirect contacts through
water bridges. Additionally, analysis of sequence alignments with
closely related MTases helped us to identify a motif in the TRD of
M.HgiDII that may be relevant to specific DNA recognition.

<>

<1>Castellano, S., Kuck, D., Sala, M., Novellino, E., Lyko, F., Sbardella, G.
<2>Constrained analogues of procaine as novel small molecule inhibitors of DNA methyltransferase-1.
<3>J. Med. Chem.
<4>51
<5>2321-2325
<6>2008
<7>Constrained analogues of procaine were synthesized, and their inhibiting activity against
DNMT1 was tested. Among them, the most
potent compound, derivative 3b, was also able to induce a recognizable
demethylation of chromosomal satellite repeats in HL60 human myeloid
leukemia cells and thus represents a lead compound for the development
of a novel class of non-nucleoside DNMT1 inhibitors.

<>

<1>Castillo, A., Peterson, S.
<2>The role of GATC flanking sequences in the methylation efficiency of Escherichia coli DNA adenine methyltransferase.
<3>FASEB J.
<4>23
<5>482.3
<6>2009
<7>Escherichia coli DNA adenine methyltransferase (Dam) methylates the adenine in GATC sequences.
The enzyme plays a crucial role in the
timing of DNA replication, gene regulation, and mismatch repair. Many
genes that Dam regulates are involved in pathogenesis. Previous in
vitro studies showed that certain GATC sites were preferentially
methylated depending upon the GATC flanking sequences. Sites rich in
A/T base pairs tend to be less preferred than those rich in G/C. Poorly
preferred GATC sites as determined by in vitro studies are also
undermethylated in vivo. The lack of methylation at these sites is
often important for the expression of pathogenic genes. Our goal is to
determine if GATC flanking sequences affect Dam methylation efficiency
at certain sites in vivo. We hypothesize that the same trend will hold
true in vivo and G/C rich flanking sequences will be preferred.
dam-minus cells were transformed with a plasmid containing the dam gene
under control of the arabinose promoter. After growth at low
concentrations of Dam we isolated the plasmid and observed the
methylation levels of GATC sites. We found that a GATC with G/C rich
flanking sequences was preferentially methylated compared to a
consecutive GATC with A/T rich flanking sequences. Our studies will
help in understanding how Dam is able to discriminate between GATC
sites in vivo which is important for its various roles, particularly
gene regulation.

<>

<1>Castro, W.O., Torres-Ballesteros, A.M., Nakayama, C.R., Melo, I.S., Pellizari, V.H., Silva, A., Ramos, R.T.
<2>Draft Genome Sequence of Haloferax sp. Strain ATB1, Isolated from a Semi-Arid Region in the Brazilian Caatinga.
<3>Genome Announcements
<4>2
<5>e00812-14
<6>2014
<7>Organisms in the Haloferax genus are extreme halophiles that grow in environments with pH
values between 4 and 12, and temperatures between 0 degrees C and 60
degrees C. In the present study, a draft of the first Haloferax sp. strain ATB1
genome isolated from the region of Cariri (in Paraiba State, Brazil) is
presented.

<>

<1>Castronovo, M., Radovic, S., Grunwald, C., Casalis, L., Morgante, M., Scoles, G.
<2>Control of Steric Hindrance on Restriction Enzyme Reactions with Surface-Bound DNA Nanostructures.
<3>Nano Lett.
<4>8
<5>4140-4145
<6>2008
<7>To understand better enzyme/DNA interactions and to design innovative detectors based on DNA
nanoarrays, we need to study the effect of
nanometric confinement on the biochemical activity of the DNA
molecules. We focus on the study of the restriction enzyme reactions
(Dpnll within DNA nanostructures on flat gold films by atomic force
microscopy (AFM). Typically we work with a few patches of DNA self
assembled monolayers (SAMs) that are hundred nm in size and are
lithographically fabricated within alkylthiol SAMs by AFM nanografting.
We start by nanografting a few patches of a single-stranded DNA (ssDNA)
molecule of 44 base pairs (bps) with a 4 bps recognition sequence
(specific for Dpnll in the middle. Afterwards, reaction-ready DNA
nanopatches are obtained by hybridization with a complementary 44bps
ssDNA sequence. The enzymatic reactions were carried out over
nanopatches with different density. By carrying out AFM height
measurements, we are able to show that the capability of the Dpnll
enzyme to reach and react at the recognition site is easily varied by
controlling the DNA packing in the nanostructures. We have found strong
evidence that inside our ordered DNA nanostructures the enzyme (that
works as a dimer) can operate down to the limit in which the space
between adjacent DNA molecules is equal to the size of the DNA/enzyme
complex. Similar experiments were carried out with a DNA sequence
without the recognition site, clearly finding that in that case the
enzymatic reaction did not lead to digestion of the molecules. These
findings suggest that it is possible to tune the efficiency of an
enzymatic reaction on a surface by controlling the steric hindrance
inside the DNA nanopatches without vary any further physical or
chemical variable. These findings are opening the door to novel
applications in both the fields of biosensing and fundamental
biophysics.

<>

<1>Catania, J., Keenan, B.C., Margison, G.P., Fairweather, D.S.
<2>Determination of 5-methylcytosine by acid hydrolysis of DNA with hydrofluoric acid.
<3>Anal. Biochem.
<4>167
<5>347-351
<6>1987
<7>Quantitation of 5-methylcytosine in DNA after acid hydrolysis has been
inaccurate because deamination of cytosine and 5-methylcytosine occurs during
the hydrolysis procedure.  There is little information in the literature
regarding the use of hydrofluoric acid (HF) for DNA hydrolysis and we have
therefore undertaken a systematic study of this process.  The
deoxyribonucleotides of cytosine and 5-methylcytosine were shown not to undergo
detectable levels of deaminatiton during prolonged periods (up to 24 h) at 80C
in 48% HF.  Kinetic studies show that the release of purine and pyrimidine
bases was complete by 4 h under these conditons.  Analysis of the
5-methylcytosine content of DNA from various tissues gave levels that were very
close to the values reported in the literature.  This method is ideally suited
for the determination of the overall cytosine methylation levels in DNA.

<>

<1>Catcheside, D.E.A.
<2>A restriction and modification model for the initiation and control of recombination in Neurospora.
<3>Genet. Res.
<4>47
<5>157-165
<6>1986
<7>It is hypothesized that the products of Neurospora rec+ genes mask
recombinators such as cog by modifying DNA and that unmodified recombinators
act as recognition sites for an endonuclease with scission properties like
those of the type I restriction enzymes found in E. coli.  These cut the DNA in
both strands at some variable distance from a recognition site.  Repair of a
two strand gap initiated in this way would require DNA synthesis using the
information contained in the homologous DNA duplex, leading to gene conversion.
Crossing over could follow from resolution of two Holliday structures formed
during gap repair.  The hypothesis explains the polarity in the frequency of
conversion events across genetic loci, the observation that chromosomes
carrying recombinators are more often converted than is the homologue, and how
recombinators can initiate conversion at a distance, as suggested by the
pattern of conversion events in the his-3 locus in crosses heterozygous for the
translocation TM429.

<>

<1>Catterall, J.F., Lees, N.D., Welker, N.E.
<2>Restriction and modification in Thermophilic bacilli.
<3>Microbiology-1976, American Society for Microbiology, Schlessinger, D., Washington
<4>0
<5>358-366
<6>1976
<7>Over the past several years we have been engaged in an effort to develop transformation in B.
stearothermophilus.  Until recently these studies have met with little success.  During this
time, however, a transfection system was established.  This system requires a 'helper phage'
which may facilitate adsorption and/or uptake of the bacteriophage DNA from the environment.
While studying the uptake and expression of infectious phage DNA, we observed that the
efficiency of transfection varied as much as 1,000-fold depending upon the strain in which the
phage were propagated.  These results suggested the presence of a restriction and modification
system.  This hypothesis was verified when we identified, by efficiency of plating (EOP)
studies, classical restriction and modification systems in these strains.  In this
communication, we describe preliminary studies on the restriction and modification systems in
B. stearothermophilus.

<>

<1>Catterall, J.F., Welker, N.E.
<2>Purification and Properties of the Bst1503 Endonuclease.
<3>Methods Enzymol.
<4>65
<5>167-170
<6>1980
<7>Site-specific endonucleases have become invaluable to the study of the
structure and function of DNA from both prokaryotic and eukaryotic organisms.
Some enzymes of this class, the restriction endonucleases, have been shown to
have a specific in vivo  function acting to protect bacterial cells from
infection.  An associated enzyme, a modification methylase, must be present in
cells producing a restriction enzyme to protect cellular DNA from digestion.
It has recently been shown that restriction enzymes may be capable of promoting
site-specific recombination in vivo.  The obligate thermophile Bacillus
stearothermophilus  exhibited typical host-specific restriction and
modification during infection by thermophilic bacteriophages.  A restriction
endonuclease (endo R.Bst15035) was purified from B. stearothermophilus strain
1503-4R.6  The purified Bst1503 retains its thermostability and is optimally
active at temperatures near the optimal growth temperature of the organism.
The enzyme is stable to long incubation at 65C in the absence of substrate and
forms limit digests after similar incubations in the presence of DNA.  Bst1503
is also stable to long term storage at 5C.

<>

<1>Catterall, J.F., Welker, N.E.
<2>Reduced efficiency of transfection in Bacillus stearothermophilus with modified phage DNA.
<3>Mol. Genetics Bulletin
<4>37
<5>9-10
<6>1974
<7>The infection of B. stearothermophilus, strain 4S with the thermophilic phage
TP-IC or TP-84 DNA requires the presence of helper phage TP-12.  The mechanism
by which the helper phage initiates transfection is unknown.

<>

<1>Catterall, J.F., Welker, N.E.
<2>Isolation and properties of a thermostable restriction endonuclease (Endo R.Bst1503) .
<3>J. Bacteriol.
<4>129
<5>1110-1120
<6>1977
<7>A restriction endonuclease was isolated from Bacillus stearothermophilus
1503-4R (Bst1503) and purified to homogeneity.  The enzyme required Mg2+ ion as
a cofactor.  Bst1503 exhibited maximal activity between pH 7.5 and 8.0 between
60 and 65C, and with about 0.2mM Mg2+.  Bst1503 was not inactivated after
exposure at 55 or 65C for up to 10h.  After 2h of incubation at 70C.  Bst1503
was inactivated by 65%.  Bst1503 was rapidly inactivated at 75C.  A single
protein-staining band having a molecular weight of 46,000 was obseved when
Bst1503 was analyzed by sodium dodecyl sufate-polyacrylamide gel
electrophoresis.  The enzyme was found to exist in two active forms, the
predominating form with an S value of 8.3 (180,000) and the second form with an
S value of 5.4 (96,000).  No conversion between the 8.3S and 5.4S forms was
observed after storage.  Bst1503 recognized six sites in TP-1C deoxyribonucleic
acid (DNA), one site in pSC101 and simian virus 40 DNAs, and three sites in
kvir DNA.  Bst1503 and BamHI were determined to be isoschizomers.  The effect
of temperatures on the activity and stability of BamHI was determined.

<>

<1>Catto, L.E., Bellamy, S.R., Retter, S.E., Halford, S.E.
<2>Dynamics and consequences of DNA looping by the FokI restriction endonuclease.
<3>Nucleic Acids Res.
<4>36
<5>2073-2081
<6>2008
<7>Genetic events often require proteins to be activated by interacting with two DNA sites,
trapping the intervening DNA in a loop. While much is known about looping equilibria, only a
few studies have examined DNA-looping dynamics experimentally. The restriction enzymes that
cut DNA after interacting with two recognition sites, such as FokI, can be used to exemplify
looping reactions. The reaction pathway for FokI on a supercoiled DNA with two sites was
dissected by fast kinetics to reveal, in turn: the initial binding of a protein monomer to
each site; the protein-protein association to form the dimer, trapping the loop; the
subsequent phosphodiester hydrolysis step. The DNA motion that juxtaposes the sites ought on
the basis of Brownian dynamics to take approximately 2 ms, but loop capture by FokI took 230
ms. Hence, DNA looping by FokI is rate limited by protein association rather than DNA
dynamics. The FokI endonuclease also illustrated activation by looping: it cut looped DNA 400
times faster than unlooped DNA.

<>

<1>Catto, L.E., Ganguly, S., Milsom, S.E., Welsh, A.J., Halford, S.E.
<2>Protein assembly and DNA looping by the FokI restriction endonuclease.
<3>Nucleic Acids Res.
<4>34
<5>1711
<6>2006
<7>The FokI restriction endonuclease recognizes an asymmetric DNA sequence and cuts both strands
at fixed positions upstream of the site. The sequence is contacted by a single monomer of the
protein, but the monomer has only one catalytic centre and forms a dimer to cut both strands.
FokI is also known to cleave DNA with two copies of its site more rapidly than DNA with one
copy. To discover how FokI acts at a single site and how it acts at two sites, its reactions
were examined on a series of plasmids with either one recognition site or with two sites
separated by varied distances, sometimes in the presence of a DNA-binding defective mutant of
FokI. These experiments showed that, to cleave DNA with one site, the monomer bound to that
site associates via a weak protein-protein interaction with a second monomer that remains
detached from the recognition sequence. Nevertheless, the second monomer catalyses
phosphodiester bond hydrolysis at the same rate as the DNA-bound monomer. On DNA with two
sites, two monomers of FokI interact strongly, as a result of being tethered to the same
molecule of DNA, and sequester the intervening DNA in a loop.

<>

<1>Caulfield, T., Medina-Franco, J.L.
<2>Molecular dynamics simulations of human DNA methyltransferase 3B with selective inhibitor nanaomycin A.
<3>J. Struct. Biol.
<4>176
<5>185-191
<6>2011
<7>DNA methyltransferases (DNMTs) are involved in epigenetic regulation of the genome and are
promising targets for therapeutic intervention in
cancer and other diseases. Until now, very limited information is
available concerning the molecular dynamics of DNMTs. The natural
product nanaomycin A is the first selective inhibitor of DNMT3B that
induce genomic demethylation. Herein we report long (>100 ns) molecular
dynamics simulations for human DNMT3B bound to nanaomycin A with and
without the presence of the cofactor S-adenosyl-L-methionine (SAM). We
concluded that SAM favors the binding of nanaomycin A to DNMT3B. Key
interactions of nanaomycin A with DNMT3B involve long lasting
interactions with Arg731, Arg733, Arg832, and the catalytic Cys651.
Results further support the previous hypothesis that nanaomycin A has
key interactions with amino acid residues involved in the mechanism of
methylation. This work represents one of the first molecular dynamics
studies of DNMT3B. Results of this work shed light on the structure and
binding recognition process of a key epigenetic enzyme with a small
molecule inhibitor.

<>

<1>Cavalca, L., Corsini, A., Andreoni, V., Muyzer, G.
<2>Draft Genome Sequence of the Arsenite-Oxidizing Strain Aliihoeflea sp. 2WW, Isolated from Arsenic-Contaminated Groundwater.
<3>Genome Announcements
<4>1
<5>e01072-13
<6>2013
<7>Here, we report the draft genome sequence of the arsenite-oxidizing bacterium Aliihoeflea sp.
strain 2WW, which consists of a 4.15-Mb chromosome and contains
different genes that are involved in arsenic transformations.

<>

<1>Cayley, S., Record, M.T.
<2>Anions affect the in vitro function of endonuclease EcoRI.
<3>J. Cell Biochem. Suppl.
<4>11C
<5>204
<6>1987
<7>Environmental variables dictate the stability and function of protein-DNA
interactions in vitro.  For instance, increasing the in vitro K+ concentration
drastically affects both the rate and extent of the formation of EcoRI-DNA
complexes (1).  Here we show the critical importance of anionic solution
components in determining the functional activity of EcoRI.  K+ and glutamate
are the predominant intracellular ionic species in E. coli and the in vivo
concentrations of these ions can accumulate to greater than 0.9 and 0.25 M,
respectively, as part of the osmotic adaptability of the organism.  Comparison
of the extent of DNA cutting by EcoRI as a function of KCl or KGlu
concentration in vitro indicates the rate of cleavage is not only highly
sensitive to the K+ concentration but also to the types and concentrations of
anions present.  Interestingly, substitution of glu- increases the rate of DNA
dramatically in solutions of high K+ content.  Since Cl- is not a primary anion
in E. coli, the results in KGlu are more likely to reflect the physiological
function of the enzyme.  Our studies indicate that anions drastically affect
the stability and function of protein-DNA interactions.  Kinetic studies
therefore provide insight into the determinants of complex formation not
obtainable from structural studies alone.

<>

<1>Cazalet, C., Gomez-Valero, L., Rusniok, C., Lomma, M., Dervins-Ravault, D., Newton, H.J., Sansom, F.M., Jarraud, S., Zidane, N., Ma, L., Bouchier, C., Etienne, J., Hartland, E.L., Buchrieser, C.
<2>Analysis of the Legionella longbeachae genome and transcriptome uncovers unique strategies to cause Legionnaires' disease.
<3>PLoS Genet.
<4>6
<5>e1000851
<6>2010
<7>Legionella pneumophila and L. longbeachae are two species of a large genus of bacteria that
are ubiquitous in nature. L. pneumophila is mainly found
in natural and artificial water circuits while L. longbeachae is mainly
present in soil. Under the appropriate conditions both species are human
pathogens, capable of causing a severe form of pneumonia termed
Legionnaires' disease. Here we report the sequencing and analysis of four
L. longbeachae genomes, one complete genome sequence of L. longbeachae
strain NSW150 serogroup (Sg) 1, and three draft genome sequences another
belonging to Sg1 and two to Sg2. The genome organization and gene content
of the four L. longbeachae genomes are highly conserved, indicating strong
pressure for niche adaptation. Analysis and comparison of L. longbeachae
strain NSW150 with L. pneumophila revealed common but also unexpected
features specific to this pathogen. The interaction with host cells shows
distinct features from L. pneumophila, as L. longbeachae possesses a
unique repertoire of putative Dot/Icm type IV secretion system substrates,
eukaryotic-like and eukaryotic domain proteins, and encodes additional
secretion systems. However, analysis of the ability of a dotA mutant of L.
longbeachae NSW150 to replicate in the Acanthamoeba castellanii and in a
mouse lung infection model showed that the Dot/Icm type IV secretion
system is also essential for the virulence of L. longbeachae. In contrast
to L. pneumophila, L. longbeachae does not encode flagella, thereby
providing a possible explanation for differences in mouse susceptibility
to infection between the two pathogens. Furthermore, transcriptome
analysis revealed that L. longbeachae has a less pronounced biphasic life
cycle as compared to L. pneumophila, and genome analysis and electron
microscopy suggested that L. longbeachae is encapsulated. These
species-specific differences may account for the different environmental
niches and disease epidemiology of these two Legionella species.

<>

<1>Cazalet, C., Rusniok, C., Bruggemann, H., Zidane, N., Magnier, A., Ma, L., Tichit, M., Jarraud, S., Bouchier, C., Vandenesch, F., Kunst, F., Etienne, J., Glaser, P., Buchrieser, C.
<2>Evidence in the Legionella pneumophila genome for exploitation of host cell functions and high genome plasticity.
<3>Nat. Genet.
<4>36
<5>1165-1173
<6>2004
<7>Legionella pneumophila, the causative agent of Legionnaires' disease, replicates as an
intracellular parasite of amoebae and persists in the
environment as a free-living microbe. Here we have analyzed the complete
genome sequences of L. pneumophila Paris (3,503,610 bp, 3,077 genes), an
endemic strain that is predominant in France, and Lens (3,345,687 bp,
2,932 genes), an epidemic strain responsible for a major outbreak of
disease in France. The L. pneumophila genomes show marked plasticity, with
three different plasmids and with about 13% of the sequence differing
between the two strains. Only strain Paris contains a type V secretion
system, and its Lvh type IV secretion system is encoded by a 36-kb region
that is either carried on a multicopy plasmid or integrated into the
chromosome. Genetic mobility may enhance the versatility of L.
pneumophila. Numerous genes encode eukaryotic-like proteins or motifs that
are predicted to modulate host cell functions to the pathogen's advantage.
The genome thus reflects the history and lifestyle of L. pneumophila, a
human pathogen of macrophages that coevolved with fresh-water amoebae.

<>

<1>Ceccaldi, A., Rajavelu, A., Champion, C., Rampon, C., Jurkowska, R., Jankevicius, G., Senamaud-Beaufort, C., Ponger, L., Gagey, N., Ali, H.D., Tost, J., Vriz, S., Ros, S., Dauzonne, D., Jeltsch, A., Guianvarc'h, D., Arimondo, P.B.
<2>C5-DNA Methyltransferase Inhibitors: From Screening to Effects on Zebrafish Embryo Development.
<3>Chembiochem
<4>12
<5>1337-1345
<6>2011
<7>DNA methylation is involved in the regulation of gene expression and plays an important role
in normal developmental processes and diseases,
such as cancer. DNA methyltransferases are the enzymes responsible for
DNA methylation on the position 5 of cytidine in a CpG context. In
order to identify and characterize novel inhibitors of these enzymes,
we developed a fluorescence-based throughput screening by using a short
DNA duplex immobilized on 96-well plates. We have screened 114 flavones
and flavanones for the inhibition of the murine catalytic Dnmt3a/3L
complex and found 36 hits with IC50 values in the lower micromolar and
high nanomolar ranges. The assay, together with inhibition tests on two
other methyltransferases, structure-activity relationships and docking
studies, gave insights on the mechanism of inhibition. Finally, two
derivatives effected zebrafish embryo development, and induced a global
demethylation of the genome, at doses lower than the control drug,
5-azacytidine.

<>

<1>Ceccaldi, A., Rajavelu, A., Ragozin, S., Senamaud-Beaufort, C., Bashtrykov, P., Testa, N., Dali-Ali, H., Maulay-Bailly, C., Amand, S., Guianvarc'h, D., Jeltsch, A., Arimondo, P.B.
<2>Identification of Novel Inhibitors of DNA Methylation by Screening of a Chemical Library.
<3>ACS Chem. Biol.
<4>8
<5>543-548
<6>2013
<7>In order to discover new inhibitors of the DNA methyltransferase 3A/3L complex, we used a
medium-throughput nonradioactive screen on a random
collection of 1120 small organic compounds. After a primary hit
detection against DNA methylation activity of the murine Dnmt3A/3L
catalytic complex, we further evaluated the EC50 of the 12 most potent
hits as well as their cytotoxicity on DU145 prostate cancer cultured
cells. Interestingly, most of the inhibitors showed low micromolar
activities and little cytotoxicity. Dichlone, a small halogenated
naphthoquinone, classically used as pesticide and fungicide, showed the
lowest EC50 at 460 nM. We briefly assessed the selectivity of a subset
of our new inhibitors against hDNMT1 and bacterial Dnmts, including M.
SssI and EcoDam, and the protein lysine methyltransferase PKMT G9a and
the mode of inhibition. Globally, the tested molecules showed a clear
preference for the DNA methyltransferases, but poor selectivity among
them. Two molecules including Dichlone efficiently reactivated YFP gene
expression in a stable HEK293 cell line by promoter demethylation.
Their efficacy was comparable to the DNMT inhibitor of reference
5-azacytidine.

<>

<1>Cecchini, D., Laville, E., Laguerre, S., Robe, P., Leclerc, M., Dore, J., Henrissat, B., Remaud-Simeon, M., Monsan, P., Potocki-Veronese, G.
<2>Functional metagenomics reveals novel pathways of prebiotic breakdown by human gut bacteria.
<3>PLoS ONE
<4>8
<5>e72766
<6>2013
<7>The human intestine hosts a complex bacterial community that plays a major role in nutrition
and in maintaining human health. A functional metagenomic approach was used to explore the
prebiotic breakdown potential of human gut bacteria, including non-cultivated ones. Two
metagenomic libraries, constructed from ileum mucosa and fecal microbiota, were screened for
hydrolytic activities on the prebiotic carbohydrates inulin, fructo-oligosaccharides,
xylo-oligosaccharides, galacto-oligosaccharides and lactulose. The DNA inserts of 17 clones,
selected from the 167 hits that were identified, were pyrosequenced in-depth, yielding in
total 407, 420 bp of metagenomic DNA. From these sequences, we discovered novel prebiotic
degradation pathways containing carbohydrate transporters and hydrolysing enzymes, for which
we provided the first experimental proof of function. Twenty of these proteins are encoded by
genes that are also present in the gut metagenome of at least 100 subjects, whatever are their
ages or their geographical origin. The sequence taxonomic assignment indicated that still
unknown bacteria, for which neither culture conditions nor genome sequence are available,
possess the enzymatic machinery to hydrolyse the prebiotic carbohydrates tested. The results
expand the vision on how prebiotics are metabolized along the intestine, and open new
perspectives for the design of functional foods.

<>

<1>Cech, D., Pein, C.-D.
<2>The influence of modifications on the cleavage of oligonucleotide duplexes by EcoRII and MvaI endonucleases.
<3>Nucleosides and Nucleotides
<4>7
<5>585-588
<6>1988
<7>To study the interaction of the restriction endonucleases MvaI and EcoRII with DNA we have
synthesized some modified oligonucleotides. The results of hydrolysis demonstrate that both
enzymes cleave their substrate by different mechanisms.

<>

<1>Cedar, H., Solage, A., Glaser, G., Razin, A.
<2>Direct detection of methylated cytosine in DNA by use of the restriction enzyme MspI.
<3>Nucleic Acids Res.
<4>6
<5>2125-2132
<6>1979
<7>The extent of methylation of the internal C in the sequence CCGG in DNA from
various eukaryotic sources has been determined using the restriction enzyme
MspI known to be specific for this sequence.  The methylation of the CCGG
sequence is reflected in the restriction pattern obtained by DNA treated with
MspI and its isoschizomer HpaII and analyzed by gel electrophoresis.  A direct
method for detection of 5-methylcytosine in the sequence CCGG has been deviced.
DNA fragments obtained with MspI were radioactively labeled at their 5' ends
and subsequently degraded to the corresponding 5'-deoxyribonucleoside
monophosphates.  5 methylcytidylic acid has been found in most of the 5' ends
of MspI fragments of calf thymus DNA (about 90%) indicating heavy methylation
of the sequence CCGG in calf thymus DNA.  The results also reveal a symmetric
methylation of both strands at this sequence in calf thymus DNA.  In contrast,
the CCGG sequence in other eukaryotic DNAs from organisms like Neurospora,
Drosophila and Herpes virus proved to be undermethylated at this sequence.

<>

<1>Cedar, H., Verdine, G.L.
<2>The amazing demethylase.
<3>Nature
<4>397
<5>568-569
<6>1999
<7>DNA methylation is important for mediating repression of gene expression during mammalian
development.  But although much is known about how methyl groups are added to DNA, little has
been done to work out how they are taken off.  In an exciting article on page 579 of this
issue, Bhattacharya et al. report that they have cloned a hitherto unknown gene which codes
for a protein with a remarkable property - it can catalyse demethylation by directly removing
methyl groups from 5-methyl-cytosine residues in DNA.

<>

<1>Cerdeira, L.T. et al.
<2>Whole-Genome Sequence of Corynebacterium pseudotuberculosis PAT10 Strain Isolated from Sheep in Patagonia, Argentina.
<3>J. Bacteriol.
<4>193
<5>6420-6421
<6>2011
<7>In this work, we report the complete genome sequence of a Corynebacterium pseudotuberculosis
PAT10 isolate, collected from a lung abscess in an
Argentine sheep in Patagonia, whose pathogen also required an
investigation of its pathogenesis. Thus, the analysis of the genome
sequence offers a means to better understanding of the molecular and
genetic basis of virulence of this bacterium.

<>

<1>Cerdeno-Tarraga, A.M. et al.
<2>The complete genome sequence and analysis of Corynebacterium diphtheriae NCTC13129.
<3>Nucleic Acids Res.
<4>31
<5>6516-6523
<6>2003
<7>Corynebacterium diphtheriae is a Gram-positive, non-spore forming, non-motile, pleomorphic rod
belonging to the genus Corynebacterium and the
actinomycete group of organisms. The organism produces a potent
bacteriophage-encoded protein exotoxin, diphtheria toxin (DT), which
causes the symptoms of diphtheria. This potentially fatal infectious
disease is controlled in many developed countries by an effective
immunisation programme. However, the disease has made a dramatic return in
recent years, in particular within the Eastern European region. The
largest, and still on-going, outbreak since the advent of mass
immunisation started within Russia and the newly independent states of the
former Soviet Union in the 1990s. We have sequenced the genome of a UK
clinical isolate (biotype gravis strain NCTC13129), representative of the
clone responsible for this outbreak. The genome consists of a single
circular chromosome of 2 488 635 bp, with no plasmids. It provides
evidence that recent acquisition of pathogenicity factors goes beyond the
toxin itself, and includes iron-uptake systems, adhesins and fimbrial
proteins. This is in contrast to Corynebacterium's nearest sequenced
pathogenic relative, Mycobacterium tuberculosis, where there is little
evidence of recent horizontal DNA acquisition. The genome itself shows an
unusually extreme large-scale compositional bias, being noticeably higher
in G+C near the origin than at the terminus.

<>

<1>Cerdeno-Tarraga, A.M. et al.
<2>Extensive DNA inversions in the B. fragilis genome control variable gene expression.
<3>Science
<4>307
<5>1463-1465
<6>2005
<7>The obligately anaerobic bacterium Bacteroides fragilis, an opportunistic pathogen and
inhabitant of the normal human colonic microbiota, exhibits
considerable within-strain phase and antigenic variation of surface
components. The complete genome sequence has revealed an unusual breadth
(in number and in effect) of DNA inversion events that potentially control
expression of many different components, including surface and secreted
components, regulatory molecules, and restriction-modification proteins.
Invertible promoters of two different types (12 group 1 and 11 group 2)
were identified. One group has inversion crossover (fix) sites similar to
the hix sites of Salmonella typhimurium. There are also four independent
intergenic shufflons that potentially alter the expression and function of
varied genes. The composition of the 10 different polysaccharide
biosynthesis gene clusters identified (7 with associated invertible
promoters) suggests a mechanism of synthesis similar to the O-antigen
capsules of Escherichia coli.

<>

<1>Cermak, T., Doyle, E.L., Christian, M., Wang, L., Zhang, Y., Schmidt, C., Baller, J.A., Somia, N.V., Bogdanove, A.J., Voytas, D.F.
<2>Efficient design and assembly of custom TALEN and other TAL effector-based constructs for DNA targeting.
<3>Nucleic Acids Res.
<4>39
<5>E82
<6>2011
<7>TALENs are important new tools for genome engineering. Fusions of transcription
activator-like (TAL) effectors of plant pathogenic Xanthomonas spp. to the FokI
nuclease, TALENs bind and cleave DNA in pairs. Binding specificity is determined
by customizable arrays of polymorphic amino acid repeats in the TAL effectors. We
present a method and reagents for efficiently assembling TALEN constructs with
custom repeat arrays. We also describe design guidelines based on naturally
occurring TAL effectors and their binding sites. Using software that applies
these guidelines, in nine genes from plants, animals and protists, we found
candidate cleavage sites on average every 35 bp. Each of 15 sites selected from
this set was cleaved in a yeast-based assay with TALEN pairs constructed with our
reagents. We used two of the TALEN pairs to mutate HPRT1 in human cells and ADH1
in Arabidopsis thaliana protoplasts. Our reagents include a plasmid construct for
making custom TAL effectors and one for TAL effector fusions to additional
proteins of interest. Using the former, we constructed de novo a functional
analog of AvrHah1 of Xanthomonas gardneri. The complete plasmid set is available
through the non-profit repository AddGene and a web-based version of our software
is freely accessible online.

<>

<1>Cerquetti, M.C., Brawer, R., Gherardi, M.M., Sordelli, D.O.
<2>Map position of the dam gene in Salmonella typhimurium.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>96
<5>491
<6>1996
<7>Escherichia coli and Salmonella typhimurium DNA is methylated in the adenine
residue in the sequence 5'-GATC-3'.  The DNA adenine methylase (dam) gene of E. coli is
located
at 74 min in a multicistronic operon containing aroK and aroB among other genes.  The S.
typhimurium dam gene has not been mapped yet, although a gene order similar to that of E. coli
(cys-G-dam-aroB) has been suggested.  The recently isolated dam::Tn10dTc mutant of S.
typhimurium enabled us to locate the dam gene more precisely.  Transductional crosses were
carried out using bacteriophage P22 lysates.  Double mutants of S. typhimurium DM6
[csG::Tn5(Kan)-aroB] and DM53 (aroB-dam::Tn10dTc) were constructed in order to perform
three-factor crosses.  Strain DM6 was transduced with a lysate grown on the dam::Tn10dTC
mutant and a total of 129 tetracycline resistant (TetR) transductants were analyzed.  More
than half
(51%) of the TetR transductants maintained the cys- aro- phenotype, 42% were cys+ aro+, 5%
were cys+ aro- and 2% were cys+ aro+.  DM53 was transduced with phage propagated on S.
typhimurium ompR::Tn5.  Kanamycin resistant transductants were screened for sensitivity to 2
aminopurine (2AP) and aro+ phenotype.  Eighty-three percent of the 72 KanR transductants were
aro- and resistant to 2AP, 13% were aro- and sensitive to 2AP, and 3% were sensitive to 2 AP
and
aro-.  Our data support the order: cysG-aroB-dam-ompR on the S. typhimurium genetic map.

<>

<1>Cerquetti, M.C., Brawer, R., Gherardi, M.M., Sordelli, D.O.
<2>Location of aroB and dam genes in the Salmonella typhimurium chromosome.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>97
<5>318
<6>1997
<7>Enterobacteria possess a functional dam methylase.  The dam gene of Escherichia coli in min 74
is part of an operon that includes at least 3 other genes, aroK, aroB and urf74.3, and the
distance between aroB and dam is less than 2 Kbp.  The dam gene of Salmonella typhimurium has
been positioned on the chromosomal map in homology to E. coli.  We have recently isolated an
insertional dam::Tn10dTc mutant of S. typhimurium.  Cotransduction frequencies of aroB and
dam::Tn10tDc found in S. typhimurium (40%) do not agree with a map distance of 2 Kbp between
these two genes, oligonucleotides matching E. coli aroB (ECaroB), S. typhimurium aroB
(STaroB), E. coli dam (Ecdam) and S. typhimurium dam (Stdam) sequences were synthesized and
tested as opposing PCR primers in the amplification of S. typhimurium and E. coli DNA.  ECaroB
and Ecdam oligonucleotide primers produced a band of the expected size (1.6 Kbp) in the
amplification of E. coli DNA.  A band of similar size was obtained using ECaroB and STdam as
primers in the amplification of E. coli DNA.  No specific PCR product was yielded when STaroB
and STdam were tested as primers in the amplification of S. typhimurium DNA.  These results
support the hypothesis that there are differences between E. coli and S. typhimurium in this
region of the chromosome.

<>

<1>Cerritelli, S., Springhorn, S.S., Lacks, S.A.
<2>DpnA, a methylase for single-strand DNA in the DpnII restriction system, and its biological function.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>86
<5>9223-9227
<6>1989
<7>The two DNA-adenine methylases encoded by the DpnII restriction gene cassette
were purified, and their activities were compared on various DNA substrates.
DpnA was able to methylate single-strand DNA and double-strand DNA, whereas
DpnM methylated only double-strand DNA.  Although both enzymes act at
5'-GATC-3' in DNA, DpnA can also methylate sequences altered in the guanine
position, but at a lower rate.  A deletion mutation in the dpnA gene was
constructed and transferred to the chromosome.  Transmission by way of the
transformation pathway of methylated and unmethylated plasmids to dpnA mutant
and wild-type recipients was examined.  The mutant cells restricted
unmethylated donor plasmid establishment much more strongly than did wild-type
cells.  In the wild type, the single strands of donor plasmid DNA that enter by
the transformation pathway are apparently methylated by DpnA prior to
conversion of the plasmid to a double-strand form, in which the plasmid would
be susceptible to the DpnII endonuclease.  The biological function of DpnA may,
therefore, be the enhancement of plasmid transfer to DpnII-containing strains
of Streptococcus pneumoniae.

<>

<1>Cerritelli, S., White, S.W., Lacks, S.A.
<2>Crystallization of the DpnM methylase from the DpnII restriction system of Streptococcus pneumoniae.
<3>J. Mol. Biol.
<4>207
<5>841-842
<6>1989
<7>Three proteins, two DNA methylases and an endonuclease, from the DpnII
restriction system of Streptococcus pneumoniae recognize the DNA sequence 5'
GATC 3' but have very different amino acid sequences, which make them
interesting subjects for structural determination.  A purification procedure
was developed that conveniently yields milligram amounts of the DpnM methylase.
The DpnM protein tends to precipitate at reduced ionic strength, and this
property was exploited to yield well-formed bipyramidal crystals.  By X-ray
diffraction, the crystals of DpnM were found to be orthorhombic, with cell
dimensions a=56.9 angstrom, b=68.2 angstrom, c=84.5 angstrom; systematic
absences identify the space group as P2/1 2/1 2/1.  Diffraction extends beyond
3 angstrom, so the crystals may allow structural determination at atomic
resolution.

<>

<1>Cerritelli, S., White, S.W., Springhorn, S.S., Lacks, S.A.
<2>Two DNA methylases in the DpnII restriction system of Streptococcus pneumoniae.
<3>FASEB J.
<4>4
<5>A2295
<6>1990
<7>The DpnII cassette contains three genes, dpnM, dpnA and dpnB, that encode,
respectively, two DNA methylases, DpnM and DpnA, and the DpnII endonuclease.
The two DNA-adenine methyltransferases were purified and characterized.  DpnM
methylates only double-strand DNA at 5'-GATC-3'.  It is the major DNA
methylating activity in S. pneumoniae and is homologous to the Dam methylase of
Escherichia coli.  We have obtained crystals of DpnM methylase suitable for
diffraction analysis.  At present we are in the process of determining the
crystal structure using the multiwavelength anomalous dispersion procedure.
DpnA, the other methylase in the DpnII restriction system, methylates
single-strand and double-strand DNA, although it is more active in methylating
DNA in single-strand form.  A deletion mutant in the dpnA gene was constructed
and introduced into the chromosome.  The mutant cells have reduced ability to
establish unmethylated donor plasmid, as compared to wild-type cells.  The role
of DpnA may be to methylate the single strands of donor plasmid DNA that enter
the cell by the transformation pathway, thereby protecting the subsequent
reconstructed plasmid from DpnII endonuclease cleavage.  The biological
function of DpnA, therefore, may be the enhancement of plasmid transfer to
DpnII-containing strains of S. pneumoniae.

<>

<1>Cervoni, N., Bhattacharya, S., Szyf, M.
<2>DNA demethylase is a processive enzyme.
<3>J. Biol. Chem.
<4>274
<5>8363-8366
<6>1999
<7>DNA methylation patterns are generated during development by a sequence of methylation and
demethylation events.  We have recently demonstrated that mammals bear a bona fide demethylase
enzyme that removes methyl groups from methylated cytosines.  A general genome wide
demethylation occurs early in development and in differentiating cell lines.  This manuscript
tests the hypothesis that the demethylase enzyme is a processive enzyme.  Using bisulfite
mapping, this report demonstrates that demethylase is a processive enzyme and that the
rate-limiting step in demethylation is the initiation of demethylation.  Initiation of
demethylation is determined by the properties of the sequence.  Once initiated, demethylation
progresses processively.  We suggest that these data provide a molecular explanation for
global hypomethylation.

<>

<1>Cervoni, N., Szyf, M.
<2>Demethylase activity is directed by histone acetylation.
<3>J. Biol. Chem.
<4>276
<5>40778-40787
<6>2001
<7>Mammalian genomes are compartmentalized into dense inactive chromatin that is hypermethylated
and active open chromatin that is hypomethylated. It is generally accepted that this bimodal
pattern of methylation is established during development and is then faithfully inherited
through subsequent cell divisions by a maintenance DNA methyltransferase (DNMT1). The pattern
of methylation is believed to direct local histone acetylation states. In contrast to this
well accepted consensus, we show here using a transient transfection model that an active
demethylase is involved in shaping patterns of methylation in somatic cells.  Demethylase
activity is directed by the state of histone acetylation, and therefore, the resulting
methylation pattern is determined by local histone acetylation states contrary to the accepted
model. Our data support a new model suggesting that the pattern of methylation is maintained
by a dynamic balance of methylation and demethylation activities and the local state of
histone acetylation. This provides a simple mechanism for explaining why active genes are not
methylated.

<>

<1>Cesnaviciene, E., Mitkaite, G., Stankevicius, K., Janulaitis, A., Lubys, A.
<2>Esp1396I restriction-modification system: Structural organization and mode of regulation.
<3>Nucleic Acids Res.
<4>31
<5>743-749
<6>2003
<7>Esp1396I restriction-modification (RM) system recognizes an interrupted palindromic DNA
sequence 5'-CCA(N)(5)TGG-3'. The Esp1396I RM system was
found to reside on pEsp1396, a 5.6 kb plasmid naturally occurring in
Enterobacter sp. strain RFL1396. The nucleotide sequence of the entire
5622 bp pEsp1396 plasmid was determined on both strands. Identified genes
for DNA methyltransferase (esp1396IM) and restriction endonuclease
(esp1396IR) are transcribed convergently. The restriction endonuclease
gene is preceded by the small ORF (esp1396IC) that possesses a strong
helix-turn-helix motif and resembles regulatory proteins found in PvuII,
BamHI and few other RM systems. Gene regulation studies revealed that
C.Esp1396I acts as both a repressor of methylase expression and an
activator of regulatory protein and restriction endonuclease expression.
Our data indicate that C protein from Esp1396I RM system activates the
expression of the Enase gene, which is co-transcribed from the promoter of
regulatory gene, by the mechanism of coupled translation.

<>

<1>Cesnaviciene, E.E., Petrusyte, M.M., Kazlauskiene, R.R., Maneliene, Z., Timinskas, A., Lubys, A., Janulaitis, A.
<2>Characterization of AloI, a Restriction-modification System of a New Type.
<3>J. Mol. Biol.
<4>314
<5>205-216
<6>2001
<7>We report the properties of the new AloI restriction and modification enzyme from
Acinetobacter lwoffi Ks 4-8 that recognizes the DNA target 5' GGA(N)(6)GTTC3' (complementary
strand 5' GAAC(N)(6)TCC3'), and the nucleotide sequence of the gene encoding this enzyme.
AloI is a bifunctional large polypeptide (deduced M(r) 143 kDa) revealing both DNA
endonuclease and methyltransferase activities. Depending on reaction cofactors, AloI cleaves
double-stranded DNA on both strands, seven bases on the 5' side, and 12-13 bases on the 3'
side of its recognition sequence, and modifies adenine residues in both DNA strands in the
target sequence yielding N6-methyladenine. For cleavage activity AloI maintains an absolute
requirement for Mg(2+) and does not depend on or is stimulated by either ATP or
S-adenosyl-l-methionine. Modification function requires the presence of
S-adenosyl-l-methionine and is stimulated by metal ions (Ca(2+)). The C-terminal and central
parts of the protein were found to be homologous to certain specificity (HsdS) and
modification (HsdM) subunits of type I R-M systems, respectively. The N-terminal part of the
protein possesses the putative endonucleolytic motif DX(n)EXK of restriction endonucleases.
The deduced amino acid sequence of AloI shares significant homology with polypeptides encoding
HaeIV and CjeI restriction-modification proteins at the N-terminal and central, but not at the
C-terminal domains. The organization of AloI implies that its evolution involved fusion of an
endonuclease and the two subunits, HsdM and HsdS, of type I restriction enzymes. According to
the structure and function properties AloI may be regarded as one more representative of a
newly emerging group of HaeIV-like restriction endonucleases. Discovery of these enzymes opens
new opportunities for constructing restriction endonucleases with a new specificity.

<>

<1>Chaboki, K.M.
<2>Characterization of a restriction endonuclease isolated from Bacillus subtilis F2.
<3>Biophys. J.
<4>80
<5>282A
<6>2001
<7>The usefulness of restriction systems with their high substrate specificities for ds-DNA, and
the processing of the free or packaged DNA during uptake into Bacillus subtilis cells made us
study the restriction enzyme activity of one of our laboratory isolate, Bacillus subtilis F2
which has been used for the propagation of the newly isolated PAK phage. During the life cycle
of PAK phage in Bacillus subtilis F2 host, it has been noticed that DNA of the phage gets
fragmented in vivo, therefore, it becomes important to explore the endonuclease activity in
this strain.  A cell bound enzyme was isolated from Bacillus subtilis F2, and was
characterized in a crude extract of the bacterial cell pellet.  The enzyme was shown to have
endonuclease activity on bacteriophage lambda DNA and other DNA as well.  Interestingly,
lambda DNA seems to have very few cut sites.  Another important characteristic of this enzyme
is it has an optimum pH shifted toward alkaline range, which is 8.5, among 182 known
restriction enzymes.  The optimum temperature was found to be 40 C.  It seems to work at high
salt concentration.  Lysates extracted at pH 6 and 6.5 have shown endonuclease activity with a
new profile of lambda DNA fragmentation of lysate extracted at pH 7.5.  Hence BsuF2 is a type
II restriction enzyme it seems to have high potential to be used as a genetic tool in
molecular biology.

<>

<1>Chae, Y.G., Hwang, D.K., Cho, J.Y.
<2>Purification and expression of XorII endonuclease in Xanthomonas oryzae pv. Oryzae, Xoo.
<3>Korean Patent Office
<4>KR 1020070098344
<5>
<6>2007
<7>This invention describes how to purify and express XorII endonuclease in Xanthomonas by using
recombinant expression vector.  This XorII can replace PvuI restriction enzyme which has been
widely used and this enzyme will be a low cost for production on a large scale.

<>

<1>Chae, Y.G., Moon, W.J., Cho, J.Y.
<2>Purification and expression of XorI endonuclease in Xanthomonas oryzae pv. Oryzae, Xoo.
<3>Korean Patent Office
<4>KR 1020070098343
<5>
<6>2007
<7>This invention describes how to purify and express XorI endonuclease in Xanthomonas by using
recombinant expression vector.  This XorI can replace PstI restriction enzyme which has been
widely used and this enzyme will be a low cost for production on a large scale.

<>

<1>Chae, Y.K.
<2>Producing restriction enzyme from Escherichia coli comprises transforming Escherichia coli by plasmid, cultivating Escherichia coli, adding culture medium, adding inducer, and cultivating the culture medium.
<3>Korean Patent Office
<4>KR 20100098204
<5>
<6>2010
<7>DERWENT ABSTRACT: NOVELTY - Producing restriction enzyme from Escherichia coli comprises
transforming Escherichia coli by plasmid including gene coding restriction enzyme; cultivating
the Escherichia coli in diluted culture medium comparing to standard concentration for about
12-20 hours; adding culture medium material to the culture medium to make standard
concentration; adding inducer which induces expression of restriction enzyme; and additionally
cultivating the culture medium for about 2-18hours. BIOTECHNOLOGY - Preferred Method: In
producing restriction enzyme, the Escherichia coli is the BL21 (DE3)/pLysS, rosetta
(DE3)/pLysS, rosetta 2 (DE3)/pLysS, arctic Express (DE3), C41 (DE3) or C43 (DE3) phosphorus.
The restriction enzyme is derived from the Xanthomonas oryzae. The plasmid is the phosphorus
of the PET28a, PET, PGEX, PQE, PDEST or the pCOLD series. The plasmid is pET28a/XorKII
phosphorus plasmid. The culture medium comprises Luria-Bertani (LB), Super Optimal Broth
(SOB), SOB including glucose, minimal culture medium, or Terrific Broth culture medium. The
inducer comprises isopropylthiogalactoside (IPTG), lactose or temperature transition
phosphorus. The concentration of IPTG is about 0.1 mM to about 1.0 mM. USE - The method is
useful for producing restriction enzyme (claimed). ADVANTAGE - The method is efficient and
stable. EXAMPLE - No example given.(15 pages)

<>

<1>Chahar, S., Elsawy, H., Ragozin, S., Jeltsch, A.
<2>Changing the DNA Recognition Specificity of the EcoDam DNA-(Adenine-N6)-Methyltransferase by Directed Evolution.
<3>J. Mol. Biol.
<4>395
<5>79-88
<6>2010
<7>EcoDam is an adenine-N6 DNA methyltransferase that methylates the GATC sites in the
Escherichia coli genome. We have changed the target
specificity of EcoDam from GATC to GATT by directed evolution, combining
different random mutagenesis methods with restriction protection at GATT
sites for selection and screening. By co-evolution of an enzyme library
and a substrate library, we identified GATT as the best non-GATC site and
discover a double mutation, R124S/P134S, as the first step to increase
enzyme activity at GATT sites. After four generations of mutagenesis and
selection, we obtained enzyme variants with new specificity for GATT.
While the wild-type EcoDam shows no detectable activity at GATT sites in
E. coli cells, some variants prefer methylation at GATT over GATC sites by
about 10-fold in cells. In vitro DNA methylation kinetics carried out
under single-turnover conditions using a hemimethylated GATC and a GATT
oligonucleotide substrate confirmed that the evolved proteins prefer
methylation of GATT sites to a similar degree. They show up to 1600-fold
change in specificity in vitro and methylate the new GATT target site with
20% of the rate of GATC methylation by the wild-type enzyme, indicating
good activity. We conclude that the new methyltransferases are fully
functional in vivo and in vitro but show a new target-site specificity.

<>

<1>Chai, B., Wang, H., Chen, X.
<2>Draft Genome Sequence of High-Melanin-Yielding Aeromonas media Strain WS.
<3>J. Bacteriol.
<4>194
<5>6693-6694
<6>2012
<7>We sequenced the genome of the high-melanin-yielding Aeromonas media strain WS and then
analyzed genes potentially involved in melanin formation. The 4.2-Mb
draft genome carries multiple genes responsible for pyomelanin synthesis and
other candidate genes identified in our separate study, which have no homolog in
other strains of Aeromonas species.

<>

<1>Chaillet, R., Cirio, C., Navara, C.
<2>The somatic form of the mouse Dnmt1 maintenance methyltransferase is expressed throughout preimplantation development.
<3>Biol. Reprod.
<4>SI
<5>90
<6>2007
<7>The inheritance of gametic epigenetic modifications of DNA is essential to the process of
genomic imprinting, which in turn is essential for normal mammalian development. Patterns of
CpG dinucleotide methylation on imprinted genes appear during gametogenesis due to the action
of de novo cytosine methyltransferase activity. Although the inheritance of these methylation
patterns during post-implantation development is due to the potent maintenance
methyltransferase activity of the somatic form of the Dnmt1 cytosine methyltransferase
(Dnmt1s), the molecular mechanism for maintaining imprinted methylation patterns during
preimplantation development remains largely unknown. The goal of this study was to determine
if Dnmt1s is a possible candidate enzyme for this mechanism by determining if Dnmt1s is
expressed in wild-type preimplantation embryos. For these studies we used the UPT82 rabbit
polyclonal antibody, which recognizes epitopes specific to Dnmt1s protein, but does not detect
the shorter, oocyte-specific Dnmt1o protein.  Improved, sensitive techniques of Dnmt1s
immunodetection with UPT82 were used. On immunoblots of extracts from 100 staged oocytes or
staged embryos, UPT82 detects Dnmt1s in fully-grown oocyte (GV and MII stages), and in embryos
at the 1-cell, 2-cell, 4-cell, 8-cell, morula and blastocyst stages. Using immunostaining and
confocal microscopy, we showed that Dnmt1s is present in the cytoplasm at all stages, and in
the nuclei of all stages except the 1-cell, pronuclear-stage embryos. Dnmt1s also failed to
appear in the maternal and paternal pronuclei even in pronuclear-stage embryos derived from
mouse strains that overexpress Dnmt1s in oocytes, and show greatly increased cytoplasmic
Dnmt1s staining. Because DNA replication is very likely to have been completed at the time of
pronuclear membrane breakdown, these observations indicate that DNA replicated at the first
embryonic cell cycle is probably not exposed to Dnmt1s until well after the replication event.
Using crosses between wild-type mice and mice carrying the Dnmt1V allele, which express only
the Dnmt1o form of Dnmt1, we showed that Dnmt1s protein expressed in 1-cell and 2-cell embryos
is derived from the oocyte, whereas Dnmt1s expressed afterwards is synthesized de novo during
embryogenesis.  These findings indicate that Dnmt1s is expressed in all diploidnuclear stages
of mouse preimplantation embryos, and suggest therefore that Dnmt1s is a strong candidate for
the maintenance methyltransferase activity required for the inheritance of methylation
imprints in the early mouse embryo.

<>

<1>Chain, B.M., Nair, S.P.
<2>Targeting polypeptide.
<3>International Patent Office
<4>WO 2005092918 A
<5>
<6>2005
<7>A targeting polypeptide is provided that may be used to target a chosen antigen to an antigen
presenting cell.  Complexes comprising such targeting polypeptide and antigen, nucleic acids
and vectors encoding them, and cells comprising the nucleic acids and vectors may be used in
methods of immunisation and enhance the immunogenicity of the antigen.

<>

<1>Chain, P., Lamerdin, J., Larimer, F., Regala, W., Lao, V., Land, M., Hauser, L., Hooper, A., Klotz, M., Norton, J., Sayavedra-Soto, L., Arciero, D., Hommes, N., Whittaker, M., Arp, D.
<2>Complete genome sequence of the ammonia-oxidizing bacterium and obligate chemolithoautotroph Nitrosomonas europaea.
<3>J. Bacteriol.
<4>185
<5>2759-2773
<6>2003
<7>Nitrosomonas europaea (ATCC 19718) is a gram-negative obligate chemolithoautotroph that can
derive all its energy and reductant for
growth from the oxidation of ammonia to nitrite. Nitrosomonas europaea
participates in the biogeochemical N cycle in the process of
nitrification. Its genome consists of a single circular chromosome of
2,812,094 bp. The GC skew analysis indicates that the genome is divided
into two unequal replichores. Genes are distributed evenly around the
genome, with approximately 47% transcribed from one strand and
approximately 53% transcribed from the complementary strand. A total of
2,460 protein-encoding genes emerged from the modeling effort, averaging
1,011 bp in length, with intergenic regions averaging 117 bp. Genes
necessary for the catabolism of ammonia, energy and reductant generation,
biosynthesis, and CO(2) and NH(3) assimilation were identified. In
contrast, genes for catabolism of organic compounds are limited. Genes
encoding transporters for inorganic ions were plentiful, whereas genes
encoding transporters for organic molecules were scant. Complex repetitive
elements constitute ca. 5% of the genome. Among these are 85 predicted
insertion sequence elements in eight different families. The strategy of
N. europaea to accumulate Fe from the environment involves several classes
of Fe receptors with more than 20 genes devoted to these receptors.
However, genes for the synthesis of only one siderophore, citrate, were
identified in the genome. This genome has provided new insights into the
growth and metabolism of ammonia-oxidizing bacteria.

<>

<1>Chain, P.S. et al.
<2>Inaugural Article: Burkholderia xenovorans LB400 harbors a multi-replicon, 9.73-Mbp genome shaped for versatility.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>15280-15287
<6>2006
<7>Burkholderia xenovorans LB400 (LB400), a well studied, effective polychlorinated
biphenyl-degrader, has one of the two largest known
bacterial genomes and is the first nonpathogenic Burkholderia isolate
sequenced. From an evolutionary perspective, we find significant
differences in functional specialization between the three replicons of
LB400, as well as a more relaxed selective pressure for genes located on
the two smaller vs. the largest replicon. High genomic plasticity,
diversity, and specialization within the Burkholderia genus are
exemplified by the conservation of only 44% of the genes between LB400 and
Burkholderia cepacia complex strain 383. Even among four B. xenovorans
strains, genome size varies from 7.4 to 9.73 Mbp. The latter is largely
explained by our findings that >20% of the LB400 sequence was recently
acquired by means of lateral gene transfer. Although a range of genetic
factors associated with in vivo survival and intercellular interactions
are present, these genetic factors are likely related to niche breadth
rather than determinants of pathogenicity. The presence of at least eleven
"central aromatic" and twenty "peripheral aromatic" pathways in LB400,
among the highest in any sequenced bacterial genome, supports this
hypothesis. Finally, in addition to the experimentally observed redundancy
in benzoate degradation and formaldehyde oxidation pathways, the fact that
17.6% of proteins have a better LB400 paralog than an ortholog in a
different genome highlights the importance of gene duplication and
repeated acquirement, which, coupled with their divergence, raises
questions regarding the role of paralogs and potential functional
redundancies in large-genome microbes.

<>

<1>Chain, P.S., Comerci, D.J., Tolmasky, M.E., Larimer, F.W., Malfatti, S.A., Vergez, L.M., Aguero, F., Land, M.L., Ugalde, R.A., Garcia, E.
<2>Whole-genome analyses of speciation events in pathogenic Brucellae.
<3>Infect. Immun.
<4>73
<5>8353-8361
<6>2005
<7>Despite their high DNA identity and a proposal to group classical Brucella species as biovars
of Brucella melitensis, the commonly recognized Brucella species can be distinguished by
distinct biochemical and fatty acid characters, as well as by a marked host range (e.g.,
Brucella suis for swine, B. melitensis for sheep and goats, and Brucella abortus for cattle).
Here we present the genome of B. abortus 2308, the virulent prototype biovar 1 strain, and its
comparison to the two other human pathogenic Brucella species and to B. abortus field isolate
9-941. The global distribution of pseudogenes, deletions, and insertions supports previous
indications that B. abortus and B. melitensis share a common ancestor that diverged from B.
suis. With the exception of a dozen genes, the genetic complements of both B. abortus strains
are identical, whereas the three species differ in gene content and pseudogenes. The pattern
of species-specific gene inactivations affecting transcriptional regulators and outer membrane
proteins suggests that these inactivations may play an important role in the establishment of
host specificity and may have been a primary driver of speciation in the genus Brucella.
Despite being nonmotile, the brucellae contain flagellum gene clusters and display
species-specific flagellar gene inactivations, which lead to the putative generation of
different versions of flagellum-derived structures and may contribute to differences in host
specificity and virulence. Metabolic changes such as the lack of complete metabolic pathways
for the synthesis of numerous compounds (e.g., glycogen, biotin, NAD, and choline) are
consistent with adaptation of brucellae to an intracellular life-style.

<>

<1>Chain, P.S., Hu, P., Malfatti, S.A., Radnedge, L., Larimer, F., Vergez, L.M., Worsham, P., Chu, M.C., Andersen, G.L.
<2>Complete Genome Sequence of Yersinia pestis Strains Antiqua and Nepal516: Evidence of Gene Reduction in an Emerging Pathogen.
<3>J. Bacteriol.
<4>188
<5>4453-4463
<6>2006
<7>Yersinia pestis, the causative agent of bubonic and pneumonic plagues, has undergone detailed
study at the molecular level. To further investigate
the genomic diversity among this group and to help characterize lineages
of the plague organism that have no sequenced members, we present here the
genomes of two isolates of the "classical" antiqua biovar, strains Antiqua
and Nepal516. The genomes of Antiqua and Nepal516 are 4.7 Mb and 4.5 Mb
and encode 4,138 and 3,956 open reading frames, respectively. Though both
strains belong to one of the three classical biovars, they represent
separate lineages defined by recent phylogenetic studies. We compare all
five currently sequenced Y. pestis genomes and the corresponding features
in Yersinia pseudotuberculosis. There are strain-specific rearrangements,
insertions, deletions, single nucleotide polymorphisms, and a unique
distribution of insertion sequences. We found 453 single nucleotide
polymorphisms in protein-coding regions, which were used to assess the
evolutionary relationships of these Y. pestis strains. Gene reduction
analysis revealed that the gene deletion processes are under selective
pressure, and many of the inactivations are probably related to the
organism's interaction with its host environment. The results presented
here clearly demonstrate the differences between the two biovar antiqua
lineages and support the notion that grouping Y. pestis strains based
strictly on the classical definition of biovars (predicated upon two
biochemical assays) does not accurately reflect the phylogenetic
relationships within this species. A comparison of four virulent Y. pestis
strains with the human-avirulent strain 91001 provides further insight
into the genetic basis of virulence to humans.

<>

<1>Chain, P.S., Lang, D.M., Comerci, D.J., Malfatti, S.A., Vergez, L.M., Shin, M., Ugalde, R.A., Garcia, E., Tolmasky, M.E.
<2>Genome of Ochrobactrum anthropi ATCC 49188T, a Versatile Opportunistic Pathogen and Symbiont of Several Eukaryotic Hosts.
<3>J. Bacteriol.
<4>193
<5>4274-4275
<6>2011
<7>Ochrobactrum anthropi is a common soil alphaproteobacterium that colonizes a wide spectrum of
organisms and is being increasingly recognized as an
opportunistic human pathogen. Potentially life-threatening infections,
such as endocarditis, are included in the list of reported O. anthropi
infections. These reports, together with the scant number of studies and
the organism's phylogenetic proximity to the highly pathogenic brucellae,
make O. anthropi an attractive model of bacterial pathogenicity. Here we
report the genome sequence of the type strain O. anthropi ATCC 49188,
which revealed the presence of two chromosomes and four plasmids.

<>

<1>Chain, P.S.G. et al.
<2>Insights into the evolution of Yersinia pestis through whole-genome comparison with Yersinia pseudotuberculosis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>13826-13831
<6>2004
<7>Yersinia pestis, the causative agent of plague, is a highly uniform clone that diverged
recently from the enteric pathogen Yersinia pseudotuberculosis. Despite their close genetic
relationship, they differ radically in their pathogenicity and transmission. Here, we report
the complete genomic sequence of Y. pseudotuberculosis IP32953 and its use for detailed genome
comparisons with available Y. pestis sequences. Analyses of identified differences across a
panel of Yersinia isolates from around the world reveal 32 Y. pestis chromosomal genes that,
together with the two Y. pestis-specific plasmids, to our knowledge, represent the only new
genetic material in Y. pestis acquired since the the divergence from Y. pseudotuberculosis. In
contrast, 149 other pseudogenes (doubling the previous estimate) and 317 genes absent from Y.
pestis were detected, indicating that as many as 13% of Y. pseudotuberculosis genes no longer
function in Y. pestis. Extensive insertion sequence-mediated genome rearrangements and
reductive evolution through massive gene loss, resulting in elimination and modification of
preexisting gene expression pathways, appear to be more important than acquisition of genes in
the evolution of Y. pestis. These results provide a sobering example of how a highly virulent
epidemic clone can suddenly emerge from a less virulent, closely related progenitor.

<>

<1>Challacombe, J.F. et al.
<2>Complete genome sequence of Halorhodospira halophila SL1.
<3>Standards in Genomic Sciences
<4>8
<5>206-214
<6>2013
<7>Halorhodospira halophila is among the most halophilic organisms known. It is an obligately
photosynthetic and anaerobic purple sulfur bacterium that exhibits
autotrophic growth up to saturated NaCl concentrations. The type strain H.
halophila SL1 was isolated from a hypersaline lake in Oregon. Here we report the
determination of its entire genome in a single contig. This is the first genome
of a phototrophic extreme halophile. The genome consists of 2,678,452 bp,
encoding 2,493 predicted genes as determined by automated genome annotation. Of
the 2,407 predicted proteins, 1,905 were assigned to a putative function. Future
detailed analysis of this genome promises to yield insights into the halophilic
adaptations of this organism, its ability for photoautotrophic growth under
extreme conditions, and its characteristic sulfur metabolism.

<>

<1>Challacombe, J.F., Duncan, A.J., Brettin, T.S., Bruce, D., Chertkov, O., Detter, J.C., Han, C.S., Misra, M., Richardson, P., Tapia, R., Thayer, N., Xie, G., Inzana, T.J.
<2>Complete Genome Sequence of Haemophilus somnus (Histophilus somni) Strain 129Pt and Comparison to Haemophilus ducreyi 35000HP and Haemophilus  influenzae Rd.
<3>J. Bacteriol.
<4>189
<5>1890-1898
<6>2007
<7>Haemophilus somnus can be either a commensal of bovine mucosal surfaces or an opportunistic
pathogen. Pathogenic strains of H. somnus are a
significant cause of systemic disease in cattle. We report the genome
sequence of H. somnus 129Pt, a nonpathogenic commensal preputial isolate,
and the results of a genome-wide comparative analysis of H. somnus 129Pt,
Haemophilus influenzae Rd, and Haemophilus ducreyi 35000HP. We found
unique genes in H. somnus 129Pt involved in lipooligosaccharide
biosynthesis, carbohydrate uptake and metabolism, cation transport, amino
acid metabolism, ubiquinone and menaquinone biosynthesis, cell surface
adhesion, biosynthesis of cofactors, energy metabolism, and electron
transport. There were also many genes in common among the three organisms.
Our comparative analyses of H. somnus 129Pt, H. influenzae Rd, and H.
ducreyi 35000HP revealed similarities and differences in the numbers and
compositions of genes involved in metabolism, host colonization, and
persistence. These results lay a foundation for research on the host
specificities and niche preferences of these organisms. Future comparisons
between H. somnus 129Pt and virulent strains will aid in the development
of protective strategies and vaccines to protect cattle against H. somnus
disease.

<>

<1>Chambaud, I., Heilig, R., Ferris, S., Barbe, V., Samson, D., Galisson, F., Moszer, I., Dybvig, K., Wroblewski, H., Viari, A., Rocha, E.P.C., Blanchard, A.
<2>The complete genome sequence of the murine respiratory pathogen  Mycoplasma pulmonis.
<3>Nucleic Acids Res.
<4>29
<5>2145-2153
<6>2001
<7>Mycoplasma pulmonis is a wall-less eubacterium belonging to the Mollicutes (trivial name,
mycoplasmas) and responsible for murine respiratory
diseases. The genome of strain UAB CTIP is composed of a single circular 963 879 bp chromosome
with a G + C content of 26.6 mol%, i.e. the lowest
reported among bacteria, Ureaplasma urealyticum apart. This genome contains 782 putative
coding sequences (CDSs) covering 91.4% of its length
and a function could be assigned to 486 CDSs whilst 92 matched the gene sequences of
hypothetical proteins, leaving 204 CDSs without significant
database match. The genome contains a single set of rRNA genes and only 29 tRNAs genes. The
replication origin oriC was localized by sequence
analysis and by using the G + C skew method. Sequence polymorphisms within stretches of
repeated nucleotides generate phase-variable protein
antigens whilst a recombinase gene is likely to catalyse the site-specific DNA inversions in
major M. pulmonis surface antigens. Furthermore, a
hemolysin, secreted nucleases and a glycoprotease are predicted virulence factors.
Surprisingly, several of the genes previously reported to be
essential for a self-replicating minimal cell are missing in the M. pulmonis genome although
this one is larger than the other mycoplasma genomes fully
sequenced until now.

<>

<1>Chames, P., Epinat, J.C., Guillier, S., Patin, A., Lacroix, E., Paques, F.
<2>In vivo selection of engineered homing endonucleases using double-strand break induced homologous recombination.
<3>Nucleic Acids Res.
<4>33
<5>e178
<6>2005
<7>Homing endonucleases, endonucleases capable of recognizing long DNA sequences, have been shown
to be a tool of choice for precise and efficient genome engineering. Consequently, the
possibility to engineer novel endonucleases with tailored specificities is under strong
investigation. In this report, we present a simple and efficient method to select
meganucleases from libraries of variants, based on their cleavage properties. The method has
the advantage of directly selecting for the ability to induce double-strand break induced
homologous recombination in a eukaryotic environment. Model selections demonstrated high
levels of enrichments. Moreover, this method compared favorably with phage display for
enrichment of active mutants from a mutant library. This approach makes possible the
exploration of large sequence spaces and thereby represents a valuable tool for genome
engineering.

<>

<1>Champion, C., Guianvarc'h, D., Senamaud-Beaufort, C., Jurkowska, R.Z., Jeltsch, A., Ponger, L., Arimondo, P.B., Guieysse-Peugeot, A.L.
<2>Mechanistic Insights on the Inhibition of C5 DNA Methyltransferases by Zebularine.
<3>PLoS ONE
<4>5
<5>e12388
<6>2010
<7>In mammals DNA methylation occurs at position 5 of cytosine in a CpG context and regulates
gene expression. It plays an important role in
diseases and inhibitors of DNA methyltransferases (DNMTs)-the enzymes
responsible for DNA methylation-are used in clinics for cancer therapy.
The most potent inhibitors are 5-azacytidine and 5-azadeoxycytidine.
Zebularine (1-(beta-D-ribofuranosyl)-2(1H)-pyrimidinone) is another
cytidine analog described as a potent inhibitor that acts by forming a
covalent complex with DNMT when incorporated into DNA. Here we bring
additional experiments to explain its mechanism of action. First, we
observe an increase in the DNA binding when zebularine is incorporated
into the DNA, compared to deoxycytidine and 5-fluorodeoxycytidine,
together with a strong decrease in the dissociation rate. Second, we
show by denaturing gel analysis that the intermediate covalent complex
between the enzyme and the DNA is reversible, differing thus from
5-fluorodeoxycytidine. Third, no methylation reaction occurs when
zebularine is present in the DNA. We confirm that zebularine exerts its
demethylation activity by stabilizing the binding of DNMTs to DNA,
hindering the methylation and decreasing the dissociation, thereby
trapping the enzyme and preventing turnover even at other sites.

<>

<1>Champion, M.D. et al.
<2>Comparative Genomic Characterization of Francisella tularensis Strains Belonging to Low and High Virulence Subspecies.
<3>PLoS Pathog.
<4>5
<5>e1000459
<6>2009
<7>Tularemia is a geographically widespread, severely debilitating, and occasionally lethal
disease in humans. It is caused by infection by a
gram-negative bacterium, Francisella tularensis. In order to better
understand its potency as an etiological agent as well as its potential
as a biological weapon, we have completed draft assemblies and report
the first complete genomic characterization of five strains belonging
to the following different Francisella subspecies (subsp.): the F.
tularensis subsp. tularensis FSC033, F. tularensis subsp. holarctica
FSC257 and FSC022, and F. tularensis subsp. novicida GA99-3548 and
GA99-3549 strains. Here, we report the sequencing of these strains and
comparative genomic analysis with recently available public Francisella
sequences, including the rare F. tularensis subsp. mediasiatica FSC147
strain isolate from the Central Asian Region. We report evidence for
the occurrence of large-scale rearrangement events in strains of the
holarctica subspecies, supporting previous proposals that further
phylogenetic subdivisions of the Type B clade are likely. We also find
a significant enrichment of disrupted or absent ORFs proximal to
predicted breakpoints in the FSC022 strain, including a genetic
component of the Type I restriction-modification defense system. Many
of the pseudogenes identified are also disrupted in the closely related
rarely human pathogenic F. tularensis subsp. mediasiatica FSC147
strain, including modulator of drug activity B (mdaB) (FTT0961), which
encodes a known NADPH quinone reductase involved in oxidative stress
resistance. We have also identified genes exhibiting sequence
similarity to effectors of the Type III (T3SS) and components of the
Type IV secretion systems (T4SS). One of the genes, msrA2 (FTT1797c),
is disrupted in F. tularensis subsp. mediasiatica and has recently been
shown to mediate bacterial pathogen survival in host organisms. Our
findings suggest that in addition to the duplication of the Francisella
Pathogenicity Island, and acquisition of individual loci, adaptation by
gene loss in the more recently emerged tularensis, holarctica, and
mediasiatica subspecies occurred and was distinct from evolutionary
events that differentiated these subspecies, and the novicida
subspecies, from a common ancestor. Our findings are applicable to
future studies focused on variations in Francisella subspecies
pathogenesis, and of broader interest to studies of genomic
pathoadaptation in bacteria.

<>

<1>Chan, G.F., Gan, H.M., Rashid, N.A.
<2>Genome Sequence of Citrobacter sp. Strain A1, a Dye-Degrading Bacterium.
<3>J. Bacteriol.
<4>194
<5>5485-5486
<6>2012
<7>Citrobacter sp. strain A1, isolated from a sewage oxidation pond, is a facultative aerobe and
mesophilic dye-degrading bacterium. This organism degrades
azo dyes efficiently via azo reduction and desulfonation, followed by the
successive biotransformation of dye intermediates under an aerobic environment.
Here we report the draft genome sequence of Citrobacter sp. A1.

<>

<1>Chan, G.F., Gan, H.M., Rashid, N.A.
<2>Genome Sequence of Enterococcus sp. Strain C1, an Azo Dye Decolorizer.
<3>J. Bacteriol.
<4>194
<5>5716-5717
<6>2012
<7>Enterococcus sp. strain C1 is a facultative anaerobe which was coisolated with Citrobacter sp.
strain A1 from a sewage oxidation pond. Strain C1 could degrade
azo dyes very efficiently via azo reduction and desulfonation in a
microaerophilic environment. Here the draft genome sequence of Enterococcus sp.
C1 is reported.

<>

<1>Chan, H.-Y., Chan, Y.-C., Kam, K.-M., Shaw, P.-C.
<2>Isolation and characterization of restriction endonuclease EclHKI from an Enterobacter cloacae strain.
<3>World J. Microbiol. Biotechnol.
<4>10
<5>30-32
<6>1994
<7>An Enterobacter cloacae strain, producing restriction enzyme EclHKI, was isolated from a
decaying potato. The enzyme is an isoschizomer of Eam1105I, which recognizes and cleaves
GACNNN^NNGTC. EclHKI was produced at high activity (40000 U/g wet cells) and was purified from
contaminants which interfere with restriction digestion by passing the cell lysate through
DEAE-Sephacel and heparin columns. Activity was optimal at 37oC in a medium salt buffer.

<>

<1>Chan, J., Halachev, M., Yates, E., Smith, G., Pallen, M.
<2>Whole-Genome Sequence of the Emerging Pathogen Mycobacterium abscessus Strain 47J26.
<3>J. Bacteriol.
<4>194
<5>549
<6>2012
<7>Mycobacterium abscessus is a rapidly growing environmental mycobacterium commonly found in
soil and water which is often also associated with
infections in humans, particularly of the lung. We report herein the draft
genome sequence of M. abscessus strain 47J26.

<>

<1>Chan, K.G., Tan, K.H., Yin, W.F., Tan, J.Y.
<2>Complete Genome Sequence of Cedecea neteri Strain SSMD04, a Bacterium Isolated from Pickled Mackerel Sashimi.
<3>Genome Announcements
<4>2
<5>e01339-14
<6>2014
<7>We report here the complete genome sequence of C. neteri SSMD04, a strain isolated from
pickled mackerel sashimi, sequenced by third-generation sequencing
technology. To the best of our knowledge, this is the first documentation that
reports the complete genome of Cedecea neteri.

<>

<1>Chan, K.G., Tee, K.K., Yin, W.F., Tan, J.Y.
<2>Complete Genome Sequence of Pluralibacter gergoviae FB2, an N-Acyl Homoserine Lactone-Degrading Strain Isolated from Packed Fish Paste.
<3>Genome Announcements
<4>2
<5>e01276-14
<6>2014
<7>Pluralibacter gergoviae FB2, a bacterial strain isolated from packed food, has been found to
exhibit quorum-quenching properties. Hence, we report the first,
complete genome of P. gergoviae sequenced using the Pacific Biosciences
single-molecule, real-time (SMRT) platform.

<>

<1>Chan, K.G., Wong, C.S., Yin, W.F., Chan, X.Y.
<2>Draft Genome Sequence of Quorum-Sensing and Quorum-Quenching Pseudomonas aeruginosa Strain MW3a.
<3>Genome Announcements
<4>2
<5>e00258-14
<6>2014
<7>Pseudomonas aeruginosa has a broad range of habitation, from aquatic environments to human
lungs. The coexistence of quorum-sensing and quorum-quenching activities occurs in P.
aeruginosa strain MW3a. In this work, we present the draft genome sequence of P. aeruginosa
MW3a, an interesting bacterium isolated from a marine environment.

<>

<1>Chan, K.G., Yin, W.F., Goh, S.Y.
<2>Complete Genome Sequence of Pandoraea pnomenusa 3kgm, a Quorum-Sensing Strain Isolated from a Former Landfill Site.
<3>Genome Announcements
<4>2
<5>e00427-14
<6>2014
<7>Pandoraea pnomenusa strain 3kgm has been identified as a quorum-sensing strain isolated from
soil. Here, we report the complete genome sequence of P. pnomenusa
strain 3kgm by using the Pacific Biosciences single-molecule real-time (PacBio RS
SMRT) sequencer high-resolution technology.

<>

<1>Chan, K.G., Yin, W.F., Lim, Y.L.
<2>Complete Genome Sequence of Pseudomonas aeruginosa Strain YL84, a Quorum-Sensing  Strain Isolated from Compost.
<3>Genome Announcements
<4>2
<5>e00246-14
<6>2014
<7>Here, we report the complete genome sequence of Pseudomonas aeruginosa strain YL84, which was
isolated from compost. This strain was found to be a chitinase-producing quorum-sensing
bacterium.

<>

<1>Chan, Q.W., Cornman, R.S., Birol, I., Liao, N.Y., Chan, S.K., Docking, T.R., Jackman, S.D., Taylor, G.A., Jones, S.J., de Graaf, D.C., Evans, J.D., Foster, L.J.
<2>Updated genome assembly and annotation of Paenibacillus larvae, the agent of American foulbrood disease of honey bees.
<3>BMC Genomics
<4>12
<5>450
<6>2011
<7>BACKGROUND: As scientists continue to pursue various 'omics-based research, there
is a need for high quality data for the most fundamental 'omics of all: genomics.
The bacterium Paenibacillus larvae is the causative agent of the honey bee
disease American foulbrood. If untreated, it can lead to the demise of an entire
hive; the highly social nature of bees also leads to easy disease spread, between
both individuals and colonies. Biologists have studied this organism since the
early 1900s, and a century later, the molecular mechanism of infection remains
elusive. Transcriptomics and proteomics, because of their ability to analyze
multiple genes and proteins in a high-throughput manner, may be very helpful to
its study. However, the power of these methodologies is severely limited without
a complete genome; we undertake to address that deficiency here. RESULTS: We used
the Illumina GAIIx platform and conventional Sanger sequencing to generate a
182-fold sequence coverage of the P. larvae genome, and assembled the data using
ABySS into a total of 388 contigs spanning 4.5 Mbp. Comparative genomics analysis
against fully-sequenced soil bacteria P. JDR2 and P. vortex showed that regions
of poor conservation may contain putative virulence factors. We used GLIMMER to
predict 3568 gene models, and named them based on homology revealed by BLAST
searches; proteases, hemolytic factors, toxins, and antibiotic resistance enzymes
were identified in this way. Finally, mass spectrometry was used to provide
experimental evidence that at least 35% of the genes are expressed at the protein
level. CONCLUSIONS: This update on the genome of P. larvae and annotation
represents an immense advancement from what we had previously known about this
species. We provide here a reliable resource that can be used to elucidate the
mechanism of infection, and by extension, more effective methods to control and
cure this widespread honey bee disease.

<>

<1>Chan, S.-H., Zhu, Z., Van Etten, J.L., Xu, S.-Y.
<2>Cloning of CviPII nicking and modification system from chlorella virus Nys-1 and application of Nt.CviPII in random DNA amplification.
<3>Nucleic Acids Res.
<4>32
<5>6187-6199
<6>2004
<7>The cloning and expression of the CviPII DNA nicking and modification system encoded by
chlorella virus NYs-1 is described. The system consists of a co-linear MTase encoding gene
(cviPIIM) and a nicking endonuclease encoding gene (cviPIINt) separated by 12 nt. M.CviPII
possesses eight conserved amino acid motifs (I to VIII) typical of C5 MTases, but, like
another chlorella virus MTase M.CviJI, lacks conserved motifs IX and X. In addition to
modification of the first cytosine in CCD (D = A, G or T) sequences, M.CviPII modifies both
the first two cytosines in CCAA and CCCG sites as well. Nt.CviPII has significant amino acid
sequence similarity to Type II restriction endonuclease CviJI that recognizes an overlapping
sequence (RG^CY). Nt.CviPII was expressed in Escherichia coli with or without a His-tag in a
host pre-modified by M.CviPII. Recombinant Nt.CviPII recognizes the DNA sequence CCD and
cleaves the phosphodiester bond 5' of the first cytosine while the other strand of DNA at
this site is not affected. Nt.CviPII displays site preferences with CCR (R = A or G) sites
preferred over CCT sites. Nt.CviPII is active from 16 to 65 degrees C with a temperature
optimum of 30-45 degrees C. Nt.CviPII can be used to generate single-stranded DNAs (ssDNAs)
for isothermal strand-displacement amplification. Nt.CviPII was used in combination with Bst
DNA polymerase I large fragment to rapidly amplify anonymous DNA from genomic DNA or from a
single bacterial colony.

<>

<1>Chan, S.H., Bao, Y., Ciszak, E., Laget, S., Xu, S.Y.
<2>Catalytic domain of restriction endonuclease BmrI as a cleavage module for engineering endonucleases with novel substrate specificities.
<3>Nucleic Acids Res.
<4>35
<5>6238-6248
<6>2007
<7>Creating endonucleases with novel sequence specificities provides more possibilities to
manipulate DNA. We have created a chimeric endonuclease
(CH-endonuclease) consisting of the DNA cleavage domain of BmrI
restriction endonuclease and C.BclI, a controller protein of the BclI
restriction-modification system. The purified chimeric endonuclease,
BmrI198-C.BclI, cleaves DNA at specific sites in the vicinity of the
recognition sequence of C.BclI. Double-strand (ds) breaks were observed at
two sites: 8 bp upstream and 18 bp within the C-box sequence. Using DNA
substrates with deletions of C-box sequence, we show that the chimeric
endonuclease requires the 5' half of the C box only for specific cleavage.
A schematic model is proposed for the mode of protein-DNA binding and DNA
cleavage. The present study demonstrates that the BmrI cleavage domain can
be used to create combinatorial endonucleases that cleave DNA at specific
sequences dictated by the DNA-binding partner. The resulting endonucleases
will be useful in vitro and in vivo to create ds breaks at specific sites
and generate deletions.

<>

<1>Chan, S.H., Opitz, L., Higgins, L., O'loane, D., Xu, S.-Y.
<2>Cofactor Requirement of HpyAV Restriction Endonuclease.
<3>PLoS ONE
<4>5
<5>e9071
<6>2010
<7>Background: Helicobacter pylori is the etiologic agent of common gastritis and a risk factor
for gastric cancer. It is also one of the richest sources of Type II restriction-modification
(R-M) systems in microorganisms. Principal/Findings: We have cloned, expressed and purified a
new restriction endonuclease HpyAV from H. pylori strain 26695. We determined the HpyAV DNA
recognition sequence and cleavage site as CCTTC 6/5. In addition, we found that HpyAV has a
unique metal ion requirement: its cleavage activity is higher with transition metal ions than
in Mg++. The special metal ion requirement of HpyAV can be attributed to the presence of a HNH
catalytic site similar to ColE9 nuclease instead of the canonical PD-X-D/EXK catalytic site
found in many other REases. Site-directed mutagenesis was carried out to verify the catalytic
residues of HpyAV. Mutation of the conserved metal-binding Asn311 and His320 to alanine
eliminated cleavage activity. HpyAV variant H295A displayed approximately 1% of wt activity.
Conclusions/Significance: Some HNH-type endonucleases have unique metal ion cofactor
requirement for optimal activities. Homology modeling and site-directed mutagenesis confirmed
that HpyAV is a member of the HNH nuclease family. The identification of catalytic residues in
HpyAV paved the way for further engineering of the metal binding site. A survey of sequenced
microbial genomes uncovered 10 putative R-M systems that show high sequence similarity to the
HpyAV system, suggesting lateral transfer of a prototypic HpyAV-like R-M system among these
microorganisms.

<>

<1>Chan, S.H., Stoddard, B.L., Xu, S.Y.
<2>Natural and engineered nicking endonucleases--from cleavage mechanism to engineering of strand-specificity.
<3>Nucleic Acids Res.
<4>39
<5>1-18
<6>2011
<7>Restriction endonucleases (REases) are highly specific DNA scissors that have facilitated the
development of modern molecular biology. Intensive studies of double strand (ds) cleavage
activity of Type IIP REases, which recognize 4-8 bp palindromic sequences, have revealed a
variety of mechanisms of molecular recognition and catalysis. Less well-studied are REases
which cleave only one of the strands of dsDNA, creating a nick instead of a ds break.
Naturally occurring nicking endonucleases (NEases) range from frequent cutters such as
Nt.CviPII (;CCD; ; denotes the cleavage site) to rare-cutting homing endonucleases (HEases)
such as I-HmuI. In addition to these bona fida NEases, individual subunits of some
heterodimeric Type IIS REases have recently been shown to be natural NEases. The discovery and
characterization of more REases that recognize asymmetric sequences, particularly Types IIS
and IIA REases, has revealed recognition and cleavage mechanisms drastically different from
the canonical Type IIP mechanisms, and has allowed researchers to engineer highly
strand-specific NEases. Monomeric LAGLIDADG HEases use two separate catalytic sites for
cleavage. Exploitation of this characteristic has also resulted in useful nicking HEases. This
review aims at providing an overview of the cleavage mechanisms of Types IIS and IIA REases
and LAGLIDADG HEases, the engineering of their nicking variants, and the applications of
NEases and nicking HEases.

<>

<1>Chan, S.H., Zhu, Z.Y., Dunigan, D.D., Van Etten, J.L., Xu, S.Y.
<2>Cloning of Nt.CviQII nicking endonuclease and its cognate methyltransferase: M.CviQII methylates AG sequences.
<3>Protein Expr. Purif.
<4>49
<5>138-150
<6>2006
<7>Chlorella virus NY-2A has a large, highly methylated dsDNA genome (45% of the cytosines are
5-methylcytosine and 37% of the adenines are
N-6-methyladenine). Here, we report the cloning, expression, and
characterization of the NY-2A-encoded CviQII nicking-modification (N-M)
system. The nicking endonuclease, Nt.CviQII, recognizes R down arrow AG
(R = A or G, down arrow indicating cleavage site) sequences and cleaves
the phosphodiester bond 5' to the adenosine. Because of the difficulty
in cloning and expressing the wild-type Nt.CviQII, C-terminal
truncation mutants were generated and full-length Nt.CviQII was
reconstructed by intein-mediated peptide ligation. The truncation
mutants and the reconstructed full-length Nt.CviQII have the same
recognition and cleavage specificity as the native enzyme. Full-length
and truncated Nt.CviQII produced by a cell-free
transcription/translation system have similar reaction rates. The
methyltransferase, M.CviQII, was also cloned and expressed. It modifies
the adenine in AG doublets of DNA in vitro and in vivo in Escherichia
coli. To our knowledge, M.CviQII is the first adenine methyltransferase
that recognizes a dinucleotide. Therefore, M.CviQII may be a useful
reagent for blocking endonuclease cleavage when restriction sites
overlap with AG sequences.

<>

<1>Chan, S.W.-L., Zilberman, D., Xie, Z., Johansen, L.K., Carrington, J.C., Jacobsen, S.E.
<2>RNA silencing genes control de novo DNA methylation.
<3>Science
<4>303
<5>1336
<6>2004
<7>Cytosine DNA methylation silences harmful DNAs such as transposons and retroviruses.
Maintenance DNA methyltransferases propagate pre-existing DNA methylation in the CG sequence
context by methylating hemi-methylated sites after DNA replication.  Much less is understood
about how invasive DNAs are initially recognized and how de novo DNA methyltransferases of the
DNMT3 family (DRM1 and DRM2 in the plant Arabidopsis thaliana) are directed to unmethylated
loci to initiate gene silencing.

<>

<1>Chan, W.Y., Dietel, K., Lapa, S.V., Avdeeva, L.V., Borriss, R., Reva, O.N.
<2>Draft Genome Sequence of Bacillus atrophaeus UCMB-5137, a Plant Growth-Promoting  Rhizobacterium.
<3>Genome Announcements
<4>1
<5>e00233-13
<6>2013
<7>Bacillus atrophaeus UCMB-5137 shows an extraordinary activity in root colonization and plant
and crop protection. Its draft genome sequence comprises
21 contigs of 4.11 Mb, harboring 4,167 coding sequences (CDS). The genome carries
several genes encoding antimicrobial lipopeptides and polyketides. Multiple
horizontally acquired genes of possible importance for plant colonization were
also found.

<>

<1>Chan, X.Y., Chua, K.H., Puthucheary, S.D., Yin, W.F., Chan, K.G.
<2>Draft Genome Sequence of an Aeromonas sp. Strain 159 Clinical Isolate That Shows  Quorum-Sensing Activity.
<3>J. Bacteriol.
<4>194
<5>6350
<6>2012
<7>Aeromonas is a pathogenic organism that is often found to infect humans. Here we  report the
draft genome of a clinical isolate in Malaysia, Aeromonas sp. strain
159, which shows N-acylhomoserine lactone production. In the draft genome of
strain 159, luxI and luxR homologue genes were found to be located at contig 47,
and these genes are believed to be important for the quorum-sensing system
present in this pathogen.

<>

<1>Chan, Y.-S., Takeuchi, R., Jarjour, J., Huen, D.S., Stoddard, B.L., Russell, S.
<2>The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive.
<3>PLoS ONE
<4>8
<5>e74254
<6>2013
<7>The homing endonuclease gene (HEG) drive system, a promising genetic approach for controlling
arthropod populations, utilises engineered nucleases to spread deleterious mutations that
inactivate individual genes throughout a target population. Previous work with a naturally
occurring LAGLIDADG homing endonuclease (I-SceI) demonstrated its feasibility in both
Drosophila and Anopheles. Here we report on the next stage of this strategy: the redesign of
HEGs with customized specificity in order to drive

<>

<1>Chandrasegaran, S.
<2>Functional domains in Flavobacterium okeanokoites (FokI) restriction endonuclease.
<3>International Patent Office
<4>WO 9418313
<5>
<6>1994
<7>The present inventors have identified the recognition and cleavage domains of the FokI
restriction endonuclease. Accordingly, the present invention relates to the DNA segments
encoding the recognition and cleavage domains of the FokI restriction endonuclease,
respectively. The 41 kDa N-terminal fragment constitutes the FokI recognition domain while the
25 kDa C-terminal fragment constitutes the FokI cleavage nuclease domain. The present
invention also relates to hybrid restriction enzymes comprising the nuclease domain of the
FokI restriction endonuclease linked to a recognition domain of another enzyme. Additionally,
the present invention relates to the construction of two insertion mutants of FokI
endonuclease.

<>

<1>Chandrasegaran, S.
<2>Functional domains in Flavobacterium okeanokoites (FokI) restriction endonuclease.
<3>US Patent Office
<4>US 5356802
<5>
<6>1994
<7>The present inventor has identified the recognition and cleavage domains of the FokI
restriction endonuclease. Accordingly, the present invention relates to DNA segments encoding
the recognition and cleavage domains of the FokI restriction endonuclease, respectively. The
41 kDa N-terminal fragment constitutes the FokI recognition domain while the 25 kDa C-terminal
fragment constitutes the FokI cleavage nuclease domain. The present invention also relates to
hybrid restriction enzymes comprising the nuclease domain of the FokI restriction endonuclease
linked to a recognition domain of another enzyme.

<>

<1>Chandrasegaran, S.
<2>Functional domains in Flavobacterium okeanokoites (FokI) restriction endonuclease.
<3>International Patent Office
<4>WO 9509233
<5>
<6>1995
<7>The present inventors have identified the recognition and cleavage domains of the FokI
restriction endonuclease.  Accordingly, the present invention relates to DNA segments encoding
the recognition and cleavage domains of the FokI restriction endonuclease, respectively.  The
41 kDa N-terminal fragment constitutes the FokI recognition domain while the 25 kDa
C-terminal fragment constitutes the FokI cleavage nuclease domain.  The present invention also
relates to hybrid restriction enzymes comprising the nuclease domain of the FokI restriction
endonuclease linked to a recognition domain of another enzyme.  One such hybrid restriction
enzyme is Ubx-FN.  This enzyme contains the homeo domain of Ubx linked to the cleavage or
nuclease domain of FokI.  Additionally, the present invention relates to the construction of
two insertion mutants of FokI endonuclease.

<>

<1>Chandrasegaran, S.
<2>Insertion and deletion mutants of FokI restriction endonuclease.
<3>US Patent Office
<4>US 5487994
<5>
<6>1996
<7>The present invention reveals the construction of several insertion (4, 8, 12, 18, 19
or 23 amino acid residues) and deletion (4 or 7 amino acid residues) mutants of the linker
region of
FokI endonuclease in Flavobacterium okeanokoites.  The mutant enzymes were purified, and their
cleavage properties were characterized.  The mutants have the same DNA sequence-specificity as
the wild-type enzyme.  However, compared with the wild-type enzyme, the insertion mutants
cleave predominantly one nucleotide further away from the recognition site on both strands of
the
DNA substrate.  The four codon deletion mutant shows relaxed specificity at the cut site while
the
seven codon deletion appears to inactivate the enzyme.  The DNA-binding and cleavage domains
of
FokI appear to be linked by a relatively malleable linker.  No simple linear relationship
exists
between the linker length and the distance of the cut site from the recognition site.
Furthermore,
the four codon insertion mutants cleave DNA substrates containing hemi-methylated FokI sites;
they do not cleave fully-methylated substrates.

<>

<1>Chandrasegaran, S.
<2>General method to clone hybrid restriction endonucleases using LIG gene.
<3>US Patent Office
<4>US 5792640
<5>
<6>1998
<7>The present invention reveals methods for cloning hybrid restriction endonucleases and for
enzymatically inactivating a target DNA.  The method for cloning hybrid restriction
endonucleases involves co-expression of a ligase.  A first plasmid contains a gene encoding a
DNA ligase.  A second plasmid contains a gene encoding a hybrid restriction endonuclease and
is compatible with the first plasmid.  The method involves transfecting host cells with the
first plasmid, so that DNA ligase is produced, followed by transfecting the cells with the
second plasmid.  The method for enzymatically inactivating a target DNA involves preparing a
plasmid, phage, virus or any other delivery vehicle such as a liposome containing a gene
encoding a nuclease, delivering the gene into cells, inducing the cells to produce the
nuclease and enzymatically inactivating the target DNA.

<>

<1>Chandrasegaran, S.
<2>Functional domains in Flavobacterium okeanokoities (FokI) restriction endonuclease.
<3>US Patent Office
<4>US 5436150
<5>
<6>1995
<7>The present inventors have identified the recognition and cleavage domains of the FokI
restriction endonuclease.  Accordingly, the present invention relates to DNA segments encoding
the recognition and cleavage domains of the FokI restriction endonuclease, respectively.  The
41 kDa N-terminal fragment constitutes the FokI recognition domain while the 25 kDa
C-terminal fragment constitutes the FokI cleavage nuclease domain.  The present invention also
relates to hybrid restriction enzymes comprising the nuclease domain of the FokI restriction
endonuclease linked to a recognition domain of another enzyme.  One such hybrid restriction
enzyme is Ubx-FN.  This enzyme contains the homeo domain of Ubx linked to the cleavage or
nuclease domain of FokI.  Additionally, the present invention relates to the construction of
two insertion mutants of FokI endonuclease.

<>

<1>Chandrasegaran, S., Lunnen, K.D., Smith, H.O., Wilson, G.G.
<2>Cloning and sequencing the HinfI restriction and modification genes.
<3>Gene
<4>70
<5>387-392
<6>1988
<7>The HinfI restriction and modification genes were cloned on a 3.9-kb PstI
fragment inserted into the PstI site of plasmid pBR322.  Both genes are
confined to an internal 2.3-kb BclI-AvaI subfragment.  This subfragment was
sequenced.  Two large open reading frames (ORF's) are present.  ORF1 codes for
the methylase [predicted 359 amino acids (aa)] and ORF2 codes for the
endonuclease (predicted 252 or 272 aa).

<>

<1>Chandrasegaran, S., Smith, H.O.
<2>Amino acid sequence homologies among twenty-five restriction endonucleases and methylases.
<3>Structure and Expression. From Proteins to Ribosomes., Adenine Press, Sarma, R.H., Sarma, M.H., New York
<4>1
<5>149-156
<6>1988
<7>The amino acid sequences of 17 DNA methylases were compared. They show extensive homologies
that tend to cluster in distinct groups sharing similar DNA recognition sequences. Altogether
they fall into 5 groups: (I) M.Dam(GATC), M.DpnII(GATC), M.T4(GATC), and M.EcoRV(GATATC) show
extensive homology (28% to 34%). (II) M.HinfI(GANTC) and M.HhaII(GANTC) are 18.8% homologous
in 128 amino acids of their amino-terminal regions. (III) M.TaqI(TCGA), M.CviBIII(TCGA),
M.PaeR7I(CTCGAG) and M.PstI(CTGCAG) show regions of homology ranging from 103 to 128 residues
in length with amino acid sequence identities of 22% to 28%. (IV) M.BspRI(GGCC),
M.BsuRI(GGCC), M.SPR(GGCC, CCGG, CC(A/T)GG), M.EcoRII(CC(A/T)GG), M.Phi3T(GGCC,GCNGC) and
M.HhaI(GCGC) show extensive homologies ranging from 22% to 66% in regions ranging from 25% to
100% of their lengths. (V)M.EcoRI is not homologous to any of the above. All the 11 DNA
adenine methylases contain a conserved DPPY or NPPY sequence within the homologous segments.
This motif is probably involved in adenine recognition and AdoMet binding at the position of
methylation. Three short regions of high homology were identified in the 6 cytosine
methylases, namely the PC, ENVK, and RER homologies. The PC motif is believed to be involved
in the catalytic mechanism of the cytosine methylases. Eight restriction enzyme sequences were
also compared. No clearly significant homologies were observed. The restriction enzymes did
not show homology to the methylases.

<>

<1>Chandrasegaran, S., Smith, H.O., Amzel, M.L., Ysern, X.
<2>Preliminary x-ray diffraction analysis of HhaII endonuclease-DNA cocrystals.
<3>Proteins
<4>1
<5>263-266
<6>1986
<7>HhaII restriction endonuclease purified from an overproducing recombinant E.
coli clone has been cocrystallized with a heptanucleotide duplex,
d-GGAGTCC:GGACTCC.  The cocrystals are monoclonic and belong to the space group
C2.  The unit cell dimensions are a = 199.0+/-1.0 angstrom, b = 100.0+/-0.5
angstrom, c = 80.3+/-0.4 angstrom, and b = 101.0+/-1.0o.  There appear to be
two dimers per asymmetric unit and the crystals diffract to 4-angstrom
resolution.

<>

<1>Chandrasegaran, S., Smith, J.
<2>Chimeric restriction enzymes: What is next?
<3>Biol. Chem.
<4>380
<5>841-848
<6>1999
<7>Chimeric restriction enzymes are a novel class of engineered nucleases in which the
non-specific DNA cleavage domain of Fokl (a type IIS restriction endonuclease) is fused to
other DNA-binding motifs. The latter include the three common eukaryotic DNA-binding motifs,
namely the helix-turn-helix motif, the zinc finger motif and the basic helix-loop-helix
protein containing a leucine zipper motif. Such chimeric nucleases have been shown to make
specific cuts in vitro very close to the expected recognition sequences. The most important
chimeric nucleases are those based on zinc finger DNA-binding proteins because of their
modular structure. Recently, one such chimeric nuclease, Zif-QQR-F(N) was shown to find and
cleave its target in vivo. This was tested by microinjection of DNA substrates and the enzyme
into frog oocytes (Carroll et al., 1999). The injected enzyme made site-specific double-strand
breaks in the targets even after assembly of the DNA into chromatin. In addition, this
cleavage activated the target molecules for efficient homologous recombination. Since the
recognition specificity of zinc fingers can be manipulated experimentally, chimeric nucleases
could be engineered so as to target a specific site within a genome. The availability of such
engineered chimeric restriction enzymes should make it feasible to do genome engineering, also
commonly referred to as gene therapy.

<>

<1>Chandrasegaran, S., Wu, L.P., Valda, E., Smith, H.O.
<2>Overproduction and purification of the M.HhaII methyltransferase from Haemophilus haemolyticus.
<3>Gene
<4>74
<5>15-21
<6>1988
<7>The HhaII methyltransferase gene from Haemophilus haemolyticus was subcloned in
an expression vector under control of the hybrid trp-lac promoter.  Induction
with isopropyl-Beta-D-thiogalactopyranoside results in overproduction of the
methyltransferase to about 3% of total cellular protein.  The methyltransferase
was purified to near electrophoretic homogeneity by phosphocellulose, DEAE, and
gel chromatography.  Its monomer Mr by sodium dodecyl sulfate-polyacrylamide
gel electrophoresis is 25 kDa, in good agreement with that predicted from the
nucleotide sequence.  Crystals of the methyltransferase were obtained in the
presence of a two-fold molar excess of the duplex oligodeoxynucleotide
substrate 5'd-GGACTCC/CCTGAGG.

<>

<1>Chandrasekhar, K., Raman, R.
<2>Restriction enzyme HincII is sensitive to methylation of cytosine that occurs 5' to the recognition sequence.
<3>Nucleic Acids Res.
<4>24
<5>1045-1046
<6>1996
<7>In this paper we demonstrate that the HincII restriction endonuclease, in addition to being
sensitive to methylation of the 3' A and C residues, is also sensitive to methylation of a
cytosine immediately 5' to the recognition sequence.  Having encountered this property in
one of the sites in the mouse c-fos gene, we confirmed the sensitivity of HincII to the 5'
cytosine methylation in in vitro methylated pUC12, pBR322 and pfos-1 plasmids.

<>

<1>Chandrashekaran, S., Babu, P., Nagaraja, V.
<2>Characterization of DNA binding activities of over-expressed KpnI restriction endonuclease and modification methylase.
<3>J. Biosci.
<4>24
<5>269-277
<6>1999
<7>The genes encoding the KpnI restriction endonuclease and methyltransferase from Klebsiella
pneumoniae have been cloned and expressed in Escherichia coli using a two plasmid strategy.
The gene for KpnI methylase with its promoter was cloned and expressed in pACYC184.  Even
though the methylase clone is in a low copy number plasmid pACMK, high level expression of
methylase is achieved.  A hyper-expressing clone of KpnI endonuclease, pETRK was engineered by
cloning the R gene into the T7 expression system.  This strategy resulted in over-expression
of KpnI endonuclease to about 15-30% of cellular protein.  Both the enzymes were purified
using a single chromatographic step to apparent homogeneity.  The yield of purified
endonuclease and methylase from one liter of culture was approximately 30 and 6 mg
respectively.  Electrophoretic mobility shift assays show that both the enzymes are capable of
binding to specific recognition sequence in the absence of any cofactors.  The complexes of
KpnI methyl transferase and endonuclease with their cognate site exhibit distinctive behavior
with respect to ionic requirement.

<>

<1>Chandrashekaran, S., Babu, P., Nagaraja, V.
<2>Purification and characterization of restriction endonuclease BasI from Bacillus species.
<3>J. Biochem. Mol. Biol. Biophys.
<4>3
<5>225-229
<6>1999
<7>After screening a variety of bacteria from soil for the DNA cleavage activity, a type II
restriction endonuclease, BasI was isolated and characterized from a Bacillus species.  Using
the purified enzyme the recognition sequence was determined.  The enzyme functions efficiently
at low ionic strength, narrow pH range and at the optimum temperature of 30oC.

<>

<1>Chandrashekaran, S., Manjunatha, U.H., Nagaraja, V.
<2>KpnI restriction endonuclease and methyltransferase exhibit contrasting mode of sequence recognition.
<3>Nucleic Acids Res.
<4>32
<5>3148-3155
<6>2004
<7>The molecular basis of the interaction of KpnI restriction endonuclease (REase) and the
corresponding methyltransferase (MTase) at their cognate recognition sequence is investigated
using a range of footprinting techniques. DNase I protection analysis with the REase reveals
the protection of a 14-18 bp region encompassing the hexanucleotide recognition sequence. The
MTase, in contrast, protects a larger region. KpnI REase contacts two adjacent guanine
residues and the single adenine residue in both the strands within the recognition sequence
5'-GGTACC-3', inferred by dimethylsulfate (DMS) protection, interference and missing
nucleotide interference analysis. In contrast, KpnI MTase does not show elaborate
base-specific contacts. Ethylation interference analysis also showed the differential
interaction of REase and MTase with phosphate groups of three adjacent bases on both strands
within the recognition sequence. The single thymine residue within the sequence is hyper-
reactive to the permanganate oxidation, consistent with MTase-induced base flipping. The REase
on the other hand does not show any major DNA distortion. The results demonstrate that the
differences in the molecular interaction pattern of the two proteins at the same recognition
sequence reflect the contrasting chemistry of DNA cleavage and methylation catalyzed by these
two dissimilar enzymes, working in combination as constituents of a cellular defense strategy.

<>

<1>Chandrashekaran, S., Saravanan, M., Radha, D.R., Nagaraja, V.
<2>Ca2+-mediated site-specific DNA cleavage and suppression of promiscuous activity of KpnI restriction endonuclease.
<3>J. Biol. Chem.
<4>279
<5>49736-49740
<6>2004
<7>The characteristic feature of type II restriction endonucleases (REases) is their exquisite
sequence specificity and obligate Mg2+
requirement for catalysis. Efficient cleavage of DNA only in the
presence of Ca2+ ions, comparable with that of Mg2+, is previously not
described. Most intriguingly, KpnI REase exhibits Ca2+-dependent
specific DNA cleavage. Moreover, the enzyme is highly promiscuous in
its cleavage pattern on plasmid DNAs in the presence of Mn2+ or Mg2+,
with the complete suppression of promiscuous activity in the presence
of Ca2+. KpnI methyltransferase does not exhibit promiscuous activity
unlike its cognate REase. The REase binds to oligonucleotides
containing canonical and mapped noncanonical sites with comparable
affinities. However, the extent of cleavage is varied depending on the
metal ion and the sequence. The ability of the enzyme to be promiscuous
or specific may reflect an evolutionary design. Based on the results,
we suggest that the enzyme KpnI represents an REase evolving to attain
higher sequence specificity from an ancient nonspecific nuclease.

<>

<1>Chandrashekaran, S., Shankar, A.B., Babu, P., Paul, B.D., Nagaraja, V.
<2>Identification and characterization of a type II restriction endonuclease, StrI from Streptomyces thermodiastaticus.
<3>Curr. Sci.
<4>77
<5>273-276
<6>1999
<7>A new type II restriction endonuclease, StrI has been identified from Streptomyces
thermodiastaticus.  The enzyme has been purified using three column chromatography steps.  The
enzyme recognizes a hexanucleotide sequence and cleaves DNA 5'-C/TCGAG-3' as indicated.  The
optimum temperature, pH, and cation requirements for the enzyme activity were determined.

<>

<1>Chandrashekharan, S., Paul, B.D., Nagaraja, V.
<2>Design of a novel regulatory circuit for expression of restriction endonucleases.
<3>Biol. Chem.
<4>379
<5>579-582
<6>1998
<7>We have developed a new strategy with a very tight control for the expression of cloned genes.
The system employed here is the T7 promoter-based expression system in which transcription
activator protein C of bacteriophage Mu (Mu C) has been cloned to serve as a repressor in the
regulatory circuit.  The system also includes pLysE, which encodes T7 lysozyme, an inhibitor
of T7 RNA polymerase.  This ensures tight regulation of cloned genes in the uninduced state.
Upon induction, the expressed Mu C protein binds to its cognate site thereby repressing lys
transcription driven by the tet promoter.  In order to evaluate the tight control achieved in
the system, and to check leaky expression, if any, we have cloned the gene for the SmaI
restriction endonuclease without its cognate methylase.  For this purpose, a dicistronic unit
was constructed by cloning the smaIR gene downstream of the Mu C gene.  SmaI expression was
observed only in the induced cell extracts, demonstrating a tight control.  The system could
be used to express the genes of other cloned restriction enzymes and has the potential for
general applications.

<>

<1>Chang, D., Zhu, Y., Chen, J., Fang, X., Li, T., Wang, J., Guo, Y., Su, L., Xu, G., Wang, Y., Chen, Z., Liu, C.
<2>Draft Genome Sequences and Annotation of Enterococcus faecium Strain LCT-EF20.
<3>Genome Announcements
<4>1
<5>e00083-12
<6>2013
<7>The space environment is reported to cause biological alterations in microorganisms, such as
growth, drug resistance, and virulence. Here, we present the model of Enterococcus faecium to
investigate the effects of space conditions on the microbe and on the whole-genome sequences
of the strain LCT-EF20 after being exposed to space flight.

<>

<1>Chang, D., Zhu, Y., Fang, X., Li, T., Wang, J., Guo, Y., Su, L., Liu, Y., Jiang, X., Wang, L., Guo, N., Liu, C.
<2>Draft Genome Sequences of the Enterococcus faecium Strain LCT-EF258.
<3>Genome Announcements
<4>1
<5>e00147-12
<6>2013
<7>The space environment has been shown to affect microbes by altering various features,
including morphology, growth rate, metabolism, virulence, drug
resistance, and gene expression and mutation. Here we present the draft genome
sequence of the Enterococcus faecium strain LCT-EF258, derived from the E.
faecium strain CGMCC 1.1736, which was exposed to 17-day space flight.

<>

<1>Chang, D., Zhu, Y., Zou, Y., Fang, X., Li, T., Wang, J., Guo, Y., Su, L., Xia, J., Yang, R., Fang, C., Liu, C.
<2>Draft Genome Sequence of Enterococcus faecium Strain LCT-EF90.
<3>J. Bacteriol.
<4>194
<5>3556-3557
<6>2012
<7>Enterococcus faecium is an opportunistic human pathogen, found widely in the human
gastrointestinal tract, and can also be isolated from a variety of plants,
animals, insects, and other environmental sources. Here, we present the fine
draft genome sequence of E. faecium LCT-EF90.

<>

<1>Chang, H.K., Zylstra, G.J., Chae, J.C.
<2>Genome Sequence of n-Alkane-Degrading Hydrocarboniphaga effusa Strain AP103T (ATCC BAA-332T).
<3>J. Bacteriol.
<4>194
<5>5120
<6>2012
<7>Hydrocarboniphaga effusa strain AP103(T) (ATCC BAA-332(T)) is a member of the
Gammaproteobacteria utilizing n-alkanes as the sole source of carbon and energy.
Here we report the draft genome sequence of AP103(T), which consists of 5,193,926
bp with a G + C content of 65.18%.

<>

<1>Chang, S., Cohen, S.N.
<2>In vivo site-specific genetic recombination promoted by the EcoRI restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>74
<5>4811-4815
<6>1977
<7>Site-specific genetic recombination promoted in vivo by the EcoRI endonuclease
has been demonstrated by using constructed hybrid plasmids in which the
chloramphenicol resistance gene was inactivated by insertion of DNA fragments
at an EcoRI site within the gene.  Such recombination can involve either the
joining of intracellularly generated cohesive termini of the same DNA fragment
or intermolecular ligation of different DNA fragments.  DNA cleavage and
ligation in vivo are precise: recombinant DNA molecules show functional
continuity of the gene sequence cleaved by the enzyme and regeneration of
nucleotide recognition sites for both the EcoRI endonuclease and the EcoRI DNA
methylase.  In other experiments, EcoRI-generated fragments of eukaryotic DNA
that had not been modified by the Escherichia coli K methylase were shown to be
taken up by bacterial cells and to undergo intracellular ligation to segments
of bacterial plasmid DNA.

<>

<1>Chang, Y.J. et al.
<2>Non-contiguous finished genome sequence and contextual data of the filamentous soil bacterium Ktedonobacter racemifer type strain (SOSP1-21).
<3>Standards in Genomic Sciences
<4>5
<5>97-111
<6>2011
<7>Ktedonobacter racemifer corrig. Cavaletti et al. 2007 is the type species of the  genus
Ktedonobacter, which in turn is the type genus of the family
Ktedonobacteraceae, the type family of the order Ktedonobacterales within the
class Ktedonobacteria in the phylum 'Chloroflexi'. Although K. racemifer shares
some morphological features with the actinobacteria, it is of special interest
because it was the first cultivated representative of a deep branching
unclassified lineage of otherwise uncultivated environmental phylotypes
tentatively located within the phylum 'Chloroflexi'. The aerobic, filamentous,
non-motile, spore-forming Gram-positive heterotroph was isolated from soil in
Italy. The 13,661,586 bp long non-contiguous finished genome consists of ten
contigs and is the first reported genome sequence from a member of the class
Ktedonobacteria. With its 11,453 protein-coding and 87 RNA genes, it is the
largest prokaryotic genome reported so far. It comprises a large number of
over-represented COGs, particularly genes associated with transposons, causing
the genetic redundancy within the genome being considerably larger than expected
by chance. This work is a part of the Genomic Encyclopedia of Bacteria and
Archaea project.

<>

<1>Chang, Y.J. et al.
<2>Complete genome sequence of Acidaminococcus fermentans type strain (VR4).
<3>Standards in Genomic Sciences
<4>3
<5>1-14
<6>2010
<7>Acidaminococcus fermentans (Rogosa 1969) is the type species of the genus Acidaminococcus, and
is of phylogenetic interest because of its isolated
placement in a genomically little characterized region of the Firmicutes. A.
fermentans is known for its habitation of the gastrointestinal tract and its
ability to oxidize trans-aconitate. Its anaerobic fermentation of glutamate has
been intensively studied and will now be complemented by the genomic basis. The
strain described in this report is a nonsporulating, nonmotile, Gram-negative
coccus, originally isolated from a pig alimentary tract. Here we describe the
features of this organism, together with the complete genome sequence, and
annotation. This is the first complete genome sequence of a member of the family
Acidaminococcaceae, and the 2,329,769 bp long genome with its 2,101
protein-coding and 81 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Chang, Z., Morgan, R.D.
<2>Method for cloning and producing the PshAI restriction endonuclease.
<3>European Patent Office
<4>EP 0794252 A
<5>
<6>1997
<7>The present invention is directed to a method for cloning and producing the PshAI restriction
endonuclease by (1) introducing the restriction endonuclease gene from Plesiomonas
shigelloides into a host whereby the restriction gene is expressed; (2) fermenting the host
which contains the vector encoding and expressing the PshAI restriction endonuclease activity,
and (3) purifying the PshAI restriction endonuclease from the fermented host which contains
the vector encoding and expressing the PshAI restriction endonuclease activity, and (3)
purifying the PshAI restriction endonuclease from the fermented host which contains the vector
encoding and expressing the PshAI restriction endonuclease activity.

<>

<1>Chang, Z., Morgan, R.D.
<2>Method for cloning and producing the PshAI restriction endonuclease.
<3>US Patent Office
<4>US 5824529
<5>
<6>1998
<7>The present invention is directed to a method for cloning and producing the PshAI restriction
endonuclease by (1) introducing the restriction endonuclease gene from Plesiomonas
shigelloides into a host whereby the restriction gene is expressed; (2) fermenting the host
which contains the vector encoding and expressing the PshAI restriction endonuclease activity,
and (3) purifying the PshAI restriction endonuclease from the fermented host which contains
the vector encoding and expressing the PshAI restriction endonuclease activity.

<>

<1>Chang, Z., Morgan, R.D.
<2>Method for cloning and producing the PmeI restriction endonuclease.
<3>European Patent Office
<4>EP 0931835 A
<5>
<6>1998
<7>The present invention relates to recombinant DNA which encodes the PmeI restriction
endonuclease and modification methylase, and production of PmeI restriction endonuclease from
the recombinant DNA.

<>

<1>Chang, Z., Morgan, R.D.
<2>Method for cloning and producing the PmeI restriction endonuclease.
<3>US Patent Office
<4>US 5945288
<5>
<6>1999
<7>The present invention relates to recombinant DNA which encodes the PmeI restriction
endonuclease and modification methylase, and production of PmeI restriction endonuclease from
the recombinant DNA.

<>

<1>Chanto, G., Occhialini, A., Gras, N., Alm, R.A., Megraud, F., Marais, A.
<2>Identification of strain-specific genes located outside the plasticity zone in nine clinical isolates of Helicobacter pylori.
<3>Microbiology
<4>148
<5>3671-3680
<6>2002
<7>Helicobacter pylori is a Gram-negative bacterium that is associated with
the development of peptic ulcers and gastric carcinoma in humans. This
species appears to be one of the most genetically variable bacteria
described to date. The overall level of heterogeneity within strains of
this organism was determined by comparing the genome sequences of two
reference strains, J99 and 26695. The aim of this study was to measure the
genetic diversity within strains of H. pylori by looking for
strain-specific genes in nine H. pylori strains isolated from patients
suffering from chronic gastritis (n=3), duodenal ulcers (n=3) or gastric
cancer (n=3). Seven loci that contained strain-specific genes in strains
J99 and 26695 were studied. These regions were subsequently amplified from
most of the clinical isolates studied and their sequences were determined.
ORFs were predicted from the sequence data and were compared to sequences
within the databases. The results showed that the genes flanking the ORFs
specific to either strain J99 or strain 26695 were also present in a
similar configuration in the genomes of the nine clinical isolates.
Moreover, in most regions, ORFs homologous to those found in the
corresponding loci in the two reference strains were detected. However, in
10 regions, genes similar to those located at another locus in the genome
of J99 or 26695 were found. Finally, six strain-specific genes were
identified in three regions of three of the H. pylori strains isolated
from patients with duodenal ulcers (n=2) and gastric cancer (n=1). Of
these six genes, five were putative genes and one was an orthologue of a
gene encoding a transposase in Thermotoga maritima. However, no
association with disease was found for these genes.

<>

<1>Chaplin, A.V., Efimov, B.A., Khokhlova, E.V., Kafarskaia, L.I., Tupikin, A.E., Kabilov, M.R., Shkoporov, A.N.
<2>Draft Genome Sequence of Coprobacter fastidiosus NSB1T.
<3>Genome Announcements
<4>2
<5>e00122-14
<6>2014
<7>Coprobacter fastidiosus is a Gram-negative obligate anaerobic bacterium belonging to the
phylum Bacteroidetes. In this work, we report the draft genome sequence of
C. fastidiosus strain NSB1(T) isolated from human infant feces.

<>

<1>Chapman, J.A. et al.
<2>The dynamic genome of Hydra.
<3>Nature
<4>464
<5>592-596
<6>2010
<7>The freshwater cnidarian Hydra was first described in 1702 and has been the object of study
for 300 years. Experimental studies of Hydra between 1736 and 1744 culminated in the discovery
of asexual reproduction of an animal by budding, the first description of regeneration in an
animal, and successful transplantation of tissue between animals. Today, Hydra is an important
model for studies of axial patterning, stem cell biology and regeneration. Here we report the
genome of Hydra magnipapillata and compare it to the genomes of the anthozoan Nematostella
vectensis and other animals. The Hydra genome has been shaped by bursts of transposable
element expansion, horizontal gene transfer, trans-splicing, and simplification of gene
structure and gene content that parallel simplification of the Hydra life cycle. We also
report the sequence of the genome of a novel bacterium stably associated with H.
magnipapillata. Comparisons of the Hydra genome to the genomes of other animals shed light on
the evolution of epithelia, contractile tissues, developmentally regulated transcription
factors, the Spemann-Mangold organizer, pluripotency genes and the neuromuscular junction.

<>

<1>Chardonnens, A., Puzio, P., McKersie, B.D.
<2>Nucleic acid sequences encoding proteins associated with abiotic stress response and plant cells and plants with increased tolerance to environmental stress.
<3>International Patent Office
<4>WO 2007020198 A
<5>
<6>2007
<7>This invention relates generally to nucleic acid sequences encoding proteins that are
associated with abiotic stress responses and abiotic stress tolerance in plants.  This
invention further relates to transformed plant cells with altered metabolic activity compared
to a corresponding non-transformed, wild-type plant cell, wherein the metabolic activity is
altered by transformation with a Stress-Related Protein (SRP) coding nucleic acid and results
in increased tolerance and/or resistance to an environmental stress as compared to a
corresponding non-transformed, wild-type plant cell.

<>

<1>Charette, S.J., Brochu, F., Boyle, B., Filion, G., Tanaka, K.H., Derome, N.
<2>Draft Genome Sequence of the Virulent Strain 01-B526 of the Fish Pathogen Aeromonas salmonicida.
<3>J. Bacteriol.
<4>194
<5>722-723
<6>2012
<7>Aeromonas salmonicida is an important fish pathogen, mainly of salmonids. This bacterium
causes a disease named furunculosis, which is particularly
detrimental for the aquaculture industry. Here, we present the draft
genome sequence of A. salmonicida 01-B526, a strain isolated from a brook
trout that is more virulent than A. salmonicida reference strain A449, for
which a genome sequence is available.

<>

<1>Charlop-Powers, Z., Banik, J.J., Owen, J.G., Craig, J.W., Brady, S.F.
<2>Selective enrichment of environmental DNA libraries for genes encoding nonribosomal peptides and polyketides by phosphopantetheine transferase-dependent complementation of siderophore biosynthesis.
<3>ACS Chem. Biol.
<4>8
<5>138-143
<6>2013
<7>The cloning of DNA directly from environmental samples provides a means to
functionally access biosynthetic gene clusters present in the genomes of the
large fraction of bacteria that remains recalcitrant to growth in the laboratory.
Herein, we demonstrate a method by which complementation of phosphopantetheine
transferase deletion mutants can be used to restore siderophore biosynthesis and
to therefore selectively enrich eDNA libraries for nonribosomal peptide
synthetase (NRPS) and polyketide synthase (PKS) gene sequences to unprecedented
levels. The common use of NRPS/PKS-derived siderophores across bacterial taxa
makes this method generalizable and should allow for the facile selective
enrichment of NRPS/PKS-containing biosynthetic gene clusters from large
environmental DNA libraries using a wide variety of phylogenetically diverse
bacterial hosts.

<>

<1>Chater, K.F.
<2>Actinophage DNA.
<3>Dev. Ind. Microbiol.
<4>21
<5>65-74
<6>1980
<7>Actinophage DNA is considered largely in relation to its possible uses in
genetic engineering.  The G + C contents (determined by density) varied from 55
to 69%.  The no. of target sites for various restriction enzymes bore some
relation to base composition, high G + C content correlating with high
frequencies of SalGI sites and very low frequencies of sites for EcoRI,
HindIII, and HpaI.  Sites for BamHI and SalPI were absent from most actinophage
DNA tested regardless of its base composition.  A transfection procedure was
characterized, in which plaques could be visualised in soft agar overlays to
which had been added actinophage DNA and protoplasts previously mixed in the
presence of polyethylene glycol.  The procedure was effective with DNA of VP5,
UC32, R4, U448, and S14 and protoplasts of Streptomyces coelicolor A3(2) or S.
lividans 66.  	Deletion mutants of several phages (detected by virtue of their
resistance to chelating agents) were obtained both spontaneously and after
transfection of protoplasts with phage DNA pretreated with EcoRI and DNA
ligase.  One deletion mutant of phage R4 lacked the single EcoRI target site,
which is therefore dispensable and available as a DNA cloning site.  The
occurrence of chelating-agent-resistant deletion mutants of some actinophages
suggested that packaging of their DNA involved staggered, site-specific cutting
of concatameric DNA.  This was corroborated by evidence that UC31, SH10, and
probably R4 DNA molecules possess cohesive ends.

<>

<1>Chater, K.F.
<2>Some Recent Developments in Streptomyces Genetics.
<3>Genetics of Industrial Microorganisms, American Society for Microbiology, Sebek, O.K., Laskin, A.I., Washington, D.C.
<4>0
<5>123-133
<6>1979
<7>This review will be concerned with "nonchromosomal" genetics.  The omission of
normal chromosomal genetics does not reflect a decline in its importance, but
rather the absence of major advances since previous reviews, with the important
exception of the development of protoplast fusion, which is dealt with by D.A.
Hopwood elsewhere in this volume.  My approach is to consider what is known
about the occurrence and genetic determination of "nonessential" (and therefore
possibly plasmid-specified) functions in streptomycetes, and then to discuss
aspects of plasmids and temperate phages relevant to the development of
potential "recombinant DNA" systems in streptomycetes.

<>

<1>Chater, K.F.
<2>A site-specific endodeoxyribonuclease from Streptomyces albus CMI 52766 sharing site-specificity with Providencia stuartii endonuclease PstI.
<3>Nucleic Acids Res.
<4>4
<5>1989-1998
<6>1977
<7>A class II site-specific endodeoxyribonuclease (SalPI) was identified in
cell-free extracts of Streptomyces albus CMI 52766 after high speed
centrifugation and fractionation through BioGel A0.5M.  SalPI cleaves lambda
DNA into at least 18 fragments.  Five cleavage sites were located in the linear
lambda map by the use of double and triple restriction enzyme digests involving
EcoRI, HindIII, SalGI and another new Streptomyces enzyme, SacI.  The results
were indistinguishable from those previously obtained for a Providencia
stuartii enzyme, PstI, by Smith, Blattner & Davies (Nucl. Acids Res. 1976 3:
343).  SalPI and PstI were shown by a double digest test to have the same site
specificity.  None of 34 phages tested was obviously restricted by S. albus CMI
52766, and correspondingly DNA from two of them was not cleaved in vitro by
SalPI.  DNA from a Streptomyces phage that does not form plaques on S. albus
CMI 52766, and plasmid SCP2 DNA from S. coelicolor A3(2), were both cleaved.

<>

<1>Chater, K.F.
<2>Streptomyces Phages and Their Applications to Streptomyces Genetics.
<3>The Bacteria, Academic Press, Queener, S.W., Day, L.E., 
<4>9
<5>119-158
<6>1986
<7>Interest in Streptomyces phages was first caused by the occasional infestation
of early antibiotic fermentation cultures.  The economic loss caused by lysed
fermentation cultures was occasionally significant so that even today there is
sometimes opposition to the import of Streptomyces phages into laboratories
attached to production plants.  However, it is now clear that laboratory
varieties of Streptomyces phages can be contained in the laboratory and turned
to good account.  Many industrial laboratories are beginning to exploit them.
This chapter is an account of recent developments in the study of Streptomyces
phages and their use in genetic manipulation.  Its starting point is a fairly
comprehensive review written several years ago (Lomovskaya et al., 1980);
information given in detail there will be only briefly summarized here.  The
most conspicuous advances since that time have been in, or have resulted from,
the use of Streptomyces phages as DNA cloning vectors.  Techniques for
isolating, assaying, and propagating Streptomyces phages from soil and from
lysogens differ only in minor detail (related to the host's mycelial growth
habit) from those used for eubacterial phages, which they resemble closely in
structure and biology (Lomovskaya et al., 1980).  This resemblance is not
surprising, since the gross molecular biology of streptomycetes is similar to
that of other gram-positive bacteria.  Of course, there are interesting
differences between streptomycetes and other bacteria (e.g., in morphology,
antibiotic production, and high G + C content in DNA), and it is mainly as
agents to help our understanding of the genetic basis of these phenomena that
Streptomyces phages have come to be important topics of research.  Streptomyces
phages are not the only tools available for such a purpose.  Some species of
Streptomyces, notably S. coelicolor A3(2), have good chromosomal genetics
(Hopwood, 1967; Hopwood et al., 1973; Hopwood and Chater, 1974; Hopwood and
Merrick, 1977; Rhodes, this volume).  Highly efficient generalized
recombination through protoplast fusion (Hopwood et al., 1977; Baltz, 1978) is
broadly applicable to Streptomyces.  Transformation for chromosomal markers by
liposome-entrapped DNA (Makins and Holt, 1981) has been reported for these
gram-positive bacteria.  Moreover, an almost extravagant range of plasmid DNA
cloning vectors for Streptomyces has come into use (reviewed by Chater et al.,
1982a; Hopwood and Chater, 1982; Chater, 1983; Bib et al., 1983; Hopwood et
al., this volume).  Streptomyces genetics has been the subject of two recent
reviews (Chater and Hopwood, 1984; Hopwood and Chater, 1984).	This chapter will
be almost wholly concerned with temperate phages (reflecting the bias in
published data), and in particular with UC31, which is the most studied
Streptomyces phage.

<>

<1>Chater, K.F.
<2>Restriction in Streptomyces.
<3>Nocardia and Streptomyces. Proceedings of the International Symposium on Nocardia and Streptomyces. Warsaw Oct 4-8 1978, Gustav Fischer Verlag, Mordarski, M., Kurytowicz, W. Jeljaszewicz, J., Stuttgart
<4>0
<5>303-311
<6>1978
<7>Restriction has been the subject of authoritative recent reviews, and I will
give only a short introduction to it before pointing out its interest for the
Streptomyces geneticist.  Many bacteria possess restriction enzymes, which are
endodeoxy-ribonucleases that recognize specific short nucleotide sequences in
double-stranded DNA and cleave both strands.  The cleavage sites may, like the
recognition site, be specific (class II enzymes) or non-specific (class I
enzymes).  The only co-factor required for class II enzymes is Mg++, whereas
class I enzymes usually require ATP, Mg++ and S-adenosyl methionine.
Restriction enzymes are obligatorily accompanied by so-called modifying enzymes
that protect the cell's own DNA from endonucleolytic cleavage, by specific
recognition and modification of the same sites as are recognized by the
restriction enzyme.  Different species or even different wild-type isolates of
the same species usually have restriction-modification (R-M) systems with
different specificities.  Consequently DNA transferred from one species or
strain to another, for example in phage infection or bacterial mating, will
often be unprotected against endonucleolytic breakdown on entering the new host
and may thus be inactivated.  In the case of a phage, a lower efficiency of
plating (e.o.p.) restriction is observed on the second host.  However, incoming
foreign DNA is also subject to host-specific modification, and if this is
achieved before restriction, that DNA molecule is safe from endonucleolytic
cleavage and retains its biological activity.  In the case of phage DNA the
normal infective cycle ensues, detectable by plaque formation.  The recognition
sites of type II enzymes are invariably palindromic, showing 2-fold rotational
symmetry.  Moreover, cleavage been due to restriction or perhaps to
inefficiency of pair formation between the species.  The e.o.p. of the
temperate phage VP5 was therefore tested on the two strains and was shown to be
independent of the previous host used for its propagation.  Either an R-M
system was absent or, if it existed, VP5 was not susceptible to it.  It
therefore seems that an R-M system which has recently been identified in S.
albus G, and which will be described later, is the only one yet identified in
Streptomycetes.

<>

<1>Chater, K.F., Carter, A.T.
<2>A New, Wide Host-range, Temperate Bacteriophage (R4) of Streptomyces and its Interaction with some Restriction-Modification Systems.
<3>J. Gen. Microbiol.
<4>115
<5>431-442
<6>1979
<7>A new temperate phage, R4, of Streptomyces was isolated from soil on a restriction-deficient
mutant of S. albus G.  In its morphology, adsorption properties and growth kinetics R4
resembled other temperate phages of Streptomyces though its requirements for Ca2+ and Mg2+
were higher than usual. It was unable to form plaques above 34.5 C.  R4-mediated transduction
was not detected.  Unlike other Streptomyces temperate phages, R4 had a wide host-range, which
correlated better with the absence of detectable class II restriction enzymes than with
conventional taxonomic divisions.  Many of the sensitive strains [but not, apparently, S.
coelicolor A3(2)] could be lysogenized. With the wild-type R4, plaques were obtained on S.
albus G only after growth on a restriction-deficient, modification-proficient mutant, and then
only at a very low efficiency of plating.  All of these plaques were of a mutant type (R4G)
which (unlike the parental R4 phage) showed conventional patterns of restriction-modification
in the S. albus G (SalGI) and S. albus P (SalPI) systems.  R4G mutants, but not R4, were
sensitive to a restriction-modification system present in two S. rimosus strains (2251 and
NRRL 2234).  DNA from SalGI-unmodified (but not from modified) R4 or R4G was cleaved by SalGI
into more than 30 fragments (mean size 1.35 kilobases; summed molecular weight (30.02x10^6).
R4 DNA was cleaved at one site by EcoRI, at one site by SalPI (0 PstI), and not at all by
HindIII or BamHI.

<>

<1>Chater, K.F., Carter, A.T.
<2>Restriction of a bacteriophage in Streptomyces albus P (CMI 52766) by Endonuclease SalPI.
<3>J. Gen. Microbiol.
<4>109
<5>181-185
<6>1978
<7>Restriction has been hard to show in streptomycetes.  For example, in
Streptomyces coelicolor A3(2) (a strain sensitive to many actinophages),
restriction could be demonstrated only in a hybrid strain (S. coelicolor A3(2)
X S. griseus kr.15) which possessed phage receptors from S. griseus kr.15 and a
restriction system from S. coelicolor A3(2) (Lomovskaya et al., 1977). This
paper reports the restriction and modification by S. albus P of a wide
host-range temperate Streptomyces phage, R4, and it is shown that the agent of
restriction is the endonuclease SalPI.

<>

<1>Chater, K.F., Wilde, L.C.
<2>Restriction of a bacteriophage of Streptomyces albus G involving endonuclease SalI.
<3>J. Bacteriol.
<4>128
<5>644-650
<6>1976
<7>The bacteriophage Pa16, isolated from soil on Streptomyces albus G, was
restricted when transferred from an alternative host back to S. albus G.
Extracted unmodified Pa16 deoxyribonucleic acid was cleaved at a single site by
a cell-free extract of S. albus G.  Fractions cleaving Pa16 deoxyribonucleic
acid contained the endonuclease SalI first described by J. Arrand, P. Myers,
and R.J. Roberts (unpublished data).  A mutant of S. albus G was isolated which
was defective in both restriction and modification of Pa16.  This mutant lacked
SalI activity.  It is concluded that SalI is the agent of restriction of Pa16
by S. albus G.

<>

<1>Chater, K.F., Wilde, L.C.
<2>Streptomyces albus G Mutants Defective in the SalGI Restriction-Modification System.
<3>J. Gen. Microbiol.
<4>116
<5>323-334
<6>1980
<7>Streptomyces albus G mutants (at least 12 of which were independent) defective
in SalGI-mediated restriction (R-) were isolated after mutagenesis.  Some of
them lacked detectable SalGI activity in cell-free extracts.  Some were also
partially or completely defective in SalGI-associated modification (M-).  Loss
of restriction rendered S. albus G sensitive to many phages to which it was
normally totally resistant.  DNA from one such phage had many SalGI target
sites (means, one site per 1.35 kilobases).  A mutant was isolated which was
heat-sensitive for growth, apparently because it was restriction-proficient but
temperature-sensitive for modification.  At a rather high frequency, this
mutant generated spontaneous heat-tolerant derivatives which were nearly all
R-.  Such R- mutants were always M- rather than being temperature-sensitive for
modification.  In a limited genetic analysis, the determinants of restriction
and modification did not recombine with each other, and since there was no
reassortment of these phenotypes among the parental output of crosses it
appeared that the determinants were located close together on the chromosome.

<>

<1>Chatterjee, D., Longo, M., Flynn, E., Oberfelder, R.
<2>Methods for production of proteins.
<3>US Patent Office
<4>US 6703484 B
<5>
<6>2004
<7>The current invention provides methods for producing a polypeptide as inclusion bodies in
bacterial host cells.  The present methods are carried out by forming a gene construct
comprising the genetic sequence encoding a polypeptide operatively linked to that of an
inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such
that host cells comprising the gene construct produce the polypeptide as intracellular
inclusion bodies.  The methods of the present invention facilitate the rapid isolation and
purification of recombinant proteins.  In addition, the present methods may be useful for
producing polypeptides or proteins which are small and are typically difficult to express, as
well as those proteins that are toxic to host cells such as E. coli.  The present invention
also provides plasmids, vectors and host cells to be used in the present invention for
production of polypeptides, and methods of production of polypeptides using these vectors and
host cells.  The invention further provides methods for producing protein molecular weight
ladders for use in protein gel electrophoresis, as well as proteins and protein molecular
weight ladders produced by these methods.

<>

<1>Chatterjee, D., Longo, M., Flynn, E., Oberfelder, R.
<2>Methods for production of proteins.
<3>Japanese Patent Office
<4>JP 2001512306 A
<5>
<6>2001
<7>
<>

<1>Chatterjee, D., Thakur, A.R., Raychaudhuri, S.
<2>Draft Genome Sequence of Ammonia-Producing Acinetobacter sp. Strain MCC2139 from  Dairy Effluent.
<3>Genome Announcements
<4>1
<5>e00410-13
<6>2013
<7>We report the draft genome sequence of an ammonia-producing, esculin-hydrolyzing,
catalase-positive, gram-negative bacterium, Acinetobacter sp. strain MCC2139.
This bacterium, isolated from dairy sludge and with optimum growth at 37 degrees
C, has a genome size of 2,967,280 bp with a G+C content of 42.3%.

<>

<1>Chatterjee, D.K.
<2>Cloning and expressing XhoII restriction endonucleases and M.XhoII modification methylase from Xanthomonas.
<3>US Patent Office
<4>US 5304480
<5>
<6>1994
<7>The present invention is directed to recombinant hosts which contain and express XhoI and
XhoII Type II restriction endonuclease and/or M.XhoI and M.XhoII modification methylase genes.
The present invention is also directed to vectors or DNA molecules which contain these genes,
and to methods of producing these enzymes. One source of these enzymes is Xanthomonas
campestris pv. holcicola, although other microorganisms may be used to isolate the restriction
endonuclease isoschizomers and modification methylase isoschizomers of this invention.

<>

<1>Chatterjee, D.K.
<2>Cloning and expressing restriction endonucleases from Haemophilus.
<3>US Patent Office
<4>US 5248605
<5>
<6>1993
<7>The present invention is directed to recombinant hosts which contain and express the HpaII
Type-II restriction endonuclease gene. The present invention is also directed to vectors or
DNA molecules which contain this gene, and to methods of producing the enzyme. One source of
this enzyme is Haemophilus parainfluenzae, although other microorganisms may be used to
isolate the restriction endonuclease isoschizomers of the invention.

<>

<1>Chatterjee, D.K.
<2>Cloning and expressing restriction endonucleases and modification methylases from Xanthomonas.
<3>US Patent Office
<4>US 5231021
<5>
<6>1993
<7>The present invention is directed to recombinant hosts which contain and express XhoI and
XhoII Type II restriction endonuclease and/or M.XhoI and M.XhoII modification methylase genes.
The present invention is also directed to vectors or DNA molecules which contain these genes,
and to methods of producing these enzymes. One source of these enzymes is Xanthomonas
campestris pv. holcicola, although other microorganisms may be used to isolate the restriction
endonuclease isoschizomers and modification methylase isoschizomers of this invention.

<>

<1>Chatterjee, D.K.
<2>Cloning and expressing restriction endonucleases and modification methylases from Xanthomonas.
<3>International Patent Office
<4>WO 9321307
<5>
<6>1993
<7>The present invention is directed to recombinant hosts which contain and express XhoI and
XhoII Type II restriction endonuclease and/or M.XhoI and M.XhoII modification methylase genes.
The present invention is also directed to vectors or DNA molecules which contain these genes,
and to methods of producing these enzymes.  One source of these enzymes is Xanthomonas
campestris pv. Holcicola, although other microorganisms may be used to isolate the restriction
endonuclease isoschizomers and modification methylase isoschizomers of this invention.

<>

<1>Chatterjee, D.K., Hammond, A.W.
<2>Cloned KpnI restriction-modification system.
<3>International Patent Office
<4>WO 9114771
<5>
<6>1991
<7>The present invention discloses the cloning and expression in a host such as Escherichia coli
of the KpnI restriction-modification system from Klebsiella pneumoniae, utilizing a two step
protocol. Initial protection of the E. coli host with methylase expressed on a vector was
required to stabilize a compatible vector carrying both the endonuclease and the methylase
genes on a single DNA fragment. A chromosomal map was generated localizing the genes for KpnI
methylase and endonuclease. An E. coli strain was constructed which produced several
thousand-fold higher levels of KpnI endonuclease than the level produced by Klebsiella
pneumoniae. This invention is also directed to cloning and expression of genes encoding for
restriction endonuclease isoschizomers of KpnI and/or modification methylase isoschizomers of
KpnI methylase.

<>

<1>Chatterjee, D.K., Hammond, A.W.
<2>Cloned KpnI restriction-modification system.
<3>US Patent Office
<4>US 5192675
<5>
<6>1993
<7>The present invention discloses the cloning and expression in a host such as Escherichia coli
of the KpnI restriction-modification system from Klebsiella pneumoniae, utilizing a two step
protocol.  Initial protection of the E. coli host with methylase expressed on a vector was
required to stabilize a compatible vector carrying both the endonuclease and the methylase
genes on a single DNA fragment.  A chromosomal map was generated localizing the genes for KpnI
methylase and endonuclease.  An E. coli strain was constructed which produced several
thousand-fold higher levels of KpnI endonuclease than the level produced by Klebsiella
pneumoniae.  This invention is also directed to cloning and expression of genes encoding for
restriction endonuclease isoschizomers of KpnI and/or modification methylase isoschizomers of
KpnI methylase.

<>

<1>Chatterjee, D.K., Hammond, A.W.
<2>Cloned KpnI restriction-modification system.
<3>US Patent Office
<4>US 5082784
<5>
<6>1992
<7>The present invention discloses the cloning and expression in a host such as Escherichia coli
of the KpnI restriction-modification system from Klebsiella pneumoniae, utilizing a two step
protocol.  Initial protection of the E. coli host with methylase expressed on a vector was
required to stabilize a compatible vector carrying both the endonuclease and the methylase
genes on a single DNA fragment.  A chromosomal map was generated localizing the genes for KpnI
methylase and endonuclease.  An E. coli strain was constructed which produced several
thousand-fold higher levels of KpnI endonuclease than the level produced by Klebsiella
pneumoniae.  This invention is also directed to cloning and expression of genes encoding for
restriction endonuclease isoschizomers of KpnI and/or modification methylase isoschizomers of
KpnI methylase.

<>

<1>Chatterjee, D.K., Hammond, A.W., Blakesley, R.W., Adams, S.M., Gerard, G.F.
<2>Genetic organization of the KpnI restriction-modification system.
<3>Nucleic Acids Res.
<4>19
<5>6505-6509
<6>1991
<7>The KpnI restriction-modification (KpnI RM) system was previously cloned and
expressed in E. coli.  The nucleotide sequences of the KpnI endonuclease
(R.KpnI) and methylase (M.KpnI) genes have now been determined.  The sequence
of the amino acid residues predicted from the endonuclease gene DNA sequence
and the sequence of the first 12 NH2-terminal amino acids determined from the
purified endonuclease protein were identical.  The KpnIR gene specifies a
protein of 218 amino acids (MW:25,115), while the KpnIM gene coes for a protein
of 417 amino acids (MW:47,582).  The two genes transcribe divergently with an
intergenenic region of 167 nucleotides containing the putative promoter regions
for both genes.  No protein sequence similarity was detected between R.KpnI and
M.KpnI.  Comparison of the amino acid sequence of M.KpnI with sequences of
various methylases revealed a significant homology to N6-adenine methylases, a
partial homology to N4-cytosine methylases, and no homology to C5-methylases.

<>

<1>Chatterjee, D.K., Longo, M., Flynn, E., Oberfelder, R.
<2>Methods for production of proteins.
<3>US Patent Office
<4>US 7223566 A
<5>
<6>2007
<7>The current invention provides methods for producing a polypeptide as inclusion bodies in
bacterial host cells.  The present methods are carried out by forming a gene construct
comprising the genetic sequence encoding a polypeptide operatively linked to that of an
inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such
that host cells comprising the gene construct produce the polypeptide as intracellular
inclusion bodies.  The methods may be useful for producing polypeptides or proteins which are
small and are typically difficult to experss, as well as those proteins that are toxic to host
cells such as E. coli.  The present invention also provides plasmids, vectors and host cells
to be used in the present invention for production of polypeptides, and methods of production
of polypeptides using these vectors and host cells.  The invention further provides methods
for producing protein molecular weight ladders for use in protein gel electrophoresis, as well
as proteins and protein molecular weight ladders produced by these methods.

<>

<1>Chatterjee, P., Brady, K.L., Solem, A., Ho, Y., Caprara, M.G.
<2>Functionally distinct nucleic acid binding sites for a group I intron encoded RNA maturase/DNA homing endonuclease.
<3>J. Mol. Biol.
<4>329
<5>239-251
<6>2003
<7>A large number of group I introns encode a family of homologous proteins that either promote
intron splicing (maturases) or are site-specific DNA
endonucleases that function in intron mobility (a process called
"homing"). Genetic studies have shown that some of these proteins have
both activities, yet how a single protein carries out both functions
remains obscure. The similarity between respective DNA-binding sites and
the RNA structure near the 5' and 3' splice sites has fueled speculation
that such proteins may use analogous interactions to perform both
functions. The Aspergillus nidulans mitochondrial COB group I intron
encodes a bi-functional protein, I-AniI, that has both RNA maturase and
site-specific DNA endonuclease activities in vitro. Here, we show that
I-AniI shows distinctive features of the endonuclease family to which it
belongs, including highly specific, tight binding and sequential DNA
strand cleavage. Competition experiments demonstrate that I-AniI binds the
COB intron RNA even in saturating concentrations of its DNA target site
substrate, suggesting that the protein has a separate binding site for
RNA. In addition, we provide evidence that two different DNA-binding site
mutants of I-AniI have little effect on the protein's RNA maturation
activity. Since RNA splicing is likely a secondary adaptation of the
protein, these observations support a model in which homing endonucleases
may have developed maturase function by utilizing a hitherto
"non-functional" protein surface.

<>

<1>Chaturvedi, D., Chakravorty, M.
<2>Identification of an unusual restriction-modification system in bacteriophage MB78.
<3>Biochem. Soc. Trans.
<4>28
<5>A171
<6>2000
<7>A restriction-modification system has been identified in bacteriophage MB78, a virulent phage
of Salmonella typhimurium isolated in this laboratory.  Genomic fragment of MB78 carrying the
corresponding genes has been cloned in both orientations in pUC19.  The clones carrying the
genes in the same orientation as that of the lacZ are mostly unstable.  However, those cloned
in opposite orientation are stable.  Viability of transformants is strain-, orientation- and
medium-dependent.  To clone the genes for restriction and modification enzymes separately, the
genomic fragment containing the restriction-modification system was subcloned using AccI and
SmaI enzymes.  One set of subclones contained active methylase gene, while the others
contained an active endonuclease gene as well as truncated (inactive) methylase gene.  The
former are stable and normal looking while the latter are unstable and looked distressed.
Both the genes have been sequenced.  The methylase and restriction genes (only 228 and 333 bp
respectively) are unusually small in comparison to respective bacterial genes.

<>

<1>Chaturvedi, D., Chakravorty, M.
<2>Restriction-modification system in bacteriophage MB78.
<3>Biochem. Biophys. Res. Commun.
<4>303
<5>884-890
<6>2003
<7>Restriction-modification system is present in bacteria to protect the cells against phage
infection. Interestingly, the bacteriophage MB78, a
virulent phage of Salmonella typhimurium possesses
restriction-modification system. Permissive host transformed with plasmid
having the genomic fragment of MB78 carrying the putative
restriction-modification genes severely restrict the growth of the phage
9NA. Growth of phage MB78 is also restricted to some extent. However, the
temperate phage P22 is not restricted at all. Cloning of the the putative
restriction-modification genes has been done in both orientations in
different vectors. The clones carrying the genes in the same orientation
as that of the lacZ in pUC19 are mostly unstable. However, those are
stable when cloned in opposite orientation. Viability of the transformants
is strain-, orientation-, and medium-dependent. The two genes have also
been cloned individually/separately. Hosts carrying only the modification
gene do not restrict growth of phages while the hosts carrying only the
restriction gene do. The former produces stable transformants while the
latter produces very unstable transformants which were viable only upto 36
h or so. The colonies carrying modification gene were normal looking while
those carrying the restriction gene were tiny, flat, and looked distressed
resembling very much the clones carrying bacterial
restriction-modification system. Amplification of the genes and subsequent
cloning in expression vector will be carried out for characterization of
the enzymes.

<>

<1>Chaudhuri, R.R. et al.
<2>Complete genome sequence and comparative metabolic profiling of the prototypical enteroaggregative Escherichia coli strain 042.
<3>PLoS ONE
<4>5
<5>e8801
<6>2010
<7>BACKGROUND: Escherichia coli can experience a multifaceted life, in some cases acting as a
commensal while in other cases causing intestinal and/or
extraintestinal disease. Several studies suggest enteroaggregative E. coli
are the predominant cause of E. coli-mediated diarrhea in the developed
world and are second only to Campylobacter sp. as a cause of
bacterial-mediated diarrhea. Furthermore, enteroaggregative E. coli are a
predominant cause of persistent diarrhea in the developing world where
infection has been associated with malnourishment and growth retardation.
METHODS: In this study we determined the complete genomic sequence of E.
coli 042, the prototypical member of the enteroaggregative E. coli, which
has been shown to cause disease in volunteer studies. We performed genomic
and phylogenetic comparisons with other E. coli strains revealing
previously uncharacterised virulence factors including a variety of
secreted proteins and a capsular polysaccharide biosynthetic locus. In
addition, by using Biolog Phenotype Microarrays we have provided a full
metabolic profiling of E. coli 042 and the non-pathogenic lab strain E.
coli K-12. We have highlighted the genetic basis for many of the metabolic
differences between E. coli 042 and E. coli K-12. CONCLUSION: This study
provides a genetic context for the vast amount of experimental and
epidemiological data published thus far and provides a template for future
diagnostic and intervention strategies.

<>

<1>Chauhan, A., Green, S., Pathak, A., Thomas, J., Venkatramanan, R.
<2>Whole-genome sequences of five oyster-associated bacteria show potential for crude oil hydrocarbon degradation.
<3>Genome Announcements
<4>1
<5>e00802-13
<6>2013
<7>Draft genome sequences of oyster-associated Pseudomonas stutzeri strain MF28, P.  alcaligenes
strain OT69, P. aeruginosa strain WC55, Stenotrophomonas maltophilia
strain MF89, and Microbacterium maritypicum strain MF109 are reported.
Genome-wide surveys of these isolates suggest that the oyster microbiome, which
remains largely understudied, has a strong potential to degrade crude oil.

<>

<1>Chauhan, A., Layton, A.C., Williams, D., Smartt, A.E., Ripp, S., Karpinets, T.V., Brown, S.D., Sayler, G.S.
<2>Draft Genome Sequence of the Polycyclic Aromatic Hydrocarbon-Degrading, Genetically Engineered Bioluminescent Bioreporter Pseudomonas fluorescens  HK44.
<3>J. Bacteriol.
<4>193
<5>5009
<6>2011
<7>Pseudomonas fluorescens strain HK44 (DSM 6700) is a genetically engineered lux-based
bioluminescent bioreporter. Here we report the draft genome
sequence of strain HK44. Annotation of  approximately 6.1 Mb sequence
indicates that 30% of the traits are unique and distributed over 5 genomic
islands, a prophage and two plasmids.

<>

<1>Chauthaiwale, V.M., Vyas, P.R., Deshpande, V.V.
<2>Genetic transformation of Chainia and heat attenuation of its restriction system.
<3>Can. J. Microbiol.
<4>37
<5>713-715
<6>1991
<7>A PEG-mediated transformation system for Chainia (NCL 82-5-1) was developed
using a broad host range Streptomyces vector, pIJ702.  Protoplasts prepared
from Chainia (NCL 81-5-1) were regenerated with 5% efficiency.  Transformation
of the protoplasts with pIJ702 gave 10-20 transformants/microgram DNA.  The low
efficiency of transformation is attributed to a restriction system in Chainia;
this could be inhibited by treating the protoplasts at 42C for 10 min just
before transformation.  The yield of transformants increased 100-fold when
pIJ702 was modified by passage in Chainia.  Because the plasmid replicon was
functional in Chainia and the modified plasmid was stably maintained, the
transformation system should be useful for self-cloning in Chainia NCL 82-5-1
of the many commercially important enzymes this strain is known to produce.

<>

<1>Chavez, S.L., Pera, R.A.R.
<2>Identification of the De Novo DNA Methyltransferase, DNMT3A2, as a Novel Regulator of Germ Cell Development.
<3>Reprod. Sci.
<4>18
<5>172A
<6>2011
<7>The methylation of DNA is mediated by a family of DNA methyltransferases which play a role in
the establishment and/or maintenance of DNA methylation patterns.  Previously, it was shown
that the targeted disruption of both active isoforms of the de novo methyltransferase, Dnmt3a,
in germ cells by conditional mouse knockout technology resulted in aberrant gametogenesis.
The aim of this study was to evaluate the expression and function of DNMT3A in human fetal
gonads and human embryonic stem cell (hESC)-derived germ cell differentiation.  In addition,
the role of Dnmt3a in mouse primordial germ cells isolated from fetal gonads throughout
development and in mouse embryonic germ cells, the pluripotent cells that PGCs form in vitro,
was also investigated.

<>

<1>Chavez-Bueno, S., Day, M.W., Toby, I.T., Akins, D.R., Dyer, D.W.
<2>Genome Sequence of SCB34, a Sequence Type 131 Multidrug-Resistant Escherichia coli Isolate Causing Neonatal Early-Onset Sepsis.
<3>Genome Announcements
<4>2
<5>e00514-14
<6>2014
<7>SCB34 is a sequence type 131, highly invasive, multidrug-resistant Escherichia coli isolate
that produced neonatal bacteremia. Whole-genome sequencing was
performed using a 250-bp library on the Illumina MiSeq platform; 5,910,264 reads
were assembled de novo using the A5 assembly pipeline. The total contig length
was 5,227,742 bp; the RAST server was used for annotation.

<>

<1>Che, J., Liu, B., Lin, Y., Tang, W., Tang, J.
<2>Draft Genome Sequence of Biocontrol Bacterium Brevibacillus brevis Strain FJAT-0809-GLX.
<3>Genome Announcements
<4>1
<5>e00160-13
<6>2013
<7>Brevibacillus brevis strain FJAT-0809-GLX had significant inhibition on many plant and animal
pathogens. The draft genome sequence of B. brevis FJAT-0809-GLX
is 6 Mb in size and consists of 5,677 genes (protein-coding sequences [CDS]),
with an average length of 933 bp and a G+C content of 47.30%. Compared with the
published B. brevis strain NBRC 100599, 618 specific genes were identified in the
strain FJAT-0809-GLX.

<>

<1>Che, S., Song, L., Song, W., Yang, M., Liu, G., Lin, X.
<2>Complete Genome Sequence of Antarctic Bacterium Psychrobacter sp. Strain G.
<3>Genome Announcements
<4>1
<5>e00725-13
<6>2013
<7>Here, we report the complete genome sequence of Psychrobacter sp. strain G, isolated from King
George Island, Antarctica, which can produce lipolytic enzymes
at low temperatures. The genomics information of this strain will facilitate the
study of the physiology, cold adaptation properties, and evolution of this genus.

<>

<1>Chedin, F.
<2>The DNMT3 Family of Mammalian De Novo DNA Methyltransferases.
<3>Prog. Mol. Biol. Transl. Sci.
<4>101
<5>255-285
<6>2011
<7>The deposition of DNA methylation at promoters of transposons, X-linked genes, imprinted
genes, and other lineage-specific genes is clearly
associated with long-term transcriptional silencing. Thus, DNA
methylation represents a key layer of epigenetic information in mammals
that is required for embryonic development, germline differentiation,
and, as shown more recently, for the function and maturation of
neuronal tissues. The DNMT3A, DNMT3B, and DNMT3L proteins are primarily
responsible for the establishment of genomic DNA methylation patterns
and, as such, play an important role in human developmental,
reproductive, and mental health. Progress in our understanding of this
important protein family has been rapid in recent years and has been
accompanied by stunning developments in the analysis of the human DNA
methylome in multiple cell types. This review focuses on recent
developments in the characterization of the DNMT3 family of DNA
methyltransferases at the biochemical, structural, and functional
levels. Interconnections between the DNA-based and histone-based layers
of epigenetic information are particularly highlighted, as it is now
clear that de now methylation occurs chiefly in the context of
nucleosomal templates.

<>

<1>Chedin, F., Lieber, M.R., Hsieh, C.L.
<2>The DNA methyltransferase-like protein DNMT3L stimulates de novo methylation by Dnmt3a.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>16916-16921
<6>2002
<7>Dnmt3L is required for the establishment of maternal methylation imprints at imprinting
centers (ICs). Dnmt3L, however, lacks the conserved
catalytic domain common to DNA methyltransferases. In an attempt to define
its function, we coexpressed DNMT3L with each of the two known de novo
methyltransferases, Dnmt3a and DNMT3B, in human cells and monitored de
novo methylation by using replicating minichromosomes carrying various ICs
as targets. Coexpression of DNMT3L with DNMT3B led to little or no change
in target methylation. However, coexpression of DNMT3L with Dnmt3a
resulted in a striking stimulation of de novo methylation by Dnmt3a.
Stimulation was observed at maternally methylated ICs such as small
nuclear ribonucleoprotein polypeptide N (SNRPN), Snrpn, and Igf2rAir, as
well as at various nonimprinted sequences present on the episomes.
Stimulation of Dnmt3a by DNMT3L was also observed at endogenous sequences
in the genome. Therefore, DNMT3L acts as a general stimulatory factor for
de novo methylation by Dnmt3a. The implications of these findings for the
function of DNMT3L and Dnmt3a in DNA methylation and genomic imprinting
are discussed.

<>

<1>Cheevadhanarak, S. et al.
<2>Draft genome sequence of Arthrospira platensis C1 (PCC9438).
<3>Standards in Genomic Sciences
<4>6
<5>43-53
<6>2012
<7>Arthrospira platensis is a cyanobacterium that is extensively cultivated outdoors on a large
commercial scale for consumption as a food for humans and animals. It can be grown in
monoculture under highly alkaline conditions, making it attractive for industrial production.
Here we describe the complete genome sequence of A. platensis C1 strain and its annotation.
The A. platensis C1 genome contains 6,089,210 bp including 6,108 protein-coding genes and 45
RNA genes, and no plasmids. The genome information has been used for further comparative
analysis, particularly of metabolic pathways, photosynthetic efficiency and barriers to gene
transfer.

<>

<1>Chen, A., Powell, L.M., Dryden, D.T.F., Murray, N.E., Brown, T.
<2>Tyrosine 27 of the specificity polypeptide of EcoKI can be UV crosslinked to a bromodeoxyuridine-substituted DNA target sequence.
<3>Nucleic Acids Res.
<4>23
<5>1177-1183
<6>1995
<7>The specificity (S) subunit of the restriction enzyme EcoKI imparts specificity for the
sequence AAC(N6)GTGC. Substitution of thymine with bromodeoxyuridine in a 25 bp DNA duplex
containing this sequence stimulated UV light-induced covalent cross-linking to the S subunit.
Crosslinking occurred only at the residue complementary to the first adenine in the AAC
sequence, demonstrating a close contact between the major groove at this sequence and the S
subunit. Peptide sequencing of a proteolytically-digested, crosslinked complex identified
tyrosine 27 in the S subunit as the site of crosslinking. This is consistent with the role of
the N-terminal domain of the S subunit in recognizing the AAC sequence. Tyrosine 27 is
conserved in the S subunits of the three type I enzymes that share the sequence AA in the
trinucleotide component of their target sequence. This suggests that tyrosine 27 may make a
similar DNA contact in these other enzymes.

<>

<1>Chen, B.S., Otten, L.G., Resch, V., Muyzer, G., Hanefeld, U.
<2>Draft genome sequence of Rhodococcus rhodochrous strain ATCC 17895.
<3>Standards in Genomic Sciences
<4>9
<5>175-184
<6>2013
<7>Rhodococcus rhodochrous ATCC 17895 possesses an array of mono- and dioxygenases,  as well as
hydratases, which makes it an interesting organism for biocatalysis.
R. rhodochrous is a Gram-positive aerobic bacterium with a rod-like morphology.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. The 6,869,887 bp long genome contains 6,609
protein-coding genes and 53 RNA genes. Based on small subunit rRNA analysis, the
strain is more likely to be a strain of Rhodococcus erythropolis rather than
Rhodococcus rhodochrous.

<>

<1>Chen, Bi.-F., Chan, W.-Yee.
<2>The de novo DNA methyltransferase DNMT3A in development and cancer.
<3>EPIGENETICS
<4>9
<5>669-677
<6>2014
<7>DNA methylation, one of the best-characterized epigenetic modifications, plays essential roles
in development, aging and diseases. The de novo DNA methyltransferase DNMT3A is responsible
for the establishment of de novo genomic DNA methylation patterns and, as such, involved in
normal development as well as in many diseases including cancer. In recent years, our
understanding of this important protein has made significant progress, which was facilitated
by stunning development in the analysis of the DNA methylome of multiple organs and cell
types. In this review, recent developments in the characterization of DNMT3A were discussed
with special emphasis on the roles of DNMT3A in development and cancer.

<>

<1>Chen, C. et al.
<2>A Glimpse of Streptococcal Toxic Shock Syndrome from Comparative Genomics of S. suis 2 Chinese Isolates.
<3>PLoS ONE
<4>2
<5>e315
<6>2007
<7>BACKGROUND: Streptococcus suis serotype 2 (SS2) is an important zoonotic pathogen, causing
more than 200 cases of severe human infection worldwide,
with the hallmarks of meningitis, septicemia, arthritis, etc. Very
recently, SS2 has been recognized as an etiological agent for
streptococcal toxic shock syndrome (STSS), which was originally associated
with Streptococcus pyogenes (GAS) in Streptococci. However, the molecular
mechanisms underlying STSS are poorly understood. METHODS AND FINDINGS: To
elucidate the genetic determinants of STSS caused by SS2, whole genome
sequencing of 3 different Chinese SS2 strains was undertaken. Comparative
genomics accompanied by several lines of experiments, including
experimental animal infection, PCR assay, and expression analysis, were
utilized to further dissect a candidate pathogenicity island (PAI). Here
we show, for the first time, a novel molecular insight into Chinese
isolates of highly invasive SS2, which caused two large-scale human STSS
outbreaks in China. A candidate PAI of approximately 89 kb in length,
which is designated 89K and specific for Chinese SS2 virulent isolates,
was investigated at the genomic level. It shares the universal properties
of PAIs such as distinct GC content, consistent with its pivotal role in
STSS and high virulence. CONCLUSIONS: To our knowledge, this is the first
PAI candidate from S. suis worldwide. Our finding thus sheds light on STSS
triggered by SS2 at the genomic level, facilitates further understanding
of its pathogenesis and points to directions of development on some
effective strategies to combat highly pathogenic SS2 infections.

<>

<1>Chen, C., Ai, L., Zhou, F., Wang, L., Zhang, H., Chen, W., Guo, B.
<2>Complete genome sequence of the probiotic bacterium Lactobacillus casei LC2W.
<3>J. Bacteriol.
<4>193
<5>3419-3420
<6>2011
<7>Lactobacillus casei LC2W, a patented probiotic strain (EP 164209630B1), is isolated from
Chinese traditional dairy products and has been implemented
in the industrial production as starter cultures. Here we present the
complete genome sequence of LC2W and the identification of a gene cluster
implicated in the biosynthesis of exopolysaccharides.

<>

<1>Chen, C., Kittichotirat, W., Chen, W., Downey, J.S., Bumgarner, R.
<2>Genome Sequence of a Serotype b Non-JP2 Aggregatibacter actinomycetemcomitans Strain, ANH9381, from a Periodontally Healthy Individual.
<3>J. Bacteriol.
<4>194
<5>1837
<6>2012
<7>Gram-negative Aggregatibacter actinomycetemcomitans can be distinguished (based on the
promoter structure of the leukotoxin operon) into JP2 and non-JP2
genotypes, with the former found to be more pathogenic than the latter. Here we
report the first complete genome sequence of a serotype b non-JP2 strain of A.
actinomycetemcomitans.

<>

<1>Chen, C., Kittichotirat, W., Chen, W., Downey, J.S., Si, Y., Bumgarner, R.
<2>Genome sequence of naturally competent Aggregatibacter actinomycetemcomitans serotype a strain D7S-1.
<3>J. Bacteriol.
<4>192
<5>2643-2644
<6>2010
<7>The major clonal lineages of the Gram-negative periodontal pathogen Aggregatibacter
actinomycetemcomitans include serotype a, b, and c
strains. Here, we report the draft genome sequence of a naturally
competent serotype a strain, D7S-1, isolated from a patient with
aggressive periodontitis.

<>

<1>Chen, C., Kittichotirat, W., Si, Y., Bumgarner, R.
<2>Genome sequence of Aggregatibacter actinomycetemcomitans serotype c strain D11S-1.
<3>J. Bacteriol.
<4>191
<5>7378-7379
<6>2009
<7>Aggregatibacter actinomycetemcomitans is a major etiological agent of periodontitis. Here we
report the complete genome sequence of a serotype c strain D11S-1, which was recovered from
the subgingival plaque of a patient diagnosed with generalized aggressive periodontitis.

<>

<1>Chen, C.-H.B., Sigman, D.S.
<2>Chemical Conversion of a DNA-binding protein into a site-specific nuclease.
<3>Science
<4>237
<5>1197-1201
<6>1987
<7>The tryptophan gene (trp) repressor of Escherichia coli has been converted into
a site-specific nuclease by covalently attaching it to the
1,10-phenanthroline-copper complex.  In its cuprous form, the coordination
complex with hydrogen peroxide as a coreactant cleaves DNA by oxidatively
attacking the deoxyribose moiety.  The chemistry for the attachment of
1,10-phenanthroline to the trp repressor involves modification of lysyl
residues with iminothiolane followed by alkylation of the resulting sulfhydryl
groups with 5-iodoacetamido-1,10-phenanthroline.  The modified trp repressor
cleaves the operators of aroH and trpEDCBA upon the addition of cupric ion and
thiol in a reaction dependent on the corepressor L-tryptophan.  Scission was
restricted to the binding site for the repressor, defined by deoxyribonuclease
I footprinting.  Since DNA-binding proteins have recognition sequences
approximately 20 base pairs long, the nucleolytic activities derived from them
could be used to isolate long DNA fragments for sequencing or chromosomal
mapping.

<>

<1>Chen, C.-K., Boucle, C.M., Blaschek, H.P.
<2>Factors involved in the transformation of previously non-transformable Clostridium perfringens type B.
<3>FEMS Microbiol. Lett.
<4>140
<5>185-191
<6>1996
<7>The pre-shock incubation of cells plus DNA and the methylation state of plasmid DNA were found
to play a role in the electroporation-based transformation of Clostridium perfringens 3626B.
Following pre-shock incubation, the highest number of C. perfringens 3626B transformants was
obtained when plasmid pGK201 was both dam+ dcm+ modified, while no transformants were obtained
when pGK201 was not methylated or only dcm methylated.  This is consistent with the
observation that plasmid pGK201 was protected against digestion by C. perfringens 3626B
cell-associated nucleases for up to 3 min when methylated by both methylases.  C. perfringens
3626B was successfully transformed only within a narrow cell recovery rate window.  The ermAM
gene associated with pGK201 and pAK102 was found to integrate into the chromosome of C.
perfringens strains 13A and 3626B.

<>

<1>Chen, C.C., Hsia, K.C., Huang, C.T., Wong, W.W., Yen, M.Y., Li, L.H., Lin, K.Y., Chen, K.W., Li, S.Y.
<2>Draft Genome Sequence of a Dominant Multidrug-resistant Neisseria gonorrhoeae strain TCDC-NG08107 from Sexual Network at High Risk of  Acquiring Human Immunodeficiency Virus and Syphilis.
<3>J. Bacteriol.
<4>193
<5>1788-1789
<6>2011
<7>Neisseria gonorrhoeae infection is the second major cause of sexually transmitted diseases
worldwide. Development of resistance to multiple classes of antimicrobials in N. gonorrhoeae
has compromised treatment and disease control. Herein, we report the availability of the draft
genome sequence of a multidrug-resistant N. gonorrhoeae isolate, TCDC-NG08107, which spread in
groups of men who have sex with men (MSM) in Taiwan.

<>

<1>Chen, C.C., Lin, Y.C., Sheng, W.H., Chen, Y.C., Chang, S.C., Hsia, K.C., Liao, M.H., Li, S.Y.
<2>Genome Sequence of a Dominant Multidrug-resistant Acinetobacter baumannii strain TCDC-AB0715.
<3>J. Bacteriol.
<4>193
<5>2361-2362
<6>2011
<7>Acinetobacter baumannii has emerged as a significant nosocomial pathogen worldwide. The
increasing trend of carbapenem and fluoroquinolone resistance in A. baumannii severely limits
the usage of therapeutic antimicrobial agents. Here, we report the genome sequence of a
multidrug-resistant A. baumannii strain TCDC-AB0715 harboring both blaOXA-23 and blaOXA-66.

<>

<1>Chen, C.C., Wang, K.Y., Shen, C.K.J.
<2>The Mammalian de Novo DNA Methyltransferases DNMT3A and DNMT3B Are Also DNA 5-Hydroxymethylcytosine Dehydroxymethylases.
<3>J. Biol. Chem.
<4>287
<5>33116-33121
<6>2012
<7>For cytosine (C) demethylation of vertebrate DNA, it is known that the TET proteins could
convert 5-methyl C (5-mC) to 5-hydroxymethyl C
(5-hmC). However, DNA dehydroxymethylase(s), or enzymes able to
directly convert 5-hmC to C, have been elusive. We present in vitro
evidence that the mammalian de novo DNA methyltransferases DNMT3A and
DNMT3B, but not the maintenance enzyme DNMT1, are also redox-dependent
DNA dehydroxymethylases. Significantly, intactness of the C methylation
catalytic sites of these de novo enzymes is also required for their
5-hmC dehydroxymethylation activity. That DNMT3A and DNMT3B function
bidirectionally both as DNA methyltransferases and as
dehydroxymethylases raises intriguing and new questions regarding the
structural and functional aspects of these enzymes and their regulatory
roles in the dynamic modifications of the vertebrate genomes during
development, carcinogenesis, and gene regulation.

<>

<1>Chen, C.C., Wang, K.Y., Shen, C.K.J.
<2>DNA 5-Methylcytosine Demethylation Activities of the Mammalian DNA Methyltransferases.
<3>J. Biol. Chem.
<4>288
<5>9084-9091
<6>2013
<7>Methylation at the 5-position of DNA cytosine on the vertebrate genomes is accomplished by the
combined catalytic actions of three DNA
methyltransferases (DNMTs), the de novo enzymes DNMT3A and DNMT3B and
the maintenance enzyme DNMT1. Although several metabolic routes have
been suggested for demethylation of the vertebrate DNA, whether active
DNA demethylase(s) exist has remained elusive. Surprisingly, we have
found that the mammalian DNMTs, and likely the vertebrates DNMTs in
general, can also act as Ca2+ ion- and redox state-dependent active DNA
demethylases. This finding suggests new directions for reinvestigation
of the structures and functions of these DNMTs, in particular their
roles in Ca2+ ion-dependent biological processes, including the
genome-wide/local DNA demethylation during early embryogenesis, cell
differentiation, neuronal activity-regulated gene expression, and
carcinogenesis.

<>

<1>Chen, C.J., Unger, C., Hoffmann, W., Lindsay, J.A., Huang, Y.C., Gotz, F.
<2>Characterization and Comparison of 2 Distinct Epidemic Community-Associated Methicillin-Resistant Staphylococcus aureus Clones of ST59 Lineage.
<3>PLoS ONE
<4>8
<5>E63210
<6>2013
<7>Sequence type (ST) 59 is an epidemic lineage of community-associated (CA)
methicillin-resistant Staphylococcus aureus (MRSA) isolates. Taiwanese CA-MRSA
isolates belong to ST59 and can be grouped into 2 distinct clones, a virulent
Taiwan clone and a commensal Asian-Pacific clone. The Taiwan clone carries the
Panton-Valentine leukocidin (PVL) genes and the staphylococcal chromosomal
cassette mec (SCCmec) VT, and is frequently isolated from patients with severe
disease. The Asian-Pacific clone is PVL-negative, carries SCCmec IV, and a
frequent colonizer of healthy children. Isolates of both clones were
characterized by their ability to adhere to respiratory A549 cells, cytotoxicity
to human neutrophils, and nasal colonization of a murine and murine sepsis
models. Genome variation was determined by polymerase chain reaction of selected
virulence factors and by multi-strain whole genome microarray. Additionally, the
expression of selected factors was compared between the 2 clones. The Taiwan
clone showed a much higher cytotoxicity to the human neutrophils and caused more
severe septic infections with a high mortality rate in the murine model. The
clones were indistinguishable in their adhesion to A549 cells and persistence of
murine nasal colonization. The microarray data revealed that the Taiwan clone had
lost the o3-prophage that integrates into the beta-hemolysin gene and includes
staphylokinase- and enterotoxin P-encoding genes, but had retained the genes for
human immune evasion, scn and chps. Production of the virulence factors did not
differ significantly in the 2 clonal groups, although more alpha-toxin was
expressed in Taiwan clone isolates from pneumonia patients. In conclusion, the
Taiwan CA-MRSA clone was distinguished by enhanced virulence in both humans and
an animal infection model. The evolutionary acquisition of PVL, the higher
expression of alpha-toxin, and possibly the loss of a large portion of the
beta-hemolysin-converting prophage likely contribute to its higher pathogenic
potential than the Asian-Pacific clone.

<>

<1>Chen, C.J., Wu, T.L., Lu, P.L., Chen, Y.T., Fung, C.P., Chuang, Y.C., Lin, J.C., Siu, L.K.
<2>Closely Related NDM-1-Encoding Plasmids from Escherichia coli and Klebsiella pneumoniae in Taiwan.
<3>PLoS ONE
<4>9
<5>E104899
<6>2014
<7>OBJECTIVE: Two plasmids carrying blaNDM-1 isolated from carbapenem-resistant
Klebsiella pneumoniae (CR-KP) and carbapenem-resistant Escherichia coli (CR-EC)
were sequenced. CR-KP and CR-EC were isolated from two Taiwanese patients without
travel histories. METHODS: Complete sequencing of the plasmids (pLK75 and pLK78)
was conducted using a shotgun approach. Annotation of the contigs was performed
using the RAST Server, followed by manual inspection and correction. RESULTS:
These similar plasmids were obtained from two patients with overlapping stays at
the same hospital. The pLK75 and pLK78 plasmids were 56,489-bp and 56,072-bp in
length, respectively. Plasmid annotation revealed a common backbone similar to
the IncN plasmid pR46. The regions flanking the blaNDM-1 genes in these plasmids
were very similar to plasmid pNDM-HU01 in Japan, which contains a complex class 1
integron located next to an ISCR1 element. The ISCR1 element has been suggested
to provide a powerful mechanism for mobilising antibiotic resistance genes.
CONCLUSION: Two indigenous NDM-1-producing Enterobacteriaceae cases were
identified for the first time in Taiwan, highlighting the alarming introduction
of NDM-1-producing Enterobacteriaceae in this region.

<>

<1>Chen, C.Y., Wu, K.M., Chang, Y.C., Chang, C.H., Tsai, H.C., Liao, T.L., Liu, Y.M., Chen, H.J., Shen, A.B., Li, J.C., Su, T.L., Shao, C.P., Lee, C.T., Hor, L.I., Tsai, S.F.
<2>Comparative genome analysis of Vibrio vulnificus, a marine pathogen.
<3>Genome Res.
<4>13
<5>2577-2587
<6>2003
<7>The halophile Vibrio vulnificus is an etiologic agent of human mortality from seafood-borne
infections. We applied whole-genome sequencing and
comparative analysis to investigate the evolution of this pathogen. The
genome of biotype 1 strain, V. vulnificus YJ016, was sequenced and
includes two chromosomes of estimated 3377 kbp and 1857 kbp in size, and a
plasmid of 48,508 bp. A super-integron (SI) was identified, and the SI
region spans 139 kbp and contains 188 gene cassettes. In contrast to
non-SI sequences, the captured gene cassettes are unique for any given
Vibrio species and are highly variable among V. vulnificus strains.
Multiple rearrangements were found when comparing the 5.3-Mbp V.
vulnificus YJ016 genome and the 4.0-Mbp V. cholerae El Tor N16961 genome.
The organization of gene clusters of capsular polysaccharide, iron
metabolism, and RTX toxin showed distinct genetic features of V.
vulnificus and V. cholerae. The content of the V. vulnificus genome
contained gene duplications and evidence of horizontal transfer, allowing
for genetic diversity and function in the marine environment. The genomic
information obtained in this study can be applied to monitoring vibrio
infections and identifying virulence genes in V. vulnificus.

<>

<1>Chen, D.
<2>Chemical modification of restriction endonuclease Bsp63I and its substrate.
<3>Hunan Jiaoyu Xueyuan Xuebao
<4>15
<5>91-94
<6>1997
<7>This paper reports on a study of the chemical modification of Bsp63I and its substrate by
using group modification reagents.  The results show that the sulfhydryl group and the lysine
residue are possibly the necessary group in the active center of Bsp63I.  The base sequence of
d(GC), which is in the recognition sequence of Bsp63I, plays an important role in the
catalysis of Bsp63I.

<>

<1>Chen, D., Liu, Q., Chen, X., Zhao, X., Chen, Y.
<2>The inhibition of restriction endonuclease PvuII cleavage activity by methylation outside its recognition sequence.
<3>Nucleic Acids Res.
<4>19
<5>5703-5705
<6>1991
<7>The recombinant plasmid pGEM4Z-ras DNA which was methylated on dam and dcm sites outside the
PvuII recognition sequence was digested with restriction endonuclease PvuII, and one of the
three PvuII sites was about 16-fold less efficient to cleave than either of the other two. On
the contrary, the three PvuII sites were cleaved at about the same rate on the unmethylated
DNA molecule. The results show that the cleavage inhibition of the methylated DNA on the
certain PvuII site was caused by methylation outside the PvuII recognition sequence. Maybe an
adjacent methylated dam site *A was responsible for the less efficient cleavage. This
observation suggests that methylation outside the recognition sequence may be considered a new
factor in the kinetic experiment of restriction endonuclease.

<>

<1>Chen, D., Sun, M., Liu, Y., Shi, G., Chen, Y.
<2>Molecular mechanism of the inhibitory effect of methylation outside the recognition sequence of restriction endonuclease PvuII on its cleavage activity.
<3>Chinese Sci. Bull.
<4>43
<5>47-53
<6>1998
<7>In 1991, we found that methylation outside the PvuII recognition sequence could partially
inhibit its cleavage activity.  To clarify the molecular mechanism, three plasmids with
different methylation states were constructed.  Then, together with the original one, four
plasmids were digested with different amounts of PvuII.  Results show that methylation on both
sites results in 90% inhibition; moving the methylated site one base further away decreases
the inhibitory effect to about 30%; with the adjacent dam methylation site eliminated, the
inhibitory effect disappears.  The data suggest that the inhibition of cleavage activity
caused by outside methylation is not "all or none", and the degree of inhibition is dependent
on the position and the number of methylated bases.

<>

<1>Chen, F., Yang, Z., Yan, M., Alvarado, J.B., Wang, G., Benner, S.A.
<2>Recognition of an expanded genetic alphabet by type-II restriction endonucleases and their application to analyze polymerase fidelity.
<3>Nucleic Acids Res.
<4>39
<5>3949-3961
<6>2011
<7>To explore the possibility of using restriction enzymes in a synthetic biology based on
artificially expanded genetic information systems
(AEGIS), 24 type-II restriction endonucleases (REases) were challenged to
digest DNA duplexes containing recognition sites where individual Cs and
Gs were replaced by the AEGIS nucleotides Z and P [respectively,
6-amino-5-nitro-3-(1'-beta-d-2'-deoxyribofuranosyl)-2(1H)-pyridone and
2-amino-8-(1'-beta-d-2'-deoxyribofuranosyl)-imidazo[1,2-a]-1,3,5-triazin-4
(8H)-one]. These AEGIS nucleotides implement complementary hydrogen bond
donor-donor-acceptor and acceptor-acceptor-donor patterns. Results allowed
us to classify type-II REases into five groups based on their performance,
and to infer some specifics of their interactions with functional groups
in the major and minor grooves of the target DNA. For three enzymes among
these 24 where crystal structures are available (BcnI, EcoO109I and NotI),
these interactions were modeled. Further, we applied a type-II REase to
quantitate the fidelity polymerases challenged to maintain in a DNA duplex
C:G, T:A and Z:P pairs through repetitive PCR cycles. This work thus adds
tools that are able to manipulate this expanded genetic alphabet in vitro,
provides some structural insights into the working of restriction enzymes,
and offers some preliminary data needed to take the next step in synthetic
biology to use an artificial genetic system inside of living bacterial
cells.

<>

<1>Chen, F.J., Lauderdale, T.L., Wang, L.S., Huang, I.W.
<2>Complete Genome Sequence of Staphylococcus aureus Z172, a Vancomycin-Intermediate and Daptomycin-Nonsusceptible Methicillin-Resistant Strain Isolated in Taiwan.
<3>Genome Announcements
<4>1
<5>e01011-13
<6>2013
<7>We report the complete genome sequence of Z172, a representative strain of sequence type
239-staphylococcal cassette chromosome mec type III (ST239-SCCmec
type III) hospital-associated methicillin-resistant Staphylococcus aureus in
Taiwan. Strain Z172 also exhibits a vancomycin-intermediate and
daptomycin-nonsusceptible phenotype.

<>

<1>Chen, G.C.C., Brown, A., Lema, M.W.
<2>Restriction endonuclease activities in the legionellae.
<3>Can. J. Microbiol.
<4>32
<5>591-593
<6>1986
<7>Studies of the restriction-modification system of bacteria led to the discovery
of a number of endonucleases which recognize and cleave specific DNA base
sequences.  These recognition sequences often consist of short palindromes and,
in a number of instances, several different enzymes have been found with the
same site specificity (isochizomers).  Because of their specificity,
restriction endonucleases have been useful for the construction of recombinant
DNA molecules and for DNA sequencing.  This system for detecting and destroying
"foreign" DNA is found in such taxonomically diverse organisms as
Acinetobacter, Nocardia, Nostoc, and Thermoplasma.  It is, therefore, not
surprising to find restriction endonucleases in new groups of organisms as they
are studied.  Because of differences in the recipient ability of several
legionellae (Chen et al. 1984), we examined these strains for the presence of
these enzymes.  We now report that several different restriction activities are
present in various legionellae strains.

<>

<1>Chen, H.-P., Zhu, S.-H., Casabon, I., Hallam, S.J., Crocker, F.H., Mohn, W.W., Indest, K.J., Eltis, L.D.
<2>Genomic and Transcriptomic Studies of an RDX (Hexahydro-1,3,5-Trinitro-1,3,5-Triazine)-Degrading Actinobacterium.
<3>Appl. Environ. Microbiol.
<4>78
<5>7798-7800
<6>2012
<7>Whole genome sequencing, transcriptomic analyses and metabolic reconstruction were used to
investigate Gordonia sp. KTR9s ability to catabolize a range of compounds including explosives
and steroids. Aspects of this mycolic acid-containing actinobacteriums catabolic potential
were experimentally verified and compared with those of rhodococci and mycobacteria.

<>

<1>Chen, J., Herzenberg, L.A., Herzenberg, L.A.
<2>Heparin inhibits EcoRI endonuclease cleavage of DNA at certain EcoRI sites.
<3>Nucleic Acids Res.
<4>18
<5>3255
<6>1990
<7>Studies presented here demonstrate that heparin inhibits EcoRI endonuclease
cleavage of DNA whereas related proteoglycans show no effect.  The inhibition
occurs at particular EcoRI sites that are near or overlap with palindromic
sequences in the murine lambda5 and Lyt-2 genes.  Endogenous heparin from
peritoneal mast cells co-isolates with DNA and inhibits digestion of peritoneal
cell DNA at the inhibitable sites.  Digestion of spleen DNA is inhibited at the
same sites when commerical heparin is added prior to digestion.  In both cases,
the inhibition is abolished by pre-treating the DNA with heparinase.  Thus,
potential artifacts in restriction fragment length analyses could occur with
DNA isolated either from cells that are naturally rich in heparin or from cells
to which heparin has been added, e.g., as an anticoagulant.

<>

<1>Chen, J., Huang, H., Chang, C.J., Stenger, D.C.
<2>Draft Genome Sequence of Xylella fastidiosa subsp. multiplex Strain Griffin-1 from Quercus rubra in Georgia.
<3>Genome Announcements
<4>1
<5>e00756-13
<6>2013
<7>The draft genome sequence of Xylella fastidiosa subsp. multiplex strain Griffin-1, isolated
from a red oak tree (Quercus rubra) in Georgia, is reported
here. The bacterium has a genome size of 2,387,314 bp, with a G+C content of
51.7%. The Griffin-1 strain genome contains 2,903 predicted open reading frames
and 50 RNA genes.

<>

<1>Chen, J., Xia, Y., Cheng, C., Fang, C., Shan, Y., Jin, G., Fang, W.
<2>Genome sequence of a non-pathogenic Listeria monocytogenes serovar 4a strain M7.
<3>J. Bacteriol.
<4>193
<5>5019-5020
<6>2011
<7>This report presents the complete and annotated genome sequence of a naturally non-pathogenic
Listeria monocytogenes serovar 4a strain M7, isolated from cow's milk in Zhejiang province,
China.

<>

<1>Chen, J., Xie, G., Han, S., Civerolo, E.L.
<2>Two whole genome sequences of Xylella fastidiosa (strains M12 and M23) causing almond leaf scorch disease in California.
<3>J. Bacteriol.
<4>192
<5>4534
<6>2010
<7>Xylella fastidiosa is a Gram negative plant pathogenic bacterium causing many economically
important diseases including almond leaf scorch disease
(ALSD) in California. Genome information greatly facilitates research in
this nutritionally fastidious organism. Here we report the complete genome
sequences of two ALSD strains, M12 and M23 of this bacterium.

<>

<1>Chen, J.H., Zhang, J., Yang, H.H., Fu, F.F., Chen, G.N.
<2>A strategy for development of electrochemical DNA biosensor based on site-specific DNA cleavage of restriction endonuclease.
<3>Biosensors and Bioelectronics
<4>26
<5>144-148
<6>2010
<7>A new strategy for development of electrochemical DNA biosensor based on site-specific DNA
cleavage of restriction endonuclease and using
quantum dots as reporter was reported in this paper The biosensor was
fabricated by immobilizing a capture hairpin probe, thiolated single
strand DNA labeled with biotin group, on a gold electrode BfuCl
nuclease, which is able to specifically cleave only double strand DNA
but not single strand DNA, was used to reduce background current and
Improve the sensitivity We demonstrated that the capture hairpin probe
can be cleaved by BfuCl nuclease in the absence of target DNA, but
cannot be cleaved in the presence of target DNA. The difference before
and after enzymatic cleavage was then monitored by electrochemical
method after the quantum dots were dissolved from the hybrids Our
results suggested that the usage of BfuCl nuclease obviously Improved
the sensitivity and selectivity of the biosensor. We successfully
applied this method to the sequence-selective discrimination between
perfectly matched and mismatched target DNA including a single-base
mismatched target DNA, and detected as low as 3 3 x 10(-14) M of
complementary target DNA Furthermore, our above strategy was also
verified with fluorescent method by designing a fluorescent molecular
beacon (MB), which combined the capture hairpin probe and a pair of
fluorophore (TAMRA) and quencher (DABCYL) The fluorescent results are
consistent with that of electroanalysis, further indicating that the
proposed new strategy indeed works as we expected.

<>

<1>Chen, J.W., Chan, K.G.
<2>Genome Sequence of Dyella japonica Strain A8, a Quorum-Quenching Bacterium That Degrades N-Acylhomoserine Lactones, Isolated from Malaysian Tropical Soil.
<3>J. Bacteriol.
<4>194
<5>6331
<6>2012
<7>Dyella japonica strain A8 is a Malaysian tropical soil bacterial strain which shows
N-acylhomoserine lactone-degrading activity. Here, we present its draft
genome sequence. A putative quorum-quenching gene was identified based on the
genome sequence analysis of strain A8. To the best of our knowledge, this is the
first genome announcement of a member from the genus of Dyella, and this is also
the first work that reports the quorum-quenching activity of Dyella japonica.

<>

<1>Chen, J.W., Gan, H.M., Yin, W.F., Chan, K.G.
<2>Genome Sequence of Roseomonas sp. Strain B5, a Quorum-Quenching N-Acylhomoserine  Lactone-Degrading Bacterium Isolated from Malaysian Tropical Soil.
<3>J. Bacteriol.
<4>194
<5>6681-6682
<6>2012
<7>Roseomonas sp. strain B5 was isolated from Malaysian tropical soil that showed
N-acylhomoserine lactone degradation. This is the first genome announcement of a
member from the genus of Roseomonas and the first report on the quorum-quenching
activity of Roseomonas spp.

<>

<1>Chen, K., Reuter, M., Sanghvi, B., Roberts, G.A., Cooper, L.P., Tilling, M., Blakely, G.W., Dryden, D.T.F.
<2>ArdA proteins from different mobile genetic elements can bind to the EcoKI Type I DNA methyltransferase of E. coli K12.
<3>Biochim. Biophys. Acta
<4>1844
<5>505-511
<6>2014
<7>Anti-restriction and anti-modification (anti-RM) is the ability to prevent cleavage by DNA
restriction-modification (RM) systems of foreign DNA entering a new bacterial host. The
evolutionary consequence of anti-RM is the enhanced dissemination of mobile genetic elements.
Homologues of ArdA anti-RM proteins are encoded by genes present in many mobile genetic
elements such as conjugative plasmids and transposons within bacterial genomes. The ArdA
proteins cause anti-RM by mimicking the DNA structure bound by Type I RM enzymes. We have
investigated ArdA proteins from the genomes of Enterococcus faecalis V583, Staphylococcus
aureus Mu50 and Bacteroides fragilis NCTC 9343, and compared them to the ArdA protein
expressed by the conjugative transposon Tn916. We find that despite having very different
structural stability and secondary structure content, they can all bind to the EcoKI
methyltransferase, a core component of the EcoKI Type I RM system. This finding indicates that
the less structured ArdA proteins become fully folded upon binding. The ability of ArdA from
diverse mobile elements to inhibit Type I RM systems from other bacteria suggests that they
are an advantage for transfer not only between closely-related bacteria but also between more
distantly related bacterial species.

<>

<1>Chen, K., Roberts, G.A., Stephanou, A.S., Cooper, L.P., White, J.H., Dryden, D.T.F.
<2>Fusion of GFP to the M.EcoKI DNA methyltransferase produces a new probe of Type I DNA restriction and modification enzymes.
<3>Biochem. Biophys. Res. Commun.
<4>398
<5>254-259
<6>2010
<7>We describe the fusion of enhanced green fluorescent protein to the C-terminus of the HsdS DNA
sequence-specificity subunit of the Type I
DNA modification methyltransferase M.EcoKI. The fusion expresses well
in vivo and assembles with the two HsdM modification subunits. The
fusion protein functions as a sequence-specific DNA methyltransferase
protecting DNA against digestion by the EcoKI restriction endonuclease.
The purified enzyme shows Forster resonance energy transfer to
fluorescently-labelled DNA duplexes containing the target sequence and
to fluorescently-labelled ocr protein, a DNA mimic that binds to the
M.EcoKI enzyme. Distances determined from the energy transfer
experiments corroborate the structural model of M.EcoKI.

<>

<1>Chen, L., Brugger, K., Skovgaard, M., Redder, P., She, Q., Torarinsson, E., Greve, B., Awayez, M., Zibat, A., Klenk, H.-P., Garrett, R.A.
<2>The genome of Sulfolobus acidocaldarius, a model organism of the Crenarchaeota.
<3>J. Bacteriol.
<4>187
<5>4992-4999
<6>2005
<7>Sulfolobus acidocaldarius is an aerobic thermoacidophilic crenarchaeon which grows optimally
at 80 degrees C and pH 2 in terrestrial solfataric springs.  Here, we describe the genome
sequence of strain DSM639, which has been used for many seminal studies on archaeal and
crenarchaeal biology.  The circular genome carries 2,225,959 bp (37% G+C) with 2,292 predicted
protein-encoding genes.  Many of the smaller genes were identified for the first time on the
basis of comparison of three Sulfolobus genome sequences.  Of the protein-coding genes, 305
are exclusive to S.  acidocaldarius and 866 are specific to the Sulfolobus genus.  Moreover,
82 genes for untranslated RNAs were identified and annotated.  Owing to the probable absence
of active autonomous and nonautonomous mobile elements, the genome stability and organization
of S.  acidocaldarius differ radically from those of Sulfolobus solfataricus and Sulfolobus
tokodaii.  The S.  acidocaldarius genome contains an integrated, and probably encaptured,
pARN-type conjugative plasmid which may facilitate intercellular chromosomal gene exchange in
S.  acidocaldarius.  Moreover, it contains genes for a characteristic restriction modification
system, a UV damage excision repair system, thermopsin, and an aromatic ring dioxygenase, all
of which are absent from genomes of other Sulfolobus species.  However, it lacks genes for
some of their sugar transporters, consistent with it growing on a more limited range of carbon
sources.  These results, together with the many newly identified protein-coding genes for
Sulfolobus, are incorporated into a public Sulfolobus database which can be accessed at
http://dac.molbio.ku.dk/dbs/Sulfolobus.

<>

<1>Chen, L., Chavda, K.D., Fraimow, H.S., Mediavilla, J.R., Melano, R.G., Jacobs, M.R., Bonomo, R.A., Kreiswirth, B.N.
<2>Complete nucleotide sequence of blaKPC-4 and blaKPC-5 harboring IncN and IncX plasmids from Klebsiella pneumoniae strains isolated in New Jersey.
<3>Antimicrob. Agents Chemother.
<4>57
<5>269-276
<6>2013
<7>Klebsiella pneumoniae carbapenemase (KPC) producing Enterobacteriaceae have
emerged as major nosocomial pathogens. bla(KPC), commonly located on Tn4401, is
found in Gram-negative bacterial strains, with the two most common variants,
bla(KPC-2) and bla(KPC-3), identified in plasmids with diverse genetic
backgrounds. In this study, we examined bla(KPC-4) and bla(KPC-5) bearing
plasmids recovered from two K. pneumoniae strains, isolated from a single New
Jersey hospital in 2005 and 2006, respectively. IncN plasmid pBK31551 is 84kb in
length, and harbors bla(KPC-4), bla(TEM-1), qnrB2, aac (3)-Ib, aph(3')-I, qacF,
qacEDelta1, sul and dfrA14, which confer resistance to ss-lactams, quinolones,
aminoglycosides, quaternary ammonium compounds and co-trimoxazole. The conserved
regions within pBK31551 are similar to that of other IncN plasmids. Surprisingly,
analysis of the Tn4401 sequence revealed a large IS110 and Tn6901carrying element
(8.3kb) inserted into the istA gene, encoding glyoxalase/bleomycin resistance,
alcohol dehydrogenase and S-formylglutathione hydrolase. Plasmid pBK31567 is 47kb
in length, and harbors bla(KPC-5), dfrA5, qacEDelta1 and sul1. pBK31567 belongs
to a novel IncX subgroup (IncX5), and possesses a highly syntenic plasmid
backbone like other IncX plasmids; however, sequence similarity at the nucleotide
level is divergent. The bla(KPC-5) gene is carried on a Tn4401 element, and
differs from the genetic environment of bla(KPC-5) described in Pseudomonas
aeruginosa strain P28 from Puerto Rico. This study underscores the genetic
diversity of multidrug resistant plasmids involved in the spread of bla(KPC)
genes, and highlights the mobility and plasticity of Tn4401. Comparative genomic
analysis provides new insights into the evolution and dissemination of KPC
plasmids belonging to different incompatibility groups.

<>

<1>Chen, L., Chavda, K.D., Melano, R.G., Hong, T., Rojtman, A.D., Jacobs, M.R., Bonomo, R.A., Kreiswirth, B.N.
<2>Molecular Survey of the Dissemination of Two blaKPC-Harboring IncFIA Plasmids in New Jersey and New York Hospitals.
<3>Antimicrob. Agents Chemother.
<4>58
<5>2289-2294
<6>2014
<7>Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae strains have
spread worldwide and become a major threat in health care facilities.
Transmission of blaKPC, the plasmid-borne KPC gene, can be mediated by clonal
spread and horizontal transfer. Here, we report the complete nucleotide sequences
of two novel blaKPC-3-harboring IncFIA plasmids, pBK30661 and pBK30683. pBK30661
is 74 kb in length, with a mosaic plasmid structure; it exhibits homologies to
several other plasmids but lacks the plasmid transfer operon (tra) and the origin
of transfer (oriT) that are required for plasmid transfer. pBK30683 is a
conjugative plasmid with a cointegrated plasmid structure, comprising a 72-kb
element that highly resembles pBK30661 (>99.9% nucleotide identities) and an
extra 68-kb element that harbors tra and oriT. A PCR scheme was designed to
detect the distribution of blaKPC-harboring IncFIA (pBK30661-like and
pBK30683-like) plasmids in a collection of clinical Enterobacteriaceae isolates
from 10 hospitals in New Jersey and New York. KPC-harboring IncFIA plasmids were
found in 20% of 491 K. pneumoniae isolates, and all carried blaKPC-3.
pBK30661-like plasmids were identified mainly in the epidemic sequence type 258
(ST258) K. pneumoniae clone, while pBK30683-like plasmids were widely distributed
in ST258 and other K. pneumoniae sequence types and among non-K. pneumoniae
Enterobacteriaceae species. This suggests that both clonal spread and horizontal
plasmid transfer contributed to the dissemination of blaKPC-harboring IncFIA
plasmids in our area. Further studies are needed to understand the distribution
of this plasmid group in other health care regions and to decipher the origins of
pBK30661-like and pBK30683-like plasmids.

<>

<1>Chen, L., Chavda, K.D., Melano, R.G., Jacobs, M.R., Koll, B., Hong, T., Rojtman, A.D., Levi, M.H., Bonomo, R.A., Kreiswirth, B.N.
<2>Comparative Genomic Analysis of KPC-Encoding pKpQIL-Like Plasmids and Their Distribution in New Jersey and New York Hospitals.
<3>Antimicrob. Agents Chemother.
<4>58
<5>2871-2877
<6>2014
<7>The global spread of Klebsiella pneumoniae carbapenemase (KPC) is predominately
associated with K. pneumoniae strains genotyped as sequence type 258 (ST258). The
first ST258-associated plasmid, pKpQIL, was described in Israel in 2006, but its
history in the northeastern United States remains unknown. Six pKpQIL-like
plasmids from four K. pneumoniae isolates (three ST258 and one ST234), one
Escherichia coli isolate, and one Enterobacter aerogenes isolate, collected from
2003 to 2010 in New York (NY) and New Jersey (NJ) hospitals, were completely
sequenced. The sequences and overall sizes of the six plasmids are highly similar
to those of pKpQIL; the major difference is that five of six NJ/NY strains harbor
blaKPC-2, while pKpQIL contains blaKPC-3. Moreover, a 26.7-kb fragment was
inverted in pKpQIL-234 (from ST234 K. pneumoniae), while a 14.5-kb region was
deleted in pKpQIL-Ec (from ST131 E. coli). PCR screening of 284 other clinical K.
pneumoniae isolates identified 101 (35.6%) harboring pKpQIL-like plasmids from 9
of 10 surveyed hospitals, demonstrating the wide dissemination of pKpQIL in this
region of endemicity. Among the positive isolates, 87.1% were typed as ST258 and
88.1% carried blaKPC-2. The finding of pKpQIL-like plasmid in this study from
strains that predate the initial report of KPC in Israel provides evidence that
pKpQIL may have originated in the United States. Our findings demonstrate that
pKpQIL plasmids are both spreading clonally in ST258 strains and spreading
horizontally to different sequence types and species, further highlighting the
clinical and public health concerns associated with carbapenem resistance.

<>

<1>Chen, L., Hu, H., Chavda, K.D., Zhao, S., Liu, R., Liang, H., Zhang, W., Wang, X., Jacobs, M.R., Bonomo, R.A., Kreiswirth, B.N.
<2>Complete sequence of a KPC-producing IncN multidrug-resistant plasmid from an epidemic Escherichia coli ST131 strain in China.
<3>Antimicrob. Agents Chemother.
<4>58
<5>2422-2425
<6>2014
<7>We report the nucleotide sequence of a novel blaKPC-2-harboring IncNplasmid,
pECN580, from a multidrug-resistant Escherichia coli ST131 isolate recovered from
Beijing, China. pECN580 harbors multiple antimicrobial resistance genes,
including blaKPC-2, blaCTX-M-3, blaTEM-1, aac(6')-Ib-cr, qnrS1, arr-3 and dfrA14
that confer beta-lactam, aminoglycoside, quinolone, rifampin and trimethoprim
resistance. The emergence of blaKPC-2-harboring multidrug-resistant plasmid in
epidemic E. coli ST131 clone poses a significant potential threat for both
community and hospital settings.

<>

<1>Chen, L., Lin, J., Liu, X., Pang, X., Lin, H., Lin, J.
<2>Transposition of IS elements induced by electroporation of suicide plasmid in Acidithiobacillus caldus.
<3>Enzyme Microb. Technol.
<4>53
<5>165-169
<6>2013
<7>Transposition insertional mutagenesis of the insertion sequences (IS elements) was discovered
for the first time in Acidithiobacillus caldus
(A. caldus), when A. caldus MTH-04 hsdM (type I
restriction-modification system M-subunit) mutant was constructed by
electroporation of a suicide plasmid. The IS element, specifically
inserting into hsdM gene, was analyzed, identified, and named ISAtc2.
The transposition frequency of ISAtc2 was ranged from 4% to 7%, and no
reverse mutation occurred in the mutants after 50 generations of
proliferation without selective pressure. These results revealed that
transposition of IS elements on A. caldus chromosome could regulate the
gene expression and metabolic pathways by gene inactivation, gene loss
and gene acquisition. Therefore, the transposition of IS elements in A.
caldus may be an important and unique regulation mechanism for
adaptation to the living condition.

<>

<1>Chen, L., MacMillan, A.M., Chang, W., Ezaz-Nikpay, K., Lane, W.S., Verdine, G.L.
<2>Direct identification of the active-site nucleophile in a DNA (Cytosine-5)-methyltransferase.
<3>Biochemistry
<4>30
<5>11018-11025
<6>1991
<7>The overproduction, purification, and determination of the active-site catalytic nucleophile
of the DNA (cytosine-5)-methyltransferase (DCMtase) enzyme, M.HaeIII are reported. Incubation
of purified M.HaeIII with an oligodeoxynucleotide specifically modified with the
mechanism-based inhibitor 5-fluoro-2'-deoxycytidine [Osterman, D.G., et al. (1988)
Biochemistry 27,5204-5210], in the presence of the cofactor S-adenosyl-L-methionine (AdoMet),
resulted in the formation of a covalent DNA-M.HaeIII complex, which was purified to
homogeneity. Characterization of the intact complex showed it to consist of one molecule of
the FdC-containing duplex oligonucleotide, one molecule of M.HaeIII, and one methyl group
derived from AdoMet. Exhaustive proteolysis, reduction, and alkylation fof the DNA-M.HaeIII
complex led to the isolation of two DNA-bound peptides-one each from treatment with Pronase or
trypsin-which were subjected to peptide sequencing in order to identify the DNA attachment
site. Both peptides were derived from the region of M.HaeIII containing a Pro-Cys sequence
that is conserved in all known DCMtases. At the position of this conserved Cys residue
(Cys71), in the sequence of each peptide, was found an unidentified amino acid residue; all
other amino acid residues were in accord with the known sequence. It it thus concluded that
Cys71 of M.HaeIII forms a covalent bond to DNA during catalytic methyl transfer. This finding
represents a direct experimental verification for the hypothesis that the conserved Cys
residue of DCMtases is the catalytic nucleophile [Wu, J.C., & Santi, D.V. (1987) J. Biol.
Chem. 262,4778-4786]. Furthermore, the present studies provide ready access to large
quantities of a homogeneous, covalent protein-DNA complex that is trapped at an intermediate
state in catalysis.

<>

<1>Chen, L., MacMillan, A.M., Verdine, G.L.
<2>Mutational separation of DNA binding from catalysis in a DNA cytosine methyltransferase.
<3>J. Am. Chem. Soc.
<4>115
<5>5318-5319
<6>1993
<7>Despite the substantial progress made toward understanding sequence-specific recognition of
DNA by regulatory proteins, the details of catalysis by DNA-modifying proteins remain poorly
understood. The inherently transient nature of catalytic protein-DNA complexes presents
problems for their characterization by X-ray crystallography or NMR spectroscopy. Catalytic
DNA-binding proteins thus present the challenge of discovering how to stall or subvert the
catalytic process so as to obtain a stable macromolecular complex. In this communication we
report the use of site-direced mutagenesis to separate catalysis from DNA binding in a DNA
(cytosine-5)-methyltransferase (DC-Mtase) enzyme.

<>

<1>Chen, L., Mediavilla, J.R., Oliveira, D.C., Willey, B.M., de Lencastre, H., Kreiswirth, B.N.
<2>Multiplex real-time PCR for rapid Staphylococcal cassette chromosome mec typing.
<3>J. Clin. Microbiol.
<4>47
<5>3692-3706
<6>2009
<7>Rapid identification and typing of methicillin (meticillin)-resistant
Staphylococcus aureus (MRSA) is important for understanding the molecular
epidemiology and evolution of MRSA and offers many advantages for
controlling transmission in both health care and community settings. We
developed a rapid molecular beacon real-time PCR (MB-PCR) assay for
staphylococcal cassette chromosome mec (SCCmec) typing. The design of this
system is based on the established definition of SCCmec types, namely, the
combination of the mec class complex with the ccr allotype. The assay
consists of two multiplex panels, the combination of which results in two
targets (mec class, ccr) for each SCCmec type. MB-PCR panel I targets
mecA, ccrB2, mecI, and the DeltamecR1-IS1272 junction (mec class B); it
can definitively identify SCCmec types II and IV. MB-PCR panel II detects
ccrC, ccrB1, ccrB3, ccrB4, and the DeltamecR1-IS431 junction (mec class
C2) and is therefore capable of identifying SCCmec types I, III, V, and VI
in combination with panel I. The method can also detect the recently
described novel SCCmec type VIII (ccrAB4 with mec class A). Our assay
demonstrated 100% concordance when applied to 162 MRSA strains previously
characterized by traditional SCCmec typing schemes. Four geographically
and temporally diverse S. aureus collections were also successfully
classified by our assay, along with 1,683 clinical isolates comprising
both hospital- and community-associated MRSA and methicillin-susceptible
S. aureus strains. As many as 96 isolates can be classified easily within
3 to 4 h, including DNA isolation, PCR cycling, and analysis. The assay is
rapid, robust, sensitive, and cost-effective, allowing for high-throughput
SCCmec typing of MRSA isolates.

<>

<1>Chen, L., Mediavilla, J.R., Smyth, D.S., Chavda, K.D., Ionescu, R., Roberts, R.B., Robinson, D.A., Kreiswirth, B.N.
<2>Identification of a novel transposon (Tn6072) and a truncated staphylococcal cassette chromosome mec element in methicillin-resistant Staphylococcus aureus ST239.
<3>Antimicrob. Agents Chemother.
<4>54
<5>3347-3354
<6>2010
<7>A novel composite transposon (Tn6072) resembling staphylococcal cassette
chromosome mercury (SCCHg) was identified in a collection of sequence type
(ST) 239 methicillin (meticillin)-resistant Staphylococcus aureus (MRSA)
isolates from Romanian hospitals. Tn6072 is homologous to the 5' region of
SCCHg found in staphylococcal cassette chromosome mec (SCCmec) type III
prototype strain 85/2082 but lacks the characteristic mer operon. SCCHg
has previously been reported to integrate downstream of orfX, at the same
chromosomal location as SCCmec. Tn6072, by contrast, is demarcated by two
IS431 elements, flanked by 8-bp direct repeats, and inserted upstream of
the origin of replication, within an open reading frame homologous to
SAR2700 of S. aureus strain MRSA252. Analysis of a geographically and
temporally diverse collection of 111 strains from the ST239 clonal group
uncovered 11 additional strains harboring Tn6072, demonstrating a
lineage-specific insertion pattern. Complete sequence analysis of the
SCCmec regions of two representative Romanian strains (BK16704, BK16691)
revealed two additional novel structures derived from a type III SCCmec
background. BK16704 possesses an SCCmec 3A.1.4 structure, with an IS256
insertion downstream of the right chromosomal junction. In contrast, the
SCCmec element of BK16691 is truncated downstream of the mec gene complex,
with a 24-kb deletion encompassing the right chromosomal junction and an
inverted downstream IS256 element. This structure, tentatively named
"psiSCCmec16691," confers methicillin resistance but lacks most of the
J1/J2 region, including the ccr gene complex. Taken together, these
findings provide evidence for the continuing evolution of SCC elements, as
well as the ST239 clonal group.

<>

<1>Chen, L., Paulsen, D.B., Scruggs, D.W., Banes, M.M., Reeks, B.Y., Lawrence, M.L.
<2>Alteration of DNA adenine methylase (Dam) activity in Pasteurella multocida causes increased spontaneous mutation frequency and attenuation in mice.
<3>Microbiology
<4>149
<5>2283-2290
<6>2003
<7>Pasteurella multocida is one of the primary bacterial pathogens associated with bovine
respiratory disease (BRD) complex. Relatively few virulence
factors of P. multocida have been characterized, and there is a need for
improved vaccines for prevention of BRD. In other Gram-negative species,
DNA adenine methylase (Dam) regulates the expression of virulence genes,
and appropriate expression of Dam is required for virulence. In this
study, the authors cloned and sequenced the P. multocida A1 dam gene and
demonstrated that it is able to restore Dam function in an Escherichia
coli dam mutant. When P. multocida dam was placed under the control of a
constitutively expressed promoter on a plasmid, it caused an increased
spontaneous mutation rate in P. multocida. In addition, the
plasmid-mediated alteration of Dam production in P. multocida caused it to
be highly attenuated in mice. These findings indicate that appropriate
expression of Dam is required for virulence of P. multocida, which is
believed to be the first report that Dam is required for virulence of a
species in the Pasteurellaceae. Therefore, Dam may function as a virulence
gene regulator in the Pasteurellaceae, similar to previously reported
findings from other Gram-negative species.

<>

<1>Chen, L., Wu, D., Cai, X., Guo, F., Blackall, P.J., Xu, X., Chen, H.
<2>Electrotransformation of Haemophilus parasuis with in vitro modified DNA based on a novel shuttle vector.
<3>Vet. Microbiol.
<4>155
<5>310-316
<6>2012
<7>The objective of the present study was to establish a valid transformation method of
Haemophilus parasuis, the causative agent of
Glasser's disease in pigs, using a novel H. parasuis-Escherichia coli
shuttle vector. A 4.2 kb endogenous plasmid pYC93 was extracted from an
H. parasuis field isolate and completely sequenced. Analysis of pYC93
revealed a region approximately 800 bp showing high homology with the
defined replication origin oriV of pLS88, a native plasmid identified
in Haemophilus ducreyi. Based on the origin region of pYC93, E. coli
cloning vector pBluescript SK(+) and the Tn903 derived kanamycin
cassette, a shuttle vector pSHK4 was constructed by overlapping PCR
strategy. When electroporation of the 15 H. parasuis serovar reference
strains and one clinical isolate SH0165 with pSHK4 was performed, only
one of these strains yielded transformants with an efficiency of 8.5 x
10(2) CFUhlg of DNA. Transformation efficiency was notably increased
(1.3 x 10(5) CFU/mu g of DNA) with vector DNA reisolated from the
homologous transformants. This demonstrated that
restriction-modification systems were involved in the barrier to
transformation of H. parasuis. By utilizing an in vitro DNA
modification method with cell-free extracts of the host H. parasuis
strains, 15 out of 16 strains were transformable. The novel shuttle
vector pSHK4 and the established electrotransformation method
constitute useful tools for the genetic manipulation of H. parasuis to
gain a better understanding of the pathogen.

<>

<1>Chen, L., Zhang, D.T., Zhang, J., Su, Y.A., Zhang, H.
<2>Whole-Genome Sequences of Two Clinical Isolates of Extensively Drug-Resistant Mycobacterium tuberculosis from Zunyi, China.
<3>Genome Announcements
<4>2
<5>e00910-14
<6>2014
<7>Before 2013, 92 countries reported extensively drug-resistant Mycobacterium tuberculosis cases
to the WHO. Here, we announce the genome sequences of two
clinical isolates of extensively drug-resistant tuberculosis (XDR-TB) from Zunyi,
China. The genome sequences are composed of 4,411,507 bp and 4,411,515 bp with
2,210 and 2,071 variants, respectively, when compared to the H37Rv genome.

<>

<1>Chen, L., Zou, G., Cao, X., Rufan, Z.
<2>The synthesis of nine kinds of oligodeoxynucleotide containing the recognition sequence for PstI and their interaction with enzyme.
<3>J. Wuhan Univ.
<4>42
<5>499-506
<6>1996
<7>In this paper, nine kinds of oligodeoxynucleotide containing the recognition sequence of
restriction endonuclease PstI have been synthesized by a solid phase phosphite-triester method
or a combination of chemical and enzymatic methods.  The results of their enzymatic hydrolysis
show that the existence of uridine in the recognition sequence of PstI has little influence on
the cleavage of this strand, but reduces the cleavage rate of its complementary strand; PstI
cleaves its substrate through  single strand cleavage in two steps.  The minimum PstI
substrate is an eight to twelve long oligodeoxynucleotide containing its recognition sequence,
and there is a requirement to the base pair numbers on both sides of PstI recognition
sequence. The two cytidines in the recognition sequence of PstI don't play the same role in
enzymatic hydrolysis.

<>

<1>Chen, M., Lin, L., Zhang, Y., Sun, L., An, Q.
<2>Genome Sequence of Klebsiella oxytoca SA2, an Endophytic Nitrogen-Fixing Bacterium Isolated from the Pioneer Grass Psammochloa villosa.
<3>Genome Announcements
<4>1
<5>e00601-13
<6>2013
<7>Klebsiella oxytoca strain SA2 is an endophytic nitrogen-fixing bacterium isolated from the
pioneer grass Psammochloa villosa, which grows in the moving sand dunes
of Ordos Plateau, China. The SA2 genome sequence provides the genetic background
for understanding its endophytic lifestyle and survival in association with grass
in nitrogen-poor environments.

<>

<1>Chen, M., Yan, Y., Zhang, W., Lu, W., Wang, J., Ping, S., Lin, M.
<2>Complete Genome Sequence of the Type Strain Pseudomonas stutzeri CGMCC 1.1803.
<3>J. Bacteriol.
<4>193
<5>6095
<6>2011
<7>Here we report the complete genome sequence of Pseudomonas stutzeri strain CGMCC 1.1803
(equivalent to ATCC 17588), the type strain of P. stutzeri,
which encodes 4,138 open reading frames on a 4,547,930-bp circular
chromosome. The CGMCC 1.1803 genome contains genes involved in
denitrification, benzoate/catechol degradation, chemotaxis, and other
functions.

<>

<1>Chen, M., Zhu, B., Lin, L., Yang, L., Li, Y., An, Q.
<2>Complete genome sequence of Kosakonia sacchari type strain SP1(T.).
<3>Standards in Genomic Sciences
<4>9
<5>1311-1318
<6>2014
<7>Kosakonia sacchari sp. nov. is a new species within the new genus Kosakonia, which was
included in the genus Enterobacter. K sacchari is a nitrogen-fixing
bacterium named for its association with sugarcane (Saccharum officinarum L.). K
sacchari bacteria are Gram-negative, aerobic, non-spore-forming, motile rods.
Strain SP1(T) (=CGMCC1.12102(T)=LMG 26783(T)) is the type strain of the K
sacchari sp. nov and is able to colonize and fix N2 in association with sugarcane
plants, thus promoting plant growth. Here we summarize the features of strain
SP1(T) and describe its complete genome sequence. The genome contains a single
chromosome and no plasmids, 4,902,024 nucleotides with 53.7% GC content, 4,460
protein-coding genes and 105 RNA genes including 22 rRNA genes, 82 tRNA genes,
and 1 ncRNA gene.

<>

<1>Chen, P., Jeannotte, R., Weimer, B.C.
<2>Exploring bacterial epigenomics in the next-generation sequencing era: a new approach for an emerging frontier.
<3>Trends Microbiol.
<4>22
<5>292-300
<6>2014
<7>Epigenetics has an important role for the success of foodborne pathogen persistence in diverse
host niches. Substantial challenges exist in determining DNA methylation to situation-specific
phenotypic traits. DNA modification, mediated by restriction-modification systems, functions
as an immune response against antagonistic external DNA, and bacteriophage-acquired
methyltransferases (MTase) and orphan MTases - those lacking the cognate restriction
endonuclease - facilitate evolution of new phenotypes via gene expression modulation via DNA
and RNA modifications, including methylation and phosphorothioation. Recent establishment of
large-scale genome sequencing projects will result in a significant increase in genome
availability that will lead to new demands for data analysis including new predictive
bioinformatics approaches that can be verified with traditional scientific rigor. Sequencing
technologies that detect modification coupled with mass spectrometry to discover new adducts
is a powerful tactic to study bacterial epigenetics, which is poised to make novel and
far-reaching discoveries that link biological significance and the bacterial epigenome.

<>

<1>Chen, P., Li, J., Zhao, J., He, L., Zhang, Z.
<2>Differential dependence on DNA ligase of type II restriction enzymes: A practical way toward ligase-free DNA automaton.
<3>Biochem. Biophys. Res. Commun.
<4>353
<5>733-737
<6>2007
<7>DNA computing study is a new paradigm in computer science and biological computing fields. As
one of DNA computing approaches, DNA automaton is
composed of the hardware, input DNA molecule and state transition
molecules. By now restriction enzymes are key hardware for DNA computing
automaton. It has been found that DNA computing efficiency may be
independent on DNA ligases when type IIS restriction enzymes like FokI are
used as hardware. In this study, we compared FokI with four other distinct
enzymes HgaI, BsmFI, BbsI, and BseMII, and found their differential
independence on T4 DNA ligase when performing automaton reactions. Since
DNA automaton is a potential powerful tool to tackle gene relationship in
genomic network scale, the feasible ligase-free DNA automaton may set an
initial base to develop functional DNA automata for various DNA technology
development and implications in genetics study in the near future.

<>

<1>Chen, P., Li, S., Yan, L., Wang, N., Yan, X., Li, H.
<2>Draft Genome Sequence of Bacillus subtilis Type Strain B7-S, Which Converts Ferulic Acid to Vanillin.
<3>Genome Announcements
<4>2
<5>e00025-14
<6>2014
<7>The Bacillus subtilis type strain B7-S was obtained through induction with ferulic acid. Here,
we present the draft genome of strain B7-S, which contains
5,313,924 bp, with a G+C content of 35.8%, 5,135 protein-coding genes, and 40
tRNA-encoding genes.

<>

<1>Chen, Q., Fischer, J.R., Benoit, V.M., Dufour, N.P., Youderian, P., Leong, J.M.
<2>In Vitro CpG Methylation Increases the Transformation Efficiency of Borrelia burgdorferi Strains Harboring the Endogenous Linear Plasmid lp56.
<3>J. Bacteriol.
<4>190
<5>7885-7891
<6>2008
<7>Borrelia burgdorferi is the causative agent of Lyme disease, the most common vector-borne
illness in the Northern hemisphere.
Low-passage-number infectious strains of B. burgdorferi exhibit
extremely low transformation efficiencies-so low, in fact, as to hinder
the genetic study of putative virulence factors. Two putative
restriction-modification (R-M) systems, BBE02 contained on linear
plasmid 25 ( lp25) and BBQ67 contained on lp56, have been postulated to
contribute to this poor transformability. Restriction barriers posed by
other bacteria have been overcome by the in vitro methylation of DNA
prior to transformation. To test whether a methylation-sensitive
restriction system contributes to poor B. burgdorferi transformability,
shuttle plasmids were treated with the CpG methylase M. SssI prior to
the electroporation of a variety of strains harboring different
putative R-M systems. We found that for B. burgdorferi strains that
harbor lp56, in vitro methylation increased transformation by at least
1 order of magnitude. These results suggest that in vitro CpG
methylation protects exogenous DNA from degradation by an
lp56-contained R-M system, presumably BBQ67. The utility of in vitro
methylation for the genetic manipulation of B. burgdorferi was
exemplified by the ease of plasmid complementation of a B. burgdorferi
B31 A3 BBK32 kanamycin-resistant ( B31 A3 BBK32:: Kan(r)) mutant,
deficient in the expression of the fibronectin- and glycosaminoglycan (
GAG)-binding adhesin BBK32. Consistent with the observation that
several surface proteins may promote GAG binding, the B. burgdorferi
B31 A3 BBK32:: Kanr mutant demonstrated no defect in the ability to
bind purified GAGs or GAGs expressed on the surfaces of cultured cells.

<>

<1>Chen, S., Wang, C., Zhao, Z., Yang, Z.L.
<2>Genome Sequence of Halorubrum sp. Strain T3, an Extremely Halophilic Archaeon Harboring a Virus-Like Element.
<3>J. Bacteriol.
<4>194
<5>6608-6609
<6>2012
<7>Halorubrum sp. strain T3, harboring a virus-like element, was isolated from a sample collected
from a solar saltern in Yunnan, China. Several strains of
Halorubrum pleomorphic viruses were reported in this genus recently; however, the
virus-host interaction in haloarchaea remains unclear. To explore this issue,
here we present the genome sequence of Halorubrum sp. strain T3 (3,168,011 bp,
68.48% G+C content).

<>

<1>Chen, S., Wilson, G.G.
<2>Method for producing the AccI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5004691
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the AccI restriction
endonuclease by (1) introducing the restriction endonuclease gene into a host whereby the
restriction gene is expressed; (2) fermenting the host which contains the plasmid encoding and
expressing the AccI restriction endonuclease activity, and (3) purifying the AccI restriction
endonuclease from the fermented host which contains the plasmid encoding and expressing the
AccI restriction endonuclease activity.

<>

<1>Chen, S.-Z., Wilson, G.
<2>Method for producing the AccI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0321267 B
<5>
<6>1995
<7>The present invention relates to clones for the AccI restriction endonuclease and modification
methylase, and to the production of these enzymes from the clones.

<>

<1>Chen, S.-Z., Wilson, G.G.
<2>Method for producing the AccI restriction endonuclease and methylase.
<3>Japanese Patent Office
<4>JP 2703786
<5>
<6>1997
<7>
<>

<1>Chen, S.L., Hung, C.S., Xu, J., Reigstad, C.S., Magrini, V., Sabo, A., Blasiar, D., Bieri, T., Meyer, R.R., Ozersky, P., Armstrong, J.R., Fulton, R.S., Latreille, J.P., Spieth, J., Hooton, T.M., Mardis, E.R., Hultgren, S.J., Gordon, J.I.
<2>Identification of genes subject to positive selection in uropathogenic strains of Escherichia coli: A comparative genomics approach.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>5977-5982
<6>2006
<7>Escherichia coli is a model laboratory bacterium, a species that is widely distributed in the
environment, as well as a mutualist and pathogen in its human hosts. As such, E. coli
represents an attractive organism to study how environment impacts microbial genome structure
and function. Uropathogenic E. coli (UPEC) must adapt to life in several microbial communities
in the human body, and has a complex life cycle in the bladder when it causes acute or
recurrent urinary tract infection (UTI). Several studies designed to identify virulence
factors have focused on genes that are uniquely represented in UPEC strains, whereas the role
of genes that are common to all E. coli has received much less attention. Here we describe the
complete 5,065,741-bp genome sequence of a UPEC strain recovered from a patient with an acute
bladder infection and compare it with six other finished E. coli genome sequences. We searched
3,470 ortholog sets for genes that are under positive selection only in UPEC strains. Our
maximum likelihood-based analysis yielded 29 genes involved in various aspects of cell surface
structure, DNA metabolism, nutrient acquisition, and UTI. These results were validated by
resequencing a subset of the 29 genes in a panel of 50 urinary, periurethral, and rectal E.
coli isolates from patients with UTI. These studies outline a computational approach that may
be broadly applicable for studying strain-specific adaptation and pathogenesis in other
bacteria.

<>

<1>Chen, S.L., Shapiro, L.
<2>Identification of long intergenic repeat sequences associated with DNA methylation sites in Caulobacter crescentus and other alpha-proteobacteria.
<3>J. Bacteriol.
<4>185
<5>4997-5002
<6>2003
<7>A systematic search for motifs associated with CcrM DNA methylation sites revealed four long
(>100-bp) motifs (CIR sequences) present in up to 21
copies in Caulobacter crescentus. The CIR1 and CIR2 motifs exhibit a
conserved inverted repeat organization, with a CcrM site in the center of
one of the repeats.

<>

<1>Chen, T., Li, E.
<2>Structure and function of eukaryotic DNA methyltransferases.
<3>Curr. Top. Dev. Biol.
<4>60
<5>55-89
<6>2004
<7>DNA methylation is a common epigenetic modification found in eukaryotic organisms ranging from
fungi to mammals. Over the past 15 years, a number
of eukaryotic DNA methyltransferases have been identified from various
model organisms. These enzymes exhibit distinct biochemical properties and
biological functions, partly due to their structural differences. The
highly variable N-terminal extensions of these enzymes harbor various
evolutionarily conserved domains and motifs, some of which have been shown
to be involved in functional specializations. DNA methylation has
divergent functions in different organisms, consistent with the notion
that it is a dynamically evolving mechanism that can be adapted to fulfill
various functions. Genetic studies using model organisms have provided
evidence suggesting the progressive integration of DNA methylation into
eukaryotic developmental programs during evolution.

<>

<1>Chen, T., Ueda, Y., Dodge, J.E., Wang, Z., Li, E.
<2>Establishment and maintenance of genomic methylation patterns in mouse embryonic stem cells by Dnmt3a and Dnmt3b.
<3>Mol. Cell. Biol.
<4>23
<5>5594-5605
<6>2003
<7>We have previously shown that the DNA methyltransferases Dnmt3a and Dnmt3b carry out de novo
methylation of the mouse genome during early
postimplantation development and of maternally imprinted genes in the
oocyte. In the present study, we demonstrate that Dnmt3a and Dnmt3b are
also essential for the stable inheritance, or "maintenance," of DNA
methylation patterns. Inactivation of both Dnmt3a and Dnmt3b in embryonic
stem (ES) cells results in progressive loss of methylation in various
repeats and single-copy genes. Interestingly, introduction of the Dnmt3a,
Dnmt3a2, and Dnmt3b1 isoforms back into highly demethylated mutant ES
cells restores genomic methylation patterns; these isoforms appear to have
both common and distinct DNA targets, but they all fail to restore the
maternal methylation imprints. In contrast, overexpression of Dnmt1 and
Dnmt3b3 failed to restore DNA methylation patterns due to their inability
to catalyze de novo methylation in vivo. We also show that
hypermethylation of genomic DNA by Dnmt3a and Dnmt3b is necessary for ES
cells to form teratomas in nude mice. These results indicate that genomic
methylation patterns are determined partly through differential expression
of different Dnmt3a and Dnmt3b isoforms.

<>

<1>Chen, T., Ueda, Y., Xie, S., Li, E.
<2>A novel Dnmt3a isoform produced from an alternative promoter localizes to euchromatin and its expression correlates with active de novo methylation.
<3>J. Biol. Chem.
<4>277
<5>38746-38754
<6>2002
<7>Previous studies have shown that the Dnmt3b gene encodes multiple variants via alternative
splicing. However, only one form of Dnmt3a has been identified so far. We now report the
discovery of a small form of Dnmt3a, termed Dnmt3a2, from both human and mouse. The transcript
encoding Dnmt3a2 is initiated from a downstream intronic promoter. As a result, the Dnmt3a2
protein lacks the N-terminal 223 (human) or 219 (mouse) amino acid residues of the full length
Dnmt3a. Recombinant Dnmt3a2 protein displayed similar cytosine methyltransferase activity as
Dnmt3a in vitro. However, Dnmt3a and Dnmt3a2 exhibited strikingly different subcellular
localization patterns. Unlike Dnmt3a, which was concentrated on heterochromatin, Dnmt3a2
displayed a localization pattern suggestive of euchromatin association. Dnmt3a2 is the
predominant form in embryonic stem cells and embryonal carcinoma cells and can also be
detected from testis, ovary, thymus, and spleen, whereas Dnmt3a is expressed at low levels
ubiquitously. Comparison of human embryonal carcinoma cell lines with breast/ovarian cancer
cell lines indicates that DNMT3A2 expression correlates with high de novo methylation
activity. These findings suggest that Dnmt3a and Dnmt3a2 may have distinct DNA targets and
different functions in development.

<>

<1>Chen, W., Wang, Y., Li, D., Li, L., Xiao, Q., Zhou, Q.
<2>Draft Genome Sequence of Brevibacillus brevis Strain X23, a Biocontrol Agent against Bacterial Wilt.
<3>J. Bacteriol.
<4>194
<5>6634-6635
<6>2012
<7>Brevibacillus brevis X23 is an appropriate biocontrol agent against bacterial wilt caused by
Ralstonia solanacearum. We report herein the draft genome sequence
(6,566,879 bp) and a circular plasmid (6,600 bp) of B. brevis X23, data which may
be helpful for mining the antagonistic activity against R. solanacearum.

<>

<1>Chen, X., Wang, Z.-C.
<2>Plant DNA methyltransferase.
<3>Shengming De Huaxue
<4>29
<5>534-538
<6>2009
<7>The addition of a methyl group to the carbon 5 of cytosine residues is the most common
modification of DNA in higher plants.  Plant DNA methyltransferases can be divided into three
classes based on their linear domain arrangement: DNA methyltransferase,
chromomethyltransferase and domains-rearranged methyltrnasferase.  MET and CMT are responsible
for the maintenance of methylation, and the latter has been found only in plants presently;
DRM appears to be the principal de novo methyltransferase.  In this article, the structure,
function and evolutionary aspects of the main plant DNA methyltransferases are analyzed in
detail.

<>

<1>Chen, X., Zhang, B., Zhang, W., Wu, X., Zhang, M., Chen, T., Liu, G., Dyson, P.
<2>Genome Sequence of Streptomyces violaceusniger Strain SPC6, a Halotolerant Streptomycete That Exhibits Rapid Growth and Development.
<3>Genome Announcements
<4>1
<5>e00494-13
<6>2013
<7>Streptomyces violaceusniger strain SPC6 is a halotolerant streptomycete isolated  from the
Linze desert in China. The strain has a very high growth rate and a
short life cycle for a streptomycete. For surface-grown cultures, the period from
spore germination to formation of colonies with mature spore chains is only 2
days at 37 degrees C. Additionally, the strain is remarkably resistant to
osmotic, heat, and UV stress compared with other streptomycetes. Analysis of the
draft genome sequence indicates that the strain has the smallest reported genome
(6.4 Mb) of any streptomycete. The availability of this genome sequence allows us
to investigate the genetic basis of adaptation for growth in an extremely arid
environment.

<>

<1>Chen, X.H. et al.
<2>Comparative analysis of the complete genome sequence of the plant growth-promoting bacterium Bacillus amyloliquefaciens FZB42.
<3>Nat. Biotechnol.
<4>25
<5>1007-1014
<6>2007
<7>Bacillus amyloliquefaciens FZB42 is a Gram-positive, plant-associated bacterium, which
stimulates plant growth and produces secondary metabolites that suppress soil-borne plant
pathogens. Its 3,918-kb genome, containing an estimated 3,693 protein-coding sequences, lacks
extended phage insertions, which occur ubiquitously in the closely related Bacillus subtilis
168 genome. The B. amyloliquefaciens FZB42 genome reveals an unexpected potential to produce
secondary metabolites, including the polyketides bacillaene and difficidin. More than 8.5% of
the genome is devoted to synthesizing antibiotics and siderophores by pathways not involving
ribosomes. Besides five gene clusters, known from B. subtilis to mediate nonribosomal
synthesis of secondary metabolites, we identified four giant gene clusters absent in B.
subtilis 168. The pks2 gene cluster encodes the components to synthesize the macrolactin core
skeleton.

<>

<1>Chen, Y., Chatterjee, S.S., Porcella, S.F., Yu, Y.S., Otto, M.
<2>Complete Genome Sequence of a Panton-Valentine Leukocidin-Negative Community-Associated Methicillin-Resistant Staphylococcus aureus Strain of Sequence type 72 from Korea.
<3>PLoS ONE
<4>8
<5>E72803
<6>2013
<7>In the past decade, community-associated (CA-) infections with
methicillin-resistant Staphylococcus aureus (MRSA) have emerged throughout the
world. Different CA-MRSA strains dominate in different geographical locations.
Many CA-MRSA lineages contain genes coding for the Panton-Valentine leukocidin.
However, the role of this leukotoxin in CA-MRSA pathogenesis is still
controversial. The genome sequences of two key PVL-positive CA-MRSA strains
(USA300, USA400) have been reported, but we lack information on the more recently
found PVL-negative CA-MRSA strains. One such strain is the PVL-negative ST72, the
main cause of CA-MRSA infections in Korea. Here, we report the entire genome
sequence of CA-MRSA ST72 and analyze its gene content with a focus on virulence
factors. Our results show that this strain does not have considerable differences
in virulence factor content compared to other CA-MRSA strains (USA300, USA400),
indicating that other toxins do not substitute for the lack of PVL in ST72. This
finding is in accordance with the notion that differential expression of
widespread virulence determinants, rather than the acquisition of additional
virulence factors on mobile genetic elements, such as PVL, is responsible for the
increased virulence of CA- compared to hospital-associated MRSA.

<>

<1>Chen, Y., Cui, Y., Pu, F., Jiang, G., Zhao, X., Yuan, Y., Zhao, W., Li, D., Liu, H., Li, Y., Liang, T., Xu, L., Wang, Y., Song, Q., Yang, J., Liang, L., Yang, R., Han, L., Song, Y.
<2>Draft genome sequence of an Acinetobacter genomic species 3 strain harboring a blaNDM-1 gene.
<3>J. Bacteriol.
<4>194
<5>204-205
<6>2011
<7>Here we report the draft genome sequence of one Acinetobacter genomic species 3 strain, D499,
which harbors the blaNDM-1 gene.  The total length of the assembled genome is 4,103,824 bp,
and 3,896 coding sequences (CDSs) were predicted within the genome.  A previously unreported
blaNDM-1-bearing plasmid was identified in this strain.

<>

<1>Chen, Y., He, Y., Zhang, B., Yang, J., Li, W., Dong, Z., Hu, S.
<2>Complete Genome Sequence of Alicyclobacillus acidocaldarius Strain Tc-4-1.
<3>J. Bacteriol.
<4>193
<5>5602-5603
<6>2011
<7>Alicyclobacillus acidocaldarius strain Tc-4-1 was initially isolated from a hot spring in
Tengchong, China. This organism is both thermophilic and
acidophilic. It can produce heat- and acid-stable enzymes, such as amylase
and esterase, which may be important in industry. Here we report the whole
genome sequence of the strain.

<>

<1>Chen, Y., Ji, Y.J., Roxby, R., Conrad, C.
<2>In vivo expression of single-stranded DNA in mammalian cells with DNA enzyme sequences targeted to C-raf.
<3>Antisense Nucleic Acid Drug Dev.
<4>10
<5>415-422
<6>2000
<7>The use of antisense oligodeoxynucleotides (AS-ODN) remains a viable method to downregulate
selected gene function.  However, limitations to the antisense approach remain, such as (1)
difficulties in delivery of the
As-ODN into target tissues, (2) instability of As-ODN in vivo, (3) uncertainties about the
precise mode of action, and (4) toxic effects in animal and human studies.  To circumvent some
of these difficulties, we designed a vector set that directs the in vivo production of
single-stranded DNA (ssDNA) of a desired target sequence with limited extraneous vector
nucleotide sequences.  One plasmid was designed to express Moloney murine leukemia virus
(MoMuLV) reverse transcriptase (RT).  Another expression plasmid contains the MoMuLV primer
binding site at the 3' end of its RNA transcript so that an ssDNA would be synthesized by RT
when both plasmids are cotransfected into cells.  To test this expression system, we
constructed a plasmid set, pssXA/pssXB that produces ssRNA-cleaving DNA 10-23 enzyme (Santoro,
S.W. and Joyce, G.F. (1977) Proc. Natl. Acad. Sci. USA 37, 13330-13342).  The DNA enzyme
sequence was placed between two oligonucleotide arms that are complementary and able to
specifically target C-raf kinase mRNA.  These plasmids were transfected into the A549 lung
carcinoma cell line.  Reduced C-raf mRNA levels by up to 34%-36%, as determined by Northern
blot analysis, were observed in the transfected cells.  Our results demonstrate the
feasibility of using this novel ssDNA expression system to generate any sequence of interest
in vivo for antisense, RNA-cleavage DNA enzyme, or triplex-forming strategies.

<>

<1>Chen, Y., Mukherjee, S., Hoffmann, M., Kotewicz, M.L., Young, S., Abbott, J., Luo, Y., Davidson, M.K., Allard, M., McDermott, P., Zhao, S.
<2>Whole Genome Sequencing of a Gentamicin-Resistant Campylobacter coli isolated from United States Retail Meats Reveals Novel Plasmid Mediated Aminoglycoside Resistance Genes.
<3>Antimicrob. Agents Chemother.
<4>57
<5>5398-5405
<6>2013
<7>Aminoglycoside resistance in Campylobacter has been routinely monitored in the
United States in clinical isolates since 1996 and in retail meats since 2002.
Gentamicin resistance first appeared in a single human isolate of Campylobacter
coli in 2000 and a single chicken meat isolate in 2007, after which it increased
rapidly to account for 11.3% human isolates and 12.5% of retail isolates in 2010.
Pulsed-field gel electrophoresis analysis indicated that gentamicin resistant C.
coli isolates from retail meat were clonal. We sequenced the genomes of two
strains in this clone using a next generation sequencing technique in order to
investigate the genetic basis for the resistance. The gaps of one strain were
closed using optical mapping and Sanger sequencing, and it is the first completed
genome of C. coli. The two genomes are highly similar to each other. A
self-transmissible plasmid carrying multiple antibiotics resistance genes was
revealed within both genomes, encoding genes resistant to gentamicin, kanamycin,
streptomycin, streptothricin, and tetracycline. Bioinformatics analysis and
experimental results showed that gentamicin resistance was due to a
phosphotransferase gene aph(2')-Ig not described previously. The phylogenetic
relationship of this newly emerged clone to other Campylobacter sp. was
determined by whole genome single nucleotide polymorphisms (SNPs) and showed that
it clustered with the other poultry isolates and was separated from isolates from
livestock.

<>

<1>Chen, Y., Stine, O.C., Badger, J.H., Gil, A.I., Nair, G.B., Nishibuchi, M., Fouts, D.E.
<2>Comparative Genomic Analysis of Vibrio parahaemolyticus: Serotype Conversion and Virulence.
<3>BMC Genomics
<4>12
<5>294
<6>2011
<7>BACKGROUND: Vibrio parahaemolyticus is a common cause of foodborne
disease. Beginning in 1996, a more virulent strain having serotype O3:K6
caused major outbreaks in India and other parts of the world, resulting in
the emergence of a pandemic. Other serovariants of this strain emerged
during its dissemination and together with the original O3:K6 were termed
strains of the pandemic clone. Two genomes, one of this virulent strain
and one pre-pandemic strain have been sequenced. We sequenced four
additional genomes of V. parahaemolyticus in this study that were isolated
from different geographical regions and time points. Comparative genomic
analyses of six strains of V. parahaemolyticus isolated from Asia and Peru
were performed in order to advance knowledge concerning the evolution of
V. parahaemolyticus; specifically, the genetic changes contributing to
serotype conversion and virulence. Two pre-pandemic strains and three
pandemic strains, isolated from different geographical regions, were
serotype O3:K6 and either toxin profiles (tdh+, trh-) or (tdh-, trh+). The
sixth pandemic strain sequenced in this study was serotype O4:K68.
RESULTS: Genomic analyses revealed that the trh+ and tdh+ strains had
different types of pathogenicity islands and mobile elements as well as
major structural differences between the tdh pathogenicity islands of the
pre-pandemic and pandemic strains. In addition, the results of single
nucleotide polymorphism (SNP) analysis showed that 94% of the SNPs between
O3:K6 and O4:K68 pandemic isolates were within a 141 kb region surrounding
the O- and K-antigen-encoding gene clusters. The "core" genes of V.
parahaemolyticus were also compared to those of V. cholerae and V.
vulnificus, in order to delineate differences between these three
pathogenic species. Approximately one-half (49-59%) of each species' core
genes were conserved in all three species, and 14-24% of the core genes
were species-specific and in different functional categories. CONCLUSIONS:
Our data support the idea that the pandemic strains are closely related
and that recent South American outbreaks of foodborne disease caused by V.
parahaemolyticus are closely linked to outbreaks in India. Serotype
conversion from O3:K6 to O4:K68 was likely due to a recombination event
involving a region much larger than the O-antigen- and K-antigen-encoding
gene clusters. Major differences between pathogenicity islands and mobile
elements are also likely driving the evolution of V. parahaemolyticus. In
addition, our analyses categorized genes that may be useful in
differentiating pathogenic Vibrios at the species level.

<>

<1>Chen, Y., Strain, E.A., Allard, M., Brown, E.W.
<2>Genome sequences of Listeria monocytogenes strains J1816 and J1-220 associated with human outbreaks.
<3>J. Bacteriol.
<4>193
<5>3424-3425
<6>2011
<7>Listeria monocytogenes has caused numerous human outbreaks. Here we report draft genomes of L.
monocytogenes J1816 and J1-220, which belongs to the
epidemic clone II and epidemic clone IV, respectively. Whole genome
sequence analysis of these strains provides a tool for studying the
short-term evolution of these epidemic clones.

<>

<1>Chen, Y., Strain, E.A., Allard, M., Brown, E.W.
<2>Genome Sequence of Cronobacter sakazakii E899, a Strain Associated with Human Illness.
<3>J. Bacteriol.
<4>193
<5>5861
<6>2011
<7>Cronobacter has caused numerous illnesses in neonates, infants, and children. Here we report
the draft genome of Cronobacter sakazakii E899.
Whole-genome sequence analysis of Cronobacter strains provides a tool for
understanding the genomic regions specific to each individual species.

<>

<1>Chen, Y.T., Lauderdale, T.L., Liao, T.L., Shiau, Y.R., Shu, H.Y., Wu, K.M., Yan, J.J., Su, I.J., Tsai, S.F.
<2>Sequencing and comparative genomic analysis of pK29, a 269-kilobase conjugative plasmid encoding CMY-8 and CTX-M-3 beta-lactamases in  Klebsiella pneumoniae.
<3>Antimicrob. Agents Chemother.
<4>51
<5>3004-3007
<6>2007
<7>A 269-kilobase conjugative plasmid, pK29, from a Klebsiella pneumoniae strain was sequenced.
The plasmid harbors multiple antimicrobial
resistance genes, including those encoding CMY-8 AmpC-type and CTX-M-3
extended-spectrum beta-lactamases in the common backbone of IncHI2
plasmids. Mechanisms for dissemination of the resistance genes are
highlighted in comparative genomic analyses.

<>

<1>Chen, Y.T., Liao, T.L., Liu, Y.M., Lauderdale, T.L., Yan, J.J., Tsai, S.F.
<2>Mobilization of qnrB2 and ISCR1 in plasmids.
<3>Antimicrob. Agents Chemother.
<4>53
<5>1235-1237
<6>2009
<7>The DNA sequences of two IncHI2 plasmids, pEC-IMP and pEC-IMPQ, from
metallo-beta-lactamase-producing Enterobacter cloacae clinical isolates
were determined. The two conjugative plasmids are almost identical, but
pEC-IMPQ carries an additional segment containing an orf513 (ISCR1), a
truncated 3' conserved sequence, and a qnrB2. Comparative analyses provide
support for the proposed ISCR1-mediated gene mobilization.

<>

<1>Chen, Y.T., Lin, J.C., Fung, C.P., Lu, P.L., Chuang, Y.C., Wu, T.L., Siu, L.K.
<2>KPC-2-encoding plasmids from Escherichia coli and Klebsiella pneumoniae in Taiwan.
<3>J. Antimicrob. Chemother.
<4>69
<5>628-631
<6>2014
<7>OBJECTIVES: Two plasmids carrying blaKPC-2 isolated from carbapenem-resistant
Escherichia coli (CR-EC) and carbapenem-resistant Klebsiella pneumoniae (CR-KP),
respectively, were completely sequenced. The CR-KP strain was selected from an
outbreak in 2012, and the CR-EC strain was the first blaKPC-2-carrying E. coli
identified in the same carbapenem resistance monitoring programme in Taiwan.
METHODS: Antimicrobial susceptibility tests, multilocus sequence typing (MLST)
and the conjugal transfer of plasmids were performed. Complete sequencing of the
plasmids was performed using a shotgun approach. RESULTS: The CR-EC and CR-KP
strains in this study were determined to be ST410 and ST11, respectively, by
MLST. From CR-EC, we identified a 145 kb conjugative plasmid that carries
blaKPC-2, blaCMY-2, blaCTX-M-3 and blaTEM-1. The plasmid is a chimera composed of
three regions related to IncI, IncN and RepFIC replicons. From CR-KP, we
identified an 86.5 kb plasmid, pKPC-LK30, which carries blaKPC-2 and blaSHV-11.
The plasmid is very similar to two blaKPC-2-carrying IncFIIK plasmids, but lacks
one of the replication origins and cannot conjugate. CONCLUSIONS: The differences
in cross-species transferability of the two plasmids can be explained by genetic
differences between their backbones and could have resulted in the confined
blaKPC-2-carrying CR-KP outbreak in Taiwan. Plasmid pKPC-LKEc is the first
blaKPC-2-carrying plasmid identified from CR-EC in Taiwan. With relatively high
transferability it should be closely monitored.

<>

<1>Chen, Y.T., Peng, H.L., Shia, W.C., Hsu, F.R., Ken, C.F., Tsao, Y.M., Chen, C.H., Liu, C.E., Hsieh, M.F., Chen, H.C., Tang, C.Y., Ku, T.H.
<2>Whole-genome sequencing and identification of Morganella morganii KT pathogenicity-related genes.
<3>BMC Genomics
<4>13
<5>S4
<6>2012
<7>BACKGROUND: The opportunistic enterobacterium, Morganella morganii, which can cause
bacteraemia, is the ninth most prevalent cause of clinical infections in patients at Changhua
Christian Hospital, Taiwan. The KT strain of M. morganii was isolated during postoperative
care of a cancer patient with a gallbladder stone who developed sepsis caused by bacteraemia.
M. morganii is sometimes encountered in nosocomial settings and has been causally linked to
catheter-associated bacteriuria, complex infections of the urinary and/or hepatobiliary
tracts, wound infection, and septicaemia. M. morganii infection is associated with a high
mortality rate, although most patients respond well to appropriate antibiotic therapy. To
obtain insights into the genome biology of M. morganii and the mechanisms underlying its
pathogenicity, we used Illumina technology to sequence the genome of the KT strain and
compared its sequence with the genome sequences of related bacteria. RESULTS: The 3,826,919-bp
sequence contained in 58 contigs has a GC content of 51.15% and includes 3,565 protein-coding
sequences, 72 tRNA genes, and 10 rRNA genes. The pathogenicity-related genes encode
determinants of drug resistance, fimbrial adhesins, an IgA protease, haemolysins, ureases, and
insecticidal and apoptotic toxins as well as proteins found in flagellae, the iron acquisition
system, a type-3 secretion system (T3SS), and several two-component systems. Comparison with
14 genome sequences from other members of Enterobacteriaceae revealed different degrees of
similarity to several systems found in M. morganii. The most striking similarities were found
in the IS4 family of transposases, insecticidal toxins, T3SS components, and proteins required
for ethanolamine use (eut operon) and cobalamin (vitamin B12) biosynthesis. The eut operon and
the gene cluster for cobalamin biosynthesis are not present in the other Proteeae genomes
analysed. Moreover, organisation of the 19 genes of the eut operon differs from that found in
the other non-Proteeae enterobacterial genomes. CONCLUSIONS: This is the first genome sequence
of M. morganii, which is a clinically relevant pathogen. Comparative genome analysis revealed
several pathogenicity-related genes and novel genes not found in the genomes of other members
of Proteeae. Thus, the genome sequence of M. morganii provides important information
concerning virulence and determinants of fitness in this pathogen.

<>

<1>Chen, Y.T., Shu, H.Y., Li, L.H., Liao, T.L., Wu, K.M., Shiau, Y.R., Yan, J.J., Su, I.J., Tsai, S.F., Lauderdale, T.L.
<2>Complete Nucleotide Sequence of pK245, a 98-Kilobase Plasmid Conferring Quinolone Resistance and Extended-Spectrum-beta-Lactamase Activity in a Clinical Klebsiella pneumoniae Isolate.
<3>Antimicrob. Agents Chemother.
<4>50
<5>3861-3866
<6>2006
<7>A plasmid containing the qnrS quinolone resistance determinant and the
gene encoding the SHV-2 beta-lactamase has been discovered from a clinical
Klebsiella pneumoniae strain isolated in Taiwan. The complete 98-kb
sequence of this plasmid, designated pK245, was determined by using a
whole-genome shotgun approach. Transfer of pK245 conferred low-level
resistance to fluoroquinolones in electroporant Escherichia coli epi300.
The sequence of the immediate region surrounding qnrS in pK245 is nearly
identical (>99% identity) to those of pAH0376 from Shigella flexneri and
pINF5 from Salmonella enterica serovar Infantis, the two other
qnrS-carrying plasmids reported to date, indicating a potential common
origin. Other genes conferring resistance to aminoglycosides (aacC2, strA,
and strB), chloramphenicol (catA2), sulfonamides (sul2), tetracycline
(tetD), and trimethoprim (dfrA14) were also detected in pK245. The dfrA14
gene is carried on a class I integron. Several features of this plasmid,
including three separate regions containing putative replicons, a
partitioning-control system, and a type II restriction modification
system, suggest that it may be able to replicate and adapt in a variety of
hosts. Although no critical conjugative genes were detected, multiple
insertion sequence elements were found scattered throughout pK245, and
these may facilitate the dissemination of the antimicrobial resistance
determinants. We conclude that pK245 is a chimera which acquired its
multiple antimicrobial resistance determinants horizontally from different
sources. The identification of pK245 plasmid expands the repertoire of the
coexistence of quinolone and extended-spectrum-beta-lactam resistance
determinants in plasmids carried by various species of the family
Enterobacteriaceae in different countries.

<>

<1>Chen, Y.Z., Mohan, V., Griffey, R.H.
<2>Spontaneous base flipping in DNA and its possible role in methyltransferase binding.
<3>Phys. Rev. E
<4>62
<5>1133-1137
<6>2000
<7>Recent crystallographic studies showed that HhaI and other methyltransferases flip their
target DNA base completely out of a DNA helix.  This base flipping is also a key feature in a
number of other enzyme-catalyzed processes involving DNA.  The mechanism of base flipping by
these enzymes remains elusive.  Based on a full atomic level description of bond rotational
motions we have studied the energetics of flipping a base in a B-DNA duplex in the absence of
the enzyme.  We have also investigated the effect of the restraints from enzyme-distorted DNA
backbone on the movement of a flipped base in several methyltransferase bound DNA crystal
structures.  Our study on crystal B-DNA helices showed that a base could be flipped at an
energy cost close to the enthalpy observed for base pair opening in premelting thermal
fluctuations.  This suggests that spontaneous base flipping in DNA due to thermal fluctuation
may be achieved.  Analysis of several crystal HhaI and HaeIII methyltransferase DNA duplex
structures showed that the enzyme induced DNA backbone distortion severely restricts the
movement of the flipped base, which indicates that during base flipping the backbone needs to
adopt a substantially different conformation than that observed in the x-ray (enzyme-bound)
structures.  Our results suggest the possible role of thermally induced transient base opening
in facilitating recognition and binding of methyltransferases and other enzymes.

<>

<1>Chen, Z., Chang, D., Zou, Y., Su, L., Zhu, Y., Fang, X., Wang, J., Guo, Y., Zhao, J., Li, D., Fang, C., Yang, R., Liu, C.
<2>Genome Sequence of Enterococcus faecium Clinical Isolate LCT-EF128.
<3>J. Bacteriol.
<4>194
<5>4765
<6>2012
<7>Enterococcus faecium, an opportunistic human pathogen that inhabits the gastrointestinal
tracts of most mammals, has emerged as an important
opportunistic nosocomial pathogen and is a prominent cause of multiresistant
nosocomial infections. Here, we report the draft genome sequence of strain
LCT-EF128, isolated from clinical specimens.

<>

<1>Chen, Z., Gui, J., Gao, X., Pei, C., Hong, Y., Zhang, Q.
<2>Genome architecture changes and major gene variations of Andrias davidianus ranavirus (ADRV).
<3>Vet. Res.
<4>44
<5>101
<6>2013
<7>Ranaviruses are emerging pathogens that have led to global impact and public
concern. As a rarely endangered species and the largest amphibian in the world,
the Chinese giant salamander, Andrias davidianus, has recently undergone
outbreaks of epidemic diseases with high mortality. In this study, we isolated
and identified a novel ranavirus from the Chinese giant salamanders that
exhibited systemic hemorrhage and swelling syndrome with high death rate in China
during May 2011 to August 2012. The isolate, designated Andrias davidianus
ranavirus (ADRV), not only could induce cytopathic effects in different fish cell
lines and yield high viral titers, but also caused severely hemorrhagic lesions
and resulted in 100% mortality in experimental infections of salamanders. The
complete genome of ADRV was sequenced and compared with other sequenced amphibian
ranaviruses. Gene content and phylogenetic analyses revealed that ADRV should
belong to an amphibian subgroup in genus Ranavirus, and is more closely related
to frog ranaviruses than to other salamander ranaviruses. Homologous gene
comparisons show that ADRV contains 99%, 97%, 94%, 93% and 85% homologues in RGV,
FV3, CMTV, TFV and ATV genomes respectively. In addition, several variable major
genes, such as duplicate US22 family-like genes, viral eukaryotic translation
initiation factor 2 alpha gene and novel 75L gene with both motifs of nuclear
localization signal (NLS) and nuclear export signal (NES), were predicted to
contribute to pathogen virulence and host susceptibility. These findings confirm
the etiologic role of ADRV in epidemic diseases of Chinese giant salamanders, and
broaden our understanding of evolutionary emergence of ranaviruses.

<>

<1>Chen, Z., Kong, H.
<2>Isolation and characterization of restriction endonuclease BstYI from Bacillus stearothermophilus Y406.
<3>FEBS Lett.
<4>234
<5>169-171
<6>1988
<7>BstYI, an isoschizomer of XhoII and MflI, has been purified from Bacillus
stearothermophilus Y406.  This enzyme recognized 5' Pu^GATCPy 3' in DNA and
cleaved between Pu and G in this sequence.  BstYI can be easily isolated and
purified by heparin-agarose column chromatography in a high yield (8000 units
BstYI can be obtained per g wet wt of cells).

<>

<1>Chen, Z., Kong, H., Wang, L.
<2>Isolation and characterization of restriction endonucleases BstFI and BstSI from Bacillus stearothermophilus.
<3>J. Fudan Univ. (Natural Sci.)
<4>28
<5>363-366
<6>1989
<7>Two type II restriction endonucleases, BstFI and BstSI, have been isolated from
Bacillus stearothermophilus FH58 and Bacillus stearothermophilus S183.  The
recognition sequence and cleavage site of BstFI is A/AGCTT; and C/PyCGPuG in
BstSI.  Therefore, BstFI is the isoschizomer of HindIII and BstSI is the
isoschizomer of AvaI.  These two enzymes can be easily purified with
heparin-agarose.  10,000 units of BstFI and 24,000 units of BstSI can be
purified per gram wet cell of FH58 or S183, respectively.  They have different
thermostability.  BstFI and BatSI are stable under incubation at 45C for as
long as 6 hours.  After 1 h incubation at 50C the relative activity of BstFI
was reduced by 50%, whereas the relative activity of BstSI was reduced only by
10% after 1 h incubation at 70C.

<>

<1>Chen, Z., Lei, X., Li, Y., Zhang, J., Zhang, H., Yang, L., Zheng, W., Xu, H., Zheng, T.
<2>Whole-Genome Sequence of Marine Bacterium Phaeodactylibacter xiamenensis Strain KD52, Isolated from the Phycosphere of Microalga Phacodactylum tricornutum.
<3>Genome Announcements
<4>2
<5>e01289-14
<6>2014
<7>Phaeodactylibacter xiamenensis KD52 is a novel bacterium isolated from a culture  of the alga
Phaeodactylum tricornutum in Xiamen, Fujian Province, China. Here, we
present the first draft genome sequence of this strain, which will provide an
opportunity to further understand the functional genes related to signing for
nutrition from the host algae and the molecular mechanisms underlying its
beneficial properties.

<>

<1>Chen, Z., Li, Y., Chang, D., An, L., Guo, Y., Wang, J., Li, T., Wang, Y., Zhang, X., Dai, W., Liu, C.
<2>Draft Genome Sequence of Enterococcus faecium Strain LCT-EF301, Which Shows Changes in Biochemical Metabolism following Space Flight.
<3>Genome Announcements
<4>2
<5>e00103-14
<6>2014
<7>An Enterococcus faecium strain was sent into space on the Shenzhou-VIII mission.  After the
space flight, the strain E. faecium LCT-EF301 was isolated and
sequenced based on the changes to its metabolic properties.

<>

<1>Chen, Z., Wen, F., Sun, N., Zhao, H.
<2>Directed evolution of homing endonuclease I-SceI with altered sequence specificity.
<3>Protein Eng. Des. Sel.
<4>22
<5>249-256
<6>2009
<7>Homing endonucleases recognize specific long DNA sequences and catalyze double-stranded breaks
that significantly stimulate homologous
recombination, representing an attractive tool for genome targeting and
editing. We previously described a two-plasmid selection system that
couples enzymatic DNA cleavage with the survival of host cells, and
enables directed evolution of homing endonucleases with altered cleavage
sequence specificity. Using this selection system, we successfully evolved
mutant I-SceI homing endonucleases with greatly increased cleavage
activity towards a new target DNA sequence that differs from the wild-type
cleavage sequence by 4 bp. The most highly evolved mutant showed a
survival rate approximately 100-fold higher than that of wild-type I-SceI
enzyme. The degree of selectivity displayed by a mutant isolated from one
round of saturation mutagenesis for the new target sequence is comparable
to that of wild-type I-SceI for the natural sequence. These results
highlight the ability and efficiency of our selection system for
engineering homing endonucleases with novel DNA cleavage specificities.
The mutant identified from this study can potentially be used in vivo for
targeting the new cleavage sequence within genomic DNA.

<>

<1>Chen, Z., Wilkins, M.R., Hunter, N., Nadkarni, M.A.
<2>Draft Genome Sequences of Two Clinical Isolates of Lactobacillus rhamnosus from Initial Stages of Dental Pulp Infection.
<3>Genome Announcements
<4>1
<5>e00073-12
<6>2013
<7>Here we report the draft genomic sequences of two clinical isolates of Lactobacillus rhamnosus
from infected dental pulps representing the initial stages of infection of pulp tissue. Based
on 454 FLX+ pyrosequencing, the two clinical isolates infecting vital pulp had a genome length
of 2.9 Mbp with distinct genomic signatures.

<>

<1>Chen, Z., Wojcik, S.F., Welker, N.E.
<2>Genetic analysis of Bacillus stearothermophilus by protoplast fusion.
<3>J. Bacteriol.
<4>165
<5>994-1001
<6>1986
<7>Efficient and reliable protoplasting, regeneration, and fusion techniques were
established for the prototrophic strain Bacillus stearothermophilus NUB36.
Auxotrophic mutants were isolated, and protoplast fusion was used to construct
isogenic mutant strains and for chromosomal mapping.  Markers were mapped using
two-, three-, and four-factor crosses.  The order of the markers was
hom-1-thr-1-his-1-(gly-1 or gly-2)-pur-1-pur-2.  These markers may be analogous
to hom, thrA, hisA, glyC, and purA markers on the Bacillus subtilis chromosome.
No analogous pur-1 marker has been reported in B. subtilis.  The relative
order of three of the markers (hom-1-thr-1-gly-1) was independently confirmed.

<>

<1>Chen, Z., Yang, R., Chen, B., Li, R.
<2>Restriction and modification in a gram-negative thermophilic bacterium and isolation of restriction endonuclease TspAI.
<3>J. Fudan Univ. (Natural Sci.)
<4>28
<5>96-101
<6>1989
<7>A restriction endonuclease TspAI had been isolated from the gram-negative and
flagellate thermophilic bacterium FD230.  TspAI functions upon phage p228 in
terms of restriction and modification.  By cleaving lambda DNA, pBR322DNA and
Phi X174 RFDNA-Hae III fragment, it had been identified as a restriction
endonuclease possessing the same recognition sequence as that of EcoRII, i.e.
the cleavage site is CCA/TGG.  TspAI could be easily isolated and purified.
The enzyme was active over a temperature range of 30-80C.  Moreover, it was
stable at 60C for as long as 30 minutes.

<>

<1>Chen, Z., Zhao, H.
<2>A highly sensitive selection method for directed evolution of homing endonucleases.
<3>Nucleic Acids Res.
<4>33
<5>e154
<6>2005
<7>Homing endonucleases are enzymes that catalyze DNA sequence specific double-strand breaks and
can significantly stimulate homologous recombination at these breaks. These enzymes have great
potential for applications such as gene correction in gene therapy or gene alteration in
systems biology and metabolic engineering. However, homing endonucleases have a limited
natural repertoire of target sequences, which severely hamper their applications. Here we
report the development of a highly sensitive selection method for the directed evolution of
homing endonucleases that couples enzymatic DNA cleavage with the survival of host cells.
Using I-SceI as a model homing endonuclease, we have demonstrated that cells with wild-type
I-SceI showed a high cell survival rate of 80-100% in the presence of the original I-SceI
recognition site, whereas cells without I-SceI showed a survival rate <0.003%. This system
should also be readily applicable for directed evolution of other DNA cleavage enzymes.

<>

<1>Chen, Z.-X., Riggs, A.D.
<2>Self-association of human de novo DNA methyltransferases, DNMT3A and DNMT3B.
<3>Biochem. Cell Biol.
<4>83
<5>566
<6>2005
<7>Proper control of cytosine methylation patterns is critical for mammalian development.  Two de
novo DNA methyltransferases, Dnmt3a and Dnmt3b, are required for the establishment of DNA
methylation patterns in germ cells and early embryos.  The mechanism by which DNMT3A and
DNMT3B carry out de novo methylation remains largely unknown.  Here, using the yeast
two-hybrid system and co-immunoprecipitation analysis, we demonstrate that human DNMT3A and
DNMT3B are capable of self-association.  Deletion analysis revealed that the C-terminal
catalytic domain is responsible for self-interaction.  Since most mutations causing
immunodeficiency, centromeric instability, and facial anomalies syndrome occur in the
catalytic domain of DNMT3B, we investigated whether these mutations can affect the ability of
DNMT3B to self-interact.  Two DNMT3B ICF mutants (H814R and D817G) show reduced ability to
interact with themselves in the yeast two-hybrid system.  In addition, DNMT3A interacts with
DNMT3B through their C-terminal catalytic domains.  Self-association of the catalytic domain
may play a role in regulating activity and function of DNMT3 methyltransferases.

<>

<1>Chen, Z.F., Pan, X.S.
<2>Identification of a new type-II restriction enzyme BsrGI from Bacillus stearothermophilus GR75.
<3>Chinese Sci. Bull.
<4>39
<5>526-528
<6>1994
<7>A new type II restriction endonuclease, BsrGI, has been isolated from Bacillus
stearothermophilus GR75. BsrGI cleaves T7 DNA at 13 sites, Lambda DNA at 5 sites and M13mp19
DNA at one site, but does not cleave pBR322 DNA and pUC19 DNA. The position of restriction
site of BsrGI on M13mp19 DNA was mapped to position 1000 using double digests with restriction
enzymes AvaII and BsrFI. The recognition sequence and cleavage site of BsrGI on M13mp19 DNA
was determined directly by using the dideoxynucleotide chain-termination method with a
synthetic counter-clockwise primer (1190-1175) downstream of the BsrGI restriction site.
Sequencing data show that the recognition sequence and cleavage site of BsrGI is
5'...T/GTACA...3'. Thus this enzyme recognizes the palindromic sequence 5'TGTACA3' and
cleaves between 5'T and G, generating 5'-protruding single-strand ends 5'GTAC3'.

<>

<1>Chen, Z.X., Mann, J.R., Hsieh, C.L., Riggs, A.D., Chedin, F.
<2>Physical and functional interactions between the human DNMT3L protein and members of the de novo methyltransferase family.
<3>J. Cell. Biochem.
<4>95
<5>902-917
<6>2005
<7>The de novo methyltransferase-like protein, DNMT3L, is required for methylation of imprinted
genes in germ cells. Although enzymatically
inactive, human DNMT3L was shown to act as a general stimulatory factor
for de novo methylation by murine Dnmt3a. Several isoforms of DNMT3A
and DNMT3B with development-stage and tissue-specific expression
patterns have been described in mouse and human, thus bringing into
question the identity of the physiological partner(s) for stimulation
by DNMT3L. Here, we used an episome-based in vivo methyltransferase
assay to systematically analyze five isoforms of human DNMT3A and
DNMT3B for activity and stimulation by human DNMT3L. Our results show
that human DNMT3A, DNMT3A2, DNMT3B1, and DNMT3B2 are catalytically
competent, while DNMT3B3 is inactive in our assay. We also report that
the activity of all four active isoforms is significantly increased
upon co-expression with DNMT3L, albeit to varying extents. This is the
first comprehensive description of the in vivo activities of the poorly
characterized human DNMT3A and DNMT3B isoforms and of their functional
interactions with DNMT3L. To further elucidate the mechanism by which
DNMT3L stimulates DNA methylation, we have mapped in detail the domains
that mediate interaction of human DNMT3L with human DNMT3A and DNMT3B.
Our results show that the C-terminus of DNMT3L is the only region
required for interaction with DNMT3A and DNMT3B and that interaction
takes place through the C-terminal catalytic domain of DNMT3A and
DNMT3B. The implications of these findings for the regulation of de
novo methyltransferases and genomic imprinting are discussed.

<>

<1>Cheng, C.K., Au, C.H., Li, L., Nong, W., Law, P.T., Cheung, W.M., Ling, J.M., Kwan, H.S.
<2>Genome Sequences of Salmonella enterica Serotype Typhimurium Blood Clinical Isolate ST4848/06 and Stool Isolate ST1489/06.
<3>Genome Announcements
<4>1
<5>e00823-13
<6>2013
<7>Salmonella enterica serotype Typhimurium human blood strains isolated from outside Africa are
rarely sequenced. Here, we report the draft genome sequences
of two S. Typhimurium clinical strains isolated in the same year, one from blood
and another from stool, in order to gain insights into the genetic basis leading
to invasive diseases.

<>

<1>Cheng, D.-F., Liu, Q., Zhao, X.-L., Dong, Y.-S., Li, Q.
<2>Quantitive study on the inhibitive effect of methylation outside the recognition sequence of restriction endonuclease PvuII on its cleavage activity.
<3>Shengwu Huazue Zazhi
<4>12
<5>36-41
<6>1996
<7>The recombinant plasmid pSV2gpt-SV40 ori-antisense-ras (P1) was constructed. Using this
plasmid as a model, it was confirmed that the DNA methylation outside the recognition sequence
could inhibit the activity of restriction endonuclease PvuII.  Quantitive observation showed
that the activity of the PvuII was reduced by 70 percent due to the DNA methylation outside
the recognition sequence.

<>

<1>Cheng, H., Fang, M.X., Jiang, X.W., Wu, M., Zhu, X.F., Zheng, G., Yang, Z.J.
<2>Draft Genome Sequence of Amphibacillus jilinensis Y1(T), a Facultatively Anaerobic, Alkaliphilic and Halotolerant Bacterium.
<3>Standards in Genomic Sciences
<4>8
<5>491-499
<6>2013
<7>The genus Amphibacillus was established in 1990, and seven additional species were described
in the past two decades. Amphibacillus jilinensis Y1(T) is a
facultatively anaerobic and alkaliphilic bacterium isolated from a soda lake in
China. Here we describe the structural and genetic features of the draft genome
about the type strain Y1(T) (3,831,075 bp, with a G+C content of 37.27%). This is
the first genome report of the Amphibacillus genus.

<>

<1>Cheng, S.-C., Kim, R., King, K., Kim, S.-H., Modrich, P.
<2>Isolation of gram quantities of EcoRI restriction and modification enzymes from an overproducing strain.
<3>J. Biol. Chem.
<4>259
<5>11571-11575
<6>1984
<7>Structural genes for EcoRI restriction endonuclease and modification methylase
have been inserted into the plasmid vector pKC3 (Shimatake, H., and Rosenberg,
M. (1981) Nature (Lond.) 292, 128-132 downstream from the bacteriophage
lambdapL promoter.  Upon induction of pL expression in strains producing a
thermolabile lambda cI857 repressor, synthesis of EcoRI polypeptides is
enhanced to the extent that after 4 h they represent several per cent of the
total cell protein.  Purification of activities overproduced in this manner
yields preparations of endonuclease and methylase which appear identical to
those obtained from conventional sources, with overall yields corresponding to
0.5 to 0.9 g of each enzyme/kg of cell paste.

<>

<1>Cheng, S.-C., Modrich, P.
<2>Positive-selection cloning vehicle useful for overproduction of hybrid proteins.
<3>J. Bacteriol.
<4>154
<5>1005-1008
<6>1983
<7>Plasmid pSCC31 contains the EcoRI endonuclease gene downstream from lambda pL.
It does not yield transformants upon introduction into Escherichia coli unless
the structural integrity of the endonuclease is destroyed.  This makes it
useful as a positive-selection cloning vehicle which can be employed for
regulated overproduction of hybrid proteins.

<>

<1>Cheng, T., Lin, P., Jin, S., Wu, Y., Fu, B., Long, R., Liu, D., Guo, Y., Peng, L., Xia, Q.
<2>Complete Genome Sequence of Bacillus bombysepticus, a Pathogen Leading to Bombyx  mori Black Chest Septicemia.
<3>Genome Announcements
<4>2
<5>e00312-14
<6>2014
<7>Bacillus bombysepticus is a Gram-positive spore-forming bacterium. Here, we announce the first
complete genome sequence of this organism isolated from the
cadavers of silkworm larvae that had been sick. The genome contains a single
circular chromosome and a circular plasmid. Analyses of the B. bombysepticus
genome will provide insights into its pathomechanisms and biology.

<>

<1>Cheng, V.W., Zhang, G., Oyedotun, K.S., Ridgway, D., Ellison, M.J., Weiner, J.H.
<2>Complete Genome of the Solvent-Tolerant Staphylococcus warneri Strain SG1.
<3>Genome Announcements
<4>1
<5>e0003813
<6>2013
<7>Staphylococcus warneri is a Gram-positive bacterium commonly found in human skin  flora. The
genome of a laboratory S. warneri isolate, strain SG1, was sequenced
to explore its mechanism of solvent tolerance and its potential as a chassis for
biofuel production.

<>

<1>Cheng, X.
<2>DNA modification by methyltransferases.
<3>Curr. Opin. Struct. Biol.
<4>5
<5>4-10
<6>1995
<7>Enzymatic methylation of DNA plays important roles in both prokaryotes and eukaryotes.
Structural study of the HhaI DNA methyltransferase has provided considerable insight into the
chemistry of C5-cytosine methylation. The DNA-protein complex reveals a substrate cytosine
flipped out of the double helix during the reaction, and a novel two-loop DNA-binding motif
used for both sequence recognition and flipping the base. Structural comparison of HhaI
C5-cytosine methyltransferase, TaqI N6-adenine methyltransferase, and catechol
O-methyltransferase reveals a common catalytic domain structure, which might be universal
among S-adenosyl-L-methionine (SAM)-dependent methyltransferases.

<>

<1>Cheng, X.
<2>Structure and function of DNA methyltransferases.
<3>Annu. Rev. Biophys. Biomol. Struct.
<4>24
<5>293-318
<6>1995
<7>Perspectives and Overview
DNA Methylation
Types of DNA methylation
C5-cytosine methyltransferases
Catalytic mechanism
HhaI methyltransferase
Crystal Structure of a C5-cytosine Methyltransferase
General protein topology: a two-domain structure
Induced fit in Protein-DNA interactions
Catalytic domain
DNA-Recognition domain
Summary and Discussion
The Catalytic Domain is a Structural Framework for the SAM-dependent Methyltransferases
TaqI methyltransferase
Catechol O-methyltransferase
Common catalytic-domain structure
N6-adenine and N4-cytosine methylation
Summary

<>

<1>Cheng, X., Balendiran, K., Schildkraut, I., Anderson, J.E.
<2>Structure of PvuII endonuclease with cognate DNA.
<3>EMBO J.
<4>13
<5>3927-3935
<6>1994
<7>We have determined the structure of PvuII endonuclease complexed with cognate DNA by X-ray
crystallography. The DNA substrate is bound with a single homodimeric protein, each subunit of
which reveals three structural regions. The catalytic region strongly resembles structures of
other restriction endonucleases, even though these regions have dissimilar primary sequences.
Comparison of the active site with those of EcoRV and EcoRI endonucleases reveals a conserved
triplet sequence close to the reactive phosphodiester group and a conserved acidic pair that
may represent the ligands for the catalytic cofactor Mg2+. The DNA duplex is not significantly
bent and maintains a B-DNA-like conformation. The subunit interface region of the homodimeric
protein consists of a pseudo-three-helix bundle. Direct contacts between the protein and the
base pairs of the PvuII recognition site occur exclusively in the major groove through two
antiparallel beta strands from the sequence recognition region of the protein. Water-mediated
contacts are made in the minor grooves to central bases of the site. If restriction enzymes do
share a common ancestor, as has been proposed, their catalytic regions have been very strongly
conserved, while their subunit interfaces and DNA sequence recognition regions have undergone
remarkable structural variation.

<>

<1>Cheng, X., Balendiran, K., Schildkraut, I., Anderson, J.E.
<2>Crystal stucture of the PvuII restriction endonuclease.
<3>Gene
<4>157
<5>139-140
<6>1995
<7>Crystal structures have now been determined for the R.PvuII restriction endonuclease as a
protein-DNA complex and in apo-form.  The structures indicate how the interaction with DNA
might proceed.

<>

<1>Cheng, X., Blumenthal, R.M.
<2>Finding a basis for flipping bases.
<3>Structure
<4>4
<5>639-645
<6>1996
<7>Rotation of a DNA nucleotide out of the double helix and into a protein binding pocket ('base
flipping') was first observed in the structure of a DNA methyltransferase.  There is now
evidence that a variety of proteins use base flipping in their interactions with DNA.  Though
the mechanism for base flipping is still unclear, we propose a three-step pathway: recognizing
the target site and increasing the interstrand phosphate-phosphate distance nearby, initiating
base flipping by protein invasion of the DNA, and trapping the flipped DNA structure.

<>

<1>Cheng, X., Blumenthal, R.M.
<2>Mammalian DNA methyltransferases: A structural perspective.
<3>Structure
<4>16
<5>341-350
<6>2008
<7>The methylation of mammalian DNA, primarily at CpG dinucleotides, has long been recognized to
play a major role in controlling gene expression, among other functions.  Given their
importance, it is surprising how many basic questions remain to be answered about the proteins
responsible for this methylation and for coordination with the parallel chromatin-marking
system that operates at the level of histone modification.  This article reviews recent
studies on, and discusses the resulting biochemical and structural insights into, the DNA
nucleotide methyltransferase (Dnmt) proteins 1, 3a, 3a2, 3b, and 3L.

<>

<1>Cheng, X., Kumar, S., Klimasauskas, S., Roberts, R.J.
<2>Crystal structure of the HhaI DNA methyltransferase.
<3>Cold Spring Harb. Symp. Quant. Biol.
<4>58
<5>331-338
<6>1993
<7>Structures of M.HhaI with AdoMet and M.HhaI, DNA and AdoHcy are described.

<>

<1>Cheng, X., Kumar, S., Posfai, J., Pflugrath, J.W., Roberts, R.J.
<2>Crystal structure of the HhaI DNA methyltransferase complexed with S-adenosyl-L-methionine.
<3>Cell
<4>74
<5>299-307
<6>1993
<7>The first three-dimensional structure of a DNA methyltranferase is presented. The crystal
structure of the DNA (cytosine-5)-methyltransferase, M.HhaI (recognition sequence: GCGC),
complexed with S-adenosyl-L-methionine has been determined and refined at 2.5 A resolution.
The core of the structure is dominated by sequence motifs conserved among all DNA
(cytosine-5)-methyltransferases, and these are responsible for cofactor binding and
methyltransferase function.

<>

<1>Cheng, X., Kumar, S., Sha, M., Roberts, R.J.
<2>Crystal structure of HhaI DNA methyltransferase complexed with S-adenosyl-L-methionine.
<3>Acta Crystallogr. A
<4>SA49
<5>61
<6>1993
<7>DNA methyltransferases are found in organisms ranging from bacteria to mammals. The DNA
methyltransferase from the bacterium Haemophilus haemolyticus catalyzes the transfer of a
methyl group from S-adenosyl-L-methionine (AdoMet) to C-5 of the internal cytosine in the DNA
sequence GCGC. The three dimensional structure of the M.HhaI-AdoMet complex has been
determined and refined at a resolution of 2.5 A. The structure is the first to be solved for
any DNA methyltransferase as well as being the first for any methyltransferase that utilizes
the ubiquitous methyl donor AdoMet. Due to the conserved nature of
(cytosine-5)-methyltransferases, the information obtained from this structure can be
generalized to the entire family, including the mammalian CpG methyltransferase.

<>

<1>Cheng, X., Roberts, R.J.
<2>AdoMet-dependent methylation, DNA methyltransferases and base flipping.
<3>Nucleic Acids Res.
<4>29
<5>3784-3795
<6>2001
<7>Twenty AdoMet-dependent methyltransferases (MTases) have been characterized structurally by
X-ray crystallography and NMR. These include seven DNA MTases, five RNA MTases, four protein
MTases and four small molecule MTases acting on the carbon, oxygen or nitrogen atoms of their
substrates. The MTases share a common core structure of a mixed seven-stranded beta-sheet
(6|7^5|4|1|2^3|) referred to as an 'AdoMet-dependent MTase fold', with the exception
of a protein arginine MTase which contains a compact consensus fold lacking the antiparallel
hairpin strands (6|7^). The consensus fold is useful to identify
hypothetical MTases during structural proteomics efforts on unannotated proteins. The same
core structure works for very different classes of MTase including those that act on
substrates differing in size from small molecules (catechol or glycine) to macromolecules
(DNA, RNA and protein). DNA MTases use a 'base flipping' mechanism to deliver a specific
base within a DNA molecule into a typically concave catalytic pocket. Base flipping involves
rotation of backbone bonds in double-stranded DNA to expose an out-of-stack nucleotide, which
can then be a substrate for an enzyme-catalyzed chemical reaction. The phenomenon is fully
established for DNA MTases and for DNA base excision repair enzymes, and is likely to prove
general for enzymes that require access to unpaired, mismatched or damaged nucleotides within
base-paired regions in DNA and RNA. Several newly discovered MTase families in eukaryotes (DNA
5mC MTases and protein arginine and lysine MTases) offer new challenges in the MTase field.

<>

<1>Cheng, X.D., Collins, R.C., Jia, D., Khan, S.I., Horton, J.R., Qiu, C., Sawada, K., Yang, Z., Zhang, X.
<2>Structural and functional analysis of methyltransferases.
<3>Cell Res.
<4>13
<5>408
<6>2003
<7>Our work involves structural characterization of AdoMet-dependent methyltransferases (MTases),
including enzymes that covalently modify DNA and histones, a process that controls many
cellular processes by affecting gene expression.  The DNA MTase structure comprises a
seven-stranded beta sheet, flanked by alpha helices to form a doubly wound open aba sandwich,
and is
henceforth referred to as the Class 1 MTase structure.  Many of the known Class I MTases act
on DNA to regulate gene expression, to repair mutations or to protect against bacterial
restriction enzymes.  Initially, it was a mystery as to how MTases acted on nucleotides that
are held inside the DNA duplex by base pairing and stacking -- seemingly inaccessible to the
active site of an enzyme.  In a process termed 'base flipping', the enzyme simply rotates
the target DNA on its flanking phosphodiester bonds such that the base projects into the
catalytic pocket.  Protein arginine MTases (PRMTs) have broad substrates including histones H3
and H4.  PRMT1 is the predominant PRMT in mammalian cells, accounting for 85% of cellular PRMT
activity and is essential for early postimplantation development.  The structure of PRMT1
forms a ring-like dimer, essential for AdoMet binding and enzymatic activity.  The AdoMet
binding domain is a compact version of the Class I MTase fold.  A recently discovered class of
these enzymes is the histone lysine MTase family, whose catalytic activity lies within a
conserved domain, the SET domain.  Using entirely different structural scaffolding, the
SET-domain MTases bind to a kinked AdoMet molecule on the opposite side of a narrow channel
from the target nitrogen of the lysine substrate.  The C-terminal tail of the SET domain forms
a pseudo knot and provides an integral part of the hydrophobic active-site pocket, including
tyrosine residues implicated in the catalytic mechanism.  On the other hand, not all histone
lysine MTases contain the SET domain; the yeast Dot1p histone H3-Lys79 MTase belongs to Class
1 MTases.

<>

<1>Chenoll, E., Codoner, F.M., Silva, A., Martinez-Blanch, J.F., Martorell, P., Ramon, D., Genoves, S.
<2>Draft Genome Sequence of Bifidobacterium animalis subsp. lactis Strain CECT 8145, Able To Improve Metabolic Syndrome In Vivo.
<3>Genome Announcements
<4>2
<5>e00183-14
<6>2014
<7>Bifidobacterium animalis subsp. lactis strain CECT 8145 is able to reduce body fat content and
improve metabolic syndrome biomarkers. Here, we report the draft
genome sequence of this strain, which may provide insights into its safety status
and functional role.

<>

<1>Cherepanova, N.A., Minero, A.S., Rakhimova, A.R., Gromova, E.S.
<2>Mechanism of CpG DNA Methyltransferases M.SssI and Dnmt3a Studied by DNA Containing 2-Aminopurine.
<3>Nucleosides Nucleotides Nucleic Acids
<4>30
<5>619-631
<6>2011
<7>Murine DNA methyltransferases Dnmt3a-CD and M.SssI from Spiroplasma methylate cytosines at CpG
sites.  The role of 6-oxo groups of guanines in DNA methylation by these enzymes has been
studied using DNA substrates, which contained 2-aminopurine at different positions.  Removal
of the 6-oxo group of the guanine located adjacent to the target cytosine in the CpG site
dramatically reduces the stability of the methyltransferase-DNA complexes and leads to a
significant decrease in the methylation.  Apparently, 06 of this guanine is involvd in the
recognition of CpG sites by the enzymes.  Cooperative binding of Dnmt3a-CD to
2-aminopurine-containing DNA and the formation of nonproductive enzyme-substrate complexes
were observed.

<>

<1>Cherepanova, N.A., Zhuze, A.L., Gromova, E.S.
<2>Inhibition of murine DNA methyltransferase Dnmt3a by DNA duplexes containing pyrimidine-2(1H)-one.
<3>Biochemistry
<4>75
<5>1244-1256
<6>2010
<7>Here we studied the inhibition of the catalytic domain of Dnmt3a methyltransferase (Dnmt3a-CD)
by DNA duplexes containing the mechanism-based inhibitor pyrimidine-2(1H)-one (P) instead of
the target cytosine. It has been shown that conjugates of Dnmt3a-CD with P-DNA (DNA containing
pyrimidine-2(1H)-one) are not stable to heating at 65A degrees C in 0.1% SDS. The yield of
covalent intermediate increases in the presence of the regulatory factor Dnmt3L. The
importance of the DNA minor groove for covalent intermediate formation during the methylation
reaction catalyzed by Dnmt3a-CD has been revealed. P-DNA was shown to inhibit Dnmt3a-CD; the
IC50 is 830 nM. The competitive mechanism of inhibition of Dnmt3a-CD by P-DNA has been
elucidated. It is suggested that therapeutic effect of zebularine could be achieved by
inhibition of not only Dnmt1 but also Dnmt3a.

<>

<1>Chernov, A.P., Belichenko, O.A., Rechkunova, N.I., Andreeva, I.S., Repin, V.E., Degtyarev, S.K.
<2>Restriction endonuclease Msp20I production by Micrococcus sp. - application in cloning and mapping.
<3>Russian Patent Office
<4>SU 1806191 A
<5>
<6>1993
<7>
<>

<1>Chernov, A.P., Kaliman, A.V.
<2>Some peculiarities of the antirestriction mechanism of bacteriophage T5.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>1
<5>14-19
<6>1987
<7>Data are cited on the peculiarities of the unique antirestriction mechanism (ARM) of
bacteriophage T5.  Phage T5 is not confined by the restriction systems of the second type,
EcoRI, EcoRII, and EcoRV, although its ARM does not inactivate the restriction endonucleases
of these systems.  There is no modification of phage T5 DNA at the EcoRII, EcoRI, and EcoRV
sites in vivo; consequently, the protection of T5 DNA from the action of restriction
endonucleases is not based on its modification by the ARM.  The ARM of phage T5 protects only
its own DNA from restriction and does not protect foreign DNA (from phage lambda).  Four
recognition sites for restriction endonuclease EcoRV have been mapped in T5 DNA and two sites
for restriction endonuclease EcoRII and three sites for restriction endonuclease HpaI in FST.
It was shown that in FST of phage T5 there are two zones with boundaries in the region of 5%
of the length of T5 DNA.  Introduction of recognition sites for restriction endonucleases by
mutagenesis into the terminal zone of FST leads to a confinement of the mutant T5 phage by the
corresponding restriction system, whereas the presence of sites in the second zone of FST does
not lead to confinement of the phage.  It is suggested that the action of (ARM) of phage T5 is
based on prevention by the antirestriction protein of contact of the T5 DNA with the enzymes
of the host restriction systems.

<>

<1>Chernov, A.V., Lepikhov, K.A., Zheleznaya, L.A., Matvienko, N.I.
<2>Two site-specific endonucleases from thermophilic strain Bacillus species OV.
<3>Biokhimiia
<4>61
<5>1837-1847
<6>1996
<7>Two site-specific endonucleases, BspOVI and BspOVII, were isolated from the thermophilic
strain Bacillus species OV.  The highest activity of both the enzymes was observed at 48 C and
did not depend on the presence of S-adenosyl-L-methionine and ATP.  BspOVI recognizes the
sequence 5'-GACNNN/NNGTC-3' 3'-CTGNN/NNNCAG-5' and cleaves it as shown by the arrows.
Consequently, it is an isoschizomer of Eam1105I and belongs to subclass IIN of endonucleases.
BspOVI has an increased stability during storage.  The enzyme can be used to create T-vectors
for direct cloning of PCR products.  BspOVII recognizes and cleaves the sequence
5'-AT/CGAT-3' 3'-TAGC/TA-5' and, consequently, it is an isoschizomer of ClaI,.  BspOVII is
blocked by methylation of adenine inside the recognition site.

<>

<1>Chernov, A.V., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>A new site-specific endonuclease-methylase from the thermophilic strain of Bacillus species LU11.
<3>Biokhimiia
<4>59
<5>1714-1729
<6>1994
<7>A new site-specific endonuclease, BspLU11III, was isolated and purified to homogeneity from
the thermophilic strain Bacillus species LU11. The enzyme recognizes the 5'-GGGAC-3'
sequence on double-stranded DNA and cleaves it at two places, 10 and 11 nucleotides from the
3'-end of the 5'-GGGAC-3' sequence and 14 and 15 nucleotides from the recognized site along
the complementary strand. In solution the enzyme is a monomer with molecular mass of about 93
kD. In the presence of S-adenosylmethionine the enzyme has methylase activity. The adenine
residue in the recognition site is methylated on one of the DNA strands. The restriction
activity of BspLU11III does not depend on ATP, but is stimulated by 80 microM
S-adenosyl-methionine. Mg2+ is required for restriction activity. Star activity is observed in
the presence of sodium chloride. BspLU11III is not a member of the three main classes of
endonucleases, but has properties similar to type IV site-specific endonucleases.

<>

<1>Chernov, A.V., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>BspLUII III, a bifunctional restriction and modification enzyme from a thermohilic strain Bacillus species LUII.
<3>Nucleic Acids Res.
<4>23
<5>1213-1214
<6>1995
<7>BspLU11III, an isomer of FinI and BsmFI, was found to cleave DNA at two points 10, 11 and 14,
15 bp in the different strands away from the recognition site, and in the presence of SAM it
exhibits the adenine specific methyltransferase activity.

<>

<1>Chernov, A.V., Vollmayr, P., Walter, J., Trautner, T.A.
<2>Masc2, a C5-DNA-methyltransferase from Ascobolus immersus with similarity to methyltransferases of higher organisms.
<3>Biol. Chem.
<4>378
<5>1467-1473
<6>1997
<7>The filamentous fungus Ascobolus immersus represents a eukaryotic model organism to study
genetic phenomena linked to DNA methylation.  Following our previous characterization of a
gene, masc1 from A. immersus, encoding the 'de novo' C5-DNA-methyltransferase, we report
here the identification of a second MTase gene, masc2.  The deduced peptide sequence of Masc2
is similar to previously identified eukaryotic MTases and distinct from Masc1 by having a
large N-terminal domain in addition to the ubiquitous C-terminal catalytic domain.  Following
cloning of the gene, Masc2 was overexpressed and purified.  Masc2 shows MTase activity with
double stranded DNAs.  Structural and biochemical properties of Masc2 suggest that it may
function as a 'maintenance' MTase.  With this finding, A. immersus represents so far the
only eukaryotic organism in which two possibly synergistically operating MTases have been
identified.

<>

<1>Chernov, A.V., Zheleznaya, L.A., Matvienko, N.I.
<2>Site-specific endonuclease BspKT8 from the Thermophile strain Bacillus species KT8.
<3>Biokhimiia
<4>60
<5>1318-1325
<6>1995
<7>A site-specific endonuclease recognizing the sequence 5'-AAGCTT-3' was isolated and purified
to homogeneity from the thermophilic strain Bacillus species KT8.  The enzyme, BspKT8, has
molecular mass 34kD and is a monomeric protein in solution.  The activity of BspKT8 does not
depend on ATP and is not stimulated by S-adenosyl-L-methionine.  The enzyme has the highest
activity in the wide temperature range from 37 to 48oC.  DNA is cleaved as indicated by the
arrows 5'-A/AGCT T-3' 3'-T TCGA/A-5' hence, the enzyme is a class II restriction
endonuclease and an isoschizomer of HindIII.

<>

<1>Chernukhin, V.A., Belichenko, O.A., Tarasova, G.V., Gonchar, D.A., Akishev, A.G., Dedkov, V.S., Mikhnenkova, N.A., Degtyarev, S.K.
<2>Bacteria Arthrobacter oxydans strain - producer of AoxI site-specific endonuclease.
<3>Russian Patent Office
<4>RU 2399663 C
<5>
<6>2010
<7>
<>

<1>Chernukhin, V.A., Boltengagen, A.A., Tarasova, G.V., Dedkov, V.S., Degtyarev, S.K.
<2>New Restriction Endonuclease AluBI from Arthrobacter luteus B - AluI Isoshizomer, nonsensitive to Presence of 5-methylcytosine in the Recognition Sequence AGCT.
<3>Ovchinnikov Bull. Biotechnol. Phys. Chem. Biol.
<4>3
<5>21-27
<6>2007
<7>A new restriction endonuclease AluBI has been discovered and characterized. AluBI is an
isoshizomer of well known restriction endonuclease AluI, which recognizes DNA sequence AGCT.
Unlike AluI, enzyme AluBI is able to cleave DNA when recognition sequence contains
5-methylcytosines. AluBI also cleaves recognition sequence with two methylated cytosines, one
modified at N4 and other at C5 positions. However, new enzyme doesn't cleave DNA with two
N4-methylcytosines in the recognition site.

<>

<1>Chernukhin, V.A., Boltengagen, A.A., Tarasova, G.V., Dedkov, V.S., Degtyarev, S.K.
<2>Bacteria Arthrobacter luteus strain B as a producer of site-specific endonuclease AluBI.
<3>Russian Patent Office
<4>RU 2340670 C
<5>
<6>2008
<7>Anthrobacter leuteus B bacteria strain is obtained, which is a producer of site-specific Alu
BI endonuclease.  Restriction endonuclease recognises and splits both strands of the DNA
5'-AGCT-3' nucleotide sequence, without methylated bases and in such a case, when in the
recognition site, one or two cytosines are methylated in the C5 position or one base is
methylated in the N4 position.

<>

<1>Chernukhin, V.A., Chmuzh, E.V., Tomilova, J.E., Nayakshina, T.N., Dedkov, V.S., Degtyarev, S.K.
<2>Bacterial strain Glacial ice bacterium - producer of GluI site-specific endonuclease.
<3>Russian Patent Office
<4>RU 2322492 C
<5>
<6>2008
<7>Strain of microorganism Glacial ice bacterium is isolated from soil and can be used for
preparing a site-specific endonuclease that recognizes and cleaves both chain of methylated
nucleotide sequence in DNA: 5'-G(m5C)/NG(m5C)-3'.  Use for the invention provides preparing
the novel site-specific endonuclease showing specificity to a methylated sequence in DNA that
can be used in DNA analysis.

<>

<1>Chernukhin, V.A., Chmuzh, E.V., Tomilova, Y.E., Nayakshina, T.N., Gonchar, D.A., Dedkov, V.S., Degtyarev, S.K.
<2>A novel site-specific endonuclease GluI recognizes methylated DNA sequence 5'-G(5mC)^NG(5mC)-3'/3'-(5mC)GN^(5mC)G.
<3>Ovchinnikov Bull. Biotechnol. Phys. Chem. Biol.
<4>3
<5>13-17
<6>2007
<7>A novel site-specific endonuclease GluI from the bacterial strain GL24 has been isolated and
characterized. The enzyme recognizes methylated DNA sequence
5'-G(5mC)^NG(5mC)-3'/3'-(5mC)GN^(5mC)G-5' and cleaves it as it is shown by arrow. Due to
its ability to cleave only modified DNA GluI may be useful for genetic engineering experiments
as well as for determination of DNA methylation status in eucaryotes.

<>

<1>Chernukhin, V.A., Golikova, L.N., Gonchar, D.A., Abdurashitov, M.A., Kashirina, Y.G., Netesova, N.A., Degtyarev, S.K.
<2>M.BstF5I-2 and M.BstF5I-4 DNA methyltransferases from BstF5I restriction-modification system of Bacillus stearothermophilus F5.
<3>Biokhimiia
<4>68
<5>967-975
<6>2003
<7>The BstF5I restriction-modification system from Bacillus stearothermophilus F5 includes four
site-specific DNA methyltransferases,
thus differing from all known restriction-modification systems. Here we
demonstrated for the first time that one bacterial cell can possess two
pairs of methylases with identical substrate specificities (methylases
BstF5I-1 and BstF5I-3 recognize GGATG, whereas methylases BstF5I-2 and
BstF5I-4 recognize CATCC) that modify adenine residues on both DNA
strands. Different chromatographic methods provide homogenous preparations
of methylases BstF5I-2 and BstF5I-4. We estimated the principal kinetic
parameters of the reaction of transfer of methyl group from the donor
S-adenosyl-L-methionine to the recognition site 5;-CATCC-3; catalyzed by
BstF5I-2 and BstF5I-4 DNA [N6-adenine]-methyltransferases from the BstF5I
restriction-modification system.

<>

<1>Chernukhin, V.A., Kashirina, Y.G., Sukhanova, K.S., Abdurashitov, M.A., Gonchar, D.A., Degtyarev, S.K.
<2>Isolation and characterization of biochemical properties of DNA methyltransferase FauIA modifying the second cytosine in the  nonpalindromic sequence 5 '-CCCGC-3 '.
<3>Biokhimiia
<4>70
<5>829-837
<6>2005
<7>A gene encoding DNA methyltransferase (methylase) FauIA of the restriction-modification system
FauI from Flavobaclerium aquatile (recognizing sequence 5'-CCCGC-3') was cloned in pJW
vector. The latter was used for transformation of E. coli RRI cells followed by subsequent
thermoinduction and biomass elaboration. Highly purified DNA methyltransferase FauIA
preparation was obtained using chromatography on different sorbents. The molecular mass of the
isolated enzyme of about 39 kD corresponds to its theoretical value. The enzyme was
characterized by temperature and pH optima of 33 degrees C and pH 7.5, respectively.
Methylation of a synthetic oligonucleotide by FauIA methylase followed by its cleavage with
various restrictases and analysis of the resultant restriction fragments revealed that FauIA
methylase modified the second cytosine residue in the sequence 5'-CCCGC-3'. Kinetic analysis
revealed Km and catalytic constant values of 0.16 mu M and 0.05 min(-1), respectively.

<>

<1>Chernukhin, V.A., Kashirina, Y.G., Tomilova, J.E., Gonchar, D.A., Dedkov, V.S., Mikhnenkova, N.A., Degtyarev, S.K.
<2>New Eight Bases Cutter AbsI from Arthrobacter species Recognizes Palindromic DNA Sequence 5'-CC^TCGAGG-3'.
<3>Ovchinnikov Bull. Biotechnol. Phys. Chem. Biol.
<4>2
<5>29-34
<6>2006
<7>Bacterial strain Arthrobacter species7M06 producing site-specific endonuclease AbsI has been
discovered. AbsI recognizes palindromic octanucleotide DNA sequence 5'-CC^TCGAGG-3' and
hydrolyzes it after second cytosine, producing 5'- sticky ends, which are compatible with
sticky ends after DNA cleavage by restriction endonucleases XhoI (5'-C^TCGAG-3'), PspXI
(5'-VC^TCGAGB-3') and SalI (5'-G^TCGAC-3'). Among all known rare-cutting site-specific
endonucleases AbsI is the only enzyme which has no recognition sequences in standard
substrates Lambda and T7 DNAs and Adenovirus type 2 DNA.

<>

<1>Chernukhin, V.A., Kuznetsov, V.V., Gonchar, D.A., Kashirina, Y.G., Netesova, N.A., Degtyarev, S.K.
<2>Substrate Specificity and Biochemical Properties of M3.BstF5I DNA Methyltransferase from the BstF5I Restriction-Modification System.
<3>Biochemistry
<4>75
<5>63-71
<6>2010
<7>Optimal conditions for DNA methylation by the M3.BstF5I enzyme from Bacillus
stearothermophilus and kinetic parameters of lambda phage DNA
modification and that of a number of oligonucleotide substrates are
established. Comparison of M1.BstF5I and M3.BstF5I kinetic parameters
revealed that with similar temperature optima and affinity for DNA,
M3.BstF5I has nearly fourfold lower turnover number (0.24 min(-1)) and
modifies the hemimethylated recognition site with lower efficiency
under optimal conditions than the unmethylated one. In contrast to
another three methylases of the BstF5I restriction-modification system,
the M3. BstF5I enzyme is able to optionally modify the noncanonical
5'-GGATC-3' DNA sequence with a rate more than one order of magnitude
lower than the methylation rate of the canonical 5'-GGATG-3'
recognition site.

<>

<1>Chernukhin, V.A., Nayakshina, T.N., Abdurashitov, M.A., Tomilova, Y.E., Mezentseva, N.V., Dedkov, V.S., Mikhnenkova, N.A., Gonchar, D.A., Degtyarev, S.K.
<2>A novel restriction endonuclease GlaI recognizes the methylated sequence 5'-G(5mC)^GC-3'.
<3>Biotekhnologiya
<4>0
<5>26-30
<6>2006
<7>A novel restriction endonuclease GlaI from soil bacterium strain GL29 has been isolated and
characterized. The enzyme recognizes the methylated DNA sequence 5'-G(m5C)^GC-3' and cleaves
it as indicated by the arrow. Due to its ability to only cleave modified DNA, GlaI can find a
practical application in genetic engineering experiments as well as in determination of
eukaryotic DNA methylation status.

<>

<1>Chernukhin, V.A., Nayakshina, T.N., Mezentseva, N.V., Tomilova, Y.E., Degtyarev, S.K., Dedkov, V.S.
<2>Strain of glacial ice bacterium I as producer of restriction endonuclease GlaI.
<3>Russian Patent Office
<4>RU 2287012 C
<5>
<6>2006
<7>
<>

<1>Chernukhin, V.A., Nayakshina, T.N., Tarasova, G.V., Golikova, L.N., Akishev, A.G., Dedkov, V.S., Mikhnenkova, N.A., Degtyarev, S.K.
<2>Bacterium Paracoccus carotinifaciens strain 3K as a producer of a new site-specific endonuclease PcsI recognizing methylated DNA.
<3>Russian Patent Office
<4>RU 2377294 C
<5>
<6>2009
<7>
<>

<1>Chernukhin, V.A., Seggewiss, J., Kashirina, Y.G., Gonchar, D.A., Degtyarev, S.K.
<2>Purification and properties of recombinant DNA methyltransferase M2.BstSE of the BstSEI nickase-modification system.
<3>Mol. Biol. (Mosk)
<4>43
<5>8-15
<6>2009
<7>The operon for the Bacillus stearothermophilus SE-589 nickase-modification system (NM.BstSEI,
recognition site 5'-GAGTC-3')
includes two DNA methyltransferase (M.) genes, bstSEIM1 and bstSEIM2.
The gene encoding M2.BstSEI was cloned in pJW and expressed in
Escherichia coli cells. M2.BstSEI was purified by chromatography and
displayed maximal activity at 55A degrees C and pH 7.5. The enzyme
modified adenine in the nickase recognition site 5'-GAGTC-3' and was
specific for 5'-GASTC-3' substrates. The kinetic parameters of the
methylation reaction were determined. The catalytic constant was 2.2
min(-1), and the Michaelis constant was 9.8 nM on T7 DNA and 5.8 mu M
on SAM.

<>

<1>Chernukhin, V.A., Tomilova, J.E., Chmuzh, E.V., Sokolova, O.O., Dedkov, V.S., Degtyarev, S.K.
<2>Bacterial strain Bacillus simplex - producer of BlsI site-specific endonuclease.
<3>Russian Patent Office
<4>RU 2322494 C
<5>
<6>2008
<7>Invention proposes the strain Bacillus simplex 23 isolated from soil and providing preparing
site-specific endonuclease.  This enzyme is able for recognizing and cleaving both chain in
nucleotide sequence of DNA comprising at least one C5-methylcytosine base in the recognition
site 5'-GCNGC-3' to form 3'-prominent ends.  The novel strain can be used for isolation of
the novel site-specific endonuclease that can be used for detection and cleavage of methylated
sites in DNA.

<>

<1>Chernukhin, V.A., Tomilova, J.E., Chmuzh, E.V., Sokolova, O.O., Dedkov, V.S., Degtyarev, S.K.
<2>Site-specific endonuclease BlsI recognizes DNA sequence 5'-G(5mC)N^GC-3' and cleaves it producing 3' sticky ends.
<3>Ovchinnikov Bull. Biotechnol. Phys. Chem. Biol.
<4>3
<5>28-33
<6>2007
<7>A bacterial strain Bacillus simplex 23, a producer of a site-specific endonuclease BlsI has
been discovered. BlsI recognizes the methylated DNA sequence 5'-G(5mC)NGC-3', like the
earlier described site-specific endonuclease BisI (recognition site 5'-G(5mC)�«NGC-3'), but
differs in positions of DNA cleavage producing 3'-protruding ends. Due to its ability to
cleave only methylated DNA, enzyme BlsI can find an application in gene engineering works as
well as in determining the methylation status of eucaryotic DNA.

<>

<1>Chernukhin, V.A., Zhuravleva, R.O., Tarasova, G.V., Boltengagen, A.A., Akishev, A.G., Mikhnenkova, N.A., Degtyarev, S.K.
<2>Strain of Kocuria rosea bacteria as producer of site-specific endonucleases KroI.
<3>Russian Patent Office
<4>RU 2394099 C
<5>
<6>2010
<7>Optimal conditions for DNA methylation by the M3.BstF5I enzyme from Bacillus
stearothermophilus and kinetic parameters of lambda phage DNA modification and that of a
number of oligonucleotide substrates are established.  Comparison of M1.BstF5I and M3.BstF5I
kinetic parameters revealed that with similar temperature optima and affinity for DNA,
M3.BstF5I has nearly fourfold lower turnover number (0.24 min-1) and modifies the
hemimethylated recognition site with lower efficiency under optimal conditions than the
unmethylated one.  In contrast to another three methylases of the BstF5I
restriction-modification system, the M3.BstF5I enzyme is able to optionally modify the
noncanonical 5'-GGATC-3' DNA sequence with a rate more than one order of magnitude lower
than the methylation rate of the canonical 5'-GGATG-3' recognition site.

<>

<1>Cherny, D.I., Kurakin, A.V.
<2>Site-selective targeting of duplex DNA with methyltransferase and biotinylated synthetic reagents.
<3>Electron Microsc.
<4>3A
<5>449-450
<6>1994
<7>Site-selective interaction of some ligands with duplex DNA may provide
the tools for developing gene-targeted drugs and for mapping of DNA and genomes.  In
this study we have examined the interaction of methyltransferase and biotinylated both
deoxyoligonucleotides and peptide nucleic acid (PNA) with duplex DNA.  It was shown
that the target sequence can be EM detected via specific complex formation either with the
enzyme per se or streptavidin as an EM marker.  These approaches allow the study  of the
site-selective interaction of these ligands with DNA and introduce novel types of sequences
for mapping of DNA and genomes.

<>

<1>Chertkov, O. et al.
<2>Complete genome sequence of Hirschia baltica type strain (IFAM 1418T).
<3>Standards in Genomic Sciences
<4>5
<5>287-297
<6>2011
<7>The family Hyphomonadaceae within the Alphaproteobacteria is largely comprised of bacte-ria
isolated from marine environments with striking morphologies and an unusual mode of cell
growth. Here, we report the complete genome sequence Hirschia baltica, which is only the
second a member of the Hyphomonadaceae with a published genome sequence. H. bal-tica is of
special interest because it has a dimorphic life cycle and is a stalked, budding bacte-rium.
The 3,455,622 bp long chromosome and 84,492 bp plasmid with a total of 3,222 pro-tein-coding
and 44 RNA genes were sequenced as part of the DOE Joint Genome Institute Program CSP 2008.

<>

<1>Chertkov, O. et al.
<2>Complete genome sequence of Tolumonas auensis type strain (TA 4).
<3>Standards in Genomic Sciences
<4>5
<5>112-120
<6>2011
<7>Tolumonas auensis Fischer-Romero et al. 1996 is currently the only validly named  species of
the genus Tolumonas in the family Aeromonadaceae. The strain is of
interest because of its ability to produce toluene from phenylalanine and other
phenyl precursors, as well as phenol from tyrosine. This is of interest because
toluene is normally considered to be a tracer of anthropogenic pollution in
lakes, but T. auensis represents a biogenic source of toluene. Other than
Aeromonas hydrophila subsp. hydrophila, T. auensis strain TA 4(T) is the only
other member in the family Aeromonadaceae with a completely sequenced type-strain
genome. The 3,471,292 bp chromosome with a total of 3,288 protein-coding and 116
RNA genes was sequenced as part of the DOE Joint Genome Institute Program JBEI
2008.

<>

<1>Chertkov, O. et al.
<2>Complete genome sequence of Thermomonospora curvata type strain (B9).
<3>Standards in Genomic Sciences
<4>4
<5>13-22
<6>2011
<7>Thermomonospora curvata Henssen 1957 is the type species of the genus Thermomonospora. This
genus is of interest because members of this clade are
sources of new antibiotics, enzymes, and products with pharmacological activity.
In addition, members of this genus participate in the active degradation of
cellulose. This is the first complete genome sequence of a member of the family
Thermomonosporaceae. Here we describe the features of this organism, together
with the complete genome sequence and annotation. The 5,639,016 bp long genome
with its 4,985 protein-coding and 76 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Chertkov, O. et al.
<2>Complete genome sequence of Aminobacterium colombiense type strain (ALA-1).
<3>Standards in Genomic Sciences
<4>2
<5>280-289
<6>2010
<7>Aminobacterium colombiense Baena et al. 1999 is the type species of the genus Aminobacterium.
This genus is of large interest because of its isolated
phylogenetic location in the family Synergistaceae, its strictly anaerobic
lifestyle, and its ability to grow by fermentation of a limited range of amino
acids but not carbohydrates. Here we describe the features of this organism,
together with the complete genome sequence and annotation. This is the second
completed genome sequence of a member of the family Synergistaceae and the first
genome sequence of a member of the genus Aminobacterium. The 1,980,592 bp long
genome with its 1,914 protein-coding and 56 RNA genes is part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Chervaux, C., Grimaldi, C., Bolotin, A., Quinquis, B., Legrain-Raspaud, S., van Hylckama, V.J.E., Denariaz, G., Smokvina, T.
<2>Genome Sequence of the Probiotic Strain Bifidobacterium animalis subsp. lactis CNCM I-2494.
<3>J. Bacteriol.
<4>193
<5>5560-5561
<6>2011
<7>Bifidobacterium animalis subsp. lactis CNCM I-2494 is part of a commercialized fermented dairy
product with documented health benefits
revealed by multiple randomized placebo-controlled clinical trials. Here
we report the complete genome sequence of this strain, which has a
circular genome of 1,943,113 bp with 1,660 open reading frames and 4
ribosomal operons.

<>

<1>Chevalier, B., Monnat, R.J. Jr., Stoddard, B.L.
<2>The LAGLIDADG homing endonuclease family.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Berlin Heidelberg
<4>16
<5>33-47
<6>2005
<7>The LAGLIDADG protein family includes the first identified and biochemically characterized
intron-encoded proteins, as described in this volume by Dujon.  It has been variously termed
the "DOD", "Dodecapeptide', "dodecamer", and "decapeptide" endonuclease family, based on the
conservation of a ten-residue sequence motif.  The LAGLIDADG endonucleases are the most
diverse of the homing endonuclease families.  Their host range includes the genomes of plant
and algal chloroplasts, fungal and protozoan mitochondria, bacteria and Archaea.  One reason
for the wide phylogenetic distribution of LAGLIDADG genes appears to be their remarkable
ability to invade unrelated types of intervening sequences, including group I introns,
archaeal introns and inteins.  Descendents of LAGLIDADG homing endonucleases also include the
yeast HO mating type switch endonuclease, which is encoded by an independent reading frame
rather than within an intron, but does carry remnants of an inactive intein domain, and
maturases that assist in RNA splicing.

<>

<1>Chevalier, B., Sussman, D., Otis, C., Noel, A.J., Turmel, M., Lemieux, C., Stephens, K., Monnat, R.J., Stoddard, B.L.
<2>Metal-dependent DNA cleavage mechanism of the I-CreI LAGLIDADG homing endonuclease.
<3>Biochemistry
<4>43
<5>14015-14026
<6>2004
<7>The LAGLIDADG homing endonucleases include free-standing homodimers, pseudosymmetric monomers,
and related enzyme domains embedded within
inteins. DNA-bound structures of homodimeric I-CreI and monomeric
I-SceI indicate that three catalytic divalent metal ions are
distributed across a pair of overlapping active sites, with one shared
metal participating in both strand cleavage reactions. These structures
differ in the precise position and binding interactions of the metals.
We have studied the metal dependence for the I-CreI homodimer using
site-directed mutagenesis of active site residues and assays of binding
affinity and cleavage activity. We have also reassessed the binding of
a nonactivating metal ion (calcium) in the wild-type enzyme-substrate
complex, and determined the DNA-bound structure of two inactive enzyme
mutants. The conclusion of these studies is that the catalytic
mechanism of symmetric LAGLIDADG homing, endonucleases, and probably
many of their monomeric cousins, involves a canonical two-metal
mechanism in each of two active sites, which are chemically and
structurally tethered to one another by a shared metal ion. Failure to
occupy the shared metal site, as observed in the presence of calcium or
when the metal-binding side chain from the LAGLIDADG motif (Asp 20) is
mutated to asparagines, prevents cleavage by the enzyme.

<>

<1>Chevalier, B., Turmel, M., Lemieux, C., Monnat, R.J., Stoddard, B.L.
<2>Flexible DNA target site recognition by divergent homing endonuclease isoschizomers I-CreI and I-MsoI.
<3>J. Mol. Biol.
<4>329
<5>253-269
<6>2003
<7>Homing endonucleases are highly specific catalysts of DNA strand breaks that induce the
transposition of mobile intervening sequences containing
the endonuclease open reading frame. These enzymes recognize long DNA
targets while tolerating individual sequence polymorphisms within those
sites. Sequences of the homing endonucleases themselves diversify to a
great extent after founding intron invasion events, generating highly
divergent enzymes that recognize similar target sequences. Here, we
visualize the mechanism of flexible DNA recognition and the pattern of
structural divergence displayed by two homing endonuclease isoschizomers.
We determined structures of I-CreI bound to two DNA target sites that
differ at eight of 22 base-pairs, and the structure of an isoschizomer,
I-MsoI, bound to a nearly identical DNA target site. This study
illustrates several principles governing promiscuous base-pair recognition
by DNA-binding proteins, and demonstrates that the isoschizomers display
strikingly different protein/DNA contacts. The structures allow us to
determine the information content at individual positions in the binding
site as a function of the distribution of direct and water-mediated
contacts to nucleotide bases, and provide an evolutionary snapshot of
endonucleases at an early stage of divergence in their target specificity.

<>

<1>Chevalier, B.S., Kortemme, T., Chadsey, M.S., Baker, D., Monnat, R.J., Stoddard, B.L.
<2>Design, activity, and structure of a highly specific artificial endonuclease.
<3>Mol. Cell
<4>10
<5>895-905
<6>2002
<7>We have generated an artificial highly specific endonuclease by fusing domains of homing
endonucleases I-Dmol and I-Crel and creating a new 1400
Angstrom(2) protein interface between these domains. Protein engineering
was accomplished by combining computational redesign and an in vivo
protein-folding screen. The resulting enzyme, E-Drel (Engineered
I-Dmol/I-Crel), binds a long chimeric DNA target site with nanomolar
affinity, cleaving it precisely at a rate equivalent to its natural
parents. The structure of an E-Drel/DNA complex demonstrates the accuracy
of the protein interface redesign algorithm and reveals how catalytic
function is maintained during the creation of the new endonuclease. These
results indicate that it may be possible to generate novel highly specific
DNA binding proteins from homing endonucleases.

<>

<1>Chevalier, B.S., Monnat, R.J. Jr., Stoddard, B.L.
<2>The homing endonuclease I-CreI uses three metals, one of which is shared between the two active sites.
<3>Nat. Struct. Biol.
<4>8
<5>312-316
<6>2001
<7>Homing endonucleases, like restriction enzymes, cleave double-stranded DNA at specific target
sites. The cleavage mechanism(s) utilized by
LAGLIDADG endonucleases have been difficult to elucidate; their active
sites are divergent, and only one low resolution cocrystal structure
has been determined, Here we report two high resolution structures of
the dimeric I-CreI homing endonuclease bound to DNA: a substrate
complex with calcium and a product complex with magnesium, The bound
metals in both complexes are verified by manganese anomalous difference
maps, The active sites are positioned close together to facilitate
cleavage across the DNA minor groove; each contains one metal ion
bound between a conserved aspartate (Asp 20) and a single scissile
phosphate. A third metal ion bridges the two active sites. This
divalent cation is bound between aspartate residues from the active
site of each subunit and is in simultaneous contact with the scissile
phosphates of both DNA strands. A metal-bound water molecule acts as
the nucleophile and is part of an extensive network of ordered water
molecules that are positioned by enzyme side chains; These structures
illustrate a unique variant of a two-metal endonuclease mechanism is
employed by the highly divergent LAGLIDADG enzyme family.

<>

<1>Chevalier, B.S., Stoddard, B.L.
<2>Homing endonucleases: structural and functional insight into the catalysts of intron/intein mobility.
<3>Nucleic Acids Res.
<4>29
<5>3757-3774
<6>2001
<7>Homing endonucleases confer mobility to their host intervening sequence, either an intron or
intein, by catalyzing a highly specific double-strand break in a cognate allele lacking the
intervening sequence. These proteins are characterized by their ability to bind long DNA
target sites (14-40 bp) and their tolerance of minor sequence changes in these sites. A wealth
of biochemical and structural data has been generated for these enzymes over the past few
years. Herein we review our current understanding of homing endonucleases, including their
diversity and evolution, DNA-binding and catalytic mechanisms, and attempts to engineer them
to bind novel DNA substrates.

<>

<1>Chi, M.-G.
<2>Isolation and purification of EcoRI restriction endonuclease.
<3>Junshi Yixue Kexueyuan Yuankan
<4>19
<5>132-135
<6>1995
<7>EcoRI restriction endonuclease was isolated from Escherichia coli RY13 and K121100.  The
purification process involved treatment with ammonium sulfate, polyethyleneimine and
phosphocellulose chromatography.  By this method a highly purified EcoRI restriction
endonuclease can be prepared.  The enzyme recognizes unique sites on DNA, with the minimal
recognition site being a hexanucleotide sequence characterized by 2-fold symmetry.  Thus, the
EcoRI enzyme is one of the simplest sequence-specific DNA enzymes known and is well suited to
a study of DNA sequence recognition by proteins.

<>

<1>Chiara, M., D'Erchia, A.M., Manzari, C., Minotto, A., Montagna, C., Addante, N., Santagada, G., Latorre, L., Pesole, G., Horner, D.S., Parisi, A.
<2>Draft Genome Sequences of Six Listeria monocytogenes Strains Isolated from Dairy  Products from a Processing Plant in Southern Italy.
<3>Genome Announcements
<4>2
<5>e00282-14
<6>2014
<7>Here we announce the draft genome sequences of 6 Listeria monocytogenes strains from ricotta
cheese produced in a dairy processing plant located in southern Italy and potentially involved
in a multistate outbreak of listeriosis in the United States.

<>

<1>Chien, H.R.
<2>Cloning and genetic analysis of restriction and modification systems from Neisseria gonorrhoeae.
<3>Ph.D. Thesis, University of Maryland, USA
<4>
<5>1-126
<6>1991
<7>Endonucleases with the specificities of S.NgoI and S.NgoIII were isolated from Neisseria
gonorrhoeae MS11; but the remaining endonucleases with the specificities of S.NgoII, S.NgoIV,
S.NgoV, S.NgoVI, S.NgoVIII, and S.NgoIX were not produced at detectable levels in MS11. The
genes encoding the RM NgoMI (recognition sequence GCCGGC), the RM NgoMII (TCACC), and the RM
NgoMIII (CCGCGG) from gonococcus MS11 were cloned in E. coli. The gene encoding the M.NgoBIII
(GGNNCC) from gonococcus MUG116 was also cloned. The cloned endonuclease R.NgoMI was purified
by column chromatography and the cutting site determined to be G/CCGGC. The base methylated in
the sequence GCCGGC by the M.NgoMI in vivo was determined to be the the first C in the
sequence GCCGGC. Plasmids encoding M.NgoMI, M.NgoMIII, and M.NgoBIII were restricted when
these plasmids were used to transform MM294 (mcr+), due to the action of the mcr gene products
on the methylated DNA. Plasmids encoding for M.NgoMII were not restricted because the mcr gene
products do not recognize S.NgoMII specificity. The DNA sequence of the NgoMI
restriction/methylation system was determined and two open reading frames were identified. The
molecular weight of the purified endonuclease was determined to be 33,000 Dal, and was in good
agreement with the size predicted from the DNA sequence (31,759 Dal). M.NgoMI contains the
four conserved domains found in cytosine-specific DNA methylases, however the size and spacing
between these domains are significantly different from that in the conserved domains in other
cytosine-specific DNA methylases. Two restriction/modification mutants MUG700 and MUG701 were
derived from MS11. Mutant MUG700 was RM NgoII negative, while mutant MUG701 was RM NgoMI
negative, as determined by appropriate endonuclease digestion and Southern hybridizations. The
RM NgoII and RM NgoMI systems seem involved in regulating the biosynthesis of LOS in N.
gonorrhoeae. The RM NgoMI system also seems involved in regulating the stability of colony
morphology. Finally, endonuclease R.NgoII was able to mediate host restriction in N.
gonorrhoeae, while endonuclease R.NgoMI was not, even though both endonucleases were not
produced at detectable levels in the strain MS11. [ The enzyme called NgoMI in this abstract
has been renamed NgoMIV, Jan/1998. ] [ The enzyme called NgoMII in this abstract has been
renamed NgoMVIII, Jan/1998. ]

<>

<1>Chien, M. et al.
<2>The genomic sequence of the accidental pathogen Legionella pneumophila.
<3>Science
<4>305
<5>1966-1968
<6>2004
<7>We present the genomic sequence of Legionella pneumophila, the bacterial agent of
Legionnaires' disease, a potentially fatal pneumonia acquired from aerosolized contaminated
fresh water. The genome includes a 45-kilobase pair element that can exist in chromosomal and
episomal forms, selective expansions of important gene families, genes for unexpected
metabolic pathways, and previously unknown candidate virulence determinants. We highlight the
genes that may account for Legionella's ability to survive in protozoa, mammalian
macrophages, and inhospitable environmental niches and that may define new therapeutic
targets.

<>

<1>Chien, R.H., Stein, D.C., Seifert, H.S., Floyd, K., So, M.
<2>Cloning and characterization of a restriction and modification system from Neisseria gonorrhoeae strain MS11.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>88
<5>213
<6>1988
<7>A type II restriction and modification (R/M) system from Neisseria gonorrhoeae MS11 was cloned
and expressed in Escherichia coli HB101.  R/M clones were constructed by the partial digestion
of DNA from MS11 with MboI and HpaII and its insertion into the BamHI and ClaI site of the
cloning vector pHSS7.  The R/M clone, pCBB49, was identified by its resistance to cleavage
with NaeI, a restriction enzyme that has 1 site in the cloning vector.  The R/M genes were
encoded by a 2.5 kb fragment in plasmid pCBB49.1, a deletion derivative of pCBB49.  The
restriction (NgoMI) and modification (M.NgoMI) enzymes were purified from HB101 (pCBB49.1) by
column chromatography.  The recognition sequence for NgoMI was GCCGGC.  When plasmid pCBB49.1
was introduced into HB101 (pFT180), the single NgoMI site of pFT180 was now resistant to
cleavage by NaeI.  When pCBB49.1 was used to transform E. coli strains ER1563 (mcrB+) and
ER1562 (mcrB), a two log difference in transformation frequencies was obtained due to the
action of the mcrB gene product on the methylated DNA.
[ The enzyme called NgoMI in this abstract has been renamed NgoMIV, Jan/1998. ]

<>

<1>Chies, J.M., de O-Dias, A.C., Maia, H.M.M., Astolfi-Filho, S.
<2>BanAI a new isoschizomer of the type II restriction endonuclease HaeIII discovered in a Bacillus anthracis isolate from Amazon Basin.
<3>FEMS Microbiol. Lett.
<4>215
<5>97-101
<6>2002
<7>Bacillus anthracis was isolated and identified from a bacterial collection of samples from the
Amazon river bank.  Type II restriction endonuclease activity was detected in this prokaryote,
the enzyme was purified, the molecular mass of the native protein estimated by gel filtration,
and optima pH, temperature and salt requirements were determined.  Quality control assays
showed complete absence of 'non-specific nucleases'.  Restriction cleavage analysis and DNA
sequencing of restriction fragments allowed unequivocal demonstration of 5'-GG/CC-3' as the
recognition sequence.  This enzyme was named BanAI and is therefore an isoschizomer of the
prototype restriction endonuclease HaeIII.  This is the first report of a type II restriction
endonuclease identified, purified from a natural isolate of B. anthracis.

<>

<1>Chies, J.M., Dias, A.C.D.O., Maia, H.M.M., Braga, L.P.D.S., Astolfi-Filho, S.
<2>Isolation and characterization of Bliai, an isoschizomer of CLAI from Bacillus licheniformis.
<3>Braz. J. Microbiol.
<4>37
<5>551-555
<6>2006
<7>The restriction endonuclease BliAI, an isoschizomer of ClaI, which recognizes the sequence
5'-AT down arrow CGAT-3', was purified from a
natural isolate identified as Bacillus licheniformis. The restriction
endonuclease was isolated from cell extracts using single-step
purification by phosphocellulose column chromatography. The restriction
endonuclease is active at 37 degrees C and over a wide range of pH and
salt concentration. The molecular weight of the purified restriction
enzyme is consistent with a value of 39 kDa.

<>

<1>Chik, J., Robins, L., Stoddard, B.
<2>Expression and purification of Ltr-II: A homing endonuclease for gene targeting.
<3>ACS Abstracts, , , San Diego
<4>243
<5>475
<6>2012
<7>Homing endonucleases are microbial enzymes that selectively form double-stranded breaks in DNA
and drive gene conversion events via homologous recombination.  One type of homing
endonuclease conaining the LADLIDADG catalytic motif recognizes long DNA sequences wqith high
specificity and has successfully been used for targeting genes involved in both medicine
(genetic diseases) and biotechnology.  LtrII is an intron-encoded LAGLIDADG homing
endonuclease from the fungus Leptogaphium truncatum.  The results of the overexpression and
purificaiton of LtrII will be shown.  In addition, initial kinetic results of LtrII with its
target DNA sequence will be presented.

<>

<1>Chikaev, N.A., Baklanov, M.M., Ryazankin, I.A., Nechaev, Y.S.
<2>Purification of restriction endonucleases using aqueous two-phase systems.
<3>Prikl. Biokhim. Mikrobiol.
<4>23
<5>84-92
<6>1987
<7>A simple procedure was developed for testing and purification of restriction
endonucleases MspI, PstI, BamHI, PvuI, PvuII that includes biomass destruction,
fractionation of cell-free extracts in the aqueous two-phase (polyethylene
glycol-dextran) system and chromatography of phosphocellulose.  Optimal
conditions for the fractionation of MspI, PstI, BamHI, PvuII, EcoRI, EcoRII,
BspRI, AluI were chosen.  For separation of PvuI and PvuII gel filtration
through Biogel A-0.5 m was additionally introduced.

<>

<1>Chikaev, N.A., Nazarenko, I.A., Matskova, L.V., Boykov, Y.H., Malygin, E.G.
<2>Purification of restriction enzyme BamHI and exonuclease III from Bacillus amyloliquefaciens.
<3>Soviet Patent Office
<4>SU 1661211 A
<5>
<6>1991
<7>The present invention provides the method of purification of the BamHI restriction
endonuclease and exonuclease III by 1) disruption of cells from Bacillus amyloliquefaciens; 2)
chromatography of the crude extract on phospocellulose and DEAE-cellulose; 3) chromatography
on hydroxylapatite and Bio-Rex 70 for BamHI and chromatography on aminohexylsepharose and
DNA-cellulose for exonuclease III. The yield was 2.5 mg BamHI restriction endonuclease with
specific activity of approximately 500,000 units/mg and 40 mg exonuclease III with specific
activity of 1,350 units/mg from 100g cells.

<>

<1>Chilley, P.M., Wilkins, B.M.
<2>Distribution of the ardA family of antirestriction genes on conjugative plasmids.
<3>Microbiology
<4>141
<5>2157-2164
<6>1995
<7>The ardA gene of I1 plasmid ColIb-P9 was previously shown to alleviate DNA restriction by type
I enzymes and to promote conjugative transmission of the unmodified plasmid to a restricting
host. To clarify the ecological role of ardA, its distribution was determined on plasmids from
23 incompatibility groups using hybridization to the coding sequence as an assay. Hybridizing
sequences, shown by nucleotide sequencing to have at least 60% identity with ardA, were
detected on plasmids belonging to the I complex (IncB, I1 and K), the F complex (IncFV) and
the IncN group. The ardA homologues were found to specify an antirestriction phenotype which
was enhanced by genetic depression of the plasmid transfer system. ardA loci map in plasmid
leading regions but show no consistent association with a particular type of
origin-of-transfer or a leading region gene of the ssb (single-stranded DNA-binding protein),
psiB (plasmid SOS inhibition) and hok (host killing) families. It may be significant that
ardA+ plasmids are authentic enterobacterial plasmids and that type I restriction systems are
associated historically with members of the Enterobacteriaceae.

<>

<1>Chin, V., Valinluck, V., Magaki, S., Ryu, J.
<2>KpnBI is the prototype of a new family (IE) of bacterial type I restriction-modification system.
<3>Nucleic Acids Res.
<4>32
<5>e138
<6>2004
<7>KpnBI is a restriction-modification (R-M) system recognized in the GM236 strain of Klebsiella
pneumoniae. Here, the KpnBI modification genes were
cloned into a plasmid using a modification expression screening method.
The modification genes that consist of both hsdM (2631 bp) and hsdS (1344
bp) genes were identified on an 8.2 kb EcoRI chromosomal fragment. These
two genes overlap by one base and share the same promoter located upstream
of the hsdM gene. Using recently developed plasmid R-M tests and a
computer program RM Search, the DNA recognition sequence for the KpnBI
enzymes was identified as a new 8 nt sequence containing one degenerate
base with a 6 nt spacer, CAAANNNNNNRTCA. From Dam methylation and HindIII
sensitivity tests, the methylation loci were predicted to be the
italicized third adenine in the 5' specific region and the adenine
opposite the italicized thymine in the 3' specific region. Combined with
previous sequence data for hsdR, we concluded that the KpnBI system is a
typical type I R-M system. The deduced amino acid sequences of the three
subunits of the KpnBI system show only limited homologies (25 to 33%
identity) at best, to the four previously categorized type I families (IA,
IB, IC, and ID). Furthermore, their identity scores to other
uncharacterized putative genome type I sequences were 53% at maximum.
Therefore, we propose that KpnBI is the prototype of a new 'type IE'
family.

<>

<1>Chin, V.R., Valinluck, V., Ryu, J.
<2>The KpnBI restriction-modification (R-M) system of Klebsiella pneumoniae strain GM236 is a prototype of a new family of type I R-M  systems.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>404-405
<6>2001
<7>From genetic evidence, the KpnBI R-M system in Klebsiella pneumoniae strain GM236 was assumed
to be either a type I or type III R-M system.
The restriction subunit (hsdR) was previously cloned and sequenced.
However, it did not show any close homology to the preexisting R-M
systems. In this project the modification genes of the KpnBI system
were cloned into a plasmid, pMECA, using a modification expression
screening method. The modification genes were identified on an 8.2 kb
EcoRI chromosomal fragment. The complete 8.2 kb fragment was sequenced
and two open reading frames (ORF) were identified. The first ORF is
2,631 bp in length and identified as the hsdM gene based on sequence
homologies. Similarily, the second ORF, which was 1,344 bp in length
and overlapped 1 base with the hsdM gene, was identified as the hsdS
gene. The presence of both hsdM and hsdS genes is a unique
characteristic of type I R-M systems. Therefore, the KpnBI system was
concluded to be a type I system. Phage and plasmid modification tests
as well as complementation tests with the HsdR subunit indicate that
the modification genes are expressed in both Klebsiella and E. coli
strains. Since the deduced amino acid sequences of all three subunits
of the KpnBI did not show sufficient (more than 40%) homologies with
any of the four categorized type I families (IA, IB, IC, and ID) we
concluded that KpnBI represents a new type I family. This family was
subsequently labeled type IE. The DNA recognition sequence of KpnBI
system has also been elucidated using a plasmid transformation method
currently developed in our laboratory. A further analysis of the inter-
and intra-family protein sequence comparisons of the subunits of type I
enzymes has allowed us to develop a criteria for grouping all the type
I systems into more than 20 new type I families in addition to the 4
existing families.

<>

<1>Chinen, A., Naito, Y., Handa, N., Kobayashi, I.
<2>Evolution of sequence recognition by restriction-modification enzymes: selective pressure for specificity decrease.
<3>Mol. Biol. Evol.
<4>17
<5>1610-1619
<6>2000
<7>Several type II restriction-modification (RM) gene complexes kill host bacterial cells that
have lost them, through attack on the chromosomal recognition sites of these cells. Two RM
gene complexes recognizing the same sequence cannot simultaneously enjoy such stabilization
through post-segregational host killing, because one will defend chromosomal sites from attack
by the other. In the present work, we analyzed intrahost competition between two RM gene
complexes when the recognition sequence of one was included in that of the other. When the
EcoRII gene complex, recognizing 5'-CCWGG (W = A, T), is lost from the host, the SsoII gene
complex, which recognizes 5'-CCNGG (N = A, T, G, C), will prevent host death by protecting
CCWGG sites on the chromosome. However, when the SsoII (CCNGG) gene complex is lost, the
EcoRII (CCWGG) gene complex will be unable to prevent host death through attack by SsoII on
5'-CCSGG (S = C, G) sites. These predictions were verified in our experiments, in which we
analyzed plasmid maintenance, cell growth, cell shape, and chromosomal DNA. Our results
demonstrate the presence of selective pressure for decrease in the specificity of recognition
sequence of RM systems in the absence of invading DNA.

<>

<1>Chinen, A., Naito, Y., Handa, N., Kobayashi, I.
<2>Evolution of sequence recognition by restriction-modification enzymes: Selective pressure for specificity decrease.
<3>Genes Genet. Syst.
<4>75
<5>381
<6>2000
<7>Several type II restriction-modification (RM) gene complexes kill host bacterial cells that
have lost them, through attack on their chromosomal recognition sites.  Two RM gene complexes
recognizing the same sequence cannot simultaneously enjoy such stabilization through
post-segregational host killing, because one will defend chromosomal sites from attack by the
other.  We analyzed intra-host competition between two RM gene complexes when the recognition
sequence of one is included in that of the other.  When the EcoRII gene complex, recognizing
5'CCWGG (W = A, T) is lost from the host, the SsoII gene complex, which recognizes 5'CCNGG
(N = A, T, G, C) will prevent host death by protecting CCWGG sites on the chromosome.
However, when the SsoII (CCNGG) gene complex is lost, the EcoRII (CCWGG) gene complex will be
unable to prevent host death through attack by SsoII on 5'CCSGG (S = C, G) sites.  These
predictions were verified in our experiments in which we analyzed plasmid maintenance, cell
growth, cell shape and chromosomal DNA.  Our results demonstrate the presence of selective
pressure for decrease in the specificity of recognition sequence of RM systems in the absence
of invading DNA.

<>

<1>Chinen, A., Uchiyama, I., Kobayashi, I.
<2>Comparison between Pyrococcus horikoshii and Pyrococcus abyssi genome sequences reveals linkage of restriction-modification genes with large genome polymorphisms.
<3>Gene
<4>259
<5>109-121
<6>2000
<7>Recent work suggests that restriction-modification gene complexes are mobile genetic elements
that insert themselves into the genome and cause various genome rearrangements. In the present
work, the complete genome sequences of Pyrococcus horikoshii and Pyrococcus abyssi, two
species in a genus of hyperthermophilic archaeon (archaebacterium), were compared to detect
large genome polymorphisms linked with restriction-modification gene homologs. Sequence
alignments, GC content analysis, and codon usage analysis demonstrated the diversity of these
homologs and revealed a possible case of relatively recent acquisition (horizontal transfer).
In two cases out of the six large polymorphisms identified, there was insertion of a DNA
segment with a modification gene homolog, accompanied by target deletion (simple
substitution).  In two other cases, homologous DNA segments carrying a modification gene
homolog were present at different locations in the two genomes (transposition). In both cases,
substitution (insertion/deletion) in one of the two loci was accompanied by inversion of the
adjacent chromosomal segment. In the fifth case, substitution by a DNA segment carrying type I
restriction, modification, and specificity gene homologs was likewise accompanied by adjacent
inversion. In the last case, two homologous DNA segments, were found at different loci in the
two genomes (transposition), but only one of them had insertion of a modification homolog and
an unknown ORF.  The possible relationship of these polymorphisms to attack by restriction
enzymes on the chromosome will be discussed.

<>

<1>Chinen, A., Uchiyama, I., Kobayashi, I.
<2>Association of putative restriction modification genes with large genome polymorphisms suggested from comparison of Pyrococcus horikoshii and Pyrococcus abyssi genome sequences.
<3>Genome Informatics
<4>11
<5>339-340
<6>2000
<7>Restriction-modification (RM) gene complexes encode two enzymatic functions, restriction and
modification.  A restriction enzyme will recognize a specific sequence in DNA and cut the DNA
unless it is methylated by a cognate modification enzyme.  RM systems will defend bacterial
cells by attacking incoming foreign DNA.  It is widely held that bacteria have evolved RM
systems and maintain them in order to protect their genome from invasion by foreign DNA such
as bacteriophages and plasmids. We earlier reported that carrying a type II RM gene complex
can increase the stability of a plasmid because cells that have lost the plasmid die.  From
this and other observations, it was proposed that the relative frequency of RM gene complexes
has increased through this post-segregational killing, in competitive exclusion, as well as
through direct attack on invading DNA.

<>

<1>Chinenova, T.A., Mkrtumian, N.M., Lomovskaia, N.D.
<2>[Genetic characteristics of a new phage resistance trait in Streptomyces coelicolor A3(2)].
<3>Genetika
<4>18
<5>1945-1952
<6>1982
<7>Phage resistance was investigated in the system of Streptomyces coelicolor A3(2) and phi C31
actinophage. Resistance of A3(2) strain to phi C31 was
shown to involve a novel mechanism responsible for the arrest of phage
intracellular growth in the whole cell population. The phage resistance
character designated Pgl+ (for 'phage growth limiting') is determined by a
gene located on the A3(2) chromosome. The gene (pgl) controls phage
'modification' which results in an inability of phage to lyse lysogenize
Pgl+ host. Instability of the Pgl+ character was revealed. Pgl+ strains
segregate Pgl variants at a high frequency, the majority of Pgl strains
reverting to the initial Pgl+ phenotype with the same high frequency.
Reversible Pgl+ in equilibrium Pgl transitions are a common feature of
A3(2) cell population.

<>

<1>Chiou, C.S., Li, H.Y., Tung, S.K., Chen, C.Y., Teng, C.H., Shu, J.C., Tseng, J.T., Hsu, C.Y., Chen, C.C.
<2>Identification of prophage gene z2389 in Escherichia coli EDL933 encoding a DNA cytosine methyltransferase for full protection of Notl sites.
<3>Int. J. Med. Microbiol.
<4>300
<5>296-303
<6>2010
<7>Pulsed-field gel electrophoresis (PFGE) analysis revealed that the genomes of some pathogenic
Escherichia coli O157:H7 strains, including
EDL933, were resistant to Noti digestion. An amino acid sequence
comparison suggested that the z2389 gene carried on prophage CP-933R in
strain EDL933 is likely to encode a C-5-cytosine methyltransferase. The
z2389-equivalent gene was found in the Notl-resistant strains tested,
but it was not detected in the Notl-susceptible strains. PFGE analysis
of the wild-type EDL933 strain and of a z2389 null mutant revealed that
z2389 was associated with full genome protection against Notl digestion
and partial protection against Eagl digestion. In vitro methylation
experiments with purified recombinant protein demonstrated that Z2389
is capable of methylating Notl and Eagl sites. Sequencing of
bisulfite-treated DNA indicated that the methylation occurred at the
first cytosine residue of the Notl recognition sequence, whereas Eagl
sites remained unmethylated or were methylated at the first cytosine
residue. Thus, z2389 encodes a DNA cytosine methyltransferase that
confers full protection to Notl sites.

<>

<1>Chirikjian, J.G., George, A., Smith, L.A.
<2>Purification of restriction endonucleases including the isolation of a novel activity using affinity chromatography.
<3>Fed. Proc.
<4>37
<5>1415
<6>1978
<7>We wish to report a new method for purifying EndoRs based on affinity
fractionation.  To date purification of restriction endonucleases have been
modifications of few early methods which are often tedious and time consuming.
We have investigated the use of affinity matrices such as covalently bound
pyran and Cibacron Blue F3Ga sepharose.  Pyran is a divinyl ether copolymer of
maleic anhydride which in carboxylic acid solution hydrolyzes to the negatively
charged polycarboxylic acid that simulates the negatively charged
phosphodiester bond.  Cibacron Blue F3Ga is a derivative of a sulfonated
polyaromatic dye and is widely used as a nonspecific affinity matrix.  Both
matrices have high capacity, rapid development time and yield enzyme free from
interfering activities.  Using these matrices we have purified several reported
EndoRs such as BamHI, HinfI, PstI and XbaI.  We have also isolated a new Endo3
from a clinical isolate H. influenzae Georgetown, which contains two major
activities: HindGUI which we have temporarily assigned to be an isoschizomer of
HhaI and HinGUII with the following fragmentation properties: PhiX174, 10;
SV40, 12-14; lambda >50; and Adeno 2 >50.

<>

<1>Chirikjian, J.G., George, J.
<2>Sequence-specific endonucleases: General properties and specific characteristics.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>73-99
<6>1981
<7>I. Introduction II. Sequence-specific endonucleases as reagents in molecular biology. III.
Matrix bound sequence-specific endonucleases IV. General methods for determining
sequence-specific endonuclease activity. V. Strategies for elucidating the recognition of DNA
by sequence-specific endonucleases VI. Catalytic diversities among isoschizomers VII. Effect
of hydrophobic reagents on sequence recognition and catalysis of sequence-specific
endonucleases VIII. Conclusions

<>

<1>Chistoserdova, L., Lapidus, A., Han, C., Goodwin, L., Saunders, L., Brettin, T., Tapia, R., Gilna, P., Lucas, S., Richardson, P.M., Lidstrom, M.E.
<2>Genome of Methylobacillus flagellatus, Molecular Basis for Obligate Methylotrophy, and Polyphyletic Origin of Methylotrophy.
<3>J. Bacteriol.
<4>189
<5>4020-4027
<6>2007
<7>Along with methane, methanol and methylated amines represent important biogenic atmospheric
constituents; thus, not only methanotrophs but also
nonmethanotrophic methylotrophs play a significant role in global carbon
cycling. The complete genome of a model obligate methanol and methylamine
utilizer, Methylobacillus flagellatus (strain KT) was sequenced. The
genome is represented by a single circular chromosome of approximately 3
Mbp, potentially encoding a total of 2,766 proteins. Based on genome
analysis as well as the results from previous genetic and mutational
analyses, methylotrophy is enabled by methanol and methylamine
dehydrogenases and their specific electron transport chain components, the
tetrahydromethanopterin-linked formaldehyde oxidation pathway and the
assimilatory and dissimilatory ribulose monophosphate cycles, and by a
formate dehydrogenase. Some of the methylotrophy genes are present in more
than one (identical or nonidentical) copy. The obligate dependence on
single-carbon compounds appears to be due to the incomplete tricarboxylic
acid cycle, as no genes potentially encoding alpha-ketoglutarate, malate,
or succinate dehydrogenases are identifiable. The genome of M. flagellatus
was compared in terms of methylotrophy functions to the previously
sequenced genomes of three methylotrophs, Methylobacterium extorquens (an
alphaproteobacterium, 7 Mbp), Methylibium petroleiphilum (a
betaproteobacterium, 4 Mbp), and Methylococcus capsulatus (a
gammaproteobacterium, 3.3 Mbp). Strikingly, metabolically and/or
phylogenetically, the methylotrophy functions in M. flagellatus were more
similar to those in M. capsulatus and M. extorquens than to the ones in
the more closely related M. petroleiphilum species, providing the first
genomic evidence for the polyphyletic origin of methylotrophy in
Betaproteobacteria.

<>

<1>Chiu, C.H., Tang, P., Chu, C., Hu, S., Bao, Q., Yu, J., Chou, Y.Y., Wang, H.S., Lee, Y.S.
<2>The genome sequence of Salmonella enterica serovar Choleraesuis, a highly invasive and resistant zoonotic pathogen.
<3>Nucleic Acids Res.
<4>33
<5>1690-1698
<6>2005
<7>Salmonella enterica serovar Choleraesuis (S.Choleraesuis), a highly invasive serovar among
non-typhoidal Salmonella, usually causes sepsis or
extra-intestinal focal infections in humans. S.Choleraesuis infections
have now become particularly difficult to treat because of the emergence
of resistance to multiple antimicrobial agents. The 4.7 Mb genome sequence
of a multidrug-resistant S.Choleraesuis strain SC-B67 was determined.
Genome wide comparison of three sequenced Salmonella genomes revealed that
more deletion events occurred in S.Choleraesuis SC-B67 and S.Typhi CT18
relative to S.Typhimurium LT2. S.Choleraesuis has 151 pseudogenes, which,
among the three Salmonella genomes, include the highest percentage of
pseudogenes arising from the genes involved in bacterial chemotaxis
signal-transduction pathways. Mutations in these genes may increase smooth
swimming of the bacteria, potentially allowing more effective interactions
with and invasion of host cells to occur. A key regulatory gene of
TetR/AcrR family, acrR, was inactivated through the introduction of an
internal stop codon resulting in overexpression of AcrAB that appears to
be associated with ciprofloxacin resistance. While lateral gene transfer
providing basic functions to allow niche expansion in the host and
environment is maintained during the evolution of different serovars of
Salmonella, genes providing little overall selective benefit may be lost
rapidly. Our findings suggest that the formation of pseudogenes may
provide a simple evolutionary pathway that complements gene acquisition to
enhance virulence and antimicrobial resistance in S.Choleraesuis.

<>

<1>Chiu, C.M., Chang, C.H., Pan, S.F., Wu, H.C., Li, S.W., Chang, C.H., Lee, Y.S., Chiang, C.M., Chen, Y.S.
<2>Draft Genome Sequence of Lactobacillus pobuzihii E100301T.
<3>Genome Announcements
<4>1
<5>e00185-13
<6>2013
<7>Lactobacillus pobuzihii E100301(T) is a novel Lactobacillus species previously isolated from
pobuzihi (fermented cummingcordia) in Taiwan. Phylogenetically,
this strain is closest to Lactobacillus acidipiscis, but its phenotypic
characteristics can be clearly distinguished from those of L. acidipiscis. We
present the draft genome sequence of strain L. pobuzihii E100301(T).

<>

<1>Chiura, H., Noro, Y., Kanayama, S., Ueda, Y., Simidu, U., Takagi, J.
<2>Site specific deoxyribonuclease produced by a marine bacterium, Flavobacterium I 16-04.
<3>Agric. Biol. Chem.
<4>52
<5>2107-2109
<6>1988
<7>Since the discovery of HindII, more than 120 kinds of site-specific
endodeoxyribonucleases (restriction enzymes) have been isolated from some 600
species of bacteria and reported.  All of them have been obtained from
terrestrial bacteria except FokI isolated from a marine bacterium,
Flavobacterium okeanokoites.  Furthermore, the marine bacteria have not been as
extensively studied as the terrestrial bacteria.  Under these circumstances, we
have been attempting to survey site-specific endonucleases in the marine
bacteria of the laboratory collection of the Ocean Research Institute,
University of Tokyo.

<>

<1>Chiura, H.X., Kamiyama, T., Hirano, H., Futagami, M., Watahiki, M., Kobayashi, K., Simidu, U., Takagi, J.
<2>Purification and characterization of AspMDI, an isoschizomer of Sau3AI, from a marine bacterium, Alcaligenes sp MD1.
<3>Nucleic Acids Res.
<4>20
<5>1996
<6>1992
<7>A new type II restriction endonuclease, Asp MD1, was isolated from a pigmented marine
bacterium Alcaligenes species MD1 which was collected from the open sea around Taiwan Island.

<>

<1>Chiurillo, M.A., Moran, Y., Canas, M., Valderrama, E., Granda, N., Sayegh, M., Ramirez, J.L.
<2>Genotyping of virulence-associated genes from biopsy specimens show high genetic diversity of Helicobacter pylori strains infecting patients in Venezuela.
<3>Int. J. Infect. Dis.
<4>17
<5>e750-6
<6>2013
<7>Background: Helicobacter pylori is a major cause of chronic gastritis and an established risk
factor for gastric adenocarcinoma. This bacterium also exhibits an extraordinarily high
genetic diversity.
Methods: The genetic diversity of H. pylori strains from Venezuelan patients with chronic
gastritis was evaluated by PCR-typing of vacA, cagA, iceA, and babA2 virulence-associated
genes using DNA extracted directly from biopsies. The nucleotide sequence and prevalence of
size variants of iceA1, iceA2, and babA2 PCR products were introduced in this analysis.
Results: The frequency of vacA s1 was associated (p < 0.01) with moderate/severe grades of
atrophic
gastritis. The cagA, iceA1, iceA2, and babA2 genotypes were found in 70.6%, 66.4%, 33.6%, and
92.3% of strains, respectively. The frequency of iceA2 and its subtype iceA2_D were higher (p
< 0.015) in cases with moderate/severe granulocytic inflammation. The most prevalent combined
genotypes were vacA
s1m1/cagA/iceA1/babA2 (26.3%), vacA s2m2/iceA1/babA2 (19.5%), and vacA s1m1/cagA/iceA2/babA2
(18.8%). Sequence analysis of iceA1, iceA2, and babA2 PCR-amplified fragments allowed us to
define allelic variants and to increase the number of genotypes detected (from 19 to 62). A
phylogenetic tree made with iceA1 sequences showed that the H. pylori strains analyzed here
were grouped with those of Western origin.
Conclusions: Our results show that patients from the western region of Venezuela have an
elevated prevalence of infection with H. pylori strains carrying known virulence genotypes
with high genetic diversity. This highlights the importance of identifying gene variants for
an early detection of virulent genotypes.

<>

<1>Chivian, D. et al.
<2>Environmental genomics reveals a single-species ecosystem deep within Earth.
<3>Science
<4>322
<5>275-278
<6>2008
<7>DNA from low-biodiversity fracture water collected at 2.8-kilometer depth in a South African
gold mine was sequenced and assembled into a single,
complete genome. This bacterium, Candidatus Desulforudis audaxviator,
composes >99.9% of the microorganisms inhabiting the fluid phase of this
particular fracture. Its genome indicates a motile, sporulating,
sulfate-reducing, chemoautotrophic thermophile that can fix its own
nitrogen and carbon by using machinery shared with archaea. Candidatus
Desulforudis audaxviator is capable of an independent life-style well
suited to long-term isolation from the photosphere deep within Earth's
crust and offers an example of a natural ecosystem that appears to have
its biological component entirely encoded within a single genome.

<>

<1>Chlebowicz, M.A., Nganou, K., Kozytska, S., Arends, J.P., Engelmann, S., Grundmann, H., Ohlsen, K., van Dijl, J.M., Buist, G.
<2>Recombination between ccrC genes in a type V (5C2 and 5) staphylococcal cassette chromosome mec (SCCmec) of Staphylococcus aureus ST398 leads to conversion from methicillin resistance to methicillin susceptibility in vivo.
<3>Antimicrob. Agents Chemother.
<4>54
<5>783-791
<6>2010
<7>Various types of the staphylococcal cassette chromosome mec (SCCmec) are known to confer
methicillin resistance on the human pathogen Staphylococcus aureus. Such cassettes are not
always stably maintained.  The present studies were aimed at identifying the mechanism
underlying the in vivo conversion of methicillin-resistant S. aureus (MRSA) to
methicillin-susceptible S. aureus (MSSA) derivatives as encountered in two patients suffering
from pneumonia and an umbilicus infection, respectively. All MRSA and MSSA isolates identified
belong to multilocus sequence type (MLST) 398, have spa type t034, and are Panton-Valentine
leukocidin positive. Sequencing of 27,616 nucleotides from the chromosomal SCCmec insertion
site in orfX to the hsdR gene for a restriction enzyme revealed a type V (5C2&5) SCCmec.
Sequence comparisons show that parts of the cassette are highly similar to sequences within
SCCmec elements from coagulase-negative staphylococci, indicating a possible common origin.
The cassette investigated contains ccrC-carrying units on either side of its class C2b mec
gene complex. In vivo loss of the mec gene complex was caused by recombination between the
recombinase genes ccrC1 allele 8 and ccrC1 allele 10. In vitro, the SCCmec was very stable,
and low-frequency MRSA-to-MSSA conversion was only observed when MRSA isolates were cultivated
at 41 degrees C for prolonged periods of time. In this case also, loss of the mec complex was
due to ccrC gene recombination. Interestingly, the MRSA and MSSA isolates studied displayed no
detectable differences in competitive growth and virulence, suggesting that the presence of
the intact type V (5C2&5) SCCmec has no negative bearing on staphylococcal fitness under the
conditions used.

<>

<1>Chmelo, R., Foltz, L., Eadie, J.S.
<2>Determination of restriction enzyme digestion completeness under low target concentration.
<3>FASEB J.
<4>7
<5>A1304
<6>1993
<7>A method for the determination of restriction enzyme digestion efficiency under low target
concentration is described. Target DNA was serially diluted from 10 to the eleventh power
molecules to 1 molecule. The DNA was digested with one unit of enzyme for one hour at 37C. The
digestion was stopped by heating the reaction at 65C for 10 minutes. PCR was performed on the
restriction digests to assay for completeness. Three primers were used: 2 forward primers and
1 reverse primer. When both forward primers are used in conjunction with the reverse primer in
PCR, products of 523 and 170 base pairs are observed. The restriction site of interest lies
between the two forward primers. If the enzyme is active at low target concentration, only the
170 base pair fragment is detected. Results have indicated that this technique is sensitive to
detect completeness of a restriction enzyme digestion at low target DNA concentration. This
protocol can be applicable for testing any restriction enzyme.

<>

<1>Chmiel, A.A., Bujnicki, J.M., Skowronek, K.J.
<2>A homology model of restriction endonuclease SfiI in complex with DNA.
<3>BMC Struct. Biol.
<4>5
<5>2
<6>2005
<7>BACKGROUND: Restriction enzymes (REases) are commercial reagents commonly used in recombinant
DNA technologies. They are attractive models for studying protein-DNA interactions and
valuable targets for protein engineering. They are, however, extremely divergent: the amino
acid sequence of a typical REase usually shows no detectable similarities to any other
proteins, with rare exceptions of other REases that recognize identical or very similar
sequences. From structural analyses and bioinformatics studies it has been learned that some
REases belong to at least four unrelated and structurally distinct superfamilies of nucleases,
PD-DxK, PLD, HNH, and GIY-YIG. Hence, they are extremely hard targets for structure prediction
and homology-based inference of sequence-function relationships and the great majority of
REases remain structurally and evolutionarily unclassified. RESULTS: SfiI is a REase which
recognizes the interrupted palindromic sequence 5'GGCCNNNN--NGGCC3' and generates 3 nt long
3' overhangs upon cleavage. SfiI is an archetypal Type IIF enzyme, which functions as a
tetramer and cleaves two copies of the recognition site in a concerted manner. Its sequence
shows no similarity to other proteins and nothing is known about the localization of its
active site or residues important for oligomerization. Using the threading approach for
protein fold-recognition, we identified a remote relationship between SfiI and BglI, a dimeric
Type IIP restriction enzyme from the PD-DxK superfamily of nucleases, which recognizes the
5'GCCNNNN--NGGC3' sequence and whose structure in complex with the substrate DNA is
available. We constructed a homology model of SfiI in complex with its target sequence and
used it to predict residues important for dimerization, tetramerization, DNA binding and
catalysis. CONCLUSIONS: The bioinformatics analysis suggest that SfiI, a Type IIF enzyme, is
more closely related to BglI, an "orthodox" Type IIP restriction enzyme, than to any other
REase, including other Type IIF REases with known structures, such as NgoMIV. NgoMIV and BglI
belong to two different, very remotely related branches of the PD-DxK superfamily: the
alpha-class (EcoRI-like), and the beta-class (EcoRV-like), respectively. Thus, our analysis
provides evidence that the ability to tetramerize and cut the two DNA sequences in a concerted
manner was developed independently at least two times in the evolution of the PD-DxK
superfamily of REases. The model of SfiI will also serve as a convenient platform for further
experimental analyses.

<>

<1>Chmiel, A.A., Radlinska, M., Pawlak, S.D., Krowarsch, D., Bujnicki, J.M., Skowronek, K.J.
<2>A theoretical model of restriction endonuclease NlaIV in complex with DNA, predicted by fold recognition and validated by site-directed  mutagenesis and circular dichroism spectroscopy.
<3>Protein Eng. Des. Sel.
<4>18
<5>181-189
<6>2005
<7>Restriction enzymes (REases) are commercial reagents commonly used in DNA manipulations and
mapping. They are regarded as very attractive
models for studying protein-DNA interactions and valuable targets for
protein engineering. Their amino acid sequences usually show no
similarities to other proteins, with rare exceptions of other REases
that recognize identical or very similar sequences. Hence, they are
extremely hard targets for structure prediction and modeling. NlaIV is
a Type II REase, which recognizes the interrupted palindromic sequence
GGNNCC (where N indicates any base) and cleaves it in the middle,
leaving blunt ends. NlaIV shows no sequence similarity to other
proteins and virtually nothing is known about its
sequence-structure-function relationships. Using protein fold
recognition, we identified a remote relationship between NlaIV and
EcoRV, an extensively studied REase, which recognizes the GATATC
sequence and whose crystal structure has been determined. Using the
'FRankenstein's monster' approach we constructed a comparative model of
NlaIV based on the EcoRV template and used it to predict the catalytic
and DNA-binding residues. The model was validated by site-directed
mutagenesis and analysis of the activity of the mutants in vivo and in
vitro as well as structural characterization of the wild-type enzyme
and two mutants by circular dichroism spectroscopy. The structural
model of the NlaIV-DNA complex suggests regions of the protein sequence
that may interact with the 'non-specific' bases of the target and thus
it provides insight into the evolution of sequence specificity in
restriction enzymes and may help engineer REases with novel
specificities. Before this analysis was carried out, neither the
three-dimensional fold of NlaIV, its evolutionary relationships or its
catalytic or DNA-binding residues were known. Hence our analysis may be
regarded as a paradigm for studies aiming at reducing 'white spaces' on
the evolutionary landscape of sequence-function relationships by
combining bioinformatics with simple experimental assays.

<>

<1>Chmuzh, E.V., Kashirina, J.G., Tomilova, J.E., Mezentseva, N.V., Dedkov, V.S., Gonchar, D.A., Abdurashitov, M.A., Degtyarev, S.K.
<2>A Novel Restriction endonuclease BisI from Bacillus subtilis T30, recognizes a methylated DNA sequence 5'- G(m5C)^NGC-3'.
<3>Biotekhnologiya
<4>3
<5>22-26
<6>2005
<7>Bacillus subtilis strain T30, producing a novel restriction endonuclease Bis I, has been
isolated and characterized. The enzyme recognizes methylated DNA sequence 5'- G(m5C)^NGC-3'
and cleaves it as shown by the arrow. Due to cleavage of only modified DNAs Bis I may find a
practical application in genetic engineering experiments as well as in determination of
eukaryotic DNA methylation status.

<>

<1>Chmuzh, E.V., Kashirina, Y.G., Tomilova, Y.E., Chernukhin, V.A., Okhapkina, S.S., Gonchar, D.A., Dedkov, V.S., Abdurashitov, M.A., Degtyarev, S.K.
<2>The Fsp4HI restriction-modification system: Gene cloning, comparison of protein structures, and biochemical properties of recombinant DNA methyltransferase M.Fsp4HI.
<3>Mol. Biol. (Mosk)
<4>41
<5>43-50
<6>2007
<7>Genes coding for the Flavobacterium sp. 4H restriction-modification (RM) system, which
recognizes the sequence 5'-GCNGC-3', were cloned in Escherichia coli ER2267 and sequenced.
The Fsp4HI RM system includes two genes: one for DNA methyltransferase (M.) and the other for
restriction endonuclease (R.), immediately following the former in the same direction. The
genes partly overlap. According to the deduced amino acid sequences, M.Fsp4HI belongs to C5
DNA methyltransferases, whereas R.Fsp4HI is only slightly similar to some restriction enzymes
recognizing similar sequences. M.Fsp4HI was purified by column chromatography. The optimal
conditions for the enzyme are 30 degrees C and pH 7.5. M.Fsp4HI modifies the first cytosine in
5'-GCNGC-3'.

<>

<1>Cho, E., Park, S.N., Kim, H.K., Kim, D.S., Jung, J., Baek, J.H., Lim, Y.K., Jo, E., Choi, M.H., Chang, Y.H., Shin, Y., Paek, J., Shin, J.H., Kim, J., Choi, S.H., Park, H.S., Kim, H., Kook, J.K.
<2>Draft Genome Sequence of the Novel Peptoniphilus sp. Strain ChDC B134, Isolated from a Human Periapical Abscess Lesion.
<3>Genome Announcements
<4>1
<5>e00822-13
<6>2013
<7>The genus Peptoniphilus comprises butyrate-producing, nonsaccharolytic species that use
peptone and amino acids as major energy sources. The novel Peptoniphilus
sp. strain ChDC B134 (=KCOM 1628) was isolated from a human periapical abscess
lesion. Here, we report the draft genome sequence of the strain.

<>

<1>Cho, K., Yoon, K., Choi, J., Oh, H., Lee, M.
<2>Novel Lactobacillus paracasei CLW-011 strain for probiotics andspecific DNA sequences of the same.
<3>Korean Patent Office
<4>KR 1020020041468 A
<5>
<6>2002
<7>
<>

<1>Cho, M.-J., Park, J.-U., Jeon, B.S., Pack, J.-W., Byun, E.Y., Lee, S.-K., Park, Y.-H., Song, J.-H., Lee, W.-K., Baik, S.-C., Choi, Y.-J., Jung, S.-A., Choe, M.-Y., Choi, S.-H., Ko, G.-H., Youn, H.-S., Rhee, K.-H.
<2>Characterization of type II restriction endonucleases (Hpy51-I) from Helicobacter pylori strain 51.
<3>J. Bact. Virol.
<4>31
<5>207-215
<6>2001
<7>This study describes the purification and characterization of a type II restriction
endonuclease of Helicobacter pylori in order to understand the DNA restriction and
modification of H. pylori.  H. pylori cell extract was subjected to polyethyleneimine
treatment, salt precipitation, heparine-sepharose column chromatography, and fast protein
liquid chromatography using Resource Q column and Mono Q column to purify the type II
restriction endonuclease.  Hpy51-I was characterized to recognize the sequence
5'-GT(G/C)AC-3', yielding 5-base 5' protruding ends.  The restriction sequence was
identical to that of Tsp45I.  The enzyme exhibited its maximal activity in the presence of
10~20 mM NaCl, but was inhibited completely in the presence of more than 80 mM NaCl.  The
enzyme showed its maximal activity in the presence of 1~10 mM MCl2.  The optimal pH and
temperature for enzyme activity was pH 9.0 and 37oC, respectively.  MnCl2 could not substitute
for MgCl2 in reaction mixture.  Addition of beta-mercaptoethanol and bovine serum albumin in
the reaction mixture led to loss of enzyme activity of Hpy51-I.  The whole cell extract of H.
pylori strain 51 was confirmed to carry the enzyme activity for methylation of
Hpy51-I-recognised sequence.  Hpy51-I digested genomic DNAs of enteric bacteria to less than I
kb while it could not cut the genomic DNAs of H. pylori isolates.  In this study, the type II
restriction enzyme (Hpy51-I) of H. pylori was identified and its biochemical properties
characterized, demonstrating that Hpy51-I might be one of the barriers for preventing the
introduction of foreign DNAs into H. pylori.

<>

<1>Cho, N.H., Kim, H.R., Lee, J.H., Kim, S.Y., Kim, J., Cha, S., Kim, S.Y., Darby, A.C., Fuxelius, H.H., Yin, J., Kim, J.H., Kim, J., Lee, S.J., Koh, Y.S., Jang, W.J., Park, K.H., Andersson, S.G., Choi, M.S., Kim, I.S.
<2>The Orientia tsutsugamushi genome reveals massive proliferation of conjugative type IV secretion system and host-cell interaction genes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>7981-7986
<6>2007
<7>Scrub typhus is caused by the obligate intracellular rickettsia Orientia tsutsugamushi
(previously called Rickettsia tsutsugamushi). The bacterium
is maternally inherited in trombicuid mites and transmitted to humans by
feeding larvae. We report here the 2,127,051-bp genome of the Boryong
strain, which represents the most highly repeated bacterial genome
sequenced to date. The repeat density of the scrub typhus pathogen is
200-fold higher than that of its close relative Rickettsia prowazekii, the
agent of epidemic typhus. A total of 359 tra genes for components of
conjugative type IV secretion systems were identified at 79 sites in the
genome. Associated with these are >200 genes for signaling and host-cell
interaction proteins, such as histidine kinases, ankyrin-repeat proteins,
and tetratrico peptide-repeat proteins. Additionally, the O. tsutsugamushi
genome contains >400 transposases, 60 phage integrases, and 70 reverse
transcriptases. Deletions and rearrangements have yielded unique gene
combinations as well as frequent pseudogenization in the tra clusters. A
comparative analysis of the tra clusters within the genome and across
strains indicates sequence homogenization by gene conversion, whereas
complexity, diversity, and pseudogenization are acquired by duplications,
deletions, and transposon integrations into the amplified segments. The
results suggest intragenomic duplications or multiple integrations of a
massively proliferating conjugative transfer system. Diversifying
selection on host-cell interaction genes along with repeated population
bottlenecks may drive rare genome variants to fixation, thereby
short-circuiting selection for low complexity in bacterial genomes.

<>

<1>Cho, S.-H., Kang, C.
<2>DNA sequence-dependent cleavage sites of restriction endonuclease HphI.
<3>Mol. Cells
<4>1
<5>81-86
<6>1990
<7>A class IIS restriction endonuclease HphI has been known to make a staggered
cut always at the 8th base pair downstream of its recognition sequence
5'GGTGA3', producing one-base 3' protruding ends, regardless of the DNA
sequence around the cleavage site.  Since the cleavage sites of HphI are
separate from its recognition sites, the effects of mutations around a cleavage
site have been studied using a phage SP6 promoter-containing plasmid pSP64
which has an HphI cleavage site near the transcription initiation site.  The
exact cutting site of the lower strand was determined to be after the 8th
nucleotide downstream of the recognition site, indicating a staggered cut at
the 9th base pair downstream.  In addition, the same cutting site in the
mutants carrying a few base pair deletions around the cleavage site is shown to
be shifted.  These variations were not affected by any changes in reaction
conditions.  These results suggest that the HphI cleavage site shifts depending
on the DNA sequence, which might affect the DNA helical structure.

<>

<1>Cho, Y.J., Choi, J.K., Kim, J.H., Lim, Y.S., Ham, J.S., Kang, D.K., Chun, J., Paik, H.D., Kim, G.B.
<2>Genome Sequence of Lactobacillus salivarius GJ-24, a probiotic strain isolated from healthy adult intestine.
<3>J. Bacteriol.
<4>193
<5>5021-5022
<6>2011
<7>The draft genome sequence of Lactobacillus salivarius GJ-24 isolated from the feces of healthy
adults was determined. The properties including milk
fermentation activity and bacteriocin production suggest its potential
uses as a probiotic lactic acid bacterium and start culture for dairy
products.

<>

<1>Choe, W., Chandrasegaran, S., Ostermeier, M.
<2>Protein fragment complementation in M.HhaI DNA methyltransferase.
<3>Biochem. Biophys. Res. Commun.
<4>334
<5>1233-1240
<6>2005
<7>The 5mC DNA methyltransferase M.HhaI can be split into two individually inactive N- and
C-terminal fragments that together can form an active
enzyme in vivo capable of efficiently methylating DNA. This active
fragment pair was identified by creating libraries of M.HhaI gene
fragment pairs and then selecting for the pairs that code for an active
5mC methyltransferase. The site of bisection for successful protein
fragment complementation in M.HhaI was in the variable region near the
target recognition domain between motif VIII and TRD. This same region
is the location of bifurcation in the naturally split 5mC
methyltransferase M.AquI, the location for circular permutation in
M.BssHII, and the location for previously engineered split versions of
M.BspRI.

<>

<1>Choi, D.H., Ryu, J.Y., Kwon, K.K., Lee, J.H., Kim, C., Lee, C.M., Noh, J.H.
<2>Draft genome sequence of Rubidibacter lacunae strain KORDI 51-2(T), a cyanobacterium isolated from seawater of Chuuk lagoon.
<3>Standards in Genomic Sciences
<4>9
<5>197-204
<6>2013
<7>A photoautotrophic cyanobacterium, Rubidibacter lacunae was reported in 2008 for  the first
time. The type strain, KORDI 51-2(T), was isolated from seawater of
Chuuk lagoon located in a tropical area. Although it belonged to a clade
exclusively comprised of extremely halotolerant strains by phylogenetic analyses,
R. lacunae is known to be incapable of growth at high salt concentration over
10%. Here we report the main features of the genome of R. lacunae strain KORDI
51-2(T). The genome of R. lacunae contains a gene cluster for phosphonate
utilization encoding three transporters, one regulator and eight C-P lyase
subunits.

<>

<1>Choi, G.-E., Cho, Y.J., Koh, W.J., Chun, J., Cho, S.N., Shin, S.J.
<2>Draft Genome Sequence of Mycobacterium abscessus subsp. bolletii BDT.
<3>J. Bacteriol.
<4>194
<5>2756-2757
<6>2012
<7>Mycobacterium abscessus subsp. bolletii is an increasing cause of human pulmonary disease and
infections of the skin and soft tissues. Consistent reports of human infections indicate that
M. bolletii is a highly pathogenic, emerging species of rapidly growing mycobacteria (RGM).
Here we report the first whole-genome sequence of M. abscessus subsp. bolletii BD(T).

<>

<1>Choi, H.S., Kwon, M.G., Kim, M.S., Park, M.A., Kim, D.W., Park, J.Y., Kim, J.S., Na, Y.J., Kim, M.Y., Kim, D.S., Chae, S.H., Seo, J.S.
<2>Draft Genome Sequence of Beta-Hemolytic Streptococcus iniae KCTC 11634.
<3>Genome Announcements
<4>1
<5>e00897-13
<6>2013
<7>Streptococcus iniae is a beta-hemolytic, Gram-positive coccus, which affects a broad range of
freshwater and marine fish species, causing substantial economic
losses in the aquaculture industry worldwide. Thus, it is very important to
derive a complete genome sequence of the bacterium to aid in the development of
vaccines and methods for preventing fish streptococcosis and zoonotic infections
in humans. Here, we present the draft genome sequence of S. iniae KCTC 11634
(1,955,615 bp, with a G+C content of 36.6%), which contains 1,868 putative coding
sequences.

<>

<1>Choi, K.D., Kim, K., Yoo, O.J.
<2>A new restriction endonuclease from Clostridium thermocellum.
<3>Sanop Misaengmul Hakhoe Chi
<4>15
<5>352-355
<6>1987
<7>The isolation and characterization of the Type II restriction endonuclease from Clostridium
thermocellum ATCC 27405 is described. This enzyme (CthI endonuclease) is an isoschizomer of
BclI endonuclease recognizing 5'-TGATCA-3'. CthI endonuclease requires Mg2+ ion for its
activity and is maximally active at pH 7.5 to 10.5 in the presence of 0 to 10mM NaCl. CthI
endonuclease is heat stable and has an optimum temperature of 60C. The activity of CthI enzyme
is sensitive to dam methylation.

<>

<1>Choi, K.D., Yoo, O.J.
<2>A new restriction endonuclease, CthII, from Clostridium thermocellum ATCC 27405.
<3>Korean Biochem. J.
<4>23
<5>418-421
<6>1990
<7>A new restriction enzyme has been isolated by DEAE-cellulose and phosphocellulose columns from
a thermophilic anaerobic bacterium, Clostridium thermocellum ATCC 27405. Following CthI, the
second peak of sequence specific endonuclease activity was eluted from the phosphocellulose
column. The enzyme was designated as CthII. The recognition sequence of CthII was determined
to be the dcm sequence, 5'-CC^(A/T) GG-3' and the cleavage site was indicated by the arrow.
CthII endonuclease requires Mg2+ ion for its activity and is maximally active at a pH range
from 8.0 to 9.0. CthII endonuclease is heat stable and has an optimum reaction temperature of
55C. Like BstNI and ZanI this enzyme was able to cleave dcm-methylated DNA.

<>

<1>Choi, O., Lim, J.Y., Seo, Y.S., Hwang, I., Kim, J.
<2>Complete Genome Sequence of the Rice Pathogen Pantoea ananatis Strain PA13.
<3>J. Bacteriol.
<4>194
<5>531
<6>2012
<7>Pantoea ananatis is the causative agent of sheath and grain rot in rice. Here, we present the
complete genome sequence of P. ananatis strain PA13,
originally isolated from a diseased rice grain.

<>

<1>Choi, S.-H.
<2>Restriction-modification systems and genetic variability of Xanthomonas oryzae pv.
<3>Diss. Abstr.
<4>54
<5>5462B-5463B
<6>1994
<7>The XorII methyltransferase gene (xorIIM) and very short patch repair endonuclease gene
(xorii.vsr) were cloned from Xanthomonas oryzae pv. oryzae, the rice bacterial leaf blight
pathogen. XorIIM encodes a polypeptide of 47 KD and was identified as a monospecific m5
cytosine methyltransferase gene. The xorii.vsr encodes a polypeptide of 19.7 KD and the gene
is similar in sequence and size to the E. coli vsr gene of the DNA cytosine methylase system
(Dcm). A population of X. oryzae pv. oryzae strains from major rice growing countries in Asia
was evaluated for the presence or absence of the XorI and XorII restriction-modification (R-M)
systems. Four clonal populations with the phenotype XorI+II+, XorI-II+, XorI+II-, XorI-II+,
and XorI-II- were distributed in Asia. The XorII R-M system was predominantly found in
southeast Asia, whereas the XorI modification system was most prevalent in northeast Asia. DNA
polymorphisms were observed between strains in genomic sequences containing the XorII R-M
genes; however, most Philippine strains and all the Indonesian and Korean strains had
identical patterns. Based on the geographic distribution of both systems and the genome
organization around the XorII system, I propose that the XorI system originated in northeast
Asia and moved to southeast Asia, while the XorII system originated in southeast Asia. The
existence of several phenotypes in some parts of Asia indicate that after movement of the
systems the populations remained clonal.

A marker-exchange mutant in which the avirulence gene locus avrXa7 was insertionally
inactivated was significantly reduced in aggressiveness to susceptible rice cultivars.
Aggressiveness was restored by complementation with a plasmid bearing the avrXa7 gene. Thus,
avrXa7 codes for not only resistance-gene-specific avirulence function, but also for
pathogenicity functions.


<>

<1>Choi, S.C., Parker, J., Richards, V.P., Ross, K., Jilly, B., Chen, J.
<2>Draft Genome Sequence of an Atypical Strain of Streptococcus pneumoniae Isolated  from a Respiratory Infection.
<3>Genome Announcements
<4>2
<5>e00822-14
<6>2014
<7>Next-generation sequencing was used to investigate an unknown clinical respiratory infection.
This new strain of Streptococcus pneumoniae, ASVL_JC_0001,
was isolated from a clinical specimen from a patient with bronchitis and
pulmonary inflammation. The draft genome sequence, obtained with an Illumina
MiSeq sequencing system, consists of 83 large contigs, a total of 2,092,532 bp
long, and has a GC content of 40.3%.

<>

<1>Choi, S.H., Heo, K., Byun, H.M., An, W., Lu, W., Yang, A.S.
<2>Identification of preferential target sites for human DNA methyltransferases.
<3>Nucleic Acids Res.
<4>39
<5>104-118
<6>2011
<7>DNA methyltransferases (DNMTs) play an important role in establishing and maintaining DNA
methylation. Aberrant expression of DNMTs and their
isoforms has been found in many types of cancer, and their contribution to
aberrant DNA methylation has been proposed. Here, we generated HEK 293T
cells stably transfected with each of 13 different DNMTs (DNMT1, two
DNMT3A isoforms, nine DNMT3B isoforms and DNMT3L) and assessed the DNA
methylation changes induced by each DNMT. We obtained DNA methylation
profiles of DNA repetitive elements and 1505 CpG sites from 808
cancer-related genes. We found that DNMTs have specific and overlapping
target sites and their DNA methylation target profiles are a reflection of
the DNMT domains. By examining H3K4me3 and H3K27me3 modifications in the
808 gene promoter regions using promoter ChIP-on-chip analysis, we found
that specific de novo DNA methylation target sites of DNMT3A1 are
associated with H3K4me3 modification that are transcriptionally active,
whereas the specific target sites of DNMT3B1 are associated with H3K27me3
modification that are transcriptionally inactive. Our data suggest that
different DNMT domains are responsible for targeting DNA methylation to
specific regions of the genome, and this targeting might be associated
with histone modifications.

<>

<1>Choi, S.H., Leach, J.E.
<2>Identification of the XorII methyltransferase gene and a vsr homolog from Xanthomonas oryzae pv. oryzae.
<3>Mol. Gen. Genet.
<4>244
<5>383-390
<6>1994
<7>The gene encoding the XorII methyltransferase (M.XorII) was cloned from Xanthomonas oryzae pv.
oryzae and characterized in Escherichia coli. The M.XorII activity was localized to a 3.1 kb
BamHI-BstXI fragment, which contained two open reading frames (ORFs) of 1272 nucleotides (424
amino acids) and 408 nucleotides (136 amino acids). Ten polypeptide domains conserved in other
M5 cytosine methyltransferases (MTases) were identified in the deduced amino acid sequence of
the 1272 ORF. E. coli Mrr+ strains were transformed poorly by plasmids containing the XorII
MTase gene, indicating the presence of at least one MeCG in the recognition sequence for
M.XorII (CGATCG). The 408 nucleotide ORF was 36% identical at the amino acid level to
sequences of the E. coli dcm-vsr gene, which is required for very short patch repair. X.
oryzae pv. oryzae genomic DNA that is resistant to digestion by PvuI and XorII hybridizes with
a 7.0 kb fragment containing the XorII MTase gene and vsr homolog, whereas DNA from strains
that lack M.XorII activity do not hybridize with the fragment.

<>

<1>Choi, S.H., Vera Cruz, C.M., Leach, J.E.
<2>Distribution of Xanthomonas oryzae pv. oryzae DNA modification systems in Asia.
<3>Appl. Environ. Microbiol.
<4>64
<5>1663-1668
<6>1998
<7>The presence or absence of two DNA modification systems, XorI and XorII, in 195 strains of
Xanthomonas oryzae pv. oryzae collected from different major rice-growing countries of Asia
was assessed.  All four possible phenotypes (XorI+ XorII+, XorI+ XorII-, XorI- XorII+ and Xor-
XorII-) were detected in the population at a ratio of approximately 1:2:2:2.  The XorI+ XorII+
and XorI- XorII+ phenotypes were observed predominantly in strains from southeast Asia
(Philippines, Malaysia, and Indonesia), whereas strains with the phenotypes XorI- XorII- and
XorI+ XorII- were distributed in south Asia (India and Nepal) and northeast Asia (China, Korea
and Japan), respectively.  Based on the prevalence and geographic distribution of the XorI and
XorII systems, we suggest that the XorI modification system originated in northeast Asia and
was later introduced to southeast Asia, while the XorII system originated in southeast Asia
and moved to northeast Asia and south Asia.  Genomic DNA from all tested strains of X. oryzae
pv. Oryzae that were resistant to digestion by endonuclease XorII or its isoschizomer PvuI
also hybridized with a 7.0-kb clone that contained the XorII modification system, whereas
strains that were digested by XorII or PvuI lacked DNA that hybridized with the clone.  Size
polymorphisms were observed in fragments that hybridized with the 7.0-kb clone.  However, a
single hybridization pattern generally was found in XorII+ strains within a country,
indicating clonal maintenance of the XorII methyltransferase gene locus.  The locus was
monomorphic for X. oryzae pv. Oryzae strains from the Philippines and all strains from
Indonesia and Korea.

<>

<1>Choi, S.K., Jeong, H., Kloepper, J.W., Ryu, C.M.
<2>Genome Sequence of Bacillus amyloliquefaciens GB03, an Active Ingredient of the First Commercial Biological Control Product.
<3>Genome Announcements
<4>2
<5>e01092-14
<6>2014
<7>Bacillus amyloliquefaciens GB03 has been used as a representative commercialized  strain of
the bacilli for biological control against a broad spectrum of plant
pathogens and as a bio-fertilizer to promote growth and yield of field crops for
more than two decades. Herein, we present the genome sequence and a brief
analysis of strain GB03.

<>

<1>Choi, T.J., Park, Y.J., Park, H.J., Jung, H.K.
<2>Dna adenine methyltransferase gene isolated from feldmannia species virus(fsv) and use thereof as dna methylation enzyme, restriction enzyme or antibiotics - involving isolation of adenine methyltransferase gene from feldmannia species virus.
<3>Korean Patent Office
<4>KR 2007062022
<5>
<6>2007
<7>DERWENT ABSTRACT: NOVELTY - A novel DNA adenine methyltransferase gene from feldmannia species
virus(FsV) and use thereof are provided to
separate DNA adenine methyltransferase present in eukaryotes which
participates in DNA methylation, so that the DNA adenine
methyltransferase is useful as a DNA methylation enzyme, restriction
enzyme or antibiotics. DETAILED DESCRIPTION - The DNA adenine
methyltransferase gene has the nucleotide sequence of SEQ ID NO:1 and
is isolated from feldmannia species virus(FsV) which infects feldmannia
as a brown algae by freezing feldmannia with liquid nitrogen, powdering
the frozen feldmannia, storing the feldmannia powder in buffer solution
containing protenase K for 12 hours so as to purely isolate the genomic
DNA from the solution, digesting the purely isolated genomic DNA with
BamH I, synthesizing genomic DNA library by using a cloning kit, and
sequencing the synthesized genomic DNA library.

<>

<1>Choi, W.S., Kang, S.C., Seo, J.S., Yoo, O.J.
<2>Inhibition of SmaI, AvaI, NaeI and XmaI endonuclease activities by the methylation of DNA with HpaII methylase.
<3>Korean J. Microbiol.
<4>24
<5>86-90
<6>1986
<7>The DNA methylated by HpaII methylase was not cleaved by SmaI, AvaI and NaeI
endonucleases.  This experimental data could be interpreted as strong evidence
that SmaI, AvaI and NaeI methylases which yet to be isolated would methylate on
the inmost cytosine nucleotide within their hexameric recognition sequences.
The facts that SmaI, AvaI and NaeI endonucleases cannot cleave the DNA
methylated by HpaII methylase are the valuable informations for protecting DNAs
upon cleavage reactions by SmaI, AvaI and NaeI endonucleases especially for
cDNA insertion experiments into vector DNAs using SmaI, AvaI and NaeI
oligonucleotide linkers.  In the case of XmaI endonuclease, partially cleaved
DNA fragments were observed although the reaction rate was greatly decreased.
This result implies that the methylation site of XmaI methylase which is yet to
be isolated would not be the same as that of HpaII methylase in XmaI sequence.

<>

<1>Choi, Y., Shin, H., Lee, J.H., Ryu, S.
<2>Identification and characterization of a novel flagellum-dependent Salmonella-infecting bacteriophage, iEPS5.
<3>Appl. Environ. Microbiol.
<4>79
<5>4829-4837
<6>2013
<7>A novel flagellatropic phage of Salmonella enterica serovar Typhimurium called
iEPS5 was isolated and characterized. iEPS5 has an icosahedral head and a long
non-contractile tail with a tail fiber. Genome sequencing revealed a
double-stranded DNA of 59,254 bp having 73 open reading frames (ORFs). To
identify the receptor for iEPS5, Tn5 transposon insertion mutants of S.
Typhimurium SL1344 that were resistant to the phage were isolated. All of the
phage-resistant mutants were found to have mutations in genes involved in
flagellar formation, suggesting that the flagellum is the adsorption target of
this phage. Analysis of phage infection using the DeltamotA mutant, which is
flagellated but non-motile, demonstrated the requirement of flagellar rotation
for iEPS5 infection. Further analysis of phage infection using the DeltacheY
mutant revealed that iEPS5 could infect host bacteria only when the flagellum is
rotating counterclockwise (CCW). These results suggested that the CCW-rotating
flagellar filament is essential for phage adsorption and required for successful
infection by iEPS5. In contrast to the well-studied flagellatropic phage Chi,
iEPS5 cannot infect the DeltafliK mutant that makes a polyhook without a
flagellar filament, suggesting that these two flagellatropic phages utilize
different infection mechanisms. Here, we present evidences that iEPS5 may inject
its DNA into the flagellar filament for infection by assessing DNA transfer from
SYBR-gold-labeled iEPS5 to the host bacteria.

<>

<1>Choi, Y.-J., Kim, S.-J., Hwang, H.-Y., Yim, J., Kim, J.-C.
<2>Characterization of restriction endonuclease EagBI from Enterobacter agglomerans CBNU45.
<3>Korean J. Microbiol.
<4>32
<5>91-95
<6>1994
<7>EagBI is a type II restriction endonuclease from Enterobacter agglomerans strain CBNU45
isolated from soil.  EagBI was partially purified by DEAE-cellulose, phosphocellulose P11 and
hydroxylapatite column chromatography.  EagBI recognizes and cleaves the sequence
5'-CGAT/CG-3' and generates 2-base 3'-protruding cohesive ends.  The optimal reaction
conditions of EagBI are 10mM Tris-HCl (pH 7.8), 6-10mM MgCl2 at 37oC.  The enzyme is maximally
active in the absence of NaCl, able to cleave both dam- and dam+ DNAs and sensitive to heat
treatment (at 65oC for 10 min).  Therefore, although EagBI is an isoschizomer of PvuI, it is
more useful than PvuI in respect of the NaCl requirement and heat-stability.

<>

<1>Choi, Y.U., Kwon, Y.K., Ye, B.R., Hyun, J.H., Heo, S.J., Affan, A., Yoon, K.T., Park, H.S., Oh, C., Kang, D.H.
<2>Draft Genome Sequence of Marinobacterium stanieri S30, a Strain Isolated from a Coastal Lagoon in Chuuk State in Micronesia.
<3>J. Bacteriol.
<4>194
<5>1260
<6>2012
<7>In this study, we isolated xylan-degrading bacteria from a coastal lagoon of Micronesia and
identified the bacteria as Marinobacterium stanieri S30. GSFLX 454
pyrosequencing and sequence analysis of the M. stanieri S30 genome generated
4,007 predicted open reading frames (ORFs) that could be candidate genes for
producing enzymes with different catalytic functions.

<>

<1>Chollet, A., Kawashima, E.
<2>DNA containing the base analogue 2-aminoadenine:  preparation, use as hybridization probes and cleavage by restriction endonucleases.
<3>Nucleic Acids Res.
<4>16
<5>305-317
<6>1988
<7>The base analogue 2-aminoadenine (2,6-diaminopurine,D) has been introduced at
selected positions into synthetic oligodeoxyribonucleotides and DNA by the
combined use of chemical and enzymatic methods.  2-aminoadenine substitution
for adenine introduces changes in the minor groove of DNA and creates an
additional hydrogen bond in the Watson-Crick base pair with thymine.
Oligonucleotide hybridization probes containing 2-aminoadenine showed increased
selectivity and hybridization strength during DNA-DNA hybridization to phage or
genomic target DNA.  Properties of the base analogue with respect to DNA
modifying enzymes were examined.  2-aminoadenine was used to probe minor groove
determinants during the treatment of DNA by 12 restriction endonucleases.
Inhibition of cleavage was found for several restriction enzymes.

<>

<1>Chong, S., Shao, Y., Paulus, H., Benner, J., Perler, F.B., Xu, M.-Q.
<2>Protein splicing involving the Saccharomyces cerevisiae VMA intein.
<3>J. Biol. Chem.
<4>271
<5>22159-22168
<6>1996
<7>Protein splicing involves the excision of an internal protein segment, the intein, from a
precursor protein and the concomitant ligation of the flanking N- and C-terminal regions.  It
occurs in mesophilic bacteria, yeast, and thermophilic archaea.  The ability to control
protein splicing of a thermophilic intein by temperature and pH in a foreign protein context
facilitated the study of the mechanism of protein splicing in thermophiles.  On the other
hand, no direct studies have been done on the mechanism of protein splicing in mesophiles.  We
examined the splicing of a chimeric protein containing the intein of the vacuolar ATPase
subunit (VMA) of Saccharomyces cerevisiae that involves cysteines rather than serines at the
reaction center.  The steps in the splicing process were deduced by analyzing intermediates
and side products that accumulate as a result of amino acid substitutions and were found to be
analogous to those occurring in thermophiles.  Moreover, appropriate amino acid replacements
allowed us to develop the first mesophilic in vitro protein splicing system as well as
strategies for modulating the rate of protein splicing and for converting the splicing
reaction to an efficient protein cleavage reaction at either splice junction.

<>

<1>Chong, T.M., Tung, H.J., Yin, W.F., Chan, K.G.
<2>Insights from the Genome Sequence of Quorum-Quenching Staphylococcus sp. Strain AL1, Isolated from Traditional Chinese Soy Sauce Brine Fermentation.
<3>J. Bacteriol.
<4>194
<5>6611-6612
<6>2012
<7>We report the draft genome sequence of Staphylococcus sp. strain AL1, which degrades
quorum-sensing molecules (namely, N-acyl homoserine lactones). To the
best of our knowledge, this is the first documentation that reports the whole
genome sequence and quorum-quenching activity of Staphylococcus sp. strain AL1.

<>

<1>Chong, T.M., Yin, W.F., Mondy, S., Grandclement, C., Dessaux, Y., Chan, K.G.
<2>Heavy-Metal Resistance of a France Vineyard Soil Bacterium, Pseudomonas mendocina Strain S5.2, Revealed by Whole-Genome Sequencing.
<3>J. Bacteriol.
<4>194
<5>6366
<6>2012
<7>Here we present the draft genome of Pseudomonas mendocina strain S5.2, possessing tolerance to
a high concentration of copper. In addition to being copper
resistant, the genome of P. mendocina strain S5.2 contains a number of
heavy-metal-resistant genes known to confer resistance to multiple heavy-metal
ions.

<>

<1>Choo, S.W., Wong, Y.L., Beh, C.Y., Lokanathan, N., Leong, M.L., Ong, C.S., Ng, K.P., Ngeow, Y.F.
<2>Whole-Genome Shotgun Sequencing of Mycobacterium abscessus M156, an Emerging Clinical Pathogen in Malaysia.
<3>Genome Announcements
<4>1
<5>e00063-12
<6>2013
<7>Mycobacterium abscessus is an emerging clinical pathogen commonly associated with
non-tuberculous mycobacterial infections. We report herein the draft genome of M. abscessus
strain M156.

<>

<1>Choo, S.W., Wong, Y.L., Leong, M.L., Heydari, H., Ong, C.S., Ng, K.P., Ngeow, Y.F.
<2>Analysis of the Genome of Mycobacterium abscessus Strain M94 Reveals an Uncommon  Cluster of tRNAs.
<3>J. Bacteriol.
<4>194
<5>5724
<6>2012
<7>Mycobacterium abscessus is a species of rapidly growing nontuberculous mycobacteria that is
frequently associated with opportunistic infections in
humans. Here, we report the annotated genome sequence of M. abscessus strain M94,
which showed an unusual cluster of tRNAs.

<>

<1>Choo, S.W., Wong, Y.L., Tan, J.L., Ong, C.S., Wong, G.J., Ng, K.P., Ngeow, Y.F.
<2>Annotated Genome Sequence of Mycobacterium massiliense Strain M154, Belonging to  the Recently Created Taxon Mycobacterium abscessus subsp. bolletii comb. nov.
<3>J. Bacteriol.
<4>194
<5>4778
<6>2012
<7>Mycobacterium massiliense has recently been proposed as a member of Mycobacterium abscessus
subsp. bolletii comb. nov. Strain M154, a clinical isolate from the
bronchoalveolar lavage fluid of a Malaysian patient presenting with lower
respiratory tract infection, was subjected to shotgun DNA sequencing with the
Illumina sequencing technology to obtain whole-genome sequence data for
comparison with other genetically related strains within the M. abscessus species
complex.

<>

<1>Choo, S.W., Wong, Y.L., Yusoff, A.M., Leong, M.L., Wong, G.J., Ong, C.S., Ng, K.P., Ngeow, Y.F.
<2>Genome Sequence of the Mycobacterium abscessus Strain M93.
<3>J. Bacteriol.
<4>194
<5>3278
<6>2012
<7>Mycobacterium abscessus is a rapid-growing species of nontuberculous mycobacteria that is
frequently associated with opportunistic infections in humans. We report
herein the draft genome sequence of M. abscessus strain M93.

<>

<1>Choo, S.W., Yusoff, A.M., Wong, Y.L., Wee, W.Y., Ong, C.S., Ng, K.P., Ngeow, Y.F.
<2>Genome Analysis of Mycobacterium massiliense Strain M172, Which Contains a Putative Mycobacteriophage.
<3>J. Bacteriol.
<4>194
<5>5128
<6>2012
<7>The genome of Mycobacterium massiliense M172, isolated from a human sputum sample, was
sequenced using Illumina GA IIX technology and found to contain
5,204,460 bp, including putative genes for virulence and antibiotic resistance as
well as a 92-kb genomic region most likely to correspond to a mycobacteriophage.

<>

<1>Choo, Y.
<2>Recognition of DNA methylation by zinc fingers.
<3>Nat. Struct. Biol.
<4>5
<5>264-265
<6>1998
<7>Zinc fingers are small DNA-binding motifs that occur in a large family of eukaryotic
transcription factors.  The DNA-binding specificity of zinc fingers can be altered by protein
engineering, for instance using phage display, to create novel protein domains which recognize
predetermined sequences.  It has been proposed that tailored DNA-binding domains of this type
can be incorporated into proteins such as restriction enzymes and transcription factors, in
order to target particular DNA sequences or genes.  The zinc finger domains studied so far -
whether naturally occurring, designed or selected - can bind specifically to various DNA sites
containing the four major DNA bases: A, G, C and T.

<>

<1>Choo, Y., Isalan, M.
<2>Nucleic acid binding proteins.
<3>International Patent Office
<4>WO 9947656
<5>
<6>1999
<7>The invention provides a method for producing a zinc finger polypeptide which binds to a
target nucleic acid sequence containing a modified base but not to an identical sequence
containing an equivalent unmodified base.

<>

<1>Choo, Y., Isalan, M.
<2>Advances in zinc finger engineering.
<3>Curr. Opin. Struct. Biol.
<4>10
<5>411-416
<6>2000
<7>Recently developments have been made in engineering sequence-specific zinc finger DNA-binding
proteins.  Advances in this area will soon make it routine to target proteins to specific DNA
sequences associated with any given gene.  The primary interest is in the regulation of gene
expression using customized transcription factors.  However, modular catalytic domains are
also being developed in order to engineer chimeric proteins with customized restriction
enzyme, methylase and integrase activity.

<>

<1>Chopin, A., Chopin, M.-C., Moillo-Batt, A., Langella, P.
<2>Two plasmid-determined restriction and modification systems in Streptococcus lactis.
<3>Plasmid
<4>11
<5>260-263
<6>1984
<7>Two restriction and modification systems were found in Streptococcus lactis
strain IL594 which was found to contain 9 plasmids designated pIL1 to pIL9.  On
the basis of protoplast-induced curing experiments, we showed that a
restriction and modification system was related to the presence of pIL6 or
pIL7.  The pIL-6-determined restriction and modification system was related to
the presence of pIL6 or pIL7.  The pIL-6-determined restriction and
modification system was confirmed by cotransfer of the plasmid and of the
restriction and modification system to a plasmid-free, nonrestricting
derivative of S. lactis IL594.

<>

<1>Chopin, M.-C., Clier, F., Ehrlich, S., Gautier, M., Schouler, C.
<2>The resistance mechanisms to IC type R/M bacteriophages of lactic acid bacteria.
<3>International Patent Office
<4>WO 9911803
<5>
<6>1999
<7>The invention concerns polypeptides constituting subunits of a resistance mechanism to Ic type
R/M bacteriophages, active against the phages of lactic acid bacteria, and nucleic acid
sequences coding for said polypeptides.  The expression of said polypeptides in lactic acid
bacteria enables to increase resistance to the attacks of bacteriophages.

<>

<1>Chopin, M.-C., Cluzel, P.-J.
<2>DNA fragments coding for a bacteriophage-resistant mechanism.
<3>US Patent Office
<4>US 5629182
<5>
<6>1997
<7>The invention relates to DNA fragments encoding an Abi-type mechanism of resistance to
bacteriophages, which fragments are capable of being obtained by cloning of chromosomal or
plasmid DNA of a bacteriophage-resistant lactic acid bacterial strain, as well as to a
polypeptide involved in an Abi-type resistance mechanism, encoded by one of the said
fragments.  The invention also encompasses recombinant vectors and transformed bacterial
strains comprising the said DNA fragments.

<>

<1>Chou, L.F., Chen, Y.T., Lu, C.W., Ko, Y.C., Tang, C.Y., Pan, M.J., Tian, Y.C., Chiu, C.H., Hung, C.C., Yang, C.W.
<2>Sequence of Leptospira santarosai serovar Shermani genome and prediction of virulence-associated genes.
<3>Gene
<4>511
<5>364-370
<6>2012
<7>Leptospirosis, a widespread zoonosis, is a re-emerging infectious disease caused
by pathogenic Leptospira species. In Taiwan, Leptospira santarosai serovar
Shermani is the most frequently isolated serovar, causing both renal and systemic
infections. This study aimed to generate a L. santarosai serovar Shermani genome
sequence and categorize its hypothetical genes, particularly those associated
with virulence. The genome sequence consists of 3,936,333 nucleotides and 4033
predicted genes. Additionally, 2244 coding sequences could be placed into
clusters of orthologous groups and the number of genes involving cell
wall/membrane/envelope biogenesis and defense mechanisms was higher than that of
other Leptospira spp. Comparative genetic analysis based on BLASTX data revealed
that about 73% and 68.8% of all coding sequences have matches to pathogenic L.
interrogans and L. borgpetersenii, respectively, and about 57.6% to saprophyte L.
biflexa. Among the hypothetical proteins, 421 have a transmembrane region, 172
have a signal peptide and 17 possess a lipoprotein signature. According to PFAM
prediction, 32 hypothetical proteins have properties of toxins and surface
proteins mediated bacterial attachment, suggesting they may have roles associated
with virulence. The availability of the genome sequence of L. santarosai serovar
Shermani and the bioinformatics re-annotation of leptospiral hypothetical
proteins will facilitate further functional genomic studies to elucidate the
pathogenesis of leptospirosis and develop leptospiral vaccines.

<>

<1>Chouaia, B., Crotti, E., Brusetti, L., Daffonchio, D., Essoussi, I., Nouioui, I., Sbissi, I., Ghodhbane-Gtari, F., Gtari, M., Vacherie, B., Barbe, V., Medigue, C., Gury, J., Pujic, P., Normand, P.
<2>Genome Sequence of Blastococcus saxobsidens DD2, a Stone-Inhabiting Bacterium.
<3>J. Bacteriol.
<4>194
<5>2752-2753
<6>2012
<7>Members of the genus Blastococcus have been isolated from sandstone monuments, as well as from
sea, soil, plant, and snow samples. We report here the genome
sequence of a member of this genus, Blastococcus saxobsidens strain DD2, isolated
from below the surface of a Sardinian wall calcarenite stone sample.

<>

<1>Choudhary, M., Mackenzie, C., Nereng, K., Sodergren, E., Weinstock, G.M., Kaplan, S.
<2>Low-resolution sequencing of Rhodobacter sphaeroides 2.4.1T: chromosome II is a true chromosome.
<3>Microbiology
<4>143
<5>3085-3099
<6>1997
<7>The photosynthetic bacterium Rhodobacter sphaeroides 2.4.1T has two chromosomes, CI
(approximately 3.0 Mb) and CII (approximately 0.9 Mb). In this study a low-redundancy
sequencing strategy was adopted to analyse 23 out of 47 cosmids from an ordered CII library.
The sum of the lengths of these 23 cosmid inserts was approximately 495 kb, which comprised
approximately 417 kb of unique DNA. A total of 1145 sequencing runs was carried out, with each
run generating 559 +/- 268 bases of sequence to give approximately 640 kb of total sequence.
After editing, approximately 2.8% bases per run were estimated to be ambiguous. After the
removal of vector and Escherichia coli sequences, the remaining approximately 565 kb of R.
sphaeroides sequences were assembled, generating approximately 291 kb of unique sequences.
BLASTX analysis of these unique sequences suggested that approximately 131 kb (45% of the
unique sequence) had matches to either known genes, or database ORFs of hypothetical or
unknown function (dORFs). A total of 144 strong matches to the database was found; 101 of
these matches represented genes encoding a wide variety of functions, e.g. amino acid
biosynthesis, photosynthesis, nutrient transport, and various regulatory functions. Two rRNA
operons (rrnB and rrnC) and five tRNAs were also identified. The remaining 160 kb of DNA
sequence which did not yield database matches was then analysed using CODONPREFERENCE from the
GCG package. This analysis suggested that 122 kb (42% of the total unique DNA sequence) could
encode putative ORFs (pORFs), with the remaining 38 kb (13%) possibly representing non-coding
intergenic DNA. From the data so far obtained, CII does not appear to be specialized for
encoding any particular metabolic function, physiological state or growth condition. These
data suggest that CII contains genes which are functionally as diverse as those found on any
other bacterial chromosome and also contains sequences (pORFs), which may prove to be unique
to this organism.

<>

<1>Choudhary, M., Zanhua, X., Fu, Y.X., Kaplan, S.
<2>Genome Analyses of Three Strains of Rhodobacter sphaeroides: Evidence of Rapid Evolution of Chromosome II.
<3>J. Bacteriol.
<4>189
<5>1914-1921
<6>2007
<7>Three strains of Rhodobacter sphaeroides of diverse origin have been under investigation in
our laboratory for their genome complexities, including
the presence of multiple chromosomes and the distribution of essential
genes within their genomes. The genome of R. sphaeroides 2.4.1 has been
completely sequenced and fully annotated, and now two additional strains
(ATCC 17019 and ATCC 17025) of R. sphaeroides have been sequenced. Thus,
genome comparisons have become a useful approach in determining the
evolutionary relationships among different strains of R. sphaeroides. In
this study, the concatenated chromosomal sequences from the three strains
of R. sphaeroides were aligned, using Mauve, to examine the extent of
shared DNA regions and the degree of relatedness among their
chromosome-specific DNA sequences. In addition, the exact intra- and
interchromosomal DNA duplications were analyzed using Mummer. Genome
analyses employing these two independent approaches revealed that strain
ATCC 17025 diverged considerably from the other two strains, 2.4.1 and
ATCC 17029, and that the two latter strains are more closely related to
one another. Results further demonstrated that chromosome II
(CII)-specific DNA sequences of R. sphaeroides have rapidly evolved, while
CI-specific DNA sequences have remained highly conserved. Aside from the
size variation of CII of R. sphaeroides, variation in sequence lengths of
the CII-shared DNA regions and their high sequence divergence among
strains of R. sphaeroides suggest the involvement of CII in the evolution
of strain-specific genomic rearrangements, perhaps requiring strains to
adapt in specialized niches.

<>

<1>Choudhry, S.
<2>Characterization of new type II restriction endonucleases from bacteria.
<3>Proc. Pakistan Acad. Sci.
<4>36
<5>165-171
<6>1999
<7>The aim of the present study was to search the available microflora of Pakistan for new type
II restriction endonucleases.  Twelve bacterial strains from the American Type Culture
Collection and CAMB Culture Collection were screened for the presence of new type II
restriction endonucleases.  Two strains, Arthrobacter picolinophilus (ATCC 27854) and
Pseudomonas aeruginosa Q2 (CAMB 2637) yielded the endonucleases.  The type II restriction
enzymes isolated and characterized from these strains are designated as ApiI and PaeQI.  These
enzymes were purified by a combination of gel filtration, ion exchange and affinity
chromatography.  The DNA sequences recognized by the two enzymes were determined by analysis
of the fragment patterns generated on different substrate DNAs, Lambda, T7, pUC19, pBR322,
PhiX174RFI.  The cleavage sites within the recognition sequences were also established by
primed synthesis.  The newly discovered enzymes ApiI and PaeQI recognize and cleave 4-6
nucleotide long DNA sequences 5'-CTGCAG-3', 5'-CCGCGG-3', respectively.

<>

<1>Choudhury, J.D., Pramanik, A., Webster, N.S., Llewellyn, L.E., Gachhui, R., Mukherjee, J.
<2>Draft Genome Sequence of Pseudoalteromonas sp. Strain NW 4327 (MTCC 11073, DSM 25418), a Pathogen of the Great Barrier Reef Sponge Rhopaloeides odorabile.
<3>Genome Announcements
<4>2
<5>e00001-14
<6>2014
<7>To date, only one marine sponge pathogen (Pseudoalteromonas sp. strain NW 4327) has fulfilled
Koch's postulates. We report the 4.48-Mbp draft genome sequence of
this strain, which is pathogenic to the Great Barrier Reef sponge Rhopaloeides
odorabile. The sequence provides valuable information on sponge-pathogen
interactions, including the mode of transmission and associated virulence
factors.

<>

<1>Choudhury, K., Leibowitz, M.J.
<2>Pentamidine-induced Alteration in Restriction Endonuclease Cleavage of Plasmid DNA.
<3>J. Biomol. Struct. Dyn.
<4>21
<5>127-134
<6>2003
<7>We have used restriction enzymes and DNaseI as probes to determine the specificity of
pentamidine binding to plasmid DNA. Cleavage of plasmid
pAZ130 by EcoRI, EcoRV and ApaI is inhibited by pentamidine, cleavage by
XbaI, NotI and AvaI is unaffected, while cleavage by XhoI, which
recognizes the same sequence as AvaI, is stimulated. DNaseI footprinting
of DNA containing these restriction sites revealed that pentamidine
protection is not strictly limited to AT-rich regions. We suggest that
perturbation of the DNA micro- environment by pentamidine binding is
responsible for its effect on nucleases.

<>

<1>Choulika, A., Perrin, A., Dujon, B., Nicolas, J.
<2>Nucleotide sequence encoding the enzyme SceI and the uses thereof.
<3>Japanese Patent Office
<4>JP 2007014347 A
<5>
<6>2007
<7>
<>

<1>Choulika, A., Perrin, A., Dujon, B., Nicolas, J.
<2>Nucelotide sequence encoding the enzyme SceI and the uses thereof.
<3>Japanese Patent Office
<4>JP 2006020640 A
<5>
<6>2006
<7>
<>

<1>Choulika, A., Perrin, A., Dujon, B., Nicolas, J.-F.
<2>Induction of homologous recombination in mammalian chromosomes by using the I-SceI system of Saccharomyces cerevisiae.
<3>Mol. Cell. Biol.
<4>15
<5>1968-1973
<6>1995
<7>The mitochondrial intron-encoded endonuclease I-SceI of Saccharomyces cerevisiae has an 18-bp
recognition sequence and, therefore, has a very low probability of cutting DNA, even within
large genomes.  We demonstrate that double-strand breaks can be initiated by the I-SceI
endonuclease at a predetermined location in the mouse genome and that the breaks can be
repaired with a donor molecule homologous with regions flanking the breaks.  This induced
homologous recombination is approximately 2 orders of magnitude more frequent than spontaneous
homologous recombination and at least 10 times more frequent than random integration near an
active promoter.  As a consequence of induced homologous recombination, a heterologous novel
sequence can be inserted at the site of the break.  This recombination can occur at a variety
of chromosomal targets in differentiated and multipotential cells.  These results demonstrate
homologous recombination involving chromosomal DNA by the double-strand break repair mechanism
in mammals and show the usefulness of very rare cutter endonucleases, such as I-SceI, for
designing genome rearrangements.

<>

<1>Choulika, A., Perrin, A., Dujon, B., Nicolas, J.-F.
<2>Nucleotide sequence encoding the enzyme I-SceI and the uses thereof.
<3>International Patent Office
<4>WO 9614408
<5>
<6>1996
<7>An isolated DNA encoding the enzyme I-SceI is provided.  The DNA sequence can be incorporated
in cloning and expression vectors, transformed cell lines and transgenic animals.  The vectors
are useful in gene mapping and site directed insertion of genes.

<>

<1>Choulika, A., Perrin, A., Dujon, B., Nicolas, J.-F.
<2>The yeast I-SceI meganuclease induces site-directed chromosomal recombination in mammalian cells.
<3>C.R. Acad. Sci. III
<4>317
<5>1013-1019
<6>1994
<7>Double-strand breaks in genomic DNA stimulate recombination.  Until now it was not possible to
induce in vivo site-directed double-strand breaks in a mammalian chromosomal target.  In this
article we describe the use of I-SceI meganuclease, a very rare cutter yeast endonuclease, to
induce site-directed double-strand breaks mediated recombination.  The results demonstrate the
potential of the I-SceI system for chromosome manipulation in mammalian cells.

<>

<1>Choulika, A., Perrin, A., Epinat, J.C., Zanghellini, A.
<2>Process of writing or rewriting a polynucleotide sequence having a predetermined content of cpg dinucleotides.
<3>International Patent Office
<4>WO 02099105 A
<5>
<6>2002
<7>The present invention is concerned with a process for (re)writing a polynucleotide sequence
containing a coding sequence for a polypeptide, whereby the content of CpG dinucleotides is
adjusted to a predetermined value.  These polynucleotides are useful to increase, stabilize,
silence and/or reduce gene expression, in particular in protein production, to generate
transgenic animal, transgenic plants or to make gene therapy.  Preferably, the present
invention also relates to process for stably expressing these (re)written polynucleotides in
in vivo and in in vivo expression systems.

<>

<1>Chovan, L.E., Hessler, P.E., Reich, K.A.
<2>Essential bacteria genes and genome scanning in Haemophilus influenzae for the identification of "essential genes".
<3>International Patent Office
<4>WO 0218601 A
<5>
<6>2002
<7>Essential bacteria genes and a method for identifying 'essential genes' (i.e., genes which
are essential to a bacterium's survival) using an in vitro transposition system, a small (975
bp) insertional element containing an antibiotic resistance cassette and mapping these inserts
relative to the deduced open reading frames of H. influenzae by PCR and Southern analysis.

<>

<1>Chovan, L.E., Hessler, P.E., Reich, K.A.
<2>Essential bacteria genes and genome scanning in Haemophilus influenzae for the identification of "essential genes".
<3>International Patent Office
<4>WO 0111033 A
<5>
<6>2001
<7>Essential bacteria genes and a method for identifying "essential genes" (i.e., genes which are
essential to a bacterium's survival) using an in vitro transposition system, a small (975 bp)
insertional element containing an antibiotic resistance cassette and mapping these inserts
relative to the deduced open reading frames of H. influenzae by PCR and Southern analysis.

<>

<1>Chovatia, M. et al.
<2>Complete genome sequence of Thermanaerovibrio acidaminovorans type strain (Su883).
<3>Standards in Genomic Sciences
<4>1
<5>254-261
<6>2009
<7>Thermanaerovibrio acidaminovorans (Guangsheng et al. 1997) Baena et al. 1999 is the type
species of the genus Thermanaerovibrio and is of phylogenetic interest
because of the very isolated location of the novel phylum Synergistetes. T.
acidaminovorans Su883(T) is a Gram-negative, motile, non-spore-forming bacterium
isolated from an anaerobic reactor of a sugar refinery in The Netherlands. Here
we describe the features of this organism, together with the complete genome
sequence, and annotation. This is the first completed genome sequence from a
member of the phylum Synergistetes. The 1,848,474 bp long single replicon genome
with its 1765 protein-coding and 60 RNA genes is part of the Genomic Encyclopedia
of Bacteria and Archaea project.

<>

<1>Chow, V. et al.
<2>Complete genome sequence of Paenibacillus sp. strain JDR-2.
<3>Standards in Genomic Sciences
<4>6
<5>1-10
<6>2012
<7>Paenibacillus sp. strain JDR-2, an aggressively xylanolytic bacterium isolated from sweetgum
(Liquidambar styraciflua) wood, is able to efficiently depolymerize, assimilate and metabolize
4-O-methylglucuronoxylan, the predominant structural component of hardwood hemicelluloses. A
basis for this capability was first supported by the identification of genes and
characterization of encoded enzymes and has been further defined by the sequencing and
annotation of the complete genome, which we describe. In addition to genes implicated in the
utilization of â-1,4-xylan, genes have also been identified for the utilization of other
hemicellulosic polysaccharides. The genome of Paenibacillus sp. JDR-2 contains 7,184,930 bp in
a single replicon with 6,288 protein-coding and 122 RNA genes. Uniquely prominent are 874
genes encoding proteins involved in carbohydrate transport and metabolism. The prevalence and
organization of these genes support a metabolic potential for bioprocessing of hemicellulose
fractions derived from lignocellulosic resources.

<>

<1>Chowdhury, P.R., Boucher, Y., Hassan, K.A., Paulsen, I.T., Stokes, H.W., Labbate, M.
<2>Genome sequence of Vibrio rotiferianus strain DAT722.
<3>J. Bacteriol.
<4>193
<5>3381-3382
<6>2011
<7>Vibrio rotiferianus is a marine pathogen capable of causing disease in various aquatic
organisms. We announce the genome sequence of V.
rotiferianus DAT722, which has a large chromosomal integron containing 116
gene cassettes and is a model organism for studying the role of this
system in vibrio evolution.

<>

<1>Chowdhury, P.R., Ingold, A., Vanegas, N., Martinez, E., Merlino, J., Merkier, A.K., Castro, M., Rocha, G.G., Borthagaray, G., Centron, D., Toledo, H.B., Marquez, C.M., Stokes, H.W.
<2>Dissemination of multiple drug resistance genes by class 1 integrons in Klebsiella pneumoniae isolates from four countries: a comparative study.
<3>Antimicrob. Agents Chemother.
<4>55
<5>3140-3149
<6>2011
<7>A comparative genetic analysis of 42 clinical Klebsiella pneumoniae isolates, resistant to two
or more antibiotics belonging to the broad-spectrum beta-lactam group, sourced from Sydney,
Australia, and three South American countries is presented. The study focuses on the genetic
contexts of class 1 integrons, mobilizable genetic elements best known for their role in the
rapid evolution of antibiotic resistance among Gram-negative pathogens. It was found that the
class 1 integrons in this cohort were located in a number of different genetic contexts with
clear regional differences. In Sydney, IS26-associated Tn21-like transposons on IncL/M
plasmids contribute greatly to the dispersal of integron-associated multiple-drug-resistant
(MDR) loci. In contrast, in the South American countries, Tn1696-like transposons on an IncA/C
plasmid(s) appeared to be disseminating a characteristic MDR region. A range of mobile genetic
elements is clearly being recruited by clinically important mobile class 1 integrons, and
these elements appear to be becoming more common with time. This in turn is driving the
evolution of complex and laterally mobile MDR units and may further complicate antibiotic
therapy.

<>

<1>Christ, C., Nelson, M.
<2>DNA modification methylases:  New uses in the manipulation of DNA.
<3>Biotechniques
<4>2
<5>218-221
<6>1984
<7>None

<>

<1>Christ, F., Schoettler, S., Wende, W., Steuer, S., Pingoud, A., Pingoud, V.
<2>The monomeric homing endonuclease PI-SceI has two catalytic centres for cleavage of the two strands of its DNA substrate.
<3>EMBO J.
<4>18
<5>6908-6916
<6>1999
<7>The monomeric homing endonuclease PI-SceI cleaves the two strands of its DNA substrate in a
concerted manner, which raises the question of whether this enzyme harbours one or two
catalytic centres. If PI-SceI has only one catalytic centre, one would expect that
cross-linking enzyme and substrate should prevent reorientation of the enzyme required to
perform the second cut after having made the first cut: PI-SceI, however, when cross-linked to
its substrate, is able to cleave both DNA strands. If PI-SceI has two catalytic centres, one
would expect that it should be possible to inactivate one catalytic centre by mutation and
obtain a variant with preference for a substrate nicked in one strand; such variants have been
found. The structural homology between the catalytic domain of PI-SceI having a pseudo 2-fold
symmetry, and I-CreI, a homodimeric homing endonuclease, suggests that in PI-SceI active site
I, which attacks the top strand, comprises Asp218, Asp229 and Lys403, while Asp326, Thr341 and
Lys301 make up active site II, which cleaves the bottom strand. Cleavage experiments with
modified oligodeoxynucleotides and metal ion mapping experiments demonstrate that PI-SceI
interacts differently with the two strands at the cleavage position, supporting a model of two
catalytic centres.

<>

<1>Christ, F., Steuer, S., Thole, H., Wende, W., Pingoud, A., Pingoud, V.
<2>A model for the PI-SceIxDNA complex based on multiple base and phosphate backbone-specific photocross-links.
<3>J. Mol. Biol.
<4>300
<5>867-875
<6>2000
<7>We have synthesized different oligodeoxynucleotides carrying, in single positions of the >36
bp recognition site of PI-SceI, photoreactive base analogues (5-iododeoxypyrimidines) or
phosphate modifications (p-azidophenacylphosphorothioates) and used them in photocross-linking
experiments with PI-SceI to probe the protein-DNA interface of the specific complex between
the homing endonuclease PI-SceI and its DNA substrate. One base-specific and several
backbone-specific cross-links were analyzed in detail: the cross-linking positions were
identified by Edman degradation of isolated cross-linked peptide x oligodeoxynucleotide
adducts
and confirmed by site-directed mutagenesis. Based on these results and the crystal structure
of PI-SceI, a model for the structure of the PI-SceI x DNA complex is proposed.

<>

<1>Christensen, L.L., Josephsen, J.
<2>The methyltransferase from the LlaDII restriction-modification system influences the level of expression of its own gene.
<3>J. Bacteriol.
<4>186
<5>287-295
<6>2004
<7>The type II restriction-modification (R-M) system LlaDII isolated from Lactococcus lactis
contains two tandemly arranged genes, llaDIIR and llaDIIM, encoding a restriction endonuclease
(REase) and a methyltransferase (MTase), respectively. Interestingly, two LlaDII recognition
sites are present in the llaDIIM promoter region, suggesting that they may influence the
activity of the promoter through methylation status. In this study, separate promoters for
llaDIIR and llaDIIM were identified, and the regulation of the two genes at the
transcriptional level was investigated. DNA fragments containing the putative promoters were
cloned in a promoter probe vector and tested for activity in the presence and absence of the
active MTase. The level of expression of the MTase was 5- to 10-fold higher than the level of
expression of the REase. The results also showed that the presence of M.LlaDII reduced the in
vivo expression of the llaDIIM promoter (P(llaDIIM)) up to 1,000-fold, whereas the activity of
the llaDIIR promoter (P(llaDIIR)) was not affected. Based on site-specific mutations it was
shown that both of the LlaDII recognition sites within P(llaDIIM) are required to obtain
complete repression of transcriptional activity. No regulation was found for llaDIIR, which
appears to be constitutively expressed.

<>

<1>Christian, V.D., Machetti, A., Kula, S., Bengezaru, M., Kosson, P.
<2>Pseudomonas Aeruginosa and Klebsiella virulence genes, proteins, and their use.
<3>Japanese Patent Office
<4>JP 2006524984 A
<5>
<6>2006
<7>
<>

<1>Christman, J.K., Sheikhnejad, G.
<2>Substrate for detection of mammalian 5-C-DNA methyltransferase.
<3>European Patent Office
<4>EP 0756008 A
<5>
<6>1995
<7>A substrate selective for detection of mammalian 5-C-DNA methyltransferase in the presence of
bacterial 5-C-DNA methyltransferase, said substrate comprising oligomeric DNA which contains
at least one 5-methylcytosine residue, and at least one cytosine or 5-fluorocytosine residue,
each of which are followed in linkage to a guanine residue.  The invention also includes a
method for measuring the presence of mammalian 5-C-DNA methyltransferase which comprises
contacting a sample containing 5-C-DNA methyltransferase with the substrate and also includes
a method for inhibiting mammalian 5-C-DNA methyltransferase which comprises contacting a
sample containing 5-C-DNA methyltransferase with the substrate.

<>

<1>Christman, J.K., Sheikhnejad, G., Marasco, C.J., Sufrin, J.R.
<2>5-methyl-2'-deoxycytidine in single-stranded DNA can act in cis to signal de novo DNA methylation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>92
<5>7347-7351
<6>1995
<7>Methylation of cytosine residues in DNA plays an important role in regulating gene expression
during vertebrate embryonic development.  Conversely, disruption of normal patterns of
methylation is common in tumors and occurs early in progression of some human cancers.  In
vertebrates, it appears that the same DNA methyltransferase maintains preexisting patterns of
methylation during DNA replication and carries out de novo methylation to create new
methylation patterns.  There are several indications that inherent signals in DNA structure
can act in vivo to initiate or block de novo methylation in adjacent DNA regions.  To identify
sequences capable of enhancing de novo methylation of DNA in vitro, we designed a series of
oligodeoxyribonucleotide substrates with substrate cytosine residues in different sequence
contexts.  We obtained evidence that some 5-methylcytosine residues in these single-stranded
DNAs can stimulate de novo methylation of adjacent sites by murine DNA 5-cytosine
methyltransferase as effectively as 5-methylcytosine residues in double-stranded DNA stimulate
maintenance methylation.  This suggests that double-stranded DNA may not be the primary
natural substrate for de novo methylation and that looped single-stranded structures formed
during the normal course of DNA replication or repair serve as "nucleation" sites for de novo
methylation of adjacent DNA regions.

<>

<1>Christopher, L.P., Kapatral, V., Vaisvil, B., Emel, G., Deveaux, L.C.
<2>Draft Genome Sequence of a New Homofermentative, Lactic Acid-Producing Enterococcus faecalis Isolate, CBRD01.
<3>Genome Announcements
<4>2
<5>e00147-14
<6>2014
<7>We report here the draft genome sequence of the novel homofermentative Enterococcus faecalis
isolate CBRD01, which is capable of high lactic acid
productivity and yields, with minimal nutritional requirements. The genome is 2.8
Mbp, with 37% G+C, and contains genes for two lactate dehydrogenase (LDH) enzymes
found in related organisms.

<>

<1>Chu, F.K., Maley, F., Wang, A.-M., Pedersen-Lane, J., Maley, G.
<2>Purification and substrate specificity of a T4 phage intron-encoded endonuclease.
<3>Nucleic Acids Res.
<4>19
<5>6863-6869
<6>1991
<7>The T4 phage td intron-encoded endonuclease (I-Tev I) cleaves the intron-deleted
td gene (td-delta-I) 23 nucleotides upstream of the intron insertion site on the noncoding
strand and
25 nucleotides upstream of this site on the coding strand, to generate a 2-base hydroxyl
overhang
in the 3' end of each DNA strand.  I-Tev I-157, a truncated form in which slightly more than
one
third (88 residues) of the endonuclease is deleted, was purified to homogeneity and shown to
possess endonuclease activity similar to that of I-Tev I, the full-length enzyme (245
residues).  The
minimal length of the td-delta-I gene that was cleaved by I-Tev I and I-Tev I-157 has been
determined to be exactly 39 basepairs, from -27 (upstream in exon1) to +12 (downstream in
exon2) relative to the intron insertion site.  Similar to the full-length endonuclease, I-Tev
I-157 cuts
the intronless thymidylate synthase genes from such diverse organisms as Escherichia coli,
Lactobacillus casei and the human.  The position and nature of the in vitro endonucleolytic
cut in
these genes are homologous to those in td-delta-I.  Point mutational analysis of the
td-delta-I
substrate based on the deduced consensus nucleotide sequence has revealed a very low degree of
specificity on either side of the cleavage site, for both the full-length and truncated I-Tev
I.

<>

<1>Chu, F.K., Maley, G., Pedersen-Lane, J., Wang, A.-M., Maley, F.
<2>Characterization of the restriction site of a prokaryotic intron-encoded endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>87
<5>3574-3578
<6>1990
<7>The 1016-base-pair (bp) intron in the T4 bacteriophage thymidylate
synthase gene (td) contains a 735-bp open reading frame that encodes a protein product
with endonucleolytic activity.  The endonuclease shows specificity for the intronless form
of the td gene.  Highly purified endonuclease cleaves the DNA of the intronless form of the
td gene in vitro at 24 bp upstream of the exon 1-exon 2 junction, generating a 2-base
staggered cut with 3'-hydroxyl overhangs.  Although the endonuclease cleaves in exon 1, it
requires some exon 2 sequence for recognition.  The maximum recognition sequence lies in
an 87-bp stretch, from 52 bp upstream to 35 bp downstream of the cleavage site, ending at
11 bp into exon 2.  The td intron endonuclease appears involved in the conversion of the
intronless form of td to intron-containing td gene in the T-even phages.  A role for intron
mobility is discussed.

<>

<1>Chu, F.K., Maley, G.F., Maley, F., Belfort, M.
<2>Intervening sequence in the thymidylate synthase gene of bacteriophage T4.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>81
<5>3049-3053
<6>1984
<7>The continuous sequence of 2.3 kilobases in a 3-kilobase DNA fragment encoding the structural
gene for coliphage T4 thymidylate synthase (5,10-methylenetetrahydrofolate:dUMP
C-methyltranferase, EC 2.1.1.45) was determined by using the M13 dideoxy chain-termination
method. From the coding information within this gene and that provided by sequence analysis of
selected CNBr peptides from the protein product, the primary structure of T4 thymidylate
synthase was determined. The most significant finding of these studies is the presence of a
1017-base-pair interruption two-thirds of the way through the nucleotide sequence of the
structural gene. The 5'- and 3'-terminal ends of this intron are demarcated by an apparent
stop and start codon, respectively. The corresponding methionine preceding the second coding
region of the synthase is not incorporated into the final protein product. Structural evidence
confirming the presence of the intervening sequence in the phage genome was obtained by
restriction and hybridization analysis. Support for the presence of the intron was also
obtained at the functional level by enzyme expression studies using selected td gene
fragments. This work also confirms the findings of Purohit and Mathews, which reveal that the
termination codon for the dihydrofolate reductase gene and the triplet intitiating thymidylate
synthase overlap by a four-base stretch, A-T-G-A. The implications of this unusual gene
arrangement are discussed.

<>

<1>Chu, F.K., Maley, G.F., West, D.K., Belfort, M., Maley, F.
<2>Characterization of the intron in the phage T4 thymidylate synthase gene and evidence for its self-excision from the primary transcript.
<3>Cell
<4>45
<5>157-166
<6>1986
<7>The td gene contains a 735 bp open reading frame within its 1017 bp intron. A 12 nucleotide
stretch may form a stable secondary structure with the putative Shine-Dalgarno sequence of the
intron open reading frame and thus impair its translation. SP6 RNA polymerase transcripts of
the td gene synthesized in vitro at 40oC encompass a 2.7 kb primary transcript, a 1.7 kb mRNA,
and a 1 kb intron RNA. The excised intron RNA consisted of linear and cyclized forms. RNAase H
studies and resistance of the cyclized intron to linearization by HeLa cell debranching enzyme
suggest it to be circular. Self-splicing of isolated td primary transcript occurred only
marginally at 28oC, but increased progressively to 50oC, and required the presence of both
Mg++ and a guanosine cofactor. An internal guide sequence is evident which may align the 5'
splice site with the 3' end, presumably for precise exon ligation.

<>

<1>Chu, Y., Gao, P., Zhao, P., He, Y., Liao, N., Jackman, S., Zhao, Y., Birol, I., Duan, X., Lu, Z.
<2>Genome Sequence of Mycoplasma capricolum subsp. capripneumoniae Strain M1601.
<3>J. Bacteriol.
<4>193
<5>6098-6099
<6>2011
<7>Mycoplasma capricolum subsp. capripneumoniae is the causative agent of contagious caprine
pleuropneumonia, a devastating disease of goats listed
by the World Organization for Animal Health. Here we report the first
complete genome sequence of this organism (strain M1601, a clinically
isolated strain from China).

<>

<1>Chua, K., Seemann, T., Harrison, P.F., Davies, J.K., Coutts, S.J., Chen, H., Haring, V., Moore, R., Howden, B.P., Stinear, T.P.
<2>Complete genome sequence of Staphylococcus aureus strain JKD6159, a unique Australian clone of ST93-IV community methicillin-resistant Staphylococcus  aureus.
<3>J. Bacteriol.
<4>192
<5>5556-5557
<6>2010
<7>Community methicillin-resistant Staphylococcus aureus (cMRSA) is an emerging issue that has
resulted in multiple worldwide epidemics. We
report the first complete genome sequence of an ST93-MRSA-IV clinical
isolate that caused severe invasive infection and a familial outbreak of
skin infection. This isolate is a representative of the most common
Australian clone of cMRSA that is more distantly related to the previously
sequenced genomes of S. aureus.

<>

<1>Chua, P., Yoo, H.S., Gan, H.M., Lee, S.M.
<2>Draft Genome Sequences of Two Cellulolytic Paenibacillus sp. Strains, MAEPY1 and  MAEPY2, from Malaysian Landfill Leachate.
<3>Genome Announcements
<4>2
<5>e00065-14
<6>2014
<7>We report the draft genome sequences of two Paenibacillus species with cellulose-degrading
abilities isolated from landfill leachate. An array of genes
putatively involved in cellulose degradation have been identified in both genome
sequences, which can benefit various biotechnological industries.

<>

<1>Chuang, L.S.-H., Ng, H.-H., Chia, J.-N., Li, B.F.L.
<2>Characterisation of independent DNA and multiple Zn-binding domains at the N terminus of human DNA-(cytosine-5) methyltransferase: Modulating the property of a DNA-binding domain by contiguous Zn-binding motifs.
<3>J. Mol. Biol.
<4>257
<5>935-948
<6>1996
<7>We report here a detailed mapping and characterisation of a DNA-binding domain
at the N terminus of human DNA-(cytosine-5) methyltransferase.  A small region, B1 (codon 202
to 369), was first identified by its Zn- and gross DNA-binding properties.  Further
fine-mapping
using deletion and point mutation analysis shows that the DNA- and Zn-binding domains involve
separate peptide motifs, KRRKTTPKEPTEKK (codons 202 to 215) for a bipartite DNA-binding
oligopeptide (DB1) and Cx2CX13HX2D(X)23EX2EX13CX3H (codons 232 to 297) for possibly
two contiguous Zn-binding domains (Azn), which can function independently.  However, B1
(containing DB1 and AZn) differs from DB1 because it does not bind to a 30 base-pair duplex.
Interestingly, H3 codons 202 to 974, which encloses B1 and B2 (containing the Zn-binding
CX2CX2CX4CX2CX2C motif from codon 533 to 550) binds preferentially to 0.8 kb duplexes,
compared with 0.4 and 0.6 kb duplexes.  As the homologous murine B1, which targets the murine
methylase to replication foci, also binds to DNA and Zn, it is possible that the N terminus of
mammalian methylase may be involved in sensing the appropriate length of newly synthesized
DNA before methylation by its C terminus.  This may enable a time delay for the transient
existence of hemi-methylation sites for their unknown biological functions in mammals.

<>

<1>Chuang, L.S.H., Ian, H.-I., Koh, T.W., Ng, H.-H., Xu, G., Li, B.F.L.
<2>Human DNA-(cytosine-5) methyltransferase-PCNA complex as a target for p21WAF1.
<3>Science
<4>277
<5>1996-2000
<6>1997
<7>DNA-(cytosine-5) methyltransferase (MCMT) methylates newly replicated mammalian DNA, but the
factors regulating this activity are unknown.  Here, MCMT is shown to bind proliferating cell
nuclear antigen, an auxiliary factor for DNA replication and repair.  Binding of PCNA requires
amino acids 163 to 174 of MCMT, occurs in intact cells at foci of newly replicated DNA, and
does not alter MCMT activity.  A peptide derived from the cell cycle regulator p21WAF1 can
disrupt the MCMT-PCNA interaction, which suggests that p21WAF1 may regulate methylation by
blocking access of MCMT to PCNA.  MCMT and p21WAF1 may be linked in a regulatory pathway,
because the extents of their expression are inversely related in both SV40-transformed and
nontransformed cells.

<>

<1>Chuang, L.S.H., Tan, E.H.H., Oh, H.K., Li, B.F.L.
<2>Selective depletion of human DNA-methyltransferase DNMT1 proteins by sulfonate-derived methylating agents.
<3>Cancer Res.
<4>62
<5>1592-1597
<6>2002
<7>5-Methylcytosine residues in the DNA (DNA methylation) are formed from the transfer of the
methyl group from S-adenosylmethionine to the C-5
position of cytosine by the DNA-(cytosine-5) methyltransferases
(DNMTs). Although regional hypermethylation and global hypomethylation
of the genome are commonly observed in neoplastic cells, how these
aberrant methylation patterns occur remains unestablished. We report
here that sulfonate-derived methylating agents, unlike
N-methylnitrosourea or iodomethane, are potent in depleting DNMT1
proteins in human cells, in addition to their DNA-damaging properties.
Their effects on cellular DNMT1 are time and dosage dependent but
independent of cell type. Unlike gamma-irradiation, these agents
apparently do not activate the p53/p21(WAF1) DNA damage response
pathway to deplete the DNMT1 proteins because cells with wild-type,
mutated, or inactivated p53 behave similarly. However, cell cycle
analysis and protease assay studies strongly suggest that
methylmethanesulfonate may activate a cellular protease to degrade
DNMT1. These results explain why reported observations on the effect of
alkylating agents on DNMT1 activities in human cells vary significantly
and provide a crucial link to understand the mechanism behind genomic
hypomethylation.

<>

<1>Chuluunbaatar, T., Ivanenko-Johnston, T., Fuxreiter, M., Meleshko, R., Rasko, T., Simon, I., Heitman, J., Kiss, A.
<2>An EcoRI-RsrI chimeric restriction endonuclease retains parental sequence specificity.
<3>Biochim. Biophys. Acta
<4>1774
<5>583-594
<6>2007
<7>To test their structural and functional similarity, hybrids were constructed between EcoRI and
RsrI, two restriction endonucleases recognizing the same DNA sequence and sharing 50% amino
acid sequence identity. One of the chimeric proteins (EERE), in which the EcoRI segment
His147-Ala206 was replaced with the corresponding RsrI segment, showed EcoRI/RsrI-specific
endonuclease activity. EERE purified from inclusion bodies was found to have approximately
100-fold weaker activity but higher specific DNA binding affinity, than EcoRI. Increased
binding is consistent with results of molecular dynamics simulations, which indicate that the
number of hydrogen bonds formed with the recognition sequence increased in the chimera as
compared to EcoRI. The success of obtaining an EcoRI-RsrI hybrid endonuclease, which differs
from EcoRI by 22 RsrI-specific amino acid substitutions and still preserves canonical cleavage
specificity, is a sign of structural and functional similarity shared by the parental enzymes.
This conclusion is also supported by computational studies, which indicate that construction
of the EERE chimera did not induce substantial changes in the structure of EcoRI.
Surprisingly, the chimeric endonuclease was more toxic to cells not protected by EcoRI
methyltransferase, than the parental EcoRI mutant. Molecular modelling revealed structural
alterations, which are likely to impede coupling between substrate recognition and cleavage
and suggest a possible explanation for the toxic phenotype.

<>

<1>Chun, J.H., Hong, K.J., Cha, S.H., Cho, M.H., Lee, K.J., Jeong, D.H., Yoo, C.K., Rhie, G.E.
<2>Complete Genome Sequence of Bacillus anthracis H9401, an Isolate from a Korean Patient with Anthrax.
<3>J. Bacteriol.
<4>194
<5>4116-4117
<6>2012
<7>Bacillus anthracis H9401 (NCCP 12889) is an isolate from a Korean patient with
gastrointestinal anthrax. The whole genome of H9401 was sequenced. It is a
circular chromosome containing 5,480 open reading frames (ORFs) and two plasmids,
pXO1 containing 202 ORFs and pXO2 containing 110 ORFs. H9401 shows high
pathogenicity and genome sequence similarity to Ames Ancestor.

<>

<1>Chung, D., Cha, M., Farkas, J., Westpheling, J.
<2>Construction of a stable replicating shuttle vector for Caldicellulosiruptor species: use for extending genetic methodologies to other members of this genus.
<3>PLoS ONE
<4>8
<5>e62881
<6>2013
<7>The recalcitrance of plant biomass is the most important barrier to its economic  conversion
by microbes to products of interest. Thermophiles have special
advantages for biomass conversion and members of the genus Caldicellulosiruptor
are the most thermophilic cellulolytic microbes known. In this study, we report
the construction of a replicating shuttle vector for Caldicellulosiruptor species
based on pBAS2, the smaller of two native C. bescii plasmids. The entire plasmid
was cloned into an E. coli cloning vector containing a pSC101 origin of
replication and an apramycin resistance cassette for selection in E. coli. The
wild-type C. bescii pyrF locus was cloned under the transcriptional control of
the regulatory region of the ribosomal protein S30EA (Cbes2105), and the
resulting vector was transformed into a new spontaneous deletion mutant in the
pyrFA locus of C. bescii that allowed complementation with the pyrF gene alone.
Plasmid DNA was methylated in vitro with a recently described cognate
methyltransferase, M.CbeI, and transformants were selected for uracil
prototrophy. The plasmid was stably maintained in low copy with selection but
rapidly lost without selection. There was no evidence of DNA rearrangement during
transformation and replication in C. bescii. A similar approach was used to
screen for transformability of other members of this genus using M.CbeI to
overcome restriction as a barrier and was successful for transformation of C.
hydrothermalis, an attractive species for many applications. Plasmids containing
a carbohydrate binding domain (CBM) and linker region from the C. bescii celA
gene were maintained with selection and were structurally stable through
transformation and replication in C. bescii and E. coli.

<>

<1>Chung, D., Farkas, J., Huddleston, J.R., Olivar, E., Westpheling, J.
<2>Methylation by a Unique alpha-class N4-Cytosine Methyltransferase Is Required for DNA Transformation of Caldicellulosiruptor bescii DSM6725.
<3>PLoS ONE
<4>7
<5>e43844
<6>2012
<7>Thermophilic microorganisms capable of using complex substrates offer special advantages for
the conversion of lignocellulosic biomass to
biofuels and bioproducts. Members of the Gram-positive bacterial genus
Caldicellulosiruptor are anaerobic thermophiles with optimum growth
temperatures between 65 degrees C and 78 degrees C and are the most
thermophilic cellulolytic organisms known. In fact, they efficiently
use biomass non-pretreated as their sole carbon source and in
successive rounds of application digest 70% of total switchgrass
substrate. The ability to genetically manipulate these organisms is a
prerequisite to engineering them for use in conversion of these complex
substrates to products of interest as well as identifying gene products
critical for their ability to utilize non-pretreated biomass. Here, we
report the first example of DNA transformation of a member of this
genus, C. bescii. We show that restriction of DNA is a major barrier to
transformation (in this case apparently absolute) and that methylation
with an endogenous unique alpha-class N4-Cytosine methyltransferase is
required for transformation of DNA isolated from E. coli. The use of
modified DNA leads to the development of an efficient and reproducible
method for DNA transformation and the combined frequencies of
transformation and recombination allow marker replacement between
non-replicating plasmids and chromosomal genes providing the basis for
rapid and efficient methods of genetic manipulation.

<>

<1>Chung, D.H., Huddleston, J.R., Farkas, J., Westpheling, J.
<2>Identification and characterization of CbeI, a novel thermostable restriction enzyme from Caldicellulosiruptor bescii DSM 6725 and a member of a new subfamily of HaeIII-like enzymes.
<3>J. Ind. Microbiol. Biotechnol.
<4>38
<5>1867-1877
<6>2011
<7>Potent HaeIII-like DNA restriction activity was detected in cell-free extracts of
Caldicellulosiruptor bescii DSM 6725 using plasmid DNA
isolated from Escherichia coli as substrate. Incubation of the plasmid
DNA in vitro with HaeIII methyltransferase protected it from cleavage
by HaeIII nuclease as well as cell-free extracts of C. bescii. The gene
encoding the putative restriction enzyme was cloned and expressed in E.
coli with a His-tag at the C-terminus. The purified protein was 38 kDa
as predicted by the 981-bp nucleic acid sequence, was optimally active
at temperatures between 75 degrees C and 85 degrees C, and was stable
for more than 1 week when stored at 35 degrees C. The cleavage sequence
was determined to be 50-GG/CC-30, indicating that CbeI is an
isoschizomer of HaeIII. A search of the C. bescii genome sequence
revealed the presence of both a HaeIII-like restriction endonuclease
(Athe 2438) and DNA methyltransferase (Athe 2437). Preliminary analysis
of other Caldicellulosiruptor species suggested that this
restriction/modification activity is widespread in this genus. A
phylogenetic analysis based on sequence alignment and conserved motif
searches identified features of CbeI distinct from other members of
this group and classified CbeI as a member of a novel subfamily of
HaeIII-like enzymes.

<>

<1>Chung, D.W., Farkas, J., Westpheling, J.
<2>Overcoming restriction as a barrier to DNA transformation in Caldicellulosiruptor species results in efficient marker replacement.
<3>Biotechnol. Biofuels.
<4>6
<5>0
<6>2013
<7>Background: Thermophilic microorganisms have special advantages for the conversion of plant
biomass to fuels and chemicals. Members of the
genus Caldicellulosiruptor are the most thermophilic cellulolytic
bacteria known. They have the ability to grow on a variety of
non-pretreated biomass substrates at or near similar to 80 degrees C
and hold promise for converting biomass to bioproducts in a single
step. As for all such relatively uncharacterized organisms with
desirable traits, the ability to genetically manipulate them is a
prerequisite for making them useful. Metabolic engineering of pathways
for product synthesis is relatively simple compared to engineering the
ability to utilize non-pretreated biomass.
Results: Here we report the construction of a deletion of cbeI
(Cbes2438), which encodes a restriction endonuclease that is as a major
barrier to DNA transformation of C. bescii. This is the first example
of a targeted chromosomal deletion generated by homologous
recombination in this genus and the resulting mutant, JWCB018 (Delta
pyrFA Delta cbeI), is readily transformed by DNA isolated from E. coli
without in vitro methylation. PCR amplification and sequencing
suggested that this deletion left the adjacent methyltransferase
(Cbes2437) intact. This was confirmed by the fact that DNA isolated
from JWCB018 was protected from digestion by CbeI and HaeIII. Plasmid
DNA isolated from C. hydrothermalis transformants were readily
transformed into C. bescii. Digestion analysis of chromosomal DNA
isolated from seven Caldicellulosiruptor species by using nine
different restriction endonucleases was also performed to identify the
functional restriction-modification activities in this genus.
Conclusion: Deletion of the cbeI gene removes a substantial barrier
to routine DNA transformation and chromosomal modification of C.
bescii. This will facilitate the functional analyses of genes as well
as metabolic engineering for the production of biofuels and bioproducts
from biomass. An analysis of restriction-modification activities in
members of this genus suggests a way forward to eliminating restriction
as a barrier to DNA transformation and efficient genetic manipulation
of this important group of hyperthermophiles.

<>

<1>Chung, G.T., Yoo, J.S., Oh, H.B., Lee, Y.S., Cha, S.H., Kim, S.J., Yoo, C.K.
<2>The Complete Genome Sequence of Neisseria gonorrhoeae NCCP11945.
<3>J. Bacteriol.
<4>190
<5>6035-6036
<6>2008
<7>Neisseria gonorrhoeae is an obligate human pathogen that is the etiological agent of
gonorrhea. We explored variations in genes of a
multidrug-resistant N. gonorrhoeae Korean patient isolate in an effort to
understand the prevalence, antibiotic resistance, and importance of
horizontal gene transfer within this important, naturally competent
organism. Here we report the complete annotated genome sequence of N.
gonorrhoeae strain NCCP11945.

<>

<1>Chung, W.C., Chen, L.L., Lo, W.S., Kuo, P.A., Tu, J., Kuo, C.H.
<2>Complete Genome Sequence of Serratia marcescens WW4.
<3>Genome Announcements
<4>1
<5>e0012613
<6>2013
<7>Serratia marcescens WW4 is a biofilm-forming bacterium isolated from paper machine aggregates.
Under conditions of phosphate limitation, this bacterium
exhibits intergeneric inhibition of Pseudomonas aeruginosa. Here, the complete
genome sequence of S. marcescens WW4, which consists of one circular chromosome
(5,241,455 bp) and one plasmid (pSmWW4; 3,248 bp), was determined.

<>

<1>Cimerman, A., Arnaud, G., Foissac, X.
<2>Stolbur phytoplasma genome survey achieved using a suppression subtractive hybridization approach with high specificity.
<3>Appl. Environ. Microbiol.
<4>72
<5>3274-3283
<6>2006
<7>Phytoplasmas are unculturable bacterial plant pathogens transmitted by phloem-feeding
hemipteran insects. DNA of phytoplasmas is difficult to
purify because of their exclusive phloem location and low abundance in
plants. To overcome this constraint, suppression subtractive hybridization
(SSH) was modified and used to selectively amplify DNA of the stolbur
phytoplasma infecting a periwinkle plant. Plasmid libraries were
constructed, and the origins of the DNA inserts were verified by
hybridization and PCR screenings. After a single round of SSH, there was
still a significant level of contamination with plant DNA (around 50%).
However, the modified SSH, which included a second round of subtraction
(double SSH), resulted in an increased phytoplasma DNA purity (97%).
Results validated double SSH as an efficient way to produce a genome
survey for microbial agents unavailable in culture. Assembly of 266 insert
sequences revealed 181 phytoplasma genetic loci which were annotated.
Comparative analysis of 113 kbp indicated that among 217 protein coding
sequences, 83% were homologous to "Candidatus Phytoplasma asteris" (OY-M
strain) genes, with hits widely distributed along the chromosome. Most of
the stolbur-specific SSH sequences were orphan genes, with the exception
of two partial coding sequences encoding proteins homologous to a
mycoplasma surface protein and riboflavin kinase.

<>

<1>Cinelli, T., Moscetti, I., Matchi, G.
<2>PsasM2I, a Type II Restriction-Modification System in Pseudomonas savastanoi pv. savastanoi: Differential Distribution of Carrier Strains in the Environment and the Evolutionary History of Homologous RM Systems in the Pseudomonas syringae Complex.
<3>Microb. Ecol.
<4>68
<5>842-858
<6>2014
<7>A type II restriction-modification system was
found in a native plasmid of Pseudomonas savastanoi pv.
savastanoi MLLI2. Functional analysis of the methyltransferase
showed that the enzyme acts by protecting the DNA sequence
CTGCAG from cleavage. Restriction endonuclease
expression in recombinant Escherichia coli cells resulted in
mutations in the REase sequence or transposition of insertion
sequence 1A in the coding sequence, preventing lethal gene
expression. Population screening detected homologous RM
systems in other P. savastanoi strains and in the Pseudomonas
syringae complex. An epidemiological survey carried out by
sampling olive and oleander knots in two Italian regions
showed an uneven diffusion of carrier strains, whose presence
could be related to a selective advantage in maintaining the RM
system in particular environments or subpopulations. Moreover,
carrier strains can coexist in the same orchards, plants,
and knot tissues with non-carriers, revealing unexpected genetic
variability on a very small spatial scale. Phylogenetic analysis
of the RM system and housekeeping gene sequences in the
P. syringae complex demonstrated the ancient acquisition of the
RM systems. However, the evolutionary history of the gene
complex also showed the involvement of horizontal gene transfer
between related strains and recombination events.

<>

<1>Ciranna, A., Larjo, A., Kivisto, A., Santala, V., Roos, C., Karp, M.
<2>Draft Genome Sequence of the Hydrogen- and Ethanol-Producing Anaerobic Alkalithermophilic Bacterium Caloramator celer.
<3>Genome Announcements
<4>1
<5>e00471-13
<6>2013
<7>Caloramator celer strain JW/YL-NZ35 is a Gram-positive thermophilic, alkalitolerant, and
strictly anaerobic bacterium capable of producing hydrogen
and ethanol under extreme conditions. The draft genome sequence presented here
will provide valuable information to further explore the physiology of this
species and its potential for biofuel production.

<>

<1>Citron, M., Velleman, M., Schuster, H.
<2>Three additional operators, Op21, Op68, and Op88, of bacteriophage P1.
<3>J. Biol. Chem.
<4>264
<5>3611-3617
<6>1989
<7>The repressor of bacteriophage P1, encoded by the c1 gene, is responsible for maintaining the
P1 prophage in the lysogenic state.  Previously, 11 c1 repressor binding sites or operators
scattered over the whole genome of P1 have been found.  From sequence analysis an asymmetric,
17-base pair consensus sequence, ATTGCTCTAATAAATTT, was derived.  Using a synthetic
15-base-long oligodeoxyribonucleotide as operator probe, we have identified three additional
operators.  We have mapped the operators at the positions 21, 68, and 88 of the P1 genome and
determined their sequence.  These operators are controlled by c1 because corresponding P1 DNA
fragments (I) require c1 repressor in vivo in order to be clonable in multicopy plasmids, (ii)
exhibit a c1-repressible promoter activity, (iii) are retarded by c1 repressor protein during
electrophoresis, and (iv) contain the 17-base pair consensus sequence with one mismatch base
each.  Furthermore, we suggest that expression of the DNA adenine methylase (dam) encoded by
P1 is controlled via Op68.

<>

<1>Claesson, M.J., Li, Y., Leahy, S., Canchaya, C., van Pijkeren, J.P., Cerdeno-Tarraga, A.M., Parkhill, J., Flynn, S., O'sullivan, G.C., Collins, J.K., Higgins, D., Shanahan, F., Fitzgerald, G.F., van Sinderen, D., O'toole, P.W.
<2>Multireplicon genome architecture of Lactobacillus salivarius.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>6718-6723
<6>2006
<7>Lactobacillus salivarius subsp. salivarius strain UCC118 is a bacteriocin-producing strain
with probiotic characteristics. The 2.13-Mb
genome was shown by sequencing to comprise a 1.83 Mb chromosome, a 242-kb
megaplasmid (pMP118), and two smaller plasmids. Megaplasmids previously
have not been characterized in lactic acid bacteria or intestinal
lactobacilli. Annotation of the genome sequence indicated an intermediate
level of auxotrophy compared with other sequenced lactobacilli. No
single-copy essential genes were located on the megaplasmid. However,
contingency amino acid metabolism genes and carbohydrate utilization
genes, including two genes for completion of the pentose phosphate
pathway, were megaplasmid encoded. The megaplasmid also harbored genes for
the Abp118 bacteriocin, a bile salt hydrolase, a presumptive conjugation
locus, and other genes potentially relevant for probiotic properties. Two
subspecies of L. salivarius are recognized, salivarius and salicinius, and
we detected megaplasmids in both subspecies by pulsed-field gel
electrophoresis of sizes ranging from 100 kb to 380 kb. The discovery of
megaplasmids of widely varying size in L. salivarius suggests a possible
mechanism for genome expansion or contraction to adapt to different
environments.

<>

<1>Clancy, C.D., Forde, B.M., Moore, S.A., O'Toole, P.W.
<2>Draft Genome Sequences of Helicobacter pylori Strains 17874 and P79.
<3>J. Bacteriol.
<4>194
<5>2402
<6>2012
<7>Helicobacter pylori is a human pathogen that colonizes the human gastric mucosa,  causing
gastritis, duodenal and gastric ulcers, and gastric carcinoma. Here we
announce the draft genomes of H. pylori strain 17874, commonly used for studying
motility, and P79, a strain for which plasmid vectors have been developed.

<>

<1>Clanton, D.J., Riggsby, W.S., Miller, R.V.
<2>NgoII, a restriction endonuclease from Neisseria gonorrhoeae.
<3>J. Bacteriol.
<4>137
<5>1299-1307
<6>1979
<7>EndoR.NgoII, a class II restriction endonuclease isolated from Neisseria gonorrhoeae, was
purified to electrophoretic homogeneity.  We were able to separate it from another restriction
endonuclease of N. gonorrhoeae, NgoI, by phosphocellulose chromatography.  NgoII is an
isoschizomer of HaeIII, a restriction endonuclease of Haemophilus aegyptius, and was found to
recognize the deoxyribonucleic acid nucleotide base sequence GGCC.  NgoII was able to digest
phage lambda deoxyribonucleic acid over a wide pH range, with optimal activity at pH 8.5.  The
enzyme has an absolute requirement for Mg2+; maximal enzyme activity was observed at 1 mM
Mg2+.  The active enzyme has a molecular weight of 65,000 and appears to be composed of six
subunits of identical molecular weight (11,000).  No methylase activity could be detected in
the purified enzyme preparation.
[ The enzyme called NgoI in this abstract has been renamed NgoCI, Jan/1998. ]
[ The enzyme called NgoII in this abstract has been renamed NgoCII, Jan/1998. ]

<>

<1>Clanton, D.J., Woodward, J.M., Miller, R.V.
<2>Identification of a new sequence-specific endonuclease NgoII, from Neisseria gonorrhoeae.
<3>J. Bacteriol.
<4>135
<5>270-273
<6>1978
<7>A class II restriction endonuclease which recognizes the same nucleotide
sequence as EndoR-HaeIII has been found in four of seven isolates of Neisseria
gonorrhoeae.

<>

<1>Clark, C.G., Chong, P.M., McCorrister, S.J., Mabon, P., Walker, M., Westmacott, G.R.
<2>DNA Sequence Heterogeneity of Campylobacter jejuni CJIE4 Prophages and Expression of Prophage Genes.
<3>PLoS ONE
<4>9
<5>E95349
<6>2014
<7>Campylobacter jejuni carry temperate bacteriophages that can affect the biology
or virulence of the host bacterium. Known effects include genomic rearrangements
and resistance to DNA transformation. C. jejuni prophage CJIE1 shows sequence
variability and variability in the content of morons. Homologs of the CJIE1
prophage enhance both adherence and invasion to cells in culture and increase the
expression of a specific subset of bacterial genes. Other C. jejuni temperate
phages have so far not been well characterized. In this study we describe
investigations into the DNA sequence variability and protein expression in a
second prophage, CJIE4. CJIE4 sequences were obtained de novo from DNA sequencing
of five C. jejuni isolates, as well as from whole genome sequences submitted to
GenBank by other research groups. These CJIE4 DNA sequences were heterogenous,
with several different insertions/deletions (indels) in different parts of the
prophage genome. Two variants of a 3-4 kb region inserted within CJIE4 had
different gene content that distinguished two major conserved CJIE4 prophage
families. Additional indels were detected throughout the prophage. Detection of
proteins in the five isolates characterized in our laboratory in isobaric Tags
for Relative and Absolute Quantitation (iTRAQ) experiments indicated that
prophage proteins within each of the two large indel variants were expressed
during growth of the bacteria on Mueller Hinton agar plates. These proteins
included the extracellular DNase associated with resistance to DNA transformation
and prophage repressor proteins. Other proteins associated with known or
suspected roles in prophage biology were also expressed from CJIE4, including
capsid protein, the phage integrase, and MazF, a type II toxin-antitoxin system
protein. Together with the results previously obtained for the CJIE1 prophage
these results demonstrate that sequence variability and expression of moron genes
are both general properties of temperate bacteriophages in C. jejuni.

<>

<1>Clark, C.G., Ng, L.-K.
<2>Sequence variability of Campylobacter temperate bacteriophages.
<3>BMC Microbiol.
<4>8
<5>49
<6>2008
<7>Background: Prophages integrated within the chromosomes of Campylobacter jejuni isolates have
been demonstrated very recently. Prior work with Campylobacter temperate bacteriophages, as
well as evidence from prophages in other enteric bacteria, suggests these prophages might have
a
role in the biology and virulence of the organism. However, very little is known about the
genetic variability of Campylobacter prophages which, if present, could lead to differential
phenotypes in isolates carrying the phages versus those that do not. As a first step in the
characterization of C. jejuni prophages, we investigated the distribution of prophage DNA
within a C. jejuni population assessed the DNA and protein sequence variability within a
subset of the putative prophages found.
Results: Southern blotting of C. jejuni DNA using probes from genes within the three putative
prophages of the C. jejuni sequenced strain RM 1221 demonstrated the presence of at least one
prophage gene in a large proportion (27/35) of isolates tested. Of these, 15 were positive for
5 or
more of the 7 Campylobacter Mu-like phage 1 (CMLP 1, also designated Campylobacter jejuni
integrated element 1, or CJIE 1) genes tested. Twelve of these putative prophages were chosen
for further analysis. DNA sequencing of a 9,000 to 11,000 nucleotide region of each prophage
demonstrated a close homology with CMLP 1 in both gene order and nucleotide sequence.
Structural and sequence variability, including short insertions, deletions, and allele
replacements, were found within the prophage genomes, some of which would alter the protein
products of the ORFs involved. No insertions of novel genes were detected within the sequenced
regions. The 12
prophages and RM 1221 had a % G+C very similar to C. jejuni sequenced strains, as well as
promoter regions characteristic of C. jejuni. None of the putative prophages were successfully
induced and propagated, so it is not known if they were functional or if they represented
remnant
prophage DNA in the bacterial chromosomes.
Conclusion: These putative prophages form a family of phages with conserved sequences, and
appear to be adapted to Campylobacter. There was evidence for recombination among groups of
prophages, suggesting that the prophages had a mosaic structure. In many of these properties,
the Mu-like CMLP 1 homologs characterized in this study resemble temperate bacteriophages of
enteric bacteria that are responsible for contributions to virulence and host adaptation.

<>

<1>Clark, J., Harrison, J.C., Mdegela, R.H., March, J.B.
<2>Extended stability of restriction enzymes at ambient temperatures.
<3>Biotechniques
<4>29
<5>536
<6>2000
<7>The stability of restriction enzymes as supplied by manufacturers without any modification has
been examined. No reduction in activity
was observed for three enzymes (HindIII, EcoRI and Tsp5091) held at
ambient temperature or 4 degrees C for the period of study (12 months),
while activity was observed for up to 12 weeks after storage at 37
degrees C, which was considerably better than following desiccation
with trehalose, a recognized preservation technique. A larger trial of
23 different restriction enzymes held at room temperature for one week
showed that all enzymes retained significant activity. As a practical
demonstration of the usefulness of this finding, enzymes were posted to
Africa by conventional mail (cost $1 US) and shown to retain activity
upon arrival after three weeks in transit (compared to a cost of $1000
US by cold-chain transportation). Supplying enzymes to third-world
markets should now be possible by removing the necessity for cold-chain
transport. After arrival, enzymes can simply be stored in a standard
domestic refrigerator.

<>

<1>Clark, J., Shevchuk, T., Kho, M.R., Smith, S.S.
<2>Methods for the design and analysis of oligodeoxynucleotide-based DNA (cytosine-5) methyltransferase inhibitors.
<3>Anal. Biochem.
<4>321
<5>50-64
<6>2003
<7>Several second-generation inhibitors of DNA (cytosine-5) methyltransferases based on studies
of modified synthetic
oligodeoxynucleoides have been described. As an aid to studies of these
inhibitors, we present an electronic structure-based algorithm that can be
used as a method for predicting the nature of the expected inhibition by
any noncytosine nucleotide target. Targeting by the major human enzyme
(hDnmt1) is governed by the presence of a three-nucleotide motif. In
hemimethylated DNA, this motif consists of a 5-methylcytosine targeting
signal that causes the enzyme to probe the opposite strand for a normally
paired guanosine or inosine residue and attempt to methylate the residue
5' to that site. As a demonstration of the method, we apply these rules to
the design and characterization of a novel oligodeoxynucleotide inhibitor
of hDnmt1. This inhibitor takes advantage of the three-nucleotide
recognition motif characteristic of hDnmt1 and shows that the enzyme is
inhibited in vitro by non-CG methylation which targets the enzyme to
normally basepaired but unproductive nucleotides such as dG, dA, and dT.
Kinetic analysis at constant S-adenosyl-L-methionine concentration shows
that representative inhibitory oligodeoxynucleotides are best viewed as
weakly productive components of systems containing two DNA substrates.
This model suggests that the most effective inhibitors are those with very
low apparent Vmax and very low Km values. Oligodeoxynucleotides containing
mispaired and unproductive targets such as dG, dA, dT, and dU are also
inhibitory as secondary substrates for the human enzyme. Biologically,
fail-safe mechanisms identified by the ab initio approach appear to be
active in preventing potentially mutagenic deamination of dihydrocytosine
and enzymatic methylation of dU.

<>

<1>Clark, N.C., Zhu, W., Patel, J.B.
<2>Missense Mutation of hsdR-Encoded Type I Restriction Enzyme Implicated in Transfer of Vancomycin-Resistance from Enterococcus to Staphylococcus aureus.
<3>Abstr. InterSci. Conf. Antimicrob. Agents Chemother.
<4>47
<5>83-84
<6>2007
<7>Background:  Seven vanA-mediated vancomycin-resistant S. aureus (VRSA) cases have been
reported. To prevent VRSA, it is important to understand the microbiological characteristics
that allow for transfer of vanA from vancomycin-resistant Enterococcus (VRE) to S. aureus.
Previously it was reported that S. aureus strain RN4220 readily acquires foreign DNA as a
result of a missense mutation within the hsdR gene of the Type I restriction-modification
system. Therefore we investigated the possibility that this mutation may have contributed to
the transfer of vanA from Enterococcus to S. aureus.
Methods:  The hsdR gene was amplified using primers, previously described by Waldron and
Lindsay (2006. J. Bacteriol.), and both strands of this PCR product were sequenced using
eleven additional primers.  The resulting sequences were compared to those of S. aureus
8325-4, the parent strain of RN4220, and the S. aureus COL strain.
Results:  The hsdR sequences of a VRSA isolate from each case were analyzed. For VRSA 1, 2, 5,
and 6 no mutation was identified that changes the amino acid sequence. For VRSA 4, a base
substitution was identified that resulted in a I to F amino acid change at position 774 and
for VRSA 7, a base substitution resulted in a T to P amino acid change at position 362. For
VRSA 3 a single-base deletion C, at base position 1382 (codon 461) created a frameshift and a
premature stop codon at base position 1405. These changes would result in a truncation of the
restriction enzyme from 930 amino acids to 469 amino acids. This truncation (50% of the
protein) is less extensive than that reported for S. aureus RN4220 (21%).  Eight other S.
aureus isolates (1 VRSA and 7 vancomycin-susceptible SA) from the same patient were
characterized. These isolates were recovered from multiple body sites on different days, but
were closely related by pulsed-field gel electrophoresis typing.  All 9 S. aureus isolates had
the same hsdR gene mutation.
Conclusions:  Three of the seven VRSA isolates has a mutation in the hsdR gene that resulted
in a protein change. The amino acid substitutions identified in VRSA 4 and 7 have not been
reported in another S. aureus and the significance of these changes is unknown. The missense
mutation identified in VRSA 3 and related S. aureus isolates from the same patient likely
results in a loss of function of the Type I restriction enzyme and may allow this strain to
more readily acquire foreign DNA than strains with a functional restriction enzyme.

<>

<1>Clark, S.J., Harrison, J., Paul, C.L., Frommer, M.
<2>High sensitivity mapping of methylated cytosines.
<3>Nucleic Acids Res.
<4>22
<5>2990-2997
<6>1994
<7>An understanding of DNA methylation and its potential role in gene control during development,
aging and cancer has been hampered by a lack of sensitive methods which can resolve exact
methylation patterns from only small quantities of DNA. We have now developed a genomic
sequencing technique which is capable of detecting every methylated cytosine on both strands
of any target sequence, using DNA isolated from fewer than 100 cells. In this method, sodium
bisulphite is used to convert cytosine residues to uracil residues in single-stranded DNA,
under conditions whereby 5-methylcytosine remains non-reactive. The converted DNA is amplified
with specific primers and sequenced. All the cytosine residues remaining in the sequence
represent previously methylated cytosines in the genome. The work described has defined
procedures that maximise the efficiency of denaturation, bisulphite conversion and
amplification, to permit methylation mapping of single genes from small amounts of genomic
DNA, readily available from germ cells and early developmental stages.

<>

<1>Clark, T.A., Lu, X., Luong, K., Dai, Q., Boitano, M., Turner, S.W., He, C., Korlach, J.
<2>Enhanced 5-methylcytosine detection in single-molecule, real-time sequencing via  Tet1 oxidation.
<3>BMC Biol.
<4>11
<5>4
<6>2013
<7>ABSTRACT: BACKGROUND: DNA methylation serves as an important epigenetic mark in both
eukaryotic and prokaryotic organisms. In eukaryotes, the most common
epigenetic mark is 5-methylcytosine, whereas prokaryotes can have
6-methyladenine, 4-methylcytosine, or 5-methylcytosine. Single-molecule,
real-time sequencing is capable of directly detecting all three types of modified
bases. However, the kinetic signature of 5-methylcytosine is subtle, which
presents a challenge for detection. We investigated whether conversion of
5-methylcytosine to 5-carboxylcytosine using the enzyme Tet1 would enhance the
kinetic signature, thereby improving detection. RESULTS: We characterized the
kinetic signatures of various cytosine modifications, demonstrating that
5-carboxylcytosine has a larger impact on the local polymerase rate than
5-methylcytosine. Using Tet1-mediated conversion, we show improved detection of
5-methylcytosine using in vitro methylated templates and apply the method to the
characterization of 5-methylcytosine sites in the genomes of Escherichia coli
MG1655 and Bacillus halodurans C-125. CONCLUSIONS: We have developed a method for
the enhancement of directly detecting 5-methylcytosine during single-molecule,
real-time sequencing. Using Tet1 to convert 5-methylcytosine to
5-carboxylcytosine improves the detection rate of this important epigenetic
marker, thereby complementing the set of readily detectable microbial base
modifications, and enhancing the ability to interrogate eukaryotic epigenetic
markers.

<>

<1>Clark, T.A., Murray, I.A., Morgan, R.D., Kislyuk, A.O., Spittle, K.E., Boitano, M., Fomenkov, A., Roberts, R.J., Korlach, J.
<2>Characterization of DNA methyltransferase specificities using single-molecule, real-time DNA sequencing.
<3>Nucleic Acids Res.
<4>40
<5>e29
<6>2012
<7>DNA methylation is the most common form of DNA modification in prokaryotic and eukaryotic
genomes. We have applied the method of single-molecule, real-time (SMRT(R)) DNA sequencing
that is capable of direct detection of modified bases at single-nucleotide resolution to
characterize the specificity of several bacterial DNA methyltransferases (MTases). In addition
to previously described SMRT sequencing of N6-methyladenine and 5-methylcytosine, we show that
N4-methylcytosine also has a specific kinetic signature and is therefore identifiable using
this approach. We demonstrate for all three prokaryotic methylation types that SMRT sequencing
confirms the identity and position of the methylated base in cases where the MTase specificity
was previously established by other methods. We then applied the method to determine the
sequence context and methylated base identity for three MTases with unknown specificities. In
addition, we also find evidence of unanticipated MTase promiscuity with some enzymes
apparently also modifying sequences that are related, but not identical, to the cognate site.

<>

<1>Clarke, C.M., Hartley, B.S.
<2>Purification, properties and specificity of the restriction endonuclease from Bacillus stearothermophilus.
<3>Biochem. J.
<4>177
<5>49-62
<6>1979
<7>The restriction endonuclease BstI was purified from 70kg of Bacillus stearothermophilus.  The
final product is at least 97% pure as judged by sodium dodecyl sulphate/polyacrylamide-gel
electrophoresis; this major protein species co-migrates with the enzyme activity on native
polyacrylamide-gel electrophoresis and isoelectric focusing.  Pure restriction endonuclease
BstI has a subunit mol. wt. of 26000 and is probably a loosely associated dimer. The enzyme
shows maximum activity at pH values between 7 and 9.5, and in the presence of 0.5-2mM-Mg2+.
NaCl inhibits the restriction enzymeactivity. Restriction endonuclease BstI cleaves DNA in a
position identical with that cleaved by endonuclease BamHI (for Bacillus amyloliquefaciens),
i.e.:
5'-G/-G-A-T-C-C-3'
3'-C-C-T-A-G/-G-5'.
In the presence of high concentrations of enzyme, DNA cleavage occurs at secondary sites.
This side-specificity is enhanced by the addition of glycerol.  Preliminary studies indicate
that these sites are of the type:
5'-/G-A-T-C-3'
3'-C-T-A-G/-5'.

<>

<1>Clarke, D.J., Chaudhuri, R.R., Martin, H.M., Campbell, B.J., Rhodes, J.M., Constantinidou, C., Pallen, M.J., Loman, N.J., Cunningham, A.F., Browning, D.F., Henderson, I.R.
<2>Complete genome sequence of the Crohn's disease-associated adherent-invasive Escherichia coli strain HM605.
<3>J. Bacteriol.
<4>193
<5>4540
<6>2011
<7>Adherent-invasive Escherchia coli strains are increasingly being associated with intestinal
pathologies. Here we present the genome
sequence of E. coli HM605, a strain isolated from colonic biopsies of a
patient with Crohn's disease.

<>

<1>Clarke, N.D.
<2>A proposed mechanism for the self-splicing of proteins.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>11084-11088
<6>1994
<7>Intervening protein sequences, called inteins, are intronlike elements that
are removed posttranslationally, apparently by self-splicing.  The conserved and essential
residues of precursor proteins consist of an asparagine as the last residue of the intein and
a
hydroxyl- or thiol-containing residue immediately following both splice junctions.
Evidence for a branched intermediate has been reported; however, the chemical nature of
the branched structure is unclear.  I propose a mechanism that includes the formation of a
branched structure, provides an explanation for the reversal of branch formation observed
at high pH, and accounts for each of the essential amino acids.  The branched structure is
formed by nucleophilic attack of the asparagine side chain on the N-terminal splice
junction.  The nature of this branched structure is a distinguishing feature of the model and
can be experimentally tested.

<>

<1>Clarke, S., Banfield, K.
<2>S-Adenosylmethionine-dependent methyltransferases.
<3>Homocysteine in Health and Disease, Cambridge University Press, Carmel, R. and Jacobsen, D.W., 
<4>
<5>63-78
<6>2001
<7>Mammalian S-adenosylmethionine-dependent methyltransferases each catalyze a reaction, giving
rise to two products - S-adenosylhomocysteine and one of a variety of methylated biomolecules
including nucleic acids, proteins, lipids, and small molecules.  S-adenosylhomocysteine is
subsequently broken down to adenosine and homocysteine by S-adenosylhomocysteine hydrolase.
The homocysteine formed can be either remethylated to methionine or converted to cysteine via
cystathionine.  As such, these methyltransferases are bifunctional; they make up an essential
part of the conduit for the conversion of methionine to cysteine in addition to generating
methylated products.

<>

<1>Claus, H., Friedrich, A., Frosch, M., Vogel, U.
<2>Differential distribution of novel restriction-modification systems in clonal lineages of Neisseria meningitidis.
<3>J. Bacteriol.
<4>182
<5>1296-1303
<6>2000
<7>Using representational difference analysis, we isolated novel meningococcal
restriction-modification (R-M) systems. NmeBI, which is a homologue of the R-M system HgaI of
Pasteurella volantium, was present in meningococci of the ET-5 complex and of lineage III.
NmeAI was found in serogroup A, ET-37 complex, and cluster A4 meningococci. NmeDI was harbored
by meningococci of the ET-37 complex and of cluster A4, but not by serogroup A meningococci.
Two of the R-M systems, NmeBI and NmeDI, were located at homologous positions between the
phenylalanyl-tRNA synthetase genes pheS and pheT, which appeared to be a preferential target
for the insertion of foreign DNA in meningococci. The distribution of the three R-M systems
was tested with 103 meningococcal strains comprising 49 sequence types. The vast majority of
the strains had either NmeBI, NmeAI, or both NmeAI and NmeDI. Using cocultivation experiments,
we could demonstrate that NmeBI, which was present in ET-5 complex meningococci, was
responsible for a partial restriction of DNA transfer from meningococci of the ET-37 complex
to meningococci of the ET-5 complex.

<>

<1>Claus, H., Stoevesandt, J., Frosch, M., Vogel, U.
<2>Genetic isolation of meningococci of the electrophoretic type 37 complex.
<3>J. Bacteriol.
<4>183
<5>2570-2575
<6>2001
<7>Neisseria meningitidis (the meningococcus) is a naturally competent bacterial species in which
intra- and interspecific horizontal gene transfer is a major source of genetic diversity. In
strains of the electrophoretic type 37 (ET-37) complex and of the A4 cluster, we identified
genomic DNA coding for a novel restriction-modification system and for the tail of a
previously unidentified prophage. Furthermore, a novel 7.2-kb DNA segment restricted to clones
of the ET-37 complex and the A4 cluster was isolated and shown to occur both as a plasmid
(pJS-B) and as a chromosomal integration. Neither the genomic loci nor pJS-B was present in
ET-5 complex, lineage 3, or serogroup A meningococci. The differential distribution of the DNA
segments described herein, as well as of opcA, porB, nmeAI, nmeBI, and nmeDI described
previously, supports the concept of genetic isolation of hypervirulent lineages responsible
for most cases of serogroup C disease worldwide.

<>

<1>Cleaver, J.E., Samson, L., Thomas, G.H.
<2>Restriction enzyme cleavage of ultraviolet-damaged DNA.
<3>Biochim. Biophys. Acta
<4>697
<5>255-258
<6>1982
<7>SV40 and pBR322 DNAs damaged by ultraviolet light were cleaved abnormally by
several restriction enzymes because of damage to pyrimidines in the recognition
sequences.  The use of a tandemly duplicated plasmid provided a particularly
sensitive target molecule for detecting pyrimidine dimers and other possible
photoproducts.  The relative efficiency with which cleavage was blocked
(HindIII>TaqI>BamI>SalI>>HhaI, HaeIII) corresponds approximately to the
relative frequency of pyrimidine dimer formation in the recognition sequences,
but at a slightly higher frequency in potential sites for the non-cyclobutane
T-C product.  The pyrimidine dimers appear to have a range of influence that
extends 1 to 3 basepairs along the DNA molecule.  These effects provide clues
to the way DNA damage from mutagens and carcinogens can interfere with specific
enzyme-DNA interactions.

<>

<1>Clements, J.B., Cortini, R., Wilkie, N.M.
<2>Analysis of Herpesvirus DNA Substructure by means of Restriction Endonucleases.
<3>J. Gen. Virol.
<4>30
<5>243-256
<6>1976
<7>The mol. wt. and molar ratios of the HindIII and HpaI fragments of HSV-1 DNA
and the EcoRI fragments of HSV-2 DNA have been determined.  Results obtained
suggest that DNA isolated from both HSV-1 and HSV-2 consists of molecules with
four different sequence arrangements which are present in similar amounts.  Our
explanation of the cleavage patterns of these four genome arrangements with the
different restriction enzymes is presented.  Some of the possible implications
of these four genome arrangements for genetic recombination are discussed.

<>

<1>Clermont, O., Denamur, E., le Gall, T., Tenaillon, O.
<2>Human specific escherichia coli strains.
<3>International Patent Office
<4>WO 2009021977 A
<5>
<6>2009
<7>The present invention relates to human specific Escherichia coli strains.

<>

<1>Cleto, S., Van der Auwera, G., Almeida, C., Vieira, M.J., Vlamakis, H., Kolter, R.
<2>Genome Sequence of Serratia plymuthica V4.
<3>Genome Announcements
<4>2
<5>e00340-14
<6>2014
<7>Serratia spp. are gammaproteobacteria and members of the family Enterobacteriaceae. Here, we
announce the genome sequence of Serratia plymuthica
strain V4, which produces the siderophore serratiochelin and antimicrobial
compounds.

<>

<1>Clifford, R.J., Hang, J., Riley, M.C., Onmus-Leone, F., Kuschner, R.A., Lesho, E.P., Waterman, P.E.
<2>Complete Genome Sequence of Providencia stuartii Clinical Isolate MRSN 2154.
<3>J. Bacteriol.
<4>194
<5>3736-3737
<6>2012
<7>Here we present the complete genome sequence of Providencia stuartii MRSN 2154, isolated from
an Afghan national. P. stuartii is a Gram-negative bacillus capable
of causing infections in a wide variety of human tissues. Because Providencia
readily acquires plasmids bearing drug resistance loci, it is of growing clinical
significance.

<>

<1>Clum, A. et al.
<2>Complete genome sequence of Pirellula staleyi type strain (ATCC 27377).
<3>Standards in Genomic Sciences
<4>1
<5>308-316
<6>2009
<7>Pirellula staleyi Schlesner and Hirsch 1987 is the type species of the genus Pirellula of the
family Planctomycetaceae. Members of this pear- or
teardrop-shaped bacterium show a clearly visible pointed attachment pole and can
be distinguished from other Planctomycetes by a lack of true stalks. Strains
closely related to the species have been isolated from fresh and brackish water,
as well as from hypersaline lakes. Here we describe the features of this
organism, together with the complete genome sequence and annotation. This is the
first completed genome sequence of the order Planctomyces and only the second
sequence from the phylum Planctobacteria/Planctomycetes. The 6,196,199 bp long
genome with its 4773 protein-coding and 49 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Clum, A. et al.
<2>Complete genome sequence of Acidimicrobium ferrooxidans type strain (ICP).
<3>Standards in Genomic Sciences
<4>1
<5>38-45
<6>2009
<7>Acidimicrobium ferrooxidans (Clark and Norris 1996) is the sole and type species  of the
genus, which until recently was the only genus within the actinobacterial
family Acidimicrobiaceae and in the order Acidomicrobiales. Rapid oxidation of
iron pyrite during autotrophic growth in the absence of an enhanced CO(2)
concentration is characteristic for A. ferrooxidans. Here we describe the
features of this organism, together with the complete genome sequence, and
annotation. This is the first complete genome sequence of the order
Acidomicrobiales, and the 2,158,157 bp long single replicon genome with its 2038
protein coding and 54 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Clyman, J.
<2>Some microbes have splicing proteins.
<3>ASM News
<4>61
<5>344-347
<6>1995
<7>The recent discovery of microbial genes whose products are spliced at the protein level,
instead of at the mRNA level, adds an unexpected dimension of complexity to the ways in which
genetic information flows from DNA to protein. These amazing protein elements, discovered
about 5 years ago and now called inteins, typically catalyze both protein and DNA
rearrangements.

<>

<1>Clyman, J., Belfort, M.
<2>Trans and cis requirements for intron mobility in a prokaryotic system.
<3>Genes Dev.
<4>6
<5>1269-1279
<6>1992
<7>Intron mobility requires cleavage of an intronless allele by an intron-encoded endonuclease,
followed by transfer of the intron into the cleaved recipient.  The mobile phage introns
provide an opportunity to identify accessory functions involved in the intron inheritance
process.  To test for trans and cis requirements of mobility in Escherichia coli, we have
exploited the td intron of phage T4 in both phage T4 and lambda genetic backgrounds.  Mobility
depends on host or phage recombinase functions, RecA or UvsX, respectively.  The process also
requires a phage-encoded 5'-->3' exonuclease activity and associated annealing function that
can be provided by phage lambda.  Finally, host-encoded 3'-->5' exonuclease activities are
also implicated in intron inheritance.  We demonstrated further that restriction enzymes could
substitute for the intron-encoded endonuclease, indicating that the endonuclease does not have
an essential role in recombination.  Neither the precise position nor the nature of the
double-strand break was critical to intron transfer.  These features provide insight into the
recombination pathway and are factors impacting on the spread of introns throughout natural
populations.

<>

<1>Coad, J.E., Lander, T.A., Litz, C.E.
<2>Inhibition of restriction endonucleases by common clinical anticoagulants.
<3>Anal. Biochem.
<4>205
<5>368-369
<6>1992
<7>Anticoagulated peripheral blood and bone marrow provide an accessible source of DNA for
molecular studies. Successful results in such investigations often depend on full activity of
labile restriction enzymes. Though occasional studies have shown heparin to be inhibitory, an
often overlooked cause of enzyme inactivity is the effect of the anticoagulant used in sample
collection. No systematic study regarding the effect of anticoagulants on restriction enzymes
or techniques to remove anticoagulant contamination has been reported. The sensitivites of 18
commonly used restriction endonucleases to sodium heparin, citric acid-sodium citrate-dextrose
(ACD), and ethylenediaminetetraacetic acid (EDTA) and various methods of decontamination are
presented.

<>

<1>Cocks, B.G., Finch, L.R.
<2>Characterization of a restriction endonuclease from Ureaplasma urealyticum 960 and differences in deoxyribonucleic acid modification of human ureaplasmas.
<3>Int. J. Syst. Bacteriol.
<4>37
<5>451-453
<6>1987
<7>Uur960I, a restriction endonuclease from Ureaplasma urealyticum 960T, cleaved
at the sequence 5'-GC/NGC-3' and is thus an isoschizomer of Fnu4HI.  Fnu4HI
cleaved deoxyribonucleic acid from human ureaplasma serovars I,III, and VI but
not II, IV, V, VII, VIII (strain 960), and IX.  This grouping of serovars,
indicative of their deoxyribonucleic acid modification, matches that previously
reported by others using different criteria.

<>

<1>Coene, M., Hoet, P., Cocito, C.
<2>Physical map of Phage 2 C DNA: Evidence for the existence of large redundant ends.
<3>Eur. J. Biochem.
<4>132
<5>69-75
<6>1983
<7>The chromosome of the Bacillus subtilis phage 2C, a linear molecule of double-stranded DNA of
about 10^8 Da, in which thymine is completely replaced by hydroxymethyluracil, was cleaved by
different endonucleases. In some cases restriction segments were much fewer than expected,
suggesting a possible interference of the unusual base with the recognition mechanism of
endonucleases. The physical map of 2C DNA was established by use of SalI and HaeIII
restriction endonucleases, which yielded a limited number of fragments. The expected number of
fragments was 240 for HaeIII and 23 for SalI; in reality, five segments were observed upon
cleavage with HaeIII and four with SalI. The terminal fragments of the genome were first
identified; the other fragments were ordered by hybridization and molecular weight
determination of restriction fragments obtained by cleavage with the two endonucleases. In
addition, hybridization of restriction fragments showed the presence of homologous regions at
the ends of the 2C genome. The structure of these direct repetitive sequences was analyzed by
cleavage with HaeIII and hybridization with EcoRI restriction fragments. Their size (9.2 MDa)
was found to be about 1/11 of that of the whole chromosome.

<>

<1>Coetzee, J.N., Smit, J.A.
<2>Restriction without modification of phage 34/13 in a strain of Proteus mirabilis.
<3>S. Afr. Med. J.
<4>43
<5>356
<6>1969
<7>Phage 34/13 prepared on its host strain Proteus mirabilis 13at does not form
plaques on P. mirabilis strain N6 when serial dilutions are spotted on a lawn
of the organism.  Low dilutions show areas of clearing due to bacterial lysis
but no phage capable of forming plaques on N6 is obtained from these zones.
Phage 34/13 adsorbs to an extent of 99% on strain N6 within 15 min.  With the
use of 32P-labelled phage 34/13 DNA it is shown that within 15 min. of
adsorption to N6, 57% of the label is in the medium in the form of acid-soluble
nucleotides.  This figure is reduced to 11% when the phage adsorbs to its
normal host 13at and 0.9% when the phage is mixed with a strain of P. mirabilis
to which it does not adsorb.  This indicates that the DNA of phage 34/13 is
restricted by strain N6.  The phage DNA which escapes restriction is not
modified as no plaques are formed.  Phenotypically strain N6 behaves as if its
genotype is r+m-.  It may be argued that r+m- strains should degrade their own
DNA and be nonviable.  Strains of this genotype have never been isolated after
mutagen treatment of wild strains.  Many bacteria which restrict foreign DNA
owe this property to a prophage.  Strain N6 carries a prophage which on UV
induction produces a defective phage which manifests itself as a bacteriocin
which kills strains of P. mirabilis, and this plasmid may contribute to the
restrictive process.  This has not been proved.  Attempts to isolate mutants of
phage 34/13 which escape restriction have been unsuccessful.  Fruitless
attempts have also been made to isolate r-m- or r-m+ mutants of strain N6 which
would allow plaque formation by the phage.

<>

<1>Coffey, A., Ross, R.P.
<2>Bacteriophage-resistance systems in dairy starter strains: molecular analysis to application.
<3>Antonie Van Leeuwenhoek
<4>82
<5>303-321
<6>2002
<7>Starter inhibition by bacteriophage infection in dairy fermentations can limit the usage of
specific bacterial strains used in the
manufacture of Cheddar, Mozzarella and other cheeses and can result in
substantial economic losses. A variety of practical measures to
alleviate the problem of phage infection have been adopted over the
years but has invariably resulted in a very limited number of strains
which can withstand intensive usage in industry. The application of
genetic techniques to improve the phage-resistance of starter cultures
for dairy fermentations has been intensively studied for the last 20
years to a point where this approach now has significant potential to
alleviate the problem. This paper highlights the recent findings and
developments that have been described in the literature that will have
an impact on improvement of the phage-resistance of starter cultures.

<>

<1>Coffey, A., Stokes, D., Fitzgerald, G.F., Ross, R.P.
<2>Traditional and molecular approaches to improving bacteriophage resistance of Cheddar and Mozzarella cheese starters.
<3>Irish J. Agr. Food Res.
<4>40
<5>239-270
<6>2001
<7>Infection by bacteriophage (bacterial viruses) during dairy fermentations remains a major
cause of starter culture failure in
Cheddar and Mozzarella manufacture, often resulting in substantial
economic losses. A variety of practical measures to alleviate the
problem of phage infection have been adopted over the years. The
application of genetic techniques to improve the phage resistance of
starter cultures for dairy fermentations is currently being explored
and this approach has significant potential to alleviate the problem.
This review highlights the significant developments that have been made
in understanding the interaction between dairy starter cultures and
bacteriophage in industry. It also describes the exploitation of
molecular methodology to genetically defend these bacteria from the
ever-present threat of bacteriophage and the scientific advances, which
formed the basis for these achievements. Attention is also given to the
development of food-grade approaches to improve genetic traits of
industrial starter strains in the context of phage resistance.

<>

<1>Coffey, B., Ross, R.P., O'Flynn, G., O'Sullivan, O., Casey, A., Callanan, M., Coffey, A., McAuliffe, O.
<2>Complete Genome Sequence of vB_EcoM_112, a T-Even-Type Bacteriophage Specific for Escherichia coli O157:H7.
<3>Genome Announcements
<4>2
<5>e00393-14
<6>2014
<7>Bacteriophage vB_EcoM_112 (formerly e11/2) is an Escherichia coli phage with specificity for
the O157:H7 serotype. The vB_EcoM_112 genome sequence shares high
degrees of similarity with the phage T4 genome sequence.

<>

<1>Coffin, S.R., Reich, N.O.
<2>Escherichia coli DNA Adenine Methyltransferase THE STRUCTURAL BASIS OF PROCESSIVE CATALYSIS AND INDIRECT READ-OUT.
<3>J. Biol. Chem.
<4>284
<5>18390-18400
<6>2009
<7>We have investigated the structural basis of processive GATC methylation by the Escherichia
coli DNA adenine methyltransferase,
which is critical in chromosome replication and mismatch repair. We
determined the contribution of the orthologically conserved phosphate
interactions involving residues Arg(95), Asn(126), Asn(132), Arg(116),
and Lys(139), which directly contact the DNA outside the cognate
recognition site (GATC) to processive catalysis, and that of residue
Arg(137), which is not conserved and contacts the DNA backbone within
the GATC sequence. Alanine substitutions at the conserved positions
have large impacts on processivity yet do not impact k(cat)/K-m(DNA) or
DNA affinity (K-D(DNA)). However, these mutants cause large preferences
for GATC sites varying in flanking sequences when considering the
pre-steady state efficiency constant k(chem)/K-D(DNA). These changes
occur mainly at the level of the methylation rate constant, which
results in the observed decreases in processive catalysis. Thus,
processivity and catalytic efficiency (k(cat)/K-m(DNA)) are uncoupled
in these mutants. These results reveal that the binding energy involved
in DNA recognition contributes to the assembly of the active site
rather than tight binding. Furthermore, the conserved residues
(Arg(95), Asn(126), Asn(132), and Arg(116)) repress the modulation of
the response of the enzyme to flanking sequence effects. Processivity
impacted mutants do not show substrate-induced dimerization as is
observed for the wild type enzyme. This study describes the structural
means by which an enzyme that does not completely enclose its substrate
has evolved to achieve processive catalysis, and how interactions with
DNA flanking the recognition site alter this processivity.

<>

<1>Coffin, S.R., Reich, N.O.
<2>Escherichia coli DNA Adenine Methyltransferase: Intrasite Processivity and Substrate-Induced Dimerization and Activation.
<3>Biochemistry
<4>48
<5>7399-7410
<6>2009
<7>Methylation of GATC sites in Escherichia coli by DNA adenine methyltransferase (EcoDam) is
essential for proper DNA replication
timing, gene regulation, and mismatch repair. The low cellular
concentration of EcoDam and the high number of GATC sites in the genome
(similar to 20000) Support the reliance on methylation
efficiency-enhancing strategies such as extensive intersite
processivity. Here, we present evidence that EcoDam has evolved other
unique mechanisms of activation not commonly observed with
restriction-modification methyltransferases, EcoDam dimerizes oil
short, synthetic DNA, resulting in enhanced catalysis; however,
dimerization is not observed on large genomic DNA where the potential
for intersite processive methylation precludes any
dimerization-dependent activation. An activated form of the enzyme is
apparent on large genomic DNA and can also be achieved with high
concentrations of short, synthetic substrates. We suggest that this
activation is inherent on polymeric DNA where either multiple GATC
sites are available for methylation or the partitioning of the enzyme
onto nonspecific DNA is favored. Unlike other restriction-modification
methyltransferases, EcoDam carries out intrasite processive catalysis
whereby the enzyme-DNA complex methylates both strands of an
unmethylated GATC site prior to dissociation From the DNA. This occurs
with short 21 bp oligonucleotides and is highly dependent upon salt
concentrations. Kinetic modeling which invokes enzyme activation by
both dimerization and excess substrate provides mechanistic insights
into key regulatory checkpoints for an enzyme involved in multiple,
diverse biological pathways.

<>

<1>Coffin, S.R., Reich, N.O.
<2>Modulation of Escherichia coli DNA methyltransferase activity by biologically derived GATC-flanking sequences.
<3>J. Biol. Chem.
<4>283
<5>20106-20116
<6>2008
<7>Escherichia coli DNA adenine methyltransferase (EcoDam) methylates the N-6 position of the
adenine in the sequence 5'-GATC-3' and plays vital
roles in gene regulation, mismatch repair, and DNA replication. It
remains unclear how the small number of critical GATC sites involved in
the regulation of replication and gene expression are differentially
methylated, whereas the similar to 20,000 GATCs important for mismatch
repair and dispersed throughout the genome are extensively methylated.
Our prior work, limited to the pap regulon, showed that methylation
efficiency is controlled by sequences immediately flanking the GATC
sites. We extend these studies to include GATC sites involved in
diverse gene regulatory and DNA replication pathways as well as sites
previously shown to undergo differential in vivo methylation but whose
function remains to be assigned. EcoDam shows no change in affinity
with variations in flanking sequences derived from these sources, but
methylation kinetics varied 12-fold. A-tracts immediately adjacent to
the GATC site contribute significantly to these differences in
methylation kinetics. Interestingly, only when the poly(A) is located
5' of the GATC are the changes in methylation kinetics revealed.
Preferential methylation is obscured when two GATC sites are positioned
on the same DNA molecule, unless both sites are surrounded by large
amounts of nonspecific DNA. Thus, facilitated diffusion and sequences
immediately flanking target sites contribute to higher order
specificity for EcoDam; we suggest that the diverse biological roles of
the enzyme are in part regulated by these two factors, which may be
important for other enzymes that sequence-specifically modify DNA.

<>

<1>Coffman, G.L., Gaubatz, J.W., Yielding, K.L., Yielding, L.W.
<2>Demonstration of specific high affinity binding sites in plasmid DNA by photoaffinity labeling with an ethidium analog.
<3>J. Biol. Chem.
<4>257
<5>13205-13207
<6>1982
<7>We have used photoaffinity labeling of pBR322 DNA with
8-azido-3-amino-5ethyl-6-phenylphenanthridinium chloride to demonstrate high affinity ethidium
binding sites. Plasmid equilibrated with as little as 1 drug/DNA molecule was photoactivated,
freed of uncomplexed drug by ethanol precipitation, and subjected to restriction analysis.
There was highly specific, rather than random blockage of HhaI sites (d(GCGC))at low drug
concentrations. Furthermore, the same 7 new digestion fragments were generated at drug to
nucleotide ratios ranging from 1:100 to 1:8000. All the new DNA fragments had chain lengths
greater than the largest HhaI fragment (393 base pairs). At higher ligand concentrations
closely approximating those needed for equilibrium binding studies, detection of the high
affinity sites was greatly masked. Drug binding to HhaI restriction fragments which had been
prepared prior to the action of drug did not induce new bands. Furthermore, the larger DNA
fragments from drug labeled plasmid were resistant to HhaI digestion over a wide range of
enzyme concentrations. These findings suggest that ligand binding can be highly selective even
between sites which have the same tetranucleotide sequence. Therefore, selective drug binding
must be dictated not only by local base sequence preference, but also by other long range
parameters.

<>

<1>Cohen, H.M., Griffiths, A.D., Tawfik, D.S., Loakes, D.
<2>Determinants of cofactor binding to DNA methyltransferases: insights from a systematic series of structural variants of  S-adenosylhomocysteine.
<3>Org. Biomol. Chem.
<4>3
<5>152-161
<6>2005
<7>S-Adenosylmethionine (AdoMet) is a commonly used cofactor, second only to ATP in the variety
of reactions in which it participates. It is the
methyl donor in the majority of methyl transfer reactions, including
methylation of DNA, RNA, proteins and small molecules. Almost all
structurally characterised methyltransferases share a conserved
AdoMet-dependent methyltransferase fold, in which AdoMet is bound in
the same orientation. Although potential interactions between the
cofactor and methyltransferases have been inferred from crystal
structures, there has not been a systematic study of the contributions
of each functional group to binding. To explore the binding interaction
we synthesised a series of seven analogues of the methyltransferase
inhibitor S-adenosylhomocysteine (AdoHcy), each containing a single
modi cation, and tested them for the ability to inhibit methylation by
HhaI and HaeIII DNA methyltransferase. Comparison of the K-i values
highlights the structural determinants for cofactor binding, and
indicates which nucleoside and amino acid functional groups contribute
significantly to AdoMet binding. An understanding of the binding of
AdoHyc to methyltransferases will greatly assist the design of AdoMet
inhibitors.

<>

<1>Cohen, H.M., Tawfik, D.S., Griffiths, A.D.
<2>Promiscuous methylation of non-canonical DNA sites by HaeIII methyltransferase.
<3>Nucleic Acids Res.
<4>30
<5>3880-3885
<6>2002
<7>The cytosine C5 methyltransferase M.HaeIII recognises and methylates the central cytosine of
its canonical site GGCC. Here we report that M.HaeIII can also, with lower efficiency,
methylate cytosines located in a wide range of non-canonical sequences. Using bisulphite
sequencing we mapped the methyl-cytosine residues in DNA methylated in vitro and in vivo by
M.HaeIII. Methyl-cytosine residues were observed in multiple sequence contexts, most commonly,
but not exclusively, at star sites (sites differing by a single base from the canonical
sequence). The most frequently used star sites had changes at positions 1 and 4, but there is
little or no methylation at star sites changed at position 2. The rate of methylation of
non-canonical sites can be quite significant: a DNA substrate lacking a canonical site was
methylated by M.HaeIII in vitro at a rate only an order of magnitude slower than an otherwise
identical substrate containing the canonical site. In vivo methylation of non-canonical sites
may therefore be significant and may have provided the starting point for the evolution of
restriction-modification systems with novel sequence specificities.

<>

<1>Cohen, H.M., Tawfik, D.S., Griffiths, A.D.
<2>Altering the sequence specificity of HaeIII methyltransferase by directed evolution using in vitro compartmentalization.
<3>Protein Eng. Des. Sel.
<4>17
<5>3-11
<6>2004
<7>Engineering the specificity of DNA-modifying enzymes has proven extremely challenging, as
sequence recognition by these enzymes is poorly understood.  Here we used directed evolution
to generate a variant of HaeIII methyltransferase that efficiently methylates a novel target
site.  M.HaeIII methylates the internal cytosine of the canonical sequence GGCC, but there is
promiscuous methylation of a variety of non-canonical sites, notably AGCC, at a reduced rate.
Using in vitro compartmentalization, libraries of M.HaeIII genes were selected for the ability
to efficiently methylate AGCC.  A two-step mutagenesis strategy, involving initial
randomization of DNA-contracting residues followed by randomization of the loop that lies
behind these residues, yielded a mutant with a 670-fold improvement in catalytic efficiency
(kcat/KmDNA) using AGCC and a preference for AGCC over GGCC.  The mutant methylates three
sites efficiently (AGCC, CGCC and GGCC).  Indeed, it methylates CGCC slightly more efficiently
than AGCC.  However, the mutant discriminates against other non-canonical sites, including
TGCC, as effectively as the wild-type enzyme.  This study provides a rate example of a
laboratory-evolved enzyme whose catalytic efficiency surpasses that of the wild-type enzyme
with the principal substrate.

<>

<1>Cohen-Gihon, I., Israeli, O., Beth-Din, A., Levy, H., Cohen, O., Shafferman, A., Zvi, A., Chitlaru, T.
<2>Whole-Genome Sequencing of the Nonproteolytic Bacillus anthracis V770-NP1-R Strain Reveals Multiple Mutations in Peptidase Loci.
<3>Genome Announcements
<4>2
<5>e00075-14
<6>2014
<7>We report the draft whole-genome sequence of the nonproteolytic Bacillus anthracis V770-NP1-R
strain. Compared to those of other B. anthracis strains, the
genome exhibits unique mutations in multiple targets potentially affecting
proteolytic functions. One of these mutations is a deletion that disrupts the
NprR quorum-sensing regulator of the NprA protease.

<>

<1>Cohen-Karni, D., Xu, D., Apone, L., Fomenkov, A., Sun, Z.Y., Davis, P.J., Kinney, S.R.M., Yamada-Mabuchi, M., Xu, S.Y., Davis, T., Pradhan, S., Roberts, R.J., Zheng, Y.
<2>The MspJI family of modification-dependent restriction endonucleases for epigenetic studies.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>11040-11045
<6>2011
<7>MspJI is a novel modification-dependent restriction endonuclease that cleaves at a fixed
distance away from the modification site. Here, we
present the biochemical characterization of several MspJI homologs,
including FspEI, LpnPI, AspBHI, RlaI, and SgrTI. All of the enzymes
specifically recognize cytosine C5 modification (methylation or
hydroxymethylation) in DNA and cleave at a constant distance
(N-12/N-16) away from the modified cytosine. Each displays its own
sequence context preference, favoring different nucleotides flanking
the modified cytosine. By cleaving on both sides of fully modified CpG
sites, they allow the extraction of 32-base long fragments around the
modified sites from the genomic DNA. These enzymes provide powerful
tools for direct interrogation of the epigenome. For example, we show
that RlaI, an enzyme that prefers (m)CWG but not (m)CpG sites,
generates digestion patterns that differ between plant and mammalian
genomic DNA, highlighting the difference between their epigenomic
patterns. In addition, we demonstrate that deep sequencing of the
digested DNA fragments generated from these enzymes provides a feasible
method to map the modified sites in the genome. Altogether, the MspJI
family of enzymes represent appealing tools of choice for method
development in DNA epigenetic studies.

<>

<1>Coil, D.A., Badger, J.H., Forberger, H.C., Riggs, F., Madupu, R., Fedorova, N., Ward, N., Robb, F.T., Eisen, J.A.
<2>Complete Genome Sequence of the Extreme Thermophile Dictyoglomus thermophilum H-6-12.
<3>Genome Announcements
<4>2
<5>e00109-14
<6>2014
<7>Here, we present the complete genome of the extreme thermophile, Dictyoglomus thermophilum
H-6-12 (phylum Dictyoglomi), which consists of 1,959,987 bp.

<>

<1>Coil, D.A., Doctor, J.I., Lang, J.M., Darling, A.E., Eisen, J.A.
<2>Draft Genome Sequence of Kocuria sp. Strain UCD-OTCP (Phylum Actinobacteria).
<3>Genome Announcements
<4>1
<5>e00172-13
<6>2013
<7>Here, we present the draft genome of Kocuria sp. strain UCD-OTCP, a member of the phylum
Actinobacteria, isolated from a restaurant chair cushion. The assembly
contains 3,791,485 bp (G+C content of 73%) and is contained in 68 scaffolds.

<>

<1>Coil, D.A., Lo, J.R., Chen, R., Ward, N., Robb, F.T., Eisen, J.A.
<2>Draft Genome Sequence of the Arsenate-Respiring Bacterium Chrysiogenes arsenatis  Strain DSM 11915.
<3>Genome Announcements
<4>1
<5>e00953-13
<6>2013
<7>Here we present the draft genome sequence of Chrysiogenes arsenatis strain DSM 11915, only the
second genome sequence from the phylum Chrysiogenetes. This
strictly anaerobic organism was isolated from arsenic-contaminated gold mine
wastewater and respires arsenate or nitrate instead of oxygen. The assembly
contains 2,824,977 bp in 22 scaffolds.

<>

<1>Coker, O.O., Regmi, S.M., Suriyaphol, P., Chininmanu, K., Prammananan, T., Chaiprasert, A.
<2>Whole-Genome Sequence of a Multidrug-Resistant Mycobacterium tuberculosis Beijing Sequence Type 10 Isolate from an Outbreak in Thailand.
<3>Genome Announcements
<4>2
<5>e00803-14
<6>2014
<7>Infections with the Beijing family of Mycobacterium tuberculosis occur worldwide  and are
endemic in Asian countries. We present the draft genome sequence of
DS6701, a multidrug-resistant M. tuberculosis Beijing strain of sequence type 10.
The isolate is a representative of strains isolated from a multidrug-resistant
tuberculosis outbreak in Thailand.

<>

<1>Col, B., Ozkeserli, Z., Kumar, D., Ozdag, H., Alakoc, Y.D.
<2>Genome Sequence of the Boron-Tolerant and -Requiring Bacterium Bacillus boroniphilus.
<3>Genome Announcements
<4>2
<5>e00935-13
<6>2014
<7>Bacillus boroniphilus is a highly boron-tolerant bacterium that also requires this element for
its growth. The complete genome sequence of B. boroniphilus was
determined by a combination of shotgun sequencing and paired-end sequencing using
454 pyrosequencing technology. A total of 84,872,624 reads from shotgun
sequencing and a total of 194,092,510 reads from paired-end sequencing were
assembled using Newbler 2.3. The estimated size of the draft genome is 5.2 Mb.

<>

<1>Colaco, C., Sen, S., Thangavelu, M., Pinder, S., Roser, B.
<2>Extraordinary stability of enzymes dried in trehalose: simplified molecular biology.
<3>Biotechnology
<4>10
<5>1007-1011
<6>1992
<7>We show that extremely fragile biomolecules such as DNA restriction and modifying enzymes can
be dried in vitro in the presence of trehalose with no loss of activity, even after prolonged
storage. A remarkable and unexpected property of the dried enzyme preparations is their
ability to withstand prolonged exposure to temperatures as high as +70oC. This stability is
unique to trehalose and is not found with other sugars irrespective of their physical or
chemical properties. The immediate significance of these observations is the ability to
convert enzymes used in molecular biology into stable reagents. The indefinite stability and
high temperature tolerance of these dried enzymes should permit the design of convenient
formats that may be of particular significance in the automation of genome mapping and
sequencing projects. The stabilization of a wide range of biomolecules by trehalose also has
practical implications for a number of areas ranging from basic science, through health care
and agriculture, to bio-electronics.

<>

<1>Colandene, J.D., Topal, M.D.
<2>The domain organization of NaeI endonuclease: Separation of binding and catalysis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>3531-3536
<6>1998
<7>NaeI is a remarkable type II restriction endonuclease.  It must bind two recognition sequences
to cleave DNA, forms a covalent protein-DNA intermediate, and is only 1 aa change away from
topoisomerase and recombinase activity.  The latter activities apparently derive from
reactivation of a cryptic DNA ligase active site.  Here, we demonstrate that NaeI has two
protease-resistant domains, involving approximately the N-terminal and C-terminal halves of
the protein, linked by a protease-accessible region of 30 aa.  The domains were purified by
cloning.  The C-terminal domain was shown by gel mobility-shift assay to have approximately
8-fold lower DNA-binding ability than intact NaeI.  Analytical ultracentrifugation showed this
domain to be a monomer in solution.  The N-terminal domain, which contains the catalytic
region defined by random mutagenesis, do not bind DNA and was a mixture of different-sized
complexes in solution implying that it mediates self-association.  DNA greatly inhibited
proteolysis of the linker region.  The results identify the DNA-binding domain, imply that DNA
cleavage and recognition are independent and separable, and lead us to speculate about a
cleft-like structure for NaeI.

<>

<1>Colandene, J.D., Topal, M.D.
<2>Evidence for mutations that break communication between the Endo and Topo domains in NaeI endonuclease/topoisomerase.
<3>Biochemistry
<4>2000
<5>13703-13707
<6>2000
<7>NaeI is a type IIe endonuclease that interacts with two DNA recognition sequences to cleave
DNA. One DNA sequence serves as a substrate and the other serves to activate cleavage. NaeI is
divided into two domains whose structures parallel the two functionalities recognized in NaeI,
endonuclease and topoisomerase. In this study, we report evidence for mutations that break
interdomain functional communication in a NaeI-DNA complex. Deletion of the initial 124 amino
acids of the N-terminal domain of NaeI converted NaeI to a monomer, consistent with
self-association being mediated by the Endo domain.  Deletions within a small region of the
C-terminal DNA binding domain of NaeI (amino acids 182-192) altered the recognition by NaeI of
sequences flanking the NaeI recognition sequence. Substituting Ala for Arg182 within this
region had no apparent effect on DNA binding but greatly reduced the extent of DNA cleavage
even though it is not part of the catalytic Endo domain. Substituting Ala for Ile185 reduced
the extent of DNA binding about 1000-fold. Substituting Ala for Lys189 altered flanking
sequence recognition.  Residues 182-192 are away from the Endo domain responsible for cleavage
and also face away from the modeled DNA binding faces of the apoprotein crystal structure. We
propose that residues 182-192 are part of a web that mediates the flow of information between
the NaeI Endo and Topo domains.

<>

<1>Colasanti, J., Sundaresan, V.
<2>Cytosine methylated DNA synthesized by Taq polymerase used to assay methylation sensitivity of restriction endonuclease HinfI.
<3>Nucleic Acids Res.
<4>19
<5>391-394
<6>1991
<7>We have studied the resistance of cytosine methylated DNA to digestion by the
restriction endonuclease HinfI, using a simple PCR procedure to synthesize DNA
of known sequence in which every cytosine is methylated at the 5 position.  We
find that HinfI cannot digest cytosine methylated DNA at the concentrations
normally used in restriction digests.  Complete digestion is possible using a
vast excess of enzyme; under these conditions, the rate of HinfI digestion for
cytosine methylated DNA is at least 1440-fold slower than for unmethylated DNA.
The presence of an additional methylated cytosine at the degenerate position
internal to the recognition sequence does not appear to increase the resistance
to HinfI digestion.  We also tested HhaII, an isoschizomer of HinfI, and found
that it is completely inactive on cytosine methylated DNA.  The procedure we
have used should be of general applicability in determination of the
methylation sensitivities of other restriction enzymes, as well as studies of
the effects of methylation on gene expression in direct DNA transfer
experiments.

<>

<1>Cole, S.
<2>Comparative mycobacterial genomics as a tool for identifying targets for the diagnosis, prophylaxis or treatment of mycobacterioses.
<3>International Patent Office
<4>WO 02074903 A
<5>
<6>2002
<7>The present invention is directed to a method of selection of purified nucleotidic sequences
or polynucleotides encoding proteins or part of proteins carrying at least an essential
function for the survival or the virulence of mycobacterium species by a comparative genomic
analysis of the sequence of the genome of M. tuberculosis aligned on the genome sequence of M.
leprae and M. tuberculosis and M. leprae marker polypeptides of nucleotides encoding the
polypeptides, and methods for using the nucleotides and the encoded polypeptides are
disclosed.

<>

<1>Cole, S.T. et al.
<2>Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence.
<3>Nature
<4>393
<5>537-544
<6>1998
<7>Countless millions of people have died from tuberculosis, a chronic infectious disease caused
by the tubercle bacillus.  The complete genome sequence of the best-characterized strain of
Mycobacterium tuberculosis, H37Rv, has been determined and analysed in order to improve our
understanding of the biology of this slow-growing pathogen and to help the conception of new
prophylactic and therapeutic interventions.  The genome comprises 4,411,529 base pairs,
contains around 4,000 genes, and has a very high guanine + cytosine content that is reflected
in the biased amino-acid content of the proteins. M. tuberculosis differs radically from other
bacteria in that a very large portion of its coding capacity is devoted to the production of
enzymes involved in lipogenesis and lipolysis, and to two new families of glycine-rich
proteins with a repetitive structure that may represent a source of antigenic variation.

<>

<1>Cole, S.T. et al.
<2>Massive gene decay in the leprosy bacillus.
<3>Nature
<4>409
<5>1007-1011
<6>2001
<7>Leprosy, a chronic human neurological disease, results from infection with the obligate
intracellular pathogen Mycobacterium leprae, a close relative of the tubercle bacillus.
Mycobacterium leprae has the longest doubling time of all known bacteria and has thwarted
every effort at culture in the laboratory.  Comparing the 3.27-megabase genome sequence of an
armadillo-derived Indian isolate of the leprosy bacillus with that of Mycobacterium
tuberculosis (4.41 Mb) provides clear explanations for these properties and reveals an extreme
case of reductive evolution.  Less than half of the genome contains functional genes but
pseudogenes, with intact counterparts in M. tuberculosis, abound.  Genome downsizing and the
current mosaic arrangement appear to have resulted from extensive recombination events between
dispersed repetitive sequences.  Gene deletion and decay have eliminated many important
metabolic activities including siderophore production, part of the oxidative and most of the
microaerophilic and anaerobic respiratory chains, and numerous catabolic systems and their
regulatory circuits.

<>

<1>Coleman, M.L., Sullivan, M.B., Martiny, A.C., Steglich, C., Barry, K., Delong, E.F., Chisholm, S.W.
<2>Genomic islands and the ecology and evolution of Prochlorococcus.
<3>Science
<4>311
<5>1768-1770
<6>2006
<7>Prochlorococcus ecotypes are a useful system for exploring the origin and function of
diversity among closely related microbes. The genetic
variability between phenotypically distinct strains that differ by less
that 1% in 16S ribosomal RNA sequences occurs mostly in genomic islands.
Island genes appear to have been acquired in part by phage-mediated
lateral gene transfer, and some are differentially expressed under light
and nutrient stress. Furthermore, genome fragments directly recovered from
ocean ecosystems indicate that these islands are variable among
cooccurring Prochlorococcus cells. Genomic islands in this free-living
photoautotroph share features with pathogenicity islands of parasitic
bacteria, suggesting a general mechanism for niche differentiation in
microbial species.

<>

<1>Coleman, N.V. et al.
<2>Genome sequence of the ethene- and vinyl chloride-oxidizing actinomycete Nocardioides sp. strain JS614.
<3>J. Bacteriol.
<4>193
<5>3399-3400
<6>2011
<7>Nocardioides sp. strain JS614 grows on ethene and vinyl chloride (VC) as sole carbon and
energy sources, and is of interest for bioremediation and
biocatalysis. Sequencing the complete genome of JS614 provides insight
into the genetic basis of alkene oxidation, supports ongoing research into
the physiology and biochemistry of growth on ethene and VC, and provides
biomarkers to facilitate detection of VC/ethene-oxidizers in the
environment. This is the first genome sequence from the genus
Nocardioides, and the first genome of a VC/ethene-oxidizing bacterium.

<>

<1>Colleaux, L., d'Auriol, L., Betermier, M., Cottarel, G., Jacquier, A., Galibert, F., Dujon, B.
<2>Universal code equivalent of a yeast mitochondrial intron reading frame is expressed into E. coli as a specific double strand endonuclease.
<3>Cell
<4>44
<5>521-533
<6>1986
<7>The intron of the mitochondrial 21S rRNA gene of Saccharomyces cerevisiae (r1 intron)
possesses a 235 codon long internal open reading frame (r1 ORF) whose translation product
determines the duplicative transposition of that intron during crosses between intron-plus
strains (omega+) and intron-minus ones (omega-). Using site-directed mutagenesis, we have
constructed a universal code equivalent of the r1 ORF that, under appropriate promoter
control, allows the overexpression in E. coli of a protein identical to the mitochondrial
intron encoded "transposase". This protein exhibits a double strand endonuclease activity
specific for the omega site. This finding demonstrates, for the first time, the enzymatic
activity of an intron encoded protein whose function is to promote the spreading of that
intron by generating double strand breaks at a specific sequence within a gene.

<>

<1>Colleaux, L., D'Auriol, L., Galibert, F., Dujon, B.
<2>Recognition and cleavage site of the intron-encoded omega transposase.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>85
<5>6022-6026
<6>1988
<7>The optional group I intron of the mitochondrial 21S rRNA gene of Saccharomyces cerevisiae
contains a 235-codon-long open reading frame the translation product of which (the omega
transposase) catalyzes the formation of a double-strand break within the intron-minus (omega-)
copies of the same gene. Purified omega transposase generates in vitro a 4-base-pair staggered
cut with 3' hydroxyl overhangs at the extact position where the intron eventually inserts in
the gene. Using randomly mutagenized synthetic olitonucleotides, single-base mutants were
produced at 21 positions around the cleavage site. Experiments with these oligonucleotides
show that the recognition site extends over an 18-base pair-long sequence within which minimal
sequence degeneracy is tolerated. The intron-encoded omega transposase is, therefore, one of
the most specific restriction endonucleases known to date.

<>

<1>Colleaux, L., Michel-Wolwertz, M.-R., Matagne, R.F., Dujon, B.
<2>The apocytochrome b gene of Chlamydomonas smithii contains a mobile intron related to both Saccharomyces and Neurospora introns.
<3>Mol. Gen. Genet.
<4>223
<5>288-296
<6>1990
<7>The mitochondrial DNA of the two interfertile algal species Chlamydomonas smithii and
Chlamydomonas reinhardtii are co-linear with the exception of ca. 1 kb insertion (the alpha
insert) present in C. smithii DNA only. In vegetative diploids resulting from interspecific
crosses, mitochondrial genomes are transmitted biparentally except for the alpha insert which
is transmitted to all C. reinhardtii molecules in a manner reminiscent of the intron-mediated
conversion event that occurs at the omega locus in yeast mitochondria, under the action of the
I-SceI endonuclease. Here we report that the alpha insert corresponds to a typical group I
intron of 1075 bp, inserted within the gene for apocytochrome b and containing a 237 codon
open reading frame (ORF). We also report the complete sequence of the apocytochrome b gene of
C. smithii. Comparison with the sequence of the same gene in C. reinhardtii reveals the
precise intron insertion site. These data, together with the previous genetic data provide the
first example of intron mobility in mitochondria of the plant kingdom. The product of the
intronic ORF shows 36% amino acid identity with the I-SceI endonuclease whereas the intron
ribozyme shows a 60% identity at the nucleotide level with the Neurospora crassa cob 1 intron.
The possibility of a recent horizontal transfer of introns between fungi and algae is
discussed.

<>

<1>Collier, D.A., Thuong, N.T., Helene, C.
<2>Sequence-specific bifunctional DNA ligands based on triple-helix-forming oligonucleotides inhibit restriction enzyme cleavage under physiological conditions.
<3>J. Am. Chem. Soc.
<4>113
<5>1457-1458
<6>1991
<7>Intermolecular triplex formation has been demonstrated to inhibit DNA-protein
interactions by the observation that selective binding of a third strand of DNA
at the recognition site of restriction/modification enzymes prevents cleavage
or methylation of the duplex.  Triplex formation has the potential to precisely
modulate gene expression if targeted at protein binding sites involved in the
regulation of a specific gene.  Intermolecular triplex formation occurs by
binding of a homopyrimidine oligonucleotide to the major grrove of a
homopurine-homopyrimidine stretch of DNA, parallel to the purine strand.
Sequence specificity results from Hoogsteen pairing between thymine and
protonated cytosine in the third pyrimidine strand and the Watson-Crick A.T and
G.C pairs of the duplex, respectively.

<>

<1>Collier, G.B.
<2>Regulation of intracellular S-adenosyl-L-methionine as a strategy for cloning restriction endonucleases.
<3>Diss. Abstr.
<4>56
<5>73B
<6>1995
<7>Available strategies to clone restriction endonucleases (REs) are dependent on selection of
the cognate DNA methyltransferase (Mtase), which are closely linked to the REs.  These
strategies are indirect and less successful at cloning REs.  Therefore, a novel RE cloning
strategy was conceived, proposed and developed.  This novel strategy is based on modulating
the intracellular S-adenosyl-L-methionine (AdoMet) levels.  DNA Mtases require the methyl
cofactor AdoMet to effect DNA methylation.  By reducing the intracellular AdoMet levels, the
DNA Mtase is unable to effect DNA methylation.  Restriction endonucleases are present only
with their cognate DNA methyltransferases to protect the host chromosomal DNA from degradation
by its own expressed RE genes.  Under reduced levels of AdoMet the DNA Mtase is rendered
ineffective and the RE will degrade the chromosomal DNA.  To reduce the intracellular AdoMet
concentrations, several strategies were considered and bacteriophage T3
S-adenosyl-L-methionine hydrolase (SAMase) was found to be useful for the development of this
system.  To tightly control the expression of SAMase a novel expression system based on the
TN10 tetracycline regulation, inducible with heated chlortetracycline, was developed.  The
effect of SAMase in this system was first optimized by using a cloned M.BamHII (BamHII Mtase)
to develop conditions to prevent methylation of this plasmid.  This in vivo reduction in DNA
methylation provides opportunities for cellular studies involved with DNA methylation.  Since
REs generate double stranded DNA breaks when the DNA is unprotected by the cognate DNA Mtase,
a system was developed to monitor and assay for these breaks.  A chromosomal
dinD1::beta-galactosidase promotor gene fusion, induced by double stranded DNA breaks, was P1
transduced into an appropriate E. coli host.  The tetracycline regulon controlled SAMase was
supplied on plasmid pACYC184 and the compatible (colE1 replicon) RM.EcoRI plasmid pRI13 was
also present.  Inducing the expression of SAMase caused the dinD1 promotor to express the
fused beta-galactosidase, suggesting that the cell was experiencing DNA double strand breaks
due to the active RE, since the DNA Mtase was affected by reduction of the level of AdoMet.
This data suggests that this novel cloning strategy is feasible.

<>

<1>Collier, G.B., Connaughton, J.F., Chirikjian, J.G.
<2>Cloning restriction enodnuclease genes by modulating methyltransferase activity.
<3>US Patent Office
<4>US 5451519
<5>
<6>1995
<7>The present invention relates to a method for cloning genes that encode restriction
endonucleases by altering the level of a methyl donor co-factor of a DNA methyltransferase
that protects the DNA of a host cell from damage by a restriction endonuclease.  The method
can be used to screen entire DNA libraries en masse to identify clones that encode restriction
enzymes by growing one library replicate under high or normal methyl donor conditions to
protect host DNA and a second library replicate under low methyl donor conditions allowing DNA
damage from the active restriction endonuclease.  Clones that encode a restriction enzyme are
identified by decreased growth or color produced in response to double stranded DNA damage
under the low methyl donor conditions.  Colorimetric methods useful in the invention can use
SOS-sensitive promoters operably linked to beta-galactosidase, which detect DNA damage.

<>

<1>Collier, G.B., Mattson, T.L., Connaughton, J.F., Kalloss, W.D.
<2>Development of a novel screening system for cloning restriction endonucleases.
<3>FASEB J.
<4>7
<5>A1303
<6>1993
<7>We have developed a novel screening system for the cloning of prokaryotic restriction
endonucleases. This system differs from existing strategies, since it is a screen for the
restriction endonuclease. Present restriction endonuclease cloning strategies are dependent on
screening or selecting for the cognate methyltransferase. The cognate methyltransferase
appears to be necessary to prevent cellular death due to enzymatic cleavage of the host
chromosomal DNA. This cloning strategy is dependent on modulating the intracellular levels of
S-adenosyl-L-methionine, the biological methyl donor used by DNA methyltransferases. In the
presence of a functional restriction endonuclease structural gene and limiting levels of
S-adenosyl-L-methionine the chromosomal DNA is not methylated but is cleaved, causing cellular
death or the induction of the SOS response. The methodology of modulating
S-adenosyl-L-methionine has further application to other cellular phenomena.

<>

<1>Collier, J., McAdams, H.H., Shapiro, L.
<2>A DNA methylation ratchet governs progression through a bacterial cell cycle.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>17111-17116
<6>2007
<7>The Caulobacter cell cycle is driven by a cascade of transient regulators, starting with the
expression of DnaA in G(1) and ending with the
expression of the essential CcrM DNA methyltransferase at the completion
of DNA replication. The timing of DnaA accumulation was found to be
regulated by the methylation state of the dnaA promoter, which in turn
depends on the chromosomal position of dnaA near the origin of replication
and restriction of CcrM synthesis to the end of the cell cycle. The dnaA
gene is preferentially transcribed from a fully methylated promoter. DnaA
initiates DNA replication and activates the transcription of the next
cell-cycle regulator, GcrA. With the passage of the replication fork, the
dnaA promoter becomes hemimethylated, and DnaA accumulation drops. GcrA
then activates the transcription of the next cell-cycle regulator, CtrA,
once the replication fork passes through the ctrA P1 promoter, generating
two hemimethylated copies of ctrA. The ctrA gene is preferentially
transcribed from a hemimethylated promoter. CtrA then activates the
transcription of ccrM, to bring the newly replicated chromosome to the
fully methylated state, promoting dnaA transcription and the start of a
new cell cycle. We show that the cell-cycle timing of CcrM is critical for
Caulobacter fitness. The sequential changes in the chromosomal methylation
state serve to couple the progression of DNA replication to cell-cycle
events regulated by the master transcriptional regulatory cascade, thus
providing a ratchet mechanism for robust cell-cycle control.

<>

<1>Collingro, A., Tischler, P., Weinmaier, T., Penz, T., Heinz, E., Brunham, R.C., Read, T.D., Bavoil, P.M., Sachse, K., Kahane, S., Friedman, M.G., Rattei, T., Myers, G.S.A., Horn, M.
<2>Unity in variety -- the pan-genome of the Chlamydiae.
<3>Mol. Biol. Evol.
<4>28
<5>3253-3270
<6>2011
<7>Chlamydiae are evolutionarily well-separated bacteria that live exclusively within eukaryotic
host cells.  They include important human pathogens such as Chlamydia trachomatis as well as
symbionts of protozoa.  As these bacteria are experimentally challenging and genetically
intractable, our knowledge about them is still limited.  In this study, we obtained the genome
sequences of Simkania negevensis Z, Waddlia chondrophila 2032/99 and Parachlamydia
acanthamoebae UV-7.  This enabled us to perform the first comprehensive comparative and
phylogenomic analysis of representative members of four major families of the Chlamydiae,
including the Chlamydiaceae.  We identifid a surprisingly large core gene set present in all
genomes and a high number of diverse accessory genes in those Chlamydiae that do not primarily
infect humans or animals, including a chemosensory system in P. acanthamoebae and a type IV
secretion system.  In S. negevensis, the type IV secretion system is encoded on a large
conjugative plasmid (pSn, 132 kb).  Phylogenetic analyses suggested that a plasmid similar to
the S. negevensis plasmid was originally acquired by the lst common ancestor of all four
families and that it was subsequently reduced, integrated into the chromosome, or lost during
diversification, ultimately giving rise to the extant virulence-associated plasmid of
pathogenic chlamydiae.  Other virulence factors, including a type III secretion system, are
conserved among the Chlamydiae to variable degrees, and together with differences in the
composition of the cell wall, reflect adaptation to different host cells including convergent
evolution among the four chlamydial families.  Phylogenomic analysis focusing on chlamydial
proteins with homology to plant proteins provided evidence for the aquisition of 53 chlamydial
genes by a plant progenitor, lending further support for the hypothesis of an early
interaction between a chlamydial ancestor and the primary photosynthetic eukaryote.

<>

<1>Collins, A.J., Nyholm, S.V.
<2>Draft genome of Phaeobacter gallaeciensis ANG1, a dominant member of the accessory nidamental gland of Euprymna scolopes.
<3>J. Bacteriol.
<4>193
<5>3397-3398
<6>2011
<7>Phaeobacter gallaeciensis strain ANG1 represents the dominant member of the bacterial
consortium within the reproductive accessory nidamental
gland (ANG) of the squid Euprymna scolopes. We present a 4.59Mb assembly
of its genome, which may provide clues as to how it benefits its host.

<>

<1>Collins, E.B.
<2>Host-controlled variations in bacteriophages active against lactic Streptococci.
<3>Virology
<4>2
<5>261-271
<6>1956
<7>Certain bacteriophages were very active against some strains of Streptococcus
cremoris, and restricted in activity against others.  Drastic changes in
activity were observed after one growth cycle.  The results of adsorption of a
restricted bacteriophage was either no detected consequence, bacterial death,
or bacterial death followed by bacteriophage reproduction.  The occurrence of
each of the last two possibilities was found to be influenced by the
multiplicity of infection.  Bacteriophage reproduction within the fruitful
cells of a restrictive host produced progeny that were fully active on the
cells of that host.  Alteration of the bacteriophgaes appeared limited to
changes in virulence for particular hosts, the alterations not being
accompanied by changes in adsorbability or changes in heat resistance.  Both
the strain of infecting bacteriophage and the particular host were important
factors in determining the specific virulence of the progeny.

<>

<1>Collins, J., Mayer, H.
<2>Restriction endonucleases or the site-specific DNA endonucleases.
<3>Arzneimittelforschung
<4>30
<5>541-547
<6>1980
<7>Our present view of the site-specific endonucleases, which appear to be
ubiquituous in the prokaryote kingdom, is probably heavily distorted by our
search for tools for recombinant DNA technology.  Only those enzymes having
recognition sequences in the range of three to seven specific bases have been
isolated.  Of course the usefulness of these enzymes in the analysis of complex
genomes, the rise of "reverse genetics", and the immediate breakthroughs in the
area of gene expression in eukaryotes, particularly the understanding of tumour
virus RNA processing and gene rearrangements in the expression of
immunoglobulin genes has dominated the consciousness of the molecular and
cell-biologists during the last five years.  There is great diversity of
staggering, symmetry, asymmetry and degeneration in the recognition sequences
found.  Taking into account also the genetic data on site-specific
recombination and/or DNA degradation suggests that our present collection of
endonucleases may only represent a narrow spectrum of specificities on an
open-ended scale of complexity.  The enzymes themselves provide a rich pool to
be exploited by the biophysicist and the biochemist to probe the subtleties of
DNA-protein interaction.

<>

<1>Collins, R.E., Rocap, G.
<2>REPK: an analytical web server to select restriction endonucleases for terminal restriction fragment length polymorphism analysis.
<3>Nucleic Acids Res.
<4>35
<5>W58-W62
<6>2007
<7>Terminal restriction fragment length polymorphism (T-RFLP) analysis is a widespread technique
for rapidly fingerprinting microbial communities.
Users of T-RFLP frequently overlook the resolving power of well-chosen
restriction endonucleases and often fail to report how they chose their
enzymes. REPK (Restriction Endonuclease Picker) assists in the rational
choice of restriction endonucleases for T-RFLP by finding sets of four
restriction endonucleases that together uniquely differentiate
user-designated sequence groups. With REPK, users can provide their own
sequences (of any gene, not just 16S rRNA), specify the taxonomic rank of
interest and choose from a number of filtering options to further narrow
down the enzyme selection. Bug tracking is provided, and the source code
is open and accessible under the GNU Public License v.2, at
http://code.google.com/p/repk. The web server is available without access
restrictions at http://rocaplab.ocean.washington.edu/tools/repk.

<>

<1>Collyn, F., Billault, A., Mullet, C., Simonet, M., Marceau, M.
<2>YAPI, a new Yersinia pseudotuberculosis pathogenicity island.
<3>Infect. Immun.
<4>72
<5>4784-4790
<6>2004
<7>Pathogenicity islands (PAIs) are chromosomal clusters of pathogen-specific virulence genes
often found at tRNA loci. In the
Yersinia pseudotuberculosis 32777 chromosome, we characterized a 98-kb
segment that has all of the characteristic features of a PAI, including
insertion in a (phenylalanine) tRNA gene, the presence of a
bacteriophage-like integrase-encoding gene, and direct repeats at the
integration sites. The G+C content of the segment ranges from 31 to
60%, reflecting a genetic mosaic: this is consistent with the notion
that the sequences were horizontally acquired. The PAI, termed YAPI
(for Yersinia adhesion pathogenicity island), carries 95 open reading
frames and includes (i) the previously described pil operon, encoding a
type IV pilus that contributes to pathogenicity (F. Collyn et al.,
Infect. Immun. 70:6196-6205, 2002); (ii) a block of genes potentially
involved in general metabolism; (iii) a gene cluster for a
restriction-modification system; and (iv) a large number of mobile
genetic elements. Furthermore, the PAI can excise itself from the
chromosome at low frequency and in a precise manner, and deletion does
not result in a significant decrease of bacterial virulence compared to
inactivation of the fimbrial gene cluster alone. The prevalence and
size of the PAI vary from one Y. pseudotuberculosis strain to another,
and it can be found integrated into either of the two phe tRNA loci
present on the species' chromosome. YAPI was not detected in the genome
of the genetically closely related species Y. pestis, whereas a
homologous PAI is harbored by the Y. enterocolitica chromosome.

<>

<1>Colson, A.M., Colson, C.
<2>Expression of the Escherichia coli K, B and Phage P1 DNA host specificities in Salmonella typhimurium.
<3>J. Gen. Microbiol.
<4>70
<5>123-128
<6>1972
<7>The Escherichia coli K,B and phage P1 host specificity genes (hsp) were
introduced into Salmonella typhimurium by means of E. coli Hfr x S. typhimurium
F- crosses.  The three systems were found to govern modification and
restriction of phages P22 and L.  The restriction coefficients were comparable
to that of lambda for the PI system and were lower (about 100) for the K and B
systems.  Phage grown on hspK+ and hspB+ recombinants was restricted by S.
typhimurium independently of the restriction governed by the LT system.  This
observation suggested that S. typhimurium possesses a previously undetected hsp
locus allelic to those of E. coli K and B.

<>

<1>Colson, A.M., Colson, C., Van Pel, A.
<2>Host-controlled restriction mutants of Salmonella typhimurium.
<3>J. Gen. Microbiol.
<4>58
<5>57-64
<6>1969
<7>Forty-eight independent restriction-deficient mutations of Salmonella typhimurium LT2 were
isolated by using selective and non-selective methods.  With phage P22 it was shown that some
mutations affected the restriction capacity only, while others affected both restriction and
modification.  The host-restriction of S. typhimurium decreased the recovery of F-lac+
infected cells and decreased the yield of recombinants in bacterial mating and in phage
P22-mediated transduction.

<>

<1>Colson, C.
<2>Genetics of R-M systems in Salmonella.
<3>Heredity
<4>41
<5>123
<6>1978
<7>Salmonella typhimurium LT2 and LT7 have three R-M systems coded by chromosomal genes.  Mutants
with r-m+ and r-m- phenotypes have been isolated for each of them.  All three were first
detected in Escherichia coli-S. typhimurium hybrids.  System LT was detected when phage P22
from lac+ hybrids between E. coli Hfr and S. typhimurium LT7 mut (with a mutator gene) was
plated on wild-type S. typhimurium.  All mutations affecting system LT are closely linked and
situated between proA and proC.  System LT is present in the many Salmonella serotypes tested,
with the exception of S. typhi.  System SA was first observed using hybrids with the hsdK
genes of E. coli: phage L. (closely related to P22) from such hybrids underwent restriction in
wild-type S. typhimurium, independently of the presence of system LT.  hsdSA is situated
between pyrB and serB and was first thought to be the Salmonella allele of hsdK and hsdB in E.
coli.  System SA was found in all S. typhimurium strains tested, but not in other Salmonella
serotypes.

<>

<1>Colson, C., Colson, A.M.
<2>A new Salmonella typhimurium DNA host specificity.
<3>J. Gen. Microbiol.
<4>69
<5>345-351
<6>1971
<7>The genetic properties of a new (hspS) host specificity of Salmonella
typhimurium were investigated using bacteriophage L.  Phage L is a better
substrate for S-specific restriction than phage P22.  Mutants deficient in
S-restriction only were found at the same frequency as mutants deficient in
both restriction and modification.  Crosses between S. typhimurium Hfr and S.
typhimurium F- or between Escherichia coli and S. typhimurium showed that the S
system has the same chromosomal location as the K system of E. coli.  The S
system was introduced in E. coli and found to be effective on phage lambda.

<>

<1>Colson, C., Colson, A.M., van Pel, A.
<2>Chromosomal location of host specificity in Salmonella typhimurium.
<3>J. Gen. Microbiol.
<4>60
<5>265-271
<6>1970
<7>The chromosomal location of the genes for host specificity in Salmonella
typhimurium has been investigated by F-mediated conjugation using host
specificity mutants isolated previously.  It was found that the sites of
mutations leading to two distinct phenotypes r-LTm+LT and r-LTm-LT, are closely
linked to each other and are located near the marker proC.

<>

<1>Colson, C., Glover, S.W., Symonds, N., Stacey, K.A.
<2>The location of the genes for host-controlled modification and restriction in Escherichia coli K-12.
<3>Genetics
<4>52
<5>1043-1050
<6>1965
<7>Recent experiments have clarified certain aspects of the processes of
modification and restriction by which certain systems of host-controlled
modification (HCM) operate in Escherichia coli (Arber 1962; Arber and Dussoix
1962; Dussoix and Arber 1962; Glover, Schell, Symonds and Stacey 1963).  The
two aspects of HCM have one thing in common:  they both involve the recognition
of a particular base sequence in DNA.  However, basically they are very
different, modification resulting in the addition of certain groups to DNA,
while restriction leaves unmodified DNA in a condition in which it is open to
attack by DNase.  This apparent dissimilarity in the processes of restriction
and modification suggests that they are under independent genetic control.  One
method of learning more about the genetic control of HCM in this system is to
analyze it genetically.

<>

<1>Colson, C., van Pel, A.
<2>DNA restriction and modification systems in Salmonella.  I.  SA and SB, two Salmonella typhimurium systems determined by genes with a chromosomal location comparable to that of the Escherichia coli hsd genes.
<3>Mol. Gen. Genet.
<4>129
<5>325-337
<6>1974
<7>Haploid hybrids between Salmonella typhimurium Hfr and Escherichia coli F-
exercise two additive types of restriction and modification (SA and SB) on
phage lambda.  System SA had been detected previously in S. typhimurium with
phage L.  Independent mutants in the SA and SB systems were isolated.  P22- and
P1-mediated transductions in S. typhimurium and in hybrids established that the
genes governing these systems are independent but linked and situated
counter-clockwise of ser B on the map, in the order:  pyrB-hsdSA-hsdSB-serB.

<>

<1>Colston, M.J., Davis, E.O.
<2>Homologous recombination, DNA repair and mycobacterial recA genes.
<3>Tuberculosis: Pathogenesis, Protection, and Control, American Society for Microbiology, Bloom, B.R., Washington, DC
<4>0
<5>217-226
<6>1994
<7>Bacterial responses to DNA damage are highly conserved.  One system, the
SOS response, involves the coordinately induced expression of over 20 genes through a
common regulatory mechanism.  The RecA protein, found in most bacteria, plays a central
role in the regulation of the SOS response.  In addition to its regulatory function, this
protein also mediates genetic recombination and DNA repair.  Virtually nothing is known
about these systems in mycobacteria.  However, since some mycobacteria are intracellular
pathogens and many of the mechanisms involved in macrophage killing of such pathogens
require the production of DNA-damaging agents such as peroxide and nitric oxide, DNA
repair mechanisms are likely to be particularly important for mycobacterial survival.  In
addition, homologous genetic recombination, a process mediated by RecA, is an important
technique for the genetic manipulation of bacteria and could play an important role in our
understanding of gene function.  In this chapter we discuss the mechanisms involved in
homologous recombination and DNA repair, what is known about these systems in
mycobacteria, and recent information on the unusual structure of the recA gene in the
pathogenic mycobacteria.

<>

<1>Comai, L., Young, K., Till, B.J., Reynolds, S., Greene, E.A., Codomo, C.A., Enns, L., Johnson, J.E., Burtner, C., Odden, A., Henikoff, S.
<2>Efficient discovery of DNA polymorphisms in natural populations by Ecotilling.
<3>Plant J.
<4>37
<5>778-786
<6>2004
<7>We have adapted the mutation detection technology used in Targeting Induced Local Lesions in
Genomes (TILLING) to the discovery of polymorphisms in natural populations.  The genomic DNA
of a queried individual is mixed with a references DNA and used to amplify a target 1-kbp
region of DNA with asymmetrically labeled fluorescent primers.  Afer heating and annealing,
heteroduplexes are nicked at mismatched sites by the endonuclease CEL I and cut strands are
visualized using Li-cor gel analyzers.  Putative polymorphisms detected in one fluorescence
channel can be verified by appearance of the opposite cut strand in the other channel.  We
demonstrated the efficiency of this technology, called Ecotilling, by the discovery in 150+
individuals of 55 haplotypes in five genes, ranging from sequences differing by a single
nucleotide polymorphism to those representing complex haplotypes in five genes, ranging from
sequences differing by a single nucleotide polymorphism to those representing complex
haplotypes.  The discovered polymorphisms were confirmed by sequencing and included base-pair
changes, small insertions and deletions, and variation in microsatellite repeat number.
Ecotilling allows the rapid detection of variation in many individuals and is cost effective
because only one individual for each haplotype needs to be sequenced.  The technology is
applicable to any organism including those that are heterozygous and polyploid.

<>

<1>Comandatore, F., Gaibani, P., Ambretti, S., Landini, M.P., Daffonchio, D., Marone, P., Sambri, V., Bandi, C., Sassera, D.
<2>Draft Genome of Klebsiella pneumoniae Sequence Type 512, a Multidrug-Resistant Strain Isolated during a Recent KPC Outbreak in Italy.
<3>Genome Announcements
<4>1
<5>e00035-12
<6>2013
<7>Here, we present the draft genome sequence of Klebsiella pneumoniae subsp. pneumoniae sequence
type 512 (ST512) isolated during a KPC-producer outbreak. This strain is resistant to
beta-lactams, cephalosporins, fluoroquinolones, aminoglycosides, macrolides, tetracyclines,
and carbapenems but susceptible to colistin. The ST512-K30BO genome is composed of 289 contigs
for 5,392,844 bp with 56.9% G+C content.

<>

<1>Comandatore, F., Sassera, D., Ambretti, S., Landini, M.P., Daffonchio, D., Marone, P., Sambri, V., Bandi, C., Gaibani, P.
<2>Draft Genome Sequences of Two Multidrug Resistant Klebsiella pneumoniae ST258 Isolates Resistant to Colistin.
<3>Genome Announcements
<4>1
<5>e00113-12
<6>2013
<7>Sequence type 258 (ST258) is the most widespread multidrug resistant (MDR) Klebsiella
pneumoniae strain worldwide. Here, we report the draft genome sequences of two
colistin-resistant MDR K. pneumoniae ST258 clinical strains isolated from hospital patients in
Italy. These strains are resistant to beta-lactams, cephalosporins, fluoroquinolones,
aminoglycosides, macrolides, tetracyclines, carbapenems, and colistin.

<>

<1>Comstock, L.R., Rajski, S.R.
<2>Conversion of DNA methyltransferases into azidonucleosidyl transferases via synthetic cofactors.
<3>Nucleic Acids Res.
<4>33
<5>1644-1652
<6>2005
<7>Aziridine-based cofactor mimics have been synthesized and are shown to undergo
methyltransferase-dependent DNA alkylation. Notably, each cofactor
mimic possesses an azide functionality, to which can be attached an
assortment of unnatural groups following methyltransferase-dependent DNA
delivery. DNA duplexes modified with these cofactor mimics are capable of
undergoing the Staudinger ligation with phosphines tethered to biological
functionalities following enzymatic modification. This methodology
provides a new tool by which to selectively modify DNA in a
methyltransferase-dependent way. The conversion of biological
methyltransferases into azidonucleosidyl transferases demonstrated here
also holds tremendous promise as a means of identifying, as yet, unknown
substrates of methylation.

<>

<1>Comstock, L.R., Rajski, S.R.
<2>Methyltransferase-directed DNA strand scission.
<3>J. Am. Chem. Soc.
<4>127
<5>14136-14137
<6>2005
<7>The study of prokaryotic DNA methyltransferases (MTases) has provided significant insight into
eukaryotic MTases and the important role that methylation plays in mammalian biology.  The
prokaryotic enzymes M.TaqI, M.EcoRI, and M.HhaI are all capable of using 5'-aziridine
adenylate 1 in place of (S)-adenosyl-L-methionine in MTase-dependent DNA alkylation reactions.
By virtue of the C8 azide, 1 is significant because it is capable of converting these DNA
MTases into azidonucleoside transferases.  Not suprising, DNA modified with 1 is capable of
undergoing very efficient Staudinger ligation with biotinylated triarylphosphines.

<>

<1>Conlan, L.H., Dupureur, C.M.
<2>Dissecting the metal ion dependence of DNA binding by PvuII endonuclease.
<3>Biochemistry
<4>41
<5>1335-1342
<6>2002
<7>Divalent cations can provide an effective means of modulating the behavior of nucleic acid
binding proteins. As a result, there is strong interest in understanding the role of metal
ions in the function of both nucleic acid binding proteins and their enzymes. We have applied
complementary fluorescence spectroscopic and nitrocellulose filter binding assays to
quantitate the role of metal ions in mediating DNA binding and sequence specificity by the
representative PvuII endonuclease. At pH 7.5 in the presence of the catalytically
nonsupportive Ca(II), this enzyme binds the PvuII target sequence with a K(d) of 50 pM. Under
strict metal-free conditions, the enzyme exhibits a K(d) of only 300 nM for the cognate
sequence, an affinity which is weak relative to those measured for other systems in the
absence of metal ions. This represents a 6000-fold increase in PvuII affinity for cognate DNA
upon the addition of Ca(II). The pH dependences of both metal ion-dependent and metal
ion-independent DNA binding are remarkably shallow throughout the physiological range; other
characterized restriction enzymes exhibit more pronounced pH dependences of DNA binding even
in the absence of metal ions. Similar measurements with noncognate sequences indicate that
divalent metal ions are not important to nonspecific DNA binding; K(d) values are
approximately 200 nM throughout the physiological pH range, a behavior shared with other
endonucleases. While some of these results extend somewhat the range of expected behavior for
restriction enzymes, these results indicate that PvuII endonuclease shares with other
characterized systems a mechanism by which cognate affinity and sequence discrimination are
most effectively achieved in the presence of divalent metal ions.

<>

<1>Conlan, L.H., Jose, T.J., Thornton, K.C., Dupureur, C.M.
<2>Modulating restriction endonuclease activities and specificities using neutral detergents.
<3>Biotechniques
<4>27
<5>955-960
<6>1999
<7>It is well known that type II restriction enzyme activities and specificities can be modulated
by altering solution conditions. The addition of co-solvents such as dimethyl sulfoxide
(DMSO), alcohols and polyols can promote star activity, which is the cleavage of non-cognate
sequences. While neutral detergents are often used to control protein aggregation, little is
known about the effect of neutral detergents on restriction enzyme activities and
specificities. We report here that BamHI, BglI, BglII, EcoRI, EcoRV, HindIII, MluI, PvuII,
SalI and XhoI restriction endonucleases are remarkably tolerant of high concentrations of
neutral detergents, Triton X-100, CHAPS and octyl glucoside. In most cases, lambda DNA
cleavage rates were comparable to those observed in the absence of detergent. Indeed, the
specific activities of SalI and XhoI were appreciably increased in the presence of Triton
X-100. For all enzymes active in the presence of detergents, sequence specificity toward
lambda DNA was not compromised. Assays of star cleavage of pUC18 by EcoRI, PvuII and BamHI
endonucleases in equimolar concentrations of Triton X-100 and sucrose revealed reduced star
activity in the detergent relative to the sucrose co-solvent. Interestingly, under star
activity-promoting
conditions, PvuII endonuclease displayed greater fidelity in Triton X-100 than in conventional
buffer. Taken altogether, these results suggest that in some cases, neutral detergents can be
used to manipulate restriction endonuclease reaction rates and specificities.

<>

<1>Conlan, L.M.
<2>The influence of divalent metal ions on DNA binding and cleavage by the restriction enzyme PvuII endonuclease.
<3>Ph.D. Thesis, Texas A and M Univ., College Station, TX, USA
<4>
<5>143
<6>2002
<7>Divalent metal ions can play an important role in the interactions beween nucleic acids and
proteins.  Here, PvuII endonuclease is developed as a model system to study DNA binding and
catalysis as a function of divalent metal ions.  A novel approach of equilibrium binding and
kinetics of binding and cleavage is used to understand the influence of divalent metal ions on
DNA recognition and catalysis.  In the presence of calcium, a divalent metal ion that does not
support catalysis, the  interaction between the specific recognition site and PvuII
endonuclease has a Kd of 53 +/- 10 pM (pH 7.5, 100 mM NaCl, 10 MM CaCl2).  A 6000 fold
reduction in the dissociation constant is seen when metal ions are absent.  Specific DNA
binding interactions exhibit an unusual shallow pH dependence.  Most protein-DNA interactions
have a more pronounced pH effect.  Nonspecific DNA binding is independent of divalent metal
ions; exhibiting a Kd near 200 nM (pH 7.5, 100 mM NaCl). Kinetic methods were developed to
understand how many metal ions are involved with both DNA binding and catalysis.  This work
presents the first comprehensive kinetic study of a restriction enzyme to show that more than
one metal ion influences both the DNA cleavage and association rate constants.  Using Ca(II)
to prevent turnover, the enzyme-DNA association rate constant is metal ion concentration
dependent, exhibiting a 100 fold increase from metal-free experiments to those done in the
presence of 10 mM Ca(II).  The association rate constant exhibited cooperative binding of at
least four metal ions per PvuII endonuclease dimer.  The dissociation rate constant showed a
shallow Ca(II) concentration dependence.  This provides new evidence that the metal ion
influence on the DNA dissociation constant is due to alterations in the association rate
constant.  Hill analysis of the cleavage rate constant as a function of Mg(II) concentration,
indicates that at least three metal ions per endonuclease dimer are involved in DNA
hydrolysis.  Combining the knowledge gained from equilibrium and kinetic studies provides
unique insights on how metal ions influence DNA binding and catalysis for restriction enzymes.

<>

<1>Conlan, S. et al.
<2>Single-molecule sequencing to track plasmid diversity of hospital-associated carbapenemase-producing Enterobacteriaceae.
<3>Sci. Transl. Med.
<4>6
<5>254ra126
<6>2014
<7>Public health officials have raised concerns that plasmid transfer between Enterobacteriaceae
species may spread resistance to carbapenems, an antibiotic
class of last resort, thereby rendering common health care-associated infections
nearly impossible to treat. To determine the diversity of carbapenemase-encoding
plasmids and assess their mobility among bacterial species, we performed
comprehensive surveillance and genomic sequencing of carbapenem-resistant
Enterobacteriaceae in the National Institutes of Health (NIH) Clinical Center
patient population and hospital environment. We isolated a repertoire of
carbapenemase-encoding Enterobacteriaceae, including multiple strains of
Klebsiella pneumoniae, Klebsiella oxytoca, Escherichia coli, Enterobacter
cloacae, Citrobacter freundii, and Pantoea species. Long-read genome sequencing
with full end-to-end assembly revealed that these organisms carry the carbapenem
resistance genes on a wide array of plasmids. K. pneumoniae and E. cloacae
isolated simultaneously from a single patient harbored two different
carbapenemase-encoding plasmids, indicating that plasmid transfer between
organisms was unlikely within this patient. We did, however, find evidence of
horizontal transfer of carbapenemase-encoding plasmids between K. pneumoniae, E.
cloacae, and C. freundii in the hospital environment. Our data, including full
plasmid identification, challenge assumptions about horizontal gene transfer
events within patients and identify possible connections between patients and the
hospital environment. In addition, we identified a new carbapenemase-encoding
plasmid of potentially high clinical impact carried by K. pneumoniae, E. coli, E.
cloacae, and Pantoea species, in unrelated patients and in the hospital
environment.

<>

<1>Conlan, S., Lawrence, C., McCue, L.A.
<2>Rhodopseudomonas palustris regulons detected by cross-species analysis of alphaproteobacterial genomes.
<3>Appl. Environ. Microbiol.
<4>71
<5>7442-7452
<6>2005
<7>Rhodopseudomonas palustris, an alpha-proteobacterium, carries out three of the chemical
reactions that support life on this planet: the conversion of sunlight to chemical-potential
energy; the absorption of carbon dioxide, which it converts to cellular material; and the
fixation of atmospheric nitrogen into ammonia. Insight into the transcription-regulatory
network that coordinates these processes is fundamental to understanding the biology of this
versatile bacterium. With this goal in mind, we predicted regulatory signals genomewide, using
a two-step phylogenetic-footprinting and clustering process that we had developed previously.
In the first step, 4,963 putative transcription factor binding sites, upstream of 2,044 genes
and operons, were identified using cross-species Gibbs sampling. Bayesian motif clustering was
then employed to group the cross-species motifs into regulons. We have identified 101 putative
regulons in R. palustris, including 8 that are of particular interest: a photosynthetic
regulon, a flagellar regulon, an organic hydroperoxide resistance regulon, the LexA regulon,
and four regulons related to nitrogen metabolism (FixK2, NnrR, NtrC, and sigma54). In some
cases, clustering allowed us to assign functions to proteins that previously had been
annotated with only putative functions; we have identified RPA0828 as the organic
hydroperoxide resistance regulator and RPA1026 as a cell cycle methylase. In addition to
predicting regulons, we identified a novel inverted repeat that likely forms a highly
conserved stem-loop and that occurs downstream of over 100 genes.

<>

<1>Connaughton, J.F., Kaloss, W.D., Vanek, P.G., Nardone, G.A., Chirikjian, J.G.
<2>The complete sequence of the Bacillus amyloliquefaciens proviral H2, BamHI methylase gene.
<3>Nucleic Acids Res.
<4>18
<5>4002
<6>1990
<7>None

<>

<1>Connaughton, J.F., Vanek, P.G., Chirikjian, J.G.
<2>Cloning of the BamHI methyl transferase gene from Bacillus amyloliquefaciens.
<3>J. Cell Biol.
<4>107
<5>535a
<6>1988
<7>We wish to report the initial characterization of a recombinant clone containing the BamHI
methylase gene.  Genomic chromosomal DNA purified from Bacillus amyloliquefaciens was
partially cleaved with HindIII, fractionated by size, and cloned into pSP64.  Plasmid DNA from
this library was challenged with BamHI endonuclease and transformed into E. coli HB101.  A
recombinant plasmid pBamM6.5 and a subclone pBamM2.5 were shown to contain the BamHI methylase
gene based on three independent observations.  Both plasmids were found to be resistent to
BamHI endonuclease cleavage, and chromosomal DNA isolated from E. coli HB101 cells harboring
either of the plasmids pBamM6.5 or pBamM2.5 were resistant to cleavage by BamHI endonuclease.
In addition, DNA isolated from lambda phage passaged through E. coli HB101 containing either
plasmid were also resistant to BamHI cleavage.  Expression of the BamHI methylase gene is
dependent on orientation in pSP64.  In these clones preliminary evidence indicates that
methylase gene expression may be under the direction of the plasmid encoded LacZ promoter.

<>

<1>Connaughton, J.F., Vanek, P.G., Lee-Lin, S.-Q., Chirikjian, J.G.
<2>Cloning of the BamHI methyltransferase gene from Bacillus amyloliquefaciens.
<3>Gene Anal. Tech.
<4>5
<5>116-124
<6>1988
<7>We wish to report the initial characterization of a recombinant clone
containing the BamHI methylase gene.  Genomic chromosomal DNA purified from
Bacillus amyloliquefaciens was partially cleaved with HindIII, fractionated by
size, and cloned into pSP64.  Plasmid DNA from this library was challenged with
BamHI endonuclease and transformed into Escherichia coli HB101.  A recombinant
plasmid pBamM6.5 and a subclone pBamM2.5 were shown to contain the BamHI
methylase gene based on three independent observations.  Both plasmids were
found to be resistant to BamHI endonuclease cleavage, and chromosomal DNA
isolated from E. coli HB101 cells harboring either of the plasmids pBamM6.5 or
pBamM2.5 was resistant to cleavage by BamHI endonuclease.  In addition, DNA
isolated from lambda phage passaged through E. coli HB101 containing either
plasmid was also resistant to BamHI cleavage.  Expression of the BamHI
methylase gene is dependent on orientation in pSP64.  In these clones
preliminary evidence indicates that methylase gene expresion may be under the
direction of the plasmid encoded LacZ promoter.

<>

<1>Connolly, B.A.
<2>Assay of restriction endonucleases using oligonucleotides.
<3>Methods Mol. Biol.
<4>30
<5>371-383
<6>1994
<7>Type II restriction endonucleases cleave double-stranded DNA at sequence-specific sites
typically 4-6 bp in length. Although large DNA molecules (viral DNA, plasmid DNA, and
chromosomal DNA) are the physiological substrates for these enzymes, activity is often shown
with small synthetic oligodeoxynucleotides providing that the recognition sequence is present.
The use of oligodeoxynucleotide substrates often allows information to be obtained concerning
the mechanism by which the particular endonuclease recognizes its cognate site so specifically
and discriminates accurately against all other sequences. Excellent examples include a very
thorough study with the EcoRI endonuclease that revealed the energetic bases of its
specificity and experiments with the EcoRV endonuclease using oligodeoxynucleotides containing
modified bases that demonstrated several direct contacts between enzyme and substrate. Central
to all these experiments are methods for the assay of the activity a particular restriction
endonuclease shows toward an oligonucleotide substrate.

<>

<1>Connolly, B.A., Eckstein, F., Pingoud, A.
<2>The Stereochemical Course of the Restriction Endonuclease EcoRI-catalyzed reaction.
<3>J. Biol. Chem.
<4>259
<5>10760-10763
<6>1984
<7>The restriction endonuclease EcoRI hydrolyzes the Rp diastereomer of
d(pGGsAATTCC), an analogue of d(pGGAATTCC) containing a chiral phosphorothioate
group at the cleavage site between the deoxyguanosine and the deoxyadenosine
residues (Connolly, B.A., Potter, B.V.L., Eckstein, F., Pingoud, A. and
Grotjahn, L. (1984) Biochemistry 23: 3343-3453).  Performing the reaction in
H2[18]O leads to d(pGG) and the hexanucleotide d([[18]O, S]pAATTCC) which has
an [18]O-containing phosphorothioate group at the 5' terminus.  Further
hydrolysis of this hexamer with nuclease P1 yields deoxyadenosine
5'-O-[18]O)phosphorothioate which can be stereospecifically phosphorylated with
adenylate kinase and pyruvate kinase to give Sp-[18]O deoxyadenosine
5'-O-(1-thiotriphosphate).  31P NMR spectroscopy shows the oxygen-18 in this
compound to be in a bridging position between the alpha- and beta-phosphorus
atoms.  Thus, the hydrolysis reaction catalyzed by EcoRI proceeds with
inversion of configuration at phosphorus.  This result is compatible with a
direct enzyme-catalyzed nucleophilic attack of H2O at phosphorus without
involvement of a covalent enzyme intermediate.

<>

<1>Connolly, B.A., Potter, B.V.L., Eckstein, F., Pingoud, A., Grotjahn, L.
<2>Synthesis and characterization of an octanucleotide containing the EcoRI recognition sequence with a phosphorothioate group at the cleavage site.
<3>Biochemistry
<4>23
<5>3443-3453
<6>1984
<7>The synthesis and characterization of an octanucleotide, d(GGsAATTCC),
containing the recognition sequence of the EcoRI restriction endonuclease with
a phosphorothioate internucleotidic linkage at the cleavage site are described.
Two approaches for the synthesis of the Rp and Sp diastereomers of this
octamer by the phosphite method are presented.  The first consists of the
addition of the sulfur instead of H2O to the phosphite at the appropriate
position during chain elongation.  This method results in a mixture of
diastereomers that can be separated by high-performance liquid chromatography
after 5'-terminal phosphorylation.  The second uses the presynthesized and
diastereomerically pure dinucleoside phosphorothioate d[Gp(S)A] for the
addition to the growing oligonucleotide chain as a block.  The products are
characterized by digestion with nuclease P1, fast atom bombardment mass
spectrometry, 31P NMR spectroscopy, and conversion to d(GGAATTCC) by
desulfurization with iodine.  Only the Rp diastereomers of d(GGsAATTCC) and its
5'-phosphorylated derivative are cleaved by EcoRI endonuclease.  The rate of
hydrolysis is slower than that of the unmodified octamer.  The phosphorothioate
octamer will be useful for the determination of the stereochemical course of
the EcoRI-catalyzed reaction.

<>

<1>Conrad, M., Topal, M.D.
<2>DNA and spermidine provide a switch mechanism to regulate the activity of restriction enzyme NaeI.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>86
<5>9707-9711
<6>1989
<7>Sequence-specific DNA-protein interactions are basic to DNA function.  To
better understand these interactions, we studied the effect of position on
cleavage of DNA by the type II restriction enzyme (EC 3.1.21.4) NaeI.  We
discovered two classes of NaeI restriction sites: sites susceptible and sites
resistant to cleavage.  Kinetic analysis showed that NaeI was activated by DNA
containing cleavable NaeI sites to rapidly cleave resistant NaeI sites by a
noncompetitive mechanism with a Km for substrate DNA of about 2 nM and a KA for
activating DNA of about 6 nM; activation increased catalysis but not substrate
binding.  Deletion mutagenesis in vitro showed that sequences flanking the NaeI
recognition site were responsible for the differences between activating and
nonactivating NaeI sites.  The polyamine spermidine had a dramatic effect on
the interaction of NaeI with DNA; in the presence of 1 mM spermidine, resistant
sites were cleaved rapidly and cleavable DNA inhibited cleavage.  The direct
regulation of enzymatic activity by DNA sequences in trans, and the modulation
of this regulation by a polyamine that is sensitive to the cell cycle, provides
a regulatory switch mechanism.  The implications of this switch for biological
control functions are discussed.

<>

<1>Conrad, M., Topal, M.D.
<2>Modified DNA fragments activate NaeI cleavage of refractory DNA sites.
<3>Nucleic Acids Res.
<4>20
<5>5127-5130
<6>1992
<7>Endonuclease NaeI cleaves DNA using a two-site mechanism. The DNA-binding sites are
nonidentical: they recognize different families of flanking sequences. A unique NaeI site that
is resistant to cleavage resides in M13 double-stranded DNA. NaeI can be activated to cleave
this site by small DNA fragments containing one or more NaeI sites. These activators are not
practical for genetic engineering because unphosphorylated activators that are consumed during
the cleavage of substrate give ends that may interfere with subsequent ligations. We show that
a DNA fragment containing phosphorothioate linkages at the NaeI scissile bonds (S-activator)
is not cleaved by NaeI, even though this S-activator binds to the substrate site. The
S-activator activates NaeI to cleave M13 DNA under conditions that completely exhaust
unsubstituted activator. These results demonstrate that activation is not coupled to cleavage
of activator, that NaeI reverts to its inactive state soon after dissociation of the EA
complex, and that S-activator makes for a nondepletable activator during prolonged
incubations.

<>

<1>Cook, L.J., Cox, J.P.L.
<2>Methylated DNA labels for marking objects.
<3>Biotechnol. Lett.
<4>25
<5>89-94
<6>2003
<7>We recently described a method for digitally labelling objects with DNA. Here we show that,
using DNA methyltransferases to create polymorphic DNA
templates, it is possible to significantly increase the number of labels
that can be generated by this method. Nine double-stranded DNA templates
of different length were methylated with either M.HaeIII or M.AluI
methyltransferase, or both. Different mixtures of methylated and
unmethylated versions of this template set were used to 'invisibly' label
paper. The mixtures were eluted from the paper and the methylated status
of the templates in each mixture successfully determined, and the labels
read, by digestion with the complementary restriction endonuclease,
followed by a polymerase chain reaction and agarose gel electrophoresis.
One methylated DNA label was read after it had been left on paper for two
months.

<>

<1>Cook, S.N., Jack, W.E., Xiong, X., Danley, L.E., Ellman, J.A., Schultz, P.G., Noren, C.J.
<2>Photochemically initiated protein splicing.
<3>Angew. Chem. Int. Ed. Engl.
<4>34
<5>1629-1630
<6>1995
<7>Protein splicing is a post-translational rearrangement process in which an
internal polypeptide segment (intein) is excised from a primary translation product, with
concomitant ligation of the flanking polypeptides (exteins).  This process results in the
production of two protein products from a single precursor polypeptide: the intein (often a
homing endonuclease thought to initiate lateral transmission of the intein coding sequence)
and the ligated exteins.

<>

<1>Coolen, J.P., Sjodin, A., Maraha, B., Hajer, G.F., Forsman, M., Verspui, E., Frenay, H.M., Notermans, D.W., de Vries, M.C., Reubsaet, F.A., Paauw, A., Roeselers, G.
<2>Draft genome sequence of Francisella tularensis subsp. holarctica BD11-00177.
<3>Standards in Genomic Sciences
<4>8
<5>539-547
<6>2013
<7>Francisella tularensis is a facultative intracellular bacterium in the class
Gammaproteobacteria. This strain is of interest because it is the etiologic agent
of tularemia and a highly virulent category A biothreat agent. Here we describe
the draft genome sequence and annotation of Francisella tularensis subsp.
holarctica BD11-00177, isolated from the first case of indigenous tularemia
detected in The Netherlands since 1953. Whole genome DNA sequence analysis
assigned this isolate to the genomic group B.FTNF002-00, which previously has
been exclusively reported from Spain, France, Italy, Switzerland and Germany.
Automatic annotation of the 1,813,372 bp draft genome revealed 2,103
protein-coding and 46 RNA genes.

<>

<1>Cooney, C.A.
<2>The restriction enzyme EheI (GGC/GCC) is sensitive to CpG methylation.
<3>Nucleic Acids Res.
<4>18
<5>3667
<6>1990
<7>None

<>

<1>Cooney, C.A., Eykholt, R.L., Bradbury, E.M.
<2>Methylation is co-ordinated on the putative replication origins of Physarum ribosomal DNA.
<3>J. Mol. Biol.
<4>204
<5>889-901
<6>1988
<7>In Physarum polycephalum, the ribosomal DNA is found as 60,000 base-pair palindromes. Each
rDNA has four symmetrically arranged replication origins flanked by ribosomal RNA genes. A
particular sequence, the putative replication origin, is repeated at the approximate position
of each origin and nowhere else in the molecule. On a typical rDNA molecule, only one origin
is active per replication cycle. We show that both the level and co-ordination of methylation
result in asymmetrically methylated rDNA molecules that are particularly hypomethylated at one
of their four putative replication origins. This pattern of methylation on a typical rDNA
molecule is consistent with a model where hypomethylation is a determinant of origin activity.

<>

<1>Cooper, A.A., Chen, Y.-J., Lindorfer, M.A., Stevens, T.H.
<2>Protein splicing of the yeast TFP1 intervening protein sequence: a model for self-excision.
<3>EMBO J.
<4>12
<5>2575-2583
<6>1993
<7>Protein splicing is the protein analogue of RNA splicing in which the
central portion (spacer) of a protein precursor is excised and the amino- and carboxy-
terminal portions of the precursor reconnected.  The yeast Tfp1 protein undergoes a rapid
protein splicing reaction to yield a spliced 69 kDa polypeptide and an excised 50 kDa spacer
protein.  We have demonstrated that the 69 kDa species arises by reformation of a bona fide
peptide bond.  Deletion analyses indicate that only sequences in the central spacer protein of
the Tfp1 precursor are critical for the protein splicing reaction.  A fusion protein in which
only the Tfp1 spacer domain was inserted into an unrelated protein also underwent efficient
splicing, demonstrating that all of the information required for protein splicing resides
within the spacer domain.  Alteration of Tfp1p splice junction residues blocked or
kinetically impaired protein splicing.  A protein splicing model is presented in which
asparagine rearrangement initiates the self-excision of the spacer protein from the Tfp1
precursor.  The Tfp1 spacer protein belongs to a new class of intervening sequences that
are excised at the protein rather than the RNA level.

<>

<1>Cooper, A.A., Stevens, T.H.
<2>Protein splicing: self-splicing of genetically mobile elements at the protein level.
<3>Trends Biochem. Sci.
<4>20
<5>351-356
<6>1995
<7>Protein splicing is a newly discovered process that is the protein equivalent
of RNA splicing.  Protein splicing proceeds through a branched protein intermediate, and
in vitro studies indicate that the reaction is autocatalytic.  The excised 'intein' proteins
are
site-specific DNA endonucleases that catalyze genetic mobility of their DNA coding
sequence by an 'intein homing' mechanism.

<>

<1>Cooper, A.A., Stevens, T.H.
<2>Protein splicing: Excision of intervening sequences at the protein level.
<3>Bioessays
<4>15
<5>667-674
<6>1993
<7>Protein splicing is an extraordinary post-translational reaction that removes
an intact central "spacer" domain (Sp) from precursor proteins (N-Sp-C) while splicing
together the N- and C-domains of the precursor, via a peptide bond, to produce a new
protein (N-C).  All of the available data on protein splicing fit a model in which these
intervening sequences excise at the protein level via a self-splicing mechanism.  Several
proteins have recently been discovered that undergo protein splicing, and in two such
cases, the excised spacer protein is an endonuclease.  Such endonucleases are capable of
conferring genetic mobility upon the intervening sequences that encodes them.  These
intervening sequences define a new family of mobile genetic elements that are translated yet
remain phenotypically silent by excising at the protein rather than the RNA level.

<>

<1>Cooper, J.E., Feil, E.J.
<2>The phylogeny of Staphylococcus aureus - which genes make the best intra-species markers?
<3>Microbiology
<4>152
<5>1297-1305
<6>2006
<7>The ability to make informed decisions on the suitability of alternative marker loci is
central for population and epidemiological investigations.
This issue was addressed using Staphylococcus aureus as a model population
by generating nucleotide sequence data from 33 gene fragments in a
representative sample of 30 strains. Supplementing the data with
pre-existing multilocus sequence typing data, an intra-species tree based
on approximately 17.8 kb of sequence was reconstructed and the goodness of
fit of each individual gene tree was computed. No strong association was
noted between gene function per se and phylogenetic reliability, but it is
suggested that candidate loci should possess at least the average degree
of nucleotide diversity for all genes in the genome. In the case of S.
aureus this threshold is >1 % mean pairwise diversity.

<>

<1>Cooper, K.K., Cooper, M.A., Zuccolo, A., Law, B., Joens, L.A.
<2>Complete Genome Sequence of Campylobacter jejuni Strain S3.
<3>J. Bacteriol.
<4>193
<5>1491-1492
<6>2011
<7>Campylobacter jejuni is one of the leading causes of bacterial gastroenteritis in the world;
however, there is only one complete genome
sequence of a poultry strain to date. Here we report the complete genome
sequence and annotation of the second poultry strain, C. jejuni strain S3.
This strain has been shown to be nonmotile, to be a poor invader in vitro,
and to be a poor colonizer of poultry after minimal in vitro passage.

<>

<1>Cooper, K.K., Mandrell, R.E., Louie, J.W., Korlach, J., Clark, T.A., Parker, C.T., Huynh, S., Chain, P.S., Ahmed, S., Carter, M.Q.
<2>Complete Genome Sequences of Two Escherichia coli O145:H28 Outbreak Strains of Food Origin.
<3>Genome Announcements
<4>2
<5>e00482-14
<6>2014
<7>Escherichia coli O145:H28 strain RM12581 was isolated from bagged romaine lettuce during a
2010 U.S. lettuce-associated outbreak. E. coli O145:H28 strain RM12761
was isolated from ice cream during a 2007 ice cream-associated outbreak in
Belgium. Here we report the complete genome sequences and annotation of both
strains.

<>

<1>Cooper, L.P., Dryden, D.T.F.
<2>The domains of a type I DNA methyltransferase: interactions and role in recognition of DNA methylation.
<3>J. Mol. Biol.
<4>236
<5>1011-1021
<6>1994
<7>The DNA methyltransferases of type I restriction-modification systems are trimeric enzymes
composed of one DNA specificity (S) subunit and two modification (M) subunits. The S subunit
contains two large regions, each of which recognizes one part of the split, asymmetrical DNA
target sequence. Each M subunit contains an amino acid motif for binding the methyl group
donor and cofactor, S-adenosyl methionine. The EcoKI methyltransferase has a strong preference
for methylating a hemimethylated DNA target rather than an unmodified target. We have used
partial proteolytic digestion of EcoKI methyltransferase to generate polypeptide domains that
we have identified by amino acid sequencing. The S subunit was cut into two large, folded
domains each containing one DNA binding region. Binding of DNA partially protected the S
subunit from digestion. The M subunit was also cut into two large domains joined together by a
short flexible loop, and a C-terminal tail region. The short loop contained part of the
S-adenosyl methionine binding motif, and cofactor binding protected the loop and the two large
domains from proteolysis. The C-terminal domain of M remained associated with the N-terminal
domain of the S subunit even after the rest of the protein had been digested. The conformation
of the tail region of the M subunit was sensitive to the methylation state of DNA in ternary
complexes also containing S-adenosyl methionine, and could differentiate between unmethylated
and hemimethylated DNA substrates.

<>

<1>Cooper, T.F., Heinemann, J.A.
<2>Postsegregational killing does not increase plasmid stability but acts to mediate the exclusion of competing plasmids.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>12643-12648
<6>2000
<7>Postsegregational killing (PSK) systems consist of a tightly linked toxin-antitoxin pair.
Antitoxin must be continually produced to prevent
the longer lived toxin from killing the cell. PSK systems on plasmids
are widely believed to benefit the plasmid by ensuring its stable
vertical inheritance. However, experimental tests of this "stability"
hypothesis were not consistent with its predictions. We suggest an
alternative hypothesis to explain the evolution of PSK: that PSK
systems have been selected through benefiting host plasmids in
environments where plasmids must compete during horizontal
reproduction. In this "competition" hypothesis, success of PSK
systems is a consequence of plasmid-plasmid competition, rather than
from an adaptive plasmid-host relationship. In support of this
hypothesis, a plasmid-encoded parDE PSK system mediated the exclusion
of an isogenic Delta parDE plasmid. An understanding of how PSK systems
influence plasmid success may provide insight into the evolution of
other determinants (e.g., antibiotic resistance and virulence) also
rendering a cell potentially dependent on an otherwise dispensable
plasmid.

<>

<1>Copeland, A. et al.
<2>Complete genome sequence of the halophilic and highly halotolerant Chromohalobacter salexigens type strain (1H11T).
<3>Standards in Genomic Sciences
<4>5
<5>379-388
<6>2011
<7>Chromohalobacter salexigens is one of nine currently known species of the genus
Chromoha-lobacter in the family Halomonadaceae. It is the most halotolerant of the so-called
'mod-erately halophilic bacteria' currently known and, due to its strong euryhaline
phenotype, it is an established model organism for prokaryotic osmoadaptation. C. salexigens
strain 1H11T and Halomonas elongata are the first and the second members of the family
Halomonada-ceae with a completely sequenced genome. The 3,696,649 bp long chromosome with a
total of 3,319 protein-coding and 93 RNA genes was sequenced as part of the DOE Joint Genome
Institute Program DOEM 2004.

<>

<1>Copeland, A. et al.
<2>Complete genome sequence of the aerobic, heterotrophy Marinithermus hydrothermalis type strain (T1T) from a deep-sea hydrothermal vent chimney.
<3>Standards in Genomic Sciences
<4>6
<5>21-30
<6>2012
<7>Marinithermus hydrothermalis Sako et al. 2003 is the type species of the monotypic genus
Marinithermus. M. hydrothermalis T1T was the first isolate within the phylum
"Thermus-Deinococcus" to exhibit optimal growth under a salinity equivalent to that of sea
water and to have an absolute requirement for NaCl for growth. M. hydrothermalis T1T is of
interest because it may provide a new insight into the ecological significance of the aerobic,
thermophilic decomposers in the circulation of organic compounds in deep-sea hydrothermal vent
ecosystems. This is the first completed genome sequence of a member of the genus Marinithermus
and the seventh sequence from the family Thermaceae. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The 2,269,167 bp long
genome with its 2,251 protein-coding and 59 RNA genes is a part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Copeland, A. et al.
<2>Complete genome sequence of Desulfomicrobium baculatum type strain (X).
<3>Standards in Genomic Sciences
<4>1
<5>29-37
<6>2009
<7>Desulfomicrobium baculatum is the type species of the genus Desulfomicrobium, which is the
type genus of the family Desulfomicrobiaceae. It is of phylogenetic
interest because of the isolated location of the family Desulfomicrobiaceae
within the order Desulfovibrionales. D. baculatum strain X(T) is a Gram-negative,
motile, sulfate-reducing bacterium isolated from water-saturated manganese
carbonate ore. It is strictly anaerobic and does not require NaCl for growth,
although NaCl concentrations up to 6% (w/v) are tolerated. The metabolism is
respiratory or fermentative. In the presence of sulfate, pyruvate and lactate are
incompletely oxidized to acetate and CO(2). Here we describe the features of this
organism, together with the complete genome sequence and annotation. This is the
first completed genome sequence of a member of the deltaproteobacterial family
Desulfomicrobiaceae, and this 3,942,657 bp long single replicon genome with its
3494 protein-coding and 72 RNA genes is part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Copeland, A. et al.
<2>Complete genome sequence of the aquatic bacterium Runella slithyformis type strain (LSU 4(T)).
<3>Standards in Genomic Sciences
<4>6
<5>145-154
<6>2012
<7>Runella slithyformis Larkin and Williams 1978 is the type species of the genus Runella, which
belongs to the Cytophagaceae, a family that was only recently
classified to the order Cytophagales in the class Cytophagia. The species is of
interest because it is able to grow at temperatures as low as 4 degrees C. This
is the first completed genome sequence of a member of the genus Runella and the
sixth sequence from the family Cytophagaceae. The 6,919,729 bp long genome
consists of a 6.6 Mbp circular genome and five circular plasmids of 38.8 to 107.0
kbp length, harboring a total of 5,974 protein-coding and 51 RNA genes and is a
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Copeland, A. et al.
<2>Complete genome sequence of the orange-red pigmented, radioresistant Deinococcus  proteolyticus type strain (MRP(T)).
<3>Standards in Genomic Sciences
<4>6
<5>240-250
<6>2012
<7>Deinococcus proteolyticus (ex Kobatake et al. 1973) Brook and Murray 1981 is one  of currently
47 species in the genus Deinococcus within the family
Deinococcaceae. Strain MRP(T) was isolated from feces of Lama glama and possesses
extreme radiation resistance, a trait is shares with various other species of the
genus Deinococcus, with D. proteolyticus being resistant up to 1.5 Mrad of gamma
radiation. Strain MRP(T) is of further interest for its carotenoid pigment. The
genome presented here is only the fifth completed genome sequence of a member of
the genus Deinococcus (and the forth type strain) to be published, and will
hopefully contribute to a better understanding of how members of this genus
adapted to high gamma- or UV ionizing-radiation. Here we describe the features of
this organism, together with the complete genome sequence and annotation. The
2,886,836 bp long genome with its four large plasmids of lengths 97 kbp, 132 kbp,
196 kbp and 315 kbp harbors 2,741 protein-coding and 58 RNA genes and is a part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Copeland, A. et al.
<2>Complete genome sequence of Atopobium parvulum type strain (IPP 1246).
<3>Standards in Genomic Sciences
<4>1
<5>166-173
<6>2009
<7>Atopobium parvulum (Weinberg et al. 1937) Collins and Wallbanks 1993 comb. nov. is the type
strain of the species and belongs to the genomically yet unstudied
Atopobium/Olsenella branch of the family Coriobacteriaceae. The species A.
parvulum is of interest because its members are frequently isolated from the
human oral cavity and are found to be associated with halitosis (oral malodor)
but not with periodontitis. Here we describe the features of this organism,
together with the complete genome sequence, and annotation. This is the first
complete genome sequence of the genus Atopobium, and the 1,543,805 bp long single
replicon genome with its 1369 protein-coding and 49 RNA genes is part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Copeland, A. et al.
<2>Complete genome sequence of Catenulispora acidiphila type strain (ID 139908).
<3>Standards in Genomic Sciences
<4>1
<5>119-125
<6>2009
<7>Catenulispora acidiphila Busti et al. 2006 is the type species of the genus Catenulispora, and
is of interest because of the rather isolated phylogenetic
location it occupies within the scarcely explored suborder Catenulisporineae of
the order Actinomycetales. C. acidiphilia is known for its acidophilic, aerobic
lifestyle, but can also grow scantly under anaerobic conditions. Under regular
conditions, C. acidiphilia grows in long filaments of relatively short aerial
hyphae with marked septation. It is a free living, non motile, Gram-positive
bacterium isolated from a forest soil sample taken from a wooded area in
Gerenzano, Italy. Here we describe the features of this organism, together with
the complete genome sequence and annotation. This is the first complete genome
sequence of the actinobacterial family Catenulisporaceae, and the 10,467,782 bp
long single replicon genome with its 9056 protein-coding and 69 RNA genes is a
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Copeland, J.C., Bryson, V.
<2>Restriction in matings of Escherichia coli strain K-12 with strain B.
<3>Genetics
<4>54
<5>441-452
<6>1966
<7>Restriction in Escherichia coli limits the acceptance of genetic elements such
as bacteriophage, sex-factor, and donated bacterial chromosomes.  As reported
in this paper, restriction of K-12 genetic material by strain B is greatest
early in the mating and later becomes constant.  Unlike restriction in B/r, no
delay in the appearance of a proximal marker is found.  Also, the linkage
between markers is not reduced by a constant amount, but is dependent upon the
distance between the markers.  Restriction in strain B is responsible for only
part of the reduction in the inheritance of genetic markers from K-12, whereas
in strain B/r restriction accounts for the entire effect.

<>

<1>Corina, L.E.
<2>Homing endonuclease I-CreII: A novel dual-motif enzyme that catalyzes group I intron homing.
<3>Ph.D. Thesis, Southwestern Medical Center, Univ. of Texas, USA
<4>
<5>1-148
<6>2005
<7>I-CreII is a homing endonuclease from the Cr.psbA intron of Chlamydomonas reinhardtii.  It
cleaves the exon 4-exon 5 junction of psbA DNA, thereby inducing a recombination event
referred to as intron homing.  Homing endonucleases are classified into four main groups based
on the presence of a catalytic domain unique to each group.  I-CreII is novel in that it
appears to contain two catalytic domains, an H-N-H and a GIY-YIG.  The role of the puative
GIY-YIG motif has been questioned, however, because I-CmoeI, a related protein, behaves like
an H-N-H endonuclease.  Recently, Kim et al. (2005) showed that DNA cleavage by I-CreII leaves
2-nt 3' OH overhands, which is a feature unique to GIY-YIG enzymes.  Thus, in order to
resolve this question, I have performed new biochemical and kinetic analyses of wild-type and
mutant proteins that have had a conserved residue in one of the motifs substituted with
alanine.  The results indicate that not only does I-CreII have a functional GIY-YIG motif, but
the data point to it as the catalytic domain.  From this new perspective, this is also the
first quantitative study of a GIY-YIG endonuclease.  These data also provide the first direct
evidence that an H-N-H motif can function primarily in DNA binding rather than catalysis- a
finding that has implications for transcription factors from lower plants and protists that
appear to have this motif.

<>

<1>Corina, L.E., Qiu, W., Desai, A., Herrin, D.L.
<2>Biochemical and mutagenic analysis of I-CreII reveals distinct but important roles for both the H-N-H and GIY-YIG motifs.
<3>Nucleic Acids Res.
<4>37
<5>5810-5821
<6>2009
<7>Homing endonucleases typically contain one of four conserved catalytic motifs, and other
elements that confer tight DNA binding. I-CreII, which
catalyzes homing of the Cr.psbA4 intron, is unusual in containing two
potential catalytic motifs, H-N-H and GIY-YIG. Previously, we showed that
cleavage by I-CreII leaves ends (2-nt 3' overhangs) that are
characteristic of GIY-YIG endonucleases, yet it has a relaxed metal
requirement like H-N-H enzymes. Here we show that I-CreII can bind DNA
without an added metal ion, and that it binds as a monomer, akin to
GIY-YIG enzymes. Moreover, cleavage of supercoiled DNA, and estimates of
strand-specific cleavage rates, suggest that I-CreII uses a sequential
cleavage mechanism. Alanine substitution of a number of residues in the
GIY-YIG motif, however, did not block cleavage activity, although DNA
binding was substantially reduced in several variants. Substitution of
conserved histidines in the H-N-H motif resulted in variants that did not
promote DNA cleavage, but retained high-affinity DNA binding-thus
identifying it as the catalytic motif. Unlike the non-specific H-N-H
colicins, however; substitution of the conserved asparagine substantially
reduced DNA binding (though not the ability to promote cleavage). These
results indicate that, in I-CreII, two catalytic motifs have evolved to
play important roles in specific DNA binding. The data also indicate that
only the H-N-H motif has retained catalytic ability.

<>

<1>Cornish-Bowden, A.
<2>Nomenclature for incompletely specified bases in nucleic acid sequences: recommendations 1984.
<3>Nucleic Acids Res.
<4>13
<5>3021-3030
<6>1985
<7>With the introduction of methods of rapid nucleic acid sequence determination, synthesis of
mixed oligonucleotide probes and computer-assisted analysis of nucleic acid sequences, the use
of a single symbol to designate a variety of possible nucleotides at a single position has
become widespread over the last few years. Whereas the use of, for example, the symbols R and
Y to designate purine (A or G) and pyrimidine (C or T) ribonucleotides respectively is
generally accepted, no agreed symbols exist for the other possible combinations. Indeed, a
plethora of diverse systems have proliferated in the last few years. It is striking that, in
one extreme case, the combination (C or G) has been represented by at least five different
symbols. A standardized set of symbols is thus required to prevent confusion.

<>

<1>Corvaglia, A.R., Francois, P., Bertrand, X., Quentin, R., Hernandez, D., van der Mee-Marquet, N.
<2>Whole-Genome Sequences of Two Staphylococcus aureus ST398 Strains of Human Origin, S94 and S100.
<3>Genome Announcements
<4>1
<5>e00691-13
<6>2013
<7>Sequence type 398 (ST398) Staphylococcus aureus was originally associated with animal
infection. We announce the complete genome sequences of two ST398
methicillin-susceptible S. aureus strains of human origin, S94 and S100. The
genome sequences assist in the characterization of interesting ST398 features
related to host specificities.

<>

<1>Corvaglia, A.R., Francois, P., Hernandez, D., Perron, K., Linder, P., Schrenzel, J.
<2>A type III-like restriction endonuclease functions as a major barrier to horizontal gene transfer in clinical Staphylococcus aureus strains.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>11954-11958
<6>2010
<7>Staphylococcus aureus is an versatile pathogen that can cause life-threatening infections.
Depending on the clinical setting, up to 50% of S. aureus infections are caused by
methicillin-resistant strains (MRSA) that in most cases are resistant to many other
antibiotics, making treatment difficult. The emergence of community-acquired MRSA drastically
changed the picture by increasing the risk of MRSA infections. Horizontal transfer of genes
encoding for antibiotic resistance or virulence factors is a major concern of
multidrug-resistant S. aureus infections and epidemiology. We identified and characterized a
type III-like restriction system present in clinical S. aureus strains that prevents
transformation with DNA from other bacterial species. Interestingly, our analysis revealed
that some clinical MRSA strains are deficient in this restriction system, and thus are
hypersusceptible to the horizontal transfer of DNA from other species, such as Escherichia
coli, and could easily acquire a vancomycin-resistance gene from enterococci. Inactivation of
this restriction system dramatically increases the transformation efficiency of clinical S.
aureus strains, opening the field of molecular genetic manipulation of these strains using DNA
of exogenous origin.

<>

<1>Cosson, P., Koehler, T., Benghezal, M., Marchetti, A., Van Delden, C.
<2>Virulence genes, proteins, and their use.
<3>International Patent Office
<4>WO 2004057018 A
<5>
<6>2004
<7>A series of genes from Pseudomonas aeruginosa and Klebsiella are shown to encode products that
are implicated in virulence.  The identification of these genes therefore allows attenuated
microorganisms to be produced.  Furthermore, the genes or their encoded products can be used
to identify antimicrobial drugs, diagnostic methods for the identification of a
pathogen-associated disease, and in the manufacture of vaccines.

<>

<1>Cosstick, R., Li, X., Tuli, D.K., Williams, D.M., Connolly, B.A., Newman, P.C.
<2>Molecular recognition in the minor groove of the DNA helix. Studies on the synthesis of oligonucleotides and polynucleotides containing 3-deaza-2'-deoxyadenosine. Interaction of the oligonucleotides with the restriction endonuclease EcoRV.
<3>Nucleic Acids Res.
<4>18
<5>4771-4778
<6>1990
<7>An improved procedure for the preparation of 3-deaza-2'-deoxyadenosine (d3CA)
is described which is suitable for the synthesis of gram quantities of this
analogue.  Using phosphoramidite chemistry d3CA has been incorporated into the
EcoRV restriction endonuclease recognition sequence present in the
self-complementary dodecamer d(GACGATATCGTC).  The modified oligonucleotides
have been thoroughly characterised by nucleoside composition analysis, circular
dichroism and thermal melting studies.  Studies with EcoRV show that
incorporation of d3CA into either the central or outer dA-dT base-pair results
in a substantial reduction in the rate of cleavage.  The two-step conversion of
d3CA to 3-deaza-2'-deoxyadenosine-5'-O-triphosphate (d3CATP) via the
5'-O-tosylate is also described.  d3CATP is not a substrate in the
poly[d(AT)].poly[d(AT)] primed polymerisation for either E. coli DNA polymerase
I or Micrococcus luteus DNA polymerase.  In a more detailed kinetic analysis
d3CATP was shown to be a competitive inhibitor of E. coli DNA polymerase I with
respect to dATP.

<>

<1>Costa, M.O., Beltrame, C.O., Ferreira, F.A., Botelho, A.M., Lima, N.C., Souza, R.C., de Almeida, L.G., Vasconcelos, A.T., Nicolas, M.F., Figueiredo, A.M.
<2>Complete Genome Sequence of a Variant of the Methicillin-Resistant Staphylococcus aureus ST239 Lineage, Strain BMB9393, Displaying Superior Ability To Accumulate  ica-Independent Biofilm.
<3>Genome Announcements
<4>1
<5>e00576-13
<6>2013
<7>Biofilm is considered an important virulence factor in nosocomial infections. Herein, we
report the complete genome sequence of a variant of
methicillin-resistant Staphylococcus aureus, strain BMB9393, which is highly
disseminated in Brazil. This strain belongs to the lineage ST239 and displays
increased ability to accumulate ica-independent biofilm and to invade human
epithelial cells.

<>

<1>Costa, N.D., Thacker, J.
<2>The Efficiency of Restriction Endonuclease Digests Determined by PCR.
<3>Biotechniques
<4>13
<5>190
<6>1992
<7>Amplification of DNA by PCR should be stopped by a double-strand break in the template
molecule. However, in the course of experiments designed to utilize restriction endonucleases
(RE) to eliminate PCR amplification, we observed that extensive digestion rarely resulted in
lack of a PCR product. We excluded one possible explanation for this obervation: "jumping" PCR
where a 5' overhang will give rise to first-round products with short end homologies that may
hybridize. Polymerization on this substrate would result in a complete template molecule.
However, this will not occur for a 3' overhang or blunt end, yet we obtained similar
amplification with DNA cut to give any type of end.

<>

<1>Cote, V., Mercier, J.P., Lemieux, C., Turmel, M.
<2>The single group-I intron in the chloroplast rrnL gene of Chlamydomonas humicola encodes a site-specific DNA endonuclease (I-ChuI).
<3>Gene
<4>129
<5>69-76
<6>1993
<7>The single group-I intron (ChLSU-1) in the chloroplast (cp) large subunit rRNA-encoding gene
(rrnL) of the green alga Chlamydomonas humicola is located at a position at which no introns
have previously been characterized in other systems. In the present study, the nucleotide (nt)
sequence of this 1118-bp intron was found to contain an internal open reading frame (ORF) that
potentially encodes a basic protein of 218 amino acid residues. The putative C. humicola
protein features two copies of the LAGLI-DADG motif and is part of the family of
intron-encoded proteins comprising the endonucleases (ENases). I-SceI, I-SceI, I-SceIV and
I-CsmI. Expression of the ChLSU.1 intron ORF in vitro in the presence of a 260-bp DNA fragment
containing the exon 1-2 junction of an intronless version of the C. humicol rrnL resulted in
specific cleavage of the DNA fragment very close to the intron insertion site. This novel
intron-encoded ENase, designated I-ChuI, was also shown to generate a staggered cut with 4-nt
(CTCG) 3'-OH overhangs 2 bp downstream from the intron insertion site.

<>

<1>Coucheron, D.H.
<2>Acetobacter strains contain DNA modified at GAATTC and GANTC.
<3>Can. J. Microbiol.
<4>43
<5>456-460
<6>1997
<7>Total DNAs from nine strains of Acetobacter xylinum, two strains of Acetobacter aceti, and one
Acetobacter pasteurianus strain were examined for the extent of digestion by various
restriction endonucleases.  The majority of the endonucleases cleaved the total DNAs with a
frequency expected from the number of sites present in DNA sequences deposited in the GenBank
database.  However, the restriction enzyme digestions identified two different genomic DNA
modifications in Acetobacter.  One sequence-specific modification protected total DNAs from
seven of the A. xylinum strains against cleavage by EcoRI (GAATTC).  Digestion of total DNAs
from A. xylinum ATCC 10245 (DNA not cut by EcoRI) and the closely related A. xylinum NRCC
17005 (DNA cut by EcoRI) with Tsp509I (AATT) revealed differences in restriction frequencies
adenine within GAATTC.  Another sequence-specific modification rendered total DNAs from all
the 12 strains recalcitrant to digestion by HinfI.  The latter modification indicated that
species of the genus Actobacter contain a solitary DNA methyltransferase that probably
methylates adenine in GANTC.

<>

<1>Coulby, J., Sternberg, N.
<2>Bacteriophage P1 encodes its own dam methylase.
<3>Plasmid
<4>17
<5>81
<6>1987
<7>The packaging of P1 DNA during its life cycle has been shown to be dependent upon the
methylation of the adenine residue in the sequence 5'-GATC-3' of the phage packaging site
(pac).  In vitro studies have shown that unmethylated P1 DNA is not cut at the pac site, and
the DNA is not packaged into the viral heads.  In an E. coli dam- host, which is unable to
methylate 5'-GATC-3', P1 phage production lags about 20 min behind that in a dam+ host. This
delayed production of phage is probably due to a need to synthesize a P1 dam analog before the
pac site can be cleaved.  To study this process we have begun to characterize the P1 dam gene.
A 13.5-kb region of P1 DNA was cloned into a lambda vector, and the hybrid was shown to encode
a dam methylase based on its ability to methylate pBR322 DNA upon infection of an E. coli dam-
host containing that plasmid.  The P1 dam gene borders the junction of P1 EcoRI fragments 3
and 8.  This region of the P1 genome is currently being subcloned in order to localize the
gene.  Following localization the gene will be mutagenized in an effort to further elucidate
the role of the dam methylase in the P1 life cycle.

<>

<1>Coulby, J.N., Sternberg, N.L.
<2>Characterization of the phage P1 dam gene.
<3>Gene
<4>74
<5>191
<6>1988
<7>Meeting Abstract

<>

<1>Coupland, P., Chandra, T., Quail, M., Reik, W., Swerdlow, H.
<2>Direct sequencing of small genomes on the Pacific Biosciences RS without library  preparation.
<3>Biotechniques
<4>53
<5>365-372
<6>2012
<7>We have developed a sequencing method on the Pacific Biosciences RS sequencer (the PacBio) for
small DNA molecules that avoids the need for a standard library
preparation. To date this approach has been applied toward sequencing
single-stranded and double-stranded viral genomes, bacterial plasmids, plasmid
vector models for DNA-modification analysis, and linear DNA fragments covering an
entire bacterial genome. Using direct sequencing it is possible to generate
sequence data from as little as 1 ng of DNA, offering a significant advantage
over current protocols which typically require 400-500 ng of sheared DNA for the
library preparation.

<>

<1>Courties, A., Riedel, T., Jarek, M., Intertaglia, L., Lebaron, P., Suzuki, M.T.
<2>Genome Sequence of Strain MOLA814, a Proteorhodopsin-Containing Representative of the Betaproteobacteria Common in the Ocean.
<3>Genome Announcements
<4>1
<5>e01062-13
<6>2013
<7>Strain MOLA814 is a marine betaproteobacterium that was isolated from seawater in the Beaufort
Sea. Here, we present its genome sequence and annotation. Genome
analysis revealed the presence of a proteorhodopsin-encoding sequence together
with its retinal-producing pathway, indicating that this strain might generate
energy by using light.

<>

<1>Cousin, S., Clermont, D., Creno, S., Ma, L., Loux, V., Bizet, C., Bouchier, C.
<2>Draft Genome Sequence of Lactobacillus pasteurii CRBIP 24.76T.
<3>Genome Announcements
<4>1
<5>e00660-13
<6>2013
<7>We report the draft genome sequence of the type strain Lactobacillus pasteurii CRBIP 24.76,
which is closely related to L. gigeriorum CRBIP 24.85(T), isolated
from a chicken crop. The total length of the 29 contigs is about 1.9 Mb, with a
G+C content of 40% and 1,946 coding sequences.

<>

<1>Cousin, S., Creno, S., Ma, L., Clermont, D., Loux, V., Bizet, C., Bouchier, C.
<2>Draft Genome Sequence of Lactobacillus hominis Strain CRBIP 24.179T, Isolated from Human Intestine.
<3>Genome Announcements
<4>1
<5>e00662-13
<6>2013
<7>We report the draft genome sequence of the strain Lactobacillus hominis CRBIP 24.179(T),
isolated from a human clinical sample. The total length of the 28
contigs is about 1.9 Mb, with a G+C content of 37% and 1,983 coding sequences.

<>

<1>Cousin, S., Loux, V., Ma, L., Creno, S., Clermont, D., Bizet, C., Bouchier, C.
<2>Draft Genome Sequences of Lactobacillus equicursoris CIP 110162T and Lactobacillus sp. Strain CRBIP 24.137, Isolated from Thoroughbred Racehorse Feces  and Human Urine, Respectively.
<3>Genome Announcements
<4>1
<5>e00663-13
<6>2013
<7>We report the draft genome sequences of strain Lactobacillus equicursoris CIP 110162(T),
isolated from racehorse breed feces, and Lactobacillus sp. strain
CRBIP 24.137, isolated from human urine; the two strains are closely related. The
total lengths of the 116 and 62 scaffolds are about 2.157 and 2.358 Mb, with G+C
contents of 46 and 45% and 2,279 and 2,342 coding sequences (CDSs), respectively.

<>

<1>Cousin, S., Ma, L., Creno, S., Clermont, D., Loux, V., Bizet, C., Bouchier, C.
<2>Draft Genome Sequence of Lactobacillus gigeriorum CRBIP 24.85T, Isolated from a Chicken Crop.
<3>J. Bacteriol.
<4>194
<5>5973
<6>2012
<7>We report the draft genome of the strain Lactobacillus gigeriorum CRBIP 24.85(T), isolated
from a chicken crop. The total length of the 60 scaffolds is about 1.9
Mb, with a GC content of 38% and 2,062 protein-coding sequences (CDS).

<>

<1>Cousineau, B., Lawrence, S., Smith, D., Belfort, M.
<2>Retrotransposition of a bacterial group II intron.
<3>Nature
<4>404
<5>1018-1021
<6>2000
<7>Self-splicing group II introns may be the evolutionary progenitors of eukaryotic spliceosomal
introns, but the route by which they invade new chromosomal sites is unknown. To address the
mechanism by which group II introns are disseminated, we have studied the bacterial L1.LtrB
intron from Lactococcus lactis. The protein product of this intron, LtrA, possesses maturase,
reverse transcriptase and endonuclease enzymatic activities. Together with the intron, LtrA
forms a ribonucleoprotein (RNP) complex which mediates a process known as retrohoming. In
retrohoming, the intron reverse splices into a cognate intronless DNA site. Integration of a
DNA copy of the intron is recombinase independent but requires all three activities of LtrA.
Here we report the first experimental demonstration of a group II intron invading ectopic
chromosomal sites, which occurs by a distinct retrotransposition mechanism. This
retrotransposition process is endonuclease-independent and recombinase-dependent, and is
likely to involve reverse splicing of the intron RNA into cellular RNA targets. These
retrotranspositions suggest a mechanism by which splicesomal introns may have become widely
dispersed.

<>

<1>Cousineau, B., Smith, D., Lawrence-Cavanagh, S., Mueller, J.E., Yang, J., Mills, D., Manias, D., Dunny, G., Lambowitz, A.M., Belfort, M.
<2>Retrohoming of a bacterial group II intron: Mobility via complete reverse splicing, independent of homologous DNA recombination.
<3>Cell
<4>94
<5>451-462
<6>1998
<7>The mobile group II intron of Lactococcus lactis, Ll.LtrB, provides the opportunity to analyze
the homing pathway in genetically tractable bacterial systems.  Here, we show that Ll.LtrB
mobility occurs by an RNA-based retrohoming mechanism in both Escherichia coli and L. lactis.
Surprisingly, retrohoming occurs efficiently in the absence of RecA function, with a relaxed
requirement for flanking exon homology and without coconversion of exon markers.  These
results lead to a model for bacterial retrohoming in which the intron integrates into
recipient DNA by complete reverse splicing and serves as the template for cDNA synthesis.  The
retrohoming reaction is completed in unprecedented fashion by a DNA repair event that is
independent of homologous recombination between the alleles.  Thus, Ll.LtrB has many features
of retrotransposons, with practical and evolutionary implications.

<>

<1>Coutinho, B.G., Passos-da-Silva, D., Previato, J.O., Mendonca-Previato, L., Venturi, V.
<2>Draft Genome Sequence of the Rice Endophyte Burkholderia kururiensis M130.
<3>Genome Announcements
<4>1
<5>e0022512
<6>2013
<7>Burkholderia kururiensis M130 is one of the few characterized rice endophytes and was isolated
from surface-sterilized rice roots. This bacterium shows strong growth-promoting effects,
being able to increase rice yields. Here we present its draft genome sequence, which contains
important traits for endophytic life and plant growth promotion.

<>

<1>Cowan, G.M.
<2>New family of Type I restriction and modification systems.
<3>Ph.D. Thesis, University of Edinburgh, Edinburgh, Scotland, , , Edinburgh
<4>
<5>1-128
<6>1988
<7>The chromosomally encoded type I restriction and modification system of Escherichia coli 15T,
EcoA, behaves physiologically as a type I system, and in genetic crosses the hsdA genes behave
as alleles of the hsd K genes of the archetypal type I system of E. coli K-12.  However,
molecular experiments failed to demonstrate relatedness between the A and K systems.  Genes
(hsdE) related to hsdA on the basis of DNA homology confer a new specificity to a natural
isolate, E. coli A58.  The genes encoding EcoA and EcoE have an organization which mimics that
of EcoK: there are three genes in the same order as those encoding EcoK, of which one, hsdS,
confers the specificity.  The polypeptides of EcoA, EcoE, and CfrA (a new A-like system) show
structural homology, demonstrated by immunological cross-reactivity.  The evidence presented
indicates that EcoA and EcoE represent an alternative family of type I restriction and
modification enzymes.  The recognition sequence of EcoE, 5'-GAG(N7)ATGC-3', is typical of
type I systems.  Furthermore, heteroduplex analyses and nucleotide sequencing have indicated
that the specificity genes of the A-like family show two large regions of variable sequence,
interspersed and flanked by conserved sequences which include a repeated sequence.  Such an
organization is similar to that of the K family specificity genes.  The hsdS genes of EcoA and
EcoE show extensive homology throughout the proximal regions: both recognize the trinucleotide
GAG.  In general no obvious similarity was detected between the primary sequences of the
specificity polypeptides of the two families, with one remarkable exception: the proximal
variable domain of the SB specificity polypeptide shows 44% identity to the proximal regions
of the specificity subunits of both EcoA and EcoE.  Significantly, the StySB enzyme also
recognizes the trinucleotide GAG.

<>

<1>Cowan, G.M.
<2>A new family of type I restriction and modification systems.
<3>Diss. Abstr.
<4>50
<5>441
<6>1989
<7>The chromosomally encoded type I restriction and modification system of Escherichia coli 15T-,
EcoA, behaves physiologically as a type I system, and in genetic crosses the hsd A genes
behave as alleles of the hsd K genes of the archetypal type I system of E. coli K-12. However,
molecular experiments failed to demonstrate relatedness between the A and K systems. Genes
(hsd E) related to hsd A on the basis of DNA homology confer a new specificity to a natural
isolate, E. coli A58. The genes encoding EcoA and EcoE have an organization which mimics that
of EcoK: there are three genes in the same order as those encoding EcoK, of which one, hsdS,
confers the specificity. The polypeptides of EcoA, EcoE, and CfrA (a new A-like system) show
structural homology, demonstrated by immunological cross-reactivity. The evidence presented
indicates that EcoA and EcoE represent an alternative family of type I restriction and
modification enzyme. The recognition sequence of EcoE, 5'-GAG(N7)ATGC-3', is typical of type
I systems. Furthermore, heteroduplex analyses and nucleotide sequencing have indicated that
the specificity genes of the A-like family show two large regions of variable sequence,
interspersed and flanked by conserved sequences which include a repeated sequence. Such an
organization is similar to that of the K family specificity genes. The hsdS genes of EcoA and
EcoE show extensive homology throughout the proximal regions: both recognize the trinucleotide
GAG. In general no obvious similarity was detected between the primary sequences of the
specificity polypeptides of the two families, with one remarkable exception the proximal
variable domain of the SB specificity polypeptide shows 44% identity to the proximal regions
of the specificity subunits of both EcoA and EcoE. Significantly, the StySB enzyme also
recognizes the trinucleotide GAG.

<>

<1>Cowan, G.M., Daniel, A.S., Gann, A.A.F., Kelleher, J.E., Murray, N.E.
<2>Defining domains in type-I restriction and modification enzymes.
<3>Gene
<4>74
<5>239-241
<6>1988
<7>Meeting Abstract

<>

<1>Cowan, G.M., Gann, A.A.F., Murray, N.E.
<2>Conservation of complex DNA recognition domains between families of restriction enzymes.
<3>Cell
<4>56
<5>103-109
<6>1989
<7>One polypeptide, designated S, confers sequence-specificity to the multisubunit
type I restriction enzymes.  Two families of such enzymes, K and A, include
members that recognize diverse, bipartite, target sequences.  The S
polypeptides of the K family, while having areas of near identity, also contain
two extensive regions of variable sequence.  We now show that one of these,
comprising the N-terminal 150 amino acids, specifies recognition of one
component of the bipartite target sequence.  We have determined the sequence
recognized by EcoE, a member of the A family.  This sequence, 5'GAG(N7)ATGC,
has the trinucleotide GAG in common with EcoA and with StySB of the K family.
We determined the nucleotide sequences of the S genes of EcoA and EcoE, and
compared their predicted amino acid sequences with each other and with those of
the five members of the K family.  There is no general sequence similarity
between families, but the domain of the S polypeptide of StySB, which specifies
GAG, shows nearly 50 per cent identity with the amino variable region of the S
polypeptides of EcoA and EcoE.  A complex domain that recognizes and directs
methylation of GAG is therefore common to enzymes of generally dissimilar amino
acid sequence.

<>

<1>Cowan, J.A.
<2>Role of metal ions in promoting DNA binding and cleavage by restriction endonucleases.
<3>Nucleic Acids Mol. Biol.
<4>14
<5>339-360
<6>2004
<7>While three major classes of restriction endonucleases have been identified (Types I, II, and
III), Type II are the most straightforward inasmuch as they require divalent magnesium as an
essential cofactor but have no need for ATP.  A complete classification of Type II restriction
nucleases has been presented elsewhere and the family is noted for the remarkable specificity
and simplicity of its function.  These enzymes cleave both strands of double-strand DNA either
at or near a recognition sequence that tends to be palindromic.  Consequently most restriction
endonucleases are dimeric and recognize symmetric DNA sequences.  While showing many
functional similarities in DNA recognition and catalytic cleavage, restriction endonucleases
also display low sequence homology, and diversity in mechanisms of recognizing DNA target
sequences and the positioning of metal cofactors.  Such diversity results in subtle variations
in both protein binding locations and potential functional roles for essential metal cofactors
that are only now coming under investigation.

<>

<1>Cox, K.L., Baltz, R.H.
<2>Restriction of bacteriophage plaque formation in Streptomyces spp.
<3>J. Bacteriol.
<4>159
<5>499-504
<6>1984
<7>Several Streptomyces species that produce restriction endonucleases were
characterized for their ability to propagate 10 different broad host range
bacteriophages.  Each species displayed a different pattern of plaque
formation.  A restrictionless mutant of S. albus G allowed plaque formation by
all 10 phages, whereas the wildtype strain showed plaques with only 2 phages.
DNA isolated from three of the phages was analyzed for the presence of
restriction sites for Streptomyces species-encoded enzymes, and a very strong
correlation was established between the failure to form plaques on Streptomyces
species that produced particular restriction enzymes and the presence of the
corresponding restriction sites in the phage DNA.  Also, the phages that lacked
restriction sites in their DNA generally formed plaques on the corresponding
restriction endonuclease-producing hosts at high efficiency.  The DNAs from the
three phages analyzed also generally contained either many or no restriction
sites for the Streptomyces species-produced enzymes, suggesting a strong
evolutionary trend to either eliminate all or tolerate many restriction sites.
The data indicate that restriction plays a major role in host range
determination for Streptomyces phages.  Analysis of bacteriophage host ranges
of many other uncharacterized Streptomyces hosts has identified four relatively
nonrestricting hosts, at least two of which may be suitable hosts for gene
cloning.  The data also suggest that several restriction systems remain to be
identified in the genus Streptomyces.

<>

<1>Cox, R., Goorha, S., Irving, C.
<2>Acrolein, a metabolite of cyclophosphamide, alters DNA methylase activity in a non-competitive fashion by reacting with -SH groups of the enzyme.
<3>Proc. Amer. Assoc. Cancer Res.
<4>28
<5>86
<6>1987
<7>Cyclophosphamide induces urinary bladder cancer both in humans and rats. Acrolein, an active
metabolite of cyclophosphamide, may be responsible for the bladder cancer induced by
cyclophosphamide.  Experiments are in progress to determine the carcinogenicity of acrolein in
rat bladder.  Studies have also been initiated to examine the biochemical lesions induced
following exposure of urothelial cells to acrolein.  In vitro studies on the effect of
acrolein on DNA methylase, a -SH containing enzyme (R. Cox, Cancer Res., 40:61, 1980), were
initiated to determine if acrolein might alter the DNA methylation pattern. Acrolein was shown
to react with the -SH group of cytochrome P450, resulting in inactivation (A.J. Marinello et
al., Cancer Res., 44:4615, 1984).  DNA methylase was isolated from the liver and urothelium of
rats and assayed as previously described (R. Cox, Biochem. Int., 5:787, 1982).  Acrolein
inhibited DNA methylase activity by 50% and 92% at final concentrations of 10 and 100 microM
acrolein.  Kinetic studies demonstrated non-competitive inhibition wiuth a Ki of 6.7 microM.
The inhibition of DNA methylase by 20 microM acrolein was restored to 85% of its original
activity when 100 microM dithiothreitol was added.  As the enzyme concentration was increased,
the inhibition of DNA methylase by 20 microM acrolein was decreased, whereas increased amounts
of DNA had no effect.  These data suggest that acrolein is a very good inhibitor of DNA
methylase, in a non-competitive fashion, by reacting with the -SH groups of the protein.

<>

<1>Cox, R., Goorha, S., Irving, C.C.
<2>Inhibition of DNA methylase activity by acrolein.
<3>Carcinogenesis
<4>9
<5>463-465
<6>1988
<7>Acrolein, a reactive metabolite of cyclophosphamide, may be responsible for bladder cancer
induced by cyclophosphamide.  DNA methylase was isolated from the liver and urothelium of rats
by high salt extraction of purified nuclei.  Acrolein at 10 microM inhibited liver and bladder
DNA methylase activity by 30-50%.  Kinetic studies with the liver enzyme showed a competitive
type of inhibition with a Ki of 6.7 microM.  Both dithiothreitol and glutathione afforded
protection to the enzyme when added to the assay.  At near equimolar concentrations of
glutathione to acrolein, the methylase retained 80-90% activity.  An increase in DNA had no
effect on the inhibition by acrolein, whereas increased amounts of protein protected against
acrolein inhibition, suggesting that acrolein reacted with the DNA methylase protein.  On the
other hand, DNA that had been reacted with acrolein was unable to serve as a substrate for DNA
methylase.  As the DNA adducts increased the methylation of the DNA decreased.  Thus, acrolein
has the ability to react with DNA and the DNA methylase protein, either of which results in
inhibition of DNA methylation.

<>

<1>Coyne, M.J., Zitomersky, N.L., McGuire, A.M., Earl, A.M., Comstock, L.E.
<2>Evidence of Extensive DNA Transfer between Bacteroidales Species within the Human Gut.
<3>MBio
<4>5
<5>e01305-14
<6>2014
<7>The genome sequences of intestinal Bacteroidales strains reveal evidence of extensive
horizontal gene transfer. In vitro studies of Bacteroides and other bacteria have addressed
mechanisms of conjugative transfer and some phenotypic outcomes of these DNA acquisitions in
the recipient, such as the acquisition of antibiotic resistance. However, few studies have
addressed the horizontal transfer of genetic elements between bacterial species coresident in
natural microbial communities, especially microbial ecosystems of humans. Here, we examine the
genomes of Bacteroidales species from two human adults to identify genetic elements that were
likely transferred among these Bacteroidales while they were coresident in the intestine.
Using seven coresident Bacteroidales species from one individual and eight from another, we
identified five large chromosomal regions, each present in a minimum of three of the
coresident strains at near 100% DNA identity. These five regions are not found in any other
sequenced Bacteroidetes genome at this level of identity and are likely all integrative
conjugative elements (ICEs). Such highly similar and unique regions occur in only 0.4% of
phylogenetically representative mock communities, providing strong evidence that these five
regions were transferred between coresident strains in these subjects. In addition to the
requisite proteins necessary for transfer, these elements encode proteins predicted to
increase fitness, including orphan DNA methylases that may alter gene expression, fimbriae
synthesis proteins that may facilitate attachment and the utilization of new substrates,
putative secreted antimicrobial molecules, and a predicted type VI secretion system (T6SS),
which may confer a competitive ecological advantage to these strains in their complex
microbial ecosystem.IMPORTANCE By analyzing Bacteroidales strains coresident in the gut
microbiota of two human adults, we provide strong evidence for extensive interspecies and
interfamily transfer of integrative conjug!
ative el
ements within the intestinal microbiota of individual humans. In the recipient strain, we show
that the conjugative elements themselves can be modified by the transposition of insertion
sequences and retroelements from the recipient's genome, with subsequent transfer of these
modified elements to other members of the microbiota. These data suggest that the genomes of
our gut bacteria are substantially modified by other, coresident members of the ecosystem,
resulting in highly personalized Bacteroidales strains likely unique to that individual. The
genetic content of these ICEs suggests that their transfer from successful adapted members of
an ecosystem confers beneficial properties to the recipient, increasing its fitness and
allowing it to better compete within its particular personalized gut microbial ecosystem.

<>

<1>Craig, J.W., Chang, F.Y., Kim, J.H., Obiajulu, S.C., Brady, S.F.
<2>Expanding Small-Molecule Functional Metagenomics through Parallel Screening of Broad-Host-Range Cosmid Environmental DNA Libraries in Diverse Proteobacteria.
<3>Appl. Environ. Microbiol.
<4>76
<5>1633-1641
<6>2010
<7>The small-molecule biosynthetic diversity encoded within the genomes of
uncultured bacteria is an attractive target for the discovery of natural
products using functional metagenomics. Phenotypes commonly associated
with the production of small molecules, such as antibiosis, altered
pigmentation, or altered colony morphology, are easily identified from
screens of arrayed metagenomic library clones. However, functional
metagenomic screening methods are limited by their intrinsic dependence on
a heterologous expression host. Toward the goal of increasing the
small-molecule biosynthetic diversity found in functional metagenomic
studies, we report the phenotypic screening of broad-host-range
environmental DNA libraries in six different proteobacteria: Agrobacterium
tumefaciens, Burkholderia graminis, Caulobacter vibrioides, Escherichia
coli, Pseudomonas putida, and Ralstonia metallidurans. Clone-specific
small molecules found in culture broth extracts from pigmented and
antibacterially active clones, as well as the genetic elements responsible
for the biosynthesis of these metabolites, are described. The host strains
used in this investigation provided access to unique sets of clones
showing minimal overlap, thus demonstrating the potential advantage
conferred on functional metagenomics through the use of multiple diverse
host species.

<>

<1>Craig, R.J., Arraj, J.A., Marinus, M.G.
<2>Induction of damage inducible (SOS) repair in dam mutants of Escherichia coli exposed to 2-aminopurine.
<3>Mol. Gen. Genet.
<4>194
<5>539-540
<6>1984
<7>2-Aminopurine induces damage inducible (SOS) repair in an Escherichia coli dam-4 strain but
not in a dam-4 mutS456 derivative or in dam+ bacteria.

<>

<1>Crampton, N., Roes, S., Dryden, D.T., Rao, D.N., Edwardson, J.M., Henderson, R.M.
<2>DNA looping and translocation provide an optimal cleavage mechanism for the type III restriction enzymes.
<3>EMBO J.
<4>26
<5>3815-3825
<6>2007
<7>EcoP15I is a type III restriction enzyme that requires two recognition sites in a defined
orientation separated by up to 3.5 kbp to efficiently
cleave DNA. The mechanism through which site-bound EcoP15I enzymes
communicate between the two sites is unclear. Here, we use atomic force
microscopy to study EcoP15I-DNA pre-cleavage complexes. From the number
and size distribution of loops formed, we conclude that the loops observed
do not result from translocation, but are instead formed by a contact
between site-bound EcoP15I and a nonspecific region of DNA. This
conclusion is confirmed by a theoretical polymer model. It is further
shown that translocation must play some role, because when translocation
is blocked by a Lac repressor protein, DNA cleavage is similarly blocked.
On the basis of these results, we present a model for restriction by type
III restriction enzymes and highlight the similarities between this and
other classes of restriction enzymes.

<>

<1>Crampton, N., Yokokawa, M., Dryden, D.T.F., Edwardson, J., Michael, R., Desirazu, N., Takeyasu, K., Yoshimura, S.H., Henderson, R.M.
<2>Fast-scan atomic force microscopy reveals that the type III restriction enzyme EcoP15I is capable of DNA translocation and looping.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>12755-12760
<6>2007
<7>Many DNA-modifying enzymes act in a manner that requires communication between two
noncontiguous DNA sites.  These sites can be brought into contact either by a
diffusion-mediated chance interaction between enzymes bound at the two sites, or by active
translocation of the intervening DNA by a site-bound enzyme.  EcoP15I, a type III restriction
enzyme, needs to interact with two recognition sites separated by up to 3,500 bp before it can
cleave DNA.  Here, we have studied the behavior of EcoP15I, using a novel fast-scan atomic
force microscope, which uses a miniaturized cantilever and scan stage to reduce the mechanical
response time of the cantilever and to prevent the onset of resonant motion at high scan
speeds.  With this instrument, we were able to achieve scan rates of up to 10 frames per s
under fluid.  The improved time resolution allowed us to image EcoP15I in real time at scan
rates of 1-3 frames per s.  EcoP15I translocated DNA in an ATP-dependent manner, at a rate of
79 +/- 33 bp/s.  The accumulation of supercoiling, as a consequence of movemennt of EcoP15I
along the DNA, could also be observed.  EcoP15I bound to its recognition site was also seen to
make nonspecific contacts with other DNA sites, thus forming DNA loops and reducing the
distance between the two recognition sites.  On the basis of our results, we conclude that
EcoP15I uses two distinct mechanisms to communicate between two recognition sites: diffusive
DNA loop formation and ATPase-driven translocation of the intervening DNA contour.

<>

<1>Cranz, S., Beck, C., Roth, M., Jeltsch, A.
<2>Molecular enzymology of the adenine-N6 DNA methyltransferase M.EcoRV: Site-directed mutagenesis analysis of DNA sequence and target base recognition.
<3>Biochem. Soc. Trans.
<4>28
<5>A309
<6>2000
<7>Methylation of DNA is important in many organisms and essential in mammals.  Nucleobases can
be methylated at adenine-N6, cytosine-N4 or cytosine-C6 atoms by specific DNA
methyltransferases.  The M.EcoRV DNA-(adenine-N6)-methyltransferase specifically transfers a
methyl group from AdoMet to the first adenine within GATATC sequences.  Here, we have
investigated the target base and DNA sequence recognition by M.EcoRV using site-directed
mutagenesis.  We show that variants of M.EcoRV in which active site residues are exchanged
(K16R, Y196W) show a >100 fold altered target base specificity and prefer methylation of
cytosine residues over adenine residues.  M.EcoRV is closely related to the dam DNA
methyltransferase which modifies adenine residues within GATC and M.EcoRV also accepts this
sequence.  To investigate DNA recognition by M.EcoRV, several amino acid residues in putative
DNA interacting regions were exchanged, i.e. in the variable domain of the enzyme as well as
at the N-terminus.  The mutants were analyzed with respect to their ability to modify GATATC,
GATC and GATCTC sequences in order to find out, which part of the recognition sequence is
contacted by the corresponding amino acid residue.  Our data show that some of the variants
display an increased specificity N136A, C140A, Lys141, Lys142 and Arg145 are important for DNA
binding of the enzyme, and that the N-terminal part of the M.EcoRV protein is not only
involved in AdoMet binding but also in DNA recognition.

<>

<1>Crasta, O.R., Folkerts, O., Fei, Z., Mane, S.P., Evans, C., Martino-Catt, S., Bricker, B., Yu, G., Du, L., Sobral, B.W.
<2>Genome sequence of Brucella abortus vaccine strain S19 compared to virulent strains yields candidate virulence genes.
<3>PLoS ONE
<4>3
<5>e2193
<6>2008
<7>The Brucella abortus strain S19, a spontaneously attenuated strain, has been used as a vaccine
strain in vaccination of cattle against brucellosis
for six decades. Despite many studies, the physiological and molecular
mechanisms causing the attenuation are not known. We have applied
pyrosequencing technology together with conventional sequencing to rapidly
and comprehensively determine the complete genome sequence of the
attenuated Brucella abortus vaccine strain S19. The main goal of this
study is to identify candidate virulence genes by systematic comparative
analysis of the attenuated strain with the published genome sequences of
two virulent and closely related strains of B. abortus, 9-941 and 2308.
The two S19 chromosomes are 2,122,487 and 1,161,449 bp in length. A total
of 3062 genes were identified and annotated. Pairwise and reciprocal
genome comparisons resulted in a total of 263 genes that were
non-identical between the S19 genome and any of the two virulent strains.
Amongst these, 45 genes were consistently different between the attenuated
strain and the two virulent strains but were identical amongst the
virulent strains, which included only two of the 236 genes that have been
implicated as virulence factors in literature. The functional analyses of
the differences have revealed a total of 24 genes that may be associated
with the loss of virulence in S19. Of particular relevance are four genes
with more than 60 bp consistent difference in S19 compared to both the
virulent strains, which, in the virulent strains, encode an outer membrane
protein and three proteins involved in erythritol uptake or metabolism.

<>

<1>Crawford, J.T., Cave, M.D., Bates, J.H.
<2>Evidence for plasmid-mediated restriction-modification in Mycobacterium avium intracellulare.
<3>J. Gen. Microbiol.
<4>127
<5>333-338
<6>1981
<7>Mycobacterium avium intracellular strain LR25 carries three plasmids with
molecular weights of 11.2, 18.3 and 107 x 10(6) as determined by electron
microscopy.  A number of phages propagated on Mycobacterium smegmatis ATCC 607
were tested for their ability to infect strain LR25.  Phage JF2 gave an
efficiency of plating of 10-4 on strain LR25, but phage JF2 propagated on
strain LR25 infected strain LR25 and M. smegmatis with equal high efficiency.
This indicated the presence of a restriction-modification (R-M) system in
strain LR25 that was not present in M. smegmatis.  Strain LR25 was grown in the
presence of acriflavine to eliminate the plasmids and tested for sensitivity to
phage JF2.  One of forty colonies was found to be R-M-deficient.  This strain,
designated strain LR163, lacks the three plasmids present in strain LR25.  The
results indicate that the R-M system is plasmid-coded.  Strain LR163 was
sensitive to several phages to which strain LR25 was resistant and for which we
were unable to isolate modified phage.  This suggests that some plasmid-coded
function in addition to restriction is involved.  An R-M system was also
demonstrated in M. avium intracellulare strain LR131 using phage JF1.  This
strain does not carry plasmids.

<>

<1>Cregg, J.M., Nguyen, A.H., Ito, J.
<2>DNA modification induced during infection of Bacillus subtilis by phage Phi3T.
<3>Gene
<4>12
<5>17-24
<6>1980
<7>The DNA of the Bacillus subtilis temperate phage Phi3T is not susceptible to
cleavage by the restriction endonuclease HaeIII, although it is cut by many
other restriction enzymes.  The host DNA from uninfected cells is cut by
HaeIII.  We show that Phi3T DNA propagated in a
restriction-modification-defective Escherichia coli cell can be digested by
HaeIII.  Thus, Phi3T DNA does contain the nucleotide recognition sequence of
HaeIII.  We suggest that this phage induces the modification of its own DNA.
In support of this mechanism we show that extracts prepared from Phi3T-infected
cells contain an activity which in the presence of S-adenosyl-L-methionine
(SAM) can modify lambda DNA against cleavage by HaeIII.  The same in
vitro-modified DNA is still susceptible to cleavage by other restriction
endonucleases.

<>

<1>Cregg, J.M., Stewart, C.R.
<2>EcoRI cleavage of DNA from Bacillus subtilis phage SPO1.
<3>Virology
<4>85
<5>601-605
<6>1978
<7>The hydroxymethyluracil-containing DNA of B. subtilis phage SPO1 is cleaved only with low
efficiency by restriction nuclease EcoRI.  The efficiency is greatly increased by EcoRI*
conditions, but even under these conditions, the efficiency is less than with
thymine-containing DNA.  In contrast with their effect on thymine-containing DNA, EcoRI*
conditions do not appear to increase the number of sites on the SPO1 genome at which cleavage
takes place.  We describe the fragments produced by a (nearly) limit digest of SPO1 DNA with
EcoRI and assign most of the known SPO1 cistrons to specific fragments.

<>

<1>Cress, B.F., Erkert, K.A., Barquera, B., Koffas, M.A.
<2>Draft Genome Sequence of Pseudoalteromonas luteoviolacea Strain B (ATCC 29581).
<3>Genome Announcements
<4>1
<5>e0004813
<6>2013
<7>We report the 4.049-Mbp high-quality draft assembly of the Pseudoalteromonas luteoviolacea
strain B (ATCC 29581) genome. This marine species is known to
biosynthesize several antimicrobial compounds, including the purple pigment
violacein. Whole-genome sequencing and genome mining will complement experimental
studies aimed at elucidating novel biosynthetic pathways capable of producing
pharmaceutically relevant molecules. Based upon 16S rRNA phylogenetic analysis,
we propose that strain ATCC 29581 be classified as a distinct phylogenetic
species of the genus Pseudoalteromonas.

<>

<1>Cress, B.F., Greene, Z.R., Linhardt, R.J., Koffas, M.A.
<2>Draft Genome Sequence of Escherichia coli Strain ATCC 23506 (Serovar O10:K5:H4).
<3>Genome Announcements
<4>1
<5>e0004913
<6>2013
<7>We report the 5.101-Mbp high-quality draft assembly of the Escherichia coli strain ATCC 23506
(serovar O10:K5:H4, also known as NCDC Bi 8337-41) genome. This
uropathogenic strain, commonly referred to as E. coli K5, produces N-acetyl
heparosan, a glycosaminoglycan-like capsular polysaccharide and precursor to the
anticoagulant pharmaceutical heparin. Metabolic reconstruction of this genome
will enable the prediction of gene deletions and overexpressions that lead to
increased heparosan production.

<>

<1>Cress, B.F., Greene, Z.R., Linhardt, R.J., Koffas, M.A.
<2>Draft Genome Sequence of Escherichia coli Strain ATCC 23502 (Serovar O5:K4:H4).
<3>Genome Announcements
<4>1
<5>e0004613
<6>2013
<7>We report the 4.682-Mbp high-quality draft assembly of the Escherichia coli strain ATCC 23502
(serovar O5:K4:H4, also known as NCDC U1-41) genome. This
uropathogenic strain, commonly referred to as E. coli K4, produces a
glycosaminoglycan-like capsular polysaccharide with a backbone similar in
structure to unsulfated chondroitin, a precursor to the nutraceutically and
potentially pharmaceutically valuable compound chondroitin sulfate. Metabolic
reconstruction of this genome will enable prediction of genetic engineering
strategies leading to increased chondroitin production.

<>

<1>Cress, B.F., Linhardt, R.J., Koffas, M.A.
<2>Draft Genome Sequence of Escherichia coli Strain Nissle 1917 (Serovar O6:K5:H1).
<3>Genome Announcements
<4>1
<5>e0004713
<6>2013
<7>We announce the availability of the 5.023-Mbp high-quality draft assembly of the  Escherichia
coli strain Nissle 1917 (serovar O6:K5:H1) genome. Short genomic
segments from this important probiotic strain have been available in public
databases, but the full genome sequence has remained inaccessible. Thus,
high-coverage, whole genome sequencing of E. coli Nissle 1917 is presented
herein. Reannotation and metabolic reconstruction will enable comparative
genomics analysis and model-guided predictions of genetic manipulations leading
to increased production of the K5 capsular polysaccharide known as N-acetyl
heparosan, a precursor to the anticoagulant pharmaceutical heparin.

<>

<1>Criscuolo, A., de la Blanchardiere, A., Coeuret, S., Passet, V., Saguet-Rysanek, V., Vergnaud, M., Verdon, R., Leclercq, A., Lecuit, M., Brisse, S.
<2>Draft Genome Sequence of Campylobacter coli Strain IPSID-1 Isolated from a Patient with Immunoproliferative Small Intestinal Disease.
<3>Genome Announcements
<4>2
<5>e00079-14
<6>2014
<7>The genome sequence and annotation of Campylobacter coli strain IPSID-1 are reported here.
This bacterial isolate is the first to be cultured from a patient
with immunoproliferative small intestinal disease (IPSID). The draft genome
sequence is 1.683 Mb long, comprises 64 contigs, and has 31.26% G+C content.

<>

<1>Croce, O., Hugon, P., Lagier, J.C., Bibi, F., Robert, C., Azhar, E.I., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Bacillus simplex Strain P558, Isolated from a Human Fecal Sample.
<3>Genome Announcements
<4>2
<5>e01241-14
<6>2014
<7>Bacillus simplex strain P558 was isolated from a fecal sample of a 25-year-old Saudi male. We
sequenced the 5.98-Mb genome of the strain and compared it to that
of B. simplex strain 1NLA3E.

<>

<1>Croce, O., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium asiaticum Strain DSM 44297.
<3>Genome Announcements
<4>2
<5>e00320-14
<6>2014
<7>We report the draft genome sequence of Mycobacterium asiaticum strain DSM 44297,  a tropical
mycobacterium seldom responsible for human infection. The genome of M. asiaticum has a size of
5,935,986 bp, with a 66.03% G+C content, encoding 5,591 proteins and 81 RNAs.

<>

<1>Croce, O., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium austroafricanum DSM 44191.
<3>Genome Announcements
<4>2
<5>e00317-14
<6>2014
<7>We announce the draft genome sequence of Mycobacterium austroafricanum DSM 44191(T) (=
E9789-SA12441(T)), a non-tuberculosis species responsible for opportunistic infection. The
genome described here has a size of 6,772,357 bp with a G+C content of 66.79% and contains
6,419 protein-coding genes and 112 RNA genes.

<>

<1>Croce, O., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium cosmeticum DSM 44829.
<3>Genome Announcements
<4>2
<5>e00315-14
<6>2014
<7>We announce the draft genome sequence of Mycobacterium cosmeticum strain DSM 44829, a
nontuberculous species responsible for opportunistic infection. The genome described here is
composed of 6,462,090 bp, with a G+C content of 68.24%. It contains 6,281 protein-coding genes
and 75 predicted RNA genes.

<>

<1>Croce, O., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium vulneris DSM 45247T.
<3>Genome Announcements
<4>2
<5>e00370-14
<6>2014
<7>We report the draft genome sequence of Mycobacterium vulneris DSM 45247(T) strain, an
emerging, opportunistic pathogen of the Mycobacterium avium complex.
The genome described here is composed of 6,981,439 bp (with a G+C content of
67.14%) and has 6,653 protein-coding genes and 84 predicted RNA genes.

<>

<1>Croce, O., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium mageritense DSM 44476T.
<3>Genome Announcements
<4>2
<5>e00354-14
<6>2014
<7>We report the draft genome sequence of Mycobacterium mageritense strain DSM 44476(T) (CIP
104973), a nontuberculosis species responsible for various
infections. The genome described here is composed of 7,966,608 bp, with a G+C
content of 66.95%, and contains 7,675 protein-coding genes and 120 predicted RNA
genes.

<>

<1>Croce, O., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium farcinogenes NCTC 10955.
<3>Genome Announcements
<4>2
<5>e00523-14
<6>2014
<7>We report the draft genome sequence of Mycobacterium farcinogenes NCTC 10955 (=DSM 43637(T)),
a nontuberculosis species responsible for bovine farcy. The
strain described here is composed of 6,139,893 bp, with a G+C content of 65.73%,
and contains 5,816 protein-coding genes and 76 RNA genes.

<>

<1>Crook, M.B., Mitra, S., Ane, J.M., Sadowsky, M.J., Gyaneshwar, P.
<2>Complete Genome Sequence of the Sesbania Symbiont and Rice Growth-Promoting Endophyte Rhizobium sp. Strain IRBG74.
<3>Genome Announcements
<4>1
<5>e00934-13
<6>2013
<7>Rhizobium sp. strain IRBG74 is the first known nitrogen-fixing symbiont in the
Agrobacterium/Rhizobium clade that nodulates the aquatic legume Sesbania sp. and
is also a growth-promoting endophyte of wetland rice. Here, we present the
sequence of the IRBG74 genome, which is composed of a circular chromosome, a
linear chromosome, and a symbiotic plasmid, pIRBG74a.

<>

<1>Crooke, H.R., Clarke, E.E., Everest, P.H., Dougan, G., Holden, D.W., Shea, J.E., Feldman, R.G.
<2>Virulence genes and proteins, and their use.
<3>Japanese Patent Office
<4>JP 2002529091 A
<5>
<6>2002
<7>
<>

<1>Crooke, H.R., Clarke, E.E., Everest, P.H., Dougan, G., Holden, D.W., Shea, J.E., Feldman, R.G.
<2>VIRULENCE GENES AND PROTEINS, AND THEIR USE.
<3>Korean Patent Office
<4>KR 1020017005731 A
<5>
<6>2001
<7>
<>

<1>Crooks, P.A., Tribe, M.J., Pinney, R.J.
<2>Inhibition of bacterial DNA cytosine-5-methyltransferase by S-adenosyl-L-homocysteine and some related compounds.
<3>J. Pharm. Pharmacol.
<4>36
<5>85-89
<6>1984
<7>S-Adenosyl-L-homocysteine and five related compounds have been evaluated as inhibitors of a
DNA cytosine-5-methyltransferase. DNA methylation was assayed in cell extracts from E. coli
strain J6-2 dcm+, proficient in DNA cytosine-5-methyltransferase activity, containing
substrate DNA isolated from E. coli strain J6-2 dcm-, a strain deficient in DNA
cytosine-5-methyltransferase. S-Adenosyl-L-homocysteine and its 7-deaza analogue,
S-tubercidinylhomocysteine, were competitive inhibitors of DNA cytosine-5-methyltransferase
with Ki's of 14.2 and 17.6 microM, respectively, in the above enzyme assay.

<>

<1>Cross, S.H., Meehan, R.R., Nan, X., Bird, A.
<2>A component of the transcriptional repressor MeCP1 shares a motif with DNA methyltransferase and HRX proteins.
<3>Nat. Genet.
<4>16
<5>256-259
<6>1997
<7>Methylation of cytosines within the sequence CpG is essential for mouse development and has
been linked to transcriptional suppression in vertebrate systems.  Methyl-CpG binding proteins
(MeCPs) 1 and 2 bind preferentially to methylated DNA and can inhibit transcription.  The gene
for MeCP2 has been cloned and a methyl-CpG binding domain within it has been defined.  A
search of DNA sequence databases with the MBD sequence identified a human cDNA with potential
to encode an MBD-like region.  Sequencing of the complete cDNA revealed that the open reading
frame also encodes two cysteine-rich domains that are found in animal DNA methyltransferases
and in the mammalian HRX protein (also known as MLL and ALL-1).  HRX is related to Drosophila
trithorax.  The protein, known as Protein Containing MBD (PCM1), was expressed in bacteria and
shown to bind specifically to methylated DNA.  PCM1 also repressed transcription in vitro in a
methylation-dependent manner.  A polyclonal antibody raised against the protein was able to
'supershift' the native MeCP1 complex from HeLa cells, indicating that PCM1 is a component
of mammalian MeCP1.

<>

<1>Crossman, L.C. et al.
<2>The complete genome, comparative and functional analysis of Stenotrophomonas maltophilia reveals an organism heavily shielded by drug  resistance determinants.
<3>Genome Biol.
<4>9
<5>R74
<6>2008
<7>ABSTRACT: BACKGROUND: Stenotrophomonas maltophilia is a nosocomial opportunistic pathogen of
the Xanthomonadaceae. The organism has been
isolated from both clinical and soil environments in addition to the
sputum of cystic fibrosis patients and the immunocompromised. Whilst
relatively distant phylogenetically, the closest sequenced relatives of S.
maltophilia are the plant pathogenic xanthomonads. RESULTS: The genome of
the bacteremia-associated isolate S. maltophilia K279a is 4,851,126 bp and
of high G+C content. The sequence reveals an organism with a remarkable
capacity for drug and heavy metal resistance. In addition to a number of
genes conferring resistance to antimicrobial drugs of different classes
via alternative mechanisms, nine resistance-nodulation-division (RND)-type
putative antimicrobial efflux systems are present. Functional genomic
analysis confirms a role in drug resistance for several of the novel RND
efflux pumps. S. maltophilia possesses potentially mobile regions of DNA
and encodes a number of pili and fimbriae likely to be involved in
adhesion and biofilm formation that may also contribute to increased
antimicrobial drug resistance. CONCLUSION: The panoply of antimicrobial
drug resistance genes and mobile genetic elements found suggests that the
organism can act as a reservoir of antimicrobial drug resistance
determinants in a clinical environment, which is an issue of considerable
concern.

<>

<1>Crossman, L.C. et al.
<2>A Commensal Gone Bad: Complete Genome Sequence of the Prototypical Enterotoxigenic Escherichia coli Strain H10407.
<3>J. Bacteriol.
<4>192
<5>5822-5831
<6>2010
<7>In most cases, Escherichia coli exists as a harmless commensal organism, but it may on
occasion cause intestinal and/or extraintestinal disease.
Enterotoxigenic E. coli (ETEC) is the predominant cause of E.
coli-mediated diarrhea in the developing world and is responsible for a
significant portion of pediatric deaths. In this study, we determined the
complete genomic sequence of E. coli H10407, a prototypical strain of
enterotoxigenic E. coli, which reproducibly elicits diarrhea in human
volunteer studies. We performed genomic and phylogenetic comparisons with
other E. coli strains, revealing that the chromosome is closely related to
that of the nonpathogenic commensal strain E. coli HS and to those of the
laboratory strains E. coli K-12 and C. Furthermore, these analyses
demonstrated that there were no chromosomally encoded factors unique to
any sequenced ETEC strains. Comparison of the E. coli H10407 plasmids with
those from several ETEC strains revealed that the plasmids had a mosaic
structure but that several loci were conserved among ETEC strains. This
study provides a genetic context for the vast amount of experimental and
epidemiological data that have been published.

<>

<1>Croucher, N.J. et al.
<2>Dominant Role of Nucleotide Substitution in the Diversification of Serotype 3 Pneumococci over Decades and during a Single Infection.
<3>PLoS Genet.
<4>9
<5>e1003868-100386
<6>2013
<7>Streptococcus pneumoniae of serotype 3 possess a mucoid capsule and cause disease associated
with high mortality rates
relative to other pneumococci. Phylogenetic analysis of a complete reference genome and 81
draft sequences from clonal
complex 180, the predominant serotype 3 clone in much of the world, found most sampled
isolates belonged to a clade
affected by few diversifying recombinations. However, other isolates indicate significant
genetic variation has accumulated
over the clonal complex's entire history. Two closely related genomes, one from the blood and
another from the
cerebrospinal fluid, were obtained from a patient with meningitis. The pair differed in their
behaviour in a mouse model of
disease and in their susceptibility to antimicrobials, with at least some of these changes
attributable to a mutation that upregulated
the patAB efflux pump. This indicates clinically important phenotypic variation can accumulate
rapidly through
small alterations to the genotype.

<>

<1>Crouzier, L., Dubois, C., Wengel, J., Veedu, R.N.
<2>Cleavage and protection of locked nucleic acid-modified DNA by restriction endonucleases.
<3>Bioorg. Med. Chem. Lett.
<4>22
<5>4836-4838
<6>2012
<7>Locked nucleic acid (LNA) is one of the most prominent nucleic acid analogues reported so far.
We herein for the first time report cleavage
by restriction endonuclease of LNA-modified DNA oligonucleotides. The
experiments revealed that RsaI is an efficient enzyme capable of
recognizing and cleaving LNA-modified DNA oligonucleotides.
Furthermore, introduction of LNA nucleotides protects against cleavage
by the restriction endonucleases PvuII, PstI, SacI, KpnI and EcoRI.

<>

<1>Crowley, S., Bottacini, F., Mahony, J., van Sinderen, D.
<2>Complete Genome Sequence of Lactobacillus plantarum Strain 16, a Broad-Spectrum Antifungal-Producing Lactic Acid Bacterium.
<3>Genome Announcements
<4>1
<5>e00533-13
<6>2013
<7>Lactobacillus plantarum strain 16 restricts the growth of various food spoilage fungi and has
the potential to be used as a biopreservative to improve the shelf
life of foods. The complete genome sequence contains 3,044,678 bp with a G+C
content of 44.74% and harbors the largest plasmid complement reported for this
species to date.

<>

<1>Cruz, A.K., Kidane, G., Pires, M.Q., Rabinovitch, L., Guaycurus, T.V., Morel, C.M.
<2>An Sau3AI restriction endonuclease isoschizomer from Bacillus cereus.
<3>FEBS Lett.
<4>173
<5>99-102
<6>1984
<7>The isolation and characterization of a restriction endonuclease from Bacillus
cereus 10C 243 are described.  The enzyme recognizes the palindromic sequence
5'-G(met-A,A)TC-3' as determined by PEI chromatography of pancreatic DNase,
snake venom phosphodiesterase digestion products of labelled fragments,
analysis of restriction digests from normal and N6-methyladenine-free DNA and
direct sequence analysis of cloned fragments.  The staggered cleavage products
with 5'-terminal pGATC extensions are efficiently labelled with polynucleotide
kinase and are easily cloned into BamHI sites.  The enzyme, denoted Bce243, is
thus an isoschizomer of SauAI.  Its use and potential advantages in
substituting Sau3AI are discussed.

<>

<1>Cruz, A.K., Pires, M.Q., Lima, V.G., Morel, C.M.
<2>Restriction endonucleases from microorganisms isolated in Brazil: an isoschizomer of HaeIII from a thermophilic Bacillus sp.
<3>Braz. J. Med. Biol. Res.
<4>22
<5>1321-1328
<6>1989
<7>The isolation and characterization of a restriction endonuclease from a
thermophilic strain of Bacillus is described.  The enzyme recognizes the
palindromic sequence 5' GGCC 3' as determined by PEI-cellulose chromatography
of pancreatic DNAse and snake venom phosphodiesterase digestion products of
labelled DNA fragments, analysis of restriction digests and direct sequence
analysis.  The enzyme, denominated BspBR, is an isoschizomer of HaeIII and
BspRI.

<>

<1>Csepregi, K., Valasek, A., Penzes, A., Toth, Z., Kiss, E.I., Kerepesi, I., Horvath, B., Nagy, I., Fekete, C.
<2>Draft Genome Sequence of an Efficient Antibiotic-Producing Industrial Strain of Saccharomonospora azurea, SZMC 14600.
<3>J. Bacteriol.
<4>194
<5>1263
<6>2012
<7>Although certain rare actinomycetes have been recognized as prolific sources of bioactive
natural products, their potential for producing biologically active
metabolites still remains unexplored. With the aim of gaining global insights
into the genetic background and the metabolic capability of Saccharomonospora
azurea SZMC 14600, whole-genome sequencing was performed.

<>

<1>Cserhati, M., Kriszt, B., Szoboszlay, S., Toth, A., Szabo, I., Tancsics, A., Nagy, I., Horvath, B., Nagy, I., Kukolya, J.
<2>De Novo Genome Project of Cupriavidus basilensis OR16.
<3>J. Bacteriol.
<4>194
<5>2109-2110
<6>2012
<7>Here we report on the complete genome sequence of Cupriavidus basilensis OR16 NCAIM BO2487.
The genome of strain OR16 contains 7,534 putative coding sequences,
including a large set of xenobiotics-degrading genes and a unique glucose
dehydrogenase gene that is absent from other Cupriavidus genomes.

<>

<1>Csirik, J., Magyar, J., Polner, G.
<2>A computer algorithm to determine the recognition site of restriction enzymes.
<3>Comput. Appl. Biosci.
<4>3
<5>245-246
<6>1987
<7>A new algorithm is proposed to determine the type-II restriction endonucleases'
recognition site knowing the digested DNA sequence and fragment lengths in an
actual case.  The algorithm is implemented for the Commodore 64 microcomputer.

<>

<1>Cuadros-Orellana, S., Martin-Cuadrado, A.B., Legault, B., D'Auria, G., Zhaxybayeva, O., Papke, R.T., Rodriguez-Valera, F.
<2>Genomic plasticity in prokaryotes: the case of the square haloarchaeon.
<3>ISME J.
<4>1
<5>235-245
<6>2007
<7>The variability in genome content among closely related strains of
prokaryotes has been one of the most remarkable discoveries of genomics.
One way to approach the description of this so-called pan-genome is to
compare one reference strain genome with metagenomic sequences from the
environment. We have applied this approach to one extreme aquatic habitat,
saturated brines in a solar saltern. The genome of Haloquadratum walsbyi
strain DSM 16790 was compared to an environmental metagenome obtained from
the exact site of its isolation. This approach revealed that some regions
of the strain genome were scarcely represented in the metagenome. Here we
have analyzed these genomic islands (GI) in the genome of DSM 16790 and
compared them with the complete sequence of some fosmids from the
environmental library. Two of the islands, GI 2 and GI 4, overlapped with
two large guanine and cytosine (GC)-rich regions that showed evidence of
high variability through mobile elements. GI 3 seemed to be a phage or
phage-remnant acquired by the reference genome, but not present in most
environmental lineages. Most differential gene content was related to
small molecule transport and detection, probably reflecting adaptation to
different pools of organic nutrients. GI 1 did not possess traces of
mobile elements and had normal GC content. This island contained the main
cluster of cell envelope glycoproteins and the variability found was
different from the other GIs. Rather than containing different genes it
consisted of homologs with low similarity. This variation might reflect a
phage evasion strategy.

<>

<1>Cuculich, P.S.
<2>Overexpression of E. coli EcoRI modification system enzymes.
<3>The Nucleus
<4>75
<5>8-10
<6>1996
<7>Escherichia coli RI (EcoRI) DNA restriction and modification enzymes are reponsible for host
specificity of E. coli.  The restriction endonuclease enzyme recognizes a hexanucleotide
repeat (5'-GAATTC-3') and cleaves double-stranded DNA in a staggered fashion between the
guanine and first adenine residues.  The methyltransferase enzyme, or methylase, adds a methyl
group to the second adenine, nearest the axis of symmetry, leaving the site resistant to
endonuclease cleavage.  The endonuclease and methylase are two independent proteins which are
coded for by distinct genes, and will be referred to collectively as the EcoRI modification
system.

<>

<1>Cue, D., Hong, L., Hanson, R.S., Flickinger, M.C.
<2>Characterization of a restriction-modification system of the thermotolerant methylotroph Bacillus methanolicus.
<3>Appl. Environ. Microbiol.
<4>62
<5>1107-1111
<6>1996
<7>We report the isolation of a restriction endonuclease, BmeTI, an isoschizomer of BclI, that
recognizes the DNA sequence 5' TGATCA 3'.  We also report that BmeTI sites are modified to
TGm6ATCA.  These findings provide the basis for devising strategies to prevent BmeTI
restriction of any DNA introduced into Bacillus methanolicus.

<>

<1>Cue, D., Lam, H., Hanson, R.S., Flickinger, M.
<2>Characterization of a Bacillus methanolicus restriction/modification system.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>95
<5>525
<6>1995
<7>A restriction endonuclease (BmeI) has been isolated from the thermotolerant, methylotrophic
bacterium Bacillus methanolicus.  BmeI is an isoschizomer of BclI, both enzymes cutting within
the sequence, TGATCA.  The same restriction patterns are evident when phage DNA or
HindIII-digested phage DNA are incubated with either BmeI or BclI or when phage DNA is
digested with both BmeI and BclI.  Destruction of the BclI restriction site present in the
integrational plasmid, pDQ500, resulted in a plasmid pDQ503, that is resistant to cleavage by
both BmeI and BclI.  The modification specificity of the cognate BmeI methyltransferase was
determined by Southern blotting of B. methanolicus chromosomal DNA that had been digested with
PstI and either BclI, DpnI, MboI or Sau3AI and using the cloned B. methanolicus, lysC gene as
the probe.  The results indicated that the single BmeI restriction site within lysC is
modified to: TGm6ATCA.  We have successfully transformed B. methanolicus with several plasmid
and integrational vectors (including pDQ503).  Our results suggest that B. methanolicus
possesses a single, dominant restriction system or, that if a second restriction system
exists, that the second restriction endonuclease is an infrequent cutter.

<>

<1>Cui, C., Li, P., Liu, G., Tang, H., Lin, K., Luo, Q., Liu, S., Xu, P., Liu, Y.
<2>Genome Sequence of Martelella sp. Strain AD-3, a Moderately Halophilic Polycyclic Aromatic Hydrocarbon-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e01189-13
<6>2014
<7>Martelella sp. strain AD-3, enriched from a petroleum-contaminated site with high salinity,
can efficiently degrade polycyclic aromatic hydrocarbons. Here, we
report the 4.75-Mb genome sequence of strain AD-3 with its genetic feature of
helping to remediate environmental organic pollutants.

<>

<1>Cui, G.-Z., Hong, W., Zhang, J., Li, W.-L., Feng, Y., Liu, Y.-J., Cui, Q.
<2>Targeted gene engineering in Clostridium cellulolyticum H10 without methylation.
<3>J. Microbiol. Methods
<4>89
<5>201-208
<6>2012
<7>Genetic engineering of Clostridium cellulolyticum has been developed slowly compared with that
of other clostridial species, and one of the
major reasons might be the restriction and modification (RM) system
which degrades foreign DNA. Here, a putative Mspl endonuclease gene,
ccel2866, was inactivated by a ClosTron-based gene disruption method.
The resulting C cellulolyticum mutant H10 Delta mspl lost the Mspl
endonuclease activity and can accept unmethylated DNA efficiently.
Following that, an oxygen-independent green fluorescence protein gene
was introduced into H10 Delta mspl without methylation, generating a
convenient reporter system to evaluate the expression of heterologous
protein in C. cellulolyticum by green fluorescence. To further
demonstrate the efficiency of the H10 Delta mspl, double mutants H10
Delta mspl Delta ldh and H10 Delta mspl Delta ack were constructed by
disrupting lactate dehydrogenase gene ccel2485 and acetate kinase gene
ccel2136 in H10 Delta mspl, respectively, without DNA methylation, and
the stability of the double mutation was confirmed after the 100th
generation. The mutant H10 Delta mspl constructed here can be used as a
platform for further targeted gene manipulation conveniently and
efficiently. It will greatly facilitate the metabolic engineering of C.
cellulolyticum aiming at faster cellulose degradation and higher
biofuel production at the molecular level.

<>

<1>Cui, X.X., Davis, G.
<2>Mobile group II intron targeting: applications in prokaryotes and perspectives in eukaryotes.
<3>Front. Biosci.
<4>12
<5>4972-4985
<6>2007
<7>Mobile group II introns are ribozymes and use a novel mechanism--target DNA-primed reverse
transcription--to proliferate in DNA. Group II introns are a unique mobile element for their
high sequence-specific, yet readily flexible target site recognition. Both the intron RNA and
the intron-encoded protein (IEP) are involved in target site recognition, and the specificity
is determined primarily by base pairing between the intron RNA and DNA target. Therefore, the
intron RNA can be modified according to the desired target sequence for specific gene
disruption. Group II intron knockout technology is mature in bacteria and is currently being
developed in eukaryotes. This technology has great potential to revolutionize fields such as
functional genomics, gene therapy, and cell line engineering.

<>

<1>Cui, Z., Gao, W., Li, Q., Xu, G., Zheng, L.
<2>Genome Sequence of the Polycyclic Aromatic Hydrocarbon-Degrading Bacterium Strain Marinobacter nanhaiticus D15-8WT.
<3>Genome Announcements
<4>1
<5>e00301-13
<6>2013
<7>Marinobacter nanhaiticus strain D15-8W(T) was isolated from a phenanthrene-degrading
consortium, enriched from sediment of the South China Sea.
Here, we present the draft genome of strain D15-8W(T), which contains 5,358,309
bp with a G+C content of 58.53% and contains 4,829 protein-coding genes and 47
tRNA genes.

<>

<1>Cui, Z., Xu, G., Li, Q., Gao, W., Zheng, L.
<2>Genome Sequence of the Pyrene- and Fluoranthene-Degrading Bacterium Cycloclasticus sp. Strain PY97M.
<3>Genome Announcements
<4>1
<5>e00536-13
<6>2013
<7>Cycloclasticus sp. strain PY97M was isolated from a phenanthrene-degrading consortium,
enriched from Yellow Sea sediment of China. Here, we present the
draft genome sequence of strain PY97M, which contains 2,359,509 bp with a G+C
content of 41.92% and contains 2, 264 protein-coding genes and 40 tRNAs.

<>

<1>Cuiv, P.O., Klaassens, E.S., Durkin, A.S., Harkins, D.M., Foster, L., McCorrison, J., Torralba, M., Nelson, K.E., Morrison, M.
<2>Draft genome sequence of Turicibacter sanguinis PC909 isolated from human faeces.
<3>J. Bacteriol.
<4>193
<5>1288-1289
<6>2010
<7>While the microbiota resident in the human gut is now known to provide a range of functions
relevant to host health many of the microbial members of the community have not yet been
cultured or are represented by a limited number of isolates. We describe here the draft genome
sequence of Turicibacter sanguinis PC909, isolated from a pooled healthy human faecal sample
as part of the Australian Human Gut Microbiome Project.

<>

<1>Culley, D.E., Shi, L., Reed, S., Romme, M.
<2>Genetic and biochemical studies of the transformation barriers present in Shewanella oneidensis MR-1.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>104
<5>320
<6>2004
<7>Shewanella oneidensis is able to enzymatically reduce and precipitate a variety of heavy
metals and is widely investigated for bioremediation of metals and radionuclides.  The
availability of the complete genome sequence and its ability to grow both aerobically and
anaerobically makes S. oneidensis MR-1 an extremely useful model system.  However, genetic
studies of this organism have been slowed by the lack of an efficient transformation system.
We are utilizing several approaches to investigate the genetic factors influencing
transformation and to improve the transformation efficiency.  Utilizing a pUC-type plasmid
isolated from MR-1, we were able to test various competent cell preparation methods,
electrical parameters, and selection strategies to develop a method that improves
electroporation efficiencies from less than 105 cfu/ug to almost 108/ug.  However, even these
improved conditions yielded less than 103/ug when transforming MR-1 with the same plasmid
isolated from E. coli.  This 100,000-fold reduction in efficiency with unmodified plasmid
indicates a very active restriction-modification system in MR-1.  Analysis of the genome
sequence indicates the presence of a dam methylase locus, three type II, and two Type I
methylases on the chromosome and a Type II methylase on a 160 kb megaplasmid.  The use of a
Type I restriction inhibitor did not improve transformation, indicating an active Type II
system.  The Type II system borne on the megaplasmid has high homology to the ClaI restriction
system, but no effect was observed with either in vivo modification using an E. coli strain
expressing ClaI methylase (pHS17; NEB) or with in vitro modification of the test plasmid with
M.TaqI (blocks ClaI cleavage).  One of the chromosomal Type II methylases is located within a
lambda-like prophage, but is lacking the corresponding restriction subunit.  We are currently
constructing MR-1 knockout mutants of the restriction subunits from the remaining two loci, as
well as attempting to modify plasmid DNA in E. coli by co-expression of the corresponding
Shewanella methylases, in hopes of overcoming the transformation barrier present in S.
oneidensis.

<>

<1>Cullik, A., Pfeifer, Y., Prager, R., von Baum, H., Witte, W.
<2>A novel IS26 structure surrounds blaCTX-M genes in different plasmids from German clinical Escherichia coli isolates.
<3>J. Med. Microbiol.
<4>59
<5>580-587
<6>2010
<7>This report focuses on the molecular characterization of 22
extended-spectrum beta-lactamase-producing Escherichia coli isolates
collected in a German university hospital during a period of 9 months in
2006. Relationship analysis of clinical isolates was done via PFGE,
multilocus sequence typing, plasmid profiling and additionally PCR for
bla(ESBL) detection and determination of phylogroups. After conjugal
transfer, plasmid isolation and subsequent PCR for bla(ESBL) detection and
determination of incompatibility groups were performed. Using one-primer
walking, up to 3600 bp upstream and downstream of different bla(CTX-M)
genes could be sequenced. beta-Lactamases found were TEM-1 (n=14), SHV-5
(n=1) and a wide variety of CTX-M types (n=21), i.e. CTX-M-15 (n=12),
CTX-M-1 (n=4), CTX-M-14 (n=2), CTX-M-9 (n=1), CTX-M-3 (n=1) and one new
type, CTX-M-65 (n=1). In 18 isolates, bla(ESBL) genes were located on
conjugative plasmids of sizes between 40 and 180 kbp belonging to
incompatibility groups FII (n=9), N (n=5) and I1 (n=4). bla(CTX-M) was
found to be associated with the common elements ISEcp1, IS26 and IS903-D,
but with unusual spacer sequences for ISEcp1 in two isolates. These
insertion sequences, connected to bla(CTX-M) as well as other genes, were
located between two IS26 elements in a configuration that has not yet been
described. The results reveal the emergence of bla(ESBL), predominantly
bla(CTX-M), located on different plasmids harboured by genotypically
different E. coli strains. The identical gene arrangement in the
bla(CTX-M) neighbourhood in plasmids of different incompatibility groups
indicates a main role of IS26 in distribution of mobile resistance
elements between different plasmids.

<>

<1>Cummings, D.J., McNally, K.L., Domenico, J.M., Matsuura, E.T.
<2>The complete DNA sequence of the mitochondrial genome of Podospora anserina.
<3>Curr. Genet.
<4>17
<5>375-402
<6>1990
<7>The complete 94,192 bp sequence of the mitochondrial genome from race s of
Podospora anserina is presented (1 kb = 10(3) base pairs). Three regions
unique to race A are also presented bringing the size of this genome to
100,314 bp. Race s contains 31 group I introns (33 in race A) and 2 group
II introns (3 in race A). Analysis shows that the group I introns can be
categorized according to families both with regard to secondary structure
and their open reading frames. All identified genes are transcribed from
the same strand. Except for the lack of ATPase 9, the Podospora genome
contains the same genes as its fungal counterparts, N. crassa and A.
nidulans. About 20% of the genome has not yet been identified. DNA
sequence studies of several excision-amplification plasmids demonstrate a
common feature to be the presence of short repeated sequences at both
termini with a prevalence of GGCGCAAGCTC.

<>

<1>Cummings, M., Adams, R.L.P.
<2>Cloning the pea DNA methylase cDNA.
<3>Biochem. Soc. Trans.
<4>21
<5>7s
<6>1992
<7>Both eukaryotic and prokaryotic cytosine methylases catalyse the transfer of a methyl group
from S-adenosyl methionine to the 5-position of the cytosine ring in DNA. The cDNA coding for
mouse and human methylase have been previously cloned. Comparison of the encoded proteins with
prokaryotic type II cytosine methylase sequences shows that the carboxy terminal one-third of
the eukaryotic enzymes shares significant homology with the prokaryotic enzymes. The
arrangement of several sequence motifs has been preserved in the eukaryotic proteins. This
includes a short amino acid sequence containing a pro-cys dipeptide which has been proved to
the the catalytic center of at least one bacterial type II methylase.

<>

<1>Cunnac, S., Bolot, S., Forero, S.N., Ortiz, E., Szurek, B., Noel, L.D., Arlat, M., Jacques, M.A., Gagnevin, L., Carrere, S., Nicole, M., Koebnik, R.
<2>High-Quality Draft Genome Sequences of Two Xanthomonas citri pv. malvacearum Strains.
<3>Genome Announcements
<4>1
<5>e00674-13
<6>2013
<7>We report high-quality draft genome sequences of two strains (race 18 and 20) of  Xanthomonas
citri pv. malvacearum, the causal agent of bacterial blight of
cotton. Comparative genomics will help to decipher mechanisms provoking disease
and triggering defense responses and to develop new molecular tools for
epidemiological surveillance.

<>

<1>Cuppels, D.A., Van Etten, J.L., Lambrecht, P., Vidaver, A.K.
<2>Survey of phytopathogenic pseudomonads for a restriction and modification system active on the double-stranded ribonucleic acid phage Phi-6.
<3>Curr. Microbiol.
<4>5
<5>247-249
<6>1981
<7>A total of 380 pseudomonad strains from 39 nomenspecies and 41 strains from 7 other bacterial
genera were screened for a double-stranded ribonucleic acid modification and restriction
system using the double-stranded ribonucleic acid modification and restriction system using
the double-stranded ribonucleic acid bacteriophage Phi-6.  Of these 421 strains, 8 showed the
low plating efficiency (10^-5 to 10^-7) characteristic of such a system.  However, the phage
propagated in 7 of the 8 were host-range mutants; the remaining strain showed some
characteristics of a host-modification system but the results were equivocal.

<>

<1>Curcio, M.J., Belfort, M.
<2>Retrohoming: cDNA-mediated mobility.
<3>Cell
<4>84
<5>9-12
<6>1996
<7>Last year was a vintage year for mobile group II introns.  In 1995 we moved from phenomenology
to mechanistic insight, from enigmatic observations to a coherent appreciation of process.
The finding that nucleated our understanding of the group II intron mobility event was the
appearance of an extraordinary double-strand break in the target DNA: a break that provides an
initiation site for reverse transcriptase, which mediates group II intron mobility; a break in
which the excised intron RNA is covalently attached to one of the DNA ends; a break that is
made by the protein and RNA products of the intron itself, with the RNA believed to be the
catalyst responsible for one of the DNA strand cleavages.  In this minireview, we piece
together the puzzle by describing the formation of this remarkable double-strand break, its
role in group II intron mobility, and the evolutionary implications of the process.  Group II
introns are catalytic RNAs that, like spliceosomal introns, splice via a lariat intermediate.
The nuclear pre-mRNA introns are thought to be descended from the group II introns, which are
found in both pro- and eukaryotes.  The functional domains of the self-splicing group II
introns are suspected to have evolved to act in trans under the guise of the snRNAs.  The
mobile group II introns are also phylogenetically linked to retroelements, which may provide
clues as to how the group II introns have become disseminated.  The mobility of group II
introns therefore engenders great evolutionary and mechanistic interest.

<>

<1>Cymerman, I.A., Obarska, A., Skowronek, K.J., Lubys, A., Bujnicki, J.M.
<2>Identification of a new subfamily of HNH nucleases and experimental characterization of a representative member, HphI restriction endonuclease.
<3>Proteins
<4>65
<5>867-876
<6>2006
<7>The restriction endonuclease (REase) R. HphI is a Type IIS enzyme that recognizes the
asymmetric target DNA sequence 5'-GGTGA-3' and in the
presence of Mg2+ hydrolyzes phosphodiester bonds in both strands of the
DNA at a distance of 8 nucleotides towards the 3' side of the target,
producing a 1 nucleotide T-staggered cut in an unspecified sequence at
this position. REases are typically ORFans that exhibit little
similarity to each other and to any proteins in the database. However,
bioinformatics analyses revealed that R.HphI is a member of a
relatively big sequence family with a conserved C-terminal domain and a
variable N-terminal domain. We predict that the C-terminal domains of
proteins from this family correspond to the nuclease domain of the HNH
superfamily rather than to the most common PD(D/E)XK superfamily of
nucleases. We constructed a three-dimensional model of the R.HphI
catalytic domain and validated our predictions by sitedirected
mutagenesis and studies of DNA-binding and catalytic activities of the
mutant proteins. We also analyzed the genomic neighborhood of R.HphI
homologs and found that putative nucleases accompanied by a DNA
methyltransferase (i.e. predicted REases) do not form a single group on
a phylogenetic tree, but are dispersed among free-standing putative
nucleases. This suggests that nucleases from the HNH superfamily were
independently recruited to become REases in the context of RM systems
multiple times in the evolution and that members of the HNH superfamily
may be much more frequent among the so far unassigned REase sequences
than previously thought.

<>

<1>Czajkowski, R., van der Wolf, J.M.
<2>Draft Genome Sequence of the Biocontrol Strain Serratia plymuthica A30, Isolated  from Rotting Potato Tuber Tissue.
<3>J. Bacteriol.
<4>194
<5>6999-7000
<6>2012
<7>Serratia plymuthica A30 is a Gram-negative bacterium expressing antagonistic activity toward
blackleg- and soft rot-causing Dickeya sp. biovar 3 ('Dickeya
solani'). Here, we present the draft genome sequence of strain A30, which has
been isolated from rotten potato tuber tissue.

<>

<1>Czank, A., Hauselmann, R., Page, A.W., Leonhardt, H., Bestor, T.H., Schafner, W., Hergersberg, M.
<2>Expression in mammalian cells of a cloned gene encoding murine DNA methyltransferase.
<3>Gene
<4>109
<5>259-263
<6>1991
<7>Mammalian DNA cytosine-5-methyltransferase (MTase, EC 2.1.1.37) is an essential
component for establishing and maintaining cell-type specific methylation
patterns in the genome.  The cDNA for the murine enzyme was previously cloned
in segments.  We have reconstructed the entire gene, encoding a protein of 1517
amino acids, from a set of overlapping cDNA clones.  We report the assembly of
two expression constructs in bacterial/mammalian shuttle vectors.
Transcription in the first construct (pEMT) is driven by the cytomegalovirus
enhancer/promoter and encodes a fusion protein with 15 additional aa at the N
terminus, while the second construct (pJMT) is driven by the simian virus 40
early promoter/enhancer upstream from the natural ATG codon.
Immunofluorescence microscopy and immunoblot analysis have shown that both
constructs direct the synthesis of MTase in COS-1 cells.  Enzyme activity in
whole-cell lysates of transfected COS-1 cells transfected with pEMT and pJMT
are on average tenfold and fivefold higher than in controls, respectively.  The
specific activities of the recombinant and endogenous mouse-cell enzyme are
similar.  These expression constructs will be of use in studies of DNA
methylation in mammals.

<>

<1>D'Afonseca, V., Prosdocimi, F., Dorella, F.A., Pacheco, L.G., Moraes, P.M., Pena, I., Ortega, J.M., Teixeira, S., Oliveira, S.C., Coser, E.M., Oliveira, L.M., Correa-de-Oliveira, G., Meyer, R., Miyoshi, A., Azevedo, V.
<2>Survey of genome organization and gene content of Corynebacterium pseudotuberculosis.
<3>Microbiol. Res.
<4>165
<5>312-320
<6>2010
<7>Corynebacterium pseudotuberculosis is an intracellular pathogen that
causes Caseous lymphadenitis (CLA) disease in sheep and goats. The
widespread occurrence and the economic importance of this pathogen have
prompted investigation of its pathogenesis. We used a genomic library of
C. pseudotuberculosis to generate 1440 genomic survey sequences (GSSs);
these were analyzed in silico with bioinformatics tools, using public
databases for comparative analyses. We employed non-redundant unique
sequences as a query for BLAST searches against the genome, the translated
genome and the proteome of four other Corynebacterium species that have
been completely sequenced. We were able to characterize approximately 8%
of the genome of C. pseudotuberculosis, including previously undescribed
functional group genes, based on the COG database; the GSSs classification
into categories gave 13% information storage and processing, 14% cellular
processes and 23% metabolism. We found a close relation between C.
pseudotuberculosis and C. diphtheriae conserved-gene synteny in
Corynebacteria species.

<>

<1>D'Arcy, A., Brown, R.S., Zabeau, M., van Resandt, R.W., Winkler, F.K.
<2>Purification and crystallization of the EcoRV restriction endonuclease.
<3>J. Biol. Chem.
<4>260
<5>1987-1990
<6>1985
<7>The type II restriction endonuclease EcoRV purified from a genetically
engineered, overproducing strain has been crystallized.  Four crystal forms all
obtained by precipitation with polyethylene glycol 4000 have been
characterized.  Two of these are suitable for high resolution structure
analysis.  Both are orthorhombic, have space group P2/12/1/2/1 and have similar
unit cell dimensions of a = 58.2 angstrom, b = 71.7 angstrom, c = 130.6
angstrom (form A) and a = 59.9 angstrom, b = 74.5 angstrom, c= 121.8 angstrom
(form B).  They diffract to about 2 angstrom resolution and appear to have one
dimer of 2 x 29,000 daltons in the asymmetric unit.

<>

<1>D'Argenio, V., Petrillo, M., Cantiello, P., Naso, B., Cozzuto, L., Notomista, E., Paolella, G., Di Donato, A., Salvatore, F.
<2>De Novo Sequencing and Assembly of the Whole Genome of Novosphingobium sp. Strain PP1Y.
<3>J. Bacteriol.
<4>193
<5>4296
<6>2011
<7>Novosphingobium sp. strain PP1Y is a marine bacterium specifically adapted to use fuels as an
energy source. We sequenced and assembled its entire
genome using the Roche 454 genome sequencer system, which led to the
identification of two plasmids and one megaplasmid, besides a 3.9-Mb
circular chromosome.

<>

<1>D'Auria, G., Dzunkova, M., Moya, A., Tomaska, M., Kolosta, M., Kmet, V.
<2>Genome Sequence of Lactobacillus plantarum 19L3, a Strain Proposed as a Starter Culture for Slovenska Bryndza Ovine Cheese.
<3>Genome Announcements
<4>2
<5>e00292-14
<6>2014
<7>The genome sequence of Lactobacillus plantarum isolated from ovine cheese is presented here.
This bacterium is proposed as a starter strain, named 19L3, for Slovenska bryndza cheese, a
traditional Slovak cheese fulfilling European Food Safety Authority (EFSA) requirements.

<>

<1>D'Auria, G., Galan, J.C., Rodriguez-Alcayna, M., Moya, A., Baquero, F., Latorre, A.
<2>Complete Genome Sequence of Acidaminococcus intestini RYC-MR95, a Gram-Negative Bacterium from the Phylum Firmicutes.
<3>J. Bacteriol.
<4>193
<5>7008-7009
<6>2011
<7>Acidaminococcus intestini belongs to the family Acidaminococcaceae, order Selenomonadales,
class Negativicutes, phylum Firmicutes. Negativicutes
show the double-membrane system of Gram-negative bacteria, although their
chromosomal backbone is closely related to that of Gram-positive bacteria
of the phylum Firmicutes. The complete genome of a clinical A. intestini
strain is here presented.

<>

<1>D'Elia, J., Stoddard, S.
<2>Ketogulonigenium endogenous plasmids.
<3>US Patent Office
<4>US 7053196 B
<5>
<6>2006
<7>The present invention relates, in general, to a genus of bacteria known as Ketogulonigenium.
The present invention further relates to transformed Ketogulonigenium, and methods of
transforming Ketogulonigenium.  The present invention also relates to nucleic acid molecules,
and vectors.

<>

<1>D'Elia, J., Stoddard, S.F.
<2>Ketogulonigenium endogenous plasmids.
<3>US Patent Office
<4>US 7053197 A
<5>
<6>2006
<7>
<>

<1>D'Elia, J., Stoddard, S.F.
<2>Ketogulonigenium endogeneous plasmids.
<3>US Patent Office
<4>US 7030233 A
<5>
<6>2006
<7>The present invention relates, in general, to a genus of bacteria known as Ketogulonigenium.
The present invention further relates to transformed Ketogulonigenium, and methods of
transforming Ketogulonigenium.  The present invention also relates to nucleic acid molecules,
and vectors.

<>

<1>D'Halluin, K., Ruiter, R.
<2>Methods and means for removal of a selected dna sequence.
<3>International Patent Office
<4>WO 2006105946 A
<5>
<6>2006
<7>A method is described for the exact removal of a selected subfragment from a DNA molecule by
intrachromosomal recombination between two directly repeated DNA sequences using a
rate-cleaving double stranded break inducing DNA endonuclease expressed under control of a
micro-spore specific promoter.  This method can be applied in a method for the exact exchange
of a target DNA fragment for a DNA fragment of interest in plant cells and plants.

<>

<1>D'Souza, D.R., Morgan, R.D., Parashar, V., Capalash, N., Sharma, P.
<2>Characterization of BflI - a thermostable, Co++-requiring isoschizomer of BsiYI from Anoxybacillus flavithermus.
<3>World J. Microbiol. Biotechnol.
<4>20
<5>593-598
<6>2004
<7>A thermophile, isolated from geothermal areas in the northern Himalayan region of India, was
identified by partial 16S rDNA sequence (GenBank
accession # AF482430) analysis as Anoxybacillus flavithermus. The
isolate produced BflI (REBASE # 4910), a Type II restriction
endonuclease, which recognized the sequence 5'-CCNNNN-N/NNGG-3' and was
the isoschizomer of BsiYI. The enzyme was purified to homogeneity by
passing through Cibacron Blue F3GA agarose, DEAE-cellulose,
heparin-agarose and MonoQ FPLC. The purified enzyme (MW 36 kDa) worked
best at 60 C in Promega's buffer C and preferentially required
Co++(0.4 mM) as cofactor followed by Mg++ (10 mM) and Mn++ (1 mM). The
enzyme showed high specific activity and worked in the presence of high
concentrations of beta-mercaptoethanol (200 mM), Triton-X-100 (25%),
urea (30%), formamide (6%) and guanidine (40 mM) and showed no star
activity in the presence of 40% glycerol. In the absence of any
stabilizing agent, BflI retained t(1/2) for at least 96 h at 37
degreesC, 6 h at 60 C and 6 months at 4 C. N-terminal
sequencing showed that its first 10 amino acid residues were
DFHEDKTIAR.

<>

<1>da Piedade, I., Skive, B., Christensen, H., Bojesen, A.M.
<2>Draft Genome Sequence of Streptococcus equi subsp. zooepidemicus Strain S31A1, Isolated from Equine Infectious Endometritis.
<3>Genome Announcements
<4>1
<5>e00683-13
<6>2013
<7>We present the draft genome sequence of Streptococcus equi subsp. zooepidemicus S31A1, a
strain isolated from equine infectious endometritis in Denmark.
Comparative analyses of this genome were done with four published reference
genomes: S. zooepidemicus strains MGCS10565, ATCC 35246, and H70 and S. equi
subsp. equi strain 4047.

<>

<1>da Silva, A.C.R. et al.
<2>Comparison of the genomes of two Xanthomonas pathogens with differing host specificities.
<3>Nature
<4>417
<5>459-463
<6>2002
<7>The genus Xanthomonas is a diverse and economically important group of bacterial
phytopathogens, belonging to the gamma-subdivision of the Proteobacteria. Xanthomonas
axonopodis pv. citri (Xac) causes citrus canker, which affects most commercial citrus
cultivars, resulting in significant losses worldwide. Symptoms include canker lesions, leading
to abscission of fruit and leaves and general tree decline. Xanthomonas campestris pv.
campestris (Xcc) causes black rot, which affects crucifers such as Brassica and Arabidopsis.
Symptoms include marginal leaf chlorosis and darkening of vascular tissue, accompanied by
extensive wilting and necrosis. Xanthomonas campestris pv. campestris is grown commercially to
produce the exopolysaccharide xanthan gum, which is used as a viscosifying and stabilizing
agent in many industries. Here we report and compare the complete genome sequences of Xac and
Xcc. Their distinct disease phenotypes and host ranges belie a high degree of similarity at
the genomic level. More than 80% of genes are shared, and gene order is conserved along most
of their respective chromosomes. We identified several groups of strain-specific genes, and on
the basis of these groups we propose mechanisms that may explain the differing host
specificities and pathogenic processes.

<>

<1>Dabe, E.C., Kohn, A.B., Bobkova, Y., Kocot, K., Citarella, M., Bostwick, C.J., Winters, G.C., Swalla, B.J., Moroz, L.L.
<2>Epigenomic Signatures in Basal Metazoans: DNA Methyltransferase in Pleurobrachia bachei.
<3>Integr. Comp. Biol.
<4>53
<5>E273
<6>2013
<7>DNA methylation is an epigenetic modification crucial to cell differentiation and development.
In the majority of bilaterians 5-methylcytosine DNA methylation occurs at CpG sites and
islands controlling gene transcription.  Contrary to Drosophila and C. elegans that have lost
this machinery, possibly due to their compact genome sizes and short life cycle, here we show
that the phylum Ctenophora has conserved methylation machinery.  Using the data from the
recently sequenced genome of Pleurobrachia bachei we cloned DNA 5-cytosine methyltransferase
(DNMT) and characterized its expression in major developmental stages and adult ctenophores.
Distinctive mRNA expression in the digestive system, (stomach, pharynx and mouth), tentacles
and unique patterns in between ciliated comb rows in adult Pleurobrachia collectively suggest
that DNMT mRNA expression levels are both cell-specific and noticeable in areas of high
proliferation.  Next using colorimetric ELISA assay for methylated DNA we directly showed that
DNA methylation does occur in the Pleurobrachia genome, although it was significantly lower
than in the molluscan (Aplysia) and mammalian (Ratus) nervous tissues. Combined, our data
suggest that the small genome of the ctenophore Pleurobrachia bachei has functional DNA
methylation machinery, possibly involved in epigenetic control of somatic cell divisions and
regulation of mRNA expression at zones of proliferation.

<>

<1>Daboussi, F. et al.
<2>Chromosomal context and epigenetic mechanisms control the efficacy of genome editing by rare-cutting designer endonucleases.
<3>Nucleic Acids Res.
<4>40
<5>6367-6379
<6>2012
<7>The ability to specifically engineer the genome of living cells at precise locations using
rare-cutting designer endonucleases has broad
implications for biotechnology and medicine, particularly for
functional genomics, transgenics and gene therapy. However, the
potential impact of chromosomal context and epigenetics on designer
endonuclease-mediated genome editing is poorly understood. To address
this question, we conducted a comprehensive analysis on the efficacy of
37 endonucleases derived from the quintessential I-CreI meganuclease
that were specifically designed to cleave 39 different genomic targets.
The analysis revealed that the efficiency of targeted mutagenesis at a
given chromosomal locus is predictive of that of homologous gene
targeting. Consequently, a strong genome-wide correlation was apparent
between the efficiency of targeted mutagenesis (0.1% to similar to 6%)
with that of homologous gene targeting (0.1% to similar to 15%). In
contrast, the efficiency of targeted mutagenesis or homologous gene
targeting at a given chromosomal locus does not correlate with the
activity of individual endonucleases on transiently transfected
substrates. Finally, we demonstrate that chromatin accessibility
modulates the efficacy of rare-cutting endonucleases, accounting for
strong position effects. Thus, chromosomal context and epigenetic
mechanisms may play a major role in the efficiency rare-cutting
endonuclease-induced genome engineering.

<>

<1>Dabrazhynetskaya, A., Soika, V., Volokhov, D., Simonyan, V., Chizhikov, V.
<2>Genome Sequence of Mycoplasma hyorhinis Strain DBS 1050.
<3>Genome Announcements
<4>2
<5>e00127-14
<6>2014
<7>Mycoplasma hyorhinis is known as one of the most prevalent contaminants of mammalian cell and
tissue cultures worldwide. Here, we present the complete
genome sequence of the fastidious M. hyorhinis strain DBS 1050.

<>

<1>Dabul, A.N., Kos, V.N., Gilmore, M.S., Camargo, I.L.
<2>Draft Genome Sequence of Methicillin-Resistant Staphylococcus aureus Strain SA16, Representative of an Endemic Clone from a Brazilian Hospital.
<3>Genome Announcements
<4>1
<5>e00754-13
<6>2013
<7>Here we report the draft genome sequence of a bloodstream isolate of methicillin-resistant
Staphylococcus aureus strain SA16. Strain SA16 is a
sequence type 5 (ST5)-staphylococcal cassette chromosome mec type II (SCCmec II)
clone and was the most prevalent isolate at a Brazilian hospital during the
second half of 2009.

<>

<1>Daccord, A., Ceccarelli, D., Burrus, V.
<2>Integrating conjugative elements of the SXT/R391 family trigger the excision and drive the mobilization of a new class of Vibrio genomic islands.
<3>Mol. Microbiol.
<4>78
<5>576-588
<6>2010
<7>In vibrios and enterobacteria lateral gene transfer is often facilitated by
integrating conjugative elements (ICEs) of the SXT/R391 family. SXT/R391 ICEs
integrate by site-specific recombination into prfC and transfer by conjugation, a
process that is initiated at a specific locus called the origin of transfer
(oriT(SXT) ). We identified genomic islands (GIs) harbouring a sequence that
shares >63% identity with oriT(SXT) in three species of Vibrio. Unlike SXT/R391
ICEs, these GIs are integrated into a gene coding for a putative stress-induced
protein and do not appear to carry any gene coding for a conjugative machinery or
for mobilization proteins. Our results show that SXT/R391 ICEs trigger the
excision and mediate the conjugative transfer in trans of the three Vibrio GIs at
high frequency. GIs' excision is independent of the ICE-encoded recombinase and
is controlled by the ICE-encoded transcriptional activator SetCD, which is
expressed during the host SOS response. Both mobI and traI, two ICE-borne genes
involved in oriT recognition, are essential for GIs' transfer. We also found that
SXT/R391 ICEs mobilize in trans over 1 Mb of chromosomal DNA located 5' of the
GIs' integration site. Together these results support a novel mechanism of
mobilization of GIs by ICEs of the SXT/R391 family.

<>

<1>Dahai, T., Ando, S., Takasaki, Y., Tadano, J.
<2>Site-directed mutagenesis of restriction endonuclease HindIII.
<3>Biosci. Biotechnol. Biochem.
<4>63
<5>1703-1707
<6>1999
<7>Site-directed mutagenesis by inverse PCR was done on the HindIII gene.  Target residues to be
mutated were chosen according to (i) the fact that a mutant obtained by sodium nitrite
treatment showed almost no HindIII activity, where Asp-123 was replaced with Asn, and (ii) the
model proposed by Stahl et al.  Seven kinds of mutants were obtained by PCR, and their
enzymatic and biochemical properties were examined.  Three mutants, P50S, D108L, and D123N,
showed fairly low HindIII activity.  On the other hand, the other four, P84Q, E85K, V106E, and
K125N, retained the activity.  In particular, E86K showed higher activity than the wild type
enzyme.  This fact was confirmed when activities of the purified wild type and E86K enzymes
were assayed.  These results coincided fairly well with data using E. coli strains that carry
the respective mutant plasmids, on their resistance to phage T7 and on growth rate.  We
conclude that the PE motif at residues 50 and 51, and DXK motif at residues 108-110, are
responsible for the enzymic reaction of HindIII.

<>

<1>Dahlem, T.J., Hoshijima, K., Jurynec, M.J., Gunther, D., Starker, C.G., Locke, A.S., Weis, A.M., Voytas, D.F., Grunwald, D.J.
<2>Simple Methods for Generating and Detecting Locus-Specific Mutations Induced with TALENs in the Zebrafish Genome.
<3>PLoS Genet.
<4>8
<5>E1002861
<6>2012
<7>The zebrafish is a powerful experimental system for uncovering gene function in
vertebrate organisms. Nevertheless, studies in the zebrafish have been limited by
the approaches available for eliminating gene function. Here we present simple
and efficient methods for inducing, detecting, and recovering mutations at
virtually any locus in the zebrafish. Briefly, double-strand DNA breaks are
induced at a locus of interest by synthetic nucleases, called TALENs. Subsequent
host repair of the DNA lesions leads to the generation of insertion and deletion
mutations at the targeted locus. To detect the induced DNA sequence alterations
at targeted loci, genomes are examined using High Resolution Melt Analysis, an
efficient and sensitive method for detecting the presence of newly arising
sequence polymorphisms. As the DNA binding specificity of a TALEN is determined
by a custom designed array of DNA recognition modules, each of which interacts
with a single target nucleotide, TALENs with very high target sequence
specificities can be easily generated. Using freely accessible reagents and
Web-based software, and a very simple cloning strategy, a TALEN that uniquely
recognizes a specific pre-determined locus in the zebrafish genome can be
generated within days. Here we develop and test the activity of four TALENs
directed at different target genes. Using the experimental approach described
here, every embryo injected with RNA encoding a TALEN will acquire targeted
mutations. Multiple independently arising mutations are produced in each growing
embryo, and up to 50% of the host genomes may acquire a targeted mutation. Upon
reaching adulthood, approximately 90% of these animals transmit targeted
mutations to their progeny. Results presented here indicate the TALENs are highly
sequence-specific and produce minimal off-target effects. In all, it takes about
two weeks to create a target-specific TALEN and generate growing embryos that
harbor an array of germ line mutations at a pre-specified locus.

<>

<1>Dai, H., He, Y., Linsley, P.S., Mao, M., Roberts, C.J., Van't-Veer, L.J., Van de Vijver, M.J., Bernards, R., Hart, A.A.M.
<2>Methods of assigning treatment to breast cancer patients.
<3>US Patent Office
<4>US 7171311 A
<5>
<6>2007
<7>The present invention relates to genetic markers whose expression is correlated with breast
cancer.  Specifically, the invention provides sets of markers whose expression patterns can be
used to differentiate clinical conditions associated with breast cancer, such as the presence
or absence of the estrogen receptor ESR1, and BRCA1 and sporadic tumors, and to provide
information on the likelihood of tumor distant metasteses within five years of initial
diagnosis.  The invention relates to methods of using these markers to distinguish these
conditions.  The invention also provides methods of classifying and treating patients based on
prognosis.  The invention also relates to kits containing ready-to-use microarrays and
computer software for data analysis using the diagnostic, prognostic and statistical methods
disclosed herein.

<>

<1>Dai, J., Wang, S., Guerlebeck, D., Laturnus, C., Guenther, S., Shi, Z., Lu, C., Ewers, C.
<2>Suppression subtractive hybridization identifies an autotransporter adhesin gene  of E. coli IMT5155 specifically associated with avian pathogenic Escherichia coli  (APEC).
<3>BMC Microbiol.
<4>10
<5>236
<6>2010
<7>BACKGROUND: Extraintestinal pathogenic E. coli (ExPEC) represent a phylogenetically diverse
group of bacteria which are implicated in a large range
of infections in humans and animals. Although subgroups of different ExPEC
pathotypes, including uropathogenic, newborn meningitis causing, and avian
pathogenic E. coli (APEC) share a number of virulence features, there still might
be factors specifically contributing to the pathogenesis of a certain subset of
strains or a distinct pathotype. Thus, we made use of suppression subtractive
hybridization and compared APEC strain IMT5155 (O2:K1:H5; sequence type complex
95) with human uropathogenic E. coli strain CFT073 (O6:K2:H5; sequence type
complex 73) to identify factors which may complete the currently existing model
of APEC pathogenicity and further elucidate the position of this avian pathotype
within the whole ExPEC group. RESULTS: Twenty-eight different genomic loci were
identified, which are present in IMT5155 but not in CFT073. One of these loci
contained a gene encoding a putative autotransporter adhesin. The open reading
frame of the gene spans a 3,498 bp region leading to a putative 124-kDa adhesive
protein. A specific antibody was raised against this protein and expression of
the adhesin was shown under laboratory conditions. Adherence and adherence
inhibition assays demonstrated a role for the corresponding protein in adhesion
to DF-1 chicken fibroblasts. Sequence analyses revealed that the flanking regions
of the chromosomally located gene contained sequences of mobile genetic elements,
indicating a probable spread among different strains by horizontal gene transfer.
In accordance with this hypothesis, the adhesin was found to be present not only
in different phylogenetic groups of extraintestinal pathogenic but also of
commensal E. coli strains, yielding a significant association with strains of
avian origin. CONCLUSIONS: We identified a chromosomally located autotransporter
gene in a highly virulent APEC strain which confers increased adherence of a
non-fimbriated E. coli K-12 strain to a chicken fibroblast cell line. Even though
flanked by mobile genetic elements and three different genetic regions upstream
of the gene, most probably indicating horizontal gene transfer events, the
adhesin gene was significantly linked with strains of avian origin. Due to the
nucleotide sequence similarity of 98% to a recently published adhesin-related
gene, located on plasmid pAPEC-O1-ColBM, the name aatA (APEC autotransporter
adhesin A) was adopted from that study.Our data substantiate that AatA might not
only be of relevance in APEC pathogenicity but also in facilitating their
reservoir life style in the chicken intestine, which might pave the way for
future intestinal preventive strategies.

<>

<1>Dai, L., Chai, D., Gu, S.Q., Gabel, J., Noskov, S.Y., Blocker, F.J., Lambowitz, A.M., Zimmerly, S.
<2>A Three-Dimensional Model of a Group II Intron RNA and Its Interaction with the Intron-Encoded Reverse Transcriptase.
<3>Mol. Cell
<4>30
<5>472-485
<6>2008
<7>Group II introns are self-splicing ribozymes believed to be the ancestors of spliceosomal
introns. Many group II introns encode reverse
transcriptases that promote both RNA splicing and intron mobility to new
genomic sites. Here we used a circular permutation and crosslinking method
to establish 16 intramolecular distance relationships within the mobile
Lactococcus lactis Ll.LtrB-DeltaORF intron. Using these new constraints
together with 13 established tertiary interactions and eight published
crosslinks, we modeled a complete three-dimensional structure of the
intron. We also used the circular permutation strategy to map RNA-protein
interaction sites through fluorescence quenching and crosslinking assays.
Our model provides a comprehensive structural framework for understanding
the function of group II ribozymes, their natural structural variations,
and the mechanisms by which the intron-encoded protein promotes RNA
splicing and intron mobility. The model also suggests an arrangement of
active site elements that may be conserved in the spliceosome.

<>

<1>Dai, Q., Restrepo, B.I., Porcella, S.F., Raffel, S.J., Schwan, T.G., Barbour, A.G.
<2>Antigenic variation by Borrelia hermsii occurs through recombination between extragenic repetitive elements on linear plasmids.
<3>Mol. Microbiol.
<4>60
<5>1329-1343
<6>2006
<7>The relapsing fever agent Borrelia hermsii undergoes multiphasic antigenic variation through
gene conversion of a unique expression site on a linear plasmid by an archived variable
antigen gene. To further characterize this mechanism we assessed the repertoire and
organization of archived variable antigen genes by sequencing approximately 85% of plasmids
bearing these genes. Most archived genes shared with the expressed gene a less than or equal
62 nucleotide (nt) region, the upstream homology sequence (UHS), that surrounded the start
codon. The 59 archived variable antigen genes were arrayed in clusters with 13 repetitive, 214
nt long downstream homology sequence (DHS) elements distributed among them. A fourteenth DHS
element was downstream of the expression locus. Informative nucleotide polymorphisms in UHS
regions and DHS elements were applied to the analysis of the expression site of relapse
serotypes from 60 infected mice in a prospective study. For most recombinations, the upstream
crossover occurred in the UHS's second half, and the downstream crossover was in the DHS's
second half. Usually the closest archival DHS element was used, but occasionally a more
distant DHS was employed. The downstream extragenic crossover site in B. hermsii contrasts
with the downstream extragenic crossover site for antigenic variation in African trypanosomes.

<>

<1>Dai, Y., Ni, Z.F., Dai, J., Zhao, T., Sun, Q.X.
<2>Isolation and expression analysis of genes encoding DNA methyltransferase in wheat (Triticum aestivum L.).
<3>Biochim. Biophys. Acta
<4>1729
<5>118-125
<6>2005
<7>DNA methylation of cytosine residues, catalyzed by DNA methyltransferases, is suggested to
play important roles in regulating
gene expression and plant development. In this study, we isolated four
wheat cDNA fragments and one cDNA with open reading frame encoding
putative DNA methyltransferase and designated TaMET1, TaMET2a, TaMET2b,
TaCMT, TaMET3, respectively. BLASTX searches and phylogenetic analysis
suggested that five cDNAs belonged to four classes (Dnmt1, Dnmt2, CMT
and Dnmt3) of DNA methyltransferase genes. TaMET2a encoded a protein of
376 aa and contained eight of ten conserved motifs characteristic of
DNA methyltransferase. Genomic sequence of TaMET2a was obtained and
found to contain ten introns and eleven exons. The expression analysis
of the five genes revealed that they were expressed in developing seed,
during germination and various vegetative tissues, but in quite
different abundance. It was interesting to note that TaMET1 and TaMET3
mRNAs were clearly detected in dry seeds. Moreover, the differential
expression patterns of five genes were observed between wheat hybrid
and its parents in leaf, stem and root of jointing stage, some were
up-regulated while some others were down-regulated in the hybrid. We
concluded that multiple wheat DNA methyltransferase genes were present
and might play important roles in wheat growth and development.

<>

<1>Daifuku, T., Yoshida, T., Kitamura, T., Kawaichi, S., Inoue, T., Nomura, K., Yoshida, Y., Kuno, S., Sako, Y.
<2>Variation of the Virus-Related Elements within Syntenic Genomes of the Hyperthermophilic Archaeon Aeropyrum.
<3>Appl. Environ. Microbiol.
<4>79
<5>5891-5898
<6>2013
<7>The increasing number of genome sequences of archaea and bacteria show their
adaptation to different environmental conditions at the genomic level. Aeropyrum
spp. are aerobic and hyperthermophilic archaea. Aeropyrum camini was isolated
from a deep-sea hydrothermal vent, and Aeropyrum pernix was isolated from a
coastal solfataric vent. To investigate the adaptation strategy in each habitat,
we compared the genomes of the two species. Shared genome features were a small
genome size, a high GC content, and a large portion of orthologous genes (86 to
88%). The genomes also showed high synteny. These shared features may have been
derived from the small number of mobile genetic elements and the lack of a RecBCD
system, a recombinational enzyme complex. In addition, the specialized physiology
(aerobic and hyperthermophilic) of Aeropyrum spp. may also contribute to the
entire-genome similarity. Despite having stable genomes, interference of synteny
occurred with two proviruses, A. pernix spindle-shaped virus 1 (APSV1) and A.
pernix ovoid virus 1 (APOV1), and clustered regularly interspaced short
palindromic repeat (CRISPR) elements. Spacer sequences derived from the A. camini
CRISPR showed significant matches with protospacers of the two proviruses
infecting A. pernix, indicating that A. camini interacted with viruses closely
related to APSV1 and APOV1. Furthermore, a significant fraction of the
nonorthologous genes (41 to 45%) were proviral genes or ORFans probably
originating from viruses. Although the genomes of A. camini and A. pernix were
conserved, we observed nonsynteny that was attributed primarily to virus-related
elements. Our findings indicated that the genomic diversification of Aeropyrum
spp. is substantially caused by viruses.

<>

<1>Daigle, K., Shenoy, S., Ehrlich, K., Gehrke, C., Ehrlich, M.
<2>Restriction at mismatched sites in DNA.
<3>Fed. Proc.
<4>45
<5>1783
<6>1986
<7>Restriction endonucleases were tested for their ability to catalyze the
cleavage of mismatch-containing recognition sites in DNA.  These mismatched
base pairs were T-G, U-G or A-C in covalently closed, circular DNA molecules
prepared by in vitro extension of chemically synthesized oligonucleotide
primers annealed to an M13-derived viral DNA.  None of the tested restriction
enzymes was able to completely cleave the mismatch-containing recognition sites
of these heteroduplexes at a normal rate.  However, three of them, SmaI, SalI,
SstI, partially digested certain T-G or U-G-substituted recognition sites under
standard digestion conditions.  In these digests, there was an accumulation of
DNA singly nicked at the mismatched recognition site.  The ability of SmaI and
SstI to partially cleave at a mismatch was shown to depend on the nature and
position of the mismatch within the corresponding recognition site.  In
contrast to such partial cleavage, little or no digestion was obtained with
AccI, HincII, HindIII, and KpnI at their corresponding tested
mismatch-containing recognition sites.  Therefore, a transition-type
substitution of only one strand of a recognition site can inhibit restriction
endonuclease-catalyzed digestion at that site.

<>

<1>Dailey, P., Kawa, D., Lu, S.D.
<2>Reagents and methods for detecting neisseria gonorrhoeae.
<3>European Patent Office
<4>EP 2050826 A
<5>
<6>2009
<7>This invention provides compositions and methods for detecting Neisseria gonorrhoeae in a
sample.  This invention also provides related reaction mixtures, kits, systems, and computers.

<>

<1>Daiyasu, H., Komori, K., Sakae, S., Ishino, Y., Toh, H.
<2>Hjc resolvase is a distantly related member of the type II restriction endonuclease family.
<3>Nucleic Acids Res.
<4>28
<5>4540-4543
<6>2000
<7>Hjc resolvase is an archaeal enzyme involved in homologous DNA recombination at the Holliday
junction intermediate. However, the structure and the catalytic mechanism of the enzyme have
not yet been identified.  We performed database searching using the amino acid sequence of the
enzyme from Pyrococcus furiosus as a query. We detected 59 amino acid sequences showing weak
but significant sequence similarity to the Hjc resolvase. The detected sequences included
DpnII, HaeII and Vsr endonuclease, which belong to the type II restriction endonuclease
family. In addition, a highly conserved region was identified from a multiple alignment of the
detected sequences, which was similar to an active site of the type II restriction
endonucleases. We substituted three conserved amino acid residues in the highly conserved
region of the Hjc resolvase with Ala residues. The amino acid replacements inactivated the
enzyme. The experimental study, together with the results of the database searching, suggests
that the Hjc resolvase is a distantly related member of the type II restriction endonuclease
family. In addition, the results of our database searches suggested that the members of the
RecB domain superfamily are evolutionarily related to the type II restriction endonuclease
family.

<>

<1>Dale, C.J.H., Moses, E.K., Ong, C.C., Morrow, C.J., Reed, M.B., Hasse, D., Strugnell, R.A.
<2>Identification and sequencing of the groE operon and flanking genes of Lawsonia intracellularis: use in phylogeny.
<3>Microbiology
<4>144
<5>2073-2084
<6>1998
<7>Proliferative enteropathy (PE) is a complex of diseases of commercial importance to the pig
industry. The obligate intracellular bacterium Lawsonia intracellularis is consistently
associated with PE and pure cultures of this bacterium have been used to reproduce PE in pigs.
In this study L. intracellularis bacteria were purified directly from PE-affected tissue. DNA
extracted from purified bacteria was used to construct a partial genomic library which was
screened using sera from L. intracellularis-immunized rabbits. Two seroreactive recombinant
clones were identified, one of which expressed proteins of 10 and 60 kDa. The sequence of the
insert from this clone, pISI-2, revealed ORFs with sequence similarity to the groES/EL operon
of Escherichia coli, the 505 ribosomal proteins L21 and L27 of E. coli, a GTP-binding protein
of Bacillus subtilis and a possible protoporphyrinogen oxidase, HemK, of E. coli. Primers
designed from unique sequences from the pISI-2 insert amplified DNA from infected, but not
non-infected, porcine ilea; the amplicon sequence obtained from tissue-cultured L.
intracellularis was identical to the corresponding sequence in pISI-2, confirming the origin
of the clone. The sequence of L. intracellularis GroEL and other GroEL sequences in the
databases were used to construct a partial phylogenetic tree. Analysis of the GroEL sequence
relationship suggested that L. intracellularis is not significantly related to other organisms
whose GroEL sequences are held in the databases and supports previous data from 16S sequence
analyses suggesting that L. intracellularis is a member of a novel group of enteric pathogens.

<>

<1>Dalgaard, J., Garrett, R.A., Kjems, J.
<2>Thermostable sequence-specific endonucleases.
<3>International Patent Office
<4>WO 9404663
<5>
<6>1994
<7>Thermostable sequence-specific DNA endonucleases are encoded by archaeal type introns of
stable RNA (ribosomal RNA or transfer RNA) or protein genes or are enzymatically active
variants thereof in which one or more amino acid residues have been deleted, inserted or
substituted by other amino acids. These endonucleases recognize relatively long sequences of
about 20 base pairs and are very rare cutters, cleaving with a frequency of about 1:5,000,000.
Thus, they are useful as endonuclease tools for gene analysis, such as genome mapping and
detection of major rearrangements in large genomes, and for gene manipulation, such as cloning
and chromosome targeting.

<>

<1>Dalgaard, J.Z.
<2>Mobile introns and inteins: friend or foe?
<3>Trends Genet.
<4>10
<5>306-307
<6>1994
<7>Since the discovery of introns, one question that has puzzled the scientific community has
been whether these genetic elements are of functional importance to their host organism. One
theory is that introns are selfish DNA elements whose mobility allows them to overcome
selection against them. This theory seems especially plausible in the case of mobile group I
introns, which encode site-specific DNA endonucleases and can invade genomes in a
site-specific manner via a double-stranded break repair (DSBR) mechanism. Interestingly, it
has recently been discovered that this mechanism also accounts for the mobility of archael
introns and inteins, introns that are spliced at the level of the protein, several of which
have been found in prokaryotic genomes. The discovery that this mechanism of mobility is not
unique to group I introns and that this family of genetic elements is not limited to the
genomes of bacteriophages and eurkaryotic nuclear and organelles revives the question of
whether encoding these 'parasitic' DNA elements confers a selective advantage on the host
genome.

<>

<1>Dalgaard, J.Z., Garrett, R.A.
<2>Protein-coding introns from the 23S rRNA-encoding gene form stable circles in the hyperthermophilic archaeon Pyrobaculum organotrophum.
<3>Gene
<4>121
<5>103-110
<6>1992
<7>Two archaeal introns have been discovered in the single-copy 23S rRNA-encoding gene of the
hyperthermophile, Pyrobaculum organotrophum.  After excision from rRNA transcripts, both
introns circularize and are stably retained in the cell.  Putative proteins encoded by the
introns and covering most of the intron sequence share a decapeptide motif with proteins
encoded by another archaeal intron and by group I introns.

<>

<1>Dalgaard, J.Z., Garrett, R.A., Belfort, M.
<2>A site-specific endonuclease encoded by a typical archaeal intron.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>5414-5417
<6>1993
<7>The protein encoded by the archaeal intron in the 23S rRNA gene of the hyperthermophile
Desulfurococcus mobilis is a double-strand DNase, that like group I intron homing
endonucleases, is capable of cleaving an intronless allele of the gene. This enzyme, I-Dmo I,
is unusual among the intron endonucleases in that it is thermostable and is expressed only
from linear and cyclized intron species and not from the precursor RNA. However, in analogy to
its eukaryotic counterparts, but unlike the bacteriophage enzymes, I-Dmo I makes a staggered
double-strand cut that generates 4-nt 3' extensions. Additionally, although the archaeal and
group I introns have entirely different structural properties and splicing pathways, I-DmoI
shares sequence similarity, in the form of the LAGLIDADG motif, with group I intron
endonucleases of eukaryotes. These observations support the independent evolutionary origin of
endonucleases and intron core elements and are consistent with the invasive potential of
endonuclease genes.

<>

<1>Dalgaard, J.Z., Garrett, R.A., Belfort, M.
<2>Purification and characterization of two forms of I-DmoI, a thermophilic site-specific endonuclease encoded by an archaeal intron.
<3>J. Biol. Chem.
<4>269
<5>28885-28892
<6>1994
<7>The archaeal intron in the 23 S rRNA gene of the hyperthermophile Desulfurococcus mobilis has
previously been shown to encode a site-specific DNA endonuclease that contains the LAGLIDADG
motif. The enzyme, I-DmoI, has been shown to be active in two forms when expressed in vitro,
from RNAs representing either the linear (I-DmoIl) or circular (I-DmoIc) intron. In this study
we have overexpressed I-DmoIl and I-DmoIc and purified the enzymes from Escherichia coli. The
optimal conditions for the enzymatic activity in vitro were determined, and the enzyme was
used to delimit the recognition boundary on its DNA substrate (14-20 nucleotides), an
intronless 23 S rRNA gene. Despite belonging to the archaeal kingdom, and being the product of
a hyperthermophile, I-DmoI shares many properties with LAGLIDADG intron and intein
endonucleases in other kingdoms. These results support the view that these phylogenetically
diverse enzymes, which function to mobilize the DNA sequences that encode them, share a common
ancestry.

<>

<1>Dalgaard, J.Z., Klar, A.J., Moser, M.J., Holley, W.R., Chatterjee, A., Mian, I.S.
<2>Statistical modeling and analysis of the LAGLIDADG family of site-specific endonucleases and identification of an intein that encodes a site-specific endonuclease of the HNH family.
<3>Nucleic Acids Res.
<4>25
<5>4626-4638
<6>1997
<7>The LAGLIDADG and HNH families of site-specific DNA endonucleases encoded by viruses,
bacteriophages as well as archaeal, eucaryotic nuclear and organellar genomes are
characterized by the sequence motifs 'LAGLIDADG' and 'HNH', respectively.  These
endonucleases have been shown to occur in different environments: LAGLIDADG endonucleases are
found in inteins, archaeal and group I introns and as free standing open reading frames; HNH
endonucleases occur in group I and group II introns and as ORFs.  Here, statistical models
(hidden Markov models, HMMs) that encompass both the conserved motifs and more variable
regions of these families have been created and employed to characterize known and potential
new family members.  A number of new, putative LAGLIDADG and HNH endonucleases have been
identified including an intein-encoded HNH sequence.  Analysis of an HMM-generated multiple
alignment of 130 LAGLIDADG family members and the three-dimensional structure of the I-CreI
endonuclease has enabled definition of the core elements of the repeated domain (~90 residues)
that is present in this family of proteins.  A conserved negatively charged residue is
proposed to be involved in catalysis.  Phylogenetic analysis of the two families indicates a
lack of exchange of endonucleases between different mobile elements (environments) and between
hosts from different phylogenetic kingdoms.  However, there does appear to have been
considerable exchange of endonuclease domains amongst elements of the same type. Such events
are suggested to be important for the formation of elements of new specificity.

<>

<1>Dalgaard, J.Z., Silva, G.H., Belfort, M., Van Roey, P.
<2>Crystallization and preliminary crystallographic analysis of the archaeal intron-encoded endonuclease I-DmoI.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>54
<5>1435-1436
<6>1998
<7>Two forms of the archaeal intron-encoded site-specific endonuclease I-DmoI, namely I-DmoIc and
I-DmoIl, have been purified and crystallized. Crystals of I-DmoIc are rod-shaped and diffract
to 3.0 A resolution, but further analysis was hampered by twinning. Crystals of I-DmoIl, which
is a six-amino-acid C-terminal truncation of I-DmoIc, are plate shaped and belong to space
group C2 with cell parameters a = 93.72, b = 37.03, c = 55.56 A, beta = 113.4 degrees, with
one molecule per asymmetric unit (Vm = 2.01 A3 Da-1). The crystals diffract to at least 2.3 A
resolution. A complete native data set has been measured and structure determination is
on-going.

<>

<1>Dalhoff, C., Lukinavicius, G., Klimasauskas, S., Weinhold, E.
<2>Direct transfer of extended groups from synthetic cofactors by DNA methyltransferases.
<3>Nat. Chem. Biol.
<4>2
<5>31-32
<6>2006
<7>S-Adenosyl-L-methionine is the major methyl donor for biological methylation reactions
catalyzed by methyltransferases.  We report the first chemical synthesis of AdoMet analogs
with extended carbon chains replacing the methyl group and their evaluation as cofactors for
all three classes of DNA methyltransferases.  Extended groups containing a double or triple
bond in the b position to the sulfonium center were transferred onto DNA in a catalytic and
sequence-specific manner, demonstrating a high utility of such synthetic cofactors for
targeted functionalization of biopolymers.

<>

<1>Dalhoff, C., Lukinavicius, G., Klimasauskas, S., Weinhold, E.
<2>Synthesis of S-adenosyl-L-methionine analogs and their use for sequence-specific transalkylation of DNA by methyltransferases.
<3>Nat. Protoc.
<4>1
<5>1879-1886
<6>2006
<7>Here we describe a one-step synthetic procedure for the preparation of S-adenosyl-L-methionine
analogs with extended carbon chains replacing the methyl group.  These AdoMet analogs function
as efficient cofactors for DNA methyltransferases, and we provide a protocol for
sequence-specific transfer of extended side chains from these AdoMet analogs to DNA by DNA
MTases.  Direct chemoselective allylation or propargylation of S-adenosyl-L-homocysteine at
sulfur is achieved under the acidic conditions needed to protect other nucleophilic positions
in AdoHcy.  The unsaturated bonds in b position to the sulfonium center of the resulting
AdoMet analogs are designed to stabilize the transition state formed upon DNA MTase-catalyzed
nucleophilic attack at the carbon next to the sulfonium center and lead to efficient transfer
of the extended side chains to DNA.  Using these protocols, sequence-specific functionalized
DNA can be obtained within one to two weeks.

<>

<1>Dalia, A.B., Lazinski, D.W., Camilli, A.
<2>Characterization of Undermethylated Sites in Vibrio cholerae.
<3>J. Bacteriol.
<4>195
<5>2389-2399
<6>2013
<7>The activities of DNA methyltransferases are important for a variety of cellular  functions in
bacteria. In this study, we developed a modified high-throughput
technique called methyl homopolymer tail mediated sequencing (methyl HTM-seq) to
identify the undermethylated sites in the Vibrio cholerae genome for the two DNA
methyltransferases, Dam, an adenine methyltransferase, and VchM, a cytosine
methyltransferase, during growth in rich medium in vitro. Many of the
undermethylated sites occurred in intergenic regions, and for most of these
sites, we identified the transcription factors responsible for undermethylation.
This confirmed the presence of previously hypothesized DNA-protein interactions
for these transcription factors and provided insight into the biological state of
these cells during growth in vitro. DNA adenine methylation has previously been
shown to mediate heritable epigenetic switches in gene regulation. However, none
of the undermethylated Dam sites tested showed evidence of regulation by this
mechanism. This study is the first to identify undermethylated adenines and
cytosines genomewide in a bacterium using second-generation sequencing
technology.

<>

<1>Daligault, H. et al.
<2>Complete genome sequence of Haliscomenobacter hydrossis type strain (O).
<3>Standards in Genomic Sciences
<4>4
<5>352-360
<6>2011
<7>Haliscomenobacter hydrossis van Veen et al. 1973 is the type species of the genus
Haliscomenobacter, which belongs to order 'Sphingobacteriales'. The species is of
interest because of its isolated phylogenetic location in the tree of life,
especially the so far genomically uncharted part of it, and because the organism
grows in a thin, hardly visible hyaline sheath. Members of the species were
isolated from fresh water of lakes and from ditch water. The genome of H.
hydrossis is the first completed genome sequence reported from a member of the
family 'Saprospiraceae'. The 8,771,651 bp long genome with its three plasmids of
92 kbp, 144 kbp and 164 kbp length contains 6,848 protein-coding and 60 RNA
genes, and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Daligault, H.E. et al.
<2>Whole-Genome Yersinia sp. Assemblies from 10 Diverse Strains.
<3>Genome Announcements
<4>2
<5>e01055-14
<6>2014
<7>Yersinia spp. are animal pathogens, some of which cause human disease. We sequenced 10
Yersinia isolates (from six species: Yersinia enterocolitica, Y.
fredericksenii, Y. kristensenii, Y. pestis, Y. pseudotuberculosis, and Y.
ruckeri) to high-quality draft or complete status. The genomes range in size from
3.77 to 4.94 Mbp.

<>

<1>Daligault, H.E. et al.
<2>Twenty Whole-Genome Bacillus sp. Assemblies.
<3>Genome Announcements
<4>2
<5>e00958-14
<6>2014
<7>Bacilli are genetically and physiologically diverse, ranging from innocuous to highly
pathogenic. Here, we present annotated genome assemblies for 20 strains
belonging to Bacillus anthracis, B. atrophaeus, B. cereus, B. licheniformis, B.
macerans, B. megaterium, B. mycoides, and B. subtilis.

<>

<1>Daligault, H.E. et al.
<2>Genome Assembly of Methicillin-Resistant Quality Control Strain Staphylococcus aureus CDC73-57501 (ATCC 29247).
<3>Genome Announcements
<4>2
<5>e00961-14
<6>2014
<7>Staphylococcus aureus is a major cause of bacterial infections in the United States, with high
percentages of serious infections resistant to a variety of
beta-lactam antibiotics. Here, we present the scaffolded genome assembly into 16
contigs of S. aureus CDC73-57501 (ATCC 29247), a methicillin-resistant quality
control strain.

<>

<1>Daligault, H.E. et al.
<2>Whole-Genome Assemblies of 56 Burkholderia Species.
<3>Genome Announcements
<4>2
<5>e01106-14
<6>2014
<7>Burkholderia is a genus of betaproteobacteria that includes three notable human pathogens: B.
cepacia, B. pseudomallei, and B. mallei. While B. pseudomallei and
B. mallei are considered potential biowarfare agents, B. cepacia infections are
largely limited to cystic fibrosis patients. Here, we present 56 Burkholderia
genomes from 8 distinct species.

<>

<1>Daligault, H.E. et al.
<2>Genome Assembly of Shigella flexneri ATCC 12022, a Quality Control Reference Strain.
<3>Genome Announcements
<4>2
<5>e01052-14
<6>2014
<7>Shigella flexneri causes shigellosis, severe and potentially life-threatening diarrhea, and
accounts for 18% of shigellosis cases in the United States. Here,
we present the 4.51-Mbp genome assembly of S. flexneri ATCC 12022, a quality
control and reference strain, in 10 scaffolds.

<>

<1>Daligault, H.E., Davenport, K.W., Minogue, T.D., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Li, P.E., Meincke, L., Munk, A.C., Palacios, G.F., Redden, C.L., Johnson, S.L.
<2>Complete Genome Assembly of Corynebacterium sp. Strain ATCC 6931.
<3>Genome Announcements
<4>2
<5>e01074-14
<6>2014
<7>The genus Corynebacterium is best known for the pathogen C. diphtheriae; however, it contains
mostly commensal and nonpathogenic, as well as several opportunistic,
pathogens. Here, we present the 2.47-Mb scaffolded assembly of the type strain,
Corynebacterium sp. ATCC 6931 (NCTC 1914), as deposited into GenBank under
accession number CP008913.

<>

<1>Daligault, H.E., Davenport, K.W., Minogue, T.D., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Lo, C.C., Meincke, L., Munk, A.C., Palacios, G.F., Redden, C.L., Johnson, S.L.
<2>Draft Genome Assembly of Klebsiella pneumoniae Type Strain ATCC 13883.
<3>Genome Announcements
<4>2
<5>e00939-14
<6>2014
<7>Klebsiella pneumoniae is a common cause of antibiotic-resistant bacterial infections in
immunocompromised individuals. Here, we present the 5.54-Mb
scaffolded assembly of the type strain K. pneumoniae type strain ATCC 13883, as
deposited in GenBank under accession no. JOOW00000000.

<>

<1>Daligault, H.E., Davenport, K.W., Minogue, T.D., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Lo, C.C., Meincke, L., Munk, C., Palacios, G.F., Redden, C.L., Johnson, S.L.
<2>Complete Genome Assembly of a Quality Control Reference Isolate, Moraxella catarrhalis Strain ATCC 25240.
<3>Genome Announcements
<4>2
<5>e00938-14
<6>2014
<7>Generally an opportunistic pathogen in the United States, Moraxella catarrhalis has acquired
resistance to multiple antibacterial/antimicrobial agents. Here, we
present the complete 1.9-Mb genome of M. catarrhalis strain ATCC 25240, as
deposited in NCBI under the accession number CP008804.

<>

<1>Daligault, H.E., Davenport, K.W., Minogue, T.D., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Lo, C.C., Meincke, L., Munk, C., Palacios, G.F., Redden, C.L., Johnson, S.L.
<2>Draft Genome Assembly of Bordetella bronchiseptica ATCC 10580, a Historical Canine Clinical Isolate.
<3>Genome Announcements
<4>2
<5>e00916-14
<6>2014
<7>We present the scaffolded genome of Bordetella bronchiseptica ATCC 10580, assembled into 98
contigs. This 5.1-Mb assembly (68.2% G+C content) contains
4,870 coding regions. The strain was originally isolated from canine lung tissue
and is used in quality control testing.

<>

<1>Daligault, H.E., Davenport, K.W., Minogue, T.D., Broomall, S.M., Bruce, D.C., Chain, P.S., Coyne, S.R., Gibbons, H.S., Jaissle, J., Lo, C.C., Meincke, L., Munk, A.C., Rosenzweig, C.N., Johnson, S.L.
<2>Draft Genome Assembly of Ralstonia pickettii Type Strain K-288 (ATCC 27853).
<3>Genome Announcements
<4>2
<5>e00973-14
<6>2014
<7>We present the genome assembly of Ralstonia pickettii K-288 (ATCC 27511), consisting of 27
contigs placed into a single scaffold. This 4.76-Mbp genome has
64.0% G+C content and 4,425 coding sequences. Because this is the type strain,
inclusion of its data set among other Ralstonia genomes should provide a
historical genomic perspective.

<>

<1>Daligault, H.E., Davenport, K.W., Minogue, T.D., Broomall, S.M., Bruce, D.C., Chain, P.S., Coyne, S.R., Gibbons, H.S., Jaissle, J., Rosenzweig, C.N., Scholz, M., Teshima, H., Johnson, S.L.
<2>Genome Assembly of Serratia marcescens Type Strain ATCC 13880.
<3>Genome Announcements
<4>2
<5>e00967-14
<6>2014
<7>Serratia marcescens ATCC 13880 is the type strain of the species and a commonly used quality
control strain. Here, we present the annotated genome assembly of
5.13 Mbp (59.8% G+C content) as submitted to NCBI under accession no.
JOVM00000000.

<>

<1>Dall'Acqua, W., Carter, P.
<2>Substrate-assisted catalysis: Molecular basis and biological significance.
<3>Protein Sci.
<4>9
<5>1-9
<6>2000
<7>Substrate-assisted catalysis (SAC) is the process by which a functional group in a substrate
contributes to catalysis by an enzyme.  SAC has been demonstrated for representatives of three
major enzyme classes: serine proteases, GTPases, and type II restriction endonucleases, as
well as lysozyme and hexose-1-phosphate uridylyltransferase.  Morover, structure-based
predictions of SAC have been made for many additional enzymes.  Examples of SAC include both
naturally occurring enzymes such as type II restriction endonucleases as well as engineered
enzymes including serine proteases.  In the latter case, a functional group from a substrate
can substitute for a catalytic residue replaced by site-directed mutagenesis. From a protein
engineering perspective, SAC provides a strategy for drastically changing enzyme substrate
specificity or even the reaction catalyzed.  From a biological viewpoint, SAC contributes
significantly to the activity of some enzymes and may represent a functional intermediate in
the evolution of catalysis.  This review focuses on advances in engineering enzyme specificity
and activity by SAC, together with the biological significance of this phenomenon.

<>

<1>Daly, C., Fitzgerald, G.
<2>Mechanisms of bacteriophage insensitivity in the lactic streptococci.
<3>Streptococcal Genetics, American Society for Microbiology, Ferretti, J.J., Curtiss, R., III, Washington, DC
<4>0
<5>259-268
<6>1987
<7>The mesophilic lactic streptococci, which comprise Streptococcus cremoris, S. lactis, and S.
lactis subsp. diacetylactis, are of major industrial importance as components of starter
cultures used in the manufacture of a variety of fermented dairy products, e.g., cheeses,
lactic butter, cultured buttermilk, sour cream, and quarg.  Bacteriophage attack has been
recognized since the 1930s as the most serious cause of starter culture inhibition in
commercial practice.  The consequence may be significant economic loss due to downgrading of
product or, in severe cases, total loss of the fermentation.  The vulnerability of dairying,
in contrast to other modern industrial fermentations, to phages exists partly because the
substrate, milk, cannot be sterilized for biochemical reasons, and in addition, the starter
culture must perform in large mechanized and automated units that demand consistent,
predictable acid production to ensure high-quality end products with the desired flavor and
texture characteristics.

<>

<1>Daly, C., Fitzgerald, G.F., Davis, R.
<2>Biotechnology of lactic acid bacteria with special reference to bacteriophage resistance.
<3>Antonie Van Leeuwenhoek
<4>70
<5>99-110
<6>1996
<7>Lactic acid bacteria play an important role in many food and feed fermentations.  In recent
years major advances have been made in unravelling the genetic and molecular basis of
significant industrial traits of lactic acid bacteria.  Bacteriophages which can infect and
destroy lactic acid bacteria pose a particularly serious threat to dairy fermentations that
can result in serious economic losses.  Consequently, these organisms and the mechanisms by
which they interact with their hosts have received much research attention.  This paper
reviews some of the key discoveries over the years that have led us to our current
understanding of bacteriophages themselves and the means by which their disruptive influence
may be minimized.

<>

<1>Daly, M.J., Gaidamakova, E.K., Matrosova, V.Y., Vasilenko, A., Zhai, M., Leapman, R.D., Lai, B., Ravel, B., Li, S.M., Kemner, K.M., Fredrickson, J.K.
<2>Protein Oxidation Implicated as the Primary Determinant of Bacterial Radioresistance.
<3>PLoS Biology
<4>5
<5>e92
<6>2007
<7>In the hierarchy of cellular targets damaged by ionizing radiation (IR), classical models of
radiation toxicity place DNA at the top. Yet, many
prokaryotes are killed by doses of IR that cause little DNA damage. Here
we have probed the nature of Mn-facilitated IR resistance in Deinococcus
radiodurans, which together with other extremely IR-resistant bacteria
have high intracellular Mn/Fe concentration ratios compared to
IR-sensitive bacteria. For in vitro and in vivo irradiation, we
demonstrate a mechanistic link between Mn(II) ions and protection of
proteins from oxidative modifications that introduce carbonyl groups.
Conditions that inhibited Mn accumulation or Mn redox cycling rendered D.
radiodurans radiation sensitive and highly susceptible to protein
oxidation. X-ray fluorescence microprobe analysis showed that Mn is
globally distributed in D. radiodurans, but Fe is sequestered in a region
between dividing cells. For a group of phylogenetically diverse
IR-resistant and IR-sensitive wild-type bacteria, our findings support the
idea that the degree of resistance is determined by the level of oxidative
protein damage caused during irradiation. We present the case that
protein, rather than DNA, is the principal target of the biological action
of IR in sensitive bacteria, and extreme resistance in Mn-accumulating
bacteria is based on protein protection.

<>

<1>Dam, B., Kube, M., Dam, S., Reinhardt, R., Liesack, W.
<2>Complete Sequence Analysis of Two Methanotroph-Specific repABC-Containing Plasmids from Methylocystis sp. Strain SC2.
<3>Appl. Environ. Microbiol.
<4>78
<5>4373-4379
<6>2012
<7>The complete nucleotide sequences of two large, low-copy-number plasmids of 229.6
kb (pBSC2-1) and 143.5 kb (pBSC2-2) were determined during assembly of the
whole-genome shotgun sequences of the methane-oxidizing bacterium Methylocystis
sp. strain SC2. The physical existence of the two plasmids in strain SC2 was
confirmed by pulsed-field gel electrophoresis followed by Southern hybridization.
Both plasmids have a conserved replication module of the repABC system and carry
genes involved in their faithful maintenance and conjugation. In addition, they
contain genes that might be involved in essential metabolic processes. These
include several heavy metal resistance genes and copper transport genes in
pBSC2-1 and a complete nitrous oxide reductase operon and a pmoC singleton in
pBSC2-2, the latter encoding the PmoC subunit of particulate methane
monooxygenase.

<>

<1>Damelin, M., Bestor, T.H.
<2>Biological functions of DNA methyltransferase 1 require its methyltransferase activity.
<3>Mol. Cell. Biol.
<4>27
<5>3891-3899
<6>2007
<7>DNA methyltransferase 1 (DNMT1) has been reported to interact with a wide variety of factors
and to contain intrinsic transcriptional
repressor activity. When a conservative point mutation was introduced
at the key catalytic residue, mutant DNMT1 failed to rescue any of the
phenotypes of Dnmt1-null embryonic stem (ES) cells, which indicated
that the biological functions of DNMT1 are exerted through the
methylation of DNA. ES cells that expressed the mutant protein did not
survive differentiation. Intracisternal A-particle family
retrotransposons were no longer methylated and were transcribed at high
levels. The proper localization of DNMT1 depended on normal genomic
methylation, and we discuss the implications of this finding for
epigenetic dysregulation in cancer.

<>

<1>Danaher, R., Stein, D.C.
<2>Characterization of a restriction enzyme isolated from a hydrothermal vent community bacterium, Hyphomonas jannaschiana.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>88
<5>213
<6>1988
<7>A restriction endonuclease has been isolated and purified from Hyphomonas
jannaschiana.  The enzyme, HjaI, was purified using an FPLC equipped with a
Mono Q column.  Yields of several thousand units of enzyme activity per gram of
cells were obtained.  Optimal activity was achieved in 25 mM Tris-HCl (pH-7.8),
100 mM NaCl, 10 mM MgCl/2, 100 micro g/ml BSA, 2 mM 2-mercaptoethanol at 37C.
This enzyme was inactivated by incubation at 65C.  By comparing the
fragmentation patterns of pBR322 and lambda DNA to computer generated patterns
provided by New England Biolabs, the recognition sequence was found to be
GATATC.  HjaI is an isoschizomer of EcoRV.  Cleavage of DNA by this enzyme
produced blunt ended fragments.  Furthermore, DNA isolated from H. jannaschiana
was not cleaved by EcoRV.

<>

<1>Danaher, R., Stein, D.C.
<2>Characterization of a restriction endonuclease and the cloning of its corresponding DNA methylase from the thermal vent bacterium Hyphomonas jannaschiana.
<3>Abstr. 1st Intl. Symp. Marine Mol. Biol., Center Marine Biotechnoogy, , Baltimore, MD
<4>0
<5>25
<6>1988
<7>The marine bacterium H. jannaschiana produces a restriction enzyme, HjaI, that
was purified by column chromatography.  Maximal enzyme activity occurred in 25
mM Tris-HCl (pH-9.0), 100 mM NaCl, 10 mM MgCl2, 100 micrograms/ml BSA, 2 mM
2-mercaptoethanol at 37C.  The recognition sequence was found to be GATATC, as
determined by comparing banding patterns obtained after digesting lambda DNA
with those predicted from its DNA sequence.  HjaI produces blunt ended DNA
fragments, as measured by a digestion/ligation scheme.  DNA isolated from H.
jannaschiana was not cleaved by EcoRV, an isoschizomer of HjaI.  The
corresponding methylase (M.HjaI) was cloned into E. coli methylase accepting
host (DH5 mcr).  H. jannaschiana chromosomal DNA was partially digested with
and inserted into the PstI site of pBR322.  Plasmid DNA was isolated from the
entire gene bank, digested with EcoRV, and transformed back into DH5 mcr.
M.HjaI clones were identified based on their plasmid DNA being partially
protected against cleavage by EcoRV.  Several plasmids were identified that
were partially resistant to EcoRV, and all shared a common 8 Kb PstI fragment.

<>

<1>Danaher, R.J., Stein, D.C.
<2>Expression of cloned restriction and modification genes, hjaIRM from Hyphomonas jannaschiana in Escherichia coli.
<3>Gene
<4>89
<5>129-133
<6>1990
<7>A type-II RM system, HjaI, was identified in the marine bacterium, Hyphomonas
jannaschiana.  The ENase recognizes GATATC, and DNA fragments generated after
cleavage with this enzyme contain blunt ends.  A DNA fragment encoding these
enzymes was cloned and expressed in Escherichia coli, although the level of
expression of the cloned genes was low.  DNA methylated by M.HjaI was not
restricted by the Mcr or Mrr restriction systems of E. coli.  Although H.
jannaschiana is a marine bacterium isolated near the thermal vents on the floor
of the Pacific Ocean, the biochemical properties of the ENase were similar to
those of EcoRV, an isoschizomer isolated from E. coli.

<>

<1>Dandekar, T., Huynen, M., Regula, J.T., Ueberle, B., Zimmermann, C.U., Andrade, M.A., Doerks, T., Sanchez-Pulido, L., Snel, B., Suyama, M., Yuan, Y.P., Herrmann, R., Bork, P.
<2>Re-annotating the Mycoplasma pneumoniae genome sequence: adding value, function and reading frames.
<3>Nucleic Acids Res.
<4>28
<5>3278-3288
<6>2000
<7>Four years after the original sequence submission, we have re-annotated the genome of
Mycoplasma pneumoniae to incorporate novel data. The total number of ORFs has been increased
from 677 to 688 (10 new proteins were predicted in intergenic regions, two further were newly
identified by mass spectrometry and one protein ORF was dismissed) and the number of RNAs from
39 to 42 genes. For 19 of the now 35 tRNAs and for six other functional RNAs the exact genome
positions were re-annotated and two new tRNA(Leu) and a small 200 nt RNA were identified.
Sixteen protein reading frames were extended and eight shortened. For each ORF a consistent
annotation vocabulary has been introduced. Annotation reasoning, annotation categories and
comparisons to other published data on M. pneumoniae functional assignments are given.
Experimental evidence includes 2-dimensional gel electrophoresis in combination with mass
spectrometry as well as gene expression data from this study. Compared to the original
annotation, we increased the number of proteins with predicted functional features from 349 to
458. The increase includes 36 new predictions and 73 protein assignments confirmed by the
published literature. Furthermore, there are 23 reductions and 30 additions with respect to
the previous annotation. mRNA expression data support transcription of 184 of the functionally
unassigned reading frames.

<>

<1>Daniel, A.S., Fuller-Pace, F.V., Legge, D.M., Murray, N.E.
<2>Distribution and diversity of hsd genes in Escherichia coli and other enteric bacteria.
<3>J. Bacteriol.
<4>170
<5>1775-1782
<6>1988
<7>We screened Salmonella typhimurium, Citrobacter freundii, Klebsiella
pneumoniae, Shigella boydii, and many isolates of Escherichia coli for DNA
sequences homologous to those encoding each of two unrelated type I restriction
and modification systems (EcoK and EcoA).  Both K- and A-related hsd genes were
identified, but never both in the same strain.  S. typhimurium encodes three
restriction and modification systems, but its DNA hybridized only to the
K-specific probe which we know to identify the StySB system.  No homology to
either probe was detected in the majority of E. coli strains, but in C.
freundii, we identified homology to the A-specific probe.  We cloned this
region of the C. freundii genome and showed that it encoded a functional,
A-related restriction system whose specificity differs from those of known type
I enzymes.  Sequences immediately flanking the hsdK genes of E.coli K-12 and
the hsdA genes of E. coli 15T- were shown to be homologous, indicating similar
or even identical positions in their respective chromosomes.  E. coli C has no
known restriction system, and the organization of its chromosome is consistent
with deletion of the three hsd genes and their neighbor, mcrB.

<>

<1>Daniels, J.S., Gates, K.S.
<2>Specificity of DNA cleavage by the type IIs restriction enzyme, HphI.
<3>ACS Abstracts
<4>208
<5>66
<6>1994
<7>Type IIs restriction enzymes are DNA-cleaving proteins that bind to short (4-8 base pair)
sequences of double-helical DNA and cleave at positions distant from the recognition site.
HphI is a type IIs restriction enzyme that recognizes the nonpalindromic sequence
5'-GGTGA-3' and is reported to hydrolyze the DNA backbone 8/7 base pairs away from the
recognition site. We will report interesting effects of varying substrate size and sequence on
the DNA-cleavage reaction of HphI.

<>

<1>Daniels, L.E., Wood, K.M., Scott, D.J., Halford, S.E.
<2>Subunit assembly for DNA cleavage by restriction endonuclease SgrAI.
<3>J. Mol. Biol.
<4>327
<5>579-591
<6>2003
<7>The SgrAI endonuclease usually cleaves DNA with two recognition sites more rapidly than DNA
with one site, often converting the former directly to
the products cut at both sites. In this respect, SgrAI acts like the
tetrameric restriction enzymes that bind two copies of their target sites
before cleaving both sites concertedly. However, by analytical
ultracentrifugation, SgrAI is a dimer in solution though it aggregates to
high molecular mass species when bound to its specific DNA sequence. Its
reaction kinetics indicate that it uses different mechanisms to cleave DNA
with one and with two SgrAI sites. It cleaves the one-site DNA in the
style of a dimeric restriction enzyme acting at an individual site,
mediating neither interactions in trans, as seen with the tetrameric
enzymes, nor subunit associations, as seen with the monomeric enzymes. In
contrast, its optimal reaction on DNA with two sites involves an
association of protein subunits: two dimers bound to sites in cis may
associate to form a tetramer that has enhanced activity, which then
cleaves both sites concurrently. The mode of action of SgrAI differs from
all restriction enzymes characterised previously, so this study extends
the range of mechanisms known for restriction endonucleases.

<>

<1>Daniels, L.L., Wais, A.C.
<2>Restriction and modification of halophage S45 in Halobacterium.
<3>Curr. Microbiol.
<4>10
<5>133-136
<6>1984
<7>A newly isolated group B1 bacteriophage, Halophage S45, is restricted and
modified in vivo by strains of Halobacterium.  Three strain-specific activities
have been observed.  Two of these occur in the same bacterial strain and appear
to be due to different kinds of enzymatic activities.  One of the restriction
specificities is shown to be associated with a strain-specific endonuclease
active on unmodified halobacterial DNA.

<>

<1>Danilevich, V.N., Livshits, V.A.
<2>The plasmid carrying a temperature-sensitive mutation in the DNA-methylase gene of the PstI system: Effect on host cells at nonpermissive temperature.
<3>Genetika
<4>35
<5>574-586
<6>1999
<7>Temperature-sensitive (ts) derivatives of plasmid pRMP1, a derivative of PBR322 containing
restriction and modification (RM) genes of the PstI system, were obtained using hydroxylamine
mutagenesis. One of the isolated plasmids responsible for the inhibition of Escherichia coli
cell growth at 42 degrees C, pRMPts, was analyzed in this work. Cells of Rec+ strains carrying
this plasmid were unable to divide at 42 degrees C and formed long non-septated filaments that
died upon prolonged cultivation. Cells of the RecA- strains carrying pRMPts did not form
filaments at 42 degrees C and rapidly disappeared. On agar media with or without ampicillin,
Rec+ and RecA- strains with this plasmid formed colonies of temperature-resistant (tr)
derivatives with frequencies ranging from 1.5 x 10^-4 to 4 x 10^-6 in independent clones. The
structure of plasmids from cells of tr- derivatives of Rec+ and RecA- strains carrying plasmid
pRMPts was analyzed by the set of restriction enzymes. Reversions to the temperature-resistant
phenotype were shown to result from the following events: (1) the insertional inactivation of
the PstI restriction enzyme gene in pRMPts (the insertion of the IS1 element); (2) deletions
in plasmid DNA fragments that partially or completely cover the restriction enzyme gene; (3)
point mutations; and (4) others. The effect of the chromosomal sulA mutation on the
maintenance of the ts-plasmid in bacterial cells was studied at 42 degrees C. High efficiency
loss of the plasmid was detected in pRMPts-carrying Rec+ cells with the sulA::Tn5 mutation
grown in liquid and solid nutrient media at this temperature.  Under similar conditions,
plasmid loss was not detected in SulA+ cells. On the basis of the data obtained, it is
concluded that the ts-mutation is located in the DNA-methylase gene of plasmid pRMPts. Mutant
DNA methylase was unable to methylate all sites in the chromosomal DNA at 42 degrees C. Some
of the unmethylated sites can be digested with the PstI enzyme, which leads to the induction
of SOS response in Rec+ cells or to total mortality in cells with the recA phenotype.

<>

<1>Danin-Poleg, Y., Elgavish, S., Raz, N., Efimov, V., Kashi, Y.
<2>Genome Sequence of the Pathogenic Bacterium Vibrio vulnificus Biotype 3.
<3>Genome Announcements
<4>1
<5>e00136-13
<6>2013
<7>We report the first genome sequence of the pathogenic Vibrio vulnificus biotype 3. This draft
genome sequence of the environmental strain VVyb1(BT3), isolated in
Israel, provides a representation of this newly emerged clonal group, which
reveals higher similarity to the clinical strains of biotype 1 than to the
environmental ones.

<>

<1>Danish-Daniel, M., Gan, H.Y., Gan, H.M., Saari, N.A., Usup, G.
<2>Genome Sequence of Nitratireductor basaltis Strain UMTGB225, a Marine Bacterium Isolated from a Green Barrel Tunicate in Bidong Island, Malaysia.
<3>Genome Announcements
<4>2
<5>e01015-14
<6>2014
<7>Nitratireductor basaltis strain UMTGB225 is a Gram-negative bacterium isolated from a marine
tunicate found in Bidong Island, Terengganu, Malaysia. In this
study, the genome of Nitratireductor basaltis UMTGB225 was sequenced to gain
insight into the role of this bacterium and its association with tunicate hosts
in a coral reef habitat.

<>

<1>Danna, K., Nathans, D.
<2>Specific cleavage of simian virus 40 DNA by restriction endonuclease of Hemophilus influenzae.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>68
<5>2913-2917
<6>1971
<7>A bacterial restriction endonuclease has been used to produce specific
fragments of SV40 DNA.  Digestion of DNA from plaque-purified stocks of SV40
with the restriction endonuclease from Hemophilus influenzae gave ll fragments
resolvable by polyacrylamide gel electrophoresis, eight of which were equimolar
with the original DNA.  The fragments ranged from about 6.5 x 10/5 to 7.4 x
10/4 daltons, as determined by electron microscopy, DNA content, or
electrophoretic mobility.

<>

<1>Danna, K.J.
<2>Daniel Nathans.
<3>Proc. Amer. Phil. Soc.
<4>154
<5>338-354
<6>2010
<7>Dr. Daniel Nathans, the brilliant and reticent Nobel Prize-winning scientist who was regarded
by many of his peers as the conscience of the Johns Hopkins University, died on 16 November
1999 of leukemia at his home.  He was seventy-one.  Proud and passionate about ideas, Dr.
Nathans helped Hopkins sustain its sense of tradition during a time that brought wrenching
changes to academic medical centers.  In addition to reaching and conducting research, Dr.
Nathans served the university as a valued adviser and interim president.  "He was the wise man
of the medical center and he was seen that way," said former Hopkins medical dean Michael M.E.
Johns, now chancellor for health affairs at Emory University in Atlanta.  "He walked quietly
but carried the responsibility of the respect he had very, very thoughtfully."  Dr. William R.
Broady, president of the Johns Hopkins University, said, "Dan Nathans was an extraordinary
human being.  He was brilliant, of course … But as one who had the privilege of knowing Dan
well, I was always most impressed with the man - modest, soft-spoken, unassuming, even
self-effacing." Colleagues described Dr. Nathans as a man who taught by example and by
encouragement, and as a researcher of vision and intellectual vigor.  "The great scientist can
separate the wheat from the chaff.  One of the great things in research is to know what
questions to ask, what subjects to focus on," said Dr. Solomon H. Snyder, the director of
Hopkins's neuroscience department.  "He would ask the very best questions."

<>

<1>Danna, K.J., Nathans, D.
<2>Bidirectional replication of simian virus 40 DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>3097-3100
<6>1972
<7>SV40 (Simian Virus 40) DNA was pulse-labeled with [3H]thymidine in infected
monkey cells, and the distribution of label within newly completed molecules
was determined by analysis of specific fragments produced by restriction
endonuclease from Hemophilus influenzae.  From these data, an order of
synthesis or temporal order of the fragments was deduced.  Comparison of the
temporal order with the physical order of the fragments in the SV40 DNA
molecule indicates a correspondence in thehse orders for two separate groups of
fragments.  From an analysis of the results, we conclude that SV40 DNA
replication begins at a specific site and proceeds bidirectionally, terminating
about halfway around the circular molecule from the initiation point.

<>

<1>Dantoft, S.H., Bielak, E.M., Seo, J.G., Chung, M.J., Jensen, P.R.
<2>Complete Genome Sequence of Pediococcus pentosaceus Strain SL4.
<3>Genome Announcements
<4>1
<5>e01106-13
<6>2013
<7>Pediococcus pentosaceus SL4 was isolated from a Korean fermented vegetable product, kimchi. We
report here the whole-genome sequence (WGS) of P. pentosaceus
SL4. The genome consists of a 1.79-Mb circular chromosome (G+C content of 37.3%)
and seven distinct plasmids ranging in size from 4 kb to 50 kb.

<>

<1>Danzitz, M.J.
<2>Analysis of a protein: DNA interface; the effect of altered DNA substrates on EcoRI methylase function.
<3>Diss. Abstr.
<4>53
<5>5180B
<6>1993
<7>EcoRI methyltransferase (MTase) binds to its double stranded recognition site 5'-GAATTC-3'
and methylates it at the inner adenosine of each strand using S-adenosyl-L-methionine as the
methyl donor. The Mtase is unperturbed by hemi-methylation at either inner adenosine in
synthetic substrates, facilitating the study of substrates harboring modified bases at unique
positions. Using this strategy the contributions of the N2 amino groups of guanosine and the
C5 methyl groups of thymine toward specificity were investigated.

As a result largely of changes in KM-DNA, deoxyinosine substitution for guanosine at either
position in the M.EcoRI recognition site reduces the specificity constant (kcat/KM-DNA) 13 and
39 fold. Thus methylation, which occurs in the major groove, is sensitive to both of these
minor groove modifications. The specificity constant obtained from the double substituted
substrate (14 fold below the control) indicates that there may be structural "communication"
between these sites.

The MTase is insensitive to removal of either one of the outer methyl groups by substitution
of deoxyuridine for the outer thymine residues. When either the inner thymine residue opposite
the hemi-methylation or both inner thymidine residues are substituted with deoxyuridine, the
KM-DNA is 5 to 10 times larger than for the control substrate leading to a specificity
constant that is 2 to 9 times larger.

Comparisons of entropic and enthalpic data clearly show that there are structural
perturbations within all of these modified substrates.

The results indicate that not only are there effects of functional group removal on MTase
function, but also structural change in the substrate beyond the altered residue is observed.
Substitutions which reduce enzyme specificity were coupled with ones that are fairly silent in
several different combinations. The resulting substrate could not only regain some of its lost
specificity, but could also become a better substrate than the control. Thus, structural
changes within the DNA substrate caused by functional group alterations can have a more
profound effect on specificity than the removal of a protein:DNA contact.


<>

<1>Danzitz, M.J., Reich, N.O.
<2>Investigations of EcoRI methylase contacts with its DNA substrate.
<3>ACS Abstracts
<4>196
<5>95
<6>1988
<7>The interactions between DNA binding proteins and their DNA substrates have
been analyzed by many researchers during the past decade.  Much of this
interest stems from the fact that DNA binding proteins are involved in a myriad
of biologically significant events.  The monomeric EcoRI methylase recognizes
the double stranded DNA sequence GAATTC and methylates the second adenine of
this site.  This methylation event protects the site from cleavage by the
corresponding dimeric endonuclease.  The portion of the DNA substrate which is
protected by EcoRI methylase binding has been determined with the use of the
hydroxyl radical DNA footprinting technique.  The ability of this enzyme to
discriminate its preferred recognition site from modified sites has also been
investigated kinetically by comparisons of specificity constants (kcat/Km).

<>

<1>Danzitz, M.J., Reich, N.O.
<2>Characterization of a protein-DNA interface:  DNA substrate functionalties important for EcoRI methylase specificity.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>84
<6>1989
<7>EcoRI methylase is a monomeric 38,050 dalton enzyme requiring the cofactor
S-adenosylmethionine (AdoMet) and its DNA substrate for activity.  The methylase recognizes
the double stranded DNA sequence 5'-GAATTC-3' and methylates the second adenine of each
strand.  Details about the DNA protein interface of a DNA methylase or any monomeric DNA
binding protein have yet to be determined.  Hydroxyl radical footprinting experiments of
recognition site containing oligonucleotides complexed with the methylase alone or with the
methylase and sinefungin (an AdoMet analog and methylase inhibitor) have been carried out.
These experiments have shown the sugar phosphate backbone regions in and around the
recognition site which are protected by the methylase both with and without sinefungin
present.  Some differences in both the binding pattern and affinity have been observed between
these complexes.  Specific functionalities have been removed from the substrate at defined
locations in the recognition site by the incorporation of non-standard nucleotides, such as
uracil.  The kinetic parameters obtained with these substrates have allowed us to evaluate the
importance of specific functional groups in substrate recognition (in the uracil example the
methyl group of each thymidine).  The effect of single asymmetric substitutions on specificity
and catalysis are being dissected out using this approach.  The thermal denaturation
characteristics of the modified substrates are being used to probe for additional substrate
structural changes.

<>

<1>Dar, M.E., Bhagwat, A.S.
<2>Mechanism of expression of DNA repair gene vsr, an Escherichia coli gene that overlaps the DNA cystosine methylase gene, dcm.
<3>Mol. Microbiol.
<4>9
<5>823-833
<6>1993
<7>The DNA cytosine methylase gene of Escherichia coli, dcm, overlaps an open reading frame (ORF)
that continues in +1 register past the end of the dcm. This ORF codes for a gene, vsr, that is
required for a T:G to C:G base mismatch correction process. In this study, mutants that affect
the level of expression of the two genes were constructed and characterized. Further, a
previously isolated mutant, dcm-6, was cloned and mutations within it were identified.
Northern blots were used to identify dcm-specific RNA species in wild type and dcm-6 cells.
Based on these studies we conclude that there is a six-codon overlap between vsr and dcm. The
two proteins appear to be made from a single RNA transcript and translation of dcm is required
for the efficient synthesis of Vsr. Further, Vsr is active by itself and may not be produced
as a fusion with Dcm. This is the first example of chromosomal genes that overlap in their
coding regions and produce proteins with distinct functions.

<>

<1>Darii, M.V., Cherepanova, N.A., Subach, O.M., Kirsanova, O.V., Rasko, T., Slaska-Kiss, K., Kiss, A., Deville-Bonne, D., Reboud-Ravaux, M., Gromova, E.S.
<2>Mutational analysis of the CG recognizing DNA methyltransferase SssI: Insight into enzyme-DNA interactions.
<3>Biochim. Biophys. Acta
<4>1794
<5>1654-1662
<6>2009
<7>To characterize important steps of DNA methylation by M.SssI, a prokaryotic DNA-(cytosine
C5)methyltransferase (C5-MTase) sharing the
specificity of eukaryotic C5-MTases (5'-CG-3'), ten amino acids,
selected on the basis of sequence alignments and a computational model,
were subjected to mutational analysis. Wild-type and mutant M.SssI
variants were studied to determine methylation activity, DNA binding
affinity, capacity to induce base flipping, and ability to form
covalent complex with a DNA substrate containing the mechanism-based
inhibitor 2-pyrimidinone. Wild-type M.SssI induced strong fluorescence
when bound to substrate DNA containing 2-aminopurine in place of the
target cytosine, indicating flipping of the target base. Reduced
fluorescence, moderate, or drastic loss of methyltransferase activity
and reduced DNA binding suggest the involvement of the conserved 5745
(motif IV), 8232 (motif VIII, QxRx (R) under bar, and T313 (variable
region, conserved (T) under barL), as well as of the non-conserved Q147
in base flipping. Replacement of E186 (motif VI, (E) under bar NV) and
8230 (motif VIII, Qx (R) under bar xR) with alanine resulted in loss of
methyltransferase activity without impairing DNA binding affinity.
These data are consistent with the catalytic role of E186 and 8230, and
provide, for the first time, experimental support for the essential
function of the hitherto not investigated invariant arginine of motif
VIII in C5-MTases.

<>

<1>Darii, M.V., Kirsanova, O.V., Drutsa, V.L., Kochetkov, S.N., Gromova, E.S.
<2>Isolation and site-directed mutagenesis of DNA methyltransferase SssI.
<3>Mol. Biol. (Mosk)
<4>41
<5>110-117
<6>2007
<7>Prokaryotic DNA methyltransferase SssI (M.SssI) methylates cytosines at C5 in CpG sequences.
Bacterial strains that produced M.SssI and its
mutants as His(6)-tagged proteins were constructed. To verify the role
of Ser300 in recognizing the CpG sequence by the enzyme, Ser300 was
replaced by Gly or Pro. The substitutions had virtually no effect on
DNA binding and methylation by M.SssI apart from a slight decrease in
binding in the case of S300P. It was assumed that no contact with DNA
is formed by the side chain of Ser300 and the carbonyl oxygen and amide
nitrogen of its peptide bonds. In addition, Ala was substituted for
highly conserved Val188, presumably involved in stabilization of the
flipped-out cytosine during the reaction. The substitution decreased
fivefold the dissociation constant of the enzyme-substrate complex and
halved the initial rate of DNA methylation. Despite the lack of a
considerable effect of V188A, it was assumed that Val188 does form a
contact with the target cytosine, but such a contact is formed with Ala
in the case of the V188A mutant.

<>

<1>Darmon, E., Leach, D.R.F.
<2>Bacterial Genome Instability.
<3>Microbiol. Mol. Biol. Rev.
<4>78
<5>1-39
<6>2014
<7>Bacterial genomes are remarkably stable from one generation to the next but are plastic on an
evolutionary time scale, substantially shaped by horizontal gene transfer, genome
rearrangement, and the activities of mobile DNA elements. This implies the existence of a
delicate balance between the maintenance of genome stability and the tolerance of genome
instability. In this review, we describe the specialized genetic elements and the endogenous
processes that contribute to genome instability. We then discuss the consequences of genome
instability at the physiological level, where cells have harnessed instability to mediate
phase and antigenic variation, and at the evolutionary level, where horizontal gene transfer
has played an important role. Indeed, this ability to share DNA sequences has played a major
part in the evolution of life on Earth. The evolutionary plasticity of bacterial genomes,
coupled with the vast numbers of bacteria on the planet, substantially limits our ability to
control disease.

<>

<1>Darrasse, A., Bolot, S., Serres-Giardi, L., Charbit, E., Boureau, T., Fisher-Le, S.M., Briand, M., Arlat, M., Gagnevin, L., Koebnik, R., Noel, L.D., Carrere, S., Jacques, M.A.
<2>High-Quality Draft Genome Sequences of Xanthomonas axonopodis pv. glycines Strains CFBP 2526 and CFBP 7119.
<3>Genome Announcements
<4>1
<5>e01036-13
<6>2013
<7>We report here the high-quality draft genome sequences of two strains of Xanthomonas
axonopodis pv. glycines, the causal agent of bacterial pustule on
soybeans. Comparison of these genomes with those of phylogenetically closely
related pathovars of Xanthomonas spp. will help to understand the mechanisms
involved in host specificity and adaptation to host plants.

<>

<1>Dartois, V., De Backer, O., Colson, C.
<2>Sequence of the Salmonella typhimurium StyLT1 restriction-modification genes: homologies with EcoP1 and EcoP15 type-III R-M systems and presence of helicase domains.
<3>Gene
<4>127
<5>105-110
<6>1993
<7>The StyLTI restriction-modification (R-M) system of Salmonella typhimurium has recently been
suggested to belong to the type-III R-M systems [De Backer and Colson. Gene 97 (1991)
103-107]. The nucleotide sequences of StyLTI mod and res have been determined. Two closely
adjacent open reading frames were found 12 bp apart with coding capacities of 651 (Mod) and
982 (Res) amino acids (aa), respectively. The genes, lying in the same direction of
transcription in the mod-res order, are transcribed as distinct units. The deduced aa
sequences reveal homologies with known type-III enzymes from the Escherichia coli P1 prophage,
E. coli P15 plasmid and Bacillus cereus chromosome. In addition, the StyLTI restriction
endonuclease (ENase), like other type-I and type-III ENases, contains sequence motifs
characteristic of superfamily-II helicases, which may be involved in DNA unwinding at the
cleavage site.

<>

<1>Dassa, B., Pietrokovski, S.
<2>Origin and evolution of inteins and other hint domains.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Derbyshire, V., Stoddard, B.L.,Wood, D.W., Berlin Heidelberg
<4>16
<5>211-231
<6>2005
<7>The intein protein family is part of the Hint superfamily, named after the characteristic
structure fold first identified in Hedgehog and intein protein domains.  Four characterized
Hint domain families are currently known: Hog-Hint, intein, and two types of bacterial
intein-like domains.  Together with sharing the same structural fold and common sequence
features, Hint domains have similar biochemical activities.  The domains post-translationally
process the proteins in which they are present by protein-splicing, self-cleavage or ligation
activities.  Hint domains are 130 to 160 amino acids long, sharing 4-6 conserved sequence
motifs.  The Hint protein fold is a compact, relatively flat, symmetrical structure, mainly
composed of b-strands, with its N- and C-termini close together.  Inteins usually include
additional homing-endonuclease and DNA-binding domains, not necessary for protein splicing,
which mediate the homing of the intein gene.  Species from the three domains of life, Eukarya,
Bacteria and Archaea, include Hint domains.  While inteins are apparently limited to
unicellular eukaryotes and prokaryotes, Hog-Hint domains are present in multicellular animals.
BIL domains and inteins overlap in their phylogenetic distributions, both are present in
bacteria, but in different types of proteins.  Data gathered on Hint domains since their
discovery in 1990 have identified their biochemical activity, their genetics and evolution.
Apparently, each Hint family has its own distinct biological role.  Nevertheless, many facets
of Hint domains are still unknown or debated.

<>

<1>Datta, J., Ghoshal, K., Sharma, S.M., Tajima, S., Jacob, S.T.
<2>Biochemical fractionation reveals association of DNA methyltransferase (Dnmt) 3b With Dnmt1 and that of Dnmt 3a with a histone H3 methyltransferase and Hdac1.
<3>J. Cell. Biochem.
<4>88
<5>855-864
<6>2003
<7>De novo DNA methyltransferases, Dnmt3a and 3b, were purified by fractionation of S-100 extract
from mouse lymphosarcoma cells through
several chromatographic matrices followed by glycerol density gradient
centrifugation. Dnmt3a was separated from Dnmt3b and Dnmt1 in the first
column, Q-Sepharose whereas Dnmt3b co-purified with Dnmt1 after further
fractionation through Mono-S and Mono-Q columns and glycerol density
gradient centrifugation. Following purification, the majority of de novo
DNA methyltransfearse activity was associated with Dnmt3b/Dnmt1 fractions.
By contrast, the fractions containing Dnmt3a alone exhibited markedly
reduced activity, which correlated with diminished expression of this
isoform in these cells. Histone deacetylase 1(Hdac1) cofractionated with
Dnmt3a throughout purification whereas Hdac1 was separated from
Dnmt3b/Dnmt1 following chromatography on Mono-Q column. Dnmt3a purified
through glycerol gradient centrifugation was also associated with a
histone H3 methyltransferase (HMTase) activity whereas purified
Dnmt3b/Dnmt1 was devoid of any HMTase activity. The activity of this
HMTase was abolished when lysine 9 of N-terminal histone H3 peptide was
replaced by leucine whereas mutation of lysine 4 to leucine inhibited this
activity only partially. This is the first report on the identification of
a few key co-repressors associated with endogenous Dnmt3a and of a complex
containing Dnmt3b and a minor form of Dnmt1 following extensive
biochemical fractionation.

<>

<1>Datta, J., Majumder, S., Bai, S.M., Ghoshal, K., Kutay, H., Smith, D.S., Crabb, J.W., Jacob, S.T.
<2>Physical and functional interaction of DNA methyltransferase 3A with Mbd3 and Brg1 in mouse lymphosarcoma cells.
<3>Cancer Res.
<4>65
<5>10891-10900
<6>2005
<7>Dnmt3a and Dnmt3b are de novo DNA methyltransferases that also act as transcriptional
repressors independent of methyltransferase activity.
To elucidate the underlying mechanism of transcriptional repression,
Dnmt3a was purified from mouse lymphosarcoma cells (P1798) by extensive
fractionation on five different chromatographic matrices followed by
glycerol density gradient centrifugation. Liquid chromatography
electrospray tandem mass spectrometry analysis of Dnmt3a-associated
polypeptides identified the methyl CpG binding protein Mbd3, histone
deacetylase 1(Hdac1), and components of Brg1 complex (Brg1, Baf155, and
Baf57) in the purified preparation. Association of Dnmt3a with Mbd3 and
Brg1 was confirmed by coinummoprecipitation and coimmunolocalization
studies. Glutathione S-transferase pulldown assay showed that the
NH2-terminal ATRX homology domain of Dnmt3a interacts with the methyl
CpG binding domain of Mbd3 and with both bromo and ATPase domains of
Brg1. Chromatin immunoprecipitation assay revealed that all three
proteins are associated with transcriptionally silent methylated
metallothionein (MT-I) promoter in the mouse lymphosarcoma cells. To
understand the functional significance of their association with the
promoter, their role on the MT-I promoter activity was analyzed by
transient transfection assay. The results showed that Mbd3 and Dnmt3a
specifically inhibited the methylated promoter, and the catalytic
activity of Dnmt3a was dispensable for the suppression. In contrast,
the wild-type but not the ATPase-inactive mutant of Brg1 suppressed
MT-I promoter irrespective of its methylation status, implicating
involvement of ATP-dependent chromatin remodeling in the process.
Coexpression of two of the three interacting proteins at a time
augmented their repressor function. This study shows physical and
functional interaction of Dnmt3a with components of nucleosome
remodeling machinery.

<>

<1>Datz, M., Janetzki-Mittmann, C., Franke, S., Gunzer, F., Schmidt, H., Karch, H.
<2>Analysis of the enterohemorrhagic Escherichia coli 0157 DNA region containing lambdoid phage gene p and Shiga-like toxin structural genes.
<3>Appl. Environ. Microbiol.
<4>62
<5>791-797
<6>1996
<7>In this study, we determined the nucleotide sequence of the p gene contained within a 5-kb
EcoRI restriction fragment cloned from Shiga-like toxin II (SLT-II)-converting phage 933W of
Escherichia coli O157:H7 strain EDL933. The p gene was 702 bp long and had 95.3% sequence
similarity to the p gene of phage lambda. Multiple hybridization patterns were obtained when
genomic DNA fragments were hybridized with both p and slt-I, slt-II, or slt-IIc sequences. All
O157 isolates also possessed an analog of lambda gene p which was not linked with either slt-I
or slt-II. Restriction fragment length polymorphism comparisons of clinical O157 isolates and
derivatives undergoing genotype turnover during infection were made, and loss of large DNA
fragments that hybridized with slt-II and p sequences was observed. To further analyze the DNA
region containing the p and slt genes, we amplified fragments by using PCR with one primer
complementary to p and the other complementary to either the slt-I or the slt-II gene. PCR
analysis with enterohemorrhagic E. coli O157 and non-O157 strains yielded PCR products that
varied in size between 5.1 and 7.8 kb. These results suggest that even within O157 isolates,
the genomes of SLT-converting phages differ.  The methods described here may assist in further
investigation of SLT-encoding phages and their role in the epidemiology of infection with
enterohemorrhagic E. coli.

<>

<1>Daujotyte, D., Liutkeviciute, Z., Tamulaitis, G., Klimasauskas, S.
<2>Chemical mapping of cytosines enzymatically flipped out of the DNA helix.
<3>Nucleic Acids Res.
<4>36
<5>e57
<6>2008
<7>Haloacetaldehydes can be employed for probing unpaired DNA structures involving cytosine and
adenine residues. Using an enzyme that was structurally proven to flip its target cytosine out
of the DNA helix, the HhaI DNA methyltransferase (M.HhaI), we demonstrate the suitability of
the chloroacetaldehyde modification for mapping extrahelical (flipped-out) cytosine bases in
protein-DNA complexes. The generality of this method was verified with two other DNA
cytosine-5 methyltransferases, M.AluI and M.SssI, as well as with two restriction
endonucleases, R.Ecl18kI and R.PspGI, which represent a novel class of base-flipping enzymes.
Our results thus offer a simple and convenient laboratory tool for detection and mapping of
flipped-out cytosines in protein-DNA complexes.

<>

<1>Daujotyte, D., Serva, S., Vilkaitis, G., Merkiene, E., Venclovas, C., Klimasauskas, S.
<2>HhaI DNA methyltransferase uses the protruding Gln237 for active flipping of its target cytosine.
<3>Structure
<4>12
<5>1047-1055
<6>2004
<7>Access to a nucleotide by its rotation out of the DNA helix (base flipping) is used by
numerous DNA modification and repair enzymes.  Despite extensive studies of the paradigm HhaI
methyltransferase, initial events leading to base flipping remained elusive.  Here we
demonstrate that the replacement of the target C:G pair with the 2-aminopurine:T pair in the
DNA or shortening of the side chain of Gln237 in the protein severely perturb base flipping,
but retain specific DNA binding.  Kinetic analyses and molecular modeling suggest that a
steric interaction between the protruding side chain of Gln237 and the target cytosine in
B-DNA reduces the energy barrier for flipping by 3 kcal/mol.  Subsequent stabilization of an
open state by further 4 kcal/mol is achieved through specific hydrogen bonding of the side
chain to the orphan guanine.  Gln237 thus plays a key role in actively opening the target C:G
pair by a ?push-and-bind? mechanism.

<>

<1>Daujotyte, D., Vilkaitis, G., Manelyte, L., Skalicky, J., Szyperski, T., Klimasauskas, S.
<2>Solubility engineering of the HhaI methyltransferase.
<3>Protein Eng.
<4>16
<5>295-301
<6>2003
<7>DNA methylation is involved in epigenetic control of numerous cellular processes in
eukaryotes, however, many mechanistic aspects of this
phenomenon are not yet understood. A bacterial prototype cytosine-C5
methyltransferase, M.HhaI, serves as a paradigm system for structural and
mechanistic studies of biological DNA methylation, but further analysis of
the 37 kDa protein is hampered by its insufficient solubility (0.15 mM).
To overcome this problem, three hydrophobic patches on the surface of
M.HhaI that are not involved in substrate interactions were subjected to
site-specific mutagenesis. Residues M51 or V213 were substituted by polar
amino acids of a similar size, and/or the C-terminal tetrapeptide FKPY was
replaced by a single glycine residue (Delta324G). Two out of six mutants,
delta324G and V213S/delta324G, showed improved solubility in initial
analyses and were purified to homogeneity using a newly developed
procedure. Biochemical studies of the engineered methyltransferases showed
that the deletion mutant delta324G retained identical DNA binding, base
flipping and catalytic properties as the wild-type enzyme. In contrast,
the engineered enzyme showed (i) a significantly increased solubility
(>0.35 mM), (ii) high-quality 2D-[(15)N,(1)H] TROSY NMR spectra, and (iii)
(15)N spin relaxation times evidencing the presence of a monomeric
well-folded protein in solution.

<>

<1>Daum, H.A., White, H.W., Seidell, C.M., Johnson, P.A.
<2>Cloning, restriction digestion and DNA labeling of large DNA fragments (>1 kb) in the presence of remelted SeaPlaque GTG agarose gels.
<3>Biotechniques
<4>11
<5>784-790
<6>1991
<7>Large DNA fragments (>1 kb), separated in low melting temperature SeaPlaque GTG
agarose gels, can be enzymatically processed directly in the presence of this
agarose (in-gel).  Time saving protocols are discussed for in-gel processing of
large DNA fragments in the presence of remelted SeaPlaque GTG agarose,
including cloning into pUC18, nick translation, random priming and restriction
digestion.  These in-gel molecular biology techniques are as efficient as those
using DNA recovered from agarose.  The effects of UV irradiation, Mg2+
concentration and agarose concentration on selected in-gel protocols are also
discussed.

<>

<1>Davalieva, K., Ziberovski, J., Efremov, G.D.
<2>Bme585I [5'-CCCGC(4/6)-3'], a new isoschizomer of restriction endonuclease FauI, isolated from a strain of Bacillus mesentericus.
<3>Microbiol. Res.
<4>159
<5>129-133
<6>2004
<7>Bme585I is a new member of the restriction endonuclease type IIS family. It was partially
purified from the heterothrophic, mesophilic
bacterial strain Bacillus mesentericus 585 by ammonium sulphate
precipitation and phosphocellulose column chromatography. Bme585I is a
monomeric protein with a molecular mass of 62 kD. The enzyme is active
over a broad pH range from 7.0 to 8.8, has a temperature optimum of
37 degrees C and tolerance of NaCl. in reaction buffer from 0 to 400 mM.
Bme585I recognizes the asymmetric sequence 5'-CCCGC(4/6)-3' and is
therefore an isoschizomer of restriction endonuclease FauI.

<>

<1>Davenport, K.W. et al.
<2>Whole-genome sequences of nine francisella isolates.
<3>Genome Announcements
<4>2
<5>e00941-14
<6>2014
<7>Primarily a zoonotic disease, Francisella tularensis is a fastidious intracellular pathogen
and is listed as a CDC category A pathogen with notably
high pathogenicity. Here we present the scaffolded genome assemblies of nine
Francisella strains: eight F. tularensis and one F. philomiragia.

<>

<1>Davenport, K.W., Daligault, H.E., Minogue, T.D., Bishop-Lilly, K.A., Broomall, S.M., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Gibbons, H.S., Jaissle, J., Redden, C.L., Rosenzweig, C.N., Scholz, M.B., Teshima, H., Johnson, S.L.
<2>Complete Genome Assembly of Staphylococcus epidermidis AmMS 205.
<3>Genome Announcements
<4>2
<5>e01059-14
<6>2014
<7>Staphylococcus epidermidis causes a large number of catheter-related sepsis infections
annually in the United States. We present the 2.54-Mbp complete genome
assembly of reference strain S. epidermidis AmMS 205, including a single 37.7-kbp
plasmid. The annotated assembly is available in GenBank under accession numbers
CP009046 and CP009047.

<>

<1>Davenport, K.W., Daligault, H.E., Minogue, T.D., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Li, P.E., Palacios, G.F., Redden, C.L., Scholz, M.B., Teshima, H., Johnson, S.L.
<2>Whole-Genome Sequence of Listeria monocytogenes Type Strain 53 XXIII.
<3>Genome Announcements
<4>2
<5>e00970-14
<6>2014
<7>Listeria monocytogenes causes the food-borne illness listeriosis that primarily infects
'vulnerable' groups (e.g., elderly adults, pregnant women, very young
children, and the immunocompromised). We sequenced the genome of L. monocytogenes
XXIII (ATCC 15313) and assembled it into a single scaffold (three contigs) and
deposited the annotated assembly into GenBank as accession no. JOOX00000000.

<>

<1>Davenport, K.W., Daligault, H.E., Minogue, T.D., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Lo, C.C., Palacios, G.F., Redden, C.L., Scholz, M.B., Teshima, H., Johnson, S.L.
<2>Complete Genome Sequence of Type Strain Pasteurella multocida subsp. multocida ATCC 43137.
<3>Genome Announcements
<4>2
<5>e01070-14
<6>2014
<7>Soft-tissue infection by Pasteurella multocida in humans is usually associated with a dog- or
cat-related injury, and these infections can become aggressive. We
sequenced the type strain P. multocida subsp. multocida ATCC 43137 into a single
closed chromosome consisting of 2,271,840 bp (40.4% G+C content), which is
currently available in the NCBI GenBank under the accession number CP008918.

<>

<1>Davenport, K.W., Daligault, H.E., Minogue, T.D., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Palacios, G.F., Redden, C.L., Scholz, M.B., Teshima, H., Johnson, S.L.
<2>Draft Genome Assembly of Delftia acidovorans Type Strain 2167.
<3>Genome Announcements
<4>2
<5>e00917-14
<6>2014
<7>The Delftia acidovorans 2167 (ATCC 15668, Delftia type strain) genome was sequenced into a
6-contig scaffolded assembly of 6.78-Mb. This environmental
microbe, previously named to both the Comamonas and Pseudomonas genera, is an
opportunistic pathogen and often the subject of phylogenetic placement debates.

<>

<1>Davenport, K.W., Daligault, H.E., Minogue, T.D., Broomall, S.M., Bruce, D.C., Chain, P.S., Coyne, S.R., Gibbons, H.S., Jaissle, J., Li, P.E., Rosenzweig, C.N., Scholz, M.B., Johnson, S.L.
<2>Complete Genome Sequence of Stenotrophomonas maltophilia Type Strain 810-2 (ATCC  13637).
<3>Genome Announcements
<4>2
<5>e00974-14
<6>2014
<7>An emerging nosocomial pathogen, Stenotrophomonas maltophila has a high mortality rate in
those it infects. Here, we present the complete genome sequence of
Stenotrophomonas maltophilia 810-2 (ATCC 13637), the type strain of the species.
The 5-Mb (66.1% G+C content) genome has been deposited in NCBI under accession
number CP008838.

<>

<1>Davenport, K.W., Daligault, H.E., Minogue, T.D., Bruce, D.C., Chain, P.S., Coyne, S.R., Jaissle, J.G., Koroleva, G.I., Ladner, J.T., Li, P.E., Palacios, G.F., Scholz, M.B., Teshima, H., Johnson, S.L.
<2>Draft Genome Assembly of Acinetobacter baumannii ATCC 19606.
<3>Genome Announcements
<4>2
<5>e00832-14
<6>2014
<7>Acinetobacter baumannii is an emerging nosocomial pathogen, and therefore high-quality genome
assemblies for this organism are needed to aid in detection,
diagnostic, and treatment technologies. Here we present the improved draft
assembly of A. baumannii ATCC 19606 in two scaffolds. This 3,953,621-bp genome
contains 3,750 coding regions and has a 39.1% G+C content.

<>

<1>David, A., Bleimling, N., Beuck, C., Lehn, J.M., Weinhold, E., Teulade-Fichou, M.P.
<2>DNA mismatch-specific base flipping by a bisacridine macrocycle.
<3>Chembiochem
<4>4
<5>1326-1331
<6>2003
<7>Most, if not all, enzymes that chemically modify nucleobases in DNA flip their target base
from the inside of the double helix into an extrahelical
position. This energetically unfavorable conformation is partly stabilized
by specific binding of the apparent abasic site being formed. Thus, DNA
base-flipping enzymes, like DNA methyltransferases and DNA glycosylases,
generally bind very strongly to DNA containing abasic sites or abasic-site
analogues. The macrocyclic bisacridine BisA has previously been shown to
bind abasic sites. Herein we demonstrate that it is able to specifically
recognize DNA base mismatches and most likely induces base flipping.
Specific binding of BisA to DNA mismatches was studied by thermal
denaturation experiments by using short duplex oligodeoxynucleotides
containing central TT, TC, or TG mismatches or a TA match. In the presence
of the macrocycle a strong increase in the melting temperature of up to
7.1 degrees C was observed for the mismatch-containing duplexes, whereas
the melting temperature of the fully matched duplex was unaffected.
Furthermore, BisA binding induced an enhanced reactivity of the mispaired
thymine residue in the DNA toward potassium permanganate oxidation. A
comparable reactivity has previously been observed for a TT target base
mismatch in the presence of DNA methyltransferase M.TaqI. This similarity
to a known base-flipping enzyme suggests that insertion of BisA into the
DNA helix displaces the mispaired thymine residue into an extrahelical
position, where it should be more prone to chemical oxidation. Thus, DNA
base flipping does not appear to be limited to DNA-modifying enzymes but
it is likely to also be induced by a small synthetic molecule binding to a
thermodynamically weakened site in DNA.

<>

<1>Davidson, R.M., Reynolds, P.R., Farias-Hesson, E., Duarte, R.S., Jackson, M., Strong, M.
<2>Genome Sequence of an Epidemic Isolate of Mycobacterium abscessus subsp. bolletii from Rio de Janeiro, Brazil.
<3>Genome Announcements
<4>1
<5>e00617-13
<6>2013
<7>Multiple isolates of Mycobacterium abscessus subsp. bolletii, collectively called BRA100, were
associated with outbreaks of postsurgical skin infections across
various regions of Brazil from 2003 to 2009. We announce the draft genome
sequence of a newly sequenced BRA100 strain, M. abscessus subsp. bolletii
CRM-0020, isolated from a patient in Rio de Janeiro, Brazil.

<>

<1>Davidson, Y.Y., Soper, S.A., Margolis, S., Sander, L.C.
<2>Immobilization of the restriction enzymes HaeIII and HindIII on porous silica particles via a glutaraldehyde linkage for the micro-digestion of dsDNA with analysis by capillary electrophoresis.
<3>J. Sep. Sci.
<4>24
<5>10-16
<6>2001
<7>Solid-phase DNA restriction digest reactors have been developed consisting of silica particles
modified with a covalently tethered
restriction enzyme. This solid-phase restriction reactor enables
digestion and separation of minute quantities of DNA with minimal
reagent consumption. In this study, the restriction enzymes, HaeIII,
PstI, and HindIII, were successfully immobilized via glutaraldehyde
linkages to porous silica micro-particles. Studies were carried out to
examine the impact of immobilization on enzymatic activity. Digestions
of phiX174-RF DNA phage and SV40 viral DNA were performed with the
immobilized enzymes by placing the silica particles in solution with
the target DNA with digestion times of 120 min and 240 min
respectively. The digests were analyzed off-line using capillary
electrophoresis (CE) with laser-induced fluorescence (LIF) detection.
Timed studies were performed to establish optimal conditions for
complete digestion. Digests utilizing immobilized HaeIII and HindIII
were similar in composition to homogeneous, free solution digests. PstI
showed no evidence of activity upon immobilization. The immobilized
restriction enzymes could also be used for multiple rounds of
digestion; however, longer incubation times were required for
successive runs probably due to partial denaturation of the restriction
enzyme. Digests also were prepared and isolated by use of a simple
micro-spin column consisting of a layer of immobilized enzyme-coated
silica on a molecular weight cut-off filter. Using this approach,
digestion times were comparable to solution digests as previously
mentioned; however, enzyme reuse and reaction product isolation was
facilitated.

<>

<1>Davie, J.J., Earl, J., de Vries, S.P., Ahmed, A., Hu, F.Z., Bootsma, H.J., Stol, K., Hermans, P.W., Wadowsky, R.M., Ehrlich, G.D., Hays, J.P., Campagnari, A.A.
<2>Comparative analysis and supragenome modeling of twelve Moraxella catarrhalis clinical isolates.
<3>BMC Genomics
<4>12
<5>70
<6>2011
<7>BACKGROUND: M. catarrhalis is a gram-negative, gamma-proteobacterium and
an opportunistic human pathogen associated with otitis media (OM) and
exacerbations of chronic obstructive pulmonary disease (COPD). With direct
and indirect costs for treating these conditions annually exceeding $33
billion in the United States alone, and nearly ubiquitous resistance to
beta-lactam antibiotics among M. catarrhalis clinical isolates, a greater
understanding of this pathogen's genome and its variability among isolates
is needed. RESULTS: The genomic sequences of ten geographically and
phenotypically diverse clinical isolates of M. catarrhalis were determined
and analyzed together with two publicly available genomes. These twelve
genomes were subjected to detailed comparative and predictive analyses
aimed at characterizing the supragenome and understanding the metabolic
and pathogenic potential of this species. A total of 2383 gene clusters
were identified, of which 1755 are core with the remaining 628 clusters
unevenly distributed among the twelve isolates. These findings are
consistent with the distributed genome hypothesis (DGH), which posits that
the species genome possesses a far greater number of genes than any single
isolate. Multiple and pair-wise whole genome alignments highlight limited
chromosomal re-arrangement. CONCLUSIONS: M. catarrhalis gene content and
chromosomal organization data, although supportive of the DGH, show modest
overall genic diversity. These findings are in stark contrast with the
reported heterogeneity of the species as a whole, as wells as to other
bacterial pathogens mediating OM and COPD, providing important insight
into M. catarrhalis pathogenesis that will aid in the development of novel
therapeutic regimens.

<>

<1>Davies, G.P., Kemp, P., Molineux, I.J., Murray, N.E.
<2>The DNA translocation and ATPase activities of restriction-deficient mutants of EcoKI.
<3>J. Mol. Biol.
<4>292
<5>787-796
<6>1999
<7>EcoKI, a type I restriction enzyme, specifies DNA methyltransferase, ATPase, endonuclease and
DNA translocation activities. One subunit (HsdR) of the oligomeric enzyme contributes to those
activities essential for restriction. These activities involve ATP-dependent DNA translocation
and DNA cleavage. Mutations that change amino acids within recognisable motifs in HsdR impair
restriction. We have used an in vivo assay to monitor the effect of these mutations on DNA
translocation. The assay follows the EcoKI-dependent entry of phage T7 DNA from the phage
particle into the host cell. Earlier experiments have shown that mutations within the seven
motifs characteristic of the DEAD-box family of proteins that comprise known or putative
helicases severely impair the ATPase activity of purified enzymes. We find that the mutations
abolish DNA translocation in vivo. This provides evidence that these motifs are relevant to
the coupling of ATP hydrolysis to DNA translocation. Mutations that identify an endonuclease
motif similar to that found at the active site of type II restriction enzymes and other
nucleases have been shown to abolish DNA nicking activity.  When conservative changes are made
at these residues, the enzymes lack nuclease activity but retain the ability to hydrolyse ATP
and to translocate DNA at wild-type levels. It has been speculated that nicking may be
necessary to resolve the topological problems associated with DNA translocation by type I
restriction and modification systems. Our experiments show that loss of the nicking activity
associated with the endonuclease motif of EcoKI has no effect on ATPase activity in vitro or
DNA translocation of the T7 genome in vivo.

<>

<1>Davies, G.P., Martin, I., Sturrock, S.S., Cronshaw, A., Murray, N.E., Dryden, D.T.F.
<2>On the structure and operation of type I DNA restriction enzymes.
<3>J. Mol. Biol.
<4>290
<5>565-579
<6>1999
<7>Type I DNA restriction enzymes are large, molecular machines possessing DNA methyltransferase,
ATPase, DNA translocase and endonuclease activities. The ATPase, DNA translocase and
endonuclease activities are specified by the restriction (R) subunit of the enzyme. We
demonstrate that the R subunit of the Eco KI type I restriction enzyme comprises several
different functional domains. An N-terminal domain contains an amino acid motif identical with
that forming the catalytic site in simple restriction endonucleases, and changes within this
motif lead to a loss of nuclease activity and abolish the restriction reaction. The central
part of the R subunit contains amino acid sequences characteristic of DNA helicases. We
demonstrate, using limited proteolysis of this subunit, that the helicase motifs are contained
in two domains. Secondary structure prediction of these domains suggests a structure that is
the same as the catalytic domains of DNA helicases of known structure. The C-terminal region
of the R subunit can be removed by elastase treatment leaving a large fragment, stable in the
presence of ATP, which can no longer bind to the other subunits of Eco KI suggesting that this
domain is required for protein assembly. Considering these results and previous models of the
methyltransferase part of these enzymes, a structural and operational model of a type I DNA
restriction enzyme is presented.

<>

<1>Davies, G.P., Powell, L.M., Webb, J.L., Cooper, L.P., Murray, N.E.
<2>EcoKI with an amino acid substitution in any one of seven DEAD-box motifs has impaired ATPase and endonuclease activities.
<3>Nucleic Acids Res.
<4>26
<5>4828-4836
<6>1998
<7>For type I restriction systems, recently determined nucleotide sequences predict conserved
amino acids in the subunit that is essential for restriction but not modification (HsdR). The
conserved sequences emphasize motifs characteristic of the DEAD-box family of proteins which
comprises putative helicases, and they identify a new candidate for motif IV. We provide
evidence based on an analysis of EcoKI which supports both the relevance of DEAD-box motifs to
the mechanism of restriction and the new definition of motif IV. Amino acid substitutions
within the newly identified motif IV and those in six other previously identified DEAD-box
motifs, but not in the original motif IV, confer restriction-deficient phenotypes. We have
examined the relevance of the DEAD-box motifs to the restriction pathway by determining the
steps permitted in vitro by the defective enzymes resulting from amino acid substitutions in
each of the seven motifs. EcoKI purified from the seven restriction-deficient mutants binds to
an unmethylated target sequence and, in the presence of AdoMet, responds to ATP by undergoing
the conformational change essential for the pathway of events leading to DNA cleavage. The
seven enzymes have little or no ATPase activity and no endonuclease activity, but they retain
the ability to nick unmodified DNA, though at reduced rates. Nicking of a DNA strand could
therefore be an essential early step in the restriction pathway, facilitating the
ATP-dependent translocation of DNA, particularly if this involves DNA helicase activity.

<>

<1>Davies, J.K.
<2>DNA restriction and modification systems in Neisseria gonorrhoeae.
<3>Clin. Microbiol. Rev.
<4>2
<5>S78-S82
<6>1989
<7>A review.  Introduces new nomenclature.

<>

<1>Davies, J.K., Normark, S.
<2>A relationship between plasmid structure, structural lability, and sensitivity to site-specific endonucleases in Neisseria gonorrhoeae.
<3>Mol. Gen. Genet.
<4>177
<5>251-260
<6>1980
<7>Nearly all gonococcal strains carry a small "phenotypically cryptic" plasmid of
approximately 4,200 basepairs.  A detailed physical map of this plasmid has
been constructed, revealing the presence of numerous putative inverted repeats.
These studies also revealed the presence on the plasmid of rercognition
sequences for several site-specific endonucleases (particularly HpaII, MspI and
AluI) that are particularly resistant to cleavage, and confirmed previous
reports of structural lability.  Both the sites that are resistant to cleavage,
and the observed structural variation are association with the inverted
repetitive sequences.

<>

<1>Davies, M.R., Shera, J., Van Domselaar, G.H., Sriprakash, K.S., McMillan, D.J.
<2>A Novel Integrative Conjugative Element Mediates Genetic Transfer from Group G Streptococcus to Other -Hemolytic Streptococci.
<3>J. Bacteriol.
<4>191
<5>2257-2265
<6>2009
<7>Lateral gene transfer is a significant contributor to the ongoing
evolution of many bacterial pathogens, including beta-hemolytic
streptococci. Here we provide the first characterization of a novel
integrative conjugative element (ICE), ICESde3396, from Streptococcus
dysgalactiae subsp. equisimilis (group G streptococcus [GGS]), a bacterium
commonly found in the throat and skin of humans. ICESde3396 is 64 kb in
size and encodes 66 putative open reading frames. ICESde3396 shares 38
open reading frames with a putative ICE from Streptococcus agalactiae
(group B streptococcus [GBS]), ICESa2603. In addition to genes involves in
conjugal processes, ICESde3396 also carries genes predicted to be involved
in virulence and resistance to various metals. A major feature of
ICESde3396 differentiating it from ICESa2603 is the presence of an 18-kb
internal recombinogenic region containing four unique gene clusters, which
appear to have been acquired from streptococcal and nonstreptococcal
bacterial species. The four clusters include two cadmium resistance
operons, an arsenic resistance operon, and genes with orthologues in a
group A streptococcus (GAS) prophage. Streptococci that naturally harbor
ICESde3396 have increased resistance to cadmium and arsenate, indicating
the functionality of genes present in the 18-kb recombinogenic region. By
marking ICESde3396 with a kanamycin resistance gene, we demonstrate that
the ICE is transferable to other GGS isolates as well as GBS and GAS. To
investigate the presence of the ICE in clinical streptococcal isolates, we
screened 69 isolates (30 GGS, 19 GBS, and 20 GAS isolates) for the
presence of three separate regions of ICESde3396. Eleven isolates
possessed all three regions, suggesting they harbored ICESde3396-like
elements. Another four isolates possessed ICESa2603-like elements. We
propose that ICESde3396 is a mobile genetic element that is capable of
acquiring DNA from multiple bacterial sources and is a vehicle for
dissemination of this DNA through the wider beta-hemolytic streptococcal
population.

<>

<1>Davis, B.M., Chao, M.C., Waldor, M.K.
<2>Entering the era of bacterial epigenomics with single molecule real time DNA sequencing.
<3>Curr. Opin. Microbiol.
<4>16
<5>192-198
<6>2013
<7>DNA modifications, such as methylation guide numerous critical biological processes, yet
epigenetic information has not routinely been collected as part of
DNA sequence analyses. Recently, the development of single molecule real time
(SMRT) DNA sequencing has enabled detection of modified nucleotides (e.g. 6mA,
4mC, 5mC) in parallel with acquisition of primary sequence data, based on
analysis of the kinetics of DNA synthesis reactions. In bacteria, genome-wide
mapping of methylated and unmethylated loci is now feasible. This technological
advance sets the stage for comprehensive, mechanistic assessment of the effects
of bacterial DNA methyltransferases (MTases)-which are ubiquitous, extremely
diverse, and largely uncharacterized-on gene expression, chromosome structure,
chromosome replication, and other fundamental biological processes. SMRT
sequencing also enables detection of damaged DNA and has the potential to uncover
novel DNA modifications.

<>

<1>Davis, E.O., Jenner, P.J.
<2>Protein splicing -- the lengths some proteins will go to.
<3>Antonie Van Leeuwenhoek
<4>67
<5>131-137
<6>1995
<7>We review the recently discovered phenomenon of protein splicing which
is the excision of an inernal protein sequence at the protein level rather than at the RNA
level. The means by which examples of protein splicing have been idenified are described,
and the similarities of the internally spliced protein products (or inteins) are discussed.
Comparisons are made between inteins and group I RNA introns.  We describe the
evidence supporting excision of inteins by a post-translational autocatalytic reaction of a
full
length polypeptide precursor, rather than by RNA splicing.  An examination is made of
some of the proposed mechanism schemes and the evidence supporting them is presented.

<>

<1>Davis, E.O., Jenner, P.J., Brooks, P.C., Colston, M.J., Sedgwick, S.G.
<2>Protein splicing in the maturation of M. tuberculosis RecA protein: a mechanism for tolerating a novel class of intervening sequence.
<3>Cell
<4>71
<5>201-210
<6>1992
<7>The M. tuberculosis recA locus comprises an 85 kd open reading frame but produced 38 kd RecA
and 47 kd products in E. coli. No RNA processing was detected; rather, an 85 kd precursor
protein was spliced, releasing a 47 kd spacer protein, and joining its terminal fragments to
form mature RecA protein. "Spacer" protein was also produced in M. tuberculosis and from a
hybrid spacer-LacZa fusion molecule. Mutagenesis at codon wobble positions at one splice
junction showed that protein rather than nucleotide sequence determined splicing activity.
Other mutants defined additional regions needed for splicing and allowed processing to be
followed. Splicing was essential for RecA activity in E. coli. The possibility that splicing
is a manifestation of a novel class of genetic element is discussed.

<>

<1>Davis, E.O., Sedgwick, S.G., Colston, M.J.
<2>Novel structure of the recA locus of Mycobacterium tuberculosis implies processing of the gene product.
<3>J. Bacteriol.
<4>173
<5>5653-5662
<6>1991
<7>A fragment of Mycobacterium tuberculosis DNA containing recA-like sequences was identified by
hybridization with the Escherichia coli recA gene and cloned. Although no expression was
detected from its own promoter in E. coli, expression from a vector promoter partially
complemented E. coli recA mutants for recombination, DNA repair, and mutagenesis, but not for
induction of phage lambda. This clone produced a protein which cross-reacts with antisera
raised against the E. coli RecA protein and was approximately the same size. However, the
nucleotide sequence of the cloned fragment revealed the presence of an open reading frame for
a protein about twice the size of other RecA proteins and the cloned product detected by
Western blotting (immunoblotting). The predicted M. tuberculosis RecA protein sequence was
homologous with RecA sequences from other bacteria, but this homology was not dispersed;
rather it was localized to the first 254 and the last 96 amino acids, with the intervening 440
amino acids being unrelated. Furthermore, the junctions of homology were in register with the
uninterrupted sequence of the E. coli RecA protein. Identical restriction fragments were found
in the genomic DNAs of M. tuberculosis H37Rv and H37Ra and of M. bovis BCG. It is concluded
that the ancestral recA gene of these species diversified via an insertional mutation of at
least 1,320 bp of DNA. Possible processing mechanisms for synthesizing a normal-size RecA
protein from this elongated sequence are discussed.

<>

<1>Davis, E.O., Thangaraj, H.S., Brooks, P.C., Colston, M.J.
<2>Evidence of selection for protein introns in the recAs of pathogenic mycobacteria.
<3>EMBO J.
<4>13
<5>699-703
<6>1994
<7>Protein introns are recently discovered genetic elements whose intervening sequences are
removed from a precursor protein by an unusual protein splicing intron, and the religation of
the amino- and carboxy-terminal fragments of the precursor. The recA gene of Mycobacterium
tuberculosis contains one such element and we now show that the other major mycobacterial
pathogen, Mycobacterium leprae, also possesses a protein intron in its recA, although other
mycobacterial recA genes do not. However, these two protein introns are different in size,
sequence and location of insertion of their coding sequences into the recAs of M. tuberculosis
and M. leprae, indicating that acquisition of the protein introns has occurred independently
in the two species, and thus suggesting that there has been selection for splicing in the
maturation of RecA in the pathogenic mycobacteria. The M. leprae protein intron provides an
example of conditional protein splicing, splicing occurring in M. leprae itself but not when
expressed in Escherichia coli, unlike most previously described protein introns. These
observations suggest that protein introns may perform a function for their host, rather than
being just selfish elements.

<>

<1>Davis, J., Hill, R.T.
<2>Draft Genome Sequence of Hawaiian Sea Slug Symbiont Vibrio sp. Strain ER1A.
<3>Genome Announcements
<4>2
<5>e00820-14
<6>2014
<7>Bacteria belonging to the genus Vibrio are prevalent in the marine environment and are known
for forming symbiotic relationships with hosts. Vibrio sp. strain
ER1A is a dominant symbiont of the Hawaiian sea slug, Elysia rufescens. Here we
report the draft genome sequence of Vibrio sp. ER1A.

<>

<1>Davis, J.R. et al.
<2>Genome Sequence of Streptomyces viridosporus Strain T7A ATCC 39115, a Lignin-Degrading Actinomycete.
<3>Genome Announcements
<4>1
<5>e00416-13
<6>2013
<7>We announce the availability of the genome sequence of Streptomyces viridosporus  strain T7A
ATCC 39115, a plant biomass-degrading actinomycete. This bacterium is
of special interest because of its capacity to degrade lignin, an underutilized
component of plants in the context of bioenergy. It has a full complement of
genes for plant biomass catabolism.

<>

<1>Davis, J.R., Goodwin, L.A., Woyke, T., Teshima, H., Bruce, D., Detter, C., Tapia, R., Han, S., Han, J., Pitluck, S., Nolan, M., Mikhailova, N., Land, M.L., Sello, J.K.
<2>Genome Sequence of Amycolatopsis sp. Strain ATCC 39116, a Plant Biomass-Degrading Actinomycete.
<3>J. Bacteriol.
<4>194
<5>2396-2397
<6>2012
<7>We announce the availability of a high-quality draft of the genome sequence of Amycolatopsis
sp. strain 39116, one of few bacterial species that are known to
consume the lignin component of plant biomass. This genome sequence will further
ongoing efforts to use microorganisms for the conversion of plant biomass into
fuels and high-value chemicals.

<>

<1>Davis, M., Guorong, X., Glickman, G., Reich, N.
<2>Eludication of sites flanking CpG dinucleotides important in the regulation of mammalian cytosine DNA methyltransferase.
<3>FASEB J.
<4>7
<5>A1291
<6>1993
<7>It is known that methylation of cytosines in eukaryotes occurs only at cytosines within CpG
dinucleotides. However, not every CpG site in eukaryotic DNA is methylated, giving rise to
various patterns of methylation for different types of cells and stages of development. How
this variability is established is not well understood, but methylation of genes at particular
sites has been shown to inhibit their expression. To examine the possibility that sequences
flanking CpG dinucleotides act differentially in the regulation of mammalian cytosine
methyltransferase, a number of dissimilar CpG flanking sequences on various substrates have
been examined. Through use of gel-shift and methylation data carried out on plasmid pBR322 and
SV40 viral DNA, sites of facilitated methyltransferase-DNA interaction have been determined.
Footprinting assays currently being employed will reveal sites critical to recognition and
catalysis. Characterization of the protein-DNA interactions involved with the mammalian
methyltransferase may lead to new insights into its gene regulatory mechanism.

<>

<1>Davis, R., Hossain, M.J., Liles, M.R., Panizzi, P.
<2>Complete Genome Sequence of Staphylococcus aureus Tager 104, a Sequence Type 49 Ancestor.
<3>Genome Announcements
<4>1
<5>e00706-13
<6>2013
<7>We report here the complete genome sequence of Staphylococcus aureus Tager 104, originally
isolated from a cutaneous abscess in 1947 by Morris Tager. Sequence
typing of the strain revealed its membership in sequence type 49 (ST49), a
previously unknown multilocus sequence type (MLST) in clinical samples.

<>

<1>Davis, R., Van der Lelie, D., Mercenier, A., Daly, C., Fitzgerald, G.F.
<2>ScrFI restriction-modification system of Lactococcus lactis subsp. cremoris UC503: Cloning and Characterization of two ScrFI methylase genes.
<3>Appl. Environ. Microbiol.
<4>59
<5>777-785
<6>1993
<7>Two genes from the total genomic DNA of dairy starter culture Lactococcus lactis subsp.
cremoris UC503, encoding ScrFI modification enzymes, have been cloned and expressed in
Escherichia coli. No homology between the two methyase genes was detected, and inverse
polymerase chain reaction of flanking chromosomal DNA indicated that both were linked on the
Lactococcus genome. Neither clone encoded the cognate endonuclease. The DNA sequence of one of
the methylase genes (encoded by pCI931M) was determined and consisted of an open reading frame
1,170 bp long, which could encode a protein of 389 amino acids (Mr,44.5). The amino acid
sequence contained the highly characteristic motifs of an m5C methylase. Extensive regions of
homology were observed with the methylases of NlaX, EcoRII, and Dcm.

<>

<1>Davis, T.O., Henderson, I., Brehm, J.K., Minton, N.P.
<2>Development of a transformation and gene reporter system for group II, non-proteolytic Clostridium botulinum type B strains.
<3>J. Mol. Microbiol. Biotechnol.
<4>2
<5>59-69
<6>2000
<7>Non-proteolytic, Group II strains of Clostridium botulinum are of particular concern to the
food industry because of their ability to survive and grow in REPFEDs (refrigerated processed
foods of extended durability). Their analysis would benefit from the availability of a gene
transfer system. In the present study we have been able, for the first time, to demonstrate
transformation in a representative Group II strain, ATCC 25765. Initial attempts to transform
ATCC 25765 with existing clostridial cloning vectors (pMTL540E and pMTL500E) were, however,
prevented by a restriction barrier. Through a combination of classical and molecular
approaches we were able to show that strain ATCC 25765 possesses a restriction endonuclease
(CboI) and a methylase activity (M.CboI) which have the same specificity as MspI and M.MspI,
respectively. CboI cleaves the palindrome 5'-CCGG-3' to generate a 3'-GC sticky end, whilst
M.CboI specifically methylates the external C residue. An E. coli host was generated which
expressed a Bacillus subtilis methylase enzyme (M.BsuFI) with equivalent specificity to
M.CboI. Plasmids (pMTL540E and pMTL500E) prepared in this strain were subsequently shown to be
capable of transforming ATCC 25765. The highest frequencies (0.8 X 10^4) transformants per
microg of DNA) were obtained when cells were cultivated in media supplemented with 1% (w/v)
glycine, and when the electroporation was undertaken at 10 kV/cm, 25 microF and at 400 ohms.
Having developed an effective transformation procedure, we went on to construct reporter
cassettes based on the Thermanaerobacterium sulfurigenes lacZ and the Vibrio fischeri luxAB
genes. Using the former, and promoter regions isolated from the botulinum toxin genes, we have
obtained preliminary evidence that reporter genes may be used to evaluate the physiological
factors that affect toxin production in the food environment.

<>

<1>Davison, J., Brunel, F.
<2>Restriction insensitivity in bacteriophage T5.  I. Genetic characterization of mutants sensitive to EcoRI restriction.
<3>J. Virol.
<4>29
<5>11-16
<6>1979
<7>Unmodified bacteriophage T5 is able to grow normally on bacterial hosts carrying three
different Escherichia coli restriction systems, EcoK, EcoPI, and EcoRI.  Under the same
conditions, the plating efficiency of bacteriophage lambda is less than 10^-9.  At least in
the case of EcoRI, this lack of in vivo restriction is not due to lack of restriction sites on
the T5 DNA molecule.  These observations suggest that bacteriophage T5 specifies one or more
restriction protection systems.  Mutants (ris) of T5 have been isolated which confer
sensitivity to EcoRI restriction but not to EcoK or EcoPI.  The mutations are located in the
pre-early region of the genetic map but are too far apart to be alleles of a single gene.
Complementation studies show that the ris mutants can be helped to grow on the
EcoRI-restricting host by coinfection with T5+.  This result provides evidence for a
restriction protection function but does not necessarily show that the ris mutants are
defective in such a system.

<>

<1>Dawoud, T.M., Jiang, T., Mandal, R.K., Ricke, S.C., Kwon, Y.Min.
<2>Improving the Efficiency of Transposon Mutagenesis in Salmonella Enteritidis by Overcoming Host-Restriction Barriers.
<3>Mol. Biotechnol.
<4>56
<5>1004-1010
<6>2014
<7>Transposon mutagenesis using transposome complex is a powerful method for functional genomics
analysis in diverse bacteria by creating a large number of random mutants to prepare a
genome-saturating mutant library. However, strong host restriction barriers can lead to
limitations with species- or strain-specific restriction-modification systems. The purpose of
this study was to enhance the transposon mutagenesis efficiency of Salmonella Enteritidis to
generate a larger number of random insertion mutants. Host-adapted Tn5 DNA was used to form a
transposome complex, and this simple approach significantly and consistently improved the
efficiency of transposon mutagenesis, resulting in a 46-fold increase in the efficiency as
compared to non-adapted transposon DNA fragments. Random nature of Tn5 insertions was
confirmed by high-throughput sequencing of the Tn5-junction sequences. The result based on S.
Enteritidis in this study should find broad applications in preparing a comprehensive mutant
library of other species using transposome complex.

<>

<1>Day, J.P., Reich, N.O., Hager, P., Rosenberg, J., McClarin, J.A., Grable, J., Boyer, H.W., Greene, P.J.
<2>Characterization of three EcoRI endonuclease mutants with altered DNA-Protein contacts designed to change specificity.
<3>J. Cell Biochem. Suppl.
<4>11C
<5>205
<6>1987
<7>Mutants of the EcoRI endonuclease were created by site directed mutagenesis in
an attempt to alter specificity.  The mutations were chosen, based on the
crystal structure, to change the hydrogen bonding characteristics of the amino
acid side chains responsible for recognition of the canonical DNA sequence.
The mutants generated were Glu144->Asp(ED144), Arg145->Lys(RK145), and
Arg200->Lys(RK200).  These were purified to homogeneity.  We measured kcat and
Km on pBR322 and pBR322 lacking the RI site.  We also determined the salt and
pH optima.  All of the mutants cleave the canonical sequence; however, they all
accumulate the nicked intermediate.  Linear DNA production by ED144 is
increased at low salt to a nearly wildtype level.  kcat values are well below
wildtype kcat in RK145 and RK200.  The specificity constants (kcat/Km) are
different from wildtype.  Canonical to noncanonical kcat/Km ratios indicate
altered specificity.  Ed144 has increased specificity for the canonical site.
The rate limiting step in the wildtype is the off rate from nonspecific DNA
after cleavage.  If this step is also limitiing in the mutants, then hydrogen
bonds involved in recognition of the canonical site are normally made to some
extent in nonspecific DNA.  Completely changing specificity is difficult or
perhaps impossible to accomplish by making a single amino acid change.  One
might expect enzymes to have a structure resistsant to specificity changes by a
single amino acid substitution, especially if a specificity change would be a
lethal event.

<>

<1>Day, M., Ibrahim, M., Dyer, D., Bulla, L.Jr.
<2>Genome Sequence of Bacillus thuringiensis subsp. kurstaki Strain HD-1.
<3>Genome Announcements
<4>2
<5>e00613-14
<6>2014
<7>We report here the complete genome sequence of Bacillus thuringiensis subsp. kurstaki strain
HD-1, which serves as the primary U.S. reference standard for all
commercial insecticidal formulations of B. thuringiensis manufactured around the
world.

<>

<1>Day, R.S.
<2>UV-induced alleviation of K-specific restriction of bacteriophage lambda.
<3>J. Virol.
<4>21
<5>1249-1251
<6>1977
<7>A partial release of K-specific restriction of phage lambda grown in
Escherichia coli C was observed when E. coli K strains AB1157 (having wild-type
repair of UV-produced DNA damage) and AB1886 (uvrA) were irradiated with UV
light before infection.  The effect occurred in AB1886 at lower UV fluences
than it did in AB1157.  Little or no release of restriction was observed when
AB2463 (recA) or AB2494 (lex-1) was used.  Such release of restriction appears
to be another of the UV-induced phenomena associated with SOS repair.

<>

<1>De Backer, O., Colson, C.
<2>Molecular nature of the restriction-modification system LT of Salmonella typhimurium.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>212
<6>1989
<7>Restriction-modification (R-M) system LT is present in most Salmonella strains.
Its genes are chromosomally located near proC.  We have cloned these genes and
determined some properties of the coded enzymes.  On appropriate S. typhimurium
strains, the phage vector lambda EMBL4 was very strongly restricted by LT.
This allowed the selection of a lambda clone carrying the modification gene and
therefore immune to the LT restriction.  This gene was subcloned into plasmid
vectors and expressed in E. coli.  A restricting recombinant clone was isolated
from a plasmid genomic library of S. typhimurium made in a modifying host
strain.  This clone proved to contain a plasmid harboring the genes coding for
both restriction and modification activities.  In contrast with other studied
R-M systems, the introduction of LT genes into a nonmodifying host is lethal,
probably because of self-restriction of the chromosomal DNA.  The sequence of
the recognition site of the LT enzymes was found to be 5' GAGAC 3'.  It is
characteristic of type III R-M systems (5 bp long, asymmetric, adenine present
on only one strand).  The methylated base is the 5' adenine.

<>

<1>De Backer, O., Colson, C.
<2>Transfer of the genes for the StyLTI restriction-modification system of Salmonella typhimurium to strains lacking modification ability results in death of the recipient cells and degradation of their DNA.
<3>J. Bacteriol.
<4>173
<5>1328-1330
<6>1991
<7>The genes encoding the restriction-modification system, StyLTI, of Salmonella
typhimurium were inserted in vivo into the conjugative plasmid pULB21.  This
allowed us to transfer the StyLTI genes at a very high frequency and to monitor
the fate of recipient cells after mating.  Transfer of the StyLTI restriction
and modification genes into a modificationless recipient was lethal and
resulted in degradation of the cell's DNA.  This indicates that, in contrast to
any other known restriction-modification systems, StyLTI cannot be established
after horizontal transfer into a naive host.

<>

<1>De Backer, O., Colson, C.
<2>Two-step cloning and expression in Escherichia coli of the DNA restriction-modification system StyLTI of Salmonella typhimurium.
<3>J. Bacteriol.
<4>173
<5>1321-1327
<6>1991
<7>The StyLTI restriction-modification system is common to most strains of the
genus Salmonella, including Salmonella typhimurium.  We report here the
two-step cloning of the genes controlling the StyLTI system.  The StyLTI
methylase gene (mod) was cloned first.  Then, the companion endonuclease gene
(res) was introduced on a compatible vector.  A strain of S. typhimurium
sensitive to the coliphage lambda was constructed and used to select
self-modifying recombinant phages from a Res-Mod+ S. typhimurium genomic
library in the lambda EMBL4 cloning vector.  The methylase gene of one of these
phages was then subcloned in pBR328 and transferred into Escherichia coli.  In
the second step, the closely linked endonuclease and methylase genes were
cloned together on a single DNA fragment inserted in pACYC184 and introduced
into the Mod+ E. coli strain obtained in the first step.  Attemps to transform
Mod- E. coli or S. typhimurium strains with this Res+ Mod+ plasmid were
unsuccessful, whereas transformation of Mod+ strains occured at a normal
frequency.  This can be understood if the introduction of the StyLTI genes into
naive hosts is lethal because of degradation of host DNA by restriction
activity; in contrast to most restriction-modification systems, StyLTI could
not be transferred into naive hosts without killing them.  In addition, it was
found that strains containing only the res gene are viable and lack restriction
activity in the absence of the companion mod gene.  This suggests that
expression of the StyLTI endonuclease activity requires at least one
polypeptide involved in the methylation activity, as is the case for types I
and III restriction-modification systems but not for type II systems.

<>

<1>De Backer, O., Colson, C.
<2>Identification of the recognition sequence for the M.StyLTI methyltransferase of Salmonella typhimurium LT7: an asymmetric site typical of type-III enzymes.
<3>Gene
<4>97
<5>103-107
<6>1991
<7>The StyLTI restriction-modification (R-M) system is encoded by chromosomal
genes of Salmonella typhimurium LT7.  We report here the identification of the
nucleotide (nt) sequence methylated by the StyLTI modification
methyltransferase (M.StyLTI).  This enzyme was partially purified from an
Escherichia coli strain expressing the cloned M.StyLTI-encoding gene, but
lacking StyLTI restriction activity, and used to methylate DNAs of known
sequence, using S-adenosyl-[methyl-3H]-methionine as the methyl donor.  The
[3H]methylated DNA was then digested with various endonucleases.  Examination
of labelled and unlabelled restriction fragments allowed us to map the M.StyLTI
sites in perfectly defined regions of the DNA.  Comparison of the nt sequences
of DNA segments with or without M.StyLTI sites permitted us to identify the
asymmetric and nondegenerate pentanucleotide, 5'-CAG(A*)G-3'/3'-GTCTC-5', as
the StyLTI sequence.  M.StyLTI was found to methylate only the 3' A (see
asterisk) in the upper strand of this sequence.  Thus, M.StyLTI recognises and
methylates the DNA in a manner very similar to that of the three known type-III
MTases, M.EcoPI, M.EcoP15, and M.HinfIII.  This strongly suggests that StyLTI
constitutes a fourth type-III R-M system.

<>

<1>De Bolle, X., Bayliss, C.D., Field, D., van de Ven, T., Saunders, N.J., Hood, D.W., Moxon, E.R.
<2>The length of a tetranucleotide repeat tract in Haemophilus influenzae determines the phase variation rate of a gene with homology to type III DNA methyltransferases.
<3>Mol. Microbiol.
<4>35
<5>211-222
<6>2000
<7>Haemophilus influenzae is an obligate commensal of the upper respiratory tract of humans that
uses simple repeats (microsatellites) to alter gene expression. The mod gene of H. influenzae
strain Rd has homology to DNA methyltransferases of type III restriction/modification systems
and has 40 tetranucleotide (5'-AGTC) repeats within its open reading frame. This gene was
found in 21 out of 23 genetically distinct H. influenzae strains, and in 13 of these strains
the locus contained repeats. H. influenzae strains were constructed in which a lacZ reporter
was fused to a chromosomal copy of mod downstream of the repeats. Phase variation occurred at
a high frequency in strains with the wild-type number of repeats. Mutation rates were derived
for similarly engineered strains, containing different numbers of repeats. Rates increased
linearly with tract length over the range 17-38 repeat units. The majority of tract
alterations were insertions or deletions of one repeat unit with a 2:1 bias towards
contractions of the tract. These results demonstrate the number of repeats to be an important
determinant of phase variation rate in H. influenzae for a gene containing a microsatellite.

<>

<1>De Feyter, R., Gabriel, D.W.
<2>Use of cloned DNA methylase genes to increase the frequency of transfer of foreign genes into Xanthomonas campestris pv. malvacearum.
<3>J. Bacteriol.
<4>173
<5>6421-6427
<6>1991
<7>In vitro-packaged cosmid libraries of DNA from the bacterium Xanthomonas
campestris pv. malvacearum were restricted 200- to 1,000-fold when introduced
into Mcr+ strains of Escherichia coli compared with restriction in the Mcr-
strain HB101.  Restriction was predominantly associated with the mcrBC+ gene in
E. coli.  A plasmid (pUFR052) encoding the XmaI and XmaIII DNA methylases was
isolated from an X. campestris pv. malvacearum library by a screening procedure
utilizing Mcr+ and Mcr- E. coli strains.  Transfer of plasmids from E. coli
strains to X. campestris pv. malvacearum by conjugation was enhanced by up to
five orders of magnitude when the donor cells contained pUFR052 as well as the
plasmid to be transferred.  Subcloning of pUFR052 revealed that at least two
regions of the plasmid were required for full modification activity.  Use of
such modifier plasmids is a simple, novel method that may allow the efficient
introduction of genes into any organism in which restriction systems provide a
potent barrier to such gene transfer.

<>

<1>De Groot, A., Dulermo, R., Ortet, P., Blanchard, L., Guerin, P., Fernandez, B., Vacherie, B., Dossat, C., Jolivet, E., Siguier, P., Chandler, M., Barakat, M., Dedieu, A., Barbe, V., Heulin, T., Sommer, S., Achouak, W., Jean, A.
<2>Alliance of Proteomics and Genomics to Unravel the Specificities of Sahara Bacterium.
<3>PLoS Genet.
<4>5
<5>e1000434
<6>2009
<7>To better understand adaptation to harsh conditions encountered in hot arid deserts, we report
the first complete genome sequence and proteome analysis of a bacterium, Deinococcus deserti
VCD115, isolated from Sahara surface sand. Its genome consists of a 2.8-Mb chromosome and
three large plasmids of 324 kb, 314 kb, and 396 kb. Accurate primary genome annotation of its
3,455 genes was guided by extensive proteome shotgun analysis. From the large corpus of MS/MS
spectra recorded, 1,348 proteins were uncovered and semiquantified by spectral counting. Among
the highly detected proteins are several orphans and Deinococcus-specific proteins of unknown
function. The alliance of proteomics and genomics highthroughput techniques allowed
identification of 15 unpredicted genes and, surprisingly, reversal of incorrectly predicted
orientation of 11 genes. Reversal of orientation of two Deinococcus-specific radiation-induced
genes, ddrC and ddrH, and
identification in D. deserti of supplementary genes involved in manganese import extend our
knowledge of the radiotolerance toolbox of Deinococcaceae. Additional genes involved in
nutrient import and in DNA repair (i.e., two extra recA, three translesion DNA polymerases, a
photolyase) were also identified and found to be expressed under standard growth conditions,
and, for these DNA repair genes, after exposure of the cells to UV. The supplementary nutrient
import and DNA repair genes are likely important for survival and adaptation of D. deserti to
its nutrient-poor, dry, and UV-exposed extreme environment.

<>

<1>De Jonckheere, J.F.
<2>Evidence for the ancestral origin of group I introns in the SSUrDNA of Naegleria spp.
<3>J. Eukaryot. Microbiol.
<4>41
<5>457-463
<6>1994
<7>The sequence variation within the group I intron in five Naegleria spp. was studied and
compared with the sequence variation within the flanking small subunit ribosomal DNA.
Considerable sequence divergence was observed in the introns as well as in the rDNA.  In the
intron deletions and insertions are only detected in the sequence contributing to the
secondary structure, not in the open reading frame.  Most of the sequence variation is
detected in the unpaired loops.  In the case of nucleotide substitution in helices,
compensating base pair changes were observed.  The sequence variation does not induce
variation in the secondary structure model.  The phylogenetic tree based on the intron
sequences is similar to the tree based on the flanking rDNA sequences.  This observation
indicates that the intron might have been acquired at an early stage in evolution, and lost in
the majority of Naegleria spp.

<>

<1>de la Campa, A.G., Kale, P., Springhorn, S.S., Lacks, S.A.
<2>Proteins encoded by the DpnII restriction gene cassette:  Two methylases and an endonuclease.
<3>J. Mol. Biol.
<4>196
<5>457-469
<6>1987
<7>Proteins encoded by three genes in the DpnII restriction enzyme cassette of
Streptococcus pneumoniae were purified and characterized.  Large amounts of the
proteins were produced by subcloning the cassette in an Escherichia coli
expression system.  All three proteins appear to be dimers composed of
identical polypeptide subunits.  One is the DpnII endonuclease, and the other
two are DNA adenine methylases active at 5'GATC3' sites.  Inactivation of
enzyme activity by insertions into the genes and comparison of the DNA sequence
with the amino-terminal sequence of amino acid residues in the proteins
demonstrated the following correspondence between genes and enzymes.  The
promoter-proximal gene in the operon, dpnM, encodes a 33000 Mr polypeptide that
gives rise to a potent DNA methylase.  The next gene, dpnA, encodes the 31000
Mr polypeptide of a weaker and less-specific methylase.  The third gene, dpnB,
encodes the 34000 Mr polypeptide of the endonuclease.  Although the
endonuclease polypeptide is initiated from an ordinary ribosome-binding site,
each of the methylase polypeptides begins at an atypical site with a consensus
sequence entirely different from that of Shine & Dalgarno.  This presumptive
novel ribosome-binding site is well recognized in both S. pneumoniae and E.
coli.

<>

<1>de la Campa, A.G., Springhorn, S.S., Kale, P., Lacks, S.A.
<2>Proteins encoded by the DpnI restriction gene cassette.
<3>J. Biol. Chem.
<4>263
<5>14696-14702
<6>1988
<7>Insertion mutations in the DpnI gene cassette of Streptococcus pneumoniae
indicated that the two genes it contains, dpnC and dpnD, were transcribed from
an adjacent promoter and that only dpnC was necessary for expression of the
DpnI endonuclease.  Large amounts of the DpnI endonuclease were produced from
the cloned cassette in an Escherichia coli expression system, and the enzyme
was purified to homogeneity.  The DpnI endonuclease is composed of a single
polypeptide of 30 kDa, which, as shown by NH2-terminal sequencing of the
protein, is encoded by the entire dpnC open reading frame.  the native protein
sedimented as a monomer of 30 kDa in 0.5 M NaCl.  A protein composed of a
20-kDa polypeptide, which is presumably encoded by dpnD, was also produced in
large amounts.  It was partially purified, but its function is unknown.
Examination of the predicted amino acid sequence of DpnI revealed a potential
metal-containing, DNA-binding finger structure.  It is suggested that this
structure provides the specificity for recognition of the methylated DNA
sequence, 5'-GmATC-3', that is cleaved by the DpnI endonuclease.

<>

<1>de la Haba, R.R., Sanchez-Porro, C., Leon, M.J., Papke, R.T., Ventosa, A.
<2>Draft Genome Sequence of the Moderately Halophilic Bacterium Pseudoalteromonas ruthenica Strain CP76.
<3>Genome Announcements
<4>1
<5>e00268-13
<6>2013
<7>Pseudoalteromonas ruthenica strain CP76, isolated from a saltern in Spain, is a moderately
halophilic bacterium belonging to the Gammaproteobacteria. Here we
report the draft genome sequence, which consists of a 4.0-Mb chromosome, of this
strain, which is able to produce the extracellular enzyme haloprotease CPI.

<>

<1>De Lencastre, H., Wu, S.W., Pinho, M.G., Ludovice, A.M., Filipe, S., Gardete, S., Sobral, R., Gill, S., Chung, M., Tomasz, A.
<2>Antibiotic resistance as a stress response: complete sequencing of a large number of chromosomal loci in Staphylococcus aureus strain COL that impact   on the expression of resistance to methicillin.
<3>Microb. Drug Resist.
<4>5
<5>163-175
<6>1999
<7>Tn551 inactivation has identified several determinants--fem or auxiliary genes--that, in
addition to the mecA gene, are also critical for the
expression of high-level and homogeneous resistance to methicillin.
Genetic and/or biochemical analysis has shown that of the nearly dozen aux
mutations described so far most are in genes involved in cell wall
synthesis (murE, pbp2, glmM, glnR, femA/B, llm, etc.) or in complex
regulatory functions (sigmaB), suggesting that optimal expression of
resistance may involve the cooperative functioning of a number of genes in
cell wall metabolism as well as stress response. The exact mechanism of
these functions is not known. In an attempt to explore this unusual aspect
of methicillin resistance more fully, a Tn551 transposon library,
constructed in the background of the highly and homogeneously
methicillin-resistant Staphylococcus aureus strain COL, was screened for
all independent insertional mutants in which the level of methicillin
resistance of the parental strain (MIC, 1,600 microg/ml) was reduced by at
least 15-fold and up to 500-fold. We now describe the sequencing of 21
Tn551-inactivated genes and their vicinities in 23 new auxiliary mutants
that have been studied before. Using the inverted polymerase chain
reaction (IPCR), we amplified fragments corresponding to the right and
left junction of the Tn551 insertions, which were then sequenced by primer
walking. The two largest groups of these new auxiliary genes encoded
either proteins of unknown functions (6 genes) or showed homology with
genes encoding proteins involved with putative sensory/regulatory
activities (7 genes: protein kinases, ABC transporters, and a catabolite
control protein). Sequencing upstream and downstream allowed the
identification of a number of additional open reading frames, some of
which may also include functions relevant for the expression of antibiotic
resistance.

<>

<1>De Leon, K.B., Young, M.L., Camilleri, L.B., Brown, S.D., Skerker, J.M., Deutschbauer, A.M., Arkin, A.P., Fields, M.W.
<2>Draft Genome Sequence of Pelosinus fermentans JBW45, Isolated during In Situ Stimulation for Cr(VI) Reduction.
<3>J. Bacteriol.
<4>194
<5>5456-5457
<6>2012
<7>Pelosinus fermentans JBW45 is an anaerobic, lactate-fermenting bacterium isolated from
Cr(VI)-contaminated groundwater at the Hanford Nuclear Reservation 100-H site (Washington)
that was collected after stimulation with a polylactate compound. The genome sequence of this
organism will provide insight into the metabolic potential of a predominant population during
stimulation for metal-reducing conditions.

<>

<1>de Lima-Morales, D., Chaves-Moreno, D., Jarek, M., Vilchez-Vargas, R., Jauregui, R., Pieper, D.H.
<2>Draft Genome Sequence of Pseudomonas veronii Strain 1YdBTEX2.
<3>Genome Announcements
<4>1
<5>e00258-13
<6>2013
<7>Pseudomonas veronii strain 1YdBTEX2 was isolated from a benzene-contaminated site. Here we
report the draft genome sequence of 1YdBTEX2 and its genes
associated with aromatic metabolism. The broad catabolic potential of this strain
is consistent with the environment from which it was isolated.

<>

<1>de los Reyes-Gavilan, C.G., Aparicio, J.F., Barbes, C., Hardisson, C., Sanchez, J.
<2>An exocytoplasmic endonuclease with restriction function in Streptomyces antibioticus.
<3>J. Bacteriol.
<4>170
<5>1339-1345
<6>1988
<7>Streptomyces antibioticus produces a strong endo-DNase which is located between
the cytoplasmic membrane and the cell wall.  All DNA substrates assayed,
including the chromosomal DNA of this species and several bacteriophage DNAs,
were completely degraded in vitro by the enzyme.  The rate of synthesis of the
nuclease depended on the growth medium.  In NBG medium, in which the enzyme is
not produced, the size of lytic plaques of several actinophages was larger than
that in GYM or GAE medium, in which synthesis of the nuclease takes place late
in growth.  In addition, one of the phages assayed, PhiA6, showed a diminution
of its efficiency of plating in GYM medium with respect to that in NBG medium;
another phage, PhiA9, grew in NBG medium but not in the other two media.  It is
postulated that the presence of the host nuclease, together with the capability
of the particular phage to adsorb on S. antibioticus of different growth
phases, determines the efficiency of growth and the plaque size of the phages
on productive media.  This hypothesis was confirmed when the growth of PhiA6
and PhiA9 in a mutant of S. antibioticus lacking the endonuclease activity was
analyzed.  It is concluded that the enzyme can assume, under some
circumstances, a role in in vivo restriction.

<>

<1>de los Reyes-Gavilan, C.G., Limsowtin, G.K.Y., Sechaud, L., Veaux, M., Accolas, J.-P.
<2>Evidence for a plasmid-linked restriction-modification system in Lactobacillus helveticus.
<3>Appl. Environ. Microbiol.
<4>56
<5>3412-3419
<6>1990
<7>The presence of a restriction-modification (R/M) system against two
bacteriophages, 328-B1 and hv, was demonstrated in three Lactobacillus
helveticus strains, CNRZ 1094, CNRZ 1095, and CNRZ 1096.  In addition, the
burst size of phage 328-B1 in the three restrictive strains CNRZ 1094, CNRZ
1095, and CNRZ 1096 was reduced with respect to the values obtained in its
propagating strain, CNRZ 328.  Heating at 60C did not inactivate the R/M
system.  Nonrestrictive variants from CNRZ 1094 were easily obtained under
several culture conditions, but treatment with novobiocin at 42C followed by
storage at -20C resulted in drastic elimination of the R+/M+ phenotype from all
clones tested.  Electrophoretic analysis of CNRZ 1084 nonrestrictive variants
revealed the concomitant loss of a 34-kb plasmid.  Four EcoRI fragments from
the 34-kb plasmid were cloned in the Escherichia coli vector pACYC184.  The use
of one or several of these fragments as probes confirmed the plasmidic location
of the genes responsible for the R/M system.  These probes also showed the
presence of R/M plasmids in the two other restriction strains, CNRZ 1095 and
CNRZ 1096.  Lactose-fermenting ability and/or proteolytic capacity was not
linked to the 34-kb plasmid.

<>

<1>De Maayer, P., Chan, W.Y., Rezzonico, F., Buhlmann, A., Venter, S.N., Blom, J., Goesmann, A., Frey, J.E., Smits, T.H., Duffy, B., Coutinho, T.A.
<2>Complete Genome Sequence of Clinical Isolate Pantoea ananatis LMG 5342.
<3>J. Bacteriol.
<4>194
<5>1615-1616
<6>2012
<7>The enterobacterium Pantoea ananatis is an ecologically versatile species. It has been found
in the environment, as plant epiphyte and endophyte, as an emerging
phytopathogen, and as a presumptive, opportunistic human pathogen. Here, we
report the complete genome sequence of P. ananatis LMG 5342, isolated from a
human wound.

<>

<1>De Maayer, P., Chan, W.Y., Venter, S.N., Toth, I.K., Birch, P.R., Joubert, F., Coutinho, T.A.
<2>Genome sequence of Pantoea ananatis LMG20103, the causative agent of Eucalyptus blight and dieback.
<3>J. Bacteriol.
<4>192
<5>2936-2937
<6>2010
<7>Pantoea ananatis is a Gram-negative plant pathogen that causes disease on a broad range of
host plants, including pineapple, maize, rice, onion, melons, and Eucalyptus, and has been
implicated in several cases of human disease. Here, we report the genome sequence of P.
ananatis LMG20103
isolated from diseased Eucalyptus in South Africa.

<>

<1>De Maayer, P., Williamson, C.E., Vennard, C.T., Danson, M.J., Cowan, D.A.
<2>Draft Genome Sequences of Geobacillus sp. Strains CAMR5420 and CAMR12739.
<3>Genome Announcements
<4>2
<5>e00567-14
<6>2014
<7>Thermophilic Geobacillus spp. can efficiently hydrolyze hemicellulose polymers and are
therefore of interest in biotechnological applications. Here we report
the genome sequences of two hemicellulolytic strains, Geobacillus sp. CAMR12739
and CAMR5420.

<>

<1>de Oliveira, L.G., Tormet, G.G.D., Samborsky, M., Marcon, J., Araujo, W.L., de Azevedo, J.L.
<2>Genome Sequence of Streptomyces wadayamensis Strain A23, an Endophytic Actinobacterium from Citrus reticulata.
<3>Genome Announcements
<4>2
<5>e00625-14
<6>2014
<7>The actinobacterium Streptomyces wadayamensis A23 is an endophyte of Citrus reticulata that
produces the antimycin and mannopeptimycin antibiotics, among
others. The strain has the capability to inhibit Xylella fastidiosa growth. The
draft genome of S. wadayamensis A23 has ~7.0 Mb and 6,006 protein-coding
sequences, with a 73.5% G+C content.

<>

<1>de Padua-Pereira, U. et al.
<2>Complete genome sequence of Streptococcus agalactiae strain SA20-06, a fish pathogen associated to meningoencephalitis outbreaks.
<3>Standards in Genomic Sciences
<4>8
<5>188-197
<6>2013
<7>Streptococcus agalactiae (Lancefield group B; GBS) is the causative agent of
meningoencephalitis in fish, mastitis in cows, and neonatal sepsis in humans.
Meningoencephalitis is a major health problem for tilapia farming and is
responsible for high economic losses worldwide. Despite its importance, the
genomic characteristics and the main molecular mechanisms involved in virulence
of S. agalactiae isolated from fish are still poorly understood. Here, we present
the genomic features of the 1,820,886 bp long complete genome sequence of S.
agalactiae SA20-06 isolated from a meningoencephalitis outbreak in Nile tilapia
(Oreochromis niloticus) from Brazil, and its annotation, consisting of 1,710
protein-coding genes (excluding pseudogenes), 7 rRNA operons, 79 tRNA genes and
62 pseudogenes.

<>

<1>de Sa, P.C., Da Silva, M.L., Carneiro, A.R., Gomes, J.C., Dias, L.M., Alves, J.T., De Oliveira, V.A.A., Barauna, R.A., Das Gracas, D.A., Matte, M.H., Sato, M.I., Hachich, E.M., Matte, G.R., Ramos, R.T., Silva, A.
<2>Draft Genome Sequence of Non-O1 and Non-O139 Vibrio cholerae Strain VCC19.
<3>Genome Announcements
<4>2
<5>e01094-14
<6>2014
<7>Vibrio cholerae O1 is the causative agent of cholera and is ubiquitous in the aquatic
environment, while V. cholerae strains non-O1 and non-O139 are recognized
as causative agents of sporadic and localized outbreaks of diarrhea. Here, we
report the complete sequence of a non-O1 and non-O139 V. cholerae strain (VCC19),
which was isolated from the environment in Brazil. The sequence includes the
integrative conjugative element (ICE). This paper is the first report of the
presence of such an element in a V. cholerae strain isolated in Brazil.

<>

<1>de Souza, J.A., Tieppo, E., Magnani, G.S., Alves, L.M., Cardoso, R.L., Cruz, L.M., de Oliveira, L.F., Raittz, R.T., de Souza, E.M., Pedrosa, F.O., Lemos, E.G.
<2>Draft Genome Sequence of the Nitrogen-Fixing Symbiotic Bacterium Bradyrhizobium elkanii 587.
<3>J. Bacteriol.
<4>194
<5>3547-3548
<6>2012
<7>The draft sequence of the genome of Bradyrhizobium elkanii 587 is presented. This was obtained
using Illumina Next-Gen DNA sequencing combined with Sanger
sequencing. Genes for the pathways involved in biological nitrogen fixation (the
nif gene cluster), nod genes including nodABC, and genes for the type III protein
secretion system (T3SS) are present.

<>

<1>de Souza, V., Piro, V.C., Faoro, H., Tadra-Sfeir, M.Z., Chicora, V.K., Guizelini, D., Weiss, V., Vialle, R.A., Monteiro, R.A., Steffens, M.B., Marchaukoski, J.N., Pedrosa, F.O., Cruz, L.M., Chubatsu, L.S., Raittz, R.T.
<2>Draft Genome Sequence of Herbaspirillum huttiense subsp. putei IAM 15032, a Strain Isolated from Well Water.
<3>Genome Announcements
<4>1
<5>e00252-12
<6>2013
<7>Here we report the one-scaffold draft genome of subsp. strain 7-2 (IAM 15032), which was
isolated from well water.

<>

<1>de Vries, G.E., Harms, N., Hoogendijk, J., Stouthamer, A.H.
<2>Isolation and characterization of Paracoccus denitrificans mutants with increased conjugation frequencies and pleiotropic loss of a (nGATCn) DNA-modifying property.
<3>Arch. Microbiol.
<4>152
<5>52-57
<6>1989
<7>A selection scheme was devised to isolate Paracoccus denitrificans mutants with
increased recipient qualities in transfer experiments, using broad host range
plasmids.  In some of the mutants obtained, a DNA modifying activity that
prevents the activity of the restriction endonucleases BamHI and BglII on
isolated P. denitrificans DNA had simultaneously been lost.  From a detailed
analysis of the restriction properties of the enzymes Sau3AI, MboI and DpnI, it
was concluded that a subset of GATC sequences in P. denitrificans DNA may be
methylated at an unusual position.  It was concluded that P. denitrificans
possesses at least one potent host-dependent restriction/modification system
which affects conjugation.  In addition to the class of enhanced transfer
mutants, at least one other class of enhanced transfer mutants with unknown
defect(s) was isolated.  Strains, in which the two mutant classes were
combined, exhibited transfer frequencies which were significantly higher than
strains containing either mutation alone.  Such double mutant strains appeared
to be well suited for future experiments like complementation analysis,
transposon mutagenesis and gene replacement by homologous recombination.

<>

<1>De Vries, N., Duinsbergen, D., Kuipers, E.J., Pot, R.G.J., Wiesenekker, P., Penn, C.W., Van Vliet, A.H.M., Vandenbroucke-Grauls, C.M.J.E., Kusters, J.G.
<2>Transcriptional phase variation of a Type III restriction-modification system in Helicobacter pylori.
<3>J. Bacteriol.
<4>184
<5>6615-6623
<6>2002
<7>Phase variation is important in bacterial pathogenesis, since it generates
antigenic variation for the evasion of immune responses and provides a
strategy for quick adaptation to environmental changes. In this study, a
Helicobacter pylori clone, designated MOD525, was identified that
displayed phase-variable lacZ expression. The clone contained a
transcriptional lacZ fusion in a putative type III DNA methyltransferase
gene (mod, a homolog of the gene JHP1296 of strain J99), organized in an
operon-like structure with a putative type III restriction endonuclease
gene (res, a homolog of the gene JHP1297), located directly upstream of
it. This putative type III restriction-modification system was common in
H. pylori, as it was present in 15 out of 16 clinical isolates. Phase
variation of the mod gene occurred at the transcriptional level both in
clone MOD525 and in the parental H. pylori strain 1061. Further analysis
showed that the res gene also displayed transcriptional phase variation
and that it was cotranscribed with the mod gene. A homopolymeric cytosine
tract (C tract) was present in the 5' coding region of the res gene.
Length variation of this C tract caused the res open reading frame (ORF)
to shift in and out of frame, switching the res gene on and off at the
translational level. Surprisingly, the presence of an intact res ORF was
positively correlated with active transcription of the downstream mod
gene. Moreover, the C tract was required for the occurrence of
transcriptional phase variation. Our finding that translation and
transcription are linked during phase variation through slipped-strand
mispairing is new for H. pylori.

<>

<1>de Vries, N., Duinsbergen, D., Kuipers, E.J., Wiesenekker, P., Vandenbroucke-Grauls, C.M., Kusters, J.G.
<2>Phase variation in a type III restriction-modification system of Helicobacter pylori.
<3>Gastroenterology
<4>118
<5>A736
<6>2000
<7>The on- and off-switching of the expression of virulence factors (phase variation) plays an
important role in the pathogenesis of many bacterial infections.  LPS phase variation in
Helicobacter pylori occurs at the translational level and has been studied well.  In contrast,
phase variation at the transcriptional level has not been demonstrated in H. pylori.
Therefore, we investigated transcriptional on- and off-switching of gene expression in H.
pylori.  Methods: A library with random genomic transcriptional lacZ fusions in H. pylori
strain 1061 (HP1061) was screened for mutants that showed blue and white sectored colonies on
X-Gal.  As the X-Gal substrate is converted into a blue product when the lacZ gene is
expressed into the beta-galactosidase, sectored colonies indicate transcriptional phase
variation.  Results: One HP1061 mutant displayed frequent on- and off-switching of lacZ
expression.  The on-to-off switch frequency was 2.67%, while the off-to-on frequency was
0.75%.  Sequencing revealed that the lacZ gene was fused to a putative type III methylase gene
(mod).  RNA spot blot hybridization demonstrated that specific lacZ and mod probes bound to
mRNA from blue colonies, but not to mRNA from white colonies.  This proved that mod switched
on and off a the transcriptional level.  An open reading frame, encoding a putative type III
restriction enzyme gene (res), is located immediately upstream of mod and contains a
polynucleotide C-tract.  This C-tract, which may cause phase variation of res through
translational frameshifts, might indirectly act on the transcription of mod.  However,
sequence analysis showed that the number of cytosines in res was not related to the on- or
off-status of mod. Conclusion: In H. pylori, the putative type III methylase gene, mod,
displays phase variation at the transcriptional level.  It is known that methylation of
promoter sequences can affect the transcription of bacterial virulence factors.  We propose a
specific role of mod and related restriction-modification systems in H. pylori in the
regulation of virulence genes.

<>

<1>de Vries, N., Duinsbergen, D., Kuipers, E.J., Wiesenekker, P., Vandenbroucke-Grauls, C.M.J., Kusters, J.G.
<2>Transcriptional phase variation in a type III restriction modification system of Helicobacter pylori.
<3>Gut
<4>47
<5>A17
<6>2000
<7>Phase variation of virulence genes is important in the pathogenesis of many bacterial
infections.  So far, phase variation at the transcriptional level has not been demonstrated in
H. pylori.  Here, we report for the first time transcriptional phase variation in H. pylori.
An H. pylori 1061 library with random genomic transcriptional lacZ fusions was screened for
transformants showing blue, white and sectored colonies on X-Gal.  This phenotype indicated a
fusion of lacZ to a gene displaying transcriptional phase variation.  In one transformant
showing sectored colonies, lacZ was inserted in a putative type III methylase gene (mod).  An
endonuclease gene (res) was located immediately upstream of mod and contained a C-tract, which
may cause translational frameshifting.  Blue colonies tended to have 14 Cs, which results in
the translation of res.  In contrast, white colonies contained C-tract lengths leading to
disruption of the open reading frame.  RNA spot blots and RT-PCR indicated that mod displayed
transcriptional phase variation, as mod mRNA was only present in blue colonies, and not in
white colonies.  Res was transcribed both in blue and in white colonies.  In H. pylori 1061 a
type III methylase gene displays transcriptional phase variation.  Translational frameshifting
of the upstream endonuclease gene may be involved in the regulation of mod phase variation.
Since DNA methylation can affect the transcription of bacterial virulence factors, we propose
that mod and related restriction-modification systems play a role in the regulation of the
expression of virulence genes in H. pylori.

<>

<1>de Waard, A., Duyvesteyn, M.
<2>Are sequence-specific deoxyribonucleases of value as taxonomic markers of cyanobacterial species?
<3>Arch. Microbiol.
<4>128
<5>242-247
<6>1980
<7>Three nucleotide sequence-specific deoxyribonucleases present in extracts of
Anabaena subcylindrica have been purified and characterized.  Endo R AsuI
recognizes and cleaves the nucleotide sequence G^GNCC (Hughes et al., 1980)
while Endo R AsuII and III split the sequences TT^CGAA and GPu^CGPyC,
respectively (this paper).  An Anabaena strain "Waterbury" converging
genetically at the 30-35% level with both A. subcylindrica and A. cylindrica
(as judged by DNA-DNA hybridization in vitro) was shown to possess the
endonuclease pattern typical for A. cylindrica (de Waard et al., 1978).  The
usefulness of these specific endonucleases as taxonomic markers for the
classifiction of cyanobacteria is discussed.

<>

<1>de Waard, A., Korsuize, J., van Beveren, C.P., Maat, J.
<2>A new sequence-specific endonuclease from Anabaena cylindrica.
<3>FEBS Lett.
<4>96
<5>106-110
<6>1978
<7>The isolation of sequence-specific endodeoxyribonucleases from the cyanophytes
Anabaena variabilis (ATCC 27892), Anabaena subcylindrica (CCAP 1403/4b) and
Anabaena catenula (CCAP 1403/1) has been reported.  We have found a new
endonuclease from Anabaena cylindrica (CCAP 1403/2a) and describe here the
procedure of its isolation as well as the elucidation of the nucleotide
sequences: 5' G Pu ^ C G Py C 3' 3' C Py G C ^ Pu G 5' recognized by the
purified enzyme AcyI.

<>

<1>de Waard, A., van Beveren, C.P., Duyvesteyn, M., van Ormondt, H.
<2>Two sequence-specific endonucleases from Anabaena oscillarioides.
<3>FEBS Lett.
<4>101
<5>71-76
<6>1979
<7>There has been an increasing number of reports on sequence-specific
endodeoxyribonucleases (endo.R.) in cyanobacteria (blue-green algae).  As their
cleavage specificities have proven to be different from those of the many
bacterial restriction enzymes already characterized, these new enzymes have
been useful additions to the ever expanding endo R. catalog.  We report here
the isolation of two such endonucleases from one strain of Anabaena
oscillarioides (AosI and II) which cleave the nucleotide sequences 5'TGC^GCA3'
and 5'GPu^CGPyC3', respectively.

<>

<1>de Wit, C.M., Dekker, B.M.M., Neele, A.C., de Waard, A.
<2>Purification and characterization of endonucleases DraII and III from Deinococcus radiophilus.
<3>FEBS Lett.
<4>180
<5>219-223
<6>1985
<7>Deinococcus radiophilus strain ATCC 27603 contains, apart from endonuclease
DraI, two additional sequence-specific endonucleases.  These enzymes,
designated DraII and DraIII, recognize nucleotide sequences with novel
specificities, PuG^GNCCPy and CACNNN^GTG, respectively.

<>

<1>Dean, A.B., Stanger, M.J., Dansereau, J.T., Van Roey, P., Derbyshire, V., Belfort, M.
<2>Zinc finger as distance determinant in the flexible linker of intron  endonuclease I-TevI.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>8554-8561
<6>2002
<7>l-Tevl, the phage T4 td intron-encoded endonuclease, recognizes a  lengthy DNA target and
initiates intron mobility by introducing a
double-strand break in the homing site. The enzyme uses both sequence and
distance determinants to cleave the DNA 23-25 bp upstream of the intron
insertion site. l-Tevl consists of an N-terminal catalytic domain and a
C-terminal DNA-binding domain separated by a long, flexible linker. The
DNA-binding domain consists of three subdomains: a zinc finger, a
minor-groove binding a-helix, and a helix-turn-helix. In this study, a
mutational analysis was undertaken to assess the roles of these subdomains
in substrate binding and cleavage. Surprisingly, the zinc finger is not
required for DNA binding or catalysis. Rather, the zinc finger is a
component of the linker and directs the catalytic domain to cleave the
homing site at a fixed distance from the intron insertion site. When the
cleavage site (CS) is shifted outside a given range, wild-type l-Tevl
defaults to the fixed distance, whereas zinc-finger mutants have lost the
distance determinant and search out the displaced cleavage sequences.
Although counterintuitive, a protein containing a 19-aa deletion of the
zinc finger can extend further than can wild-type l-Tevl to cleave a
distant CS sequence, and a Cys-to-Ala mutant of the ligands for zinc,
nominally a longer protein, can retract to cleave at a closer CS sequence.
Models are presented for the novel function of the zinc finger, as a
molecular constraint, whereby intramolecular protein-protein interactions
position the catalytic domain by "catalytic clamp" and/or
"linker-organizer" mechanisms.

<>

<1>Dean, P.D.C., Walker, J.N.B.
<2>Recent advances in high-performance liquid affinity chromatography columns.
<3>Biochem. Soc. Trans.
<4>13
<5>1055-1058
<6>1985
<7>For the initial isolation of restriction endonucleases from crude extracts we
have found that 1 ml of quaternary aminoethyl 'Mono-Q' strong anion-exchange
column is ideal.  All the restriction enzymes we studies were eluted between
0.2 and 0.6m-KCl.  This has enabled us to get to up a rapid screening program
for novel type-II restriction endonuclease activities.  the profile of enzyme
activities from an extract of a cyanobacterium Nostoc SA, incubated with lambda
DNA, shows four separate restriction enzymes of different specificity.  These
enzyme fractions were separately digested with other DNA substrates.  These and
other experiments lead us to conclude that the enzymes were NspSAI
(C-Y-C-G-R-G), NspSAII (G-G-T-N-A-C-C), NspSAIII (C-C-A-T-G-G), NspSAIV
(G-G-A-T-C-C) and were isoschizomers of AvaI, BstEII, NcoI and BamHI,
respectively.  The distribution of cytosines and guanines in each cutting site
is interestingly consistent.  We have found that the purity of these and other
enzymes after a single Mono Q column step is sufficient not only for
characterization of their specificity on a series of substrates, but also to
carry out analysis of the termini of the cutting sites using a modified form of
M13 sequencing.

<>

<1>Deangelis, K.M., D'Haeseleer, P., Chivian, D., Fortney, J.L., Khudyakov, J., Simmons, B., Woo, H., Arkin, A.P., Davenport, K.W., Goodwin, L., Chen, A., Ivanova, N., Kyrpides, N.C., Mavromatis, K., Woyke, T., Hazen, T.C.
<2>Complete genome sequence of 'Enterobacter lignolyticus' SCF1.
<3>Standards in Genomic Sciences
<4>5
<5>69-85
<6>2011
<7>In an effort to discover anaerobic bacteria capable of lignin degradation, we isolated
'Enterobacter lignolyticus' SCF1 on minimal media with alkali lignin as
the sole source of carbon. This organism was isolated anaerobically from tropical
forest soils collected from the Short Cloud Forest site in the El Yunque National
Forest in Puerto Rico, USA, part of the Luquillo Long-Term Ecological Research
Station. At this site, the soils experience strong fluctuations in redox
potential and are net methane producers. Because of its ability to grow on lignin
anaerobically, we sequenced the genome. The genome of 'E. lignolyticus' SCF1 is
4.81 Mbp with no detected plasmids, and includes a relatively small arsenal of
lignocellulolytic carbohydrate active enzymes. Lignin degradation was observed in
culture, and the genome revealed two putative laccases, a putative peroxidase,
and a complete 4-hydroxyphenylacetate degradation pathway encoded in a single
gene cluster.

<>

<1>DeBacker, O., Chomez, P., DePlaen, E.
<2>Positive selection of recombinant plasmids based on the EcoK restriction activity of Escherichia coli K-12.
<3>Gene
<4>150
<5>197-198
<6>1994
<7>We have constructed a pTZ19R-derived vector which allows efficient positive selection of
recombinant plasmids. The system uses the EcoK restriction activity of Escherichia coli K-12
to select against non-recombinant plasmids. The vector contains an EcoK site which, if deleted
or disrupted by ligating a DNA fragment, yields recombinant plasmids that are no longer
suceptible to EcoK restriction when transformed into a restriction-proficient E. coli host.

<>

<1>Debov, S.S., Karyagina, A.S., Lunin, V.G., Lopatina, N.G., Gruber, I.M., Polyachenko, V.M., Nikolskaya-Sanovich, I.I.
<2>Recombinant plasmid DNA d-33 which codes the SsoII methylase - consists of specified vector plasmid DNA and plasmid DNA from Shigella sonnei 47.
<3>Soviet Patent Office
<4>SU 1532585 A
<5>
<6>1989
<7>The recombinant plasmid DNA d33 has mol. wt. 4.5 MDa and contains vector plasmid DNA pUC19,
plasmid DNA P4 with methylase gene SsoII from Shigella sonnei 47, one site for cleavage by
endonucleases ClaI, EcoRV, SacII, PstI, SphI, HindIII, SacI, KpnI, SmaI, XbaI, a cleavage site
for Cfr10I, SalGI, four sites of cleavage for EcoRI, the Bla gene which codes beta-lactamase,
fragments of the lac gene lac with a synthetic polylinker and located on both sides of the
cloned DNA R4.  The SsoII methylase gene located on the cloned DNA plasmid R4 which determines
the synthesis of the SsoII methylase, and the gene for the restriction endonuclease SsoII
consisting of two fragments located on both sides of the DNA of vector pUC19, which thus
cannot determine the synthesis of restriction endonuclease SsoII. Preparation of the
recombinant plasmid DNA d33 involves digesting vector plasmid DNA pUC19 by the restriction
endonuclease BamHI and combining the product by means of DNA-ligase with plasmid DNA P4 that
has been digested by restriction endonuclease BglII. Then the combined fragments are treated
with restriction endonuclease BamHI and transformed in E. coli cells PS200. The transformants
are then grown in a broth of LB medium containing 50 micro.g/ml ampicillin, and the plasmid
DNA d33 is separated.

<>

<1>Debov, S.S., Karyagina, A.S., Lunin, V.G., Lopatina, N.G., Gruber, I.M., Polyachenko, V.M., Nikolskaya-Sanovich, I.I.
<2>Recombinant plasmid DNA - is used for synthesis of restrictase and methylase and is produced by the specified strain of Escherichia coli.
<3>Soviet Patent Office
<4>SU 1539205 A
<5>
<6>1990
<7>Recombinant DNA plasmid d 24 which codes the synthesis of the SsoII restrictase and methylase
more efficiently is present in E. coli GISK-178 which is used as their producer. The plasmid d
24 contains the DNA plasmid of vector pUC19 of 2.69 units size and DNA plasmid from S. sonnei
47 which codes the above ferments, of 4.4 units size, and the strain of E. coli is obtained
by transforming the cells of E.coli B834 with d 24.

<>

<1>Debov, S.S., Nikolskaya, I.I., Lopatina, N.G.
<2>Preparation of modified methylase enzymes from Shigella sonnei - includes chromatography of treated Shigella cells on phenyl-sepharose with reducing ammonium sulphate and increasing triton X-100 concentration.
<3>Soviet Patent Office
<4>SU 1437394
<5>
<6>1988
<7>The process involves culturing Shigella sonnei 47 cells, breaking down the obtained cells
using ultrasound, centrifuging at 105000g, removal of nucleic acids by using Streptomycin
sulphate, salting out the proteins with 80% saturated ammonium sulphate, and subjecting to
hydrophobic chromatography.  The chromatographic stage is carried out on phenyl-sepharose with
the utilization of a combined linear gradient comprising a reducing concentration of ammonium
sulphate from 1.0 to 0.0M and an increasing concentration of Triton X-100 from 0.30 to 0.60%.
Under these conditions, the modifying methylases SsoII and SsoI are eluted at Triton X-100
concentrations of 0.43 and 0.52 wt.%, respectively.  The enzymes obtained in this process do
not contain exo- and endonucleases, as indicated by complete preservation of DNA phage gamma
with long-term incubation of excess amounts of enzymes under conditions assisting the activity
of nucleases (in the presence of magnesium ions).  This allows SsoI and SsoII methylases to be
used in molecular biological experiments on the production of recombinant DNA in vitro.

<>

<1>Debov, S.S., Nikolskaya-Sanovich, I.I., Uporova, T.M., Suchkov, S.V., Kartashova, I.M.
<2>Site-specific endonuclease EcoRI production by disintegrating cells of E. Coli strain RY-13, chromatographing on blue sepharose and isoelectric-focussing.
<3>Soviet Patent Office
<4>SU 1120019 A
<5>
<6>1984
<7>Site-specific endonuclease EcoRI is obtained by: culturing E. coli strain RY13; disintegrating
the cells; and purifying the resulting enzyme by chromatography on blue Sepharose and
isoelectric focussing at pH 3.5-10 on a glycerol density gradient of 0.1-60% recovering the pH
7.7-8.0 fraction.

<>

<1>DeBoy, R.T., Mongodin, E.F., Fouts, D.E., Tailford, L.E., Khouri, H., Emerson, J.B., Mohamoud, Y., Watkins, K., Henrissat, B., Gilbert, H.J., Nelson, K.E.
<2>Insights into plant cell wall degradation from the genome sequence of the soil bacterium Cellvibrio japonicus.
<3>J. Bacteriol.
<4>190
<5>5455-5463
<6>2008
<7>The plant cell wall, which consists of a highly complex array of interconnecting
polysaccharides, is the most abundant source of organic
carbon in the biosphere. Microorganisms that degrade the plant cell wall
synthesize an extensive portfolio of hydrolytic enzymes that display
highly complex molecular architectures. To unravel the intricate
repertoire of plant cell wall-degrading enzymes synthesized by the
saprophytic soil bacterium Cellvibrio japonicus, we sequenced and analyzed
its genome, which predicts that the bacterium contains the complete
repertoire of enzymes required to degrade plant cell wall and storage
polysaccharides. Approximately one-third of these putative proteins (57)
are predicted to contain carbohydrate binding modules derived from 13 of
the 49 known families. Sequence analysis reveals approximately 130
predicted glycoside hydrolases that target the major structural and
storage plant polysaccharides. In common with that of the colonic
prokaryote Bacteroides thetaiotaomicron, the genome of C. japonicus is
predicted to encode a large number of GH43 enzymes, suggesting that the
extensive arabinose decorations appended to pectins and xylans may
represent a major nutrient source, not just for intestinal bacteria but
also for microorganisms that occupy terrestrial ecosystems. The results
presented here predict that C. japonicus possesses an extensive range of
glycoside hydrolases, lyases, and esterases. Most importantly, the genome
of C. japonicus is remarkably similar to that of the gram-negative marine
bacterium, Saccharophagus degradans 2-40(T). Approximately 50% of the
predicted C. japonicus plant-degradative apparatus appears to be shared
with S. degradans, consistent with the utilization of plant-derived
complex carbohydrates as a major substrate by both organisms.

<>

<1>Debroas, D., Humbert, J.F., Enault, F., Bronner, G., Faubladier, F., Cornillot, C.
<2>Metagenomic approach studying the taxonomic and functional diversity of the bacterial community in a mesotrophic lake (Lac du Bourget - France).
<3>Environ. Microbiol.
<4>11
<5>2412-2424
<6>2009
<7>The main goals of this work were to identify the metabolic pathways of the
bacterial community in a lacustrine ecosystem and to establish links
between taxonomic composition and the relative abundances of these
metabolic pathways. For this purpose, we analysed a 16S rRNA gene library
obtained by gene amplification together with a sequence library of both
insert ends on c. 7700 fosmids. Whatever the library used, Actinobacteria
was the most abundant bacterial group, followed by Proteobacteria and
Bacteroidetes. Specific aquatic clades such as acI and acIV
(Actinobacteria) or LD12 and GOBB-C201 (Alphaproteobacteria) were found in
both libraries. From comparative analysis of metagenomic libraries, the
metagenome of this lake was characterized by overrepresentation of genes
involved in the degradation of xenobiotics mainly associated with
Alphaproteobacteria. Actinobacteria were mainly related to metabolic
pathways involved in nucleotide metabolism, cofactors, vitamins, energy,
replication and repair. Betaproteobacteria appeared to be characterized by
the presence of numerous genes implicated in environmental information
processing (membrane transport and signal transduction) whereas glycan and
carbohydrate metabolism pathways were overrepresented in Bacteroidetes.
These results prompted us to propose hypotheses on the ecological role of
these bacterial classes in lacustrine ecosystems.

<>

<1>Debrouwere, L., Zabeau, M., Van Montagu, M., Schell, J.
<2>The ral gene of phage lambda.
<3>Mol. Gen. Genet.
<4>179
<5>75-80
<6>1980
<7>The lambda ral function modulates the restriction and modification activities
of the Escherichia coli K12 and B restriction enzymes (Zabeau et al., 1980).
In order to further analyse this function, ral deficient mutants have been
isolated, using a method which exploits the property of the strong mutagen
N-methyl-N'-nitro-N-nitrosoguanidine (N.G.) to induce multiple closely linked
mutations.  Hence, mutagenized phages carrying mutations in one locus were
frequently found to contain additional mutations in adjacent loci.  This very
efficient mutagenesis procedure enabled us to isolate 27 independent Ral
deficient mutants.  Seven mutants were found to affect the ral gene directly
and were located between the genes N and cIII.  Detailed mapping of two of
these mutants showed that the lambda ral gene is located at position 70.6-70.9%
on the physical map.  The isolation and characterization of these mutants
further supports the conclusion that ral is a gene different from the N gene,
and demonstrates that the ral gene product is responsible for both
counteracting restriction and enhancing modification.

<>

<1>Debroy, S., Bhattacharjee, A., Thakur, A.R., Raychaudhuri, S.
<2>Draft Genome Sequence of the Nitrate- and Phosphate-Accumulating Bacillus sp. Strain MCC0008.
<3>Genome Announcements
<4>1
<5>e00189-12
<6>2013
<7>Here, we report the draft genome sequence of the nitrate- and phosphate-accumulating Bacillus
sp. strain MCC0008, isolated from a consortium
enriched from municipal sewage in nitrate broth (HiMedia M439). The total size of
the genome is 5,609,456 bp, with a G+C content of 35.1%.

<>

<1>Debroy, S., Mukherjee, P., Roy, S., Thakur, A.R., Raychaudhuri, S.
<2>Draft Genome Sequence of a Phosphate-Accumulating Bacillus sp., WBUNB004.
<3>Genome Announcements
<4>1
<5>e00251-12
<6>2013
<7>The draft genome sequence of a nitrate- and phosphate-removing, Gram-positive sp. with optimum
growth at 37 degrees C and pH 7 in nitrate broth (HiMedia M439)
isolated from rhizosphere of a water lily, with a genome size of 5,465,157 bp and
a G+C content of 35.0%, is reported here.

<>

<1>Debroy, S., Mukherjee, P., Roy, S., Thakur, A.R., Raychaudhuri, S.
<2>Draft Genome Sequence of a Nitrate- and Phosphate-Removing Bacillus sp., WBUNB009.
<3>Genome Announcements
<4>1
<5>e00254-12
<6>2013
<7>The draft genome sequence (5,868,741 bp) of a nitrate- and phosphate-removing sp., WBUNB009,
isolated from a raw sewage canal in nitrate broth (Himedia M439)
with a G+C content of 34.9% is reported. It removes 60.23% nitrate and 96%
phosphate within 16 h at 37 degrees C.

<>

<1>Debruyn, J.M., Radosevich, M., Wommack, K.E., Polson, S.W., Hauser, L.J., Fawaz, M.N., Korlach, J., Tsai, Y.C.
<2>Genome Sequence and Methylome of Soil Bacterium Gemmatirosa kalamazoonensis KBS708T, a Member of the Rarely Cultivated Gemmatimonadetes Phylum.
<3>Genome Announcements
<4>2
<5>e00226-14
<6>2014
<7>Bacteria belonging to the phylum Gemmatimonadetes are found in a wide variety of  environments
and are particularly abundant in soils. Here, we present the complete genome sequence and
methylation pattern of the newly described Gemmatirosa kalamazoonensis type strain.

<>

<1>Decatur, W.A., Johansen, S., Vogt, V.M.
<2>Expression of the Naegleria intron endonuclease is dependent on a functional group I self-cleaving ribozyme.
<3>RNA
<4>6
<5>616-627
<6>2000
<7>NaSSU1 is a complex nuclear group I intron found in several species of Naegleria, consisting
of a large self-splicing group I ribozyme (NaGIR2), which itself is interrupted by a small,
group I-like ribozyme (NaGIR1) and an open reading frame (ORF) coding for a homing
endonuclease. The GIR1 ribozyme cleaves in vitro transcripts of NaSSU1 at two internal
processing sites about 400 nt downstream of the 5' end of the intron, proximal to the
endonuclease ORF. Here we demonstrate that self-cleavage of the excised intron also occurs in
vivo in Naegleria gruberi, generating an ORF-containing RNA that possesses a short leader with
a sequence element likely to be involved in gene expression. To assess the functional
significance of self-cleavage, we constructed a genetic system in Saccharomyces cerevisiae.
First, a mutant yeast strain was selected with a mutation in all the rRNA genes, rendering the
rDNA resistant to cleavage by the Naegleria endonuclease. Active endonuclease, which is
otherwise lethal, could be expressed readily in these cells. Endonuclease activity also could
be detected in extracts of yeast harboring plasmids in which the endonuclease ORF was embedded
in its native context in the intron. Analysis of the RNA from these yeast cells showed that
the excised intron RNA was processed as in N. gruberi. A mutant intron constructed to prevent
self-cleavage of the RNA failed to express endonuclease activity. These results support the
hypothesis that the NaGIR1-catalyzed self-cleavage of the intron RNA is a key event in
expression of the endonuclease.

<>

<1>Deckert, G., Warren, P.V., Gaasterland, T., Young, W.G., Lenox, A.L., Graham, D.E., Overbeek, R., Snead, M.A., Keller, M., Aujay, M., Huber, R., Feldman, R.A., Short, J.M., Olson, G.J., Swanson, R.V.
<2>The complete genome of the hyperthermophilic bacterium Aquifex aeolicus.
<3>Nature
<4>392
<5>353-358
<6>1998
<7>Aquifex aeolicus was one of the earliest diverging, and is one of the most thermophilic,
bacteria known. It can grow on hydrogen, oxygen, carbon dioxide, and mineral salts. The
complex metabolic machinery needed for A. aeolicus to function as a chemolithoautotroph (an
organism which uses an inorganic carbon source for biosynthesis and an inorganic chemical
energy source) is encoded within a genome that is only one-third the size of the E. coli
genome. Metabolic flexibility seems to be reduced as a result of the limited genome size. The
use of oxygen (albeit at very low concentrations) as an electron acceptor is allowed by the
presence of a complex respiratory apparatus. Although this organism grows at 95 degrees C, the
extreme thermal limit of the Bacteria, only a few specific indications of thermophily are
apparent from the genome. Here we describe the complete genome sequence of 1,551,335 base
pairs of this evolutionarily and physiologically interesting organism.

<>

<1>Dedkov, V.C., Rechkunova, N.I., Degtyarev, S.K., Zhilkin, P.A., Kolykhalov, A.A., Verhosina, V.A.
<2>Bacillus sphaericus producing restriction endonuclease BsiI - enzyme purification and characterization.
<3>Soviet Patent Office
<4>SU 1784642
<5>
<6>1992
<7>
<>

<1>Dedkov, V.S.
<2>Analysis of specificity of DNA-methyltransferase M.AspS9I in cell lysate by means of restriction endonuclease blocking.
<3>Biotekhnologiya
<4>0
<5>30-39
<6>2009
<7>Specificity of DNA-methyltransferase M.AspS9I has been determined using cell lysate of
Arthrobacter species S9.  To this end, we employed methylation sensitivity of restriction
endonucleases and also modeling of the methylation process.  Modeling consisted of editing DNa
sequences by substitution of letters for methylated bases and their complementary bases.
Substrate DNA treated by M.AspS9I were used for studying of sensitivity of some restriction
endonucleases to methylation.  Thus it was shown that the overlapping dcm-methylation did not
block the M.AspS9I activity.  The suggested approach can appear universal and simple enough
for determining DNA-methyltransferases specificity.

<>

<1>Dedkov, V.S.
<2>Defining specificity of DNA methyltransferase M.Bsc4I in cellular lysate by blocking restriction endonucleases and computer modeling.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>3-8
<6>2009
<7>The specificity of DNA methyltransferase M.Bsc4I was determined in cellular lysate of Bacillus
schlegelii 4.  The methylation sensitivity of restriction endonucleases and methylation
modeling were used for this purpose.  Modeling consisted of editing DNA sequences using
replacements of methylated bases and their complementary bases.  Substrate DNA treated with
M.Bsc4I were used to study the sensitivity of some restrictases to methylation.  It was shown
that M.Bsc4I methylated 5'-Cm4CNNNNNNNGG-3' and overlapping dcm-methylation blocked its
activity.  The suggested approach would be universal and simple for determining the
specificity of DNA methyltransferases.

<>

<1>Dedkov, V.S.
<2>New DNA methyltransferase M.AjnI from the bacterium Acinetobacter johnsonii R2 produces the 5'-m5CCWGG-3' sequence.
<3>Prikl. Biokhim. Mikrobiol.
<4>46
<5>849-853
<6>2010
<7>Optimum conditions for the activity of the new DNA methylase in cell lysate were determined.
Methylation of DNAs of bacteriophages lambda
and T7 and plasmid pBR322 (dcm+) in the 5'-Cm5CWGG-3' region blocked
M.AjnI activity. The specificity of M.AjnI was determined using lambda
DNA methylated by this enzyme as well as computer modeling and data on
the sensitivity of restriction endonucleases Mval, HinfI, and BstMAI to
methylation.

<>

<1>Dedkov, V.S.
<2>New DNA methyltransferase M.AjnI from Acinetobacter johnsonii R2 produces 5'-m5CCWGG-3' sequence.
<3>Biotekhnologiya
<4>0
<5>36-40
<6>2010
<7>
<>

<1>Dedkov, V.S.
<2>Novel M.BstC8I methyltransferase forms 5'-G(m5C)NNGC-3'. Investigation of restriction endonuclease sensitivity to M.BstC8I methylation.
<3>Mol. Genet. Microbiol. Virol.
<4>27
<5>40-47
<6>2012
<7>A novel M.BstC8I DNA methylase was detected in cell lysate of Bacillus stearothermophilus C8
grown on Luria agar at 37A degrees C. DNA
methylation of bacteriophages lambda and T7 in the 5'-G(m5C)NNGC-3'
segment blocked the activity of the BstC8I restrictase. The specificity
of the M.BstC8I was analyzed on methylated lambda DNA and using
computer modeling and the data on the sensitivity of BstC8I, BsuRI,
AjnI, and PvuII restrictases to methylation. The sensitivity of a
number of restrictases to the novel type of methylation was shown. The
results can be used for study of DNA methylation.

<>

<1>Dedkov, V.S.
<2>Determining of G+C Content in Bacterial DNA using Restriction Endonucleases.
<3>Biotekhnologiya
<4>4
<5>77-82
<6>2004
<7>Bacterial DNAs were cleaved by restriction endonucleases (restrictases) recognizing sequences
of G and C or A and T nucleotides. Experimental curves were obtained for determination G+C
content in bacterial DNA. The developed express-method is supposed to be helpful for
restriction cleavage of bacterial lysates DNA, bacteria identification separation of microbial
isolates in to strains and also for determining DNA methylases.

<>

<1>Dedkov, V.S., Bondar, T.S., Shevchenco, A.V., Degtyarev, S.K.
<2>New rare-cutting restriction endonuclease SmiI from Streptococcus milleri recognizes 5'-ATTT/AAAT-3'.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>1
<5>23-27
<6>2000
<7>A new restriction endonuclease (restrictase) SmiI of type II was detected in the bacterial
strain Streptococcus milleri. Cellular lysate enzyme cut T7 and adenovirus-2 DNAs at site
5'-ATTT/AAAT-3' but not lambda DNA, which does not contain this sequence. Intensive aeration
inhibited the growth of S. milleri. The content of restrictase in cells was the greatest
during the logarithmic growth phase. A total of 20,000 units of SmiI were isolated from 4 g of
cells by cellular extract fractionation with ammonium sulfate and subsequent chromatography on
columns with Bio Gel A 0.5 m, heparin agarose, and phosphocellulose. The purified enzyme cut
the synthetic oligonucleotide duplex in the center of the recognized site 5'-ATTT/AAAT-3'.
SmiI restrictase is a true isoschizomer of the rare-cutting SwaI enzyme. SmiI belongs to a
small group of enzymes which recognize octanucleotide sites and can be used for large-block
fragmentation of DNA. Comparison of specificities of rare-cutting and other restrictases
suggests that the enzymes recognizing octanucleotides can evolutionarily originate from
enzymes recognizing both hexanucleotides and tetranucleotides.

<>

<1>Dedkov, V.S., Degtyarev, S.K.
<2>Detection of restriction endonucleases in Streptomyces and Nocardia cells.
<3>Prikl. Biokhim. Mikrobiol.
<4>28
<5>309-313
<6>1992
<7>A simple technique is proposed for the detection of restriction endonucleases in Streptomyces
and Nocardia cells. The analysis was performed directly in the cells collected from colonies
cultivated on Petri dishes with an innoculation loop. The cells were treated with lyzozyme,
EDTA and Triton X-100. The lysates were tested for restriction endonucleases. The technique
enables the detection of enzymes NcoI, NotI, NruI, Sfr303I, and SfiI in the lysates of the
respective strains-producers.

<>

<1>Dedkov, V.S., Degtyarev, S.K.
<2>Actinobacillus and Streptococcus: producers of isoschizomers of the restriction endonucleases R.HphI, R.SauI, R.NheI, R.MboI and R.SwaI.
<3>Biol. Chem.
<4>379
<5>573-574
<6>1998
<7>New restriction endonucleases have been found in microorganisms isolated from the microflora
of human teeth.  The strain-producers are Actinobacillus suis and Streptococcus milleri.  The
new enzymes are isoschizomers of the prototypes as follows: AsuHPI-HphI; AsuSAI-SauI;
AsuNHI-NheI; AsuMBI and SmiMBI-MboI; SmiI-rare-cutter SwaI.

<>

<1>Dedkov, V.S., Degtyarev, S.K., Rechkunova, N.I., Serov, G.D., Puchkova, L.I., Vaitkevicius, D.P., Kiuduliene, L.J., Janulaitis, A.
<2>Staphylococcus saprophyticus strain b-4069 - is used as a producer of a site-specific restriction endonuclease recognising and splitting the nucleotide sequence GTTAAC.
<3>Soviet Patent Office
<4>SU 1486512 A
<5>
<6>1989
<7>Strain B-4069 of Staphylococcus saprophyticus is used as producer of the site-specific
restriction endonuclease SsrI which recognizes and splits the nucleotide sequence GTTAAC. The
strain is not toxic to humans and is obtained by sedimentation of the microorganisms present
in air and deep cultivation in a medium containing fermentative peptone, yeast extract,
glucose, mineral salts and water, yielding 3.4-3.7 g. of moist biomass/l. of medium. The
preparation (1.5 ml) has an activity of 10000 units/ml.

<>

<1>Dedkov, V.S., Kileva, E.V., Popichenko, D.V., Degtyarev, S.K.
<2>FatI restriction endonuclease from Flavobacterium aquatile NL3 cleaves DNA at 5'-^CATG-3' site.
<3>Biotekhnologiya
<4>5
<5>3-7
<6>2002
<7>
<>

<1>Dedkov, V.S., Mikhnenkova, N.A., Djanobilova, Z.K., Tarasova, M.V.
<2>A M.BssECI DNA Methyltransferase Forms 5 '-m4CCNNGG-3 '. Sensitivity of Restriction Endonucleases to the New Methylation.
<3>Biotekhnologiya
<4>2
<5>32-42
<6>2012
<7>A novel DNA methyltransferase, M.BssECI, has been isolated and purified from Bacillus
stearothermophilus EC cells. The enzyme methylates a
surface cytosine residue in a 5'-CCNNGG-3' sequence with the formation
of N4-methylcytosine, 5'-m4CCNNGG-3'. The approaches to enzyme
isolation, purification (gel-filtration on a Biogel A-0,5m with the
following chromatography on benzyl-DEAE-cellulose and
heparin-Sepharose) and identification of the methylated DNA sequence
were developed. The optimum conditions and the activity of the novel
methyltransferase were determined using phage X. DNA on the basis of
the BssECI restriction endonuclease blockage. A base that was
methylated within the recognized sequence was detected using the
[H-3]-labeling of the oligonucleotide duplex. The specificity of
M.BssECI was investigated using self-methylated Adeno-2 DNA taking into
account the sensitivity of the BssECI, MvaI and MspI restriction
endonucleases to methylated DNA; computer modeling was also employed.
The isolated enzyme can be used in studying of character of DNA
methylation; in particular, it permitted to distinguish the
m4C-methylated cytosine from m5C. DNAs of phages lambda and T7
methylated by M.BssECI were applied to the investigation of the
sensitivity of some restriction endonucleases to the methylation of
their recognition sites.

<>

<1>Dedkov, V.S., Mikhnenkova, N.A., Vlasenko, L.P., Tomilova, J.E., Kashirina, J.G., Gonchar, D.A., Degtyarev, S.K.
<2>Restriction Endonuclease AjnI from Acinetobacter johnsonii R2, an Isoschizomer of EcoRII, recognizes 5'- CCWGG-3' and cleaves dcm-methylated sites.
<3>Biotekhnologiya
<4>3
<5>19-24
<6>2004
<7>A producer of restriction endonuclease AjnI has been isolated from natural resources and
identified as bacterial species Acinetobacter johnsonii R2. The restriction enzyme
purification and estimation of its activity is described. It has been shown that AjnI produces
DNA fragments with 5'-CCWGG sticky ends similar to EcoRII, but cleavage is not blocked by
dcm-methylation. A new restriction endonuclease AjnI may be widely used in genetic
engineering.

<>

<1>Dedkov, V.S., Nayakshina, T.N., Popichenko, D.V., Degtyarev, S.K.
<2>Restriction endonuclease BmtI from Bacillus megaterium S2 cleaves DNA at 5'-GCTAG^C-3' site.
<3>Biotekhnologiya
<4>1
<5>11-15
<6>2003
<7>A new strain producer of a BmtI restriction endonuclease (restrictase) has been found out and
identified as bacteria species Bacillus
megaterium S2. The scheme of purification and several characteristics
of the enzyme are described. BmtI is a heteroschizomer of the well
known NheI enzyme (recognition sequence 5'-GdwnarwCTAGC-3'). It
produces DNA fragments with CTAG-3' extending ends. BmtI restrictase
may be used in genetic engineering.

<>

<1>Dedkov, V.S., Prihodko, G.G., Puchkova, L.I., Serov, G.D., Rechkunova, N.I., Degtyarev, S.K.
<2>SsrI - a Type II restriction endonuclease from Staphylococcus saprophyticus cells.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>11
<5>24-27
<6>1989
<7>The recognition sequence and cleavage site for restriction endonuclease SsrI have been
determined, the latter being 5'-GTT^AAC-3'.  The enzyme was isolated from Staphyloccus
saprophyticus and may be used in DNA investigation instead of its isoschizomer HpaI.

<>

<1>Dedkov, V.S., Rechkunova, N.I., Prihodko, E.A., Kileva, E.V., Kusner, Y.S., Verchozina, V.A., Degtyarev, S.K.
<2>CciNI, an isoschizomer of NotI from Curtobacterium citreum recognizes 5'-GC/GGCCGC-3'.
<3>Gene
<4>157
<5>99-100
<6>1995
<7>CciNI, an isoschizomer of NotI, has been isolated from Curtobacterium citreum.  The enzyme
cleaves within the recognition sequence 5'-GC/GGCCGC-3' as indicated by the slash.

<>

<1>Dedkov, V.S., Repin, V.E., Rechkunova, N.I., Degtyarev, S.K., Verhosina, V.A., Vinogradova, T.P.
<2>Screening of strains producing restriction endonucleases in Lake Baikal.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>1
<5>35-37
<6>1990
<7>Screening of bacterial strains which produce restriction endonucleases was
performed.  It was shown that strains Flavobacterium aquatile, Hafnia alvei,
Acinetobacter calcoaceticus, Pseudomonas gladioli were producers of
restrictases FauI, HalI and HalII, Aca I, PgaI, respectively.

<>

<1>Dedkov, V.S., Sinichkina, S.A., Abdurashitov, M.A., Popichenko, D.V., Degtyarev, S.K.
<2>Restriction endonucleases FalI and FalII from Flavobacterium aquatile Ob10 recognize 5'-(8/13)AAGN5CTT(13/8)-3' and 5'-CG^CG-3',  respectively.
<3>Biotekhnologiya
<4>6
<5>24-29
<6>2003
<7>We have isolated from water supplies a strain Flavobacterium aquatile Ob10 which produces two
restriction endonucleases FalI and FalII.  FalI is a new prototype and recognizes the DNA
sequence that follows: 5'-(8/13)AAGN5CTT(13/8)-3'.  FalI is stimulated by
S-adenosylmethionine and cleaves DNA on both sides of its recognition sequence.  Thus, FalI
belongs to the BcgI-subtype of restriction endonucleases.  The recognition sequence for FalII
restriction
endonuclease is 5'-CG/CG-3' and this enzyme is thus an isoschizomer of FnuDII.

<>

<1>Dedkov, V.S., Sinichkina, S.A., Popichenko, D.V., Degtyarev, S.K.
<2>Restriction endonuclease ZraI from Zoogloea ramigera 11 recognizes 5'-GAC/GTC-3'.
<3>Biotekhnologiya
<4>6
<5>3-7
<6>2001
<7>A producer of restriction endonuclease (restrictase) from natural isolates has been obtained
and identified as bacteria species Zoogloea ramigera II, while the restrictase was named ZraI.
The way of the purification and estimation of the enzyme activity is described.  It was shown
that ZraI produces blunt ended DNA fragments.  The restrictase is promising for the wide use
in gene engineering.

<>

<1>Dedkov, V.S., Zernov, Y.P., Rechkunova, N.I., Degtyarev, S.K.
<2>Isolation of aquatic microorganisms producing the restriction endonucleases from the Black Sea.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>17-18
<6>1990
<7>300 clones of microorganisms isolated at different stations and from different
depths in the Black Sea were screened for restriction endonucleases was found
in 17 clones screened.  Three of them were identified to be Alteromonas
haloplanktis Bl. Restriction endonuclease AhaBI is an isoschizomer of Sau96I.
An identified Alteromonas haloplanktis clone B8 produces AhaB8I restriction
endonuclease the prototype of which is KpnI.  Of the clones isolated three are
Moraxella species B4 producing MspB4I restriction endonuclease analogous to
BanI, three are Bacillus species producing BspB2I and one is Micrococcus lylae
113 producing Mly1131 analogue of NarI, six Moraxella species B6 produce
MspB6I.  The isolated producer strains may be used for isolation of
above-mentioned restriction endonucleases.

<>

<1>DeFilippes, F.M.
<2>A simple assay for DNA restriction endonucleases.
<3>Anal. Biochem.
<4>52
<5>637-641
<6>1973
<7>Recently several groups have used restriction enzymes to produce a limited
number of unique, double stranded DNA fragments from the genome of SV40 virus
and the double stranded replicative form of bacteriophage PhiX174.

<>

<1>DeFilippes, F.M.
<2>A new method for isolation of a restriction enzyme from Haemophilus parainfluenzae.
<3>Biochem. Biophys. Res. Commun.
<4>58
<5>586-596
<6>1974
<7>A rapid procedure which gives high yields of the restriction enzyme HpaI from
Hemophilus parainfluenzae is described.  The procedure effectively removes a
second restriction enzyme HpaII as well as exonucleolytic activity.  The
optimal ionic conditions for the enzyme are similar to those found for one of
the enzymes isolated from Hemophilus influenzae.  The enzyme is stable at 37C
for several hours but it is rapidly inactivated at 60C.  Patterns are presented
which show the electropohoretic separation of the digestion products of two
viral DNAs by this enzyme.

<>

<1>Defossez, P.-A.
<2>Ceci n'est pas une DNMT: Recently discovered functions of DNMT2 and their relation to methyltransferase activity (Comment on DOI 10.1002/bies.201300088).
<3>Bioessays
<4>35
<5>1024
<6>2013
<7>
<>

<1>Degnan, P.H., Yu, Y., Sisneros, N., Wing, R.A., Moran, N.A.
<2>Hamiltonella defensa, genome evolution of protective bacterial endosymbiont from pathogenic ancestors.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>9063-9068
<6>2009
<7>Eukaryotes engage in a multitude of beneficial and deleterious interactions with bacteria.
Hamiltonella defensa, an endosymbiont of
aphids and other sap-feeding insects, protects its aphid host from attack
by parasitoid wasps. Thus H. defensa is only conditionally beneficial to
hosts, unlike ancient nutritional symbionts, such as Buchnera, that are
obligate. Similar to pathogenic bacteria, H. defensa is able to invade
naive hosts and circumvent host immune responses. We have sequenced the
genome of H. defensa to identify possible mechanisms that underlie its
persistence in healthy aphids and protection from parasitoids. The 2.1-Mb
genome has undergone significant reduction in size relative to its closest
free-living relatives, which include Yersinia and Serratia species
(4.6-5.4 Mb). Auxotrophic for 8 of the 10 essential amino acids, H.
defensa is reliant upon the essential amino acids produced by Buchnera.
Despite these losses, the H. defensa genome retains more genes and
pathways for a variety of cell structures and processes than do obligate
symbionts, such as Buchnera. Furthermore, putative pathogenicity loci,
encoding type-3 secretion systems, and toxin homologs, which are absent in
obligate symbionts, are abundant in the H. defensa genome, as are
regulatory genes that likely control the timing of their expression. The
genome is also littered with mobile DNA, including phage-derived genes,
plasmids, and insertion-sequence elements, highlighting its dynamic nature
and the continued role horizontal gene transfer plays in shaping it.

<>

<1>deGraaff, J., Kreuning, P.C., van de Putte, P.
<2>Host controlled restriction and modification of bacteriophage mu and mu-promoted chromosome mobilization in Citrobacter freundii.
<3>Mol. Gen. Genet.
<4>123
<5>283-288
<6>1973
<7>Bacteriophage Mu grown on Escherichia coli K12 (Mu.K) is restricted by wild
type Citrobacter freundii.  In two C. freundii mutants, where the restriction
of foreign F' factors is absent (de Graaff and Stouthamer, 1971), the
restriction for Mu.K., although at a lower level, still exists.  Consequently
two host specificity systems exist in C. freundii, one affecting mainly the
acceptance of foreign plasmid and chromosomal DNA and one affecting foreign DNA
of bacteriophage Mu.  Mu is able to lysogenize C. freundii and to induce
mutations at random in its chromosome.  Furthermore Mu is able to promote the
mobilization of the C. freundii chromosome in strains carrying F' factors.  Mu
promoted integration of F ts 114 lac+ into the C. freundii chromosome was
observed, resulting in the formation of stable Hfr strains.  In this way it is
possible to devise a method for chromosome transfer in other genera than E.
coli to which plasmids of E. coli can be transferred, but in which no
chromosome mobilization is possible because of poor DNA homology between the
foreign plasmid and the host chromosome.

<>

<1>Degtyarev, S.K., Abdurashitov, M.A., Kolyhalov, A.A., Rechkunova, N.I.
<2>AclI, a new restriction endonuclease from Acinetobacter calcoaceticus recognizing 5'-AA^CGTT-3' .
<3>Nucleic Acids Res.
<4>20
<5>3787
<6>1992
<7>AclI, a new restriction endonuclease, has been purified from Acinetobacter calcoaceticus M4.
AclI recognizes the sequence 5'AA^CGTT3' and cleaves DNA as indicated. The enzyme was
purified using the following chromatographic steps: 1) gel-filtration through biogel A-0.5m,
2) DEAE-cellulose, 3) phosphocellulose, 4) heparin sepharose.

<>

<1>Degtyarev, S.K., Belavin, P.A., Repin, V.E., Malygin, E.G.
<2>Method of producing restriction endonuclease FokI from Flavobacterium okeanokoites.
<3>Soviet Patent Office
<4>SU 1406160
<5>
<6>1988
<7>
<>

<1>Degtyarev, S.K., Belavin, P.A., Shishkina, I.G., Zarytova, V.F., Gavryuchenkova, L.P., Morozov, S.M.
<2>Immobilized oligonucleotides as affinity ligands for restriction endonucleases.
<3>Bioorg. Khim.
<4>15
<5>358-362
<6>1989
<7>Affinity chromatography of Type IIS restriction endonucleases is proposed. It is shown that
endonucleases HgaI, FokI, and SfaNI have affinity to the matrix with immobilized
oligonucleotides which contain the endonuclease's recognition sites resistant to hydrolysis.

<>

<1>Degtyarev, S.K., Belichenko, O.A., Lebedeva, N.A., Dedkov, V.S., Abdurashitov, M.A.
<2>BtrI, a novel restriction endonuclease, recognises the non-palindromic sequence 5'-CACGTC(-3/-3)-3'.
<3>Nucleic Acids Res.
<4>28
<5>e56
<6>2000
<7>The recognition sequence and cleavage positions of a new restriction endonuclease BtrI
isolated from Bacillus stearothermophilus SE-U62 have been determined.  BtrI belongs to a rare
type IIQ of restriction endonucleases, which recognise non-palindromic nucleotide sequences
and cleave DNA symmetrically within them.

<>

<1>Degtyarev, S.K., Chmuzh, E.V., Abdurashitov, M.A., Kashirina, Y.G., Dedkov, V.S., Tomilova, Y.E., Mezentseva, N.V., Gonchar, D.A.
<2>Strain of bacterium Bacillus subtilis as producer of restriction endonuclease BisI.
<3>Russian Patent Office
<4>RU 2270859 C
<5>
<6>2006
<7>Invention relates to preparing new strain used for the isolation of the new restriction
endonuclease BisI that can be used in the detection of modified DNA.  The strain Bacillus
subtilis 230 was isolated from soil as a result of a search for producers of restriction
endonucleases. BisI recognizes and cleaves both chains of a DNA nucleotide sequence containing
at least one C5-methylcytosine base in the recognition site 5'-GCNGC-3'

<>

<1>Degtyarev, S.K., Kolyhalov, A.A., Rechkunova, N.I., Abdurashitov, M.A.
<2>AcsI, a new restriction endonuclease from Arthrobacter citreus 310 recognizing 5'-Pu^AATTPy-3'.
<3>Nucleic Acids Res.
<4>20
<5>3789
<6>1992
<7>AcsI, an isoschizomer of FsiI, has been purified from Arthrobacter citreus 310. AcsI
recognizes the sequence 5'Pu^AATTPy3' and cleaves DNA as indicated by the arrow. The enzyme
was purified using two chromatographic steps: phosphocellulose and heparin sepharose. The
enzyme was free of contaminating nuclease activity. After 20-fold over-digestion on lambda DNA
greater than 95% of the DNA fragments can be ligated and then recut by AcsI. Optimal
conditions for AcsI activity are 10 mM Tris-HCl (pH 8.5), 10 mM MgCl2, 100 mM NaCl at 37C. The
fragments produced by AcsI digestion of lambda and T7 DNAs match those predicted by cleavage
at the sequence PuAATTPy.

<>

<1>Degtyarev, S.K., Kolyhalov, A.A., Rechkunova, N.I., Dedkov, V.S.
<2>Determination of substrate specificity of restriction endonuclease FauI.
<3>Bioorg. Khim.
<4>15
<5>130-132
<6>1989
<7>The recognition sequence and cleavage point of restriction endonuclease FauI
have been determined as 5'-CCCGC(4/6).  Not being an isoschizomer of any known
restriction endonuclease, this enzyme may be used in genetic engineering.

<>

<1>Degtyarev, S.K., Kolyhalov, A.A., Rechkunova, N.T., Tepavicharova, I.I., Mechandjiska, L.I., Builieva, E.I.
<2>Bsp1720I, an isoschizomer of EspI from Bacillus species 1720 recognizing 5'-GC^TNAGC-3'.
<3>Nucleic Acids Res.
<4>19
<5>2504
<6>1991
<7>Bsp1720I, an isoschisomer of EspI (1) has been purified from Bacillus species 1720.  Bsp1720I
recognizes the sequence 5'GC^TNAGC3' and cleaves DNA as indicated by the arrow.  The enzyme
was purified using the following chromatographic steps: 1) gel-filtration through biogel A0.5
m, 2) DEAE-cellulose, 3) phosphocellulose.  The enzyme was free of contaminating nuclease
activity.  After 40-fold overdigestion on lambda DNA greater than 95% of the DNA fragments can
be ligated and then recut by Bsp1720I.  Optimal conditions for Bsp1720I activity are 20 mM
Tris-HCl (pH 7.5), 10 mM MgCl2, 100 mM NaCl at 37C. The fragments produced by Bsp1720I
digestion of lambda DNA match those predicted by cleavage at the sequence GCTNAGC (figure 1,
lane 2). The cleavage site was determined accordingly to earlier published procedure (2).  DNA
of pUC 8 with the insert containing a Bsp1720I cleavage site (pVE27) was digested by the
enzymes XmaI and PvuII; reaction products were labelled with [a-32P]dCTP by Klenow Fragment.
350 bp DNA fragment was eluted from gel after electrophoresis in 6% PAAG, its structure and
Bsp1720I hydrolysis site were determined (figure 2).  The results show that Bsp1720I cleaves
DNA as indicated by arrows:
5'-GC^TNAGC-3'
3'-CGANT^CG-5'.

<>

<1>Degtyarev, S.K., Kolykhalov, A.A., Rechkunova, N.I., Dedkov, V.S., Zhilkin, P.A.
<2>BsiI - A new unusual restriction endonuclease.
<3>Mol. Biol. (Mosk)
<4>24
<5>244-247
<6>1990
<7>The restriction endonuclease BsiI from Bacillus sphaericus was isolated.  The
recognition sequence and cleavage point of enzyme BsiI have been determined as
C^TCGTG
GAGCA^C.
This restriction endonuclease is not an isoschizomer of any known restriction
endonucleases and differs from other enzymes: it hydrolyses DNA at an
assymmetrical recognition sequence.

<>

<1>Degtyarev, S.K., Netesova, N.A., Abdurashitov, M.A., Shevchenko, A.V.
<2>Primary structure and strand specificity of BstF5I-1 DNA methyltransferase which recognizes 5'-GGATG-3'.
<3>Gene
<4>187
<5>217-219
<6>1997
<7>The gene for BstF5I-1 DNA-methyltransferase (Mtase) from Bacillus stearothermophilus F5 (a
FokI isoschizomer, recognizing 5'-GGATG-3') was cloned and its nucleotide sequence was
determined.  Analysis of deduced amino acid sequence shows that M.BstF5I-1 belongs to D/21
class of Mtases and has a little homology with M.FokI.  M.BstF5I-1 modifies only the upper
strand of the recognition sequence (5'-GGATG-3').

<>

<1>Degtyarev, S.K., Netesova, N.A., Chizhikov, V.E., Abdurashitov, M.A.
<2>Cloning and characterization of the gene encoding M.FauI DNA methyltransferase.
<3>Biol. Chem.
<4>379
<5>567-568
<6>1998
<7>The enzymes of the FauI restriction-modification system from the Flavobacterium aquatile
strain recognize the non-palindromic sequence 5'-CCCGC-3'/3'-GG-GCG-5'.  We have cloned
the gene encoding the DNA modifying component of this system and determined its nucleotide
sequence.  The deduced amino acid sequence contains ten conserved motifs characteristic for
[cytosine-5] DNA methyltransferases.  Part of the gene sequence that encodes the putative
target recognizing domain of the M.FauI shows some homology with the downstream region, thus
indicating that duplication of the DNA segment was probably involved in the gene evolution.

<>

<1>Degtyarev, S.K., Prihodko, G.G., Rechkunova, N.I.
<2>Determination of the substrate specificity of restriction endonuclease SfeI.
<3>Bioorg. Khim.
<4>14
<5>848-849
<6>1988
<7>The recognition sequence and cleavage site C^TRYAG of a new restriction
endonuclease SfeI have been determined.

<>

<1>Degtyarev, S.K., Prikhodko, E.A., Prikhodko, G.G., Krasnykh, V.N.
<2>VspI methylase belongs to m6A-gamma class of adenine methylases.
<3>Nucleic Acids Res.
<4>21
<5>2015
<6>1993
<7>We have successfully isolated a genomic clone of Vibrio species strain 343 which encodes
methyltransferases and prevents plasmid and bacterial DNA degredation by restriction
endonuclease VspI. There are three open reading frames starting from nucleotides #503, #57,
#812 and ending at #1780, which encode 47.5, 45.5 and 36.1 kd proteins respectively. The
Shine-Dalgarno signal are present in the last two open reading frames. Gel-filtration of
native methylase VspI showed a molecular weight of 42.7 kd. Thus, the open reading frame of
the methylase gene is #557-1780 and it encodes a 408 amino acid protein. The comparison of
deduced amino acid structure of VspI methylase and others has been done according to
Klimasauskas et al. According to mutual positions of two conservative domains this enzyme
belongs to m6A-gamma class. M.VspI has a NPPW-motif instead of an NPPY one, but replacement of
aromatic amino acid tyrosine by another aromatic amino acid tryptophan might be insignificant.

<>

<1>Degtyarev, S.K., Prikhodko, E.A., Rechkunova, N.I., Gorbunov, Y.A.
<2>Interaction of VspI and Tru9I restriction endonucleases with synthetic oligonucleotides.
<3>Biochim. Biophys. Acta
<4>1172
<5>89-94
<6>1993
<7>We describe the properties of two new restriction endonucleases VspI and Tru9I which recognize
sequences AT^TAAT and T^TAA respectively. The molecular weights, subunit structure and
steady-state kinetic constants of these enzymes for native and modified substrates have been
determined. We have investigated the interaction of VspI and Tru9I with synthetic
oligonucleotides containing modifications either within the recognition sites or around them.
These modifications represent the substitution of different DNA deoxyribonucleosides by
1,2-dideoxy-D-ribofuranose, which corresponds to loss of the heterocyclic base while the
sugar-phosphate chain remains intact. The effects of the substitutions were analyzed by
determining the steady-state kinetic values of the hydrolysis reaction by VspI and Tru9I. The
enzymes exhibited Michaelis-Menten kinetics for hydrolyzable substrates. The initial rates
(Vo) of hydrolysis of modified and unmodified strands of the duplexes varied as a result of
these substitutions. The substrates for VspI and Tru9I which contain modifications around the
bond to be hydrolyzed or within the complementary nucleosides were unreactive.

<>

<1>Degtyarev, S.K., Prikhodko, E.A., Rechkunova, N.I., Prikhodko, G.G., Krasnykh, V.N.
<2>Biochemical characterization of VspI methyltransferase.
<3>Gene
<4>157
<5>65-66
<6>1995
<7>The gene (vspIM) encoding VspI methyltransferase (MTase) has previously been cloned and
sequenced, and shown to belong to the gamma class of m6-adenine MTases.  Here it is shown that
the MTase modifies the third adenine within the recognition sequence 5'-ATTAAT-3'.

<>

<1>Degtyarev, S.K., Rechkunova, N.I.
<2>Determination of purity of restriction endonucleases.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>14
<5>102-105
<6>1988
<7>A method for the determination of exonucleases and phosphatase impurities in preparations of
restriction endonucleases using 5'-[32P]-labelled double-stranded 20-base long
deoxyribooligonucleotides is suggested. Incubation of enzyme with substrate is carried out in
restriction buffer with subsequent electrophoresis of the reaction mixture in 20%
polyacrylamide in the presence of 7M urea and autoradiography of the gel.  The percent of
label in oligonucleotide is determined.

<>

<1>Degtyarev, S.K., Rechkunova, N.I., Grinev, A.A., Dedkov, V.S.
<2>Determination of substrate specificity of restriction endonucleases Bme18I and Kzo9I.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>15
<5>25-26
<6>1989
<7>The recognition sequence and cleavage point of restriction endonucleases Bme18I and Kzo9I have
been determined as G^G(A/T) CC and ^GATC, respectively.

<>

<1>Degtyarev, S.K., Rechkunova, N.I., Kolyhalov, A.A., Dedkov, V.S., Zhilkin, P.A.
<2>II-Q restriction endonucleases - new class of type II enzymes.
<3>Nucleic Acids Res.
<4>18
<5>5807-5810
<6>1990
<7>Unique restriction endonucleases Bpu10I and BsiI have been isolated from
Bacillus pumilus and Bacillus sphaericus, respectively.  The recognition
sequences and cleavage points of these enzymes have been determined as
5'-CC^TNAGC-3'
3'-GGANT^CG-5' for Bpu10I
and
5'-C^TCGTG-3'
3'-GAGCA^C-5' for BsiI.
Restriction endonucleases Bpu10I and BsiI represent a new class of enzymes
which recognize nonpalindromic nucleotide sequences and hydrolyze DNA within
the recognition sequences may be regarded as quasi-palindromic and the enzymes
may be designated as type II-Q restriction endonucleases.

<>

<1>Degtyarev, S.K., Rechkunova, N.I., Netesova, N.A., Tchigikov, V.E., Malygin, E.G., Kochkin, A.V., Mikhajlov, V.V., Rasskazov, V.A.
<2>Determination of substrate specificity of restriction endonuclease VneI.
<3>Bioorg. Khim.
<4>13
<5>422-423
<6>1987
<7>The recognition sequence and cleavage point of restriction endonuclease VneI
have been determined as 5'-G^TGCAC.  This enzyme is not isoschizomer of any
known restriction endonucleases and therefore may be widely used in
investigation of DNA structure.

<>

<1>Degtyarev, S.K., Rechkunova, N.I., Novozhilov, M.I., Semenchenko, G.V., Galimdayeva, R.S.
<2>Brevibacterium immotum bacterial strain - produces restriction endonuclease BimI which recognises and cleaves specified nucleotide sequence.
<3>Soviet Patent Office
<4>SU 1532584 A
<5>
<6>1989
<7>The bacterial strain Brevibacterium immotum VKPM V-4151 (I) produces the restriction
endonuclease BimI, which is an isoschizomer of the restriction endonuclease MlaI, and which is
able to recognise and cleave the nucleotide sequence TTCGAA. (I) has been identified as a
result of research into restriction endonuclease-producing strains of microorganisms
inhabiting Lake Balkhash. (I) contains the restriction endonuclease BimII (isoschizomer of
BspRI) as well as BimI, and they are separated from each other by chromatography on
phosphocellulose P-11. (I) grows on in inexpensive, readily available nutrient medium. 2500
units of the restriction endonuclease BimI with activity 10000 units/ml can be separated from
one gram of raw biomass.

<>

<1>Degtyarev, S.K., Rechkunova, N.I., Novozhilova, M.I., Semenchenko, G.V., Galimdayeva, R.S.
<2>Paracoccus denitrificans bacterial strain - produces restriction endonuclease Pde12I which recognises and cleaves the specified nucleotide sequence.
<3>Soviet Patent Office
<4>SU 1532583 A
<5>
<6>1989
<7>The bacterial strain Paracoccus denitrificans VKPM V-4152 (I) produces the restriction
endonuclease Pde12I, which is an isoschizomer of the restriction endonuclease Sau96I, and
which is able to recognise and cleave the nucleotide sequence GGNCC. (I) has been identified
as a result of research into restriction endonuclease-producing strains of microorganisms
inhabiting Lake Balkhash. It is easily destroyed by ultrasound and provides a high yield of
the restriction endonuclease Pde12I. (I) is cultured in a nutrient medium containing (in g/l):
tryptone 10, yeast extract 5, and water the remainder. The process is carried out at 30
degrees C with good aeration for 2 days. Biomass yield is 2-3 g/l of culture medium and Pde12I
yield is 3000 units/g biomass.

<>

<1>Degtyarev, S.K., Rechkunova, N.I., Repin, V.E., Kolyhalov, A.A., Netesov, S.V.
<2>The recognition sequence and cleavage point of restriction endonuclease Bse 21 I.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>1
<5>138-139
<6>1990
<7>The recognition sequence and cleavage point of restriction endonuclease Bse21I
have been determined as 5'-CC^TNAGG.  This enzyme is an isoschizomer of SauI
and may replace it in investigation of DNA structure.

<>

<1>Degtyarev, S.K., Rechkunova, N.I., Zernov, Y.P., Dedkov, V.S., Chizikov, V.E., Van Calligan, M., Williams, R., Murray, E.
<2>Bsp24I, a new unusual restriction endonuclease.
<3>Gene
<4>131
<5>93-95
<6>1993
<7>*
A new restriction endonuclease, Bsp24I, from Bacillus species 24, recognizing:

   5'-^N8 GACNNNNNNTGGN12^-3'
   3'-^N13CTGNNNNNNACCN7^-5',

has been isolated. Its specificity and cleavage points were determined.


<>

<1>Degtyarev, S.K., Rechkunova, N.I., Zhilkina, O.A., Semenchenko, G.V., Novozhilova, M.I.
<2>Kurthia zopfii as a producer of the restriction endonuclease Kzo 9I.
<3>Soviet Patent Office
<4>SU 1440919 A
<5>
<6>1988
<7>The present invention is directed to produce the restriction endonuclease Kzo9I recognizing
the nucleotide sequence GATC from Kurthia zopfii. The yield of the enzyme was 2,500 units/g
cells with specific activity of 10,000 units/ml.

<>

<1>Degtyarev, S.K., Repin, V.E.
<2>Obtaining HgaI-restrictase-restriction endonuclease HgaI preparation by Haemophilus gallinarum and purification.
<3>Soviet Patent Office
<4>SU 1389292 A
<5>
<6>1996
<7>
<>

<1>Degtyarev, S.K., Repin, V.E., Rechkunova, N.I., Semchenko, L.C., Ivanova, E.P., Rasskazov, V.A.
<2>Novel strain of bacteria, Bacillus species 21 - is used as a producer of the site-specific restriction endonuclease Bse21I.
<3>Soviet Patent Office
<4>SU 1518372 A
<5>
<6>1989
<7>Novel Bacillus species 21 strain is a producer of the site-specific restriction endonuclease
Bse21I, which can recognise and split the sequence of nucleotides 5'-CCTNAGG-3'. The strain
is isolated from a sample of Japan sea water, does not require special nutrients, the cells
are easily disrupted with ultrasonication, and gives high yield of ferment equal to 30000
units/g of biomass.

<>

<1>Degtyarev, S.K., Repin, V.E., Rechkunova, N.I., Shevchenko, A.V., Abdurashitov, M.A.
<2>Strain of bacterium Bacillus badius - a producer of restriction endonuclease recognizing and splitting the nucleotide sequence 5'-(A/T)CCGG(A/T)-3'.
<3>Russian Patent Office
<4>RU 2053299 C
<5>
<6>1996
<7>
<>

<1>Degtyarev, S.K., Repin, V.E., Rechkunova, N.I., Tchigikov, V.E., Malygin, E.G., Mikhajlov, V.V., Rasskazov, V.A.
<2>Determination of the substrate specificity of restriction endonuclease VspI.
<3>Bioorg. Khim.
<4>13
<5>420-421
<6>1987
<7>The recognition sequence and cleavage point of restriction endonuclease VspI
have been determined as 5'-AT^TAAT.  This enzyme is not an isoschizomer of any
known restriction endonucleases.  DNA pBR322 contains a single VspI recognition
sequence in position 3539.  Therefore this enzyme may be used for cloning DNA
in the VspI site in AmpR-gene of pBR322.

<>

<1>Degtyarev, S.K., Zernov, Y.P.
<2>Type II restriction enzymes:  Possible evolutionary links between the enzymes and their palindromic tetranucleotide recognition sequences.
<3>Mol. Biol. (Mosk)
<4>24
<5>1393-1398
<6>1990
<7>The distribution of restriction enzymes with tetranucleotide recognition sequences in nature
was analyzed.  A rule for the conversion of these recognition sites was formulated, and a
scheme showing the evolutionary links of restriction enzymes with their palindromic
tetranucleotide sequences is proposed.

<>

<1>Degtyarev, S.K., Zilkin, P.A., Prihodko, G.G., Repin, V.E., Rechkunova, N.I.
<2>Determination of unusual substrate specificity of restriction endonuclease Bpu10I.
<3>Mol. Biol. (Mosk)
<4>23
<5>1051-1056
<6>1989
<7>A new enzyme Bpu10I was isolated form Bacillus pumilus.  This enzyme is not an isoschizomer of
any known restriction endonucleases.  The search of possible recognition sequences was carried
out in sequences ABCNiDEF (i=0-6) on substrate DNA lambda CI857, T7, pBR322.  The recognition
sequence and cleavage sites of restriction endonuclease Bpu10I have been determined as
CC^TNAGC
GGANT^CG.

<>

<1>Deibert, M., Grazulis, S., Janulaitis, A., Siksnys, V., Huber, R.
<2>Crystal structure of MunI restriction endonuclease in complex with cognate DNA at 1.7 A resolution.
<3>EMBO J.
<4>18
<5>5805-5816
<6>1999
<7>The MunI restriction enzyme recognizes the palindromic hexanucleotide sequence C/AATTG (the
'/' indicates the cleavage site).  The crystal structure of its active site mutant D83A
bound to cognate DNA has been determined at 1.7 A resolution.  Base-specific contacts between
MunI and DNA occur exclusively in the major groove. While DNA-binding sites of most other
restriction enzymes are comprised of discontinuous sequence segments, MunI combines all
residues involved in the base-specific contacts within one short stretch (residues R115-R121)
located at the N-terminal region of the 3(10)4 helix. The outer CG base pair of the
recognition sequence is recognized solely by R115 through hydrogen bonds made by backbone and
side chain atoms to both bases. The mechanism of recognition of the central AATT nucleotides
by MunI is similar to that of EcoRI, which recognizes the G/AATTC sequence. The local
conformation of AATT deviates from the typical B-DNA form and is remarkably similar to
EcoRI-DNA. It appears to be essential for specific hydrogen bonding and recognition by MunI
and EcoRI.

<>

<1>Deibert, M., Grazulis, S., Sasnauskas, G., Siksnys, V., Huber, R.
<2>Structure of the tetrameric restriction endonuclease NgoMIV in complex with cleaved DNA.
<3>Nat. Struct. Biol.
<4>7
<5>792-799
<6>2000
<7>The crystal structure of the NgoMIV restriction endonuclease in complex with cleaved DNA has
been determined at 1.6 A resolution. The crystallographic asymmetric unit contains a protein
tetramer and two DNA molecules cleaved at their recognition sites. This is the first structure
of a tetrameric restriction enzyme-DNA complex. In the tetramer, two primary dimers are
arranged back to back with two oligonucleotides bound in clefts on opposite sides of the
tetramer. The DNA molecules retain a B-type conformation and have an enclosed angle between
their helical axes of 60 degrees. Sequence-specific interactions occur in both the major and
minor grooves. Two Mg2+ ions are located close to the cleaved phosphate at the active site of
NgoMIV. Biochemical experiments show that interactions between the recognition sites within
the tetramer greatly increase DNA cleavage efficiency.

<>

<1>Deissler, H., Genc, B., Doerfler, W.
<2>Restriction endonuclease BsoFI is sensitive to the 5'-methylation of deoxycytidines in its recognition sequence.
<3>Nucleic Acids Res.
<4>23
<5>4227-4228
<6>1995
<7>The isoschizomeric restriction endonucleases Fnu4HI and BsoFI cleave DNA at 5'-GC/NGC-3'
sequences.  Fnu4HI has been shown to be inhibited by 5'-CG-3' methylation in the sequences
5'-GmCGGC-3' or 5'-GCGGmCG-3'.  We have now investigated the methylation sensitivity of
BsoFI by testing its activity on plasmid DNA 5'-CG-3' methylated with the M.SssI DNA
methyltransferase or on synthetic (CGG)n repetitive oligodeoxyribonucleotides which have been
partly or completely C methylated.  The data demonstrate that BsoFI cannot cleave at its
recognition sequence when it is completely 5'-CG-3' methylated.  These enzymes have proven
to be useful in analyses of the methylation status in (CGG)n repeats of the human genome.

<>

<1>DeJonckheere, J.F., Brown, S.
<2>Three different group I introns in the nuclear large subunit ribosomal DNA of the amoeboflagellate Naegleria.
<3>Nucleic Acids Res.
<4>26
<5>456-461
<6>1998
<7>We have amplified the large subunit ribosomal DNA of the 12 described Naegleria spp. and of 34
other Naegleria lineages that might be distinct species.  Two strains yielded a product that
is longer than 3 kb, which is the length of the LSUrDNA of all described Naegleria spp.
Sequencing data revealed that the insert in one of these strains is a group I intron without
an open reading frame, while the other strain contains two different group I introns, of which
the second intron has an ORF of 175 amino acids.  In the latter ORF there is a conserved
His-Cys box as in the homing endonucleases present in group I introns in the small subunit
ribosomal DNA of Naegleria spp.  Although the group I introns in the LSUrDNA differ in
sequence, they are more related to each other than they are to the group I introns in the
SSUrDNA of Naegleria spp.  The three group I introns in the LSUrDNA in Naegleria are at
different locations and are probably acquired by horizontal transfer, contrary to the SSUrDNA
group I introns in this genus which are of ancestral origin and are transmitted vertically.

<>

<1>Del Castillo, C.S., Hikima, J.I., Jang, H.B., Nho, S.W., Jung, T.S., Wongtavatchai, J., Kondo, H., Hirono, I., Takeyama, H., Aoki, T.
<2>Comparative sequence analysis of a multi-drug resistant plasmid from Aeromonas hydrophila.
<3>Antimicrob. Agents Chemother.
<4>57
<5>120-129
<6>2013
<7>Aeromonas hydrophila is a pathogenic bacterium that has been implicated in fish,
animal, and human disease. Recently, a multi-drug resistance (MDR) plasmid pR148
was isolated from A. hydrophila obtained from a tilapia (Oreochromis niloticus)
farm in Thailand. pR148 is a 165,906 bp circular plasmid containing 147 coding
regions showing highest similarity to pNDM-1_Dok1, an MDR plasmid isolated from a
human pathogen. It was also very similar to other IncA/C plasmids isolated from
humans, animals, food, and fish. pR148 contains a mercuric resistance operon and
encodes the complete set of genes for the type 4 secretion system. pR148 encodes
for a Tn21 type transposon. This contains the drug resistance genes qacH,
bla(OXA-10), aadA1, and sul1 in a class 1 integron; tetA and tetR in a transposon
Tn1721; and catA2 and a duplicate sul1 in a locus showing 100% similarity to IncU
plasmids isolated from fish. The bla(OXA-10) and aadA1 genes showed 100%
similarity with those from the Acinetobacter baumannii AYE chromosomal genome.
The similarity of pR148 to a human pathogen-derived plasmid indicates that the
plasmids were either transferred between them or that they are derived from a
common origin. Previous studies have shown that IncA/C plasmids retain a
conserved backbone, while the accessory region points to lateral gene transfer.
These observations point out the dangers of indiscriminate use of antibiotics in
humans and in animals and the necessity of understanding how drug resistance
determinants are disseminated and transferred.

<>

<1>Del Cerro, C., Felpeto-Santero, C., Rojas, A., Tortajada, M., Ramon, D., Garcia, J.L.
<2>Genome Sequence of the Butanol Hyperproducer Clostridium saccharoperbutylacetonicum N1-4.
<3>Genome Announcements
<4>1
<5>e0007013
<6>2013
<7>Clostridium saccharoperbutylacetonicum is one of the most important acetone-butanol-ethanol
(ABE)-generating industrial microorganisms and one of the
few bacteria containing choline in its cell wall. Here, we report the draft
genome sequence of C. saccharoperbutylacetonicum strain N1-4 (6.6 Mbp; G+C
content, 29.4%) and the findings obtained from the annotation of the genome.

<>

<1>Del Cerro, C., Garcia, J.M., Rojas, A., Tortajada, M., Ramon, D., Galan, B., Prieto, M.A., Garcia, J.L.
<2>Genome Sequence of the Methanotrophic Poly-beta-Hydroxybutyrate Producer Methylocystis parvus OBBP.
<3>J. Bacteriol.
<4>194
<5>5709-5710
<6>2012
<7>Methylocystis parvus OBBP is an obligate methylotroph considered the type species of the genus
Methylocystis. Two pmoCAB particulate methane monooxygenase operons
and one additional singleton pmoC paralog were identified in the sequence. No
evidence of genes encoding soluble methane monooxygenase was found. Comparison of
M. parvus OBBP and Methylocystis sp. strain Rockwell (ATCC 49242) suggests that
both species should be taxonomically classified in different genera.

<>

<1>del Gaudio, R., Di Giaimo, R., Potenza, N., Branno, M., Aniello, F., Geraci, G.
<2>Characterization of a new variant DNA (cytosine-5)-methyltransferase unable to methylate double stranded DNA isolated from the marine annelid worm Chaetopterus variopedatus.
<3>FEBS Lett.
<4>460
<5>380-384
<6>1999
<7>The enzyme S-adenosylmethionine-DNA (cytosine-5)-methyltransferase has been identified, first
time for invertebrates, in embryos of the marine polychaete annelid worm Chaetopterus
variopedatus. The molecule has been isolated from embryos at 15 h of development. It is a
single peptide of about 200 kDa molecular weight, cross-reacting with antibodies against sea
urchin DNA methyltransferase. The enzymatic properties of the molecule are similar to those of
Dnmt1 methyltransferases isolated from other organisms, but with the peculiarity to be unable
to make 'de novo' methylation on double stranded DNA.

<>

<1>Del Giudice, L.
<2>Method for isolating restriction- and modificationless mutants of Escherichia coli K-12.
<3>J. Bacteriol.
<4>137
<5>673-676
<6>1979
<7>A simple method is described for the selection and isolation of restriction- and
modificationless mutants in Escherichia coli K-12 by using the following properties: (i) the
temperature-sensitive repressor activity of phage lambda-cI857; (ii) a mutant of lambda phage
defective in integration and the establishment of repression (lambda-b2cI); (iii) a virulent
phage insensitive to the repressor activity.  The final yield of spontaneously arising rK- mK+
and rK- mK- mutants from stationary-phase cultures was about 5% of the surviving cells.

<>

<1>Del Rio, T.G. et al.
<2>Complete genome sequence of Intrasporangium calvum type strain (7 KIP).
<3>Standards in Genomic Sciences
<4>3
<5>294-303
<6>2010
<7>Intrasporangium calvum Kalakoutskii et al. 1967 is the type species of the genus
Intrasporangium, which belongs to the actinobacterial family Intrasporangiaceae.
The species is a Gram-positive bacterium that forms a branching mycelium, which
tends to break into irregular fragments. The mycelium of this strain may bear
intercalary vesicles but does not contain spores. The strain described in this
study is an airborne organism that was isolated from a school dining room in
1967. One particularly interesting feature of I. calvum is that the type of its
menaquinone is different from all other representatives of the family
Intrasporangiaceae. This is the first completed genome sequence from a member of
the genus Intrasporangium and also the first sequence from the family
Intrasporangiaceae. The 4,024,382 bp long genome with its 3,653 protein-coding
and 57 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Delahodde, A., Goguel, V., Becam, A.M., Creusot, F., Perea, J., Banroques, J., Jacq, C.
<2>Site-specific DNA endonuclease and RNA maturase activities of two homologous intron-encoded proteins from yeast mitochondria.
<3>Cell
<4>56
<5>431-444
<6>1989
<7>Two introns of the mitochondrial genome 777-3A of S. cerevisiae, bI4 in cob and aI4 in cox1
genes, contain ORFs that can be translated into two homologous proteins. We changed the UGA,
AUA, and CUN codons of these ORFs to the universal genetic code, in order to study the
functions of their translated products in E. coli and in yeast, by retargeting the nuclear
encoded protein into mitochondria. The p27b14 protein has been shown to be required for the
splicing of both introns bI4 and aI4. The homologous p28aI4 protein is highly toxic to E.
coli. It can specifically cleave double-stranded DNA at a sequence representing the junction
of the two fused flanking exons. We present evidence that this system is a good model for
studying the role of mitochondrial intron-encoded proteins in the rearrangement of genetic
information at both the RNA (RNA splicing-bI4 maturase) and DNA levels (intron
transposition-aI4 transposase).

<>

<1>Delannoy, C.M., Zadoks, R.N., Lainson, F.A., Ferguson, H.W., Crumlish, M., Turnbull, J.F., Fontaine, M.C.
<2>Draft Genome Sequence of a Nonhemolytic Fish-Pathogenic Streptococcus agalactiae  Strain.
<3>J. Bacteriol.
<4>194
<5>6341-6342
<6>2012
<7>Streptococcus agalactiae is a significant Gram-positive bacterial pathogen of terrestrial and
aquatic animals. A subpopulation of nonhemolytic strains which
appear to be pathogenic only for poikilotherms exists. We report here the first
draft genome sequence of a nonhemolytic S. agalactiae isolate recovered from a
diseased fish.

<>

<1>Deleo, F.R., Chen, L., Porcella, S.F., Martens, C.A., Kobayashi, S.D., Porter, A.R., Chavda, K.D., Jacobs, M.R., Mathema, B., Olsen, R.J., Bonomo, R.A., Musser, J.M., Kreiswirth, B.N.
<2>Molecular dissection of the evolution of carbapenem-resistant multilocus sequence type 258 Klebsiella pneumoniae.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>111
<5>4988-4993
<6>2014
<7>Infections caused by drug-resistant bacteria are a major problem worldwide.
Carbapenem-resistant Klebsiella pneumoniae, most notably isolates classified as
multilocus sequence type (ST) 258, have emerged as an important cause of hospital
deaths. ST258 isolates are predominantly multidrug resistant, and therefore
infections caused by them are difficult to treat. It is not known why the ST258
lineage is the most prevalent cause of multidrug-resistant K. pneumoniae
infections in the United States and other countries. Here we tested the
hypothesis that carbapenem-resistant ST258 K. pneumoniae is a single genetic
clone that has disseminated worldwide. We sequenced to closure the genomes of two
ST258 clinical isolates and used these genomes as references for comparative
genome sequencing of 83 additional clinical isolates recovered from patients at
diverse geographic locations worldwide. Phylogenetic analysis of the SNPs in the
core genome of these isolates revealed that ST258 K. pneumoniae organisms are two
distinct genetic clades. This unexpected finding disproves the single-clone
hypothesis. Notably, genetic differentiation between the two clades results from
an approximately 215-kb region of divergence that includes genes involved in
capsule polysaccharide biosynthesis. The region of divergence appears to be a
hotspot for DNA recombination events, and we suggest that this region has
contributed to the success of ST258 K. pneumoniae. Our findings will accelerate
research on novel diagnostic, therapeutic, and vaccine strategies designed to
prevent and/or treat infections caused by multidrug resistant K. pneumoniae.

<>

<1>Delhalle, E.
<2>Restriction and modification of bacteriophages by Escherichia coli K12 involving a cryptic P1 prophage associated with different plasmids.
<3>Ann. Microbiol. (Paris)
<4>124A
<5>173-178
<6>1973
<7>By transducing hybrid plasmids with P1, two transductants were found to carry a cryptic P1
phage closely associated with plasmids R(T), Ton or Col(VI), Trp.  The presence of these
cryptic P1 phages was manifested by restriction and modification of phages T1 and T7.  Linkage
on a single genetic structure of the cryptic P1 phage and the other extrachromosomal genes was
demonstrated by associated transfer by conjugation and P1-transduction.  The two cryptic P1
phages were not identical.

<>

<1>Deligios, M., Bacciu, D., Deriu, E., Corti, G., Bordoni, R., De Bellis, G., Leori, G.S., Rubino, S., Uzzau, S.
<2>Draft Genome Sequence of the Host-Restricted Salmonella enterica Serovar Abortusovis Strain SS44.
<3>Genome Announcements
<4>2
<5>e00261-14
<6>2014
<7>Salmonella enterica serovar Abortusovis is a pathogen strictly adapted to ovines, in which it
causes abortion. To enhance our understanding of this pathogen, we assembled the first draft
sequence of an S. Abortusovis genome (strain SS44). The obtained genomic data might facilitate
the study of S. enterica evolution and host adaptation.

<>

<1>DeLong, E.F., Preston, C.M., Mincer, T., Rich, V., Hallam, S.J., Frigaard, N.U., Martinez, A., Sullivan, M.B., Edwards, R., Brito, B.R., Chisholm, S.W., Karl, D.M.
<2>Community genomics among stratified microbial assemblages in the ocean's interior.
<3>Science
<4>311
<5>496-503
<6>2006
<7>Microbial life predominates in the ocean, yet little is known about its genomic variability,
especially along the depth continuum. We report here
genomic analyses of planktonic microbial communities in the North Pacific
Subtropical Gyre, from the ocean's surface to near-sea floor depths.
Sequence variation in microbial community genes reflected vertical
zonation of taxonomic groups, functional gene repertoires, and metabolic
potential. The distributional patterns of microbial genes suggested
depth-variable community trends in carbon and energy metabolism,
attachment and motility, gene mobility, and host-viral interactions.
Comparative genomic analyses of stratified microbial communities have the
potential to provide significant insight into higher-order community
organization and dynamics.

<>

<1>Delorme, C., Bartholini, C., Luraschi, M., Pons, N., Loux, V., Almeida, M., Guedon, E., Gibrat, J.F., Renault, P.
<2>Complete Genome Sequence of the Pigmented Streptococcus thermophilus Strain JIM8232.
<3>J. Bacteriol.
<4>193
<5>5581-5582
<6>2011
<7>Streptococcus thermophilus is a dairy species commonly used in the manufacture of cheese and
yogurt. Here, we report the complete sequence of
S. thermophilus strain JIM8232, isolated from milk and which produces a
yellow pigment, an atypical trait for this bacterium.

<>

<1>Delorme, C., Guedon, E., Pons, N., Cruaud, C., Couloux, A., Loux, V., Chiapello, H., Poyart, C., Gautier, C., Sanchez, N., Almeida, M., Kennedy, S., Ehrlich, S.D., Gibrat, J.F., Wincker, P., Renault, P.
<2>Complete Genome Sequence of the clinical Streptococcus salivarius strain CCHSS3.
<3>J. Bacteriol.
<4>193
<5>5041-5042
<6>2011
<7>Streptococcus salivarius is a commensal species commonly found in the human oral cavity and
digestive tract, although it is also associated with
human infections such as meningitis, endocarditis and bacteremia. Here, we
report the complete sequence of S. salivarius strain CCHSS3 isolated from
human blood.

<>

<1>DelVecchio, V.G. et al.
<2>The genome sequence of the facultative intracellular pathogen Brucella melitensis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>443-448
<6>2002
<7>Brucella melitensis is a facultative intracellular bacterial pathogen that causes abortion in
goats and sheep and Malta fever in humans. The genome of B. melitensis strain 16M was
sequenced and found to contain 3,294,935 bp distributed over two circular chromosomes of
2,117,144 bp and 1,177,787 bp encoding 3,197 ORFs. By using the bioinformatics suite ERGO,
2,487 (78%) ORFs were assigned functions. The origins of replication of the two chromosomes
are similar to those of other -proteobacteria. Housekeeping genes, including those involved in
DNA replication, transcription, translation, core metabolism, and cell wall biosynthesis, are
distributed on both chromosomes. Type I, II, and III secretion systems are absent, but genes
encoding sec-dependent, sec-independent, and flagella-specific type III, type IV, and type V
secretion systems as well as adhesins, invasins, and hemolysins were identified. Several
features of the B. melitensis genome are similar to those of the symbiotic Sinorhizobium
meliloti.

<>

<1>Delver, E.P., Agofanova, O.V., Tupikova, E.E., Vorobeva, E.P., Belogurov, A.A.
<2>System controlling expression of antirestriction genes ardA and ardB of IncN transmission plasmid pKM101 (R46).
<3>Mol. Biol. (Mosk)
<4>32
<5>208-213
<6>1998
<7>A system regulating the expression of antirestriction genes ardA and ardB of transmission
plasmid pKM101 (R46) was studied.  These phylogenetically distant genes are involved in
protecting plasmid DNA from cell restriction enzymes and adapting to a new host after conjugal
transfer.  The regulatory system involves two conserved upstream repeats, which flank the 5'
end of ard and contain a potent promoter, and regulatory proteins ArdK and ArdR.  At least one
of the proteins, ArdK, can bind with the promoter and act as a suppressor.  Both proteins
control the expression of their own genes.  The regulatory system was assumed to trigger the
coordinated expression of the ard genes and efficient synthesis of the Ard antirestriction
proteins, which are essential to conjugal transfer of the plasmid and its adaptation to the
new host.

<>

<1>Delver, E.P., Kotova, V.U., Zavilgelsky, G.B., Belogurov, A.A.
<2>Nucleotide sequence of the Gene (ard) encoding the antirestriction protein of plasmid colIb-P9.
<3>J. Bacteriol.
<4>173
<5>5887-5892
<6>1991
<7>The IncI1 plasmid ColIb-P9 was found to encode an antirestriction function.
The relevant gene, and (alleviation of restriction of DNA), maps about 5 kb
from the origin of transfer, in the region transferred early during bacterial
conjugation.  Ard inhibits both restriction and modification by each of the
four type I systems of Escherichia coli tested, but it had no effect on
restriction by EcoRI, a type II system, or EcoP1, a type III system.  The
nucleotide sequence of the ColIb ard gene was determined; the predicted
molecular weight of the Ard polypeptide is 19,193.  The proposed polypeptide
chain contains an excess of 25 negatively charged amino acids, suggesting that
its overall character is very acidic.  Deletion analysis of the gene revealed
that the Ard protein contained a distinct functional domain located in the
COOH-terminal half of the polypeptide.  We suggest that the biological role of
the ColIb Ard protein is associated with overcoming host-controlled restriction
during bacterial conjugation.

<>

<1>Demidova, G.V., Goncharov, E.K., Tynianova, V.I.
<2>Comparison of specific recognition sites of adenine and cytosine DNA-methyltransferase of Yersinia pestis EV 76C dam and dcm by Escherichia coli methylases.
<3>Biokhimiia
<4>49
<5>1594-1597
<6>1984
<7>Using enzymatic modelling of in vitro methylation of chromosome DNAs from Yersinia pestis EV
76, E. coli 834 and E. coli C600 RII by DNA methylases of EcoRII and EcoDam as well as of DNA
hydrolysis of plasmid pBR 322 from the cells of Y. pestis EV 76, E. coli C600 and E. coli 834
by restrictases of EcoRII and CfuI, it was found that cytosine DNA methylase from plaque
bacteria does not correspond to the type of RII methylases of E. coli.  Adenine DNA methylase
is related to E. coli methylases type dam and modified adenine in the nucleotide sequence of
GATC.

<>

<1>Dempsey, R.M., Carroll, D., Kong, H.M., Higgins, L., Keane, C.T., Coleman, D.C.
<2>Sau421, a Bcgl-like restriction-modification system encoded by the Staphylococcus aureus quadruple-converting phage phi 42.
<3>Microbiology
<4>151
<5>1301-1311
<6>2005
<7>The serotype F phage 042 of Staphylococcus aureus is a triple-converting bacteriophage that
encodes the staphylokinase gene
(sak) and the enterotoxin A gene (entA). Lysogeny results in loss of
expression of the chromosomal beta-haemolysin gene (hlb) (negative
conversion), the expression of staphylokinase and enterotoxin A
(positive conversion), and the acquisition of resistance to lysis by
all 23 phages of the International Basic Set (IBS) of S. aureus typing
phages. Until this study, the basis of 042 resistance to lysis by
exogenous phages was unknown. The authors report here that phage 042
encodes a restriction-modification (R-M) system, termed Sau42I,
adjacent to and in the same orientation to the phage integrase gene
int. The genes encoding Sau42I were cloned and sequenced, and found to
consist of two overlapping reading frames, ORF S (specificity) and ORF
RM (restriction-modification), in the same orientation. The ORFs share
a high degree of DNA and amino acid sequence homology with the
previously characterized BcgI R-M system of Bacillus coagulans.
Expression of the cloned Sau421 ORF S and ORF RM in S. aureus 80CR3
transformants from a plasmid vector conferred resistance to lysis by
all 23 IBS phages. Similarly, transformants of S. aureus RN4220
harbouring recombinant plasmids containing both ORFs were resistant to
lysis by the IBS typing phages. However, transformants harbouring
plasmids encoding either ORF S or ORF RM were susceptible to lysis by
the IBS phages, and they had the same phage-susceptibility pattern as
the respective parental isolates. In vitro analysis of crude and
partially purified extracts of S. aureus transformants harbouring both
the 042 ORF S and ORF RM genes indicated that Sau421 has endonuclease
activity and requires co-factors Mg2+ and S-adenosylmethionine in order
to function, and activity is optimized at pH 8, although the precise
recognition sequence has yet to be determined. The findings of this
study confirm that phi 42 is a quadruple-converting phage, believed to
be the first described for S. aureus, and show that it encodes a novel
R-M system termed Sau421.

<>

<1>den Bakker, H.C., Moreno, S., A, I., Govoni, G., Cummings, C.A., Ranieri, M.L., Degoricija, L., Hoelzer, K., Rodriguez-Rivera, L.D., Brown, S., Bolchacova, E., Furtado, M.R., Wiedmann, M.
<2>Genome sequencing reveals diversification of virulence factor content and possible host adaptation in distinct subpopulations of Salmonella enterica.
<3>BMC Genomics
<4>12
<5>425
<6>2011
<7>BACKGROUND: Divergence of bacterial populations into distinct subpopulations is
often the result of ecological isolation. While some studies have suggested the
existence of Salmonella enterica subsp. enterica subclades, evidence for these
subdivisions has been ambiguous. Here we used a comparative genomics approach to
define the population structure of Salmonella enterica subsp. enterica, and
identify clade-specific genes that may be the result of ecological
specialization. RESULTS: Multi-locus sequence analysis (MLSA) and single
nucleotide polymorphisms (SNPs) data for 16 newly sequenced and 30 publicly
available genomes showed an unambiguous subdivision of S. enterica subsp.
enterica into at least two subpopulations, which we refer to as clade A and clade
B. Clade B strains contain several clade-specific genes or operons, including a
beta-glucuronidase operon, a S-fimbrial operon, and cell surface related genes,
which strongly suggests niche specialization of this subpopulation. An additional
set of 123 isolates was assigned to clades A and B by using qPCR assays targeting
subpopulation-specific SNPs and genes of interest. Among 98 serovars examined,
approximately 20% belonged to clade B. All clade B isolates contained two
pathogenicity related genomic islands, SPI-18 and a cytolethal distending toxin
islet; a combination of these two islands was previously thought to be exclusive
to serovars Typhi and Paratyphi A. Presence of beta-glucuronidase in clade B
isolates specifically suggests an adaptation of this clade to the vertebrate
gastrointestinal environment. CONCLUSIONS: S. enterica subsp. enterica consists
of at least two subpopulations that differ specifically in genes involved in host
and tissue tropism, utilization of host specific carbon and nitrogen sources and
are therefore likely to differ in ecology and transmission characteristics.

<>

<1>Denes, T., Vongkamjan, K., Ackermann, H.W., Moreno, S., A, I., Wiedmann, M., den Bakker, H.C.
<2>Comparative genomic and morphological analysis of Listeria phages isolated from farm environments.
<3>Appl. Environ. Microbiol.
<4>80
<5>4616-4625
<6>2014
<7>The genus Listeria is ubiquitous in the environment and includes the globally
important foodborne pathogen Listeria monocytogenes. While the genomic diversity
of Listeria has been well studied, considerably less is known about the genomic
and morphological diversity of Listeria bacteriophages. In this study, we
sequenced and analyzed the genomes of 14 Listeria phages mostly isolated from New
York dairy farm environments, as well as one related Enterococcus faecalis phage,
to obtain information on genome characteristics and diversity. We also examined
12 of the phages by electron microscopy to characterize their morphology. These
Listeria phages, based on gene orthology and morphology, together with previously
sequenced Listeria phages could be classified into five orthoclusters, including
one novel orthocluster. One orthocluster (I) consists of large genome (
approximately 135 kb) myoviruses belonging to the genus "Twort-like viruses",
three orthoclusters (II-IV) contain small genome (36-43 kb) siphoviruses with
icosahedral heads, and the novel orthocluster V contains medium-sized genome (
approximately 66 kb) siphoviruses with elongated heads. A novel orthocluster (VI)
of E. faecalis phages, with medium-sized genomes ( approximately 56 kb), was
identified which grouped together and shares morphological features with the
novel Listeria phage orthocluster V. This new group of phages (i.e.,
orthoclusters V and VI) is composed of putative lytic phages that may prove to be
useful in phage-based applications for biocontrol, detection, and therapeutic
purposes.

<>

<1>Deng, H., Yang, X., Yeo, S.Pei.Xing., Gao, Z.
<2>Highly Sensitive Electrochemical Methyltransferase Activity Assay.
<3>Anal. Chem.
<4>86
<5>2117-2123
<6>2014
<7>A simple and highly sensitive electrochemical DNA methyltransferase (MTase) activity assay is
presented in this report. The assay employs the electrocatalytic oxidation of ascorbic acid
(AA) by a threading intercalator (N,N'-bis(3propylimidazole)-1,4,5,8-naphthalene diimide
(PIND) functionalized with electrocatalytic redox Os(bpy)(2)Cl+ moieties (PIND-Os)). Briefly,
a double-stranded DNA (ds-DNA) containing the symmetric sequence of 5'-CCGG-3' is first
immobilized on a gold electrode. The electrode is then incubated with M.SssI CpG
methyltransferase (M.SssI MTase) which catalyzes the methylation of the specific CpG
dinucleotides, and the electrode is subsequently treated with a restriction endonuclease HpaII
which recognizes the 5'-CCGG-3' sequence. Once the CpG site in the 5'-CCGG-3' is
methylated, HpaII recognition is blocked. Higher M.SssI MTase activity leads to more CpG sites
being methylated and consequently impedes more the restriction endonuclease HpaII digestion
process. Thus, a larger amount of ds-DNA remains on the electrode surface after the HpaII.
treatment. Thereafter, the electrode is incubated with PIND-Os during which PIND-Os
specifically inserts itself between base pairs of ds-DNA and catalyzes the electrooxidation of
AA. The methylation event corresponding to the MTase activity can therefore be monitored and
amplified by the electrocatalytic oxidation of AA. A linear correlation between the catalytic
oxidation current of AA and the activity of M.SssI MTase ranged from 0 to 120 U/mL with a
current sensitivity of 0.046 mu A mL U-1 is obtained. The inhibitor screening ability of the
developed MTase activity assay is also demonstrated.

<>

<1>Deng, J., Szyf, M.
<2>Multiple isoforms of DNA methyltransferase are encoded by the vertebrate cytosine DNA methyltransferase gene.
<3>J. Biol. Chem.
<4>273
<5>22869-22872
<6>1998
<7>This manuscript tests the hypothesis that multiple forms of cytosine-DNA methyltransferase are
expressed in vertebrates in vivo.  Vertebrate genomes are distinguished by tissue- and
gene-specific DNA methylation patterns.  Specific methylation patterns are believed to encode
epigenetic information.  In distinction from the remarkable diversity of DNA methylation
patterns, only one functional DNA MeTase cDNA has been identified to date in different
vertebrate organisms.  Using reverse transcription-polymerase chain reaction and RNase
protection analyses, we show that the methyltransferase domain of the rat DNA MeTase is
alternatively spliced in vivo, generating different in-frame variants of DNA MeTase in
specific tissues.  This process is developmentally regulated and is induced in PC12 cells by a
known inducer of neuronal differentiation, nerve growth factor.  The data presented here point
toward a new mechanism for generating diversity of DNA MeTases and possibly diverse DNA
methylation patterns.

<>

<1>Deng, W. et al.
<2>Genome sequence of Yersinia pestis KIM.
<3>J. Bacteriol.
<4>184
<5>4601-4611
<6>2002
<7>We present the complete genome sequence of Yersinia pestis KIM, the etiologic agent of bubonic
and pneumonic plague. The strain KIM, biovar Mediaevalis, is associated with the second
pandemic, including the Black Death. The 4.6-Mb genome encodes 4,198 open reading frames
(ORFs). The origin, terminus, and most genes encoding DNA replication proteins are similar to
those of Escherichia coli K-12. The KIM genome sequence was compared with that of Y. pestis
CO92, biovar Orientalis, revealing homologous sequences but a remarkable amount of genome
rearrangement for strains so closely related. The differences appear to result from multiple
inversions of genome segments at insertion sequences, in a manner consistent with present
knowledge of replication and recombination. There are few differences attributable to
horizontal transfer. The KIM and E. coli K-12 genome proteins were also compared, exposing
surprising amounts of locally colinear "backbone," or synteny, that is not discernible at the
nucleotide level. Nearly 54% of KIM ORFs are significantly similar to K-12 proteins, with
conserved housekeeping functions. However, a number of E. coli pathways and transport systems
and at least one global regulator were not found, reflecting differences in lifestyle between
them. In KIM-specific islands, new genes encode candidate pathogenicity proteins, including
iron transport systems, putative adhesins, toxins, and fimbriae.

<>

<1>Deng, W., Liou, S.R., Plunkett, I.I.I.G., Mayhew, G.F., Rose, D.J., Burland, V., Kodoyianni, V., Schwartz, D.C., Blattner, F.R.
<2>Comparative Genomics of Salmonella enterica Serovar Typhi Strains Ty2 and CT18.
<3>J. Bacteriol.
<4>185
<5>2330-2337
<6>2003
<7>We present the 4.8-Mb complete genome sequence of Salmonella enterica serovar Typhi strain
Ty2, a human-specific pathogen causing typhoid fever.
A comparison with the genome sequence of recently isolated S. enterica
serovar Typhi strain CT18 showed that 29 of the 4,646 predicted genes in
Ty2 are unique to this strain, while 84 genes are unique to CT18. Both
genomes contain more than 200 pseudogenes; 9 of these genes in CT18 are
intact in Ty2, while 11 intact CT18 genes are pseudogenes in Ty2. A
half-genome interreplichore inversion in Ty2 relative to CT18 was
confirmed. The two strains exhibit differences in prophages, insertion
sequences, and island structures. While CT18 carries two plasmids, one
conferring multiple drug resistance, Ty2 has no plasmids and is sensitive
to antibiotics.

<>

<1>Deng, X., Salazar, J.K., Frezet, S., Maccannell, D., Ribot, E.M., Fields, P.I., Fricke, W.F., Zhang, W.
<2>Genome Sequence of Salmonella enterica Serotype Tennessee Strain CDC07-0191, Implicated in the 2006-2007 Multistate Food-Borne Outbreak Linked to Peanut  Butter in the United States.
<3>Genome Announcements
<4>1
<5>e00260-13
<6>2013
<7>Salmonella enterica serotype Tennessee strain CDC07-0191 was isolated from the 2006-2007
multistate food-borne outbreak linked to peanut butter in the United
States. Here we report a high-quality draft assembly of the genome sequence of
this strain, derived from a patient. This is the first reported high-quality
draft genome sequence for S. enterica serotype Tennessee, which will enable
in-depth studies of its transmission and virulence.

<>

<1>Deng, Y., Zhu, Y., Wang, P., Zhu, L., Zheng, J., Li, R., Ruan, L., Peng, D., Sun, M.
<2>Complete Genome Sequence of Bacillus subtilis BSn5, an Endophytic Bacterium of Amorphophallus konjac with Antimicrobial Activity for the  Plant Pathogen Erwinia carotovora subsp. carotovora.
<3>J. Bacteriol.
<4>193
<5>2070-2071
<6>2011
<7>Here, we present the complete genome sequence of Bacillus subtilis strain BSn5, isolated from
Amorphophallus konjac calli tissue and showing strong
inhibitory activity to Erwinia carotovora subsp. carotovora, which causes
Amorphophallus soft rot disease and affects the industry development of
this organism.

<>

<1>Deng, Y.M., Liu, C.Q., Dunn, N.W.
<2>LldI, a plasmid-encoded type I restriction and modification system in Lactococcus lactis.
<3>DNA Seq.
<4>11
<5>239-245
<6>2000
<7>A plasmid-encoded type I restriction and modification (R-M) system, designated LldI, was
identified in Lactococcus lactis biovar diacetylactis LD10-1. LldI consists of three genes
encoding endonuclease, methylase and specificity subunits, respectively. RT-PCR analysis
revealed that the three genes are co-transcribed as a polycistronic mRNA in L. lactis. The
specificity subunit of LldI differs significantly in the target recognition domains from those
of other type I R-M systems, suggesting that LldI confers a novel specificity in L. lactis.

<>

<1>Denjmuchametov, M.M., Ruban, N.M., Zakharova, M.V., Beletzkaja, I.V., Kravetz, A.N., Solonin, A.S.
<2>Eco27kI, a new isoschizomer of AvaI from Escherichia coli.
<3>Nucleic Acids Res.
<4>20
<5>1992
<6>1992
<7>A small collection (108) of Escherichia coli strains isolated in Kiev was surveyed for type II
restriction endonucleases. 95 strains produced these enzymes; among them Eco27kI, a new
isoschizomer of AvaI, recognized the palindromic sequence 5'-C/PyCGPuG-3'.

<>

<1>Denjmukhametov, M.M., Brevnov, M.G., Zakharova, M.V., Repyk, A.V., Solonin, A.S., Petrauskene, O.V., Gromova, E.S.
<2>The Ecl18kI restriction-modification system: cloning, expression, properties of the purified enzymes.
<3>FEBS Lett.
<4>433
<5>233-236
<6>1998
<7>Ecl18kI is a type II restriction-modification system isolated from Enterobacter cloaceae 18kI
strain.  Genes encoding Ecl18kI methyltransferase (M.Ecl18kI) and Ecl18kI restriction
endonuclease (R.Ecl18kI) have been cloned and expressed in Escherichia coli.  These enzymes
recognize the 5'.../CCNGG...3' sequence in DNA; M.Ecl18kI methylates the C5 carbon atom of
the inner dC residue and R.Ecl18kI cuts DNA as shown by the arrow.  The restriction
endonuclease and the methyltransferase were purified from E. coli B834 [p18Ap1] cells to near
homogeneity.  The restriction endonuclease is present in the solution as a tetramer, while the
methyltransferase is a monomer.  The interactions of M.Ecl18kI and R.Ecl18kI with
1,2-dideoxy-D-ribofuranose containing DNA duplexes were investigated.  The target base
flipping-out mechanism is applicable in the case of M.Ecl18kI.  Correct cleavage of the abasic
substrates by R.Ecl18kI is accompanied by non-canonical hydrolysis of the modified strand.

<>

<1>Denjmukhametov, M.M., Kravets, A.N.
<2>New Escherichia coli strain produces restriction endonuclease Eco27KI - new strain for large-scale production of restriction enzyme.
<3>Russian Patent Office
<4>RU 2044053
<5>
<6>1995
<7>
<>

<1>Denjmukhametov, M.M., Kravets, A.N., Solonin, A.S.
<2>An Escherichia coli strain, a producer of restriction endonuclease Eco71kl - in high yield.
<3>Russian Patent Office
<4>RU 2070925 C
<5>
<6>1996
<7>
<>

<1>Denjmukhametov, M.M., Zakharova, M.V., Kravets, A.N., Pertsev, A.V., Sineva, E.V., Repik, A.V., Beletskaya, I.V., Gromova, E.S., Solonin, A.S.
<2>Characterization of plasmids encoding type II restriction-modification systems, isoschizomers of SsoII.
<3>Mol. Biol. (Mosk)
<4>31
<5>831-838
<6>1997
<7>Five strains of Enterobacteriaceae family have been revealed that have
restriction-modification systems of type II, isoschizomers of SsoII.  The genes coding for the
restriction endonucleases and DNA methyltransferases are located on plasmids of two types. The
plasmids differ in size, capacity for mobilization by conjugative plasmids, and structure of
locus cer.  Recombinant two-plasmid strains were obtained containing plasmids with genes for
Ecl18kI and EcoRII restriction endonucleases in the presence of DNA methyltransferase Ecl18kI.
The stability of the two-plasmid system with the ecoRIIR and ecl18kIM genes implies similar
mechanisms of concerted regulation of gene expression in the restriction-modification systems
Ecl18kI and EcoRI.  Genes for Ecl18kI and SsoII restriction-modification systems have
essentially the same sequence.

<>

<1>Denou, E., Bruttin, A., Barretto, C., Ngom-Bru, C., Brussow, H., Zuber, S.
<2>T4 phages against Escherichia coli diarrhea: potential and problems.
<3>Virology
<4>388
<5>21-30
<6>2009
<7>A combination of in vitro and in vivo experiments with comparative phage
genomics was used for the rational design of a phage cocktail against E.
coli diarrhea. Orally applied T4 coliphages representing three different
subgroups (T4-, RB49- and JS98-like phages) had no negative impact on the
murine gut microbiota. T4 phages were found with high titers in the cecum
and colon and lower titers in the small intestine, but were not detected
in the blood, liver or spleen. No adverse effects were observed after
one-month exposure to phage nor were serum anti-T4 antibodies detected. T4
phages belonging to the same subgroup showed closely related genomes that
differed by 12 (phage JS10 vs. JS98 reference) to 17 (phage JSE vs. RB49
reference) insertion/deletions mostly representing single small ORFs.
Bioinformatic analysis did not reveal undesired genes in the T4 genomes.
Sequence variability was seen over the tail fibre genes, but the
variability did not correlate with phage host range. The investigated T4
phages were not only species- but also strain-specific, necessitating the
use of phage cocktails consisting of 10 and 16 T4 phage isolates to cover
half to two thirds of E. coli strains representing the five main
pathotypes isolated from diarrhea patients.

<>

<1>Denovan-Wright, E.M., Nedelcu, A.M., Lee, R.W.
<2>Complete sequence of the mitochondrial DNA of Chlamydomonas eugametos.
<3>Plant Mol. Biol.
<4>36
<5>285-295
<6>1998
<7>The complete  nucleotide sequence of the Chlamydomonas eugametos mitochondrial genome has been
determined (22,897 bp, 34.6% G+C).  The genes identified in this circular-mapping genome
include those for apocytochrome b, subunit 1 of the cytochrome oxidase complex.  Subunits
1,2,4,5, and 6 of the N dehydrogenase complex, discontinuous large and small subunit ribosomal
rRNAs and three tRNAs whose anticodons CAU, CCA and UUG are specific for methionine,
tryptophan and glutamine, respectively.  The C. eugametos mitochondrial DNA, therefore, shares
almost the same reduced set of coding functions and similar unusual features of rRNA gene
organization with the linear 15.8 kb mtDNA of Chlamydomonas reinhardtii, the only other
completely sequenced chlamydomonadalean mtDNA.  However, sequence analysis of the C. eugametos
mtDNA has revealed the following distinguishing features relative to those of C. reinhardtii:
(1) the absence of a reverse transcriptase-like gene homologue, (2) the presence of an
addition gene for tRNAmet that may be a pseudogene, (3) a completely different gene order, (4)
transcription of all genes from the same mtDNA strand, (5) a lower G+ C content, (6) less
pronounced bias in codon usage, and (7) nine group I introns, several of which contain open
reading frames coding for potential maturases/endonucleases and two have a nucleotide at the
5' or 3' splice site of the deduced precursor RNAs that deviates from highly conserved
nucleotides reported in other group I introns.  The features of mitochondrial genome
organization and gene content shared by C. eugametos and C. reinhardtii contrast with their
green algal mtDNAs that have been characterized in detail.  The deep evolutionary divergence
between these two Chlamydomonas taxa within the Chlamydomonadales suggests that their shared
features of mitochondrial genome organization evolved prior to the origin of this group.

<>

<1>deOliveira-Cunha, C., Goda-Zuleta, L.F., Paula-deAlmeida, L.G., Prioli-Ciapina, L., Lustrino-Borges, W., Pitard, R.M., Baldani, J.I., Straliotto, R., deFaria, S.M., Hungria, M., Sousa-Cavada, B., Mercante, F.M., Ribeiro-deVasconcelos, A.T.
<2>Complete Genome Sequence of Burkholderia phenoliruptrix BR3459a (CLA1), a Heat-Tolerant, Nitrogen-Fixing Symbiont of Mimosa flocculosa.
<3>J. Bacteriol.
<4>194
<5>6675-6676
<6>2012
<7>The genus Burkholderia represents a challenge to the fields of taxonomy and phylogeny and,
especially, to the understanding of the contrasting roles as
either opportunistic pathogens or bacteria with biotechnological potential. Few
genomes of nonpathogenic strains, especially of diazotrophic symbiotic bacteria,
have been sequenced to improve understanding of the genus. Here, we contribute
with the complete genome sequence of Burkholderia phenoliruptrix strain BR3459a
(CLA1), an effective diazotrophic symbiont of the leguminous tree Mimosa
flocculosa Burkart, which is endemic to South America.

<>

<1>DePaoli-Roach, A., Roach, P.J., Zucker, K.E., Smith, S.S.
<2>Selective phosphorylation of human DNA methyltransferase by protein kinase C.
<3>FEBS Lett.
<4>197
<5>149-153
<6>1986
<7>Human DNA methyltransferase, the enzyme thought to be responsible for the somatic inheritance
of patterns of DNA methylation, is an effective substrate for phosphorylation by protein
kinase C. This provides a plausible mechanistic link between the action of tumor promoting
phorbol esters, which stimulate protein kinase C, and abnormal patterns of DNA methylation
often observed in transformed cells.

<>

<1>Deplus, R., Brenner, C., Burgers, W.A., Putmans, P., Kouzarides, T., de Launoit, Y., Fuks, F.
<2>Dnmt3L is a transcriptional repressor that recruits histone deacetylase.
<3>Nucleic Acids Res.
<4>30
<5>3831-3838
<6>2002
<7>The Dnmt3L protein belongs to the Dnmt3 family of DNA methyltransferases by virtue of its
sequence homology in the plant homeodomain (PHD)-like motif. Dnmt3L is essential for the
establishment of maternal genomic imprints and, given its lack of key methyltransferase
motifs, is more likely to act as a regulator of methylation rather than as an enzyme that
methylates DNA. Here, we show that Dnmt3L, like Dnmt3a and Dnmt3b, interacts both in vitro and
in vivo with the histone deacetylase HDAC1. Consistent with the binding to a deacetylase,
Dnmt3L purifies histone deacetylase activity from nuclear extracts. We find that Dnmt3L can
repress transcription and that this repression is dependent on HDAC1 and is relieved by
treatment with the HDAC inhibitor trichostatin A. Binding of Dnmt3L to HDAC1 as well as its
repressive function require the PHD-like motif. Our results indicate that Dnmt3L plays a role
in transcriptional regulation and that recruitment of the HDAC repressive machinery is a
shared and conserved feature of the Dnmt3 family. The fact that, despite the absence of a
methyltransferase domain, Dnmt3L retains the capacity to contact deacetylase further
substantiates the notion that the Dnmts can repress transcription independently of their
methylating activities.

<>

<1>Deppenmeier, U. et al.
<2>The genome of Methanosarcina mazei: evidence for lateral gene transfer between bacteria and archaea.
<3>J. Mol. Microbiol. Biotechnol.
<4>4
<5>453-461
<6>2002
<7>The Archaeon Methanosarcina mazei and related species are of great ecological importance as
they are the only organisms fermenting acetate, methylamines and methanol to methane, carbon
dioxide and ammonia (in case of methylamines). Since acetate is the precursor of 60% of the
methane produced on earth these organisms contribute significantly to the production of this
greenhouse gas, e.g. in rice paddies. The 4,096,345 base pairs circular chromosome of M. mazei
is more than twice as large as the genomes of the methanogenic Archaea currently completely
sequenced (Bult et al., 1996; Smith et al., 1997). 3,371 open reading frames (ORFs) were
identified. Based on currently available sequence data 376 of these ORFs are
Methanosarcina-specific and 1,043 ORFs find their closest homologue in the bacterial domain.
544 of these ORFs reach significant similarity values only in the bacterial domain. They
include 56 of the 102 transposases, and proteins involved in gluconeogenesis, proline
biosynthesis, transport processes, DNA-repair, environmental sensing, gene regulation, and
stress response. Striking examples are the occurrence of the bacterial GroEL/GroES chaperone
system and the presence of tetrahydrofolate-dependent enzymes. These findings might indicate
that lateral gene transfer has played an important evolutionary role in forging the physiology
of this metabolically versatile methanogen.

<>

<1>Deraspe, M., Alexander, D.C., Xiong, J., Ma, J.H., Low, D.E., Jamieson, F.B., Roy, P.H.
<2>Genomic analysis of Pseudomonas aeruginosa PA96, the host of carbapenem resistance plasmid pOZ176.
<3>FEMS Microbiol. Lett.
<4>356
<5>212-216
<6>2014
<7>Pseudomonas aeruginosa PA96 is a clinical isolate from Guangzhou, China, that is
multiresistant to antibiotics. We previously described the 500-kb IncP-2 plasmid, pOZ176 that
encodes many resistance genes including the IMP-9 carbapenemase. Whole-genome sequencing of
PA96 enabled characterization of its genomic islands, virulence factors, and chromosomal
resistance genes. We filled gaps using PCR and used optical mapping to confirm the correct
contig order. We automatically annotated the core genome and manually annotated the genomic
islands. The genome is 6 444 091 bp and encodes 5853 ORFs. From the whole-genome sequence, we
constructed a physical map and constructed a phylogenetic tree for comparison with sequenced
P. aeruginosa strains. Analysis of known core genome virulence factors and resistance genes
revealed few differences with other strains, but the major virulence island is closer to that
of DK2 than to PA14. PA96 most closely resembles the environmental strain M18, and notably
shares a common serotype, pyoverdin type, flagellar operon, type IV pilin, and several genomic
islands with M18.

<>

<1>Derbise, A., Chenal-Francisque, V., Huon, C., Fayolle, C., Demeure, C.E., Chane-Woon-Ming, B., Medigue, C., Hinnebusch, B.J., Carniel, E.
<2>Delineation and Analysis of Chromosomal Regions Specifying Yersinia pestis.
<3>Infect. Immun.
<4>78
<5>3930-3941
<6>2010
<7>Yersinia pestis, the causative agent of plague, has recently diverged from the less virulent
enteropathogen Yersinia pseudotuberculosis. Its
emergence has been characterized by massive genetic loss and
inactivation and limited gene acquisition. The acquired genes include
two plasmids, a filamentous phage, and a few chromosomal loci. The aim
of this study was to characterize the chromosomal regions acquired by
Y. pestis. Following in silico comparative analysis and PCR screening
of 98 strains of Y. pseudotuberculosis and Y. pestis, we found that
eight chromosomal loci (six regions [R1(pe) to R6(pe)] and two coding
sequences [CDS1(pe) and CDS2(pe)]) specified Y. pestis. Signatures of
integration by site specific or homologous recombination were
identified for most of them. These acquisitions and the loss of
ancestral DNA sequences were concentrated in a chromosomal region
opposite to the origin of replication. The specific regions were
acquired very early during Y. pestis evolution and were retained during
its microevolution, suggesting that they might bring some selective
advantages. Only one region (R3(pe)), predicted to carry a lambdoid
prophage, is most likely no longer functional because of mutations.
With the exception of R1(pe) and R2(pe), which have the potential to
encode a restriction/modification and a sugar transport system,
respectively, no functions could be predicted for the other Y.
pestis-specific loci. To determine the role of the eight chromosomal
loci in the physiology and pathogenicity of the plague bacillus, each
of them was individually deleted from the bacterial chromosome. None of
the deletants exhibited defects during growth in vitro. Using the
Xenopsylla cheopis flea model, all deletants retained the capacity to
produce a stable and persistent infection and to block fleas.
Similarly, none of the deletants caused any acute flea toxicity. In the
mouse model of infection, all deletants were fully virulent upon
subcutaneous or aerosol infections. Therefore, our results suggest that
acquisition of new chromosomal materials has not been of major
importance in the dramatic change of life cycle that has accompanied
the emergence of Y. pestis.

<>

<1>Derbyshire, V., Kowalski, J.C., Dansereau, J.T., Hauer, C.R., Belfort, M.
<2>Two-domain structure of the td intron-encoded endonuclease I-TevI correlates with the two-domain configuration of the homing site.
<3>J. Mol. Biol.
<4>265
<5>494-506
<6>1997
<7>I-TevI, the T4 td intron-encoded endonuclease, catalyzes the first step in intron homing by
making a double-strand break in the intronless allele within a sequence designated the homing
site.  The 28 kDa enzyme, which interacts with the homing site over a span of 37 bp, binds as
a monomer, contacting two domains of the substrate.  In this study, limited proteolysis
experiments indicate that I-TevI consists of two domains that behave as discrete physical
entities as judged by a number of functional and structural criteria.  Overexpression clones
for each domain were constructed and the proteins were purified.  The carboxy-terminal domain
has DNA-binding activity coincident with the primary binding region of the homing site and
binds with the same affinity as the full-length enzyme.  The isolated amino-terminal domain,
contains the conserved GIY-YIG motif, consistent with its being the catalytic domain.
Furthermore, site-directed mutagenesis of a conserved arginine residue within the extended
motif rendered the full-length protein catalytically inactive, although DNA-binding was
maintained.  This is the first evidence that the GIY-YIG motif is important for catalytic
activity.  An enzyme with an N-terminal catalytic domain and a C-terminal DNA-binding domain
connected by a flexible linker is in accord with the bipartite structure of the homing site.

<>

<1>Desai, P.T., Porwollik, S., Long, F., Cheng, P., Wollam, A., Bhonagiri-Palsikar, V., Hallsworth-Pepin, K., Clifton, S.W., Weinstock, G.M., McClelland, M.
<2>Evolutionary Genomics of Salmonella enterica Subspecies.
<3>MBio
<4>4
<5>e00198-13
<6>2013
<7>Six subspecies are currently recognized in Salmonella enterica. Subspecies I (subspecies
enterica) is responsible for nearly all infections in humans and warm-blooded animals, while
five other subspecies are isolated principally from coldblooded animals. We sequenced 21
phylogenetically diverse strains, including two representatives from each of the previously
unsequenced five subspecies and 11 diverse new strains from S. enterica subspecies enterica,
to put this species into an evolutionary perspective. The phylogeny of the subspecies was
partly obscured by abundant recombination events between lineages and a relatively short
period of time within which subspeciation took place. Nevertheless, a variety of different
tree-building methods gave congruent evolutionary tree topologies for subspeciation. A total
of 285 gene families were identified that were recruited into subspecies enterica, and most of
these are of unknown function. At least 2,807 gene families were identified in one or more of
the other subspecies that are not found in subspecies I or Salmonella bongori. Among these
gene families were 13 new candidate effectors and 7 new candidate fimbrial clusters. A third
complete type III secretion system not present in subspecies enterica (I) isolates was found
in both strains of subspecies salamae (II). Some gene families had complex taxonomies, such as
the type VI secretion systems, which were recruited from four different lineages in five of
six subspecies. Analysis of nonsynonymous-to-synonymous substitution rates indicated that the
more-recently acquired regions in S. enterica are undergoing faster fixation rates than the
rest of the genome. Recently acquired AT-rich regions, which often encode virulence functions,
are under ongoing selection to maintain their high AT content.

<>

<1>Deschamps, P., Zivanovic, Y., Moreira, D., Rodriguez-Valera, F., Lopez-Garcia, P.
<2>Pangenome evidence for extensive interdomain horizontal transfer affecting lineage core and shell genes in uncultured planktonic thaumarchaeota and euryarchaeota.
<3>Genome Biol. Evol.
<4>6
<5>1549-1563
<6>2014
<7>Horizontal gene transfer (HGT) is an important force in evolution, which may
lead, among other things, to the adaptation to new environments by the import of
new metabolic functions. Recent studies based on phylogenetic analyses of a few
genome fragments containing archaeal 16S rRNA genes and fosmid-end sequences from
deep-sea metagenomic libraries have suggested that marine planktonic archaea
could be affected by high HGT frequency. Likewise, a composite genome of an
uncultured marine euryarchaeote showed high levels of gene sequence similarity to
bacterial genes. In this work, we ask whether HGT is frequent and widespread in
genomes of these marine archaea, and whether HGT is an ancient and/or recurrent
phenomenon. To answer these questions, we sequenced 997 fosmid archaeal clones
from metagenomic libraries of deep-Mediterranean waters (1,000 and 3,000 m depth)
and built comprehensive pangenomes for planktonic Thaumarchaeota (Group I
archaea) and Euryarchaeota belonging to the uncultured Groups II and III
Euryarchaeota (GII/III-Euryarchaeota). Comparison with available reference
genomes of Thaumarchaeota and a composite marine surface euryarchaeote genome
allowed us to define sets of core, lineage-specific core, and shell gene ortholog
clusters for the two archaeal lineages. Molecular phylogenetic analyses of all
gene clusters showed that 23.9% of marine Thaumarchaeota genes and 29.7% of
GII/III-Euryarchaeota genes had been horizontally acquired from bacteria. HGT is
not only extensive and directional but also ongoing, with high HGT levels in
lineage-specific core (ancient transfers) and shell (recent transfers) genes.
Many of the acquired genes are related to metabolism and membrane biogenesis,
suggesting an adaptive value for life in cold, oligotrophic oceans. We
hypothesize that the acquisition of an important amount of foreign genes by the
ancestors of these archaeal groups significantly contributed to their divergence
and ecological success.

<>

<1>Deschavanne, P., Radman, M.
<2>Counterselection of GATC sequences in enterobacteriophages by the components of the methyl-directed mismatch repair system.
<3>J. Mol. Evol.
<4>33
<5>125-132
<6>1991
<7>Weak to severe deficit of GATC sequences in the DNA of enterobacteriophages appears to be
correlated with their undermethylation during growth in dam+ (GATC ade-methylase) bacteria.
This observation is corroborated by the sequence analysis showing no evidence for
site-specific mutagenicity of 6meAde. The MutH protein of the methyl-directed mismatch repair
system recognizes and cleaves the undermethylated GATC sequences in the course of mismatch
repair. To enquire whether the MutH function of the methyl-directed mismatch repair system
participates in counterselection of GATC sequences in enterobacteriophages, we have studied
the yield of bacteriophage PhiX174 containing either 0, 1, or 2 GATC sequences, in wild type,
dam, and mut (H,L,S,U) Escherichia coli. Following transfection with unmethylated DNA
containing two GATC sequences, a net decrease in the yield of infective particles was observed
in all bacterial mutH+ dam- strains, whereas no detectable decrease was observed in bacteria
infected by DNA without GATC sequence. This effect of the MutH function is maximum in wild
type and mutL and mutS bacteria whereas the effect is not significant in mutU bacteria,
suggesting an interaction of the helicase II with the MutH protein.

However, in dam+ bacteria, the presence of GATC sequences leads to an increased yield of
infective particles. The effect of GATC sequence and its Dam methylation system on phage yield
in mutH- bacteria reveals that methylated GATC sequences are advantageous to the phage. These
results suggest that the methyl-directed mismatch repair system, and in particular its MutH
protein, may have participated in severe counterselection of GATC sequences from
enterobacteriophages, presumably by DNA cleavage or by interfering with DNA replication or
packaging when GATC sequences are undermethylated. Coevolution of the Dam and MutH proteins
could then account for the loss of GATC sequences from DNA of bacteriophages growing in dam+
hosts.


<>

<1>Descloux, S., Rossano, A., Perreten, V.
<2>Characterization of new Staphylococcal Cassette Chromosome mec (SCCmec) and topoisomerase genes in fluoroquinolone and methicillin-resistant Staphylococcus pseudintermedius.
<3>J. Clin. Microbiol.
<4>46
<5>1818-1823
<6>2008
<7>Fluoroquinolone and methicillin-resistant Staphylococcus pseudintermedius isolates harbor two
new Staphylococcal Cassette Chromosome (SCCmec) which belong to class A, allotype 3 (SCCmec
II-III) and to the new allotype 5 (SCCmec VII). Analysis of the complete nucleotide sequence
of the topoisomerase loci gyrB/A and grlB/A revealed mutations involved in fluoroquinolone
resistance.

<>

<1>Deshpande, N.P., Wong, Y.K., Manefield, M., Wilkins, M.R., Lee, M.
<2>Genome Sequence of Dehalobacter UNSWDHB, a Chloroform-Dechlorinating Bacterium.
<3>Genome Announcements
<4>1
<5>e00720-13
<6>2013
<7>The chloroform-respiring bacterium Dehalobacter UNSWDHB was isolated from subsurface soil
contaminated with a mixture of organohalides, including
chloroform. Here, we present its 3.2-Mb genome.

<>

<1>Deshuillers, P.L., Santos, A.P., do Nascimento, N.C., Hampel, J.A., Bergin, I.L., Dyson, M.C., Messick, J.B.
<2>Complete Genome Sequence of Mycoplasma ovis Strain Michigan, a Hemoplasma of Sheep with Two Distinct 16S rRNA Genes.
<3>Genome Announcements
<4>2
<5>e01235-13
<6>2014
<7>We report the complete genome sequence of Mycoplasma ovis strain Michigan. Its single circular
chromosome has 702,511 bp and contains 2 copies of the 16S rRNA
gene, one corresponding to M. ovis and the other to 'Candidatus Mycoplasma
haemovis.' All housekeeping genes and the 5S-23S rRNA genes are present in single
copies.

<>

<1>Desiere, F., McShan, W.M., van Sinderen, D., Ferretti, J.J., Brussow, H.
<2>Comparative Genomics Reveals Close Genetic Relationships between Phages from Dairy Bacteria and Pathogenic Streptococci: Evolutionary Implications for Prophage-Host Interactions.
<3>Virology
<4>288
<5>325-341
<6>2001
<7>The genome of the highly pathogenic M1 serotype Streptococcus pyogenes isolate SF370 contains
eight prophage elements. Only prophage SF370.1 could be induced by mitomycin C treatment.
Prophage SF370.3 showed a 33.5-kb-long genome that closely resembled the genome organization
of the cos-site temperate Siphovirus r1t infecting the dairy bacterium Lactococcus lactis. The
two-phage genomes shared between 60 and 70% nucleotide sequence identity over the DNA
packaging, head and tail genes. Analysis of the SF370.3 genome revealed mutations in the
replisome organizer gene that may prevent the induction of the prophage. The mutated phage
replication gene was closely related to a virulence marker identified in recently emerged M3
serotype S. pyogenes strains in Japan. This observation suggests that prophage genes confer
selective advantage to the lysogenic host. SF370.3 encodes a hyaluronidase and a DNase that
may facilitate the spreading of S. pyogenes through tissue planes of its human host. Prophage
SF370.2 showed a 43-kb-long genome that closely resembled the genome organization of pac-site
temperate Siphoviridae infecting the dairy bacteria S. thermophilus and L. lactis. Over part
of the structural genes, the similarity between SF370.2 and S. thermophilus phage O1205
extended to the nucleotide sequence level. SF370.2 showed two probable inactivating mutations:
one in the replisome organizer gene and another in the gene encoding the portal protein.
Prophage SF370.2 also encodes a hyaluronidase and in addition two very likely virulence
factors: prophage-encoded toxins acting as superantigens that may contribute to the immune
deregulation observed during invasive streptococcal infections. The superantigens are encoded
between the phage lysin and the right attachment site of the prophage genome. The genes were
nearly sequence identical with a DNA segment in S. equi, suggesting horizontal gene transfer.
The trend for prophage genome inactivation was even more evident for the remaining five
prophage sequences that showed massive losses of prophage DNA. In these prophage remnants only
13-0.3 kb of putative prophage DNA was detected. We discuss the genomics data from S. pyogenes
strain SF370 within the framework of Darwinian coevolution of prophages and lysogenic bacteria
and suggest elements of genetic cooperation and elements of an arms race in this host-parasite
relationship.

<>

<1>Desiniotis, A., Kouvelis, V.N., Davenport, K., Bruce, D., Detter, C., Tapia, R., Han, C., Goodwin, L.A., Woyke, T., Kyrpides, N.C., Typas, M.A., Pappas, K.M.
<2>Complete Genome Sequence of the Ethanol-Producing Zymomonas mobilis subsp. mobilis Centrotype ATCC 29191.
<3>J. Bacteriol.
<4>194
<5>5966-5967
<6>2012
<7>Zymomonas mobilis is an ethanologenic bacterium that has been studied for use in  biofuel
production. Of the sequenced Zymomonas strains, ATCC 29191 has been
described as the phenotypic centrotype of Zymomonas mobilis subsp. mobilis, the
taxon that harbors the highest ethanol-producing Z. mobilis strains. ATCC 29191
was isolated in Kinshasa, Congo, from palm wine fermentations. This strain is
reported to be a robust levan producer, while in recent years it has been
employed in studies addressing Z. mobilis respiration. Here we announce the
finishing and annotation of the ATCC 29191 genome, which comprises one chromosome
and three plasmids.

<>

<1>Desirazu, N.R., Reddy, Y.V.
<2>Targetted DNA distortion by EcoP15I DNA methyltransferase.
<3>Biochem. Soc. Trans.
<4>28
<5>A309
<6>2000
<7>EcoP15I DNA methyltransferase, a member of the type III restriction-modification system, binds
to the sequence 5'-CAGCAG-3' transferring a methyl group from S-adenosyl-L-methionine to the
second adenine.  We have investigated protein-DNA interactions in the methylase-DNA complex by
three methods.  Determination of equilibrium dissociation constants indicated that the enzyme
had higher affinity for DNA containing mismatches at the target base within the recognition
sequence.  Potassium permanganate footprinting studies revealed that there was a
hyper-reactive cleavage site coincident with adenine that is the target base for methylation.
More importantly, to detect DNA conformational alterations within the enzyme-DNA complexes, we
have used a fluorescence-based assay, when the enzyme bound to DNA containing 2-aminopurine
substitutions within the cognate sequence, an 8-10 fold fluorescence enhancement resulting
from enzymatic flipping of the target adenine was observed, fluorescence spectroscopy analysis
showed that the changes attributable to structural distortion were specific for only the bases
within the recognition sequence.  Interestingly, we observed that both the adenines in the
recognition site appear to be structurally distorted to the same extent.  While the target
adenine is probably flipped out of the DNA duplex, our results also suggest that fluorescence
enhancements could be derived from protein-DNA interactions other than base flipping.  Taken
together, our results support the proposed base flipping mechanism for adenine
methyltransferases.

<>

<1>Detich, N., Hamm, S., Just, G., Knox, J.D., Szyf, M.
<2>The methyl donor S-adenosylmethionine inhibits active demethylation of DNA: a candidate novel mechanism for the pharmacological effects of s-adenosylmethionine.
<3>J. Biol. Chem.
<4>278
<5>20812-20820
<6>2003
<7>S-Adenosylmethionine (AdoMet) is the methyl donor of numerous methylation reactions. The
current model is that an increased concentration of AdoMet
stimulates DNA methyltransferase reactions, triggering hypermethylation
and protecting the genome against global hypomethylation, a hallmark of
cancer. Using an assay of active demethylation in HEK 293 cells, we show
that AdoMet inhibits active demethylation and expression of an ectopically
methylated CMV-GFP (green fluorescent protein) plasmid in a dose-dependent
manner. The inhibition of GFP expression is specific to methylated GFP;
AdoMet does not inhibit an identical but unmethylated CMV-GFP plasmid.
S-Adenosylhomocysteine (AdoHcy), the product of methyltransferase
reactions utilizing AdoMet does not inhibit demethylation or expression of
CMV-GFP. In vitro, AdoMet but not AdoHcy inhibits methylated DNA-binding
protein 2/DNA demethylase as well as endogenous demethylase activity
extracted from HEK 293, suggesting that AdoMet directly inhibits
demethylase activity, and that the methyl residue on AdoMet is required
for its interaction with demethylase. Taken together, our data support an
alternative mechanism of action for AdoMet as an inhibitor of
intracellular demethylase activity, which results in hypermethylation of
DNA.

<>

<1>Detich, N., Ramchandani, S., Szyf, M.
<2>A conserved 3'-untranslated element mediates growth regulation of DNA methyltransferase 1 and inhibits its transforming activity.
<3>J. Biol. Chem.
<4>276
<5>24881-24890
<6>2001
<7>Ectopic expression of DNA methyltransferase 1 (DNMT1) has been proposed to play an important
role in cancer. dnmt1 mRNA is undetectable in
growth-arrested cells but is induced upon entrance into the S phase of the cell cycle, and
until now, the mechanisms responsible for this regulation were unknown. In this report, we
demonstrate that the 3'-untranslated region (3'-UTR) of the dnmt1 mRNA can confer a
growth-dependent regulation on its own message as well as a heterologous beta-globin mRNA Our
results indicate that a 54-nucleotide highly conserved element within the 3'-UTR is necessary
and sufficient to mediate this regulation. Cell-free mRNA decay experiments demonstrate that
this element increases mRNA turnover rates and does so to a greater extent in the presence of
extracts prepared from arrested cells. A specific RNA-protein complex is formed with the
3'-UTR only in growth-arrested cells, and a UV cross-linking analysis revealed a 40-kDa
protein (p40), the binding of which is dramatically increased in growth-arrested cells and is
inversely correlated with dnmt1 mRNA levels as cells are induced into the cell cycle. Although
ectopic expression of human DNMT1 lacking the 3'-UTR can transform NIH-3T3 cells, inclusion
of the 3'-UTR prevents transformation. These
results support the hypothesis that deregulated expression of DNMT1
with the cell cycle is important for cellular transformation.

<>

<1>Detich, N., Theberge, J., Szyf, M.
<2>Promoter-specific activation and demethylation by MBD2/demethylase.
<3>J. Biol. Chem.
<4>277
<5>35791-35794
<6>2002
<7>MBD2 is the only member of a family of methyl-CpG-binding proteins that has been reported to
be both a transcriptional repressor and a DNA
demethylase (dMTase). To understand the apparently contradictory function
of MBD2/dMTase, we studied the effects of dMTase overexpression on the
activity of various in vitro methylated promoters transiently transfected
into HEK293 cells. We found that forced expression of a MBD2/dMTase
expression vector (His-dMTase) differentially activated two methylated
reporters, pSV40-CAT (the SV40 enhancerless promoter adjacent to the
chloramphenicol acetyltransferase (CAT) reporter gene) and pGL2T+I4xTBRE
(a region of the p21 promoter next to the luciferase reporter gene), in a
time- and dose-dependent manner. His-dMTase increased pSV40-CAT expression
by 3-10-fold after 96 h, while pGL2T+I4xTBRE expression was increased by
2-3-fold after only 48 h and did not further increase at 96 h. Gene
activation was not universal because no effect was seen with the p19-ARF
promoter. We then assessed whether activation might be due to
demethylation within the promoter region. Using bisulfite mapping, we
found that exogenous expression of His-dMTase induced demethylation at 8
of the 10 CpG sites within the SV40 promoter. The observation that
His-dMTase increases the demethylase activity in the cells was also
confirmed using an in vitro CpG demethylase assay with a mC32pG
oligonucleotide substrate and purified Q-Sepharose fractions from HEK293
cells transfected with His-dMTase or empty pcDNA3.1His vector. We propose
that a single protein possessing both repressor and demethylase functions
has evolved to coordinate a program that requires suppression of some
methylated genes and activation of others.

<>

<1>Deutsch, J., Razin, A., Sedat, J.
<2>Analysis of 5-methylcytosine in DNA.
<3>Anal. Biochem.
<4>72
<5>586-592
<6>1976
<7>A method is described for the unambiguous rapid identification and quantitation of the minor
base, 5-methylcytosine, in DNA using high resolution mass spectrometry.  This method can
detect one 5-methylcytosine residue per 5500 nucleotides in Phi X174 DNA, in a sample of less
than 10 micrograms and requires less than 1 micrograms of calf thymus DNA in which the molar
ratio of 5-methylcytosine/cytosine is about 0.05.

<>

<1>Devajyothi, C., Brahmachari, V.
<2>Detection of a CpA methylase in an insect system: Characterization and substrate specificity.
<3>Mol. Cell. Biochem.
<4>110
<5>103-111
<6>1992
<7>a cytosine-specific DNA methyltransferase (EC 2.1.1.37) has been purified to near homogeneity
from a mealybug (Planococcus lilacinus). The enzyme can methylate cytosine residues in CpG
sequences as well as CpA sequences. The apparent molecular weight of the enzymes was estimated
as 135,000 daltons by FPLC. The enzyme exhibits a processive mode of action and a salt
dependance similar to mammalian methylases. Mealybug methylase exhibits a preference for
denatured DNA substrates.

<>

<1>Deveau, A., Gross, H., Morin, E., Karpinets, T., Utturkar, S., Mehnaz, S., Martin, F., Frey-Klett, P., Labbe, J.
<2>Genome Sequence of the Mycorrhizal Helper Bacterium Pseudomonas fluorescens BBc6R8.
<3>Genome Announcements
<4>2
<5>e01152-13
<6>2014
<7>We report the draft genome sequence of the mycorrhizal helper bacterium Pseudomonas
fluorescens strain BBc6R8. This is the first genome of a mycorrhizal
helper bacterium. The draft genome contains 6,952,353 bp and is predicted to
encode 6,317 open reading frames. Comparative genomic analyses will help to
identify helper traits.

<>

<1>Devi, S.H., Taylor, T.D., Avasthi, T.S., Kondo, S., Suzuki, Y., Megraud, F., Ahmed, N.
<2>Genome of Helicobacter pylori Strain 908.
<3>J. Bacteriol.
<4>192
<5>6488-6489
<6>2010
<7>Helicobacter pylori is a genetically diverse and coevolved pathogen inhabiting human gastric
niches and leading to a spectrum of gastric
diseases in susceptible populations. We describe the genome sequence of H.
pylori 908, which was originally isolated from an African patient living
in France who suffered with recrudescent duodenal ulcer disease. The
strain was found to be phylogenetically related to H. pylori J99, and its
comparative analysis revealed several specific genome features and novel
insertion-deletion and substitution events. The genome sequence revealed
several strain-specific deletions and/or gain of genes exclusively present
in HP908 compared with different sequenced genomes already available in
the public domain. Comparative and functional genomics of HP908 and its
subclones will be important in understanding genomic plasticity and the
capacity to colonize and persist in a changing host environment.

<>

<1>Devi, U., Khatri, I., Kumar, N., Kumar, L., Sharma, D., Subramanian, S., Saini, A.K.
<2>Draft Genome Sequence of a Plant Growth-Promoting Rhizobacterium, Serratia fonticola Strain AU-P3(3).
<3>Genome Announcements
<4>1
<5>e00946-13
<6>2013
<7>Plant growth-promoting rhizobacteria (PGPR), found in the rhizospheric region of  plants, not
only suppress plant disease, but also directly improve plant health
by improving the availability of nutrients and by providing phytostimulants.
Herein, we report the high-quality genome sequence of Serratia fonticola strain
AU-P3(3), a PGPR of the pea plant, which confers phosphate solubilization,
indole-3-acetic acid production, ammonia production, hydrogen cyanide (HCN)
production, and siderophore production and also confers activity against
Rhizoctonia species. The 5.02-Mb genome sequence contains genes related to plant
growth promotion and biocontrol activities.

<>

<1>Devi, U., Khatri, I., Kumar, N., Sharma, D., Subramanian, S., Saini, A.K.
<2>Draft Genome Sequence of Plant-Growth-Promoting Rhizobacterium Serratia fonticola Strain AU-AP2C, Isolated from the Pea Rhizosphere.
<3>Genome Announcements
<4>1
<5>e01022-13
<6>2013
<7>Plant health can be augmented by plant-growth-promoting rhizobacteria (PGPR) that confer
biofertilizer, phytostimulation, and biocontrol activities. Herein, we
provide the high-quality draft genome sequence of Serratia fonticola strain
AU-AP2C, a Gram-negative motile PGPR of the pea plant, conferring phosphate
solubilization, ammonia production, and antifungal activity against Fusarium sp.
The 4.9-Mb genome contains genes related to plant growth promotion and synthesis
of siderophores.

<>

<1>Devor, E.J.
<2>The relative efficiency of restriction enzymes:  an update.
<3>Am. J. Hum. Genet.
<4>42
<5>179-182
<6>1988
<7>The question of which restriction enzymes to use in screening newly cloned
probes for RFLPs continues to come up.  The importance of this question is
obvious when it is recognised that, as of this writing, more than 140
restriction endonucleases are listed as commerically available.  By way of
aiding the decision process, Wijsman (1984) produced a model with which the
efficiency of any restriction enzyme relative to that of any other might be
assessed.  The model involves a calculation of the relative efficiency (RE) of
the enzyme, which takes into account the recognition sequence of the enzyme,
the size of the DNA fragment being used to probe the digests, and the minimum
restriction-fragment size detectable in the usual 0.7%-1.25% agarose gels.  RE
is thus a unitless value by means of which any two or more enzymes might be
compared.  All other things being equal, the higher the RE value the more
efficient the enzyme is in detecting sequence variants.  In this letter this
model is applied to an expanded and updated list of restriction enzymes, with
the result that some apparently good screening candidates and a few poor
candidates are observed.

<>

<1>Dey, M.S.
<2>General buffer for restriction endonucleases.
<3>European Patent Office
<4>EP 0555797 A
<5>
<6>1992
<7>Restriction endonuclease buffer compositions which are suitable for use with essentially all
restriction endonucleases. The compositions comprise Tris-HCl, MgCl2, and at least one of KCl,
beta-mercaptoethanol and deoxyribonucleoside triphosphate. The buffer compositions provide
efficient restriction of DNA using a wide variety of restriction endonucleases, regardless of
the specific buffer requirements of the restriction endonuclease when using conventional
restriction buffers.

<>

<1>Dey, R., Pal, K.K., Sherathia, D., Dalsania, T., Savsani, K., Patel, I., Sukhadiya, B., Mandaliya, M., Thomas, M., Ghorai, S., Vanpariya, S., Rupapara, R., Rawal, P., Saxena, A.K.
<2>Draft Genome Sequence of Bacillus sp. Strain NSP2.1, a Nonhalophilic Bacterium Isolated from the Salt Marsh of the Great Rann of Kutch, India.
<3>Genome Announcements
<4>1
<5>e00909-13
<6>2013
<7>The 5.52-Mbp draft genome sequence of Bacillus sp. strain NSP2.1, a nonhalophilic bacterium
isolated from the salt marsh of the Great Rann of Kutch, India, is
reported here. An analysis of the genome of this organism will facilitate the
understanding of its survival in the salt marsh.

<>

<1>Dey, R., Pal, K.K., Sherathia, D., Dalsania, T., Savsani, K., Patel, I., Thomas, M., Ghorai, S., Vanpariya, S., Rupapara, R., Rawal, P., Sukhadiya, B., Mandaliya, M., Saxena, A.K.
<2>Draft Genome Sequence of Bacillus sp. Strain NSP9.1, a Moderately Halophilic Bacterium Isolated from the Salt Marsh of the Great Rann of Kutch, India.
<3>Genome Announcements
<4>1
<5>e00835-13
<6>2013
<7>We report the 4.52-Mbp draft genome sequence of Bacillus sp. strain NSP9.1, a moderately
halophilic bacterium isolated from the salt marsh of the Great Rann of
Kutch, India. Analysis of the genome of this organism will lead to a better
understanding of the genes and metabolic pathways involved in imparting
osmotolerance.

<>

<1>Dey, R., Pal, K.K., Sherathia, D., Sukhadiya, B., Dalsania, T., Patel, I., Savsani, K., Thomas, M., Vanpariya, S., Mandaliya, M., Rupapara, R., Rawal, P., Ghorai, S., Bhayani, S., Shah, A., Saxena, A.K.
<2>Insight into the First Draft Genome Sequence of the Genus Sediminibacillus, Sediminibacillus halophilus Strain NSP9.3.
<3>Genome Announcements
<4>2
<5>e01133-13
<6>2014
<7>We report the 3.98-Mbp first draft genome sequence of Sediminibacillus halophilus strain
NSP9.3, a moderate halophile isolated from a seasonal salt marsh of the
Great Rann of Kutch, India. Exploring the genome of this organism will facilitate
the understanding of the mechanism(s) of osmotolerance and survival in
differential osmolarity.

<>

<1>Dey, R., Pal, K.K., Sherathia, D., Vanpariya, S., Patel, I., Dalsania, T., Savsani, K., Sukhadiya, B., Mandaliya, M., Thomas, M., Ghorai, S., Rupapara, R., Rawal, P.
<2>Draft Genome Sequence of the Obligate Halophilic Bacillus sp. Strain NSP22.2, Isolated from a Seasonal Salt Marsh of the Great Rann of Kutch, India.
<3>Genome Announcements
<4>1
<5>e01104-13
<6>2013
<7>Here, we report the 4.0-Mbp draft genome of an obligate halophile, Bacillus sp. strain
NSP22.2, isolated from a seasonal salt marsh of the Great Rann of Kutch,
India. To understand the mechanism(s) of obligate halophilism and to isolate the
relevant gene(s), the genome of Bacillus sp. NSP22.2 was sequenced.

<>

<1>Dhakal, R., Seale, R., Brent, D., Hilton, C., Craven, H., Turner, M.S.
<2>Draft Genome Comparison of Representatives of the Three Dominant Genotype Groups of Dairy Bacillus licheniformis Strains.
<3>Appl. Environ. Microbiol.
<4>80
<5>3453-3462
<6>2014
<7>The spore-forming bacterium Bacillus licheniformis is a common contaminant of milk and milk
products. Strains of this species isolated from dairy products can be differentiated into
three major groups, namely, G, F1, and F2, using random amplification of polymorphic DNA
(RAPD) analysis; however, little is known about the genomic differences between these groups
and the identity of the fragments that make up their RAPD profiles. In this work we obtained
high-quality draft genomes of representative strains from each of the three RAPD groups
(designated strain G-1, strain F1-1, and strain F2-1) and compared them to each other and to
B. licheniformis ATCC 14580 and Bacillus subtilis 168. Whole-genome comparison and multilocus
sequence typing revealed that strain G-1 contains significant sequence variability and belongs
to a lineage distinct from the group F strains. Strain G-1 was found to contain genes coding
for a type I restriction modification system, urease production, and bacitracin synthesis, as
well as the 8-kbp plasmid pFL7, and these genes were not present in strains F1-1 and F2-1. In
agreement with this, all isolates of group G, but no group F isolates, were found to possess
urease activity and antimicrobial activity against Micrococcus. Identification of RAPD band
sequences revealed that differences in the RAPD profiles were due to differences in gene
lengths, 3' ends of predicted primer binding sites, or gene presence or absence. This work
provides a greater understanding of the phylogenetic and phenotypic differences observed
within the B. licheniformis species.

<>

<1>Dharmalingam, K., Goldberg, E.B.
<2>Restriction in vivo.  V. Induction of SOS functions in Escherichia coli by restricted T4 phage DNA and alleviation of restriction by SOS functions.
<3>Mol. Gen. Genet.
<4>178
<5>51-58
<6>1980
<7>Degradation products of restricted T4 DNA induced filamentation, mutagenesis,
and to a lesser extent, synthesis of recA protein in wild type cells but not in
recA, lexA or recBC mutants of Escherichia coli.  We conclude that the
structural damage to the DNA caused by restriction cleavage and exonuclease V
degradation can induce SOS functions.  Degradation of restricted
nonglucosylated T4 DNA by exonuclease V delayed cell division and induced
filament formation and mutagenesis in lexA+ but not in lexA- cells.  Delay of
cell division was also dependent upon recA and recBC functions.  Such
degradation of DNA also dramatically increased mutagenesis in tif- Sfi- cells
at 42.  The synthesis of recA protein continued in the restricting host after
infection by the nonglucosylated T4 phage, but enhanced synthesis is not
induced to the extent seen in SOS induced tif cells grown at 42.  We also found
that restriction of nonglucosylated T4 was alleviated in UV irradiated cells.
The UV induced alleviation of rgl and rK restriction depended upon post
irradiation protein synthesis and was not observed in recA, lexA or recBC
mutants.

<>

<1>Dharmalingam, K., Goldberg, E.B.
<2>Phage-coded protein prevents restriction of unmodified progeny T4 DNA.
<3>Nature
<4>260
<5>454-455
<6>1976
<7>None

<>

<1>Dharmalingam, K., Goldberg, E.B.
<2>Restriction in vivo.  IV. Effect of restriction of parental DNA on the expression of restriction alleviation systems in phage T4.
<3>Virology
<4>96
<5>404-411
<6>1979
<7>Under restricting conditions expression of early T4 genes is determined in part by the
location of the particular gene relative to the cleavage site.  Some genes are inactivated
directly by cleavage of the DNA.  Other genes are inactivated by secondary exonucleolytic
degradation of cleaved DNA.  A third class of genes continues to be expressed from the small
fragments which remain after the exonucleolytic degradation of the cleaved fragments.
Expression of phage genes coding for the inhibitors of the restriction endonuclease and
restriction exonuclease are prevented by endonucleolytic cleavage and exonucleolytic
degradation, respectively.

<>

<1>Dharmalingam, K., Goldberg, E.B.
<2>Mechanism localisation and control of restriction cleavage of phage T4 and lambda chromosomes in vivo.
<3>Nature
<4>260
<5>406-410
<6>1976
<7>The primary action of restriction endonuclease, cleaving infecting DNA, has
been demonstrated in vivo.  This primary cleavage is followed rapidly by
hydrolysis of the cleaved DNA at its newly exposed termini.  Infecting viruses
can inactivate cytoplasmic and membrane restriction endonucleases to prevent
cleavage of unmodified DNA replicas.

<>

<1>Dharmalingam, K., Revel, H.R., Goldberg, E.B.
<2>Physical mapping and cloning of bacteriophage T4 anti-restriction ednonuclease gene.
<3>J. Bacteriol.
<4>149
<5>694-699
<6>1982
<7>We have proposed that the ability of T4 to produce non-glucosylated progeny
after a single cycle of growth on a galU rglA rglB+ mutant of Escherichia coli
is due to the inhibition of the rglB+ function by a phage-coded,
anti-restriction endonuclease protein.  Based on this hypothesis, we screened
T4 deletion mutants for failure to give a burst in this host.  The absence of
an arn gene in phage mutants secondary infecting non-glucosylated phage from
rglB-controlled cleavage.  A functional arn gene was cloned on plasmid pBR325,
and the 0.8-kilobase insert DNA was shown to be homologous to the DNA missing
in the arn deletion phage.

<>

<1>Dherbecourt, J., Falentin, H., Canaan, S., Thierry, A.
<2>A genomic search approach to identify esterases in Propionibacterium freudenreichii involved in the formation of flavour in Emmental cheese.
<3>Microb. Cell Fact.
<4>7
<5>16
<6>2008
<7>ABSTRACT: BACKGROUND: Lipolysis is an important process of cheese ripening
that contributes to the formation of flavour. Propionibacterium
freudenreichii is the main agent of lipolysis in Emmental cheese; however,
the enzymes involved produced by this species have not yet been
identified. Lipolysis is performed by esterases (carboxylic ester
hydrolases, EC 3.1.1.-) which are able to hydrolyse acylglycerols bearing
short, medium and long chain fatty acids. The genome sequence of P.
freudenreichii type strain CIP103027T was recently obtained in our
laboratory.The aim of this study was to identify as exhaustively as
possible the potential esterases in P. freudenreichii that could be
involved in the hydrolysis of acylglycerols in Emmental cheese. The
proteins identified were produced in a soluble and active form by
heterologous expression in Escherichia coli for further study of their
activity and specificity of hydrolysed substrates. RESULTS: The approach
chosen was a genomic search approach that combined and compared four
methods based on automatic and manual searches of homology and motifs
among P. freudenreichii CIP103027T predicted proteins. Twenty-three
putative esterases were identified in this step. Then a selection step
permitted to focus the study on the 12 most probable esterases, according
to the presence of the GXSXG motif of the alpha/beta hydrolase fold
family. The 12 corresponding coding sequences were cloned in expression
vectors, containing soluble N-terminal fusion proteins. The best
conditions to express each protein in a soluble form were found thanks to
an expression screening, using an incomplete factorial experimental
design. Eleven out of the 12 proteins were expressed in a soluble form in
E. coli and six showed esterase activity on 1-naphthyl acetate and/or
propionate, as demonstrated by a zymographic method. CONCLUSION: We were
able to demonstrate that our genomic search approach was efficient to
identify esterases from the genome of a P. freudenreichii strain, more
exhaustively than classical approaches. This study highlights the interest
in using the automatic search of motifs, with the manual search of
homology to previously characterised enzymes as a complementary method.
Only further characterisations would permit the identification of the
esterases of P. freudenreichii involved in the lipolysis in Emmental
cheese.

<>

<1>Dhillon, B., Cavaletto, J.R., Wood, K.V., Goodwin, S.B.
<2>Accidental Amplification and Inactivation of a Methyltransferase Gene Eliminates Cytosine Methylation in Mycosphaerella graminicola.
<3>Genetics
<4>186
<5>67
<6>2010
<7>A de novo search for repetitive elements in the genome sequence of the wheat pathogen
Mycosphaerella graminicola identified a family of
repeats containing a DNA cytosine methyltransferase sequence (MgDNMT).
All 23 MgDNMT sequences identified carried signatures of repeat induced
point mutation (RIP). All copies were subtelomeric in location except
for one on chromosome 6. Synteny with M. fijiensis implied that the
nontelomeric copy on chromosome 6 served as a template for subsequent
amplifications. Southern analysis revealed that the MgDNMT sequence
also was amplified in 15 additional M. graminicola isolates from
various geographical regions. However, this amplification event was
specific to M. graminicola; a search for MgDNMT homologs identified
only a single, unmutated copy in the genomes of 11 other ascomycetes. A
genome-wide methylation assay revealed that M. graminicola lacks
cytosine methylation, as expected if its MgDNMT gene is inactivated.
Methylation was present in several other species tested, including the
closest known relatives of M. graminicola, species S1 and S2.
Therefore, the observed changes most likely occurred within the past
10,500 years since the divergence between M. graminicola and S1. Our
data indicate that the recent amplification of a single-copy MgDNMT
gene made it susceptible to RIP, resulting in complete loss of cytosine
methylation in M. graminicola.

<>

<1>Di Bonaventura, M.P., Desalle, R., Pop, M., Nagarajan, N., Figurski, D.H., Fine, D.H., Kaplan, J.B., Planet, P.J.
<2>Complete Genome Sequence of Aggregatibacter (Haemophilus) aphrophilus NJ8700.
<3>J. Bacteriol.
<4>191
<5>4693-4694
<6>2009
<7>We report the finished and annotated genome sequence of Aggregatibacter aphrophilus strain
NJ8700, a strain isolated from the oral flora of a healthy individual, and discuss
characteristics that may affect its dual roles in human health and disease. This strain has a
rough appearance, and its genome contains genes encoding a type VI secretion system and
several factors that may participate in host colonization.

<>

<1>Di Cagno, R., De Angelis, M., Cattonaro, F., Gobbetti, M.
<2>Draft Genome Sequence of Lactobacillus rossiae DSM 15814T.
<3>J. Bacteriol.
<4>194
<5>5460-5461
<6>2012
<7>The draft genome sequence of Lactobacillus rossiae DSM 15814(T) (CS1, ATCC BAA-88) was
determined by a whole-genome shotgun approach. Reads were assembled
to a 2.9-Mb draft version. RAST genome annotation evidenced 2,723 predicted
coding sequences. Many carbohydrate, amino acid, and amino acid derivative
subsystem features were found.

<>

<1>Di Gennaro, P., Zampolli, J., Presti, I., Cappelletti, M., D'Ursi, P., Orro, A., Mezzelani, A., Milanesi, L.
<2>Genome Sequence of Rhodococcus opacus Strain R7, a Biodegrader of Mono- and Polycyclic Aromatic Hydrocarbons.
<3>Genome Announcements
<4>2
<5>e00827-14
<6>2014
<7>Rhodococcus opacus strain R7 (CIP107348) degrades several mono- and polycyclic aromatic
hydrocarbons. Here, we present the high-quality draft genome sequence of
strain R7, consisting of 10,118,052 bp, with a G+C content of 67.0%, 9,602
protein-coding genes, and 62 RNAs genes.

<>

<1>Di Giaimo, R., Locascio, A., Aniello, F., Branno, M., del Gaudio, R., Potenza, N., Geraci, G.
<2>DNA (cytosine-5) methyltransferase turnover and cellular localization in developing Paracentrotus lividus sea urchin embryo.
<3>Gene
<4>272
<5>199-208
<6>2001
<7>The turnover and localization of the enzyme DNA (cytosine-5) methyltransferase (Dnmt1) were
studied during Paracentrotus lividus sea urchin embryo development using antibody preparations
against the NH(2) and COOH-terminal regions of the molecule. The antibodies reveal, by Western
blots and whole-mount analyses, that the enzyme is differentially required during embryonic
development. The changeover point is at blastula stage, where a proteolytic mechanism
hydrolyses the enzyme present in all embryonic cells by removing a peptide of about 45 kDa
from the amino terminal region of the 190 kDa enzyme initially synthesized on maternal
transcripts. The resulting 145 kDa enzyme shows modified catalytic properties, different
antibody reactivity and is rapidly destroyed in the few hours before gastrulation. At more
advanced stages of development the enzyme is newly synthesized but only in particular cell
types, among which are neurons. The data show that Dnmt1 is removed from embryonic cells
before gastrulation to be synthesized again at different levels in different cell types,
indicating that the concentration of Dnmt1 is critical for the various differentiated cells of
the developing sea urchin embryo.

<>

<1>Di Lallo, G., Evangelisti, M., Mancuso, F., Ferrante, P., Marcelletti, S., Tinari, A., Superti, F., Migliore, L., D'Addabbo, P., Frezza, D., Scortichini, M., Thaller, M.C.
<2>Isolation and partial characterization of bacteriophages infecting Pseudomonas syringae pv. actinidiae, causal agent of kiwifruit bacterial canker.
<3>J. Basic Microbiol.
<4>54
<5>1-12
<6>2014
<7>The phytopathogen Pseudomonas syringae pv. actinidiae (Psa) is the causal agent of bacterial
canker of kiwifruit. In the last years, it has caused severe economic losses to Actinidia spp.
cultivations, mainly in Italy and New Zealand. Conventional strategies adopted did not provide
adequate control of infection. Phage therapy may be a realistic and safe answer to the urgent
need for novel antibacterial agents aiming to control this bacterial pathogen. In this study,
we described the isolation and characterization of two bacteriophages able to specifically
infect Psa. phiPSA1, a member of the Siphoviridae family, is a temperate phage with a narrow
host range, a long latency, and a burst size of 178; phiPSA2 is a lytic phage of Podoviridae
family with a broader host range, a short latency, a burst size of 92 and a higher
bactericidal activity as determined by the TOD value. The genomic sequence of phiPSA1 has a
length of 51,090 bp and a low sequence homology with the other siphophages, whereas phiPSA2
has a length of 40 472 bp with a 98% homology with Pseudomonas putida bacteriophage gh-1. Of
the two phages examined, phiPSA2 may be considered as a candidate for phage therapy of
kiwifruit disease, while phiPSA1 seems specific toward the recent outbreak's isolates and
could be useful for Psa typing.

<>

<1>Di Pilato, V., Pollini, S., Rossolini, G.M.
<2>Characterization of plasmid pAX22, encoding VIM-1 metallo-beta-lactamase, reveals a new putative mechanism of In70 integron mobilization.
<3>J. Antimicrob. Chemother.
<4>69
<5>67-71
<6>2014
<7>Objectives: VIM-type enzymes are among the most widespread acquired metallo-b-lactamases among
Gramnegative pathogens. Integron In70 is a class 1 integron that has emerged as a successful
genetic support for
blaVIM-1 (one of themostprevalent blaVIM allelic variants) in
Gram-negativenon-fermenters,andis usually chromosome borne. The objective of this study was to
characterize plasmid pAX22 from Achromobacter xylosoxidans
AX22, which represents the only In70-harbouring plasmid known so far, to gather insights into
the mechanisms of evolution and dissemination of In70-like elements.
Methods: The complete sequence of pAX22was obtained by pyrosequencing and assembled with Roche
Newbler software. The draft sequence, completed using a PCR-based strategy, was annotated via
the BASys tool and compared with known sequences using BLAST algorithms.
Results: The backbone of pAX22 showed significant similarity with that of pNOR-2000, a
blaVIM-2-harbouring plasmid from Pseudomonas aeruginosa, and with the TnCP23 transposon. The
three elements differed from each other mainly by the class 1 integron cassette arrays and by
some integron-associated structures. In pAX22, In70was associated with a novel putative
transposon, Tn7017, composed of a defective Tn402-like transposon
carrying In70 and the ISPa17 insertion sequence.
Conclusion: Plasmid pAX22 belongs to a lineage of plasmids circulating among Gram-negative
non-fermenters. In70 was probably acquired by pAX22 by transposition of Tn7017, revealing a
novel putative mechanism of In70
mobilization. Our results highlight the potential role that ISPa17couldhave in mobilizing
defective Tn402-like transposons carrying class 1 integrons.

<>

<1>Dias, G.M., Thompson, C.C., Fishman, B., Naka, H., Haygood, M.G., Crosa, J.H., Thompson, F.L.
<2>Genome Sequence of the Marine Bacterium Vibrio campbellii DS40M4, Isolated from Open Ocean Water.
<3>J. Bacteriol.
<4>194
<5>904
<6>2012
<7>Vibrio sp. strain DS40M4 is a marine bacterium that was isolated from open ocean water. In
this work, using genomic taxonomy, we were able to
classify this bacterium as V. campbellii. Our genomic analysis revealed
that V. campbellii DS40M4 harbors genes related to iron transport,
virulence, and environmental fitness, such as those encoding anguibactin
and vanchrobactin biosynthesis proteins, type II, III, IV, and VI
secretion systems, and proteorhodopsin.

<>

<1>Diaz, L.A., Hardisson, C., Rodicio, M.R.
<2>Isolation and characterization of actinophages infecting Streptomyces species and their interaction with host restriction-modification systems.
<3>J. Gen. Microbiol.
<4>135
<5>1847-1856
<6>1989
<7>Nine different phages, PhiA1 to PhiA9, were isolated from soil samples on
Streptomyces antibioticus ATCC 11891, a strain which produces the macrolide
antibiotic oleandomycin.  Each phage displayed a different host-range which did
not extend beyond Streptomyces species.  Host-range was mainly limited by
adsorption specificity and host-controlled restriction-modification systems.
All the phages except PhiA3 and PhiA9 formed turbid plaques on S. antibioticus,
but did not lysogenize this host.  However, three of the phages (PhiA5, PhiA7
and PhiA8) were identified as temperate, since they were able to lysogenize
other Streptomyces strains.  All of the phages were morphologically similar and
belonged to group B of Bradley's classification.  They had polyhedral heads and
long, non-contractile tails.  PhiA5, PhiA6 and PhiA7 had a base plate at the
terminal end of the tail.  Analysis with restriction endonucleases indicated
that the nine phages contained double-stranded DNA.  Hybridization studies
between the phage genomes, together with results on genome structure, allowed
classification of the phages into five groups: (I) PhiA2, PhiA4 and PhiA9, (II)
PhiA3 and PhiA8, (III) PhiA7, (IV) PhiA5 and PhiA6, and (V) PhiA1.

<>

<1>Diaz-Quinonez, J.A., Hernandez-Monroy, I., Lopez-Martinez, I., Ortiz-Alcantara, J., Gonzalez-Duran, E., Ruiz-Matus, C., Kuri-Morales, P., Ramirez-Gonzalez, J.E.
<2>Genome Sequence of Vibrio cholerae Strain O1 Ogawa El Tor, Isolated in Mexico, 2013.
<3>Genome Announcements
<4>2
<5>e01123-14
<6>2014
<7>We present the draft genome sequence of Vibrio cholerae InDRE 3140 recovered in 2013 during a
cholera outbreak in Mexico. The genome showed the Vibrio 7th
pandemic islands VSP1 and VSP2, the pathogenic islands VPI-1 and VPI-2, the
integrative and conjugative element SXT/R391 (ICE-SXT), and both prophages CTXphi
and RS1phi.

<>

<1>Dick, G.J., Andersson, A.F., Baker, B.J., Simmons, S.L., Thomas, B.C., Yelton, A.P., Banfield, J.F.
<2>Community-wide analysis of microbial genome sequence signatures.
<3>Genome Biology
<4>10
<5>R85
<6>2009
<7>BACKGROUND: Analyses of DNA sequences from cultivated microorganisms have
revealed genome-wide, taxa-specific nucleotide compositional characteristics,
referred to as genome signatures. These signatures have far-reaching implications
for understanding genome evolution and potential application in classification of
metagenomic sequence fragments. However, little is known regarding the
distribution of genome signatures in natural microbial communities or the extent
to which environmental factors shape them. RESULTS: We analyzed metagenomic
sequence data from two acidophilic biofilm communities, including composite
genomes reconstructed for nine archaea, three bacteria, and numerous associated
viruses, as well as thousands of unassigned fragments from strain variants and
low-abundance organisms. Genome signatures, in the form of tetranucleotide
frequencies analyzed by emergent self-organizing maps, segregated sequences from
all known populations sharing < 50 to 60% average amino acid identity and
revealed previously unknown genomic clusters corresponding to low-abundance
organisms and a putative plasmid. Signatures were pervasive genome-wide. Clusters
were resolved because intra-genome differences resulting from translational
selection or protein adaptation to the intracellular (pH approximately 5) versus
extracellular (pH approximately 1) environment were small relative to
inter-genome differences. We found that these genome signatures stem from
multiple influences but are primarily manifested through codon composition, which
we propose is the result of genome-specific mutational biases. CONCLUSIONS: An
important conclusion is that shared environmental pressures and interactions
among coevolving organisms do not obscure genome signatures in acid mine drainage
communities. Thus, genome signatures can be used to assign sequence fragments to
populations, an essential prerequisite if metagenomics is to provide ecological
and biochemical insights into the functioning of microbial communities.

<>

<1>Dick, G.J., Podell, S., Johnson, H.A., Rivera-Espinoza, Y., Bernier-Latmani, R., McCarthy, J.K., Torpey, J.W., Clement, B.G., Gaasterland, T., Tebo, B.M.
<2>Genomic insights into Mn(II) oxidation by the marine alphaproteobacterium Aurantimonas sp. strain SI85-9A1.
<3>Appl. Environ. Microbiol.
<4>74
<5>2646-2658
<6>2008
<7>Microbial Mn(II) oxidation has important biogeochemical consequences in
marine, freshwater, and terrestrial environments, but many aspects of the
physiology and biochemistry of this process remain obscure. Here, we
report genomic insights into Mn(II) oxidation by the marine
alphaproteobacterium Aurantimonas sp. strain SI85-9A1, isolated from the
oxic/anoxic interface of a stratified fjord. The SI85-9A1 genome harbors
the genetic potential for metabolic versatility, with genes for
organoheterotrophy, methylotrophy, oxidation of sulfur and carbon
monoxide, the ability to grow over a wide range of O(2) concentrations
(including microaerobic conditions), and the complete Calvin cycle for
carbon fixation. Although no growth could be detected under autotrophic
conditions with Mn(II) as the sole electron donor, cultures of SI85-9A1
grown on glycerol are dramatically stimulated by addition of Mn(II),
suggesting an energetic benefit from Mn(II) oxidation. A putative Mn(II)
oxidase is encoded by duplicated multicopper oxidase genes that have a
complex evolutionary history including multiple gene duplication, loss,
and ancient horizontal transfer events. The Mn(II) oxidase was most
abundant in the extracellular fraction, where it cooccurs with a putative
hemolysin-type Ca(2+)-binding peroxidase. Regulatory elements governing
the cellular response to Fe and Mn concentration were identified, and 39
targets of these regulators were detected. The putative Mn(II) oxidase
genes were not among the predicted targets, indicating that regulation of
Mn(II) oxidation is controlled by other factors yet to be identified.
Overall, our results provide novel insights into the physiology and
biochemistry of Mn(II) oxidation and reveal a genome specialized for life
at the oxic/anoxic interface.

<>

<1>Dickerson, R.E., Drew, H.R.
<2>Structure of a B-DNA dodecamer  II.  Influence of base sequence on helix structure.
<3>J. Mol. Biol.
<4>149
<5>761-786
<6>1981
<7>Detailed examination of the structure of the B-DNA dodecamer
C-G-C-G-A-A-T-T-C-G-C-G, obtained by single-crystal X-ray analysis (Drew et
al., 1981), reveals that the local helix parameters, twist, tilt and roll, are
much more strongly influenced by base sequence than by crystal packing or any
other external forces.  The central EcoRI restriction endonuclease recognition
site, G-A-A-T-T-C, is a B helix with an average of 9.8 base-pairs per turn.  It
is flanked on either side by single base-pair steps having aspects of an A-like
helix character.  The dodecamer structure suggests several general principles,
whose validity must be tested by other B-DNA analyses.  (1) When an external
bending moment is applied to a B-DNA double helix, it bends smoothly, without
kinks or breaks, and with relatively little effect on local helix parameters.
(2) Purine-3', 5'-pyrimidine steps open their base planes towards the major
groove, pyrimidine-purine steps open toward the minor groove, and homopolymer
(Pur-Pur, Pyr-Pyr) steps resist rolling in either direction.  This behavior is
related to the preference of pyrimidines for more negative glycosyl torsion
angles.  (3) CpG steps have smaller helical twist angles than do GpC, as though
in compensation for their smaller intrinsic base overlap.  Data on A-T steps
are insufficient for generalization.  (4) G-C base-pairs have smaller propellor
twist than A-T, and this arises mainly from interstrand base overlap rather
than the presence of the third hydrogen bond.  (5) DNAase I cuts preferentially
at positions of high helical twist, perhaps because of increased exposure of
the backbone to attack.  The correlation of the digestion patterns in solution
and helical twist in the crystal argues for the essential identity of the helix
structure in the two environments.  (6) In the two places where the sequence
TpCpG occurs, the C slips from under T in order to stack more efficiently over
G.  At the paired bases of this CpG step, the G and C are tilted so the angle
between base planes is splayed out to the outside of the helix.  This TpC is
the most favored cutting site for DNAse I by a factor of 4-5 (Lomonossoff et
al., 1981).  (7) The EcoRI restriction endonuclease and methylase both appear
to prefer a cutting site of the type purine-purine-A-T-T-pyrimidine, involving
two adjacent homopolymer triplets, and this may be a consequence of the
relative stiffness of homopolymer base-stacking observed in the dodecamer.

<>

<1>Dickman, S.
<2>Lithuanian biochemist builds enzyme empire.
<3>Science
<4>257
<5>1473-1474
<6>1992
<7>Vilnius -If you like to browse through laboratory catalogs looking for the latest equipment
and reagents, you might have come across a surprising entry in the most recent offering from
New England Biolabs.  There, on page 46, you'll find a whole set of new restriction enzymes
-the enzymes that chop up DNA and are a vital part of every molecular biologist's toolbox.
The surprise: The enzymes are all labeled "Made in Lithuania".  Lithuania? How could a small
Baltic state, independent for less than a year, compete with hot shot Western biotech
companies in supplying enzymes to the United States? Ask Rich Roberts, the former Cold Spring
Harbor Laboratory molecular biologist who is now director of research for New England Biolabs
and he will answer in a word: "Janulaitis".  Vidas Janulaitis (pronounced Yanoo-LITEis), he
will tell you, is professor of biochemistry a the University of Vilnius, head of the Institute
of Applied Enzymology -and creator of one of the world's largest collections of restriction
enzymes, with more than 100 on offer.  He also appears to be the first successful
biotechnology entrepreneur to emerge from the former Soviet Union -and New England Biolabs'
competitors are well aware of his talents.  "Formidable", is how Jeremy Walker of Amersham
International describes Janulaitis' contribution to the number of new restriction enzymes
marketed each year.

<>

<1>Dicosimo, D.J., Sharpe, P.L.
<2>Chromosomal expression of foreign genes in the hsdM region of a methylotrophic microbial host cell.
<3>US Patent Office
<4>US 20070065903 A
<5>
<6>2007
<7>Provided is a method for stably expressing an introduced gene or genes in a methylotrophic
microorganism host wherein the gene(s) are integrated into the hsdM region of the chromosome.
This method provides stable, high-level expression of the integrated genes in which growth
rate of the host strain is not highly affected and a selection marker is not required.  The
use of this method for expressing carotenoid biosynthetic genes and resulting production of
astaxanthin is also described.

<>

<1>Didier, S., Lazzaroni, J.C., Portalier, R.
<2>Cell localization of EcoRI endonuclease in Escherichia coli K-12.
<3>Appl. Microbiol. Biotechnol.
<4>28
<5>468-470
<6>1988
<7>Cell compartmentation of EcoRI endonuclease was analyzed using either parental
or tolA excretory strains of Escherichia coli.  Cells were subjected to various
fractionation procedures such as osmotic shock or spheroplast formation.  Our
results showed that EcoRI activity was almost entirely recovered into
cytoplasmic fractions and consequently was not released into the extracellular
medium by a tolA mutant.  These results did not support previous reports
suggesting a periplasmic location for the EcoRI enzyme and did not allow to
develop a simple method for EcoRI purification from culture supernatants of
excretory mutants.

<>

<1>Didovyk, A.
<2>The structural basis of DNA recognition and base extrusion by a DNA cytosine-5 methyltransferase M.HaeIII.
<3>Ph.D. Thesis, Harvard University, USA
<4>
<5>
<6>2010
<7>The goal of this study is to elucidate the mechanism of sequence specific DNA recognition and
base extrusion by DNA cytosine-5 methyltransferase from Haemophilus aegyptius M.Haelll. We
have solved the crystal structure of the C71S mutant of M.Haelll in complex with the substrate
DNA at 2.4A resolution (InC below for brevity). For the first time an X-ray structure reveals
a fully intrahelical target cytosine
poised for extrusion by a cytosine-5 methyltransferase. The target cytosine
is destabilized, having lost most of its stacking interactions with both neighbouring bases
and making longer hydrogen bonds with the complementary guanine. In addition the protein
competes for Watson-Crick hydrogen bonding of the target base pair.  Both the protein and the
DNA conformations are remarkably different from those in the structure where the target
cytosine is extrahelical (ExC for brevity) [57]. In the ExC structure the cytosine 3 ' of the
target base is the one forming a base pair with the guanine of the target base pair, whereas
in the InC structure the bases within the
recognition sequence stay correctly paired. The conformation of the DNA backbone 3' to the
target cytosine changes significantly as well - it shifts further away from the protein. The
catalytic loop of M.Haelll (residues 71-89) is retracted away from the DNA as well. The
results suggest that M.Haelll actively participates in base flipping.  It destabilizes the
target base by altering both stacking and Watson-Crick hydrogen bonding - two fundamental
interactions that keep DNA bases intrahelical.  In order to elucidate the role of the
intercalating residue Ile-221 in base extrusion,
a glycine mutant was constructed. Its structure in complex with the specific DNA has been
determined using X-ray crystallography. This structure is essentially identical to the ExC,
suggesting that the extrahelical state, observed in ExC, can be achieved in the absence of the
DNA helix stabilization provided by Ile-221.

<>

<1>Didovyk, A., Verdine, G.L.
<2>Structural Origins of DNA Target Selection and Nucleobase Extrusion by a DNA Cytosine Methyltransferase.
<3>J. Biol. Chem.
<4>287
<5>40099-40105
<6>2012
<7>Epigenetic methylation of cytosine residues in DNA is an essential element of genome
maintenance and function in organisms ranging from
bacteria to humans. DNA 5-cytosine methyltransferase enzymes (DCMTases)
catalyze cytosine methylation via reaction intermediates in which the
DNA is drastically remodeled, with the target cytosine residue extruded
from the DNA helix and plunged into the active site pocket of the
enzyme. We have determined a crystal structure of M.HaeIII DCMTase in
complex with its DNA substrate at a previously unobserved state, prior
to extrusion of the target cytosine and frameshifting of the DNA
recognition sequence. The structure reveals that M.HaeIII selects the
target cytosine and destabilizes its base-pairing through a precise,
focused, and coordinated assault on the duplex DNA, which isolates the
target cytosine from its nearest neighbors and thereby facilitates its
extrusion from DNA.

<>

<1>Diekmann, S., McLaughlin, L.W.
<2>DNA curvature in native and modified EcoRI recognition sites and possible influence upon the endonuclease cleavage reaction.
<3>J. Mol. Biol.
<4>202
<5>823-834
<6>1988
<7>The ligation of a decadeoxynucleotide containing the EcoRI recognition site
forms a series of multimers which appear to be curved based on observed
anomalous gel migration in polyacrylamide gels.  The degree of DNA curvature
present in the recognition sequence, based upon the observed migration anomaly,
can be altered by modifications to the purine functional groups at the 2- and
6-positions.  Deletion of the guanine 2-amino group, occurring in the minor
groove of the B-DNA helix, is most effective in increasing the observed DNA
curvature.  Conversely, the displacement of an amino group from the major
groove to the minor groove eliminates curvature.  DNA curvature is also
modulated by the exocyclic group at the purine 6-position with decreasing
curvature observed when changing the amino group to a carbonyl or proton
substitute.

<>

<1>Diene, S.M., Merhej, V., Henry, M., El Filali, A., Roux, V., Robert, C., Azza, S., Gavory, F., Barbe, V., La Scola, B., Raoult, D., Rolain, J.M.
<2>The Rhizome of the Multidrug-Resistant Enterobacter aerogenes Genome Reveals How New 'Killer Bugs' Are Created because of a Sympatric Lifestyle.
<3>Mol. Biol. Evol.
<4>30
<5>369-383
<6>2013
<7>Here, we sequenced the 5,419,609 bp circular genome of an Enterobacter aerogenes
clinical isolate that killed a patient and was resistant to almost all current
antibiotics (except gentamicin) commonly used to treat Enterobacterial
infections, including colistin. Genomic and phylogenetic analyses explain the
discrepancies of this bacterium and show that its core genome originates from
another genus, Klebsiella. Atypical characteristics of this bacterium (i.e.,
motility, presence of ornithine decarboxylase, and lack of urease activity) are
attributed to genomic mosaicism, by acquisition of additional genes, such as the
complete 60,582 bp flagellar assembly operon acquired "en bloc" from the genus
Serratia. The genealogic tree of the 162,202 bp multidrug-resistant conjugative
plasmid shows that it is a chimera of transposons and integrative conjugative
elements from various bacterial origins, resembling a rhizome. Moreover, we
demonstrate biologically that a G53S mutation in the pmrA gene results in
colistin resistance. E. aerogenes has a large RNA population comprising 8 rRNA
operons and 87 cognate tRNAs that have the ability to translate transferred genes
that use different codons, as exemplified by the significantly different codon
usage between genes from the core genome and the "mobilome." On the basis of our
findings, the evolution of this bacterium to become a "killer bug" with new
genomic repertoires was from three criteria that are "opportunity, power, and
usage" to indicate a sympatric lifestyle: "opportunity" to meet other bacteria
and exchange foreign sequences since this bacteria was similar to sympatric
bacteria; "power" to integrate these foreign sequences such as the acquisition of
several mobile genetic elements (plasmids, integrative conjugative element,
prophages, transposons, flagellar assembly system, etc.) found in his genome; and
"usage" to have the ability to translate these sequences including those from
rare codons to serve as a translator of foreign languages.

<>

<1>Diep, A.L., Lang, J.M., Darling, A.E., Eisen, J.A., Coil, D.A.
<2>Draft Genome Sequence of Dietzia sp. Strain UCD-THP (Phylum Actinobacteria).
<3>Genome Announcements
<4>1
<5>e00197-13
<6>2013
<7>Here, we present the draft genome sequence of an actinobacterium, Dietzia sp. strain UCD-THP,
isolated from a residential toilet handle. The assembly contains
3,915,613 bp. The genome sequences of only two other Dietzia species have been
published, those of Dietzia alimentaria and Dietzia cinnamea.

<>

<1>Diep, B.A., Gill, S.R., Chang, R.F., Phan, T.H., Chen, J.H., Davidson, M.G., Lin, F., Lin, J., Carleton, H.A., Mongodin, E.F., Sensabaugh, G.F., Perdreau-Remington, F.
<2>Complete genome sequence of USA300, an epidemic clone of community-acquired meticillin-resistant Staphylococcus aureus.
<3>Lancet
<4>367
<5>731-739
<6>2006
<7>BACKGROUND: USA300, a clone of meticillin-resistant Staphylococcus aureus, is a major source
of community-acquired infections in the USA, Canada, and
Europe. Our aim was to sequence its genome and compare it with those of
other strains of S aureus to try to identify genes responsible for its
distinctive epidemiological and virulence properties. METHODS: We
ascertained the genome sequence of FPR3757, a multidrug resistant USA300
strain, by random shotgun sequencing, then compared it with the sequences
of ten other staphylococcal strains. FINDINGS: Compared with closely
related S aureus, we noted that almost all of the unique genes in USA300
clustered in novel allotypes of mobile genetic elements. Some of the
unique genes are involved in pathogenesis, including Panton-Valentine
leucocidin and molecular variants of enterotoxin Q and K. The most
striking feature of the USA300 genome is the horizontal acquisition of a
novel mobile genetic element that encodes an arginine deiminase pathway
and an oligopeptide permease system that could contribute to growth and
survival of USA300. We did not detect this element, termed arginine
catabolic mobile element (ACME), in other S aureus strains. We noted a
high prevalence of ACME in S epidermidis, suggesting not only that ACME
transfers into USA300 from S epidermidis, but also that this element
confers a selective advantage to this ubiquitous commensal of the human
skin. INTERPRETATION: USA300 has acquired mobile genetic elements that
encode resistance and virulence determinants that could enhance fitness
and pathogenicity.

<>

<1>Dikic, J., Menges, C., Clarke, S., Kokkinidis, M., Pingoud, A., Wende, W., Desbiolles, P.
<2>The rotation-coupled sliding of EcoRV.
<3>Nucleic Acids Res.
<4>40
<5>4064-4070
<6>2012
<7>It has been proposed that certain type II restriction enzymes (REs), such as EcoRV, track the
helical pitch of DNA as they diffuse along DNA, a so-called
rotation-coupled sliding. As of yet, there is no direct experimental observation
of this phenomenon, but mounting indirect evidence gained from single-molecule
imaging of RE-DNA complexes support the hypothesis. We address this issue by
conjugating fluorescent labels of varying size (organic dyes, proteins and
quantum dots) to EcoRV, and by fusing it to the engineered Rop protein scRM6.
Single-molecule imaging of these modified EcoRVs sliding along DNA provides us
with their linear diffusion constant (D(1)), revealing a significant size
dependency. To account for the dependence of D(1) on the size of the EcoRV label,
we have developed four theoretical models describing different types of motion
along DNA and find that our experimental results are best described by
rotation-coupled sliding of the protein. The similarity of EcoRV to other type II
REs and DNA binding proteins suggests that this type of motion could be widely
preserved in other biological contexts.

<>

<1>Dila, D., Raleigh, E.A.
<2>Genetic dissection of the methylcytosine-specific restriction system mcrB of Escherichia coli K-12.
<3>Gene
<4>74
<5>23-24
<6>1988
<7>Meeting Abstract

<>

<1>Dila, D., Sutherland, E., Moran, L., Slatko, B., Raleigh, E.A.
<2>Genetic and sequence organization of the mcrBC locus of Escherichia coli K-12.
<3>J. Bacteriol.
<4>172
<5>4888-4900
<6>1990
<7>The mcrB (rglB) locus of Escherichia coli K-12 mediates sequence-specific restriction of
cytosine-modified DNA.  Genetic and sequence analysis shows that the locus actually comprises
two genes, mcrB and mcrC.  We show here that in vivo, McrC modifies the specificity of McrB
restriction by expanding the range of modified sequences restricted.  That is, the sequences
sensitive to McrB+-dependent restriction can be divided into two sets:  some modified
sequences containing 5-methylcytosine are restricted by McrB+McrC+ cells.  The sequences
restricted only by McrB+C+ include T-even bacteriophage containing 5-hydroxymethylcytosine
(restriction of this phage is the RglB+ phenotype), some sequences containing
N4-methylcytosine, and some sequences containing 5-methylcytosine.  The sequence codes for two
polypeptides of 54 (McrB) and 42 (McrC) kilodaltons, whereas in vitro translation yields four
products, of ~29 and ~49 (McrB) and of ~38 and ~40 (McrC) kilodaltons.  The McrB polypeptide
sequence contains a potential GTP-binding motif, so this protein presumably binds the
nucleotide cofactor.  The deduced McrC polypeptide is somewhat basic and may bind to DNA,
consistent with its genetic activity as a modulator of the specificity of McrB.  At the
nucleotide sequence level, the G+C content of mcrBC is very low for E. coli, suggesting that
the genes may have been acquired recently during the evolution of the species.

<>

<1>Dinakaran, V., Shankar, M., Jayashree, S., Rathinavel, A., Gunasekaran, P., Rajendhran, J.
<2>Genome Sequence of Staphylococcus arlettae Strain CVD059, Isolated from the Blood of a Cardiovascular Disease Patient.
<3>J. Bacteriol.
<4>194
<5>6615-6616
<6>2012
<7>We have isolated a Staphylococcus arlettae strain, strain CVD059, from the blood  of a
rheumatic mitral stenosis patient. Here, we report the genome sequence and
potential virulence factors of this clinical isolate. The draft genome of S.
arlettae CVD059 is 2,565,675 bp long with a G+C content of 33.5%.

<>

<1>Ding, F., Chaillet, J.R.
<2>In vivo stabilization of the Dnmt1 (cytosine-5)-methyltransferase protein.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>14861-14866
<6>2002
<7>The Dnmt1o form of the Dnmt1 (cytosine-5)-methyltransferase enzyme is synthesized and stored
in the cytoplasm of the oocyte and is used after fertilization to maintain methylation
patterns on imprinted genes. After implantation of the blastocyst, Dnmt1o is replaced by the
Dnmt1 form, which has an additional 118 aa at its amino terminus. To investigate functional
differences between Dnmt1o and Dnmt1, mice were generated with a mutant allele, Dnmt1(V),
which synthesized Dnmt1o instead of Dnmt1 in all somatic cells. Homozygous Dnmt1(V) mice were
phenotypically normal, and had normal levels of genomic methylation, indicating that Dnmt1o
adopts the maintenance methyltransferase function of Dnmt1. Despite the apparent equivalence
of Dnmt1o and Dnmt1 maintenance methyltransferase function in somatic cells, the Dnmt1o
protein was found at high levels (with a corresponding high enzymatic activity) in Dnmt1(V)
mice. In heterozygous Dnmt1(V)/+ embryonic stem cells and early embryos, equal steady-state
levels of Dnmt1o and Dnmt1 proteins were produced from the Dnmt1(V) and the WT Dnmt1 alleles,
respectively. However, in older embryos and adults, the Dnmt1(V) allele produced five times
the steady-state level of protein of the WT Dnmt1 allele. The difference in Dnmt1o and Dnmt1
levels is due to a developmentally regulated mechanism that degrades the Dnmt1 protein. The
intrinsic stability of the Dnmt1o protein is the most likely reason for its use as a
maternal-effect protein; stable ooplasmic stores of Dnmt1o would be available to traffick into
the nuclei of the eight-cell stage embryo and maintain methylation patterns on alleles of
imprinted genes during the fourth embryonic S phase.

<>

<1>Ding, F., Patel, C., Ratnam, S., McCarrey, J.R., Chaillet, J.R.
<2>Conservation of Dnmt1o Cytosine methyltransferase in the marsupial Monodelphis domestica.
<3>Genesis
<4>36
<5>209-213
<6>2003
<7>Imprinted genes have been identified in both eutherian mammals and in marsupials. In eutherian
species, there is a conservation of the
imprinting process, both in terms of the genes imprinted and the
epigenetic inheritance mechanism. In the mouse, the inheritance of gametic
methylation patterns depends on an oocyte-derived isoform of the Dnmt1
(cytosine-5)-methyltransferase protein, Dnmt1o, which functions during
preimplantation development to maintain methylation patterns on imprinted
alleles. To determine if this component of genomic imprinting is also
found in marsupials, Dnmt1 isoforms were examined in somatic cells and
germ cells of the South American opossum Monodelphis domestica. There is a
Dnmt1o protein in Monodelphis oocytes that is synthesized, as in the
mouse, from a different transcript than the somatic Dnmt1 protein. Thus,
an essential component of imprinting in eutherian mammals is found in a
marsupial species, suggesting that marsupials and eutherian mammals
imprint their genes with the same methylation-dependent mechanism.

<>

<1>Ding, H., Moksa, M.M., Hirst, M., Beatty, J.T.
<2>Draft Genome Sequences of Six Rhodobacter capsulatus Strains, YW1, YW2, B6, Y262, R121, and DE442.
<3>Genome Announcements
<4>2
<5>e00050-14
<6>2014
<7>Rhodobacter capsulatus is a model organism for studying a novel type of horizontal gene
transfer mediated by a phage-like gene transfer agent (RcGTA).
Here we report the draft genome sequences of six R. capsulatus strains that
exhibit different RcGTA properties, including RcGTA overproducers, RcGTA
nonproducers, and/or RcGTA nonreceivers.

<>

<1>Ding, J., Dou, Y., Wang, Y., Chang, Y.
<2>Draft Genome Sequence of Vibrio fortis Dalian14 Isolated from Diseased Sea Urchin (Strongylocentrotus intermedius).
<3>Genome Announcements
<4>2
<5>e00409-14
<6>2014
<7>Here, we report the draft genome sequence of Vibrio fortis Dalian14 isolated from diseased sea
urchin (Strongylocentrotus intermedius) during disease outbreaks in
North China. The availability of this genome sequence will facilitate the study
of the mechanisms of pathogenicity and evolution of Vibrio species.

<>

<1>Ding, J., Pan, Y., Jiang, H., Cheng, J., Liu, T., Qin, N., Yang, Y., Cui, B., Chen, C., Liu, C., Mao, K., Zhu, B.
<2>Whole genome sequences of four Brucella strains.
<3>J. Bacteriol.
<4>193
<5>3674-3675
<6>2011
<7>Brucella melitensis and Brucella suis are intracellular pathogens to livestock and humans.
Here we report four genome sequences, the virulent strain B. melitensis M28-12 and vaccine
strains B. melitensis M5, M111 and B. suis S2 that show varied virulence and pathogenicity,
which will help to design more effective brucellosis vaccine.

<>

<1>Ding, J.Y., Chiang, P.W., Hong, M.J., Dyall-Smith, M., Tang, S.L.
<2>Complete Genome Sequence of the Extremely Halophilic Archaeon Haloarcula hispanica Strain N601.
<3>Genome Announcements
<4>2
<5>e00178-14
<6>2014
<7>Haloarcula hispanica has been widely used in haloarchaeal studies, particularly in the
isolation of haloviruses. The genome of strain N601, a laboratory
derivative of the type strain ATCC 33960, was sequenced. Several potentially
significant differences from the published sequence of the type strain (CGMCC
1.2049 = ATCC 33960) were observed.

<>

<1>Ding, R., Li, Y., Qian, C., Wu, X.
<2>Draft Genome Sequence of Paenibacillus elgii B69, a Strain with Broad Antimicrobial Activity.
<3>J. Bacteriol.
<4>193
<5>4537
<6>2011
<7>Here, we report the draft genome sequence of Paenibacillus elgii B69, which was isolated from
soil and with broad-spectrum antimicrobial
activity. As far as we know, the P. elgii genome is the biggest one among
Paenibacillus genus with genome sequence available. Multiple sets of genes
related to antibiotic biosynthetic pathways have been found in the genome.

<>

<1>Dingman, D.W.
<2>Presence of N6-methyladenine in GATC sequences of Bacillus popilliae and Bacillus lentimorbus KLN2.
<3>J. Bacteriol.
<4>172
<5>6156-6159
<6>1990
<7>Nine strains of Bacillus popilliae and Bacillus lentimorbus KLN2 contain N6-methyladenine in
GATC sequences, as determined by using the restriction enzymes MboI and DpnI. Among eight
other Bacillus species examined, all, except one strain of Bacillus brevis (ATCC 9999), lacked
adenine methylation in GATC. A methylase with Escherichia coli dcm site specificity was not
present in any of the Bacillus species studied.

<>

<1>Dinsmore, P.K., Klaenhammer, T.R.
<2>Bacteriophage resistance in Lactococcus.
<3>Mol. Biotechnol.
<4>4
<5>297-314
<6>1995
<7>Lactic acid bacteria are industrial micoorganisms used in many food fermentations.
Lactococcus species are susceptible to bacteriophage infections that may result in slowed or
failed fermentations.  A substantial amount of research has focused on characterizing natural
mechanisms by which bacterial cells defend themselves against phage.  Numerous natural phage
defense mechanisms have been identified and studied, and recent efforts have improved phage
resistance by using molecular techniques.  The study of how phages overcome these resistance
mechanisms is also an important objective.  New strategies to minimize the presence,
virulence, and evolution of phage are being developed and are likely to be applied
industrially.

<>

<1>Dix, T.I., Kieronska, D.H.
<2>Errors between sites in restriction site mapping.
<3>Comput. Appl. Biosci.
<4>4
<5>117-123
<6>1988
<7>Restriction site mapping programs construct maps by generating permutations of
fragments and checking for consistency.  Unfortunately many consistent maps
often are obtained within the experimental error bounds, even though there is
only one actual map.  A particularly efficient algorithm is presented that aims
to minimize error bounds between restriction sites.  The method is generalized
for linear and circular maps.  The time complexity is derived and execution
times are given for multiple enzymes and a range of error bounds.

<>

<1>Djao, O.D. et al.
<2>Complete genome sequence of Syntrophothermus lipocalidus type strain (TGB-C1).
<3>Standards in Genomic Sciences
<4>3
<5>268-275
<6>2010
<7>Syntrophothermus lipocalidus Sekiguchi et al. 2000 is the type species of the genus
Syntrophothermus. The species is of interest because of its strictly
anaerobic lifestyle, its participation in the primary step of the degradation of
organic maters, and for releasing products which serve as substrates for other
microorganisms. It also contributes significantly to maintain a regular pH in its
environment by removing the fatty acids through beta-oxidation. The strain is
able to metabolize isobutyrate and butyrate, which are the substrate and the
product of degradation of the substrate, respectively. This is the first complete
genome sequence of a member of the genus Syntrophothermus and the second in the
family Syntrophomonadaceae. Here we describe the features of this organism,
together with the complete genome sequence and annotation. The 2,405,559 bp long
genome with its 2,385 protein-coding and 55 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Djordjevic, G.M.
<2>Development of a triggered suicide system for bacteriophage defense.
<3>Diss. Abstr.
<4>58
<5>1095B
<6>1997
<7>A novel bacteriophage defense mechanism was developed for Lactococcus lactis where an
inducible phage promoter was used to activate bacterial suicide system after the infection.
The LlaIR+ restriction endonuclease was exploited as a lethal gene of a phage-inducible
suicide system.  When expressed from a constitutive promoter, the LlaIR+ endonuclease was
lethal across a wide range of Gram-positive bacteria, including L. lactis.  Lethality of the
LlaIR+ was exploited to develop several novel, positive selection cloning vectors for these
organisms.  A middle, phage-inducible promoter (Phi31P) from lytic lactococcal bacteriophage
Phi31 was cloned upstream of the LlaIR+ on the high-copy plasmid (pTRK414H).  When L. lactis
(pTRK414h) was infected with 10^7 pfu/ml phage in broth culture, at multiplicity of infection
(MOI) of 0.1, no lysis was observed and the culture developed normally.  The efficiency of
plaquing (EOP) for Phi31 on L. lactis (pTRK414H) was lowered to 10^-4.  Center of infection
assays revealed that 85% of the infected L. lactis (pTRK414H) cells did not release progeny
phage.  The burst size of Phi31 in L. lactis (pTRK414H) was 41, four-fold lower than in
control cells.  The Phi31P/LlaIR+ cassette also inhibited four Phi31-recombinant derivatives,
at levels at least ten-fold greater than Phi31.  However, mutant phages could be enriched that
were less sensitive to the Phi31P/LlaIR+-encoded restriction.  These phages were altered in
the strength and timing of Phi31P induction.  The efficiency of the Phi31P/LlaIR+-based
suicide system was improved by increasing promoter strength, providing a restriction enhancer
llaIC, and by combination with other abortive defenses, per31 and abiA.  When either per31 or
abiA were combined with llaIC and Phi31P/LlaIR+, the EOP was reduced to <10^ -10 and virulent
phages were eliminated from the infected population.  Broader application of bacterial suicide
systems depends on the availability of phage-specific promoters.  A rapid method for isolation
of these promoters, based on the "capping" activity of the vaccinia virus guanylytransferase,
was developed in this study and used successfully to identify a phage-specific promoter from
lytic lactococcal bacteriophage sk1.

<>

<1>Djordjevic, G.M., Klaenhammer, T.R.
<2>Positive selection, insertion cloning vectors for lactic acid bacteria based on a restriction endonuclease cassette.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>95
<5>527
<6>1995
<7>Lactococcus lactis contains numerous restriction and modification (R/M) systems of different
specificity.  A novel IIS type R/M system has been characterized from the Lactococcus lactis
conjugative plasmid pTR2030.  The LlaI operon is composed of six genes; the methylase gene
llaM is followed by three genes (llaI.1, llaI.2, and llaI.3), all of which are essential for
restriction activity.  We have successfully subcloned the llaI.1, llaI.2, and llaI.3 genes,
without llaIM, as a suicide cassette into the E. coli-lactococcal shuttle vectors pTRKL2 and
pBV5030.  A promoter (P6) from Lactobacillus acidophilus ATCC4356, which is functional in E.
coli, lactococci, and lactobacilli, was cloned upstream of the three gene cassette.
Restriction activity (R+) was evaluated in Escherichia coli and various lactic acid bacteria
(LAB).  The R+ cassette was not functional in E. coli, but was lethal to L. lactis,
Lactobacillus plantarum, Lactobacillus gasseri, Lactobacillus johnsonii, Enterococcus
faecalis, and Carnobacterium pisicola.  The R+ suicide vector has several unique restriction
cloning sites located within the R+ three gene cassette that can facilitate cloning, including
NdeI, StuI, NarI and EcoRV.  Random genomic fragments from Lb. johnsonii were cloned into the
NdeI site resulting in complete inactivation of R+ activity and providing unconditional
selection for recombinant plasmids in surviving transformants.  These positive selection
cloning vectors are the first for lactic acid bacteria that are based on a restriction
endonuclease cassette.  Functional activity of the llaI genes in various lactic acid bacteria
will also enable use of the R+ vector for positive screening of promoter and terminator
sequences in these genera.

<>

<1>Djordjevic, G.M., Klaenhammer, T.R.
<2>Positive selection, cloning vectors for gram-positive bacteria based on a restriction endonuclease cassette.
<3>Plasmid
<4>35
<5>37-45
<6>1996
<7>Lactococcus lactis contains numerous restriction and modification (R/M) systems of different
specificities.  A novel IIS type R/M system encoded by the LlaI operon has previously been
characterized from the L. lactis conjugative plasmid pTR2030.  The LlaI operon is composed of
six genes: First, a small regulatory gene IIaIC precedes the methylase gene llaIM.  The
following three genes, llaI.1, llaI.2, llaI.3, are all essential for restriction endonuclease
activity and are designated as the restriction cassette llaIR.  The fourth open reading frame
of unknown function follows the llaIR gene cassette.  We have successfully subcloned the three
llaIR genes, llaI.1, llaI.2, and llaI.3, without llaIM, as a suicide cassette into the three
shuttle vectors pTRKL2, pTRKH2, and pBV5030.  A promoter (P6) from Lactobacillus acidophilus
ATCC4356, which is functional in E. coli, lactococci, and lactobacilli was cloned upstream of
the three gene cassette.  Restriction activity was evaluated in Escherichia coli and several
gram-positive bacteria.  The llaIR restriction cassette was not functional in E. coli, but its
presence was lethal to L. lactis, Lactobacillus gasseri, Lactobacillus plantarum,
Lactobacillus johnsonii, Lactobacillus acidophilus, Carnobacterium pisicola, Enterococcus
faecalis, Bacillus subtilis, and Leuconostoc gelidum.  Several novel, positive selection
cloning vectors were developed that can exploit unique cloning sites within the llaIR
cassette.  Insertions in llaI.1 resulted in complete inactivation of restriction activity and
provided unconditional selection for recombinant plasmids in surviving transformants.  These
positive selection cloning vectors are the first for gram-positive bacteria that are based on
a restriction endonuclease cassette.  Functional activity of the llaIR genes in various
gram-positive bacteria would also enable use of these cloning vectors for positive selection
of promoters, terminators, and regulatory sequences across these genera.

<>

<1>Djordjevic, G.M., O'Sullivan, D.J., Walker, S.A., Conkling, M.A., Klaenhammer, T.R.
<2>A triggered-suicide system designed as a defense against bacteriophages.
<3>J. Bacteriol.
<4>179
<5>6741-6748
<6>1997
<7>A novel bacteriophage protection system for Lactococcus lactis based on a genetic trap, in
which a strictly phage-inducible promoter isolated from the lytic phage Phi31 is used to
activate a bacterial suicide system after infection, was developed.  The lethal gene of the
suicide system consists of the three-gene restriction cassette LlaIR+, which is lethal across
a wide range of gram-positive bacteria.  The phage-inducible trigger promoter (Phi31P) and the
LlaIR+ restriction cassette were cloned in Escherichia coli on a high-copy-number replicon to
generate pTRK414H.  Restriction activity was not apparent in E. coli or L. lactis prior to
phage infection.  In phage challenges of L. lactis (pTRK414H) with Phi31, the efficiency of
plaquing was lowered to a 10^-4 and accompanied by a fourfold reduction in burst size.
Center-of-infection assays revealed that only 15% of infected cells released progeny phage.
In addition to phage Phi31, the Phi31P/LlaIR+ suicide cassette also inhibited four
Phi31-derived recombinant phages at levels at least 10-fold greater than that of Phi31.  The
Phi31P/LlaIR+-based suicide system is a genetically engineered form of abortive infection that
traps and eliminates phages potentially evolving in fermentation environments by destroying
the phage genome and killing the propagation host.  This type of phage-triggered suicide
system could be designed for any bacterium-phage combination, given a universal lethal gene
and an inducible promoter which is triggered by the infecting bacteriophage.

<>

<1>Djordjevic, M.
<2>Modeling bacterial immune systems: Strategies for expression of toxic - but useful - molecules.
<3>Biosystems
<4>112
<5>139-144
<6>2013
<7>Protection of bacterial cells against virus infection requires expression of molecules that
are able to destroy the incoming foreign
DNA. However, these molecules can also be toxic for the host cell. In
both restriction-modification (R-M), and the recently discovered
CRISPR/Cas systems, the toxicity is (in part) avoided through rapid
transition of the expression of the toxic molecules from 'OFF' to 'ON'
state. In restriction-modification systems the rapid transition is
achieved through a large binding cooperativity, and low translation
rate of the control protein. On the other hand, CRISPR array expression
in CRISPR/Cas systems involves a mechanism where a small decrease of
unprocessed RNAs leads to a rapid increase of processed small RNAs.
Surprisingly, this rapid amplification crucially depends on fast
non-specific degradation of the unprocessed molecules by an
unidentified nuclease, rather than on large cooperativity in protein
binding. Furthermore, the major control elements that are responsible
for fast transition of R-M and CRISPR/Cas systems from 'OFF' to 'ON'
state, are also directly involved in increased stability of the steady
states of these systems. We here discuss mechanisms that allow rapid
transition of toxic molecules from the unproductive to the productive
state in R-M and CRISPR/Cas systems. The main purpose of this
discussion is to put relevant theoretical and experimental work in a
perspective that points to general similarities in otherwise
mechanistically very different bacterial immune systems.

<>

<1>Djukic, M., Becker, D., Poehlein, A., Voget, S., Daniel, R.
<2>Genome Sequence of Paenibacillus alvei DSM 29, a Secondary Invader during European Foulbrood Outbreaks.
<3>J. Bacteriol.
<4>194
<5>6365
<6>2012
<7>Paenibacillus alvei is known as a secondary invader during European foulbrood of  honeybees.
Here, we announce the 6.83-Mb draft genome sequence of P. alvei type
strain DSM 29. Putative genes encoding an antimicrobial peptide, a binary toxin,
a mosquitocidal toxin, alveolysin, and different polyketides and nonribosomal
peptides were identified.

<>

<1>Djukic, M., Poehlein, A., Thurmer, A., Daniel, R.
<2>Genome Sequence of Brevibacillus laterosporus LMG 15441, a Pathogen of Invertebrates.
<3>J. Bacteriol.
<4>193
<5>5535-5536
<6>2011
<7>Here we announce the genome sequence of the bacterium Brevibacillus laterosporus LMG 15441,
which is a pathogen of invertebrates. The genome
consists of one chromosome and two circular plasmids. Sequence analysis
revealed a large potential to produce polyketides, nonribosomal peptides,
and toxins.

<>

<1>Dmitrenko, O.A., Sidorenko, S.V., Zhukhovitsky, V.G., Terekhova, R.V., Karabak, V.I., Tarasevich, N.N., Vasilyeva, E.I., Prokhorov, V.Y.
<2>Comparative effectiveness of typing Staphylococcus aureus methicillin-resistant strains, isolated in Moscow hospitals, with the use of three collections of bacteriophages.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>1
<5>3-9
<6>2003
<7>The typing of S. aureus methicillin-resistant strains, isolated in different hospitals of
Moscow; was carried out with the use of three
collections of phages: the International Set of Phages; the set of phages
of the International Center of S. aureus phage typing in London (L); and
the experimental collection of phages of the Gamaleya Institute of
Epidemiology and Microbiology in Moscow (M). In this study made with the
use of both the phages of the International Diagnostic Set and phages L in
the standard typing dose of 1 TP about 6% of the cultures under study
proved to be sensitive. When the typing dose was increased to 100 TP the
phages of the international diagnostic set lyzed 75.5% of the cultures.
The typed strains were found to belong to phage types 77 (71.7%), 77/84/85
(19.6%) and 94/96 (6.5%). At a concentration of 100 TP phages L lyzed
83.7% of the cultures, but the dominating phage types could not be
determined due to a great variety of phage markers. In contrast to the two
preceding collections, the third phage collection M was composed in such a
way that in the study of the investigated culture the specificity of its
restriction modification was primarily evaluated and only then the
presence of antiphage immunity was determined. This latter collection was
used in the evaluation of 93.1% of the cultures. By the specificity of
their restriction specification system the majority of them were
classified with two new groups, heretofore not described. Only this
collection M made it possible to differentiate epidemic and sporadic
strains and to evaluate the epidemic situation in all 6 hospitals.

<>

<1>do Nascimento, N.C., Dos Santos, A.P., Chu, Y., Guimaraes, A.M., Pagliaro, A., Messick, J.B.
<2>Genome Sequence of Mycoplasma parvum (Formerly Eperythrozoon parvum), a Diminutive Hemoplasma of the Pig.
<3>Genome Announcements
<4>1
<5>e00986-13
<6>2013
<7>We report the complete genome sequence of Mycoplasma parvum strain Indiana. Its circular
chromosome is 564,395 bp, which is smaller than that of Mycoplasma
genitalium, which was previously considered the smallest member of the
Mollicutes. Comparative analyses of the genomes of M. parvum and Mycoplasma suis
will provide novel insights into the molecular basis of their virulence.

<>

<1>do Nascimento, N.C., Guimaraes, A.M., Santos, A.P., Sanmiguel, P.J., Messick, J.B.
<2>Complete Genome Sequence of Mycoplasma haemocanis Strain Illinois.
<3>J. Bacteriol.
<4>194
<5>1605-1606
<6>2012
<7>Mycoplasma haemocanis is a blood pathogen that may cause acute disease in immunosuppressed or
splenectomized dogs. The genome of the strain Illinois is a
single circular chromosome with 919,992 bp and a GC content of 35%. Analyses of
the M. haemocanis genome will provide insights into its biology and in vitro
cultivation requirements.

<>

<1>Do, J.W., Moon, C.H., Kim, H.J., Ko, M.S., Kim, S.B., Son, J.H., Kim, J.S., An, E.J., Kim, M.K., Lee, S.K., Han, M.S., Cha, S.J., Park, M.S., Park, M.A., Lee, J.S., Kim, Y.C., Choi, D.L., Kim, J.W., Park, J.W.
<2>Complete genomic DNA sequence of rock bream iridovirus.
<3>Virology
<4>325
<5>351-363
<6>2004
<7>Iridovirus is a causative agent of epizootics among cultured rock bream
(Oplegnathus fasciatus) in Korea. Here, we report the complete genomic
sequence of rock bream iridovirus (RBIV). The genome of RBIV was 112080 bp
long and contained at least 118 putative open reading frames (ORFs), and
its genome organization was similar to that of infectious spleen and
kidney necrosis virus (ISKNV). Of the RBIV's 118 ORFs, 85 ORFs showed
60-99% amino acid identity to those of ISKNV. Phylogenetic analysis of
major capsid protein (MCP), DNA repair protein RAD2, and DNA polymerase
type-B family indicated that RBIV is closely related to red sea bream
iridovirus (RSIV), Grouper sleepy disease iridovirus (GSDIV), Dwarf
gourami iridovirus (DGIV), and ISKNV. The genome sequence provides useful
information concerning the evolution and divergence of iridoviruses in
cultured fish.

<>

<1>Doberva, M., Sanchez-Ferandin, S., Ferandin, Y., Intertaglia, L., Croue, J., Suzuki, M., Lebaron, P., Lami, R.
<2>Genome Sequence of the Sponge-Associated Ruegeria halocynthiae Strain MOLA R1/13b, a Marine Roseobacter with Two Quorum-Sensing-Based Communication Systems.
<3>Genome Announcements
<4>2
<5>e00998-14
<6>2014
<7>Ruegeria halocynthiae MOLA R1/13b is an alphaproteobacterium isolated from the Mediterranean
sea sponge Crambe crambe. We report here the genome sequence and
its annotation, revealing the presence of quorum-sensing genes. This is the first
report of the full genome of a Ruegeria halocynthiae strain.

<>

<1>Doberva, M., Sanchez-Ferandin, S., Ferandin, Y., Intertaglia, L., Joux, F., Lebaron, P., Lami, R.
<2>Genome Sequence of Maribius sp. Strain MOLA 401, a Marine Roseobacter with a Quorum-Sensing Cell-Dependent Physiology.
<3>Genome Announcements
<4>2
<5>e00997-14
<6>2014
<7>Maribius sp. strain MOLA401 is an alphaproteobacterium isolated from a coral reef lagoon
located in New Caledonia, France. We report the genome sequence and its
annotation which, interestingly, reveals the presence of genes involved in quorum
sensing. This is the first report of a full genome within the genus Maribius.

<>

<1>Dobkin, C., Ferrando, C., Brown, W.T.
<2>PFGE of human DNA: 5-azacytidine improves restriction.
<3>Nucleic Acids Res.
<4>15
<5>3183
<6>1987
<7>Pulsed field gel electrophoresis of human DNA and the development of long-range restriction
maps are frequently hampered by the cytosine methylation that occurs at CG dinucleotides in
human DNA.  This methylation can interfere with restriction and change the size, number and
concentration of fragments detected in Southern blots.  We have found that these effects,
which vary at different loci and in different cell lines, can be partially overcome by growing
cells in 5-azacytidine prior to DNA isolation.  This treatment increases the number of enzymes
that can be used to map a locus; it can help distinguish between polymorphic methylation
patterns and polymorphic restriction patterns; and it can establish distinguishing
characteristics for particular restriction fragments.

<>

<1>Dobrindt, U., Blum-Oehler, G., Nagy, G., Schneider, G., Johann, A., Gottschalk, G., Hacker, J.
<2>Genetic structure and distribution of four pathogenicity islands (PAI I(536) to PAI IV(536)) of uropathogenic Escherichia coli strain 536.
<3>Infect. Immun.
<4>70
<5>6365-6372
<6>2002
<7>For the uropathogenic Escherichia coli strain 536 (O6:K15:H31), the DNA
sequences of three pathogenicity islands (PAIs) (PAI I(536) to PAI
III(536)) and their flanking regions (about 270 kb) were determined to
further characterize the virulence potential of this strain. PAI I(536) to
PAI III(536) exhibit features typical of PAIs, such as (i) association
with tRNA-encoding genes; (ii) G+C content differing from that of the host
genome; (iii) flanking repeat structures; (iv) a mosaic-like structure
comprising a multitude of functional, truncated, and nonfunctional
putative open reading frames (ORFs) with known or unknown functions; and
(v) the presence of many fragments of mobile genetic elements. PAI I(536)
to PAI III(536) range between 68 and 102 kb in size. Although these
islands contain several ORFs and known virulence determinants described
for PAIs of other extraintestinal pathogenic E. coli (ExPEC) isolates,
they also consist of as-yet-unidentified ORFs encoding putative virulence
factors. The genetic structure of PAI IV(536), which represents the core
element of the so-called high-pathogenicity island encoding a siderophore
system initially identified in pathogenic yersiniae, was further
characterized by sample sequencing. For the first time, multiple PAI
sequences (PAI I(536) to PAI IV(536)) in uropathogenic E. coli were
studied and their presence in several wild-type E. coli isolates was
extensively investigated. The results obtained suggest that these PAIs or
at least large fragments thereof are detectable in other pathogenic E.
coli isolates. These results support our view that the acquisition of
large DNA regions, such as PAIs, by horizontal gene transfer is an
important factor for the evolution of bacterial pathogens.

<>

<1>Dobritsa, A.P., Dobritsa, S.V.
<2>DNA protection with the DNA methylase M.BbvI from Bacillus brevis var. GB against cleavage by the restriction endonucleases PstI and PvuII.
<3>Gene
<4>10
<5>105-112
<6>1980
<7>BamHI fragments of the Bacillus brevis var. GB plasmid pAD1 have been cloned in Escherichia
coli HB101 using pBR322 plasmid as a vector.  The analysis of the recombinant plasmids showed
that additional PstI sites had appeared in cloned fragments of pAD1.  Methylation of the
recombinant plasmids in vitro by enzymes from B. brevis GB cells blocks cleavage at these
additional PstI sites of cloned pAD1 fragments and at the PstI site of pBR322.  Among DNA
methylases of B. brevis GB, the cytosine DNA methylase M.BbvI is the most likely agent
modifying the recognition sequences of PstI.  The methylase can modify cytosine residues in
PstI or PvuII sites if these recognition sequences are linked to G at 5'- or to C at
3'-termini.  In particular, in vitro methylation of the SV40 DNA by B. brevis GB methylases
protects one of the two PstI sites and two of the three PvuII sites.  The described effect of
the protection of the specific PstI and PvuII sites may be used for physical mapping of
genomes and DNA cloning.

<>

<1>Dobritsa, A.P., Mikhailov, A.A., Vanyushin, B.F.
<2>Methylation of phage 1P+f DNA of Bacillus brevis var. G-B.
<3>Biokhimiia
<4>40
<5>1269-1274
<6>1975
<7>The DNA of a virulent mutant of temperate phage 1P+f of Bacillus brevis var.
G-B belongs to the AT type (GC=34.5 mole %) and contains 5-methylcytosine (0.17
mole %) and N6-methyl-adenine (0.32 mole %) as minor bases.  The amount of
these bases in the phage DNA does not depend on the nature of the host (P- and
S variants of B. brevis var. G-B).  In contrast to the host DNA and
heterologous DNA of Pseudomonas aeruginosa the DNAs of phages 1P+f (P-) and
1P+f (S) do not accept methyl groups during in vitro methylation by enzymes
from B. brevis var. G-B cells.  Consequently, these phage DNAs are completely
methylated in vivo.  The nature of the methylation of phage DNA in cells of
different variants of B. brevis var. G-B is the same, i.e., dissociation of the
culture is not accompanied by a change in the specificity of the methylation of
the DNA.  In phage 1P+f DNA 5-methylcytosine is present in all of the
pyrimidine isopliths; the maximum amount of this base (~27%) is found in the
dipyrimidine clusters.  In the host DNA all of the 5-methylcytosine is located
only in mono- and dipyrimidine fragments in a ratio of 1:1.  This means that
the specificities of the methylation of the cytosine residues in the host and
phage DNAs are different.  This difference is apparently due to the
participation of some specific methylase, induced by phage 1P+f, in the
methylation of the phage DNA.

<>

<1>Dodd, A., Swanevelder, D., Featherston, J., Rumbold, K.
<2>Draft Genome Sequence of Streptomyces albulus Strain CCRC 11814, an {varepsilon}-Poly-L-Lysine-Producing Actinomycete.
<3>Genome Announcements
<4>1
<5>e00696-13
<6>2013
<7>Here, we report the draft genome sequence of Streptomyces albulus strain CCRC 11814, a
soil-dwelling, Gram-positive bacterium. S. albulus produces
epsilon-poly-l-lysine, which has diverse antimicrobial activity. The genome is
9.43 Mb in size, with a G+C content of 72.2%, and contains 9,177 protein-coding
sequences.

<>

<1>Dodge, A.G., Wackett, L.P., Sadowsky, M.J.
<2>Plasmid Localization and Organization of Melamine Degradation Genes in Rhodococcus sp. Strain Mel.
<3>Appl. Environ. Microbiol.
<4>78
<5>1397-1403
<6>2012
<7>Rhodococcus sp. strain Mel was isolated from soil by enrichment and grew in
minimal medium with melamine as the sole N source with a doubling time of 3.5 h.
Stoichiometry studies showed that all six nitrogen atoms of melamine were
assimilated. The genome was sequenced by Roche 454 pyrosequencing to 13x
coverage, and a 22.3-kb DNA region was found to contain a homolog to the melamine
deaminase gene trzA. Mutagenesis studies showed that the cyanuric acid hydrolase
and biuret hydrolase genes were clustered together on a different 17.9-kb contig.
Curing and gene transfer studies indicated that 4 of 6 genes required for the
complete degradation of melamine were located on an approximately 265-kb
self-transmissible linear plasmid (pMel2), but this plasmid was not required for
ammeline deamination. The Rhodococcus sp. strain Mel melamine metabolic pathway
genes were located in at least three noncontiguous regions of the genome, and the
plasmid-borne genes encoding enzymes for melamine metabolism were likely recently
acquired.

<>

<1>Doerfler, W.
<2>Patterns of DNA methylation-evolutionary vestiges of foreign DNA inactivation as a host defense mechanism.
<3>Biol. Chem. Hoppe Seyler
<4>372
<5>557-564
<6>1991
<7>The proposals in this review are based on experimental work on the integration
of foreign DNA in mammalian cells, on the establishment of specific de novo
patterns of DNA methylation, and on the inhibition of transcription by the
sequence-specific methylation of promoter sequences.  It is suggested that
eukaryotic cells have developed several mechanisms of defense against the
uptake, integration, and continued expression of foreign DNA.  In the course of
evolution and continuing at present, cells have been exposed to foreign DNA,
entire genomes or fragments of them.  A particularly problematic organ system
in that respect must be the digestive tract in higher organisms.  The defense
mechanisms are thought to be the folling:  (1) degradation and/or excretion of
foreign DNA; (ii) excision and loss of previously integrated DNA from the host
genome; (iii) targeted inactivation of foreign genes by sequence-specific
methylation.  Genes whose products could be advantageous to the transformed
cells can somehow be selectively excluded from this silencing mechanism.  In
part, the specificity of de novo methylation must reside in the DNA
methyltransferase systems of the host cell.  However, nucleotide sequence,
structure, and chromatin arrangement in the foreign DNA could also play an
important role.  Since defense processes must have been activated many times in
evolution, patterns of DNA methylation as they can be observed today, may
represent vestiges of evolution, i.e. the sum total of selective de novo
methylations, possibly demethylations, and mutations.  Could existing patterns
of DNA methylation be altered during embryogenesis?  One has also to consider
the possibility that the insertion and progressive methylation of foreign DNA
can lead to alterations in the methylation of flanking host cell DNA-sequences
abutting the site of integration.  It will be interesting to investigate to
what extent these changes can contribute to the oncogenic transformation of
cells, particularly after the insertion of foreign (viral) genomes in cells
transformed by oncogenic viruses.

<>

<1>Doggett, N.A., Stubben, C.J., Chertkov, O., Bruce, D.C., Detter, J.C., Johnson, S.L., Han, C.S.
<2>Complete Genome Sequence of Bacillus thuringiensis Serovar Israelensis Strain HD-789.
<3>Genome Announcements
<4>1
<5>e01023-13
<6>2013
<7>Bacillus thuringiensis is an important microbial insecticide for controlling agricultural
pests. We report the finished genome sequence of Bacillus
thuringiensis serovar israelensis strain HD-789, which contains genes encoding 7
parasporal crystals consisting of Cry4Aa3, Cry4Ba5 (2 genes), Cry10Aa3, Cry11Aa3,
Cry60Ba3, and Cry60Aa3, plus 3 Cyt toxin genes and 1 hemagglutinin gene.

<>

<1>Dogs, M. et al.
<2>Genome sequence of Phaeobacter inhibens type strain (T5(T)), a secondary metabolite producing representative of the marine Roseobacter clade, and  emendation of the species description of Phaeobacter inhibens.
<3>Standards in Genomic Sciences
<4>9
<5>334-350
<6>2013
<7>Strain T5(T) is the type strain of the species Phaeobacter inhibens Martens et al. 2006, a
secondary metabolite producing bacterium affiliated to the
Roseobacter clade. Strain T5(T) was isolated from a water sample taken at the
German Wadden Sea, southern North Sea. Here we describe the complete genome
sequence and annotation of this bacterium with a special focus on the secondary
metabolism and compare it with the genomes of the Phaeobacter inhibens strains
DSM 17395 and DSM 24588 (2.10), selected because of the close phylogenetic
relationship based on the 16S rRNA gene sequences of these three strains. The
genome of strain T5(T) comprises 4,130,897 bp with 3.923 protein-coding genes and
shows high similarities in genetic and genomic characteristics compared to P.
inhibens DSM 17395 and DSM 24588 (2.10). Besides the chromosome, strain T5(T)
possesses four plasmids, three of which show a high similarity to the plasmids of
the strains DSM 17395 and DSM 24588 (2.10). Analysis of the fourth plasmid
suggested horizontal gene transfer. Most of the genes on this plasmid are not
present in the strains DSM 17395 and DSM 24588 (2.10) including a nitrous oxide
reductase, which allows strain T5(T) a facultative anaerobic lifestyle. The G+C
content was calculated from the genome sequence and differs significantly from
the previously published value, thus warranting an emendation of the species
description.

<>

<1>Dogs, M. et al.
<2>Genome sequence of Phaeobacter daeponensis type strain (DSM 23529(T)), a facultatively anaerobic bacterium isolated from marine sediment, and emendation  of Phaeobacter daeponensis.
<3>Standards in Genomic Sciences
<4>9
<5>142-159
<6>2013
<7>TF-218(T) is the type strain of the species Phaeobacter daeponensis Yoon et al. 2007, a
facultatively anaerobic Phaeobacter species isolated from tidal flats.
Here we describe the draft genome sequence and annotation of this bacterium
together with previously unreported aspects of its phenotype. We analyzed the
genome for genes involved in secondary metabolite production and its anaerobic
lifestyle, which have also been described for its closest relative Phaeobacter
caeruleus. The 4,642,596 bp long genome of strain TF-218(T) contains 4,310
protein-coding genes and 78 RNA genes including four rRNA operons and consists of
five replicons: one chromosome and four extrachromosomal elements with sizes of
276 kb, 174 kb, 117 kb and 90 kb. Genome analysis showed that TF-218(T) possesses
all of the genes for indigoidine biosynthesis, and on specific media the strain
showed a blue pigmentation. We also found genes for dissimilatory nitrate
reduction, gene-transfer agents, NRPS/ PKS genes and signaling systems homologous
to the LuxR/I system.

<>

<1>Doherty, J.P., Lindeman, R., Trent, R.J., Graham, M.W., Woodcock, D.M.
<2>Escherichia coli host strains SURE and SRB fail to preserve a palindrome cloned in lambda phage: improved alternate host strains.
<3>Gene
<4>124
<5>29-35
<6>1993
<7>We have attempted to produce Escherichia coli strains with the optimal combination of host
mutations required for the construction of genomic libraries in lambda and cosmid vectors. For
lambda vectors, we defined this as a strain that combined high efficiency of phage plating
with optimal tolerance to DNA methylation and the ability to propagate recombinants containing
regions of potential secondary structure. To optimize this latter property, we have tested a
series of strains for the ability to propagate a lambda phage containing a palindromic
sequence. These included an mcr- derivative of a strain shown by Ishiura et al. [J. Bacteriol.
171 (1989)] 1068-1074] to allow optimal stability of inserts in cosmid clones. All the sbcC
strains allowed plaque formation of the palindrome-containing lambda phage. However, while the
palindrome-containing phage plated with reasonable efficiency of SURE (recB sbcC recJ umuC
uvrC) and SRB (sbcC recJ umuC uvrC), the majority of phage recovered from these strains no
longer required an sbcC host for subsequent plating. These two strains also gave poorer titres
with a low-yielding phage clone from the huma Prader-Willi chromosome region. Optimal phage
hosts appear to be those that are McrA (mcrBC-hse-mrr)combined with mutations in sbcC plus
recBC or recD and without mutations in additional recombination functions such as recJ or recJ
umuC uvrC.

<>

<1>Dohra, H., Suzuki, H., Suzuki, T., Tanaka, K., Fujishima, M.
<2>Draft Genome Sequence of Holospora undulata Strain HU1, a Micronucleus-Specific Symbiont of the Ciliate Paramecium caudatum.
<3>Genome Announcements
<4>1
<5>e00664-13
<6>2013
<7>Holospora undulata is a micronucleus-specific symbiont of the ciliate Paramecium  caudatum. We
report here the draft genome sequence of H. undulata strain HU1.
This genome information will contribute to the study of symbiosis between H.
undulata and the host P. caudatum.

<>

<1>Doi, A., Pack, S.P., Kodaki, T., Makino, K.
<2>Reinvestigation of the Molecular Influence of Hypoxanthine on the DNA Cleavage Efficiency of Restriction Endonucleases BglII, EcoRI and BamHI.
<3>J. Biochem. (Tokyo)
<4>146
<5>201-208
<6>2009
<7>Hypoxanthine (Hyp), a deaminated base of adenine (Ade), can be employed as a good probe
molecule to reveal the significance of the minor groove
of guanine (Gua) in biomolecular interactions because Hyp possesses a
similar structure to Gua lacking its 2-amino group. In this study, we
examined cleavage efficiencies of restriction endonuclease enzymes on
DNA substrates with Hyp in their recognition sequences. As a substrate
for BglII, EcoRI and BamHI, 24-mer DNA oligomer with Hyp (in place of
Gua) was prepared together with its complementary sequences with
cytosine (Cyt) or thymine (Thy) as the counter base. At 37 degrees C
incubation for 1h, BglII and EcoRI showed higher DNA cleavage
reactivity on Hyp-containing DNA substrates than on normal ones,
whereas BamHI showed lower values on Hyp-containing substrates. Such
high cleavage performance of BglII and EcoRI on Hyp-containing DNA
substrates is in contrast to the results obtained 20 years ago, in
which short DNA substrates (8- or 10-mer) and low reaction temperatures
(15-20 degrees C) were employed. These new results suggest that the
lack of the exocyclic 2-amino group of Gua could contribute to enhanced
recognition access of BglII and EcoRI to DNA substrates.

<>

<1>Doi, K., Mori, K., Martono, H., Nagayoshi, Y., Fujino, Y., Tashiro, K., Kuhara, S., Ohshima, T.
<2>Draft Genome Sequence of Geobacillus kaustophilus GBlys, a Lysogenic Strain with  Bacteriophage OH2.
<3>Genome Announcements
<4>1
<5>e00634-13
<6>2013
<7>Geobacillus kaustophilus strain GBlys was isolated along with the bacteriophage OH2, which
infects G. kaustophilus NBRC 102445(T). Here we present a draft
sequence of this strain's genome, which consists of 216 contigs for a total of
3,541,481 bp, 3,679 predicted coding sequences, and a G+C content of 52.1%.

<>

<1>Doi, K., Mori, K., Mutaguchi, Y., Tashiro, K., Fujino, Y., Ohmori, T., Kuhara, S., Ohshima, T.
<2>Draft Genome Sequence of D-Branched-Chain Amino Acid Producer Lactobacillus otakiensis JCM 15040T, Isolated from a Traditional Japanese Pickle.
<3>Genome Announcements
<4>1
<5>e00546-13
<6>2013
<7>Lactobacillus otakiensis strain JCM 15040(T) was isolated from an unsalted pickling solution
used in the production of sunki, a traditional Japanese pickle.
Here, we prepared a draft genome sequence for this strain consisting of 40
contigs containing a total of 2,347,132 bp, 2,310 predicted coding sequences, and
a G+C content of 42.4%.

<>

<1>Doi, K., Mori, K., Tashiro, K., Fujino, Y., Nagayoshi, Y., Hayashi, Y., Kuhara, S., Ohshima, T.
<2>Draft Genome Sequence of Pediococcus lolii NGRI 0510Q(T) Isolated from Ryegrass Silage.
<3>Genome Announcements
<4>1
<5>e00156-12
<6>2013
<7>Pediococcus lolii NGRI 0510Q(T) was isolated from ryegrass silage produced on Ishigaki Island,
Okinawa Prefecture, Japan. Here we present a draft genome
sequence for this strain, consisting of 103 contigs for a total of 2,047,078 bp,
2,154 predicted coding sequences, and a G+C content of 42.1%.

<>

<1>Doi, N., Kumadaki, S., Oishi, Y., Matsumura, N., Yanagawa, H.
<2>In vitro selection of restriction endonucleases by in vitro compartmentalization.
<3>Nucleic Acids Res.
<4>32
<5>e95
<6>2004
<7>Restriction endonucleases are widely used in laboratory applications from recombinant DNA
technology to diagnostics, but engineering of restriction
enzymes by structure-guided design and in vivo directed evolution is at an
early stage. Here, we report the use of an in vitro compartmentalization
system for completely in vitro selection of restriction enzymes.
Compartmentalization of a single gene in a rabbit reticulocyte in vitro
transcription/translation system serves to isolate individually
synthesized enzymes from each other. In each compartment, an active enzyme
cleaves only its own encoding gene, whereas genes encoding inactive
enzymes remain intact. Affinity selection of the cleaved DNA encoding
active restriction endonucleases was accomplished by the use of
streptavidin-immobilized beads and dUTP-biotin, which was efficiently
incorporated into the cohesive end of the cleaved DNA using a DNA
polymerase. We confirmed that genes encoding active restriction
endonuclease FokI could be selected from a randomized library. This method
overcomes the limitations of current in vivo technologies and should prove
useful for rapid screening and evolution of novel restriction enzymes from
diverse mutant libraries, as well as for studies of catalytic and
evolutionary mechanisms of restriction enzymes.

<>

<1>Doi, N., Yanagawa, H.
<2>In vitro selection of restriction enzyme and receptor ligands by microcapsulated cell free protein synthesis system.
<3>Baiotekunoroji Janaru
<4>5
<5>84-86
<6>2005
<7>
<>

<1>Doiron, K.M., Lavigne-Nicolas, J., Cupples, C.G.
<2>Effect of interaction between 5-azacytidine and DNA (cytosine-5) methyltransferase on C-to-G and C-to-T mutations in Escherichia coli.
<3>Mutat. Res.
<4>429
<5>37-44
<6>1999
<7>The purpose of this study was to determine the effect of the Dcm
cytosine methyltransferase on 5-azacytidine (5-azaC) mutagenesis in
Escherichia coli. We used a Lac reversion assay to measure C-to-G and C-
to-T mutations at a single, methylatable cytosine in the lacZ gene, in
the presence and absence of Dcm. C-to-G mutations are stimulated by 5-
azaC but are largely independent of Dcm. In contrast, C-to-T mutations
are not stimulated by 5-azaC in either wild type or dcm cells. However,
in cells which contain Dcm but are defective in very short patch
repair, the normally high frequency of spontaneous C-to-T mutations is
decreased by the analog in a dose-dependent manner.

<>

<1>Doiron, K.M.J., Viau, S., Koutroumanis, M., Cupples, C.G.
<2>Overexpression of vsr in Escherichia coli is mutagenic.
<3>J. Bacteriol.
<4>178
<5>4294-4296
<6>1996
<7>Overexpression of vsr in Escherichia coli stimulates transition and frameshift mutations.  The
pattern of mutations suggests that mutagenesis is due to saturation or inactivation of
dam-directed mismatch repair.

<>

<1>Dombroski, D.F., Morgan, A.R.
<2>Restriction nuclease digestions driven to completion by Escherichia coli RNA polymerase and T4 gene 32 protein.
<3>J. Biol. Chem.
<4>260
<5>415-417
<6>1985
<7>Restriction enzyme digestions of large scale DNA preparations often do not go
to completion.  This is due to product inhibition by the newly generated ends
of the digested DNA.  The addition of exogenous proteins that bind tightly to
the free ends of DNA or to single-stranded DNA will relieve this inhibition.
We show that a considerable savings on restriction nucleases can be attained by
the addition of RNA polymerase or T4 gene 32 protein in stoichiometric amounts
to the newly produced DNA ends.

<>

<1>Domingos, D.F., Dellagnezze, B.M., Greenfield, P., Reyes, L.R., Melo, I.S., Midgley, D.J., Oliveira, V.M.
<2>Draft Genome Sequence of the Biosurfactant-Producing Bacterium Gordonia amicalis  Strain CCMA-559, Isolated from Petroleum-Impacted Sediment.
<3>Genome Announcements
<4>1
<5>e00894-13
<6>2013
<7>Gordonia amicalis strain CCMA-559 was isolated from an oil-contaminated mangrove  swamp and
shown to produce biosurfactants. This strain is a strict aerobe that
readily degrades an array of carbon sources, including N-acetylglucosamine,
cellobiose, Tween 80, and 4-hydroxybenzoic acid, and, like other G. amicalis
strains, likely desulfurizes dibenzothiophene.

<>

<1>Domingos, D.F., Dellagnezze, B.M., Greenfield, P., Reyes, L.R., Melo, I.S., Midgley, D.J., Oliveira, V.M.
<2>Draft Genome Sequence of Bacillus pumilus CCMA-560, Isolated from an Oil-Contaminated Mangrove Swamp.
<3>Genome Announcements
<4>1
<5>e00707-13
<6>2013
<7>Bacillus pumilus strain CCMA-560 was isolated from an oil-contaminated mangrove swamp and was
shown to produce biosurfactants. The strain appears to be capable
of degrading some plant cell wall-related compounds, including hemicelluose and
pectin. Genes for biopolymer export and polysaccharide intercellular adhesin
synthesis were also annotated.

<>

<1>Dominguez, M.A. Jr., Thornton, K.C., Melendez, M.G., Dupureur, C.M.
<2>Differential effects of isomeric incorporation of fluorophenylalanines into PvuII endonuclease.
<3>Proteins
<4>45
<5>55-61
<6>2001
<7>Incorporation of fluorine into proteins has long served as a means of probing structure and
function, yet there are few studies that examine the impact of fluorine substitution,
particularly at locations distant from the active sites of enzymes. The flexibility of
isomeric fluorine incorporation at Phe is used to explore subtle substitution effects on
enzyme activity and conformation. The unnatural amino acids o-, m-, and p-fluorophenylalanines
were incorporated biosynthetically into the representative PvuII restriction endonuclease.
Interestingly, m-fluoro-Phe-PvuII endonuclease exhibits very similar conformational stability
to that of the native enzyme, but it exhibits a reproducible, 2-fold higher average specific
activity. Given the level of incorporation and the distribution of species, the species of
modified enzyme responsible for this increase in specific activity is most likely even faster.
Further, moving the fluorine atom from the meta- to the para-position of Phe results in a
4-fold decrease in specific activity and a decrease in conformational stability of 1.5
kcal/mol. Since none of the Phe residues in PvuII endonuclease lies near the DNA recognition
or catalytic sites, this differential behavior alludes to the impact of subtle changes in
enzyme conformation on endonuclease activity and suggests novel ways to influence catalytic
behavior.

<>

<1>Dominguez-Bello, M.G., Maldonado, A.L.
<2>Population dynamics and Restriction-Modification (RM) systems in Helicobacter pylori.
<3>Zoonoses Public Health
<4>54
<5>114
<6>2007
<7>H. pylori is highly recombinant, but RMs provide a barrier to recombination and preserve
species identity.  A strain with higher RM numbers is expected to have high barriers to
recombination.  As human hosts mix, H. pylori strains recombine.  In Latin America we have
evidence of an increasing dominance of European strains at the expense of Amerindian, and
African strains.  We hypothesize that European strains of H. pylori have higher number
methylases than Amerindian strains.  We compared the number of restriction sites (using
pDRAW32) in 3406 bp DNA sequences from H. pylori strains from different geographic phylotypes.
We also determined in vitro, the restriction profile of 16 RE to infer the number of active
methylases.  The number of possible restriction sites for 16 RE on multilocus sequences from
102 strains of diverse geographical origin did not vary among strain groups.  All strains had
cognate sequences for at least 13 of the 16 tested RE.  Average restriction sites were 5.2;
5.7; 6.1 and 6.3 for strains hpWAfrica, hpEurope, hspEAsia and hspAmerind respectively.  Only
Hpy8I, HpyCH4IV, HpyCH4V, had variable number of restriction sites by strain group.  MLST
sequences do not indicate a substantial variation in the number of RE cognate restriction
sites between strain phylotypes.  Preliminary results indicate however that the number of
active methylases is very variable among strains.  So far, based on restriction profiles to 17
RE, 2 Amerindian strains show 0 and 4 methylases and one European strain shows 5 methylases.
In conclusion, variations in RE activity in H. pylori populations do not seem to be due to
differences in the number of restriction sites in the DNA, but rather to the number of active
methylases.

<>

<1>Dominova, I.N., Kublanov, I.V., Podosokorskaya, O.A., Derbikova, K.S., Patrushev, M.V., Toshchakov, S.V.
<2>Complete Genomic Sequence of 'Thermofilum adornatus' Strain 1910bT, a Hyperthermophilic Anaerobic Organotrophic Crenarchaeon.
<3>Genome Announcements
<4>1
<5>e00726-13
<6>2013
<7>The complete genomic sequence of a novel hyperthermophilic crenarchaeon, strain 1910b(T), was
determined. The genome comprises a 1,750,259-bp circular chromosome
containing single copies of 3 rRNA genes, 43 tRNA genes, and 1,896 protein-coding
sequences. In silico genome-genome hybridization suggests the proposal of a novel
species, 'Thermofilum adornatus' strain 1910b(T).

<>

<1>Dominova, I.N., Sorokin, D.Y., Kublanov, I.V., Patrushev, M.V., Toshchakov, S.V.
<2>Complete Genome Sequence of Salinarchaeum sp. Strain HArcht-Bsk1T, Isolated from  Hypersaline Lake Baskunchak, Russia.
<3>Genome Announcements
<4>1
<5>e00505-13
<6>2013
<7>The complete genome sequence of a novel halophilic archaeon, Salinarchaeum sp. strain
HArcht-Bsk1(T), was determined using next-generation sequencing. The
genome comprises a 3,255,260-bp circular chromosome with a G+C content of 66.7%.
Automatic annotation of the genome revealed a single rRNA operon, 45 tRNAs, and
3,013 protein-coding gene sequences.

<>

<1>Domotor, D., Becsagh, P., Rakhely, G., Schneider, G., Kovacs, T.
<2>Complete Genomic Sequence of Erwinia amylovora Phage PhiEaH2.
<3>J. Virol.
<4>86
<5>10899
<6>2012
<7>Erwinia amylovora is the causative agent of fire blight, a serious disease of
some Rosaceae plants. The newly isolated bacteriophage PhiEaH2 is able to lyse E.
amylovora in the laboratory and has reduced the occurrence of fire blight cases
in field experiments. This study presents the sequenced complete genome and
analysis of phage PhiEaH2.

<>

<1>Donahue, J.P., Israel, D.A., Peek, R.M., Blaser, M.J., Miller, G.G.
<2>Overcoming the restriction barrier to plasmid transformation of Helicobacter pylori.
<3>Mol. Microbiol.
<4>37
<5>1066-1074
<6>2000
<7>Helicobacter pylori strains demonstrate substantial variability in the efficiency of
transformation by plasmids from Escherichia coli, and many strains are completely resistant to
transformation. Among the barriers to transformation are numerous strain-specific
restriction-modification systems in H. pylori. We have developed a method to protect plasmid
DNA from restriction by in vitro site-specific methylation using cell-free extracts of H.
pylori before transformation. In two cases, plasmid DNA treated with cell-free extracts in
vitro acquired the restriction pattern characteristic of genomic DNA from the source strain.
Among three strains examined in detail, the transformation frequency by treated plasmid
shuttle and suicide vectors was significantly increased compared with mock-treated plasmid
DNA. The results indicate that the restriction barrier in H. pylori can be largely overcome by
specific DNA methylation in vitro. The approach described should significantly enhance the
ability to manipulate gene function in H. pylori and other organisms that have substantial
restriction barriers to transformation.

<>

<1>Donahue, J.P., Israel, D.A., Torres, V.J., Necheva, A.S., Miller, G.G.
<2>Inactivation of a Helicobacter pylori DNA methyltransferase alters dnaK operon expression following host-cell adherence.
<3>FEMS Microbiol. Lett.
<4>208
<5>295-301
<6>2002
<7>The Helicobacter pylori hpyIM gene encodes a type II DNA methyltransferase that is highly
conserved among strains. To investigate the potential role of M.HpyI methyltransferase
activity in controlling gene expression in H. pylori, we analyzed gene transcription profiles
in wild-type strain J166 and an isogenic hpyIM mutant strain using gene arrays. This analysis
showed that the expression of a majority of genes was unaffected by hpyIM mutation, especially
in exponential phase cultures. However, in stationary phase cultures and in cells adherent to
AGS gastric epithelial cells in vitro, loss of hpyIM function altered the expression of the
stress-responsive dnaK operon. Complementation of the hpyIM mutation using a shuttle plasmid
encoding a wild-type copy of the gene re-established the wild-type pattern of dnaK operon
expression. These data suggested that hpyIM, encoding a DNA methyltransferase, may have a role
in H. pylori physiology that supersedes its original function in a type II
restriction-modification system.

<>

<1>Donahue, J.P., Peek, R.M. Jr.
<2>Restriction and modification systems.
<3>Helicobacter pylori: Physiology and Genetics, ASM Press, Mobley, H.L.T., Mendz, G.L., Hazell, S.L., Washington, DC
<4>
<5>269-276
<6>2001
<7>Prokaryotic restriction-modification (R-M) systems were first recognized in Escherichia coli
nearly 50 years ago and are now known to be ubiquitous among bacterial species.  In general,
R-M systems consist of two distinct enzymatic activities: first, a restriction endonuclease
that cleaves DNA at a specific recognition sequence, and second, a DNA methyltransferase that
methylates DNA at the same site and thus prevents cleavage by the cognate restriction enzyme.
The genomic sequences of Helicobacter pylori strains J99 and 26695 have revealed that this
bacterium contains an abundance of restriction and modification genes, some of which have been
subsequently shown to function as authentic R-M systems or as partial systems, composed of the
DNA methyltransferase component alone.  Interestingly, R-M genes comprise a significant
percentage of H. pylori strain-specific genes and are more prevalent in H. pylori than in
other bacterial species whose genomes have been fully sequenced.  R-M systems in H. pylori
have been identified on the basis of sequence similarity to known restriction enzymes and
methyltransferases, genetic organization, and specific enzyme isolation and characterization.
This chapter summarizes the current state of knowledge regarding the structure and function of
the large number of putative R-M genes and systems that are now recognized to be present in H.
pylori.

<>

<1>Donati, C. et al.
<2>Structure and dynamics of the pan-genome of Streptococcus pneumoniae and closely related species.
<3>Genome Biology
<4>11
<5>R107
<6>2010
<7>BACKGROUND: Streptococcus pneumoniae is one of the most important causes of
microbial diseases in humans. The genomes of 44 diverse strains of S. pneumoniae
were analyzed and compared with strains of non-pathogenic streptococci of the
Mitis group. RESULTS: Despite evidence of extensive recombination, the S.
pneumoniae phylogenetic tree revealed six major lineages. With the exception of
serotype 1, the tree correlated poorly with capsular serotype, geographical site
of isolation and disease outcome. The distribution of dispensable genes--genes
present in more than one strain but not in all strains--was consistent with
phylogeny, although horizontal gene transfer events attenuated this correlation
in the case of ancient lineages. Homologous recombination, involving short
stretches of DNA, was the dominant evolutionary process of the core genome of S.
pneumoniae. Genetic exchange occurred both within and across the borders of the
species, and S. mitis was the main reservoir of genetic diversity of S.
pneumoniae. The pan-genome size of S. pneumoniae increased logarithmically with
the number of strains and linearly with the number of polymorphic sites of the
sampled genomes, suggesting that acquired genes accumulate proportionately to the
age of clones. Most genes associated with pathogenicity were shared by all S.
pneumoniae strains, but were also present in S. mitis, S. oralis and S. infantis,
indicating that these genes are not sufficient to determine virulence.
CONCLUSIONS: Genetic exchange with related species sharing the same ecological
niche is the main mechanism of evolution of S. pneumoniae. The open pan-genome
guarantees the species a quick and economical response to diverse environments.

<>

<1>Dong, A., Yoder, J.A., Zhang, X., Zhou, L., Bestor, T.H., Cheng, X.
<2>Structure of human DNMT2, an enigmatic DNA methyltransferase homolog that displays denaturant-resistant binding to DNA.
<3>Nucleic Acids Res.
<4>29
<5>439-448
<6>2001
<7>DNMT2 is a human protein that displays strong sequence similarities to DNA
(cytosine-5)-methyltransferases (m(5)C MTases) of both prokaryotes and eukaryotes. DNMT2
contains all 10 sequence motifs that are conserved among m(5)C MTases, including the consensus
S:-adenosyl-L-methionine-binding motifs and the active site ProCys dipeptide. DNMT2 has close
homologs in plants, insects and Schizosaccharomyces pombe, but no related sequence can be
found in the genomes of  Saccharomyces cerevisiae or Caenorhabditis elegans.  The crystal
structure of a deletion mutant of DNMT2 complexed with S-adenosyl-L-homocysteine (AdoHcy) has
been determined at 1.8 A resolution. The structure of the large domain that contains the
sequence motifs involved in catalysis is remarkably similar to that of M.HhaI, a confirmed
bacterial m(5)C MTase, and the smaller target recognition domains of DNMT2 and M.HhaI are also
closely related in overall structure. The small domain of DNMT2 contains three short helices
that are not present in M.HhaI. DNMT2 binds AdoHcy in the same conformation as confirmed m(5)C
MTases and, while DNMT2 shares all sequence and structural features with m(5)C MTases, it has
failed to demonstrate detectable transmethylase activity. We show here that homologs of DNMT2,
which are present in some organisms that are not known to methylate their genomes, contain a
specific target-recognizing sequence motif including an invariant CysPheThr tripeptide. DNMT2
binds DNA to form a denaturant-resistant complex in vitro. While the biological function of
DNMT2 is not yet known, the strong binding to DNA suggests that DNMT2 may mark specific
sequences in the genome by binding to DNA through the specific target-recognizing motif.

<>

<1>Dong, A.P., Zhou, L., Zhang, X., Stickel, S., Roberts, R.J., Cheng, X.D.
<2>Structure of the Q237W mutant of HhaI DNA methyltransferase: an insight into protein-protein interactions.
<3>Biol. Chem.
<4>385
<5>373-379
<6>2004
<7>We have determined the structure of a mutant (Q237W) of HhaI DNA methyltransferase, complexed
with the methyl-donor product AdoHcy. The
Q237W mutant proteins were crystallized in the monoclinic space group
C2 with two molecules in the crystallographic asymmetric unit.
Protein-protein interface calculations in the crystal lattices suggest
that the dimer interface has the specific characteristics for homodimer
protein-protein interactions, while the two active sites are spatially
independent on the outer surface of the dimer. The solution behavior
suggests the formation of HhaI dimers as well. The same HhaI dimer
interface is also observed in the previously characterized binary
(M.HhaI-AdoMet) and ternary (M.HhaI-DNA-AdoHcy) complex structures,
crystallized in different space groups. The dimer is characterized
either by a noncrystallographic twofold symmetry or a crystallographic
symmetry. The dimer interface involves three segments: the
amino-terminal residues 2-8, the carboxy-terminal residues 313-327, and
the linker (amino acids 179-184) between the two functional domains the
catalytic methylation domain and the DNA target recognition domain.
Both the amino and carboxyterminal segments are part of the methylation
domain. We also examined protein-protein interactions of other
structurally characterized DNA MTases, which are often found as a
2-fold related dimer with the largest dimer interface area for the
group-beta MTases. A possible evolutionary link between the Type I and
Type II restriction-modification systems is discussed.

<>

<1>Dong, C., Bai, X., Lai, Q., Xie, Y., Chen, X., Shao, Z.
<2>Draft Genome Sequence of Sphingobium sp. Strain C100, a Polycyclic Aromatic Hydrocarbon-Degrading Bacterium from the Deep-Sea Sediment of the Arctic Ocean.
<3>Genome Announcements
<4>2
<5>e01210-13
<6>2014
<7>Sphingobium sp. strain C100 was isolated from a polycyclic aromatic hydrocarbon
(PAH)-degrading consortium from the deep-sea sediment of the Arctic Ocean. It can
degrade two- to four-ring PAHs at 25 degrees C. Here we present the draft genome
sequence of this strain, which is 4,776,810 bp with a G+C content of 63.9%.

<>

<1>Dong, C., Bai, X., Lai, Q., Xie, Y., Chen, X., Shao, Z.
<2>Draft Genome Sequence of Marinomonas sp. Strain D104, a Polycyclic Aromatic Hydrocarbon-Degrading Bacterium from the Deep-Sea Sediment of the Arctic Ocean.
<3>Genome Announcements
<4>2
<5>e01211-13
<6>2014
<7>Marinomonas sp. strain D104 was isolated from a polycyclic aromatic hydrocarbon-degrading
consortium enriched from deep-sea sediment from the Arctic
Ocean. The draft genome sequence of D104 (approximately 3.83 Mbp) contains 62
contigs and 3,576 protein-encoding genes, with a G+C content of 44.8%.

<>

<1>Dong, C., Chen, X., Xie, Y., Lai, Q., Shao, Z.
<2>Complete genome sequence of Thalassolituus oleivorans R6-15, an obligate hydrocarbonoclastic marine bacterium from the Arctic Ocean.
<3>Standards in Genomic Sciences
<4>9
<5>893-901
<6>2014
<7>Strain R6-15 belongs to the genus Thalassolituus, in the family Oceanospirillaceae of
Gammaproteobacteria. Representatives of this genus are
known to be the obligate hydrocarbonoclastic marine bacteria. Thalassolituus
oleivorans R6-15 is of special interest due to its dominance in the crude
oil-degrading consortia enriched from the surface seawater of the Arctic Ocean.
Here we describe the complete genome sequence and annotation of this strain,
together with its phenotypic characteristics. The genome with size of 3,764,053
bp comprises one chromosome without any plasmids, and contains 3,372
protein-coding and 61 RNA genes, including 12 rRNA genes.

<>

<1>Dong, H., Zhang, Y., Li, Y.
<2>Method for inactivation of restriction modification system in butanol-producing Clostridium mutant.
<3>Chinese Patent Office
<4>CN 102071211 A
<5>
<6>2011
<7>
<>

<1>Dong, H.J. et al.
<2>Engineering Clostridium Strain to Accept Unmethylated DNA.
<3>PLoS ONE
<4>5
<5>e9038
<6>2010
<7>It is difficult to genetically manipulate the medically and biotechnologically important genus
Clostridium due to the existence of
the restriction and modification (RM) systems. We identified and
engineered the RM system of a model clostridial species, C.
acetobutylicum, with the aim to allow the host to accept the
unmethylated DNA efficiently. A gene CAC1502 putatively encoding the
type II restriction endonuclease Cac8241 was identified from the genome
of C. acetobutylicum DSM1731, and disrupted using the ClosTron system
based on group II intron insertion. The resulting strain SMB009 lost
the type II restriction endonuclease activity, and can be transformed
with unmethylated DNA as efficiently as with methylated DNA. The
strategy reported here makes it easy to genetically modify the
clostridial species using unmethylated DNA, which will help to advance
the understanding of the clostridial physiology from the molecular
level.

<>

<1>Dong, Q., Cao, X., Zou, G., Zhu, R.
<2>A simple and practical quantitative method for measuring the activity of restriction endonucleases.
<3>Yichuan
<4>9
<5>34-36
<6>1987
<7>None

<>

<1>Dong, Q., Zou, G., Cao, X., Zhu, R.
<2>Chemical modification and inhibition kinetics of restriction endonuclease Bsp63I.
<3>Wuhan Daxue Xuebao
<4>42
<5>237-240
<6>1996
<7>Restriction endonuclease Bsp63I has been isolated and purified by affinity chromatography.
The role of specific amino acid residues in Bsp63I was determined by chemical modification.
Sulfhydryl groups were modified with p-chloromercuribenzoic acid, lysine residues with
pyridoxal-5'-phosphate (PLP) and arginine residues with 2,3-butanedione.  The results show
that these residues are related to the activity of Bsp63I.  Dynamic analysis shows that the
type of PLP inhibition is analogous anticompetitive inhibition. *>

<>

<1>Dong, X., El Karkouri, K., Robert, C., Gavory, F., Raoult, D., Fournier, P.E.
<2>Genomic Comparison of Rickettsia helvetica and Other Rickettsia Species.
<3>J. Bacteriol.
<4>194
<5>2751
<6>2012
<7>We report the complete and annotated genome sequence of Rickettsia helvetica strain C9P9,
which was first isolated in 1979 from Ixodes ricinus ticks in
Switzerland and is considered a human pathogen.

<>

<1>Dong, X., El Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Rickettsia australis, the Agent of Queensland Tick Typhus.
<3>J. Bacteriol.
<4>194
<5>5129
<6>2012
<7>Rickettsia australis strain Phillips(T) was isolated in Queensland, Australia, in 1950. It is
the tick-borne agent of Queensland tick typhus, a disease endemic in
Australia. The 1.29-Mb genome sequence of this bacterium is highly similar to
that of Rickettsia akari but contains two plasmids.

<>

<1>Dong, X., El-Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Genomic Analysis of Rickettsia japonica Strain YHT.
<3>J. Bacteriol.
<4>194
<5>6992
<6>2012
<7>Rickettsia japonica strain YH, isolated in 1984 in Japan, is the type strain of R. japonica,
the tick-borne agent of Japanese spotted fever. Here, we report the
1.33-Mb genome of this rickettsial species.

<>

<1>Doolitle, M.M., Sirotkin, K.
<2>Bacteriophage T2 and T4, dam+ and damh and Eco dam+ methylation: preference at different sites.
<3>Biochim. Biophys. Acta
<4>949
<5>240-246
<6>1988
<7>We present a method for determining preference for methylation at minor methylation sites. The
target DNA sequence is first subjected to computer-assisted analysis to predict which
restriction endonuclease(s) will generate fragments that will contain only one or two likely
minor methylation site(s). The target DNA is then methylated in vitro with a radioactive
methyl-group donor and subjected to digestion by the chosen restriction enzyme(s). The amount
of radioactivity in the various fragments is determined, after separating them using
polyacrylamide gel electrophoresis. We documented the effect of nearby bases on the
methylation preference and the relative preference for methylation at some specific minor
methylation sites.

<>

<1>Doolittle, R.F.
<2>The comings and goings of homing endonucleases and mobile introns.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>5379-5381
<6>1993
<7>In this issue Belfort and coworkers report the isolation and characterization of a
thermostable endonuclease from the archaebacterium Desulfurococcus mobilis. Remarkably, the
enzyme is encoded in an intron in the single gene for 23S rRNA and is expressed only after the
corresponding RNA segment is excised from the newly made rRNA. Archaebacterial introns are
unique in that their excision is catalyzed by an enzyme. The endonuclease has a number of
features characteristic of the homing endonucleases found in fungi and their organelles, and
these have led the authors to wonder whether trans-kingdom gene transfer may have been
involved. As the authors suggest, all of these topics--introns, archaebacteria, homing
endonucleases, and horizontal transfers--are lightning rods for debate.

<>

<1>Dopazo, J., Mendoza, A., Herrero, J., Caldara, F., Humbert, Y., Friedli, L., Guerrier, M., Grand-Schenk, E., Gandin, C., DeFrancesco, M., Polissi, A., Buell, G., Feger, G., Garcia, E., Peitsch, M., Garcia-Bustos, J.F.
<2>Annotated draft genomic sequence from a Streptococcus pneumoniae type 19F clinical isolate.
<3>Microb. Drug Resist.
<4>7
<5>99-125
<6>2001
<7>The public availability of numerous microbial genomes is enabling the analysis of bacterial
biology in great detail and with an unprecedented, organism-wide and taxon-wide, broad scope.
Streptococcus pneumoniae is one of the most important bacterial pathogens throughout the
world.  We present here sequences and functional annotations for 2.1-Mbp of pneumococcal DNA,
covering more than 90% of the total estimated size of the genome.  The sequenced strain is a
clinical isolate resistant to macrolides and tetracycline.  It carries a type 19F capsular
locus, but multilocus sequence typing for several conserved genetic loci suggests that the
strain sequence belongs to a pneumococcal lineage that most often expresses a serotype 15
capsular polysaccharide.  A total of 2,046 putative open reading frames longer than 100 amino
acids were identified (average of 1,009 bp per ORF), including all described two-component
systems and aminoacyl tRNA synthetases.  Comparisons to other complete, or nearly complete,
bacterial genomes were made and are presented in a graphical form for all the predicted
proteins.

<>

<1>Dordet-Frisoni, E. et al.
<2>Draft Genome Sequences of Mycoplasma auris and Mycoplasma yeatsii, Two Species of the Ear Canal of Caprinae.
<3>Genome Announcements
<4>1
<5>e00280-13
<6>2013
<7>We report here the draft genome sequences of Mycoplasma auris and Mycoplasma yeatsii, two
species commonly isolated from the external ear canal of Caprinae.

<>

<1>Dornberger, U., Leijon, M., Fritzsche, H.
<2>High base pair opening rates in tracts of GC base pairs.
<3>J. Biol. Chem.
<4>274
<5>6957-6962
<6>1999
<7>Sequence-dependent structural features of the DNA double helix have a strong influence on the
base pair opening dynamics. Here we report a detailed study of the kinetics of base pair
breathing in tracts of GC base pairs in DNA duplexes derived from 1H NMR measurements of the
imino proton exchange rates upon titration with the exchange catalyst ammonia. In the limit of
infinite exchange catalyst concentration, the exchange times of the guanine imino protons of
the GC tracts extrapolate to much shorter base pair lifetimes than commonly observed for
isolated GC base pairs. The base pair lifetimes in the GC tracts are below 5 ms for almost all
of the base pairs. The unusually rapid base pair opening dynamics of GC tracts are in striking
contrast to the behavior of AT tracts, where very long base pair lifetimes are observed. The
implication of these findings for the structural principles governing spontaneous helix
opening as well as the DNA-binding.

<>

<1>Dorner, L.F., Bitinaite, J., Whitaker, R.D., Schildkraut, I.
<2>Genetic analysis of the base-specific contacts of BamHI restriction endonuclease.
<3>J. Mol. Biol.
<4>285
<5>1515-1523
<6>1999
<7>Here, we investigate the highly specific interaction of the BamHI endonuclease with its
cognate recognition sequence GGATCC by determining which amino acid residues can be
substituted at the DNA interface while maintaining specificity.  Mutational studies, together
with the structural determination of the restriction endonuclease BamHI have revealed the
amino acid residues which are involved in DNA catalysis and those which play a role in the
specific binding of the enzyme to its cognate DNA recognition sequence.  Amino acid residues
N116, S118, R122, D154 and R155 are involved in DNA sequence recognition and are located in
the major groove in close proximity to the nucleotide bases comprising the recognition
sequence.  Cassette mutagenesis of these amino acids, together with in vivo transcriptional
interference selection, was used to identify an array of substitutions which maintain
site-specific binding to the cognate GGATCC sequence.  This approach has demonstrated the
extent of acceptable variation among amino acid residues which are directly involved in
site-specific binding.  One variant, double mutant N116H, S118G was found to cleave DNA only
when the adenine base in the recognition site is methylated.

<>

<1>Dorner, L.F., Schildkraut, I.
<2>Direct selection of binding proficient/catalytic deficient variants of BamHI endonuclease.
<3>Nucleic Acids Res.
<4>22
<5>1068-1074
<6>1994
<7>Variants of BamHI endonuclease in which the glutamate 113 residue has been changed to lysine
or the aspartate 94 to asparagine were shown to behave as repressor molecules in vivo. This
was demonstrated by placing a BamHI recognition sequence, GGATCC, positioned as an operator
sequence in an antisense promoter for the aadA gene (spectinomycin resistance). Repression of
this promoter relieved the inhibition of expression of spectinomycin resistance. This system
was then used to select new binding proficient/cleavage deficient BamHI variants. The BamHI
endonuclease gene was mutagenized either by exposure to hydroxylamine or by PCR. The
mutagenized DNA was reintroduced into E.coli carrying the aadA gene construct, and
transformants that conferred spectinomycin resistance were selected. Twenty SP'transformants
were sequenced. Thirteen of these were newly isolated variants of the previously identified
D94 and E113 residues which are known to be involved in catalysis. The remaining seven
variants were all located at residue 111 and the glutamate 111 residue was shown to be
involved with catalysis.

<>

<1>Doroghazi, J.R., Ju, K.S., Brown, D.W., Labeda, D.P., Deng, Z., Metcalf, W.W., Chen, W., Price, N.P.
<2>Genome Sequences of Three Tunicamycin-Producing Streptomyces Strains, S. chartreusis NRRL 12338, S. chartreusis NRRL 3882, and S. lysosuperificus  ATCC 31396.
<3>J. Bacteriol.
<4>193
<5>7021-7022
<6>2011
<7>We announce the sequencing of Streptomyces chartreusis NRRL 12338 and NRRL 3882 and
Streptomyces lysosuperificus ATCC 31396. These are producers of
tunicamycins, chartreusins, cephalosporins, holomycins, and calcimycin.
The announced genomes, together with the published Streptomyces
clavuligerus genome, will facilitate data mining of these secondary
metabolites.

<>

<1>Doronina, V.A., Murray, N.E.
<2>Proteolytic control of restriction by the type I restriction enzyme EcoKI.
<3>Biochem. Soc. Trans.
<4>2000
<5>A177
<6>2000
<7>Type I restriction systems are sophisticated molecular machines that methylate (modify) or cut
duplex DNA depending upon the methylation status of their target sequences.  When unmodified
DNA enters the bacterial cells this DNA is a substrate for restriction, a process in which
extensive DNA translocation precedes DNA breakage.  An amino acid substitution in EcoKI, which
changes a motif essential for the active site of the modification activity, leads to an enzyme
that is unable to modify the DNA but retains the ability to recognize unmodified target
sequences and make double-stranded breaks in the DNA.  In vivo, breakage of unmodified
chromosomes is prevented by a posttranslational mechanism that depends on ClpXP-dependent
proteolysis and leads to the degradation of the subunit within the restriction enzyme that is
essential for restriction but not modification.  Analysis of mutants that affect different
stages of the restriction reaction suggests a model in which the degradation occurs after the
enzyme has recognized unmodified target sequences and initiated the DNA translocation that
precedes DNA breakage.  The mechanisms that allow cells to distinguish between chromosomal and
foreign DNA are under investigation.

<>

<1>Doronina, V.A., Murray, N.E.
<2>The proteolytic control of restriction activity in Escherichia coli K-12.
<3>Mol. Microbiol.
<4>39
<5>416-428
<6>2001
<7>The endonuclease activity of EcoKI is regulated by the ClpXP-dependent degradation of the
subunit that is essential for restriction, but not modification.  We monitored proteolysis in
mutants blocked at different steps in the restriction pathway.  Mutations that prevent DNA
translocation render EcoKI refractory to proteolysis, whereas those that permit DNA
translocation, but block endonuclease activity, do not.  Although proteolysis alleviates
restriction in a mutant that lacks modification activity, some restriction activity remains;
our evidence indicates residual EcoKI associated with the membrane fraction.  ClpXP protects
the bacterial chromosome, but little effect was detected on unmodified foreign DNA within the
cytoplasm of a restriction-proficient cell.  The molecular basis for the distinction between
unmodified resident and foreign DNA remains to be determined.

<>

<1>Dorscht, J., Klumpp, J., Bielmann, R., Schmelcher, M., Born, Y., Zimmer, M., Calendar, R., Loessner, M.J.
<2>Comparative genome analysis of Listeria bacteriophages reveals extensive mosaicism, programmed translational frameshifting, and a novel prophage insertion site.
<3>J. Bacteriol.
<4>191
<5>7206-7215
<6>2009
<7>The genomes of six Listeria bacteriophages were sequenced and analyzed.
Phages A006, A500, B025, P35, and P40 are members of the Siphoviridae and
contain double-stranded DNA genomes of between 35.6 kb and 42.7 kb. Phage
B054 is a unique myovirus and features a 48.2-kb genome. Phage B025
features 3' overlapping single-stranded genome ends, whereas the other
viruses contain collections of terminally redundant, circularly permuted
DNA molecules. Phages P35 and P40 have a broad host range and lack
lysogeny functions, correlating with their virulent lifestyle. Phages
A500, A006, and B025 integrate into bacterial tRNA genes, whereas B054
targets the 3' end of translation elongation factor gene tsf. This is the
first reported case of phage integration into such an evolutionarily
conserved genetic element. Peptide fingerprinting of viral proteins
revealed that both A118 and A500 utilize +1 and -1 programmed
translational frameshifting for generating major capsid and tail shaft
proteins with C termini of different lengths. In both cases, the unusual
+1 frameshift at the 3' ends of the tsh coding sequences is induced by
overlapping proline codons and cis-acting shifty stops. Although Listeria
phage genomes feature a conserved organization, they also show extensive
mosaicism within the genome building blocks. Of particular interest is
B025, which harbors a collection of modules and sequences with relatedness
not only to other Listeria phages but also to viruses infecting other
members of the Firmicutes. In conclusion, our results yield insights into
the composition and diversity of Listeria phages and provide new
information on their function, genome adaptation, and evolution.

<>

<1>Dorvel, B., Sigalov, G., Zhao, Q., Comer, J., Dimitrov, V., Mirsaidov, U., Aksimentiev, A., Timp, G.
<2>Analyzing the forces binding a restriction endonuclease to DNA using a synthetic nanopore.
<3>Nucleic Acids Res.
<4>37
<5>4170-4179
<6>2009
<7>Restriction endonucleases are used prevalently in recombinant DNA technology because they bind
so stably to a specific target sequence and,
in the presence of cofactors, cleave double-helical DNA specifically at a
target sequence at a high rate. Using synthetic nanopores along with
molecular dynamics (MD), we have analyzed with atomic resolution how a
prototypical restriction endonuclease, EcoRI, binds to the DNA target
sequence-GAATTC-in the absence of a Mg(2+) ion cofactor. We have
previously shown that there is a voltage threshold for permeation of DNA
bound to restriction enzymes through a nanopore that is associated with a
nanonewton force required to rupture the complex. By introducing mutations
in the DNA, we now show that this threshold depends on the recognition
sequence and scales linearly with the dissociation energy, independent of
the pore geometry. To predict the effect of mutation in a base pair on the
free energy of dissociation, MD is used to qualitatively rank the
stability of bonds in the EcoRI-DNA complex. We find that the second base
in the target sequence exhibits the strongest binding to the protein,
followed by the third and first bases, with even the flanking sequence
affecting the binding, corroborating our experiments.

<>

<1>Dos Santos, A.P., Guimaraes, A.M., do Nascimento, N.C., Sanmiguel, P.J., Messick, J.B.
<2>Complete Genome Sequence of Mycoplasma wenyonii Strain Massachusetts.
<3>J. Bacteriol.
<4>194
<5>5458-5459
<6>2012
<7>Mycoplasma wenyonii is a hemotrophic mycoplasma that causes acute and chronic infections in
cattle. Here, we announce the first complete genome sequence of
this organism. The genome is a single circular chromosome with 650,228 bp and
G+C% of 33.9. Analyses of M. wenyonii genome will provide insights into its
biology.

<>

<1>Dos Santos, R.A., Berretta, A.A., Barud, H.S., Ribeiro, S.J., Gonzalez-Garcia, L.N., Zucchi, T.D., Goldman, G.H., Riano-Pachon, D.M.
<2>Draft Genome Sequence of Komagataeibacter rhaeticus Strain AF1, a High Producer of Cellulose, Isolated from Kombucha Tea.
<3>Genome Announcements
<4>2
<5>e00731-14
<6>2014
<7>Here, we present the draft genome sequence of Komagatabaeicter rhaeticus strain AF1, which was
isolated from Kombucha tea and is capable of producing high levels
of cellulose.

<>

<1>Doskar, J.
<2>Determination of the restriction-modification system of the polyvalent bacteriophage 821 in the cells of Staphylococcus-aureus strains.
<3>Scripta Fac. Sci. Nat. Univ. Purk. Brun.
<4>18
<5>421
<6>1988
<7>None

<>

<1>Doskar, J.
<2>Characteristics of restriction-modification system of the polyvalent phage 812 in strains of staphylococcus aureus.
<3>Scripta Fac. Sci. Nat. Univ. Purk. Brun.
<4>19
<5>391-401
<6>1989
<7>On the basis of the study of restriction-deficient mutants (r-) a
restriction-modification (RM) system concerning the polyvalent phage 812 has
been characterized in some strains of S. aureus.  Capability to modify DNA of
the phage 812 has been evidenced only in one of the total number of twelve r-
mutant strains under study isolated earlier (Doskar and Rosypal 1986).
Modification acquired by the polyvalent phage 812 in this mutant strain enables
this phage to grow not only on the parent restrictive strain r+ but also on
certain strains insensitive to the non-modified phage.  A similar pattern of
sensitivity of these strains both to the modified phage and to certain
host-range mutants of the phage of the phage 812 leads us to the conclusion
that RM systems in these strains are similar and that the mutations widening
the host-range of the polyvalent phage concern restriction-modification
mechanisms.  Our attempt to detect restriction endonucleases of the class II in
crude extracts from strains insensitive to the polyvalent phage 812 have not
been successful.  RM system of the polyvalent phage 812 is believed not to be
of class II because of the fact that most of r-mutants are
modification-deficient.

<>

<1>Dou, D., Inagaki, K., Kita, K., Ohshima, A., Hiraoka, N., Kishimoto, N., Sugio, T., Tano, T.
<2>Restriction endonuclease AfaI from Acidiphilium facilis, a new isoschizomer of RsaI: purification and properties.
<3>Biochim. Biophys. Acta
<4>1009
<5>83-86
<6>1989
<7>We have purified AfaI endonuclease, an isoschizomer of RsaI, from Acidiphilium
facilis strain 28H.  The enzyme is homogeneous as judged by polyacrylamide gel
electrophoresis, and composed of a single polypeptide chain with a molecular
weight of 30,000.  AfaI endonuclease, like RsaI, recognizes the tetranucleotide
sequence 5'-G-T-A-C-3', and cleaves between the T and A to produce blunt-ended
fragments.  The yield of the enzyme is 50-100 times that of the RsaI, which is
from a phototrophic bacterium, Rhodopseudomonas sphaeroides strain 28/5.

<>

<1>Douc-Rasy, S., Kolb, A., Prunell, A.
<2>Protein-induced unwinding of DNA:  measurement by gel electrophoresis of complexes with DNA minicircles.  Application to restriction endonuclease EcoRI, catabolite gene activator protein and lac repressor.
<3>Nucleic Acids Res.
<4>17
<5>5173-5189
<6>1989
<7>An electrophoretic procedure for the measurement of the helix unwinding induced
by a sequence-specific protein is described.  The method, which was applied
here to EcoRI, CAP and lac repressor, involved the migration of the complexes
with positively and negatively supercoiled DNA minicircles carrying a single
protein binding site.  Mobility shifts of complexes relative to naked DNAs
appeared to be a result of 1) the unwinding; of 2) an increase in the molecular
frictional coefficient, which led to a retardation; of 3) bending, in the
particular case of CAP, which induced an acceleration; and of 4) looping, in
the case of lac repressor, which also resulted in an acceleration.  Under
conditions where the migration of the naked topoisomers was V-like (topoisomer
mobility showed the same linear increase with both negative and positive
supercoilings; Zivanovic et al. (1986) J. Mol. Biol., 192, 645-660), the
protein unwinding contribution to mobility was assumed to be identical to that
experimentally observed in the case of a thermal unwinding:  all negatively
supercoiled topoisomers were retarded and all positively supercoiled
topoisomers were accelerated to the same extent.  In contrast, the mobility
contribution of the frictional term, as well as those of bending and looping,
appeared to vary strongly with the magnitude of the supercoiling, but only
weakly with its polarity.  As a consequence, these latter contributions may
approximately cancel when one is measuring the difference between the shifts
observed for two comigrating, negatively and positively supercoiled,
topoisomers, allowing the unwinding to be calculated.  While estimates obtained
for EcoRI, 23 +/- 3, and CAP, about 29, were in good agreement with previous
measurements using topoisomerase I, the value found for lac repressor, 13 to
16, was significantly smaller.

<>

<1>Doucette-Stamm, L., Bush, D.
<2>Nucleic acid and amino acid sequences relating to Staphylococcus epidermidis for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7060458 B
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Staphylococcus epidermidis that are useful in diagnosis and therapy of pathological
conditions; antibodies against the polypeptides; and methods for the production of the
polypeptides.  The invention also provides methods for the detection, prevention and treatment
of pathological conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D.
<2>Nucleic acid and amino acid sequences relating to Staphylococcus epidermidis for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7183083 B
<5>
<6>2007
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Staphylococcus epidermidis that are useful in diagnosis and therapy of pathological
conditions; antibodies against the polypeptides; and methods for the production of the
polypeptides.  The invention also provides methods for the detection, prevention and treatment
of pathological conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, O., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7115731 B
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid
sequences derived from Streptococcus pneumoniae that are useful in
diagnosis and therapy of pathological conditions; antibodies against the
polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment
of pathological conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6699703 B
<5>
<6>2004
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7081530 B
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7098023 B
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid
sequences derived from Streptococcus pneumoniae that are useful in
diagnosis and therapy of pathological conditions; antibodies against the
polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment
of pathological conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7122368 A
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7129339 A
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7129340 A
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7135560 A
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7151171 A
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7153952 A
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7074914 B
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7326544 A
<5>
<6>2008
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7335493 A
<5>
<6>2008
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7335494 A
<5>
<6>2008
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment for pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L., Bush, D., Zeng, Q., Opperman, T., Houseweart, C.E.
<2>Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7338786 A
<5>
<6>2008
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Streptococcus pneumoniae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doucette-Stamm, L.A., Bush, D.
<2>Nucleic acid sequences and expression system relating to Enterococcus faecium for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6583275 A
<5>
<6>2003
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Enterococcus
faecium that are useful in diagnosis and therapy of pathological conditions; antibodies
against the polypeptides; and methods for the production of the polypeptides.  The invention
also provides methods for the detection, prevention and treatment of pathological conditions
resulting from bacterial infection.

<>

<1>Doucette-Stamm, L.A., Bush, D.
<2>Nucleic acid and amino acid sequences relating to Enterococcus faecalis for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6617156 B
<5>
<6>2003
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Enterococcus faecalis that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Doughty, B., Kazer, S.W., Eisenthal, K.B.
<2>Binding and cleavage of DNA with the restriction enzyme EcoR1 using time-resolved second harmonic generation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>19979-19984
<6>2011
<7>The binding of EcoR1 to a 90-bp DNA duplex attached to colloidal microparticles and the
subsequent cleavage by the enzyme was observed in real time and label-free with time-resolved
second harmonic (SH) spectroscopy. This method provides a unique way to investigate
biomolecular interactions based on its sensitivity to changes in structure and electrical
charge on formation of a complex and subsequent dynamics. The binding of EcoR1 to the
recognition sequence in DNA appears as a rapid increase in the SH signal, which is attributed
to the enzyme-induced change in the DNA conformation, going from a rod-like to a bent shape.
In the presence of the cofactor Mg(2+), the subsequent decay in the SH signal was monitored in
real time as the following processes occurred: cleavage of DNA, dissociation of the enzyme
from the DNA, and diffusion of the 74-bp fragment into the bulk solution leaving the 16-bp
fragment attached to the microparticle. The observed decay was dependent on the concentration
of Mg(2+), which functions as a cofactor and as an electrolyte. With SH spectroscopy the
rehybridization dynamics between the rehybridized microparticle bound and free cleaved DNA
fragments was observed in real time and label-free following the cleavage of DNA.
Collectively, the experiments reported here establish SH spectroscopy as a powerful method to
investigate equilibrium and time-dependent biological processes in a noninvasive and
label-free way.

<>

<1>Downing, M.E., Brady, K.L., Caprara, M.G.
<2>A C-terminal fragment of an intron-encoded maturase is sufficient for promoting group I intron splicing.
<3>RNA
<4>11
<5>437-446
<6>2005
<7>Group I introns often encode proteins that catalyze site-specific DNA hydrolysis. Some of
these proteins have acquired the ability to promote
splicing of their cognate intron, but whether these two activities
reside in different regions of the protein remains obscure. A crystal
structure of I-AniI, a dual function intron-encoded protein, has shown
that the protein has two pseudo-symmetric domains of equal size. Each
domain contacts its DNA substrate on either side of two cleavage sites.
As a first step to identify the RNA binding surface, the N- and
C-terminal domains of I-AniI were separately expressed and tested for
promoting the splicing of the mitochondrial (mt) COB pre-RNA. The
N-terminal protein showed no splicing activation or RNA binding,
suggesting that this domain plays a minimal role in activity or is
improperly folded. Remarkably, the 16-kDa C-terminal half facilitates
intron splicing with a rate similar to that of the full-length protein.
Both the C-terminal fragment and full-length proteins bind tightly to
the COB intron. RNase footprinting shows that the C-terminal and
full-length proteins bind to the same regions and induce the same
conformational changes in the COB intron. Together, these results show
that the C-terminal fragment of I-AniI is necessary and sufficient for
maturase activity and suggests that I-AniI acquired splicing function
by utilizing a relatively small protein surface that likely represents
a novel RNA binding motif. This fragment of I-AniI represents the
smallest group I intron splicing cofactor described to date.

<>

<1>Doyon, J.B., Pattanayak, V., Meyer, C.B., Liu, D.R.
<2>Directed evolution and substrate specificity profile of homing endonuclease I-SceI.
<3>J. Am. Chem. Soc.
<4>128
<5>2477-2484
<6>2006
<7>The laboratory evolution of enzymes with tailor-made DNA cleavage specificities would
represent new tools for manipulating genomes and may enhance our understanding of
sequence-specific DNA recognition by nucleases.  Below we describe the development and
successful application of an efficient in vivo positive and negative selection system that
applies evolutionary pressure either to favor the cleavage of a desired target sequence or to
disfavor the cleavage of nontarget sequences.  We also applied a previously described in vitro
selection method to reveal the comprehensive substrate specificity profile of I-SceI homing
endonucleases with altered DNA cleavage specificities.  The most highly evolved enzyme cleaves
the target mutant DNA sequence with a selectivity that is comparable to wild-type I-SceI's
preference for its cognate substrate.

<>

<1>Dreier, J., Bickle, T.A.
<2>ATPase activity of the type IC restriction-modification system EcoR124II.
<3>J. Mol. Biol.
<4>257
<5>960-969
<6>1996
<7>We have investigated the ATPase activity of the type IC restriction-modification
(R-M) system EcoR124II.  As with all type I R-M systems EcoR124II requires ATP hydrolysis to
cut DNA.  We determined the KM for ATP to be 10^-5 to 10^-4 M.  By measuring ATP
hydrolysis under different conditions and by simultaneously monitoring DNA restriction,
methylation and ATP hydrolysis we propose that the order of events during restriction is: (1)
binding of EcoR124II to a non-methylated recognition sequence, (2) start of DNA-dependent ATP
hydrolysis which continues even after restriction is complete, (3) restriction of DNA, (4)
methylation of the product.  Non-cleavable DNA substrates, such as recognition site containing
oligonucleotides, also support ATP hydrolysis.  Methylation can also occur prior to ATP
hydrolysis and prevent DNA degradation.

<>

<1>Dreier, J., MacWilliams, M.P., Bickle, T.A.
<2>DNA cleavage by the type IC restriction-modification enzyme EcoR124II.
<3>J. Mol. Biol.
<4>264
<5>722-733
<6>1996
<7>Type I restriction-modification systems bind to non-palindromic, bipartite recognition
sequences.  Although these enzymes methylate specific adenine residues within their
recognition sequences, they cut DNA at sites up to several thousand base-pairs away.  We have
investigated the mechanism of how EcoR124II, a type IC restriction-modification system,
selects the cleavage site.  Restriction studies with different DNA constructs revealed that
circular DNA requires only one non-methylated recognition sequence to be cut, whereas linear
DNA needs at least two such sites.  Cleavage of linear DNA is independent of site orientation.
Further investigations of the linear substrates revealed a mechanism whereby the double-strand
break is introduced between two recognition sequences.  We propose a model for the selection
of restriction sites by type I enzymes where two EcoR124II complexes bind to two recognition
sequences.  Lack of methylation at a site stimulates the enzyme to translocate DNA on both
sides of the recognition sequence.  Thus the two complexes approach each other and, at the
point where they meet, they interact to introduce a double-strand break in the DNA.

<>

<1>Dreiseikelmann, B., Eichenlaub, R., Wackernagel, W.
<2>The effect of differential methylation by Escherichia coli of plasmid DNA and phage T7 and lambda DNA on the cleavage by restriction endonuclease MboI from Moraxella bovis.
<3>Biochim. Biophys. Acta
<4>562
<5>418-428
<6>1979
<7>The nucleotide sequence recognized and cleaved by the restriction endonuclease MboI is
5'^GATC and is identical to the central tetranucleotide of the restriction sites of BamHI and
BglII. Experiments on the restriction of DNA from Escherichia coli dam and dam+ confirm the
notion that GATC sequences are adenosyl-methylated by the dam function of E. coli and thereby
are made refractory to cleavage by MboI. On the basis of this observation the degree of dam
methylation of various DNAs was examined by cleavage with MboI and other restriction
endonuclease. In plasmid DNA essentially all of the GATC sequences are methylated by the dam
function. The DNA of phage lambda is only partially methylated. Extended methylation is
observed in the DNA of a substitution mutant of lambda, lambda gal8bio256, and in the lambda
derived plasmid, lambda dv93, which is completely methylated. In contrast, phage T7 DNA is not
methylated by dam. A suppression of dam methylation of T7 DNA appears to act only in cis since
plasmid DNA replicated in a T7-infected cell is completely methylated. The results are
discussed with respect to the participation of the dam methylase in different replication
systems.

<>

<1>Dreiseikelmann, B., Wackernagel, W.
<2>Absence in Bacillus subtilis and Staphylococcus aureus of the sequence-specific deoxyribonucleic acid methylation that is conferred in Escherichia coli K-12 by the dam and dcm enzymes.
<3>J. Bacteriol.
<4>147
<5>259-261
<6>1981
<7>Restriction analysis of plasmid pHV14 deoxyribonucleic acid isolated from Escherichia coli
K-12, Bacillus subtilis, and Staphylococcus aureus with restriction endonucleases MboI,
Sau3AI, and EcoRII was used to study the methylation of those nucleotide sequences which in E.
coli contain the major portions of N6-methyladenine and 5-methylcytosine.

<>

<1>Drew, H.R., Travers, A.
<2>Structural junctions in DNA: the influence of flanking sequence on nuclease digestion specificities.
<3>Nucleic Acids Res.
<4>13
<5>4445-4467
<6>1985
<7>When a protein binds to DNA, the affinity of this protein for its primary site
of interaction may be influenced by the nature of flanking sequences.  This is
thought to be a consequence of local cooperativity in the DNA molecule, where
the conformation at one point along the helix can influence the conformation at
another, and thereby modulate the free energy of protein-DNA recognition.In
order to learn more about this procesas, we have carried out experiments of two
sorts.  First, we have constructed sequences of the type (dA)11(dG)8, where the
conformational preferences of the DNA molecule switch from one extreme to
another over just a single base pair, and subjected them to digestion by DNAase
I and DNAase II.  This is to learn whether the structure changes abruptly at
the junction point, or more gradually with an influence extending into residues
on either side.  Secondly, we have subjected long plasmid DNA to digestion by
restriction enzymes Fnu DII, HaeIII, HhaI and MspI, to look for correlations
between cutting rate and the identity of nucleotides on either side of the
restriction site.  The influence of flanking sequence on nuclease digestion
specificities is clearly evident in both kinds of experiment, but the rules
governing this seem complex and not easily formulated.  The best that can be
done at present is to divide the problem into two parts, "analogue" and
"digital", representing sugar-phosphate and base components of recognition.

<>

<1>Drexler, H., Christensen, J.R.
<2>Genetic crosses between restricted and unrestricted phage T1 in lysogenic and nonlysogenic hosts.
<3>Virology
<4>13
<5>31-39
<6>1961
<7>When Shigella dysenteriae is lysogenized by the phage P1, it becomes immune to
infection with "ordinary" T1 phage ("restricted T1" or "rT1").  A
host-controlled variant of T1 ("unrestricted T1" or "uT1") can multiply in the
lysogenic S. dysenteriae.  Three-factor crosses were done between suitably
marked strains of rT1 and uT1 in both lysogenic and nonlysogenic S.
dysenteriae.  When lysogenic cells are infected with both rT1 and uT1, one
finds some recombinants among the progeny, but few, if any phage bearing the
genotype of the restricted parent.  The proportion of recombinants in the total
yield is about one-eighth to one-twelfth that in control crosses (in
nonlysogenic hosts) and is independent of the order of addition of either
parent and of the time elapsing between the addition of each parent.  We infer,
then, that (1) the genome of rT1 enters the lysogenic cells; (2) it is neither
replicated nor is it destroyed; and (3) it can "mate" with genomes of uT1 which
may be present.  The proportions of the various recombinant classes are
unusual.  Complementary classes are not equal; certain markers may be recovered
together with a high frequency, whereas certain others, more closely linked,
are seldom found together.  Certain "double crossover" classes are more
frequent than certain "single crossover" classes.  These observations can be
explained in terms of a "copy choice" mechanism of recombination by assuming
that the rT1 genome contains at least two "bad spots," the location of which
can be determined approximately and which force a "switch" in replication from
the restricted genome to the unrestricted genome.

<>

<1>Dreyer, K., Schulte-Holthausen, H.
<2>Casein is a potent enhancer for restriction enzyme activity.
<3>Nucleic Acids Res.
<4>19
<5>4295
<6>1991
<7>Restriction enzymes class II are ATP independent and require magnesium ions for
their activity.  Bovine serum albumin (BSA) and spermidine are added in some
reaction buffers.  In our experience addition of spermidine sometimes causes
severe problems with high molecular weight DNA and BSA has only a slight effect
on enzyme activity.  Here we show that casein, the major protein component of
milk is a potent stimulator of restriction enzyme activity.

<>

<1>Dreyfus-Fourcade, M., Sebald, M., Zavadova, M.
<2>Host-controlled restriction and modification of phage r in Clostridium perfringens NCTC 8798 and some of its sporulation mutants.
<3>Ann. Inst. Pasteur Microbiol.
<4>122
<5>1117-1127
<6>1972
<7>The development of phage r, a virulent bacteriophage, in a wild type strain of
C. perfringens and in its sporulation mutants is described.  The phage is fully
accepted by the wild type strain and most of the Spo mutants (r-m+).  Some
mutants have altered host-controlled restriction and modification functions,
i.e., the phage r is restricted and may be partially modified by the mutant
strains.  Strains of phenotype r+m+/-, r+/-m+, r+/-m- and r-m- are described.

<>

<1>Droge, M., Puhler, A., Selbitschka, W.
<2>Phenotypic and molecular characterization of conjugative antibiotic resistance plasmids isolated from bacterial communities of activated sludge.
<3>Mol. Gen. Genet.
<4>263
<5>471-482
<6>2000
<7>In order to isolate antibiotic resistance plasmids from bacterial communities found in
activated sludge, derivatives of the
3-chlorobenzoate-degrading strain Pseudomonas sp. B13, tagged with the
green fluorescent protein as an identification marker, were used as
recipients in filter crosses. Transconjugants were selected on agar
plates containing 3-chlorobenzoate as the sole carbon source and the
antibiotic tetracycline, streptomycin or spectinomycin, and were
recovered at frequencies in the range of 10(-5) to 10(-8) per
recipient. A total of 12 distinct plasmids, designated pB1-pB12, was
identified. Their sizes ranged between 41 to 69 kb and they conferred
various patterns of antibiotic resistance on their hosts. Two of the
plasmids, pB10 and pB11, also mediated resistance to inorganic mercury.
Seven of the 12 plasmids were identified as broad-host-range plasmids
displaying extremely high transfer frequencies in filter crosses,
ranging from 10^-1 to 10^-2 per recipient cell. Ten of the 12
plasmids belonged to the IncP incompatibility group, based on replicon
typing using IncP group-specific PCR primers. DNA sequencing of PCR
amplification products further revealed that eight of the 12 plasmids
belonged to the IncP beta subgroup, whereas two plasmids were
identified as IncP alpha plasmids. Analysis of the IncP-specific PCR
products revealed considerable differences among the IncP beta plasmids
at the DNA sequence level. In order to characterize the gene 'load'
of the IncP plasmids, restriction fragments were cloned and their DNA
sequences established. A remarkable diversity of putative proteins
encoded by these fragments was identified. Besides transposases and
proteins involved in antibiotic resistance, two putative DNA invertases
belonging to the Din family, a methyltransferase of a type I
restriction/modification system, a superoxide dismutase. parts of a
putative efflux system belonging to the RND family, and proteins of
unknown function were identified.

<>

<1>Drotschmann, K., Aronshtam, A., Fritz, H.-J., Marinus, M.G.
<2>The Escherichia coli MutL protein stimulates binding of Vsr and MutS to heteroduplex DNA.
<3>Nucleic Acids Res.
<4>26
<5>948-953
<6>1998
<7>Vsr DNA mismatch endonuclease is the key enzyme of very short patch (VSP) DNA mismatch repair
and nicks the T-containing strand at the site of a T-G mismatch in a sequence-dependent
manner. MutS is part of the mutHLS repair system and binds to diverse mismatches in DNA. The
function of the mutL gene product is currently unclear but mutations in the gene abolish
mutHLS-dependent repair. The absence of MutL severely reduces VSP repair but does not abolish
it. Purified MutL appears to act catalytically to bind Vsr to its substrate; one-hundredth of
an equivalent of MutL is sufficient to bring about a significant effect. MutL enhances binding
of MutS to its substrate 6-fold but does so in a stoichiometric manner. Mutational studies
indicate that the MutL interaction region lies within the N-terminal 330 amino acids and that
the MutL multimerization region is at the C-terminal end. MutL mutant monomeric forms can
stimulate MutS binding.

<>

<1>Drouin, M., Lucas, P., Otis, C., Lemieux, C., Turmel, M.
<2>Biochemical characterization of I-CmoeI reveals that this H-N-H homing endonuclease shares functional similarities with H-N-H colicins.
<3>Nucleic Acids Res.
<4>28
<5>4566-4572
<6>2000
<7>Endonuclease assays of the H-N-H proteins encoded by two group I introns in the Chlamydomonas
moewusii chloroplast psbA gene revealed that the CmpsbA.1 intron specifies a site-specific DNA
endonuclease, designated I-CmoeI. Like most previously reported intron-encoded endonucleases,
I-CmoeI generates a double-strand break near the insertion site of its encoding intron,
leaving 3' extensions of 4 nt. This enzyme was purified from Escherichia coli as a fusion
protein with a His tag at its N-terminus. The recombinant protein (rI-CmoeI) requires a
divalent alkaline earth cation for DNA cleavage (Mg(2+) > Ca(2+) > Sr(2+) > Ba(2+)).  It also
requires a metal cofactor for DNA binding, a property shared with H-N-H colicins but not with
the homing endonucleases characterized to date. rI-CmoeI binds its recognition sequence as a
monomer, as revealed by gel retardation assays. K:(m) and k(cat) values of 100 +/- 40 pM and
0.26 +/- 0.04 min(-1), respectively, were determined. Replacement of the first histidine of
the H-N-H motif by an alanine residue abolishes both rI-CMOE:I activity and binding to its
substrate. We propose that this conserved histidine residue plays a role in binding the metal
cofactor and that such binding induces a structural modification of the enzyme which is
required for DNA recognition.

<>

<1>Drozdz, M., Piekarowicz, A., Bujnicki, J.M., Radlinska, M.
<2>Novel non-specific DNA adenine methyltransferases.
<3>Nucleic Acids Res.
<4>40
<5>2119-2130
<6>2012
<7>The mom gene of bacteriophage Mu encodes an enzyme that converts adenine to
N(6)-(1-acetamido)-adenine in the phage DNA and thereby protects the viral genome from
cleavage by a wide variety of restriction endonucleases. Mu-like prophage sequences present in
Haemophilus influenzae Rd (FluMu), Neisseria meningitidis type A strain Z2491 (Pnme1) and H.
influenzae biotype aegyptius ATCC 11116 do not possess a Mom-encoding gene. Instead, at the
position occupied by mom in Mu they carry an unrelated gene that encodes a protein with
homology to DNA adenine N(6)-methyltransferases (hin1523, nma1821, hia5, respectively).
Products of the hin1523, hia5 and nma1821 genes modify adenine residues to N(6)-methyladenine,
both in vitro and in vivo. All of these enzymes catalyzed extensive DNA methylation; most
notably the Hia5 protein caused the methylation of 61% of the adenines in lambda DNA. Kinetic
analysis of oligonucleotide methylation suggests that all adenine residues in DNA, with the
possible exception of poly(A)-tracts, constitute substrates for the Hia5 and Hin1523 enzymes.
Their potential 'sequence specificity' could be summarized as AB or BA (where B = C, G or
T). Plasmid DNA isolated from Escherichia coli cells overexpressing these novel DNA
methyltransferases was resistant to cleavage by many restriction enzymes sensitive to adenine
methylation.

<>

<1>Dryden, D.T.
<2>Reeling in the bases.
<3>Nat. Struct. Mol. Biol.
<4>11
<5>804-806
<6>2004
<7>EcoR124I is a type I DNA restriction and modification enzyme. The single-molecule magnetic
tweezers technique reveals that this
sophisticated molecular machine is capable of moving thousands of base
pairs of DNA in one binding event.

<>

<1>Dryden, D.T., Davies, G.D., Martin, I., Powell, L.M., Murray, N.E., Ellis, D.J., Berge, T., Edwardson, J.M., Henderson, R.M.
<2>The assembly of the EcoKI type I DNA restriction/modification enzyme and its interaction with DNA.
<3>Biochem. Soc. Trans.
<4>27
<5>691-696
<6>1999
<7>
<>

<1>Dryden, D.T., Edwardson, J.M., Henderson, R.M.
<2>DNA translocation by type III restriction enzymes: a comparison of current models of their operation derived from ensemble and single-molecule  measurements.
<3>Nucleic Acids Res.
<4>39
<5>4525-4531
<6>2011
<7>Much insight into the interactions of DNA and enzymes has been obtained using a number of
single-molecule techniques. However, recent results  generated using two of these
techniques-atomic force microscopy (AFM) and  magnetic tweezers (MT)-have produced apparently
contradictory results when applied to the action of the ATP-dependent type III restriction
endonucleases on DNA. The AFM images show extensive looping of the DNA  brought about by the
existence of multiple DNA binding sites on each enzyme and enzyme dimerisation. The MT
experiments show no evidence for looping being a requirement for DNA cleavage, but instead
support a diffusive sliding of the enzyme on the DNA until an enzyme-enzyme collision occurs,
leading to cleavage. Not only do these two methods appear to disagree, but also the models
derived from them have difficulty explaining some ensemble biochemical results on DNA
cleavage. In this 'Survey and Summary', we describe several different models put forward for
the action of type III restriction enzymes and their inadequacies. We also attempt to
reconcile the different models and indicate areas for further experimentation to elucidate the
mechanism of these enzymes.

<>

<1>Dryden, D.T., Murray, N.E., Rao, D.N.
<2>Nucleoside triphosphate-dependent restriction enzymes.
<3>Nucleic Acids Res.
<4>29
<5>3728-3741
<6>2001
<7>The known nucleoside triphosphate-dependent restriction enzymes are hetero-oligomeric proteins
that behave as molecular machines in response to their target sequences. They translocate DNA
in a process dependent on the hydrolysis of a nucleoside triphosphate. For the ATP-dependent
type I and type III restriction and modification systems, the collision of translocating
complexes triggers hydrolysis of phosphodiester bonds in unmodified DNA to generate
double-strand breaks. Type I endonucleases break the DNA at unspecified sequences remote from
the target sequence, type III endonucleases at a fixed position close to the target sequence.
Type I and type III restriction and modification (R-M) systems are notable for effective
post-translational control of their endonuclease activity. For some type I enzymes, this
control is mediated by proteolytic degradation of that subunit of the complex which is
essential for DNA translocation and breakage. This control, lacking in the well-studied type
II R-M systems, provides extraordinarily effective protection of resident DNA should it
acquire unmodified target sequences. The only well-documented GTP-dependent restriction
enzyme, McrBC, requires methylated target sequences for the initiation of phosphodiester bond
cleavage.

<>

<1>Dryden, D.T.F.
<2>Bacterial DNA methyltransferases.
<3>S-Adenosylmethionine-dependent Methyltransferases: Structures and Functions., World Scientific Publishing, Cheng, X., Blumenthal, R.M., Singapore
<4>
<5>283-340
<6>1999
<7>There are four entries for DNA MTases in the E.C. database.  Classes 2.1.1.37 and 2.1.1.73
both methylate carbon 5 of the cytosine base and differ in that the former group contains a
large regulatory domain and is only found in multicellular eukarotes.  Classes 2.1.1.72 and
2.1.1.113 act on the exocyclic amino groups of adenine and cytosine respectively.  Thus, three
modifications carried out by MTases have now been identified. Cytosine can be modified at
either the C5 or N4 position, and adenine at the N6 position.  These methylation sites are all
in the major groove of B-form DNA which is also the region which allows the most effective
recognition of a DNA sequence via hydrogen bonding and hydrophobic interactions.

<>

<1>Dryden, D.T.F.
<2>The Architecture of Restriction Enzymes.
<3>Structure
<4>21
<5>1720-1721
<6>2013
<7>In this issue of Structure, Lyumkis and colleagues describe a high resolution structure of a
polymerized form of the SgrAl restriction enzyme, which shows that it forms a helical assembly
with four enzyme molecules per turn of the helix. The DNA is arranged on the periphery of the
protein helix pointing away from the helical axis.

<>

<1>Dryden, D.T.F., Cooper, L.P., Murray, N.E.
<2>Purification and characterization of the methyltransferase from the Type I restriction and modification system of Escherichia coli K12.
<3>J. Biol. Chem.
<4>268
<5>13228-13236
<6>1993
<7>The DNA methyltransferase component of the type I restriction and modification enzyme of
Escherichia coli K12 has been purified. The active component, a trimer of molecular mass 170
kDa consisting of one DNA recognition subunit(S) and two modification subunits(M), showed the
expected preference for modifying a hemimethylated substrate rather than an unmethylated one.
Small amounts of the dimers M2 and M1S1 were also isolated. Subunit rearrangements of the
three protein species occurred on ion exchange and heparin-agarose chromatography.
Denaturation of the trimer gave folding intermediates, and these and the dimer forms isolated
during purification may reflect the assembly of the protein in vivo. Enzyme activity was
recovered on refolding the denatured protein by dilution of the denaturant. A comparison of
the predicted isoelectric points of all known S subunits of type I restriction and
modification enzymes revealed values that correlated with the arrangement of type I systems in
several families. Electrostatic interactions may explain the different subunit stoichiometries
observed during purification of type I enzymes and the differing preferences for
hemimethylated DNA displayed by the three type I families.

<>

<1>Dryden, D.T.F., Cooper, L.P., Murray, N.E.
<2>Assembly of the multifunctional EcoKI DNA restriction enzyme in vitro.
<3>Tech. Prot. Chem.
<4>8
<5>593-601
<6>1997
<7>Type I DNA restriction/modification systems have been found in many strains of Escherichia
coli and Salmonella enterica and several other gram negative and positive bacteria.  They
maintain the modification of the host chromosome after DNA replication by methylating adenine
bases on the newly synthesized DNA strand within specific DNA target sequences.  This
methylation reaction is triggered by the recognition of targets which are methylated on the
parental DNA stand.  If methylation is not detected on either strand then the restriction
reaction is triggered.  Unmodified target sequences will exist on foreign DNA, usually of
viral origin.  A type I system cleaves the foreign DNA thereby preventing (restricting) its
replication and propagation.  In contrast to the widely used type II restriction/modification
systems which have separate restriction endonucleases and modification methyltransferases, the
type I systems combine both activities in one large oligomeric enzyme.

<>

<1>Dryden, D.T.F., Cooper, L.P., Thorpe, P.H., Byron, O.
<2>The in vitro assembly of the EcoKI type I DNA restriction/modification enzyme and its in vivo implications.
<3>Biochemistry
<4>36
<5>1065-1076
<6>1997
<7>Type I DNA restriction/modification enzymes protect the bacterial cell from viral infection by
cleaving foreign DNA which lacks N6-adenine methylation within a target sequence and
maintaining the methylation of the targets on the host chromosome.  It has been noted that the
genes specifying type I systems can be transferred to a new host lacking the appropriate,
protective methylation without any adverse effect.  The modification phenotype apparently
appears before the restriction phenotype, but no evidence for transcriptional or translational
control of the genes and the resultant phenotypes has been found.  Type I enzymes contain
three types of subunit, S for sequence recognition, M for DNA modification (methylation), and
R for DNA restriction (cleavage), and can function solely as an M2S1 methylase or as a R2M2S1
bifunctional methylase/nuclease.  We show that the methylase is not stable at the
concentrations expected to exist in vivo, dissociating into free M subunit and M1S1, whereas
the complete nuclease is a stable structure.  The M1S1 form can bind the R subunit as
effectively as the M2S1 methylase but possesses no activity; therefore, upon establishment of
the system in a new host, we propose that most of the R subunit will initially be trapped in
an inactive complex until the methylase has been able to modify and protect the host
chromosome.  We believe that the in vitro assembly pathway will reflect the in vivo situation,
thus allowing the assembly process to at least partially explain the observations that the
modification phenotype appears before the restriction phenotype upon establishment of a type I
system in a new host cell.

<>

<1>Dryden, D.T.F., Davies, G.D., Powell, L.M., Murray, N.E., Ellis, D.J., Berge, T., Edwardson, J.M., Henderson, R.M.
<2>The assembly of the EcoKI type I DNA restriction/modification enzyme and its interaction with DNA.
<3>Biochem. Soc. Trans.
<4>27
<5>A87
<6>1999
<7>Type I DNA restriction-modification enzymes protect the bacterial cell from viral infection by
cleaving foreign DNA which lacks N6-adenine methylation within a target sequence and
maintaining the methylation of the targets on the host chromosome.  They comprise three types
of subunit, S M and R and can function solely as an M2S1 methylase or as an R2M2S1
bifunctional methylase/nuclease.  The subunits contain domains related to those in other
smaller methylases, nucleases and helicases.  The nuclease is a stable structure, whereas the
methylase dissociates into nonfunctional forms M and M1S1 at the concentrations expected to
exist in vivo.  The restriction reaction relies upon extensive DNA translocation driven by ATP
hydrolysis prior to endonucleolytic DNA cleavage.  Cleavage occurs between two,
widely-separated, target sites.  This is consistent with the translocation process causing the
collision of two enzymes on the DNA.  However, atomic force microscopy has suggested that DNA
binding induces dimerization of the enzyme prior to the initiation of translocation and that
cleavage occurs once the DNA loop bound between the two enzymes has been pulled in towards the
enzymes.

<>

<1>Dryden, D.T.F., Sturrock, S.S., Winter, M.
<2>Structural modelling of a type I DNA methyltransferase.
<3>Nat. Struct. Biol.
<4>2
<5>632-635
<6>1995
<7>Amino-acid sequence comparison and tertiary structure modelling suggest a structure for type I
DNA methyltransferases and an evolutionary link to type II methyltransferases.

<>

<1>Dryden, D.T.F., Willcock, D.F., Murray, N.E.
<2>Mutational analysis of conserved amino-acid motifs in EcoKI adenine methyltransferase.
<3>Gene
<4>157
<5>123-124
<6>1995
<7>The EcoKI methyltransferase (M.EcoKI, MTase) contains the amino acid (aa) sequences AAGTA and
NPPF believed to represent the two sequences that are strongly conserved in adenine MTases.
We have analysed a mutation in the first sequence that abolishes cofactor binding and enzyme
activity, and mutations in the second sequence that reduce or abolish activity without
affecting cofactor and DNA binding.

<>

<1>Du J, Z.X., Bernatavichute, Y.V., Stroud, H., Feng, S., Caro, E., Vashisht, A.A., Terragni, J., Chin, H.G., Tu, A., Hetzel, J., Wohlschlegel, J.A., Pradhan, S., Patel, D.J., Jacobsen, S.E.
<2>Dual binding of chromomethylase domains to H3K9me2-containing nucleosomes directs DNA methylation in plants.
<3>Cell
<4>151
<5>167-180
<6>2012
<7>DNA methylation and histone modification exert epigenetic control over gene expression. CHG
methylation by CHROMOMETHYLASE3 (CMT3) depends on histone H3K9
dimethylation (H3K9me2), but the mechanism underlying this relationship is poorly
understood. Here, we report multiple lines of evidence that CMT3 interacts with
H3K9me2-containing nucleosomes. CMT3 genome locations nearly perfectly correlated
with H3K9me2, and CMT3 stably associated with H3K9me2-containing nucleosomes.
Crystal structures of maize CMT3 homolog ZMET2, in complex with H3K9me2 peptides,
showed that ZMET2 binds H3K9me2 via both bromo adjacent homology (BAH) and chromo
domains. The structures reveal an aromatic cage within both BAH and chromo
domains as interaction interfaces that capture H3K9me2. Mutations that abolish
either interaction disrupt CMT3 binding to nucleosomes and show a complete loss
of CMT3 activity in vivo. Our study establishes dual recognition of H3K9me2 marks
by BAH and chromo domains and reveals a distinct mechanism of interplay between
DNA methylation and histone modification.

<>

<1>Du Y, S.L., Feng, W., Pei, G., Zheng, P., Yu, Z., Sun, J., Qiao, J.
<2>Draft Genome Sequence of Lactococcus lactis subsp. lactis Strain YF11.
<3>Genome Announcements
<4>1
<5>e00599-13
<6>2013
<7>Lactococcus lactis subsp. lactis strain YF11 is a food preservative bacterium with a high
capacity to produce nisin. Here, we announce the draft genome
sequence of Lactococcus lactis subsp. lactis YF11 (2,527,433 bp with a G+C
content of 34.81%).

<>

<1>Du, L., Zhang, M., Burton, R., Spielberger, K., Mollova, E., Gilmanshin, R.
<2>Rapid mapping of large DNA molecules using fluorescent labeled restriction enzyme.
<3>Biophys. J.
<4>S
<5>331A
<6>2007
<7>Rapid DNA mapping at the single molecule level is a powerful tool for research and clinical
application. We report here a development of new tags for high throughput mapping of large DNA
based on our Direct Linear Analysis (DLA) technology. Double-stranded DNA molecules were
stained with two fluorescent tags: a nonspecific intercalator for uniform backbone staining
and a sequence-specific tag for physical mapping. These DNA molecules were then stretched by
elongational flow to a linear conformation in a high-throughput microfluidic device, and the
positions of site-specific tags were detected. We applied restriction enzymes for DNA tagging
due to their unique features including high binding affinity, sequence specificity, and fast
target localization. The restriction enzymes selected for tagging retained high binding
efficiency in the presence of Ca++ after fluorescent-labeling; however, their cleavage
function was inactivated. We show that DNA fragments ranging from 50-200 kb, or 16-66
microns-long, can be tagged with fluorescently labeled AgeI or BamHI, intercalated, stretched
to their contour length in our device, and then interrogated molecule by molecule. More than
5,000 molecules could be analyzed through the microfluidic device in 10-15 minutes and
identity of the DNA fragments revealed by tagging patterns as well as lengths. We analyzed
multiple genomic DNA fragments from E. coli stains, demonstrating great potential of using
this method to obtain information regarding DNA structure and to discriminate DNA fragments
originated from different organisms.

<>

<1>Du, X.J., Jia, S.R., Yang, Y., Wang, S.
<2>Genome Sequence of Gluconacetobacter sp. Strain SXCC-1, Isolated from Chinese Vinegar Fermentation Starter.
<3>J. Bacteriol.
<4>193
<5>3395-3396
<6>2011
<7>Gluconacetobacter are prominent bacteria during traditional vinegar fermentation. Here, we
report a draft genome sequence of Gluconacetobacter
sp. strain SXCC-1. This strain was isolated from fermentation starter
(Daqu) that used for commercial production of Shanxi vinegar, the best
known vinegar of China.

<>

<1>Du, Z., Zhang, Z., Miao, T., Wu, J., Lu, G., Yu, J., Xiao, J., Chen, G.
<2>Draft Genome Sequence of the Novel Agar-Digesting Marine Bacterium HQM9.
<3>J. Bacteriol.
<4>193
<5>4557
<6>2011
<7>Strain HQM9, an aerobic rod-shaped marine bacterium from red alga, can produce agarases and
liquefy solid plating medium efficiently when agar
was used as coagulant. Here we report the draft genome sequence of strain
HQM9, which should be a novel species of Flavobacteriaceae, and initial
findings from its preliminary analysis.

<>

<1>Dua, A., Sangwan, N., Kaur, J., Saxena, A., Kohli, P., Gupta, A.K., Lal, R.
<2>Draft Genome Sequence of Agrobacterium sp. Strain UHFBA-218, Isolated from Rhizosphere Soil of Crown Gall-Infected Cherry Rootstock Colt.
<3>Genome Announcements
<4>1
<5>e00302-13
<6>2013
<7>We report here the draft genome sequence of the alphaproteobacterium Agrobacterium sp. strain
UHFBA-218, which was isolated from rhizosphere soil of
crown gall-infected cherry rootstock Colt. The draft genome of strain UHFBA-218
consists of 112 contigs (5,425,303 bp) and 5,063 coding sequences with a G+C
content of 59.8%.

<>

<1>Duan, C., Tong, Y., Huang, Y., Wang, X., Xiong, X., Wen, B.
<2>Complete Genome Sequence of Rickettsia heilongjiangensis, an Emerging Tick-Transmitted Human Pathogen.
<3>J. Bacteriol.
<4>193
<5>5564-5565
<6>2011
<7>Rickettsia heilongjiangensis is an emerging tick-transmitted human pathogen causing
far-Eastern spotted fever. Here we report the complete
sequence and the main features of the genome of R. heilongjiangensis
(strain 054).

<>

<1>Duan, X., Gimble, F.S., Quiocho, F.A.
<2>Crystal structure of PI-SceI, a homing endonuclease with protein splicing activity.
<3>Cell
<4>89
<5>555-564
<6>1997
<7>PI-SceI is a bifunctional yeast protein that propagates its mobile gene by catalyzing protein
splicing and site-specific DNA double-strand cleavage.  Here, we report the 2.4 A crystal
structure of the PI-SceI protein.  The structure is composed of two separate domains (I and
II) with novel folds and different functions.  Domain I, which is elongated and formed largely
from seven beta sheets, harbors the N and C termini residues and two His residues that are
implicated in protein splicing.  Domain II, which is compact and is primarily composed of two
similar alpha/beta motifs related by local two-fold symmetry, contains the putative nuclease
active site with a cluster of two acidic residues and one basic residue commonly found in
restriction endonucleases.  This report presents prototypic structures of domains with single
endonuclease and protein splicing active sites.

<>

<1>Duan, Y., Wilkosz, P., Rosenberg, J.M.
<2>Dynamic contributions to the DNA binding entropy of the EcoRI and EcoRV restriction endonucleases.
<3>J. Mol. Biol.
<4>264
<5>546-555
<6>1996
<7>Molecular dynamics simulations on DNA-EcoRI and DNA-EcoRV complexes suggest that the DNA
within these complexes is significantly more ordered than free DNA.  Similarly, both the
protein and the DNA are more ordered in the specific (cognate) DNA-EcoRV complex than they are
in the non-cognate DNA-protein complex, consistent with recently proposed analogies between
protein folding and sequence-specific DNA-protein recognition.  Analysis of the trajectories
shows that the net entropy gain upon specific binding to be the result of opposing
contributions.  Solvent release, which increases entropy versus configurational terms (as
measured by the magnitude of the atomic fluctuations), and collective terms from tight
coupling between the motions of the protein and the DNA.

<>

<1>Dubey, A.K., Bhattacharya, S.K.
<2>Angle and locus of the bend induced by the MspI DNA methyltransferase in a sequence-specific complex with DNA.
<3>Nucleic Acids Res.
<4>25
<5>2025-2029
<6>1997
<7>Bending of DNA induced by M.MspI, one of the m5C-DNA methyltransferases, has been investigated
using circular permutation analysis.  The M.MspI Mtase induced sharp bends in DNA containing
its recognition sequence 5'-CCGG-3' which was estimated to be 142 +/- 4 degrees and 132 +/-
4 degrees for circularly permuted DNA fragments of 127 and 1459 bp respectively.  The bend
center was found to be asymmetric with respect to the CCGG sequence and appeared to exclude
the 'target cytosine'.  An estimate of -15 kcal/mol was obtained for the free energy
associated with M.MspI-induced DNA bending.

<>

<1>Dubey, A.K., Bisaria, V.S., Mukhopadhyay, S.N., Ghose, T.K.
<2>Stabilization of restriction endonuclease BamHI by cross-linking reagents.
<3>Biotechnol. Bioeng.
<4>33
<5>1311-1316
<6>1989
<7>Bacillus amyloliquefaciens H produces a restriction endonuclease BamHI which is
heat labile even at low temperatures.  Studies were conducted to enhance
thermal stability of BamHI using cross-linking reagents, namely,
glutaraldehyde, dimethyl adipimidate (DMA), dimethyl suberimidate (DMS), and
dimethyl 3,3'-dithiobispropionimidate (DTBP).  Reaction with glutaraldehyde did
not result in a preparation with enhanced thermal stability.  However, the
DMA-, DMS-, and DTBP-cross-linked preparations of BamHI exhibited significant
improvement in thermal stability.  Studies on thermal denaturation of the
cross-linked enzyme preparations revealed that these do not follow a true
first-order kinetics.  A possible deactivation scheme has been proposed in
which the enzyme has been envisaged to go through a fully active but more
susceptible transient state which, on prolonged heat exposure, exhibits a
first-order decay kinetics.  At 35C, which is close to the optimum reaction
temperature of 37C for BamHI activity, the half-life of DMA-, DMS-, and
DTBP-cross-linked preparations were 4.0, 5.25, and 5.5 h, respectively, whereas
the native enzyme exhibited a half-life of 1.2 h only.  The apparent values of
deactivation rate constants for native, DMA-, DMS-, and DTBP-cross-linked BamHI
were 1.13, 0.39, 0.29, and 0.26 h-1, respectively, at the same temperature, and
the apparent values of activation energies for denaturation of native, DMA-,
DMS-, and DTBP-cross-linked BamHI were 2.63, 5.24, 6.55, and 9.2 kcal/mol,
respectively.  The DTBP-cross-linked BamHI was, therefore, the best heat-stable
preparation among those tested.  The unusually low values of activation
energies for denaturation of BamHI show their highly thermolabile nature
compared to other commonly encountered enzymes such as trypsin, having
activation energies of more than 40 kcal/mol for their denaturation.

<>

<1>Dubey, A.K., Mollet, B., Roberts, R.J.
<2>Purification and characteriation of the MspI DNA methyltransferase cloned and overexpressed in E. coli.
<3>Nucleic Acids Res.
<4>20
<5>1579-1585
<6>1992
<7>The MspI restriction-modification system, which recognizes the sequence 5'-CCGG-3', has been
previously cloned and sequenced. We subcloned the methyltransfearse gene (M.MspI) downstream
of the ptac promoter in the multicopy vector pUC119 and overexpressed it in E. coli. Upon
induction with IPTG, M.MspI constitutes more than 10% of cellular protein. A scheme has been
devised to purify large amounts of biologically active M.MspI to apparent homogeneity from
these overexpressing E. coli cells. Approximately 0.8 mg of pure M.MspI per gram of cells (wet
weight) can be obtained. The apparent molecular weight of M.MspI is 49 kD, by SDS gel
electrophoresis and 48-54 kD by gel filtration. At low concentrations (less than 0.4 mg/ml),
the methyltransferase is a monomer in solution but at higher concentrations (greater than 3.0
mg/ml) it exists predominantly as a dimer. Polyclonal antibodies raised against M.MspI
cross-react with the DNA-methyltransferases of several other restriction-modification systems.

<>

<1>Dubey, A.K., Mukhopadhyay, S.N., Bisaria, V.S., Ghose, T.K.
<2>Sources, production, and purification of restriction enzymes.
<3>Process. Biochem.
<4>22
<5>25-34
<6>1987
<7>At present restriction enzymes are low volume, high value microbial products
but information on their production, downstream processing, stability
characteristics and applications have not been discussed from a
biotechnological view point.  In this article an attempt has been made to bring
out relevant information on restriction enzymes in collated form.

<>

<1>Dubey, A.K., Roberts, R.J.
<2>Sequence-specific DNA binding by the MspI DNA methyltransferase.
<3>Nucleic Acids Res.
<4>20
<5>3167-3173
<6>1992
<7>The MspI methyltransferase (M.MspI) recognizes the sequence CCGG and catalyzes the formation
of 5-methylcytosine at the first C-residue. We have investigated the sequence-specific
DNA-binding properties of M.MspI under equilibrium conditions, using gel-mobility shift assays
and DNaseI footprinting. M.MspI binds to DNA in a sequence-specific manner either alone or in
the presence of the normal methyl donor S-adenosyl-L-methionine as well as the analogues,
sinefungin and S-adenosyl-L-homocysteine. In the presence of S-adenosyl-L-homocysteine, M.MspI
shows the highest binding affinity to DNA containing a hemimethylated recognition sequence
(Kd=3.6x10-7M), but binds less well to unmethylated DNA (Kd=8.3x10-7M). Surprisingly it shows
specific, although poor, binding to fully methylated DNA (Kd=4.2x10-6M). M.MspI binds
approximately 5-fold more tightly to DNA containing its recognition sequence, CCGG, than to
nonspecific sequences in the absence of cofactors. In the presence of S-adenosyl-L-methionine,
S-adenosyl-L-homocysteine or sinefungin the discrimination between specific and nonspecific
sequences increases up to 100-fold. DNaseI footprinting studies indicate that 16 base pairs of
DNA are covered by M.MspI, with the recognition sequence CCGG located asymmetrically within
the footprint.

<>

<1>Duchateau, P., Grizot, S., Paques, F.
<2>Meganuclease engineering.
<3>BIOFUTUR
<4>0
<5>38-42
<6>2008
<7>
<>

<1>Duchateau, P., Paques, F.
<2>New heterodimeric meganuclease, useful for inducing a double-strand nucleic acid break in target DNA sequence, for preventing, improving or curing a genetic disease or a disease caused by an infectious agent - involving vector-mediated gene transfer.
<3>International Patent Office
<4>WO 200697854
<5>
<6>2006
<7>DERWENT ABSTRACT: NOVELTY - A heterodimeric meganuclease comprising two domains of different
meganucleases, where the domains are in two
separate polypeptides which are able to assemble and to cleave a
chimeric DNA target sequence comprising one different half of each
parent meganuclease DNA target sequence, is new. DETAILED DESCRIPTION -
INDEPENDENT CLAIMS are also included for: (1) a recombinant vector
comprising two polynucleotide fragments, each encoding a different
polypeptide; (2) a host cell comprising two polynucleotide fragments or
a vector; (3) a non-human transgenic animal comprising two
polynucleotide fragments; (4) a transgenic plant comprising two
polynucleotide fragments; (5) use of a heterodimeric meganuclease, two
polynucleotides, a vector, a host cell, a transgenic plant, or a
non-human transgenic mammal, for molecular biology, for in vivo or in
vitro genetic engineering, and for in vivo or in vitro genome
engineering; (6) genetic engineering comprising a step of double-strand
nucleic acid breaking in a site of interest; (7) genome engineering
comprising double-strand breaking a genomic locus comprising a chimeric
DNA target of a heterodimeric meganuclease;(8) a composition comprising
a heterodimeric meganuclease, two polynucleotides, or a vector; and (9)
use of a heterodimeric meganuclease, two polynucleotides, or a vector,
for (i) the preparation of a medicament for preventing, improving or
curing a genetic disease or a disease caused by an infectious agent
that presents a DNA intermediate, in an individual, the medicament is
administrated by any means to the individual, or (ii) in vitro, for
inhibiting the propagation, inactivating or deleting an infectious
agent that presents a DNA intermediate, in biological derived products
or products intended for biological uses or for disinfecting an object.
BIOTECHNOLOGY - Preferred Sequences: Each polypeptide comprises the
LAGLIDADG Homing Endonuclease Core Domain of a different LAGLIDADG
homing endonuclease or its variant. The LAGLIDADG homing endonuclease
is a homodimeric enzyme, where the homodimeric LAGLIDADG homing
endonuclease is I-CreI. The LAGLIDADG homing endonuclease is a
monomeric enzyme, where the monomeric LAGLIDADG homing endonuclease is
I-DmoI. One of the polypeptide comprises the LAGLIDADG Homing
Endonuclease Core Domain of an I-CreI variant having a substitution in
positions 44, 68, and/or 70 of I-CreI, where the residues in positions
44, 68, and/or 70 of I-CreI are replaced with an amino acid selected
from A, D, E, G, H, K, N, P, Q, R, S, T, or Y. The I-CreI variant
comprises the replacement of the aspartic acid in position 75 with an
uncharged amino acid, where the uncharged residue is an asparagine or a
valine. The heterodimeric meganuclease cleaves a chimeric DNA target
comprising the sequence of SEQ ID NO: 2. One of the polypeptides has a
glutamine (Q) in position 44 of I-CreI for cleaving a chimeric DNA
target, wheren -4 is t or n+4 is a. One of the polypeptide has an
alanine (A) or an asparagine in position 44 of I-CreI for cleaving a
chimeric DNA target, where n-4 is a or n+4 is t. One of the
polypeptides has a lysine (K) in position 44 of I-CreI for cleaving a
chimeric DNA target, where n-4 is c or n+4 is g. Preferred Recombinant
Vector: The recombinant vector includes a targeting construct
comprising sequences sharing homologies with the region surrounding the
chimeric DNA target sequence, where the targeting construct comprises
(i) sequences sharing homologies with the region surrounding the
chimeric DNA target sequence, and (ii) sequences to be introduced
flanked by sequence as in (i). Preferred Method: Genetic engineering
comprises double-strand nucleic acid breaking in a site of interest
located on a vector comprising a chimeric DNA target of a heterodimeric
meganuclease, by contacting the vector with a heterodimeric
meganuclease, therefore inducing a homologous recombination with
another vector presenting homology with the sequence surrounding the
cleavage site.

<>

<1>Duchateau, P., Paques, F.
<2>Novel heterodimeric meganuclease, useful for preventing, improving or curing a genetic disease and a disease caused by an infectious agent such as virus - involving vector-mediated gene transfer and expression in Saccharomyces cerevisiae for use in gen.
<3>International Patent Office
<4>WO 200734262
<5>
<6>2007
<7>NOVELTY - A heterodimeric meganuclease comprising two domains of different meganucleases,
where the domains are in two
separate polypeptides which are able to assemble and to cleave a
chimeric DNA target sequence comprising one different half of each
parent meganuclease DNA target sequence, is new. DETAILED DESCRIPTION -
INDEPENDENT CLAIMS are included for the following: (1) a recombinant
vector comprising two polynucleotide fragments, each encoding the
different polypeptides; (2) a host cell comprising the polynucleotide
present in the vector; (3) a non-human transgenic animal comprising the
polynucleotide fragments present in the vector; (4) a transgenic plant
comprising two polynucleotide fragments present in the vector; and (5)
a composition comprising the heterodimeric meganuclease, and the
polynucleotides present in the vector. BIOTECHNOLOGY - Preparation: The
heterodimeric meganuclease is prepared by standard recombinant methods
(disclosed). Preferred Heterodimeric Meganuclease: The each polypeptide
comprises the (S1) homing endonuclease core domain of a different (S1)
homing endonuclease or its variant. The (S1) homing endonuclease is a
homodimeric or monomeric enzyme. The homodimeric (S1) homing
endonuclease is I-CreI. The monomeric (S1) homing endonuclease is
I-DmoI. The polypeptide comprises the (S1) homing endonuclease core
domain of an I-CreI variant having at least one substitution in
positions 44, 68, and/or 70 of I-CreI. The residues in positions 44,
68, and/or 70 of I-CreI are replaced with an amino acid chosen from
Ala, Asp, Glu, Gly, His, Lys, Asp, Pro, Gln, Arg, Ser, Thr, Asp, Glu,
His, Lys, His, Gln, Arg, Ser, Thr, Tyr. The I-CreI variant comprises
the replacement of the aspartic acid in position 75 with an uncharged
amino acid. The uncharged residue is an asparagine or a valine. The
heterodimeric meganuclease cleaves chimeric DNA target comprising the
sequence (SEQ ID No. 2). The polypeptide has (a) a Gln in position 44
of I-CreI for cleaving a chimeric DNA target, where n-4 is Thr or n+4
is Ala, (b) an alanine or an asparagine in position 44 of I-CreI for
cleaving a chimeric DNA target, where n-4 is Ala or n+4 is Thr, or (c)
a lysine in position 44 of I-CreI for cleaving a chimeric DNA target,
where n-4 is Cys or n+4 is Gly. Preferred Recombinant Vector: The
recombinant vector includes a targeting construct comprising sequences
sharing homologies with the region surrounding the chimeric DNA target
sequence. The targeting construct comprises (a) sequences sharing
homologies with the region surrounding the chimeric DNA target
sequence, and (b) sequences to be introduced flanked by the sequence as
in (a). Preferred Composition: The composition further comprises a
targeting DNA construct having the sequence which repairs the site of
interest flanked by sequences sharing homologies with the targeted
locus. Leu-Ala-Gly-Leu-Ile-Asp-Ala-Asp-Gly (S1), and c-11a-10a-
9a-8a-7c-6n-5n-4n-3n-2n-1n+1n+2n+3n+4n+5g+6t+7t+8t+9t+10g+11 (SEQ ID
No. 2), where n is a, t, c or g. ACTIVITY - Antimicrobial; Virucide. No
biological data given. MECHANISM OF ACTION - Nuclease agonist; Gene
therapy. USE - For genetic engineering, involves double-strand nucleic
acid breaking in a site of interest located on a vector comprising a
chimeric DNA target of the heterodimeric meganuclease, by contacting
the vector with the heterodimeric meganuclease, thus inducing a
homologous recombination with another vector presenting homology with
the sequence surrounding the cleavage site of the heterodimeric
meganuclease and maintaining the broken genomic locus appropriate for
homologous recombination with a targeting DNA construct comprising the
sequence to be introduced in the locus, flanked by sequences sharing
homologies with the targeted locus. The heterodimeric meganuclease or
the polynucleotides presenting the vector for (a) preventing, improving
or curing a genetic disease in an individual or a disease caused by an
infectious agent.

<>

<1>Duchateau, P., Valton, J., Daboussi, F.
<2>CpG methylation inhibition of rare-cutter endonucleases and methylation-insensitive variants of a LAGLIDADG meganuclease.
<3>International Patent Office
<4>WO 201201527 A
<5>
<6>2012
<7>The present invention concerns novel methods for improving cleavage of DNA by rare-cutting
endonucleases, overcoming DNA modification constraints, particularly DNA methylation, thereby
giving new tools for genome engineering, particularly to increase the integration efficiency
of a transgene into a genome at a predetermined location, including therapeutic applications
and cell line engineering.

<>

<1>Duchaud, E. et al.
<2>The genome sequence of the entomopathogenic bacterium Photorhabdus luminescens.
<3>Nat. Biotechnol.
<4>21
<5>1307-1313
<6>2003
<7>Photorhabdus luminescens is a symbiont of nematodes and a broad-spectrum insect pathogen. The
complete genome sequence of strain TT01 is 5,688,987
base pairs (bp) long and contains 4,839 predicted protein-coding genes.
Strikingly, it encodes a large number of adhesins, toxins, hemolysins,
proteases and lipases, and contains a wide array of antibiotic
synthesizing genes. These proteins are likely to play a role in the
elimination of competitors, host colonization, invasion and bioconversion
of the insect cadaver, making P. luminescens a promising model for the
study of symbiosis and host-pathogen interactions. Comparison with the
genomes of related bacteria reveals the acquisition of virulence factors
by extensive horizontal transfer and provides clues about the evolution of
an insect pathogen. Moreover, newly identified insecticidal proteins may
be effective alternatives for the control of insect pests.

<>

<1>Duchaud, E., Boussaha, M., Loux, V., Bernardet, J.F., Michel, C., Kerouault, B., Mondot, S., Nicolas, P., Bossy, R., Caron, C., Bessieres, P., Gibrat, J.F., Claverol, S., Dumetz, F., Henaff, M.L., Benmansour, A.
<2>Complete genome sequence of the fish pathogen Flavobacterium psychrophilum.
<3>Nat. Biotechnol.
<4>25
<5>763-769
<6>2007
<7>We report here the complete genome sequence of the virulent strain JIP02/86 (ATCC 49511) of
Flavobacterium psychrophilum, a widely
distributed pathogen of wild and cultured salmonid fish. The genome
consists of a 2,861,988-base pair (bp) circular chromosome with 2,432
predicted protein-coding genes. Among these predicted proteins, stress
response mediators, gliding motility proteins, adhesins and many putative
secreted proteases are probably involved in colonization, invasion and
destruction of the host tissues. The genome sequence provides the basis
for explaining the relationships of the pathogen to the host and opens new
perspectives for the development of more efficient disease control
strategies. It also allows for a better understanding of the physiology
and evolution of a significant representative of the family
Flavobacteriaceae, whose members are associated with an interesting
diversity of lifestyles and habitats.

<>

<1>Duchaud, E., Taourit, S., Glaser, P., Frangeul, L., Kunst, F., Danchin, A., Buchrieser, C.
<2>Sequence of the Photorhabdus luminescens strain TT01 genome and uses.
<3>International Patent Office
<4>WO 02094867 A
<5>
<6>2002
<7>The invention concerns the genomic sequence and nucleotide sequences coding for polypeptides
or Photorhabdus luminescens, such as polypeptides involved in operons of biosynthesis of
antibiotics, or of toxins, or polypeptides exhibiting a toxin or antibiotic type activity
capable of being used as pesticide, bactericide or fungicide as well as vectors comprising
said sequences and cells or animals transformed by said vectors.

<>

<1>Duda, E.G., Izsvak, Z., Orosz, A.
<2>Isolation and purification of CeqI endonuclease, an isoschizomer of EcoRV.
<3>Nucleic Acids Res.
<4>15
<5>1334
<6>1987
<7>None

<>

<1>Dudnik, A., Dudler, R.
<2>Genome and Transcriptome Sequences of Pseudomonas syringae pv. syringae B301D-R.
<3>Genome Announcements
<4>2
<5>e00306-14
<6>2014
<7>Strains of the plant pathogen Pseudomonas syringae are commonly found in the phylosphere and
are able to infect a number of agriculturally important crops. Here, we report a high-quality
draft genome sequence of Pseudomonas syringae pv. syringae B301D-R, isolated from pears, which
is a model strain for phytotoxin research in P. syringae.

<>

<1>Dudnik, A., Dudler, R.
<2>Non contiguous-finished genome sequence of Pseudomonas syringae pathovar syringae strain B64 isolated from wheat.
<3>Standards in Genomic Sciences
<4>8
<5>420-429
<6>2013
<7>The Gram-negative gammaproteobacterium Pseudomonas syringae is one of the most wide-spread
plant pathogens and has been repeatedly reported to cause significant
damage to crop plantations. Research on this pathogen is very intensive, but most
of it is done on isolates that are pathogenic to Arabidopsis, tomato, and bean.
Here, we announce a high-quality draft genome sequence of Pseudomonas syringae
pv. syringae B64 which is the first published genome of a P. syringae strain
isolated from wheat up to date. The genome sequence will assist in gaining
insights into basic virulence mechanisms of this pathogen which has a relatively
small complement of type III effectors.

<>

<1>Dudnik, A., Dudler, R.
<2>High-Quality Draft Genome Sequence of Pseudomonas syringae pv. Syringae Strain SM, Isolated from Wheat.
<3>Genome Announcements
<4>1
<5>e00610-13
<6>2013
<7>Pseudomonas syringae is one of the most widespread plant pathogens that can cause significant
damage to crop plantations. Here, we announce a noncontiguous
finished genome sequence of Pseudomonas syringae pv. syringae strain SM, isolated
from hexaploid wheat. The genome sequence revealed the smallest described
complement of type III effectors.

<>

<1>Dueger, E.L., House, J.K., Heithoff, D.M., Mahan, M.J.
<2>Salmonella DNA adenine methylase mutants elicit protective immune responses to homologous and heterologous serovars in chickens.
<3>Infect. Immun.
<4>69
<5>7950-7954
<6>2001
<7>Salmonella DNA adenine methylase (Dam) mutants that lack or
overproduce Dam are highly attenuated for virulence in mice and confer
protection against murine typhoid fever. To determine whether vaccines
based on Dam are efficacious in poultry, a Salmonella Dam-vaccine was
evaluated in the protection of chicken broilers against oral challenge
with homologous and heterologous Salmonella serovars. A Salmonella
enterica serovar Typhimurium Dam-vaccine strain was
attenuated for virulence in day-of-hatch chicks more than 100,000-fold.
Vaccination of chicks elicited cross-protective immune responses, as
evidenced by reduced colonization (10- to 10,000-fold) of the
gastrointestinal tract (ileum, cecum, and feces) and visceral organs
(bursa and spleen) after challenge with homologous (Typhimurium F98)
and heterologous (Enteritidis 4973 and S. enterica
O6,14,24: e,h-monophasic) Salmonella serovars that are
implicated in Salmonella infection of poultry. The
protection conferred was observed for the organ or the maximum
CFU/tissue/bird as a unit of analysis, suggesting that Dam mutant
strains may serve as the basis for the development of efficacious
poultry vaccines for the containment of Salmonella.

<>

<1>Dueger, E.L., House, J.K., Heithoff, D.M., Mahan, M.J.
<2>Salmonella DNA adenine methylase mutants prevent colonization of newly hatched chickens by homologous and heterologous serovars.
<3>Int. J. Food Microbiol.
<4>80
<5>153-159
<6>2003
<7>Salmonella mutants lacking DNA adenine methylase (Dam) are highly attenuated for virulence and
confer protection against oral challenge with
homologous and heterologous Salmonella serovars in mice and chicken
broilers. To determine whether vaccines based on Dam are efficacious in
preventing early colonization of newly hatched chickens, a Salmonella
typhimurium Dam(-) vaccine was evaluated for the protection of chicks
against oral challenge with homologous and heterologous Salmonella
serovars. Vaccination of chicks elicited protection 2 and 6 days
post-challenge as evidenced by a significant reduction in colonization of
the gastrointestinal tract (ileum, cecum and feces) and visceral organs
(spleen and bursa) when challenged with homologous S. typhimurium.
Moderate protection was observed following challenge with heterologous S.
enteritidis and Salmonella O6, 14, 24:e, h-monophasic) serovars. These
data suggest that Salmonella Dam mutant strains conferred
cross-protection, presumably via competitive exclusion mechanisms that
prevent superinfection of chicks by other Salmonella strains. Such
protection may reduce pre-harvest Salmonella contamination in poultry,
decreasing the potential for food-borne transmission of this pathogen to
humans.

<>

<1>Dueger, E.L., House, J.K., Heithoff, D.M., Mahan, M.J.
<2>Salmonella DNA adenine methylase mutants elicit early and late onset protective immune responses in calves.
<3>Vaccine
<4>21
<5>3249-3258
<6>2003
<7>Salmonellosis is an important disease of livestock and Salmonella contamination of
livestock-derived food products and effluents pose a
significant risk to human health. Salmonella vaccines currently available
to prevent salmonellosis in cattle have limited efficacy. Here we
evaluated a Salmonella enterica serovar Typhimurium vaccine strain lacking
the DNA adenine methylase (Dam) for safety and efficacy in calves.
Vaccination was safe in calves, and following challenge with virulent
Typhimurium 4 weeks post-immunization, vaccinated animals exhibited
significantly lower mortality, diarrhea, and rectal temperatures, as well
as reduced colonization of gastrointestinal tract and visceral organs
compared to non-vaccinated control animals. Additionally, early onset
protection (competitive exclusion) in vaccinated neonatal calves was
demonstrated by attenuated clinical disease (as measured by rectal
temperatures and attitude scores) and reduced mortality when challenged
with virulent Typhimurium 24h after immunization. Taken together, these
data suggest that vaccination with Salmonella Dam mutant strains confer
significant protection against Salmonella infections in cattle via both
adaptive immunity and competitive exclusion mechanisms.

<>

<1>Dueholm, M.S., Albertsen, M., D'Imperio, S., Tale, V.P., Lewis, D., Nielsen, P.H., Nielsen, J.L.
<2>Complete Genome of Rhodococcus pyridinivorans SB3094, a Methyl-Ethyl-Ketone-Degrading Bacterium Used for Bioaugmentation.
<3>Genome Announcements
<4>2
<5>e00525-14
<6>2014
<7>Here, we present the complete genome of Rhodococcus pyridinivorans SB3094, a
methyl-ethyl-ketone (MEK)-degrading strain used for bioaugmentation relating to
the treatment of wastewater contamination with petrochemical hydrocarbons. The
genome highlights important features for bioaugmentation, including the genes
involved in the degradation of MEK.

<>

<1>Dueholm, M.S., Danielsen, H.N., Nielsen, P.H.
<2>Complete Genome Sequence of Pseudomonas sp. UK4, a Model Organism for Studies of  Functional Amyloids in Pseudomonas.
<3>Genome Announcements
<4>2
<5>e00898-14
<6>2014
<7>Here, we present the complete genome of Pseudomonas sp. UK4. This bacterium was the first
Pseudomonas strain shown to produce functional amyloids, and it
represents a model organism for studies of functional amyloids in Pseudomonas
(Fap).

<>

<1>Duff, M.K., Davies, J.K.
<2>Multiple restriction-modification systems in Neisseria gonorrhoeae.
<3>Gene
<4>74
<5>227-228
<6>1988
<7>Meeting Abstract

<>

<1>Duff, M.K., Davies, J.K.
<2>DNA enzymes of Neisseria gonorrhoeae.
<3>Gonococci and Meningococci, Kluwer Academic Publishers, Poolman, J.T., Zanen, H.C., Meyer, T.F., Heckels, J.E., Makela, P.R.H., Dordrecht, The Netherlands
<4>
<5>251-256
<6>1988
<7>Two gonococcal strains, the Dam+ strain D109, and the Dam- strain D211, were investigated for
enzyme content. Enzymes were obtained from surface washes, by sonication and by osmotic shock.
They were partially purified by precipitation and heparin-sepharose affinity chromatography.
Endonuclease activity was assayed by using both dam-methylated and nonmethylated
bacteriophage lambda DNA.  Two endonuclease activities that have not been previously described
in the gonococcus were discovered.  One, designated R.NgoIX, is an isoschizomer of DpnI. [The
enzyme named NgoIX has been renamed NgoDXVII. Enzymes with the specificity NgoX are mixtures
of NgoI and NgoVIII specificities.]

<>

<1>Dufresne, A. et al.
<2>Genome sequence of the cyanobacterium Prochlorococcus marinus SS120, a nearly minimal oxyphototrophic genome.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>10020-10025
<6>2003
<7>Prochlorococcus marinus, the dominant photosynthetic organism in the ocean, is found in two
main ecological forms: high-light-adapted genotypes
in the upper part of the water column and low-light-adapted genotypes at
the bottom of the illuminated layer. P. marinus SS120, the complete genome
sequence reported here, is an extremely low-light-adapted form. The genome
of P. marinus SS120 is composed of a single circular chromosome of
1,751,080 bp with an average G+C content of 36.4%. It contains 1,884
predicted protein-coding genes with an average size of 825 bp, a single
rRNA operon, and 40 tRNA genes. Together with the 1.66-Mbp genome of P.
marinus MED4, the genome of P. marinus SS120 is one of the two smallest
genomes of a photosynthetic organism known to date. It lacks many genes
that are involved in photosynthesis, DNA repair, solute uptake,
intermediary metabolism, motility, phototaxis, and other functions that
are conserved among other cyanobacteria. Systems of signal transduction
and environmental stress response show a particularly drastic reduction in
the number of components, even taking into account the small size of the
SS120 genome. In contrast, housekeeping genes, which encode enzymes of
amino acid, nucleotide, cofactor, and cell wall biosynthesis, are all
present. Because of its remarkable compactness, the genome of P. marinus
SS120 might approximate the minimal gene complement of a photosynthetic
organism.

<>

<1>Dugaiczyk, A., Boyer, H.W., Goodman, H.M.
<2>Ligation of EcoRI endonuclease-generated DNA fragments into linear and circular structures.
<3>J. Mol. Biol.
<4>96
<5>171-184
<6>1975
<7>Double-stranded DNA fragments terminated at their 5'-ends by the
single-stranded sequence pA-A-T-T-, generated by digestion of DNA with EcoRI
restriction endonuclease, were ligated with Escherichia coli polynucleotide
ligase under various conditions of temperature, concentration and time.  The
linear and circular products of ligation were separated by electrophoresis in
agarose gel and quantitated by densitometry.  The rate of ligation of
(EcoRI-cleaved) simian virus (SV40) DNA at a concentration of 100 microgram/ml
increased from 0C to 5C to 10C (6-fold increase overall); raising the
temperature to 15C did not further increase the rate of ligation.  At the
appropriate DNA concentrations, the predominant products of ligation are either
linear concatemers that are integral multimers of the starting DNA fragment, or
covalently closed circular structures of the monomeric DNA fragment.  Ligating
a mixture of two different length DNA fragments gives rise to all of the
possible expected recombinant molecules.  Linear or circular products of
ligation were predicted by consideration of the total concentration of DNA
termini, i, and the local concentration of one terminus in the neighborhood of
the other on the same DNA molecule, j.  The parameter j is a function of the
length of a DNA molecule, providing this length is greater than the random coil
segment of DNA.  Experimentally it was found that circular structures are
formed in significant amounts only under conditions when the value of j is
several times greater than that of i.  When j = i, equal amounts of linear and
circular products would be expected, but most of the molecules were ligated
into linear concatemers.  No circular structure of a DNA fragment whose contour
length l (6 x 10-2 microm) is smaller than the random coil segment value b
(7.17 x 10-2 microm) was observed, while circular structures of the dimer of
the same molecule (12 x 10-2 microm) were detected.

<>

<1>Dugaiczyk, A., Hedgpeth, J., Boyer, H.W., Goodman, H.M.
<2>Physical identity of the SV40 deoxyribonucleic acid sequence recognized by the EcoRI restriction endonuclease and modification methylase.
<3>Biochemistry
<4>13
<5>503-512
<6>1974
<7>The EcoRI modification methylase introduces two methyl groups into one SV40 DNA
molecule.  The only base methylated has been identified as N6-methyladenine.
Both of the methyl groups are introduced into the same fragment (designated F,
about 400 base pairs long) of a HindII endonuclease digest of SV40 DNA.  The
EcoRI endonuclease makes one double-strand cleavage in SV40 DNA.  The site of
this cleavage is also contained within the F fragment.  Analysis of
dinucleoside monophosphates, trinucleoside diphosphates, and tetranucleoside
triphosphates generated by partial digestion of the methylated DNA with
pancreatic DNase I gives the following sequence of nucleotides at the site of
methylation by the EcoRI methylase:  GpApm6ApTpTpC.  This sequence (with A in
place of m6A) is also found at the site of phosphodiester-bond cleavage by the
EcoRI restriction endonuclease.  Using [c-32P]rATP and polynucleotide kinase,
SV40 DNA has been labeled in each strand with 32P specifically at the
phosphodiester bonds cleaved by the EcoRI endonuclease.  The DNA was
polymerized at 4o by hydrogen bonding of the cohesive termini of the EcoRI
endonuclease break.  The labeled 5'-monophosphates at the staggered
single-strand breaks were esterified with the adjacent 3'-hydroxyl groups by
polynucleotide ligase at low temperature, and the covalently polymerized DNA
was methylated by the EcoRI modification methylase using
S-adenosyl-L-[methyl-3H]methionine.  Analysis of the radioactive labels in the
mono- and dinucleotides from a partial digest of this double-labeled DNA
identifies physically the same sequence of base pairs in SV40 DNA as the
substrate site for the EcoRI endonuclease and for the EcoRI modification
methylase.

<>

<1>Dugaiczyk, A., Kimball, M., Linn, S., Goodman, H.M.
<2>Location and nucleotide sequence of the site on SV40 DNA methylated by the EcoB modification methylase.
<3>Biochem. Biophys. Res. Commun.
<4>61
<5>1133-1140
<6>1974
<7>The modification methylase from Escherichia coli strain B, EcoB, introduces two
methyl groups into one SV40 DNA molecule.  The only base methylated has been
identified as N6-methyladenine.  Digestion of the methylated SV40 DNA with two
restriction endonucleases, one from Hemophilus influenzae, HindIII, and another
from Hemophilus aegyptius, Hae, locates the methyl groups in the same DNA
fragment, about 250 base pairs long, between 0.860 and 0.907 fractional lengths
of SV40 DNA clockwise from the EcoRI site on the circular SV40 geneome.
Analysis of dinucleoside monophosphates, trinucleoside diphosphates, and
tetranucleoside triphosphates, generated by partial digestion of the methylated
DNA with pancreatic DNaseI gave the following sequences of nucleotides at the
site(s) of methylation by the EcoB methylase: 5'...C-m6A-G-C-T...3' and
5'...T-G-m6A-A...3' These cannot be incorporated into a simple sequence with
2-fold rotational symmetry.

<>

<1>Duhaime, M.B., Wichels, A., Waldmann, J., Teeling, H., Glockner, F.O.
<2>Ecogenomics and genome landscapes of marine Pseudoalteromonas phage H105/1.
<3>ISME J.
<4>5
<5>107-121
<6>2011
<7>Marine phages have an astounding global abundance and ecological impact.
However, little knowledge is derived from phage genomes, as most of the
open reading frames in their small genomes are unknown, novel proteins. To
infer potential functional and ecological relevance of sequenced marine
Pseudoalteromonas phage H105/1, two strategies were used. First,
similarity searches were extended to include six viral and bacterial
metagenomes paired with their respective environmental contextual data.
This approach revealed 'ecogenomic' patterns of Pseudoalteromonas phage
H105/1, such as its estuarine origin. Second, intrinsic genome signatures
(phylogenetic, codon adaptation and tetranucleotide (tetra) frequencies)
were evaluated on a resolved intra-genomic level to shed light on the
evolution of phage functional modules. On the basis of differential codon
adaptation of Phage H105/1 proteins to the sequenced Pseudoalteromonas
spp., regions of the phage genome with the most 'host'-adapted proteins
also have the strongest bacterial tetra signature, whereas the least
'host'-adapted proteins have the strongest phage tetra signature. Such a
pattern may reflect the evolutionary history of the respective phage
proteins and functional modules. Finally, analysis of the structural
proteome identified seven proteins that make up the mature virion, four of
which were previously unknown. This integrated approach combines both
novel and classical strategies and serves as a model to elucidate
ecological inferences and evolutionary relationships from phage genomes
that typically abound with unknown gene content.

<>

<1>Duhring, U., Enke, H., Grundel, M., Lockau, W., Smith, C.R., Woods, P., Ziegler, K., Oesterhelt, C., Coleman, J.R., Kramer, D.
<2>Genetically modified photoautotrophic ethanol producing host cells, method for producing the host cells, constructs for the transformation of the host cells, method for testing a photoautotrophic strain for a desired growth property etc.
<3>International Patent Office
<4>WO 2009098089 A
<5>
<6>2009
<7>In one embodiment the invention provides a genetically modified photoautotrophic, ethanol
producing host cell comprising: - at least one first genetic modification changing the
enzymatic activity or affinity of an endogenous host cell enzyme, - the first genetic
modification resulting in an enhanced level of biosynthesis of acetaldehyde, pyruvate, acetyl
- CoA or precursors thereof compared to the respective wild type host cell, - at least one
second genetic modification different from the first genetic modification comprising an
overexpressed enzyme for the formation of ethanol.

<>

<1>Dujardin, G., Jacq, C., Slonimski, P.P.
<2>Single base substitution in an intron of oxidase gene compensates splicing defects of the cytochrome b gene.
<3>Nature
<4>298
<5>628-632
<6>1982
<7>An extragenic suppressor mutation, mim2-1, which compensates yeast
mitochondrial mutants deficient in splicing of the cytochrome b gene, has
been mapped and sequenced. The mutation is due to a single G leads to A
transition in the long open reading frame of the fourth intron of the
oxidase subunit one gene. It causes the replacement of a glutamic codon by
a lysine codon and the expression of a novel mRNA maturase active in
splicing. Evolution and regulatory connections between homologous introns
of nonhomologous genes are discussed.

<>

<1>Dujon, B.
<2>Sequence of the intron and flanking exons of the mitochondrial 21S rRNA gene of yeast strains having different alleles at the omega and rib-1 loci.
<3>Cell
<4>20
<5>185-197
<6>1980
<7>The complete nucleotide sequence has been determined for the intron, its junctions and the
flanking exon regions of the 21S rRNA gene in three genetically characterized strains
differing by their omega alleles (omega+, omega- and omega n) and by their
chloramphenicol-resistant mutations at the rib-1 locus. Comparison of these DNA sequences
shows that:  omega+ differs from omega- and omega n by the presence of the intron (1143bp), as
well as by a second and unexpected mini-insert (66bp) located 156bp upstream within the exon,
whose nature and functions are still unknown but whose striking palindromic structure may
suggest a mitochondrial transposable element. The two mutations CR321 and CR323 correspond to
two different monosubstitutions, 56bp apart in the omega- and omega n strains but separated by
the intron in the omega+ strains. In relation to previous genetic results, a model is
discussed assuming that the interactions of two different regions or genetic loci determine
the chloramphenicol resistance, one of which contains the omega n mutations. A long
uninterrupted coding sequence able to specify a 235 amino acid polypeptide exists within the
intron. This remarkable observation gives new insight into the origin of the mitochondrial
introns and raises the question of the possible functions of intron-encoded polypeptides.
Finally, sequence comparisons with evolutionarily distant organisms, showing that different
rRNA introns are inserted at different positions of an otherwise highly conserved region of
the gene, suggest a recent insertion of these introns and a mechanism for splicing after the
assembly of the large ribosomal subunit.

<>

<1>Dujon, B.
<2>Homing endonucleases and the yeast mitochondrial omega locus - A historical perspective.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Berlin Heidelberg
<4>16
<5>11-31
<6>2005
<7>It was May 1985.  Outside the laboratory, near Paris, nature was exulting in its colorful
mid-spring glory.  The last technical details and experimental pitfalls had been fixed in the
preceding weeks.  Now, the site-specific endonucleolytic activity of the intron-encoded
protein I that had been carefully engineered to express in Escherichia coli was detectable.
According to my autoradiogram, it was cleaving the intron-less DNA exactly where I expected.
This experiment opened the way to a series of yet unexpected developments, but for me it
concluded a long and rather solitary quest.  Long, because the route that led to the first
intron-encoded homing endonuclease, I-SceI according to the present nomenclature, had started
no less than 15 years before, from a peculiarity of mitochondrial inheritance in yeast.
Solitary, because, over this long period, the phenomenon that led to this discovery had
remained a unique oddity of nature, limiting its interest for many.  Indeed, after the
discovery of I-SceI, it took 3 additional years before the next examples of homing
endonucleases could be identified, suggesting the generality of the phenomenon.  By this time,
the enzymatic properties of I-SceI and its unusual specificity were already characterized, and
it was clear that we were in the presence of a novel class of enzymes.

<>

<1>Dujon, B.
<2>Group I introns as mobile genetic elements: facts and mechanistic speculations--a review.
<3>Gene
<4>82
<5>91-114
<6>1989
<7>Group I introns form a structural and functional group of introns with widespread but
irregular distribution among very diverse organisms and genetic systems. Evidence is now
accumulating that several group I introns are mobile genetic elements with properties similar
to those originally described for the omega system of Saccharomyces cerevisiae: mobile group I
introns encode sequence-specific double-strand (ds) endoDNases, which recognize and cleave
intronless genes to insert a copy of the intron by a ds-break repair mechanism. This mechanism
results in: the efficient propagation of group I introns into their cognate sites; their
maintenance at the site against spontaneous loss; and, perhaps, their transposition to
different sites. The spontaneous loss of group I introns occurs with low frequency by an
RNA-mediated mechanism. This mechanism eliminates introns defective for mobility and/or for
RNA splicing. Mechanisms of intron acquisition and intron loss must create an equilibrium,
which explains the irregular distribution of group I introns in various genetic systems.
Furthermore, the observed distribution also predicts that horizontal transfer of intron
sequences must occur between unrelated species, using vectors yet to be discovered.

<>

<1>Dujon, B., Choulika, A., Colleaux, L., Fairhead, C., Perrin, A., Plessis, A., Thierry, A.
<2>Nucleotide sequence encoding the enzyme I-SceI and the uses thereof.
<3>US Patent Office
<4>US 5474896
<5>
<6>1995
<7>An isolated DNA encoding the enzyme I-SceI is provided.  The DNA sequence can be incorporated
in cloning and expression vectors, transformed cell lines and transgenic animals.  The vectors
are useful in gene mapping and site-directed insertion of genes.

<>

<1>Dujon, B., Choulika, A., Perrin, A., Nicolas, J.-F.
<2>Nucleotide sequence encoding the enzyme I-SceI and the uses thereof.
<3>US Patent Office
<4>US 5792632
<5>
<6>1998
<7>An isolated DNA encoding the enzyme I-SceI is provided.  The DNA sequence can be incorporated
in cloning and expression vectors, transformed cell lines and transgenic animals.  The vectors
are useful in gene mapping and site-directed insertion of genes.

<>

<1>Dujon, B., Choulika, A., Perrin, A., Nicolas, J.-F.
<2>Nucleotide sequence encoding the enzyme I-SceI and the use thereof.
<3>US Patent Office
<4>US 6395959
<5>
<6>2002
<7>An isolated DNA encoding the enzyme I-SceI is provided.  The DNA sequence can be incorporated
in cloning and expression vectors, transformed cell lines and transgenic animals.  The vectors
are useful in gene mapping and site-directed insertion of genes.

<>

<1>Dujon, B., Colleaux, L., Jacquier, A., Michel, F., Monteilhet, C.
<2>Mitochondrial introns as mobile genetic elements: The role of intron-encoded proteins.
<3>Extrachromosomal elements in lower eukaryotes, Plenum, Wickner, R.B., Hinnebush, A., Lambowitz, A.M., Gunsalus, I.C., Hollaender, A., NY
<4>0
<5>5-27
<6>1986
<7>Introns of organelle genes share distinctive RNA secondary structures that allow their
classification into two known families. These structures are believed to play an essential
role in splicing, and members of both structural classes have recently been shown to perform
self-splicing reactions in vitro. In lower eukaryotes, many structured introns also contain
long internal open reading frames (ORFs), which are able to code for hydrophilic proteins.
Several properties of self-splicing structured introns suggest that they resemble mobile
genetic elements, even though no actual transposition event involving these introns has yet
been found. We report here on the characterization of two intron-encoded proteins that
strongly support this attractive idea. First, we show that the class I intron of the 21S
ribosomal RNA (rRNA) gene of Saccharomyces cerevisiae omega+ strains (r1 intron) encodes a
specific transposase. This protein has been partially purified from Escherichia coli cells
that overexpress it from an artificial universal code equivalent to the r1 intronic ORF. The
omega transposase shows double-strand endonuclease activity in vitro. This activity creates a
4-bp staggered cut with 3' OH overhangs within a specific sequence of the 21S rRNA gene of
omega strains. It is precisely within this sequence that the r1 intron inserts by a
duplicative transposition. Second, we report on the synthesis, in E. coli, of a putative
reverse transcriptase encoded by the class II intron of the cytochrome b gene of
Schizosaccharomyces pombe. This synthesis was obtained from E. coli expression vectors, using
the class I intronic ORF linked to an artificial initiator sequence.

<>

<1>Dujon, B., Colleaux, L., Jacquier, A., Michel, F., Monteilhet, C.
<2>Mitochondrial introns as mobile genetic elements: the role of intron-encoded proteins.
<3>Basic Life Sci.
<4>40
<5>5-27
<6>1986
<7>Introns of organelle genes share distinctive RNA secondary structures that allow
their classification into two known families.  These structures are believed to play an
essential role
in splicing, and members of both structural classes have recently been shown to perform self-
splicing reactions in vitro.  In lower eukaryotes, many structured introns also contain long
internal
open reading frames (ORFs), which are able to code for hydrophilic proteins.  Several
properties
of self-splicing structured introns suggest that they resemble mobile genetic elements, even
though
no actual transposition event involving these introns has yet been found.  We report here on
the
characterization of two intron-encoded proteins that strongly support this attractive idea.
First, we
show that the class I intron of the 21S ribosomal RNA (rRNA) gene of Saccharomyces cerevisiae
omega+ strains (r1 intron) encodes a specific transposase.  This protein has been partially
purified
from Escherichia coli cells that overexpress it from an artificial universal code equivalent
to the r1
intronic ORF.  The omega transposase shows a double-strand endonuclease activity in vitro.
This
activity creates a 4-bp staggered cut with 3' OH overhangs within a specific sequence of the
21S
rRNA gene of omega- strains.  It is precisely within this sequence that the r1 intron inserts
by a
duplicative transposition.  Second, we report on the synthesis, in E. coli, of a putative
reverse
transcriptase encoded by the Class II intron of the cytochrome b gene of Schizosaccharomyces
pombe.  This synthesis was obtained from E. coli expression vectors, using the class II
intronic
ORF linked to an artificial initiator sequence.  As further support of the idea that
structured introns
are mobile, we show, from a systematic screening of introns in various yeast species, that the
r1
intron has transposed into the ATPase subunit 9 gene of Kluyveromyces fragilis.  Structural
features observed at the new intron homing site may be relevant to the transposition event.

<>

<1>Dujon, B., Cottarel, G., Colleaux, L., Betermier, M., Jacquier, A., D'Auriol, L., Galibert, F.
<2>Mechanism of integration of an intron within a mitochondrial gene: a double strand break and the transposase function of an intron encoded protein as revealed by in vivo and in vitro assays.
<3>Achievements and Perspectives of Mitochondrial Research. Volume II: Biogenesis., Elsevier Science Publishers B.V., Quagliariello, E., Amsterdam
<4>0
<5>215-225
<6>1985
<7>The intron of the mitochondrial 21S rRNA gene (rl intron) is optional in the yeast
Saccharomyces cerevisiae, being found in some strains (omega+) but not in others (omega-) with
no phenotypic difference related to its presence or absence. Crosses between omega+ strains
and omega- strains determine a unique phenomenon that results in the integration of that
intron into the previously intron minus copies of the 21S rRNA gene. This phenomenon resembles
a duplicative transposition, with the intron itself representing the transposable element,
except that the recipient site seems unique and that the transposition is accompanied by a
unidirectional gene conversion affecting the genetic and molecular markers located in flanking
exon sequences. In addition, the transposition is nearly quantitative, converting almost all
intron minus copies of the 21S rRNA gene into intron plus copies, hence efficiently spreading
that intron within the populations of interbreeding yeasts.

<>

<1>Dumonceaux, T.J., Town, J., Links, M.G., Boyetchko, S.
<2>High-Quality Draft Genome Sequence of Pseudomonas sp. BRG100, a Strain with Bioherbicidal Properties against Setaria viridis (Green Foxtail) and Other Pests   of Agricultural Significance.
<3>Genome Announcements
<4>2
<5>e00995-14
<6>2014
<7>Pseudomonas sp. BRG100 inhibits the growth of certain agricultural pests and is a potentially
useful biopesticide for weeds and plant diseases. We have sequenced
the 6.25-Mbp genome of this strain and assembled it into 4 scaffolds. Genome
sequence comparisons revealed that this strain may represent a novel species of
Pseudomonas.

<>

<1>Duncan, C.H., Wilson, G.A., Young, F.E.
<2>Biochemical and genetic properties of site-specific restriction endonucleases in Bacillus globigii.
<3>J. Bacteriol.
<4>134
<5>338-344
<6>1978
<7>Bacillus globigii contains two site-specific endonucleases, BglI and BglII.  A
rapid technique for selection of mutants deficient in each of these enzymes was
developed using sensitivity to infection by bacteriophage SP50 as an indication
of the levels of enzyme.  Mutants defective in BglI, BglII, and both BglI and
BglII retained the wild-type modification phenotype.  Genetic and biochemical
studies have established that these enzymes are involved in restriction in
vivo.  Simplified purification procedures for BglI and BglII using these
mutants are described.

<>

<1>Duncan, S.S., Bertoli, M.T., Kersulyte, D., Valk, P.L., Tamma, S., Segal, I., McClain, M.S., Cover, T.L., Berg, D.E.
<2>Genome Sequences of Three hpAfrica2 Strains of Helicobacter pylori.
<3>Genome Announcements
<4>1
<5>e00729-13
<6>2013
<7>We present the genome sequences of three hpAfrica2 strains of Helicobacter pylori, which are
postulated to have evolved in isolation for many millennia in
people of San ethnicity. Although previously considered to be ancestral to
Helicobacter acinonychis, the hpAfrica2 strains differ markedly from H.
acinonychis in their gene arrangement. These data provide new insights into
Helicobacter evolution.

<>

<1>Dunin-Horkawicz, S., Feder, M., Bujnicki, J.M.
<2>Phylogenomic analysis of the GIY-YIG nuclease superfamily.
<3>BMC Genomics
<4>7
<5>98
<6>2006
<7>BACKGROUND: The GIY-YIG domain was initially identified in homing endonucleases and later in
other selfish mobile genetic elements
(including restriction enzymes and non-LTR retrotransposons) and in
enzymes involved in DNA repair and recombination. However, to date no
systematic search for novel members of the GIY-YIG superfamily or
comparative analysis of these enzymes has been reported. RESULTS: We
carried out database searches to identify all members of known GIY-YIG
nuclease families. Multiple sequence alignments together with predicted
secondary structures of identified families were represented as Hidden
Markov Models (HMM) and compared by the HHsearch method to the
uncharacterized protein families gathered in the COG, KOG, and PFAM
databases. This analysis allowed for extending the GIY-YIG superfamily to
include members of COG3680 and a number of proteins not classified in COGs
and to predict that these proteins may function as nucleases, potentially
involved in DNA recombination and/or repair. Finally, all old and new
members of the GIY-YIG superfamily were compared and analyzed to infer the
phylogenetic tree. CONCLUSION: An evolutionary classification of the
GIY-YIG superfamily is presented for the very first time, along with the
structural annotation of all (sub)families. It provides a comprehensive
picture of sequence-structure-function relationships in this superfamily
of nucleases, which will help to design experiments to study the mechanism
of action of known members (especially the uncharacterized ones) and will
facilitate the prediction of function for the newly discovered ones.

<>

<1>Dunitz, M.I., Coil, D.A., Jospin, G., Eisen, J.A., Adams, J.Y.
<2>Draft Genome Sequences of Escherichia coli Strains Isolated from Septic Patients.
<3>Genome Announcements
<4>2
<5>e01278-14
<6>2014
<7>We present the draft genome sequences of six strains of Escherichia coli isolated from blood
cultures collected from patients with sepsis. The strains were
collected from two patient sets, those with a high severity of illness, and those
with a low severity of illness. Each genome was sequenced by both Illumina and
PacBio for comparison.

<>

<1>Dunitz, M.I., James, P.M., Jospin, G., Eisen, J.A., Coil, D.A., Chandler, J.A.
<2>Draft Genome Sequence of Tatumella sp. Strain UCD-D_suzukii (Phylum Proteobacteria) Isolated from Drosophila suzukii Larvae.
<3>Genome Announcements
<4>2
<5>e00349-14
<6>2014
<7>Here we present the draft genome of Tatumella sp. strain UCD-D_suzukii, the first member of
this genus to be sequenced. The genome contains 3,602,931 bp in 72 scaffolds. This strain was
isolated from Drosophila suzukii larvae as part of a larger project to study the microbiota of
D. suzukii.

<>

<1>Dunlap, C.A., Bowman, M.J., Schisler, D.A.
<2>Genomic analysis and secondary metabolite production in Bacillus amyloliquefaciens AS 43.3: a biocontrol antagonist of Fusarium Head Blight.
<3>Biol. Control
<4>64
<5>166-175
<6>2013
<7>The complete genome of the biocontrol antagonist Bacillus amyloliquefaciens AS 43.3 is
reported. B. amyloliquefaciens AS 43.3 has previously been shown to be effective in reducing
Fusarium head blight in wheat. The 3.9Mbp genome was sequenced, assembled, and annotated.
Genomic analysis of the strain identified 9 biosynthetic gene clusters encoding secondary
metabolites associated with biocontrol activity. The analysis identified five non-ribosomal
peptide synthetase clusters encoding three lipopeptides (surfactin, iturin, and fengycin), a
siderophore (bacillibactin), and the antibiotic dipeptide bacilysin. In addition, three
polyketide synthetase clusters were identified which encoded for the antibacterials:
bacillaene, difficidin, and macrolactin. In addition to the non-ribosomal mediated
biosynthetic clusters discovered, we identified a ribosomally encoded biosynthetic cluster
that produces the antibiotic plantazolicin. To confirm the gene clusters were functional,
cell-free culture supernatant was analyzed using LC=96MS/MS. The technique confirmed the
presence of all nine metabolites or their derivatives. The study suggests the strain is most
likely a member of the B. amyloliquefaciens subsp. plantarium clade. Comparative genomics of
eight completed genomes of B. amyloliquefaciensidentify the core and pan-genomes for the
species, including identifying genes unique to the biocontrol strains. This study demonstrates
the growing importance of applying genomic-based studies to biocontrol organisms of plant
pathogens which can enable the rapid identification of bioactive metabolites produced by a
prospective biological control organism. In addition, this work provides a foundation for a
mechanistic understanding of the B. amyloliquefaciens AS 43.3/Fusarium head blight biocontrol
interaction.

<>

<1>Dunn, N.W., Holloway, B.W.
<2>Transduction and host controlled modification.  The role of the phage.
<3>Informative molecules in biological systems., North Holland, Ledoux, L., Amsterdam
<4>0
<5>223-233
<6>1971
<7>Three unrelated general transducing phages of Pseudomonas aeruginosa have been
studied to determine if the bacterial chromosomal fragments transferred in
transduction are always susceptible to restriction.  Using restriction and
modification mutants of the parent donor and recipient strains, to avoid
effects on recombination resulting from non-homology, two of the phages, B3 and
G101, were restricted for both their plaque forming and transducing properties.
However, a third transducing phage, F116, showed no such restriction when
transducing from a donor strain with a different DNA specificity to that of the
recipient strain.  This immunity to restriction appears to be associated with
the F115 phage genome and a variant of F116 has been found which, while immune
to restriction of plaque forming ability, is not immune to restriction of its
transducing ability.  The implication from these results is that a phage may be
able to protect bacterial DNA from inactivation by the restriction mechanisms
of host controlled modification.  With phage B3 a second type of immunity was
found, in that not all bacterial markers showed the same reduction of
transduction frequency caused by restriction and the results suggest an uneven
distribution of recognition sites on the bacterial chromosome.

<>

<1>Dunny, G.M., McKay, L.L.
<2>Group II introns and expression of conjugative transfer functions in lactic acid bacteria.
<3>Antonie Van Leeuwenhoek
<4>76
<5>77-88
<6>1999
<7>The homologous lactococcal conjugative elements pRS01 and the sex factor of Lactococcus lactis
strain 712 both contain a Group II intron within a gene believed to encode a conjugative
relaxase enzyme. This enzyme is responsible for nicking of DNA at the origin of transfer
(oriT) sequence of the sex factor DNA to initiate the strand transfer process. Group II
introns have been studied in eukaryotes, and several of these elements in yeast mitochondrial
genes have received considerable attention. These introns are relatively large in size and
generally encode a protein within the intron sequence. In addition to splicing activity. Group
II introns are mobile genetic elements. The intron-encoded proteins (IEPs) contain
endonuclease and reverse transcriptase domains believed to play an enzymatic role in genetic
mobility reactions, while a putative maturase domain is thought to promote splicing by
stabilizing the folding of the intron RNA into an active ribozyme structure which carries out
the splicing reaction. The lactococcal introns represent the first examples of Group II
introns shown to be functional in vivo in prokaryotes. Because of the advantages of a
bacterial system for genetic and molecular studies, the Ll.ltrB intron from pRS01 has
attracted the attention of several laboratories interested in Group II intron biology.
Recently, it has been shown that the system can be adapted to function in Escherichia coli
(although at somewhat reduced efficiency). In addition, it has been recently proven that the
best studied form of mobility, the homing of the intron into an intronless allele of the
cognate exon gene, occurs via an RNA intermediate and does not require DNA homology or
generalized host recombination functions. Current efforts are analysis of the role Ll.ltrB
splicing in regulating expression of pRS01 conjugation functions. The lactococcal Group II
introns represent the first demonstrated genetically mobile prokaryotic retroelements, and
they also have considerable potential as genetic engineering tools for Lactic Acid Bacteria
(LAB) and other organisms.

<>

<1>Dunten, P.W., Little, E.J., Gregory, M.T., Manohar, V.M., Dalton, M., Hough, D., Bitinaite, J., Horton, N.C.
<2>The structure of SgrAI bound to DNA; recognition of an 8 base pair target.
<3>Nucleic Acids Res.
<4>36
<5>5405-5416
<6>2008
<7>The three-dimensional X-ray crystal structure of the 'rare cutting' type II restriction
endonuclease SgrAI bound to cognate DNA is presented. SgrAI
forms a dimer bound to one duplex of DNA. Two Ca(2+) bind in the enzyme
active site, with one ion at the interface between the protein and DNA,
and the second bound distal from the DNA. These sites are differentially
occupied by Mn(2+), with strong binding at the protein-DNA interface, but
only partial occupancy of the distal site. The DNA remains uncleaved in
the structures from crystals grown in the presence of either divalent
cation. The structure of the dimer of SgrAI is similar to those of Cfr10I,
Bse634I and NgoMIV, however no tetrameric structure of SgrAI is observed.
DNA contacts to the central CCGG base pairs of the SgrAI canonical target
sequence (CR|CCGGYG, | marks the site of cleavage) are found to be very
similar to those in the NgoMIV/DNA structure (target sequence G|CCGGC).
Specificity at the degenerate YR base pairs of the SgrAI sequence may
occur via indirect readout using DNA distortion. Recognition of the outer
GC base pairs occurs through a single contact to the G from an arginine
side chain located in a region unique to SgrAI.

<>

<1>Dunten, P.W., Little, E.J., Horton, N.C.
<2>The restriction enzyme SgrAI: structure solution via combination of poor MIRAS and MR phases.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>65
<5>393-398
<6>2009
<7>Uninterpretable electron-density maps were obtained using either MIRAS phases or MR phases in
attempts to determine the structure of the type
II restriction endonuclease SgrAI bound to DNA. While neither solution
strategy was particularly promising (map correlation coefficients of
0.29 and 0.22 with the final model, respectively, for the MIRAS and MR
phases and Phaser Z scores of 4.0 and 4.3 for the rotation and
translation searches), phase combination followed by density
modification gave a readily interpretable map. MR with a distantly
related model located a dimer in the asymmetric unit and provided the
correct transformation to use in averaging electron density between
SgrAI subunits. MIRAS data sets with low substitution and MR solutions
from only distantly related models should not be ignored, as
poor-quality starting phases can be significantly improved. The
bootstrapping strategy employed to improve the initial MIRAS phases is
described.

<>

<1>Dupuis, M.-E., Villion, M., Magadan, A.H., Moineau, S.
<2>CRISPR-Cas and restriction-modification systems are compatible and increase phage resistance.
<3>Nat. Commun.
<4>4
<5>2087
<6>2013
<7>Bacteria have developed a set of barriers to protect themselves against invaders such as phage
and plasmid nucleic acids. Different prokaryotic defence systems exist and at least two of
them directly target the incoming DNA: restriction-modification (R-M) and CRISPR-Cas systems.
On their own, they are imperfect barriers to invasion by foreign DNA. Here, we show that R-M
and CRISPR-Cas systems are compatible and act together to increase the overall phage
resistance of a bacterial cell by cleaving their respective target sites. Furthermore, we show
that the specific methylation of phage DNA does not impair CRISPR-Cas acquisition or
interference activities. Taken altogether, both mechanisms can be leveraged to decrease phage
contaminations in processes relying on bacterial growth and/or fermentation.

<>

<1>Dupureur, C.M.
<2>NMR studies of restriction enzyme-DNA interactions: Role of conformation in sequence specificity.
<3>Biochemistry
<4>44
<5>5065-5074
<6>2005
<7>Sequence specific DNA binding proteins are thought to adopt distinct conformations when
binding to target (cognate) and nontarget
(noncognate) sequences. There is both biochemical and crystallographic
evidence that this behavior is important in mediating sequence
recognition by the Mg(II)-dependent Type II restriction enzymes.
Despite this, there are few systematic comparisons of the structural
behavior of these enzymes in various complexes. Here, H-1-N-15 HSQC NMR
spectroscopy is applied to PvuII endonuclease (2 x 18 kDa) in an effort
to better understand the relationship between sequence recognition and
enzyme conformational behavior. Spectra of the free enzyme collected in
the absence and presence of metal ions indicate that while there is a
modest backbone conformational response upon binding Ca(II), this does
not occur with Mg(II). Substrate binding itself is accompanied by very
dramatic spectral changes consistent with a large-scale conformational
response. HSQC spectra of the enzyme bound to cognate (specific) and
noncognate (nonspecific) oligonucleotides in the presence of Ca(II) are
dramatically distinct, revealing for the first time the structural
uniqueness of a PvuII cognate complex in solution. The strong
correlation between NMR spectral overlap and crystallographic data
(C-alpha rmsd) permits characterization of the nonspecific Pvull
complex as being more similar to the free enzyme than to the specific
complex. Collectively, these data support the notion that it is the
DNA, not the metal ion, which promotes a unique conformational response
by the enzyme. It therefore follows that the principle role of metal
ions in complex formation is one of driving substrate affinity and
stability rather than conformationally priming the enzyme for substrate
binding and sequence recognition. These results not only provide
valuable insights into the mechanism of protein-DNA interactions but
also demonstrate the utility of NMR spectroscopy in structure-function
studies of these representative nucleic acid systems.

<>

<1>Dupureur, C.M.
<2>Unique P-31 spectral response to the formation of a specific restriction enzyme-DNA complex.
<3>Nucleosides Nucleotides Nucleic Acids
<4>25
<5>747-764
<6>2006
<7>Protein-induced distortion is a dramatic but not universally observed feature of
sequence-specific DNA interactions. This is illustrated by
the crystal structures of restriction enzyme-DNA complexes: While some
of these structures exhibit DNA distortion, others do not. Among the
latter is PvuII endonuclease, a small enzyme that is also amenable to
NMR spectroscopic studies. Here P-31 NMR spectroscopy is applied to
demonstrate the unique spectral response of DNA to sequence-specific
protein interactions. The P-31 NMR spectrum of a noncognate DNA
exhibits only spectral broadening upon the addition of enzyme. However,
when enzyme is added to target DNA, a number of P-31 resonances ship
dramatically. The magnitudes of the chemical shifts (2-3 ppm) are among
the largest observed. Site-specific substitution with phosphoramidates
and phosphorothioates are used analyze these effects. While such
spectral features have been interpreted as indicative of DNA backbone
distortions, FRET analysis indicates that this does not occur in
PvuII-cognate DNA complexes in solution. The distinct P-31 spectral
signature observed for cognate DNA mirrors that observed for the
enzyme, underscoring the unique features of cognate complex formation.

<>

<1>Dupureur, C.M.
<2>Metal ion-dependent DNA-induced conformational changes in PvuII restriction endonucleases.
<3>SAAS Bulletin: Biochem. and Biotech.
<4>12
<5>43-47
<6>1999
<7>Using 19F NMR spectroscopy as a solution structural probe, important preliminary
conformational studies of the interactions between 3-fluorotyrosine-PvuII endonuclease and a
cognate DNA are described.  Even at high microM to mM concentrations of enzyme and DNA, Ca(II)
is required to observe enzyme conformational changes associated with the binding of a short
oligonucleotide.  Further, these structural changes are global: The resonances of no fewer
than 6 of the 10 3-fluorotyrosine residues per PvuII subunit shift by at least 0.1 ppm
relative to the free enzyme, far more than can be accounted for as local effects of DNA
binding.  Partial titration experiments are consistent with a slow off-rate for
oligonucleotide binding and correspondingly slow (ms) exchange between free and DNA-bound
conformations of enzyme.  These results demonstrate that this system provides a means to
explore solution conformational aspects of restriction enzyme-DNA interactions.

<>

<1>Dupureur, C.M., Conlan, L.H.
<2>A catalytically deficient active site variant of PvuII endonuclease binds Mg(II) ions.
<3>Biochemistry
<4>39
<5>10921-10927
<6>2000
<7>In efforts to understand the mechanisms of many nucleic acid enzymes, the first site-directed
mutations are made at conserved acidic active residues. Almost without exception, the low or
null activities of the resulting variants are attributed to the importance of the acidic
residue(s) to the ligation of required metal ions. Using Mg^25 NMR spectroscopy as a direct
probe of metal ion binding and the homodimeric PvuII restriction endonuclease as a model
system, this interpretation is examined and clarified. Our results indicate that Mg(II) binds
wild-type PvuII endonuclease in the absence of DNA with a K(d,app) of 1.9 mM. Hill analysis
yields an n(H) coefficient of 1.4, a value consistent with the binding of more than one Mg(II)
ion per monomer active site. Variable pH studies indicate that two ionizable groups are
responsible for Mg(II) binding by wild-type PvuII endonuclease near physiological pH. The
pK(a,app) for these ionizations is 6.7, a value which is unusual for acidic residues but
consistent with data obtained for critical groups in MunI endonuclease and a number of other
hydrolases. To assign residues critical to ligating Mg(II), binding measurements were
performed on the low activity catalytic site mutants E68A and D58A. As expected, E68A binds
Mg(II) ions very weakly (K(d,app) approximately 40 mM), implicating Glu68 as critical to
Mg(II) binding. Interestingly, while D58A has only residual specific activity, it retains an
affinity for Mg(II) with a K(d,app) of 3.6 mM and exhibits a Hill coefficient of 0.7.
Moreover, in this variant, multiple ionizable groups with pK(a,app) of 7.2 are involved in
Mg(II) binding, suggesting a shuffling of Mg(II) ligands in the active site. These data
indicate that Asp58 is important for the critical positioning of metal ion(s) required for
catalysis.

<>

<1>Dupureur, C.M., Dominguez, M.A.
<2>The PD ... (D/E)XK motif in restriction enzymes: A link between function and conformation.
<3>Biochemistry
<4>40
<5>387-394
<6>2001
<7>The active sites of Mg(II)-dependent nucleases feature a cluster of conserved charged residues
which includes both acidic (Asp and Glu) and
basic (Lys) side chains. In restriction enzymes, these side chains are
part of the conserved PD...(D/E)XK functional sequence motif which has
been implicated as being important in metal ion binding and catalytic
steps. Recent work revealing the unusual behavior of the active site
variant D58A of the representative PvuII endonuclease prompted
speculation that the array of charged groups in the nuclease active
site may also be linked to conformational behavior [Dupureur, C. M.
and Conlan, L. H. (2000) Biochemistry 39, 10921-10927]. To address this
issue, we analyzed the conformational behavior of active site variants
of PvuII endonuclease using both NMR spectroscopic and thermodynamic
methods. NMR spectroscopic analysis via F-19 and H-1-N-15 HSQC
experiments indicates that a number of side chain and backbone amide
groups are perturbed upon Ala substitution at conserved active site
residues Asp58, Glu68, and Lys70. Spectral changes are particularly
pronounced for the lowest-activity mutants (D58A and K70A). These
changes are accompanied by perturbations in conformational stability.
Ala substitution at each of these positions results in 2-5 kcal/mol of
stabilization over the wild-type enzyme at pH 7.7, changes which
constitute increases in DeltaG(d)(H2O) of 20-50%. The pH dependencies
of mutant enzyme stabilities are distinct from those of the wild type,
results which confirm that these ionizable groups strongly influence
stability. Wild-type enzyme stability is correlated with the ionization
of groups shown to be important to metal ion binding and orientation.
Correlations between spectral changes and conformational stability
indicate that the latter measurements may prove useful in the
evaluation of site-directed mutant restriction enzymes. More
importantly, these results indicate that structure-function
relationships in restriction enzyme active sites can be complex, and
that the ensemble of conserved charged residues which mediate DNA
hydrolysis in Mg(II)-dependent nucleases constitutes a critical link
between function and conformation.

<>

<1>Dupureur, C.M., Hallman, L.M.
<2>Effects of divalent metal ions on the activity and conformation of native and 3-fluorotyrosine-PvuII endonucleases.
<3>Eur. J. Biochem.
<4>261
<5>261-268
<6>1999
<7>The activities of restriction enzymes are important examples of Mg(II)-dependent hydrolysis of
DNA. While a number of crystallographic studies of enzyme-DNA complexes have also involved
metal ions, there have been no solution studies exploring the relationship between enzyme
conformation and metal-ion binding in restriction enzymes.  Using PvuII restriction
endonuclease as a model system, we have successfully developed biosynthetic fluorination and
NMR spectroscopy as a solution probe of restriction-enzyme conformation.  The utility of this
method is demonstrated with a study of metal-ion binding by PvuII endonuclease.  Replacement
of 74% (+-10%) of the Tyr residues in PvuII endonuclease by 3-fluorotyrosine produces an
enzyme with Mg(II)-supported specific activity and sequence specificity that is
indistinguishable from that of the native enzyme.  Mn(II) supports residual activity of both
the native and fluorinated enzymes; Ca(II) does not support activity in either enzyme, a
result consistent with previous studies.  1H- and 19F-NMR spectroscopic studies reveal that
while Mg(II) does not alter the enzyme conformation, the paramagnetic Mn(II) produces both
short-range spectral broadening and longer range changes in chemical shift.  Most
interestingly, Ca(II) binding perturbs a larger number of different resonances than Mn(II).
Coupled with earlier mutagenesis studies that place Ca(II) in the active site these data
suggest that the enzyme makes conformational adjustments to accommodate the distinct geometric
preferences of Ca(II) and may play a role in the inability of this metal ion to support
activity in restriction enzymes.

<>

<1>Dupuy, V. et al.
<2>Complete Genome Sequence of Mycoplasma putrefaciens Strain 9231, One of the Agents of Contagious Agalactia in Goats.
<3>Genome Announcements
<4>1
<5>e00354-13
<6>2013
<7>Mycoplasma putrefaciens is one of the etiologic agents of contagious agalactia in goats. We
report herein the complete genome sequence of Mycoplasma putrefaciens
strain 9231.

<>

<1>Dupuy, V., Thiaucourt, F.
<2>Complete Genome Sequence of Mycoplasma capricolum subsp. capripneumoniae Strain 9231-Abomsa.
<3>Genome Announcements
<4>2
<5>e01067-14
<6>2014
<7>Mycoplasma capricolum subsp. capripneumoniae is the etiological agent of contagious caprine
pleuropneumonia. We report here the complete and annotated
genome sequence of M. capricolum subsp. capripneumoniae strain 9231-Abomsa.

<>

<1>Duquesne, K., Prima, V., Ji, B., Rouy, Z., Medigue, C., Talla, E., Sturgis, J.N.
<2>Draft Genome Sequence of the Purple Photosynthetic Bacterium Phaeospirillum molischianum DSM120, a Particularly Versatile Bacterium.
<3>J. Bacteriol.
<4>194
<5>3559-3560
<6>2012
<7>Here we present the draft genome sequence of the versatile and adaptable purple photosynthetic
bacterium Phaeospirillum molischianum DSM120. This study advances
the understanding of the adaptability of this bacterium, as well as the
differences between the Phaeospirillum and Rhodospirillum genera.

<>

<1>Duquesne, K., Sturgis, J.N.
<2>Shotgun Genome Sequence of the Large Purple Photosynthetic Bacterium Rhodospirillum photometricum DSM122.
<3>J. Bacteriol.
<4>194
<5>2380
<6>2012
<7>Here, we present the shotgun genome sequence of the purple photosynthetic bacterium
Rhodospirillum photometricum DSM122. The photosynthetic apparatus of
this bacterium has been particularly well studied by microscopy. The knowledge of
the genome of this oversize bacterium will allow us to compare it with the other
purple bacterial organisms to follow the evolution of the photosynthetic
apparatus.

<>

<1>Durai, S., Mani, M., Kandavelou, K., Wu, J., Porteus, M.H., Chandrasegaran, S.
<2>Zinc finger nucleases: custom-designed molecular scissors for genome engineering of plant and mammalian cells.
<3>Nucleic Acids Res.
<4>33
<5>5978-5990
<6>2005
<7>Custom-designed zinc finger nucleases (ZFNs), proteins designed to cut at specific DNA
sequences, are becoming powerful tools in gene targeting--the
process of replacing a gene within a genome by homologous recombination
(HR). ZFNs that combine the non-specific cleavage domain (N) of FokI
endonuclease with zinc finger proteins (ZFPs) offer a general way to
deliver a site-specific double-strand break (DSB) to the genome. The
development of ZFN-mediated gene targeting provides molecular biologists
with the ability to site-specifically and permanently modify plant and
mammalian genomes including the human genome via homology-directed repair
of a targeted genomic DSB. The creation of designer ZFNs that cleave DNA
at a pre-determined site depends on the reliable creation of ZFPs that can
specifically recognize the chosen target site within a genome. The
(Cys2His2) ZFPs offer the best framework for developing custom ZFN
molecules with new sequence-specificities. Here, we explore the different
approaches for generating the desired custom ZFNs with high
sequence-specificity and affinity. We also discuss the potential of
ZFN-mediated gene targeting for 'directed mutagenesis' and targeted 'gene
editing' of the plant and mammalian genome as well as the potential of
ZFN-based strategies as a form of gene therapy for human therapeutics in
the future.

<>

<1>Duran, E., Ramsauer, V.P., Ballester, M., Torrenegra, R.D., Rodriguez, O.E., Winkle, S.A.
<2>Qualitative analysis of sequence specific binding of flavones to DNA using restriction endonuclease activity assays.
<3>Biopolymers
<4>99
<5>530-537
<6>2013
<7>Flavones, found in nature as secondary plant metabolites, have shown efficacy as anti-cancer
agents. We have examined the binding of two
flavones, 5,7-dihydroxy-3,6,8-trimethoxy-2-phenyl-4H-chromen-4-one
(5,7-dihydroxy-3,6,8-trimethoxy flavone; FlavA) and
3,5-dihydroxy-6,7,8-trimethoxy-2-phenyl-4H-chromen-4-one
(3,5-dihydroxy-6,7,8-trimethoxy flavone; FlavB), to phiX174 RF DNA
using restriction enzyme activity assays employing the restriction
enzymes Alw44, AvaII, BssHII, DraI, MluI, NarI, NciI, NruI, PstI, and
XhoI. These enzymes possess differing target and flanking sequences
allowing for observation of sequence specificity analysis. Using
restriction enzymes that cleave once with a mixture of supercoiled and
relaxed DNA substrates provides for observation of topological effects
on binding. FlavA and FlavB show differing sequence specificities in
their respective binding to phiX. For example, with relaxed DNA, FlavA
shows inhibition of cleavage with DraI (reaction site 5TTTAAA) but not
BssHII (5GCGCGC) while FlavB shows the opposite results. Evidence for
tolological specificity is also observed, Molecular modeling and
conformational analysis of the flavones suggests that the phenyl ring
of FlavB is coplanar with the flavonoid ring while the phenyl ring of
FlavA is at an angle relative to the flavonoid ring. This may account
for aspects of the observed sequence and topological specificities in
the effects on restriction enzyme activity.

<>

<1>Duranti, S., Turroni, F., Lugli, G.A., Milani, C., Viappiani, A., Mangifesta, M., Gioiosa, L., Palanza, P., van Sinderen, D., Ventura, M.
<2>Genomic Characterization and Transcriptional Studies of the Starch-Utilizing Strain Bifidobacterium adolescentis 22L.
<3>Appl. Environ. Microbiol.
<4>80
<5>6080-6090
<6>2014
<7>Bifidobacteria are members of the gut microbiota but the genetic basis for their adaptation to
the human gut is poorly understood.  Analysis of the 2,203,222 bp genome of Bifidobacterium
adolescentis 22L revealed a nutrient-acquisition strategy that targets diet/plant-derived
glycans, in particular starch and starch-like carbohydrates.  Starch-like carbohydrates were
shown to support the growth of B. acelescentis 22L.  Transcriptome profiling of 22L cultures
grown under in vitro conditions or during colonization of the murine gut by RNAseq and qRT-PCR
assays revealed the expression of a set of chromosomal loci responsible for starch metabolism
as well as for pili production.  Such extracellular structures include so-called
sortase-dependent and type IVb pili, which may be involved in gut colonization of 22L through
adhesion to extracellular matrix proteins.

<>

<1>Durfee, T., Nelson, R., Baldwin, S., Plunkett, G. III, Burland, V., Mau, B., Petrosino, J.F., Qin, X., Muzny, D.M., Ayele, M., Gibbs, R.A., Csorgo, B., Posfai, G., Weinstock, G.M., Blattner, F.R.
<2>The complete genome sequence of Escherichia coli DH10B: insights into the biology of a laboratory workhorse.
<3>J. Bacteriol.
<4>190
<5>2597-2606
<6>2008
<7>Escherichia coli DH10B was designed for the propagation of large insert DNA library clones. It
is used extensively, taking advantage of properties
such as high DNA transformation efficiency and maintenance of large
plasmids. The strain was constructed by serial genetic recombination
steps, but the underlying sequence changes remained unverified. We report
the complete genomic sequence of DH10B by using reads accumulated from the
bovine sequencing project at Baylor College of Medicine and assembled with
DNAStar's SeqMan genome assembler. The DH10B genome is largely colinear
with that of the wild-type K-12 strain MG1655, although it is
substantially more complex than previously appreciated, allowing DH10B
biology to be further explored. The 226 mutated genes in DH10B relative to
MG1655 are mostly attributable to the extensive genetic manipulations the
strain has undergone. However, we demonstrate that DH10B has a 13.5-fold
higher mutation rate than MG1655, resulting from a dramatic increase in
insertion sequence (IS) transposition, especially IS150. IS elements
appear to have remodeled genome architecture, providing homologous
recombination sites for a 113,260-bp tandem duplication and an inversion.
DH10B requires leucine for growth on minimal medium due to the deletion of
leuLABCD and harbors both the relA1 and spoT1 alleles causing both
sensitivity to nutritional downshifts and slightly lower growth rates
relative to the wild type. Finally, while the sequence confirms most of
the reported alleles, the sequence of deoR is wild type, necessitating
reexamination of the assumed basis for the high transformability of DH10B.

<>

<1>Durham, B.P. et al.
<2>Draft genome sequence of marine alphaproteobacterial strain HIMB11, the first cultivated representative of a unique lineage within the Roseobacter clade  possessing an unusually small genome.
<3>Standards in Genomic Sciences
<4>9
<5>632-645
<6>2014
<7>Strain HIMB11 is a planktonic marine bacterium isolated from coastal seawater in  Kaneohe Bay,
Oahu, Hawaii belonging to the ubiquitous and versatile Roseobacter
clade of the alphaproteobacterial family Rhodobacteraceae. Here we describe the
preliminary characteristics of strain HIMB11, including annotation of the draft
genome sequence and comparative genomic analysis with other members of the
Roseobacter lineage. The 3,098,747 bp draft genome is arranged in 34 contigs and
contains 3,183 protein-coding genes and 54 RNA genes. Phylogenomic and 16S rRNA
gene analyses indicate that HIMB11 represents a unique sublineage within the
Roseobacter clade. Comparison with other publicly available genome sequences from
members of the Roseobacter lineage reveals that strain HIMB11 has the genomic
potential to utilize a wide variety of energy sources (e.g. organic matter,
reduced inorganic sulfur, light, carbon monoxide), while possessing a reduced
number of substrate transporters.

<>

<1>Durmaz, E., Higgins, D.L., Klaenhammer, T.R.
<2>Molecular characterization of a second abortive phage resistance gene present in Lactococcus lactis subsp. lactis ME2.
<3>J. Bacteriol.
<4>174
<5>7463-7469
<6>1992
<7>The fifth phage resistance factor from the prototype phage-insensitive strain Lactococcus
lactis subsp. lactis ME2 has been characterized and sequenced. The genetic determinant for Prf
(phage resistance five) was subcloned from the conjugative plasmid pTN20, which also encodes a
restriction and modification system. Typical of other abortive resistance mechanisms, Prf
reduces the efficiency of plaquing to 10-2 to 10-3 and decreases the plaque size and burst
size of the small isometric-headed phage p2 in L. lactis subsp. lactis LM0230. However,
normal-size plaques occurred at a frequency of 10-4 and contained mutant phages that were
resistant to Prf, even after repeated propagation through a sensitive host. Prf does not
prevent phage adsorption or promote restriction and modification activities, but 90% of Prf+
cells infected with phage p2 die. Thus, phage infections in Prf+ cells are aborted. Prf is
effective in both L. lactis subsp. lactis and L. lactis subsp. cremoris strains against
several small isometric-headed phages but not against prolate-headed phages. The Prf
determinant was localized by Tn5 mutagenesis and subcloning. DNA sequencing identified a
1,056-nucleotide strutural gene designated abiC, Prf+ expression was obtained when abiC was
subcloned into the lactococcal expression vector pMG36e, abiC is distinct from two other
lactococcal abortive phage resistance genes, abiA (Hsp+, from L. lactis subsp. lactis ME2) and
abi416 (Abi+, from L. lactis subsp. lactis IL416). Unlike abiA, the action of abiC does not
appear to affect DNA replication. Thus, abiC represents a second abortive system found in ME2
that acts at a different point of the phage lytic cycle.

<>

<1>Durmaz, E., Klaenhammer, T.R.
<2>A starter culture rotation strategy incorporating paired restriction/modification and abortive infection bacteriophage defenses in a single lactococcus lactis strain.
<3>Appl. Environ. Microbiol.
<4>61
<5>1266-1273
<6>1995
<7>Three derivatives of Lactococcus lactis subsp. lactis NCK203, each with a different pair of
restriction/modification (R/M) and abortive infection (Abi) phage defense systems, were
constructed and then rotated in repeated cycles of a milk starter culture activity test (SAT).
The rotation proceeded successfully through nine successive SATs in the presence of phage and
whey containing phage from previous cycles. Lactococcus cultures were challenged with 2 small
isometric-headed phages, omega-31 and ul36, in one rotation series and with a composite of 10
industrial phages in another series. Two native lactococcal R+/M+ plasmids, pTRK68 and pTRK11,
and one recombinant plasmid, pTRK308, harboring a third distinct R/M system were incorporated
into three NCK203 derivatives constructed separately for the rotation. The R+/M+ NCK203
derivatives were transformed with high-copy-number plasmids encoding four Abi genes, abiA,
abiC, per31, and per50. Various Abi and R/M combinations constructed in NCK203 were evaluated
for their effects on cell growth, level of phage resistance, and retardation of phage
development during repeated cycles of the SAT. The three NCK203 derivatives chosen for use in
the SAT exhibited additive effects of the R/M and Abi phenotypes against sensitive phages. In
such combinations, phage escaping restriction are prevented from completing their infective
cycle by an abortive response that kills the host cell. The rotation series successfully
controlled modified, recombinant, and mutant phages which were resistant to any one of the
individual defense systems by presenting a different set of R/M and Abi defenses in the next
test of the rotation.

<>

<1>Durmaz, E., Klaenhammer, T.R.
<2>Abortive phage resistance mechanism AbiZ speeds the lysis clock to cause premature lysis of phage-infected Lactococcus lactis.
<3>J. Bacteriol.
<4>189
<5>1417-1425
<6>2007
<7>The conjugative plasmid pTR2030 has been used extensively to confer phage resistance in
commercial Lactococcus starter cultures. The
plasmid harbors a 16-kb region, flanked by insertion sequence (IS)
elements, that encodes the restriction/modification system L1aI and
carries an abortive infection gene, abiA. The AbiA system inhibits both
prolate and small isometric phages by interfering with the early stages
of phage DNA replication. However, abiA alone does not account for the
full abortive activity reported for pTR2030. In this study, a 7.5-kb
region positioned within the IS elements and downstream of abiA was
sequenced to reveal seven additional open reading frames (ORFs). A
single ORF, designated abiZ, was found to be responsible for a
significant reduction in plaque size and an efficiency of plaquing
(EOP) of 10(-6), without affecting phage adsorption. AbiZ causes phage
phi 31-infected Lactococcus lactis NCK203 to lyse 15 min early,
reducing the burst size of phi 31 100-fold. Thirteen of 14 phages of
the P335 group were sensitive to AbiZ, through reduction in either
plaque size, EOP, or both. The predicted AbiZ protein contains two
predicted transmembrane helices but shows no significant DNA
homologies. When the phage phi 31 lysin and holin genes were cloned
into the nisin-inducible shuttle vector pMSP3545, nisin induction of
holin and lysin caused partial lysis of NCK203. In the presence of
AbiZ, lysis occurred 30 min earlier. In holin-induced cells, membrane
permeability as measured using propidium iodide was greater in the
presence of AbiZ. These results suggest that AbiZ may interact
cooperatively with holin to cause premature lysis.

<>

<1>Durmaz, E., Miller, M.J., Azcarate-Peril, M.A., Toon, S.P., Klaenhammer, T.R.
<2>Genome sequence and characteristics of Lrm1, a prophage from industrial Lactobacillus rhamnosus strain M1.
<3>Appl. Environ. Microbiol.
<4>74
<5>4601-4609
<6>2008
<7>Prophage Lrm1 was induced with mitomycin C from an industrial Lactobacillus rhamnosus starter
culture, M1. Electron microscopy of the
lysate revealed relatively few intact bacteriophage particles among
empty heads and disassociated tails. The defective Siphoviridae phage
had an isometric bead of approximately 55 nm and noncontractile tail of
about 275 nm with a small baseplate. In repeated attempts, the prophage
could not be cured from L. rhamnosus M1, nor could a sensitive host be
identified. Sequencing of the phage Lrm1 DNA revealed a genome of
39,989 bp and a G+C content of 45.5%. A similar genomic organization
and mosaic pattern of identities align Lrm1 among the closely related
Lactobacillus casei temperate phages A2, Phi AT3, and LeaI and with L.
rhamnosus virulent phage Lu-Nu. Of the 54 open reading frames (ORFs)
identified, all but 8 shared homology with other phages of this group.
Five unknown ORFs were identified that had no homologies in the
databases nor predicted functions. Notably, Lrm1 encodes a putative
endonuclease and a putative DNA methylase with homology to a methylase
in Lactococcus lactis phage Tuc2009. Possibly, the DNA methylase,
endonuclease, or other Lrm1 genes provide a function crucial to L.
rhamnosus M1 survival, resulting in the stability of the defective
prophage in its lysogenic state. The presence of a defective prophage
in an industrial strain could provide superinfection immunity to the
host but could also contribute DNA in recombination events to produce
new phages potentially infective for the host strain in a large-scale
fermentation environment.

<>

<1>Durrenberger, F., Rochaix, J.-D.
<2>Chloroplast ribosomal intron of Chlamydomonas reinhardtii: in vitro self-splicing, DNA endonuclease activity and in vivo mobility.
<3>EMBO J.
<4>10
<5>3495-3501
<6>1991
<7>All chloroplast 23S ribosomal RNA genes of the unicellular alga Chlamydomonas reinhardtii
contain an 888 bp group I intron with an internal open reading frame (ORF). A precursor RNA
encompassing the intron with its 5' and 3' flanking sequences was shown to self-splice both
during in vitro transcription and upon incubation of the isolated pre-RNA under self-splicing
conditions. Expression of the internal ORF in Escherichia coli in the presence of a plasmid
containing a cDNA corresponding to the intronless form of the 23S rRNA gene resulted in
specific cleavage of the cDNA at or close to the exon junction sequence. To test whether this
ORF-encoded double-strand DNA endonuclease is involved in intron mobility in vivo, the same
ribosomal cDNA was stably integrated into the C. reinhardtii chloroplast genome using particle
gun mediated transformation. All the transformants with the cDNA integrated at the expected
site in the chloroplast genome had the intron precisely inserted at the artificial exon
junction site. These experiments demonstrate that the chloroplast ribosomal intron of C.
reinhardtii behaves as a ribozyme in vitro and also as a mobile genetic element in vivo
provided a target site is present.

<>

<1>Durrenberger, F., Rochaix, J.-D.
<2>Characterization of the cleavage site and the recognition sequence of the I-CreI DNA endonuclease encoded by the chloroplast ribosomal intron of Chlamydomonas reinhardtii.
<3>Mol. Gen. Genet.
<4>236
<5>409-414
<6>1993
<7>The chloroplast ribosomal intron of Chlamydomonas reinhardtii encodes a sequence-specific DNA
endonuclease (I-CreI), which is most probably involved in the mobility of this intron. Here we
show that I-CreI generates a 4 bp staggered cleavage just downstream of the intron insertion
site. The I-CreI recognition sequence is 19-24 bp in size and is located asymmetrically around
the intron insertion site. Screening of natural variants of the I-CreI recognition sequence
indicates that the I-CreI endonuclease tolerates single and even multiple base changes within
its recognition sequence.

<>

<1>Durrett, R., Miras, M., Mirouze, N., Narechania, A., Mandic-Mulec, I., Dubnau, D.
<2>Genome Sequence of the Bacillus subtilis Biofilm-Forming Transformable Strain PS216.
<3>Genome Announcements
<4>1
<5>e00288-13
<6>2013
<7>Bacillus subtilis PS216, a strain isolated in Slovenia, has been sequenced. PS216 is
transformable and forms robust biofilms, making it useful for the study of
competence regulation in an undomesticated bacterium.

<>

<1>Dussoix, D., Arber, W.
<2>Host Specificity of DNA Produced by Escherichia Coli:  II. Control over acceptance of DNA from infecting phage lambda.
<3>J. Mol. Biol.
<4>5
<5>37-49
<6>1962
<7>DNA of lambda.K (lambdaphage grown on E. coli strain K12) is shown to be
degraded upon infection of the new host strains E. coli K12(P1) or E. coli B.
This breakdown begins shortly after phage attachment and successful DNA
injection.  32P label from the lambda.K DNA submitted to this degradation
appears partly in acid-soluble components (organic and inorganic) and partly in
acid-insoluble compounds.  The host cell survives such an infection and permits
diffusion of a fraction of the degradation products into the medium, while
probably re-using another fraction.  Genetic markers from lambda.K are rescued
in K12(P1) host cells infected with both restricted lambda.K and unrestricted
lambda.K(P1).  Since DNA breakdown competes in time with the rescue, the
probability of marker rescue is high if the unrestricted phage infects first
and low if the restricted phage infects first.  Only closely linked markers
have a good chance to be rescued together.  The host specificity imparted to
phage DNA by the bacterial strain on which it was produced is thought to be
responsible for its recognition as incompatible with a new host strain.
Bacterial mutants are described which, despite the presence of prophage P1,
accept infecting lambda.K at relatively high rates.

<>

<1>Dussoix, D., Arber, W.
<2>Host specificity of DNA produced by Escherichia coli IV.  Host specificity of infectious DNA from bacteriophage lambda.
<3>J. Mol. Biol.
<4>11
<5>238-246
<6>1965
<7>DNA extracted with phenol from bacteriophage lambda gives rise to phage
production after uptake by helper-infected, competent recipient cells (Kaiser,
2962).  Under our experimental conditions, the number of infective centres
obtained by infection of Escherichia coli K12 is about 10-3 per phage
equivalent of DNA from lambda.K or from lambda.K(P1).  But on K12(P1) recipient
cells only lambda-K(P1) DNA infects with an efficiency of 10-3, while lambda.K
DNA gives about 100 times less infective centres.  The same factor of
restriction for lambda.K is found in controls done by infection of the
competent cells with intact phage particles instead of the phage DNA.
Similarly, restrictions displayed by K12 against phage grown on E. coli B or E.
coli C and those displayed by B against phage grown on K12 or C are found to
hold true for DNA preparations.  E. coli C accepts all tested lambda DNA with
about the same efficiency.  We conclude that the phenol extraction does not
affect the host specificity of the phage DNA.  One cycle growth of lambda
initiated by infection of K12 with lambda.K(P1) DNA confirms this result:  the
parental P1-directed host specificity is transferred into the phage progeny,
and it is found only in such phage particles that also inherit one strand of
the infecting DNA molecule.  The stability of the association of DNA with its
host specificity is further revealed by its resistance to various physical,
chemical and enzymic treatments of the lambda DNA.  It is significant with
respect to the understanding of the mechanism of competence of bacteria for
infection with lambda DNA that only non-restricted phage acts as a good helper.

<>

<1>Dusterhoft, A.
<2>Cloning of the restriction-modification systems from Herpetosiphon giganteus Hpa1, Hpa2 and Hpg5 in E. coli - Characterization and comparative analysis of the genetic organization and structure.
<3>Ph.D. Thesis, Justus-Liebig University, Geissen, W. Germany
<4>
<5>1-160
<6>1990
<7>None - but this thesis shows that M.HgiBI, M.HgiDI, M.HgiDII and M.HgiGI are
m5C-methylases.  Also, because SalI was used to select M.HgiDII gene and SalI
can cleave when the last cytosine in the recognition sequence (GTCGAC) is m5C,
M.HgiDII must methylate the first cytosine in the sequence.

<>

<1>Dusterhoft, A., Erdmann, D., Kroger, M.
<2>Stepwise cloning and molecular characterization of the HgiDI restriction-modification system from Herpetosiphon giganteus Hpa2.
<3>Nucleic Acids Res.
<4>19
<5>1049-1056
<6>1991
<7>The restriction-modification system HgiDI from Herpetosiphon giganteus strain
Hpa2 has been cloned in E. coli in a two-step procedure.  Selection of the
methyltransferase (M.HgiDI) gene in vitro was performed using the heterologous
restriction endonuclease AhaII, an isoschizomer of AcyI and HgiDI (GRCGYC).
Cloning of the complete HgiDI endonuclease (R.HgiDI) gene could only be
achieved in recipient cells harbouring a recombinant plasmid, which was
expressing the corresponding methyltransferase and could thereby prevent the
host from self-destruction of its genetic material.  The HgiDI
restriction-modification system was sequenced and functionally correlated with
two open reading frames of 309 (M) and 359 (R) codons.  In homology studies
M.HgiDI showed significant similarities to 20 other m5C-methyltransferases and
turned out to be the most compact enzyme of this group described so far.
Initial attempts for overexpression of M.HgiDI and partial purification of
R.HgiDI have been successful.

<>

<1>Dusterhoft, A., Erdmann, D., Kroger, M.
<2>Isolation and genetic structure of the AvaII isoschizomeric restriction-modification system HgiBI from Herpetosiphon giganteus Hpg5:  M.HgiBI reveals high homology to M.BanI.
<3>Nucleic Acids Res.
<4>19
<5>3207-3211
<6>1991
<7>The complete type II restriction-modification system HgiBI of Herpetosiphon
giganteus strain Hpg5 recognizing the AvaII specific DNA sequence GGWCC has
been cloned and expressed functionally active in Escherichia coli.  A
considerable acceleration in cloning could be achieved by preparing a size
restricted library after application of a related hybridization probe.  Both
methyltransferase (437 codons) and restriction endonuclease gene (274 codons)
were found to be encoded on a 3.6 kilobases ClaI/HincII fragment in the same
transcriptional orientation separated by one triplet only.  Protein sequence
comparisons revealed a close resemblance of M.HgiBI to the group of
m5C-methyltransferases, especially to M.BanI from Bacillus aneurinolyticus with
the related recognition sequence GGYRCC.  In contrast, no significant
similarities have been observed for the associated endonuclease R.HgiBI with
any other restriction enzyme described so far, even not with the isoschizomeric
R.SinI from Salmonella infantis, or with R.BanI.

<>

<1>Dusterhoft, A., Kroger, M.
<2>Site directed mutagenesis on the restriction endonuclease EcoRI using mixed oligonucleotides.
<3>Nucl. Nucl.
<4>7
<5>737-740
<6>1988
<7>The restriction endonuclease EcoRI could be modified via site directed
mutagenesis at position Arg200.  Using the thiophosphate system we introduced
either Lys, Glu or Gly in a one pot procedure.  Although G recognition should
be affected, Lys200 showed wildtype specificity.

<>

<1>Dusterhoft, A., Kroger, M.
<2>Cloning, sequence and characterization of m5C-methyltransferase-encoding gene, hgiDIIM (GTCGAC), from Herpetosiphon giganteus strain Hpa2.
<3>Gene
<4>106
<5>87-92
<6>1991
<7>We have cloned the gene (hgiDIIM) encoding the methyltransferase (MTase) of the SalI
isoschizomeric restriction-modification (R-M) system, HgiDII (GTCGAC), into Escherichia coli.
The hgiDIIM gene has been isolated from the same plasmid library of Herpetosiphon giganteus
strain Hpa2, as was the previously cloned R-M system, HgiDI [AcyI/GRCGYC; Dusterhoft et al.,
Nucl. Acids Res. 19 (1991) 1049-1056]. Sequencing and functional localization of hgiDIIM
revealed an open reading frame (ORF) of 354 codons (39786 Da) with significant homologies to
the group of m5C-, rather than the m4C-/m6A-, MTases. Subsequent cloning and analysis of
adjacent chromosomal segments led to the identification of two additional ORFs upstream
(ORF15, 139 codons) and downstream (ORF68, 611 codons) from hgiDIIM with the same
transcriptional orientation as the hgiDIIM gene. However, the expected restriction enzyme
function was not found in either of these ORFs.

<>

<1>Duyvesteyn, M.G.C., de Waard, A.
<2>A new sequence-specific endonuclease from a thermophilic cyanobacterium, Mastigocladus laminosus.
<3>FEBS Lett.
<4>111
<5>423-426
<6>1980
<7>The screening of a number of cyanobacteria (blue-green algae) for new
sequence-specific deoxyribonucleases has been very rewarding.  We report here
the purification of a new enzyme of this type from Mastigocladus laminosus
(named MlaI) which recognizes and cleaves the nucleotide sequence TT^CGAA.

<>

<1>Duyvesteyn, M.G.C., de Waard, A., van Ormondt, H.
<2>Two sequence-specific deoxyribonucleases from Rhodospirillum rubrum.
<3>FEBS Lett.
<4>117
<5>241-246
<6>1980
<7>The usefulness of sequence-specific endonucleases in solving fundamental
problems in current molecular biology is well established.  The catalog of
enzymes has increased steadily.  We report here the isolation and partial
characterization of two enzymes from Rhodospirillum rubrum, endonucleases
R.RruI and R.RruII.  The first enzyme is unique in that it recognizes and
cleaves the hexanucleotide sequence AGT^ACT in DNA molecules of various origin.
The latter enzyme (endo R.RruII) recognizes the nucleotide sequence CC(A/T)GG
as does endo R.EcoRII; however, it cleaves the DNA within the site like the
isoschizomer endo R.BstNI (CC^(A/T)GG).

<>

<1>Duyvesteyn, M.G.C., Korsuize, J., de Waard, A.
<2>Isolation and characterization of a sequence-specific deoxyriboendonuclease from Calothrix scopulorum.
<3>Plant Mol. Biol.
<4>1
<5>75-79
<6>1981
<7>Sequence-specific deoxyriboendonucleases have been isolated from bacteria and
fungi.  Except in a few cases the nucleotide sequences recognized by the
cyanobacerial enzymes have been shown to be unique.  In this report we describe
the isolation and characterization of an endonuclease (endo R. CscI) from a
cyanobacterium, Calothrix scopulorum which cleaves the deoxynucleotide sequence
CCGC^GG.  Isoschizomers have been found previously in a fungus and in two
bacterial strains.

<>

<1>Duyvesteyn, M.G.C., Korsuize, J., de Waard, A., Vonshak, A., Wolk, C.P.
<2>Sequence-specific endonucleases in strains of Anabaena and Nostoc.
<3>Arch. Microbiol.
<4>134
<5>276-281
<6>1983
<7>The complements of restriction endonucleases of 12 strains of cyanobacteria were determined in
cell-free extracts, and were compared with the complements of restriction activities assessed
by measuring the relative efficiencies of plating of cyanophages on those cyanobacteria.  The
hosts which were susceptible to all of the phages contained endo R.AvaI and endo R.AvaII, and
in several cases probably endo R.AvaIII, or isoschizomers of these enzymes. Three hosts which
were lysed by only a subset (1 or 3) of the phages contained different restriction
endonucleases.  Anabaena sp. PCC 7120 showed apparent phenotypic restriction of phage AN-22
grown in hosts with (isoschizomers of) AvaI, II and III, but no corresponding endonuclease has
yet been detected in vitro.  Nostoc sp. ATCC 29131 (PCC 6705) was found to contain a
restriction enzyme, NspBII, with hitherto unknown specificity, C A/C GC T/G G.

<>

<1>Dwivedi, G.R., Sharma, E., Rao, D.N.
<2>Helicobacter pylori DprA alleviates restriction barrier for incoming DNA.
<3>Nucleic Acids Res.
<4>41
<5>3274-3288
<6>2013
<7>Helicobacter pylori is a Gram-negative bacterium that colonizes human stomach and causes
gastric inflammation. The species is naturally competent and displays remarkable diversity.
The presence of a large number of restriction-modification (R-M) systems in this bacterium
creates a barrier against natural transformation by foreign DNA. Yet, mechanisms that protect
incoming double-stranded DNA (dsDNA) from restriction enzymes are not well understood. A
DNA-binding protein, DNA Processing Protein A (DprA) has been shown to facilitate natural
transformation of several Gram-positive and Gram-negative bacteria by protecting incoming
single-stranded DNA (ssDNA) and promoting RecA loading on it. However, in this study, we
report that H. pylori DprA (HpDprA) binds not only ssDNA but also dsDNA thereby conferring
protection to both from various exonucleases and Type II restriction enzymes. Here, we
observed a stimulatory role of HpDprA in DNA methylation through physical interaction with
methyltransferases. Thus, HpDprA displayed dual functional interaction with H. pylori R-M
systems by not only inhibiting the restriction enzymes but also stimulating
methyltransferases. These results indicate that HpDprA could be one of the factors that
modulate the R-M barrier during inter-strain natural transformation in H. pylori.

<>

<1>Dwivedi, V., Sangwan, N., Nigam, A., Garg, N., Niharika, N., Khurana, P., Khurana, J.P., Lal, R.
<2>Draft Genome Sequence of Thermus sp. Strain RL, Isolated from a Hot Water Spring  Located atop the Himalayan Ranges at Manikaran, India.
<3>J. Bacteriol.
<4>194
<5>3534
<6>2012
<7>Thermus sp. strain RL was isolated from a hot water spring (90 degrees C to 98 degrees C) at
Manikaran, Himachal Pradesh, India. Here we report the draft genome
sequence (20,36,600 bp) of this strain. The draft genome sequence consists of 17
contigs and 1,986 protein-coding sequences and has an average G+C content of
68.77%.

<>

<1>Dwyer, P.A., Riftina, F., Agarwal, K.L.
<2>Interaction of HpaI endonuclease with chemically synthesized oligonucleotides.
<3>Fed. Proc.
<4>38
<5>294
<6>1979
<7>The octanucleotide dG-G-T-T-A-A-C-C is cleaved by the restriction endonuclease
HpaI at the phosphodiester bond between thymidine and adenosine.
Octanucleotides, containing modified nucleosides in the recognition sequence
for HpaI, were chemically synthesized by our modified triester method.
Interaction of the enzyme with dG-G-T-U-A-A-C-C, dG-G-T-U(Br)-A-A-C-C, and
dG-I-T-T-A-A-C-C will be described.  An octanucleotide, in which the
phosphodiester bond at the site of cleavage is replaced by its methyl
phosphonate analogue, was also synthesized and is a reversible inhibitor of the
HpaI endonuclease.

<>

<1>Dwyer-Hallquist, P., Kezdy, F.J., Agarwal, K.L.
<2>Interaction of the HpaI endonuclease with synthetic oligonucleotides.
<3>Biochemistry
<4>21
<5>4693-4700
<6>1982
<7>To determine which functional groups of bases within the grooves of
double-helical DNA interact with the HpaI endonuclease, we have employed
chemically synthesized octanucleotides containing base analogues.  The 5-methyl
group of thymine was probed as a contact between the HpaI endonuclease and its
recognition sequence by using the ologonucleotides d(G-G-T-T-A-A-C-C),
d(G-G-T-U-A-A-C-C), and d(G-G-T-B-A-A-C-C).  The 2-amino group of guanine was
probed as a contact for the HpaI endonuclease by using the octanucleotide
d(G-I-T-T-A-A-C-C).  The HpaI endonuclease cleaves octanucleotides
d(G-G-T-T-A-A-C-C) and d(G-G-T-B-A-A-C-C) according to Michaelis-Menten
kinetics.  However, both the Km and turnover number for d(G-G-T-B-A-A-C-C) were
severalfold lower than those for cleave of d(G-G-T-T-A-A-C-C).  In addition,
d(G-G-T-U A-A-C-C) was not cleaved by HpaI endonuclease, suggesting that the
5-methyl group of thymine is a contact between the HpaI endonuclease and its
recognition sequence.  d(G-I-T-T-A-A-C-C) was not cleaved by the HpaI
endonuclease which may be due in part to the low thermal stability of the
duplex.  Nevertheless, our results suggest that the 2-amino group of guanine is
a contact for the HpaI endonuclease.  A phosphate group 5' external to the HpaI
recognition sequence has been identified as a contact between the HpaI
endonuclease and DNA.  The HpaI endonuclease cleaved 5'-phosphorylated
octanucleotide 30-fold faster than unphosphorylated octanucleotide.  In
addition, the Km of the d(G-G-T-T-A-A-C-C) was 8000-fold higher than the Km of
the phage f1 RFI DNA, suggesting that the octanucleotide is too short to take
advantage of the entire DNA binding site of the enzyme.

<>

<1>Dy, L., Chalasani, S., Essani, K.
<2>Isolaton of Escherichia coli mutants lacking methylcytosine-dependent restriction systems for cloning extensively methylated frog virus 3 DNA.
<3>Gene
<4>131
<5>87-91
<6>1993
<7>Many bacterial strains possess methylation-dependent restriction systems (MDRS) that
demonstrate methylcytosine-dependent restriction endonuclease activity for the dinucleotide
sequence, dCpdG. This makes these strains unsuitable for cloning methylated DNA. Some
commercially available bacterial cells are recommended for cloning DNA fragments with
methylated cytosines and adenines, e.g., Escherichia coli DH5-alphaMCR. Our attempts to clone
frog virus 3 (FV3) DNA, which has the highest degree of cytosine methylation ever reported,
using DH5-alphaMCR cells, were not successful. This and other observations suggested the
existence of additional MDRS that have not yet been eliminated from DH5-alphaMCR cells. In
order to isolate a mutant from this bacterial strain that is suitable to clone highly
methylated FV3 DNA, we transformed these cells with a recombinant pUC19 plasmid containing a
methylated 1.4-kb genomic DNA fragment from FV3, and selected for ampicillin (Ap) resistance.
Three such attempts yielded only one colony that contained a fully methylated 1.4-kb FV3
genomic DNA fragment. Furthermore, plasmid-cured Ap-sensitive colonies originating from this
clone were isolated and have been successfully employed to clone the highly methylated FV3
genomic DNA fragment.

<>

<1>Dyachenko, O.V., Tarlachkov, S.V., Marinitch, D.V., Shevchuk, T.V., Buryanov, Y.I.
<2>Expression of exogenous DNA methyltransferases: Application in molecular and cell biology.
<3>Biokhimiia
<4>79
<5>77-87
<6>2014
<7>DNA methyltransferases might be used as powerful tools for studies in molecular and cell
biology due to their ability to recognize and modify nitrogen bases in specific sequences of
the genome. Methylation of the eukaryotic genome using exogenous DNA methyltransferases
appears to be a promising approach for studies on chromatin structure. Currently, the
development of new methods for targeted methylation of specific genetic loci using DNA
methyltransferases fused with DNA-binding proteins is especially interesting. In the present
review, expression of exogenous DNA methyltransferase for purposes of in vivo analysis of the
functional chromatin structure along with investigation of the functional role of DNA
methylation in cell processes are discussed, as well as future prospects for application of
DNA methyltransferases in epigenetic therapy and in plant selection.

<>

<1>Dyall-Smith, M., Pfeiffer, F., Klee, K., Palm, P., Gross, K., Schuster, S.C., Rampp, M., Oesterhelt, D.
<2>Haloquadratum walsbyi: limited diversity in a global pond.
<3>PLoS ONE
<4>6
<5>e20968
<6>2011
<7>Background: Haloquadratum walsbyi commonly dominates the microbial flora of hypersaline
waters. Its cells are extremely fragile squares requiring .14%(w/v) salt for growth,
properties that should limit its dispersal and promote geographical isolation and divergence.
To assess this, the genome sequences of two isolates recovered from sites at near maximum
distance on Earth, were compared.  Principal Findings: Both chromosomes are 3.1 MB in size,
and 84% of each sequence was highly similar to the other (98.6% identity), comprising the core
sequence. ORFs of this shared sequence were completely synteneic (conserved in genomic
orientation and order), without inversion or rearrangement. Strain-specific
insertions/deletions could be precisely mapped, often allowing the genetic events to be
inferred. Many inferred deletions were associated with short direct repeats (4- 20 bp).
Deletion-coupled insertions are frequent, producing different sequences at identical
positions. In cases where the inserted and deleted sequences are homologous, this leads to
variant genes in a common synteneic background (as already described by others). Cas/CRISPR
systems are present in C23T but have been lost in HBSQ001 except for a few spacer remnants.
Numerous types of mobile genetic elements occur in both strains, most of which appear to be
active, and with some specifically targetting others. Strain C23T carries two ,6 kb plasmids
that show similarity to halovirus His1 and to sequences nearby halovirus/plasmid gene clusters
commonly found in haloarchaea. Conclusions: Deletion-coupled insertions show that Hqr. walsbyi
evolves by uptake and precise integration of foreign DNA, probably originating from close
relatives. Change is also driven by mobile genetic elements but these do not by themselves
explain the atypically low gene coding density found in this species. The remarkable genome
conservation despite the presence of active systems for genome rearrangement implies both an
efficient global dispersal system, and a high selective fitness for this species.

<>

<1>Dyall-Smith, M.L., Pfeiffer, F., Oberwinkler, T., Klee, K., Rampp, M., Palm, P., Gross, K., Schuster, S.C., Oesterhelt, D.
<2>Genome of the Haloarchaeon Natronomonas moolapensis, a Neutrophilic Member of a Previously Haloalkaliphilic Genus.
<3>Genome Announcements
<4>1
<5>e0009513
<6>2013
<7>The genus Natronomonas contains two species, one haloalkaliphile (N. pharaonis) and one
neutrophile (N. moolapensis). Here, we report the genome sequence of N.
moolapensis strain 8.8.11. The overall genome properties are similar for the two
species. Only the neutrophile contains bacteriorhodopsin and a membrane
glycolipid.

<>

<1>Dybvig, K., Cao, Z., French, C.T., Yu, H.
<2>Evidence for type III restriction and modification systems in Mycoplasma pulmonis.
<3>J. Bacteriol.
<4>189
<5>2197-2202
<6>2007
<7>Mycoplasma pulmonis possesses a cassette of genes that are predicted to code for type III
restriction and modification (R-M) enzymes. Transposon
disruption of a gene predicted to code for the endonuclease subunit of the
enzyme resulted in loss of R-M activity. Genomic data indicate that the
cassette was acquired by horizontal gene transfer and possibly located on
a mobile element.

<>

<1>Dybvig, K., Sitaraman, R., French, C.T.
<2>A family of phase-variable restriction enzymes with differing specificities generated by high-frequency gene rearrangements.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>13923-13928
<6>1998
<7>The hsd genes of Mycoplasma pulmonis encode restriction and modification enzymes exhibiting a
high degree of sequence similarity to the type I enzymes of enteric bacteria.  The S subunits
of type I systems dictate the DNA sequence specificity of the holoenzyme and are required for
both the restriction and the modification reactions.  The M. pulmonis chromosome has two hsd
loci, both of which contains two hsdS genes each and are complex, site-specific DNA inversion
systems.  Embedded within the coding regions of each hsdS gene are a minimum of three sites at
which DNA inversions occur to generate extensive amino acid sequence variations in the
predicted S subunits.  We show that the polymorphic hsdS genes produced by gene rearrangement
encode a family of functional S subunits with differing DNA sequence specificities.  In
addition to creating polymorphisms in hsdS sequences, DNA inversions regulate the
phase-variable production of restriction activity because the other genes required for
restriction activity (hsdR and hsdM) are expressed only from loci that are oriented
appropriately in the chromosome relative to the hsd promoter.  These data cast doubt on the
prevailing paradigms that restriction systems are either selfish or function to confer
protection from invasion by foreign DNA.

<>

<1>Dybvig, K., Swinton, D., Maniloff, J., Hattman, S.
<2>Cytosine methylation of the sequence GATC in a mycoplasma.
<3>J. Bacteriol.
<4>151
<5>1420-1424
<6>1982
<7>Mycoplasma virus L2 is subject to host-specific restriction and modification in
Acholeplasma laidlawii strains JA1 and K2.  We have examined the DNAs from both
host cells and viruses propagated on these strains with respect to
susceptibility to cleavage by restriction endonucleases and for DNA base
modifications.  We show that, in strain K2 and L2 virus grown on K2 cells,
cytosine in the sequence GATC is methylated to 5-methylcytosine and, although
strain K2 and L2 viruses grown on K2 contain N6-methyladenine in their DNA,
adenine in the sequence GATC is not methylated.  In contrast to K2, strain JA1
and L2 virus grown on JA1 cells contain no detectable methylated bases.  It is
not known which of the methylated bases in K2 is the basis for the K2
restriction-modification system operative on L2 virus.

<>

<1>Dybvig, K., Voelker, L.L.
<2>Molecular biology of mycoplasmas.
<3>Annu. Rev. Microbiol.
<4>50
<5>25-57
<6>1996
<7>Although mycoplasmas lack cell walls, they are in many respects similar to the gram-positive
bacteria with which they share a common ancestor.  The molecular biology of mycoplasmas is
intriguing because the chromosome is uniquely small (<600 kb in some species) and extremely
A-T rich (as high as 75 mol% in some species).  Perhaps to accommodate DNA with a lower G+C
content, most mycoplasmas do not have the "universal" genetic code.  In these species, TGA is
not a stop codon; instead it encodes tryptophan at a frequency 10 times greater than TGG, the
usual codon for this amino acid.  Because of the presence of TGA codons, the translation of
mycoplasmal proteins terminates prematurely when cloned genes are expressed in other
eubacteria, such as Escherichia coli.  Many mycoplasmas possess strikingly dynamic chromosomes
in which high-frequency changes result from errors in DNA repair or replication and from
highly active recombination systems.  Often, high-frequency changes in the mycoplasmal
chromosome are associated with antigenic and phase variation, which regulate the production of
factors critical to disease pathogenesis.

<>

<1>Dybvig, K., Yu, H.
<2>Regulation of a restriction and modification system via DNA inversion in Mycoplasma pulmonis.
<3>Mol. Microbiol.
<4>12
<5>547-560
<6>1994
<7>An invertible DNA element of 6.8kb, designated the hsd1 locus, was identified in the
chromosome of Mycoplasma pulmonis. Infection of host cells with mycoplasma virus P1 revealed
that the organism's restriction and modification (R-M) properties are controlled by inversion
of hsd1. The nucleotide sequence of hsd1 revealed several genes, the predicted amino acids of
which bear striking similarity to the subunits of the type I R-M enzymes previously found only
in enteric bacteria.

<>

<1>Dynan, W., Fox, K., Stoddard, B.
<2>Editorial: NAR Surveys the Past, Present and Future of Restriction Endonucleases.
<3>Nucleic Acids Res.
<4>42
<5>1-2
<6>2014
<7>In this issue, Nucleic Acids Research presents five Survey and Summary articles that describe
the historical development of studies on restriction endonucleases and summarize much of our
current understanding of this diverse and complex group of enzymes. The first of these
articles, entitled 'Highlights of the DNA cutters: a short history of the restriction
enzymes' (1), describes seminal studies on bacteriophage host restriction, details subsequent
work on type I and type III enzymes that established the restriction-modification (RM)
paradigm and summarizes other landmark events that led to restriction enzymes becoming a main
driving force in the development of modern biotechnology and molecular medicine. Other Survey
and Summary articles in this issue describe three of the major types of RM systems as they are
understood today (2-4). The different types of RM systems-of which there are currently four-
vary in their cofactor dependence, in the spatial relationship of DNA binding and cleavage
sites and in the way in which endonuclease and modification activities are physically and
mechanistically coupled to one another. The last of the Survey and Summary articles in this
issue discusses RM systems in the broader context of toxin-antitoxin genetic systems, which
exist in great variety throughout the microbial world (5).

<>

<1>Dyson, P., Evans, M.
<2>Novel post-replicative DNA modification in Streptomyces: analysis of the preferred modification site of plasmid pIJ101.
<3>Nucleic Acids Res.
<4>26
<5>1248-1253
<6>1998
<7>Both Streptomyces lividans and Streptomyces avermitilis have the ability to site  specifically
modify their DNA, rendering it susceptible to in vitro
Tris-dependent double-strand cleavage. We have cloned a 160 bp fragment
containing the preferred modification site of plasmid pIJ101 and, employing an in
vitro primer extension assay, determined that the modifications occur at guanine
residues on either strand separated by 3 bp. These guanines are located within a
6 bp palindromic 'core' sequence. A cloned copy of a 35 bp region of the plasmid
containing this core sequence was not recognized by the modifying activity in
vivo. To further investigate the nature of the site specificity a set of deletion
mutants of the 160 bp sequence were analysed. This revealed that a substantial
portion of this sequence is essential for authentic modification. The essential
region contains three 13 bp direct repeats, the central one containing the core
sequence, while the left-hand and right-hand copies overlap two potential
stem-loop structures. Deletion of either left- or right-hand repeat structures
abolishes modification within the core sequence, although the left-hand deletion
resulted in modification at a secondary site within the right-hand direct repeat.
These data support a post-replicative mechanism of modification, underlined by
the observation that the modifications are not detected in single-stranded
plasmid replication intermediates.

<>

<1>Dziewit, L., Adamczuk, M., Szuplewska, M., Bartosik, D.
<2>DIY series of genetic cassettes useful in construction of versatile = vectors specific for Alphaproteobacteria.
<3>J. Microbiol. Methods
<4>86
<5>166-174
<6>2011
<7>We have developed a DIY (Do It Yourself) series of genetic casset=
tes, which facilitate construction of novel versatile vectors for
Alphaproteobacteria. All the cassettes are based on defined genetic
modules derived from three natural plasmids of Paracoccus aminophilus JCM
7686. We have constructed over 50 DIY cassettes, which differ in structure
and specific features. All of them are functional in eight strains
representing three orders of Alphaproteobacteria: Rhodobacterales,
Rhizobiales and Caulobacterales. Besides various replication and
stabilization systems, many of the cassettes also contain selective
markers appropriate for Alphaproteobacteria (40 cassettes) and genetic
modules responsible for mobilization for conjugal transfer (24 cassettes).
All the DIY cassettes are bordered by different types of polylinkers,
which facilitate vector construction. Using these DIY cassettes, we have
created a set of compatible Escherichia coli-Alphaproteobacteria
mobilizable shuttle vectors (high or low copy number in E. coli), which
will greatly assist the genetic manipulation of Alphaproteobacteria.

<>

<1>Dziewit, L., Cegielski, A., Romaniuk, K., Uhrynowski, W., Szych, A., Niesiobedzki, P., Zmuda-Baranowska, M.J., Zdanowski, M.K., Bartosik, D.
<2>Plasmid diversity in arctic strains of Psychrobacter spp.
<3>Extremophiles
<4>17
<5>433-444
<6>2013
<7>Six strains of Psychrobacter spp. isolated from guano of little auks collected on Spitsbergen
island (Arctic) carried nine plasmids that
were fully sequenced. These replicons (ranging in size from 2917 to
14924 bp) contained either repA (ColE2-type) or repB (iteron-type)
replication systems of a relatively narrow host range, limited to
Psychrobacter spp. All but one of the plasmids carried predicted
mobilization for conjugal transfer systems, encoding relaxases of the
MOBQ, MOBV or MOBP families. The plasmids also contained diverse
additional genetic load, including a type II restriction-modification
system and a gene encoding a putative subunit C of alkyl hydroperoxide
reductase (AhpC)-an antioxidant enzyme and major scavenger of reactive
oxygen species. Detailed comparative sequence analyses, extended to all
plasmids identified so far in psychrophilic bacteria, distinguished
groups of the most ubiquitous replicons, which play a key role in
horizontal gene transfer in cold environments.

<>

<1>Dziewit, L., Kuczkowska, K., Adamczuk, M., Radlinska, M., Bartosik, D.
<2>Functional characterization of the type II PamI restriction-modification system derived from plasmid pAMI7 of Paracoccus aminophilus JCM 7686.
<3>FEMS Microbiol. Lett.
<4>324
<5>56-63
<6>2011
<7>Plasmid pAMI7 of the methylotrophic bacterium Paracoccus aminophilus JCM 7686
(Alphaproteobacteria) encodes a functional type II
restriction-modification (R-M) system designated PamI. Homologous
systems were identified in the genomes of distinct taxonomic groups of
Bacteria and Archaea, which provides evidence that horizontal gene
transfer has contributed to the wide dissemination of R-M modules -
even between domains. Analysis of the cleavage specificity of the R.
PamI endonuclease revealed that this protein is an isoschizomer of
restriction enzyme NcoI. Interestingly, bioinformatic analyses suggest
that R. PamI and NcoI are accompanied by methyltransferases of
different methylation specificities (C5-methylcytosine and
N4-methylcytosine methyltransferases, respectively), which possibly
exemplifies recombinational shuffling of genes coding for individual
components of R-M systems. The PamI system can stabilize plasmid pAMI7
in a bacterial population, most probably at the postsegregational
level. Therefore, it functions in an analogous manner to
plasmid-encoded toxin-antitoxin (TA) systems. Since the TA system of
pAMI7 is nonfunctional, it is highly probable that this lack is
compensated by the stabilizing activity of PamI. This indicates the
crucial role of the analyzed R-M system in the stable maintenance of
pAMI7, which is, to our knowledge, the first report of 'symbiosis'
between a R-M system and a plasmid in the Alphaproteobacteria.

<>

<1>Dziewit, L., Oscik, K., Bartosik, D., Radlinska, M.
<2>Molecular Characterization of a Novel Temperate Sinorhizobium Bacteriophage, Phi LM21, Encoding DNA Methyltransferase with CcrM-Like Specificity.
<3>J. Virol.
<4>88
<5>13111-13124
<6>2014
<7>Phi LM21 is a temperate phage isolated from Sinorhizobium sp. strain LM21
(Alphaproteobacteria). Genomic analysis and electron microscopy suggested that Phi LM21 is a
member of the family Siphoviridae. The phage has an isometric head and a long noncontractile
tail. The genome of Phi LM21 has 50,827 bp of linear double-stranded DNA encoding 72 putative
proteins, including proteins responsible for the assembly of the phage particles, DNA
packaging, transcription, replication, and lysis. Virion proteins were characterized using
mass spectrometry, leading to the identification of the major capsid and tail components, tape
measure, and a putative portal protein. We have confirmed the activity of two gene products, a
lytic enzyme (a putative chitinase) and a DNA methyltransferase, sharing sequence specificity
with the cell cycle-regulating methyltransferase (CcrM) of the bacterial host. Interestingly,
the genome of Sinorhizobium phage Phi LM21 shows very limited similarity to other known phage
genome sequences and is thus considered unique.IMPORTANCEProphages are known to play an
important role in the genomic diversification of bacteria via horizontal gene transfer. The
influence of prophages on pathogenic bacteria is very well documented. However, our knowledge
of the overall impact of prophages on the survival of their lysogenic, nonpathogenic bacterial
hosts is still limited. In particular, information on prophages of the agronomically important
Sinorhizobium species is scarce. In this study, we describe the isolation and molecular
characterization of a novel temperate bacteriophage, Phi LM21, of Sinorhizobium sp. LM21.
Since we have not found any similar sequences, we propose that this bacteriophage is a novel
species. We conducted a functional analysis of selected proteins. We have demonstrated that
the phage DNA methyltransferase has the same sequence specificity as the cell cycle-regulating
methyltransferase CcrM of its host. We point out that this phenomenon of mimicking the host
regulatory mechanisms by viruses is quite common in bacteriophages.

<>

<1>Earl, A.M., Desjardins, C.A., Fitzgerald, M.G., Arachchi, H.M., Zeng, Q., Mehta, T., Griggs, A., Birren, B.W., Toney, N.C., Carr, J., Posey, J., Butler, W.R.
<2>High quality draft genome sequence of Segniliparus rugosus CDC 945(T)= (ATCC BAA-974(T)).
<3>Standards in Genomic Sciences
<4>5
<5>389-397
<6>2011
<7>Segniliparus rugosus represents one of two species in the genus Segniliparus, the sole genus
in the family Segniliparaceae. A unique and interesting feature of
this family is the presence of extremely long carbon-chain length mycolic acids
bound in the cell wall. S. rugosus is also a medically important species because
it is an opportunistic pathogen associated with mammalian lung disease. This
report represents the second species in the genus to have its genome sequenced.
The 3,567,567 bp long genome with 3,516 protein-coding and 49 RNA genes is part
of the NIH Roadmap for Medical Research, Human Microbiome Project.

<>

<1>Earl, A.M., Eppinger, M., Fricke, W.F., Rosovitz, M.J., Rasko, D.A., Daugherty, S., Losick, R., Kolter, R., Ravel, J.
<2>Whole-Genome Sequences of Bacillus subtilis and Close Relatives.
<3>J. Bacteriol.
<4>194
<5>2378-2379
<6>2012
<7>We sequenced four strains of Bacillus subtilis and the type strains for two closely related
species, Bacillus vallismortis and Bacillus mojavensis. We report
the high-quality Sanger genome sequences of B. subtilis subspecies subtilis
RO-NN-1 and AUSI98, B. subtilis subspecies spizizenii TU-B-10(T) and DV1-B-1,
Bacillus mojavensis RO-H-1(T), and Bacillus vallismortis DV1-F-3(T).

<>

<1>Eastberg, J.H., Eklund, J., Monnat, R., Stoddard, B.L.
<2>Mutability of an HNH nuclease imidazole general base and exchange of a deprotonation mechanism.
<3>Biochemistry
<4>46
<5>7215-7225
<6>2007
<7>Several unique protein folds that catalyze the hydrolysis of phosphodiester bonds have arisen
independently in nature, including the
PD(D/E)XK superfamily (typified by type II restriction endonucleases
and many recombination and repair enzymes) and the HNH superfamily
(found in an equally wide array of enzymes, including bacterial
colicins and homing endonucleases). Whereas the identity and position
of catalytic residues within the PD(D/E)XK superfamily are highly
variable, the active sites of HNH nucleases are much more strongly
conserved. In this study, the ability of an HNH nuclease to tolerate a
mutation of its most conserved catalytic residue (its histidine general
base), and the mechanism of the most active enzyme variant, were
characterized. Conversion of this residue into several altered
chemistries, glutamine, lysine, or glutamate, resulted in measurable
activity. The histidine to glutamine mutant displays the highest
residual activity and a pH profile similar to that of the wild-type
enzyme. This activity is dependent on the presence of a neighboring
imidazole ring, which has taken over as a less efficient general base
for the reaction. This result implies that mutational pathways to
alternative HNH-derived catalytic sites do exist but are not as
extensively or successfully diverged or reoptimized in nature as
variants of the PD(D/E)XK nuclease superfamily. This is possibly due to
multiple steric constraints placed on the compact HNH motif, which is
simultaneously involved in protein folding, DNA binding, and catalysis,
as well as the use of a planar, aromatic imidazole group as a general
base.

<>

<1>Eastberg, J.H., Smith, A.M., Zhao, L., Ashworth, J., Shen, B.W., Stoddard, B.L.
<2>Thermodynamics of DNA target site recognition by homing endonucleases.
<3>Nucleic Acids Res.
<4>35
<5>7209-7221
<6>2007
<7>The thermodynamic profiles of target site recognition have been surveyed for homing
endonucleases from various structural families. Similar to
DNA-binding proteins that recognize shorter target sites, homing
endonucleases display a narrow range of binding free energies and
affinities, mediated by structural interactions that balance the magnitude
of enthalpic and entropic forces. While the balance of DeltaH and TDeltaS
are not strongly correlated with the overall extent of DNA bending,
unfavorable DeltaH(binding) is associated with unstacking of individual
base steps in the target site. The effects of deleterious basepair
substitutions in the optimal target sites of two LAGLIDADG homing
endonucleases, and the subsequent effect of redesigning one of those
endonucleases to accommodate that DNA sequence change, were also measured.
The substitution of base-specific hydrogen bonds in a wild-type
endonuclease/DNA complex with hydrophobic van der Waals contacts in a
redesigned complex reduced the ability to discriminate between sites, due
to nonspecific DeltaS(binding).

<>

<1>Eastman, A.W., Weselowski, B., Nathoo, N., Yuan, Z.C.
<2>Complete Genome Sequence of Paenibacillus polymyxa CR1, a Plant Growth-Promoting  Bacterium Isolated from the Corn Rhizosphere Exhibiting Potential for Biocontrol,  Biomass Degradation, and Biofuel Production.
<3>Genome Announcements
<4>2
<5>e01218-13
<6>2014
<7>Here we report the complete genome sequence of the bacterium Paenibacillus polymyxa CR1
(accession no. CP006941), which consists of one circular chromosome
of 6,024,666 bp with 5,283 coding sequences (CDS), 87 tRNAs, and 12 rRNA operons.
Data presented will allow for further insights into the mechanisms underpinning
agriculturally and industrially relevant processes.

<>

<1>Eberhard, J., Oza, J., Reich, N.O.
<2>Cloning, sequence analysis and heterologous expression of the DNA adenine-(N-6) methyltransferase from the human pathogen Actinobacillus actinomycetemcomitans.
<3>FEMS Microbiol. Lett.
<4>195
<5>223-229
<6>2001
<7>We cloned and sequenced the DNA adenine-N-6 methyltransferase gene of the human pathogen
Actinobacillus actinomycetemcomitans (M.AacDAM).
Restriction digestion shows that the enzyme methylates adenine in the
sequence GATC. Expression of the enzyme in a DAM(-) background shows in
vivo activity. A PSI-BLAST search revealed that M.AacDAM is most
related to M.HindIV. M.EcoDAM, M.StyDAM. and M.Small. The ClustalW
alignment shows highly conserved regions in the enzyme characteristic
for type a MTases. Phylogenetic tree analysis shows a cluster of
enzymes recognizing the sequence GATC, within a branch of orphan MTases
harboring M.AacDAM, The cloning and sequencing of this first
methyltransferase gene described for A. actinomycetemcomitans open the
path for studies on the potential regulatory impact of DNA methylation
on gene regulation and virulence in this organism.

<>

<1>Eberhart, L., Deringer, J.R., Brayton, K.A., Sawant, A., Besser, T.E., Call, D.R.
<2>Characterization of a novel microcin that kills enterohemorrhagic E. coli O157:H7 and O26.
<3>Appl. Environ. Microbiol.
<4>78
<5>6592-6599
<6>2012
<7>A novel phenotype was recently identified whereby specific strains of Escherichia
coli inhibit competing E. coli via a mechanism that was designated
"proximity-dependent inhibition" (PDI). PDI-expressing E. coli (PDI(+)) is known
to inhibit susceptible E. coli strains (PDI(-)), including several
enterohemorrhagic (EHEC) and enterotoxigenic (ETEC) strains. In this study every
strain from a genetically diverse panel of E. coli O157:H7 (n=25) and additional
strains of E. coli serovar O26 were susceptible to the PDI phenotype. Live-dead
staining was consistent with inhibition by killing of susceptible cells.
Comparative genome analysis identified the genetic component of PDI, which is
composed of a plasmid-borne (Incl1) operon encoding a putative microcin and
associated genes for transport, immunity, and microcin activation. Transfer of
the plasmid to a PDI(-) strain resulted in transfer of the phenotype and deletion
of the genes within the operon resulted in loss of the inhibition phenotype.
Deletion of chromosomally encoded tolC also resulted in loss of the inhibitory
phenotype and this confirmed that the putative microcin is most likely secreted
via a type I secretion pathway. Deletion of an unrelated plasmid gene did not
affect the PDI phenotype. Quantitative RT-PCR demonstrated that microcin
expression is correlated with logarithmic-phase growth. The ability to inhibit a
diversity of E. coli strains indicates this microcin may influence gut community
composition and could be useful for control of important enteric pathogens.

<>

<1>Eckstein, F.
<2>Action of restriction endonucleases on phosphorothioate DNA.
<3>Biochem. Soc. Trans.
<4>14
<5>204-205
<6>1986
<7>Phosphorothioate analogues of nucleotides possess a number of properties which
make then interesting compounds for biochemists (Eckstein, 1983).  One of these
is the often slow rate of enzyme-catalysed hydrolysis of these analogues in
comparson with the natural compounds.  Thus it has been shown that
phosphorothioate internucleotidic linkages in DNA are either not or only slowly
hydrolysed by exonuclease III and the 5' - 3' exonuclease activity of
polymerase I.  An early observation indicated that at least some restriction
endonucleases might hydrolyse such groups more slowly than the unmodified
linkages (Vosberg & Eckstein, 1982).  As restriction endonucleases have found
wide application in the manipulation of DNA and as this observation might offer
the possibility of blocking specific sites in DNA against hydrolysis by these
enzymes, it was decided to initiate a more detailed investigation on the
interaction of restriction endonucleases with phosphorothioate-containing
oligonucleotides and DNA.

<>

<1>Eckstein, F.
<2>Interaction of DNA containing phosphorothioate groups with restriction enzymes.
<3>Ann. NY Acad. Sci.
<4>471
<5>217-225
<6>1986
<7>Phosphorothioate analogues of nucleotides have become very valuable tools in
enzymology.  The interest in these compounds is based on a variety of
properties that distinguish them from the naturally occurring nucleotides.
These are the generally slow rate of enzymatic hydrolysis of phosphorothioates,
the chirality of the generally slow rate of enzymatic hydrolysis of
phosphorothioates, the chirality of the phosphorous when the phosphorothioate
is linked to two nonidentical groups, and the affinity to metal ions.  The
various applications of phosphorothioate analogues of nucleotides to the study
of enzyme mechanisms have been reviewed extensively in recent years.  In this
article the effect of incorporation of phosphorothioate groups into DNA will be
discussed.

<>

<1>Eckstein, F.
<2>Phosphorothioation of DNA in bacteria.
<3>Nat. Chem. Biol.
<4>3
<5>689-690
<6>2007
<7>A phosphorothioate modificaiton of DNA has been identified in bacteria.  This first observed
alteration of the DNA phosphate backbone opens many questions about themechanism of sulfur
incoporation and the function of this modification.

<>

<1>Eckstein, F., Connolly, B.A., Potter, V.V.L.
<2>Cleavage of phosphorothioate-containing oligonucleotides and DNA by restriction endonucleases.
<3>Abteil. Chem.
<4>8
<5>23-27
<6>1987
<7>The oligonucleotide d(GGsAATTCC) containing the recognition sequence of the
EcoRI restriction endonuclease with a phosphorothioate group at the site of
cleavage was synthesized by two approaches both based on the polymer support
phophoroamidite method.  Only the Rp-diastereomer was cleaved by EcoRI, at a
rate approximately 20 times slower than the all-phosphate-containing octamer.
To study the interaction of restriction endonucleases with phosphorothioate
internucleotide linkages double stranded M13mp2 DNA was prepared in which the
(+)strand contained only phosphate groups but the (-)strand contained phosphate
groups as well as base specifically introduced phosphorothioate groups of the
Rp-configuration.  This hybrid DNA was cleaved by several restriction enzymes.
The results obtained allow the classification of these enzymes into three
groups:  those which produce nicked DNA as an isolatable intermediate, those
where the nicked DNA is the final product and a third where ony linearised DNA
can be detected.  Particularly the second class of enzymes should be of
interest for the manipulation of DNA.

<>

<1>Eckweiler, D., Bunk, B., Sproer, C., Overmann, J., Haussler, S.
<2>Complete Genome Sequence of Highly Adherent Pseudomonas aeruginosa Small-Colony Variant SCV20265.
<3>Genome Announcements
<4>2
<5>e01232-13
<6>2014
<7>The evolution of small-colony variants within Pseudomonas aeruginosa populations  chronically
infecting the cystic fibrosis lung is one example of the emergence of
adapted subpopulations. Here, we present the complete genome sequence of the
autoaggregative and hyperpiliated P. aeruginosa small-colony variant SCV20265,
which was isolated from a cystic fi brosis (CF) patient.

<>

<1>Eddy, S.R., Gold, L.
<2>Artificial mobile DNA element constructed from the EcoRI endonuclease gene.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>1544-1547
<6>1992
<7>There exist several examples of mobile group I introns.  These introns appear
to use a straightforward mechanism to achieve highly site-specific and
efficient insertion into homologous intronless genes.  Because the only
intron-specific function required by the prevailing model for the mechanism  of
intron mobility is the introduction of a site-specific double-stranded break in
the intronless recipient DNA molecule, we reasoned that it should in principle
be possible to construct an artificially mobile element from the gene for the
restriction enzyme EcoRI that is capable of site-specific insertion at rates
near those of authentic mobile introns.  The generality of the mobility
mechanism may enable high-efficiency targeted gene replacements or disruptions
in a variety of organisms.

<>

<1>Eddy, S.R., Gold, L.
<2>The phage T4 nrdB intron: a deletion mutant of a version found in the wild.
<3>Genes Dev.
<4>5
<5>1032-1041
<6>1991
<7>Bacteriophage T4 possesses three self-splicing group I introns. Two of the three introns are
mobile elements; the third, in the gene encoding a subunit of the phage nucleotide reductase
(nrdB), is not mobile. Because intron mobility offers a reasonable explanation for the
paradoxical occurrence of large intervening sequences in a space-efficient eubacterial phage,
it is puzzling that the nrdB intron is not mobile like its compatriots. We have discovered a
larger nrdB intron in a closely related phage, and we infer from comparative sequence data
that the T4 intron is a deletion mutant derived from this larger intron. This larger nrdB
intron encodes an open reading frame of 269 codons, which we have cloned and overexpressed.
The overexpressed protein shows a dsDNA endonuclease activity for the intronless nrdB gene,
typical of mobile introns. Thus, we believe that all three introns of T4 are or were mobile
"infectious introns" and that they have entered into and been maintained in the phage
population by virtue of this efficient mobility.

<>

<1>Edgell, D., Kleinstiver, B., Wolfs, J.M., Wang, L., Kolaczyk, T., McDowell, B., Bogdanove, A.
<2>Genome engineering nucleases derived from GIY-YIG homing endonucleases.
<3>J. Biomol. Struct. Dyn.
<4>31
<5>64-65
<6>2013
<7>Efficient targeted manipulation of complex genomes requires highly specific endonucleases to
generate double-strand breaks at defined locations.  The predominantly engineered nucleases,
zinc-finger nucleases and TAL effector nucleases use the catalytic domain of FokI as the
nuclease portion.  This domain, however, functions as a dimer to nonspecifically cleave DNA
meaning that ZFNs and TALENs must be designed in head-to-head pairs to target a desired
sequence.  To overcome this limitation and expand the toolbox of genome editing reagents, we
used the N-terminal catalytic domain and interdomain linker of the monomeric GIY-YIG homing
endonuclease I-TevI to create I-TevI-zinc-fingers, and I-TevI-TAL effectors.  We also made
I-TevI fusions to LAGLIDADGs homing endonucleases.  All the three fusions showed activity on
model substates on par with ZFNs and TALENs in yeast-based recombination assays.  These
proof-of-concept experiments demonstrate that the catalytic domain of GIY-YIG homing
endonucleases can be targeted to relevant loci by fusing the domain to characterize
DNA-binding platforms.  Recent efforts have focused on improving the Tev-TAL platform by (1)
understanding the spacing requirements between the nuclease cleavage site and the DNA binding
site, (2) probing the DNA binding requirements of the I-TevI linker domain, and (3)
demonstrating activity in mammalian systems.

<>

<1>Edgell, D.R.
<2>Free-standing homing endonucleases of T-even phage: Freeloaders or functionaries?
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Berlin Heidelberg
<4>16
<5>147-160
<6>2005
<7>The ability of group I and II introns, and of inteins, to promote their own mobility to
cognate genes lacking the intron or intein is a property that is now well documented in the
scientific literature.  This so-called homing of introns and inteins is mediated by homing
endonucleases encoded within the genetic elements.  Other chapters within this book have
detailed the different classes of homing endonucleases, how homing endonuclease specifically
recognize their target sequences, and the mechanisms of DNA cleavage.  This chapter deals with
the surprising observation that homing endonucleases are not always found encoded within
introns or inteins, and are often found inserted between genes.  Such free-standing homing
endonucleases are particularly abundant in the well-studied Escherichia coli bacteriophage T4.
In phage T4, the endonucleases themselves are mobile genetic elements, promoting their spread
to related T-even phage genomes that lack the endonuclease gene by a double-strand break
repair pathway.  This process is remarkably similar to endonuclease-mediated intron homing,
and has been termed intronless homing.  Free-standing endonucleases have been identified in
most sequenced genomes, but it is unlikely that all practice intronless homing, as some have
been co-opted by cellular genomes to function in pathways unrelated to endonuclease mobility.
Here, I will focus on free-standing endonucleases of phage genomes, and highlight differences
in DNA-binding and cleavage strategies of intron-encoded versus free-standing endonucleases as
it relates to promoting mobility between genomes.

<>

<1>Edgell, D.R.
<2>Selfish DNA: Homing Endonucleases Find a Home.
<3>Curr. Biol.
<4>19
<5>R115-R117
<6>2009
<7>Self-splicing group I introns come in two flavours - those with a homing endonuclease to
promote mobility of the intron, and those without an endonuclease. How homing endonucleases
and self-splicing introns associate to form a composite selfish genetic element is a question
of long-standing interest. Recent work has revealed that a shared characteristic of both
introns and endonucleases, the targeting of conserved sequences, may provide the impetus for
the evolution of composite mobile genetic elements.

<>

<1>Edgell, D.R., Belfort, M., Shub, D.A.
<2>Barriers to intron promiscuity in bacteria.
<3>J. Bacteriol.
<4>182
<5>5281-5289
<6>2000
<7>The first bacterial intron, a self-splicing group I intron, was found to interrupt the
thymidylate synthase (td) gene of the Escherichia coli phage T4.  The second and third
bacterial group I introns were found to interrupt the aerobic (nrdB) and anaerobic (nrdD
[initially named sunY]) ribonucleotide reductases of phage T4, and another group I intron was
soon discovered in the DNA polymerase gene of SPO1, a Bacillus phage.  From this (admittedly)
small sampling of phage genomes, one might have naively expected that group I introns would be
abundant in phage or bacterial genomes, especially since subsequent laboratory experiments
demonstrated that group I introns could propagate themselves (by a process called homing)
throughout populations of intron-minus alleles with near 100% efficiency.  That a similar
homing phenomenon had also been previously demonstrated for a group I intron in the large rRNA
gene of yeast mitochondrial gave additional support to the notion that group I introns should
be able to spread efficiently throughout populations.  However, this expected outcome has
never been realized in natural phage populations; some phage populations harbor many introns,
while other related phage populations harbor many introns, while other related phage
populations are strangely lacking in any introns whatsoever.  Why do group I introns have an
unusual distribution in phage and bacterial genomes, and what potential barriers might exist
to prevent their spread?

<>

<1>Edgell, D.R., Derbyshire, V., Van Roey, P., LaBonne, S., Stanger, M.J., Li, Z., Boyd, T.M., Shub, D.A., Belfort, M.
<2>Intron-encoded homing endonuclease I-TevI also functions as a transcriptional autorepressor.
<3>Nat. Struct. Mol. Biol.
<4>11
<5>936-944
<6>2004
<7>Customary binding sites of intron-encoded homing endonucleases lie within cognate intronless
alleles, at the so-called homing sites. Here,
we describe a novel, high-affinity binding site for I-TevI
endonuclease, encoded within the group I td intron of phage T4. This
site is an operator that overlaps the T4 late promoter, which drives
I-TevI expression from within the td intron. I-TevI binds the operator
and homing sites with equal affinity, and functions as a
transcriptional autorepressor. Distinct sequence and spacing
requirements of the catalytic domain result in reduced cleavage
activity on operator DNA. Crystallographic studies showed that the
overall interactions of the DNA-binding domain with the operator and
homing sites are similar, but have some different hydrogen-bonding
contacts. We present a model in which the flexibility in protein-DNA
interactions allows I-TevI to bind variant intronless alleles to
promote intron mobility while facilitating its function in
autorepression, and thereby persistence in its host.

<>

<1>Edgell, D.R., Fast, N.M., Doolittle, W.F.
<2>Selfish DNA: The best defense is a good offense.
<3>Curr. Biol.
<4>6
<5>385-388
<6>1996
<7>The recent discovery of novel biochemical activities of intron-encoded endonucleases
emphasizes the selfish nature of mobile genetic elements.

<>

<1>Edgell, D.R., Gibb, E.A., Belfort, M.
<2>Mobile DNA elements in T4 and related phages.
<3>Virol. J.
<4>7
<5>290
<6>2010
<7>Mobile genetic elements are common inhabitants of virtually every genome where they can exert
profound influences on genome structure and function in addition to promoting their own spread
within and between genomes. Phage T4 and related phage have long served as a model system for
understanding the molecular mechanisms by which a certain class of mobile DNA, homing
endonucleases, promote their spread. Homing endonucleases are site-specific DNA endonucleases
that initiate mobility by introducing double-strand breaks at defined positions in genomes
lacking the endonuclease gene, stimulating repair and recombination pathways that mobilize the
endonuclease coding region. In phage T4, homing endonucleases were first discovered as encoded
within the self-splicing td, nrdB and nrdD introns of T4. Genomic data has revealed that
homing endonucleases are extremely widespread in T-even-like phage, as evidenced by the
astounding fact that similar to 11% of the T4 genome encodes homing endonuclease genes, with
most of them located outside of self-splicing introns. Detailed studies of the mobile td
intron and its encoded endonuclease, I-TevI, have laid the foundation for genetic, biochemical
and structural aspects that regulate the mobility process, and more recently have provided
insights into regulation of homing endonuclease function. Here, we summarize the current state
of knowledge regarding T4-encoded homing endonucleases, with particular emphasis on the
td/I-TevI model system. We also discuss recent progress in the biology of free-standing
endonucleases, and present areas of future research for this fascinating class of mobile
genetic elements.

<>

<1>Edgell, D.R., Shub, D.A.
<2>Related homing endonucleases I-BmoI and I-TevI use different strategies to cleave homologous recognition sites.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>7898-7903
<6>2001
<7>A typical homing endonuclease initiates mobility of its group I intron by recognizing DNA both
upstream and downstream of the intron insertion
site of intronless alleles, preventing the endonuclease from binding
and cleaving its own intron-containing allele. Here, we describe a
GIY-YIG family homing endonuclease. I-BmoI, that possesses an unusual
recognition sequence, encompassing 1 base pair upstream but 38 base
pairs downstream of the intron insertion site. I-BmoI binds
intron-containing and intronless substrates with equal affinity but can
nevertheless discriminate between the two for cleavage. I-BmoI is
encoded by a group I intron that interrupts the thymidylate synthase
CTS) gene (thyA) of Bacillus mojavensis s87-18. This intron resembles
one inserted 21 nucleotides further downstream in a homologous TS gene
ltd) of Escherichia coli phage T4. I-TevI, the T4 td intron-encoded
GIY-YIG endonuclease, is very similar to I-BmoI, but each endonuclease
gene is inserted within a different position of its respective intron.
Remarkably, I-TevI and I-BmoI bind a homologous stretch of TS-encoding
DNA and cleave their intronless substrates in very similar positions.
Our results suggest that each endonuclease has independently evolved
the ability to distinguish intron-containing from intronless alleles
while maintaining the same conserved recognition sequence centered on
DNA-encoding active site residues of TS.

<>

<1>Edgell, D.R., Stanger, M.J., Belfort, M.
<2>Importance of a single base pair for discrimination between intron-containing and intronless alleles by endonuclease I-Bmol.
<3>Curr. Biol.
<4>13
<5>973-978
<6>2003
<7>Homing endonucleases initiate mobility of their host group I introns by binding to and
cleaving lengthy recognition sequences that are typically
centered on the intron insertion site (IS) of intronless alleles. Because
the intron interrupts the endonucleases' recognition sequence,
intron-containing alleles are immune to cleavage by their own
endonuclease. I-TevI and I-BmoI are related GIY-YIG endonucleases that
bind a homologous stretch of thymidylate synthase (TS)-encoding DNA but
use different strategies to distinguish intronless from intron-containing
substrates. I-TevI discriminates between substrates at the level of DNA
binding, as its recognition sequence is centered on the intron IS. I-BmoI,
in contrast, possesses a very asymmetric recognition sequence with respect
to the intron IS, binds both intron-containing and intronless TS-encoding
substrates, but efficiently cleaves only intronless substrate. Here, we
show that I-BmoI is extremely tolerant of multiple substitutions around
its cleavage sites and has a low specific activity. However, a single G-C
base pair, at position -2 of a 39-base pair recognition sequence, is a
major determinant for cleavage efficiency and distinguishes intronless
from intron-containing alleles. Strikingly, this G-C base pair is
universally conserved in phylogenetically diverse TS-coding sequences;
this finding suggests that I-BmoI has evolved exquisite cleavage
requirements to maximize the potential to spread to variant intronless
alleles, while minimizing cleavage at its own intron-containing allele.

<>

<1>Edgell, D.R., Stanger, M.J., Belfort, M.
<2>Coincidence of cleavage sites of intron endonuclease I-TevI and critical sequences of the host thymidylate synthase gene.
<3>J. Mol. Biol.
<4>343
<5>1231-1241
<6>2004
<7>To maximize spread of their host intron or intein, many homing endonucleases recognize
nucleotides that code for important and
conserved amino acid residues of the target gene. Here, we examine the
cleavage requirements for I-TevI, which binds a stretch of thymidylate
synthase (TS) DNA that codes for functionally critical residues in the
TS active site. Using an in vitro selection scheme, we identified two
base-pairs in the I-TevI cleavage site region as important for cleavage
efficiency. These were confirmed by comparison of I-TevI cleavage
efficiencies on mutant and on wild-type substrates. We also showed that
nicking of the bottom strand by I-TevI is not affected by mutation of
residues surrounding the bottom-strand cleavage site, unlike other
homing endonucleases. One of these two base-pairs is universally
conserved in all TS sequences, and is identical with a previously
identified cleavage determinant of I-BmoI, a related GIY-YIG
endonuclease that binds a homologous stretch of TS-encoding DNA. The
other base-pair is conserved only in a subset of TS genes that includes
the I-TevI, but not the I-BmoI, target sequence. Both the I-TevI and
I-BmoI cleavage site requirements correspond to functionally critical
residues involved in an extensive hydrogen bond network within the TS
active site. Remarkably, these cleavage requirements correlate with TS
phylogeny in bacteria, suggesting that each endonuclease has
individually adapted to efficiently cleave distinct TS substrates.

<>

<1>Edgell, M.H., Hutchison, C.A. III, Sclair, M.
<2>Specific endonuclease R fragments of bacteriophage PhiX174 deoxyribonucleic acid.
<3>J. Virol.
<4>9
<5>574-582
<6>1972
<7>The restriction enzyme from Hemophilus influenzae, endonuclease R, cleaves
PhiX174 replicative-form deoxyribonucleic acid (DNA) into at least 13 specific
limit fragments.  The molecular weights of 12 of the fragments have been
estimated by gel electrophoresis and electron microscopy.  Using the genetic
assay for small fragments of PhiX DNA, we have shown that we can salvage
markers from the endonuclease R PhiX-RF fragments.

<>

<1>Edmonds, P., Hall, B.M., Edwards, W.R., Hartline, K.M.
<2>Presence of methylated adenine in GATC Sequences in chromosomal DNAs from Campylobacter species.
<3>J. Bacteriol.
<4>174
<5>8156-8157
<6>1992
<7>We digested chromosomal DNAs from 12 Campylobacter strains (C. jejuni, 4 strains; C. coli, 2
strains; C. fetus subsp. fetus, 2 strains; C. hyointestinalis, 2 strains; and C. upsaliensis,
2 strains) and from 4 Helicobacter strains (H. pylori, 2 strains: and H. mustelae, 2 strains)
with HindIII, SstI, BamHI, DpnI, MboI, and Sau3AI. Restriction fragments were then separated
by electrophoresis in 1% agarose or 10% polyacrylamide gels. Only DNAs from three
Campylobacter species (C. jejuni, C. coli, and C. upsaliensis) were digested with DpnI (an
enzyme that recognizes only methylated adenine in GATC sequences). We used MboI and Sau3AI to
confirm these findings.

<>

<1>Edouard, S., Bibi, F., Dhamodharan, R., Lagier, J.C., Azhar, E.I., Robert, C., Caputo, A., Yasir, M., Jiman-Fatani, A.A., Alawi, M., Fournier, P.E., Raoult, D.
<2>Non-contiguous finished genome sequence and description of Corynebacterium jeddahense sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>987-1002
<6>2014
<7>Corynebacterium jeddahense sp. nov., strain JCB(T), is the type strain of Corynebacterium
jeddahense sp. nov., a new species within the genus
Corynebacterium. This strain, whose genome is described here, was isolated from
fecal flora of a 24-year-old Saudi male suffering from morbid obesity.
Corynebacterium jeddahense is a Gram-positive, facultative anaerobic,
nonsporulating bacillus. Here, we describe the features of this bacterium,
together with the complete genome sequencing and annotation, and compare it to
other member of the genus Corynebacterium. The 2,472,125 bp-long genome (1
chromosome but not plasmid) contains 2,359 protein-coding and 53 RNA genes,
including 1 rRNA operon.

<>

<1>Edouard, S., Sankar, S., Dangui, N.P., Lagier, J.C., Michelle, C., Raoult, D., Fournier, P.E.
<2>Genome sequence and description of Nesterenkonia massiliensis sp. nov. strain NP1(T.).
<3>Standards in Genomic Sciences
<4>9
<5>866-882
<6>2014
<7>Nesterenkonia massiliensis sp. nov., strain NP1(T), is the type strain of Nesterenkonia
massiliensis sp. nov., a new species within the genus
Nesterenkonia. This strain, whose genome is described here, was isolated from the
feces of a 32-year-old French woman suffering from AIDS and living in Marseille.
Nesterenkonia massiliensis is a Gram-positive aerobic coccus. Here, we describe
the features of this bacterium, together with the complete genome sequencing and
annotation. The 2,726,371 bp long genome (one chromosome but no plasmid) contains
2,663 protein-coding and 51 RNA genes, including 1 rRNA operon.

<>

<1>Edwards, A. et al.
<2>Novel essential bacterial polypeptides.
<3>International Patent Office
<4>WO 2004058809 A
<5>
<6>2004
<7>The present invention relates to polypeptide targets for pathogenic bacteria.  The invention
also provides biochemical and biophysical characteristics of those polypeptides.

<>

<1>Edwards, R.A., Helm, R.A., Maloy, S.R.
<2>Increasing DNA transfer efficiency by temporary inactivation of host restriction.
<3>Biotechniques
<4>26
<5>892-900
<6>1999
<7>E. coli and Salmonella typhimurium are widely used bacterial hosts for genetic manipulation of
DNA from prokaryotes and eukaryotes.  Introduction of foreign DNA by electroporation or
transduction into E. coli and Salmonella is limited by host restriction of incoming DNA by the
recipient cells.  Here, we describe a simple method that temporarily inactivates host
restriction, allowing high-frequency DNA transfer.  This technique might be readily applied to
a wide range of bacteria to increase DNA transfer between strains and species.

<>

<1>Ee, R., Lim, Y.L., Yin, W.F., Chan, K.G.
<2>De Novo Assembly of the Quorum-Sensing Pandoraea sp. Strain RB-44 Complete Genome Sequence Using PacBio Single-Molecule Real-Time Sequencing Technology.
<3>Genome Announcements
<4>2
<5>e00245-14
<6>2014
<7>We report the first complete genome sequence of Pandoraea sp. strain RB-44, which was found to
possess quorum-sensing properties. To the best of our knowledge, this is the first
documentation of both a complete genome sequence and quorum-sensing properties of a Pandoraea
species.

<>

<1>Efimova, E.P., Delver, E.P., Belogurov, A.A.
<2>Alleviation of type I restriction in adenine methylase (dam) mutants of Escherichia coli.
<3>Mol. Gen. Genet.
<4>214
<5>313-316
<6>1988
<7>The host-controlled EcoKI-restriction of unmodified phage lambda.O is alleviated in dam
mutants of Escherichia coli by 100- to 300-fold. In addition, the EcoKI modification activity
is substantially decreased in dam- strains. We show that type I restriction (EcoBI, EcoDI and
EcoKI) is detectably alleviated in dam mutants. However, no relief of EcoRI restriction (Type
II) occurs in dam- strains and only a slight effect of dam mutation on EcoPI restriction (Type
III) is observed. We interpret the alleviation of the type I restriction in dam- strains to be
a consequence of induction of the function which interferes with type I restriction systems.

<>

<1>Efimova, E.P., Delver, E.P., Belogurov, A.A.
<2>2-aminopurine and 5-bromouracil induce alleviation of type I restriction in Escherichia coli:  Mismatches function as inducing signals?
<3>Mol. Gen. Genet.
<4>214
<5>317-320
<6>1988
<7>The EcoKI restriction of unmodified phage lambda is 1000-fold alleviated in Escherichia coli
grown in the presence of base analogs 2-aminopurine (2AP) and 5-bromouracil (5BU). 2AP
treatment of bacteria affects specifically the type I restriction systems (EcoAI, EcoBI, EcoDI
and EcoKI) and does not influence type II (EcoRI) and type III (EcoP1) restriction.
2AP-induced alleviation of restriction occurs in bacteria which are deficient in the SOS
response (recA and lexA) and mismatch repair (mutH, mutL and mutS) and can be distinguished
from the alleviation of restriction observed in dam-strains. We suggest that mismatches
induced by 2AP and 5BU may function as an inducing signal for the alleviation of restriction
observed in the presence of base analogs.

<>

<1>Egas, C., Barroso, C., Froufe, H.J., Pacheco, J., Albuquerque, L., da Costa, M.S.
<2>Complete genome sequence of the Radiation-Resistant bacterium Rubrobacter radiotolerans RSPS-4.
<3>Standards in Genomic Sciences
<4>9
<5>1062-1075
<6>2014
<7>Rubrobacter radiotolerans strain RSPS-4 is a slightly thermophilic member of the  phylum
'Actinobacteria' isolated from a hot spring in Sao Pedro do Sul, Portugal.
This aerobic and halotolerant bacterium is also extremely resistant to gamma and
UV radiation, which are the main reasons for the interest in sequencing its
genome. Here, we present the complete genome sequence of strain RSPS-4 as well as
its assembly and annotation. We also compare the gene sequence of this organism
with that of the type strain of the species R. radiotolerans isolated from a hot
spring in Japan. The genome of strain RSPS-4 comprises one circular chromosome of
2,875,491 bp with a G+C content of 66.91%, and 3 circular plasmids of 190,889 bp,
149,806 bp and 51,047 bp, harboring 3,214 predicted protein coding genes, 46 tRNA
genes and a single rRNA operon.

<>

<1>Ehara, A., Suzuki, H., Kanesaki, Y., Yoshikawa, H., Amachi, S.
<2>Draft genome sequence of strain q-1, an iodide-oxidizing alphaproteobacterium isolated from natural gas brine water.
<3>Genome Announcements
<4>2
<5>e00659-14
<6>2014
<7>Here we report the draft genome sequence of strain Q-1, an iodide (I(-))-oxidizing
heterotrophic bacterium in the class Alphaproteobacteria
isolated from natural gas brine water. The genome sequence contained a
multicopper oxidase gene probably responsible for iodide oxidation. A
photosynthetic gene cluster was found but genes for carbon-fixation were absent.

<>

<1>Ehbrecht, H.-J., Pingoud, A., Urbanke, C., Maass, G., Gualerzi, C.
<2>Linear diffusion of restriction endonucleases on DNA.
<3>J. Biol. Chem.
<4>260
<5>6160-6166
<6>1985
<7>We have investigated the dependence of the rate of cleavage of DNA by EcoRI,
HindIII, and BamHI on the chain length of the substrate.  In order to keep the
influence of flanking sequences and of nonspecific binding identical for all
substrates we have carried out all experiments with the same plasmid DNA which
had been digested previously with a variety of different restriction enzymes to
give a set of substrates of different lengths.  Our results show that depending
on the buffer conditions long substrates are cleaved faster than small ones.
We interpret these findings to mean that under certain conditions a linear
diffusion of the enzymes on the DNA is involved in localizing the recognition
sites.  For EcoRI the mean diffusion length is approximately 1000 base pairs at
1 mM MgCl2 which can be shown by diffusion theory to correspond to a linear
diffusion coefficient of 5.10-10 cm2 s-1.  At 10 mM MgCl2 the linear diffusion
of EcoRI is negligible and does not lead to a significant enhancement of the
rate of site localization.  In the presence of nonsaturating amounts of one of
the prokaryotic histone-like protein Hu (NS 2) small and large DNa substrate
are cleaved with identical rate by EcoRI indicating that other proteins bound
to the DNA constitute a barrier across wich linear diffusion cannot take place.
We conclude that linear diffusion, albeit detectable under certain conditions
in vitro, probably is of little importance for the process of site localization
in vivo.

<>

<1>Ehrlich, M., Ehrlich, K., Mayo, J.A.
<2>Unusual properties of the DNA from Xanthomonas phage XP-12 in which 5-methylcytosine completely replaces cytosine.
<3>Biochim. Biophys. Acta
<4>395
<5>109-119
<6>1975
<7>Xanthomonas phage XP-12 contains 5-methylcytosine completely replacing
cytosine.  This substitution confers several unusual properties upon XP-12 DNA.
The buoyant density of XP-12 DNA in CsCl gradients is 1.710 g/cm3, 0.016 g/cm3
lower than that expected for a normal DNA with the same percentage of adenine
plus thymine.  The melting temperature for XP-12 DNA in 0.012 M Na+ is the
highest reported for any naturally occurring DNA, 83.2C, 6.1C higher than that
of normal DNAs with the same percentage of adenine plus thymine.  Unlike the
minor amounts of 5-methylcytosine found in most plant and animal DNAs, the
5-methylcytosine residues of XP-12 derive their methyl group from the 3-carbon
of serine instead of from the thiomethyl carbon of methionine.

<>

<1>Ehrlich, M., Gama-Sosa, M.A., Carreira, L.H., Ljungdahl, L.G., Kuo, K.C., Gehrke, C.W.
<2>DNA methylation in thermophilic bacteria: N4-methylcytosine, 5-methylcytosine, and N6-methyladenine.
<3>Nucleic Acids Res.
<4>13
<5>1399-1412
<6>1985
<7>While determining the minor and major base composition of the DNA from 17 types of
thermophilic bacteria by high performance liquid chromatography (HPLC) of enzymatic digests,
we have discovered a novel base, N4-methylcytosine (m4C). Its structure was proven by
comparison of the DNA-derived nucleoside to the analogous authentic compound by HPLC, UV
spectroscopy, and mass spectroscopy. Eight of the bacterial DNAs contained m4C. Only two
contained the common minor base, 5-methylcytosine (m5C), and neither of these was from an
extreme thermophile. The other prevalent modified base of bacterial DNA, N6-methyladenine
(m6A), was found in nine of the DNAs. Restriction analysis revealed that four of the DNAs had
dam-type (Gm6ATC) methylation patterns. Due to the propensity of m5C residues to be deaminated
by heat to thymine residues and to inefficient repair of the resulting mismatched base pairs,
thermophiles with optimal growth temperatures of >60C generally may avoid having m5C in their
genomes. Instead, some of them have deamination-resistant m4C residues.

<>

<1>Ehrlich, M., Wang, R.Y.-H.
<2>5-Methylcytosine in eukaryotic DNA.
<3>Science
<4>212
<5>1350-1357
<6>1981
<7>A small portion of the cytosine residues in the DNA of higher eukaryotes as
well as in that of many lower eukaryotes is methylated.  The resulting
5-methylcytosine residues occur in specific sequences in the DNA, usually
adjacent to guanine residues on the 3' side.  This methylation of eukaryotic
DNA has been proposed to function in ways, including control of transcription,
maintenance of chromosome structure, repair of DNA, establishment of preferred
sites for mutation, oncogenic transformation, and, in certain systems,
protection of DNA against enzymatic degradation.

<>

<1>Ehrlich, M., Wilson, G.G., Kenneth, C.K., Gehrke, C.W.
<2>N4-Methylcytosine as a minor base in bacterial DNA.
<3>J. Bacteriol.
<4>169
<5>939-943
<6>1987
<7>The DNA base composition, including the minor base content, of 26 strains of
bacteria was determined.  The studied bacteria are sources of widely used
restriction endonucleases.  Approximately 35% of the bacterial DNAs contained
N4-methylcytosine, about 60% contained 5-methylcytosine, and about 90% had
N6-methyladenine.

<>

<1>Eickbush, T.H.
<2>Mobile introns: Retrohoming by complete reverse splicing.
<3>Curr. Biol.
<4>9
<5>R11-R14
<6>1999
<7>A mobile bacterial group II intron can integrate into DNA by the reverse splicing into a
target site of its RNA transcript, which then acts as a template for DNA synthesis by an
encoded reverse transcriptase.  Mobility does not require homologous recombination, which has
important practical and evolutionary implications.

<>

<1>Eidam, C., Poehlein, A., Brenner, M.G., Kadlec, K., Liesegang, H., Brzuszkiewicz, E., Daniel, R., Sweeney, M.T., Murray, R.W., Watts, J.L., Schwarz, S.
<2>Complete Genome Sequence of Mannheimia haemolytica Strain 42548 from a Case of Bovine Respiratory Disease.
<3>Genome Announcements
<4>1
<5>e00318-13
<6>2013
<7>Mannheimia haemolytica is the major bacterial component in the bovine respiratory disease
complex, which accounts for considerable economic losses to the cattle
industry worldwide. The complete genome sequence of M. haemolytica strain 42548
was determined. It has a size of 2.73 Mb and contains 2,888 genes, including
several antibiotic resistance genes.

<>

<1>Eigner, J., Block, S.
<2>Host-controlled restriction of T-even bacteriophages:  relation of four bacterial deoxyribonucleases to restriction.
<3>J. Virol.
<4>2
<5>320-326
<6>1968
<7>Escherichia coli strains B and K-12, which restrict growth of nonglucosylated
T-even phage (T*phage), and nonrestricting strains (Shigella sonnei and mutants
of E. coli B) were tested for levels of endonuclease I and exonucleases I,II,
and III, by means of in vitro assays.  Cell-free extracts freed from
deoxyribonucleic acid (DNA) were examined with three substrates: E. coli DNA,
T*2 DNA, and T2 DNA.  Both restricting and nonrestricting strains had
comparable levels of the four nuclease activities and had similar patterns of
preference for the three substrates.  In addition, mutants of E. coli B and
K-12 that lack endonuclease I were as effective as their respective wild types
in restricting T* phage.

<>

<1>Eijkelkamp, B.A., Stroeher, U.H., Hassan, K.A., Papadimitrious, M.S., Paulsen, I.T., Brown, M.H.
<2>Adherence and motility characteristics of clinical Acinetobacter baumannii isolates.
<3>FEMS Microbiol. Lett.
<4>323
<5>44-51
<6>2011
<7>Acinetobacter baumannii continues to be a major health problem especially in
hospital settings. Herein, features that may play a role in persistence and
disease potential were investigated in a collection of clinical A. baumannii
strains from Australia. Twitching motility was found to be a common trait in A.
baumannii international clone I strains and in abundant biofilm formers, whereas
swarming motility was only observed in isolates not classified within the
international clone lineages. Bioinformatic analysis of the type IV fimbriae
revealed a correlation between PilA sequence homology and motility. A high level
of variability in adherence to both abiotic surfaces and epithelial cells was
found. We report for the first time the motility characteristics of a large
number of A. baumannii isolates and present a direct comparison of A. baumannii
binding to nasopharyngeal and lung epithelial cells.

<>

<1>Eikmeyer, F., Hadiati, A., Szczepanowski, R., Wibberg, D., Schneiker-Bekel, S., Rogers, L.M., Brown, C.J., Top, E.M., Puhler, A., Schluter, A.
<2>The complete genome sequences of four new IncN plasmids from wastewater treatment plant effluent provide new insights into IncN plasmid diversity and evolution.
<3>Plasmid
<4>68
<5>13-24
<6>2012
<7>The dissemination of antibiotic resistance genes among bacteria often occurs by means of
plasmids. Wastewater treatment plants (WWTP) were previously recognized as hot spots for the
horizontal transfer of genetic material. One of the plasmid groups that is often associated
with drug resistance is the incompatibility group IncN. The aim of this study was to gain
insights into the diversity and evolutionary history of IncN plasmids by determining and
comparing the complete genome sequences of the four novel multi-drug resistance plasmids
pRSB201, pRSB203, pRSB205 and pRSB206 that were exogenously isolated from the final effluent
of a municipal WWTP. Their sizes range between 42,875bp and 56,488bp and they share a common
set of backbone modules that encode plasmid replication initiation, conjugative transfer, and
plasmid maintenance and control. All plasmids are transferable at high rates between
Escherichia coli strains, but did not show a broad host range. Different genes conferring
resistances to ampicillin, streptomycin, spectinomycin, sulfonamides, tetracycline and
trimethoprim were identified in accessory modules inserted in these plasmids. Comparative
analysis of the four WWTP IncN plasmids and IncN plasmids deposited in the NCBI database
enabled the definition of a core set of backbone genes for this group. Moreover, this approach
revealed a close phylogenetic relationship between the IncN plasmids isolated from
environmental and clinical samples. Phylogenetic analysis also suggests the existence of
host-specific IncN plasmid subgroups. In conclusion, IncN plasmids likely contribute to the
dissemination of resistance determinants between environmental bacteria and clinical strains.
This is of particular importance since multi-drug resistance IncN plasmids have been
previously identified in members of the Enterobacteriaceae that cause severe infections in
humans.

<>

<1>Einvik, C., Decatur, W.A., Embley, T.M., Vogt, V.M., Johansen, S.
<2>Naegleria nucleolar introns contain two group I ribozymes with different functions in RNA splicing and processing.
<3>RNA
<4>3
<5>710-720
<6>1997
<7>We have characterized the structural organization and catalytic properties of the large
nucleolar group I introns (NaSSU1) of different Naegleria species N. jamiesoni, N. andersoni,
N. italica, and N. gruberi.  NaSSU1 consists of three distinct RNA domains: an open reading
frame encoding a homing-type endonuclease, and a small group I ribozyme (NaGIR1) inserted into
the P6 loop of a second group I ribozyme (NaGIR2).  The two ribozymes have different functions
in RNA splicing and processing.  NaGIR1 is an unusual self-cleaving group I ribozyme
responsible for intron processing at two internal sites (IPS1 and IPS2), both close to the 5'
end of the open reading frame.  This processing is hypothesized to lead to formation of a
messenger RNA for the endonuclease.  Structurally, NaGIR2 is a typical group IC1 ribozyme,
catalyzing intron excision and exon ligation reactions.  NaGIR2 is responsible for
circularization of the excised intron, a reaction that generates full-length RNA circles of
wild-type intron.  Although it is only distantly related in primary sequence, NaSSU1 RNA has a
predicted organization and function very similar to that of the mobile group I intron DiSSU1
of Didymium, the only other group I intron known to encode two ribozymes.  We propose that
these twin-ribozyme introns define a distinct category of group I introns with a conserved
structural organization and function.

<>

<1>Eisen, J.A. et al.
<2>The complete genome sequence of Chlorobium tepidum TLS, a photosynthetic, anaerobic, green-sulfur bacterium.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>9509-9514
<6>2002
<7>The complete genome of the green-sulfur eubacterium Chlorobium tepidum TLS was determined to
be a single circular chromosome of 2,154,946 bp. This represents the first genome sequence
from the phylum Chlorobia, whose members perform anoxygenic photosynthesis by the reductive
tricarboxylic acid cycle. Genome comparisons have identified genes in C. tepidum that are
highly conserved among photosynthetic species. Many of these have no assigned function and may
play novel roles in photosynthesis or photobiology. Phylogenomic analysis reveals likely
duplications of genes involved in biosynthetic pathways for photosynthesis and the metabolism
of sulfur and nitrogen as well as strong similarities between metabolic processes in C.
tepidum and many Archaeal species.

<>

<1>Eisenschmidt, K.
<2>Development of a programmable restriction enzyme.
<3>Ph.D. Thesis, Germany
<4>
<5>
<6>2005
<7>
<>

<1>Eisenschmidt, K., Lanio, T., Jeltsch, A., Pingoud, A.
<2>A fluorimetric assay for on-line detection of DNA cleavage by restriction endonucleases.
<3>J. Biotechnol.
<4>96
<5>185-191
<6>2002
<7>We have developed an assay for online detection of DNA cleavage by restriction endonucleases,
suitable for the high throughput screening of the activity and flanking sequence preference of
restriction endonuclease variants. For this purpose oligodeoxynucleotides were used, labeled
with either 6-FAM or TAMRA whose fluorescence is quenched by a neighboring DABCYL group. After
endonucleolytic cleavage the products are too short to remain double-stranded and the
fluorophor labeled strand is released with concomitant increase in fluorescence which can be
easily quantified.
Employing this method, cleavage reactions can be monitored continuously, allowing for fast
detection of specific activity as well as determination of kinetic parameters. To demonstrate
the reliability of our
assay we measured K(M) and k(cat) values for the restriction endonuclease EcoRV and obtained
results similar to those obtained with established assays. Moreover, our method makes it
possible to observe
the cleavage of two different substrates differing in the sequences flanking the EcoRV site
and labeled with different fluorophors in competition in a single experiment. This assay can
be carried out in a
microplate format, which allows for the analysis of many restriction endonuclease variants in
parallel.

<>

<1>Eisenschmidt, K., Lanio, T., Simoncsits, A., Jeltsch, A., Pingoud, V., Wende, W., Pingoud, A.
<2>Developing a programmed restriction endonuclease for highly specific DNA cleavage.
<3>Nucleic Acids Res.
<4>33
<5>7039-7047
<6>2005
<7>Specific cleavage of large DNA molecules at few sites, necessary for the analysis of genomic
DNA or for targeting individual genes in complex genomes, requires endonucleases of extremely
high specificity. Restriction endonucleases (REase) that recognize DNA sequences of 4-C8 bp
are not sufficiently specific for this purpose. In principle, the specificity of REases can be
extended by fusion to sequence recognition modules, e.g. specific DNA-binding domains or
triple-helix forming oligonucleotides (TFO). We have chosen to extend the specificity of
REases using TFOs, given the combinatorial flexibility this fusion offers in addressing a
short, yet precisely recognized restriction site next to a defined triple-helix forming site
(TFS). We demonstrate here that the single chain variant of PvuII (scPvuII) covalently coupled
via the bifunctional cross-linker N-(gamma-maleimidobutryloxy) succinimide ester to a TFO
(5¡'-NH2-[CH2]6 or 12-MPMPMPMPMPPPPPPT-3¡', with M being 5-methyl-2¡'-deoxycytidine and P
being 5-[1-propynyl]-2¡'-deoxyuridine), cleaves DNA specifically at the recognition site of
PvuII (CAGCTG) if located in a distance of approximately one helical turn to a TFS
(underlined) complementary to the TFO (¡'addressed¡' site:
5¡'-TTTTTTTCTCTCTCTCN~10CAGCTG-3¡'), leaving ¡'unaddressed¡' PvuII sites intact. The
preference for cleavage of an ¡'addressed¡' compared to an ¡'unaddressed¡' site is
>1000-fold, if the cleavage reaction is initiated by addition of Mg2+ ions after preincubation
of scPvuII-TFO and substrate in the absence of Mg2+ ions to allow triple-helix formation
before DNA cleavage. Single base pair substitutions in the TFS prevent addressed DNA cleavage
by scPvuII-TFO.

<>

<1>Ekino, K., Kwon, I., Goto, M., Yoshino, S., Furukawa, K.
<2>Functional analysis of HO gene in delayed homothallism in Saccharomyces cerevisiae wy2.
<3>Yeast
<4>15
<5>451-458
<6>1999
<7>Saccharomyces cerevisiae wy2 exhibits a novel life cycle, with delayed homothallism caused by
a defective HO gene. In this strain, gradual diploidization occurs during successive
subcultures. Three amino acids of wy2 HO were different from those of wild-type (wt) HO, which
included a nonsense mutation (TAG) from Trp-292 and two amino acid changes of His-475 to Leu
and Glu-530 to Lys. The ho gene of heterothallic strain CG379 was also sequenced in this
study. Four amino acids of ho were different from those of HO. Among different amino acids in
wy2 HO and ho, the alteration of His-475 to Leu was common between them. His-475 in HO was
previously suggested to be involved in the DNA binding. We constructed a variety of chimeric
HO genes by exchanging the corresponding restriction fragments generated from the wt HO, wy2
HO and ho genes. These results and the site-directed mutagenesis studies allowed us to draw
the following conclusions: (a) Gly-223 is essential for HO activity; (b) mutation of His-475
to Leu significantly reduces the HO activity; (c) amber mutation (TAG) in wy2 HO car be
suppressed inefficiently.

<>

<1>Eklund, J.L.
<2>Design and characterization of homing endonuclease I-PpoI variants with novel DNA sequence specificity.
<3>Ph.D. Thesis, University of Washington, Seattle, USA
<4>
<5>
<6>2005
<7>Homing endonucleases bind and cleave long DNA target sites with high degree of sequence
specificity.  The homing endonuclease I-PpoI recognizes a 15 bp, semi-palindromic homing site
sequence in the rDNA of all eukaryotes.  The co-crystal structure indicates taht a b-sheet in
the major groove is responsible for most of the DNA-protein contacts that determine sequence
specificity of the enzyme.  Base pair changes in the +/-6 position of the I-PpoI binding site
disrupt cleavage.  To better understand the specificity at the +/- site, I have explored
variants with altered specificity for a +6C/-6G change.  Three libraries of I-PpoI variants
with varying degrees of rationally designed changes in the b-sheet were generated and screened
for altered DNA sequence specificity to a +6C/-6G basepair change using a yeast one-hybrid
assay.  A total of thirteen unique variants were isolated and characterized for binding
affinity and cleavage activity on the WT site and the +6C/-6G site; the other seven variants
had generally relaxed specificity or a higher affinity for the WT site.  Only one variant
showed cleavage activity on the WT site and none of the variants cleaves the +6C/-6G site.
Select variants were further investigated for their binding affinity for 4 other binding site
sequences.  Five variants with single amino acid changes to the WT protein sequence and three
variants with single amino acid changes to a protein variant with a specificity shift were
generated and biochemically characterized for cleavage activity and binding affinity.  I-PpoI
variant proteins provide insight into how DNA-protein contacts determine DNA sequence
specificity.  A deeper understanding of homing endonuclease sequence specificity determinants
and how specificity shifts occur will aid in the design and generation of homing endonuclease
variants as useful tools for genomic research and therapy.

<>

<1>Eklund, J.L., Ulge, U.Y., Eastberg, J., Monnat, R.J. Jr.
<2>Altered target site specificity variants of the I-PpoI His-Cys box homing endonuclease.
<3>Nucleic Acids Res.
<4>35
<5>5839-5850
<6>2007
<7>We used a yeast one-hybrid assay to isolate and characterize variants of the eukaryotic homing
endonuclease I-PpoI that were able to bind a mutant,
cleavage-resistant I-PpoI target or 'homing' site DNA in vivo. Native
I-PpoI recognizes and cleaves a semi-palindromic 15-bp target site with
high specificity in vivo and in vitro. This target site is present in the
28S or equivalent large subunit rDNA genes of all eukaryotes. I-PpoI
variants able to bind mutant target site DNA had from 1 to 8 amino acid
substitutions in the DNA-protein interface. Biochemical characterization
of these proteins revealed a wide range of site-binding affinities and
site discrimination. One-third of variants were able to cleave target site
DNA, but there was no systematic relationship between site-binding
affinity and site cleavage. Computational modeling of several variants
provided mechanistic insight into how amino acid substitutions that
contact, or are adjacent to, specific target site DNA base pairs determine
I-PpoI site-binding affinity and site discrimination, and may affect
cleavage efficiency.

<>

<1>El Aamri, F., Acosta, F., Real, F., Padilla, D.
<2>Whole-Genome Sequence of the Fish Virulent Strain Streptococcus iniae IUSA-1, Isolated from Gilthead Sea Bream (Sparus aurata) and Red Porgy (Pagrus pagrus).
<3>Genome Announcements
<4>1
<5>e0002513
<6>2013
<7>Streptococcus iniae is a major fish pathogen that produces invasive infections that result in
economic losses in aquaculture. In this study, the draft genome
sequence of Streptococcus iniae strain IUSA-1, isolated from a natural outbreak
affecting gilthead sea bream (Sparus aurata) and red porgy (Pagrus pagrus), is
presented.

<>

<1>El Kafsi, H., Binesse, J., Loux, V., Buratti, J., Boudebbouze, S., Dervyn, R., Hammani, A., Maguin, E., van de Guchte, M.
<2>Genome Sequence of Lactobacillus delbrueckii subsp. lactis CNRZ327, a Dairy Bacterium with Anti-Inflammatory Properties.
<3>Genome Announcements
<4>2
<5>e00328-14
<6>2014
<7>Lactobacillus delbrueckii subsp. lactis CNRZ327 is a dairy bacterium with anti-inflammatory
properties both in vitro and in vivo. Here, we report the
genome sequence of this bacterium, which appears to contain no less than 215
insertion sequence (IS) elements, an exceptionally high number regarding the
small genome size of the strain.

<>

<1>El-Arabi, T.F., Griffiths, M.W., She, Y.M., Villegas, A., Lingohr, E.J., Kropinski, A.M.
<2>Genome sequence and analysis of a broad-host range lytic bacteriophage that infects the Bacillus cereus group.
<3>Virol. J.
<4>10
<5>48
<6>2013
<7>ABSTRACT: BACKGROUND: Comparatively little information is available on members of
the Myoviridae infecting low G+C content, Gram-positive host bacteria of the
family Firmicutes. While numerous Bacillus phages have been isolated up till now
only very few Bacillus cereus phages have been characterized in detail. RESULTS:
Here we present data on the large, virulent, broad-host-range B. cereus phage
vB_BceM_Bc431v3 (Bc431v3). Bc431v3 features a 158,618 bp dsDNA genome,
encompassing 239 putative open reading frames (ORFs) and, 20 tRNA genes encoding
17 different amino acids. Since pulsed-field gel electrophoresis indicated that
the genome of this phage has a mass of 155-158 kb Bc431v3 DNA appears not to
contain long terminal repeats that are found in the genome of Bacillus phage
SPO1. CONCLUSIONS: Bc431v3 displays significant sequence similarity, at the
protein level, to B. cereus phage BCP78, Listeria phage A511 and Enterococcus
phage [latin capital letter o with stroke]EF24C and other morphologically related
phages infecting Firmicutes such as Staphylococcus phage K and Lactobacillus
phage LP65. Based on these data we suggest that Bc431v3 should be included as a
member of the Spounavirinae; however, because of all the diverse taxonomical
information has been addressed recently, it is difficult to determine the genus.
The Bc431v3 phage contains some highly unusual genes such as gp143 encoding
putative tRNAHis guanylyltransferase. In addition, it carries some genes that
appear to be related to the host sporulation regulators. These are: gp098, which
encodes a putative segregation protein related to FstK/SpoIIIE DNA transporters;
gp105, a putative segregation protein; gp108, RNA polymerase sigma factor F/B;
and, gp109 encoding RNA polymerase sigma factor G.

<>

<1>El-Deiry, W.S., Nelkin, B.D., Celano, P., Yen, R.-W.C., Falco, J.P., Hamilton, S.R., Baylin, S.B.
<2>High expression of the DNA methyltransferase gene characterizes human neoplastic cells and progression stages of colon cancer.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>88
<5>3470-3474
<6>1991
<7>DNA methylation abnormalities occur consistently in human neoplasia including widespread
hypomethylation and more recently recognized local increases in DNA methylation that hold
potential for gene inactivation events. To study this imbalance further, we have cloned and
localized to chromosome 19 a portion of the human DNA methyltransferase gene that codes for
the enzyme catalyzing DNA methylation. Expression of this gene is low in normal human cells,
significantly increased (30- to 50-fold by PCR analysis) in virally transformed cells, and
strikingly elevated in human cancer cells (several hundredfold). In comparison to colon mucosa
from patients without neoplasia, median levels of DNA methyltransferse transcripts are 15-fold
increased in histologically normal nucosa from patients with cancers of the benign polyps that
can precede cancers, 60-fold increased in the premalignant polyps, and >200-fold increased in
the cancers. Thus, increases in DNA methyltansferase gene expression precede development of
colonic neoplasia and continue during progression of colonic neoplasms. These increases may
play a role in the genetic instability of cancer and mark early events in cell transformation.

<>

<1>El-Sayed, E.S.A., El-Didamony, G., Mansour, K.
<2>Isolation and characterization of two types of actinophage infecting Streptomyces scabies.
<3>Folia Microbiol. (Praha)
<4>46
<5>519-526
<6>2001
<7>Two types of actinophages, phiS and phiL, were isolated from soil samples by using
Streptomyces scabies, a potato scab pathogen, as
indicator strain. The phages were partially characterized according to
their physicochemical properties, plaques and particles morphology, and
their host range; this varied from narrow (for phiS) to wide (for
phiL). The adsorption rate constants of the phiS and phiL were 3.44 and
3.18 pL/min, and their burst sizes were 1.61 and 3.75 virions per mL,
respectively. One-step growth indicated that phiS and phiL have a
latent period of 1/2 h followed by a rise period of 1/2 h. The
temperate character of these phages was tested in other isolates of
Streptomyces. Four of the phages ( phiSS3, phiSS12, phiSS13 and
phiSS17) were identified as temperate phages, since they were able to
lysogenize SS3, SS12, SS13 and SS17. phiSS3, phiSS12 and, phiSS13 were
homoimmune, and they were heteroimmune with respect to SS17. The
restriction barriers of lysogenic isolates (SS12, SS13 and SS17)
interfered with the blockage of plaque formation by phages ( phiSS12,
phiSS13 or phiSS17) propagated on them, about 75 % of lysogenic
isolates had restriction systems. The exposure of the lysogenic
isolates (SS12, SS13 and SS17) to UV-irradiation prevented the possible
restriction barriers of these isolates so that these barriers could be
overcome.

<>

<1>Elbir, H., Gimenez, G., Robert, C., Bergstrom, S., Cutler, S., Raoult, D., Drancourt, M.
<2>Complete Genome Sequence of Borrelia crocidurae.
<3>J. Bacteriol.
<4>194
<5>3723-3724
<6>2012
<7>We announce the draft genome sequence of Borrelia crocidurae (strain Achema). The 1,557,560-bp
genome (27% GC content) comprises one 919,477-bp linear chromosome
and 638,083-bp plasmids that together carry 1,472 open reading frames, 32 tRNAs,
and three complete rRNAs, with almost complete colinearity between B. crocidurae
and Borrelia duttonii chromosomes.

<>

<1>Elbir, H., Larsson, P., Normark, J., Upreti, M., Korenberg, E., Larsson, C., Bergstrom, S.
<2>Genome Sequence of the Asiatic Species Borrelia persica.
<3>Genome Announcements
<4>2
<5>e01127-13
<6>2014
<7>We report the complete genome sequence of Borrelia persica, the causative agent of tick-borne
relapsing fever borreliosis on the Asian continent. Its genome of
1,784,979 bp contains 1,850 open reading frames, three ribosomal RNAs, and 32
tRNAs. One clustered regularly interspaced short palindromic repeat (CRISPR) was
detected.

<>

<1>Elbir, H., Larsson, P., Upreti, M., Normark, J., Bergstrom, S.
<2>Genome Sequence of the Relapsing Fever Borreliosis Species Borrelia hispanica.
<3>Genome Announcements
<4>2
<5>e01171-13
<6>2014
<7>Borrelia hispanica is the etiological pathogen of tick-borne relapsing fever, transmitted to
humans by infected Ornithodoros erraticus ticks. Here we present
the 1,783,846-bp draft genome sequence, with an average G+C content of 28%. It
has 2,140 open reading frames, 3 ribosomal RNAs, and 32 transfer RNAs.

<>

<1>Elbir, H., Robert, C., Nguyen, T.T., Gimenez, G., El Sanousi, S.M., Flock, J.I., Raoult, D., Drancourt, M.
<2>Staphylococcus aureus subsp. anaerobius strain ST1464 genome sequence.
<3>Standards in Genomic Sciences
<4>9
<5>1-13
<6>2013
<7>Staphylococcus aureus subsp. anaerobius is responsible for Morel's disease in animals and a
cause of abscess in humans. It is characterized by a
microaerophilic growth, contrary to the other strains of S. aureus. The
2,604,446-bp genome (32.7% GC content) of S. anaerobius ST1464 comprises one
chromosome and no plasmids. The chromosome contains 2,660 open reading frames
(ORFs), 49 tRNAs and three complete rRNAs, forming one complete operon. The size
of ORFs ranges between 100 to 4,600 bp except for two ORFs of 6,417 and 7,173 bp
encoding segregation ATPase and non-ribosomal peptide synthase, respectively. The
chromosome harbors Staphylococcus phage 2638A genome and incomplete
Staphylococcus phage genome PT1028, but no detectable CRISPRS. The antibiotic
resistance gene for tetracycline was found although Staphylococcus aureus subsp.
anaerobius is susceptible to tetracycline in-vitro. Intact oxygen detoxification
genes encode superoxide dismutase and cytochrome quinol oxidase whereas the
catalase gene is impaired by a stop codon. Based on the genome, in-silico
multilocus sequence typing indicates that S. aureus subsp. anaerobius emerged as
a clone separated from all other S. aureus strains, illustrating host-adaptation
linked to missing functions. Availability of S. aureus subsp. anaerobius genome
could prompt the development of post-genomic tools for its rapid discrimination
from S. aureus.

<>

<1>Eldarov, M.A., Karpichev, I.V., Samko, O.T., Anikeitcheva, N.V., Kalugin, A.A., Khoroshoutina, E.B., Sokolov, N.N.
<2>Characterization of BciBII, an isoschizomer of BstNI from a strain of Bacillus circulans B.
<3>Nucleic Acids Res.
<4>20
<5>2896
<6>1992
<7>BciBII, an isoschizomer of BstNI (figure 1) has been isolated from Bacillus circulans B. The
enzyme was purified by precipitation with polyethylenimine, ammonium sulfate and subsequent
column chromatography on DEAE-sepharose, Blue sepharose and phosphocellulose. Restriction
endonuclease BciBII showed maximal activity at 37C, pH between 7.4-7.8, MgCl2 concentration in
the range of 8-12 mM and at high ionic strenghth.

<>

<1>Eldarov, M.A., Karpichev, I.V., Samko, O.T., Anikeitcheva, N.V., Kalugin, A.A., Khoroshoutina, E.B., Sokolov, N.N.
<2>Isolation and characterization of restriction endonuclease BcuAI from Bacillus cereus A.
<3>Nucleic Acids Res.
<4>20
<5>2898
<6>1992
<7>Restriction endonuclease BcuAI, an isoschizomer of AvaII (Figure 1), has been purified from
Bacillus cereus A. The procedure of isolation of BcuAI included the fractionation with
ammonium sulfate and column chromatography on DEAE-Sepharose (elution buffer 10 mM
K-phosphate, pH 7.4, 0.1 mM EDTA, 1 mM DTT, 0.0-1.0 M KCl; elution zone of enzyme activity
0.2-0.3 M KCl). 1400 u. BcuAI can be obtained from 1 g. of wet cells. BcuAI showed maximal
activity at 30-37C, pH between 7.6-8.2, MgCl2 concentration in the range of 5-10 mM and at
high ionic strength.

<>

<1>Elde, M., Haugen, P., Willassen, N.P., Johansen, S.
<2>I-NjaI, a nuclear intron-encoded homing endonuclease from Naegleria, generates a pentanucleotide 3' cleavage-overhang within a 19 base-pair partially symmetric DNA recognition site.
<3>Eur. J. Biochem.
<4>259
<5>281-288
<6>1999
<7>Different species of the amoebo-flagellate Naegleria harbor optional group I introns in the
nuclear ribosomal DNA that contain open reading frames.  Intron proteins from Naegleria
jamiesoni, Naegleria andersoni, and Naegleria italica (named I-NjaI, I-NanI and I-NitI,
respectively) were expressed in Escherichia coli and found to be isoschizomeric homing
endonucleases that specifically recognize and cleave intron-lacking homologous alleles of
ribosomal DNA.  The I-NjaI endonuclease was affinity purified, characterized in more detail,
and found to generate five-nucleotide 3' staggered ends at the intron insertion site which
differs from the ends generated by all other known homing endonucleases.  The recognition site
was delimited and found to cover an ~19 base-pair partially symmetric sequence spanning both
the cleavage site and the intron insertion site.  The palindromic feature was supported by
mutational analysis of the target DNA.  All single-site substitutions within the recognition
sequence were cleaved by the purified I-NjaI endonuclease, but at different efficiencies.  The
center for symmetry and cleavage was found to be completely degenerate in specificity, which
resembles that of the subclass IIW bacterial restriction enzymes.

<>

<1>Elde, M., Willassen, N.P., Johansen, S.
<2>Functional characterization of isoschizomeric His-Cys box homing endonucleases from Naegleria.
<3>Eur. J. Biochem.
<4>267
<5>7257-7266
<6>2000
<7>Several species within the amoeboflagellate genus Naegleria harbor an optional ORF containing
group I introns in their nuclear small subunit ribosomal DNA. The different ORFs encode homing
endonucleases with 65 to 95% identity at the amino-acid level. I-NjaI, I-NanI and I-NitI, from
introns in Naegleria jamiesoni, N. andersoni and N. italica, respectively, were analyzed in
more detail and found to be isoschizomeric endonucleases that recognize and cleave an
approximately 19-bp partially symmetrical sequence, creating a pentanucleotide 3' overhang
upon
cleavage. The optimal conditions for cleavage activity with respect to temperature, pH, salt
and divalent metal ions were investigated. The optimal cleavage temperature for all three
endonucleases was found to be 37 degrees C and the activity was dependent on the concentration
of NaCl with an optimum at 200 mM. Divalent metal ions, primarily Mg2+, are essential for
Naegleria endonuclease activity. Whereas both Mn2+ and Ca2+ could substitute for Mg2+, but
with a slower cleavage rate, Zn2+ was unable to support cleavage. Interestingly, the pH
dependence of DNA cleavage was found to vary significantly between the I-NitI and
I-NjaI/I-NanI endonucleases with optimal pH values at 6.5 and 9, respectively. Site-directed
mutagenesis of conserved I-NjaI residues strongly supports the hypothesis that Naegleria
homing endonucleases share a similar zinc-binding structure and active site with the His-Cys
box homing endonuclease I-PpoI.

<>

<1>Elhai, J.
<2>Determination of bias in the relative abundance of oligonucleotides in DNA sequences.
<3>J. Comput. Biol.
<4>8
<5>151-175
<6>2001
<7>Different statistical measures of bias of oligonucleotide sequences in DNA sequences were
compared, both by theoretical analysis and according to their abilities to predict the
relative abundances of oligonucleotides in the genome of Escherichia coli. The expected
frequency of an oligonucleotide calculated from a maximal order Markov model was shown to be a
degenerate case of the expected frequency calculated from biases of all subwords arising when
noncontiguous subwords exhibit no bias. Since (at least in E, coli) noncontiguous sequences
exhibit significant bias, the total compositional bias
approach is expected to represent biases in genomic sequences more
faithfully than Markov approaches. In fact, the efficacy of statistics
based on Markov analysis even at the highest order were inferior in
predicting actual frequencies of oligonucleotides to methods that
factored out biases of internal subwords with gaps. Using total
compositional bias as a measure of relative abundance, tetranucleotide
and hexanucleotide palindromes were found to be distributed differently
from nonpalindromic sequences, with their means shifted somewhat
towards underrepresentation, A subpopulation of palindromic
hexanucleotides, however, was highly underrepresented, and this group
consisted almost entirely of targets for Type II restriction enzymes
found within strains of E, coli, Sites recognized by Type I
endonucleases from related strains were not markedly biased, and with
pentanucleotides, palindromic and nonpalindromic sequences had nearly
identical distributions. The loss of restriction sites may be explained
by the free transfer of plasmids encoding restriction enzymes and
episodic selection for the presence of the enzymes.

<>

<1>Elhai, J., Cai, Y., Wolk, C.P.
<2>Conduction of pEC22, a plasmid coding for MR.EcoT22I, mediated by a resident Tn3-like transposon, Tn5396.
<3>J. Bacteriol.
<4>176
<5>5059-5067
<6>1994
<7>pEC22 is a small plasmid that encodes the restriction-modification system MR.EcoT22I.
Restriction and functional analysis of the plasmid identified the positions of genes encoding
that system. The plasmid is able to be conducted by conjugal plasmids, a process mediated by a
transposon contained within pEC22. This cryptic transposon, called Tn5396, was isolated from
pEC22 and partially sequenced. The sequence of Tn5396 is for the most part typical of
transposons of the Tn3 family and is most similar to that of Tn1000. The transposon differs
from closely related transposons in that it lacks well-conserved sequences in the
inverted-repeat region and has an unusually long terminal inverted repeat. Consideration of
regions of internal sequence similarity in this and other transposons in the Tn3 family
supports a theory of the mechanism by which the ends of Tn3-like transposons may maintain
substantial identity between their inverted repeats over the course of evolutionary time.

<>

<1>Elhai, J., Vepritskiy, A., Muro-Pastor, A.M., Flores, E., Wolk, C.P.
<2>Reduction of conjugal transfer efficiency by three restriction activities of Anabaena sp. strain PCC 7120.
<3>J. Bacteriol.
<4>179
<5>1998-2005
<6>1997
<7>The efficiency of conjugal transfer of plasmids from Escherichia coli to the cyanobacterium
Anabaena sp. strain PCC 7120 was quantitated as a function of the number of restriction sites
for the restriction enzymes carried by the recipient.  In addition to the previously
recognized isoschizomers of AvaI and AvaII, PCC 7120 was found to possess an isoschizomer of
AvaIII.  Plasmids modified in E. coli with methylases that protect in vitro against
restriction by the three enzymes were transferred with high efficiency, nearly independent of
the number of restriction sites on the plasmid.  Plasmids left unprotected against one of the
three restriction enzymes were transferred with lower efficiencies.  For low numbers of sites,
the efficiency of conjugal transfer decreased as an exponential function of the number of
unprotected sites.  The methods presented may be used to increase the efficiency of conjugal
transfer into restriction-competent bacteria.

<>

<1>Elkins, J.G. et al.
<2>A korarchaeal genome reveals insights into the evolution of the Archaea.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>8102-8107
<6>2008
<7>The candidate division Korarchaeota comprises a group of uncultivated microorganisms that, by
their small subunit rRNA phylogeny, may have
diverged early from the major archaeal phyla Crenarchaeota and
Euryarchaeota. Here, we report the initial characterization of a member of
the Korarchaeota with the proposed name, "Candidatus Korarchaeum
cryptofilum," which exhibits an ultrathin filamentous morphology. To
investigate possible ancestral relationships between deep-branching
Korarchaeota and other phyla, we used whole-genome shotgun sequencing to
construct a complete composite korarchaeal genome from enriched cells. The
genome was assembled into a single contig 1.59 Mb in length with a G + C
content of 49%. Of the 1,617 predicted protein-coding genes, 1,382 (85%)
could be assigned to a revised set of archaeal Clusters of Orthologous
Groups (COGs). The predicted gene functions suggest that the organism
relies on a simple mode of peptide fermentation for carbon and energy and
lacks the ability to synthesize de novo purines, CoA, and several other
cofactors. Phylogenetic analyses based on conserved single genes and
concatenated protein sequences positioned the korarchaeote as a deep
archaeal lineage with an apparent affinity to the Crenarchaeota. However,
the predicted gene content revealed that several conserved cellular
systems, such as cell division, DNA replication, and tRNA maturation,
resemble the counterparts in the Euryarchaeota. In light of the known
composition of archaeal genomes, the Korarchaeota might have retained a
set of cellular features that represents the ancestral archaeal form.

<>

<1>Elkins, J.G. et al.
<2>Complete Genome Sequence of the Cellulolytic Thermophile Caldicellulosiruptor obsidiansis OB47T.
<3>J. Bacteriol.
<4>192
<5>6099-6100
<6>2010
<7>Caldicellulosiruptor obsidiansis OB47(T) (ATCC BAA-2073, JCM 16842) is an extremely
thermophilic, anaerobic bacterium capable of hydrolyzing
plant-derived polymers through the expression of
multidomain/multifunctional hydrolases. The complete genome sequence
reveals a diverse set of carbohydrate-active enzymes and provides further
insight into lignocellulosic biomass hydrolysis at high temperatures.

<>

<1>Elkins, J.G. et al.
<2>Complete Genome Sequence of the Hyperthermophilic Sulfate-Reducing Bacterium Thermodesulfobacterium geofontis OPF15T.
<3>Genome Announcements
<4>1
<5>e00162-13
<6>2013
<7>Thermodesulfobacterium geofontis OPF15(T) (ATCC BAA-2454, JCM 18567) was isolated from
Obsidian Pool, Yellowstone National Park, and grows optimally at 83 degrees
C. The 1.6-Mb genome sequence was finished at the Joint Genome Institute and has
been deposited for future genomic studies pertaining to microbial processes and
nutrient cycles in high-temperature environments.

<>

<1>Ellegaard, K.M., Klasson, L., Naslund, K., Bourtzis, K., Andersson, S.G.
<2>Comparative genomics of wolbachia and the bacterial species concept.
<3>PLoS Genet.
<4>9
<5>E1003381
<6>2013
<7>The importance of host-specialization to speciation processes in obligate
host-associated bacteria is well known, as is also the ability of recombination
to generate cohesion in bacterial populations. However, whether divergent strains
of highly recombining intracellular bacteria, such as Wolbachia, can maintain
their genetic distinctness when infecting the same host is not known. We first
developed a protocol for the genome sequencing of uncultivable endosymbionts.
Using this method, we have sequenced the complete genomes of the Wolbachia
strains wHa and wNo, which occur as natural double infections in Drosophila
simulans populations on the Seychelles and in New Caledonia. Taxonomically, wHa
belong to supergroup A and wNo to supergroup B. A comparative genomics study
including additional strains supported the supergroup classification scheme and
revealed 24 and 33 group-specific genes, putatively involved in host-adaptation
processes. Recombination frequencies were high for strains of the same supergroup
despite different host-preference patterns, leading to genomic cohesion. The
inferred recombination fragments for strains of different supergroups were of
short sizes, and the genomes of the co-infecting Wolbachia strains wHa and wNo
were not more similar to each other and did not share more genes than other A-
and B-group strains that infect different hosts. We conclude that Wolbachia
strains of supergroup A and B represent genetically distinct clades, and that
strains of different supergroups can co-exist in the same arthropod host without
converging into the same species. This suggests that the supergroups are
irreversibly separated and that barriers other than host-specialization are able
to maintain distinct clades in recombining endosymbiont populations. Acquiring a
good knowledge of the barriers to genetic exchange in Wolbachia will advance our
understanding of how endosymbiont communities are constructed from vertically and
horizontally transmitted genes.

<>

<1>Elliott, S.L., Brazier, J., Cosstick, R., Connolly, B.A.
<2>Mechanism of the Escherichia coli DNA T:G-mismatch endonuclease (Vsr protein) probed with thiophosphate-containing oligodeoxynucleotides.
<3>J. Mol. Biol.
<4>353
<5>692-703
<6>2005
<7>The mechanism of the Escherichia coli DNA T:G mismatch endonuclease (Vsr) has been
investigated using oligodeoxynucleotides substituted, at the scissile phosphate, with isomeric
phosphorothioates and a 3'-phosphorothiolate. Binding and kinetic data with the
phosphorothioates/phosphorothiolate indicate that the two magnesium ions, which constitute
essential co-factors, are required to stabilise the extra negative charge developed on the
phosphate as the transition state is formed. Additionally one of the magnesium ions serves to
activate the leaving group (the non-bridging 3'-oxygen atom of the scissile phosphate) during
the hydrolysis reaction. Stereochemical analysis, using the R(p) phosphorothioate isomer,
indicates that Vsr carries out a hydrolytic reaction with inversion of stereochemistry at
phosphorus, compatible with an in-line attack of water and a pentacovalent transition state
with trigonal bipyramidal geometry. In conjunction with structures of Vsr bound to its
products, these data allow the reconstruction of the enzyme-substrate complex and a
comprehensive description of the hydrolysis mechanism.

<>

<1>Elliott, T.
<2>Cloning, genetic characterization, and nucleotide sequence of the hemA-prfA operon of Salmonella typhimurium.
<3>J. Bacteriol.
<4>171
<5>3948-3960
<6>1989
<7>The first step in heme biosynthesis is the formation of 5-aminolevulinic acid (ALA). Mutations
in two genes, hemA and hemL, result in auxotrophy for ALA in Salmonella typhimurium, but the
roles played by these genes and the mechanism of ALA synthesis are not understood. I have
cloned and sequenced the S. typhimurium hemA gene. The predicted polypeptide sequence for the
HemA protein shows no similarity to known ALA synthases, and no ALA synthase activity was
detected in extracts prepared from strains carrying the cloned hemA gene. Genetic analysis,
DNA sequencing of amber mutations, and maxicell studies proved that the open reading frame
identified in the DNA sequence encodes HemA. Another surprising finding of this study is that
hemA lies directly upstream of prfA, which encodes peptide chain release factor 1 (RF-1). A
hemA::Kan insertion mutation, constructed in vitro, was transferred to the chromosome and used
to show that these two genes form an operon. The hemA gene ends with an amber codon,
recognized by RF-1. I suggest a model for autogenous control of prfA expression by translation
reinitiation.

<>

<1>Ellis, D.J., Dryden, D.T., Berge, T., Edwardson, J.M., Henderson, R.M.
<2>Direct observation of DNA translocation and cleavage by the EcoKI endonuclease using atomic force microscopy.
<3>Nat. Struct. Biol.
<4>6
<5>15-17
<6>1999
<7>Type I DNA restriction and modification enzymes such as EcoKI are multimeric enzymes that
cleave DNA molecules lacking an appropriate pattern of methylation on a target sequence,
thereby protecting the host bacterium from invasion by foreign DNA.  The reaction involves the
initial recognition of an unmethylated DNA target sequence with the aid of the cofactor
S-adenosyl methionine.  Recognition is followed by extensive ATP-dependent translocation of
the DNA in both directions toward the enzyme, which remains bound at the target sequence.
Eventually, endonucleolytic cleavage of the DNA occurs.  Recently, protein complexes with
nucleic acids have been imaged successfully using fluid tapping-mode atomic force microscopy.
In this study, we have used AFM to explore the type I DNA restriction and modification
pathway, and specifically to follow the ATP-dependent translocation and cleavage of a single
plasmid by the EcoKI enzyme complex under near-physiological conditions.

<>

<1>Ellison, D.W., Clark, T.R., Sturdevant, D.E., Virtaneva, K., Porcella, S.F., Hackstadt, T.
<2>Genomic comparison of virulent Rickettsia rickettsii Sheila Smith and avirulent Rickettsia rickettsii Iowa.
<3>Infect. Immun.
<4>76
<5>542-550
<6>2008
<7>Rickettsia rickettsii is an obligate intracellular pathogen that is the causative agent of
Rocky Mountain spotted fever. To identify genes
involved in the virulence of R. rickettsii, the genome of an avirulent
strain, R. rickettsii Iowa, was sequenced and compared to the genome of
the virulent strain R. rickettsii Sheila Smith. R. rickettsii Iowa is
avirulent in a guinea pig model of infection and displays altered plaque
morphology with decreased lysis of infected host cells. Comparison of the
two genomes revealed that R. rickettsii Iowa and R. rickettsii Sheila
Smith share a high degree of sequence identity. A whole-genome alignment
comparing R. rickettsii Iowa to R. rickettsii Sheila Smith revealed a
total of 143 deletions for the two strains. A subsequent single-nucleotide
polymorphism (SNP) analysis comparing Iowa to Sheila Smith revealed 492
SNPs for the two genomes. One of the deletions in R. rickettsii Iowa
truncates rompA, encoding a major surface antigen (rickettsial outer
membrane protein A [rOmpA]) and member of the autotransporter family, 660
bp from the start of translation. Immunoblotting and immunofluorescence
confirmed the absence of rOmpA from R. rickettsii Iowa. In addition, R.
rickettsii Iowa is defective in the processing of rOmpB, an
autotransporter and also a major surface antigen of spotted fever group
rickettsiae. Disruption of rompA and the defect in rOmpB processing are
most likely factors that contribute to the avirulence of R. rickettsii
Iowa. Genomic differences between the two strains do not significantly
alter gene expression as analysis of microarrays revealed only four
differences in gene expression between R. rickettsii Iowa and R.
rickettsii strain R. Although R. rickettsii Iowa does not cause apparent
disease, infection of guinea pigs with this strain confers protection
against subsequent challenge with the virulent strain R. rickettsii Sheila
Smith.

<>

<1>Ellison, E.L., Muscarella, D.E., Vogt, V.M.
<2>Characterization of I-PpoI, an intron-encoded endonuclease from Physarum polycephalum.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>177
<6>1993
<7>The genetic mobility of group I introns is dependent upon the site-specific endonucleases
which they encode. We have characterized several properties of I-PpoI, the endonuclease that
mediates "homing" of intron 3 (PpLSU3) in the nuclear ribosomal DNA (rDNA) of the slime mold
Physarum polycephalum. From deletion analysis, we conclude that the minimum recognition site
is a sequence of 15 bp, which is partially symmetric. The purified enzyme behaves as a dimer
in gel filtration and sedimentation. We have studied the interaction of I-PpoI with DNA by
bandshift analysis, MPE footprinting and DMS protection. The enzyme, which binds in the major
groove, protects ca. 21bp of DNA surrounding the cleavage site. The I-PpoI recognition site is
present in the nuclear rDNA of all eucaryotes. In yeast, expression of the enzyme is lethal,
presumably because of cleavage of rDNA repeats on chromosome XII. By pulse field gel analysis,
this is the only chromosome cut in vitro. Yeast mutants that survive the lethal effects of the
endonuclease occur at surprisingly high frequencies, and are of at least two classes. The
first consists of cells carrying point mutations in the recognition sequence. All ca. 150
copies of the rDNA are mutant, as evidenced by the inability of I-PpoI to cleave the rDNA in
vitro. The second class consists of cells in which the intron has become inserted into all
copies of the rDNA. These cells express active endonuclease.

<>

<1>Ellison, E.L., Vogt, V.M.
<2>Interaction of the intron-encoded mobility endonuclease I-PpoI with its target site.
<3>Mol. Cell. Biol.
<4>13
<5>7531-7539
<6>1993
<7>Endonucleases encoded by mobile group I introns are highly specific DNases that induce a
double-strand break near the site to which the intron moves. I-PpoI from the acellular slime
mold Physarum polycephalum mediates the mobility of intron 3 (Pp LSU3) in the extrachromosomal
nuclear ribosomal DNA of this organism. We showed previously that cleavage by I-PpoI creates a
four-base staggered cut near the point of intron insertion. We have now characterized several
further properties of the endonuclease. As determined by deletion analysis, the minimal target
site recognized by I-PpoI was a sequence of 13 to 15 bp spanning the cleavage site. The
purified protein behaved as a globular dimer in sedimentation and gel filtration. In gel
mobility shift assays in the presence of EDTA, I-PpoI formed a stable and specific complex
with DNA, dissociating with a half-life of 45 minutes. By footprinting and interference assays
with methidiumpropyl-EDTA-intron(II), I-PpoI contacted a 22-to 24-bp stretch of DNA. The
endonuclease protected most of the purines found in both the major and minor grooves of the
DNA helix from modification by dimethylsulfate (DMS). However, the reactivity to DMS was
enhanced at some purines, suggesting that binding leads to a conformational change in the DNA.
The pattern of DMS protection differed fundamentally in the two partially symmetrical halves
of the recognition sequence.

<>

<1>Ellrott, K.P., Kasarjian, J.K.A., Jiang, T., Ryu, J.
<2>Restriction enzyme recognition sequence search program.
<3>Biotechniques
<4>33
<5>1322-1326
<6>2002
<7>A critical and difficult part of characterizing restriction enzymes and methylases is the
identification of recognition sequences.  To simplify this process, we have developed a
plasmid transformation method along with a computer program named RM search that determines
the exact recognition sequences for given restriction and modification systems.

<>

<1>Embleton, M.L., Siksnys, V., Halford, S.E.
<2>DNA cleavage reactions by type II restriction enzymes that require two copies of their recognition sites.
<3>J. Mol. Biol.
<4>311
<5>503-514
<6>2001
<7>Several type II restriction endonucleases interact with two copies of their target sequence
before they cleave DNA. Three such enzymes,
NgoMIV, Cfr10I and NaeI, were tested on plasmids with one or two copies
of their recognition sites, and on catenanes containing two interlinked
rings of DNA with one site in each ring. The enzymes showed distinct
patterns of behaviour. NgoMIV and NaeI cleaved the plasmid with two
sites faster than that with one site and the catenanes at an
intermediate rate, while Cfr10I gave similar steady-state rates on all
three substrates. Both Cfr10I and NgoMIV converted the majority of the
substrates with two sites directly to the products cut at both sites,
while NaeI cleaved just one site at a time. All three enzymes thus
synapse two DNA sites through three-dimensional space before cleaving
DNA. With Cfr10I and NgoMIV, both sites are cleaved in one turnover, in
a manner consistent with their tetrameric structures, while the
cleavage of a single site by NaeI indicates that the second site acts
not as a substrate but as an activator, as reported previously. The
complexes spanning two sites have longer lifetimes on catenanes with
one site in each ring than on circular DNA with two sites, which
indicates that the catenanes have more freedom for site juxtaposition
than plasmids with sites in cis.

<>

<1>Embleton, M.L., Vologodskii, A.V., Halford, S.E.
<2>Dynamics of DNA loop capture by the Sfil restriction endonuclease on supercoiled and relaxed DNA.
<3>J. Mol. Biol.
<4>339
<5>53-66
<6>2004
<7>The SfiI endonuclease is a prototype for DNA looping. It binds two copies of its recognition
sequence and, if Mg2+ is present, cuts both
concertedly. Looping was examined here on supercoiled and relaxed forms
of a 5.5 kb plasmid with three SfiI sites: sites 1 and 2 were separated
by 0.4 kb, and sites 2 and 3 by 2.0 kb. SfiI converted this plasmid
directly to the products cut at all three sites, though DNA species
cleaved at one or two sites were formed transiently during a burst
phase. The burst revealed three sets of doubly cut products,
corresponding to the three possible pairings of sites. The equilibrium
distribution between the different loops was evaluated from the burst
phases of reactions initiated by adding MgCl2 to SfiI bound to the
plasmid. The short loop was favored over the longer loops, particularly
on supercoiled DNA. The relative rates for loop capture were assessed
after adding SfiI to solutions containing the plasmid and MgCl2. On
both supercoiled and relaxed DNA, the rate of loop capture across 0.4
kb was only marginally faster than over 2.0 kb or 2.4 kb. The relative
strengths and rates of looping were compared to computer simulations of
conformational fluctuations in DNA. The simulations concurred broadly
with the experimental data, though they predicted that increasing site
separations should cause a shallower decline in the equilibrium
constants than was observed but a slightly steeper decline in the rates
for loop capture. Possible reasons for these discrepancies are
discussed.

<>

<1>Embleton, M.L., Williams, S.A., Watson, M.A., Halford, S.E.
<2>Specificity from the synapsis of DNA elements by the SfiI endonuclease.
<3>J. Mol. Biol.
<4>289
<5>785-797
<6>1999
<7>The synapsis of DNA sites is a prerequisite for the reactions of many proteins that act at
specific DNA sequences. The requirement for synapsis was investigated by analysing the
reactions of SfiI, a tetrameric restriction enzyme that cleaves DNA only after interacting
with two recognition sites. In the presence of Mg2+, oligonucleotide duplexes with the cognate
recognition sequence were cleaved rapidly, with cooperative kinetics, while non-cognate
duplexes were not cleaved. In the absence of Mg2+, the primary complex formed by SfiI with
cognate DNA contained two duplexes synapsed by the tetramer: a secondary complex containing
one duplex was seen only at elevated SfiI concentrations. In contrast, the principal complex
with non-cognate DNA contained one duplex bound to SfiI. Pairs of non-cognate duplexes, or one
cognate and one non-cognate duplex, generally failed to form synaptic complexes. On adding
Mg2+to complexes with cognate DNA, cleavage occurred much more rapidly in the synaptic complex
than in the secondary complex. DNA synapsis thus acts to enhance the specificity of SfiI for
its recognition sequence, by demanding two cognate sites for a catalytically active complex
and by excluding non-cognate sites from the synaptic complex.

<>

<1>Embley, T.M., Dyal, P., Kilvington, S.
<2>A group I intron in the small subunit ribosomal RNA gene from Naegleria andersoni ssp. andersoni strain PPMFB-6.
<3>Nucleic Acids Res.
<4>20
<5>6411
<6>1992
<7>PCR reactions designed to amplify the entire small subunit (SSU) ribosomal RNA gene from
Naegleria species for phylogenetic analyses, demonstrated a single band of approximately 2 kb
in most species but a band of approximately 3.2 kb in strains of Naegleria andersoni,
Naegleria andersoni ssp. andersoni and Naegleria australiensis ssp. italica.  The extra length
in N.andersoni spp. andersoni strain PPMFB-6 is due to an insertion of approximately 1277 base
pairs in the coding sequence of the rRNA gene.  The insertion is sited near the tip of helix
19 immediately prior to the conserved SSU rRNA sequence CC-AG.  Extraction and sizing of total
rRNA by gel electrophoresis revealed that the insertion was removed in vivo to produce a
mature SSU rRNA of the same size as strains lacking the insertion in their SSU gene.  The
insertion in N.andersoni ssp. andersoni was amplified and directly sequenced using a linear
PCR reaction.  Sequence analysis revealed that it contains motifs which identify it as a group
I intron.  Thus, the short sequences P, Q, R and S, which represent the conserved core
structure of group I introns, are all present and they occur in that order.  The intron also
contains a long open reading frame situated between R and S, which codes for a putative
protein of unknown function and containing 245 amino acids.  Group I introns are rare in
nuclear SSU rRNA genes and have hitherto only been reported in Ustilago maydis, Pneumocytis
carini and Ankistrodesmus stipitatus.  The introns in these taxa are all small (394-480
bases), they do not contain long open reading frames, and they are inserted at different
positions in the SSU rRNA gene.  Furthermore, Naegleria branches at a point in the eukaryote
phylogenetic tree which is much deeper than these taxa.  Thus, our observation significantly
extends the phylogenetic range of group I introns in Eukaryote nuclear SSU genes.

<>

<1>Emond, E., Ee, E.L., Drolet, G., Moineau, S., LaPointe, G.
<2>Molecular characterization of a theta replication plasmid and its use for development of a two-component food-grade cloning system for Lactococcus lactis.
<3>Appl. Environ. Microbiol.
<4>67
<5>1700-1709
<6>2001
<7>pCD4, a small, highly stable theta-replicating lactococcal plasmid, was
used to develop a food-grade cloning system. Sequence analysis revealed
five open reading frames and two putative cis-acting regions. None appears
to code for undesirable phenotypes with regard to food applications.
Functional analysis of the replication module showed that only the
cis-acting ori region and the repB gene coding for the replication
initiator protein were needed for the stable replication and maintenance
of pCD4 derivatives in Lactococcus lactis. A two-component food-grade
cloning system was derived from the pCD4 replicon. The vector pVEC1, which
carries the functional pCD4 replicon, is entirely made up of L. lactis DNA
and has no selection marker. The companion pCOM1 is a repB-deficient pCD4
derivative that carries an erythromycin resistance gene as a dominant
selection marker. The pCOM1 construct can only replicate in L. lactis if
trans complemented by the RepB initiator provided by pVEC1. Since only the
cotransformants that carry both pVEC1 and pCOM1 can survive on plates
containing erythromycin, pCOM1 can be used transiently to select cells
that have acquired pVEC1. Due to the intrinsic incompatibility between
these plasmids, pCOM1 can be readily cured from the cells grown on an
antibiotic-free medium after the selection step. The system was used to
introduce a phage resistance mechanism into the laboratory strain MG1363
of L. lactis and two industrial strains. The introduction of the antiphage
barrier did not alter the wild-type plasmid profile of the industrial
strains. The phenotype was stable after 100 generations and conferred an
effective resistance phenotype against phages of the 936 and c2 species.

<>

<1>Emond, E., Holler, B.J., Boucher, I., Vandenbergh, P.A., Vedamuthu, E.R., Kondo, J.K., Moineau, S.
<2>Phenotypic and genetic characterization of the bacteriophage abortive infection mechanism AbiK from Lactococcus lactis.
<3>Appl. Environ. Microbiol.
<4>63
<5>1274-1283
<6>1997
<7>The natural plasmid pSRQ800 isolated from Lactococcus lactis subsp. lactis W1 conferred strong
phage resistance against small isometric phages of the 936 and P335 species when introduced
into phage-sensitive L. lactis strains.  It had very limited effect on prolate phages of the
c2 species.  The phage resistance mechanism encoded on pSRQ800 is a temperature-sensitive
abortive infection system (Abi).  Plasmid pSRQ800 was mapped, and the Abi genetic determinant
was localized on a 4.5-kb EcoRI fragment.  Cloning and sequencing of the 4.5-kb fragment
allowed the identification of two large open reading frames.  Deletion mutants showed that
only orf1 was needed to produce the Abi phenotype.  orf1 (renamed abiK) coded for a predicted
protein of 599 amino acids (AbiK) with an estimated molecular size of 71.4 kDa and a pI of
7.98.  DNA and protein sequence alignment programs found no significant homology with
databases.  However, a database query based on amino acid composition suggested that AbiK
might be in the same protein family as AbiA.  No phage DNA replication nor phage structural
protein production was detected in infected AbiK+ L. lactis cells.  This system is believed to
act at or prior to phage DNA replication.  When cloned into a high-copy vector, AbiK
efficiency increased 100-fold.  AbiK provides another powerful tool that can be used in
controlling phages during lactococcal fermentations.

<>

<1>Emond-Rheault, J.G., Vincent, A.T., Trudel, M.V., Brochu, F., Boyle, B., Tanaka, K.H., Attere, S.A., Jubinville, E., Loch, T.P., Winters, A.D., Faisal, M., Frenette, M., Derome, N., Charette, S.J.
<2>Variants of a genomic island in Aeromonas salmonicida subsp. salmonicida link isolates with their geographical origins.
<3>Vet. Microbiol.
<4>175
<5>68-76
<6>2014
<7>Aeromonas salmonicida subsp. salmonicida is a fish pathogen. Analysis of its
genomic characteristics is required to determine the worldwide distribution of
the various populations of this bacterium. Genomic alignments between the 01-B526
pathogenic strain and the A449 reference strain have revealed a 51-kb chromosomal
insertion in 01-B526. This insertion (AsaGEI1a) has been identified as a new
genomic island (GEI) bearing prophage genes. PCR assays were used to detect this
GEI in a collection of 139 A. salmonicida subsp. salmonicida isolates. Three
forms of this GEI (AsaGEI1a, AsaGEI1b, AsaGEI2a) are now known based on this
analysis and the sequencing of the genomes of seven additional isolates. A new
prophage (prophage 3) associated with AsaGEI2a was also discovered. Each GEI
appeared to be strongly associated with a specific geographic region. AsaGEI1a
and AsaGEI2a were exclusively found in North American isolates, except for one
European isolate bearing AsaGEI2a. The majority of the isolates bearing AsaGEI1b
or no GEI were from Europe. Prophage 3 has also a particular geographic
distribution and was found only in North American isolates. We demonstrated that
A. salmonicida subsp. salmonicida possesses unsuspected elements of genomic
heterogeneity that could be used as indicators to determine the geographic
origins of isolates of this bacterium.

<>

<1>Endlich, B., Linn, S.
<2>The DNA restriction endonuclease of Escherichia coli B.  I. Studies of the DNA translocation and the ATPase activities.
<3>J. Biol. Chem.
<4>260
<5>5720-5728
<6>1985
<7>Electron microscopic examination of DNA intermediates formed by the restriction
endonuclease of Escherichia coli B revealed supercoiled loops that are
presumably formed during an ATP-dependent DNA translocation process in which
the enzyme remains bound to the recognition site while tracking along the DNA
helix to a cleavage site.  The rate of DNA translocation during this process is
at least 5000 base pairs/min at 37C.  Even after all cleavages have been
completed, complexes are seen that contain terminal loops or loop plus tail
structures.  During this later phase of the reaction, ATP is hydrolyzed at a
rate which is dependent upon the size of the largest possible loop (or loop
plus tail); this ATP hydrolysis can be terminated by one double-strand cleavage
within the loop region between the recognition site and the terminus.  To
explain these results, it is hypothesized that after cleavage the enzyme cycles
between a tracking (and possibly backtracking) mode which is fueled by ATP
hydrolysis and a relatively long static period in which ATP hydrolysis does not
occur.  While tracking, the enzyme would be bound both to the recognition site
and to a distal site but, while static, the enzyme would be bound only at the
recognition site of nonlooped molecules.  This postnuclease phase of the
reaction is hypothesized to reflect a reaction whereby the enzyme initially
scans DNA molecules before making a strand cleavage.

<>

<1>Endlich, B., Linn, S.
<2>The DNA restriction endonuclease of Escherichia coli B.  II.  Further studies of the structure of DNA intermediates and products.
<3>J. Biol. Chem.
<4>260
<5>5729-5738
<6>1985
<7>The DNA intermediates and final products formed by the Type I restriction
endonuclease, EcoB, were further characterized.  DNA cleaved on only one strand
(hemi-restricted DNA) contains gaps of approximately 70-100 nucleotides, while
the fully restricted products contain 3'-single-stranded tails averaging
approximately 70-100 nucleotides for each strand cleaved.  The gaps and tails
are formed with the release of an equal number of nucleotides as small
oligonucleotides that are soluble in acid.  After purification, neither the
hemi-restricted nor the fully restricted DNAs are cleaved again by EcoB.  There
is no apparent specificity for which strand of a duplex is initially cleaved by
EcoB, nor is there specificity with respect to the composition of the
3'-terminal nucleotide formed on the DNA or the 3'- or 5'-terminal nucleotides
of the acid-soluble oligonucleotides released during DNA cleavage.  The
structure formed at the 5' terminus of the DNA product which blocks
phosphorylation by T4 polynucleotide kinase remains unknown, but its removal
with phage lambda exonuclease allows at least some reutilization of recognition
sites by EcoB as well as phosphorylation of the newly formed 5' termini.  To
explain the complex mechanism of this enzyme, it is suggested that the
unidentified 5'-tails prevent wasteful rerestriction from occurring, whereas
the 3'-single-stranded tails create DNA which, when nonhomologous to
chromosomal DNA, cannot be rescued because such tails are not a substrate for
DNA polymerases.  However, when homologous chromosomal DNA exists, the randomly
cleaved large fragments with these tails can easily be assimilated by
recA-mediated genetic recombination, thus stimulating DNA exchange between
related organisms.

<>

<1>Endlich, B., Linn, S.
<2>Type I restriction enzymes.
<3>The Enzymes, Academic Press, Boyer, P., 
<4>14
<5>137-156
<6>1981
<7>Many bacteria contain enzymatic systems that act to restrict the expression of
foreign DNA introduced through phage infection, conjugation, or transformation.
This phenomenon of host-controlled specificity was first described in the
early 1950s by Luria and Human, who studied T-even phages, and by Bertani and
Weigle, who investigated the restriction of the host ranges of lambda and P2
phages.  It was observed that a particular phage could have widely different
efficiencies of infection on several closely related bacterial strains, but
when phages that had initially plated with low efficiency were replated on the
same bacterial strain, the efficiency of infection increased dramatically.
This modification of host range was not a hereditary genetic adaptation,
however, since it could be lost by subsequent propagation of the phage in
another bacterial strain.  It was subsequently shown that the host-specificity
system described by Luria and Human was unique to the T-even phages, whereas
the system described by Bertani and Weigle was more widespread.  The type I
restriction-modification systems described in this review are of the latter,
more general, class.  Modification of the T-even phages involves the
glycosylation of hydroxymethylcytosine residues, and has been reviewed
elsewhere.  Further investigation of lambda phage host specificity by Arber and
coworkers led to the hypothesis of a molecular mechanism in which special DNA
sequences are acted upon by appropriate restriction and modification enzymnes.
Foreign DNA that happens to contain such specific sequences is cleaved by a
restriction endonuclease upon entering the cell.  When these same sequence
exist in the bacterium's own DNA, however, they are protected by a modification
methylase that imparts methylase that imparts methyl groups to bases within the
sequences, rendering them resistant to endonuclease action.  This hypothesis
was substantiated in the late 1960s when restriction endonucleases from the E.
coli strains K and B were discovered and isolated in highly purified form.
These nucleases were genetically identified as the restriction enzymes and
observed to be complex in terms of cofactor requirements, subunit composition,
and interactions with the DNA substrate.  Other restriction enzymes were soon
isolated from other bacteria such as Haemophilus influenzae.  Many of these
endonucleases have distinctly different properties from those of the E. coli B
and K nucleases, and are distinguished as two additional types of restriction
endonucleases.  Whether all of these latter enzymes function in situ in a
restriction capacity is not clear.

<>

<1>Endo, M., Majima, T.
<2>Protein variants with photo-reactivity and therapeutic products containing the protein variants.
<3>Japanese Patent Office
<4>JP 2003310253 A
<5>
<6>2003
<7>
<>

<1>Endo, M., Nakayama, K., Majima, T.
<2>Design and synthesis of photochemically controllable restriction endonuclease BamHI by manipulating the salt-bridge network in the dimer  interface.
<3>J. Org. Chem.
<4>69
<5>4292-4298
<6>2004
<7>The strategy for the design of photochemically controllable enzymes by manipulating the dimer
interface is described. Employing a restriction
endonuclease BamHI, the selective incorporation of amino acids having a
photoremovable 6-nitroveratryl group into the specific position
(Lys132) in the dimer interface of the BamHI mutant (H133A) was
performed. The activity of the photofunctionalized BamHI mutant was
significantly suppressed, and the following photoirradiation induced
the recovery of the activity. In addition, uncaging of the
6-nitroveratryl group introduced to Lys132 did not seriously reduce the
catalytic activity and affinity for the substrate. These results
indicate that the activity of the enzyme can be effectively regulated
by caging and uncaging of the specific amino acid in the dimer
interface using the photoremovable group.

<>

<1>Endow, S.A.
<2>Analysis of Drosophila melanogaster Satellite IV with restriction endonuclease MboII.
<3>J. Mol. Biol.
<4>114
<5>441-449
<6>1977
<7>The products of digestion of Drosophila melanogaster satellite IV DNA with
restriction endonuclease MboII have been analysed and found to be consistent
with a repeating pentamer sequence (A-G-A-A-G)n for satellite IV.  More than
95% of the satellite DNA is digested to fragments less than 25 base-pairs in
length, suggesting that the DNA sequence is highly conserved.

<>

<1>Endow, S.A., Roberts, R.J.
<2>Two restriction-like enzymes from Xanthomonas malvacearum.
<3>J. Mol. Biol.
<4>112
<5>521-529
<6>1977
<7>Two sequence-specific endonucleases, XmaI and XmaII, have been purified from
Xanthomonas malvacearum.  XmaI makes cuts in bacteriophage lambda and
adenovirus-2 DNA identical with those produced by SmaI, a restriction-like
enzyme previously isolated from Serratia marcescens Sb.XmaI cleaves within the
sequence 5'-C^-C-C-G-G-G-3' 3'-G-G-G-C-G-^C-5' at the sites indicated by the
arrows.  XmaII is an isoschizomer of the specific endonuclease PstI previously
isolated from Providencia stuartii.

<>

<1>Engel, P.
<2>Plasmid transformation of Streptomyces tendae after heat attenuation of restriction.
<3>Appl. Environ. Microbiol.
<4>53
<5>1-3
<6>1987
<7>Streptomyces tendae ATCC 31160 produces nikkomycin, a fungicide and insecticide
that inhibits chitin synthases.  Exposure of S. tendae protoplasts to 50C for
30 min is required for transformation (10-2 thiostrepton-resistant
transformants per microgram of DNA) with plasmid pIJ702 or pIJ680 from
Streptomyces lividans.  pIJ702 and pIJ680 DNA isolated from the S. tendae
transformants is efficient (10-6 to 10-7 transformants per microgram DNA) in
subsequent transformations of S. tendae protoplasts generated at 30C.  PstI
fails to cut the single PstI site in pIJ702 and cuts only one of the two PstI
sites in pIJ680 DNA isolated from S. tendae transformants.  Digests of plasmid
DNA mixtures showed that plasmid DNA from S. tendae does not inhibit PstI
activity.  pIJ702 and pIJ680 DNA from S. tendae transformants was used to
transform S. lividans to show that plasmid DNA remains unchanged, except for
modification at some PstI sites in S. tendae, as a consequence of passage
through S. tendae.  The DNA modification is lost when S. lividans is
transformed with plasmid DNA from S. tendae transformants.  Since S. tendae
modifies only some PstI sites, it appears the modification (presumably
restriction activity also) activity in S. tendae recognizes a sequence that
includes or overlaps the PstI hexanucleotide recognition sequence.

<>

<1>Engel, P., Vizcaino, M.I., Crawford, J.M.
<2>Gut symbionts from distinct hosts exhibit genotoxic activity via divergent colibactin biosynthetic pathways.
<3>Appl. Environ. Microbiol.
<4>0
<5>0
<6>2014
<7>Secondary metabolites produced by nonribosomal peptide synthetase (NRPS) or
polyketide synthase (PKS) pathways are chemical mediators of microbial
interactions in diverse environments. However, little is known about their
distribution, evolution, and functional roles in bacterial symbionts associated
with animals. A prominent example is "colibactin", a largely unknown family of
secondary metabolites produced by Escherichia coli via a hybrid NRPS-PKS
biosynthetic pathway, inflicting DNA damage upon eukaryotic cells and
contributing to colorectal cancer and tumor formation in the mammalian gut. Thus
far, homologs of this pathway have only been found in closely related
Enterobacteriaceae, while a divergent variant of this gene cluster was recently
discovered in a marine alphaproteobacterial Pseudovibrio strain. Herein, we
sequenced the genome of Frischella perrara PEB0191, a bacterial gut symbiont of
honey bees, and identified a homologous colibactin biosynthetic pathway related
to those found in Enterobacteriaceae. We show that the colibactin genomic island
(GI) has conserved gene synteny and biosynthetic module architecture across F.
perrara, Enterobacteriaceae and the Pseudovibrio strain. Comparative metabolomics
analyses of F. perrara and E. coli further reveal that these two bacteria produce
related colibactin pathway-dependent metabolites. Finally, we demonstrate that F.
perrara, like E. coli, causes DNA damage in eukaryotic cells in vitro in a
colibactin pathway-dependent manner. Together, these results support that
divergent variants of the colibactin biosynthetic pathway are widely distributed
among bacterial symbionts, producing related secondary metabolites and likely
endowing its producer with functional capabilities important for diverse
symbiotic associations.

<>

<1>Engin, D., Hascelik, G.
<2>iceA genotypes may play a role in acquiring metronidazole resistance in Helicobacter pylori.
<3>Helicobacter
<4>8
<5>356
<6>2003
<7>Homologous recombination contributes to the extraordinary genetic diversity of Helicobacter
pylori.  Recently, evidence of in vivo horizontal gene transfer was detected in the rdxA gene,
which is involved in metronidazole resistance.  This bacterium possesses a large number of
restriction - modification systems that afford protection against transforming DNA, apparently
regulating the natural transformation of H. pylori.  The newly discovered virulence factor
icaA exists in two variants, iceA1 which encodes an NlaIIIR homologue restriction enzyme and
iceA2.  In this study, we evaluated the relationship between the metronidazole resistance and
iceA genotypes in 76 H. pylori strains.  We determined metronidazole MIC values of H. pylori
strains by agar dilution.  Following DNA purification iceA genotypes of H. pylori strains were
determined by using PCR.  Thirty-eight (50.0%) out of 76 H. pylori strains were found
metronidazole resistant.  MIC50 was 8 and MIC90 value was 128 ug/ml.  Forty-eight (63.2%) and
19 (25.0%) of the strains were typed as iceA1 and iceA2, respectively.  Whereas, 9 (11.8%)
strains yielded PCR products of unexpected length and typed as iceA1a/2a.  No significant
difference was found between metronidazole MIC values of iceA1 and iceA2 strains.
Metronidazole MIC values of iceA1a/2a strains were marginally lower than iceA2 strains
(Mann-Whitney p=0.075).  We conclude that, restriction-modification systems may play an
important role in acquiring metronidazole resistance in H. pylori.  The organization of iceA
locus in iceA1a/2a strains should be further evaluated.

<>

<1>Engler, L.E., Sapienza, P., Dorner, L.F., Kucera, R., Schildkraut, I., Jen-Jacobson, L.
<2>The energetics of the interaction of BamHI endonuclease with its recognition site GGATCC.
<3>J. Mol. Biol.
<4>307
<5>619-636
<6>2001
<7>The interaction of BamHI endonuclease with DNA has been studied crystallographically, but has
not been characterized rigorously in solution. The enzyme binds in solution as a homodimer to
its recognition site GGATCC. Only six base-pairs are directly recognized, but binding affinity
(in the absence of the catalytic cofactor Mg(2+)) increases 5400-fold as oligonucleotide
length increases from 10 to 14 bp. Binding is modulated by sequence context outside the
recognition site, varying about 30-fold from the best (GTG or TAT) to the worst (CGG)
flanking triplets. BamHI, EcoRI and EcoRV endonucleases all have different context
preferences, suggesting that context affects binding by influencing the free energy levels of
the complexes rather than that of the free DNA. Ethylation interference footprinting in the
absence of divalent metal shows a localized and symmetrical pattern of phosphate contacts,
with strong contacts at NpNpNpGGApTCC. In the presence of Mg(2+), first-order cleavage rate
constants are identical in the two GGA half-sites, are the same for the two nicked
intermediates and are unaffected by substrate length in the range 10-24 bp. DNA binding is
strongly enhanced by mutations D94N, E111A or E113K, by binding of Ca(2+) at the active site,
or by deletion of the scissile phosphate GpGATCC, indicating that a cluster of negative
charges at the catalytic site contributes at least 3-4 kcal/mol of unfavorable binding free
energy. This electrostatic repulsion destabilizes the enzyme-DNA complex and favors metal ion
binding and progression to the transition state for cleavage.

<>

<1>Engler, L.E., Welch, K.K., Jen-Jacobson, L.
<2>Specific binding by EcoRV endonuclease to its DNA recognition site GATATC.
<3>J. Mol. Biol.
<4>269
<5>82-101
<6>1997
<7>Restriction endonuclease EcoRV has been reported to be unable to distinguish its specific DNA
site, GATATC, from non-specific DNA sites in the absence of the catalytic cofactor Mg2+, and
thus to exercise sequence specificity solely in the catalytic step.  In contrast, we show here
that under appropriate conditions of pH and salt concentration, specific complexes with
oligonucleotides containing the GATATC site can be detected by either filter-binding or
gel-retardation.  Equilibrium binding constants (KA) are easily measured by both direct
equilibrium and equilibrium-competition methods.  The preference for "specific" over
"non-specific" binding at pH 7 in the absence of divalent cations is about 1000-fold (per mole
of oligonucleotide) or 12,000-fold (per mole of binding sites).  Ethylation-interference
footprinting shows that the "specific" complex includes strong contacts to the phosphate
groups GpApTpApTC.  Specific DNA binding is strongly pH-dependent, decreasing about 15-fold
for each increase of one pH unit above pH 6, but non-specific binding is not; thus, binding
specificity decreases with increasing pH.  Gel retardation and filter-binding at pH values
less than 7 yielded essentially identical values of KA for specific-site binding, but at pH>7
gel retardation significantly underestimates KA.  Specific-site binding is stimulated about
700-fold by Ca2+ (not a cofactor for cleavage), but with non-cleavable 3'-phosphorothioate
and 4'-thiodeoxyribose derivatives whose response to Ca2+ is similar to that of the parent
oligonucleotide, Mg2+ stimulates binding only fourfold and twofold, respectively.  Thus,
binding specificity is not dramatically enhanced by Mg2+.  Taking into account discrimination
in binding and in the first-order rate constant for phosphodiester bond scission, the overall
discrimination exercised against the incorrect site GTTATC is about 10^7-fold.  EcoRV
endonuclease is thus not a "new paradigm" for site-specific interaction without binding
specificity, but like other type II restriction endonucleases achieves sequence specificity by
discriminating both in DNA binding and in catalysis.

<>

<1>Enikeeva, F.N., Severinov, K.V., Gelfand, M.S.
<2>Restriction-modification systems and bacteriophage invasion: Who wins?
<3>J. Theor. Biol.
<4>266
<5>550-559
<6>2010
<7>The success of a phage that infects a bacterial cell possessing a restriction-modification
(R-M) system depends on the activities of the
host methyltransferase and restriction endonuclease, and the number of
susceptible sites in the phage genome. However, there is no model
describing this dependency and linking it to observable parameters such
as the fraction of surviving cells under excess phage, or probability
of plating at low amount of phages. We model the phage infection of a
cell with a R-M system as a pure birth process with a killing state. We
calculate the transitional probabilities and the stationary
distribution for this process. We generalize the model developed for a
single cell to the case of multiple identical cells invaded by a
Poisson-distributed number of phages. The R-M enzyme activities are
assumed to be constant, time-dependent, or random. The obtained results
are used to estimate the ratio of the methyltransferase and
endonuclease activities from the observed fraction of surviving cells.

<>

<1>Epinat, J.-C., Lacroix, E.
<2>Homing endonuclease I-DmoI mutants with enhanced activity at 37.degree.C and use thereof.
<3>European Patent Office
<4>EP 1591521 A
<5>
<6>2005
<7>I-DmoI derivatives with enhanced cleavage activity at 37oC, said mutant comprising a sequence
of a mutant of a I-DmoI endonuclease or a chimeric derivative thereof including at least the
first I-DmoI domain, said sequence comprising the substitution of at least: (i) one of the
residues in positions 4, 20, 49, 52, 92, 94 and/or 95 of said first I-DmoI domain, and/or (ii)
one of the residues in positions 101, 102, and/or 109 of the linker or the beginning of the
second domain of I-DmoI, if present.  Polynucleotide encoding said derivatives, cell, animal
or plant comprising said polynucleotide and use thereof for isolating meganucleases with new
DNA target specificity.

<>

<1>Epinat, J.C., Arnould, S., Chames, P., Rochaix, P., Desfontaines, D., Puzin, C., Patin, A., Zanghellini, A., Paques, F., Lacroix, E.
<2>A novel engineered meganuclease induces homologous recombination in yeast and mammalian cells.
<3>Nucleic Acids Res.
<4>31
<5>2952-2962
<6>2003
<7>Homologous gene targeting is the ultimate tool for reverse genetics, but its use is often
limited by low efficiency. In a number of recent studies,
site- specific DNA double-strand breaks (DSBs) have been used to induce
efficient gene targeting. Engineering highly specific, dedicated DNA
endonucleases is the key to a wider usage of this technology. In this
study, we present two novel, chimeric meganucleases, derived from homing
endonucleases. The first one is able to induce recombination in yeast and
mammalian cells, whereas the second cleaves a novel (chosen) DNA target
site. These results are a first step toward the generation of custom
endonucleases for the purpose of targeted genome engineering.

<>

<1>Eppinger, M. et al.
<2>Genome Sequences of the Biotechnologically Important Bacillus megaterium Strains QM B1551 and DSM319.
<3>J. Bacteriol.
<4>193
<5>4199-4213
<6>2011
<7>Bacillus megaterium is deep-rooted in the Bacillus phylogeny, making it an evolutionarily key
species and of particular importance in understanding
genome evolution, dynamics, and plasticity in the bacilli. B. megaterium
is a commercially available, nonpathogenic host for the biotechnological
production of several substances, including vitamin B(12), penicillin
acylase, and amylases. Here, we report the analysis of the first complete
genome sequences of two important B. megaterium strains, the plasmidless
strain DSM319 and QM B1551, which harbors seven indigenous plasmids. The
5.1-Mbp chromosome carries approximately 5,300 genes, while QM B1551
plasmids represent a combined 417 kb and 523 genes, one of the largest
plasmid arrays sequenced in a single bacterial strain. We have documented
extensive gene transfer between the plasmids and the chromosome. Each
strain carries roughly 300 strain-specific chromosomal genes that account
for differences in their experimentally confirmed phenotypes. B.
megaterium is able to synthesize vitamin B(12) through an
oxygen-independent adenosylcobalamin pathway, which together with other
key energetic and metabolic pathways has now been fully reconstructed.
Other novel genes include a second ftsZ gene, which may be responsible for
the large cell size of members of this species, as well as genes for gas
vesicles, a second beta-galactosidase gene, and most but not all of the
genes needed for genetic competence. Comprehensive analyses of the global
Bacillus gene pool showed that only an asymmetric region around the origin
of replication was syntenic across the genus. This appears to be a
characteristic feature of the Bacillus spp. genome architecture and may be
key to their sporulating lifestyle.

<>

<1>Eppinger, M., Baar, C., Linz, B., Raddatz, G., Lanz, C., Keller, H., Morelli, G., Gressmann, H., Achtman, M., Schuster, S.C.
<2>Who ate whom? Adaptive Helicobacter genomic changes that accompanied a host jump from early humans to large felines.
<3>PLoS Genet.
<4>2
<5>e120
<6>2006
<7>Helicobacter pylori infection of humans is so old that its population genetic structure
reflects that of ancient human migrations. A closely
related species, Helicobacter acinonychis, is specific for large felines,
including cheetahs, lions, and tigers, whereas hosts more closely related
to humans harbor more distantly related Helicobacter species. This
observation suggests a jump between host species. But who ate whom and
when did it happen? In order to resolve this question, we determined the
genomic sequence of H. acinonychis strain Sheeba and compared it to
genomes from H. pylori. The conserved core genes between the genomes are
so similar that the host jump probably occurred within the last 200,000
(range 50,000-400,000) years. However, the Sheeba genome also possesses
unique features that indicate the direction of the host jump, namely from
early humans to cats. Sheeba possesses an unusually large number of highly
fragmented genes, many encoding outer membrane proteins, which may have
been destroyed in order to bypass deleterious responses from the feline
host immune system. In addition, the few Sheeba-specific genes that were
found include a cluster of genes encoding sialylation of the bacterial
cell surface carbohydrates, which were imported by horizontal genetic
exchange and might also help to evade host immune defenses. These results
provide a genomic basis for elucidating molecular events that allow
bacteria to adapt to novel animal hosts.

<>

<1>Eppinger, M., Daugherty, S., Agrawal, S., Galens, K., Sengamalay, N., Sadzewicz, L., Tallon, L., Cebula, T.A., Mammel, M.K., Feng, P., Soderlund, R., Tarr, P.I., Debroy, C., Dudley, E.G., Fraser, C.M., Ravel, J.
<2>Whole-Genome Draft Sequences of 26 Enterohemorrhagic Escherichia coli O157:H7 Strains.
<3>Genome Announcements
<4>1
<5>e0013412
<6>2013
<7>First identified in 1982, Escherichia coli O157:H7 is the dominant enterohemorrhagic serotype
underlying food-borne human infections in North
America. Here, we report the genomes of twenty-six strains derived from patients
and the bovine reservoir. These resources enable detailed whole-genome
comparisons and permit investigations of genotypic and phenotypic plasticity.

<>

<1>Eppinger, M., Guo, Z., Sebastian, Y., Song, Y., Lindler, L.E., Yang, R., Ravel, J.
<2>Draft Genome Sequences of Yersinia pestis Isolates From Natural Foci of Endemic Plague in China.
<3>J. Bacteriol.
<4>191
<5>7628-7629
<6>2009
<7>To gain insights into the evolutionary origin, emergence and pathogenicity of the etiologic
agent of plague, we have sequenced the genomes of four  Yersinia pestis strains isolated from
the zoonotic rodent reservoir in  endemic plague foci in China (24). These resources enable
in-depth studies  of Y. pestis sequence variations, and detailed whole genome comparisons of
very closely related genomes from the supposed site of the origin and emergence of global
pandemics of plague.

<>

<1>Eppinger, M., Mammel, M.K., Leclerc, J.E., Ravel, J., Cebula, T.A.
<2>Genome Signatures of Escherichia coli O157:H7 Isolates from the Bovine Host Reservoir.
<3>Appl. Environ. Microbiol.
<4>77
<5>2916-2925
<6>2011
<7>Cattle comprise a main reservoir of Shiga toxin-producing Escherichia coli
O157:H7 (STEC). The significant differences in host prevalence,
transmissibility, and virulence phenotypes among strains from bovine and
human sources are of major interest to the public health community and
livestock industry. Genomic analysis revealed divergence into three
lineages: lineage I and lineage I/II strains are commonly associated with
human disease, while lineage II strains are overrepresented in the
asymptomatic bovine host reservoir. Growing evidence suggests that
genotypic differences between these lineages, such as polymorphisms in
Shiga toxin subtypes and synergistically acting virulence factors, are
correlated with phenotypic differences in virulence, host ecology, and
epidemiology. To assess the genomic plasticity on a genome-wide scale, we
have sequenced the whole genome of strain EC869, a bovine-associated E.
coli O157:H7 isolate. Comparative phylogenomic analysis of this key
isolate enabled us to place accurately bovine lineage II strains within
the genetically homogenous E. coli O157:H7 clade. Identification of
polymorphic loci that are anchored both in the chromosomal backbone and
horizontally acquired regions allowed us to associate bovine genotypes
with altered virulence phenotypes and host prevalence. This study
catalogued numerous novel lineage II-specific genome signatures, some of
which appear to be associated intimately with the altered pathogenic
potential and niche adaptation within the bovine rumen. The presented
extended list of polymorphic markers is valuable in the development of a
robust typing system critical for forensic, diagnostic, and
epidemiological studies of this emerging human pathogen.

<>

<1>Eppinger, M., McNair, K., Zogaj, X., Dinsdale, E.A., Edwards, R.A., Klose, K.E.
<2>Draft Genome Sequence of the Fish Pathogen Piscirickettsia salmonis.
<3>Genome Announcements
<4>1
<5>e00926-13
<6>2013
<7>Piscirickettsia salmonis is a Gram-negative intracellular fish pathogen that has  a
significant impact on the salmon industry. Here, we report the genome sequence
of P. salmonis strain LF-89. This is the first draft genome sequence of P.
salmonis, and it reveals interesting attributes, including flagellar genes,
despite this bacterium being considered nonmotile.

<>

<1>Epstein, D.M.
<2>DNA-methylase linking reaction.
<3>US Patent Office
<4>US 5856090
<5>
<6>1999
<7>The activity of sequence-specific DNA binder proteins, such as DNA methylases, provides a
method of obtaining a covalent linkage between a nucleic acid segment and a polypeptide
determinant encoded by the nucleic acid segment.  The polypeptide determinant is expressed as
a fusion protein together with the DNA methylase, which binds in vivo to a cytidine suicide
analog when present in a nucleotide sequence.  A plasmid suitable for use in this linkage
reaction can comprise: (1) a gene fusion construct including a gene encoding a DNA methylase
and a gene encoding a polypeptide determinant; (2) a promoter for transcription of the gene
fusion construct as messenger RNA; and (3) a methylase conjugation element linked to the gene
fusion sequence, the methylase conjugation element including a methylase binding site having
at least one copy of a nucleotide sequence including a cytidine suicide analog capable of
irreversibly binding the DNA methylase.  The plasmid can form a plasmid-polypeptide
determinant conjugate.  The plasmids and methods of the present invention are useful for in
vitro evolution of proteins.

<>

<1>Erdmann, D.
<2>Cloning of restriction-modification systems from Herpetosiphon giganteus Hpg9 and Hpg24 in E. coli - Characterization and comparative analysis of the genetic organization and structure.
<3>Ph.D. Thesis, Justus-Liebig University, Geissen, W. Germany
<4>
<5>1-204
<6>1991
<7>None

<>

<1>Erdmann, D., Dusterhoft, A., Kroger, M.
<2>Cloning and molecular characterization of the HgiCI restriction/modification system from Herpetosiphon giganteus Hpg9 reveals high similarity to BanI.
<3>Eur. J. Biochem.
<4>202
<5>1247-1256
<6>1991
<7>The genes coding for the GGYRCC specific restriction/modification system HgiCI from
Herpetosiphon giganteus Hpg9 have been cloned in Escherichia coli in three steps. As an
initial step, the methyltransferase gene could be obtained after heterologous in vitro
selection of a plasmid gene bank by cleavage with the isoschizomeric restriction endonuclease
BanI. The adjacent endonuclease gene was cloned following Southern blot analysis of flanking
genomic regions. The two genes code for polypeptides of 420 amino acids (M.HgiCI) and 345
amino acids (R.HgiCI). Establishing a functional endonuclease gene could only be achieved
using a tightly regulated expression system or by methylation of the genomic DNA prior to
transformation of the endonuclease gene. The methyltransferase M.HgiCI shows significant
similarities to the family of 5-methylcytidine methyltransferases. Striking similarities could
be found with both the isoschizomeric endonuclease and methyltransferase of the BanI
restriction/modification system from Bacillus aneurinolyticus.

<>

<1>Erdmann, D., Horst, G., Dusterhoft, A., Kroger, M.
<2>Stepwise cloning and genetic organization of the seemingly unclonable HgiCII restriction-modification system from Herpetosiphon giganteus strain Hpg9, using PCR technique.
<3>Gene
<4>117
<5>15-22
<6>1992
<7>The genes, hgiCIIR and hgiCIIM, that encode the HgiCII restriction and modification (R-M)
system from Herpetosiphon giganteus strain Hpg9, an AvaII isoschizomer recognizing the
sequence, CCA/TCC, were cloned in Escherichia coli. Cloning the respective hgiCIIM gene was
achieved via in vitro selection both from a Sau3AI- and an NheI-generated plasmid gene library
using AvaII, a commercially available isoschizomer of HgiCII. However, all attempts to clone
the closely linked hgiCIIR and M genes in a single step resulted in deletions spanning parts
of the coding region of hgiCIIR. Therefore, cloning of the missing 3'-terminal part of this
gene was achieved by applying the inverse polymerase-chain-reaction technique. All attempts to
construct an enzymatically active R.HgiCII failed; only the inactivated hgiCIIR gene could be
cloned. Sequencing of the hgiCIIRM region (carrying predesigned small mutations in the R gene)
disclosed three open reading frames (ORFs):one small ORF preceding the methyltransferase
(MTase)-encoding gene, plus those encoding M.HgiCII (49620 Da) and R.HgiCII (30891 Da).
M.HgiCII exhibits the common motif of ten conserved amino-acid blocks typically found within
the group of m5C-MTases. The R-M system of HgiCII reveals strong homologies to the
isoschizomeric R-M system of HgiBI from H. giganteus strain Hpg5, which in contrast, could be
cloned in one step.

<>

<1>Erkel, C., Kube, M., Reinhardt, R., Liesack, W.
<2>Genome of Rice Cluster I archaea--the key methane producers in the rice rhizosphere.
<3>Science
<4>313
<5>370-372
<6>2006
<7>Rice fields are a global source of the greenhouse gas methane, which is
produced by methanogenic archaea, and by methanogens of Rice Cluster I
(RC-I) in particular. RC-I methanogens are not yet available in pure
culture, and the mechanistic reasons for their prevalence in rice fields
are unknown. We reconstructed a complete RC-I genome (3.18 megabases)
using a metagenomic approach. Sequence analysis demonstrated an
aerotolerant, H2/CO2-dependent lifestyle and enzymatic capacities for
carbohydrate metabolism and assimilatory sulfate reduction, hitherto
unknown among methanogens. These capacities and a unique set of
antioxidant enzymes and DNA repair mechanisms as well as
oxygen-insensitive enzymes provide RC-I with a selective advantage over
other methanogens in its habitats, thereby explaining the prevalence of
RC-I methanogens in the rice rhizosphere.

<>

<1>Erlanson, D.A., Chen, L., Verdine, G.L.
<2>DNA methylation through a locally upaired intermediate.
<3>J. Am. Chem. Soc.
<4>115
<5>12583-12584
<6>1993
<7>Methylation of DNA serves essential roles in mammalian development and in bacterial resistance
to viral pathogens. In this process, a DNA (cytosine-5)-methyltransferase (DCMtase) mediates
delivery of a methyl group from S-adenosyl-L-methionine to the 5-position of cytosine residues
in DNA. DCMtases operate by conjugate addition of a cysteine thiolate to the 6-carbon (C6) of
the substrate cytosine followed by transfer of a methyl group to C5. __-Elimination
regenerates the free enzyme (Figure 1). We have noted that the stereoelectronic attack
trajectories for thiolate addition and methyl transfer cannot be accommodated in a canonical
B-form duplex, suggesting that DCMtases cause transient helical disruption during the
catalytic event. Here we report evidence in favor of DCMtase-induced distortion of DNA and
propose a structural model for the enzyme-DNA intermediate.

<>

<1>Erlanson, D.A., Wolfe, S.A., Chen, L., Verdine, G.L.
<2>Selective base-pair destabilization enhances binding of a DNA methyltransferase.
<3>Tetrahedron
<4>53
<5>12041-12056
<6>1997
<7>Disulfide crosslinks were introduced into the minor groove of DNA using the convertible
nucleoside approach.  Depending upon the length of the tether, the modified base pairs were
either stabilized or destabilized.  When the base-pairs were destabilized, the oligonucleotide
was bound by a unmodified oligonucleotide.  Insights into the mechanism of DCMtases are
discussed.

<>

<1>Erova, T., Sha, J., Fadl, A., Khajanchi, B., Chopra, A.
<2>Mutations within the catalytic site of DNA methyltransferase (Dam) of Aeromonas hydrophila reverts the virulence of the dam-overproducing strain to that of the wild-type bacterium.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>106
<5>76
<6>2006
<7>A. hydrophila is an opportunistic human pathogen. In our recently published studies, we
demonstrated that DNA adenine methyltransferase (Dam) of A. hydrophila is crucial for
bacterial viability, and that it affects virulence of the bacterium by altering biological
activities associated with type 2 and type 3 secreted proteins. Overall, overproduction of Dam
in A. hydrophila using an arabinose-inducible pBAD vector attenuated bacterial virulence in a
mouse model of infection. In this study, we demonstrated that the virulence potential of
Dam-overproducing strain of A. hydrophila was reverted back to that of the wild-type (WT)
bacterium when the catalytic site of Dam was mutated. Using Altered Sites in vitro Mutagenesis
System, we mutated aspartic acid (D) and tyrosine (Y) residues to alanine (A) within the
conserved catalytic motif (DPPY) of Dam. To confirm that the mutated enzyme lost its
methyltransferase (MTase) activity, we transformed pBAD/dam*D/A, pBAD/dam*Y/A, and
pBAD/damnative (as a control) recombinant plasmids into E. coli GM33 (Dam-) strain. Genomic
DNA (gDNA) isolated from either the E. coli GM33 (pBAD/dam*D/A) or the E. coli GM33
(pBAD/dam*Y/A) strain grown in the presence of arabinose was sensitive to DpnII digestion and
resistant to DpnI restriction endonuclease cutting. These data verified that the gDNA were not
methylated as the gDNA from E. coli GM33 strain with pBAD/damnative plasmid which exhibited
opposite sensitivity to DpnI and resistance to DpnII digestion. Overproduction of mutated Dam
in A. hydrophila resulted in bacterial motility, T3SS-associated cytotoxicity as well as
hemolytic and cytotoxic activity associated with the cytotoxic enterotoxin and that of the
protease activity similar to that of the WT bacterium which harbored the pBAD vector alone. In
addition, we noted that lactone production, an indicator of quorum sensing, was increased when
the native dam gene was over-expressed, with its levels returning to that of the WT bacterium
when the damgene was mutated. Taken together, our data indicated that MTase activity is
essential for altered virulence of A. hydrophila.

<>

<1>Erova, T.E., Fadl, A.A., Sha, J., Khajanchi, B.K., Pillai, L.L., Kozlova, E.V.C.A.K.
<2>Mutations within the catalytic motif of DNA adenine methyltransferase (Dam) of Aeromonas hydrophila cause the virulence of the Dam-overproducing strain to revert to that of the wild-type phenotype - bacterium DNA denine-methyltransferase catalytic.
<3>Infect. Immun.
<4>74
<5>5763-5772
<6>2006
<7>AUTHOR ABSTRACT - In this study, we demonstrated that the methyltransferase activity
associated with Dam was essential for
attenuation of Aeromonas hydrophila virulence. We mutated aspartic acid
and tyrosine residues to alanine within the conserved DPPY catalytic
motif of Dam and transformed the pBAD/dam(D/A), pBAD/dam(Y/A), and
pBAD/damA(AhSSu) (with the native dam gene) recombinant plasmids into
the Escherichia coli GM33 (dam-deficient) strain. Genomic DNA (gDNA)
isolated from either of the E. coli GM33 strains harboring the pBAD
vector with the mutated dam gene was resistant to DpnI digestion and
sensitive to DpnII restriction endonuclease cutting. These findings
were contrary to those with the gDNA of E. coli GM33 strain containing
the pBAD/dam(AhSSU) plasmid, indicating nonmethylation of E. coli gDNA
with mutated Dam. Overproduction of mutated Dam in A. hydrophild
resulted in bacterial motility, hemolytic and cytotoxic activities
associated with the cytotoxic enterotoxin (Act), and protease activity
similar to that of the wild-type (WT) bacterium, which harbored the
pBAD vector and served as a control strain. On the contrary,
overproduction of native Dam resulted in decreased bacterial motility,
increased Act-associated biological effects, and increased protease
activity. Lactone production, an indicator of quorum sensing, was
increased when the native dam gene was overexpressed, with its levels
returning to that of the control strain when the dam gene was mutated.
These effects of Dam appeared to be mediated through a regulatory
glucose-inhibited division A protein. Infection of mice with the
mutated Dam-overproducing strains resulted in mortality rates similar
to those for the control strain, with 100% of the animals dying within
2 to 3 days with two 50% lethal doses (LD(50)s) of the WT bacterium.
Importantly, immunization of mice with a native-Dam-overproducing
strain at the same LD50 did not result in any lethality and provided
protection to animals after subsequent challenge with a lethal dose of
the control strain.

<>

<1>Erova, T.E., Kosykh, V.G., Sha, J., Chopra, A.K.
<2>DNA adenine methyltransferase (Dam) controls the expression of the cytotoxic enterotoxin (act) gene of Aeromonas hydrophila via tRNA modifying enzyme-glucose-inhibited division protein (GidA).
<3>Gene
<4>498
<5>280-287
<6>2012
<7>Aeromonas hydrophila is both a human and animal pathogen, and the cytotoxic enterotoxin (Act)
is a crucial virulence factor of this bacterium because of its associated hemolytic,
cytotoxic, and enterotoxic activities.
Previously, to define the role of some regulatory genes in modulating Act production, we
showed that deletion of a glucose-inhibited division gene (gidA) encoding tRNA methylase
reduced Act levels, while overproduction
of DNA adenine methyltransferase (Dam) led to a concomitant increase in Act-associated
biological activities of a diarrheal isolate SSU of A.  hydrophila. Importantly, there are
multiple GATC binding sites for Dam within an upstream sequence of the gidA gene and one such
target site in the act gene upstream region.  We showed the dam gene to be essential for the
viability of A. hydrophila SSU, and, therefore, to better understand
the interaction of the encoding genes, Dam and GidA, in act gene regulation, we constructed a
gidA in-frame deletion mutant of Escherichia coli GM28 (dam+) and GM33 (Ädam) strains. We then
tested the expressional activity of the act and gidA genes by using a promoterless pGlow-TOPO
vector containing a
reporter green fluorescent protein (GFP). Our data indicated that in GidA+ strains of E. coli,
constitutive methylation of the GATC site(s) by Dam negatively regulated act and gidA gene
expression as measured by
GFP production. However, in the ÄgidA strains, irrespective of the presence or absence of
constitutively active Dam, we did not observe any alteration in the expression of the act gene
signifying the role of GidA in positively
regulating Act production. To determine the exact mechanism of how Dam and GidA influence Act,
a real-time quantitative PCR (RT-qPCR) assay was performed. The analysis indicated an increase
in gidA and act gene expression in the A. hydrophila Dam-overproducing strain, and these data
matchedwith Act production in the E. coli GM28 strain. Thus, the extent of DNA methylation
caused by constitutive versus overproduction of Dam, as well as possible conformation of DNA
influence the expression of act and gidA genes in A. hydrophila SSU.  Our results indicate
that the act gene is under the control of both Dam and GidA modification methylases, and Dam
regulates Act production via GidA.

<>

<1>Erova, T.E., Pillai, L., Fadl, A.A., Sha, J., Wang, S.F., Galindo, C.L., Chopra, A.K.
<2>DNA adenine methyltransferase influences the virulence of Aeromonas hydrophila.
<3>Infect. Immun.
<4>74
<5>410-424
<6>2005
<7>Among the various virulence factors produced by Aeromonas hydrophila, a type II secretion
system (T2SS)-secreted cytotoxic enterotoxin (Act) and the T3SS are crucial in the
pathogenesis of Aeromonas-associated infections. Our laboratory molecularly characterized both
Act and the T3SS from a diarrheal isolate, SSU of A. hydrophila, and defined the role of some
regulatory genes in modulating the biological effects of Act. In this study, we cloned,
sequenced, and expressed the DNA adenine methyltransferase gene of A. hydrophila SSU
(dam(AhSSU)) in a T7 promoter-based vector system using Escherichia coli ER2566 as a host
strain, which could alter the virulence potential of A. hydrophila. Recombinant Dam,
designated as M.AhySSUDam, was produced as a histidine-tagged fusion protein and purified from
an E. coli cell lysate using nickel affinity chromatography. The purified Dam had
methyltransferase activity, based on its ability to transfer a methyl group from
S-adenosyl-l-methionine to N(6)-methyladenine-free lambda DNA and to protect methylated lambda
DNA from digestion with DpnII but not against the DpnI restriction enzyme. The dam gene was
essential for the viability of the bacterium, and overproduction of Dam in A. hydrophila SSU,
using an arabinose-inducible, P(BAD) promoter-based system, reduced the virulence of this
pathogen. Specifically, overproduction of M.AhySSUDam decreased the motility of the bacterium
by 58%. Likewise, the T3SS-associated cytotoxicity, as measured by the release of lactate
dehydrogenase enzyme in murine macrophages infected with the Dam-overproducing strain, was
diminished by 55% compared to that of a control A. hydrophila SSU strain harboring the pBAD
vector alone. On the contrary, cytotoxic and hemolytic activities associated with Act as well
as the protease activity in the culture supernatant of a Dam-overproducing strain were
increased by 10-, 3-, and 2.4-fold, respectively, compared to those of the control A.
hydrophila SSU strain. The Dam-overproducing strain was not lethal to mice (100% survival)
when given by the intraperitoneal route at a dose twice that of the 50% lethal dose, which
within 2 to 3 days killed 100% of the animals inoculated with the A. hydrophila control
strain. Taken together, our data indicated alteration of A. hydrophila virulence by
overproduction of Dam.

<>

<1>Ershova, A.S., Karyagina, A.S., Vasiliev, M.O., Lyashchuk, A.M., Lunin, V.G., Spirin, S.A., Alexeevski, A.V.
<2>Solitary restriction endonucleases in prokaryotic genomes.
<3>Nucleic Acids Res.
<4>40
<5>10107-10115
<6>2012
<7>Prokaryotic restriction-modification (R-M) systems defend the host cell from the  invasion of
a foreign DNA. They comprise two enzymatic activities: specific DNA
cleavage activity and DNA methylation activity preventing cleavage. Typically,
these activities are provided by two separate enzymes: a DNA methyltransferase
(MTase) and a restriction endonuclease (RE). In the absence of a corresponding
MTase, an RE of Type II R-M system is highly toxic for the cell. Genes of the R-M
system are linked in the genome in the vast majority of annotated cases. There
are only a few reported cases in which the genes of MTase and RE from one R-M
system are not linked. Nevertheless, a few hundreds solitary RE genes are present
in the Restriction Enzyme Database (http://rebase.neb.com) annotations. Using the
comparative genomic approach, we analysed 272 solitary RE genes. For 57 solitary
RE genes we predicted corresponding MTase genes located distantly in a genome. Of
the 272 solitary RE genes, 99 are likely to be fragments of RE genes. Various
explanations for the existence of the remaining 116 solitary RE genes are also
discussed.

<>

<1>Erskine, S.G., Baldwin, G.S., Halford, S.E.
<2>Rapid-reaction analysis of plasmid DNA cleavage by the EcoRV restriction endonuclease.
<3>Biochemistry
<4>36
<5>7567-7576
<6>1997
<7>Rapid-reaction methods have been used previously to identify intermediates in the reaction of
the EcoRV restriction endonuclease on oligonucleotide substrates.  In this study, the pathway
on macromolecular DNA was elucidated by using the quench-flow method to analyze EcoRV
reactions on a plasmid with one recognition site.  Some reactions were carried out by first
allowing the EcoRV enzyme to bind nonspecifically to the DNA and then initiating DNA cleavage
by adding magnesium ions.  The subsequent transfer of the enzyme from nonspecific to specific
sites was extremely rapid, at a random walk rate of at least 5 x 10^5 base pairs per second.
The two strands of the DNA at the EcoRV recognition site were then cleaved sequentially, at
rates that were faster than the turnover number of the enzyme.  The rates recorded for the
cleavage steps were direct measurements of phosphodiester hydrolysis, while the turnover is
limited by the dissociation of the product cleaved in both strands.  Other reactions were
initiated by adding EcoRV and MgCl2 to the DNA: these are the processes observed in reactions
starting from DNA-bound enzyme but also the bimolecular association of the protein with the
plasmid.  The association rate was limited by diffusion but its rate constant, 1.2 x 10^8 M-1
s-1, was unusually small for the binding of a protein to DNA.  The slowness of this
diffusion-controlled process may be due to a rapid oscillation of the protein between closed
and open conformations, with only the open form capable of binding DNA.

<>

<1>Erskine, S.G., Halford, S.E.
<2>Fluorescent substrates for the EcoRV restriction endonuclease.
<3>Biochem. Soc. Trans.
<4>22
<5>299s
<6>1994
<7>In the presence of Mg2+, the EcoRV restriction endonuclease cleaves DNA specifically at the
sequence GAT/ATC (where / denotes the point of scission). Even the change of a single base
pair within this sequence will lead to a million fold reduction of EcoRV activity.
Paradoxically, gel retardation experiments in the absence of Mg2+ show that, unlike EcoRI and
many other type II restriction endonucleases, EcoRV binds to DNA without any sequence
preference. The specificity of EcoRV is in fact dependent on the production of a high affinity
binding site for Mg2+ between the protein and the cognate DNA. X-ray crystal structures show
that the cognate DNA adopts a highly distorted, kinked conformation in its complex with EcoRV,
in contrast to noncognate DNA which retains a B-like conformation. The difference between the
delta-G0 for the binding of cognate and noncognate sequences is near to zero and hence the
energy from the additional contacts with the specific DNA appears to be neutralized by the
unfavorable energy change from the distortion of the DNA. In addition, the crystal structures
show that the protein itself must undergo a conformation change before it can bind DNA.
Therefore, during one cycle of DNA cleavage, the EcoRV protein will undergo several
conformational changes: due first to binding nonspecific DNA and then specific DNA, and later
by the sequence of events leading to cleavage and product release.

<>

<1>Erskine, S.G., Halford, S.E.
<2>Interactions of the EcoRV restriction endonuclease with fluorescent oligodeoxynucleotides.
<3>Gene
<4>157
<5>153-156
<6>1995
<7>A self-complementary dodecadeoxyribonucleotide that contains the recognition sequence for the
R.EcoRV Enase was synthesized with a primary amino group at its 5' terminus.  The 5' amino
function was labeled with the fluorescent dye 5-[dimethylamino] napthalene-1-sulfonyl
chloride.  The labeled oligodeoxyribonucleotide in its duplex form was shown to be a suitable
substrate for kinetic studies on the ENase and that no significant dye-DNA or dye-protein
interactions occurred.  Finally, the binding of R.EcoRV to the labeled DNA was followed by
detecting the fluorescence resonance energy transfer between the tryptophans of the protein
and the fluorescent labels of the DNA.

<>

<1>Erskine, S.G., Halford, S.E.
<2>Reactions of the EcoRV restriction endonuclease with fluorescent oligodeoxynucleotides: identical equilibrium constants for binding to specific and non-specific DNA.
<3>J. Mol. Biol.
<4>275
<5>759-772
<6>1998
<7>The EcoRV restriction endonuclease cleaves DNA specifically at its recognition sequence in the
presence of magnesium ions, but several studies have indicated that it binds to DNA in the
absence of Mg2+ without any preference for its recognition site.  However, specific binding to
the recognition site has also been reported.  To distinguish between these reports,
oligodeoxynucleotides were tagged with either dansyl or eosin fluorophores at their 5'
termini and annealed to form duplexes of 12 to 16 base-pairs.  For each length of duplex, one
derivative had the EcoRV recognition sequence while another lacked this sequence.  For the
duplexes with the recognition site, the fluorophores had no effect on DNA cleavage rates by
EcoRV in the presence of Mg2+.  The binding of the specific and non-specific duplexes to EcoRV
in the absence of Mg2+ was measured by fluorescence resonance energy transfer and by
fluorescence depolarization.  In both procedures, the signal from the specific complex
differed from the complex with non-specific DNA, with the depolarization data indicating that
non-specific DNA bound to EcoRV retains a higher rotational freedom than specific DNA.  Even
so, the equilibrium constant for the binding of specific DNA was identical, within error
limits, to that for non-specific DNA.

<>

<1>Eruslanov, B.V., Kramarov, V.M., Smolyanivov, V.V., Borovik, R.V.
<2>Isolation of the site-specific endonuclease EcoRI with the aid of an immunoabsorbent.
<3>Bioorg. Khim.
<4>6
<5>1361-1369
<6>1980
<7>A simple and satisfactorily reproducible method of isolating the site-specific endonuclease
EcoRI which is based on the chromatography of the enzyme on a column filled with antibodies
immobilized on Sepharose 4B is described.  The binding of the enzyme to the antibodies is
carried out directly from a cell extract, and after the support has been washed free from
unbound protein the enzyme is eluted.  The isolation of the homogeneous enzyme (20,000
activity units per 1g of biomass) takes place in one stage and lasts 3 h.  The preparation
obtained is stable on storage in 50% glycerol at -15C for more than six months.  The paper
also describes a method of purifying the endonuclease EcoRI to the homogeneous state in two
chromatographic stages:  in columns containing phosphocellulose and Sephadex G-150.  The
enzyme obtained by this method contains no nonspecific nucleases and possesses a specific
activity of 750,000 activity units/mg of protein (3750 activity units per 1g of biomass).

<>

<1>Erwin, A.L., Gotschlich, E.C.
<2>Cloning of a Neisseria meningitidis gene for L-lactate dehydrogenase (L-LDH): evidence for a second meningococcal L-LDH with different   regulation.
<3>J. Bacteriol.
<4>178
<5>4807-4813
<6>1996
<7>We report the cloning of lldA, a Neisseria meningitidis gene for L-lactate dehydrogenase
(L-LDH). Escherichia coli contains a single L-LDH gene
(lldD) in the lld operon (previously lct). E. coli grown in complex media
does not have L-LDH activity, but the activity is induced by growth in
defined medium with L-lactate as the carbon source. In contrast,
meningococci contain at least one L-LDH in addition to the lldA gene
product. These enzymes are active in meningococci grown in complex media
and are not dependent on growth in L-lactate. The predicted amino acid
sequence of lldA is homologous to that of E. coli lldD and of other
prokaryotic and eukaryotic flavin mononucleotide-containing enzymes that
catalyze the oxidation of L-lactate and other small alpha-hydroxy acids. A
mutant with a deletion in lldA was found to have reduced L-LDH activity.
However, this mutant was able to grow on L-lactate, indicating that a
second L-LDH must exist. Activity of the lldA enzyme was affected by
growth conditions, being increased by growth on a defined medium with
either L-lactate or pyruvate as the carbon source. For meningococci grown
on a complex medium, activity of the lldA enzyme was increased by growth
on plates or in well-aerated broth. A second L-lactate-oxidizing activity
was seen in bacteria grown in poorly aerated broth. Neisseria gonorrhoeae
contains a homolog of lldA. As for meningococci, mutation of the
gonococcal lldA reduced L-LDH activity but did not affect growth on
L-lactate.

<>

<1>Erwin, A.L., Sandstedt, S.A., Bonthuis, P.J., Geelhood, J.L., Nelson, K.L., Unrath, W.C., Diggle, M.A., Theodore, M.J., Pleatman, C.R., Mothershed, E.A., Sacchi, C.T., Mayer, L.W., Gilsdorf, J.R., Smith, A.L.
<2>Analysis of genetic relatedness of Haemophilus influenzae isolates by multilocus sequence typing.
<3>J. Bacteriol.
<4>190
<5>1473-1483
<6>2008
<7>The gram-negative bacterium Haemophilus influenzae is a human-restricted
commensal of the nasopharynx that can also be associated with disease. The
majority of H. influenzae respiratory isolates lack the genes for capsule
production and are nontypeable (NTHI). Whereas encapsulated strains are
known to belong to serotype-specific phylogenetic groups, the structure of
the NTHI population has not been previously described. A total of 656 H.
influenzae strains, including 322 NTHI strains, have been typed by
multilocus sequence typing and found to have 359 sequence types (ST). We
performed maximum-parsimony analysis of the 359 sequences and calculated
the majority-rule consensus of 4,545 resulting equally most parsimonious
trees. Eleven clades were identified, consisting of six or more ST on a
branch that was present in 100% of trees. Two additional clades were
defined by branches present in 91% and 82% of trees, respectively. Of
these 13 clades, 8 consisted predominantly of NTHI strains, three were
serotype specific, and 2 contained distinct NTHI-specific and
serotype-specific clusters of strains. Sixty percent of NTHI strains have
ST within one of the 13 clades, and eBURST analysis identified an
additional phylogenetic group that contained 20% of NTHI strains. There
was concordant clustering of certain metabolic reactions and putative
virulence loci but not of disease source or geographic origin. We conclude
that well-defined phylogenetic groups of NTHI strains exist and that these
groups differ in genetic content. These observations will provide a
framework for further study of the effect of genetic diversity on the
interaction of NTHI with the host.

<>

<1>Erxleben, A., Wunsch-Palasis, J., Gruning, B.A., Luzhetska, M., Bechthold, A., Gunther, S.
<2>Genome Sequence of Streptomyces sp. Strain Tu6071.
<3>J. Bacteriol.
<4>193
<5>4278-4279
<6>2011
<7>Streptomyces sp. Tu6071 is a soil-dwelling bacterium which has a highly active isoprenoid
biosynthesis. Isoprenoids are important precursors for
biopharmaceutical molecules such as antibiotics or anticancer agents,
e.g., landomycin. Streptomyces sp. Tu6071 produces the industrially
important terpene glycosides phenalinolactones, which have antibacterial
activity against several Gram-positive bacteria. The availability of the
genome sequence of Streptomyces sp. Tu6071 allows for understanding the
biosynthesis of these pharmaceutical molecules and will facilitate
rational genome modification to improve industrial use.

<>

<1>Eskes, R., Liu, L., Ma, H., Chao, M.Y., Dickson, L., Lambowitz, A.M., Perlman, P.S.
<2>Multiple homing pathways used by yeast mitochondrial group II introns.
<3>Mol. Cell. Biol.
<4>20
<5>8432-8446
<6>2000
<7>The yeast mitochondrial DNA group II introns aI1 and aI2 are retroelements that insert site
specifically into intronless alleles by a process called homing. Here, we used patterns of
flanking marker coconversion in crosses with wild-type and mutant aI2 introns to distinguish
three coexisting homing pathways: two that were reverse transcriptase (RT) dependent
(retrohoming) and one that was RT independent. All three pathways are initiated by cleavage of
the recipient DNA target site by the intron-encoded endonuclease, with the sense strand
cleaved by partial or complete reverse splicing, and the antisense strand cleaved by the
intron-encoded protein.  The major retrohoming pathway in standard crosses leads to insertion
of the intron with unidirectional coconversion of upstream exon sequences. This pattern of
coconversion suggests that the major retrohoming pathway is initiated by target DNA-primed
reverse transcription of the reverse-spliced intron RNA and completed by double-strand break
repair (DSBR) recombination with the donor allele. The RT-independent pathway leads to
insertion of the intron with bi-directional coconversion and presumably occurs by a
conventional DSBR recombination mechanism initiated by cleavage of the recipient DNA target
site by the intron-encoded endonuclease, as for group I intron homing. Finally, some mutant
DNA target sites shift up to 43% of retrohoming to another pathway not previously detected for
aI2 in which there is no coconversion of flanking exon sequences. This new pathway presumably
involves synthesis of a full-length cDNA copy of the inserted intron RNA, with completion by a
repair process independent of homologous recombination, as found for the Lactococcus lactis
Ll.LtrB intron. Our results show that group II intron mobility can occur by multiple pathways,
the ratios of which depend on the characteristics of both the intron and the DNA target site.
This remarkable flexibility enables group II introns to use different recombination and repair
enzymes in different host cells.

<>

<1>Eskes, R., Yang, J., Lambowitz, A.M., Perlman, P.S.
<2>Mobility of yeast mitochondrial group II introns: Engineering a new site specificity and retrohoming via full reverse splicing.
<3>Cell
<4>88
<5>865-874
<6>1997
<7>The mobile group II introns aI1 and aI2 of yeast mtDNA encode endonuclease activities that
cleave intronless DNA target sites to initiate mobility by target DNA-primed reverse
transcription.  For aI2, sense-strand cleavage occurs mainly by a partial reverse splicing
reaction, whereas for aI1, complete reverse splicing occurs, leading to insertion of the
linear intron RNA into double-stranded DNA.  Here, we show that aI1 homing and reverse
splicing depend on the EBS1 (RNA)/IBS1(DNA) pairing and that target specificity can be changed
by compensatory changes in the target site and the donor intron.  Using well-marked strains to
follow coconversion of flanking DNA, we show that homing occurs by both RT-dependent and
-independent pathways.  Remarkably, in most RT-dependent events, the reverse spliced intron is
the initial template for first-strand cDNA synthesis.

<>

<1>Eskin, B.
<2>The host-controlled restriction enzyme of Escherichia coli B.
<3>Ph.D. Thesis, University of California, Berkeley, USA
<4>
<5>1-191
<6>1973
<7>The restriction endonuclease of Escherichia coli B has been
purified and is free of non-specific endonuclease.  On sucrose gradients it
sediments in a broad band with an S20,w of 11 through 18.  As judged by
polyacrylamide gel electrophoresis the enzyme exists in at leas two active
forms, each of which possesses three nonidentical polypeptides, alpha,
beta, and gamma, of molecular weights 135,000, 60,000 and 55,000,
respectively.  The molecular weights of these two forms have been estimated
to be around 200,000 and 700,000.  Combining this information with
estimates of the ratios of subunits present in each form suggests that the
molecular formulae of the two forms are alpha1beta1gamma1 and
alpha2beta4gamma2, respectively.  The subunits, beta and gamma, are
indistinguishable by polyacrylamide gel electrophoresis from the two
subunits found in the modification methylase of Escherichia coli B.

<>

<1>Eskin, B., Lautenberger, J.A., Linn, S.
<2>Host-controlled modification and restriction of bacteriophage T7 by Escherichia coli B.
<3>J. Virol.
<4>11
<5>1020-1023
<6>1973
<7>T7 phage resists Escherichia coli B host-controlled modification and
restriction in vivo, but its DNA carries roughly five sites which are
susceptible to the purified enzymes.

<>

<1>Eskin, B., Linn, S.
<2>The deoxyribonucleic acid modification and restriction enzymes of Escherichia coli B. III.  Studies of the restriction adenosine triphosphatase.
<3>J. Biol. Chem.
<4>247
<5>6192-6196
<6>1972
<7>The restriction endonuclease of Escherichia coli B catalyzes a massive
hydrolysis of ATP to ADP and Pi. The ATPase requires S-adenosylmethionine and
DNA containing unmodified restriction sites.  The apparent Km for fd
replicative form DNA is 20 microM DNA nucleotide, and ATP is half-saturating at
100 microM.  Like the nuclease, the ATPase is inhibited strongly by
S-adenosylethionine and 5'-methylthioadenosine, but only weakly by
S-adenosylhomocysteine.  The hydrolysis of ATP continues long after DNA
degradation has ceased.  Whereas no ATPase is observed when restricted DNA but
no unmodified DNA is present in a reaction mixture, restricted DNA is required
for the maintenance of ATPase once initiated.  A hypothetical scheme is
presented which involves the conversion of the enzyme during DNA hydrolysis
from a form capable of nuclease activity to one catalyzing the breakdown of
ATP.

<>

<1>Eskin, B., Linn, S.
<2>The deoxyribonucleic acid modification and restriction enzymes of Escherichia coli B. II. Purification, subunit structure, and catalytic properties of the restriction endonuclease.
<3>J. Biol. Chem.
<4>247
<5>6183-6191
<6>1972
<7>The restriction endonuclease of Escherichia coli B has been purified and is
free of nonspecific endonuclease.  On sucrose gradients it sediments in a broad
band with an S20,w of 11 through 18.  As judged by polyacrylamide gel
electrophoresis the enzyme exists in at least two active forms, each of which
possess three nonidentical polypeptides, a, b, and c, of molecular weights
135,000, 60,000 and 55,000, respectively.  The subunits, b, and c, are
indistinguishable by polyacrylamide gel electrophoresis from the two subunits
found in the modification methylase of E. coli B.  ATP is half-saturating at 80
to 100 lM, S-adenosylmethionine has an apparent Km of 0.3 to 0.4 microM, and fd
replicative form DNA is saturating at greater than 10 to 20 microM
DNA-nucleotide.  S-adenosylethionine and 5'-methylthioadenosine, but not
S-adenosylhomocysteine are potent inhibitors of the enzyme.  Modified DNA
inhibits by 50% when added in a 5-fold excess over unmodified substrate.  The
restriction nuclease activity ceases after 5 or 10 min, and the enzyme does not
appear to turn over in the nuclease reaction.  The cleaved DNA product is not
adenylylated, phosphorylated, or methylated during hydrolysis.  It is
susceptible to lambda-exonuclease, exonuclease III, the recBC nuclease, and,
after denaturation, exonuclease I.  These results imply that the termini of the
restricted DNA have hydroxyl and 5'-phosphoryl groups, and that they have a
duplex structure.  However, restricted DNA cannot be phosphorylated by
polynucleotide kinase, even after treatment of the DNA with alkaline
phosphatase.  Finally, in order to make the enzyme more accessible, relatively
rapid assay procedures are suggested which are based on the ATPase activity of
the enzyme, or on the rendering of circular DNA to a linear form which is
susceptible to the recBC nuclease.

<>

<1>Eskridge, R.W., Weinfeld, H., Paigen, K.
<2>Susceptibility of different coliphage genomes to host-controlled variation.
<3>J. Bacteriol.
<4>93
<5>835-844
<6>1967
<7>Twenty-eight coliphages were studied for their susceptibility to four systems
of host control variation in Escherichia coli.  Both temperate and virulent
phages were studied, including phages with ribonucleic acid, double- and
single-stranded deoxyribonucleic acid (DNA) and glucosylated DNA.  The systems
examined were E. coli C-K, K-B, B-K, and K-K(P1).  The C-K, K-B, and B-K
systems affected temperate phages and nonlysogenizing mutants derived from
temperate phages.  In general, these systems did not restrict virulent phages.
Phage 21e, a variant of phage 21, lost the ability to undergo restriction in
the C-K and B-K systems, but retained susceptibility to the K-B and K-K(P(1)
systems.  This suggests that the genetic site(s) on the phage, as well as in
the host, determines susceptibility to host-controlled variation.  Both
temperate and dependent virulent phages were susceptible to the host control
system resulting from the presence of prophage P1.  The autonomous and small
virulents were not susceptible.  In a given system, the various susceptible
phages differed widely in their efficiency of plating on the restricting host.
If the few infections that occur arise in rare special cells, then different
populations of special cells are available to different phage species.  For
most phage types, when a susceptible phage infected a nonrestricting host, the
progeny showed the specificity appropriate to that host. Behavior of T3 was
exceptional, however,  When T3 obtained from E. coli K infected E. coli C or B,
some of the progeny phages retained K host specificity, whereas others acquired
the specificity of the new host.

<>

<1>Espada, J., Ballestar, E., Fraga, M.F., Garea, A.V., Juarranz, A., Stockert, J.C., Robertson, K.D., Fuks, F.O., Esteller, M.
<2>Human DNA methyltransferase 1 is required for maintenance of the histone H3 modification pattern.
<3>J. Biol. Chem.
<4>279
<5>37175-37184
<6>2004
<7>DNA methyltransferase 1 (DNMT1) plays an essential role in murine development and is thought
to be the enzyme primarily responsible for
maintenance of the global methylation status of genomic DNA. However,
loss of DNMT1 in human cancer cells affects only the methylation status
of a limited number of pericentromeric sequences. Here we show that
human cancer cells lacking DNMT1 display at least two important
differences with respect to wild type cells: a profound disorganization
of nuclear architecture, and an altered pattern of histone H3
modification that results in an increase in the acetylation and a
decrease in the dimethylation and trimethylation of lysine 9.
Additionally, this phenotype is associated with a loss of interaction
of histone deacetylases (HDACs) and HP1 (heterochromatin protein 1)
with histone H3 and pericentromeric repetitive sequences (satellite 2).
Our data indicate that DNMT1 activity, via maintenance of the
appropriate histone H3 modifications, contributes to the preservation
of the correct organization of large heterochromatic regions.

<>

<1>Espedido, B.A., Steen, J.A., Barbagiannakos, T., Mercer, J., Paterson, D.L., Grimmond, S.M., Cooper, M.A., Gosbell, I.B., van Hal, S.J., Jensen, S.O.
<2>Carriage of an ACME II Variant may have contributed to MRSA ST239-like Strain Replacement in Liverpool Hospital, Sydney, Australia.
<3>Antimicrob. Agents Chemother.
<4>56
<5>3380-3383
<6>2012
<7>Approximately 39% of MRSA ST239-like bloodstream isolates from Liverpool Hospital
(1997-2008) carry an arginine catabolic mobile element (ACME). Whole genome
sequencing revealed that an ACME II variant is located between orfX and SCCmec
III, and based on pulsed-field gel electrophoresis patterns and temporal
relationships of all ST239-like isolates (n=360), ACME carriage may have
contributed to sub-pulsotype strain replacement.

<>

<1>Esperito-Santo, C., Lin, Y., Hao, X., Wei, G., Rensing, C., Grass, G.
<2>Draft Genome Sequence of Pseudomonas psychrotolerans L19, Isolated from Copper Alloy Coins.
<3>J. Bacteriol.
<4>194
<5>1623-1624
<6>2012
<7>We report the draft genome sequence of Pseudomonas psychrotolerans strain L19, isolated from a
European 50-cent copper alloy coin. Multiple genes potentially involved in copper resistance
were identified; however, it is unknown if these copper ion resistance determinants contribute
to prolonged survival of this strain on dry metallic copper.

<>

<1>Espinoza-Miranda, S.S., Gomez-Rodriguez, J.A., Huete-Perez, J.A.
<2>Mining for restriction endonucleases in Nicaragua.
<3>Encuentro
<4>93
<5>49-62
<6>2012
<7>The Molecular Biology Center at the University of Central America in Nicaragua (CBM-UCA) was
founded in 1999 to strengthen biotechnology research capacity and education in Nicaragua and
the Central American region.  One of the first projects launched by the CBM-UCA was
bio-prospecting for key industrial enzymes.  This ongoing study seeks to discover and
characterize restriction enzymes (RE) in bacteria, and to create a database of microorganisms
isolated and identified by 16S rDNA sequencing methodology.  In this paper we highlight the
importance of studying the extreme environmental conditions for building knowledge of
Nicaraguan biodiversity through modern molecular biology techniques such as metagenomics.  The
isolation of prototype enzymes such as EcoRV and ClaI is presented as an update and extension
of previously undertaken work.

<>

<1>Espinoza-Valles, I., Soto-Rodriguez, S., Edwards, R.A., Wang, Z., Vora, G.J., Gomez-Gil, B.
<2>Draft Genome Sequence of the Shrimp Pathogen Vibrio harveyi CAIM 1792.
<3>J. Bacteriol.
<4>194
<5>2104
<6>2012
<7>Vibrio harveyi is a Gram-negative bacterium found in tropical and temperate marine
environments as a free-living organism or in association with aquatic
animals. We report the first sequenced genome of a Vibrio harveyi strain, CAIM
1792, the etiologic agent of the 'bright red' syndrome of the Pacific white
shrimp Litopenaeus vannamei.

<>

<1>Esposito, D., Fitzmaurice, W.P., Benjamin, R.C., Goodman, S.D., Waldman, A.S., Socca, J.J.
<2>The complete nucleotide sequence of bacteriophage HP1 DNA.
<3>Nucleic Acids Res.
<4>24
<5>2360-2368
<6>1996
<7>The complete nucleotide sequence of the temperate phage HP1 of Haemophilus influenzae was
determined. The phage contains a linear, double-stranded genome of 32 355 nt with cohesive
termini. Statistical methods were used to identify 41 probable protein coding segments
organized into five plausible transcriptional units. Regions encoding proteins involved in
recombination, replication, transcriptional control, host cell lysis and phage production were
identified. The sizes of proteins in the mature HP1 particle were determined to assist in
identifying genes for structural proteins. Similarities between HP1 coding sequences and those
in databases, as well as similar gene organizations and control mechanisms, suggest that HP1
is a member of the P2-like phage family, with strong similarities to coliphages P2 and 186 and
some similarity to the retronphage Ec67.

<>

<1>Essani, K., Goorha, R., Granoff, A.
<2>An animal virus-induced DNA-methyltransferase.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>222
<6>1989
<7>The DNA genome of frog virus 3 (FV3), an iridovirus, is highly methylated; more
than 20% of cytosine residues are methylated at the 5-carbon position.
Methylation of the viral DNA occurs in the cytoplasm of infected cells by an
FV3-specified DNA-methyltransferase (DNA-mt).  To determine the role of this
enzyme in virus replication and gene expression we have isolated a number of
FV3 mutants defective in the expression of DNA-mt activity.  Combined genetic
and biochemical analyses of one of the mutants have revealed a 26K polypeptide
associated with DNA-mt activity.  Attempts to purify the 26K polypeptide have
resulted in co-purification of two other polypeptides.  30K and 18K, along with
the 26K one.  Additional experiments directed toward identifying DNA-mt
activity with the individual polypeptides, together with reconstitution
experiments, have indicated that at least two polypeptides (26K and 18K) are
required for functional DNA-mt activity.  These data support the conclusion
that FV3-induced DNA-mt, unlike any known eukaryotic DNA-mt, resides in a
complex of at least two polypeptides.

<>

<1>Essani, K., Goorha, R., Granoff, A.
<2>Mutation in a DNA-binding protein reveals an association between DNA-methyltransferase activity and a 26,000-Da polypeptide in frog virus 3-infected cells.
<3>Virology
<4>161
<5>211-217
<6>1987
<7>The DNA of frog virus 3, an iridovirus, is highly methylated; more than 20% of the cytosine
bases are methylated at the 5-carbon position by an FV3-induced DNA methyltransferase.  To
determine the role of this enzyme in virus replication and regulation of gene expression, we
have analyzed an FV3 mutant that lacks DNA-mt activity and is resistant to 5-azacytidine (an
inhibitor of DNA-mt).  Comparative polypeptide analysis using cytoplasmic extracts from the
wild-type FV3 and mutant-infected cells, revealed that a single protein of 26,000 molecular
weight was altered in the mutant-infected cells.  The altered polypeptide migrated faster in
SDS-polyacrylamide gel as compared to the wild-type FV3 26K protein.  Five spontaneous
revertants derived from the mutant regained the migrational characteristic of the wild-type
26K protein, DNA-mt activity, and methylation of their DNA.  We further show that the 26K
polypeptide is a DNA-binding protein and that 80% of the enzyme activity can be eluted from an
ssDNA affinity column.  Taken together, these data support the conclusion that the 26K
polypeptide is associated with DNA-mt activity.

<>

<1>Estabrook, R.A., Lipson, R., Hopkins, B., Reich, N.
<2>The coupling of tight DNA binding and base flipping: identification of a conserved structural motif in base flipping enzymes.
<3>J. Biol. Chem.
<4>279
<5>31419-31428
<6>2004
<7>Val(121) is positioned immediately above the extrahelical cytosine in HhaI DNA C(5)-cytosine
methyltransferase, and replacement with alanine
dramatically interferes with base flipping and catalysis. DNA binding and
k(cat) are decreased 10^5-fold for the Val(121) --> Ala mutant that has a
normal circular dichroism spectrum and AdoMet affinity. The magnitude of
this loss of function is comparable with removal of the essential
catalytic Cys(81). Surprisingly, DNA binding is completely recovered
(increase of 10^5-fold) with a DNA substrate lacking the target cytosine
base (abasic). Thus, interfering with the base flipping transition results
in a dramatic loss of binding energy. Our data support an induced fit
mechanism in which tight DNA binding is coupled to both base flipping and
protein loop rearrangement. The importance of the proximal protein segment
(His(127)-Thr(132)) in maintaining this critical interaction between
Val(121) and the flipped cytosine was probed with single site alanine
substitutions. None of these mutants are significantly altered in
secondary structure, AdoMet or DNA affinity, k(methylation),
k(inactivation), or k(cat). Although Val(121) plays a critical role in
both extrahelical base stabilization and catalysis, its position and
mobility are not influenced by individual residues in the adjacent peptide
region. Structural comparisons with other DNA methyltransferases and DNA
repair enzymes that stabilize extrahelical nucleotides reveal a motif that
includes a positively charged or polar side chain and a hydrophobic
residue positioned adjacent to the target DNA base and either the 5'- or
3'-phosphate.

<>

<1>Estabrook, R.A., Reich, N.
<2>Observing an induced-fit mechanism during sequence-specific DNA methylation.
<3>J. Biol. Chem.
<4>281
<5>37205-37214
<6>2006
<7>The characterization of conformational changes that drive induced-fit mechanisms and their
quantitative importance to enzyme specificity are
essential for a full understanding of enzyme function. Here, we report on
M.HhaI, a sequence-specific DNA cytosine C(5) methyltransferase that
reorganizes a flexible loop (residues 80-100) upon binding cognate DNA as
part of an induced-fit mechanism. To directly observe this approximately
26A conformational rearrangement and provide a basis for understanding its
importance to specificity, we replaced loop residues Lys-91 and Glu-94
with tryptophans. The double mutants W41F/K91W and W41F/E94W are
relatively unperturbed in kinetic and thermodynamic properties. W41F/E94W
shows DNA sequence-dependent changes in fluorescence: significant changes
in equilibrium and transient state fluorescence that occur when the enzyme
binds cognate DNA are absent with nonspecific DNA. These real-time,
solution-based results provide direct evidence that binding to cognate DNA
induces loop reorganization into the closed conformer, resulting in the
correct assembly of the active site. We propose that M.HhaI scans
nonspecific DNA in the loop-open conformer and rearranges to the closed
form once the cognate site is recognized. The fluorescence data exclude
mechanisms in which loop motion precedes base flipping, and we show loop
rearrangements are directly coupled to base flipping, because the
sequential removal of single hydrogen bonds within the target
guanosine:cytosine base pair results in corresponding changes in loop
motion.

<>

<1>Estrada-Acosta, M., Medrano-Felix, A., Jimenez, M., Gomez-Gil, B., Leon-Felix, J., Amarillas, L., Chaidez, C.
<2>Draft Genome Sequence of Salmonella enterica subsp. enterica Serotype Saintpaul Strain S-70, Isolated from an Aquatic Environment.
<3>Genome Announcements
<4>1
<5>e01016-13
<6>2013
<7>Salmonella is a pathogen of worldwide importance, causing disease in a vast range of hosts,
including humans. We report the genome sequence of Salmonella enterica
subsp. enterica serotype Saintpaul strain S-70, isolated from an aquatic
environment.

<>

<1>Etson, C.M., Todorov, P., Walt, D.R.
<2>Elucidating Restriction Endonucleases Reaction Mechanisms via Dwell-Time Distribution Analysis.
<3>Biophys. J.
<4>106
<5>22A
<6>2014
<7>We have developed a Total Internal Reflection Fluorescence microscopy based assay that allows
us to simultaneously measure the length of the catalytic cycle for hundreds of restriction
endonuclease molecules in one experiment.  We stably attach thousands of short duplex DNA
molecules, each labeled with a single quantum dot semiconductor nanocrystal, to a passivated
glass surface within a flow channel.  The disappearance of a quantum dot indicates that its
DNA tether has been cleaved.  We introduce restriction endonuclease molecules into the channel
in the absence of magnesium, which permits binding to, but not cleavage of the surface
immobilized DNA substrate.  When buffer containing magnesium is introduced into the flow
channel, DNA cleavage by the pre-bound restriction endonuclease molecules is initiated.  This
synchronization allows us to measure the lag time between the introduction of magnesium and
the completion of DNA cleavage for the entire population of enzymes.  Analysis of the
dwell-time distributions can provide insights into the DNA cleavage mechanism.  Our
observations suggest that EcoRV, a dimeric Type II restriction endonuclease that cleaves the
palindromic sequence GAT/ATCF (where / is the cut site), requires two kinetic steps to
complete duplex cleavage after prebinding.  However, dwell-time distributions suggest that
BcnI, which is active as a monomer and cleaves the pseudopalindromic sequence 5'-CC/SGG-5'
(where S stands for C or G), requires more than four kinetic steps to complete duplex
cleavage.  Furthermore, experiments performed with strand-specific DNA substrates suggest that
the number of steps indicated by the dwell-time distribution depends on which strand of the
recognition site must be cleaved to result in quantum dot release.  By designing additional
substrates that mimic the various intermediate states, we plan to dissect the mechanism by
which BcnI cleaves each strand of the intact restriction site.

<>

<1>Etson, C.M., Wilburn, F., Moody, T., Fashakin, V., Walt, D.R.
<2>Single-Molecule Studies of Restriction Endonuclease Kinetics.
<3>Biophys. J.
<4>102
<5>486A
<6>2012
<7>Under optimal conditions, restriction endonucleases are capable of mediating remarkably
specific DNA cleavage. This quality makes the restriction endonuclease an indispensible tool
for genetic modification and manipulation.  However, the mechanism by which restriction
endonucleases effectively discriminate between their cognate site and other DNA sequences is
not fully understood. Under certain conditions, many restriction endonucleases display 'star
activity' - relaxed specificity resulting in DNA cleavage at sequences that differ from their
normal recognition sequence - but the mechanism by which specificity is relaxed is not fully
understood. Although at least 600 of the almost 4000 restriction endonucleases that have been
identified are commercially available in purified form, DNA cleavage kinetics of only a few of
these enzymes have been studied in detail. We have developed a fluorescence-based approach
with which we can track the progress of the cleavage reaction in real time, and simultaneously
determine the values of the kinetic constants for a particular restriction endonuclease at a
specific sequence. Modeling restriction endonuclease-mediated DNA cleavage as a
Michaelis-Menten-like process, we expected reaction rates to display a hyperbolic dependence
on substrate concentration, but our measurements deviate from this dependence, especially
under conditions associated with increased star activity (such as low ionic strength). These
observations suggest that substrate inhibition may be a part of the reaction mechanism under
normal conditions, and that star activity may be a result of an increase in the population of
this pathway. Using high density arrays of femtoliter-sized reaction
vessels created by selectively etching bundled optical fibers, we can
observe the cleavage activity of hundreds of individual restriction endonuclease molecules in
solution. By characterizing the population distribution of single-enzyme turnover rates under
a variety of conditions, we hope to gain insight into the reaction mechanisms of both specific
cleavage and star activity.

<>

<1>Etzkorn, C., Horton, N.C.
<2>Mechanistic insights from the structures of HincII bound to cognate DNA cleaved from addition of Mg2+ and Mn2+.
<3>J. Mol. Biol.
<4>343
<5>833-849
<6>2004
<7>The three-dimensional X-ray crystal structures of HincII bound to cognate DNA containing
GTCGAC and Mn2+ or Mg2+, at 2.50 Angstrom and
2.95 Angstrom resolution, respectively, are presented. In both
structures, the DNA is found cleaved, and the positions of the
active-site groups, cleaved phosphate group, and 3' oxygen atom of the
leaving group are in very similar positions. Two highly occupied Mn2+
positions are found in each active site of the four
crystallographically independent subunit copies in the HincII/DNA/Mn2+
structure. The manganese ion closest to the previously identified
single Ca2+ position of HincII is shifted 1.7 Angstrom and has lost
direct ligation to the active-site aspartate residue, Asp127. A
Mn2+-ligated water molecule in a position analogous to that seen in the
HincII/DNA/Ca2+ structure, and proposed to be the attacking
nucleophile, is beyond hydrogen bonding distance from the active-site
lysine residue, Lys129, but remains within hydrogen bonding distance
from the proRp oxygen atom of the phosphate group 3' to the scissile
phosphate group. In addition, the position of the cleaved phosphate
group is on the opposite side of the axis connecting the two metal ions
relative to that found in the BamHI/product DNA/Mn2+ structure.
Mechanistic implications are discussed, and a model for the
two-metal-ion mechanism of DNA cleavage by HincII is proposed.

<>

<1>Etzkorn, C., Horton, N.C.
<2>Ca2+ binding in the active site of HincII: implications for the catalytic mechanism.
<3>Biochemistry
<4>43
<5>13256-13270
<6>2004
<7>The 2.8 A crystal structure of the type II restriction endonuclease HincII bound to Ca(2+) and
cognate DNA containing GTCGAC is presented. The DNA is
uncleaved, and one calcium ion is bound per active site, in a position
previously described as site I in the related blunt cutting type II
restriction endonuclease EcoRV [Horton, N. C., Newberry, K. J., and
Perona, J. J. (1998) Proc. Natl. Acad. Sci. U.S.A. 95 (23), 13489-13494],
as well as that found in other related enzymes. Unlike the site I metal in
EcoRV, but similar to that of PvuII, NgoMIV, BamHI, BglII, and BglI, the
observed calcium cation is directly ligated to the pro-S(p) oxygen of the
scissile phosphate. A calcium ion-ligated water molecule is well
positioned to act as the nucleophile in the phosphodiester bond cleavage
reaction, and is within hydrogen bonding distance of the conserved active
site lysine (Lys 129), as well as the pro-R(p) oxygen of the phosphate
group 3' of the scissile phosphate, suggesting possible roles for these
groups in the catalytic mechanism. Kinetic data consistent with an
important role for the 3'-phosphate group in DNA cleavage by HincII are
presented. The previously observed sodium ion [Horton, N. C., Dorner, L.
F., and Perona, J. J. (2002) Nat. Struct. Biol. 9, 42-47] persists in the
active sites of the Ca(2+)-bound structure; however, kinetic data show
little effect on the single-turnover rate of DNA cleavage in the absence
of Na(+) ions.

<>

<1>Euler, C.W., Ryan, P.A., Martin, J.M., Fischetti, V.A.
<2>M.SpyI, a DNA methyltransferase encoded on a mefA chimeric element, modifies the genome of Streptococcus pyogenes.
<3>J. Bacteriol.
<4>189
<5>1044-1054
<6>2007
<7>While screening the clonality of Streptococcus pyogenes isolates from an outbreak of
erythromycin-resistant pharyngitis in Pittsburgh, PA, we found
a correlation between the presence of the chimeric element Phi10394.4
(carrying the macrolide efflux gene, mefA) and genomic DNA being resistant
to cleavage by SmaI restriction endonuclease. A search of the open reading
frames in Phi10394.4 identified a putative type II
restriction-modification (R-M) cassette containing a cytosine
methyltransferase gene (spyIM). Heterologous expression of the cloned
spyIM gene, as well as allelic-replacement experiments, showed that the
action of this methyltransferase (M.SpyI) was responsible for the
inhibition of SmaI digestion of genomic DNA in the Phi10394.4-containing
isolates. Analysis of the methylation patterns of streptococcal genomic
DNA from spyIM-positive strains, a spyIM deletion mutant, and a
spyIM-negative strain determined that M.SpyI specifically recognized and
methylated the DNA sequence to generate 5'-C(m)CNGG. To our knowledge,
this is the first methyltransferase gene from S. pyogenes to be cloned and
to have its activity characterized. These results reveal why pulsed field
gel electrophoresis analysis of SmaI-digested genomic DNA cannot be used
to analyze the clonality of some streptococci containing Phi10394.4 and
may explain the inability of previous epidemiological studies to use SmaI
to analyze DNAs from macrolide-resistant streptococci. The presence of the
SpyI R-M cassette in Phi10394.4 could impart a selective advantage to host
strain survival and may provide another explanation for the observed
increase in macrolide-resistant streptococci.

<>

<1>Evans, D.A., Bronowska, A.K.
<2>Implications of fast-time scale dynamics of human DNA/RNA cytosine methyltransferases (DNMTs) for protein function.
<3>Theor. Chem. Acc.
<4>125
<5>407-418
<6>2010
<7>The role of protein dynamics in the control of substrate recognition, catalysis, and
protein-protein interactions is often underestimated. Recently, a number of studies have
examined the contribution of protein dynamics to the thermodynamics of ligand binding in
detail,
mostly using NMR relaxation measurements and molecular dynamics (MD) simulations. The results
unequivocally demonstrate that conformational dynamics play a pivotal role in the properties
and functions of proteins, and
ignoring this contribution is likely to lead to substantial errors when explaining the
biological function of proteins and in predictions of the binding affinities of their cognate
ligands. However, the details of the interplay between structure and dynamics and the way it
affects the biological function of the target protein remain poorly understood. In
this study, the changes in fast (picosecond-to-nanosecond time scale) dynamics of catalytic
domains of four human cytosine DNA methyltransferases (DNMTs) were studied using molecular
dynamics (MD) simulations. The results
provide insight into the protein dynamics changes that occur upon binding of the cofactor,
S-adenosylmethionine (SAM). Contrary to expectations, increased amplitude of motions of
backbone amide (N-H) and terminal heavy
atom (C-C) bond vectors was observed in all studied DNMTs upon binding of SAM. These results
imply that the cofactor binding causes a global increase in the extent of protein dynamics in
the short time scale. This global
dynamic change constitutes a favourable entropic contribution to the free energy of SAM
binding. These results suggest that cytosine DNA methyltransferases may exploit changes in
their fast scale dynamics to reduce the entropic cost of the substrate binding.

<>

<1>Evans, M., Kaczmarek, F.S., Stutzman-Engwall, K., Dyson, P.
<2>Characterization of the Streptomyces-lividans-type site-specific DNA modification system in the avermectin-producer Streptomyces avermitilis permits investigation of two novel giant linear plasmids, pSA1 and pSA2.
<3>Microbiology
<4>140
<5>1367-1371
<6>1994
<7>The degradation of Streptomyces avermitilis DNA samples analysed by conventional pulsed-field
gel electrophoresis was shown to be due to Tris-dependent, double-strand cleavage.  Using
alternative electrophoretic conditions, separation of intact DNA molecules was achieved,
permitting the identification of two novel giant linear plasmids: the 100 kb pSA1 and 250 kb
pSA2.  Use of pSA2 DNA as a probe showed that pSA1 does not cross-hybridize, indicating that
the plasmids are not closely related.  The site-specificity of the DNA modifications, which
render the DNA susceptible to Tris-dependent cleavage, was found to be essentially identical
to that of similar modifications found in the DNA of S. lividans.

<>

<1>Evans, P.S., Luo, Y., Muruvanda, T., Ayers, S., Hiatt, B., Hoffman, M., Zhao, S., Allard, M.W., Brown, E.W.
<2>Complete Genome Sequences of Salmonella enterica Serovar Heidelberg Strains Associated with a Multistate Food-Borne Illness Investigation.
<3>Genome Announcements
<4>2
<5>e01154-13
<6>2014
<7>Next-generation sequencing is being evaluated for use with food-borne illness investigations,
especially when the outbreak strains produce patterns that cannot
be discriminated from non-outbreak strains using conventional procedures. Here we
report complete genome assemblies of two Salmonella enterica serovar Heidelberg
strains with a common pulsed-field gel electrophoresis pattern isolated during an
outbreak investigation.

<>

<1>Evans, T.C. Jr., Benner, J., Xu, M.Q.
<2>Semisynthesis of cytotoxic proteins using a modified protein splicing element.
<3>Protein Sci.
<4>7
<5>2256-2264
<6>1998
<7>Two cytotoxic proteins, bovine pancreatic ribonuclease A (RNase A), and a restriction
endonuclease from Haemophilus parainfluenzae (HpaI), were produced using a novel semisynthetic
approach that utilizes a protein splicing element, an intein, to generate a reactive thioester
at the C-terminus of a recombinant protein. Nucleophilic attack on this thioester by the
N-terminal cysteine of a synthetic peptide ultimately leads to the ligation of the two
reactants through a native peptide bond. This strategy was used to produce RNase A and HpaI by
isolating inactive truncated forms of these proteins, the first 109 and 223 amino acids of
RNase A and HpaI, respectively, as fusion proteins consisting of the target protein, an
intein, and a chitin binding domain. Thiol-induced cleavage of the precursor led to the
liberation of the target protein with a C-terminal thioester-tag. Addition of synthetic
peptides representing the amino acids missing from the truncated forms led to the generation
of full-length products that displayed catalytic activity indicative of the wild-type enzymes.
The turnover numbers and Km for ligated and renatured RNase A were 8.2 s(-1) and 1.5 mM, in
good agreement with reported values of 8.3 s(-1) and 1.2 mM (Hodges & Merrifield, 1975).
Ligated HpaI had a specific activity of 0.5-1.5 x 10(6) U/mg, which compared favorably with
the expected value of 1-2 x 10(6) U/mg (J. Benner, unpubl. obs.). Besides assisting in the
production of cytotoxic proteins, this technique could allow the easy insertion of unnatural
amino acids into a protein sequence.

<>

<1>Evdokimov, A.A., Sclavi, B., Zinoviev, V.V., Malygin, E.G., Hattman, S., Buckle, M.
<2>Study of bacteriophage T4-encoded dam DNA (Adenine-N-6)-methyltransferase binding with substrates by rapid laser  UV cross-linking.
<3>J. Biol. Chem.
<4>282
<5>26067-26076
<6>2007
<7>DNA methyltransferases of the Dam family ( including bacteriophage T4-encoded Dam DNA
(adenine- N-6)-methyltransferase ( T4Dam)) catalyze
methyl group transfer from S-adenosyl-L-methionine ( AdoMet), producing
S-adenosyl-Lhomocysteine ( AdoHcy) and methylated adenine residues in
palindromic GATC sequences. In this study, we describe the application
of direct ( i. e. no exogenous cross-linking reagents) laser UV
cross-linking as a universal non-perturbing approach for studying the
characteristics of T4Dam binding with substrates in the equilibrium and
transient modes of interaction. UV irradiation of the enzyme center dot
substrate complexes using an Nd3(+): yttrium aluminum garnet laser at
266nm resulted in up to 3 and > 15% yields of direct T4Dam
cross-linking to DNA and AdoMet, respectively. Consequently, we were
able to measure equilibrium constants and dissociation rates for enzyme
center dot substrate complexes. In particular, we demonstrate that both
reaction substrates, specific DNA and AdoMet( or product AdoHcy),
stabilized the ternary complex. The improved substrate affinity for the
enzyme in the ternary complex significantly reduced dissociation rates
( up to 2 orders of magnitude). Several of the parameters obtained (
such as dissociation rate constants for the binary T4Dam center dot
AdoMet complex and for enzyme complexes with a non-fluorescent
hemimethylated DNA duplex) were previously inaccessible by other means.
However, where possible, the results of laser UV cross-linking were
compared with those of fluorescence analysis. Our study suggests that
rapid laser UV cross-linking efficiently complements standard DNA
methyltransferase-related tools and is a method of choice to probe
enzyme-substrate interactions in cases in which data cannot be acquired
by other means.

<>

<1>Evdokimov, A.A., Zinoviev, V.V., Kuznetsov, V.V., Netesova, N.A., Malygin, E.G.
<2>Design of oligonucleotide inhibitors for human DNA methyltransferase 1.
<3>Mol. Biol. (Mosk)
<4>43
<5>418-425
<6>2009
<7>Mammalian DNA methyltransferase 1 (Dnmt1) is responsible for copying the DNA methylation
pattern during cell division. Since Dnmt1 plays an
important role in carcinogenesis, it is of particular interest to
search for its specific inhibitors. To design oligonucleotide
inhibitors of human Dnmt1, a number of singlestranded, double-stranded,
and hairpin DNA structures containing a canonical or a modified Dnmt1
recognition site (5'-CG) were constructed on the basis of a 22-nt
sequence. Structural features such as a C:A mismatch,
phosphorothioates, and hairpins proved capable of incrementally
increasing the oligonucleotide affinity for Dnmt1. An improvement of
inhibitory properties was also achieved by replacing the target
cytosine with 5,6-dihydro-5-azacytosine, 5-methyl-2-pyrimidinone, or
6-methyl-pyrrolo-[2,3-d]-2-pyrimidinone. The concentration that caused
50% inhibition of methylation of 1 mu M poly(dI-dC) center dot
poly(dI-dC), a conventional DNA substrate, was approximately 10(-7) M
for the most efficient oligonucleotides. Under the same in vitro
conditions, these oligonucleotide inhibitors demonstrated a
substantially stronger effect compared to known Dnmt1 inhibitors, which
were used as controls.

<>

<1>Evdokimov, A.A., Zinoviev, V.V., Malygin, E.G.
<2>Effect of S-adenosyl-L-methionine and its analogues on site-specific binding of DNA-(adenine-N6)-methyltransferase of T4 phage with the oligonucleotide substrate.
<3>Bioorg. Khim.
<4>26
<5>797-800
<6>2000
<7>Using fluorescence of 2-aminopurine-substituted oligonucleotide duplexes, "flipping" of the
target base in the process of interaction of T4 DNA-(adenine-N6)-methyltransferase (EC
2.1.1.72) with the substrate double-stranded DNA was revealed.  It was shown that
S-adenosyl-L-methionine, the methyl group donor, induces the reorientation of the enzyme
relative to the asymmetrically modified recognition site.

<>

<1>Evdokimov, A.A., Zinoviev, V.V., Malygin, E.G.
<2>The Kinetic Mechanism of Phage T4 DNA-[N6-Adenine]-Methyltransferase.
<3>Mol. Biol. (Mosk)
<4>36
<5>849-861
<6>2002
<7>Kinetic analysis of methyl group transfer from S-adenosyl-L-methionine to the GATC recognition
site catalyzed by the phage T4 DNA-[N6-adenine]-methyltransferase [EC 2.1.1.72] showed that
the reverse reaction is at least 500 times lower than the direct one.  The overall pattern of
product inhibition corresponds to an ordered steady-state mechanism following the sequence
SAM/DNA/metDNA/SAH/.  Pronounced inhibition was observed at high concentrations of the
20-meric substrate duplex, which may be attributed to formation of a dead-end complex
MTase-SAH-DNA.  In contrast, high SAM concentrations proportionally accelerated the reaction.
Thus, the reaction may include a stage whereby the binding of SAM and the release of SAH are
united into one concerted event.  Computer fitting of alternative kinetic schemes to the
aggregate of experimental data revealed that the most plausible mechanism involves
isomerization of the enzyme.

<>

<1>Evdokimov, A.A., Zinoviev, V.V., Malygin, E.G., Schlagman, S.L., Hattman, S.
<2>Bacteriophage T4 Dam DNA-[N6-adenine]Methyltransferase. Kinetic evidence for a catalytically essential conformational change in the ternary complex.
<3>J. Biol. Chem.
<4>277
<5>279-286
<6>2002
<7>We carried out a steady state kinetic analysis of the bacteriophage T4
DNA-[N(6)-adenine]methyltransferase (T4 Dam) mediated methyl group transfer reaction from
S-adenosyl-l-methionine (AdoMet) to Ade in the palindromic recognition sequence, GATC, of a
20-mer oligonucleotide duplex. Product inhibition patterns were consistent with a steady
state-ordered bi-bi mechanism in which the order of substrate binding and product (methylated
DNA, DNA(Me) and S-adenosyl-l-homocysteine, AdoHcy) release was AdoMet, DNA, DNA(Me), Hcy. A
strong reduction in the rate of methylation was observed at high concentrations of the
substrate 20-mer DNA duplex. In contrast, increasing substrate AdoMet concentration led to
stimulation in the reaction rate with no evidence of saturation. We propose the following
model. Free T4 Dam (initially in conformational form E) randomly interacts with substrates
AdoMet and DNA to form a ternary T4 Dam-AdoMet-DNA complex in which T4 Dam has isomerized to
conformational state F, which is specifically adapted for catalysis. After the chemical step
of methyl group transfer from AdoMet to DNA, product DNA(Me) dissociates relatively rapidly
(k(off) = 1.7 s(-1)) from the complex. In contrast, dissociation of product AdoHcy proceeds
relatively slowly (k(off) = 0.018 s(-1)), indicating that its release is the rate-limiting
step, consistent with k(cat) = 0.015 s(-1). After AdoHcy release, the enzyme remains in the F
conformational form and is able to preferentially bind AdoMet (unlike form E, which randomly
binds AdoMet and DNA), and the AdoMet-F binary complex then binds DNA to start another
methylation cycle. We also propose an alternative pathway in which the release of AdoHcy is
coordinated with the binding of AdoMet in a single concerted event, while T4 Dam remains in
the isomerized form F. The resulting AdoMet-F binary complex then binds DNA, and another
methylation reaction ensues. This route is preferred at high AdoMet concentrations.

<>

<1>Evdokimova, N.M., Aleshkin, G.I., Skavronskaya, A.G.
<2>Inheritance and phenotypic expression of plasmids coding EcoRI restriction endonuclease in Vibrio cholerae cells.
<3>Biull. Eksp. Biol. Med.
<4>100
<5>472-474
<6>1985
<7>Restriction endonucleases are widely used nowadays to clone DNA fragments of different origin
with the aid of vector molecules of plasmids and bacteriophages.  The process of DNA
recombination, in which restriction endonucleases take part, also takes place in vitro.  The
obtaining of hybrid molecules in vivo, due to the action of restriction endonucleases can in
many cases facilitate the task of cloning foreign DNA in bacterial cells.  For instance, by
means of a technique based on completion of a recA-independent recombination process,
EcoRI-dependent cloning of DNA has been carried out in Escherichia coli cells in vivo, so that
it was possible to obtain plasmids carrying recB+C+ genes or enterotoxin Ent genes.
Reproduction of this process in Vibrio cholerae opens up a new approach to the obtaining of
recombinant plasmids carrying genes of V. cholerae and, in particular, genes responsible for
the leading pathogenetic sign of V. cholerae, namely toxin formation.  For this purpose it was
necessary to transfer into V. cholerae cells a plsmid coding restriction endonuclease EcoRI,
to preserve it in V. cholerae cells, and to express EcoRI activity in them.  The aim of this
investigaton was to construct such a plasmid and to investigate the phenotypic expression of
its genetic determinants in V. cholerae cells.

<>

<1>Everett, E.A.
<2>Structure-function analysis of the EcoRI DNA Methylase.
<3>Ph.D. Thesis, Univ. of California, Santa Barbara, UMI, , Ann Arbor
<4>
<5>1-190
<6>1990
<7>The structural characteristics of the E. coli EcoRI methylase involved in methyl transfer from
S-adenosyl-L-methionine to DNA and in binding these moities were analyzed. This study provides
a "picture" of the methylase structure binding regions, critical residues and catalytic state.
Not much structure-function information is available about DNA methylases or S-
adenosyl-L-methionine binding enzymes. For DNA methylases, little is known about
protein-cofactor interactions or the mechanisms of methyl transfer to DNA. Information about
the E. coli EcoRI methylase is of interest since S-adenosyl-L-methionine is the methyl group
donor for a large range of enzymes and the importance of DNA methylation is becoming
increasingly obvious. Determination of structure-function relationships used traditional and
innovative protein chemistries. These included proteolytic patterns, photoaffinity labeling
with [methyl-3H]8-azido-S-adenosyl-L-methionine, chemical modification of cysteine and
histidine residues, fluorescence spectroscopy and mass spectrometry.

<>

<1>Everett, E.A., Falick, A.M., Reich, N.O.
<2>Identification of a critical cysteine in EcoRI DNA methyltransferase by mass spectrometry.
<3>J. Biol. Chem.
<4>265
<5>17713-17719
<6>1990
<7>EcoRI DNA methyltransferase (MTase) is rapidly inactivated by N-ethylmaleimide
with concomitant incorporation of 2 mol of N-ethyl[2-3H]maleimide/mol of
functional monomer.  Preincubation of the enzyme with either
S-adenosylmethionine or DNA reduces the rate of activity loss, whereas
preincubation with DNA and the S-adenosylmethionine analog sinefungin
completely protects the enzyme from inactivation.  An endo proteinase Glu-C
digest of N-ethyl[2-3H]maleimide-modified enzyme was prepared and separated by
high pressure liquid chromatography.  Modified and unmodified
cysteine-containing peptides were located and identified by radioactivity, mass
spectrometry, and tandem mass spectrometry.  In the absence of any ligands,
cysteines 25, 116, and 223 are modified by N-ethylmaleimide; in the presence of
DNA and sinefungin Cys-223 is essentially unmodified.  Thus, N-ethylmaleimide
modification of Cys-223 in EcoRI DNA MTase is responsible for the loss of
enzyme activity.  Cys-223 is preceded by Asn, and this (or Cys-Asn) occurs with
high frequency in adenine and cytosine (N-4) DNA MTases.  Direct involvement of
cysteine in methyl transfer reactions to adenine N-6 and cytosine N-4 is
supported by the similarity of the reactions catalyzed by adenine N-6 and
cytosine N-4 DNA MTases, the frequent presence of Asn-flanking Cys, and the
importance of Cys-223 to EcoRI MTase function.

<>

<1>Everett, E.A., Reich, N.O.
<2>Determination of the EcoRI methylase Adomet binding region.
<3>J. Cell Biol.
<4>107
<5>854a
<6>1988
<7>Although S-Adenosylmethionine (Adomet) is a methyl group donor in many
enzymatic reactions including DNA methylation, structural features of DNA
methylases necessary for Adomet binding and DNA methylation have not yet been
determined.  Investigating structural determinants of activity is critical in
understanding methylation and its role in gene expression, DNA repair,
restriction-modification and carcinogenesis.  Our research involves the E. coli
EcoRI methylase which transfers the methyl group from Adomet to the second
adenine in GAATC DNA sequences.  Methylation protects the site from cleavage by
the corresponding endonuclease.  We have synthesized an Adomet photoaffinity
analog, 3H8-AzidoAdomet, which binds specifically to the Adomet site with
similar binding and catalytic constants.  Covalent modification of the binding
site with 3H8-AzAdomet and subsequent chromatographic, electrophoretic,
autoradiographic and sequence analyses of proteolytic digests are being
employed in isolating the methylase portion critical for Adomet binding.  A
peptide segment of the Adomet binding site to which 3H8-AzAdomet binds has been
obtained and is being identified along with the covalently modified amino acid
residue or residues.

<>

<1>Everett, E.A., Reich, N.O.
<2>Determination of the EcoRI methylase Adomet binding region.
<3>ACS Abstracts
<4>196
<5>94
<6>1988
<7>Although S-Adenosylmethionine (Adomet) is a methyl group donor in many
enzymatic reactions including DNA methylation, structural features of DNA
methylases necessary for Adomet binding and DNA methylation have not yet been
determined.  Investigating structural determinants of activity is critical in
understanding methylation and its role in gene expression, DNA repair,
restriction-modification and carcinogenesis.  Our research involves the E. coli
EcoRI methylase which transfers the methyl group from Adomet to the second
adenine in GAATTC DNA sequences.  Methylation protects the site from cleavage
by the corresponding endonuclease.  We have synthesized an Adomet photoaffinity
analog.  3H8-AzidoAdomet, which binds specifically to the Adomet site with
similar binding and catalytic constants.  Covalent modification of the binding
site with 3H8-AzAdomet and subsequent chromatographic, electrophoretic,
autoradiographic and sequence analyses of proteolytic digests are being
employed in isolating the methylase portion critical for Adomet binding.  A
peptide segment of the Adomet binding site to which 3H8-AzAdomet binds has been
obtained and is being identified along with the covalently modified amino acid
residue or residues.

<>

<1>Everett, E.A., Reich, N.O.
<2>EcoRI DNA methylase activity is eliminated upon histidine residue modification.
<3>Biochem. Biophys. Res. Commun.
<4>164
<5>233-237
<6>1989
<7>The E. coli EcoRI DNA methylase activity is completely eliminated in five
minutes upon incubation with the histidine residue specific reagent diethyl
pyrocarbonate.  In that two moles of N-ethoxyformylimidazole per mole of
methylase are detected spectroscopically upon inactivation and activity is not
restored by hydroxylamine, it is likely that activity loss is due to double
modification of a single histidine residue.  This information is critical in
determining the enzymatic mechanism, causes of the pH-activity curve, designing
protein mutants and interpreting previous structure-function data.

<>

<1>Everett, K.D., Kahane, S., Bush, R.M., Friedman, M.G.
<2>An unspliced group I intron in 23S rRNA links chlamydiales, chloroplasts, and mitochondria.
<3>J. Bacteriol.
<4>181
<5>4734-4740
<6>1999
<7>Chlamydia was the only genus in the order Chlamydiales until the
recent characterization of Simkania negevensis Z(T) and Parachlamydia
acanthamoebae strains. The present study of Chlamydiales 23S ribosomal
DNA (rDNA) focuses on a naturally occurring group I intron in the I-
CpaI target site of 23S rDNA from S. negevensis. The intron, SnLSU. 1,
belonged to the IB4 structural subgroup and was most closely related to
large ribosomal subunit introns that express single-motif, LAGLIDADG
endonucleases in chloroplasts of algae and in mitochondria of amoebae.
RT-PCR and electrophoresis of in vivo rRNA indicated that the intron
was not spliced out of the 23S rRNA. The unspliced 658-nt intron is the
first group I intron to be found in bacterial rDNA or rRNA, and it may
delay the S. negevensis developmental replication cycle by affecting
ribosomal function.

<>

<1>Everroad, R.C., Woebken, D., Singer, S.W., Burow, L.C., Kyrpides, N., Woyke, T., Goodwin, L., Detweiler, A., Prufert-Bebout, L., Pett-Ridge, J.
<2>Draft Genome Sequence of an Oscillatorian Cyanobacterium, Strain ESFC-1.
<3>Genome Announcements
<4>1
<5>e00527-13
<6>2013
<7>The nonheterocystous filamentous cyanobacterium strain ESFC-1 has recently been isolated from
a marine microbial mat system, where it was identified as belonging
to a recently discovered lineage of active nitrogen-fixing microorganisms. Here,
we report the draft genome sequence of this isolate. The assembly consists of 3
scaffolds and contains 5,632,035 bp with a GC content of 46.5%.

<>

<1>Evers, S., Bessler, C., Feesche, J., Maurer, K.-H., Ehrenreich, A., Veith, B., Liesegang, H., Henne, A., Herzberg, C., Gottschalk, G.
<2>Restriction modification systems of Bacillus licheniformis and improved biotechnological production procedures based on them.
<3>German Patent Office
<4>DE 102004041717 A
<5>
<6>2006
<7>
<>

<1>Ewens, W.J.
<2>Inference problems in population genetics:  DNA sequences, restriction endonucleases and ascertainment sampling.
<3>Proc. R. Soc. Lond. B Biol. Sci.
<4>219
<5>223-239
<6>1983
<7>A major trend of population genetics theory in the 1970s was the increased
emphasis on inductive arguments, based on observed genetic data, rather than on
deductive arguments based on theory and models.  This occurred in part because
the deductive theory had largely fulfilled its role of describing evolution as
a genetic process, and in part because of the increasing amounts of data
available on the genetic constitution of natural populations.  Inference
procedures raise difficulties not present in the deductive theory.  Often
conditional arguments are necessary since the data often must fulfil some
condition to be observed.  Different inference procedures, having different
efficiencies, apply for data from different apparatuses.  Care must be taken in
deciding what it is the inference concerns.  These problems are illustrated by
reference to restriction endonuclease techniques and ascertainment sampling.

<>

<1>Ezekiel, U.R., Zassenhaus, P.
<2>Evidence for a site-specific endonuclease in yeast mitochondria which recognizes the sequence 5'GCCCGGC.
<3>Biochem. Biophys. Res. Commun.
<4>201
<5>208-214
<6>1994
<7>We have discovered a mitochondrial, site-specific DNase in Saccharomyces cerevisiae with
properties like that of a type II restriction endonuclease. The enzyme, termed SceIII, cleaves
the palindromic sequence, 5'GCCGGC, to give 3' ends recessed by 4 bases. SceIII is the first
restriction-like endonuclease to be described in yeast mitochondria.

<>

<1>Fabbri, M., Garzon, R., Cimmino, A., Liu, Z., Zanesi, N., Callegari, E., Liu, S., Alder, H., Costinean, S., Fernandez-Cymering, C., Volinia, S., Guler, G., Morrison, C.D., Chan, K.K., Marcucci, G., Calin, G.A., Huebner, K., Croce, C.M.
<2>MicroRNA-29 family reverts aberrant methylation in lung cancer by targeting DNA methyltransferases 3A and 3B.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>15805-15810
<6>2007
<7>MicroRNAs (miRNAs) are small, noncoding RNAs that regulate expression of many genes. Recent
studies suggest roles of miRNAs in
carcinogenesis. We and others have shown that expression profiles of
miRNAs are different in lung cancer vs. normal lung, although the
significance of this aberrant expression is poorly understood. Among
the reported down-regulated miRNAs in lung cancer, the miRNA (miR)-29
family (29a, 29b, and 29c) has intriguing complementarities to the
3'-UTRs of DNA methyltransferase (DNMT)3A and -3B (de novo
methyltransferases), two key enzymes involved in DNA methylation, that
are frequently up-regulated in lung cancer and associated with poor
prognosis. We investigated whether miR-29s could target DNMT3A and -B
and whether restoration of miR-29s could normalize aberrant patterns of
methylation in non-small-cell lung cancer. Here we show that expression
of miR-29s is inversely correlated to DNMT3A and -3B in lung cancer
tissues, and that miR-29s directly target both DNMT3A and -3B. The
enforced expression of miR-29s in lung cancer cell lines restores
normal patterns of DNA methylation, induces reexpression of
methylation-silenced tumor suppressor genes, such as FHIT and WWOX, and
inhibits tumorigenicity in vitro and in vivo. These findings support a
role of miR-29s in epigenetic normalization of NSCLC, providing a
rationale for the development of miRNA-based strategies for the
treatment of lung cancer.

<>

<1>Fabian, J.S., Mattson, T., Bazar, L.S., Chirikjian, J.G.
<2>Characterization of the Bacillus amyloliquefaciens HI methyltransferase.
<3>FASEB J.
<4>12
<5>A1446
<6>1998
<7>Type II DNA methyltransferases are part of bacterial restriction-modification systems.
Methyltransferases function as monomers, recognize a palindromic DNA sequence and use
S-adenosyl-methionine as the methyl donor.  The Bacillus amyloliquefaciens methyltransferase
is a 49 kDa protein which methylates the N4 position of cytosine in the DNA sequence GGATCC.
Protein sequence alignment shows that M.BamHI has structural homology to M.TaqI.  The
carboxyl-terminal region of M.BamHI also contains a region showing 72% identity to the
predicted DNA binding region of M.DpnA.  Here, we report the properties of truncated proteins
missing either the N or C terminus amino acids.  These truncated proteins have been tested for
SAM and DNA binding as well as methylation activity.  However, specificity appears to be
relaxed. In contrast, deletion of the carboxyl-terminus removes both DNA binding and
methylating activity although SAM binding activity is retained.  The peptide produced from in
vitro transcription/translation of the carboxyl-terminal region is being tested for DNA
binding activity.  These results assign the DNA binding activity of M.BamHI to the
carboxyl-terminal of the protein as predicted from sequence alignment studies.  The N-terminus
region is necessary for increasing the specificity of the enzyme.

<>

<1>Fabian, J.S., Mattson, T., Chirikjian, J.G.
<2>Comparative study of the biochemical properties of cellular and viral DNA-methyltransferases from Bacillus amyloliquefaciens H1.
<3>FASEB J.
<4>10
<5>A1103
<6>1996
<7>Methyltransferases (Mtases) are a class of enzymes which place methyl groups onto a variety of
substrates using S-adenosyl-methionine as a cofactor.  In prokaryotes, Mtases are often part
of a restriction-modification system in which their role is to protect the host DNA from
cleavage by the restriction enzyme.  Bacillus amyloliquefaciens HI contains two distinct
methyltransferases (M.BamHI and M.BamHII) which recognize and methylate the same double
stranded DNA sequence 5'-GGATCC-3'.  Both methylases have been cloned, sequenced, purified and
characterized.  The two Mtases have little primary amino acid sequence homology, and therefore
provide an ideal model for a variety of comparative studies.  The cellular enzyme (M.BamHI),
is active as a 49KD monomer and methylates the exocyclic N4 nitrogen of the internal cytosine.
M.BamHI, associated with a prophage, is a 31KD monomeric enzyme that also methylates the
internal cytosine, although it is not yet known if it has an N4 or a C5 methylation activity.
Time course experiments will be presented indicating that the specific activity of the viral
methyltransferase (M.BamHII) is significantly greater than that of the cellular methylase
(M.BamHI).  The nature of this difference in activity is being investigated.  Gel shift
experiments will be presented that allow a comparison of the dissociation constants.  If the
activity of either BamHI or BamHII is similar to that for HhaI, which extrudes the target base
from the double helix, one might expect a bilobal conformation of the enzyme, at least when
associated with its target sequence.  The results of limited proteolysis experiments designed
to address this possibility will also be presented.

<>

<1>Fabian, J.S., Mattson, T., Chirikjian, J.G.
<2>Characterization of the Bacillus amyloliquefaciens HI methyltransferase.
<3>FASEB J.
<4>11
<5>A900
<6>1997
<7>Type II DNA methyltransferases are part of bacterial restriction-modification systems.  Most
of the Mtases function as monomers, recognize a palindromic DNA sequence and use
S-Adenosyl-Methionine as the methyl donor.  The Bacillus amyloliquefaciens methyltransferase
is a 49kDa protein which methylates at the N4 position of cytosine in the sequence GGATCC.
Crystal structures of three DNA Mtases show that they are folded into two domains.  The
C-terminal domain is responsible for specific recognition and binding of the target DNA
sequence.  The N-terminal domain contains the catalytic site formed by the conserved motif IV
(NPPY in M.TaqI and PCQ in M.HhaI and M.HaeIII) and a deep cleft that harbors the cofactor
SAM.  Crystal structures have also revealed that these enzymes extrude the target base from
the double helix into the catalytic pocket where the methyl transfer occurs.  Partial
proteolytic digestion of the BamHI methylase using Proteinase K generates a 38kDa peptide
fragment that maintains methylase activity in vitro.  N-terminal peptide sequencing indicates
that the fragment is missing 103 amino acids from the N-terminal end.  Gel shift assays
indicate that the 38kDa fragment is able to interact with the DNA in a sequence dependent
manner.  Preliminary kinetic analysis of the proteolytic fragment shows a decreased Km, Kcat
and Vmax.  We are currently constructing C-terminal deletion mutants and will test them for
methylase and DNA binding activity.

<>

<1>Faelker, S., Schilling, J., Schmidt, M.A., Heusipp, G.
<2>Overproduction of DNA adenine methyltransferase alters motility, invasion, and the lipopolysaccharide O-antigen composition of Yersinia enterocolitica.
<3>Infect. Immun.
<4>75
<5>4990-4997
<6>2007
<7>DNA adenine methyltransferase not only regulates basic cellular functions but also interferes
with the proper expression of virulence factors in various pathogens.  We showed previously
that for the human pathogen Yersinia enterocolitica, overproduction of Dam results in
increased invasion of epithelial cells.  Since invasion and motility are coordinately
regulated in Y. enterocolitica, we analyzed the motility of a Dam-overproducting (DamOP)
strain and found it to be highly motile.  In DamOP strains, the operon encoding the master
regulator of flagellum biosynthesis, flh DC, is upregulated.  We show that the increased
invasion is not due to enhanced expression of known and putative Y. enterocolitica invasion
and adhesion factors, such as Inv, YadA, Ail, Myf fibrils, Pil, or Flp pili.  However,
overproduction of Dam results in an increased amount of rough lipopolysaccharide molecules
lacking O-antigen side chains, this implies that reduced steric hindrance by LPS might
contribute to increased invasion by a Y. enterocolitica DamOP strain.  Our data add an
important new aspect to the various virulence-associated phenotypes influenced by DNA
methylation in Y. enterocolitica and indicate that Dam targets regulatory processes modulating
the composition and function of the bacterial surface.

<>

<1>Faelker, S., Schmidt, M.A., Heusipp, G.
<2>DNA methylation in Yersinia enterocolitica: Role of the DNA adenine methyltransferase in mismatch repair and regulation of virulence factors.
<3>Microbiology
<4>151
<5>2291-2299
<6>2005
<7>DNA adenine methyltransferase plays an important role in physiological processes of
Gram-negative bacteria such as mismatch repair and replication.  In addition, Dam regulates
the expression of virulence genes in various species.  The authors cloned the dam gene of
Yersinia enterocolitica and showed that Dam is essential for viability.  Dam overproduction in
Y. enterocolitica resulted in an increased frequency of spontaneous mutation and decreased
resistance to 2-aminopurine; however, these effects were only marginal compared to the effect
of overproduction of Escherichia coli-derived Dam in Y. enterocolitica, implying different
roles or activities of Dam in mismatch repair of the two species.  These differences in Dam
function are not the cause for the essentiality of Dam in Y. enterocalitica, as Dam of E. coli
can complement a dam defect in Y. enterocolitica.  Instead, Dam seems to interfere with
expression of essential genes.  Furthermore, Dam mediates virulence of Y. enterocolitica.  Dam
overproduction results in increased tissue culture invasion of Y. enterocolitica, while the
expression of specifically in vivo-expressed genes is not altered.

<>

<1>Fajardo-Sanchez, E., Stricher, F., Paques, F., Isalan, M., Serrano, L.
<2>Computer design of obligate heterodimer meganucleases allows efficient cutting of custom DNA sequences.
<3>Nucleic Acids Res.
<4>36
<5>2163-2173
<6>2008
<7>Meganucleases cut long (>12 bp) unique sequences in genomes and can be used to induce targeted
genome engineering by homologous recombination
in the vicinity of their cleavage site. However, the use of natural
meganucleases is limited by the repertoire of their target sequences,
and considerable efforts have been made to engineer redesigned
meganucleases cleaving chosen targets. Homodimeric meganucleases such
as I-CreI have provided a scaffold, but can only be modified to
recognize new quasi-palindromic DNA sequences, limiting their general
applicability. Other groups have used dimer-interface redesign and
peptide linkage to control heterodimerization between related
meganucleases such as I-DmoI and I-CreI, but until now there has been
no application of this aimed specifically at the scaffolds from
existing combinatorial libraries of I-CreI. Here, we show that
engineering meganucleases to form obligate heterodimers results in
functional endonucleases that cut non-palindromic sequences. The
protein design algorithm (FoldX v2.7) was used to design specific
heterodimer interfaces between two meganuclease monomers, which were
themselves engineered to recognize different DNA sequences. The new
monomers favour functional heterodimer formation and prevent homodimer
site recognition. This design massively increases the potential
repertoire of DNA sequences that can be specifically targeted by
designed I-CreI meganucleases and opens the way to safer targeted
genome engineering.

<>

<1>Falb, M., Pfeiffer, F., Palm, P., Rodewald, K., Hickmann, V., Tittor, J., Oesterhelt, D.
<2>Living with two extremes: conclusions from the genome sequence of Natronomonas pharaonis.
<3>Genome Res.
<4>15
<5>1336-1343
<6>2005
<7>Natronomonas pharaonis is an extremely haloalkaliphilic archaeon that was isolated from
salt-saturated lakes of pH 11. We sequenced its 2.6-Mb
GC-rich chromosome and two plasmids (131 and 23 kb). Genome analysis
suggests that it is adapted to cope with severe ammonia and heavy metal
deficiencies that arise at high pH values. A high degree of nutritional
self-sufficiency was predicted and confirmed by growth in a minimal medium
containing leucine but no other amino acids or vitamins. Genes for a
complex III analog of the respiratory chain could not be identified in the
N. pharaonis genome, but respiration and oxidative phosphorylation were
experimentally proven. These studies identified protons as coupling ion
between respiratory chain and ATP synthase, in contrast to other
alkaliphiles using sodium instead. Secretome analysis predicts many
extracellular proteins with alkaline-resistant lipid anchors, which are
predominantly exported through the twin-arginine pathway. In addition, a
variety of glycosylated cell surface proteins probably form a protective
complex cell envelope. N. pharaonis is fully equipped with archaeal signal
transduction and motility genes. Several receptors/transducers signaling
to the flagellar motor display novel domain architectures. Clusters of
signal transduction genes are rearranged in haloarchaeal genomes, whereas
those involved in information processing or energy metabolism show a
highly conserved gene order.

<>

<1>Faldu, P.R., Kothari, V.V., Kothari, C.R., Rawal, C.M., Domadia, K.K., Patel, P.A., Bhimani, H.D., Raval, V.H., Parmar, N.R., Nathani, N.M., Koringa, P.G., Joshi, C.G., Kothari, R.K.
<2>Draft Genome Sequence of Textile Azo Dye-Decolorizing and -Degrading Pseudomonas  aeruginosa Strain PFK10, Isolated from the Common Effluent Treatment Plant of the  Ankleshwar Industrial Area of Gujarat, India.
<3>Genome Announcements
<4>2
<5>e00019-14
<6>2014
<7>Here, we report the draft genome sequence of Pseudomonas aeruginosa strain PFK10, isolated
from the common effluent treatment plant (CETP) of the Ankleshwar
industrial area of Gujarat, India. The 6.04-Mb draft genome sequence of strain
PFK10 provides information about the genes encoding enzymes that enable the
strain to decolorize and degrade textile azo dye.

<>

<1>Falentin, H., Cousin, S., Clermont, D., Creno, S., Ma, L., Chuat, V., Loux, V., Rudiger, P., Bizet, C., Bouchier, C.
<2>Draft Genome Sequences of Five Strains of Lactobacillus acidophilus, Strain CIP 76.13T, Isolated from Humans, Strains CIRM-BIA 442 and CIRM-BIA 445, Isolated  from Dairy Products, and Strains DSM 20242 and DSM 9126 of Unknown Origin.
<3>Genome Announcements
<4>1
<5>e00658-13
<6>2013
<7>Lactobacillus acidophilus is a natural inhabitant of mammalian gastrointestinal systems and is
used in dairy and pharmaceutical products. Five draft genome
sequences, covering 1,995,790 nucleotides (nt) on average, are divided into 19 to
34 scaffolds covering 1,995 to 2,053 genes. The draft genome sequences were
compared to the sequence of the L. acidophilus NCFM dairy strain.

<>

<1>Falentin, H., Naquin, D., Loux, V., Barloy-Hubler, F., Loubiere, P., Nouaille, S., Lavenier, D., Le Bourgeois, P., Francois, P., Schrenzel, J., Hernandez, D., Even, S., Le Loir, Y.
<2>Genome Sequence of Lactococcus lactis subsp. lactis bv. diacetylactis LD61.
<3>Genome Announcements
<4>2
<5>e01176-13
<6>2014
<7>Lactococcus lactis is widely used in the dairy industry. We report the draft genome sequence
of L. lactis subsp. lactis bv. diacetylactis LD61, an industrial
and extensively studied strain. In contrast to the closely related and
plasmidless strain IL1403, LD61 contains 6 plasmids, and the genome sequence
provides additional information related to adaptation to the dairy environment.

<>

<1>Falgenhauer, L., Yao, Y., Fritzenwanker, M., Schmiedel, J., Imirzalioglu, C., Chakraborty, T.
<2>Complete Genome Sequence of Phage-Like Plasmid pECOH89, Encoding CTX-M-15.
<3>Genome Announcements
<4>2
<5>e00356-14
<6>2014
<7>A nonconjugative and nontypable plasmid of a clinical Escherichia coli isolate expressing
resistance to extended-spectrum cephalosporins (ESCs) was isolated and sequenced. The plasmid
pECOH89 contains a CTX-M-15 resistance cassette and comprises 111,741 bp, with strong homology
to bacteriophage-like plasmids and to the Salmonella-specific bacteriophage SSU5.

<>

<1>Falkow, S., Formal, S.B.
<2>Restriction in genetic crosses between Escherichia coli and Shigella flexneri.
<3>J. Bacteriol.
<4>100
<5>540-541
<6>1969
<7>Shigella flexneri restricts Escherichia coli deoxyribonucleic acid (DNA) and
can modify phage DNA so that it is restricted in E. coli.

<>

<1>Fallico, V., Ross, R.P., Fitzgerald, G.F., McAuliffe, O.
<2>Novel conjugative plasmids from the natural isolate Lactococcus lactis subspecies cremoris DPC3758: A repository of genes for the potential improvement of dairy starters.
<3>J. Dairy Sci.
<4>95
<5>3593-3608
<6>2012
<7>A collection of 17 natural lactococcal isolates from raw milk cheeses were studied in terms of
their plasmid distribution, content, and
diversity. All strains in the collection harbored an abundance of
plasmids, including Lactococcus lactis ssp. cremoris DPC3758, whose
8-plasmid complement was selected for sequencing. The complete
sequences of pAF22 (22,388 kb), pAF14 (14,419 kb), pAF12 (12,067 kb),
pAF07 (7,435 kb), and pAF04 (3,801 kb) were obtained, whereas gene
functions of technological interest were mapped to pAF65 (65 kb) and
pAF45 (45 kb) by PCR. The plasmids of L. lactis DPC3758 were found to
encode many genes with the potential to improve the technological
properties of dairy starters. These included 3 anti-phage
restriction/modification (R/M) systems (1 of type I and 2 of type II)
and genes for immunity/resistance to nisin, lacticin 481, cadmium, and
copper. Regions encoding conjugative/mobilization functions were
present in 6 of the 8 plasmids, including those containing the R/M
systems, thus enabling the food-grade transfer of these mechanisms to
industrial strains. Using cadmium selection, the sequential stacking of
the R/M plasmids into a plasmid-free host provided the recipient with
increased protection against 936- and c2-type phages. The association
of food-grade selectable markers and mobilization functions on L.
lactis DPC3758 plasmids will facilitate their exploitation to obtain
industrial strains with enhanced phage protection and robustness. These
natural plasmids also provide another example of the major role of
plasmids in contributing to host fitness and preservation within its
ecological niche.

<>

<1>Falquet, L., Calderon-Copete, S.P., Frey, J.
<2>Draft Genome Sequence of the Virulent Clostridium chauvoei Reference Strain JF4335.
<3>Genome Announcements
<4>1
<5>e00593-13
<6>2013
<7>Clostridium chauvoei is the etiological agent of blackleg, a disease of cattle and sheep with
high mortality rates, causing severe economic losses in livestock
production. Here, we report the draft genome sequence of the virulent C. chauvoei
strain JF4335 (2.8 Mbp and 28% G+C content) and the annotation of the genome.

<>

<1>Falquet, L., Liljander, A., Schieck, E., Gluecks, I., Frey, J., Jores, J.
<2>Complete Genome Sequences of Virulent Mycoplasma capricolum subsp. capripneumoniae Strains F38 and ILRI181.
<3>Genome Announcements
<4>2
<5>e01041-14
<6>2014
<7>Contagious caprine pleuropneumonia (CCPP) caused by Mycoplasma capricolum subsp.
capripneumoniae is a severe epidemic affecting mainly domestic Caprinae species
but also affects wild Caprinae species. M. capricolum subsp. capripneumoniae
belongs to the 'Mycoplasma mycoides cluster.' The disease features prominently in
East Africa, in particular Kenya, Tanzania, and Ethiopia. CCPP also endangers
wildlife and thus affects not only basic nutritional resources of large
populations but also expensively built-up game resorts in affected countries.
Here, we report the complete sequences of two M. capricolum subsp.
capripneumoniae strains: the type strain F38 and strain ILRI181 isolated druing a
recent outbreak in Kenya. Both genomes have a G+C content of 24% with sizes of
1,016,760 bp and 1,017,183 bp for strains F38 and ILRI181, respectively.

<>

<1>Falsafi, S., Reich, N.
<2>Molecular dynamic-based prediction of DNA structure changes in DNA sequence and correlation to substrate specificity in EcoRI DNA methyltransferase.
<3>FASEB J.
<4>6
<5>A221
<6>1992
<7>The object of this computational investigation is to determine the structural
consequences of single base-pair substitutions in the canonical site GAATTC of
14 base-pair oligonucleotide duplexes.  The computational schemes reported
herein are guided by the earlier approaches established by Kollman and
co-workers and include 100 picoseconds of molecular dynamics.  The atomic
trajectories are analyzed for temporal variations of the phosphodiester and
glycosidic torsion angles and the helicoidal parameters using both AMBER (1986)
AMBER 3.0, UCSF) and Dials and Windows (J. Biomol. Str. Dyn., 6, 669(1989))
algorithms.  We first performed two computations using the crystal structure of
the Dickerson dodecamer and the standard Arnott B-DNA structure as the starting
points.  The average backbone torsion angles alpha, beta, gamma, delta,
epsilon, chi 1, chi 2 taken over the entire simulation time show 3% difference
between the two structures, suggesting convergence towards a unique final
structure.  The degree of agreement is even greater for the average helical
twist omega.  These results corroborate Kollman's earlier report, therefore
establishing the validity of our approach.  In view of our ongoing
investigation with a series of 14 base-pair oligonucleotides (Biochemistry
(1991) 30, 2933-2939; Nucleic Acids Research, in press), the sequence
d(GGCGGAATTCGCGG) was subjected to 100 picoseconds of molecular dynamics.  The
calculated backbone torsion angles show good agreement with the identical
conformational values obtained from an earlier calculation on the dodecamer.
In view of these consistencies, we have used the results from this later
computation to gauge the structural variations that result from single
base-pair substitutions within the canonical site GAATTC of d(GGCGGAATTCGCGG).
It is hoped that these structural differences will ultimately help elucidate
the mechanisms that describe the enzyme-DNA binding and catalysis for EcoRI
methyltransferase.

<>

<1>Fan, L., Bo, S., Chen, H., Ye, W., Kleinschmidt, K., Baumann, H.I., Imhoff, J.F., Kleine, M., Cai, D.
<2>Genome sequence of Bacillus subtilis subsp. spizizenii gtP20b isolated from the Indian Ocean.
<3>J. Bacteriol.
<4>193
<5>1276
<6>2010
<7>Bacillus subtilis is a model organism of aerobic spore-forming Gram-positive bacteria and is
of great industrial significance as the source of diverse novel functional molecules. Here we
present to our knowledge the first genome sequence of a Bacillus subtilis strain gtP20b
isolated from the marine environment. A subset of candidate genes and gene clusters were
identified, which are potentially involved in production of diverse functional molecules like
novel ribosomal and non-ribosomal antimicrobial peptides. The genome sequence described in
this paper is due to its high strain-specificity of great importance for basic as well as
applied researches on marine organisms.

<>

<1>Fan, L., Liu, Y., Li, Z., Baumann, H.I., Kleinschmidt, K., Ye, W., Imhoff, J.F., Kleine, M., Cai, D.
<2>Draft genome sequence of a marine Streptomyces sp. strain PP-C42, isolated from the Baltic Sea.
<3>J. Bacteriol.
<4>193
<5>3691-3692
<6>2011
<7>Streptomyces, a branch of aerobic Gram-positive bacteria represents the largest genus of
actinobacteria. The streptomycetes are characterized by a
complex secondary metabolism and produce over two-thirds of the clinically
used natural antibiotics today. Here we report the draft genome sequence
of a Streptomyces strain PP-C42 isolated from the marine environment. A
subset of unique genes and gene clusters for diverse secondary metabolites
as well as antimicrobial peptides (AMPs) could be identified from the
genome, showing great promise as a source for novel bioactive compounds.

<>

<1>Fang, G. et al.
<2>Genome-wide mapping of methylated adenine residues in pathogenic Escherichia coli using single-molecule real-time sequencing.  LID - 10.1038/nbt.2432 [doi].
<3>Nat. Biotechnol.
<4>30
<5>1232
<6>2012
<7>Single-molecule real-time (SMRT) DNA sequencing allows the systematic detection of chemical
modifications such as methylation but has not previously been applied on a genome-wide scale.
We used this approach to detect 49,311 putative 6-methyladenine (m6A) residues and 1,407
putative 5-methylcytosine (m5C) residues in the genome of a pathogenic Escherichia coli
strain. We obtained strand-specific information for methylation sites and a quantitative
assessment of the frequency of methylation at each modified position. We deduced the sequence
motifs recognized by the methyltransferase enzymes present in this strain without prior
knowledge of their specificity. Furthermore, we found that deletion of a phage-encoded
methyltransferase-endonuclease (restriction-modification; RM) system induced global
transcriptional changes and led to gene amplification, suggesting that the role of RM systems
extends beyond protecting host genomes from foreign DNA.

<>

<1>Fang, N., Xu, S.-Y.
<2>Method for cloning and expression of BsrGI restriction endonuclease and BsrGI methyltransferase in E. coli.
<3>International Patent Office
<4>WO 200463328
<5>
<6>2004
<7>The present invention relates to recombinant DNA encoding the BsrGI restriction endonuclease
as well as BsrGI methyltransferase, expression of BsrGI restriction endonuclease and BsrGI
methyltransferase in E. coli cells containing the recombinant DNA.
NOVELTY - An isolated DNA encoding the BsrGI restriction endonuclease (and
BsrGI methylase) which is obtainable from
Bacillus stearothermophilus GR75 (or ATCC PTA-4892), is new. DETAILED
DESCRIPTION - INDEPENDENT CLAIMS are also included for the following:
(1) a recombinant DNA vector comprising a vector into which a DNA
segment encoding the BsrGI restriction endonuclease has been inserted
or a vector which comprises the isolated DNA above; (2) a host cell
transformed by the vector of (1); and (3) a method of producing
recombinant BsrGI restriction endonuclease comprising culturing a host
cell transformed with the vector above under conditions for expression
of the endonuclease and methylase. USE - The restriction endonucleases
and methylases are useful in the laboratory for cleaving DNA molecules
into small fragments for molecular cloning and gene characterization.
EXAMPLE - Genomic DNA was prepared from Bacillus stearothermophilus
GR75 strain collection. The genomic DNA and a cloning vector pUC-Cm
were digested completely with BamHI, HindIII, PstI, SacI, SalI, SphI
and XbaI at 37degreesC for an hour. Genomic DNA was ligated to the
vector with compatible ends overnight using T4 DNA ligase. Plasmid DNA
was prepared, generating a mixed plasmid library. This was challenged
and transformed in Escherichia coli. After digestion with BsrGI and gel
electrophoresis, one true resistant clone was found. This was again
subjected to restriction mapping. The bsrGIM gene generated was
analyzed. One amino acid sequence translated from the DNA sequence
indicated conserved amino-methyltransferase motifs. The bsrGIM gene has
1947 bp encoding a protein of 76.2 kDa. M.BsrGI was predicted to be an
amino-methyltransferase. A Bst genomic gene was also subjected to the
same treatment generating a BsrGI thermostable restriction
endonuclease. (49 pages)

<>

<1>Fang, N., Xu, S.-Y.
<2>Method for cloning and expression of BsrGI restriction endonuclease and BsrGI methyltransferase in E. coli.
<3>US Patent Office
<4>US 06869786
<5>
<6>2005
<7>The present invention relates to recombinant DNA encoding the BsrGI restriction endonuclease
as well as BsrGI methyltransferase, expression
of BsrGI restriction endonuclease and BsrGI methyltransferase in E.
coli cells containing the recombinant DNA.

<>

<1>Fang, X., Fang, Z., Zhao, J., Zou, Y., Li, T., Wang, J., Guo, Y., Chang, D., Su, L., Ni, P., Liu, C.
<2>Draft Genome Sequence of Pseudomonas aeruginosa Strain ATCC 27853.
<3>J. Bacteriol.
<4>194
<5>3755
<6>2012
<7>Pseudomonas aeruginosa is a common bacterium that can cause disease. The versatility of
Pseudomonas aeruginosa enables the organism to infect damaged
tissues or those with reduced immunity which cause inflammation and sepsis. Here
we report the genome sequence of the strain ATCC 27853.

<>

<1>Fang, Y., Huang, Y., Li, Q., Chen, H., Yao, Z., Pan, J., Gu, J., Tang, B., Wang, H.G., Yu, B., Tong, Y.G., Zou, Q.M., Mao, X.H.
<2>First Genome Sequence of a Burkholderia pseudomallei Isolate in China, Strain BPC006, Obtained from a Melioidosis Patient in Hainan.
<3>J. Bacteriol.
<4>194
<5>6604-6605
<6>2012
<7>Melioidosis, caused by Burkholderia pseudomallei, is considered to be endemic to  Northern
Australia and Southeast Asia, with high mortality and relapse rates,
regardless of powerful antibiotic therapy. Here we report the first genome
sequence of Burkholderia pseudomallei strain BPC006, obtained from a melioidosis
patient in Hainan, China. The genome sizes of the 2 chromosomes were determined
to be 4,001,777 bp and 3,153,284 bp.

<>

<1>Fanning, T.G., Kreutzfeldt, H.-J., Davies, R.W., Schreier, P.H.
<2>Detection of restriction endonucleases with a lac repressor-lac operator filter binding assay.
<3>FEBS Lett.
<4>61
<5>237-239
<6>1976
<7>The use of restriction enzymes as experimental tools has progressed rapidly
within the last several years.  The purification of these enzymes, however,
often poses a problem: since acid-soluable material is normally not released a
convenient assay for enzyme activity does not exist.  Thus, gel electrophoresis
or viscometry of digested DNA has often been employed to detect restriction
enzymes during purification.  These methods are, however, time consuming and
involve large quantities of DNA and/or expensive equipment.  We report here a
test for restriction enzymes that avoids these complications; the test is
inexpensive, rapid and can be done with very small quantities of DNA (0.02
microgram per test is normally sufficient).

<>

<1>Farfan, M., Spataro, N., Sanglas, A., Albarral, V., Loren, J.G., Bosch, E., Fuste, M.C.
<2>Draft Genome Sequence of the Aeromonas diversa Type Strain.
<3>Genome Announcements
<4>1
<5>e00330-13
<6>2013
<7>We present here the first genome sequence of the Aeromonas diversa type strain (CECT 4254(T)).
This strain was isolated from the leg wound of a patient in New
Orleans (Louisiana) and was originally described as enteric group 501 and
distinguished from A. schubertii by DNA-DNA hybridization and phenotypical
characterization.

<>

<1>Farias, M.E., Revale, S., Mancini, E., Ordonez, O., Turjanski, A., Cortez, N., Vazquez, M.P.
<2>Genome sequence of Sphingomonas sp. S17, isolated from an alkaline, hyperarsenic and hypersaline volcanic-associated lake at high altitude in the Argentinean Puna.
<3>J. Bacteriol.
<4>193
<5>3686-3687
<6>2011
<7>The High-Altitude Andean Lakes (HAAL) in the Argentinian Puna-High Andes region represent an
almost unexplored ecosystem exposed to extreme
conditions (high UV irradiation, hypersalinity, drastic temperature
changes, desiccation, high pH). Here we present the first genome sequence
isolated from this extreme environment, a Sphingomonas sp.

<>

<1>Farlow, J., Filippov, A.A., Sergueev, K.V., Hang, J., Kotorashvili, A., Nikolich, M.P.
<2>Comparative whole genome analysis of six diagnostic brucellaphages.
<3>Gene
<4>541
<5>115-122
<6>2014
<7>Whole genome sequencing of six diagnostic brucellaphages, Tbilisi (Tb), Firenze
(Fz), Weybridge (Wb), S708, Berkeley (Bk) and R/C, was followed with genomic
comparisons including recently described genomes of the Tb phage from Mexico
(TbM) and Pr phage to elucidate genomic diversity and candidate host range
determinants. Comparative whole genome analysis revealed high sequence
homogeneity among these brucellaphage genomes and resolved three genetic groups
consistent with defined host range phenotypes. Group I was composed of Tb and Fz
phages that are predominantly lytic for Brucella abortus and Brucella neotomae;
Group II included Bk, R/C, and Pr phages that are lytic mainly for B. abortus,
Brucella melitensis and Brucella suis; Group III was composed of Wb and S708
phages that are lytic for B. suis, B. abortus and B. neotomae. We found that the
putative phage collar protein is a variable locus with features that may be
contributing to the host specificities exhibited by different brucellaphage
groups. The presence of several candidate host range determinants is illustrated
herein for future dissection of the differential host specificity observed among
these phages.

<>

<1>Farrance, I.K., Eadie, J.S., Ivarie, R.
<2>Improved chemistry for oligodeoxyribonucleotide synthesis substantially improves restriction enzyme cleavage of a synthetic 35-mer.
<3>Nucleic Acids Res.
<4>17
<5>1231-1245
<6>1989
<7>Two DNA duplexes of identical sequence and 35 nt in length were synthesized by
an original and a highly improved version of phosphoramidite chemistry.  By
base composition analysis, DNA synthesized by improved chemistry (termed
DMTS-imp) contained no detectable modified bases while DNA synthesized by the
original chemistry (termed DMTS-std) had a large number of modifications.
Under optimal reaction conditions, HhaI and RsaI cleaved the DMTS-std duplex to
76-77% completion and the DMTS-imp duplex to 96-99% completion.  Restriction
analysis and piperidine treatment yielded estimates of approx. 3.0% modified
nucleotides in DMTS-std and approx. 1.0% in DMTS-imp.  Overall, the
improvements in chemistry inreased the restriction efficiency of synthetic DNA
up to 10-fold.

<>

<1>Farrell, S.O.
<2>A fast restriction enzyme experiment for the undergraduate biochemistry lab.
<3>J. Chem. Educ.
<4>71
<5>1095-1096
<6>1994
<7>We recently updated out laboratory curriculum to include some popular and important techniques
of molecular biology. We have designed a rapid experiment using restriction enzymes that can
be done in one three-hour lab period. This experiment is one of the highlights of the course,
and almost every student gets positive results.

<>

<1>Farrugia, D.N., Elbourne, L.D.H., Hassan, K.A., Eijkelkamp, B.A., Tetu, S.G., Brown, M.H., Shah, B.S., Peleg, A.Y., Mabbutt, B.C., Paulsen, I.T.
<2>The Complete Genome and Phenome of a Community-Acquired Acinetobacter baumannii.
<3>PLoS ONE
<4>8
<5>e58628
<6>2013
<7>Many sequenced strains of Acinetobacter baumannii are established nosocomial pathogens capable
of resistance to multiple antimicrobials.
Community-acquired A. baumannii in contrast, comprise a minor
proportion of all A. baumannii infections and are highly susceptible to
antimicrobial treatment. However, these infections also present acute
clinical manifestations associated with high reported rates of
mortality. We report the complete 3.70 Mbp genome of A. baumannii
D1279779, previously isolated from the bacteraemic infection of an
Indigenous Australian; this strain represents the first
community-acquired A. baumannii to be sequenced. Comparative analysis
of currently published A. baumannii genomes identified twenty-four
accessory gene clusters present in D1279779. These accessory elements
were predicted to encode a range of functions including polysaccharide
biosynthesis, type I DNA restriction-modification, and the metabolism
of novel carbonaceous and nitrogenous compounds. Conversely, twenty
genomic regions present in previously sequenced A. baumannii strains
were absent in D1279779, including gene clusters involved in the
catabolism of 4-hydroxybenzoate and glucarate, and the A. baumannii
antibiotic resistance island, known to bestow resistance to multiple
antimicrobials in nosocomial strains. Phenomic analysis utilising the
Biolog Phenotype Microarray system indicated that A. baumannii D1279779
can utilise a broader range of carbon and nitrogen sources than
international clone I and clone II nosocomial isolates. However,
D1279779 was more sensitive to antimicrobial compounds, particularly
beta-lactams, tetracyclines and sulphonamides. The combined genomic and
phenomic analyses have provided insight into the features
distinguishing A. baumannii isolated from community-acquired and
nosocomial infections.

<>

<1>Faruqi, A.F., Ahmed, N.
<2>Isolation and partial purification of a restriction endonuclease from Pseudomonas ovalis.
<3>Pak. J. Sci. Ind. Res.
<4>30
<5>390-392
<6>1987
<7>A restriction endonuclease, PovI, has been isolated and partially purified from
Pseudomonas ovalis by high speed centrifugation and fractionation on a column
of Biogel followed by dialysis and ion exchange chromatography on a
phosphocellulose column.  The presence of the restriction endonuclease was
detected by a simple assay procedure using agarose slab gel electrophoresis.

<>

<1>Fassbender, A., Lesche, R., Piepenbrock, C., Rujan, T., Berlin, K., Distler, J., Koenig, T.
<2>Method for determining the methylation pattern of a polynucleic acid.
<3>International Patent Office
<4>WO 200688978
<5>
<6>2006
<7>Particular aspects relate to a method for determining the methylation pattern of a polynucleic
acid, comprising: a) preparing a solution comprising a mixture of fragments of the polynucleic
acid; b) coupling the fragments with a substance being detectable with a detection  method; c)
contacting a solution comprising the fragments of b) with a DNA microarray having a plurality
of different immobilized oligonucleotides, each comprising at least one methylation site, at
respectively assigned different locations thereon, the contacing under conditions affording
hybridization of fragments with correlated immobilized oligonucleotides under defined
stringency, and wherein the immobilized oligonucleotides have a length of less than 200 bases;
d) optionally performing a washing step; and e) detecting, using the physical detection
method, such as immobilized nucleic acids to which solution fragments are hybridized and/or to
which solution fragments are not hybridized.

<>

<1>Fatemi, M., Hermann, A., Gowher, H., Jeltsch, A.
<2>Dnmt3a and Dnmt1 functionally cooperate during de novo methylation of DNA.
<3>Eur. J. Biochem.
<4>269
<5>4981-4984
<6>2002
<7>Dnmt3a is a de novo DNA methyltransferase that modifies unmethylated DNA. In contrast Dnmt1
shows high preference for hemimethylated DNA. However, Dnmt1 can be activated for the
methylation of unmodified DNA. We show here that the Dnmt3a and Dnmt1 DNA methyltransferases
functionally cooperate in de novo methylation of DNA, because a fivefold stimulation of
methylation activity is observed if both enzymes are present. Stimulation is observed if
Dnmt3a is used before Dnmt1, but not if incubation with Dnmt1 precedes Dnmt3a, demonstrating
that methylation of the DNA by Dnmt3a stimulates Dnmt1 and that no physical interaction of
Dnmt1 and Dnmt3a is required. If Dnmt1 and Dnmt3a were incubated together a slightly increased
stimulation is observed that could be due to a direct interaction of these enzymes. In
addition, we show that Dnmt1 is stimulated for methylation of unmodified DNA if the DNA
already carries some methyl groups. We conclude that after initiation of de novo methylation
of DNA by Dnmt3a, Dnmt1 becomes activated by the pre-existing methyl groups and further
methylates the DNA. Our data suggest that Dnmt1 also has a role in de novo methylation of DNA.
This model agrees with the biochemical properties of these enzymes and provides a mechanistic
basis for the functional cooperation of different DNA MTases in de novo methylation of DNA
that has also been observed in vivo.

<>

<1>Fatemi, M., Hermann, A., Pradhan, S., Jeltsch, A.
<2>The activity of the murine DNA methyltransferase Dnmt1 is controlled by interaction of the catalytic domain with the N-terminal part of the enzyme leading to an allosteric activation of the enzyme after binding to methylated DNA.
<3>J. Mol. Biol.
<4>309
<5>1189-1199
<6>2001
<7>The mammalian DNA methyltransferase Dnmt1 is responsible for the maintenance of the pattern of
DNA methylation in vivo. It is a large multidomain enzyme comprising 1620 amino acid residues.
We have purified and characterized individual domains of Dnmt1 (NLS-containing domain, NlsD,
amino acid residues: 1-343; replication foci-directing domain, 350-609; Zn-binding domain
(ZnD), 613-748; polybromo domain, 746-1110; and the catalytic domain (CatD), 1124-1620). CatD,
ZnD and NlsD bind to DNA, demonstrating the existence of three independent DNA-binding sites
in Dnmt1. CatD shows a preference for binding to hemimethylated CpG-sites; ZnD prefers
methylated CpGs; and NlsD specifically binds to CpG-sites, but does not discriminate between
unmethylated and methylated DNA. These results are not compatible with the suggestion that the
target recognition domain of Dnmt1 resides in the N terminus of the enzyme. We show by
protein-protein interaction assays that ZnD and CatD interact with each other. The isolated
catalytic domain does not methylate DNA, neither alone nor in combination with other domains.
Full-length Dnmt1 was purified from baculovirus-infected insect cells. Under the experimental
conditions, Dnmt1 has a strong (50-fold) preference for hemimethylated DNA. Dnmt1 is
stimulated to methylate unmodified CpG sites by the addition of fully methylated DNA. This
effect is dependent on Zn, suggesting that binding of methylated DNA to ZnD triggers the
allosteric activation of the catalytic center of Dnmt1. The allosteric activation model can
explain kinetic data obtained by others. It suggests that Dnmt1 might be responsible for
spreading of methylation, a process that is observed during aging and carcinogenesis but may
be important for de novo methylation of DNA.

<>

<1>Fauman, E.B., Blumenthal, R.M., Cheng, X.
<2>Structure and evolution of AdoMet-dependent methyltransferases.
<3>S-Adenosylmethionine-dependent Methyltransferases: Structures and Functions., World Scientific Publishing, Cheng, X., Blumenthal, R.M., Singapore
<4>
<5>1-38
<6>1999
<7>A review of many different types of SAM-sependent Mtases, including DNA methyltransferases,
RNA methyltransferases and many others.

<>

<1>Fauvart, M., Sanchez-Rodriguez, A., Beullens, S., Marchal, K., Michiels, J.
<2>Genome sequence of Rhizobium etli CNPAF512, a nitrogen-fixing symbiont isolated from bean root nodules in Brazil.
<3>J. Bacteriol.
<4>193
<5>3158-3159
<6>2011
<7>Rhizobium etli is a Gram-negative soil-dwelling alpha-proteobacterium that carries out
symbiotic biological nitrogen fixation in close association
with legume hosts. R. etli strains exhibit high sequence divergence and
are geographically structured, with a potentially dramatic influence on
the outcome of symbiosis. Here, we present the genome sequence of R. etli
CNPAF512, a Brazilian isolate from bean nodules. We anticipate that the
availability of genome sequences of R. etli strains from distinctly
different areas will provide valuable new insights into the geographic
mosaic of the R. etli pangenome and the evolutionary dynamics that shape
it.

<>

<1>Fazakerley, G.V., Gabarro-Arpa, J., Lebret, M., Guy, A., Guschlbauer, W.
<2>The GTm6AC sequence is overwound and bent.
<3>Nucleic Acids Res.
<4>17
<5>2541-2556
<6>1989
<7>By a combination of distance constraints obtained from NMR spectra and
molecular mechanics calculations we have determined the three dimensional
structure of the self-complementary decanucleotide d(CGCGTm6ACGCG).
Methylation of an adenine at a position 3' to T induces significant
conformational changes relative to B-DNA.  This arises from the close proximity
of the four methyl groups in the large groove in the centre of the sequence.
The helical twist between the two T.m6A base pairs is found to be 45, as for
D-DNA, and is accompanied by a high negative value of the wedge roll angle
between these base pairs.  The overall nonzero wedge roll observed shows that
the helix is bent.  These constraints appear to be material for the absence of
the sequence T-m6A in natural DNAs.

<>

<1>Fazakerley, G.V., Quignard, E., Teoule, R., Guy, A., Guschlbauer, W.
<2>A two-dimensional H-NMR study of the dam methylase site:  comparison between the hemimethylated GATC sequence, its unmethylated analogue and a hemimethylated CATG sequence:  The sequence dependence of methylation upon base-pair lifetimes.
<3>Eur. J. Biochem.
<4>167
<5>397-404
<6>1987
<7>We report two-dimensional NOE (NOESY) spectra on the sequence d(GCGATCATGG) d(CCATGATCGC)
which contains the unmethylated dam site. As expected the DNA adopts a B-form conformation but
appears to be distorted at the TG step of the second strand. This distorsion, probably
bending, is not seen on the opposite strand. When the first strand is methylated on adenine in
the GATC or CATG sequence the NOESY spectra indicate little or no change in the conformation.
However the single strand-duplex change is slowed down to the slow-exchange region on a proton
NMR time scale. We have assigned the exchangeable imino and cytidine amino resonances of the
three duplexes. From the imino linewidths as a function of temperature, we observe that the
unmethylated and the hemimethylated Gm6ATC duplexes melt normally from the ends. However, this
is not so for the hemimethylated Cm6ATG duplex which, apart from the terminal base pairs,
melts cooperatively and at high temperature. In spectra recorded in H2O a second duplex is
observed, for the Gm6ATC sequence, which we have not been able to identify. It is however
unlikely to be a hairpin structure. Ultraviolet-melting curves also indicate the presence of
two transitions for this duplex. The effect of methylation upon base-pair lifetimes has been
studied by comparing the above three duplexes. Little effect is observed upon methylation in
the GATC sequence but a drastic increase in the lifetimes of all base pairs is observed upon
methylation in the CATG sequence.

<>

<1>Fedoreyeva, L.I., Vanyushin, B.F.
<2>N6-Adenine DNA-methyltransferase in wheat seedlings.
<3>FEBS Lett.
<4>514
<5>305-308
<6>2002
<7>The N(6)-adenine DNA-methyltransferase was isolated from the vacuolar vesicle fraction of
wheat coleoptiles. In the presence of S-adenosyl-L-methionine the enzyme de novo methylates
the first adenine residue in the TGATCA sequence in the single- or double-stranded DNA
substrates but it prefers single-stranded structures. Wheat adenine DNA-methyltransferase
(wadmtase) is a Mg(2+)- or Ca(2+)-dependent enzyme with a maximum activity at pH 7.5-8.0.
Wadmtase seems to be responsible for mitochondrial DNA modification that might be involved in
the regulation of replication of mitochondria in plants.

<>

<1>Fedotova, E.A., Protsenko, A.S., Zakharova, M.V., Lavrova, N.V., Alekseevsky, A.V., Oretskaya, T.S., Karyagina, A.S., Solonin, A.S., Kubareva, E.A.
<2>SsoII-like DNA-methyltransferase Ecl18kI: Interaction between regulatory and methylating functions.
<3>Biochemistry
<4>74
<5>85-91
<6>2009
<7>The interaction of DNA-methyltransferase Ecl18kI (M.Ecl18kI) with a fragment of promoter
region of restriction-modification system SsoII
was studied. It is shown that dissociation constants of M.Ecl18kI and
M.SsoII complexes with DNA ligand carrying a regulatory site previously
characterized for M.SsoII have comparable values. A deletion derivative
of M.Ecl18kI, Delta(72-379)Ecl18kI, representing the N-terminal protein
region responsible for regulation, was obtained. It is shown that such
polypeptide fragment has virtually no interaction with the regulatory
site. Therefore, the existence of a region responsible for methylation
is necessary for maintaining M.Ecl18kI regulatory function. The
properties of methyl-transferase NlaX, which is actually a natural
deletion derivative of M.Ecl18kI and M.SsoII lacking the first 70 amino
acid residues and not being able to regulate gene expression of the
SsoII restriction-modification system, were studied. The ability of
mutant forms of M.Ecl18kI incorporating single substitutions in regions
responsible for regulation and methylation to interact with both sites
of DNA recognition was characterized. The data show a correlation
between DNA-binding activity of two M.Ecl18kI regions-regulatory and
methylating.

<>

<1>Feher, Z., Kiss, A., Venetianer, P.
<2>Expression of a bacterial modification methylase gene in yeast.
<3>Nature
<4>302
<5>266-268
<6>1983
<7>Methylation of specific cytosines in the DNA is generally believed to play some
role in the regulation of gene expression in eukaryotes.  However, some
eukaryotes, such as Drosophila and yeast (S. Hattman, personal communication)
seem not to contain 5-methylcytosine in their DNA.  It would be interesting to
test, how gene expression in such organisms would respond to the methylation of
specific cytosines in the genome.  As a first step towards this goal, we have
introduced the gene encoding the Bacillus sphaericus R modification methylase,
which methylates the internal cytosine within the recognition sequence 5'-GGCC,
into yeast cells.  Southern-type hybridization to DNAs isolated from the
transformed yeast clones revealed that the yeast plasmid carrying the
prokaryotic methylase gene, as well as the two chromosomal genes tested (his3
and leu2) were methylated, whereas the bulk of the yeast DNA remained largely
unmethylated.  This indicates that the Bacillus sphaericus modification
methylase was expressed in yeast but it modified only certain parts of the
yeast DNA.

<>

<1>Feher, Z., Schlagman, S.L., Miner, Z., Hattman, S.
<2>The UV excision-repair system of Saccharomyces cerevisiae is involved in the removal of methylcytosines formed in vivo by a cloned prokaryotic DNA methyltransferase.
<3>Curr. Genet.
<4>16
<5>461-464
<6>1989
<7>DNA methyltransferase activity is not normally found in yeast.  To investigate
the response of Saccharomyces cerevisiae to the presence of methylated bases,
we introduced the Bacillus subtilis SPR phage DNA-[cytosine-5]
methyltransferase gene on the shuttle vector, YEp51.  The methyltransferase
gene was functionally expressed in yeast under the control of the inducible
yeast GAL10 promoter.  Following induction we observed a time-dependent
methylation of yeast DNA in RAD+ and rad2 mutant strains; the rad2 mutant is
defective in excision-repair of UV-induced DNA damage.  Analysis of restriction
endonuclease digestion patterns revealed that the relative amount of methylated
DNA was greater in the excision defective rad2 mutant than in the RAD+ strain.
These data indicate that the yeast excision-repair system is capable of
recognizing and removing m5C residues.

<>

<1>Feher, Z., Schlagman, S.L., Miner, Z., Hattman, S.
<2>Expression of prokaryotic DNA methyltransferase genes in yeast.
<3>Zentralbl. Mikrobiol.
<4>145
<5>343
<6>1990
<7>DNA methyltransferase activity is not normally found in yeast. To investigate the response of
Saccharomyces cerevisiae to the presence of methylated bases, we introduced two phage-encoded
DNA methyltransferase genes on the shuttle vector, YEp51. The Bacillus subtilis phage SPR
enzyme methylates cytosine residues in three distinct recognition sequences [GGCC, CCGG, and
CC(A/T)GG], whereas the Escherichia coli phage T4 Dam enzyme methylates adenines primarily in
GATC sequences. Both phage-encoded genes were functionally expressed in yeast under the
control of the inducible yeast GAL10 promoter. DNA methyltransferase activity was assayed by
determining the susceptibility of yeast DNA to cleavage by restriction endonucleases:
resistance to HaeIII and sensitivity to DpnI indicate SPR- and T4 Dam-specific in vivo
methylation, respectively. Following induction of the GAL10 promoter we observed a
time-dependent methylation of yeast DNA in the RAD+, as well as rad2 and rad3 mutant strains;
these rad mutant strains are defective in excision-repair. Analysis of the restriction
endonuclease digestion patterns revealed that the relative amount of high molecular weight
methylated DNA was greater in the excision-defective rad mutants than in the RAD+ strain.
Furthermore, after methyltransferase gene expression was turned off (by the addition of
glucose to previously induced cells), we observed a decrease in methylation in the RAD+, but
not in the DNA of the rad mutant. These data indicate that the yeast excision-repair system is
capable of recognizing and removing m5C and m6A residues. All of our plasmid constructions
contain more than one AUG initiation codon at the 5' end of the mRNA transcript and some of
the AUGs are followed by an in-frame stop codon. Since we observed expression of the
methyltransferase genes, it would appear that, in yeast, one or two short ORFs in the leader
region do not necessarily prevent translation of a downstream gene.

<>

<1>Feher, Z., Schlagman, S.L., Miner, Z., Hattman, S.
<2>In vivo methylation of yeast DNA by prokaryotic DNA methyltransferases.
<3>Gene
<4>74
<5>193-195
<6>1988
<7>No detectable amounts of methylated bases are present in the DNA of the yeast, Saccharomyces
cerevisiae.  This feature and the availability of an Escherichia coli-yeast cloning system
make yeast an excellent host to study the factors affecting de novo DNA methylation, as well
as their attendant effects in a eukaryotic cell.  Earlier, Feher et al. (1983) observed
heterologous expression of the Bacillus sphaericus RI DNA-cytosine methyltransferase in yeast.
Methylation of two known chromosomal genes was detected, while the bulk DNA remained largely
unmethylated.  Heterologous expression of the E. coli DNA (N6-adenine)methyltransferase gene
(dam) in yeast was reported by Brooks et al. (1983) and Hoekstra and Malone (1985; 1986).

<>

<1>Feil, H. et al.
<2>Comparison of the complete genome sequences of Pseudomonas syringae pv. syringae B728a and pv. tomato DC3000.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>11064-11069
<6>2005
<7>The complete genomic sequence of Pseudomonas syringae pv. syringae B728a (Pss B728a) has been
determined and is compared with that of P. syringae
pv. tomato DC3000 (Pst DC3000). The two pathovars of this economically
important species of plant pathogenic bacteria differ in host range and
other interactions with plants, with Pss having a more pronounced
epiphytic stage of growth and higher abiotic stress tolerance and Pst
DC3000 having a more pronounced apoplastic growth habitat. The Pss B728a
genome (6.1 Mb) contains a circular chromosome and no plasmid, whereas the
Pst DC3000 genome is 6.5 mbp in size, composed of a circular chromosome
and two plasmids. Although a high degree of similarity exists between the
two sequenced Pseudomonads, 976 protein-encoding genes are unique to Pss
B728a when compared with Pst DC3000, including large genomic islands
likely to contribute to virulence and host specificity. Over 375
repetitive extragenic palindromic sequences unique to Pss B728a when
compared with Pst DC3000 are widely distributed throughout the chromosome
except in 14 genomic islands, which generally had lower GC content than
the genome as a whole. Content of the genomic islands varies, with one
containing a prophage and another the plasmid pKLC102 of Pseudomonas
aeruginosa PAO1. Among the 976 genes of Pss B728a with no counterpart in
Pst DC3000 are those encoding for syringopeptin, syringomycin, indole
acetic acid biosynthesis, arginine degradation, and production of ice
nuclei. The genomic comparison suggests that several unique genes for Pss
B728a such as ectoine synthase, DNA repair, and antibiotic production may
contribute to the epiphytic fitness and stress tolerance of this organism.

<>

<1>Feingersch, R., Suzuki, M.T., Shmoish, M., Sharon, I., Sabehi, G., Partensky, F., Beja, O.
<2>Microbial community genomics in eastern Mediterranean Sea surface waters.
<3>ISME J.
<4>4
<5>78-87
<6>2010
<7>Offshore waters of the eastern Mediterranean Sea are one of the most oligotrophic regions on
Earth in which the primary productivity is phosphorus limited. To study the unexplored
function and physiology of microbes inhabiting this system, we have analyzed a genomic library
from the eastern Mediterranean Sea surface waters by sequencing both termini of nearly 5000
clones. Genome recruitment strategies showed that the majority of high-scoring pairs
corresponded to genomes from the Alphaproteobacteria (SAR11-like and Rhodobacterales),
Cyanobacteria (Synechococcus and high-light
adapted Prochlorococcus) and diverse uncultured Gammaproteobacteria. The community structure
observed, as evaluated by both protein similarity scores or metabolic potential, was similar
to that found in the euphotic zone of the ALOHA station off Hawaii but very different from
that of deep aphotic
zones in both the Mediterranean Sea and the Pacific Ocean. In addition, a strong enrichment
toward phosphate and phosphonate uptake and utilization metabolism was also observed.

<>

<1>Fekete, P.Z., Brzuszkiewicz, E., Blum-Oehler, G., Olasz, F., Szabo, M., Gottschalk, G., Hacker, J., Nagy, B.
<2>DNA sequence analysis of the composite plasmid pTC conferring virulence and antimicrobial resistance for porcine enterotoxigenic Escherichia coli.
<3>Int. J. Med. Microbiol.
<4>302
<5>4-9
<6>2012
<7>In this study the plasmid pTC, a 90kb self-conjugative virulence plasmid
of the porcine enterotoxigenic Escherichia coli (ETEC) strain EC2173
encoding the STa and STb heat-stable enterotoxins and tetracycline
resistance, has been sequenced in two steps. As a result we identified
five main distinct regions of pTC: (i) the maintenance region responsible
for the extreme stability of the plasmid, (ii) the TSL (toxin-specific
locus comprising the estA and estB genes) which is unique and
characteristic for pTC, (iii) a Tn10 transposon, encoding tetracycline
resistance, (iv) the tra (plasmid transfer) region, and (v) the colE1-like
origin of replication. It is concluded that pTC is a self-transmissible
composite plasmid harbouring antibiotic resistance and virulence genes.
pTC belongs to a group of large conjugative E. coli plasmids represented
by NR1 with a widespread tra backbone which might have evolved from a
common ancestor. This is the first report of a completely sequenced animal
ETEC virulence plasmid containing an antimicrobial resistance locus,
thereby representing a selection advantage for spread of pathogenicity in
the presence of antimicrobials leading to increased disease potential.

<>

<1>Feldmann, K.A., Brover, V., Makarov, V., Swaller, T.
<2>Nucleotide sequences and polypeptides encoded thereby useful for modifying plant characteristics.
<3>US Patent Office
<4>US 7569389 A
<5>
<6>2009
<7>Isolated polynucleotides and polypeptides encoded thereby are described, together with the use
of those products for making transgenic plants.

<>

<1>Felgner, P., Davies, D.H., Liang, X.
<2>Compositions and methods for immunodominant antigens.
<3>International Patent Office
<4>WO 2008140478 A
<5>
<6>2008
<7>Contemplated compositions, devices, and methods comprise immunodominant antigenes from
selected human pathogens (Burkholderia pseudomallei, Borrelia burgdorferi, Brucella
malitensis, Chlamydia muridarum, Coxiella burnetii, Francisella tularensis, human Herpes virus
1 and 2, Mycobacterium tuberculosis, Plasmodium falciparum, and Vaccinia virus) can be used as
a vaccine, as diagnostic markers, and as therapeutic agents.  In particularly preferred
aspects, the antigens have quantified and known relative reactivities with respect to sera of
a population infected with the pathogen, and have a known association with a disease
parameter.

<>

<1>Felle, M., Joppien, S., Nemeth, A., Diermeier, S., Thalhammer, V., Dobner, T., Kremmer, E., Kappler, R., Langst, G.
<2>The USP7/Dnmt1 complex stimulates the DNA methylation activity of Dnmt1 and regulates the stability of UHRF1.
<3>Nucleic Acids Res.
<4>39
<5>8355-8365
<6>2011
<7>Aberrant DNA methylation is often associated with cancer and the formation of tumors; however,
the underlying mechanisms, in particular the
recruitment and regulation of DNA methyltransferases remain largely
unknown. In this study, we identified USP7 as an interaction partner of
Dnmt1 and UHRF1 in vivo. Dnmt1 and USP7 formed a soluble dimer complex
that associated with UHRF1 as a trimeric complex on chromatin. Complex
interactions were mediated by the C-terminal domain of USP7 with the
TS-domain of Dnmt1, whereas the TRAF-domain of USP7 bound to the
SRA-domain of UHRF1. USP7 was capable of targeting UHRF1 for
deubiquitination and affects UHRF1 protein stability in vivo. Furthermore,
Dnmt1, UHRF1 and USP7 co-localized on silenced, methylated genes in vivo.
Strikingly, when analyzing the impact of UHRF1 and USP7 on Dnmt1-dependent
DNA methylation, we found that USP7 stimulated both the maintenance and de
novo DNA methylation activity of Dnmt1 in vitro. Therefore, we propose a
dual role of USP7, regulating the protein turnover of UHRF1 and
stimulating the enzymatic activity of Dnmt1 in vitro and in vivo.

<>

<1>Felley-Bosco, E., Pourzand, C., Zijlstra, J., Amstad, P., Cerutti, P.
<2>A genotypic mutation system measuring mutations in restriction recognition sequences.
<3>Nucleic Acids Res.
<4>19
<5>2913-2919
<6>1991
<7>The RFLP/PCR approach (restriction fragment length polymorphism/polymerase
chain reaction) to genotypic mutation analysis described here measures
mutations in restriction recognition sequences.  Wild-type DNA is restricted
before the resistant, mutated sequences are amplified by PCR and cloned.  We
tested the capacity of this experimental design to isolate a few copies of a
mutated sequence of the human c-Ha-ras1 gene from a large excess of wild-type
DNA.  For this purpose we constructed a 272 bp fragment with 2 mutations in the
PvuII recognition sequence 1727-1732 and studied the rescue by RFLP/PCR of a
few copies of this PvuII mutant standard.  Following amplification with
Taq-polymerase induced bp changes were quantitated by hybridization with
specific oligonucleotide probes.  Our results indicate that 10 PvuII mutant
standard copies can be rescued from 10/8 to 10/9 wild-type sequences.  Taq
polymerase errors originating from unrestricted, residual wild-type DNA were
sequence dependent and consisted mostly of transversions originating at G.C bp.
In contrast to a doubly mutated standard the capacity to rescue single bp
mutations by RFLP/PCR is limited by Taq-polymerase errors.  Therefore, we
assessed the capacity of our protocol to isolate a G to T transversion mutation
at base pair 1698 of the MspI-site 1695-1698 of the c-Ha-ras1 gene from excess
wild-type ras1 DNA.  We found that 100 copies of the mutated ras1 fragment
could be readily rescued from 10/8 copies of wild-type DNA.

<>

<1>Fellinger, K., Rothbauer, U., Felle, M., Laengst, G., Leonhardt, H.
<2>Dimerization of DNA methyltransferase 1 is mediated by its regulatory domain.
<3>J. Cell. Biochem.
<4>106
<5>521-528
<6>2009
<7>DNA methylation is a major epigenetic modification and plays a crucial role in the regulation
of gene expression. Within the family of DNA
methyltransferases (Dnmts), Dnmt3a and 3b establish methylation marks during early
development, while Dnmt1 maintains methylation
patterns after DNA replication. The maintenance function of Dnmt1 is regulated by its large
regulatory N-terminal domain that interacts with
other chromatin factors and is essential for the recognition of hemi-methylated DNA.
Gelfiltration analysis showed that purified Dnmt1 elutes
at an apparent molecular weight corresponding to the size of a dimer. With protein interaction
assays we could show that Dnmt1 interacts with
itself through its N-terminal regulatory domain. By deletion analysis and
co-immunoprecipitations we mapped the dimerization domain to
the targeting sequence TS that is located in the center of the N-terminal domain (amino acids
310-629) and was previously shown to mediate
replication independent association with heterochromatin at chromocenters. Further mutational
analyses suggested that the dimeric complex
has a bipartite interaction interface and is formed in a head-to-head orientation. Dnmt1 dimer
formation could facilitate the discrimination of
hemi-methylated target sites as has been found for other palindromic DNA sequence recognizing
enzymes. These results assign an additional
function to the TS domain and raise the interesting question how these functions are spatially
and temporarily co-ordinated.

<>

<1>Feng, H., Zhi, Y., Sun, Y., Wei, X., Luo, Y., Zhou, P.
<2>Draft Genome Sequence of a Novel Streptomyces griseorubens Strain, JSD-1, Active  in Carbon and Nitrogen Recycling.
<3>Genome Announcements
<4>2
<5>e00650-14
<6>2014
<7>Streptomyces griseorubens JSD-1, isolated from compost-treated soil, is able to utilize
lignocellulose and nitrate as its sole carbon and nitrogen source for
growth. Here, we announce the draft genome map of this actinomycete. The genes
participating in lignocellulose and nitrate metabolism were picked out and
identified.

<>

<1>Feng, L., Ma, T., Zhang, J., Xu, F., Shi, L.
<2>Draft Genome Sequence of Bacillus subtilis QH-1, a Chromium-Reducing Bacterial Strain Isolated in Qinghai Province, China.
<3>Genome Announcements
<4>2
<5>e00182-14
<6>2014
<7>Bacillus subtilis strain QH-1, a chromium-reducing bacterial strain, was isolated from a soil
sample from a chromium-containing slag heap. The draft genome
sequence of this bacterium is 4,034,036 bp in length, with a G+C content of
43.71%, and it is predicted to contain 4,082 protein-coding genes.

<>

<1>Feng, L., Reeves, P.R., Lan, R., Ren, Y., Gao, C., Zhou, Z., Ren, Y., Cheng, J., Wang, W., Wang, J., Qian, W., Li, D., Wang, L.
<2>A recalibrated molecular clock and independent origins for the cholera pandemic clones.
<3>PLoS ONE
<4>3
<5>E4053
<6>2008
<7>Cholera, caused by Vibrio cholerae, erupted globally from South Asia in 7
pandemics, but there were also local outbreaks between the 6(th)
(1899-1923) and 7(th) (1961-present) pandemics. All the above are serotype
O1, whereas environmental or invertebrate isolates are antigenically
diverse. The pre 7th pandemic isolates mentioned above, and other minor
pathogenic clones, are related to the 7(th) pandemic clone, while the
6(th) pandemic clone is in the same lineage but more distantly related,
and non-pathogenic isolates show no clonal structure. To understand the
origins and relationships of the pandemic clones, we sequenced the genomes
of a 1937 prepandemic strain and a 6(th) pandemic isolate, and compared
them with the published 7(th) pandemic genome. We distinguished mutational
and recombinational events, and allocated these and other events, to
specific branches in the evolutionary tree. There were more mutational
than recombinational events, but more genes, and 44 times more base pairs,
changed by recombination. We used the mutational single-nucleotide
polymorphisms and known isolation dates of the prepandemic and 7(th)
pandemic isolates to estimate the mutation rate, and found it to be 100
fold higher than usually assumed. We then used this to estimate the
divergence date of the 6(th) and 7(th) pandemic clones to be about 1880.
While there is a large margin of error, this is far more realistic than
the 10,000-50,000 years ago estimated using the usual assumptions. We
conclude that the 2 pandemic clones gained pandemic potential
independently, and overall there were 29 insertions or deletions of one or
more genes. There were also substantial changes in the major integron,
attributed to gain of individual cassettes including copying from within,
or loss of blocks of cassettes. The approaches used open up new avenues
for analysing the origin and history of other important pathogens.

<>

<1>Feng, L., Wang, W., Cheng, J., Ren, Y., Zhao, G., Gao, C., Tang, Y., Liu, X., Han, W., Peng, X., Liu, R., Wang, L.
<2>Genome and proteome of long-chain alkane degrading Geobacillus thermodenitrificans NG80-2 isolated from a deep-subsurface oil reservoir.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>5602-5607
<6>2007
<7>The complete genome sequence of Geobacillus thermodenitrificans NG80-2, a thermophilic
bacillus isolated from a deep oil reservoir in Northern
China, consists of a 3,550,319-bp chromosome and a 57,693-bp plasmid. The
genome reveals that NG80-2 is well equipped for adaptation into a wide
variety of environmental niches, including oil reservoirs, by possessing
genes for utilization of a broad range of energy sources, genes encoding
various transporters for efficient nutrient uptake and detoxification, and
genes for a flexible respiration system including an aerobic branch
comprising five terminal oxidases and an anaerobic branch comprising a
complete denitrification pathway for quick response to dissolved oxygen
fluctuation. The identification of a nitrous oxide reductase gene has not
been previously described in Gram-positive bacteria. The proteome further
reveals the presence of a long-chain alkane degradation pathway; and the
function of the key enzyme in the pathway, the long-chain alkane
monooxygenase LadA, is confirmed by in vivo and in vitro experiments. The
thermophilic soluble monomeric LadA is an ideal candidate for treatment of
environmental oil pollutions and biosynthesis of complex molecules.

<>

<1>Feng, Y., Xu, H.F., Li, Q.H., Zhang, S.Y., Wang, C.X., Zhu, D.L., Cao, F.L., Li, Y.G., Johnston, R.N., Zhou, J., Liu, G.R., Liu, S.L.
<2>Complete Genome Sequence of Salmonella enterica Serovar Pullorum RKS5078.
<3>J. Bacteriol.
<4>194
<5>744
<6>2012
<7>Salmonella enterica serovar Pullorum is a chicken-adapted pathogen, causing pullorum disease.
Its strict host adaptation has been suspected to
result in gene decay. To validate this hypothesis and identify the decayed
genes, we sequenced the complete genome of S. Pullorum RKS5078. We found
263 pseudogenes in this strain and conducted functional analyses of the
decayed genes.

<>

<1>Feng, Z., Fang, G., Korlach, J., Clark, T., Luong, K., Zhang, X., Wong, W., Schadt, E.
<2>Detecting DNA modifications from SMRT sequencing data by modeling sequence context dependence of polymerase kinetic.
<3>PLOS Comp. Biol.
<4>9
<5>e1002935
<6>2013
<7>DNA modifications such as methylation and DNA damage can play critical regulatory roles in
biological systems. Single molecule, real time (SMRT) sequencing technology generates DNA
sequences as well as DNA polymerase kinetic information that can be used for the direct
detection of DNA modifications. We demonstrate that local sequence context has a strong impact
on DNA polymerase kinetics in the neighborhood of the incorporation site during the DNA
synthesis reaction, allowing for the possibility of estimating the expected kinetic rate of
the enzyme at the incorporation site using kinetic rate information collected from existing
SMRT sequencing data (historical data) covering the same local sequence contexts of interest.
We develop an Empirical Bayesian hierarchical model for incorporating historical data. Our
results show that the model could greatly increase DNA modification detection accuracy, and
reduce requirement of control data coverage. For some DNA modifications that have a strong
signal, a control sample is not even needed by using historical data as alternative to
control. Thus, sequencing costs can be greatly reduced by using the model. We implemented the
model in a R package named seqPatch, which is available at
https://github.com/zhixingfeng/seqPatch.

<>

<1>Feng, Z., Li, J., Zhang, J.R., Zhang, X.
<2>qDNAmod: a statistical model-based tool to reveal intercellular heterogeneity of  DNA modification from SMRT sequencing data.
<3>Nucleic Acids Res.
<4>42
<5>13488-13499
<6>2014
<7>In an isogenic cell population, phenotypic heterogeneity among individual cells is common and
critical for survival of the population under different environment conditions. DNA
modification is an important epigenetic factor that can regulate phenotypic heterogeneity. The
single molecule real-time (SMRT) sequencing technology provides a unique platform for
detecting a wide range of DNA modifications, including N6-methyladenine (6-mA),
N4-methylcytosine (4-mC) and 5-methylcytosine (5-mC). Here we present qDNAmod, a novel
bioinformatic tool for genome-wide quantitative profiling of intercellular heterogeneity of
DNA modification from SMRT sequencing data. It is capable of estimating proportion of isogenic
haploid cells, in which the same loci of the genome are differentially modified. We tested the
reliability of qDNAmod with the SMRT sequencing data of Streptococcus pneumoniae strain ST556.
qDNAmod detected extensive intercellular heterogeneity of DNA methylation (6-mA) in a clonal
population of ST556. Subsequent biochemical analyses revealed that the recognition sequences
of two type I restriction-modification (R-M) systems are responsible for the intercellular
heterogeneity of DNA methylation initially identified by qDNAmod. qDNAmod thus represents a
valuable tool for studying intercellular phenotypic heterogeneity from genome-wide DNA
modification.

<>

<1>Ferat, J.-L., Michel, F.
<2>Group II self-splicing introns in bacteria.
<3>Nature
<4>364
<5>358-361
<6>1993
<7>Like nuclear premessenger introns, group II self-splicing introns are excised from primary
transcripts as branched molecules, containing a 2'-5' phosphodiester bond.  For this reason,
it is widely believed that the ribozyme (catalytic RNA) core of group II introns, or some
evolutionarily related molecule, gave rise to the RNA components of the spliceosomal splicing
machinery of the eukaryotic nucleus.  One difficulty with this hypothesis has been the
restricted distribution of group II introns.  Unlike group I self-splicing introns, which
interrupt not only organelle primary transcripts, but also some bacterial and nuclear genes,
group II introns seemed to be confined to mitochondrial and chloroplast genomes.  We now
report the discovery of group II introns both in cyanobacteria (the ancestors of chloroplasts)
and the gamma subdivision of purple bacteria, or proteobacteria, whose alpha subdivision
probably gave rise to mitochondria.  At least one of these introns actually self-splices in
vitro.

<>

<1>Ferdous, A., Akaike, T., Maruyama, A.
<2>Inhibition of sequence-specific protein-DNA interaction and restriction endonuclease cleavage via triplex stabilization by poly(L-lysine)-graft-dextran copolymer.
<3>Biomacromolecules
<4>1
<5>186-193
<6>2000
<7>Triplex stabilization by poly(L-lysine)-graft-dextran copolymer within a mammalian gene
promoter inhibits the DNA binding activity of nuclear proteins from HeLa cells as well as
restriction endonuclease cleavage at physiological pH and ionic conditions in vitro.
Electrophoretic mobility shift assays using a 30-mer homopurine-homopyrimidine stretch
(located between -165 and -146 bp) of the promoter is engineered at the BamHI and PstI sites
of a plasmid DNA, copolymer-mediated triplex stabilization also remarkably competes
endonuclease activity of BamHI.  Finally, the triplex-stabilizing efficiency of the copolymer
is remarkably higher than that of spermine and benzo[e]pyridoindole.  Our results indicate
that the copolymer, regardless of the length of the target duplex, stabilizes triplexes for
significant inhibition of protein-DNA interaction and endonuclease activity.  Since stable
triplex formation within a short region out of a long native duplex is a prerequisite to
confer the therapeutic potential of antigene strategy, triplex stabilization on a long target
duplex and inhibition of nuclear protein - DNA interaction may open the possible in vivo
applicability of the copolymer.

<>

<1>Ferenci, T., Zhou, Z., Betteridge, T., Ren, Y., Liu, Y., Feng, L., Reeves, P.R., Wang, L.
<2>Genomic sequencing reveals regulatory mutations and recombinational events in the widely used MC4100 lineage of Escherichia coli K-12.
<3>J. Bacteriol.
<4>191
<5>4025-4029
<6>2009
<7>The genome of an Escherichia coli MC4100 strain with a lambda placMu50
fusion revealed numerous regulatory differences from MG1655, including one
that arose during laboratory storage. The 194 mutational differences
between MC4100(MuLac) and other K-12 sequences were mostly allocated to
specific lineages, indicating the considerable mutational divergence
between K-12 strains.

<>

<1>Fernandez, A., Gil, E., Cartelle, M., Perez, A., Beceiro, A., Mallo, S., Tomas, M.M., Perez-Llarena, F.J., Villanueva, R., Bou, G.
<2>Interspecies spread of CTX-M-32 extended-spectrum {beta}-lactamase and the role of the insertion sequence IS1 in down-regulating blaCTX-M gene expression.
<3>J. Antimicrob. Chemother.
<4>59
<5>841-847
<6>2007
<7>OBJECTIVES: To characterize the extended-spectrum beta-lactamases (ESBLs)
as well as their genetic environment in different isolates of
Enterobacteriaceae from a patient with repeated urinary tract infections.
METHODS: Two isolates of Escherichia coli and one Proteus mirabilis, all
with ESBL phenotypes, were studied. Conjugation experiments and
restriction fragment length polymorphisms (RFLPs) were performed. Cloning
of the bla genes was by plasmid restriction and fragments ligation.
Antibiotic susceptibility testing was by Etest. The genetic environment
was analysed by direct sequencing of the DNA surrounding the bla gene.
RT-PCR was performed to study the differences in the bla(CTX-M) gene
expression. RESULTS: The bla gene was transferred by conjugation from the
three clinical isolates, which by RFLP showed the same plasmid. The bla
gene and surrounding sequences were cloned, an approximately 9 kbp AccI
fragment was sequenced and the bla(CTX-M-32) gene was identified. The MICs
of ceftazidime for transconjugants and transformants bearing the
bla(CTX-M-32) gene were lower than those previously reported. Analysis of
the DNA surrounding the ESBL gene revealed a new genetic structure with
two insertion sequences, IS5 and IS1, located immediately upstream of the
bla(CTX-M-32) gene; IS1 was located between the bla gene and IS5, and
within the -10 and -35 promoter boxes of the bla(CTX-M-32) gene.
Microbiological and biochemical studies revealed lower bla(CTX-M-32) gene
expression in bacterial isolates with IS1 between the promoter boxes.
CONCLUSIONS: Data suggest putative in vivo horizontal bla(CTX-M-32) gene
transfer between two different genera of Enterobacteriaceae. A new complex
structure, IS5-IS1, was detected upstream of the bla gene and IS1
negatively modulated expression of the bla(CTX-M-32) gene because its
location modified the bla promoter region.

<>

<1>Fernandez, M., Olek, A., Walter, J., Sanchez, J.
<2>Analysis of DNA methylation processes related to the inhibition of DNA synthesis by 5-azacytidine in Streptomyces antibioticus ETH 7451.
<3>Biol. Chem.
<4>379
<5>559-562
<6>1998
<7>5-Azacytidine inhibits DNA synthesis and to a lesser proportion RNA synthesis in S.
antibioticus.  The biosynthesis of proteins is not affected.  The main inhibitory effect of
5-azacytidine on DNA and RNA synthesis is probably caused by its incorporation into newly
synthesized DNA or RNA and the formation of covalent complexes between cytosine-specific
methyltransferases and the modified DNA or RNA templates.  To analyze whether such effects
could occur at the oriC region of S. antibioticus we analyzed the methylation status of this
region using the bisulphite assisted genomic sequencing method.  One of the cytosine residues
found to be partially methylated was contained within a unique NaeI sequence (GCCGGC) in oriC.
Subsequent analysis shows chromosomal DNA from S. antibioticus to be resistant to R.NaeI
restriction indicating that this strain contains a NaeI-specific cytosine C5-methyltransferase
activity.  Following 5-azacytidine treatment the NaeI site within the oriC region becomes
partially demethylated.  Our results suggest that some of the 5-azacytidine effects on DNA and
RNA synthesis might indeed be related to the complex formation and inhibition of a
cytosine-specific DNA methyltransferase.

<>

<1>Fernandez, M., Soliveri, J., Novella, I.S., Yebra, M.J., Barbes, C., Sanchez, J.
<2>Effect of  5-azacytidine and sinefungin on Streptomyces development.
<3>Gene
<4>157
<5>221-223
<6>1995
<7>The effect of two DNA-methyltransferase inhibitors, 5-azacytidine (5azaC) and sinefungin (Sf),
on the development of Streptomyces antibioticus ETH7451 (Sa) was studied. Pulse labeling
experiments and SDS-PAGE analysis of proteins from cells grown in sporulation synthetic medium
showed that both inhibitors affect a limited number of systems. Synthesis of the antibiotic
rhodomycin was increased in the presence of 5azaC. 5azaC also stimulated the production of
actinorhodin in cultures of S. coelicolor A3(2) grown in minimal medium. The analog did not
affect the expression of whiB and whiG, two sporulation genes from S. coelicolor A3(2) whose
homologues are present in Sa. Overall results indicated that 5azaC and Sf affect specific
events associated with differentiation and secondary metabolism in Streptomyces.

<>

<1>Fernandez-de-Las-Heras, L., Alonso, S., de la Vega-de-Leon, A., Xavier, D., Perera, J., Navarro-Llorens, J.M.
<2>Draft Genome Sequence of the Steroid Degrader Rhodococcus ruber Strain Chol-4.
<3>Genome Announcements
<4>1
<5>e00215-13
<6>2013
<7>The whole-genome shotgun sequence of Rhodococcus ruber strain Chol-4 is presented here. This
organism was shown to be able to grow using many steroids as the sole carbon and energy
sources. These sequence data will help us to further explore the metabolic abilities of this
versatile degrader.

<>

<1>Ferreira, A.C., Tenreiro, R., Correa-de-Sa, M.I., Dias, R.
<2>Complete Genome Sequences of Two Central European Brucella suis bv. 2 Haplotype 2c Strains Isolated from Wild Boars.
<3>Genome Announcements
<4>2
<5>e00686-14
<6>2014
<7>The Brucella suis haplotype 2c is commonly isolated from wild boars and domestic  pigs across
Central Europe, though it is rarely described in the Iberian Peninsula. We report here the
complete and annotated genome sequences of two haplotype 2c strains isolated from wild boars
in the northeast region of Spain, above the Ebro River.

<>

<1>Ferreira, A.C., Tenreiro, R., Correa-de-Sa, M.I., Dias, R.
<2>Complete Genome Sequences of Three Iberian Brucella suis Biovar 2 Strains Isolated from Wild Boars.
<3>Genome Announcements
<4>2
<5>e00618-14
<6>2014
<7>Brucella suis biovar 2 is the most common biovar isolated from wild boars (Sus scrofa)
associated with transmission to outdoor-reared pigs in Europe. We report here the complete and
annotated genome sequences of three strains isolated from wild boars in Portugal and Spain and
belonging to the Iberian clone (haplotypes 2d and 2e).

<>

<1>Ferreras, E.R., De Maayer, P., Makhalanyane, T.P., Guerrero, L.D., Aislabie, J.M., Cowan, D.A.
<2>Draft Genome Sequence of Microbacterium sp. Strain CH12i, Isolated from Shallow Groundwater in Cape Hallett, Antarctica.
<3>Genome Announcements
<4>2
<5>e00789-14
<6>2014
<7>The Antarctic continent is largely covered by an expansive ice sheet, but it harbors diverse
terrestrial and aquatic habitats in the coastal ice-free
continental margins. Here we present the draft genome of Microbacterium sp.
CH12i, which was isolated from hypersaline, alkaline, and nutrient-rich
groundwater from Cape Hallett, northern Victoria Land, Antarctica.

<>

<1>Ferretti, J.J. et al.
<2>Complete genome sequence of an M1 strain of Streptococcus pyogenes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>4658-4663
<6>2001
<7>The 1,852,442-bp sequence of an M1 strain of Streptococcus pyogenes, a Gram-positive pathogen,
has been determined and contains 1,752 predicted protein-encoding genes. Approximately
one-third of these genes have no identifiable function, with the remainder falling into
previously characterized categories of known microbial function. Consistent with the
observation that S. pyogenes is responsible for a wider variety of human disease than any
other bacterial species, more than 40 putative virulence-associated genes have been
identified. Additional genes have been identified that encode proteins likely associated with
microbial "molecular mimicry" of host characteristics and involved in rheumatic fever or acute
glomerulonephritis. The complete or partial sequence of four different bacteriophage genomes
is also present, with each containing genes for one or more previously undiscovered
superantigen-like proteins. These prophage-associated genes encode at least six potential
virulence factors, emphasizing the importance of bacteriophages in horizontal gene transfer
and a possible mechanism for generating new strains with increased pathogenic potential.

<>

<1>Ferrin, L.J., Camerini-Otero, R.D.
<2>Selective cleavage of human DNA: RecA-assisted restriction endonuclease (RARE) cleavage.
<3>Science
<4>254
<5>1494-1497
<6>1991
<7>Current methods for sequence-specific cleavage of large segments of DNA are
severely limited because of the paucity of possible cleavage sites.  A method
is described whereby any EcoRI site can be targeted for specific cleavage.  The
technique is based on the ability of RecA protein from Escherichia coli to pair
an oligonucleotide to its homologous sequence in duplex DNA and to form a
three-stranded complex.  This complex is protected from EcoRI methylase; after
methylation and RecA protein removal, EcoRI restriction enzyme cleavage was
limited to the site previously protected from methylation.  When pairs of
oligonucleotides are used, a specific fragment can be cleaved out of genomes.
The method was tested on lambda phage, Escherichia coli, and human DNA.
Fragments exceeding 500 kilobases in length and yields exceeding 80 percent
could be obtained.

<>

<1>Ferris, P.J., Vogt, V.M.
<2>Structure of the central spacer region of extrachromosomal ribosomal DNA in Physarum polycephalum.
<3>J. Mol. Biol.
<4>159
<5>359-381
<6>1982
<7>We have analyzed the sequence organization of the central spacer region of the
extrachromosomal ribosomal DNA from two strains of the acellular slime mold
Physarum polycephalum.  It had been inferred previously from electron
microscopy that this region, which comprises about one third of the 60 kb++
palindromic rDNA, contains a complex series of inverted repetitious sequences.
By partial digestion of end-labeled fragments isolated from purified rDNA and
from rDNA fragments cloned in Escherichia coli, we have constructed a detailed
restriction map of this region.  The 22 kb of spacer DNA of each half molecule
of rDNA contains the following elements:  (a) two separate regions, one of 1.1
kb and one of 2.1 kb, composed of many direct repeats of the same 30 base-pair
unit; (b) a region of 4.4 kb composed of a complex series of inverted repeats
of a 310 base-pair unit; (c) another region of 1.6 kb composed of inverted
repeats of the same 310 base-pair unit located directly adjacent to the center
of the rDNA; (d) two copies of a unique sequence of 0.85 kb, which probably
cocntains a replication origin.  Some of the CpG sequences in the spacer resist
cleavage by certain restriction endonucleases and thus appear to be methylated.
The lack of perfect symmetry about the central axis and the arrangement of
inverted repeated sequences explain the complex pattern of branches and forks
of the fold-back molecules previously observed by electron microscopy.
Comparison of the rDNA restriction maps from the two strains of Physarum
suggests that the repeat units in the spacer are undergoing concerted
evolution.  We propose a model to explain the evolutionary origin of the
several palindromic axes in the Physarum rDNA spacer.

<>

<1>Ferrucci, L., Rossino, R., Mezzanotee, R.
<2>Factors affecting the digestion of restriction endonucleases in situ.
<3>Cytobios
<4>68
<5>45-51
<6>1991
<7>The influence of incubation buffers and glycerol on the enzyme activity of naked DNA of lambda
phage and mouse, and of mouse chromosomal DNA was investigated. The results obtained varied in
part from previously known data, but confirmed the importance of these factors in determining
the patterns of in situ restriction enzyme digestion so far attributed exclusively to
endonuclease activity.

<>

<1>Fettweis, J.M., Serrano, M.G., Huang, B., Brooks, J.P., Glascock, A.L., Sheth, N.U., Strauss, J.F. III, Jefferson, K.K., Buck, G.A.
<2>An emerging Mycoplasma associated with trichomoniasis, vaginal infection and disease.
<3>PLoS ONE
<4>9
<5>E110943
<6>2014
<7>Humans are colonized by thousands of bacterial species, but it is difficult to
assess the metabolic and pathogenic potential of the majority of these because
they have yet to be cultured. Here, we characterize an uncultivated vaginal
mycoplasma tightly associated with trichomoniasis that was previously known by
its 16S rRNA sequence as "Mnola." In this study, the mycoplasma was found almost
exclusively in women infected with the sexually transmitted pathogen Trichomonas
vaginalis, but rarely observed in women with no diagnosed disease. The genomes of
four strains of this species were reconstructed using metagenome sequencing and
assembly of DNA from four discrete mid-vaginal samples, one of which was obtained
from a pregnant woman with trichomoniasis who delivered prematurely. These
bacteria harbor several putative virulence factors and display unique metabolic
strategies. Genes encoding proteins with high similarity to potential virulence
factors include two collagenases, a hemolysin, an O-sialoglycoprotein
endopeptidase and a feoB-type ferrous iron transport system. We propose the name
"Candidatus Mycoplasma girerdii" for this potential new pathogen.

<>

<1>Fevre, C., Passet, V., Deletoile, A., Barbe, V., Frangeul, L., Almeida, A.S., Sansonetti, P., Tournebize, R., Brisse, S.
<2>PCR-based identification of Klebsiella pneumoniae subsp. rhinoscleromatis, the agent of rhinoscleroma.
<3>PLoS Neglected Trop. Dis.
<4>5
<5>E1052
<6>2011
<7>Rhinoscleroma is a chronic granulomatous infection of the upper airways caused by
the bacterium Klebsiella pneumoniae subsp. rhinoscleromatis. The disease is
endemic in tropical and subtropical areas, but its diagnosis remains difficult.
As a consequence, and despite available antibiotherapy, some patients evolve
advanced stages that can lead to disfiguration, severe respiratory impairment and
death by anoxia. Because identification of the etiologic agent is crucial for the
definitive diagnosis of the disease, the aim of this study was to develop two
simple PCR assays. We took advantage of the fact that all Klebsiella pneumoniae
subsp. rhinoscleromatis isolates are (i) of capsular serotype K3; and (ii) belong
to a single clone with diagnostic single nucleotide polymorphisms (SNP). The
complete sequence of the genomic region comprising the capsular polysaccharide
synthesis (cps) gene cluster was determined. Putative functions of the 21 genes
identified were consistent with the structure of the K3 antigen. The K3-specific
sequence of gene Kr11509 (wzy) was exploited to set up a PCR test, which was
positive for 40 K3 strains but negative when assayed on the 76 other Klebsiella
capsular types. Further, to discriminate Klebsiella pneumoniae subsp.
rhinoscleromatis from other K3 Klebsiella strains, a specific PCR assay was
developed based on diagnostic SNPs in the phosphate porin gene phoE. This work
provides rapid and simple molecular tools to confirm the diagnostic of
rhinoscleroma, which should improve patient care as well as knowledge on the
prevalence and epidemiology of rhinoscleroma.

<>

<1>Ffrench-Constant, R.H., Waterfield, N., Burland, V., Perna, N.T., Daborn, P.J., Bowen, D., Blattner, F.R.
<2>A genomic sample sequence of the entomopathogenic bacterium Photorhabdus luminescens W14: Potential implications for virulence.
<3>Appl. Environ. Microbiol.
<4>66
<5>3310-3329
<6>2000
<7>Photorhabdus luminescens is a pathogenic bacterium that lives in the guts of insect-pathogenic
nematodes. After invasion of an insect host
by a nematode, bacteria are released from the nematode gut and help
kill the insect, in which both the bacteria and the nematodes
subsequently replicate. However, the bacterial virulence factors
associated with this 'symbiosis of pathogens' remain largely obscure.
In order to identify genes encoding potential virulence factors, we
performed approximately 2,000 random sequencing reads from a P.
luminescens W14 genomic library. We then compared the sequences
obtained to sequences in existing gene databases and to the Escherichia
coli K-12 genome sequence. Here we describe the different classes of
potential virulence factors found. These factors include genes that
putatively encode Tc insecticidal toxin complexes, Rbi-like toxins,
proteases and lipases, colicin and pyocins, and various antibiotics.
They also include a diverse array of secretion (e.g., type III), iron
uptake, and lipopolysaccharide production systems. We speculate on the
potential functions of each of these gene classes in insect infection
and also examine the extent to which the invertebrate pathogen P,
luminescens shares potential antivertebrate virulence factors. The
implications for understanding both the biology of this insect pathogen
and links between the evolution of vertebrate virulence factors and the
evolution of invertebrate virulence factors are discussed.

<>

<1>Fiala, E.S., Staretz, M.E., Pandya, G., El-Bayoumy, K., Hamilton, S.R.
<2>Inhibition of DNA (cytosine-5) methyltransferase (Mtase) by selenium compounds, determined by an improved method for Mtase and global DNA methylation.
<3>Proc. Amer. Assoc. Cancer Res.
<4>39
<5>88
<6>1998
<7>Organoselenium compounds such as benzyl selenocyanate and
1,4-phenylenebis(methylene)selenocyanate are efficient chemopreventive agents, inhibiting a
variety of chemically induced tumors in animal models at both the initiation and
postinitiation stages.  Because several lines of evidence suggest that inhibition of Mtase in
tumor cells may be a sufficient condition for the suppression or reversion of carcinogenesis,
we examined the effects of sodium selenite, BSC, p-XSC and benzyl thiocyanate on Mtase
activity in extracts from human colon carcinomas, and in HCT116 human colon carcinoma cells in
culture.  For this purpose, we developed an improved assay of the enzyme, in which label
derived from S-adenosyl[methyl-3H]methionine is specifically determined in
5-methyldeoxycytidine by HPLC with radioflow detection.  Selenite, BSC and p-XSC inhibited
Mtase extracted from a human colon carcinoma with IC50s of 3.8, 8.1, and 5.2 uM, respectively.
BTC had no effect.  Selenite, BSC and p-XSC also strongly inhibited the growth and Mtase
activity of HCT116 cells.  We suggest that inhibition of Mtase may be central to the mechanism
of chemoprevention by selenium compounds at the stage of postinitiation.

<>

<1>Fiala, E.S., Staretz, M.E., Pandya, G.A., El-Bayoumy, K., Hamilton, S.R.
<2>Inhibition of DNA cytosine methyltransferase by chemopreventive selenium compounds, determined by an improved assay for DNA cytosine methyltransferase and DNA cytosine methylation.
<3>Carcinogenesis
<4>19
<5>597-604
<6>1998
<7>The organoselenium compounds benzyl selenocyanate and 1,4-phenylenebis(methylene)selenocyanate
as well as sodium selenite, are effective chemopreventive agents for various chemically
induced tumors in animal models at both the initiation and postinitiation stages.  The
mechanisms involved at the postinitiation stage are not clear.  Because several lines of
evidence indicate that inhibition of excess DNA (cytosine-5)-methyltransferase may be a
sufficient factor for the suppression or reversion of carcinogenesis, we examined the effects
of sodium selenite, Bsc, p-XSC and benzyl thiocyanate, the sulfur analog of BSC, on Mtase
activity in nuclear extracts of human colon carcinomas, and of p-XSC on the Mtase activity of
HCT116 human colon carcinoma cells in culture.  For this purpose, we developed an improved
Mtase assay, in which the incorporation of the methyl-[3-H] group from
S-adenosyl[methyl-3H]methionine into deoxycytidine of poly(dI-dC)-poly(dI-dC), is specifically
determined by HPLC with radioflow detection after enzymatic hydrolysis, enhancing specificity
and reliability.  In a variation, using SssI methyltransferase and labeled
S-adenosylmethionine, the overall methylation status of DNA in various tissues can also be
compared. Selenite, BSC and p-XSC inhibited Mtase extracted from a human colon carcinoma with
IC50S of 3.8, 8.1 and 5.2 uM, respectively; BTC had no effect.  P-XSC also inhibited the Mtase
activity and growth of human colon carcinoma HCT116 cells, with an IC50 of ~20 uM.  The
improved Mtase assay should prove to be a reliable method for screening potential Mtase
inhibitors, especially using cells in culture.  We suggest that inhibition of Mtase may be a
major mechanism of chemoprevention by selenium compounds at the postinitiation stage of
carcinogenesis.

<>

<1>Fiebig, A., Pradella, S., Petersen, J., Michael, V., Pauker, O., Rohde, M., Goker, M., Klenk, H.P., Wagner-Dobler, I.
<2>Genome of the marine alphaproteobacterium Hoeflea phototrophica type strain (DFL-43(T)).
<3>Standards in Genomic Sciences
<4>7
<5>440-448
<6>2013
<7>Hoeflea phototrophica Biebl et al. 2006 is a member of the family Phyllobacteriaceae in the
order Rhizobiales, which is thus far only partially
characterized at the genome level. This marine bacterium contains the
photosynthesis reaction-center genes pufL and pufM and is of interest because it
lives in close association with toxic dinoflagellates such as Prorocentrum lima.
The 4,467,792 bp genome (permanent draft sequence) with its 4,296 protein-coding
and 69 RNA genes is a part of the Marine Microbial Initiative.

<>

<1>Fiebig, A., Pradella, S., Petersen, J., Pauker, O., Michael, V., Lunsdorf, H., Goker, M., Klenk, H.P., Wagner-Dobler, I.
<2>Genome of the R-body producing marine alphaproteobacterium Labrenzia alexandrii type strain (DFL-11(T)).
<3>Standards in Genomic Sciences
<4>7
<5>413-426
<6>2013
<7>Labrenzia alexandrii Biebl et al. 2007 is a marine member of the family Rhodobacteraceae in
the order Rhodobacterales, which has thus far only partially
been characterized at the genome level. The bacterium is of interest because it
lives in close association with the toxic dinoflagellate Alexandrium lusitanicum.
Ultrastructural analysis reveals R-bodies within the bacterial cells, which are
primarily known from obligate endosymbionts that trigger 'killing traits' in
ciliates (Paramecium spp.). Genomic traits of L. alexandrii DFL-11(T) are in
accordance with these findings, as they include the reb genes putatively involved
in R-body synthesis. Analysis of the two extrachromosomal elements suggests a
role in heavy-metal resistance and exopolysaccharide formation, respectively. The
5,461,856 bp long genome with its 5,071 protein-coding and 73 RNA genes consists
of one chromosome and two plasmids, and has been sequenced in the context of the
Marine Microbial Initiative.

<>

<1>Fiebig, A., Riedel, T., Gronow, S., Petersen, J., Klenk, H.P., Goker, M.
<2>Genome sequence of the reddish-pigmented Rubellimicrobium thermophilum type strain (DSM 16684(T)), a member of the Roseobacter clade.
<3>Standards in Genomic Sciences
<4>8
<5>480-490
<6>2013
<7>Rubellimicrobium thermophilum Denner et al. 2006 is the type species of the genus
Rubellimicrobium, a representative of the Roseobacter clade within the
Rhodobacteraceae. Members of this clade were shown to be abundant especially in
coastal and polar waters, but were also found in microbial mats and sediments.
They are metabolically versatile and form a physiologically heterogeneous group
within the Alphaproteobacteria. Strain C-Ivk-R2A-2(T) was isolated from colored
deposits in a pulp dryer; however, its natural habitat is so far unknown. Here we
describe the features of this organism, together with the draft genome sequence
and annotation and novel aspects of its phenotype. The 3,161,245 bp long genome
contains 3,243 protein-coding and 45 RNA genes.

<>

<1>Fiedoruk, K., Daniluk, T., Swiecicka, I., Murawska, E., Sciepuk, M., Leszczynska, K.
<2>First Complete Genome Sequence of Escherichia albertii Strain KF1, a New Potential Human Enteric Pathogen.
<3>Genome Announcements
<4>2
<5>e00004-14
<6>2014
<7>Escherichia albertii has been recently recognized as an emerging human and bird enteric
pathogen. Here, we report the first complete chromosome nucleotide
sequence of a clinical isolate of E. albertii strain KF1, which may provide
information about the pathogenic potential of this new species and the mechanisms
of evolution of enteropathogenic Escherichia spp.

<>

<1>Field, D. et al.
<2>The minimum information about a genome sequence (MIGS) specification.
<3>Nat. Biotechnol.
<4>26
<5>541-547
<6>2008
<7>With the quantity of genomic data increasing at an exponential rate, it is imperative that
these data be captured electronically, in a standard
format. Standardization activities must proceed within the auspices of
open-access and international working bodies. To tackle the issues
surrounding the development of better descriptions of genomic
investigations, we have formed the Genomic Standards Consortium (GSC).
Here, we introduce the minimum information about a genome sequence (MIGS)
specification with the intent of promoting participation in its
development and discussing the resources that will be required to develop
improved mechanisms of metadata capture and exchange. As part of its wider
goals, the GSC also supports improving the 'transparency' of the
information contained in existing genomic databases.

<>

<1>Fierer, N., Breitbart, M., Nulton, J., Salamon, P., Lozupone, C., Jones, R., Robeson, M., Edwards, R.A., Felts, B., Rayhawk, S., Knight, R., Rohwer, F., Jackson, R.B.
<2>Metagenomic and small-subunit rRNA analyses reveal the genetic diversity of bacteria, archaea, fungi, and viruses in soil.
<3>Appl. Environ. Microbiol.
<4>73
<5>7059-7066
<6>2007
<7>Recent studies have highlighted the surprising richness of soil bacterial
communities; however, bacteria are not the only microorganisms found in
soil. To our knowledge, no study has compared the diversities of the four
major microbial taxa, i.e., bacteria, archaea, fungi, and viruses, from an
individual soil sample. We used metagenomic and small-subunit RNA-based
sequence analysis techniques to compare the estimated richness and
evenness of these groups in prairie, desert, and rainforest soils. By
grouping sequences at the 97% sequence similarity level (an operational
taxonomic unit [OTU]), we found that the archaeal and fungal communities
were consistently less even than the bacterial communities. Although total
richness levels are difficult to estimate with a high degree of certainty,
the estimated number of unique archaeal or fungal OTUs appears to rival or
exceed the number of unique bacterial OTUs in each of the collected soils.
In this first study to comprehensively survey viral communities using a
metagenomic approach, we found that soil viruses are taxonomically diverse
and distinct from the communities of viruses found in other environments
that have been surveyed using a similar approach. Within each of the four
microbial groups, we observed minimal taxonomic overlap between sites,
suggesting that soil archaea, bacteria, fungi, and viruses are globally as
well as locally diverse.

<>

<1>Figueiredo, C., Quint, W.G., Sanna, R., Sablon, E., Donahue, J.P., Xu, Q., Miller, G.G., Peek, R.M., Blaser, M.J., van Doorn, L.
<2>Genetic organization and heterogeneity of the iceA locus of Helicobacter pylori.
<3>Gene
<4>246
<5>59-68
<6>2000
<7>The genetic organization and sequence heterogeneity of the iceA locus of Helicobacter pylori
was studied, and the existence of two distinct gene families, iceA1 and iceA2, at this locus
was confirmed. iceA1 has significant sequence homology to nlaIIIR, encoding an endonuclease in
Neisseria lactamica, but the similarity at the protein level is limited, due to frameshift
mutations of iceA1 in most H. pylori strains. In only five of the 19 iceA1 strains studied, a
full-length open reading frame (ORF), capable of encoding a 228aa protein, with 52% homology
to NlaIII was observed. The region upstream of iceA2 is highly variable in length, containing
up to 15 copies of 8bp tandem repeats. iceA2 can encode proteins of 24, 59, 94, or 129 amino
acids, consisting of 14 and 10aa domains, conserved in all iceA2 strains, flanking 0, 1, 2, or
3 copies of a 35aa cassette. This 35aa cassette consists of domains of 13, 16 and 6aa,
respectively. The 13aa and 6aa domains are highly conserved, but the 16aa domain exists in two
variants. In total, five distinct iceA2 subtypes were defined. Database searches did not
reveal any homologous sequences. Recombinant IceA1 and IceA2 proteins were expressed in
Escherichia coli, confirming the predicted ORFs. Genotype-specific PCR primers permitted iceA
genotyping in 318 (99.1%) of a worldwide collection of 321 H. pylori strains. The conserved
sizes of the amplification products confirmed the worldwide distribution of discrete variants
of iceA1 and iceA2.

<>

<1>Figueiredo, H.C., Leal, C.A., Dorella, F.A., Carvalho, A.F., Soares, S.C., Pereira, F.L., Azevedo, V.A.
<2>Complete Genome Sequences of Fish Pathogenic Weissella ceti Strains WS74 and WS105.
<3>Genome Announcements
<4>2
<5>e01014-14
<6>2014
<7>We describe here the genome sequencing and annotation of Weissella ceti strains WS74 and
WS105, isolated from diseased rainbow trout in Brazil. The two genomes
were sequenced with an Ion Torrent personal genome machine (PGM) using a fragment
library. The genomes of strains WS74 and WS105 consist of circular chromosomes
1,389,513 bp and 1,390,396 bp long, respectively, both presenting a G+C content
of 40.75%.

<>

<1>Figueiredo, H.C., Leal, G., Pereira, F.L., Soares, S.C., Dorella, F.A., Carvalho, A.F., Pereira, U.P., Azevedo, V.A.
<2>Whole-Genome Sequence of Weissella ceti Strain WS08, Isolated from Diseased Rainbow Trout in Brazil.
<3>Genome Announcements
<4>2
<5>e00851-14
<6>2014
<7>We report here the complete genome sequence of Weissella ceti strain WS08, an emerging
pathogen to farm-raised rainbow trout. The genome of strain WS08 is
composed of a circular chromosome with 1,355,853 bp and a G+C content of 40.78%.

<>

<1>Figueiredo, S.C., Neves-Borges, A.C., Coelho, A.
<2>The neuraminidase gene is present in the non-toxigenic Vibrio cholerae Amazonia strain: a different allele in comparison to the pandemic strains.
<3>Memorias do Instituto Oswaldo Cruz
<4>100
<5>563-569
<6>2005
<7>The neuraminidase gene, nanH, is present in the O1, non-toxigenic Vibrio
cholerae Amazonia strain. Its location has been assigned to a 150 kb NotI
DNA fragment, with the use of pulsed-field gel electrophoresis and DNA
hybridization. This NotI fragment is positioned inside 630 kb SfiI and
1900 kb I-CeuI fragments of chromosome 1. Association of the pathogenicity
island VPI-2, carrying nanH and other genes, with toxigenic strains has
been described by other authors. The presence of nanH in a non-toxigenic
strain is an exception to this rule. The Amazonia strain nanH was
sequenced (Genbank accession No. AY825932) and compared to available V.
cholerae sequences. The sequence is different from those of pandemic
strains, with 72 nucleotide substitutions. This is the first description
of an O1 strain with a different nanH allele. The most variable domain of
the Amazonia NanH is the second lectin wing, comprising 13 out of 17 amino
acid substitutions. Based on the presence of nanH in the same region of
the genome, and similarity of the adjacent sequences to VPI-2 sequences,
it is proposed that the pathogenicity island VPI-2 is present in this
strain.

<>

<1>Figueiredo, T.A., Aguiar, S.I., Melo-Cristino, J., Ramirez, M.
<2>DNA methylase activity as a marker for the presence of a family of phage-like elements conferring efflux-mediated macrolide resistance in  streptococci.
<3>Antimicrob. Agents Chemother.
<4>50
<5>3689-3694
<6>2006
<7>Recently, two related chimeric genetic elements (Tn1207.3 and Phi10394.4) were shown to carry
the macrolide efflux gene mef in Streptococcus
pyogenes (group A streptococci [GAS]). The dissemination of elements
belonging to the Tn1207.3/Phi10394.4 family in recent isolates of GAS,
Streptococcus dysgalactiae subsp. equisimilis, Streptococcus pneumoniae,
and Streptococcus agalactiae recovered in Portugal was surveyed. In total,
149 GAS, 18 S. pneumoniae, 4 S. dysgalactiae subsp. equisimilis, and 5 S.
agalactiae isolates from infections, presenting the M phenotype of
macrolide resistance and containing the mef gene, were screened for the
presence of Tn1207.3/Phi10394.4 by PCR targeting open reading frames
(ORFs) specific for these related elements. All the GAS isolates tested
and one of the S. dysgalactiae subsp. equisimilis isolates carried
Tn1207.3. However, neither of these elements was found in the isolates of
the other streptococcal species. It was also noted that the DNAs of the
isolates carrying Tn1207.3 were resistant to cleavage by the endonuclease
SmaI. Cloning and expression of ORF12 of Tn1207.3 in Escherichia coli
showed that it encoded a methyltransferase that rendered DNA refractory to
cleavage by SmaI (M.Spy10394I). Using this characteristic as a marker for
the presence of the Tn1207.3/Phi10394.4 family, we reviewed the literature
and concluded that these genetic elements are widely distributed among
tetracycline-susceptible GAS isolates presenting the M phenotype from
diverse geographic origins and may have played an important role in the
dissemination of macrolide resistance in this species.

<>

<1>Filipchenkova, L.P., Kosykh, V.G., Buryanov, Y.I., Baev, A.A., Ansberga, S.E.
<2>Purification of restriction endonuclease EcoRII from Escherichia coli B834/pSK323.
<3>Soviet Patent Office
<4>SU 1321060
<5>
<6>1994
<7>
<>

<1>Filippini, M., Qi, W., Blom, J., Goesmann, A., Smits, T.H., Bagheri, H.C.
<2>Genome Sequence of Fibrella aestuarina BUZ 2T, a Filamentous Marine Bacterium.
<3>J. Bacteriol.
<4>194
<5>3555
<6>2012
<7>Fibrella aestuarina BUZ 2(T) is the type strain of the recently characterized genus Fibrella.
Here we report the draft genome sequence of this strain, which
consists of a single scaffold representing the chromosome (with 11 gaps) and a
161-kb circular plasmid.

<>

<1>Filippini, M., Qi, W., Jaenicke, S., Goesmann, A., Smits, T.H., Bagheri, H.C.
<2>Genome Sequence of the Filamentous Bacterium Fibrisoma limi BUZ 3T.
<3>J. Bacteriol.
<4>194
<5>4445
<6>2012
<7>Fibrisoma limi strain BUZ 3(T), a Gram-negative bacterium, was isolated from coastal mud from
the North Sea (Fedderwardersiel, Germany) and characterized
using a polyphasic approach in 2011. The genome consists of a chromosome of about
7.5 Mb and three plasmids.

<>

<1>Finan, T.M., Weidner, S., Wong, K., Buhrmester, J., Chain, P., Vorholter, F.J., Hernandez-Lucas, I., Becker, A., Cowie, A., Gouzy, J., Golding, B., Puhler, A.
<2>The complete sequence of the 1,683-kb pSymB megaplasmid from the N2-fixing endosymbiont Sinorhizobium meliloti.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>9889-9894
<6>2001
<7>Analysis of the 1,683,333-nt sequence of the pSymB megaplasmid from the symbiotic N(2)-fixing
bacterium Sinorhizobium meliloti revealed that the
replicon has a high gene density with a total of 1,570 protein-coding
regions, with few insertion elements and regions duplicated elsewhere in
the genome. The only copies of an essential arg-tRNA gene and the minCDE
genes are located on pSymB. Almost 20% of the pSymB sequence carries genes
encoding solute uptake systems, most of which were of the ATP-binding
cassette family. Many previously unsuspected genes involved in
polysaccharide biosynthesis were identified and these, together with the
two known distinct exopolysaccharide synthesis gene clusters, show that
14% of the pSymB sequence is dedicated to polysaccharide synthesis. Other
recognizable gene clusters include many involved in catabolic activities
such as protocatechuate utilization and phosphonate degradation. The
functions of these genes are consistent with the notion that pSymB plays a
major role in the saprophytic competence of the bacteria in the soil
environment.

<>

<1>Fineran, P.C., Iglesias, C.M.C., Ramsay, J.P., Wilf, N.M., Cossyleon, D., McNeil, M.B., Williamson, N.R., Monson, R.E., Becher, S.A., Stanton, J.A., Brugger, K., Brown, S.D., Salmond, G.P.
<2>Draft Genome Sequence of Serratia sp. Strain ATCC 39006, a Model Bacterium for Analysis of the Biosynthesis and Regulation of Prodigiosin, a Carbapenem, and Gas  Vesicles.
<3>Genome Announcements
<4>1
<5>e01039-13
<6>2013
<7>Serratia sp. strain ATCC 39006 is a Gram-negative bacterium and a member of the
Enterobacteriaceae that produces various bioactive secondary metabolites,
including the tripyrrole red pigment prodigiosin and the beta-lactam antibiotic
1-carbapenen-2-em-3-carboxylic acid (a carbapenem). This strain is the only
member of the Enterobacteriaceae known to naturally produce gas vesicles, as
flotation organelles. Here we present the genome sequence of this strain, which
has served as a model for analysis of the biosynthesis and regulation of
antibiotic production.

<>

<1>Finkbeiner, S.R., Allred, A.F., Tarr, P.I., Klein, E.J., Kirkwood, C.D., Wang, D.
<2>Metagenomic analysis of human diarrhea: viral detection and discovery.
<3>PLoS Pathog.
<4>4
<5>e1000011
<6>2008
<7>Worldwide, approximately 1.8 million children die from diarrhea annually,
and millions more suffer multiple episodes of nonfatal diarrhea. On
average, in up to 40% of cases, no etiologic agent can be identified. The
advent of metagenomic sequencing has enabled systematic and unbiased
characterization of microbial populations; thus, metagenomic approaches
have the potential to define the spectrum of viruses, including novel
viruses, present in stool during episodes of acute diarrhea. The detection
of novel or unexpected viruses would then enable investigations to assess
whether these agents play a causal role in human diarrhea. In this study,
we characterized the eukaryotic viral communities present in diarrhea
specimens from 12 children by employing a strategy of "micro-mass
sequencing" that entails minimal starting sample quantity (<100 mg stool),
minimal sample purification, and limited sequencing (384 reads per
sample). Using this methodology we detected known enteric viruses as well
as multiple sequences from putatively novel viruses with only limited
sequence similarity to viruses in GenBank.

<>

<1>Finnegan, E.J., Dennis, E.S.
<2>Isolation and identification by sequence homology of a putative cytosine methyltransferase from Arabidopsis thaliana.
<3>Nucleic Acids Res.
<4>21
<5>2383-2388
<6>1993
<7>A plant cytosine methyltransferase cDNA was isolated using degenerate oligonucleotides, based
on homology between prokaryote and mouse methyltransferases, and PCR to amplify a short
fragment of a methyltansferase gene. A fragment of the predicted size was amplified from
genomic DNA from Arabidopsis thaliana. Overlapping cDNA clones, some with homology to the PCR
amplifed fragment, were identified and sequenced. The assembled nucleic acid sequence is 4720
bp and encodes a protein of 1534 amino acids which has significant homology to prokaryote and
mammalian cytosine methyltransferases. Like mammalian methylases, this enzyme has a C terminal
methyltransferase domain linked to a second larger domain. The Arabidopsis methylase has eight
of the ten conserved sequence motifs found in prokaryote cytosine-5 methyltransferases and
shows 50% homology to the murine enzyme in the methyltransferase domain. The amino terminal
domain is only 24% homologous to the murine enzyme and lacks the zinc binding region that has
been found in methyltransferases from both mouse and man. In contrast to mouse where a single
methyltransferase gene has been identified, a small multigene family with homology to the
region amplified in PCR has been identified in Arabidopsis thaliana.

<>

<1>Finnegan, E.J., Genger, R.K., Peacock, W.J., Dennis, E.S.
<2>DNA methylation in plants.
<3>Annu. Rev. Plant Physiol. Plant Mol. Biol.
<4>49
<5>223-247
<6>1998
<7>Methylation of cytosine residues in DNA provides a mechanism of gene control.  There are two
classes of methyltransferase in Arabidopsis; one has a carboxy-terminal methyltransferase
domain fused to an amino-terminal regulatory domain and is similar to mammalian
methyltransferases.  The second class apparently lacks an amino-terminal domain and is less
well conserved.  Methylcytosine can occur at any cytosine residue, but it is likely that
clonal transmission of methylation patterns only occurs for cytosines in strand-symmetrical
sequences CpG and CpNpG.  In plants, as in mammals, DNA methylation has dual roles in defense
against invading DNA and transposable elements and in gene regulation.  Although originally
reported as having no phenotypic consequence, reduced DNA methylation disrupts normal plant
development.

<>

<1>Finnegan, E.J., Kovac, K.A.
<2>Plant DNA methyltransferases.
<3>Plant Mol. Biol.
<4>43
<5>189-201
<6>2000
<7>DNA methylation is an important modification of DNA that plays a role in genome management and
in regulating gene expression during
development. Methylation is carried out by DNA methyltransferases which
catalyse the transfer of a methyl group to bases within the DNA helix.
Plants have at least three classes of cytosine methyltransferase which
differ in protein structure and function. The METI family, homologues
of the mouse Dnmt1 methyltransferase, most likely function as
maintenance methyltransferases, but may also play a role in de novo
methylation. The chromomethylases, which are unique to plants, may
preferentially methylate DNA in heterochromatin; the remaining class,
with similarity to Dnmt3 methyltransferases of mammals, are putative de
novo methyltransferases. The various classes of methyltransferase may
show differential activity on cytosines in different sequence contexts.
Chromomethylases may preferentially methylate cytosines in CpNpG
sequences while the Arabidopsis METI methyltransferase shows a
preference for cytosines in CpG sequences. Additional proteins, for
example DDM1, a member of the SNF2/SWI2 family of chromatin remodelling
proteins, are also required for methylation of plant DNA.

<>

<1>Finnegan, E.J., Peacock, W.J., Dennis, E.S.
<2>Reduced DNA methylation in Arabidopsis thaliana results in abnormal plant development.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>8449-8454
<6>1996
<7>Arabidopsis plants transformed with an antisense construct of an Arabidopsis methyltransferase
cDNA (METI) have reduced cytosine methylation in CG dinucleotides. Methylation levels in
progeny of five independent transformants ranged from 10% to 100% of the wild type. Removal of
the antisense construct by segregation in sexual crosses did not fully restore methylation
patterns in the progeny, indicating that methylation patterns are subject to meiotic
inheritance in Arabidopsis. Plants with decreased methylation displayed a number of phenotypic
and developmental abnormalities, including reduced apical dominance, smaller plant size,
altered leaf size and shape, decreased fertility, and altered flowering time. Floral organs
showed homeotic transformations that were associated with ectopic expression of the floral
homeotic genes AGAMOUS and APETALA3 in leaf tissue. These observations suggest that DNA
methylation plays an important role in regulating many developmental pathways in plants and
that the developmental abnormalities seen in the methyltransferase antisense plants may be due
to dysregulation of gene expression.

<>

<1>Finster, K.W., Kjeldsen, K.U., Kube, M., Reinhardt, R., Mussmann, M., Amann, R., Schreiber, L.
<2>Complete genome sequence of Desulfocapsa sulfexigens, a marine deltaproteobacterium specialized in disproportionating inorganic sulfur  compounds.
<3>Standards in Genomic Sciences
<4>8
<5>58-68
<6>2013
<7>Desulfocapsa sulfexigens SB164P1 (DSM 10523) belongs to the deltaproteobacterial  family
Desulfobulbaceae and is one of two validly described members of its genus.
This strain was selected for genome sequencing, because it is the first marine
bacterium reported to thrive on the disproportionation of elemental sulfur, a
process with a unresolved enzymatic pathway in which elemental sulfur serves both
as electron donor and electron acceptor. Furthermore, in contrast to its
phylogenetically closest relatives, which are dissimilatory sulfate-reducers, D.
sulfexigens is unable to grow by sulfate reduction and appears metabolically
specialized in growing by disproportionating elemental sulfur, sulfite or
thiosulfate with CO2 as the sole carbon source. The genome of D. sulfexigens
contains the set of genes that is required for nitrogen fixation. In an acetylene
assay it could be shown that the strain reduces acetylene to ethylene, which is
indicative for N-fixation. The circular chromosome of D. sulfexigens SB164P1
comprises 3,986,761 bp and harbors 3,551 protein-coding genes of which 78% have a
predicted function based on auto-annotation. The chromosome furthermore encodes
46 tRNA genes and 3 rRNA operons.

<>

<1>Finta, C., Kiss, A.
<2>Footprint analysis of the BspRI DNA methyltransferase -- DNA interaction.
<3>Nucleic Acids Res.
<4>25
<5>2841-2846
<6>1997
<7>The interaction between the GGCC-specific BspRI DNA methyltransferase (M.BspRI) and substrate
DNA was studied with footprinting techniques usiung a DNA fragment that was unmodified on both
strands.  Footprinting with DNase I revealed an ~14 bp protected region.  Footprinting with
dimethylsulfate detected major groove interactions with the guanine bases of the recognition
sequence.  Reaction with 1,10-phenanthroline-copper did not show protection, suggesting that
minor groove interactions play little role in sequence-specific recognition by M.BspRI.
Hydroxyl radical footprinting revealed a protected stretch of 6 nt.  The hydroxyl radical
footprint of M.BspRI differs markedly from the footprint reported for the HhaI and SssI
methyltransferases.  The pattern of protection from dimethylsulfate and hydroxyl radicals
suggests that the interactions of M.BspRI with DNA are similar to those detected in the
co-crystal structure of the HaeIII methyltransferase.

<>

<1>Finta, C., Sulima, U., Venetianer, P., Kiss, A.
<2>Purification of the KpnI DNA methyltransferase and photolabeling of the enzyme with S-adenosyl-L-methionine.
<3>Gene
<4>164
<5>65-69
<6>1995
<7>An Escherichia coli strain overproducing the KpnI DNA methyltransferase (M.KpnI) was
constructed by cloning the kpnIM gene downstream from the inducible T7 phage omega 10
promoter.  A method involving three chromatographic steps has been developed to purify M.KpnI
to homogeneity.  The purified enzyme has a pH optimum around 7.3 and is inhibited by salts.
M.KpnI can be photolabeled by UV-irradiation of the enzyme in the presence of
S-adenosyl-L-[methyl-3H]methionine ([methyl-3H]AdoMet).  Photolabeling results from a specific
interaction betweeen M.KpnI and AdoMet, as indicated by the dependence of photolabeling on
native enzyme conformation and by the inhibitory effect of the AdoMet analogs, sinefungin and
S-adenosyl-L-homocysteine (AdoHcy).

<>

<1>Fiore, M.F., Alvarenga, D.O., Varani, A.M., Hoff-Risseti, C., Crespim, E., Ramos, R.T., Silva, A., Schaker, P.D., Heck, K., Rigonato, J., Schneider, M.P., Jeong, H., Sim, Y.M., Kim, H.J., Lee, Y.J., Lee, D.W., Lim, S.K., Lee, S.J.
<2>Draft Genome Sequence of the Brazilian Toxic Bloom-Forming Cyanobacterium Microcystis aeruginosa Strain SPC777.
<3>Genome Announcements
<4>1
<5>e00547-13
<6>2013
<7>Microcystis aeruginosa strain SPC777 is an important toxin-producing cyanobacterium, isolated
from a water bloom of the Billings reservoir (Sao Paulo
State, Brazil). Here, we report the draft genome sequence and initial findings
from a preliminary analysis of strain SPC777, including several gene clusters
involved in nonribosomal and ribosomal synthesis of secondary metabolites.

<>

<1>Firczuk, M., Wojciechowski, M., Czapinska, H., Bochtler, M.
<2>DNA intercalation without flipping in the specific ThaI-DNA complex.
<3>Nucleic Acids Res.
<4>39
<5>744-754
<6>2011
<7>The PD-(D/E)XK type II restriction endonuclease ThaI cuts the target sequence CG/CG with blunt
ends. Here, we report the 1.3 A resolution
structure of the enzyme in complex with substrate DNA and a sodium or
calcium ion taking the place of a catalytic magnesium ion. The structure
identifies Glu54, Asp82 and Lys93 as the active site residues. This agrees
with earlier bioinformatic predictions and implies that the PD and (D/E)XK
motifs in the sequence are incidental. DNA recognition is very unusual:
the two Met47 residues of the ThaI dimer intercalate symmetrically into
the CG steps of the target sequence. They approach the DNA from the minor
groove side and penetrate the base stack entirely. The DNA accommodates
the intercalating residues without nucleotide flipping by a doubling of
the CG step rise to twice its usual value, which is accompanied by drastic
unwinding. Displacement of the Met47 side chains from the base pair
midlines toward the downstream CG steps leads to large and compensating
tilts of the first and second CG steps. DNA intercalation by ThaI is
unlike intercalation by HincII, HinP1I or proteins that bend or repair
DNA.

<>

<1>Firman, K.
<2>Polynucleotide motor, a motor system, their preparation and uses.
<3>US Patent Office
<4>US 20060160117 A
<5>
<6>2006
<7>A polynucleotide motor is disclosed, comprising an enzyme capable of binding to a duplex
nucleic acid sequence, which enzyme is also capable of translocating the nucleic acid sequence
without causing cleavage thereof.  The motor may be associated with a substance bound to the
nucleic acid sequence so that the bound substance, such as magnetic bead, biotin,
streptavidin, a scintillant or the like, can itself be translocated, relative to the region of
binding of the enzyme, during translocation.  The enzyme remains bound to the original
recognition site of the nucleic acid sequence.  Such a system has applications in screening or
testing for a pre-determined biological, chemical or physical activity; for example, in
screening for new pharmacologically-effective ligands.

<>

<1>Firman, K., Creasey, W.A., Watson, G., Price, C., Glover, S.W.
<2>Genetic and physical studies of restriction-deficient mutants of the Inc FIV plasmids R124 and R124/3.
<3>Mol. Gen. Genet.
<4>191
<5>145-153
<6>1983
<7>R124 and R124/3 are R plasmids that carry the genes for two different
restriction and modification systems.  The phenotype of strains carrying either
of these plasmids along with the F'lac+ plasmid, is restriction-deficient
(Res-).  The Res- phenotype is not due to selection of preexisting mutants but
rather to a complex mutational event caused by the F plasmid.
Restriction-deficient mutants carry extensive deletions and other DNA
rearrangements.  Tn7 insertion is used to locate the restriction gene.  Many of
the Res- mutants are genetically unstable and revert at exceptionally high
frequencies.  Reversion is accompanied by DNA rearrangements which result in a
net gain of 9kb of DNA.  F- derivatives of F+ which do not cause
restriction-deficiency but do cause deletion were used to distinguish between
the DNA rearrangements associated with restriction-deficiency and those
associated with deletion.  From Res+ revertants of strains carrying F'lac+ and
R124 or R124/3 we have isolated F plasmids that now carry the genes for the
R124 or R124/3 restriction and modification systems.  It is suggested that
interaction between part of the F plasmid and that segment of the R plasmid
which controls the switch in Res-Mod specificity which has been observed
(Glover et al. 1983) is responsible for the production of
restriction-deficiency.

<>

<1>Firman, K., Dutta, C., Weiserova, M., Janscak, P.
<2>The role of subunit assembly in the functional control of Type I restriction-modification enzymes.
<3>Mol. Biol. Today
<4>1
<5>35-41
<6>2000
<7>Type I restriction-modification enzymes are multifunctional, multisubunit enzymes that provide
their host bacteria with protection
against invading DNA. This protection is accomplished through a very
efficient cleavage of foreign DNA using a complex mechanism involving
hydrolysis of ATP and subsequent translocation of the substrate DNA. In
addition, the same enzymes provide protection for the host chromosome
against cleavage by methylating the target recognition sites. However,
the restriction (cleavage) activity of the enzyme must be carefully
controlled both in terms of what constitutes substrate DNA ("foreign"
versus "host") and the timing of the two activities when transferred to
a new host (modification must precede restriction to allow
establishment of the R-M system). The mechanism by which the enzyme
differentiates between "foreign" and "host" DNA is described and we
discuss recent evidence showing post-translational control as the
primary mechanism for temporal control of restriction. The importance
of subunit assembly in these processes is detailed and extended to
include novel assemblies with unexpected function.

<>

<1>Firman, K., Dutta, C.F.
<2>Construction of restriction-modification endonuclease subunit R fusion protein EcoprrI/EcoR124I and its use in a molecular motor capable of  moving DNA.
<3>International Patent Office
<4>WO 200340302 A
<5>26
<6>2003
<7>An R sub-unit protein of an E. coli Type IC restriction-modification (R-M), endonuclease in
which an N-terminal portion of the R sub-unit, preferably at least amino acids 1-140,
especially 1-146, is derived from EcoprrI and the remainder is derived from another E.coli
Type IC R-M endonuclease, preferably EcoR1241.  The R-M endonuclease formed from the R
sub-unit substantially in the R1M2S stoichiometric form, complexes with DNA and can be powered
by Mg2+ and ATP to form a molecular motor capable of moving DNA.

<>

<1>Firman, K., Price, C., Glover, S.W.
<2>The EcoR124 and EcoR124/3 restriction and modification systems:  cloning the genes.
<3>Plasmid
<4>14
<5>224-234
<6>1985
<7>The Escherichia coli plasmid R124 codes for a type I restriction and
modification system EcoR124 and carries genetic information, most probably in
the form of a "silent copy", for the expression of a different R-M specificity
R124/3.  Characteristic DNA rearrangements have been shown to accompany the
switch in specificity from R124 to R124/3 and vice versa.  We have cloned a
14.2-kb HindIII fragment from R124 and shown that it contains the hsdR, hsdM,
and hsdS genes which code for the EcoR124 R-M system.  An equivalent fragment
from the plasmid R124/3 following the switch in R-M specificity has also been
cloned and shown to contain the genes coding for the EcoR124/3 R-M system.
These fragments, however, lack a component present on the wild-type plasmid
essential for the switch in specificity.  Restriction fragment maps and
preliminary heteroduplex analysis indicate the near identity of the genes that
encode the two different DNA recognition specificities.  Transposon mutagenesis
was used to locate the positions of the hsdR, hsdM, and hsdS genes on the
cloned fragments in conjunction with complementation tests for gene function.
Indirect evidence indicates that hsdR is expressed from its own promoter and
that hsdM and hsdS are expressed from a single promoter, unidirectionally.

<>

<1>Firman, K., Szczelkun, M.D.
<2>Measuring motion on DNA by the type I restriction endonuclease EcoR124I using triplex displacement.
<3>EMBO J.
<4>19
<5>2094-2102
<6>2000
<7>The type I restriction enzyme EcoR124I cleaves DNA following extensive linear translocation
dependent upon ATP hydrolysis. Using protein-directed displacement of a DNA triplex, we have
determined the kinetics of one-dimensional motion without the necessity of measuring DNA or
ATP hydrolysis. The triplex was pre-formed specifically on linear DNA, 4370 bp from an
EcoR124I site, and then incubated with endonuclease. Upon ATP addition, a distinct lag phase
was observed before the triplex-forming oligonucleotide was displaced with exponential
kinetics. As the distance between type I and triplex sites was shortened, the lag time
decreased whilst the displacement reaction remained exponential. This is indicative of
processive DNA translocation followed by collision with the triplex and oligonucleotide
displacement. A linear relationship between lag duration and inter-site distance gives a
translocation velocity of 400 +/- 32 bp/s at 20 degrees C. Furthermore, the data can only be
explained by bi-directional translocation. An endonuclease with only one of the two HsdR
subunits responsible for motion could still catalyse translocation. The reaction is less
processive, but can 'reset' in either direction whenever the DNA is released.

<>

<1>Firnhaber, C., Gerber, M., Tooley, K., Scoggin, C.
<2>DNA Contamination in Commercial Restriction Endonucleases.
<3>Am. J. Hum. Genet.
<4>39
<5>145
<6>1986
<7>None

<>

<1>Fischer, A., Santana-Cruz, I., Giglio, M., Nadendla, S., Drabek, E., Vilei, E.M., Frey, J., Jores, J.
<2>Genome Sequence of Mycoplasma feriruminatoris sp. nov., a Fast-Growing Mycoplasma Species.
<3>Genome Announcements
<4>1
<5>e00216-12
<6>2013
<7>Members of the ' cluster' represent important livestock pathogens worldwide. We report the
genome sequence of sp. nov., the closest relative to the ' cluster'
and the fastest-growing species described to date.

<>

<1>Fischer, M.G., Allen, M.J., Wilson, W.H., Suttle, C.A.
<2>Giant virus with a remarkable complement of genes infects marine zooplankton.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>19508-19513
<6>2010
<7>As major consumers of heterotrophic bacteria and phytoplankton,
microzooplankton are a critical link in aquatic foodwebs. Here, we
show that a major marine microflagellate grazer is infected by
a giant virus, Cafeteria roenbergensis virus (CroV), which has the
largest genome of any described marine virus (¡Ö730 kb of doublestranded
DNA). The central 618-kb coding part of this AT-rich genome
contains 544 predicted protein-coding genes; putative early
and late promoter motifs have been detected and assigned to 191
and 72 of them, respectively, and at least 274 genes were expressed
during infection. The diverse coding potential of CroV includes predicted
translation factors, DNA repair enzymes such as DNA mismatch
repair protein MutS and two photolyases, multiple ubiquitin
pathway components, four intein elements, and 22 tRNAs. Many
genes including isoleucyl-tRNA synthetase, eIF-2¦Ã, and an Elp3-
like histone acetyltransferase are usually not found in viruses.
We also discovered a 38-kb genomic region of putative bacterial
origin, which encodes several predicted carbohydrate metabolizing
enzymes, including an entire pathway for the biosynthesis of 3-
deoxy-D-manno-octulosonate, a key component of the outer membrane
in Gram-negative bacteria. Phylogenetic analysis indicates
that CroV is a nucleocytoplasmic large DNA virus, with Acanthamoeba
polyphaga mimivirus as its closest relative, although less
than one-third of the genes of CroV have homologs in Mimivirus.
CroV is a highly complex marine virus and the only virus studied in
genetic detail that infects one of the major groups of predators in
the oceans.

<>

<1>Fischer, W., Breithaupt, U., Kern, B., Smith, S.I., Spicher, C., Haas, R.
<2>A comprehensive analysis of Helicobacter pylori plasticity zones reveals that they are integrating conjugative elements with intermediate integration specificity.
<3>BMC Genomics
<4>15
<5>310
<6>2014
<7>BACKGROUND: The human gastric pathogen Helicobacter pylori is a paradigm for
chronic bacterial infections. Its persistence in the stomach mucosa is
facilitated by several mechanisms of immune evasion and immune modulation, but
also by an unusual genetic variability which might account for the capability to
adapt to changing environmental conditions during long-term colonization. This
variability is reflected by the fact that almost each infected individual is
colonized by a genetically unique strain. Strain-specific genes are dispersed
throughout the genome, but clusters of genes organized as genomic islands may
also collectively be present or absent. RESULTS: We have comparatively analysed
such clusters, which are commonly termed plasticity zones, in a high number of H.
pylori strains of varying geographical origin. We show that these regions contain
fixed gene sets, rather than being true regions of genome plasticity, but two
different types and several subtypes with partly diverging gene content can be
distinguished. Their genetic diversity is incongruent with variations in the rest
of the genome, suggesting that they are subject to horizontal gene transfer
within H. pylori populations. We identified 40 distinct integration sites in 45
genome sequences, with a conserved heptanucleotide motif that seems to be the
minimal requirement for integration. CONCLUSIONS: The significant number of
possible integration sites, together with the requirement for a short conserved
integration motif and the high level of gene conservation, indicates that these
elements are best described as integrating conjugative elements (ICEs) with an
intermediate integration site specificity.

<>

<1>Fisher, E.W., Yang, M.-T., Jeng, S.-T., Gardner, J.F., Gumport, R.I.
<2>Selection of mutations altering specificity in restriction-modification enzymes using the bacteriophage P22 challenge-phage system.
<3>Gene
<4>157
<5>119-121
<6>1995
<7>A method for selecting mutants of site-specific DNA-binding proteins has been applied to the
study of the EcoRI and RsrI restriction-modification enzymes.  Catalytically inactive variants
of both endonucleases are shown to function as pseudo-repressors in the bacteriophage P22
challenge-phage assay, and, upon further mutagenesis of the gene encoding R.EcoRI, a variant
of that enzyme has been selected which appears to bind EcoRI-methylated GAATTC sequences to
the exclusion of unmethylated sites: this specificity is the opposite of that belonging to the
native enzyme.  Variants of the EcoRI methylase have also been found that lack either
catalytic activity or both binding and catalytic activities.

<>

<1>Fisher, E.W., Yang, M.T., Gardner, J.F., Gumport, R.I.
<2>Isolation of a mutant EcoRI endonuclease with enhanced affinity for duplex methylated d(GAATTC).
<3>FASEB J.
<4>7
<5>A1290
<6>1993
<7>The EcoRI endonuclease, one of the best characterized type II restriction endonucleases,
cleaves duplex DNA at the sequence GAATTC with extreme specificity, catalyzing incorrect
cleavage fewer than once in 10 to the 7th binding events. A companion DNA methyltransferase
uniquely methylates the inner adenosine residue of the same sequence, inhibiting recognition
of the site by the endonuclease. We have applied the bacteriophage P22 challenge-phage system
to the selection of a mutant of the endonuclease which binds the methylated sequence in
preference to the normal, unmethylated target sequence. The challenge-phage assay links
survival of the infected cell to the presence of a protein capable of binding a cloned
recognition sequence of the user's choice, and permits selection of mutant proteins with this
capability from a randomly mutagenized population. In this case a methylated EcoRI site was
chosen as the target recognition sequence by including in the host cells a plasmid bearing the
EcoRI methylase gene. We have isolated a mutant of EcoRI endonuclease, generated by
hydroxylamine treatment of a plasmid encoding a catalytically inactive form of the enzyme
(E111Q), which binds the methylated site tightly enough to form a stable lysogen in
challenge-phage assays. The challenge-phage data suggests that the mutant protein requires the
methyl group for binding, because cells lacking the EcoRI methylase gene are unable to form
lysogens. Current efforts are directed toward identifying the mutational change and examining
by gel mobility shift assays whether the mutant protein discriminates the methylated from the
unmethylated site in vitro. This work was supported in part by a grant (GM25621) from the
National Institute of Health to R.I.G.

<>

<1>Fisher, O., Siman-Tov, R., Ankri, S.
<2>Characterization of cytosine methylated regions and 5-cytosine DNA methyltransferase (Ehmeth) in the protozoan parasite Entamoeba histolytica.
<3>Nucleic Acids Res.
<4>32
<5>287-297
<6>2004
<7>The DNA methylation status of the protozoan parasite Entamoeba histolytica was heretofore
unknown.  In the present study, we developed a new technique, based on the affinity of
methylated DNA to 5-methyl-cytosine antibodies, to identify methylated DNA in this parasite.
Ribosomal. DNA and ribosomal DNA circles were isolated by this method and we confirmed the
validity of our approach by sodium bisulfite sequencing.  We also report the identification
and the characterization of a gene, Ehmeth, encoding a DNA methyltransferase strongly
homologous to the human DNA methyltransferase 2 (Dnmt2).  Immunofluorescence microscopy using
an antibody raised against a recombinant Ehmeth showed that Ehmeth is concentrated in the
nuclei of trophozoites.  The recombinant Ehmeth has a weak but significant methyltransferase
activity when E. histolytica genomic DNA is used as substrate.  5-Azacytidine (5-AzaC), an
inhibitor of DNA methyltransferase, was used to study in vivo the role of DNA methylation in
E. histolytica.  Genomic DNA of trophozoites grown with 5-AzaC (23 uM) was undermethylated and
the ability of 5-AzaC-treated trophozoites to kill mammalian cells or to cause liver abscess
in hamsters was strongly impaired.

<>

<1>Fisunov, G.Y., Alexeev, D.G., Bazaleev, N.A., Ladygina, V.G., Galyamina, M.A., Kondratov, I.G., Zhukova, N.A., Serebryakova, M.V., Demina, I.A., Govorun, V.M.
<2>Core proteome of the minimal cell: comparative proteomics of three mollicute species.
<3>PLoS ONE
<4>6
<5>E21964
<6>2011
<7>Mollicutes (mycoplasmas) have been recognized as highly evolved prokaryotes with
an extremely small genome size and very limited coding capacity. Thus, they may
serve as a model of a 'minimal cell': a cell with the lowest possible number of
genes yet capable of autonomous self-replication. We present the results of a
comparative analysis of proteomes of three mycoplasma species: A. laidlawii, M.
gallisepticum, and M. mobile. The core proteome components found in the three
mycoplasma species are involved in fundamental cellular processes which are
necessary for the free living of cells. They include replication, transcription,
translation, and minimal metabolism. The members of the proteome core seem to be
tightly interconnected with a number of interactions forming core interactome
whether or not additional species-specific proteins are located on the periphery.
We also obtained a genome core of the respective organisms and compared it with
the proteome core. It was found that the genome core encodes 73 more proteins
than the proteome core. Apart of proteins which may not be identified due to
technical limitations, there are 24 proteins that seem to not be expressed under
the optimal conditions.

<>

<1>Fittipaldi, N., Beres, S.B., Olsen, R.J., Kapur, V., Shea, P.R., Watkins, M.E., Cantu, C.C., Laucirica, D.R., Jenkins, L., Flores, A.R., Lovgren, M., Ardanuy, C., Linares, J., Low, D.E., Tyrrell, G.J., Musser, J.M.
<2>Full-Genome Dissection of an Epidemic of Severe Invasive Disease Caused by a Hypervirulent, Recently Emerged Clone of Group A Streptococcus.
<3>Am. J. Pathol.
<4>180
<5>1522-1534
<6>2012
<7>Group A Streptococcus (GAS) causes an exceptionally broad range of infections in humans, from
relatively mild pharyngitis and skin infections to life-threatening necrotizing fasciitis and
toxic shock syndrome. An epidemic of severe invasive human infections caused by type emm59
GAS, heretofore an exceedingly rare cause of disease, spread west to east across Canada over a
3-year period (2006 to 2008). By sequencing the genomes of 601 epidemic, historic, and other
emm59 organisms, we discovered that a recently emerged, genetically distinct emm59 clone is
responsible for the Canadian epidemic. Using near-real-time genome sequencing, we were able to
show spread of the Canadian epidemic clone into the United States. The extensive genome data
permitted us to identify patterns of geographic dissemination as well as links between emm59
subclonal lineages that cause infections. Mouse and nonhuman primate models of infection
demonstrated that the emerged clone is unusually virulent. Transmission of epidemic emm59
strains may have occurred primarily by skin contact, as suggested by an experimental model of
skin transmission. In addition, the emm59 strains had a significantly impaired ability to
persist in human saliva and to colonize the oropharynx of mice, and seldom caused human
pharyngitis. Our study contributes new information to the rapidly emerging field of molecular
pathogenomics of bacterial epidemics and illustrates how full-genome data can be used to
precisely illuminate the landscape of strain dissemination during a bacterial epidemic.

<>

<1>Fitz-Gibbon, S., Tomida, S., Chiu, B.H., Nguyen, L., Du, C., Liu, M., Elashoff, D., Erfe, M.C., Loncaric, A., Kim, J., Modlin, R.L., Miller, J.F., Sodergren, E., Craft, N., Weinstock, G.M., Li, H.
<2>Propionibacterium acnes Strain Populations in the Human Skin Microbiome Associated with Acne.
<3>J. Invest. Dermatol.
<4>133
<5>2152-2160
<6>2013
<7>The human skin microbiome has important roles in skin health and disease.
However, bacterial population structure and diversity at the strain level is
poorly understood. We compared the skin microbiome at the strain level and genome
level of Propionibacterium acnes, a dominant skin commensal, between 49 acne
patients and 52 healthy individuals by sampling the pilosebaceous units on their
noses. Metagenomic analysis demonstrated that although the relative abundances of
P. acnes were similar, the strain population structures were significantly
different in the two cohorts. Certain strains were highly associated with acne,
and other strains were enriched in healthy skin. By sequencing 66 previously
unreported P. acnes strains and comparing 71 P. acnes genomes, we identified
potential genetic determinants of various P. acnes strains in association with
acne or health. Our analysis suggests that acquired DNA sequences and bacterial
immune elements may have roles in determining virulence properties of P. acnes
strains, and some could be future targets for therapeutic interventions. This
study demonstrates a previously unreported paradigm of commensal strain
populations that could explain the pathogenesis of human diseases. It underscores
the importance of strain-level analysis of the human microbiome to define the
role of commensals in health and disease.Journal of Investigative Dermatology
advance online publication, 28 February 2013; doi:10.1038/jid.2013.21.

<>

<1>Fitz-Gibbon, S.T., Ladner, H., Kim, U.-J., Stetter, K.O., Simon, M.I., Miller, J.H.
<2>Genome sequence of the hyperthermophilic crenarchaeon Pyrobaculum aerophilum.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>984-989
<6>2002
<7>We determined and annotated the complete 2.2-megabase genome sequence of Pyrobaculum
aerophilum, a facultatively aerobic nitrate-reducing hyperthermophilic (Topt=100 C)
crenarchaeon.  Clues were found suggesting explanations of the organism's surprising
intolerance to sulfur, which may aid in the development of methods for genetic studies of the
organism.  Many interesting features worthy of further genetic studies were revealed.  Whole
genome computational analysis confirmed experiments showing that P. aerophilum (and perhaps
all crenarchaea) lack 5' untranslated regions in their mRNAs and thus appear not to use a
ribosome-binding site (Shine-Delgarno)-based mechanism for translation initiation at the 5'
end of transcripts.  Inspection of the lengths and distribution of mononucleotide
repeat-tracts revealed some interesting features.  For instance, it was seen that
mononucleotide repeat-tracts of Gs (or Cs) are highly unstable, a pattern expected for an
organism deficient in mismatch repair.  This result, together with an independent study on
mutation rates, suggests a "mutator" phenotype.

<>

<1>Fitzgerald, G.F., Twomey, D.P., Daly, C., Coffey, A.G.
<2>Bacteriophage resistance in Lactococcus: Molecular characterization of the ScrFI restriction/modification system from Lactococcus lactis subsp. cremoris UC503.
<3>Genetics and Molecular Biology of Streptococci, Lactococci, and Enterococci, Karger, Ferretti, J.J., Gilmore, M.S., Klaenhammer, T.R., Brown, F., Basel
<4>85
<5>581-590
<6>1995
<7>Members of the genus Lactococcus are used as starter cultures in the manufacture of a range of
ripened and unripened fermented dairy products such as cheese, lactic butter and cottage
cheese.  In many cases the production of these foods is concentrated in large manufacturing
units where starter cultures are used on a more or less continuous and intensive basis.  In
such modern, highly efficient manufacturing regimes, poor starter performance is undesirable.
Thus, considerable effort has been targeted to successfully eliminate those factors, such as
poor milk quality, which could have a negative impact on the activity of the starter culture.
While there has been considerable success in countering the major problem of bacteriophage
infection in cheese plants, phage still represent a very significant threat to the efficient
performance of lactococcal starter cultures.  Obvious physical approaches such as careful
design of manufacturing plants, aseptic handling of cultures and proper separation of whey
from the culture preparation area have had considerable success but on their own will not
guarantee a phage-free environment.  A further approach has been to examine the cultures
themselves and to use isolates which display inherent resistance to phage.  Thus, carefully
selected strains, exhibiting considerable levels of natural resistance to phage, have been
included in defined starter culture systems used in many parts of the world, particularly for
Cheddar cheese production.  These defined systems have proved to be quite successful in
countering phage, especially when they are combined with good sanitization and asepsis
regimes.  The quest for lactococcal hosts exhibiting high levels of resistance to phage has
stimulated the analysis of phage/host interactions at a fundamental level.  One outcome of
this research has been the recognition that some of these hosts possess one or more specific
mechanisms which can confer on the host significant levels of resistance to phage.  These are
usually, although not always, plasmid-encoded and quite often these plasmids are
self-transmissible.  In general, three distinct types of resistance have been identified:
adsorption inhibition, abortive infection (Abi) and restriction/modification (R/M).  Since the
topic of phage resistance in lactococci has been reviewed in considerable depth in a number of
recent publications, these specific areas will not be discussed in detail.  The major emphasis
of this paper will be on the molecular analysis of the ScrFI R/M system, which mediates
insensitivity to phage in vivo and has a number of unusual and interesting features.

<>

<1>Fitzgerald, L.A., Graves, M.V., Li, X., Feldblyum, T., Hartigan, J., Van Etten, J.L.
<2>Sequence and annotation of the 314-kb MT325 and the 321-kb FR483 viruses that infect Chlorella Pbi.
<3>Virology
<4>358
<5>459-471
<6>2007
<7>Viruses MT325 and FR483, members of the family Phycodnaviridae, genus Chlorovirus, infect the
fresh water, unicellular, eukaryotic, chlorella-like green alga, Chlorella Pbi. The 314,335-bp
genome of MT325 and the 321,240-bp genome of FR483 are the first viruses that infect Chlorella
Pbi to have their genomes sequenced and annotated. Furthermore, these genomes are the two
smallest chlorella virus genomes sequenced to date, MT325 has 331 putative protein-encoding
and 10 tRNA-encoding genes and FR483 has 335 putative protein-encoding and 9 tRNA-encoding
genes. The protein-encoding genes are almost evenly distributed on both strands, and
intergenic space is minimal. Approximately 40% of the viral gene products resemble entries in
public databases, including some that are the first of their kind to be detected in a virus.
For example, these unique gene products include an aquaglyceroporin in MT325, a potassium ion
transporter protein and an alkyl sulfatase in FR483, and a dTDP-glucose pyrophosphorylase in
both viruses. Comparison of MT325 and FR483 protein-encoding genes with the prototype
chlorella virus PBCV-1 indicates that approximately 82% of the genes are present in all three
viruses.

<>

<1>Fitzgerald, L.A., Graves, M.V., Li, X., Feldblyum, T., Nierman, W.C., Van Etten, J.L.
<2>Sequence and annotation of the 369-kb NY-2A and the 345-kb AR158 viruses that infect Chlorella NC64A.
<3>Virology
<4>358
<5>472-484
<6>2007
<7>Viruses NY-2A and AR158, members of the family Phycodnaviridae, genus
Chlorovirus, infect the fresh water, unicellular, eukaryotic,
chlorella-like green alga, Chlorella NC64A. The 368,683-bp genome of NY-2A
and the 344,690-bp genome of AR158 are the two largest chlorella virus
genomes sequenced to date; NY-2A contains 404 putative protein-encoding
and 7 tRNA-encoding genes and AR158 contains 360 putative protein-encoding
and 6 tRNA-encoding genes. The protein-encoding genes are almost evenly
distributed on both strands, and intergenic space is minimal. Two of the
NY-2A genes encode inteins, the large subunit of ribonucleotide reductase
and a superfamily II helicase. These are the first inteins to be detected
in the chlorella viruses. Approximately 40% of the viral gene products
resemble entries in the public databases, including some that are
unexpected for a virus. These include GDP-d-mannose dehydratase, fucose
synthase, aspartate transcarbamylase, Ca(++) transporting ATPase and
ubiquitin. Comparison of NY-2A and AR158 protein-encoding genes with the
prototype chlorella virus PBCV-1 indicates that 85% of the genes are
present in all three viruses.

<>

<1>Fitzgerald, M.C., Skowron, P., Van Etten, J.L., Smith, L.M., Mead, D.A.
<2>Rapid shotgun cloning utilizing the two base recognition endonuclease CviJI.
<3>Nucleic Acids Res.
<4>20
<5>3753-3762
<6>1992
<7>A new approach has been developed for the rapid fragmentation and fractionation of DNA into a
size suitable for shotgun cloning and sequencing. The restriction endonuclease CviJI normally
cleaves the recognition sequence PuGCPy between the G and C to leave blunt ends. Atypical
reaction conditions which alter the specificity of this enzyme (CviJI**) yield a quasi-random
distribution of DNA fragments from the small molecule pUC19 (2686 base pairs). To
quantitatively evaluate the randomness of this fragmentation strategy, a CviJI** digest of
pUC19 was size fractionated by a rapid gel filtration method and directly ligated, without end
repair, to a lacZ minus M13 cloning vector. Sequence analysis of 76 clones showed that CviJI**
restricts PyGCPy and PuCGPu, in addition to PuGCPy sites, and that new sequence data is
accumulated at a rate consistent with random fragmentation. Advantages of this approach
compared to sonication and agarose gel fractionation include: smaller amounts of DNA are
required (0.2-0.5 ug instead of 2-5 ug), fewer steps are involved (no pre-ligation, end
repair, chemical extraction, or agarose gel electrophoresis and elution are needed), and
higher cloning efficiencies are obtained (CviJI** digested and column fractionated DNA
transforms 3-16 times more efficiently than sonicated, end-repaired, and agarose fractionated
DNA).

<>

<1>Fitzgerlad, G.F., Daly, C., Brown, L.R., Gingeras, T.R.
<2>ScrFI: a new sequence-specific endonuclease from Streptococcus cremoris.
<3>Nucleic Acids Res.
<4>10
<5>8171-8179
<6>1982
<7>A novel sequence-specific endonuclease has been isolated from Streptococcus cremoris F. ScrFI
recognizes the sequence:5'CC^NGG3'3'GGN^CC5'and cleaves as indicated by the arrow.  It is
the first enzyme to recognize this sequence and the first endonuclease reported from the
lactic streptococci used in dairy fermentations.

<>

<1>Flanagan, J.C., Lang, J.M., Darling, A.E., Eisen, J.A., Coil, D.A.
<2>Draft Genome Sequence of Curtobacterium flaccumfaciens Strain UCD-AKU (Phylum Actinobacteria).
<3>Genome Announcements
<4>1
<5>e00244-13
<6>2013
<7>Here we present the draft genome of an actinobacterium, Curtobacterium flaccumfaciens strain
UCD-AKU, isolated from a residential carpet. The genome
assembly contains 3,692,614 bp in 130 contigs. This is the first member of the
Curtobacterium genus to be sequenced.

<>

<1>Fleischman, R.A., Campbell, J.L., Richardson, C.C.
<2>Modification and restriction of T-even bacteriophages.
<3>J. Biol. Chem.
<4>251
<5>1561-1570
<6>1976
<7>Using the single-stranded circular DNA of bacteriophage fd as template,
double-stranded circular DNA has been prepared in vitro with either 5-
hydroxymethylcytosine ([hmdC]DNA) or cytosine ([dC]DNA) in the product strand.
Extracts prepared from Escherichia coli cells restrictive to T-even phage containing
nonglucosylated DNA degrade [hmdC]DNA to acid-soluble material in vitro, but do not
degrade [dC]DNA.  In contrast, extracts prepared from E. coli K12 rglA- rglB-, a strain
permissive to T-even phage containing nonglucosylated DNA, do not degrade [hmdC]DNA
or [dC]DNA.  In addition, glucosylation of the [hmdC]DNA renders it resistant to
degradation by extracts from restrictive strains.  The conversion of [hmdC]DNA to acid-
soluble material in vitro consists of an HmCyt-specific endonucleolytic cleavage requiring
the presence of the RglB gene product to form a linear molecule, followed by a non-
HmCyt-specific hydrolysis of the linear DNA to acid-soluble fragments, catalyzed in part
by exonuclease V.  The RglB protein present in extracts of E. coli K12 rglA- rglB+ has
been purified 200-fold by complementation with extracts from E. coli K12 rglA- rglB-.
The purified RglB protein does not contain detectable HmCyt-specific endonuclease or
exonuclease activity.  In vitro endonucleolytic cleavage of [hmdC]DNA thus requires
additional factors present in cell extracts.

<>

<1>Fleischman, R.A., Richardson, C.C.
<2>Analysis of host range restriction in Escherichia coli treated with toluene.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>68
<5>2527-2531
<6>1971
<7>Escherichia coli cells treated with toluene replicate DNA when they are
provided with deoxyribonucleoside 5'-triphosphates, ATP, Mg++, and K+.
However, when deoxycytidine 5'-triphosphate is replaced by hydroxymethyl
deoxycytidine 5'-triphosphate, incorporation of nucleotides into
acid-precipitable material by toluene-treated strains restrictive to
nonglucosylated T-even phage is reduced to less than 5% of that normally
observed.  Even when dCTP is present in the reaction mixture, a similar effect
of the hydroxymethyl analogue on DNA replication is observed.  In contrast,
toluene-treated E. coli K12 r6-r2,4-, a strain permissive to the
nonglucosylated T-even phage, incorporates hydroxymethyl deoxycytosine into its
DNA, and replication proceeds at only a slightly reduced rate in the presence
of the hydroxymethyl deoxycytidine 5'-triphosphate.  The presence of the
hydroxymethyl deoxycytidine 5'-triphosphate in the reaction mixture does not
lead to degradation of preexisting DNA of the restrictive host, but it does
lead to an irreversible inhibition of further DNA replication; the inhibition
is observed only when the hydroxymethyl deoxycytidine 5'-triphosphate is
present during replication.  Thus phage-specific enzymes are not necessary for
the incorporation of hydroxymethylcytosine into phage DNA, and the restrictive
mechanism, present in the host cell before infection, can recognize
hydroxymethylcytosine residues in its own DNA, as well as the DNA of the T-even
phage.

<>

<1>Fleischmann, R.D. et al.
<2>Whole-genome random sequencing and assembly of Haemophilus influenzae Rd.
<3>Science
<4>269
<5>496-512
<6>1995
<7>An approach for genome analysis based on sequencing and assembly of unselected pieces of DNA
from the whole chromosome has been applied to obtain the complete nucleotide sequence
(1,830,137 base pairs) of the genome from the bacterium Haemophilus influenzae Rd.  This
approach eliminates the need for initial mapping efforts and is therefore applicable to the
vast array of microbial species for which genome maps are unavailable.  The H. influenzae Rd
genome sequence (Genome Sequence Database accession number L42023) represents the only
complete genome sequence from a free-living organism.

<>

<1>Fleischmann, R.D., Adams, M.D., White, O., Smith, H.O., Venter, J.C.
<2>Nucleotide sequence of the Haemophilus influenzae Rd genome, fragments thereof, and uses thereof.
<3>US Patent Office
<4>US 6506581 A
<5>
<6>2003
<7>The present invention provides the sequencing of the entire genome of Haemophilus influenzae
Rd, SEQ ID NO: 1.  The present invention further provides the sequence information stored on
computer readable media, and computer-based systems and methods which facilitate its use.  In
addition to the entire genomic sequence, the present invention identifies over 1700 protein
encoding fragments of the genome and identifies, by position relative to a unique Not I
restriction endonuclease site, any regulatory elements which modulate the expression of the
protein encoding fragments of the Haemophilus genome.

<>

<1>Fleischmann, R.D., Adams, M.D., White, O., Smith, H.O., Venter, J.C.
<2>Genomic Sequence of the Haemophilus influenzae Rd, its fragment and Use Thereof.
<3>Japanese Patent Office
<4>JP 2004041185 A
<5>
<6>2004
<7>Problem to be solved: To provide the entire genome sequence of Haemophilus influenzae Rd.
Solution: The invention provides a sequence expressed by a specific sequence, the sequence
information stored on a computer readable medium, a computer-based system, a method to make
the easy use of the system and over 1,700 protein encoding fragments in the sequence expressed
by a specific sequence.

<>

<1>Fleischmann, R.D., Adams, M.D., White, O., Smith, H.O., Venter, J.C.
<2>Nucleotide sequence of the Haemophilus influenzae Rd genome, fragments thereof, and uses thereof.
<3>US Patent Office
<4>US 6846651 A
<5>
<6>2005
<7>The present invention provides the sequencing of the entire genome of Haemophilus influenzae
Rd, SEQ ID NO:1.  The present invention further provides the sequence information stored on
computer readable media, and computer-based systems and methods which facilitate its use.  In
addition to the entire genomic sequence, the present invention identifies over 1700 protein
encoding fragments of the genome and identifies, by position relative to a unique NotI
restriction endonuclease site, any regulatory elements which modulate the expression of the
protein encoding fragments of the Haemophilus genome.

<>

<1>Fletcher, B.S.
<2>Development and validation of an approach to produce large-scale quantities of CpG-methylated plasmid DNA.
<3>Micro. Biotech.
<4>1
<5>62-67
<6>2008
<7>The prokaryotic CpG-specific DNA methylase from Spiroplasma, SssI methylase, has been
extensively used to methylate plasmid DNA in vitro
to investigate the effects of methylation in vertebrate systems.
Currently available methods to produce CpG-methylated plasmid DNA have
certain limitations and cannot generate large quantities of methylated
DNA without cost or problems of purity. Here we describe an approach in
which the SssI methylase gene has been introduced into the Escherichia
coli bacterial genome under the control of an inducible promoter.
Plasmid DNA propagated in this bacterium under conditions which induce
the methylase gene result in significant (> 90%) CpG methylation.
Methylated DNA produced by this approach behaves similarly to
methylated DNA produced in vitro using the purified methylase. The
approach is scalable allowing for the production of milligram
quantities of methylated plasmid DNA.

<>

<1>Flett, F., Mersinias, V., Smith, C.P.
<2>High efficiency intergeneric conjugal transfer of plasmid DNA from Escherichia coli to methyl DNA-restricting streptomycetes.
<3>FEMS Microbiol. Lett.
<4>155
<5>223-229
<6>1997
<7>Many streptomycetes, including S. coelicolor A3(2), possess a potent methyl-specific
restriction which can present an effective barrier to the
introduction of heterologous DNA. We have compared the efficiency of intergeneric
conjugal transfer of different types of plasmids to S. coelicolor and S. lividans
66 using two E. coli donors: the standard, methylation proficient strain S17-1,
and the methylation deficient donor, ET12567(pUB307). We demonstrate that the
methylation deficient donor can yield > 10(4)-fold more S. coelicolor
exconjugants than the standard donor. In the case of pSET152 derivatives, which
integrate into the host chromosome by site-specific recombination, up to 10% of
streptomycete spores in the conjugation mixture inherit the plasmid. The
conjugation procedure is efficient enough to obtain exconjugants with 'suicide'
delivery plasmids and therefore provides a simple route for conducting gene
disruptions in methyl DNA-restricting streptomycetes, and possibly other
bacteria.

<>

<1>Flett, F., Wotton, S.F., Kirby, R.
<2>A common host specificity in the restriction and modification of a bacteriophage by three distinct Streptomyces species.
<3>J. Gen. Microbiol.
<4>110
<5>465-467
<6>1979
<7>Restriction/modification of bacteriophage is a well characterized phenomenon in many bacterial
species and has been identified among streptomycetes in Streptomyces albus G, Streptomyces
griseus Kr. 15 and Streptomyces coelicolor.  Direct correlation between bacterial restriction
of bacteriophage and a specific endonuclease has only been demonstrated for a few species,
including one Streptomyces species.  The discovery of extrachromosomal elements in
Streptomyces opened up the possibility of detecting such an element carrying
restriction/modification, as has been shown to occur in eubacteria.

<>

<1>Flick, K.E., Jurica, M.S., Monnat, R.J. Jr., Stoddard, B.L.
<2>DNA binding and cleavage by the nuclear intron-encoded homing endonuclease I-PpoI.
<3>Nature
<4>394
<5>96-101
<6>1998
<7>Homing endonucleases are a diverse collection of proteins that are encoded by genes with
mobile, self-splicing introns.  They have also been identified in self-splicing inteins
(protein introns).  These enzymes promote the movement of the DNA sequences that encode them
from one chromosome location to another; they do this by making a site-specific double-strand
break at a target site in an allele that lacks the corresponding mobile intron.  The target
site recognized by these small endonucleases are generally long (14-44 base pairs).  Four
families of homing endonucleases have been identified, including the LAGLIDADG, the His-Cys
box, the GIY-YIG and the H-N-H endonucleases.  The first identified His-Cys box homing
endonuclease was I-PpoI from the slime mould Physarum polycephalum.  Its gene residues in one
of only a few nuclear introns known to exhibit genetic mobility.  Here we report the structure
of the I-PpoI homing endonuclease bound to homing-site DNA determined to 1.8 Angstroms
resolution.  I-PpoI displays an elongated fold of dimensions 25 x 35 x 80 Angstroms, with
mixed alpha/beta topology.  Each I-PpoI monomer contains three antiparallel beta-sheets
flanked by two long alpha-helices and a long carboxy-terminal tail, and is stabilized by two
bound zinc ions 15 Angstroms apart.  The enzyme possesses a new zinc-bound fold and
endonuclease active site.  The structure has been determined in both uncleaved substrate and
cleaved product complexes.

<>

<1>Flick, K.E., McHugh, D., Heath, J.D., Stephens, K.M., Monnat, R.J. Jr., Stoddard, B.L.
<2>Crystallization and preliminary X-ray studies of I-PpoI: A nuclear, intron-encoded homing endonuclease from Physarum polycephalum.
<3>Protein Sci.
<4>6
<5>2677-2680
<6>1997
<7>The homing endonuclease I-PpoI is encoded by an optional third intron, Pp LSU 3, found in
nuclear, extrachromosomal copies of the Physarum polycephalum 26S rRNA gene.  This
endonuclease promotes the lateral transfer or "homing" of its encoding intron by recognizing
and cleaving a partially symmetric, 15 bp homing site in 26S rDNA alleles that lack the Pp LSU
3 intron.  The open reading frame encoding I-PpoI has been subcloned, and the endonuclease has
been overproduced in E. coli.  Purified recombinant I-PpoI has been co-crystallized with a 21
bp homing site DNA duplex.  The crystals belong to space group P3I21, with unit cell
dimensions a=b=114 A, c= 89 A.  The results of initial X-ray diffraction experiments indicate
that the asymmetric unit contains an enzyme homodimer and one duplex DNA molecule, and that
the unit cell has a specific volume of 3.4 A3/dalton.  These experiments also provide strong
evidence that I-PpoI contains several bound zinc ions as part of its structure.

<>

<1>Fliess, A., Wolfes, H., Rosenthal, A., Schwellnus, K., Blocker, H., Frank, R., Pingoud, A.
<2>Role of thymidine residues in DNA recognition by the EcoRI and EcoRV restriction endonucleases.
<3>Nucleic Acids Res.
<4>14
<5>3463-3474
<6>1986
<7>We have synthesized a series of oligonucleotides containing the EcoRI (GAATTC)
or EcoRV (GATATC) recognition site within which or adjacent to which thymidine
was substituted by uridine or derivatives of uridine.  The effects of these
substitutions on the rate of the EcoRI and EcoRV catalyzed cleavage reaction
were investigated.  Our results show that most of the substitutions within the
site are quite well tolerated by EcoRI, not, however, by EcoRV.  We conclude
that the thymin residues most likely are not dirctly involved in the
recognition process of the EcoRI reaction.  In contrast, they are major points
of contact, between substrate and enzym in the EcoRV reaction.  The effects of
substitutions in the position adjacent to the recognition site is also markedly
different for EcoRI and EcoRV.  Here, EcoRI seems to be considerably more
selective than EcoRV.

<>

<1>Fliess, A., Wolfes, H., Seela, F., Pingoud, A.
<2>Analysis of the recognition mechanism involved in the EcoRV catalyzed cleavage of DNA using modified oligodeoxynucleotides.
<3>Nucleic Acids Res.
<4>16
<5>11781-11793
<6>1988
<7>We have prepared a series of undecadeoxynucleotides that contain changes in the
functional group pattern present within the EcoRV recognition site - GATATC -.
Oligonucleotides were synthesized on solid phase using normal and modified
beta-cyanoethylphosphoramidites and analyzed in steady state cleavage
experiments with the EcoRV restriction endonuclease.  The following groups
appear to interact strongly with the enzyme, since their modification or
substitution renders the oligonucleotides refractory to cleavage:  the
exocyclic NH2-groups of both A residues, the N7 of the first A residue, the
exocyclic NH2-group of the C residue and the CH3-groups of both T residues.
The exocyclic NH-group of the G residue supports effective recognition, since
its absence lowers the kcat of the cleavage reaction.  The N7 of the second A
residue and the C5 position of the C residue apparently are not recognized by
EcoRV; their substitution by -CH- or modification with -Br or -CH3 does not
considerably change the rate of cleavage.  All oligonucleotides investigated
compete with the unmodified substrate for binding to the enzyme.  We conclude
that EcoRV recognizes its substrate presumably through hydrogen bonds to the
exocyclic NH2-group and the N7 of the first A residue, the exocyclic NH2-groups
of the second A and the C residue, as well as through hydrophobic interactions
with both T residues.

<>

<1>Flinspach, K., Ruckert, C., Kalinowski, J., Heide, L., Apel, A.K.
<2>Draft Genome Sequence of Streptomyces niveus NCIMB 11891, Producer of the Aminocoumarin Antibiotic Novobiocin.
<3>Genome Announcements
<4>2
<5>e01146-13
<6>2014
<7>Streptomyces niveus NCIMB 11891 is the producer of the gyrase inhibitor novobiocin, which
belongs to the aminocoumarin class of antibiotics. The genome
sequence of this strain was found to contain, besides the gene cluster for
novobiocin, a putative gene cluster for the macrolactam antibiotic BE-14106 and
further secondary metabolite gene clusters.

<>

<1>Flores, H., Osuna, J., Heitman, J., Soberon, X.
<2>Saturation mutagenesis of His114 of EcoRI reveals relaxed-specificity mutants.
<3>Gene
<4>157
<5>295-301
<6>1995
<7>EcoRI recognizes and cleaves DNA at GAATTC sites and is one of the best characterized
sequence-specific restriction endonucleases (ENases).  In previous studies, an EcoRI mutant,
which exhibited relaxed substrate specificity and cleaved both canonical and EcoRI star sites,
was isolated.  This mutant enzyme has Tyr instead of His114.  Here, we subjected residue 114
of the EcoRI ENase to saturation mutagenesis.  The resulting mutant enzymes were characterized
both in vivo and in vitro, resulting in the identification of mutants with canonical (H114K,
Q, D, I) or relaxed (H114Y, F, S, T) specificity, as well as one mutant with severely impaired
activity (H114P).  In the X-ray structure of an EcoRI-substrate complex, His 114 is located
between the catalytic and recognition regions of EcoRI and may directly contact the DNA
phosphate backbone.  Based on our genetic and biochemical findings and the X-ray structure, we
propose that His114 participates in substrate recognition and catalysis, either directly, via
protein-DNA interactions, or indirectly, by mediating conformational changes that trigger DNA
cleavage in response to substrate recognition.

<>

<1>Flores, V., Lopez-Merino, A., Mendoza-Hernandez, G., Guarneros, G.
<2>Comparative genomic analysis of two brucellaphages of distant origins.
<3>Genomics
<4>99
<5>233-240
<6>2012
<7>Here, we present the first complete genome sequence of brucellaphage Tbilisi (Tb)
and compared it with that of Pr, a broad host-range brucellaphage recently
isolated in Mexico. The genomes consist of 41,148bp (Tb) and 38,253bp (Pr), they
differ mainly in the region encoding structural proteins, in which the genome of
Tb shows two major insertions. Both genomes share 99.87% nucleotide identity, a
high percentage of identity among phages isolated at so globally distant
locations and temporally different occasions. Sequence analysis revealed 57
conserved ORFs, three transcriptional terminators and four putative
transcriptional promoters. The co-occurrence of an ORF encoding a putative
DnaA-like protein and a putative oriC-like origin of replication was found in
both brucellaphages genomes, a feature not described in any other phage genome.
These elements suggest that DNA replication in brucellaphages differs from other
phages, and might resemble that of bacterial chromosomes.

<>

<1>Florez, A.B., Reimundo, P., Delgado, S., Fernandez, E., Alegria, A., Guijarro, J.A., Mayo, B.
<2>Genome Sequence of Lactococcus garvieae IPLA 31405, a Bacteriocin-Producing, Tetracycline-Resistant Strain Isolated from a Raw-Milk Cheese.
<3>J. Bacteriol.
<4>194
<5>5118-5119
<6>2012
<7>This work describes the draft genome sequence of Lactococcus garvieae IPLA 31405, isolated
from a traditional Spanish cheese. The genome contains a
lactose-galactose operon, a bacteriocin locus, two integrated phages, a
transposon harboring an active tet(M) gene, and two theta-type plasmid replicons.
Genes encoding virulence factors were not recorded.

<>

<1>Flusberg, B.A., Webster, D.R., Lee, J.H., Travers, K.J., Olivares, E.C., Clark, T.A., Korlach, J., Turner, S.W.
<2>Direct detection of DNA methylation during single-molecule, real-time sequencing.
<3>Nat. Methods
<4>7
<5>461-465
<6>2010
<7>We describe the direct detection of DNA methylation, without bisulfite conversion, through
single-molecule, real-time (SMRT) sequencing. In SMRT sequencing, DNA polymerases catalyze the
incorporation of fluorescently labeled nucleotides into complementary nucleic acid strands.
The arrival times and durations of the resulting fluorescence pulses yield information about
polymerase kinetics and allow direct detection of modified nucleotides in the DNA template,
including N6-methyladenine, 5-methylcytosine and 5-hydroxymethylcytosine.  Measurement of
polymerase kinetics is an intrinsic part of SMRT sequencing and does not adversely affect
determination of primary DNA sequence. The various modifications affect polymerase kinetics
differently, allowing discrimination between them. We used these kinetic signatures to
identify adenine methylation in genomic samples and found that, in combination with circular
consensus
sequencing, they can enable single-molecule identification of epigenetic
modifications with base-pair resolution. This method is amenable to long read
lengths and will likely enable mapping of methylation patterns in even highly
repetitive genomic regions.

<>

<1>Flynn, E., Chatterjee, D., Longo, M., Oberfelder, R.
<2>Methods for Production of Proteins.
<3>Japanese Patent Office
<4>JP 2008119000 A
<5>
<6>2008
<7>
<>

<1>Flynn, J.
<2>Murine DNA cytosine C-5 methyltransferase: Insights into epigenetic control. from enzymological studies.
<3>Diss. Abstr.
<4>58
<5>3614B
<6>1998
<7>The processes that contribute to mammalian development are numerous.  Genomic methylation
patterns change dynamically with each differentiating cell division.  The enzyme responsible
for DNA methylation is essential for normal development.  A precise functional description is
catalysis and epigenesis. A rigorous DNA binding assay defined how sequences flanking CpG
dideoxynucleotides affect the stability of the enzyme:DNA complex.  Oligonucleotides form
reversible 1:1 complexes with the enzyme sequence-specifically.  A guanine/cytosine-rich
sequence that mimics the GC-box boundthree-fold more tightly than the adenine/thymine rich
cyclic AMP responsive element.  Binding discrimination between hemi- and unmethylated forms
was small and single-stranded substrates bound more weakly than double-stranded DNA.  An in
vitro screening method selected for CpG flanking sequence preferences from a large, divergent
population.  After five iterative rounds of increasing selective pressure,
guanosine/cytosine-rich sequences were abundant.  The enzyme uses sequence-dependent
conformational features over two helical turns for discrimination, rather than specific base
interactions.  The constants KmDNA, kcat, and kmethylation were determined.  The rate limiting
step for most substrates was the methylation step itself.  Contrastingly, hemimethylated
substrates exhibited burst kinetics, consistent with a rapid methylation event followed by a
slower step which determines kcat.  Large substrates with multiple recognition sites do not
show burst kinetics and have turnover constants of 30 hr-1.  Maintenance of genomic
methylation patterns is preferred over de novo establishment of new patterns.  Four types of
steady-state analyses were used to identify the order of substrate addition and product
release.  Steady-state initial velocity studies including product and dead-end inhibition
studies support an ordered Bi-Bi kinetic model in which DNA binds first, followed by
S-adenosyl methionine and then release of S-adenosyl homocysteine.

<>

<1>Flynn, J., Azzam, R., Reich, N.
<2>DNA binding discrimination of the murine DNA cytosine-C5 methyltransferase.
<3>J. Mol. Biol.
<4>279
<5>101-116
<6>1998
<7>Mammalian DNA cytosine-C5 methyltransferase modifies the CpG dinucleotide in the context of
many different genomic sequences.  A rigorous DNA binding assay was developed for the murine
enzyme and used to define how sequences flanking the CpG dinucleotide affect the stability of
the enzyme:DNA complex.  Oligonucleotides containing a single CpG site form reversible 1:1
complexes with the enzyme that are sequence-specific.  A guanine/cytosine-rich 30 base-pair
sequence, a mimic of the GC-box cis-element, bound threefold more tightly than an
adenine/thymine-rich sequence, a mimic of the cyclic AMP responsive element.  However, the
binding discrimination between hemi- and unmethylated forms of these DNA substrates was small,
as we previously observed at the KmDNA level.  Single-stranded substrates are bound much more
weakly than double-stranded DNA forms.  An in vitro screening method was used to select for
CpG flanking sequence preferences of the DNA methyltransferase from a large, divergent
population of DNA substrates.  After five interactive rounds of increasing selective pressure,
guanosine/cytosine-rich sequences were abundant and contributed to binding stabilization for
at least 12 base-pairs on either side of a central CpG.  Our results suggest a read-out of
sequence-dependent conformational features, such as helical flexibility, minor groove
dimensions and critical phosphate orientation and mobility, rather than interactions with
specific bases over the course of two complete helical turns.  Thus, both studies reveal a
preference for guanosine/cytosine deoxynucleotides flanking the cognate CpG.  The enzyme
specificity for similar sequences in the genome may contribute to the in vivo functions of
this vital enzyme.

<>

<1>Flynn, J., Fang, J.Y., Mikovits, J.A., Reich, N.O.
<2>A potent cell-active allosteric inhibitor of murine DNA cytosine C5 methyltransferase.
<3>J. Biol. Chem.
<4>278
<5>8238-8243
<6>2003
<7>The major DNA cytosine methyltransferase isoform in mouse erythroleukemia cells, Dnmt1,
exhibits potent dead-end inhibition with a single-stranded
nucleic acid by binding to an allosteric site on the enzyme. The
previously reported substrate inhibition with double-stranded substrates
also involves binding to an allosteric site. Thus, both forms of
inhibition involve ternary enzyme-DNA-DNA complexes. The inhibition
potency of the single-stranded nucleic acid is determined by the sequence,
length, and most appreciably the presence of a single 5-methylcytosine
residue. A single-stranded phosphorothioate derivative inhibits DNA
methylation activity in nuclear extracts. Mouse erythroleukemia cells
treated with the phosphorothioate inhibitor show a significant decrease in
global genomic methylation levels. Inhibitor treatment of human colon
cancer cells causes demethylation of the p16 tumor suppressor gene and
subsequent p16 re-expression. Allosteric inhibitors of mammalian DNA
cytosine methyltransferases, representing a new class of molecules with
potential therapeutic applications, may be used to elucidate novel
epigenetic mechanisms that control development.

<>

<1>Flynn, J., Glickman, J.F., Reich, N.O.
<2>Murine DNA cytosine-C5 methyltransferase: pre-steady- and steady-state kinetic analysis with regulatory DNA sequences.
<3>Biochemistry
<4>35
<5>7308-7315
<6>1996
<7>We present the first description of KmDNA, KdDNA, kcat, and kmethylation for a mammalian DNA
methyltransferase.  Homogeneous, 190 000 MrDNA (cytosine-5-)-methyltransferase isolated from
mouse erythroleukemia cells has turnover constants of 0.15-0.59 h-1 with single-stranded and
unmethylated double-stranded oligonucleotides containing a single CpG dinucleotide.  These
substrates were designed to mimic DNA transcriptional cis elements previously reported to have
cytosine C-5-methylated regulation.  The rate-limiting step for these substrates is the
methylation step itself.  In contrast, hemimethylated double-stranded substrates show burst
kinetics, consistent with a rapid methylation event (3 h-1) followed by a slower step which
determines steady-state kcat.  Hemimethylated and unmethylated double-stranded DNA shows
similar binding affinities; these results reveal the molecular basis for the enzyme's
preference for hemimethylated DNA to be the methyl transfer step.  Substrates with multiple
recognition sites do not show burst kinetics and have turnover rate constants of 6 h-1.
Catalytic turnover for the mammalian enzyme is thus approximately 10-fold slower than that for
the related bacterial enzymes.  Our combined results show quantitatively that one enzyme is
certainly capable of both maintenance and de novo methylation and that maintenance of the
genomic methylation pattern is preferred over the de novo establishment of new patterns.
Direct comparison of the mammalian enzyme with the bacterial DNA cytosine-C5
methyltransferase, M.SssI, indicates dramatic differences in preferences for single-stranded,
double-stranded, and hemimethylated double-stranded substrates.  Moreover, the specificity
hierarchy shown for the M.SssI is derived from very different changes in Km and catalysis than
those observed for the mammalian DCMTase.  These results demonstrate that the M.SssI, and
perhaps other DNA cytosine methyltransferases from bacteria, is functionally dissimilar to the
mammalian enzyme.

<>

<1>Flynn, J., Reich, N.
<2>Murine DNA (cytosine-5-)-methyltransferase: Steady-state and substrate trapping analyses of the kinetic mechanism.
<3>Biochemistry
<4>37
<5>15162-15169
<6>1998
<7>DNA (cytosine-5-)-methyltransferase is essential for viable mammalian development and has a
central function in the determination and maintenance of epigenetic methylation patterns.
Steady-state and substrate trapping studies were performed to better understand how the enzyme
functions.  The catalytic efficiency was dependent on substrate DNA length.  A 14-fold
increase in KmDNA was observed as the length decreased from 5000 to 100 base pairs and kcat
decreased by a third.  Steady-state analyses were used to identify the order of substrate
addition onto the enzyme and the order of product release.  Double-reciprocal patterns of
velocity versus substrate concentration intersected far from the origin and were nearly
parallel.  The kinetic mechanism does not appear to change when the DNA substrate is either
6250 or 100 base pairs in length.  Isotope trapping studies showed that the initial enzyme -
AdoMet complex was not catalytically competent; however, the initial enzyme-poly(dI.dC-dI.dC)
complex was observed to be competent for catalysis.  Product inhibition studies also support a
sequential ordered bi-bi kinetic mechanism in which DNA binds to the enzyme first, followed by
S-adenosyl-L-methionine, and then the products S-adenosyl-L-homocysteine and methylated DNA
are released.  The proposed mechanism is similar to the mechanism proposed for M.HhaI, a
bacterial DNA (cytosine-5-)-methyltransferase.  Evidence for an enzyme - DNA-DNA ternary
complex is also presented.

<>

<1>Flynn, J., Reich, N.O.
<2>Identification of flanking sequence dependence of DNA recognition for mammalian DNA cytosine methyltransferase.
<3>FASEB J.
<4>7
<5>A1219
<6>1993
<7>Mammalian DNA cytosine 5-methyltransferase (DNA Mtase) is a DNA modification enzyme which has
been implicated to have an important role in gene regulation. Cytosines in the context of the
dinucleotide CpG are differentially methylated in the different tissue types of higher
eukaryotes. The pattern of methylation throughout the genome is believed to be heritable and
important for the development of differentiated cell types. The aim of our studies are to
answer to what extent sequences flanking a CpG determine the binding and catalytic activity of
DNA Mtase. Our procedure uses a random population of oligonucleotide substrates which contain
a central CpG dinucleotide flanked by 12 base pair degenerate regions on each side. The
degenerate regions have been made such that no additional CpG's can be introduced into the
oligos and contain equivalent proportions of all permutations. Partially purified enzyme was
used to create ternary complexes with cofactors and 32P end labeled DNA. Complexes using the
cofactors S-Adenosyl methionine, S-adenosyl homocysteine or sinefungin were shown to mobility
shift the double stranded DNA in polyacrylamide gels. The addition of DNA Mtase specific
antibodies to the reaction supershifts the DNA to a less mobile complex. The shifted DNA was
isolated and amplified by the polymerase chain reaction. When subjecting this DNA to further
rounds of selection preferential DNA flanking sequences survive. Through this application of
in vitro genetics CpG flanking sequence dependencies are being resolved.

<>

<1>Foerg, E., Saporito, L., Huang, S., Yang, J., Allen, M.M.
<2>Isolation and characterization of two sequence-specific endonucleases from the cyanobacterium Synechocystis sp. PCC 6308.
<3>FEMS Microbiol. Lett.
<4>69
<5>105-108
<6>1990
<7>The first two restriction endonucleases to be characterized in the cyanobacterium
Synechocystis sp. PCC 6308 are described.  SynI, an AvaII isoschizomer, recognizes the base
sequence 5'-GG[AT]CC-3'.  SynII, an XmnI isoschizomer, recognizes the sequence
5'-GAANNNNTTC-3'.

<>

<1>Foley, S., Bruttin, A., Brussow, H.
<2>Widespread distribution of a group I intron and its three deletion derivatives in the lysin gene of streptococcus thermophilus bacteriophages.
<3>J. Virol.
<4>74
<5>611-618
<6>2000
<7>Of 62 Streptococcus thermophilus bacteriophages isolated from various ecological settings,
half contain a lysin gene interrupted by a group IA2 intron. Phage mRNA splicing was
demonstrated. Five phages possess a variant form of the intron resulting from three distinct
deletion events located in the intron-harbored open reading frame (orf 253). The predicted orf
253 gene sequence showed a significantly lower GC content than the surrounding intron and
lysin gene sequences, and the predicted protein shared a motif with endonucleases found in
phages from both gram-positive and gram-negative bacteria. A comparison of the phage lysin
genes revealed a clear division between intron-containing and intron-free alleles, leading to
the establishment of a 14-bp consensus sequence associated with intron possession. The
conserved intron was not found elsewhere in the phage or S. thermophilus bacterial genomes.
Folding of the intron RNA revealed secondary structure elements shared with other phage
introns: first, a 38-bp insertion between regions P3 and P4 that can be folded into two
stem-loop structures (shared with introns from Bacillus phage SPO1 and relatives); second, a
conserved P7.2 region (shared with all phage introns); third, the location of the stop codon
from orf 253 in the P8 stem (shared with coliphage T4 and Bacillus phage SPO1 introns);
fourth, orf 253, which has sequence similarity with the H-N-H motif of putative endonuclease
genes found in introns from Lactococcus, Lactobacillus, and Bacillus phages.

<>

<1>Follman, H., Balzer, H.-J., Schleicher, R.
<2>Biosynthesis and distribution of methylcytosine in wheat DNA.  How different are plant DNA methyltransferases?
<3>Nucleic Acid Methylation, Wiley-Liss, Clawson, G.A., Willis, D.B., Weissbach, A., Jones, P.A., New York
<4>0
<5>199-210
<6>1990
<7>One reason for the high methylcytosine content of plant DNA may lie in the
specificity and activity of plant DNA methyltransferases.  We have analyzed the
properties of the DNA methylase system previously purified from wheat embryo
and find that it differs markedly enough from the known mammalian enzymes to
establish a plant-specific type of DNA methyltransferase.  These differences
reside, inter alia, in molecular weight, specificity towards DNA substrates of
varying methylation, and insensitivity towards inhibition by 5-azacytidine.

<>

<1>Fomenkov, A., Clark, T., Spittle, K., Anton, B.M., Vincze, T., Korlach, J., Roberts, R.J.
<2>Molecular dissection of the methylome of Burkholderia cenocepacia J2315.
<3>FEBS J.
<4>280
<5>72-73
<6>2013
<7>B. cenocepacia is a pathogenic gram-negative bacterium that often causes an opportunistic
infection in cystic fibrosis patients.  It is highly antibiotic-resistant, transmissible, and
often lethal.  Although genome sequences of several strains are available, the study of this
organism, like many pathogens, has been relatively limited, and genetic study is difficult.
The aim of this work was to analyze the genomic DNA methylation patterns from the sequenced
strain, J2315, and identify specificities of putative DNA methyltransferases.  This might
facilitate the development of an efficient transformation system to enhance genetic studies of
this strain.  We took advantage of the recently developed platform for single-molecule real
time sequencing by Pacific Biosciences.  This next generation sequencing technology allowed us
not only to perform high throughput DNA sequencing, but also to identify the epigenetic status
of the DNA using the polymerase's kinetic signature on the template during the reaction.  The
kinetic signature analysis of B. cenocepacia J2315 genomic DNA revealed three modified motifs.
Additionally, a number of ORF's encoding putative DNA methyltransferases were cloned into
plasmid vectors, and their specificities were determined using SMRT sequecing.  A type III
restriction-modificaiton system called M.BceJI resulted in m6A modification in the recognition
sequence CA-CAG.  98.4% of all CACAG sequences in the genome were modified on just one strand
as indicated, which is typical of a Type III methyltransferase.  Cloning ORF BCAL3494 showed
it to be the gene responsible.  The genomic motif GTWWAC, containing a symmetrical
double-stranded m6A modification, was 95.7% modified in the genome and was caused by the
product of ORF BCAL992 and was called M.BceJIV.  Another modified motif with m4C modification
was detected at the low rate of 4.8% within the eight base pair motif GCGGCCGC.  This was
probably caused by the product of ORF BCAL1036, although when this was cloned and expressed at
high levels the central tetranucleotide GGCC was m4C modified at high levels.  We called this
methyltransferase M.BceJII.  Surprisingly, we also detected an unusual kinetic signature on
the G base that is also associated with the four base core of the GCGGCCGC motif.  We still do
not know which enzyme is responsible for this G modification.  Preliminary results suggest
that M.BceJORF178P might also lead to G modification on the first base in the sequence GGNNTA
albeit with a low IPD ratio in vitro.  This enzyme is highly toxic in E. coli, which has so
far made a determination of its specificity very difficult.  One rather interesting
methyltransferase is encoded by ORF pBCA072, which is located on a plasmid.  It results in
single-stranded, non-specific m6A modification, but was only detected on plasmid DNA.  We have
called this enzyme M.BceJIII.  A similar enzyme has been found on an E. coli plasmid and both
could possibly play a restricted role during DNA replication.  The M and S subunits of a
putative Type I restriction-modification system BceJORF418P have been cloned, but no
modifications were detected either on genomic or plasmid DNA indicating that the system is
probably inactive in this strain.

<>

<1>Fomenkov, A., Dila, D., Raleigh, E.A., Xu, S.-Y.
<2>A method for direct cloning of nuclease genes in E. coli.
<3>International Patent Office
<4>WO 9532281
<5>
<6>1995
<7>The present invention discloses a novel method for the direct cloning of nuclease
genes such as restriction endonuclease genes in E. coli.  In addition, there is provided a
novel
strain which facilitates application of the method.  This method has been successfully
employed to
clone a number of genes coding for endonuclease including restriction endonuclease genes.

<>

<1>Fomenkov, A., Dila, D.K., Raleigh, E.A., Xu, S.-Y.
<2>A method for direct cloning of nuclease genes in E. coli.
<3>European Patent Office
<4>EP 1431388 A
<5>
<6>2004
<7>The present invention discloses a novel method for the direct cloning of nuclease genes such
as restriction endonuclease genes in E. coli.  In addition, there is provided a novel strain
which facilitates application of the method.  This method has been successfully employed to
clone a number of genes coding for endonuclease including restriction endonuclease genes.

<>

<1>Fomenkov, A., Xiao, J.-P., Dila, D., Raleigh, E., Xu, S.-Y.
<2>The 'endo-blue method' for direct cloning of restriction endonuclease genes in E. coli.
<3>Nucleic Acids Res.
<4>22
<5>2399-2403
<6>1994
<7>A new E. coli strain has been constructed that contains the dinD1::LacZ+ fusion and is
deficient in methylation-dependent restriction systems (McrA-, McrBC-, Mrr-). This strain has
been used to clone restriction endonuclease genes directly into E. coli. When E. coli cells
are not fully protected by the cognate methylase, the restriction enzyme damages the DNA in
vivo and induces the SOS response. The SOS-induced cells form blue colonies on indicator
plates containing X-gal. Using this method the genes coding for the thermostable restriction
enzymes TaqI (5'TCGA3') and Tth111I (5'GACNNNGTC3') have been successfully cloned in E.
coli. The new strain will be useful to clone other genes involved in DNA metabolism.

<>

<1>Fomenkov, A., Xu, S.-Y.
<2>Isolation of temperature-sensitive mutants of the BamHI restriction endonuclease.
<3>Gene
<4>157
<5>303-310
<6>1995
<7>Two heat-sensitive R.BamHI mutants, T157I and P173L, and one cold-sensitive R.BamHI mutant,
T114I, were isolated after chemical mutagenesis of the bamhIR gene that codes for the
restriction endonuclease BamHI (R.BamHI).  The thermosensitivity of T114I, T157I and P173L is
revealed by the 10/2-10/3 lower plating efficiency at the non-permissive temperature of
strains bearing these alleles.  The conditional-lethal phenotype can be rescued by
introduction of the cognate bamhIM gene into the same cell.  The mutant enzymes induce the SOS
response in vivo and display reduced phage restriction activity.  The P173L protein, when
expressed at 30oC and purified, shows reduced thermostability at 65oC.  T157I and P173L
mutants yield different intermediates during partial trypsin digestion.  The
conditional-lethal BamHI mutants could be used to deliver in vivo DNA cleavage and for further
isolation of relaxed-specificity mutants.

<>

<1>Fomenkov, A.I., Kramarov, V.M., Andreev, L.V., Mochalov, V.V., Smolyaninov, V.V., Matvienko, N.I.
<2>Isolation and properties of a new site specific endonuclease Bme142I from Bacillus megaterium 142.
<3>Nucleic Acids Res.
<4>16
<5>10399
<6>1988
<7>A new type II restriction endonuclease, Bme142I, was partially purified and characterized from
Bacillus megaterium 142.  It has been seen in a crude extract, because this bacterial strain
does not have significant amounts of contaminating nucleases.  The enzyme was purified by
chromatography on phosphocellulose P11 (elution buffer - 10 mM K-phosphate, pH 7.0, 0.1 mM
EDTA, 2 mM DTT, 0.2 - 1 M NaCl) and hydroxyapatite (elution buffer - 0.01 - 0.5 M K-phosphate,
pH 7.0, 1 mM EDTA, 2 mM DTT).  Yield of enzyme is 2000 units per gram of wet cells.  The
enzyme is stable in elution buffer and may be stored at -20C after addition of glycerol to
50%.  The recognition site was determined from the cleavage pattern of pBR322 plasmid.  The
5'-end nucleotide is G.  This has been shown by incubation of the 5'-end labelled DNA
fragments with nuclease P1 followed by thin-layer chromatography on a PEI cellulose plate.
The points of cleavage of the recognition site were determined by the Maxam-Gilbert method.
In contrast to its isoschizomer, the restriction endonuclease HaeII, which cleaves the same
recognition site to produce a 4 base 3' extension, the Bme142I produces blunt-ended DNA
fragments:
5' PuGC^GCPy 3'
3' PyCG^CGPu 5'.

<>

<1>Fondi, M., Orlandini, V., Emiliani, G., Papaleo, M.C., Maida, I., Perrin, E., Vaneechoutte, M., Dijkshoorn, L., Fani, R.
<2>Draft Genome Sequence of the Hydrocarbon-Degrading and Emulsan-Producing Strain Acinetobacter venetianus RAG-1T.
<3>J. Bacteriol.
<4>194
<5>4771-4772
<6>2012
<7>We report the draft genome sequence of Acinetobacter venetianus strain RAG-1(T),  which is
able to degrade hydrocarbons and to synthesize a powerful biosurfactant
(emulsan) that can be employed for oil removal and as an adjuvant for vaccine
delivery. The genome sequence of A. venetianus RAG-1(T) might be useful for
bioremediation and/or clinical purposes.

<>

<1>Fondi, M., Orlandini, V., Maida, I., Perrin, E., Papaleo, M.C., Emiliani, G., de Pascale, D., Parrilli, E., Tutino, M.L., Michaud, L., Lo, G.A., Fani, R.
<2>Draft Genome Sequence of the Volatile Organic Compound-Producing Antarctic Bacterium Arthrobacter sp. Strain TB23, Able To Inhibit Cystic Fibrosis Pathogens  Belonging to the Burkholderia cepacia Complex.
<3>J. Bacteriol.
<4>194
<5>6334-6335
<6>2012
<7>Arthrobacter sp. strain TB23 was isolated from the Antarctic sponge Lissodendoryx nobilis.
This bacterium is able to produce antimicrobial compounds and volatile
organic compounds (VOCs) that inhibit the growth of other Antarctic bacteria and
of cystic fibrosis opportunistic pathogens, respectively. Here we report the
draft genome sequence of Arthrobacter sp. TB23.

<>

<1>Fonfara, I., Curth, U., Pingoud, A., Wende, W.
<2>Creating highly specific nucleases by fusion of active restriction endonucleases and catalytically inactive homing endonucleases.
<3>Nucleic Acids Res.
<4>40
<5>847-860
<6>2012
<7>Zinc-finger nucleases and TALE nucleases are produced by combining a specific DNA-binding
module and a non-specific DNA-cleavage module,
resulting in nucleases able to cleave DNA at a unique sequence. Here a new
approach for creating highly specific nucleases was pursued by fusing a
catalytically inactive variant of the homing endonuclease I-SceI, as DNA
binding-module, to the type IIP restriction enzyme PvuII, as cleavage
module. The fusion enzymes were designed to recognize a composite site
comprising the recognition site of PvuII flanked by the recognition site
of I-SceI. In order to reduce activity on PvuII sites lacking the flanking
I-SceI sites, the enzymes were optimized so that the binding of I-SceI to
its sites positions PvuII for cleavage of the composite site. This was
achieved by optimization of the linker and by introducing amino acid
substitutions in PvuII which decrease its activity or disturb its dimer
interface. The most specific variant showed a more than 1000-fold
preference for the addressed composite site over an unaddressed PvuII
site. These results indicate that using a specific restriction enzyme,
such as PvuII, as cleavage module, offers an alternative to the otherwise
often used catalytic domain of FokI, which by itself does not contribute
to the specificity of the engineered nuclease.

<>

<1>Fontana, M.R., Pizza, M., Masiganani, V., Monaci, E.
<2>Gonococcal proteins and nucleic acids.
<3>US Patent Office
<4>US 7504111 A
<5>
<6>2009
<7>The invention provides proteins from gonococcus (Neisseria gonorrhoeae), including amino acid
sequences, the corresponding nucleotide sequences, expression data, and serological data.  The
proteins are useful antigens for vaccines, immunogenic compositions, and/or diagnostics.  They
are also useful for distinguishing between gonococcus and meningococcus and, in particular,
between genococcus and serogroup B meningococcus.

<>

<1>Fontana, M.R., Pizza, M., Masignani, V., Monaci, E.
<2>Gonococcal proteins and nucleic acids.
<3>European Patent Office
<4>EP 1777233 A
<5>
<6>2007
<7>The invention provides proteins from genococcus (Neisseria gonorrhoeae), including amino acid
sequences, the corresponding nucleotide sequences, expression data, and serological data.  The
proteins are useful antigens for vaccines, immunogenic compositions, and/or diagnostics.  They
are also useful for distinguishing between gonococcus and meningococcus and, in particular,
between gonococcus and serogroup B meningococcus.

<>

<1>Fookes, M., Yu, J., De Majumdar, S., Thomson, N., Schneiders, T.
<2>Genome Sequence of Klebsiella pneumoniae Ecl8, a Reference Strain for Targeted Genetic Manipulation.
<3>Genome Announcements
<4>1
<5>e00027-12
<6>2013
<7>We report the genome sequence of Klebsiella pneumoniae subsp. pneumoniae Ecl8, a  spontaneous
streptomycin-resistant mutant of strain ECL4, derived from NCIB 418.
K. pneumoniae Ecl8 has been shown to be genetically tractable for targeted gene
deletion strategies and so provides a platform for in-depth analyses of this
species.

<>

<1>Foray, V., Grigorescu, A.S., Sabri, A., Haubruge, E., Lognay, G., Francis, F., Fauconnier, M.L., Hance, T., Thonart, P.
<2>Whole-Genome Sequence of Serratia symbiotica Strain CWBI-2.3T, a Free-Living Symbiont of the Black Bean Aphid Aphis fabae.
<3>Genome Announcements
<4>2
<5>e00767-14
<6>2014
<7>The gammaproteobacterium Serratia symbiotica is one of the major secondary symbionts found in
aphids. Here, we report the draft genome sequence of S.
symbiotica strain CWBI-2.3(T), previously isolated from the black bean aphid
Aphis fabae. The 3.58-Mb genome sequence might provide new insights to understand
the evolution of insect-microbe symbiosis.

<>

<1>Ford, E., Boyer, H.W.
<2>Degradation of enteric bacterial deoxyribonucleic acid by the Escherichia coli B restriction endonuclease.
<3>J. Bacteriol.
<4>104
<5>594-595
<6>1970
<7>the deoxyribonucleic acid of five different genera of enteric microorganisms
was shown to be degraded by the Escherichia coli B restriction endonuclease.

<>

<1>Ford, K., Taylor, C., Connolly, B., Hornby, D.P.
<2>Effects of co-factor and deoxycytidine substituted oligonucleotides upon sequence-specific interations between MspI DNA methyltransferase and DNA.
<3>J. Mol. Biol.
<4>230
<5>779-786
<6>1993
<7>MspI methyltransferase (M.MspI) catalyses the transfer of a methyl group from
S-adenosyl-L-methionine to the C-5 position of the outer deoxycytidine base in the DNA
sequence 5'-CCGG-3'. Recombinant M.MspI when expressed and purified as a translational
fusion with glutathione-S-transferase, shows all of the properties of the wild-type enzyme. We
report the kinetic analysis of M.MspI binding to DNA, which suggests a two-stage methylation
process, whose initial DNA binding rate is governed by the presence of a positively charged
sulphonium centre of the coafactor. Results are also presented that indicate that M.MspI binds
preferentially to hemi-methylated DNA and that full methylation of either deoxycytidine on
both strands significantly impairs sequence-specific protein-DNA interactions. Furthermore,
the importance of the 4-amino group of the inner deoxycytidine for sequence-specific
protein-DNA interactions is demonstrated by substituting deoxycytidine with
2-pyrimidinone-1-beta-D-2-deoxyriboside. In addition, we detail the intrinsic structural
elements of a cofactor, required to enhance the binding of M.MspI to its recognition sequence
by using S-adenosyl-L-methionine and a range of derivatives.

<>

<1>Forde, A., Daly, C., Fitzgerald, G.F.
<2>Identification of four phage resistance plasmids from Lactococcus lactis subsp. cremoris HO2.
<3>Appl. Environ. Microbiol.
<4>65
<5>1540-1547
<6>1999
<7>The bacteriophage-host sensitivity patterns of 16 strains of Lactococcus lactis originally
isolated from a mixed strain Cheddar cheese starter culture were determined.  Using phages
obtained from cheese factory whey, four of the strains were found to be highly phage
resistant.  One of these isolates, Lactococcus lactis subsp. cremoris HO2, was studied in
detail to determine the mechanisms responsible for the phage insensitivity phenotypes.
Conjugal transfer of plasmid DNA from strain HO2 allowed a function to be assigned to four of
its six plasmids.  A 460kb molecule, designated pCI646, was found to harbor the lactose
utilization genes, while this and plasmids of 58 kb (pCI658), 42 kb (pCI642), and 4.5 kb
(pCI605) were shown to be responsible for the phage resistance phenotypes observed against the
small isometric-headed phage omega-712 (936 phage species) and the prolate-headed phage
omega-c2 (c2 species).  PCI658 was found to mediate an adsorption-blocking mechanism and was
also responsible for the fluffy pellet phenotype of cells containing the molecule.  PCI642 and
pCI605 were both shown to be required for the operation of a restriction-modification system.

<>

<1>Forde, A., Fitzgerald, G.F.
<2>Bacteriophage defence systems in lactic acid bacteria.
<3>Antonie Van Leeuwenhoek
<4>76
<5>89-113
<6>1999
<7>The study of the interactions between lactic acid bacteria and their bacteriophages has been a
vibrant and rewarding research activity for a considerable number of years. In the more recent
past, the application of molecular genetics for the analysis of phage-host relationships has
contributed enormously to the unravelling of specific events which dictate insensitivity to
bacteriophage infection and has revealed that while they are complex and intricate in nature,
they are also extremely effective. In addition, the strategy has laid solid foundations for
the construction of phage resistant strains for use in commercial applications and has
provided a sound basis for continued investigations into existing, naturally-derived and
novel, genetically-engineered defence systems. Of course, it has also become clear that phage
particles are highly dynamic in their response to those defence systems which they do
encounter and that they can readily adapt to them as a consequence of their genetic
flexibility and plasticity. This paper reviews the exciting developments that have been
described in the literature regarding the study of phage-host interactions in lactic acid
bacteria and the innovative approaches that can be taken to exploit this basic information for
curtailing phage infection.

<>

<1>Forde, B.M., Neville, B.A., O'Donnell, M.M., Riboulet-Bisson, E., Claesson, M.J., Coughlan, A., Ross, R.P., O'Toole, P.W.
<2>Genome sequences and comparative genomics of two Lactobacillus ruminis strains from the bovine and human intestinal tracts.
<3>Microb. Cell Fact.
<4>10
<5>S13
<6>2011
<7>Background: The genus Lactobacillus is characterized by an extraordinary degree of phenotypic
and genotypic
diversity, which recent genomic analyses have further highlighted. However, the choice of
species for sequencing
has been non-random and unequal in distribution, with only a single representative genome from
the L. salivarius
clade available to date. Furthermore, there is no data to facilitate a functional genomic
analysis of motility in the
lactobacilli, a trait that is restricted to the L. salivarius clade.
Results: The 2.06 Mb genome of the bovine isolate Lactobacillus ruminis ATCC 27782 comprises a
single circular
chromosome, and has a G+C content of 44.4%. In silico analysis identified 1901 coding
sequences, including genes
for a pediocin-like bacteriocin, a single large exopolysaccharide-related cluster, two sortase
enzymes, two CRISPR
loci and numerous IS elements and pseudogenes. A cluster of genes related to a putative pilin
was identified, and
shown to be transcribed in vitro. A high quality draft assembly of the genome of a second L.
ruminis strain, ATCC
25644 isolated from humans, suggested a slightly larger genome of 2.138 Mb, that exhibited a
high degree of
synteny with the ATCC 27782 genome. In contrast, comparative analysis of L. ruminis and L.
salivarius identified a
lack of long-range synteny between these closely related species. Comparison of the L.
salivarius clade core
proteins with those of nine other Lactobacillus species distributed across 4 major
phylogenetic groups identified
the set of shared proteins, and proteins unique to each group.
Conclusions: The genome of L. ruminis provides a comparative tool for directing functional
analyses of other
members of the L. salivarius clade, and it increases understanding of the divergence of this
distinct Lactobacillus
lineage from other commensal lactobacilli. The genome sequence provides a definitive resource
to facilitate
investigation of the genetics, biochemistry and host interactions of these motile intestinal
lactobacilli.

<>

<1>Forde, G.K., Kedzierski, P., Sokalski, W.A., Forde, A.E., Hill, G.A., Leszczynski, J.
<2>Physical nature of interactions within the active site of cytosine-5-methyltransferase.
<3>J. Phys. Chem. A
<4>110
<5>2308-2313
<6>2006
<7>The physical nature of interactions within the active site of cytosine-5-methyltransferase
(CMT) was studied using a
variation-perturbation energy decomposition scheme defining a sequence
of approximate intermolecular interaction energy models. These models
have been used to analyze the catalytic activity of residues
constituting cytosine-5-methyltransferase active site as well their
role in the binding group of de novo designed inhibitors. Our results
indicate that Glu119, Arg163, and Arg165 appear to play the dominant
role in stabilizing the protonated transition state structure and their
influence can be qualitatively approximated by electrostatic
interactions alone. The stabilization of neutral structures of the
alternative reaction pathway is small, which might suggest the
protonated pathway as preferred by the enzyme. Exchange and
delocalization terms are negligible in most cases, or they cancel each
other to some extent. Interactions of inhibitors with the CMT active
site are dominated by electrostatic multipole contributions in analogy
with previously studied transition state analogue inhibitors of leucyl
aminopeptidase.

<>

<1>Foret, S., Kucharski, R., Pellegrini, M., Feng, S., Jacobsen, S.E., Robinson, G.E., Maleszka, R.
<2>DNA methylation dynamics, metabolic fluxes, gene splicing, and alternative phenotypes in honey bees.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>109
<5>4968-4973
<6>2012
<7>In honey bees (Apis mellifera), the development of a larva into either a queen or worker
depends on differential feeding with royal jelly and involves epigenomic
modifications by DNA methyltransferases. To understand the role of DNA
methylation in this process we sequenced the larval methylomes in both queens and
workers. We show that the number of differentially methylated genes (DMGs) in
larval head is significantly increased relative to adult brain (2,399 vs. 560)
with more than 80% of DMGs up-methylated in worker larvae. Several highly
conserved metabolic and signaling pathways are enriched in methylated genes,
underscoring the connection between dietary intake and metabolic flux. This
includes genes related to juvenile hormone and insulin, two hormones shown
previously to regulate caste determination. We also tie methylation data to
expressional profiling and describe a distinct role for one of the DMGs encoding
anaplastic lymphoma kinase (ALK), an important regulator of metabolism. We show
that alk is not only differentially methylated and alternatively spliced in Apis,
but also seems to be regulated by a cis-acting, anti-sense non-protein-coding
transcript. The unusually complex regulation of ALK in Apis suggests that this
protein could represent a previously unknown node in a process that activates
downstream signaling according to a nutritional context. The correlation between
methylation and alternative splicing of alk is consistent with the recently
described mechanism involving RNA polymerase II pausing. Our study offers
insights into diet-controlled development in Apis.

<>

<1>Formighieri, E.F. et al.
<2>The mitochondrial genome of the phytopathogenic basidiomycete Moniliophthora perniciosa is 109 kb in size and contains a stable integrated plasmid.
<3>Mycol. Res.
<4>112
<5>1136-1152
<6>2008
<7>We present here the sequence of the mitochondrial genome of the basidiomycete phytopathogenic
hemibiotrophic fungus Moniliophthora perniciosa, causal agent of the Witches' Broom Disease
in Theobroma cacao.
The DNA is a circular molecule of 109,103 base pairs, with 31.9% GC, and is the largest
sequenced so far. This size is due essentially to the presence of numerous non-conserved
hypothetical ORFs. It contains the 14 genes coding for proteins involved in the oxidative
phosphorylation, the two rRNA genes, one ORF coding for a ribosomal protein (rps3), and a set
of 26 tRNA genes that recognize codons for all amino acids. Seven homing endonucleases are
located inside introns. Except atp8, all conserved known genes are in the same orientation.
Phylogenetic analysis based on the cox genes agrees with the commonly accepted fungal
taxonomy. An uncommon feature of this mitochondrial genome is the presence of a region that
contains a set of four, relatively small, nested, inverted repeats enclosing two genes coding
for polymerases with an invertron-type structure and three conserved hypothetical genes
interpreted as the stable integration of a mitochondrial linear plasmid. The integration of
this plasmid seems to be a recent evolutionary event that could have implications in fungal
biology. This sequence is available under GenBank accession number AY376688.

<>

<1>Formusa, P.A., Hsiang, T., Habash, M.B., Lee, H., Trevors, J.T.
<2>Genome Sequence of Pseudomonas mandelii PD30.
<3>Genome Announcements
<4>2
<5>e00713-14
<6>2014
<7>The genome sequence of Pseudomonas mandelii PD30 is reported in this announcement. The genes
for the reduction of nitrate to dinitrogen were
identified in the genome assembly and subsequently used in gene expression
research.

<>

<1>Forrow, S., Lee, M., Souhami, R.L., Hartley, J.A.
<2>The effect of AT and GC sequence specific minor groove binding agents on restriction endonuclease activity.
<3>ACS Abstracts
<4>208
<5>97
<6>1994
<7>The naturally occuring DNA minor groove-binders, netropsin and distamycin A, recognize (A/T)4
and (A/T)5 sequences respectively. These ligands have well-documented effects on proteins such
as restriction endonucleases and transcription factors whose DNA recognition sequences have
high A/T content. We have investigated the ability of two synthetic imidazole containing and
G/C selective oligopeptides to interfere with the catalytic activity of restriction
endonucleases, and compared this with the effects of netropsin and distamycin. The
endonucleases were chosen to have either A/T rich (EcoRI, EcoRV) or G/C rich (BalI, NruI,
Fnu4HI, BanII) recognition sequences. An agarose gel assay was used to measure the degree of
cleavage of 32P-labelled DNA in the presence or absence of minor groove-binding ligand, and
ligand-DNA binding data was obtained using methidium-propyl EDTA (MPE) footprinting
methodology. The results from these studies will be presented.

<>

<1>Forrow, S.M., Lee, M., Souhami, R.L., Hartley, J.A.
<2>The effect of AT and GC sequence specific minor groove-binding agents on restriction endonuclease activity.
<3>Chem. Biol. Interact.
<4>96
<5>125-142
<6>1995
<7>The ability of the naturally occurring A/T specific DNA minor groove binders netropsin and
distamycin A and two synthetic G/C selective oligopeptide analogues (1 and 2), to interfere
with the catalytic activity of restriction endonucleases has been investigated. Enzymes were
chosen to have A/T rich (EcoRI, EcoRV) or G/C rich (BalI, NruI) recognition sequences. An
agarose gel assay was used to measure the cleavage of 32P-labelled DNA and ligand-DNA binding
data was obtained using methidium-propyl EDTA footprinting. Netropsin and distamycin bind at
the recognition sites, and dose-dependently inhibited cleavage by, EcoRI and EcoRV, (EcoRI >
EcoRV). They were also more effective at inhibiting the catalytic activity of BalI than either
1 or 2. NruI was inhibited by distamycin and 2, but not by netropsin or 1. DNA footprinting
revealed that neither 1 or 2 bound to the BalI or NruI recognition sequences under the
conditions used whereas netropsin and distamycin footprint at adjacent sites. 1 binds to two
of the three recognition sequences for the enzyme Fnu4HI (GCNGC) in the fragment studied and
was shown to inhibit DNA cleavage only at these two sites. 2 binds strongly to two GGGCTC
sequences which are recognition sites for the enzyme BanII. In this case a pronounced
stimulation of cleavage was observed in the presence of 2 over a wide dose range. The results
indicate that enzyme inhibition does not necessarily result from simultaneous occupancy of a
common site, or at nearby flanking sequences, and in some circumstances, a pronounced
stimulation of enzyme cleavage can occur.

<>

<1>Forsberg, K.J., Patel, S., Gibson, M.K., Lauber, C.L., Knight, R., Fierer, N., Dantas, G.
<2>Bacterial phylogeny structures soil resistomes across habitats.
<3>Nature
<4>509
<5>612-616
<6>2014
<7>Ancient and diverse antibiotic resistance genes (ARGs) have previously been
identified from soil, including genes identical to those in human pathogens.
Despite the apparent overlap between soil and clinical resistomes, factors
influencing ARG composition in soil and their movement between genomes and
habitats remain largely unknown. General metagenome functions often correlate
with the underlying structure of bacterial communities. However, ARGs are
proposed to be highly mobile, prompting speculation that resistomes may not
correlate with phylogenetic signatures or ecological divisions. To investigate
these relationships, we performed functional metagenomic selections for
resistance to 18 antibiotics from 18 agricultural and grassland soils. The 2,895
ARGs we discovered were mostly new, and represent all major resistance
mechanisms. We demonstrate that distinct soil types harbour distinct resistomes,
and that the addition of nitrogen fertilizer strongly influenced soil ARG
content. Resistome composition also correlated with microbial phylogenetic and
taxonomic structure, both across and within soil types. Consistent with this
strong correlation, mobility elements (genes responsible for horizontal gene
transfer between bacteria such as transposases and integrases) syntenic with ARGs
were rare in soil by comparison with sequenced pathogens, suggesting that ARGs
may not transfer between soil bacteria as readily as is observed between human
pathogens. Together, our results indicate that bacterial community composition is
the primary determinant of soil ARG content, challenging previous hypotheses that
horizontal gene transfer effectively decouples resistomes from phylogeny.

<>

<1>Forsblom, S., Rigler, R., Ehrenberg, M., Pettersson, U., Philipson, L.
<2>Kinetic studies on the cleavage of adenovirus DNA by restriction endonuclease EcoRI.
<3>Nucleic Acids Res.
<4>3
<5>3255-3269
<6>1976
<7>The kinetics of cleavage of DNA from Adenovirus Type 1 (Ad1), Type 5 (Ad5) and
Type 6 (Ad6) by restriction endonuclease EcoRI was investigated by quantitative
evaluation of the fluorescence from ethidium stained DNA fragments separated on
agarose gels.  The apparent rate constants of cleavage at different cleavage
sites have been determined and large differences in the cleavage sites of the
individual sites within one type of DNA were found.  From the kinetics of
cleavage information on the sequence of the DNA fragments can be obtained.  The
order of the fragments A,B,C,D of Ad6 DNA obtained after complete cleavage by
restriction endonuclease EcoRI was found to be ADCB; the order of the
corresponding fragments A,B,C of Ad1 and Ad5 DNA was found to be ACB.

<>

<1>Forsyth, R.A., Ohlsen, K., Zyskind, J.
<2>Genes essential for microbial proliferation and antisense thereto.
<3>International Patent Office
<4>WO 0134810 A
<5>
<6>2001
<7>The sequences of nucleic acids encoding proteins required for E. coli proliferation are
disclosed.  The nucleic acids can be used to express proteins or portions thereof, to obtain
antibodies capable of specifically binding to the expressed proteins, and to use those
expressed proteins as a screen to isolate candidate molecules for rational drug discovery
programs.  The nucleic acids can also be used to screen for homologous genes that are required
for proliferation in microorganisms other than E. coli.  The nucleic acids can also be used to
design expression vectors and secretion vectors.  The nucleic acids of the present invention
can also be used in various assay systems to screen for proliferation required genes in other
organisms as well as to screen for antimicrobial agents.

<>

<1>Forsyth, R.A., Ohlsen, K., Zyskind, J.W.
<2>Genes essential for microbial proliferation and antisense thereto.
<3>US Patent Office
<4>US 6589738 A
<5>
<6>2003
<7>
<>

<1>Foss, H.M., Roberts, C.J., Claeys, K.M., Selker, E.U.
<2>Abnormal chromosome behavior in Neurospora mutants defective in DNA methylation.
<3>Science
<4>262
<5>1737-1741
<6>1993
<7>The function and regulation of DNA methylation in eukaryotes remain unclear. Genes affecting
methylation were identified in the fungus Neurospora crassa. A mutation in one gene, dim-2,
resulted in the loss of all detectable DNA methylation. Abnormal segregation of the
methylation defects in crosses led to the discovery that the methylation mutants frequently
generate strains with extra chromosomes or chromosomal parts. Starvation for
S-adenosylmethionine, the presumed methyl group donor for DNA methylation, also produced
aneuploidy. These results suggest that DNA methylation plays a role in the normal control of
chromosome behavior.

<>

<1>Foster, B. et al.
<2>Complete genome sequence of Xylanimonas cellulosilytica type strain (XIL07T).
<3>Standards in Genomic Sciences
<4>2
<5>1-8
<6>2010
<7>Xylanimonas cellulosilytica Rivas et al. 2003 is the type species of the genus Xylanimonas of
the actinobacterial family Promicromonosporaceae. The species X. cellulosilytica is of
interest because of its ability to hydrolyze cellulose and xylan. Here we describe the
features of this organism, together with the complete genome sequence, and annotation. This is
the first complete genome sequence of a member of the large family Promicromonosporaceae, and
the 3,831,380 bp long genome (one chromosome plus an 88,604 bp long plasmid) with its 3485
protein-coding and 61 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Foster, P.L., Rosche, W.A.
<2>Levels of the Vsr endonuclease do not regulate stationary-phase reversion of a Lac- frameshift allele in Escherichia coli.
<3>J. Bacteriol.
<4>180
<5>1944-1946
<6>1998
<7>Vsr endonuclease, which initiates very short patch repair, has been hypothesized to regulate
mutation in stationary-phase cells.  Overexpression of Vsr does dramatically increase the
stationary-phase reversion of a Lac- frameshift allele, but the absence of Vsr has no effect.
Thus, at least in this case, Vsr has no regulatory role in stationary-phase mutation, and the
effects of Vsr overproduction are likely to be artifactual.

<>

<1>Foster, S., Brummel, K., McDowell, P., Clarke, S.
<2>Antigenic polypeptides.
<3>European Patent Office
<4>EP 1710311 A
<5>
<6>2006
<7>The invention relates to a method for the identification of antigenic polypeptides expressed
by pathogenic microbes; vaccines comprising said polypeptides; recombinant methods to
manufacture said polypeptides; and therapeutic antibodies directed to said polypeptides.

<>

<1>Foster, S., Brummel, K., Mcdowell, P., Clarke, S.
<2>Antigenic polypeptides.
<3>European Patent Office
<4>EP 2269634 A
<5>
<6>2011
<7>The invention relates to a method for the identification of antigenic polypeptides expressed
by pathogenic microbes; vaccines comprising said polypeptides; recombinant methods to
manufacture said polypeptides; and therapeutic antibodies directed to said polypeptides.

<>

<1>Foster, S., Clark, S., Mcdowell, P., Brummell, K., Mond, J.
<2>Antigenic Polypeptides.
<3>Japanese Patent Office
<4>JP 2004536885 A
<5>
<6>2004
<7>
<>

<1>Foster, S., Clarke, S., Mcdowell, P., Brummel, K.
<2>Antigenic polypeptides.
<3>European Patent Office
<4>EP 2287316 A
<5>
<6>2011
<7>The invention relates to a method for the identification of antigenic polypeptides expressed
by pathogenic microbes; vaccines comprising said polypeptides; recombinant methods to
manufacture said polypeptides; and therapeutic antibodies directed to said polypeptides.

<>

<1>Fotso, F.A., Mediannikov, O., Padmanabhan, R., Robert, C., Fournier, P.E., Raoult, D., Drancourt, M.
<2>Genome Sequence of Borrelia crocidurae Strain 03-02, a Clinical Isolate from Senegal.
<3>Genome Announcements
<4>2
<5>e01150-14
<6>2014
<7>The draft genome sequence of Borrelia crocidurae strain 03-02, a blood isolate from a febrile
Senegalese patient, comprises a 920,021-bp linear chromosome
(27.7% G+C content), 32 tRNAs, 818 open reading frames, and one cluster of
regularly interspaced short palindromic repeats. Its genotype differs from that
of the Achema reference strain.

<>

<1>Foulks, J.M., Parnell, K.M., Chau, S., Swierczek, K., Saunders, M., Wright, K., Hendrickson, T.F., McCullar, M.V., Kanner, S.B.
<2>Epigenetic Drug Discovery: Targeting DNA Methyltransferases (vol 17, pg 2, 2012).
<3>J. Biomol. Screen.
<4>17
<5>700
<6>2012
<7>Foulks, J.M.; Parnell, K.M.; Nix, R.N.; Chau, S.; Swierczek, K.; Saunders, M.; Wright, K.;
Hendrickson, T.F.; Ho, K.K.; McCullar, M.V.; Kanner, S.B. Epigenetic Drug Discovery: Targeting
DNA Methyltransferases. J. Biomol. Screen. 2012, 17, 2-17. (Original doi:
10.1177/1087057111421212).

<>

<1>Fournier, P.E., El Karkouri, K., Leroy, Q., Robert, C., Giumelli, B., Renesto, P., Socolovschi, C., Parola, P., Audic, S., Raoult, D.
<2>Analysis of the Rickettsia africae genome reveals that virulence acquisition in Rickettsia species may be explained by genome reduction.
<3>BMC Genomics
<4>10
<5>166
<6>2009
<7>ABSTRACT: BACKGROUND: The Rickettsia genus includes 25 validated species,
17 of which are proven human pathogens. Among these, the pathogenicity
varies greatly, from the highly virulent R. prowazekii, which causes
epidemic typhus and kills its arthropod host, to the mild pathogen R.
africae, the agent of African tick-bite fever, which does not affect the
fitness of its tick vector. RESULTS: We evaluated the clonality of R.
africae in 70 patients and 155 ticks, and determined its genome sequence,
which comprises a circular chromosome of 1,278,540 bp including a tra
operon and an unstable 12,377-bp plasmid. To study the genetic
characteristics associated with virulence, we compared this species to R.
prowazekii, R. rickettsii and R. conorii. R. africae and R. prowazekii
have, respectively, the less and most decayed genomes. Eighteen genes are
present only in R. africae including one with a putative protease domain
upregulated at 37degreesC. CONCLUSION: Based on these data, we speculate
that a loss of regulatory genes causes an increase of virulence of
rickettsial species in ticks and mammals. We also speculate that in
Rickettsia species virulence is mostly associated with gene loss.

<>

<1>Fournier, P.E., El Karkouri, K., Robert, C., Medigue, C., Raoult, D.
<2>Complete Genome Sequence of Rickettsia slovaca, the Agent of Tick-Borne Lymphadenitis.
<3>J. Bacteriol.
<4>194
<5>1612
<6>2012
<7>The present study reports the complete and annotated genome sequence of the human pathogen
Rickettsia slovaca strain 13-B, which was isolated from a Dermacentor
tick in Slovakia in 1968. The 1.27-Mb genome provides further insights into the
acquisition of virulence related to genome reduction in Rickettsia species.

<>

<1>Fournier, P.E., Rouli, L., El Karkouri, K., Nguyen, T.T., Yagupsky, P., Raoult, D.
<2>Genomic Comparison of Kingella kingae Strains.
<3>J. Bacteriol.
<4>194
<5>5972
<6>2012
<7>Kingella kingae is a betaproteobacterium from the order Neisseriales, and it is an agent of
invasive infections in children. We sequenced the genome from the
septic arthritis strain 11220434. It is composed of a 1,990,794-bp chromosome but
no plasmid, and it contains 2,042 protein-coding genes and 52 RNA genes,
including 3 rRNA genes.

<>

<1>Fournier, P.E., Vallenet, D., Barbe, V., Audic, S., Ogata, H., Poirel, L., Richet, H., Robert, C., Mangenot, S., Abergel, C., Nordmann, P., Weissenbach, J., Raoult, D., Claverie, J.M.
<2>Comparative genomics of multidrug resistance in Acinetobacter baumannii.
<3>PLoS Genet.
<4>2
<5>e7
<6>2006
<7>Acinetobacter baumannii is a species of nonfermentative gram-negative bacteria commonly found
in water and soil. This organism was susceptible
to most antibiotics in the 1970s. It has now become a major cause of
hospital-acquired infections worldwide due to its remarkable propensity to
rapidly acquire resistance determinants to a wide range of antibacterial
agents. Here we use a comparative genomic approach to identify the
complete repertoire of resistance genes exhibited by the
multidrug-resistant A. baumannii strain AYE, which is epidemic in France,
as well as to investigate the mechanisms of their acquisition by
comparison with the fully susceptible A. baumannii strain SDF, which is
associated with human body lice. The assembly of the whole shotgun genome
sequences of the strains AYE and SDF gave an estimated size of 3.9 and 3.2
Mb, respectively. A. baumannii strain AYE exhibits an 86-kb genomic region
termed a resistance island--the largest identified to date--in which 45
resistance genes are clustered. At the homologous location, the SDF strain
exhibits a 20 kb-genomic island flanked by transposases but devoid of
resistance markers. Such a switching genomic structure might be a hotspot
that could explain the rapid acquisition of resistance markers under
antimicrobial pressure. Sequence similarity and phylogenetic analyses
confirm that most of the resistance genes found in the A. baumannii strain
AYE have been recently acquired from bacteria of the genera Pseudomonas,
Salmonella, or Escherichia. This study also resulted in the discovery of
19 new putative resistance genes. Whole-genome sequencing appears to be a
fast and efficient approach to the exhaustive identification of resistance
genes in epidemic infectious agents of clinical significance.

<>

<1>Foury, F., Roganti, T., Lecrenier, N., Purnelle, B.
<2>The complete sequence of the mitochondrial genome of Saccharomyces cerevisiae.
<3>FEBS Lett.
<4>440
<5>325-331
<6>1998
<7>The currently available yeast mitochondrial DNA (mtDNA) sequence is incomplete, contains many
errors and is derived from several polymorphic
strains. Here, we report that the mtDNA sequence of the strain used for
nuclear genome sequencing assembles into a circular map of 85,779 bp which
includes 10 kb of new sequence. We give a list of seven small hypothetical
open reading frames (ORFs). Hot spots of point mutations are found in
exons near the insertion sites of optional mobile group I intron-related
sequences. Our data suggest that shuffling of mobile elements plays an
important role in the remodelling of the yeast mitochondrial genome.

<>

<1>Fouts, D.E. et al.
<2>Major structural differences and novel potential virulence mechanisms from the genomes in multiple Campylobacter species.
<3>PLoS Biology
<4>3
<5>e15
<6>2005
<7>Sequencing and comparative genome analysis of four strains of Campylobacter including C. lari
RM2100, C. upsaliensis RM3195, and C. coli RM2228 has revealed major structural differences
that are associated with the insertion of phage- and plasmid-like genomic islands, as well as
major variations in the lipooligosaccharide complex. Poly G tracts are longer, are greater in
number, and show greater variability in C. upsaliensis than in the other species. Many genes
involved in host colonization, including racR/S, cadF, cdt, ciaB, and flagellin genes, are
conserved across the species, but variations that appear to be species specific are evident
for a lipooligosaccharide locus, a capsular (extracellular) polysaccharide locus, and a novel
Campylobacter putative licABCD virulence locus. The strains also vary in their metabolic
profiles, as well as their resistance profiles to a range of antibiotics. It is evident that
the newly identified hypothetical and conserved hypothetical proteins, as well as
uncharacterized two-component regulatory systems and membrane proteins, may hold additional
significant information on the major differences in virulence among the species, as well as
the specificity of the strains for particular hosts.

<>

<1>Fouts, D.E., Tyler, H.L., DeBoy, R.T., Daugherty, S., Ren, Q., Badger, J.H., Durkin, A.S., Huot, H., Shrivastava, S., Kothari, S., Dodson, R.J., Mohamoud, Y., Khouri, H., Roesch, L.F., Krogfelt, K.A., Struve, C., Triplett, E.W., Methe, B.A.
<2>Complete genome sequence of the N2-fixing broad host range endophyte Klebsiella pneumoniae 342 and virulence predictions verified in mice.
<3>PLoS Genet.
<4>4
<5>E1000141
<6>2008
<7>We report here the sequencing and analysis of the genome of the
nitrogen-fixing endophyte, Klebsiella pneumoniae 342. Although K.
pneumoniae 342 is a member of the enteric bacteria, it serves as a model
for studies of endophytic, plant-bacterial associations due to its
efficient colonization of plant tissues (including maize and wheat, two of
the most important crops in the world), while maintaining a mutualistic
relationship that encompasses supplying organic nitrogen to the host
plant. Genomic analysis examined K. pneumoniae 342 for the presence of
previously identified genes from other bacteria involved in colonization
of, or growth in, plants. From this set, approximately one-third were
identified in K. pneumoniae 342, suggesting additional factors most likely
contribute to its endophytic lifestyle. Comparative genome analyses were
used to provide new insights into this question. Results included the
identification of metabolic pathways and other features devoted to
processing plant-derived cellulosic and aromatic compounds, and a robust
complement of transport genes (15.4%), one of the highest percentages in
bacterial genomes sequenced. Although virulence and antibiotic resistance
genes were predicted, experiments conducted using mouse models showed
pathogenicity to be attenuated in this strain. Comparative genomic
analyses with the presumed human pathogen K. pneumoniae MGH78578 revealed
that MGH78578 apparently cannot fix nitrogen, and the distribution of
genes essential to surface attachment, secretion, transport, and
regulation and signaling varied between each genome, which may indicate
critical divergences between the strains that influence their preferred
host ranges and lifestyles (endophytic plant associations for K.
pneumoniae 342 and presumably human pathogenesis for MGH78578). Little
genome information is available concerning endophytic bacteria. The K.
pneumoniae 342 genome will drive new research into this less-understood,
but important category of bacterial-plant host relationships, which could
ultimately enhance growth and nutrition of important agricultural crops
and development of plant-derived products and biofuels.

<>

<1>Fox, K.L., Dowideit, S.J., Erwin, A.L., Srikhanta, Y.N., Smith, A.L., Jennings, M.P.
<2>Haemophilus influenzae phasevarions have evolved from type III DNA restriction systems into epigenetic regulators of gene expression.
<3>Nucleic Acids Res.
<4>35
<5>5242-5252
<6>2007
<7>Phase variably expressed (randomly switching) methyltransferases associated with type III
restriction-modification (R-M) systems have been
identified in a variety of pathogenic bacteria. We have previously shown
that a phase variable methyltransferase (Mod) associated with a type III
R-M system in Haemophilus influenzae strain Rd coordinates the random
switching of expression of multiple genes, and constitutes a phase
variable regulon-'phasevarion'. We have now identified the recognition
site for the Mod methyltransferase in H. influenzae strain Rd as
5'-CGAAT-3'. This is the same recognition site as the previously described
HinfIII system. A survey of 59 H. influenzae strains indicated significant
sequence heterogeneity in the central, variable region of the mod gene
associated with target site recognition. Intra- and inter-strain
transformation experiments using Mod methylated or non-methylated
plasmids, and a methylation site assay demonstrated that the sequence
heterogeneity seen in the region encoding target site specificity does
correlate to distinct target sites. Mutations were identified within the
res gene in several strains surveyed indicating that Res is not
functional. These data suggest that evolution of this type III R-M system
into an epigenetic mechanism for controlling gene expression has, in some
strains, resulted in loss of the DNA restriction function.

<>

<1>Fox, K.L., Srikhanta, Y.N., Jennings, M.P.
<2>Phase variable type III restriction-modification systems of host-adapted bacterial pathogens.
<3>Mol. Microbiol.
<4>65
<5>1375-1379
<6>2007
<7>Phase variation, the high-frequency on/off switching of gene expression, is a common feature
of host-adapted bacterial pathogens. Restriction-modification (R-M) systems, which are
ubiquitous among bacteria, are classically assigned the role of cellular defence against
invasion of foreign DNA. These enzymes are not obvious candidates for phase variable
expression, a characteristic usually associated with surface-expressed molecules subject to
host immune selection. Despite this, numerous type III R-M systems in bacterial pathogens
contain repetitive DNA motifs that suggest the potential for phase variation. Several roles
have been proposed for phase variable R-M systems based on DNA restriction function. However,
there is now evidence in several important human pathogens, including Haemophilus influenzae,
Neisseria meningitidis and Neisseria gonorrhoeae, that these systems are 'phasevarions'
(phasevariable regulons) controlling expression of multiple genes via a novel epigenetic
mechanism.

<>

<1>Fox, K.R.
<2>DNAse I footprinting of restriction enzymes.
<3>Biochem. Biophys. Res. Commun.
<4>155
<5>779-785
<6>1988
<7>DNAse I footprint of restriction enzymes has been achieved by using calcium
containing digestion buffers so that the enzymes bind to but do not cleave DNA.
EcoRI produces a footprint 17 bases long, overestimating the region of contact
with DNA by about 7-8 base pairs.  Restriction enzymes HaeIII and HinP1I
generate smaller footprints of 15 and 13 base pairs respectively.

<>

<1>Fox, K.R., Allinson, S.L., Sahagun-Krause, H., Brown, T.
<2>Recognition of GT mismatches by Vsr mismatch endonuclease.
<3>Nucleic Acids Res.
<4>28
<5>2535-2540
<6>2000
<7>The Vsr mismatch endonuclease recognises the sequence CTWGG (W=A or T) in which the indicated
thymine is paired with guanine and nicks the DNA backbone on the 5'-side of the mispaired
thymine.  By using base analogues of G and T we have explored the functional groups on the
mismatch pair which are recognized by the enzyme.  Removal of the thymine 5-methyl group
causes a 60% reduction in activity, while removing the 2-amino group of guanine reduces
cleavage by 90%.  Placing 2-aminopurine or nebularine opposite T generates mismatches which
are cut at a much lower rate (0.1%).  When either base is removed, generating a pseudoabasic
site (1',2'-dideoxyribose), the enzyme still produces site-specific cleavage, but at only 1%
of the original rate.  Although TT and CT mismatches at this position are cleaved at a low
rate (~1%), mismatches with other bases (such as GA and AC) and Watson-Crick base pairs are
not cleaved by the enzyme.  There is also no cleavage when the mismatched T is replaced with
difluorotoluene.

<>

<1>Fraga, M.F., Ballestar, E., Montoya, G., Taysavang, P., Wade, P.A., Esteller, M.
<2>The affinity of different MBD proteins for a specific methylated locus depends on their intrinsic binding properties.
<3>Nucleic Acids Res.
<4>31
<5>1765-1774
<6>2003
<7>The methyl-CpG binding domain (MBD) family of proteins was defined based on sequence
similarity in their DNA binding domains. In light of their
high degree of conservation, it is of inherent interest to determine the
genomic distribution of these proteins, and their associated co-repressor
complexes. One potential determinant of specificity resides in differences
in the intrinsic DNA binding properties of the various MBD proteins. In
this report, we use a capillary electrophoretic mobility shift assay
(CEMSA) with laser-induced fluorescence (LIF) and neutral capillaries to
calculate MBD-DNA binding affinities. MBD proteins were assayed on pairs
of methylated and unmethylated duplex oligos corresponding to the promoter
regions of the BRCA1, MLH1, GSTP1 and p16(INK4a) genes, and binding
affinities for each case were calculated by Scatchard analyses. With the
exception of mammalian MBD3 and Xenopus MBD3 LF, all the MBD proteins
showed higher affinity for methylated DNA (in the nanomolar range) than
for unmethylated DNA (in the micromolar range). Significant differences
between MBD proteins in the affinity for methylated DNA were observed,
ranging within two orders of magnitude. By mutational analysis of MBD3 and
using CEMSA, we demonstrate the critical role of specific residues within
the MBD in conferring selectivity for methylated DNA. Interestingly, the
binding affinity of specific MBD proteins for methylated DNA fragments
from naturally occurring sequences are affected by local methyl-CpG
spacing.

<>

<1>Franchina, M., Hooper, J., Kay, P.H.
<2>Five novel alternatively spliced transcripts of DNA (cytosine-5) methyltransferase 2 in human peripheral blood leukocytes.
<3>Int. J. Biochem. Cell Biol.
<4>33
<5>1104-1115
<6>2001
<7>Alternative splicing of RNA molecules transcribed from DNA (cytosine-5) methyltransferases has
been proposed as a mechanism by which methylation is able to effect diverse biological
processes in higher eukaryotes. This study has investigated transcriptional versatility of DNA
(cytosine-5) methyltransferase 2, which may methylate cytosine residues within 5'-CCTGG-3'
pentanucleotides in regions of the human genome devoid of 5'-CG-3' methylation. Five novel
splice variants of DNA (cytosine-5) methyltransferase 2 were identified in the peripheral
blood leukocytes of healthy subjects following cloning and sequencing of RT-PCR products
amplified using gene specific oligodeoxyribonucleotide primers. The generation of some of
these splice variants may be influenced by the formation of secondary structures within
pre-mRNA due to the repetition of sequences flanking alternatively spliced exons in a reverse
and complementary orientation on the same strand. These findings enable novel approaches to
investigate the role of RNA secondary structures in alternative splicing. The DNA (cytosine-5)
methyltransferase 2 splice variants are generated in all the major cell types of peripheral
blood, as well as in neoplastic lymphoid cells indicating that they are unlikely to generate
proteins involved in control of the cell cycle or cellular differentiation. Interestingly, the
gene products generated by some splice variants completely or partially lack highly conserved
amino acid motifs shown to be important for the catalysis of cytosine  methylation. The
possibility cannot be excluded, therefore, that alternative splicing of DNA (cytosine-5)
methyltransferase 2 pre-mRNA may generate protein isoforms which have different methylating
capabilities or which are involved in biological processes other than the catalysis of
cytosine methylation.

<>

<1>Franchina, M., Kay, P.H.
<2>Allele-specific variation in the gene copy number of human cytosine 5-methyltransferase.
<3>Hum. Hered.
<4>50
<5>112-117
<6>2000
<7>Previously, we have identified two alternate allelic forms of cytosine 5-methyltransferase,
5-MT I and 5-MT II, specified by polymorphic fragments of 1.5 and 1.1 kb, respectively. In the
presence study, a 0.8-kb genomic probe was prepared which was confirmed to be included within
the polymorphic fragments. The 0.8-kb probe hybridised with greater intensity to the 1.1-kb
fragment than the 1.5-kb fragment. Densitometric analysis indicated that there is 1 copy of
5-MT associated with 5-MT I, whereas there may be 1-4 copies of the gene associated with the
5-MT II allele. Segregation studies demonstrated that the multiple copies of 5-MT II are
inherited in a Mendelian fashion. These results allow novel approaches to investigating the
underlying mechanisms of cytosine methylation and gene duplication.

<>

<1>Franchina, M., Kay, P.H.
<2>Evidence that cytosine residues within 5'-CCTGG-3' pentanucleotides can be methylated in human DNA independently of the methylating system that modifies 5'-CG-3' dinucleotides.
<3>DNA Cell Biol.
<4>19
<5>521-526
<6>2000
<7>In contrast to the complex sequence specificities of the prokaryotic DNA methylating systems,
the mammalian machinery identified thus far methylates cytosine residues within the context of
a 5'-CG-3' dinucleotide. To explore the possibility that cytosine residues that do not
precede guanine may be independently methylated in mammalian DNA, we have examined a region of
the human myogenic gene, Myf-3, which is not targeted by the methylating system that
methylates 5'-CG-3' dinucleotides. Our investigations have revealed cytosine methylation
within the 5'-CCTGG-3' pentanucleotides specified by the 0.8-kb Myf-3 probe. We have also
found that in DNA from neoplastic cells, in which 5'-CG-3' dinucleotides within Myf-3 become
abnormally hypermethylated, cytosine residues within 5'-CCTGG-3' pentanucleotides are not
methylated. Moreover, methylation of 5'-CCTGG-3' pentanucleotides was not detected within
the closely related Myf-4 gene, which is normally 5'-CG-3' hypermethylated. These findings
indicate the existence of a system that methylates 5'-CCTGG-3' pentanucleotides
independently of the system that methylates cytosine residues within 5'-CG-3' dinucleotides.
It is possible that the 5'-CCTGG-3' methylating system influences the fate of foreign
integrated DNA.

<>

<1>Francois, J.-C., Saison-Behmoaras, T., Barbier, C., Chassignol, M., Thuong, N.T., Helene, C.
<2>Sequence-specific recognition and cleavage of duplex DNA via triple-helix formation by oligonucleotides covalently linked to a phenanthroline-copper chelate.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>86
<5>9702-9706
<6>1989
<7>Homopyrimidine oligodeoxynucleotides recognize the major groove of the DNA double helix at
homopurine-homopyrimidine sequences by forming local triple helices. Phenanthroline was
covalently attached to the 5' end of an 11-mer homopyrimidine oligonuceotide of sequence
d(TTTCCTCCTCT). Simian virus 40 DNA, which contains a single target site for this
oligonucleotide, was used as a substrate for the phenanthroline-oligonucleotide conjugate. In
the presence of copper ions and a reducing agent, a single specific double-strand cleavage
site was observed at 20oC by agarose gel electrophoresis. The efficiency of double-strand
cleavage was >70% at 20oC and pH 7.4. Secondary cleavage sites were observed when binding of
the oligonucleotide to mismatched sequences was allowed to take place at low temperature. The
exact location of the cleavage sites was determined by polyacrylamide gel electrophoresis of
denatured fragments by using both simian virus 40 DNA and a synthetic DNA fragment containing
the target sequence. The asymmetric distribution of the cleavage sites on the two strands
revealed that the cleavage reaction took place in the minor groove even though the
phenanthroline linker was located in the major groove. Linkers of different lengths were used
to tether phenanthroline to the oligonucleotide and their relative efficacies of DNA cleavage
were compared. Based on these comparative studies and on model building, it is proposed that
the phenanthroline ring carried by the oligonucleotide intercalates from the major groove and
that copper chelation locks the complex in place from within the minor groove where the
cleavage reaction occurs.

<>

<1>Francois, J.-C., Saison-Behmoaras, T., Thuong, N.T., Helene, C.
<2>Inhibition of restriction endonuclease cleavage via triple helix formation by homopyrimidine oligonucleotides.
<3>Biochemistry
<4>28
<5>9617-9619
<6>1989
<7>A 17-mer homopyrimidine oligonucleotide was designed to bind to the major
groove of SV40 DNA at a 17 base pair homopurine-homopyrimidine sequence via
Hoogsteen base pairing.  This sequence contains the recognition site for the
class II-S restriction enzyme Ksp632I.  The oligonucleotide was shown to
inhibit enzymatic cleavage under conditions that allow for triple helix
formation.  Inhibition is sequence-specific and occurs in the micromolar
concentration range.  Triple helix formation by oligonucleotides opens new
possibilities for sequence-specific regulation of gene expression.

<>

<1>Frank, O., Pradella, S., Rohde, M., Scheuner, C., Klenk, H.P., Goker, M., Petersen, J.
<2>Complete genome sequence of the Phaeobacter gallaeciensis type strain CIP 105210(T) (= DSM 26640(T) = BS107(T)).
<3>Standards in Genomic Sciences
<4>9
<5>914-932
<6>2014
<7>Phaeobacter gallaeciensis CIP 105210(T) (= DSM 26640(T) = BS107(T)) is the type strain of the
species Phaeobacter gallaeciensis. The genus Phaeobacter belongs to
the marine Roseobacter group (Rhodobacteraceae, Alphaproteobacteria). Phaeobacter
species are effective colonizers of marine surfaces, including frequent
associations with eukaryotes. Strain BS107(T) was isolated from a rearing of the
scallop Pecten maximus. Here we describe the features of this organism, together
with the complete genome sequence, comprising eight circular replicons with a
total of 4,448 genes. In addition to a high number of extrachromosomal replicons,
the genome contains six genomic island and three putative prophage regions, as
well as a hybrid between a plasmid and a circular phage. Phylogenomic analyses
confirm previous results, which indicated that the originally reported P.
gallaeciensis type-strain deposit DSM 17395 belongs to P. inhibens and that CIP
105210(T) (= DSM 26640(T)) is the sole genome-sequenced representative of P.
gallaeciensis.

<>

<1>Frank, S.A.
<2>Polymorphism of bacterial restriction-modification systems: the advantage of diversity.
<3>Evolution
<4>48
<5>1470-1477
<6>1994
<7>Bacterial restriction-modification systems provide defense against foreign DNA by using a self
versus nonself recognition mechanism. A great diversity of recognition motifs is maintained in
natural populations. Circumstantial evidence suggests that defense against bacteriophage
viruses favors this diversity. (1) Bacterial restriction enzymes can destroy invading phage
DNA. (2) Phage DNA can mimic the host's self-recognition mechanism. The ability of the virus
to pose as a mimic favors diversification of the host's recognition motif. Other observations
suggest that restriction modification (RM) does not provide any significant defensive
advantages in mature communities. (1) In laboratory experiments, bacteria evolve resistance to
phage by mutation and selection of the receptors to which phage adsorb. The outcome of these
experiments is a community dominated by bacteria with receptor-based resistance, with a low
abundance of phage and susceptible bacteria. (2) Phage are rare and receptor-based resistance
is common in samples from natural communities. I present a model that shows two factors
determine community composition: resources and RM diversity. Communities in resource-rich
habitats are dominated by receptor-based resistance and support few phage; communities in poor
habitats are dominated by restriction-modification defense and relatively abundant phage. RM
diversity is itself a direct cause of community composition. As diversity increases from a low
level, the abundance of phage increases and the relative abundance of receptor-based
resistance declines. Further increases in diversity cause a crash in phage abundance, yielding
a stable community of diverse RM types but an absence of the selective pressure--the
phage--that drove the diversification. Empirical studies must sample a range of resource
levels and RM diversity to analyze the forces that determine community composition.

<>

<1>Frankel, A.D.
<2>Sequence-specific recognition of DNA by the HinfI restriction endonuclease.
<3>Ph.D. Thesis, Johns Hopkins University, USA
<4>
<5>1-92
<6>1983
<7>A method has been developed for measuring association constants of DNA-protein
complexes using gel chromatography.  It is suitable for the study of a variety
of systems and can be used over a wide range of concentrations.  This technique
has been used to study the sequence-specific interaction of the HinfI
restriction endonuclease with DNA.  HinfI has a monomer molecular weight of
31000 daltons and appears to be active as a dimer.  Its turnover number is 25
sites cleaved min-1 dimer-1 and its Km is less than 1x10-11M.  The protein was
found to bind to supercoiled plasmid molecules with an observed free energy of
association of -13.9 kcal mole-1.  The binding decreases at concentrations of
NaCl above 50mM and this dependence corresponds to the apparent release of 3.4
ion pairs.  The affinity of the nuclease for its site was found to be
independent of pH over the range studied and showed a small dependence on
temperature.  When the degenerate middle position of the HinfI recognition
site, 5'GANTC, was varied, no change was observed in the binding constant.  The
salt and pH dependencies of the cleavage reaction are similar to those of the
binding constants and each of the degenerate sites studied was equally well
cleaved.  Linear fragments containing the site are bound as tightly as
supercoiled molecules.  The enzyme binds to non-specific DNA about 6 orders of
magnitude more weakly than to its site and is unable to bind to DNA methylated
at the A position of its recognition site.

<>

<1>Frankel, A.D., Ackers, G.K., Smith, H.O.
<2>Measurement of DNA-protein equilibria using gel chromatography:  application to the HinfI restriction endonuclease.
<3>Biochemistry
<4>24
<5>3049-3054
<6>1985
<7>A method is described for measuring equilibrium constants of DNA-protein
interactions using gel chromatography.  This technique has been used to study
the sequence-specific interaction of the HinfI restriction endonuclease with
DNA.  HinfI has a monomeric molecular weight of 31000 and exists as a dimer in
its active form.  The protein binds to supercoiled DNA molecules containing its
recognition site with an apparent free energy of -13.9 kcal/mol of sites.  This
interaction is highly salt sensitive and causes a release of 3.4 ion pairs.
The affinity of the nuclease for its recognition site is largely independent of
both pH (6.5-8.5) and temperature (7-35C) and was not affected by variations in
the degenerate middle position of the site.  Linear DNA fragments containing
the HinfI recognition site were bound as tightly as supercoiled molecules.
Binding to nonspecific DNA sites or to methylated DNA sites was approximately 6
orders of magnitude weaker.  In general, enzyme activity and binding affinity
parallel each other.

<>

<1>Frankel, A.D., Smith, H.O.
<2>Restriction and modification enzymes detect no allosteric changes in DNA with bound lac repressor or RNA polymerase.
<3>J. Mol. Biol.
<4>146
<5>611-619
<6>1981
<7>A 203 base-pair fragment containing the lac operator/promoter region of
Escherichia coli was inserted into the EcoRI site of the plasmid vector pKC7.
Rates of restriction endonuclease cleavage of the flanking EcoRI sites and of
several other restriction sites on the DNA molecule were then compared in the
presence and absence of bound RNA polymerase or lac repressor.  The rates were
identical whether or not protein had been bound, even for sites as close as 40
base-pairs from a protein binding site.  No difference was detected using
supercoiled, nicked circular, or linear DNA substrates.  No apparent change in
the rates of methylation of EcoRI sites by EcoRI methylase was produced by
binding the regulatory proteins.

<>

<1>Franklin, N.C., Dove, W.F.
<2>Genetic evidence for restriction targets in the DNA of phages lambda and Phi80.
<3>Genet. Res.
<4>14
<5>151-157
<6>1969
<7>The phenomenon of restriction, recently reviewed in depth by Arber (1968) and
by Arber and Linn (1969), is observed as the inactivation of a genome following
transfer from one host to another.  Thus, phages propagated on one host may
form plaques on a second host with an efficiency lower than that on the first.
Those progeny phages which do emerge from the second host generally will have
become modified so that they are no longer restricted in that host.  This
modification is not replicated, and is diluted out upon further propagation of
the phages in the first host.  The ability of a bacterium to restrict or to
modify depends upon three linked bacterial cistrons:  one required for
restriction, the second for modification and the third required for both
functions and determining the specificity of each (Glover, Schell, Symonds &
Stacey, 1963; Wood, 1966, Boyer & Roulland-Dussoix, 1969; Glover & Colson,
1969).  Restriction of a phage by a bacterium was shown some years ago to be
exerted directly upon the DNA of the phage after it is injected into the
bacterial cell (Dussoix & Arber, 1962).  Restricting enzymes have now been
purified and found to act at only a limited number of sites in the target DNA
molecule, making double-strand breaks (Meselson & Yuan, 1968; Linn & Arber,
1968; Roulland-Dussoix & Boyer, 1969).  Subsequent degradation in vivo
presumably occurs by non-specific nuclease action on these fragments.
Comparison of the restriction properties of several phages has now led to
observations of a genetic nature which confirm the conclusion, based on
biochemical evidence, that restriction is directed at target sites localized
within the phage genome.  The restriction system of E. coli K12 is far more
active on phage lambda than on the related phage Phi80.  The restriction
properties of these phages are unaffected by each other in a trans
complementation test.  Rather, the restriction properties can be exchanged only
by genetic recombination, behaving as a small set of mappable restriction
targets.  Sensitivity of lambda to K restriction can be lost by genetic
deletion.

<>

<1>Franzon, V.L., Barker, A., Manning, P.A.
<2>Nucleotide sequence encoding the mannose-fucose-resistant hemagglutinin of Vibrio cholerae O1 and construction of a mutant.
<3>Infect. Immun.
<4>61
<5>3032-3037
<6>1993
<7>The region of DNA encoding the mannose-fucose-resistant hemagglutinin (MFRHA) of Vibrio
cholerae O1 has been localized, and the nucleotide sequence has been determined. The region
contains a single open reading frame encoding 230 amino acids, corresponding to a protein of
26.9 kDa. The N terminus of this protein is atypical for a protein localized in the outer
membrane. A mutant lacking MFRHA activity has been constructed by allelic exchange after
inactivation via the insertion of a kanamycin resistance gene cartridge. The MFRHA-negative
mutant has been assessed for virulence in the infant mouse cholera model. This mutant shows a
marked defect in its ability to persist in the infant mouse gut and is incapable of competing
with the wild-type organism, even when given in 25-fold excess. This defect also leads to a >
100-fold increase in the 50% lethal dose. These data suggest that the MFRHA is an important
colonization factor in the infant mouse model.

<>

<1>Fraser, C., Galeotti, C., Grandi, G., Hickey, E., Masignani, V., Mora, M., Petersen, J., Pizza, M., Rappuoli, R., Ratti, G., Scalato, E., Scarselli, M., Tettelin, H., Venter, C.J.
<2>Neisseria meningitidis antigens and compositions.
<3>European Patent Office
<4>EP 1944371 A
<5>
<6>2008
<7>The invention provides proteins from Neisseria meningitidis, including the amino acid
sequences and the corresponding nucleotide sequences.  The proteins are predicted to be useful
antigens for vaccines and/or diagnostics.

<>

<1>Fraser, C., Masignani, V., Tetterin, H.
<2>STREPTOCOCCUS PNEUMONIAE PROTEINS AND NUCLEIC ACIDS.
<3>Japanese Patent Office
<4>JP 2005503119 A
<5>
<6>2005
<7>
<>

<1>Fraser, C.M. et al.
<2>The minimal gene complement of Mycoplasma genitalium.
<3>Science
<4>270
<5>397-403
<6>1995
<7>The complete nucleotide sequence (580,070 base pairs) of the Mycoplasma genitalium genome, the
smallest known genome of any free-living organism, has been determined by whole-genome random
sequencing and assembly.  A total of only 470 predicted coding regions were identified that
include genes required for DNA replication, transcription and translation, DNA repair,
cellular transport, and energy metabolism.  Comparison of this genome to that of Haemophilus
influenzae suggests that differences in genome content are reflected as profound differences
in physiology and metabolic capacity between these two organisms.

<>

<1>Fraser, C.M. et al.
<2>Genomic sequence of a Lyme disease spirochaete, Borrelia burgdorferi.
<3>Nature
<4>390
<5>580-586
<6>1997
<7>The genome of the bacterium Borrelia burgdorferi B31, the aetiologic agent of Lyme disease,
contains a linear chromosome of 910,725 base pairs and at least 17 linear and circular
plasmids with a combined size of more than 53,000 base pairs.  The chromosome contains 853
genes encoding a basic set of proteins for DNA replication, transcription, translation, solute
transport and energy metabolism, but, like Mycoplasma genitalium, it contains no genes for
cellular biosynthetic reactions.  Because B. burgdorferi and M. genitalium are distantly
related eubacteria, we suggest that their limited metabolic capacities reflect convergent
evolution by gene loss from more metabolically competent progenitors.  Of 430 genes on 11
plasmids, most have no known biological function; 39% of plasmid genes are paralogues that
form 47 gene families.  The biological significance of the multiple plasmid-encoded genes is
not clear, although they may be involved in antigenic variation or immune evasion.

<>

<1>Fraser, C.M. et al.
<2>Complete genome sequence of Treponema pallidum, the syphilis spirochete.
<3>Science
<4>281
<5>375-388
<6>1998
<7>The complete genome sequence of Treponema pallidum was determined and shown to be 1,138,006
pairs containing 1041 predicted coding sequences (open reading frames).  Systems for DNA
replication, transcription, translation, and repair are intact, but catabolic and biosynthetic
activities are minimized.  The number of identifiable transporters is small, and no
phosphoenolpyruvate: phosphotransferase carbohydrate transporters were found.  Potential
virulence factors include a family of 12 potential membrane proteins and several putative
hemolysins.  Comparison of the T. pallidum genome sequence with that of another pathogenic
spirochete, Borrelia burgdorferi, the agent of Lyme disease, identified unique and common
genes and substantiates the considerable diversity observed among pathogenic spirochetes.

<>

<1>Fraser, C.M., Adams, M.D., Gocayne, J.D., Hutchison, C.A. III, Smith, H.O., Venter, J.C., White, O.
<2>Nucleotide sequence of the Mycoplasma genitalium genome, fragments thereof, and uses thereof.
<3>US Patent Office
<4>US 6537773 A
<5>
<6>2003
<7>The present invention provides the nucleotide sequence of the entire genome of Mycoplasma
genitalium, SEQ ID NO: 1.  The present invention further provides the sequence information
stored on computer readable media, and computer-based systems and methods which facilitate its
use.  In addition to the entire genomic sequence, the present invention identifies protein
encoding fragments of the genome, and identifies, by position relative to two (2) genes known
to flank the origin of replication, any regulatory elements which modulate the expression of
the protein encoding fragments of the Mycoplasma genitalium genome.

<>

<1>Frauer, C., Leonhardt, H.
<2>A versatile non-radioactive assay for DNA methyltransferase activity and DNA binding.
<3>Nucleic Acids Res.
<4>37
<5>e22
<6>2009
<7>We present a simple, non-radioactive assay for DNA methyltransferase activity and DNA binding.
As most proteins are studied as GFP fusions in
living cells, we used a GFP binding nanobody coupled to agarose beads (GFP
nanotrap) for rapid one-step purification. Immobilized GFP fusion proteins
were subsequently incubated with different fluorescently labeled DNA
substrates. The absolute amounts and molar ratios of GFP fusion proteins
and bound DNA substrates were determined by fluorescence spectroscopy. In
addition to specific DNA binding of GFP fusion proteins, the enzymatic
activity of DNA methyltransferases can also be determined by using suicide
DNA substrates. These substrates contain the mechanism-based inhibitor
5-aza-dC and lead to irreversible covalent complex formation. We obtained
covalent complexes with mammalian DNA methyltransferase 1 (Dnmt1), which
were resistant to competition with non-labeled canonical DNA substrates,
allowing differentiation between methyltransferase activity and DNA
binding. By comparison, the Dnmt1(C1229W) catalytic site mutant showed
DNA-binding activity, but no irreversible covalent complex formation. With
this assay, we could also confirm the preference of Dnmt1 for
hemimethylated CpG sequences. The rapid optical read-out in a multi-well
format and the possibility to test several different substrates in direct
competition allow rapid characterization of sequence-specific binding and
enzymatic activity.

<>

<1>Frauer, C., Leonhardt, H.
<2>Twists and turns of DNA methylation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>8919-8920
<6>2011
<7>DNA methylation, the post-replicative transfer of a methyl group to the C5 position of
cytosine bases, was the first epigenetic modification identified and has been intensively
studied for more than half a century.  By now it is clear that Dnmt1, the major eukaryotic DNA
methyltransferase, faithfully maintains genome-wide methylation patterns and plays an
essential role in the epigenetic network controlling gene expression and genome stability
during development.  However, the molecular mechanisms that ultimately control DNA methylation
still remain elusive.  This is, in part, attributable to the remarkable complexity of the DNA
methylation reaction, the apparent involvement of several inter- and intra-molecular protein
interactions, and the limited structural information.  The crystal structure of Dnmt1
presented in PNAS now provides detailed insights into the inner workings and possible
regulation of one of the most intriguing enzymes.

<>

<1>Fraunholz, M., Bernhardt, J., Schuldes, J., Daniel, R., Hecker, M., Sinha, B.
<2>Complete Genome Sequence of Staphylococcus aureus 6850, a Highly Cytotoxic and Clinically Virulent Methicillin-Sensitive Strain with Distant Relatedness to  Prototype Strains.
<3>Genome Announcements
<4>1
<5>e00775-13
<6>2013
<7>Staphylococcus aureus is a frequent human commensal bacterium and pathogen. Here  we report
the complete genome sequence of strain 6850 (spa type t185; sequence
type 50 [ST50]), a highly cytotoxic and clinically virulent methicillin-sensitive
strain from a patient with complicated S. aureus bacteremia associated with
osteomyelitis and septic arthritis.

<>

<1>Frederick, C.A., Grable, J., Melia, M., Samudzi, C., Jen-Jacobson, L., Wang, B.-C., Greene, P., Boyer, H.W., Rosenberg, J.M.
<2>Kinked DNA in crystalline complex with EcoRI endonuclease.
<3>Nature
<4>309
<5>327-331
<6>1984
<7>The 3 angstrom electron density map of a co-crystalline recognition complex
between EcoRI endonuclease and the oligonucleotide TCGCGAATTCGCG reveals that a
tight, complementary interface between the enzyme and the major groove of the
DNa is the major determinant of sequence specificity.  The DNA contains a
torsional kink and other departures from the B conformation which unwind the
DNA and thereby widen the major groove in the recognition site.

<>

<1>Frederick, C.A., Quigley, G.J., van der Marel, G.A., van Boom, J.H., Wang, A.H.-J., Rich, A.
<2>Methylation of the EcoRI recognition site does not alter DNA conformation:  The crystal structure of d(CGCGAm6ATTCGCG) at 2.0-A resolution.
<3>J. Biol. Chem.
<4>263
<5>17872-17879
<6>1988
<7>Methylation of nucleic acid bases is known to prevent the cleavage of DNA by
restriction endonucleases.  The effect on the conformation of the DNA molecule
itself and hence its interactions with other DNA binding proteins has been a
subject of general interest.  To help address this question, we have solved the
crystal structure at 2.0 A of the methylated dodecamer, d(CGCGAm6ATTCGCG),
which contains the EcoRI recognition sequence and have compared the
conformation of the methylated molecule with that of its nonmethylated
counterpart.  This methylation produces a bulky hydrophobic patch on the floor
of the major groove of B-DNA which plays an important role in the mechanism of
inhibition of EcoRI restriction activity.  However, with the exception of small
perturbations in the immediate vicinity of the methyl groups, the structure is
virtually unchanged.  Given the lack of a conformational change upon
methylation, we have extended this thesis of the recognition process to other
types of restriction systems and found that different restriction enzymes seem
to have their own characteristic protein-DNA interactions.  The relative
spatial orientations of methylation sites and cleavage sites must play a major
role in ordering protein secondary structure elements as well as
subunit-subunit interactions along the DNA strand.

<>

<1>Free, A., Wakefield, R.I., Smith, B.O., Dryden, D.T., Barlow, P.N., Bird, A.P.
<2>DNA recognition by the methyl-CpG binding domain of MeCP2.
<3>J. Biol. Chem.
<4>276
<5>3353-3360
<6>2001
<7>The methyl-CpG binding domain (MBD) of the transcriptional repressor MeCP2 has been proposed
to recognize a single symmetrically methylated CpG base pair via hydrophobic patches on an
otherwise positively charged DNA binding surface. We have tested this binding model by
analysis of mutant derivatives of the MeCP2 MBD in electrophoretic mobility shift assays
complemented by NMR structural analysis. Exposed arginine side chains on the binding face, in
particular Arg-111, were found to be critical for binding. Arg-111 was found to interact with
the conserved aspartate side chain Asp-121, which is proposed to orientate the arginine side
chain to allow specific contacts with the DNA. The conformational flexibility of the
disordered B-C loop region, which forms part of the binding face, was also shown to be
important. In contrast, mutation of the exposed hydrophobic side chains had a less severe
effect on DNA binding. This suggests that the Arg-111 side chain may contribute to
sequence-specific recognition of the CpG site rather than simply making nonspecific contacts
with the phosphate backbone. The majority of missense mutations within the MBD found in the
human genetic disorder Rett syndrome were shown or predicted to affect folding of the domain
rather than the DNA recognition event directly.

<>

<1>Freese, H.M. et al.
<2>Genome sequence of the phage-gene rich marine Phaeobacter arcticus type strain DSM 23566(T.).
<3>Standards in Genomic Sciences
<4>8
<5>450-464
<6>2013
<7>Phaeobacter arcticus Zhang et al. 2008 belongs to the marine Roseobacter clade whose members
are phylogenetically and physiologically diverse. In contrast to
the type species of this genus, Phaeobacter gallaeciensis, which is well
characterized, relatively little is known about the characteristics of P.
arcticus. Here, we describe the features of this organism including the annotated
high-quality draft genome sequence and highlight some particular traits. The
5,049,232 bp long genome with its 4,828 protein-coding and 81 RNA genes consists
of one chromosome and five extrachromosomal elements. Prophage sequences
identified via PHAST constitute nearly 5% of the bacterial chromosome and
included a potential Mu-like phage as well as a gene-transfer agent (GTA). In
addition, the genome of strain DSM 23566(T) encodes all of the genes necessary
for assimilatory nitrate reduction. Phylogenetic analysis and intergenomic
distances indicate that the classification of the species might need to be
reconsidered.

<>

<1>Freitag, M., Selker, E.U.
<2>Controlling DNA methylation: many roads to one modification.
<3>Curr. Opin. Genet. Dev.
<4>15
<5>191-199
<6>2005
<7>Genetic, biochemical and cytological studies on DNA methylation in several eukaryotic
organisms have resulted in leaps of understanding in the past three years. Discoveries of
mechanistic links between DNA methylation and histone methylation, and between these processes
and RNA interference (RNAi) machineries have reinvigorated the field. The details of the
connections between DNA methylation, histone modifications and RNA silencing remain to be
elucidated, but it is already clear that no single pathway accounts for all DNA methylation
found in eukaryotes. Rather, different taxa use one or more of several general mechanisms to
control methylation. Despite recent progress, classic questions remain, including: What are
the signals for DNA methylation? Are 'de novo' and 'maintenance' methylation truly
separate processes? How is DNA methylation regulated?

<>

<1>Freitag, M., Williams, R.L., Kothe, G.O., Selker, E.U.
<2>A cytosine methyltransferase homologue is essential for repeat-induced point mutation in Neurospora crassa.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>8802-8807
<6>2002
<7>During sexual development, Neurospora crassa inactivates genes in duplicated DNA segments by a
hypermutation process, repeat-induced point mutation (RIP). RIP introduces C:G to T:A
transition mutations and creates targets for subsequent DNA methylation in vegetative tissue.
The mechanism of RIP and its relationship to DNA methylation are not fully understood.
Mutations in DIM-2, a DNA methyltransferase (DMT) responsible for all known cytosine
methylation in Neurospora, does not prevent RIP. We used RIP to disrupt a second putative DMT
gene in the Neurospora genome and tested mutants for defects in DNA methylation and RIP. No
effect on DNA methylation was detected in the tissues that could be assayed, but the mutants
showed recessive defects in RIP. Duplications of the am and mtr genes were completely stable
in crosses homozygous for the mutated potential DMT gene, which we call rid (RIP defective).
The same duplications were inactivated normally in heterozygous crosses. Disruption of the rid
gene did not noticeably affect fertility, growth, or development. In contrast, crosses
homozygous for a mutation in a related gene in Ascobolus immersus, masc1, reportedly fail to
develop and heterozygous crosses reduce methylation induced premeiotically .  We isolated
homologues of rid from Neurospora tetrasperma and Neurospora intermedia to identify conserved
regions. Homologues possess all motifs characteristic of eukaryotic DMTs and have large
distinctive C- and N-terminal domains.

<>

<1>French, C.T., Sitaraman, R., Dybvig, K.
<2>DNA inversions regulate a family of phase-variable restriction and modification enzymes in Mycoplasma pulmonis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>0
<5>274-275
<6>1998
<7>Mycoplasma pulmonis is a murine pathogen that produces chronic disease in the reproductive and
respiratory tracts and joints.  The ability to undergo rapid phenotypic changes is probably an
important pathogenic attribute allowing the organism to quickly adapt to a changing
environment.  The hsd1 and hsd2 loci of M. pulmonis are highly homologous, site-specific DNA
inversion systems that encode proteins exhibiting significant amino acid similarity with the
type I restriction and modification systems of enteric bacteria.  DNA inversions regulate
transcription of the hsdS genes, acting as an on/off switches controlling which particular
hsdS genes are expressed in the cell.  The hsdS genes encode the S subunits which dictate the
sequence specificity of the type I holoenzyme.  We hypothesized that the various hsdS genes
that can be induced by DNA inversion encode S subunits with different DNA recognition
specificities, resulting in the production of R-M enzymes with different specificities.  To
test this possibility, cultures of M. pulmonis was subcloned using filter cloning methods.
R-M properties were examined by quantitating PFU of mycoplasma virus P1 on lawns of each
individual subclone.  The R-M properties of over 100 subclones have been examined thus far.
Subclones have been divided into five groups, each with unique R-M properties.  Group I is R-M
negative.  Southern blot and hsd mRNA transcription anlayses suggest that R-M systems are
"off" in these subclones because the hsd1 and hsd2 loci are oriented in the chromosome such
that the hsdR and hsdM genes (encoding the R and M subunits of the type I holoenzyme) are not
transcribed.  Groups II, III, IV and V are R-M positive, with the specificity of the R-M
activity of each group being distinct.  Gene analyses (Southern hybridization, PCR and
nucleotide sequencing indicate that different hsdS genes are transcribed in the different
groups, explaining the unique R-M properties of each group.  These results indicate that M.
pulmonis possesses a novel family of phase-variable R-M enzymes.  Because hsd inversions have
previously been correlated with other gene rearrangements that regulate the phase-variable
production of the V-1 surface proteins, it is possible that R-M enzymes in this system have an
important but undefined role in disease pathogenesis.

<>

<1>Fresco-Taboada, A., Del Cerro, C., Fernandez-Lucas, J., Arroyo, M., Acebal, C., Garcia, J.L., de la Mata, I.
<2>Genome of the Psychrophilic Bacterium Bacillus psychrosaccharolyticus, a Potential Source of 2'-Deoxyribosyltransferase for Industrial Nucleoside  Synthesis.
<3>Genome Announcements
<4>1
<5>e00309-13
<6>2013
<7>Here we report the draft genome sequence of Bacillus psychrosaccharolyticus, a cold-adapted
bacterium with biotechnological interest. The genome contains genes
related to the ability of this microorganism to grow at low temperatures and
includes a nucleoside 2'-deoxyribosyltransferase, which can be used in the
industrial synthesis of modified nucleosides with therapeutic activity.

<>

<1>Frey, B., Kahle, C., Zolch, C., Kaluza, K., Herz, G., Lechner, M., Auer, J., Schmitz, G.
<2>Screening for novel class-II restriction endonucleases.
<3>Fresenius Z. Anal. Chem.
<4>343
<5>123-124
<6>1992
<7>More than 1500 class-II restriction endonucleases have been isolated from eu- and
archaebacteria. In addition only a few restriction enzymes were described from eukaryotic
sources like virus infected chlorella-like green algae. These enzymes represent more than 170
different sequence specificities. Class-II restriction endonucleases are important tools in
recombinant DNA technology. In addition restriction enzymes recognizing octa- and
heptanucleotide sequences are necessary for the mapping of genomes because they produce very
large fragments that can be resolved by pulse field gel electrophoresis. Our aim was to find
and characterize new restriction enzymes and determine the specificities with respect to their
recognition sequence and cleavage position.

<>

<1>Frey, B., Kaluza, K., Auer, J., Stratidakis, I., Rina, M., Bouriotis, V., Schmitz, G.
<2>AspEI, a novel Eam11051 isoschizomer from Aureobacterium species recognizing 5'-GACnnn/nnGTC-3' .
<3>Nucleic Acids Res.
<4>20
<5>3782
<6>1992
<7>We have isolated AspEI, a novel class II restriction endonuclease from Aureobacterium species
3676 recognizing the palindromic sequence 5'-GACnnn/nnGTC-3' generating a 3'-protruding
mononucleotide. With respect to its isoschizomer from Eam1105I, the novel enzyme can be easily
isolated in high purity because of its occurrence as a single restriction enzyme in the
Aureobacterium strain and the presence of high specific activity even in the crude extract. A
comparison of cleavage patterns experimentally obtained with AspEI on standard lambda,
phiX174RF, pBR322, T7 and Ad2 DNAs of known nucleotide sequence, with computer-derived mapping
data predicts the sequence 5'-GACn5GTC-3'. The cut positions within the AspEI recognitin
site were determined according to the enzymatic sequencing approach.

<>

<1>Frey, K.G., Bishop-Lilly, K.A., Daligault, H.E., Davenport, K.W., Bruce, D.C., Chain, P.S., Coyne, S.R., Chertkov, O., Freitas, T., Jaissle, J., Koroleva, G.I., Ladner, J.T., Minogue, T.D., Palacios, G.F., Redden, C.L., Xu, Y., Johnson, S.L.
<2>Full-Genome Assembly of Reference Strain Providencia stuartii ATCC 33672.
<3>Genome Announcements
<4>2
<5>e01082-14
<6>2014
<7>A member of the normal human gut microflora, Providencia stuartii is of clinical  interest due
to its role in nosocomial infections of the urinary tract and
because it readily acquires antibiotic resistance. Here, we present the complete
genome of P. stuartii strain ATCC 33672, consisting of a 4.28-Mbp chromosome and
a 48.9-kbp plasmid.

<>

<1>Fricke, W.F., Mammel, M.K., McDermott, P.F., Tartera, C., White, D.G., Leclerc, J.E., Ravel, J., Cebula, T.A.
<2>Comparative genomics of 28 Salmonella enterica isolates: evidence for CRISPR-mediated adaptive sublineage evolution.
<3>J. Bacteriol.
<4>193
<5>3556-3568
<6>2011
<7>Despite extensive surveillance, food-borne Salmonella enterica infections
continue to be a significant burden on public health systems worldwide. As
the S. enterica species comprises sublineages that differ greatly in
antigenic representation, virulence, and antimicrobial resistance
phenotypes, a better understanding of the species' evolution is critical
for the prediction and prevention of future outbreaks. The roles that
virulence and resistance phenotype acquisition, exchange, and loss play in
the evolution of S. enterica sublineages, which to a certain extent are
represented by serotypes, remains mostly uncharacterized. Here, we compare
17 newly sequenced and phenotypically characterized nontyphoidal S.
enterica strains to 11 previously sequenced S. enterica genomes to carry
out the most comprehensive comparative analysis of this species so far.
These phenotypic and genotypic data comparisons in the phylogenetic
species context suggest that the evolution of known S. enterica
sublineages is mediated mostly by two mechanisms, (i) the loss of coding
sequences with known metabolic functions, which leads to functional
reduction, and (ii) the acquisition of horizontally transferred phage and
plasmid DNA, which provides virulence and resistance functions and leads
to increasing specialization. Matches between S. enterica clustered
regularly interspaced short palindromic repeats (CRISPR), part of a
defense mechanism against invading plasmid and phage DNA, and plasmid and
prophage regions suggest that CRISPR-mediated immunity could control
short-term phenotype changes and mediate long-term sublineage evolution.
CRISPR analysis could therefore be critical in assessing the evolutionary
potential of S. enterica sublineages and aid in the prediction and
prevention of future S. enterica outbreaks.

<>

<1>Fricke, W.F., McDermott, P.F., Mammel, M.K., Zhao, S., Johnson, T.J., Rasko, D.A., Fedorka-Cray, P.J., Pedroso, A., Whichard, J.M., Leclerc, J.E., White, D.G., Cebula, T.A., Ravel, J.
<2>Antimicrobial resistance-conferring plasmids with similarity to virulence plasmids from avian pathogenic Escherichia coli strains in Salmonella enterica serovar Kentucky isolates from poultry.
<3>Appl. Environ. Microbiol.
<4>75
<5>5963-5971
<6>2009
<7>Salmonella enterica, a leading cause of food-borne gastroenteritis
worldwide, may be found in any raw food of animal, vegetable, or fruit
origin. Salmonella serovars differ in distribution, virulence, and host
specificity. Salmonella enterica serovar Kentucky, though often found in
the food supply, is less commonly isolated from ill humans. The
multidrug-resistant isolate S. Kentucky CVM29188, isolated from a chicken
breast sample in 2003, contains three plasmids (146,811 bp, 101,461 bp,
and 46,121 bp), two of which carry resistance determinants (pCVM29188_146
[strAB and tetRA] and pCVM29188_101 [bla(CMY-2) and sugE]). Both
resistance plasmids were transferable by conjugation, alone or in
combination, to S. Kentucky, Salmonella enterica serovar Newport, and
Escherichia coli recipients. pCVM29188_146 shares a highly conserved
plasmid backbone of 106 kb (>90% nucleotide identity) with two virulence
plasmids from avian pathogenic Escherichia coli strains (pAPEC-O1-ColBM
and pAPEC-O2-ColV). Shared avian pathogenic E. coli (APEC) virulence
factors include iutA iucABCD, sitABCD, etsABC, iss, and iroBCDEN. PCR
analyses of recent (1997 to 2005) S. Kentucky isolates from food animal,
retail meat, and human sources revealed that 172 (60%) contained similar
APEC-like plasmid backbones. Notably, though rare in human- and
cattle-derived isolates, this plasmid backbone was found at a high
frequency (50 to 100%) among S. Kentucky isolates from chickens within the
same time span. Ninety-four percent of the APEC-positive isolates showed
resistance to tetracycline and streptomycin. Together, our findings of a
resistance-conferring APEC virulence plasmid in a poultry-derived S.
Kentucky isolate and of similar resistance/virulence plasmids in most
recent S. Kentucky isolates from chickens and, to lesser degree, from
humans and cattle highlight the need for additional research in order to
examine the prevalence and spread of combined virulence and resistance
plasmids in bacteria in agricultural, environmental, and clinical
settings.

<>

<1>Fricke, W.F., Seedorf, H., Henne, A., Kruer, M., Liesegang, H., Hedderich, R., Gottschalk, G., Thauer, R.K.
<2>The genome sequence of Methanosphaera stadtmanae reveals why this human intestinal archaeon is restricted to methanol and H2 for methane formation and ATP synthesis.
<3>J. Bacteriol.
<4>188
<5>642-658
<6>2006
<7>Methanosphaera stadtmanae has the most restricted energy metabolism of all methanogenic
archaea. This human intestinal inhabitant can generate methane only by reduction of methanol
with H2 and is dependent on acetate as a carbon source. We report here the genome sequence of
M. stadtmanae, which was found to be composed of 1,767,403 bp with an average G+C content of
28% and to harbor only 1,534 protein-encoding sequences (CDS). The genome lacks 37 CDS present
in the genomes of all other methanogens. Among these are the CDS for synthesis of
molybdopterin and for synthesis of the carbon monoxide dehydrogenase/acetyl-coenzyme A
synthase complex, which explains why M. stadtmanae cannot reduce CO2 to methane or oxidize
methanol to CO2 and why this archaeon is dependent on acetate for biosynthesis of cell
components. Four sets of mtaABC genes coding for methanol:coenzyme M methyltransferases were
found in the genome of M. stadtmanae. These genes exhibit homology to mta genes previously
identified in Methanosarcina species. The M. stadtmanae genome also contains at least 323 CDS
not present in the genomes of all other archaea. Seventy-three of these CDS exhibit high
levels of homology to CDS in genomes of bacteria and eukaryotes. These 73 CDS include 12 CDS
which are unusually long (>2,400 bp) with conspicuous repetitive sequence elements, 13 CDS
which exhibit sequence similarity on the protein level to CDS encoding enzymes involved in the
biosynthesis of cell surface antigens in bacteria, and 5 CDS which exhibit sequence similarity
to the subunits of bacterial type I and III restriction-modification systems.

<>

<1>Fridman, O., Goldberg, A., Ronin, I., Shoresh, N., Balaban, N.Q.
<2>Optimization of lag time underlies antibiotic tolerance in evolved bacterial populations.
<3>Nature
<4>513
<5>418-421
<6>2014
<7>The great therapeutic achievements of antibiotics have been dramatically undercut by the
evolution of bacterial strategies that overcome antibiotic stress. These strategies fall into
two classes. 'Resistance' makes it possible for a microorganism to grow in the constant
presence of the antibiotic, provided that the concentration of the antibiotic is not too high.
'Tolerance' allows a microorganism to survive antibiotic treatment, even at high antibiotic
concentrations, as long as the duration of the treatment is limited. Although both resistance
and tolerance are important reasons for the failure of antibiotic treatments, the evolution of
resistance is much better understood than that of tolerance. Here we followed the evolution of
bacterial populations under intermittent exposure to the high concentrations of antibiotics
used in the clinic and characterized the evolved strains in terms of both resistance and
tolerance. We found that all strains adapted by specific genetic mutations, which became fixed
in the evolved populations. By monitoring the phenotypic changes at the population and
single-cell levels, we found that the first adaptive change to antibiotic stress was the
development of tolerance through a major adjustment in the single-cell lag-time distribution,
without a change in resistance. Strikingly, we found that the lag time of bacteria before
regrowth was optimized to match the duration of the antibiotic-exposure interval. Whole genome
sequencing of the evolved strains and restoration of the wild-type alleles allowed us to
identify target genes involved in this antibiotic-driven phenotype: 'tolerance by lag'
(tbl). Better understanding of lag-time evolution as a key determinant of the survival of
bacterial populations under high antibiotic concentrations could lead to new approaches to
impeding the evolution of antibiotic resistance.

<>

<1>Friedhoff, P., Franke, I., Krause, K.L., Pingoud, A.
<2>Cleavage experiments with deoxythymidine 3',5'-bis-(p-nitrophenyl phosphate) suggest that the homing endonuclease I-PpoI follows the same mechanism of phosphodiester bond hydrolysis as the non-specific Serratia nuclease.
<3>FEBS Lett.
<4>443
<5>209-214
<6>1999
<7>We show here that two nucleases, Serratia nuclease and I-PpoI, with contrasting specificities,
i.e. non-specific vs. highly sequence specific, share a structurally similar active site
region with conservation of the catalytically relevant histidine and asparagine residues.  On
the basis of a comparison of the available structures and biochemical data for wild type and
mutant variants of Serratia nuclease and I-PpoI we propose that both enzymes have a catalytic
mechanism, a proposition that is supported by our finding that both enzymes accept
deoxythymidine 3',5'-bis-(p-nitrophenyl phosphate) as a substrate and cleave it in an
identical manner.  According to this mechanism a histidine residue functions as a general base
and Mg2+ bound to an asparagine residue as a Lewis acid in phosphodiester bond cleavage.

<>

<1>Friedhoff, P., Lurz, R., Lueder, G., Pingoud, A.
<2>Sau3AI, a monomeric type II restriction endonuclease that dimerizes on the DNA and thereby induces DNA loops.
<3>J. Biol. Chem.
<4>276
<5>23581-23588
<6>2001
<7>Here, we report that Sau3AI, an unusually large type II restriction enzyme with sequence
homology to the mismatch repair protein MutH, is a monomeric enzyme as shown by gel filtration
and ultracentrifugation. Structural similarities in the N- and C-terminal halves of the
protein suggest that Sau3AI is a pseudo-dimer, i.e. a polypeptide with two similar domains.
Since Sau3AI displays a nonlinear dependence of cleavage activity on enzyme concentration and
a strong preference for substrates with two recognition sites over those with only one, it is
likely that the functionally active form of Sau3AI is a dimer of a pseudo-dimer. Indeed,
electron microscopy studies demonstrate that two distant recognition sites are brought
together through DNA looping induced by the simultaneous binding of two Sau3AI molecules to
the DNA. We suggest that the dimeric form of Sau3AI supplies two DNA-binding sites, one that
is associated with the catalytic center and one that serves as an effector site.

<>

<1>Friedman, J., Friedmann, A., Razin, A.
<2>Studies on the biological role of DNA methylation:  III.  Role in excision of one-genome long single-stranded Phi X174 DNA.
<3>Nucleic Acids Res.
<4>4
<5>3483-3496
<6>1977
<7>Accumulation of replicative intermediates of the bacteriophage Phi X174 was
observed in E. coli C infected cells when phage DNA methylation has been
inhibited by nicotinamide or when cells were infected with a
temperature-sensitive mutant in gene A.  Analysis of the accumulating
replicative intermediates by electron microscopy revealed that these molecules
are composed of double-stranded DNA rings with multiple-genome length
single-stranded "tails".  These results suggest that the single
5-methylcytosine residue present in the phage DNA serves as a recognition site
for the gene A protein mediating the excision of one-genome long phage DNA.

<>

<1>Friedman, J., Razin, A.
<2>Studies on the biological role of DNA methylation.  II.  Role of Phi X174 DNA methylation in the process of viral progeny DNA synthesis.
<3>Nucleic Acids Res.
<4>3
<5>2665-2675
<6>1976
<7>In vivo inhibition of bacteriophage Phi X174 DNA methylation by nicotinamide resulted in the
accumulation of replicative intermediates with multiple-genome length single-stranded "tails".
These abnormal replicative intermediates could not be chased into viral single-stranded
circular DNA.  The effect of nicotinamide on phage maturation and accumulation of abnormal
replicative intermediates could be reversed by washing out the inhibitor.  The results suggest
that the single methyl group present in the viral DNA serves as a recognition site for a
specific endonuclease, probably the gene A protein product, that is responsible for the
excision of the single-stranded one-genome long viral DNA, before final maturation of the
virus occurs.

<>

<1>Friedman, S.
<2>The irreversible binding of Azacytosine-containing DNA fragments to bacterial DNA (cytosine-5)methyltransferases.
<3>J. Biol. Chem.
<4>260
<5>5698-5705
<6>1985
<7>DNA containing 5-azacytosine is an irreversible inhibitor of DNA
(cytosine-5)methyltransferase.  This paper describes the binding of DNA
methyltransferase to 32P-labeled fragments of DNA containing 5-azacytosine.
The complexes were identified by gel electrophoresis.The EcoRII
methyltransferase specified by the R15 plasmid was purified from Escherichia
coli B(R15).  This enzyme methylates the second C in the sequence CCAGG and has
a molecular mass of 60,000 Da.  Specific binding of enzyme to DNA fragments
could be detected if either excess unlabeled DNA or 0.8% sodium dodecyl sulfate
was added to the reaction mixture prior to electrophoresis.  Binding was
dependent upon the presence of both the CCAGG sequence and azacytosine in the
DNA fragment.  S-Adenosylmethionine stimulated the formation of the complex.
The complex was stable to 6M urea but could be digested with pronase.  These
DNA fragments could be used to detect the presence of several different
methyltransferases in crude extracts of E. coli.  No DNA protein complexes
could be detected in E. coli B extracts, a strain that contains no
DNA(cytosine-5)methyltransferases.  The chromosomally determined methylase with
the same specificity as the purified EcoRII methylase could be detected in
crude extracts of E. coli K12 strains.  The MspI methylase cloned in E. coli
HB101 could also be detected in crude extracts.  These enzymes are the only
proteins that bind azacytosine-containing DNA in crude extracts of E. coli.

<>

<1>Friedman, S.
<2>The effect of 5-azacytidine on E. coli DNA methylase.
<3>Biochem. Biophys. Res. Commun.
<4>89
<5>1328-1333
<6>1979
<7>5-Azacytidine, when added to growing E. coli K12, causes a decrease in DNA
methylation assayed in vitro.  This decrease is greater when E. coli DNA is
used as substrate than when calf thymus DNA is used.  The decrease in activity
is not due to the inhibition of protein synthesis caused by this drug, since
neither chloramphenicol nor rifampin causes a decrease in enzyme activity.  The
effect is specific for the DNA(cytosine-5)methylase; the methylation of adenine
is not affected.  The concentration of drug that inhibits the DNA methylase by
50% is the same concentration that inhibits cell growth by 50%.

<>

<1>Friedman, S.
<2>The inhibition of DNA(Cytosine-5)methylases by 5-azacytidine:  The effect of azacytosine-containing DNA.
<3>Mol. Pharmacol.
<4>19
<5>314-320
<6>1981
<7>DNA extracted from Escherichia coli grown in the presence of 5-azacytidine
(azaC-DNA) inhibits the DNA(cytosine-5)methyltransferase extracted from E. coli
K12 cells without affecting the DNA(adenine-N6)methyltransferase present in
these same cells.  The inhibition is time-dependent and the rate of inhibition
can be decreased by addition of substrate DNA.  The inhibitory capacity of the
DNA is destroyed by incubation with pancreatic deoxyribonuclease or micrococcal
nuclease; however, the inhibited enzyme cannot be reactivated by treatment with
these enzymes.  The cytosine methylases methylate only double-stranded DNA.
Similarly, the inhibitory DNA loses activity if it is heat-denatured, and
regains activity upon reannealing.  The DNA will also inhibit the EcoRII and
HpaII modification methylases which also synthesize 5-methylcytosine in DNA.
Digestion of the inhibitory DNA with the respective restriction endonuclease
destroys, in part, the inhibitory activity of the DNA for the respective
methylase.  Base analysis indicated that 5-azacytidine replaced 8.4% of the
cytosine in the azaC-DNA.  These results suggest that DNA containing
5-azacytidine irreversibly inhibits DNA(cytosine-5)methylases.

<>

<1>Friedman, S.
<2>Adducts of the EcoRII methylase and 5-fluorocytosine-containing DNA.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>A436
<6>1991
<7>We have investigated the properties of the adduct formed between the EcoRII
methyltransferase and DNA containing 5-fluorocytosine.  A polymer of CCCA/TGGG
containing 75% substitution of cytosine by 5-fluorocytosine was synthesized
labeled with alpha-32P dATP.  Irreversible inhibition of the enzyme by the DNA
occurred over a 5 h time course.  This inhibition only occurred in the presence
of AdoMet.  The DNA was methylated by the enzyme when incubated with
(methyl-3H)AdoMet with a similar time course.  Digestion with DNase I yielded
an adduct that was stable to boiling in 1% SDS but was unstable to a pH <5 and
to ultrafiltration.  However, if the adduct were digested with trypsin a
portion of the DNA remained bound to a peptide as defined by its altered
mobility.  The adduct was therefore treated with staphylococcal protease and
the products purified by HPLC.  The radioactive peak containing both 32P was
further purified by polyacrylamide gel electrophoresis.  The region containing
32P was subjected to peptide sequencing for 20 cycles.  A sequence was obtained
which included a region of the protein that contains cysteine 186, a residue
that we had previously identified as being susceptible to photolabeling with
AdoMet and therefore part of the active site.

<>

<1>Friedman, S.
<2>Binding of the EcoRII methylase to azacytosine-containing DNA.
<3>Nucleic Acids Res.
<4>14
<5>4543-4556
<6>1986
<7>Binding of DNA (cytosine-5) methyltransferases to azacytosine containing DNA is
stimulated by the presence of S-adenosyl-methionine or its analogs sinefungin
or S-adenosyl-L-homocysteine.  Methylation of the DNA is therefore not
necessary for binding to occur.  There is no relationship between the affinity
of the analog for the EcoRII enzyme and its ability to stimulate binding.  The
DNA-enzyme complex partially dissociates on incubation in 0.1% sodium dodecyl
sulfate and 0.5 M ammonium acetate.  Some of this DNA could again form a tight
complex with enzyme, indicating that DNA-enzyme complex formation is
reversible.  Binding occurs when the second cytosine in the sequence CCAGG is
substituted by azacytosine.  This is the cytosine that would normally be
methylated by the enzyme.  The binding is therefore due to specific interaction
of the methylase with azacytosine at the site it would normally methylate.

<>

<1>Friedman, S., Ansari, N.
<2>Binding of the EcoRII methyltransferase to 5-fluorocytosine-containing DNA.  Isolation of a bound peptide.
<3>Nucleic Acids Res.
<4>20
<5>3241-3248
<6>1992
<7>The properties of the interaction of 5-fluorocytosine-containing DNA with the EcoRII
methyltransferase were studied. The DNA used was either a polymer synthesized in vitro, or a
20-mer containing one CCA/TGG sequence. The DNA could be methylated by the enzyme. In the
process the enzyme formed a tight binding adduct with the DNA that could be identified by
sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Enzyme activity was inhibited by
this interaction. The 20-mer could be used to titrate the active site of the enzyme. The DNA
polymer formed a tight binding complex that could be identified following digestion of the DNA
with pancreatic deoxyribonuclease or micrococcal nuclease. A peptide-DNA adduct could be
isolated after digestion of the EcoRII-DNA adduct with staphylococcal protease V8 by high
pressure liquid chromatography and polyacrylamide gel electrophoresis. Sequencing of the
peptide indicated the DNA bound to a region of the protein that is conserved in all
procaryotic DNA(cytosine-5)-methyltransferases. We have previously shown that this region
contains a cysteine that can be photomethylated with adenosylmethionine. This region, in
addition to forming part of, or being adjacent to, the AdoMet binding site, also forms part of
the DNA binding site.

<>

<1>Friedman, S., Cheong, L.C.
<2>Effect of 5-azacytidine on deoxyribonucleic acid methylation in Escherichia coli K12.
<3>Biochem. Pharmacol.
<4>33
<5>2675-2679
<6>1984
<7>5-azacytidine inhibits Escherichia coli DNA (cytosine-5) methylase when added to growing
cells.  The time-course of recovery of methylase activity and the appearance of
5-methylcytosine in DNA following removal of the drug was studied.  When E. coli K12 was
treated with 5-azacytidine for 30 min, DNA (cytosine-5) methylase levels decreased to less
than 10% of control levels and slowly recovered to control levels for three generations after
treatment and returned to control levels after six generations of growth.  In contrast,
beta-galactosidase levels in induced cells, which declined to 66% of control one generation
after treatment, returned to control by the third generation of growth.  The rate of induction
of beta-galactosidase had returned to the control rate two generations after growth resumed.
Since azacytidine-containing DNA inhibits DNA-cytosine methylases in vitro, the prolonged
inhibition of cytosine methylation in E. coli K12 following treatment with the drug could be
due to the persistence of the drug in DNA and thus inhibition of newly synthesized enzymes.

<>

<1>Friedman, S., Som, S.
<2>Induction of EcoRII methyltranserase:  Evidence for autogenous control.
<3>J. Bacteriol.
<4>175
<5>6293-6298
<6>1993
<7>The cytosine analog 5-azacytidline kills Escherichia coli cells that carry plamids expressing
EcoRII DNA (cytosine 5)methyltransferase under control of its own promoter. We previously
showed that this enzyme binds tightly to azacytidine-containing DNA in vitro and proposed that
such binding is lethal in vivo. In support of this proposal, we now show that the enzyme
sediments with the nucleoid of azabytidine-treated cells. Azacytidine treatment led to an
increase in the amount of enzyme, and this increase required sequences in the ecoRIIM promoter
region. Enzyme inducibility correlated with drug sensitivity: plasmids carrying the
methyltransferase gene but lacking the wild-type promoter did not confer sensitivity. These
results suggested that the ecoRIIM gene was under autogenous control. Transcriptional
ecoRIIM' -lacZ fusions in E. coli were, therefore, constructed. They showed that expression
from the ecoRIIM promoter was reversed by treating the cells with azacytidine. These results
provide evidence that the expression of the ecoRIIM gene is under autogenous regulation and
that call death induced by azacytidine is due, in part, to the disruption of autoregulation.

<>

<1>Friedman, S., Som, S., Yang, L.-F.
<2>The core element of the EcoRII methylase as defined by protease digestion and deletion analysis.
<3>Nucleic Acids Res.
<4>19
<5>5403-5408
<6>1991
<7>Binding of the EcoRII DNA methyltransferase to azacytosine-containing DNA
protects the enzyme from digestion by proteases.  The limit digest yields a
product having a Mr, on SDS-PAGE 20% less than the intact protein.  The N
terminus of the tryptic digestion product was sequenced and found to be missing
the N terminal 82 amino acids.  Under the conditions used unbound enzyme was
digested to small peptides.  Protection of the enzyme undergoes major
conformational changes when bound to DNA.  The trypsin sensitive region of the
EcoRII methyltransferase occurs prior to the first constant region shared with
other procaryotic DNA(cytosine-5)methyltransferases.  To determine if this
region played a role in substrate binding or specificity, N-terminal deletion
mutants were studied.  Deletion of 97 amino acids resulted in a decrease of
enzyme activity.  Further deletions caused a complete loss of activity.  Enzyme
deleted through amino acid 85 was purified and found to have the same
specificity as wild type however there was an increase in Km for both
S-adenosylmethionine (AdoMet) and DNA of 27 and 18 fold respectively.  The
N-terminus of the EcoRII methylase, although a variable region present in many
procaryotic DNA(cytosine-5)methylases, plays no role in determining enzyme
specificity, although it does contribute to the interaction with both AdoMet
and DNA.

<>

<1>Friedrich, T., Fatemi, M., Gowhar, H., Leismann, O., Jeltsch, A.
<2>Specificity of DNA binding and methylation by the M.FokI DNA methyltransferase.
<3>Biochim. Biophys. Acta
<4>1480
<5>145-159
<6>2000
<7>The M.FokI adenine-N(6) DNA methyltransferase recognizes the asymmetric DNA sequence
GGATG/CATCC. It consists of two domains each containing all motifs characteristic for
adenine-N(6) DNA methyltransferases. We have studied the specificity of DNA-methylation by
both domains using 27 hemimethylated oligonucleotide substrates containing recognition sites
which differ in one or two base pairs from GGATG or CATCC. The N-terminal domain of M.FokI
interacts very specifically with GATG-sequences, because only one of the altered sites is
modified. In contrast, the C-terminal domain shows lower specificity. It prefers
CATCC-sequences but only two of the 12 star sites (i.e. sites that differ in 1 bp from the
recognition site) are not accepted and some star sites are modified with rates reduced only
2-3-fold. In addition, GGATGC- and CGATGC-sites are modified which differ at two positions
from CATCC. DNA binding experiments show that the N-terminal domain preferentially binds to
hemimethylated GGATG/C(m)ATCC sequences whereas the C-terminal domain binds to DNA with higher
affinity but without specificity. Protein-protein interaction assays show that both domains of
M.FokI are in contact with each other. However, several DNA-binding experiments demonstrate
that DNA-binding of both domains is mutually exclusive in full-length M.FokI and both domains
do not functionally influence each other. The implications of these results on the molecular
evolution of type IIS restriction/modification systems are discussed.

<>

<1>Friedrich, T., Roth, M., Helm-Kruse, S., Jeltsch, A.
<2>Functional mapping of the EcoRV DNA methyltransferase by random mutagenesis and screening for catalytically inactive mutants.
<3>Biol. Chem.
<4>379
<5>475-480
<6>1998
<7>M.EcoRV is an alpha-adenine DNA methyltransferase.  According to structure predictions, the
enzyme consists of a catalytic domain, which has a structure similar to all other
DNA-methyltransferases, and a smaller DNA-recognition domain.  We have investigated this
enzyme by random mutagenesis, using error-prone PCR, followed by selection for catalytically
inactive mutants.  20 single mutants were identified that are completely inactive in vivo as
His6- and GST-fusion proteins.  13 of them could be overexpressed and purified.  All of these
mutants are also inactive in vitro.  5 of the mutations are located near the putative binding
site for a flipped adenine residue (C192R, D193G, E212G, W231R, N239H).  All of these variants
bind to DNA, demonstrating the importance of this region of the protein in catalysis.  Only
the W231R mutant could be purified with high yields.  It binds to DNA and AdoMet and, thus,
behaves like a bona fine active site mutant.  According to the structure prediction Trp231
corresponds to Val121 in M.HhaI, which forms a hydrophobic contact to the flipped target
cytosine.  4 of the remaining purified variants are located within a small region of the
putative DNA-recognition domain (F115S, F117L, S121P, C122Y).  F117L, S121P and C122Y are
unable to bind to DNA, suggesting a critical role of this region in DNA binding.  Taken
together, these results are in good agreement with the structural model of M.EcoRV.

<>

<1>Friend, P.L.
<2>Association of chloramphenicol resistance of group A streptococci with host-controlled restriction and modification of bacteriophages.
<3>Can. J. Microbiol.
<4>17
<5>1573-1576
<6>1971
<7>Bacteriophages grown on chloramphenicol (CM)-sensitive group A streptococci
plate efficiently only on other CM-sensitive strains, while phages propagated
on CM-resistant strains plate well only on CM-resistant hosts.  Data are
presented showing that the phages are subject to host-controlled modification
and restriction, and that these processes are related to the CM resistance
level of the host.

<>

<1>Friis, C., Wassenaar, T.M., Javed, M.A., Snipen, L., Lagesen, K., Hallin, P.F., Newell, D.G., Toszeghy, M., Ridley, A., Manning, G., Ussery, D.W.
<2>Genomic Characterization of Campylobacter jejuni Strain M1.
<3>PLoS ONE
<4>5
<5>e12253
<6>2010
<7>Campylobacter jejuni strain M1 (laboratory designation 99/308) is a rarely
documented case of direct transmission of C. jejuni from chicken to a
person, resulting in enteritis. We have sequenced the genome of C. jejuni
strain M1, and compared this to 12 other C. jejuni sequenced genomes
currently publicly available. Compared to these, M1 is closest to strain
81116. Based on the 13 genome sequences, we have identified the C. jejuni
pan-genome, as well as the core genome, the auxiliary genes, and genes
unique between strains M1 and 81116. The pan-genome contains 2,427 gene
families, whilst the core genome comprised 1,295 gene families, or about
two-thirds of the gene content of the average of the sequenced C. jejuni
genomes. Various comparison and visualization tools were applied to the 13
C. jejuni genome sequences, including a species pan- and core genome plot,
a BLAST Matrix and a BLAST Atlas. Trees based on 16S rRNA sequences and on
the total gene families in each genome are presented. The findings are
discussed in the background of the proven virulence potential of M1.

<>

<1>Fritsch, A., Tiollais, P., Buc, H.
<2>Preparative purification of lambda-DNA fragments obtained after EcoRI digestion.
<3>FEBS Lett.
<4>52
<5>121-126
<6>1975
<7>The study of the expression, and structure of bacterial or eukaryotic genes, as
well as the study of the interaction of a particular gene with specific
proteins, require the isolation of individual genes, or groups of genes.  In
bacteria, this can be done with specialized transducing phages, whose DNA is
hydrolysed by restriction enzymes, and the interesting DNA fragment is then
purified.  In a previous publication, the purification of the E. coli lac gene,
via the DNA of a lac tranducing lambda bacteriophage has been reported.  In the
present work, we describe the purification of all the EcoRI digestion products
of the DNA of two mutants of lambda bacteriophage.  Two methods have been used:
polyacrylamide gel electrophoresis, and isopycnic centrifugation in cesium
sulfate gradients in the presence of silver ions.  As will be seen, the proper
combination of both methods allows the purification of all fragments of lambda
genomes with either two, or six sites sensitive to EcoRI digestion.

<>

<1>Fritsche, P., Alves, J.
<2>A monomeric mutant of restriction endonuclease EcoRI nicks DNA without sequence specificity.
<3>Biol. Chem.
<4>385
<5>975-985
<6>2004
<7>We have mutated the monomer-monomer interface of the restriction endonuclease EcoRI in order
to destabilize the homodimer and to stabilize heterodimers. Mutations of Leu158 to charged
amino acid
residues result in strong destabilization of the dimer. The largest
effect was detected for the L158D mutant which is monomeric even at
higher concentrations. It unspecifically degrades DNA by cleaving both
single strands independently every 15 nucleotides on the average.
Although cleavage is reproducible, it is not determined by nucleotide
sequence but by general properties like conformation or deformability
as has been found for other unspecific nucleases.
Mutations of Ile230, which is in direct contact with Leu158 of the
other subunit, cause structural changes with the loss of about ten
percent alpha-helix content, but interfere only marginally with
homodimerization and double strand cleavage. Again the mutation to
aspartate shows the strongest effects. Mixtures of single mutants, one
containing aspartate at one of the two positions and the other lysine
at the corresponding position, form heterodimers. These are mainly
stabilized compared to the homodimers by reestablishment of the
wildtype hydrophobic interaction at the not mutated residues while an
interaction of aspartate and lysine seems energetically unfavorable in
this structural context.

<>

<1>Fritz, A., Kuster, W., Alves, J.
<2>Asn141 is essential for DNA recognition by EcoRI restriction endonuclease.
<3>FEBS Lett.
<4>438
<5>66-70
<6>1998
<7>The amino acid residue Asn141 of the restriction endonuclease EcoRI was proposed to make three
hydrogen bonds to both adenine residues within the recognition sequence GAATTC.  We have
mutated Asn141 to alanine, aspartate, serine, and tyrosine.  Only the serine mutant is active
under normal buffer conditions although 1000-fold less than wild-type EcoRI.  The alanine and
aspartate mutants can be activated by Mn2+.  At acidic pH the latter mutant becomes even more
active than the wild-type enzyme in the presence of Mn2+.  We conclude that Asn141 is
essential for DNA recognition and that serine can partly substitute it.

<>

<1>Fritzenwanker, M., Kuenne, C., Billion, A., Hain, T., Zimmermann, K., Goesmann, A., Chakraborty, T., Domann, E.
<2>Complete Genome Sequence of the Probiotic Enterococcus faecalis Symbioflor 1 Clone DSM 16431.
<3>Genome Announcements
<4>1
<5>e00165-12
<6>2013
<7>Here, we report the complete and annotated genome sequence of the probiotic Enterococcus
faecalis Symbioflor 1 clone DSM 16431, included in a commercial
probiotic product used for more than 50 years without any reports of infection.
This sequence will provide new insights into the biology of this nonpathogenic
and probiotic microorganism.

<>

<1>Froman, B.E., Tait, R.C., Kado, C.I., Rodriguez, R.L.
<2>Purification of restriction endonuclease XcyI from Xanthomonas cyanopsidis.
<3>Gene
<4>28
<5>331-335
<6>1984
<7>A new type II restriction endonuclease XcyI, purified from Xanthomonas cyanopsidis 13D5, is an
isoschizomer of SmaI and XmaI that cleaves at the nucleotide sequence 5'-C^CCGG-3' of
double-stranded DNA. The single restriction activity present in this strain permits rapid
purification of 8000 units of cleavage activity from 10 g of freshly harvested cells. The
resulting XcyI preparation is free of contaminating nuclease activities that interfere with in
vitro manipulation of DNA.

<>

<1>Frommer, M., McDonald, L.E., Millar, D.S., Collis, C.M., Watt, F., Grigg, G.W., Molloy, P.L., Paul, C.L.
<2>A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>1827-1831
<6>1992
<7>The modulation of DNA-protein interactions by methylation of protein-binding
sites in DNA and the occurrence in genomic imprinting, X chromosome
inactivation, and fragile X syndrome of different methylation patterns in DNA
of different chromosomal origin have underlined the need to establish
methylation patterns in individual strands of particular genomic sequences.  We
report a genomic sequencing method that provides positive identification of
5-methylcytosine residues and yields strand-specific sequences of individual
molecules in genomic DNA.  The method utilizes bisulfite-induced modification
of genomic DNA, under conditions whereby cytosine is converted to uracil, but
5-methylcytosine remains nonreactive.  The sequence under investigation is then
amplified by PCR with two sets of strand-specific primers to yield a pair of
fragments, one from each strand, in which all uracil and thymine residues have
been amplified as thymine and only 5-methylcytosine residues have been
amplified as cytosine.  The PCR products can be sequenced directly to provide a
strand-specific average sequence for the population of molecules or can be
cloned and sequenced to provide methylation maps of single DNA molecules.  We
tested the method by defining the methylation status within single DNA strands
of two closely spaced CpG dinucleotides in the promoter of the human kininogen
gene.  During the analysis, we encountered in sperm DNA an unusual methylation
pattern, which suggests that the high methylation level of single-copy
sequences in sperm may be locally modulated by binding of protein factors in
germ-line cells.

<>

<1>Froseth, B.R., Harlander, S.K., McKay, L.L.
<2>Plasmid-mediated reduced phage sensitivity in Streptococcus lactis KR5.
<3>J. Dairy Sci.
<4>71
<5>275-284
<6>1988
<7>The phage insensitivity of Streptococcus lactis KR5 was evaluated for its
possible linkage to plasmid DNA.  This strain possessed plasmids of
40,29,26,21,16.5,10.5,7.8, and 1.5 Mdal.  Plasmid curing using novobiocin
resulted in derivatives with increased sensitivity to prolate-headed phage,
suggesting the involvement of plasmid DNA in phage insensitivity.
Transformation of S. lactis LM0230 protoplasts with the KR5 plasmid DNA pool
produced transformants containing a plasmid of about 27 Mdal.  These
erythromycin-resistant transformants were lactose-positive phage-sensitive or
were lactose-negative and exhibited a reduced sensitivity to phage.  Agarose
gel electrophoresis and restriction endonuclease digestion analysis showed the
17-Mdal plasmid band to be composed of two distinct plasmids of 26 Mdal (pBF61)
and 29 Mdal (pBF62), which coded for reduced phage sensitivity and
lactose-positive phenotypes, respectively.  The mechanisms of reduced phage
sensitivity encoded by pBF61 included a restriction/modification system and a
mechanism that resulted in reduced plaque size independent of incubation
temperature.  These results further support the involvement of plasmid DNA in
the mechanisms for reduced phage sensitivity in dairy streptococci.

<>

<1>Frostesjo, L., Holm, I., Grahn, B., Page, A.W., Bestor, T.H., Heby, O.
<2>Interference with DNA methyltransferase activity and genome methylation during F9 teratocarcinoma stem cell differentiation induced by polyamine depletion.
<3>J. Biol. Chem.
<4>272
<5>4359-4366
<6>1997
<7>When ornithine decarboxylase, the initial and highly regulated enzyme in polyamine
biosynthesis, is irreversibly inactivated by alpha-difluoromethylornithine, F9 teratocarcinoma
stem cells are depleted of putrescine and spermidine and as a result differentiate into a cell
type which phenotypically resembles the parietal endoderm cells of the early mouse embryo.
Simultaneously the level of decarboxylated S-adenosylmethionine (dcAdoMet), the amino propyl
group donor in spermidine and spermine synthesis, increases dramatically, as the aminopropyl
group acceptor molecules (putrescine and spermidine) become limiting.  When this excessive
accumulation of dcAdoMet is prevented by specific inhibition of the AdoMet decarboxylase
activity, the differentiative effect is counteracted, despite the fact that the extent of
polyamine depletion remains almost identical.  Therefore, it may be concluded that dcAdoMet
plays an important role in the induction of differentiation.  Moreover, this key metabolite
acts as a competitive inhibitor of DNA methyltransferase and is therefore capable of
interfering with the maintenance methylation of newly replicated DNA.  During the course of F9
cell differentiation, the highly methylated genome is gradually demethylated, and its pattern
of gene expression is changed.  Our present findings, that the DNA remains highly methylated
and that the differentiative process is counteracted when the build-up of dcAdoMet is
prevented, provide strong evidence for a causative relation between the level of cdAdoMet and
the state of DNA methylation as well as cell differentiation.

<>

<1>Fry, P.R., Calcutt, M.J., Foecking, M.F., Hsieh, H.Y., Suntrup, D.G., Perry, J., Stewart, G.C., Middleton, J.R.
<2>Draft Genome Sequence of Staphylococcus chromogenes Strain MU 970, Isolated from  a Case of Chronic Bovine Mastitis.
<3>Genome Announcements
<4>2
<5>e00835-14
<6>2014
<7>Coagulase-negative staphylococcal species are a common cause of subclinical bovine mastitis,
with Staphylococcus chromogenes being one of the most frequently
identified species in these cases. The draft genome sequence of an S. chromogenes
isolate (MU 970) recovered from the milk of a cow with a chronic intramammary
infection is reported here.

<>

<1>Fsihi, H., Vincent, V., Cole, S.T.
<2>Homing events in the gyrA gene of some mycobacteria.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>3410-3415
<6>1996
<7>The A subunit of DNA gyrase in Mycobacterium leprae, unlike its
counterpart in Mycobacterium tuberculosis, is produced by protein splicing as its gene,
gyrA, harbors a 1260-bp in-frame insertion encoding an intein, a putative homing
endonuclease.  Analysis of the gyrA locus from different mycobacterial species revealed the
presence of inteins in Mycobacterium flavescens, Mycobacterium gordonae, and
Mycobacterium kansasii but not in 10 other pathogenic or saprophytic mycobacteria.  In all
four cases where intein coding sequences were found, they were localized in the same
position in gyrA, immediately downstream of the codon for the key active-site residue Tyr-
130.  The intein products were similar, but not identical, in sequence and the splice
junctions displayed all the features found in other polypeptides known to be produced by
protein splicing from a precursor protein.  Paired motifs, found in homing endonucleases
encoded by some group I RNA introns, and inteins showing endonuclease activity, were
present in the gyrA inteins as were other intein-specific signatures.  Some strains of
M.flavescens, M. gordonae, and M. kansasii were shown by PCR analysis to have
inteinless gyrA genes, in contrast to the situation in M. leprae where all the isolates
possessed insertions in gyrA.  Sequencing of the corresponding regions revealed that,
although the GyrA protein sequence was conserved, the nucleotide sequences differed in
gyrA genes with and without inteins, suggesting that the homing endonuclease displays
sequence specificity.

<>

<1>Fu, A.Q., Genereux, D.P., Stoger, R., Burden, A.F., Laird, C.D., Stephens, M.
<2>Statistical Inference of In Vivo Properties of Human DNA Methyltransferases from Double-Stranded Methylation Patterns.
<3>PLoS ONE
<4>7
<5>e32225
<6>2012
<7>DNA methyltransferases establish methylation patterns in cells and transmit these patterns
over cell generations, thereby influencing each
cell's epigenetic states. Three primary DNA methyltransferases have
been identified in mammals: DNMT1, DNMT3A and DNMT3B. Extensive in
vitro studies have investigated key properties of these enzymes, namely
their substrate specificity and processivity. Here we study these
properties in vivo, by applying novel statistical analysis methods to
double-stranded DNA methylation patterns collected using
hairpin-bisulfite PCR. Our analysis fits a novel Hidden Markov Model
(HMM) to the observed data, allowing for potential bisulfite conversion
errors, and yields statistical estimates of parameters that quantify
enzyme processivity and substrate specificity. We apply this model to
methylation patterns established in vivo at three loci in humans: two
densely methylated inactive X (Xi)-linked loci (FMR1 and G6PD), and an
autosomal locus (LEP), where methylation densities are tissue-specific
but moderate. We find strong evidence for a high level of processivity
of DNMT1 at FMR1 and G6PD, with the mean association tract length being
a few hundred base pairs. Regardless of tissue types, methylation
patterns at LEP are dominated by DNMT1 maintenance events, similar to
the two Xi-linked loci, but are insufficiently informative regarding
processivity to draw any conclusions about processivity at that locus.
At all three loci we find that DNMT1 shows a strong preference for
adding methyl groups to hemi-methylated CpG sites over unmethylated
sites. The data at all three loci also suggest low (possibly 0)
association of the de novo methyltransferases, the DNMT3s, and are
consequently uninformative about processivity or preference of these
enzymes. We also extend our HMM to reanalyze published data on mouse
DNMT1 activities in vitro. The results suggest shorter association
tracts (and hence weaker processivity), and much longer non-association
tracts than human DNMT1 in vivo.

<>

<1>Fuchs, B.M., Spring, S., Teeling, H., Quast, C., Wulf, J., Schattenhofer, M., Yan, S., Ferriera, S., Johnson, J., Glockner, F.O., Amann, R.
<2>Characterization of a marine gammaproteobacterium capable of aerobic anoxygenic photosynthesis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>2891-2896
<6>2007
<7>Members of the gammaproteobacterial clade NOR5/OM60 regularly form an abundant
part, up to 11%, of the bacterioplankton community in coastal systems during the
summer months. Here, we report the nearly complete genome sequence of one
cultured representative, Congregibacter litoralis strain KT71, isolated from
North Sea surface water. Unexpectedly, a complete photosynthesis superoperon,
including genes for accessory pigments, was discovered. It has a high sequence
similarity to BAC clones from Monterey Bay [Beja O, Suzuki MT, Heidelberg JF,
Nelson WC, Preston CM, et al. (2002) Nature 415:630-633], which also share a
nearly identical gene arrangement. Although cultures of KT71 show no obvious
pigmentation, bacteriochlorophyll a and spirilloxanthin-like carotenoids could be
detected by HPLC analysis in cell extracts. The presence of two potential BLUF
(blue light using flavin adenine dinucleotide sensors), one of which was found
adjacent to the photosynthesis operon in the genome, indicates a light- and
redox-dependent regulation of gene expression. Like other aerobic anoxygenic
phototrophs (AAnPs), KT71 is able to grow neither anaerobically nor
photoautotrophically. Cultivation experiments and genomic evidence show that KT71
needs organic substrates like carboxylic acids, oligopeptides, or fatty acids for
growth. The strain grows optimally under microaerobic conditions and actively
places itself in a zone of approximately 10% oxygen saturation. The genome
analysis of C. litoralis strain KT71 identifies the gammaproteobacterial marine
AAnPs, postulated based on BAC sequences, as members of the NOR5/OM60 clade. KT71
enables future experiments investigating the importance of this group of
gammaproteobacterial AAnPs in coastal environments.

<>

<1>Fuchs, C., Rosenvold, E., Honigman, A., Szybalski, W.
<2>Identification of palindromic sequences recognized by restriction endonucleases, as based on the tabularized sequencing data for seven viral and plasmid DNAs.
<3>Gene
<4>10
<5>357-370
<6>1980
<7>Computer search  of DNA sequences for phages PhiX174, G4, M13 and fd, plasmids
pBR322 and pA3, and virus SV40, was employed to prepare tables specifying the
size classes and frequencies of DNA segments located between all possible
tetra-, penta, and hexanucleotide palindromes.  As described earlier (Fuchs et
al., 1978), these tables permit identifying sequences recognized by most of the
restriction endonucleases.  The effect of sequencing errors on the accuracy of
the present identification method is evaluated.  Only four of the 224 listed
sequences do not appear in any of the seven DNAs, leading to discussion (see
Appendix) on the natural sequence distribution.

<>

<1>Fuchs, C., Rosenvold, E.C., Honigman, A., Szybalski, W.
<2>A simple method for identifying the palindromic sequences recognized by restriction endonucleases: the nucleotide sequence of the AvaII site.
<3>Gene
<4>4
<5>1-23
<6>1978
<7>Tables specifying the frequencies, distances between and positions of all
possible tetra-, penta-, and hexanucleotide palindromes*** in PhiX174 and SV40
viral DNAs were prepared by a computer search of their base sequences.  A
simple method based on these tables is described for identifying the sequence
recognized by any specific restriction endonuclease.  The method requires
experimental determination of the number and approximate sizes of the fragments
obtained by digestion of PhiX174 RF and SV40 DNAs.  Using this method we
identified the sequence for the AvaII restriction endonuclease as 5'-GG(AT)CC.

<>

<1>Fuchs, L.Y., Covarrubias, L., Escalante, L., Sanchez, S., Bolivar, F.
<2>Characterization of a site-specific restriction endonuclease SphI from Streptomyces phaeochromogenes.
<3>Gene
<4>10
<5>39-46
<6>1980
<7>A new type-II restriction endonuclease SphI, has been partially purified from
Streptomyces phaeochromogenes.  SphI recognizes the hexanucleotide sequence
5'-GCATG^C and cleaves it at the postiion marked by the arrow.  This nucleotide
sequence is present twice in SV40 DNA, four times in lambda DNA and only once
in the cloning vehicles pBR322, pBR325, pBR327 and PBR328.

<>

<1>Fuchs, R., Blakesley, R.
<2>Guide to the Use of Type II Restriction  Endonucleases.
<3>Methods Enzymol.
<4>100
<5>3-38
<6>1983
<7>Type II restriction endonucleases are DNases that recognize specific
oligonucleotide sequences, make double-strand cleavages, and generate unique,
equal molar fragments of a DNA molecule.  By the nature of their controllable,
predictable, infrequent, and site-specific cleavage of DNA, restriction
endonucleases proved to be extremely useful as tools in dissecting, analyzing,
and reconfiguring genetic information at the molecular level.  Over 350
different restriction endonucleases have been isolated from a wide variety of
prokaryotic sources, representing at least 85 different recognition sequences.
A number of excellent reviews detail the variety of restriction enzymes and
their sources, their purification and determination of their sequence
specificity, ad their physical properties, kinetics, and reaction mechanism.
Here we provide a summary, based on the literature and our experience in this
laboratory, emphasizing the practical aspects for using restriction
endonucleases as tools.  This review focuses on the reaction, its components
and the conditions that affect enzymic activity and sequence fidelity, methods
for terinating the reaction, some reaction variations, and a troubleshooting
guide to help identify and solve restriction endonuclease-related problems.

<>

<1>Fuchs, R., Kane, J.F.
<2>Expression of the genes for the Pst restriction-modification enzymes after cloning into a temperature-sensitive replication plasmid.
<3>Miami BioTechnology Winter Symposium, Academic Press, Ahmad, F., Schultz, J., Smith, E.E., Whelan, W.J., 
<4>19
<5>523
<6>1982
<7>Our objective was to develop a model system to study the use of an
overexpression plasmid to increase synthesis of a restriction enzyme in
Escherichia coli.  We chose the restriction (PstI)-modification system of
Providencia stuartii that has been cloned into the HindIII site of pBR322
(Walder, et al., 1981, P.N.A.S. 78:1503).  One hybrid plasmid, designated
pPst102, contained a 5.9 kb insert with an internal HindIII site, producing 1.9
and 4.0 kb fragments.  Although both enzymes were specified by the 4.0 kb
fragment, the genes were poorly expressed in E. coli.  We subcloned the Pst
genes into a temperature sensitive runaway plasmid (pBEU I) and transformed E.
coli HB101.  Four independent transformants were isolated and characterized:
pRF I (1.9 kb insert); pRF 2 (4.0 kb insert); pRF 3 and pRF 4 (4.9 kb insert).
All four strains were subjected to temperature shifts to allow for
overexpression of the plasmid.  Crude cell extracts were prepared and examined
for PstI.  When the PstI activity of E. coli/pPst 102 is assigned a value
enzymatic activities were estimated in the four strains: pRF I <0.5; pRF 2
<0.5; pRF3 <0.5; pRF 4-10.0.  These results demonstrate that the overexpression
plasmid produces a 10-fold increase in the activity of PstI in E. coli.
Furthermore, this system provides a basis for studying the regulation of the
PstI gene.

<>

<1>Fuchu, G., Ohtsubo, Y., Ito, M., Miyazaki, R., Ono, A., Nagata, Y., Tsuda, M.
<2>Insertion sequence-based cassette PCR: cultivation-independent isolation of gamma-hexachlorocyclohexane-degrading genes from soil DNA.
<3>Appl. Microbiol. Biotechnol.
<4>79
<5>627-632
<6>2008
<7>gamma-Hexachlorocyclohexane (gamma-HCH) is a highly chlorinated pesticide
that has caused serious environmental problems. Based on the frequently
observed association of insertion sequence IS6100 with lin genes for
gamma-HCH degradation in several gamma-HCH-degrading bacterial strains
isolated to date, DNA fragments flanked by two copies of IS6100 were
amplified by nested polymerase chain reaction (PCR) technique using a DNA
sample extracted from soil contaminated with HCH. Four distinct DNA
fragments with sizes of 6.6, 2.6, 1.6, and 1.3 kb were obtained, three of
which carried lin genes: the 6.6-kb fragment carried linD and linE as well
as linR; the 2.6-kb fragment showed a truncated form of linF; and the
1.6-kb fragment carried linB. Our approach, named as insertion sequence
(IS)-based cassette PCR, was successful in the isolation of the lin genes
from HCH-contaminated soil without cultivation of host cells and is
applicable for the culture-independent isolation of other functional genes
bordered by other IS elements.

<>

<1>Fucik, V., Grunnerova, H., Zadrazil, S.
<2>Restriction and modification in Bacillus subtilis 168: Regulation of hsrM(nonB) expression in spoOA mutants and effects on permissiveness for Phi15 and Phi105 phages.
<3>Mol. Gen. Genet.
<4>186
<5>118-121
<6>1982
<7>Gene hsrM (nonB) of Bacillus subtilis 168, causing non-permissiveness to phage
SP10 (Saito et al. 1979) and reduced plating efficiency of unmodified phage
Phi105, is responsible for non-permissiveness of B. subtilis 168 for phages
Phi15 and PZA.  Upon transformation to sporulation deficiency (allele spoOA) B.
subtilis 168 becomes permissive for Phi15 and PAZ and loses the ability to
restrict Phi105.  spoOA str-1 double transformants of B. subtilis 168, however,
retain the restriction 168 and non-permissiveness for Phi15 and PZA phages, in
spite of their Spo- phenotype.  Therefore it appears that a functional product
of the spoOA gene is required for expression of gene hsrM in wild-type
bacteria, but is not essential in streptomycin-resistant bacteria.  Phage
genomes (PZA) were trapped in spores of the restriction deficient strain with
much higher efficiency than in the wild-type.

<>

<1>Fuerst, C.R.
<2>Plating efficiencies of modified lambda-bio particles on temperature-sensitive hsd mutants of Escherichia coli K12.
<3>Virology
<4>143
<5>352-356
<6>1985
<7>Two mutants of Escherichia coli K12 that are temperature sensitive in cell growth and lambda
phage production are shown to contain at least two mutations.  One of the mutations in each of
the isolates is in the hsd locus, and modification and restriction of lambda exhibits
temperature sensitivity. One of the hsd mutations causes plaque formation by modified
lambda-bio particles that do not contain an intact ral gene to be temperature dependent.

<>

<1>Fuglsang, A.
<2>Distribution of potential type II restriction sites (palindromes) in prokaryotes.
<3>Biochem. Biophys. Res. Commun.
<4>310
<5>280-285
<6>2003
<7>Restriction-modification systems are used as a defensive mechanism against inappropriate
invasion of foreign DNA. The recognition sequences for the
common type II restriction enzymes and their corresponding methylases are
usually palindromes. In this study, we identified the most over- and
underrepresented words in DNA of four bacteria: Escherichia coli, Bacillus
subtilis, Clostridium perfringens, and Pseudomonas aeruginosa. Using
maximum order Markov chain analysis, we found that palindromic words were
most often more underrepresented than their non-palindromic counterparts.
No strict rule for the intragenic palindrome content could be derived, but
for three of the bacteria there was a weak correlation between codon usage
bias and palindrome content. A clear drop in palindrome counts was
observed in the Shine-Dalgarno region for B. subtilis and C. perfringens,
but not in E. coli or P. aeruginosa. It was also shown that palindromes in
eubacteria and archaebacteria seem to occur slightly more infrequently
than expected on the basis of the genomic GC-content, but some exceptions
to this principle exist.

<>

<1>Fujihara, Y., Miyasako, H., Kato, K., Hayashi, T., Toraya, T.
<2>Molecular Cloning, Expression, and Characterization of Starfish DNA (Cytosine-5)-methyltransferases.
<3>Biosci. Biotechnol. Biochem.
<4>76
<5>1661-1671
<6>2012
<7>To determine whether and if so how a DNA methylation-dependent epigenetic mechanism for
transcriptional gene silencing functions in
Echinoderms, we cloned and sequenced dnmt1 and dnmt3 cDNAs of the
starfish Asterina pectinifera. Since the Strongylocentrotus purpuratus
genome has only two loci of DNA (cytosine-5)-methyltransferase genes
encoding Dnmt1 and Dnmt3, they might constitute a sufficient set of
timid genes in Echinoderms. The starfish Dnmt3 whose cDNA we cloned
showed highest homology to a mammalian Dnmt3a2 splicing variant.
Essentially all the characteristic motifs and sequences of the
mammalian counterparts were found in the starfish Dnmts as well, except
that a typical PCNA binding domain motif was lacking in the starfish
Dnmt1. RT-PCR analysis indicated that the dnmt1 mRNA exists in both
ovary and oocytes, but its levels in other tissues were very low or
almost negligible. In contrast, the dnmt3 mRNA was detected only in the
ovary, and not at all in the oocytes. The size of a dnmt1 transcript
was about 6.5 kb on Northern blot analysis. On heterologous expression,
the starfish Dnmt1 protein was expressed in insect cells in
catalytically active form.

<>

<1>Fujimoto, D., Srinivasan, P.R., Borek, E.
<2>On the nature of the deoxyribonucleic acid methylases.  Biological evidence for the multiple nature of the enzymes.
<3>Biochemistry
<4>4
<5>2849-2855
<6>1965
<7>The minor components of DNA of several bacterial species were determined by growing the
organisms in the presence of [methyl-14C]methionine and analyzing the labeled bases by isotope
dilution.  The DNA's were found to contain either 6-methylaminopurine or both
6-methylaminopurine and 5-methylcytosine.  The interaction of extracts derived from some of
these organisms with a variety of DNA's was also examined.  The results of these studies
suggest that the DNA methylases as well as the substrate DNA's possess directive specificity
regarding the pattern of methylation.  The findings confirm that the DNA methylases are
species specific and also demonstrate the existence of two types of DNA methylases: one
capable of methylating adenine and the other cytosine.  Infection by T2 bacteriophage elevates
the DNA methylase activity of both E. coli B and E. coli K12.  Extracts of E. coli B can
methylate only adenine of DNA, whereas in E. coli K12 two methylating capacities are normally
present, producing 5-methylcytosine and 6-methylaminopurine.  However, on phage infection only
the level of the adenine methylase was elevated.

<>

<1>Fujimoto, K., Iida, H., Kawakami, M., Bando, T., Tao, Z.-F., Sugiyama, H.
<2>Sequence-specific protection of plasmid DNA from restriction endonuclease hydrolysis by pyrrole-imidazole-cyclopropapyrroloindole conjugates.
<3>Nucleic Acids Res.
<4>30
<5>3748-3753
<6>2002
<7>The pyrrole-imidazole (Py-Im) triamide-cyclopropapyrroloindole (CPI) conjugates ImPyImLDu86
(7) and ImImPyLDu86 (14) were synthesized and their alkylating activities and inhibitory
effects on DNA hydrolysis by restriction endonucleases were examined. Sequencing gel analysis
demonstrated that conjugates 7 and 14 specifically alkylated DNA at 5'-CGCGCG-3' and
5'-PyGGCCPu-3', respectively. Agarose gel electrophoresis indicated that incubation of a
supercoiled plasmid, pSPORT I (4109 bp), with conjugate 7 effectively inhibited its hydrolysis
by BssHII (5'-G_CGCGC-3'), whereas conjugate 14 had no effect on this hydrolysis. These
results suggest that conjugate 7 sequence-specifically inhibits the hydrolysis of DNA by
BssHII. Sequence-specific alkylation by the Py-Im triamide-CPI conjugates was further
confirmed by inhibition of the Eco52I (5'-C_GGCCG-3') hydrolysis of conjugate 14-treated
pQBI PGK (5387 bp). In clear contrast, hydrolysis of pQB1 PGK by DraI (3'-TTT_AAA-3') was
not inhibited by 5 micro M conjugate 14. That ImImPy did not inhibit the hydrolysis of pQB1
PGK indicates that covalent bond formation is necessary for inhibition. A similar experiment,
using linear pQBI PGK, achieved the same extent of protection of the DNA with approximately
half the concentration of conjugate 14 as was required to protect supercoiled DNA from
hydrolysis.

<>

<1>Fujimoto, R., Sasaki, T., Nishio, T.
<2>Characterization of DNA methyltransferase genes in Brassica rapa.
<3>Plant Cell Physiol.
<4>47
<5>S238
<6>2006
<7>Cytosine methylation is an epigenetic process that plays a role in regulating gene expression.
Plants have at least three classes of cytosine methyltransferase, i.e., MET1 class of
methyltransferase, Chromomethylases, and de novo DNA methyltransferase.  In our study three
putative methyltransferase genes, BrMET1a, BrMET1b, and BrCMT were isolated from genome
library of Brassica rapa.  Identities of amino acid sequences between BrMET1a and atMET1 and
between BrMET1b and AtMET1 were 72.0% and 66.7%, respectively.  Pfam analysis using deduced
amino acid sequence showed the presence of BAH domain, CHROMO domain, and methyltransferase
domain in BrCMT.  RT-PCR analysis revealed the expression of BrMET1a, BrMET1b, and BrCMT.
These results suggested that these three genes function as methyltransferase in B. rapa.

<>

<1>Fujimoto, R., Sasaki, T., Nishio, T.
<2>Characterization of DNA methyltransferase genes in Brassica rapa.
<3>Genes Genet. Syst.
<4>81
<5>235-242
<6>2006
<7>DNA methylation is essential for normal development and plays important roles in regulating
gene expression in plants. Analysis of the key
enzymes catalyzing DNA methylation is important to understand
epigenetic phenomena. In this study, three putative methyltransferase
genes, BrMET1a, BrMET1b, and BrCMT, were isolated from a genome library
of Brassica rapa. Structural conservation of the amino acid sequence
between BrMET1a/BrMET1b and AtMET1 and that between BrCMT and AtCMT3
suggests that they may function as DNA methyltransferase. BrMET1a was
expressed in vegetative and reproductive organs, while BrMET1b was
expressed only in pistils, indicating that these two genes have
different functions. BrCMT was expressed especially in stamens at the
stage of 2-4 days before anthesis. We isolated three DNA
methyltransferase genes in Brassica rapa and indicated differences of
expression patterns of these DNA methyltransferase genes and expression
levels in different tissues and developmental stages, suggesting that
these genes might play important roles in epigenetic gene regulation in
B. rapa.

<>

<1>Fujinami, S., Oikawa, Y., Araki, T., Shinmura, Y., Midorikawa, R., Ishizaka, H., Kato, C., Horikoshi, K., Ito, M., Tamegai, H.
<2>Genome Sequence of the Deep-Sea Denitrifier Pseudomonas sp. Strain MT-1, Isolated from the Mariana Trench.
<3>Genome Announcements
<4>2
<5>e01313-14
<6>2014
<7>Pseudomonas sp. strain MT-1 was the first deep-sea denitrifier isolated and characterized from
mud recovered from a depth of 11,000 m in the Mariana Trench.
We report here the genome sequence of this bacterium, which contributes to our
understanding of denitrification and bioenergetics in the deep sea.

<>

<1>Fujinami, S., Takarada, H., Kasai, H., Sekine, M., Omata, S., Harada, T., Fukai, R., Hosoyama, A., Horikawa, H., Kato, Y., Nakazawa, H., Fujita, N.
<2>Complete genome sequence of Ilumatobacter coccineum YM16-304(T.).
<3>Standards in Genomic Sciences
<4>8
<5>430-440
<6>2013
<7>Ilumatobacter coccineum YM16-304(T) (=NBRC 103263(T)) is a novel marine actinobacterium
isolated from a sand sample collected at a beach in Shimane
Prefecture, Japan. Strain YM16-304(T) is the type strain of the species.
Phylogenetically, strain YM16-304(T) is close to Ilumatobacter nonamiense
YM16-303(T) (=NBRC 109120(T)), Ilumatobacter fluminis YM22-133(T) and some
uncultured bacteria including putative marine sponge symbionts. Whole genome
sequence of these species has not been reported. Here we report the complete
genome sequence of strain YM16-304(T). The 4,830,181 bp chromosome was predicted
to encode a total of 4,291 protein-coding genes.

<>

<1>Fujinami, S., Takeda, K., Onodera, T., Satoh, K., Sano, M., Narumi, I., Ito, M.
<2>Draft Genome Sequence of Sodium-Independent Alkaliphilic Microbacterium sp. Strain TS-1.
<3>Genome Announcements
<4>1
<5>e01043-13
<6>2013
<7>Alkaliphilic Microbacterium sp. strain TS-1, newly isolated from the jumping spider, showed
Na(+)-independent growth and motility. Here, we report the draft
genome sequence of this bacterium, which may provide beneficial information for
Na(+)-independent alkaline adaptation mechanisms.

<>

<1>Fujinami, S., Takeda, K., Onodera, T., Satoh, K., Sano, M., Narumi, I., Ito, M.
<2>Draft Genome Sequence of Potassium-Dependent Alkaliphilic Bacillus sp. Strain TS-2, Isolated from a Jumping Spider.
<3>Genome Announcements
<4>2
<5>e00458-14
<6>2014
<7>The potassium-dependent alkaliphilic Bacillus sp. strain TS-2 was isolated from the mashed
extract of a jumping spider, and its draft genome sequence was
obtained. Comparative genomic analysis with a previously sequenced
sodium-dependent alkaliphilic Bacillus species may reveal potassium-dependent
alkaline adaptation mechanisms.

<>

<1>Fujinami, S., Takeda-Yano, K., Onodera, T., Satoh, K., Sano, M., Takahashi, Y., Narumi, I., Ito, M.
<2>Draft Genome Sequence of Calcium-Dependent Paenibacillus sp. Strain TCA20, Isolated from a Hot Spring Containing a High Concentration of Calcium Ions.
<3>Genome Announcements
<4>2
<5>e00866-14
<6>2014
<7>Calcium-dependent Paenibacillus sp. strain TCA20 was isolated from a water sample of a hot
spring containing a high concentration of calcium ions. Here, we report
the draft genome sequence of this bacterium, which may be the basis for the
research of calcium ion homeostasis.

<>

<1>Fujisawa, H., Yonesaki, T., Minagawa, T.
<2>Sequence of the T4 recombination gene, uvsX, and its comparison with that of the recA gene of Escherichia coli.
<3>Nucleic Acids Res.
<4>13
<5>7473-7481
<6>1985
<7>We have determined the nucleotide sequence of the uvsX gene of
bacteriophage T4 which is involved in DNA recombination and damage repair,
and whose product catalyzes in vitro reactions related to recombination
process in analogous manners to E. coli recA gene product. The coding
region consisted of 1170 nucleotides directing the synthesis of a
polypeptide of 390 amino acids in length with a calculated molecular
weight of 43,760. Amino acid composition, the sequence of seven
NH2-terminal amino acids and molecular weight of the protein deduced from
the nucleotide sequence were consistent with the data from the analysis of
the purified uvsX protein. The nucleotide sequence and the deduced amino
acid sequence were compared with those of the recA gene. Although a
significant homology was not found in the nucleotide sequences, the amino
acid sequences included 23% of identical and 15% of conservatively
substituted residues.

<>

<1>Fujisawa, T. et al.
<2>Genomic Structure of an Economically Important Cyanobacterium, Arthrospira (Spirulina) platensis NIES-39.
<3>DNA Res.
<4>17
<5>85-103
<6>2010
<7>A filamentous non-N-2-fixing cyanobacterium, Arthrospira (Spirulina) platensis, is an
important organism for industrial applications and as
a food supply. Almost the complete genome of A. platensis NIES-39 was
determined in this study. The genome structure of A. platensis is
estimated to be a single, circular chromosome of 6.8 Mb, based on
optical mapping. Annotation of this 6.7 Mb sequence yielded 6630
protein-coding genes as well as two sets of rRNA genes and 40 tRNA
genes. Of the protein-coding genes, 78% are similar to those of other
organisms; the remaining 22% are currently unknown. A total 612 kb of
the genome comprise group II introns, insertion sequences and some
repetitive elements. Group I introns are located in a protein-coding
region. Abundant restriction-modification systems were determined.
Unique features in the gene composition were noted, particularly in a
large number of genes for adenylate cyclase and haemolysin-like
Ca2+-binding proteins and in chemotaxis proteins. Filament-specific
genes were highlighted by comparative genomic analysis.

<>

<1>Fukao, M., Oshima, K., Morita, H., Toh, H., Suda, W., Kim, S.W., Suzuki, S., Yakabe, T., Hattori, M., Yajima, N.
<2>Genomic Analysis by Deep Sequencing of the Probiotic Lactobacillus brevis KB290 Harboring Nine Plasmids Reveals Genomic Stability.
<3>PLoS ONE
<4>8
<5>e60521
<6>2013
<7>We determined the complete genome sequence of Lactobacillus brevis KB290, a probiotic lactic
acid bacterium isolated from a traditional Japanese fermented vegetable. The genome contained
a 2,395,134-bp chromosome that housed 2,391 protein-coding genes and nine plasmids that
together accounted for 191 protein-coding genes. KB290 contained no virulence factor genes,
and several genes related to presumptive cell wall-associated polysaccharide biosynthesis and
the stress response were present in L. brevis KB290 but not in the closely related L. brevis
ATCC 367. Plasmid-curing experiments revealed that the presence of plasmid pKB290-1 was
essential for the strain's gastrointestinal tract tolerance and tendency
to aggregate. Using next-generation deep sequencing of current and 18-year-old stock strains
to detect low frequency variants, we evaluated genome stability. Deep sequencing of four
periodic KB290 culture stocks with more than 1,000-fold coverage revealed 3 mutation sites and
37 minority variation sites, indicating long-term stability and providing a useful method for
assessing the stability of industrial bacteria at the nucleotide level.

<>

<1>Fuks, F., Burgers, W.A., Brehm, A., Hughes-Davies, L., Kouzarides, T.
<2>DNA methyltransferase Dnmt1 associates with histone deacetylase activity.
<3>Nat. Genet.
<4>24
<5>88-91
<6>2000
<7>The DNA methyltransferase Dnmt1 is responsible for cytosine methylation in mammals and has a
role in gene silencing. DNA methylation represses genes partly by recruitment of the
methyl-CpG-binding protein MeCP2, which in turn recruits a histone deacetylase activity. Here
we show that Dnmt1 is itself associated with histone deacetylase activity in vivo. Consistent
with this association, we find that one of the known histone deacetylases, HDAC1, has the
ability to bind Dnmt1 and can purify methyltransferase activity from nuclear extracts. We have
identified a transcriptional repression domain in Dnmt1 that functions, at least partly, by
recruiting histone deacetylase activity and shows homology to the repressor domain of the
trithorax-related protein HRX (also known as MLL and ALL-1). Our data show a more direct
connection between DNA methylation and histone deacetylation than was previously considered.
We suggest that the process of DNA methylation, mediated by Dnmt1, may depend on or generate
an altered chromatin state via histone deacetylase activity.

<>

<1>Fuks, F., Burgers, W.A., Godin, N., Kasai, M., Kouzarides, T.
<2>Dnmt3a binds deacetylases and is recruited by a sequence-specific repressor to silence transcription.
<3>EMBO J.
<4>20
<5>2536-2544
<6>2001
<7>The Dnmt3a DNA methyltransferase is essential for mammalian development and is responsible for
the generation of genomic methylation patterns, which lead to transcriptional silencing. Here,
we show that Dnmt3a associates with RP58, a DNA-binding transcriptional repressor protein
found at transcriptionally silent heterochromatin. Dnmt3a acts as a co-repressor for RP58 in a
manner that does not require its de novo methyltransferase activity. Like other characterized
co-repressors, Dnmt3a associates with the histone deacetylase HDAC1 using its ATRX-homology
domain. This domain of Dnmt3a represents an independent transcriptional repressor domain whose
silencing functions require HDAC activity. These results identify Dnmt3a as a co-repressor
protein carrying deacetylase activity and show that Dnmt3a can be targeted to specific
regulatory foci via its association with DNA-binding transcription factors.

<>

<1>Fukuda, E., Kaminska, K.H., Bujnicki, J.M., Kobayashi, I.
<2>Cell death upon epigenetic genome methylation: a novel function of methyl-specific deoxyribonucleases.
<3>Genome Biology
<4>9
<5>R163
<6>2008
<7>Background Alteration in epigenetic methylation can affect gene expression and
other processes. In Prokaryota, DNA methyltransferase genes frequently
move between genomes and present a potential threat. A methyl-specific
deoxyribonuclease, McrBC, of Escherichia coli cuts invading methylated
DNAs. Here we examined whether McrBC competes with genome methylation
systems through host killing by chromosome cleavage.
Results
McrBC inhibited the establishment of a plasmid carrying a PvuII
methyltransferase gene but lacking its recognition sites, likely
through the lethal cleavage of chromosomes that become methylated.
Indeed, its phage-mediated transfer caused McrBC-dependent chromosome
cleavage. Its induction led to cell death accompanied by chromosome
methylation, cleavage and degradation. RecA/RecBCD functions affect the
chromosome processing and, together with SOS response, reduce the
lethality. Our evolutionary/genomic analyses of mcrBC homologs revealed
(i) wide distribution in Prokaryota, (ii) frequent distant horizontal
transfer and linkage with mobility-related genes, (iii) diversification
in the DNA binding domain. In these features, McrBCs resemble Type II
restriction-modification systems, which behave as selfish mobile
elements maintaining their frequency by host killing. McrBCs are
frequently found linked with a methyltransferase homolog, which
suggests a functional association.
Conclusions
Our experiments indicate McrBC can respond to genome methylation
systems by host killing. Combined with our evolutionary/genomic
analyses, they support our hypothesis McrBCs have evolved as mobile
elements competing with specific genome methylation systems through
host killing. To our knowledge, this represents the first report of a
defense system against epigenetic systems through cell death.

<>

<1>Fukuda, E., Kobayashi, I.
<2>Suicidal defense against an epigenetic system: a role for a Type IV methyl-dependent restriction enzyme.
<3>Genes Genet. Syst.
<4>81
<5>408
<6>2006
<7>Attack on the host cell chromosome by restriction enzymes is a subject of various biological
interactions. When several Type II restriction-modification gene complexes enter a new host
cell, they try to avoid chromosome breakage and cell killing by expressing the modification
enzyme first. Once they establish themselves, they attack the host chromosome when their
presence is threatened. Through this postsegregational killing strategy, they force their
maintenance on the host. On the other
hand, Escherichia coli cells employ a Type IV restriction enzyme McrBC, which recognizes
methylated DNAs and cleaves DNA between two of these recognition sites. We hypothesize that
McrBC serves for suicidal defense against invasion of specific DNA methylation systems. In
this work, we analyze host attack by McrBC. We demonstrate McrBC-mediated abortion of
establishment of a modification enzyme gene in E. coli. We also demonstrate McrBC-dependent
cell death and chromosomal DNA degradation following genome methylation by RM systems. These
results support our hypothesis. To our knowledge, this represents the first example of direct
suicidal defense against an epigenetic system.

<>

<1>Fukuda, K., Hosoyama, A., Tsuchikane, K., Ohji, S., Yamazoe, A., Fujita, N., Shintani, M., Kimbara, K.
<2>Complete Genome Sequence of Polychlorinated Biphenyl Degrader Comamonas testosteroni TK102 (NBRC 109938).
<3>Genome Announcements
<4>2
<5>e00865-14
<6>2014
<7>Comamonas testosteroni TK102 (NBRC 109938; JCM 19603) can utilize biphenyl as a sole carbon
source and degrade polychlorinated biphenyls (PCBs). The complete
nucleotide sequence of the TK102 genome was determined. TK102 possesses several
integrative and conjugative element-like regions, and one of them carries
biphenyl-degradative genes.

<>

<1>Fukuda, S., Toh, H., Hase, K., Oshima, K., Nakanishi, Y., Yoshimura, K., Tobe, T., Clarke, J.M., Topping, D.L., Suzuki, T., Taylor, T.D., Itoh, K., Kikuchi, J., Morita, H., Hattori, M., Ohno, H.
<2>Bifidobacteria can protect from enteropathogenic infection through production of acetate.
<3>Nature
<4>469
<5>543-547
<6>2011
<7>The human gut is colonized with a wide variety of microorganisms,
including species, such as those belonging to the bacterial genus Bifidobacterium, that have
beneficial effects on human physiology and pathology1-3. Among the most distinctive benefits
of bifidobacteria are modulation of host defence responses and protection against infectious
diseases4-6. Nevertheless, the molecular mechanisms underlying these effects have barely been
elucidated. To investigate these mechanisms, we used mice associated with certain
bifidobacterial strains and a simplified model of lethal infection with enterohaemorrhagic
Escherichia coli O157:H7, together with an integrated 'omics' approach. Here we show that
genes encoding an ATP-binding-cassette-type carbohydrate transporter present in certain
bifidobacteria contribute to protecting mice against death induced by E. coli O157:H7. We
found that this effect can be attributed, at least in part, to increased production of acetate
and that translocation of the E. coli O157:H7 Shiga toxin from the gut lumen to the blood was
inhibited. We propose that acetate produced by protective bifidobacteria improves intestinal
defence mediated by epithelial cells and thereby protects the host against lethal infection.

<>

<1>Fukuda, T., Nogami, S., Ohya, Y.
<2>VDE-initiated intein homing in Saccharomyces cerevisiae proceeds in a meiotic recombination-like manner.
<3>Genes Cells
<4>8
<5>587-602
<6>2003
<7>BACKGROUND: Inteins and group I introns found in prokaryotic and eukaryotic organisms
occasionally behave as mobile genetic elements.
During meiosis of the yeast Saccharomyces cerevisiae, the site-specific
endonuclease encoded by VMA1 intein, VDE, triggers a single double-strand
break (DSB) at an inteinless allele, leading to VMA1 intein homing.
Besides the accumulating information on the in vitro activity of VDE, very
little has been known about the molecular mechanism of intein homing in
yeast nucleus. RESULTS: We developed an assay to detect the product of
VMA1 intein homing in yeast genome. We analysed mutant phenotypes of RecA
homologs, Rad51p and Dmc1p, and their interacting proteins, Rad54p and
Tid1p, and found that they all play critical roles in intein inheritance.
The absence of DSB end processing proteins, Sae2p and those in the
Mre11-Rad50-Xrs2 complex, also causes partial reduction in homing
efficiency. As with meiotic recombination, crossover events are frequently
observed during intein homing. We also observed that the absence of
premeiotic DNA replication caused by hydroxyurea (HU) or clb5delta
clb6delta mutation reduces VDE-mediated DSBs. CONCLUSION: The repairing
system working in intein homing shares molecular machinery with meiotic
recombination induced by Spo11p. Moreover, like Spo11p-induced DNA
cleavage, premeiotic DNA replication is a prerequisite for a VDE-induced
DSB. VMA1 intein thus utilizes several host factors involved in meiotic
and recombinational processes to spread its genetic information and
guarantee its progeny through establishment of a parasitic relationship
with the organism.

<>

<1>Fukuda, T., Ohta, K., Ohya, Y.
<2>Investigation of the mechanism of meiotic DNA cleavage by VMA1-derived endonuclease uncovers a meiotic alteration in chromatin structure  around the target site.
<3>Euk. Cell
<4>5
<5>981-990
<6>2006
<7>VMA1-derived endonuclease (VDE), a homing endonuclease in Saccharomyces cerevisiae, is encoded
by the mobile intein-coding sequence within the
nuclear VMA1 gene. VDE recognizes and cleaves DNA at the 31-bp VDE
recognition sequence (VRS) in the VMA1 gene lacking the intein-coding
sequence during meiosis to insert a copy of the intein-coding sequence
at the cleaved site. The mechanism underlying the meiosis specificity
of VMA1 intein-coding sequence homing remains unclear. We studied
various factors that might influence the cleavage activity in vivo and
found that VDE binding to the VRS can be detected only when DNA
cleavage by VDE takes place, implying that meiosis-specific DNA
cleavage is regulated by the accessibility of VDE to its target site.
As a possible candidate for the determinant of this accessibility, we
analyzed chromatin structure around the VRS and revealed that local
chromatin structure near the VRS is altered during meiosis. Although
the meiotic chromatin alteration exhibits correlations with DNA binding
and cleavage by VDE at the VMA1 locus, such a chromatin alteration is
not necessarily observed when the VRS is embedded in ectopic gene loci.
This suggests that nucleosome positioning or occupancy around the VRS
by itself is not the sole mechanism for the regulation of
meiosis-specific DNA cleavage by VDE and that other mechanisms are
involved in the regulation.

<>

<1>Fukuda, T., Ohta, K., Ohya, Y.
<2>Strategies of VDE for propagation in yeast nuclear genome.
<3>Genes Genet. Syst.
<4>80
<5>438
<6>2005
<7>VDE, a homing endonuclease in Saccharomyces cerevisiae, is encoded by a mobile intein within
the nuclear gene, VMA1.  VDE causes double-strand  break at the 31-bp VDE-recognition sequence
in intein-less VMA1 allele during meiosis. The DSB is then repaired using the
intein-containing allele as a template, leading to the conversion of intein-less allele to
intein-containing allele. These processes are called "homing", by which VDE spreads throughout
the population.  The study on DSB repair process of homing revealed that VDE-introduced DSB is
repaired by homologous recombination system working in meiotic recombination as if it is one
of the programmed meiotic DSBs.  To elucidate the meiosis-specificity of DSB formation by VDE,
we examined several possibilities.  We found that none of expression, endonuclease activity,
and nuclear import necessarily regulates the meiosis-specific DSB by VDE.  On the other hand,
chromatin structure around VRS alters during meiosis. Chromatin immunoprecipitation revealed
that VDE loading onto VRS occurs only when chromatin changes around VRS.  These results raise
the possibility that chromatin configuration, which is modulated by the intrinsic property of
VRS, reulates the meiosis-specific integration of VDE.  Thus, several host factors are
involved in homing at meiosis-specific DSB formation and DSB repair processes to increase the
copy number of VDE.

<>

<1>Fukui, T., Atomi, H., Kanai, T., Matsumi, R., Fujiwara, S., Imanaka, T.
<2>Complete genome sequence of the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1 and comparison with Pyrococcus genomes.
<3>Genome Res.
<4>15
<5>352-363
<6>2005
<7>The genus Thermococcus, comprised of sulfur-reducing hyperthermophilic archaea, belongs to the
order Thermococcales in Euryarchaeota along with
the closely related genus Pyrococcus. The members of Thermococcus are
ubiquitously present in natural high-temperature environments, and are
therefore considered to play a major role in the ecology and metabolic
activity of microbial consortia within hot-water ecosystems. To obtain
insight into this important genus, we have determined and annotated the
complete 2,088,737-base genome of Thermococcus kodakaraensis strain KOD1,
followed by a comparison with the three complete genomes of Pyrococcus
spp. A total of 2306 coding DNA sequences (CDSs) have been identified,
among which half (1165 CDSs) are annotatable, whereas the functions of 41%
(936 CDSs) cannot be predicted from the primary structures. The genome
contains seven genes for probable transposases and four virus-related
regions. Several proteins within these genetic elements show high
similarities to those in Pyrococcus spp., implying the natural occurrence
of horizontal gene transfer of such mobile elements among the order
Thermococcales. Comparative genomics clarified that 1204 proteins,
including those for information processing and basic metabolisms, are
shared among T. kodakaraensis and the three Pyrococcus spp. On the other
hand, among the set of 689 proteins unique to T. kodakaraensis, there are
several intriguing proteins that might be responsible for the specific
trait of the genus Thermococcus, such as proteins involved in additional
pyruvate oxidation, nucleotide metabolisms, unique or additional metal ion
transporters, improved stress response system, and a distinct restriction
system.

<>

<1>Fuller, T.E., Wilson, T.L., Martin, S., Klein, L.K.
<2>Anti-bacterial vaccine compositions.
<3>International Patent Office
<4>WO 2006048753 A
<5>
<6>2006
<7>The present invention relates generally to the identification of genes responsible for
virulence of streptococci, thereby allowing for production of novel attenuated mutant strains
useful in vaccines and identification of new anti-bacterial agents that target the virulence
genes and their products.

<>

<1>Fuller-Pace, F.V., Bullas, L.R., Delius, H., Murray, N.E.
<2>Genetic recombination can generate altered restriction specificity.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>81
<5>6095-6099
<6>1984
<7>A recombinant strain, isolated following the transduction of an Escherichia
coli recipient carrying the Salmonella typhimurium (SB) specificity genes with
DNA from a donor having the Salmonella potsdam (SP) specificity, was shown
[Bullas, L.R., Colson, C. and Van Pel, A. (1976) J. Gen. Microbiol. 95,
166-172] to have neither SB nor SP specificity but to encode a novel
restriction specificity, SQ.  The heteroduplex analysis of the hsdS
(specificity) genes of the SB and SP restriction and modification systems
described here identifies a conserved sequence of around 100 base pairs flanked
by two nonhomologous regions each of approximately 500 base pairs.  This
organization parallels that previously deduced from the DNA sequences of the
hsdS genes of the related E. coli K-12, B, and D restriction systems.  The
present heteroduplex analyses further show that the hsdS gene conferring the SQ
specificity deserves one nonhomologous region from the SB gene and the other
from the SP gene, as predicted from genetic exchange within the conserved
sequence.  This finding supports the idea that two domains of an hsdS
polypeptide, which are different for each specificity, may correlate with two
regions of the DNA sequence recognized.  It has been shown that the recognition
sequences for E. coli K-12 and B each consist of two short oligonucleotide
sequences interrupted by a nonspecific sequence.  A similar organization is
suggested for the Salmonella specificity systems, providing the potential for
evolutionary diversification of restriction specificities as a result of
recombination within the conserved sequence of the hsdS gene.

<>

<1>Fuller-Pace, F.V., Cowan, G.M., Murray, N.E.
<2>EcoA and EcoE:  Alternatives to the EcoK Family of Type I Restriction and Modification Systems of Escherichia coli.
<3>J. Mol. Biol.
<4>186
<5>65-75
<6>1985
<7>The genes (hsd A) encoding EcoA, a restriction and modification system first
identified in Escherichia coli 15T-, behave in genetic crosses as alleles of
the genes (hsd K) encoding the archetypal type I restriction and modification
system of E. coli K12.  Nevertheless, molecular experiments have failed to
detect relatedness between the A and K systems.  We have cloned the hsd A genes
and have identified, on the basis of DNA homology, related genes (hsd E)
conferring a new specificity to a natural isolate of E. coli.  We show that the
overall organization of the genes encoding EcoA and EcoE closely parallels that
for EcoK.  Each enzyme is encoded by three genes, of which only one, hsdS,
confers the specificity of DNA interaction.  The three genes are in the same
order as those encoding EcoK, i.e. hsdR, hsdM and hsdS and, similarly, they
include a promter between hsdR and hsdM from which the M and S genes can be
transcribed.  The evidence indicates that EcoA and EcoE are type I restriction
and modification enzymes, but they appear to identify an alternative family to
EcoK.  For both families, the hsdR polypeptide is by far the largest, but the
sizes of the other two polypeptides are reversed, with the smallest polypeptide
of EcoK being the product of hsdS, and the smallest for the EcoA family being
the product of hsdM.  Physiologically, the A restriction and modification
system differs from that of K and its relatives, in that A-specific methylation
of unmodified DNA is particularly effective.

<>

<1>Fuller-Pace, F.V., Murray, N.E.
<2>Two DNA recognition domains of the specificity polypeptides of a family of type I restriction enzymes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>83
<5>9368-9372
<6>1986
<7>The hsd genes of Salmonella typhimurium and Salmonella potsdam encode related
type I restriction and modification systems designated SB and SP, respectively;
the polypeptide encoded by the hsdS gene dictates the DNA sequence recognized.
The hsdS genes of the SB and SP systems have a conserved sequence of around 100
base pairs flanked by two nonhomologous (variable) regions of around 500 base
pairs.  Recombination between the hsdS genes of SB and SP generated a system
(SQ) with a different recognition specificity.  We have localized the position
of the crossover in the central conserved region by analysis of nucleotide
sequences.  Concomitant with the generation of a new combination of flanking
variable regions is the recombination of minor differences in the central
conserved region.  A polypeptide domain encoded on the 5' side of the crossover
dictates recognition of the trinucleotide component of the target sequence, and
a second domain encoded on the 3' side of the crossover, similarly governs
recognition of the tetra- or penta-nucleotide component.  Our analysis
implicates at least parts of the variable regions in the determination of the
specificity of interaction between protein and DNA.  Furthermore, the
trinucleotide components of the recognition sequences of S. typhimurium and
Escherichia coli K-12 are identical, and the 5' segments of their hsdS genes
are strikingly homologous rather than variable.

<>

<1>Fung, W.T.
<2>Functional analysis of the two subunits of DNA methyltransferase EcoHK31I.
<3>Ph.D. Thesis, Chinese Univ. of Hong Kong, China
<4>
<5>1-224
<6>2006
<7>Methylation of cytosine residues in DNA occurs in diverse organisms from bacteria to humans.
In higher eukaryotic organisms cytosine-C5 methyltransferase is the only type of DNA MTase and
it plays an important role in controlling a number of cellular processes including
transcription, genomic imprinting and DNA repair.  In bacteria, there are three types of
MTases, mC4-, mC5- and mA6-, classified according to the methylation site of the DNA.  MTase
and its cognate restriction endonuclease form restriction-modification system.  The role of
MTase is to protect the host from its own ENase digestion while the ENase acts to degrade the
invasion of foreign DNA.  Sequence comparison of nearly 50 bacterial mC5-MTases has shown that
these enzymes share an overall common protein architecture.  Ten conserved motifs (I to X),
each 10 to 20 amino acids in length, have been identified, five of which are highly conserved
(I, IV, VI, VIII and X).  In addition, all of these enzymes have a hypervariable region lying
between motifs VIII and IX.  It is called the target recognition domain, and is responsible
for the specificity of DNA recognition and the choice of base to be methylated.  All
mC5-MTases are monomeric enzymes, except M. EcoHK31I and M.AquI which are MTases composed of
two polypeptides.  M.EcoHK31I is a mC5-MTase which recognizes the sequence 5-YGGCCR-3' and
consists of polypeptide a and b, with the latter gene encoded in an alternative reading frame
of the former.  All of the conserved motifs in mC5-MTases can be found in polypeptide a,
except motif IX, which is located in polypeptide b.  Both polypeptides are required for in
vitro methylation.  Since both of the polypeptides a and b of M.EcoHK31I are sequenced and
cloned into the expression vector separately, the role of DNA recognition and subunits
interaction of individual polypeptides can be studied.  By electromobility shift assay, we
found that polypeptides a and b complex recognize specific double strand oligos substrate.
Polypeptide a-DNA formed aggregates and polypeptide b alone did not bind DNA.  Therefore,
polypeptide b assists the proper binding of polypeptide a to DNA substrate.  Complex of
polypeptide a and a polypeptide b variant with N-terminal deletion of 41 amino acids showed a
16-fold reduction in methylation activity.  Further deletion resulted in an inactive MTase. By
surface plasmon resonance assay, the dissociation equilibrium constant of polypeptides a and b
complex was found to be 56.2nM and the KD for polypeptide a and deltaN46-polypeptide b complex
was increased by about 95 folds, contributing by a drastic decrease in dissociate rate
constant and an increase in association rate constant.  This indicated that the N-terminal
region of polypeptide b takes part in subunit interaction.  To pinpoint which amino acid
residues located at the variable region of polypeptide a are important for DNA binding and
subunits interaction, "charge-to-alanine scanning mutagenesis" were performed on 16 charge
residues between Asp213 and Glu271 in the small domain.  It was found that the five charge
residues upstream of motif X are not required for activity.  For other residues except K225,
E240 and D245, the protein is active when the same charge is maintained.

<>

<1>Fung, W.T., Sze, K.H., Lee, K.F., Shaw, P.C.
<2>Functional studies of the small subunit of EcoHK31I DNA methyltransferase.
<3>Biol. Chem.
<4>387
<5>507-513
<6>2006
<7>EcoHK311 DNA methyltransferase recognizes the sequence 5'-YGGCCR-3' and adds a methyl group
to the fifth position of the internal cytosine to
protect the DNA from cleavage by its cognate endonuclease. M.EcoHK311
is composed of polypeptides alpha and beta. Polypepticle beta only
contains the conserved IX motif of the C5-MTase family, and provides a
unique example to show that this motif alone may be dislocated to
another polypeptide. By electromobility shift assay, we found that the
alpha/beta complex recognizes specific oligonucleotide substrates.
Polypeptide alpha formed aggregates with DNA, while polypeptide p alone
did not bind DNA. Therefore, polypeptide beta assists in the proper
binding of polypeptide alpha to DNA substrate. The complex of
polypeptide alpha and a polypeptide beta variant with an N-terminal
deletion of 41 amino acids showed a 16-fold reduction in methylation
activity. Further deletion resulted in an inactive methyltransferase.
The dissociation equilibrium constant (K-d) of the alpha/beta complex
was 56.4 nM, while the K-d value for the alpha/Delta N46-polypeptide
beta complex was increased approximately 95-fold, caused by a drastic
decrease in dissociate rate constant (k(d)) and an increase in the
association rate constant (k(a)). This indicates that the N-terminal
region of polypeptide beta takes part in subunit interaction, while the
C-terminal region is involved in DNA binding.

<>

<1>Funo, K., Kitagawa, W., Tanaka, M., Sone, T., Asano, K., Kamagata, Y.
<2>Draft Genome Sequence of Tomitella biformata AHU 1821T, Isolated from a Permafrost Ice Wedge in Alaska.
<3>Genome Announcements
<4>2
<5>e00066-14
<6>2014
<7>Tomitella biformata AHU 1821(T) was isolated and cultured from a permafrost ice wedge, aged
presumably about 25,000 years, in the Fox permafrost tunnel (64.952
degrees N 147.617 degrees W), Alaska. These genome data provide the basis for
investigating T. biformata AHU 1821(T), identified as a long-term survivor of the
extremely cold and closed environment.

<>

<1>Furmanek, B., Gromek, K., Sektas, M., Kaczorowski, T.
<2>Isolation and characterization of type IIS restriction endonuclease from Neisseria cuniculi ATCC 14688.
<3>FEMS Microbiol. Lett.
<4>196
<5>171-176
<6>2001
<7>Neisseria cuniculi produces the restriction enzyme NcuI which is an isoschizomer of MboII. We
have demonstrated that NcuI recognizes a
pentanucleotide sequence (5'-GAAGA-3'/3'-CTTCT-5'), and cleaves the DNA
8 and 7 nucleotides downstream from the recognition site leaving a
single 3'-protruding nucleotide. We have purified this enzyme to
electrophoretic homogeneity using a four-step chromatographic procedure.
NcuI endonuclease is a monomeric protein with an Mr = 48,000 +/- 1,000
under denaturing conditions. The properties of NcuI are consistent with
those for MboII, the position of the cleavage site being identical and
the pH profile and divalent cation requirements being similar.
Moreover, NcuI cross-reacts strongly with anti-MboII serum suggesting
the presence of similar antigenic determinants. We have determined the
sequence of 20 N-terminal amino acids for NcuI and concluded that this
sequence is identical to the N-terminal portion of the MboII enzyme.

<>

<1>Furmanek, B., Sektas, M., Wons, E., Kaczorowski, T.
<2>Molecular characterization of the DNA methyltransferase M1.NcuI from Neisseria cuniculi ATCC 14688.
<3>Res. Microbiol.
<4>158
<5>164-174
<6>2007
<7>The methyltransferase M1.NcuI is a member of the restriction-modification system in Neisseria
cuniculi ATCC14688 and recognizes the asymmetric
pentanucleotide sequence 5'-GAAGA-3'/3'-CTTCT-5'. We purified M1.NcuI to
electrophoretic homogeneity using a four-step chromatographic procedure.
M1.NcuI is a protein with M(r)=32,000+/-1000 under denaturing conditions.
It modifies the recognition sequence by transferring the methyl group from
S-adenosyl-l-methionine to the 3' adenine of the pentanucleotide sequence
5'-GAAGA-3'. M1.NcuI, like many other methyltransferases, occurs as a
monomer in solution, as determined by gel filtration. Divalent cations
inhibit the methylation activity of M1.NcuI. Optimal enzyme activity was
observed at a pH of 8.0. M1.NcuI cross-reacted with anti-M1.MboII serum
which reflects the similarity of M1.NcuI with M1.MboII at the amino acid
level. The gene coding for the enzyme, designated ncuIM1, was cloned,
sequenced and overexpressed in Escherichia coli. The structural gene is
780 nucleotides in length coding for a protein of 259 amino acids (M(r)
30,098). The presence and distribution of nine highly conserved amino acid
sequence motifs and a putative target recognition domain in the enzyme
structure suggest that M1.NcuI, similar to M1.MboII and M1.HpyAII, belongs
to N(6)-adenine beta-class DNA methyltransferases.

<>

<1>Furmanek-Blaszk, B.
<2>Phenotypic and molecular characteristics of an Aeromonas hydrophila strain isolated from the River Nile.
<3>Microbiol. Res.
<4>169
<5>547-552
<6>2014
<7>Aeromonas hydrophila, an inhabitant of aquatic ecosystems found in most parts of  the world,
has considerable virulence potential. The polymerase chain reaction
technique was used to assay for the presence of five virulence factor genes:
haemolytic toxins aerA and ahh1, elastase ahyB, the enterotoxin act, and the
polar flagella flaA/flaB in the A. hydrophila strain isolated from the River
Nile. Drug screening showed high levels of resistance to beta-lactam antibiotics
and tetracycline. Slime production was determined by the Congo red agar plate
test. The isolate produced two restriction enzymes named AehI and AehII which are
isoschizomers of XhoI and StuI respectively. The complete nucleotide sequence of
the cryptic plasmid pAhy2.5 (2524 bp) from this strain was determined. Sequence
analysis revealed the presence of two open reading frames (ORFs) encoding
putative proteins. The protein coded by ORF1 is homologous with Rep proteins of
plasmids belonging to the pC194 family, which are known to replicate by the
rolling-circle mechanism. The putative double-strand origin of replication and a
region with palindromic sequences that could function as a single-strand origin
were detected in pAhy2.5.

<>

<1>Furmanek-Blaszk, B., Boratynski, R., Zolcinska, N., Sektas, M.
<2>M1.Mboll and M2.Mboll type IIS methyltransferases: different specificities, the same target.
<3>Microbiology
<4>155
<5>1111-1121
<6>2009
<7>Methylation of a base in a specific DNA sequence protects the DNA from nucleolytic cleavage by
restriction enzymes recognizing the same
sequence. The Mboll restriction-modification (R-M) system of Moraxella
bovis ATCC 10900 consists of a restriction endonuclease gene and two
methyltransferase genes. The enzymes encoded by this system recognize
an asymmetrical sequence 5'-GAAGA-3'/3'-CTTCT-5'. M1.Mboll modifies the
last adenine in the recognition sequence 5'-GAAGA-3' to
N-6-methyladenine. A second methylase, M2.Mboll, was cloned and
purified to electrophoretic homogeneity using a four-step
chromatographic procedure. It was demonstrated that M2.Mboll modifies
the internal cytosine in the recognition sequence 3'-CTTCT-5', yielding
N-4-methylcytosine, and moreover is able to methylate single-stranded
DNA. The protein exists in solution as a monomer of molecular mass 30
000 +/- 1000 Da under denaturing conditions, Divalent cations (Ca2+,
Mg2+, Mn2+, and Zn2+) inhibit M2.Mboll methylation activity. It was
found that the isomethylomer M2.Ncul from Neisseria cuniculi ATCC 14688
behaves in the same manner. Functional analysis showed that the
complete Mboll R-M system, consisting of two methyltransferases genes
and the mbollR gene, is the most stable and the least harmful to
bacterial cells.

<>

<1>Furness, L.M., Buchbinder, J.L.
<2>Genes expressed in C3A liver cell cultures treated with steroids.
<3>US Patent Office
<4>US 6673549 B
<5>
<6>2004
<7>The present invention relates to a combination comprising a plurality of cDNAs which are
differentially expressed in human C3A liver cell cultures treated with steroids or synthetic
steroid analogues and which may be used entirely or in part to detect metabolic and
toxicological responses to treatment with steroids and steroid antagonists, to diagnose, to
stage, to treat, or to monitor the treatment of a subject with an steroid responsive disorder.

<>

<1>Furuta, Y., Abe, K., Kobayashi, I.
<2>Genome comparison and context analysis reveals putative mobile forms of restriction-modification systems and related rearrangements.
<3>Nucleic Acids Res.
<4>38
<5>2428-2443
<6>2010
<7>The mobility of restriction-modification (RM) gene complexes and their association with genome
rearrangements is a subject of active
investigation. Here we conducted systematic genome comparisons and genome
context analysis on fully sequenced prokaryotic genomes to detect
RM-linked genome rearrangements. RM genes were frequently found to be
linked to mobility-related genes such as integrase and transposase
homologs. They were flanked by direct and inverted repeats at a
significantly high frequency. Insertion by long target duplication was
observed for I, II, III and IV restriction types. We found several RM
genes flanked by long inverted repeats, some of which had apparently
inserted into a genome with a short target duplication. In some cases,
only a portion of an apparently complete RM system was flanked by inverted
repeats. We also found a unit composed of RM genes and an integrase
homolog that integrated into a tRNA gene. An allelic substitution of a
Type III system with a linked Type I and IV system pair, and allelic
diversity in the putative target recognition domain of Type IIG systems
were observed. This study revealed the possible mobility of all types of
RM systems, and the diversity in their mobility-related organization.

<>

<1>Furuta, Y., Abe, K., Kobayashi, I.
<2>Search for genomic rearrangements related to restriction-modification systems through genome comparison.
<3>Genes Genet. Syst.
<4>84
<5>441
<6>2009
<7>We are proposing that restriction-modification systems are mobile element and the data which
supports the hypothesis is accumulating.  For example, Type II restriction modification
inserted in genome with long target duplication was found by the intra-genomic comparison
analysis of Helicobacter pylori. First, we searched for more genome rearrangements related to
restriction-modification systems by comprehensive bacterial intra-genomic comparison analysis
and found the examples of (i) insertion with long target duplication of another type of
restriction-modification systems, (ii) alleles consisted of different types of
restriction-modification systems and (iii) alleles which have diversity in recognition
domains. Next, we searched for the restriction-modification systems which are flanked by
repeat sequences and found that restriction-modifiction systems are flanked by repeats much
frequently than other genes significantly.  By the genome comparison analysis of those repeats
flanked restriction-modification systems, we found that the transposon like structure of
restriction-modification, which was flanked by long inverted repeats, was inserted with short
direct repeats in the genome.

<>

<1>Furuta, Y., Kawai, M., Uchiyama, I., Kobayashi, I.
<2>Domain Movement within a Gene: A Novel Evolutionary Mechanism for Protein Diversification.
<3>PLoS ONE
<4>6
<5>e18819
<6>2011
<7>A protein function is carried out by a specific domain localized at a specific position. In
the present study, we report that, within a gene, a
specific amino acid sequence can move between a certain position and
another position. This was discovered when the sequences of
restriction-modification systems within the bacterial species Helicobacter
pylori were compared. In the specificity subunit of Type I
restriction-modification systems, DNA sequence recognition is mediated by
target recognition domain 1 (TRD1) and TRD2. To our surprise, several
sequences are shared by TRD1 and TRD2 of genes (alleles) at the same locus
(chromosomal location); these domains appear to have moved between the two
positions. The gene/protein organization can be represented as
x-(TRD1)-y-x-(TRD2)-y, where x and y represent repeat sequences. Movement
probably occurs by recombination at these flanking DNA repeats. In
accordance with this hypothesis, recombination at these repeats also
appears to decrease two TRDs into one TRD or increase these two TRDs to
three TRDs (TRD1-TRD2-TRD2) and to allow TRD movement between genes even
at different loci. Similar movement of domains between TRD1 and TRD2 was
observed for the specificity subunit of a Type IIG restriction enzyme.
Similar movement of domain between TRD1 and TRD2 was observed for Type I
restriction-modification enzyme specificity genes in two more eubacterial
species, Streptococcus pyogenes and Mycoplasma agalactiae. Lateral domain
movements within a protein, which we have designated DOMO (domain
movement), represent novel routes for the diversification of proteins.

<>

<1>Furuta, Y., Kobayashi, I.
<2>Movement of DNA sequence recognition domains between non-orthologous proteins.
<3>Nucleic Acids Res.
<4>40
<5>9218-9232
<6>2012
<7>Comparisons of proteins show that they evolve through the movement of domains. However, in
many cases, the underlying mechanisms remain
unclear. Here, we observed the movements of DNA recognition domains
between non-orthologous proteins within a prokaryote genome.
Restriction-modification (RM) systems, consisting of a
sequence-specific DNA methyltransferase and a restriction enzyme,
contribute to maintenance/evolution of genomes/epigenomes. RM systems
limit horizontal gene transfer but are themselves mobile. We compared
Type III RM systems in Helicobacter pylori genomes and found that
target recognition domain (TRD) sequences are mobile, moving between
different orthologous groups that occupy unique chromosomal locations.
Sequence comparisons suggested that a likely underlying mechanism is
movement through homologous recombination of similar DNA sequences that
encode amino acid sequence motifs that are conserved among Type III DNA
methyltransferases. Consistent with this movement, incongruence was
observed between the phylogenetic trees of TRD regions and other
regions in proteins. Horizontal acquisition of diverse TRD sequences
was suggested by detection of homologs in other Helicobacter species
and distantly related bacterial species. One of these RM systems in H.
pylori was inactivated by insertion of another RM system that likely
transferred from an oral bacterium. TRD movement represents a novel
route for diversification of DNA-interacting proteins.

<>

<1>Furuta, Y., Kobayashi, I.
<2>Restriction-modification systems as mobile epigenetic elements.
<3>Bacterial Integrative Mobile Genetic Elements, Landes Bioscience, Roberts, A.P., Mullany, P., Austin, TX
<4>
<5>
<6>2011
<7>Transfer of mobile genetic elements between prokaryotes is limited by restriction-modification
systems.  Restriction-modification systems consist of a modification enzyme that
epigenetically methylates a specific DNA sequence, and a restriction endonuclease (restriction
enzyme) that cuts DNA lacking this epigenetic mark.  These elements were discovered because
they attack mobile genetic elements.  However, recent studies have revealed that they are
themselves mobile.  In some cases, the mobility of restriction-modification systems is through
symbiosis with other forms of mobile elements.  In other cases, movement is unlinked to other
mobile elements.  The systems may insert into the genome with long and variable target
duplication, or into the intergenic region of an operon.  Insertion of
restriction-modification systems induces other genome rearrangements such as amplification and
inversion.  Even a domain within a protein can be the unit of mobility: some
restriction-modification system subunits that recognize a target DNA sequence contain mobile
amino acid sequences that can apparently move between different domains of a protein through
recombination of DNA sequences encoding them.  This mobility extends the biological
significance of restriction-modification systems beyond defense: the systems define, and
sometimes even force, epigenetic order on a genome.  The multilevel conflicts involving these
mobile epigenetic elements may drive prokaryotic evolution.

<>

<1>Furuta, Y., Namba, H., Shibata, T., Nishiyama, T., Shigenobu, S., Suzuki, Y., Sugano, S., Hasebe, M., Kobayashi, I.
<2>Methylome diversification through changes in the sequence specificity of DNA methyltransferases.
<3>Genes Genet. Syst.
<4>88
<5>347
<6>2013
<7>Helicobacter pylori, a human gastric pathogen, have a large number of DNA methyltransferase
genes with each strain carrying a unique repertoire.  Previous genome comparison works
suggested that these methyltransferases often change DNA sequence specificity through movement
of amino-acid sequences in the target recognition domains between genes and within a gene
(Domain Movement).  By Single-Molecule Real-Time sequencing technology, we detected methylated
DNA sites throughout several closely related genomes.  We successfully deduced DNA sequence
motifs for methylation and assigned each of them to a specific amino-acid sequence group of
target recognition domains in the specificity determinant genes.  Overall, the methylome
turned out to be quite variable among the closely-related strains, although there are
hypermethylated loci in all the strains.  As expected from their effects on gene expression,
knockout of a specificity gene led to changes in the transcriptome.  These results provide
evidence for proposed mechanisms of sequence-specificity changes in the DNA methyltransferases
and lend support to the concept of epigenetics-driven adaptive evolution.

<>

<1>Furuta, Y., Namba-Fukuyo, H., Shibata, T.F., Nishiyama, T., Shigenobu, S., Suzuki, Y., Sugano, S., Hasebe, M., Kobayashi, I.
<2>Methylome Diversification through Changes in DNA Methyltransferase Sequence Specificity.
<3>PLoS Genet.
<4>10
<5>e1004272
<6>2014
<7>Epigenetic modifications such as DNA methylation have large effects on gene expression and
genome maintenance. Helicobacter pylori, a human gastric pathogen,
has a large number of DNA methyltransferase genes, with different strains having
unique repertoires. Previous genome comparisons suggested that these
methyltransferases often change DNA sequence specificity through domain
movement-the movement between and within genes of coding sequences of target
recognition domains. Using single-molecule real-time sequencing technology, which
detects N6-methyladenines and N4-methylcytosines with single-base resolution, we
studied methylated DNA sites throughout the H. pylori genome for several closely
related strains. Overall, the methylome was highly variable among closely related
strains. Hypermethylated regions were found, for example, in rpoB gene for RNA
polymerase. We identified DNA sequence motifs for methylation and then assigned
each of them to a specific homology group of the target recognition domains in
the specificity-determining genes for Type I and other restriction-modification
systems. These results supported proposed mechanisms for sequence-specificity
changes in DNA methyltransferases. Knocking out one of the Type I specificity
genes led to transcriptome changes, which suggested its role in gene expression.
These results are consistent with the concept of evolution driven by DNA
methylation, in which changes in the methylome lead to changes in the
transcriptome and potentially to changes in phenotype, providing targets for
natural or artificial selection.

<>

<1>Futterer, O., Angelov, A., Liesegang, H., Gottschalk, G., Schleper, C., Schepers, B., Dock, C., Antranikian, G., Liebl, W.
<2>Genome sequence of Picrophilus torridus and its implications for life around pH 0.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>9091-9096
<6>2004
<7>The euryarchaea Picrophilus torridus and Picrophilus oshimae are able to grow around pH 0 at
up to 65 C, thus they represent the most thermoacidophilic organisms known. Several features
that may contribute to the thermoacidophilic survival strategy of P. torridus were deduced
from analysis of its 1.55-megabase genome. P. torridus has the smallest genome among
nonparasitic aerobic microorganisms growing on organic substrates and simultaneously the
highest coding density among thermoacidophiles. An exceptionally high ratio of secondary over
ATP-consuming primary transport systems demonstrates that the high proton concentration in the
surrounding medium is extensively used for transport processes. Certain genes that may be
particularly supportive for the extreme lifestyle of P. torridus appear to have been
internalized into the genome of the Picrophilus lineage by horizontal gene transfer from
crenarchaea and bacteria. Finally, it is noteworthy that the thermoacidophiles from
phylogenetically distant branches of the Archaea apparently share an unexpectedly large pool
of genes.

<>

<1>Fuxreiter, M., Osman, R.
<2>Probing the general base catalysis in the first step of BamHI action by computer simulations.
<3>Biochemistry
<4>40
<5>15017-15023
<6>2001
<7>BamHI is a type II restriction endonuclease that catalyzes the scission of the phoshodiester
bond in the GAGTCC cognate sequence in the presence of two divalent metal ions. The first step
of the reaction is the preparation of water for nucleophilic attack by Glu-113, which has been
proposed to abstract the proton from the attacking water molecule. Alternatively, the
3'-phosphate group to the susceptible phosphodiester bond has been suggested to play a role
as the general base. The two hypotheses have been tested by computer simulations using the
semiempirical protein dipoles Langevin dipoles (PDLD/S) method. Deprotonation of water by
Glu-113 has been found to be less favorable by 5.7 kcal/mol than metal-catalyzed deprotonation
with a concomitant proton transfer to bulk solvent. The preparation of the nucleophile by the
3'-phosphate group is less favorable by 12.3 kcal/mol. These results suggest that both the
general base and the substrate-assisted mechanisms in the first step of BamHI action are less
likely than the metal-catalyzed reaction. The metal ions in the active site of BamHI make the
largest contributions to the reduction of the free energy of hydroxide ion formation. On the
basis of these findings we propose that the first step of endonuclease catalysis does not
require a general base; rather, the essential attacking nucleophile in BamHI catalytic action
is stabilized by the metal ions.

<>

<1>Fuxreiter, M., Osman, R., Simon, I.
<2>Computational approaches to restriction endonucleases.
<3>Theochem.
<4>666-7
<5>469-479
<6>2003
<7>Type II restriction endonucleases catalyze phosphodiester bond hydrolysis in bacteria to
protect the host cell from invading phage DNA.  Due to their exquisite sequence selectivity
type II restriction endonucleases serve as excellent model systems for studying protein -
nucleic acid interactions.  Crystal structures of the PD-(D/E)XK superfamily revealed a common
a/b core motif and similar active site.  In contrast, these enzymes show little sequence
similarity and use different strategies to interact with their substrate DNA.  Computational
approaches have been applied to unify the mechanism of restriction endonucleases and
rationalize their diversity.  The first step of type II restriction endonuclease catalysis has
been studied on BamHI by semi-microscopic version of the Protein Dipoles Langevin Dipoles
method.  The substrate-assisted catalysis and the general base mechanism have been concluded
as less likely than the metal-catalyzed reaction.  A general model for catalysis has been
proposed based on the group contributions to the reduction of the activation free energy.
Factors contributing to structural stability of PD-(D/E)XK type II restriction endonucleases
have been analyzed to elucidate evolutionary relationship between these enzymes.  Residues
playing role in catalysis and recognition were highly correlated with those participating in
stabilization centers.  Thus the main functional motifs were concluded to be evolutionary more
conserved than other parts of the structure.  This observation is consistent with the proposal
that these enzymes have developed from a common ancestor with divergent evolution.

<>

<1>Fuxreiter, M., Simon, I.
<2>Protein stability indicates divergent evolution of PD-(D/E)XK type II restriction endonucleases.
<3>Protein Sci.
<4>11
<5>1978-1983
<6>2002
<7>Type II restriction endonucleases recognize 4-8 base-pair-long DNA sequences and catalyze
their cleavage with remarkable specificity.
Crystal structures of the PD-(DE)XK superfamily revealed a common
alpha/beta core motif and similar active site. In contrast, these
enzymes show little sequence similarity and use different strategies to
interact with their substrate DNA. The intriguing question is whether
this enzyme family could have evolved from a common origin. In our
present work, protein structure stability elements were analyzed and
compared in three parts of PD-(DE)XK type II restriction endonucleases:
(1) core motif, (2) active-site residues, and (3) residues playing role
in DNA recognition. High correlation was found between the active-site
residues and those stabilization factors that contribute to preventing
structural decay. DNA recognition sites were also observed to
participate in stabilization centers. It indicates that recognition
motifs and active sites in PD-(DE)XK type II restriction endonucleases
should have been evolutionary more conserved than other parts of the
structure. Based on this observation it is proposed that PD-(DE)XK type
II restriction endonucleases have developed from a common ancestor with
divergent evolution.

<>

<1>Gabbara, S., Bhagwat, A.S.
<2>Interaction of EcoRII endonuclease with DNA substrates containing single recognition sites.
<3>J. Biol. Chem.
<4>267
<5>18623-18630
<6>1992
<7>EcoRII is unusual among type II restriction enzymes in that, while it cleaves substrates such
as pBR322 and bacteriophage lambda that contain several recognition sites for the enzyme
efficiently, substrates such as the genomes of bacteriophages T3 and T7 which contain a small
number of recognition sites are cut poorly by it. Interestingly, pBR322, or a short DNA duplex
containing a single site for the enzyme, can activate the enzyme to cleave resistant
substrates. We show here that at low concentrations, activator short duplexes are themselves
cleaved poorly by the enzyme. Further, the reaction shows substrate cooperativity, and at high
concentration, the duplexes are both activators and good substrates for the enzyme. This
supports the model that the activation of EcoRII involves binding of more than one DNA
molecule and provides a simple system to study the mechanism of activation. Using a gel
mobility shift assay, we show that the enzyme forms sequence-specific, methylation-sensitive
complexes with the duplexes in the absence of activating DNA. Therefore, resistance of the
short duplexes to the enzyme at low concentrations cannot be due to an inability of the enzyme
to bind the duplexes. Interestingly, these complexes are stable in the presence of Mg2+, the
cofactor for the enzyme, and the complexes obtained in the presence of Mg2+ do not contain DNA
that is cleaved by the enzyme. The inefficient step in the action of EcoRII on resistant
substrates must occur subsequent to initial substrate binding and it is this step that the
activating DNA must regulate.

<>

<1>Gabbara, S., Bhagwat, A.S.
<2>The mechanism of inhibition of DNA (cytosine-5-)-methyltransferases by 5-azacytosine is likely to involve methyl transfer to the inhibitor.
<3>Biochem. J.
<4>307
<5>87-92
<6>1995
<7>The mechanism of inhibition of DNA (cytosine-5-)-methyltransferases by the mechanism-based
inhibitor 5-azacytosine has remained unclear, mainly because of the unavailability of a
substrate in which the inhibitor, but not normal cytosine, is present at the target site. We
synthesized an oligonucleotide duplex containing a single target site for the EcoRII
methyltransferase, in which the target base is 5-azacytosine. This substrate formed a stable
covalent complex with EcoRII methyltransferase in the absence and in the presence of the
cofactor S-adenosylmethionine. The complex formed in the presence of the cofactor was
resistant to SDS and moderate heat treatment, and a methyl group was incorporated into the
complex. Enzyme titration and kinetic studies of inhibition suggest that methyl transfer to
the complex occurred only during the first turnover of the reaction. These results suggest
that, when the enzyme binds to 5-azacytosine in the presence of the cofactor, a methyl group
is transferred to the N-5 position of the base, resulting in the inactivation of the enzyme.

<>

<1>Gabbara, S., Sheluho, D., Bhagwat, A.S.
<2>Cytosine methyltransferase from Escherichia coli in which active site cysteine is replaced with serine is partially active.
<3>Biochemistry
<4>34
<5>8914-8923
<6>1995
<7>EcoRII methyltransferase (M.EcoRII) catalyzes the transfer of methyl groups from
S-adenosyl-L-methionine (SAM) to C-5 position of second cytosine in the DNA sequence
5'-CCWGG (W=A or T).  The reaction is initiated by a nucleophilic attack of the C-6 target
cytosine
by a cysteine that is conserved among all cytosine methyltransferases.  We have replaced this
cysteine in M.EcoRII with serine or alanine and purified the proteins to homogeneity.  The
catalytic
efficiency (Kcat/Km) of the mutant enzyme with serine (C186S) for methyl transfer is about
10,000 times less than that of WT but is substantially higher than the efficiency of the C186A
mutant.  We show that the WT enzyme and C186S mutant are proficient in exchange of proton at
C-5 and that this activity is reduced in the mutant to the same extent as the methyl transfer
activity.
The C186S mutant is insensitive to a cysteine-specific inhibitor, and it transfers methyl
groups to
the same position of cytosine as the WT enzyme.  The ability of serine to act as a nucleophile
in the
enzyme reaction suggests that it - and probably the cysteine in the WT enzyme - is activated
by a
nearby base.  Like the WT enzyme, C186S forms stable SDS-resistant complexes with DNA
containing 5-azacytosine; but unlike the WT enzyme, the mutant reacts faster with
5-azacytosine
than with normal cytosine.  Apparently, greater reactivity of 5-azacytosine assists the C186S
mutant in catalysis.

<>

<1>Gabbara, S., Wyszynski, M., Bhagwat, A.S.
<2>A DNA repair process in Escherichia coli corrects U:G and T:G mismatches to C:G at sites of cytosine methylation.
<3>Mol. Gen. Genet.
<4>243
<5>244-248
<6>1994
<7>Escherichia coli contains a base mismatch correction system called VSP repair that is known to
correct T:G mismatches to C:G when they occur in certain sequence contexts. The preferred
sequence context for this process is the site for methylation by the E. coli DNA cytosine
methylase (Dcm). For this reason, VSP repair is thought to counteract potential mutagenic
effects of deamination of 5-methylcytosine to thymine. We have developed a genetic reversion
assay that quantitates the frequency of C to T mutations at Dcm sites and the removal of such
mutations by DNA repair processes. Using this assay, we have studied the repair of U:G
mismatches in DNA to C:G and have found that VSP repair is capable of correcting these
mismatches. Although VSP repair substantially affects the reversion frequency, it may not be
as efficient at correcting U:G mismatches as the uracil DNA glycosylase-mediated repair
process.

<>

<1>Gabbara, S.S.
<2>Molecular mechanism and mutagenic effects of DNA 5-cytosine methyltransferases.
<3>Diss. Abstr.
<4>55
<5>403B-404B
<6>1994
<7>DNA (C-S)cytosine methyltransferases (C5 methylases) catalyze the transfer of methyl group to
position 5 of cytosine in DNA. This type of modification occurs in most organisms, from
bacteria to humans. In prokaryotes C5 methylases function primarily in protecting the cell
from viral invasions. In eukaryotes C5 methylases play important roles in the regulation of
gene expression. All known cytosine methylases, including the mouse and the human methylases,
share a strong sequence conservation, suggesting a common reaction mechanism. I used the
Escherichia coli EcoRII methylase in this study as a model enzyme to understand the mechanism
and function of all cytosine methylases. The reaction catalyzed by the methylase is proposed
to occur in a Michael fashion. My data using cytosine analogs support this mechanism.
Furthermore, I show that a cysteine conserved among all cytosine methylases is required for
catalysis. This role is likely to be in the nucleophilic attack at carbon 6 of cytosine, a
first step in the reaction. In support of this role substitution of the conserved cysteine by
serine or alanine reduces kcat of methyl transferase activity by 4 and 6 orders of magnitude,
respectively, but does not appreciably affect Km. While studying the properties of the mutants
with serine and alanine changes, I found that the hydroxyl group of the serine, replacing the
conserved cysteine, can covalently link to DNA. Sites of cytosine methylation are hot-spots
for cytosine (C) to thymine (T) mutations. In humans such mutations are found to be the cause
of genetic diseases and cancer. Using a genetic system based on the reversion of a mutation in
a kanamycin-resistance gene, I show that EcoRII methylase can cause cytosine to uracil (U)
deaminations at a site of cytosine methylation, and that the uracil formed can propagate in
vivo to cause C:G to T:A transition mutations. Thus the enzyme-mediated C to U deamination is
one pathway by which cytosine methylases cause C to T mutations. The contribution of the C to
U to T pathway in C to T mutagenesis was assessed by analyzing the barriers to its occurrence.

<>

<1>Gabrielsen, C., Brede, D.A., Hernandez, P.E., Nes, I.F., Diep, D.B.
<2>Genome Sequence of the Bacteriocin-Producing Strain Lactococcus garvieae DCC43.
<3>J. Bacteriol.
<4>194
<5>6976-6977
<6>2012
<7>This work describes the draft genome sequence of Lactococcus garvieae DCC43. The  2.2-Mb draft
genome contains 2,227 predicted protein-coding genes, among which is
a region encoding the bacteriocin garvicin ML. No antibiotic resistance genes or
capsule-related virulence genes were identified. Two plasmid replication regions
indicate that this strain likely contains plasmids. Comparative genomics suggests
that this strain displays a high degree of sequence variation from the previously
sequenced L. garvieae strains.

<>

<1>Gabs, S., Josephsen, J.
<2>Improvement of phage defence in Lactococcus lactis by introduction of the plasmid encoded restriction and modification system LlaAI.
<3>Lett. Appl. Microbiol.
<4>36
<5>332-336
<6>2003
<7>AIMS: To study the ability of the plasmid-encoded restriction and modification (R/M) system
LlaAI to function as a bacteriophage resistance
mechanism in Lactococcus lactis during milk fermentations. METHODS AND
RESULTS: Plasmid pAIcat4, carrying the R/M system LlaAI and a
chloramphenicol resistance cassette, was introduced into the plasmid-free
strain L. lactis MG1614 and the industrial strain L. lactis 964. By
measuring changes in conductivity the influence of different phage on the
growth was determined. CONCLUSIONS: The plasmid-encoded R/M system LlaAI
significantly improves the bacteriophage resistance of L. lactis during
milk fermentations. SIGNIFICANCE AND IMPACT OF THE STUDY: It is essential
to determine the potential of a phage defence mechanism in L. lactis
starter culture strains during growth in milk before steps are taken to
improve starter cultures. This study shows that LlaAI is useful for
improvement of starter cultures.

<>

<1>Gabsalilow, L., Schierling, B., Friedhoff, P., Pingoud, A., Wende, W.
<2>Site- and strand-specific nicking of DNA by fusion proteins derived from MutH and I-SceI or TALE repeats.
<3>Nucleic Acids Res.
<4>41
<5>e83
<6>2013
<7>Targeted genome engineering requires nucleases that introduce a highly specific double-strand
break in the genome that is either processed by homology-directed repair in the presence of a
homologous repair template or by non-homologous end-joining (NHEJ) that usually results in
insertions or deletions. The error-prone NHEJ can be efficiently suppressed by 'nickases'
that produce a single-strand break rather than a double-strand break. Highly specific nickases
have been produced by engineering of homing endonucleases and more recently by modifying zinc
finger nucleases (ZFNs) composed of a zinc finger array and the catalytic domain of the
restriction endonuclease FokI. These ZF-nickases work as heterodimers in which one subunit has
a catalytically inactive FokI domain. We present two different approaches to engineer highly
specific nickases; both rely on the sequence-specific nicking activity of the DNA mismatch
repair endonuclease MutH which we fused to a DNA-binding module, either a catalytically
inactive variant of the homing endonuclease I-SceI or the DNA-binding domain of the TALE
protein AvrBs4. The fusion proteins nick strand specifically a bipartite recognition sequence
consisting of the MutH and the I-SceI or TALE recognition sequences, respectively, with a more
than 1000-fold preference over a stand-alone MutH site. TALE-MutH is a programmable nickase.

<>

<1>Gachechiladze, K.K., Balardshishvili, N.S., Adamia, R.S., Chanishvili, T.G., Kruger, D.H.
<2>Host-controlled modification and restriction as a criterion of evaluating the therapeutical potential of Pseudomonas phage.
<3>J. Basic Microbiol.
<4>31
<5>101-106
<6>1991
<7>The recently isolated phages Phi ST3 and Phi ST1 were compared as to their
lysis behaviour in about 100 different P. aeruginosa strains.  The growth of
Phi ST3 varies greatly in different host strains.  We demonstrated one case of
non-classical, host-dependent modification and restriction.  Here the
capability to adsorb, and consequently to reproduce in a given host strain
differs, depending on which modification the phage acquired in its former host.
The DNA-containing phage Phi ST1 displays stable lysis properties in the
majority of the host strains.  This make Phi ST1 a candidate for therapeutic
phage preparations.  One of the reasons for stable lysis properties is the
apparent selection against recognition sites of restriction enzymes in its
genome.

<>

<1>Gado, I., Laszlo, V.G., Negy, B., Milch, H., Drin, I., Awad-Masalmeh, M., Horvath, J.
<2>Phage restriction and the presence of small plasmids in Salmonella enteritidis.
<3>Zentralbl. Bakteriol.
<4>287
<5>509-519
<6>1998
<7>Between 1990-1994, a total of 16,505 S. enteritidis strains of human, animal and food origin
were phage-typed, using the Hungarian scheme and the changes of incidence of the dominant
phage types were monitored.  The incidence of PT1 (corresponding to Ward's PT1 was very high
between 1990 and 1992 (67.9071.0% of the total S. enteritidis isolates), later, it decreased.
The prevalence of PT6 (corresponding to Ward's PT4) was rare until 1992, then it gradually
increased.  The phage type and plasmid content of 78 Salmonella enteritidis strains were
determined.  Small plasmids were present in 59% of the isolates, together with a
serotype-specific (38 MDa) plasmid.  A correlation was found between the presence of the small
plasmid and phage restriction to two phages used for subdividing the Hungarian phage types 1
(PT1) and 6 (PT6) of S. enteritidis (corresponding to PT1 and PT4 in Ward's typing scheme,
respectively).

<>

<1>Gaechter, T., Wunderlin, C., Schmidheini, T., Solioz, M.
<2>Genome Sequence of Enterococcus hirae (Streptococcus faecalis) ATCC 9790, a Model Organism for the Study of Ion Transport, Bioenergetics, and Copper Homeostasis.
<3>J. Bacteriol.
<4>194
<5>5126-5127
<6>2012
<7>Enterococcus hirae ATCC 9790 is a Gram-positive lactic acid bacterium that has been used in
basic research for over 4 decades. Here we report the sequence and
annotation of the 2.8-Mb genome of E. hirae and its endemic 29-kb plasmid
pTG9790.

<>

<1>Gagnevin, L., Bolot, S., Gordon, J.L., Pruvost, O., Verniere, C., Robene, I., Arlat, M., Noel, L.D., Carrere, S., Jacques, M.A., Koebnik, R.
<2>Draft Genome Sequence of Xanthomonas axonopodis pv. allii Strain CFBP 6369.
<3>Genome Announcements
<4>2
<5>e00727-14
<6>2014
<7>We report here the draft genome sequence of Xanthomonas axonopodis pv. allii strain CFBP 6369,
the causal agent of bacterial blight of onion. The draft genome
has a size of 5,425,942 bp and a G+C content of 64.4%.

<>

<1>Gai, Z., Wang, X., Liu, X., Tai, C., Tang, H., He, X., Wu, G., Deng, Z., Xu, P.
<2>The genes coding for the conversion of carbazole to catechol are flanked by IS6100 elements in Sphingomonas sp. strain XLDN2-5.
<3>PLoS ONE
<4>5
<5>E10018
<6>2010
<7>BACKGROUND: Carbazole is a recalcitrant compound with a dioxin-like
structure and possesses mutagenic and toxic activities. Bacteria respond
to a xenobiotic by recruiting exogenous genes to establish a pathway to
degrade the xenobiotic, which is necessary for their adaptation and
survival. Usually, this process is mediated by mobile genetic elements
such as plasmids, transposons, and insertion sequences. FINDINGS: The
genes encoding the enzymes responsible for the degradation of carbazole to
catechol via anthranilate were cloned, sequenced, and characterized from a
carbazole-degrading Sphingomonas sp. strain XLDN2-5. The car gene cluster
(carRAaBaBbCAc) and fdr gene were accompanied on both sides by two copies
of IS6100 elements, and organized as
IS6100::ISSsp1-ORF1-carRAaBaBbCAc-ORF8-IS6100-fdr-IS6100. Carbazole was
converted by carbazole 1,9a-dioxygenase (CARDO, CarAaAcFdr), meta-cleavage
enzyme (CarBaBb), and hydrolase (CarC) to anthranilate and
2-hydroxypenta-2,4-dienoate. The fdr gene encoded a novel ferredoxin
reductase whose absence resulted in lower transformation activity of
carbazole by CarAa and CarAc. The ant gene cluster (antRAcAdAbAa) which
was involved in the conversion of anthranilate to catechol was also
sandwiched between two IS6100 elements as IS6100-antRAcAdAbAa-IS6100.
Anthranilate 1,2-dioxygenase (ANTDO) was composed of a reductase (AntAa),
a ferredoxin (AntAb), and a two-subunit terminal oxygenase (AntAcAd).
Reverse transcription-PCR results suggested that carAaBaBbCAc gene
cluster, fdr, and antRAcAdAbAa gene cluster were induced when strain
XLDN2-5 was exposed to carbazole. Expression of both CARDO and ANTDO in
Escherichia coli required the presence of the natural reductases for full
enzymatic activity. CONCLUSIONS/SIGNIFICANCE: We predict that IS6100 might
play an important role in the establishment of carbazole-degrading
pathway, which endows the host to adapt to novel compounds in the
environment. The organization of the car and ant genes in strain XLDN2-5
was unique, which showed strong evolutionary trail of gene recruitment
mediated by IS6100 and presented a remarkable example of rearrangements
and pathway establishments.

<>

<1>Gai, Z., Wang, X., Tang, H., Tai, C., Tao, F., Wu, G., Xu, P.
<2>Genome Sequence of Sphingobium yanoikuyae XLDN2-5, an Efficient Carbazole-Degrading Strain.
<3>J. Bacteriol.
<4>193
<5>6404-6405
<6>2011
<7>Sphingobium yanoikuyae XLDN2-5 is an efficient carbazole-degrading strain. Carbazole-degrading
genes are accompanied on both sides by two copies of
IS6100 elements. Here, we describe the draft genome sequence of strain
XLDN2-5, which may provide important clues as to how it recruited
exogenous genes to establish pathways to degrade the xenobiotics.

<>

<1>Gai, Z., Wang, X., Zhang, X., Su, F., Wang, X., Tang, H., Tai, C., Tao, F., Ma, C., Xu, P.
<2>Genome Sequence of Sphingomonas elodea ATCC 31461, a Highly Productive Industrial Strain of Gellan Gum.
<3>J. Bacteriol.
<4>193
<5>7015-7016
<6>2011
<7>The commercial gelling agent gellan gum is a heteropolysaccharide produced by Sphingomonas
elodea ATCC 31461. However, the genes involved in the
biosynthesis, regulation, and modification of gellan gum have not been
fully characterized. Here we describe the draft genome sequence of stain
ATCC 31461 and major findings from its annotation.

<>

<1>Gai, Z., Zhang, Z., Wang, X., Tao, F., Tang, H., Xu, P.
<2>Genome Sequence of Pseudomonas aeruginosa DQ8, an Efficient Degrader of n-Alkanes and Polycyclic Aromatic Hydrocarbons.
<3>J. Bacteriol.
<4>194
<5>6304-6305
<6>2012
<7>Pseudomonas aeruginosa DQ8, which was isolated from the crude oil polluted soil in the Daqing
oilfield of China, can efficiently degrade diesel, crude oil,
n-alkanes, and polycyclic aromatic hydrocarbons (PAHs). Here, we present a 6.8-Mb
assembly of its genome sequence. We have annotated 23 coding sequences (CDSs)
responsible for catabolism of n-alkanes and PAHs.

<>

<1>Gaido, M.L.
<2>Purification of BsuE methylase to investigate the role of DNA methylation in rat growth hormone gene expression.
<3>Diss. Abstr.
<4>48
<5>2876B
<6>1988
<7>DNA methylation at specific cytosine bases is one mechanism believed to be
involved in determining tissue specific gene expression.  A tissue specific
methylation pattern has been observed for the rat growth hormone (rGH) gene.
In rat pituitary tissue which expresses GH a CGCG sequence 144 basepairs
upstream from the rGH transcription initiation site is unmethylated.  This site
is methylated in rat liver, spleen and kidney which do not express GH.  We have
directly tested the effect of site specific methylation at the CGCG site on rGH
promoter activity.  1.5 kilobasepairs of rGH promoter sequences were inserted
in front of two bacterial indicator genes Neo and CAT.  A CGCG specific
methylase was isolated from Bacillus subtilis strain ISE15 by three column
chromatography steps:  phosphocellulose, heparin-sepharose and DEAE-Sepharose.
Its molecular weight was 41,000 by gel filtration.  It exhibits optimal
activity in mM KCl, 50 mM Tris.HCl, pH 7.3-8.3 and 5 mM 2-mercaptoethanol.
This enzyme was used to methylate the CGCG sequence in GH1 Neo and GH1.CAT.
Methylated and unmethylated fusion genes were transferred into GH3 rat
pituitary tissue culture cells by calcium-phosphate coprecipitation or
electroporation.  Cells transfected with GH1 Neo were harvested after 2 days,
replated at 5x10/5 cells/100 cm dish in selective media (400 ug/ml G418), and
counted after 2 weeks.  Cells were harvested 36 hours after transfection with
GH1.CAT and acetylation of 14C-chloramphenicol measured in whole cell extracts.
BsuE methylation resulted in a 68% decrease in GH1-Neo fusion gene activity
and in a lesser 44% decrease in the control RSV-Neo fusion gene activity.  The
control methylase, HhaI, which has over twice as many sites on the plasmid,
inhibited RS V-Neo and GH1-Neo activity by 81%-83%.  We conclude that extensive
methylation can nonspecifically inhibit fusion gene expression.  BsuE
methylation of GH1-CAT inhibited fusion gene expression by 49% while the
control HhaI methylase had no inhibitory effect on GH1-CAT activity in
transient expression assays.  In addition, neither BsuE or HhaI methylation had
any inhibitory effect on RSV-CAT activity.  Thus, we conclude that methylation
of the rGH promoter at CGCG sequences does specifically inhibit rGH promoter
activity.

<>

<1>Gaido, M.L., Prostko, C.R., Strobl, J.S.
<2>Isolation and characterization of BsuE methyltransferase, a CGCG specific DNA methyltransferase from Bacillus subtilis.
<3>J. Biol. Chem.
<4>263
<5>4832-4836
<6>1988
<7>The DNA methyltransferase M.BsuEI that recognizes the sequence 5'-CGCG-3' has
been isolated from Bacillus subtilis strain ISE15.  A 1600-fold purification of
M.BsuEI was achieved by column chromatography on phosphocellulose,
heparin-Sepharose, and DEAE-Sepharose.  DNA methyltransferase activity was
monitored in the column eluants radiochemically by the transfer of tritiated
methyl groups from radiolabeled S-adenosylmethionine to poly(dGdC)-poly(dGdC)
DNA, a sensitive and specific substrate for M.BsuEI activity.  The DNA sequence
specificity of this methyltransferase activity was confirmed enzymatically by
demonstrating that M.BsuEI-methylated DNA was selectively protected from
cleavage by the restriction enzyme isoschizomers, ThaI and FnuDII.  Purified
M.BsuEI has an apparent molecular size of 41,000-43,000 as determined by gel
filtration and migrates as a 41-kDa protein in a sodium dodecyl
sulfate-poly-acrylamide gel.  DNA methylation by M.BsuEI is dependent upon the
presence of S-adenosylmethionine and 2-mercaptoethanol.  M.BsuEI
methyltransferase activity is optimal at 37C in the presence of 50 mM Tris-HCl,
pH 7.8, 25 mM KCl, 6 micromolar S-adenosylmethionine, 5 mM 2-mercaptoethanol,
and 10 mM EDTA.  M.BsuEI methylates the external cytidine in its recognition
sequence in both linear and supercoiled DNA.  A unique property of M.BsuEI is
its ability to methylate 5'-CGCG-3' in Z-DNA.

<>

<1>Gaido, M.L., Strobl, J.S.
<2>Methylation sensitivity of the restriction enzymes FnuDII and AccII.
<3>Arch. Microbiol.
<4>46
<5>338-340
<6>1987
<7>The restriction enzymes FnuDII and AccII are isoschizomers of the DNA sequence
5'-CGCG-3'.  We have determined that 5-methylcytidine at either cytidine
position in this recognition sequence inhibits DNA cleavage by FnuDII and
AccII.  A third isoschizomer, ThaI was previously shown to exhibit an identical
methylation sensitivity.  It is remarkable that 3 restriction enzymes derived
from diverse microbiological sources exhibit this identical methylation
sensitivity.

<>

<1>Gaigalas, M., Maneliene, Z., Kazlauskiene, R., Petrusyte, M., Janulaitis, A.
<2>PfoI, a unique type II restriction endonuclease that recognizes the sequence 5'-T/CCNGGA-3'.
<3>Nucleic Acids Res.
<4>30
<5>e98
<6>2002
<7>A new type II restriction endonuclease designated PfoI has been partially purified from
Pseudomonas fluorescens biovar 126.  PfoI recognizes the interrupted hexanucleotide
palindromic sequence 5'-T/CCNGGA-3' and cleaves DNA to produce protruding pentanucleotide
5'-ends.

<>

<1>Gal, S., Monteith, N., Shkalim, S., Huang, H., Head, T.
<2>Methylation of dna may be useful as a computational tool: Experimental evidence.
<3>Current Developments in Mathematical Biology, , Mahdavi, K., Culshaw, R., Boucher, J., 
<4>38
<5>1-14
<6>2007
<7>Previously we have explained the abstract concept we call 'aqueous computing' and
illustrated it with concrete wet lab results. Here, we
explore the use of methylase enzymes to 'write' on double-stranded DNA
molecules at sites where restriction enzymes will cut if, and only if,
the sites have not previously been methylated. A site represents the
bit zero (False, F) if the site has been methylated and the bit one
(True, T) if it has not been methylated. 'Reading' is done by
attempting a cut at each of the sites. We found 8 commercially
available methylases and 8 corresponding restriction enzymes that would
not cut after the action of one of the methylases. We were able to
confirm that methylation by each of these 8 enzymes individually
blocked cleavage only by I he restriction enzyme associated with that
site and not any other enzyme. We I hen used these enzymes to approach
a 3-variable, 4-clause satisfiability (SAT) problem using either
plasmid DNA (pBluescript) or PCR product made from the region
containing the restriction enzyme sites on the plasmid. Pairs of
methylases were defined to represent each of the states of the
operators p, q and r, one methylase for p and another for p', etc. We
methylated the DNA in parallel at the two sites so either the p site
was methylated (making p false) or the P' site was methylated (making
p' false). We did that for the other two variables as well to create a
set of logically consistent DNA fragments. Then we applied the 4
clauses using restriction enzymes to cut DNA fragments that did riot
satisfy them. At the end, we found evidence for intact DNA indicating
an answer satisfying all of the clauses. To confirm the state of each
of the Boolean operators, we used cleavage by the appropriate
restriction enzyme. We found in the computation with both the plasmid
and the PCR product, one site pair to show false in both sites; q and
q', for instance. This should not be possible. We suspected incomplete
cutting during the clauses by one of these restriction Enzymes,
specifically BssHII In summary, we did successfully show the usefulness
of DNA methylation in a scheme to do a mathematical computation. Thus,
we have added to our arsenal of potential methods of performing DNA
computing in the aqueous style.

<>

<1>Galagan, J.E. et al.
<2>The genome of Methanosarcina acetivorans reveals extensive metabolic and physiological diversity.
<3>Genome Res.
<4>12
<5>532-542
<6>2002
<7>Methanogenesis, the biological production of methane, plays a pivotal role in the global
carbon cycle and contributes significantly to global warming. The majority of methane in
nature is derived from acetate. Here we report the complete genome sequence of an
acetate-utilizing methanogen, Methanosarcina acetivorans C2A. Methanosarcineae are the most
metabolically diverse methanogens, thrive in a broad range of environments, and are unique
among the Archaea in forming complex multicellular structures. This diversity is reflected in
the genome of M. acetivorans. At 5,751,492 base pairs it is by far the largest known archaeal
genome. The 4524 open reading frames code for a strikingly wide and unanticipated variety of
metabolic and cellular capabilities. The presence of novel methyltransferases indicates the
likelihood of undiscovered natural energy sources for methanogenesis, whereas the presence of
single-subunit carbon monoxide dehydrogenases raises the possibility of nonmethanogenic
growth. Although motility has not been observed in any Methanosarcineae, a flagellin gene
cluster and two complete chemotaxis gene clusters were identified. The availability of genetic
methods, coupled with its physiological and metabolic diversity, makes M. acetivorans a
powerful model organism for the study of archaeal biology.

<>

<1>Galagan, J.E. et al.
<2>The genome sequence of the filamentous fungus Neurospora crassa.
<3>Nature
<4>422
<5>859-868
<6>2003
<7>Neurospora crassa is a central organism in the history of twentieth-century genetics,
biochemistry and molecular biology. Here, we
report a high-quality draft sequence of the N. crassa genome. The
approximately 40-megabase genome encodes about 10,000 protein-coding
genes--more than twice as many as in the fission yeast Schizosaccharomyces
pombe and only about 25% fewer than in the fruitfly Drosophila
melanogaster. Analysis of the gene set yields insights into unexpected
aspects of Neurospora biology including the identification of genes
potentially associated with red light photobiology, genes implicated in
secondary metabolism, and important differences in Ca2+ signalling as
compared with plants and animals. Neurospora possesses the widest array of
genome defence mechanisms known for any eukaryotic organism, including a
process unique to fungi called repeat-induced point mutation (RIP). Genome
analysis suggests that RIP has had a profound impact on genome evolution,
greatly slowing the creation of new genes through genomic duplication and
resulting in a genome with an unusually low proportion of closely related
genes.

<>

<1>Galardini, M. et al.
<2>Permanent draft genome sequences of the symbiotic nitrogen fixing Ensifer meliloti strains BO21CC and AK58.
<3>Standards in Genomic Sciences
<4>9
<5>325-333
<6>2013
<7>Ensifer (syn. Sinorhizobium) meliloti is an important symbiotic bacterial species that fixes
nitrogen. Strains BO21CC and AK58 were previously investigated for
their substrate utilization and their plant-growth promoting abilities showing
interesting features. Here, we describe the complete genome sequence and
annotation of these strains. BO21CC and AK58 genomes are 6,985,065 and 6,974,333
bp long with 6,746 and 6,992 genes predicted, respectively.

<>

<1>Galarza, M., Tarazona, D., Borda, V., Agapito, J.C., Guio, H.
<2>Evidence of Clonal Expansion in the Genome of a Multidrug-Resistant Mycobacterium tuberculosis Clinical Isolate from Peru.
<3>Genome Announcements
<4>2
<5>e00089-14
<6>2014
<7>We report the genome sequence of Mycobacterium tuberculosis INS-MDR from Peru, a
multidrug-resistant tuberculosis (MDR-TB) and Latin American-Mediterranean (LAM)
lineage strain. Our analysis showed mutations related to drug resistance in the
rpoB (D516V), katG (S315T), kasA (G269S), and pncA (Q10R) genes. Our evidence
suggests that INS-MDR may be a clonal expansion related to the African strain KZN
1435.

<>

<1>Galburt, E.A., Chadsey, M.S., Jurica, M.S., Chevalier, B.S., Erho, D., Tang, W., Monnat, R.J. Jr., Stoddard, B.L.
<2>Conformational changes and cleavage by the homing endonuclease I-PpoI: A critical role for a leucine residue in the active site.
<3>J. Mol. Biol.
<4>300
<5>877-887
<6>2000
<7>The homing endonuclease I-PpoI severely bends its DNA target, resulting in significant
deformations of the minor and major groove near the scissile phosphate groups. To study the
role of conformational changes within the protein catalyst and the DNA substrate, we have
determined the structure of the enzyme in the absence of bound DNA, performed gel retardation
analyses of DNA binding and bending, and have mutagenized a leucine residue that contacts an
adenine nucleotide at the site of cleavage. The structure of the L116A/DNA complex has been
determined and the effects of the mutation on affinity and catalysis have been measured. The
wild-type protein displays a rigid-body rotation of its individual subunits upon DNA binding.
Homing site DNA is not detectably bent in the absence of protein, but is sharply bent in both
the wild-type and L116A complexes. These results indicate that binding involves a large
distortion of the DNA and a smaller change in protein conformation. Leucine 116 is critical
for binding and catalysis: it appears to be important for forming a well-ordered protein-DNA
complex at the cleavage site, for maximal deformation of the DNA, and for desolvation of the
nucleotide bases that are partially unstacked in the enzyme complex.

<>

<1>Galburt, E.A., Chevalier, B., Tang, W., Jurica, M.S., Flick, K.E., Monnat, R.J. Jr., Stoddard, B.L.
<2>A novel endonuclease mechanism directly visualized for I-PpoI.
<3>Nat. Struct. Biol.
<4>6
<5>1096-1099
<6>1999
<7>A novel mechanism of DNA endonucleolytic cleavage has been visualized for the homing
endonuclease I-PpoI by trapping the uncleaved enzyme-substrate complex and comparing it to the
previously visualized product complex. This enzyme employs a unique single metal mechanism. A
magnesium ion is coordinated by an asparagine residue and two DNA oxygen atoms and stabilizes
the phosphoanion transition state and the 3'oxygen leaving group. A hydrolytic water molecule
is activated by a histidine residue for an in-line attack on the scissile phosphate. A
strained enzyme-substrate-metal complex is formed before cleavage, then relaxed during the
reaction.

<>

<1>Galburt, E.A., Jurica, M.S.
<2>His-Cys box homing endonuclease.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Derbyshire, V., Stoddard, B.L., Wood, D.W., Berlin Heidelberg
<4>16
<5>85-102
<6>2005
<7>Homing endonucleases are often grouped into four families based on distinct sequence motifs.
One of these families is known as the His-Cys box homing endonucleases and contains two
clusters of conserved histidine and cysteine residues over a central 100 amino acid region.
At last count, 23 members of this family had been identified.  The open reading frames of
these proteins are contained within mobile group I introns found in nuclear rDNA genes of
several protists.  The nuclear location of these introns and ORFs is currently unique among
the homing endonuclease families and poses an intriguing puzzle regarding their expression
from non-coding rRNA transcripts.  The best-studied member of the His-Cys box homing
endonucleases is I-PpoI from the myxomycete Physarum polycephalum.  Following an introduction
to all of the known members of the His-Cys box endonuclease family, much of the following
chapter will outline the extensive characterization of  I-PpoI structure and function.
Although our understanding of how I-PpoI is expressed in cells is still not fully complete,
the means by which I-PpoI specifically recognizes a single cleavage site in the host genome to
mediate homing of its host intron is widely accepted.  Details of DNA recognition and the
catalytic mechanism of nucleolytic cleavage have been ascertained from both in vivo and in
vitro activity assays as well as from extensive X-ray crystallographic structural analyses of
the enzyme bound to its DNA substrate.

<>

<1>Galburt, E.A., Stoddard, B.L.
<2>Restriction endonucleases: one of these things is not like the others.
<3>Nat. Struct. Biol.
<4>7
<5>89-91
<6>2000
<7>The crystal structure of the restriction endonuclease BglII in complex with its DNA target
site has been determined. The DNA binding mode and chemistry of catalysis are observed to
differ from BamHI which cleaves a similar target site. These observations indicate that more
divergence has occurred within this family of proteins than originally thought.

<>

<1>Galburt, E.A., Stoddard, B.L.
<2>Catalytic mechanisms of restriction and homing endonucleases.
<3>Biochemistry
<4>41
<5>13851-13860
<6>2002
<7>The catalytic mechanisms of type II restriction endonucleases and homing endonucleases are
discussed and compared. Brief reviews of the
chemistry of phosphoryl transfers and canonical one-metal and two-metal
endonucleolytic mechanisms are provided along with possible future
directions in the study of endonuclease active sites. The discussion of
type II restriction endonucleases is comprised of a description of the
general architecture of the canonical active site structural motif
followed by more in-depth examples of one- and two-metal mechanisms.
The homing endonuclease section is comprised of four sections
describing what is known regarding the cleavage mechanisms of the four
group I intron homing endonuclease families: LAGLIDADG, His-Cys box,
H-N-H, and GIY-YIG.

<>

<1>Galibert, F. et al.
<2>The composite genome of the legume symbiont Sinorhizobium meliloti.
<3>Science
<4>293
<5>668-672
<6>2001
<7>The scarcity of usable nitrogen frequently limits plant growth. A tight metabolic association
with rhizobial bacteria allows legumes to obtain nitrogen compounds by bacterial reduction of
dinitrogen (N2) to ammonium (NH4+). We present here the annotated DNA sequence of the
alpha-proteobacterium Sinorhizobium meliloti, the symbiont of alfalfa. The tripartite
6.7-megabase (Mb) genome comprises a 3.65-Mb chromosome, and 1.35-Mb pSymA and 1.68-Mb pSymB
megaplasmids. Genome sequence analysis indicates that all three elements contribute, in
varying degrees, to symbiosis and reveals how this genome may have emerged during evolution.
The genome sequence will be useful in understanding the dynamics of interkingdom associations
and of life in soil environments.

<>

<1>Gallagher, L.A., McKevitt, M., Ramage, E.R., Manoil, C.
<2>Genetic dissection of the Francisella novicida restriction barrier.
<3>J. Bacteriol.
<4>190
<5>7830-7837
<6>2008
<7>Francisella tularensis is the causative agent of tularemia and is a category A select agent.
Francisella novicida, considered by some to be one of four subspecies of F. tularensis, is
used as a model in pathogenesis studies because it causes a disease similar to tularemia in
rodents but is not harmful to humans. F. novicida exhibits a strong restriction barrier which
reduces the transformation frequency of foreign DNA up to 10(6)-fold. To identify the genetic
basis of this barrier, we carried out a mutational analysis of restriction genes identified in
the F. novicida genome. Strains carrying combinations of insertion mutations in eight
candidate loci were created and assayed for reduced restriction of unmodified plasmid DNA
introduced by transformation. Restriction was reduced by mutations in four genes,
corresponding to two type I, one type II, and one type III restriction system. Restriction was
almost fully eliminated in a strain in which all four genes were inactive. The strongest
contributor to the restriction barrier, the type II gene, encodes an enzyme which specifically
cleaves Dam-methylated DNA. Genome comparisons show that most restriction genes in the F.
tularensis subspecies are pseudogenes, explaining the unusually strong restriction barrier in
F. novicida and suggesting that restriction was lost during evolution of the human pathogenic
subspecies. As part of this study, procedures were developed to introduce unmodified plasmid
DNA into F. novicida efficiently, to generate defined multiple mutants, and to produce
chromosomal deletions of multiple adjacent genes.

<>

<1>Gallagher, M.L., Burke, W.F.
<2>Sequence-specific endonuclease from the transformable cyanobacterium Anacystis nidulans R2.
<3>FEMS Microbiol. Lett.
<4>26
<5>317-321
<6>1985
<7>Extracts of the transformable cyanobacterial strain Anacystis nidulans R2 were
analyzed for the presence of restriction endonuclease.  One enzyme, AniI, was
found and determined to be sequence-specific on the basis of its ability to
cleave several Bacillus plasmids at a limited number of sites.  The activity of
this enzyme is significantly reduced in extracts prepared from cell cultures
grown at 38C.

<>

<1>Gallagher, M.L., Burke, W.F.
<2>Unique cyanobacterial recognition sequence for the restriction endonuclease AniI.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>87
<5>224
<6>1987
<7>In previous studies, we have shown that the transformable cyanobacterium
Anacystis nidulans R2 possesses a sequence specific endonuclease AniI.
Restriction digest patterns of site-specific endonucleases of cyanobacterial
origin were compared with AniI generated digests to determine if AniI is an
isoschizomer of any of these enzymes.  We have previously shown that pBR322 is
not cleaved by AniI.  We determined that only eight characterized
cyanobacterial restriction-endonucleases are unable to cleave pBR322.  These
enzymes or isoschizomers of them, if commercially available, were obtained and
used to digest plasmid DNA substrates.  Although several enzymes were not
available, we were able to make a comparison based on published data which
indicated the number of cleavage sities on various substrate DNA's.  None of
the commercially available endonucleases produced restriction patterns similar
to AniI digests.  None of the enzymes which were compared indirectly were found
to have the same number of cleavage sites on PhiX174 substrate DNA.  We
therefore conclude that the AniI recognition sequence is unique with respect to
other cyanobacterial restriction endonucleases.

<>

<1>Galli, D.M., Kerr, M.S., Fair, A.D., Permpanich, P., LeBlanc, D.J.
<2>Parameters associated with cloning in Actinobacillus actinomycetemcomitans.
<3>Plasmid
<4>47
<5>138-147
<6>2002
<7>Characterization of virulence traits in Actinobacillus actinomycetemcomitans requires the
application of recombinant DNA
techniques. To develop appropriate genetic tools it is necessary to
identify suitable host-vector systems. The Current Study assessed
cloning parameters in A. actinomycetemcomitans for two preciously
described vectors. pDMG4 and pMMB67. It was determined that the maximum
size of recombinant molecule that could be transferred to A.
actinomycetemcomitans strain ATCC29522 via electroporation was 33 kb.
The size limit for transformation of the same strain with ligation
mixtures (direct cloning), however, was limited to 23-24 kb. Additional
experiments included electroporation of various A.
actinomycetemcomitans strains with plasmid DNA isolated from
Escherichia coli and different A. actinomycetemcomitans sources.
Differences in transformation efficiencies, suggested the presence of a
restriction modification system for pDMG4 in some strains of A. actinomycetemcomitans. Cloning
of portions of the enterococcal plasmid pJH1 into A. actinomycetemcomitans resulted in the
insertion of the intact vector
into the chromosome.

<>

<1>Gallien, S., Perrodou, E., Carapito, C., Deshayes, C., Reyrat, J.M., Van Dorsselaer, A., Poch, O., Schaeffer, C., Lecompte, O.
<2>Ortho-proteogenomics: multiple proteomes investigation through orthology and a new MS-based protocol.
<3>Genome Res.
<4>19
<5>128-135
<6>2009
<7>The progress in sequencing technologies irrigates biology with an ever-increasing
number of genome sequences. In most cases, the gene repertoire is predicted in
silico and conceptually translated into proteins. As recently highlighted, the
predicted genes exhibit frequent errors, particularly in start codons, with a
serious impact on subsequent biological studies. A new "ortho-proteogenomic"
approach is presented here for the annotation refinement of multiple genomes at
once. It combines comparative genomics with an original proteomic protocol that
allows the characterization of both N-terminal and internal peptides in a single
experiment. This strategy was applied to the Mycobacterium genus with
Mycobacterium smegmatis as the reference, and identified 946 distinct proteins,
including 443 characterized N termini. These experimental data allowed the
correction of 19% of the characterized start codons, the identification of 29
proteins missed during the annotation process, and the curation, thanks to
comparative genomics, of 4328 sequences of 16 other Mycobacterium proteomes.

<>

<1>Gallo, K.A., Shao, K.L., Phillips, L.R., Regan, J.B., Koziolkiewicz, M., Uznanski, B., Stec, W.J., Zon, G.
<2>Alkyl phosphotriester modified oligodeoxyribonucleotides.  V. Synthesis and absolute configuration of Rp and Sp diastereomers of an ethyl phosphotriester (Et) modified EcoRI recognition sequence, d[GGAA(Et)TTCC].
<3>Nucleic Acids Res.
<4>14
<5>7405-7420
<6>1986
<7>Protected deoxynucleoside 3'-O-ethyl-N, N-diisopropylphosphoramidite reagents
were prepared for use in the automated synthesis of ethyl phosphotriester (Et)
modified oligonucleotides.  The title diastereomers were separated by
reversed-phase HPLC, and chirality at phosphorus was assigned by an improved
configurational correlation scheme that was verified by NMR spectroscopic
studies (accompanying paper, Part VI).  This generally applicable correlation
scheme involved (1.) enzymatic digestions of each diastereomer to give the
corresponding diastereomer of d[A(Et)T]; (2.) phosphite triester sulfurization
to obtain diastereomeric 0-ethyl phosphorothioates, d[AS(Et)T], which were
separated by HPLC for (3.) stereoretentive oxidation with H2O2 to give
d[A(Et)T], and (4.) stereoretentive de-ethylation with PhSH-Et3N to give
diastereomeric phosphorothioates, d[AST], whose configurations at phosphorus
had been assigned previously.  Neither the Rp-Rp nor Sp-Sp duplex,
{d[GGAA(Et)TTCC]}2, was cleaved by EcoRI endonuclease under conditions that led
to cleavage of both the unmodified duplex, [d(GGAATTCC)]2, and the mixture of
diastereomeric phosphorothioate-modified duplexes, [d(GGAASTTCC)]2.  Cleavage
of the latter substrates was Sp-selective.

<>

<1>Gamon, K., Hansen, D.
<2>Improving transformation rate for Bacillus licheniformis host - by incubating foreign DNA with host cell modifying enzyme before transfer.
<3>German Patent Office
<4>DE 4005025 A
<5>
<6>1990
<7>The number of transformants produced when transferring foreign DNA into a receptor
microorganism is increased by (1) releasing restriction and modification enzymes from a
receptor (Bacillus licheniformis) cell; (2) inhibiting the restriction enzymes: (3) the
modification system is allowed to act in vitro on the foreign DNA in presence of a c-factor;
then (4) transferring the modified DNA by standard methods into the same receptor organism.
Also new are (A) cell lysates with methylase activity obtained from B. licheniformis DSM
641,3406 and/or 3407 (or their mutants or variants) by growing the cells mechanically or
enzymatically disintegrating them, separation of large components or high mol. wt. materials
by filtration and/or centrifugation, and (B) isolated DNA containing, at least once, the
recognition site 5'-GCNGC-3', which is methylated in vitro by the cell lysate.

<>

<1>Gan, H.M., Chew, T.H., Hudson, A.O., Savka, M.A.
<2>Genome Sequence of Methylobacterium sp. Strain GXF4, a Xylem-Associated Bacterium Isolated from Vitis vinifera L. Grapevine.
<3>J. Bacteriol.
<4>194
<5>5157-5158
<6>2012
<7>Methylobacterium sp. strain GXF4 is an isolate from grapevine. Here we present the sequence,
assembly, and annotation of its genome, which may shed light on its
role as a grapevine xylem inhabitant. To our knowledge, this is the first genome
announcement of a plant xylem-associated strain of the genus Methylobacterium.

<>

<1>Gan, H.M., Chew, T.H., Hudson, A.O., Savka, M.A.
<2>Genome Sequence of Novosphingobium sp. Strain Rr 2-17, a Nopaline Crown Gall-Associated Bacterium Isolated from Vitis vinifera L. Grapevine.
<3>J. Bacteriol.
<4>194
<5>5137-5138
<6>2012
<7>Novosphingobium sp. strain Rr 2-17 is an N-acyl homoserine lactone (AHL)-producing bacterium
isolated from the crown gall tumor of a grapevine. To
our knowledge, this is the first draft genome announcement of a plant-associated
strain from the genus Novosphingobium.

<>

<1>Gan, H.M., Chew, T.H., Tay, Y.L., Lye, S.F., Yahya, A.
<2>Genome Sequence of Hydrogenophaga sp. Strain PBC, a 4-Aminobenzenesulfonate-Degrading Bacterium.
<3>J. Bacteriol.
<4>194
<5>4759-4760
<6>2012
<7>Hydrogenophaga sp. strain PBC is an effective degrader of 4-aminobenzenesulfonate isolated
from textile wastewater. Here we present the assembly and annotation of
its genome, which may provide further insights into its metabolic potential. This
is the first announcement of the draft genome sequence of a strain from the genus
Hydrogenophaga.

<>

<1>Gan, H.M., Chew, T.H., Tay, Y.L., Lye, S.F., Yahya, A.
<2>Genome Sequence of Ralstonia sp. Strain PBA, a Bacterium Involved in the Biodegradation of 4-Aminobenzenesulfonate.
<3>J. Bacteriol.
<4>194
<5>5139-5140
<6>2012
<7>Ralstonia sp. strain PBA was isolated from textile wastewater in a coculture with
Hydrogenophaga sp. strain PBC. Here we present the assembly and annotation of its
genome, which may provide further insights into the mechanism of its interaction
with strain PBC during 4-aminobenzenesulfonate degradation.

<>

<1>Gan, H.M., Lean, S.S., Suhaili, Z., Thong, K.L., Yeo, C.C.
<2>Genome Sequence of Acinetobacter baumannii AC12, a Polymyxin-Resistant Strain Isolated from Terengganu, Malaysia.
<3>J. Bacteriol.
<4>194
<5>5979-5980
<6>2012
<7>Acinetobacter baumannii is a major cause of nosocomial infection worldwide. We report the
draft genome sequence of A. baumannii AC12, a multidrug-resistant
nosocomial strain with additional resistance to carbapenems and polymyxin. The
genome data will provide insights into the genetic basis of antimicrobial
resistance and its adaptive mechanism.

<>

<1>Gan, H.M., McGroty, S.E., Chew, T.H., Chan, K.G., Buckley, L.J., Savka, M.A., Hudson, A.O.
<2>Whole-Genome Sequence of Enterobacter sp. Strain SST3, an Endophyte Isolated from Jamaican Sugarcane (Saccharum sp.) Stalk Tissue.
<3>J. Bacteriol.
<4>194
<5>5981-5982
<6>2012
<7>Enterobacter sp. strain SST3 is an endophytic bacterium isolated from Saccharum spp. Here we
present its annotated draft genome that may shed light on its role
as a bacterial endophyte of sugarcane. To our knowledge, this is the first genome
announcement of a sugarcane-associated bacterium from the genus Enterobacter.

<>

<1>Gan, H.M., Triassi, A.J., Wheatley, M.S., Savka, M.A., Hudson, A.O.
<2>High-Quality Draft Whole-Genome Sequences of Three Strains of Enterobacter Isolated from Jamaican Dioscorea cayenensis (Yellow Yam).
<3>Genome Announcements
<4>2
<5>e00170-14
<6>2014
<7>Here we report the whole-genome sequences of three endophytic bacteria, Enterobacter sp.
strain DC1, Enterobacter sp. strain DC3, and Enterobacter sp.
strain DC4, from root tubers of the yellow yam plant, Dioscorea cayenensis.
Preliminary analyses suggest that the genomes of the three bacteria contain genes
involved in acetoin and indole-3-acetic acid metabolism.

<>

<1>Gan, H.Y., Gan, H.M., Savka, M.A., Triassi, A.J., Wheatley, M.S., Smart, L.B., Fabio, E.S., Hudson, A.O.
<2>Whole-genome sequences of 13 endophytic bacteria isolated from shrub willow (salix) grown in geneva, new york.
<3>Genome Announcements
<4>2
<5>e00288-14
<6>2014
<7>Shrub willow, Salix spp. and hybrids, is an important bioenergy crop. Here we report the
whole-genome sequences and annotation of 13 endophytic bacteria from
stem tissues of Salix purpurea grown in nature and from commercial cultivars and
Salix viminalis x Salix miyabeana grown in bioenergy fields in Geneva, New York.

<>

<1>Gan, H.Y., Gan, H.M., Tarasco, A.M., Busairi, N.I., Barton, H.A., Hudson, A.O., Savka, M.A.
<2>Whole-Genome Sequences of Five Oligotrophic Bacteria Isolated from Deep within Lechuguilla Cave, New Mexico.
<3>Genome Announcements
<4>2
<5>e01133-14
<6>2014
<7>Here, we report the whole-genome sequences and annotation of five oligotrophic bacteria from
two sites within the Lechuguilla Cave in the Carlsbad Caverns
National Park, NM. Three of the five genomes contain an acyl-homoserine lactone
signal synthase ortholog (luxI) that is involved in cell-to-cell communication
via quorum sensing.

<>

<1>Ganesan, A.T.
<2>Genetic recombination during transformation in Bacillus subtilis:  appearance of a deoxyribonucleic acid methylase.
<3>J. Bacteriol.
<4>139
<5>270-279
<6>1979
<7>In Bacillus subtilis the ability to take up deoxyribonucleic acid (DNA) and
undergo genetic transformation may coincide with the induction of defective
phage(s) and the expression of possibly related cryptic genes.  A
restriction-modification enzyme system appears to be expressed.  Targets of the
restriction activity on the DNA can be blocked by methylation catalyzed by the
methyl transferase.  It is shown that cellular DNA becomes progressively
methylated and reaches the maximum level during the peak of competency.
Deoxycytidine residues of both incoming donor and resident DNA are methylated.
The possible participation of these enzymes in recombination and the general
role of crytptic genes in inducible functions are discussed.

<>

<1>Ganesan, A.T.
<2>Uptake, restriction, modification and recombination of DNA molecules during transformation in B. subtilis.
<3>Molecular Cloning and Gene Regulation in Bacilli, Academic Press, Ganesan, A.T., Chang, S., Hoch, J.A., 
<4>0
<5>261-268
<6>1982
<7>Bacillus subtilis can be subjected to regimens of growth conditions that would
permit them to take up single, double and plasmid superhelical DNA molecules.
We know very little of transport mechanisms the cell has in store to handle
different types of molecules.

<>

<1>Gann, A.A.F.
<2>Recognition domains of type I restriction enzymes.
<3>Diss. Abstr.
<4>50
<5>4377B
<6>1990
<7>Type I restriction and modification enzymes recognize asymmetric, bipartite target sequences,
the specificity of which is dictated by a single subunit encoded by the HsdS gene.  Within the
K-family, the S genes of members with different specificities have been sequenced (Gough and
Murray, 1983; Gann et al. 1987).  Comparisons of these reveal two large variable regions, each
of ~450 base pairs, separated by a highly conserved region of ~100 base pairs. Recombination
between the central conserved regions of two S genes, those of StySP and StySB, has produced a
new S gene (StySQ) encoding a functional polypeptide that confers a novel, hybrid specificity
(Fuller-Pace et al., 1984; Nagaraja, et al. 1985).  In this thesis I describe the formation of
a second recombinant S gene, StySJ, which is of reciprocal structure to StySQ.  StySJ
recognizes a target sequence predicted by a model wherein each S polypeptide contains two
structurally independent DNA recognition domains which act together in defining an enzyme's
target sequence.  Site directed mutagenesis was then used to demonstrate that the variable
N-terminal 150 amino acids of an S polypeptide alone constitute one DNA recognition domain.
Two S polypeptides, each deleted for a single recognition domain were also produced.  Though
showing no enzymatic activity in vivo, these truncated polypeptide were capable of inhibiting
the activities of complete restriction and modification enzymes from their own family, but not
from another.  This is interpreted as being due to the truncated S polypeptides binding other
enzyme subunits, thereby disrupting the formation of function restriction complexes.

<>

<1>Gann, A.A.F.
<2>Recognition domains of Type I restriction enzymes.
<3>Ph.D. Thesis, , , Edinburgh
<4>
<5>
<6>1988
<7>Type I restriction and modification enzymes recognize asymmetric, bipartite target sequences,
the specificity of which is dictated by a single subunit encoded by the hsdS gene.  Within the
K-family, the S genes of members with different specificities have been sequenced.
Comparisons of these reveal two large variable regions, each of ~450 base pairs, separated by
a highly conserved region of ~100 base pairs.  Recombination between the central conserved
regions of two S genes, those of StySP and StySB, has produced a new S gene (StySQ) encoding a
functional polypeptide that confers a novel, hybrid specificity.  In this thesis I describe
the formation of a second recombinant S gene, StySJ, which is of reciprocal structure to
StySQ.  StySJ recognizes a target sequence predicted by a model wherein each S polypeptide
contains two structurally independent DNA recognition domains which act together in defining
an enzyme's target sequence.  Site directed mutagenesis was then used to demonstrate that the
variable N-terminal 150 amino acids of an S polypeptide alone constitute one DNA recognition
domain.  Two S polypeptides, each deleted for a single recognition domain were also produced.
Though showing no enzymatic activity in vivo, these truncated polypeptides were capable of
inhibiting the activities of complete restriction and modification enzymes from their own
family, but not from another.  This is interpreted as being due to the truncated S
polypeptides binding other enzyme subunits, thereby disrupting the formation of functional
complexes.

<>

<1>Gann, A.A.F., Campbell, A.J.B., Collins, J.F., Coulson, A.F.W., Murray, N.E.
<2>Reassortment of DNA recognition domains and the evolution of new specificities.
<3>Mol. Microbiol.
<4>1
<5>13-22
<6>1987
<7>TypeI restriction enzymes comprise three subunits only one of which, the S polypeptide,
dictates the specificity of the DNA sequence recognized. Recombination between two different
hsdS genes, SP and SB, led to the isolation of a system, SQ, which had a different specificity
from that of either parent. The finding that the nucleotide sequence recognized by SQ is a
hybrid containing components from both the SP and SB target sequences suggested that DNA
recognition is carried out by two separable domains within each specificity polypeptide.  To
test this we have made the recombinant gene of reciprocal structure and demonstrate that it
encodes a polypeptide whose recognition sequence, deduced in vivo, is as predicted by this
model.  We also report the sequence of the SB specificity gene, so that information is now
available for the five known members of this family of enzymes.  All show a similar
organization of conserved and variable regions.  Comparisons of the predicted amino acid
sequences reveal large non-conserved areas which may not even be structurally similar.  This
is remarkable since these different S subunits are functionally identical, except for the
specificity with respect to the DNA sequence with which they interact.  We discuss the
correlation of the variation in polypeptide sequence with recognition specificities.

<>

<1>Gao, C., Hu, C., Ma, C., Su, F., Yu, H., Jiang, T., Dou, P., Wang, Y., Qin, T., Lv, M., Xu, P.
<2>Genome Sequence of the Lactate-Utilizing Pseudomonas aeruginosa Strain XMG.
<3>J. Bacteriol.
<4>194
<5>4751-4752
<6>2012
<7>Pseudomonas aeruginosa XMG, isolated from soil, utilizes lactate. Here we present a 6.45-Mb
assembly of its genome sequence. Besides the lactate utilization
mechanism of the strain, the genome sequence may also provide other useful
information related to P. aeruginosa, such as identifying genes involved in
virulence, drug resistance, and aromatic catabolism.

<>

<1>Gao, E.B., Gui, J.F., Zhang, Q.Y.
<2>A Novel Cyanophage with a Cyanobacterial Nonbleaching Protein A Gene in the Genome.
<3>J. Virol.
<4>86
<5>236-245
<6>2012
<7>A cyanophage, PaV-LD, has been isolated from harmful filamentous cyanobacterium
Planktothrix agardhii in Lake Donghu, a shallow freshwater lake in China. Here,
we present the cyanophage's genomic organization and major structural proteins.
The genome is a 95,299-bp-long, linear double-stranded DNA and contains 142
potential genes. BLAST searches revealed 29 proteins of known function in
cyanophages, cyanobacteria, or bacteria. Thirteen major structural proteins
ranging in size from 27 kDa to 172 kDa were identified by SDS-PAGE and
mass-spectrometric analysis. The genome lacks major genes that are necessary to
the tail structure, and the tailless PaV-LD has been confirmed by an electron
microscopy comparison with other tail cyanophages and phages. Phylogenetic
analysis of the major capsid proteins also reveals an independent branch of
PaV-LD that is quite different from other known tail cyanophages and phages.
Moreover, the unique genome carries a nonbleaching protein A (NblA) gene (open
reading frame [ORF] 022L), which is present in all phycobilisome-containing
organisms and mediates phycobilisome degradation. Western blot detection
confirmed that 022L was expressed after PaV-LD infection in the host filamentous
cyanobacterium. In addition, its appearance was companied by a significant
decline of phycocyanobilin content and a color change of the cyanobacterial cells
from blue-green to yellow-green. The biological function of PaV-LD nblA was
further confirmed by expression in a model cyanobacterium via an integration
platform, by spectroscopic analysis and electron microscopy observation. The data
indicate that PaV-LD is an exceptional cyanophage of filamentous cyanobacteria,
and this novel cyanophage will also provide us with a new vision of the
cyanophage-host interactions.

<>

<1>Gao, F., Wang, Y., Liu, Y.J., Wu, X.M., Lv, X., Gan, Y.R., Song, S.D., Huang, H.
<2>Genome sequence of Acinetobacter baumannii MDR-TJ.
<3>J. Bacteriol.
<4>193
<5>2365-2366
<6>2011
<7>Acinetobacter baumannii is a species of pathogenic bacteria, originally included in the
aerobic gram-negative bacterium, which is resistant to most antibiotics. In this paper, the
MDR-TJ strain was isolated by the Second Hospital of Tianjin Medical University, China, which
was found resistant to penicillin, spore class, aminoglycosides, quinolones and also imipenem.
The genome sequence of Acinetobacter baumannii strain MDR-TJ was determined using a
combination of 454 pyrosequencing and paired-end sequencing performed by the Roche Genome
Sequencer FLX Systems to generate a scaffolded assembly.

<>

<1>Gao, G., Li, J., Li, T., Zhang, Z., Wang, L., Yuan, X., Wang, Y., Xu, J., Ke, Y., Huang, L., Wang, D., Chen, Z., Xu, X.
<2>Complete Genome Sequence of Brucella canis Strain 118, a Strain Isolated from Canine.
<3>J. Bacteriol.
<4>194
<5>6680
<6>2012
<7>Brucella canis infects several species of animals, and canine is the preferred host. Genome
sequences of strains from different hosts are valuable for
comparative analysis of host adaptation and microevolution. Here, we report the
genome sequence of Brucella canis strain 118, a strain isolated from canine.

<>

<1>Gao, J., Yu, X., Xie, Z.
<2>Draft Genome Sequence of High-Siderophore-Yielding Pseudomonas sp. Strain HYS.
<3>J. Bacteriol.
<4>194
<5>4121
<6>2012
<7>We sequenced the genome of the high-siderophore-yielding strain Pseudomonas sp. HYS and then
analyzed its iron acquisition systems. The 5.6-Mb draft genome
sequence has a special pattern of pyoverdine synthesis clusters and contains an
hmuRSTUV heme uptake cluster, which has a homolog only in some strains of the
order Enterobacteriales.

<>

<1>Gao, L., Zhou, J., Liu, J., Du G, C.J.
<2>Draft Genome Sequence of Gluconobacter oxydans WSH-003, a Strain That Is Extremely Tolerant of Saccharides and Alditols.
<3>J. Bacteriol.
<4>194
<5>4455-4456
<6>2012
<7>Gluconobacter oxydans is known for its incomplete oxidation of a wide range of alcohols,
sugars, and acids in a bioprocess. The corresponding oxidation products
are secreted almost completely into the medium. Here, we present the high-quality
draft genome sequence of G. oxydans WSH-003, an industrial strain with both high
l-sorbose productivity and extreme tolerance to saccharides and alditols.

<>

<1>Gao, P., Tang, Q., An, X.M., Yan, X.X., Liang, D.C.
<2>Structure of HsdS Subunit from Thermoanaerobacter tengcongensis Sheds Lights on Mechanism of Dynamic Opening and Closing of Type I Methyltransferase.
<3>PLoS ONE
<4>6
<5>e17346
<6>2011
<7>Type I DNA methyltransferases contain one specificity subunit (HsdS) and two modification
subunits (HsdM). The electron microscopy model of
M.EcoKI-M2S1 methyltransferase shows a reasonable closed state of this
clamp-like enzyme, but the structure of the open state is still
unclear. The 1.95 angstrom crystal structure of the specificity subunit
from Thermoanaerobacter tengcongensis (TTE-HsdS) shows an unreported
open form inter-domain orientation of this subunit. Based on the
crystal structure of TTE-HsdS and the closed state model of
M.EcoKI-M2S1, we constructed a potential open state model of type I
methyltransferase. Mutational studies indicated that two alpha-helices
(aa30-59 and aa466-495) of the TTE-HsdM subunit are important
inter-subunit interaction sites in the TTE-M2S1 complex. DNA binding
assays also highlighted the importance of the C-terminal region of
TTE-HsdM for DNA binding by the TTE-M2S1 complex. On the basis of
structural analysis, biochemical experiments and previous studies, we
propose a dynamic opening and closing mechanism for type I
methyltransferase.

<>

<1>Gao, Q., Thorson, J.S.
<2>The biosynthetic genes encoding for the production of the dynemicin enediyne core in Micromonospora chersina ATCC53710.
<3>FEMS Microbiol. Lett.
<4>282
<5>105-114
<6>2008
<7>Dynemicin is a novel anthraquinone-fused member of the 10-membered
enediyne antitumor antibiotic family. The development of a genetic system
for the dynemicin producer Micromonospora chersina confirmed, for the
first time, the requirement of the putative enediyne core biosynthetic
genes (dynE8, U14 and U15) and a tailoring oxidase gene (orf23) for
dynemicin production. Cloning and sequence analysis of a 76 kb of genomic
sequence region containing dynE8 revealed a variety of genes conserved
among known enediyne loci. Surprisingly, this fragment and flanking
chromosomal DNA lacked any obvious genes encoding for the biosynthesis of
the anthraquinone, suggesting that the location of genes encoding for the
biosynthesis of the dynemicin enediyne core and the dynemicin
anthraquinone are chromosomally distinct. The demonstrated trace
production of a shunt product from mutant strain QGD23 (Deltaorf23) also
sets the stage for subsequent studies to delineate the key steps in
enediyne core biosynthesis and tailoring.

<>

<1>Gao, X., Liu, K., Qiu, B.-S.
<2>An investigation on the genetic background of  Nostoc flagelliforme by similarity analysis of its partial genomic DNA and  phylogenetic comparison of deduced related species.
<3>Acta Physiol. Plant.
<4>33
<5>1301-1318
<6>2011
<7>Nostoc flagelliforme, which is distributed on arid and semi-arid steppes of northwestern parts
of China, has attracted increasing interest for its stress tolerance. In order to gain more
insight into the genetic background of N. flagelliforme, we sequenced its partial genomic DNA
for
similarity analyses against current public databases, followed
by phylogenetic comparison of N. flagelliforme and the potentially related species deduced
from the similarity analyses. Approximately 430 kb genomic sequence (*5% of genome as a rough
estimate) was determined from 106
distinct genomic clones. Nucleotide BLAST showed that *23.1% of the partial genomic sequence
was similar to N. punctiforme genomic DNA and *12.4% to its plasmid DNA. Similar protein
search by online FASTA-protein program showed 46.2% of the similar proteins had their
corresponding orthologs in N. punctiforme genome. Furthermore, phylogenetic comparison based
on 16S rRNA
sequences showed N. flagelliforme and N. punctiforme clustered closer among the deduced
related species. These results indicated that N. punctiforme might also be potentially close
neighbor species of N. flagelliforme, in addition to the formerly regarded close neighbor
species N. commune
and N. sphaeroids. In general, these data enriched our recognition of the evolutionary
relationship between N. flagelliforme and other Nostoc species, especially N. punctiforme.

<>

<1>Gao, Z., Liu, X., Ruan, L.
<2>Genome Sequence of Anaerophaga sp. Strain HS1, a Novel, Moderately Thermophilic, Strictly Anaerobic Bacterium Isolated from Hot Spring  Sediment.
<3>J. Bacteriol.
<4>193
<5>5572
<6>2011
<7>Anaerophaga sp. strain HS1 was isolated from offshore hot spring sediment in Xiamen, China. It
was identified as a novel, moderately thermophilic,
strictly anaerobic bacterium affiliated with the family Marinilabiaceae
and showed xylanase activity. Here, we describe the 3.88-Mb draft genome
sequence of Anaerophaga sp. strain HS1 and the annotation analysis of
related xylanase genes.

<>

<1>Garbeva, P., van Elsas, J.D., de Boer, W.
<2>Draft Genome Sequence of the Antagonistic Rhizosphere Bacterium Serratia plymuthica Strain PRI-2C.
<3>J. Bacteriol.
<4>194
<5>4119-4120
<6>2012
<7>Serratia plymuthica strain PRI-2C is a rhizosphere bacterial strain with antagonistic activity
against different plant pathogens. Here we present the
5.39-Mb (G+C content, 55.67%) draft genome sequence of S. plymuthica strain
PRI-2C with the aim of providing insight into the genomic basis of its
antagonistic activity.

<>

<1>Garcia, J.C., Urakawa, H., Le VQ, S.L.Y., Klotz, M.G., Nielsen, J.L.
<2>Draft Genome Sequence of Nitrosospira sp. Strain APG3, a Psychrotolerant Ammonia-Oxidizing Bacterium Isolated from Sandy Lake Sediment.
<3>Genome Announcements
<4>1
<5>e00930-13
<6>2013
<7>Bacteria in the genus Nitrosospira play vital roles in the nitrogen cycle. Nitrosospira sp.
strain APG3 is a psychrotolerant betaproteobacterial
ammonia-oxidizing bacterium isolated from freshwater lake sediment. The draft
genome revealed that it represents a new species of cluster 0 Nitrosospira, which
is presently not represented by described species.

<>

<1>Garcia, J.L., Rozas, D., Del Cerro, C., Nogales, J., El-Said, M.M., Diaz, E.
<2>Genome Sequence of Pseudomonas azelaica HBP1, Which Catabolizes 2-Hydroxybiphenyl Fungicide.
<3>Genome Announcements
<4>2
<5>e01248-13
<6>2014
<7>Pseudomonas azelaica HBP1 (DSM 8897) is one of the few bacteria able to completely mineralize
the 2-hydroxybiphenyl biocide. Here, we report the draft
genome sequence of this strain (7.4 Mbp; G+C content, 63.5%) and the findings
obtained from its genome annotation.

<>

<1>Garcia, L.R., Molineux, I.J.
<2>Translocation and specific cleavage of bacteriophage T7 DNA in vivo by EcoKI.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>12430-12435
<6>1999
<7>Infection of Escherichia coli containing the type I restriction enzyme EcoKI by bacteriophage
T7 0.3 mutants leads to restriction during the late stages of genome entry and during DNA
replication. Patterns of cleavage in vivo suggest that some cutting occurs near the midpoint
of two recognition sites, consistent with the idea that EcoKI translocates DNA bidirectionally
through itself and cuts when two enzyme molecules collide. Rapid ejection of a 0.3(+) T7
genome from a bacteriophage lambda particle results in degradation of the infecting DNA by
EcoKI, showing that the normal T7 DNA translocation process delays restriction. A unique
recognition site inserted at the genomic left end allows EcoKI to function as a molecular
motor and to translocate the remaining 39 kilobases of T7 DNA into the cell.

<>

<1>Garcia, P., Monjardin, C., Martin, R., Madera, C., Soberon, N., Garcia, E., Meana, A., Suarez, J.E.
<2>Isolation of new Stenotrophomonas bacteriophages and genomic characterization of temperate phage S1.
<3>Appl. Environ. Microbiol.
<4>74
<5>7552-7560
<6>2008
<7>Twenty-two phages that infect Stenotrophomonas species were isolated
through sewage enrichment and prophage induction. Of them, S1, S3, and S4
were selected due to their wide host ranges compared to those of the other
phages. S1 and S4 are temperate siphoviruses, while S3 is a virulent
myovirus. The genomes of S3 and S4, about 33 and 200 kb, were resistant to
restriction digestion. The lytic cycles lasted 30 min for S3 and about 75
min for S1 and S4. The burst size for S3 was 100 virions/cell, while S1
and S4 produced about 75 virus particles/cell. The frequency of
bacteriophage-insensitive host mutants, calculated by dividing the number
of surviving colonies by the bacterial titer of a parallel, uninfected
culture, ranged between 10(-5) and 10(-6) for S3 and 10(-3) and 10(-4) for
S1 and S4. The 40,287-bp genome of S1 contains 48 open reading frames
(ORFs) and 12-bp 5' protruding cohesive ends. By using a combination of
bioinformatics and experimental evidence, functions were ascribed to 21
ORFs. The morphogenetic and lysis modules are well-conserved, but no
lysis-lysogeny switch or DNA replication gene clusters were recognized.
Two major clusters of genes with respect to transcriptional orientation
were observed. Interspersed among them were lysogenic conversion genes
encoding phosphoadenosine phosphosulfate reductase and GspM, a protein
involved in the general secretion system II. The attP site of S1 may be
located within a gene that presents over 75% homology to a
Stenotrophomonas chromosomal determinant.

<>

<1>Garcia, R.A., Bustamante, C.J., Reich, N.O.
<2>Sequence-specific recognition by cytosine C5 and adenine N6 DNA methyltransferases requires different deformations of DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>7618-7622
<6>1996
<7>DNA methyltransferases modify specific cytosines and adenines within 2-6 bp recognition
sequences.  We used scanning force microscopy and gel shift analysis to show that M.HhaI, a
cytosine C5 methyltransferase, causes only a 2 degree bend upon binding its recognition site.
Our results are consistent with prior crystallographic analysis showing that the enzyme
stabilizes an extrahelical base while leaving the DNA duplex otherwise unperturbed.  In
contrast, similar analysis of M.EcoRI, an adenine N6 DNA methyltransferase, shows an average
bend angle of approximately 52 degrees.  This distortion of DNA conformation by M.EcoRI is
shown to be important for sequence-specific binding.

<>

<1>Garcia, R.G., Brank, A.S., Christman, J.K., Marquez, V.E., Eritja, R.
<2>Synthesis of oligonucleotide inhibitors of DNA (cytosine-C5) methyltransferase containing 5-azacytosine residues at specific sites.
<3>Antisense Nucleic Acid Drug Dev.
<4>11
<5>369-378
<6>2001
<7>The incorporation of 5-azacytosine residues into DNA causes potent inhibition of DNA
(cytosine-C5) methyltransferases.  The synthesis of oligodeoxyribonucleotides incorporating
single or multiple 5-aza-2'-deoxycytidine residues at precise sites was undertaken to
generate an array of sequences containing the reactive 5-azacytosine base as specific target
sites for enzymatic methylation.  Preparation of these modified oligonucleotides requires the
use of 2-(p-nitrophenyl) ethyloxycarbonyl (NPEOC) groups for the protection of exocyclic amino
functions.  These groups are removed under mild conditions, thus avoiding conventional
protocols that are detrimental to the integrity of the 5-azacytosine ring.

<>

<1>Garcia-Alvarez, L. et al.
<2>Meticillin-resistant Staphylococcus aureus with a novel mecA homologue in human and bovine populations in the UK and Denmark: a descriptive study.
<3>Lancet Infect Dis
<4>11
<5>595-603
<6>2011
<7>BACKGROUND: Animals can act as a reservoir and source for the emergence of
novel meticillin-resistant Staphylococcus aureus (MRSA) clones in human
beings. Here, we report the discovery of a strain of S aureus (LGA251)
isolated from bulk milk that was phenotypically resistant to meticillin
but tested negative for the mecA gene and a preliminary investigation of
the extent to which such strains are present in bovine and human
populations. METHODS: Isolates of bovine MRSA were obtained from the
Veterinary Laboratories Agency in the UK, and isolates of human MRSA were
obtained from diagnostic or reference laboratories (two in the UK and one
in Denmark). From these collections, we searched for mecA PCR-negative
bovine and human S aureus isolates showing phenotypic meticillin
resistance. We used whole-genome sequencing to establish the genetic basis
for the observed antibiotic resistance. FINDINGS: A divergent mecA
homologue (mecA(LGA251)) was discovered in the LGA251 genome located in a
novel staphylococcal cassette chromosome mec element, designated type-XI
SCCmec. The mecA(LGA251) was 70% identical to S aureus mecA homologues and
was initially detected in 15 S aureus isolates from dairy cattle in
England. These isolates were from three different multilocus sequence type
lineages (CC130, CC705, and ST425); spa type t843 (associated with CC130)
was identified in 60% of bovine isolates. When human mecA-negative MRSA
isolates were tested, the mecA(LGA251) homologue was identified in 12 of
16 isolates from Scotland, 15 of 26 from England, and 24 of 32 from
Denmark. As in cows, t843 was the most common spa type detected in human
beings. INTERPRETATION: Although routine culture and antimicrobial
susceptibility testing will identify S aureus isolates with this novel
mecA homologue as meticillin resistant, present confirmatory methods will
not identify them as MRSA. New diagnostic guidelines for the detection of
MRSA should consider the inclusion of tests for mecA(LGA251). FUNDING:
Department for Environment, Food and Rural Affairs, Higher Education
Funding Council for England, Isaac Newton Trust (University of Cambridge),
and the Wellcome Trust.

<>

<1>Garcia-Fernandez, A., Villa, L., Carta, C., Venditti, C., Giordano, A., Venditti, M., Mancini, C., Carattoli, A.
<2>Klebsiella pneumoniae ST258 Producing KPC-3 Identified in Italy Carries Novel Plasmids and OmpK36/OmpK35 Porin Variants.
<3>Antimicrob. Agents Chemother.
<4>56
<5>2143-2145
<6>2012
<7>A carbapenemase-resistant Klebsiella pneumoniae strain, clone ST258 producing
KPC-3, was fully characterized. The entire plasmid content was investigated,
thereby identifying plasmids of the IncFII(k) (two of them similar to pKPQIL and
pKPN3, respectively), IncX, and ColE types, carrying a formidable set of
resistance genes against toxic compounds, metals, and antimicrobial drugs and a
novel iron(III) uptake system.

<>

<1>Garcia-Heredia, I., Martin-Cuadrado, A.-B., Mojica, F.J., Santos, F., Mira, A., Anton, J., Rodriguez-Valera, F.
<2>Reconstructing viral genomes from the environment using fosmid clones: the case of haloviruses.
<3>PLoS ONE
<4>7
<5>e33802
<6>2012
<7>Background: Metaviriomes, the viral genomes present in an environment, have been studied by
direct sequencing of the viral DNA or by cloning in small insert libraries. The short reads
generated by both approaches make it very difficult to assemble and annotate such flexible
genomic entities. Many environmental viruses belong to unknown groups or prey on uncultured
and little known cellular lineages, and hence might not be present in databases.
Methodology and Principal Findings: Here we have used a different approach, the cloning of
viral DNA into fosmids before sequencing, to obtain natural contigs that are close to the size
of a viral genome. We have studied a relatively low diversity extreme environment: saturated
NaCl brines, which simplifies the analysis and interpretation of the data. Forty-two different
viral genomes were retrieved, and some of these were almost complete, and could be tentatively
identified as head-tail phages (Caudovirales).
Conclusions and Significance: We found a cluster of phage genomes that most likely infect
Haloquadratum walsbyi, the square archaeon and major component of the community in these
hypersaline habitats. The identity of the prey could be confirmed by the presence of CRISPR
spacer sequences shared by the virus and one of the available strain genomes. Other viral
clusters detected appeared to prey on the Nanohaloarchaea and on the bacterium Salinibacter
ruber, covering most of the diversity of microbes found in this type of environment. This
approach appears then as a viable alternative to describe
metaviriomes in a much more detailed and reliable way than by the more common approaches based
on direct sequencing. An example of transfer of a CRISPR cluster including repeats and spacers
was accidentally found supporting the dynamic nature and frequent transfer of this peculiar
prokaryotic mechanism of cell protection.

<>

<1>Gardes, A., Kaeppel, E., Shehzad, A., Seebah, S., Teeling, H., Yarza, P., Glockner, F.O., Grossart, H.P., Ullrich, M.S.
<2>Complete genome sequence of Marinobacter adhaerens type strain (HP15), a diatom-interacting marine microorganism.
<3>Standards in Genomic Sciences
<4>3
<5>97-107
<6>2010
<7>Marinobacter adhaerens HP15 is the type strain of a newly identified marine species, which is
phylogenetically related to M. flavimaris, M. algicola, and M.
aquaeolei. It is of special interest for research on marine aggregate formation
because it showed specific attachment to diatom cells. In vitro it led to
exopolymer formation and aggregation of these algal cells to form marine snow
particles. M. adhaerens HP15 is a free-living, motile, rod-shaped, Gram-negative
gammaproteobacterium, which was originally isolated from marine particles sampled
in the German Wadden Sea. M. adhaerens HP15 grows heterotrophically on various
media, is easy to access genetically, and serves as a model organism to
investigate the cellular and molecular interactions with the diatom Thalassiosira
weissflogii. Here we describe the complete and annotated genome sequence of M.
adhaerens HP15 as well as some details on flagella-associated genes. M. adhaerens
HP15 possesses three replicons; the chromosome comprises 4,422,725 bp and codes
for 4,180 protein-coding genes, 51 tRNAs and three rRNA operons, while the two
circular plasmids are ~187 kb and ~42 kb in size and contain 178 and 52
protein-coding genes, respectively.

<>

<1>Gardiner, D.M., Stiller, J., Covarelli, L., Lindeberg, M., Shivas, R.G., Manners, J.M.
<2>Genome Sequences of Pseudomonas spp. Isolated from Cereal Crops.
<3>Genome Announcements
<4>1
<5>e00209-13
<6>2013
<7>Compared to those of dicot-infecting bacteria, the available genome sequences of  bacteria
that infect wheat and barley are limited. Herein, we report the draft
genome sequences of four pseudomonads originally isolated from these cereals.
These genome sequences provide a useful resource for comparative analyses within
the genus and for cross-kingdom analyses of plant pathogenesis.

<>

<1>Gardiner, D.M., Stiller, J., Kazan, K.
<2>Genome Sequence of Fusarium graminearum Isolate CS3005.
<3>Genome Announcements
<4>2
<5>e00227-14
<6>2014
<7>Fusarium graminearum is one of the most important fungal pathogens of wheat, barley, and maize
worldwide. This announcement reports the genome sequence of a highly virulent Australian
isolate of this species to supplement the existing genome of the North American F. graminearum
isolate Ph1.

<>

<1>Gardner, A.F., Kumar, S., Perler, F.B.
<2>Genome Sequence of the Model Hyperthermophilic Archaeon Thermococcus litoralis NS-C.
<3>J. Bacteriol.
<4>194
<5>2375-2376
<6>2012
<7>The hyperthermophilic archaeon Thermococcus litoralis strain NS-C, first isolated in 1985, has
been a foundational organism for archaeal research in biocatalysis,
DNA replication, metabolism, and the discovery of inteins. Here, we present the
genome sequence of T. litoralis with a focus on the replication machinery and
inteins.

<>

<1>Gardner, R.C., Howarth, A.J., Messing, J., Shepherd, R.J.
<2>Cloning and sequencing of restriction fragments generated by EcoRI*.
<3>DNA
<4>1
<5>109-115
<6>1982
<7>Thirty-four EcoRI* sites have been identified on the nucleotide sequence of
CaMV, following cloning of EcoRI* fragments in M13mp2.  From this sequencing
data, we have deduced that EcoRI* recognizes sites at any one of the six
positions in the recognition site, with the exception of A5T or T5A changes
within the central tetramer.  The EcoRI* restriction patterns of PhiX174 and
pBR322 are consistent with these recognition criteria.  Similarly, BamHI*
cleavage of PhiX174 and SV40 (George et al., 1980) produces restriction
patterns that are consistent with single-position degeneracy in the canonical
BamHI recognition site.  Cohesive termini produced by EcoRI* cleavage were
ligated into the EcoRI site of M13mp2, even when there was a base pair mismatch
within the four nucleotide overlap.  Mismatches were corrected asymmetrically
during subsequent replication of M13 in E. coli.

<>

<1>Gardner, S.P., Olson, J.W.
<2>Barriers to Horizontal Gene Transfer in Campylobacter jejuni.
<3>Adv. Appl. Microbiol.
<4>79
<5>19-42
<6>2012
<7>Campylobacter jejuni is among the most frequent agent of food-borne gastroenteritis in the
world, but its physiology and pathogenesis. is
less well understood than other bacterial enteric pathogens. This is
due in part to the incompatibility of the molecular tools that have
enabled advances in the characterization of other bacterial species.
Most notably, the dearth of plasmid-based complementation, reporter
assays, and plasmid-based unmarked mutagenesis procedures in many of
the type strains has hindered research progress. The techniques
themselves are not inadequate in Cam pylobacter species, but rather the
barrier to genetic transfer of these genetic constructs from
non-Campylobacter cloning stains such as Escherichia coli. Here, we
review the modes of genetic transfer in C. jejuni and review the
current state of research into the mechanism of each. Also reviewed are
two systems (CRISPR-Cas and restriction modification) that are common
to many strains of C. jejuni and are at least partly responsible for
these barriers.

<>

<1>Garfin, D.E., Boyer, H.W., Goodman, H.M.
<2>Sequences spanning the EcoRI substrate site.
<3>Nucleic Acids Res.
<4>2
<5>1851-1865
<6>1975
<7>Substrate recognition by the EcoRI restriction endonuclease was investigated by
analysis of the nucleotide sequences at the sites of enzymatic cleavage in
various DNA molecules.  5'-end labeling and homochromatographic fingerprinting
led to the determination of a 17-base-pair sequence spanning the EcoRI site of
simian virus 40 DNA and a 15-base-pair sequence overlapping the EcoRI site of
ColE1 plasmid DNA.  Three other DNAs were similarly tested, although extended
sequences were not determined in these cases.  The EcoRI site was shown to be
the symmetric, double-stranded equivalent of -N-G-A-A-T-T-C-N-.

<>

<1>Garfin, D.E., Goodman, H.M.
<2>Nucleotide sequences at the cleavage sites of two restriction endonucleases from Hemophilus parainfluenzae.
<3>Biochem. Biophys. Res. Commun.
<4>59
<5>108-116
<6>1974
<7>The nucleotide sequences at the cleavage sites of two restriction endonucleases from
Hemophilus parainfluenzae, HpaI and HpaII, have been determined.  Terminal labeling at both
the 3'- and 5'-termini was used to show that the HpaI enzyme cleaves the sequence
5'...N-G-T-T^pA-A-C-N...3' 3'...N-C-A-Ap^T-T-G-N...5' and the sequence cleaved by the
HpaII enzyme is 5'...N-C^pC-G-G-N...3' 3'...N-G-G-Cp^C-N...5' where the arrows indicate
the points of strand scission.

<>

<1>Garg, R., Kumari, R., Tiwari, S., Goyal, S.
<2>Genomic Survey, Gene Expression Analysis and Structural Modeling Suggest Diverse Roles of DNA Methyltransferases in Legumes.
<3>PLoS ONE
<4>9
<5>14
<6>2014
<7>DNA methylation plays a crucial role in development through inheritable gene silencing. Plants
possess three types of DNA methyltransferases (MTases), namely Methyltransferase (MET),
Chromomethylase (CMT) and Domains Rearranged Methyltransferase (DRM), which maintain
methylation at CG, CHG and CHH sites. DNA MTases have not been studied in legumes so far.
Here, we report the identification and analysis of putative DNA MTases in five legumes,
including chickpea, soybean, pigeonpea, Medicago and Lotus. MTases in legumes could be
classified in known MET, CMT, DRM and DNA nucleotide methyltransferases (DNMT2) subfamilies
based on their domain organization. First three MTases represent DNA MTases, whereas DNMT2
represents a transfer RNA (tRNA) MTase. Structural comparison of all the MTases in plants with
known MTases in mammalian and plant systems have been reported to assign structural features
in context of biological functions of these proteins. The structure analysis clearly specified
regions crucial for protein-protein interactions and regions important for nucleosome binding
in various domains of CMT and MET proteins. In addition, structural model of DRM suggested
that circular permutation of motifs does not have any effect on overall structure of DNA
methyltransferase domain. These results provide valuable insights into role of various domains
in molecular recognition and should facilitate mechanistic understanding of their function in
mediating specific methylation patterns. Further, the comprehensive gene expression analyses
of MTases in legumes provided evidence of their role in various developmental processes
throughout the plant life cycle and response to various abiotic stresses. Overall, our study
will be very helpful in establishing the specific functions of DNA MTases in legumes.

<>

<1>Garido, M. et al.
<2>Listeria monocytogenes genome,polypeptides and uses.
<3>Japanese Patent Office
<4>JP 2004507217 A
<5>
<6>2004
<7>
<>

<1>Garita-Cambronero, J., Sena-Velez, M., Palacio-Bielsa, A., Cubero, J.
<2>Draft Genome Sequence of Xanthomonas arboricola pv. pruni Strain Xap33, Causal Agent of Bacterial Spot Disease on Almond.
<3>Genome Announcements
<4>2
<5>e00440-14
<6>2014
<7>We report the annotated genome sequence of Xanthomonas arboricola pv. pruni strain Xap33,
isolated from almond leaves showing bacterial spot disease symptoms
in Spain. The availability of this genome sequence will aid our understanding of
the infection mechanism of this bacterium as well as its relationship to other
species of the same genus.

<>

<1>Garnett, J., Halford, S.E.
<2>Properties and subunit structure of EcoRV methyltransferase.
<3>Gene
<4>74
<5>73-76
<6>1988
<7>Meeting Abstract

<>

<1>Garnier, F., Taourit, S., Glaser, P., Courvalin, P., Galimand, M.
<2>Characterization of transposon Tn1549, conferring VanB-type resistance in Enterococcus spp.
<3>Microbiology
<4>146
<5>1481-1489
<6>2000
<7>Transfer of VanB-type resistance to glycopeptides among enterococci has been reported to be
associated with the movement of large chromosomal
genetic elements or of plasmids. The authors report the characterization
of the 34 kb transposon Tn1549 borne by a plasmid related to pAD1 and
conferring vancomycin resistance in clinical isolates of Enterococcus spp.
Tn1549 contained 30 ORFs and appeared to be organized like the Tn916
family of conjugative transposons into three functional regions: (i) the
right end, implicated in the excision-integration process; (ii) the
central part, in which the vanB2 operon replaces the tet(M) gene; and
(iii) the left extremity, in which eight of the 18 ORFs could be
implicated in the conjugative transfer.

<>

<1>Garnier, T. et al.
<2>The complete genome sequence of Mycobacterium bovis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>7877-7882
<6>2003
<7>Mycobacterium bovis is the causative agent of tuberculosis in a range of animal species and
man, with worldwide annual losses to agriculture of $3
billion. The human burden of tuberculosis caused by the bovine tubercle
bacillus is still largely unknown. M. bovis was also the progenitor for
the M. bovis bacillus Calmette-Guerin vaccine strain, the most widely used
human vaccine. Here we describe the 4,345,492-bp genome sequence of M.
bovis AF2122/97 and its comparison with the genomes of Mycobacterium
tuberculosis and Mycobacterium leprae. Strikingly, the genome sequence of
M. bovis is >99.95% identical to that of M. tuberculosis, but deletion of
genetic information has led to a reduced genome size. Comparison with M.
leprae reveals a number of common gene losses, suggesting the removal of
functional redundancy. Cell wall components and secreted proteins show the
greatest variation, indicating their potential role in host-bacillus
interactions or immune evasion. Furthermore, there are no genes unique to
M. bovis, implying that differential gene expression may be the key to the
host tropisms of human and bovine bacilli. The genome sequence therefore
offers major insight on the evolution, host preference, and pathobiology
of M. bovis.

<>

<1>Garrett, M., Parker, J., Stephens, C.M.
<2>Draft Genome Sequences of Antibiotic-Resistant Commensal Escherichia coli.
<3>Genome Announcements
<4>2
<5>e00873-14
<6>2014
<7>Antimicrobial resistance is a significant public health issue. We report here the draft genome
sequences of three drug-resistant strains of commensal Escherichia
coli isolated from a single healthy college student. Each strain has a distinct
genome, but two of the three contain an identical large plasmid with multiple
resistance genes.

<>

<1>Garrett, R.A., Prangishvili, D., Shah, S.A., Reuter, M., Stetter, K.O., Peng, X.
<2>Metagenomic analyses of novel viruses and plasmids from a cultured environmental sample of hyperthermophilic neutrophiles.
<3>Environ. Microbiol.
<4>12
<5>2918-2930
<6>2010
<7>Summary Two novel viral genomes and four plasmids were assembled from an
environmental sample collected from a hot spring at Yellowstone National
Park, USA, and maintained anaerobically in a bioreactor at 85 degrees C
and pH 6. The double-stranded DNA viral genomes are linear (22.7 kb) and
circular (17.7 kb), and derive apparently from archaeal viruses HAV1 and
HAV2. Genomic DNA was obtained from samples enriched in filamentous and
tadpole-shaped virus-like particles respectively. They yielded few
significant matches in public sequence databases reinforcing, further, the
wide diversity of archaeal viruses. Several variants of HAV1 exhibit major
genomic alterations, presumed to arise from viral adaptation to different
hosts. They include insertions up to 350 bp, deletions up to 1.5 kb, and
genes with extensively altered sequences. Some result from recombination
events occurring at low complexity direct repeats distributed along the
genome. In addition, a 33.8 kb archaeal plasmid pHA1 was characterized,
encoding a possible conjugative apparatus, as well as three cryptic
plasmids of thermophilic bacterial origin, pHB1 of 2.1 kb and two closely
related variants pHB2a and pHB2b, of 5.2 and 4.8 kb respectively.
Strategies are considered for assembling genomes of smaller genetic
elements from complex environmental samples, and for establishing possible
host identities on the basis of sequence similarity to host CRISPR immune
systems.

<>

<1>Garrigues, C., Johansen, E., Pedersen, M.B.
<2>Complete genome sequence of Bifidobacterium animalis subsp. lactis BB-12, a widely consumed probiotic strain.
<3>J. Bacteriol.
<4>192
<5>2467-2468
<6>2010
<7>Bifidobacterium animalis subsp. lactis BB-12 is a commercially available probiotic strain used
throughout the world in a variety of functional
foods and dietary supplements. The benefits of BB-12 have been documented
in a number of independent clinical trials. Determination of the complete
genome sequence reveals a single circular chromosome of 1,942,198 bp with
1,642 predicted protein-encoding genes, 4 rRNA operons, and 52 tRNA genes.
Knowledge of this sequence will lead to insight into the specific features
which give this strain its probiotic properties.

<>

<1>Garvey, P., van Sinderen, D., Twomey, D.P., Hill, C., Fitzgerald, G.F.
<2>Molecular genetics of bacteriophage and natural phage defense systems in the genus Lactococcus.
<3>Int. Dairy Journal
<4>5
<5>905-947
<6>1995
<7>Bacteriophage infection of starter cultures used in a range of milk fermentation processes,
particularly those involving Lactococcus lactis, poses a significant problem in industrial
practice.  The application of genetic and molecular technologies to the study of lactococcal
bacteriophages has proven to be very rewarding in terms of understanding the nature of phage
with respect to their physical and genetic organisation.  The availability of the full genomic
sequence of a number of phages provides an unambiguous basis for determining the relationship
between them, for elucidating their evolutionary progression and will also yield strategies
for obstructing successful phage proliferation on previously sensitive hosts.  The genetic
analysis of plage/host interactions has also highlighted the presence of natural defense
systems (e.g. adsorption blocking, inhibition of phage DNA entry, restriction modification and
abortive infection) in lactococci.  A number of restriction modification systems and abortive
infection mechanisms have been characterized at a molecular level and the genes involved have
been cloned and sequenced.  Plasmid-encoded phage resistance mechanisms can be exploited to
generate strains which can successfully counter phage proliferation and will provide a basis
for understanding the complex interactions between phages and their target hosts at a
molecular level.

<>

<1>Garzetti, D., Heesemann, J., Rakin, A.
<2>Genome Sequences of Four Yersinia enterocolitica Bioserotype 4/O:3 Isolates from  Mammals.
<3>Genome Announcements
<4>1
<5>e00466-13
<6>2013
<7>We report here the complete genome sequences of four European Yersinia enterocolitica
mammalian isolates of bioserotype 4/O:3. The genomes have an
average size of 4.50 Mb, a G+C content of 47%, and between 4,231 and 4,330 coding
sequences (CDSs). No relevant differences were detected by genome comparison
between mammalian and human isolates.

<>

<1>Gasiunas, G., Sasnaukas, G., Tamulaitis, G., Siksnys, V.
<2>Tetrameric restriction enzyme Cfr42I belongs to the GIY-YIG nuclease family.
<3>FEBS J.
<4>276
<5>140-141
<6>2009
<7>The GIY-YIG nuclease domain was originally identified in homing endonucleases and enzymes
involved in DNA repair and recombination.  Despite wide distribution of this domain,
biochemical and structural studies of GIY-YIG proteins are often hampered by their
multi-domain organization and complex functional requirements.  Many of the GIY-YIG family
enzymes are functional as monomers.  We show that the Cfr42I restriction endonuclease which
belongs to the GIY-YIG family and recognizes the symmetric sequence 5'-CCGC/GG-3' ('/'
indicates the cleavage site) is a tetramer in solution.  Moreover, biochemical and kinetic
studies demonstrate that the Cfr42I tetramer is catalytically active only upon simultaneous
binding of two copies of its recognition sequence.  In that respect Cfr42I resembles the
homotetrameric Type IIF restriction enzymes that belong to the distinct PD-D/E)XK nuclease
superfamily.  Unlike the PD-(D/E)XK enzymes, the GIY-YIG nuclease Cfr42I accommodates an
extrmely wide selection of metal ion cofactors, including Mg2+, Mn2+, Co2+, Zn2+, Ni2+, Cu2+
and Ca2+.  To our knowledge, Cfr42I is the first tetrameric GIY-YIG family enzyme.  Similar
structural arrangement and phenotypes displayed by restriction enzymes belonging to the
PD-(D/E)XK and GIY-YIG nuclease families point to the functional significance of
tetramerization.  In order to understand structure-function relationship within Cfr42I
restriction enzyme we aim to determine its 3D structure by X-ray crystallography.

<>

<1>Gasiunas, G., Sasnauskas, G., Tamulaitis, G., Urbanke, C., Razaniene, D., Siksnys, V.
<2>Tetrameric restriction enzymes: expansion to the GIY-YIG nuclease family.
<3>Nucleic Acids Res.
<4>36
<5>938-949
<6>2008
<7>The GIY-YIG nuclease domain was originally identified in homing endonucleases and enzymes
involved in DNA repair and recombination. Many of the GIY-YIG family enzymes are functional as
monomers. We show here that the Cfr42I restriction endonuclease which belongs to the GIY-YIG
family and recognizes the symmetric sequence 5'-CCGC/GG-3' ('/' indicates the cleavage
site) is a tetramer in solution. Moreover, biochemical and kinetic studies provided here
demonstrate that the Cfr42I tetramer is catalytically active only upon simultaneous binding of
two copies of its recognition sequence. In that respect Cfr42I resembles the homotetrameric
Type IIF restriction enzymes that belong to the distinct PD-(E/D)XK nuclease superfamily.
Unlike the PD-(E/D)XK enzymes, the GIY-YIG nuclease Cfr42I accommodates an extremely wide
selection of metal-ion cofactors, including Mg(2+), Mn(2+), Co(2+), Zn(2+), Ni(2+), Cu(2+) and
Ca(2+). To our knowledge, Cfr42I is the first tetrameric GIY-YIG family enzyme. Similar
structural arrangement and phenotypes displayed by restriction enzymes of the PD-(E/D)XK and
GIY-YIG nuclease families point to the functional significance of tetramerization.

<>

<1>Gasperik, J., Godany, A., Hostinova, E., Zelinka, J.
<2>Specific endonuclease Sau3239I from Streptomyces aureofaciens CCM 3239.
<3>Biologia (Bratisl)
<4>38
<5>315-319
<6>1983
<7>In the research of relations of plasmid DNA to antibiotic production in
Streptomyces aureofaciens also the prototrophic chlortetracycline (CTC)
producing strain of S. aureofaciens CCM 3239 was studied.  In this strain the
presence of plasmid DNA was proven (unpublished data).  After elimination of
the plasmid there were considerations on using this strain as the acceptor of
DNA transformation.  The possibility of detection of a restriction-modifying
system on this extrachromosomal element as found in eubacteria, (Arber, 1974)
remained open.  Detection of restrictase activity in S. aureofaciens CCM 3239
was interest as restriction endonucleases were isolated from more than 15 types
of Streptomyces (Roberts, 1982).  Restriction endonuclease, type II, was also
found in the non-producing mutant of S. aureofaciens IKA 18/4 (Timko, et al.,
1978).  The substrate specificity and the cleavage site of this enzyme, SauI,
is described in the paper by Timko et al., (1981).

<>

<1>Gassner, C., Schneider-Scherzer, E.S., Lottspeich, F., Schweiger, M., Auer, B.
<2>Escherichia coli bacteriophage T1 DNA methyltransferase appears to interact with Escherichia coli enolase.
<3>Biol. Chem.
<4>379
<5>621-623
<6>1998
<7>Infection of Escherichia coli cells with bacteriophage T1 induces synthesis of a
bacteriophage-specific DNA methyltransferase with a specificity for adenine residues in the
sequence 5'-GATC-3'.  Purification of M.EcoT1 allowed the determination of the coding
sequence of the gene.  The peptide of the entire coding sequence was over-expressed as a
histidine-hexapeptide tagged protein in E. coli.  Affinity purification using a Ni2+ chelating
(Ni-NTA) resin yielded a recombinant enzyme with almost the same enzymatic properties as the
protein purified from T1 infected E. coli cells.  Interestingly, in both purification
procedures, a protein with a molecular weight of 50000 was found to copurify with M.EcoT1.
The N-terminal amino acid sequence identified these proteins in both cases as E. coli enolase.

<>

<1>Gast, F.U., Brinkmann, T., Pieper, U., Kruger, T., Noyer-Weidner, M., Pingoud, A.
<2>The recognition of methylated DNA by the GTP-dependent restriction endonuclease McrBC resides in the N-terminal domain of McrB.
<3>Biol. Chem.
<4>378
<5>975-982
<6>1997
<7>McrBC is a GTP-dependent restriction endonuclease of E. coli K12, selectively directed against
DNA containing modified cytosine residues.  McrB, one of its components, is responsible for
the binding and, together with McrC, for the cleavage of DNAs containing two 5'-PumC sites
separated by 40-80 base pairs.  Gel retardation assays with wild-type and mutant McrB reveal
that (i) single 5'-PumC sites in DNA can be sufficient to elicit binding by McrB.  Binding to
such substrates is, however, weak and strongly dependent on the sequence context of PumC
sites.  (ii) Strong DNA binding (Kass ~ 10^7M-1) is dependent on the presence of at least two
PumC sites, even if they are separated by less than 40 bp, and is modulated by the sequence
context (-AmCCGGT->-AmCTc/gAGT->-AGGmCCT->-AAGmCTT-).  (iii) DNA binding by McrB is
accompanied by formation of distinct multiple complexes whose distribution is modulated by
GTP. (iv) McrC, which cannot bind DNA by itself, moderately stimulates the DNA binding of McrB
and converts McrB-DNA complexes to large aggregates.  (v) Deletion of the C-terminal half of
McrB, which harbors the three consensus sequences characteristic for guanine nucleotide
binding proteins, leads to protein inactive in GTP binding and/or hydrolysis and in
McrC-assisted DNA cleavage; the protein, however, remains fully competent in DNA binding.
(vi) Mutations in McrB which lead to a reduction in GTP binding and/or hydrolysis can affect
DNA binding, suggesting that the two activities are coupled in the full-length protein.

<>

<1>Gaudet, F., Talbot, D., Leonhardt, H., Jaenisch, R.
<2>A short DNA methyltransferase isoform restores methylation in vivo.
<3>J. Biol. Chem.
<4>273
<5>32725-32729
<6>1998
<7>Two murine DNA methyltransferase isoforms (MTases) have been observed, a longer form in
somatic and embryonic stem cells and a shorter form in oocytes and preimplantation embryos.
While the longer MTase is associated with maintenance methyltransferase activity in
replicating cells, little is known about the shorter form.  We present genetic and biochemical
evidence that both isoforms are expressed from the same Dnmt1 gene by using different
translation initiation sites in exons 1 and 4.  We further demonstrate that the shorter
isoform can functionally rescue Dnmt1 null ES cells that have a hypomethylated genome.  These
rescued ES cells differentiate in vivo into a variety of cell types, unlike the Dnmt1 null ES
cells that die upon induction of differentiation.  These results show that the shorter isoform
can substitute for the longer maintenance MTase in ES and differentiated cells.  Our data
further indicate that the shorter MTase isoform found in oocytes is fully functional in vivo
and may play an active role in the regulation of DNA methylation and the establishment of
imprinting patterns.

<>

<1>Gauntlett, J.C., Nilsson, H.-O., Fulurija, A., Marshall, B.J., Benghezal, M.
<2>Phase-variable restriction/modification systems are required for Helicobacter pylori colonization.
<3>Gut Pathog.
<4>6
<5>35
<6>2014
<7>Background: One mechanism utilized by bacterial pathogens for host adaptation and immune
evasion is the generation of phenotypic diversity by the phasevarion that results from the
differential expression of a suite of genes regulated by the activity of a phase-variable
methyltransferase within a restriction modification (RM) system. Phasevarions are active in
Helicobacter pylori, however there have been no studies investigating the significance of
phase-variable RM systems on host colonization.Methods: Two mutant types incapable of phase
variation were constructed; a clean deletion mutant ('DEL') and a mutant ('ON') where the
homopolymeric repeat was replaced with a non-repeat synonymous sequence, resulting in
expression of the full-length protein. The resulting mutants were assessed for their
colonisation ability in the mouse model.Results: Five phase-variable genes encoding either
methyltransferases or members of RM systems were found in H. pylori OND79. Our mutants fell
into three categories; 1, those with little effect on colonization, 2, those where expression
of the full-length protein was detrimental, 3, those where both mutations were
detrimental.Conclusions: Our results demonstrated that phase-variable methyltransferases are
critical to H. pylori colonization, suggesting that genome methylation and generation of
epigenetic diversity is important for colonization and pathogenesis. The third category of
mutants suggests that differential genome methylation status of H. pylori cell populations,
achieved by the phasevarion, is essential for host adaptation. Studies of phase-variable RM
mutants falling in the two other categories, not strictly required for colonization, represent
a future perspective to investigate the role of phasevarion in persistence of H. pylori.

<>

<1>Gauthier, A., Turmel, M., Lemieux, C.
<2>A group I intron in the chloroplast large subunit rRNA gene of Chlamydomonas eugametos encodes a double-strand endonuclease that cleaves the homing site of this intron.
<3>Curr. Genet.
<4>19
<5>43-47
<6>1991
<7>During interspecific crosses between Chlamydomonas eugametos and Chlamydomonas moewusii, an
optional group I intron of 955 base pairs (CeLSU 5) in the C. eugametos chloroplast large
subunit rRNA gene undergoes a duplicative transposition event which is associated with
frequent co-conversion of flanking cpDNA sequences. In the present study, we show that the
basic protein of 218 amino acids encoded by CeLSU 5 could mediate the phenomenon of intron
transposition, also called intron homing. We overexpressed the ORF specifying this protein in
E. coli using expression vectors that contain a C. moewusii cpDNA sequence encompassing the
intron homing site. The expression product was found to exhibit a double-strand DNA
endonuclease activity that is specific for the homing site. This activity was detected in vivo
by self linearization of the expression plasmids.

<>

<1>Gautier, F., Bunemann, H., Grotjahn, L.
<2>Analysis of calif-thymus satellite DNA:  evidence for specific methylation of cytosine in C-G sequences.
<3>Eur. J. Biochem.
<4>80
<5>175-183
<6>1977
<7>Digestion of purified calf thymus satellite I (density = 1.714 g/cm3) with a
series of restriction enzymes shows that modification in this satellite occurs
preferentially in the sequence C-G. This was also shown to be the case in the
other satellites and in bulk chromosomal calf thymus DNA.  Cloning of purified
satellite I DNA in Escherichia coli makes sites, previously modified, available
for cutting with certain restriction enzymes.  All these new sites contain the
sequence C-G.  High-resolution mass spectroscopy establishes that the
satellites contain a low concentration of 5-methylcytosine.  This infers that
methylation which inhibits restriction enzyme cutting must occur preferentially
in the sequence C-G.  Hybridization of cRNA of cloned satellite I DNA with the
satellites III (density = 1.706 g/cm3) and IV (density = 1.710 g/cm3) shows
that there is no or little sequence homology between these satellites.
Digestion of calf thymus satellite I DNA with endoR.EcoRI and subsequent
hybridization studies with the fragments shows two EcoRI fragments in addition
to the usual 1400-base-pair EcoRI repeat unit.

<>

<1>Gautier, M., Chopin, M.-C.
<2>Plasmid-determined systems for restriction and modification activity and abortive infection in Streptococcus cremoris.
<3>Appl. Environ. Microbiol.
<4>53
<5>923-927
<6>1987
<7>Streptococcus cremoris strain IL964 possessed a restriction and modification
(R/M) activity which resulted in a bacteriophage efficiency of plating 5x10-6.
Phage sensitivity of protoplast-induced plasmid-cured derivatives indicated
that two plasmids called pIL103 (5.7 kiobases) and pIL107 (15.2 kilobases) were
each coding for one R/M system.  Plasmid pIL103-encoded R/M was ascertained by
transfer into the plasmid-free, R-/M- strain IL1403 of S. lactis, using
protoplast cotransformation.  This procedure failed for pIL107 because of some
degree of incompatibility between pIL107 and the indicator plasmid pHV1301 used
in cotransformation experiments.  We also observed that plasmid pIL105 (8.7
kilobases) which showed no incidence of phage sensitivity in the parental
strain IL964, mediated abortive infection in strain IL1403.  In 97% of the
infected cells, the phage infection was abortive, while in the remaining 3%
phages were produced with a decreased burst size (50 instead of 180).

<>

<1>Gautier, M., Veaux, M., Chopin, M.-C.
<2>Cloning of three plasmid-determined systems for restriction/modification and abortive infection.
<3>FEMS Microbiol. Rev.
<4>46
<5>44
<6>1987
<7>We have previously described three plasmids encoding phage-defense mechanisms in Streptococcus
lactis and S. cremoris. Plasmids pIL7 (33 kilobases) and pIL103 (5.7 kb) each encodes for a
restriction/modification (R/M) system and pIL105 (9.5 kb) encodes phage abortive infection. We
cloned each of these mechanisms into the plasmid-free, S. lactis strain IL1403. The entire
small pIL103 plasmid was inserted into the single EcoRI site of vector plasmid pIL204.
Recombinant DNA molecules formed were used in transformation of S. lactis IL1403 protoplasts.
Transformants selected for the pIL204-conferred erythromycin resistance were checked for
resistance to phage 66. All the phage-resistant clones harbored pIL103::pIL204 recombinant
DNA. In some cases deletions occured allowing the isolation of a 3.5 kb fragment carrying
genes coding for R/M activity. In the same way, the entire pIL105 plasmid together with
abortive infection activity was cloned into the single XbaI site of plasmid vector pIL252. A
6.5 kb fragment carrying genes coding for R/M activity was isolated from the large pIL7
plasmid following partial digestion with Sau3A and insertion of the fragments into the single
BamHI site of plasmid vector pIL252. Experiments are currently in progress to bring together
the 3 cloned fragments on the same vector plasmid in order to stack phage defense mechanisms
and construct strains improved in their phage resistance.

<>

<1>Gawthorne, J.A., Beatson, S.A., Srikhanta, Y.N., Fox, K.L., Jennings, M.P.
<2>Origin of the Diversity in DNA Recognition Domains in Phasevarion Associated modA Genes of Pathogenic Neisseria and Haemophilus influenzae.
<3>PLoS ONE
<4>7
<5>e32337
<6>2012
<7>Phase variable restriction-modification (R-M) systems have been identified in a range of
pathogenic bacteria. In some it has been
demonstrated that the random switching of the mod (DNA
methyltransferase) gene mediates the coordinated expression of multiple
genes and constitutes a phasevarion (phase variable regulon). ModA of
Neisseria and Haemophilus influenzae contain a highly variable, DNA
recognition domain (DRD) that defines the target sequence that is
modified by methylation and is used to define modA alleles. 18 distinct
modA alleles have been identified in H. influenzae and the pathogenic
Neisseria. To determine the origin of DRD variability, the 18 modA DRDs
were used to search the available databases for similar sequences.
Significant matches were identified between several modA alleles and
mod gene from distinct bacterial species, indicating one source of the
DRD variability was via horizontal gene transfer. Comparison of DRD
sequences revealed significant mosaicism, indicating exchange between
the Neisseria and H. influenzae modA alleles. Regions of high inter-and
intra-allele similarity indicate that some modA alleles had undergone
recombination more frequently than others, generating further
diversity. Furthermore, the DRD from some modA alleles, such as modA12,
have been transferred en bloc to replace the DRD from different modA
alleles.

<>

<1>Gayle, R., Bennett, G.
<2>Synthesis of a specific sequence of DNA using restriction enzymes.
<3>Miami BioTechnology Winter Symposium, Academic Press, Ahmad, F., Schultz, J., Smith, E.E., Whelan, W.J., 
<4>19
<5>526
<6>1982
<7>The restriction enzyme MboII recognizes the DNA sequence GAAGA and cleaves the
DNA eight bases downstream from this site, leaving a single 3' protruding base.
There are no apparent sequence requirements in the stretch of DNA between the
recognition sequence and the point of cleavage.  Using this property, a segment
of DNA from one fragment can be transferred to another fragment.

<>

<1>Gayle, R.B., Auger, E.A., Gough, G.R., Gilham, P.T., Bennett, G.N.
<2>Formation of MboII vectors and cassettes using asymmetric MboII linkers.
<3>Gene
<4>54
<5>221-228
<6>1987
<7>Class-IIS restriction endonucleases such as MboII cleave DNA at a specified
distance away from their recognition sequences.  This feature was exploited to
cleave DNA at previously inaccessible locations by preparing special asymmetric
linker/adapters containing the MboII recognition sequence.  These could be
joined to DNA fragments and subsequently cleaved by MboII.  Attachment of a 3'
phosphate to one of the two different oligodeoxynucleotides comprising the
asymmetric duplex prevented ligation at the improper end of the linker.
Plasmids were contructed containing a unique BamHI or BclI site between the
recognition and cleavage site of MboII.  These sites were used to introduce a
foreign fragment into the plasmid at a position permitting MboII to cleave
within the newly inserted fragment.  Once cleaved at the unique MboII site,
another DNA fragment was inserted.  DNA was thus inserted at a sequence not
previously accessible to specific cleavage by a restriction enzyme.  A cassette
containing an identifiable marker, the lac operator, between two oppositely
oriented MboII/BamHI linkers was made and tested in a random insertion linker
mutagenesis experiment.

<>

<1>Gazzola, S., Pietta, E., Bassi, D., Fontana, C., Puglisi, E., Cappa, F., Cocconcelli, P.S.
<2>Draft Genome Sequence of Vancomycin-Heteroresistant Staphylococcus epidermidis Strain UC7032, Isolated from Food.
<3>Genome Announcements
<4>1
<5>e00709-13
<6>2013
<7>Staphylococcus epidermidis strain UC7032 was isolated from ready-to-eat cured meat and is
heteroresistant to glycopeptide antibiotics. The draft whole-genome
analysis revealed that this strain shows common characteristics typical of
strains that are involved in nosocomial infections.

<>

<1>Ge, F., Li, W., Chen, G., Liu, Y., Zhang, G., Yong, B., Wang, Q., Wang, N., Huang, Z., Li, W., Wang, J., Wu, C., Xie, Q., Liu, G.
<2>Draft genome sequence of Gordonia neofelifaecis NRRL B-59395, a cholesterol-degrading actinomycete.
<3>J. Bacteriol.
<4>193
<5>5045-5046
<6>2011
<7>We report a draft sequence of the genome of Gordonia neofelifaecis NRRL B-59395, a
cholesterol-degrading actinomycete isolated from fresh faeces
of a clouded leopard (Neofelis nebulosa). As predicted, the reported
genome contains several gene clusters for cholesterol degradation. This is
the second available genome sequence of the family Gordoniaceae.

<>

<1>Ge, X., Zhao, Y., Hou, W., Zhang, W., Chen, W., Wang, J., Zhao, N., Lin, J., Wang, W., Chen, M., Wang, Q., Jiao, Y., Yuan, Z., Xiong, X.
<2>Complete Genome Sequence of the Industrial Strain Gluconobacter oxydans H24.
<3>Genome Announcements
<4>1
<5>e00003-13
<6>2013
<7>is characterized by its ability to incompletely oxidize carbohydrates and alcohols. The high
yields of its oxidation products and complete secretion into
the medium make it important for industrial use. We report the finished genome
sequence of H24, an industrial strain with high l-sorbose productivity.

<>

<1>Ge, Y.Z., Pu, M.T., Gowher, H., Wu, H.P., Ding, J.P., Jeltsch, A., Xu, G.L.
<2>Chromatin targeting of de novo DNA methyltransferases by the PWWP domain.
<3>J. Biol. Chem.
<4>279
<5>25447-25454
<6>2004
<7>DNA methylation patterns of mammalian genomes are generated in gametogenesis and early
embryonic development. Two de novo DNA
methyltransferases, Dnmt3a and Dnmt3b, are responsible for the process.
Both enzymes contain a long N-terminal regulatory region linked to a
conserved C-terminal domain responsible for the catalytic activity.
Although a PWWP domain in the N-terminal region has been shown to bind
DNA in vitro, it is unclear how the DNA methyltransferases access their
substrate in chromatin in vivo. We show here that the two proteins are
associated with chromatin including mitotic chromosomes in mammalian
cells, and the PWWP domain is essential for the chromatin targeting of
the enzymes. The functional significance of PWWP-mediated chromatin
targeting is suggested by the fact that a missense mutation in this
domain of human DNMT3B causes immunodeficiency, centromeric
heterochromatin instability, facial anomalies (ICF) syndrome, which is
characterized by loss of methylation in satellite DNA, pericentromeric
instability, and immunodeficiency. We demonstrate that the mutant
protein completely loses its chromatin targeting capacity. Our data
establish the PWWP domain as a novel chromatin/chromosome-targeting
module and suggest that the PWWP-mediated chromatin association is
essential for the function of the de novo methyltransferases during
development.

<>

<1>Ge, Z., Taylor, D.E.
<2>Contributions of genome sequencing to understanding the biology of Helicobacter pylori.
<3>Annu. Rev. Microbiol.
<4>53
<5>353-387
<6>1999
<7>About half of the world's population carries Helicobacter pylori, a gram-negative, spiral
bacterium that colonizes the human stomach. The link between H. pylori and, ulceration as well
as its association with the development of both gastric cancer and mucosa-associated lymphoid
tissue lymphoma in humans is a serious public health concern. The publication of the genome
sequences of two stains of H. pylori gives rise to direct evidence on the genetic diversity
reported previously with respect to gene organization and nucleotide variability from strain
to strain. The genome size of H. pylori strain 26695 is 1,6697,867 bp and is 1,643,831 bp for
strain J99. Approximately 89% of the predicted open reading frames are common to both of the
strains, confirming H. pylori as a single species. A region containing approximately 45% of H.
pylori strain-specific open reading frames, termed the plasticity zone, is present on the
chromosomes, verifying that some strain variability exists. Frequent alteration of nucleotides
in the third position of the triplet codons and various copies of insertion elements on the
individual chromosomes appear to contribute to distinct polymorphic fingerprints among strains
analyzed by restriction fragment length polymorphisms, random amplified polymorphic DNA
method, and repetitive element-polymerase chain reaction. Disordered chromosomal locations of
some genes seen by pulsed-field gel electrophoresis are likely caused by rearrangement or
inversion of certain segments in the genomes. Cloning and functional characterization of the
genes involved in acidic survival, vacuolating toxin, cag-pathogenicity island, motility,
attachment to epithelial cells, natural transformation, and the biosynthesis of
lipopolysaccharides have considerably increased our understanding of the molecular genetic
basis for the pathogenesis of H. pylori. The homopolymeric nucleotide tracts and dinucleotide
repeats, which potentially regulate the on- and off-status of the target genes by the
strand-slipped mispairing mechanism, are often found in the genes encoding the outer-membrane
proteins, in enzymes for lipopolysaccharide synthesis, and within DNA modification/restriction
systems. Therefore, these genes may be involved in the H. pylori-host interaction.

<>

<1>Geahigan, K.B., Meints, G.A., Hatcher, M.E., Orban, J., Drobny, G.P.
<2>The dynamic impact of CpG methylation in DNA.
<3>Biochemistry
<4>39
<5>4939-4946
<6>2000
<7>Solid-state deuterium NMR is used to investigate perturbations of the local, internal dynamics
in the EcoRI restriction binding site, -GAATTC- induced by cytidine methylation. Methylation
of the cytidine base in this sequence is known to suppress hydrolysis by the EcoRI restriction
enzyme. Previous solid-state deuterium NMR studies have detected large amplitude motions of
the phosphate-sugar backbone at the AT-CG junction of the unmethylated DNA sequence. This
study shows that methylation of the cytidine base in a CpG dinucleotide reduces the amplitudes
of motions of the phosphate-sugar backbone. These observations suggest a direct link between
suppression of the amplitudes of localized, internal motions of the sugar-phosphate backbone
of the DNA and inhibition of restriction enzyme cleavage.

<>

<1>Gebreyesus, K.H., Owens, R.A.
<2>Selective protection of restriction endonuclease sites.
<3>Biotechniques
<4>34
<5>512-523
<6>2003
<7>A difficulty that is encountered when attempting to insert a PCR-amplified product or DNA
fragment of interest into a particular vector is the
presence within the insert of one or more internal restriction
endonuclease (RE) sites identical to those selected for the flanks of the
insert. Our method circumvents this problem by partially protecting
internal RE sites while flanking sites for the same RE are cleaved. The
amplified product is first heat denatured in the presence of excess
amounts of perfectly complementary oligonucleotides that can anneal to the
flanks of the insert. The mixture is allowed to anneal and is subsequently
digested with the appropriate endonucleases. This results in the cleavage
of the flanking RE sites while digestion at the internal RE site is not
efficient. The mixture is subsequently heat denatured and column purified
to remove the oligonucleotides. The product is then allowed to anneal and
can be used directly in a ligation reaction with the plasmid vector. This
method facilitates the construction of recombinant molecules by creating
desired flanks while preserving internal RE sites.

<>

<1>Geck, P., Molnar, A., Nasz, I.
<2>Identification of ATGCAT sequence at sites of SmuCI restriction endonuclease by computer and physical mapping of Adenovirus Type I DNA.
<3>Acta Microbiol. Hung.
<4>38
<5>47-53
<6>1991
<7>Physical mapping of adenovirus type I DNA was carried out in order to analyze the recognition
sequence of a novel Streptococcus restriction endonuclease. In addition to the new map and
homology data on this poorly analyzed serotype, the result offers the definite evidence for
the ATGCAT recognition sequence on adenovirus DNA and the physical map of cleavage points.

<>

<1>Geck, P., Molnar, A., Nasz, I.
<2>A mathematical method for the identification of the ATGCAT recognition sequence of a Streptococcus mutans restriction endonuclease.
<3>Acta Microbiol. Hung.
<4>38
<5>43-46
<6>1991
<7>Taking advantage of a mathematical equation applied so far in different fields the calculation
of ATGCAT recognition sequence of restriction endonuclease from Streptococcus mutans serotype
C (SmuCI) is reported together with confirming computer and physical mapping data.

<>

<1>Geck, P., Molnar, A., Nasz, J.
<2>Elimination of non-specific nucleases from restriction endonuclease preparations by different binding on free DNA ligand.
<3>Acta Microbiol. Hung.
<4>34
<5>241-245
<6>1987
<7>In the purification of a novel restriction endonuclease (an AvaIII
isoschizomer, isolated in this laboratory) standard methods were insufficient
to eliminate non-specific nuclease contaminations.  Taking advantage of the
specific site recognition and binding of the restriction endonuclease on DNAs,
a method is described for the simple extraction of non-specific nucleases.  DNA
substrate without recognizable sites do not bind the restriction endonuclease,
while non-specific nucleases are absorbed to, and eliminated with, the DNA via
gel filtration chromatography under special conditions.

<>

<1>Gee, J.E., Marston, C.K., Sammons, S.A., Burroughs, M.A., Hoffmaster, A.R.
<2>Draft Genome Sequence of Bacillus cereus Strain BcFL2013, a Clinical Isolate Similar to G9241.
<3>Genome Announcements
<4>2
<5>e00469-14
<6>2014
<7>Bacillus cereus strains, such as G9241, causing anthrax-like illnesses have recently been
discovered. We report the genome sequence of a clinical strain, B.
cereus BcFL2013, which is similar to G9241, recovered from a patient in Florida.

<>

<1>Geese, W.J., Kwon, Y.K., Wen, X.P., Waring, R.B.
<2>In vitro analysis of the relationship between endonuclease and maturase activities in the bi-functional group I intron-encoded protein, I-AniI.
<3>Eur. J. Biochem.
<4>270
<5>1543-1554
<6>2003
<7>The AnCOB group I intron from Aspergillus nidulans encodes a homing DNA endonuclease called
I-AniI which also functions as a maturase, assisting
in AnCOB intron RNA splicing. In this investigation we biochemically
characterized the endonuclease activity of I-AniI in vitro and utilized
competition assays to probe the relationship between the RNA- and
DNA-binding sites. Despite functioning as an RNA maturase, I-AniI still
retains several characteristic properties of homing endonucleases
including relaxed substrate specificity, DNA cleavage product retention
and instability in the reaction buffer, which suggest that the protein has
not undergone dramatic structural adaptations to function as an
RNA-binding protein. Nitrocellulose filter binding and kinetic burst
assays showed that both nucleic acids bind I-AniI with the same 1 : 1
stoichiometry. Furthermore, in vitro competition activity assays revealed
that the RNA substrate, when prebound to I-AniI, stoichiometrically
inhibits DNA cleavage activity, yet in reciprocal experiments, saturating
amounts of prebound DNA substrate fails to inhibit RNA splicing activity.
The data suggest therefore that both nucleic acids do not bind the same
single binding site, rather that I-AniI appears to contain two binding
sites.

<>

<1>Geese, W.J., Waring, R.B.
<2>A comprehensive characterization of a group IB intron and its encoded maturase reveals that protein-assisted splicing requires an almost intact intron RNA.
<3>J. Mol. Biol.
<4>308
<5>609-622
<6>2001
<7>The group I intron (AnCOB) of the mitochondrial apocytochrome b gene from Aspergillus nidulans
encodes a bi-functional maturase protein that is also a DNA endonuclease. Although the AnCOB
intron self-splices, the encoded maturase protein greatly facilitates splicing, in part, by
stabilizing RNA tertiary structure. To determine their role in self-splicing and in
protein-assisted splicing, several peripheral RNA sub-domains in the 313 nucleotide intron
were deleted (P2, P9, P9.1) or truncated (P5ab, P6a). The sequence in two helices (P2 and P9)
was also inverted. Except for P9, the deleted regions are not highly conserved among group I
introns and are often dispensable for catalytic activity. Nevertheless, despite the very tight
binding of AnCOB RNA to the maturase and the high activity of the bimolecular complex (the
rate of 5' splice-site cleavage was >20 min(-1) with guanosine as the cofactor), the intron
was surprisingly sensitive to these modifications. Several mutations inactivated splicing
completely and virtually all impaired splicing to varying degrees. Mutants containing
comparatively small deletions in various regions of the intron significantly decreased binding
affinity (generally >10(4)-fold), indicating that none of the domains that remained
constitutes the primary recognition site of the maturase. The data argue that tight binding
requires tertiary interactions that can be maintained by only a relatively intact intron RNA,
and that the binding mechanism of the maturase differs from those of two other
well-characterized group I intron splicing factors, CYT-18 and Cpb2. A model is proposed in
which the protein promotes widespread cooperative folding of an RNA lacking extensive initial
tertiary structure.

<>

<1>Gefter, M., Hausmann, R., Gold, M., Hurwitz, J.
<2>The enzymatic methylation of ribonucleic acid and deoxyribonucleic acid.
<3>J. Biol. Chem.
<4>241
<5>1995-2006
<6>1966
<7>An enzyme activity which leads to the degradation of S-adenosylmethionine appears after T3
phage infection of Escherichia coli B.  The purification and properties of this activity have
been described.  The enzyme catalyzes the conversion of S-adenosylmethionine to
thiomethyladenosine and homoserine.  It has been suggested that a gamma-amino-butyrolactone is
an intermediate in this reaction.  The enzyme appears only after infection with T3 phage.
Infection with phages T1, T2, T4, T5, T6, T7, or lambda does not result in the appearance of
any activity.  The rate of enzyme formation and the requirement for protein synthesis de novo
suggests that this activity is similar to early enzymes formed after infection with the T-even
phages.  The enzyme has been isolated both from normal phage T3-infected E. coli and from
ultraviolet light-inactivated phage T3-infected E. coli.  In the later case, the specific
activity of cell-free extracts was several-fold higher than extracts obtained from normal
phage T3-infected cells.  Infection of e. coli with ultraviolet light-inactivated phage T3
does not result in the cessation of host cell deoxyribonucleic or ribonucleic acid synthesis.
Thus, newly synthesized DNA and RNA formed after infection with phage T3 are devoid of
methylated bases.  Phage T2, grown in ultraviolet light-inactivated, phage T3-infected E.
coli, is likewise devoid of methylated bases.  Such methyl-deficient T2 phage appears to be
normal in its biological properties so far examined.

<>

<1>Geier, G.E., Modrich, P.
<2>Recognition sequence of the dam methylase of Escherichia coli K12 and mode of cleavage of DpnI endonuclease.
<3>J. Biol. Chem.
<4>254
<5>1408-1413
<6>1979
<7>The recognition sequence for the dam methylase of Escherichia coli K12 has been determined
directly by use of in vivo methylated ColE1 DNA or DNA methylated in vitro with purified
enzyme. The methylase recognizes the symmetric tetranucleotide d(pG-A-T-C) and introduces two
methyl groups per site in duplex DNA with the product of methylation being
6-methylaminopurine. This work has also demonstrated that DpnI restriction endonuclease
cleaves on the 3' side of the modified adenine within the methylated sequence to yield DNA
fragments possessing fully base-paired termini. All sequences in ColE1 DNA methylated by the
dam enzyme are subject to double strand cleavage by DpnI endonuclease. Therefore, this
restriction enzyme can be employed for mapping the location of sequences possessing the dam
modification.

<>

<1>Geiger, R., Ruter, T., Alves, J., Fliess, A., Wolfes, H., Pingoud, V., Urbanke, C., Maass, G., Pingoud, A., Dusterhoft, A., Kroger, M.
<2>Genetic engineering of EcoRI mutants with altered amino acid residues in the DNA binding site:  Physiochemical investigations give evidence for an altered monomer/dimer equilibrium for the Gln144Lys145 and Gln144Lys145Lys200 mutants.
<3>Biochemistry
<4>28
<5>2667-2677
<6>1989
<7>We have genetically engineered the Arg200 ->Lys mutant, the Glu144Arg145
->GlnLys double mutant, and the Glu144Arg145Arg200 ->GlnLysLys triple mutant of
the EcoRI endonuclease in extension of previously published work on
site-directed mutagenesis of the EcoRI endonuclease in which Glu144 had been
exchanged for Gln and Arg145 for Lys [Wolfes et al. (1986) Nucleic Acids Res.
14, 9063].  All these mutants carry modifications in the DNA binding site.
Mutant EcoRI proteins were purified to homogeneity and characterized by
physiochemical techniques.  All mutants have a very similar secondary structure
composition.  However, whereas the Lys200 mutant is not impaired in its
capacity to form a dimer, the Gln144Lys145 and Gln144Lys145Lys200 mutants have
a very much decreased propensity to form a dimer or tetramer depending on
concentration as shown by gel filtration and analytical ultracentrifugation.
This finding may explain the results of isoelectric focusing experiments which
show that these two mutants have a considerably more basic pI than expected for
a protein in which an acidic amino acid was replaced by a neutral one.
Furthermore, while wild-type EcoRI and the Lys200 mutant are denatured in an
irreversible manner upon heating to 60C, the thermal denaturation process as
shown by circular dichroism spectroscopy is fully reversible with the
Gln144Lys145 double mutant and the Gln144Lys145Lys200 triple mutant.  All EcoRI
endonuclease mutants described here have a residual enzymatic activity with
wild-type specificity, since Escherichia coli cells overexpressing the mutant
proteins can only survive in the presence of EcoRI methylase.  The detailed
analysis of the enzymatic activity and specificity of the purified mutant
proteins is the subject of the accompanying paper.

<>

<1>Geiman, T.M., Sankpal, U.T., Robertson, A.K., Chen, Y., Mazumdar, M., Heale, J., Schmiesing, J.A., Kim, W., Yokomori, K., Zhao, Y., Robertson, K.D.
<2>Isolation and characterization of a novel DNA methyltransferase complex linking DNMT3B with components of the mitotic chromosome condensation machinery.
<3>Nucleic Acids Res.
<4>32
<5>2716-2729
<6>2004
<7>Proper patterns of genome-wide DNA methylation, mediated by DNA methyltransferases DNMT1, -3A
and -3B, are essential for embryonic development and genomic stability in mammalian cells. The
de novo DNA methyltransferase DNMT3B is of particular interest because it is frequently
overexpressed in tumor cells and is mutated in immunodeficiency, centromere instability and
facial anomalies (ICF) syndrome. In order to gain a better understanding of DNMT3B, in terms
of the targeting of its methylation activity and its role in genome stability, we
biochemically purified endogenous DNMT3B from HeLa cells. DNMT3B co-purifies and interacts,
both in vivo and in vitro, with several components of the condensin complex (hCAP-C, hCAP-E
and hCAP-G) and KIF4A. Condensin mediates genome-wide chromosome condensation at the onset of
mitosis and is critical for proper segregation of sister chromatids. KIF4A is proposed to be a
motor protein carrying DNA as cargo. DNMT3B also interacts with histone deacetylase 1 (HDAC1),
the co-repressor SIN3A and the ATP-dependent chromatin remodeling enzyme hSNF2H. Further more,
DNMT3B co-localizes with condensin and KIF4A on condensed chromosomes throughout mitosis.
These studies therefore reveal the first direct link between the machineries regulating DNA
methylation and mitotic chromosome condensation in mammalian cells.

<>

<1>Geiman, T.M., Sankpal, U.T., Robertson, A.K., Zhao, Y.X., Zhao, Y.M., Robertson, K.D.
<2>DNMT3B interacts with hSNF2H chromatin remodeling enzyme, HDACs 1 and 2, and components of the histone methylation system.
<3>Biochem. Biophys. Res. Commun.
<4>318
<5>544-555
<6>2004
<7>The non-random pattern of genome-wide DNA methylation in mammalian cells is established and
maintained by DNA methyltransferases DNMT1,
3A, and 3B. De novo DNA methyltransferase DNMT3B is critical for
embryonic development and is mutated in ICF syndrome. Despite its
importance in normal cellular functioning, little is known about how
DNMT3B operates in the context of chromatin. Here we demonstrate that
DNMT3B associates with four chromatin-associated enzymatic activities
common to transcriptionally repressed, heterochromatic regions of the
genome: DNA methyltransferase, histone deacetylase, ATPase, and histone
methylase activities. By immunoprecipitation and GST pull-down, we show
that DNMT3B interacts with HDAC1, HDAC2, HP1 proteins, Suv39h1, and the
ATP-dependent chromatin remodeling enzyme hSNF2H. Endogenous hSNF2H is
also associated with DNA methyltransferase activity. These proteins
co-localize extensively with DNMT3B in heterochromatic regions. Our
results therefore link DNMT3B to three other components of the
epigenetic machinery and provide important insights into how DNA
methylation patterns may be established within the chromatin
environment.

<>

<1>Geis, A., El Demerdash, H.A., Heller, K.J.
<2>Sequence analysis and characterization of plasmids from Streptococcus thermophilus.
<3>Plasmid
<4>50
<5>53-69
<6>2003
<7>The nucleotide sequences of eight plasmids isolated from seven Streptococcus thermophilus
strains have been determined. Plasmids pSt04,
pER1-1, and pJ34 are related and replicate via a rolling circle mechanism.
Plasmid pJ34 encodes for a replication initiation protein (RepA) and a
small polypeptide with unknown function. Plasmids pSt04 and pER1-1 carry
in addition to repA genes coding for small heat shock proteins (sHsp).
Expression of these proteins is induced at elevated temperatures or low pH
and increases the thermo- and acid resistance. Plasmids pER1-2 and pSt22-2
show identical sequences with five putative open reading frames (ORFs).
The gene products of ORF1 and ORF4 reveal some similarities to transposon
encoded proteins of Bacillus subtilis and Tn916. ORF1 of plasmid pSt106
encodes a protein similar to resolvases of different Gram-positive
bacteria. Integrity of ORF2 and 3, encoding a putative DNA primase and a
replication protein, is essential for replication. ORF1 to 3 of plasmid
pSt08, which are organized in a tricistronic operon, encode a RepA
protein, an adenosine-specific methyltransferase, and a type II
restriction endonuclease. Another type II restriction-modification (R/M)
system is encoded on plasmid pSt0 which is highly similar to those encoded
on lactococcal plasmid pHW393 and B. subtilis plasmid pXH13. Plasmid-free
derivatives of strains St0 and St08 show increased phage sensitivity,
indicating that in the wild-type strains the R/M systems are functionally
expressed. Recombinant plasmids based on the replicons of plasmids pSt04,
pJ34, pSt106, pSt08, and pSt0, are able to replicate in Lactococcus lactis
and B. subtilis, respectively, whereas constructs carrying pER1-2 only
replicate in S. thermophilus.

<>

<1>Gelfand, M.S., Koonin, E.V.
<2>Avoidance of palindromic words in bacterial and archaeal genomes: a close connection with restriction enzymes.
<3>Nucleic Acids Res.
<4>25
<5>2430-2439
<6>1997
<7>Short palindromic sequences (4, 5 and 6 bp palindromes) are avoided at a statistically
significant level in the genomes of several bacteria, including the completely sequenced
Haemophilus influenzae and Synechocystis sp. genomes and in the complete genome of the
archaeon Methanococcus jannaschii.  In contrast, there is only moderate avoidance of
palindromes in the small genome of the bacterium Mycoplasma genitalium and no detectable
avoidance in the genomes of chloroplasts and mitochondria.  The sites for type II
restriction-modification enzymes detected in the given species tend to be among the most
avoided palindromes in a particular genome, indicating a direct connection between the
avoidance of short oligonucleotide words and restriction-modification systems with the
respective specificity.  Palindromes corresponding to sites for restriction enzymes from other
species are also avoided, albeit less significantly, suggesting that in the course of
evolution bacterial DNA has been exposed to a wide spectrum of restriction enzymes, probably
as the result of lateral transfer mediated by mobile genetic elements, such as plasmids and
prophages.  Palindromic words appear to accumulate in DNA once it becomes isolated from
restriction-modification systems, as demonstrated by the case of organellar genomes.  By
combining these observations with protein sequence analysis, we show that the most avoided
4-palindrome and the most avoided 6-palindrome in the archaeon M. jannaschii are likely to be
recognition sites for two novel restriction-modification systems.

<>

<1>Gelinas, R.E., Myers, P.A., Roberts, R.J.
<2>Two sequence-specific endonucleases from Moraxella bovis.
<3>J. Mol. Biol.
<4>114
<5>169-179
<6>1977
<7>Two new sequence-specific endodeoxyribonucleases have been partially purified from Moraxella
bovis. These restriction-like enzymes, MboI and MboII, each cleave bacteriophage lambda DNA
and adenovirus-2 DNA at more than 50 sites. MboI recognizes the sequence 5'^G-A-T-C 3' 3'
C-T-A-T^5' and cleaves at the sites indicated by the arrows. A specific endonuclease, MosI,
has also been purified from Moraxella osloenis and recognizes the same sequence as MboI.

<>

<1>Gelinas, R.E., Myers, P.A., Weiss, G.H., Roberts, R.J., Murray, K.
<2>A specific endonuclease from Brevibacterium albidum.
<3>J. Mol. Biol.
<4>114
<5>433-440
<6>1977
<7>A new restriction-like endonuclease, BalI, has been partially purified from
Brevibacterium albidum.  This enzyme cleaves bacteriophage lambda DNA at least
18 times and adenovirus-2 DNA at least 16 times, but does not cleave simian
virus 40 DNA.  All sites cleaved by BalI are also cut by the specific
endonuclease HaeIII from Haemophilus aegyptius.  The recognition sequence of
BalI is 5'-T-G-G ^ C-C-A-3' 3'-A-C-C ^ G-G-T-5' and the cleavage site is
indicated by the arrows.

<>

<1>Gemmen, G.J., Millin, R., Smith, D.E.
<2>DNA looping by two-site restriction endonucleases: heterogeneous probability distributions for loop size and unbinding force.
<3>Nucleic Acids Res.
<4>34
<5>2864-2877
<6>2006
<7>Proteins interacting at multiple sites on DNA via looping play an important role in many
fundamental biochemical processes. Restriction
endonucleases that must bind at two recognition sites for efficient
activity are a useful model system for studying such interactions. Here we
used single DNA manipulation to study sixteen known or suspected two-site
endonucleases. In eleven cases (BpmI, BsgI, BspMI, Cfr10I, Eco57I, EcoRII,
FokI, HpaII, NarI, Sau3AI and SgrAI) we found that substitution of Ca2+
for Mg2+ blocked cleavage and enabled us to observe stable DNA looping.
Forced disruption of these loops allowed us to measure the frequency of
looping and probability distributions for loop size and unbinding force
for each enzyme. In four cases we observed bimodal unbinding force
distributions, indicating conformational heterogeneity and/or complex
binding energy landscapes. Measured unlooping events ranged in size from 7
to 7500 bp and the most probable size ranged from less than 75 bp to
nearly 500 bp, depending on the enzyme. In most cases the size
distributions were in much closer agreement with theoretical models that
postulate sharp DNA kinking than with classical models of DNA elasticity.
Our findings indicate that DNA looping is highly variable depending on the
specific protein and does not depend solely on the mechanical properties
of DNA.

<>

<1>Gemmen, G.J., Millin, R., Smith, D.E.
<2>Tension-dependent DNA cleavage by restriction endonucleases: Two-site enzymes are "switched off" at low force.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>11555-11560
<6>2006
<7>DNA looping occurs in many important protein-DNA interactions, including those regulating
replication, transcription, and recombination. Recent
theoretical studies predict that tension of only a few piconewtons acting
on DNA would almost completely inhibit DNA looping. Here, we study
restriction endonucleases that require interaction at two separated sites
for efficient cleavage. Using optical tweezers we measured the dependence
of cleavage activity on DNA tension with 15 known or suspected two-site
enzymes (BfiI, BpmI, BsgI, BspMI, Cfr9I, Cfr10I, Eco57I, EcoRII, FokI,
HpaII, MboII, NarI, SacII, Sau3AI, and SgrAI) and six one-site enzymes
(BamHI, EcoRI, EcoRV, HaeIII, HindIII, and DNaseI). All of the one-site
enzymes were virtually unaffected by 5 pN of tension, whereas all of the
two-site enzymes were completely inhibited. These enzymes thus constitute
a remarkable example of a tension sensing "molecular switch." A detailed
study of one enzyme, Sau3AI, indicated that the activity decreased
exponentially with tension and the decrease was approximately 10-fold at
0.7 pN. At higher forces ( approximately 20-40 pN) cleavage by the
one-site enzymes EcoRV and HaeIII was partly inhibited and cleavage by
HindIII was enhanced, whereas BamHI, EcoRI, and DNaseI were largely
unaffected. These findings correlate with structural data showing that
EcoRV bends DNA sharply, whereas BamHI, EcoRI, and DNaseI do not. Thus,
DNA-directed enzyme activity involving either DNA looping or bending can
be modulated by tension, a mechanism that could facilitate mechanosensory
transduction in vivo.

<>

<1>Gemmen, G.J., Millin, R., Smith, D.E.
<2>Dynamics of single DNA looping and cleavage by Sau3AI and effect of tension applied to the DNA.
<3>Biophys. J.
<4>91
<5>4154-4165
<6>2006
<7>Looping and cleavage of single DNA molecules by the two-site restriction endonuclease Sau3AI
were measured with optical tweezers. A DNA template
containing many recognition sites was used, permitting loop sizes from
approximately 10 to 10,000 basepairs. At high enzyme concentration,
cleavage events were detected within 5 s and nearly all molecules were
cleaved within 5 min. Activity decreased approximately 10-fold as the DNA
tension was increased from 0.03 to 0.7 pN. Substituting Ca(2+) for Mg(2+)
blocked cleavage, permitting measurement of stable loops. At low tension,
the initial rates of cleavage and looping were similar (approximately
0.025 s(-1) at 0.1 pN), suggesting that looping is rate limiting. Short
loops formed more rapidly than long loops. The optimum size decreased from
approximately 250 to 45 basepairs and the average number of loops (in 1
min) from 4.2 to 0.75 as tension was increased from 0.03 to 0.7 pN. No
looping was detected at 5 pN. These findings are in qualitative agreement
with recent theoretical predictions considering only DNA mechanics, but we
observed weaker suppression with tension and smaller loop sizes. Our
results suggest that the span and elasticity of the protein complex,
nesting of loops, and protein-induced DNA bending and wrapping play an
important role.

<>

<1>Geng, J., Chiu, C.H., Tang, P., Chen, Y., Shieh, H.R., Hu, S., Chen, Y.Y.
<2>Complete Genome and Transcriptomes of Streptococcus parasanguinis FW213: Phylogenic Relations and Potential Virulence Mechanisms.
<3>PLoS ONE
<4>7
<5>E34769
<6>2012
<7>Streptococcus parasanguinis, a primary colonizer of the tooth surface, is also an
opportunistic pathogen for subacute endocarditis. The complete genome of strain
FW213 was determined using the traditional shotgun sequencing approach and
further refined by the transcriptomes of cells in early exponential and early
stationary growth phases in this study. The transcriptomes also discovered 10
transcripts encoding known hypothetical proteins, one pseudogene, five
transcripts matched to the Rfam and additional 87 putative small RNAs within the
intergenic regions defined by the GLIMMER analysis. The genome contains five
acquired genomic islands (GIs) encoding proteins which potentially contribute to
the overall pathogenic capacity and fitness of this microbe. The differential
expression of the GIs and various open reading frames outside the GIs at the two
growth phases suggested that FW213 possess a range of mechanisms to avoid host
immune clearance, to colonize host tissues, to survive within oral biofilms and
to overcome various environmental insults. Furthermore, the comparative genome
analysis of five S. parasanguinis strains indicates that albeit S. parasanguinis
strains are highly conserved, variations in the genome content exist. These
variations may reflect differences in pathogenic potential between the strains.

<>

<1>Geng, J., Huang, S.C., Li, S., Hu, S., Chen, Y.Y.
<2>Complete Genome Sequence of the Ureolytic Streptococcus salivarius Strain 57.I.
<3>J. Bacteriol.
<4>193
<5>5596-5597
<6>2011
<7>Streptococcus salivarius 57.I is one of the most abundant and highly ureolytic bacteria in the
human mouth. It can utilize urea as the sole
nitrogen source via the activity of urease. Complete genome sequencing of
S. salivarius 57.I revealed a chromosome and a phage which are absent in
strain SK126.

<>

<1>Geng, W., Cao, M., Song, C., Xie, H., Liu, L., Yang, C., Feng, J., Zhang, W., Jin, Y., Du, Y., Wang, S.
<2>Complete genome sequence of Bacillus amyloliquefaciens LL3, which exhibits glutamic acid-independent production of poly-{gamma}-glutamic acid.
<3>J. Bacteriol.
<4>193
<5>3393-3394
<6>2011
<7>Bacillus amyloliquefaciens is one of most prevalent Gram-positive aerobic spore-forming
bacteria with the ability of synthesizing polysaccharides
and polypeptides. Here, we report the complete genome sequence of B.
amyloliquefaciens LL3, which was isolated from fermented food and presents
the glutamic acid-independent production of poly-gamma-glutamic acid.

<>

<1>Genger, R.K., Kovac, K.A., Dennis, E.S., Peacock, W.J., Finnegan, E.J.
<2>Multiple DNA methyltransferase genes in Arabidopsis thaliana.
<3>Plant Mol. Biol.
<4>41
<5>269-278
<6>1999
<7>Methylation of plant DNA occurs at cytosines in any sequence context, and as the Arabidopsis
methyltransferase, METI, preferentially methylates cytosines in CG dinucleotides, it is likely
that Arabidopsis has other methyltransferases with different target specificities. We have
identified five additional genes encoding putative DNA methyltransferases. Three of these
genes are very similar to METI throughout the coding region; these genes probably arose by a
series of gene duplication events, the most recent giving rise to METIIa and METIIb. METIIa
and b are expressed at low levels in vegetative and floral organs and the level of transcripts
is not affected by the introduction of a METI antisense transgene, nor do the METII enzymes
substitute for the reduced activity of METI in methylating CG dinucleotides. METIII is not
essential as it encodes a truncated protein. Two other genes encode a second class of DNA
methyltransferase with the conserved motifs characteristic of cytosine methyltransferases, but
with little homology to the METI-like methyltransferases through the remainder of the protein.
These two methyltransferases are characterized by the presence of a chromodomain inserted
within the methyltransferase domain, suggesting that they may be associated with
heterochromatin. Both these genes are transcribed at low levels in vegetative and reproductive
tissues.

<>

<1>Gentzbittel, L., Nicolas, P.
<2>A basic program to construct evolutionary trees from restriction endonuclease data.
<3>J. Hered.
<4>80
<5>254
<6>1989
<7>None

<>

<1>Gentzbittel, L., Nicolas, P.
<2>Improvement of "A BASIC program to construct evolutionary trees from restriction endonuclease data" with the use of PASCAL.
<3>J. Hered.
<4>81
<5>491-492
<6>1990
<7>None

<>

<1>George, J., Blakesley, R.W., Chirikjian, J.G.
<2>Sequence-specific endonuclease BamHI.
<3>J. Biol. Chem.
<4>255
<5>6521-6524
<6>1980
<7>The specificity of cleavage of BamHI is altered in the presence of hydrophobic
reagents, such as glycerol and M2SO.  The enzyme with altered specificity,
designated BamHI.1, generated digestion patterns of various DNAs, which were
distinct from those generated by BamHI.  Cleavage sites recognized in PhiX174
RF DNA in the presence of these hydrophobic reagents are not related to the
BamHI palindrome.  BamHI.1 appears to be an endogenous form of BamHI that can
be expressed by altering the hydrophobicity of the reaction.

<>

<1>George, J., Chirikjian, J.G.
<2>Biospecific fractionation matrices for sequence specific endonucleases.
<3>Nucleic Acids Res.
<4>5
<5>2223-2232
<6>1978
<7>Fractionation of several type II specific restriction endonucleases was
achieved by separation on two novel biospecific matrices.  The matrices are
pyran, a copolymer of divinyl ether of maleic anhydride, and Cibacron Blue
F3Ga, a blue dye commonly used for the calibration of molecular sieves.  Both
compounds are insolubilized by coupling to sepharose through a cyanogen bromide
linkage and in their soluable form inhibit the restriction endonucleases which
we have tested.  These affinity matrices can be used to obtain restriction
endonucleases from crude extracts after removal of nucleic acids.  They have
also proven to have a high capacity when used as subsequent step in enzyme
purification.  Their additional advantage is the rapid development time and
reusability of columns packed with the two matrices.

<>

<1>George, J., Chirikjian, J.G.
<2>Sequence-specific endonuclease BamHI: Relaxation of sequence recognition.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>79
<5>2432-2436
<6>1982
<7>The effect of glycerol on the specificity of DNA cleavage by the restriction
endonuclease BamHI has been examined.  In addition to the canonical G ^ G-A-T-C
site, BamHI cuts DNA at several sites that we have named noncanonical BamHI.1
sites.  The number of BamHI.1 sites is simian virus 40 and pBR322 was
determined to be 13 for each DNA.  Cutting sites determined by DNA sequence
analysis include G ^ G-A-A-C-C, G ^ G-C-T-C-C, G ^ G-G-T-C-C, and G-A-A-T-C-C
with the complementary strand sequence assignments of G-G-T-T-C-C, G-G-A-G-C-C,
G-G-A-C-C-C, and G-G-A-T-T-C.  The relaxation in specificity was related to
hydrogen bond acceptor and donor sites in the recognition sequence, in an
attempt to generate a model of BamHI recognition of cognate sites in DNA.

<>

<1>George, J., Hamada, Y.-Y.T., Chirikjian, J.G.
<2>Chemical modifications as structural probes for the recognition of DNA palindromes by BamHI and BglI.
<3>Fed. Proc.
<4>40
<5>1848
<6>1981
<7>BamHI and BglI have been previously purified to apparent homogeneity in our
laboratory.  The catalytically active form of BamHI is a dimer comprised of two
apparently identical subunits of 23,000 daltons.  In comparison the active form
of BglI was isolated as a single polypeptide of 34,000 daltons.  Chemical
modification of the enzyme and both oligonucleotide and plasmid DNA substrates
have been undertaken.  Enzyme which has been reductively alkylated (lysine
modification) with formaldehyde is inactivated.  In contrast when such
modifications are performed in the presence of a DNA substrate enzymatic
activity is maintained.  Likewise DNAs that contain no recognition sequences
for BamHI confer little protection against activity losses.  This suggests that
upon specific binding to DNA, at least one or a group of lysine residues, are
protected from chemical modification. BamHI recognizes the palindromic sequence
G^GATCC.  We have evidence to suggest that the information required for
recognition specificity resides in the major groove of the DNA double helix.
This recognition may involve the N6 position of adenine and the 04 position of
thymine.  The involvement of N7 and N2 of guanine and N3 of adenine are under
curretn investigation by chemical modification procedures.

<>

<1>George, J., Nardone, G., Chirikjian, J.G.
<2>Sequence-specific BamHI endonuclease.
<3>J. Biol. Chem.
<4>260
<5>14387-14392
<6>1985
<7>Arginyl residues in BamHI endonuclease were examined because of their alleged
role in proteins that contain nucleotide- or phosphate-binding sites.
Butanedione, an arginine-specific reagent, inhibited the endonuclease in the
presence of sodium borate.  The inhibition was decreased by preliminary
incubation of the enzyme with DNA or competitive inhibitors which were the
5'-phosphoryl deoxydinucleotide subsets of the BamHI recognition sequence.  The
dinucleotide pdGpdG protected the enzyme most efficiently against the
butanedione modification.  Dinucleotides that were unrelated to the recognition
sequence failed to protect the enzyme from inactivation.  These studies
indicate that arginine residues may reside in the enzyme's active site and
might function in the sequence-specific recognition of the BamHI palindrome.

<>

<1>Gerasimaite, R., Klimasauskas, S.
<2>Novel approaches to engineering sequence-specificity of DNA cytosine-5 methyltransferases.
<3>FEBS J.
<4>274
<5>259
<6>2007
<7>Bacterial DNA cytosine-5 methyltransferases (C5-MTases) recognize
2-8 bp DNA sequences and transfer the methyl group from
the cofactor S-adenosyl-L-methionine onto the C5 position of a
cytosine residue in that particular target sequence. Besides their
utmost biological importance, DNA MTases were shown to be
useful tools for site-specific DNA labelling [1] and for the construction
of bionanodevices [2]. Here we describe the conversion of the
HhaI methyltransferase (M.HhaI), which recognizes the palindromic
GCGC site, into a GCG-specific or CGC-specific MTases.
Due to a non-palindromic nature of the new targets, such engineered
MTases could be used for creating hemimethylated CpG sites
in DNA, which are desired models for mechanistic studies of the
maintenance of genomic methylation patterns in higher eukaryotes
[3]. Numerous crystal structures available for M.HhaI show that
the recognition of the GCGC target is accomplished by two recognition
loops. Directed evolution and rational design approaches
were employed to obtain variants with the novel substrate specificities.
Interestingly, one of the most efficient MTase variants contains
a mutation outside the recognition loops, which allows the
formation of a non-specific contact to the phosphodiester backbone
of the DNA. Our results thus demonstrate that new asymmetric
target specificities can be created by alterations of the
recognition elements in DNA MTases, and that de novo non-specific
contacts can compensate for the loss of base-specific DNA
interactions.
References
1. Lukinavicius, G, et al. (2007). J Am Chem Soc, DOI: 10.1021/
ja0691876.
2. Singer, E, et al. (2006). Nano Lett, 6(6): 1184.
3. Vilkaitis, G, et al. (2005). J Biol Chem, 280(1): 64.

<>

<1>Gerasimaite, R., Merkiene, E., Klimasauskas, S.
<2>Direct observation of cytosine flipping and covalent catalysis in a DNA methyltransferase.
<3>Nucleic Acids Res.
<4>39
<5>3771-3780
<6>2011
<7>Methylation of the five position of cytosine in DNA plays important roles in epigenetic
regulation in diverse organisms including humans. The transfer of methyl groups from the
cofactor S-adenosyl-l-methionine is carried out by methyltransferase enzymes. Using the
paradigm bacterial methyltransferase M.HhaI we demonstrate, in a chemically unperturbed
system, the first direct real-time analysis of the key mechanistic events-the flipping of the
target cytosine base and its covalent activation; these changes were followed by monitoring
the hyperchromicity in the DNA and the loss of the cytosine chromophore in the target
nucleotide, respectively. Combined with studies of M.HhaI variants containing redesigned
tryptophan fluorophores, we find that the target base flipping and the closure of the mobile
catalytic loop occur simultaneously, and the rate of this concerted motion inversely
correlates with the stability of the target base pair. Subsequently, the covalent activation
of the target cytosine is closely followed by but is not coincident with the methyl group
transfer from the bound cofactor. These findings provide new insights into the temporal
mechanism of this physiologically important reaction and pave the way to in-depth studies of
other base-flipping systems.

<>

<1>Gerasimaite, R., Vilkaitis, G., Klimasauskas, S.
<2>A directed evolution design of a GCG-specific DNA hemimethylase.
<3>Nucleic Acids Res.
<4>37
<5>7332-7341
<6>2009
<7>DNA cytosine-5 methyltransferases (C5-MTases) are valuable models to study sequence-specific
modification of DNA and are becoming increasingly
important tools for biotechnology. Here we describe a structure-guided
rational protein design combined with random mutagenesis and selection to
change the specificity of the HhaI C5-MTase from GCGC to GCG. The
specificity change was brought about by a five-residue deletion and
introduction of two arginine residues within and nearby one of the target
recognizing loops. DNA protection assays, bisulfite sequencing and enzyme
kinetics showed that the best selected variant is comparable to wild-type
M.HhaI in terms of sequence fidelity and methylation efficiency, and
supersedes the parent enzyme in transalkylation of DNA using synthetic
cofactor analogs. The designed C5-MTase can be used to produce
hemimethylated CpG sites in DNA, which are valuable substrates for studies
of mammalian maintenance MTases.

<>

<1>Germann, M.W., Kalisch, B.W., Lundberg, P., Vogel, H.J., van de Sande, J.H.
<2>Perturbation of DNA hairpins containing the EcoRI recognition site by hairpin loops of varying size and composition: physical (NMR and UV) and enzymatic (EcoRI) studies.
<3>Nucleic Acids Res.
<4>18
<5>1489-1498
<6>1990
<7>We have investigated loop-induced structural perturbation of the stem structure
in hairpins d(GAATTCXnGAATTC) (X = A, T, and n = 3, 4, 5 and 6) that contain an
EcoRI restriction site in close proximity to the hairpin loop.
Oligonucleotides containing either a T3 or an A3 loop were not hydrolyzed by
the restriction enzyme and also showed only weak binding to EcoRI in the
absence of the cofactor Mg2+.  In contrast, hairpins with larger loops are
hydrolyzed by the enzyme at the scission site next to the loop although the
substrate with a A4 loop is significantly more resistant than the
oligonucleotide containing a T4 loop.  The hairpin structures with 3 loop
residues were found to be thermally most stable while larger hairpin loops
resulted in structures with lower melting temperatures.  The T-loop hairpins
are thermally more stable than the hairpins containing the same number of A
residues in the loop.  As judged from proton NMR spectroscopy and the
thermodynamic data, the base pair closest to the hairpin loop did form in all
cases studied.  The hairpin loops did, however, affect the conformation of the
stem structure of the hairpins.  From 31P and 1H NMR spectroscopy we conclude
that the perturbation of the stem structure is stronger for smaller hairpin
loops and that the extent of the perturbation is limited to 2 - 3 base pairs
for hairpins with T3 or A4 loops.  Our results demonstrate that hairpin loops
modulate the conformation of the stem residues close to the loop and that this
in turn reduces the substrate activity for DNA sequence specific proteins.

<>

<1>Germann, M.W., Kalisch, B.W., Varnum, J.M., Vogel, H.J., van de Sande, J.H.
<2>NMR spectroscopic and enzymatic studies of DNA hairpins containing mismatches in the EcoRI recognition site.
<3>Biochem. Cell Biol.
<4>76
<5>391-402
<6>1998
<7>We have correlated the structural perturbations caused by DNA mismatches with the enzymatic
data of the interaction of the restriction endonuclease EcoRI with DNA. Oligonucleotides
d(CGAGAATTCTCA5GAXAATTCT) (X = G, A, T) and d(CGCGAATTYGCGT4CGCXAATTCGCG) (Y = C, X = G, T and
Y = A, X = T) containing single mismatches within the EcoRI recognition site were
characterized by NMR spectroscopy and by their EcoRI substrate properties. UV melting and gel
electrophoresis studies confirm that the oligonucleotides form hairpin structures. The
presence of either a CT or a CA mismatch results in markedly lower Tm and van't Hoff
enthalpies compared with the fully base paired control. NMR imino proton spectra of these
hairpins demonstrate that the perturbation caused by the two mispairs or a noncanonical AT
pair is localized and limited to one or two base pairs on either side of the perturbation. The
DNA hairpin structures containing single mismatches, and to a lesser extent also sequences
with a single noncanonical base pair, are substrates for the restriction endonuclease. In
addition to the strand scission at the nonperturbed GpA phosphodiester bond some cleavage is
observed at the mismatched position. The interactions of the CA and CT mismatched hairpin with
the enzyme are characterized by binding constants that are only 33 and 57 times lower,
respectively, than that for the canonical sequence, corresponding to 8-10 kJ x mol(-1) less
favorable free binding energy. This, taken together with the NMR data, indicates that the CA
and CT mismatches have only small effects on the EcoRI recognition of the DNA substrate. We
conclude that two out of the three hydrogen bonds that characterize the interaction of EcoRI
with the CG base pair in the canonical sequence can still be formed for either the CT or CA
mismatched recognition site.

<>

<1>Gerome, P., Le Fleche, P., Blouin, Y., Scholz, H.C., Thibault, F.M., Raynaud, F., Vergnaud, G., Pourcel, C.
<2>Yersinia pseudotuberculosis ST42 (O:1) Strain Misidentified as Yersinia pestis by Mass Spectrometry Analysis.
<3>Genome Announcements
<4>2
<5>e00435-14
<6>2014
<7>We report here the draft sequence of strain CEB14_0017, alias HIAD_DUP, recovered from a human
patient and initially identified as Yersinia pestis by mass
spectrometry analysis. Genotyping based on tandem repeat polymorphism assigned
the strain to Yersinia pseudotuberculosis sequence type 42 (ST42). The total
assembly length is 4,894,739 bp.

<>

<1>Gerrish, R.S., Gill, A.L., Fowler, V.G., Gill, S.R.
<2>Development of pooled suppression subtractive hybridization to analyze the pangenome of Staphylococcus aureus.
<3>J. Microbiol. Methods
<4>81
<5>56-60
<6>2010
<7>We describe the development and application of a Pooled Suppression
Subtractive Hybridization (PSSH) method to describe differences between
the genomic content of a pool of clinical Staphylococcus aureus isolates
and a sequenced reference strain. In comparative bacterial genomics,
Suppression Subtractive Hybridization (SSH) is normally utilized to
compare genomic features or expression profiles of one strain versus
another, which limits its ability to analyze communities of isolates.
However, a PSSH approach theoretically enables the user to characterize
the entirety of gene content unique to a related group of isolates in a
single reaction. These unique fragments may then be linked to individual
isolates through standard PCR. This method was applied to examine the
genomic diversity found in pools of S.aureus isolates associated with
complicated bacteremia infections leading to endocarditis and
osteomyelitis. Across four pools of 10 isolates each, four hundred and
twenty seven fragments not found in or significantly divergent from the S.
aureus NCTC 8325 reference genome were detected. These fragments could be
linked to individual strains within its pool by PCR. This is the first use
of PSSH to examine the S. aureus pangenome. We propose that PSSH is a
powerful tool for researchers interested in rapidly comparing the genomic
content of multiple unstudied isolates.

<>

<1>Gfeller, K.Y., Roth, M., Melle, L., Teuber, M.
<2>Sequence and genetic organization of the 19.3-kb erythromycin- and dalfopristin-resistance plasmid pLME300 from Lactobacillus fermentum  ROT1.
<3>Plasmid
<4>50
<5>190-201
<6>2003
<7>Lactobacillus fermentum ROT1 was isolated from a raw milk dairy product. It is resistant to
novobiocin, tetracycline, erythromycin and
dalfopristin. A chromosomal tetracycline-resistance determinant was
identified as tetM. A 19,398-bp plasmid (pLME300), present in several
erythromycin-resistant strains of Lb. fermentum, was isolated from strain
ROT1 and completely sequenced. Based on putative open reading frames,
pLME300 contains at least four different functional regions. In region I,
ORF1 shows high homologies to replication proteins of different
theta-replicating plasmids. In addition, a tandem repeat of a 22-bp
sequence appears 4.5 times. In region II, ORF3 may code for a methylase,
and ORF4 has homologies to Mrr restriction system proteins of Deinococcus
radiodurans and Escherichia coli suggesting a restriction-modification
system. Region III harbours antibiotic-resistance genes, coding for a
macrolide-lincosamide-streptogramin B (MLS) methylase Erm(LF) and the
streptogramin A acetyltransferase Vat(E), which is identical to Vat(E)
from Enterococcus faecium. Furthermore, region III shows a 91% nucleotide
sequence identity to an erm-vat linkage of E. faecium. Region IV carries
ORFs that appear to be involved in plasmid mobilization as characterized
by a putative origin of transfer and a mobilization protein. pLME300 is
the largest completely sequenced multi-resistance plasmid isolated from
any Lactobacillus strain so far.

<>

<1>Ghai, R., Martin-Cuadrado, A.B., Molto, A.G., Heredia, I.G., Cabrera, R., Martin, J., Verdu, M., Deschamps, P., Moreira, D., Lopez-Garcia, P., Mira, A., Rodriguez-Valera, F.
<2>Metagenome of the Mediterranean deep chlorophyll maximum studied by direct and fosmid library 454 pyrosequencing.
<3>ISME J.
<4>0
<5>1-13
<6>2010
<7>The deep chlorophyll maximum (DCM) is a zone of maximal photosynthetic activity, generally
located toward the base of the photic zone in lakes and oceans. In the tropical waters, this
is a permanent feature, but in the Mediterranean and other temperate waters, the DCM is a
seasonal phenomenon. The metagenome from a single sample of a mature Mediterranean DCM
community has been 454 pyrosequenced both directly and after cloning in fosmids. This study is
the first to be carried out at this sequencing depth (ca. 600 Mb combining direct and fosmid
sequencing) at any DCM. Our results indicate a microbial community massively dominated by the
high-light-adapted Prochlorococcus marinus subsp. pastoris, Synechococcus sp., and the
heterotroph Candidatus Pelagibacter. The sequences retrieved were remarkably similar to the
existing genome of P. marinus subsp. pastoris with a nucleotide identity over 98%. Besides, we
found a large number of cyanophages that could prey on this microbe, although sequence
conservation was much lower. The high abundance of phage sequences in the cellular size
fraction indicated a remarkably high proportion of cells suffering phage lytic attack. In
addition, several fosmids clearly belonging to Group II Euryarchaeota were retrieved and
recruited many fragments from the total direct DNA sequencing suggesting that this group might
be quite abundant in this habitat. The comparison between the direct and fosmids sequencing
revealed a bias in the fosmid libraries against low-GC DNA and specifically against the two
most dominant members of the community, Candidatus Pelagibacter and P. marinus subsp.
pastoris, thus unexpectedly providing a feasible method to obtain large genomic fragments from
other less prevalent members of this community.

<>

<1>Ghaskadbi, S., Bharathi, S., Modak, S.P.
<2>Inhibition of cleavage by restriction endonucleases due to modifications induced in SV40 DNA by methyl methanesulfonate.
<3>Cell. Mol. Biol. Res.
<4>41
<5>59-66
<6>1995
<7>Methyl methanesulfonate (MMS), a direct mutagen, methylates DNA bases and causes distortions
in DNA structure.  Supercoiled SV40 DNA was treated in vitro with varying concentrations of
MMS from 0.001 mM to 10 mM MMS either for 30 min or 3 h and analyzed by electrophoresis in 1%
neutral and alkaline agarose gels.  The electrophoretic mobility (EPM) of native DNA did not
change after treatment with the mutagen, while alkaline gels revealed low MW DNA fragments due
to single strand breaks at alkali-sensitive sites generated by the action of MMS.  By
two-dimensional electrophoresis, we find that all three native DNA forms contain
alkali-sensitive sites after treatment with MMS.  To examine the effect of base modification
by MMS on DNA-protein interactions, we have used as probes, restriction endonucleases.  These
cleave DNA in a sequence-specific manner, and their activity is dependent upon the methylation
status of the substrate DNA.  We find that cleavage by these restriction endonucleases is
inhibited due to methylation by MMS.

<>

<1>Ghazal, S., Hurst, S.G.I.V., Morris, K., Abebe-Akele, F., Thomas, W.K., Badr, U.M., Hussein, M.A., Abouzaied, M.A., Khalil, K.M., Tisa, L.S.
<2>Draft Genome Sequence of Photorhabdus luminescens Strain BA1, an Entomopathogenic Bacterium Isolated from Nematodes Found in Egypt.
<3>Genome Announcements
<4>2
<5>e00396-14
<6>2014
<7>Photorhabdus luminescens strain BA1 is an entomopathogenic bacterium that forms a symbiotic
association with Heterorhabditis nematodes. We report here a 5.0-Mbp
draft genome sequence for P. luminscens strain BA1, with a G+C content of 42.46%
and 4,250 candidate protein-coding genes.

<>

<1>Ghequire, M.G., Rokni-Zadeh, H., Zarrineh, P., De Mot, R.
<2>Draft Genome Sequence of Pseudomonas fluorescens LMG 5329, a White Line-Inducing  Principle-Producing Bioindicator for the Mushroom Pathogen Pseudomonas tolaasii.
<3>Genome Announcements
<4>1
<5>e00383-13
<6>2013
<7>Pseudomonas tolaasii, the causative agent of Agaricus bisporus brown blotch disease, can be
identified by the white line reaction, occurring upon
confrontation of the tolaasin-producing mushroom pathogen with 'Pseudomonas
reactans,' producing the lipopeptide white line-inducing principle (WLIP). The
draft genome sequence of the WLIP-producing indicator Pseudomonas fluorescens
strain LMG 5329 is reported here.

<>

<1>Ghodhbane-Gtari, F. et al.
<2>Draft Genome Sequence of Frankia sp. Strain CN3, an Atypical, Noninfective (Nod-) Ineffective (Fix-) Isolate from Coriaria nepalensis.
<3>Genome Announcements
<4>1
<5>e0008513
<6>2013
<7>We report here the genome sequence of Frankia sp. strain CN3, which was isolated  from
Coriaria nepalensis. This genome sequence is the first from the fourth
lineage of Frankia, strains of which are unable to reinfect actinorhizal plants.
At 10 Mb, it represents the largest Frankia genome sequenced to date.

<>

<1>Ghodhbane-Gtari, F., Hurst, S.G.I.V., Oshone, R., Morris, K., Abebe-Akele, F., Thomas, W.K., Ktari, A., Salem, K., Gtari, M., Tisa, L.S.
<2>Draft Genome Sequence of Frankia sp. Strain BMG5.23, a Salt-Tolerant Nitrogen-Fixing Actinobacterium Isolated from the Root Nodules of Casuarina  glauca Grown in Tunisia.
<3>Genome Announcements
<4>2
<5>e00520-14
<6>2014
<7>Nitrogen-fixing actinobacteria of the genus Frankia are symbionts of woody dicotyledonous
plants termed actinorhizal plants. We report here a 5.27-Mbp draft genome sequence for Frankia
sp. strain BMG5.23, a salt-tolerant nitrogen-fixing actinobacterium isolated from root nodules
of Casuarina glauca collected in Tunisia.

<>

<1>Gholizadeh, A., Faizi, M.H., Kohnehrouz, B.B.
<2>Induced expression of EcoRI endonuclease as an active maltose-binding fusion protein in Escherichia coli.
<3>Microbiology
<4>79
<5>167-172
<6>2010
<7>Among the numerous bacterial Type II restriction enzymes, EcoRI endonuclease is the most
extensively studied and is widely used in
recombinant DNA technology. Its heterologous overexpression as
recombinant protein has already been studied. However, very limited
information concerning its fused product is available thus far. In the
present study, the EcoRI restriction endonuclease gene was cloned and
expressed as a part of maltose-binding fusion protein under the control
of strong inducible tac promoter in TB1 strain of Escherichia coli
cells. Transformed cells containing pMALc2X-EcoRI recombinant plasmid
were unable to grow under experimental conditions. However, fused EcoRI
protein was purified (with the yield of 0.01 mg/l of bacterial culture)
by affinity chromatography from E. coli cells induced at the late
exponential phase of growth. Restriction quality test revealed that the
purified product could restrict a control plasmid DNA in vitro.

<>

<1>Gholizadeh, A., Kohnehrouz, B.B.
<2>Carborundum-dependent entrance of EcoRI restriction enzyme into plant cells and specific cleavage of genomic DNA.
<3>Indian J. Exp. Biol.
<4>47
<5>684-689
<6>2009
<7>In a basic research to determine the morpho-molecular interactions of plant tissues with EcoRI
DNA restriction enzyme, it was demonstrated
that this protein is capable of entering the sunflower and maize leaf
cells using a plant tissue-abrading material and cleaving the genomic
DNA at specific sites. This was inferred from the analysis of
morphological patterns of EcoRI-treated leaf areas as well as using
some molecular tests, including the cleavage pattern analysis of
genomic DNA isolated from treated locations followed by ligation of
cleaved fragments into EcoRI site of a DNA cloning vector system. The
overall results indicated that the specific restriction of genomic DNA
may happen following the entrance of EcoRI protein most likely into the
nucleus of plant cells.

<>

<1>Ghosh, A., Chaudhary, S.A., Apurva, S.R., Tiwari, T., Gupta, S., Singh, A.K., Katudia, K.H., Patel, M.P., Chikara, S.K.
<2>Whole-Genome Sequencing of Micrococcus luteus Strain Modasa, of Indian Origin.
<3>Genome Announcements
<4>1
<5>e0007613
<6>2013
<7>The hydrocarbon-degrading bacterium Micrococcus luteus strain Modasa was isolated from
contaminated soil from Modasa, North Gujarat, India. Whole-genome sequencing
and analysis provide an insight into the potentially important genes responsible
for bioremediation.

<>

<1>Ghosh, A., Passaris, I., Tesfazgi, M.M., Rocha, S., Vanoirbeek, K., Hofkens, J., Aertsen, A.
<2>Cellular localization and dynamics of the Mrr type IV restriction endonuclease of Escherichia coli.
<3>Nucleic Acids Res.
<4>42
<5>3908-3918
<6>2014
<7>In this study, we examined the intracellular whereabouts of Mrr, a cryptic type IV restriction
endonuclease of Escherichia coli K12, in response to different
conditions. In absence of stimuli triggering its activity, Mrr was found to be
strongly associated with the nucleoid as a number of discrete foci, suggesting
the presence of Mrr hotspots on the chromosome. Previously established elicitors
of Mrr activity, such as exposure to high (hydrostatic) pressure (HP) or
expression of the HhaII methyltransferase, both caused nucleoid condensation and
an unexpected coalescence of Mrr foci. However, although the resulting
Mrr/nucleoid complex was stable when triggered with HhaII, it tended to be only
short-lived when elicited with HP. Moreover, HP-mediated activation of Mrr
typically led to cellular blebbing, suggesting a link between chromosome and
cellular integrity. Interestingly, Mrr variants could be isolated that were
specifically compromised in either HhaII- or HP-dependent activation,
underscoring a mechanistic difference in the way both triggers activate Mrr. In
general, our results reveal that Mrr can take part in complex spatial
distributions on the nucleoid and can be engaged in distinct modes of activity.

<>

<1>Ghosh, I., Sun, L., Evans, T.C., Xu, M.Q.
<2>An improved method for utilization of peptide substrates for antibody characterization and enzymatic assays.
<3>J. Immunol. Methods
<4>293
<5>85-95
<6>2004
<7>Synthetic peptides have become an important tool in antibody production and enzyme
characterization. The small size of peptides, however, has
hindered their use in assays systems, such as Western blots, and as
immunogens. Here, we present a facile method to improve the properties
of peptides for multiple applications by ligating the peptides to
intein-generated carrier proteins. The stoichiometric ligation of
peptide and carrier achieved by intein-mediated protein ligation (IPL)
results in the ligation product migrating as a single band on a
SDS-PAGE gel. The carrier proteins, HhaI methylase (M.HhaI) and
maltose-binding protein (MBP), were ligated to various peptides; the
ligated carrier-peptide products gave sharp, reproducible bands when
used as positive controls for antibodies raised against the same
peptides during Western blot analysis. We further show that ligation of
the peptide antigens to a different thioester-tagged carrier protein,
paramyosin, produced immunogens for the production of antisera in
rabbits or mice. Furthermore, we demonstrate the generation of a
substrate for enzymatic assays by ligating a peptide containing the
phosphorylation site for Abl protein tyrosine kinase to a carrier
protein. This carrier-peptide protein was used as a kinase substrate
that could easily be tested for phosphorylation using a phosphotyrosine
antibody in Western blot analysis. These techniques do not require
sophisticated equipment, reagents, or skills thereby providing a simple
method for research and development.

<>

<1>Ghosh, S., LaPara, T.M., Sadowsky, M.J.
<2>Draft Genome Sequence of Sphingobacterium sp. Strain PM2-P1-29, a Tetracycline-Degrading TetX-Expressing Aerobic Bacterium Isolated from  Agricultural Soil.
<3>Genome Announcements
<4>2
<5>e00963-14
<6>2014
<7>The genome of Sphingobacterium sp. strain PM2-P1-29 was sequenced. The bacterium  contains a
physiologically active tet(X) gene, encoding a tetracycline-degrading
monooxygenase. To our knowledge, this is the only bacterium naturally harboring
tet(X) for which tetracycline degradation has been demonstrated.

<>

<1>Ghosh, S.S., Eis, P.S., Blumeyer, K., Fearon, K., Millar, D.P.
<2>Real time kinetics of restriction endonuclease cleavage monitored by fluorescence resonance energy transfer.
<3>Nucleic Acids Res.
<4>22
<5>3155-3159
<6>1994
<7>The kinetics of PaeR7I endonuclease-catalysed cleavage reactions of fluorophor-labeled
oligonucleotide substrates have been examined using fluorescence resonance energy transfer
(FRET). A series of duplex substrates were synthesized with an internal CTCGAG PaeR7I
recognition site and donor (fluorescein) and acceptor (rhodamine) dyes conjugated to the
opposing 5' termini. The time-dependent increase in donor fluorescence resulting from
restriction cleavage of these substrates was continuously monitored and the initial rate data
was fitted to the Michaelis-Menten equation. The steady state kinetic parameters for these
substrates were in agreement with the rate constants obtained from a gel electrophoresis-based
fixed time point assay using radiolabeled substrates. The FRET method provides a rapid
continuous assay as well as high sensitivity and reproducibility. These features should make
the technique useful for the study of DNA-cleaving enzymes.

<>

<1>Ghosh, S.S., Obermiller, P.S., Kwoh, T.J., Gingeras, T.R.
<2>Analysis of substrate specificity of the PaeR7I endonuclease: effect of base methylation on the kinetics of cleavage.
<3>Nucleic Acids Res.
<4>18
<5>5063-5068
<6>1990
<7>In murine cells expressing the PaeR7I endonuclease and methylase genes, the
recognition sites (CTCGAG) of these enzymes can be methylated at the adenine
residue by the PaeR7I methylase and at the internal cytosine by the mouse DNA
methyltransferase.  Using nonadecameric duplex deoxyoligonucleotide substrates,
the specificity of the PaeR7I endonuclease for unmethylated, hemi-methylated,
and fully methylated N6-methyladenine (m6A) and C5-methylcytosine (m5C)
versions of these substrates has been studied.  The Km, kcat, and Ki values for
these model substrates have been measured and suggest that fully or
hemi-m6A-methylated PaeR7I sites in the murine genome are completely protected.
However, the reactivity of fully or hemi-m5C-methylated PaeR7I sites is
depressed 2900- and 100-fold respectively, compared to unmodified PaeR7I sites.
The implications of the kinetic constants of the PaeR7I endonuclease for these
methylated recognition sites as they occur in murine cells expressing this
endonuclease gene are discussed.

<>

<1>Ghosh, W., George, A., Agarwal, A., Raj, P., Alam, M., Pyne, P., Das Gupta, S.K.
<2>Whole-Genome Shotgun Sequencing of the Sulfur-Oxidizing Chemoautotroph Tetrathiobacter kashmirensis.
<3>J. Bacteriol.
<4>193
<5>5553-5554
<6>2011
<7>The chemolithoautotrophic betaproteobacterium Tetrathiobacter kashmirensis belongs to the
family Alcaligenaceae and is phylogenetically closely
related to pathogens such as Taylorella and Bordetella species. While a
complete inorganic sulfur oxidation gene cluster, soxCDYZAXWB, is present
in its genome, pathogenicity islands or genes associated with virulence,
disease, cellular invasion, and/or intracellular resistance are completely
absent.

<>

<1>Giacani, L., Iverson-Cabral, S.L., King, J.C., Molini, B.J., Lukehart, S.A., Centurion-Lara, A.
<2>Complete Genome Sequence of the Treponema pallidum subsp. pallidum Sea81-4 Strain.
<3>Genome Announcements
<4>2
<5>e00333-14
<6>2014
<7>Using the rabbit model of syphilis, the Sea81-4 strain of Treponema pallidum subsp. pallidum
has been found to be more likely than other strains to invade the central nervous system
(CNS). To identify possible explanations for this important phenotype at the genomic level, we
sequenced the Sea81-4 strain genome.

<>

<1>Giacani, L., Jeffrey, B.M., Molini, B.J., Le HT, L.S.A., Centurion-Lara, A., Rockey, D.D.
<2>Complete genome sequence and annotation of the Treponema pallidum subsp. pallidum Chicago strain.
<3>J. Bacteriol.
<4>192
<5>2645-2646
<6>2010
<7>In syphilis research, the Nichols strain of Treponema pallidum, isolated in 1912, has been the
most widely studied. Recently, important differences
among T. pallidum strains emerged; therefore, we sequenced and annotated
the Chicago strain genome to facilitate and encourage the use of this
strain in studying the pathogenesis of syphilis.

<>

<1>Giammanco, G.M., Grimont, F., Grimont, P.A.
<2>MboII endonuclease heat inactivation before agarose gel electrophoresis to prevent artifactual bands in restriction patterns.
<3>Biotechniques
<4>27
<5>886-887
<6>1999
<7>MboII restriction enzyme belongs to class-IIS endonucleases group.  Like all of the enzymes
belonging to this class, MboII cleaves DNA at a specific distance from its recognition
sequence and still binds to its recognition sequence after DNA has been cleaved, because
binding and cleaving domains have separate functions.

<>

<1>Giannakis, M., Chen, S.L., Karam, S.M., Engstrand, L., Gordon, J.I.
<2>Helicobacter pylori evolution during progression from chronic atrophic gastritis to gastric cancer and its impact on gastric stem cells.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>4358-4363
<6>2008
<7>We have characterized the adaptations of Helicobacter pylori to a rarely
captured event in the evolution of its impact on host biology-the
transition from chronic atrophic gastritis (ChAG) to gastric
adenocarcinoma-and defined the impact of these adaptations on an
intriguing but poorly characterized interaction between this bacterium and
gastric epithelial stem cells. Bacterial isolates were obtained from a
single human host colonized with a single dominant strain before and after
his progression from ChAG to gastric adenocarcinoma during a 4-year
interval. Draft genome assemblies were generated from two isolates, one
ChAG-associated, the other cancer-associated. The cancer-associated strain
was less fit in a gnotobiotic transgenic mouse model of human ChAG and
better able to establish itself within a mouse gastric epithelial
progenitor-derived cell line (mGEP) that supports bacterial attachment.
GeneChip-based comparisons of the transcriptomes of mGEPs and a control
mouse gastric epithelial cell line revealed that, upon infection, the
cancer-associated strain regulates expression of GEP-associated signaling
and metabolic pathways, and tumor suppressor genes associated with
development of gastric cancer in humans, in a manner distinct from the
ChAG-associated isolate. The effects on GEP metabolic pathways, some of
which were confirmed in gnotobiotic mice, together with observed changes
in the bacterial transcriptome are predicted to support aspects of an
endosymbiosis between this microbe and gastric stem cells. These results
provide insights about how H. pylori may adapt to and influence stem cell
biology and how its intracellular residency could contribute to gastric
tumorigenesis.

<>

<1>Giannino, D., Mele, G., Cozza, R., Bruno, L., Testone, G., Ticconi, C., Frugis, G., Bitonti, M.B., Innocenti, A.M., Mariotti, D.
<2>Isolation and characterization of a maintenance DNA-methyltransferase gene from peach (Prunus persica [L.] Batsch): transcript localization in vegetative and reproductive meristems of triple buds.
<3>J. Exp. Bot.
<4>54
<5>2623-2633
<6>2003
<7>A cDNA coding for a DNA (cytosine-5)-methyltransferase (METase) was
isolated from peach (Prunus persica [L.] Batsch) and the corresponding
gene designated as PpMETI. The latter encoded a predicted polypeptide of
1564 amino acid residues and harboured all the functional domains
conserved in the maintenance METases group type I. PpMETI was a single
copy in the cultivar Chiripa which was used as a model in the present
study. Expression analyses revealed that PpMETI transcripts were more
abundant in tissues with actively proliferating cells such as apical tips,
uncurled leaves, elongating herbaceous stems, and small immature fruits.
Peach plants bear bud clusters (triads or triple buds), consisting of two
lateral and one central bud with floral and vegetative fates,
respectively. PpMETI in situ hybridization was performed in triple buds
during their entire developmental cycle. High and low levels of PpMETI
transcript were related to burst and quiescence of vegetative growth,
respectively. Message localization distinguished lateral from central buds
during the meristem switch to the floral phase. In fact, the PpMETI
message was abundant in the L1 layer of protruding domes, a morphological
trait marking the beginning of floral transition. The PpMETI transcript
was also monitored during organ flower formation. Altogether, these data
suggest a relationship between DNA replication and PpMETI gene expression.

<>

<1>Gibb, E.A., Edgell, D.R.
<2>Better late than early: delayed translation of intron-encoded endonuclease I-TevI is required for efficient splicing of its host group I intron.
<3>Mol. Microbiol.
<4>78
<5>35-46
<6>2010
<7>The td group I intron interrupting the thymidylate synthase (TS) gene of phage T4 is a mobile
intron that encodes the homing endonuclease
I-TevI. Efficient RNA splicing of the intron is required to restore
function of the TS gene, while expression of I-TevI from within the
intron is required to initiate intron mobility. Three distinct layers
of regulation temporally limit I-TevI expression to late in the T4
infective cycle, yet the biological rationale for stringent regulation
has not been tested. Here, we deleted key control elements to
deregulate I-TevI expression at early and middle times post T4
infection. Strikingly, we found that deregulation of I-TevI, or of a
catalytically inactive variant, generated a thymidine-dependent
phenotype that is caused by a reduction in td intron splicing.
Prematurely terminating I-TevI translation restores td splicing,
full-length TS synthesis, and rescues the thymidine-dependent
phenotype. We suggest that stringent translational control of I-TevI
evolved to prevent the ribosome from disrupting key structural elements
of the td intron that are required for splicing and TS function at
early and middle times post T4 infection. Analogous translational
regulatory mechanisms in unrelated intron-open reading frame
arrangements may also function to limit deleterious consequences on
splicing and host gene function.

<>

<1>Gibson, D.G. et al.
<2>Complete chemical synthesis, assembly, and cloning of a Mycoplasma genitalium genome.
<3>Science
<4>319
<5>1215-1220
<6>2008
<7>We have synthesized a 582,970-base pair Mycoplasma genitalium genome. This synthetic genome,
named M. genitalium JCVI-1.0, contains all the genes of wild-type M. genitalium G37 except
MG408, which was disrupted by an antibiotic marker to block pathogenicity and to allow for
selection. To identify the genome as synthetic, we inserted "watermarks" at intergenic sites
known to tolerate transposon insertions. Overlapping "cassettes" of 5 to 7 kilobases (kb),
assembled from chemically synthesized oligonucleotides, were joined by in vitro recombination
to produce intermediate assemblies of approximately 24 kb, 72 kb ("1/8 genome"), and 144 kb
("1/4 genome"), which were all cloned as bacterial artificial chromosomes in Escherichia coli.
Most of these intermediate clones were sequenced, and clones of all four 1/4 genomes with the
correct sequence were identified. The complete synthetic genome was assembled by
transformation-associated recombination cloning in the yeast Saccharomyces cerevisiae, then
isolated and sequenced. A clone with the correct sequence was identified. The methods
described here will be generally useful for constructing large DNA molecules from chemically
synthesized pieces and also from combinations of natural and synthetic DNA segments.

<>

<1>Gibson, D.G. et al.
<2>Creation of a bacterial cell controlled by a chemically synthesized genome.
<3>Science
<4>329
<5>52-56
<6>2010
<7>We report the design, synthesis, and assembly of the 1.08-mega-base pair
Mycoplasma mycoides JCVI-syn1.0 genome starting from digitized genome
sequence information and its transplantation into a M. capricolum
recipient cell to create new M. mycoides cells that are controlled only by
the synthetic chromosome. The only DNA in the cells is the designed
synthetic DNA sequence, including "watermark" sequences and other designed
gene deletions and polymorphisms, and mutations acquired during the
building process. The new cells have expected phenotypic properties and
are capable of continuous self-replication.

<>

<1>Gil, R., Silva, F.J., Zientz, E., Delmotte, F., Gonzalez-Candelas, F., Latorre, A., Rausell, C., Kamerbeek, J., Gadau, J., Holldobler, B., Van Ham, R.C., Gross, R., Moya, A.
<2>The genome sequence of Blochmannia floridanus: Comparative analysis of reduced genomes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>9388-9393
<6>2003
<7>Bacterial symbioses are widespread among insects, probably being one of the key factors of
their evolutionary success. We present the complete
genome sequence of Blochmannia floridanus, the primary endosymbiont of
carpenter ants. Although these ants feed on a complex diet, this symbiosis
very likely has a nutritional basis: Blochmannia is able to supply
nitrogen and sulfur compounds to the host while it takes advantage of the
host metabolic machinery. Remarkably, these bacteria lack all known genes
involved in replication initiation (dnaA, priA, and recA). The
phylogenetic analysis of a set of conserved protein-coding genes shows
that Bl. floridanus is phylogenetically related to Buchnera aphidicola and
Wigglesworthia glossinidia, the other endosymbiotic bacteria whose
complete genomes have been sequenced so far. Comparative analysis of the
five known genomes from insect endosymbiotic bacteria reveals they share
only 313 genes, a number that may be close to the minimum gene set
necessary to sustain endosymbiotic life.

<>

<1>Gilbert, M.J., Miller, W.G., Yee, E., Blaser, M.J., Wagenaar, J.A., Duim, B.
<2>Complete Genome Sequence of Campylobacter fetus subsp. testudinum Strain 03-427T.
<3>Genome Announcements
<4>1
<5>e01002-13
<6>2013
<7>Campylobacter fetus subsp. testudinum has been isolated from reptiles and humans. This
Campylobacter subspecies is genetically distinct from other C. fetus
subspecies. Here, we present the first whole-genome sequence for this C. fetus
subspecies.

<>

<1>Gilbert, M.J., Miller, W.G., Yee, E., Kik, M., Wagenaar, J.A., Duim, B.
<2>Complete Genome Sequence of Campylobacter iguaniorum Strain 1485ET, Isolated from a Bearded Dragon (Pogona vitticeps).
<3>Genome Announcements
<4>2
<5>e00844-14
<6>2014
<7>Campylobacter iguaniorum has been isolated from reptiles. This Campylobacter species is
genetically related to Campylobacter fetus and Campylobacter
hyointestinalis. Here we present the first whole-genome sequence for this
species.

<>

<1>Gill, J.J., Berry, J.D., Russell, W.K., Lessor, L., Escobar, G., D, A., Hernandez, D., Kane, A., Keene, J., Maddox, M., Martin, R., Mohan, S., Thorn, A.M., Russell, D.H., Young, R.
<2>The Caulobacter crescentus phage phiCbK: genomics of a canonical phage.
<3>BMC Genomics
<4>13
<5>542
<6>2012
<7>ABSTRACT: BACKGROUND: The bacterium Caulobacter crescentus is a popular model for
the study of cell cycle regulation and senescence. The large prolate siphophage
phiCbK has been an important tool in C. crescentus biology, and has been studied
in its own right as a model for viral morphogenesis. Although a system of some
interest, to date little genomic information is available on phiCbK or its
relatives. RESULTS: Five novel phiCbK-like C. crescentus bacteriophages,
CcrMagneto, CcrSwift, CcrKarma, CcrRogue and CcrColossus, were isolated from the
environment. The genomes of phage phiCbK and these five environmental phage
isolates were obtained by 454 pyrosequencing. The phiCbK-like phage genomes range
in size from 205 kb encoding 318 proteins (phiCbK) to 280 kb encoding 448
proteins (CcrColossus), and were found to contain nonpermuted terminal
redundancies of 10 to 17 kb. A novel method of terminal ligation was developed to
map genomic termini, which confirmed termini predicted by coverage analysis. This
suggests that sequence coverage discontinuities may be useable as predictors of
genomic termini in phage genomes. Genomic modules encoding virion morphogenesis,
lysis and DNA replication proteins were identified. The phiCbK-like phages were
also found to encode a number of intriguing proteins; all contain a clearly
T7-like DNA polymerase, and five of the six encode a possible homolog of the C.
crescentus cell cycle regulator GcrA, which may allow the phage to alter the host
cell's replicative state. The structural proteome of phage phiCbK was determined,
identifying the portal, major and minor capsid proteins, the tail tape measure
and possible tail fiber proteins. All six phage genomes are clearly related;
phiCbK, CcrMagneto, CcrSwift, CcrKarma and CcrRogue form a group related at the
DNA level, while CcrColossus is more diverged but retains significant similarity
at the protein level. CONCLUSIONS: Due to their lack of any apparent relationship
to other described phages, this group is proposed as the founding cohort of a new
phage type, the phiCbK-like phages. This work will serve as a foundation for
future studies on morphogenesis, infection and phage-host interactions in C.
crescentus.

<>

<1>Gill, S.R. et al.
<2>Insights on evolution of virulence and resistance from the complete genome analysis of an early methicillin-resistant Staphylococcus aureus strain and a biofilm-producing methicillin-resistant Staphylococcus epidermidis strain.
<3>J. Bacteriol.
<4>187
<5>2426-2438
<6>2005
<7>Staphylococcus aureus is an opportunistic pathogen and the major causative agent of numerous
hospital- and community-acquired infections. Staphylococcus epidermidis has emerged as a
causative agent of infections often associated with implanted medical devices. We have
sequenced the  2.8-Mb genome of S. aureus COL, an early methicillin-resistant isolate, and the
2.6-Mb genome of S. epidermidis RP62a, a methicillin-resistant biofilm isolate. Comparative
analysis of these and other staphylococcal genomes was used to explore the evolution of
virulence and resistance between these two species. The S. aureus and S. epidermidis genomes
are syntenic throughout their lengths and share a core set of 1,681 open reading frames.
Genome islands in nonsyntenic regions are the primary source of variations in pathogenicity
and resistance. Gene transfer between staphylococci and low-GC-content gram-positive bacteria
appears to have shaped their virulence and resistance profiles. Integrated plasmids in S.
epidermidis carry genes encoding resistance to cadmium and species-specific LPXTG surface
proteins. A novel genome island encodes multiple phenol-soluble modulins, a potential S.
epidermidis virulence factor. S. epidermidis contains the cap operon, encoding the
polyglutamate capsule, a major virulence factor in Bacillus anthracis. Additional phenotypic
differences are likely the result of single nucleotide polymorphisms, which are most numerous
in cell envelope proteins. Overall differences in pathogenicity can be attributed to genome
islands in S. aureus which encode enterotoxins, exotoxins, leukocidins, and leukotoxins not
found in S. epidermidis.

<>

<1>Gillings, M.R., Labbate, M., Sajjad, A., Giguere, N.J., Holley, M.P., Stokes, H.W.
<2>Mobilization of a Tn402-like class 1 integron with a novel cassette array via flanking miniature inverted-repeat transposable element-like structures.
<3>Appl. Environ. Microbiol.
<4>75
<5>6002-6004
<6>2009
<7>A Tn402-like class 1 integron was recovered from a prawn-associated
bacterium. One of its cassettes included methionine sulfoxide reductase
genes, the first example of such genes being captured by an integron. The
integron was flanked by direct repeats that resemble miniature
inverted-repeat transposable element sequences. Excision of the integron
by homologous recombination through these sequences was demonstrated.

<>

<1>Gimble, F.S.
<2>Invasion of a multitude of genetic niches by mobile endonuclease genes.
<3>FEMS Microbiol. Lett.
<4>185
<5>99-107
<6>2000
<7>Persistence of a mobile DNA element in a population reflects a balance between the ability of
the host to eliminate the element and the ability of the element to survive and to disseminate
to other individuals. In each of the three biological kingdoms, several families of a mobile
DNA element have been identified which encode a single protein that acts on nucleic acids.
Collectively termed homing endonuclease genes (HEGs), these elements employ varied strategies
to ensure their survival. Some members of the HEG families have a minimal impact on host
fitness because they associate with genes having self-splicing introns or inteins that remove
the HEGs at the RNA or protein level. The HEG and the intron/intein gene spread throughout the
population by a gene conversion process initiated by the HEG-encoded endonuclease called
'homing' in which the HEG and intron/intein genes are copied to cognate alleles that lack
them. The endonuclease activity also contributes to a high frequency of lateral transmission
of HEGs between species as has been documented in plants and other systems. Other HEGs have
positive selection value because the proteins have evolved activities that benefit their host
organisms. The success of HEGs in colonizing diverse genetic niches results from the
flexibility of the encoded endonucleases in adopting new specificities.

<>

<1>Gimble, F.S.
<2>Degeneration of a homing endonuclease and its target sequence in a wild yeast strain.
<3>Nucleic Acids Res.
<4>29
<5>4215-4223
<6>2001
<7>Mobile introns and inteins self-propagate by 'homing', a gene conversion process initiated
by site-specific homing endonucleases. The VMA intein, which encodes the PI-SceI endonuclease
in Saccharomyces cerevisiae, is present in several different yeast strains. Surprisingly, a
wild wine yeast (DH1-1A) contains not only the intein(+) allele, but also an inteinless allele
that has not undergone gene conversion. To elucidate how these two alleles co-exist, we
characterized the endonuclease encoded by the DH1-1A intein(+) allele and the target site in
the intein(-) allele. Sequence analysis reveals seven mutations in the 31 bp recognition
sequence, none of which occurs at positions that are individually critical for activity.
However, binding and cleavage of the sequence by PI-SceI is reduced 10-fold compared to the S.
cerevisiae target. The PI-SceI analog encoded by the DH1-1A intein(+) allele contains 11
mutations at residues in the endonuclease and protein splicing domains. None affects protein
splicing, but one, a R417Q substitution, accounts for most of the decrease in DNA cleavage and
DNA binding activity of the DH1-1A protein. Loss of activity in the DH1-1A endonuclease and
target site provides one explanation for co-existence of the intein(+) and intein(-) alleles.

<>

<1>Gimble, F.S.
<2>Broken symmetry in homing endonucleases.
<3>Structure
<4>14
<5>804-806
<6>2006
<7>Homing DNA endonucleases are highly site-specific enzymes that initiate the transfer of mobile
DNA elements. In this issue of Structure, Spiegel et al. report the structure of the I-CeuI
homing enzyme and describe how a symmetric homodimeric enzyme acquired specificity for an
asymmetric substrate.

<>

<1>Gimble, F.S.
<2>Engineering homing endonucleases for genomic applications.
<3>Nucleic Acids Mol. Biol.
<4>16
<5>177-192
<6>2005
<7>The rapid progress in molecular biology that has occurred over the last three decades is due
in large part to the availability of sequence-specific nucleases.  These enzymes have been
indispensable tools in recombinant DNA protocols.  Most notably, the type II bacterial
restriction enzymes have permitted the rapid and low-cost production of recombinant DNAs for
cloning and other methods.  One drawback of these enzymes, however, is the small size of their
recognition sequences, which range in length from 4-8 base pairs.  An enzyme that recognizes a
6-bp target sequence cleaves DNA on average approximately once every 4000 bp.  When these
enzymes digest DNA from a mammalian genome, which is typically >100 megabases in length, many
thousands of DNA fragments are generated, and purification of individual species from this
pool is difficult.

<>

<1>Gimble, F.S.
<2>Engineering homing endonucleases to modify complex genomes.
<3>Gene Ther. Regul.
<4>3
<5>33-50
<6>2006
<7>Gene targeting to selected chromosomal loci is greatly stimulated when free DNA ends are
created that initiate double-strand break repair.  Gene therapy reagents can be developed by
engineering DNA endonucleases that cleave genomes at desired target sequences.  Homing
endonucleases are naturally occurring rare-cutting enzymes that have well understood DNA
binding and DNA cleavage properties.  Rational design methods as well as directed evolution
strategies that involve genetic selections and screens using combinatorial libraries generate
homing endonucleases with altered sequence specificities.  Molecular switches are being
introduced into these enzymes to regulate their activity.  This article reviews the progress
that has been made in constructing homing endonucleases for gene therapy and genome
engineering, and discusses the challenges that remain.

<>

<1>Gimble, F.S., Duan, X., Hu, D., Quiocho, F.A.
<2>Identification of lys-403 in the PI-SceI homing endonuclease as part of a symmetric catalytic center.
<3>J. Biol. Chem.
<4>273
<5>30524-30529
<6>1998
<7>Superposition of the PI-SceI and I-CreI homing endonuclease three-dimensional x-ray structures
indicates general similarity between the I-CreI homodimer and the PI-SceI endonuclease domain.
Saddle-shaped structures are present in each protein that are proposed to bind DNA. At the
putative endonucleolytic active sites, the superposition reveals that two lysine (Lys-301 and
Lys-403 in PI-SceI and Lys-98 and Lys-98' in I-CreI) and two aspartic acid residues (Asp-218
and Asp-326 in PI-SceI and Asp-20 and Asp-20' in I-CreI) are related by 2-fold symmetry. The
critical role of Lys-301, Asp-218, and Asp-326 in the PI-SceI reaction pathway was reported
previously. Here, we demonstrate the significance of the active-site symmetry by showing that
alanine substitution at Lys-403 reduces cleavage activity by greater than 50-fold but has
little effect on the DNA binding activity of the mutant enzyme. Substitution of Lys-403 with
arginine, which maintains the positive charge, has only a modest effect on activity.
Interestingly, even though the Lys-301 and Lys-403 residues display pseudosymmetry, PI-SceI
mutant proteins with substitutions at these positions have different behaviors. The presence
of similar basic and acidic residues in many LAGLIDADG homing endonucleases suggests that
these enzymes use a common reaction mechanism to cleave double-stranded DNA.

<>

<1>Gimble, F.S., Moure, C.M., Posey, K.L.
<2>Assessing the plasticity of DNA target site recognition of the PI-SceI homing endonuclease using a bacterial two-hybrid selection system.
<3>J. Mol. Biol.
<4>334
<5>993-1008
<6>2003
<7>The PI-SceI protein from Saccharomyces cerevisiae is a member of the LAGLIDADG family of
homing endonucleases that have been used in genomic
engineering. To assess the flexibility of the PI-SceI-binding interaction
and to make progress towards the directed evolution of homing
endonucleases that cleave specified DNA targets, we applied a two-hybrid
method to select PI-SceI variants from a randomized expression library
that bind to different DNA substrates. In particular, the codon for Arg94,
which is located in the protein splicing domain and makes essential
contacts to two adjacent base-pairs, and the codons for four proximal
residues were randomized. There is little conservation of the wild-type
amino acid residues at the five randomized positions in the variants that
were selected to bind to the wild-type site, yet one of the purified
derivatives displays DNA-binding specificity and DNA endonuclease activity
that is similar to that of the wild-type enzyme. A spectrum of DNA-binding
behaviors ranging from partial relaxation of specificity to marked shifts
in target site recognition are present in variants selected to bind to
sites containing mutations at the two base-pairs. Our results illustrate
the inherent plasticity of the PI-SceI/DNA interface and demonstrate that
selection based on DNA binding is an effective means of altering the DNA
cleavage specificity of homing endonucleases. Furthermore, it is apparent
that homing endonuclease target specificity derives, in part, from
constraints on the flexibility of DNA contacts imposed by hydrogen bonds
to proximal residues.

<>

<1>Gimble, F.S., Stephens, B.W.
<2>Substitutions in conserved dodecapeptide motifs that uncouple the DNA binding and DNA cleavage activities of PI-SceI endonuclease.
<3>J. Biol. Chem.
<4>270
<5>5849-5856
<6>1995
<7>The PI-SceI endonuclease from yeast belongs to a protein family whose members contain two
conserved dodecapeptide motifs within their primary sequences. The function of two acidic
residues within these motifs, Asp218 and Asp326, was examined by substituting alanine,
asparagine, and glutamic acid residues at these positions. All of the purified mutant proteins
bind to the PI-SceI recogniton site with the same affinity and specificity as the wild-type
enzyme. By contrast, substituting alanine or asparagine amino acids at the two positions
completely eliminates strand cleavage of substrate DNA, whereas substitution with glutamic
acid markedly reduces the cleavage activity. Experiments using nicked substrates demonstrate
that the wild-type enzyme shows no strand preference during cleavage. These results are
consistent with a model in which both acidic residues are part of a single catalytic center
that cleaves both DNA strands. Furthermore, substrate binding by wild-type PI-SceI stimulates
hydroxyl radical or hydroxide ion attack at the cleavage site while binding by the
alanine-substituted proteins either stimulates this attack significantly less or protects the
DNA at this position. These findings are discussed in terms of possible reaction mechanisms
for PI-SceI-mediated endonucleolytic cleavage.

<>

<1>Gimble, F.S., Thorner, J.
<2>Purification and characterization of VDE, a site-specific endonuclease from the yeast Saccharomyces cerevisiae.
<3>J. Biol. Chem.
<4>268
<5>21844-21853
<6>1993
<7>The 119-kDa primary translation product of the VMA1 gene of Saccharomyces cerevisiae undergoes
a self-catalyzed rearrangement (protein splicing) that excises an internal 50-kDa segment of
the polypeptide and joins the amino-terminal and carboxyl-terminal segments to generate the
69-kDa subunit of the vacuolar membrane-associated H+-ATPase. We have shown previously that
the internal segment is a site-specific endonuclease (Gimble, F.S., and Thorner, J. (1992)
Nature 357, 301-306). Here we describe methods for the high level expression and purification
to near homogeneity of both the authentic VMA1-derived endonuclease (or VDE) from yeast (yield
18%) and a recombinant form of VDE made in bacteria (yield 29%). Detailed characterization of
these preparations demonstrated that the yeast-derived and bacterially produced enzymes were
indistinguishable, as judged by: (a) behavior during purification; (b) apparent native
molecular mass (50 kDa); (c) immunological reactivity; and (d) catalytic properties (specific
activity; cleavage site recognition; and optima for pH, temperature, divalent cation and ionic
strength). The minimal site required for VDE cleavage was delimited to a 30-base pair sequence
within its specific substrate ( the VMA1 delvde allele).

<>

<1>Gimble, F.S., Thorner, J.
<2>Homing of a DNA endonuclease gene by meiotic gene conversion in Saccharomyces cerevisiae.
<3>Nature
<4>357
<5>301-306
<6>1992
<7>An unusual protein splicing reaction joins the N-terminal segment (A) and the C-terminal
segment (C) of the 119K primary translation product (ABC) of the yeast VMA1 gene to yield a
69K vacuolar H+-ATPase subunit (AC) and an internal 50K polypeptide (B). This 50K protein is a
site-specific DNA endonuclease that shares 34% indentity with the homothallic switching
endonuclease. The site cleaved by the VMA1-derived endonuclease exists in a VMA1 allele that
lacks the derived endonuclease segment of the open reading frame. Cleavage at this site only
occurs during meiosis and initiates 'homing', a gentic event that converts a VMA1 allele
lacking the endonuclease coding sequence into one that contains it.

<>

<1>Gimble, F.S., Wang, J.
<2>Substrate recognition and induced DNA distortion by the PI-SceI endonuclease, an enzyme generated by protein splicing.
<3>J. Mol. Biol.
<4>263
<5>163-180
<6>1996
<7>PI-SceI, a double-stranded DNA endonuclease from Saccharomyces cerevisiae, is generated by
protein splicing of an intein, which is an internal polypeptide within a larger precursor
protein.  The enzyme initiates the mobility of the intein by cleaving at inteinless alleles of
the VMA1 gene.  Genetic and biochemical studies reveal that the enzyme makes numerous
base-specific and phosphate backbone contacts with its 31 bp asymmetrical recognition site.
This site can be divided into two regions, both of which contain nucleotides that are
essential for cleavage by PI-SceI.  Region I contains the PI-SceI cleavage site while Region
II includes an adjacent sequence that covers two helical turns.  Mutational, interference and
DNA mobility shift analyses demonstrate that Region II is sufficient for high-affinity PI-SceI
binding.  Within this region, PI-SceI uses primarily phosphate backbone and some major groove
interactions to contact the DNA, while within Region I , protein binding involves
predominantly major groove interactions that overlap and lie proximal to the cleavage site.
Interestingly, DNA binding by PI-SceI induces DNA conformational changes within Region II that
are entirely exclusive of Region I sequences.  Furthermore, additional distortion occurs when
PI-SceI binds to Region I in conjunction with Region II.  The importance of this latter
distortion in the cleavage pathway is underscored by substrate mutations at or near the
cleavage site that reduce or eliminate both Region I DNA bending and substrate cleavage.
Based on these findings, we propose a model in which sequence-specific contacts made by
PI-SceI contribute to its localization to the cleavage site and to its stabilization of a DNA
conformation that is required for catalysis.  Finally, we discuss how the recognition
characteristics of PI-SceI may have allowed the evolution of other endonucleases with altered,
but similar, specificities.

<>

<1>Gingeras, T.R.
<2>Restriction endonucleases and their recognition sequences.
<3>Biochem. Soc. Trans.
<4>8
<5>397-398
<6>1980
<7>None

<>

<1>Gingeras, T.R.
<2>Restriction-modification systems: Genetic sentries and useful systems in the study of molecular genetics.
<3>Modern Microbial Genetics, Wiley-Liss, Inc., Streips, U.N., Yasbin, R.E., New York
<4>0
<5>301-321
<6>1991
<7>This chapter describes two important roles which restriction-modification
systems have in molecular genetics.  The first of these roles concerns the
operation of these systems as genetic sentries.  The second role involves the
part which restriction-modification systems are playing in the study of several
important topics in molecular genetics.

<>

<1>Gingeras, T.R., Blumenthal, R.M., Roberts, R.J., Brooks, J.E.
<2>The isolation and characterization of the E. coli dam methylase gene.
<3>Metabolism and Enzymology of Nucleic Acids, Publishing House of the Slovak Academy of Sciences, Zelinka, J., Balan, J., Bratislava
<4>4
<5>329-340
<6>1982
<7>The E. coli dam (DNA adenine methylase) codes for an enzyme which methylates the DNA sequence
GATC. When DNA has been modified by dam it is no longer susceptible to endonucleolytic
cleavage by the restriction endonuclease MboI. Several DNA methylases have been shown to be
part of a restriction-modification system (Roberts, 1981). However, the dam methylase does not
seem to be part of such a system. Rather, the dam methylase has been implicated as part of a
post-replicational mismatch repair system in E. coli. In heteroduplex lambda DNA containing
only one methylated strand, the repair system will correct the unmethylated strand to match
the methylated strand. (Wagner and Meselson, 1976, Meselson et al., 1980). Fully methylated
mismatched heteroduplexes are not corrected. In addition, it has been demonstrated that when
the dam methylase is not produced (dam-) or is over-produced (damS), such E. coli strains are
hypermutable. (Herman and Modrich, 1980). An additional function for the dam methylase in E.
coli may involve its role in DNA replication. It has been shown that the sequence methylated
by the dam methylase, GATC, occurs at a very high frequency (i.e., 11 times within 245 base
pairs) at the E. coli origin of replication (J. Zyskind, personal communication). Furthermore,
such sites have been shown to occur near or at the ends of Okazaki fragments (Gomez-Eichelmann
and Lark, 1977). Because of the possible important roles that the dam methylasae plays in the
biology of E. coli, as well as sharing an identical recognition sequence with a set of type II
restriction and modification enzymes (MboI), we have isolated and characterized clones
containing the dam methylase gene.

<>

<1>Gingeras, T.R., Brooks, J.E.
<2>Cloned restriction/modification system from Pseudomonas aeruginosa.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>80
<5>402-406
<6>1983
<7>DNA fragments from Pseudomonas aeruginosa carrying the PaeR7 restriction/modification genes
have been cloned in the plasmid vector pBR322 and propagated in Escherichia coli. A subclone
(pPAORM3.8) has been constructed that contains the complete restriction/modification system on
a 3.8-kilobase DNA fragment. Digestion of the pPAORM3.8 plasmid with nuclease BAL-31 has
yielded two types of clones. One type contains an active methylase gene but no active
endonuclease gene; such clones will modify the DNA but not restrict the growth of incoming
phage in vivo. The second type contains an active endonuclease gene but no active methylase
gene, as judged both by in vivo tests and by the activity of the cell extracts in vitro.
Although extracts of cells containing these plasmids display restriction endonuclease
activity, these bacteria are unable to restrict the growth of incoming phage. Furthermore,
chromosomal and phage DNA isolated from these host cells are not protected against cleavage by
PaeR7 in vitro. The properties of PaeR7 endonuclease and methylase enzymes have also been
examined. The PaeR7 restriction endonuclease recognizes and cleaves the sequence C^T-C-G-A-G,
as does XhoI. However, there exists a canonical XhoI site at 26.5% on the adenovirus 2
genome which is totally refractory to PaeR7 cleavage but is cut by XhoI. Under conditions of
low salt, high glycerol, and high enzyme concentrations, a "PaeR7*" activity is found that is
similar to that observed for EcoRI. Finally, evidence is presented that the PaeR7 methylase
modifies the adenine residue within the recognition sequence. the restriction enzyme, the
third describes the purification procedure for the methylase and the fourth describes the
recognition sequence of the methylase. In some cases, several references appear in one of
these categories when independent groups have reached similar conclusions.

<>

<1>Gingeras, T.R., Ghosh, S., Blumeyer, K., Eis, R., Millar, D.
<2>A spectrofluorometric method for measurement of restriction endonuclease activity based on fluorescence polarization.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>173
<6>1993
<7>A method is described for monitoring the enzymatic activity of type II restriction
endonucleases using fluorophore-labeled oligonucleotide substrates. Cleavage of the duplex
oligonucleotide substrates, in which one strand is covalently labeled with fluorescein,
results in a time-dependent change of fluorescence polarization due to the formation of short,
labeled fragments which are single-stranded at the temperature of the assay. This non-isotopic
homogeneous approach is extremely sensitive and is much simpler than the current radiolabeled
procedures which require fixed timepoint assays and the use of filter binding or gel
electrophoresis for kinetic analysis of the cleavage reaction. The PaeR7I endonuclease has
been used as an example for the applicability of the method. The kinetic parameters for the
hydrolysis of a fluorescein-modified oligonucleotide substrate of the enzyme have been
determined by using this approach to follow reaction rates.

<>

<1>Gingeras, T.R., Greenough, L., Schildkraut, I., Roberts, R.J.
<2>Two new restriction endonucleases from Proteus vulgaris.
<3>Nucleic Acids Res.
<4>9
<5>4525-4536
<6>1981
<7>Two novel sequence-specific endonucleases have been isolated from Proteus
vulgaris, ATCC 13315.  PvuI recognizes the sequence: 5' CGAT^CG 3' 3' GC^TAGC
5' and PvuII recognizes the sequence: 5' CAG^CTG 3' 3' GTC^GAC 5' and cleave as
indicated by the arrow.  PvuI is an isoschizomer of XorII, RshI, and XniI.  No
enzyme with the specificity of PvuII has been described previously.

<>

<1>Gingeras, T.R., Milazzo, J.P., Roberts, R.J.
<2>A computer assisted method for the determination of restriction enzyme recognition sites.
<3>Nucleic Acids Res.
<4>5
<5>4105-4127
<6>1978
<7>A computer program has been developed which aids in the determination of
restriction enzyme recognition sequences.  This is achieved by cleaving DNAs of
known sequence with a restriction endonuclease and comparing the fragmentation
pattern with a computer-generated set of patterns.  The feasibility of this
approach has been tested using fragmentation patterns of PhiX174 DNA produced
by enzymes of both known and unknown specificity.  Recognition sequences are
predicted for two restriction endonucleases (BbvI and SfaNI) using this method.
In addition, recognition sequnces are predicted for two other new enzymes
(PvuI and MstI) using another computer-assisted method.

<>

<1>Gingeras, T.R., Myers, P.A., Olson, J.A., Hanberg, F.A., Roberts, R.J.
<2>A new specific endonuclease present in Xanthomonas holcicola, Xanthomonas papavericola and Brevibacterium luteum.
<3>J. Mol. Biol.
<4>118
<5>113-122
<6>1978
<7>A new specific endonuclease, XhoI, has been partially purified from Xanthomonas
holcicola.  This enzyme cleaves adenovirus-2 DNA at five sites, bacteriophage
DNA at one site, PhiX174 DNA at one site, but does not cleave simian virus DNA.
It recognizes the sequence 5'-C-^T-C-G-A-G-3' 3'-G-A-G-C-T^-C-5' and cuts at
the sites indicated by the arrows.  Enzymes with identical specificity have
also been found in Xanthomonas papavericola and Brevibacterium luteum.

<>

<1>Gingeras, T.R., Theriault, G., Brooks, J.E.
<2>Organization and expression of a type II restriction-modification system from Pseudomonas aeruginosa.
<3>Metabolism and Enzymology of Nucleic Acids, Publishing House of the Slovak Academy of Sciences., Zelinka, J., Balan, J., Bratislava
<4>5
<5>267-275
<6>1984
<7>A series of BAL-31 deletion mutants have been generated affecting both the structural and
regulatory regions of the PaeR7 restriction-modification system. This system is organized as
an operon with both the genes sharing a common regulatory region. Deletions in this regulatory
region lead to a gradient of expression levels for both enzymes. Most striking is the
observation that depressed levels of expression of endonuclease, as exemplified by both
pPAOR1.9 and pPAOdel1.3, result in clones that do not restrict infecting phage.
Cotransformation of pPAOR1.9 and pPACYCM2.7 into the same host did not result in recovery of a
restriction phenotype. Consequently, phage restriction seems to be critically dependent on the
levels of endonuclease produced, and not on the effect of an active methylase gene. DNA
sequences of the control region of this system indicate two PaeR7 sites which are located in a
critical position in the PaeR7 operon to determine if the levels of methylase are sufficient
to protect both the plasmid and the DNA of the host cell. The effect of methylation on these
sites during transcription is being studied as another possible means of controlling
expression in this system.

<>

<1>Gioia, J. et al.
<2>Paradoxical DNA Repair and Peroxide Resistance Gene Conservation in Bacillus pumilus SAFR-032.
<3>PLoS ONE
<4>2
<5>e928
<6>2007
<7>BACKGROUND: Bacillus spores are notoriously resistant to unfavorable conditions such as UV
radiation, gamma-radiation, H(2)O(2), desiccation,
chemical disinfection, or starvation. Bacillus pumilus SAFR-032 survives
standard decontamination procedures of the Jet Propulsion Lab spacecraft
assembly facility, and both spores and vegetative cells of this strain
exhibit elevated resistance to UV radiation and H(2)O(2) compared to other
Bacillus species. PRINCIPAL FINDINGS: The genome of B. pumilus SAFR-032
was sequenced and annotated. Lists of genes relevant to DNA repair and the
oxidative stress response were generated and compared to B. subtilis and
B. licheniformis. Differences in conservation of genes, gene order, and
protein sequences are highlighted because they potentially explain the
extreme resistance phenotype of B. pumilus. The B. pumilus genome includes
genes not found in B. subtilis or B. licheniformis and conserved genes
with sequence divergence, but paradoxically lacks several genes that
function in UV or H(2)O(2) resistance in other Bacillus species.
SIGNIFICANCE: This study identifies several candidate genes for further
research into UV and H(2)O(2) resistance. These findings will help explain
the resistance of B. pumilus and are applicable to understanding
sterilization survival strategies of microbes.

<>

<1>Gioia, J., Qin, X., Jiang, H., Clinkenbeard, K., Lo, R., Liu, Y., Fox, G.E., Yerrapragada, S., McLeod, M.P., McNeill, T.Z., Hemphill, L., Sodergren, E., Wang, Q., Muzny, D.M., Homsi, F.J., Weinstock, G.M., Highlander, S.K.
<2>The Genome Sequence of Mannheimia haemolytica A1: Insights into Virulence, Natural Competence, and Pasteurellaceae Phylogeny.
<3>J. Bacteriol.
<4>188
<5>7257-7266
<6>2006
<7>The draft genome sequence of Mannheimia haemolytica A1, the causative agent of bovine
respiratory disease complex (BRDC), is presented. Strain
ATCC BAA-410, isolated from the lung of a calf with BRDC, was the DNA
source. The annotated genome includes 2,839 coding sequences, 1,966 of
which were assigned a function and 436 of which are unique to M.
haemolytica. Through genome annotation many features of interest were
identified, including bacteriophages and genes related to virulence,
natural competence, and transcriptional regulation. In addition to
previously described virulence factors, M. haemolytica encodes adhesins,
including the filamentous hemagglutinin FhaB and two trimeric
autotransporter adhesins. Two dual-function
immunoglobulin-protease/adhesins are also present, as is a third
immunoglobulin protease. Genes related to iron acquisition and drug
resistance were identified and are likely important for survival in the
host and virulence. Analysis of the genome indicates that M. haemolytica
is naturally competent, as genes for natural competence and DNA uptake
signal sequences (USS) are present. Comparison of competence loci and USS
in other species in the family Pasteurellaceae indicates that M.
haemolytica, Actinobacillus pleuropneumoniae, and Haemophilus ducreyi form
a lineage distinct from other Pasteurellaceae. This observation was
supported by a phylogenetic analysis using sequences of predicted
housekeeping genes.

<>

<1>Giongo, A., Tyler, H.L., Zipperer, U.N., Triplett, E.W.
<2>Two genome sequences of the same bacterial strain, Gluconacetobacter diazotrophicus PAl 5, suggest a new standard in genome sequence submission.
<3>Standards in Genomic Sciences
<4>2
<5>309-317
<6>2010
<7>Gluconacetobacter diazotrophicus PAl 5 is of agricultural significance due to its ability to
provide fixed nitrogen to plants. Consequently, its genome sequence
has been eagerly anticipated to enhance understanding of endophytic nitrogen
fixation. Two groups have sequenced the PAl 5 genome from the same source (ATCC
49037), though the resulting sequences contain a surprisingly high number of
differences. Therefore, an optical map of PAl 5 was constructed in order to
determine which genome assembly more closely resembles the chromosomal DNA by
aligning each sequence against a physical map of the genome. While one sequence
aligned very well, over 98% of the second sequence contained numerous
rearrangements. The many differences observed between these two genome sequences
could be owing to either assembly errors or rapid evolutionary divergence. The
extent of the differences derived from sequence assembly errors could be assessed
if the raw sequencing reads were provided by both genome centers at the time of
genome sequence submission. Hence, a new genome sequence standard is proposed
whereby the investigator supplies the raw reads along with the closed sequence so
that the community can make more accurate judgments on whether differences
observed in a single stain may be of biological origin or are simply caused by
differences in genome assembly procedures.

<>

<1>Giordano, M., Mattachini, M.E., Cella, R., Pedrali-Noy, G.
<2>Purification and properties of a novel DNA methyltransferase from cultured rice cells.
<3>Biochem. Biophys. Res. Commun.
<4>177
<5>711-719
<6>1991
<7>DNA methyltransferase activity has been observed in a total crude homogenate of rice cells
grown in suspension culture using either native plant DNA or, under the conditions used, the
more responsive hemimethylated poly (dI-MedC).poly(dI-dC).  Using the latter substrate we have
purified an enzyme fraction 380-fold by salt extraction of chromatin, DEAE cellulose and
phosphocellulose.  This purified fraction showed enzyme activity only with poly
(dI-MedC).poly(dI-dC) thus suggesting the occurrence in plants of a DNA methyltransferase
specific for hemimethylated DNA.  A Mr value of 54,000 was calculated on the basis of the
sedimentation coefficient which was determined by sucrose density gradient centrifugation.
Apparent Km values for poly(dI-MedC).poly(dI-dC) and S-adenosyl-L-methionine were found to be
17 ug/ml and 2.6 uM, respectively.

<>

<1>Giovannelli, D. et al.
<2>Complete genome sequence of Thermovibrio ammonificans HB-1(T), a thermophilic, chemolithoautotrophic bacterium isolated from a deep-sea hydrothermal vent.
<3>Standards in Genomic Sciences
<4>7
<5>82-90
<6>2012
<7>type strain HB-1 is a thermophilic (T: 75 degrees C), strictly anaerobic,
chemolithoautotrophic bacterium that was isolated from an active, high
temperature deep-sea hydrothermal vent on the East Pacific Rise. This organism
grows on mineral salts medium in the presence of CO/H, using NO or S as electron
acceptors, which are reduced to ammonium or hydrogen sulfide, respectively. is
one of only three species within the genus , a member of the family , and it
forms a deep branch within the phylum . Here we report the main features of the
genome of strain HB-1 (DSM 15698).

<>

<1>Giovannelli, D., Ferriera, S., Johnson, J., Kravitz, S., Perez-Rodriguez, I., Ricci, J., O'Brien, C., Voordeckers, J.W., Bini, E., Vetriani, C.
<2>Draft genome sequence of Caminibacter mediatlanticus strain TB-2, an epsilonproteobacterium isolated from a deep-sea hydrothermal vent.
<3>Standards in Genomic Sciences
<4>5
<5>135-143
<6>2011
<7>Caminibacter mediatlanticus strain TB-2(T) [1], is a thermophilic, anaerobic,
chemolithoautotrophic bacterium, isolated from the walls of an active deep-sea
hydrothermal vent chimney on the Mid-Atlantic Ridge and the type strain of the
species. C. mediatlanticus is a Gram-negative member of the Epsilonproteobacteria
(order Nautiliales) that grows chemolithoautotrophically with H(2) as the energy
source and CO(2) as the carbon source. Nitrate or sulfur is used as the terminal
electron acceptor, with resulting production of ammonium and hydrogen sulfide,
respectively. In view of the widespread distribution, importance and
physiological characteristics of thermophilic Epsilonproteobacteria in deep-sea
geothermal environments, it is likely that these organisms provide a relevant
contribution to both primary productivity and the biogeochemical cycling of
carbon, nitrogen and sulfur at hydrothermal vents. Here we report the main
features of the genome of C. mediatlanticus strain TB-2(T).

<>

<1>Giovannoni, S.J., Tripp, H.J., Givan, S., Podar, M., Vergin, K.L., Baptista, D., Bibbs, L., Eads, J., Richardson, T.H., Noordewier, M., Rappe, M.S., Short, J.M., Carrington, J.C., Mathur, E.J.
<2>Genome streamlining in a cosmopolitan oceanic bacterium.
<3>Science
<4>309
<5>1242-1245
<6>2005
<7>The SAR11 clade consists of very small, heterotrophic marine alpha-proteobacteria that are
found throughout the oceans, where they
account for about 25% of all microbial cells. Pelagibacter ubique, the
first cultured member of this clade, has the smallest genome and encodes
the smallest number of predicted open reading frames known for a
free-living microorganism. In contrast to parasitic bacteria and archaea
with small genomes, P. ubique has complete biosynthetic pathways for all
20 amino acids and all but a few cofactors. P. ubique has no pseudogenes,
introns, transposons, extrachromosomal elements, or inteins; few paralogs;
and the shortest intergenic spacers yet observed for any cell.

<>

<1>Girault, G., Parisot, N., Peyretaillade, E., Peyret, P., Derzelle, S.
<2>Draft Genomes of Three Strains Representative of the Bacillus anthracis Diversity Found in France.
<3>Genome Announcements
<4>2
<5>e00736-14
<6>2014
<7>We report here the draft genomes of three Bacillus anthracis strains isolated in  France:
08-8_20 (A.Br.001/002), 99-100 (A.Br.011/009), and 00-82 (B.Br CNEVA).
The total lengths of assemblies are 5,440,708 bp, 5,446,472 bp, and 5,436,014 bp
for 08-8_20, 99-100, and 00-82, respectively.

<>

<1>Girlich, D., Poirel, L., Nordmann, P.
<2>A diversity of clavulanic acid-inhibited extended-spectrum {beta}-lactamases in Aeromonas sp. from the Seine River, Paris, France.
<3>Antimicrob. Agents Chemother.
<4>55
<5>1256-1261
<6>2011
<7>Environmental Aeromonas sp. isolates resistant to ceftazidime were
recovered during an environmental survey performed with water samples from
the Seine River, in Paris, France, in November 2009. Selected isolates
were identified by sequencing of the 16S rRNA and rpoB genes. PCR and
cloning experiments were used to identify
broad-spectrum-beta-lactamase-encoding genes and their genetic context.
Clavulanic acid-inhibited extended-spectrum-beta-lactamase (ESBL) genes
were identified in 71% of the Aeromonas sp. isolates. A variety of ESBL
genes were detected, including bla(VEB-1a), bla(SHV-12), bla(PER-1),
bla(PER-6), bla(TLA-2), and bla(GES-7), suggesting an aquatic reservoir of
those ESBL genes. Moreover, the repeated elements and different insertion
sequences were identified in association with the bla(PER-6) and the
bla(VEB-1a) genes, respectively, indicating a wide diversity of
mobilization events, making Aeromonas spp. a vehicle for ESBL
dissemination.

<>

<1>Girlich, D., Poirel, L., Szczepanowski, R., Schluter, A., Nordmann, P.
<2>Carbapenem-Hydrolyzing GES-5-Encoding Gene on Different Plasmid Types Recovered from a Bacterial Community in a Sewage Treatment Plant.
<3>Appl. Environ. Microbiol.
<4>78
<5>1292-1295
<6>2012
<7>Plasmids pRSB113 and pRSB115 were recovered from an activated sludge bacterial
community of a municipal wastewater treatment plant in Germany. Both plasmids
carry the same bla(GES-5) carbapenemase gene, located within two distinct class 1
integrons. These plasmids have different backbones, belong to different
incompatibility groups, and could replicate in both Pseudomonas aeruginosa and
Escherichia coli.

<>

<1>Gitschier, J.
<2>A Half-Century of Inspiration: An Interview with Hamilton Smith.
<3>PLoS Genet.
<4>8
<5>e1002466
<6>2012
<7>In 1962, Hamilton Smith abandoned a career in medicine to follow his passion for the emerging
field of molecular biology; within six years, he had made the discovery of a lifetime.  As a
new Johns Hopkins faculty member, Smith, together with his first graduate student, Kent
Wilcox, geared up to study recombination in vitro but instead discoverd the restriction enzyme
"R" in Haemophilus influenzae.  By cobbling together crude techniques, Smith, along with
Wilcox and later Tom Kelly, showed the R cleaves DNA at a specific recognition sequence, a
palindromic site, yielding blunt-ended DNA fragments.  Now known as HindII, R proved to be the
first of an enormous class of Type II restriction enzymes, and as such, presaged gene cloning,
allowed DNA to be reproducibly fragmented and then sequenced, and enabled physical mapping of
genomes.  Smith went on to discover DNA methylases that constitute the other half of the
bacterial host restriction and modification systems, as hypothesized by Werner Arber of
Switzerland.  Together with Arber and his Hopkins colleague Daniel Nathans, who first used the
enzyme on SV40 DNA and demonstrated discrete bands on a tube gel, Smith shared the Nobel Prize
for Physiology or Medicine in 1978.

<>

<1>Giuliani, M., Pizza, M., Rappuoli, R.
<2>Combination neisserial compositions.
<3>European Patent Office
<4>EP 2258851 A
<5>
<6>2010
<7>Compositions comprising a first biological molecule from a Neisseria bacterium and a second
biological molecule from a Neisseria bacterium.  The term "biological molecule" includes
proteins and nucleic acids.  Preferred Neisseria species are N. meningitidis and N.
gonorrhoeae.

<>

<1>Giuliani, M.M., Pizza, M., Rappuoli, R.
<2>Combination neisserial compositions.
<3>International Patent Office
<4>WO 2000071725 A
<5>
<6>2000
<7>Compositions comprising a first biological molecule from a Neisseria bacterium and a second
biological molecule from a Neiseria bacterium.  The term "biological molecule" includes
proteins and nucleic acids.  Preferred Neisseria species are N. meningitidis and N.
gonorrhoeae.

<>

<1>Giuliani, M.M., Pizza, M., Rappuoli, R.
<2>Combination neisserial compositions.
<3>European Patent Office
<4>EP 1860191 A
<5>
<6>2007
<7>Compositions comprising a first biological molecule from a Neisseria bacterium and a second
biological molecule from a Neisseria bacterium.  The term "biological molecule" includes
proteins and nucleic acids.  Preferred Neisseria species are N. meningitidis and N.
gonorrhoeae.

<>

<1>Gizard, Y., Zbinden, A., Schrenzel, J., Francois, P.
<2>Whole-Genome Sequences of Streptococcus tigurinus Type Strain AZ_3a and S. tigurinus 1366, a Strain Causing Prosthetic Joint Infection.
<3>Genome Announcements
<4>1
<5>e00210-12
<6>2013
<7>Streptococcus tigurinus, a novel member of the Streptococcus mitis group, was recently
identified as a causative agent of invasive infections. We report the
complete genome sequences of the S. tigurinus type strain AZ_3a and S. tigurinus
strain 1366. The genome sequences assist in the characterization of virulence
determinants of S. tigurinus.

<>

<1>Gladchenko, T.N., Zernov, Y.P.
<2>Method for isolation of the restriction endonuclease ApaI.
<3>Soviet Patent Office
<4>SU 1655986 A
<5>
<6>1991
<7>The present invention is directed to a method for purification of the ApaI restriction
endonuclease by 1) extraction of a lysate in a two-phase mixture of polyethyleneglycol,
dextran, and NaCl; 2) chromatography on phosphocellulose and hydroxylapatite. The yield of
ApaI was 51%, and specific activity was 150,000 units/mg with no contaminating nuclease
activity.

<>

<1>Gladitz, J., Shen, K., Antalis, P., Hu, F.Z., Post, J.C., Ehrlich, G.D.
<2>Codon usage comparison of novel genes in clinical isolates of Haemophilus influenzae.
<3>Nucleic Acids Res.
<4>33
<5>3644-3658
<6>2005
<7>A similarity statistic for codon usage was developed and used to compare novel gene sequences
found in clinical isolates of Haemophilus influenzae with a reference set of 80 prokaryotic,
eukaryotic and viral genomes. These analyses were performed to obtain an indication as to
whether individual genes were Haemophilus-like in nature, or if they probably had more
recently entered the H.influenzae gene pool via horizontal gene transfer from other species.
The average and SD values were calculated for the similarity statistics from a study of the
set of all genes in the H.influenzae Rd reference genome that encoded proteins of 100 amino
acids or longer. Approximately 80% of Rd genes gave a statistic indicating that they were most
like other Rd genes. Genes displaying codon usage statistics >1 SD above this range were
either considered part of the highly expressed group of H.influenzae genes, or were considered
of foreign origin. An alternative determinant for identifying genes of foreign origin was when
the similarity statistics produced a value that was much closer to a non-H.influenzae
reference organism than to any of the Haemophilus species contained in the reference set.
Approximately 65% of the novel sequences identified in the H.influenzae clinical isolates
displayed codon usages most similar to Haemophilus sp. The remaining novel sequences produced
similarity statistics closer to one of the other reference genomes thereby suggesting that
these sequences may have entered the H.influenzae gene pool more re

<>

<1>Gladney, L.M., Katz, L.S., Knipe, K.M., Rowe, L.A., Conley, A.B., Rishishwar, L., Marino-Ramirez, L., Jordan, I.K., Tarr, C.L.
<2>Genome Sequences of Vibrio navarrensis, a Potential Human Pathogen.
<3>Genome Announcements
<4>2
<5>e01188-14
<6>2014
<7>Vibrio navarrensis is an aquatic bacterium recently shown to be associated with human illness.
We report the first genome sequences of three V. navarrensis
strains obtained from clinical and environmental sources. Preliminary analyses of
the sequences reveal that V. navarrensis contains genes commonly associated with
virulence in other human pathogens.

<>

<1>Glaser, P. et al.
<2>Comparative genomics of Listeria species.
<3>Science
<4>294
<5>849-852
<6>2001
<7>Listeria monocytogenes is a food-borne pathogen with a high mortality rate that has also
emerged as a paradigm for intracellular parasitism. We present and compare the genome
sequences of L. monocytogenes (2,944,528 base pairs) and a nonpathogenic species, L. innocua
(3,011,209 base pairs). We found a large number of predicted genes encoding surface and
secreted proteins, transporters, and transcriptional regulators, consistent with the ability
of both species to adapt to diverse environments. The presence of 270 L. monocytogenes and 149
L. innocua strain-specific genes (clustered in 100 and 63 islets, respectively) suggests that
virulence in Listeria results from multiple gene acquisition and deletion events.

<>

<1>Glaser, P., Rusniok, C., Buchrieser, C., Chevalier, F., Frangeul, L., Msadek, T., Zouine, M., Couve, E., Lalioui, L., Poyart, C., Trieu-Cuot, P., Kunst, F.
<2>Genome sequence of Streptococcus agalactiae, a pathogen causing invasive neonatal disease.
<3>Mol. Microbiol.
<4>45
<5>1499-1513
<6>2002
<7>Streptococcus agalactiae is a commensal bacterium colonizing the intestinal tract of a
significant proportion of the human population.
However, it is also a pathogen which is the leading cause of invasive
infections in neonates and causes septicaemia, meningitis and pneumonia.
We sequenced the genome of the serogroup III strain NEM316, responsible
for a fatal case of septicaemia. The genome is 2,211,485 base pairs long
and contains 2118 protein coding genes. Fifty-five per cent of the
predicted genes have an ortholog in the Streptococcus pyogenes genome,
representing a conserved backbone between these two streptococci. Among
the genes in S. agalactiae that lack an ortholog in S. pyogenes, 50% are
clustered within 14 islands. These islands contain known and putative
virulence genes, mostly encoding surface proteins as well as a number of
genes related to mobile elements. Some of these islands could therefore be
considered as pathogenicity islands. Compared with other pathogenic
streptococci, S. agalactiae shows the unique feature that pathogenicity
islands may have an important role in virulence acquisition and in genetic
diversity.

<>

<1>Glaser, P., Rusniok, C., Chevalier, F., Frangeul, L., Lalioui, L., Zouine, M., Couve, E., Buchrieser, C., Poyart, C., Trieu-Cuot, P., Kunst, F.
<2>Streptococcus agalactiae genome sequence, use for developing vaccines, diagnostic tools, and for identifying therapeutic targets.
<3>International Patent Office
<4>WO 02092818 A
<5>
<6>2002
<7>The invention concerns the genome sequence and nucleotide sequences coding for Streptococcus
agalactiae polypeptides, such as cellular envelope polypeptides, or secreted or specific
polypeptides, or polypeptides involved in the metabolism and the replication process, as well
as vectors or cells comprising said sequences.  The invention also concerns the use thereof
for developing vaccines, diagnostic tools, DNA chips and for identifying therapeutic targets.

<>

<1>Glasner, J.D. et al.
<2>Genome Sequence of the Plant-Pathogenic Bacterium Dickeya dadantii 3937.
<3>J. Bacteriol.
<4>193
<5>2076-2077
<6>2011
<7>Dickeya dadantii is a plant-pathogenic enterobacterium responsible for the soft rot disease of
many plants of economic importance. We present here
the sequence of strain 3937, a strain widely used as a model system for
research on the molecular biology and pathogenicity of this group of
bacteria.

<>

<1>Glasner, J.D., Marquez-Villavicencio, M., Kim, H.S., Jahn, C.E., Ma, B., Biehl, B.S., Rissman, A.I., Mole, B., Yi, X., Yang, C.H., Dangl, J.L., Grant, S.R., Perna, N.T., Charkowski, A.O.
<2>Niche-specificity and the variable fraction of the Pectobacterium pan-genome.
<3>Mol. Plant Microbe Interact.
<4>21
<5>1549-1560
<6>2008
<7>We compare genome sequences of three closely related soft-rot pathogens that vary
in host range and geographical distribution to identify genetic differences that
could account for lifestyle differences. The isolates compared, Pectobacterium
atrosepticum SCRI1043, P. carotovorum WPP14, and P. brasiliensis 1692, represent
diverse lineages of the genus. P. carotovorum and P. brasiliensis genome contigs,
generated by 454 pyrosequencing ordered by reference to the previously published
complete circular chromosome of P. atrosepticum genome and each other, account
for 96% of the predicted genome size. Orthologous proteins encoded by P.
carotovorum and P. brasiliensis are approximately 95% identical to each other and
92% identical to P. atrosepticum. Multiple alignment using Mauve identified a
core genome of 3.9 Mb conserved among these Pectobacterium spp. Each core genome
is interrupted at many points by species-specific insertions or deletions
(indels) that account for approximately 0.9 to 1.1 Mb. We demonstrate that the
presence of a hrpK-like type III secretion system-dependent effector protein in
P. carotovorum and P. brasiliensis and its absence from P. atrosepticum is
insufficient to explain variability in their response to infection in a plant.
Additional genes that vary among these species include those encoding peptide
toxin production, enzyme production, secretion proteins, and antibiotic
production, as well as differences in more general aspects of gene regulation and
metabolism that may be relevant to pathogenicity.

<>

<1>Glasner, W., Hennecke, F., Fritz, H.J.
<2>Enzymatic properties and biological functions of VSR DNA mismatch endonuclease.
<3>Structural tools for the analysis of protein-nucleic acid complexes advances in life sciences., Birkhauser Verlag, , Basel
<4>0
<5>165-173
<6>1992
<7>The protein encoded by the vsr gene of E. coli K-12 was produced by construction and
expression of a chimeric gene. The corresponding fusion protein contains b-lactamase as the
N-terminal and the Vsr gene product as the C-terminal component. Enzymological studies of the
fusion protein reveal the Vsr gene product as the first known example of a DNA mismatch
endonuclease. Its substrate recognition properties are characterized by a combination of
sequence and structure specificity as follows. Vsr endonuclease recognizes T/G mismatches in
sequence contexts related to the cognate sequence of Dcm DNA cytosine methyltransferase and
cleaves the phosphodiester bond on the 5' side of the mismatched T residue, forming a
phosphorylated 5' end (Hennecke et al. 1991, Nature 353: 775-778). Studies concerning the
influence of individual functional groups of the mismatched bases on substrate recognition and
the detailed requirements of DNA sequence context are reported. The enzymology of Vsr
endonuclease is summarized and its biological role is discussed.

<>

<1>Glasner, W., Merkl, R., Schellenberger, V., Fritz, H.J.
<2>Substrate preferences of Vsr DNA mismatch endonuclease and their consequences for the evolution of the Escherichia coli K-12 genome.
<3>J. Mol. Biol.
<4>245
<5>1-7
<6>1995
<7>The substrate spectrum of Vsr DNA mismatch endonuclease of Escherichia coli K-12 was
investigated using fluorescence-labelled oligonucleotide substrates and a DNA sequencer for
detection and quantification of substrates and reaction products. Fourteen substrates were
found to be processed by the enzyme, which differ in one or two positions from the canonical
pentanucleotide sequence CTA/TGG (T mismatched to G). Relative second-order rate constants of
these substrates were determined in groups of four by multiple substrate kinetics and compared
to the underrepresentation of the corresponding pentanucleotides in the E. coli K-12 genome.
The high quality of correlation further establishes active mutagenesis by VSP repair as a
significant driving force of the evolution of the E. coli K-12 genome and provides clues to
its possible selective value.

<>

<1>Glass, J.I., Lefkowitz, E.J., Glass, J.S., Heiner, C.R., Chen, E.Y., Cassell, G.H.
<2>The complete sequence of the mucosal pathogen Ureaplasma urealyticum.
<3>Nature
<4>407
<5>757-762
<6>2000
<7>The comparison of the genomes of two very closely related human mucosal pathogens, Mycoplasma
genitalium and Mycoplasma pneumoniae, has helped define the essential functions of a
self-replicating minimal cell, as well as what constitutes a mycoplasma.  Here we report the
complete sequence of a more distant phylogenetic relative of those bacteria, Ureaplasma
urealyticum (parvum biovar), which is also a mucosal pathogen of humans.  It is the third
mycoplasma to be sequenced, and has the smallest sequenced prokaryotic genome except for M.
genitalium.  Although the U. urealyticum genome is similar to the two sequenced mycoplasma
genomes, features make this organism unique among mycoplasmas and all bacteria.  Almost all
ATP synthesis is the result of urea hydrolysis, which generates an energy-producing
electrochemical gradient.  Some highly conserved eubacterial enzymes appear not to be encoded
by U. urealyticum, including the cell-division protein FtsZ, chaperonins GroES and GroEL, and
ribonucleoside-diphosphate reductase.  U. urealyticum has six closely related iron
transporters, which apparently arose through gene duplication, suggesting that it has a kind
of respiration system not present in other small genome bacteria.  The genome is only 25.5%
G+C in nucleotide content, and the G+C content of individual genes may predict how essential
those genes are to ureaplasma survival.

<>

<1>Glatman, L.I., Iablokova, M.B., Kravets, A.I., Terekhov, A.A., Samoilenko, I.I.
<2>Identification of the plasmid pLG13 coding for the DNA modification-restriction system EcoRV and properties of strains carrying this plasmid.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>10
<5>39-42
<6>1985
<7>
<>

<1>Glatman, L.I., Kravets, A.N.
<2>Systems of DNA restriction-modification.
<3>Antibiot. Khimioter
<4>34
<5>932-938
<6>1989
<7>None

<>

<1>Glatman, L.I., Moroz, A.F., Iablokova, M.B., Rebentish, B.A., Kholmina, G.V.
<2>New DNA modification-restriction plasmid system detected in a clinical strain of Escherichia coli.
<3>Dokl. Akad. Nauk.
<4>252
<5>993-995
<6>1980
<7>
<>

<1>Glatman, L.I., Moroz, A.F., Yablokova, M.B., Rebentish, B.A., Kcholmina, G.V.
<2>A novel plasmid-mediated DNA restriction-modification system in E. coli.
<3>Plasmid
<4>4
<5>350-351
<6>1980
<7>R plasmids from 101 clinical isolates were transferred to E. coli J62 by conjugation and
tested for the presence of R plasmid-mediated restriction-modification DNA systems.  Thirty R
plasmids were found to inhibit phage lambda vir development.  Ten plasmids determined
restriction-modification system; nine of them proved identical with R.M. EcoRII.  One
transconjugant, E. coli J62 pLG74, was shown to have a restriction-modification system
different from all the known R plasmid-mediated systems.  Site-specific endonuclease has been
isolated from E. coli J62 pLG74 which differed from all the known restriction endonucleases in
the number of cleavage sites on phages lambda, phiX 174, virus SV40, plasmid pBR322 DNA
molecules.

<>

<1>Glatman, L.I., Terekhov, A.A., Kalnin, K.V., Bolotin, A.P., Rebentish, B.A.
<2>New site-specific endodesoxyribonuclease EcoHI.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>3
<5>32
<6>1990
<7>A new Type II site-specific endonuclease, EcoHI, has been isolated from a strain of
Escherichia coli and characterized.  Restriction endonuclease EcoHI recognizes the nucleotide
sequence C C (C/G) G'G with the cleavage site between the fourth and fifth nucleotides.  It
is an isoschizomer of the restriction endonuclease CauII.  The yield of enzyme is 2500 units
of activity per 1 g of biomass.  The producing strain Escherichia coli HI is nonpathogenic,
easily grown with the antiobiotic resistance markers allowing the strain to be cultivated
under selective conditions.

<>

<1>Glaub, A., Bomholt, C., Gravermann, K., Brinkrolf, K., Albersmeier, A., Ruckert, C., Tauch, A.
<2>Complete Genome Sequence of Corynebacterium falsenii DSM 44353 To Study the Evolution of Corynebacterium Cluster 3 Species.
<3>Genome Announcements
<4>2
<5>e00158-14
<6>2014
<7>Corynebacterium falsenii is a member of the natural microflora of wild and domesticated birds
and is rarely detected in human clinical specimens. The
chromosomal sequence of the type strain C. falsenii DSM 44353 comprises 2,677,607
bp and provides detailed insights into the evolution of Corynebacterium species
assigned to the highly diverse cluster 3.

<>

<1>Glavina, D.Rio.T. et al.
<2>Complete genome sequence of Chitinophaga pinensis type strain (UQM 2034).
<3>Standards in Genomic Sciences
<4>2
<5>87-95
<6>2010
<7>Chitinophaga pinensis Sangkhobol and Skerman 1981 is the type strain of the species which is
the type species of the rapidly growing genus Chitinophaga in
the sphingobacterial family 'Chitinophagaceae'. Members of the genus Chitinophaga
vary in shape between filaments and spherical bodies without the production of a
fruiting body, produce myxospores, and are of special interest for their ability
to degrade chitin. Here we describe the features of this organism, together with
the complete genome sequence, and annotation. This is the first complete genome
sequence of a member of the family 'Chitinophagaceae', and the 9,127,347 bp long
single replicon genome with its 7,397 protein-coding and 95 RNA genes is part of
the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Glenn, T.C., Waller, D.R., Braun, M.J.
<2>Increasing proportions of uracil in DNA substrates increases inhibition of restriction enzyme digests.
<3>Biotechniques
<4>17
<5>1086-1090
<6>1994
<7>Techniques that rely upon the incorporation of uracil into DNA are being published with
increasing frequency, especially in PCR protocols. We report here the efficiency of 18 type II
restriction enzymes to digest PCR amplicons synthesized with varying proportions of TTP to
dUTP in the PCR mixture. We find that most enzymes with A:T/U bp in their recognition site
digest the amplicons less efficiently as the percentage of dUTP in the reaction mixture is
increased. This effect is most dramatic when the proportion of dUTP in the nucleotide mixture
exceeds 50%. All but one of the enzymes which fail to digest amplicons that are synthesized
with 100% dUTP digest some amplicons which are synthesized with 90% dUTP.

<>

<1>Glickman, J.F.
<2>The murine DNA methyltransferase: Purification, heterologous expression, steady-state kinetics and post-translational processing.
<3>Diss. Abstr.
<4>58
<5>1265B
<6>1997
<7>The mammalian DNA methyltransferase catalyzes the transfer of methyl groups from S-adenosyl
methionine to the C5-cytosine within the d(CpG) dinucleotides of double-stranded DNA.  DNA
methylation is critical for normal embryonic development, and regulates gene expression by
controlling the binding of proteins to the regulatory regions of DNA.  Despite the biological
importance of this enzyme, the DNA methyltransferase itself has not been characterized in
detail, with respect to size, steady-state kinetic parameters, post-translational
modifications and N-terminal sequence.  This lack of information has been due to the
difficulty in  the purification of suitable quantities of the protein for enzymological and
biochemical studies.  The murine erthroleukemia cell-derived DNA methyltransferase (300-800
micrograms) of 3 x 10^10 cells grown in 10 liter cultures.  The recombinant enzyme was
expressed as a fusion protein containing a nickel-affinity leader peptide using a baculovirus
expression system.  Expression was 50-fold higher per cell than in the MEL-cells.  The
recombinant enzyme (1-2mg) was purified from 5 x 10^8 Sf21 cells using immobilized nickel
affinity chromatography.  The steady-state kinetic parameters of kcat, Km for the substrate,
double-stranded poly(dI-dC) and the cofactor, S-adenosylmethionine were determined for both
the MEL cell-derived and the recombinant DNA methyltransferases, demonstrating that the two
enzymes were functionally similar.  The DNA methyltransferase was a noticeably slow enzyme
(kcat, 7/hr); however, based on an estimate of the number of DNA methyltransferase molecules
per nucleus, there should be enough methyltransferase activity to account for the levels of
DNA methylation in a vertebrate cell.  Metabolic labeling experiments revealed that the DNA
methyltransferase was post-translationally phosphorylated on serine/threonine residues, and
peptide mapping using HPLC-electrospray mass spectrometry resulted in the identification of a
single, predominant phosphorylation site on the MEL-derived DNA methyltransferase in the
domain responsible for targeting of the enzyme to the replication foci.  Data demonstrating a
single start of translation at the first methionine, as revised by a newly reported cDNA, is
presented.

<>

<1>Glickman, J.F., Flynn, J., Reich, N.O.
<2>Purification and characterization of recombinant baculovirus-expressed mouse DNA methyltransferase.
<3>Biochem. Biophys. Res. Commun.
<4>230
<5>280-284
<6>1997
<7>DNA methylation is essential for normal embryonic development in mice.  An understanding of
how DNA methylation is controlled is largely dependent upon the isolation and characterization
of the cellular components of the DNA methylation system.  The enzyme which methylates DNA in
eukaryotic cells is a C-5 cytosine DNA methyltransferase.  Historically, the characterization
of this enzyme has been limited by its availability and purity.  Here, we present a
single-step purification of 4 mg of baculovirus-expressed mouse DNA methyltransferase
containing a nickel-affinity leader peptide.  The recombinant DNA methyltransferase copurified
with inhibitory RNA, which was removed by treatment with ribonuclease A.  Like its
non-recombinant counterpart, the recombinant enzyme is activated by hemi-methylation.  A
direct steady-state kinetic comparison between the recombinant baculovirus-expressed enzyme
with its MEL cell-derived counterpart is presented.

<>

<1>Glickman, J.F., Pavlovich, J.G., Reich, N.O.
<2>Peptide mapping of the murine DNA methyltransferase reveals a major phosphorylation site and the start of translation.
<3>J. Biol. Chem.
<4>272
<5>17851-17857
<6>1997
<7>The murine DNA methyltransferase catalyzes the transfer of methyl groups from
S-adenosylmethionine to cytosines within d(CpG) dinucleotides.  The enzyme is necessary for
normal embryonic development and is implicated in a number of important processes, including
the control of gene expression and cancer.  Metabolic labeling and high pressure liquid
chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS) were performed on DNA
methyltransferase purified from murine erythroleukemia cells.  Serine 514 was identified as a
major phosphorylation site that lies in a domain required for targeting of the enzyme to the
replication foci.  These results present a potential mechanism for the regulation of DNA
methylation.  HPLC-ESI-MS peptide mapping data demonstrated that the purified murine DNA
methyltransferase protein contains the N-terminal regions predicted by the recently revised
5' gene sequences.  The evidence suggests a start of translation at the first predicted
methionine, with no alternate translational start sites.  Our peptide mapping results provide
a more detailed structural characterization of the DNA methyltransferase that will facilitate
future structure/function studies.

<>

<1>Glickman, J.F., Reich, N.O.
<2>The expression of the murine DNA cytosine methyltransferase in cultured cells from Spodoptera frugiperda using a recombinant Baculovirus.
<3>FASEB J.
<4>7
<5>A1291
<6>1993
<7>The murine DNA methyltransferase (MTase, EC 2.1.1.371) is a 169.6 kD enzyme which transfers
methyl groups from the S-adenosyl methionine cofactor to the 5' carbon of cytosine in DNA.
DNA methylation in mammals has been implicated in the regulation of gene expression,
immunoglobulin gene re-arrangement, X-chromosome inactivation and carcinogenesis. We have
expressed the MTase in cultured insect cells using a lethal linear baculovirus system. Our
data indicates 100-fold enhancement in MTase expression over previous methods. This system
will enable us to perform more precise enzymology on the MTase because prior studies were
limited by the quantity of purified enzyme available. To our knowledge, this is the largest
nuclear protein expressed using the baculovirus system. We are currently optimizing the
expression system and investigating the role of post-translational modification and
sub-cellular localization in the regulation of function of MTase function.

<>

<1>Glickman, J.F., Reich, N.O.
<2>Baculovirus-mediated high level expression of a mammalian DNA methyltransferase.
<3>Biochem. Biophys. Res. Commun.
<4>204
<5>1003-1008
<6>1994
<7>The murine C-5 cytosine DNA methyltransferase (Mtase, E.C.2.1.1.37) containing a hexahistidine
affinity leader peptide has been expressed at levels which are at least 50-fold higher than
previously reported.  The recombinant enzyme has activity levels similar to the wild-type
enzyme.  The recombinant polypeptide binds to and elutes from a nickel affinity resin (IMAC
resin).  No dramatic differences in post-translational modification between the wild-type and
recombinant enzyme were observed.  The recombinant system will be useful in performing
site-directed mutagenesis and will facilitate enzymological and biological investigations of
this enzyme.

<>

<1>Glockner, G., Lehmann, R., Romualdi, A., Pradella, S., Schulte-Spechtel, U., Schilhabel, M., Wilske, B., Suhnel, J., Platzer, M.
<2>Comparative analysis of the Borrelia garinii genome.
<3>Nucleic Acids Res.
<4>32
<5>6038-6046
<6>2004
<7>Three members of the genus Borrelia (B.burgdorferi, B.garinii, B.afzelii) cause tick-borne
borreliosis. Depending on the Borrelia species involved,
the borreliosis differs in its clinical symptoms. Comparative genomics
opens up a way to elucidate the underlying differences in Borrelia
species. We analysed a low redundancy whole-genome shotgun (WGS) assembly
of a B.garinii strain isolated from a patient with neuroborreliosis in
comparison to the B.burgdorferi genome. This analysis reveals that most of
the chromosome is conserved (92.7% identity on DNA as well as on amino
acid level) in the two species, and no chromosomal rearrangement or larger
insertions/deletions could be observed. Furthermore, two collinear
plasmids (lp54 and cp26) seem to belong to the basic genome inventory of
Borrelia species. These three collinear parts of the Borrelia genome
encode 861 genes, which are orthologous in the two species examined. The
majority of the genetic information of the other plasmids of
B.burgdorferii is also present in B.garinii although orthology is not easy
to define due to a high redundancy of the plasmid fraction. Yet, we did
not find counterparts of the B.burgdorferi plasmids lp36 and lp38 or their
respective gene repertoire in the B.garinii genome. Thus, phenotypic
differences between the two species could be attributable to the presence
or absence of these two plasmids as well as to the potentially positively
selected genes.

<>

<1>Gloeckner, F.O., Kube, M., Bauer, M., Teeling, H., Lombardot, T., Ludwig, W., Gade, D., Beck, A., Borzym, K., Heitmann, K., Rabus, R., Schlesner, H., Amann, R., Reinhardt, R.
<2>Complete genome sequence of the marine planctomycete Pirellula sp. strain 1.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>8298-8303
<6>2003
<7>Pirellula sp. strain 1 ("Rhodopirellula baltica") is a marine representative of the globally
distributed and environmentally important
bacterial order Planctomycetales. Here we report the complete genome
sequence of a member of this independent phylum. With 7.145 megabases,
Pirellula sp. strain 1 has the largest circular bacterial genome sequenced
so far. The presence of all genes required for heterolactic acid
fermentation, key genes for the interconversion of C1 compounds, and 110
sulfatases were unexpected for this aerobic heterotrophic isolate.
Although Pirellula sp. strain 1 has a proteinaceous cell wall, remnants of
genes for peptidoglycan synthesis were found. Genes for lipid A
biosynthesis and homologues to the flagellar L- and P-ring protein
indicate a former Gram-negative type of cell wall. Phylogenetic analysis
of all relevant markers clearly affiliates the Planctomycetales to the
domain Bacteria as a distinct phylum, but a deepest branching is not
supported by our analyses.

<>

<1>Gloeckner, G., Schulte-Spechtel, U., Schilhabel, M., Felder, M., Suehnel, J., Wilske, B., Platzer, M.
<2>Comparative genome analysis: Selection pressure on the Borrelia vls cassettes is essential for infectivity.
<3>BMC Genomics
<4>7
<5>211
<6>2006
<7>ABSTRACT: BACKGROUND: At least three species of Borrelia burgdorferi sensu lato (Bbsl) cause
tick-borne Lyme disease. Previous work including the
genome analysis of B. burgdorferi B31 and B. garinii PBi suggested a
highly variable plasmid part. The frequent occurrence of duplicated
sequence stretches, the observed plasmid redundancy, as well as the mainly
unknown function and variability of plasmid encoded genes rendered the
relationships between plasmids within and between species largely
unresolvable. RESULTS: To gain further insight into Borreliae genome
properties we completed the plasmid sequences of B. garinii PBi, added the
genome of a further species, B. afzelii PKo, to our analysis, and compared
for both species the genomes of pathogenic and apathogenic strains. The
core of all Bbsl genomes consists of the chromosome and two plasmids
collinear between all species. We also found additional groups of
plasmids, which share large parts of their sequences. This makes it very
likely that these plasmids are relatively stable and share common
ancestors before the diversification of Borrelia species. The analysis of
the differences between B. garinii PBi and B. afzelii PKo genomes of low
and high passages revealed that the loss of infectivity is accompanied in
both species by a loss of similar genetic material. Whereas B. garinii PBi
suffered only from the break-off of a plasmid end, B. afzelii PKo lost
more material, probably an entire plasmid. In both cases the vls gene
locus encoding for variable surface proteins is affected. CONCLUSIONS: The
complete genome sequences of a B. garinii and a B. afzelii strain
facilitates further comparative studies within the genus Borrellia. Our
study shows that loss of infectivity can be traced back to only one single
event in B. garinii PBi: the loss of the vls cassettes possibly due to
error prone gene conversion. Similar albeit extended losses in B. afzelii
PKo support the hypothesis that infectivity of Borrelia species depends
heavily on the evasion from the host response.

<>

<1>Glover, S.W.
<2>Restriction Endonucleases.  Genetics of restriction/modification systems.
<3>Biochem. Soc. Trans.
<4>8
<5>395-400
<6>1980
<7>Three different types of restriction/modification system are now recognized: type I, type II
and type III.  The genetics of type-I systems has been investigated by several groups of
workers and there is common agreement that three genes are involved: hsdS, hsdR and hsdM.  The
specificity of the system is determined by the product of the hsdS gene.  Mutations in the
hsdS gene produce an r-m- phenotype; mutations in the hsdR gene produce an r-m+ phenotype and
mutations in the hsdM gene produce an r-m- phenotype.  Several such systems have been
recognized and analysed genetically in Escherichia coli and in Salmonella typhimurium.
Complementation tests employing partial diploids constructed by using F' strains show that
the E. coli systems K and B and the S. typhimurium system SB are functionally related, and
mapping experiments show that they are genetically allelic.  It has been shown that
restriction endonucleases isolated from type-I systems have the three different polypeptide
subunits that the three-gene model predicts, and that type-I-modification methylases have the
two subunits expected from the genetic model.  However, unambiguous assignment of polypeptide
subunit to coding gene is still lacking. No convincing evidence for regulatory-gene functions
has been obtained so far.

<>

<1>Glover, S.W.
<2>Functional analysis of host-specificity mutants in Escherichia coli.
<3>Genet. Res.
<4>15
<5>237-250
<6>1970
<7>Evidence from a functional analysis of host-specificity mutants in merodiploids
is presented which supports the suggestion that three genes, hss, hsr and hsm,
are necessary for the expression of host-controlled restriction and
modification.  The host-specificity phenotype expressed by the merodiploids
provides evidence that at least two genes, hss and hsr, are concerned in the
expression of host-specific restriction of DNA and one of these genes, hss, is
responsible for the strain specificity of the restriction enzyme.  A class of
modification-deficient mutants isolated from restriction-deficient,
modification-proficient mutants, was also tested for complementation in
merodiploids and the phenotype of these merodiploids provides evidence that at
least two genes, hss and hsm, are concerned in the expression of host-specific
modification of DNA and one of these genes, hss, is responsible for the strain
specificity of the modification enzyme.  How these three genes function at the
molecular level is discussed in terms of models based on the interaction of
subunits to form oligomeric enzymes.

<>

<1>Glover, S.W.
<2>Host specificity in F' heterogenotes of Escherichia coli.
<3>J. Gen. Microbiol.
<4>53
<5>i-ii
<6>1968
<7>Host-controlled modification of DNA in strains of Escherichia coli has two
clearly defined characteristics.  First, modification, a process which acts
directly on DNA and involves the specific alteration of certain base sequences
by methylation.  As a result, the DNA synthesized in a particular strain may
carry a characteristic, strain-specific pattern.  Secondly, restriction, a
process which may lead to the rapid degradation of DNA introduced into a cell
of different host strain specificity.  The host specificities of E. coli strain
K12 and E. coli strain B are different such that DNA from K12 may be degraded
in B and vice versa.  In both strains mutants have been isolated which either
have lost the ability to restrict but are still able to modify DNA, or have
lost both the ability to restrict and to modify DNA.  The genetic location of
both these mutations has been determined in E. coli K and E. coli B.  Both
mutations map in the thr-leu region.  An F' factor from E. coli K12 carrying
thr-leu and the genes determining host specificity has been used to examine the
host-specificity properties of diploids.  Restriction was assayed simply by
measuring the efficiency of plating of bacteriophage lambda grown on either E.
coli K or E. coli B and designated lambda.K and lambda.B respectively.
Modification was assayed again indirectly by growing phage lambda on the
diploid strains and measuring its efficiency of plating on standard indicator
strains.  The results obtained so far indicate that:  1) In strain K the
wild-type alleles are dominant to both of the mutants described above.  2) The
diploid K/K may plate lambda.B less efficiently than the haploid strain K.  3)
The diploid K/B restricts both lambda.K and lambda.B.  4) The diploid K/B
produces lambda particles which are able to plate on both strains K and B.  5)
The diploid constructed between K and a mutant of B deficient in both
restriction and modification is indistinguishable from the haploid K strain.
6) The diploid constructed between K and a mutant of B deficient in restriction
only, restricts both lambda.K and lambda.B and produces lambda particles which
are able to plate on both strains K and B.  These results will be discussed in
relation to the several models which have been proposed to explain the genetic
basis of host-controlled modification of DNA.

<>

<1>Glover, S.W.
<2>Genetics of the R-M systems in Haemophilus.
<3>Heredity
<4>41
<5>123
<6>1978
<7>The genus Haemophilus is an extremely rich source of restriction endonucleases of both type I
and type II.  Altogether 22 such enzymes have been detected and partially characterized.
About half of these enzymes have been isolated from Haemophilus influenzae and each of the H.
influenzae serotypes a, b, c, d, e and f are believed to possess restriction and modification
(R-M) systems.  The R-M systems from serotypes a, b, d, e and f have been analyzed genetically
and the restriction endonucleases from serotypes a, f (HinaII, HinaIII and HinfIII) have been
partially characterized.  All of the evidence indicates that the enzymes which form part of
the H. influenzae R-M systems are type I enzymes and their role appears to be similar to that
of other restriction endonucleases that can be detected by in vivo tests.  Type II restriction
endonucleases have been isolated from all of the H. influenzae serotypes other than serotype a
and characterized using bacteriophage lambda or other well-defined DNA molecules as substrate.
These enzymes do not appear to form part of recognized R-M systems and they have little or no
activity on Haemophilus phage DNA.

<>

<1>Glover, S.W., Colson, C.
<2>Genetics of host-controlled restriction and modification in Eschericia coli.
<3>Genet. Res.
<4>13
<5>227-240
<6>1969
<7>In recent years many aspects of the processes of restriction and modification
which characterize the host controlled modification (CM) of bacteriophages in
Escherica coli have been clarified (reviewed by Arber (1965a) and Klein
(1965)).  In this paper we describe:  (i) the results of genetic experiments
which locate the sites of these mutations close to the serB locus, (ii) the
results of genetic crosses between different HCM mutants, and (iii) the results
of experiments designed to elucidate the number of functional units involved in
the control of HCM and the relationships between them.

<>

<1>Glover, S.W., Colson, C.
<2>Stable and unstable alterations of the host-induced modification properties of Escherichia coli B, K and C.
<3>Genet. Res.
<4>7
<5>223-234
<6>1966
<7>The system of host-induced modification (HIM) in E. coli B, K and C (Arber &
Dussoix, 1962) has the following properties.  Normal K, which is able to
restrict phage lambda grown in either B or C and confers a K-specific
modification to lambda-DNA synthesized in it, can be represented as K r+m+.
Similarly, normal B can be represented as B r+m+, indicating that it restricts
phage lambda grown in either K or C and that it confers a B-specific
modification to lambda-DNA synthesized in it.  Strain C accepts phage lambda
grown in K, B or C without restriction.  Recently the genes controlling
restriction and modification have been mapped close to the marker threonine and
on the opposite side of it to leucine (Colson, Glover, Symonds & Stacey, 1965).
In the course of this work, strains were isolated which had HIM properties
unlike those of B, K or C.  These strains display different patterns of HIM.
One such pattern can be represented as B r-m+, indicating that the ability to
restrict lambda grown in K or C has been lost while the ability to confer the
B-specific modification is retained.  Another can be represented as K r-m- or B
r-m-:  these strains have lost the ability to restrict lambda.C and to confer
respectively either the K- or the B-specific modificatiton.  Others are more
complex and comprise strains which on first isolation appear to bear HIM
properties intermediate between those of B,K and C, but which on further
analysis turn out to be unstable.  We shall describe the isolation of these
strains and present some details of their behaviour.

<>

<1>Glover, S.W., Colson, C.
<2>The breakdown of the restriction mechanism in zygotes of Escherichia coli.
<3>Genet. Res.
<4>6
<5>153-155
<6>1965
<7>A few cells in a culture of E. coli K or E. coli B, usually about 1 in 10-4,
permit the growth of the restricted phage lambda.C (Arber & Dussoix, 1962).
Such cells may be genetically different from the majority of the cells in the
culture or they may be temporarily in such a physiological condition that the
restriction mechanism breaks down or cannot be expressed.  Mutants which are
genetically unable to restrict the growth of phage normally restricted by
wild-type cells have been isolated (Glover et al., 1963; Colson et al., 1964;
Wood, 1964).  But several environmental factors are known which decrease the
capacity of wild-type bacteria to restrict the growth of unmodified phage
particles (Luria, 1953; Lederberg, 1957; Uetake et al., 2964).  We shall report
here observations which suggest that E. coli zygotes are in a special
physiological condition such that their ability to restrict phage is reduced by
more than a factor of 100.

<>

<1>Glover, S.W., Firman, K.
<2>A novel class of restriction-deficient mutants.
<3>Heredity
<4>41
<5>122-123
<6>1978
<7>R124 is a unique R factor in compatability group F IV which confers tetracycline resistance
(Tc) and carries the genes for a restriction-modification (R-M) system.  R124/3 is a
derivative plasmid which determines a R-M system of different biological specificity to R124.
During experiments designed to isolate restriction-deficient (r-) mutants of these R factors
in which F-lac+.0 was transferred to R124 or R124/3 carrying bacteria virtually all of the
colonies isolated were restriction-deficient.  Restriction-deficient mutants can be isolated
just as readily following transfer of these R factors to F-lac+ carrying bacteria.  Many of
the restriction-deficient mutants isolated are also modification-deficient and some
unexpectedly express the modifications characteristic of both R124 and R124/3.  Evidence is
presentd to indicate that the restriction-deficient phenotype of these R factors is not
dependent on the continued presence of F-lac+ which lead to their isolation and that other
F-prime factors can interact with R124 to produce r- mutants also at very high frequency.
Many of these r- mutants show high "reversion" frequencies to r+ and evidence is presented to
indicate that mutation to Tc sensitivity may also arise frequently following transfer to
F-lac+ to R+ bacteria.  These results are discussed in relation to what is known about the DNA
of the plasmid molecules from studies conducted by Dr. S.G. Hughes.

<>

<1>Glover, S.W., Firman, K.
<2>Genetic control of DNA specificity.
<3>Genetic Exchange, Marcel Dekker, Streips, U.N., Goodgal, S.H., Guild, W.R., Wilson, G.A., New York
<4>0
<5>331-338
<6>1982
<7>The plasmid R124 codes for a unique restriction and modification (RM) system and the
derivative plasmid R124/3 codes for an RM system with a different DNA specificity.  Evidence
has been obtained that the genes coding for each RM system are carried by both R124 and R124/3
and that normally only one set of genes is expressed.  The mechanism that ensues that only one
set of genes is expressed has been investigated in a series of experiments in which a plasmid
expressing the genes for the R124 RM system and a plasmid expressing the genes for the R124/3
can be transposed to F plasmids that are compatible with the R factors.  The results of these
experiments indicate that a trans-acting regulatory mechanism coded for by a resident plasmid
switches off the expression of RM genes on an introduced plasmid.  Evidence is also presented
for an additional trans-acting regulatory mechanism which can switch on the expression of the
alternative set of genes carried by the introduced plasmid.  The regulatory mechanism(s)
controlling the switch in expression of R124 and R124/3 RM genes involves a physical
rearrangement of DNA segments, and a possible model for this regulatory mechanism is
discussed.

<>

<1>Glover, S.W., Firman, K., Watson, G., Price, C., Donaldson, S.
<2>The alternate expression of two restriction and modification systems.
<3>Mol. Gen. Genet.
<4>190
<5>65-69
<6>1983
<7>Plasmids R124 and R124/3 carry genes coding for two different R-M systems and
normally only one set of genes is expressed.  These genes can be translocated
to F plasmids that are compatible with the R factors and in strains carrying
these F plasmids and an R factor a transacting regulatory mechanism switches
off the expression of R-M genes on the introduced plasmid.  Additionally the
unexpressed genes on the introduced plasmid are expressed.  The regulatory
mechanism controlling the alternative expression of R124 and R124/3 R-M genes
involves a physical rearrangement of DNA sequences.

<>

<1>Glover, S.W., Piekarowicz, A.
<2>Host specificity of DNA in Haemophilus influenzae:  Restriction and modification in strain Rd.
<3>Biochem. Biophys. Res. Commun.
<4>46
<5>1610-1617
<6>1972
<7>Wild type strains of Haemophilus influenzae Rd consist of two phenotypic
classes of bacteria which differ in their abilities to restrict and modify
phage HP1.  Each of these classes r- m- and r+ m+ is unstable and segregates
several percent of bacteria of the alternative phenotype.  In addition stable
r- m- bacteria occur spontaneously in r+ m+ cultures and an r- m+ mutant was
isolated after mutagenesis of an r+ m+ strain.

<>

<1>Glover, S.W., Schell, J., Symonds, N., Stacey, K.A.
<2>The control of host-induced modification by phage P1.
<3>Genet. Res.
<4>4
<5>480-482
<6>1963
<7>The phenomenon of host-induced modification has two facets, restriction and
modification.  Recently these two processes have been clarified by Arber and
Dussoix in the system where phage lambda multiplies either in Escherichia coli
K12, or in K12 cells lysogenic for the phage P1.  The phage lambda.K is
accepted only by 1 in 10-4 recipient K(P1) cells; these produce modified
lambda.P1 phage particles, which then are able to infect either K or K(P1)
cells with an efficiency of one.  In a beautiful series of experiments Arber
and Dussoix showed that modification must be a process which acts directly on
the DNA of phage lambda, and that where unmodified lambda-DNA enters K(P1)
cells it is rapidly degraded into small molecular weight fragments.  These
findings led Arber to the generalization that modification should affect all
forms of DNA which are synthesized in K(P1) cells, and that restriction would
apply to all DNA that can be transferred from K to K(P1).  He was able to
demonstrate that restriction and modification did occur during the transfer of
the bacterial genome in conjugation, and also during the transfer of DNA by
transforming principle and by transduction (Arber & Dussoix, 1962, Dussoix &
Arber, 1962; Arber, 1962).  In this note we shall first present some data
extending Arber's observations to the transfer of a variety of episomes, and
then show how these results can be utilized to demonstrate that the roles of
restriction and modification imposed by P1 are under independent genetic
control.

<>

<1>Glukhov, I.L., Ishtvan, F., Baev, A.A.
<2>Separation of bacterial methylase includes fractionation of Escherichia coli cells content on columns of polyethylene-imine-agarose, heparin-agarose and phospho-cellulose.
<3>Soviet Patent Office
<4>SU 929705 A
<5>
<6>1982
<7>EcoRI methylase, useful in DNA function investigations in genetic engineering, is recovered
from Escherichia coli by: disintegrating; fractionating with streptomycin sulphate;
concentrating with ammonium sulphate; dialysing; and fractionating on columns containing
polyethylene imine-agarose, heparin-agarose and phospho-cellulose P-11. Inclusion of the
latter fractionating steps enhances the enzyme yield and quickens the overall process.

<>

<1>Glushka, J., Barany, F., Cowburn, D.
<2>Observation of arginyl-deoxyoligonucleotide interactions in TaqI endonuclease by detection of specific 1H NMR signals from 140 kD [Nn1, Nn2, 15N Arg]TaqI/oligomer complexes.
<3>Biochem. Biophys. Res. Commun.
<4>164
<5>88-93
<6>1989
<7>Proton and nitrogen signals of the guanidinium amines in [Nn1, Nn2 15N Arg]TaqI
endonuclease were observed using isotope filtered experiments and proton
detected H{15N} heterocorrelated two dimensional NMR spectroscopy.  These
rapidly exchanging protons could be detected in the free enzyme only at pH 4.5;
at pH 8.5, no signals were measured after extensive signal averaging.  Addition
of deoxyribonucleotide oligomers resulted in the appearance of two groups of
signals at about 5.8 and 7.5 ppm.  Since these signals are independent of the
presence of cognate sequence or Mg2+, it is assumed they represent nonspecific
arginyl-DNA interactions.  This labeling/NMR approach provides a new method for
investigating the role of arginine in protein-DNA interactions.

<>

<1>Godany, A., Bukovska, G., Farkasovska, J., Brnakova, Z., Dmitriev, A., Tkacikova, L., Ayele, T., Mikula, I.
<2>Characterization of a complex restriction-modification system detected in Staphylococcus aureus and Streptococcus agalactiae strains isolated  from infections of domestic animals.
<3>Folia Microbiol. (Praha)
<4>49
<5>307-314
<6>2004
<7>Characterization of classic type II restriction-modification systems (RMS) (restriction
endonucleases and modification methyltransferases)
was carried out in isolates of Staphylococcus aureus and Streptococcus
agalactiae obtained from clinical material. Among the 100 isolates of
S. aureus two different RMS type II were detected. The first was
expressed in isolates 32 and 33 (Sau32 I and Sau33 I); the targeting
sequence was determined as 5'-GGN CC-3' (Sau96 I isoschizomer). The
second was found in isolates no. 90, 93, 96*, and 98 (Sau90 I, Sau93 I,
Sau96* I, Sau98 I) and enzymes recognized sequence 5'-CTY RAG-3' (SmlI
isoschizomer). Analysis of 40 isolates of S. agalactiae revealed only
one RMS; it was detected in two isolates (no. 16 and 23; Sag16 I and
Sag23 I). Restriction endonuclease expressed by these isolates cleaved
DNA in sequence 5'-CTG CA/G-3' (PstI isoschizomer). In RMS-positive S.
aureus and S. agalactiae isolates plasmid DNA capable of replication in
Escherichia coli and Bacillus subtilis was also detected and isolated.

<>

<1>Godany, A., Farkasovska, J., Bukovska, G., Timko, J.
<2>Connection between foreign DNA replication and induced expression of the restriction-modification system in Streptomyces aureofaciens.
<3>Folia Microbiol. (Praha)
<4>46
<5>193-196
<6>2001
<7>Tetracycline-producing strains of Streptomyces aureofaciens expressed the SauLPI
restriction-modification system, which recognized the specific DNA sequence 5-GCCGGC-3'
(isoschizomer NaeI).  The activation of the second R-M system SauLPII (5'-GAGCTC-3',
isoschizomer of XhoI), which was silent during the growth cycle, after a foreign DNA transfer
into this strain was observed.  This phenomenon was tentatively explained as a response of the
cells against the exogenous DNA entering the cells.  The involvement of a SOS-like response in
induction of R-M system genes in S. aureofaciens strains has been considered.

<>

<1>Godany, A., Pristas, P., Oktavcova, B., Farkosovska, J., Ziffova, M., Sevcikova, B.
<2>Characterization of an XhoI isoschizomer in Streptomyces aureofaciens after actinophage infection.
<3>FEMS Microbiol. Lett.
<4>138
<5>123-127
<6>1996
<7>After infection of tetracycline producing strains of S. aureofaciens with
actinophages Mu1/6 and B1 some phage resistant colonies were obtained in each experiment.
These colonies expressed a new restriction-modification (RM) system of type II, which  was
different from the common RM system (SauLPI) of these strains recognizing the  sequence
GCCGGC.  This new RM system was not detected before in parental strains.   The new
endonuclease was purified from a phage resistant strain of S. aureofaciens B96,  using two
step column chromatography to the grade without non specific nucleolytic  activity.  SauLPII
endonuclease recognized and cleaved the palindromic hexanucleotide  sequence 5'-C/TCGAG-3',
thus it was a true isoschizomer of XhoI.

<>

<1>Goddard, M.R., Burt, A.
<2>Recurrent invasion and extinction of a selfish gene.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>13880-13885
<6>1999
<7>Homing endonuclease genes show super-Mendelian inheritance, which allows them to spread in
populations even when they are of no benefit to the host organism. To test the idea that
regular horizontal transmission is necessary for the long-term persistence of these genes, we
surveyed 20 species of yeasts for the omega-homing endonuclease gene and associated group I
intron. The status of omega could be categorized into three states (functional, nonfunctional,
or absent), and status was not clustered on the host phylogeny. Moreover, the phylogeny of
omega differed significantly from that of the host, strong evidence of horizontal
transmission. Further analyses indicate that horizontal transmission is more common than
transposition, and that it occurs preferentially between closely related species. Parsimony
analysis and coalescent theory suggest that there have been 15 horizontal transmission events
in the ancestry of our yeast species, through simulations indicate that this value is probably
an underestimate. Overall, the data support a cyclical model of invasion, degeneration, and
loss, followed by reinvasion, and each of these transitions is estimated to occur about once
every 2 million years. The data are thus consistent with the idea that frequent horizontal
transmission is necessary for the long-term persistence of homing endonuclease genes, and
further, that this requirement limits these genes to organisms with easily accessible germ
lines. The data also show that mitochondrial DNA sequences are transferred intact between
yeast species; if other genes do not show such high levels of horizontal transmission, it
would be due to lack of selection, rather than lack of opportunity.

<>

<1>Godley, L.A., Mondragon, A.
<2>Molecular biology. Preference by exclusion.
<3>Science
<4>331
<5>1017-1018
<6>2011
<7>DNA methylation is a modification that controls gene expression and contributes to mammalian
development, aging, and cancer cell biology.  In mice and humans, the addition of a methyl
group to a cytosine within a cytosine-guanine dinucleotide is catalyzed by DNA
methyltransferase enzymes DNMT3A, DNMT3B, or DNMT1.  The latter is the main "maintenance
methylase" because it adds a methyl group primarily to double-strand DNA that is already
methylated on one strand (hemimethylated).  How DNMT1 prefers hemimethylated over unmethylated
DNA, in contrast to DNMT3A and DNMT3B, has not been clear.  On page 1036 of this issue, Song
et al. present the crystal structures of human and mouse DNMT1 in complex with unmethylated
DNA, providing an explanation for the mechanism of substrate selection by this crucial enzyme.

<>

<1>Godson, G.N., Roberts, R.J.
<2>A catalogue of cleavages of PhiX174, S13, G4, and ST-1 DNA by 26 different restriction endonucleases.
<3>Virology
<4>73
<5>561-567
<6>1976
<7>The number of cleavages produced in PhiX174, S12, G4, and ST-1 double-stranded
RFI DNA by 26 different enzymes is given.  Some enzymes, but not all, produce
similar-sized fragments from PhiX174 and S13 DNA, but all enzymes produce
completely different-sized fragments from G4 and ST-1 compared with PhiX174.
PhiX174 RFI is cleaved once to linear RFIII DNA by PstI, AvaI, and XhoI (BluI).
S13 RFI is cleaved once by PstI, AvaI, and MboI; G4 RF is cleaved once by
EcoRI, KpnI, PstI, and BglII; and ST-1 RFI DNA is cleaved once by KpnI and BglI
and BglII.  The sites of these single cleavages have been mapped.  At least
eight enzymes cleave single-stranded PhiX174 DNA as well as PhiX174
double-stranded DNA.

<>

<1>Goedecke, K., Pignot, M., Goody, R.S., Scheidig, A.J., Weinhold, E.
<2>Structure of the N6-adenine DNA methyltransferase M.TaqI in complex with DNA and a cofactor analog.
<3>Nat. Struct. Biol.
<4>8
<5>121-125
<6>2001
<7>The 2.0 angstrom crystal structure of the N6-adenine DNA methyltransferase M.TaqI in complex
with specific DNA and a non-reactive cofactor analog reveals a previously unrecognized
stabilization of the extrahelical target base.  To catalyze the transfer of the methyl group
from the cofactor S-adenosyl-L-methionine to the 6-amino group of adenine within the
double-stranded DNA sequence 5'-TCGA-3', the target nucleoside is rotated out of the DNA
helix.  Stabilization of the extrahelical conformation is achieved by DNA compression
perpendicular to the DNA helix axis at the target base pair position and relocation of the
partner base thymine in an interstrand Pi-stacked position, where it would sterically overlap
with an inner-helical target adenine.  The extrahelical target adenine is specifically
recognized in the active site, and the 6-amino group of adenine donates two hydrogen bonds to
Asn 105 and Pro 106, which both belong to the conserved catalytic motif IV of N6-adenine DNA
methyltransferases.  These hydrogen bonds appear to increase the partial negative charge of
the N6 atom of adenine and activate it for direct nucleophilic attack on the methyl group of
the cofactor.

<>

<1>Goff, S.P., Rambach, A.
<2>SstI: A restriction endonuclease from Streptomyces sp. stanford.
<3>Gene
<4>3
<5>347-352
<6>1978
<7>A strain of Streptomyces has been isolated which is a convenient source of a
new restriction endonuclease.  The enzyme has been prepared from extracts of
these cells and its cleavage sites localized on phage lambda DNA.  The enzyme,
termed SstI, produces cohesive ends and should be useful for molecular cloning
experiments.

<>

<1>Goffin, J., Eisenhauer, E.
<2>DNA methyltransferase inhibitors-state of the art.
<3>Ann. Oncol.
<4>13
<5>1699-1716
<6>2002
<7>Background: DNA methylation is the addition of a methyl group to the 5 position of cytosine.
It is an epigenetic process with several effects, including chromatin structure modulation,
transcriptional repression and the suppression of transposable elements.  In malignancy,
methylation patterns change, resulting in global hypomethylation with regional
hypermethylation.  This can lead to genetic instability and the repression of tumor suppressor
genes.
Design: A review of the DNA methyltransferase inhibitor literature was conducted.
Results: DNA methylation inhibitors have demonstrated the ability to inhibit hypermethylation,
restore suppressor gene expression and exert antitumor effects in in vitro and in vivo
laboratory models.  Four inhibitors, which are analogs of the nucleoside deoxycytidine, have
been clinically tested: 5-azacytidine, 5-aza-2'-deoxycytidine, 1-beta-D-arabinofuranosyl-5
-azacytosine and dihydro-5-azacytidine.  The first two have demonstrated encouraging
antileukemic activity but little activity in solid tumors, while the latter two are no longer
under study due to lack of efficacy.  A fifth agent, MG98, is an antisense
oligodeoxynucleotide directed against the 3' untranslated region of the DNA
methyltransferase-1 enzyme mRNA, and is now under phase II study.
Conclusions: While some positive clinical results with DNA methyltransferase inhibitors have
been seen, a definitive clinical role for these agents will most likely require combination
therapy, and good phase III studies are needed.

<>

<1>Gogarten, J.P., Hilario, E.
<2>Inteins, introns, and homing endonucleases: recent revelations about the life cycle of parasitic genetic elements.
<3>BMC Evol. Biol.
<4>6
<5>94
<6>2006
<7>Self splicing introns and inteins that rely on a homing endonuclease for propagation are
parasitic genetic elements. Their life-cycle and
evolutionary fate has been described through the homing cycle.
According to this model the homing endonuclease is selected for
function only during the spreading phase of the parasite. This phase
ends when the parasitic element is fixed in the population. Upon
fixation the homing endonuclease is no longer under selection, and its
activity is lost through random processes. Recent analyses of these
parasitic elements with functional homing endonucleases suggest that
this model in its most simple form is not always applicable.
Apparently, functioning homing endonuclease can persist over long
evolutionary times in populations and species that are thought to be
asexual or nearly asexual. Here we review these recent findings and
discuss their implications. Reasons for the long-term persistence of a
functional homing endonuclease include: More recombination (sexual and
as a result of gene transfer) than previously assumed for these
organisms; complex population structures that prevent the element from
being fixed; a balance between active spreading of the homing
endonuclease and a decrease in fitness caused by the parasite in the
host organism; or a function of the homing endonuclease that increases
the fitness of the host organism and results in purifying selection for
the homing endonuclease activity, even after fixation in a local
population. In the future, more detailed studies of the population
dynamics of the activity and regulation of homing endonucleases are
needed to decide between these possibilities, and to determine their
relative contributions to the long term survival of parasitic genes
within a population. Two outstanding publications on the amoeba
Naegleria group I intron (Wikmark et al. BMC Evol Biol 2006, 6: 39) and
the PRP8 inteins in ascomycetes (Butler et al. BMC Evol Biol 2006, 6:
42) provide important stepping stones towards integrated studies on how
these parasitic elements evolve through time together with, or despite,
their hosts.

<>

<1>Gogarten, J.P., Senejani, A.G., Zhaxybayeva, O., Olendzenski, L., Hilario, E.
<2>Inteins: Structure, function, and evolution.
<3>Annu. Rev. Microbiol.
<4>56
<5>263-287
<6>2002
<7>Inteins are genetic elements that disrupt the coding sequence of genes. However, in contrast
to introns, inteins are transcribed and translated
together with their host protein. Inteins appear most frequently in
Archaea, but they are found in organisms belonging to all three domains of
life and in viral and phage proteins. Most inteins consist of two domains:
One is involved in autocatalytic splicing, and the other is an
endonuclease that is important in the spread of inteins. This review
focuses on the evolution and technical application of inteins and only
briefly summarizes recent advances in the study of the catalytic
activities and structures of inteins. In particular, this review considers
inteins as selfish or parasitic genetic elements, a point of view that
explains many otherwise puzzling aspects of inteins.

<>

<1>Goguel, V., Delahodde, A., Jacq, C.
<2>Connections between RNA splicing and DNA intron mobility in yeast mitochondria: RNA maturase and DNA endonuclease switching experiments.
<3>Mol. Cell. Biol.
<4>12
<5>696-705
<6>1992
<7>The intron-encoded proteins bI4 RNA maturase and aI4 DNA endonuclease can
be faithfully expressed in yeast cytoplasm from engineered forms of their
mitochondrial coding sequences. In this work we studied the relationships
between these two activities associated with two homologous intron-encoded
proteins: the bI4 RNA maturase encoded in the fourth intron of the
cytochrome b gene and the aI4 DNA endonuclease (I-SceII) encoded in the
fourth intron of the gene coding for the subunit I of cytochrome oxidase.
Taking advantage of both the high recombinogenic properties of yeast and
the similarities between the two genes, we constructed in vivo a family of
hybrid genes carrying parts of both RNA maturase and DNA endonuclease
coding sequences. The presence of a sequence coding for a mitochondrial
targeting peptide upstream from these hybrid genes allowed us to study the
properties of their translation products within the mitochondria in vivo.
We thus could analyze the ability of the recombinant proteins to
complement RNA maturase deficiencies in different strains. Many
combinations of the two parental intronic sequences were found in the
recombinants. Their structural and functional analysis revealed the
following features. (i) The N-terminal half of the bI4 RNA maturase could
be replaced in total by its equivalent from the aI4 DNA endonuclease
without affecting the RNA maturase activity. In contrast, replacing the
C-terminal half of the bI4 RNA maturase with its equivalent from the aI4
DNA endonuclease led to a very weak RNA maturase activity, indicating that
this region is more differentiated and linked to the maturase activity.
(ii) None of the hybrid proteins carrying an RNA maturase activity kept
the DNA endonuclease activity, suggesting that the latter requires the
integrity of the aI4 protein. These observations are interesting because
the aI4 DNA endonuclease is known to promote the propagation, at the DNA
level, of the aI4 intron, whereas the bI4 RNA maturase, which is required
for the splicing of its coding intron, also controls the splicing process
of the aI4 intron. We propose a scenario for the evolution of these
intronic proteins that relies on a switch from DNA endonuclease to RNA
maturase activity.

<>

<1>Goh, S., Ong, P.F., Song, K.P., Riley, T.V., Chang, B.J.
<2>The complete genome sequence of Clostridium difficile phage {phi}C2 and comparisons to {phi}CD119 and inducible prophages of CD630.
<3>Microbiology
<4>153
<5>676-685
<6>2007
<7>The complete genomic sequence of a previously characterized temperate
phage of Clostridium difficile, C2, is reported. The genome is 56 538 bp
and organized into 84 putative ORFs in six functional modules. The head
and tail structural proteins showed similarities to that of C. difficile
phage CD119 and Streptococcus pneumoniae phage EJ-1, respectively.
Homologues of structural and replication proteins were found in prophages
1 and 2 of the sequenced C. difficile CD630 genome. A putative holin
appears unique to the C. difficile phages and was functional when
expressed in Escherichia coli. Nucleotide sequence comparisons of C2 to
CD119 and the CD630 prophage sequences showed relatedness between C2 and
the prophages, but less so to CD119. C2 integrated into a gene encoding a
putative transcriptional regulator of the gntR family. C2, CD119 and CD630
prophage 1 genomes had a Cdu1-attP-integrase arrangement, suggesting that
the pathogenicity locus (PaLoc) of C. difficile, flanked by cdu1, has
phage origins. The attP sequences of C2, CD119 and CD630 prophages were
dissimilar. C2-related sequences were found in 84 % of 37 clinical C.
difficile isolates and typed reference strains.

<>

<1>Gohda, K., Matsuo, N., Oda, Y., Ikehara, M., Uesugi, S.
<2>Effects of 2'-substituents of the first deoxyguanosine residue in the recognition sequence on EcoRI restriction endonuclease activity.
<3>J. Biochem. (Tokyo)
<4>121
<5>219-224
<6>1997
<7>The effects of 2'-substituents of the first deoxyguanosine on EcoRI activity were examined
using synthetic octadeoxynucleotides d(GG*AATTCC) containing 2'-substituted derivatives (G*),
i.e., 2'-fluoro-2'-deoxyguanosine (dGfl), 2'-chloro-2'-deoxyguanosine (dGcl), and
guanosine (rG).  The overall structures of the octamers were very similar, as shown by CD and
UV measurements, although their EcoRI reactivities were very different: 100% in 60 min for
d(GGAATTCC) and d(GGflAATTCC), 5% in 24h for d[G(rG)AATTCC], and no cleavage at all in 24h for
d(GGclAATTCC).  However, the kinetics showed the octamers exhibit similar binding-affinity to
the enzyme (10^6-10^7 M).  31P-NMR analysis suggested the modified octamers change the
phosphate backbone conformation in a duplex, since an unusual downfield-shifted signal in the
spectra was commonly observed for the modified octamers at low temperature (i.e., a single
strand state), which was shifted upfield at high temperature (i.e., a single strand state).
The order of the differences was dGcl>rG>dGfl-containing octamers, coinciding with that of the
vdW volume of 2'-substituents (Cl>OH>F) and the cleavage reactivities.  These findings
suggest the steric hindrance by the 2'-substituents causes a conformational change of the
phosphate backbone close to the scissile bond, and then interferes with the EcoRI reaction.

<>

<1>Goker, M. et al.
<2>Complete genome sequence of the termophilic sulfur-reducer Desulfurobacterium thermolithotrophum type strain (BSA) from a deep-sea hydrothermal vent.
<3>Standards in Genomic Sciences
<4>5
<5>407-415
<6>2011
<7>Desulfurobacterium thermolithotrophum L'Haridon et al. 1998 is the type species of the ge-nus
Desulfurobacterium which belongs to the family Desulfurobacteriaceae. The species is of
interest because it represents the first thermophilic bacterium that can act as a primary
pro-ducer in the temperature range of 45-75°C (optimum 70°C) and is incapable of growing
un-der microaerophilic conditions. Strain BSAT preferentially synthesizes high-melting-point
fatty acids (C18 and C20) which is hypothesized to be a strategy to ensure the functionality
of the membrane at high growth temperatures. This is the second completed genome sequence of a
member of the family Desulfurobacteriaceae and the first sequence from the genus
Desulfurobacterium. The 1,541,968 bp long genome harbors 1,543 protein-coding and 51 RNA genes
and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Goker, M. et al.
<2>Genome sequence of the Thermotoga thermarum type strain (LA3(T)) from an African  solfataric spring.
<3>Standards in Genomic Sciences
<4>9
<5>1105-1117
<6>2014
<7>Thermotoga thermarum Windberger et al. 1989 is a member to the genomically well characterized
genus Thermotoga in the phylum 'Thermotogae'. T. thermarum is of
interest for its origin from a continental solfataric spring vs. predominantly
marine oil reservoirs of other members of the genus. The genome of strain LA3T
also provides fresh data for the phylogenomic positioning of the
(hyper-)thermophilic bacteria. T. thermarum strain LA3(T) is the fourth sequenced
genome of a type strain from the genus Thermotoga, and the sixth in the family
Thermotogaceae to be formally described in a publication. Phylogenetic analyses
do not reveal significant discrepancies between the current classification of the
group, 16S rRNA gene data and whole-genome sequences. Nevertheless, T. thermarum
significantly differs from other Thermotoga species regarding its iron-sulfur
cluster synthesis, as it contains only a minimal set of the necessary proteins.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. The 2,039,943 bp long chromosome with its 2,015
protein-coding and 51 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Goker, M. et al.
<2>Genome sequence of the mud-dwelling archaeon Methanoplanus limicola type strain (DSM 2279(T)), reclassification of Methanoplanus petrolearius as Methanolacinia  petrolearia and emended descriptions of the genera Methanoplanus and  Methanolacinia.
<3>Standards in Genomic Sciences
<4>9
<5>1076-1088
<6>2014
<7>Methanoplanus limicola Wildgruber et al. 1984 is a mesophilic methanogen that was isolated
from a swamp composed of drilling waste near Naples, Italy, shortly
after the Archaea were recognized as a separate domain of life. Methanoplanus is
the type genus in the family Methanoplanaceae, a taxon that felt into disuse
since modern 16S rRNA gene sequences-based taxonomy was established.
Methanoplanus is now placed within the Methanomicrobiaceae, a family that is so
far poorly characterized at the genome level. The only other type strain of the
genus with a sequenced genome, Methanoplanus petrolearius SEBR 4847(T), turned
out to be misclassified and required reclassification to Methanolacinia. Both,
Methanoplanus and Methanolacinia, needed taxonomic emendations due to a
significant deviation of the G+C content of their genomes from previously
published (pre-genome-sequence era) values. Until now genome sequences were
published for only four of the 33 species with validly published names in the
Methanomicrobiaceae. Here we describe the features of M. limicola, together with
the improved-high-quality draft genome sequence and annotation of the type
strain, M3(T). The 3,200,946 bp long chromosome (permanent draft sequence) with
its 3,064 protein-coding and 65 RNA genes is a part of the G enomic E ncyclopedia
of B acteria and Archaea project.

<>

<1>Goker, M. et al.
<2>Complete genome sequence of the acetate-degrading sulfate reducer Desulfobacca acetoxidans type strain (ASRB2).
<3>Standards in Genomic Sciences
<4>4
<5>393-401
<6>2011
<7>Desulfobacca acetoxidans Elferink et al. 1999 is the type species of the genus Desulfobacca,
which belongs to the family Syntrophaceae in the class
Deltaproteobacteria. The species was first observed in a study on the competition
of sulfate-reducers and acetoclastic methanogens for acetate in sludge. D.
acetoxidans is considered to be the most abundant acetate-degrading sulfate
reducer in sludge. It is of interest due to its isolated phylogenetic location in
the 16S rRNA-based tree of life. This is the second completed genome sequence of
a member of the family Syntrophaceae to be published and only the third genome
sequence from a member of the order Syntrophobacterales. The 3,282,536 bp long
genome with its 2,969 protein-coding and 54 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Goker, M. et al.
<2>Genome sequence of the moderately thermophilic, amino-acid-degrading and sulfur-reducing bacterium Thermovirga lienii type strain (Cas60314(T)).
<3>Standards in Genomic Sciences
<4>6
<5>230-239
<6>2012
<7>Thermovirga lienii Dahle and Birkeland 2006 is a member of the genus Thermovirga  in the
genomically moderately well characterized phylum 'Synergistetes'. Members
of this relatively recently proposed phylum 'Synergistetes' are of interest
because of their isolated phylogenetic position and their diverse habitats, e.g.
from humans to oil wells. The genome of T. lienii Cas60314(T) is the fifth genome
sequence (third completed) from this phylum to be published. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. The 1,999,646 bp long genome (including one plasmid) with its 1,914
protein-coding and 59 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Goker, M. et al.
<2>Complete genome sequence of Isosphaera pallida type strain (IS1B).
<3>Standards in Genomic Sciences
<4>4
<5>63-71
<6>2011
<7>Isosphaera pallida (ex Woronichin 1927) Giovannoni et al. 1995 is the type species of the
genus Isosphaera. The species is of interest because it was the
first heterotrophic bacterium known to be phototactic, and it occupies an
isolated phylogenetic position within the Planctomycetaceae. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. This is the first complete genome sequence of a member of the genus
Isosphaera and the third of a member of the family Planctomycetaceae. The
5,472,964 bp long chromosome and the 56,340 bp long plasmid with a total of 3,763
protein-coding and 60 RNA genes are part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Goker, M. et al.
<2>Complete genome sequence of Odoribacter splanchnicus type strain (1651/6).
<3>Standards in Genomic Sciences
<4>4
<5>200-209
<6>2011
<7>Odoribacter splanchnicus (Werner et al. 1975) Hardham et al. 2008 is the type species of the
genus Odoribacter, which belongs to the family Porphyromonadaceae
in the order 'Bacteroidales'. The species is of interest because members of the
Odoribacter form an isolated cluster within the Porphyromonadaceae. This is the
first completed genome sequence of a member of the genus Odoribacter and the
fourth sequence from the family Porphyromonadaceae. The 4,392,288 bp long genome
with its 3,672 protein-coding and 74 RNA genes and is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Goker, M. et al.
<2>Complete genome sequence of Ignisphaera aggregans type strain (AQ1.S1).
<3>Standards in Genomic Sciences
<4>3
<5>66-75
<6>2010
<7>Ignisphaera aggregans Niederberger et al. 2006 is the type and sole species of genus
Ignisphaera. This archaeal species is characterized by a coccoid-shape and
is strictly anaerobic, moderately acidophilic, heterotrophic hyperthermophilic
and fermentative. The type strain AQ1.S1(T) was isolated from a near neutral,
boiling spring in Kuirau Park, Rotorua, New Zealand. This is the first completed
genome sequence of the genus Ignisphaera and the fifth genome (fourth type
strain) sequence in the family Desulfurococcaceae. The 1,875,953 bp long genome
with its 2,009 protein-coding and 52 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Goker, M. et al.
<2>Complete genome sequence of Olsenella uli type strain (VPI D76D-27C).
<3>Standards in Genomic Sciences
<4>3
<5>76-84
<6>2010
<7>Olsenella uli (Olsen et al. 1991) Dewhirst et al. 2001 is the type species of the genus
Olsenella, which belongs to the actinobacterial family Coriobacteriaceae.
The species is of interest because it is frequently isolated from dental plaque
in periodontitis patients and can cause primary endodontic infection. The species
is a Gram-positive, non-motile and non-sporulating bacterium. The strain
described in this study was isolated from human gingival crevices. This is the
first completed sequence of the genus Olsenella and the fifth sequence from a
member of the family Coriobacteriaceae. The 2,051,896 bp long genome with its
1,795 protein-coding and 55 RNA genes is a part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Gold, M., Hausmann, R., Maitra, U., Hurwitz, J.
<2>The enzymatic methylation of RNA and DNA, VIII.  Effects of bacteriophage  infection on the activity of the methylating enzymes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>52
<5>292-297
<6>1964
<7>We, as well as others, have previously reported on the presence in Escherichia coli of several
enzymes which catalyze the transfer of methyl groups from S-adenosylmethionine to sRNA,
ribosomal RNA, and DNA.  Although the biological function of the methylated bases which these
enzymes produce is still obscure, the species and strain specificity of the methylation
reactions suggest that they provide a basis for a recognition mechanism.  The virulent
bacteriophage-host cell system is an example of a phenomenon involving recognition by the host
of a foreign nucleic acid; in some instances, phage DNA is rapidly synthesized while the host
DNA is rapidly degraded.  If methylated bases are involved in controlling such a recognition
mechanism, then a study of the methylated base content of DNA's of various bacteriophages
grown in different hosts might provide a clue as to the biological function of the methylating
enzymes.  In order to establish a suitable system for further investigation, we have studied
the effects of phage infection on the activities of the various methylating enzymes in the
host cell.  This communication summarizes such studies.  It has been found that while the RNA
methylases are apparently unchanged, DNA methylation activity increases markedly after
infection with T2.  In contrast, T3 infection induces an enzyme which cleaves
S-adenosylmethionine to thiomethyladenosine and homoserine.

<>

<1>Gold, M., Hurwitz, J.
<2>The enzymatic methylation of ribonucleic acid and deoxyribonucleic acid. V. Purification and properties of the deoxyribonucleic acid-methylating activity of Escherichia coli.
<3>J. Biol. Chem.
<4>239
<5>3858-3865
<6>1964
<7>The classical investigations of Wyatt, and Dunn and Smith, established the existence of
5-methylcytosine and 6-methylaminopurine as constituents of  the deoxyribonucleic acid of
higher plants and animals, and of bacteria and bacterial viruses, respectively.  Until
recently, the biochemical pathways for the  incorporation of these "trace bases" into
deoxyribonucleic acid was unknown although it had been shown that the deoxyribonucleoside
triphosphate of 5-methylcytosine could quantitatively replace deoxycytidine triphosphate as a
substrate for the enzyme deoxyribonucleic acid polymerase.  The discovery that the
glucosylation of A of the T-even bacteriophages and methylation of soluble ribonucleic acid
occurred at the polynucleotide level suggested that the incorporation of methyl groups might
also take place after polymerization in the biosynthesis of DNA.  In previous communications
from this laboratory, the presence of an enzyme activity of Escherichia coli which catalyzes
the transfer of methyl groups to cytosine and adenine moieties of a variety of DNAs has been
reported.  This reaction can be presented by the following equation.  DNA +
S-adenosylmethionine  + DNA methylase  gives methyl-DNA (containing 5-methylcytosine and
6-methylaminopurine) + S-adenosylhomocysteine.  In this report the purification and properties
of this enzyme activity are described.

<>

<1>Gold, M., Hurwitz, J., Anders, M.
<2>THE ENZYMATIC METHYLATION OF RNA AND DNA, II. ON THE SPECIES SPECIFICITY OF THE METHYLATION ENZYMES.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>50
<5>164-169
<6>1963
<7>There is evidence that methylated bases in DNA and sRNA are not randomly distributed in
polynucleotide chains.  In wheat germ DNA, the two 6-aminopyrimidines, cytosine and
5-methylcytosine, do not appear to substitute randomly for each other, as determined by
chemical analysis of oligonucleotides.  This specificity appears to be related to the direct
methylation of deoxycytidylate of DNA at the polynucleotide level, as has also been found to
apply to the origin of the base, 6-methylaminopurine.  These observations are in keeping with
the incorporation studies of Bessman et al. with the DNA polymerase system.  This enzyme
readily catalyzes the incorporation of dCMP and 5-methyl dCMP into DNA without distinguishing
between these deoxynucleotides, and therefore does not appear responsible for localization of
methylated bases.  In RNA, the methylated bases are uniquely localized to soluble RNA as
indicated by a large body of information.  Here, RNA polymerase, the enzyme which appears to
synthesize all RNA species from a DNA template of normal cells, lacks specificity in
differentiating between methylated bases and normal bases.  Thus, for example, ribothymidylate
is readily incorporated into RNA in place of uridylate with the same nearest neighbor
frequency.  However, the distribution of methylated bases of sRNA is not random, and analyses
of purified sRNA molecules, specific for particular amino acids, indicate that they contain
varied amounts as well as different methylated bases.  As in the case of DNA, this specific
distribution of methylated bases has been explained by the observation that methylation occurs
at the polynucleotide level rather than the mononucleotide stage.  While with DNA it appears
that methylation is catalyzed by a single enzyme, in the case of sRNA many enzymes are
involved.  To date, 5 different enzymes catalyzing specific methylation reactions with sRNA
have been isolated.

<>

<1>Gold, M., Hurwitz, J., Anders, M.
<2>The enzymatic methylation of RNA and DNA.
<3>Biochem. Biophys. Res. Commun.
<4>11
<5>107-114
<6>1963
<7>The methionine origin of the methyl group of the "trace bases" of Escherichia coli and ascites
cells has been demonstrated by Borek et al. and Biswas et al.  Fleissner and Borek have
recently reported that extracts of E. coli catalyse the transfer of the methyl group from
C14-methyl-labeled methionine to E. coli soluble RNA provided this S-RNA was isolated from a
methionine auxotroph which continues to synthesize RNA when deprived of its essential amino
acid.  Although the DNA dependent RNA polymerase system incorporated ribothymidylate from
ribothymidine triphosphate into RNA specifically in place of uridylate, no demonstrable
phosphorylation of ribothymidylate was detected in extracts of E. coli.  These observations,
and especially the report of Fleissner and Borek, suggest that the trace nucleotide
ribothymidylate, is synthesized at the polynucleotide level.

<>

<1>Gold, S.E., Blacutt, A.A., Meinersmann, R.J., Bacon, C.W.
<2>Whole-Genome Shotgun Sequence of Bacillus mojavensis Strain RRC101, an Endophytic Bacterium Antagonistic to the Mycotoxigenic Endophytic Fungus Fusarium  verticillioides.
<3>Genome Announcements
<4>2
<5>e01090-14
<6>2014
<7>Here, we report the whole-genome shotgun sequence of Bacillus mojavensis strain RRC101,
isolated from a maize kernel. This strain is antagonistic to the
mycotoxigenic plant pathogen Fusarium verticillioides and grows within maize
tissue, suggesting potential as an endophytic biocontrol agent.

<>

<1>Golding, G.R., Bryden, L., Levett, P.N., McDonald, R.R., Wong, A., Graham, M.R., Tyler, S., Van Domselaar, G., Mabon, P., Kent, H., Butaye, P., Smith, T.C., Kadlec, K., Schwarz, S., Weese, S.J., Mulvey, M.R.
<2>Whole-Genome Sequence of Livestock-Associated ST398 Methicillin-Resistant Staphylococcus aureus Isolated from Humans in Canada.
<3>J. Bacteriol.
<4>194
<5>6627-6628
<6>2012
<7>Despite reports of high colonization rates of ST398 livestock-associated methicillin-resistant
Staphylococcus aureus (LA-MRSA) among pigs and pig farmers,
the incidence of LA-MRSA infection in the general population in Canada appears to
be rare in comparison to that in some European countries. In this study, the
complete genome sequence of a Canadian representative LA-MRSA isolate (08BA02176)
from a human postoperative surgical site infection was acquired and compared to
the sequenced genome of an LA-MRSA isolate (S0385) from Europe to identify
genetic traits that may explain differences in the success of these particular
strains in some locales.

<>

<1>Golding, M.C., Westhusin, M.E.
<2>Analysis of DNA (cytosine 5) methyltransferase mRNA sequence and expression in bovine preimplantation embryos, fetal and adult tissues.
<3>Gene Expr. Patterns
<4>3
<5>551-558
<6>2003
<7>Mammalian preimplantation development is a critical stage for establishment of the genomic
methylation pattern and proper function of
the enzymes responsible for this appear essential for normal development.
To date, the vast majority of work concerning the developmental expression
of the DNA cytosine 5-methyltansferases (Dnmts) has been conducted in
mice. Here we report the sequence and expression of the Dnmt family during
bovine preimplantation and fetal development. Bovine Dnmt mRNAs display
strong sequence homology to those of human and mouse and similar to other
species, exist as multiple isoforms. Two of these splice variants, which
have been termed Dnmt2gamma and Dnmt3a4, represent previously unreported
sequence combinations. Work presented here demonstrates early bovine
embryos express mRNA coding for the somatic form of Dnmt1 and that this
transcript fractionates with the ribosome. Unlike the murine model, mRNA
encoding the de novo methyltransferases, Dnmt3a and 3b are present during
preimplantation development and can also be found in the ribosomal
subcellular fraction. Further, results of Real Time PCR analysis indicate
significant differences in Dnmt mRNA expression levels exist among
different tissue types as well as between fetal and adult stages.
Recently, it has been postulated that the cause of abnormal methylation
observed in cloned embryos may be due in part to misexpression of the
Dnmt1o isoform during preimplantation development. Work presented here
raises new and significant hypotheses that must be considered both
regarding the cadre of DNA methyltranferases that direct epigenetic
programming during normal development and regarding the implication of
abnormal DNMT expression in cloned embryos.

<>

<1>Goldman, B.S., Hinkle, G.J., Slater, S.C., Wiegand, R.C.
<2>Myxococcus xanthus genome sequences and uses thereof.
<3>US Patent Office
<4>US 6833447 A
<5>
<6>2004
<7>The presesnt invention relates to nulceic acid sequences from the bacterium, Myxococcus
xanthus and, in particular, to genomic DNA sequences.  The invention encompasses nucleic acid
molecules present in non-coding regions as well as nucleic acid molecules that encode proteins
and fragments of proteins.  In addition, proteins and fragments of proteins so encoded and
antibodies capable of binding the proteins are encompassed by the present invention.  The
invention also encompasses oligonucleotides including primers, e.g. useful for amplifying
nucleic acid molecules, and collections of nucleic acid molecules and oligonucleotides, e.g.
in microarrays.  The invention also provides constructs and transgenic cells and organisms
comprising nucleic acid molecules of the invention.  The invention also relates to methods of
using the disclosed nucleic acid molecules, oligonucleotides, proteins, fragments of proteins,
and antibodies, for example, for gene identification and analysis, and preparation of
constructs and transgenic cells and organisms.

<>

<1>Goldman, B.S., Krasomil-Osterfeld, K., Malvar, T.M., Pitkin, J.W., Slater, S.C., Wu, W., Zeng, J.
<2>Nucleotide and amino acid sequences from Xenorhabdus and uses thereof.
<3>US Patent Office
<4>US 7319142 A
<5>
<6>2008
<7>The invention provides isolated nucleotide sequences from Xenorhabdus nematophila species
Xs86068, and, in particular, nucleotide sequences that encode insect inhibitory proteins, the
insecticidal proteins, and compositions that comprise one or more of the insecticidal proteins
for use in controlling insect infestation.

<>

<1>Goldstein, A.S., Hughes, A.J. Jr.
<2>Nucleic acid-free thermostable enzymes and methods of production thereof.
<3>US Patent Office
<4>US 5861295
<5>
<6>1999
<7>The present invention provides thermostable enzymes, such as DNA polymerases and restriction
endonucleases, that are substantially free from contamination with nucleic acids.  The
invention also provides methods for the production of these enzymes, and kits comprising these
enzymes which may be used in amplifying or sequencing nucleic acid molecules, including
through use of the polymerase chain reaction.

<>

<1>Goldstein, S.W., Mylari, B.L., Perez, J.R., Glazer, E.A.
<2>Preparation of bisamidinophenylindoles as antiproliferative DNA methyltransferase inhibiting agents.
<3>US Patent Office
<4>US 6699862 B
<5>
<6>2004
<7>Novel indolyl-2-phenyl bisamidines are described which are DNA methyltransferase inhibiting
agents, and pharmaceutical compositions containing them are used as antiproliferative agents
for treating a disease, especially a neoplastic disease, characterized by abnormally rapid
proliferation of tissue involved in said disease; said indolyl-2-phenyl bisamidines comprising
a (figure) and a pharmaceutically acceptable salt thereof, wherein: R1 is hydrogen or
(c1-C3)alkyl; R3, R4, R5, R8, R9, and R10 are hydrogen or (C1-C3)alkyl, or R3 and R4 may be
taken together, or R8 and R9 may be taken together with the nitrogen atoms to which they are
attached, to form an imidazolinyl group; R14 is -H; -NHC(=O) (CH2)m R20; -(CH2)mR20;
-CH(CH3)R20; -(CH2)m (C6H3)-R17; -(CH2)m(C6H3)-R20; -(CH2)m (heterocyclyl)-R17;
-(CH2)m(heterocyclyl)-R20; -CH2CH=CHR20; -(CH2)m C(=O)NH-CHR20R21; or -CH2)m C(=O)NH-CH2-C(=O)
NHCHR20R21; where R17 is hydrogen; halogen; (C1-C3)alkyl; -CF3; -CN; -NO2; -N(R1)2; -OH; or
(C1-C3) alkyl(C1-C3) alkyl(C1-C3)alkoxy; R21 is -C(=O)OR1; CH(OH)CH2OH; -C(=O)NH2; or -C(=O)H;
R21 is hydrogen; (C1-C6)alkyl; -(CH2)nR22; -CH(CH3)CH2C(=O)OR1; and -CH2-(C6H5); and R22 is
-H; -NH2; -OR1; -SR1; -CN; -OCH2-(C6H5); -O(CH2)m -OR1; -C(=O)OR1; thienyl; tetrahydropyranyl;
-CH(OH)CH2OH; -C(=O)C(CH3)=CH2; -NHC(=O)OCH2- (C6H5); and -S(=O)2R1; where m is 1, 2, or 3;
and n is 1 through 5, inclusive.

<>

<1>Goldstone, R.J., Amos, M., Talbot, R., Schuberth, H.J., Sandra, O., Sheldon, I.M., Smith, D.G.
<2>Draft Genome Sequence of Trueperella pyogenes, Isolated from the Infected Uterus  of a Postpartum Cow with Metritis.
<3>Genome Announcements
<4>2
<5>e00194-14
<6>2014
<7>Trueperella pyogenes is a common commensal bacterium and an opportunistic pathogen associated
with chronic purulent disease, particularly in ruminants. We report here the genome sequence
of a T. pyogenes isolate from a severe case of bovine metritis. This is the first full record
of a T. pyogenes genome.

<>

<1>Goldstone, R.J., Talbot, R., Schuberth, H.J., Sandra, O., Sheldon, I.M., Smith, D.G.
<2>Draft Genome Sequence of Escherichia coli MS499, Isolated from the Infected Uterus of a Postpartum Cow with Metritis.
<3>Genome Announcements
<4>2
<5>e00217-14
<6>2014
<7>Specific Escherichia coli strains associated with bovine postpartum uterine infection have
recently been described. Many recognized virulence factors are
absent in these strains; therefore, to define a prototypic strain, we report here
the genome sequence of E. coli isolate MS499 from a cow with the postpartum
disease metritis.

<>

<1>Golemboski, D., Eardly, B.D.
<2>Draft Genome Sequences of Respiratory and Urinary Tract Isolates of Acinetobacter baumannii from the Same Patient.
<3>Genome Announcements
<4>2
<5>e00692-14
<6>2014
<7>Acinetobacter baumannii is a frequent hospital-acquired human pathogen. This report describes
the draft genome sequences of two distinct A. baumannii clinical
isolates from the same patient. A comparison of the genomes revealed differences
in antibiotic resistance and will enable the determination of genomic differences
responsible for virulence at each body site.

<>

<1>Golikova, L.N., Gutorov, V.V., Evdokimov, A.A., Shchelkunov, S.N., Gonchar, D.A., Okhapkina, S.S., Degtyarev, S.K., Netesova, N.A.
<2>M.BstF5I-4, the fourth DNA-methyltransferase of the BstF5I restriction-modification system from Bacillus stearothermophilus F5.
<3>Bioorg. Khim.
<4>28
<5>84-86
<6>2002
<7>The fourth DNA-methyltransferase of the BstF5I restriction-modification (RM) system from
Bacillus stearothermophilus F5 (M.BstF5I-4) was discovered, which modifies the adenine residue
within the upper strand of the recognition site 5'-GGATG-3'/5'-CATCC-3'. Thus, unlike
other known RM systems, the BstF5I RM system comprises four genes encoding
DNA-methyltransferases, three of which possess the same substrate specificity and methylate
adenine within the 5'-GGATG sequence.

<>

<1>Golikova, L.N., Netosova, N.A., Gutorov, V.V., Belavin, P.A., Abdurashitov, M.A., Gonchar, D.A., Degtyarev, S.K.
<2>Multiplicity of site-specific DNA-methyltransferases of the BstF5I restriction modification system from Bacillus stearothermophilus F5.
<3>Mol. Biol. (Mosk)
<4>34
<5>443-447
<6>2000
<7>A fragment located downstream of the genes for DNA methyltransferases of Bacillus
stearothermophilus F5 (M.BstF5I-I and M.BstF5I-2) was
sequenced. The fragment contains a gene for another methylase,
M.BstF5I-3, structurally and functionally similar to the N-terminal
domain of M.FokI. Thus, in contrast to other restriction-modification
systems, the BstF5I system includes three methylases, two being
homologous to the individual M.FokI domains.

<>

<1>Goll, M.G., Bestor, T.H.
<2>Eukaryotic cytosine methyltransferases.
<3>Annu. Rev. Biochem.
<4>74
<5>481-514
<6>2005
<7>Large-genome eukaryotes use heritable cytosine methylation to silence promoters, especially
those associated with transposons and imprinted
genes. Cytosine methylation does not reinforce or replace ancestral
gene regulation pathways but instead endows methylated genomes with the
ability to repress specific promoters in a manner that is buffered
against changes in the internal and external environment. Recent
studies have shown that the targeting of de novo methylation depends on
multiple inputs; these include the interaction of repeated sequences,
local states of histone lysine methylation, small RNAs and components
of the RNAi pathway, and divergent and catalytically inert cytosine
methyltransferase homologues that have acquired regulatory roles. There
are multiple families of DNA (cytosine-5) methyltransferases in
eukaryotes, and each family appears to be controlled by different
regulatory inputs. Sequence-specific DNA-binding proteins, which
regulate most aspects of gene expression, do not appear to be involved
in the establishment or maintenance of genomic methylation patterns.

<>

<1>Gololobova, N.S., Okhapkina, S.S., Abdurashitov, M.A., Degtyarev, S.K.
<2>Determination and analysis of the primary structure of NM.BstSEI operon from Bacillus stearothermophilus SE-589 which produces N.BstSEI site-specific nickase.
<3>Mol. Biol. (Mosk)
<4>39
<5>960-964
<6>2005
<7>Nucleotide sequence of Bacillus stearothermophilus SE-589 DNA fragment which includes an
operon for site-specific NM-system with a gene for BstSEI nickase has been determined.
Analysis of the regions adjacent to nickase gene has revealed two genes encoding DNA
methyltransferases, which belong to different classes. Three genes which form system operon
are separated with short open reading frames (ORFs). Analysis of these ORFs has shown that
they encode polypeptides which are homologous to different parts of BstSEI nickase, NatB
protein and arginase. A difference in GC-content of the beginning and ending regions of the
cloned DNA fragment as well as presence of short ORFs similar to genes for known proteins may
indicate that NM.BstSEI system operon has evolved by horizonthal DNA transfer.

<>

<1>Golovenko, D., Manakova, E., Tamulaitiene, G., Grazulis, S., Siksnys, V.
<2>Structural mechanisms for the 5'-CCWGG sequence recognition by the N- and C-terminal domains of EcoRII.
<3>Nucleic Acids Res.
<4>37
<5>6613-6624
<6>2009
<7>EcoRII restriction endonuclease is specific for the 5'-CCWGG sequence (W stands for A or T);
however, it shows no activity on a single recognition site. To activate cleavage it requires
binding of an additional target site as an allosteric effector. EcoRII dimer consists of three
structural units: a central catalytic core, made from two copies of the C-terminal domain
(EcoRII-C), and two N-terminal effector DNA binding domains (EcoRII-N). Here, we report
DNA-bound EcoRII-N and EcoRII-C structures, which show that EcoRII combines two radically
different structural mechanisms to interact with the effector and substrate DNA. The catalytic
EcoRII-C dimer flips out the central T:A base pair and makes symmetric interactions with the
CC:GG half-sites. The EcoRII-N effector domain monomer binds to the target site asymmetrically
in a single defined orientation which is determined by specific hydrogen bonding and van der
Waals interactions with the central T:A pair in the major groove. The EcoRII-N mode of the
target site recognition is shared by the large class of higher plant transcription factors of
the B3 superfamily.

<>

<1>Golovenko, D., Manakova, E., Zakrys, L., Zaremba, M., Sasnauskas, G., Grazulis, S., Siksnys, V.
<2>Structural insight into the specificity of the B3 DNA-binding domains provided by the co-crystal structure of the C-terminal fragment of BfiI restriction enzyme.
<3>Nucleic Acids Res.
<4>42
<5>4113-4122
<6>2014
<7>The B3 DNA-binding domains (DBDs) of plant transcription factors (TF) and DBDs of EcoRII and
BfiI restriction endonucleases (EcoRII-N and BfiI-C) share a common
structural fold, classified as the DNA-binding pseudobarrel. The B3 DBDs in the
plant TFs recognize a diverse set of target sequences. The only available
co-crystal structure of the B3-like DBD is that of EcoRII-N (recognition sequence
5'-CCTGG-3'). In order to understand the structural and molecular mechanisms of
specificity of B3 DBDs, we have solved the crystal structure of BfiI-C
(recognition sequence 5'-ACTGGG-3') complexed with 12-bp cognate oligoduplex.
Structural comparison of BfiI-C-DNA and EcoRII-N-DNA complexes reveals a
conserved DNA-binding mode and a conserved pattern of interactions with the
phosphodiester backbone. The determinants of the target specificity are located
in the loops that emanate from the conserved structural core. The BfiI-C-DNA
structure presented here expands a range of templates for modeling of the
DNA-bound complexes of the B3 family of plant TFs.

<>

<1>Goltsman, D.S., Denef, V.J., Singer, S.W., VerBerkmoes, N.C., Lefsrud, M., Mueller, R.S., Dick, G.J., Sun, C.L., Wheeler, K.E., Zemla, A., Baker, B.J., Hauser, L., Land, M., Shah, M.B., Thelen, M.P., Hettich, R.L., Banfield, J.F.
<2>Community genomic and proteomic analyses of chemoautotrophic iron-oxidizing 'Leptospirillum rubarum' (Group II) and 'Leptospirillum ferrodiazotrophum' (Group III) bacteria in acid mine drainage biofilms.
<3>Appl. Environ. Microbiol.
<4>75
<5>4599-4615
<6>2009
<7>We analyzed near-complete population (composite) genomic sequences for
coexisting acidophilic iron-oxidizing Leptospirillum group II and III
bacteria (phylum Nitrospirae) and an extrachromosomal plasmid from a
Richmond Mine, Iron Mountain, CA, acid mine drainage biofilm. Community
proteomic analysis of the genomically characterized sample and two other
biofilms identified 64.6% and 44.9% of the predicted proteins of
Leptospirillum groups II and III, respectively, and 20% of the predicted
plasmid proteins. The bacteria share 92% 16S rRNA gene sequence identity
and >60% of their genes, including integrated plasmid-like regions. The
extrachromosomal plasmid carries conjugation genes with detectable
sequence similarity to genes in the integrated conjugative plasmid, but
only those on the extrachromosomal element were identified by proteomics.
Both bacterial groups have genes for community-essential functions,
including carbon fixation and biosynthesis of vitamins, fatty acids, and
biopolymers (including cellulose); proteomic analyses reveal these
activities. Both Leptospirillum types have multiple pathways for osmotic
protection. Although both are motile, signal transduction and
methyl-accepting chemotaxis proteins are more abundant in Leptospirillum
group III, consistent with its distribution in gradients within biofilms.
Interestingly, Leptospirillum group II uses a methyl-dependent and
Leptospirillum group III a methyl-independent response pathway. Although
only Leptospirillum group III can fix nitrogen, these proteins were not
identified by proteomics. The abundances of core proteins are similar in
all communities, but the abundance levels of unique and shared proteins of
unknown function vary. Some proteins unique to one organism were highly
expressed and may be key to the functional and ecological differentiation
of Leptospirillum groups II and III.

<>

<1>Golubov, A., Neubauer, H., Nolting, C., Heesemann, J., Rakin, A.
<2>Structural organization of the pFra virulence-associated plasmid of rhamnose-positive Yersinia pestis.
<3>Infect. Immun.
<4>72
<5>5613-5621
<6>2004
<7>The 137,036-bp plasmid pG8786 from rhamnose-positive Yersinia pestis G8786
isolated from the high mountainous Caucasian plague focus in Georgia is an
enlarged form of the pFra virulence-associated plasmid containing genes
for synthesis of the antigen fraction 1 and phospholipase D. In addition
to the completely conserved genes of the pFra backbone, pG8786 contains
two large regions consisting of 4,642 and 32,617 bp, designated regions 1
and 2, respectively. Region 1 retains a larger part of Salmonella enterica
serovar Typhi plasmid pHCM2 resembling the backbone of pFra replicons,
while region 2 contains 25 open reading frames with high levels of
similarity to the transfer genes of the F-like plasmids. Surprisingly,
region 1 is also present in the pFra plasmid of avirulent Y. pestis strain
91001 isolated in Inner Mongolia, People's Republic of China. Despite the
fact that some genes typically involved in conjugative transfer of the
F-like replicons are missing in pG8786, we cannot exclude the possibility
that pG8786 might be transmissive under certain conditions. pG8786 seems
to be an ancient form of the pFra group of plasmids that were conserved
due to the strict geographical isolation of rhamnose-positive Y. pestis
strains in the high mountainous Caucasian plague locus.

<>

<1>Golyshin, P.N., Werner, J., Chernikova, T.N., Tran, H., Ferrer, M., Yakimov, M.M., Teeling, H., Golyshina, O.V.
<2>Genome Sequence of Thalassolituus oleivorans MIL-1 (DSM 14913T).
<3>Genome Announcements
<4>1
<5>e00141-13
<6>2013
<7>Thalassolituus oleivorans is one of the most prevalent marine gammaproteobacteria in microbial
communities, emerging after oil spills in coastal, estuarine, and
surface seawaters. Here, we present the assembled genome of strain T. oleivorans
MIL-1 (DSM 14913(T)), which is 3,920,328 bp with a G+C content of 46.6%.

<>

<1>Gomes, L.H., Otto, T.D., Vasconcellos, E.A., Ferrao, P.M., Maia, R.M., Moreira, A.S., Ferreira, M.A., Castello-Branco, L.R., Degrave, W.M., Mendonca-Lima, L.
<2>Genome Sequence of Mycobacterium bovis BCG Moreau, the Brazilian Vaccine Strain against Tuberculosis.
<3>J. Bacteriol.
<4>193
<5>5600-5601
<6>2011
<7>Mycobacterium bovis bacillus Calmette-Guerin (BCG) is the only vaccine available against
tuberculosis, and the strains used worldwide represent a
family of daughter strains with distinct genotypic characteristics. Here
we report the complete genome sequence of M. bovis BCG Moreau, the strain
in continuous use in Brazil for vaccine production since the 1920s.

<>

<1>Gomez, P., Ribas-Aparicio, R.M., Pelaez, A.I., Gomez, A., Rodicio, M.R.
<2>Isolation and nucleotide sequence of the gene encoding the XamI DNA methyltransferase of Xanthomonas campestris pv. amaranthicola.
<3>Biochim. Biophys. Acta
<4>1351
<5>261-266
<6>1997
<7>The gene (xamIM) encoding the DNA methyltransferase of the XamI restriction-modification
system from Xanthomonas campestris pv. amaranthicola (M.XamI) has been cloned in Escherichia
coli and its nucleotide sequence determined.  The sequence predicts a protein of 527 amino
acids that contains nine conserved motifs characteristic of DNA amino methyltransferases.  In
fact, M.XamI shows significant similarity with N6-adenine methyltransferases of the gamma
group of amino methyltransferases, including M.SalI (from the isoschizomeric SalI
restriction-modification system) and M.TaqI (the only N6-adenine methyltransferase for which a
three-dimensional structure is available).  M.XamI and M.SalI share two highly conserved
regions within the C-terminal domain, one of which aligns with one of the DNA recognition
loops proposed for M.TaqI.  Analysis of the chromosomal DNA adjacent to xamIM led to the
identification of an additional ORF (275 codons), downstream, in the same transcriptional
orientation. Although some limited similarities between the SalI restriction enzyme and the
product deduced from this ORF were found, the clone carrying xamIM did not express the
expected endonuclease function.

<>

<1>Gomez, P., Ribas-Aparicio, R.M., Pelaez, A.I., Rodicio, M.R.
<2>Characterization of IS1389, a new member of the IS3 family of insertion sequences isolated from Xanthomonas campestris pv. amaranthicola.
<3>Arch. Microbiol.
<4>172
<5>15-21
<6>1999
<7>IS1389, a new insertion sequence belonging to the IS3 family, has been identified in
Xanthomonas campestris pv. amaranthicola. The genome of this bacterium contains at least 11
copies of the element, whereas no hybridizing sequences were detected in other Xanthomonas
species [X. axonopodis, X. fragaridae, X. phaseoli, and X. (Stenotrophomonas) maltophila]. Two
nearly identical copies of the element (IS1389-A and IS1389-B) were characterized. According
to analysis of sequence alignments and similar structural features, IS1389 belongs to the
IS407 subgroup of the IS3 family, which duplicates 4 bp of target DNA upon insertion. IS1389-A
was found in the proximity of the modification gene of the XamI restriction-modification
system.

<>

<1>Gomez-Consarnau, L., Gonzalez, J.M., Coll-Llado, M., Gourdon, P., Pascher, T., Neutze, R., Pedros-Alio, C., Pinhassi, J.
<2>Light stimulates growth of proteorhodopsin-containing marine Flavobacteria.
<3>Nature
<4>445
<5>210-213
<6>2007
<7>Proteorhodopsins are bacterial light-dependent proton pumps. Their discovery
within genomic material from uncultivated marine bacterioplankton caused
considerable excitement because it indicated a potential phototrophic function
within these organisms, which had previously been considered strictly
chemotrophic. Subsequent studies established that sequences encoding
proteorhodopsin are broadly distributed throughout the world's oceans.
Nevertheless, the role of proteorhodopsins in native marine bacteria is still
unknown. Here we show, from an analysis of the complete genomes of three marine
Flavobacteria, that cultivated bacteria in the phylum Bacteroidetes, one of the
principal components of marine bacterioplankton, contain proteorhodopsin.
Moreover, growth experiments in both natural and artificial seawater (low in
labile organic matter, which is typical of the world's oceans) establish that
exposure to light results in a marked increase in the cell yield of one such
bacterium (Dokdonia sp. strain MED134) when compared with cells grown in
darkness. Thus, our results show that the phototrophy conferred by
proteorhodopsin can provide critical amounts of energy, not only for respiration
and maintenance but also for active growth of marine bacterioplankton in their
natural environment.

<>

<1>Gomez-Eichelmann, M.C., Levy-Mustri, A., Ramirez-Santos, J.
<2>Presence of 5-methylcytosine in CC(A/T)GG sequences (Dcm methylation) in DNAs from different bacteria.
<3>J. Bacteriol.
<4>173
<5>7692-7694
<6>1991
<7>The presence of CC(A/T)GG sequences with methylated internal cytosine (Dcm methylation) was
determined in DNA from different genera of eubacteria. This methylation was studied by using
restriction enzymes EcoRII and BstNI, which cleave unmethylated or methylated CC(A/T)GG
sequences. Dcm methylation was only detected in genera of the family Enterobacteriaceae
closely related to Escherichia: Shigella, Citrobacter, Salmonella, and Klebsiella.

<>

<1>Gomez-Eichelmann, M.C., Ramirez-Santos, J.
<2>Methylated cytosine at Dcm (CCA/TGG) sites in Escherichia coli:  Possible function and evolutionary implications.
<3>J. Mol. Evol.
<4>37
<5>11-24
<6>1993
<7>The frequency and distribution of methylated cytosine (5-MeC) at CCA/TGG (Dcm sites) in 49 E.
coli DNA loci (207,530 bp) were determined. Principal observations of this analysis were: (1)
Dcm frequency was higher than expected from random occurrence but lower than calculated with
Markov chain analysis; (2) CCTGG sites were found more frequently in coding than in noncoding
regions, while the opposite was true for CCAGG sites; (3) Dcm site distribution does not
exhibit any identifiably regular pattern on the chromosome; (4) Dcm sites at oriC are probably
not important for accurate initiation of DNA replication; (5) 5-MeC in codons was more
frequently found in first than in second and third positions; (6) there are probably few genes
in which the mutation rate is determined mainly be DNA methylation. It is proposed that the
function of Dcm methylase is to protect chromosomal DNA from restriction-enzyme EcoRII. The
Dcm methylation contribution to determine frequency of oligonucleotides, mutation rate, and
recombination level, and thus evolution of the E. coli genome, could be interpreted as a
consequence of the acquisition of this methylation.

<>

<1>Gomez-Gil, B., Soto-Rodriguez, S., Lozano, R., Betancourt-Lozano, M.
<2>Draft Genome Sequence of Vibrio parahaemolyticus Strain M0605, Which Causes Severe Mortalities of Shrimps in Mexico.
<3>Genome Announcements
<4>2
<5>e00055-14
<6>2014
<7>Acute hepatopancreatic necrosis disease (AHPND), also known as early mortality syndrome (EMS),
causes high mortalities in cultured shrimps in Asia (L. Tran et
al., Dis. Aquat. Organ. 105:45-55, 2013, http://dx.doi.org/10.3354/dao02621).
Here, we report the draft genome sequence of one Mexican strain of Vibrio
parahaemolyticus that causes similar clinical signs in diseased shrimps.

<>

<1>Gomez-Jimenez, S., Noriega-Orozco, L., Sotelo-Mundo, R.R., Cantu-Robles, V.A., Cobian-Guemes, A.G., Cota-Verdugo, R.G., Gamez-Alejo, L.A., Del Pozo-Yauner, L., Guevara-Hernandez, E., Garcia-Orozco, K.D., Lopez-Zavala, A.A., Ochoa-Leyva, A.
<2>High-Quality Draft Genomes of Two Vibrio parahaemolyticus Strains Aid in Understanding Acute Hepatopancreatic Necrosis Disease of Cultured Shrimps in  Mexico.
<3>Genome Announcements
<4>2
<5>e00800-14
<6>2014
<7>The high-quality draft genomes of two Vibrio parahaemolyticus strains, one that causes the
acute hepatopancreatic necrosis disease (AHPND) in cultured shrimps
(FIM-S1708(+)), and another that does not (FIM-S1392(-)) are reported. A
chromosome-scale assembly for the FIM-S1392(-) genome is reported here. The
analysis of the two genomes gives some clues regarding the genomic differences
between the strains.

<>

<1>Gomez-Pereira, P.R., Schuler, M., Fuchs, B.M., Bennke, C., Teeling, H., Waldmann, J., Richter, M., Barbe, V., Bataille, E., Glockner, F.O., Amann, R.
<2>Genomic content of uncultured Bacteroidetes from contrasting oceanic provinces in the North Atlantic Ocean.
<3>Environ. Microbiol.
<4>14
<5>52-66
<6>2012
<7>Bacteroidetes are widespread in marine systems where they play a crucial role in
organic matter degradation. Whole genome analysis of several strains has revealed
a broad glycolytic and proteolytic potential. In this study, we used a targeted
metagenomic approach to investigate the degradation capabilities of distinct
Bacteroidetes clades from two contrasting regions of the North Atlantic Ocean,
the Polar Biome (BPLR) and the North Atlantic Subtropical (NAST). We present here
the analysis of 76 Bacteroidetes fosmids, of which 28 encode the 16S rRNA gene as
phylogenetic marker, and their comparison to complete Bacteroidetes genomes.
Almost all of the 16S rRNA harbouring fosmids belonged to clades that we
previously identified in BPLR and NAST. The majority of sequenced fosmids could
be assigned to Bacteroidetes affiliated with the class Flavobacteria. We also
present novel genomic information on the classes Cytophagia and Sphingobacteria,
suggesting a capability of the latter for attachment to algal surfaces. In our
fosmid set we identified a larger potential for polysaccharide degradation and
cell surface attachment in the phytoplankton-rich BPLR. Particularly, two
flavobacterial fosmids, one affiliated with the genus Polaribacter, showed a
whole armoury of enzymes that likely function in degradation of sulfated
polysaccharides known to be major constituents of phytoplankton cell walls. Genes
involved in protein and peptidoglycan degradation, although present in both
fosmid sets, seemed to have a slight preponderance in NAST. This study provides
support for the hypothesis of a distinct specialization among marine
Bacteroidetes for the degradation of certain types of polymers.

<>

<1>Gomez-Rodriguez, J.A., Huete-Perez, J.A.
<2>Bioprospeccion de enzimas de restriccion en bacterias de suelos y ambientes volcanicos de Nicaragua.
<3>Encuentro
<4>15
<5>70-87
<6>2008
<7>
<>

<1>Gonchar, D.A., Abdurashitov, M.A., Belichenko, O.A., Dedkov, V.S., Mezentseva, N.V., Tomilova, J.E., Degtyarev, S.K.
<2>PspXI, a novel restriction endonuclease that recognizes the unusual  DNA sequence 5'-VC^TCGAGB-3'.
<3>Ovchinnikov Bull. Biotechnol. Phys. Chem. Biol.
<4>1
<5>18-23
<6>2005
<7>We have discovered a bacterial strain Pseudomonas species X11 that produces the novel
restriction endonuclease PspXI. This enzyme recognizes an unusual degenerate octanucleotide
sequence 5'-VCTCGAGB-3', where V stands for A, C or G and B stands for T, C or G.  The PspXI
restriction endonuclease preparation with concentration of 10000 units/ml was isolated using
four chromatographic steps. PspXI cuts its recognition sequence between C and T producing
cohesive ends compatible with those of produced by XhoI(5'-C^TCGAG-3') and
SalI(5'-G^TCGAC-3') restriction endonucleases.

<>

<1>Gonchar, D.A., Abdurashitov, M.A., Okhapkina, S.S., Shagin, D.A., Kileva, E.V., Degtyarev, S.K.
<2>Sse9I restriction-modification system: Organization of genes and structural comparison of proteins.
<3>Mol. Biol. (Mosk)
<4>41
<5>491-498
<6>2007
<7>The nucleotide sequence was established for the operon of the Sse9I type II
restriction-modification system of Sporosarcina species 9D.  The enzymes of the Sse9I system
recognize the 5'-AATT-3' tetranucleotide.  The operon includes three genes,
sse9IC-sse9IR-sse9IM, which are transcribed unidirectionally and code, respectively, for the
controller protein (C.Sse9I), restriction endonuclease (R.Sse9I), and DNA methyltransferase
(M.Sse9I).  The region immediately upstream of sse9IC was found to contain a conserved
nucleotide sequence (C box) providing a binding site for C.Sse9I.  The amino acid sequences of
C.Sse9I and R.Sse9I were compared with those of related proteins.  In the case of R.Sse9I, the
highest homology was observed with the R.MunI (5'CAATTG-3') and R.EcoRI (5'GAATTC-3')
regions that harbor the amino acid residues involved in recognizing the AATT inner
tetranucleotide.  The sse9IR gene was cloned in an expression vector, and recombinant R.Sse9I
was isolated.

<>

<1>Gonchar, D.A., Akishev, A.G., Degtyarev, S.Kh.
<2>BlsI- and GlaI-PCR assays - a new method of DNA methylation study.
<3>Ovchinnikov Bull. Biotechnol. Phys. Chem. Biol.
<4>6
<5>5-12
<6>2010
<7>Regulation of genes activity in mammalians genomes is based on DNA methylation of CG
dinucleotides with formation of 5-methylcytosine (5mC) in both DNA strands.  Mammalian
DNA-methyltransferases Dnmt1, Dnmt3a and Dnmt3b catalyze a reaction of DND methylation.  Dnmt1
maintains DNA methylation pattern in vivo modifying a new strand after replication.  Dnmt3a
and Dnmt3b are responsible for DNA methylation de novo and, likely, differ in their function
and preferable region of modification.  Study of Dnmt3a and Dnmnt3b substrate specificity has
shown that both enzymes methylate CG-dinucleotide4 mostly in DNA sequence PuCGPy.  At present
time 5mC determination is represented mostly by a chemical treatment of DNA with sodium
bisulphite, which results in cytosine transformation into uracil, and native DNA allows to
locate positions of methylated cytosines in studied DNA.  Method of bisulphite conversion is
quite expensive, time-consuming and often results in obtaining false positive data.  Because
of substantial DNA degradation this method is used for analysis of only short (100 - 150 bp)
DNA sequences.  Among enzymatic methods of 5mC determination, so called methyl-sensitive PCR
assay is the most popular.  This method is based on inability of restriction enzymes, which
contain CG dinucleotide in the recognition site, to cut this site if 5mC is present in the
dinucleotide.  A subsequent PCR from primers, which are located around a chosen recognition
site, produces a corresponding DNA fragment if there is a methylated CG-dinucleotide within
this site.  On the contrary, DNA fragment is not produced in PCR if there is no methylated
CG-dinucleotide in a recognition sequence of restriction enzyme.  Application of
methyl-sensitive PCR assay is limited by a very short list of recognition sequences.  Recently
we have discovered and characterized absolutely new enzymes BlsI and GlaI, which belong to the
type of methyl-directed site-specific DNA endonucleases and cleave only methylated DNA.  GlaI
recognizes DNA sequence 5'-Pu(5mC)GPy-3'/3'-PyG(5mC)Pu-5', whereas BlsI hydrolyzes DNA
sequence 5'-GCNGC-3' if at least one 5-methylcytosine (N isn't considering) is present in
each DNA strand of the recognition site.  In this work we have developed a new method of DNA
methylation determination -
BlsI- and GlaI-PCR assays, and have applied this method to study a methylation of regulation
of tumor suppressor genes in DNA from different human cell lines.

<>

<1>Gonchar, D.A., Dedkov, V.S., Verkhozina, V.A., Kusner, Y.S., Shevchenko, A.V., Degtyarev, S.K.
<2>Restriction endonuclease Sse9I from Sporosarcina sp. strain 9D recognizes the 5'-AATT-3' DNA sequence.
<3>Prikl. Biokhim. Mikrobiol.
<4>34
<5>139-141
<6>1998
<7>A new restriction endonuclease Sse9I was isolated from the bacterial strain Sporosarcina sp.
9D. The enzyme belongs to type II restrictases and recognizes the tetranucleotide sequence
5'-AATT-3'.  The enzyme cleaves DNA before the first adenine residue, so it is a true
isoschizomer of Tsp509I restrictase.  However, unlike the prototype, Sse9I digests DNA at 55oC
and loses its activity after 20 min storage at 65oC.

<>

<1>Gonchar, D.A., Dedkov, V.S., Verkhozina, V.A., Kusner, Y.S., Shevchenko, A.V., Degtyarev, S.K.
<2>Sse9I, a restriction endonuclease from Sporosarcina sp. 9D which recognizes the sequence 5'AATT-3'.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>2
<5>32-34
<6>1998
<7>Sse9I, a type II restriction endonuclease, has been isolated from Sporosarcina species 9D.
The recognition sequence and cleavage point of restriction endonuclease Sse9I have been
determined as 5'-/AATT-3'.  The new enzyme is an isoschizomer of Tsp509I, but its optimal
incubation temperature is 55 C and it is inactivated at 65 C for 20 min.

<>

<1>Gonchar, D.A., Wolf, Y.I., Degtyarev, S.K.
<2>Cloning and characterization of Sse9I DNA-methyltransferase recognizing 5'-AATT-3'.
<3>Nucleic Acids Res.
<4>24
<5>2790-2792
<6>1996
<7>The gene from Sporosarcina species 9D encoding Sse9I DNA-methyltransferase (M.Sse9I) was
cloned and expressed in Escherichia coli.  The recombinant plasmid pMSse-1 contains the
M.Sse9I gene 1086 bp in length, corresponding to a protein of 362 amino acid residues.
M.Sse9I recognizes the tetranucleotide sequence 5'-AATT-3' and modifies the second adenine
within the recognition sequence.  The amino acid sequence of M.Sse9I was compared with those
of other methylases.  According to mutual positions of four conservative domains the new
enzyme belongs to a subgroup of D12 class.  This subgroup includes Sse9I, CviAII, NlaIII and
N-terminal domains of LlaI, FokI and StsI DNA-methyltransferases.

<>

<1>Gong, W., Kisiela, M., Schilhabel, M.B., Xiong, G., Maser, E.
<2>Genome Sequence of Comamonas testosteroni ATCC 11996, a Representative Strain Involved in Steroid Degradation.
<3>J. Bacteriol.
<4>194
<5>1633-1634
<6>2012
<7>Comamonas testosteroni strains belong to the family of Comamonadaceae and are known for their
ability to utilize steroid compounds as carbon source. Here, we
present the draft genome sequence of strain ATCC 11996, with a G+C content of
61.48%.

<>

<1>Gong, W., O'Gara, M., Blumenthal, R.M., Cheng, X.
<2>Structure of PvuII DNA-(cytosine N4) methyltransferase, an example of domain permutation and protein fold assignment.
<3>Nucleic Acids Res.
<4>25
<5>2702-2715
<6>1997
<7>We have determined the structure of PvuII methyltransferase (M.PvuII) complexed with
S-adenosyl-L-methionine (AdoMet) by multiwavelength anomalous diffraction, using a crystal of
the selenomethionine-substituted protein.  M.PvuII catalyzes transfer of the methyl group from
AdoMet to the exocyclic amino (N4) nitrogen of the central cytosine in its recognition
sequence 5'-CAGCTG-3'.  The protein is dominated by an open alpha/beta-sheet stucture with a
prominent V-shaped cleft: AdoMet and catalytic amino acids are located at the bottom of this
cleft.  The size and the basic nature of the cleft are consistent with duplex DNA binding.
The target (methylatable) cytosine, if flipped out of the double helical DNA as seen for DNA
methyltransferases that generate 5-methylcytosine, would fit into the concave active site next
to the AdoMet.  This M.PvuII alpha/beta-sheet structure is very similar to those of M.HhaI (a
cytosine C5 methyltransferase) and M.TaqI (an adenine N6 methyltransferase), consistent with a
model predicting that DNA methyltransferases share a common structural fold while having the
major functional regions permuted into three distinct linear orders.  The main feature of the
common fold is a seven-stranded beta-sheet (6/7/5/4/1/2/3/) formed by five parallel
beta-strands and an antiparallel beta-hairpin.  The beta-sheet is flanked by size parallel
alpha-helices, three on each side.  The AdoMet binding site is located at the C-terminal ends
of strands beta1 and beta2 and the active site is at the C-terminal ends of strands beta4 and
beta5 and the N-terminal end of strand beta7.  The AdoMet-protein interactions are almost
identical among M.PvuII, M.HhaI and M.TaqI, as well as in an RNA methyltransferase and at
least one small molecule methyltransferase.  The structural similarity among the active sites
of M.PvuII, M.TaqI and M.HhaI reveals that catalytic amino acids essential for cytosine N4 and
adenine N6 methylation coincide spatially with those for cytosine C5 methylation, suggesting a
mechanism for amino methylation.

<>

<1>Gonzaga, A., Lopez-Perez, M., Martin-Cuadrado, A.B., Ghai, R., Rodriguez-Valera, F.
<2>Complete Genome Sequence of the Copiotrophic Marine Bacterium Alteromonas macleodii Strain ATCC 27126T.
<3>J. Bacteriol.
<4>194
<5>6998
<6>2012
<7>The genome of Alteromonas macleodii strain ATCC 27126(T) has been resequenced and closed into
a single contig. We describe here the genome of this important and
globally distributed marine bacterium.

<>

<1>Gonzaga, A., Martin-Cuadrado, A.B., Lopez-Perez, M., Megumi, M., C, G.-H., I, K., N, E., Lopez-Garcia, P., Moreira, D., Ussery, D., Zaballos, M., Ghai, R., Rodriguez-Valera, F.
<2>Polyclonality of Concurrent Natural Populations of Alteromonas macleodii.
<3>Genome Biol. Evol.
<4>4
<5>1360-1374
<6>2012
<7>We have analyzed a natural population of the marine bacterium, Alteromonas
macleodii, from a single sample of seawater to evaluate the genomic diversity
present. We performed full genome sequencing of four isolates and 161 metagenomic
fosmid clones, all of which were assigned to A. macleodii by sequence similarity.
Out of the four strain genomes, A. macleodii deep ecotype (AltDE1) represented a
different genome, whereas AltDE2 and AltDE3 were identical to the previously
described AltDE. Although the core genome ( approximately 80%) had an average
nucleotide identity of 98.51%, both AltDE and AltDE1 contained flexible genomic
islands (fGIs), that is, genomic islands present in both genomes in the same
genomic context but having different gene content. Some of the fGIs encode cell
surface receptors known to be phage recognition targets, such as the O-chain of
the lipopolysaccharide, whereas others have genes involved in physiological
traits (e.g., nutrient transport, degradation, and metal resistance) denoting
microniche specialization. The presence in metagenomic fosmids of genomic
fragments differing from the sequenced strain genomes, together with the presence
of new fGIs, indicates that there are at least two more A. macleodii clones
present. The availability of three or more sequences overlapping the same genomic
region also allowed us to estimate the frequency and distribution of
recombination events among these different clones, indicating that these
clustered near the genomic islands. The results indicate that this natural A.
macleodii population has multiple clones with a potential for different phage
susceptibility and exploitation of resources, within a seemingly unstructured
habitat.

<>

<1>Gonzalez, D., Kozdon, J.B., McAdams, H.H., Shapiro, L., Collier, J.
<2>The functions of DNA methylation by CcrM in Caulobacter crescentus: a global approach.
<3>Nucleic Acids Res.
<4>42
<5>3720-3735
<6>2014
<7>DNA methylation is involved in a diversity of processes in bacteria, including maintenance of
genome integrity and regulation of gene expression. Here, using
Caulobacter crescentus as a model, we exploit genome-wide experimental methods to
uncover the functions of CcrM, a DNA methyltransferase conserved in most
Alphaproteobacteria. Using single molecule sequencing, we provide evidence that
most CcrM target motifs (GANTC) switch from a fully methylated to a
hemi-methylated state when they are replicated, and back to a fully methylated
state at the onset of cell division. We show that DNA methylation by CcrM is not
required for the control of the initiation of chromosome replication or for DNA
mismatch repair. By contrast, our transcriptome analysis shows that >10% of the
genes are misexpressed in cells lacking or constitutively over-expressing CcrM.
Strikingly, GANTC methylation is needed for the efficient transcription of dozens
of genes that are essential for cell cycle progression, in particular for DNA
metabolism and cell division. Many of them are controlled by promoters methylated
by CcrM and co-regulated by other global cell cycle regulators, demonstrating an
extensive cross talk between DNA methylation and the complex regulatory network
that controls the cell cycle of C. crescentus and, presumably, of many other
Alphaproteobacteria.

<>

<1>Gonzalez, E., Padilla, C., Saavedra, C., Vasquez, C.
<2>The expression of the bstVIM gene from Bacillus stearothermophilus V is restricted to vegetative cell growth.
<3>Microbiology
<4>140
<5>1337-1340
<6>1994
<7>The activity of BstVI DNA methyltransferase was monitored during the sporulative cycle of
Bacillus stearothermophilus V. Significant methylase activity was found only in bacteria
growing vegetatively. This was confirmed by Northern hybridization, which indicated that the
bstVIM gene was not transcribed in cells undergoing sporulation. Supporting evidence came from
experiments which demonstrated that the RNA polymerase holoenzyme from these cells did not
recognize the promoter elements upstream of the bstVIM gene.

<>

<1>Gonzalez, E., Vasquez, C.
<2>Characterization of the bstVIRM genes encoding the Bacillus stearothermophilus V restriction-modification system.
<3>Gene
<4>131
<5>103-106
<6>1993
<7>The nucleotide (nt) sequence of a 2.7-kb HindIII-EcoRI DNA fragment encoding the bstVIR and
bstVIM genes has been determined. The sequence predicts a restriction endonuclease of 224
amino acids (aa), Mr 25,104, and a methyltransferase of 561 aa, Mr 65,702. Both genes are
aligned in the same orientation and are separated by a 102-nt intergenic region. No homology
was found between R-BstVI and M-BstVI when their deduced aa sequences were compared.
Significant similarity at the aa level was found, however, when both enzymes were compared to
their equivalents in the paeR71RM system of Pseudomonas aeruginosa PAO303.

<>

<1>Gonzalez, V., Santamaria, R.I., Bustos, P., Hernandez-Gonzalez, I., Medrano-Soto, A., Moreno-Hagelsieb, G., Janga, S.C., Ramirez, M.A., Jimenez-Jacinto, V., Collado-Vides, J., Davila, G.
<2>The partitioned Rhizobium etli genome: Genetic and metabolic redundancy in seven interacting replicons.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>3834-3839
<6>2006
<7>We report the complete 6,530,228-bp genome sequence of the symbiotic nitrogen fixing bacterium
Rhizobium etli. Six large plasmids comprise
one-third of the total genome size. The chromosome encodes most functions
necessary for cell growth, whereas few essential genes or complete
metabolic pathways are located in plasmids. Chromosomal synteny is
disrupted by genes related to insertion sequences, phages, plasmids, and
cell-surface components. Plasmids do not show synteny, and their orthologs
are mostly shared by accessory replicons of species with multipartite
genomes. Some nodulation genes are predicted to be functionally related
with chromosomal loci encoding for the external envelope of the bacterium.
Several pieces of evidence suggest an exogenous origin for the symbiotic
plasmid (p42d) and p42a. Additional putative horizontal gene transfer
events might have contributed to expand the adaptive repertoire of R.
etli, because they include genes involved in small molecule metabolism,
transport, and transcriptional regulation. Twenty-three putative sigma
factors, numerous isozymes, and paralogous families attest to the
metabolic redundancy and the genomic plasticity necessary to sustain the
lifestyle of R. etli in symbiosis and in the soil.

<>

<1>Gonzalez-Ceron, G., Miranda-Olivares, O.J., Servin-Gonzalez, L.
<2>Characterization of the methyl-specific restriction system of Streptomyces coelicolor A3(2) and of the role played by laterally acquired nucleases.
<3>FEMS Microbiol. Lett.
<4>301
<5>35-43
<6>2009
<7>The methyl-specific restriction system of Streptomyces coelicolor A3(2) was analyzed by
carrying out transformations with unmethylated and
methylated pSET152 DNA. Streptomyces coelicolor was found to strongly
restrict DNA methylated in vivo by the Dam, Dcm and Hsd modification
systems of Escherichia coli. Hsd-modified DNA was restricted as
strongly as Dam-modified DNA, even though there are significantly fewer
sites on the plasmid; Dcm-modified plasmid was restricted more strongly
then either Dam-or Hsd-modified DNA. Restriction of plasmid DNA
modified in vitro by different methylases also showed a greater
dependence on the methylated sequence than on the number of methylated
sites. Streptomyces coelicolor mutants were constructed that lacked
genes identified as the likely candidates for encoding methyl-specific
restriction nucleases (the products of the SCO4213, SCO4631 and SCO2863
genes, as well as the SCO3261-SCO3262 operon) that are located in the
laterally acquired genomic islands of the S. coelicolor chromosome;
these mutants showed partial alleviation of methylated DNA restriction.
Cloning of these genes in the close relative Streptomyces lividans
increased the restriction of methylated DNA by this species, confirming
their role as part of the methyl-specific restriction system of S.
coelicolor.

<>

<1>Gonzalez-Escalona, N., McFarland, M.A., Rump, L.V., Payne, J., Andrzejewski, D., Brown, E.W., Evans, P.S., Croley, T.R.
<2>Draft Genome Sequences of Two O104:H21 Escherichia coli Isolates Causing Hemorrhagic Colitis during a 1994 Montana Outbreak Provide Insight into Their  Pathogenicity.
<3>Genome Announcements
<4>1
<5>e00805-13
<6>2013
<7>We sequenced the genomes of two strains of O104:H21 enterohemorrhagic Escherichia coli (EHEC)
isolated during an outbreak of hemorrhagic colitis in Montana in
1994. These strains carried a plasmid that contains several virulence genes not
present in pO157. The genome sequences will improve phylogenetic analysis of
other non-O157 E. coli strains in the future.

<>

<1>Gonzalez-Escalona, N., Strain, E.A., De Jesus, A.J., Jones, J.L., Depaola, A.
<2>Genome sequence of the clinical O4:K12 serotype Vibrio parahaemolyticus strain 10329.
<3>J. Bacteriol.
<4>193
<5>3405-3406
<6>2011
<7>Vibrio parahaemolyticus is the leading cause of foodborne illnesses worldwide. Here we report
a draft genome of V. parahaemolyticus strain
Vp10329 of O4:K12 serotype. It belongs to the main U. S. West Coast clonal
complex of V. parahaemolyticus (ST36) causing oyster-associated human
illness. It contains the virulence determinants tdh and trh but appears to
infect at much lower doses than V. parahaemolyticus strains with these
same determinants from other areas such as the U.S. Gulf and Atlantic
coasts.

<>

<1>Gonzalez-Escalona, N., Thirunavukkarasu, N., Singh, A., Toro, M., Brown, E.W., Zink, D., Rummel, A., Sharma, S.K.
<2>Draft Genome Sequence of Bivalent Clostridium botulinum Strain IBCA10-7060, Encoding Botulinum Neurotoxin B and a New FA Mosaic Type.
<3>Genome Announcements
<4>2
<5>e01275-14
<6>2014
<7>Here we report the genome sequence of a Clostridium botulinum strain IBCA10-7060  producing
botulinum neurotoxin serotype B and a new toxin serotype. Multilocus
sequence typing analysis revealed that this strain belongs to a new sequence
type, and whole-genome single nucleotide polymorphism analysis showed that this
strain clustered with strains in lineage 2 from group I.

<>

<1>Gonzalez-Nicieza, R., Turner, D.P., Connolly, B.A.
<2>DNA binding and cleavage selectivity of the Escherichia coli DNA G:T-mismatch endonuclease (vsr protein).
<3>J. Mol. Biol.
<4>310
<5>501-508
<6>2001
<7>The Escherichia coli vsr endonuclease recognises T:G base-pair mismatches in double-stranded
DNA and initiates a repair pathway by
hydrolysing the phosphate group 5' to the incorrectly paired T. The
gene encoding the vsr endonuclease is next to the gene specifying the
E. coli dcm DNA-methyltransferase; an enzyme that adds CH3 groups to
the first dC within its target sequence CC[A/T]GG, giving
C5MeC[A/T]GG. Deamination of the d(5Me)C results in CT[A/T]GG in
which the first T is mis-paired with dG and it is believed that the
endonuclease preferentially recognises T:G mismatches within the dcm
recognition site. Here, the preference of the vsr endonuclease for
bases surrounding the T:G mismatch has been evaluated. Determination of
specificity constant (k(st)/K-D; k(st) = rate constant for single
turnover, K-D = equilibrium dissociation constant) confirms vsr's
preference for a T:G mismatch within a dcm sequence i.e. C (T) under
bar [A/T]GG (the underlined T being mis-paired with dG) is the best
substrate. However, the enzyme is capable of binding and hydrolysing
sequences that differ from the dcm target site by a single base-pair
(dcm star sites). Individual alteration of any of the four bases
surrounding the mismatched T gives a substrate, albeit with reduced
binding affinity and slowed turnover rates. The vsr endonuclease has a
much lower selectivity for the dcm sequence than type II restriction
endonucleases have for their target sites. The results are discussed in
the light of the known crystal structure of the vsr protein and its
possible physiological role.

<>

<1>Gonzalez-Perez, M., Murcia, M.I., Landsman, D., Jordan, I.K., Marino-Ramirez, L.
<2>Genome Sequence of the Mycobacterium colombiense Type Strain, CECT 3035.
<3>J. Bacteriol.
<4>193
<5>5866-5867
<6>2011
<7>We report the first whole-genome sequence of the Mycobacterium colombiense type strain, CECT
3035, which was initially isolated from Colombian
HIV-positive patients and causes respiratory and disseminated infections.
Preliminary comparative analyses indicate that the M. colombiense lineage
has experienced a substantial genome expansion, possibly contributing to
its distinct pathogenic capacity.

<>

<1>Gonzalgo, M.L., Jones, P.A.
<2>Mutagenic and epigenetic effects of DNA methylation.
<3>Mutat. Res.
<4>386
<5>107-118
<6>1997
<7>Tumorigenesis begins with the disregulated growth of an abnormal cell that has acquired the
ability to divide more rapidly than its normal counterparts.  Alterations in global levels and
regional changes in the patterns of DNA methylation are among the earliest and most frequent
events known to occur in human cancers.  These changes in methylation may impair the proper
expression and/or function of cell-cycle regulatory genes and thus confer a selective growth
advantage to affected cells.  Developments in the field of cancer research over the past few
years have led to an increased understanding of the role DNA methylation may play in
tumorigenesis.  Many of these studies have investigated two major mechanisms by which DNA
methylation may lead to aberrant cell cycle control: (1) through the generation of transition
mutations via deamination-driven events resulting in the inactivation of tumor suppressor
genes, or (2) by altering levels of gene expression through epigenetic effects at CpG islands.
The mechanisms by which the normal function of growth regulatory genes may become affected by
the mutagenic and epigenetic properties of DNA methylation will be discussed in the framework
of recent discoveries in the field.

<>

<1>Goodgal, S.H., Gromkova, R.
<2>Separation of specific segments of transforming DNA after treatment with endodeoxyribonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>70
<5>503-506
<6>1973
<7>Hemophilus parainfluenzae endodeoxyribonuclease was used to degrade the DNA of
H. influenzae and to follow the biological activity of 14 markers associated
with this DNA.  It was found that some H. influenzae markers were completely
inactivated by endodeoxyribonuclease treatment, while others appeared to retain
all or almost all of their original activity.  The bulk of the H. influenzae
DNA was reduced to double-stranded pieces of the order of 0.8 1 Mdaltons.
Velocity sedimentation of the DNA in sucrose gradients disclosed that markers
that retained biological activity were present in DNA particles that were of
the order of 1 Mdaltons or larger, and indicated a close correlation between
the size of the DNA fragment and the amount of biological activity retained.
These data suggest that H. parainfluenzae endodeoxyribonuclease breaks DNA at
specific sites.  The nalr marker was shown to have twice as much biological
activity after treatment with endodeoxyribonuclease when assayed at saturating
DNA concentrations.  In the linear portion of the DNA dose-response curve, the
biological activity of this marker was reduced 3- to 10-fold compared to
untreated DNA (in accord with the reduced size of its DNA).  These data
demonstrate a specific enrichment of the nalr marker by about 6- to 20-fold,
and suggest a technique for the separation and purification of specific
segments of DNA.

<>

<1>Goodison, S., Urquidi, V., Kumar, D., Reyes, L., Rosser, C.J.
<2>Complete Genome Sequence of Mycoplasma hyorhinis Strain SK76.
<3>Genome Announcements
<4>1
<5>e00101-12
<6>2013
<7>Mycoplasma hyorhinis is a eubacterium belonging to the Mollicutes class and is responsible for
porcine respiratory and arthritic diseases. It is also the major
contaminant of mammalian tissue cultures in laboratories worldwide. Here, we
report the complete genome sequence of M. hyorhinis strain SK76.

<>

<1>Goodman, H.M., Greene, P.J., Garfin, D.E., Boyer, H.W.
<2>DNA site recognition by the EcoRI restriction endonuclease and modification methylase.
<3>Nucleic Acid - Protein Recognition, Academic Press, Vogel, H.J., New York
<4>0
<5>239-259
<6>1977
<7>None

<>

<1>Goodner, B. et al.
<2>Genome sequence of the plant pathogen and biotechnology agent Agrobacterium tumefaciens C58.
<3>Science
<4>294
<5>2323-2328
<6>2001
<7>Agrobacterium tumefaciens is a plant pathogen capable of transferring a defined segment of DNA
to a host plant, generating a gall tumor. Replacing the transferred  tumor-inducing genes with
exogenous DNA allows the introduction of any desired gene into the plant. Thus, A. tumefaciens
has been critical for the development of modern plant genetics and agricultural biotechnology.
Here we describe the genome of A. tumefaciens strain C58, which has an unusual structure
consisting of one circular and one linear chromosome. We discuss genome architecture and
evolution and additional genes potentially involved in virulence and metabolic parasitism of
host plants.

<>

<1>Goodrich-Blair, H., Scarlato, V., Gott, J.M., Xu, M.-Q., Shub, D.A.
<2>A self-splicing group I intron in the DNA polymerase gene of Bacillus subtilis bacteriophage SPO1.
<3>Cell
<4>63
<5>417-424
<6>1990
<7>We report a self-splicing intron in bacteriophage SPO1, whose host is the gram-positive
Bacillus subtilis. The intron contains all the conserved features of primary sequence and
secondary structure previously described for the group IA introns of eukaryotic organelles and
the gram-negative bacteriophage T4. The SPO1 intron contains an open reading frame of 522
nucleotides. As in the T4 introns, this open reading frame begins in a region that is looped
out of the secondary structure, but ends in a highly conserved region of the intron core. The
exons encode SPO1 DNA polymerase, which is highly similar to E. coli DNA polymerase I. The
demonstration of self-splicing introns in viruses of both gram-positive and gram-negative
eubacteria lends further evidence for their early origin in evolution.

<>

<1>Goodrich-Blair, H., Shub, D.A.
<2>A site- and strand-specific intron-endonuclease is required for marker exclusion.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>177
<6>1993
<7>The virulent Bacillus subtilis bacteriophage SPO1 and SP82 belong to a closely related family
that contain hydroxymethyluracil (HMU) in place of thymine in their DNA. The DNA polymerase
gene of these phage is interrupted by a self-splicing group I intron. We have characterized
two endonucleases, I-HmuI (encoded by SPO1) and I-HmuII (encoded by SP82) that are encoded
entirely within the DNA polymerase intron. Other intron-endonucleases are initiators of
"intron homing". They introduce a double-strand cleavage specifically on intron-less alleles
and repair of this cleavage, using the intron-plus allele as a template, results in two
intron-plus copies of DNA via unidirectional gene conversion. The HMU-phage
intron-endonucleases have several unique features. First, they are strand- as well as
site-specific, introducing a nick in the noncoding strand 4 nucleotides (nt) (I-HmuI) or 54 nt
(I-HmuII) downstream of the 3' splice site of the intron, within exon II of the DNA
polymerase gene. Second, the endonucleases cleave intron-plus as well as intron-less DNA.
Third, the endonucleases show a distinct preference, both in vitro and in vivo, for the DNA of
the heterologous phage. These differences in the HMU intron-endonucleases could reflect a
difference in function. Previous reports indicated that in mixed infections genetic markers of
SP82 are more likely to be carried by progeny than those of SP01. This exclusion occurs over
10 kilobases of DNA surrounding the DNA polymerase gene. We have found that expression of
I-HmuII by SP82 is required for this exclusion process. We hypothesize that the intron homing
mechanism of I-HmuII has been expanded to confer a selective advantage on its host in
competition with close relatives.

<>

<1>Goodrich-Blair, H., Shub, D.A.
<2>The DNA polymerase genes of several HMU-bacteriophages have similar group I introns with highly divergent open reading frames.
<3>Nucleic Acids Res.
<4>22
<5>3715-3721
<6>1994
<7>A previous report described the discovery of a group I, self-splicing intron
in the DNA
polymerase gene of the Bacillus subtilis bacteriophage SPO1.  In this study, the DNA
polymerase genes of
three close relatives of SPO1: SP82, 2C and Phi e, were also found to be interrupted by an
intron.  All of these
introns have group I secondary structures that are extremely similar to one another in
primary sequence.
Each is interrupted by an open reading frame that, unlike the intron core or exon sequences,
are highly
diverged.  Unlike the relaives of Escherichia coli bacteriophage T4, most of which do not
have introns, this
intron seems to be common among the relatives of SPO1.

<>

<1>Goodrich-Blair, H., Shub, D.A.
<2>Beyond homing: Competition between intron endonucleases confers a selective advantage on flanking genetic markers.
<3>Cell
<4>84
<5>211-221
<6>1996
<7>The closely related B. subtilis bacteriophages SPO1 and SP82 have similar introns inserted
into a conserved domain of their DNA polymerase genes.  These introns encode endonucleases
with unique properties.  Other intron-encoded "homing" endonucleases cleave both strands of
intronless DNA; subsequent repair results in unidirectional gene conversion to the
intron-containing allele.  In contrast, the enzymes described here cleave one strand on both
intron-containing and intronless targets at different distances from their common intron
insertion site.  Most surprisingly, each enzyme prefers DNA of the heterologous phage.  The
SP82-encoded endonuclease is responsible for exclusion of the SPO1 intron and flanking genetic
markers from the progeny of mixed infections, a novel selective advantage imparted by an
intron to the genome in which it resides.

<>

<1>Goodsell, D.S.
<2>The molecular perspective: Restriction endonucleases.
<3>Stem Cells
<4>20
<5>190-191
<6>2002
<7>We live in a remarkable age. Physicians now have an unprecedented range of options for helping
their patients. Surgery and radiology are using advanced technologies to pinpoint treatments.
Chemotherapeutic approaches are showing ever-greater effectiveness and specificity,
approaching the goal of a "magic bullet." And we are now entering a time when we can make
changes at the most basic level, introducing changes directly into the genetic code. Gene
therapy, the ability to introduce new genes into living cells, holds great promise for the
treatment of many diseases, including cancer. Restriction endonucleases are the molecules that
spawned the field of biotechnology, the first of the many molecular tools that make gene
therapy a reality. Bacteria build restriction endonucleases to protect themselves from attack
by viruses. These enzymes cleave DNA at a specific sequence of nucleotides: for instance, one
enzyme from Escherichia coli cuts the sequence GAATTC and another enzyme from Bacillus
amyloliquefaciens will not cut it, but instead cuts the similar sequence GGATCC. Each species
of bacterium protects its own DNA at its personal sequence by adding a bulky methyl group on a
few of the bases, so only invading viral DNA, which does not have the protective methyl
groups, is chopped up and destroyed. There are hundreds of forms of these restriction enzymes,
made by different bacteria to cut DNA at different sequences.

<>

<1>Goodsell, D.S.
<2>The Molecular Perspective: Restriction Endonucleases.
<3>The Oncologist
<4>7
<5>82-83
<6>2002
<7>We live in a remarkable age. Physicians now have an unprecedented range of options for helping
their patients. Surgery and radiology are using advanced technologies to pinpoint treatments.
Chemotherapeutic approaches are showing ever-greater effectiveness and specificity,
approaching the goal of a "magic bullet." And we are now entering a time when we can make
changes at the most basic level, introducing changes directly into the genetic code. Gene
therapy, the ability to introduce new genes into living cells, holds great promise for the
treatment of many diseases, including cancer. Restriction endonucleases are the molecules that
spawned the field of biotechnology, the first of the many molecular tools that make gene
therapy a reality. Bacteria build restriction endonucleases to protect themselves from attack
by viruses. These enzymes cleave DNA at a specific sequence of nucleotides: for instance, one
enzyme from Escherichia coli cuts the sequence GAATTC and another enzyme from Bacillus
amyloliquefaciens will not cut it, but instead cuts the similar sequence GGATCC. Each species
of bacterium protects its own DNA at its personal sequence by adding a bulky methyl group on a
few of the bases, so only invading viral DNA, which does not have the protective methyl
groups, is chopped up and destroyed. There are hundreds of forms of these restriction enzymes,
made by different bacteria to cut DNA at different sequences.

<>

<1>Goodsell, D.S.
<2>Recognition in action: flipping pyrimidine dimers.
<3>J. Mol. Recognit.
<4>18
<5>193-195
<6>2005
<7>DNA bases are normally sheltered within a double helix, but enzymes that modify and repair DNA
gain access by flipping individual bases out
of the double helix.

<>

<1>Goodwin, S.B. et al.
<2>Finished Genome of the Fungal Wheat Pathogen Mycosphaerella graminicola Reveals Dispensome Structure, Chromosome Plasticity, and Stealth Pathogenesis.
<3>PLoS Genet.
<4>7
<5>E1002070
<6>2011
<7>The plant-pathogenic fungus Mycosphaerella graminicola (asexual stage:
Septoria tritici) causes septoria tritici blotch, a disease that greatly
reduces the yield and quality of wheat. This disease is economically
important in most wheat-growing areas worldwide and threatens global food
production. Control of the disease has been hampered by a limited
understanding of the genetic and biochemical bases of pathogenicity,
including mechanisms of infection and of resistance in the host. Unlike
most other plant pathogens, M. graminicola has a long latent period during
which it evades host defenses. Although this type of stealth pathogenicity
occurs commonly in Mycosphaerella and other Dothideomycetes, the largest
class of plant-pathogenic fungi, its genetic basis is not known. To
address this problem, the genome of M. graminicola was sequenced
completely. The finished genome contains 21 chromosomes, eight of which
could be lost with no visible effect on the fungus and thus are
dispensable. This eight-chromosome dispensome is dynamic in field and
progeny isolates, is different from the core genome in gene and repeat
content, and appears to have originated by ancient horizontal transfer
from an unknown donor. Synteny plots of the M. graminicola chromosomes
versus those of the only other sequenced Dothideomycete, Stagonospora
nodorum, revealed conservation of gene content but not order or
orientation, suggesting a high rate of intra-chromosomal rearrangement in
one or both species. This observed "mesosynteny" is very different from
synteny seen between other organisms. A surprising feature of the M.
graminicola genome compared to other sequenced plant pathogens was that it
contained very few genes for enzymes that break down plant cell walls,
which was more similar to endophytes than to pathogens. The stealth
pathogenesis of M. graminicola probably involves degradation of proteins
rather than carbohydrates to evade host defenses during the biotrophic
stage of infection and may have evolved from endophytic ancestors.

<>

<1>Goossens, M.D., Dumez, Y., Kaplan, L., Lupker, M., Chabret, C., Henrion, R., Rosa, J.
<2>Prenatal Diagnosis of Sickle-Cell anemia in the First Trimester of Pregnancy.
<3>N. Engl. J. Med.
<4>309
<5>831-833
<6>1983
<7>To investigate the usefulness of chorionic biopsy for prenatal diagnosis of
sickle-cell anemia by restriction-endonuclease analysis of fetal DNA, we
studies 30 pregnancies before elective abortion.  When the reproductibility of
the technique for obtaining adequate DNA samples was established, we
successfully applied the test to five pregnancies at risk for sickle-cell
anemia.  In two cases, sickle-cell disease of the fetus led to a decision to
terminate the pregnancy.  In three other cases, a normal or AS genotype was
demonstrated.  One normal infant has been born, and one other pregnancy is
continuing normally.  In one case in which fetal death was observed three weeks
after ampling, placental abnormalties found on histologic examination were
compatible with a chromosomal aberration.  Our study shows that chorionic
biopsy is feasible for the prenatal diagnosis of sickle-cell disease before the
10th gestational week.  If subsequent experience demonstrates this technique to
be safe enough for mother and fetus, the ability to test in early pregnancy may
make prenatal diagnosis acceptable to more couples at risk for serious genetic
disorders.

<>

<1>Gootz, T.D., Lescoe, M.K., Dib-Hajj, F., Dougherty, B.A., He, W., Della-Latta, P., Huard, R.C.
<2>Genetic organization of transposase regions surrounding blaKPC carbapenemase genes on plasmids from Klebsiella strains isolated in a New York City hospital.
<3>Antimicrob. Agents Chemother.
<4>53
<5>1998-2004
<6>2009
<7>Carbapenem-resistant Klebsiella strains carrying Klebsiella pneumoniae
carbapenemases (KPC) are endemic to New York City and are spreading across
the United States and internationally. Recent studies have indicated that
the KPC structural gene is located on a 10-kb plasmid-borne element
designated Tn4401. Fourteen Klebsiella pneumoniae strains and one
Klebsiella oxytoca strain isolated at a New York City hospital in 2005
carrying either bla(KPC-2) or bla(KPC-3) were examined for isoforms of
Tn4401. Ten of the Klebsiella strains contained a 100-bp deletion in
Tn4401, corresponding to the Tn4401a isoform. The presence of this
deletion adjacent to the upstream promoter region of bla(KPC) in Tn4401a
resulted in a different -35 promoter sequence of TGGAGA than that of
CTGATT present in isoform Tn4401b. Complete sequencing of one plasmid
carrying bla(KPC) from each of three nonclonal isolates indicated the
presence of genes encoding other types of antibiotic resistance
determinants. The 70.6-kb plasmid from K. pneumoniae strain S9 carrying
bla(KPC-2) revealed two identical copies of Tn4401b inserted in an inverse
fashion, but in this case, one of the elements disrupted a group II
self-splicing intron. In K. pneumoniae strain S15, the Tn4401a element
carrying bla(KPC-2) was found on both a large 120-kb plasmid and a smaller
24-kb plasmid. Pulsed-field gel electrophoresis results indicate that the
isolates studied represent a heterogeneous group composed of unrelated as
well as closely related Klebsiella strains. Our results suggest that
endemic KPC-positive Klebsiella strains constitute a generally nonclonal
population comprised of various alleles of bla(KPC) on several distinct
plasmid genetic backgrounds. This study increases our understanding of the
genetic composition of the evolving and expanding role of KPC-producing,
healthcare-associated, gram-negative pathogens.

<>

<1>Gopal, J.
<2>Molecular cloning and biochemical characterization of DsaV methyltransferase from Dactylococcopsis salina.
<3>Diss. Abstr.
<4>57
<5>121
<6>1995
<7>The methyltransferase (Mtase) in the DsaV restriction-modification system methylates within
5'-CCNGG sequences.  I have cloned the gene for this Mtase and characterized it.  The
predicted sequence of the Mtase protein contains sequence motifs conserved among all
cytosine-5 Mtases and is most similar to other Mtases that methylate CCNGG sequences -- namely
M.ScrFI and M.SsoII.  The "variable" region within the three enzymes that methylate CCNGG can
be aligned with the sequences of two enzymes that methylate CCWGG sequences.  Remarkably, two
segments within this region contain significant similarity with the region of M.HhaI that is
known to contact DNA bases.  These aligments suggest that many cytosine-5 Mtases are likely to
interact with DNA using a similar structural framework.  I have developed a simple new method
that can identify the base methylated by a sequence-specific DNA methyltransferase and have
used it to identify the cytosine that is methylated by DsaV methyltransferase (M.DsaV) within
its recognition sequence 5'-CCNGG.  The method utilizes the fact that exonuclease III of E.
coli does not degrade DNA ends with 3' overhangs and cannot hydrolyze a phosphorothioate
linkage.  DNA duplexes containing phosphorothioate linkages at specific positions were
methylated with M.DsaV in the presence of [methyl-3H] S-adenosylmethionine and were subjected
to exonuclease III digestion.  The pattern of [methyl-3H] dCMP release from the duplexes was
consistent with the methylation of the internal cytosine in CCNGG, but not of the outer
cytosine.  Methylation of CCWGG (W = A or T) sequences at the internal cytosines is native to
E. coli and is not restricted by the modified cytosine restriction (Mcr) systems.
Surprisingly, the gene for M.DsaV was significantly restricted by the McrBC system.  I
interpret this to mean that M.DsaV may occasionally methylate at sequences other than CCNNGG
or may occasionally methylate the outer cytosine in its recognition sequence.  I have also
shown for the first time that M.EcoRII methylates at CCSGG sites and other non-canonical sites
using a plasmid that substantially overproduces M.EcoRII.  This result suggests that C5
methylates may occasionally methylate cellular DNA at non-canonical sites and that E. coli
methylation specific restriction system and DNA mismatch correction systems may have evolved
to accommodate this fact.

<>

<1>Gopal, J., Bhagwat, A.S.
<2>Determination of methylation specificity of DsaV methyltransferase by a simple biochemical method.
<3>Nucleic Acids Res.
<4>23
<5>29-35
<6>1995
<7>We have developed a simple new method that can identify the base methylated by a
sequence-specific DNA methyltransferase and have used it to identify the cytosine that is
methylated by DsaV methyltransferase (M.DsaV) within its recognition sequence 5'-CCNGG. The
method utilizes the fact that exonuclease III of E.coli does not degrade DNA ends with 3'
overhangs and cannot hydrolyze a phosphorothioate linkage. DNA duplexes containing
phosphorothioate linkages at specific positions were methylated with M.DsaV in the presence of
[methyl-3H] S-adenosylmethionine and were subjected to exonuclease III digestion. The pattern
of [methyl-3H] dCMP release from the duplexes was consistent with the methylation of the
internal cytosine in CCNGG, but not of the outer cytosine. To establish the accuracy of this
method, we confirmed the known specificity of EcoRII methyltransferase by the method. We also
confirmed the specificity of M.DsaV using an established biochemical method that involves the
use of a type IIS restriction enzyme. Methylation of CCWGG (W=A or T) sequences at the
internal cytosines is native to E. coli and is not restricted by the modified cytosine
restriction (Mcr) systems. Surprisingly, the gene for M.DsaV was significantly restricted by
the McrBC system. We interpret this to mean that M.DsaV may occasionally methylate at
sequences other than CCNGG or may occasionally methylate the outer cytosine in its recognition
sequence.

<>

<1>Gopal, J., Yebra, M.J., Bhagwat, A.S.
<2>DsaV methyltransferase and its isoschizomers contain a conserved segment that is similar to the segment in HhaI methyltransferase that is in contact with DNA bases.
<3>Nucleic Acids Res.
<4>22
<5>4482-4488
<6>1994
<7>The methyltransferase (MTase) in the DsaV restriction-modification system methylates within
5'-CCNGG sequences. We have cloned the gene for this MTase and determined its sequence. The
predicted sequence of the MTase protein contains sequence motifs conserved among all
cytosine-5 MTases and is most similar to other MTases that methylate CCNGG sequences, namely
M.ScrFI and M.SsoII. All three MTases methylate the internal cytosine within their recognition
sequence. The 'variable' region within the three enzymes that methylate CCNGG can be aligned
with the sequences of two enzymes that methylate CCWGG sequences. Remarkably, two segments
within this region contain significant similarity with the region of M.HhaI that is known to
contact DNA bases. These alignments suggest that many cytosine-5 MTases are likely to interact
with DNA using a similar structural framework.

<>

<1>Gopal, J., Yebra, M.J., Bhagwat, A.S.
<2>Cloning and characterization of the gene encoding the DsaV methyltransferase.
<3>Gene
<4>157
<5>61-63
<6>1995
<7>A gene encoding the M.DsaV methyltransferase was cloned and characterized.  The enzyme
methylates the internal cytosines in the 5'-CCTGG recognition sequence, as determined by a
novel rapid method employing 3H label and exonuclease III.

<>

<1>Gopalakrishnan, S.
<2>Functional characterization of the de novo DNA methyltransferase DNMT3B.
<3>Ph.D. Thesis, Univ. of Florida, Gainesville
<4>
<5>
<6>2009
<7>DNA methylation is an epigenetic mark that is required for transcriptional repression in
mammalian development, imprinting, and in the maintenance of genome stability. Genome-wide
methylation patterns are established and maintained by three DNA methyltransferases (DNMTs)-
DNMT1, DNMT3A, and DNMT3B. DNMT3B is specifically involved in silencing the satellite repeats
at the centromeric and pericentromeric regions. The role of DNMT3B at the centromeric region
is also emphasized by mitotic defects arising due to chromosome instability observed in ICF
syndrome, a disease caused by germline mutations in DNMT3B. Although the mechanism of DNA
methylation is well known, the targeting of DNA methylation and DNMT3B to certain genomic loci
remains poorly understood. DNMT3B is also regulated by alternative splicing. Several DNMT3B
splice variants are overexpressed in tumor cells and negatively regulate normal DNMT3B
mediated DNA methylation. Therefore it is important to understand the significance of DNMT3B
splice variants in development and tumorigenesis. In the present study, a yeast two-hybrid
screening was performed and several novel DNMT3B protein interactions were identified. Of the
several proteins identified, the interaction
of DNMT3B with the mammalian chromatin associated factor MCAF and the chromodomain helicase
DNA binding protein CHD3 were confirmed, and need further characterization. The interaction
between DNMT3B and the constitutive centromeric protein CENP-C was confirmed in mammalian
cells. Results from siRNA knock downs, bisulfite genomic sequencing and ChIP,
demonstrate that CENP-C recruits DNA methylation and DNMT3B to both centromeric and
pericentromeric satellite repeats. CENP-C and DNMT3B influence the histone modifications in
satellite repeat regions, including marks characteristic of centromeric chromatin and
disruption of this interaction causes elevated transcription of centromeric repeats. Loss of
CENP-C or DNMT3B leads to elevated chromosome misalignment and segregation defects during
mitosis. Taken together, the interaction between CENP-C and DNMT3B suggests a novel mechanism
by which DNA methylation is targeted to discrete regions of the genome and contributes to
chromosomal stability.
In another study, a novel alternatively spliced form of DNMT3B lacking exon 5 was identified
and characterized. This variant was termed DNMT3B3"5 because of its close resemblance with the
ubiquitously expressed DNMT3B3 isoform. The novel splice variant lacking exon 5 is highly
expressed in pluripotent cells and neural tissues, and is conserved in the mouse and is
re-expressed on converting differentiated mEFs into pluripotent iPS cells.
DNMT3B3"5 also displays altered expression in human tumor cell lines as well as an altered
subcellular localization. Ectopic overexpression of DNMT3B3"5 resulted in repetitive element
hypomethylation. Taken together, these results demonstrate that alternative splicing of exon 5
may play an important role in stem cell maintenance or differentiation and exon 5 could
influence the functional properties of DNMT3B.

<>

<1>Gopinath, G., Jean-Gilles, B.J., Grim, C., Blaylock, M., Blackwell, R., Merid, S., Diallo, A., Hanes, D.
<2>Whole-Genome Sequences of Six Salmonella enterica Serovar Bovismorbificans Isolates Associated with a 2011 Multistate Hummus-Borne Outbreak.
<3>Genome Announcements
<4>2
<5>e01239-13
<6>2014
<7>We present six draft genome sequences of Salmonella enterica serovar Bovismorbificans from
isolates associated with the 2011 hummus-borne multistate
outbreak. All six genome sequences indicate the presence of two plasmids, one of
which demonstrates similarity to the 93-kb pSLT2 IncF-type plasmid of Salmonella
enterica serovar Typhimurium.

<>

<1>Gopinath, G., Jean-Gilles, B.J., Grim, C., Hanes, D.
<2>Draft Genome Sequences of Nine Salmonella enterica Serovar Bovismorbificans Isolates from Various Sources.
<3>Genome Announcements
<4>2
<5>e01249-13
<6>2014
<7>The sequences of nine genomes of Salmonella enterica serovar Bovismorbificans were compared to
study the diversity and distribution of this emerging virulent
serovar. These whole-genome sequences fill some gaps in knowledge of the
diversity of the isolates used in this investigation.

<>

<1>Gopinath, G.R., Grim, C.J., Tall, B.D., Mammel, M.K., Sathyamoorthy, V., Trach, L.H., Chase, H.R., Fanning, S., Stephan, R.
<2>Genome Sequences of Two Enterobacter pulveris Strains, 601/05T (=LMG 24057T =DSM  19144T) and 1160/04 (=LMG 24058 =DSM 19146), Isolated from Fruit Powder.
<3>Genome Announcements
<4>1
<5>e00991-13
<6>2013
<7>We report the draft genome sequences of the Enterobacter pulveris strains 601/05(T)
(=LMG24057(T) =DSM19144(T)) and 1160/04 (=LMG24058 =DSM19146), isolated
from fruit powder. The genome assemblies for the E. pulveris type strain,
LMG24057, and strain LMG24058 have sizes of 4,708,624 and 4,811,103 bp and G+C
contents of 56.6% and 56.5%, respectively.

<>

<1>Goppelt, M., Langowski, J., Pingoud, A., Haupt, W., Urbanke, C., Mayer, H.
<2>The effect of several nucleic acid binding drugs on the cleavage of d(GGAATTCC) and pBR322 by the EcoRI restriction endonuclease.
<3>Nucleic Acids Res.
<4>9
<5>6115-6127
<6>1981
<7>The endonucleolytic action of the EcoRI restriction enzyme on the
double-stranded oligonucleotide d(GGAATTCC) and the supercoiled plasmid DNA
pBR322 is inhibited by actinomycin D, ethidium bromide, proflavin, distamycin A
and netropsin.  Half-maximal inhibition is observed at around 100 micromolar
concentrations for the intercalating drugs, and around 0.1 to 1 micromolar
concentrations for netropsin and distamycin A.  The inhibitory activity of
these drugs can be correlated with their affinity to the oligonucleotide and
the plasmid DNA.  Since at high concentrations of the drugs a complete
inhibition is observed, it is concluded that the effect of the drugs on the
sterochemistry of the EcoRI site is such that recognition is excluded.

<>

<1>Goppelt, M., Pingoud, A., Maass, G., Mayer, H., Koster, H., Frank, R.
<2>The interaction of the EcoRI restriction endonuclease with its substrate:  A physico-chemical study employing natural and synthetic oligonucleotides and polynucleotides.
<3>Eur. J. Biochem.
<4>104
<5>101-107
<6>1980
<7>The interaction of the EcoRI restriction endodeoxyribonuclease with
polynucleotides has been studied in a qualitative manner by an affinity
adsorption technique using polynucleotides immobilized on cellulose.  It is
shown that EcoRI binds to single-stranded and double-stranded
polyribonucleotides and polydeoxyribonucleotides.  Mg2+ ions are not required
for binding.  In order to define differences between specific and non-specific
binding we have investigated the interaction of EcoRI with d(TAAATG),
d(TTACAT), d(GAATTC) and d(GGAATTCC).  We have synthesized for this purpose
d(TAAATG), d(TTACAT) and d(GAATTC) by the diester approach.  d(GAATTC) and
d(GGAATTCC) are self complementary.  Differential melting experiments show that
the octanucleotide has a melting point of 28C under ionic conditions where the
hexanucleotide is single-stranded even below 0C.  Correspondingly, the
octanucleotide is cleaved by EcoRI, while the hexanucleotide is not.  The
binding of oligonucleotides to EcoRI can be monitored by the circular dichroism
of the enzyme.  Titrations show that in the absence of Mg2+ ions all
oligonucleotides are bound with similar magnitude:  Ka - 10/7 M-1.  Complex
formation is weakened with increasing temperature, corresponding to a delta Ho
of -21 kJ/mol and increasing ionic strength, corresponding to an involvement of
two ion-pair bonds between DNA and enzyme.  Mg2+ ions have no significant
influence on the binding of d(TAAATG), d(TTACAT) and d(GAATTC) to the enzyme.
The binding of d(GGAATTCC) to EcoRI is strengthened by a factor of 50 in the
presence of Mg2+ ions, as measured by cleavage experiments using d(GAATTC) as a
competing inhibitor in the enzymatic assay.

<>

<1>Gorbalenya, A.E.
<2>Self-splicing group I and group II introns encode homologous (putative) DNA endonucleases of a new family.
<3>Protein Sci.
<4>3
<5>1117-1120
<6>1994
<7>A new family of protein domains consisting of 50-80 amino acid residues is described.  It is
composed of nearly 40 members, including domains encoded by plastid and phage group I introns;
mitochondrial, plastid, and bacterial group II introns; eubacterial genomes and plasmids; and
phages.  The name "EX1HH-HX3H" was coined for both domain and family.  It is based on 2 most
prominent amino acid sequene motifs, each encompassing a pair of highly conserved histidine
residues in a specific arrangement: EX1HH and HX3H.  The "His" motifs often alternate with
amino- and carboxy-terminal motifs of a new type of Zn-finger-like structure
CX2,4CX29-54[CH]X2,3[CH].  The EX1HH-HX3H domain in eubacterial E2-type bacteriocins and in
phage RB3 (wild variant of phage T4) product of the nrdB group I intron was reported to be
essential for DNA endonuclease activity of these proteins.  In other proteins, the EX1HH-HX3H
domain is hypothesized to possess DNase activity as well.  Presumably, this activity promotes
movement (rearrangement) of group I and group II introns encoding the EX1HH-HX3H domain and
other gene targets.  In the case of Escherichia coli restrictase McrA and possibly several
related proteins, it appears to mediate the restriction of alien DNA molecules.

<>

<1>Gorbalenya, A.E., Koonin, E.V.
<2>Endonuclease (R) subunits of type-I and type-III restriction-modification enzymes contain a helicase-like domain.
<3>FEBS Lett.
<4>291
<5>277-281
<6>1991
<7>A statistically significant amino acid sequence similarity is demonstrated
between the endonuclease (R) subunit of the EcoKI restriction-modification
(R-M) enzyme, and RNA and DNA helicases of the so-called 'DEAD' family.  It is
further shown that all three known sequences of R subunits of type-I and
type-III R-M enzymes contain the conserved amino acid sequence motifs typical
of the previously described helicase superfamily II (1989) Nucl. Acids Res.
17,4713-4730.  A hypothesis is proposed that these enzymes may exert helicase
activity possibly required for local unwinding of DNA in the cleavage sites.

<>

<1>Gorbunov, Y.A., Zinovev, V.V., Rechkunova, N.I., Ovechkina, L.G., Popov, S.G., Malygin, E.G.
<2>Chemical synthesis and characteristics of oligonucleotide substrates for restriction endonuclease BamHI and methylase Eco dam.
<3>Bioorg. Khim.
<4>13
<5>1629-1637
<6>1987
<7>Oligodeoxyribonucleotides which form a number of duplexes, containing the recognition
sequences for endonuclease BamHI and DNA methylase Eco dam, were synthesized by the
phosphotriester approach. Furthermore, synthesis of 3'-phosphorylated
oligodeoxyribonucleotides from corresponding S-methyl phosphorothioate triester oligomers is
described. The synthetic duplexes are characterized by some defects in the recognition
sequences for endonuclease BamHI and methylase Eco dam, viz. nick, absence of an
internucleotide phosphate, modifications (including partial single-strandedness) of the
recognition site. Interaction of the enzymes with these synthetic substrates was investigated.

<>

<1>Gordon, S.V., Brosch, R., Billault, A., Garnier, T., Eiglmeier, K., Cole, S.T.
<2>Identification of variable regions in the genomes of tubercle bacilli using bacterial artificial chromosome arrays.
<3>Mol. Microbiol.
<4>32
<5>643-655
<6>1999
<7>Whole-genome comparisons of the tubercle bacilli were undertaken using ordered bacterial
artificial chromosome (BAC) libraries of Mycobacterium
tuberculosis and the vaccine strain, Mycobacterium bovis BCG-Pasteur,
together with the complete genome sequence of M. tuberculosis H37Rv.
Restriction-digested BAC arrays of M. tuberculosis H37Rv were used in
hybridization experiments with radiolabelled M. bovis BCG genomic DNA to
reveal the presence of 10 deletions (RD1-RD10) relative to M.
tuberculosis. Seven of these regions, RD4-RD10, were also found to be
deleted from M. bovis, with the three M. bovis BCG-specific deletions
being identical to the RD1-RD3 loci described previously. The distribution
of RD4-RD10 in Mycobacterium africanum resembles that of M. tuberculosis
more closely than that of M. bovis, whereas an intermediate arrangement
was found in Mycobacterium microti, suggesting that the corresponding
genes may affect host range and virulence of the various tubercle bacilli.
Among the known products encoded by these loci are a copy of the proposed
mycobacterial invasin Mce, three phospholipases, several PE, PPE and
ESAT-6 proteins, epoxide hydrolase and an insertion sequence. In a
complementary approach, direct comparison of BACs uncovered a third class
of deletions consisting of two M. tuberculosis H37Rv loci, RvD1 and RvD2,
deleted from the genome relative to M. bovis BCG and M. bovis. These
deletions affect a further seven genes, including a fourth phospholipase,
plcD. In summary, the insertions and deletions described here have
important implications for our understanding of the evolution of the
tubercle complex.

<>

<1>Gorecki, R.K., Bardowski, J.K.
<2>Molecular mechanisms of bacteriophage resistance of lactic acid bacteria.
<3>Postepy Mikrobiologii
<4>50
<5>265-273
<6>2011
<7>Lactic acid bacteria (LAB) constitute a heterogeneous group of bacteria, which are found in
diverse environments, such as the human body or plants, and are traditionally used to produce
fermented food. Food bio-transformation in industrial processes increases the economical
importance of LAB. However, conditions that exist in industrial facilities do not seem to be
an optimal environment for bacteria. During technological processes, which take place in
enclosed space, the intensity of physical (temperature shift), chemical (acids) or biological
(phages) stress factors raises dramatically. In the dairy industry, bacteriophage
contamination is regarded as a serious problem due to the disturbance or arrest of the
production processes, which results in significant economical losses. It is well documented
that LAB evolved defense systems against bacteriophages, which allow them to survive in harsh
conditions. Therefore, bacteria used in food industry are selected for high level of
bacteriophage resistance. According to the mode of action, natural bacterial defense systems
against their predators were divided into 5 categories: (i) inhibition of phage adsorption,
(ii) blocking of phage DNA injection, (iii) phage abortive infection systems, (iv) restriction
modification systems, (v) CRISPR/Cas systems. Remarkably, the majority of known bacteriophage
resistance systems are plasmid-encoded. In this context, future studies on phage resistance
mechanisms as well as plasmid sequencing may have an impact on solving the problem of phage
infections in the dairy industry.

<>

<1>Gorecki, R.K., Koryszewska-Baginska, A., Golebiewski, M., Zylinska, J., Grynberg, M., Bardowski, J.K.
<2>Adaptative potential of the Lactococcus lactis IL594 strain encoded in its 7 plasmids.
<3>PLoS ONE
<4>6
<5>e22238
<6>2011
<7>The extrachromosomal gene pool plays a significant role both in evolution and in the
environmental adaptation of bacteria.  The L. lactis subsp. lactis IL594 strain contains seven
plasmids, named pIL1 to pIL7, and is the parental strain of the plasmidfree L. lactis IL1403,
which is one of the best characterized lactococcal strains of LAB. Complete nucleotide
sequences of pIL1 (6,382 bp), pIL2 (8,277 bp), pIL3 (19,244 bp), pIL4 (48,979), pIL5 (23,395),
pIL6 (28,435 bp) and pIL7 (28,546) were
established and deposited in the generally accessible database (GeneBank). Nine highly
homologous repB-containing replicons, belonging to the lactococcal theta-type replicons, have
been identified on the seven plasmids. Moreover, a putative region involved in conjugative
plasmid mobilization was found on four plasmids, through identification of the
presence of mob genes and/or oriT sequences. Detailed bioinformatic analysis of the plasmid
nucleotide sequences provided new insight into the repertoire of plasmid-encoded functions in
L. lactis, and indicated that plasmid genes from IL594 strain can be important for L. lactis
adaptation to specific environmental conditions (e.g. genes coding for proteins involved in
DNA repair or cold shock response) as well as for technological processes (e.g. genes encoding
citrate and lactose utilization, oligopeptide transport, restriction-modification system).
Moreover, global gene analysis indicated
cooperation between plasmid- and chromosome-encoded metabolic pathways.

<>

<1>Gorkiewicz, G., Kienesberger, S., Schober, C., Scheicher, S.R., Gully, C., Zechner, R., Zechner, E.L.
<2>A genomic island defines subspecies-specific virulence features of the host-adapted pathogen Campylobacter fetus subsp. venerealis.
<3>J. Bacteriol.
<4>192
<5>502-517
<6>2010
<7>The pathogen Campylobacter fetus comprises two subspecies, C. fetus subsp.
fetus and C. fetus subsp. venerealis. Although these taxa are highly
related on the genome level, they are adapted to distinct hosts and
tissues. C. fetus subsp. fetus infects a diversity of hosts, including
humans, and colonizes the gastrointestinal tract. In contrast, C. fetus
subsp. venerealis is largely restricted to the bovine genital tract,
causing epidemic abortion in these animals. In light of their close
genetic relatedness, the specific niche preferences make the C. fetus
subspecies an ideal model system to investigate the molecular basis of
host adaptation. In this study, a subtractive-hybridization approach was
applied to the genomes of the subspecies to identify different genes
potentially underlying this specificity. The comparison revealed a genomic
island uniquely present in C. fetus subsp. venerealis that harbors several
genes indicative of horizontal transfer and that encodes the core
components necessary for bacterial type IV secretion. Macromolecular
transporters of this type deliver effector molecules to host cells,
thereby contributing to virulence in various pathogens. Mutational
inactivation of the putative secretion system confirmed its involvement in
the pathogenicity of C. fetus subsp. venerealis.

<>

<1>Gorlas, A., Gimenez, G., Raoult, D., Roux, V.
<2>Draft Genome Sequences of Actinomyces timonensis Strain 7400942T and Its Prophage.
<3>J. Bacteriol.
<4>194
<5>6613-6614
<6>2012
<7>A draft genome sequence of Actinomyces timonensis, an anaerobic bacterium isolated from a
human clinical osteoarticular sample, is described here.
CRISPR-associated proteins, insertion sequence, and toxin-antitoxin loci were
found on the genome. A new virus or provirus, AT-1, was characterized.

<>

<1>Gorlas, A., Robert, C., Gimenez, G., Drancourt, M., Raoult, D.
<2>Complete Genome Sequence of Methanomassiliicoccus luminyensis, the Largest Genome of a Human-Associated Archaea Species.
<3>J. Bacteriol.
<4>194
<5>4745
<6>2012
<7>The present study describes the complete and annotated genome sequence of
Methanomassiliicoccus luminyensis strain B10 (DSM 24529(T), CSUR P135), which was
isolated from human feces. The 2.6-Mb genome represents the largest genome of a
methanogenic euryarchaeon isolated from humans. The genome data of M. luminyensis
reveal unique features and horizontal gene transfer events, which might have
occurred during its adaptation and/or evolution in the human ecosystem.

<>

<1>Gorlatova, N.V., Kryuchkova, E.G., Koltovaya, N.A., Dolgova, I.N., Ananyin, V.M., Velkov, V.V.
<2>Synthesis of EcoRV restriction endonuclease in Escherichia coli continuous culture.  II. Effects of cultivation conditions on efficiency of thermoinduction of synthesis of cloned EcoRV restriction endonuclease.
<3>Biotekhnologiya
<4>0
<5>27-30
<6>1991
<7>Effects of growth rate of recombinant cells of E. coli K802 (plLRV8) and of induction
conditions on the efficiency of thermoinduction of synthesis of EcoRV restriction endonuclease
cloned in PILRV8 plasmid controlled by the pR promoter of lambda phage were studied. The
efficiency of thermoinduction was shown to be 2-2.5 times higher in E. coli cells growing with
mu=0.35 h-1 than in fast growing cells with mu=0.6 h-1. Introduction of casamino acids did not
influence the thermoinduction level in slow (mu=0.35 h-1) growing cells, but stimulated
restrictase synthesis insignificantly in fast growing cells (mu=0.6 h-1). Addition of glucose
and glucose + casamino acids into EcoRV induction medium resulted in lowering the
thermoinduction efficiency of the PR promoter by 13-31%, respectively, as compared with
induction on nutrient-deficient medium for slow growing cells. At the same time, addition of
glucose while thermoinducing fast growing cells enhanced synthesis of EcoRV restrictase.
Dynamics of the induced synthesis of EcoRV restriction endonuclease were observed to be
different in fast and slow growing cultures of recombinant cells.

<>

<1>Gorlatova, N.V., Kryukova, E.G., Koltovaya, N.A., Dolgova, I.N., Velkov, V.V.
<2>Synthesis of EcoRV restriction endonuclease by chemostatic cultivation of an Escherichia coli K 802 recombinant strain.  1. Effects of growth rate on expression of plasmid genes of EcoRV restriction endonuclease and beta-lactamase.
<3>Biotekhnologiya
<4>3
<5>34-38
<6>1991
<7>Stability and expression of pILRV8 plasmid in E. coli cells growing on minimal
glucose medium were studied in chemostatic mode within a wide range of dilution
rates (D=0; V=0.1-0.6 hr-1) and in turbidostatic mode (Vmax=0.7 hr-1).  High
segregational and structural stability was characteristic for the strain under
these conditions.  Beta-lactamase and EcoRV restriction endonuclease determined
by genes of pILRV8 plasmid were synthesized concertedly, activity increasing
4-fold with increasing dilution rate in chemostatic mode.

<>

<1>Gormley, N.A., Bath, A.J., Halford, S.E.
<2>Reactions of BglI and other Type II restriction endonucleases with discontinuous recognition sites.
<3>J. Biol. Chem.
<4>275
<5>6928-6936
<6>2000
<7>Type II restriction enzymes generally recognize continuous sequences of 4-8 consecutive base
pairs on DNA, but some recognize discontinuous
sites where the specified sequence is interrupted by a defined length
of nonspecific DNA. To date, a mechanism has been established for only
one type II endonuclease with a discontinuous site, SfiI at
GGCCNNNNNGGCC (where N is any base). In contrast to orthodox enzymes
such as EcoRV, dimeric proteins that act at a single site, SfiI is a
tetramer that interacts with two sites before cleaving DNA. BglI has a
similar recognition sequence (GCCNNNNNGGC) to SfiI but a crystal
structure like EcoRV. BglI and several other endonucleases with
discontinuous sites were examined to see if they need two sites for
their DNA cleavage reactions. The enzymes included some with sites
containing lengthy segments of nonspecific DNA, such as XcmI
(CCANNNNNNNNNTGG). In all cases, they acted at individual sites.
Elongated recognition sites do not necessitate unusual reaction
mechanisms. Other experiments on BglI showed that it bound to and
cleaved DNA in the same manner as EcoRV, thus further delineating a
distinct group of restriction enzymes with similar structures and a
common reaction mechanism.

<>

<1>Gormley, N.A., Hillberg, A.L., Halford, S.E.
<2>The type IIs restriction endonuclease BspMI is a tetramer that acts concertedly at two copies of an asymmetric DNA sequence.
<3>J. Biol. Chem.
<4>277
<5>4034-4041
<6>2002
<7>Type IIs endonucleases recognize asymmetric DNA sequences and cleave both strands at fixed
positions downstream of the sequence. Many type IIs enzymes, including BspMI, cleave
substrates with two sites more rapidly than those with one site. They usually act sequentially
on DNA with two sites, but BspMI converted such a substrate directly to the final products cut
at both sites. The BspMI endonuclease was found to be a tetramer, in contrast to the monomeric
structures for many type IIs enzymes. No change in subunit association occurred during the
BspMI reaction. Plasmids with two BspMI sites were cleaved in cis, in reactions spanning sites
in the same DNA, even when the sites were separated by just 38 bp. Plasmids with one BspMI
site were cleaved in trans, with the enzyme bridging sites in separate DNA molecules: these
slow reactions could be accelerated by adding a second DNA with the recognition sequence.
Thus, whereas many type IIs enzymes dimerize before cleaving DNA, a process facilitated by two
recognition sites in cis, the BspMI tetramer binds two copies of its recognition sequence
before cleaving the DNA in both strands at both sites.

<>

<1>Gorovsky, M.A., Hattman, S., Pleger, G.L.
<2>[6N]Methyl adenine in the nuclear DNA of a eucaryote, Tetrahymena pyriformis.
<3>J. Cell Biol.
<4>56
<5>697-701
<6>1973
<7>DNA isolated from macronuclei of the ciliate, Tetrahymena pyriformis, has been found to
contain [6N]methyl adenine (MeAde); this represents the first clear demonstration of
significant amounts of MeAde in the DNA of a eucaryote. The amounts of macronuclear MeAde
differed slightly between different strains of Tetrahymena, with approximately 0.65-0.80% of
the adenine bases being methylated. The MeAde content of macronuclear DNA did not seem to vary
in different physiological states. The level of MeAde in DNA isolated from micronuclei, on the
other hand, was quite low (at least tenfold lower than in macronuclear DNA).

<>

<1>Goto, T., Ogura, Y., Hirakawa, H., Tomida, J., Morita, Y., Akaike, T., Hayashi, T., Kawamura, Y.
<2>Complete Genome Sequence of Helicobacter cinaedi Strain PAGU611, Isolated in a Case of Human Bacteremia.
<3>J. Bacteriol.
<4>194
<5>3744-3745
<6>2012
<7>We report the complete genome sequence of Helicobacter cinaedi strain PAGU611, isolated in a
case of human bacteremia. The PAGU611 genome comprises a
2,078,348-bp chromosome and a 23,054-bp plasmid. The chromosome contains a unique
genomic island, encoding a type VI secretion system and clustered regularly
interspaced short palindromic repeat (CRISPR) loci.

<>

<1>Gott, J.M.
<2>Genes within genes: independent expression of phage T4 intron open reading frames and the genes in which they reside.
<3>Genes Dev.
<4>2
<5>1791-1799
<6>1988
<7>The td, nrdB, and sunY introns of bacteriophage T4 each contain a long open reading frame
(ORF). These ORFs are preceded by functional T4 late promoters and, in the case of the nrdB
intron ORF, a functional middle promoter. Expression of phage-encoded intron ORF-lacZ fusions
indicates that these T4 genes are highly regulated. The lack of translation of these ORFs from
the early pre-mRAs can be accounted for by the presence of secondary structures that are
absent from the late RNAs. Because translation of the intron ORFs could disrupt core
structural elements required for pre-mRNA splicing, such regulation may be necessary to allow
expression of the genes in which they reside.

<>

<1>Gottschling, D.E.
<2>Telomere-proximal DNA in Saccharomyces cerevisiae is refractory to methyltransferase activity in vivo.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>4062-4065
<6>1992
<7>Genes located near telomeres in Saccharomyces cerevisiae undergo position-effect variegation;
their transcription is subject to reversible but mitotically heritable repression. This
position effect and the finding that telomeric DNA is late replicating suggest that yeast
telomeres exist in a heterochromatin-like state. Mutations in genes that suppress the
telomeric position effect suggest that a special chromatin structure exists near chromosomal
termini. Thus transcriptional repression may be explained by the inability of DNA binding
proteins to access the DNA near telomeres. To test this hypothesis, the Escherichia coli Dam
DNA methyltransferase, which modifies the sequence GATC, was introduced into S. cerevisiae
cells. DNA sequences near the telomere were highly refractive to Dam methylation but were
modified when located at positions more internal on the chromosome. Telomeric sequences were
accessible to methyltransferase activity in strains that contained a mutation that suppressed
the telomeric position effect. These data support the model that sequence-specific DNA binding
proteins are excluded from telomere-proximal sequences in vivo and show that expression of DNA
methyltransferase activity may serve as a useful tool for mapping chromosomal structural
domains in vivo.

<>

<1>Gotz, F., Seeber, S.
<2>Recombinant restriction enzyme Sau3AI.
<3>European Patent Office
<4>EP 460673 A
<5>
<6>1990
<7>New DNA (I) contains at least one of the coding regions for a protein (P1) with Sau3AI
methylase activity or for a protein (P2) with Sau3AI endonuclease activity, these coding
regions being present in a specified 3360 bp sequence reproduced in the specification. Also
included are analogues of (I) within the degeneracy of the genetic code or sequences able to
hybridise with such DNA under stringent conditions. Also new are (1) recombinant vectors
containing one or more copies of (I); (2) microorganisms transformed with such vectors, and
(3) recombinant proteins with Sau3AI-endonuclease activity produced by the microorganisms.
More specifically, (I) may have, inside and/or outside the P1 coding region, at least one
mutation to reduce total methylase activity when expressed in cells. In particular, a stop
codon is introduced within the P1 coding region, esp. CGA (2134-2136) is replaced by TGA.

<>

<1>Gotz, F., Seeber, S.
<2>Recombinant restriction enzyme Sau3AI.
<3>US Patent Office
<4>US 5175101
<5>
<6>1992
<7>The invention concerns a DNA which contains (1) at least one of the two regions of the
nucleotide sequence SEQ ID No:1 coding for a protein with a Sau3AI methylase activity or for a
protein with a Sau3AI endonuclease activity. (2) a sequence corresponding to the DNA from (1)
within the scope of the degeneration of the genetic code or (3) a sequence which hybridizes
under stringent hybridization conditions with a DNA from (1) or (2).  In addition a vector is
disclosed which contains a DNA according to the present invention, in particular under the
control of a regulatable closed promoter, as well as microorganisms which are transformed with
one or several vectors according to the present invention and the isolation of recombinant
Sau3AI endonuclease from the transformed microorganisms.

<>

<1>Goudenege, D., Labreuche, Y., Krin, E., Ansquer, D., Mangenot, S., Calteau, A., Medigue, C., Mazel, D., Polz, M.F., Le Roux, F.
<2>Comparative genomics of pathogenic lineages of Vibrio nigripulchritudo identifies virulence-associated traits.
<3>ISME J.
<4>7
<5>1985-1996
<6>2013
<7>Vibrio nigripulchritudo is an emerging pathogen of farmed shrimp in New Caledonia
and other regions in the Indo-Pacific. The molecular determinants of V.
nigripulchritudo pathogenicity are unknown; however, molecular epidemiological
studies have suggested that pathogenicity is linked to particular lineages. Here,
we performed high-throughput sequencing-based comparative genome analysis of 16
V. nigripulchritudo strains to explore the genomic diversity and evolutionary
history of pathogen-containing lineages and to identify pathogen-specific genetic
elements. Our phylogenetic analysis revealed three pathogen-containing V.
nigripulchritudo clades, including two clades previously identified from New
Caledonia and one novel clade comprising putatively pathogenic isolates from
septicemic shrimp in Madagascar. The similar genetic distance between the three
clades indicates that they have diverged from an ancestral population roughly at
the same time and recombination analysis indicates that these genomes have, in
the past, shared a common gene pool and exchanged genes. As each contemporary
lineage is comprised of nearly identical strains, comparative genomics allowed
differentiation of genetic elements specific to shrimp pathogenesis of varying
severity. Notably, only a large plasmid present in all highly pathogenic (HP)
strains encodes a toxin. Although less/non-pathogenic strains contain related
plasmids, these are differentiated by a putative toxin locus. Expression of this
gene by a non-pathogenic V. nigripulchritudo strain resulted in production of
toxic culture supernatant, normally an exclusive feature of HP strains. Thus,
this protein, here termed 'nigritoxin', is implicated to an extent that remains
to be precisely determined in the toxicity of V. nigripulchritudo.

<>

<1>Gough, J.A., Murray, N.E.
<2>Sequence diversity among related genes for recognition of specific targets in DNA molecules.
<3>J. Mol. Biol.
<4>166
<5>1-19
<6>1983
<7>Escherichia coli strains K12 and B, and a new strain designed D, each encode a
characteristic restriction and modification enzyme.  These enzymes (EcoK, EcoB
and presumably EcoD) comprise three subunits of which one, that encoded by the
so-called specificity gene (hsdS), is responsible for recognition of the DNA
sequence speific to that system.  The other two subunits, encoded by hsdR and
hsdM, are interchangeable between systems, and the available molecular evidence
suggests that the hsdR and hsdM genes are highly conserved.  The DNA sequence
of a segment of the hsd region that includes the hsdS gene has been determined
for each of the three strains.  The hsdS gene varies in length from 1335 to
1425 base-pairs and the only regions showing obvious homology, one of about 100
base-pairs and a second of about 250 base-pairs, are highly conserved.  The
remainder of each hsdS gene shares little, or no, homology with either of the
other related specificity genes.  Thus, the specificity subunits, though
components of a family of closely related enzymes with very similar functions,
have remarkably dissimilar primary structure.

<>

<1>Gough, M., Lederberg, S.
<2>Methylated bases in the host-modified deoxyribonucleic acid of Escherichia coli and bacteriophage lambda.
<3>J. Bacteriol.
<4>91
<5>1460-1468
<6>1966
<7>The deoxyribonucleic acid (DNA) from strains of Escherichia coli and phage
lambda was examined to determine whether the types or amounts of
methionine-derived methylated bases present correlated with the host-specific
modification of that DNA.  The DNA of strain C600 (which has K-12 modification
specificity) and of a modificationless mutant of C600 are similar in their
content of 5-methylcytosine and 6-methylaminopurine.  Strains Bc251 and its
Pl-lysogen differ in P1-controlled specificity, but they have the same content
of 6-methylaminopurine, and both lack 5-methylcytosine in their DNA.  Phage
lambda contains the same methylated bases as its host of origin, but in reduced
amounts and in different proportions.  Although minor amounts of these
methylated bases may have importance as a result of their location, the
presence of the majority of these methylated bases is irrelevant to the
specificity of host modification of DNA.

<>

<1>Gouvea-Taketani, R., Domingues, Z.T., Soares-de-Melo, I., Mendes, R.
<2>Whole-Genome Shotgun Sequencing of Rhodococcus erythropolis Strain P27, a Highly  Radiation-Resistant Actinomycete from Antarctica.
<3>Genome Announcements
<4>1
<5>e00763-13
<6>2013
<7>Here, we report the draft genome sequence of radiation-resistant Rhodococcus erythropolis
strain P27, isolated from leaves of Deschampsia antarctica Desv. (Poaceae) in the Admiralty
Bay area, Antarctica.

<>

<1>Govind, R., Fralick, J.A., Rolfe, R.D.
<2>Genomic Organization and Molecular Characterization of Clostridium difficile Bacteriophage {Phi}CD119.
<3>J. Bacteriol.
<4>188
<5>2568-2577
<6>2006
<7>In this study, we have isolated a temperate phage (PhiCD119) from a
pathogenic Clostridium difficile strain and sequenced and annotated its
genome. This virus has an icosahedral capsid and a contractile tail
covered by a sheath and contains a double-stranded DNA genome. It belongs
to the Myoviridae family of the tailed phages and the order Caudovirales.
The genome was circularly permuted, with no physical ends detected by
sequencing or restriction enzyme digestion analysis, and lacked a cos
site. The DNA sequence of this phage consists of 53,325 bp, which carries
79 putative open reading frames (ORFs). A function could be assigned to 23
putative gene products, based upon bioinformatic analyses. The PhiCD119
genome is organized in a modular format, which includes modules for
lysogeny, DNA replication, DNA packaging, structural proteins, and host
cell lysis. The PhiCD119 attachment site attP lies in a noncoding region
close to the putative integrase (int) gene. We have identified the phage
integration site on the C. difficile chromosome (attB) located in a
noncoding region just upstream of gene gltP, which encodes a carrier
protein for glutamate and aspartate. This genetic analysis represents the
first complete DNA sequence and annotation of a C. difficile phage.

<>

<1>Gowers, D.M., Bellamy, S.R.W., Halford, S.E.
<2>One recognition sequence, seven restriction enzymes, five reaction mechanisms.
<3>Nucleic Acids Res.
<4>32
<5>3469-3479
<6>2004
<7>The diversity of reaction mechanisms employed by Type II restriction enzymes was investigated
by analysing the reactions of seven endonucleases at the same DNA sequence. NarI, KasI,
Mly113I, SfoI, EgeI, EheI and BbeI cleave DNA at several different positions in the sequence
5'-GGCGCC-3'. Their reactions on plasmids with one or two copies of this sequence revealed
five distinct mechanisms. These differ in terms of the number of sites the enzyme binds, and
the number of phosphodiester bonds cleaved per turnover. NarI binds two sites, but cleaves
only one bond per DNA-binding event. KasI also cuts only one bond per turnover but acts at
individual sites, preferring intact to nicked sites. Mly113I cuts both strands of its
recognition sites, but shows full activity only when bound to two sites, which are then
cleaved concertedly. SfoI, EgeI and EheI cut both strands at individual sites, in the manner
historically considered as normal for Type II enzymes. Finally, BbeI displays an absolute
requirement for two sites in close physical proximity, which are cleaved concertedly. The
range of reaction mechanisms for restriction enzymes is thus larger than commonly imagined, as
is the number of enzymes needing two recognition sites.

<>

<1>Gowers, D.M., Halford, S.E.
<2>Protein motion from non-specific to specific DNA by three-dimensional routes aided by supercoiling.
<3>EMBO J.
<4>22
<5>1410-1418
<6>2003
<7>DNA-binding proteins are generally thought to locate their target sites by first associating
with the DNA at random and then translocating to the
specific site by one-dimensional (1D) diffusion along the DNA. We report
here that non-specific DNA conveys proteins to their target sites just as
well when held near the target by catenation as when co-linear with the
target. Hence, contrary to the prevalent view, proteins move from random
to specific sites primarily by three-dimensional (3D) rather than 1D
pathways, by multiple dissociation/re-association events within a single
DNA molecule. We also uncover a role for DNA supercoiling in target-site
location. Proteins find their sites more readily in supercoiled than in
relaxed DNA, again indicating 3D rather than 1D routes.

<>

<1>Gowers, D.M., Wilson, G.G., Halford, S.E.
<2>Measurement of the contributions of 1D and 3D pathways to the translocation of a protein along DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>15883-15888
<6>2005
<7>Proteins that act at specific DNA sequences bind DNA randomly and then translocate to the
target site. The translocation is often ascribed to the protein sliding along the DNA while
maintaining continuous contact with it. Proteins also can move on DNA by multiple cycles of
dissociation/reassociation within the same chain. To distinguish these pathways, a strategy
was developed to analyze protein motion between DNA sites. The strategy reveals whether the
protein maintains contact with the DNA as it transfers from one site to another by sliding or
whether it loses contact by a dissociation/reassociation step. In reactions at low salt, the
test protein stayed on the DNA as it traveled between sites, but only when the sites were <50
bp apart. Transfers of >30 bp at in vivo salt, and over distances of >50 bp at any salt,
always included at least one dissociation step. Hence, for this enzyme, 1D sliding operates
only over short distances at low salt, and 3D dissociation/reassociation is its main mode of
translocation.

<>

<1>Gowher, H., Ehrlich, K.C., Jeltsch, A.
<2>DNA from Aspergillus flavus contains 5-methylcytosine.
<3>FEMS Microbiol. Lett.
<4>205
<5>151-155
<6>2001
<7>DNA from Aspergillus sp. has been reported not to contain 5-methylcytosine. However, it has
been found that Aspergillus nidulans responds to 5-azacytidine, a drug that is a strong
inhibitor of DNA methyltransferases. Therefore, we have re-examined the occurrence of
5-methylcytosine in DNA from Aspergillus flavus by using a highly sensitive and specific
method for detection of modified bases in genomic DNA comprising high-performance liquid
chromatography separation of nucleosides, labeling of the nucleoside with deoxynucleoside
kinase and two-dimensional thin-layer chromatography. Our results show that 5-methylcytosine
is present in DNA from A. flavus. We estimate the relative amounts of 5-methylcytosine to
cytosine to be approximately 1/400.

<>

<1>Gowher, H., Jeltsch, A.
<2>Molecular enzymology of the EcoRV DNA-(Adenine-N6)-methyltransferase: Kinetics of DNA binding and bending, kinetic mechanism and linear diffusion of the enzyme on DNA.
<3>J. Mol. Biol.
<4>303
<5>93-110
<6>2000
<7>The EcoRV DNA-(adenine-N(6))-methyltransferase recognizes GATATC sequences and modifies the
first adenine residue within this site. We show here, that the enzyme binds to the DNA and the
cofactor S-adenosylmethionine (AdoMet) in an ordered bi-bi fashion, with AdoMet being bound
first. M.EcoRV binds DNA in a non-specific manner and the enzyme searches for its recognition
site by linear diffusion with a range of approximately 1800 bp. During linear diffusion the
enzyme continuously scans the DNA for the presence of recognition sites. Upon specific
M.EcoRV-DNA complex formation a strong increase in the fluorescence of an oligonucleotide
containing a 2-aminopurine base analogue at the GAT-2AP-TC position is observed which, most
likely, is correlated with DNA bending. In contrast to the GAT-2AP-TC substrate, a G-2AP-TATC
substrate in which the target base is replaced by 2-aminopurine does not show an increase in
fluorescence upon M.EcoRV binding, demonstrating that 2-aminopurine is not a general tool to
detect base flipping. Stopped-flow experiments show that DNA bending is a fast process with
rate constants >10 s(-1).  In the presence of cofactor, the specific complex adopts a second
conformation, in which the target sequence is more tightly contacted by the enzyme. M.EcoRV
exists in an open and in a closed state that are in slow equilibrium. Closing the open state
is a slow process (rate constant approximately 0.7 min(-1)) that limits the rate of DNA
methylation under single turnover conditions. Product release requires opening of the closed
complex which is very slow (rate constant approximately 0.05-0.1 min(-1)) and limits the rate
of DNA methylation under multiple turnover conditions. M.EcoRV methylates DNA sequences
containing more than one recognition site in a distributive manner. Since the dissociation
rate from non-specific DNA does not depend on the length of the DNA fragment, DNA dissociation
does not preferentially occur at the ends of the DNA.

<>

<1>Gowher, H., Jeltsch, A.
<2>Molecular enzymology of the catalytic domains of the Dnmt3a and Dnmt3b DNA methyltransferases.
<3>J. Biol. Chem.
<4>277
<5>20409-20414
<6>2002
<7>The C-terminal domains of the mammalian DNA methyltransferases Dnmt1, Dnmt3a, and Dnmt3b
harbor all the conserved motifs characteristic for
cytosine-C5 methyltransferases. Whereas the isolated catalytic domain
of Dnmt1 is inactive, we show here that the C-terminal domains of
Dnmt3a and Dnmt3b are catalytically active. Neither Dnmt3a nor Dnmt3b
shows a significant preference for the satellite 2 sequence, although
Dnmt3b is required for methylation of these regions in vivo. However,
the catalytic domain of Dnmt3a methylates DNA in a distributive
reaction, whereas Dnmt3b is processive, which accelerates methylation
of macromolecular DNA in vitro. This property could make Dnmt3b a
preferred enzyme for methylation at satellite 2 repeats, since they are
highly CG-rich. We have also analyzed the catalytic activities of six
different mutations found in ICF (immunodeficiency, centromeric
instability, and facial abnormalities) patients in the catalytic domain
of Dnmt3b. Five of them display catalytic activities reduced by
10-50-fold; one mutant was inactive in our assay (residual activity
<1%). These results confirm that a reduced catalytic activity of Dnm3b
causes ICF. However, the mutations in general do not completely
abrogate catalytic activity. This finding may explain why ICF patients
are viable, whereas nmt3b knock-out mice die during embryogenesis.

<>

<1>Gowher, H., Jeltsch, A.
<2>Enzymatic properties of recombinant Dnmt3a DNA methyltransferase from mouse: the enzyme modifies DNA in a non-processive manner and also methylates non-CpG Sites.
<3>J. Mol. Biol.
<4>309
<5>1201-1208
<6>2001
<7>We present the first in vitro study investigating the catalytic properties of a mammalian de
novo DNA methyltransferase. Dnmt3a from mouse was cloned and expressed in Escherichia coli. It
was shown to be catalytically active in E. coli cells in vivo. The methylation activity of the
purified protein was highest at pH 7.0 and 30 mM KCl. Our data show that recombinant Dnmt3a
protein is indeed a de novo methyltransferase, as it catalyzes the transfer of methyl groups
to unmethylated substrates with similar efficiency as to hemimethylated substrates. With
oligonucleotide substrates, the catalytic activity of Dnmt3a is similar to that of Dnmt1: the
Km values for the unmethylated and hemimethylated oligonucleotide substrates are 2.5 microM,
and the k(cat) values are 0.05 h^-1 and 0.07 h^-1, respectively. The enzyme catalyzes the
methylation of DNA in a distributive manner, suggesting that Dnmt3a and Dnmt1 may cooperate
during de novo methylation of DNA. Further, we investigated the methylation activity of Dnmt3a
at non-canonical sites. Even though the enzyme shows maximum activity at CpG sites, with
oligonucleotide substrates, a high methylation activity was also found at CpA sites, which are
modified only twofold slower than CpG sites. Therefore, the specificity of Dnmt3a is
completely different from that of the maintenance methyltransferase Dnmt1, which shows a 40 to
50-fold preference for hemimethylated over unmethylated CpG sites and has almost no
methylation activity at non-CpG sites. Copyright 2001 Academic Press.

<>

<1>Gowher, H., Leismann, O., Jeltsch, A.
<2>DNA of Drosophila melanogaster contains 5-methylcytosine.
<3>EMBO J.
<4>19
<5>6918-6923
<6>2000
<7>It is commonly accepted that the DNA of Drosophila melanogaster does not contain
5-methylcytosine, which is essential in the development of most eukaryotes. We have developed
a new, highly specific and sensitive assay to detect the presence of 5-methylcytosine in
genomic DNA. The DNA is degraded to nucleosides, 5-methylcytosine purified by HPLC and, for
detection by 1D- and 2D-TLC, radiolabeled using deoxynucleoside kinase and [-32P]ATP. Using
this assay, we show here that 5- methylcytosine occurs in the DNA of D. melanogaster at a
level of 1 in 1000-2000 cytosine residues in adult flies. DNA methylation is detectable in all
stages of D. melanogaster development.

<>

<1>Gowher, H., Liebert, K., Hermann, A., Xu, G.L., Jeltsch, A.
<2>Mechanism of stimulation of catalytic activity of Dnmt3A and Dnmt3B DNA-(cytosine-C5)-methyltransferases by Dnmt3L.
<3>J. Biol. Chem.
<4>280
<5>13341-13348
<6>2005
<7>Dnmt3L has been identified as a stimulator of the catalytic activity of de novo DNA
methyltransferases. It is essential in the development of
germ cells in mammals. We show here that Dnmt3L stimulates the
catalytic activity of the Dnmt3A and Dnmt3B enzymes by directly binding
to their respective catalytic domains via its own C-terminal domain.
The catalytic activity of Dnmt3A and -3B was stimulated similar to
15-fold, and Dnmt3L directly binds to DNA but not to
S-adenosyl-L-methionine (AdoMet). Complex formation between Dnmt3A and
Dnmt3L accelerates DNA binding by Dnmt3A 20-fold and lowers its Km for
DNA. Interaction of Dnmt3L with Dnmt3A increases the binding of the
coenzyme AdoMet to Dnmt3A, and it lowers the Km of Dnmt3A for AdoMet.
On the basis of our data we propose a model in which the interaction of
Dnmt3A with Dnmt3L induces a conformational change of Dnmt3A that opens
the active site of the enzyme and promotes binding of DNA and the
AdoMet. We demonstrate that the interaction of Dnmt3A and Dnmt3L is
transient, and after DNA binding to Dnmt3A, Dnmt3L dissociates from the
complex. Following dissociation of Dnmt3L, Dnmt3A adopts a closed
conformation leading to slow rates of DNA release. Therefore, Dnmt3L
acts as a substrate exchange factor that accelerates DNA and AdoMet
binding to de novo DNA methyltransferases.

<>

<1>Gowher, H., Loutchanwoot, P., Vorobjeva, G., Handa, V., Jurkowska, R.Z., Jurkowski, T.P., Jeltsch, A.
<2>Mutational analysis of the catalytic domain of the murine Dnmt3a DNA-(cytosine C5)-methyltransferase.
<3>J. Mol. Biol.
<4>357
<5>928-941
<6>2006
<7>On the basis of amino acid sequence alignments and structural data of related enzymes, we have
performed a mutational analysis of 14 amino
acid residues in the catalytic domain of the murine Dnmt3a
DNA-(cytosine C5)-methyltransferase. The target residues are located
within the ten conserved amino acid sequence motifs characteristic for
cytosine-C5 methyltransferases and in the putative DNA recognition
domain of the enzyme (TRD). Mutant proteins were purified and tested
for their catalytic properties and their abilities to bind DNA and
AdoMet. We prepared a structural model of Dnmt3a. to interpret our
results. We demonstrate that Phe50 (motif I) and Glu74 (motif II) are
important for AdoMet binding and catalysis. D96A (motif III) showed
reduced AdoMet binding but increased activity under conditions of
saturation with S-adenosyl-L-methionine (AdoMet), indicating that the
contact of Asp96 to AdoMet is not required for catalysis. R130A
(following motif IV), R241A and R246A (in the TRD), R292A, and R297A
(both located in front of motif X) showed reduced DNA binding. R130A
displayed a strong reduction in catalytic activity and a complete
change in flanking sequence preferences, indicating that Arg130 has an
important role in the DNA interaction of Dnmt3a. R292A also displayed
reduced activity and changes in the flanking sequence preferences,
indicating a potential role in DNA contacts farther away from the CG
target site. N167A (motif VI) and R202A (motif VIII) have normal AdoMet
and DNA binding but reduced catalytic activity. While Asn167 might
contribute to the positioning of residues from motif VI, according to
structural data Arg202 has a role in catalysis of cytosine-C5
methyltransferases. The R295A variant was catalytically inactive most
likely because of destabilization of the hinge sub-domain of the
protein.

<>

<1>Gowher, H., Stockdale, C.J., Goyal, R., Ferreira, H., Owen-Hughes, T., Jeltsch, A.
<2>De novo methylation of nucleosomal DNA by the mammalian Dnmt1 and Dnmt3A DNA methyltransferases.
<3>Biochemistry
<4>44
<5>9899-9904
<6>2005
<7>In the cell, DNA is wrapped on histone octamers, which reduces its accessibility for DNA
interacting enzymes. We investigated de novo
methylation of nucleosomal DNA in vitro and show that the Dnmt3a and
Dnmt1 DNA methyltransferases efficiently methylate nucleosomal DNA
without dissociation of the histone octamer from the DNA. In contrast,
the prokaryotic SssI DNA methyltransferase and the catalytic domain of
Dnmt3a are strongly inhibited by nucleosomes. We also found that
full-length Dnmt1 and Dnmt3a bind to nucleosomes much stronger than
their isolated catalytic domains, demonstrating that the N-terminal
parts of the MTases are required for the interaction with nucleosomes.
Variations of the DNA sequence or the histone tails did not
significantly influence the methylation activity of Dnmt3a. The
observation that mammalian methyltransferases directly modify
nucleosomal DNA provides an insight into the mechanisms by which
histone tail and DNA methylation patterns can influence each other
because the DNA methylation pattern can be established while histones
remain associated to the DNA.

<>

<1>Goyal, R., Rathert, P., Laser, H., Gowher, H., Jeltsch, A.
<2>Phosphorylation of serine-515 activates the mammalian maintenance methyltransferase Dnmt1.
<3>EPIGENETICS
<4>2
<5>155-160
<6>2007
<7>DNA methyltransferase 1 methylates hemi-methylated CG sites generated during DNA replication.
Serine 515 of this enzyme has been shown to be
phosphorylated. To explore the importance of S515 phosphorylation, we
generated mutants of Dnmt1 which removed the phosphorylation potential
(S515A) or mimic phosphoserine (S515E), purified the proteins from
insect cells and analyzed their DNA methylation activity in vitro. The
S515E mutant was found to be active, while S515A mutant had severe loss
in activity when compared to the wild type protein. The loss of
activity of the S515A variant was not due to loss of DNA binding
capacity. Furthermore, we show that a phosphorylated peptide whose
sequence mimics the surrounding of Ser515 ((EKIYISKIVVE)-K-P) inhibited
the activity of wild type Dnmt1 ten-fold more than the
non-phosphorylated peptide. The inhibition was specific for Dnmt1 and
for the particular peptide sequence. Our data suggest that
phosphorylation of Ser515 is important for an interaction between the
N-terminal domain of Dnmt1 and its catalytic domain that is necessary
for activity and that this interaction is specifically disrupted by the
phosphorylated peptide. We conclude that phosphorylation of Dnmt1 at
Ser515 could be an important regulator of Dnmt1 activity during cell
cycle and after proliferative stimuli.

<>

<1>Goyal, R., Reinhardt, R., Jeltsch, A.
<2>Accuracy of DNA methylation pattern preservation by the Dnmt1 methyltransferase.
<3>Nucleic Acids Res.
<4>34
<5>1182-1188
<6>2006
<7>DNA methyltransferase 1 (Dnmt1) has a central role in copying the pattern of DNA methylation
after replication which is one manifestation of
epigenetic inheritance. With oligonculeotide substrates we show that mouse
Dnmt1 has a 30- to 40-fold preference for hemimethylated DNA that is
almost lost after addition of fully methylated oligonucleotides. Using
long hemimethylated DNA substrates that carry defined methylation patterns
and bisulfite analysis of the methylation reaction products, we show a
15-fold preference for hemimethylated CG sites. Dnmt1 moves along the DNA
in a random walk methylating hemimethylated substrates with high
processivity (>50 sites are visited on average which corresponds to linear
diffusion over 6000 bp). The frequency of skipping sites is very low
(<0.3%) and there is no detectable flanking sequence preference. CGCTC
sites tend to terminate the processive methylation of DNA by Dnmt1.
Unmethylated DNA is modified non-processively with a preference for
methylation at CCGG sites. We simulate the propagation of methylation
patterns using a stochastic model with the specificity of Dnmt1 observed
here and conclude that either methylation of several sites is required to
propagate the methylation information over several cellular generations or
additional epigenetic information must be used.

<>

<1>Goyon, C.
<2>Isolation and identification by sequence homology of a second putative C5-DNA-methyltransferase gene from Ascobolus immersus.
<3>DNA Seq.
<4>9
<5>109-112
<6>1998
<7>I report the cloning of a new Ascobolus gene (masc 2) that potentially encodes a
C5-DNA-methyltransferase. The putative protein exhibits the two domains characteristic of
eukaryotic maintenance DNA-methyltransferase: a large N-terminal domain and a C-terminal
domain containing all ten catalytic motifs arranged in the canonical order. A new type of
eukaryotic DNA-methylase gene (masc 1) has been recently found in Ascobolus. Masc1 is
essential for the de novo methylation and dispensable for methylation maintenance. The masc2
gene could encode the methylase involved in this maintenance.

<>

<1>Goyon, C., Nogueira, T.I.V., Faugeron, G.
<2>Perpetuation of cytosine methylation in Ascobolus immersus implies a novel type of maintenance methylase.
<3>J. Mol. Biol.
<4>240
<5>42-51
<6>1994
<7>In the ascomycete Ascobolus immersus, duplicated DNA segments are subject to the methylation
induced premeiotically (MIP) process. Affected sequences are heavily methylated at their
cytosine residues. We used the bisulphite genomic sequencing method to determine the
methylation status of every cytosine residue in a gene which had undergone MIP. Several
individual DNA molecules, all issued from the replication of a single molecule initially
subject to MIP, were sequenced. In each molecule, methylation extended over almost the whole
length of the previously duplicated segment. The methylation extent was precisely delimited
and constant in each of the molecules, leaving unmethylated a nearly 100-nucleotide region
next to each end. In none of the molecules did methylation resulting from MIP extend beyond
the ends. Although the DNA molecules were not all methylated with the same intensity, all
cytosine residues in the methylated portion could be methylated, most of them belonging to
non-symmetrical sequences. This finding contrasts with the situation in higher eukaryotes in
which most, if not all, methylation is at short symmetrical sequences such as CpG or CpNpG,
ensuring perpetuation of methylation. Methylation at non-symmetrical sequences implies that in
A. immersus maintenance involves a novel sequence-non-specific methyltransferase.

<>

<1>Grable, J., Frederick, C.A., Samudzi, C., Jen-Jacobson, L., Lesser, D., Greene, P., Boyer, H.W., Itakura, K., Rosenberg, J.M.
<2>Two-fold symmetry of crystalline DNA-EcoRI endonuclease recognition complexes.
<3>J. Biomol. Struct. Dyn.
<4>1
<5>1149-1160
<6>1984
<7>Recognition complexes between EcoRI endonuclease and either of two synthetic
oligonucleotides (sequences CGCGAATTCGCG and TCGCGAATTCGCG) crystallize in
space Group P321 with unit cell parameters a=128 and c=47 angstroms and a=118.4
and c=49.7 angstroms, respectively.  Native diffraction data to 3 angstrom
resolution have been collected from the form containing the tridecameric
sequence.  Electrophoretic analyses of dissolved crystals demonstrate that this
form contains DNA and protein in a ratio of one double helix per enzyme dimer.
The most likely asymmetric unit contents are one 31,000 dalton enzyme subunit
and one strand of DNA, yielding VM values of 3.1 cubic angstroms/dal and 2.8
cubic angstroms/dal for the forms containing dodecameric and tridecameric DNA,
respectively.  This implies that the DNA-protein complex possesses two-fold
rotational symmetry, which has been incorporated in the crystalline lattice.

<>

<1>Grabowski, G., Alves, J.
<2>Transformation of the EcoRI restriction endonuclease to an enzyme with altered specificity: Development of a positive in vivo selection system.
<3>Biol. Chem. Hoppe Seyler
<4>376
<5>S102
<6>1995
<7>The EcoRI restriction endonuclease is one of the best studied enzymes so far.  It cleaves the
DNA sequence G/AATTC with very high specificity.  All attempts to change the sequence
specificity of this enzyme by using site-directed mutagenesis methods have not been successful
until today.  This is caused by the redundancy of enzyme-substrate interactions leading to
specific sequence recognition and cleavage.  Therefore, we have developed a positive in vivo
selection system that allows screening for an active enzyme with altered specificity.
Bacterial cells are transformed with a pool of plasmids containing the randomly mutated
endonuclease gene.  In order to protect the host DNA from cleavage a methyltransferase
corresponding to the altered specificity is required.  We used the MunI methylase which is
specific for the cognate sequence CAATTG.  Selection is carried out by infecting the cells
with a lambda phage, which carried a gene coding for a toxic protein.  The sequence of this
gene was altered to generate several sites of the desired new specificity.  The transcription
of the toxic gene is prevented only when a restriction activity with this specificity is
present.

<>

<1>Grabowski, G., Jeltsch, A., Wolfes, H., Maass, G., Alves, J.
<2>Site-directed mutagenesis in the catalytic center of the restriction endonuclease EcoRI.
<3>Gene
<4>157
<5>113-118
<6>1995
<7>The catalytic center of the restriction endonuclease (ENase) EcoRI is structurally homologous
to that of EcoRV, BamHI and PvuII.  Each of these ENases contains a short motif of three to
four amino acid (aa) residues which are positioned in a similar orientation to the scissile
phosphodiester bond.  We have mutated these aa (Pro90, Asp91, Glu111 and Lys113) in EcoRI to
determine their individual roles in catalysis.  The replacement of Asp91 and Lys113,
respectively, by conservative mutations (Ala91, Asn91, Ala113, Gln113, His113 and Leu113)
resulted in a reduction of binding affinity and complete loss of cleavage activity.  Only
Lys113-Arg substitution still allows to cleave DNA, albeit with a rate reduced by at least
four orders of magnitude.  Lys113 seems to stabilize the structure of the wild-type (wt) ENase
since all five ENase variants with mutations at this position show a strongly enhanced
tendency to aggregate.  The Ala and Gln mutants of Glu111 bind the recognition sequence
slightly stronger than wt EcoRI and cleave it with a low, but detectable rate.  Only the
Glu111-Lys mutant, in which the charge is reversed, shows neither binding nor cleavage
activity.  Pro90 is not important for catalysis, because the Ala90 mutant cleaves DNA with an
only slightly reduced rate.  Under star conditions, however, this mutant is even more active
than wt EcoRI.  Therefore, the charged aa Asp91, Glu111 and Lys113 are essential for catalytic
activity of the EcoRI ENase.  Differences in the individual contributions of these aa to
binding and catalysis, as compared with results obtained with EcoRV and BamHI mutants, show
that similar catalytic centers are used in a slightly different way by these three ENases.

<>

<1>Grabowski, G., Maass, G., Alves, J.
<2>Asp-59 is not important for the catalytic activity of the restriction endonuclease EcoRI.
<3>FEBS Lett.
<4>381
<5>106-110
<6>1996
<7>The amino acid Asp-59 was proposed to be involved in EcoRI catalyzed DNA cleavage.  We have
tested this hypothesis by site directed mutagenesis experiments.  The four mutants D59A, D59E,
D59G, and D59N bind with similar stability to the specific recognition sequence as wild type
EcoRI.  The D59E mutant cleaves DNA as fast as the wild type enzyme.  Specific activities of
the other three mutants are five to tenfold lower.  Therefore, we conclude that Asp-59  is not
involved in catalysis of the EcoRI restriction endonuclease.  Consequences for catalytic
mechanisms of EcoRI and other restriction enzymes are discussed.

<>

<1>Grachev, S.A., Mamaev, S.V., Gurevich, A.I., Igoshin, A.V., Kolosov, M.N., Slyusarenko, A.G.
<2>Restriction endonuclease TaqXI from Thermus aquaticus.
<3>Bioorg. Khim.
<4>7
<5>628-630
<6>1981
<7>A new restriction endonuclease TaqXI has been isolated from an unidentified
strain of Thermus aquaticus.  The enzyme recognizes the pentanucleotide
sequence CC(A/T)GG and cleaves it between C and A or T, the methylation of the
C residue is not protecting the sequence from the cleavage.

<>

<1>Grad, Y.H. et al.
<2>Genomic epidemiology of the Escherichia coli O104:H4 outbreaks in Europe, 2011.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>109
<5>3065-3070
<6>2012
<7>The degree to which molecular epidemiology reveals information about the sources  and
transmission patterns of an outbreak depends on the resolution of the
technology used and the samples studied. Isolates of Escherichia coli O104:H4
from the outbreak centered in Germany in May-July 2011, and the much smaller
outbreak in southwest France in June 2011, were indistinguishable by standard
tests. We report a molecular epidemiological analysis using multiplatform
whole-genome sequencing and analysis of multiple isolates from the German and
French outbreaks. Isolates from the German outbreak showed remarkably little
diversity, with only two single nucleotide polymorphisms (SNPs) found in isolates
from four individuals. Surprisingly, we found much greater diversity (19 SNPs) in
isolates from seven individuals infected in the French outbreak. The German
isolates form a clade within the more diverse French outbreak strains. Moreover,
five isolates derived from a single infected individual from the French outbreak
had extremely limited diversity. The striking difference in diversity between the
German and French outbreak samples is consistent with several hypotheses,
including a bottleneck that purged diversity in the German isolates, variation in
mutation rates in the two E. coli outbreak populations, or uneven distribution of
diversity in the seed populations that led to each outbreak.

<>

<1>Graentzdoerffer, A., Lindenstrauss, U., Pich, A., Andreesen, J.R.
<2>New restriction endonuclease EacI, useful for specific cleavage of double stranded DNA e.g. for analysis, derived from Eubacterium acidaminophilum - recombinant enzyme production via plasmid expression in host cell for DNA cleavage.
<3>German Patent Office
<4>DE 10060525
<5>
<6>2002
<7>NOVELTY - Restriction endonuclease EacI from Eubacterium acidaminophilum. The sequence
encoding EacI, 1713 bp, and the derived 570 amino acids protein sequence are fully defined in
the specification. DETAILED DESCRIPTION - An INDEPENDENT CLAIM is alo included for preparation
of EacI by biochemical isolation from E. acidaminophilum or by overexpressing the coding
sequence in Escherichia coli. BIOTECHNOLOGY - EacI recognizes the motif 5'-GGATC and cuts the
strand containing this motif 4 bases downstream from the C and the complementary strand 5
bases downstream. Preparation: EacI (located in the cytoplasm) is isolated conventionally from
cultures of E. acidaminophilum, purified to homogeneity and its N-terminal sequence
determined. To isolate the coding sequence, total DNA from E. acidaminophilum was digested
with restriction enzymes, and the fragments cloned into vectors for transformation of a
methylase-negative strain of E. coli. The plasmid pools were digested with EacI, the products
used to transform E. coli again and only protected, i.e. properly methylated, plasmids were
able to replicate. Inserts were isolated from replicating plasmids and one insert contained
two open reading frames; one for the methyltransferase and the other for EacI. The EacI
sequence was cloned into a commercial expression plasmid for transformation of E. coli so that
a fusion with a peptide tag (to facilitate affinity purification) was produced. USE - EacI is
used for specific cleavage of double-stranded DNA, to produce well-defined fragments or for
analysis. ADVANTAGE - EacI can be isolated easily, in stable form, from E. acidaminophilum,
and since its coding sequence has been determined, it can be overexpressed in E. coli for
large scale production, optionally in mutated form (with modified properties and better
stability). EXAMPLE - Escherichia coli transformed with the vector pASK-IBA7 that contained
the coding sequence for an EacI-Strep-tagII fusion protein was cultured to optical density
(600 nm) 0.5. EacI expression was then induced by adding anhydrotetracycline and after 3 hr
the cells were harvested, lysed in pH 7.5 buffer and centrifuged again. The clear supernatant
was applied to a Strep-Tactin column and this eluted with buffer containing 2.5 mg/ml
dethiobiotin to recover a fraction containing pure EacI.

<>

<1>Graentzdoerffer, A., Lindenstrauss, U., Pich, A., Andreesen, J.R.
<2>New DNA-methyltransferase M.EacI, useful for protecting double stranded DNA against cleavage by restriction enzymes, derived from Eubacterium acidaminophilum.
<3>German Patent Office
<4>DE 10060526
<5>
<6>2002
<7>NOVELTY - DNA-methyltransferase M.EacI from Eubacterium acidaminophilum. The sequence encoding
M.EacI of 1929 base pairs (bp), and the derived protein sequence, 642 amino acids, are fully
defined in the specification. DETAILED DESCRIPTION - An INDEPENDENT CLAIM is also included for
preparation of M.EacI by biochemical isolation from E. acidaminophilum or by overexpressing
the coding sequence in Escherichia coli. BIOTECHNOLOGY - Preparation: M.EacI (located in the
cytoplasm) is isolated conventionally from cultures of E. acidaminophilum and purified to
homogeneity. To isolate the coding sequence, total DNA from E. acidaminophilum was digested
with restriction enzymes, and the fragments cloned into vectors for transformation of a
methylase-negative strain of E. coli. The plasmid pools were digested with EacI (the
restriction endonuclease associated with M.EacI), the products used to transform E. coli again
and only protected, i.e. properly methylated, plasmids were able to replicate. Inserts were
isolated from replicating plasmids and one insert contained two open reading frames; one for
M.EacI and the other for EacI. The M.EacI sequence was cloned into a commercial expression
plasmid for transformation of E. coli so that a fusion with a peptide tag (to facilitate
affinity purification) was produced. USE - M.EacI is used to inhibit specific cleavage of
double-stranded DNA by restriction enzymes, e.g. EacI or AlwI, that recognize the motif
5'-GGATC. EXAMPLE - Escherichia coli transformed with the vector pASK-IBA7 that contained the
coding sequence for an M.EacI-Strep-tagII fusion protein was cultured to optical density (600
nm) 0.5. M.EacI expression was then induced by adding anhydrotetracycline and after 3 hours
the cells were harvested, lysed in pH 7.5 buffer and centrifuged again. The clear supernatant
was applied to a Strep-Tactin column and this eluted with buffer containing 2.5 mg/ml
dethiobiotin to recover a fraction containing pure M.EacI.

<>

<1>Graessmann, M., Graessmann, A., Wagner, H., Werner, E., Simon, D.
<2>Complete DNA methylation does not prevent polyoma and simian virus 40 virus early gene expression.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>80
<5>6470-6474
<6>1983
<7>The effect of DNA methylation on polyoma virus and simian virus 40 gene
expression was investigated.  For this purpose, the cytosines of all C-G
dinucleotides of the viral DNAs were methylated by the use of rat liver
methylase and the completeness of methylation was verified by dinucleotide
analysis and restriction endonuclease treatment.  The biological activity of
unmethylated and fully methylated DNAs was tested by microinjecting them into
tissue culture cells.  The functions analyzed included early and late viral
gene expression, viral DNA replication, oncogenic transformation efficiency,
and virus maturation.  No difference in any of these biological functions was
observed between methylated and unmethylated DNA.  Early gene expression of
methylated DNA is not the result of demethylation because viral DNA reextracted
from the injected cells, under nonpermissive conditions, retained the
methylation pattern of the input DNA.  In contrast, viral DNA extracted from
transformed cells or from intact virus particles was partially or completely
demethylated.

<>

<1>Grande, L., Michelacci, V., Tozzoli, R., Ranieri, P., Maugliani, A., Caprioli, A., Morabito, S.
<2>Whole genome sequence comparison of vtx2-converting phages from Enteroaggregative Haemorrhagic Escherichia coli strains.
<3>BMC Genomics
<4>15
<5>574
<6>2014
<7>BACKGROUND: Enteroaggregative Haemorrhagic E. coli (EAHEC) is a new pathogenic
group of E. coli characterized by the presence of a vtx2-phage integrated in the
genomic backbone of Enteroaggregative E. coli (EAggEC). So far, four distinct
EAHEC serotypes have been described that caused, beside the large outbreak of
infection occurred in Germany in 2011, a small outbreak and six sporadic cases of
HUS in the time span 1992-2012. In the present work we determined the whole
genome sequence of the vtx2-phage, termed Phi-191, present in the first described
EAHEC O111:H2 isolated in France in 1992 and compared it with those of the
vtx-phages whose sequences were available. RESULTS: The whole genome sequence of
the Phi-191 phage was identical to that of the vtx2-phage P13374 present in the
EAHEC O104:H4 strain isolated during the German outbreak 20 years later.
Moreover, it was also almost identical to those of the other vtx2-phages of EAHEC
O104:H4 strains described so far. Conversely, the Phi-191 phage appeared to be
different from the vtx2-phage carried by the EAHEC O111:H21 isolated in the
Northern Ireland in 2012.The comparison of the vtx2-phages sequences from EAHEC
strains with those from the vtx-phages of typical Verocytotoxin-producing E. coli
strains showed the presence of a 900 bp sequence uniquely associated with EAHEC
phages and encoding a tail fiber. CONCLUSIONS: At least two different
vtx2-phages, both characterized by the presence of a peculiar tail fiber-coding
gene, intervened in the emergence of EAHEC. The finding of an identical
vtx2-phage in two EAggEC strains isolated after 20 years in spite of the high
variability described for vtx-phages is unexpected and suggests that such
vtx2-phages are kept under a strong selective pressure.The observation that
different EAHEC infections have been traced back to countries where EAggEC
infections are endemic and the treatment of human sewage is often ineffective
suggests that such countries may represent the cradle for the emergence of the
EAHEC pathotype. In these regions, EAggEC of human origin can extensively
contaminate the environment where they can meet free vtx-phages likely spread by
ruminants excreta.

<>

<1>Grandi, G., Fraser, C.M., Rappuoli, R., Scarlato, V., Scarselli, M., Ratti, G., Galeotti, C., Mora, M., Masignan, V., Venter, C.J., Tetterin, H., Peterson, J., Hickey, E., Pizza, M.
<2>Neisseria Genomic Research.
<3>Japanese Patent Office
<4>JP 2003527079 A
<5>
<6>2003
<7>
<>

<1>Grandi, G., Masignani, V., Venter, C.J., Mora, M., Scarseri, M., Scarlato, V., Rappuoli, R., Pizza, M., Galeotti, C., Ratti, G., Teterine, H., Peterson, J., Hickey, E., Fraser, C.M.
<2>Neisseria Genomic Sequences And Methods Of Their Use.
<3>Japanese Patent Office
<4>JP 2004511201 A
<5>
<6>2004
<7>
<>

<1>Grandori, R., Sander, C.
<2>Identification by computer sequence analysis of transcriptional regulator proteins in Dictyostelium discoideum and Serratia marcescens.
<3>Nucleic Acids Res.
<4>19
<5>2359-2362
<6>1991
<7>We have performed computer searches in the database of known protein sequences
for proteins similar in sequence to bacteriophage regulatory proteins of known
3-D structure.  The searches are more selective than other methods due to the
use of a length-dependent threshold in sequence similarity, above which
structural homology is implied with high certainty.  Two probable DNA binding
proteins were identified which are predicted to have a three-dimensional
structure very similar to bacteriophage cro and repressor proteins.
Approximate three-dimensional model coordinates are available from the authors.
Both proteins contain the helix-turn-helix sequence motif typical of a wide
class of DNA binding proteins and their function is deduced by analogy to
sequence-similar proteins of known function.  We predict that the Y.SmaI
protein in the restriction-modification enzyme gene locus of the
enterobacterium serratia marcescens is a regulator of endonuclease expression;
and, that the vegetative specific gene VSH7 of the slime mold dictyostelium
discoideum codes for a regulator of gene expression specific for the slime mold
growth phage before the onset of the developmental program.  Point mutations
that would have a strong effect on growth regulation phenotype are suggested.
The VSH7 protein would be the first eukaryotic representative of the cro /
phage repressor class.

<>

<1>Grant, S.G., Jessee, J., Bloom, F.R., Hanahan, D.
<2>Differential plasmid rescue from transgenic mouse DNAs into Escherichia coli methylation-restriction mutants.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>87
<5>4645-4649
<6>1990
<7>Plasmids comprising transgene insertions in four lines of transgenic mice have been retrieved
by plasmid rescue into a set of Escherichia coli strains with
mutations in different members of the methylation-dependent restriction system
(MDRS). Statistical analysis of plasmid rescue frequencies has revealed that the
MDRS loci detect differential modifications of the transgene insertions among
mouse lines that show distinctive patterns of transgene expression. Plasmids in
mice that express hybrid insulin transgenes during development can be readily
cloned into E. coli strains carrying mutations in two of the MDRS loci, mcrA and
mcrB. In mice in which transgene expression is inappropriately delayed into
adulthood, plasmids can only be cloned into E. coli that carry mutations in all
known MDRS activities. Differential cloning frequencies in the presence or
absence of the various methylation-dependent restriction genes represent a
further way to distinguish regions of mammalian chromosomes. These multiply
deficient E. coli strains will also facilitate the molecular cloning of modified
chromosomal DNA.

<>

<1>Grasby, J.A., Connolly, B.A.
<2>Stereochemical outcome of the hydrolysis reaction catalyzed by the EcoRV restriction endonuclease.
<3>Biochemistry
<4>31
<5>7855-7861
<6>1992
<7>The stereochemical course of the reaction catalyzed by the EcoRV restriction and endonuclesae
has been determined. This endonuclease recognizes GATATC sequences and cuts between the
central T and dA bases. The Rp isomer of d(GACGATsATCGTC) (this dodecamer contains a
phosphorothioate rather than the usual phosphate group between the central T and dA residues,
indicated by the s) was a substrate for the endonuclease. Performing this reaction in H2[18O]
gave [18O]dps(ATCGTC)(a pentamer containing an 18 O-labeled 5'-phosphorothioate) which was
converted to [18O]dAMPS with nuclese P1. This deoxynucleoside 5'-[18O]phosphorothioate was
sterospecifically converted to [18O]dATPalphaS with adenylate kinase and pyruvate kinase
[Brody, R.S., & Frey, P.A.(1981) Biochemistry 20, 1245-1251]. Analysis of the position of the
18O in this product by 31P NMR spectroscopy showed that it was in a bridging position between
the alpha- and beta-phosphorus atoms. This indicates that the EcoRV hydrolysis proceeds with
inversion of configuration at phosphorus. The simplest interpretation is that the mechanism of
this endonuclease involves a direct in-line attack at phosphorus by H2O with a trigonal
bipyramidal transition state. A covalent enzyme oligodeoxynucleotide species can be discounted
as an intermediate. An identical result has been previously observed with the EcoRI
endonuclease [Connolly,B.A., Eckstein,F., & Pingoud,A.(1984) J. Biol. Chem. 259, 10760-10763].
X-ray crystallography has shown that both of these endonucleases contain a conserved array of
amino acids at their active sites. Possible mechanistic roles for these conserved amino acids
in the light of the stereochemical findings are discussed.

<>

<1>Grasso, R.J., Paigen, K.
<2>Loss of host-controlled restriction of lambda bacteriophage in Escherichia coli following methionine deprivation.
<3>J. Virol.
<4>2
<5>1368-1373
<6>1968
<7>Lambda bacteriophages produced in Escherichia coli C (designated as lambda.C)
are restricted in their ability to grow in E. coli K-12.  The rare successful
infections that arise in the K-12 population occur in "special" cells which
have lost their capacity to restrict lambda.C.  These infections yield modified
progeny phage (designated as lambda.K) which unlike lambda.C, plate equally
well on E. coli C and E. coli K-12.  When methionine, but no other amino acid,
was removed from the growth medium of a mutant strain of E. coli K-12, the
number of special cells rapidly increased 500- to 3,000-fold.  These new
special cells retain their capacity to produce modified lambda.K progeny.  This
conversion of restricting cells into special cells does not require the
synthesis of new protein.  The special cells formed when methionine was removed
from the culture did not revert into restricting cells when methionine was
restored.  Such cells have also lost the ability to divide for at least 4 hr
after methionine supplementation.  When methionine was restored, the remaining
restricting cells, but not the special cells, immediately resumed growth.
Removing methionine from cultures of E. coli B caused a similar increase in the
number of special cells able to support the growth of lambda.C and lambda.K.
However, when E. coli K-12 (P1) cultures were deprived of methionine, the
number of special cells increased for lambda.C but not for lambda.K.  Thus,
retention of the P1-restriction system, unlike the B- and the K-12 systems,
does not require the presence of methionine.

<>

<1>Grasso, R.J., Paigen, K.
<2>The effect of amino acids on host-controlled restriction of lambda phage.
<3>Virology
<4>36
<5>1-8
<6>1968
<7>Phage lambda grown in Escherichia coli C (lambda.C) plates with an efficiency
of approximately 4 X 10-4 on log phage E. coli K12 grown in broth.  These few
successful infections occur in "special cells" in the K12 population which have
lost their ability to restrict lambda.C.  E. coli K12 grown in minimal medium
has 30-50 times as many special cells.  Supplementation of minimal medium with
an amino acid mixture reduced the frequency of special cells to that observed
in broth cultures.  When amino acids were tested individually, the addition of
either alainine or leucine alone to minimal medium reduced the special cell
population.  This reduction in the frequency of special cells was reversed by
the futher addition of several other amino acids to the growth medium.  Medium
shift experiments with alanine and leucine suggest that these amino acids do
not act at the time of infection; instead they determine the presence of a
metabolic system involved in the expression of host-controlled restriction.
Removal of methionine from K12 cultures containing an otherwise complete amino
acid mixture caused an increase in the number of special cells.  Unlike the
alanine and leucine effects, this increase was rapid and occurred in the
presence of chloramphenicol.  Prolonged growth in methionine was required to
restore the reduced special cell frequency.  It appears that the removal of
methionine can convert a restricting cell into a special cell which is then
incapable of undergoing cell division.  The various amino acid effects are
specific to the K12-restriction system and do not influence the B- or
P1-restriction systems of E. coli.

<>

<1>Grasso, R.J., Paigen, K.
<2>Loss of host-controlled restriction and modification of phage lambda in Escherichia coli K12 previously infected with UV-irradiated coli-phage T3.
<3>Virology
<4>38
<5>191-194
<6>1969
<7>Host range properties of bacteriophages are altered not only by genetic
mutation, but also by the process of host-controlled modification.

<>

<1>Graves, K.L., Butler, M.M., Hardy, L.W.
<2>Site-directed mutagenesis of the gene encoding phage T4 dCMP hydroxymethylase.
<3>FASEB J.
<4>7
<5>A1197
<6>1993
<7>The proposed roles of several amino acid residues in deoxycytidylate (dCMP) hydroxymethylase
(CH) have been tested. CH catalyzes the formation of 5-hydroxymethyl-dCMP, an essential
component of phage T4 DNA, from dCMP and methylenetetrahydrofolate (CH2THF). CH resemble
thymidylate synthase (TS), an enzyme of known crystallographic structure, in both amino acid
sequence and reaction catalyzed. Conversion of Cys148 to Asp, Gly, or Ser decreases CH
activity at least 10/5-fold, consistent with a nucleophilic role for Cys148 (analogous to the
catalytic Cys residue in TS). In TS, hydrogen bonds connect O4 and N3 of the substrate dUMP to
an Asn. Conversion of the corresponding residue in CH, Asp179 to an Asn reverses the substrate
preference of CH from dCMP to dUMP. Asp179 is proposed to stabilize covalent catalytic
intermediates, by protonating N3 of the pyrimidine-CH adduct. CH is inactivated by
5-fluoro-deoxyuridylate (FdUMP), a mechanism-based inactivator of TS, by formation of a
ternary complex between enzyme, CH2THF and FdUMP. Replacement of Cys148 prevents formation of
such a complex. The secondary equilibrium isotope effect upon the formation of the ternary
complex, HK/TK, was measured using a mixture of 2(14C)-FdUMP and 6[3H]-FdUMP. The observed
inverse effect is consistent with the formation of a covalent complex where C6 of FdUMP is sp3
hybridized and covalently linked to the thiol of Cys148. The proposed roles of a number of
other residues in CH are currently under investigation.

<>

<1>Gray, T.A., Palumbo, M.J., Derbyshire, K.M.
<2>Draft Genome Sequence of MKD8, a Conjugal Recipient Mycobacterium smegmatis Strain.
<3>Genome Announcements
<4>1
<5>e00148-13
<6>2013
<7>We report an annotated draft genome sequence of the Mycobacterium smegmatis strain MKD8. This
strain acts as a recipient during conjugation with the
reference M. smegmatis strain mc(2)155. While the genomes of the two strains are
colinear and have similar sizes, extensive genome-wide sequence variation
suggests rich diversity within the M. smegmatis clade.

<>

<1>Grazulis, S., Deibert, M., Rimseliene, R., Skirgaila, R., Sasnauskas, G., Lagunavicius, A., Repin, V., Urbanke, C., Huber, R., Siksnys, V.
<2>Crystal structure of the Bse634I restriction endonuclease: comparison of two enzymes recognizing the same DNA sequence.
<3>Nucleic Acids Res.
<4>30
<5>876-885
<6>2002
<7>Crystal structures of Type II restriction endonucleases demonstrate a conserved common core
and active site residues but diverse structural elements involved in DNA sequence
discrimination. Comparative structural analysis of restriction enzymes recognizing the same
nucleotide sequence might therefore contribute to our understanding of the structural
diversity of specificity determinants within restriction enzymes. We have solved the crystal
structure of the Bacillus stearothermophilus restriction endonuclease Bse634I by the multiple
isomorphous replacement technique to 2.17 Angstroms resolution. Bse634I is an isoschisomer of
the Cfr10I restriction enzyme whose crystal structure has been reported previously.
Comparative structural analysis of the first pair of isoschisomeric enzymes revealed conserved
structural determinants of sequence recognition and catalysis. However, conformations of the
N-terminal subdomains differed between Bse634I/Cfr10I, suggesting a rigid body movement that
might couple DNA recognition and catalysis. Structural similarities extend to the quaternary
structure level: crystal contacts suggest that Bse634I similarly to Cfr10I is arranged as a
tetramer. Kinetic analysis reveals that Bse634I is able to interact simultaneously with two
recognition sites supporting the tetrameric architecture of the protein. Thus, restriction
enzymes Bse634I, Cfr10I and NgoMIV, recognizing overlapping nucleotide sequences, exhibit a
conserved tetrameric architecture that is of functional importance.
* To whom correspondence should be addressed at: Institute of Biotechnolo

<>

<1>Grazulis, S., Manakova, E., Roessle, M., Bochtler, M., Tamulaitiene, G., Huber, R., Siksnys, V.
<2>Structure of the metal-independent restriction enzyme BfiI reveals fusion of a specific DNA-binding domain with a nonspecific nuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>15797-15802
<6>2005
<7>Among all restriction endonucleases known to date, BfiI is unique in cleaving DNA in the
absence of metal ions. BfiI represents a different
evolutionary lineage of restriction enzymes, as shown by its crystal
structure at 1.9-A resolution. The protein consists of two structural
domains. The N-terminal catalytic domain is similar to Nuc, an
EDTA-resistant nuclease from the phospholipase D superfamily. The
C-terminal DNA-binding domain of BfiI exhibits a beta-barrel-like
structure very similar to the effector DNA-binding domain of the
Mg(2+)-dependent restriction enzyme EcoRII and to the B3-like DNA-binding
domain of plant transcription factors. BfiI presumably evolved through
domain fusion of a DNA-recognition element to a nonspecific nuclease akin
to Nuc and elaborated a mechanism to limit DNA cleavage to a single
double-strand break near the specific recognition sequence. The crystal
structure suggests that the interdomain linker may act as an autoinhibitor
controlling BfiI catalytic activity in the absence of a specific DNA
sequence. A psi-blast search identified a BfiI homologue in a
Mesorhizobium sp. BNC1 bacteria strain, a plant symbiont isolated from an
EDTA-rich environment.

<>

<1>Green, N.M., Zhang, S., Porcella, S.F., Nagiec, M.J., Barbian, K.D., Beres, S.B., Lefebvre, R.B., Musser, J.M.
<2>Genome Sequence of a Serotype M28 Strain of Group A Streptococcus: Potential New Insights into Puerperal Sepsis and Bacterial Disease   Specificity.
<3>J. Infect. Dis.
<4>192
<5>760-770
<6>2005
<7>Puerperal sepsis, a major cause of death of young women in Europe in the 1800s, was due
predominantly to the gram-positive pathogen group A
Streptococcus. Studies conducted during past decades have shown that
serotype M28 strains are the major group A Streptococcus organisms
responsible for many of these infections. To begin to increase our
understanding of their enrichment in puerperal sepsis, we sequenced the
genome of a genetically representative strain. This strain has genes
encoding a novel array of prophage virulence factors, cell-surface
proteins, and other molecules likely to contribute to host-pathogen
interactions. Importantly, genes for 7 inferred extracellular proteins are
encoded by a 37.4-kb foreign DNA element that is shared with group B
Streptococcus and is present in all serotype M28 strains. Proteins encoded
by the 37.4-kb element were expressed extracellularly and in human
infections. Acquisition of foreign genes has helped create a
disease-specialist clone of this pathogen.

<>

<1>Greenaway, P.J.
<2>The isolation of a restriction enzyme from Bordetella pertussis.
<3>Biochem. Biophys. Res. Commun.
<4>95
<5>1282-1287
<6>1980
<7>A restriction enzyme was isolated from Bordetella pertussis cells by a
single-step purification procedure using chromatography on phosphocellulose.
Different DNA molecules were digested with this enzyme; the fragmentation
patterns obtained were compared to those obtained after digestion with the
HindIII enzyme isolated from Haemophilus influenzae strain Rd.  It was
concluded that the cleavage site specificities of these enzymes were identical.

<>

<1>Greenberg, D.E., Porcella, S.F., Zelazny, A.M., Virtaneva, K., Sturdevant, D.E., Kupko, J.J., Barbian, K.D., Babar, A., Dorward, D.W., Holland, S.M.
<2>Genome sequence analysis of the emerging human pathogenic acetic acid bacterium Granulibacter bethesdensis.
<3>J. Bacteriol.
<4>189
<5>8727-8736
<6>2007
<7>Chronic granulomatous disease (CGD) is an inherited immune deficiency characterized by
increased susceptibility to infection with
Staphylococcus, certain gram-negative bacteria, and fungi. Granulibacter
bethesdensis, a newly described genus and species within the family
Acetobacteraceae, was recently isolated from four CGD patients residing in
geographically distinct locales who presented with fever and
lymphadenitis. We sequenced the genome of the reference strain of
Granulibacter bethesdensis, which was isolated from lymph nodes of the
original patient. The genome contains 2,708,355 base pairs in a single
circular chromosome, in which 2,437 putative open reading frames (ORFs)
were identified, 1,470 of which share sequence similarity with ORFs in the
nonpathogenic but related Gluconobacter oxydans genome. Included in the
967 ORFs that are unique to G. bethesdensis are ORFs potentially important
for virulence, adherence, DNA uptake, and methanol utilization. GC% values
and best BLAST analysis suggested that some of these unique ORFs were
recently acquired. Comparison of G. bethesdensis to other known CGD
pathogens demonstrated conservation of some putative virulence factors,
suggesting possible common mechanisms involved in pathogenesis in CGD.
Genotyping of the four patient isolates by use of a custom microarray
demonstrated genome-wide variations in regions encoding DNA uptake systems
and transcriptional regulators and in hypothetical ORFs. G. bethesdensis
is a genetically diverse emerging human pathogen that may have recently
acquired virulence factors new to this family of organisms.

<>

<1>Greene, P., Reich, N.O., McClarin, J., Boyer, H.W., Rosenberg, J.
<2>Site-directed mutagenesis of the EcoRI endonuclease.
<3>J. Cell Biochem. Suppl.
<4>9B
<5>94
<6>1985
<7>The EcoRI endonuclease has been crystallized with its substrate and the X-ray
structure has been solved to 3 angstroms (Nature 309: 327, 1984).  Sequence
recognition is mediated by a series of hydrogen bonds between complementary
surfaces of the protein and the major groove of the DNA.  We have used the
crystallographic analysis to design putative specificity mutations in the
endonuclease and to predict possible new specificities that might arise.  In
particular, a glutamic acid residue which is postulated to interact with the N6
positions of the adjacent adenines in the recognition sequence has been
replaced with glutamic acid.  The predicted recognition sequence would contain
a guanine in place of one of the adenines.  This alteration has been introduced
into the endonuclease gene by means of mismatch primer mutagenesis.  The
expression and characterization of the mutant protein is underway.

<>

<1>Greene, P.J., Ballard, B.T., Stephenson, F., Kohr, W.J., Rodriguez, H., Rosenberg, J.M., Boyer, H.W.
<2>Purification and characterization of the restriction endonuclease RsrI, an isoschizomer of EcoRI.
<3>Gene
<4>68
<5>43-52
<6>1988
<7>Rhodobacter sphaeroides strain 630 produces restriction enzyme RsrI which is an
isoschizoemr of EcoRI.  We have purified this enzyme and initiated a comparison
with the EcoRI endonuclease.  The properties of RsrI are consistent with a
reaction mechanism similar to that of EcoRI:  the position of cleavage within
the -GAATTC- site is identical, the MgCl2 optimum for the cleavage is
identical, and the pH profile is similar.  Methylation of the substrate
sequence by the EcoRI methylase protects the site from cleavage by the RsrI
endonuclease.  RsrI cross-reacts strongly with anti-EcoRI serum indicating
three-dimensional structural similarities.  We have determined the sequence of
34 N-terminal amino acids for RsrI and this sequence possesses significant
similarity to the EcoRI N terminus.

<>

<1>Greene, P.J., Betlach, M.C., Boyer, H.W., Goodman, H.M.
<2>The EcoRI restriction endonuclease.
<3>Methods Mol. Biol.
<4>7
<5>87-105
<6>1974
<7>Type II bacterial restriction endonucleases have become increasingly useful in
the analysis of small DNA genomes, while attempts to use the Type I restriction
endonucleases have not been satisfactory because they appear to be less
specific than originally imagined.  The current purification procedure for the
EcoRI restriction endonuclease and some of its properties, as well as a
procedure for partial purification of the related EcoRI modification methylase,
are described.

<>

<1>Greene, P.J., Boyer, H.W.
<2>Genetic and molecular analysis of the EcoRI restriction and modification system.
<3>Fed. Proc.
<4>40
<5>293
<6>1981
<7>The structure and function of the EcoRI endonuclease and methylase are being
studied in order to elucidate how proteins recognize specific DNA sequences.
We have crystallized the EcoRI endonuclease and measured space group and unit
cell parameters.  We plan to sequence the endonuclease and methylase by
sequencing the DNA which encodes them.  The genes are closely linked on a
multicopy plasmid, pMB1.  The approximate boundaries of the genes have been
determined by cloning different restriction fragments.  Based on subunit
molecular weight estimates of 36,000 daltons and 29,000 daltons for the
methylase and endonuclease respectively, the required length of DNA for both
structural genes is ~ 1800 base pairs.  A 2300 base pair restriction fragment
has been shown to contain all of the information necessary for the expression
of normal levels of both enzymes.  A detailed restriction map of this fragment
has been prepared using the technique of end labeling followed by partial
digestion.  DNA sequencing is underway using chain terminating inhibitors.
Clones have been isolated with this fragment in both orientations relative to
the lac promoter and operator in pBH20.  The effect of the lac control system
on the specific activity of EcoRI enzymes suggests that the direction of
transcription is from the endonuclease to the methylase.

<>

<1>Greene, P.J., Gupta, M., Boyer, H.W., Brown, W.E., Rosenberg, J.M.
<2>Sequence Analysis of the DNA Encoding the EcoRI Endonuclease and Methylase.
<3>J. Biol. Chem.
<4>256
<5>2143-2153
<6>1981
<7>The EcoRI endonuclease and methylase recognize the same hexanucleotide substrate sequence.  We
have determined the sequence of a fragment of DNA which encodes these enzymes using the
chain-termination method of Sanger (Sanger, F., Nicklen, S., and Coulson, A.R. (1977) Proc.
Natl. Acad. Sci. U.S.A. 74, 5463-5467).  The amino acid sequences of both enzymes were derived
from the DNA sequence.  The coding regions selected include the only open translational frames
of sufficient length to accommodate the enzymes.  They coincide with previously established
gene boundaries and orientation.  The predicted amino acid sequences correlate well with
analyses of the purified protein. Comparison of the nucleotide and protein sequences reveals
no homology between the endonuclease and methylase which might provide insight into the origin
of the restriction-modification system or the mechanism of common substrate recognition.
Based on secondary structure predictions, the two enzymes also have grossly different
molecular architecture.  The base composition of the sequence is 65% A + T, and the codon
usage is signficantly different from that observed in several Escherichia coli chromosomal
genes.  In some cases, frequently selected codons are recognized by minor tRNA species.  A
spontaneous mutation in the endonuclease gene was isolated.  Serine replaces arginine at
residue 187.  In crude extracts, EcoRI specific cleavage is ~0.3% wild type.

<>

<1>Greene, P.J., Heyneker, H.L., Bolivar, F., Rodriguez, R.L., Betlach, M.C., Covarrubias, A.A., Backman, K., Russel, D.J., Tait, R., Boyer, H.W.
<2>A general method for the purification of restriction enzymes.
<3>Nucleic Acids Res.
<4>5
<5>2373-2380
<6>1978
<7>An abbreviated procedure has been developed for the purification of restriction
endonucleases.  This procedure uses chromatography on phosphocellulose and
hydroxylapatite and results in enzymes of sufficient purity to permit their use
in the sequencing, molecular cloning, and physical mapping of DNA.

<>

<1>Greene, P.J., Poonian, M.S., Nussbaum, A.L., Tobias, L., Garfin, D.E., Boyer, H.W., Goodman, H.M.
<2>Restriction and modification of a self-complementary octanucleotide containing the EcoRI substrate.
<3>J. Mol. Biol.
<4>99
<5>237-261
<6>1975
<7>In order to study the interactions of the EcoRI restriction endonuclease and
modification methylase with DNA, we have synthesized the self-complementary
octanucleotide, pT-G-A-A-T-T-C-A, which contains the nucleotide sequence of the
EcoRI substrate.  This octamer can act as a substrate for both the endonuclease
and methylase; the enzymatic alteration of this molecule is the same as that of
DNA, with cleavage and methylation both occurring at the same positions on
either substrate.  The optimum temperature for the reaction of the
octanucleotide with the endonuclease is 15C and that with the methylase is
12.5C.  The optimum temperature for the reactions of both of the enzymes with
DNA is 37C.  The low temperatures for reactions with the octanucleotide reflect
the conditions under which this molecule can serve as a substrate for the
enzymes.

<>

<1>Greene, P.J., Yanofsky, S., Reich, N., Day, J., Hager, P., Boyer, H.W.
<2>Structure and function of the EcoRI endonuclease.
<3>Protein Structure, Folding and Design 2, Alan R. Liss, Inc., Oxender, D.L., NY
<4>0
<5>3-7
<6>1987
<7>None

<>

<1>Gregoire, S.T.
<2>The effect of DNA-mediated restriction on conjugation in Neisseria gonorrhoeae.
<3>M.Sc. Thesis. Univ. Maryland.
<4>0
<5>1-138
<6>1986
<7>A derivative of the gonococcal shuttle-vector pLES2 containing the proAB genes of Neisseria
gonorrhoeae strain KH45 was mobilized by the gonococcal conjugal plasmid into Escherichia coli
recipients independent of the host strain restriction-modification phenotype.  Introduction of
this plasmid into E. coli via conjugation does not result in deletions.  Expression of the
proline biosynthesis genes was confirmed by cultivation of transconjugants on media devoid of
proline.  The plasmid was mobilized to various gonococcal strains.  Transfer frequencies of
10^-5 were obtained.  When isogenic crosses between the recipient D13 strain, and D13
containing pLES4, were compared, transfer occurred at 10^-2.  The role of restriction during
conjugation was investigated as an explanation for the increase in transfer efficiency.  The
self-mobilizable plasmid pFT6, propagated in an R.NgoII-, M.NgoII- gonococcal strain, was used
in filter matings with R.NgoII- M.NgoII+, R.NgoII+ M.NgoII+, and R.NgoII- M.NgoII- recipient
strains.  The results demonstrated that the plasmid mobilized to the gonococcal recipients at
equal frequencies, indicating that restriction was not a barrier to conjugation.  Introduction
of pLES4 into commensal species of the Neisseriaceae via conjugation was unsuccessful.  A
mechanism other than restriction is presented as a possible factor influencing conjugation in
the gonococcus.

<>

<1>Gregorova, D., Pravcova, M., Karpiskova, R., Rychlik, I.
<2>Plasmid pC present in Salmonella enterica serovar Enteritidis PT14b strains encodes a restriction modification system.
<3>FEMS Microbiol. Lett.
<4>214
<5>195-198
<6>2002
<7>Salmonella enterica serovar Enteritidis (S. enteritidis) possesses plasmids of different sizes
and roles. Besides the serovar-specific
virulence plasmid present in most field strains, S. enteritidis can
harbour plasmids of low molecular mass whose biological role is poorly
understood. We therefore sequenced plasmid pC present in S. enteritidis
strains belonging to phage type PT14b. The size of plasmid was
determined to be 5269 bp and it was predicted to encode four open
reading frames (ORFs). The first two ORFs were found (initial 3230 bp)
to be highly homologous to rom and mbeA genes of Co1E1 plasmid of
Escherichia coli. Proteins encoded by the other two ORFs were 99%
homologous to a restriction methylase and restriction endonuclease
encoded by plasmid pECO29 of a field strain of E. coli. Using
insertional mutagenesis we confirmed experimentally that the plasmid
pC-encoded restriction modification system was functional and could
explain the high resistance of S. enteritidis PT14b strains to phage
infection.

<>

<1>Gregory, R., Saunders, V.A., Saunders, J.R.
<2>Rule-based simulation of temperate bacteriophage infection: Restriction-modification as a limiter to infection in bacterial populations.
<3>Biosystems
<4>100
<5>166-177
<6>2010
<7>An individual-based model (IbM) for bacterial adaptation and evolution, COSMIC-Rules, has been
employed to simulate interactions of virtual
temperate bacteriophages (phages) and their bacterial hosts. Outcomes
of infection mimic those of a phage such as lambda, which can enter
either the lytic or lysogenic cycle, depending on the nutritional
status of the host. Infection of different hosts possessing differing
restriction and modification systems is also simulated. Phages
restricted upon infection of one restricting host can be adapted (by
host-controlled modification of the phage genome) and subsequently
propagate with full efficiency on this host. However, such ability is
lost if the progeny phages are passaged through a new host with a
different restriction and modification system before attempted
re-infection of the original restrictive host. The simulations show
that adaptation and re-adaptation to a particular host-controlled
restriction and modification system result in lower efficiency and
delayed lysis of bacterial cells compared with infection of
non-restricting host bacteria.
Such biologically realistic simulations validate the use of the IbM
approach to predicting behaviour of bacteriophages in bacterial
populations. The applicability of the model for more complex scenarios
aimed at predictive modelling of bacterial evolution in a changing
environment and the implications for the spread of viruses in a wider
context are discussed.

<>

<1>Greif, G., Iraola, G., Berna, L., Coitinho, C., Rivas, C.M., Naya, H., Robello, C.
<2>Complete Genome Sequence of Mycobacterium tuberculosis Strain MtURU-001, Isolated from a Rapidly Progressing Outbreak in Uruguay.
<3>Genome Announcements
<4>2
<5>e01220-13
<6>2014
<7>Despite efficient control programs, large clonal outbreaks of tuberculosis (TB) may arise in
low-risk populations. Recently, an unusual TB outbreak was reported
in Uruguay, reaching an elevated disease attack rate (53 to 69%). Here, we report
the genome sequence of the Mycobacterium tuberculosis strain associated with this
rapidly progressing outbreak, named MtURU-001.

<>

<1>Greil, F., Moorman, C., van Steensel, B.
<2>DamID: Mapping of in vivo protein-genome interactions using tethered DNA adenine methyltransferase.
<3>Methods Enzymol.
<4>410
<5>342-359
<6>2006
<7>A large variety of proteins bind to specific parts of the genome to regulate gene expression,
DNA replication, and chromatin structure.
DamID is a powerful method used to map the genomic interaction sites of
these proteins in vivo. It is based on fusing a protein of interest to
Escherichia coli DNA adenine methyltransferase (dam). Expression of
this fusion protein in vivo leads to preferential methylation of
adenines in DNA surrounding the native binding sites of the dam fusion
partner. Because adenine methylation does not occur endogenously in
most eukaryotes, it provides a unique tag to mark protein interaction
sites. The adenine-methylated DNA fragments are isolated by selective
polymerase chain reaction amplification and can be identified by
microarray hybridization. We and others have successfully applied DamID
to the genome-wide identification of interaction sites of several
transcription factors and other chromatin-associated proteins. This
chapter discusses DamID technology in detail, and a step-by-step
experimental protocol is provided for use in Drosophila cell lines.

<>

<1>Grenier, F., Matteau, D., Baby, V., Rodrigue, S.
<2>Complete Genome Sequence of Escherichia coli BW25113.
<3>Genome Announcements
<4>2
<5>e01038-14
<6>2014
<7>Escherichia coli BW25113 is the parent strain of the Keio collection comprising nearly 4,000
single-gene deletion mutants. We report the complete 4,631,469-bp
genome sequence of this strain and the key variations from the type strain E.
coli MG1655.

<>

<1>Grepinet, O., Boumart, Z., Virlogeux-Payant, I., Loux, V., Chiapello, H., Gendrault, A., Gibrat, J.F., Chemaly, M., Velge, P.
<2>Genome Sequence of the Persistent Salmonella enterica subsp. enterica Serotype Senftenberg Strain SS209.
<3>J. Bacteriol.
<4>194
<5>2385-2386
<6>2012
<7>Salmonella enterica subsp. enterica serotype Senftenberg is an emerging serotype  in poultry
production which has been found to persist in animals and the farm
environment. We report the genome sequence and annotation of the SS209 strain of
S. Senftenberg, isolated from a hatchery, which was identified as persistent in
broiler chickens.

<>

<1>Grepinet, O., Rossignol, A., Loux, V., Chiapello, H., Gendrault, A., Gibrat, J.F., Velge, P., Virlogeux-Payant, I.
<2>Genome Sequence of the Invasive Salmonella enterica subsp. enterica Serotype Enteritidis Strain LA5.
<3>J. Bacteriol.
<4>194
<5>2387-2388
<6>2012
<7>Salmonella enterica subsp. enterica serotype Enteritidis is one of the major causes of
gastroenteritis in humans due to consumption of poultry derivatives.
Here we report the whole-genome sequence and annotation, including the virulence
plasmid, of S. Enteritidis LA5, which is a chicken isolate used by numerous
laboratories in virulence studies.

<>

<1>Gressmann, H., Linz, B., Ghai, R., Pleissner, K.P., Schlapbach, R., Yamaoka, Y., Kraft, C., Suerbaum, S., Meyer, T.F., Achtman, M.
<2>Gain and loss of multiple genes during the evolution of Helicobacter pylori.
<3>PLoS Genet.
<4>1
<5>e43
<6>2005
<7>Sequence diversity and gene content distinguish most isolates of Helicobacter pylori. Even
greater sequence differences differentiate distinct populations of H. pylori from different
continents, but it was not clear whether these populations also differ in gene content. To
address this question, we tested 56 globally representative strains of H. pylori and four
strains of Helicobacter acinonychis with whole genome microarrays. Of the weighted average of
1,531 genes present in the two sequenced genomes, 25% are absent in at least one strain of H.
pylori and 21% were absent or variable in H. acinonychis. We extrapolate that the core genome
present in all isolates of H. pylori contains 1,111 genes. Variable genes tend to be small and
possess unusual GC content; many of them have probably been imported by horizontal gene
transfer. Phylogenetic trees based on the microarray data differ from those based on sequences
of seven genes from the core genome. These discrepancies are due to homoplasies resulting from
independent gene loss by deletion or recombination in multiple strains, which distort
phylogenetic patterns. The patterns of these discrepancies versus population structure allow a
reconstruction of the timing of the acquisition of variable genes within this species.
Variable genes that are located within the cag pathogenicity island were apparently first
acquired en bloc after speciation. In contrast, most other variable genes are of unknown
function or encode restriction/modification enzymes, transposases, or outer membrane proteins.
These seem to have been acquired prior to speciation of H. pylori and were subsequently lost
by convergent evolution within individual strains. Thus, the use of microarrays can reveal
patterns of gene gain or loss when examined within a phylogenetic context that is based on
sequences of core genes.

<>

<1>Greub, G., Kebbi-Beghdadi, C., Bertelli, C., Collyn, F., Riederer, B.M., Yersin, C., Croxatto, A., Raoult, D.
<2>High throughput sequencing and proteomics to identify immunogenic proteins of a new pathogen: the dirty genome approach.
<3>PLoS ONE
<4>4
<5>E8423
<6>2009
<7>BACKGROUND: With the availability of new generation sequencing
technologies, bacterial genome projects have undergone a major boost.
Still, chromosome completion needs a costly and time-consuming gap
closure, especially when containing highly repetitive elements. However,
incomplete genome data may be sufficiently informative to derive the
pursued information. For emerging pathogens, i.e. newly identified
pathogens, lack of release of genome data during gap closure stage is
clearly medically counterproductive. METHODS/PRINCIPAL FINDINGS: We thus
investigated the feasibility of a dirty genome approach, i.e. the release
of unfinished genome sequences to develop serological diagnostic tools. We
showed that almost the whole genome sequence of the emerging pathogen
Parachlamydia acanthamoebae was retrieved even with relatively short reads
from Genome Sequencer 20 and Solexa. The bacterial proteome was analyzed
to select immunogenic proteins, which were then expressed and used to
elaborate the first steps of an ELISA. CONCLUSIONS/SIGNIFICANCE: This work
constitutes the proof of principle for a dirty genome approach, i.e. the
use of unfinished genome sequences of pathogenic bacteria, coupled with
proteomics to rapidly identify new immunogenic proteins useful to develop
in the future specific diagnostic tests such as ELISA,
immunohistochemistry and direct antigen detection. Although applied here
to an emerging pathogen, this combined dirty genome sequencing/proteomic
approach may be used for any pathogen for which better diagnostics are
needed. These genome sequences may also be very useful to develop DNA
based diagnostic tests. All these diagnostic tools will allow further
evaluations of the pathogenic potential of this obligate intracellular
bacterium.

<>

<1>Grewal, S., Vakhlu, J., Gupta, V., Sangwan, N., Kohli, P., Nayyar, N., Rani, P., Sance, S.S., Lal, R.
<2>Draft Genome Sequence of Pseudomonas sp. Strain JMM, a Sediment-Hosted Environmental Isolate.
<3>Genome Announcements
<4>2
<5>e00879-14
<6>2014
<7>Pseudomonas sp. strain JMM was isolated from the sediments of a natural water reservoir (pH, 6
to 7) located at Chambyal village in Samba district of Jammu and
Kashmir, India. Here we report the annotated draft genome sequence of strain JMM
having 52 contigs with 5,884 genes and an average G+C content of 66.5%.

<>

<1>Griffin, S., Branch, P., Xu, Y.-Z., Karran, P.
<2>DNA mismatch binding and incision at modified guanine bases by extracts of mammalian cells: implications for tolerance to DNA methylation damage.
<3>Biochemistry
<4>33
<5>4787-4793
<6>1994
<7>Two activities involved in separate pathways for correcting G.T mispairs in DNA have been
assayed on duplex substrates containing modified guanine bases. The first, the G.T mismatch
incision activity, is specifically involved in short-patch repair of mispairs arising via
deamination of 5-methylcytosine. The second activity can be detected by its ability to bind to
G.T mispairs and may initiate correction by a long-patch mechanism. 6-Thioguanine and
06-methylguanine paired with thymine were efficiently incised by cell extracts if the modified
guanine was in a CpG dinucleotide. Incision was not observed when either purine was paired
with cytosine. Extracts of cells that are tolerant both to methylation damage and to
6-thioguanine in DNA also incised 6-thioguanine.T and O6-methylguanine.T base pairs. The data
suggest that this activity is unlikely to contribute significantly to the biological effects
of 06-methylguanine in DNA. A defect in this pathway is therefore unlikely to explain the
cross-resistance of tolerant cells to the two base analogs in DNA. In binding assays,
6-thioguanine-T base pairs were recognized efficiently and to an equivalent extent by the same
protein complex as G.T mispairs. O6-methylguanine.T base pairs were also recognized but with
reduced efficiency. No binding was observed to 6-thioguanine.C or O6-methylguanine.C base
pairs. Recognition by the binding complex was essentially independent of the base immediately
5' to the mismatched guanine but was somewhat more efficient if O6-methylguanine was preceded
by a purine. Extracts of two tolerant lines with a known defect in G.T mismatch binding failed
to form complexes with substrates containing the modified bases. The ability of the G.T
binding factor to recognize both O6-methylguanine.T and 6-thioguanine.T pairs indicates that
the long-patch repair pathway is more likely to be involved in mediating the cytotoxicity of
the two-base analogs.

<>

<1>Griffiths, A., Tawfik, D.
<2>In vitro sorting method.
<3>International Patent Office
<4>WO 9902671
<5>
<6>1999
<7>The invention describes a method for isolating one or more genetic elements encoding a gene
product having a desired activity, comprising the steps of: (a) compartmentalizing genetic
elements into microcapsules; (b) expressing the genetic elements to produce their respective
gene products within the microcapsules; (c) sorting the genetic elements which produce the
gene product having a desired activity.  The invention enables the in vitro evolution of
nucleic acids by repeated mutagenesis and iterative applications of the method of the
invention.

<>

<1>Grigaite, R., Maneliene, Z., Janulaitis, A.
<2>AarI, a restriction endonuclease from Arthrobacter aurescens SS2-322, which recognizes the novel non-palindromic sequence 5'-CACCTGC(N)4/8-3'.
<3>Nucleic Acids Res.
<4>30
<5>e123
<6>2002
<7>A new type II restriction endonuclease AarI has been isolated from Arthrobacter aurescens
SS2-322. AarI recognizes the non-palindromic heptanucleotide sequence 5'-CACCTGC(N)(4/8)-3'
and makes a staggered cut at the fourth and eighth bases downstream of the target duplex
producing a four base 5'-protruding end. AarI activity is stimulated by
oligodeoxyribonucleotide duplexes containing an enzyme-specific recognition sequence.

<>

<1>Grigorescu, A., Horvath, M., Wilkosz, P.A., Chandrasekhar, K., Rosenberg, J.M.
<2>The integration of recognition and cleavage: X-ray structures of pre-transition state complex, post-reactive complex, and the DNA-free endonuclease.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>137-177
<6>2004
<7>DNA has been selected as the biological information storage molecule for many reasons; one of
which is its stability.  The rate of spontaneous hydrolysis of DNA (at 24oC and pH 7.4) was
estimated at 5.7x10-14S-1; more recently, Radzicka and Wolfenden have estimated this value at
1.7x10-13 s-1.  This corresponds to an estimated half-life of 130,000 years for a DNA
phosphodiester bond in solution, placing DNA hydrolysis among the slowest of biochemical
reactions in the absence of enzymes.

<>

<1>Grigorescu, A.A., Rosenberg, J.M.
<2>A local folding transition coupled to site-specific binding in EcoRI restriction endonuclease.
<3>Biophys. J.
<4>86
<5>591a
<6>2004
<7>Restriction endonuclease EcoRI is not only an invaluable tool in genetic engineering but also
a paradigm for understanding the molecular
mechanisms of protein-DNA recognition. Recent crystallographic data
demonstrate that a major localized folding transition accompanies
binding of the EcoRI restriction endonuclease to its cognate DNA site;
this result is in accordance with previous biophysical studies which
indicated a large conformational change taking place in the protein
upon site-specific binding. The three-dimensional structure of the apo
EcoRI enzyme reveals an unusual molecular architecture: a rigid
scaffold with a small minidomain on top of it. In the absence of
stabilizing inter-molecular contacts the minidomain does not adopt a
well-ordered conformation. A structural analysis indicates that the
metastability of this region of the protein is dictated by a
combination of favorable and unfavorable tertiary interactions. The
conformation of the minidomain observed in the specific protein-DNA
complex is apparently stabilized by a set of favorable inter-molecular
contacts. Many of these are highly specific contacts (recognition
interactions) between the protein and the DNA substrate. All the amino
acid substitutions that have been shown to alter (i.e. reduce) the DNA
sequence specificity of the enzyme are localized in regions that
exhibit low structural stability in the unbound protein. This, and
other lines of evidence suggest that local native metastability is
important for the molecular recognition function of the EcoRI
endonuclease.

<>

<1>Grigoryeva, T.V., Laikov, A.V., Naumova, R.P., Manolov, A.I., Larin, A.K., Karpova, I.Y., Semashko, T.A., Alexeev, D.G., Kostryukova, E.S., Muller, R., Govorun, V.M.
<2>Draft Genome of the Nitrogen-Fixing Bacterium Pseudomonas stutzeri Strain KOS6 Isolated from Industrial Hydrocarbon Sludge.
<3>Genome Announcements
<4>1
<5>e00072-12
<6>2013
<7>Here we present a draft genome of Pseudomonas stutzeri strain KOS6. This strain was isolated
from industrial hydrocarbon sludge as a diazotrophic microorganism.
It represents one of the major parts of the culturable community of the waste and
has potential importance for phytoremediation technology.

<>

<1>Grim, C.J., Gopinath, G.R., Mammel, M.K., Sathyamoorthy, V., Trach, L.H., Chase, H.R., Tall, B.D., Fanning, S., Stephan, R.
<2>Genome Sequence of an Enterobacter helveticus Strain, 1159/04 (LMG 23733), Isolated from Fruit Powder.
<3>Genome Announcements
<4>1
<5>e01038-13
<6>2013
<7>We report the draft genome sequence of Enterobacter helveticus strain LMG 23733,  isolated
from fruit powder. The draft genome assembly for E. helveticus strain
LMG 23733 has a size of 4,635,476 bp and a G+C content of 55.9%.

<>

<1>Grim, C.J., Hasan, N.A., Taviani, E., Haley, B., Chun, J., Brettin, T.S., Bruce, D.C., Detter, J.C., Han, C.S., Chertkov, O., Challacombe, J., Huq, A., Nair, G.B., Colwell, R.R.
<2>Genome Sequence of Hybrid Vibrio cholerae O1 MJ-1236, B-33, and CIRS101 and Comparative Genomics with V. cholerae.
<3>J. Bacteriol.
<4>192
<5>3524-3533
<6>2010
<7>The genomes of Vibrio cholerae O1 Matlab variant MJ-1236, Mozambique O1 El Tor variant B33,
and altered O1 El Tor CIRS101 were sequenced. All three
strains were found to belong to the phylocore group 1 clade of V.
cholerae, which includes the 7th-pandemic O1 El Tor and serogroup O139
isolates, despite displaying certain characteristics of the classical
biotype. All three strains were found to harbor a hybrid variant of CTXPhi
and an integrative conjugative element (ICE), leading to their
establishment as successful clinical clones and the displacement of
prototypical O1 El Tor. The absence of strain- and group-specific genomic
islands, some of which appear to be prophages and phage-like elements,
seems to be the most likely factor in the recent establishment of
dominance of V. cholerae CIRS101 over the other two hybrid strains.

<>

<1>Grimaud, R., Ghiglione, J.F., Cagnon, C., Lauga, B., Vaysse, P.J., Rodriguez-Blanco, A., Mangenot, S., Cruveiller, S., Barbe, V., Duran, R., Wu, L.F., Talla, E., Bonin, P., Michotey, V.
<2>Genome Sequence of the Marine Bacterium Marinobacter hydrocarbonoclasticus SP17,  Which Forms Biofilms on Hydrophobic Organic Compounds.
<3>J. Bacteriol.
<4>194
<5>3539-3540
<6>2012
<7>Marinobacter hydrocarbonoclasticus SP17 forms biofilms specifically at the interface between
water and hydrophobic organic compounds (HOCs) that are used as
carbon and energy sources. Biofilm formation at the HOC-water interface has been
recognized as a strategy to overcome the low availability of these nearly
water-insoluble substrates. Here, we present the genome sequence of SP17, which
could provide further insights into the mechanisms of enhancement of HOCs
assimilation through biofilm formation.

<>

<1>Grime, S.K., Martin, R.L., Holaway, B.L.
<2>Inhibition of restriction enzyme cleavage of DNA modified with 7-deaza-dGTP.
<3>Nucleic Acids Res.
<4>19
<5>2791
<6>1991
<7>DNA templates containing G+C rich regions or stable secondary structures can
potentially pose problems for PCR amplification.  Incorporation of the
structure destabilizing analogue, 7-deaza-dGTP into the PCR reaction allows
successful amplification of such template sequences.  It has been reported that
when 7-deaza-dGTP is substituted for dGTP in a DNA-fragment, the cleavage rate
by the restriction enzyme EcoRI is reduced.  We find that EcoRI as well as
other commonly used restriction enzymes will not cut DNA containing
7-deaza-dGTP when it is incorporated into PCR product.  Substrates for the
restriction enzymes were amplified by PCR using pUC19 and bacteriophage lambda
sequences.  Each 100 microliter reaction contained 200 microM each of dATP,
dCTP, dTTP, dGTP or 7-deaza-dGTP, 1 microM of each primer, 2.5 units Taq DNA
Polymerase, 1 ng DNA template, 50 mM KCl, 10 mM Tris-HCl, pH 8.3 @ 37C, 1.5 mM
MgCl2, 0.01% gelatin.  Five hundred nanograms of the PCR product from reactions
containing either 7-deaza-dGTP or dGTP were incubated with commonly-used
restriction enzymes.  Restriction digests were resolved on 1.2% agarose gels
with ethidium bromide staining.  Table 1 summarizes the results of the
restriction digests where 7-deaza-dGTP was substituted for dGTP in the PCR
reaction.  For most enzymes tested, inclusion of 7-deaza-dGTP in the PCR
product prevented the restriction enzyme from cutting the DNA.  It is not yet
clear to us how this inhibition occurs.  We assume that the presence of the
deaza residue in the recognition sequence affects proper binding and/or cutting
of the DNA substrate.  As yet the details of the spatial interaction between
the modified G residue and various restriction enzyme active/binding sites is
unclear.  Inhibition of cutting can occur when the G residues are 5 prime (eg.
EcoRI), 3 prime (eg. PstI), immediately adjacent (eg. SalI), or two bases away
from the cut site (eg. AccI).  Three enzymes (XbaI, HindIII, MaeII) are not
inhibited by the presence of modified G residues in their recognition sequence.
We have no knowledge of how modified G residues may inhibit cutting from a
complementary strand of DNA.  7-deaza-dGTP could be beneficial for applications
such as cDNA cloning where multiple internal restriction sites are protected
during linker digestion. This would be similar to the strategy of methylating
cDNA prior to digestion with methyl-sensitive restriction enzymes.  Use of
7-deaza-dGTP to this end may allow the use of a greater number of enzymes.

<>

<1>Grimes, E., Koob, M., Szybalski, W.
<2>Achilles' heel cleavage: creation of rare restriction sites in lambda phage genomes and evaluation of additional operators, repressors and restriction/modification systems .
<3>Gene
<4>90
<5>1-7
<6>1990
<7>A novel technique for the creation of rare restriction sites was described by Koob et al.
[Science 241 (1988) 1084-1086]. This technique, Achilles' heel cleavage (AC), relies on the
use of a bound repressor molecule to protect only one of many identical restriction sites from
a modification methyltransferase that inactivates all other restriction sites. The technique
was applied to a small plasmid and shown to work efficiently with two repressor/operator
systems: lac repressor/lacO operator and lambda repressor/lambda OL1 operator. Here, we have
extended these results to a lac operator carried by a much larger vector, namely a 44-kb phage
lambda construct. In addition, we have evaluated the effect of altering the stability of the
lac repressor/lac operator complex by varying both the operator and the repressor. We have
also evaluated several more restriction/modification systems (MboI, Dam, MspI and AluI) in
addition to HhaI and HaeII used earlier. Finally, we extended the AC technique to a third
system, that of the phage 434 repressor and a synthetic 434 operator. From our results we
conclude that the ACT method should be applicable to the mapping of large genomes and to
measuring the strength of operator-repressor interactions. AC could also be applied to
identifying and evaluating many different DNA-binding proteins and their sites of action.

<>

<1>Grindl, W., Wende, W., Pingoud, V., Pingoud, A.
<2>The protein splicing domain of the homing endonuclease PI-SceI is responsible for specific DNA binding.
<3>Nucleic Acids Res.
<4>26
<5>1857-1862
<6>1998
<7>The homing endonuclease PI-SceI consists of a protein splicing domain (I) and an
endonucleolytic domain (II).  To characterize the two domains with respect to their
contribution to DNA recognition we cloned, purified and characterized the isolated domains.
Both domains have no detectable endonucleolytic activity.  Domain I binds specifically to the
PI-SceI recognition sequence, whereas domain II displays only weak non-specific DNA binding.
In the specific complex with domain I the DNA is bent to a similar extent as observed with the
initial complex formed between PI-SceI and DNA.  Our results indicate that protein splicing
domain I is also involved in recognition of the DNA substrate.

<>

<1>Grishin, A., Fonfara, I., Alexeevski, A., Spirin, S., Zanegina, O., Karyagina, A., Alexeyevsky, D., Wende, W.
<2>IDENTIFICATION OF CONSERVED FEATURES OF LAGLIDADG HOMING ENDONUCLEASES.
<3>J. Bioinform. Comput. Biol.
<4>8
<5>453-469
<6>2010
<7>LAGLIDADG family of homing endonucleases are rare-cutting enzymes which recognize long target
sequences and are of great interest in genome
engineering. Despite advances in homing endonuclease engineering,
effective methods of broadening the range of cleaved sequences are
still lacking. Here, we present a study of conserved structural
features of LAGLIDADG homing endonucleases that might aid further
development of such methods. The protein-DNA interface of LAGLIDADG
homing endonucleases differs considerably with the particular nuclease,
and the analysis of conserved protein-DNA interactions could not
identify any residues crucial for DNA binding and common to most
nucleases of the family. For the homing endonuclease PI-SceI, a
comparison of structural and experimental data derived from literature
helped to identify 23 residues that are likely to be important for DNA
binding. Analysis of the LAGLIDADG domain dimerization interface
allowed the choosing of six positions that contribute to dimerization
specificity most, while comparison of 446 sequences of LAGLIDADG
endonucleases revealed groups of residues in these positions that
appear to be most favorable for dimerization.

<>

<1>Gritsenko, O.M., Koudan, E.V., Mikhailov, S.N., Ermolinsky, B.S., VanAerschot, A., Herdewijn, P., Gromova, E.S.
<2>Affinity modification of EcoRII DNA methyltransferase by the dialdehyde-substituted DNA duplexes: mapping the enzyme region that interacts with DNA.
<3>Nucleosides Nucleotides Nucleic Acids
<4>21
<5>753-764
<6>2002
<7>Affinity modification of EcoRII DNA methyltransferase (M.EcoRII) by DNA duplexes containing
oxidized 2'-O-beta-D-ribofuranosylcytidine (Crib*) or 1-(beta-D-galactopyranosyl)thymine
(Tgal*) residues was performed. Cross-linking yields do not change irrespective of whether
active Crib* replaces an outer or an inner (target) deoxycytidine within the EcoRII
recognition site. Chemical hydrolysis of M.EcoRII in the covalent cross-linked complex with
the Tgal*-substituted DNA indicates the region Gly268-Met391 of the methylase that is likely
to interact with the DNA sugar-phosphate backbone. Both specific and non-specific DNA interact
with the same M.EcoRII region. Our results support the theoretically predicted DNA binding
region of M.EcoRII.

<>

<1>Gritsenko, O.M., Mikhailov, S.N., Efimtseva, E.V., Van Aerschot, A., Herdewijn, P., Gromova, E.S.
<2>Probing the MvaI methyltransferase region that interacts with DNA: Affinity labeling with the dialdehyde-containing DNA duplexes.
<3>Nucleosides Nucleotides Nucleic Acids
<4>19
<5>1805-1820
<6>2000
<7>Affinity labeling of methyltransferase MvaI by DNA duplexes containing oxidized
2'-O-beta-D-ribofuranosylcytidine or 1-(beta-D-galactopyranosyl)thymine residues was
performed. Partial chemical hydrolysis of the covalently bound methylase in the conjugates
with the dialdehyde-containing DNA allowed us to determine the amino acid region in the C
terminus of methylase MvaI that interacts with DNA.

<>

<1>Grivell, L.A.
<2>Transposition: Mobile introns get into line.
<3>Curr. Biol.
<4>6
<5>48-51
<6>1996
<7>Group II introns encode highly structured, frequently self-splicing RNAs; they are also mobile
genetic elements.  This mobility has been found to involve DNA-primed reverse transcription,
with similarities to retrotransposition and telomere maintenance.

<>

<1>Grivell, L.A.
<2>Homing in on an endosymbiotic endonuclease.
<3>Curr. Biol.
<4>2
<5>450-452
<6>1992
<7>The "spacer protein" generated by a type of protein splicing in yeast turns out to be an
endonuclease with the precise DNA-sequence specificity to promote the spread of its own coding
sequence.

<>

<1>Grizot, S.
<2>I-MsoI homing endonuclease variants having novel substrate specificity and use for genetic engineering, genome therapy and antiviral therapy.
<3>International Patent Office
<4>WO 200968937 A
<5>
<6>2009
<7>An I-MsoI homing endonuclease variant able to cleave mutant I-MsoI sites having variation at
positions +/-8 to +/-10, a vector encoding said variant, a cell, an animal or a plant modified
by said vector.  Use of said I-MsoI endonuclease variant and derived products for genetic
engineering, genome therapy and antiviral therapy.

<>

<1>Grizot, S., Duchateau, P.
<2>Substitution variants of meganuclease I-DmoI with modified target sequence selection and increased catalytic activity.
<3>International Patent Office
<4>WO 200974873 A
<5>
<6>2009
<7>The current invention relates to polypeptides encoding mutant I-DmoI derivatives with enhanced
cleavage activity and altered sequence specificity and uses of these polypeptides.  These
polypeptides comprise at leaast the first I-DmoI domain, and the peptide sequence comprises
the substitution of at least one of residues 15, 19 and/or 20 as well as at least one of the
residues in positions 27, 29, 33, 35, 37, 75, 76, 81 of the first I-DmoI domain.

<>

<1>Grizot, S., Duchateau, P.
<2>Improved chimeric meganuclease enzymes and their uses for cleavage of specific nucleic acid sequences.
<3>International Patent Office
<4>WO 200974842 A
<5>
<6>2009
<7>The current invention relates to polypeptides encoding mutant I-DmoI derivatives with enhanced
cleavage activity and altered sequence specificity and uses of these polypeptides.  These
polypeptides comprise at least the first I-DmoI domain, and the peptide sequence comprises the
substitution of at least one of residues 15, 19, and/or 20 as well as at least one of the
residues in positions 29, 33, 35, 75, 76, 77 of the first I-DmoI domain.

<>

<1>Grizot, S., Duclert, A., Thomas, S., Duchateau, P., Paques, F.
<2>Context dependence between subdomains in the DNA binding interface of the I-CreI homing endonuclease.
<3>Nucleic Acids Res.
<4>39
<5>6124-6136
<6>2011
<7>Homing endonucleases (HE) have emerged as precise tools for achieving gene targeting events.
Redesigned HEs with tailored specificities can be used to cleave new sequences, thereby
considerably expanding the number of targetable genes and loci. With HEs, as well as with
other protein scaffolds, context dependence of DNA/protein interaction patterns remains one of
the major limitations for rational engineering of new DNA binders. Previous studies have shown
strong crosstalk between different residues and regions of the DNA binding interface. To
investigate this phenomenon, we systematically combined mutations from three groups of amino
acids in the DNA binding regions of the I-CreI HE. Our results confirm that important
crosstalk occurs throughout this interface in I-CreI. Detailed analysis of success rates
identified a nearest-neighbour effect, with a more pronounced level of dependence between
adjacent regions. Taken together, these data suggest that combinatorial engineering does not
necessarily require the identification of separable functional or structural regions, and that
groups of amino acids provide acceptable building blocks that can be assembled, overcoming the
context dependency of the DNA binding interface. Furthermore, the present work describes a
sequential method to engineer tailored HEs, wherein three contiguous regions are individually
mutated and assembled to create HEs with engineered specificity.

<>

<1>Grizot, S., Epinat, J.C., Thomas, S., Duclert, A., Rolland, S., Paques, F., Duchateau, P.
<2>Generation of redesigned homing endonucleases comprising DNA-binding domains derived from two different scaffolds.
<3>Nucleic Acids Res.
<4>38
<5>2006-2018
<6>2010
<7>Homing endonucleases have become valuable tools for genome engineering. Their sequence
recognition repertoires can be expanded by modifying their
specificities or by creating chimeric proteins through domain swapping
between two subdomains of different homing endonucleases. Here, we show
that these two approaches can be combined to create engineered
meganucleases with new specificities. We demonstrate the modularity of the
chimeric DmoCre meganuclease previously described, by successfully
assembling mutants with locally altered specificities affecting both
I-DmoI and I-CreI subdomains in order to create active meganucleases with
altered specificities. Moreover these new engineered DmoCre variants
appear highly specific and present a low toxicity level, similar to
I-SceI, and can induce efficient homologous recombination events in
mammalian cells. The DmoCre based meganucleases can therefore offer new
possibilities for various genome engineering applications.

<>

<1>Grizot, S., Smith, J., Daboussi, F., Prieto, J., Redondo, P., Merino, N., Villate, M., Thomas, S., Lemaire, L., Montoya, G., Blanco, F.J., Paques, F., Duchateau, P.
<2>Efficient targeting of a SCID gene by an engineered single-chain homing endonuclease.
<3>Nucleic Acids Res.
<4>37
<5>5405-5419
<6>2009
<7>Sequence-specific endonucleases recognizing long target sequences are emerging as powerful
tools for genome engineering. These endonucleases
could be used to correct deleterious mutations or to inactivate viruses,
in a new approach to molecular medicine. However, such applications are
highly demanding in terms of safety. Mutations in the human RAG1 gene
cause severe combined immunodeficiency (SCID). Using the I-CreI dimeric
LAGLIDADG meganuclease as a scaffold, we describe here the engineering of
a series of endonucleases cleaving the human RAG1 gene, including obligate
heterodimers and single-chain molecules. We show that a novel single-chain
design, in which two different monomers are linked to form a single
molecule, can induce high levels of recombination while safeguarding more
effectively against potential genotoxicity. We provide here the first
demonstration that an engineered meganuclease can induce targeted
recombination at an endogenous locus in up to 6% of transfected human
cells. These properties rank this new generation of endonucleases among
the best molecular scissors available for genome surgery strategies,
potentially avoiding the deleterious effects of previous gene therapy
approaches.

<>

<1>Grogan, D.W.
<2>Cytosine methylation by the SuaI restriction-modification system: implications for genetic fidelity in a hyperthermophilic archaeon.
<3>J. Bacteriol.
<4>185
<5>4657-4661
<6>2003
<7>5-methylcytosine in chromosomal DNA represents a potential source of frequent spontaneous
mutation for hyperthermophiles. To determine the
relevance of this threat for the archaeon Sulfolobus acidocaldarius, the
mode of GGCC methylation by its restriction-modification system, SuaI, was
investigated. Distinct isoschizomers of the SuaI endonuclease were used to
probe the methylation state of GGCC in native S. acidocaldarius DNA. In
addition, the methylation sensitivity of the SuaI endonuclease was
determined with synthetic oligonucleotide substrates and modified natural
DNAs. The results show that the SuaI system uses N(4) methylation to block
cleavage of its recognition site, thereby avoiding the creation of G. T
mismatches by spontaneous deamination at extremely high temperature.

<>

<1>Groll, D.H., Jeltsch, A., Selent, U., Pingoud, A.
<2>Does the restriction endonuclease EcoRV employ a two-metal-ion mechanism for DNA cleavage?
<3>Biochemistry
<4>36
<5>11389-11401
<6>1997
<7>Two models for the catalytic mechanism of the restriction endonuclease EcoRV exist which
differ in the number and function of metal ions proposed to be directly involved in catalysis.
In one model, two metal ions bound by Glu45, Asp74, and Asp90 are assumed to have a direct
catalytic function; in the other, only one metal ion bound by Asp74 and Asp90.  We show here
that in the presence of Mn2+, the catalytic activity of an EcoRV-E45A mutant is only slightly
reduced (1.8-fold) as compared to wild type EcoRV and that the single-turnover rate constant
of DNA cleavage by E45A is reduced only 39-fold, whereas the D74A and D90A mutants are
catalytically inactive under all conditions.  These findings make an important catalytic
function of Glu45, like binding of an essential divalent metal ion, unlikely.  In addition, we
have analyzed the dependence of the DNA cleavage rate by EcoRV and EcoRV mutants on the
concentration of Mg2+ and Mn2+.  We found for the wild type enzyme a sigmoidal dependence of
the rate of DNA cleavage on the concentration of Mg2+ or Mn2+, indicative of at least two
metal ions involved in DNA binding and catalysis.  This, however, does not mean that EcoRV
follows a two-metal-ion mechanism in DNA cleavage, because also for the E45A mutant a
sigmoidal dependence of the rate of DNA cleavage on the Mg2+ concentration was found, making
metal ion binding to the E45/D74 site unlikely.  In contrast, the Y219C mutant shows a
hyperbolic dependence.  In agreement with results obtained earlier, these findings demonstrate
binding of a Mg2+ ion at a site influenced by Tyr219, an amino acid residue that is far away
from the active site.  Metal binding at this site does not have a catalytic role but rather
supports specific DNA binding.  We conclude that on the basis of our data a two-metal-ion
mechanism of DNA cleavage is unlikely for EcoRV and that the complex metal ion effects
observed are due to metal ion binding at sites that are not directly involved in catalysis.

<>

<1>Gromkova, R., Bendler, J., Goodgal, S.
<2>Restriction and modification of bacteriophage S2 in Haemophilus influenzae.
<3>J. Bacteriol.
<4>114
<5>1151-1157
<6>1973
<7>The major conclusion from these studies is that variants of Haemophilus
influenzae Rd which restrict and modify phage S2 are metastable and capable of
giving rise to one another with high frequency.  Nonrestrictive RdS cells
segregate spontaneously to the restricting, modifying phenotype in about 5% of
the progeny of a single clone.  The restriction cells derived from RdS revert
to the nonrestrictive phenotype in 15 to 25% of the progeny of a single clone.
These frequencies are not appreciably affected by treatment with acriflavine or
ethidium bromide, compounds which affect plasmid stability, or by
nitrosoguanidine, a powerful mutagen.  The genetic locus for restriction and
modification of bacteriophage S2 is found to have a chromosomal position
between the biotin and proline loci.  Restriction-modification of phage S2 has
been shown to be a function of its deoxyribonucleic acid (DNA) in that
transfection with S2 phage DNA or prophage DNA is subject to host restriction
and modification.  An enzyme preparation which contains endodeoxyribonuclease
but no appreciable exonuclease activity, from mutant H. influenzae com-10 did
not restrict phage S2.RdS DNA or prophage DNA transfecting activity, indicating
that this endodeoxyribonuclease is not responsible for phage restriction.  A
new restriction enzyme isolated from H. influenzae Rd was found to be the major
enzyme involved in the restriction of bacteriophage S2.  The enzyme inactivated
the transfecting activity of unmodified phage DNA but did not attack modified
phage DNA.  Unlike endodeoxyribonuclease R, this enzyme requires adenosine
triphosphate and S-adenosylmethionine.

<>

<1>Gromkova, R., Goodgal, S.
<2>On the restriction-modification systems of Haemophilus aegyptius.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>75
<5>103
<6>1975
<7>In order to assess the role of type I and type II restriction systems in vivo
in Haemophilus we have attempted to isolate mutants deficient in restriction
and modification.  Because no mutants of the type II system were found in H.
influenze Rd, H. aegyptius was used since it contained different type II
restriction activities.  Bacteriophage S2 does not form plaques on wild type H.
aegyptius although it is efficiently absorbed.  Mutants capable of plating S2
with an efficiency of 10-7 were obtained.  Restriction was completely
eliminated by three more stepwise mutagenic treatments.  Analysis of these
mutants suggested that the wild type had at least three independent type I
systems.  Mutants deficient in restriction-modification were also deficient in
transforming activity.  The type II restriction systems of H. aegyptius did not
restrict bacteriophage S2 from H. influenze although its DNA was attacked in
vitro by purified type II restriction enzymes.  In the same way unmodified S2
grown on H. aegyptius was not restricted in vivo by the type II restriction
systems of H. influenzae Rd, but was excluded by its type I restriction system.
These data support the conclusion that only type I restriction is involved in
the exclusion of bacteriophage S2 in Haemophilus.  A type I activity was
isolated from H. aegyptius and appears responsible for restriction of S2.

<>

<1>Gromkova, R., Goodgal, S.H.
<2>On the role of restriction enzymes in transformation and transfection.
<3>Mechanisms of recombination, Plenum, Grell, R.F., New York
<4>0
<5>209-215
<6>1974
<7>None

<>

<1>Gromkova, R., Goodgal, S.H.
<2>Biological Properties of a Haemophilus influenzae Restriction Enzyme, HindI.
<3>J. Bacteriol.
<4>127
<5>848-854
<6>1976
<7>A type I restriction enzyme from Haemophilus influenzae, HindI, which requires
adenosine 5'-triphosphate and 5-adenosyl methionine, was studied for its
activity on transfecting and transforming deoxyribonucleic acid (DNA).  The
enzyme reduced the size of unmodified bacteriophage S2 DNA from 37 Mdaltons to
approximately 10 Mdaltons, but did not affect modified S2 DNA.  Unmodified
transforming DNA was attacked in vitro by HindI; however, relatively low levels
of inactivation were obtained for single markers, and linked transformants were
inactivated as a function of the distance between markers. In contrast,
unmodified bacterial DNA was not inactivated in vivo for either single or
linked markers by the HindI restriction system, probably because the segments
generated by HindI were still capable of being integrated in vivo.  The lack of
preferential inactivation of markers by the enzyme suggests that it makes
random breaks in the DNA.

<>

<1>Gromkova, R., Goodgal, S.H.
<2>On the effect of a restriction modification system on transformation in Haemophilus influenzae.
<3>Fed. Proc.
<4>34
<5>588
<6>1975
<7>Type I restriction enzymes, those that require ATP and S-adenosyl methionine
for activity, have been isolated and shown to be responsible for restriction of
bacteriophages S2 and HPlcl in Haemophilus. H. influenzae serotypes Rb and Rd
have different Type I restriction and modification systems although their type
II endodeoxyribonucleases have the same specificities.  The H. influenzae Rd
system has been shown to reversibly segregate, with a high frequency, to a
restriction modification deficient phenotype, however, the Rb system is stable
and can be used to study the effect of the type I restriction modification
system on bacterial transformation.  Mutants with different restriction and
modification phenotypes were examined for their efficiency of transformation.
The mutants studied included the first step mutants, r+/-m+, and r-m-; and the
second step mutants r-m-, r-m+, and r-m+/- obtained by a second round of
mutation.  A reduction in normal transforming activity ranging from 0.5 to 0.01
was obtained with all mutants tested with the greatest reductions associated
with the second step r-m- strains.  Transformation of mutants to the r+m+
phenotype resulted in recovery of normal transforming efficiency, and,
conversely transformation of r+m+ wild type recipients to the r-m- phenotype
resulted in lowered transformability.  The mutants were found to have
deficiences in their type I restriction and modification enzymes but no changes
in their levels of type II restriction enzymes.

<>

<1>Gromkova, R., Goodgal, S.H.
<2>Action of Haemophilus endodeoxyribonuclease on biologically active deoxyribonucleic acid.
<3>J. Bacteriol.
<4>109
<5>987-992
<6>1972
<7>An enzyme similar to that described by Smith and Wilcox (15) for Haemophilus
influenzae which attacks foreign deoxyribonucleic acid (DNA) but not its own
has been isolated and purified from H. parainfluenzae.  The enzyme degrades
foreign DNA to limited sizes and can destroy the transforming activity of H.
influenzae and Bacillus subtilis DNA.  The enzyme can also destroy the
biological activity of H. influenzae phage and prophage DNA.  On the other
hand, the H. influenzae endodeoxyribonuclease can destroy the transforming
activity of H. parainfluenzae DNA but not its own DNA.  It also attacks B.
subtilis DNA and its transforming activity.

<>

<1>Gromkova, R., LaPorte, J., Goodgal, S.H.
<2>A restriction enzyme from Haemophilus influenzae requiring adenosine triphosphate and S-adenosyl methionine.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>73
<5>72
<6>1973
<7>The endodeoxyribonuclease isolated from H. influenzae does not attack H. influenzae DNA nor H.
influenzae phage S2 transfecting activity.  Since there is restriction and modification of
phage S2 in H. influenzae it was clear that another restriction system was present.  A new
restriction activity (Hd) that destroys the transfecting activity of unmodified DNA from phage
grown in nonrestrictive recipients and does not inactivate modified DNA has been purified from
H. influenzae.  Unlike endo-R the enzyme activity which attacks phage DNA requires ATP and
S-adenosyl methionine.  Transformation by foreign bacterial DNA was also inactivated by the Hd
system.  This inactivation was stimulated by ATP and SAM.  The enzyme is considerably larger
than endo-R; it elutes earlier in Bio-Gel fractionation, and sediments further in a glycerol
sedimentation velocity gradient.  A similar enzyme has been isolated from H. influenzae Reid.
Since the H. influenzae Reid enzyme can attack the DNA from phage grown in restrictive cells
of H. influenzae, one can conclude that the H. influenzae and H. influenzae Reid restriction
systems have different specificities.

<>

<1>Gromova, E.S., Elob, A.A., Kubareva, E.A., Metelev, V.G., Shabarova, Z.A.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic fragments of DNA.  IV. DNA duplexes with phosphoamide and pyrophosphate internucleotide bonds -- substrates for the study of single-strand breaks.
<3>Mol. Biol. (Mosk)
<4>20
<5>29-40
<6>1986
<7>A set of DNA duplexes containing repetitive recognition sites for restriction
endonucleases EcoRII, EcoRI, and AluI, in which the phosphodiester bonds in the EcoRII
cleavage sites were replaced by phosphoamide or unbreakable pyrophosphate bonds, was
synthesized.  It was found that the restriction endonuclease EcoRII does not cleave the
substrate at the phosphoamide bond.  Using substrates containing modified bonds in the
EcoRII recognition sites in one of the strands, it was shown that this enzyme is capable of
catalyzing single-strand breaks both in the dA- and in the dT-containing strand of the
recognition site.  The restriction endonuclease EcoRII interacts with both strands of the
recognition site of DNA, whereas cleavage of each of them occurs independently of the
cleavage of the other.  Restriction endonucleases EcoRI and AluI specifically digest the
synthesized substrates at the phosphodiester bonds; however, this cleavage is hindered if
the modified bond is situated in the recognition site (EcoRI) or adjoins it (AluI).  For
EcoRII and AluI this effect is more pronounced in the case of substrates with
pyrophosphate bonds than with phosphoamide bonds.

<>

<1>Gromova, E.S., Elob, A.A., Kubareva, E.A., Metelev, V.G., Shabarova, Z.A.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic fragments of DNA.
<3>Mol. Biol. (Mosk)
<4>20
<5>22-32
<6>1986
<7>A set of DNA duplexes containing repetitive recognition sites for restriction endonucleases
EcoRII, EcoRI, and AluI, in which the phosphodiester bonds in the EcoRII cleavage sites were
replaced by phosphoamide or unbreakable pyrophosphate bonds, was synthesized.  It was found
that the restriction endonuclease EcoRII does not cleave the substrate at the phosphoamide
bond.  Using substrates containing modified bonds in the EcoRII recognition sites in one of
the strands, it was shown that this enzyme is capable of catalyzing single-strand breaks both
in the dA- and in the dT-containing strand of the recognition site.  The restriction
endonuclease EcoRII interacts with both strands of the recognition site of DNA, whereas
cleavage of each of them occurs independently of the cleavage of the other.  Restriction
endonucleases EcoRI and AluI specifically digest the synthesized substrates at the
phosphodiester bonds; however, this cleavage is hindered if the modified bond is situated in
the recognition site (EcoRI) or adjoins it (AluI).  For EcoRII and AluI this effect is more
pronounced in the case of substrates with pyrophosphate bonds than with phosphoamide bonds.

<>

<1>Gromova, E.S., Khoroshaev, A.V.
<2>Prokaryotic DNA methyltransferases: The structure and the mechanism of interaction with DNA.
<3>Mol. Biol. (Mosk)
<4>37
<5>300-314
<6>2003
<7>The review considers current views on the function of DNA methyltransferases (MTases) that
belong to prokaryotic type II
restriction-modification systems. A commonly accepted classification of
MTases is described along with their primary and tertiary structures and
molecular mechanisms of their specific interaction with DNA (including
methylation). MTase inhibitors are also considered. Special emphasis is
placed on the flipping of the target heterocyclic base out of the double
helix and on the methods employed in its analysis. Base flipping is a
fundamentally new type of DNA conformational changes and is also of
importance in the case of other DNA-operating enzymes. MTases show unique
sequence homology, and are similar in structure of functional centers and
in the mechanism of methylation. These data contribute to the
understanding of the general biological significance of methylation, since
prokaryotic and eukaryotic MTases are structurally and functionally
similar.

<>

<1>Gromova, E.S., Kubareva, E.A., Vinogradova, M.N., Oretskaya, T.S., Shabarova, Z.A.
<2>Peculiarities of recognition of CCA/TGG sequences in DNA by restriction endonucleases MvaI and EcoRII.
<3>J. Mol. Recognit.
<4>4
<5>133-141
<6>1991
<7>To elucidate the mechanism of action of restriction endonucleases MvaI and EcoRII a study was
made of their interaction with a set of synthetic substrates in which the heterocyclic bases
or the sugar-phosphate backbone has been modified; individual nucleotide residues had been
removed or replaced with hydrocarbon bridges, and mismatched base pairs had been introduced.
The groups of atoms in the heterocyclic bases and the phosphates in the recognition site that
produce the most significant influence on the functioning of endonucleases MvaI and EcoRII
were discerned. Profound differences were found in the functioning of the MvaI and EcoRII
neoschizomers. The catalytic activity of EcoRII is significantly affected by any alteration in
the recognition site structure and conformation, with a modification in one strand of the
substrate causing the same decrease in the hydrolysis rate of both strands. Endonuclease MvaI
is tolerant to a number of structural abnormalities; the latter sometimes affect only
hydrolysis of one strand of the recognition site. The enzyme can preferentially cleave one of
the substrate strands. Mismatched base pairs retard and sometimes block the hydrolysis. The
effect depends on the particular enzyme, mismatch and its location.

<>

<1>Gromova, E.S., Kubareva, E.A., Yolov, A.A., Akatova, E.A., Nikolskaya, I.I., Shabarova, Z.A.
<2>Cleavage of concatemeric substrates by restriction endonucleases MvaI and SsoII.
<3>Biokhimiia
<4>56
<5>552-559
<6>1991
<7>The interaction of enzymes SsoII (^CCNGG) and MvaI (CC^A/TGG) with concatemeric
duplexes used earlier to study EcoRII (CCA/TGG) was investigated with a view to
elucidating the general principles of restriction endonuclease function.  A
pattern common to all three enzymes was observed with DNA duplexes containing
AA or TT pairs in the central position of the recognition site.  The AA pair
blocks or substantially hinders the endonuclease action, whereas the TT pair is
either less inhibitory or altogether inert.  SsoII, like EcoRII was able to
processively cleave the concatemeric substrates and to interact with (or to be
close to) the hydrogen in the 5th position of the outer dC residue of the
recognition site.  MvaI was found to differ from EcoRII in the way they
recognize and cleave the same nucleotide sequence.  The substrate-bound MvaI
molecule is incapable of linear diffusion along the DNA.  Effective hydrolysis
of dU- and m5dC-containing polymers rules out the participation of hydrophobic
contacts of the enzyme with the methyl group of the dT residue and with the 5th
hydrogen of the outer dC residue of the recognition site in DNA-protein
interactions.

<>

<1>Gromova, E.S., Kuchava, E.A., Pein, C.-D., Oreshkaya, T.S., Shabarova, Z.A., Check, D.
<2>Study of the DNA cleavage mechanism of the restriction endonuclease MvaI.
<3>Dokl. Akad. Nauk.
<4>295
<5>1493-1497
<6>1987
<7>None

<>

<1>Gromova, E.S., Oretskaya, T.S., Eritja, R., Guschlbauer, W.
<2>Kinetic studies of MvaI DNA methyltransferase interaction with modified oligonucleotide duplexes.
<3>Biochem. Mol. Biol. Int.
<4>36
<5>247-255
<6>1995
<7>We have measured steady-state kinetics of an N4-cytosine methylase, M.MvaI,
using as substrates modified non-selfcomplementary tetradecanucleotide duplexes containing the
CCWGG target sequence.  The inner or outer localisation of the dI residue in the MvaI
recognition
site seems to be of little importance since the specificity constants kcat/KM are only 2 to 7
fold
smaller than that of the canonical substrate.  Replacement of dG residues by dI in both
strands
resulted in a 25 to 60-fold decrease of the specificity constant.  Modifications of the
phosphate
backbone or opening of the sugar ring of one of the dG residues had only little influence on
the
action of M.MvaI.  The enzyme appears to be rather tolerant to different kinds of modification
in its
substrate in the minor groove.

<>

<1>Gromova, E.S., Petrauskene, O.V., Kubareva, E.A.
<2>Study of the mechanism of EcoRII endonuclease activation with synthetic substrates.
<3>Nucleic Acids Symp. Ser.
<4>24
<5>217-218
<6>1991
<7>EcoRII restriction endonuclease is not able to cleave phage DNA with isolated EcoRII
recognition sites.  However addition of the EcoRII sensitive DNA or short substrates may
stimulate such a cleavage.  It has been suggested that EcoRII requires at least two
recognition sites for its activation.  In order to substantiate the mechanism of
EcoRII-substrate interaction we report a model system convenient for stimulation effect
studies and the results of testing as activators a wide set of modified and unmodified DNA.

<>

<1>Gromova, E.S., Shabarova, A.Z.
<2>DNA-Protein interactions:  The use of synthetic oligo- and polynucleotides for studying single-stranded-DNA-binding proteins and restriction endonucleases.
<3>Prog. Nucleic Acid Res. Mol. Biol.
<4>39
<5>1-47
<6>1990
<7>None

<>

<1>Gromova, E.S., Vinogradova, M.N., Uporova, T.M., Gryaznova, O.I., Isagulyants, M.G., Kosykh, V.G., Nikolskaya, I.I., Shabarova, Z.A.
<2>DNA duplexes with phosphoamide bonds:  the interaction with EcoRII and SsoII restriction endonucleases.
<3>Bioorg. Khim.
<4>13
<5>269-272
<6>1987
<7>We studied the interaction of EcoRII and SsoII restriction endonucleases with
synthetic DNA duplexes, containing 3'N -> 5'P and 3'P -> 5'N phosphoamide
internucleotide bonds in one of the cleavage points.  Enzymatic hydrolysis of
the modified strand of the duplexes is blocked in all cases.  The presence of
phosphoamide bonds was found to reduce the rate of cleavage of the natural
strand by EcoRII and to have no influence in case of SsoII.  Properties of the
EcoRII endonuclease complex with its substrate, containing non-cleavable 3'N ->
5'P internucleotide bonds in each cleavage point, were examined.  In the
presence of Mg2+ ions the equilibrium association constant of the
enzyme-substrate complex is 3-fold reduced, and the dissociation rate constant
of the complex is increased by 1.5 times.

<>

<1>Gronenborn, B., Messing, J.
<2>Methylation of single-stranded DNA in vitro introduces new restriction endonuclease cleavage sites.
<3>Nature
<4>272
<5>375-377
<6>1978
<7>Restriction endonucleases recognise specific sequences in DNA, and these
endonucleases, especially those which generate cohesive ends, have been widely
used to clone DNA.  However, many DNAs lack sequences which are recognised by
endonucleases such as EcoRI, HindIII or BamHI.  A general method of overcoming
this problem has been described recently. This approach involves the synthesis
of oligonucleotides sensitive to a specific endonuclease and the blunt end
ligation of these molecules to the DNA to be cloned.  In contrast, we sought a
method which avoids the insertion of addtional nucleotides into a DNA sequence,
but depends on direct modification of DNA.  If a DNA sequence differs in only
one base pair from the recognition sequence of a restriction endonuclease, a
particular change of this base pair will generate the proper sequence.  Here we
describe a way of generating restriction endonuclease cleavage sites by single
base changes derived after in vitro methylation of single-stranded DNA.

<>

<1>Grones, J., Turna, J.
<2>ApaCI, an isoschizomer of BamHI isolated from Acetobacter pasteurianus.
<3>Nucleic Acids Res.
<4>20
<5>3513
<6>1992
<7>
<>

<1>Grones, J., Turna, J.
<2>Some properties of restriction endonucleases from Acetobacter pasteurianus.
<3>Biologia (Bratisl)
<4>46
<5>1103-1108
<6>1991
<7>Restriction endonucleases ApaBI, ApaCI and ApaDI have been purified from three different
strains of Acetobacter pasteurianus. The enzyme ApaBI recognized 35 cleavage sites on
bacteriophage lambda DNA, 20 sites on adeno-virus-2 DNA and 2 sites on plasmid pBR322 DNA.
The recognition sequence for this enzyme is
5'-GCANNNNN/TGC-3'
3'-CGT/NNNNNACG-5'
The ApaCI enzyme is an isoschizomer of the restriction endonuclease BamHI. ApaDI is the
third enzyme with 6 sites of cleavage on pBR 327 DNA, 7 sites on pAT 153 DNA more than 20
sites on bacteriophage lambda DNA.

<>

<1>Grones, J., Turna, J.
<2>Isolation of a new restriction enzyme, ApaCI, an isoschizomer of BamHI produced by Acetobacter pasteurianus.
<3>Folia Microbiol. (Praha)
<4>37
<5>353-356
<6>1992
<7>A new Type II restriction endonuclease ApaCI purified from Acetobacter pasteurianus is an
isoschizomer of BamHI that cleaves at the nucleotide sequence 5'-G/GATCC-3' of
double-stranded DNA. The single restriction activity present in this strain permits the rapid
purification of 30,000 units of cleavage activity from 10 g of freshly harvested cells. The
resulting ApaCI preparation is free of contaminant nuclease activities that might interfere
with in vitro manipulation of DNA.

<>

<1>Grones, J., Turna, J.
<2>Some properties of restriction endonuclease ApaBI from Acetobacter pasteurianus.
<3>Biochim. Biophys. Acta
<4>1162
<5>323-325
<6>1993
<7>A number of restriction endonucleases have been isolated from many species of prokaryotes and
their specificities documented. Recently, several restriction endonucleases have been isolated
from Acetobacter genera. Up to date, two restriction endonucleases, ApaI and ApaLI have been
isolated from Acetobacter pasteurianus cells. The isolation of a new restriction endonuclease,
type-II ApaBI is described in this article.

<>

<1>Gronow, S. et al.
<2>Complete genome sequence of Bacteroides salanitronis type strain (BL78).
<3>Standards in Genomic Sciences
<4>4
<5>191-199
<6>2011
<7>Bacteroides salanitronis Lan et al. 2006 is a species of the genus Bacteroides, which belongs
to the family Bacteroidaceae. The species is of interest because it
was isolated from the gut of a chicken and the growing awareness that the
anaerobic microflora of the cecum is of benefit for the host and may impact
poultry farming. The 4,308,663 bp long genome consists of a 4.24 Mbp chromosome
and three plasmids (6 kbp, 19 kbp, 40 kbp) containing 3,737 protein-coding and
101 RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Gronow, S. et al.
<2>Complete genome sequence of Paludibacter propionicigenes type strain (WB4).
<3>Standards in Genomic Sciences
<4>4
<5>36-44
<6>2011
<7>Paludibacter propionicigenes Ueki et al. 2006 is the type species of the genus Paludibacter,
which belongs to the family Porphyromonadaceae. The species is of
interest because of the position it occupies in the tree of life where it can be
found in close proximity to members of the genus Dysgonomonas. This is the first
completed genome sequence of a member of the genus Paludibacter and the third
sequence from the family Porphyromonadaceae. The 3,685,504 bp long genome with
its 3,054 protein-coding and 64 RNA genes consists of one circular chromosome and
is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Gronow, S. et al.
<2>Complete genome sequence of Veillonella parvula type strain (Te3).
<3>Standards in Genomic Sciences
<4>2
<5>57-65
<6>2010
<7>Veillonella parvula (Veillon and Zuber 1898) Prevot 1933 is the type species of the genus
Veillonella in the family Veillonellaceae within the order
Clostridiales. The species V. parvula is of interest because it is frequently
isolated from dental plaque in the human oral cavity and can cause opportunistic
infections. The species is strictly anaerobic and grows as small cocci which
usually occur in pairs. Veillonellae are characterized by their unusual
metabolism which is centered on the activity of the enzyme methylmalonyl-CoA
decarboxylase. Strain Te3(T), the type strain of the species, was isolated from
the human intestinal tract. Here we describe the features of this organism,
together with the complete genome sequence, and annotation. This is the first
complete genome sequence of a member of the large clostridial family
Veillonellaceae, and the 2,132,142 bp long single replicon genome with its 1,859
protein-coding and 61 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Groot, M.N., Nieboer, F., Abee, T.
<2>Enhanced Transformation Efficiency of Recalcitrant Bacillus cereus and Bacillus weihenstephanensis Isolates upon In Vitro Methylation of Plasmid DNA.
<3>Appl. Environ. Microbiol.
<4>74
<5>7817-7820
<6>2008
<7>Digestion patterns of chromosomal DNAs of Bacillus cereus and Bacillus weihenstephanensis
strains suggest that Sau3AI-type restriction
modification systems are widely present among the isolates tested. In
vitro methylation of plasmid DNA was used to enhance poor plasmid
transfer upon electroporation to recalcitrant strains that carry Sau3AI
restriction barriers.

<>

<1>Gros, C., Chauvigne, L., Poulet, A., Menon, Y., Ausseil, F., Dufau, I., Arimondo, P.B.
<2>Development of a universal radioactive DNA methyltransferase inhibition test for high-throughput screening and mechanistic studies.
<3>Nucleic Acids Res.
<4>41
<5>12
<6>2013
<7>DNA methylation is an important epigenetic mark in eukaryotes, and aberrant pattern of this
modification is involved in numerous diseases such as cancers. Interestingly, DNA methylation
is reversible and thus is considered a promising therapeutic target. Therefore, there is a
need for identifying new small inhibitors of C5 DNA methyltransferases (DNMTs). Despite the
development of numerous in vitro DNMT assays, there is a lack of reliable tests suitable for
high-throughput screening, which can also  ive insights into inhibitor mechanisms of action.
We developed a new test based on scintillation proximity assay meeting these requirements.
After optimizing our assay on human DNMT1 and calibrating it with two known inhibitors, we
carried out S-Adenosyl-L-Methionine and DNA competition studies on three inhibitors and were
able to determine each mechanism of action. Finally, we showed that our test was applicable to
3 other methyltransferases sources: human DNMT3A, bacterial M. SssI and cellular extracts a s
well.

<>

<1>Grosskopf, R., Kessler, C.
<2>Restriction endonuclease DraII.
<3>US Patent Office
<4>US 4840901
<5>
<6>1989
<7>The present invention teaches a restriction endonuclease which recognizes and cleaves
palindromic sequence 5'Pu GGNCC Py3' 3'Py CCNGG Pu5' as well as a process for obtaining
this endonuclease. One source of the endonuclease is Deinococcus radiophilus DSM 20551 (ATCC
27063). The endonuclease is useful in analysis of DNA molecules.

<>

<1>Grosskopf, R., Kessler, C.
<2>Restriction endonuclease DraIII.
<3>US Patent Office
<4>US 4863858
<5>459
<6>1989
<7>*
A restriction endonuclease which recognizes palindromic sequence:

   5' CACNNNGTG 3'
   3' GTGNNNCAC 5'

and cleaves said sequence between the third and fourth base from the 3' end.


<>

<1>Grosskopf, R., Kessler, C.
<2>New restriction endonuclease DraIII prepared by cultivating Deinococcus radiophilus useful in DNA analysis.
<3>German Patent Office
<4>DE 3425957
<5>
<6>1986
<7>
<>

<1>Grosskopf, R., Kessler, C.
<2>Restriction endonuclease DraII.
<3>German Patent Office
<4>DE 3420298
<5>
<6>1989
<7>The present invention teaches a restriction endonuclease which recognizes and cleaves
palindromic sequence 5'Pu GGNCC Py3' 3'Py CCNGG Pu5' as well as a process for obtaining
this endonuclease. One source of the endonuclease is Deinococcus radiophilus DSM 20551 (ATCC
27063). The endonuclease is useful in analysis of DNA molecules.

<>

<1>Grosskopf, R., Wolf, W., Kessler, C.
<2>Two new restriction endonucleases DraII and DraIII from Deinococcus radiophilus.
<3>Nucleic Acids Res.
<4>13
<5>1517-1528
<6>1985
<7>In addition to recently characterized DraI (1), two new Type II restriction
endonucleases, DraII and DraIII, with novel site-specificities were isolated
and purified from Deinococcus radiophilus ATCC 27603.  DraII and DraIII
recognize the hepta- and nonanucleotide sequences(DraII)5'-Pu G ^ G N C C Py-3'
3'-Py C C N G ^ G Pu-5'and(DraIII)5'-C A C N N N ^ G T G-3' 3'-G T G ^ N N N C
A C-5'The cleavage sites within both strands are indicated by arrows.  The
recognition sequences were established by mapping of the cleavage sites on
pBR322 (DraII) and fd109 RF DNA (DraIII).  The sequence specificities were
confirmed by computer-assisted restriction analyses of the generated fragment
patterns of the sequenced DNA's of the bacteriophages lambda, PhiX174 RF,
M13mp8 RF and fd109 RF, the viruses Adeno2 and SV40, and the plasmids pBR322
and pBR328.  The cleavage positions within the recognition sequences were
determined by sequencing experiments.

<>

<1>Grote, J. et al.
<2>Draft genome sequence of strain HIMB100, a cultured representative of the SAR116 clade of marine Alphaproteobacteria.
<3>Standards in Genomic Sciences
<4>5
<5>269-278
<6>2011
<7>Strain HIMB100 is a planktonic marine bacterium in the class Alphaproteobacteria. This strain
is of interest because it is one of the first known isolates from a globally ubiquitous clade
of marine bacteria known as SAR116 within the family Rhodospirillaceae. Here we de-scribe
preliminary features of the organism, together with the draft genome sequence and an-notation.
This is the second genome sequence of a member of the SAR116 clade. The 2,458,945 bp genome
contains 2,334 protein-coding and 42 RNA genes.

<>

<1>Grote, J., Schott, T., Bruckner, C.G., Glockner, F.O., Jost, G., Teeling, H., Labrenz, M., Jurgens, K.
<2>Genome and physiology of a model Epsilonproteobacterium responsible for sulfide detoxification in marine oxygen depletion zones.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>109
<5>506-510
<6>2012
<7>Eutrophication and global climate change lead to expansion of hypoxia in the ocean, often
accompanied by the production of hydrogen sulfide, which
is toxic to higher organisms. Chemoautotrophic bacteria are thought to
buffer against increased sulfide concentrations by oxidizing hydrogen
sulfide before its diffusion to oxygenated surface waters. Model organisms
from such environments have not been readily available, which has
contributed to a poor understanding of these microbes. We present here a
detailed study of 'Sulfurimonas gotlandica' str. GD1, an
Epsilonproteobacterium isolated from the Baltic Sea oxic-anoxic interface,
where it plays a key role in nitrogen and sulfur cycling. Whole-genome
analysis and laboratory experiments revealed a high metabolic flexibility,
suggesting a considerable capacity for adaptation to variable redox
conditions. S. gotlandica str. GD1 was shown to grow
chemolithoautotrophically by coupling denitrification with oxidation of
reduced sulfur compounds and dark CO(2) fixation. Metabolic versatility
was further suggested by the use of a range of different electron donors
and acceptors and organic carbon sources. The number of genes involved in
signal transduction and metabolic pathways exceeds those of other
Epsilonproteobacteria. Oxygen tolerance and environmental-sensing systems
combined with chemotactic responses enable this organism to thrive
successfully in marine oxygen-depletion zones. We propose that S.
gotlandica str. GD1 will serve as a model organism in investigations that
will lead to a better understanding how members of the
Epsilonproteobacteria are able to cope with water column anoxia and the
role these microorganisms play in the detoxification of sulfidic waters.

<>

<1>Grouzdev, D.S., Dziuba, M.V., Sukhacheva, M.S., Mardanov, A.V., Beletskiy, A.V., Kuznetsov, B.B., Skryabin, K.G.
<2>Draft Genome Sequence of Magnetospirillum sp. Strain SO-1, a Freshwater Magnetotactic Bacterium Isolated from the Ol'khovka River, Russia.
<3>Genome Announcements
<4>2
<5>e00235-14
<6>2014
<7>Here, we present the draft genome sequence of Magnetospirillum sp. strain SO-1, a freshwater
magnetotactic spirillum isolated from the sediments of the Ol'khovka River, Russia.

<>

<1>Grover, S., Sharma, V.K., Mallapa, R.H., Batish, V.K.
<2>Draft Genome Sequence of Lactobacillus fermentum Lf1, an Indian Isolate of Human  Gut Origin.
<3>Genome Announcements
<4>1
<5>e00883-13
<6>2013
<7>Lactobacillus fermentum is a normal inhabitant of the human gastrointestinal tract. Here, we
report the draft genome sequence of an Indian isolate of the
probiotic strain L. fermentum Lf1, isolated from the human gut.

<>

<1>Grover, S., Sharma, V.K., Mallapa, R.H., Batish, V.K.
<2>Draft Genome Sequence of Lactobacillus plantarum Strain Lp91, a Promising Indian  Probiotic Isolate of Human Gut Origin.
<3>Genome Announcements
<4>1
<5>e00976-13
<6>2013
<7>Lactobacillus plantarum is a highly versatile species among lactic acid bacteria  that has
been widely isolated from highly diversified ecological niches,
including the gastrointestinal tract. Here, we report the first draft genome
sequence of an Indian isolate of the probiotic strain L. plantarum Lp91, isolated
from human gut.

<>

<1>Gruber, I.M., Nikolskaya, I.I., Uporova, T.M., Nisilevich, V.F.
<2>Growth and bioenergetic characteristics of Escherichia coli CK, capable of producing restriction endonuclease, under the conditions of batch cultivation.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>6
<5>50-55
<6>1984
<7>The conditions necessary for the controlled single and multicycle process of
the batch cultivation of E. coli CK, capable of producing E. coli CK specific
endonuclease, have been established.  This process can be regulated with
respect to a number of parameters (temperature, pH, pO2, eH).  The possibility
of using thermodynamic characteristics, calculated on the basis of the redox
potential and disclosing the energetics of growth, for evaluating the
effectiveness of controlled batch cultivation.  The optimum results have been
obtained during the isolation of E. coli CK restriction endonuclease, active
and containing no admixture of other endonucleases, at the period from the
maximum specific growth rate to the end of the exponential growth phase, i.e.
to the beginning of the stationary phase.

<>

<1>Gruen, M., Chang, K., Serbanescu, I., Liu, D.R.
<2>An in vivo selection system for homing endonuclease activity.
<3>Nucleic Acids Res.
<4>30
<5>e29
<6>2002
<7>Homing endonucleases are enzymes that catalyze the highly sequence-specific cleavage of DNA.
We have developed an in vivo selection in Escherichia coli that links cell survival with
homing endonuclease-mediated DNA cleavage activity and sequence specificity. Using this
selection, wild-type and mutant variants of three homing endonucleases were characterized
without requiring protein purification and in vitro analysis. This selection system may
facilitate the study of sequence-specific DNA cleaving enzymes, and selections based on this
work may enable the evolution of homing endonucleases with novel activities or specificities.

<>

<1>Gruenbaum, Y., Cedar, H., Razin, A.
<2>Restriction enzyme digestion of hemimethylated DNA.
<3>Nucleic Acids Res.
<4>9
<5>2509-2515
<6>1981
<7>Hemimethylated duplex DNA of the bacteriophage PhiX174 was synthesized using
primed repair synthesis in vitro with E. coli DNA polymerase I followed by
ligation to produce the covalently closed circular duplex (RFI).
Single-stranded PhiX DNA was used as template, a synthetic oligonucleotide as
primer and 5-methyldeoxycytidine-5'-triphosphate (5mdCTP) was used in place of
dCTP.  The hemimethylated product was used as substrate for cleavage by various
restriction enzymes.  Out of the 17 enzymes tests, only 5 (BstNI, TaqI, HincII,
HinfI and HpaI) cleaved the hemimethylated DNA.  Two enzymes (MspI and HaeIII)
were able to produce nicks on the unmethylated strand of the cleavage site.
MspI, which is known to cleave at CCGG when the internal cytosine residue is
methylated, does not cleave when both cytosines are methylated.  Another
enzyme, ApyI, cleaves at the sequence CC(A/T)GG when the internal cytosine is
methylated, but is inactive on hemimethylated DNA in which both cytosines are
methylated.  Hemimethylated molecules should be useful for studying DNA
methylation both in vivo and in vitro.

<>

<1>Grunau, C., Schattevoy, R., Mache, N., Rosenthal, A.
<2>MethTools-a toolbox to visualize and analyze DNA methylation data.
<3>Nucleic Acids Res.
<4>28
<5>1053-1058
<6>2000
<7>The Bisulfite Genomic Sequencing technique has found wide acceptance for the generation of
DNA-methylation maps with single-base resolution. The method is based on the selective
deamination of cytosine to uracil (and subsequent conversion to thymine via PCR), whereas
5-methylcytosine residues remain unchanged. Methylation maps are created by the comparison of
bisulfite converted sequences with the untreated genomic sequence. 'MethTools' is a
collection of software tools that replaces the time-consuming manual comparison process,
generates graphical outputs of methylation patterns and methylation density, estimates the
systematic error of the experiment and searches for conserved methylated nucleotide patterns.
The programs are written in Perl 5 and C, and the source code can be downloaded. All tools run
independently but the programs are interfaced. Thus, a script can perform the entire analysis
procedure automatically. In addition, a web-based remote analysis service is offered. Both the
source code and the remote analysis are available at http://genome.imb-jena.de/methtools/

<>

<1>Gruning, B.A., Erxleben, A., Hahnlein, A., Gunther, S.
<2>Draft Genome Sequence of Streptomyces viridochromogenes Strain Tu57, Producer of  Avilamycin.
<3>Genome Announcements
<4>1
<5>e00384-13
<6>2013
<7>Here we present the draft genome sequence of Streptomyces viridochromogenes Tu57. This strain
is a producer of avilamycin A, an oligosaccharide antibiotic from the
orthosomycin group, which is active against Gram-positive bacteria.

<>

<1>Grunwald, S., Driever, P.H., Hoelzer, D., Drahovsky, D.
<2>Reduced methyl group acceptance of 1-beta-D-arabinofuranosylcytosine-containing DNA polymers.
<3>Biochim. Biophys. Acta
<4>950
<5>366-373
<6>1988
<7>Previous studies have shown that 1-beta-D-arabinofuranosylcytosine (ara-C) can
induce differentiation of various malignant cells and that DNA methylation
patterns become altered under ara-C treatment of those cells.  The aim of this
study was to investigate whether this influence on DNA methylation is caused by
a direct effect of DNA-incorporated ara-C molecules on nuclear DNA methylase.
For this reason, we constructed various ara-C-substituted DNA polymers and used
them as substrates for highly purified eukaryotic DNA methylase isolated from
murine P815 mastocytoma cells.  The ara-C incorporation into DNA polymers was
measured by either an ara-C-specific radioimmunoassay or by use of
radioactive-labelled ara-C during the synthesis of those polymers.  We found an
inverse correlation between the level of ara-C substitution of the DNA polymers
and their methyl group acceptance.  Kinetic experiments performed with
ara-C-modified DNA polymers pointed out that the mode of action of DNA
methylase remains unaltered.  DNA methylase is neither detached nor fixed at an
ara-C site, but is somehow hindered in its enzymatic activity, probably by
slowing down the walking mechanism.  Hence, the previously observed
hypermethylation of DNA of some eukaryotic cells, propagated in the presence of
ara-C, is apparently not due to a direct effect of DNA-incorporated ara-C
molecules on endogenous DNA methylase.

<>

<1>Gu, H.H., Xu, J., Gallagher, M., Dean, G.E.
<2>Peptide splicing in the vacuolar ATPase subunit A from Candida tropicalis.
<3>J. Biol. Chem.
<4>268
<5>7372-7381
<6>1993
<7>Subunit A of the vacuolar proton pump appears to be responsible for the ATP hydrolysis which
is coupled to the pumping of protons into a variety of intracellular acid compartments,
including the fungal vacuole.  We report here the cloning and sequence determination of the
gene encoding subunit A from Candida tropicalis.  Southern blot hybridization analysis
indicates that there is a single gene which encodes this protein.  The gene contains a single
intron at the extreme 5'-end  of the coding region.  The gene is predicted to encode a
polypeptide of 1088 residues with a calculated molecular mass of 119,019 daltons, yet the
mature polypeptide appears to be approximately 67 kDa, indicating that this protein probably
undergoes the same sort of processing that is evidenced in the homologous protein from
Saccharomyces cerevisiae in which an approximately 50-kDa polypeptide (the spacer) is spliced
out of the mature protein.  The Candida gene, with and without this middle portion, has been
expressed in S. cerevisiae and found to restore a Saccharomyces subunit A deletion mutant
(tfp1-delta8) to apparently wild-type growth at pH 7.6, and normal vacuolar acidification.
The peptide sequence of the two predicted mature ends is very similar to the sequences of the
analogous proteins from Daucus carota, S. cerevisiae, and Neurospora crassa (60.5, 87.4, and
72.9% identity, respectively), but the middle portion bears only very limited homology with
the Saccharomyces protein sequence.  Processing of the gene product occurs in S. cerevisiae,
Escherichia coli, and in rabbit reticulocyte-mediated in vitro translation, indicating that
the excision is probably autocatalytic.  The limited sequence identity seen between the
Saccharomyces and Candida spacer domains may considerably narrow the functionally important
regions responsible for the excision event.

<>

<1>Gu, Y., Yang, C., Wang, X., Geng, W., Sun, Y., Feng, J., Wang, Y., Quan, Y., Che, Y., Zhang, C., Gong, T., Zhang, W., Gao, W., Zuo, Z., Song, C., Wang, S.
<2>Genome Sequence of the epsilon-Poly-l-Lysine-Producing Strain Streptomyces albulus NK660, Isolated from Soil in Gutian, Fujian Province, China.
<3>Genome Announcements
<4>2
<5>e00532-14
<6>2014
<7>We determined the complete genome sequence of a soil bacterium, Streptomyces albulus NK660. It
can produce epsilon-poly-l-lysine, which has antimicrobial
activity against a spectrum of microorganisms. The genome of S. albulus NK660
contains a 9,360,281-bp linear chromosome and a 12,120-bp linear plasmid.

<>

<1>Gualtieri, M., Ogier, J.C., Pages, S., Givaudan, A., Gaudriault, S.
<2>Draft Genome Sequence and Annotation of the Entomopathogenic Bacterium Xenorhabdus szentirmaii Strain DSM16338.
<3>Genome Announcements
<4>2
<5>e00190-14
<6>2014
<7>We report the genome sequence of Xenorhabdus szentirmaii DSM16338 (4.84 Mb), a symbiont of the
entomopathogenic nematode Steinernema rarum. This strain produces
antimicrobial activity.

<>

<1>Guan, P., Ai, P., Dai, X., Zhang, J., Xu, L., Zhu, J., Li, Q., Deng, Q., Li, S., Wang, S., Liu, H., Wang, L., Li, P., Zheng, A.
<2>Complete Genome Sequence of Bacillus thuringiensis Serovar Sichuansis Strain MC28.
<3>J. Bacteriol.
<4>194
<5>6975
<6>2012
<7>Bacillus thuringiensis is an important microbial insecticide used in the control  of
agricultural pests. Here we report the finished, annotated genome sequence of
Bacillus thuringiensis serovar Sichuansis strain MC28, which can form parasporal
crystals consisting of Cry4Cc1, Cry30Fa1, Cry53Ab1, Cry54Aa1, Cry54Ab1, Cry68Aa1,
Cry69Aa1, Cry69Aa2, Cry70Ba1, Cyt1Da1, and Cyt2Aa3. It is also highly toxic to
lepidopterous and dipterous insects.

<>

<1>Guan, S., Zhu, Z., Blanchard, A.
<2>Methods for engineering high fidelity restriction endonuclease BtsIM1 and BtsIM2 variants from Bacillus thermoglucosidasius with altered cleavage specificity and reduction of star activity.
<3>International Patent Office
<4>WO 201128841 A
<5>
<6>2011
<7>Mutations introduced into a restriction endonuclease may alter not only fidelity of cleavage
but also specificity.  Examples of novel variant endonucleases with altered specificities are
described and also variants with high fidelity of cleavage.  Examples describe muations in
BtsI that result in at least one of altered cleavage and nicking and high fidelity cleavage.

<>

<1>Guan, S.X., Blanchard, A., Zhang, P.H., Zhu, Z.Y.
<2>Alteration of Sequence Specificity of the Type IIS Restriction Endonuclease BtsI.
<3>PLoS ONE
<4>5
<5>e11787
<6>2010
<7>The Type IIS restriction endonuclease BtsI recognizes and digests at GCAGTG(2/0). It comprises
two subunits: BtsIA and BtsIB. The BtsIB
subunit contains the recognition domain, one catalytic domain for
bottom strand nicking and part of the catalytic domain for the top
strand nicking. BtsIA has the rest of the catalytic domain that is
responsible for the DNA top strand nicking. BtsIA alone has no activity
unless it mixes with BtsIB to reconstitute the BtsI activity. During
characterization of the enzyme, we identified a BtsIB mutant R119A
found to have a different digestion pattern from the wild type BtsI.
After characterization, we found that BtsIB(R119A) is a novel
restriction enzyme with a previously unreported recognition sequence
CAGTG(2/0), which is named as BtsI-1. Compared with wild type BtsI,
BtsI-1 showed different relative activities in NEB restriction enzyme
reaction buffers NEB1, NEB2, NEB3 and NEB4 and less star activity.
Similar to the wild type BtsIB subunit, the BtsI-1 B subunit alone can
act as a bottom nicking enzyme recognizing CAGTG(-/0). This is the
first successful case of a specificity change among this restriction
endonuclease type.

<>

<1>Guan, W., Shao, J., Davis, R.E., Zhao, T., Huang, Q.
<2>Genome Sequence of a Xylella fastidiosa Strain Causing Sycamore Leaf Scorch Disease in Virginia.
<3>Genome Announcements
<4>2
<5>e00773-14
<6>2014
<7>Xylella fastidiosa causes bacterial leaf scorch in landscape trees including sycamore. We
determined the draft genome of X. fastidiosa strain Sy-Va, isolated
in Virginia from a sycamore tree displaying leaf scorch symptoms. The Sy-VA
genome contains 2,477,829 bp, and has a G+C content of 51.64 mol%.

<>

<1>Guan, W., Shao, J., Zhao, T., Huang, Q.
<2>Genome Sequence of a Xylella fastidiosa Strain Causing Mulberry Leaf Scorch Disease in Maryland.
<3>Genome Announcements
<4>2
<5>e00916-13
<6>2014
<7>Xylella fastidiosa causes bacterial leaf scorch in landscape trees, including mulberry. We
determined the draft genome of the mulberry strain Mul-MD in order
to gain a better understanding of the molecular basis of strain divergence, host
specificity, nutrient requirements, and pathogenicity, as well as to develop
genome-based specific detection methods.

<>

<1>Guan, Y., Ngugi, D.K., Blom, J., Ali, S., Ferry, J.G., Stingl, U.
<2>Draft Genome Sequence of an Obligately Methylotrophic Methanogen, Methanococcoides methylutens, Isolated from Marine Sediment.
<3>Genome Announcements
<4>2
<5>e01184-14
<6>2014
<7>Methanococcoides methylutens, the type species of the genus Methanococcoides, is  a slightly
halophilic methanogenic archaeon with a methylotrophic metabolism.
Here, we present the annotated draft genome sequence of M. methylutens, which
comprises 2,508,511 bp with 2,482 coding sequences, 51 tRNA genes, and a G+C
content of 42.5%.

<>

<1>Guardiola-Avila, I., Acedo-Felix, E., Noriega-Orozco, L., Yepiz-Plascencia, G., Sifuentes-Romero, I., Gomez-Gil, B.
<2>Draft Genome Sequence of Vibrio mimicus Strain CAIM 602T.
<3>Genome Announcements
<4>1
<5>e0008413
<6>2013
<7>Vibrio mimicus is a Gram-negative bacterium associated with gastrointestinal diseases in
humans around the world. We report the complete genome sequence of
the Vibrio mimicus strain CAIM 602(T) (CDC1721-77, LMG 7896(T), ATCC 33653(T)).

<>

<1>Gubler, M., Bickle, T.A.
<2>Increased protein flexibility leads to promiscuous protein-DNA interactions in type IC restriction-modification systems.
<3>EMBO J.
<4>10
<5>951-957
<6>1991
<7>We have investigated the role of a four amino acid element that is repeated twice and three
times, respectively, in the specificity polypeptides of the two allelic
restriction-modification systems EcoR124 and EcoR124/3. We had earlier shown that this
difference in amino acid sequence between the two systems is solely responsible for the
different DNA sequence specificities of the two systems. The effect of single amino acid
substitutions and small insertion and deletion mutations on restriction activity and
modification specificity was determined in vivo by phage infection assays and in vitro by
methylation of DNA with purified modification methylases. Mutant restriction-modification
systems with changes in the number and the length of the central amino acid repeats exhibited
decreased restriction activity and in some cases relaxed substrate specificity. Our data
strongly support the idea that the repetitive amino acid motif in the specificity polypeptides
forms part of a flexible interdomain linker. It may be responsible for positioning on the DNA
the two major specificity polypeptide domains which are thought to contact independently the
half sites of the split recognition sequences typical for all type I restriction-modification
systems.

<>

<1>Gubler, M., Braguglia, D., Meyer, J., Piekarowicz, A., Bickle, T.A.
<2>Recombination of constant and variable modules alters DNA sequence recognition by type IC restriction-modification enzymes.
<3>EMBO J.
<4>11
<5>233-240
<6>1992
<7>EcoR124I and EcoDXXI are allelic type I restriction-modification (R-M) systems
whose specificity genes consist of common structural elements: two variable
regions are separated by a constant, homologous region containing a number of
repetitive sequence elements.  In vitro recombination of variable and constant
elements has led to fully active, hybrid R-M systems exhibiting new and
predictable target site specificities.  Methylation of synthetic DNA sequences
with purified, hybrid modification methylases was used to confirm the proposed
recognition sequences.  The results clearly demonstrate the correlation between
protein domains and target site specificity.  Our data suggest that a bacterial
population may switch the recognition sequences of its type I R-M system by
single recombination events and thus is able to maintain a prokaryotic analogue
of the immune system of variable specificity.

<>

<1>Guedon, E., Delorme, C., Pons, N., Cruaud, C., Loux, V., Couloux, A., Gautier, C., Sanchez, N., Layec, S., Galleron, N., Almeida, M., van de Guchte, M., Kennedy, S.P., Ehrlich, S.D., Gibrat, J.F., Wincker, P., Renault, P.
<2>Complete Genome Sequence of the commensal Streptococcus salivarius strain JIM8777.
<3>J. Bacteriol.
<4>193
<5>5024-5025
<6>2011
<7>The commensal bacterium Streptococcus salivarius is a prevalent species of the human
oropharyngeal tract with an important role in oral ecology.
Here, we report the complete 2.2-Mb genome sequence and annotation of
strain JIM8777, which was recently isolated from the oral cavity of a
healthy, dentate infant.

<>

<1>Gueimonde, M., Bottacini, F., van Sinderen, D., Ventura, M., Margolles, A., Sanchez, B.
<2>Genome Sequence of Parascardovia denticolens IPLA 20019, Isolated from Human Breast Milk.
<3>J. Bacteriol.
<4>194
<5>4776-4777
<6>2012
<7>This work describes the draft genome of Parascardovia denticolens IPLA 20019, isolated from
human milk. This species, usually isolated from caries lesions, is
taxonomically related to the genus Bifidobacterium. The genetic information of
IPLA 20019 enhances our understanding of the adaptation of this P. denticolens
strain from human breast milk.

<>

<1>Gueimonde, M., Ventura, M., Margolles, A., Sanchez, B.
<2>Genome Sequence of the Immunomodulatory Strain Bifidobacterium bifidum LMG 13195.
<3>J. Bacteriol.
<4>194
<5>6997
<6>2012
<7>In this work, we report the genome sequences of Bifidobacterium bifidum strain LMG13195.
Results from our research group show that this strain is able to
interact with human immune cells, generating functional regulatory T cells.

<>

<1>Guenthner, C., Kim, S., Bednarik, D.P.
<2>Cloning, isolation, and characterization of the human T-cell DNA-cytosine 5-methyltransferase gene.
<3>J. Cell Biochem. Suppl.
<4>16E
<5>179
<6>1992
<7>DNA methylation can affect the latency of HIV and HTLV by effectively silencing
transcriptional expression. Previous studies have shown that a CpG island in the HIV LTR, when
methylated by the human DNA-cytosine 5-methyltransferase (MeTase), inactivates HIV
transcription in cis. Further characterization of the methyltransferase enzyme is therefore of
obvious interest. A human T-cell cDNA library from the cell line Jurkatt has been screened,
using a murine DNA methyltransferase cDNA clone as a probe, and a clone of approximately 0.7
Kb has been isolated (pMET2) and sequenced with 87% homology to an area in the 3' region of
the murine cDNA. Based on the size of the mature protein (-172 Kdaltons), a gene of at least 5
Kb is expected. Southern blot analysis of human genomic DNA yielded single fragments of 4.2 Kb
to over 12 Kb, using the pMET2 fragment as a probe. Northern blot analysis has been employed
to analyze transcriptional expression, and to determine the size of the methyltranferase
message. A human genomic library has been screened and four putative clones isolated and shown
positive by PCR analysis, using primers to pMET2. Southern analysis on phage DNA digested with
XbaI yields fragments large enough to include a full-length copy of the gene. Sub-cloning into
an expression vector is underway in order to obtain sequence information, and to further
characterize this gene.

<>

<1>Guerrero, L.D., Makhalanyane, T.P., Aislabie, J.M., Cowan, D.A.
<2>Draft Genome Sequence of Williamsia sp. Strain D3, Isolated From the Darwin Mountains, Antarctica.
<3>Genome Announcements
<4>2
<5>e01230-13
<6>2014
<7>Actinobacteria are the dominant taxa in Antarctic desert soils. Here, we describe the first
draft genome of a member of the genus Williamsia (strain D3) isolated
from Antarctic soil. The genome of this psychrotolerant bacterium may help to
elucidate crucial survival mechanisms for organisms inhabiting cold desert soil
systems.

<>

<1>Guha, S.
<2>Determination of DNA Sequences Containing Methylcytosine in Bacillus subtilis Marburg.
<3>J. Bacteriol.
<4>163
<5>573-579
<6>1985
<7>The methylcytosine-containing sequences in the DNA of Bacillus subtilis 168
Marburg (Restriction-modification Type BSUM) were determined by three different
methods: (i) examination of in vivo-methylated DNA by restriction enzyme
digestion and, whenever possible, analysis for methycytosine at the 5' end;
(ii) methylation in vitro of unmethylated DNA with B. subtilis DNA
methyltransferase and determination of the methylated sites; and (iii) the
methylatability of unmethylated DNA by B. subtilis methyltransferase after
potential sites have been destroyed by digestion with restriction
endonucleases.  The results obtained by these methods, taken together, show
that methylcytosine was present only within the sequence 5'-TCGA-3'.  The
presence of methylcytosine at the 5' end of the DNA fragments generated by
restriction endonuclease AsuII digestion and the fact that in vivo-methylated
DNA could not be digested by the enzyme XhoI showed that the recognition
sequences of these two enzymes contained methylcytosine.  As these two enzymes
recognized a similar sequence containing a 5' pyrimidine (Py) and a 3' purine
(pu), 5'-PyTCGAPu'3', the possibility that methylcytosine is present in the
complementary sequences 5'TTCGAG-3' and 5'CTCGAA-3' was postulated.  This was
verified by the methylation in vitro, with B. subtilis enzyme, of a
2.6-kilobase fragment of lambda DNA containing two such sites and devoid of
AsuII or XhoI recognition sequences.  By analyzing the methylatable sites, it
was found that in one of the two PyTCGAPu sequences, cytosine was methylated in
vitro in both DNA strands.  It is concluded that the sequence 5'-PyTCGAPu'3' is
methylated by the DNA methyltransferase (of cytosine) of B. subtilis Marburg.

<>

<1>Guha, S.
<2>DNA methyltransferase of Bacillus subtilis Marburg:  purification, properties and further evidence of specificity.
<3>Gene
<4>74
<5>77-81
<6>1988
<7>Bacillus subtilis Marburg strain displays DNA methyltransferase activity. This enzyme, M.BsuM,
methylates cytosine in the sequence 5'-YTCGAR-3' (Y=pyrimidine; R=purine). M.BsuM was
purified from exponentially growing cells of B. subtilis 168M. This enzyme (45+/- 1 kDa) is
monomeric and recognizes only double-stranded DNA. It is inhibited partially by Mg2+, Mn2+
ions and spermidine and almost totally by sodium dodecyl sulfate, urea and agarose. This
enzyme methylates specifically the three methylatable sites of the plasmid pBM3. Relaxation of
specificity (star activity) was observed in the presence of organic solvents. A very low
amount of M.BsuM was obtained in the standard Marburg strain. To obtain sufficient enzyme
attempts are being made to clone the M.BsuM gene in Escherichia coli by using a constructed
plasmid (pBM14) vector. Only one transformant containing a 3-kb insert and showing a low level
of expression, was obtained.

<>

<1>Guha, S., Guschlbauer, W.
<2>Expression of Escherichia coli dam gene in Bacillus subtilis provokes DNA damage response: N6-methyladenine is removed by two repair pathways.
<3>Nucleic Acids Res.
<4>20
<5>3607-3615
<6>1992
<7>The dam gene of Excherichia coli encodes a DNA methyltransferase that methylates the N6
position of adenine in the sequence GATC. It was stably expressed from a shuttle vector in a
repair- and recombination-proficient strain of Bacillus subtilis. In this strain the majority
of plasmid DNA molecules was modified at dam sites whereas most chromosomal DNA remained
unmethylated during exponential growth. During stationary phase the amount of unmethylated DNA
increased, suggesting that methylated bases were being removed. An ultraviolet damage
repair-deficient mutant (uvrB) contained highly methylated chromosomal and plasmid DNA. High
levels of Dam methylation were detrimental to growth and viability of this mutant strain and
some features of the SOS response were also induced. A mutant defective in the synthesis of
adaptive DNA alkyltransferases and induction of the adaptive response (ada) also showed high
methylation and properties similar to that of the dam gene expressing uvrB strain. When
protein extracts from B. subtilis expressing the Dam methyltransferase or treated with
N-methyl-N-nitro-N-nitroso-guanidine were incubated with [3H]-labelled Dam methylated DNA, the
methyl label was bound to two proteins of 14 and 9 kD. Some free N6-methyladenine residues are
excised by proteins involved in both excision (uvrB) and the adaptive response (ada) DNA
repair pathways in B. subtilis.

<>

<1>Guha, S., Guschlbauer, W.
<2>Improved plasmids containing the Escherichia coli dam gene under the control of the tac promoter.
<3>Biochim. Biophys. Acta
<4>1132
<5>309-310
<6>1992
<7>We report the construction of a series of plasmids containing the dam gene under the control
of the tac promoter. Cells containing these plasmids produce about 8 to 10-fold more Dam
methyltransferase (Mtase) than the previously used plasmid pTP166 and avoid the use of a high
temperature step necessary for the expression of Dam Mtase in the plasmid pDOX1 and thus
allows its use for the study of thermosensitive mutants.

<>

<1>Guhan, N., Muniyappa, K.
<2>Mycobacterium tuberculosis RecA intein possesses a novel ATP-dependent site-specific double-stranded DNA endonuclease activity.
<3>J. Biol. Chem.
<4>277
<5>16257-16264
<6>2002
<7>Mycobacterium tuberculosis recA harbors an intervening sequence in its open reading frame,
presumed to encode an endonuclease (PI-MtuI) required for intein homing in inteinless recA
allele. Although the protein-splicing ability of PI-MtuI has been characterized, the
identification of its putative endonuclease activity has remained elusive. To investigate
whether PI-MtuI possesses endonuclease activity, recA intervening sequence was cloned,
overexpressed, and purified to homogeneity. Here we show that PI-MtuI bound both single- and
double-stranded DNA with similar affinity but failed to cleave DNA in the absence of
cofactors. Significantly, PI-MtuI nicked supercoiled DNA in the presence of alternative
cofactors but required both Mn2+ and ATP to generate linear double-stranded DNA. We observed
that PI-MtuI was able to inflict a staggered double-strand break 24 bp upstream of the
insertion site in the inteinless recA allele. Similar to a few homing endonucleases, DNA
cleavage by PI-MtuI was specific with an exceptionally long cleavage site spanning 22 bp. The
kinetic mechanism of PI-MtuI promoted cleavage supports a sequential rather than concerted
pathway of strand cleavage with the formation of nicked double-stranded DNA as an
intermediate. Together, these results reveal that RecA intein is a novel Mn2+-ATP-dependent
double-strand specific endonuclease, which is likely to be important for homing process in
vivo.

<>

<1>Guhan, N., Muniyappa, K.
<2>The RecA intein of Mycobacterium tuberculosis promotes cleavage of ectopic DNA sites: Implications for the dispersal of inteins in natural populations.
<3>J. Biol. Chem.
<4>277
<5>40352-40361
<6>2002
<7>The RecA intein of Mycobacterium tuberculosis, a novel double-stranded DNA endonuclease,
requires both Mn2+ and ATP for efficient cleavage of the inteinless recA allele. In this
study, we show that Mg2+ alone was sufficient to stimulate PI-MtuI to cleave double-stranded
DNA at ectopic sites. In the absence of Mg2+, PI-MtuI formed complexes with topologically
different forms of DNA containing ectopic recognition sequences with equal affinity but failed
to cleave DNA. We observed that PI-MtuI was able to inflict double-strand breaks robustly
within the ectopic recognition sequence to generate either a blunt end or 1-2 nucleotide
3'-hydroxyl overhangs. Mutational analyses of the presumptive metal-ion binding ligands
(D122, D222 and E220) together with immunoprecipitation assays provided compelling evidence to
link both the Mg2+- and Mn2+ and ATP-dependent endonuclease activities to PI-MtuI. The kinetic
mechanism of PI-MtuI promoted cleavage of ectopic DNA sites proceeded through a sequential
mechanism with transient accumulation of nicked circular duplex DNA as an intermediate.
Together, these data suggest that PI-MtuI, like group II introns, might mediate ectopic DNA
transposition and hence its lateral transfer in natural populations.

<>

<1>Guhan, N., Muniyappa, K.
<2>Mycobacterium tuberculosis RecA intein, a LAGLIDADG homing endonuclease, displays Mn2+ and DNA-dependent ATPase activity.
<3>Nucleic Acids Res.
<4>31
<5>4184-4191
<6>2003
<7>Mycobacterium tuberculosis RecA intein (PI-MtuI), a LAGLIDADG homing endonuclease, displays
dual target specificity in response to alternative
cofactors. While both ATP and Mn(2+) were required for optimal cleavage of
an inteinless recA allele (hereafter referred to as cognate DNA), Mg(2+)
alone was sufficient for cleavage of ectopic DNA sites. In this study, we
have explored the ability of PI-MtuI to catalyze ATP hydrolysis in the
presence of alternative metal ion cofactors and DNA substrates. Our
results indicate that PI-MtuI displays maximum ATPase activity in the
presence of cognate but not ectopic DNA. Kinetic analysis revealed that
Mn(2+) was able to stimulate PI-MtuI catalyzed ATP hydrolysis, whereas
Mg(2+) failed to do so. Using UV crosslinking, limited proteolysis and
amino acid sequence analysis, we show that (32)P-labeled ATP was bound to
a 14 kDa peptide containing the putative Walker A motif. Furthermore, the
limited proteolysis approach disclosed that cognate DNA was able to induce
structural changes in PI-MtuI. Mutation of the presumptive metal
ion-binding ligands (Asp122 and Asp222) in the LAGLIDADG motifs of PI-MtuI
impaired its affinity for ATP, thus resulting in a reduction in or loss of
its endonuclease activity. Together, these results suggest that PI-MtuI is
a (cognate) DNA- and Mn(2+)-dependent ATPase, unique from the LAGLIDADG
family of homing endonucleases, and implies a possible role for ATP
hydrolysis in the recognition and/or cleavage of homing site DNA sequence.

<>

<1>Guhan, N., Muniyappa, K.
<2>Structural and functional characteristics of homing endonucleases.
<3>Crit. Rev. Biochem. Mol. Biol.
<4>38
<5>199-248
<6>2003
<7>Mobile genetic elements constitute a remarkably diverse group of nonessential selfish genes
that provide no apparent function to the host.
These selfish genes have been implicated in host extinction, speciation
and architecture of genetic systems. Homing endonucleases, encoded by the
open reading frames embedded in introns or inteins of mobile genetic
elements, possess double-stranded DNA-specific endonuclease activity. They
inflict sequence-specific double-strand breaks at or near the homing site
in intron- or intein-less allele. Subsequently, through nonreciprocal
exchange the insertion sequence (intron or intein) is transferred from an
intein- or intron-containing allele to an intein- or intron-less allele.
The components of host double-strand break repair pathway are thought to
finish the "homing" process. Several lines of evidence suggest that homing
endonucleases are capable of promoting transposition into ectopic sites
within or across genomes for their survival as well as dispersal in
natural populations. The occurrence of inteins at high frequencies serves
as instructive models for understanding the mechanistic aspects of the
process of homing and its evolution. This review focuses on genetic,
biochemical, structural, and phylogenetic aspects of homing endonucleases,
and their comparison with restriction endonucleases.

<>

<1>Guild, W.R., Smith, M.D., Shoemaker, N.B.
<2>Conjugative transfer of chromosomal R determinants in Streptococcus pneumoniae.
<3>Microbiology-1982, American Society for Microbiology, Schlessinger, D., Washington, DC
<4>0
<5>88-92
<6>1982
<7>
<>

<1>Guimaraes, A.M., Toth, B., Santos, A.P., do Nascimento, N.C., Kritchevsky, J.E., Messick, J.B.
<2>Genome Sequence of 'Candidatus Mycoplasma haemolamae' Strain Purdue, a Red Blood  Cell Pathogen of Alpacas (Vicugna pacos) and Llamas (Lama glama).
<3>J. Bacteriol.
<4>194
<5>6312-6313
<6>2012
<7>We report the complete genome sequence of 'Candidatus Mycoplasma haemolamae,' an  endemic
red-cell pathogen of camelids. The single, circular chromosome has
756,845 bp, a 39.3% G+C content, and 925 coding sequences (CDSs). A great
proportion (49.1%) of these CDSs are organized into paralogous gene families,
which can now be further explored with regard to antigenic variation.

<>

<1>Guimaraes, L.C., Soares, S.C., Albersmeier, A., Blom, J., Jaenicke, S., Azevedo, V., Soriano, F., Tauch, A., Trost, E.
<2>Complete Genome Sequence of Corynebacterium urealyticum Strain DSM 7111, Isolated from a 9-Year-Old Patient with Alkaline-Encrusted Cystitis.
<3>Genome Announcements
<4>1
<5>e00264-13
<6>2013
<7>Corynebacterium urealyticum is a common skin colonizer with potent urease activity. It is
clinically recognized as an opportunistic pathogen causing
urinary tract infections. The annotated genome sequence of strain DSM 7111,
isolated from the urine of a young boy with an ectopic kidney, provides new
insights into the pathomechanisms of this bacterium.

<>

<1>Guimont, C., Henry, P., Linden, G.
<2>Restriction/modification in Streptococcus thermophilus: isolation and characterization of a type II restriction endonuclease Sth455I.
<3>Appl. Microbiol. Biotechnol.
<4>39
<5>216-220
<6>1993
<7>Streptococcus thermophilus strain CNRZ 455 produces a type II restriction endonuclease
designated Sth455I. This enzyme was isolated from cell extracts by anionic and cationic
exchange chromatography. This yielded an enzyme preparation free of non-specific nucleases.
The optimal reaction conditions for Sth455I are: MgCl2, 30 mM; pH range, 8-9; incubation
temperature, 37-40oC; and high NaCl concentration, 100-200 mM. The results of single- and
double-digestion experiments indicates that Sth455I is an isoschizomer of BstNI and EcoRII
showing different sensitivity to methylation The enzyme exhibits restriction activity on the
DNA of three bacteriophages of S. thermophilus and no activity on the phage lytic for strain
CNRZ 455. The restriction/modification system associated with this strain is discussed.

<>

<1>Guinane, C.M., Barrett, E., Fitzgerald, G.F., van Sinderen, D., Ross, R.P., Stanton, C.
<2>Genome Sequence of Bifidobacterium breve DPC 6330, a Strain Isolated from the Human Intestine.
<3>J. Bacteriol.
<4>193
<5>6799-6800
<6>2011
<7>The draft genome of Bifidobacterium breve DPC 6330, isolated from an elderly patient, was
determined. B. breve DPC 6330 was previously
identified to synthesize the beneficial metabolite conjugated linoleic
acid from free linoleic acid. The sequence will allow identification and
characterization of the genetic determinants of its putative beneficial
properties.

<>

<1>Guinane, C.M., Kent, R.M., Norberg, S., Hill, C., Fitzgerald, G.F., Stanton, C., Ross, R.P.
<2>Host Specific Diversity in Lactobacillus johnsonii as Evidenced by a Major Chromosomal Inversion and Phage Resistance Mechanisms.
<3>PLoS ONE
<4>6
<5>e18740
<6>2011
<7>Genetic diversity and genomic rearrangements are a driving force in bacterial evolution and
niche adaptation. We sequenced and annotated
the genome of Lactobacillus johnsonii DPC6026, a strain isolated from
the porcine intestinal tract. Although the genome of DPC6026 is similar
in size (1.97mbp) and GC content (34.8%) to the sequenced human isolate
L. johnsonii NCC 533, a large symmetrical inversion of approximately
750 kb differentiated the two strains. Comparative analysis among 12
other strains of L. johnsonii including 8 porcine, 3 human and 1
poultry isolate indicated that the genome architecture found in DPC6026
is more common within the species than that of NCC 533. Furthermore a
number of unique features were annotated in DPC6026, some of which are
likely to have been acquired by horizontal gene transfer (HGT) and
contribute to protection against phage infection. A putative type III
restriction-modification system was identified, as were novel Clustered
Regularly Interspaced Short Palindromic Repeats (CRISPR) elements.
Interestingly, these particular elements are not widely distributed
among L. johnsonii strains. Taken together these data suggest
intra-species genomic rearrangements and significant genetic diversity
within the L. johnsonii species and indicate towards a host-specific
divergence of L. johnsonii strains with respect to genome inversion and
phage exposure.

<>

<1>Guio, H., Tarazona, D., Galarza, M., Borda, V., Curitomay, R.
<2>Genome analysis of 17 extensively drug-resistant strains reveals new potential mutations for resistance.
<3>Genome Announcements
<4>2
<5>e00759-14
<6>2014
<7>We report the whole-genome sequence of an extensively drug-resistant (XDR) tuberculosis (TB)
strain of Latin American-Mediterranean (LAM) lineage. This
strain is phenotypically resistant to aminoglycosides, but carries no related
mutations in rrs, tlyA, and eis. Through genome analysis comparison with 16 XDR
strains, we found 218 non-synonymous single nucleotide polymorphisms (SNPs)
shared that could confer resistance.

<>

<1>Gumerov, V.M., Mardanov, A.V., Beletsky, A.V., Prokofeva, M.I., Bonch-Osmolovskaya, E.A., Ravin, N.V., Skryabin, K.G.
<2>Complete genome sequence of 'Vulcanisaeta moutnovskia' strain 768-28, a novel member of the hyperthermophilic crenarchaeal genus Vulcanisaeta.
<3>J. Bacteriol.
<4>193
<5>2355-2356
<6>2011
<7>Strain 768-28 was isolated from a hot spring in Kamchatka, Russia and represents a novel
member of the Vulcanisaeta genus. The complete genome sequence of this thermoacidophilic
anaerobic crenarchaeon reveals genes for protein and carbohydrate-active enzymes, the
Embden-Meyerhof and Entner-Doudoroff pathways for glucose metabolism, the tricarboxylic acid
cycle, beta-oxidation of fatty acids, and sulfate reduction.

<>

<1>Gumulak-Smith, J., Teachman, A., Tu, A.H., Simecka, J.W., Lindsey, J.R., Dybvig, K.
<2>Variations in the surface proteins and restriction enzyme systems of Mycoplasma pulmonis in the respiratory tract of infected rats.
<3>Mol. Microbiol.
<4>40
<5>1037-1044
<6>2001
<7>Restriction and modification (R-M) systems are generally thought to protect bacteria from
invasion by foreign DNA. This paper proposes the existence of an alternative role for the
phase-variable R-M systems encoded by the hsd loci of Mycoplasma pulmonis. Populations of M.
pulmonis cells that arose during growth in different environments were compared with respect
to R-M activity and surface antigen production. When M. pulmonis strain X1048 was propagated
in laboratory culture medium, > 95% of colony-forming units (cfu) lacked R-M activity and
produced the variable surface protein VsaA. Mycoplasmas isolated from the nose of
experimentally infected rats also lacked R-M activity and produced VsaA. In contrast, the cell
population of mycoplasmas isolated from the lower respiratory tract of the infected rats was
more complex. The most dramatic results were obtained for mycoplasmas isolated from the
trachea. At 14 days postinfection, 38% of mycoplasma isolates produced a Vsa protein other
than VsaA, and 34% of isolates had active restriction systems. These data suggest that
differences in selection pressures in animal tissues affect the surface proteins and the R-M
activity of the mycoplasmal cell population. We propose that variations in the production of
R-M activity and cell surface proteins are important for the survival of the mycoplasma within
the host.

<>

<1>Gunaletchumy, S.P., Teh, X., Khosravi, Y., Ramli, N.S., Chua, E.G., Kavitha, T., Mason, J.N., Lee, H.T., Alias, H., Zaidan, N.Z., Yassin, N.B., Tay, L.C., Rudd, S., Mitchell, H.M., Kaakoush, N.O., Loke, M.F., Goh, K.L., Vadivelu, J.
<2>Draft Genome Sequences of Helicobacter pylori Isolates from Malaysia, Cultured from Patients with Functional Dyspepsia and Gastric Cancer.
<3>J. Bacteriol.
<4>194
<5>5695-5696
<6>2012
<7>Helicobacter pylori is the main bacterial causative agent of gastroduodenal disorders and a
risk factor for gastric adenocarcinoma and mucosa-associated
lymphoid tissue (MALT) lymphoma. The draft genomes of 10 closely related H.
pylori isolates from the multiracial Malaysian population will provide an insight
into the genetic diversity of isolates in Southeast Asia. These isolates were
cultured from gastric biopsy samples from patients with functional dyspepsia and
gastric cancer. The availability of this genomic information will provide an
opportunity for examining the evolution and population structure of H. pylori
isolates from Southeast Asia, where the East meets the West.

<>

<1>Gunderson, K.
<2>Methods for detecting methylation state of CpG islands in genomic DNA using methylation-sensitive restriction endonuclease.
<3>US Patent Office
<4>US 20060134650 A
<5>
<6>2006
<7>The invention provides methods of identifying a plurality of reactive recognition sites for a
restriction endonuclease in genomic DNA.  In particular embodiments, the methods can be used
to identify methylation state of a plurality of CpG target sites in genomic DNA.  The method
can include steps of treating genomic DNA with a restriction endonuclease, thereby producing
genomic DNA fragments; ligating the fragments, thereby forming a concatenated DNA; and
identifying sequence portions of the concatenated DNA that are re-ordered compared to the
genomic DNA.

<>

<1>Gunn, J.S., Piekarowicz, A., Chien, R., Stein, D.C.
<2>Cloning and linkage analysis of Neisseria gonorrhoeae DNA methyltransferases.
<3>J. Bacteriol.
<4>174
<5>5654-5660
<6>1992
<7>We have cloned DNA methyltransferases (MTases) from various strains of Neisseria gonorrhoeae.
Each of these clones represents a single specificity, indicating that the multiple gonococcal
MTase specificities are encoded by monospecific MTases. The DNAs of five strains (FA5100, F62,
MS11, Pgh3-2, and WR302) were digested with NheI, SpeI, or NheI plus SpeI and subjected to
pulsed-field gel electrophoresis. The DNA MTase clones were used to probe Southern blots of
these pulsed-field gels to determine whether the MTase genes are linked and whether there are
stain-to-strain differences. The results indicate that none of these genes are closely linked,
but variable hybridization patterns indicate that there exist restriction fragment length
polymorphisms between the strains tested. Most of the chromosomal regions containing these
restriction fragment length polymorphisms are clustered in regions containing gonococcal genes
known or suspected to antigenically vary via genetic recombination.

<>

<1>Gunn, J.S., Stein, D.C.
<2>Natural variation of the NgoII restriction-modification of Neisseria gonorrhoeae.
<3>Gene
<4>132
<5>15-20
<6>1993
<7>The NgoII restriction-modification (R-M) system of Neisseria gonorrhoeae recognizes the
sequence 5'-GGCC-3'. This system is encoded by two separate genes, dcmB for the
methyltransferase (MTase) and dcrB for the restriction endonuclease (ENase). Three strains
that vary in their NgoII phenotype were examined. Strain Pgh3-2 produced detectable levels of
both enzymes, strain F62 lacked detectable levels of the dcrB gene product, and strain WR302
failed to produce either gene product. Strains that lacked either enzyme activity still
possessed the genes that encode them. Transcriptional fusions of dcrB in strains F62 and
Pgh3-2 indicate that this gene is transcribed at nearly identical levels in each strain. The
DNA encoding the NgoII R-M system was cloned from the three strains, and the nucleotide
sequence was determined. The dcrB genes of WR302 and F62 possess the same frameshift mutation
(base position 1435) which would result in a truncated protein. The WR302 dcmB was found to
have a point mutation that changed Arg 288 (a residue that is conserved in all prokaryotic and
phage cytosine MTases sequenced to date) to Trp.
[ The enzyme called NgoII in this abstract has been renamed NgoCII, Jan/1998. ]

<>

<1>Gunn, J.S., Stein, D.C.
<2>Use of a non-selective transformation technique to construct a multiply restriction/modification-deficient mutant of Neisseria gonorrhoeae.
<3>Mol. Gen. Genet.
<4>251
<5>509-517
<6>1996
<7>A technique that allows for easy identification of transformants of Neisseria gonorrhoeae in
the absence of selective pressure has been developed.  A suicide vector that contains a
gonococcal DNA uptake sequence was constructed to aid in DNA uptake.  In this transformation
procedure, a limiting number of cells is incubated with an excess amount of DNA, and the
mixture is plated onto a non-selective medium.  At least 20% of the resulting colonies
contained cells that had been transformed.  This strategy was utilized to construct specific
deletions of the S.NgoI, II, IV, V, and VII restriction-modification (R/M) genes.  All five
deletions were successfully incorporated into the chromosome of FA19, producing strain JUG029.
Strain JUG029 could be transformed with non-methylated plasmid DNA while strain FA19 could not
be transformed with such DNA.  The development of a simple, non-slective transformation
technique, coupled with the construction of a strain that is more permissive for DNA-mediated
transformation, will aid in genetic manipulations of the gonococcus.

<>

<1>Gunn, J.S., Stein, D.C.
<2>The Neisseria gonorrhoeae S.NgoVIII restriction/modification system: a type IIs system homologous to the Haemophilus parahaemolyticus HphI restriction/modification system.
<3>Nucleic Acids Res.
<4>25
<5>4147-4152
<6>1997
<7>Strains of Neisseria gonorrhoeae possess numerous restriction-modification systems.  One of
these systems, which has been found in all strains tested, encodes the S.NgoVIII specificity
(5' TCACC 3') R-M system.  We cloned two adjacent methyltransferase genes (dcmH and damH),
each encoding proteins whose actions protect DNA from digestion by R.HphI or R.NgoBI (%'
TCACC 3').  The damH gene product is a N6-methyladenine methyltransferase that recognizes
this sequence.  We constructed a plasmid containing multiple copies of the S.NgoVIII sequence,
grew it in the presence of damH and used the HPLC to demonstrate the presence of
N6-methyladenine in the DNA.  A second plasmid, containing overlapping damH and Escherichia
coli dam recognition sequences in combination with various restriction digests, was used to
identify which adenine in the recognition sequence was modified by damH.  The predicted dcmH
gene product is homologous to 5-methylcytosine methyltransferases.  The products of both the
dcmH and damH genes, as well as an open reading frame downstream of the damH gene are highly
similar to the Haemophilus parahaemolyticus hphIMC, hphIMA and hphIR gene products, encoding
the HphI type IIs R-M system.  The S.NgoVIII R-M genes are flanked by a 97 bp direct repeat
that may be involved in the mobility of this R-M system.

<>

<1>Gunthert, U., Freund, M., Trautner, T.A.
<2>Restriction and modification in Bacillus subtilis:  Two DNA methyltransferases with BsuRI specificity.  I.  Purification and Physical Properties.
<3>J. Biol. Chem.
<4>256
<5>9340-9345
<6>1981
<7>Two S-adenosyl-L-methionine: DNA (cytosine 5)-methyltransferases, termed
M.BsuRIa and M.BsuRIb, were purified 3,000- and 4,000-fold, respectively, from
Bacillus subtilis strain OG3R (r+m+) by successive column chromatography.  The
molecular weights determined by gel filtration were 37,000 for M.BsuRIa and
40,000 for M.BsuRIb.  The sedimentation coefficients s20,w were 3.55 for both
enzymes as determined by glycerol gradient centrifugation, corresponding to
molecular weights of 43,000.  Analysis of the two methyltransferases by agarose
gel electrophoresis, showed correspondence of the M.BsuRIa activity with one
protein band at a molecular weight of 41,000, whereas M.BsuRIb activity was
associated with two protein bands with molecular weights of 42,000 and 39,000,
respectively.

<>

<1>Gunthert, U., Jentsch, S., Freund, M.
<2>Restriction and modification in Bacillus subtilis:  Two DNA Methyltransferases with BsuRI specificity.  II.  Catalytic properties, substrate specificity, and mode of action.
<3>J. Biol. Chem.
<4>256
<5>9346-9351
<6>1981
<7>The properties of two DNA methyltransferases, termed M.BsuRIa and M.BsuRIb,
whose isolation was described in the preceding paper (Gunthert, U., Freund, M.,
and Trautner, T.A. (1981) J. Biol. Chem. 256: 9340-9345) were compared.  Both
enzymes recognize the same target sequence in double-stranded DNA, leading to
methylation of the internal cytosine:  5'GGC*C  The enzymes have identical
reaction constants with their substrates, DNA (km = 2.7 nM for the 5'GGCC
sequence), and S-adenosyl-L-methionine (km = 0.7 microM).  Initial rates of
methyl group transfer were proportional to enzyme concentration over a range of
50-fold, indicating absence of aggregation.  The enzymes are different in their
ionic strength requirements using Tris-HCl, pH 8.4.  M.BsuRIa is most active at
100 mM, M.BsuRIb at 440 mM.  As measured by incorporation kinetics and heat
inactivation, M.BsuRIa is the more stable enzyme of the two.  Equilibrium
dialysis was used to study the mode of methyl group transfer to the DNA with
either enzyme.  The data indicate that initially S-adenosyl-L-methionine binds
to methyltransferase.  This complex attaches to either modified or nonmodified
DNA.  The methyl group will then be transfered to a nonmodified target
sequence, leading to the disociation of enzyme and S-adenosyl-L-homocysteine
from the DNA.

<>

<1>Gunthert, U., Lauster, R., Reiners, L.
<2>Multispecific DNA methyltransferases from Bacillus subtilis phages Properties of wild-type and various mutant enzymes with altered DNA affinity.
<3>Eur. J. Biochem.
<4>159
<5>485-492
<6>1986
<7>Temperate Bacillus subtilis phages SPR, Phi3T, d11 and SPbeta code for DNA
methyltransferases, each having mutliple sequence specificities.  The SPR
wild-type and various mutant methyltransferases were overproduced 1000-fold in
Escherichia coli and were purified by three consecutive chromatographic steps.
The stable form of these multispecific enzymes in solution are monomers with a
relative molecular mass (Mr) of about 50,000.  The methyl-transfer kinetics of
the SPR wild-type and mutant enzymes were determined with DNA substrates
carrying either none or one of the three recognition sequences (GGCC, CCGG,
CCA/TGG).  Evaluation of the catalytic properties for DNA and
S-adenosylmethionine binding suggested that the NH2-terminal part of the
protein is important for both non-sequence-specific DNA binding and
S-adenosylmethionine binding as well as transfer of methyl groups.  On the
other hand, mutations in the COOH-terminal part lead to weaker site-specific
interactions of the enzyme.  Antibodies raised against the purified SPR enzyme
specifically immunoprecipitated the Phi3T,d11 and SPbeta methyltransferases,
but failed to precipitate the chromosomally coded enzymes from B. subtilis
(BsuRI) and B. sphaericus (BspRI).  Immunoaffinity chromatography is an
efficient purification step for the related phage methyltransferases.

<>

<1>Gunthert, U., Pawlek, B., Stutz, J., Trautner, T.A.
<2>Restriction and Modification in Bacillus subtilis: Inducibility of a DNA Methylating Activity in Nonmodifying Cells.
<3>J. Virol.
<4>20
<5>188-195
<6>1976
<7>The nonrestricting/nonmodifying strain Bacillus subtilis 222 (4-m-) can be
induced to synthesize a DNA-modifying activity upon treatment with either
mitomycin C (MC) or UV light.  This is shown by the following facts.  (i)
Infection of MC-pretreated 222 cells with unmodified SPP1 phage yields about 3%
modified phage that are resistant to restriction in B. subtilis R (r+m+).  The
induced modifying activity causes the production of a small fraction of fully
modified phage in a minority class of MC-treated host cells.  (ii) The
MC-pretreated host cells contain a DNA cytosine methylating activity; both
bacterial and phage DNAs have elevated levels of 5-methylcytosine.  (iii) The
MC-induced methylation of SPP1 DNA takes place at the recognition nucleotide
sequences of restriction endonuclease R from B. subtilis R.  (iv) Crude
extracts of MC-pretreated 222 cells have enhanced DNA methyltransferase
activities, with a substrate specificity similar to that found in modification
enzymes present in (constituitively) modifying strains.

<>

<1>Gunthert, U., Reiners, L.
<2>Bacillus subtilis phage SPR codes for a DNA methyltransferase with triple sequence specificity.
<3>Nucleic Acids Res.
<4>15
<5>3689-3701
<6>1987
<7>SPR, a temperate Bacillus subtilis phage, codes for a DNA methyltransferase
that can methylate the sequences GGCC (or GGCC) and CCGG at the cytosines
indicated.  We show here that it can also methylate the sequence CC(A/T)GG and
protect it from cleavage with EcoRII and ApyI.  This methylation can be seen in
vivo as well as in vitro with purified SPR methyltransferase.  SPR19 and SPR83
are two mutant phages, defective in GGCC or CCGG methylation, respectively.
These mutants have not lost their ability to methylate CC(A/T)GG sites.
Mutation SPR26 has lost the ability to methylate all three sites.  Thus the SPR
methyltransferase codes for three genetically distinguishable methylation
abilities.

<>

<1>Gunthert, U., Reiners, L., Lauster, R.
<2>Cloning and expression of Bacillus subtilis phage DNA methyltransferase genes in Escherichia coli and B. subtilis.
<3>Gene
<4>41
<5>261-270
<6>1986
<7>The DNA methyltransferase (Mtase) genes of the temperate Bacillus subtilis
phages SPR (wild type and various mutants), Phi3T, p11 and SBb have been cloned
and expressed in Escherichia coli and B. subtilis host-plasmid vector systems.
Mtase activity has been quantitated in these clones by performing in vitro
methylation assays of cell-free extracts.  The four-phage Mtase genes differ in
the amount of Mtase synthesized when transcribed from their genuine promoters.
In B. subtilis as well as in E. coli the SPR Mtase is always produced in
smaller amounts than the other phage Mtases.  Expression levels of the SPR
Mtase are dependent on the strength of the upstream vector promoter sequences.
Overproduction of the SPR wild-type and mutant enzymes was achieved in E. coli
(inducible expression) by fusions to the lambda pL or the tac promoter and in
B. subtilis (constitutive expression) by means of the phage SP02 promoter.

<>

<1>Gunthert, U., Storm, K., Bald, R.
<2>Restriction and modification in Bacillus subtilis. Localization of the methylated nucleotide in the BsuRI recognition sequence.
<3>Eur. J. Biochem.
<4>90
<5>581-583
<6>1978
<7>Calf thymus DNA was methylated in vitro with cell extracts of Bacillus subtilis OG3R (r+m+)
and S-adenosyl[Me-3H]methionine.  After depurination of the [3H]methylated DNA, the analysis
of the pyrimidine dinucleotides revealed the following positions of the methylated nucleosides
(indicated by an asterisk) with the BsuRI recognition sequence:
5' dG-dG-dC*-dC
dC-dC*-dG-dG 5'.

<>

<1>Gunthert, U., Stutz, J., Klotz, G.
<2>Restriction and modification in B. subtilis.
<3>Mol. Gen. Genet.
<4>142
<5>185-191
<6>1975
<7>The content of 5-methylcytosine (5MC) and 6-methyladenine (6MA) in modified and
nonmodified DNAs from B. subtilis and B. subtilis phage SPP1 were determined.
Non-modified SPP1-O DNA contains about 15 5MC residues/molecule.  Each modified
SPP1-R DNA molecule carries 190 modification specific methyl groups.  This
number is sufficient to account for modification of the 80 restriction sites in
SPP1 DNA (Bron and Murray, 1975) against endo R.BsuR, assuming each modified
site contains two 5MC residues.  Resistance of SPO1 DNA against endo R.BsuR
restriction both in vivo and in vitro is probably not due to methylation of
endo R.BsuR recognition sites.

<>

<1>Gunthert, U., Trautner, T.A.
<2>DNA methyltransferases of Bacillus subtilis and its bacteriophages.
<3>Curr. Top. Microbiol. Immunol.
<4>108
<5>11-22
<6>1984
<7>Postreplicative DNA methylation occurs with a high incidence in bacteria and may affect
resident DNA and that of infecting bacteriophages. By far the most widespread role of DNA
methylation is to provide the DNA with "modification" i.e., protection against the
endonucleolytic attack of cellular restriction enzymes. Important aspects of this role in
connection with type-I enzymes are discussed in the review of Suri et al. (this volume).
Another well-studied physiological function of DNA methylation observed in Escherichia coli is
its role in strand recognition for proofreading during DNA synthesis, which is covered in the
review by Radman and Wagner (this volume). Some role of host-mediated DNA methylation is
implemented in the regulation of gene expression of bacteriophage mu; this work is the subject
of the review by Kahmann (this volume).

<>

<1>Guo, H., Karberg, M., Long, M., Jones, J.P. III, Sullenger, B., Lambowitz, A.M.
<2>Group II introns designed to insert into therapeutically relevant DNA target sites in human cells.
<3>Science
<4>289
<5>452-457
<6>2000
<7>Mobile group II intron RNA's insert directly into DNA target sites and are then
reverse-transcribed into genomic DNA by the associated intron-encoded protein.  Target site
recognition involves modifiable base-pairing interactions between the intron RNA and a
>14-nucleotide region of the DNA target site, as well as fixed interactions between the
protein and flanking regions.  Here, we developed a highly efficient Escherichia coli genetic
assay to determine detailed target site recognition rules for the Lactococcus lactis group II
intron Ll.LtrB and to select introns that insert into desired target sites.  Using human
immunodeficiency virus-type 1 (HIV-1) proviral DNA and the human CCR5 gene as examples, we
show that group II introns can be retargeted to insert efficiently into virtually any target
DNA and that the retargeted introns retain activity in human cells.  This work provides the
practical basis for potential applications of targeted group II introns in genetic
engineering, functional genomics, and gene therapy.

<>

<1>Guo, H., Zimmerly, S., Perlman, P.S., Lambowitz, A.M.
<2>Group II intron endonucleases use both RNA and protein subunits for recognition of specific sequences in double-stranded DNA.
<3>EMBO J.
<4>16
<5>6835-6848
<6>1997
<7>Group II introns use intron-encoded reverse transcriptase, maturase and DNA endonuclease
activities for site-specific insertion into DNA.  Remarkably, the endonucleases are
ribonucleoprotein complexes in which the excised intron RNA cleaves the sense strand of the
recipient DNA by reverse splicing, while the intron-encoded protein cleaves the antisense
strand.  Here, studies with the yeast group II intron aI2 indicate that both the RNA and
protein components of the endonuclease contribute to recognition of an ~30 bp DNA target site.
Our results lead to a model in which the protein component first recognizes specific
nucleotides in the most distal 5' exon region of the DNA target site (E2-21 to -11).  Binding
of the protein then leads to DNA unwinding, enabling the intron RNA to base pair to a 13
nucleotide DNA sequence (E2-12 to E3+1) for reverse splicing.  Antisense-strand cleavage
requires additional interactions of the protein with the 3' exon DNA (E3 +1 to +10).  Our
results show how enzymes can use RNA and protein subunits cooperatively to recognize specific
sequences in double-stranded DNA.

<>

<1>Guo, H.-C.
<2>Type II restriction endonucleases: Structures and applications.
<3>Recent Res. Devel. Macromol.
<4>7
<5>225-245
<6>2003
<7>Restriction endonucleases are enzymes that recognize specific double-strand DNA sequences and
cleave at a defined point within or close to that sequence.  These enzymes have become
indispensable tools in modern biochemistry, recombinant DNA technology, genome mapping, and
genetic manipulation.  Diverse recognition strategies and high specificity make restriction
enzymes an ideal system for structural studies of DNA recognition and cleavage by proteins.
For various applications such as DNA fingerprinting to study cancer or infectious diseases,
there have also been long-standing interests in rational design of artificial restriction
enzymes with desired specificities.  However, restriction enzymes have proven to be a
difficult case for protein engineering.  In general initial attempts were not successful,
mainly because the tight coupling of specific binding to catalysis was not fully understood.
New structures reported reveal that these enzymes are more diverse than expected.  Comparisons
of twelve available structures of Type II restriction enzyme/DNA complexes indicate that
structural elements responsible for allosteric coupling of recognition to catalysis, as well
as dimer structures, correlate well with their cleavage patterns on DNA.  It is thus likely
that enzymes in the existing pool with different cleavage patterns reflect different control
mechanisms that couple DNA recognition to DNA cleavage and/or dimer structures.  A systematic
study of restriction enzymes with unique DNA cleavage patterns may illuminate alternative
coupling mechanisms suitable for manipulation in order to create tailor-made restriction
enzymes with desired specificities.

<>

<1>Guo, H.T., Xu, X.D.
<2>Broad host range plasmid-based gene transfer system in the cyanobacterium Gloeobacter violaceus which lacks thylakoids.
<3>Prog. Nat. Sci.
<4>14
<5>31-35
<6>2004
<7>Gloeobacter violaceus, a cyanobacterium lack of thylakoids, is refractory to genetic
manipulations because its cells are enveloped by
a thick gelatinous sheath and in colonial form. In this study, a large
number of single cells were obtained by repeated pumping with a syringe
with the gelatinous sheath removed. And an exogenous broad host range
plasmid pKT210 was conjugatively transferred into G. violaceus.
Analyses with dot-blot hybridization and restriction mapping showed
that the exogenous plasmid pKT210 had been introduced into G. violaceus
and stably maintained with no alteration in its structure. pKT210
extracted from G. violaceus exconjugants could be transformed into the
mcr - mrr - E. coli strain DH10B but not the mcr(+) mrr(+) strain
DH5alpha, which suggests that a methylase system may be present in G.
violaceus.

<>

<1>Guo, J., Gaj, T., Barbas, C.F.I.I.I.
<2>Directed Evolution of an Enhanced and Highly Efficient FokI Cleavage Domain for Zinc Finger Nucleases.
<3>J. Mol. Biol.
<4>400
<5>96-107
<6>2010
<7>Zinc finger nucleases (ZFNs) are powerful tools for gene therapy and genetic engineering. The
high specificity and affinity of these chimeric
enzymes are based on custom-designed zinc finger proteins (ZFPs). To
improve the performance of existing ZFN technology, we developed an in
vivo evolution-based approach to improve the efficacy of the FokI cleavage
domain (FCD). After multiple rounds of cycling mutagenesis and DNA
shuffling, a more efficient nuclease variant (Sharkey) was generated. In
vivo analyses indicated that Sharkey is >15-fold more active than
wild-type FCD on a diverse panel of cleavage sites. Further, a mammalian
cell-based assay showed a three to sixfold improvement in targeted
mutagenesis for ZFNs containing derivatives of the Sharkey cleavage
domain. We also identified mutations that impart sequence specificity to
the FCD that might be utilized in future studies to further refine ZFNs
through cooperative specificity. In addition, Sharkey was observed to
enhance the cleavage profiles of previously published and newly selected
heterodimer ZFN architectures. This enhanced and highly efficient cleavage
domain will aid in a variety of ZFN applications in medicine and biology.

<>

<1>Guo, M., Han, X., Jin, T., Zhou, L., Yang, J., Li, Z., Chen, J., Geng, B., Zou, Y., Wan, D., Li, D., Dai, W., Wang, H., Chen, Y., Ni, P., Fang, C., Yang, R.
<2>Genome sequences of three species in the family planctomycetaceae.
<3>J. Bacteriol.
<4>194
<5>3740-3741
<6>2012
<7>Most of the species in the family Planctomycetaceae are of interest for their eukaryotic-like
cell structures and characteristics of resistance to extreme
environments. Here, we report draft genome sequences of three aquatic parasitic
species of this family, Singulisphaera acidiphila (DSM 18658T), Schlesneria
paludicola (DSM 18645T), and Zavarzinella formosa (DSM 19928T).

<>

<1>Guo, Q., Li, S., Lu, X., Zhang, X., Wang, P., Ma, P.
<2>Complete Genome Sequence of Bacillus subtilis BAB-1, a Biocontrol Agent for Suppression of Tomato Gray Mold.
<3>Genome Announcements
<4>2
<5>e00744-14
<6>2014
<7>Bacillus subtilis BAB-1, isolated from cotton rhizosphere soil, is an excellent biocontrol
agent for tomato gray mold. The genome of B. subtilis strain BAB-1 was
fully sequenced and annotated, genes encoding the antifungal active compound were
identified, and multiple sets of regulatory systems were found in the genome.

<>

<1>Guo, S., Mao, Z., Wu, Y., Hao, K., He, P., He, Y.
<2>Genome Sequencing of Bacillus subtilis Strain XF-1 with High Efficiency in the Suppression of Plasmodiophora brassicae.
<3>Genome Announcements
<4>1
<5>e0006613
<6>2013
<7>The genome of the rhizobacterium Bacillus subtilis XF-1 is 4.06 Mb in size and harbors 3,853
coding sequences (CDS). Giant gene clusters were dedicated to the
nonribosomal synthesis of antimicrobial lipopeptides and polyketides. Remarkably,
XF-1 possesses a gene cluster involved in the synthesis of chitosanase that is
related to the suppression of the pathogen Plasmodiophora brassicae.

<>

<1>Guo, W., Wang, Y., Song, C., Yang, C., Li, Q., Li, B., Su, W., Sun, X., Song, D., Yang, X., Wang, S.
<2>Complete genome of Pseudomonas mendocina NK-01, which synthesizes medium-chain-length polyhydroxyalkanoates and alginate oligosaccharides.
<3>J. Bacteriol.
<4>193
<5>3413-3414
<6>2011
<7>Pseudomonas mendocina NK-01 can synthesize medium-chain-length polyhydroxyalkanoate (PHA(MCL))
and alginate oligosaccharides (AO)
simultaneously from glucose in the condition of limited nitrogen source.
Here we report the complete sequence of the 5.4-Mbp genome of Pseudomonas
mendocina NK-01, that was isolated from farmland soil in Tianjin, China.

<>

<1>Guo, Y., Cen, Z., Zou, Y., Fang, X., Li, T., Wang, J., Chang, D., Su, L., Liu, Y., Chen, Y., Yang, R., Liu, C.
<2>Whole-Genome Sequence of Klebsiella pneumonia Strain LCT-KP214.
<3>J. Bacteriol.
<4>194
<5>3281
<6>2012
<7>Klebsiella pneumoniae is a Gram-negative, nonmotile, encapsulated, lactose-fermenting,
facultative anaerobic, rod-shaped bacterium found in the
normal flora of the mouth, skin, and intestines. Here we present the fine-draft
genome sequence of K. pneumoniae strain LCT-KP214, which originated from K.
pneumoniae strain CGMCC 1.1736.

<>

<1>Guo, Y., Jiao, Z., Li, L., Wu, D., Crowley, D.E., Wang, Y., Wu, W.
<2>Draft Genome Sequence of Rahnella aquatilis Strain HX2, a Plant Growth-Promoting  Rhizobacterium Isolated from Vineyard Soil in Beijing, China.
<3>J. Bacteriol.
<4>194
<5>6646-6647
<6>2012
<7>Rahnella aquatilis strain HX2 is a plant growth-promoting, disease-suppressive rhizobacterium
that was isolated from a vineyard soil in Beijing, China. Here, we
report the genome sequence of this strain, which provides a valuable resource for
future research examining the mechanisms of traits associated with plant growth
promotion and biocontrol.

<>

<1>Guo, Y., Wang, H., Li, Y., Song, Y., Chen, C., Liao, Y., Ren, L., Guo, C., Tong, W., Shen, W., Chen, M., Mao, X., Guo, G., Zou, Q.
<2>Genome of Helicobacter pylori Strain XZ274, an Isolate from a Tibetan Patient with Gastric Cancer in China.
<3>J. Bacteriol.
<4>194
<5>4146-4147
<6>2012
<7>The infection rate of Helicobacter pylori is high all over the world, especially  in the
Chinese Tibetan Plateau. Here, we report the genome sequence of
Helicobacter pylori strain XZ274 isolated from a Tibetan patient with gastric
cancer. The strain contains 1,634,138 bp with 1,654 coding sequences and a pXZ274
plasmid of 22,406 bp with 26 coding sequences. This is the first complete genome
sequence of Helicobacter pylori from the Tibetan Plateau in China.

<>

<1>Guo, Z., Chen, P., Ren, P., Kuang, S., Zhou, Z., Li, Z., Liu, M., Shi, D., Xiao, Y., Wang, X., Zhou, R., Jin, H., Bi, D.
<2>Genome Sequence of Duck Pathogen Mycoplasma anatis Strain 1340.
<3>J. Bacteriol.
<4>193
<5>5883-5884
<6>2011
<7>Mycoplasma anatis, a member of the class Mollicutes, is the causative agent of a contagious
infectious disease of domestic ducklings, wild
birds, and eggs. Increasing reports show that coinfection of M. anatis
with Escherichia coli results in substantial economic impacts on the duck
farms in China. Here, we announce the first genome sequence of M. anatis.

<>

<1>Guo, Z., Xu, X., Zheng, Q., Li, T., Kuang, S., Zhang, Z., Chen, Y., Lu, X., Zhou, R., Bi, D., Jin, H.
<2>Genome Sequence of Mycoplasma columbinum Strain SF7.
<3>Genome Announcements
<4>1
<5>e00157-13
<6>2013
<7>Mycoplasma columbinum is a member of nonglycolytic Mycoplasma species which can hydrolyze
arginine. Increasingly research has revealed that M. columbinum is
associated with respiratory disease of pigeons and that the respiratory disease
symptoms could be eliminated via the use of mycoplasma treatment medicine. Here
we report the genome sequence of M. columbinum strain SF7, which is the first
genome report for M. columbinum.

<>

<1>Guolin, A., Shiwen, L., Yong, Z., Mingyi, Q., Tong, Z.
<2>Effects of organic solvents on the specificity and activity of restriction endonuclease Bsp63I and Bsp78I.
<3>J. Wuhan Univ.
<4>3
<5>135-138
<6>1991
<7>
<>

<1>Gupta, H.K., Gupta, R.D., Singh, A., Chauhan, N.S., Sharma, R.
<2>Genome Sequence of Rheinheimera sp. Strain A13L, Isolated from Pangong Lake, India.
<3>J. Bacteriol.
<4>193
<5>5873-5874
<6>2011
<7>Rheinheimera sp. strain A13L, which has antimicrobial activity, was isolated from alkaline
brackish water of the high-altitude Pangong Lake of
Ladakh, India. Here we report the draft genome sequence of Rhienheimera
sp. strain A13L (4,523,491 bp with a G+C content of 46.23%). The genome is
predicted to contain genes for marinocine and colicin V production, which
may be responsible for the antimicrobial activity of the strain.

<>

<1>Gupta, R., Capalash, N., Sharma, P.
<2>Restriction endonucleases: natural and directed evolution.
<3>Appl. Microbiol. Biotechnol.
<4>94
<5>583-599
<6>2012
<7>Type II restriction endonucleases (REs) are highly sequence-specific compared with other
classes of nucleases. PD-(D/E)XK nucleases,
initially represented by only type II REs, now comprise a large and
extremely diverse superfamily of proteins and, although sharing a
structurally conserved core, typically display little or no detectable
sequence similarity except for the active site motifs. Sequence
similarity can only be observed in methylases and few isoschizomers. As
a consequence, REs are classified according to combinations of
functional properties rather than on the basis of genetic relatedness.
New alignment matrices and classification systems based on structural
core connectivity and cleavage mechanisms have been developed to
characterize new REs and related proteins. REs recognizing more than
300 distinct specificities have been identified in RE database (REBASE:
http://rebase.neb.com/cgi-bin/statlist) but still the need for newer
specificities is increasing due to the advancement in molecular biology
and applications. The enzymes have undergone constant evolution through
structural changes in protein scaffolds which include random mutations,
homologous recombinations, insertions, and deletions of coding DNA
sequences but rational mutagenesis or directed evolution delivers
protein variants with new functions in accordance with defined
biochemical or environmental pressures. Redesigning through random
mutation, addition or deletion of amino acids, methylation-based
selection, synthetic molecules, combining recognition and cleavage
domains from different enzymes, or combination with domains of
additional functions change the cleavage specificity or substrate
preference and stability. There is a growing number of patents awarded
for the creation of engineered REs with new and enhanced properties.

<>

<1>Gupta, R., Vakhlu, J., Agarwal, A., Nilawe, P.D.
<2>Draft Genome Sequence of Plant Growth-Promoting Bacillus amyloliquefaciens Strain W2 Associated with Crocus sativus (Saffron).
<3>Genome Announcements
<4>2
<5>e00862-14
<6>2014
<7>Bacillus sp. strain W2 is a plant growth-promoting rhizobacterium isolated from saffron fields
of Kashmir, India. Here, we report the draft genome sequence (3.9
Mb) of Bacillus amyloliquefaciens strain W2 having 65 contigs (3, 997, 511 bp),
4,163 coding sequences, and an average 46.45% GC content. Despite the 99%
identity of the 16S rRNA gene with that of Bacillus amyloliquefaciens subsp.
plantarum FZB42, the genome comparison revealed that only 48.7% of the W2 genome
has homology with that of FZB42.

<>

<1>Gupta, R., Xu, S.-Y., Sharma, P., Capalsh, N.
<2>Characterization of MspNI (G/GWCC) and MspNII (R/GATCY), novel thermostable Type II restriction endonucleases from Meiothermus sp., isoschizomers of AvaII and BstYI.
<3>Mol. Biol. Rep.
<4>39
<5>5607-5614
<6>2012
<7>MspNI and MspNII, isoschizomers of prototype Type II restriction endonucleases AvaII and
BstYI, were extracted from an extreme thermophile bacterium belonging to the genus
Meiothermus, isolated from the hot sulphur springs in north Himalayan region of India where
temperature and pH ranged from 60 to 80C and 7.5 to 8.5, respectively. The two enzymes were
purified to homogeneity using Cibacron-Blue 3GA Agarose, Q-Sepharose and SP-Sepharose
chromatography and were homodimers with subunit molecular weights of 27 and 45 kDa,
respectively. Restriction mapping and run-off sequencing of MspNI and MspNII cleaved pBR322
DNA showed that they recognized and cleaved 5'-G/GWCC-3' and 5'-R/GATCY-3' sites,
respectively.
MspNI and MspNII worked optimally at 60 and 70C, 6 and 5 mM MgCl(2), respectively and showed
no star activity in organic solvents. Both were resistant to sequence methylation and were
stable up to 25 PCR cycles.

<>

<1>Gupta, Y.K., Yang, L., Chan, S.H., Samuelson, J.C., Xu, S.Y., Aggarwal, A.K.
<2>Structural Insights into the Assembly and Shape of Type III Restriction-Modification (R-M) EcoP15I Complex by Small-Angle X-ray Scattering.
<3>J. Mol. Biol.
<4>420
<5>261-268
<6>2012
<7>EcoP15I is the prototype of the Type III restriction enzyme family, composed of two
modification (Mod) subunits to which two (or one)
restriction (Res) subunits are then added. The Mod subunits are
responsible for DNA recognition and methylation, while the Res subunits
are responsible for ATP hydrolysis and cleavage. Despite extensive
biochemical and genetic studies, there is still no structural
information on Type III restriction enzymes. We present here
small-angle X-ray scattering (SAXS) and analytical ultracentrifugation
analysis of the EcoP15I holoenzyme and the Mod(2) subcomplex. We show
that the Mod(2) subcomplex has a relatively compact shape with a radius
of gyration (R-G) of similar to 37.4 angstrom and a maximal dimension
of similar to 110 angstrom. The holoenzyme adopts an elongated crescent
shape with an R-G of similar to 65.3 angstrom and a maximal dimension
of similar to 218 angstrom. From reconstructed SAXS envelopes, we
postulate that Mod(2) is likely docked in the middle of the holoenzyme
with a Res subunit at each end. We discuss the implications of our
model for EcoP15I. action, whereby the Res subunits may come together
and form a 'sliding clamp' around the DNA.

<>

<1>Guray, A., Erarslan, A., Ozbilen, O.
<2>A buffer system suitable for most restriction enzymes.
<3>DOGA TU J. Biol.
<4>13
<5>14-17
<6>1989
<7>For the digestion of DNA samples by several restriction enzymes, the use of a
single potassium glutamate buffer instead of different buffers recommended by
the manufacturers was investigated.  Almost all of the restriction enzymes that
we have tested to digest lambda DNA, showed the same activity with both buffer
systems.  Thus the convenience of potassium glutamate buffer was clearly
understood to be used with most restriction enzymes in place of the buffers
recommended by the vendors.

<>

<1>Guschlbauer, W.
<2>The DNA and S-adenosylmethionine-binding regions of EcoDam and related methyltransferases.
<3>Gene
<4>74
<5>211-214
<6>1988
<7>Previous comparison of the amino acid sequences of the GATC-methylating Escherichia coli Dam
methyltransferase (MTase) with those of other adenine MTases (M.EcoRV, M.DpnII and T4Dam)
localized four conserved regions. Regions III and IV have similarities with many other MTases.
The sequence DPPY (or NPPY) is always present in region IV. It was suggested to be the AdoMet
binding site. Publication of the nucleotide and amino acid sequences of M.CviBIII, M.DpnA and
MutH give further credence to this assignment: M.DpnA, which also methylates GATC, has strong
similarities with regions III and IV; M.CviBIII, a cytosine methylase, has a characteristic
NPPY sequence in region IV, and only limited resemblance in region III; MutH, the
GATC-specific endonuclease in DNA mismatch repair, has significant similarities uniquely in
region III. The presently available evidence suggests that region III is the GAT(C) binding
site and region IV is the AdoMet binding site. This hypothesis is strengthened by recent
genetic findings.

<>

<1>Guseinov, O.A., Bogdarina, I.G., Kossykh, V.G., Buryanov, Y.I., Baev, A.A.
<2>Sensitivity of chromosomal and plasmid E. coli DNA to restriction endonuclease EcoRII.
<3>Biokhimiia
<4>43
<5>1718-1720
<6>1978
<7>It was shown that E. coli C, E. coli MRE 600 DNA, and also plasmid DNA of Col E1, RSF 2124
from E. coli K-12, and plasmid DNA from E. coli MRE 600 were completely resistant against
restriction endonuclease R. EcoRII.  Plasmid DNA's of Col E1, RSF 2124 amplified for 4 hours
in the presence of chloramphenicol are sensitive to R.EcoRII but after 16-hour amplification
in the presence of chloramphenicol these DNA's acquire complete resistance against R.EcoRII.
These data point to the slower rate of modification of DNA in vivo by DC-methylases of EcoRII
type in comparison with DNA methylase EcoRII.

<>

<1>Guss, A.M., Olson, D.G., Caiazza, N.C., Lynd, L.R.
<2>Dcm methylation is detrimental to plasmid transformation in Clostridium thermocellum.
<3>Biotechnol. Biofuels.
<4>5
<5>30
<6>2012
<7>Background: Industrial production of biofuels and other products by cellulolytic
microorganisms is of interest but hindered by the nascent
state of genetic tools. Although a genetic system for Clostridium
thermocellum DSM1313 has recently been developed, available methods
achieve relatively low efficiency and similar plasmids can transform C.
thermocellum at dramatically different efficiencies.
Results: We report an increase in transformation efficiency of C.
thermocellum for a variety of plasmids by using DNA that has been
methylated by Escherichia coli Dam but not Dcm methylases. When
isolated from a dam + dcm + E. coli strain, pAMG206 transforms C.
thermocellum 100-fold better than the similar plasmid pAMG205, which
contains an additional Dcm methylation site in the pyrF gene. Upon
removal of Dcm methylation, transformation with pAMG206 showed a four-
to seven-fold increase in efficiency; however, transformation
efficiency of pAMG205 increased 500-fold. Removal of the Dcm
methylation site from the pAMG205 pyrF gene via silent mutation
resulted in increased transformation efficiencies equivalent to that of
pAMG206. Upon proper methylation, transformation efficiency of plasmids
bearing the pMK3 and pB6A origins of replication increased ca. three
orders of magnitude.
Conclusions: E. coli Dcm methylation decreases transformation
efficiency in C. thermocellum DSM1313. The use of properly methylated
plasmid DNA should facilitate genetic manipulation of this industrially
relevant bacterium.

<>

<1>Guthrie, E., Meda, M.
<2>Method for producing and cloning SacII restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0475195 B
<5>
<6>1995
<7>The present invention is directed to a method for cloning and producing the SacII restriction
endonuclease by 1) introducing the restriction endonuclease gene from Streptomyces
achromogenes into a host whereby the restriction gene is expressed; 2) fermenting the host
which contains the plasmid encoding and expressing the SacII restriction endonuclease
activity, and 3) purifying the SacII restriction endonuclease from the fermented host which
contains the plasmid encoding and expressing the SacII restriction endonuclease activity.

<>

<1>Guthrie, E.P.
<2>Isolated DNA encoding the SphI restriction endonuclease and related methods for producing the same.
<3>US Patent Office
<4>US 5262318
<5>
<6>1993
<7>The present invention is directed to a method for cloning and producing the SphI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Streptomyces
phaeochromogenes into a host whereby the restriction gene is expressed; 2) fermenting the host
which contains the plasmid encoding and expressing the SphI restriction endonuclease activity,
and 3) purifying the SphI restriction endonuclease from the fermented host which contains the
plasmid encoding and expressing the SphI restriction endonuclease activity.

<>

<1>Guthrie, E.P., Meda, M.M.
<2>Method for producing and cloning SacII restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5288696
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the SacII restriction
endonuclease by 1) introducing the restriction endonuclease gene from Streptomyces
achromogenes into a host whereby the restriction gene is expressed; 2)fermenting the host
which contains the plasmid encoding and expressing the SacII restriction endonuclease
activity, and 3) purifying the SacII restriction endonuclease from the fermented host which
contains the plasmid encoding and expressing the SacII restriction endonuclease activity.

<>

<1>Guthrie, E.P., Quinton-Jager, T., Moran, L.S., Slatko, B.E., Kucera, R.B., Benner, J.S., Wilson, G.G., Brooks, J.E.
<2>Cloning, expression and sequence analysis of the SphI restriction-modification system.
<3>Gene
<4>180
<5>107-112
<6>1996
<7>SphI, a type II restriction-modification system from the bacterium Streptomyces
phaeochromogenes, recognizes the sequence 5'-GCATGC.  The SphI methyltransferase encoding
gene, sphIM, was cloned into Escherichia coli using Mtase selection to isolate the clone.
However, none of these clones contained the restriction endonuclease gene.  Repeated attempts
to clone the complete Enase gene along with sphIM in one step failed, presumably due to
expression of SphI Enase gene, sphIR, in the presence of inadequate expression of sphIM.  The
complete sphIR was finally cloned using a two-step process.  PCR was used to isolate the 3'
end of sphIR from a library.  The intact sphIR, reconstructed under control of an inducible
promoter, was introduced into an E. coli strain containing a plasmid with the NlaIII
Mtase-encoding gene (nlaIIIM).  The nucleotide sequence of the SphI system was determined,
analyzed and compared to previously sequenced R-M systems.  The sequence was also examined for
features which would help explain why sphIR unlike other actinomycete Enase genes seemed to be
expressed in E. coli.

<>

<1>Guthrie, E.P., Van Cott, E.M., Taron, C.H.
<2>Method for cloning and producing the NaeI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5292651
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the NaeI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Nocardia aerocolonigenes
into a host whereby the restriction gene is expressed; 2) fermenting the host which contains
the plasmid encoding and expressing the NaeI restriction endonuclease activity, and 3)
purifying the NaeI restriction endonuclease from the fermented host which contains the plasmid
encoding and expressing the NaeI restriction endonuclease activity.

<>

<1>Guthrie, E.P., Van Cott, E.M., Taron, C.H.
<2>Method for cloning and producing the NaeI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0577254
<5>
<6>1997
<7>The present invention relates to recombinant DNA which encodes the NaeI restriction
endonuclease and modification methylase, and the production of these enzymes from the
recombinant DNA.

<>

<1>Gutierrez, G.
<2>Draft Genome Sequence of Methanobacterium formicicum DSM 3637, an Archaebacterium Isolated from the Methane Producer Amoeba Pelomyxa palustris.
<3>J. Bacteriol.
<4>194
<5>6967-6968
<6>2012
<7>Here is reported the draft genome sequence of Methanobacterium formicicum DSM 3637, which was
isolated from the methane-producing amoeba Pelomyxa palustris.
This bacterium was determined to be an endosymbiont living in the cytoplasm of P.
palustris and the source of methane; however, the global characteristics of its
genome suggest a free-living lifestyle rather than an endosymbiotic one.

<>

<1>Gutjahr, A., Xu, S.Y.
<2>Engineering nicking enzymes that preferentially nick 5-methylcytosine-modified DNA.
<3>Nucleic Acids Res.
<4>42
<5>e77
<6>2014
<7>N.Gamma is a strand-specific and site-specific DNA nicking enzyme (YCG downward arrowGT or AC
upward arrowCGR). Here we describe the isolation of single and
double mutants of N.Gamma with attenuated activity. The nicking domains (NDs) of
E59A and 11 double mutants were fused to the 5mCG-binding domain of MBD2 and
generated fusion enzymes that preferentially nick 5mCG-modified DNA. The CG
dinucleotide can be modified by C5 methyltransferases (MTases) such as M.SssI,
M.HhaI or M.HpaII to create composite sites AC upward arrowYGG N(8-15) 5mCG. We
also constructed a fusion enzyme 2xMBD2-ND(N.BceSVIII) targeting more frequent
composite sites AS upward arrowYS N(5-12) 5mCG in Mn2+ buffer. 5mCG-dependent
nicking requires special digestion conditions in high salt (0.3 M KCl) or in Ni2+
buffer. The fusion enzyme can be used to nick and label 5mCG-modified plasmid and
genomic DNAs with fluorescently labeled Cy3-dUTP and potentially be useful for
diagnostic applications, DNA sequencing and optical mapping of epigenetic
markers. The importance of the predicted catalytic residues D89, H90, N106 and
H115 in N.Gamma was confirmed by mutagenesis. We found that the wild-type enzyme
N.Gamma prefers to nick 5mCG-modified DNA in Ni2+ buffer even though the nicking
activity is sub-optimal compared to the activity in Mg2+ buffer.

<>

<1>Guyot, J.-B., Caudron, B.
<2>Statistical significance of amino acid sequence similarity in type II DNA methyltransferases.
<3>C.R. Acad. Sci. III
<4>317
<5>20-24
<6>1994
<7>The statistical significance of amino acid sequence similarities previously observed in type
II DNA methyltransferases has been investigated. It is shown: (1) that the intramolecular
similarities observed among various type II Mtases are not statistically significant and thus
cannot be used to support a gene duplication model; (2) that the intermolecular similarities
observed in a peptide in various type II adenine methylases are statistically confirmed; (3)
that the similarities observed between MutH and these proteins for this peptide are not
statistically significant and therefore cannot be used to propose a functional role in DNA
recognition for this peptide.

<>

<1>Guyot, J.B., Grassi, J., Hahn, U., Guschlbauer, W.
<2>The role of the preserved sequences of Dam methylase.
<3>Nucleic Acids Res.
<4>21
<5>3183-3190
<6>1993
<7>We have undertaken a site directed mutational analysis of two of the preserved regions in the
amino acids sequence of Dam methylase in order to characterize their role. Mutations in region
IV (sequence DPPY) abolish catalytic activity and greatly affect AdoMet crosslinking. Mutants
in region III display a lowered specific activity with an unchanged AdoMet crosslinking
capacity. We have also made a series of deletions both at the N and C terminal parts of the
protein, which have been found to provide inactive enzyme. We discuss the significance of
these results for the understanding of the functional properties of the enzyme.

<>

<1>Gwinn, D.D., Lawton, W.D.
<2>Alteration of host specificity in Bacillus subtilis.
<3>Bacteriol. Rev.
<4>32
<5>297-301
<6>1968
<7>Restriction of bacteriophages in host bacteria has been reported by many investigators, most
of whom have observed the phenomenon to be associated with host-induced modification.  This
report describes the restriction of bacteriophage without any apparent host-induced
modification.  After a period of incubation at 53C, Bacillus subtilis 168 (ind-) cells were
rendered permissive in their response to infection by phages SP-10 and SP-20.  Because phage
SP-20 infected heated 168 cells with an efficiency 100 times greater than phage SP-10, most of
the work described here was done with phage SP-20.

<>

<1>Gyllborg, M.C., Sahl, J.W., Cronin, D.C.I.I.I., Rasko, D.A., Mandel, M.J.
<2>Draft Genome Sequence of Vibrio fischeri SR5, a Strain Isolated from the Light Organ of the Mediterranean Squid Sepiola robusta.
<3>J. Bacteriol.
<4>194
<5>1639
<6>2012
<7>Here, we describe the draft genome sequence of Vibrio fischeri SR5, a squid symbiotic isolate
from Sepiola robusta in the Mediterranean Sea. This 4.3-Mbp
genome sequence represents the first V. fischeri genome from an S. robusta
symbiont and the first from outside the Pacific Ocean.

<>

<1>Ha, J.-H., Spolar, R.S., Record, M.T.
<2>Role of the hydrophobic effect in stability of site-specific protein-DNA complexes.
<3>J. Mol. Biol.
<4>209
<5>801-816
<6>1989
<7>The site-specific binding interaction of lac repressor with a symmetric
operator sequence and of EcoRI endonuclease with its specific recognition site
both exhibit a characteristic dependence of equilibrium binding constant (Kobs)
on temperature, in which Kobs attains a relative maximum in the physiologically
relevant temperature range.  This behavior, which appears to be quite general
for site-specific protein-DNA interactions, is indicative of a large negative
standard heat capacity change (DeltaCoP,obs) in the association process.  By
analogy with model compound transfer studies and protein folding data, we
propose that this DeltaCoP,obs results primarily from the removal of non-polar
surface from water in the association process.  From DeltaCoP,obs we obtain
semiquantitative information regarding the change in water-exposed non-polar
surface area (DeltaAnp) and the corresponding hydrophobic driving force for
association (DeltaGohyd):Delta Go hyd~8(+/- 1) x 10/1 DeltaCoP,obs~ -22 (+/-5)
DeltaAnp.  We propose that removal of non-polar surface from water (the
hydrophobic effect) and release of cations (the polyelectrolyte effect) drive
the thermodynamically unfavorable processes (e.g. conformational distortions)
necessary to achieve mutually complementary recognition surfaces (at a steric
and functional-group level) in the specific complex.

<>

<1>Haake, S.K., Yoder, S.C., Attarian, G., Podkaminer, K.
<2>Native plasmids of Fusobacterium nucleatum: Characterization and use in development of genetic systems.
<3>J. Bacteriol.
<4>182
<5>1176-1180
<6>2000
<7>Three native plasmids of Fusobacterium nucleatum were characterized, including DNA sequence
analysis of one plasmid, pFN1. A shuttle plasmid, pHS17, capable of transforming Escherichia
coli and F. nucleatum ATCC 10953 was constructed with pFN1. pHS17 was stably maintained in the
F. nucleatum transformants, and differences in the transformation efficiencies suggested the
presence of a restriction-modification system in F. nucleatum.

<>

<1>Habe, H., Kobuna, A., Hosoda, A., Kouzuma, A., Yamane, H., Nojiri, H., Omori, T., Watanabe, K.
<2>Subtractive hybridization and random arbitrarily primed PCR analyses of a benzoate-assimilating bacterium, Desulfotignum balticum.
<3>Appl. Microbiol. Biotechnol.
<4>79
<5>87-95
<6>2008
<7>Subtractive hybridization (SH) and random arbitrarily primed PCR (RAP-PCR) were used to detect
genes involved in anaerobic benzoate degradation by
Desulfotignum balticum. Through SH, we obtained 121 DNA sequences specific
for D. balticum but not for D. phosphitoxidans (a
non-benzoate-assimilating species). Furthermore, RAP-PCR analysis showed
that a 651-bp DNA fragment, having 55% homology with the solute-binding
protein of the ABC transporter system in Methanosarcina barkeri, was
expressed when D. balticum was grown on benzoate, but not on pyruvate. By
shotgun sequencing of the fosmid clone (38,071 bp) containing the DNA
fragment, 33 open reading frames (ORFs) and two incomplete ORFs were
annotated, and several genes within this region corresponded to the DNA
fragments obtained by SH. An 11.3-kb gene cluster (ORF10-17) revealed
through reverse transcription-PCR showed homology with the ABC transporter
system and TonB-dependent receptors, both of which are presumably involved
in the uptake of siderophore/heme/vitamin B(12), and was expressed in
response to growth on benzoate.

<>

<1>Haber, J.E., Wolfe, K.H.
<2>Function and evolution of HO and VDE endonucleases in fungi.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Berlin Heidelberg
<4>16
<5>161-175
<6>2005
<7>The site-specific HO and VDE endonucleases are unusual members of a family of so-called group
I LAGLIDADG homing endonucleases that are generally implicated in the homing of intron and
intein sequences.  The great majority of these endonucleases are found in mitochondria and
plastids of eukaryotes, but in budding yeast two members of this family apparently "escaped"
into the nucleus.  In each case, the endonuclease creates a site-specific double-strand break
in a target and promotes mobility of DNA sequences by homologous recombination requiring the
Rad52 and Rad51 group of recombination proteins.  The HO endonuclease has been the subject of
a great deal of interest, both because of its remarkable evolution and because of its great
utility in the detailed analysis of DSB-mediated recombination.  Unlike most homing
endonucleases, the HO endonuclease does not promote its own amplification, but rather
catalyzes the switching/replacement of yeast's mating-type genes.  In addition, budding
yeasts harbor the VDE endonuclease, which is found as an intein in the VMA1 gene.  VDE
promotes its propagation from a VMA1 gene containing the VDE intein into a VMA1 gene lacking
such a sequence.  VDE is remarkable also in sharing a close sequence and presumably
evolutionary relationship with HO.

<>

<1>Haber, K., Neumark, T., Foldes, I.
<2>Restriction-modification like phenomenon observed in Mycobacterium smegmatis strains are consequences of natural polylysogeny.
<3>Acta Microbiol. Hung.
<4>35
<5>180-181
<6>1988
<7>Titrating phage butyricum (By) on its original host, Mycobacterium smegmatis
butyricum, and on M. smegmatis SN2 (SN2), phenomena similar to
restriction-modification could be observed.  On the contrary, observations
showing that By phages propagated on SN2 cells have altered morphological
structure - instead of the icosahedral shape of the head of phage By, the head
of phage By propagated on SN2 cells is elongated - could not be explained by
the effect of DNA modification.  It has been assumed that the phenomena
observed resulted as a consequence of natural lysogeny of the M. smegmatis
strains.  This hypothesis was proved by biological methods and electron
microscopic observations.  Phages with non-contractile tails of both types of
head morphology were found to be present in lysates of M. smegmatis butyricum
and SN2 cells, showing the polylysogenic property of their strains.

<>

<1>Haberhauer, G., Zelder, O., Pompejus, M., Schroeder, H., Kroeger, B.
<2>Corynebacterium glutamicum genes encoding proteins involved in homeostasis and adaptation.
<3>Japanese Patent Office
<4>JP 2007259859 A
<5>
<6>2007
<7>
<>

<1>Haberman, A.
<2>The bacteriophage P1 restriction endonuclease.
<3>J. Mol. Biol.
<4>89
<5>545-563
<6>1974
<7>The bacteriophage P1 restriction endonuclease has been purified from
Escherichia coli lysogenic for P1.  This restriction endonuclease P has a
sedimentation coefficient of 9.3S.  Unlike the E. coli K restriction
endonuclease, endonuclease P does not require S-adenosylmethionine for breakage
of DNA.  S-adenosylmethionine does, however, stimulate the rate of
double-strand breakage of DNA by endonuclease P.  Hydrolysis of ATP by
endonuclease P could not be detected under conditions in which the K
restriction endonuclease massively degrades ATP.  The enzyme makes a limited
number of double-strand breaks in unmodified or heterologously modified lambda
DNA.  In the presence of S-adenosylmethionine, it does not cut every DNA
molecule to the same extent.  Incubation of lambda DNA with excess amounts of
the enzyme results in less breakage of the DNA than with smaller amounts of
enzyme.  This effect is not seen in the absence of S-adenosylmethionine appears
to be greater than the maximum amount of cutting in its presence.  This is most
likely due to the modification methylase activity of P1 restriction
endonuclease.

<>

<1>Haberman, A., Heywood, J., Meselson, M.
<2>DNA modification methylase activity of Escherichia coli restriction endonucleases K and P.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>3138-3141
<6>1972
<7>The highly purified restriction endonucleases of E. coli K and coliphage P1
transfer methyl groups from S-adenosylmethionine to adenine residues of
unmodified DNA.  Incubation of unmodified DNA with endonucleases K or P and
S-adenosylmethionine renders the DNA resistant to restriction.  The enzymes,
therefore, have both restriction endonuclease and modification methylase
activities.

<>

<1>Hadi, S.M.
<2>A site-specific DNA methylase from Escherichia coli K.
<3>Indian J. Biochem. Biophys.
<4>18
<5>295-297
<6>1981
<7>A DNA methylase has been partially purified from E. coli K using heparin-agarose and
DEAE-sephacel chromatography.  Plasmid pBR322 DNA was methylated by the enzyme and cleaved
with restriction endonuclease HaeIII.  Among the DNA fragments obtained, only a few were
methylated demonstrating that the enzyme recognizes specific sites on DNA.  Since all the
EcoRII cleavage sites fall within the methylated fragments, the DNA methylase may be the
cytosine methylase of E. coli K which is known to render the methylated sites resistant to
EcoRII cleavage.

<>

<1>Hadi, S.M., Bachi, B., Iida, S., Bickle, T.A.
<2>DNA restriction-modification enzymes of phage P1 and plasmid p15B.
<3>J. Mol. Biol.
<4>165
<5>19-34
<6>1983
<7>We have purified the type III restriction enzymes EcoPI and EcoP15 to
homogeneity from bacteria that contain the structural genes for the enzymes
cloned on small, multicopy plasmids and which overproduce the enzymes.  Both of
the enzymes contain two different subunits.  The molecular weights of the
subunits are the same for both enzymes and antibodies prepared against one
enzyme cross-react with both subunits of the other.  Bacteria containing a
plasmid derivative in which a large part of one of the structural genes has
been deleted have a restriction- modification+ phenotype and contain only the
smaller of the two subunits.  This subunit therefore must be the one that both
recognizes the specific DNA sequence and methylates it in the modification
reaction (the restriction enzyme itself also acts as a modification methylase).
We have purified the PI and P15 modification subunits from these deletion
derivatives and have shown that in vitro they have the expected properties:
they are sequence-specific modification methylases.  In addition, we have
demonstrated that strains carrying the full restriction/modification system
also contain a pool of free modification subunits that might be responsible for
in vivo modification.

<>

<1>Hadi, S.M., Bachi, B., Shepherd, J.C.W., Yuan, R., Ineichen, K., Bickle, T.A.
<2>DNA Recognition and Cleavage by the EcoP15 Restriction Endonuclease.
<3>J. Mol. Biol.
<4>134
<5>655-666
<6>1979
<7>EcoP15 is a restriction-modification enzyme coded by the P15 plasmid of
Escherichia coli.  We have determined the sites recognized by this enzyme on
pBR322 and simian virus 40 DNA.  The enzyme recognizes the sequence: 5'
C-A-G-C-A-G 3'. . . . . . 3' G-T-C-G-T-C 5' for restriction, the enzyme cleaves
the DNA 25 to 26 base-pairs 3' to this sequence and cleaves single-stranded 5'
protrusions two bases long.

<>

<1>Hadi, S.M., Bickle, T.A., Yuan, R.
<2>The role of S-adenosylmethionine in the cleavage of deoxyribonucleic acid by the restriction endonuclease from Escherichia coli K.
<3>J. Biol. Chem.
<4>250
<5>4159-4164
<6>1975
<7>The restriction endonuclease from Escherichia coli K specifically cleaves
foreign DNA in the presence of S-adenosylmethionine, ATP, and Mg++.  The role
of S-adenosylmethionine in this reaction has been studied by following the
specific binding of the enzyme to unmodified DNA.  The results indicate that
S-adenosylmethionine acts as an allosteric effector.  However, the
rate-limiting step in the activation of the enzyme is not the binding of the
effector itself, but an event subsequent to it.  The interaction of the
S-adenosylmethionine with two mutant K restriction endonucleases isolated
previously has also been investigated.  One of them, which is defective in
restriction, can be activated in a manner similar to the wild type enzyme,
while the other one, which lacks both restriction and modification activities
(due to a mutation in the subunit responsible for DNA recognition), shows no
such effect.

<>

<1>Hadi, S.M., Yuan, R.
<2>Complementation in vitro by restriction enzymes from mutant strains E. coli K12.
<3>Experientia
<4>29
<5>752
<6>1973
<7>The restriction endonuclease from E. coli K12 has previously been shownw to a)
bind and then cleave unmodified lambda DNA in the presence of SAM, ATP and
Mg++, b) hydrolyze the ATP to ADP and inorganic phosphate in the course of the
restriction reaction, and c) methylate unmodified lambda DNA when only SAM is
present.  The restriction deficient mutant strains from E. coli K12, gamma
K-mK+ and gamma K-mK- have been shown to complement in vivo to yield a wild
type phenotype, gamma K+mK+.  Using a binding assay based on complementation in
vitro between the two mutant extracts, the restriction endonucleases from the
two mutant strains were purified extensively.  Complementation in vitro could
be detected by specific binding and also by cleavage of the unmodified DNA when
both mutant proteins were present.  Either mutant protein by itself showed no
activity.  Studies on the methylase and ATPase activities of these mutant
proteins, either alone or in combination are currently in progress.

<>

<1>Hadi, S.M., Yuan, R.
<2>Complementation in vitro by mutant restriction enzymes from Escherichia coli K.
<3>J. Biol. Chem.
<4>249
<5>4580-4586
<6>1974
<7>Two mutant strains of Escherichia coli K, K-18 lacking restriction activity
(endonucleolytic cleavage of foreign DNA), and the other, K-19 lacking both
restriction and modification activities (specific DNA methylation that protects
against the homologous restriction activity), complement in vivo to yield a
wild type phenotype.  Although neither mutant extract alone binds unmodified
DNA, the wild type extract and the mixture of mutant extracts do.  Retention of
the DNA-enzyme complex on membrane filters was used as an assay to purify the
two mutant restriction enzymes.  Both of these sediment like the wild type
enzyme.  Complementation in vitro by the two mutant enzymes could be
demonstrated by specific DNA binding, cleavage of the unmodified DNA, and
restriction-dependent ATP hydrolysis.  All of these activities are absent in
the individual mutant endonucleases but present in the wild type restriction
endonuclease from E. coli K.  However, both mutant enzymes show an activity
that hydrolyzes ATP which is different from that of the wild type enzyme since
it does not require unmodified DNA, but is dependent on the presence of
S-adenosylmethionine.

<>

<1>Haendiges, J., Timme, R., Allard, M., Myers, R.A., Payne, J., Brown, E.W., Evans, P., Gonzalez-Escalona, N.
<2>Draft Genome Sequences of Clinical Vibrio parahaemolyticus Strains Isolated in Maryland (2010 to 2013).
<3>Genome Announcements
<4>2
<5>e00776-14
<6>2014
<7>Vibrio parahaemolyticus is the leading cause of food-borne illnesses associated with the
consumption of raw shellfish worldwide. Here, we report 45 draft genomes
of V. parahaemolyticus. Thirty-five of them are strains that were isolated from
clinical cases in the state of Maryland from 2010 to 2013. The remaining 10
strains were historical isolates, isolated mostly from the West Coast of the
United States during the period of 1988 to 2004. The availability of these
genomes will allow for future phylogenetic analyses with other V.
parahaemolyticus strains.

<>

<1>Hafez, M., Guha, T.K., Hausner, G.
<2>I-OmiI and I-OmiII: Two intron-encoded homing endonucleases within the Ophiostoma minus rns gene.
<3>Fungal Biol.
<4>118
<5>721-731
<6>2014
<7>The mitochondrial small subunit ribosomal RNA (rns) gene of the ascomycetous fungus Ophiostoma
minus [strain WIN(M)371] was found to contain a group IC2 and a group IIB1 intron at positions
mS569 and mS952 respectively. Both introns have open reading frames (ORFs) embedded that
encode double motif LAGLIDADG homing endonucleases (I-OmiI and I-OmiII respectively).
Codon-optimized versions of I-OmiI and I-OmiII were synthesized for overexpression in
Escherichia coli. The in vitro characterization of I-OmiII showed that it is a functional
homing endonuclease that cleaves the rns target site two nucleotides upstream (sense strand)
of the intron insertion site generating 4 nucleotide 3' overhangs. The endonuclease activity
of I-OmiII was tested using linear and circular substrates and cleavage activity was evaluated
at various temperatures. The I-OmiI protein was expressed in E. coli, but purification was
difficult, thus the endonuclease activity of this protein was tested via in vivo assays.
Overall this study showed that there are many native forms of functional homing endonucleases
yet to be discovered among fungal mtDNA genomes.

<>

<1>Hafstrom, T., Jansson, D.S., Segerman, B.
<2>Complete Genome Sequence of Brachyspira intermedia Reveals Unique Genomic Features in Brachyspira Species and Phage-mediated Horizontal Gene Transfer.
<3>BMC Genomics
<4>12
<5>395
<6>2011
<7>ABSTRACT: BACKGROUND: Brachyspira spp. colonize the intestines of some
mammalian and avian species and show different degrees of
enteropathogenicity. Brachyspira intermedia can cause production losses in
chickens and strain PWS/AT now becomes the fourth genome to be completed
in the genus Brachyspira. RESULTS: 15 classes of unique and shared genes
were analyzed in B. intermedia, B. murdochii, B. hyodysenteriae and B.
pilosicoli. The largest number of unique genes was found in B. intermedia
and B. murdochii. This indicates the presence of larger pan-genomes. In
general, hypothetical protein annotations are overrepresented among the
unique genes. A 3.2 kb plasmid was found in B. intermedia strain PWS/AT.
The plasmid was also present in the B. murdochii strain but not in nine
other Brachyspira isolates. Within the Brachyspira genomes, genes had been
translocated and also frequently switched between leading and lagging
strands, a process that can be followed by different AT-skews in the third
positions of synonymous codons. We also found evidence that bacteriophages
were being remodeled and genes incorporated into them. CONCLUSIONS: The
accessory gene pool shapes species-specific traits. It is also influenced
by reductive genome evolution and horizontal gene transfer. Gene-transfer
events can cross both species and genus boundaries and bacteriophages
appear to play an important role in this process. A mechanism for
horizontal gene transfer appears to be gene translocations leading to
remodeling of bacteriophages in combination with broad tropism.

<>

<1>Hager, P.W., Boyer, H.W., Day, J.O., Reich, N.O., Greene, P.
<2>Analysis of noncanonical DNA cleavage by EcoRI endonuclease mutants.
<3>J. Cell Biochem. Suppl.
<4>11C
<5>206
<6>1987
<7>It has been proposed that DNA sequence specificity for the EcoRI endonuclease
results from the formation of twelve hydrogen bonds between bases exposed at
the major groove of the EcoRI site and six amino acid side chains (3 from each
subunit) in the enzyme.  (Rosenberg and Greene, DNA 1, 117-124 (1982),
Rosenberg et al., Science, in press) Conservative replacements in the 3 amino
acids identified in the x-ray crystallographic structure as responsible for
specificity were made (Glu 144 to Asp, Arg 145 to Lys, and Arg 200 to Lys).
These changes failed to alter the site of primary cleavage.  However, all of
these proteins nick noncanonical DNA sequences in pBR322 lacking the EcoRI
site.  The location of these single strand cleavage sites is being determined
for the wild type and mutant enzymes.  The identification of the sequences
recognized by the different enzymes and the hierarchy of the rates at which
these sequences are hydrolyzed will contribute to our understanding of how the
endonuclease discriminates between DNA squences.

<>

<1>Hager, P.W., Reich, N.O., Day, J.P., Coche, T.G., Boyer, H.W., Rosenberg, J.M., Greene, P.J.
<2>Probing the role of glutamic acid 144 in the EcoRI endonuclease using aspartic acid and glutamine replacements.
<3>J. Biol. Chem.
<4>265
<5>21520-21526
<6>1990
<7>The x-ray structure of the EcoRI endonuclease-DNA complex suggests that
hydrogen bonds between amino acids, glutamic acid 144, arginine 145, and
arginine 200, and major groove base moieties are the molecular determinants of
specificity.  We have investigated residue 144 using aspartate and glutamine
substitutions introduced by site-directed mutagenesis.  Substitution with
glutamine results in a null phenotype (at least a 2000-fold reduction in
activity).  On the other hand, the aspartic acid mutant (ED144) retained in
vivo activity.  Substrate binding and catalytic studies were done with purified
ED144 enzyme.  The affinity of the ED144 enzyme for the canonical sequence
5'-GAATTC-3' is about 340-fold less than the wild-type (WT) enzyme, while its
affinity for nonspecific DNA is about 50 times greater.  The ED144 enzyme
cleaves one strand in the EcoRI site in plasmid pBR322 with a kcat/Km similar
to WT.  In contrast to the WT enzyme, the ED144 enzyme dissociates after the
first strand cleavage.  Partitioning between cleavage and dissociation at the
first and second cleavage steps for the ED144 enzyme is extremely
salt-sensitive.  The altered partitioning results largely from a
destabilization of the enzyme-DNA complex, particularly the enzyme-nicked DNA
complex, with only small changes in the respective cleavage rates.  The
hydrogen bonds of Glu-144 are critical, they appear to act cooperatively with
other specificity contacts to stabilize the enzyme-DNA complex.

<>

<1>Hahn, D.R., McHenney, M.A., Baltz, R.H.
<2>Characterization of FP22, a large streptomycete bacteriophage with DNA insensitive to cleavage by many restriction enzymes.
<3>J. Gen. Microbiol.
<4>136
<5>2395-2404
<6>1990
<7>Bacteriophage FP22 has a very broad host range within streptomycetes and
appeared to form lysogens of Streptomyces ambofaciens ATCC 15154.  FP22 shared
strong cross-immunity and antibody cross-reactivity with bacteriophage P23, but
not with seven other streptomycete bacteriophages.  FP22 particles had a head
diameter of 71 nm and a tail length of 307 mm.  The FP22 genome was 131 kb,
which is the largest bacteriophage genome reported for streptomycetes.  The G+C
content of the genome was 46 mol% and restriction mapping indicated that FP22
DNA had discrete ends.  NaCl- and pyrophosphate-resistant deletion mutants were
readily isolated and the extent of the deletions defined at least 23 kb of
dispensable DNA in two regions of the genome.  The DNA was not cleaved by most
restriction endonucleases (or isoschizomers) which have been identified in the
streptomycetes, including the tetranucleotide cutter MboI (GATC).

<>

<1>Haima, P., Bron, S., Venema, G.
<2>The effect of restriction on shotgun cloning and plasmid stability in Bacillus subtilis Marburg.
<3>Mol. Gen. Genet.
<4>209
<5>335-342
<6>1987
<7>Using the bifunctional cloning vehicle pHP13, which carries the replication
functions of the cryptic Bacillus subtilis plasmid pTA1060, the effects of BsuM
restriction on the efficiency of shotgun cloning of heterologous Escherichia
coli DNA were studied.  In a restriction-deficient but modification-proficient
mutant of B. subtilis, clones were obtained at a high frequency, comparable to
frequencies normally obtained in E. coli (10/4 clones per microgram target
DNA).  Large inserts were relatively abundant (26% of the clones contained
inserts in the range of 6 to 15 kb), which resulted in a high average insert
length (3.6 kb).  In the restriction-proficient B. subtilis strain, the class
of large inserts was underrepresented.  Transformation of B. subtilis with E.
coli-derived individual recombinant plasmids was affected by BsuM restriction
in two ways.  First, the transforming activities of recombinant plasmids
carrying inserts larger than 4kb, were, in comparison with the vector pHP13,
reduced to varying degrees in the restricting host.  The levels of the
reduction increased with insert length, resulting in a 7800-fold reduction for
the largest plasmid used (pC23; insert length 16 kb).  Second, more than 80% of
the pC23 transformants in the restricting strain contained a deleted plasmid.
In the non-restricting strain, the transforming activities of the plasmids were
fairly constant as a function of insert length (in the range of 0-16 kb), and
no structural instability was observed.  It is concluded that for shotgun
cloning in B. subtillis, the use of restriction-deficient strains is highly
preferable.  Evidence is presented that in addition to XhoI other sequences are
involved in BsuM restriction.  It is postulated that AsuII sites are additional
target sites for BsuM restriction.

<>

<1>Hain, T. et al.
<2>Whole-Genome Sequence of Listeria welshimeri Reveals Common Steps in Genome Reduction with Listeria innocua as Compared to Listeria  monocytogenes.
<3>J. Bacteriol.
<4>188
<5>7405-7415
<6>2006
<7>We present the complete genome sequence of Listeria welshimeri, a nonpathogenic member of the
genus Listeria. Listeria welshimeri harbors a
circular chromosome of 2,814,130 bp with 2,780 open reading frames.
Comparative genomic analysis of chromosomal regions between L. welshimeri,
Listeria innocua, and Listeria monocytogenes shows strong overall
conservation of synteny, with the exception of the translocation of an
F(o)F(1) ATP synthase. The smaller size of the L. welshimeri genome is the
result of deletions in all of the genes involved in virulence and of
"fitness" genes required for intracellular survival, transcription
factors, and LPXTG- and LRR-containing proteins as well as 55 genes
involved in carbohydrate transport and metabolism. In total, 482 genes are
absent from L. welshimeri relative to L. monocytogenes. Of these, 249
deletions are commonly absent in both L. welshimeri and L. innocua,
suggesting similar genome evolutionary paths from an ancestor. We also
identified 311 genes specific to L. welshimeri that are absent in the
other two species, indicating gene expansion in L. welshimeri, including
horizontal gene transfer. The species L. welshimeri appears to have been
derived from early evolutionary events and an ancestor more compact than
L. monocytogenes that led to the emergence of nonpathogenic Listeria spp.

<>

<1>Halazonetis, T.D., Kandil, A.N.
<2>Predicted structural similarities of the DNA binding domains of c-Myc and endonuclease EcoRI.
<3>Science
<4>255
<5>464-466
<6>1992
<7>The c-Myc oncoprotein belongs to a family of proteins whose DNA binding domains
contain a basic region-helix-loop-helix (bHLH) motif.  Systematic mutagenesis
of c-Myc revealed that dimerized bHLH motifs formed a parallel four-helix
bundle with the amino termini of helices 1 and 2 directed toward the inner and
outer nucleotides of the DNA binding site, respectively.  Both the basic region
and the carboxyl-terminal end of the loop contributed to DNA binding
specificity.  The DNA binding domain of c-Myc may therefore be structurally
similar to that of restriction endonuclease EcoRI.

<>

<1>Halden, N.F., Wolf, J.B., Cross, S.L., Leonard, W.J.
<2>Identification and characterization of a novel restriction enzyme derived from Mycoplasma fermentans.
<3>Clin. Res.
<4>36
<5>404a
<6>1988
<7>During our studies defining proteins that bind to the 5' regulatory regions of
the human interleukin-2 receptor alpha chain (IL2Ralpha,p55, Tac antigen) gene,
we have unexpectedly identified a new restriction enzyme specific for a
sequence not cleaved by any known restriction enzyme.  Nuclear extracts from
various cell lines were incubated with 32P-labeled DNA from the IL2Ralpha
promoter region.  In the absence of any exogenous nuclease, extracts from
Jurkat and MT-2 cells, but not from HeLa and HUT-102B2 cells, were capable of
cleaving the DNA.  Subsequent analysis confirmed that only the Jurkat and MT-2
T cells were infected with mycoplasma.  When extracts were prepared from the
same Jurkat cell line cured with BM cycline, the activity was not present.  We
have therefore concluded that the nuclease activity was derived from the
mycoplasma rather than the T cells.  The mycoplasma strain containing both MT-2
and Jurkat cells was identified as M. fermentans.  The new enzyme specifically
recognizes the palindrome CAATTG, is inactivated by 15 min treatment at 65C, by
proteinase K, and by 20 mM EDTA, and works optimally in low salt (<10mM NaCl)
restriction enzyme buffer.  In conclusion, this enzyme recognizes a new
recognition sequence and therefore has potential for general use in molecular
biology.  In addition, it is possible that a rapid diagnostic test for
mycoplasma may result from the use of mycoplasma specific restriction
endonuclease assays on appropriate sequences of DNA.

<>

<1>Halden, N.F., Wolf, J.B., Leonard, W.J.
<2>Identification of a novel site specific endonuclease produced by Mycoplasma fermentans:  discovery while characterizing DNA binding proteins in T lymphocyte cell lines.
<3>Nucleic Acids Res.
<4>17
<5>3491-3499
<6>1989
<7>We have discovered a new restriction endonuclease, MfeI, in nuclear extracts
from T cells contaminated with Mycoplasma fermentans.  This endonuclease was
identified while studying proteins binding to the interleukin-2 receptor alpha
chain gene promoter.  MfeI cuts at the recognition sequence C^AATTG generating
EcoRI compatible cohesive ends.  Potential applications are discussed.

<>

<1>Hale, W.B., van der Woude, M.W., Low, D.A.
<2>Analysis of nonmethylated GATC sites in the Escherichia coli chromosome and identification of sites that are differentially methylated in response to environmental stimuli.
<3>J. Bacteriol.
<4>176
<5>3438-3441
<6>1994
<7>Seven GATC sites that are nonmethylated in logarithmic growth phase cells using glycerol as a
carbon source were isolated from the Escherichia coli chromosome.  Three of these GATC sites
are located upstream of the operons gut, mtl, and ppiA, whereas DNA sequences adjacent to
three other nonmethylated GATC sites are not homologous to previously identified genes.  The
seventh nonmethylated GATC site is located downstream of uspA.  The protection of this site
from DNA methylation requires leucine-responsive regulatory protein and is leucine responsive.
The carbon source and the growth phase influenced the protection of the GATC site 5' of the
ppiA gene.  The other five sites were protected under all the environmental conditions
examined.

<>

<1>Haley, B.J., Choi, S.Y., Hasan, N.A., Abdullah, A.S., Cebula, T.A., Huq, A., Colwell, R.R.
<2>Genome Sequences of Clinical Vibrio cholerae Isolates from an Oyster-Borne Cholera Outbreak in Florida.
<3>Genome Announcements
<4>1
<5>e00966-13
<6>2013
<7>Between November 2010 and April 2011, 11 cases of cholera were identified and associated with
the consumption of raw oysters harvested from Apalachicola Bay,
Florida. The etiological agent was the ctxAB-positive Vibrio cholerae serogroup
O75. The genome sequences of the isolates provide useful information and are
deposited in the public genome databases.

<>

<1>Haley, B.J., Luo, Y., Wang, C., Pettengill, J., Allard, M., Brown, E., Karns, J.S., Van Kessel, J.A.
<2>Genome Sequences of Eight Salmonella enterica subsp. enterica Serovars Isolated from a Single Dairy Farm.
<3>Genome Announcements
<4>2
<5>e00082-14
<6>2014
<7>Here, we report draft genome sequences of 26 isolates of Salmonella enterica subsp. enterica,
representing eight serotypes, which were isolated from cows in a Pennsylvania dairy herd, the
farm on which they were reared, and the associated off-site heifer-raising facility over an
8-year sampling period.

<>

<1>Halford, S.
<2>Restriction enzymes in DNA-protein interactions.
<3>SERC Bulletin
<4>3
<5>12-13
<6>1986
<7>Within any organism from Escherichia coli to man, the genetic information is stored as a
sequence of bases in the DNA but this constitutes only a blue-print for that organism.  The
retrieval of the genetic information is absolutely dependent upon proteins that interact with
the DNA.  The molecular basis of DNA-protein interactions is being studied by a group at the
Bristol University who have used the EcoRI restriction endonuclease, an enzyme widely used in
genetic engineering, as the test system.

<>

<1>Halford, S.E.
<2>The specificity of the EcoRI restriction endonuclease.
<3>Biochem. Soc. Trans.
<4>8
<5>399-400
<6>1980
<7>None

<>

<1>Halford, S.E.
<2>Restriction enzymes.
<3>Encyclopaedia of Molecular Biology, Blackwell, Kendrew, J., Oxford
<4>
<5>954-956
<6>1994
<7>Restriction enzymes are endonucleases that cleave DNA in response to a recognition site on the
DNA.  The recognition site consists of a specific sequence of nucleotides in the DNA duplex,
typically 4-8 base pairs long.  These endonucleases are found in many species of bacteria,
where they function in restriction and modification systems.  The bacterial R/M system is
analogous to an immune system, in that it enables the bacterium to distinguish its own DNA
from foreign DNA and to eliminate the latter.  The discovery of restriction-modification
systems followed the observation more than 30 years ago that a single cycle of phage growth in
a particular bacterial host could alter the host range of the progeny phage; they had become
'restricted' in their host range as a result of 'modification' in the original host.
Restriction-modification is achieved by a combination of two enzyme activities: a modification
methyltransferase and a restriction endonuclease.  The former transfers methyl groups from
S-adenosylmethionine to specific bases within the recognition sequence, generally one in each
strand: if one strand is already methylated, the second strand remains a substrate.  The
latter cleaves the DNA but only if the recognition site is not methylated in either strand.
Methylation of just one strand blocks restriction activity, so the bacterial DNA is protected
from the endonuclease even after its semiconservative replication.  However, DNA that is
foreign to the cell carrying the R/M system will lack the appropriate methylation and, if such
DNA enters the cell, it is likely to be cleaved by the restriction enzyme.

<>

<1>Halford, S.E.
<2>Restriction enzymes that act simultaneously at two DNA sites.
<3>Biochem. Soc. Trans.
<4>27
<5>A88
<6>1999
<7>The reaction of a type II restriction enzyme commonly results in DNA cleavage at a single
site.  DNA with multiple recognition sites is cleaved by a succession of separate reactions at
individual sites.  However, it has been discovered recently that many of the type II
endonucleases can convert a substrate with two recognition sites directly into the product
cleaved at both sites, without liberating DNA cut at a single site.  The coupled cleavage of
two sites can be due to a processive mechanism involving the intramolecular transfer of the
enzyme from one site to another.  Whilst such transfers have generally been considered to
occur by "sliding", the linear diffusion of the protein along the DNA, studies on the EcoRV
endonuclease have excluded "sliding" as a major pathway for intramolecular transfer and have
indicated instead that the transfer occurs by "hopping", the dissociation of the protein from
one site on the DNA followed by its reassociation to another site on the same molecule of DNA.
In other cases, concerted action at two DNA sites is achieved by the restriction enzyme being
unable to cleave DNA until it has bound to two copies of its recognition sequence, either by
looping out the DNA between sites on the same DNA molecule or by bridging sites on separate
molecules.  The SfiI endonuclease provides an illustration of the latter system.

<>

<1>Halford, S.E.
<2>Hopping, jumping and looping by restriction enzymes.
<3>Biochem. Soc. Trans.
<4>29
<5>363-373
<6>2001
<7>Type II restriction endonucleases recognize specific DNA sequences and cleave both strands of
the DNA at fixed locations at or near their
recognition sites. Many of these enzymes are dimeric proteins that
recognize, in symmetrical fashion, palindromic DNA sequences. They
generally catalyse independent reactions at each recognition site on
the DNA, although in some cases they act processively; cutting the DNA
first at one site, then translocating along the DNA to another site and
cutting that before leaving the DNA. The way in which the degree of
processivity varies with the length of DNA between the sites can reveal
the mechanism of translocation. In contrast with the common view that
proteins move along DNA by 'sliding', the principal mode of transfer of
the EcoRV endonuclease is by 'hopping' and 'jumping', i.e. the
dissociation of the protein from one site followed by its
re-association with another site in the same DNA molecule, either close
to or distant from the original site. Other type II restriction enzymes
require two copies of their recognition sites for their DNA cleavage
reactions. Many of these enzymes, such as SfiI, are tetramers with two
DNA-binding surfaces. SfiI has no activity when bound to just one
recognition site, and instead both DNA-binding surfaces have to be
filled before it becomes active. Although the two sites can be on
separate DNA molecules, SfiI acts optimally with two sites on the same
DNA, where it traps the DNA between the sites in a loop. SfiI thus
constitutes a test system for the analysis of DNA looping.

<>

<1>Halford, S.E.
<2>How does EcoRI cleave its recognition site on DNA?
<3>Trends Biochem. Sci.
<4>8
<5>455-460
<6>1983
<7>The two protein subunits of the EcoRI restriction enzyme interact symmetrically
with the recognition site on DNA, so that each subunit is in position to cleave
one strand of the DNA.  But each subunit seems to require a protein
conformation change before it can cleave DNA.  Depending upon whether one or
both subunits change conformation during the life-time of the enzyme-DNA
complex, a single reaction of the EcoRI enzyme cleaves either one or both
strands of the DNA.  Reaction profiles with other restriction enzymes differ
from EcoRI, though the underlying mechanisms may be the same.

<>

<1>Halford, S.E.
<2>The (billion dollar) consequences of studying why certain isolates of phage lambda infect only certain strains of E. coli: restriction enzymes.
<3>Biochemist
<4>31
<5>10-13
<6>2009
<7>In 1953, Bertani and Weigle1 reported that samples of bacteriophage ë that had been propagated
on certain strains of Escherichia coli retained the ability to infect that same strain of E.
coli but were unable to infect other strains. A contemporary version of their study is shown
in Figure 1; phage ë, obtained by infecting E. coli strain K (ëK), is applied to two different
E. coli strains, K and B. The ëK particles infect E. coli K with an efficiency of 1: i.e.,
every phage produces a plaque in a lawn of E. coli K cells. But the same preparation of ëK
shows a pathetic efficiency of infection on E. coli B, about 10-4: i.e., 10,000 phage
particles yield just one plaque on the lawn of E. coli B cells. The phage obtained from the
few productive infections of E. coli B (ëB) are then tested against the same two strains, K
and B. This time, the phage have largely lost the ability to infect E. coli K, as they now
show an efficiency of 10-4 instead of 1, but they infect E. coli B much more readily than
before, with an efficiency raised from 10-4 to 1.

<>

<1>Halford, S.E., Baldwin, G.S., Vipond, I.B.
<2>DNA recognition by EcoRV.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>152
<6>1993
<7>In the presence of magnesium ions, the EcoRV restriction endonuclease cleaves DNA specifically
at its recognition sequence, GATATC.  Sequences that differ from the recognition site by one
base pair are cleaved at least a million times more slowly.  Yet, in binding to DNA in the
absence of magnesium ions, the EcoRV restriction enzyme shows no sequence specificity.  The
protein binds all sequences with equal affinity, and it can transfer readily from one site to
another along the DNA molecule without dissociating from the DNA.  However, the DNA cleavage
activity that is observed at any particular sequence is a function of the fractional
saturation of the relevant enzyme-DNA complex with magnesium ions.  The observed difference in
cleavage rates is due to the fact that the EcoRV enzyme has a high affinity for magnesium when
it is located at its recognition site on DNA, but it has a low affinity for magnesium when it
is located on any other DNA sequence.  Once it has bound the metal ion, the intrinsic activity
of the EcoRV enzyme at noncognate sites is similar to that at the recognition site.  This
mechanism can be correlated to the crystal structures of the EcoRV endonuclease that have been
determined by F.K. Winkler.  Three structures were solved: the free enzyme in the absence of
DNA; the specific complex at its recognition sequence, a nonspecific complex at a different
DNA sequence.  The specific complex displays a large number of sequence-specific interactions
between the protein and the DNA.  These are missing in the nonspecific complex.  But the
additional interactions that are seen in the specific complex contribute nothing to the net
free energy change for DNA binding.  Instead, all of the energy from the specific interactions
is used to distort the DNA, and in other conformational changes: the structure of the specific
DNA bound to EcoRV is extensively distorted while the nonspecific DNA is close to B-form.  The
distortion results in only the specific complex being able to bind magnesium ions and to
catalyse the reaction.  The communication between DNA recognition and catalysis thus seems to
be mediated by the structure of the DNA itself.  This model is currently being tested by
mutational analyses of EcoRV and by using fluorescence methods to monitor the conformational
changes in the DNA and in the protein.

<>

<1>Halford, S.E., Bilcock, D.T., Stanford, N.P., Williams, S.A., Milsom, S.E., Gormley, N.A., Watson, M.A., Bath, A.J., Embleton, M.L., Gowers, D.M., Daniels, L.E., Parry, S.H., Szczelkun, M.D.
<2>Restriction endonuclease reactions requiring two recognition sites.
<3>Biochem. Soc. Trans.
<4>27
<5>696-699
<6>1999
<7>The standard perception of a restriction enzyme is of an endonuclease that recognizes a short
palindrome of DNA and which cleaves both strands of the DNA at fixed locations in that
sequence, in a reaction that requires only Mg2+ ions as a cofactor.  The perception thus
refers to the type II restriction enzymes, as opposed to the type I and the type III systems.
Many of the type II restriction enzymes conform to this perception.  Their recognition sites
are often symmetrical palindromes, 4, 6 or occasionally 8 bp long, although in some instances
the palindrome is interrupted by a fixed length of unspecified sequence.  They are usually
dimers of identical subunits that interact symmetrically with their recognition sequences,
even when the recognition site is an interrupted palindrome.  One active site in the dimer
cleaves one strand of the DNA, while the second active site cleaves the symmetrically
equivalent position in the other strand of the DNA.  The reactions of these enzymes are thus
independent events at individual recognition sites, unless the enzyme hops along the DNA from
one site to another.  The type II enzymes that match the standard perception include several
well-characterized endonucleases, such as EcoRV, BglI and BamHI.

<>

<1>Halford, S.E., Catto, L.E., Pernstich, C., Rusling, D.A., Sanders, K.L.
<2>The reaction mechanism of FokI excludes the possibility of targeting zinc finger nucleases to unique DNA sites.
<3>Biochem. Soc. Trans.
<4>39
<5>584-588
<6>2011
<7>The FokI endonuclease is a monomeric protein with discrete DNA-recognition and catalytic
domains. The latter has only one active site so, to cut both
strands, the catalytic domains from two monomers associate to form a
dimer. The dimer involving a monomer at the recognition site and another
from free solution is less stable than that from two proteins tethered to
the same DNA. FokI thus cleaves DNA with two sites better than one-site
DNA. The two sites can be immediately adjacent, but they can alternatively
be many hundreds of base pairs apart, in either inverted or repeated
orientations. The catalytic domain of FokI is often a component of zinc
finger nucleases. Typically, the zinc finger domains of two such nucleases
are designed to recognize two neighbouring DNA sequences, with the
objective of cutting the DNA exclusively between the target sequences.
However, this strategy fails to take account of the fact that the
catalytic domains of FokI can dimerize across distant sites or even at a
solitary site. Additional copies of either target sequence elsewhere in
the chromosome must elicit off-target cleavages.

<>

<1>Halford, S.E., Goodall, A.J.
<2>Modes of DNA cleavage by the EcoRV restriction endonuclease.
<3>Biochemistry
<4>27
<5>1771-1777
<6>1988
<7>The mechanism of action ofthe EcoRV restriction endonuclease at its single
recognition site on the plasmid pAT153 was analyzed by kinetic methods.  In
reactions at pH 7.5, close to the optimum for this enzyme, both strands of the
DNA were cut in a single concerted reaction:  DNA cut in only one strand of the
duplex was neither liberated from the enzyme during the catalytic turnover nor
accumulated as a steady-state intermediate.  In contrast, reactions at pH 6.0
involved the sequential cutting of the two strands of the DNA.  Under these
conditions, DNA cut in a single strand was an obligatory intermediate in the
reaction pathway and a fraction of the nicked DNA dissociated from the enzyme
during the turnover.  The different reaction profiles are shown to be
consistent with a single mechanism in which the kinetic activity of each
subunit of the dimeric protein is governed by its affinity for Mg2+ ions.  At
pH 7.5, Mg2+ is bound to both subunits of the dimer for virtually the complete
period of the catalytic turnover, while at pH 6.0 Mg2+ is bound transiently to
one subunit at a time.  The kinetics of the EcoRV nuclease were unaffected by
the DNA supercoiling.

<>

<1>Halford, S.E., Gowers, D.M., Sessions, R.B.
<2>Two are better than one.
<3>Nat. Struct. Biol.
<4>7
<5>705-707
<6>2000
<7>A crystal structure of a tetrameric restriction enzyme, NgoMIV, bound to two DNA duplexes has
been determined.  Two subunits contact each duplex in much the same manner as a dimeric
restriction enzyme recognizing a single site, but the dimeric units are packed back-to-back,
placing the duplexes on opposite sides of the tetramer.  Interaction with two recognition
sites, presumably via looping, enhances NgoMIV activity.

<>

<1>Halford, S.E., Johnson, N.P.
<2>The EcoRI restriction endonuclease, covalently closed DNA and ethidium bromide.
<3>Biochem. J.
<4>199
<5>767-777
<6>1981
<7>The reactions of the EcoRI restriction endonuclease on the covalently closed
DNA of plamid pMB9 were studied in the presence of ethidium bromide.  At the
concentrations of ethidium bromide tested, which covered the range over which
the DNA is changed from negatively to positively supercoiled, the dye caused no
alteration to the rate at which this enzyme cleaved the covalently closed DNA
to yield the open-circle form, but the rate at which these open circles were
cleaved to the linear product could be inhibited.  The fluorescence change,
caused by ethidium bromide binding with different stoichiometries to covalently
clodsed and open-circle DNA, provided a direct and sensitive signal for
monitoring the cleavage of DNA by this enzyme.  This method was used for a
steady-state kinetic analysis of the reaction catalysed by the EcoRI
restriction enzyme.  Reaction mechanisms where a complex between DNA and Mg2+
is the substrate for this enzyme were eliminated, and instead DNA and Mg2+ must
bind to the enzyme in separate stages.  The requisite controls for this
fluorimetric assay in both steady-state and transient kinetics studies, and its
application to other enzymes that alter the structure of covalently closed DNA,
are described.

<>

<1>Halford, S.E., Johnson, N.P.
<2>The EcoRI restriction endonuclease with bacteriophage lambda DNA.  Equilibrium binding sites.
<3>Biochem. J.
<4>191
<5>593-604
<6>1980
<7>The EcoRI restriction endonuclease was found by the filter binding technique to
form stable complexes, in the absence of Mg2+, with the DNA from derivatives of
bacteriophage lambda that either contain or lack EcoRI recognition sites.  The
amount of complex formed at different enzyme concentrations followed a
hyperbolic equilibrium-binding curve with DNA molecules containing EcoRI
recognition sites, but a sigmoidal equilibrium-binding curve was obtained with
a DNA molecule lacking EcoRI recognition sites.  The EcoRI enzyme displayed the
same affinity for individual recognition sites on lambda DNA, even under
conditions where it cleaves these sites at different rates.  The binding of the
enzyme to a DNA molecule lacking EcoRI sites was decreased by Mg2+.  These
observations indicate that (a) the EcoRI restriction enzyme binds
preferentially to its recognition site on DNA, and that different reaction
rates at different recognition sites are due to the rate of breakdown of this
complex; (b) the enzyme also binds to other DNA sequences, but that two
molecules of enzyme, in a different protein conformation, are involved in the
formation of the complex at non-specific sequences; (c) the different
affinities of the enzyme for the recognition site and for other sequences on
DNA, coupled with the different protein conformations, account for the
specificity of this enzyme for the cleavage of DNA at its recognition site; (d)
the decrease in the affinity of the enzyme for DNA, caused by Mg2+, liberates
binding energy from the DNA-protein complex that can be used in the catalytic
reaction.

<>

<1>Halford, S.E., Johnson, N.P.
<2>Single turnovers of the EcoRI restriction endonuclease.
<3>Biochem. J.
<4>211
<5>405-415
<6>1983
<7>Single turnovers of the EcoRI restriction endonuclease, cleaving its recognition site on the
covalently closed form of plasmid pMB9, were examined. Two methods were used to monitor the
progress of the reactions: one involved quenching the reaction at various times followed by
the electrophoretic separation of the products cleaved in one and in both strands of the
duplex; the other employed a stopped-flow fluorimeter to measure the amount of ethidium
bromide bound to the DNA as it changes when the DNA, cleaved in at least one strand,
dissociates from the enzyme.  Two procedures were used to initiate the reactions.  For some,
one solution containing the enzyme was mixed with a second containing both DNA and MgCl2: in
these reactions, the fluorescence changed at the same rate as the cleavage of the first strand
of the duplex. Other reactions were started by the addition of MgCl2 to a pre-equilibrium of
enzyme and DNA: here, both strands of the DNA were cleaved faster than before, with the
fluorescence signal now occurring at the same time as the cleavage of the second strand.  The
different kinetics from the two assays and the two mixing procedures are consistent with the
rates of these reactions being controlled by protein conformational changes.  These may affect
either one subunit alone within the dimeric EcoRI enzyme, allowing the enzyme to cleave only
one strand of the DNA in each turnover.  Alternatively, both subunits of the dimer may change,
so that the enzyme then cleaves both strands during the life-time of one enzyme-DNA complex.

<>

<1>Halford, S.E., Johnson, N.P., Grinsted, J.
<2>The EcoRI restriction endonuclease with bacteriophage lambda DNA.  Kinetic studies.
<3>Biochem. J.
<4>191
<5>581-592
<6>1980
<7>The kinetics of the reactions of the EcoRI restriction endonuclease at
individual recognition sites on the DNA from bacteriophage lambdawere found to
differ markedly from site to site.  Under certain conditions of pH and ionic
strength, the rates for the cleavage of the DNA were the same at each
recognition site.  But under altered experimental conditions, different
reaction rates were observed at each recognition site.  These results are
consistent with a mechanism in which the kinetic stability of the complex
between the enzyme and the recognition site on the DNA differs among the sites,
due to the effect of interactions between the enzyme and DNA sequences
surrounding each recognition site upon the transition state of the reaction.
Reactions at individual sites on a DNA molecule containing more than one
recognition site were found to be independent of each other, thus excluding the
possibility of a processive mechanism for the EcoRI enzyme.  The consequences
of these observations are discussed with regard to both DNA-protein
interactions and to the application of restriction enzymes in the study of the
structure of DNA molecules.

<>

<1>Halford, S.E., Johnson, N.P., Grinsted, J.
<2>The reactions of the EcoRI and other restriction endonucleases.
<3>Biochem. J.
<4>179
<5>353-365
<6>1979
<7>The reaction of the EcoRI restriction endonuclease was studied with both the
plasmid pMB9 and DNA from bacteriophage lambda as the substrates.  With both
circular and linear DNA molecules, the only reaction catalysed by the EcoRI
restriction endonuclease was the hydrolysis of the phosphodiester bond within
one strand of the recognition site on the DNA duplex.  The cleavage of both
strands of the duplex was achieved only after two independent reactions, each
involving a single-strand scission.  The reactivity of the enzyme for
single-strand scissions was the same for both the first and the second cleavage
within its recognition site.  No differences were observed between the
mechanism of action on supercoiled and linear DNA substrates.  Other
restriction endonucleases were tested against plasmid pMB9.  The HindIII
restriction endonuclease cleaved DNA in the same manner as the EcoRI enzyme.
However, in contrast with EcoRI, the SalI and the BamHI restriction
endonucleases appeared to cleave both strands of the DNA duplex almost
simultaneously.  The function of symmetrical DNA sequences and the conformation
of the DNA involved in these DNA-protein interactions are discussed in the
light of these observations.  The fact that the same reactions were observed on
both supercoiled and linear DNA substrates implies that these interactions do
not involve the unwinding of the duplex before catalysis.

<>

<1>Halford, S.E., Lovelady, B.M., McCallum, S.
<2>Altered specificity of the EcoRV restriction endonuclease.
<3>Biochem. Soc. Trans.
<4>14
<5>260-261
<6>1986
<7>Type II restriction endonucleases recognize specific sequences of nucleotides on DNA and, in
the presence of Mg2+, cleave both strands of the DNA at fixed locations relative to the
recognition site.  These enzymes generally show very high specificities: the rates at which
they cleave their recognition sites can be several orders of magnitude faster than that for
any cleavage at alternative DNA sequences.  However, in altered environments such as at high
pH or in the presence of water-miscible organic solvents, many restriction enzymes show
reduced specificity and cleave DNA at several sites in addition to the recognition site.  This
phenomena was first observed with the EcoRI restriction enzyme, the altered activity at high
pH being noted as EcoRI.  We describe here a similar alteration in the specificity of the
EcoRV restriction enzyme and show that each additional site cleaved by EcoRV differs from the
canonical EcoRV recognition sequence, 5'-G-A-T-A-T-C-3', by one nucleotide.

<>

<1>Halford, S.E., Lovelady, B.M., McCallum, S.A.
<2>Relaxed specificity of the EcoRV restriction endonuclease.
<3>Gene
<4>41
<5>173-181
<6>1986
<7>The EcoRV restriction endonuclease normally shows a high specificity for its recognition site
on DNA, GATATC. In standard reactions, it cleaves DNA at this site several orders of magnitude
more readily than at any alternative sequence. But in the presence of dimethyl sulphoxide and
at high pH, the EcoRV enzyme cleaves DNA at several sites that differ from its recognition
site by one nucleotide. Of the 18 (3 x 6) possible sequences that differ from GATATC by one
base, all were cleaved readily except for the following 4 sites: TATATC, CATATC, GATATA and
GATATG. However, two of the sites that could be cleaved by EcoRV in the presence of dimethyl
suphoxide, GAGATC and GATCTC, were only cleaved on DNA that lacked dam methylation: both
contain the sequence GATC, the recognition site for the dam methylase of Escherichia coli.

<>

<1>Halford, S.E., Marko, J.F.
<2>How do site-specific DNA-binding proteins find their targets?
<3>Nucleic Acids Res.
<4>32
<5>3040-3052
<6>2004
<7>Essentially all the biological functions of DNA depend on site-specific DNA-binding proteins
finding their targets, and therefore ?searching? through megabases of non-target DNA. In this
article, we review current understanding of how this sequence searching is done. We review how
simple diffusion through solution may be unable to account for the rapid rates of association
observed in experiments on some model systems, primarily the Lac repressor. We then present a
simplified version of the ?facilitated diffusion? model of Berg, Winter and von Hippel,
showing how non-specific DNA?protein interactions may account for accelerated targeting, by
permitting the protein to sample many binding sites per DNA encounter. We discuss the
1-dimensional ?sliding? motion of protein along non-specific DNA, often proposed to be the
mechanism of this multiple site sampling, and we discuss the role of short-range diffusive
?hopping? motions. We then derive the optimal range of sliding for a few physical situations,
including simple models of chromosomes in vivo, showing that a sliding range of ~100bp before
dissociation optimizes targeting in vivo. Going beyond first-order binding kinetics, we
discuss how processivity, the interaction of a protein with two or more targets on the same
DNA, can reveal the extent of sliding and we review recent experiments studying processivity
using the restriction enzyme EcoRV. Finally, we discuss how single molecule techniques might
be used to study the dynamics of DNA site-specific targeting of proteins.

<>

<1>Halford, S.E., Taylor, J.D., Vermote, C.L.M., Vipond, I.B.
<2>Mechanism of action of restriction endonuclease EcoRV.
<3>Nucl. Acids and Mol. Biol., Springer-Verlag, Eckstein, F., Lilley, D.M.J., Berlin
<4>7
<5>47-69
<6>1993
<7>Type II restriction/modification (R/M) systems, such as EcoRV, consist of two enzymes that act
at the same DNA sequence; a modification methyltransferase and a restriction endonuclease
(Bennett and Halford 1989; Wilson and Murray 1991). For EcoRV, the recognition sequence is
GATATC (Kholmina et al. 1980). The EcoRV modification enzyme methylates the first adenine
within this sequence (Nwosu et al. 1988) while, in the presence of Mg2+ ions, the EcoRV
restriction enzymes cleaves DNA specifically at this site (Schilkdraut et al. 1984). The
endonuclease cuts both strands at the center of the sequence, to leave blunt-ended DNA
fragments. However, prior methylation of the recognition site blocks restriction activity. The
basic tenet of R/M systems is that, in vivo, the restriction enzyme cuts only DNA molecules
that have not been exposed previously to the methyltransferase. Hence, they enable the cell to
degrade foreign DNA entering the cell without destroying host DNA. E. coli strains carrying
the EcoRV system show this phenotype (Bougueleret et al. 1984).

<>

<1>Halford, S.E., Taylor, J.D., Vermote, C.L.M., Vipond, I.B.
<2>Assays for restriction endonucleases using plasmid substrates.
<3>Methods Mol. Biol.
<4>30
<5>385-396
<6>1994
<7>A type II restriction enzyme purchased from a commercial supplier comes with a specified
number of units of enzyme activity. The units of restriction enzyme activity are defined by
the minimal amount of enzyme needed to complete the digestion of 1 ug of bacteriophage lambda
DNA in 1 h. These units are usually measured by making serial dilutions of the stock solution
of the enzyme, adding 1 uL from each dilution to 1 ug of phage lambda DNA in a suitable
buffer, incubating the reactions for 1 h at 37oC, and then analyzing the DNA by
electrophoresis through agarose. This is, at best, a semiquantitative assay. It cannot yield
quantitative data about the rate of the reaction of a restricton enzyme on a DNA substrate.

<>

<1>Halford, S.E., Welsh, A.J., Szczelkun, M.D.
<2>Enzyme-mediated DNA looping.
<3>Annu. Rev. Biophys. Biomol. Struct.
<4>33
<5>1-24
<6>2004
<7>Most reactions on DNA are carried out by multimeric protein complexes that interact with two
or more sites in the DNA and thus loop out the DNA
between the sites. The enzymes that catalyze these reactions usually have
no activity until they interact with both sites. This review examines the
mechanisms for the assembly of protein complexes spanning two DNA sites
and the resultant triggering of enzyme activity. There are two main routes
for bringing together distant DNA sites in an enzyme complex: either the
proteins bind concurrently to both sites and capture the intervening DNA
in a loop, or they translocate the DNA between one site and another into
an expanding loop, by an energy-dependent translocation mechanism. Both
capture and translocation mechanisms are discussed here, with reference to
the various types of restriction endonuclease that interact with two
recognition sites before cleaving DNA.

<>

<1>Hall, M.F., Noren, C.J., Perler, F.B., Schildkraut, I.
<2>Creation of an artificial bifunctional intein by grafting a homing endonuclease into a mini-intein.
<3>J. Mol. Biol.
<4>323
<5>173-179
<6>2002
<7>The majority of inteins are comprised of a protein splicing domain and a homing endonuclease
domain. Experimental evidence has demonstrated that the splicing domain and the endonuclease
domain in a bifunctional intein are largely independent of each other with respect to both
structure and activity. Here, an artificial bifunctional intein has been created through the
insertion of an existing homing endonuclease into a mini-intein that is naturally lacking this
functionality. The gene for I-CreI, an intron-encoded homing endonuclease, was grafted into
the monofunctional Mycobacterium xenopi GyrA intein at the putative site of the missing
endonuclease. The resulting fusion protein was found to be capable of protein splicing similar
to that of the parent intein. In addition, the protein demonstrated site-specific endonuclease
activity that is characteristic of the I-CreI homing endonuclease. The function of each domain
therefore remained unaffected by the presence of the other domain. This artificial fusion of
the two domains is a potential novel mobile genetic element.

<>

<1>Hall, R.K., Larcom, L.L.
<2>Blockage of restriction endonuclease cleavage by thymine dimers.
<3>Photochem. Photobiol.
<4>36
<5>429-432
<6>1982
<7>Viral DNAs were subjected to 254 nm irradiation and then digested with type II
restriction endonucleases.  At the fluences used, irradiation inhibited
cleavage by nucleases which recognize sites containing neighboring thymines.
Cleavage by endonucleases with other recognition sequences was not affected.
In viral and plasmid DNAs, this effect could be used to study thymine dimer
formation at a few specific, mapped sites of defined base sequence.

<>

<1>Hall, R.M.
<2>The DNA adenine methyltransferase (dam+) gene of bacteriophage T4 reverses the mutator phenotype of an Escherichia coli dam mutant.
<3>J. Bacteriol.
<4>172
<5>2812-2813
<6>1990
<7>The mutator phenotype of Escherichia coli dam mutants was found to be reversed
by introduction of the bacteriophage T4 gene for DNA adenine methyltransferase.
This precludes a direct role for the E. coli DNA adenine methyltransferase in
mismatch repair, in addition to its role in strand discrimination, as suggested
by earlier studies.

<>

<1>Hallam, S.J., Konstantinidis, K.T., Putnam, N., Schleper, C., Watanabe, Y., Sugahara, J., Preston, C., de la Torre, J., Richardson, P.M., DeLong, E.F.
<2>Genomic analysis of the uncultivated marine crenarchaeote Cenarchaeum symbiosum.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>18296-18301
<6>2006
<7>Crenarchaeota are ubiquitous and abundant microbial constituents of soils, sediments, lakes,
and ocean waters. To further describe the cosmopolitan
nonthermophilic Crenarchaeota, we analyzed the genome sequence of one
representative, the uncultivated sponge symbiont Cenarchaeum symbiosum. C.
symbiosum genotypes coinhabiting the same host partitioned into two
dominant populations, corresponding to previously described a- and b-type
ribosomal RNA variants. Although they were syntenic, overlapping a- and
b-type ribotype genomes harbored significant variability. A single tiling
path comprising the dominant a-type genotype was assembled and used to
explore the genomic properties of C. symbiosum and its planktonic
relatives. Of 2,066 ORFs, 55.6% matched genes with predicted function from
previously sequenced genomes. The remaining genes partitioned between
functional RNAs (2.4%) and hypotheticals (42%) with limited homology to
known functional genes. The latter category included some genes likely
involved in the archaeal-sponge symbiotic association. Conversely, 525 C.
symbiosum ORFs were most highly similar to sequences from marine
environmental genomic surveys, and they apparently represent orthologous
genes from free-living planktonic Crenarchaeota. In total, the C.
symbiosum genome was remarkably distinct from those of other known Archaea
and shared many core metabolic features in common with its free-living
planktonic relatives.

<>

<1>Hallam, S.J., Mincer, T.J., Schleper, C., Preston, C.M., Roberts, K., Richardson, P.M., DeLong, E.F.
<2>Pathways of carbon assimilation and ammonia oxidation suggested by environmental genomic analyses of marine Crenarchaeota.
<3>PLoS Biology
<4>4
<5>e95
<6>2006
<7>Marine Crenarchaeota represent an abundant component of oceanic microbiota with potential to
significantly influence biogeochemical cycling in marine
ecosystems. Prior studies using specific archaeal lipid biomarkers and
isotopic analyses indicated that planktonic Crenarchaeota have the
capacity for autotrophic growth, and more recent cultivation studies
support an ammonia-based chemolithoautotrophic energy metabolism. We
report here analysis of fosmid sequences derived from the uncultivated
marine crenarchaeote, Cenarchaeum symbiosum, focused on the reconstruction
of carbon and energy metabolism. Genes predicted to encode multiple
components of a modified 3-hydroxypropionate cycle of autotrophic carbon
assimilation were identified, consistent with utilization of carbon
dioxide as a carbon source. Additionally, genes predicted to encode a near
complete oxidative tricarboxylic acid cycle were also identified,
consistent with the consumption of organic carbon and in the production of
intermediates for amino acid and cofactor biosynthesis. Therefore, C.
symbiosum has the potential to function either as a strict autotroph, or
as a mixotroph utilizing both carbon dioxide and organic material as
carbon sources. From the standpoint of energy metabolism, genes predicted
to encode ammonia monooxygenase subunits, ammonia permease, urease, and
urea transporters were identified, consistent with the use of reduced
nitrogen compounds as energy sources fueling autotrophic metabolism.
Homologues of these genes, recovered from ocean waters worldwide,
demonstrate the conservation and ubiquity of crenarchaeal pathways for
carbon assimilation and ammonia oxidation. These findings further
substantiate the likely global metabolic importance of Crenarchaeota with
respect to key steps in the biogeochemical transformation of carbon and
nitrogen in marine ecosystems.

<>

<1>Hallam, S.J., Putnam, N., Preston, C.M., Detter, J.C., Rokhsar, D., Richardson, P.M., DeLong, E.F.
<2>Reverse methanogenesis: Testing the hypothesis with environmental genomics.
<3>Science
<4>305
<5>1457-1462
<6>2004
<7>Microbial methane consumption in anoxic sediments significantly impacts the global environment
by reducing the flux of greenhouse gases from ocean to atmosphere. Despite its significance,
the biological mechanisms controlling anaerobic methane oxidation are not well characterized.
One current model suggests that relatives of methane-producing Archaea developed the capacity
to reverse methanogenesis and thereby to consume methane to produce cellular carbon and
energy. We report here a test of the "reverse-methanogenesis" hypothesis by genomic analyses
of methane-oxidizing Archaea from deep-sea sediments. Our results show that nearly all genes
typically associated with methane production are present in one specific group of archaeal
methanotrophs. These genome-based observations support previous hypotheses and provide an
informed foundation for metabolic modeling of anaerobic methane oxidation.

<>

<1>Hallet, B.
<2>Playing Dr Jekyll and Mr Hyde: combined mechanisms of phase variation in bacteria.
<3>Curr. Opin. Microbiol.
<4>4
<5>570-581
<6>2001
<7>Phase variation is the adaptive process by which bacteria undergo frequent and reversible
phenotypic changes resulting from genetic alterations in specific loci
of their genomes. This process is crucial for the survival of pathogens and
commensals in hostile and ever-changing host environments. Despite important
differences in the molecular mechanisms that mediate and regulate phase
variation, related strategies have evolved to generate high levels of genetic
diversity through complex and combinatorial reshuffling of genetic information.
Recent studies, supported by the emergence of global genomic approaches, have
revealed that bacterial pathogens often use a combination of different mechanisms
to vary the expression of a variety of biological functions, providing new
insights into bacterial adaptation and virulence mechanisms. Recent advances in
the understanding of the molecular mechanisms of phase variation are reviewed,
and differences in these mechanisms outlined.

<>

<1>Halling, S.M., Peterson-Burch, B.D., Bricker, B.J., Zuerner, R.L., Qing, Z., Li, L.L., Kapur, V., Alt, D.P., Olsen, S.C.
<2>Completion of the genome sequence of Brucella abortus and comparison to the highly similar genomes of Brucella melitensis and Brucella suis.
<3>J. Bacteriol.
<4>187
<5>2715-2726
<6>2005
<7>Brucellosis is a worldwide disease of humans and livestock that is caused by a number of very
closely related classical Brucella species in the
alpha-2 subdivision of the Proteobacteria. We report the complete genome
sequence of Brucella abortus field isolate 9-941 and compare it to those
of Brucella suis 1330 and Brucella melitensis 16 M. The genomes of these
Brucella species are strikingly similar, with nearly identical genetic
content and gene organization. However, a number of insertion-deletion
events and several polymorphic regions encoding putative outer membrane
proteins were identified among the genomes. Several fragments previously
identified as unique to either B. suis or B. melitensis were present in
the B. abortus genome. Even though several fragments were shared between
only B. abortus and B. suis, B. abortus shared more fragments and had
fewer nucleotide polymorphisms with B. melitensis than B. suis. The
complete genomic sequence of B. abortus provides an important resource for
further investigations into determinants of the pathogenicity and
virulence phenotypes of these bacteria.

<>

<1>Ham, J.S., Kim, H.W., Seol, K.H., Jang, A., Jeong, S.G., Oh, M.H., Kim, D.H., Kang, D.K., Kim, G.B., Cha, C.J.
<2>Genome Sequence of Lactobacillus salivarius NIAS840, Isolated from Chicken Intestine.
<3>J. Bacteriol.
<4>193
<5>5551-5552
<6>2011
<7>Lactobacillus salivarius is a well-known lactic acid bacterium to which increasing attention
has been paid recently for use as probiotics for
humans and animals. L. salivarius NIAS840 was first isolated from broiler
chicken feces, displaying antimicrobial activities against
multidrug-resistant Staphylococcus aureus and Salmonella enterica serovar
Typhimurium. Here, we report the genome sequence of L. salivarius NIAS840
(2,046,557 bp) including a small plasmid and two megaplasmids.

<>

<1>Ham, J.S., Lee, T., Byun, M.J., Lee, K.T., Kim, M.K., Han, G.S., Jeong, S.G., Oh, M.H., Kim, D.H., Kim, H.
<2>Complete Genome Sequence of Bifidobacterium longum subsp. longum KACC 91563.
<3>J. Bacteriol.
<4>193
<5>5044
<6>2011
<7>Bifidobacterium longum strains predominate the colonic microbiota of breast-fed infants. Here
we report a complete genome sequence of B. longum
subsp. longum KACC 91563 isolated from feces of neonates. A single
circular chromosome of 2,385,301 bp contains 1,980 protein coding genes,
56 tRNA genes, and 3 rRNA operons.

<>

<1>Hamablet, L., Chen, G.C., Brown, A., Roberts, R.J.
<2>LpnI, from Legionella pneumophila, is a neoschiozmer of HaeII.
<3>Nucleic Acids Res.
<4>17
<5>6417
<6>1989
<7>LpnI is a Type II restriction endonuclease that was previously isolated from Legionella
pneumophila strain 11 EJ and partially purified (1). Further purification by phosphocellulose
and DNA-agrose chromatography, with an intermediate 50-75% ammonium sulphate
concentration/fractionation step gave enzyme sufficiently pure for detailed characterization.
LpnI cleaves pUC19 DNA at three sites. Double digests of pUC19 DNA with LpnI and either AatII,
EcoRI, PvuI or RsaI mapped the LpnI cleavage site to approximately 230, 690 and 1090
nucleotides. These sites lie close to those predicted for HaeII. Double digest between HaeII
and LpnI on bacteriophage lambda DNA confirmed that these enzymes are isoschizomers (Fig 1a).

<>

<1>Hamada, M., Ichikawa, N., Oguchi, A., Fujita, N.
<2>Draft Genome Sequence of Lysinimicrobium mangrovi NBRC 105856T, Isolated from the Rhizosphere of a Mangrove.
<3>Genome Announcements
<4>2
<5>e01131-14
<6>2014
<7>Here, we report the draft genome sequence of the only species of the genus Lysinimicrobium,
Lysinimicrobium mangrovi NBRC 105856(T), isolated from the
rhizosphere of a mangrove. The first genomic sequence of this genus and species
presented here will facilitate taxonomical, ecological, and functional studies of
this rare actinobacterial group.

<>

<1>Hammerl, J.A., Jackel, C., Reetz, J., Beck, S., Alter, T., Lurz, R., Barretto, C., Brussow, H., Hertwig, S.
<2>Campylobacter jejuni Group III Phage CP81 Contains Many T4-Like Genes without Belonging to the T4-Type Phage Group: Implications for the Evolution of T4 Phages.
<3>J. Virol.
<4>85
<5>8597-8605
<6>2011
<7>CP81 is a virulent Campylobacter group III phage whose linear genome
comprises 132,454 bp. At the nucleotide level, CP81 differs from other
phages. However, a number of its structural and replication/recombination
proteins revealed a relationship to the group II Campylobacter phages
CP220/CPt10 and to T4-type phages. Unlike the T4-related phages, the CP81
genome does not contain conserved replication and virion modules. Instead,
the respective genes are scattered throughout the phage genome. Moreover,
most genes for metabolic enzymes of CP220/CPt10 are lacking in CP81. On
the other hand, the CP81 genome contains nine similar genes for homing
endonucleases which may be involved in the attrition of the conserved gene
order for the virion core genes of T4-type phages. The phage apparently
possesses an unusual modification of C or G bases. Efficient cleavage of
its DNA was only achieved with restriction enzymes recognizing pure A/T
sites. Uncommonly, phenol extraction leads to a significant loss of CP81
DNA from the aqueous layer, a property not yet described for other phages
belonging to the T4 superfamily.

<>

<1>Hammerl, J.A., Jackel, C., Reetz, J., Hertwig, S.
<2>The Complete Genome Sequence of Bacteriophage CP21 Reveals Modular Shuffling in Campylobacter Group II Phages.
<3>J. Virol.
<4>86
<5>8896
<6>2012
<7>Campylobacter group II phages described so far share a high degree of sequence
similarity. We report the 182,833-bp genomic sequence of the closely related
group II phage CP21 and show that it has a completely different genomic
organization. As in other group II phages, the CP21 genome is composed of large
modules separated by long DNA repeat regions which obviously trigger
recombination and modular shuffling.

<>

<1>Hammerl, J.A., Klein, I., Lanka, E., Appel, B., Hertwig, S.
<2>Genetic and functional properties of the self-transmissible Yersinia enterocolitica plasmid pYE854, which mobilizes the virulence plasmid pYV.
<3>J. Bacteriol.
<4>190
<5>991-1010
<6>2008
<7>Yersinia strains frequently harbor plasmids, of which the virulence
plasmid pYV, indigenous in pathogenic strains, has been thoroughly
characterized during the last decades. Yet, it has been unknown whether
the nonconjugative pYV can be transferred by helper plasmids naturally
occurring in this genus. We have isolated the conjugative plasmids pYE854
(95.5 kb) and pYE966 (70 kb) from a nonpathogenic and a pathogenic
Yersinia enterocolitica strain, respectively, and demonstrate that both
plasmids are able to mobilize pYV. The complete sequence of pYE854 has
been determined. The transfer proteins and oriT of the plasmid reveal
similarities to the F factor. However, the pYE854 replicon does not belong
to the IncF group and is more closely related to a plasmid of
gram-positive bacteria. Plasmid pYE966 is very similar to pYE854 but lacks
two DNA regions of the larger plasmid that are dispensable for
conjugation.

<>

<1>Hammond, A.W., Chatterjee, D.K.
<2>Cloning and expressing various restriction endonucleases and modification methylases from Neisseria.
<3>International Patent Office
<4>WO 9118916
<5>
<6>1991
<7>The present invention is directed to recombinant hosts which contain and express various Type
II restriction endonuclease and/or modification methylase genes.  In particular, the present
invention is concerned with the cloned restriction endonucleases, NgoAIII and NgoAI, which
recognize and cleave within or near the double-stranded DNA sequence, 5'CCGCGG3' and
5'PuGCGCPy3', respectively.  Also provided in this invention are cloned modification
methylase genes corresponding to said restriction endonucleases.  This invention is further
concerned with a cloned modification methylase, M.NgoAII.  One source of these enzymes is
Neisseria gonorrhoeae, although other microorganisms may be used to isolate the restriction
endonuclease isoschizomers and modification methylase isoschizomers of this invention.

<>

<1>Hammond, A.W., Chatterjee, D.K.
<2>Host expressing NgoAIII restriction endonuclease and modification methylase from Neisseria.
<3>US Patent Office
<4>US 5147800
<5>
<6>1992
<7>The present invention is directed to recombinant hosts which contain and express various Type
II restriction endonuclease and/or modification methylase genes.  In particular, the present
invention is concerned with the cloned restriction endonucleases, NgoAIII and NgoAI, which
recognize and cleave within or near the double-stranded DNA sequence, 5' CCGCGG 3' and 5'
PuGCGCPy 3', respectively.  Also provided in this invention are cloned modification methylase
genes corresponding to said restriction endonucleases.  This invention is further concerned
with a cloned modification methylase, M.NgoAII.  One source of these enzymes is Neisseria
gonorrhoeae, although other microorganisms may be used to isolate the restriction endonuclease
isoschizomers and modification methylase isoschizomers of this invention.

<>

<1>Hammond, A.W., Chatterjee, D.K., Gerard, G.F.
<2>Cloning and expression of restriction endonucleases from Neisseria.
<3>International Patent Office
<4>WO 9113975
<5>
<6>1991
<7>The present invention is directed to a restriction endonuclease which recognizes and cleaves
the palindromic sequence GCCGGC between the first G and C residues from the 5' end, producing
a four-base 5' extension. Also provided in this invention is a modification methylase
corresponding to said restriction endonuclease. One source of these enzymes is Neisseria
gonorrhoeae. This invention is also concerned with cloning and expressing the genes coding for
said restriction endonuclease and said modification methylase.

<>

<1>Hammond, A.W., Gerard, G.F., Campbell, J.H., Chatterjee, D.K.
<2>Characterization of a restriction enzyme from a strain of Neisseria gonorrhoea which recognizes 5'G^CCGGC3', an isoschizomer of NaeI.
<3>Nucleic Acids Res.
<4>17
<5>3320
<6>1989
<7>A type II restriction enzyme, NgoAIV, has been isolated from a strain of Neisseria gonorrhoea.
NgoAIV recognizes the palindromic sequence, 5'G^CCGGC3', and cleaves between the first G and
C residues producing a 4- base 5' extension.

<>

<1>Hammond, A.W., Gerard, G.F., Chatterjee, D.K.
<2>Characterization of NgoAIII, an isoschizomer of SstII from a strain of Neisseria gonorrhoea.
<3>Nucleic Acids Res.
<4>17
<5>6750
<6>1989
<7>Note that this enzyme has been renamed NgoFIII because the strain it is from is
FA1090 and not WR220, from which the NgoA series of enzymes have been reported.

<>

<1>Hammond, A.W., Gerard, G.F., Chatterjee, D.K.
<2>Cloning the KpnI restriction-modification system in Escherichia coli.
<3>Gene
<4>97
<5>97-102
<6>1991
<7>The genes encoding the KpnI restriction and modification (R-M) system from
Klebsiella pneumoniae, recognizing the sequence, 5'-GGTAC^C-3', were cloned and
expressed in Escherichia coli.  Although the restriction endonuclease (ENase)-
and methyltransferase (MTase)-encoding genes were closely linked, initial
attempts to clone both genes as a single DNA fragment in a plasmid vector
resulted in deletions spanning all or part of the gene coding for the ENase.
Initial protection of the E. coli host with MTase expressed on a plasmid was
required to stabilize a compatible plasmid carrying both the ENase- and the
MTase-encoding genes on a single DNA fragment.  However, once established, the
MTase activity can be supplied in cis to the kpnIR gene, without an extra copy
of kpnIM.  A chromosomal map was generated localizing the kpnIR and kpnIM genes
on 1.7-kb and 3.5-kb fragments, respectively.  A final E. coli strain was
constructed, AH29, which contained two compatible plasmids: an inducible
plasmid carrying the kpnIR gene which amplifies copy number at elevated
temperatures and a pBR322 derivative expressing M.KpnI.  This strain produces
approx. 10 million units of R.KpnI/g of wet-weight cells, which is several
1000-fold higher than the level of R.KpnI produced by K. pneumoniae.  In
addition, DNA methylated with M.KpnI in vivo does not appear to be restricted
by the mcrA, mcrB or mrr systems of E. coli.

<>

<1>Han, C. et al.
<2>Complete genome sequence of Treponema succinifaciens type strain (6091).
<3>Standards in Genomic Sciences
<4>4
<5>361-370
<6>2011
<7>Treponema succinifaciens Cwyk and Canale-Parola 1981 is of interest because this  strictly
anaerobic, apathogenic member of the genus Treponema oxidizes
carbohydrates and couples the Embden-Meyerhof pathway via activity of a
pyruvate-formate lyase to the production of acetyl-coenzyme A and formate. This
feature separates this species from most other anaerobic spirochetes. The genome
of T. succinifaciens 6091(T) is only the second completed and published type
strain genome from the genus Treponema in the family Spirochaetaceae. The
2,897,425 bp long genome with one plasmid harbors 2,723 protein-coding and 63 RNA
genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Han, C. et al.
<2>Complete genome sequence of Syntrophobotulus glycolicus type strain (FlGlyR).
<3>Standards in Genomic Sciences
<4>4
<5>371-380
<6>2011
<7>Syntrophobotulus glycolicus Friedrich et al. 1996 is currently the only member of the genus
Syntrophobotulus within the family Peptococcaceae. The species is of
interest because of its isolated phylogenetic location in the genome-sequenced
fraction of tree of life. When grown in pure culture with glyoxylate as carbon
source the organism utilizes glyoxylate through fermentative oxidation, whereas,
when grown in syntrophic co-culture with homoacetogenic or methanogenic bacteria,
it is able to oxidize glycolate to carbon dioxide and hydrogen. No other organic
or inorganic carbon source is utilized by S. glycolicus. The subdivision of the
family Peptococcaceae into genera does not reflect the natural relationships,
particularly regarding the genera most closely related to Syntrophobotulus. Both
Desulfotomaculum and Pelotomaculum are paraphyletic assemblages, and the
taxonomic classification is in significant conflict with the 16S rRNA data. S.
glycolicus is already the ninth member of the family Peptococcaceae with a
completely sequenced and publicly available genome. The 3,406,739 bp long genome
with its 3,370 protein-coding and 69 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Han, C. et al.
<2>Complete genome sequence of Thermaerobacter marianensis type strain (7p75a).
<3>Standards in Genomic Sciences
<4>3
<5>337-345
<6>2010
<7>Thermaerobacter marianensis Takai et al. 1999 is the type species of the genus
Thermaerobacter, which belongs to the Clostridiales family Incertae Sedis XVII.
The species is of special interest because T. marianensis is an aerobic,
thermophilic marine bacterium, originally isolated from the deepest part in the
western Pacific Ocean (Mariana Trench) at the depth of 10.897m. Interestingly,
the taxonomic status of the genus has not been clarified until now. The genus
Thermaerobacter may represent a very deep group within the Firmicutes or
potentially a novel phylum. The 2,844,696 bp long genome with its 2,375
protein-coding and 60 RNA genes consists of one circular chromosome and is a part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Han, C. et al.
<2>Complete genome sequence of the sulfur compounds oxidizing chemolithoautotroph Sulfuricurvum kujiense type strain (YK-1(T)).
<3>Standards in Genomic Sciences
<4>6
<5>94-103
<6>2012
<7>Sulfuricurvum kujiense Kodama and Watanabe 2004 is the type species of the monotypic genus
Sulfuricurvum, which belongs to the family Helicobacteraceae in
the class Epsilonproteobacteria. The species is of interest because it is
frequently found in crude oil and oil sands where it utilizes various reduced
sulfur compounds such as elemental sulfur, sulfide and thiosulfate as electron
donors. Members of the species do not utilize sugars, organic acids or
hydrocarbons as carbon and energy sources. This genome sequence represents the
type strain of the only species in the genus Sulfuricurvum. The genome, which
consists of a circular chromosome of 2,574,824 bp length and four plasmids of
118,585 bp, 71,513 bp, 51,014 bp, and 3,421 bp length, respectively, harboring a
total of 2,879 protein-coding and 61 RNA genes and is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Han, C. et al.
<2>Complete genome sequence of Kangiella koreensis type strain (SW-125).
<3>Standards in Genomic Sciences
<4>1
<5>226-233
<6>2009
<7>Kangiella koreensis (Yoon et al. 2004) is the type species of the genus and is of phylogenetic
interest because of the very isolated location of the genus
Kangiella in the gammaproteobacterial order Oceanospirillales. K. koreensis
SW-125(T) is a Gram-negative, non-motile, non-spore-forming bacterium isolated
from tidal flat sediments at Daepo Beach, Yellow Sea, Korea. Here we describe the
features of this organism, together with the complete genome sequence, and
annotation. This is the first completed genome sequence from the genus Kangiella
and only the fourth genome from the order Oceanospirillales. This 2,852,073 bp
long single replicon genome with its 2647 protein-coding and 48 RNA genes is part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Han, C. et al.
<2>Complete genome sequence of Pedobacter heparinus type strain (HIM 762-3).
<3>Standards in Genomic Sciences
<4>1
<5>54-62
<6>2009
<7>Pedobacter heparinus (Payza and Korn 1956) Steyn et al. 1998 comb. nov. is the type species of
the rapidly growing genus Pedobacter within the family
Sphingobacteriaceae of the phylum 'Bacteroidetes'. P. heparinus is of interest,
because it was the first isolated strain shown to grow with heparin as sole
carbon and nitrogen source and because it produces several enzymes involved in
the degradation of mucopolysaccharides. All available data about this species are
based on a sole strain that was isolated from dry soil. Here we describe the
features of this organism, together with the complete genome sequence, and
annotation. This is the first report on a complete genome sequence of a member of
the genus Pedobacter, and the 5,167,383 bp long single replicon genome with its
4287 protein-coding and 54 RNA genes is part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Han, C.S. et al.
<2>Pathogenomic Sequence Analysis of Bacillus cereus and Bacillus thuringiensis Isolates Closely Related to Bacillus anthracis.
<3>J. Bacteriol.
<4>188
<5>3382-3390
<6>2006
<7>Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis are closely related
gram-positive, spore-forming bacteria of the B. cereus
sensu lato group. While independently derived strains of B. anthracis
reveal conspicuous sequence homogeneity, environmental isolates of B.
cereus and B. thuringiensis exhibit extensive genetic diversity. Here we
report the sequencing and comparative analysis of the genomes of two
members of the B. cereus group, B. thuringiensis 97-27 subsp. konkukian
serotype H34, isolated from a necrotic human wound, and B. cereus E33L,
which was isolated from a swab of a zebra carcass in Namibia. These two
strains, when analyzed by amplified fragment length polymorphism within a
collection of over 300 of B. cereus, B. thuringiensis, and B. anthracis
isolates, appear closely related to B. anthracis. The B. cereus E33L
isolate appears to be the nearest relative to B. anthracis identified thus
far. Whole-genome sequencing of B. thuringiensis 97-27and B. cereus E33L
was undertaken to identify shared and unique genes among these isolates in
comparison to the genomes of pathogenic strains B. anthracis Ames and B.
cereus G9241 and nonpathogenic strains B. cereus ATCC 10987 and B. cereus
ATCC 14579. Comparison of these genomes revealed differences in terms of
virulence, metabolic competence, structural components, and regulatory
mechanisms.

<>

<1>Han, G.H., Kim, W., Chun, J., Kim, S.W.
<2>Draft Genome Sequence of Methylophaga aminisulfidivorans MPT.
<3>J. Bacteriol.
<4>193
<5>4265
<6>2011
<7>Methylophaga aminisulfidivorans MP(T) is a restricted facultatively marine methylotrophic
bacterium that grows on methanol, methylated amines,
dimethyl sulfide, and dimethyl sulfoxide. Here we present the high-quality
draft genome sequence of M. aminisulfidivorans MP(T) (KCTC 12909(T) = JCM
14647(T)), consisting of a chromosome (3,092,085 bp) and a plasmid (16,875
bp).

<>

<1>Han, J., Xu, H., Huang, J., Wu, J.
<2>Recombinant preparation of restriction endonuclease DUF820-2 by prokaryotic expression.
<3>Chinese Patent Office
<4>CN 102433350 A
<5>
<6>2012
<7>
<>

<1>Han, J., Zhang, F., Hou, J., Liu, X., Li, M., Liu, H., Cai, L., Zhang, B., Chen, Y., Zhou, J., Hu, S., Xiang, H.
<2>Complete Genome Sequence of the Metabolically Versatile Halophilic Archaeon Haloferax mediterranei, a Poly(3-Hydroxybutyrate-co-3-Hydroxyvalerate) Producer.
<3>J. Bacteriol.
<4>194
<5>4463-4464
<6>2012
<7>Haloferax mediterranei, an extremely halophilic archaeon, has shown promise for production of
poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) from unrelated
cheap carbon sources. Here we report the complete genome (3,904,707 bp) of H.
mediterranei CGMCC 1.2087, consisting of one chromosome and three megaplasmids.

<>

<1>Han, J.E., Kim, J.H., Choresca, C., Shin, S.P., Jun, J.W., Park, S.C.
<2>Draft Genome Sequence of a Clinical Isolate, Aeromonas hydrophila SNUFPC-A8, from a Moribund Cherry Salmon (Oncorhynchus masou masou).
<3>Genome Announcements
<4>1
<5>e00133-12
<6>2013
<7>We present the genome of a clinical isolate, Aeromonas hydrophila SNUFPC-A8, from a moribund
cherry salmon. The completed draft genome of this strain shows high
sequence homology to the reference strain A. hydrophila ATCC 7966 (NC008570.1)
and known plasmids pAsa2 and pAAk1 from other Aeromonas species (NC004925.1 and
NC019014.1).

<>

<1>Han, J.E., Kim, J.H., Shin, S.P., Jun, J.W., Chai, J.Y., Park, S.C.
<2>Draft Genome Sequence of Aeromonas salmonicida subsp. achromogenes AS03, an Atypical Strain Isolated from Crucian Carp (Carassius carassius) in the Republic   of Korea.
<3>Genome Announcements
<4>1
<5>e00791-13
<6>2013
<7>We present the draft genome sequence of Aeromonas salmonicida subsp. achromogenes strain AS03,
an atypical A. salmonicida strain that causes erythrodermatitis in
crucian carp (Carassius carassius). This is the first genome sequence report of
A. salmonicida subsp. achromogenes, one of the four subspecies of atypical A.
salmonicida.

<>

<1>Han, J.I., Choi, H.K., Lee, S.W., Orwin, P.M., Kim, J., Laroe, S.L., Kim, T.G., O'Neil, J., Leadbetter, J.R., Lee, S.Y., Hur, C.G., Spain, J.C., Ovchinnikova, G., Goodwin, L., Han, C.
<2>Complete genome sequence of the metabolically versatile plant growth-promoting endophyte, Variovorax paradoxus S110.
<3>J. Bacteriol.
<4>193
<5>1183-1190
<6>2010
<7>Variovorax paradoxus is a microorganism of special interest due to its diverse metabolic
capabilities, including the biodegradation of both
biogenic compounds and anthropogenic contaminants. V. paradoxus also
engages in mutually beneficial interactions with both bacteria and plants.
The complete genome sequence of V. paradoxus S110 is composed of 6,754,997
base pairs with 6,279 predicted protein-coding sequences within two
circular chromosomes. The genomic analysis has revealed multiple metabolic
features for autotrophic and heterotrophic lifestyles. These metabolic
diversities enable independent survival as well as a symbiotic lifestyle.
Consequently, S110 appears to have evolved into a superbly adaptable
microorganism, able to survive in ever-changing environmental conditions.
Based on our findings, we suggest V. paradoxus S110 as a potential
candidate for agrobiotechnological applications, such as biofertilizer and
biopesticide. Because it has many associations with other biota, it is
also suited to serve as an additional model system for studies of
microbe-plant and microbe-microbe interactions.

<>

<1>Han, J.I., Spain, J.C., Leadbetter, J.R., Ovchinnikova, G., Goodwin, L.A., Han, C.S., Woyke, T., Davenport, K.W., Orwin, P.M.
<2>Genome of the Root-Associated Plant Growth-Promoting Bacterium Variovorax paradoxus Strain EPS.
<3>Genome Announcements
<4>1
<5>e00843-13
<6>2013
<7>Variovorax paradoxus is a ubiquitous betaproteobacterium involved in plant growth promotion,
the degradation of xenobiotics, and quorum-quenching activity. The
genome of V. paradoxus strain EPS consists of a single circular chromosome of
6,550,056 bp, with a 66.48% G+C content.

<>

<1>Han, K., Li, Z.F., Peng, R., Zhu, L.P., Zhou, T., Wang, L.G., Li, S.G., Zhang, X.B., Hu, W., Wu, Z.H., Qin, N., Li, Y.Z.
<2>Extraordinary expansion of a Sorangium cellulosum genome from an alkaline milieu.
<3>Sci. Rep.
<4>3
<5>2101
<6>2013
<7>Complex environmental conditions can significantly affect bacterial genome size
by unknown mechanisms. The So0157-2 strain of Sorangium cellulosum is an
alkaline-adaptive epothilone producer that grows across a wide pH range. Here, we
show that the genome of this strain is 14,782,125 base pairs, 1.75-megabases
larger than the largest bacterial genome from S. cellulosum reported previously.
The total 11,599 coding sequences (CDSs) include massive duplications and
horizontally transferred genes, regulated by lots of protein kinases, sigma
factors and related transcriptional regulation co-factors, providing the So0157-2
strain abundant resources and flexibility for ecological adaptation. The
comparative transcriptomics approach, which detected 90.7% of the total CDSs, not
only demonstrates complex expression patterns under varying environmental
conditions but also suggests an alkaline-improved pathway of the insertion and
duplication, which has been genetically testified, in this strain. These results
provide insights into and a paradigm for how environmental conditions can affect
bacterial genome expansion.

<>

<1>Han, K., Tano, H., Kasai, K.
<2>Immobilization on polymeric particles of nucleic acids exhibiting restriction endonuclease-recognition site and its use for nucleic acid and protein purification.
<3>Japanese Patent Office
<4>JP 9600296 A
<5>
<6>1996
<7>
<>

<1>Han, X., Li, M., Ding, Z., Zhao, J., Ji, K., Wen, M., Lu, T.
<2>Genome Sequence of Streptomyces auratus Strain AGR0001, a Phoslactomycin-Producing Actinomycete.
<3>J. Bacteriol.
<4>194
<5>5472-5473
<6>2012
<7>Streptomyces auratus strain AGR0001 produces neophoslactomycin A, a novel analog  of
phoslactomycin that possesses potent activity against some phytopathogenic
fungi. Here, the draft genome sequence of S. auratus strain AGR0001 is presented,
which would provide insight into the biosynthetic mechanism of neophoslactomycin
A.

<>

<1>Hanahan, D.
<2>Mechanisms of DNA Transformation:  Restrictionlike effects in Transformation in Escherichia coli and Salmonella typhimurium.
<3>Escherichia coli and Salmonella typhimurium: Cellular and molecular biology., American Society for Microbiology, Neidhardt, F.C., Washington D.C.
<4>0
<5>1-7
<6>1986
<7>The introduction of naked DNA into Escherichia coli was first demonstrated by
Mandel and Higa, who observed that incubation of a suspension of E. coli  cells
and bacteriophage lambda DNA in a solution of CaCl2 at 0C resulted in the
subsequent appearance of infectious centers.  They further showed that a heat
pulse, in which the mixture of cells and DNA was briefly incubated at 42C,
chilled on ice, and then diluted into growth medium, improved the frequency of
transfection.  The general applicability of these conditions to DNA transfer
was demonstrated by their use to effect plasmid transformation, in which
circular plasmids carrying antibiotic resistance genes were stably established
as replicating episomes; genetic transformation, in which linear E. coli DNA
was transformed into auxotrophic strains to restore the mutant alleles; and
transfection of other bacteriophages.  These observations proved to be
applicable to DNA transformation of Salmonella typhimurium, suggesting that
induction of the artificial or natural ability to take up DNA reflected general
strutural or physiological features of these two closely related organisms.
Most subsequent studies of DNA transformation have employed plasmids carrying
antibiotic resistance genes, scoring transformation by the appearance of
drug-resistant colonies under selective conditions.  Primary attention has been
focused on E. coli, but it appears the observations are, in general, applicable
to S. typhimurium as well.  Transformation with linear DNAs generally shows the
same response to conditions, given the genetic distinction that linear DNA
(chromosomal or bacteriophage) will most efficiently transform strains
deficient in recBC nuclease, due to its degradative activity on the ends of DNA
molecules.

<>

<1>Hanak, A.M., Nagler, M., Weinmaier, T., Sun, X., Fragner, L., Schwab, C., Rattei, T., Ulrich, K., Ewald, D., Engel, M., Schloter, M., Bittner, R., Schleper, C., Weckwerth, W.
<2>Draft Genome Sequence of the Growth-Promoting Endophyte Paenibacillus sp. P22, Isolated from Populus.
<3>Genome Announcements
<4>2
<5>e00276-14
<6>2014
<7>Paenibacillus sp. P22 is a Gram-negative facultative anaerobic endospore-forming  bacterium
isolated from poplar hybrid 741 (female symbol[Populus alba x (P. davidiana + P. simonii) x P.
tomentosa]). This bacterium shows strong similarities to Paenibacillus humicus, and important
growth-promoting effects on in vitro grown explants of poplar hybrid 741 have been described.

<>

<1>Hanawa, F., Okamoto, M., Towers, G.H.N.
<2>Inhibition of restriction enzyme's DNA sequence recognition by PUVA treatment.
<3>Photochem. Photobiol.
<4>74
<5>269-273
<6>2001
<7>Applying various restriction enzymes on a specially designed 1.5 kb DNA fragment revealed that
the inhibitory effects of psoralens + UVA
irradiation (PUVA) treatment on restriction endonuclease activities are
caused by recognition inhibition. In this study restriction enzymes
that have a 5'-TpA sequence at the cleaving site (KpnI, XbaI, PmeI and
DraI), and the noncleaving site (PacI) in recognition sites, or have
two 5'-TpA sequences at the recognition site, and a nonspecific
sequence between the recognition and the cleaving sites (BciVI), were
inhibited by PUVA treatment. Most of the other restriction enzymes used
in this study, which do not have a 5'-TpA sequence at their restriction
site, were not inhibited by PUVA treatment, although a 5'-TpA sequence
is located adjacent (SmaI) or very close (BamHI, SacI and PstI) to the
recognition and cleaving sites for these enzymes. Because SphI, which
does not have 5'-TpA at its restriction site, was strongly inhibited by
PUVA treatment, the 5'-CpA sequence is suggested to be a new binding
site of psoralens after UVA irradiation.

<>

<1>Hanck, T., Gerwin, N., Fritz, H.-J.
<2>Nucleotide sequence of the dcm locus of Escherichia coli K12.
<3>Nucleic Acids Res.
<4>17
<5>5844
<6>1989
<7>None

<>

<1>Hanck, T., Schmidt, S., Fritz, H.J.
<2>Sequence-specific and mechanism-based crosslinking of Dcm DNA cytosine-C5 methyltransferase of E.coli K-12 to synthetic oligonucleotides containing 5-fluoro-2'-deoxycytidine.
<3>Nucleic Acids Res.
<4>21
<5>303-309
<6>1993
<7>The product of the dcm gene is the only DNA cytosine-C5 methyltransferase of Escherichia coli
K-12; it catalyses transfer of a methyl group from S-adenosyl methionine (SAM) to the C-5
position of the inner cytosine residue of the cognate sequence CCA/TGG. Sequence-specific,
covalent crosslinking of the enzyme to synthetic oligonucleotides containing
5-fluoro-2'-deoxycytidine is demonstrated. This reaction is abolished if serine replaces the
cysteine at residue #177 of the enzyme. These results lend strong support to a catalytic
mechanism in which an enzyme sulfhydryl group undergoes Michael addition to the C5-C6 double
bond, thus activating position C-5 of the substrate DNA cytosine residue for electrophilic
attack by the methyl donor SAM. The enzyme is capable of self-methylation in a DNA-independent
reaction requiring SAM and the presence of cysteine at position #177.

<>

<1>Hancox, E.L., Connolly, B.A., Walker, R.T.
<2>Synthesis and properties of oligodeoxynucleotides containing the analogue 2'-deoxy-4'-thiothymidine.
<3>Nucleic Acids Res.
<4>21
<5>3485-3491
<6>1993
<7>The 2'-deoxythymidine analogue 2'-deoxy-4'thiothymidine has been incorporated, using
standard methodology, into a series of dodecadeoxynucleotides containing the EcoRV restriction
endonuclease recognition site (GATATC). The stability of these oligodeoxynucleotides and their
ability to act as substrates for the restriction endonuclease and associated methylase have
been compared with a normal unmodified oligodeoxynucleotide. No problems were encountered in
the synthesis despite the presence of a potentially oxidisable sulfur atom in the sugar ring.
The analogue had very little effect on the melting temperature of the self-complementary
oligodeoxynucleotides so synthesised and all had a CD spectrum compatible with a B-DNA
structure. The oligodeoxynucleotide containing one analogue in each strand within the
recognition site, adjacent to the bond to be cleaved (i.e. GAXATC, where X is
2'-deoxy-4'-thiothymidine), was neither a substrate for the endonuclease nor was recognized
by the associated methylase. When still within the recognition hexanucleotide but two further
residues removed from the site of cleavage (i.e. GATAXC), the oligodeoxynucleotide was a poor
substrate for both the endonuclease and methylase. Binding of the oligodeoxynucleotide to the
endonuclease was unaffected but the kcat value was only 0.03% of the value obtained for the
parent oligodeoxynucleotide. These results show that the incorporation of
2'-deoxy-4'-thionucleosides into synthetic oligodeoxynucleotides may shed light on subtle
interations between proteins and their normal substrates and may also show why
2'-deoxy-4'-thiothymidine itself is so toxic in cell culture.

<>

<1>Hancox, E.L., Halford, S.E.
<2>Kinetic analysis of a mutational hot spot in the EcoRV restriction endonuclease.
<3>Biochemistry
<4>36
<5>7577-7585
<6>1997
<7>The EcoRV endonuclease contacts the minor groove of DNA through a peptide loop encompassing
residues 67-72.  This loop adapts to distorted DNA in the specific complex and to regular DNA
in the nonspecific complex.  Random mutagenesis had previously identified glutamine 69 as the
key component of the loop and this study reports on mutants with glutamate (Q69E), lysine
(Q69K), or leucine (Q69L) at this position.  The mutants bound DNA specifically at the EcoRV
recognition site in the presence of Ca2+, in the same manner as wild-type EcoRV.  In the
absence of divalent metals, Q69K and Q69L showed the same nonspecific binding as native EcoRV
while Q69E failed to bind DNA.  Glutamate at position 69 presumably repels nonspecific DNA
whilst allowing the adaptations to specific DNA.  Both Q69E and Q69K had severely impaired DNA
cleavage activities, while Q69L had a steady-state kcat within an order of magnitude of
wild-type EcoRV though its primary product was nicked DNA, in contrast to double strand breaks
by wild-type EcoRV.  The activity of Q69L required higher concentrations of Mg2+ than the
wild-type and showed a sigmoidal dependence upon the Mg2+ concentration, indicating two metal
ions per strand scission.  Transient kinetics on Q69L gave lower rate constants for
phosphodiester hydrolysis than wild-type EcoRV and its reaction also involved a slow
conformational change preceding DNA cleavage that had no equivalent with the wild-type.  Gln69
in EcoRV thus plays key roles in the adjustments of the protein to varied DNA structures and
in the alignment of the catalytic functions for DNA cleavage.

<>

<1>Handa, N., Ichige, A., Kobayashi, I.
<2>Contribution of RecFOR machinery of homologous recombination to cell survival after loss of a restriction-modification gene complex.
<3>Microbiology
<4>155
<5>2320-2332
<6>2009
<7>Loss of a type II restriction-modification gene complex, such as EcoRI, from a bacterial cell
leads to death of its descendant cells through
attack by residual restriction enzyme molecules on under-methylated target
sites of newly synthesized chromosomes. Through such post-segregational
host killing, these gene complexes force their maintenance on their host
cells. This finding led to re-discovery of type II
restriction-modification systems as selfish mobile elements. The host
prokaryote cells were found to cope with such attacks through a variety of
means. RecBCD pathway of homologous recombination in Escherichia coli
repairs the lethal lesions on the chromosome while it destroys restricted
non-self DNA. The recBCD homologs, however, appears very limited in
distribution among bacterial genomes, while homologs of RecFOR proteins
responsible for another pathway are widespread in eubacteria, just as the
restriction-modification systems are. In the present work, therefore, we
examined possible contribution of RecFOR pathway in cell survival after
loss of a restriction-modification gene complex. A recF mutation reduced
the survival in otherwise rec-positive background and, more severely, in a
recBC sbcBC background. We also found that its effect is prominent in the
presence of specific non-null mutant forms of RecBCD enzyme: the
resistance to killing seen with recC1002, recC1004, recC2145 and recB2154
is much reduced to the level of a null recBC allele when combined with a
recF, recO or recR mutant allele. Such resistance was also dependent on
RecJ and RecQ functions. UV resistance of these non-null recBCD mutants is
also decreased by recF, recJ or recQ mutation. These results demonstrate
that RecFOR pathway of recombination can greatly contribute to resistance
to restriction-modification-mediated host killing depending on genetic
backgrounds.

<>

<1>Handa, N., Ichige, A., Kusano, K., Kobayashi, I.
<2>Cellular responses to postsegregational killing by restriction-modification genes.
<3>J. Bacteriol.
<4>182
<5>2218-2229
<6>2000
<7>Plasmids that carry one of several type II restriction modification gene complexes are known
to show increased stability. The underlying mechanism was proposed to be the lethal attack by
a restriction enzyme at chromosomal recognition sites in cells that had lost the restriction
modification gene complex. In order to examine bacterial responses to this postsegregational
cell killing, we analyzed the cellular processes following loss of the EcoRI restriction
modification gene complex carried by a temperature-sensitive plasmid in an Escherichia coli
strain that is wild type with respect to DNA repair. A shift to the nonpermissive temperature
blocked plasmid replication, reduced the increase in viable cell counts and resulted in loss
of cell viability. Many cells formed long filaments, some of which were multinucleated and
others anucleated. In a mutant defective in RecBCD exonuclease/recombinase, these cell death
symptoms were more severe and cleaved chromosomes accumulated. Growth inhibition was also more
severe in recA, ruvAB, ruvC, recG, and recN mutants. The cells induced the SOS response in a
RecBC-dependent manner. These observations strongly suggest that bacterial cells die as a
result of chromosome cleavage after loss of a restriction modification gene complex and that
the bacterial RecBCD/RecA machinery helps the cells to survive, at least to some extent, by
repairing the cleaved chromosomes. These and previous results have led us to hypothesize that
the RecBCD/Chi/RecA system serves to destroy restricted "nonself" DNA and repair restricted
"self" DNA.

<>

<1>Handa, N., Kobayashi, I.
<2>Post-segregational killing by restriction modification gene complexes: Observations of individual cell deaths.
<3>Biochimie
<4>81
<5>931-938
<6>1999
<7>Through a mechanism known as post-segregational killing, several plasmids mediate their stable
maintenance by carrying genes that kill plasmid-free segregant cells. We demonstrated earlier
that loss of plasmids carrying type II restriction modification (RM) gene complexes inhibits
the propagation of a cell population and causes chromosome breakage. We now show the
morphology of individual cells changes following loss of thermosensitive plasmids carrying
EcoRI RM or PaeR7I RM after a shift to a non-permissive temperature. After a lag, many cells
formed long filaments containing multiple nuclei as detected by DAPI staining. Several hours
after the shift, many of these long filaments lacked nuclei. Fragmentation of chromosomal DNA
down to 5 kb was detected by electrophoresis. These observations lend strong support to the
concept of post-segregational cell killing by type II restriction modification gene complexes.

<>

<1>Handa, N., Kobayashi, I.
<2>Type III restriction is alleviated by bacteriophage (RecE) homologous recombination function but enhanced by bacterial (RecBCD) function.
<3>J. Bacteriol.
<4>187
<5>7362-7373
<6>2005
<7>Previous works have demonstrated that DNA breaks generated by restriction enzymes stimulate,
and are repaired by, homologous recombination with an intact, homologous DNA region through
the function of lambdoid bacteriophages lambda and Rac. In the present work, we examined the
effect of bacteriophage functions, expressed in bacterial cells, on restriction of an
infecting tester phage in a simple plaque formation assay. The efficiency of plaque formation
on an Escherichia coli host carrying EcoRI, a type II restriction system, is not increased by
the presence of Rac prophage-presumably because, under the single-infection conditions of the
plaque assay, a broken phage DNA cannot find a homologue with which to recombine. To our
surprise, however, we found that the efficiency of plaque formation in the presence of a type
III restriction system, EcoP1 or EcoP15, is increased by the bacteriophage-mediated homologous
recombination functions recE and recT of Rac prophage. This type III restriction alleviation
does not depend on lar on Rac, unlike type I restriction alleviation. On the other hand,
bacterial RecBCD-homologous recombination function enhances type III restriction. These
results led us to hypothesize that the action of type III restriction enzymes takes place on
replicated or replicating DNA in vivo and leaves daughter DNAs with breaks at nonallelic
sites, that bacteriophage-mediated homologous recombination reconstitutes an intact DNA from
them, and that RecBCD exonuclease blocks this repair by degradation from the restriction
breaks.

<>

<1>Handa, N., Naito, Y., Kobayashi, I.
<2>Experimental genome evolution: Large-scale genome rearrangements associated with resistance of a chromosomal restriction-modification gene complex to replacement.
<3>Genes Genet. Syst.
<4>75
<5>381
<6>2000
<7>Type II restriction enzymes are paired with modification enzymes that protect type II
restriction sites from cleavage by methylating them.  A plasmid carrying a type II restriction
modification gene complex is not easily replaced by an incompatible plasmid because loss of
the former leads to cell death through chromosome cleavage.  In the present work, we looked to
see if a chromosomally located restriction modification gene complex could be replaced by a
homologous stretch of DNA.  We tried to replace the PaeR7I gene complex on the Escherichia
coli chromosome by transducing a homologous stretch of PaeR7I-modified DNA.  The replacement
efficiency of the restriction modification complex was lower than expected.  Some of the
resulting recombinant clones retained the recipient restriction modification gene complex as
well as the homologous DNA (donor allele), and slowly lost the donor allele in the absence of
selection.  Analysis of their genome-wide rearrangements by Southern hybridization, inverse
PCR, and sequence determination demonstrated the occurrence of unequal homologous
recombination between copies of the transposon IS3.  It was strongly suggested that multiple
rounds of unequal IS3-IS3 recombination caused large-scale duplication and inversion of the
chromosome, and that only one of the duplicated copies of the recipient PaeR7I was replaced.

<>

<1>Handa, N., Nakayama, Y., Sadykov, M., Kobayashi, I.
<2>Experimental genome evolution: large-scale genome rearrangements associated with resistance to replacement of a chromosomal restriction-modification gene complex.
<3>Mol. Microbiol.
<4>40
<5>932-940
<6>2001
<7>Type II restriction enzymes are paired with modification enzymes that protect type II
restriction sites from cleavage by methylating them. A plasmid carrying a type II
restriction-modification gene complex is not easily replaced by an incompatible plasmid
because loss of the former leads to cell death through chromosome cleavage. In the present
work, we looked to see whether a chromosomally located restriction-modification gene complex
could be replaced by a homologous stretch of DNA. We tried to replace the PaeR7I gene complex
on the Escherichia coli chromosome by transducing a homologous stretch of PaeR7I-modified DNA.
The replacement efficiency of the restriction-modification complex was lower than expected.
Some of the resulting recombinant clones retained the recipient restriction-modification gene
complex as well as the homologous DNA (donor allele), and slowly lost the donor allele in the
absence of selection. Analysis of their genome-wide rearrangements by Southern hybridization,
inverse polymerase chain reaction (iPCR) and sequence determination demonstrated the
occurrence of unequal homologous recombination between copies of the transposon IS3. It was
strongly suggested that multiple rounds of unequal IS3-IS3 recombination caused large-scale
duplication and inversion of the chromosome, and that only one of the duplicated copies of the
recipient PaeR7I was replaced.

<>

<1>Handa, V., Jeltsch, A.
<2>Profound flanking sequence preference of Dnmt3a and Dnmt3b mammalian DNA methyltransferases shape the human epigenome.
<3>J. Mol. Biol.
<4>348
<5>1103-1112
<6>2005
<7>Mammalian DNA methyltransferases methylate cytosine residues within CG dinucleotides. By
statistical analysis of published data of the Human
Epigenome Project we have determined flanking sequences of up to
+/- four base-pairs surrounding the central CG site that are
characteristic of high (5'-CTTGCGCAAG-3') and low (5'-TGTTCGGTGG-3')
levels of methylation in human genomic DNA. We have investigated the
influence of flanking sequence on the catalytic activity of the Dnmt3a
and Dnmt3b de novo DNA methyltransferases using a set of synthetic
oligonucleotide substrates that covers all possible +/- 1 flanks
in quantitative terms. Methylation kinetics experiments revealed a >13-fold difference between
the preferred (RCGY) and disfavored +/-
1 flanking base-pairs (YCGR). In addition, AT-rich flanks are preferred
over GC-rich ones. These experimental preferences coincide with the
genomic methylation patterns. Therefore, we have expanded our
experimental analysis and found a >500-fold difference in the
methylation rates of the consensus sequences for high and low levels of
methylation in the genome. This result demonstrates a very pronounced
flanking sequence preference of Dnmt3a and Dnmt3b. It suggests that the
methylation pattern of human DNA is due, in part, to the flanking
sequence preferences of the de novo DNA MTases and that flanking
sequence preferences could be involved in the origin of CG islands.
Furthermore, similar flanking sequence preferences have been found for
the stimulation of the immune system by unmethylated CGs, suggesting a
co-evolution of DNA MTases and the immune system.

<>

<1>Handfield, M., Hillman, J.D., Progulske-Fox, A.
<2>Identification of Actinobacillus actinomycetemcomitans antigens for use in the diagnosis, treatment, and monitoring of periodontal diseases.
<3>US Patent Office
<4>US 7052860 B
<5>
<6>2006
<7>Antibodies, polypeptides, and polynucleotides are provided for the detection, prevention,
amelioration and treatment of diseases caused by Actinobacillus actinomycetemcomitans.

<>

<1>Handique, A.K.
<2>New technique for thermostability of restriction and modifying enzymes.
<3>Curr. Sci.
<4>66
<5>103-104
<6>1994
<7>Commentary on the use of trehalose to stabilize restriction enzymes

<>

<1>Hanish, J., McClelland, M.
<2>Activity of DNA modification and restriction enzymes in KGB, a potassium glutamate buffer.
<3>Gene Anal. Tech.
<4>5
<5>105-107
<6>1988
<7>The most abundant intracellular cation in bacteria is potassium, and the most abundant anion
is glutamate. However, most recommended restriction endonuclease buffers contain Na+ and Cl-.
Restriction endonucleases retain their ability to cleave DNA over a much broader range of
potassium glutamate (KGlu) concentrations than NaCl concentrations. These facts encouraged us
to investigate the possibility that we could use KGlu in an NaCl-free buffer and achieve
normal levels of activity for all restriction endonuclease. In this paper we present data
comparing the activity of 85 restriction endonucleases and 11 DNA methylases in a series of
KGlu buffers (KGC) against that found under optimal conditions recommended by the vendors (New
England Biolabs, Boehringer Mannheim Biochem., and International Biotech Inc.).

<>

<1>Hanish, J., McClelland, M.
<2>Controlled partial restriction digestions of DNA by competition with modification methyltransferases.
<3>Anal. Biochem.
<4>179
<5>357-360
<6>1989
<7>Competitive reactions, using defined ratios of DNA restriction
methyltransferase to endonuclease, are shown to result in reliable partial
restriction digests of DNA.  This method is suitable over a wide range of DNA
concentrations and works on DNA in liquid or embedded in agarose.  Simultaneous
methylase/endonuclease reactions using endonucleases that cleave human DNA very
infrequently, such as ClaI or NotI, should generate very large discrete DNA
fragments suitable for physical mapping in the million base-pair range.
Another possible application of methylase/endonuclease competitive reactions is
the production of defined partial digests for making cosmid, lambda, or other
genomic libraries.

<>

<1>Hanish, J., McClelland, M.
<2>Methylase-limited partial NotI cleavage for physical mapping of genomic DNA.
<3>Nucleic Acids Res.
<4>18
<5>3287-3291
<6>1990
<7>Partial cleavage of DNA with the restriction endonuclease NotI (5'...GC/GGCCGC...3') is an
important technique for genomic mapping. However, partial genomic cleavage with this enzyme is
impaired by the agarose matrix in which the DNA must be suspended. To solve this problem we
have purified the blocking methylase M.BspRI (5'...GGmCC...3') for competititon digests with
NotI. The resulting methylase-limited partial DNA cleavage is shown to be superior to standard
techniques on bacterial genomic DNA.

<>

<1>Hanish, J., McClelland, M.
<2>Enzymatic cleavage of a bacterial chromosome at a transposon-inserted rare site.
<3>Nucleic Acids Res.
<4>19
<5>829-832
<6>1991
<7>The sequential use of the methylase M.XbaI (5'-TCTAGm6A) and the methylation-dependent
endonuclease DpnI (5'-Gm6A/TC) results in cleavage at 5'-TCTAGA/TCTAGA. This recognition
sequence was introduced into a transposon derived from the Mu bacteriophage and transposed
into the genome of the bacterium Salmonella typhimurium. M.XbaI methylation was provided in
vivo by a plasmid containing the M.XbaI gene and the S. typhimurium genome was cleaved to
completion by DpnI at one or more sites, depending on the number of transposon insertions. The
resulting genomic fragments were resolved by pulsed-field electrophoresis. The potential use
of single M.XbaI/DpnI cleavage sites as reference positions to map rare restriction sites is
discussed.

<>

<1>Hanish, J., Rebelsky, M., McClelland, M., Westbrook, C.
<2>Application of methylase-limited partial NotI cleavage for a long-range restriction map of the human ABL locus.
<3>Genomics
<4>10
<5>681-685
<6>1991
<7>The use of partial restriction digests for mapping complex genomes by
pulsed-field gel electrophoresis has been limited by the difficulty of
consistently obtaining these digests in agarose, which is a necessary matrix
for high-molecular-weight DNA.  Enzyme cleavage in agarose is faster then
diffusion for most of the enzymes which cleave infrequently.  We have developed
a method for the production of partial digests in agarose for the endonuclease
NotI (5' ...GC/GGCCGC...3') which circumvents the diffusion problem by using
the blocking methylase M.BspRI (5' ...GGmCC...3'), which competes for the same
sites.  Using various ratios of the methylase and endonuclease results in
partial digests in any size range desired.  We report the successful
application of this technique to the production of NotI partial digests of
human genomic DNA for the mapping of the ABL locus of human chromosome 9.

<>

<1>Hanley, A.B., Furniss, C.S.M., Kwiatkowska, C.A., Mackie, A.R.
<2>The manipulation of DNA with restriction enzymes in low water systems.
<3>Biochim. Biophys. Acta
<4>1074
<5>40-44
<6>1991
<7>The cleavage of phage lambda DNA by the restriction enzyme HindIII in low water
systems has been investigated.  Two types of low water systems have been
studied - those which contain a surfactant in a reverse micelle environment and
a surfactant-free system in which a solid support (celite) is used.  The effect
of the surfactants themselves in a normal aqueous environment has also been
studied.  Charged surfactants were found to greatly inhibit HindIII activity in
aqueous buffer, while non-ionic surfactants did not affect either the activity
or the specificity of the restriction enzyme.  The rate of cleavage by HindIII
in a reverse micelle system consisting of sodium dioctylsulphosuccinate is very
slow, however, in a Triton B system the expected fragments are observed.  In a
surfactant-free low water environment, cleavage occurs at the expected sites
but in a different order to that observed in normal aqueous systems.  These
results sugest that DNA tertiary structure in low water systems is different to
that in aqueous solution and that this influences cleavage by the restriction
enzyme HindIII.

<>

<1>Hanley, A.B., Grinfeld, E., Baxter, R.L.
<2>The cleavage of nucleic acids in reversed micelles using site specific endonucleases.
<3>Biocatalysis
<4>3
<5>253-258
<6>1990
<7>Plasmid and lambda DNA molecules of between 2.2 and 48.5 kb pairs can be
solubilised in n-hexane containing the surfactant sodium dioctyl sulfosuccinate
(AOT) and aqueous buffers.  Linear lambda phage DNA fragments (2.2-23.1 kb
pairs) and intact lambda bio 1 DNA (48.5 kb pairs) are efficiently cleaved by
BamHI and EcoRI in systems containing 100 mM AOT.  Under these conditions,
lambda bio 1 DNA undergoes regioselective restriction by HindIII at only one
site but is completely cleaved when the surfactant concentration is lowered to
50 mM.  Covalent closed circular plasmid DNA (pUC8, 2.73 kb pairs) is only
partially linearised by EcoRI and BamHI in reversed micelles; HaeII cleavage
affords both complete and partial restriction fragments.  The results suggest
that the tertiary structures adopted by substrate DNA in reversed micelles
influence the availability of restriction sites.

<>

<1>Hansen, R.S., Wijmenga, C., D'Esposito, M., Weemaes, C.M.R., Gartier, S.M.
<2>Mutations in the DNMT3B DNA methyltransferase gene cause the ICF syndrome.
<3>Am. J. Hum. Genet.
<4>66
<5>1724
<6>2000
<7>Immunodeficiency, Centromeric instability and Facial anomalies are characteristics of a rare
genetic disorder termed the ICF syndrome.  The centromeric instability of chromosomes 1, 9,
and 16 is associated with the abnormal hypomethylation of their pericentromeric satellite
regions.  Hypomethylation has also been reported for other types of heterochromatin, including
the inactive X chromosome.  We examined these phenomena further at the molecular level and
report here examples of extensive hypomethylation of these regions in ICF cells that are
associated with nuclease hypersensitivity, advanced replication time and a variable escape
from silencing for genes on the inactive X and Y chromosomes.  The ICF locus has been mapped
to a 9 c-M region of chromosome 20 by homozygosity mapping.  By searching for homologies to
known DNA methyltransferases, we identified a genomic sequence located in the ICF region that
contains a full length homologue of the mouse DNMT3B methyltransferase gene.  The human
sequence was used to screen ICF kindreds and we discovered mutations in 4 patients from 3
families.  Restriction enzyme and bisulfite methylation analyses revealed extensive
hypomethylation of 5' CpG islands at all 10 genes examined on the inactive X and 1 gene on
the Y in two ICF females and 3 ICF males.  Abnormal hypomethylation in ICF was also associated
with advanced replication time for several loci examined, including satellite II sequences.
Consistent with these data, we discovered novel examples of escape from inactivation in
untreated diploid fibroblasts and lymphoblasts from ICF patients for G5PD, MPP1, and SYBL1.
This escape is variable, and appears to correlate with the degree of advanced replication time
for these loci.  The ICF phenotype, therefore, is likely to involve abnormalities in gene
silencing at multiple loci that arise from defects in methylation and late replication.  We
found that the satellite hypomethylation defect in ICF cells can be complemented by somatic
cell fusion to CHO cells.  We presume that this de novo methylation results from
complementation of the defective human ICF gene with a functional hamster homologue and we
sought to identify and localize DNA methyltransferases with de novo activity.  Dnmt3a and
Dnmt3b are two very homologous DNA methyltransferases with de novo activity that were recently
identified in the mouse.  Because the ICF locus maps to the proximal long arm of chromosome
20, we searched the unfinished chromosome 20 sequence data at the Sanger Centre for homology
to these genes and identified a PAC sequence containing a full length coding sequence that is
highly homologous to the murine Dnmt3b gene.  The predicted DNMT38 sequence was verified by
sequencing RT-PCR and PCR products amplified with derived primers.  We examined the DNMT3B
gene in ICF patients for mutations and found three ICF-specific mutations: a T to G
transversion resulting in a V726G missense substitution that is homozygous in the two affected
brothers of Family 1, a CpG to CpA missense transition (A603T) that is heterozygous in the P4
ICF female of Family 3, and an intronic CpG to CpA splice mutation resulting in an STP
insertion before codon 897 that is homozygous in the P3 patient of Family 2 and heterozygous
in the P4 patient of Family 3.  None of the mutations were present in over 200 normal
chromosomes examined from unrelated individuals.  All three DNMT3B mutations are predicted to
be in all major splice forms and occur in regions that are invariant among the DNMT3-like
methyltransferases that have been identified in zebra-fish, mouse and man and are in or near
regions of homology to certain bacteriophage methyltransferases.  These mutations all appear
to be in the catalytic domain of the enzyme as they are within or near motifs present in
nearly all m5C-methyltransferases.  Our observations of escape from X inactivation,
complementation for the ICF methylation defect in somatic cells, and of DNMT3B mutations in
ICF patients provide a useful model for examining the role of this enzyme in the establishment
and maintenance of somatic methylation patterns.  This is the first example of a mutation in a
human gene that alters DNA methylation patterns.

<>

<1>Hansen, R.S., Wijmenga, C., Luo, P., Stanek, A.M., Canfield, T.K., Weemaes, C.M., Gartler, S.M.
<2>The DNMT3B DNA methyltransferase gene is mutated in the ICF immunodeficiency syndrome.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>14412-14417
<6>1999
<7>DNA methylation is an important regulator of genetic information in species ranging from
bacteria to humans. DNA methylation appears to be critical for mammalian development because
mice nullizygous for a targeted disruption of the DNMT1 DNA methyltransferase die at an early
embryonic stage. No DNA methyltransferase mutations have been reported in humans until now. We
describe here the first example of naturally occurring mutations in a mammalian DNA
methyltransferase gene. These mutations occur in patients with a rare autosomal recessive
disorder, which is termed the ICF syndrome, for immunodeficiency, centromeric instability, and
facial anomalies. Centromeric instability of chromosomes 1, 9, and 16 is associated with
abnormal hypomethylation of CpG sites in their pericentromeric satellite regions. We are able
to complement this hypomethylation defect by somatic cell fusion to Chinese hamster ovary
cells, suggesting that the ICF gene is conserved in the hamster and promotes de novo
methylation. ICF has been localized to a 9-centimorgan region of chromosome 20 by homozygosity
mapping. By searching for homologies to known DNA methyltransferases, we identified a genomic
sequence in the ICF region that contains the homologue of the mouse Dnmt3b methyltransferase
gene. Using the human sequence to screen ICF kindreds, we discovered mutations in four
patients from three families. Mutations include two missense substitutions and a 3-aa
insertion resulting from the creation of a novel 3' splice acceptor. None of the mutations
were found in over 200 normal chromosomes. We conclude that mutations in the DNMT3B are
responsible for the ICF syndrome.

<>

<1>Hanson, D.K., Lamb, M.R., Mahler, H.R., Perlman, P.S.
<2>Evidence for translated intervening sequences in the mitochondrial genome of Saccharomyces cerevisiae.
<3>J. Biol. Chem.
<4>257
<5>3218-3224
<6>1982
<7>In yeast, the mitochondrial genes for subunit I of cytochrome oxidase (oxi3) and for
apocytochrome b (cob) are known to be split. In some strains, the latter contains five
intervening sequences, three of which coincide with clusters of mutational sites referred to
in their order of transcription as the loci box3, 10, and 7, respectively. Mutations at the
first of these result in the accumulation of novel, large polypeptides (apparent Mr = about
43,000) believed to originate from a fusion of sequences found in the NH2-terminal segment of
apocytochrome b to others encoded in the intervening sequence itself. We now provide evidence
for close similarities of at least a part of translated intron sequences between (a) mutants
in box7 in "long" form and "short" form strains (which lack the first three introns including
the one for the box3 locus); (b) mutants in a subset of box7 mutants and those in box3, and
(c) between intron sequences in box7 and a sequence presumably encoded in oxi3. These
structural homologies presumably encoded in oxi3. These structural homologies have been
analyzed and shown to be referable to sequence homologies in two proteins, one derived from
the second intron (box3) in cob and the other from oxi3. The accumulation in certain cob
mutants of proteins and of a transcript containing a sequence specified by oxi3 provides
additional strong evidence for the previously suggested regulation of oxi3 by the penultimate,
box7-containing intron of cob.

<>

<1>Hanvey, J.C., Shimizu, M., Wells, R.D.
<2>Site-specific inhibition of EcoRI restriction/modification enzymes by a DNA triple helix.
<3>Nucleic Acids Res.
<4>18
<5>157-161
<6>1990
<7>The abilility of oligopyrimidines to inhibit, through triple helix formation,
the specific protein-DNA interactions of the EcoRI restriction and modification
enzymes (EcoRI and M.EcoRI) with their recognition sequence (GAATTC) was
studied.  The oligonucleotides (CTT)4 and (CTT)8 formed triplexes in plasmids
at (GAA)n repeats containing EcoRI sites.  Cleavage and methylation of EcoRI
sites within these sequences were specifically inhibited by the
oligonucleotides, whereas an EcoRI site adjacent to a (GAA)n sequence was
inhibited much less.  Also, other EcoRI sites within the plasmid, or in
exogenously added lambda DNA, were not inhibited.  These results demonstrate
the potential of using triplex-forming oligonucleotides to block protein-DNA
interactions at specific sites, and thus this technique may be useful in
chromosome mapping and in the modulation of gene expression.

<>

<1>Hao, K., He, P., Blom, J., Rueckert, C., Mao, Z., Wu, Y., He, Y., Borriss, R.
<2>The Genome of Plant Growth-Promoting Bacillus amyloliquefaciens subsp. plantarum  Strain YAU B9601-Y2 Contains a Gene Cluster for Mersacidin Synthesis.
<3>J. Bacteriol.
<4>194
<5>3264-3265
<6>2012
<7>The genome of rhizobacterium Bacillus amyloliquefaciens subsp. plantarum YAU B9601-Y2 was 4.24
Mb in size and harbored 3,991 coding sequences (CDS). Giant
gene clusters were dedicated to nonribosomal synthesis of antimicrobial
lipopeptides and polyketides. Remarkably, CAU B946 possessed a gene cluster
involved in synthesis of mersacidin.

<>

<1>Hao, P., Zheng, H., Yu, Y., Ding, G., Gu, W., Chen, S., Yu, Z., Ren, S., Oda, M., Konno, T., Wang, S., Li, X., Ji, Z.-S., Zhao, G.
<2>Complete Sequencing and Pan-Genomic Analysis of Lactobacillus delbrueckii subsp bulgaricus Reveal Its Genetic Basis for Industrial Yogurt Production.
<3>PLoS ONE
<4>6
<5>e15964
<6>2011
<7>Lactobacillus delbrueckii subsp. bulgaricus (Lb. bulgaricus) is an important species of Lactic
Acid Bacteria (LAB) used for cheese and
yogurt fermentation. The genome of Lb. bulgaricus 2038, an industrial
strain mainly used for yogurt production, was completely sequenced and
compared against the other two ATCC collection strains of the same
subspecies. Specific physiological properties of strain 2038, such as
lysine biosynthesis, formate production, aspartate-related
carbon-skeleton intermediate metabolism, unique EPS synthesis and
efficient DNA restriction/modification systems, are all different from
those of the collection strains that might benefit the industrial
production of yogurt. Other common features shared by Lb. bulgaricus
strains, such as efficient protocooperation with Streptococcus
thermophilus and lactate production as well as well-equipped stress
tolerance mechanisms may account for it being selected originally for
yogurt fermentation industry. Multiple lines of evidence suggested that
Lb. bulgaricus 2038 was genetically closer to the common ancestor of
the subspecies than the other two sequenced collection strains,
probably due to a strict industrial maintenance process for strain 2038
that might have halted its genome decay and sustained a gene network
suitable for large scale yogurt production.

<>

<1>Hao, X., Lin, Y., Johnstone, L., Baltrus, D.A., Miller, S.J., Wei, G., Rensing, C.
<2>Draft Genome Sequence of Plant Growth-Promoting Rhizobium Mesorhizobium amorphae, Isolated from Zinc-Lead Mine Tailings.
<3>J. Bacteriol.
<4>194
<5>736-737
<6>2012
<7>Here, we describe the draft genome sequence of Mesorhizobium amorphae strain CCNWGS0123,
isolated from nodules of Robinia pseudoacacia growing
on zinc-lead mine tailings. A large number of metal(loid) resistance
genes, as well as genes reported to promote plant growth, were identified,
presenting a great future potential for aiding phytoremediation in
metal(loid)-contaminated soil.

<>

<1>Hao, X., Lin, Y., Johnstone, L., Liu, G., Wang, G., Wei, G., McDermott, T., Rensing, C.
<2>Genome Sequence of the Arsenite-Oxidizing Strain Agrobacterium tumefaciens 5A.
<3>J. Bacteriol.
<4>194
<5>903
<6>2012
<7>Microbial transformations of arsenic influence its mobility and toxicity. We report the draft
genome sequence of the arsenite-oxidizing strain
Agrobacterium tumefaciens 5A isolated from an As-contaminated soil in the
Madison River Valley, MT. A large number of metal (or metalloid)
resistance genes, especially contributing to arsenite oxidation, were
identified.

<>

<1>Hao, Y., Huang, D., Guo, H., Xiao, M., An, H., Zhao, L., Zuo, F., Zhang, B., Hu, S., Song, S., Chen, S., Ren, F.
<2>Complete Genome Sequence of Bifidobacterium longum subsp. longum BBMN68, a New Strain from Healthy Chinese Centenarian.
<3>J. Bacteriol.
<4>193
<5>787-788
<6>2010
<7>Bifidobacterium longum subsp. longum BBMN68 was isolated from the fecal of healthy centenarian
living in longevity area of BaMa, Guangxi, China.
Here, we reported the main genome features of B. longum BBMN68 strain and
the identification of several predicted proteins that tailored to the
ecological niche of longevity.

<>

<1>Hao, Z., Li, L., Liu, J., Ren, Y., Wang, L., Bartlam, M., Egli, T., Wang, Y.
<2>Genome Sequence of a Freshwater Low-Nucleic-Acid-Content Bacterium, Betaproteobacterium Strain CB.
<3>Genome Announcements
<4>1
<5>e00135-13
<6>2013
<7>Betaproteobacterium strain CB is a typical minute freshwater bacterium, representing the
small-cell bacteria that are numerically dominant in most
freshwater environments. The genome of betaproteobacterium CB consists of a
circular 2,045,720-bp chromosome, and the information we report will provide
insights into the mechanisms underlying its survival and ecological function.

<>

<1>Haq, I., O'Brien, R., Lagunavicius, A., Siksnys, V., Ladbury, J.E.
<2>Specific DNA Recognition by the Type II Restriction Endonuclease MunI: The Effect of pH.
<3>Biochemistry
<4>40
<5>14960-14967
<6>2001
<7>To investigate the effect of pH on sequence-specific binding, a thermodynamic characterization
of the interaction of the protein MunI with a specific, and a nonspecific, oligonucleotide was
performed. MunI is a type II restriction endonuclease which is able to bind specifically, but
loses its enzymatic activity in the absence of magnesium ions. Comparison of the specific and
nonspecific interactions at 10 and 25 degrees C shows that the latter is accompanied by a
small change in enthalpy, and a negligible change in constant pressure heat capacity. On going
through the pH range 5.75-9.0 at 25 degrees C, the affinity of specific complex formation is
reduced by 20-fold. The interaction is accompanied by the protonation of groups assumed to be
on the protein. Based on the simplest model that will fit the data, two distinct protonation
events are observed. At low pH, two groups per protein molecule undergo protonation with a
pK(a) of 6.0 and 6.9 in the free and bound forms, respectively. At high pH, a further
independent protonation occurs involving two groups with pK(a) values of 8.9 and approximately
10.7 in the free and bound forms, respectively. The change in heat capacity ranges from -2.7
to -1.7 kJ mol(-)(1) K(-)(1) in going from pH 6.5 to 8.5. This range of variation of change in
heat capacity can be accounted for by the effects of protonation of the interacting molecules.
The change in heat capacity, calculated from surface area burial using a previously
established relationship (1.15 kJ mol(-)(1) K(-)(1)), does not correlate well with the
experimentally determined values.

<>

<1>Haqqi, T.M., Ahmad, S., Ahmad, N.S., Ahmad, M., Hasnain, A., Siddiqi, M., Hadi, S.M.
<2>Cloning and expression of EcoRI specific restriction modification system.
<3>Indian J. Biochem. Biophys.
<4>22
<5>252-254
<6>1985
<7>RI-specific restriction-modification genes have been cloned in the multicopy
plasmid pBR322.  Plasmid DNA, isolated from E. coli RY13, was cleaved with
restriction enzyme BamHI and inserted at the BamHI site in pBR322 and cloned in
E. coli HB101.  The recombinant DNA transnformants were selected by their
resistance to ampicillin and sensitivity to tetracycline.  EcoRI-specific
clones were identified by determining the efficiency of plating on
transformants of modified and unmodified lambda phage.  Crude enzyme prepared
from several of the transformants by dextran polyethylene glycol phase
partition procedure yielded the same restriction pattern as the parental strain
and purified EcoRI.

<>

<1>Hardwick, J.M., von Sprecken, R.S., Yielding, K.L., Yielding, L.W.
<2>Ethidium binding sites on plasmid DNA determined by photoaffinity labeling.
<3>J. Biol. Chem.
<4>259
<5>11090-11097
<6>1984
<7>Photoaffinity labeling of pBR322 with ethidium monoazide
(8-azido-3-amino-5-ethyl-6-phenylphenanthridinium chloride) was used to provide evidence for
the sequence specifity of ethidium binding to native DNA. DNA-drug interactions were examined
at concentrations of eight covalently bound ethidium drugs per molecule of pBR322 (4363 base
pairs). Restriction enzyme cutting was blocked by the covalent binding of a drug molecule at
(or near) the enzyme recognition sequence. This phenomenon was observed with all restriction
enzymes tested and was not limited to specific regions of the pBR322 molecule.
Double-digestion experiments indicated that a drug molecule may bind 2 to 3 base pairs outside
the recognition sequence and still block restriction enzyme digestion. Intact plasmid was
treated with [3H] ethidium monoazide and digested with restriction enzymes. The amount of
covalently-linked ethidium analog was quantitated for different restriction fragments and the
G-C content of each fragment was determined from the DNA sequence. In approximately half of
the fragments the drug appeared to preferentially bind at a G-C base pair. However, a
preference for specific sequences such as 5'-C-G-3' was detected, as had been suggested by
previous modeling studies with ethidium bromide. The other fragments were located in specific
map regions of the plasmid and did not bind drug with a strict dependence on GC content
suggesting that binding specificity may depend on more than one structural feature of the DNA.

<>

<1>Harhay, G.P., Koren, S., Phillippy, A.M., McVey, D.S., Kuszak, J., Clawson, M.L., Harhay, D.M., Heaton, M.P., Chitko-McKown, C.G., Smith, T.P.
<2>Complete Closed Genome Sequences of Mannheimia haemolytica Serotypes A1 and A6, Isolated from Cattle.
<3>Genome Announcements
<4>1
<5>e00188-13
<6>2013
<7>Mannheimia haemolytica is a respiratory pathogen affecting cattle and related ruminants
worldwide. M. haemolytica is commonly associated with bovine
respiratory disease complex (BRDC), a polymicrobial multifactorial disease. We
present the first two complete closed genome sequences of this species,
determined using an automated assembly pipeline requiring no manual finishing.

<>

<1>Harhay, G.P., McVey, D.S., Koren, S., Phillippy, A.M., Bono, J., Harhay, D.M., Clawson, M.L., Heaton, M.P., Chitko-McKown, C.G., Korlach, J., Smith, T.P.
<2>Complete Closed Genome Sequences of Three Bibersteinia trehalosi Nasopharyngeal Isolates from Cattle with Shipping Fever.
<3>Genome Announcements
<4>2
<5>e00084-14
<6>2014
<7>Bibersteinia trehalosi is a respiratory pathogen affecting cattle and related ruminants
worldwide. B. trehalosi is closely related to Mannheimia haemolytica
and is often associated with bovine respiratory disease complex (BRDC), a
polymicrobial multifactorial disease. We present three complete closed genome
sequences of this species generated using an automated assembly pipeline.

<>

<1>Harhay, G.P., Murray, R.W., Lubbers, B., Griffin, D., Koren, S., Phillippy, A.M., Harhay, D.M., Bono, J., Clawson, M.L., Heaton, M.P., Chitko-McKown, C.G., Smith, T.P.
<2>Complete Closed Genome Sequences of Four Mannheimia varigena Isolates from Cattle with Shipping Fever.
<3>Genome Announcements
<4>2
<5>e00088-14
<6>2014
<7>Mannheimia varigena is an occasional respiratory pathogen of cattle and pigs. We  present the
first four complete closed genome sequences of this species.

<>

<1>Harjes, J., Ryu, T., Abdelmohsen, U.R., Moitinho-Silva, L., Horn, H., Ravasi, T., Hentschel, U.
<2>Draft Genome Sequence of the Antitrypanosomally Active Sponge-Associated Bacterium Actinokineospora sp. Strain EG49.
<3>Genome Announcements
<4>2
<5>e00160-14
<6>2014
<7>The marine sponge-associated bacterium Actinokineospora sp. strain EG49 produces  the
antitrypanosomal angucycline-like compound actinosporin A. The draft genome
of Actinokineospora sp. EG49 has a size of 7.5 megabases and a GC content of
72.8% and contains 6,629 protein-coding sequences (CDS). antiSMASH predicted 996
genes residing in 36 secondary metabolite gene clusters.

<>

<1>Harmon-Smith, M. et al.
<2>Complete genome sequence of Sebaldella termitidis type strain (NCTC 11300).
<3>Standards in Genomic Sciences
<4>2
<5>220-227
<6>2010
<7>Sebaldella termitidis (Sebald 1962) Collins and Shah 1986, is the only species in the genus
Sebaldella within the fusobacterial family 'Leptotrichiaceae'. The sole
and type strain of the species was first isolated about 50 years ago from
intestinal content of Mediterranean termites. The species is of interest for its
very isolated phylogenetic position within the phylum Fusobacteria in the tree of
life, with no other species sharing more than 90% 16S rRNA sequence similarity.
The 4,486,650 bp long genome with its 4,210 protein-coding and 54 RNA genes is
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Harper, S.H., Firman, K., Glover, S.W.
<2>The interactions of mini-F with the plasmids R124 and R124/3.
<3>Plasmids in Bacteria, Plenum Press, Helsinki, D.R., Cohen, S.N., Clewell, D.B., Jackson, D.A., Hollaender, A., New York
<4>0
<5>920
<6>1984
<7>The Inc FIV plasmid R124 encodes a unique restriction and modification (R-M) system.  R124/3
is a derivative plasmid which encodes an R-M system of different specificity.  When R124 or
R124/3 are present in the same cell as the sex Factor F a restriction deficient (r-) phenotype
results.  This r- phenotype reverts back to r+ with a frequency dependent upon the particular
isolate.  When the F plasmids are isolated from these reverted strains they are found to
encode the R-M system of the donor R124 or R124/3 plasmid.  The R-M system is translocated
into the region of EcoRI fragments 5 and 7 of F, which encodes the Phi and Ori VI functions of
F.  During this transfer of DNA some F DNA is lost.  Recent work with mini-F (cloned EcoRI
fragments 5 and 7 of F) have shown that they alone do not result in the translocation of R-M
genes from R124 or R124/3.  EcoRI fragments 5 and 7 of F when present in the same cell as R124
or R124/3 result in a fraction of the cells expressing a r- phenotype which reverts back to a
r+ phenotype.  This reversion process is not accompanied by any DNA transfer or rearrangement
between F, R124, or R124/3 plasmids.  This strongly suggests that some other region of F is
involved in the DNA rearrangement.  Studies with mini-F have also shown that under certain
circumstances the rearrangement results in R124 expressing incF1 incompatability instead of
the normal Inc FIV.

<>

<1>Harrington, A., Hill, C.
<2>Plasmid involvement in the formation of a spontaneous bacteriophage insensitive mutant of Lactococcus lactis.
<3>FEMS Microbiol. Lett.
<4>96
<5>135-141
<6>1992
<7>Lactococcus lactis subsp. lactis biovar. diacetylactis DPC721 is a spontaneous bacteriophage
insensitive mutant of stain DPC220, isolated after challenge with an industrial bacteriophage,
phiD1. Plasmid analysis demonstrated that the bacteriophage insensitivity was associated with
the absence of two native DPC220 plasmids (pAH82 and pAH33), and the presence of a novel
plasmid (pAH90) in DPC721. The plasmids were transferred by conjugative mobilization to a
plasmid free background where it was confirmed by restriction mapping that pAH90 is a
co-integrate formed by the precise recombination of pAH82 and pAH33. The resistance phenotype
encoded by pAH90 was also active against two bacteriophages homologous for the plasmid-free
strain. Plasmid pAH90 was shown to encode at least two independent resistance mechanisms,
including an adsorption-inhibition mechanism and a restriction and modification system. The
adsorption-inhibition mechanism encoded by the co-integrate plasmid was specific for one of
the phage used in this study.

<>

<1>Harrington, R., Ondov, B.D., Radune, D., Friss, M.B., Klubnik, J., Diviak, L., Hnath, J., Cendrowski, S.R., Blank, T.E., Karaolis, D., Friedlander, A.M., Burans, J.P., Rosovitz, M.J., Treangen, T., Phillippy, A.M., Bergman, N.H.
<2>Genome Sequence of the Attenuated Carbosap Vaccine Strain of Bacillus anthracis.
<3>Genome Announcements
<4>1
<5>e00067-12
<6>2013
<7>The Bacillus anthracis Carbosap genome, which includes the pXO1 and pXO2 plasmids, has been
shown to encode the major B. anthracis virulence factors, yet this strain's attenuation has
not yet been explained. Here we report the draft genome sequence of this strain, and a
comparison to fully virulent B. anthracis.

<>

<1>Harris, A.P., Techtmann, S.M., Stelling, S.C., Utturkar, S.M., Alshibli, N.K., Brown, S.D., Hazen, T.C.
<2>Draft Genome Sequence of Pseudoalteromonas sp. Strain ND6B, an Oil-Degrading Isolate from Eastern Mediterranean Sea Water Collected at a Depth of 1,210  Meters.
<3>Genome Announcements
<4>2
<5>e01212-14
<6>2014
<7>Here, we report the draft genome of Pseudoalteromonas sp. strain ND6B, which is able to grow
with crude oil as a carbon source. Strain ND6B was isolated from
eastern Mediterranean Sea deep water at a depth of 1,210 m. The genome of strain
ND6B provides insight into the oil-degrading ability of the Pseudoalteromonas
species.

<>

<1>Harris, V.H., Hamilton, A., Williams, D.M., Hornby, D.P.
<2>Studying the function of methyltransferases using nucleotide analogue based random mutagenesis.
<3>Collection Symposium Series
<4>2
<5>209-212
<6>1999
<7>The function of two methyltransferases M.SPR and M.HhaI have been investigated using a PCR
based random mutagenesis procedure using nucleotide analogues with ambiguous base pairing
properties.  The mutagenesis procedure was compared with traditional protocols in which
manganese ions are introduced to decrease the fidelity of the DNA polymerase.

<>

<1>Harris-Warrick, R.M., Elkana, Y., Ehrlich, S.D., Lederberg, J.
<2>Electrophoretic separation of Bacillus subtilis genes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>72
<5>2207-2211
<6>1975
<7>The cleavage of Bacillus subtilis DNA by EcoRI restriction endonuclease
produces segments which retain various degrees of genetic transforming
activity.  The active segments analyzed thus far, range in size from 23 to 3
kilobases and can be partially separated by agarose gel electrophoresis.
Various markers can thus be enriched from 30- to 60-fold.

<>

<1>Harrison, A., Dyer, D.W., Gillaspy, A., Ray, W.C., Mungur, R., Carson, M.B., Zhong, H., Gipson, J., Gipson, M., Johnson, L.S., Lewis, L., Bakaletz, L.O., Munson, R.S. Jr.
<2>Genomic sequence of an otitis media isolate of nontypeable Haemophilus influenzae: Comparative study with H. influenzae serotype d, strain KW20.
<3>J. Bacteriol.
<4>187
<5>4627-4636
<6>2005
<7>In 1995, the Institute for Genomic Research completed the genome sequence of a rough
derivative of Haemophilus influenzae serotype d, strain KW20. Although extremely useful in
understanding the basic biology of H. influenzae, these data have not provided significant
insight into disease caused by nontypeable H. influenzae, as serotype d strains are not
pathogens. In contrast, strains of nontypeable H. influenzae are the primary pathogens of
chronic and recurrent otitis media in children. In addition, these organisms have an important
role in acute otitis media in children as well as other respiratory diseases. Such strains
must therefore contain a gene repertoire that differs from that of strain Rd. Elucidation of
the differences between these genomes will thus provide insight into the pathogenic mechanisms
of nontypeable H. influenzae. The genome of a representative nontypeable H. influenzae strain,
86-028NP, isolated from a patient with chronic otitis media was therefore sequenced and
annotated. Despite large regions of synteny with the strain Rd genome, there are large
rearrangements in strain 86-028NP's genome architecture relative to the strain Rd genome. A
genomic island similar to an island originally identified in H. influenzae type b is present
in the strain 86-028NP genome, while the mu-like phage present in the strain Rd genome is
absent from the strain 86-028NP genome. Two hundred eighty open reading frames were identified
in the strain 86-028NP genome that were absent from the strain Rd genome. These data provide
new insight that complements and extends the ongoing analysis of nontypeable H. influenzae
virulence determinants.

<>

<1>Harrison, J., Addison, G., Ramsden, N., Drewes, G.
<2>Capturing and identification of methyltransferase and identification of methyltransferase interacting molecules using affinity matrix.
<3>International Patent Office
<4>WO 201216704 A
<5>
<6>2012
<7>The present invention relates to immobilization compounds, immobilization products and
preparations thereof as well as methods and uses for the identification of methyltransferase
interacting compounds or for the purification of identification of methyl-transferase
proteins.

<>

<1>Harro, J.M., Daugherty, S., Bruno, V.M., Jabra-Rizk, M.A., Rasko, D.A., Shirtliff, M.E.
<2>Draft Genome Sequence of the Methicillin-Resistant Staphylococcus aureus Isolate  MRSA-M2.
<3>Genome Announcements
<4>1
<5>e00037-12
<6>2013
<7>We report the draft genome sequence of a methicillin-resistant strain of Staphylococcus
aureus, designated MRSA-M2. This clinical isolate was obtained
from an osteomyelitis patient undergoing treatment at the University of Texas
Medical Branch (Galveston, TX). This strain is an ST30, spa type T019, agr III
strain and has been utilized as a model S. aureus strain in a number of
proteomic, transcriptomic, and animal model studies.

<>

<1>Hartke, A., Benachour, A., Boutibonnes, P., Auffray, Y.
<2>Characterization of a complex restriction/modification system detected in a Bifidobacterium longum strain.
<3>Appl. Microbiol. Biotechnol.
<4>45
<5>132-136
<6>1996
<7>Two type-II restriction endonucleases, BloI and BloII, have been detected in a
Bifidobacterium longum strain.  BloI is influenced by dam methylation: it cleaves dam- but not
dam+ DNA.  It shows a temperature and pH optimum of 45oC and pH 7.5.  Restriction analysis
and cloning experiments showed that the recognition sequence is RGATCY and that the enzyme
cuts 5' to the guanine residue.  It is an isoschizomer of commercial enzymes, BstYI and XhoII.
The second activity is not inhibited by dam methylation.  It has a temperature optimum between
25oC and 30oC and shows a broad pH optimum between 4.5 and 7.0.  The activity is thermolabile
and can be heat-killed by a 5 min incubation at 60oC.  Cloning and sequencing experiments
revealed that its recognition sequence is CTGCAG and that it cuts 5' to the second guanine
residue
in the sequence.  This enzyme is the first described isoschizomer of PstI.

<>

<1>Hartman, A.L., Norais, C., Badger, J.H., Delmas, S., Haldenby, S., Madupu, R., Robinson, J., Khouri, H., Ren, Q., Lowe, T.M., Maupin-Furlow, J., Pohlschroder, M., Daniels, C., Pfeiffer, F., Allers, T., Eisen, J.A.
<2>The Complete Genome Sequence of Haloferax volcanii DS2, a Model Archaeon.
<3>PLoS ONE
<4>5
<5>E9605
<6>2010
<7>BACKGROUND: Haloferax volcanii is an easily culturable moderate halophile
that grows on simple defined media, is readily transformable, and has a
relatively stable genome. This, in combination with its biochemical and
genetic tractability, has made Hfx. volcanii a key model organism, not
only for the study of halophilicity, but also for archaeal biology in
general. METHODOLOGY/PRINCIPAL FINDINGS: We report here the sequencing and
analysis of the genome of Hfx. volcanii DS2, the type strain of this
species. The genome contains a main 2.848 Mb chromosome, three smaller
chromosomes pHV1, 3, 4 (85, 438, 636 kb, respectively) and the pHV2
plasmid (6.4 kb). CONCLUSIONS/SIGNIFICANCE: The completed genome sequence,
presented here, provides an invaluable tool for further in vivo and in
vitro studies of Hfx. volcanii.

<>

<1>Hartman, N., Zinder, N.D.
<2>The effect of B specific restriction and modification of DNA on linkage relationships in f1 bacteriophage II.  Evidence for a heteroduplex intermediate in f1 recombination.
<3>J. Mol. Biol.
<4>85
<5>357-369
<6>1974
<7>We have analyzed the results of three gene II amber x gene II amber f1 phage
crosses.  Each was done in a non-restricting (K) host, and in a restricting (B)
host.  In each cross, only one parent was sensitive to B restriction.  The
other parent was protected from B restriction, either because of a combination
of genetic mutation at one site governing sensitivity to B restriction, and B
specific modification at the other, or because of genetic mutation at both
sites.  In all cases, B restriction resulted in the disruption of linkage
relationships between the gene II region and the unselected sensitivity site
markers.  Previously we have shown that when such crosses involve a protected
parent which is B modified at both sensitivity sites, linkage relationships
remain the same under restricting and non-restricting conditions.  Hence, the
SB protection is conferred by mutation rather than modification.  Taken
together, these results imply that, in a restricted cross, the B host
specificity system can distinguish a protected parent which is B modified from
one which is a sensitivity site mutant.  Since such a distinction could be made
most easily on an intermediate structure containing hybrid DNA, we interpret
these results in terms of a recombination mechanism mediated by an asymmetric
heteroduplex.

<>

<1>Hartman, N., Zinder, N.D.
<2>The effect of B specific restriction and modification of DNA on linkage relationships in f1 bacteriophage.  I.  Studies on the mechanism of B restriction in vivo.
<3>J. Mol. Biol.
<4>85
<5>345-356
<6>1974
<7>We have examined the effect of B specific restriction and modification of DNA
on bacteriophage f1 recombination, using a procedure which enabled us to
isolate the products of individual recombination events.  We have analyzed the
results of a series of recombination experiments, each consisting of a cross
between two gene II amber mutants, carried out both under conditions in which
neither parent was restricted, and under conditions in which one parent was
restricted while the other was protected from restriction because of B specific
modification of its genome.  At least half of the recombination events under
both restricting and nonrestricting conditions generated one parent and one
recombinant.  By considering the ratio of recombinant types emerging wiwth each
parent, linkage relationships between selected and unselected markers were
established.  These linkage relationships remained the same under restricting
and non-restricting conditions, except that under restricting conditions,
neither the sensitive parent nor the class of recombinants normally associated
with that parent was found.  We interpret this result as evidence that
restriction cleavage does not occur at the sites governing sensitivity to B
restriction, and perhaps is random.

<>

<1>Hartmann, H., Goebel, W.
<2>A new restriction enzyme from Enterobacter cloacae (EclI).
<3>FEBS Lett.
<4>80
<5>285-287
<6>1977
<7>Deoxyribonucleases, which recognize specific nucleotide sequences in a duplex
deoxypolynucleotide chain, are designated as restriction enzymes.  These
enzymes, which may be partially involved in the phenomenon of genetic
restriction, have been isolated from many bacterial species.  Among them are
relatively few members of the large family of enterobacteriaceae.  We wish to
report here the isolation of a restriction enzyme of type II from an
Enterobacter cloacae strain.  As judged from the lambdacleavage pattern
obtained with this enzyme, which is designated EclI, it recognizes a sequence
different from that of a large variety of other type II restriction enzymes.

<>

<1>Harvey, Z.H., Snider, M.J.
<2>Draft Genome Sequence of the Nicotinate-Metabolizing Soil Bacterium Bacillus niacini DSM 2923.
<3>Genome Announcements
<4>2
<5>e01251-14
<6>2014
<7>Bacillus niacini is a member of a small yet diverse group of bacteria able to catabolize
nicotinic acid. We report here the availability of a draft genome for
B. niacini, which we will use to understand the evolution of its namesake
phenotype, which appears to be unique among the species in its phylogenetic
neighborhood.

<>

<1>Harvill, E.T., Goodfield, L.L., Ivanov, Y., Meyer, J.A., Newth, C., Cassiday, P., Tondella, M.L., Liao, P., Zimmerman, J., Meert, K., Wessel, D., Berger, J., Dean, J.M., Holubkov, R., Burr, J., Liu, T., Brinkac, L., Kim, M., Losada, L.
<2>Genome Sequences of 28 Bordetella pertussis U.S. Outbreak Strains Dating from 2010 to 2012.
<3>Genome Announcements
<4>1
<5>e01075-13
<6>2013
<7>Despite the availability of highly effective vaccines, Bordetella pertussis incidence has been
rapidly rising in highly vaccinated populations. Recent
outbreaks have received media attention, feeding concerns about the emergence of
dangerous new strains with increased virulence or that escape vaccine-induced
immunity. To accelerate the study of this reemerging pathogen, we sequenced the
genomes of 28 B. pertussis strains isolated during outbreaks from 2010 through
2012, making both strains and sequence data available to the scientific
community.

<>

<1>Harvill, E.T., Goodfield, L.L., Ivanov, Y., Smallridge, W.E., Meyer, J.A., Cassiday, P.K., Tondella, M.L., Brinkac, L., Sanka, R., Kim, M., Losada, L.
<2>Genome Sequences of Nine Bordetella holmesii Strains Isolated in the United States.
<3>Genome Announcements
<4>2
<5>e00438-14
<6>2014
<7>An increasing number of pertussis-like cases are attributed to the emergent pathogen
Bordetella holmesii. The genomes of 9 clinical isolates show that they
are clonal, lack the virulence factors encoded by B. pertussis, and are more
similar to nonpertussis bordetellae. New markers for B. holmesii can be developed
using these sequences.

<>

<1>Harwich, M.D. Jr., Alves, J.M., Buck, G.A., Strauss, J.F. III, Patterson, J.L., Oki, A.T., Girerd, P.H., Jefferson, K.K.
<2>Drawing the line between commensal and pathogenic Gardnerella vaginalis through genome analysis and virulence studies.
<3>BMC Genomics
<4>11
<5>375
<6>2010
<7>BACKGROUND: Worldwide, bacterial vaginosis (BV) is the most common vaginal
disorder. It is associated with risk for preterm birth and HIV infection. The
etiology of the condition has been debated for nearly half a century and the lack
of knowledge about its cause and progression has stymied efforts to improve
therapy and prevention. Gardnerella vaginalis was originally identified as the
causative agent, but subsequent findings that it is commonly isolated from
seemingly healthy women cast doubt on this claim. Recent studies shedding light
on the virulence properties of G. vaginalis, however, have drawn the species back
into the spotlight. RESULTS: In this study, we sequenced the genomes of a strain
of G. vaginalis from a healthy woman, and one from a woman with bacterial
vaginosis. Comparative analysis of the genomes revealed significant divergence
and in vitro studies indicated disparities in the virulence potential of the two
strains. The commensal isolate exhibited reduced cytotoxicity and yet the
cytolysin proteins encoded by the two strains were nearly identical, differing at
a single amino acid, and were transcribed at similar levels. The BV-associated
strain encoded a different variant of a biofilm associated protein gene and
demonstrated greater adherence, aggregation, and biofilm formation. Using filters
with different pore sizes, we found that direct contact between the bacteria and
epithelial cells is required for cytotoxicity. CONCLUSIONS: The results indicated
that contact is required for cytotoxicity and suggested that reduced cytotoxicity
in the commensal isolate could be due to impaired adherence. This study outlines
two distinct genotypic variants of G. vaginalis, one apparently commensal and one
pathogenic, and presents evidence for disparate virulence potentials.

<>

<1>Hasan, N., Kim, S.C., Podhajska, A.J., Szybalski, W.
<2>A novel multistep method for generating precise unidirectional deletions using BspMI, a class-IIS restriction enzyme.
<3>Gene
<4>50
<5>55-62
<6>1986
<7>A novel approach is described that permits the introduction of unidirectional
deletions into a cloned DNA fragment, in a precisely controlled manner.  The
method is based on the use of a special vector and a class-IIS restriction
endonuclease, BspMI, which produces staggered cuts 4 and 8 nucleotides (nt) to
the 3' from its recognition site 5' -ACCTGC-3'.  The DNA fragment is inserted
into the pUC19-based plasmid, which contains a unique BspMI recognition site,
and the appropriate number of cleavage-and-deletion cycles is performed, each
cycle removing 4 bp.  Since the recognition site is not affected by the BspMI
cleavage, no recloning of the DNA fragment is necessary.  Each cycle consists
of (i) BspMI cleavage, (ii) removal of the 4-nt single-stranded cohesive ends
with mung bean nuclease (MB), and (iii) blunt-end ligation to recircularize the
plasmid.  The shortened plasmid is reintroduced into the host, after one or
after several such 4-bp deletion cycles.  When DNA is inserted into the
multiple cloning site in the lacZ-alpha gene, the progress of 4-bp removal can
be followed by determining the Lac phenotype, since removal of multiples of 3
bp retains the reading frame while other kinds of deletions distort (or
restore) the reading frame.  Loss of pre-existing restriction sites or creation
of new ones also permits monitoring the progress of the deletion process.  The
partial fill-in of the cohesive ends with less than four deoxyribonucleotide
triphosphates, as mediated by Klenow fragment of E. coli DNA polymerase I
(PolIk), would result in 1,2 or 3-bp deletions or additions in a given cleavage
cycle, which permits one to create precise deletions.  An analogous series of
cycles of BspMI cleavage employing a PolIk-mediated fill-in reaction instead of
the MB digestion step, and followed by ligation, would permit one to synthesize
various 4-bp direct-repeat sequences and desired variants of them.

<>

<1>Hasan, N.A., Grim, C.J., Haley, B.J., Chun, J., Alam, M., Taviani, E., Hoq, M., Munk, A.C., Saunders, E., Brettin, T.S., Bruce, D.C., Challacombe, J.F., Detter, J.C., Han, C.S., Xie, G., Nair, G.B., Huq, A., Colwell, R.R.
<2>Comparative genomics of clinical and environmental Vibrio mimicus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>21134-21139
<6>2010
<7>Whether Vibrio mimicus is a variant of Vibrio cholerae or a separate species has
been the subject of taxonomic controversy. A genomic analysis was undertaken to
resolve the issue. The genomes of V. mimicus MB451, a clinical isolate, and
VM223, an environmental isolate, comprise ca. 4,347,971 and 4,313,453 bp and
encode 3,802 and 3,290 ORFs, respectively. As in other vibrios, chromosome I
(C-I) predominantly contains genes necessary for growth and viability, whereas
chromosome II (C-II) bears genes for adaptation to environmental change. C-I
harbors many virulence genes, including some not previously reported in V.
mimicus, such as mannose-sensitive hemagglutinin (MSHA), and enterotoxigenic
hemolysin (HlyA); C-II encodes a variant of Vibrio pathogenicity island 2
(VPI-2), and Vibrio seventh pandemic island II (VSP-II) cluster of genes.
Extensive genomic rearrangement in C-II indicates it is a hot spot for evolution
and genesis of speciation for the genus Vibrio. The number of virulence regions
discovered in this study (VSP-II, MSHA, HlyA, type IV pilin, PilE, and integron
integrase, IntI4) with no notable difference in potential virulence genes between
clinical and environmental strains suggests these genes also may play a role in
the environment and that pathogenic strains may arise in the environment.
Significant genome synteny with prototypic pre-seventh pandemic strains of V.
cholerae was observed, and the results of phylogenetic analysis support the
hypothesis that, in the course of evolution, V. mimicus and V. cholerae diverged
from a common ancestor with a prototypic sixth pandemic genomic backbone.

<>

<1>Hashimoto, H., Shimizu, T., Imasaki, T., Kato, M., Shichijo, N., Kita, K., Sato, M.
<2>Crystal structures of type II restriction endonuclease Eco0109I and its complex with cognate DNA.
<3>J. Biol. Chem.
<4>280
<5>5605-5610
<6>2005
<7>EcoO109I is a type II restriction endonuclease that recognizes the DNA sequence of RGGNCCY.
Here we describe the crystal structures of
EcoO109I and its complex with DNA. A comparison of the two structures
shows that the catalytic domain moves drastically to capture the DNA.
One metal ion and two water molecules are observed near the active site
of the DNA complex. The metal ion is a Lewis acid that stabilizes the
pentavalent phosphorus atom in the transition state. One water
molecule, activated by Lys-126, attacks the phosphorus atom in an S(N)2
mechanism, whereas the other water interacts with the W-leaving oxygen
to donate a proton to the oxygen. EcoO109I is similar to EcoRI family
enzymes in terms of its DNA cleavage pattern and folding topology of
the common motif in the catalytic domain, but it differs in the manner
of DNA recognition. Our findings propose a novel classification of the
type II restriction endonucleases and lead to the suggestion that
EcoO109I represents a new subclass of the EcoRI family.

<>

<1>Hashimoto, H., Takahashi, H., Nishioka, M., Fujiwara, S., Takagi, M., Imanaka, T., Inoue, T., Kai, Y.
<2>Crystallographic study of intein homing endonuclease II encoded in the archaeal DNA polymerase gene.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>56
<5>1185-1186
<6>2000
<7>Intein homing endonucleases are proteins spliced out from a precursor protein and
site-specific enzymes that make double-strand breaks in inteinless alleles. Crystals of intein
homing endonuclease II from the hyperthermophilic archaeon Pyrococcus kodakaraensis strain
KOD1 (PI-PkoII) have been grown at room temperature using ammonium sulfate as a precipitant.
The diffraction pattern of the crystal extends to 3.0 A resolution at room temperature upon
exposure to synchrotron X-rays at KEK-PF, Japan. The crystals have symmetry consistent with
space group C222(1), with unit-cell parameters a = 107.6, b = 150.5, c = 146.8 A. A full set
of X-ray diffraction data were collected to 3.0 A Bragg spacing from a native crystal with an
overall R(merge) of 4.8% and a completeness of 96.6%.

<>

<1>Hashimoto-Gotoh, T.
<2>Quantitative determination of effective nibbling activities contaminating restriction endonuclease preparations.
<3>Anal. Biochem.
<4>231
<5>230-236
<6>1995
<7>A simple and sensitive procedure with which to detect residual exonucleolytic nibbling
activities contaminating restriction endonuclease preparations is described.  The procedure
uses the kyosei-plasmid, pKF4, which confers kanamycin resistance and enforces streptomycin
sensitivity encoded by the trp promoter/operator-driven rpsL+4amber (POtrp-rpsL+4am) gene onto
Escherichia coli streptomycin-resistant, amber-suppressive, trp repressor-negative strains
such as TH5.  When TH5 cells transformed by pKF4 were selected on agar medium containing
kanamycin plus streptomycin, the efficiency of transformation plating was substantially lower
than that on agar containing kanamycin alone.  However, when pKF4 DNA was digested by
restriction enzymes that cut once per molecule within POtrp-rpsL+4am and religated, the
plating efficiency increased depending on the degree of contamination of exonucleolytic
nibbling activities in the enzyme preparations, due to deletion mutation at the ligation
junction.  Plating efficiency was converted to "effective nibbling activity" corresponding to
Bal31 nuclease-equivalent units.  Using this procedure, effective nibbling activities were
detected in 17 of 34 commercial samples of restriction enzymes tested.  The method is simple
and more sensitive than the procedures used by the commercial suppliers and it is applicable
to the quality control testing of more than 100 restriction enzymes.

<>

<1>Hashimoto-Gotoh, T., Tsujimura, A., Ogasahara, Y.
<2>Detection of exonuclease activities in restriction endonuclease preparations using an enforcement plasmid for kanamycin-resistance selection.
<3>Gene
<4>164
<5>41-44
<6>1995
<7>A new enforcement (kyosei-) cloning plasmid vector, designated pKF4, was constructed which
confers kanamycin resistance (KmR) and enforces streptomycin sensitivity (SmS).  Since it is
important to employ restriction endonuclease (ENase) preparations free of exonuclease (Exo)
activities for effective use of the kyosei-cloning procedure, ENases such as HpaI and SmaI
purchased from four different suppliers were examined for possible contamination by
exonucleases using pKF4.  The plasmid DNA was digested with either ENase, ligated and
transformed into Escherichia coli mutants, rpsL, supE, trpR.  With pKF4 intact DNA (approx. 8
ng), 2.3 x 10/5 KmR transformant and four KmRsmR transformant colonies were obtained: the
efficiency of transformation plating (ETP) of the intact DNA was approx. 2 x 10-5.  On the
other hand, the ETP values were significantly higher by one to three orders of magnitude when
cut and rejoined DNAs were used under the same conditions in six out of either ENase samples
examined.  The results indicate that even commercially supplied ENases, that should have
passed their quality control test, could have been contaminated with Exo sufficient to
interfere with effective use of the kyosei-cloning method.  Therefore, it is advisable to
examine ENase samples for possible contamination with Exo activities, in order to choose the
right preparations for this method at the beginning of the experiments.

<>

<1>Hasman, H., Mevius, D., Veldman, K., Olesen, I., Aarestrup, F.M.
<2>beta-Lactamases among extended-spectrum beta-lactamase (ESBL)-resistant Salmonella from poultry, poultry products and human patients in The Netherlands.
<3>J. Antimicrob. Chemother.
<4>56
<5>115-121
<6>2005
<7>OBJECTIVES: The purpose of this work was to study the genetic determinants
responsible for extended-spectrum beta-lactamase (ESBL) resistance of
Salmonella isolated from Dutch poultry, poultry meat and hospitalized
humans. METHODS: Thirty-four ESBL-resistant Salmonella isolates from The
Netherlands were tested towards 21 antimicrobial agents. PCR and
sequencing were used to determine the underlying genetic determinants
responsible for the ESBL phenotypes. The transferability of the ESBL
phenotypes was tested by conjugation to a susceptible Salmonella enterica
serovar Dublin and plasmid purification, restriction fragment length
polymorphism (RFLP) and pulsed-field gel electrophoresis (PFGE) were
employed to further characterize a subset of the isolates. RESULTS: A
great genetic diversity was seen among the isolates. The bla(TEM-52) gene
was most predominant and was found among Salmonella enterica serovars
Blockley, Thomson, London, Enteritidis phage type 14b, Paratyphi B,
Virchow and Typhimurium phage types 11 and 507. We also found the
bla(TEM-20) gene in S. Paratyphi B var. Java and the bla(TEM-63) gene in
S. Isangi. Furthermore, we detected the bla(CTX-M-28) gene in S. Isangi
and the bla(CTX-M-3) gene in S. Typhimurium phage type 507. The
bla(CTX-M-2) gene was identified in S. Virchow, which also contained a
copy of the bla(SHV-2) gene and a copy of the bla(TEM-1) gene. The
bla(SHV-12) gene was found alone in S. Concord and together with the
bla(TEM-52) gene in S. Typhimurium. Finally, the bla(ACC-1) gene was
cloned from a S. Bareilly isolate and was found to be present on
indistinguishable plasmids in all S. Bareilly isolates examined as well as
in a S. Braenderup isolate and a S. Infantis isolate. CONCLUSIONS: Our
data underscore the diversity of ESBL genes in Salmonella enterica
isolated from animals, food products and human patients.

<>

<1>Hassan, K.A., Elbourne, L.D., Tetu, S.G., Johnson, E.A., Paulsen, I.T.
<2>Genome Sequence of the Neurotoxigenic Clostridium butyricum Strain 5521.
<3>Genome Announcements
<4>2
<5>e00632-14
<6>2014
<7>Clostridium strains from six phylogenetic groups, C. botulinum groups I to IV, C. baratii, and
C. butyricum, display the capacity to produce botulinum neurotoxin.
Here, we present the genome sequence of a C. butyricum isolate, the
neurotoxigenic strain 5521, which encodes the type E botulinum neurotoxin.

<>

<1>Hassan, K.A., Tetu, S.G., Elbourne, L.D., Johnson, E.A., Paulsen, I.T.
<2>Genome Sequence of the Group III Clostridium botulinum Strain Eklund-C.
<3>Genome Announcements
<4>1
<5>e0004413
<6>2013
<7>The neurotoxins produced by Clostridium botulinum strains are among the world's most potent
toxins and are the causative agents of paralytic botulism. Here, we
present the draft genome sequence of the group III C. botulinum strain Eklund-C,
including a pseudolysogen-like bacteriophage that harbors the type C neurotoxin
operon.

<>

<1>Hassan, S.S. et al.
<2>Complete genome sequence of Corynebacterium pseudotuberculosis biovar ovis strain P54B96 isolated from antelope in South Africa obtained by rapid next generation  sequencing technology.
<3>Standards in Genomic Sciences
<4>7
<5>189-199
<6>2012
<7>The Actinobacteria, Corynebacterium pseudotuberculosis strain P54B96, a nonmotile,
non-sporulating and a mesophile bacterium, was isolated from liver,
lung and mediastinal lymph node lesions in an antelope from South Africa. This
strain is interesting in the sense that it has been found together with
non-tuberculous mycobacteria (NTMs) which could nevertheless play a role in the
lesion formation. In this work, we describe a set of features of C.
pseudotuberculosis P54B96, together with the details of the complete genome
sequence and annotation. The genome comprises of 2.34 Mbp long, single circular
genome with 2,084 protein-coding genes, 12 rRNA, 49 tRNA and 62 pseudogenes and a
G+C content of 52.19%. The analysis of the genome sequence provides means to
better understanding the molecular and genetic basis of virulence of this
bacterium, enabling a detailed investigation of its pathogenesis.

<>

<1>Hassan, Y.I., Lepp, D., He, J., Zhou, T.
<2>Draft Genome Sequences of Devosia sp. Strain 17-2-E-8 and Devosia riboflavina Strain IFO13584.
<3>Genome Announcements
<4>2
<5>e00994-14
<6>2014
<7>Here we report the draft genome of Devosia sp. strain 17-2-E-8, isolated from Ontario
agricultural soil (Canada) with promising deoxynivalenol
biotransformation capabilities. In addition, we report the draft genome of
Devosia riboflavina strain IFO13584, used as a control strain in our studies
aimed at highlighting unique gene clusters involved in deoxynivalenol
epimerization.

<>

<1>Hassani, I.I., Robert, C., Michelle, C., Raoult, D., Hacene, H., Desnues, C.
<2>Non-contiguous finished genome sequence and description of Halopiger djelfamassiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>160-174
<6>2013
<7>Halopiger djelfamassiliensis strain IIH2(T) sp. nov. is the type strain of Halopiger
djelfamassiliensis sp. nov., a new species within the genus Halopiger.
This strain, whose genome is described here, was isolated from evaporitic
sediment of the hypersaline Lake Zahrez Gharbi in the Djelfa region (Algeria). H.
Djelfamassiliensis is a Gram-negative, polymorphic-shaped and strictly aerobic
archaeon. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 3,771,216 bp long genome-contains
3,761 protein-coding and 51 RNA genes, including 4 rRNA genes.

<>

<1>Hassaninasab, A., Hashimoto, Y., Tomita-Yokotani, K., Kobayashi, M.
<2>Discovery of the curcumin metabolic pathway involving a unique enzyme in an intestinal microorganism.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>6615-6620
<6>2011
<7>Polyphenol curcumin, a yellow pigment, derived from the rhizomes of a plant (Curcuma longa
Linn) is a natural antioxidant exhibiting a variety of pharmacological activities and
therapeutic properties. It has long been used as a traditional medicine and as a preservative
and coloring agent in foods. Here, curcumin-converting microorganisms were isolated from human
feces, the one exhibiting the highest activity being identified as Escherichia coli. We are
thus unique in discovering that E. coli was able to act on curcumin. The curcumin-converting
enzyme was purified from E. coli and characterized. The native enzyme had a molecular mass of
about 82 kDa and consisted of two identical subunits. The enzyme has a narrow substrate
spectrum, preferentially acting on curcumin. The microbial metabolism of curcumin by the
purified enzyme was found to comprise a two-step reduction, curcumin being converted
NADPH-dependently into an intermediate product, dihydrocurcumin, and then the end product,
tetrahydrocurcumin. We named this enzyme 'NADPH-dependent curcumin/dihydrocurcumin
reductase' (CurA). The gene (curA) encoding this enzyme was also identified. A homology
search with the BLAST program revealed that a unique enzyme involved in curcumin metabolism
belongs to the medium-chain dehydrogenase/reductase superfamily.

<>

<1>Hata, E.
<2>Complete Genome Sequence of Mycoplasma canadense Strain HAZ 360_1 from Bovine Mastitic Milk in Japan.
<3>Genome Announcements
<4>2
<5>e00984-14
<6>2014
<7>Bovine mycoplasmal mastitis is spreading quickly among cows. Mycoplasma canadense, a causal
species of bovine mastitis, reduces milk quality and quantity
via the infiltration of numerous inflammatory cells. Presented here is the
complete 693,241-bp genome sequence of M. canadense strain HAZ 360_1, which was
isolated in Japan.

<>

<1>Hata, E., Murakami, K.
<2>Complete Genome Sequence of Mycoplasma californicum Strain HAZ160_1 from Bovine Mastitic Milk in Japan.
<3>Genome Announcements
<4>2
<5>e00684-14
<6>2014
<7>Bovine mycoplasmal mastitis is spreading quickly among cows. It often leads to clinical
mastitis outbreaks and often results in huge economic losses. Mycoplasma
californicum is an important causal species of bovine mastitis. Presented here is
the 799,088-bp complete genome sequence of M. californicum strain HAZ160_1, which
was isolated in Japan.

<>

<1>Hata, K., Okano, M., Lei, H., Li, E.
<2>Dnmt3L cooperates with the Dnmt3 family of de novo DNA methyltransferases to establish maternal imprints in mice.
<3>Development
<4>129
<5>1983-1993
<6>2002
<7>Genomic imprinting is regulated by differential methylation of the paternal and maternal
genome. However, it remains unknown how parental imprinting is established during
gametogenesis. In this study, we demonstrate that Dnmt3L, a protein sharing homology with DNA
methyltransferases, Dnmt3a and Dnmt3b, but lacking enzymatic activity, is essential for the
establishment of maternal methylation imprints and appropriate expression of maternally
imprinted genes. We also show that Dnmt3L interacts with Dnmt3a and Dnmt3b and co-localizes
with these enzymes in the nuclei of transfected cells, suggesting that Dnmt3L may regulate
genomic imprinting via the Dnmt3 family enzymes. Consistent with this model, we show that
[Dnmt3a(-/-), Dnmt3b()] mice also fail to establish maternal methylation imprints. In
addition, both Dnmt3a and Dnmt3L are required for spermatogenesis. Together, our findings
suggest that Dnmt3L may cooperate with Dnmt3 family methyltransferases to carry out de novo
methylation of maternally imprinted genes in oocytes.

<>

<1>Hatfull, G.F.
<2>Complete Genome Sequences of 138 Mycobacteriophages.
<3>J. Virol.
<4>86
<5>2382-2384
<6>2012
<7>Bacteriophages are the most numerous biological entities in the biosphere, and
although their genetic diversity is high, it remains ill defined.
Mycobacteriophages-the viruses of mycobacterial hosts-provide insights into this
diversity as well as tools for manipulating Mycobacterium tuberculosis. We report
here the complete genome sequences of 138 new mycobacteriophages, which-together
with the 83 mycobacteriophages previously reported-represent the largest
collection of phages known to infect a single common host, Mycobacterium
smegmatis mc(2) 155.

<>

<1>Hatfull, G.F. et al.
<2>Exploring the mycobacteriophage metaproteome: phage genomics as an educational platform.
<3>PLoS Genet.
<4>2
<5>e92
<6>2006
<7>Bacteriophages are the most abundant forms of life in the biosphere and carry genomes
characterized by high genetic diversity and mosaic
architectures. The complete sequences of 30 mycobacteriophage genomes show
them collectively to encode 101 tRNAs, three tmRNAs, and 3,357 proteins
belonging to 1,536 "phamilies" of related sequences, and a statistical
analysis predicts that these represent approximately 50% of the total
number of phamilies in the mycobacteriophage population. These phamilies
contain 2.19 proteins on average; more than half (774) of them contain
just a single protein sequence. Only six phamilies have representatives in
more than half of the 30 genomes, and only three-encoding tape-measure
proteins, lysins, and minor tail proteins-are present in all 30 phages,
although these phamilies are themselves highly modular, such that no
single amino acid sequence element is present in all 30 mycobacteriophage
genomes. Of the 1,536 phamilies, only 230 (15%) have amino acid sequence
similarity to previously reported proteins, reflecting the enormous
genetic diversity of the entire phage population. The abundance and
diversity of phages, the simplicity of phage isolation, and the relatively
small size of phage genomes support bacteriophage isolation and
comparative genomic analysis as a highly suitable platform for
discovery-based education.

<>

<1>Hattman, S.
<2>The functioning of T-even phages with unglucosylated DNA in restricting Escherichia coli host cells.
<3>Virology
<4>24
<5>333-348
<6>1964
<7>The DNA of the T-even bacteriophages contains glucose bound to the pyrimidine
base 5-hydroxymethylcytosine.  Modified forms of T-even phage, designated T*
phage, which contain little or no glucose, are produced by growth in certain
bacterial mutants blocked at some step in the synthesis of uridine
diphosphoglucose.  The T* phage is able to grow on some strains of Shigella
(permissive hosts) but grows poorly or not at all on various strains of
Escherichia coli (restrictive hosts).  Evidence is presented that the DNA of T*
phage undergoes extensive degradation to acid-soluble fragments following
infection of restricting E. coli cells, whereas infection of E. coli with
wild-type phage, or of Shigella with T* phage, causes very little degradation
of phage DNA.  T* phage can perform certain functions after infection of
restricting hosts.  In these respects the T* forms of phages T2,T4, and T6
differ to some extent.  Also, phage T*2 can undergo extensive growth activation
in E. coli cells in multiple infection, whereas T*4 and T*6 do not exhibit this
multiplicity activation.  In mixed infection of E. coli B with the mutant
T4am122, unable to induce synthesis of the enzyme
deoxycytidylate-hydroxymethylase, T*2 and T*6 are capable of complementing the
missing function, whereas T*4 cannot.  Evidence is presented that T* phages can
in fact direct synthesis of some early enzymes in restricting hosts.  Despite
the occurrence of some early enzyme synthesis, phage DNA synthesis does not
occur after infection of restricting bacteria with T* phage except in the cells
where restriction fails.  The ability of permissive hosts to support
replication of T* phage is not due to an initial glucosylation of the incoming
nonglucosylated DNA.

<>

<1>Hattman, S.
<2>Methylation of adenine residues in bacteriophage T2 DNA.
<3>Virology
<4>49
<5>404-412
<6>1972
<7>It has been previously shown that uP1 mutants, derived from phage T2 gt rP1, determine an
altered form of the phage-induced DNA methylase. The uP1 enzyme methylates two- to threefold
more adenine residues on phage DNA than does the wild-type methylase. The question of whether
the presence of 5-hydroxymethylcytosine (HMC) in T2 DNA plays a role in determining methylase
specificity was investigated. A triple amber mutantn was constructed which is defective in the
synthesis of the enzymes, deoxycytidine triphosphatase (dCTPase) and deoxycytidylate
hydroxymethylase (dCMP-HMase) as well as in the ability to degrade completely host DNA (genes
56,42 and 47, respectively). In host cells lacking an amber suppressor (su-), the mutant is
shown to synthesize DNA which contains cytosine (C) in place of HMC. This C-containing DNA has
the same level of N6-methyladenine (MeAde) as observed in the HMC-containing DNA of T2 gt. rP1
DNAs behaved similarly as substrates for in vitro methylation by extracts of phage-infected
cells. These data indicate that the 5-hydroxymethyl group on HMC does not interfere with the
activity of the wilde-type DNA methylase, nor does it enhance the activity of the uP1 enzyme.

<>

<1>Hattman, S.
<2>Partial purification of the Escherichia coli K-12 mec+ deoxyribonucleic acid-cytosine methylase:  in vitro methylation completely protects bacteriophage lambda deoxyribonucleic acid against cleavage by R.EcoRII.
<3>J. Bacteriol.
<4>129
<5>1330-1334
<6>1977
<7>A procedure is described for the partial purification of the deoxyribonucleic
acid (DNA)-cytosine methylases controlled by the RII plasmid and by the
Escherichia coli mec+ gene.  The two enzymes exhibit similar but distinct
chromatographic behavior on diethylaminoethyl-cellulose and phosphocellulose.
Preliminary studies on the two methylases indicate that they are
indistinguishable with respect to their Km for S-adenosylmethionine and their
pH [in tris(hydroxymethyl)aminomethane buffer] and NaCl concentration optima.
In vitro methylation of various phage lambda DNA substrates by the mec+ or RII
enzyme modifies the DNA to a form that is completely resistant to
double-stranded cleavage by the RII restriction endonuclease (R.EcoRII).  These
results are consistent with our earlier proposal that the mec+ methylase
recognizes RII host specificity sites.

<>

<1>Hattman, S.
<2>Specificity of the bacteriophage Mu mom+-controlled DNA modification.
<3>J. Virol.
<4>34
<5>277-279
<6>1980
<7>Bacteriophage Mu DNA was labeled after induction in the presence of
[8-3H]adenine.  Purified DNA was enzymatically digested, and the 3H-labeled
dinucleotides were isolated.  Approximately 15 to 20% of the adenine residues
were modified to a new form, Ax, as observed previously (S. Hattman, J. Virol.
32:468-475, 1979) in bulk DNA.  Paper electrophoretic analysis revealed that
only two dinucleotide species contain Ax, namely, (Ax,C) and (Ax,G).  The
observation that only C and G are the nearest neighbors of Ax is consistent
with the proposal of Kahmann and Kamp (R. Kahmann and D. Kamp, J. Mol. Biol.,
in press) that modification of Mu DNA occurs at the A residue within the
pentanucleotide sequence 5'...(C/G)-A-(G/C)-N-Py...3'.

<>

<1>Hattman, S.
<2>Plasmid-controlled variation in the content of methylated bases in bacteriophage lambda deoxyribonucleic acid.
<3>J. Virol.
<4>10
<5>356-361
<6>1972
<7>The N6-methyladenine (MeAde) and 5-methylcytosine (MeC) contents in deoxyribonucleic acid
(DNA) of bacteriophage lambda has been analyzed as a function of host specificity. The
following facts have emerged: (i) lambda grown on strains harboring the P1 prophage contain
ca. 70 more MeAde residues/DNA molecule than lambda grown either in the P1-sensitive parent,
or in a P1 immune-defective lysogen which does not confer P1 modification; (ii) lambda grown
on strains harboring the N-3 drug-resistance factor contain ca. 60 more MeC residues/DNA
molecule than lambda grown on the parental strain lacking the factor; (iii) lambda grown in
Escherichia coli B strains is devoid of MeC, whereas lambda grown in a B (N-3) host contains a
high level of MeC; (iv) the MeAde content in lambda DNA is not affected by the N-3 factor.
These results suggest that P1 controls an adenine-specific DNA methylase, and that the N-3
plasmid controls a cytosine-specific DNA methylase. The N-3 factor has been observed
previously to direct cytosine-specific methylation of phage P22 DNA and E. coli B DNA in vivo;
in vitro studies presented here demonstrate this activity.

<>

<1>Hattman, S.
<2>Unusual Modification of Bacteriophage Mu DNA.
<3>J. Virol.
<4>32
<5>468-475
<6>1979
<7>Bacteriophage mu DNA was labeled after induction in the presence of [2-3H]adenine or
[8-3H]adenine. Both Mu mom+.dam+ DNA and Mu mom-.dam+ DNA have similar N6 -methyladenine
(MeAde) contents, as well as similar frequencies of MeAde nearest neighbors. Both DNAs are
sensitive to in vitro cleavage by R.DpnI but resistant to cleavage by R.DpnI but resistant to
cleavage by R.DpnII. These results indicate that the mom+ protein does not alter the sequence
specificity of the host dam+ methylase to produce MeAde at new sites. However, we have
discovered a new modified base, denoted Ax, in Mu mom+.dam+ DNA; approximately 15% of the
adenine residues are modified to Ax. Although the precise nature of the modification is not
yet defined, analysis be electrophoresis and chromatography indicates that the N6-amino group
is not the site of modification, and that the added moiety contains a free carboxyl group. Ax
is not present in Mu mom+.dam+ or Mu mom-.dam+ phage DNA or in cellular DNA from unonduced Mu
mom+.dam+ lysogens. These results suggest that expression of the dam+ and mom+ genes are
required for the Ax modification and that this modification is responsible for protecting Mu
DNA against certain restriction nucleases. Mu mom+.dam+ DNA and Mu mom-.dam- DNA contain a
very low level of MeAde (ca. 1 MeAde per 5,000 adenine residues). Since the only nearest
neighbor to MeAde appears to be cytosine, we suggest that the methylated sequence is
5'...C-A-C...3' and that this methylation is mediated by the EcoK modification enzyme.

<>

<1>Hattman, S.
<2>Plasmid-controlled variation in the content of methylated bases in single-stranded DNA phages M13 and fd.
<3>J. Mol. Biol.
<4>74
<5>749-752
<6>1973
<7>The N6-methyladenine and 5-methylcytosine contents in the DNA of bacteriophages
M13 and fd have been analyzed.  The results are summarized as follows. (1)
After growth in bacteria harboring the N-3 fi- drug resistance-factor, fd and
M13 are observed to contain approximately 1 to 2 more 5-methylcytosine residues
per DNA molecule than after rgrowth in the parental drug-sensitive host; no
effect on the N6-methyladenine content is produced by the plasmid.  (2) After
growth in bacteria harboring P1 prophage, fd and M13 are observed to contain
approximately 2 to 3 more N6-methyladenine residues per DNA molecule than after
growth in the parental P1-sensitive host; no apparent effect on the
5-methylcytosine content was produced by the P1 plasmid.  (3) In agreement with
others, fd carrying B-host specificity (fd.B) is observed to contain 2 more
N6-methyladenine residues/DNA molecule than fd.K.

<>

<1>Hattman, S.
<2>DNA Modification:  Methylation.
<3>Bacteriophage T4, ASM Publications, Mathews, C.K., Berget, P., Mosig, G., Kutter, B., 
<4>0
<5>152-155
<6>1983
<7>None

<>

<1>Hattman, S.
<2>DNA methylation of T-even bacteriophages and of their nonglucosylated mutants:  its role in P1-directed restriction.
<3>Virology
<4>42
<5>359-367
<6>1970
<7>The 6-methylaminopurine (MAP) content of bacteriophages T2,T4,T6 and their nonglucosylated gt
mutants has been analyzed. Phage T2 contains 25% more MAP than T4; the nonglucosylated forms
of T2 and T4 contain 30-100% more MAP than their respective wild-type parents; the level of
MAP is affected by the growth temperature and by the level of glucosylation. T6 and its
nonglucosylated mutants are devoid of MAP. Support for the hypothesis that methylation has a
role in P1-directed restriction of gt mutants of T-even phages (Revel and Georgopoulos, 1969)
is presented: (1) mutants of T2gt and T4gt insensitive to P1-restriction, designated uP1
(Revel and Georgopoulos, 1969) contain hypermethylated DNA; (2) cells simultaneously infected
with P1-sensitive T2gt or T6gt (designated rP1) and ultraviolet-irradiated T2gt uP1 yield
progeny rP1 phage which contain hypermethylated DNA and are partially resistant to P1
restriction. It is proposed that the uP1 mutations occur in the structural gene for phage DNA
methylase. Methylation does not appear to be involved in the restriction of T2gt and T4gt by
certain bacterial hosts that do not restrict T5gt (these bacteria are designated r6-r2,4+)
because of the findings that (1) a T4gt mutant lacking MAP is still restricted in r6-r2,4+;
(2) phenotypically methylated T6gt is accepted in r6-r2,4+.

<>

<1>Hattman, S.
<2>Variation of 6-methylaminopurine content in bacteriophage P22 deoxyribonucleic acid as a function of host specificity.
<3>J. Virol.
<4>7
<5>690-691
<6>1971
<7>The 6-methylaminopurine (MAP) content of P22 deoxyribonucleic acid has been
analyzed as a function of the host specificity it carries.  A 40 to 50%
reduction in MAP level occurs as a result of growth in host cells defective in
the ability to confer LT specificity.

<>

<1>Hattman, S.
<2>DNA methylation.
<3>The Enzymes, Academic Press, Boyer, P., 
<4>14
<5>517-548
<6>1981
<7>Almost all biological macromolecules undergo some form of processing event
subsequent to their biosynthesis.  Polypeptides are specifically folded,
cleaved, phosphorylated, acetylated, methylated, or covalently bound to other
polypeptides.  RNA molecules are specifically cleaved, lengthened, spliced,
methylated, thiolated, or otherwise base-modified.  These processes are
obligatory events in the establishment of functional expression of these
molecules.  It has been known for almost three decades that DNA is also subject
to post-replication modification.  The main topic of this chapter is to
consider the phenomenon of DNA methylation, its nature, distribution, analysis,
specificity, and biological function.  In addition, I briefly review other DNA
modifications known to occur among prokaryotes and eukaryotes.

<>

<1>Hattman, S.
<2>DNA methyltransferase-dependent transcription of the phage Mu mom gene.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>79
<5>5518-5521
<6>1982
<7>The phage Mu mom gene controls an unusual DNA modification. Expression of the mom function
requires an active host (dam+) DNA adenine methylase [S-adenosyl-L-methionine:DNA
(6-aminopurine)-methyltranasferase]; in dam- hosts, Mu development is normal except that the
viral DNA does not undergo the mom modification. The present communication compares
transcription of the mom gene in dam+ versus dam- cells. 32P-labeled probes were prepared by
nick-translation of a purified mom gene-containing restriction fragment and of virion DNA,
respectively. These probes were hybridized with various RNAs blotted onto nitrocellulose
filters (after fractionation by agarose gel electrophoresis). The salient findings are: (i)
mom-specific RNA was readily detected in dam+ lysogenic cells, but only after induction of the
Mu prophage; (ii) the level of mom RNA was decreased at least to 1/20th in induced dam- Mu
lysogens; and (iii) little difference, if any, was observed between dam+ and dam- cells with
respect to total Mu transcripts produced after prophage induction. These results are in accord
with the known pattern of mom gene expression and Mu development. They show that the host
(dam+) DNA adenine methylase activity is required for transcription of the mom gene. This
represents a unique example where a DNA methylase exerts a positive regulatory role in mRNA
transcription; alternative mechanisms for this process will be discussed.

<>

<1>Hattman, S., Brooks, J.E., Masurekar, M.
<2>Sequence specificity of the P1 modification methylase (M.EcoP1) and the DNA methylase (M.Ecodam) controlled by the Escherichia coli dam gene.
<3>J. Mol. Biol.
<4>126
<5>367-380
<6>1978
<7>Labeled oligonucleotides have been fractionated from pancreatic DNase digests of DNA that had
been methylated in vitro with the P1 modification enzyme (M.EcoP1) or with the DNA-adenine
methylase (M.Ecodam) controlled by the Escherichia coli dam gene. The sequences of methylated
oligonucleotides were established for M.Eco dam modification of calf thymus DNA. The results
show that M.Ecodam methylates adenine residues contained in the twofold symmetrical sequence,
5'...G-A-T-C...3'. The sequence for the site methylated by M.EcoP1 has also been deduced; we
proposed that M.EcoP1 modification produces the following methylated pentameric sequence:
5'...A-G-A*-C-Py...3' (where A* = N6 methyladenine and Py is C or T).

<>

<1>Hattman, S., Cousens, L.
<2>Location of the region controlling host specificity (hsII) with respect to drug resistance markers on the fi- R factor, N-3.
<3>J. Bacteriol.
<4>112
<5>1428-1430
<6>1972
<7>The region controlling host specificity (hsII) has been mapped, by P22
transduction, with respect to drug resistance markers carried on the fi- R
factor, N-3.  The relative linear sequence of contransducible markers is
Tc-Su-Sm-hsII.

<>

<1>Hattman, S., Fukasawa, T.
<2>Host-induced modification of T-even phages due to defective glucosylation of their DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>50
<5>297-300
<6>1963
<7>None

<>

<1>Hattman, S., Gold, E., Plotnik, A.
<2>Methylation of cytosine residues in DNA controlled by a drug resistance factor.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>187-190
<6>1972
<7>The proportion of 5-methylcytosine (5MeCyt) and 6-methylaminopurine
(N6-methyladenine, 6MeAde) in bacteriophage P22 DNA was analyzed as a function
of the host-specificity the phage carried.  In the DNA of P22 grown in stsrains
harboring the modifying drug-resistance-transfer-factor N-3, the 5MeCyt content
was at least twice that after growth in strains lacking the factor.  In
contrast, the 6MeAde level of P33 DNA was unaffected by the presence or absence
of the factor.  The 6MeAde and 5MeCyt levels were unaffected by factors 222 and
N-1, which do not modify phage DNA.  The 5MeCyt/6MeAde ratio was only slightly
higher int he DNA of Salmonella strains that had received the N-3 factor.
After transfer of the N-3 factor to Escherichia coli strain B, which normally
lacks 5MeCyt, a high content of 5MeCyt is observed.  We conclude that the N-3
factor controls a DNA methylase specific for cytosine residues.  If the N-3
host specificity is imparted by cytosine methylation, this would be the first
instance where a biological role for 5MeCyt has been elucidated.

<>

<1>Hattman, S., Goradia, M., Monaghan, C., Bukhari, A.I.
<2>Regulation of the DNA-modification function of bacteriophage Mu.
<3>Cold Spring Harb. Symp. Quant. Biol.
<4>47
<5>647-653
<6>1983
<7>The DNA-modification function of bacteriophage Mu, termed the mom function,
presents very interesting examples of DNA modification and regulation of gene
expression.  On both counts it sets new precedents.  The modification involved
is new, and the expression of the mom gene appears to require methylation of
sequences adjacent to the gene.

<>

<1>Hattman, S., Gribbin, C., Hutchison, C.A.
<2>In vivo methylation of bacteriophage PhiX174 DNA.
<3>J. Virol.
<4>32
<5>845-851
<6>1979
<7>A mutant (designated mec-) has been isolated from Escherichia coli C which has
lost DNA-cytosine methylase activity and the ability to protect phage lambda
against in vivo restriction by the RII endonuclease.  This situation is
analogous to that observed with an E. coli K-12 mec- mutant; thus, the E. coli
C methylase appears to have overlapping sequence specificity with the K-12 and
RII enzymes; (the latter methylases have been shown previously to recognize the
same sequence).  Covalently closed, supertwisted double-stranded DNA (RFI) was
isolated from C mec+ and C mec- cells infected with bacteriophage PhiX174.
PhiX.mec- RFI is sensitive to in vitro cleavage by R.EcoRII and is cut twice to
produce two fragments of almost equal size.  In contrast, PhiX.mec+ RFI is
relatively resistant to in vitro cleavage by R.EcoRII.  R.BstI, which cleaves
mec+/RII sites independent of the presence or absence of 5-methylcytosine,
cleaves both forms of the RFI and produces two fragments similar in size to
those with R.EcoRII.  These results demonstrate that PhiX.mec+ RFI is
methylated in vivo by the host mec+ enzyme and that this methylation protects
the DNA against cleavage by R.EcoRII.  This is consistent with the known
location of two mec+/RII sequences (viz., 5' ... C-C-A/T-G-G ...3') on the
PhiX174 map.  Mature single-stranded virion DNA was isolated from PhiX174
propagated in C mec+ or C mec- in the presence of L-[methyl-3H]methionine.
Paper chromatographic analyses of acid hydrolysates revealed that PhiX.mec+ DNA
had a 10-fold-higher ratio of [3H]5-methylcytosine to [3H]cytosine compared to
PhiX.mec-.  Since PhiX.mec+ contains, on the average, approximately 1
5-methylcytosine residue per viral DNA, we conclude that methylation of PhiX174
is mediated by the host mec+ enzyme only.  These results are not consistent
with the conclusions of previous reports that PhiX174 methylation is mediated
by a phage-induced enzyme and that methylation is essential for normal phage
development.

<>

<1>Hattman, S., Keister, T., Gottehrer, A.
<2>Sequence specificity of DNA methylases from Bacillus amyloliquefaciens and Bacillus brevis.
<3>J. Mol. Biol.
<4>124
<5>701-711
<6>1978
<7>DNA methylation in Bacillus amyloliquefaciens strain H (Bam) and Bacillus
brevis (Bbv) has been examined by a variety of techniques.  In vivo labelling
studies revealed that Bam DNA contains no N6-methyladenine (MeAde), but
contains 5-methylcytosine (MeCyt); approximately 0.7% of the cytosine residues
are methylated.  DNA methylase activity was partially purified from both Bam
and Bbv; the Bam enzyme preparation transferred methyl groups from
S-adenosyl-L-[methyl-3H]methionine ([3H]AdoMet) to specific DNA cytosine
residues only; in agreement with Vanyushin & Dobritsa (1975), the Bbv enzyme
preparation methylated both DNA adenine and cytosine residues.  The (partial)
sequence specificity of the methylases was determined by analyzing
[3H]methyl-labelled dinucleotides obtained from enzymatic digests of DNA
methylated in vitro.  Bam and Bbv each contain a DNA-cytosine methylase with
overlapping sequence specificity; e.g. both enzymes produce G-C*, C*-A and
C*-T.  This is consistent with a single, twofold symmetrical methylation
sequence of 5'...G-C*-(A or T)-G-C...3'; this was observed by Vanyushin &
Dobritsa (1975) for a different Bbv strain.  Bam contains a second DNA-cytosine
methylase (not present in Bbv), which produces T-C* and C*-T.  We propose that
this methylase is the BamI modification enzyme, and that the modified sequence
is 5'...G-G-A-G-C*-C...3'.  Bbv appears to contain two DNA-adenine methylases
which produce the (partial) methylated sequences, 5'...G-A*-T...3' and
5'...A-A*-G...3', respectively; in the former case, all the G-A-T-C sites on
Bbv DNA appear to be methylated.

<>

<1>Hattman, S., Malygin, E.G.
<2>Bacteriophage T2Dam and T4Dam DNA-[N6-adenine]-methyltransferases.
<3>Prog. Nucleic Acid Res. Mol. Biol.
<4>77
<5>67-126
<6>2004
<7>DNA mewthyltransferases are important enzymes that methylate DNA as a post replicative event.
DNA MTases, encoded by both cellular and viral genes, catalyze methyl group transfer from
S-adenosyl-L-methionine, producing S-adenosyl-L-homocysteine and methylated DNA.  The methyl
group acceptor atom is either an exocyclic amino nitrogen (N6-Ade or N4-Cyt) or a ring carbon
(C5-Cyt).  The DNA-[amino]-MTases [EC2.1.1.72 and 113]transfer methyl groups directly to the
exocyclic nitrogen without the formation of a covalent enzyme-DNA intermediate, which occurs
with the C5-Cyt MTases [EC2.1.73].  While most phokaryote DNA MTases are components of
restriction-modification systems important in protecting cells from foreign DNAs, certain
MTases do not have cognate restriction enzymes associated with them.  These include a family
(Dam) of prokaryotic DNA-adenine MTases that methylate Ade in GATC sequences.  Several
bacteriophages, such as T2 and T4, also encode Dam MTases.  Generally speaking, the Dam MTases
are not essential for viability of bacteria or phage; however, they do have a variety of
functions including regulation of transcription of certain genes, timing of DNA replication
initiation, and protection against restriction endonucleases, and they play a crucial role in
pathogenicity of intestinal bacteria.  Because they methylate specific nucleotide sequences,
they provide excellent objects for studies on protein-DNA interactions.  Valuable insights
into the organization/function of MTases have come from the identification of common motifs
discovered by amino acid-seqence alignments.  The solution of a number of MTase crystal
structures has added key details on the specific protein-DNA and protein-cofactor
interactions.  However, genetic and biochemical analyses are essential for a more complete
understanding of the functioning of these enzymes.  Here, we present results from all three
approaches directed at characterizing the bacteriophage T2/T4 Dam DNA-[N6-adenine]-MTase.

<>

<1>Hattman, S., Revel, H.R., Luria, S.E.
<2>Enzyme synthesis directed by nonglucosylated T-even bacteriophages in restriction hosts.
<3>Virology
<4>30
<5>427-438
<6>1966
<7>Phage T2gt, defective in ability to initiate production of Alpha-glucosyl
transferase, elicits the synthesis of other phage directed early enzymes in the
host bacterium Escherichia coli B in which it cannot grow.  This early-enzyme
synthesis is arrested at about the same time in infection with either T2gt or
T2, despite the fact that T2gt DNA is apparently not replicated in E. coli B.
A similar pattern of phage-enzyme synthesis is observed with other T even
phages with nonglucosylated DNA.  Experiments with UV-irradiated phage and with
inhibitors of protein synthesis indicate that the arrest of enzyme synthesis in
infection with nonglucosylated phage is due not to the regulatory process
operative in infection with normal phage, but to a different mechanism.  The
evidence suggests that this mechanism is the rapid degradation of the
nonglucosylated phage DNA.  The experiments also provide estimates for the
half-life of phage-specific mRNA in the presence or absence of protein
synthesis.

<>

<1>Hattman, S., Schlagman, S., Cousens, L.
<2>Isolation of a mutant of Escherichia coli defective in cytosine-specific deoxyribonucleic acid methylase activity and in partial protection of bacteriophage lambda against restriction by cells containing the N-3 drug-resistance factor.
<3>J. Bacteriol.
<4>115
<5>1103-1107
<6>1973
<7>A mutant (designated mec-) of Escherichia coli F+ 100 endoI- su+ rk-mk+ has
been isolated which is defective in cytosine-specific deoxyribonucleic acid
(DNA) methylase activity.  The DNA of this mutant, as well as the DNA of phage
lambda and fd propagated in it, is virtually devoid of 5-methyl-cytosine (MeC);
in contrast, the mutation has no significant effect on the level of
N6-methyladenine in DNA.  Phage lambda grown on the mec- mutant is more
strongly restricted by N-3-containing cells than is lambda grown on the mec+
parent.  These results suggest that methylation of certain cytosine residues by
the E. coli K-12 enzyme partially protects lambda DNA from either the N-3
restriction nuclease or against secondary degradation subsequent to
N-3-specific degradation.  Analysis of the MeC level in viral and cellular DNA
obtained from mec+, mec+ (mN3+), and mec- (mN3+) strains has led to the
conclusion that the R-factor controlled DNA-cytosine methylase may be capable
of methylating a sequence(s) which is a substrate for the K-12 enzyme.

<>

<1>Hattman, S., Schlagman, S., Goldstein, L., Frohlich, M.
<2>Salmonella typhimurium SA Host Specificity System is based on Deoxyribonucleic Acid-Adenine Methylation.
<3>J. Bacteriol.
<4>127
<5>211-217
<6>1976
<7>We have determined the nature of the deoxyribonucleic acid (DNA) modification
governed by the SA host specificity system of Salmonella typhimurium.  Two
lines of evidence indicate that SA modification is based on methylation of
DNA-adenine residues.  (i) The SA+ locus of Salmonella was transferred into
Escherichia coli B, a strain that does not contain 5-methylcytosine in its DNA;
although the hybrid strain was able to confer SA modification, its DNA still
did not contain 5-methylcytosine.  (ii) the N6-methyladenine content of phage L
DNA was measured after growth in various host strains; phage lacking SA
modification contained fewer N6-methyladenine residues per DNA.  We also
investigated the possibility, suggested by others, that SA modification
protects phage DNA against restriction by the RII host specificity system.
Phages lambda, P3, and L were grown in various SA+ and SA- hosts and tested for
their relative plating ability on strains containing or lacking RII
restriction; the presence or absence of SA modification had no effect on RII
restriction.  In vitro studies revealed, however, that Salmonella DNA is
protected against cleavage by purified RII restriction endonuclease (R-EcoRII).
This protection is not dependent on SA modification; rather, it appears to be
due to methylation by a DNA-cytosine methylase which has overlapping
specificity with the RII modification enzyme, but which is not involved in any
other known host specificity system.

<>

<1>Hattman, S., van Ormondt, H., de Waard, A.
<2>Sequence specificity of the wild-type (dam+) and mutant (damh) forms of bacteriophage T2 DNA adenine methylase.
<3>J. Mol. Biol.
<4>119
<5>361-376
<6>1978
<7>Non-glucosylated, non-methylated phage T2 DNA was methylated in vitro with partially purified
wild-type (dam+) or mutant (damh) T2 DNA adenine methylase. The radioactively labeled
methyladenine-containing DNA was enzymatically degraded and the resulting oligonucleotides
were separated according to chain length by DEAE-cellulose chromatography. Following
"fingerprinting" by two-dimensional electrophoresis, we determined the sequence for vaious
di-, tri- and tetranucleotides containing radioactive N6-methyldeoxyadenosine. From this
analysis we conclude that both T2 dam+ and T2 damh contain the sequence 5'...G-mA-Py...3'.

<>

<1>Hattman, S., Wilkinson, J., Swinton, D., Schlagman, S., MacDonald, P.M., Mosig, G.
<2>Common evolutionary origin of the Phage T4 dam and host Escherichia coli dam DNA-adenine methyltransferase genes.
<3>J. Bacteriol.
<4>164
<5>932-937
<6>1985
<7>We compared the known DNA nucleotide and encoded amino acid sequences of the Escherichia coli
and bacteriophage T4 dam (DNA-adenine methyltransferase) genes. Despite the absence of any DNA
sequence homology, there were four regions (11 to 33 residues long) of amino acid sequence
homology containing 45 to 64% identity. These results suggest that the genes for these two
enzymes have a common evolutionary origin.

<>

<1>Hattori, M. et al.
<2>The DNA sequence of human chromosome 21.
<3>Nature
<4>405
<5>311-319
<6>2000
<7>Chromosome 21 is the smallest human autosome. An extra copy of chromosome 21 causes Down
syndrome, the most frequent genetic cause of significant mental retardation, which affects up
to 1 in 700 live births. Several anonymous loci for monogenic disorders and predispositions
for common complex disorders have also been mapped to this chromosome, and loss of
heterozygosity has been observed in regions associated with solid tumours. Here we report the
sequence and gene catalogue of the long arm of chromosome 21. We have sequenced 33,546,361
base pairs (bp) of DNA with very high accuracy, the largest contig being 25,491,867 bp. Only
three small clone gaps and seven sequencing gaps remain, comprising about 100 kilobases. Thus,
we achieved 99.7% coverage of 21q. We also sequenced 281,116 bp from the short arm. The
structural features identified include duplications that are probably involved in chromosomal
abnormalities and repeat structures in the telomeric and pericentromeric regions. Analysis of
the chromosome revealed 127 known genes, 98 predicted genes and 59 pseudogenes.

<>

<1>Haugen, P., Bhattacharya, D.
<2>The spread of LAGLIDADG homing endonuclease genes in rDNA.
<3>Nucleic Acids Res.
<4>32
<5>2049-2057
<6>2004
<7>Group I introns that encode homing endonuclease genes (HEGs) are highly invasive genetic
elements. Their movement into a homologous position in
an intron-less allele is termed homing. Although the mechanism of
homing is well understood, the evolutionary relationship between HEGs
and their intron partners remains unclear. Here we have focused on the
largest family of HEGs (encoding the protein motif, LAGLIDADG) to
understand how HEGs and introns move in rDNA. Our analysis shows the
phylogenetic clustering of HEGs that encode a single copy of the
LAGLIDADG motif in neighboring, but often evolutionarily distantly
related, group I introns. These endonucleases appear to have inserted
into existing introns independent of ribozymes. In contrast, our data
support a common evolutionary history for a large family of
heterologous introns that encode HEGs with a duplicated LAGLIDADG
motif. This finding suggests that intron/double-motif HEG elements can
move into heterologous sites as a unit. Our data also suggest that a
subset of the double-motif HEGs in rDNA originated from the duplication
and fusion of a single-motif HEG encoded by present-day ribozymes in
LSU rDNA.

<>

<1>Haugen, P., De Jonckheere, J.F., Johansen, S.
<2>Characterization of the self-splicing products of two complex Naegleria LSU rDNA group I introns containing homing endonuclease genes.
<3>Eur. J. Biochem.
<4>269
<5>1641-1649
<6>2002
<7>The two group I introns Nae.L1926 and Nmo.L2563, found at two different sites in nuclear LSU
rRNA genes of Naegleria amoebo-flagellates, have been characterized in vitro. Their structural
organization is related to that of the mobile Physarum intron Ppo.L1925 (PpLSU3) with ORFs
extending the L1-loop of a typical group IC1 ribozyme. Nae.L1926, Nmo.L2563 and Ppo.L1925 RNAs
all self-splice in vitro, generating ligated exons and full-length intron circles as well as
internal processed excised intron RNAs. Formation of full-length intron circles is found to be
a general feature in RNA processing of ORF-containing nuclear group I introns. Both Naegleria
LSU rDNA introns contain a conserved polyadenylation signal at exactly the same position in
the 3' end of the ORFs close to the internal processing sites, indicating an RNA polymerase
II-like expression pathway of intron proteins in vivo. The intron proteins I-NaeI and I-NmoI
encoded by Nae.L1926 and Nmo.L2563, respectively, correspond to His-Cys homing endonucleases
of 148 and 175 amino acids. I-NaeI contains an additional sequence motif homologous to the
unusual DNA binding motif of three antiparallel beta sheets found in the I-PpoI endonuclease,
the product of the Ppo.L1925 intron ORF.

<>

<1>Haugen, P., Huss, V.A.R., Nielsen, H., Johansen, S.
<2>Complex group-I introns in nuclear SSU rDNA of red and green algae: Evidence of homing-endonuclease pseudogenes in the Bangiophyceae.
<3>Curr. Genet.
<4>36
<5>345-353
<6>1999
<7>The green alga Scenedesmus pupukensis and the red alga Porphyra spiralis contain large
group-IC1 introns in their nuclear small subunit
ribosomal RNA genes due to the presence of open reading frames at the
5' end of the introns. The putative 555 amino-acid Scenedesmus-encoded
protein harbors a sequence motif resembling the bacterial S9 ribosomal
proteins. The Porphyra intron self-splices in vitro, and generates both
ligated exons and a full-length intron RNA circle. The Porphyra intron
has an unusual structural organization by encoding a potential 149
amino-acid homing-endonuclease-like protein on the complementary
strand. A comparison between related group-I introns in the
Bangiophyceae revealed homing-endonuclease-like pseudogenes due to
frame-shifts and deletions in Porphyra and Bangia. The Scenedesmus and
Porphyra introns provide new insights into the evolution and possible
novel functions of nuclear group-I intron proteins.

<>

<1>Haugen, P., Simon, D.M., Bhattacharya, D.
<2>The natural history of group I introns.
<3>Trends Genet.
<4>21
<5>111-119
<6>2005
<7>There are four major classes of introns: self-splicing group I and group II introns, tRNA
and/or archaeal introns and spliceosomal introns
in nuclear pre-mRNA. Group I introns are widely distributed in
protists, bacteria and bacteriophages. Group II introns are found in
fungal and land plant mitochondria, algal plastids, bacteria and
Archaea. Group II and spliceosomal introns share a common splicing
pathway and might be related to each other. The tRNA and/or archaeal
introns are found in the nuclear tRNA of eukaryotes and in archaeal
tRNA, rRNA and mRNA. The mechanisms underlying the self-splicing and
mobility of a few model group I introns are well understood. By
contrast, the role of these highly distinct processes in the evolution
of the 1500 group I introns found thus far in nature (e.g. in algae and
fungi) has only recently been clarified. The explosion of new sequence
data has facilitated the use of comparative methods to understand group
I intron evolution in a broader context and to generate hypotheses
about intron insertion, splicing and spread that can be tested
experimentally.

<>

<1>Haugen, P., Wikmark, O.-G., Vader, A., Coucheron, D.H., Sjottem, E., Johansen, S.D.
<2>The recent transfer of a homing endonuclease gene.
<3>Nucleic Acids Res.
<4>33
<5>2734-2741
<6>2005
<7>The myxomycete Didymium iridis (isolate Panama 2) contains a mobile group I intron named
Dir.S956-1 after position 956 in the nuclear small subunit (SSU) rRNA gene. The intron is
efficiently spread through homing by the intron-encoded homing endonuclease I-DirI. Homing
endonuclease genes (HEGs) usually spread with their associated introns as a unit, but
infrequently also spread independent of introns (or inteins). Clear examples of HEG mobility
are however sparse. Here, we provide evidence for the transfer of a HEG into a group I intron
named Dir.S956-2 that is inserted into the SSU rDNA of the Costa Rica 8 isolate of D.iridis.
Similarities between intron sequences that flank the HEG and rDNA sequences that flank the
intron (the homing endonuclease recognition sequence) suggest that the HEG invaded the intron
during the recent evolution in a homing-like event. Dir.S956-2 is inserted into the same SSU
site as Dir.S956-1. Remarkably, the two group I introns encode distantly related splicing
ribozymes with phylogenetically related HEGs inserted on the opposite strands of different
peripheral loop regions. The HEGs are both interrupted by small spliceosomal introns that must
be removed during RNA maturation.

<>

<1>Hauglund, M.J., Tatum, F.M., Bayles, D.O., Maheswaran, S.K., Briggs, R.E.
<2>Genome Sequences of Mannheimia haemolytica Serotype A1 Strains D153 and D193 from Bovine Pneumonia.
<3>Genome Announcements
<4>1
<5>e00848-13
<6>2013
<7>Here we report two genome sequences, one complete and one draft, from virulent bovine strains
of Mannheimia haemolytica serotype A1 recovered prior to the field
usage of modern antimicrobial drugs.

<>

<1>Hauptmann, A.L., Glaring, M.A., Hallin, P.F., Prieme, A., Stougaard, P.
<2>Draft Genome Sequence of the Psychrophilic and Alkaliphilic Rhodonellum psychrophilum Strain GCM71T.
<3>Genome Announcements
<4>1
<5>e01014-13
<6>2013
<7>Rhodonellum psychrophilum GCM71(T), isolated from the cold and alkaline submarine ikaite
columns in the Ikka Fjord in Greenland, displays optimal growth at 5 to 10
degrees C and pH 10. Here, we report the draft genome sequence of this strain,
which may provide insight into the mechanisms of adaptation to these extreme
conditions.

<>

<1>Hauser, H., Richter, D.C., van Tonder, A., Clark, L., Preston, A.
<2>Comparative genomic analyses of the Taylorellae.
<3>Vet. Microbiol.
<4>159
<5>195-203
<6>2012
<7>Contagious equine metritis (CEM) is an important venereal disease of horses that
is of concern to the thoroughbred industry. Taylorella equigenitalis is a
causative agent of CEM but very little is known about it or its close relative
Taylorella asinigenitalis. To reveal novel information about Taylorella biology,
comparative genomic analyses were undertaken. Whole genome sequencing was
performed for the T. equigenitalis type strain, NCTC11184. Draft genome sequences
were produced for a second T. equigenitalis strain and for a strain of T.
asinigenitalis. These genome sequences were analysed and compared to each other
and the recently released genome sequence of T. equigenitalis MCE9. These
analyses revealed that T. equigenitalis strains appear to be very similar to each
other with relatively little strain-specific DNA content. A number of genes were
identified that encode putative toxins and adhesins that are possibly involved in
infection. Analysis of T. asinigenitalis revealed that it has a very similar gene
repertoire to that of T. equigenitalis but shares surprisingly little DNA
sequence identity with it. The generation of genome sequence information greatly
increases knowledge of these poorly characterised bacteria and greatly
facilitates study of them.

<>

<1>Hausmann, R., Gold, M.
<2>The enzymatic methylation of ribonucleic acid and deoxyribonucleic acid.
<3>J. Biol. Chem.
<4>241
<5>1985-1994
<6>1966
<7>Infection of Escherichia coli B with bacteriophages T1, T2, or T4 results in increase in the
activity of deoxyribonucleic acid methylase assayed in crude extracts of the infected cells.
The largest increase is observed after infection with T2 phage; T4 and T1 phages lead to
smaller increases in that order of decreasing magnitude.  After infection with T3, T5, or T6
phages, a decrease in activity was observed; T7 and lambda phages had no effect.  The kinetics
of the increase in methylase activity after T2 infection is similar to those found with the
"early enzymes" induced by T-even phages, and protein synthesis is necessary for the increase
to occur.  The properties of the methylase activity of phage T2-infected bacteria suggest that
it is a new phage-directed enzyme.

<>

<1>Hausner, G., Iranpour, M., Kim, J.-J., Breuil, C., Davis, C.N., Gibb, E.A., Reid, J., Loewen, P.C., Hopkin, A.A.
<2>Fungi vectored by the introduced bark beetle Tomicus piniperda in Ontario, Canada and comments on the taxonomy of Leptographium lundbergii, L. terebrantis, L. truncatum and L. wingfieldii.
<3>Can. J. Bot.
<4>83
<5>1222-1237
<6>2005
<7>Fungi isolated from Tomicus piniperda (L.) galleries in infected trap logs, standing trees,
and directly from insects were identified using morphological features and molecular data
obtained from the mitochondrial and nuclear DNA region. Identified strains represented
Leptographium wingfieldii Morelet, Leptographium procerum (Kendr.) Wingf., Leptographium
lundbergii Lag.

<>

<1>Hausner, G., Sethuraman, J., Edgell, D.
<2>Methods for producing homing endonucleases and their use in site-directed homologous recombination.
<3>US Patent Office
<4>US 20110256607 A
<5>
<6>2011
<7>The present disclosure provides, in part, polypeptides having endonuclease activity, nucleic
acid sequences for such a polypeptide, target sequences for the endonuclease, as well as
vectors, cells, kits, methods, and uses of the same.

<>

<1>Havelaar, K.J., Korsuize, J.
<2>Differences in susceptibility to restriction by E. coli B between various heteroduplex molecules of bacteriophage FD DNA.
<3>Mol. Biol. Rep.
<4>1
<5>453-455
<6>1974
<7>Fragments of B-modified bacteriophage fd sB1o sB2 RF DNA were prepared with the help of
purified endonuclease R from Haemophilus parainfluenzae.  These were hybridized with
unmodified circular single stranded fd DNA.  The resulting partial heteroduplex molecules were
assayed for infectivity on competent cells of B-restricting and non-restricting strains of E.
coli.  Three of such heteroduplexes originating from neighboring fragments on the physical map
of fd RF DNA were shown to be more resistant to EcoB restriction than six others and the
unmodified control.  It is suggested that the three corresponding vicinal fragments contain
essential parts of the EcoB recognition site on this phage DNA.

<>

<1>Havelsrud, O.E., Sorum, H., Gaustad, P.
<2>Genome Sequences of Corynebacterium pseudotuberculosis Strains 48252 (Human, Pneumonia), CS_10 (Lab Strain), Ft_2193/67 (Goat, Pus), and CCUG 27541.
<3>Genome Announcements
<4>2
<5>e00869-14
<6>2014
<7>Here we report the genome sequencess of four Corynebacterium pseudotuberculosis strains. These
include a strain isolated from a patient with C.
pseudotuberculosis pneumonia (48252), a strain isolated from pus in goat
(Ft_2193/67), a laboratory strain originating from strain Ft_2193/67 (CS_10), and
the draft genome of an equine reference strain, CCUG 27541.

<>

<1>Hawkey, J., Edwards, D.J., Dimovski, K., Hiley, L., Billman-Jacobe, H., Hogg, G., Holt, K.E.
<2>Evidence of microevolution of Salmonella Typhimurium during a series of egg-associated outbreaks linked to a single chicken farm.
<3>BMC Genomics
<4>14
<5>800
<6>2013
<7>BACKGROUND: The bacterium Salmonella enterica serovar Typhimurium (S.
Typhimurium) is one of the most frequent causes of foodborne outbreaks of
gastroenteritis. Between 2005-2008 a series of S. Typhimurium outbreaks occurred
in Tasmania, Australia, that were all traced to eggs originating from a single
chicken farm. We sequenced the genomes of 12 isolates linked to these outbreaks,
in order to investigate the microevolution of a pathogenic S. Typhimurium clone
in a natural, spatiotemporally restricted population. RESULTS: The isolates,
which shared a phage type similar to DT135 known locally as 135@ or 135a, formed
a clade within the S. Typhimurium population with close similarity to the
reference genome SL1334 (160 single nucleotide polymorphisms, or SNPs). Ten of
the isolates belonged to a single clone (<23 SNPs between isolate pairs) which
likely represents the population of S. Typhimurium circulating at the chicken
farm; the other two were from sporadic cases and were genetically distinct from
this clone. Divergence dating indicated that all 12 isolates diverged from a
common ancestor in the mid 1990 s, and the clone began to diversify in 2003-2004.
This clone spilled out into the human population several times between 2005-2008,
during which time it continued to accumulate SNPs at a constant rate of 3-5 SNPs
per year or 1x10-6 substitutions site-1 year-1, faster than the longer-term (~50
year) rates estimated previously for S. Typhimurium. Our data suggest that
roughly half of non-synonymous substitutions are rapidly removed from the S.
Typhimurium population, after which purifying selection is no longer important
and the remaining substitutions become fixed in the population. The S.
Typhimurium 135@ isolates were nearly identical to SL1344 in terms of gene
content and virulence plasmids. Their phage contents were close to SL1344, except
that they carried a different variant of Gifsy-1, lacked the P2 remnant found in
SL1344 and carried a novel P2 phage, P2-Hawk, in place SL1344's P2 phage
SopEvarphi. DT135 lacks P2 prophage. Two additional plasmids were identified in
the S. Typhimurium 135@ isolates, pSTM2 and pSTM7. Both plasmids were IncI1, but
phylogenetic analysis of the plasmids and their bacterial hosts shows these
plasmids are genetically distinct and result from independent plasmid acquisition
events. CONCLUSIONS: This study provides a high-resolution insight into
short-term microevolution of the important human pathogen S. Typhimurium. It
indicates that purifying selection occurs rapidly in this population (</= 6
years) and then declines, and provides an estimate for the short-term
substitution rate. The latter is likely to be more relevant for foodborne
outbreak investigation than previous estimates based on longer time scales.

<>

<1>Hawtrey, A.O., Ariatti, M.
<2>A possible model for the methylation of deoxycytidine in DNA.
<3>Med. Hypotheses
<4>15
<5>125-134
<6>1984
<7>The modified base 5-methylcytidine has been found in the DNA of a number of
different eukaryotic cells where it occurs principally in the dinucleotide
sequence -CmpG- which is present as a palindrome in double-strand nucleic acid
molecules.  There is considerable evidence to indicate and suggest that
5-methylcytosine serves as a regulatory signal in eukaryotic gene expression.
Replication of DNA containing -CmpG- gives rise to daughter DNA molecules
containing new -CpG- dinucleotide sequences in which the cytidine residues are
not methylated.  Methylation of these residues is carried out by a methylase
enzyme using S-adenosyl-L-methionine as a specific methyl group donor.  This
model discussed in the present communication tries to explain in chemical and
biological terms the mechanism of the methylation reaction.  The first
reactions of the scheme are well known through the work of other investigators.
However, we introduce a new concept into our reaction mechanism by postulating
the direct involvement of S-adenosyl-L-methionine in the reaction through its
covalent attachment to the cytosine ring followed by a specific ring closure
and methylation involving transfer of a hydride ion.  The model also gives a
possible explanation of mechanism of interaction of dimethyl sulphoxide with
the enzyme systems of certain eukaryotic cells, which are altered or changed in
the regulation of gene expression by this chemical reagent.

<>

<1>Hayakawa, T., Ono, A., Ueda, T.
<2>Synthesis of decadeoxyribonucleotides containing 5-modified uracils and their interactions with restriction endonucleases BglII, Sau3AI and MboI (Nucleosides and Nucleotides 82).
<3>Nucleic Acids Res.
<4>16
<5>4761-4776
<6>1988
<7>Decadeoxyribonucleotides containing uracil, 5-bromouracil, 5-cyanouracil and 5-ethyluracil in
recognition sequences of restriction endonucleases BglII, Sau3AI, MboI were synthesized.
Decanucleotides containing 5-bromouracil in place of thymine had essentially the same
susceptibility to all the restriction endonucleases.  Uracil-containing decanucleotides were
however very resistant to attack.  Decanucleotides containing 5-cyanouracil in the recognition
sequence were strongly resistant to hydrolysis by Sau3AI, but were hydrolysed by BglII and
MboI as well as the parent decanucleotide.  Decanucleotides containing 5-ethyluracil were
strongly resistant to hydrolysis by Sau3AI, but were partially resistant to hydrolysis by
BglII and MboI.

<>

<1>Hayano-Kanashiro, C., Lopez-Arredondo, D.L., Cruz-Morales, P., Alcaraz, L.D., Olmedo, G., Barona-Gomez, F., Herrera-Estrella, L.
<2>First draft genome sequence of a strain from the genus citricoccus.
<3>J. Bacteriol.
<4>193
<5>6092-6093
<6>2011
<7>Bacteria of the genus Citricoccus have been isolated from ecological niches characterized by
diverse abiotic stress conditions. Here we report
the first genome draft of a strain of the genus Citricoccus isolated from
the extremely oligotrophic Churince system in the Cuatro Cienegas Basin
(CCB) in Coahuila, Mexico.

<>

<1>Hayashi, H., Shinagawa, H., Makino, K., Hayashi, T., Onishi, S., Hattori, M., Kurokawa, K.
<2>Nucleic acid molecule and polypeptide specific to intestinal hemorrhagic pathogenic Escherichia coli O157:H7, and method of use.
<3>Japanese Patent Office
<4>JP 2002355074 A
<5>
<6>2002
<7>
<>

<1>Hayashi, K., Morooka, N., Yamamoto, Y., Fujita, K., Isono, K., Choi, S., Ohtsubo, E., Baba, T., Wanner, B.L., Mori, H., Horiuchi, T.
<2>Highly accurate genome sequences of Escherichia coli K-12 strains MG1655 and W3110.
<3>Mol. Syst. Biol.
<4>2
<5>2006.0007
<6>2006
<7>With the goal of solving the whole-cell problem with Escherichia coli K-12 as a model cell,
highly accurate genomes were determined for two closely related K-12 strains, MG1655 and
W3110. Completion of the W3110 genome and comparison with the MG1655 genome revealed
differences at 267 sites, including 251 sites with short, mostly single-nucleotide, insertions
or deletions (indels) or base substitutions (totaling 358 nucleotides), in addition to 13
sites with an insertion sequence element or defective prophage in only one strain and two
sites for the W3110 inversion. Direct DNA sequencing of PCR products for the 251 regions with
short indel and base disparities revealed that only eight sites are true differences. The
other 243 discrepancies were due to errors in the original MG1655 sequence, including 79
frameshifts, one amino-acid residue deletion, five amino-acid residue insertions, 73 missense,
and 17 silent changes within coding regions. Errors in the original MG1655 sequence (<1 per
13,000 bases) were mostly within portions sequenced with out-dated technology based on
radioactive chemistry.

<>

<1>Hayashi, T. et al.
<2>Complete genome sequence of Enterohemorrhagic Escherichia coli O157:H7 and genomic comparison with a laboratory strain K-12.
<3>DNA Res.
<4>8
<5>11-22
<6>2001
<7>Escherichia coli O157:H7 is a major food-borne infectious pathogen that causes diarrhea,
hemorrhagic colitis, and hemolytic uremic syndrome. Here we report the complete chromosome
sequence of an O157:H7 strain isolated from the Sakai outbreak, and the results of genomic
comparison with a benign laboratory strain, K-12 MG1655. The chromosome is 5.5 Mb in size, 859
Kb larger than that of K-12. We identified a 4.1-Mb sequence highly conserved between the two
strains, which may represent the fundamental backbone of the E. coli chromosome. The remaining
1.4-Mb sequence comprises of O157:H7-specific sequences, most of which are horizontally
transferred foreign DNAs. The predominant roles of bacteriophages in the emergence of O157:H7
is evident by the presence of 24 prophages and prophage-like elements that occupy more than
half of the O157:H7-specific sequences. The O157:H7 chromosome encodes 1632 proteins and 20
tRNAs that are not present in K-12. Among these, at least 131 proteins are assumed to have
virulence-related functions.  Genome-wide codon usage analysis suggested that the
O157:H7-specific tRNAs are involved in the efficient expression of the strain-specific genes.
A complete set of the genes specific to O157:H7 presented here sheds new insight into the
pathogenicity and the physiology of O157:H7, and will open a way to fully understand the
molecular mechanisms underlying the O157:H7 infection.

<>

<1>Hayden, H.S., Gillett, W., Saenphimmachak, C., Lim, R., Zhou, Y., Jacobs, M.A., Chang, J., Rohmer, L., D'Argenio, D.A., Palmieri, A., Levy, R., Haugen, E., Wong, G.K., Brittnacher, M.J., Burns, J.L., Miller, S.I., Olson, M.V., Kaul, R.
<2>Large-insert genome analysis technology detects structural variation in Pseudomonas aeruginosa clinical strains from cystic fibrosis patients.
<3>Genomics
<4>91
<5>530-537
<6>2008
<7>Large-insert genome analysis (LIGAN) is a broadly applicable,
high-throughput technology designed to characterize genome-scale
structural variation. Fosmid paired-end sequences and DNA fingerprints
from a query genome are compared to a reference sequence using the Genomic
Variation Analysis (GenVal) suite of software tools to pinpoint locations
of insertions, deletions, and rearrangements. Fosmids spanning regions
that contain new structural variants can then be sequenced. Clonal pairs
of Pseudomonas aeruginosa isolates from four cystic fibrosis patients were
used to validate the LIGAN technology. Approximately 1.5 Mb of inserted
sequences were identified, including 743 kb containing 615 ORFs that are
absent from published P. aeruginosa genomes. Six rearrangement breakpoints
and 220 kb of deleted sequences were also identified. Our study expands
the "genome universe" of P. aeruginosa and validates a technology that
complements emerging, short-read sequencing methods that are better suited
to characterizing single-nucleotide polymorphisms than structural
variation.

<>

<1>Hayward, G.S., Frenkel, N., Roizman, B.
<2>Anatomy of herpes simplex virus DNA: Strain differences and heterogeneity in the locations of restriction endonuclease cleavage sites.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>72
<5>1768-1772
<6>1975
<7>Digestion of herpes simplex virus DNA by the HindII or EcoRI restriction
endonucleases yielded 11 to 15 fragments with molecular weights between 2 and
28 million.  The electrophoretic profiles obtained in 0.3% agarose gels with
DNA fragments from nine different strains of herpes simplex virus type 1 could
be readily differentiated from the patterns exhibited by the corresponding
fragments from four separate strains of type 2 virus; however, within each
serotype, the laboratory strains differed significantly among themselves and
also from isolates passaged a minimum number of times outside the human host.
Digestion of all DNAs of herpes simplex virus with either enzyme reproducibly
generated two classes of fragments (major and minor) which differed in molar
concentration.  Moreover, although the molecular weight of an intact herpes
simplex 1 (F1) DNA molecule is approximately 98 Md, the summed molecular
weights of all major and minor HindIII fragments totalled 160 Md, and the seven
major fragments alone accounted for only 60 Md.  These unusual features
indicate the existence of limited heterogeneity in the positions of cleavage
sites along individual molecules.  We have eliminated the possibility that
minor fragments arose from contamination with the defective DNA of high buoyant
density which appears on serial undiluted passage of the virus.  In fact, this
latter type of DNA was resistant to cleavage by HindIII and gave large amounts
of only two species of EcoRI fragments, suggesting that the defective molecules
consist of many tandem repeats of a small segment of viral DNA.  The
heterogeneity in the viral DNA of normal density appears to be related to the
structural organization of the molecules and does not necessarily imply
differences in genetic content.

<>

<1>Hazen, T.H., Humphrys, M.S., Ochieng, J.B., Parsons, M., Bopp, C.A., O'Reilly, C.E., Mintz, E., Rasko, D.A.
<2>Draft Genome Sequences of Nine Enteropathogenic Escherichia coli Strains from Kenya.
<3>Genome Announcements
<4>2
<5>e00582-14
<6>2014
<7>We report here the draft genome sequences of nine enteropathogenic Escherichia coli (EPEC)
strains isolated from children in Kenya who died during
hospitalization with diarrhea. Each of the isolates possess the EPEC adherence
factor (EAF) plasmid encoding the bundle-forming pilus, which is characteristic
of EPEC. These isolates represent diverse serogroups and EPEC phylogenomic
lineages.

<>

<1>Hazen, T.H., Robinson, G.L., Harris, A.D., Rasko, D.A., Johnson, J.K.
<2>Genome Sequence of Klebsiella oxytoca 11492-1, a Nosocomial Isolate Possessing a  FOX-5 AmpC beta-Lactamase.
<3>J. Bacteriol.
<4>194
<5>3028-3029
<6>2012
<7>Klebsiella oxytoca strain 11492-1 was isolated from a perianal swab culture from  a patient at
the University of Maryland Medical Center in 2005. The K. oxytoca 11492-1 draft genome
contains multiple antibiotic resistance genes, including a FOX-5 AmpC beta-lactamase encoded
on a large IncA/C plasmid.

<>

<1>Hazen, T.H., Sahl, J.W., Fraser, C.M., Donnenberg, M.S., Scheutz, F., Rasko, D.A.
<2>Draft Genome Sequences of Three O157 Enteropathogenic Escherichia coli Isolates.
<3>Genome Announcements
<4>1
<5>e00516-13
<6>2013
<7>We report the draft genome sequences of three enteropathogenic Escherichia coli (EPEC)
isolates that display the O157 serogroup but do not have the Shiga toxin
genes (stx), which are characteristic of O157 enterohemorrhagic E. coli (EHEC).
E. coli strain RN587/1 has the O157:H8 serotype and possesses the EAF plasmid
characteristic of typical EPEC (J. B. Kaper, J. P. Nataro, and H. L. Mobley, Nat.
Rev. Microbiol. 2:123-140, 2004). The other two isolates, strains C844-97 and
C639-08, are both O157:H45 and possess the locus of enterocyte effacement (LEE)
pathogenicity island; however, they do not contain the EAF plasmid or the
stx-carrying phage.

<>

<1>Hazen, T.H., Sahl, J.W., Redman, J.C., Morris, C.R., Daugherty, S.C., Chibucos, M.C., Sengamalay, N.A., Fraser-Liggett, C.M., Steinsland, H., Whittam, T.S., Whittam, B., Manning, S.D., Rasko, D.A.
<2>Draft Genome Sequences of the Diarrheagenic Escherichia coli Collection.
<3>J. Bacteriol.
<4>194
<5>3026-3027
<6>2012
<7>We report the draft genome sequences of the collection referred to as the Escherichia coli
DECA collection, which was assembled to contain representative isolates of the 15 most common
diarrheagenic clones in humans
(http://shigatox.net/new/). These genomes represent a valuable resource to the community of
researchers who examine these enteric pathogens.

<>

<1>Hazen, T.H., Wu, D., Eisen, J.A., Sobecky, P.A.
<2>Sequence Characterization and Comparative Analysis of Three Plasmids Isolated from Environmental Vibrio spp.
<3>Appl. Environ. Microbiol.
<4>73
<5>7703-7710
<6>2007
<7>The horizontal transfer of genes by mobile genetic elements such as
plasmids and phages can accelerate genome diversification of Vibrio spp.,
affecting their physiology, pathogenicity, and ecological character. In
this study, sequence analysis of three plasmids from Vibrio spp.
previously isolated from salt marsh sediment revealed the remarkable
diversity of these elements. Plasmids p0908 (81.4 kb), p23023 (52.5 kb),
and p09022 (31.0 kb) had a predicted 99, 64, and 32 protein-coding
sequences and G+C contents of 49.2%, 44.7%, and 42.4%, respectively. A
phylogenetic tree based on concatenation of the host 16S rRNA and rpoA
nucleotide sequences indicated p23023 and p09022 were isolated from
strains most closely related to V. mediterranei and V. campbellii,
respectively, while the host of p0908 forms a clade with V. fluvialis and
V. furnissii. Many predicted proteins had amino acid identities to
proteins of previously characterized phages and plasmids (24 to 94%).
Predicted proteins with similarity to chromosomally encoded proteins
included RecA, a nucleoid-associated protein (NdpA), a type IV helicase
(UvrD), and multiple hypothetical proteins. Plasmid p0908 had striking
similarity to enterobacteria phage P1, sharing genetic organization and
amino acid identity for 23 predicted proteins. This study provides
evidence of genetic exchange between Vibrio plasmids, phages, and
chromosomes among diverse Vibrio spp.

<>

<1>He, H., Deng, M., He, J., Weng, S.
<2>Structure and sequence analysis of cytosine DNA methyltransferase gene form infectious spleen and kidney necrosis virus.
<3>Bingdu Xuebao
<4>17
<5>349-355
<6>2001
<7>In the infectious spleen and kidney necrosis virus genome, an iridovirus, we have identified
an open reading frame whose deduced amino acid sequence contains motifs characteristic of
cytosine DNA methyltransferases.  The ORF consists of 684bp which codes for a protein of 227
aa with a predicted molecular mass of 25,855 Da. Compared with some MTases found in bacteria,
ISKNV MTase ORF contains the first four highly conserved  motifs of cytosine MTase, but the
motif, responsible for DNA binding specificity, is missing.  Compared with 6 vertebrate
iridoviruses, ISKNV is considered as a new group of Iridoviridae.  Partial sequences of 7
vertebrate iridovirus MTases are highly conserved, they can be used to design primers to
identify vertebrate iridovirus by PCR method.

<>

<1>He, J., Shao, X., Zheng, H., Li, M., Wang, S., Zhang, Q., Li, L., Liu, Z., Sun, M., Wang, S., Yu, Z.
<2>The Complete Genome Sequence of Bacillus thuringiensis Mutant Strain BMB171.
<3>J. Bacteriol.
<4>192
<5>4074-4075
<6>2010
<7>Bacillus thuringiensis is widely used as biopesticide for a long time. Here we report the
finished and annotated genome sequence of B.
thuringiensis mutant strain BMB171, a crystalliferous mutant strain
obtained and stocked in our laboratory, with a high transformation
frequency.

<>

<1>He, J., Wang, J., Yin, W., Shao, X., Zheng, H., Li, M., Zhao, Y., Sun, M., Wang, S., Yu, Z.
<2>Complete Genome Sequence of Bacillus thuringiensis subsp. chinensis strain CT-43.
<3>J. Bacteriol.
<4>193
<5>3407-3408
<6>2011
<7>Bacillus thuringiensis is widely used as agricultural biopesticide for a long time. As a
producing strain, B. thuringiensis subsp. chinensis strain
CT-43 has high toxin to lepidopterous and dipterous insects. It can form
various parasporal crystals consisting of Cry1Aa3, Cry1Ba1, Cry1Ia14,
Cry2Aa9, and Cry2Ab1. During fermentation, it simultaneously generates
vegetative insecticidal protein Vip3Aa10, as well as insecticidal
nucleotide analogue thuringiensin. Here we report the finished, annotated
genome sequence of B. thuringiensis strain CT-43.

<>

<1>He, J.G., Deng, M., Weng, S.P., Li, Z., Zhou, S.Y., Long, Q.X., Wang, X.Z., Chan, S.M.
<2>Complete genome analysis of the mandarin fish infectious spleen and kidney necrosis iridovirus.
<3>Virology
<4>291
<5>126-139
<6>2001
<7>The nucleotide sequence of the infectious spleen and kidney necrosis virus
(ISKNV) genome was determined and found to comprise 111,362 bp with a G+C
content of 54.78%. It contained 124 potential open reading frames (ORFs)
with coding capacities ranging from 40 to 1208 amino acids. The analysis
of the amino acid sequences deduced from the individual ORFs revealed that
35 of the 124 potential gene products of ISKNV show significant homology
to functionally characterized proteins of other species. Some of the
putative gene products of ISKNV showed significant homologies to proteins
in the GenBank/EMBL/DDBJ databases including enzymes and structural
proteins involved in virus replication, transcription, protein
modification, and virus-host interaction. In addition, one major repeated
sequence showing significant homology to the Red Sea bream iridovirus
(RSIV) genome was identified. Based on the information obtained from
biological properties (including histopathology, tissue tropisms, natural
host range, and geographic distribution), physiochemical and physical
properties, and genome analysis, we suggest that ISKNV, RSIV, sea bass
iridovirus, grouper iridovirus, and African lampeye iridovirus may belong
to a new genus of the Iridoviridae family and are tentatively referred to
as cell hypertrophy iridoviruses.

<>

<1>He, M. et al.
<2>Evolutionary dynamics of Clostridium difficile over short and long time scales.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>7527-7532
<6>2010
<7>Clostridium difficile has rapidly emerged as the leading cause of antibiotic-associated
diarrheal disease, with the transcontinental spread of
various PCR ribotypes, including 001, 017, 027 and 078. However, the genetic
basis for the emergence of C. difficile as a human pathogen is unclear. Whole
genome sequencing was used to analyze genetic variation and virulence of a
diverse collection of thirty C. difficile isolates, to determine both macro and
microevolution of the species. Horizontal gene transfer and large-scale
recombination of core genes has shaped the C. difficile genome over both short
and long time scales. Phylogenetic analysis demonstrates C. difficile is a
genetically diverse species, which has evolved within the last 1.1-85 million
years. By contrast, the disease-causing isolates have arisen from multiple
lineages, suggesting that virulence evolved independently in the highly epidemic
lineages.

<>

<1>He, P., Hao, K., Blom, J., Rueckert, C., Vater, J., Mao, Z., Wu, Y., Hou, M., He, P., He, Y., Borriss, R.
<2>Genome sequence of the plant growth promoting strain Bacillus amyloliquefaciens subsp plantarum B9601-Y2 and expression of mersacidin and other secondary metabolites.
<3>J. Biotechnol.
<4>164
<5>281-291
<6>2012
<7>The plant-associated Bacillus amyloliquefaciens subsp. plantarum strain B9601-Y2, isolated
from wheat rhizosphere, is a powerful plant
growth-promoting rhizobacterium. Its relative large genome size of 4.24
Mbp, exceeding that of other representatives of the B.
amyloliquefaciens subsp. plantarum taxon, is mainly due to the presence
of 18 DNA-islands containing remnants of phages, a unique restriction
modification system, a gene cluster for mersacidin synthesis, and an
orphan gene cluster devoted to non-ribosomal synthesis of an
unidentified peptide. Like other members of the taxon, the Y2 genome
contains giant gene clusters for non-ribosomal synthesis of the
polyketides macrolactin, difficidin, and bacillaene, the antifungal
lipopeptides bacillomycin D, and fengycin, the siderophore
bacillibactin, and the dipeptide bacilysin. A gene cluster encoding
enzymes for a degradative pathway with 2-keto-3-deoxygluconate and
2-keto-3-deoxy-phosphogluconate as intermediates was explored by genome
mining and found as being a unique feature for representatives of the
plantarum subspecies. A survey of the Y2 genome against other B.
amyloliquefaciens genomes revealed 130 genes only occurring in subsp.
plantarum but not in subsp. amyloliquefaciens. Notably, the surfactin
gene cluster is not functional due to a large deletion removing parts
of the Srf synthetases B and C. Expression of polyketides,
lipopeptides, mersacidin, and of the growth hormone indole-3-acetic
acid in Y2 was demonstrated by matrix-assisted laser desorption
ionization-time of flight mass spectroscopy and high-performance liquid
chromatography, respectively.

<>

<1>He, X., Ou, H.Y., Yu, Q., Zhou, X., Wu, J., Liang, J., Zhang, W., Rajakumar, K., Deng, Z.
<2>Analysis of a genomic island housing genes for DNA S-modification system in Streptomyces lividans 66 and its counterparts in other distantly related  bacteria.
<3>Mol. Microbiol.
<4>65
<5>1034-1048
<6>2007
<7>The complete sequence (92 770 bp) of a genomic island (GI) named SLG from Streptomyces
lividans 66, encoding a novel DNA S-modification system (dnd), was
determined. Its overall G+C content was 67.8%, lower than those of three
sequenced Streptomyces genomes. Among 85 predicted open reading frames (ORFs) in
SLG, 22 ORFs showed little homology with previously known proteins. SLG displays
a mosaic structure composed of four modules, indicative of multiple recombination
events in its formation. Spontaneous excision and circularization of SLG was
observed, and the excision rate appeared to be induced at least fivefold by MNNG
exposure. Using constructed mini-islands of SLG, we demonstrated that Slg01, a
P4-like integrase, was sufficient to promote SLG integration, excision and
circularization. Eleven counterpart dnd clusters, which also mapped to GIs in 10
chromosomes and a plasmid, were found in taxonomically unrelated bacterial
species from various geographic niches. Additionally, c. 10% of actinomycetes
were found to possess a dnd cluster in a survey involving 74 strains. Comparison
of dnd clusters in the 12 bacteria strongly suggests that these dnd-bearing
elements might have evolved from a common ancestor similar to plasmid-originated
chromosome II of Pseudoalteromonas haloplanktis TAC125.

<>

<1>He, Z., Crist, M., Yen, H.-C., Duan, X., Quiocho, F.A., Gimble, F.S.
<2>Amino acid residues in both the protein splicing and endonuclease domains of the PI-SceI intein mediate DNA binding.
<3>J. Biol. Chem.
<4>273
<5>4607-4615
<6>1998
<7>A structure-based model describing the interaction of the two-domain PI-SceI endonuclease with
its 31-base pair DNA substrate suggests that the endonuclease domain (domain II) contacts the
cleavage site region of the substrate, while the protein splicing domain (domain I) interacts
with a distal region that is sufficient for high affinity binding.  To support this model,
alanine-scanning mutagenesis was used to assemble a set of 49 PI-SceI mutant proteins that
were purified and assayed for their DNA binding and cleavage properties.  Fourteen mutant
proteins were 4- to >500-fold less active than wild-type PI-SceI in cleavage assays, and one
mutant (T225A) was 3-fold more active.  Alanine substitution at two positions in domain I
reduces overall binding >60-fold by perturbing the interaction of PI-SceI with the minimal
binding region.  Conversely, mutations in domain II have little effect on binding, reduce
binding to the cleavage site region only, or affect binding to both regions.  Interestingly,
substitutions at Lys301, which is part of the endonucleolytic active site, eliminate binding
to the cleavage site region but permit contact with the minimal binding region.  This
experimental evidence demonstrates that the protein splicing domain as well as the
endonuclease domain is involved in binding of a DNA substrate with the requisite length.

<>

<1>He, Z., Jiang, X., Xue, J., Zheng, W.
<2>Purification of BamHI DNA methylase and protection function of DNA methylation against restriction endonuclease BamHI.
<3>Weishengwuxue Tongbao
<4>16
<5>217-220
<6>1989
<7>None

<>

<1>Heard, E., Fried, M.
<2>The use of 5-azacytidine to increase cleavage of methylation sensitive rare cutting restriction enzymes sites in amplified DNA.
<3>Nucleic Acids Res.
<4>18
<5>6147-6148
<6>1990
<7>None

<>

<1>Heath, P.J., Stephens, K.M., Monnat, R.J. Jr., Stoddard, B.L.
<2>The structure of I-Crel, a group I intron-encoded homing endonuclease.
<3>Nat. Struct. Biol.
<4>4
<5>468-476
<6>1997
<7>The structure of I-Crel provides the first view of a protein encoded by a gene within an
intron. This endonuclease recognizes a long DNA site approximately 20 base pairs in length and
facilitates the lateral transfer of that intron. The protein exhibits a DNA-binding surface
consisting of four antiparallel beta-strands that form a 20 A wide groove which is over 70 A
long. The architecture of this fold is different from that of the TATA binding protein, TBP,
which also contains an antiparallel beta-saddle. The conserved LAGLIDADG motif, which is found
in many mobile intron endonucleases, maturases and inteins, forms a novel helical interface
and contributes essential residues to the active site.

<>

<1>Heather, Z., Holden, M.T., Steward, K.F., Parkhill, J., Song, L., Challis, G.L., Robinson, C., Davis-Poynter, N., Waller, A.S.
<2>A novel streptococcal integrative conjugative element involved in iron acquisition.
<3>Mol. Microbiol.
<4>70
<5>1274-1292
<6>2008
<7>In this study, we determined the function of a novel non-ribosomal peptide
synthetase (NRPS) system carried by a streptococcal integrative
conjugative element (ICE), ICESe2. The NRPS shares similarity with the
yersiniabactin system found in the high-pathogenicity island of Yersinia
sp. and is the first of its kind to be identified in streptococci. We
named the NRPS product 'equibactin' and genes of this locus eqbA-N.
ICESe2, although absolutely conserved in Streptococcus equi, the causative
agent of equine strangles, was absent from all strains of the closely
related opportunistic pathogen Streptococcus zooepidemicus. Binding of
EqbA, a DtxR-like regulator, to the eqbB promoter was increased in the
presence of cations. Deletion of eqbA resulted in a small-colony
phenotype. Further deletion of the irp2 homologue eqbE, or the genes eqbH,
eqbI and eqbJ encoding a putative ABC transporter, or addition of the iron
chelator nitrilotriacetate, reversed this phenotype, implicating iron
toxicity. Quantification of (55)Fe accumulation and sensitivity to
streptonigrin suggested that equibactin is secreted by S. equi and that
the eqbH, eqbI and eqbJ genes are required for its associated iron import.
In agreement with a structure-based model of equibactin synthesis,
supplementation of chemically defined media with salicylate was required
for equibactin production.

<>

<1>Heavens, D., Tailford, L.E., Crossman, L., Jeffers, F., Mackenzie, D.A., Caccamo, M., Juge, N.
<2>Genome sequence of the vertebrate gut symbiont Lactobacillus reuteri ATCC 53608.
<3>J. Bacteriol.
<4>193
<5>4015-4016
<6>2011
<7>Lactobacillus reuteri inhabiting the gastrointestinal tract of a range of vertebrates is a
true symbiont with established beneficial effects to the host. Here we describe the draft
genome of L. reuteri ATCC 53608 strain isolated from pig. The genome sequence provides
important insights into the evolutionary changes underlying host specialisation.

<>

<1>Hebert, E.M., Raya, R.R., Brown, L., Font-de-Valdez, G., Savoy-de-Giori, G., Taranto, M.P.
<2>Genome Sequence of the Cheese-Starter Strain Lactobacillus delbrueckii subsp. lactis CRL 581.
<3>Genome Announcements
<4>1
<5>e00602-13
<6>2013
<7>We report the genome sequence of Lactobacillus delbrueckii subsp. lactis CRL 581  (1,911,137
bp, GC 49.7%), a proteolytic strain isolated from a homemade
Argentinian hard cheese which has a key role in bacterial nutrition and releases
bioactive health-beneficial peptides from milk proteins.

<>

<1>Hebert, E.M., Saavedra, L., Taranto, M.P., Mozzi, F., Magni, C., Nader, M.E., Font-de-Valdez, G., Sesma, F., Vignolo, G., Raya, R.R.
<2>Genome Sequence of the Bacteriocin-Producing Lactobacillus curvatus Strain CRL705.
<3>J. Bacteriol.
<4>194
<5>538-539
<6>2012
<7>Lactobacillus curvatus is one of the most prevalent lactic acid bacteria found in fermented
meat products. Here, we present the draft genome
sequence of Lactobacillus curvatus CRL705, a bacteriocin producer strain
isolated from an Argentinean artisanal fermented sausage, which consists
of 1,833,251 bp (GC content, 41.9%) and two circular plasmids of 12,342 bp
(pRC12; GC, 43.9%) and 18,664 bp (pRC18; GC, 34.4%).

<>

<1>Hebert, L., Moumen, B., Duquesne, F., Breuil, M.F., Laugier, C., Batto, J.M., Renault, P., Petry, S.
<2>Genome sequence of Taylorella equigenitalis MCE9, the causative agent of contagious equine metritis.
<3>J. Bacteriol.
<4>193
<5>1785
<6>2011
<7>Taylorella equigenitalis is the causative agent of contagious equine metritis (CEM), a
sexually-transmitted infection of horses. We herein report the genome sequence of T.
equigenitalis strain MCE9, isolated in 2005 from the urethral fossa of a 4-year-old stallion
in France.

<>

<1>Hebert, L., Touzain, F., de Boisseson, C., Breuil, M.F., Duquesne, F., Laugier, C., Blanchard, Y., Petry, S.
<2>Draft Genome Sequence of Taylorella equigenitalis Strain MCE529, Isolated from a  Belgian Warmblood Horse.
<3>Genome Announcements
<4>2
<5>e01214-14
<6>2014
<7>Taylorella equigenitalis is the causative agent of contagious equine metritis (CEM), a
sexually transmitted infection of horses. We herein report the genome
sequence of T. equigenitalis strain MCE529, isolated in 2009 from the urethral
fossa of a 15-year-old Belgian Warmblood horse in France.

<>

<1>Hedges, R.W.
<2>Phenotypic characterization of fi- R factors determining the restriction and modification hspII specificity.
<3>Mol. Gen. Genet.
<4>115
<5>225-233
<6>1972
<7>All known determinants of the restriction, modification specificity hspII are
plasmids of the compatibility class N.  Two I-like R factors, R56 and R64 are
able to interfere with the lytic cycle of phage lambda (in liberation of phage
by lysogens or newly infected cells) by a different mechanism.

<>

<1>Hedges, R.W., Datta, N.
<2>R124, an fi+ R factor of a new compatibility class.
<3>J. Gen. Microbiol.
<4>71
<5>403-405
<6>1972
<7>The plasmids of Gram-negative bacteria can be classified as fi+ or fi-
(Watanabe et al. 1964).  The former are capable of repressing the fertility
functions, notably the production of sex pili, determined by the F factor
(Nishimura, Ishibashi, Meynell & Hirota, 1967).  Most fi+ plasmids carry genes
determining the production of sex pili similar to those determined by the F
factor (F-like pili) in antigenic specificity and phage adsorption (Meynell &
Datta, 1966; Lawn & Meynell, 1970; Dennison & Hedges, 1972).  Among the fi-
plasmids a number of compatibility classes has been described (Watanabe, 1968;
Datta & Hedges, 1971; Datta et al. 1971; Hedges & Datta, 1971).  Two plasmids
belonging to the same compatibility class cannot stably co-exisit in a single
cell.  Among the fi+ plasmids four compatibility classes have been described,
three of which we propose to name as follows:  FI, the class including F,
ColV2, ColV3 (MacFarren & Clowes, 1967) and R386 (Dennison, 1972); FII, the
class including RI and many other fi+ R factors (Meynell, Meynell & Datta,
1968); FIII, the class including ColB-K98 and ColB-K166 (Frydman & Meynell,
1969); fourthly, the class including R62 (R62, though fi+, determines I-like
pili and has the compatibility of a typical I-like plasmid) (lawn, Meynell,
Meynell & Datta, 1967; N. Datta & R.W. Hedges, unpublished).  R124 is an fi+ R
factor carrying tetracycline resistance and specifying F-like pili (Meynell &
Datta, 1966).  It is the only fi+ R factor carrying tetracycline resistance and
specifying F-like pili (Meynell & Datta, 1966).  It is the only fi+ R factor so
far tested to determine restriction and modification of a number of DNA phages
including lambda.  The specificity of this restriction, which is unique, is
termed hsp I (Bannister & Glover, 1968).  Among the fi- R factors, all those
capable of restriction or modification fall into a single compatibility group N
(Hedges, 1972) and it seemed interesting to determine the compatibility
specificity of R124.  In this paper we show that this plasmid co-exists stably
with members of all the compatibility groups listed above and is thus the first
example of a new compatibility group, FIV.

<>

<1>Hedgpeth, J., Boyer, H.W.
<2>Hydrolysis of adenosine triphosphate accompanying interaction between Escherichia coli B restriction endonuclease and unmodified deoxyribonucleic acid.
<3>Biochim. Biophys. Acta
<4>331
<5>310-317
<6>1973
<7>The Escherichia coli B restriction endonuclease hydrolyzes ATP to ADP and Pi in
the presence of unmodified, double-strand DNA and Mg2+.  The requirements for
maximum ATP hydrolysis are essentially the same as those for phosphodiester
bond cleavage.  However, ATP is hydrolyzed under conditions where
phosphodiester bond cleavage is prevented, namely, in the absence of
S-adenosyl-L-methionine and in the presence of DNA which is resistant to the
endonuclease by virtue of mutations.  ATP is shown to be necessary for the
formation of the endonuclease-unmodified DNA complex.

<>

<1>Hedgpeth, J., Goodman, H.M., Boyer, H.W.
<2>DNA nucleotide sequence restricted by the RI Endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>3448-3452
<6>1972
<7>The sequence of DNA base pairs adjacent to the phosphodiester bonds cleaved by
the RI restriction endonuclease in unmodified DNA from coliphage lambda has
been determined.  The 5'-terminal nucleotide labeled with 32P and
oligonucleotides up to the heptamer were analyzed from a pancreatic DNase
digest.  The following sequence of nucleotides adjacent to the RI break made in
lambda DNA was deduced from these data and from the 3'-dinucleotide sequence
and nearest-neighbor analysis obtained from repair synthesis with the DNA
polymerase of Rous sarcoma virus 5'....A/TpG^pApApTpTpCpT/A....3'
3'....T/ApCpTpTpApAp^GpA/T....5' The RI endonuclease cleavage of the
phosphodiester bonds (indicated by arrows) generates 5'-phosphodiester bonds
(indicated by arrows) generates 5'-phosphoryls and short cohesive termini of
four nucleotides, pApApTpT.  The most striking feature of the sequence is its
symmetry.

<>

<1>Hegna, I.K., Bratland, H., Kolsto, A.
<2>BceS1, a new addition to the type III restriction and modification family.
<3>FEMS Microbiol. Lett.
<4>202
<5>189-193
<6>2001
<7>The nucleotide sequence of an 11-kb chromosomal BglII fragment from Bacillus cereus American
Type Culture Collection (ATCC) 10987 strain revealed two closely adjacent open reading frames
organized in an operon, of which the deduced amino acids showed identity to the type III
restriction and modification (R/M) subunits described in Gram-negative bacteria. An enhanced
transcription level was revealed when the culture was grown in the presence of foreign DNA. A
cell-free extract from this culture restricted pUC19, whereas from a plain medium the
restriction was very weak. The in vitro methylation protected pUC 19 from restriction. The R/M
system was designated BceS1 as this endonuclease required ATP and Mg(2+) as cofactors like
other type III endonucleases. BceS1 is the first chromosomal type III R/M system characterized
in a Gram-positive bacterium.

<>

<1>Hegna, I.K., Karlstrom, E.S., Lopez, R., Kristensen, T., Kolsto, A.-B.
<2>A type-II DNA restriction and modification system in Bacillus cereus?
<3>Gene
<4>114
<5>149-150
<6>1992
<7>The deduced amino acid (aa) sequence of an open reading frame, present on a fragment of the
Bacillus cereus ATCC10987 geneome, shows 29.3% identity within a 368-aa segment of the
type-III EcoPI modification and restriction operon.

<>

<1>Hehlmann, R., Hattman, S.
<2>Mutants of bacteriophage T2 gt with altered DNA methylase activity.
<3>J. Mol. Biol.
<4>67
<5>351-360
<6>1972
<7>Certain non-glucosylated (gt) T-even bacteriophage mutants are restricted by
prophage P1 (these are designated rP1).  Unrestricted mutants (uP1) have been
shown earlier to contain hypermethylated DNA compared to their rP1 parent.  The
DNA methylating enzyme induced by rP1 and uP1 phage has been partially purified
by affinity chromatography on DNA cellulose. Both rP1 and uP1 methylase
methylate adenine exclusively.  The uP1 methylase shows a greater thermal
lability than the rP1 enzyme; both enzymic forms are stabilized against heat
inactivation in the presence of substrate S-adenosylmethionine.  In terms of
their ability to methylate non-viral cytosine-containing DNA's, rP1 and uP1
methylase exhibit similar Km values and extents of methylation.  However, the
uP1 methylase shows a higher affinity (lowered Km) and greater site recognition
(higher extent of methylation) than the rP1 methylase with various T-even phage
(5-hydroxymethylcytosine-containing) DNA's.  These results are consistent with
the notion that mutation from rP1 to uP1 produces an alteration in the
viral-induced DNA methylase.

<>

<1>Heidelberg, J.F. et al.
<2>DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae.
<3>Nature
<4>406
<5>477-483
<6>2000
<7>Here we determine the complete genomic sequence of the gram negative, gamma-Proteobacterium
Vibrio cholerae El Tor N16961 to be 4,033,460 base pairs (bp). The genome consists of two
circular chromosomes of 2,961,146 bp and 1,072,314 bp that together encode 3,885 open reading
frames. The vast majority of recognizable genes for essential cell functions (such as DNA
replication, transcription, translation and cell-wall biosynthesis) and pathogenicity (for
example, toxins, surface antigens and adhesins) are located on the large chromosome. In
contrast, the small chromosome contains a larger fraction (59%) of hypothetical genes compared
with the large chromosome (42%), and also contains many more genes that appear to have origins
other than the gamma-Proteobacteria. The small chromosome also carries a gene capture system
(the integron island) and host 'addiction' genes that are typically found on plasmids; thus,
the small chromosome may have originally been a megaplasmid that was captured by an ancestral
Vibrio species. The V. cholerae genomic sequence provides a starting point for understanding
how a free-living, environmental organism emerged to become a significant human bacterial
pathogen.

<>

<1>Heidelberg, J.F. et al.
<2>Genome sequence of the dissimilatory metal ion-reducing bacterium Shewanella oneidensis.
<3>Nat. Biotechnol.
<4>20
<5>1118-1123
<6>2002
<7>Shewanella oneidensis is an important model organism for bioremediation studies because of its
diverse respiratory capabilities, conferred in part by multicomponent, branched electron
transport systems. Here we report the sequencing of the S. oneidensis genome, which consists
of a 4,969,803-base pair circular chromosome with 4,758 predicted protein-encoding open
reading frames (CDS) and a 161,613-base pair plasmid with 173 CDSs. We identified the first
Shewanella lambda-like phage, providing a potential tool for further genome engineering.
Genome analysis revealed 39 c-type cytochromes, including 32 previously unidentified in S.
oneidensis, and a novel periplasmic [Fe] hydrogenase, which are integral members of the
electron transport system. This genome sequence represents a critical step in the elucidation
of the pathways for reduction (and bioremediation) of pollutants such as uranium (U) and
chromium (Cr), and offers a starting point for defining this organism's complex electron
transport systems and metal ion-reducing capabilities.

<>

<1>Heidmann, S., Seifert, W., Kessler, C., Domdey, H.
<2>Cloning, characterization and heterologous expression of the SmaI restriction-modification system.
<3>Nucleic Acids Res.
<4>17
<5>9783-9796
<6>1989
<7>The genes coding for the class-II Serratia marcescens restriction-modification
system have been cloned and expressed in E. coli.  Recombinant clones
restricted incoming phage only poorly; the recombinant plasmids, however,
became fully modified in vivo, i.e. completely resistant against digestion with
R.SmaI.  The determined nucleotide sequence of the cloned system revealed three
open reading frames with lengths of 252 bp, 741 bp, and 876 bp.  Through
various deletion experiments and an insertion-mutation experiment the 876 bp
open reading frame could be assigned to the SmaI DNA modification enzyme and
the 741 bp open reading frame to the SmaI restriction endonuclease.  Mapping of
the transcription start sites of the genes revealed that the SmaI endonuclease
is transcribed as a polycistronic mRNA together with a 252 bp long preceding
open reading frame of unknown function.  No homology was found when comparing
the amino acid sequence of M.SmaI with the published sequences of m5C-specific
DNA modification methyltransferases.  On the other hand, a stretch of 14 amino
acids in the C-proximal region of M.SmaI shows a signficant homology to the
C-proximal amino acid sequences of the N6A-methyltransferases M.HinfI and
M.DpnIIA and the N4C-methyltransferase M.PvuII.

<>

<1>Heilig, R., Flament, D., Guegen, Y., Raffin, J., Henneke, G., Rolland, J., Weissenbach, J., Saurin, W., Querellou, J., Lecompte, O., Dietrich, J., Prieur, D., Thierry, J., Forterre, P.
<2>GENOME SEQUENCE AND POLYPEPTIDES OF PYROCOCCUS ABYSSI, FRAGMENT AND USES THEREOF.
<3>Japanese Patent Office
<4>JP 2004500802 A
<5>
<6>2004
<7>
<>

<1>Heininger, K., Horz, W., Zachau, H.G.
<2>Specificity of cleavage by a restriction nuclease from Bacillus subtilis.
<3>Gene
<4>1
<5>291-303
<6>1977
<7>The restriction nuclease from B. subtilis (Bsu) which cleaves in the middle of
the tetra-nucleotide sequence 5'-GGCC-3'	 3'-CCGG-5' has been found to decrease
its substrate specificity at high nuclease concentrations.  There are special
conditions, high pH, low ionic strength, and high glycerol contents, which
strongly enhance splitting with decreased specificity and also lead to
splitting of single-stranded DNA.  By sequence analyses it is shown that the
reduction in specificity of Bsu corresponds to cleavage predominantly at
5'-GC-3' 3'-CG-5' sequences.  No comparable change in specificity has been
observed in a restriction nuclease from Haemophilus aegyptius (HaeIII), an
isoschizomer of Bsu.

<>

<1>Heinl, S., Wibberg, D., Eikmeyer, F., Szczepanowski, R., Blom, J., Linke, B., Goesmann, A., Grabherr, R., Schwab, H., Puhler, A., Schluter, A.
<2>Insights into the completely annotated genome of Lactobacillus buchneri CD034, a strain isolated from stable grass silage.
<3>J. Biotechnol.
<4>161
<5>153-166
<6>2012
<7>Lactobacillus buchneri belongs to the group of heterofermentative lactic acid
bacteria and is a common member of the silage microbiome. Here we report the
completely annotated genomic sequence of L. buchneri CD034, a strain isolated
from stable grass silage. The whole genome of L. buchneri CD034 was sequenced on
the Roche Genome Sequencer FLX platform. It was found to consist of four
replicons, a circular chromosome, and three plasmids. The circular chromosome was
predicted to encode 2319 proteins and contains a genomic island and two prophages
which significantly differ in G+C-content from the remaining chromosome. It
possesses all genes for enzymes of a complete phosphoketolase pathway, whereas
two enzymes necessary for glycolysis are lacking. This confirms the
classification of L. buchneri CD034 as an obligate heterofermentative lactic acid
bacterium. A set of genes considered to be involved in the lactate degradation
pathway and genes putatively involved in the breakdown of plant cell wall
polymers were identified. Moreover, several genes encoding putative S-layer
proteins and two CRISPR systems, belonging to the subclasses I-E and II-A, are
located on the chromosome. The largest plasmid pCD034-3 was predicted to encode
57 genes, including a putative polysaccharide synthesis gene cluster, whereas the
functions of the two smaller plasmids, pCD034-1 and pCD034-2, remain cryptic.
Phylogenetic analysis based on sequence comparison of the conserved marker gene
rpoA reveals that L. buchneri CD034 is more closely related to Lactobacillus
hilgardii strains than to Lactobacillus brevis and Lactobacillus plantarum
strains. Comparison of the L. buchneri CD034 core genome to other fully sequenced
and closely related members of the genus Lactobacillus disclosed a high degree of
conservation between L. buchneri CD034 and the recently sequenced L. buchneri
strain NRRL B-30929 and a more distant relationship to L. buchneri ATCC 11577 and
L. brevis ssp. gravesensis ATCC 27305, which cluster together with L. hilgardii
type strain ATCC 8290. L. buchneri CD034 genome information will certainly
provide the basis for further postgenome studies with the objective to optimize
application of the strain in silage production.

<>

<1>Heintschel von Heinegg, E., Nalik, H.P., Schmid, E.N.
<2>Characterisation of a Helicobacter pylori phage (HP1).
<3>J. Med. Microbiol.
<4>38
<5>245-249
<6>1993
<7>The infection of two Helicobacter pylori strains with a
phage-containing supernate of the lysogenic H. pylori strain IMMi 290/89
resulted in a lytic cycle and propagation of phage HP1. In
negatively-stained preparations, the empty phage heads measured 55-60 nm
in diameter and mature heads measured 50 nm. The flexible, striated
phage tail was c. 170 nm in length and 9.5 nm in diameter. The phage
showed a mean density of 1.40 g/cm3 in sucrose-density gradients and
contained double-stranded DNA c. 22,000 bp in length.

<>

<1>Heinze, R.J., Giron-Monzon, L., Solovyova, A., Elliot, S.L., Geisler, S., Cupples, C.G., Connolly, B.A., Friedhoff, P.
<2>Physical and functional interactions between Escherichia coli MutL and the Vsr repair endonuclease.
<3>Nucleic Acids Res.
<4>37
<5>4453-4463
<6>2009
<7>DNA mismatch repair (MMR) and very-short patch (VSP) repair are two pathways involved in the
repair of T:G mismatches. To learn about
competition and cooperation between these two repair pathways, we analyzed
the physical and functional interaction between MutL and Vsr using
biophysical and biochemical methods. Analytical ultracentrifugation
reveals a nucleotide-dependent interaction between Vsr and the N-terminal
domain of MutL. Using chemical crosslinking, we mapped the interaction
site of MutL for Vsr to a region between the N-terminal domains similar to
that described before for the interaction between MutL and the strand
discrimination endonuclease MutH of the MMR system. Competition between
MutH and Vsr for binding to MutL resulted in inhibition of the
mismatch-provoked MutS- and MutL-dependent activation of MutH, which
explains the mutagenic effect of Vsr overexpression. Cooperation between
MMR and VSP repair was demonstrated by the stimulation of the Vsr
endonuclease in a MutS-, MutL- and ATP-hydrolysis-dependent manner, in
agreement with the enhancement of VSP repair by MutS and MutL in vivo.
These data suggest a mobile MutS-MutL complex in MMR signalling, that
leaves the DNA mismatch prior to, or at the time of, activation of
downstream effector molecules such as Vsr or MutH.

<>

<1>Heip, J., Rolfe, B., Schell, J.
<2>Abolition of host cell restriction by high multiplicity of phage infection.
<3>Virology
<4>59
<5>356-370
<6>1974
<7>When restricting host cells (Escherichia coli K12 or E. coli B) are infected
first with a nonmodified lambda phage (called the helper phage) at high
multiplicity and subsequently superinfected with a second nonmodified lambda
vir phage (called the test phage), it can be shown that an increased number of
infected cells produce a successful infection of phage which carry the virulent
marker of the test phage.  This multiplicity effect has been studied both in
normal and in recombination defective strains, in the presence or the absence
of chloramphenicol and mitomycin C and in circumstances where the gene
expression of the nonmodified helper phage was prevented by immunity.  The
results indicate that the nonmodified helper-phage causes a temporary
inactivation of the restricting system and that the multiplicity effect is not
due to gene complementation or cooperation or to recombination.  Furthermore,
this multiplicity effect can be observed in different hosts and with various
nonmodified phages.  When similar experiments were performed with modified
helper phage no multiplicity effect was observed.

<>

<1>Heiter, D., Lunnen, K., Wilson, G.G.
<2>Engineering of strand-specific, sequence-specific, DNA-nicking enzymes from restriction endonucleases.
<3>US Patent Office
<4>US 20030100094 A
<5>62
<6>2003
<7>Methods are provided for converting into a sequence specific strand specific and location
specific DNA nicking endonuclease, a restriction endonuclease that recognizes an asymmetric
DNA sequence, the endonuclease having two catalytic sites and one or more single sequence
specific DNA-binding domains.  In one embodiment the method requires inactivating one of the
catalytic sites of the restriction endonuclease.  In another embodiment, the restriction
endonuclease is a dimer having a first and second subunit each comprising a sequence specific
DNA binding domain, a catalytic site and a dimerization domain.  The nicking endonuclease is
formed from combining one subunit having an inactivated catalytic site and a second subunit
having an inactivated DNA binding domain.  The nicking endonuclease may be converted into a
restriction endonuclease by the addition of manganese cations in the digestion buffer.

<>

<1>Heiter, D., Lunnen, K., Wilson, G.G.
<2>Method for engineering strand-specific, sequence-specific, DNA-nicking enzymes.
<3>US Patent Office
<4>US 7081358 A
<5>
<6>2006
<7>Methods are provided for converting into a sequence specific strand specific and location
specific DNA nicking endonuclease, a restriction endonuclease that recognizes an asymmetric
DNA sequence, the endonuclease having two catalytic sites and one or more single sequence
specific DNA-binding domains.  In one embodiment the method requires inactivating one of the
catalytic sites of the restriction endonuclease.  In another embodiment, the restriction
endonuclease is a dimer having a first and second subunit each comprising a sequence specific
DNA binding domain, a catalytic site and a dimerization domain.  The nicking endonuclease is
formed from combining one subunit having an inactivated catalytic site and a second subunit
having an inactivated DNA binding domain.  The nicking endonuclease may be converted into a
retriction endonuclease by the addition of manganese cations in the digestion buffer.

<>

<1>Heiter, D., Lunnen, K., Wilson, G.G.
<2>A method for engineering strand-specific, sequence-specific, DNA-nicking enzymes.
<3>International Patent Office
<4>WO 03055975 A
<5>
<6>2003
<7>Methods are provided for converting into a sequence specific strand specific and location
specific DNA nicking endonuclease, a restriction endonuclease that recognizes an asymmetric
DNA sequence, the endonuclease having two catalytic sites and one or more single sequence
specific DNA-binding domains.  In one embodiment the method requires inactivating one of the
catalytic sites of the restriction endonuclease.  In another embodiment, the restriction
endonuclease is a dimmer having a first and second subunit each comprising a sequence specific
DNA binding domain, a catalytic site and a dimerization domain.  The nicking endonuclease is
formed from combining one subunit having an inactivated catalytic site and a second subunit
having an inactivated DNA binding domain.  The nicking endonuclease may be converted into a
restriction endonuclease by the addition of manganese cations in the digestion buffer.

<>

<1>Heiter, D.F., Lunnen, K.D., Wilson, G.G.
<2>Site-specific DNA-nicking mutants of the heterodimeric restriction endonuclease R.BbvCI.
<3>J. Mol. Biol.
<4>348
<5>631-640
<6>2005
<7>The restriction enzyme R.BbvCI cleaves duplex DNA within a seven base-pair asymmetric
recognition sequence, thus: CCTCAGC/GCGAGG -> CC^TCAGC/GC^TGAGG. We show that R.BbvCI
comprises
two different subunits, R-1 and R-2; that each subunit contains a
catalytic site for DNA strand hydrolysis; and that these sites act
independently and strand-specifically. In turn, each catalytic site was
inactivated by mutagenesis to form dimeric enzymes in which only one
bite remained functional. The altered enzymes hydrolyzed just one
strand of the recognition sequence, nicking the DNA rather than
cleaving it. Enzymes in which the catalytic site in the R-1 subunit
remained functional nicked the bottom strand of the sequence, producing
CCTCAGC/GC^TGAGG, while those in which the catalytic site
in the R-2 subunit remained functional nicked the top strand, producing
CC^TCAGC/GCTGAGG. These DNA-nicking enzymes could prove
useful for investigation of DNA repair, recombination, and replication,
and for laboratory procedures that initiate from nicks, such as DNA
degradation, synthesis, and amplification.

<>

<1>Heithoff, D.M., Enioutina, E.Y., Daynes, R.A., Sinsheimer, R.L., Low, D.A., Mahan, M.J.
<2>Salmonella DNA adenine methylase mutants confer cross-protective immunity.
<3>Infect. Immun.
<4>69
<5>6725-6730
<6>2001
<7>Salmonella isolates that lack or overproduce DNA adenine methylase elicited a cross-protective
immune response to different Salmonella serovars.  The protection afforded by the Salmonella
enterica serovar Typhimurium Dam vaccine was greater than that elicited in mice that survived
a virulent infection.  S. enterica serovar Typhimurium Dam mutant strains exhibited enhanced
sensitivity to mediators of innate immunity such as antimicrobial peptides, bile, salts, and
hydrogen peroxide.  Also, S. enterica serovar Typhimurium Dam- vaccines were not
immunosuppressive; unlike wild-type vaccines, they failed to induce increased nitric oxide
levels and permitted a subsequent robust humoral response to diptheria toxoid antigen in
infected mice.  Dam mutant strains exhibited a low-grade persistence which, coupled with the
nonimmunosuppression and the ectopic protein expression caused by altered levels of Dam, may
provide an expanded source of potential antigens in vaccinated hosts.

<>

<1>Heithoff, D.M., Julio, S.M., Mahan, M.J.
<2>The role of DNA adenine methylation in controlling bacterial virulence.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>59
<6>2001
<7>Salmonella DNA adenine methylase (Dam) mutants are avirulent and are effective as live
vaccines against murine typhoid fever. The role of
Dam in virulence and in the elicitation of protective immune responses
may rely on its capacity as a global regulator of gene expression. Dam
was shown to regulate the expression of many genes that are induced
during infection, and Dam- and Dam over-producer mutants expressed
several proteins distinct from each other and from wild-type Salmonella
grown in vitro. To explore whether Dam mutants were attenuated for
virulence in pathogens other than Salmonella, we attempted to construct
Dam-mutations in V. cholerae and Y. pseudotuberculosis. Dam is
essential for viability in V. cholerae and in Yersinia. However, Dam
overproduction in both organisms was not lethal and lead to reduced
virulence in both organisms. Overproduction of Dam in Yersinia resulted
in the ectopic secretion of Yersinia outer proteins (Yops) and a fully
protective immune response in vaccinated mice. Since DNA adenine
methylases are highly conserved in a wide variety of virulent bacteria,
dysregulation of Dam activity is potentially a general strategy for the
development of vaccines against varied bacterial pathogens.

<>

<1>Heithoff, D.M., Julio, S.M., Mahan, M.J.
<2>Regulation of bacterial virulence by DNA methylation.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>102
<5>73
<6>2002
<7>Salmonella that lack or overproduce DNA adenine methylase (Dam) elicited a cross-protective
immune response to different Salmonella
serovars. Mice immunized with Dam mutant strains conferred
significantly more protection than that conferred by mice exposed to a
sub-lethal challenge with the virulent strain. In contrast to
Salmonella and E. coli, Dam is essential for viability in Vibrio
cholerae and Yersinia pseudotuberculosis; however, Dam overproduction
is not lethal and significantly attenuated the virulence of both
pathogens. Y. pseudotuberculosis mutants that overproduce Dam conferred
fully protective immune responses and secreted several Yersinia outer
proteins (Yops) under conditions that are nonpermissive for secretion
in wild-type strains. Dam overproduction disrupted both the thermal and
calcium regulation of the synthesis of the YopE cytotoxin, and relaxed
the thermal but not the calcium dependence of YopE secretion. Because
alterations of Dam activity attenuate such a diverse set of pathogenic
organisms, the role of Dam in virulence may emerge as a general theme
in bacterial pathogenesis.

<>

<1>Heithoff, D.M., Sinsheimer, R.L., Low, D.A., Mahan, M.J.
<2>An essential role for DNA adenine methylation in bacterial virulence.
<3>Science
<4>284
<5>967-970
<6>1999
<7>Salmonella typhimurium lacking DNA adenine methylase were fully proficient in colonization of
mucosal sites but showed severe defects in colonization of deeper tissue sites.  These Dam-
mutants were totally avirulent and were effective as live vaccines against murine typhoid
fever.  Dam regulated the expression of at least 20 genes known to be induced during
infection; a subset of these genes are among those activated by the PhoP global virulence
regulator.  PhoP, in turn, affected Dam methylation at specific genomic sites, as evidenced by
alterations in DNA methylation patterns.  Dam inhibitors are likely to have broad
antimicrobial action, and Dam- derivatives of these pathogens may serve as live attenuated
vaccines.

<>

<1>Heithoff, D.M., Sinsheimer, R.L., Low, D.A., Mahan, M.J.
<2>In vivo gene expression and the adaptive response: From pathogenesis to vaccines and antimicrobials.
<3>Philos. Trans. R. Soc. Lond. B. Biol. Sci.
<4>355
<5>633-642
<6>2000
<7>Microbial pathogens possess a repertoire of virulence determinants that each make unique
contributions to fitness during infection. Analysis of
these in vivo-expressed functions reveals the biology of the infection
process, encompassing the bacterial infection strategies and the host
ecological and environmental retaliatory strategies designed to combat
them (e.g. thermal, osmotic, oxygen, nutrient and acid stress). Many of
the bacterial virulence functions that contribute to a successful
infection are normally only expressed during infection. A genetic
approach was used to isolate mutants that ectopically expressed many of
these functions in a laboratory setting. Lack of DNA adenine methylase
(Dam) in Salmonella typhimurium abolishes the preferential expression
of many bacterial virulence genes in host tissues. Dam- Salmonella were
proficient in colonization of mucosal sites but were defective in
colonization of deeper tissue sites. Additionally, Dam- mutants were
totally avirulent and effective as live vaccines against murine typhoid
fever. Since dam is highly conserved in many pathogenic bacteria that
cause significant morbidity and mortality worldwide, Dams are
potentially excellent targets for both vaccines and antimicrobials.

<>

<1>Heitman, J.
<2>On the repair of DNA breaks and the specificity of the EcoRI restriction enzyme.
<3>Ph.D. Thesis, Rockefeller University, NY, USA
<4>
<5>1-209
<6>1989
<7>We would like to understand how proteins and enzymes interact with DNA.  We
describe here our studies of the Mrr methylation dependent restriction enzyme,
DNA single- and double-strand break repair in E. coli, and substrate
recognition by the EcoRI endonuclease.  Many species of bacteria make
restriction-modification systems to destroy foreign DNA that enters the cell.
These systems usually consist of an endonuclease that cleaves a specific DNA
sequence and a methylase which modifies the DNA to protect the host chromosome.
We observed that when foreign site-specific methylases are expressed in E.
coli, the SOS DNA repair response is induced.  This DNA damage is inflicted by
E. coli restriction enzymes that cleave adenine (Mrr) or cytosine (McrB)
methylated DNA.  The genes encoding four of the five known E. coli restriction
systems lie clustered together, perhaps to coordinate or sequester the cellular
defense system.  Several of these restriction systems differ between E. coli
species, suggesting that their action may establish species boundaries.  The
EcoRI endonuclease cleaves DNA molecules at the sequence GAATTC.  This enzyme
is well characterized biochemically, the sequence of its gene is known, and the
X-ray crystal structure of an EcoRI-DNA complex has been solved at 3 A
resolution.  (McClarin, et al., 1986).  EcoRI serves as a paradigm for other
restriction enzymes and as a model of DNA-protein interactions.  We took a
genetic approach to study the EcoRI endonuclease.  We first asked if EcoRI DNA
double-strand breaks are repaired in E. coli.  To this end, a series of
temperature-sensitive EcoRI endouclease alleles were isolated.  Temperature
shifts with these alleles revealed that in vivo DNA scission induces the E.
coli SOS DNA  repair response.  However, neither SOS induction nor
recombination are required to repair these lesions.  DNA ligase is required and
may suffice to repair EcoRI breaks in the E. coli chromosome.  An in vivo DNA
scission assay was devised based on the finding that DNA breaks induce the SOS
response.  SOS induction was monitored with strains carrying the lactose operon
fused to an SOS inducible promoter.  After DNA scission, these strains produce
Beta-galactosidase and form blue colonies on X-Gal medium.  With this blue
colony phenotype as a screen, two approaches were taken to isolate EcoRI
mutants altered or disrupted in substrate specificity.  First, amino acids
(E144, R200) implicated in substrate binding by the crystal structure were
subjected to site-directed mutagenesis.  Of 50 of the 60 possible
substitutions, several alleles retain weak endonuclease activity which, in vivo
and in vitro, is of wild-type specificity.  Therefore the simple hydrogen bond
model proposed from the crystal structure is insufficient to explain substrate
recognition and additional interactions must participate in the
substrate-enzyme complex.  In the second approach, mutants of an EcoRI ts
allele were isolated which conditionally induce the SOS response and impair
cell growth in spite of the normally protective methylase.  In vitro, these
mutant proteins exhibit enhanced cleavage activity at EcoRI* sites, sequences
which differ by one nucleotide from the normal recognition site and are also
cleaved by the wild-type enzyme under altered buffer conditions.  Four of the
five mutations of this type lie at the DNA-protein interface and may directly
alter or disrupt substrate recognition.  One other (H114Y) lies far from the
binding and cleavage sites.  This mutation falls three amino acids away from a
previously described mutation, E111G (King et al., 1986, 1988), which severly
impairs cleavage activity without altering DNA binding.  These two mutations
support a model whereby DNA scission by the EcoRI endonuclease is
allosterically activated upon substrate binding:  we suggest that the E111G
mutation inhibits this conformational change while the H114Y mutation renders
it more facile such that additional DNA sequences act as allosteric effectors
and trigger cleavage.

<>

<1>Heitman, J.
<2>How the EcoRI endonuclese recognizes and cleaves DNA.
<3>Bioessays
<4>14
<5>445-454
<6>1992
<7>One popular recombinant DNA tool is the EcoRI endonuclease, which cleaves DNA at GAATTC sites
and serves as a paradigm for sequence specific DNA-enzyme interactions. The recently revised
X-ray crystal structure of an EcoRI-DNA complex reveals EcoRI employs novel DNA recognition
motifs, a four a-helix bundle and two extended chains, which project into the major groove to
contact substrate purines and pyrimidines. Interestingly, pyrimidine contacts had been
predicted based on genetic and biochemical studies. Current work focuses on the EcoRI active
site structure, enzyme and substrate conformational changes during catalysis, and
host-restriction system interactions.

<>

<1>Heitman, J.
<2>On the origins, structures and functions of restriction-modification enzymes.
<3>Genet. Eng. (N Y), Plenum Press, Setlow, J.K., New York
<4>15
<5>57-108
<6>1993
<7>Typically, restriction-modification (RM) systems consist of two enzymes: an endonuclease that
recognizes and cleaves a specific DNA sequence, and a methyltransferase that modifies the same
sequence to protect the host chromosome from cleavage. These enzymes also play an important
role in genetic engineering and provide insight into the basis of sequence-specific
DNA-protein interactions. A growing number of type II restriction endonucleases and
methyltransferases are being subjected to biochemical and genetic studies which, when combined
with ongoing X-ray crystallographic analyses, promise to provide detailed models for
mechanisms of DNA recognition and catalysis. Studies on anti-restriction systems, the repair
of DNA-single- and double-strand breaks, and the roles of DNA lesions in recombination
initiation, suggest RM systems may provoke genome rearrangements and assimilate foreign DNA
into the host genome and point towards possible evolutionary sources of this interesting group
of DNA metabolizing enzymes.

<>

<1>Heitman, J., Fulford, W., Model, P.
<2>Phage Trojan horses: a conditional expression system for lethal genes.
<3>Gene
<4>85
<5>193-197
<6>1989
<7>The EcoRI restriction enzyme (ENase) cleaves DNA molecules within the sequence
GAATTC.  Cells expressing this lethal activity normally make a second enzyme,
the M.EcoRI methyltransferase (MTase), which protects their chromosomal DNA by
modifying the EcoRI recognition sites.  To isolate mutants of the EcoRI ENase,
its gene was cloned into a filamentous phage vector (M13mp18) under control of
the lac promoter.  Normally, filamentous phages (M13, f1 and their derivatives)
form turbid plaques by impairing the growth of their host cell without killing
it.  In contrast, phages expressing the EcoRI ENase kill the host cell, but
survive long enough to produce plaques which are very clear.  Expression of the
M.EcoRI MTase rescues the host and restores turbid plaque formation.  EcoRI
ENase mutants were isolated by screening for mutants that make turbid, instead
of clear, plaques on an M- host.  This conditional expression system may be
useful for cloning and mutating genes for other toxic proteins.

<>

<1>Heitman, J., Ivanenko, T., Kiss, A.
<2>DNA nicks inflicted by restriction endonucleases are repaired by a RecA- and RecB-dependent pathway in Escherichia coli.
<3>Mol. Microbiol.
<4>33
<5>1141-1151
<6>1999
<7>Two mutants of the EcoRI endonuclease (R200K and E144C) predominantly nick only one strand of
the DNA substrate. Temperature sensitivity of the mutant enzymes allowed us to study the
consequences of inflicting DNA nicks at EcoRI sites in vivo. Expression of the EcoRI
endonuclease mutants in the absence of the EcoRI methyltransferase induces the SOS DNA repair
response and greatly reduces viability of recA56, recB21 and lexA3 mutant strains of
Escherichia coli. In parallel studies, overexpression of the EcoRV endonuclease in cells also
expressing the EcoRV methyltransferase was used to introduce nicks at non-cognate EcoRV sites
in the bacterial genome. EcoRV overproduction was lethal in recA56 and recB21 mutant strains
and moderately toxic in a lexA3 mutant strain. The toxic effect of EcoRV overproduction could
be partially alleviated by introduction into the cells of multiple copies of the E. coli DNA
ligase gene. These observations suggest that an increased number of DNA nicks can overwhelm
the repair capacity of DNA ligase, resulting in the conversion of a proportion of DNA nicks
into DNA lesions that require recombination for repair.

<>

<1>Heitman, J., Model, P.
<2>Site-specific methylases induce the SOS DNA repair response in Escherichia coli.
<3>J. Bacteriol.
<4>169
<5>3243-3250
<6>1987
<7>Expression of the site-specific adenine methylase HhaII (GmeANTC, where me
is methyl) or PstI (CTGCmeAG) induced the SOS DNA repair response in Escherichia
coli. In contrast, expression of methylases indigenous to E. coli either did not induce SOS
(EcoRI-GAmeATTC) or induced SOS to a lesser extent (dam-GmeATC). Recognition of
adenine-methylated DNA required the product of a previously undescribed gene, which we
named mrr (methylated adenine recognition and restriction). We suggest that mrr encodes
an endonuclease that cleaves DNA containing N6-methyladenine and that DNA double-
strand breaks induce the SOS response. Cytosine methylases foreign to E. coli (MspI
[meCCGG], HaeIII [GGmeCC], BamHI [GGATmeCC],HhaI [GmeCGC],
BsuRI[GGmeCC], and M.SPR) also induced SOS, whereas one indigenous to E. coli
(EcoRII [CmeCA/TGG]) did not. SOS induction by cytosine methylation required the rglB
locus, which encodes an endonuclease that cleaves DNA containing 5-hydroxymethyl- or
5-methylcytosine (E. A. Raleigh and G. Wilson, Proc. Natl. Acad. Sci. USA 83:9070-
9074, 1986).

<>

<1>Heitman, J., Model, P.
<2>Substrate recognition by the EcoRI endonuclease.
<3>Proteins
<4>7
<5>185-197
<6>1990
<7>The EcoRI restriction endonuclease is one of the most widely used tools for
recombinant DNA maniulations.  Because the EcoRI enzyme has been extremely well
characterized biochemically and its structure is known at 3 angstrom resolution
as an enzyme-DNA complex, EcoRI also serves as a paradigm for other restriction
enzymes and as an important model of DNA-protein interactions.  To facilitate a
genetic analysis of the EcoRI enzyme, we devised an in vivo DNA scission assay
based on our finding that DNA double-strand breaks induce the Escherichia coli
SOS response and thereby increase beta-galactosidase expression from SOS::lacZ
gene fusions.  By site-directed mutagenesis, 50 of 60 possible point mutations
were generated at three amino acids (E144, R145, and R200) implicated in
substrate recognition by the crystal structure.  Although several of these
mutant enzymes retain partial endonuclease activity, none are altered in
substrate specificity in vivo or in vitro.  These findings argue that, in
addition to the hydrogen bond interactions revealed by the crystal structure,
the EcoRI enzyme must make additional contacts to recognize its substrate.

<>

<1>Heitman, J., Model, P.
<2>Mutants of the EcoRI endonuclease with promiscuous substrate specificity implicate residues involved in substrate recognition.
<3>EMBO J.
<4>9
<5>3369-3378
<6>1990
<7>The EcoRI restriction endonuclease cleaves DNA molecules at the sequence GAATTC. We devised a
genetic screen to isolate EcoRI mutants with altered or broadened substrate specificity. In
vitro, the purified mutant enzymes cleave both the wild-type substrate and sites which differ
from this by one nucleotide (EcoRI star sites). These mutations identify four residues
involved in substrate recognition and catalysis that are different from the amino acids
proposed to recognize the substrate based on the EcoRI-DNA co-crystal structure. In fact,
these mutations suppress EcoRI mutants altered at some of the proposed substrate binding
residues (R145, R200). We argue that these mutations permit cleavage of additional DNA
sequences either by perturbing or removing direct DNA-protein interactions or by facilitating
conformational changes that allosterically couple substrate binding to DNA scission.

<>

<1>Heitman, J., Model, P.
<2>SOS induction as an in vivo assay of enzyme-DNA interactions.
<3>Gene
<4>103
<5>1-9
<6>1991
<7>We have constructed strains which are convenient and sensitive indicators of
DNA damage and describe their use.  These strains utilize an SOS::lacZ fusion
constructed by Kenyon and Walker [Proc. Natl. Acad. Sci. USA 77 (1980)
2819-2823] and respond to DNA damage by producing Beta-galactosidase.  They can
be used to characterize restriction systems and screen for restriction
endonuclease mutants.  Applications include the study of other enzymes involved
in DNA metabolism, such as DNA methyltransferases, topoisomerases,
recombinases, and DNA replication and repair enzymes.

<>

<1>Heitman, J., Zinder, N.D., Model, P.
<2>Repair of the Escherichia coli chromosome after in vivo scission by the EcoRI endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>86
<5>2281-2285
<6>1989
<7>We prepared a set of temperature-sensitive mutants of the EcoRI endonuclease.
Under semipermissive conditions, Escherichia coli strains bearing these alleles
form poorly growing colonies in which intracellular substrates are cleaved at
EcoRI sites and the SOS DNA repair response is induced.  Strains defective in
SOS induction (lexA3 mutant) or SOS induction and recombination (recA56 and
recB21 mutants) are not more sensitive to this in vivo DNA scission, whereas
strains deficient in DNA ligase (lig4 and lig ts7 mutants) are extremely
sensitive.  We conclude that although DNA scission induces the SOS response,
neither this induction nor recombination are required for repair.  DNA ligase
is necessary and may be sufficient to repair EcoRI-mediated DNA breaks in the
E. coli chromosome.

<>

<1>Heitman, J.B.
<2>On the repair of DNA breaks and the specificity of the EcoRI restriction enzyme.
<3>Diss. Abstr.
<4>51
<5>562B
<6>1990
<7>We took a genetic approach to study the well characterized EcoRI restriction
enzyme, an endonuclease that cleaves DNA molecules at the sequence GAATTC.
First, a series of temperature-sensitive EcoRI endonuclease alleles were
isolated.  Temperature shifts with these alleles revealed that in vivo DNA
scission induces the E. coli SOS DNA repair response.  However, neither SOS
induction nor recombination are required to repair these lesions.  DNA ligase
is required and may suffice to repair EcoRI breaks in the E. coli chromosome.
To monitor DNA scission in vivo we employed strains carrying the lactose operon
fused to an SOS inducible promoter.  After DNA scission, these strains produce
beta-galactosidase and form blue colonies on X-Gal medium.  Using this assay,
two approaches were taken to isolate EcoRI mutants altered or disrupted in
substrate specificity.  First, amino acids (E144, R145, R200) implicated in
substrate binding by the crystal structure were subjected to site-directed
mutagenesis.  Of 50 of the 60 possible substitutions, several alleles retain
weak endonuclease activity which is of wild-type specificity.  Therefore the
simple hydrogen bond model proposed from the crystal structure is insufficient
to explain substrate recognition and additional interactions must participate
in the substrate-enzyme complex.  In the second approach, mutants of an EcoRI
ts allele were isolated which conditionally induce the SOS response in spite of
the protective methylase.  These mutant proteins exhibit enhanced cleavage
activity at EcoRI sites.  Four of five isolated mutations lie at the
DNA-protein interface and may directly alter or disrupt substrate recognition.
One other (H114Y) lies far from the binding or cleavage sites and falls three
amino acids away from a previously described mutation, E111G (King et al, 1986,
1988), which severely impairs DNA cleavage without altering DNA binding.  These
mutations support a model whereby DNA scission by the EcoRI endonuclease is
allosterically activated upon substrate binding: the E111G mutation may inhibit
this conformational change while the H114Y mutation may render it more facile
such that additional DNA sequences act as allosteric effectors and trigger
cleavage.

<>

<1>Hejdankova, J., Rypackova, B.
<2>Method for improving the recovery of restriction endonuclease PstI from lysates of Providencia stuartii.
<3>Czech Patent Office
<4>CS 258940 B
<5>
<6>1989
<7>
<>

<1>Hejnova, J., Dobrindt, U., Nemcova, R., Rusniok, C., Bomba, A., Frangeul, L., Hacker, J., Glaser, P., Sebo, P., Buchrieser, C.
<2>Characterization of the flexible genome complement of the commensal Escherichia coli strain A0 34/86 (O83 : K24 : H31).
<3>Microbiology
<4>151
<5>385-398
<6>2005
<7>Colonization by the commensal Escherichia coli strain A0 34/86 (O83 : K24 : H31) has proved to
be safe and efficient in the prophylaxis and
treatment of nosocomial infections and diarrhoea of preterm and newborn
infants in Czech paediatric clinics over the past three decades. In
searching for traits contributing to this beneficial effect related to the
gut colonization capacity of the strain, the authors have analysed its
genome by DNA-DNA hybridization to E. coli K-12 (MG1655) genomic DNA
arrays and to 'Pathoarrays', as well as by multiplex PCR, bacterial
artificial chromosome (BAC) library cloning and shotgun sequencing. Four
hundred and ten E. coli K-12 ORFs were absent from A0 34/86, while 72 out
of 456 genes associated with pathogenicity islands of E. coli and Shigella
were also detected in E. coli A0 34/86. Furthermore, extraintestinal
pathogenic E. coli-related genes involved in iron uptake and adhesion were
detected by multiplex PCR, and genes encoding the HlyA and cytotoxic
necrotizing factor toxins, together with 21 genes of the uropathogenic E.
coli 536 pathogenicity island II, were identified by analysis of 2304
shotgun and 1344 BAC clone sequences of A0 34/86 DNA. Multiple sequence
comparisons identified 31 kb of DNA specific for E. coli A0 34/86; some of
the genes carried by this DNA may prove to be implicated in the
colonization capacity of the strain, enabling it to outcompete pathogens.
Among 100 examined BAC clones roughly covering the A0 34/86 genome, one
reproducibly conferred on the laboratory strain DH10B an enhanced capacity
to persist in the intestine of newborn piglets. Sequencing revealed that
this BAC clone carried gene clusters encoding gluconate and mannonate
metabolism, adhesion (fim), invasion (ibe) and restriction/modification
functions. Hence, the genome of this clinically safe and highly efficient
colonizer strain appears to harbour many 'virulence-associated' genes.
These results highlight the thin line between bacterial 'virulence' and
'fitness' or 'colonization' factors, and question the definition of
enterobacterial virulence factors.

<>

<1>Helene, C.
<2>Sequence-selective recognition and cleavage of double-helical DNA.
<3>Curr. Opin. Biotechnol.
<4>4
<5>29-36
<6>1993
<7>Single sites within long double-helical DNA molecules can be recognized by a variety of
mechanisms. Different strategies have been used to adapt sequence-specific recognition to
sequence-specific cleavage of duplex DNA. Any nucleic acid can be converted into an artificial
nuclease by the attachment of a cleaving reagent. Alternatively, a sequence-specific ligand
can be used to protect a methylase recognition site from methylation. The protected site may
then be cleaved selectively by a restriction endonuclease (the so-called 'Achilles heel'
cleavage technique). Recent developments in this area have shown that it is possible to cleave
chromosomal DNA at single sites within bacterial and eukaryotic genomes.

<>

<1>Helling, R.B., Goodman, H.M., Boyer, H.W.
<2>Analysis of endonuclease R EcoRI fragments of DNA from lambdoid bacteriophages and other viruses by agarose-gel electrophoresis.
<3>J. Virol.
<4>14
<5>1235-1243
<6>1974
<7>By means of agarose-gel electrophoresis, endonuclease R EcoRI-generated
fragments of DNA from various viruses were separated, their molecular weights
were determined, and complete or partial fragment maps for lambda, Phi80, and
hybrid phages were constructed.

<>

<1>Hellinga, H.W., Smith, J.J., Jantz, D.
<2>New recombinant meganuclease with altered specificity for a recognition sequence half-site relative to a wild-type I-CreI, I-MsoI, I-SceI or I-CeuI meganuclease, useful for treating a prokaryotic infection in a eukaryotic host.
<3>International Patent Office
<4>WO 200747859
<5>
<6>2007
<7>DERWENT ABSTRACT: NOVELTY - A new recombinant meganuclease has altered specificity for at
least one recognition sequence half-site relative to
a wild-type I-Crel, I-MsoI, I-SceI or I-CeuI meganuclease. DETAILED
DESCRIPTION - The new recombinant meganuclease, having altered
specificity for at least one recognition sequence half-site relative to
a wild-type 1-Crel meganuclease, comprises a polypeptide having at
least 85% sequence similarity to residues 2-153 of the I-CreI
meganuclease comprising a fully defined 163-amino acid sequence (SEQ ID
NO: 1) and having specificity for a recognition sequence half-site
which differs by at least one base pair from a half-site within an
I-CreI meganuclease recognition sequence consisting of a fully defined
sequence comprising 22 (each of the sequences) amino acids (SEQ ID NO:
2-5); where the recombinant meganuclease comprises at least one
modification of Table 1 which is not an excluded modification. The new
recombinant meganuclease, having altered specificity for at least one
recognition sequence half-site relative to a wild-type I-MsoI
meganuclease, comprises a polypeptide having at least 85% sequence
similarity to residues 6-160 of the I-MsoI meganuclease comprising a
fully defined 170-amino acid sequence (SEQ ID NO: 6) and having
specificity for a recognition sequence half-site which differs by at
least one base pair from a half-site within an I-MsoI meganuclease
recognition sequence consisting of 22 (each of the sequences) amino
acids (SEQ ID NO: 7 or 8); where the recombinant meganuclease comprises
at least one modification of Table 2 which is not an excluded
modification. The new recombinant meganuclease, having altered
specificity for a recognition sequence relative to a wild-type I-SceI
meganuclease, comprises a polypeptide having at least 85% sequence
similarity to residues 3-186 of the I-SceI meganuclease comprising a
fully defined 235-amino acid sequence (SEQ ID NO: 9) and having
specificity for a recognition sequence which differs by at least one
base pair from an I-SceI meganuclease recognition sequence comprising a
fully defined sequence having 18-amino acid sequence each (SEQ ID NO:
10 or 11); where the recombinant meganuclease comprises at least one
modification of Table 3 which is not an excluded modification. The new
recombinant meganuclease having altered specificity for at least one
recognition sequence half-site relative to a wild-type I-CeuI
meganuclease, comprises a polypeptide having at least 85% sequence
similarity to residues 5-211 of the I-CeuI meganuclease comprising a
fully defined 218-amino acid sequence (SEQ ID NO: 12) and having
specificity for a recognition sequence half-site which differs by at
least one base pair from a half-site within an 1-Ceul meganuclease
recognition sequence consisting of a fully defined sequence having 22
amino acids each (SEQ ID NO: 13 or 14); where the recombinant
meganuclease comprises at least one modification of Table 4 which is
not an excluded modification. INDEPENDENT CLAIMS are: (1) a recombinant
meganuclease heterodimer; (2) a recombinant meganuclease monomer having
altered affinity for dimer formation with a reference meganuclease
monomer; (3) a method for producing a genetically-modified eukaryotic
cell including an exogenous sequence of interest inserted in a
chromosome of the eukaryotic cell; (4) a method for producing a
genetically-modified eukaryotic cell by disrupting a target sequence in
a chromosome of the eukaryotic cell; (5) a method of producing a
genetically-modified organism; (6) a method for treating a disease by
gene therapy in a eukaryote; (7) a method for treating a disease by
gene therapy in a eukaryote by disrupting a target sequence in a
chromosome of the eukaryotic cell;(8) a method for treating a viral
pathogen infection in a eukaryotic host by disrupting a target sequence
in a genome of the viral pathogen; (9) a method for treating a
prokaryotic pathogen infection in a eukaryotic host by disrupting a
target sequence in a genome

<>

<1>Hemavathy, K.C., Nagaraja, V.
<2>DNA methylation in mycobacteria: absence of methylation at GATC (Dam) and CCA/TGG (Dcm) sequences.
<3>FEMS Immunol Med Microbiol
<4>11
<5>291-296
<6>1995
<7>The presence of 6-methyladenine and 5-methylcytosine at Dam (GATC) and Dcm (CCA/TGG) sites in
DNA of mycobacterial species was investigated using
isoschizomer restriction enzymes. In all species examined, Dam and Dcm
recognition sequences were not methylated indicating the absence of these
methyltransferases. On the other hand, high performance liquid chromatographic
analysis of genomic DNA from Mycobacterium smegmatis and Mycobacterium
tuberculosis showed significant levels of 6-methyladenine and 5-methylcytosine
suggesting the presence of DNA methyltransferases other than Dam and Dcm.
Occurrence of methylation was also established by a sensitive genetic assay.

<>

<1>Hemeon, I., Gutierrez, J.A., Ho, M.-C., Schramm, V.L.
<2>Characterizing DNA methyltransferases with an ultrasensitive luciferase-linked continuous assay.
<3>Anal. Chem.
<4>83
<5>4996-5004
<6>2011
<7>DNA (cytosine-5)-methyltransferases (DNMTs) catalyze the transfer of a methyl group from
S-adenosyl-L-methionine (AdoMet) to the 5-position of cytosine residues and thereby silence
transcription of regulated genes. DNMTs are important epigenetic targets. However, isolated
DNMTs are weak catalysts and are difficult to assay. We report an ultrasensitive
luciferase-linked continuous assay that converts the S-adenosyl-L-homocysteine product of DNA
methylation to a quantifiable luminescent signal. Results with this assay are compared with
the commonly used DNA labeling from [methyl-3H]AdoMet. A
50-methylthioadenosine-adenosylhomocysteine nucleosidase is used to hydrolyze AdoHcy to
adenine. Adenine phosphoribosyl transferase converts adenine to AMP and pyruvate
orthophosphate dikinase converts AMP to ATP. Firefly luciferase gives a stable luminescent
signal that results from continuous AMP recycling to ATP. This assay exhibits a broad dynamic
range (0.11000 pmol of AdoHcy). The rapid response time permits continuous assays of DNA
methylation detected by light output. The assay is suitable for high-throughput screening of
chemical libraries for DNMT inhibition activity. The kinetic properties of human and bacterial
CpG methyltransferases are characterized using this assay. Human catalytic domain DNMT3b
activation byDNMT3L is shown to involve two distinct kinetic states that alter kcat but not Km
for AdoMet. The assay is shown to be
robust in the presence of high concentrations of the pyrimidine analogues 5-azacytidine and
5-azacytosine.

<>

<1>Hemme, C.L. et al.
<2>Sequencing of multiple clostridial genomes related to biomass conversion and biofuel production.
<3>J. Bacteriol.
<4>192
<5>6494-6496
<6>2010
<7>Modern methods to develop microbe-based biomass conversion processes require a system-level
understanding of the microbes involved. Clostridium
species have long been recognized as ideal candidates for processes
involving biomass conversion and production of various biofuels and other
industrial products. To expand the knowledge base for clostridial species
relevant to current biofuel production efforts, we have sequenced the
genomes of 20 species spanning multiple genera. The majority of species
sequenced fall within the class III cellulosome-encoding Clostridium and
the class V saccharolytic Thermoanaerobacteraceae. Species were chosen
based on representation in the experimental literature as model organisms,
ability to degrade cellulosic biomass either by free enzymes or by
cellulosomes, ability to rapidly ferment hexose and pentose sugars to
ethanol, and ability to ferment synthesis gas to ethanol. The sequenced
strains significantly increase the number of noncommensal/nonpathogenic
clostridial species and provide a key foundation for future studies of
biomass conversion, cellulosome composition, and clostridial systems
biology.

<>

<1>Henaut, A., Rouxel, T., Gleizes, A., Moszer, I., Danchin, A.
<2>Uneven distribution of GATC motifs in the Escherichia coli chromosome, its plasmids and its phages.
<3>J. Mol. Biol.
<4>257
<5>574-585
<6>1996
<7>This work reconsiders the GATC motif distribution in a 1.6 Mb segment of the Escherichia coli
genome, compared to its distribution in phages and plasmids.  At first sight the distribution
of GATC words looks random.  But when a realistic model of the chromosome (made of average
genes having the same codon usage as in the real chromosome), is used as a theoretical
reference, strong biases are observed.  GATC pairs such as GATCNNGATC are under-represented
while there is a strong positive selection for motifs separated by 10, 19, 70 and 1100 bp.
The last class is the only one present in E. coli parasites.  It can be ascribed to the
triggering sequences of the long-patch mismatch repair system.  The 6 bp class overlaps with
the consensus of CAP (catabolite activator protein) and FNR (fumarate/nitrate regulator)
binding sites, thus accounting for counter-selection.  The other classes, which could be
targets for a nucleic acid binding protein, are almost always present inside protein coding
sequences, and are members of clusters of GATC motifs.  Analysis of the genes containing these
motifs suggests that they correspond to a regulatory process monitoring the shift from
anaerobic to aerobic growth conditions  In particular this regulation, closing down
transcription of a large number of genes involved in intermediary metabolism would be well
suited for the cold and oxygen shift from the mammal's gut to the standard environmental
conditions.  In this process the methylation status of GATC clusters would be very important
for tuning transcription, and a DNA binding protein, probably a member of the cold-shock
proteins family would be needed for alleviating the effects mediated by slackening of the pace
of methylation during the shift.

<>

<1>Henderson, I.R., Deleris, A., Wong, W., Zhong, X.H., Chin, H.G., Horwitz, G.A., Kelly, K.A., Pradhan, S., Jacobsen, S.E.
<2>The De Novo Cytosine Methyltransferase DRM2 Requires Intact UBA Domains and a Catalytically Mutated Paralog DRM3 during RNA-Directed DNA Methylation in Arabidopsis thaliana.
<3>PLoS Genet.
<4>6
<5>e1001182
<6>2010
<7>Eukaryotic DNA cytosine methylation can be used to transcriptionally silence repetitive
sequences, including transposons and retroviruses.
This silencing is stable between cell generations as cytosine
methylation is maintained epigenetically through DNA replication. The
Arabidopsis thaliana Dnmt3 cytosine methyltransferase ortholog DOMAINS
REARRANGED METHYLTRANSFERASE2 (DRM2) is required for establishment of
small interfering RNA (siRNA) directed DNA methylation. In mammals PIWI
proteins and piRNA act in a convergently evolved RNA-directed DNA
methylation system that is required to repress transposon expression in
the germ line. De novo methylation may also be independent of RNA
interference and small RNAs, as in Neurospora crassa. Here we identify
a clade of catalytically mutated DRM2 paralogs in flowering plant
genomes, which in A. thaliana we term DOMAINS REARRANGED
METHYLTRANSFERASE3 (DRM3). Despite being catalytically mutated, DRM3 is
required for normal maintenance of non-CG DNA methylation,
establishment of RNA-directed DNA methylation triggered by repeat
sequences and accumulation of repeat-associated small RNAs. Although
the mammalian catalytically inactive Dnmt3L paralogs act in an
analogous manner, phylogenetic analysis indicates that the DRM and
Dnmt3 protein families diverged independently in plants and animals. We
also show by site-directed mutagenesis that both the DRM2 N-terminal
UBA domains and C-terminal methyltransferase domain are required for
normal RNA-directed DNA methylation, supporting an essential targeting
function for the UBA domains. These results suggest that plant and
mammalian RNA-directed DNA methylation systems consist of a combination
of ancestral and convergent features.

<>

<1>Hendrich, B., Bird, A.
<2>Identification and characterization of a family of mammalian methyl-CpG binding proteins.
<3>Mol. Cell. Biol.
<4>18
<5>6538-6547
<6>1998
<7>Methylation at the DNA sequence 5'-CpG is required for mouse development. MeCP2 and MBD1
(formerly PCM1) are two known proteins that bind specifically to methylated DNA via a related
amino acid motif and that can repress transcription. We describe here three novel human and
mouse proteins (MBD2, MBD3, and MBD4) that contain the methyl-CpG binding domain. MBD2 and
MBD4 bind specifically to methylated DNA in vitro. Expression of MBD2 and MBD4 tagged with
green fluorescent protein in mouse cells shows that both proteins colocalize with foci of
heavily methylated satellite DNA. Localization is disrupted in cells that have greatly reduced
levels of CpG methylation. MBD3 does not bind methylated DNA in vivo or in vitro. MBD1, MBD2,
MBD3, and MBD4 are expressed in somatic tissues, but MBD1 and MBD2 expression is reduced or
absent in embryonic stem cells which are known to be deficient in MeCP1 activity. The data
demonstrate that MBD2 and MBD4 bind specifically to methyl-CpG in vitro and in vivo and are
therefore likely to be mediators of the biological consequences of the
methylation signal.

<>

<1>Hendrich, B., Bird, A.
<2>Mammalian methyltransferases and methyl-CpG-binding domains: proteins involved in DNA methylation.
<3>Curr. Top. Microbiol. Immunol.
<4>249
<5>55-74
<6>2000
<7>The modified base 5-methylcytosine has been known to exist in mammalian DNA since 1950.  It
wasn't until 1988 that the gene encoding the enzyme reponsible for the maintenance of
5-methylcytosine in mammals, DNA-(cytosine-5) methyltransferase 1, was identified.  The
following year, a protein activity was reported; it was able to bind DNA containing methylated
cytosine followed by guanosine but, otherwise, it was indifferent to the sequence context.  A
different protein activity, which was also capable of binding the sequence MeCpG, was
identified in 1992, and the corresponding gene was cloned.  This provided the first molecular
handle on MeCpG-binding proteins.  For the following 5 years, however, no further proteins
were identified that were able to either methylate DNA or to bind specifically to methylated
DNA.  The past 2 years have seen a flurry of activity in this field, with the reporting of
three new candidate methyltransferases and four new candidate MeCPs.  Also developing is an
ever-more-precise molecular picture of exactly how DNA methylation affects transcription. In
this chapter, we will review that is known about the mammalian proteins involved in both
methylating DNA and in interpreting the signal that DNA methylation represents.  For an
evolutionary discussion of the known eukaryotic DNMTs, we refer the reader to a recent review
by Colot and Rossignol.  In this review, we will only discuss MeCPs that do not require
additional DNA sequence for specific binding to DNA.  For a summary of MeCPs in general we
refer the reader to a review by Tate and Bird.

<>

<1>Hendrickson, E.L. et al.
<2>Complete genome sequence of the genetically tractable hydrogenotrophic methanogen Methanococcus maripaludis.
<3>J. Bacteriol.
<4>186
<5>6956
<6>2004
<7>The genome sequence of the genetically tractable, mesophilic, hydrogenotrophic methanogen
Methanococcus maripaludis contains 1,722 protein-coding genes in a single circular chromosome
of 1,661,137 bp. Of the protein-coding genes (open reading frames [ORFs]), 44% were assigned a
function, 48% were conserved but had unknown or uncertain functions, and 7.5% (129 ORFs) were
unique to M. maripaludis. Of the unique ORFs, 27 were confirmed to encode proteins by the mass
spectrometric identification of unique peptides. Genes for most known functions and pathways
were identified. For example, a full complement of hydrogenases and methanogenesis enzymes was
identified, including eight selenocysteine-containing proteins, with each being paralogous to
a cysteine-containing counterpart. At least 59 proteins were predicted to contain iron-sulfur
centers, including ferredoxins, polyferredoxins, and subunits of enzymes with various redox
functions. Unusual features included the absence of a Cdc6 homolog, implying a variation in
replication initiation, and the presence of a bacterial-like RNase HI as well as an RNase HII
typical of the Archaea. The presence of alanine dehydrogenase and alanine racemase, which are
uniquely present among the Archaea, explained the ability of the organism to use L- and
D-alanine as nitrogen sources. Features that contrasted with the related organism
Methanocaldococcus jannaschii included the absence of inteins, even though close homologs of
most intein-containing proteins were encoded. Although two-thirds of the ORFs had their
highest Blastp hits in Methanocaldococcus jannaschii, lateral gene transfer or gene loss has
apparently resulted in genes, which are often clustered, with top Blastp hits in more
distantly related groups.

<>

<1>Hendrix, J.D., Welker, N.E.
<2>Isolation of a Bacillus stearothermophilus mutant exhibiting increased thermostability in its restriction endonuclease.
<3>J. Bacteriol.
<4>162
<5>682-692
<6>1985
<7>A procedure was developed for the selection of spontaneous mutants of Bacillus
stearothermophilus NUB31 that are more efficient than the wild type in the
restriction of phage at elevated temperatures.  Inactivation studies revealed
that two mutants contained a more thermostable restriction enzyme and one
mutant contained three times more enzyme than the wild type.  The restriction
endonucleases from the wild type and one of the mutants were purified to
apparent homogeneity.  The mutant enzyme was more thermostable than the
wild-type enzyme.  The subunit molecular weight, amino acid composition,
N-terminal and C-terminal amino acid residues, tryptic peptide map, and
catalytic properties of the two enzymes were determined.  The two enzymes have
similar catalytic properties, but the molecular size of the mutant enzyme is
approximately 6 to 7 kilodaltons larger than that of the wild-type enzyme.  The
mutant enzyme contains 54 additional amino acid residues, of which 26 to 28 are
aspartate/asparagine, 8 to 15 are glutamate/glutamine, and 8 to 9 are tyrosine
residues.  The two enzymes contained similar amounts of the other amino acids,
identical N-terminal residues, and different C-terminal residues.  Tryptic
peptide analyses revealed a high degree of homology between the two enzymes.
The increased thermostability observed in the mutant enzyme appears to have
been achieved by a mutation that resulted in the addition of amino acid
residues to the wild-type enzyme.  A number of mechanisms are discussed that
could account for the observed difference between the mutant and wild-type
enzymes.

<>

<1>Hendrix, R.W., Smith, M.C.M., Burns, R.N., Ford, M.E., Hatfull, G.F.
<2>Evolutionary relationships among diverse bacteriophages and prophages: All the world's a phage.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>2192-2197
<6>1999
<7>We report DNA and predicted protein sequence similarities, implying homology, among genes of
double-stranded DNA (dsDNA) bacteriophages and prophages spanning a broad phylogenetic range
of host bacteria. The sequence matches reported here establish genetic connections, not always
direct, among the lambdoid phages of Escherichia coli, phage phiC31 of Streptomyces, phages of
Mycobacterium, a previously unrecognized cryptic prophage, phiflu, in the Haemophilus
influenzae genome, and two small prophage-like elements, phiRv1 and phiRv2, in the genome of
Mycobacterium tuberculosis. The results imply that these phage genes, and very possibly all of
the dsDNA tailed phages, share common ancestry. We propose a model for the genetic structure
and dynamics of the global phage population in which all dsDNA phage genomes are mosaics with
access, by horizontal exchange, to a large common genetic pool but in which access to the gene
pool is not uniform for all phage.

<>

<1>Heng, N.C., Haji-Ishak, N.S., Kalyan, A., Wong, A.Y., Lovric, M., Bridson, J.M., Artamonova, J., Stanton, J.A., Wescombe, P.A., Burton, J.P., Cullinan, M.P., Tagg, J.R.
<2>Genome Sequence of the Bacteriocin-Producing Oral Probiotic Streptococcus salivarius Strain M18.
<3>J. Bacteriol.
<4>193
<5>6402-6403
<6>2011
<7>Streptococcus salivarius is a Gram-positive bacterial commensal and pioneer colonizer of the
human oral cavity. Many strains produce
ribosomally synthesized proteinaceous antibiotics (bacteriocins), and some
strains have been developed for use as oral probiotics. Here, we present
the draft genome sequence of the bacteriocin-producing oral probiotic S.
salivarius strain M18.

<>

<1>Henikoff, S., Comai, L.
<2>A DNA methyltransferase homolog with a chromodomain exists in multiple polymorphic forms in Arabidopsis.
<3>Genetics
<4>149
<5>307-318
<6>1998
<7>Chromodomains are thought to mediate protein-protein interactions between chromatin
components.  We have detected a chromodomain embedded within the catalytic region of a
predicted Arabidopsis DNA methyltransferase that is diverged from other eukaryotic enzymes.
The 791 residue "chromomethylase" is encoded by a floral transcript that is spliced from 20
exons and is present at only ~1/10^-7 of total mRNA.  Genomic sequencing reveals an ancient
haplotype split at CMT1 between Col-0 + Metz and the other ecotypes examined.  In the Col-0 +
Metz haplotype, alternative mRNA processing at intron 13 truncates the coding region.  In Ler,
RLD, and No-0, similar truncation is caused by insertion of an intact retrotransposon,
Evelknievel, which is present as a single copy in Ler and RLD and is currently methylated and
inactive.  Evelknievel is found at this site on a single branch that connects the Ler, RLD,
and No-0 ecotypes but is absent from the genomes of all other ecotypes examined.  A stop codon
within exon 6 of the Metz ecotype confirms that CMT1 is nonessential.  Nevertheless,
comparison to CMT1 of Cardaminopsis arenosa, an outcrossing relative, indicates conservation
for DNA methyltransferase function.  We discuss how allelic diversity of CMT1 may reflect
loosened selective constraints in a self-fertilizing species such as Arabidopsis thaliana.

<>

<1>Henke, R.M.
<2>Molecular and biochemical studies on a bifunctional group I intron encoded protein of yeast mitochondria.
<3>Diss. Abstr.
<4>61
<5>2899
<6>2000
<7>Intron 4a (aI4a) of the yeast mitochondrial COXI gene is a mobile group I intron that contains
a reading frame encoding both the homing endonuclease I-SceII and a latent maturase capable of
splicing both aI4a and the fourth intron of the cytochrome b (COB) gene (bI4).  The aI4a
reading frame is a member of a large gene family recognized by the presence of related
dodecapeptide sequence motifs called P1 and P2.  In this study missense mutations of P1 and P2
were placed in mtDNA by biolistic transformation and the effects of the mutations on intron
mobility, I-SceII activity and maturase function were tested.  The mutations of P1 strongly
affected intron mobility and I-SceII activity but had little or no effect on maturase
function, while mutations of P2 affected splicing but not mobility or I-SceII activity.
Surprisingly, the conditional (ts) mutations at P1 and P2 block one or the other function of
the protein but not both.  This study indicates that the two functions depend on separate
domains of the intron-encoded protein.  Normally splicing of the aI4a intron requires the bI4
maturase.  In strains lacking the bI4 maturase, second site-suppressors that activate the aI4a
maturase have been isolated; some map to the nuclear NAM2 gene (NAM2-1) and another to the
aI4a ORF (MIM2-1, glu 117) and then transformed into yeast mitochondria.  These changes were
designed to determine if activation of aI4a maturase results from the loss of a negative
charge or the gain of a positive charge at the mim2 site.  Substitution of a positively
charged amino acid (lys or arg) results in an active aI4a maturase, while the substitution of
either a negatively charged amino acid (wild-type, glu) or a neutral amino acid (gln) was
insufficient to activate the maturase.  These data show that the presence of a positive charge
at the mim2 residue is sufficient to activate aI4a's latent maturase activity.  The mim2
alleles were further characterized by assaying them for intron mobility.  All the mim2 alleles
retain wild-type levels of intron mobility, indicating that they encode I-SceII activity.  The
presence of both maturase and I-SceII activity in several alleles, (lys and arg) clearly
demonstrates that both functions can co-exist within the same poly-peptide.  This study also
characterizes several new in vitro DNA binding properties of I-SceII and speculates on their
in vivo significance.

<>

<1>Henke, R.M., Butow, R.A., Perlman, P.S.
<2>Maturase and endonuclease functions depend on separate conserved domains of the bifunctional protein encoded by the group I intron aI4a of yeast mitochondrial DNA.
<3>EMBO J.
<4>14
<5>5094-5099
<6>1995
<7>Intron 4a (aI4a) of the yeast mitochondrial COXI gene is a mobile group I intron that contains
a reading frame encoding both the homing endonuclease I-SceII and a latent maturase capable
of splicing both aI4a and the fourth intron of the cytochrome b (COB) gene (bI4).  The aI4a
reading frame is a member of a large gene family recognized by the presence of related
dodecapeptide sequence motifs called P1 and P2.  In this study, missense mutations of P1 and
P2 were placed in mitochondrial DNA by biolistic transformation.  The effects of the mutations
on intron mobility, endonuclease I-SceII activity and maturase function were tested.  The
mutations of P1 strongly affected mobility and endonuclease I-SceII activity, but had little
or no effect on maturase function; mutations of P2 affected splicing but not mobility or
endonuclease I-SceII activity.  Surprisingly, the conditional (temperature-sensitive)
mutations at P1 and P2 block one or the other function of the protein but not both.  This
study indicates that the two functions depend on separate domains of the intron-encoded
protein.

<>

<1>Henkel, C.V., den Dulk-Ras, A., Zhang, X., Hooykaas, P.J.
<2>Genome Sequence of the Octopine-Type Agrobacterium tumefaciens Strain Ach5.
<3>Genome Announcements
<4>2
<5>e00225-14
<6>2014
<7>We have sequenced the complete genome of the plant pathogen Agrobacterium tumefaciens strain
LBA4213, a derivative of the wild-type strain A. tumefaciens
Ach5 and the ancestor of A. tumefaciens strain LBA4404 used in genetic
engineering. The genome consists of a circular chromosome and a linear
chromosome, as well as a megaplasmid and a tumor-inducing plasmid.

<>

<1>Hennecke, F., Kolmar, H., Brundl, K., Fritz, H.-J.
<2>The vsr gene product of E. coli K-12 is a strand- and sequence-specific DNA mismatch endonuclease.
<3>Nature
<4>253
<5>776-778
<6>1991
<7>In Escherichia coli K-12, the Dcm methyltransferase catalyses methylation of the inner
cytosine residue in the sequence CCA/TGG. Hydrolytic deamination of 5-methylcytosine bases in
DNA leads to thymine residues, and hence to T/G mismatches, pre-mutagenic DNA lesions
consisting of two natural DNA constituents and thus devoid of an obvious marker of the damaged
DNA strand. These mismatches are corrected by the VSP repair pathway, which is characterized
by very short patches of DNA repair synthesis. It depends on genes vsr and polA and is
strongly stimulated by mutL and mutS. The vsr gene product (Vsr; Mr 18,000) was purified and
characterized as a DNA mismatch endonuclease, a unique and hitherto unknown type of enzyme.
Vsr endonuclease nicks double-stranded DNA within the sequence CTA/TGN or NTA/TGG next to the
underlined thymidine residue, which is mismatched to 2'-deoxyguanosine. The incision is
mismatch-dependent and strand-specific. These results illustrate how Vsr endonuclease
initiates VSP mismatch repair.

<>

<1>Henriques, A.O., Beall, B.W., Roland, K., Moran, C.P. Jr.
<2>Characterization of cotJ, a sigma E-controlled operon affecting the polypeptide composition of the coat of Bacillus subtilis spores.
<3>J. Bacteriol.
<4>177
<5>3394-3406
<6>1995
<7>The outermost protective structure found in endospores of Bacillus subtilis is a thick protein
shell known as the coat, which makes a key
contribution to the resistance properties of the mature spore and also
plays a role in its interaction with compounds able to trigger
germination. The coat is organized as a lamellar inner layer and an
electron-dense outer layer and has a complex polypeptide composition. Here
we report the cloning and characterization of an operon, cotJ, located at
about 62 degrees on the B. subtilis genetic map, whose inactivation
results in the production of spores with an altered pattern of coat
polypeptides. The cotJ operon was identified by screening a random library
of lacZ transcriptional fusions for a conditional (inducer-dependent) Lac+
phenotype in cells of a strain in which the structural gene (spoIIGB) for
the early-acting, mother-cell-specific transcriptional factor sigma E was
placed under the control of the IPTG
(isopropyl-beta-D-thiogalactopyranoside)-inducible Pspac promoter.
Sequence analysis of cloned DNA from the cotJ region complemented by
genetic experiments revealed a tricistronic operon preceded by a strong
sigma E-like promoter. Expression of an SP beta-borne cotJ-lacZ fusion
commences at around h 2 of sporulation, as does expression of other sigma
E-dependent genes, and shows an absolute requirement for sigma E. Studies
with double-reporter strains bearing a cotJ-gusA fusion and lacZ fusions
to other cot genes confirmed that expression of cotJ is initiated during
sporulation prior to activation of genes known to encode coat structural
proteins (with the sole exception of cotE). An in vitro-constructed
insertion-deletion mutation in cotJ resulted in the formation of spores
with no detectable morphological or resistance deficiency. However,
examination of the profile of electrophoretically separated spore coat
proteins from the null mutant revealed a pattern that was essentially
identical to that of a wild-type strain in the range of 12 to 65 kDa,
except for polypeptides of 17 and 24 kDa, the putative products of the
second (cotJB) and third (cotJC) cistrons of the operon, that were missing
or reduced in amount in the coat of the mutant. Polypeptides of the same
apparent sizes are detected in spores of a cotE null mutant, on which
basis we infer that the products of the cotJ operon are required for the
normal formation of the inner layers of the coat or are themselves
structural components of the coat. Because the onset of cotJ transcription is temporally
coincident with the appearance of active sigmaE, we speculate that cotJ-encoded products may
be involved in an early stage of coat assembly.

<>

<1>Henriques, A.O., Reis-Serra, C.A.D., Schyns, G.
<2>Probiotic.
<3>International Patent Office
<4>WO 2008110325 A
<5>
<6>2008
<7>The present invention relates to newly identified probiotics.  The invention also relates to
polynucleotide sequences comprising genes that encode proteins which are involved into
probiotic behavior.

<>

<1>Henriques, I., Juca, R.R.T., Barauna, R.A., de Sa, P.H., Marinho, A.D., Carneiro, A.R., Barbosa, S., Pereira, A., Alves, A., Saavedra, M.J., Egas, C., Silva, A., Correia, A.
<2>Draft Genome Sequence of Serratia fonticola UTAD54, a Carbapenem-Resistant Strain Isolated from Drinking Water.
<3>Genome Announcements
<4>1
<5>e00970-13
<6>2013
<7>Serratia fonticola UTAD54 is an environmental isolate that is resistant to carbapenems due to
the presence of a class A carbapenemase and a
metallo-beta-lactamase that are unique to this strain. Its draft genome sequence
was obtained to clarify the molecular basis of its carbapenem resistance and
identify the genomic context of its carbapenem resistance determinants.

<>

<1>Hensgens, L.A.M., Bonen, L., de Haan, M., van der Horst, G., Grivell, L.A.
<2>Two intron sequences in yeast mitochondrial COX1 gene:  homology among URF-containing introns and strain-dependent variation in flanking exons.
<3>Cell
<4>32
<5>379-389
<6>1983
<7>The DNA sequences of two optional introns in the gene for subunit I of cytochome c oxidase in
yeast mitochondrial DNA have been determined. Both contain long unassigned reading frames
(URFs). These display regions of amino acid homology with six other URFs, two of which encode
proteins involved in mitochondrial RNA splicing. Such conserved regions may thus define
functionally important domains of proteins involved in RNA processing. This homology also
implies that these URFs had a common ancestral sequence, which has been duplicated and
sipersed around the genome. Comparison of the flanking exons in the long strain KL14-4A with
their unsplit counterpart in D273-10B reveals clustered sequence differences, which lead in
D273-10B to codonas rearely used in exons. These differences may be linked to the loss or
absence of one of the optional introns.

<>

<1>Hensley, P., Nardone, G., Chirikjian, J.G., Wastney, M.E.
<2>The time-resolved kinetics of superhelical DNA cleavage by BamHI restriction endonuclease.
<3>J. Biol. Chem.
<4>265
<5>15300-15307
<6>1990
<7>The kinetics of the cleavage of superhelical plasmid DNA (pBR322) by the
restriction endonuclease, BamHI, have been analyzed in terms of a compartmental
model consistent with the chemistry first proposed by Rubin and Modrich (Rubin,
R.A., and Modrich, P. (1978) Nucleic Acids Res. 5, 2991-2997) for analysis of
the kinetics of the restriction endonuclease, EcoRI.  The model was defined in
terms of two compartments representing DNA substrate (bound and free), two
compartments representing nicked intermediate (bound and free), one compartment
representing linear product, and one compartment for free enzyme.  A
simultaneous analysis of concentration changes over time of the three DNA forms
(superhelical, nicked, and linear) at six different enzyme concentrations was
undertaken employing this compartmental model using SAAM (Simulation Analysis
and Modeling) software.  Results showed that rate constants characterizing the
association of enzyme with superhelical DNA (6.0 x 10/5M-1S-1) and nicked DNA
(2.8 x 10/5M-1S-1) were similar in magnitude and rate constants characterizing
cleavage of the first (1.2 x 10-2S-1) and second phosphodiester bonds (3.1 x
10-2S-1) were also similar.  The analysis yields a kinetically determined
equilibrium constant of 12.9 nM for the dissociation of nicked intermediate
from the enzyme.  The rate constant describing the release of the nicked
intermediate from the enzyme has a value of 3.7 x 10-3S-1.  By comparing the
value of this release rate constant to the value of the constant describing the
second cleavage event, it can be determined that only 10% of the nicked
intermediate bound to the enzyme is released as free nicked DNA and that 90% of
the nicked intermediate is processed to the linear form without being released.
Hence, most of the DNA is cleaved as the result of a single enzyme-DNA
recognition event.  No steady state assumptions were made in the analysis.  The
approach was to directly solve the differential equations which described the
kinetic processes using an interactive method.  This study demonstrates the
usefulness of this approach for the analysis of kinetics of protein-DNA
interactions for the restriction endonucleases.

<>

<1>Hentosh, P., McCastlain, J.C.
<2>Effects of 2-chloroadenine substitution in DNA on restriction endonuclease cleavage reactions.
<3>Nucleic Acids Res.
<4>19
<5>3143-3148
<6>1991
<7>The purine analog, 2-chloro-2'-deoxyadenosine triphosphate (CldATP), was
incorporated enzymatically in place of dATP into the minus strand of M13mp18
duplex DNA.  Its effect on protein-DNA interactions was assessed by determining
the amount of DNA cleavage by type II restriction endonucleases.  Substitution
of chloroadenine (ClAde) for adenine (Ade) in DNA appreciably decreased the
amount and rate of DNA cleavage of the minus strand when the analog was
situated within the appropriate endonuclease recognition site.  ClAde residues
flanking a restriction site had variable effects.  SmaI cleaved both
ClAde-containing and control substrates with equal efficiency.  NarI, however,
was stimulated 1.5-fold by the presence of ClAde outside its recognition site.
The effects of analog incorporation on restriction enzyme cleavage of an
opposing unsubstituted strand of duplex DNA was examined by enzymatically
incorporating CldATP into complementary minus strand of a 36-base
oligonucleotide.  Endonucleolytic cleavage of both plus and minus strands was
reduced on 36-mers containing ClAde residues located within only the minus
strand.  These data suggest that ClAde residues incorporated into a single DNA
strand may have an appreciable effect of DNA-protein interactions that involve
one or both strands of duplex DNA.

<>

<1>Hepburn, P.A., Margison, G.P., Tisdale, M.J.
<2>Enzymatic methylation of cytosine in DNA is prevented by adjacent 06-methylguanine residues.
<3>J. Biol. Chem.
<4>266
<5>7985-7987
<6>1991
<7>The effect of 06-alkylation of guanine residues on the enzymatic methylation of
cytosine has been studied using synthetic oligonucleotides in which all
guanines in cytosine-guanine sequences at potentially methylatable sites are
replaced by 06-methylguanine.  In contrast with the unmodified forms, which
showed high acceptance activity for methyl-3H-labeled groups from
S-adenosyl-L-[methyl-3H]methionine in the presence of DNA methylase, the
modified oligonucleotides were not substrates for the enzyme neither in the
single-stranded or annealed forms.  In view of the importance of cytosine
methylation in the down-regulation of certain genes, the potential to affect
gene expression by this mechanism may be a contributory factor in the toxic and
carcinogenic effects of chemical methylating agents.

<>

<1>Hepburn, P.A., Tisdale, M.J.
<2>Importance of the O6 position of guanine residues in the binding of DNA methylase to DNA.
<3>Biochim. Biophys. Acta
<4>1088
<5>341-344
<6>1991
<7>Methylation of Micrococcus lysodeikticus DNA by purified DNA methylase isolated
from L1210 leukaemia cells is potently and specifically inhibited by both
hetero and homoribo and deoxyribopolynucleotides containing guanine residues.
The inhibitory effect is unaffected by chain length, but is abolished when the
O6 residue of guanine is substituted as in poly[d(O6MeG)]20.  Potent inhibition
is also shown by polyinosinic and polyxanthylic acids, but not by polyadenylic
acid or by heteropolymers containing adenine and thymine.  These results
suggest that the 6-position of the purine nucleus is important in binding of
the DNA methylase to a particular region of the DNA duplex and that the
hydrogen bonding properties of this group are important in enzyme recognition.

<>

<1>Hepworth, P.J., Ashelford, K.E., Hinds, J., Gould, K.A., Witney, A.A., Williams, N.J., Leatherbarrow, H., French, N.P., Birtles, R.J., Mendonca, C., Dorrell, N., Wren, B.W., Wigley, P., Hall, N., Winstanley, C.
<2>Genomic variations define divergence of water/wildlife-associated Campylobacter jejuni niche specialists from common clonal complexes.
<3>Environ. Microbiol.
<4>13
<5>1549-1560
<6>2011
<7>Although the major food-borne pathogen Campylobacter jejuni has been
isolated from diverse animal, human and environmental sources, our
knowledge of genomic diversity in C. jejuni is based exclusively on human
or human food-chain-associated isolates. Studies employing multilocus
sequence typing have indicated that some clonal complexes are more
commonly associated with particular sources. Using comparative genomic
hybridization on a collection of 80 isolates representing diverse sources
and clonal complexes, we identified a separate clade comprising a group of
water/wildlife isolates of C. jejuni with multilocus sequence types
uncharacteristic of human food-chain-associated isolates. By genome
sequencing one representative of this diverse group (C. jejuni 1336), and
a representative of the bank-vole niche specialist ST-3704 (C. jejuni
414), we identified deletions of genomic regions normally carried by human
food-chain-associated C. jejuni. Several of the deleted regions included
genes implicated in chicken colonization or in virulence. Novel genomic
insertions contributing to the accessory genomes of strains 1336 and 414
were identified. Comparative analysis using PCR assays indicated that
novel regions were common but not ubiquitous among the water/wildlife
group of isolates, indicating further genomic diversity among this group,
whereas all ST-3704 isolates carried the same novel accessory regions.
While strain 1336 was able to colonize chicks, strain 414 was not,
suggesting that regions specifically absent from the genome of strain 414
may play an important role in this common route of Campylobacter infection
of humans. We suggest that the genomic divergence observed constitutes
evidence of adaptation leading to niche specialization.

<>

<1>Herbert, M., Deadman, M., Hood, D., Moxon, E.
<2>Virulence genes in H. influenzae.
<3>International Patent Office
<4>WO 02077020 A
<5>
<6>2002
<7>Virulence genes in H. influenzae are identified using signature tagged mutagenesis (STM).  The
genes and their encoded products may be useful in the production of vaccines or antibiotics to
prevent or treat H. influenzae infection.

<>

<1>Herbert, M., O'Keeffe, T.A., Purdy, D., Elmore, M., Minton, N.P.
<2>Gene transfer into Clostridium difficile CD630 and characterisation of its methylase genes.
<3>FEMS Microbiol. Lett.
<4>229
<5>103-110
<6>2003
<7>Ignorance of pathogenesis in Clostridium difficile may be attributable to a lack of effective
genetic tools. We have now shown that oriT-based
shuttle vectors may be conjugated from Escherichia coli donors to the C.
difficile strain CD630, at frequencies of around 10(-6) transconjugants
per donor cell. Transfer is unaffected by either sequences present on the
vector or its methylation status. Whilst the genome of this strain carries
five methylase genes, there is no in silico or experimental evidence for
cognate restriction enzymes. It would seem that the identified methylases
do not participate in restriction-modification, and must, therefore,
fulfil another role. A similar situation most likely applies to other
clostridia.

<>

<1>Herd, M., Kocks, C.
<2>Gene fragments distinguishing an epidemic-associated strain from a virulent prototype strain of Listeria monocytogenes belong to a distinct   functional subset of genes and partially cross-hybridize with other   Listeria species.
<3>Infect. Immun.
<4>69
<5>3972-3979
<6>2001
<7>Most major food-borne outbreaks of listeriosis in Europe and in the United States have been
caused by genetically closely related Listeria
monocytogenes strains of serotype 4b. In order to assess whether genomic
loci exist that could underlie this increased epidemic potential, we
subtracted the genome of the virulent prototype L. monocytogenes strain
EGD from a prototype epidemic strain. A total of 39 DNA fragments
corresponding to 20% of an estimated total of 150 to 190 kb of
differential genome material were isolated. For 21 of these fragments, no
function on the basis of homology could be predicted. Of the remaining 18
fragments, 15 had homologies to bacterial surface proteins, some of which
have been implicated in virulence mechanisms such as cell invasion,
adhesion, or immune escape. Southern hybridization of arrays containing
the epidemic-clone-specific DNA segments with genomic DNA of different L.
monocytogenes strains was consistent with the current lineage division.
Surprisingly, however, some of the fragments hybridized in a mosaic-like
fashion to genomes of two other Listeria species, the animal pathogen L.
ivanovii and the nonpathogen L. innocua. Taken together, our results
provide a starting point for the identification of
epidemic-trait-associated genes.

<>

<1>Herman, G.E., Modrich, P.
<2>Escherichia coli K-12 clones that overproduce dam methylase are hypermutable.
<3>J. Bacteriol.
<4>145
<5>644-646
<6>1981
<7>A strain of Escherichia coli K-12 that overproduces dam methylase 50-fold was found to be
hypermutable, and mutations which resulted in loss of excess methylase activity restored
mutation frequencies to wild-type levels. These results are consistent with involvement of
this deoxyribonucleic acid methylase in mismatch correction.

<>

<1>Herman, G.E., Modrich, P.
<2>Escherichia coli dam methylase physical and catalytic properties of the homogeneous enzyme.
<3>J. Biol. Chem.
<4>257
<5>2605-2612
<6>1982
<7>The Escherichia coli dam methylase has been purified 3000-fold to a purity of 95% from a clone
which overproduces the enzyme 10- to 20-fold. Physical properties of enzyme purified from the
overproducing clone were identical with those of enzyme previously obtained from a
non-overproducing E. coli strain (Geier, G.E., and Modrich, P. (1979) J. Biol. Chem. 254,
1408-1413). The methylase is comprised of a single polypeptide chain of Mr = 31,000 has an
S20,W of 2.8 S, a Stokes radius of 24 A, and exists in solution as a monomer. Its aggregation
state is not affected by the presence of S-adenosyl-L-methionine. The simple kinetic behavior
of the methylase indicates that it functions as a monomer. Initial rates of methyl transfer
are first order in enzyme concentration, and Michaelis-Menten behavior is obeyed with respect
to both substrates. At 37C, in the presence of saturating DNA, the enzyme has a turnover
number of 19 methyl transfers/min with a KM for S-adenosyl-L-methionine of 12.2 lM. At
half-saturating S-adenosyl-L-methionine, the apparent KM for d(G-A-T-C) sites in ColE1 DNA is
3.6NM. The mechanism of methyl transfer is also consistent with the monomer being the
functional form of the enzyme. Studies with G4 RFI DNA (two d(G-A-T-C) sites) indicate that
the methylase transfers 1 methyl group to a recognition site and then dissociates from this
DNA prior to subsequent catalysis. It appears that kinetic parameters for methyl transfer to
sites already modified on one DNA strand may be slightly more favorable than those for
transfer to sites in which both strands are unmethylated.

<>

<1>Herman, J., Nelkin, B., Mabry, M., Wilson, G., Baylin, S.
<2>Introduction and expression of HhaI DNA methyltransferase in eukaryotic cells.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>223
<6>1989
<7>DNA methylation abnormalities, which include both widespread hypomethylation
and regional hypermethylation, hve been reported for many malignancies.  In
culture, tumor cells often have increased DNA methyltransferase activity with
unknown consequences.  To assess functional consequences of increased cytosine
methylation in eukaryotic cells, we are using a constitutively expressed
prokaryotic DNA methyltransferase enzyme in murine fibroblasts.  We inserted
the cloned DNA sequence for the bacterial DNA methyltransferase M.HhaI
(methylated sequence G^mCGC) into the retroviral expression vector pZIPneo
SV(X).  Abundant G418 resistant colonies were produced in PA 317 amphotropic
packaging cells after infection with either the pZIPneo SV(X) vector alone or
M.HhaI inserted in the antisense direction.  However, insertion of M.HhaI in
the sense orientation resulted in a total of two G418 resistant clones in 5
independent experiments.  Southern and Northern blots probed with M.HhaI
sequences demonstrated integration and expression.  Each M.HhaI clone in the
sense orientation was tumorigenic in nude mice, and one of the clones was
morphologically distinct from control PA 317 cells.  Other data suggest that
constitutive expression of M.HhaI is often lethal to eukaryotic cells in
culture, and may cause transformation of surviving cells.

<>

<1>Herman, J., Nelkin, B., Mabry, M., Wilson, G., Baylin, S.
<2>Introduction of prokaryotic HhaI DNA methyltransferase alters the phenotype of 3T3 cells.
<3>Proc. Amer. Assoc. Cancer Res.
<4>30
<5>428
<6>1989
<7>DNA methylation abnormalities are common in cancer.  Recently, we described
hypermethylation on chromosome 11p (PNAS USA 85:5693-5697, 1988) which could
result from the fact that tumor cells often have increased DNA
methyltransferase activity.  To assess the functional consequences of
increasing cytosine methylation capacity in eukaryotic cells, we have
constitutively expressed two prokaryotic DNA methyltransferase enzymes in
murine fibroblasts.  DNA sequences for the bacterial DNA methyltransferases
M.HhaI (methylated sequence GmCGC), and M.HpaII (CmCGG) were introduced via the
retroviral expression vector pZIPneo SV(X).  Abundant G418 resistant colonies,
typically 60/experiment, were produced in PA317 amphotropic packaging cells
after infection with the pZIPneo SV(X) vector alone, the M.HhaI inserted in the
antisense direction, and M.HpaII in both directions.  However, insertion of
M.HhaI in the sense orientation resulted in a total of only two G418 resistant
clones in 5 independent experiments.  M.HhaI sequences were integrated and
expressed in each clone.  Both clones were tumorigenic in nude mice, and one of
the clones was morphologically distinct from control PA317 cells.  We also
observed a similar discrepancy in the ability to select stably transfected Psi2
cells used to infect the PA317 cells with the sense M.HhaI construct.  Our data
suggest that constitutive expression of M.HhaI is most often lethal to 3T3
cells in culture, and may cause transformation of those cells which survive.

<>

<1>Hermann, A., Fatemi, M., Jeltsch, A.
<2>Molecular enzymology of the Dnmt1 DNA methyltransferase explains the mechanism of cis-spreading of DNA methylation.
<3>Biochem. Soc. Trans.
<4>28
<5>A248
<6>2000
<7>In mammals, methylation of DNA within CpG-sequences is involved in epigenetic control of gene
expression, chromatin condensation and genetic imprinting.  So far, three active DNA
methyltransferases have been identified in mice: Dnmt1 which is responsible for maintenance
methylation, as well as Dnmt3a and 3b which are required for de novo methylation.  Methylation
of DNA is essential in mammals because mice deficient in either Dnmt1, Dnmt3a or Dnmt3b die
during development.  During tumorigenesis or aging, spreading of methylation is observed, i.e.
starting from one modified CpG-site methylation spreads to neighboring CpGs until one region
of the DNA is completely methylated.  The Dnmt1 enzyme consists of a catalytic domain (500
amino acid residues) which closely resembles prokaryotic DNA-(cytosine-C5)-methyltransferases
and a large regulatory N-terminal domain comprising about 1100 amino acid residues.  By
cloning and characterizing several fragments of the Dnmt1 enzyme, we demonstrate that the
N-terminal domain forms a DNA binding site distinct from the active center.  In kinetic assays
with purified full-length Dnmt1, we show that binding of methylated CpGs to this additional
DNA binding site allosterically activates the enzyme.  These enzymological properties of Dnmt1
can explain the phenomenon of spreading of DNA methylation.

<>

<1>Hermann, A., Gowher, H., Jeltsch, A.
<2>Biochemistry and biology of mammalian DNA methyltransferases.
<3>Cell. Mol. Life Sci.
<4>61
<5>2571-2587
<6>2004
<7>DNA methylation is a stable but not irreversible epigenetic signal that silences gene
expression. It has a variety of important functions in mammals, including control of gene
expression, cellular differentiation and development, preservation of chromosomal integrity,
parental imprinting and X-chromosome inactivation. In addition, it has been implicated in
brain function and the development of the immune system. Somatic alterations in genomic
methylation patterns contribute to the etiology of human cancers and ageing. It is tightly
interwoven with the modification of histone tails and other epigenetic signals. Here we review
our current understanding of the molecular enzymology of the mammalian DNA methyltransferases
Dnmt1, Dnmt3a, Dnmt3b and Dnmt2 and the roles of the enzymes in the above-mentioned biological
processes.

<>

<1>Hermann, A., Goyal, R., Jeltsch, A.
<2>The Dnmt1 DNA-(cytosine-C5)-methyltransferase methylates DNA processively with high preference for hemimethylated target sites.
<3>J. Biol. Chem.
<4>279
<5>48350-48359
<6>2004
<7>In the cell Dnmt1 is the major enzyme to maintain the pattern of DNA methylation after DNA
replication. Evidence suggests the protein is
located at the replication fork where it could directly modify nascent DNA
immediately after replication. To elucidate the potential mechanism of
this process, we investigate the processivity of DNA methylation and
accuracy of copying an existing pattern of methylation in this study using
purified Dnmt1 and hemimethylated substrate DNA. We demonstrate that Dnmt1
methylates a hemimethylated 958mer substrate in a highly processive
reaction. Fully methylated and unmethylated CG sites do not inhibit
processive methylation of the DNA. Extending previous work, we show that
unmethylated sites embedded in a hemimethylated context are modified at an
approx. 24-fold reduced rate which demonstrates that the enzyme accurately
copies existing patterns of methylation. Completely unmodified DNA is
methylated even more slowly due to an allosteric activation of Dnmt1 by
methylcytosine-containing DNA. Interestingly, Dnmt1 is not able to
methylate hemimethylated CG sites on different strands of the DNA in a
processive manner indicating that Dnmt1 keeps its orientation with respect
to the DNA while methylating the CG sites on one strand of the DNA.

<>

<1>Hermann, A., Jeltsch, A.
<2>Methylation sensitivity of restriction enzymes interacting with GATC sites.
<3>Biotechniques
<4>34
<5>924-930
<6>2003
<7>DNA methylation plays an important role in many species ranging from bacteria to man, by
controlling the expression of genes and protection of the genome from selfish DNA like
transposons and viruses.  In E. coli and other gamma-proteobacteria, methylation of adenine
residues (dam, DNA adenine methylation) occurs at GATC sites.  It serves to discriminate
between parental and daughter strand during post-replicative mismatch repair, to coordinate
cell cycle, DNA replication, and to regulate gene expression.  It also controls the
pathogenicity of different gamma-proteobacteria.  To investigate the methylation state of DNA
from various bacteria, archaea as well as lower and higher eukaryotes, digestion of the DNA
with GATC-interacting restriction enzymes is often used.

<>

<1>Hermann, A., Schmitt, S., Jeltsch, A.
<2>The human Dnmt2 has residual DNA-(cytosine-C5)-methyltransferase activity.
<3>J. Biol. Chem.
<4>278
<5>31717-31721
<6>2003
<7>The human Dnmt2 protein is one member of a protein family conserved from Schizosaccharomyces
pombe and Drosophila melanogaster to Mus musculus and
Homo sapiens. It contains all of the amino acid motifs characteristic for
DNA-(Cytosine-C5) methyltransferases, and its structure is very similar to
prokaryotic DNA methyltransferases. Nevertheless, so far all attempts to
detect catalytic activity of this protein have failed. We show here by two
independent assay systems that the purified Dnmt2 protein has weak DNA
methyltransferase activity. Methylation was observed at CG sites in a
loose ttnCGga(g/a) consensus sequence, suggesting that Dnmt2 has a more
specialized role than other mammalian DNA methyltransferases.

<>

<1>Hernandez, D., Seidl, K., Corvaglia, A.R., Bayer, A.S., Xiong, Y.Q., Francois, P.
<2>Genome Sequences of Sequence Type 45 (ST45) Persistent Methicillin-Resistant Staphylococcus aureus (MRSA) Bacteremia Strain 300-169 and ST45 Resolving MRSA  Bacteremia Strain 301-188.
<3>Genome Announcements
<4>2
<5>e00174-14
<6>2014
<7>Persistent methicillin-resistant Staphylococcus aureus (MRSA) bacteremia (positive blood
cultures after >/=7 days) represents a challenging subset of
invasive MRSA infections. The comparison of genome sequences of persistent
(300-169) and resolving (301-188) MRSA bacteremia isolates with similar genetic
background (sequence type 45 [ST45]) will help us to better understand underlying
mechanisms of persistent MRSA bacteremia.

<>

<1>Hernandez, D., van der Mee-Marquet, N., Kluytmans, J., Donnio, P.Y., Quentin, R., Corvaglia, A.R., Francois, P.
<2>Whole-Genome Sequences of Staphylococcus aureus ST398 Strains of Animal Origin.
<3>Genome Announcements
<4>1
<5>e00689-13
<6>2013
<7>Staphylococcus aureus sequence type 398 (ST398) was originally associated with animal
infections. We announce the complete genome sequences of two ST398
methicillin-susceptible S. aureus strains from the livestock environment. These
genome sequences assist in the characterization of interesting ST398 features
relying on host tropism and epidemiological settings.

<>

<1>Hernandez-Mendoza, A., Martinez-Ocampo, F., Lozano-Aguirre, B.L.F., Popoca-Ursino, E.C., Ortiz-Hernandez, L., Sanchez-Salinas, E., Dantan-Gonzalez, E.
<2>Draft Genome Sequence of the Organophosphorus Compound-Degrading Burkholderia zhejiangensis Strain CEIB S4-3.
<3>Genome Announcements
<4>2
<5>e01323-14
<6>2014
<7>Burkholderia species are widely distributed in the environment. A Burkholderia zhejiangensis
strain was isolated from pesticide-contaminated soil from an
agricultural field in Mexico and identified as an organophosphorus
compound-degrading bacterium. In this study, we report the draft genome sequence
of Burkholderia zhejiangensis strain CEIB S4-3.

<>

<1>Herrin, D.L., Chen, Y.-F., Schmidt, G.W.
<2>RNA splicing in Chlamydomonas chloroplasts.
<3>J. Biol. Chem.
<4>265
<5>21134-21140
<6>1990
<7>The 23 rRNA gene of the Chlamydomonas reinhardtii chloroplast contains an 888-base pair intron
with structural features characteristic of Group I introns. The nuclear, chloroplast
ribosome-deficient mutant of C. reinhardtii, ac20, overaccumulates an approx. 3.6-kilobase
unspliced 23 S preRNA compared to wild-type cells. We have used [a-32P]GTP labeling of total
RNA preparations from ac20 to rapidly determine that 23 S preRNA is capable of self-splicing.
The ability of the 23 S intron (with flanking exon sequences) to correctly catalyze its own
splicing was confirmed using RNA produced by in vitro transcription of cloned DNA. These
results identify the first example of a self-splicing RNA of chloroplast origin.

<>

<1>Hershberge, C.L., Rosteck, P.R.
<2>Protecting bacteria from phage infection by transformation with cloning vector containing segment with restriction and modification activity.
<3>United Kingdom Patent Office
<4>GB 2126237 A
<5>
<6>1982
<7>Recombinant DNA cloning vector comprises a DNA segment that confers restriction and a cognate
modification activity to a bacterium; a replicon which functions in the given bacterium; and a
gene which expresses a functional polypeptide in the same bacterium; such that the
modification activity is expressed in the bacterium before restriction; and that the gene and
the DNA segment do not confer antibiotic resistance to the bacterium. The expressed
polypeptide may be human proinsulin.

<>

<1>Hertwig, S., Klein, I., Schmidt, V., Beck, S., Hammerl, J.A., Appel, B.
<2>Sequence analysis of the genome of the temperate Yersinia enterocolitica phage PY54.
<3>J. Mol. Biol.
<4>331
<5>605-622
<6>2003
<7>The temperate Yersinia phage PY54 belongs to the unusual group of phages that replicate as
linear plasmids with covalently closed ends. Besides
Escherichia coli phage N15, PY54 is the only member of this group to be
identified. We have determined the complete sequence (46,339 bp) of the
PY54 genome. Bioinformatic analyses revealed 67 open reading frames (ORFs)
with good coding potential located on both DNA strands. The comparison of
the deduced PY54 gene products with known proteins encoded by other phages
and bacteria along with functional studies have enabled us to assign the
possible functions of 25 ORFs. In the left arm of the PY54 genome, we
identified a number of ORFs that obviously code for head and tail
proteins. Furthermore, this part of the phage genome contains genes
probably involved in plasmid partitioning. Regarding the predicted gene
functions and gene order, the PY54 and N15 left arms are similar. However,
there are only weak DNA homologies and, in contrast to N15, the Yersinia
phage harbours only a few ORFs related to genes found in lambdoid phages.
The PY54 right arm comprises mainly regulatory genes as well as genes
important for plasmid replication, DNA methylation, and host cell lysis.
Out of 36 deduced products of the right arm, 13 revealed strongest
database homologies to N15 proteins, of which the protelomerase and the
Rep protein are exclusively homologous to their N15 counterparts. A number
of PY54 genes essential for the lytic or lysogenic cycle were identified
by functional analysis and characterization of phage mutants. In order to
study transcription during the lytic and lysogenic stage, we analysed 34
PY54 ORFs by reverse transcriptase (RT)-PCR. The phage transcription
patterns in lysogenic bacteria and at the late lytic stage of infection
are nearly identical. The reasons for this finding are spontaneous release
of phages during lysogeny and a high rate of phages that lysogenize their
Yersinia host upon infection.

<>

<1>Hess, J.F., FitzGerald, P.G.
<2>Protection of a restriction enzyme from heat inactivation by alpha-crystallin.
<3>Mol. Vis.
<4>4
<5>29-32
<6>1998
<7>To determine whether the chaperone activity of human alpha-crystallin can protect a
restriction enzyme from heat inactivation.  The restriction enzyme NdeI was heated in the
presence or absence of purified bovine alpha-crystallin.  Following heat treatment, the
enzymatic activity of the heat treated samples was assayed by cleavage of plasmid DNA.  The
extent of digestion was monitored by agarose gel electrophoresis and visualization of DNA
fragments by ethidium bromide staining.  Heating of NdeI in the absence of alpha-crystallin
resulted in inactivation.  However, NdeI heated in the presence of alpha-crystallin remained
active.  Furthermore, an increased amount of alpha-crystallin provided a longer period of
thermal protection.  The chaperone activity and thermo-protective effect of alpha-crystallin
extend to protection of enzymatic activity, not merely the protection from thermally induced
aggregation/denaturation.  In addition, inclusion of alpha-crystallin during some enzymatic
reactions may be beneficial.

<>

<1>Heuer, H., Kopmann, C., Binh, C.T., Top, E.M., Smalla, K.
<2>Spreading antibiotic resistance through spread manure: characteristics of a novel plasmid type with low %G+C content.
<3>Environ. Microbiol.
<4>11
<5>937-949
<6>2009
<7>Bioactive amounts of antibiotics as well as resistant bacteria reach the
soil through manure fertilization. We investigated plasmids that may
stimulate the environmental spread and interspecies transfer of antibiotic
resistance. After treatment of two soils with manure, either with or
without the sulfonamide antibiotic sulfadiazine, a significant increase in
copies of the sulfonamide resistance gene sul2 was detected by qPCR. All
sul2 carrying plasmids, captured in Escherichia coli from soil, belonged
to a novel class of self-transferable replicons. Manuring and sulfadiazine
significantly increased the abundance of this replicon type in a
chemically fertilized but not in an annually manured soil, as determined
by qPCR targeting a transfer gene. Restriction patterns and antibiograms
showed a considerable diversity within this novel plasmid group. Analysis
of three complete plasmid sequences revealed a conserved 30 kbp backbone
with only 36% G+C content, comprised of transfer and maintenance genes
with moderate homology to plasmid pIPO2 and a replication module (rep and
oriV) of other descent. The plasmids differed in composition of the
27.0-28.3 kbp accessory region, each of which carried ISCR2 and several
resistance genes. Acinetobacter spp. was identified as a potential host of
such LowGC-type plasmids in manure and soil.

<>

<1>Heuer, H., Szczepanowski, R., Schneiker, S., Puhler, A., Top, E.M., Schluter, A.
<2>The complete sequences of plasmids pB2 and pB3 provide evidence for a recent ancestor of the IncP-1{beta} group without any accessory genes.
<3>Microbiology
<4>150
<5>3591-3599
<6>2004
<7>The nucleotide sequences of the broad-host-range antibiotic resistance
plasmids pB2 (61 kb) and pB3 (56 kb), which were isolated from a
wastewater treatment plant, were determined and analysed. Both have a
nearly identical IncP-1beta backbone, which diverged early from the
sequenced IncP-1beta plasmids R751, pB10, pJP4, pADP1 and pUO1. In
contrast to the latter plasmids, the pB2 and pB3 backbone does not seem to
have undergone any deletions. The complete partition gene parA is located
downstream of the mating pair formation (trb) module. A 14.4 kb or 19.0 kb
mobile genetic element is present between traC and parA of pB3 and pB2,
respectively. This region is typical for insertions in IncP-1beta
plasmids, but the insertion site is unique. Both elements differ only by a
duplication in pB2 of a tetA(C)-tetR-tnpA(IS26) fragment. The 5 bp target
site duplication and the 26 bp inverted repeats flanking the mobile
genetic elements are still intact, indicating that the insertion occurred
recently. The element consists of three nested transposable elements: (i)
a relict of a Tn402-like transposon with a gene for a new class D
beta-lactamase (bla(NPS-2)); (ii) within that, another Tn402-like element
with a class 1 integron harbouring the gene cassettes cmlA1 for a
chloramphenicol efflux protein and aadA2 encoding a
streptomycin/spectinomycin adenylyltransferase, and a copy of IS6100;
(iii) into the integrase gene intI1 a tetracycline resistance module
tetA(C)-tetR flanked by copies of IS26 is inserted. Interestingly, in
contrast to all other IncP-1beta plasmids analysed so far, the oriV region
between trfA and klcA is not interrupted by accessory genes, and there is
no indication that previously inserted accessory genes have subsequently
been deleted. The genes kluAB are also missing in that region and should
thus be considered acquired genes. These findings, together with the fact
that IncP-1beta plasmids acquired accessory elements at various positions
in the backbone, suggest that IncP-1beta plasmids without any accessory
genes exist in microbial communities. They must occasionally acquire
accessory genes by transposition events, resulting in those plasmids that
have been found based on selectable phenotypic traits.

<>

<1>Heuermann, D., Haas, R.
<2>A stable shuttle vector system for efficient genetic complementation of Helicobacter pylori strains by transformation and conjugation.
<3>Mol. Gen. Genet.
<4>257
<5>519-528
<6>1998
<7>A versatile plasmid shuttle vector system was constructed, which is useful for genetic
complementation of Helicobacter pylori strains or mutants with cloned genes of homologous or
heterologous origin.  The individual plasmid vectors consist of the minimal essential genetic
elements, including an origin of replication for Escherichia coli, a H. pylori-specific
replicon originally identified on a small cryptic H. pylori plasmid, an oriT sequence and a
multiple cloning site.  Shuttle plasmid pHel2 carries a chloramphenicol resistance cassette
(catGC) and pHel3 contains a kanamycin resistance gene (aphA-3) as the selectable marker; both
are functional in E. coli and H. pylori.  The shuttle plasmids were introduced into the H.
pylori strain P1 by natural transformation.  An efficiency of 7.0 x 10^-7 and 4.7 x 10^-7
transformants per viable recipient was achieved with pHel2 and pHel3, respectively, and both
vectors showed stable, autonomous replication of H. pylori.  An approximately 100-fold higher
H. pylori transformation rate was obtained when the shuttle vectors for transformation were
isolated from the homologous H. pylori strain, rather than E. coli, indicating that DNA
restriction and modification mechanisms play a crucial role in plasmid transformation.
Interestingly, both shuttle vectors could also be mobilized efficiently from E. coli into
different H. pylori recipients, with pHel2 showing an efficiency of 2.0 x 10^-5
transconjugants per viable H. pylori P1 recipient.  Thus, DNA restriction seems to be strongly
reduced or absent during conjugal transfer.  The functional complementation of a
recA-deficient H. pylori mutant by the cloned H. pylori recA+ gene, and the expression of the
heterologous green fluorescent protein in H. pylori demonstrate the general usefulness of this
system, which will significantly facilitate the molecular analysis of H. pylori virulence
factors in the future.

<>

<1>Heumann, W.
<2>Rhizobium lupini genetics.
<3>Curr. Top. Microbiol. Immunol.
<4>88
<5>1-24
<6>1979
<7>The purpose of this article is to review a genetic system that differs in many
aspects to the well-known Escherichia coli system.  It is not the aim of the
author to deal directly with the complex issues of Rhizobium symbiosis with
Legumes nor with the problem of symbiotic nitrogen fixation by the Rhizobia.
However, genetical investigations of the rhizobial strains involved may
contribute to some understanding of the genetics of this complex system of
symbiotic nitrogen fixation.  The mutants used in this study were obtained by
mutagenesis of Rhizobium lupini strains isolated directly from Lupinus luteus
root nodules in 1960.  These strains lost certain characteristics of the
wild-type Rhizobium, especially its capacity to nodulate Lupins.  They are,
however, characterized by high conjugational fertility, a prerequisite for the
investigation of their genetics, by star formation, a recognition mechanism;
and also by their bright yellow colony pigmentation.  This pigmentation arises
from carotenoids localized in the plasma membrane of the cells which protect
the cell's cytochromes from photoinactivation.  These three characteristics,
conjugation, star formation, and carotenoid pigmentation, proved very useful
for genetic investigation of these Rhizobium mutants.

<>

<1>Heurgue-Hamard, V., Champ, S., Engstrom, A., Ehrenberg, M., Buckingham, R.H.
<2>The hemK gene in Escherichia coli encodes the N5-glutamine methyltransferase that modifies peptide release factors.
<3>EMBO J.
<4>21
<5>769-778
<6>2002
<7>Class 1 peptide release factors (RFs) in Escherichia coli are N5-methylated on the glutamine
residue of the universally conserved GGQ motif. One other protein alone has been shown to
contain N5-methylglutamine: E.coli ribosomal protein L3. We identify the L3 methyltransferase
as YfcB and show that it methylates ribosomes from a yfcB strain in vitro, but not RF1 or RF2.
HemK, a close orthologue of YfcB, is shown to methylate RF1 and RF2 in vitro. hemK is
immediately downstream of and co-expressed with prfA. Its deletion in E.coli K12 leads to very
poor growth on rich media and abolishes methylation of RF1. The activity of unmethylated RF2
from K12 strains is extremely low due to the cumulative effects of threonine at position 246,
in place of alanine or serine present in all other bacterial RFs, and the lack of
N5-methylation of Gln252. Fast-growing spontaneous revertants in hemK K12 strains contain the
mutations Thr246Ala or Thr246Ser in RF2. HemK and YfcB are the first identified
methyltransferases modifying glutamine, and are widely distributed in nature.

<>

<1>Heymann, S., Gutter, H., Schulke, U., Kirstein, D., Muller-Uri, F., Hess, W., Borner, T.
<2>Procedure for the isolation of restriction endonucleases.
<3>DDR Patent Office
<4>DD 281412 A
<5>
<6>1990
<7>The finding concerns a procedure for the isolation of restriction endonucleases. A
pre-purified restriction endonuclease containing cell extract was applied one or more times to
a chromatography column, which contained stable porous glassbeads with phosphate groups bound
to it as solid phase sorbents, which was equilibrated with binding buffer. After washing with
binding buffer, the bound restriction endonuclease was eluted with a Phosphate-buffer gradient
or eluted in steps of increasing phosphate buffer concentration. The solid-phase sorbent is
reusable after regeneration. Areas of application are modern genetics and the chemical
industry for the production of Bio, Laboratory, and fine chemicals.

<>

<1>Hickman, A.B., Li, Y., Mathew, S.V., May, E.W., Craig, N.L., Dyda, F.
<2>Unexpected structural diversity in DNA recombination: the restriction endonuclease connection.
<3>Mol. Cell
<4>5
<5>1025-1034
<6>2000
<7>Transposition requires a coordinated series of DNA breakage and joining reactions. The Tn7
transposase contains two proteins: TnsA, which carries out DNA breakage at the 5' ends of the
transposon, and TnsB, which carries out breakage and joining at the 3' ends of the
transposon. TnsB is a member of the retroviral integrase superfamily whose hallmark is a
conserved DDE motif. We report here the structure of TnsA at 2.4 A resolution. Surprisingly,
the TnsA fold is that of a type II restriction endonuclease. Thus, Tn7 transposition involves
a collaboration between polypeptides, one containing a DDE motif and one that does not. This
result indicates that the range of biological processes that utilize restriction enzyme-like
folds also includes DNA transposition.

<>

<1>Hicks, M.R., Rodger, A., Thomas, C.M., Batt, S.M., Dafforn, T.R.
<2>Restriction enzyme kinetics monitored by UV linear dichroism.
<3>Biochemistry
<4>45
<5>8912-8917
<6>2006
<7>The use of linear dichroism (LD) spectroscopy for biological applications has been brought to
the forefront recently by our
development of thermostated microvolume Couette cells. We present a
method for following the digestion of DNA by restriction endonucleases
in real time without the use of any extrinsic dyes or labels. This is
accomplished using linear dichroism spectroscopy (the differential
absorbance of light polarized parallel and perpendicular to the sample
orientation axis). The differential absorbance signal depends on the
degree of alignment of the molecules. In this case the DNA is aligned
by Couette flow (flowing the solution in the annular gap between two
concentric cylinders), and we monitor the increase in alignment upon
linearization of a circular DNA molecule. In addition, we observe a
decrease in alignment upon further digestion and subsequent shortening
of the DNA. Ten enzymes were investigated: seven enzymes with a single
cut site (EcoRI, KpnI, NdeI, NotI, NruI, SmaI, XbaI), two enzymes with
two cut sites (BstZ17I, EagI), and one enzyme with no cut site (ClaI).
LD, as implemented in this new assay, is broadly applicable across a
wide range of DNA-modifying enzymes and compounds and, as such, is a
useful addition to the toolbox of biological characterization.

<>

<1>Hien, L.-T., Zatsepin, T.S., Schierling, B., Volkov, E.M., Wende, W., Pingoud, A., Kubareva, E.A., Oretskaya, T.S.
<2>Restriction Endonuclease SsoII with Photoregulated Activity: A 'Molecular Gate' Approach.
<3>Bioconjugate Chem.
<4>22
<5>1366-1373
<6>2011
<7>A novel method for regulating the activity of homodimeric
proteins-"molecular gate" approach-was proposed and its usefulness illustrated for the type II
restriction endonuclease SsoII (R.SsoII) as a model. The "molecular gate" approach is based on
the modification of R.SsoII with azobenzene derivatives, which allows regulating DNA binding
and cleavage via illumination with light. R. SsoII variants with single cysteine residues
introduced at selected positions were obtained and modified with maleimidoazobenzene
derivatives. A twofold change in the enzymatic activity after illumination with light of
wavelengths of 365 and 470 nm, respectively,
was demonstrated when one or two molecules of azobenzene derivatives were attached to the
R.SsoII at the entrance of or within the DNA-binding site.

<>

<1>Hiessl, S., Schuldes, J., Thurmer, A., Halbsguth, T., Broker, D., Angelov, A., Liebl, W., Daniel, R., Steinbuchel, A.
<2>Involvement of Two Latex-Clearing Proteins during Rubber Degradation and Insights into the Subsequent Degradation Pathway Revealed by the Genome Sequence of Gordonia polyisoprenivorans Strain VH2.
<3>Appl. Environ. Microbiol.
<4>78
<5>2874-2887
<6>2012
<7>The increasing production of synthetic and natural poly(cis-1,4-isoprene) rubber
leads to huge challenges in waste management. Only a few bacteria are known to
degrade rubber, and little is known about the mechanism of microbial rubber
degradation. The genome of Gordonia polyisoprenivorans strain VH2, which is one
of the most effective rubber-degrading bacteria, was sequenced and annotated to
elucidate the degradation pathway and other features of this actinomycete. The
genome consists of a circular chromosome of 5,669,805 bp and a circular plasmid
of 174,494 bp with average GC contents of 67.0% and 65.7%, respectively. It
contains 5,110 putative protein-coding sequences, including many candidate genes
responsible for rubber degradation and other biotechnically relevant pathways.
Furthermore, we detected two homologues of a latex-clearing protein, which is
supposed to be a key enzyme in rubber degradation. The deletion of these two
genes for the first time revealed clear evidence that latex-clearing protein is
essential for the microbial utilization of rubber. Based on the genome sequence,
we predict a pathway for the microbial degradation of rubber which is supported
by previous and current data on transposon mutagenesis, deletion mutants, applied
comparative genomics, and literature search.

<>

<1>Hiett, K., Meinersmann, R.
<2>Isolation of a putative retriction/modification system from Campylobacter jejuni.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>224
<6>1994
<7>Molecular studies of the Campylobacter jejuni genome can often prove difficult; one reason is
that C. jejuni genomic DNA can be digested with only a small number of restriction
endonucleases. Our goal was to isolate, characterize, and eventually disrupt a
restriction/modification system in C. jejuni so that future genetic manipulation might prove
easier.

A lambda-zapII library was constructed using C. jejuni A74/O genomic DNA. The library was
then transfected simultaneously with XL-1 blue and XL-1 Blue/pAN4, a plasmid containing the
EcoRI loci in pBR322. Primary plaques were counted and compared from both transfection
conditions; the number of plaques on XL-1 Blue were significantly higher than those on XL-1
Blue/pAN4. Forty-two primary plaques were then picked from the XL-1 Blue/pAN4 lysates and
again transfected using both XL-1 Blue and XL-1Blue/pAN4 lysates and again transfected using
both XL-1 Blue and XL-1 Blue/pAN4. Primary plaques that gave similar lysate numbers when
transfected into both E. coli were chosen on the hypothesis that they carried a C. jejuni
restriction/modification system. Six of these secondary plaques were transfected into XL-1
Blue for phage DNA isolation; isolated DNA was digested using a series of six restriction
endonucleases. Only one of the six enzymes chosen allowed digestion of the phage DNA's.
Future studies will include further characterization of the C. jejuni inserts to determine if
a restriction/modification system is present.


<>

<1>Higashide, M., Kuroda, M., Omura, C.T., Kumano, M., Ohkawa, S., Ichimura, S., Ohta, T.
<2>Methicillin-resistant Staphylococcus saprophyticus Carrying Staphylococcal Cassette Chromosome mec (SCCmec) Have Emerged in Urogenital Tract  Infections.
<3>Antimicrob. Agents Chemother.
<4>52
<5>2061-2068
<6>2008
<7>Staphylococcus saprophyticus is a uropathogenic bacterium that causes acute uncomplicated
urinary tract infections (UTIs), particularly in
female outpatients. We investigated the dissemination and antimicrobial
susceptibility of 101 S. saprophyticus isolates from the genitourinary
tracts of patients in Japan. Eight of these isolates were mecA-positive
and showed beta-lactam resistance. Pulsed field gel electrophoresis (PFGE)
showed that only some isolates were isogenic, indicating that the mecA
gene was apparently acquired independently by mecA-positive isolates
through staphylococcal cassette chromosome mec (SCCmec). Type
determination of SCCmec by multiplex PCR showed non-typeable element in
the 8 mecA-positive isolates. Sequence analysis of the entire SCCmec
element from a prototype S. saprophyticus strain revealed that it is
non-typeable with the current SCCmec classification due to the novel
composition of the class A mec gene complex (IS431-mecA-mecR1-mecI genes)
and the ccrA1/ccrB3 gene complex. Intriguingly, the attachment sites of
SCCmec are similar to those of type I SCCmec in S. aureus NCTC 10442.
Further, the genes around the mec gene complex are similar to those of
type II/III SCCmec in S. aureus, while those around the ccr gene complex
are similar to those of SCC15305RM found in S. saprophyticus ATCC 15305.
In comparison with known SCCmec elements, this S. saprophyticus SCCmec is
a novel type.

<>

<1>Higgins, D.L., Sanozky-Dawes, R., Klaenhammer, T.R.
<2>Characterization of a cointegrate conjugal plasmid encoding phage restriction and modification activities in lactic streptococci.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>87
<5>154
<6>1987
<7>The conjugal plasmid pTN1060 (Tra+) encodes lactose-fermenting ability (Lac+),
resistance to nisin (Nisr), and restriction and modification activities (R/M+)
against phages of lactic streptococci.  Physical characterization of pTN1060
revealed the plasmid was a cointegrate formed from pTR1040 (Lac+, Nisr) and a
30 kb fragment that conferred Tra+ and R/M+ functions.  To identify the origin
of the 30kb fragment, exconjugants from S. lactis N1 were characterized for
Tra+ and R/M+ activities.  Exconjugants harboring either 60 Md pTN1060-like
plasmids, or pTR1040 in combination with a 30 kb plasmid (pTN20) exhibited
Lac+, Nisr, R/M+ and Tra+.  Phenotypic analysis, curing studies, and genetic
transfer experiments with derivatives harboring pTN20 demonstrated a direct
correlation of pTN20 with R/M+ activities and conjugal transfer ability.  pTN20
probes hybridized with pTN1060 but not with pTR1040.  Restriction mapping
further confirmed that pTN20 fragments were present in pTN1060.  The data
demonstrated that pTN1060 was a cointegrate plasmid comprised of pTR1040 and
pTN20.

<>

<1>Higgins, D.L., Sanozky-Dawes, R.B., Klaenhammer, T.R.
<2>Restriction and modification activities from Streptococcus lactis ME2 are encoded by a self-transmissible plasmid, pTN20, that forms cointegrates during mobilization of lactose-fermenting ability.
<3>J. Bacteriol.
<4>170
<5>3435-3442
<6>1988
<7>A self-transmissible (Tra+) plasmid encoding determinants for restriction and
modification activities (R+/M+) from Streptococcus lactis ME2 was isolated and
characterized.  The 28-kilobase (kb) plasmid (pTN20) was detected in
lactose-fermenting (Lac+) transconjugants generated from matings between S.
lactis N1, an ME2 variant, and a plasmid-free recipient, S. lactis LM2301.  The
plaquing efficiencies of prolate- and small isometric-headed phages were
reduced on transconjugants containing either pTN20 (R+/M+ Tra+) or 100-kb
plasmids encoding Lac+, R+/M+, and Tra+.  Lac+ transconjugants which harbored
pTR1040 (Lac+) and pTN20 (R+/M+) were phenotypically R-/M- and transferred Lac+
at low frequency in subsequent matings to give rise to 100-kb R+/M+ plasmids.
R+/M+ activities and high-frequency conjugal transfer ability were detected in
Lac+ transconjugants that contained pTR1041 (Lac+) and pTN20 (R+/M+).  No
100-kb R+/M+ plasmids were recovered after these matings, suggesting that
pTR1041 was mobilized by pTN20 through a process that resembled plasmid
donation.  pTR1041 was identical to pTR1040 but contained an additional 3.3-kb
DNA fragment.  These data suggested that phenotypic expression of R+/M+ and
Tra+ is affected by coresident Lac+ plasmids.  Restriction enzyme analysis and
hybridization reactions demonstrated that the 100-kb R+/M+ plasmid was formed
by a cointegration event between pTR1040 (Lac+) and pTN20 (R+/M+Tra+) during
conjugal transfer via a conductive-type process.  This is the first report that
defines self-transmissible restriction and modification plasmids in the lactic
streptococci.

<>

<1>Higgins, L.S., Besnier, C., Kong, H.
<2>The nicking endonuclease N.BstNBI is closely related to Type IIs restriction endonucleases MlyI and PleI.
<3>Nucleic Acids Res.
<4>29
<5>2492-2501
<6>2001
<7>N.BstNBI is a nicking endonuclease that recognizes the sequence GAGTC and nicks the top strand
preferentially. The Type IIs restriction endonucleases PleI and MlyI also recognize GAGTC, but
cleave both DNA strands. Cloning and sequencing the genes encoding each of these three
endonucleases discloses significant sequence similarities. Mutagenesis studies reveal a
conserved set of catalytic residues among the three endonucleases, suggesting that they are
closely related to each other. Furthermore, PleI and MlyI contain a single active site for DNA
cleavage. The results from cleavage assays show that the reactions catalyzed by PleI and MlyI
are sequential two step processes. The double-stranded DNA is first nicked on one DNA strand
and then further cleaved on the second strand to form linear DNA. Gel filtration analysis
shows that MlyI dimerizes in the presence of a cognate DNA and Ca(2+) whereas N.BstNBI remains
a monomer, implicating dimerization as a requisite for the second strand cleavage. We suggest
that N.BstNBI, MlyI and PleI diverged from a common ancestor and propose that N.BstNBI differs
from MlyI and PleI in having an extremely limited second strand cleavage activity, resulting
in a site-specific nicking endonuclease.

<>

<1>Highlander, S.K. et al.
<2>Subtle genetic changes enhance virulence of methicillin resistant and sensitive Staphylococcus aureus.
<3>BMC Microbiol.
<4>7
<5>99
<6>2007
<7>ABSTRACT: BACKGROUND: Community acquired (CA) methicillin-resistant Staphylococcus aureus
(MRSA) increasingly causes disease worldwide. USA300
has emerged as the predominant clone causing superficial and invasive
infections in children and adults in the USA. Epidemiological studies
suggest that USA300 is more virulent than other CA-MRSA. The genetic
determinants that render virulence and dominance to USA300 remain unclear.
RESULTS: We sequenced the genomes of two pediatric USA300 isolates: one
CA-MRSA and one CA-methicillin susceptible (MSSA), isolated at Texas
Children's Hospital in Houston. DNA sequencing was performed by Sanger
dideoxy whole genome shotgun (WGS) and 454 Life Sciences pyrosequencing
strategies. The sequence of the USA300 MRSA strain was rigorously
annotated. In USA300, MRSA 2685 chromosomal open reading frames were
predicted and 3.1 and 27 kilobase (kb) plasmids were identified. USA300
MSSA contained a 20 kb plasmid with some homology to the 27 kb plasmid.
Two regions found in US300 MRSA were absent in USA300 MSSA. The USA300
sequence was aligned with other sequenced S. aureus genomes and regions
unique to USA300 MRSA were identified. CONCLUSIONS: USA300-MRSA is highly
similar to other MRSA strains based on whole genome alignments and gene
content, indicating that the differences in pathogenesis are due to subtle
changes rather than to large-scale acquisition of virulence factor genes.
The USA300 Houston isolate differs from another sequenced USA300 strain
isolate, derived from a patient in San Francisco, in plasmid content and a
number of sequence polymorphisms. Such differences will provide new
insights into the evolution of pathogens.

<>

<1>Highlander, S.K., Garza, O.
<2>The restriction-modification system of Pasteurella haemolytica is a member of a new family of type I enzymes.
<3>Gene
<4>178
<5>89-96
<6>1996
<7>Genes encoding the type I restriction-modification (R-M) system of the bovine pathogen,
Pasteurella haemolytica, have been identified immediately downstream of a locus that encodes a
transcriptional activator of P. haemolytica leukotoxin expression.  Type I enzymes are encoded
by three genes called hsdM, hsdS and hsdR, and have fallen into three groups, called Ia, Ib
and Ic.  HsdS provides a sequence recognition function which in concert with HsdM forms an
active methyltransferase (Mtase).  Inclusion of the HsdR subunit in the complex creates an
active restriction endonuclease (Enase) capable of cleaving unmethylated target DNA.  The P.
haemolytica hsdMSR genes were mapped using transposon Tn10d-Cam insertions, and bacteriophage
restriction and modification assays in Escherichia coli.  We determined the nucleotide
sequences of hsdM, hsdS and hsdR, and observed that the deduced amino acid (aa) sequences were
very similar to predicted R-M subunits in the respiratory pathogen, Haemophilus influenzae.
Phylogenetic comparisons of all known Hsd aa sequences placed the P. haemolytica and H.
influenzae proteins into a new group which we labeled the Type Id R-M family.  Expression of
the P. haemolytica R-M genes in E. coli was inefficient and is likely to be a consequence of
the unusual codon usage in P. haemolytica genes.

<>

<1>Highlander, S.K., Hang, V.T.
<2>A putative leucine zipper activator of Pasteurella haemolytica leukotoxin transcription and the potential for modulation of its synthesis by slipped-strand  mispairing.
<3>Infect. Immun.
<4>65
<5>3970-3975
<6>1997
<7>A Pasteurella haemolytica cosmid clone that activates leukotoxin transcription in Escherichia
coli has been isolated. The activator locus, alxA, is part of a
continuous open reading frame that includes the type I hsdM methylase gene. AlxA
and HsdM peptides are processed from a precursor, and translation of the
polyprotein can be modulated by slipped-strand mispairing across a
pentanucleotide repeat, ACAGC, within the 5' end of alxA-hsdM. Extracts
containing AlxA can bind to a leukotoxin promoter fragment.

<>

<1>Hilbert, H., Himmelreich, R., Plagens, H., Herrmann, R.
<2>Sequence analysis of 56 kb from the genome of the bacterium Mycoplasma pneumoniae comprising the dnaA region, the atp operon and a cluster of ribosomal protein genes.
<3>Nucleic Acids Res.
<4>24
<5>628-639
<6>1996
<7>To sequence the entire 800 kilobase pair genome of the bacterium Mycoplasma pneumoniae, a
plasmid library was established which contained the majority of the EcoRI fragments from M.
pneumoniae.  The EcoRI fragments were subcloned from an ordered cosmid library comprising the
complete M. pneumoniae genome.  Individual plasmid clones were sequenced in an ordered fashion
mainly by primer walking.  We report here the initial results from the sequence analysis of
about 56 kb comprising the dnaA region as a potential origin of replication, the ATPase operon
and a region coding for a cluster of ribosomal protein genes.  The data were compared with the
corresponding genes/operons from Bacillus subtilis, Escherichia coli, Mycoplasma capricolum
and Mycoplasma gallisepticum.

<>

<1>Hill, A.E., Lainson, F.A.
<2>Survey of restriction-modification systems and transformation in Mannheimia haemolytica and Pasteurella trehalosi.
<3>Vet. Microbiol.
<4>92
<5>103-109
<6>2003
<7>A significant obstacle to molecular studies of Mannheimia (Pasteurella) haemolytica, has been
its resistance to genetic transformation. The lack
of competence of many M. haemolytica strains has been attributed to the
presence of restriction modification systems. In this study,
representative strains of 12 M. haemolytica serotypes and four Pasteurella
trehalosi serotypes were successfully transformed by electroporation using
a recombinant vector derived from the native M. haemolytica A1 serotype
plasmid pNSF2176. Transformation was achieved despite PCR-based evidence
for the presence of genes encoding a type I restriction enzyme, phaI, and
a type II restriction enzyme hsdM, in each of the M. haemolytica strains.

<>

<1>Hill, C.
<2>Bacteriophage and bacteriophage resistance in lactic acid bacteria.
<3>FEMS Microbiol. Rev.
<4>12
<5>87-108
<6>1993
<7>The study of bacteriophage-host interactions has been instrumental in the development of
genetic systems in many genera, and laid many of the foundations of modern molecular genetics.
Research into bacteriophage and bacteriophage resistance in the lactic acid bacteria has moved
into a new and exciting dimension in recent years. Mechanisms such as adsorption inhibition,
restriction and modification, and abortive infection which have been detected and described
phenotypically over the past decade are now being subjected to molecular analysis, and this
has led to a better understanding of the nature and variety of resistance systems employed by
lactic acid bacteria to combat phage attack. In addition, analysis of different bacteriophage
has increased our knowledge of these ubiquitous particles to the point where it is possible to
construct novel phage resistances based on the phage genome itself. This review outlines the
recent progress in the molecular analysis of bacteriophage, bacteriophage resistance and
counter resistance, and the construction of novel resistance mechansims.

<>

<1>Hill, C., Garvey, P., Fitzgerald, G.F.
<2>Bacteriophage-host interactions and resistance mechanisms, analysis of the conjugative bacteriophage resistance plasmid pNP40.
<3>Lait
<4>76
<5>67-79
<6>1996
<7>Research into bacteriophage and bacteriophage resistance in the lactic acid bacteria
has moved into a new and exciting dimension in recent years.  Mechanisms such as adsorption
inhibition, restriction and modification, and abortive infection which have been described
phenotypically over the past decade are now being subjected to molecular analysis, and this
has led
to a better understanding of the nature and variety of resistance systems employed by lactic
acid
bacteria to combat phage attack.  In addition, analysis of different bacteriophage has
increased our
knowlege of these ubiquitous particles to the point where it is possible to construct novel
phage
resistances based on the phage genome.  This review will briefly outline the recent progress
in the
molecular analysis of bacteriophage-host interactions, bacteriophage resistance and counter
resistance, and the construction of novel resistance mechanisms.  In particular, recent
evidence
regarding the mechanisms of resistance employed by the conjugative plasmid pNP40 will be
described in some detail.  In addition, an instance will be described in which pNP40 has been
used
in the construction of phage resistant starters which have been successfully exploited by the
dairy
industry.

<>

<1>Hill, C., Miller, L., Klaenhammer, T.
<2>Molecular characterization of a Type II methylase gene exchanged between pTR2030 and a virulent phage in Lactococci.
<3>Plasmid
<4>23
<5>167-168
<6>1990
<7>None

<>

<1>Hill, C., Miller, L.A., Klaenhammer, T.R.
<2>Nucleotide sequence and distribution of the pTR2030 resistance determinant (hsp) which aborts bacteriophage infection in Lactococci.
<3>Appl. Environ. Microbiol.
<4>56
<5>2255-2258
<6>1990
<7>The lactococcal plasmid pTR2030 encodes resistance to bacteriophage attack via
two mechanisms, an abortive-infection mechanism, designated Hsp, and a
restriction and modification system.  We present the complete sequence of the
hsp structural gene.  The gene is 1,887 base pairs in length and encodes a
protein with a predicted molecular mass of 73.8 kilodaltons.  The upstream
region was cloned in a promoter-screening vector and shown to direct the
constitutive expression of the cat-86 gene.  An internal probe was used to
determine the distribution of the hsp sequence in industrially significant
lactococcal strains and to evaluate its relatedness to another lactococcal
plasmid implicated in an abortive-infection-type mechanism, pNP40.  No homology
was detected, suggesting that this gene is not widely distributed in
lactococci.  Therefore, there are at least two independent abortive-infection
genotypes in lactococci.

<>

<1>Hill, C., Miller, L.A., Klaenhammer, T.R.
<2>Cloning, expression, and sequence determination of a bacteriophage fragment encoding bacteriophage resistance in Lactococcus lactis.
<3>J. Bacteriol.
<4>172
<5>6419-6426
<6>1990
<7>A number of host-encoded phage resistance mechanisms have been described in
lactococci.  However, the phage genome has not been exploited as a source of
additional resistance determinants.  A 4.5-kb BamHI-HindIII fragment of phage
nck202.50 (Phi50) was subcloned in streptococcus-Escherichia coli shuttle
plasmid pSA3 and introduced into Lactoccus lactis NCK203 and MG1363 by
protoplast transformation.  This cloned phage fragment directed a bacteriophage
resistance phenotype designated Per (phage-encoded resistance).  Both Phi50 and
a distantly related phage, nck202.48 (Phi48), formed small plaques on strain
NCK213 at a slightly reduced efficiency of plaquing on the Per+ host.  The per
locus was further reduced to a 1.4-kb fragment through in vitro deletion
analysis.  The 1.4-kb fragment was sequenced, and the Per phenotype was found
to be associated with a ca. 500-bp region rich in direct and inverted repeats.
We present evidence that the Per region contains a phage origin of replication
which, in trans, may interfere with phage replication by titration of DNA
polymerase or other essential replication factors.  It was demonstrated that
the Per+ activity was not detected against six independent phages which were
previously shown to be sensitive to the Hsp+ mechanism.  The mutually exclusive
resistance mechanisms could be combined to confer resistance to both types of
phages (Hsp resistant and Per resistant) in a single host.  This is the first
description in lactococci of a phage resistance phenotype, other than
superinfection immunity, originating from a lactococcal phage genome.

<>

<1>Hill, C., Miller, L.A., Klaenhammer, T.R.
<2>In vivo genetic exchange of a functional domain from a Type II A methylase between lactococcal plasmid pTR2030 and a virulent bacteriophage.
<3>J. Bacteriol.
<4>173
<5>4363-4370
<6>1991
<7>The conjugative plasmid pTR2030 confers bacteriophage resistance to lactococci by two
independent mechanisms, an abortive infection mechanism (Hsp+) and a restriction and
modification system (R+/M+).  pTR2030 transconjugants of lactococcal strains are used in the
dairy industry to prolong the usefulness of mesophilic starter cultures.  One bacteriophage
which has emerged against a pTR2030 transconjugant is not susceptible to either of the two
defense systems encoded by the plasmid.  Phage nck202.50 (Phi50) is completely resistant to
restriction by pTR2030.  A region of homology between pTR2030 Phi50 was subcloned, physically
mapped, and sequenced.  A region of 1,273 bp was identical in both plasmid and phage,
suggesting that the fragment had recently been transferred between the two genomes.  Sequence
analysis confirmed that the transferred region encoded >55% of the amino domain of the
structural gene for a type II methylase designated LlaI.  The LlaI gene is 1,869 bp in length
and shows organizational similarities to the type II A methylase FokI.  In addition to the
amino domain, upstream sequences, possibly containing the expression signals, were present on
the phage genome.  The phage Phi50 fragment containing the methylase amino domain, designated
LlaPI, when cloned onto the shuttle vector pSA3 was capable of modifying another phage genome
in trans.  This is the first report of the genetic exchange between a bacterium and a phage
which confers a selective advantage on the phage.  Definition of the LlaI system on pTR2030
provides the first evidence that type II systems contribute to restriction and modification
phenotypes during host-dependent replication of phages in lactococci.

<>

<1>Hill, C., Miller, L.A., Klaenhammer, T.R.
<2>The bacteriophage resistance plasmid pTR2030 forms high-molecular-weight multimers in Lactococci.
<3>Plasmid
<4>25
<5>105-112
<6>1991
<7>Lactococcus lactis ME2 can transfer a 46-kb plasmid, pTR2030, which encodes
abortive phage infection (Hsp) and restriction/modification (R/M) activities.
pTR2030 can be detected as a monomeric plasmid in transconjugants at low copy
number, but not in ME2.  pTR2030-specific probes were cloned and used to
determine the location of the element in ME2.  No homology was observed between
these pTR2030-specific probes and the CsCl-purified plasmid content of ME2.
However, probes specific for pTR2030 hybridized strongly to a
high-molecular-weight moiety, and not to chromosomal DNA, in total DNA isolated
by a gentle lysis procedure.  The absence of junction fragments indicates that
pTR2030 forms high-molecular-weight multimers in lactococci.  A phage-sensitive
derivative of ME1, L. lactis N1, is cured of pTR2030 and no longer possesses
the high-molecular-weight species.  When pTR2030 was reintroduced to N1 via
conjugation, an ME2-like phage-insensitive phenotype was restored.  pTR2030
could remain as a detectable monomeric plasmid in the N1 transconjugants or
could revert to the high-molecular-weight structure.

<>

<1>Hill, C., Pierce, K., Klaenhammer, T.R.
<2>The conjugative plasmid pTR2030 encodes two bacteriophage defense mechanisms in Lactococci, restriction modification (R+/M+) and abortive infection (Hsp+).
<3>Appl. Environ. Microbiol.
<4>55
<5>2416-2419
<6>1989
<7>pTR2030 is a conjugative plasmid which encodes resistance to bacteriophage in
lactococci by a mechanism that aborts the phage infection (Hsp+).  Subcloning
and in vivo deletion events showed that two independent mechanisms of
resistance are located on a 13.6-kilobase BglII fragment cloned in pSA3; one
mechanism is responsible for the abortive infection, and the other encodes a
restriction modification system.  The introduction of pTR2030 or the
recombinant plasmid pTK6 resulted in the loss of a resident restriction
modification plasmid in Lactococcus lactis NCK202 which was not previously
identified.

<>

<1>Hill, C., Romero, D.A., McKenney, D.S., Finer, K.R., Klaenhammer, T.R.
<2>Localization, cloning, and expression of genetic determinants for bacteriophage resistance (Hsp) from the conjugative plasmid pTR2030.
<3>Appl. Environ. Microbiol.
<4>55
<5>1684-1689
<6>1989
<7>Genetic determinants for a bacteriophage resistance mechanism (Hsp+) encoded by plasmid
pTR2030 (46.2 kilobases [kb]) were localized by mapping an 11.5 -kb deletion that accompanied
the transition of Lactococcus lactis LMA 12-4 transconjugants (M.E. Sanders, P.J. Leonard,
W.D. Sing and T.R. Klaenhammer, Appl. Environ. Microbiol. 52:1001-1007, 1986) from phage
resistance to phage sensitivity. The deleted 34.7-kb replicon (pTR2023, Hsp-) retained its
conjugative ability, demonstrating that the phage resistance and conjugal transfer
determinants were genetically distinct. The Hsp region of pTR2030, which was contained within
a 13.6-kb BglII fragment, was cloned into the BamHI site of bacteriophage lambda EMBL3, and
Hsp was subcloned into the Escherichia coli-Streptococcus shuttle vector pSa3. The recombinant
plasmids pTK6 and pTK9 were recovered in E. coli HB101 and contained a 13.6-kb insert in
opposite orientations. L. lactis MG1363 transformants carrying pTK6 or pTK9 exhibited a
significant reduction in plaque size, in addition to a slight reduction in the efficiency of
plaquing for both prolate and small isometric phages. Phenotypic reactions observed for the
recombinant plasmids suggest that pTR2030-encoded Hsp acts similarly against both prolate and
small isometric phages. Tn5 mutagenesis was used to define the region essential for the
expression of the Hsp+ phenotype. Any of four insertions within a 3-kb region resulted in the
loss of phagge resistance, whereas a further 26 insertions outside this locus had no effect on
Hsp expression. In vitro deletion analysis confirmed that the 3-kb region contained all the
information necessary for the observed resistance.

<>

<1>Hill, S.A.
<2>Cell to cell transmission of donor DNA overcomes differential incorporation of non-homologous and homologous markers in Neisseria gonorrhoeae.
<3>Gene
<4>240
<5>175-182
<6>1999
<7>The neisseriae are naturally competent for DNA transformation. This genetic study examines
whether the modification status of chromosomal donor DNA affects transformation of Neisseria
gonorrhoeae to drug resistance. When a single modification system was inactivated, unmodified
chromosomal donor DNA was not restricted when used to transform the cognate restriction+ host,
irrespective of whether the donor DNA carried a point mutation (homologous marker) or a
drug-resistance gene cassette (non-homologous marker). These observations contrasted
transformations performed with unmodified plasmid donor DNAs, where the incoming DNA was
excluded. However, during the study, it became apparent that certain strains of gonococci
showed differential incorporation of non-homologous markers when compared with the
incorporation of the homologous marker, even when the donor DNAs were prepared from parental
strains. Differential incorporation of markers could be rescued either through cell to cell
transmission of donor DNA, or by performing in vitro transformations with donor DNA
preparations that were obtained from spent culture supernatants. Overall, the data indicate
that, in addition to the exclusion of foreign DNA through the requirement for a genus-specific
uptake sequence, gonococci appear capable of excluding DNA on the basis of homology.

<>

<1>Hiller, D.A.
<2>Mechanism of DNA bending and its role in the specificity of EcoRV restriction endonuclease.
<3>Ph.D. Thesis, Univ. of California, Santa Barbara
<4>
<5>
<6>2005
<7>Many DNA binding proteins involved in transcription, packaging and other processes bend their
target sequences as part of their function.  The mechanism of DNA bending and its role in
specificity, therefore, are topics of fundamental interest.  The homodimeric EcoRV restriction
endonuclease, which recognizes the six base pair sequence GATATC, has emerged as one of the
best studied nucleic acid binding proteins.  EcoRV bends the center TA step by 50 degrees to
facilitate cleavage of the phosphodiester backbone.  A fluorescence resonance energy transfer
assay was developed to monitor the DNA bending step, in addition to the binding and cleavage
steps previously observed.  Bending of cognate DNA is rapid and appears to be simultaneous
with binding.  Furthermore, no bending was observed in the absence of divalent metal. To
elucidate the mechanism of DNA bending by EcoRV, electrostatic contacts between the protein
and DNA were deleted.  This was done by mutating positively charged residues to alanine,
replacing negatively charged DNA phosphates with uncharged methylphosphonates, and making both
substitutions simultaneously.  It was found that asymmetric neutralization of the phosphate
backbone improves DNA binding, bending, and cleavage, which indicates that DNA bending begins
during the initial association process and has increasing importance throughout the catalytic
pathway.  DNA bending was also stabilized by the diffuse charge present in the
carboxy-terminal domain of each monomer, as observed by fluorescence in solution.  The crystal
structure of a mutant lacking these domains also showed small changes at the active site,
leading to destabilized binding of the divalent metal needed for catalysis.  To further
understand how induced fit contributes to specificity, the transient kinetic techniques
developed were used in partnership with x-ray crystallography to study discrimination against
a noncognate sequence, GAATTC.  Again, binding and bending were simultaneous; however the bend
angle decreased in response to a steric block at the center step.  The crystal structure
showed that metal binding was disrupted, and this was suffient to reduce the cleavage rate
105-fold.  Together, these studies demonstrate that specificity is coupled to catalysis by
induced fit and partially establish the timing and mechanism of DNA bending by a site-specific
nucleic acid enzyme.

<>

<1>Hiller, D.A., Fogg, J.M., Martin, A.M., Beechem, J.M., Reich, N.O., Perona, J.J.
<2>Simultaneous DNA binding and bending by EcoRV endonuclease observed by real-time fluorescence.
<3>Biochemistry
<4>42
<5>14375-14385
<6>2003
<7>The complete catalytic cycle of EcoRV endonuclease has been observed by combining fluorescence
anisotropy with fluorescence resonance energy
transfer (FRET) measurements. Binding, bending, and cleavage of substrate
oligonucleotides were monitored in real time by rhodamine-x anisotropy and
by FRET between rhodamine and fluorescein dyes attached to opposite ends
of a 14-mer DNA duplex. For the cognate GATATC site binding and bending
are found to be nearly simultaneous, with association and bending rate
constants of (1.45-1.6) x 10(8) M(-1) s(-1). On the basis of the
measurement of k(off) by a substrate-trapping approach, the equilibrium
dissociation constant of the enzyme-DNA complex in the presence of
inhibitory calcium ions was calculated as 3.7 x 10(-12) M from the kinetic
constants. Further, the entire DNA cleavage reaction can be observed in
the presence of catalytic Mg(2+) ions. These measurements reveal that the
binding and bending steps occur at equivalent rates in the presence of
either Mg(2+) or Ca(2+), while a slow decrease in fluorescence intensity
following bending corresponds to k(cat), which is limited by the cleavage
and product dissociation steps. Measurement of k(on) and k(off) in the
absence of divalent metals shows that the DNA binding affinity is
decreased by 5000-fold to 1.4 x 10(-8) M, and no bending could be detected
in this case. Together with crystallographic studies, these data suggest a
model for the induced-fit conformational change in which the role of
divalent metal ions is to stabilize the sharply bent DNA in an orientation
suitable for accessing the catalytic transition state.

<>

<1>Hiller, D.A., Perona, J.J.
<2>Positively charged C-terminal subdomains of EcoRV endonuclease: contributions to DNA binding, bending, and cleavage.
<3>Biochemistry
<4>45
<5>11453-11463
<6>2006
<7>The carboxy-terminal subdomains of the homodimeric EcoRV restriction endonuclease each bear a
net charge of +4 and are positioned on the inner
concave surface of the 50 degree DNA bend that is induced by the enzyme. A
complete kinetic and structural analysis of a truncated EcoRV mutant
lacking these domains was performed to assess the importance of this
diffuse charge in facilitating DNA binding, bending, and cleavage. At the
level of formation of an enzyme-DNA complex, the association rate for the
dimeric mutant enzyme was sharply decreased by 10(3)-fold, while the
equilibrium dissociation constant was weakened by nearly 10(6)-fold
compared with that of wild-type EcoRV. Thus, the C-terminal subdomains
strongly stabilize the enzyme-DNA ground-state complex in which the DNA is
known to be bent. Further, the extent of DNA bending as observed by
fluorescence resonance energy transfer was also significantly decreased.
The crystal structure of the truncated enzyme bound to DNA and calcium
ions at 2.4 A resolution reveals that the global fold is preserved and
suggests that a divalent metal ion crucial to catalysis is destabilized in
the active site. This may explain the 100-fold decrease in the rate of
metal-dependent phosphoryl transfer observed for the mutant. These results
show that diffuse positive charge associated with the C-terminal
subdomains of EcoRV plays a key role in DNA association, bending, and
cleavage.

<>

<1>Hiller, D.A., Rodriguez, A.M., Perona, J.J.
<2>Non-cognate enzyme-DNA complex: structural and kinetic analysis of EcoRV endonuclease bound to the EcoRI recognition site GAATTC.
<3>J. Mol. Biol.
<4>354
<5>121-136
<6>2005
<7>The crystal structure of EcoRV endonuclease bound to non-cognate DNA at 2.0 angstroms
resolution shows that very small structural adaptations are sufficient to ensure the extreme
sequence specificity characteristic of restriction enzymes. EcoRV bends its specific GATATC
site sharply by 50 degrees into the major groove at the center TA step, generating unusual
base-base interactions along each individual DNA strand. In the symmetric non-cognate complex
bound to GAATTC, the center step bend is relaxed to avoid steric hindrance caused by the
different placement of the exocyclic thymine methyl groups. The decreased base-pair unstacking
in turn leads to small conformational rearrangements in the sugar-phosphate backbone,
sufficient to destabilize binding of crucial divalent metal ions in the active site. A second
crystal structure of EcoRV bound to the base-analog GAAUTC site shows that the 50 degrees
center-step bend of the DNA is restored. However, while divalent metals bind at high occupancy
in this structure, one metal ion shifts away from binding at the scissile DNA phosphate to a
position near the 3'-adjacent phosphate group. This may explain why the 10(4)-fold attenuated
cleavage efficiency toward GAATTC is reconstituted by less than tenfold toward GAAUTC.
Examination of DNA binding and bending by equilibrium and stopped-flow florescence quenching
and fluorescence resonance energy transfer (FRET) methods demonstrates that the capacity of
EcoRV to bend the GAATTC non-cognate site is severely limited, but that full bending of GAAUTC
is achieved at only a threefold reduced rate compared with the cognate complex. Together, the
structural and biochemical data demonstrate the existence of distinct mechanisms for ensuring
specificity at the bending and catalytic steps, respectively. The limited conformational
rearrangements observed in the EcoRV non-cognate complex provide a sharp contrast to the
extensive structural changes found in a non-cognate BamHI-DNA crystal structure, thus
demonstrating a diversity of mechanisms by which restriction enzymes are able to achieve
specificity.

<>

<1>Hiller, N.L., Janto, B., Hogg, J.S., Boissy, R., Yu, S., Powell, E., Keefe, R., Ehrlich, N.E., Shen, K., Hayes, J., Barbadora, K., Klimke, W., Dernovoy, D., Tatusova, T., Parkhill, J., Bentley, S.D., Post, J.C., Ehrlich, G.D., Hu, F.Z.
<2>Comparative genomic analyses of seventeen Streptococcus pneumoniae strains: insights into the pneumococcal supragenome.
<3>J. Bacteriol.
<4>189
<5>8186-8195
<6>2007
<7>The distributed-genome hypothesis (DGH) states that pathogenic bacteria
possess a supragenome that is much larger than the genome of any single
bacterium and that these pathogens utilize genetic recombination and a
large, noncore set of genes as a means of diversity generation. We
sequenced the genomes of eight nasopharyngeal strains of Streptococcus
pneumoniae isolated from pediatric patients with upper respiratory
symptoms and performed quantitative genomic analyses among these and nine
publicly available pneumococcal strains. Coding sequences from all strains
were grouped into 3,170 orthologous gene clusters, of which 1,454 (46%)
were conserved among all 17 strains. The majority of the gene clusters,
1,716 (54%), were not found in all strains. Genic differences per strain
pair ranged from 35 to 629 orthologous clusters, with each strain's genome
containing between 21 and 32% noncore genes. The distribution of the
orthologous clusters per genome for the 17 strains was entered into the
finite-supragenome model, which predicted that (i) the S. pneumoniae
supragenome contains more than 5,000 orthologous clusters and (ii) 99% of
the orthologous clusters ( approximately 3,000) that are represented in
the S. pneumoniae population at frequencies of >or=0.1 can be identified
if 33 representative genomes are sequenced. These extensive genic
diversity data support the DGH and provide a basis for understanding the
great differences in clinical phenotype associated with various
pneumococcal strains. When these findings are taken together with previous
studies that demonstrated the presence of a supragenome for Streptococcus
agalactiae and Haemophilus influenzae, it appears that the possession of a
distributed genome is a common host interaction strategy.

<>

<1>Hillman, J.L., Lal, P., Corley, N.C., Guegler, K.J., Yue, H.
<2>Human nucleic acid methylases.
<3>US Patent Office
<4>US 6096526
<5>
<6>2000
<7>The invention provides a human nucleic acid methylases (HNAM) and polynucleotides which
identify and encode HNAM. The invention also
provides expression vectors, host cells, antibodies, agonists, and
antagonists. The invention also provides methods for diagnosing,
treating or preventing disorders associated with expression of HNAM.

<>

<1>Himmelreich, R., Hilbert, H., Plagens, H., Pirkl, E., Li, B.-C., Herrmann, R.
<2>Complete sequence analysis of the genome of the bacterium Mycoplasma pneumoniae.
<3>Nucleic Acids Res.
<4>24
<5>4420-4449
<6>1996
<7>The entire genome of the bacterium Mycoplasma pneumoniae M129 has been sequenced.  It has a
size of 816,394 base pairs with an average G+C content of 40.0 mol%.  We predict 677 open
reading frames (ORFs) and 39 genes coding for various RNA species.  Of the predicted ORFs,
75.9% showed significant similarity to genes/proteins of other organisms while only 9.9% did
not reveal any significant similarity to gene sequences in databases.  This permitted us
tentatively to assign a functional classification to a large number of ORFs and to deduce the
biochemical and physiological properties of this bacterium.  The reduction of the genome size
of M.pneumoniae during its reductive evolution from ancestral bacteria can be explained by the
loss of complete anabolic (e.g. no amino acid synthesis) and metabolic pathways.  Therefore,
M.pneumoniae depends in nature on an obligate parasitic lifestyle which requires the provision
of exogenous essential metabolites.  All the major classes of cellular processes and metabolic
pathways are briefly described.  For a number of activities/functions present in M.pneumoniae
according to experimental evidence, the corresponding genes could not be identified by
similarity search.  For instance we failed to identify genes/proteins involved in motility,
chemotaxis and management of oxidative stress.

<>

<1>Himmelreich, R., Plagens, H., Hilbert, H., Reiner, B., Herrmann, R.
<2>Comparative anlaysis of the genomes of the bacteria Mycoplasma pneumoniae and Mycoplasma genitalium.
<3>Nucleic Acids Res.
<4>25
<5>701-712
<6>1997
<7>The sequenced genomes of the two closely related bacteria Mycoplasma genitalium and Mycoplasma
pneumoniae were compared with emphasis on genome organization and coding capacity.  All the
470 proposed open reading frames (ORFs) of the smaller M.genitalium genome (580 kb) were
contained in the larger genome (816 kb) of M.pneumoniae.  There were some discrepancies in
annotation, but inspection of the DNA sequences showed that the corresponding DNA was always
present in M.pneumoniae.  The two genomes could be subdivided into six segments.  The order of
orthologous genes was well conserved within individual segments but the order of these
segments in both bacteria was different.  We explain the different organization of the
segments by translocation via homologous recombination.  The translocations did not disturb
the continuous bidirectional course of transcription in both genomes, starting at the proposed
origin of replication.  The additional 236 kb in M.pneumoniae, compared with the M.genitalium
genome, were coding for 209 proposed ORFs not identified in M.genitalium.  Of these ORFs, 110
were amplifications of ORFs existing mainly as single copies in M.genitalium.  In addition, 23
ORFs containing a copy of either one of the three repetitive DNA sequences RepMP2/3, RepMP4
and RepMP5 were annotated in M.pneumoniae but not in M.genitalium, although similar DNA
sequences were present.  The M.pneumoniae-specific genes included a restriction-modification
system, two transport systems for carbohydrates, the complete set of three genes coding for
the arginine dihydrolase pathway and 14 copies of the repetitive DNA sequence RepMP1 which
were part of several different translated genes with unknown function.

<>

<1>Hines, J.L., Agarwal, K.L.
<2>Purification and characterization of HpaI and HpaII.
<3>Fed. Proc.
<4>38
<5>294
<6>1979
<7>The restriction endonucleases from Haemophilus parainfluenzae, HpaI and HpaII,
have been purified to homogeneity.  HpaI has been purified 10,000 fold and has
a specific activity of 2.2 x 106 units/mg.  (1 unit corresponds to the amount
of enzyme required to cleave 1 microgram of lambda DNA to completion in 60
minutes.) HpaI has a monomer molecular weight of 29,000 daltons and exists as a
58,000 dalton dimer under native conditions.  A specific volume (V) of 0.72
cc/g used in the molecular weight determinations was calculated from the amino
acid composition.  The sedimentation velocity is 3.61 x 10-13 sec.  A ratio of
frictional coefficients (f/fo) was found to be 1.5 indicating an elongated
molecule.  HpaII has been purified 8,000 fold and has a specific activity of
1.8 x 106 units/mg.  (Same units as above.)  HpaII has a monomer molecular
weight of 41,000 and exists as an 82,000 dalton dimer under native conditions.
A specific volume (v) of 0.73 cc/g used in the molecular weight determinations
was calculated from the amino acid composition.  The sedimentation velocity is
4.45 x 10-13 sec.  A ratio of frictional coefficients (f/fo) was found to be
1.45 indicating that HpaII is also an elongated molecule.

<>

<1>Hines, J.L., Chauncey, T.R., Agarwal, K.L.
<2>Preparation and properties of HpaI and HpaII endonucleases.
<3>Methods Enzymol.
<4>65
<5>153-163
<6>1980
<7>Two restriction endonuclease activities, HpaI and HpaII, have been isolated
from Haemophilus parainfluenzae.  Endonuclease HpaI has been isolated in
homogeneous form, while HpaII has been purified free of contaminating nuclease
and phosphatase activies.  HpaI recognizes the DNA sequence and cleaves the
phosphodiester bonds in both strands as indicated.  HpaII recognizes the DNA
sequence and cleaves the phosphodiester bonds in a staggered fashion as
indicated.  The reaction products, in both cases, contain 5'-phosphoryl and
3'-hydroxyl termini.

<>

<1>Hingorani-Varma, K., Bitinaite, J.
<2>Kinetic analysis of the coordinated interaction of SgrAI restriction endonuclease with different DNA targets.
<3>J. Biol. Chem.
<4>278
<5>40392-40399
<6>2003
<7>SgrAI restriction endonuclease cooperatively interacts and cleaves two target sites that
include both the canonical sites, CPuCCGGPyG, and the
secondary sites, CPuCCGGPy(A/T/C). It has been observed that the cleaved
canonical sites stimulate SgrAI cleavage at the secondary sites.
Equilibrium binding studies show that SgrAI binds to its canonical sites
with a high affinity (Ka = 4-8 x 10(10) M-1) and that it has a 15-fold
lower affinity for the cleaved canonical sites and a 30-fold lower
affinity for the secondary sites. Steady-state kinetics reveals substrate
cooperativity for SgrAI cleavage on both canonical and secondary sites.
The specificity of SgrAI for the secondary site CACCGGCT, as measured by
kcat/K is about 500-fold lower than that for the canonical site CACCGGCG,
but this difference is reduced to 10-fold in the presence of the cleaved
canonical sites. The efficiency of canonical site cleavage also increases
by 3-fold when the cleaved canonical sites are present in the reaction.
Furthermore, the substrate cooperativity for SgrAI cleavage is abolished
for both types of sites in the presence of cleaved canonical sites. These
results indicate that target site cleavage occurs via a coordinated
interaction of two SgrAI protein subunits, where the subunit bound to the
cleaved site stimulates the cleavage of the uncut site bound by the other
subunit. The free subunits of SgrAI have the flexibility to bind different
target sites and, consequently, assemble into various catalytically active
complexes, which differ in their catalytic efficiencies.

<>

<1>Hinkle, N.F., Miller, R.V.
<2>pMG7-mediated restriction of Pseudomonas aeruginosa phage DNAs is determined by a class II restriction endonuclease.
<3>Plasmid
<4>2
<5>387-393
<6>1979
<7>A class II restriction endonuclease, PaeR7, has been isolated from a Pseudomonas aeruginosa
strain containing the resistance plasmid pMG7. The activity cannot be isolated from an
isogenic strain which does not contain this resistance plasmid. EndoR-PaeR7 requires Mg2+ for
activity but does not require ATP or S-adenosyl-L-methionine. Specific digestion patterns are
produced upon agarose gel electrophoresis of substrate DNAs which have been digested with the
enzyme. The enzyme is the biochemical basis for the pMG7-mediated phage interference reported
by Jacoby and Sutton (1977). DNAs isolated from restricted phages act as substrates for the
enzyme while DNAs isolated from unrestricted phages and phages which have been modified by
growth on strains containing pMG7 do not act as substrates for the enzyme.

<>

<1>Hinsch, B., Kula, M.-R.
<2>Physical and kinetic properties of the site specific endonuclease BamHI from Bacillus amyloliquefaciens.
<3>Nucleic Acids Res.
<4>8
<5>623-633
<6>1980
<7>The site specific endonuclease BamHI which is composed of subunits of a
molecular weight of 22,000 can aggregate to complexes of a molecular weight of
36,0000.  It is an acidic protein with an iselectric point at pH 5.3.  Optimal
activity is reached at 13 mM MgCl2.  A very simple method is presented to
determine kinetic constants of restriction enzymes directly from agarose gel
photographs without any further equipment applying the integrated Michaelis
Menten equation.  With pJC 80 DNA as a substrate KM was found to be 3.6 10-10
M.  The method can be used to redefine the unit activity of site specific
endonucleases unambiguously.

<>

<1>Hinsch, B., Kula, M.-R.
<2>Reaction kinetics of some important site-specific endonucleases.
<3>Nucleic Acids Res.
<4>9
<5>3159-3174
<6>1981
<7>Reaction kinetics of the site-specific endonucleases BamHI, BglII, ClaI, EcoRI,
HpaII, PstI, SalI, SmaI, and XorII were investigated employing some frequently
used substrates.  Six of these enzymes could be analyzed under steady-state
conditions.  Kinetic data were obtained from progres curves applying an
integrated Michaelis-Menten equation.  KM ranged from 4 - 10-9M to 4 - 10-11 M.
Activities also spanned two orders of magnitude.  In the case of ClaI the
analysis of the pre-steady-state kinetics ("burst reaction") allowed the
assessment of several rate constants.  The rate-limiting step is the very slow
dissociation of the enzyme-product complex (0.22 min-1).  This complex is
formed from the enzyme-bound nicked intermediate at a rate of about 6.  SmaI
and XorII resembled ClaI in their kinetics.  The burst reaction can be used for
the easy and unambiguous determination of molar concentrations of site-specific
endonucleases in any preparation, which is free of non-specific DNases.

<>

<1>Hinsch, B., Mayer, H., Kula, M.-R.
<2>Binding of non-substrate nucleotides to a restriction endonuclease: a model for the interaction of Bam HI with its recognition sequence.
<3>Nucleic Acids Res.
<4>8
<5>2547-2559
<6>1980
<7>The kinetic constants of the site-specific endonuclease Bam HI for various
substrates were determined and binding of non-substrate nucleotides to the
enzyme was studied.  Agarose gel assays in combination with an integrated
Michaelis-Menten equation were used for the evaluation of data.  The turnover
number was 2.2 min-1 at 37C with pJC80 DNA as the substrate.  It depends on the
conformation and base composition of the substrate.  Michaelis constants also
depend on substrate conformation.  Non-substrate polynucleotides were found to
inhibit Bam competitively with KI ranging from 10-6 to >10-3 M depending on
base composition, base pairing, and helix conformation.  Dinucleotides showed
sequence-specific, competitive inhibition with KIs ranging from 10-5 to >10-3
M.  Mononucleotides and -nucleosides acted noncompetitively.  Binding was
influenced by the extent of phosphorylation, but not by the nature of the base.
KIs varied between 10-3 and 10-2 M.  The results are discussed with respect to
the recognition requirements of Bam HI.

<>

<1>Hiom, K., Sedgwick, S.G.
<2>Simple methods for making EcoK and McrA restrictionless mutants.
<3>Nucleic Acids Res.
<4>19
<5>2502
<6>1991
<7>Restriction systems present a major obstacle to the cloning and expression of
heterologous DNAs in bacteria such as E. coli.  Many modern cloning strains
therefore carry disabling mutations in genes encoding restriction endonucleases
to prevent degradation of these heterologous DNAs, however many laboratory
strains of E. coli are not deficient in host restriction activity and are
therefore inaccessible to foreign DNAs.  Here we describe simple methods for
genetically eliminating EcoK and McrA restriction from E. coli that are
generally applicable to strain improvement programs with other bacterial
species.

<>

<1>Hiom, K., Sedgwick, S.G.
<2>Cloning and structural characterization of the mcrA locus of Escherichia coli.
<3>J. Bacteriol.
<4>173
<5>7368-7373
<6>1991
<7>Escherichia coli has DNA restriction systems which are able to recognize and
attack modified cytosine residues in the DNA of incoming bacteriophages and
plasmids.  The locus for the McrA/RglA system of modified cytosine restriction
was located near the pin gene of the defective element, e14.  Hence, loss of
the e14 element through abortive induction after UV irradiation caused a
permanent loss of McrA restriction activity.  e14 DNA encoding McrA restriction
was cloned and sequenced to reveal a single open reading frame of 831 bp with a
predicted gene product of 31 kDa.  Clones expressing the complete open reading
frame conferred both McrA and RglA phenotypes; however, a deletion derivative
was found which complemented RglA restriction against nonglucosylated T6gt
phage but did not complement for McrA restriction of methylated plasmid DNA.
Possible explanations for this activity and a comparison with the different
organization of the McrB/RglB restriction system are discussed.

<>

<1>Hiom, K., Thomas, S.M., Sedgwick, S.G.
<2>Different mechanisms for SOS induced alleviation of DNA restriction in Escherichia coli.
<3>Biochimie
<4>73
<5>399-405
<6>1991
<7>The alleviation of DNA restriction during the SOS response in Escherichia coli has been
further investigated.  With the EcoK DNA restriction system UV irradiated wild-type cells show
a 10^4-fold increase in ability to plate non-modified lambda phage and a 3-4 fold increase in
transformation by non-modified plasmid DNA.  A role for the umuDC genes of E. coli in the
process of SOS-induced restriction alleviation was identified by showing that a umuC122::Tn5
mutant could alleviate EcoK restriction to only 5% that of wild-type levels.  Although umuDC
are better characterized for their pivotal role in SOS induced mutagenesis, it is demonstrated
here that umu-dependent alleviation of EcoK restriction is a transient process in which
umu-dependent mutagenesis plays little part.  A second form of SOS induced alleviation of DNA
restriction is described in this paper involving the McrA restriction system.  The mcrA gene
is shown to be encoded within a defective prophage called e14 situated at the 25 min region on
the Escherichia coli genetic map.  e14 is known to abortively excise from the chromosome after
SOS induction and it is demonstrated in this report that mcrA is lost from the genome after
SOS induction as part of e14.  This results in a co-ordinate decrease in the level of mcrA
restriction within a population of cells.

<>

<1>Hiom, K.J., Sedgwick, S.G.
<2>Alleviation of EcoK DNA restriction in Escherichia coli and involvement of umuDC activity.
<3>Mol. Gen. Genet.
<4>231
<5>265-275
<6>1992
<7>The activity of the EcoKI DNA restriction system of Escherichia coli reduces
both the plating efficiency of unmodified phage lambda and the transforming
ability of unmodified pBR322 plasmid DNA.  However, restriction can be
alleviated in wild-type cells, by UV irradiation and expression of the SOS
response, so that 10/3- to 10/4-fold increases in phage growth and fourfold
increases in plasmid transformation occurred with unmodified DNA.  Restriction
alleviation was found to be a transient effect because induced cells, which
initially failed to restrict unmodified plasmid DNA, later restricted
unmodified phage lambda.  Although the SOS response was needed for restriction
alleviation, constitutive SOS induction, elicited genetically with a recA730
mutation, did not alleviate restriction and UV irradiation was still needed.  A
hitherto unsuspected involvement of the umuDC operon in this alleviation of
restriction is characterized and, by differential complementation, was
separated from the better known role of umuDC in mutagenic DNA repair.  The
need for cleavage of UmuD for restriction alleviation was shown with plasmids
encoding cleavable, cleaved, and non-cleavable forms of UmuD.  However, UV
irradiation was still needed even when cleaved UmuD was provided.  The
possibility that restriction alleviation occurs by a general inhibition of the
EcoK restriction/modification complex was tested and discounted because
modification of lambda was not reduced by UV irradiation.  An alternative idea,
that restriction activity was competitively reduced by an increase in EcoK
modification, was also discounted by the lack of any increase in the
modification of lambda Ral-, a naturally undermodified phage.  Other possible
mechanisms for restriction alleviation are discussed.

<>

<1>Hiramatsu, K., Aritaka, N., Hanaki, H., Kawasaki, S., Hosoda, Y., Hori, S., Fukuchi, Y., Kobayashi, I.
<2>Dissemination in Japanese hospitals of strains of Staphylococcus aureus heterogeneously resistant to vancomycin.
<3>Lancet
<4>350
<5>1670-1673
<6>1997
<7>BACKGROUND: Since the discovery of the vancomycin-resistant Staphylococcus aureus (VRSA)
strain Mu50 (minimum inhibitory concentration [MIC] 8 mg/L),
there has been concern about the potential spread of such strains
throughout Japanese hospitals. Two important questions need to be
answered: (1) what is the prevalence of VRSA, and (2) by what mechanism
does vancomycin resistance occur. METHODS: The vancomycin susceptibilities
of three methicillin-resistant S aureus (MRSA) strains (Mu50, Mu3, and H1)
and the methicillin-susceptible S aureus type strain FDA209P were compared
by MIC determinations and population analysis. Mu3 (MIC 3 mg/L) was
isolated from the sputum of a patient with pneumonia after surgery who had
failed vancomycin therapy. H1 (MIC 2 mg/L), which is a representative
vancomycin-susceptible MRSA strain, was isolated from a patient with
pneumonia who responded favourably to vancomycin therapy. Subclones of Mu3
with increased resistance against vancomycin were selected with serial
concentrations of vancomycin and their MICs were determined. The
prevalence of VRSA and Mu3-like strains in Japanese hospitals was
estimated by population analysis from 1149 clinical MRSA isolates obtained
from 203 hospitals throughout Japan. The genetic traits of the Mu3 and
Mu50 strains were compared with clonotypes of MRSA from around the world.
FINDINGS: Mu3 and Mu50 had an identical pulsed-field gel electrophoresis
banding pattern. When grown in a drug-free medium, Mu3 produced
subpopulation of cells with varying degrees of vancomycin resistance, thus
demonstrating natural heterogeneity, or variability, in susceptibility to
vancomycin. In the presence of vancomycin, Mu3 produced subclones with
resistance roughly proportional to the concentrations of vancomycin used.
Selection of Mu3 with 8 mg/L or more of vancomycin gave rise to subclones
with vancomycin resistance equal to that of Mu50 (MIC 8 mg/L) at a
frequency of 1/1,000,000. During screening of Japanese MRSA strains, no
strain of VRSA additional to Mu50 was found. The prevalence of MRSA
isolates heterogeneously resistant to vancomycin was 20% in Juntendo
University Hospital, 9.3% in the other seven university hospitals, and
1.3% in non-university hospitals or clinics. INTERPRETATION:
Heterogeneously resistant VRSA is a preliminary stage that allows
development into VRSA upon exposure to vancomycin. Heterogeneously
resistant VRSA was found in hospitals throughout Japan. This finding could
explain, at least partly, the frequent therapeutic failure of MRSA
infection with vancomycin in Japan.

<>

<1>Hiramatsu, K., Itou, T., Awaya, A.
<2>DIAGNOSTIC AGENT.
<3>Japanese Patent Office
<4>JP 1997224700 A
<5>
<6>1997
<7>
<>

<1>Hiraoka, N., Kita, K., Nakajima, H., Kimizuka, F., Obayashi, A.
<2>Site-Specific Restriction Endonucleases of Several Non-Pathogenic Bacteria.
<3>J. Ferment. Technol.
<4>63
<5>151-157
<6>1985
<7>Seventy-one non-pathogenic bacterial strains were surveyed for the presence of Type II
restriction endonucleases in aerobic culture. Five strains were found to contain specific
enzymes: CflI from Cellulomonas flavignea, GalI from Gluconobacter albidus, GceI from
Gluconobacter cerinus, HacI from Halococcus acetoinfaciens, and MflI from Microbacterium
flavum. These enzymes cleaved respectively the sequences of 5'-CTGCA^G-3', 5'-CCGC^GG-3',
5'-CCGC^GG-3', 5'-^GATC-3', and 5'Pu^GATCPy'3' at the positions indicated. HacI and
MflI digested DNA from the Escherichia coli Dam-strain but not from the Dam+ strain,
indicating that they react only with unmodified sequences. With NaCl or KCl, the optimal salt
concentrations were 40-60 mM for CflI and 175-300mM for HacI; GalI and GceI did not require
either salts. Activity was greatest at pH 7.1-7.5 for CflI and HacI, and pH 7.5-8.0 for GalI
and GceI.

<>

<1>Hiraoka, N., Kita, K., Nakajima, H., Obayashi, A.
<2>Purification and characterization of sequence-specific restriction endonuclease MflI.
<3>J. Ferment. Technol.
<4>62
<5>583-588
<6>1984
<7>Class II restriction endonuclease MflI was purified 790-fold from the crude extract of
Microbacterium flavum by chromatography on Phosphocellulose, DEAE-cellulose, and
Heparin-sepharose columns. The purified preparation was free from non-specific nucleases and
phosphatases. The enzyme required 7 to 20 mM Mg++ ions but not monovalent cations for its
activity, and the maximum activity was obtained at pH 8.0-8.5 in the Tris-HCl buffer system.
This enzyme recognized 5'-PuGATCPy-3' in DNA and cleaved between Pu and G in this sequence.
However, MflI digested DNA from Escherichia coli Dam- strain, but not DNA from its Dam+ (wild
type) strain, indicating that this enzyme only restricts the unmodified sequence.

<>

<1>Hirata, R., Anraku, Y.
<2>Mutations at the putative junction sites of the yeast Vma1 protein, the catalytic subunit of the vacuolar membrane H+-ATPase, inhibit its processing by protein splicing.
<3>Biochem. Biophys. Res. Commun.
<4>188
<5>40-47
<6>1992
<7>A single gene, VMA1, encodes the 69-kDa subunit of the vacuolar
membrane H+-ATPase in the yeast Saccharomyces cerevisiae.  We have proposed that the
subunit is synthesized as a precursor of 120 kDa (1,071 amino acids) and then converted to
the 69-kDa form by an unusual processing reaction, which removes the internal domain of
454 amino acids (residues 284-737) and joins the N- and C-terminal domains.  Cysteine to
serine mutations at residues 284 and 738, the residues that bracket the internal domain,
were introduced into the VMA1 gene by site-directed mutagenesis, and the mutant genes
were expressed in a null vma1 mutant.  Cells harboring either of the mutant vma1 genes
accumulate nonfunctional fragments of the subunit.  The mutation of Cys-284 inhibited the
cleavage of the N-terminal junction site.  Cys-738 -> Ser mutation appeared to block the
processing at both junction sites although the mutant gene yielded a small fraction of the
functional 69-kDa subunit.

<>

<1>Hirata, R., Ohsumi, Y., Nakano, A., Kawasaki, H., Suzuki, K., Anraku, Y.
<2>Molecular structure of a gene, VMA1, encoding the catalytic subunit of H+-translocating adenosine triphosphatase from vacuolar membranes of Saccharomyces cerevisiae.
<3>J. Biol. Chem.
<4>265
<5>6726-6733
<6>1990
<7>Subunit alpha of the vacuolar membrane H+-translocating adenosine triphosphatase of the yeast
Saccharomyces cerevisiae contains a catalytic site for ATP hydrolysis. N-terminal sequences of
six tryptic peptides of the subunit were determined. Based on the peptide sequence
information, a 39-base oligonucleotide probe was synthesized, and the gene encoding the
subunit (VMA1) was isolated from a genomic DNA library by hybridization. The nucleotide
sequence of the gene predicts a polypeptide of 1,071 amino acids with a calculated molecular
mass of 118,635 daltons, which is much larger than the value 67 kDa estimated on sodium
dodecyl sulfate-polyacrylamide gels. N- and C-terminal regions of the deduced sequence
(residues 1-284 and 739-1,071) are very similar to those of the catalytic subunits of carrot
(69 kDa) and Neurospora crassa (67 kDa) vacuolar membrane H+-ATPases (62 and 73% identity over
600 residues, respectively). The homologous regions also show about 25% sequence identity over
400 residues with beta-subunits of F0F1-ATPases. In contrast, the internal region containing
454 amino acid residues (residues 285-738) shows no detectable sequence similarities to any
known ATPase subunits and instead is similar to a yeast endonuclease encoded by the HO gene.
None of the six tryptic peptides is located in this internal region. Northern blotting
analysis detected a single mRNA of 3.5 kilobases, indicating that the gene has no introns.
Although the reason for the discrepancy in molecular mass is unclear at present, these results
suggest that a novel processing mechanism, which might involve a post-translational excision
of the internal region followed by peptide ligation, operates on the yeast VMA1 product. The
VMA1 gene has proved to be the same gene as the TFP1 gene whose dominant mutant allele
(TFP1-408) confers a dominant trifluoperazine resistance and Ca2+-sensitive growth. This and
our findings suggest that the vacuolar membrane H+-ATPase participates in maintenance of
cytoplasmic Ca2+ homeostasis.

<>

<1>Hirayama, N.
<2>Molecules controlling genetic information, Part 4, Restriction endonucleases.
<3>Gendai Kagaku
<4>368
<5>52-54
<6>2001
<7>
<>

<1>Hirsch, A.M. et al.
<2>Complete Genome Sequence of Micromonospora Strain L5, a Potential Plant-Growth-Regulating Actinomycete, Originally Isolated from Casuarina  equisetifolia Root Nodules.
<3>Genome Announcements
<4>1
<5>e00759-13
<6>2013
<7>Micromonospora species live in diverse environments and exhibit a broad range of  functions,
including antibiotic production, biocontrol, and degradation of
complex polysaccharides. To learn more about these versatile actinomycetes, we
sequenced the genome of strain L5, originally isolated from root nodules of an
actinorhizal plant growing in Mexico.

<>

<1>Hirsch, J.A., Wah, D.A., Dorner, L.F., Shildkraut, I., Aggarwal, A.K.
<2>Crystallization and preliminary X-ray analysis of restriction endonuclease FokI bound to DNA.
<3>FEBS Lett.
<4>403
<5>136-138
<6>1997
<7>FokI is a type IIs restriction endonuclease which recognizes an asymmetric DNA sequence and
cleaves DNA a short distance away from the sequence.  The enzyme is bipartite in nature with
its DNA recognition and cleavage functions located on distinct domains.  We report here
cocrystals of the complete FokI enzyme (579 amino acids) bound to a 20-bp DNA fragment
containing its recognition sequence.  The complex is amongst the largest protein-DNA complexes
to be crystallized, and required macroseeding techniques for optimal crystal growth.  The
cocrystals diffract to at least 2.8 Angstroms in resolution and belong to space group P21 with
unit cell dimensions of a=67.9, b=119.8, c=69.1 Angstroms, b=96.6 degrees.  Using specific
amino acid analysis we show that asymmetric unit contains a single FokI molecule bound to the
20-bp DNA fragment.  This paper reports the first cocrystals of a type IIs restriction
endonuclease.

<>

<1>Hirsch-Kauffmann, M., Sauerbier, W.
<2>Inhibition of modification and restriction for phages lambda and Tl by co-infecting T3.
<3>Mol. Gen. Genet.
<4>102
<5>89-94
<6>1968
<7>The host controlled modifications of phage lambda DNA by Escherichia coli B,K, and C(P1) can
be suppressed by preinfecting the bacteria with UV-irradiated phage T3.  Since UV-irradiated
T3 induces an enzyme which cleaves S-adenosylmethionine into homoserine and thiomethyl
adenosine, and since S-adenosylmethionine is the only methyl group donor for DNA methylation,
we conclude that methylation is a required step in the host controlled modification of
lambda-DNA.  T3 itself successfully infects E. coli K and B with its nonmethylated DNA.  Also,
restricted phage lambda or Tl will be accepted by the restrictive hosts E. coli B,K, and C(P1)
if these are preinfected with UV-T3.  It thus appears that T3 is capable of blocking the
restriction mechanisms in these hosts.  The inability of T3 to grow on C(P1) is not
understood.  Since T3-DNA is restricted but not degraded into nucleotides by E. coli C(P1) we
presume that degradation is not the initial step in restriction.

<>

<1>Hisata, K., Ito, T., Matsunaga, N., Komatsu, M., Jin, J., Li, S., Watanabe, S., Shimizu, T., Hiramatsu, K.
<2>Dissemination of multiple MRSA clones among community-associated methicillin-resistant Staphylococcus aureus infections from Japanese children with impetigo.
<3>J. Infect. Chemother.
<4>17
<5>609-621
<6>2011
<7>The proportion of MRSA strains that cause skin
and soft infections has recently increased. In 3 months we
have characterized 17 MRSA strains isolated from children
with impetigo at a Japanese hospital. Seventeen MRSA
strains belonged to 7 clones defined by clonal complex
(CC) in MLST genotype and type of SCCmec, which were
rarely identified among healthcare-associated MRSA: CC
91-SCCmecIIb (4 strains); CC91-SCCmecIIn (2 strains);
CC91-SCCmecIVa (2 strains); CC91-SCCmecV (4 strains);
CC88-SCCmecIVg (3 strains); CC1-SCCmecIVc (1 strain);
and CC5-SCCmecIVn (1 strain). Although one strain
belonged to CC5, which has been commonly identified in
healthcare-associated MRSA, it did not carry type II
SCCmec, but carried type IV SCCmec. Fourteen of the 17
strains carried exfoliative toxin a or b gene, and none
carried Panton-Valentine leukocidine gene. Furthermore,
we determined the entire nucleotide sequences of two type
V SCCmec elements carried by strains JCSC5952, a CC91
strain, and TSGH17, a Taiwanese CC59 strain. The structure
of SCCmecJCSC5952 was more than 99% homologous
in nucleotide identity with those of Taiwanese
PVL-positive ST59 MRSA strains TSGH17 and PM1,
which were designated as type V

<>

<1>Hjerde, E., Pierechod, M.M., Williamson, A.K., Bjerga, G.E., Willassen, N.P., Smalas, A.O., Altermark, B.
<2>Draft Genome Sequence of the Actinomycete Rhodococcus sp. Strain AW25M09, Isolated from the Hadsel Fjord, Northern Norway.
<3>Genome Announcements
<4>1
<5>e0005513
<6>2013
<7>The cold-adapted Rhodococcus sp. strain AW25M09 was isolated from an Atlantic hagfish caught
off the shore of northern Norway as part of an ongoing
bioprospecting project that aims to identify novel bacteria with biotechnological
potential. Here, we present the 5.8-Mb draft genome sequence, together with
details regarding the origin of the strain and its sequence assembly.

<>

<1>Hjorleifsdottir, S., Pelursdottir, S., Kristjansson, J.K., Korpela, J., Torsti, A.-M., Mattila, P.
<2>Screening of Rhodothermus marinus for restriction endonucleases.
<3>Thermophiles Sci. Technol.
<4>20
<5>17
<6>1992
<7>A screening of 43 Rhodothermus marinus strains for restriction endonucleases was done.  Two
types of DNA substrates were used in the screening assay.  They were T7 DNA and lambda DNA.  A
total of 27 strains (63%) showed cleavage of one or both substrates.  Restriction
endonuclease positive strains were compared with a dendrogram based on a number of
characteristics of the Rhodothermus marinus group genus.  The comparison revealed that
restriction endonucleases exist in certain groups of strains.  Characterization of the
restriction endonucleases indicate that all positive strains contain RmaI, an isoschizomer of
MaeI and 3 strains contain a second restriction endonuclease, an isoschizomer of EcoRV as
well.  RmaI, a type II restriction endonuclease, has been put on the market.

<>

<1>Hjorleifsdottir, S., Petursdottir, S.K., Korpela, J., Torsti, A.-M., Mattila, P., Kristjansson, J.K.
<2>Screening for restriction endonucleases in aerobic, thermophilic eubacteria.
<3>Biotechnol. Tech.
<4>10
<5>13-18
<6>1996
<7>A total of 216 Icelandic aerobic, heterotrophic, thermophiles belonging to three different
genera were screened for type II restriction endonucleases.  The frequency of positive strains
was 44% for both Thermus and Bacillus but 63% for Rhodothermus.  Approximately half of the
enzymes from each group were characterised and a total of 14 different restriction enzymes
were found.  In all cases they were isoschizomers of known enzymes.  Thermus contained 9
different types, Bacillus 6 and Rhodothermus had 3.  This is the first time that isoschizomers
of BspEI, BglI, EagI and EcoRV are found in Thermus and BstBI and EcoRV are found in
Rhodothermus.

<>

<1>Hlavaty, J.J., Benner, J.S., Hornstra, L.J., Schildkraut, I.
<2>Identification of the metal-binding sites of restriction endonucleases by Fe(2+)-mediated oxidative cleavage.
<3>Biochemistry
<4>39
<5>3097-3105
<6>2000
<7>Fenton chemistry [Fenton (1894) J. Chem. Soc. 65, 899-910] techniques were employed to
identify the residues involved in metal binding located at the active sites of restriction
endonucleases. This process uses transition metals to catalytically oxidize the peptide
linkage that is in close proximity to the amino acid residues involved in metal ligation.
Fe(2+) was used as the redox-active transition metal. It was expected that Fe(2+) would bind
to the endonucleases at the Mg(2+)-binding site [Liaw et al. (1993) Biochemistry 32,
7999-4003; Ermacora et al. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 6383-6387; Soundar and
Colman (1993) J. Biol. Chem. 268, 5264-5271; Wei et al. (1994) Biochemistry 33, 7931-7936;
Ettner et al. (1995) Biochemistry 34, 22-31; Hlavaty and Nowak (1997) Biochemistry 36,
15515-15525). Fe(2+)-mediated oxidation was successfully performed on TaqI endonuclease,
suggesting that this approach could be applied to a wide array of endonucleases [Cao and
Barany (1998) J. Biol. Chem. 273, 33002-33010]. The restriction endonucleases BamHI, FokI,
BglI, BglII, PvuII, SfiI, BssSI, BsoBI, EcoRI, EcoRV, MspI, and HinP1I were subjected to
oxidizing conditions in the presence of Fe(2+) and ascorbate. All proteins were inactivated
upon treatment with Fe(2+) and ascorbate. BamHI, FokI, BglI, BglII, PvuII, SfiI, BssSI, and
BsoBI were specifically cleaved upon treatment with Fe(2+)/ascorbate. The site of
Fe(2+)/ascorbate-induced protein cleavage for each enzyme was determined. The Fe(2+)-mediated
oxidative cleavage of BamHI occurs between residues Glu77 and Lys78. Glu77 has been shown by
structural and mutational studies to be involved in both metal ligation and catalysis [Newman
et al. (1995) Science 269, 656-663; Viadiu and Aggarwal (1998) Nat. Struct. Biol. 5, 910-916;
Xu and Schildkraut (1991) J. Biol. Chem. 266, 4425-4429]. The sites of
Fe(2+)/ascorbate-induced cleavage for PvuII, FokI, BglI, and BsoBI agree with the
metal-binding sites identified in their corresponding three-dimensional structures or from
mutational studies [Cheng et al. (1994) EMBO J. 13, 3297-3935; Wah et al. (1997) Nature 388,
97-100; Newman et al. (1998) EMBO J. 17, 5466-5476; Ruan et al. (1997) Gene 188, 35-39]. The
metal-binding residues of BglII, SfiI, and BssSI are proposed based on amino acid sequencing
of their Fe(2+)/ascorbate-generated cleavage fragments. These results suggest that Fenton
chemistry may be a useful methodology in identifying amino acids involved in metal binding in
endonucleases.

<>

<1>Ho, A.Y., Chow, K.H., Law, P.Y., Tse, H., Ho, P.L.
<2>Draft Genome Sequences of Two Multidrug-Resistant Acinetobacter baumannii Strains of Sequence Type ST92 and ST96.
<3>Genome Announcements
<4>1
<5>e00296-13
<6>2013
<7>The global epidemiology of multidrug-resistant Acinetobacter baumannii is dominated by a
limited number of clones. Here, we announce the draft genome
sequences of two multidrug-resistant A. baumannii strains, 1H8 and 4A3,
representing the major epidemic clones, sequence type 92 (ST92) and ST96,
respectively.

<>

<1>Ho, D.K., Wu, J.C., Santi, D.V., Floss, H.G.
<2>Stereochemical studies of the C-methylation of deoxycytidine catalyzed by HhaI methylase and the N-methylation of deoxyadenosine catalyzed by EcoRI methylase.
<3>Arch. Biochem. Biophys.
<4>284
<5>264-269
<6>1991
<7>The steric course of methyl group transfer catalyzed by two DNA methylases, HhaI methylase,
HhaI methylase, a DNA (cytosine-5) methyltransferase, and EcoRI methylase, which methylates at
N6 of adenosine, has been studied with (methyl-R)- and (methyl-S)-[methyl-2H1,3H]
adenosylmethionine as the methyl donor, using as substrates poly-d(GC) (HhaI) and the
dodecamer oligonucleotide duplex d(CGCGAATTCGCG) (EcoRI), respectively. The methylated
nucleotides were degraded to convert the chiral methyl groups into acetic acid for
configurational analysis. It was found that both enzymatic reactions proceed with inversion of
configuration of the methyl group.

<>

<1>Ho, N., Lingohr, E.J., Villegas, A., Cole, L., Kropinski, A.M.
<2>Genomic characterization of two new Salmonella bacteriophages: vB_SosS_Oslo and vB_SemP_Emek.
<3>Ann. Agrar. Sci.
<4>10
<5>18-23
<6>2012
<7>Salmonella are classified on the basis of their surface antigens of which the O-antigen, which
is the immunodominant portion of the lipopolysaccharide molecule, is highly variable among
different strains.  Enzymes modifying O-antigenes are often encoded by prophages which
represent the genomes of integrated temperate phages.  The expression of certain prophage
genes by the lysogen can often result in the change in serotype of the host.  It was
hypothesized that O-antigenes 14 and 20 of Salmonella strains might be encoded on prophages.
This report outlines the isolation and sequencing of two novel bacteriophages,one from each fo
Salmonella enterica serovars Oslo and Emek thought to have given rise to O-antigens 14 and 20,
respectively.  Further analysis through lysogen isolation and serotyping has proven otherwise.

<>

<1>Ho, P.L., Lo, W.U., Chan, J., Cheung, Y.Y., Chow, K.H., Yam, W.C., Lin, C.H., Que, T.L.
<2>pIMP-PH114 Carrying bla IMP-4 in a Klebsiella pneumoniae Strain is Closely Related to Other Multidrug-Resistant IncA/C2 Plasmids.
<3>Curr. Microbiol.
<4>68
<5>227-232
<6>2014
<7>The IncA/C plasmids are broad host-range vehicles which have been associated with
wide dissemination of CMY-2 among Enterobacteriaceae of human and animal origins.
Acquired metallo-beta-lactamases (MBLs) such as the IMP-type enzymes are
increasingly reported in multidrug-resistant Gram-negative bacteria worldwide,
particularly in Enterobacteriaceae. We described the complete sequence of the
first IMP-4-encoding IncA/C2 plasmid, pIMP-PH114 (151,885 bp), from a sequence
type 1 Klebsiella pneumoniae strain that was recovered from a patient who was
hospitalized in the Philippines. pIMP-PH114 consists of a backbone from the
IncA/C2 plasmids, with the insertion of a novel Tn21-like class 1 integron
composite structure (containing the cassette array bla IMP-4-qacG-aacA4-catB3,
followed by a class C beta-lactamase bla DHA-1 and the mercury resistance operon,
merRTPCADE) and a sul2-floR encoding region. Phylogenetic analysis of the IncA/C
repA sequences showed that pIMP-PH114 formed a subgroup with other IncA/C
plasmids involved in the international spread of CMY-2, TEM-24 and NDM-1.
Identical bla IMP-4 arrays have been described among different Enterobacteriaceae
and Acinetobacter spp. in China, Singapore and Australia but the genetic context
is different. The broad host range of IncA/C plasmids may have facilitated
dissemination of the bla IMP-4 arrays among different diverse groups of bacteria.

<>

<1>Ho, P.L., Poon, W.W., Loke, S.L., Leung, M.S., Chow, K.H., Wong, R.C., Yip, K.S., Lai, E.L., Tsang, K.W.
<2>Community emergence of CTX-M type extended-spectrum beta-lactamases among urinary Escherichia coli from women.
<3>J. Antimicrob. Chemother.
<4>60
<5>140-144
<6>2007
<7>OBJECTIVES: To conduct a territory-wide study of extended-spectrum
beta-lactamases (ESBLs) among community isolates of urinary Escherichia
coli from women in Hong Kong. METHODS: Up to 50 consecutive single-patient
E. coli isolates, collected from 13 laboratories in 2004, were studied.
The ESBLs were characterized by PCR sequencing using specific primers. The
epidemiological relationship of the isolates was studied by PFGE and
phylogenetic group PCRs. RESULTS: Forty-two ESBL producers were found
among 600 consecutive isolates tested. The ESBL prevalence was 7.3%
(15/205) for women aged 18-35 years, 5% (11/219) for women aged 36-50
years, 6.3% (4/63) for women aged 51-64 years and 10.6% (12/113) for women
aged >or=65 years (P=0.3). The ESBL-producing isolates were often
multidrug-resistant and CTX-M-14 was found in 37 isolates, CTX-M-15 in 3
isolates and CTX-M-3 in 2 isolates. PFGE revealed no significant clusters
among the ESBL producers. Overall, CTX-M-14 producers were significantly
more likely to belong to group D than non-ESBL producers [18/37 (48.6%)
versus 13/57 (22.8%), P=0.009]. However, 7 of 13 (53.8%) CTX-M-14
producers from women aged 18-35 years represented phylogenetic group B2,
compared with 7 of 24 (29.2%) for women of all other ages (P=0.1).
CONCLUSIONS: The study documented the community emergence of CTX-M as the
predominant ESBL type among urinary isolates from women. The spread of
CTX-M enzymes among isolates from young women is concerning and deserves
close monitoring.

<>

<1>Ho, W.S., Gan, H.M., Yap, K.P., Balan, G., Yeo, C.C., Thong, K.L.
<2>Genome Sequence of Multidrug-Resistant Escherichia coli EC302/04, Isolated from a Human Tracheal Aspirate.
<3>J. Bacteriol.
<4>194
<5>6691-6692
<6>2012
<7>Escherichia coli is an important etiologic agent of lower respiratory tract infections (LRTI).
Multidrug-resistant E. coli EC302/04 was isolated from a
tracheal aspirate, and its genome sequence is expected to provide insights into
antimicrobial resistance as well as adaptive and virulence mechanisms of E. coli
involved in LRTI.

<>

<1>Ho, Y., Kim, S.-J., Waring, R.B.
<2>A protein encoded by a group I intron in Aspergillus nidulans directly assists RNA splicing and is a DNA endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>8994-8999
<6>1997
<7>Some group I introns self-splice in vitro, but almost all are thought to be assisted by
proteins in vivo.  Mutational analysis has shown that the splicing of certain group I introns
depends upon a maturase protein encoded by the intron itself.  However the effect of a protein
on splicing can be indirect.  We now provide evidence that a mitochondrial intron-encoded
protein from Aspergillus nidulans directly facilitates splicing in vitro.  This demonstrates
that a maturase is an RNA splicing protein.  The protein-assisted reaction is as fast as that
of any other known group I intron.  Interestingly the protein is also a DNA endonuclease, an
activity required for intron mobilization.  Mobile elements frequently encode proteins that
promote their propagation.  Intron-encoded proteins that also assist RNA splicing would
facilitate both the transposition and horizontal transmission of introns.

<>

<1>Ho, Y., Waring, R.B.
<2>The maturase encoded by a group I intron from Aspergillus nidulans stabilizes RNA tertiary structure and promotes rapid splicing.
<3>J. Mol. Biol.
<4>292
<5>987-1001
<6>1999
<7>The AnCOB group I intron from Aspergillus nidulans self-splices, providing the
Mg(2+)concentration is >/=15 mM. The splicing reaction is greatly stimulated by a maturase
protein encoded within the intron itself. An initial structural and biochemical analysis of
the splicing reaction has now been performed. The maturase bound rapidly to the precursor RNA
(kon approximately 3x10^9 M^-1 min^-1) and remained tightly bound (koff</=0.04 min^-1). The
catalytic step of 5' splice-site cleavage occurred at a rate of up to 11 min^-1 under single
turnover conditions. The maturase- assisted reaction of heat-denatured RNA proceeded at a rate
of about 1 min^-1, arguing that there are early steps of folding that cannot be readily
facilitated by the protein. pH analysis revealed a biphasic profile with a pKa of 7.0. The
rate of the maturase-assisted reaction was independent of the Mg(2+)concentration down to 3
mM. Self-splicing in optimal Mg(2+)(>/=150 mM) was tenfold slower, in part because of the
existence of an equilibrium between folded and partially folded RNA. In contrast, the maturase
very effectively stabilized tertiary structure in 5 mM Mg(2+), a noticeable example being an
interaction between the P8 helix and a GNRA sequence that constitutes the L2 terminal loop of
the P2 helix. Formation of the 5' splice-site recognition helix was assisted by either the
maturase or high concentrations of Mg(2+). The maturase was required during splicing so it is
not a true chaperone. However, RNase protection assays and kinetic studies suggest that the
maturase recognizes and facilitates folding of an intron with limited tertiary structure and
even incomplete secondary structure.

<>

<1>Ho, Y.S., Adroub, S.A., Abadi, M., Al, A.B., Alkhateeb, R., Gao, G., Ragab, A., Ali, S., van Soolingen, D., Bitter, W., Pain, A., Abdallah, A.M.
<2>Complete Genome Sequence of Mycobacterium vaccae Type Strain ATCC 25954.
<3>J. Bacteriol.
<4>194
<5>6339-6340
<6>2012
<7>Mycobacterium vaccae is a rapidly growing, nontuberculous Mycobacterium species that is
generally not considered a human pathogen and is of major pharmaceutical
interest as an immunotherapeutic agent. We report here the annotated genome
sequence of the M. vaccae type strain, ATCC 25954.

<>

<1>Ho, Y.S., Adroub, S.A., Aleisa, F., Mahmood, H., Othoum, G., Rashid, F., Zaher, M., Ali, S., Bitter, W., Pain, A., Abdallah, A.M.
<2>Complete Genome Sequence of Mycobacterium fortuitum subsp. fortuitum Type Strain  DSM46621.
<3>J. Bacteriol.
<4>194
<5>6337-6338
<6>2012
<7>Mycobacterium fortuitum is a member of the rapidly growing nontuberculous mycobacteria (NTM).
It is ubiquitous in water and soil habitats, including
hospital environments. M. fortuitum is increasingly recognized as an
opportunistic nosocomial pathogen causing disseminated infection. Here we report
the genome sequence of M. fortuitum subsp. fortuitum type strain DSM46621.

<>

<1>Hobley, G., McKelvie, J.C., Harmer, J.E., Howe, J., Oyston, P.C.F., Roach, P.L.
<2>Development of rationally designed DNA N6 adenine methyltransferase inhibitors.
<3>Bioorg. Med. Chem. Lett.
<4>22
<5>3079-3082
<6>2012
<7>A series of bisubstrate inhibitors for DNA N6 adenine methyltransferase (Dam) have been
synthesized by linking an amine analogue of
S-adenosylmethionine to an aryl moiety designed to probe the binding
pocket of the DNA adenine base. An initial structure-activity
relationship study has identified substituents that increase inhibitor
potency to the similar to 10 mu M range and improve selectivity against
the human cytosine methyltransferase Dnmt1.

<>

<1>Hobom, G., Schwarz, E., Melzer, M., Mayer, H.
<2>Restriction endonuclease EcaI from Enterobacter cloacae.
<3>Nucleic Acids Res.
<4>9
<5>4823-4832
<6>1981
<7>Restriction endonuclease EcaI obtained from Enterobacter cloacae DSM30056
recognizes the group of heptanucleotide palindromes 5'-G-^G-T-N-A-C-C-3', and
on cleavage (arrow) produces fragments with 5' - terminal pentanucleotide
extensions.  It is identical in specificity with restriction endonuclease
BstEII from Bacillus stearothermophiulus ET.

<>

<1>Hobson, N., Price, N.L., Ward, J.D., Raivio, T.L.
<2>Generation of a restriction minus enteropathogenic Escherichia coli E2348/69 strain that is efficiently transformed with large, low copy plasmids.
<3>BMC Microbiol.
<4>8
<5>13
<6>2008
<7>Background: Many microbes possess restriction-modification systems that protect them from
parasitic DNA molecules. Unfortunately, the presence of a restriction-modification system in a
given microbe also hampers genetic analysis. Although plasmids can be successfully conjugated
into the enteropathogenic Escherichia coli strain E2348/69 and optimized protocols for
competent cell preparation have been developed, we found that a large, low copy (similar to
15) bioluminescent reporter plasmid, pJW15, that we modified for use in EPEC, was exceedingly
difficult to transform into E2348/69. We reasoned that a restriction-modification system could
be responsible for the low transformation efficiency of E2348/69 and sought to identify and
inactivate the responsible gene(s), with the goal of creating an easily transformable strain
of EPEC that could complement existing protocols for genetic manipulation of this important
pathogen.Results: Using bioinformatics, we identified genes in the unfinished enteropathogenic
Escherichia coli (EPEC) strain E2348/69 genome whose predicted products bear homology to the
HsdM methyltransferases, HsdS specificity subunits, and HsdR restriction endonucleases of type
I restriction-modification systems. We constructed a strain carrying a deletion of the
conserved enzymatic domain of the EPEC HsdR homologue, NH4, and showed that its transformation
efficiency was up to four orders of magnitude higher than that of the parent strain. Further,
the modification capacity of NH4 remained intact, since plasmids that were normally
recalcitrant to transformation into E2348/69 could be transformed upon passage through NH4.
NH4 was unaffected in virulence factor production, since bundle forming pilus (BFP) subunits
and type III secreted (T3S) proteins were present at equivalent levels to those seen in
E2348/69. Further, NH4 was indistinguishable from E2348/69 in tissue culture infection model
assays of localized adherence and T3S.Conclusion: We have shown that EPEC strain E2348/69
utilizes a type 1 restriction- modification system to limit entry of new DNA. This
restriction- modification system does not appear to be involved in virulence determinant
expression or infection phenotypes. The hsdR mutant strain should prove useful in genetic
analysis of the important diarrheal pathogen EPEC.

<>

<1>Hobson, S.D., Falick, A.M., Reich, N.O.
<2>Identification of a critical histidine in EcoRI DNA methyltransferase using protein modification and tandem mass spectrometry.
<3>FASEB J.
<4>7
<5>A1275
<6>1993
<7>Diethyl pyrocarbonate (DEPC) is a known histidine specific reagent. Reaction of DEPC with
EcoRI DNA methyltransferase causes a time-dependent loss of activity due to modification of a
critical histidine(s) in the enzyme. The second order rate constant of the inactivation of the
enzyme is 570 M-1s-1. A kinetic analysis of the inactivation by DEPC has been used to
determine that only one of the seven histidines in the enzyme is critical. The number of
critical histidines has also been determined spectrophotometrically. This analysis revealed
that approximately 1.75 histidines are modified by DEPC-mediated inactivation suggests that
the residue has a pKa of about 6.0. The modified histidines are being identified via tandem
mass spectrometry methods in a novel LC-m.s. setup.

<>

<1>Hochhut, B., Wilde, C., Balling, G., Middendorf, B., Dobrindt, U., Brzuszkiewicz, E., Gottschalk, G., Carniel, E., Hacker, J.
<2>Role of pathogenicity island-associated integrases in the genome plasticity of uropathogenic Escherichia coli strain 536.
<3>Mol. Microbiol.
<4>61
<5>584-595
<6>2006
<7>The genome of uropathogenic Escherichia coli isolate 536 contains five well-characterized
pathogenicity islands (PAIs) encoding key virulence
factors of this strain. Except PAI IV(536), the four other PAIs of strain
536 are flanked by direct repeats (DRs), carry intact integrase genes and
are able to excise site-specifically from the chromosome. Genome screening
of strain 536 identified a sixth putative asnW-associated PAI. Despite the
presence of DRs and an intact integrase gene, excision of this island was
not detected. To investigate the role of PAI-encoded integrases for the
recombination process the int genes of each unstable island of strain 536
were inactivated. For PAI I(536) and PAI II(536), their respective P4-like
integrase was required for their excision. PAI III(536) carries two
integrase genes, intA, encoding an SfX-like integrase, and intB, coding
for an integrase with weak similarity to P4-like integrases. Only intB was
required for site-specific excision of this island. For PAI V(536),
excision could not be abolished after deleting its P4-like integrase gene
but additional deletion of the PAI II(536)-specific integrase gene was
required. Therefore, although all mediated by P4-like integrases, the
activity of the PAI excision machinery is most often restricted to its
cognate island. This work also demonstrates for the first time the
existence of a cross-talk between integrases of different PAIs and shows
that this cross-talk is unidirectional.

<>

<1>Hochwind, K., Weinmaier, T., Schmid, M., van Hemert, S., Hartmann, A., Rattei, T., Rothballer, M.
<2>Draft Genome Sequence of Lactobacillus casei W56.
<3>J. Bacteriol.
<4>194
<5>6638
<6>2012
<7>We announce the draft genome sequence of Lactobacillus casei W56 in one contig. This strain
shows immunomodulatory and probiotic properties. The strain is also
an ingredient of commercially available probiotic products.

<>

<1>Hodges, R.A., Perler, F.B., Noren, C.J., Jack, W.E.
<2>Protein splicing removes intervening sequences in an archaea DNA polymerase.
<3>Nucleic Acids Res.
<4>20
<5>6153-6157
<6>1992
<7>The Vent DNA polymerase gene from Thermococcus litoralis contains two in-frame insertions that
must be spliced out to form the mature polymerase. Primer extension and cDNA PCR revealed no
evidence of spliced RNA to account for this editing. In contrast, pulse-chase analysis
indicated that expression constructs lacking the first insertion produced a protein precursor
in Escherichia coli that was processed post-translationally to form polymerase and I-TliI, the
endonuclease protein that is the product of the second insertion. At least one intermediate,
which migrated more slowly than the precursor and may be branched, was also detected. Amino
acid substitutions at the splice junction slowed or blocked the protein splicing reaction.
Processing occurs in several heterologous systems, indicating either self-splicing or
ubiquitous splicing factors. Processing occurs in a mutant lacking I-TliI endonuclease
activity, establishing the independence of splicing and endonuclease activities.

<>

<1>Hodges-Garcia, Y., Hagerman, P.J.
<2>Cytosine methylation can induce local distortion in the structure of duplex DNA.
<3>Biochemistry
<4>31
<5>7595-7599
<6>1992
<7>Methyl groups at the C5 position of pyrimidines located within oligopurine-oligopyrimidine
tracts in DNA have been shown previously to modulate curvature generated by those tracts.
However, it was not known whether the influence of such methyl groups is consequent to the
altered helical structure within the tracts themselves. In the current study, it is
demonstrated that methylation of cytosines up to three base pairs away from a (dA)5(dT)5 tract
(A-tract) can still result in alterations of the net curvature of the A-tract-containing DNA,
as measured by alterations in electrophoretic mobility. This latter effect depends strongly on
both the sequence of the non-A-tract DNA and the positions of the methylated C residues. The
current results lend further support to the notion that the biological consequences of
cytosine methylation may be effected through local alteration in DNA structure as well as
through direct protein-DNA interactions.

<>

<1>Hodgson, C.P.
<2>A rapid visual method for the identification of 4- or 6-base restriction endonuclease sites.
<3>Biotechniques
<4>7
<5>148-149
<6>1989
<7>In this report, we describe a rapid approach to identification of restriction
endonuclease sites which makes computer searching unnecessary.  Increased speed
is possible because searching with specific sequences is eliminated,
substituting a one-time visual identification of palindromes.  These can be
marked and identified with the aid of a chart at a later time.  In this manner,
it is possible to identify 10-20 sites per minute with practice.  The technique
will be especially useful for short pieces of DNA (2 kb or less), for newly
published sequences, and for cloning experiments involving complex DNA
constructions which are not readily available on a database.

<>

<1>Hoefler, B.C., Konganti, K., Straight, P.D.
<2>De Novo Assembly of the Streptomyces sp. Strain Mg1 Genome Using PacBio Single-Molecule Sequencing.
<3>Genome Announcements
<4>1
<5>e00535-13
<6>2013
<7>We report a draft genome assembly of Streptomyces sp. strain Mg1, a competitive soil isolate
with multiple secondary metabolite gene clusters.

<>

<1>Hoekstra, M.F., Malone, R.E.
<2>Expression of the Escherichia coli dam methylase in Saccharomyces cerevisiae:  Effect of in vivo adenine methylation on genetic recombination and mutation.
<3>Mol. Cell. Biol.
<4>5
<5>610-618
<6>1985
<7>The Escherichia coli DNA adenine methylase (dam) gene has been introduced into Saccharomyces
cerevisiae on a yeast-E. coli shuttle vector. Sau3AI, MboI, and DpnI restriction enzyme
digests and Southern hybridization analysis indicated that the dam gene is expressed in yeast
cells and methylates GATC sequences. Analysis of digests of total genomic DNA indicated that
some GATC sites are not sensitive to methylation. The failure to methylate may reflect an
inaccessibility to the methylase due to chromosome structure. The effects of this in vivo
methylation on the processes of recombination and mutation in mitotic cells were determined. A
small but definite general increase was found in the frequency of mitotic recombination. A
similar increase was observed for reversion of some auxotrophic markers; other markers
demonstrated a small decrease in mutation frequency. The effects on mutation appear to be
locus (or allele) specific. Recombination in meiotic cells was measured and was not detectably
altered by the presence of 6-methyladenine in GATC sequences.

<>

<1>Hoekstra, W.P.M., DeHaan, P.G.
<2>The location of the restriction locus for lambda.K in Escherichia coli.
<3>Mutat. Res.
<4>2
<5>204-212
<6>1965
<7>Analysis of recombinants from E. coli K12 Hfr x E. coli B F- crosses showed
that one locus on the chromososme of Escherichia coli, controlling restriction
and probably also the modification of phage lambda, is located between the
leading point of the Hfr H chromosome and the locus for threonine synthesis.
The restriction locus also controls the restriction of the fertility factor of
K12 and the restriction of chromosomal DNA of K12 in E. coli B.  The presence
of the defective prophage X in E. coli B causes an additional restriction for
phage lambda and for the fertility factor.

<>

<1>Hoelzer, K., Shackelton, L.A., Parrish, C.R.
<2>Presence and role of cytosine methylation in DNA viruses of animals.
<3>Nucleic Acids Res.
<4>36
<5>2825-2837
<6>2008
<7>Nucleotide composition varies greatly among DNA viruses of animals, yet the evolutionary
pressures and biological mechanisms driving these patterns are unclear. One of the most
striking discrepancies lies in the frequency of CpG (the dinucleotide CG, linked by a
phosphate group), which is underrepresented in most small DNA viruses (those with genomes
below 10 kb) but not in larger DNA viruses. Cytosine methylation might be partially
responsible, but research on this topic has focused on a few virus groups. For several viruses
that integrate their genome into the host genome, the methylation status during this stage has
been studied extensively, and the relationship between methylation and viral-induced tumor
formation has been examined carefully. However, for actively replicating viruses-particularly
small DNA viruses-the methylation status of CpG motifs is rarely known and the effects on the
viral life cycle are obscure. In vertebrate host genomes, most cytosines at CpG sites are
methylated, which in vertebrates acts to regulate gene expression and facilitates the
recognition of unmethylated, potentially pathogen-associated DNA. Here we briefly introduce
cytosine methylation before reviewing what is currently known about CpG methylation in DNA
viruses.

<>

<1>Hoet, P.P., Coene, M.M., Cocito, C.G.
<2>Replication cycle of Bacillus subtilis hydroxymethyluracil-containing phages.
<3>Annu. Rev. Microbiol.
<4>46
<5>95-116
<6>1992
<7>The present review focuses on phage 2C, a member of a family of virulent phages that multiply
in Bacillus subtilis. The best known members of this group are SP01, Phie, H1, 2C, SP8, and
Sp82, the genomes of which are made of double-stranded DNA of about 150 kilobase pairs (kbp).
The two DNA strands have different buoyant densities. Moreover, thymine (T) is completely
replaced by hydroxymethyluracil (hmUra). Comparison of the phage DNAs has shown that both base
substitutions and deletions have contributed to the evolution of their genomes. In addition,
all of the hmUra-phage genomes contain colinear redundant ends, amounting to 10% of total
bases. Two lines of evidence suggest that the redundant ends of 2C DNA, in spite of extensive
homology, contain unique sequences. Further studies focused on DNA replication during the
lytic cycle. The semiconservative replication of the infecting viral genome is followed by
extensive recombination. At the level of replication forks, viral DNA synthesis in
permeabilized infected bacteria, was incorporated in small amounts into phage DNA. The
putative primary origin of replication has been cloned and localized on the viral genome. Some
viral promoters have been successfully cloned in Escherichia coli. These sequences, however,
did not promote transcription in B. subtilis. The abnormal base might be required for promoter
activity in the natural host.

<>

<1>Hofer, B.
<2>The sensitivity of DNA cleavage by SpeI and ApaLI to methylation by M.EcoK.
<3>Nucleic Acids Res.
<4>16
<5>5206
<6>1988
<7>During work on site-directed mutagenesis of the human interleukin-2 gene an
EcoK site was created which overlapped recognition sequences for SpeI and ApaLI
(Fig. 1a).  Isolation of the DNA from m+K strain DH1 and m-K strain HB101 and
subsequent incubation with the two restriction endonucleases revealed that EcoK
methylation completely or almost completely protected both DNA strands from
cleavage by SpeI, but did not prevent cleavage of either strand by ApaLI (Fig.
1b).  Thus, methylation of only one of the 5'-terminal A's of the SpeI site is
sufficient to protect it against SpeI, whereas methylation of one of the two
A's of the ApaLI sequence does not interfere with its cleavage by ApaLI.

<>

<1>Hofer, B., Koster, H.
<2>On the influence of thymidine analogues on the activity of phage fd promoters in vitro.
<3>Nucleic Acids Res.
<4>8
<5>6143-6162
<6>1980
<7>RF I DNA of phage fd containing 5-bromo-deoxyuridine (br5Ud) or deoxyuridine
(Ud) instead of deoxythymidine (Td) in the codogenic strand was synthesized in
vitro.  The modified genomes could be cleaved by restriction endonuclease
HpaII.  Although the recognition site of HpaII is CCGG, the cleavage rate was
significantly reduced with Ud-containing DNA.  Both base substitutions altered
the mobilities of several DNA fragments under the conditions of polyacrylamide
gel electrophoresis.  The fragments containing binding sites for RNA polymerase
were assayed for the rates of stable complex formation.  The substitution of Td
for both, Ud and br5Ud, strongly influenced this parameter.  Thus the methyl
group of Td has to be regarded as one of the sites in DNA which determine the
rate of stable RNA polymerase binding and thereby possibly mediate promoter
activity in vitro.  In most cases the rate of complex formation was decreased
by Ud, but increased by br5Ud.

<>

<1>Hofer, B., Kuhlein, B.
<2>The sensitivity of DNA cleavage by SnoI to methylation by M.EcoK.
<3>Nucleic Acids Res.
<4>17
<5>8009
<6>1989
<7>None

<>

<1>Hofer, B., Ruhe, G., Koch, A., Koster, H.
<2>Primary and secondary structure specificity of the cleavage of single-stranded DNA by endonuclease HinfI.
<3>Nucleic Acids Res.
<4>10
<5>2763-2773
<6>1982
<7>The interaction of endonuclease HinfI with single-stranded fd DNA was examined.
The sizes of the cleavage products indicate that the enzyme cuts this
substrate at the same sequences as double-stranded DNA (GANTC).  To determine
whether or not the recognition sites in a single-stranded DNA have to be
present in double-stranded form in order to be cleaved, DNA fragments
containing complementary or non-complementary HinfI sequences were prepared and
tested as substrates.  The results suggest that completely base-paired
recognition sites are necessary for cleavage.  Sequences surrounding the HinfI
pentanucleotides significantly modulate the reaction rates.

<>

<1>Hoffman, L.M., Jendrisak, J.J.
<2>Improving DNA transformation efficiency comprises introducing into the cell an OCR (Overcomes Classical Restriction) protein and the unmodified transforming DNA - involving shuttle vector plasmid, cosmid or fosmid-mediated phage T3 or phage T7 gene.
<3>US Patent Office
<4>US 7101713
<5>
<6>2006
<7>DERWENT ABSTRACT: NOVELTY - Improving transformation efficiency of a prokaryotic host cell
having a Type I restriction and modification
(R-M) system by an unmodified transforming DNA comprises introducing
into the prokaryotic host cell an isolated OCR (overcomes classical
restriction) protein, which protein is encoded by a T3 or T7
bacteriophage gene, together with the unmodified transforming DNA.
BIOTECHNOLOGY - Preferred Method: The prokaryotic host cell is
Eubacteria or Archaebacteria. The OCR protein is encoded by the 0.3 ocr
gene of bacteriophage T7 or bacteriophage T3. The OCR protein and the
unmodified transforming DNA are introduced into the prokaryotic host
cell by electroporation. The unmodified transforming DNA is a
polynucleotide comprising recombinant DNA in a vector. The unmodified
transforming DNA is a transposon or an artificial transposon. The
unmodified transforming DNA is a polynucleotide selected from a
plasmid, a suicide plasmid, a shuttle vector, a cosmid, a fosmid, an
oligonucleotide, an amplicon, an episome, a bacterial artificial
chromosome (BAC), a yeast artificial chromosome (YAC), or a replicon.
USE - The method is useful for improving transformation efficiency of a
prokaryotic host cell having a Type I restriction and modification
(R-M) system by an unmodified transforming DNA (claimed). ADVANTAGE -
The method improves the transforming efficiency of a prokaryotic host
cell having a Type I restriction and modification (R-M) system
(claimed). 1 microgram of the T7 OCR orotein was added to an
electroporation cuvette along with electrocompetent Salmonella
typhimurium strain LT2, and then pUC19 plasmid was added and the cells
were electroporated as before. The transformation efficiency of the 65
pUC19 DNA for the S. typhimurium strain LT2 was approximately one
hundred-fold higher in the presence of OCR protein than in its
absence.

<>

<1>Hoffmann, M., Luo, Y., Lafon, P.C., Timme, R., Allard, M.W., McDermott, P.F., Brown, E.W., Zhao, S.
<2>Genome Sequences of Salmonella enterica Serovar Heidelberg Isolates Isolated in the United States from a Multistate Outbreak of Human Salmonella Infections.
<3>Genome Announcements
<4>1
<5>e00004-12
<6>2013
<7>Salmonella enterica is recognized as one of the most common bacterial agents of foodborne
illness. We report draft genomes of four Salmonella serovar Heidelberg isolates associated
with the recent multistate outbreak of human Salmonella Heidelberg infections linked to kosher
broiled chicken livers in the United States in 2011. Isolates 2011K-1259 and 2011K-1232 were
recovered from humans, whereas 2011K-1724 and 2011K-1726 were isolated from chicken liver.
Whole genome sequence analysis of these isolates provides a tool for studying the short-term
evolution of these epidemic clones and can be used for characterizing potentially new
virulence factors.

<>

<1>Hoffmann, M., Muruvanda, T., Allard, M.W., Korlach, J., Roberts, R.J., Timme, R., Payne, J., McDermott, P.F., Evans, P., Meng, J., Brown, E.W., Zhao, S.
<2>Complete Genome Sequence of a Multidrug-Resistant Salmonella enterica Serovar Typhimurium var. 5- Strain Isolated from Chicken Breast.
<3>Genome Announcements
<4>1
<5>e01068-13
<6>2013
<7>Salmonella enterica subsp. enterica serovar Typhimurium is a leading cause of salmonellosis.
Here, we report a closed genome sequence, including sequences of 3
plasmids, of Salmonella serovar Typhimurium var. 5- CFSAN001921 (National
Antimicrobial Resistance Monitoring System [NARMS] strain ID N30688), which was
isolated from chicken breast meat and shows resistance to 10 different
antimicrobials. Whole-genome and plasmid sequence analyses of this isolate will
help enhance our understanding of this pathogenic multidrug-resistant serovar.

<>

<1>Hoffmann, M., Muruvanda, T., Allard, M.W., Korlach, J., Roberts, R.J., Timme, R., Payne, J., McDermott, P.F., Evans, P., Meng, J., Brown, E.W., Zhao, S.
<2>Complete Genome Sequence of a Multidrug-Resistant Salmonella enterica Serovar Typhimurium var. 5- Strain Isolated from Chicken Breast.
<3>Genome Announcements
<4>2
<5>e00294-14
<6>2014
<7>Volume 1, no. 6, e01068-13, 2013.  Page 2: The following should be added to the
Acknowledgments.  "Research reported in this publication was supported by the Small Business
Innovation Research Program (NIGMS) of the National Institutes of Health under award number
R44GM105125 to R.J.R.  The content is solely the responsibility of the authors and does not
necessarily represent the official views of the National Institutes of Health."

<>

<1>Hoffmann, M., Muruvanda, T., Pirone, C., Korlach, J., Timme, R., Payne, J., Evans, P., Meng, J., Brown, E.W., Allard, M.W.
<2>First Fully Closed Genome Sequence of Salmonella enterica subsp. enterica Serovar Cubana Associated with a Food-Borne Outbreak.
<3>Genome Announcements
<4>2
<5>e01112-14
<6>2014
<7>Salmonella enterica subsp. enterica serovar Cubana (Salmonella serovar Cubana) is associated
with human and animal disease. Here, we used third-generation,
single-molecule, real-time DNA sequencing to determine the first complete genome
sequence of Salmonella serovar Cubana CFSAN002050, which was isolated from fresh
alfalfa sprouts during a multistate outbreak in 2012.

<>

<1>Hoffmann, M., Zhao, S., Luo, Y., Li, C., Folster, J.P., Whichard, J., Allard, M.W., Brown, E.W., McDermott, P.F.
<2>Genome Sequences of Five Salmonella enterica Serovar Heidelberg Isolates Associated with a 2011 Multistate Outbreak in the United States.
<3>J. Bacteriol.
<4>194
<5>3274-3275
<6>2012
<7>Salmonella enterica serovar Heidelberg has caused numerous outbreaks in humans. Here, we
report draft genomes of five isolates of serovar Heidelberg associated
with the recent (2011) multistate outbreak linked to ground turkey in the United
States. Isolates 2011K-1110 and 2011K-1132 were recovered from humans, while
isolates 2011K-1138, 2011K-1224, and 2011K-1225 were recovered from ground
turkey. Whole-genome sequence analysis of these isolates provides a tool for
studying the short-term evolution of these epidemic clones.

<>

<1>Hofreuter, D., Tsai, J., Watson, R.O., Novik, V., Altman, B., Benitez, M., Clark, C., Perbost, C., Jarvie, T., Du, L., Galan, J.E.
<2>Unique features of a highly pathogenic Campylobacter jejuni strain.
<3>Infect. Immun.
<4>74
<5>4694-4707
<6>2006
<7>Campylobacter jejuni, a major human enteric pathogen, exhibits significant strain-to-strain
differences which result in differences in pathogenic potential.  C. jejuni 81-176 is a highly
virulent strain that exhibits unique pathogenic features and is used by many research
laboratories.  We have determined the nucleotide sequence of its genome and compared it to the
genomes of other sequenced C. jejuni strains.  We identified a number of unique genetic
features which may confer specific metabolic and pathogenic properties on this strain.  We
have also identified regions of the C. jejuni genome that are hot spots for the integration of
horizontally acquired genetic material.  This information should help the understanding of the
pathogenesis of C. jejuni and, in particular, the unique features of this highly pathogenic
strain.

<>

<1>Hogg, G., Dimovski, K., Hiley, L., Holt, K.E.
<2>Draft Genome Sequences for Ten Salmonella enterica Serovar Typhimurium Phage Type 135 Variants.
<3>Genome Announcements
<4>1
<5>e00293-13
<6>2013
<7>Salmonella enterica serovar Typhimurium (S. Typhimurium) is a common cause of gastroenteritis
in humans. Here, we report the draft genome sequences of 10
isolates of an S. Typhimurium phage type 135 variant that is linked to
egg-associated outbreaks in Tasmania, Australia.

<>

<1>Hogg, J.S., Hu, F.Z., Janto, B., Boissy, R., Hayes, J., Keefe, R., Post, J.C., Ehrlich, G.D.
<2>Characterization and modeling of the Haemophilus influenzae core and supragenomes based on the complete genomic sequences of Rd and 12 clinical nontypeable strains.
<3>Genome Biology
<4>8
<5>R103
<6>2007
<7>BACKGROUND: The distributed genome hypothesis (DGH) posits that chronic
bacterial pathogens utilize polyclonal infection and reassortment of genic
characters to ensure persistence in the face of adaptive host defenses.
Studies based on random sequencing of multiple strain libraries suggested
that free-living bacterial species possess a supragenome that is much
larger than the genome of any single bacterium. RESULTS: We derived high
depth genomic coverage of nine nontypeable Haemophilus influenzae (NTHi)
clinical isolates, bringing to 13 the number of sequenced NTHi genomes.
Clustering identified 2,786 genes, of which 1,461 were common to all
strains, with each of the remaining 1,328 found in a subset of strains;
the number of clusters ranged from 1,686 to 1,878 per strain. Genic
differences of between 96 and 585 were identified per strain pair.
Comparisons of each of the NTHi strains with the Rd strain revealed
between 107 and 158 insertions and 100 and 213 deletions per genome. The
mean insertion and deletion sizes were 1,356 and 1,020 base-pairs,
respectively, with mean maximum insertions and deletions of 26,977 and
37,299 base-pairs. This relatively large number of small rearrangements
among strains is in keeping with what is known about the transformation
mechanisms in this naturally competent pathogen. CONCLUSION: A finite
supragenome model was developed to explain the distribution of genes among
strains. The model predicts that the NTHi supragenome contains between
4,425 and 6,052 genes with most uncertainty regarding the number of rare
genes, those that have a frequency of <0.1 among strains; collectively,
these results support the DGH.

<>

<1>Hoheisel, J.D., Nizetic, D., Lehrach, H.
<2>Control of partial digestion combining the enzymes dam methylase and MboI.
<3>Nucleic Acids Res.
<4>17
<5>9571-9582
<6>1989
<7>A method is described which allows the preparation of reproducible partial digests without
previous establishment of the incubation conditions. It is based on a combined application of
dam methylase and the restriction endonuclease MboI, both recognizing the sequence
5'-GATC-3' but MboI unable to cut the methylated site. Due to their competition for the same
substrate the DNA is partially digested with the size of the resulting fragments strongly
dependent on the ratio of enzymes. The Km of the dam methylase was determined to be 115 ng
DNA/microliter indicating a variance in fragment sizes generated at low DNA-concentrations.
This effect is minimized above 150 ng/microliter. Any influence of digestion time is avoided,
because the reaction runs until complete modification of all sites. The dependence on enzyme
concentration and presence of agarose was checked. Knowledge of these parameters allows an
accurate prediction of fragment sizes generated at different conditions. The technique was
successfully used to construct libraries from different sources, in particular
chromosome-specific libraries from small amounts of flow-sorted material.

<>

<1>Holden, M.T. et al.
<2>Complete genome of acute rheumatic fever-associated serotype M5 Streptococcus pyogenes strain manfredo.
<3>J. Bacteriol.
<4>189
<5>1473-1477
<6>2007
<7>Comparisons of the 1.84-Mb genome of serotype M5 Streptococcus pyogenes strain Manfredo with
previously sequenced genomes emphasized the role of
prophages in diversification of S. pyogenes and the close relationship
between strain Manfredo and MGAS8232, another acute rheumatic
fever-associated strain.

<>

<1>Holden, M.T. et al.
<2>The genome of Burkholderia cenocepacia J2315, an epidemic pathogen of cystic fibrosis patients.
<3>J. Bacteriol.
<4>191
<5>261-277
<6>2009
<7>Bacterial infections of the lungs of cystic fibrosis (CF) patients cause major complications
in the treatment of this common genetic disease. Burkholderia
cenocepacia infection is particularly problematic since this organism has high
levels of antibiotic resistance, making it difficult to eradicate; the resulting
chronic infections are associated with severe declines in lung function and
increased mortality rates. B. cenocepacia strain J2315 was isolated from a CF
patient and is a member of the epidemic ET12 lineage that originated in Canada or
the United Kingdom and spread to Europe. The 8.06-Mb genome of this highly
transmissible pathogen comprises three circular chromosomes and a plasmid and
encodes a broad array of functions typical of this metabolically versatile genus,
as well as numerous virulence and drug resistance functions. Although B.
cenocepacia strains can be isolated from soil and can be pathogenic to both
plants and man, J2315 is representative of a lineage of B. cenocepacia rarely
isolated from the environment and which spreads between CF patients. Comparative
analysis revealed that ca. 21% of the genome is unique in comparison to other
strains of B. cenocepacia, highlighting the genomic plasticity of this species.
Pseudogenes in virulence determinants suggest that the pathogenic response of
J2315 may have been recently selected to promote persistence in the CF lung. The
J2315 genome contains evidence that its unique and highly adapted genetic content
has played a significant role in its success as an epidemic CF pathogen.

<>

<1>Holden, M.T., Lindsay, J.A., Corton, C., Quail, M.A., Cockfield, J.D., Pathak, S., Batra, R., Parkhill, J., Bentley, S.D., Edgeworth, J.D.
<2>Genome Sequence of a Recently Emerged, Highly Transmissible, Multi-Antibiotic- and Antiseptic-Resistant Variant of  Methicillin-Resistant Staphylococcus aureus, Sequence Type 239 (TW).
<3>J. Bacteriol.
<4>192
<5>888-892
<6>2010
<7>The 3.1-Mb genome of an outbreak methicillin-resistant Staphylococcus aureus (MRSA) strain
(TW20) contains evidence of recently acquired DNA,
including two large regions (635 kb and 127 kb). The strain is resistant
to a wide range of antibiotics, antiseptics, and heavy metals due to
resistance genes encoded on mobile genetic elements and also mutations in
housekeeping genes.

<>

<1>Holden, M.T.G. et al.
<2>Complete genomes of two clinical Staphylococcus aureus strains: Evidence for the rapid evolution of virulence and drug resistance.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>9786-9791
<6>2004
<7>Staphylococcus aureus is an important nosocomial and community-acquired pathogen. Its genetic
plasticity has facilitated the evolution of many virulent and drug-resistant strains,
presenting a major and constantly changing clinical challenge. We sequenced the ~2.8-Mbp
genomes of two disease-causing S. aureus strains isolated from distinct clinical settings: a
recent hospital-acquired representative of the epidemic methicillin-resistant S. aureus
EMRSA-16 clone (MRSA252), a clinically important and globally prevalent lineage; and a
representative of an invasive community-acquired methicillin-susceptible S. aureus clone
(MSSA476). A comparative-genomics approach was used to explore the mechanisms of evolution of
clinically important S. aureus genomes and to identify regions affecting virulence and drug
resistance. The genome sequences of MRSA252 and MSSA476 have a well conserved core region but
differ markedly in their accessory genetic elements. MRSA252 is the most genetically diverse
S. aureus strain sequenced to date: ~6% of the genome is novel compared with other published
genomes, and it contains several unique genetic elements. MSSA476 is methicillin-susceptible,
but it contains a novel Staphylococcal chromosomal cassette (SCC) mec-like element (designated
SCC476), which is integrated at the same site on the chromosome as SCCmec elements in MRSA
strains but encodes a putative fusidic acid resistance protein. The crucial role that
accessory elements play in the rapid evolution of S. aureus is clearly illustrated by
comparing the MSSA476 genome with that of an extremely closely related MRSA community-acquired
strain; the differential distribution of large mobile elements carrying virulence and
drug-resistance determinants may be responsible for the clinically important phenotypic
differences in these strains.

<>

<1>Holden, M.T.G. et al.
<2>Genomic plasticity of the causative agent of melioidosis, Berkholderia pseudomallei.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>14240-14245
<6>2004
<7>Burkholderia pseudomallei is a recognized biothreat agent and the causative agent of
melioidosis. This Gram-negative bacterium exists as a soil saprophyte in melioidosis-endemic
areas of the world and accounts for 20% of community-acquired septicaemias in northeastern
Thailand where half of those affected die. Here we report the complete genome of B.
pseudomallei, which is composed of two chromosomes of 4.07 megabase pairs and 3.17 megabase
pairs, showing significant functional partitioning of genes between them. The large chromosome
encodes many of the core functions associated with central metabolism and cell growth, whereas
the small chromosome carries more accessory functions associated with adaptation and survival
in different niches. Genomic comparisons with closely and more distantly related bacteria
revealed a greater level of gene order conservation and a greater number of orthologous genes
on the large chromosome, suggesting that the two replicons have distinct evolutionary origins.
A striking feature of the genome was the presence of 16 genomic islands (GIs) that together
made up 6.1% of the genome. Further analysis revealed these islands to be variably present in
a collection of invasive and soil isolates but entirely absent from the clonally related
organism B. mallei. We propose that variable horizontal gene acquisition by B. pseudomallei is
an important feature of recent genetic evolution and that this has resulted in a genetically
diverse pathogenic species.

<>

<1>Holden, M.T.G. et al.
<2>Rapid Evolution of Virulence and Drug Resistance in the Emerging Zoonotic Pathogen Streptococcus suis.
<3>PLoS ONE
<4>4
<5>e6072-e6072
<6>2009
<7>Background: Streptococcus suis is a zoonotic pathogen that infects pigs and can occasionally
cause serious infections in
humans. S. suis infections occur sporadically in human Europe and North America, but a recent
major outbreak has been
described in China with high levels of mortality. The mechanisms of S. suis pathogenesis in
humans and pigs are poorly
understood.
Methodology/Principal Findings: The sequencing of whole genomes of S. suis isolates provides
opportunities to
investigate the genetic basis of infection. Here we describe whole genome sequences of three
S. suis strains from the same
lineage: one from European pigs, and two from human cases from China and Vietnam. Comparative
genomic analysis was
used to investigate the variability of these strains. S. suis is phylogenetically distinct
from other Streptococcus species for
which genome sequences are currently available. Accordingly, ,40% of the ,2 Mb genome is
unique in comparison to
other Streptococcus species. Finer genomic comparisons within the species showed a high level
of sequence conservation;
virtually all of the genome is common to the S. suis strains. The only exceptions are three
,90 kb regions, present in the two
isolates from humans, composed of integrative conjugative elements and transposons. Carried in
these regions are coding
sequences associated with drug resistance. In addition, small-scale sequence variation has
generated pseudogenes in
putative virulence and colonization factors.
Conclusions/Significance: The genomic inventories of genetically related S. suis strains,
isolated from distinct hosts and
diseases, exhibit high levels of conservation. However, the genomes provide evidence that
horizontal gene transfer has
contributed to the evolution of drug resistance.

<>

<1>Holert, J., Alam, I., Larsen, M., Antunes, A., Bajic, V.B., Stingl, U., Philipp, B.
<2>Genome Sequence of Pseudomonas sp. Strain Chol1, a Model Organism for the Degradation of Bile Salts and Other Steroid Compounds.
<3>Genome Announcements
<4>1
<5>e00014-12
<6>2013
<7>Bacterial degradation of steroid compounds is of high ecological and biotechnological
relevance. Pseudomonas sp. strain Chol1 is a model organism for studying the degradation of
the steroid compound cholate. Its draft genome sequence is presented and reveals one gene
cluster responsible for the metabolism of steroid compounds.

<>

<1>Holland-Moritz, H.E., Bevans, D.R., Lang, J.M., Darling, A.E., Eisen, J.A., Coil, D.A.
<2>Draft Genome Sequence of Leucobacter sp. Strain UCD-THU (Phylum Actinobacteria).
<3>Genome Announcements
<4>1
<5>e00325-13
<6>2013
<7>Here we present the draft genome of Leucobacter sp. strain UCD-THU. The genome contains
3,317,267 bp in 11 scaffolds. This strain was isolated from a
residential toilet as part of an undergraduate project to sequence reference
genomes of microbes from the built environment.

<>

<1>Holland-Moritz, H.E., Coil, D.A., Badger, J.H., Dmitrov, G.I., Khouri, H., Ward, N.L., Robb, F.T., Eisen, J.A.
<2>Draft Genome Sequence of the Pyridinediol-Fermenting Bacterium Synergistes jonesii 78-1.
<3>Genome Announcements
<4>2
<5>e00833-14
<6>2014
<7>Here we present the draft genome of Synergistes jonesii 78-1, ATCC 49833, a member of the
Synergistes phylum. This organism was isolated from the rumen of a
Hawaiian goat and ferments pyridinediols. The assembly contains 2,747,397 bp in
61 contigs.

<>

<1>Holloway, S.P., Deshpande, N.N., Herrin, D.L.
<2>The catalytic group-I introns of the psbA gene of Chlamydomonas reinhardtii: core structures, ORFs and evolutionary implications.
<3>Curr. Genet.
<4>36
<5>69-78
<6>1999
<7>The sequences and predicted secondary structures of the four catalytic group-I introns in the
psbA gene of Chlamydomonas reinhardtii, Cr.psbA-1-Cr.psbA-4, have been determined. Cr.psbA-1
and Cr.psbA-4 are subgroup-IA1 introns and have similar secondary structures, except at the
3' end where Cr.psbA-1 contains a large inverted-repeat domain. Cr.psbA-4 is closely related
to intron 1 of the Chlamydomonas moewusii psbA gene, with which it shares the same location,
high nucleotide identity in the core, and an identically placed ORF that shows 58% amino-acid
identity. Cr.psbA-2 is a subgroup-IA3 intron, and shows similarities to the Chlamydomonas
eugametos rRNA intron, Ce.LSU-1. Cr.psbA-3 is a subgroup-IA2 intron, and is remarkably similar
to the T4 phage intron, sunY.  Interestingly, a degenerate version of Cr.psbA-3 is located in
the intergenic region between the chloroplast petA and petD genes. All four introns contain
ORFs, which potentially code for basic proteins of 11-38 kDa. The ORFs in introns 2 and 3
contain variants of the GIY-YIG motif; however, the Cr.psbA-2 ORF is free-standing, whereas
the Cr.psbA-3 ORF is contiguous and in-frame with the upstream exon. The Cr.psbA-4 ORF
contains an H-N-H motif, and possibly a GIY-YIG motif. These data indicate that the C.
reinhardtii psbA introns have multiple origins, and illustrate some of the evolutionary DNA
dynamics associated with group-I introns in Chlamydomonas.

<>

<1>Holmes, M.L., Nuttall, S.D., Dyall-Smith, M.L.
<2>Construction and use of Halobacterial shuttle vectors and further studies on Haloferax DNA gyrase.
<3>J. Bacteriol.
<4>173
<5>3807-3813
<6>1991
<7>We report here on advances made in the construction of plasmid shuttle vectors suitable for
genetic manipulations in both Escherichia coli and halobacteria. Starting with a 20.4-kb
construct, pMDS1, new vectors were engineered which were considerably smaller yet retained
several alternative cloning sites. A restriction barrier observed when plasmid DNA was
transferred into Haloferax volcanii cells was found to operate via adenine methylation,
resulting in a 10^3 drop in transformation efficiency and the loss of most constructs by
incorporation of the resistance marker into the chromosome. Passing shuttle vectors through E.
coli dam mutants effectively avoided this barrier. Deletion analysis revealed that the gene(s)
for autonomous replication of pHK2 (the plasmid endogenous to Haloferax strain Aa2.2 and used
in the construction of pMDS1) was located within a 4.2-kb SmaI-KpnI fragment. Convenient
restriction sites were identified near the termini of the novobiocin resistance determinant
(gyrB), allowing the removal of flanking sequences (including gyrA). These deletions did not
appear to significantly affect transformation efficiencies or the novobiocin resistance
phenotype of halobacterial transformants. Northern blot hybridization with strand- and
gene-specific probes identified a single gyrB-gyrA transcript of 4.7 kb. This is the first
demonstration in prokaryotes that the two subunits of DNA gyrase may be cotranscribed.

<>

<1>Holmfeldt, K., Howard-Varona, C., Solonenko, N., Sullivan, M.B.
<2>Contrasting genomic patterns and infection strategies of two co-existing Bacteroidetes podovirus genera.
<3>Environ. Microbiol.
<4>16
<5>2501-2513
<6>2014
<7>Bacterial viruses (phages) are abundant, ecologically important biological
entities. However, our understanding of their impact is limited by model systems
that are primarily not well represented in nature, e.g. Enterophages and their
hosts. Here, we investigate genomic characteristics and infection strategies
among six aquatic Bacteroidetes phages that represent two genera of exceptionally
large ( approximately 70-75 kb genome) podoviruses, which were isolated from the
same seawater sample using Cellulophaga baltica as host. Quantitative host range
studies reveal that these genera have contrasting narrow (specialist) and broad
(generalist) host ranges, with one-step growth curves revealing reduced burst
sizes for the generalist phages. Genomic comparisons suggest candidate genes in
each genus that might explain this host range variation, as well as provide
hypotheses about receptors in the hosts. One generalist phage, phi38:1, was more
deeply characterized, as its infection strategy switched from lytic on its
original host to either inefficient lytic or lysogenic on an alternative host. If
lysogenic, this phage was maintained extrachromosomally in the alternative host
and could not be induced by mitomycin C. This work provides fundamental knowledge
regarding phage-host ranges and their genomic drivers while also exploring the
'host environment' as a driver for switching phage replication mode.

<>

<1>Hols, P., Hancy, F., Fontaine, L., Grossiord, B., Prozzi, D., Leblond-Bourget, N., Decaris, B., Bolotin, A., Delorme, C., Ehrlich, S.D., Guedon, E., Monnet, W., Renault, P., Kleerebezem, M.
<2>New insights in the molecular biology and physiology of Streptococcus thermophilus revealed by comparative genomics.
<3>FEMS Microbiol. Rev.
<4>29
<5>435-463
<6>2005
<7>Streptoeoccus thermophilus is a major dairy starter used for the manufacture of yoghurt and
cheese. The access to three genome
sequences, comparative genomics and multilocus sequencing analyses
suggests that this species recently emerged and is still undergoing a
process of regressive evolution towards a specialised bacterium for
growth in milk. Notably, S. thermophilus has maintained a
well-developed nitrogen metabolism whereas its sugar catabolism has
been subjected to a high level of degeneracy due to a paucity of carbon
sources in milk. Furthermore, while pathogenic streptococci are
recognised for a high capacity to expose proteins at their cell surface
in order to achieve cell adhesion or to escape the host immune system,
S. thermophilus has nearly lost this unique feature as well as many
virulence-related functions. Although gene decay is obvious in S.
thermophilus genome evolution, numerous small genomic islands, which
were probably acquired by horizontal gene transfer, comprise important
industrial phenotypic traits such as polysaccharide biosynthesis,
bacteriocin production, restriction-modification systems or oxygen
tolerance.

<>

<1>Holt, D.C., Holden, M.T., Tong, S.Y., Castillo-Ramirez, S., Clarke, L., Quail, M.A., Currie, B.J., Parkhill, J., Bentley, S.D., Feil, E.J., Giffard, P.M.
<2>A very early-branching Staphylococcus aureus lineage lacking the carotenoid pigment staphyloxanthin.
<3>Genome Biol. Evol.
<4>3
<5>881-895
<6>2011
<7>Here we discuss the evolution of the northern Australian Staphylococcus aureus
isolate MSHR1132 genome. MSHR1132 belongs to the divergent clonal complex 75
lineage. The average nucleotide divergence between orthologous genes in MSHR1132
and typical S. aureus is approximately sevenfold greater than the maximum
divergence observed in this species to date. MSHR1132 has a small accessory
genome, which includes the well-characterized genomic islands, nuSAalpha and
nuSabeta, suggesting that these elements were acquired well before the expansion
of the typical S. aureus population. Other mobile elements show mosaic structure
(the prophage varphiSa3) or evidence of recent acquisition from a typical S.
aureus lineage (SCCmec, ICE6013 and plasmid pMSHR1132). There are two differences
in gene repertoire compared with typical S. aureus that may be significant clues
as to the genetic basis underlying the successful emergence of S. aureus as a
pathogen. First, MSHR1132 lacks the genes for production of staphyloxanthin, the
carotenoid pigment that confers upon S. aureus its characteristic golden color
and protects against oxidative stress. The lack of pigment was demonstrated in
126 of 126 CC75 isolates. Second, a mobile clustered regularly interspaced short
palindromic repeat (CRISPR) element is inserted into orfX of MSHR1132. Although
common in other staphylococcal species, these elements are very rare within S.
aureus and may impact accessory genome acquisition. The CRISPR spacer sequences
reveal a history of attempted invasion by known S. aureus mobile elements. There
is a case for the creation of a new taxon to accommodate this and related
isolates.

<>

<1>Holt, J.P., Grant, A.J., Coward, C., Maskell, D.J., Quinlan, J.J.
<2>Identification of Cj1051c as a Major Determinant for the Restriction Barrier of Campylobacter jejuni Strain NCTC11168.
<3>Appl. Environ. Microbiol.
<4>78
<5>7841-7848
<6>2012
<7>Campylobacter jejuni is a leading cause of human diarrheal illness in the world, and research
on it has benefitted greatly by the completion
of several genome sequences and the development of molecular biology
tools. However, many hurdles remain for a full understanding of this
unique bacterial pathogen. One of the most commonly used strains for
genetic work with C. jejuni is NCTC11168. While this strain is readily
transformable with DNA for genomic recombination, transformation with
plasmids is problematic. In this study, we have identified a
determinant of this to be cj1051c, predicted to encode a
restriction-modification type IIG enzyme. Knockout mutagenesis of this
gene resulted in a strain with a 1,000-fold-enhanced transformation
efficiency with a plasmid purified from a C. jejuni host. Additionally,
this mutation conferred the ability to be transformed by plasmids
isolated from an Escherichia coli host. Sequence analysis suggested a
high level of variability of the specificity domain between strains and
that this gene may be subject to phase variation. We provide evidence
that cj1051c is active in NCTC11168 and behaves as expected for a type
IIG enzyme. The identification of this determinant provides a greater
understanding of the molecular biology of C. jejuni as well as a tool
for plasmid work with strain NCTC11168.

<>

<1>Holt, K.E., Baker, S., Weill, F.X., Holmes, E.C., Kitchen, A., Yu, J., Sangal, V., Brown, D.J., Coia, J.E., Kim, D.W., Choi, S.Y., Kim, S.H., da Silveira, W.D., Pickard, D.J., Farrar, J.J., Parkhill, J., Dougan, G., Thomson, N.R.
<2>Shigella sonnei genome sequencing and phylogenetic analysis indicate recent global dissemination from Europe.
<3>Nat. Genet.
<4>44
<5>1056-1059
<6>2012
<7>Shigella are human-adapted Escherichia coli that have gained the ability to invade the human
gut mucosa and cause dysentery, spreading efficiently via
low-dose fecal-oral transmission. Historically, S. sonnei has been predominantly
responsible for dysentery in developed countries but is now emerging as a problem
in the developing world, seeming to replace the more diverse Shigella flexneri in
areas undergoing economic development and improvements in water quality.
Classical approaches have shown that S. sonnei is genetically conserved and
clonal. We report here whole-genome sequencing of 132 globally distributed
isolates. Our phylogenetic analysis shows that the current S. sonnei population
descends from a common ancestor that existed less than 500 years ago and that
diversified into several distinct lineages with unique characteristics. Our
analysis suggests that the majority of this diversification occurred in Europe
and was followed by more recent establishment of local pathogen populations on
other continents, predominantly due to the pandemic spread of a single, rapidly
evolving, multidrug-resistant lineage.

<>

<1>Holt, K.E., Thomson, N.R., Wain, J., Phan, M.D., Nair, S., Hasan, R., Bhutta, Z.A., Quail, M.A., Norbertczak, H., Walker, D., Dougan, G., Parkhill, J.
<2>Multidrug-Resistant Salmonella enterica Serovar Paratyphi A Harbors IncHI1 Plasmids Similar to Those Found in Serovar Typhi.
<3>J. Bacteriol.
<4>189
<5>4257-4264
<6>2007
<7>Salmonella enterica serovars Typhi and Paratyphi A cause systemic
infections in humans which are referred to as enteric fever.
Multidrug-resistant (MDR) serovar Typhi isolates emerged in the 1980s, and
in recent years MDR serovar Paratyphi A infections have become established
as a significant problem across Asia. MDR in serovar Typhi is almost
invariably associated with IncHI1 plasmids, but the genetic basis of MDR
in serovar Paratyphi A has remained predominantly undefined. The DNA
sequence of an IncHI1 plasmid, pAKU_1, encoding MDR in a serovar Paratyphi
A strain has been determined. Significantly, this plasmid shares a common
IncHI1-associated DNA backbone with the serovar Typhi plasmid pHCM1 and an
S. enterica serovar Typhimurium plasmid pR27. Plasmids pAKU_1 and pHCM1
share 14 antibiotic resistance genes encoded within similar mobile
elements, which appear to form a 24-kb composite transposon that has
transferred as a single unit into different positions into their IncHI1
backbones. Thus, these plasmids have acquired similar antibiotic
resistance genes independently via the horizontal transfer of mobile DNA
elements. Furthermore, two IncHI1 plasmids from a Vietnamese isolate of
serovar Typhi were found to contain features of the backbone sequence of
pAKU_1 rather than pHCM1, with the composite transposon inserted in the
same location as in the pAKU_1 sequence. Our data show that these serovar
Typhi and Paratyphi A IncHI1 plasmids share highly conserved core DNA and
have acquired similar mobile elements encoding antibiotic resistance genes
in past decades.

<>

<1>Holtz, J.K., Topal, M.D.
<2>Location of putative binding and catalytic sites of NaeI by random mutagenesis.
<3>J. Biol. Chem.
<4>269
<5>27286-27290
<6>1994
<7>Endonuclease NaeI is a prototype for an unusual group of type II restriction endonucleases
that must bind two DNA recognition sequences to cleave DNA. The naeIR gene, expressed from a
Ptac promoter construct, was toxic to Escherichia coli in the absence of NaeI-sequence
specific methylases. The naeIR gene was mutagenized with N-methyl-N'-nitrosoguanidine; four
classes of NaeI variants were isolated in the absence of protecting methylase activity. Class
I variants (T60I, E70K) lacked detectable cleavage activity, but displayed good
sequence-specific DNA binding. Class II variants (D95N, G141D) displayed 1-5% of the wild-type
cleavage activity and normal DNA binding. Class III variants (G131E, G131R/A219T, G236S,
L241P, G245E, G245R, G250E, G270E) lacked both cleavage and binding activities. These results
imply two amino acids (Thr-60, Glu-70) essential for catalysis. In addition, two domains are
indicated in NaeI: one (Thr-60 to Gly-155) mediates substrate binding and catalysis, the other
(Gly-197 to Gly-270) may mediate binding of the activating DNA sequence. Our results are
compared with the active site residues of EcoRI, EcoRV, and BamHI.

<>

<1>Holubova, I., Vejsadova, I., Firman, K., Weiserovda, M.
<2>Cellular localization of type I restriction-modification enzymes is family dependent.
<3>Biochem. Biophys. Res. Commun.
<4>319
<5>375-380
<6>2004
<7>Cellular localization of Type I restriction-modification enzymes EcoKI, EcoAI, and
EcoR124I-the most frequently studied representatives of IA,
113, and IC families-was analyzed by immunoblotting of subcellular
fractions isolated from Escherichia coli strains harboring the
corresponding hsd genes. EcoR124I shows characteristics similar to
those of EcoKI. The complex enzymes are associated with the cytoplasmic
membrane via DNA interaction as documented by the release of the Hsd
subunits from the membrane into the soluble fraction following
benzonase treatment. HsdR subunits of the membrane-bound enzymes EcoKI
and EcoR124I are accessible, though to a different extent, at the
external surface of cytoplasmic membrane as shown by trypsinization of
intact spheroplasts. EcoAI strongly differs from EcoKI and EcoR124I,
since neither benzonase nor trypsin affects its association with the
cytoplasmic membrane. Possible reasons for such a different
organization are discussed in relation of the control of the
restriction-modification activities in vivo.

<>

<1>Holubova, I., Vejsadova, S., Weiserova, M., Firman, K.
<2>Localization of the Type I Restriction-Modification Enzyme EcoKI in the Bacterial Cell.
<3>Biochem. Biophys. Res. Commun.
<4>270
<5>46-51
<6>2000
<7>To localise the type I restriction-modification (R-M) enzyme EcoKI within the bacterial cell,
the Hsd subunits present in subcellular fractions were analysed using immunoblotting
techniques. The endonuclease (ENase) as well as the methylase (MTase) were found to be
associated with the cytoplasmic membrane. HsdR and HsdM subunits produced individually were
soluble, cytoplasmic polypeptides and only became membrane-associated when coproduced with the
insoluble HsdS subunit. The release of enzyme from the membrane fraction following benzonase
treatment indicated a role for DNA in this interaction. Trypsinization of spheroplasts
revealed that the HsdR subunit in the assembled ENase was accessible to protease, while HsdM
and HsdS, in both ENase and MTase complexes, were fully protected against digestion. We
postulate that the R-M enzyme EcoKI is associated with the cytoplasmic membrane in a manner
that allows access of HsdR to the periplasmic space, while the MTase components are localised
on the inner side of the plasma membrane.

<>

<1>Holubova, J., Josephsen, J.
<2>Potential of AbiS as defence mechanism determined by conductivity measurement.
<3>J. Appl. Microbiol.
<4>103
<5>2382-2391
<6>2007
<7>Aim: To compare pH and conductivity used in the determination of growth in reconstituted skim
milk (RSM), to determine whether the presence of
one or two plasmids in Lactococcus lactis had any influence on growth,
and whether AbiS improved bacteriophages resistance of L. lactis.
Methods and Results: Conductivity and pH were used to determine
growth in RSM. A small increase in the generation time was found with
increasing number of plasmids, while their size was unimportant. The
introduction of a plasmid-encoding AbiS did only enhance the level of
phage resistance significant when other plasmids encoding either AbiS1
or the restriction modification system LlaBIII was present.
Conclusions: The earliest detection of growth was observed by
measuring pH, rather than conductance. The plasmid-encoded AbiS system
has a potential to be used as a phage resistance mechanisms in L.
lactis during milk fermentations, especially when combined with other
anti-phage mechanisms.
Significance and Impact of the Study: This study widened the
knowledge about the influence of plasmid introduction on the growth
rate of L. lactis, which is important for the construction of new
strains. The level of protection against 936 groups of phages was only
significant when the mechanism was present together with the RM system
LlaBIII.

<>

<1>Holz, B., Dank, N., Eickhoff, J.E., Lipps, G., Krauss, G., Weinhold, E.
<2>Identification of the binding site for the extrahelical target base in N6-adenine DNA methyltransferases by photo-cross-linking with duplex oligodeoxyribonucleotides containing 5-iodouracil at the target position.
<3>J. Biol. Chem.
<4>274
<5>15066-15072
<6>1999
<7>DNA methyltransferases flip their target bases out of the DNA double helix for catalysis. Base
flipping of C5-cytosine DNA methyltransferases was directly observed in the protein-DNA
cocrystal structures of M.HhaI and M.HaeIII. Indirect structural evidence for base flipping of
N6-adenine and N4-cytosine DNA methyltransferases was obtained by modeling DNA into the
three-dimensional structures of M.TaqI and M.PvuII in complex with the cofactor. In addition,
biochemical evidence of base flipping was reported for different N6-adenine DNA
methyltransferases. As no protein-DNA cocrystal structure for the related N6-adenine and
N4-cytosine DNA methyltransferases is available, we used light-induced photochemical
cross-linking to identify the binding site of the extrahelical target bases. The N6-adenine
DNA methyltransferases M.TaqI and M.CviBIII, which both methylate adenine within the
double-stranded 5'-TCGA-3' DNA sequence, were photo-cross-linked to duplex
oligodeoxyribonucleotides containing 5-iodouracil at the target position in 50-60% and almost
quantitative yield, respectively. Proteolytic fragmentation of the M.CviBIII-DNA complex
followed by Edman degradation and electrospray ionization mass spectrometry indicates
photo-cross-linking to tyrosine 122. In addition, the mutant methyltransferases M.TaqI/Y108A
and M.TaqI/F196A were photo-cross-linked with 6-fold and 2-fold reduced efficiency,
respectively, which suggests that tyrosine 108 is the primary site of modification in M.TaqI.
Our results indicate a close proximity between the extrahelical target base and tyrosine 122
in M.CviBIII or tyrosine 108 in M.TaqI. As both residues belong to the conserved motif IV
((N/D/S)(P/I)P(Y/F/W)) found in all N6-adenine and N4-cytosine DNA as well as in N6-adenine
RNA methyltransferases, a similar spatial relationship between the target bases and the
aromatic amino acid residue within motif IV is expected for all these methyltransferases.

<>

<1>Holz, B., Klimasauskas, S., Serva, S., Weinhold, E.
<2>2-Aminopurine as a fluorescent probe for DNA base flipping by methyltransferases.
<3>Nucleic Acids Res.
<4>26
<5>1076-1083
<6>1998
<7>DNA base flipping, which was first observed for the C5-cytosine  methyltransferase M.HhaI,
results in a complete removal of the stacking interactions between the target base and its
neighboring bases.  We have investigated whether duplex oligodeoxynucleotides containing the
fluorescent base analogue 2-aminopurine can be used to sense DNA base flipping.  Using M.HhaI
as a paradigm for a base flipping enzyme, we find that the fluorescence intensity of duplex
oligodeoxynucleotides containing 2-aminopurine at the target site is dramatically enhanced
(54-fold) in the presence of M.HhaI.  Duplex oligodeoxynucleotides containing 2-aminopurine
adjacent to the target cytosine show little fluorescence increase upon addition of M.HhaI.
These results clearly demonstrate that duplex oligodeoxynucleotides containing 2-aminopurine
at the target site can serve as fluorescence probes for base flipping.  Another enzyme
hypothesized to use a base flipping mechanism is the N6-adenine DNA methyltransferase M.TaqI.
Addition of M.TaqI to duplex oligodeoxynucleotides bearing 2-aminopurine at the target
position, also results in a strongly enhanced fluorescence (13-fold), whereas addition to
duplex oligodeoxynucleotides containing 2-aminopurine at the 3'- or 5'-neighboring position
leads only to small fluorescence increases.  These results give the first experimental
evidence that the adenine-specific DNA methyltransferase M.TaqI also flips its target base.

<>

<1>Holz, B., Pues, H., Wolcke, J., Weinhold, E.
<2>Fluorescence studies on the base flipping mechanism of the DNA methyltransferase M.TaqI.
<3>FASEB J.
<4>11
<5>A1151
<6>1997
<7>The DNA methyltransferase from Thermus aquaticus catalyzes the methyl group transfer from
S-adenosyl-L-methionine to the N6-position of adenine in the double-stranded DNA sequence
5'-TCGA-3'.  A model of the ternary complex built with the crystal structure of M.TaqI-SAM
complex and DNA duplex shows that the distance between the cofactor and the target adenine is
too large to allow a direct methyl group transfer.  This distance can be decreased if the
adenine rotates out of the DNA helix as described for a C5-DNA methyltransferase.  DNA
containing the adenine analogue 2-aminopurine and intrinsic tryptophan fluorescence were used
to examine such a base flipping mechanism for M.TaqI.  The fluorescence of 2-Ap is highly
quenched in duplex DNA due to stacking of the neighboring bases.  Addition of M.TaqI to a DNA
duplex with 2-Ap at the target site causes a large fluorescence increase which points to an
extrahelical base in this complex.  In stopped-flow experiments this increase is single
exponential with a rate constant of 20 s^-1.  Binding of DNA leads to a 30% increase of the
tryptophan fluorescence intensity of M.TaqI.  This increase was resolved into two steps: a
rapid first step (k+2 = 20 s-1) and a second step with a k+3 = 2 s-1.  Since both steps are
not concentration dependent we suggest a very fast formation of an initial complex in a time
scale not resolved by the stopped-flow method.  These data suggest that DNA binding of M.TaqI
is at least a three step process, where the second step gives rise not only to a
conformational change of the DNA methyltransferase but also of the DNA.

<>

<1>Holz, B., Weinhold, E.
<2>Probes for DNA base flipping by DNA methyltransferases.
<3>Bioorganic Chemistry: Highlights and New Aspects, Wiley-VCH, Diederichsen, U., Lindhorst, T.K., Westermann, B., Wessjohann, L.A., Weinheim
<4>
<5>337-345
<6>1999
<7>In addition to the normal nucleobases adenine, thymine, guanine, and cytosine, the DNA of most
organisms contains the methylated bases C5-methylcytosine, N6-methyladenine, or
N4-methylcytosine.  These methylated bases are formed by DNA methyltransferases which catalyze
the transfer of the activated methyl group from the cofactor S-adenosyl-L-methionine to the C5
carbon of cytosine, the N6 nitrogen of adenine, or the N4 nitrogen of cytosine within specific
DNA sequences.  Most DNA Mtases recognize palindromic DNA sequences, which contain two
symmetry-related target bases.  After DNA replication, only the parental strand contains
methylated bases (hemi-methylated DNA), and methylation of the daughter strand restores the
fully methylated DNA.

<>

<1>Holz-Schietinger, C., Reich, N.O.
<2>The Inherent Processivity of the Human de Novo Methyltransferase 3A (DNMT3A) Is Enhanced by DNMT3L.
<3>J. Biol. Chem.
<4>285
<5>29091-29100
<6>2010
<7>Human DNMT3A is responsible for de novo DNA cytosine methylation patterning during
development. Here we show that DNMT3A methylates 5-8
CpG sites on human promoters before 50% of the initially bound enzyme
dissociates from the DNA. Processive methylation is enhanced 3-fold in
the presence of DNMT3L, an inactive homolog of DNMT3A, therefore
providing a mechanism for the previously described DNMT3L activation of
DNMT3A. DNMT3A processivity on human promoters is also regulated by DNA
topology, where a 2-fold decrease in processivity was observed on
supercoiled DNA in comparison with linear DNA. These results are the
first observation that DNMT3A utilizes this mechanism of increasing
catalytic efficiency. Processive de novo DNA methylation provides a
mechanism that ensures that multiple CpG sites undergo methylation for
transcriptional regulation and silencing of newly integrated viral DNA.

<>

<1>Holz-Schietinger, C., Reich, N.O.
<2>RNA modulation of the human DNA methyltransferase 3A.
<3>Nucleic Acids Res.
<4>40
<5>8550-8557
<6>2012
<7>DNA methyltransferase 3A (DNMT3A) is one of two human de novo DNA methyltransferases essential
for transcription regulation during cellular
development and differentiation. There is increasing evidence that RNA plays a
role in directing DNA methylation to specific genomic locations within mammalian
cells. Here, we describe two modes of RNA regulation of DNMT3A in vitro. We show
a single-stranded RNA molecule that is antisense to the E-cadherin promoter binds
tightly to the catalytic domain in a structurally dependent fashion causing
potent inhibition of DNMT3A activity. Two other RNA molecules bind DNMT3A at an
allosteric site outside the catalytic domain, causing no change in catalysis. Our
observation of the potent and specific in vitro modulation of DNMT3A activity by
RNA supports in vivo data that RNA interacts with DNMT3A to regulate
transcription.

<>

<1>Honda, S., Selker, E.U.
<2>Direct interaction between DNA methyltransferase DIM-2 and HP1 is required for DNA methylation in Neurospora crassa.
<3>Mol. Cell. Biol.
<4>28
<5>6044-6055
<6>2008
<7>DNA methylation is involved in gene silencing and genomic stability in mammals, plants, and
fungi. Genetics studies of Neurospora crassa have
revealed that a DNA methyltransferase (DIM-2), a histone H3K9
methyltransferase (DIM-5), and heterochromatin protein 1 (HP1) are
required for DNA methylation. We explored the interrelationships of
these components of the methylation machinery. A yeast two-hybrid
screen revealed that HP1 interacts with DIM-2. We confirmed the
interaction in vivo and demonstrated that it involves a pair of
PXVXL-related motifs in the N-terminal region of DIM-2 and the chromo
shadow domain of HP1. Both regions are essential for proper DNA
methylation. We also determined that DIM-2 and HP1 form a stable
complex independently of the trimethylation of histone H3K9, although
the association of DIM-2 with its substrate sequences depends on
trimethyl-H3K9. The DIM-2/HP1 complex does not include DIM-5. We
conclude that DNA methylation in Neurospora is largely or exclusively
the result of a unidirectional pathway in which DIM-5 methylates
histone H3K9 and then the DIM-2/HP1 complex recognizes the resulting
trimethyl-H3K9 mark via the chromo domain of HP1.

<>

<1>Honda, T., Liang, Y., Arai, D., Ito, Y., Yoshino, T., Tanaka, T.
<2>Draft Genome Sequence of Marine Cyanobacterium Synechococcus sp. Strain NKBG042902, Which Harbors a Homogeneous Plasmid Available for Metabolic  Engineering.
<3>Genome Announcements
<4>2
<5>e00704-14
<6>2014
<7>The marine cyanobacterium Synechococcus sp. strain NKBG042902 was isolated from coastal areas
in Japan. Strain NKBG042902 has four plasmids: pSY8, pSY9, pSY10,
and pSY11. Moreover, the hybrid plasmid pUSY02 containing pSY11 and Escherichia
coli plasmid pUC18 was constructed for this strain. The genetic manipulation
technique using pUSY02 was established for this strain and used in metabolic
engineering. Here, we report the draft genome sequence of this strain, which has
77 contigs comprising a total length of 3,319,479 bp, with a G+C content of
49.4%.

<>

<1>Hong, K.W., Gan, H.M., Low, S.M., Lee, P.K., Chong, Y.M., Yin, W.F., Chan, K.G.
<2>Draft Genome Sequence of Pantoea sp. Strain A4, a Rafflesia-Associated Bacterium  That Produces N-Acylhomoserine Lactones as Quorum-Sensing Molecules.
<3>J. Bacteriol.
<4>194
<5>6610
<6>2012
<7>Pantoea sp. strain A4 is a Gram-negative bacterium isolated from the Rafflesia flower. We
present here, for the first time, the genome sequence of
Rafflesia-associated Pantoea sp. strain A4, which exhibited quorum-sensing
activity.

<>

<1>Hong, K.W., Koh, C.L., Sam, C.K., Yin, W.F., Chan, K.G.
<2>Complete Genome Sequence of Burkholderia sp. Strain GG4, a Betaproteobacterium That Reduces 3-Oxo-N-Acylhomoserine Lactones and Produces Different  N-Acylhomoserine Lactones.
<3>J. Bacteriol.
<4>194
<5>6317
<6>2012
<7>Burkholderia sp. strain GG4, isolated from the ginger rhizosphere, possesses a unique
N-acylhomoserine lactone (AHL)-modifying activity that reduces 3-oxo-AHLs
to 3-hydroxy-AHLs. To the best of our knowledge, this is the first sequenced
genome from a bacterium of the genus Burkholderia that shows both quorum-sensing
and signaling confusion activities.

<>

<1>Hong, K.W., Koh, C.L., Sam, C.K., Yin, W.F., Chan, K.G.
<2>Whole-Genome Sequence of N-Acylhomoserine Lactone-Synthesizing and -Degrading Acinetobacter sp. Strain GG2.
<3>J. Bacteriol.
<4>194
<5>6318
<6>2012
<7>Acinetobacter sp. strain GG2 is a quorum-sensing and quorum-quenching bacterium isolated from
the ginger rhizosphere. It degrades a broad range of
N-acylhomoserine lactone molecules via lactonase. The genome sequence of strain
GG2 may provide insights on the regulation of quorum-sensing and quorum-quenching
mechanisms in this bacterium.

<>

<1>Hong, K.W., Thinagaran, D.A., Gan, H.M., Yin, W.F., Chan, K.G.
<2>Whole-Genome Sequence of Cupriavidus sp. Strain BIS7, a Heavy-Metal-Resistant Bacterium.
<3>J. Bacteriol.
<4>194
<5>6324
<6>2012
<7>Cupriavidus sp. strain BIS7 is a Malaysian tropical soil bacterium that exhibits  broad
heavy-metal resistance [Co(II), Zn(II), Ni(II), Se(IV), Cu(II), chromate,
Co(III), Fe(II), and Fe(III)]. It is particularly resistant to Fe(II), Fe(III),
and Zn(II). Here we present the assembly and annotation of its genome.

<>

<1>Hong, S.H., Kim, J.S., Lee, S.Y., In, Y.H., Choi, S.S., Rih, J.K., Kim, C.H., Jeong, H., Hur, C.G., Kim, J.J.
<2>The genome sequence of the capnophilic rumen bacterium Mannheimia succiniciproducens.
<3>Nat. Biotechnol.
<4>22
<5>1275-1281
<6>2004
<7>The rumen represents the first section of a ruminant animal's stomach, where feed is
collected and mixed with microorganisms for initial
digestion. The major gas produced in the rumen is CO(2) (65.5 mol%), yet
the metabolic characteristics of capnophilic (CO(2)-loving) microorganisms
are not well understood. Here we report the 2,314,078 base pair genome
sequence of Mannheimia succiniciproducens MBEL55E, a recently isolated
capnophilic Gram-negative bacterium from bovine rumen, and analyze its
genome contents and metabolic characteristics. The metabolism of M.
succiniciproducens was found to be well adapted to the oxygen-free rumen
by using fumarate as a major electron acceptor. Genome-scale metabolic
flux analysis indicated that CO(2) is important for the carboxylation of
phosphoenolpyruvate to oxaloacetate, which is converted to succinic acid
by the reductive tricarboxylic acid cycle and menaquinone systems. This
characteristic metabolism allows highly efficient production of succinic
acid, an important four-carbon industrial chemical.

<>

<1>Hongoh, Y., Sharma, V.K., Prakash, T., Noda, S., Taylor, T.D., Kudo, T., Sakaki, Y., Toyoda, A., Hattori, M., Ohkuma, M.
<2>Complete genome of the uncultured Termite Group 1 bacteria in a single host protist cell.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>5555-5560
<6>2008
<7>Termites harbor a symbiotic gut microbial community that is responsible for their ability to
thrive on recalcitrant plant matter. The community comprises diverse microorganisms, most of
which are as yet uncultivable; the detailed symbiotic mechanism remains unclear. Here, we
present the first complete genome sequence of a termite gut symbiont-an uncultured bacterium
named Rs-D17 belonging to the candidate phylum Termite Group 1 (TG1). TG1 is a dominant group
in termite guts, found as intracellular   symbionts of various cellulolytic protists, without
any physiological information. To acquire the complete genome sequence, we collected Rs-D17
cells from only a single host protist cell to minimize their genomic  variation and performed
isothermal whole-genome amplification. This
strategy enabled us to reconstruct a circular chromosome (1,125,857 bp)
encoding 761 putative protein-coding genes. The genome additionally contains 121 pseudogenes
assigned to categories, such as cell wall biosynthesis, regulators, transporters, and defense
mechanisms. Despite its apparent reductive evolution, the ability to synthesize 15 amino acids
and various cofactors is retained, some of these genes having been duplicated. Considering
that diverse termite-gut protists harbor TG1 bacteria, we suggest that this bacterial group
plays a key role in the gut symbiotic system by stably supplying essential nitrogenous
compounds deficient in lignocelluloses to their host protists and the termites. Our results
provide a breakthrough to clarify the functions of and the interactions among the individual
members of this multilayered symbiotic complex.

<>

<1>Hongoh, Y., Sharma, V.K., Prakash, T., Noda, S., Toh, H., Taylor, T.D., Kudo, T., Sakaki, Y., Toyoda, A., Hattori, M., Ohkuma, M.
<2>Genome of an Endosymbiont Coupling N2 Fixation to Cellulolysis within Protist Cells in Termite Gut.
<3>Science
<4>322
<5>1108-1109
<6>2008
<7>Termites harbor diverse symbiotic gut microorganisms, the majority of which are as yet
uncultivable and their interrelationships unclear.  Here, we present the complete genome
sequence of the uncultured Bacteroidales endosymbiont of the cellulolytic protest
Pseudotrichonympha grassii, which accounts for 70% of the bacterial cells in the gut of the
termite Coptotermes formosanus.  Functional annotation of the chromosome (1,114,206 base
pairs) unveiled its ability to fix dinitrogen and recycle putative host nitrogen wastes for
biosynthesis of diverse amino acids and cofactors, and import glucose and xylose as energy and
carbon sources.  Thus, nitrogen fixation and cellulolysis are coupled within the protist's
cells.  This highly evolved symbiotic system probably underlies the ability of the worldwide
pest termites Coptotermes to use wood as their sole food.

<>

<1>Honma, Y., Fernandez, R.E., Maurelli, A.T.
<2>A DNA adenine methylase mutant of Shigella flexneri shows no significant attenuation of virulence.
<3>Microbiology
<4>150
<5>1073-1078
<6>2004
<7>Mutants of Salmonella defective in DNA adenine methylase (dam) have been reported to be
attenuated for virulence and to provide protective
immunity when used as vaccine strains. To determine whether these
observations could be extended to Shigella, a dam mutant of Shigella
flexneri 2a was characterized and examined for the role of dam in
pathogenesis. The Shigella dam mutant showed some unique
characteristics; however, it retained virulence in vivo as well as in
vitro. The mutant invaded cultured L2 monolayer cells as efficiently as
the wild-type parent, but its intracellular growth was suppressed up to
7 h post-invasion. Furthermore, the invading dam mutant formed smaller
plaques in cell monolayers compared to the parent strain. However, the
mutant produced keratoconjunctivitis in the Sereny test in guinea pigs
only slightly more slowly than the wild-type. While the effect of the
dam mutation on virulence was modest, the rate of spontaneous mutation
in the dam mutant was 1000-fold greater compared with the wild-type.
The virulence and high mutability displayed by the dam mutant of Sh.
flexneri suggest that a general anti-bacterial pathogen vaccine
strategy based on mutations in dam needs to be re-evaluated.

<>

<1>Honsha, K.K.
<2>Stable restriction enzyme preparation by culturing Bifidobacterium bifidum.
<3>Japanese Patent Office
<4>JP 5863386 A
<5>
<6>1981
<7>Preparation comprises culturing the bacterial strain of Bifidobacterium bifidum that produces
restriction enzyme which can recognise the base arrangement CTGCAG in two chain-type DNA and
collecting the restriction enzyme from the bacterial body. Preferably B. bifidum YIT-4005
(FERM-P3372) and YIT-4007 (FERM-P 5871) can be used. The culture is practiced at 25-45 degrees
C preferably at 36-38 degrees C at pH 5-7. Preferably 6-7. After the culture the bacterial
body is treated with lysozyme and crushed by sonication. Noncellular extract is obtained by
centrifuging the crushed product and is refined by applying known methods. The restriction
enzyme (abbr. BbiI) produced by B. bifidum is the isoschizomer of PstI. B. bifidum is not a
disease-causing germ and can be easily cultured in large quantities. Further BbiI is much
stabler than PstI and can be stored at 4C for longer than two weeks with little inactivation.

<>

<1>Hood, D.W., Deadman, M.E., Jennings, M.P., Bisercic, M., Fleischmann, R.D., Venter, J.C., Moxon, E.R.
<2>DNA repeats identify novel virulence genes in Hemophilus influenzae.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>11121-11125
<6>1996
<7>The whole genome sequence (1.83 Mbp) of Haemophilus influenzae strain Rd was searched to
identify tandem oligonucleotide repeat sequences.  Loss or gain of one or more nucleotide
repeats through a recombination-independent slippage mechanism is known to mediate phase
variation of surface molecules of pathogenic bacteria, including H. influenzae.  This
facilitates evasion of host defenses and adaptation to the varying microenvironments of the
host.  We reasoned that iterative nucleotides could identify novel genes relevant to
microbe-host interactions.  Our search of the Rd genome sequence identified 9 novel loci with
multiple (range 6-36, mean 22) tandem tetranucleotide repeats.  All were found to be located
within putative open reading frames and included homologues of hemoglobin-binding proteins of
Neisseria, a glycosyltransferase (lgtC gene product) of Neisseria, and an adhesin of Yersinia.
These tetranucleotide repeat sequences were also shown to be present in two other
epidemiologically different H. influenzae type b strains, although the number and distribution
of repeats was different.  Further characterization of the lgtC gene showed that it was
involved in phenotypic switching of a lipopolysaccharide epitope and that this variable
expression was associated with changes in the number of tetranucleotide repeats.  Mutation of
lgtC resulted in attenuated virulence of H., influenzae in an infant rate model of invasive
infection.  These data indicate the rapidity, economy, and completeness with which whole
genome sequences can be used to investigate the biology of pathogenic bacteria.

<>

<1>Hooper, C.
<2>Molecular Evolution: Sampling the New Synthesis.
<3>J. NIH Res.
<4>5
<5>66-70
<6>1993
<7>Contains a discussion of the evolution of restriction enzymes.

<>

<1>Hoose, S.A., Kladde, M.P.
<2>DNA methyltransferase probing of DNA-protein interactions.
<3>Methods Mol. Biol.
<4>338
<5>225-244
<6>2006
<7>Effective methods of probing chromatin structure without disrupting DNA-protein interactions
and associations are necessary for creating an accurate picture of chromatin and its processes
in vivo.  Expression of cytidine-5 DNA methyltransferases (C5 DMTases) in Saccharomyces
cerevisiae provides a powerful noninvasive method for asaying relative DNA accessibility in
chromatin.  DNA MTases are occluded from protein-associated DNA based on the strength and span
of the DNA-protein interation.  Ectopic regulation of C5 DMTase expression systems allows for
minimal disruption of yeast physiology.  Methylated sites are detectd by bisulfite genomic
sequencing, which leads to a positive signal corresponding to modified cytidine residues.
High-resolution C5 DMTases with dinucleotide recognition specificity are shown to provide
sufficient coverage to map interactions spanning a relatively short distance.

<>

<1>Hooton, S.P., Timms, A.R., Moreton, J., Wilson, R., Connerton, I.F.
<2>Complete Genome Sequence of Salmonella enterica Serovar Typhimurium U288.
<3>Genome Announcements
<4>1
<5>e00467-13
<6>2013
<7>Salmonella enterica serovar Typhimurium U288 has firmly established itself within the United
Kingdom pig production industry. The prevalence of this highly
pathogenic multidrug-resistant serovar at such a critical point in the food chain
is therefore of great concern. To enhance our understanding of this
microorganism, whole-genome and plasmid sequencing was performed.

<>

<1>Hopkins, B.B., Reich, N.O.
<2>Simultaneous DNA binding, bending, and base flipping - Evidence for a novel M. EcoRI methyltransferase-DNA complex.
<3>J. Biol. Chem.
<4>279
<5>37049-37060
<6>2004
<7>We measured the kinetics of DNA bending by M.EcoRI using DNA labeled at both 5'-ends and
observed changes in fluorescence resonance energy
transfer. Although known to bend its cognate DNA site, energy transfer
is decreased upon enzyme binding. This unanticipated effect is shown to
be robust because we observe the identical decrease with different dye
pairs, when the dye pairs are placed on the respective 3'-ends, the
effect is cofactor- and protein-dependent, and the effect is observed
with duplexes ranging from 14 through 17 base pairs. The same labeled
DNA shows the anticipated increased energy transfer with EcoRV
endonuclease, which also bends this sequence, and no change in energy
transfer with EcoRI endonuclease, which leaves this sequence unbent. We
interpret these results as evidence for an increased end-to-end
distance resulting from M.EcoRI binding, mediated by a mechanism novel
for DNA methyltransferases, combining DNA bending and an overall
expansion of the DNA duplex. The M.EcoRI protein sequence is poorly
accommodated into well defined classes of DNA methyltransferases, both
at the level of individual motifs and overall alignment. Interestingly,
M.EcoRI has an intercalation motif observed in the FPG DNA glycosylase
family of repair enzymes. Enzyme-dependent changes in anisotropy and
fluorescence resonance energy transfer have similar rate constants,
which are similar to the previously determined rate constant for base
flipping; thus, the three processes are nearly coincidental. Similar
fluorescence resonance energy transfer experiments following
AdoMet-dependent catalysis show that the unbending transition
determines the steady state product release kinetics.

<>

<1>Horiuchi, K.
<2>Studies on restriction endonucleases.
<3>Tanpakushitsu Kakusan Koso
<4>22
<5>981-991
<6>1977
<7>
<>

<1>Horiuchi, K., Vovis, G.F., Enea, V., Zinder, N.D.
<2>Cleavage mapping of the Escherichia coli B-specific modification sites of bacteriophage fl.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>75
<5>225
<6>1975
<7>Replicative form DNA (RF) of bacteriophage fl was cleaved into specific
fragments by two endonucleases, R.HaeII and R.HaeIII, isolated from Hemophilus
aegyptius and an endonuclease R.HindII isolated from H. influenzae.  The
fragments were ordered so as to construct a circular map of the fl genome by:
1) digesting the isolated restriction fragments with a second restriction
enzyme; and 2) testing in a transfection system for the ability of the
fragments to rescue amber mutations contained on single-strand viral DNA that
was hybridized to a particular fragment.  Both of two SB sites, genetic sites
which confer upon a DNA molecule susceptibility to the restriction-modification
system of E. coli B, of fl were located on specific restriction fragments by
transfection experiments.  In vitro modification of RF of fl and its SB mutants
and the subsequent cleavage of the DNA by the Hemophilus endonucleases showed
that the B-specific methylation occurs at, or very near, the SB site.

<>

<1>Horiuchi, K., Vovis, G.F., Enea, V., Zinder, N.D.
<2>Cleavage map of bacteriophage f1: Location of the Escherichia coli B-specific modification sites.
<3>J. Mol. Biol.
<4>95
<5>147-165
<6>1975
<7>Replicative form DNA of bacteriophage f1 was cleaved into specific fragments by
two endonucleases isolated from Hemophilus aegyptius and an endonuclease
isolated fom H. influenzae.  The fragments were ordered so as to construct a
circular map of the phage f1 genome by: (1) digesting the isolated restriction
fragments with a second restriction enzyme; and (2) testing in a transfection
system for the ability of the fragments to rescue amber mutations contained on
single-stranded viral DNA that was hybridized to a particular fragment.  The
genome of bacteriophage f1 contains two SB sites, genetic sites which confer
upon a DNA molecule susceptibility to the restriction-modification system of
Escherichia coli B.  Each of these sites was located on a specific restriction
fragment by transfection experiments.  In vitro modification of replicative
form DNA of f1 and its SB mutants by endonuclease R. EcoB and the subsequent
cleavage of the DNA by the Hemophilus endonucleases showed that the B-specific
methylation occurs within at least 200 nucleotide pairs of the SB site.

<>

<1>Horiuchi, K., Vovis, G.F., Zinder, N.D.
<2>Effect of deoxyribonucleic acid length on the adenosine triphosphatase activity of Escherichia coli restriction endonuclease B.
<3>J. Biol. Chem.
<4>249
<5>543-552
<6>1974
<7>Restriction endonuclease B possesses a DNA-dependent ATPase activity.  The ATP
hydrolysis can continue for hours after the DNA hydrolysis has stopped and is
due to a stable DNA-enzyme complex.  RF1 of bacteriophage f1 is much more
active than its full length linear form (RFIII) in stimulating the ATP
hydrolysis.  Experiments with lambda phage DNA sheared into various size
molecules indicate that the ATPase activity is a direct function of the
molecular weight of linear DNA in a range between 0.9 to 13 Mdaltons.  While
sonicated, unmodified lambda DNA molecules (0.5 Md) fail to stimulate the
ATPase activity, they do inhibit the intact DNA-stimulated ATP hydrolysis.  The
results can be interpreted as follows. (a) Endonuclease R-B recognizes DNA at
the SB sites; this recognition step is independent of DNA length.  (b) The
probability that a linear DNA molecule is cleaved by an enzyme molecule bound
to the DNA depends upon the length of the DNA: the greater the number of
nucleotide pairs the higher the probability of cleavage.  Circularization of a
small DNA duplex also increases the probability of cleavage.  One possible
explanation is that the enzyme travels along the DNA molecule before cleaving
the DNA.  If the enzyme reaches an end of the DNA molecule before cleavage
occurs, the enzyme molecule is inactivated. (c) After DNA hydrolysis has
occurred, the enzyme (or one of its components) remains on the DNA molecule and
causes the massive ATP hydrolysis.

<>

<1>Horiuchi, K., Zinder, N.
<2>Cleavage of bacteriophage f1 DNA by the restriction enzyme of Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>3220-3224
<6>1972
<7>We studied the cleavage of the replicative form DNA (RFI) of bacteriophage f1
and its SB mutants by purified restriction endonuclease of E. coli B.  The
results indicate that: (i) Circular replicative forms are broken once to yield
full-length linear molecules (RFIII).  Such linear molecules are less
susceptible than RFI to endonuclease R-B.  (ii) The genetic sites (SB sites)
that confer on the DNA susceptibility to B-restriction are not the actual sites
of cleavage.  The number of possible cleavage sites is larger than the number
of SB sites.  We conclude this because an RFIII molecule produced by
endonuclease R-B from RFI of a mutant that has only one SB site can be
circularized by denaturation and renaturation.  (iii) The SB site is not
modified when the DNA is cleaved, since an SB site can be used repeatedly by
endonuclease R-B; the RF III described in ii can be cleaved by the same enzyme
after denaturation and renaturation.

<>

<1>Horiuchi, K., Zinder, N.D.
<2>Site-specific cleavage of single-stranded DNA by a Hemophilus restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>72
<5>2555-2558
<6>1975
<7>Single-stranded viral DNA of bacteriophage f1 is cleaved into specific
fragments by endo R.HaeIII, a restriction endonuclease isolated from Hemophilus
aegyptius.  The sites of the single strand cleavage correspond to those of the
double strand cleavage.  A single-stranded DNA fragment containing only one
HaeIII site is also cleaved by this enzyme.  This observation suggests that the
reaction of single-stranded DNA cleavage does not require the formation of a
symmetrical double-stranded structure that would result from the intramolecular
base-pairing between two different HaeIII sites.  Other restriction
endonucleases may also cleave single-stranded DNA.

<>

<1>Horn, F., Schroeckh, V., Netzker, T., Guthke, R., Brakhage, A.A., Linde, J.
<2>Draft Genome Sequence of Streptomyces iranensis.
<3>Genome Announcements
<4>2
<5>e00616-14
<6>2014
<7>Streptomyces iranensis HM 35 has been shown to exhibit 72.7% DNA-DNA similarity to the
important drug rapamycin (sirolimus)-producing Streptomyces rapamycinicus
NRRL5491. Here, we report the genome sequence of HM 35, which represents a
partially overlapping repertoire of secondary metabolite gene clusters with S.
rapamycinicus, including the gene cluster for rapamycin biosynthesis.

<>

<1>Horn, G., Taubeneck, U.
<2>Comparative experiments with DNA restriction factors Escherichia coli and Shigella sonnei.
<3>Z. Allg. Mikrobiol.
<4>10
<5>103-119
<6>1970
<7>None

<>

<1>Hornby, D., Matin, M.
<2>Method for detection of truncated proteins.
<3>International Patent Office
<4>WO 0206527 A
<5>
<6>2002
<7>The present invention can be used to detect and characterize a genetic mutation in a nucleic
acid sample of interest.  The invention involves the use of a detector nucleotide sequence
encoding a polypeptide, the activity of which can be attenuated or modified when a target
sequence including a mutation is cloned into a linker region of the detector nucleotide
sequence.  In one aspect, the detector nucleotide sequence encodes a detector polypeptide
comprising two or more catalytically-essential domains separated by a linker region, where the
catalytically-essential domains can function in concert to catalyze a detectable reaction.  A
mutation in the linker region that results in the elimination of one of the
catalytically-essential domains, either by truncation or by substantially altering its amino
acid sequence, will result in a detectable activity loss, thereby signaling the presence of a
mutation.

<>

<1>Hornby, D.P.
<2>DNA Methyltransferases (EC 2.1.1.72 and EC 2.1.1.73).
<3>Methods Mol. Biol.
<4>16
<5>201-211
<6>1993
<7>DNA methyltransferases (Mtases) catalyze the transfer of the S-methyl group of
S-adenosylmethionine (SAM) to deoxycytosine (dC) or deoxyadenine (dA) bases within defined DNA
sequences. Individual enzymes are specific for one or the other base, and modify at the 6-NH2
of dA(EC 2.1.1.72) or at the N4 or 5-C position of dC (EC 2.1.1.73) depending on the
particular enzyme. The reaction is predominantly irreversible. Enzymes, such as
O6-methylguanine DNA Mtase, that participate in DNA repair processes are not discussed.

<>

<1>Hornby, D.P., Bickle, T.A.
<2>Characterization of EcoPI DNA adenine methylase.
<3>Biochem. Soc. Trans.
<4>15
<5>1039
<6>1987
<7>Three categories of restriction and modification systems have been documented in bacteria and
their phage.  Type I enzymes are complex multifunctional assemblies which catalyze DNA
methylation, DNA hydrolysis, ATP hydrolysis and DNA topoisomerization.  Type II restriction
and modification genes res and mod encode independent restriction endonuclease and DNA
methylase activities which both recognize a common DNA sequence motif.  The latter enzymes are
widely used in molecular cloning.  The most recent family to emerge is a functional hybrid
between the previous types.  Thus EcoPI, a type III enzyme produced by phage PI is a tetramer
of subunit stoichiometry (mod)2(res)2.  The methylase subunit recognizes and methylates the
sequence AGACC at the central adenine, while the endonuclease subunit hydrolyses the DNA
simultaneously some 25 base pairs downstream.  We are presently investigating the behavior of
the enzyme on gels in the presence and absence of oligonucleotide duplexes and co-factors to
elucidate the detailed mechanism of catalysis.

<>

<1>Hornby, D.P., Ford, G.C.
<2>Protein-mediated base flipping.
<3>Curr. Opin. Biotechnol.
<4>9
<5>354-358
<6>1998
<7>Since the discovery that the DNA methyltransferase M.HhaI utilizes a base flipping mechanism
to expose its target cytosine during catalysis, this phenomenon has been observed in other
nucleic acid modifying enzymes.  The crystallographic analyses of such enzyme-DNA complexes
have revealed the molecular features of extrahelical base stabilization, but have been less
informative about the flipping process itself.

<>

<1>Hornby, D.P., Muller, M., Bickle, T.A.
<2>High level expression of the EcoPI modification methylase gene and characterization of the gene product.
<3>Gene
<4>54
<5>239-245
<6>1987
<7>We have cloned the gene coding for the EcoPI modification methylase in an
expression system based on the phage lambda pL promoter and the cI857-coded
thermoinducible repressor.  We have used this system to purify the enzyme on
the 20-30 mg scale and have examined some of its enzymatic properties.  The
enzyme is a tetramer of Mr 72000 subunits and is approximately 40%
alpha-helical.  Experiments with the methyl donor, S-adenosyl methionine,
radioactively labelled in different positions indicate that a methyl group is
transferred to the enzyme during the reaction in what is most likely a covalent
bond.

<>

<1>Hornby, D.P., Whitmarsh, A., Pinarbasi, H., Kelly, S.M., Price, N.C., Shore, P., Baldwin, G.S., Waltho, J.
<2>The DNA recognition subunit of a DNA methyltransferase is predominantly a molten globule in the absence of DNA.
<3>FEBS Lett.
<4>355
<5>57-60
<6>1994
<7>Enzyme-catalysed DNA methylation provides an opportunity for the modulation of protein-DNA
recognition in biological systems. Recently we have demonstrated that the smaller of the two
subunits of the heterodimeric, cytosine-specific DNA methyltransferase, M.AquI, is largely
responsible for sequence-specific DNA recognition. Here we present evidence from a series of
NMR, fluorescence and circular dichroism spectroscopy experiments that the DNA binding subunit
of M.AquI has the characteristics of a molten globule in the absence of the catalytic
machinery. In this metastable state this subunit retains its ability to bind DNA in a
sequence-specific manner. We believe this finding offers an insight into the structural
flexibility which underpins the mechanism of action of these enzymes, and may provide a
possible biological role for molten globules in protein function.

<>

<1>Hornby, D.P.J., Matin, M.M.
<2>Cloning vectors.
<3>International Patent Office
<4>WO 0125451 A
<5>
<6>2001
<7>Disclosed are novel, one-step, non-plating, methods for isolating transformed host cells which
contain a vector incorporating a nucleic acid insert.  A typical method comprises the steps
of: (a) providing a vector which comprises (i) a gene permitting positive selection, which
gene is a cytotoxic gene (e.g. a methyltransferase such as MspI) arranged such as to be
insertionally inactivated by the incorporation of a heterologous nucleic acid insert into the
vector (ii) a gene which provides a host cell transformed with the vector with a protective
effect against a selection agent, but which does not significantly degrade the selection agent
in a medium in which the host is present (e.g. an antibiotic resistance gene such as TetA(c)
which encodes a product which does not break down the selection agent but which operates as a
membrane associated antibiotic efflux pump capable of lowering the intracellular concentration
of the selection agent while substantially maintaining the intercellular concentration
thereof); (b) cloning a heterologous nuclei acid insert into the vector; (c) transforming a
population of host cells with the vector; (d) selecting host cells transformed with the vector
in a single homogenous selection step, which step does not include growing the host cells on
an agar medium e.g. by selecting host cells transformed with the vector in a liquid
propagation medium comprising the selection agent.  Also disclosed are corresponding methods
for cloning or isolating genes.  The heterologous nucleic acid insert may be obtained by use
of nucleic acid chromatography to predetermine its size (e.g. using Transgenomic, Inc. WAVE
RTM analysis system).  Also provided are vectors adapted for use in the methods (e.g. pMTet1
vector shown in Fig 2, and SEQ ID No. 1), plus compositions, kits, and systems, and methods
and processes of use thereof.

<>

<1>Horst, J.-P., Fritz, H.-J.
<2>Counteracting the mutagenic effect of hydrolytic deamination of DNA 5-methylcytosine residues at high temperature: DNA mismatch N-glycosylase Mig.Mth of the thermophilic archaeon Methanobacterium thermoautotrophicum THF.
<3>EMBO J.
<4>15
<5>5459-5469
<6>1996
<7>Spontaneous hydrolytic deamination of DNA 5-methyl-cytosine residues gives rise to T/G
mismatches which are pre-mutagenic lesions requiring DNA repair.  For fundamental reasons, the
significance of this and other processes lowering genetic fidelity must be accentuated at
elevated temperatures, making thermophilic organisms attractive objects for studying how cells
cope with thermal noise threatening the integrity of their genetic information.  Gene mig of
Methanobacterium thermoautotrophicum THF, an anaerobic archaeon with an optimal growth
temperature of 65oC, was isolated and is product (Mig.Mth; EC3.2.2-) shown to be a
T/G-selective DNA thymine N-glycosylase with the properties required for counteracting the
mutagenic effect of hydrolytic 5-meC deamination.  The enzyme acts on T/G and U/G oppositions
with similar efficiency; G/G, A/G, T/C and U/C are minor substrates; no other opposition of
common nucleobases is attacked and no removal of U from single-stranded DNA is observed.
Substrate preferences are modulated by sequence context.  Together with the results presented
here, one example of an enzyme directed against the hydrolytic deamination damage of 5-meC is
known from each of the three phylogenetic kingdoms; entry into the repair pathway is
glycosylytic in the eukaryotic and the archaeal case, whereas the eubacterial repair starts
with an endonucleolytic DNA incision.

<>

<1>Horton, J.R., Blumenthal, R.M., Cheng, X.
<2>Restriction endonucleases: Structure of the conserved catalytic core and the role of metal ions in DNA cleavage.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin, Heidelberg
<4>14
<5>361-392
<6>2004
<7>Type II restriction endonucleases are a fascinating group of proteins.  With the REBase
database currently listing ~3500 Type II REases having nearly 240 distinct DNA sequence
specificities, they constitute one of the larger known families of enzymes.  These
DNA-cleaving enzymes combine very high catalytic efficiencies (kcat/kuncat=~1016) with
exquisite DNA sequence selectivity.  Restriction enzymes that are classified Type II cleave
specifically within or close to their recognition sites, and do not require ATP hydrolysis for
their nucleolytic activity.  DNA cleavage by these enzymes can result in DNA with either 5'
or 3' overhangs or blunt ends.

<>

<1>Horton, J.R., Bonventre, J., Cheng, X.
<2>How is modification of the DNA substrate recognized by the PvuII restriction endonuclease?
<3>Biol. Chem.
<4>379
<5>451-458
<6>1998
<7>In restriction-modification systems, cleavage of substrate sites in cellular DNA by the
restriction endonuclease is prevented by the action of a cognate methyltransferase that acts
on the same substrate sites.  The PvuII restriction endonuclease has been structurally
characterized in a complex with substrate DNA and as an apoenzyme.  We report here a
structure, determined to 1.9 A resolution by crystallography, of a complex between R.PvuII and
iodinated DNA.  The presence of an iodine at the 5-carbon of the methylatable cytosine results
in the following changes in the protein: His84 moved away from the modified base; this
movement was amplified in His85 and disrupts an intersubunit hydrogen bond; and the base
modification disturbs the distribution of water molecules that associate with these histidine
residues and the area of the scissile bond.  Considering these observations, hypotheses are
given as to why a similar oligonucleotide, where a methyl group resides on the 5-carbon of the
methylatable cytosine, is slowly cleaved by R.PvuII.

<>

<1>Horton, J.R., Borgaro, J.G., Griggs, R.M., Quimby, A., Guan, S., Zhang, X., Wilson, G.G., Zheng, Y., Zhu, Z., Cheng, X.
<2>Structure of 5-hydroxymethylcytosine-specific restriction enzyme, AbaSI, in complex with DNA.
<3>Nucleic Acids Res.
<4>42
<5>7947-7959
<6>2014
<7>AbaSI, a member of the PvuRts1I-family of modification-dependent restriction endonucleases,
cleaves deoxyribonucleic acid (DNA) containing
5-hydroxymethylctosine (5hmC) and glucosylated 5hmC (g5hmC), but not DNA
containing unmodified cytosine. AbaSI has been used as a tool for mapping the
genomic locations of 5hmC, an important epigenetic modification in the DNA of
higher organisms. Here we report the crystal structures of AbaSI in the presence
and absence of DNA. These structures provide considerable, although incomplete,
insight into how this enzyme acts. AbaSI appears to be mainly a homodimer in
solution, but interacts with DNA in our structures as a homotetramer. Each AbaSI
subunit comprises an N-terminal, Vsr-like, cleavage domain containing a single
catalytic site, and a C-terminal, SRA-like, 5hmC-binding domain. Two N-terminal
helices mediate most of the homodimer interface. Dimerization brings together the
two catalytic sites required for double-strand cleavage, and separates the 5hmC
binding-domains by approximately 70 A, consistent with the known activity of
AbaSI which cleaves DNA optimally between symmetrically modified cytosines
approximately 22 bp apart. The eukaryotic SET and RING-associated (SRA) domains
bind to DNA containing 5-methylcytosine (5mC) in the hemi-methylated CpG
sequence. They make contacts in both the major and minor DNA grooves, and flip
the modified cytosine out of the helix into a conserved binding pocket. In
contrast, the SRA-like domain of AbaSI, which has no sequence specificity,
contacts only the minor DNA groove, and in our current structures the 5hmC
remains intra-helical. A conserved, binding pocket is nevertheless present in
this domain, suitable for accommodating 5hmC and g5hmC. We consider it likely,
therefore, that base-flipping is part of the recognition and cleavage mechanism
of AbaSI, but that our structures represent an earlier, pre-flipped stage, prior
to actual recognition.

<>

<1>Horton, J.R., Cheng, X.
<2>PvuII endonuclease contains two calcium ions in active sites.
<3>J. Mol. Biol.
<4>300
<5>1049-1056
<6>2000
<7>Restriction endonucleases differ in their use of metal cofactors despite having remarkably
similar folds for their catalytic regions. To explore this, we have characterized the
interaction of endonuclease PvuII with the catalytically incompetent cation Ca(2+). The
structure of a glutaraldehyde-crosslinked crystal of the endonuclease PvuII-DNA complex,
determined in the presence of Ca(2+) at a pH of approximately 6.5, supports a two-metal
mechanism of DNA cleavage by PvuII. The first Ca(2+) position matches that found in all
structurally examined endonucleases, while the second position is similar to that of EcoRV but
is distinct from that of BamHI and BglI. The location of the second metal in PvuII, unlike
that in BamHI/BglI, permits no direct interaction between the second metal and the O3' oxygen
leaving group. However, the interactions between the DNA scissile phosphate and the metals,
the first metal and the attacking water, and the attacking water and DNA are the same in PvuII
as they are in the two-metal models of BamHI and BglI, but are distinct from the proposed
three-metal or the two-metal models of EcoRV.

<>

<1>Horton, J.R., Liebert, K., Bekes, M., Jeltsch, A., Cheng, X.D.
<2>Structure and Substrate Recognition of the Escherichia coli DNA Adenine Methyltransferase.
<3>J. Mol. Biol.
<4>358
<5>559-570
<6>2006
<7>The structure of the Escherichia coli Dam DNA-(adenine-N6)-methyltransferase in complex with
cognate DNA was determined at 1.89 (A) over circle resolution in the presence of
S-adenosyl-L-homocysteine. DNA recognition and the dynamics of base-flipping were studied by
site-directed mutagenesis, DNA methylation kinetics and fluorescence stopped-flow experiments.
Our data illustrate the mechanism of coupling of DNA recognition and base-flipping. Contacts
to the non-target strand in the second (3') half of the GATC site are established by R124 to
the fourth base-pair, and by L122 and P134 to the third base-pair. The aromatic ring of Y119
intercalates into the DNA between the second and third base-pairs, which is essential for
base-flipping to occur. Compared to previous published structures of bacteriophage T4 Dam,
three major new observations are made in E. coli Dam. (1) The first Gua is recognized by K9,
removal of which abrogates the first base-pair recognition. (2) The flipped target Ade binds
to the surface of EcoDam in the absence of S-adenoSyl-L-methionine, which illustrates a
possible intermediate in the base-flipping pathway. (3) The orphaned Thy residue displays
structural flexibility by adopting an extrahelical or intrahelical position where it is in
contact to N120.

<>

<1>Horton, J.R., Liebert, K., Hattman, S., Jeltsch, A., Cheng, X.
<2>Transition from nonspecific to specific DNA interactions along the substrate-recognition pathway of Dam methyltransferase.
<3>Cell
<4>121
<5>349-361
<6>2005
<7>DNA methyltransferases methylate target bases within specific nucleotide sequences. Three
structures are described for bacteriophage T4 DNA-adenine methyltransferase (T4Dam) in ternary
complexes with partially and fully specific DNA and a methyl-donor analog. We also report the
effects of substitutions in the related Escherichia coli DNA methyltransferase (EcoDam),
altering residues corresponding to those involved in specific interaction with the canonical
GATC target sequence in T4Dam. We have identified two types of protein-DNA interactions:
discriminatory contacts, which stabilize the transition state and accelerate methylation of
the cognate site, and antidiscriminatory contacts, which do not significantly affect
methylation of the cognate site but disfavor activity at noncognate sites. These structures
illustrate the transition in enzyme-DNA interaction from nonspecific to specific interaction,
suggesting that there is a temporal order for formation of specific contacts.

<>

<1>Horton, J.R., Mabuchi, M.Y., Cohen-Karni, D., Zhang, X., Griggs, R.M., Samaranayake, M., Roberts, R.J., Zheng, Y., Cheng, X.
<2>Structure and cleavage activity of the tetrameric MspJI DNA modification-dependent restriction endonuclease.
<3>Nucleic Acids Res.
<4>40
<5>9763-9773
<6>2012
<7>The MspJI modification-dependent restriction endonuclease recognizes 5-methylcytosine or
5-hydroxymethylcytosine in the context of CNN(G/A) and cleaves both strands at fixed distances
(N(12)/N(16)) away from the modified cytosine at the 3'-side. We determined the crystal
structure of MspJI of Mycobacterium sp. JLS at 2.05-A resolution. Each protein monomer harbors
two domains: an N-terminal DNA-binding domain and a C-terminal endonuclease. The N-terminal
domain is structurally similar to that of the eukaryotic SET and RING-associated domain, which
is known to bind to a hemi-methylated CpG dinucleotide. Four protein monomers are found in the
crystallographic asymmetric unit. Analytical gel-filtration and ultracentrifugation
measurements confirm that the protein exists as a tetramer in solution. Two monomers form a
back-to-back dimer mediated by their C-terminal endonuclease domains. Two back-to-back dimers
interact to generate a tetramer with two double-stranded DNA cleavage modules. Each cleavage
module contains two active sites facing each other, enabling double-strand DNA cuts.
Biochemical, mutagenesis and structural characterization suggest three different monomers of
the tetramer may be involved respectively in binding the modified cytosine, making the first
proximal N(12) cleavage in the same strand and then the second distal N(16) cleavage in the
opposite strand. Both cleavage events require binding of at least a second recognition site
either in cis or in trans.

<>

<1>Horton, J.R., Nastri, H.G., Riggs, P.D., Cheng, X.
<2>Asp34 of PvuII endonuclease is directly involved in DNA minor groove recognition and indirectly involved in catalysis.
<3>J. Mol. Biol.
<4>284
<5>1491-1504
<6>1998
<7>The PvuII restriction endonuclease is a homodimer that recognizes and cleaves the DNA sequence
5'-CAGCTG-3' in double-stranded DNA, and the structure of this enzyme has been reported.  In
the wild-type enzyme, Asp34 interacts with the internal guanine of the recognition sequence on
the minor groove side.  The Asp34 codon was altered to specify Gly (D34G), and in vitro
studies have revealed that the D34G protein has lost binding specificity for the central G.C
base-pairs, and that it cuts the canonical sequence with 10^-4-fold reduced activity as
compared to the wild-type enzyme.  We have now determined the structure at 1.59 angstroms
resolution of the D34G PvuII endonuclease complexed with a 12 bp duplex deoxyoligonucleotide
containing the cognate sequence.  The D34G alteration results in several structural changes
relative to wild-type protein/DNA complexes.  First, the sugar moiety of the internal guanine
changes from a C2'-endo to C3'-endo pucker while that of the 3' guanine changes from
C3'-endo to C2'-endo pucker.  Second, the axial rise between the internal G.C base-pairs is
reduced while that between the G.C and flanking base-pairs is expanded.  Third, two distinct
monomeric active sites are observed that we refer to as being "primed" and "unprimed" for
phosphodiester bond cleavage.  The primed and unprimed sites differ in the conformation of the
Asp58 side-chain, and in the absence from unprimed sites of four networked water molecules.
These water molecules, present in the primed site, have been implicated in the catalytic
mechanism of this and other endonucleases; some of them can be replaced by the Mg2+ necessary
for cleavage.  Taken together, these structural changes imply that the Asp34 side-chains from
the two subunits maintain a distinct conformation of its DNA substrate, properly situating the
target backbone phosphates and indirectly manipulating the active sites.  This provides some
insight into how recognition of the specific DNA sequence is linked to catalysis by the highly
specific restriction endonucleases, and reveals one way in which the structural conformation
of the DNA is modulated coordinately with that of the PvuII protein.

<>

<1>Horton, J.R., Nugent, R.L., Li, A., Mabuchi, M.Y., Fomenkov, A., Cohen-Karni, D., Griggs, R.M., Zhang, X., Wilson, G.G., Zheng, Yu., Xu, S.-Y., Cheng, X.
<2>Structure and mutagenesis of the DNA modification-dependent restriction endonuclease AspBHI.
<3>Sci. Rep.
<4>4
<5>9
<6>2014
<7>The modification-dependent restriction endonuclease AspBHI recognizes 5-methylcytosine (5mC)
in the double-strand DNA sequence context of (C/T)(C/G)(5mC) N(C/G) (N = any nucleotide) and
cleaves the two strands a fixed distance (N-12/N-16) 3' to the modified cytosine. We
determined the crystal structure of the homo-tetrameric AspBHI. Each subunit of the protein
comprises two domains: an N-terminal DNA-recognition domain and a C-terminal DNA cleavage
domain. The N-terminal domain is structurally similar to the eukaryotic SET and
RING-associated (SRA) domain, which is known to bind to a hemi-methylated CpG dinucleotide.
The C-terminal domain is structurally similar to classic Type II restriction enzymes and
contains the endonuclease catalytic-site motif of DX(20)EAK. To understand how specific amino
acids affect AspBHI recognition preference, we generated a homology model of the AspBHI-DNA
complex, and probed the importance of individual amino acids by mutagenesis. Ser41 and Arg42
are predicted to be located in the DNA minor groove 5' to the modified cytosine. Substitution
of Ser41 with alanine (S41A) and cysteine (S41C) resulted in mutants with altered cleavage
activity. All 19 Arg42 variants resulted in loss of endonuclease activity.

<>

<1>Horton, J.R., Ratner, G., Banavali, N.K., Huang, N., Choi, Y., Maier, M.A., Marquez, V.E., MacKerell, A.D. Jr., Cheng, X.
<2>Caught in the act: visualization of an intermediate in the DNA base-flipping pathway induced by HhaI methyltransferase.
<3>Nucleic Acids Res.
<4>32
<5>3877-3886
<6>2004
<7>Rotation of a DNA or RNA nucleotide out of the double helix and
into a protein pocket ('base flipping') is a mechanistic feature common to
some DNA/RNA-binding proteins. Here, we report the structure of HhaI
methyltransferase in complex with DNA containing a south-constrained abasic
carbocyclic sugar at the target site in the presence of the methyl donor
byproduct AdoHcy. Unexpectedly, the locked south pseudosugar appears to be
trapped in the middle of the flipping pathway via the DNA major groove,
held in place primarily through Van der Waals contacts with a set of
invariant amino acids. Molecular dynamics simulations indicate that the
structural stabilization observed with the south-constrained pseudosugar
will not occur with a north-constrained pseudosugar, which explains its
lowered binding affinity. Moreover, comparison of structural transitions of
the sugar and phosphodiester backbone observed during computational studies
of base flipping in the M.HhaIM-DNA-AdoHcy ternary  complex indicate that the
south-constrained pseudosugar induces a conformation on the phosphodiester
backbone that corresponds to that of a discrete intermediate of the
base-flipping pathway. As previous crystal structures of M.HhaI ternary
complex with DNA displayed the flipped sugar moiety in the antipodal north
conformation, we suggest that conversion of the sugar pucker from south to
north beyond the middle of the pathway is an essential part of the
mechanism through which flipping must proceed to reach its final
destination. We also discuss the possibility of the south-constrained
pseudosugar mimicking a transition state in the phosphodiester and sugar
moieties that occurs during DNA base flipping in the presence of M.HhaI.

<>

<1>Horton, J.R., Wang, H., Mabuchi, M.Y., Zhang, X., Roberts, R.J., Zheng, Y., Wilson, G.G., Cheng, X.
<2>Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix.
<3>Nucleic Acids Res.
<4>42
<5>12092-12101
<6>2014
<7>MspJI belongs to a family of restriction enzymes that cleave DNA containing 5-methylcytosine
(5mC) or 5-hydroxymethylcytosine (5hmC). MspJI is specific for the sequence 5(h)mC-N-N-G or A
and cleaves with some variability 9/13 nucleotides downstream. Earlier, we reported the
crystal structure of MspJI without DNA and proposed how it might recognize this sequence and
catalyze cleavage. Here we report its co-crystal structure with a 27-base pair oligonucleotide
containing 5mC. This structure confirms that MspJI acts as a homotetramer and that the
modified cytosine is flipped from the DNA helix into an SRA-like-binding pocket. We expected
the structure to reveal two DNA molecules bound specifically to the tetramer and engaged with
the enzyme's two DNA-cleavage sites. A coincidence of crystal packing precluded this
organization, however. We found that each DNA molecule interacted with two adjacent tetramers,
binding one specifically and the other non-specifically. The latter interaction, which
prevented cleavage-site engagement, also involved base flipping and might represent the
sequence-interrogation phase that precedes specific recognition. MspJI is unusual in that DNA
molecules are recognized and cleaved by different subunits. Such interchange of function might
explain how other complex multimeric restriction enzymes act.

<>

<1>Horton, J.R., Zhang, X., Maunus, R., Yang, Z., Wilson, G.G., Roberts, R.J., Cheng, X.
<2>DNA nicking by HinP1I endonuclease: bending, base flipping and minor groove expansion.
<3>Nucleic Acids Res.
<4>34
<5>939-948
<6>2006
<7>HinP1I recognizes and cleaves the palindromic tetranucleotide sequence G downward arrowCGC in
DNA. We report three structures of HinP1I-DNA
complexes: in the presence of Ca(2+) (pre-reactive complex), in the
absence of metal ion (binary complex) and in the presence of Mg(2+)
(post-reactive complex). HinP1I forms a back-to-back dimer with two active
sites and two DNA duplexes bound on the outer surfaces of the dimer facing
away from each other. The 10 bp DNA duplexes undergo protein-induced
distortions exhibiting features of A-, B- and Z-conformations: bending on
one side (by intercalation of a phenylalanine side chain into the major
groove), base flipping on the other side of the recognition site (by
expanding the step rise distance of the local base pair to Z-form) and a
local A-form conformation between the two central C:G base pairs of the
recognition site (by binding of the N-terminal helix in the minor groove).
In the pre- and post-reactive complexes, two metals (Ca(2+) or Mg(2+)) are
found in the active site. The enzyme appears to cleave DNA sequentially,
hydrolyzing first one DNA strand, as seen in the post-reactive complex in
the crystalline state, and then the other, as supported by the observation
that, in solution, a nicked DNA intermediate accumulates before
linearization.

<>

<1>Horton, N.C., Connolly, B.A., Perona, J.J.
<2>Inhibition of EcoRV endonuclease by deoxyribo-3'-S-phosphorothiolates: A high-resolution X-ray crystallographic study.
<3>J. Am. Chem. Soc.
<4>122
<5>3314-3324
<6>2000
<7>Three high-resolution structures of the restriction endonuclease EcoRV bound to a duplex DNA
substrate analogue with deoxyribo-3'-S-phosphorothiolate linkages at both scissile phosphates
are presented.  In each of these structures cocrystallized with Mg2+, Mn2+, or Ca2+ ions, the
nonesterified pro-S oxygen of the scissile phosphate no longer directly ligates a divalent
cation, as is observed for the unmodified complex.  Instead, one metal ion in all three
structures is shifted toward the adjacent 3'-phosphate of the DNA, to occupy a position
nearly identical to that previously observed in an EcoRV T93A/DNA/Ca2+ complex (N.C. Horton et
al., Proc. Natl. Acad. Sci. U.S.A. 1998, 95, 13489).  A second divalent metal ion in each
structure bridges the carboxylate groups of Asp74 and Glu45 (74/45 site), as also seen in both
wild-type and T93A cocrystals.  The uncleaved 3'-S-phosphorothiolate DNAs in these complexes
are only slightly distorted from the conformation of the unmodified duplex.  Kinetic
measurements show that the rate of the chemical step for analogue cleavage is severely reduced
for each of the active metals Mg2+, Mn2+, and Co2+, and that the thiophilic Mn2+, Cd2+, and
Zn2+ cations do not provide a measurable reconstitution of activity.  The inability of
thiophilic metals to improve activity is consistent with models for catalysis derived from
previous crystal structures, which indicate that ligation of a metal ion to the 3'-oxygen is
mediated through an inner-sphere water molecule rather than by direct interaction.  The
structures suggest that 3'-S-phosphorothiolate analogues resist cleavage because the bridging
sulfur excludes inner-sphere ligation of divalent metal ions to any position on the scissile
phosphate.  This distinguishes the inhibitory mechanism in EcoRV from that operative in the
3'-5' exonuclease active site of DNA polymerase I (C.A. Brautigam et al., Biochemistry,
1999, 38, 696), and likely as well from other enzymes which also catalyze phosphoryl transfer
via direct metal ligation to the 3'-oxygen leaving group.

<>

<1>Horton, N.C., Dorner, L.F., Perona, J.J.
<2>Sequence selectivity and degeneracy of a restriction endonuclease mediated by DNA intercalation.
<3>Nat. Struct. Biol.
<4>9
<5>42-47
<6>2002
<7>The crystal structure of the HincII restriction endonuclease-DNA complex shows that degenerate
specificity for blunt-ended cleavage at GTPyPuAC sequences arises from indirect readout of
conformational preferences at the center pyrimidine-purine step. Protein-induced distortion of
the DNA is accomplished by intercalation of glutamine side chains into the major groove on
either side of the recognition site, generating bending by either tilt or roll at three
distinct loci. The intercalated side chains propagate a concerted shift of all six target-site
base pairs toward the minor groove, producing an unusual cross-strand purine stacking at the
center pyrimidine-purine step. Comparison of the HincII and EcoRV cocrystal structures
suggests that sequence-dependent differences in base-stacking free energies are a crucial
underlying factor mediating protein recognition by indirect readout.

<>

<1>Horton, N.C., Dorner, L.F., Schildkraut, I., Perona, J.J.
<2>Crystallization and preliminary diffraction analysis of the HincII restriction endonuclease-DNA complex.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>55
<5>1943-1945
<6>1999
<7>Crystals of the 60 kDa dimeric HincII restriction enzyme bound to a 12 base-pair
dyad-symmetric duplex DNA carrying the specific 5'-GTCGAC recognition site have been
obtained. Crystals grew by hanging-drop vapor diffusion from solutions containing polyethylene
glycol 4000 as precipitating agent. The rod-shaped crystals belong to space group I222 (or
I2(1)2(1)2(1)), with unit-cell dimensions a = 66.9, b = 176.7, c = 256.0 A. There are most
likely to be two dimeric complexes in the asymmetric unit. A complete native data set has been
collected from a high-energy synchrotron source to a resolution of 2.5 A at 100 K, with an
R(merge) of 4.8%.

<>

<1>Horton, N.C., Newberry, K.J., Perona, J.J.
<2>Metal ion-mediated substrate-assisted catalysis in type II restriction endonucleases.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>13489-13494
<6>1998
<7>The 2.15-Angstrom resolution cocrystal structure of EcoRV endonuclease mutant T93A complexed
with DNA and Ca2+ ions reveals two divalent metals bound in one of the active sites.  One of
these metals is ligated through an inner-sphere water molecule to the phosphate group located
3' to the scissile phosphate.  A second inner-sphere water on this metal is positioned
approximately in-line for attack on the scissile phosphate.  This structure corroborates the
observation that the pro-SP phosphoryl oxygen on the adjacent 3' phosphate cannot be modified
without severe loss of catalytic efficiency.  The structural equivalence of key groups,
conserved in the active sites of EcoRV, EcoRI, PvuII, and BamHI endonucleases, suggests that
ligation of a catalytic divalent metal ion to this phosphate may occur in many type II
restriction enzymes.  Together with previous  cocrystal structures, these data allow
construction of a detailed model for the pretransition state configuration in EcoRV.  This
model features three divalent metal ions per active site and invokes assistance in the
bond-making step by a conserved lysine, which stabilizes the attacking hydroxide ion
nucleophile.

<>

<1>Horton, N.C., Otey, C., Lusetti, S., Sam, M.D., Kohn, J., Martin, A.M., Ananthnarayan, V., Perona, J.J.
<2>Electrostatic contributions to site specific DNA cleavage by EcoRV endonuclease.
<3>Biochemistry
<4>41
<5>10754-10763
<6>2002
<7>Mutational analysis of amino acids at the periphery of the EcoRV endonuclease active site
suggests that moderate-range electrostatic effects play a significant role in modulating the
efficiency of phosphoryl transfer. Asp36 and Lys38 located on minor-groove binding surface
loops approach within 7-9 angstroms of the scissile phosphates of the DNA. While the rates of
single-site mutations removing the carboxylate or amine moieties at these positions are
decreased 10^3-10^5-fold compared to that of wild-type EcoRV, we find that double mutants
which rebalance the charge improve catalysis by up to 500-fold. Mutational analysis also
suggests that catalytic efficiency is influenced by Lys173, which is buried at the base of a
deep depression penetrating from a distal surface of the enzyme. The Lys173 amine group lies
just 6 angstroms from the amine group of the conserved essential Lys92 side chain in the
active site. Kinetic and crystallographic analyses of the EcoRV E45A mutant enzyme further
show that the Glu45 carboxylate group facilitates an extensive set of conformational
transitions which occur upon DNA binding. The crystal structure of E45A bound to DNA and Mn2+
ions reveals significant conformational alterations in a small alpha-helical portion of the
dimer interface located adjacent to the DNA minor groove. This leads to a tertiary
reorientation of the two monomers as well as shifting of the key major-groove binding
recognition loops. Because the Glu45 side chain does not appear to play a direct structural
role in maintaining the active site, these rearrangements may instead originate in an altered
electrostatic potential caused by removal of the negative charge. A Mn2+ binding site on the
scissile phosphate is also disrupted in the E45A structure such that inner-sphere metal
interactions made by the scissile DNA phosphate and conserved Asp90 carboxylate are each
replaced with water molecules in the mutant. These findings argue against a proposed role for
Asp36 as the general base in EcoRV catalysis, and reveal that the induced-fit conformational
changes necessary for active site assembly and metal binding are significantly modulated by
the electrostatic potential in this region.

<>

<1>Horton, N.C., Perona, J.J.
<2>Role of protein-induced bending in the specificity of DNA recognition: crystal structure of EcoRV endonuclease complexes with d(AAAGAT) + d(ATCTT).
<3>J. Mol. Biol.
<4>277
<5>779-787
<6>1998
<7>The crystal structure of EcoRV endonuclease has been determined at 2.1 A resolution complexed
to two five-base-pair DNA duplexes each containing the cognate recognition half-site.  The
highly localized 50o bend into the major groove seen at the center TA-step of the continuous
GATATC site is preserved in this discontinuous DNA complex lacking the scissile phosphates.
Thus, this crystal structure provides evidence that covalent constraints associated with a
continuous target site are not essential to enzyme-induced DNA bending, even when these
constraints are removed directly at the locus of the bend.  The scissile phosphates are also
absent in the crystal structure of EcoRV bound to the non-specific site TCGCGA, which shows a
straight B-like conformation.  We conclude that DNA bending by EcoRV is governed only by the
sequence and is not influenced by the continuity of the phosphodiester backbone.  Together
with other data showing that cleavable non-cognate sites different by one or two base-pairs
from GATATC, but does not bend non-specific sites that are less similar.  Structural and
thermodynamic considerations suggest that the sequence-dependent energy cost of DNA bending is
likely to play an important role in determining the specificity of EcoRV.  This differential
cost is manifested at the binding step for bent non-cognate sequences and at the catalytic
step for unbent non-specific sequences.

<>

<1>Horton, N.C., Perona, J.J.
<2>Recognition of flanking DNA sequences by EcoRV endonuclease involves alternative patterns of water-mediated contacts.
<3>J. Biol. Chem.
<4>273
<5>21721-21729
<6>1998
<7>The 2.1-A cocrystal structure of EcoRV endonuclease bound to 5'-CGGGATATCCC, in a crystal
lattice isomorphous with the cocrystallized undecamer 5'-AAAGATATCTT previously determined,
shows novel base recognition in the major groove of the DNA flanking the GATATC target site.
Lys104 of the enzyme interacts through water molecules with the exocyclic N-4 amino groups of
flanking cytosines.  Steric exclusion of water molecule-binding sites by the 5'methyl group
of thymine drives the adoption of alternative water-mediated contacts with AT versus GC
flanks.  This structure provides a rare example of structural adaptability in the recognition
of different DNA sequences by a protein and suggests preferred strategies for the expansion of
target site specificity by EcoRV.

<>

<1>Horton, N.C., Perona, J.J.
<2>Crystallographic snapshots along a protein-induced DNA-bending pathway.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>5729-5734
<6>2000
<7>Two new high-resolution cocrystal structures of EcoRV endonuclease bound to DNA show that a
large variation in DNA-bending angles is sampled in the ground state binary complex. Together
with previous structures, these data reveal a contiguous series of protein conformational
states delineating a specific trajectory for the induced-fit pathway. Rotation of the
DNA-binding domains, together with movements of two symmetry-related helices binding in the
minor groove, causes base unstacking at a key base-pair step and propagates structural changes
that assemble the active sites. These structures suggest a complex mechanism for DNA bending
that depends on forces generated by interacting protein segments, and on selective
neutralization of phosphate charges along the inner face of the bent double helix.

<>

<1>Horton, N.C., Perona, J.J.
<2>Making the most of metal ions.
<3>Nat. Struct. Biol.
<4>8
<5>290-293
<6>2001
<7>Crystal structures of the homing endonuclease I-Crel bound to substrate DNA and divalent
metals show that one metal ion is shared between the
two active sites of the enzyme. This arrangement appears uniquely
suited to the formation of double-stranded DNA breaks via a concerted
reaction.

<>

<1>Horton, N.C., Perona, J.J.
<2>DNA cleavage by EcoRV endonuclease: Two metal ions in three metal ion binding sites.
<3>Biochemistry
<4>43
<5>6841-6857
<6>2004
<7>Four crystal structures of EcoRV endonuclease mutants K92A and K38A provide new insight into
the mechanism of DNA bending and the structural basis for metal-dependent phosphodiester bond
cleavage. The removal of a key active site positive charge in the uncleaved K92A-DNA-M(2+)
substrate complex results in binding of a sodium ion in the position of the amine nitrogen,
suggesting a key role for a positive charge at this position in stabilizing the sharp DNA bend
prior to cleavage. By contrast, two structures of K38A cocrystallized with DNA and Mn(2+) ions
in different lattice environments reveal cleaved product complexes featuring a common, novel
conformation of the scissile phosphate group as compared to all previous EcoRV structures. In
these structures, the released 5'-phosphate and 3'-OH groups remain in close juxtaposition
with each other and with two Mn(2+) ions that bridge the conserved active site carboxylates.
The scissile phosphates are found midway between their positions in the prereactive substrate
and postreactive product complexes of the wild-type enzyme. Mn(2+) ions occupy two of the
three sites previously described in the prereactive complexes and are plausibly positioned to
generate the nucleophilic hydroxide ion, to compensate for the incipient additional negative
charge in the transition state, and to ionize a second water for protonation of the
3'-oxyanion. Reconciliation of these findings with earlier X-ray and fluorescence studies
suggests a novel mechanism in which a single initially bound metal ion in a third distinct
site undergoes a shift in position together with movement of the scissile phosphate deeper
into the active site cleft. This reconfigures the local environment to permit binding of the
second metal ion followed by movement toward the pentacovalent transition state. The new
mechanism suggested here embodies key features of previously proposed two- and three-metal
catalytic models, and offers a view of the stereochemical pathway that integrates much of the
copious structural and functional data that are available from exhaustive studies in many
laboratories.

<>

<1>Horton, N.C., Sam, M.D., Connolly, B.A., Perona, J.J.
<2>Structural mechanism of phosphoryl transfer in EcoRV restriction endonuclease.
<3>Biophys. J.
<4>78
<5>417A
<6>2000
<7>The structural mechanism of phosphoryl transfer in the type II restriction endonuclease EcoRV
has been investigated using thermodynamic, kinetic, and crystallographic methods on modified
substrates and site directed mutants.  A three metal ion mechanism has been proposed based on
the observation of divalent cation binding sites in the enzyme active site.  pH rate profiles
using Mg2+ and Mn2+ as the catalytic cofactor support the role of the metal ion in increasing
the electrophilicity of the scissile phosphorus by direct ligation to the proS oxygen of the
scissile phosphate.  Thio substitution at the 3'O position, in conjunction with metal
substitution experiments, support the lack of direct metal ligation at this position.
Structures of 3'S phosphorothiolate DNA containing the EcoRV cognate sequence, bound to
EcoRV, and the divalent cations Ca2+, Mg2+ and Mn2+ show loss of direct ligation by the metal
ion to the proS oxygen of the scissile phosphate, explaining the inability of EcoRV to cleave
this modified substrate.  A structure of the mutant K38A bound to cognate DNA nd Mn2+ shows a
new conformation of the scissile phosphate which is pulled deep into the active site, and
suggests the conformation of an intermediate in the phosphoryl transfer reaction pathway.

<>

<1>Horvath, B., Hunyadkurti, J., Voros, A., Fekete, C., Urban, E., Kemeny, L., Nagy, I.
<2>Genome sequence of Propionibacterium acnes Type II strain ATCC 11828.
<3>J. Bacteriol.
<4>194
<5>202-203
<6>2011
<7>Propionibacterium acnes is an anaerobic Gram-positive bacterium that forms part of the normal
human cutaneous microbiota and is occasionally associated with inflammatory diseases (I.
Kurokawa et al., Exp. Dermatol. 18:821- 832, 2009). Here we present the complete genome
sequence for the commercially available P. acnes type II reference strain ATCC 11828 (I. Nagy
et al., Microbes Infect. 8:2195-2205, 2006) recovered from a subcutaneous abscess.

<>

<1>Horvath, P., Barrangou, R.
<2>Protection against Foreign DNA.
<3>Bacterial Stress Responses, 2nd Edition, Amer. Soc. Microbiology, Storz, G., Hengge, R., Washington, D.C.
<4>0
<5>333-348
<6>2011
<7>Bacteria rely on several defense systems that allow them to survive exposure to invading
nucleic acids. Predatory exposure to abundant and ubiquitous viruses, combined with
competition from a vast array of microbes, has lead to exogenous DNA exposure via
transduction, conjugation, and transformation. Consequently, bacterial immune systems have
been developed that allow the cell to recognize and distinguish incoming exogenous 'foreign'
DNA, from endogenous 'self' DNA. These systems maintain genetic integrity, species identity,
and genetic uniqueness, yet allow occasional exogenous DNA uptake and conservation of
advantageous genetic material for adaptation to the environment. In addition to defense
strategies such as prevention of adsorption, blocking of injection, and abortive infection,
which are effective against phage, the chapter briefly addresses the well-characterized
restriction-modification system (R-M), non-sugar-specific nucleases, and histone-like nucleoid
structuring (H-NS). The chapter more specifically elaborates on clustered regularly
interspaced short palindromic repeats (CRISPR). CRISPR/Cas, a recently described microbial
system, provides acquired immunity against phages and plasmids by targeting nucleic acids in a
sequence-specific manner. CRISPR features may be exploited for typing purposes, ecological and
epidemiological studies, and also for enhancing phage resistance in bacteria.

<>

<1>Hoshino, T., Uozumi, T., Horinouchi, S., Ozaki, A., Beppu, T., Arima, K.
<2>Purification of cohesive-end-producing restriction endonuclease from Bacillus subtilis G.
<3>Biochim. Biophys. Acta
<4>479
<5>367-369
<6>1977
<7>A new restriction endonuclease was partially purified from Bacillus subtilis G
(IAM1247).  This restriction endonuclease (endonuclease RBsuG) seems to produce
cohesive ends at its cleavage site.

<>

<1>Hoskins, I.C., Lax, A.J.
<2>Identification of restriction barriers in Pasteurella multocida.
<3>FEMS Microbiol. Lett.
<4>156
<5>223-226
<6>1997
<7>Several naturally occurring antibiotic resistance plasmids were isolated from Pasteurella
multocida type D strains.  One plasmid, pPM1, was used to study transfer of DNA among P.
multocida strains, and could be transferred into Escherichia coli and some P. multocida
isolates.  However, pPM1 could only be transferred into the toxigenic P. multocida LFB3 at
very low frequency.  Plasmid recovered from the electrotransformants could be transferred to
LFB3 at high frequency.  These plasmid DNAs were resistant to PstI, and sensitive to DpnI
digestion.  Sensitivity to DpnI was common to all the P. multocida DNAs, but resistance to
PstI was confined to LFB3.  Plasmid pPM1 treated with PstI methylase was able to transform
LFB3 at an increased frequency compared to unmethylated DNA, suggesting that LFB3 has a
restriction system which cleaves at or near PstI sites.

<>

<1>Hoskins, J. et al.
<2>Genome of the bacterium Streptococcus pneumoniae strain R6.
<3>J. Bacteriol.
<4>183
<5>5709-5717
<6>2001
<7>Streptococcus pneumoniae is among the most significant causes of bacterial disease in humans.
Here we report the 2,038,615-bp genomic sequence of the gram-positive bacterium S. pneumoniae
R6. Because the R6 strain is avirulent and, more importantly, because it is readily
transformed with DNA from homologous species and many heterologous species, it is the
principal platform for investigation of the biology of this important pathogen. It is also
used as a primary vehicle for genomics-based development of antibiotics for gram-positive
bacteria. In our analysis of the genome, we identified a large number of new uncharacterized
genes predicted to encode proteins that either reside on the surface of the cell or are
secreted. Among those proteins there may be new targets for vaccine and antibiotic
development.

<>

<1>Hoskisson, P.A., Smith, M.C.M.
<2>Hypervariation and phase variation in the bacteriophage resistome.
<3>Curr. Opin. Microbiol.
<4>10
<5>396-400
<6>2007
<7>Most bacteria encode proteins for defence against infection by bacteriophages. The mechanisms
that bring about phage defence are
extremely diverse, suggesting frequent independent evolution of novel
processes. Phage defence determinants are often plasmid or
phage-encoded and many that are chromosomal show evidence of lateral
transfer. Recent studies on restriction-modification (R-M) systems show
that these genes are amongst the most rapidly evolving. Some bacteria
have contingency genes that encode alternative target specificity
determinants for Type I or Type III R-M systems, thus expanding the
range of phages against which the host population is immune. The most
counter-intuitive observation, however, is the prevalence of phase
variation in many restriction systems, but recent arguments suggest
that switching off expression of R-M systems can aid phage defence.

<>

<1>Hossain, M.J., Beaz-Hidalgo, R., Figueras, M.J., Liles, M.R.
<2>Draft Genome Sequences of Two Novel Aeromonas Species Recovered in Association with Cyanobacterial Blooms.
<3>Genome Announcements
<4>2
<5>e01181-14
<6>2014
<7>Aeromonas aquatica and Aeromonas lacus are two new species that have been found in association
with cyanobacterial blooms from recreational Finnish lakes where
adverse human health effects have been recorded. Here, we present the draft
genome sequences of their type strains.

<>

<1>Hotchkiss, R.D.
<2>The quantitative separation of purines, pyrimidines, and nucleosides by paper chromatography.
<3>J. Biol. Chem.
<4>175
<5>315-332
<6>1948
<7>The separation of amino acid mixtures by migration with
organic solvents in filter paper has been successfully accomplished by
many workers since it was first described by Consden, Gordon, and
Martin.  Each amino acid travels in a more or less well defined spot in
the body of uniformly migrating solvent and can be visualized in the
dried paper as a local spot giving a color reaction with ninhydrin.  The
present paper reports the separation of the purines and pyrimidines
contained in nucleic acids, and several related compounds, by the
movement of a boundary of n-butyl alcohol along paper strips.  Vischer
and Chargaff have described the principal steps of a procedure for
separating the two bases, guanine and adenine, from nucleic acid in a
moving body of a quinoline-collidine mixture.  These purines were
located in the paper by precipitation of mercury sulfide after formation
of the insoluble mercury salts and washing in dilute acid.  After
complete removal of the quinoline and collidine it was possible to
identify the guanine and adenine by their absorption maxima in the
ultraviolet range.

<>

<1>Hotz, H.R., Peters, A.H.
<2>Protein demethylation required for DNA methylation.
<3>Nat. Genet.
<4>41
<5>10-11
<6>2009
<7>DNA methylation and histone modifications have essential roles in the transcriptional
regulation of gene expression. Lysine-specific demethylase-1 (Lsd1), also called KDM1, is an
enzyme with specificity toward the di and monomethylation states of lysine 4 and lysine 9 of
histone H3 (H3K4 and H3K9), and of lysine 370 of p53 (ref. 1). Lsd1 serves transcriptional
co-activator and co-repressor functions during development. On page 125 of this issue, En Li,
Taiping Chen and colleagues3 identify DNA methyltransferase-1 (Dnmt1) as a novel substrate for
Lsd1 and show that demethylation of Dnmt1 is required for the maintenance of global DNA
methylation.

<>

<1>Hou, P., Ji, M., He, N., Lu, Z.
<2>A microarray method to evaluate the effect of CA mispairs on the accuracy of BstUI restriction endonuclease.
<3>Anal. Biochem.
<4>317
<5>276-279
<6>2003
<7>Protein-DNA interaction is an essential event in many biological processes, such as
transcription, replication, restriction, and modification.  The sequence selectivity of
DNA-binding proteins plays an important role in controlling these processes in the cell.
Understanding how DNA-binding proteins select the correct DNA sequence from the nonspecific
sequences has attracted more and more interest in recent years.  Among DNA-binding proteins,
restriction endonucleases are perhaps the most extreme in their DNA selectivity.  Research on
the effects of sequence variations of the canonical recognition sites on the cleavage
efficiency of restriction endonucleases is important for understanding its specificity.  These
sequence variations include both a single base pair change and mispairs within the recognition
site.  The former can result in over a million-fold decrease in activity, and the latter can
impart a local destabilization on a double helix and affect the deformability and dynamic
properties of the DNA at the mismatched site.

<>

<1>Hou, P., Ji, M.J., He, N.Y., Lu, Z.H.
<2>Microarray-based method to evaluate the accuracy of restriction endonucleases HpaII and MspI.
<3>Biochem. Biophys. Res. Commun.
<4>314
<5>110-117
<6>2004
<7>A double-strand DNA (ds DNA) microarray was fabricated to analyze the structural perturbations
caused by methylation and the different base
mismatches in the interaction of the restriction endonucleases HpaII
and MspI with DNA. First, a series of synthesized oligonucleotides were
arrayed on the aldehyde-coated glass slides. Second, these
oligonucleotides were hybridized with target sequences to obtain a ds DNA
microarray, which includes several types of double strands with-the
fully methylated, semi-methylated, and unmethylated canonical
recognition sequences, semi-methylated and unmethylated base mismatches
within the recognition sequences. The cleavage experiments were carried
out under normal buffer conditions. The results indicated that MspI
could partially cleave methylated and semi-methylated canonical
recognition sequences. In contrast, HpaII could not cleave methylated
and semi-methylated canonical recognition sequences. HpaII and MspI
could both cleave the unmethylated canonical recognition sequence.
However, HpaII could partially cleave the sequence containing one GG
mismatch and not cleave other base mismatches in the corresponding
recognition site. In contrast, MspI could not recognize the base
mismatches within the recognition sequence. A good reproducibility was
observed in several parallel experiments. The experiment indicates that
the microarray technology has great potentials in high-throughput
identifying important interactions between protein and DNA.

<>

<1>Hou, S. et al.
<2>Genome sequence of the deep-sea {gamma}-proteobacterium Idiomarina loihiensis reveals amino acid fermentation as a source of carbon and   energy.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>18036-18041
<6>2004
<7>We report the complete genome sequence of the deep-sea gamma-proteobacterium, Idiomarina
loihiensis, isolated recently from a
hydrothermal vent at 1,300-m depth on the Loihi submarine volcano, Hawaii.
The I. loihiensis genome comprises a single chromosome of 2,839,318 base
pairs, encoding 2,640 proteins, four rRNA operons, and 56 tRNA genes. A
comparison of I. loihiensis to the genomes of other gamma-proteobacteria
reveals abundance of amino acid transport and degradation enzymes, but a
loss of sugar transport systems and certain enzymes of sugar metabolism.
This finding suggests that I. loihiensis relies primarily on amino acid
catabolism, rather than on sugar fermentation, for carbon and energy.
Enzymes for biosynthesis of purines, pyrimidines, the majority of amino
acids, and coenzymes are encoded in the genome, but biosynthetic pathways
for Leu, Ile, Val, Thr, and Met are incomplete. Auxotrophy for Val and Thr
was confirmed by in vivo experiments. The I. loihiensis genome contains a
cluster of 32 genes encoding enzymes for exopolysaccharide and capsular
polysaccharide synthesis. It also encodes diverse peptidases, a variety of
peptide and amino acid uptake systems, and versatile signal transduction
machinery. We propose that the source of amino acids for I. loihiensis
growth are the proteinaceous particles present in the deep sea
hydrothermal vent waters. I. loihiensis would colonize these particles by
using the secreted exopolysaccharide, digest these proteins, and
metabolize the resulting peptides and amino acids. In summary, the I.
loihiensis genome reveals an integrated mechanism of metabolic adaptation
to the constantly changing deep-sea hydrothermal ecosystem.

<>

<1>Houlston, C.E., Lindsay, H., Pradhan, S., Adams, R.L.P.
<2>DNA substrate specificity of pea DNA methylase.
<3>Biochem. J.
<4>293
<5>617-624
<6>1993
<7>DNA methylase, present in low-salt extracts of nuclei prepared from Pisum sativum shoot tips,
methylates model DNA substrates containing CNG trinucleotides or CI dinucleotides only. The
binding to the hemimethylated trinucleotide substrates is very much stronger and more
persistent than the binding to the unmethylated substrates or to the hemimethylated
dinucleotide substrate. When the DNA concentration is limiting, the rate of methyl-group
transfer with the hemimethylated CNG substrate is much greater than that with the unmethylated
CNG. However, the Vmax. is similar for the two CNG substrates. On fractionation using
Q-Sepharose, two peaks of activity are seen with different relative activities using the di-
and trinucleotide substrates. The relative activity with these substrates changes during
purification, during plant growth and on heating at 35oC as well, indicating that more than
one enzyme or more than one form of the enzyme may be present.

<>

<1>Howard, K.A., Card, C., Benner, J.S., Callahan, H.L., Maunus, R., Silber, K., Wilson, G., Brooks, J.E.
<2>Cloning the DdeI restriction-modification system using a two-step method.
<3>Nucleic Acids Res.
<4>14
<5>7939-7951
<6>1986
<7>DdeI, a Type II restriction-modification system from the gram-negative
anaerobic bacterium Desulfovibrio desulfuricans, recognizes the sequence CTNAG.
The system has been cloned into E. coli in two steps.  First the methylase
gene was cloned into pBR322 and a derivative expressing higher levels was
constructed.  Then the endonuclease gene was located by Southern blot analyses;
BamHI fragments large enough to contain the gene were cloned into pACYC184,
introduced into a host containing the methylase gene, and screened for
endonuclease activity.  Both genes are stably maintained in E. coli on separate
but compatible plasmids.  The DdeI methylase is shown to be a cytosine
methylase.  DdeI methylase clones decrease in viability as methylation activity
increases in E. coli RR1 (our original cloning strain).  Therefore the DdeI
system has been cloned and maintained in ER1467, a new E. coli cloning strain
engineered to accept cytosine methylases.  Finally, it has been demonstrated
that a very high level of methylation was necessary in the DdeI system for
successful introduction of the active endonuclease gene into E. coli.

<>

<1>Howden, B.P., Seemann, T., Harrison, P.F., McEvoy, C.R., Stanton, J.A., Rand, C.J., Mason, C.W., Jensen, S.O., Firth, N., Davies, J.K., Johnson, P.D., Stinear, T.P.
<2>Complete Genome Sequence of Staphylococcus aureus Strain JKD6008, an ST239 Clone of Methicillin-Resistant Staphylococcus aureus with  Intermediate-Level Vancomycin Resistance.
<3>J. Bacteriol.
<4>192
<5>5848-5849
<6>2010
<7>We report here the complete 2.92-Mb genome sequence of a clinical isolate of
methicillin-resistant Staphylococcus aureus subsp. aureus that
demonstrates intermediate-level vancomycin resistance. The strain, named
JKD6008, belongs to multilocus sequence type 239 and was isolated from the
bloodstream of a patient in New Zealand in 2003.

<>

<1>Hsieh, C.-L.
<2>In vivo activity of murine de novo methyltransferases, Dnmt3a and Dnmt3b.
<3>Mol. Cell. Biol.
<4>19
<5>8211-8218
<6>1999
<7>The putative de novo methyltransferases, Dnmt3a and Dnmt3b, were reported to have weak
methyltransferase activity in methylating the 3' long terminal repeat of Moloney murine
leukemia virus in vitro. The activity of these enzymes was evaluated in vivo, using a stable
episomal system that employs plasmids as targets for DNA methylation in human cells. De novo
methylation of a subset of the CpG sites on the stable episomes is detected in human cells
overexpressing the murine Dnmt3a or Dnmt3b1 protein.  This de novo methylation activity is
abolished when the cysteine in the P-C motif, which is the catalytic site of cytosine
methyltransferases, is replaced by a serine. The pattern of methylation on the episome is
nonrandom, and different regions of the episome are methylated to different extents.
Furthermore, Dnmt3a also methylates the sequence methylated by Dnmt3a on the stable episome in
the corresponding chromosomal target. Overexpression of human DNMT1 or murine Dnmt3b does not
lead to the same pattern or degree of de novo methylation on the episome as overexpression of
murine Dnmt3a. This finding suggests that these three enzymes may have different targets or
requirements, despite the fact that weak de novo methyltransferase activity has been
demonstrated in vitro for all three enzymes. It is also noteworthy that both Dnmt3a and Dnmt3b
proteins coat the metaphase chromosomes while displaying a more uniform pattern in the
nucleus. This is the first evidence that Dnmt3a and Dnmt3b have de novo methyltransferase
function in vivo and the first indication that the Dnmt3a and Dnmt3b proteins may have
preferred target sites.

<>

<1>Hsieh, P.-C., Xiao, J.-P., O'Loane, D., Xu, S.-Y.
<2>Cloning, expression, and purification of a thermostable nonhomodimeric restriction enzyme, BslI.
<3>J. Bacteriol.
<4>182
<5>949-955
<6>2000
<7>BslI is a thermostable type II restriction endonuclease with interrupted recognition sequence
CCNNNNN/NNGG (/, cleavage position).  The BslI restriction-modification system from Bacillus
species was cloned and expressed in Escherichia coli.  The system is encoded by three genes:
the 2,739-bp BslI methylase gene (bslIM), the bslIRalpha gene, and the bslIRbeta gene.  The
alpha and beta subunits of BslI can be expressed independently in E. coli in the absence of
BslI methylase (M.BslI) protection.  BslI endonuclease activity can be reconstituted in vitro
by mixing the two subunits together.  Gel filtration chromatography and native polyacrylamide
gel electrophoresis indicated that BslI forms heterodimers (alphabeta), heterotetramers
(alpha2beta2), and possibly oligomers in solution.  Two beta subunits can be cross-linked by a
chemical cross-linking agent, indicating formation of heterotetramer BslI complex
(alpha2beta2).  In DNA mobility shift assays, neither subunit alone can bind DNA.  DNA
mobility shift activity was detected after mixing the two subunits together.  Because of the
symmetric recognition sequence of the BslI endonuclease, we propose that its active form is
alpha2beta2.  M.BslI contains nine conserved motifs of N-4 cytosine DNA methylases within the
beta group of aminomethyltransferases.  Synthetic duplex deoxyoligonucleotides containing
cytosine hemimethylated or fully methylated at N-4 in BslI sites in the first or second
cytosine are resistant to BslI digestion.  C-5 methylation of the second cytosine on both
strands within the recognition sequence also renders the site refractory to BslI digestion.
Two putative zinc fingers are found in the alpha subunit of BslI endonuclease.

<>

<1>Hsu, D.W., Lin, M.J., Lee, T.L., Wen, S.C., Chen, X., Shen, C.J.
<2>Two major forms of DNA (cytosine-5) methyltransferase in human somatic tissues.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>9751-9756
<6>1999
<7>Thus far, only one major form of vertebrate DNA (cytosine-5) methyltransferase (CpG MTase, EC
2.1.1.37) has been identified, cloned,
and extensively studied. This enzyme, dnmt1, has been hypothesized to
be responsible for most of the maintenance as well as the de novo
methylation activities occurring in the somatic cells of vertebrates.
We now report the discovery of another abundant species of CpG MTase in
various types of human cell lines and somatic tissues. Interestingly,
the mRNA encoding this CpG MTase results from alternative splicing of
the primary transcript from the Dnmt1 gene, which incorporates in-frame
an additional 48 nt. between exons 4 and 5. Furthermore, this 48-nt exon
sequence is derived from the first, or the most upstream, copy of a set
of seven different Alu repeats located in intron 4. The ratios of
expression of this mRNA to the expression of the previously known,
shorter Dnmt1 mRNA species, as estimated by semiquantitative reverse
transcription-PCR analysis, range from two-thirds to three-sevenths.
This alternative splicing scheme of the Dnmt1 transcript seems to be
conserved in the higher primates. We suggest that the originally
described and the recently discovered forms of CpG MTase be named dnmt1-
a and dnmt1-b, respectively. The evolutionary and biological
implications of this finding are discussed in relation to the cellular
functions of the CpG residues and the CpG MTases.

<>

<1>Hsu, M., Berg, P.
<2>Altering the specificity of restriction endonuclease:  effect of replacing Mg2+ with Mn2+.
<3>Biochemistry
<4>17
<5>131-138
<6>1978
<7>In the presence of 100 mM Tris buffer (pH 7.5) and 1-10 mM Mg2+ EcoRI
endonuclease cleaves DNA at a specific nucleotide sequence and in a
characteristic way: -G^AATTC-.  But if Mg2+ is replaced by Mn2+, the
specificity of the cleavage is relaxed and cleavages occur at many other sites;
moreover, there appears to be a hierarchy of cleavage rates at the pseudo-EcoRI
restriction sites.  For example, SV40 DNA is cleaved only once in the usual
digestion conditions, but with Mn2+ more than ten cleavages are made; the five
most rapidly cleaved SV40 DNA map locations are 0/1.0>0.93>0.33 ~0.49>0.25.
Mn2+ also alters the restriction specificity of HindIII but not HpaII
endonuclease.

<>

<1>Hsu, M.-Y., Inouye, M., Inouye, S.
<2>Retron for the 67-base multicopy single-stranded DNA from Escherichia coli: A potential transposable element encoding both reverse transcriptase and Dam methylase functions.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>87
<5>9454-9458
<6>1990
<7>The region (retron-Ec67) required for the biosynthesis of a branched-RNA linked multicopy
single-stranded DNA (msDNA-Ec67) from a clinical isolate of Escherichia coli was mapped at a
position equivalent to 19 min on the K-12 chromosome. The element containing the retron
consisted of a unique 34-kilobase sequence that was flanked by direct repeats of a
26-base-pair sequence found in the K-12 chromosomal DNA. This suggests that the 34-kilobase
element was probably integrated into the E. coli genome by a mechanism related to
transposition or phage integration. In the 34-kilobase sequence an open reading frame of 285
residues was found, which displays 44% sequence identity with the E. coli Dam methylase.
Interestingly, there are three GATC sequences, the site of Dam methylation, in the promoter
region of the gene for reverse transcriptase.

<>

<1>Hu, A., He, J., Chu, K.H., Yu, C.P.
<2>Genome Sequence of the 17{beta}-Estradiol-Utilizing Bacterium Sphingomonas Strain KC8.
<3>J. Bacteriol.
<4>193
<5>4266-4267
<6>2011
<7>Sphingomonas strain KC8 is known for its ability to utilize 17beta-estradiol, a natural
estrogen and an environmental
endocrine-disrupting compound, as the sole carbon and energy source. Here,
we report the draft genome sequence of the strain KC8 (4,074,265 bp, with
a GC content of 63.7%) and major findings from its annotation.

<>

<1>Hu, A., Lv, M., Yu, C.P.
<2>Draft Genome Sequence of the Bisphenol A-Degrading Bacterium Sphingobium sp. Strain YL23.
<3>Genome Announcements
<4>1
<5>e00549-13
<6>2013
<7>Sphingobium sp. strain YL23, a novel bacterium isolated from sewage sludge of a domestic
wastewater treatment plant, has been shown to completely degrade
bisphenol A under aerobic conditions. Here, we describe a 3.8-Mb assembly of its
genome sequence and major findings from its annotation.

<>

<1>Hu, A.-L.W.
<2>NgoAIV--a new restriction endonuclease compared to its commercially available isoschizomers.
<3>BRL Focus
<4>15
<5>42-43
<6>1994
<7>The restriction endonuclease NaeI recognizes the sequence GCC^GGC and generates blunt-ended
DNA fragments. However, NaeI demonstrates a strong site preference in its cleavage rate that
may result in incomplete cleavage in some DNA. Therefore, there is a need to search for an
isoschizomer to circumvent the problem. NgoAIV is an isoschizomer of NaeI that was purified
from a strain of E. coli bearing the cloned NgoAIV gene from Neisseria gonorrhoea. NgoAIV
cleaves between the first G and C residues, providing cohesive ends for more effective
ligation. The enzyme has an apparent molecular weight of 31,000 Da on an SDS-PAGE gel.

<>

<1>Hu, A.W., Marschel, A.H.
<2>Endonuclease NciI generates atypical termini.
<3>Fed. Proc.
<4>41
<5>1119
<6>1982
<7>Cleavage of DNA by the type II restriction endonuclease, NciI, produces termini
that differ from those produced by most other type II enzymes.  This difference
is manifested by the inability of T4 DNA ligase to ligate NciI generated
termini, even under conditions that normally enhance ligation.  Tests for
contaminating enzymes present in NciI preparations showed negligible levels of
3' or 5' single or double strand specific exonucleases.  Similarly, no other
endonuclease was detected in these preparations.  The presence of a ligation
inhibitor in NciI preparations was eliminated by mixing DNA fragments produced
by HpaII or TaqI with NciI fragments and measuring ligation.  Only the NciI
generated fragments remained resistant to the ligation reaction precluding the
presence of a nonspecific ligation inhibitor.  A successful ligation procedure
was designed and included a two step incubation with T4 polynucleotide kinase
and NciI formed fragments.  Step one utilized conditions that enhanced the
inherent 3' phosphatase activity of T4 polynucleotide kinase.  The 2nd
incubation step optimized the phosphorylation of the 5' hydroxyl terminus.  DNA
fragments treated in this manner were ligated with more than a 50% efficiency
under normal conditions.  The above result suggests that NciI cleaves DNA
leaving a 3' phosphate and 5' hydroxyl, and explains the inability of T4 DNA
ligase to ligate NciI generated fragments.

<>

<1>Hu, D., Crist, M., Duan, X., Gimble, F.S.
<2>Mapping of a DNA binding region of the PI-SceI homing endonuclease by affinity cleavage and alanine-scanning mutagenesis.
<3>Biochemistry
<4>38
<5>12621-12628
<6>1999
<7>The PI-SceI protein is a member of the LAGLIDADG family of homing endonucleases that is
generated by a protein splicing reaction. PI-SceI has a bipartite domain structure, and the
protein splicing and endonucleolytic reactions are catalyzed by residues in domains I and II,
respectively. Structural and mutational evidence indicates that both domains mediate DNA
binding. Treatment of the protein with trypsin breaks a peptide bond within a disordered
region of the endonuclease domain situated between residues Val-270 and Leu-280 and interferes
with the ability of this domain to bind DNA. To identify specific residues in this region that
are involved in DNA binding and/or catalysis, alanine-scanning mutagenesis was used to create
a set of PI-SceI mutant proteins that were assayed for activity. One of these mutants, N281A,
was >300-fold less active than wild-type PI-SceI, and two other proteins, R277A and N284A,
were completely inactive. These decreases in cleavage activity parallel similar decreases in
substrate binding by the endonuclease domains of these mutant proteins. We mapped the
approximate position of the disordered region to one of the ends of the 31 base pair PI-SceI
recognition sequence using mutant proteins that were substituted with cysteine at residues
Asn-274 and Glu-283 and tethered to the chemical nuclease FeBABE. These mutational and
affinity cleavage data strongly support a model of PI-SceI docked to its DNA substrate that
suggests that one or more residues identified here are responsible for contacting base pair
A/T(-)(9), which is essential for substrate binding.

<>

<1>Hu, D., Crist, M., Duan, X., Quiocho, F.A., Gimble, F.S.
<2>Probing the Structure of the PI-SceI-DNA Complex by Affinity Cleavage and Affinity Photocross-linking.
<3>J. Biol. Chem.
<4>275
<5>2705-2712
<6>2000
<7>The PI-SceI protein is an intein-encoded homing endonuclease that initiates the mobility of
its gene by making a double strand break at a single site in the yeast genome. The PI-SceI
protein splicing and endonucleolytic active sites are separately located in each of two
domains in the PI-SceI structure. To determine the spatial relationship between bases in the
PI-SceI recognition sequence and selected PI-SceI amino acids, the PI-SceI-DNA complex was
probed by photocross-linking and affinity cleavage methods. Unique solvent-accessible cysteine
residues were introduced into the two PI-SceI domains at positions 91, 97, 170, 230, 376, and
378, and the mutant proteins were modified with either 4-azidophenacyl bromide or iron
(S)-1-(p-bromoacetamidobenzyl)-ethylenediaminetetraacetate (FeBABE). The phenyl azide-coupled
proteins cross-linked to the PI-SceI target sequence, and the FeBABE-modified proteins cleaved
the DNA proximal to the derivatized amino acid. The results suggest that an extended
beta-hairpin loop in the endonuclease domain that contains residues 376 and 378 contacts the
major groove near the PI-SceI cleavage site. Conversely, residues 91, 97, and 170 in the
protein splicing domain are in close proximity to a distant region of the substrate. To
interpret our results, we used a new PI-SceI structure that is ordered in regions of the
protein that bind DNA. The data strongly support a model of the PI-SceI-DNA complex derived
from this structure.

<>

<1>Hu, D., Li, X., Chang, Y., He, H., Zhang, C., Jia, N., Li, H., Wang, Z.
<2>Genome Sequence of Streptomyces sp. Strain TOR3209, a Rhizosphere Microecology Regulator Isolated from Tomato Rhizosphere.
<3>J. Bacteriol.
<4>194
<5>1627
<6>2012
<7>Streptomyces sp. strain TOR3209, isolated from tomato rhizosphere, can regulate the
rhizosphere microecology of a variety of crops. Strain TOR3209 could improve
plant systemic resistance and promote plant growth. Here, the genome sequence of
strain TOR3209 is reported, providing the molecular biological basis of the
regulation mechanism of rhizosphere microecology.

<>

<1>Hu, K.-Y., Wuu, J.-A., Kao, M.-C., Liu, Y.-T., Pai, S.-H.
<2>Isolation and characterization of a newly identified type II restriction endonuclease from a local Streptomyces sp. in Taiwan.
<3>Appl. Biochem. Biotechnol.
<4>73
<5>231-241
<6>1998
<7>Streptomyces chusanensis ZS-2, isolated from a soil sample in Chusan in Taiwan, was found to
produce a new Type II restriction endonuclease. This restriction enzyme was designated as
SchI. The purified enzyme was characterized as having a subunit mol wt of 28 kDa, and was
apparently free from exonuclease activities. It cleaves the phosphodiester bond between the
fourth C and the fifth G on the 5'-CCGCGG-3' sequence of DNAs, leaving a 2-nucleotide
protruding end at its 3' site. This data suggests that SchI is an isoschizomer of SacII. In
addition, based on the comparison between SchI and SacII regarding reaction parameters, it
seems that SchI is a better choice of restriction enzyme for genetic analysis and mapping.

<>

<1>Hu, K.-Y., Wuu, J.-A., Kao, M.-C., Liu, Y.-T., Pai, S.-H.
<2>A newly identified restriction enzyme, SchI, from a local strain of streptomyces sp. in Taiwan.
<3>Taiwanese Patent Office
<4>TW 432110 B
<5>14
<6>2001
<7>Streptomyces chusanensis ZS-2, isolated from a soil sample in Chusan of Taiwan, was found to
produce a new type II restriction endonuclease.  This restriction enzyme was designated as
SchI.  The purified enzyme was characterized to be a homodimer with a subunit molecular weight
of 28 kDa.  It was apparently free from exonuclease activities.  It cleaves the phosphodiester
bond between the 4th 'C' and the 5th 'G' on the 5'-CCGCGG-3' sequence of DNAs leaving a
2-nucleotide protruding end at its 3' site.  This data suggests that SchI is an isoschizomer
of SacII.  In addition, based on the comparison between SchI and SacII regarding reaction
parameters, it seems that SchI is a better choice of restriction enzyme for genetic analysis
and mapping.

<>

<1>Hu, L., Li, N., Xu, C., Zhong, S., Lin, X., Yang, J., Zhou, T., Yuliang, A., Wu, Y., Chen, Y.R., Cao, X., Zemach, A., Rustgi, S., von Wettstein, D., Liu, B.
<2>Mutation of a major CG methylase in rice causes genome-wide hypomethylation, dysregulated genome expression, and seedling lethality.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>111
<5>10642-10647
<6>2014
<7>Cytosine methylation at CG sites (mCG) plays critical roles in development, epigenetic
inheritance, and genome stability in mammals and plants. In the dicot
model plant Arabidopsis thaliana, methyltransferase 1 (MET1), a principal CG
methylase, functions to maintain mCG during DNA replication, with its null
mutation resulting in global hypomethylation and pleiotropic developmental
defects. Null mutation of a critical CG methylase has not been characterized at a
whole-genome level in other higher eukaryotes, leaving the generality of the
Arabidopsis findings largely speculative. Rice is a model plant of monocots, to
which many of our important crops belong. Here we have characterized a null
mutant of OsMet1-2, the major CG methylase in rice. We found that seeds
homozygous for OsMet1-2 gene mutation (OsMET1-2-/-), which directly segregated
from normal heterozygote plants (OsMET1-2+/-), were seriously maldeveloped, and
all germinated seedlings underwent swift necrotic death. Compared with wild type,
genome-wide loss of mCG occurred in the mutant methylome, which was accompanied
by a plethora of quantitative molecular phenotypes including dysregulated
expression of diverse protein-coding genes, activation and repression of
transposable elements, and altered small RNA profiles. Our results have revealed
conservation but also distinct functional differences in CG methylases between
rice and Arabidopsis.

<>

<1>Hu, P., Elliott, J., McCready, P., Skowronski, E., Garnes, J., Kobayashi, A., Brubaker, R.R., Garcia, E.
<2>Structural organization of virulence-associated plasmids of Yersinia pestis.
<3>J. Bacteriol.
<4>180
<5>5192-5202
<6>1998
<7>The complete nucleotide sequence and gene organization of the three virulence plasmids from
Yersinia pestis KIM5 were determined. Plasmid
pPCP1 (9,610 bp) has a GC content of 45.3% and encodes two previously
known virulence factors, an associated protein, and a single copy of
IS100. Plasmid pCD1 (70,504 bp) has a GC content of 44.8%. It is known to
encode a number of essential virulence determinants, regulatory functions,
and a multiprotein secretory system comprising the low-calcium response
stimulation that is shared with the other two Yersinia species pathogenic
for humans (Y. pseudotuberculosis and Y. enterocolitica). A new
pseudogene, which occurs as an intact gene in the Y. enterocolitica and Y.
pseudotuberculosis-derived analogues, was found in pCD1. It corresponds to
that encoding the lipoprotein YlpA. Several intact and partial insertion
sequences and/or transposons were also found in pCD1, as well as six
putative structural genes with high homology to proteins of unknown
function in other yersiniae. The sequences of the genes involved in the
replication of pCD1 are highly homologous to those of the cognate plasmids
in Y. pseudotuberculosis and Y. enterocolitica, but their localization
within the plasmid differs markedly from those of the latter. Plasmid pMT1
(100,984 bp) has a GC content of 50.2%. It possesses two copies of IS100,
which are located 25 kb apart and in opposite orientations. Adjacent to
one of these IS100 inserts is a partial copy of IS285. A single copy of an
IS200-like element (recently named IS1541) was also located in pMT1. In
addition to 5 previously described genes, such as murine toxin, capsule
antigen, capsule anchoring protein, etc., 30 homologues to genes of
several bacterial species were found in this plasmid, and another 44 open
reading frames without homology to any known or hypothetical protein in
the databases were predicted.

<>

<1>Hu, P., Lang, J., Wawrousek, K., Yu, J., Maness, P.C., Chen, J.
<2>Draft Genome Sequence of Rubrivivax gelatinosus CBS.
<3>J. Bacteriol.
<4>194
<5>3262
<6>2012
<7>Rubrivivax gelatinosus CBS, a purple nonsulfur photosynthetic bacterium, can grow
photosynthetically using CO and N(2) as the sole carbon and nitrogen nutrients,
respectively. R. gelatinosus CBS is of particular interest due to its ability to
metabolize CO and yield H(2). We present the 5-Mb draft genome sequence of R.
gelatinosus CBS with the goal of providing genetic insight into the metabolic
properties of this bacterium.

<>

<1>Hu, P., Yang, M., Zhang, A., Wu, J., Chen, B., Hua, Y., Yu, J., Chen, H., Xiao, J., Jin, M.
<2>Complete Genome Sequence of Streptococcus suis Serotype 3 Strain ST3.
<3>J. Bacteriol.
<4>193
<5>3428-3429
<6>2011
<7>Streptococcus suis is a zoonotic pathogen, causing economic loss in swine industry, and is
also a threat to human health. To date, the mechanism of
pathogenisis is not fully understood. Here, we report the complete genome
sequence of S. suis strain ST3 of serotype 3, which provides opportunities
to reveal genetic basis of infection of S. suis non-serotype 2 strains.

<>

<1>Hu, P., Yang, M., Zhang, A., Wu, J., Chen, B., Hua, Y., Yu, J., Xiao, J., Jin, M.
<2>Complete Genome Sequence of Streptococcus suis Serotype 14 Strain JS14.
<3>J. Bacteriol.
<4>193
<5>2375-2376
<6>2011
<7>Streptococcus suis is an important zoonotic agent leading to a variety of diseases in swine
and can be transmitted to human being upon close contact. Here, we report the complete genome
sequence of S. suis serotype 14 strain JS14 which was isolated from a diseased pig in Jiangsu
Province, China.

<>

<1>Hu, S., Zheng, H., Gu, Y., Zhao, J., Zhang, W., Yang, Y., Wang, S., Zhao, G., Yang, S., Jiang, W.
<2>Comparative genomic and transcriptomic analysis revealed genetic characteristics related to solvent formation and xylose utilization in Clostridium acetobutylicum EA 2018.
<3>BMC Genomics
<4>12
<5>93
<6>2011
<7>ABSTRACT: BACKGROUND: Clostridium acetobutylicum, a gram-positive and
spore-forming anaerobe, is a major strain for the fermentative production
of acetone, butanol and ethanol. But a previously isolated hyper-butanol
producing strain C. acetobutylicum EA 2018 does not produce spores and has
greater capability of solvent production, especially for butanol, than the
type strain C. acetobutylicum ATCC 824. RESULTS: Complete genome of C.
acetobutylicum EA 2018 was sequenced using Roche 454 pyrosequencing.
Genomic comparison with ATCC 824 identified many variations which may
contribute to the hyper-butanol producing characteristics in the EA 2018
strain, including a total of 46 deletion sites and 26 insertion sites. In
addition, transcriptomic profiling of gene expression in EA 2018 relative
to that of ATCC824 revealed expression-level changes of several key genes
related to solvent formation. For example, spo0A and adhEII have higher
expression level, and most of the acid formation related genes have lower
expression level in EA 2018. Interestingly, the results also showed that
the variation in CEA_G2622 (CAC2613 in ATCC 824), a putative
transcriptional regulator involved in xylose utilization, might accelerate
utilization of substrate xylose. CONCLUSIONS: Comparative analysis of C.
acetobutylicum hyper-butanol producing strain EA 2018 and type strain ATCC
824 at both genomic and transcriptomic levels, for the first time,
provides molecular-level understanding of non-sporulation, higher solvent
production and enhanced xylose utilization in the mutant EA 2018. The
information could be valuable for further genetic modification of C.
acetobutylicum for more effective butanol production.

<>

<1>Hu, W., Wang, C., Liang, J., Zhang, T., Hu, Z., Wang, Z., Lan, W., Li, F., Wu, H., Ding, J., Wu, G., Deng, Z., Cao, C.
<2>Structural insights into DndE from Escherichia coli B7A involved in DNA phosphorothioation modification.
<3>Cell Res.
<4>22
<5>1203-1206
<6>2012
<7>DNA phosphorothioate modification, originally developed as an artificial tool to stabilize
oligodeoxynucleotides against nuclease degradation, was recently found to be incorporated with
sulfur into DNA backbone as a novel physiological variation by the five-gene dnd cluster
(dndA-dndE) products in a sequence- and stereo-specific manner.  This PT modification causes
the DNA degradation (Dnd) phenotype and is widespread and quantized in bacterial genomes,
working as a part of a restriction modification system.  This modification can be specifically
cleaved in vitro by type IV restriction endonuclease.  DndA works as a cysteine desulfurase
and assembles DndC as a 4Fe-4S cluster protein.  DndC possesses ATP pyrophosphatase activity
and is predicted to have 3'-phosphoadenosine-5'-phosphosulfate reductase activity, whereas
DndB has homology to a group of transcriptional regulators.  DndD, known as SpfD in
Pseudomonas fluorescens Pf0-1, has ATPase activity possibly related to DNA structure
alteration of nicking during PT incorporation.  Sequence identity (46%) and similarity (61%)
to phosphoribosylaminoimidazole carboxylase (NCAIR synthetase) from Anabaena variabilis
suggest that DndE could be an NCAIR synthase analogue.  However, DndE may also act as a
sulfotransferase due to a specific PAPS binding sequence AAVGK-TLLIHLHR contained in the
C-terminus of DndE from Streptomyces lividans.  Therefore, the exact function of DndE remains
unknown.

<>

<1>Hu, W.P., Yu, H.S., Hsieh, M.C., Chen, Y.C., Wang, J.J.
<2>Evaluation of DNA-binding ability of antibiotic DC-81 - indole conjugates by restriction endonuclease BamHI and molecular modeling studies.
<3>Chin. Pharm. J.
<4>54
<5>465-470
<6>2002
<7>DC-81, an antitumor antibiotic produced by Streptomyces species, belongs to the
pyrrolo-{2,1-c}{1,4}benzodiazepines which are potent inhibitors of nucleic acid synthesis
because of their ability to recognize and bind to specific sequence of DNA to form a labile
covalent adduct.  The hybrid agents comprised of DC-81 and indole carboxylate moiety 2 through
carbon chain linkers (comprised of zero and two to four carbons) have been designed and
synthesized by our group.  These compounds with DNA binding ability were determined by
restriction endonuclease BamHI and molecular modeling studies.  The results demonstrated that
DC-81 - IC hybrid 3c (with a three carbon chain linker) showed a higher DNA-binding affinity
and gave rise to a maximal stabilization of the complex with DNA at the minor groove as
compared to the other complexes formed with 3a, 3b and 3d.

<>

<1>Hu, X., Fan, W., Han, B., Liu, H., Zheng, D., Li, Q., Dong, W., Yan, J., Gao, M., Berry, C., Yuan, Z.
<2>Complete genome sequence of the mosquitocidal bacterium Bacillus sphaericus C3-41 and comparison with those of closely related Bacillus  species.
<3>J. Bacteriol.
<4>190
<5>2892-2902
<6>2008
<7>Bacillus sphaericus strain C3-41 is an aerobic, mesophilic, spore-forming bacterium that has
been used with great success in mosquito control
programs worldwide. Genome sequencing revealed that the complete genome of
this entomopathogenic bacterium is composed of a chromosomal replicon of
4,639,821 bp and a plasmid replicon of 177,642 bp, containing 4,786 and
186 potential protein-coding sequences, respectively. Comparison of the
genome with other published sequences indicated that the B. sphaericus
C3-41 chromosome is most similar to that of Bacillus sp. strain NRRL
B-14905, a marine species that, like B. sphaericus, is unable to
metabolize polysaccharides. The lack of key enzymes and sugar transport
systems in the two bacteria appears to be the main reason for this
inability, and the abundance of proteolytic enzymes and transport systems
may endow these bacteria with exclusive metabolic pathways for a wide
variety of organic compounds and amino acids. The genes shared between B.
sphaericus C3-41 and Bacillus sp. strain NRRL B-14905, including mobile
genetic elements, membrane-associated proteins, and transport systems,
demonstrated that these two species are a biologically and
phylogenetically divergent group. Knowledge of the genome sequence of B.
sphaericus C3-41 thus increases our understanding of the bacilli and may
also offer prospects for future genetic improvement of this important
biological control agent.

<>

<1>Hu, X., Li, A., Lv, L., Yuan, C., Guo, L., Jiang, X., Jiang, H., Qian, G., Zheng, B., Guo, J., Li, L.
<2>High quality draft genome sequence of Staphylococcus cohnii subsp. cohnii strain  hu-01.
<3>Standards in Genomic Sciences
<4>9
<5>755-762
<6>2014
<7>Staphylococcus cohnii subsp. cohnii belongs to the family Staphylococcaceae in the order
Bacillales, class Bacilli and phylum Firmicutes. The increasing
relevance of S. cohnii to human health prompted us to determine the genomic
sequence of Staphylococcus cohnii subsp. cohnii strain hu-01, a
multidrug-resistant isolate from a hospital in China. Here we describe the
features of S. cohnii subsp. cohnii strain hu-01, together with the genome
sequence and its annotation. This is the first genome sequence of the species
Staphylococcus cohnii.

<>

<1>Hu, X., Wang, J., Wang, F., Chen, Q., Huang, Y., Cui, Z.
<2>Complete Genome Sequence of the p-Nitrophenol-Degrading Bacterium Pseudomonas putida DLL-E4.
<3>Genome Announcements
<4>2
<5>e00596-14
<6>2014
<7>The first complete genome sequence of a p-nitrophenol (PNP)-degrading bacterium is reported
here. Pseudomonas putida DLL-E4, a Gram-negative bacterium isolated
from methyl-parathion-polluted soil, can utilize PNP as the sole carbon and
nitrogen source. P. putida DLL-E4 has a 6,484,062 bp circular chromosome that
contains 5,894 genes, with a G+C content of 62.46%.

<>

<1>Hu, Y., Li, T., Yang, Z., Zhang, B., Li, Y.
<2>Phage resistance of Corynebacterium crenatum conferred by the restriction and modification system cglI.
<3>Sheng Wu Gong Cheng Xue Bao
<4>24
<5>760-765
<6>2008
<7>
<>

<1>Hu, Y., Xu, X., Song, P., Jiang, L., Zhang, Z., Huang, H.
<2>Draft Genome Sequence of Deinococcus xibeiensis R13, a New Carotenoid-Producing Strain.
<3>Genome Announcements
<4>1
<5>e00987-13
<6>2013
<7>Deinococcus xibeiensis strain R13, isolated from radiation-contaminated soils, synthesizes a
unique ketocarotenoid, deinoxanthin. Here, we present a 3.49-Mb
assembly of its genome sequence, which can help us find the key genes of the
deinoxanthin biosynthesis pathways and modify genes obtaining a high yield of the
new carotenoid.

<>

<1>Hu, Y., Zhang, W., Liang, H., Liu, L., Peng, G., Pan, Y., Yang, X., Zheng, B., Gao, G.F., Zhu, B., Hu, H.
<2>Whole-Genome Sequence of a Multidrug-Resistant Clinical Isolate of Acinetobacter lwoffii.
<3>J. Bacteriol.
<4>193
<5>5549-5550
<6>2011
<7>Acinetobacter lwoffii has been considered an opportunistic pathogen that can cause nosocomial
infections in humans. Here, we present the genome
sequence of A. lwoffii WJ10621, a multidrug-resistant clinical isolate
that carries a plasmid with the NDM-1 resistance gene.

<>

<1>Hu, Z.Y., Wang, Y.Z., Im, W.T., Wang, S.Y., Zhao, G.P., Zheng, H.J., Quan, Z.X.
<2>The first complete genome sequence of the class fimbriimonadia in the phylum armatimonadetes.
<3>PLoS ONE
<4>9
<5>E100794
<6>2014
<7>In this study, we present the complete genome of Fimbriimonas ginsengisoli Gsoil
348T belonging to the class Fimbriimonadia of the phylum Armatimonadetes,
formerly called as candidate phylum OP10. The complete genome contains a single
circular chromosome of 5.23 Mb including a 45.5 kb prophage. Of the 4820 open
reading frames (ORFs), 3,000 (62.2%) genes could be classified into Clusters of
Orthologous Groups (COG) families. With the split of rRNA genes, strain Gsoil
348T had no typical 16S-23S-5S ribosomal RNA operon. In this genome, the GC skew
inversion which was usually observed in archaea was found. The predicted gene
functions suggest that the organism lacks the ability to synthesize histidine,
and the TCA cycle is incomplete. Phylogenetic analyses based on ribosomal
proteins indicated that strain Gsoil 348T represents a deeply branching lineage
of sufficient divergence with other phyla, but also strongly involved in
superphylum Terrabacteria.

<>

<1>Hua, N.M., Karska-Wysocki, B., Szatmari, G., Mamet-Bratley, M.D.
<2>Purification and characterization of the LlaGI restriction  enodnuclease from Lactococcus lactis subsp. cremoris G2.
<3>Life Sc. Confer. Ottawa
<4>0
<5>P001
<6>1998
<7>In the dairy industry, Lactococcus lactis are widely used as starter cultures in the
manufacture of the dairy products.  Bacteriophage infection is a serious problem for the dairy
industry.  To date, four categories of natural phage defense mechanisms have been identified
in lactococci based on the mode of action: adsorption inhibition, penetration blocking,
abortive infection, and restriction-modification system.  L. lactis subsp. cremoris G2 has
previously been isolated and purified in our laboratory.  Strain G2, which encodes a type-II
R-M endonuclease, designated LlaGI, contains at least five plasmids.  When L. lactis strain G2
was cured from most of its plasmids, the specific nuclease activity was lost.  These results
strongly indicate that the R-M system is plasmid-encoded.  LlaGI, an isoschizomer of NheI,
which recognizes the palindromic DNA sequence 5'_GCTAGC_3' has been purified to homogeneity
by a two-step procedure using ion-exchange and affinity chromatography.  The enzyme yield was
in excess of 1000 units/g of wet cells.  The purified enzyme was free of nonspecific nuclease
and requires only magnesium ion for its activity.  Physical properties indicate that LlaGI is
present in solution as monomer of about 40kDa.

<>

<1>Hua, X., Zhou, H., Jiang, Y., Feng, Y., Chen, Q., Ruan, Z., Yu, Y.
<2>Genome Sequences of Two Multidrug-Resistant Acinetobacter baumannii Strains Isolated from a Patient before and after Treatment with Tigecycline.
<3>J. Bacteriol.
<4>194
<5>6979-6980
<6>2012
<7>Acinetobacter baumannii is a Gram-negative bacterium which emerged as a significant nosocomial
pathogen worldwide. To investigate the molecular basis of
the tigecycline-resistant mechanism, we determined the genome sequences of two
multidrug-resistant A. baumannii strains isolated from a patient before and after
treatment with tigecycline.

<>

<1>Huai, Q., Colandene, J.D., Chen, Y., Luo, F., Zhao, Y., Topal, M.D., Ke, H.
<2>Crystal structure of NaeI - an evolutionary bridge between DNA endonuclease and topoisomerase.
<3>EMBO J.
<4>19
<5>3110-3118
<6>2000
<7>NaeI is transformed from DNA endonuclease to DNA topoisomerase and recombinase by a single
amino acid substitution. The crystal structure of NaeI was solved at 2.3 A resolution and
shows that NaeI is a dimeric molecule with two domains per monomer. Each domain contains one
potential DNA recognition motif corresponding to either endonuclease or topoisomerase
activity. The N-terminal domain core folds like the other type II restriction endonucleases as
well as lambda-exonuclease and the DNA repair enzymes MutH and Vsr, implying a common
evolutionary origin and catalytic mechanism. The C-terminal domain contains a catabolite
activator protein (CAP) motif present in many DNA-binding proteins, including the type IA and
type II topoisomerases. Thus, the NaeI structure implies that DNA processing enzymes evolved
from a few common ancestors. NaeI may be an evolutionary bridge between endonuclease and DNA
processing enzymes.

<>

<1>Huai, Q., Colandene, J.D., Topal, M.D., Ke, H.
<2>Structure of NaeI-DNA complex reveals dual-mode DNA recognition and complete dimer rearrangement.
<3>Nat. Struct. Biol.
<4>8
<5>665-669
<6>2001
<7>NaeI, a novel DNA endonuclease, shows topoisomerase and recombinase activities when a Lys
residue is substituted for Leu 43. The NaeI-DNA structure demonstrates that each of the two
domains of NaeI recognizes one molecule of DNA duplex. DNA recognition induces dramatic
rearrangements: narrowing the binding site of the Topo domain 16 angstroms to grip DNA,
widening that
of the Endo domain 8 angstroms to encircle and bend DNA 45 degrees for cleavage, and
completely
rebuilding the homodimer interface. The NaeI-DNA structure presents the first example of novel
recognition of two copies of one DNA sequence by two different amino acid sequences and two
different structural motifs in one polypeptide.

<>

<1>Huang, B., Schaeffer, C.J., Li, Q., Tsai, M.-D.
<2>Splase: A new class IIS zinc-finger restriction endonuclease with specificity for Sp1 binding sites.
<3>J. Protein Chem.
<4>15
<5>481-489
<6>1996
<7>A new restriction endonuclease, named Sp1ase, was constructed by genetically fusing the
DNA-cleavage domain of the restriction endonuclease FokI with the zinc-finger DNA-binding
domain of the transcription factor Sp1.  The resulting protein was expressed in Escherichia
coli, partially purified, and shown to selectively digest plasmid DNA harboring consensus Sp1
sites.  Sp1ase was also shown to selectively digest the long terminal repeat of the HIV-1 DNA
at Sp1 sites.  Sp1ase recognizes a 10-bp DNA sequence and hydrolyzes phosphodiester bonds
upstream of the binding sequence.  The binding specificity of Sp1ase makes this a "rare
cutter" restriction enzyme which could be valuable in creating large DNA fragments for genome
sequencing projects.  The result also presents the opportunity to create other restriction
enzymes by altering the binding specificity of the zing-finger recognition helix.

<>

<1>Huang, E., Guo, Y., Yousef, A.E.
<2>Draft Genome Sequence of Paenibacillus sp. Strain OSY-SE, a Bacterium Producing the Novel Broad-Spectrum Lipopeptide Antibiotic Paenibacterin.
<3>J. Bacteriol.
<4>194
<5>6306
<6>2012
<7>A strain of Paenibacillus sp., OSY-SE, was isolated from soil and found to produce a novel
lipopeptide antibiotic. The antibiotic, paenibacterin, is active
against Gram-negative and Gram-positive bacterial pathogens. Paenibacterin is
biosynthesized by a nonribosomal peptide synthetase pathway. Here we report the
draft genome sequence of Paenibacillus sp. OSY-SE.

<>

<1>Huang, E., Yousef, A.E.
<2>Draft Genome Sequence of Paenibacillus polymyxa OSY-DF, Which Coproduces a Lantibiotic, Paenibacillin, and Polymyxin E1.
<3>J. Bacteriol.
<4>194
<5>4739-4740
<6>2012
<7>Paenibacillus polymyxa OSY-DF is a Gram-positive rod-shaped bacterium isolated from a
fermented vegetable food. This bacterial strain displays potent
antimicrobial activities against Gram-positive and Gram-negative pathogenic
bacteria, attributed to the production of the lantibiotic paenibacillin and the
colistin peptide polymyxin E1. Here we report the draft genome sequence of
Paenibacillus polymyxa OSY-DF.

<>

<1>Huang, E.-S., Newbold, J.E., Pagano, J.S.
<2>Analysis of simian virus 40 DNA with the restriction enzyme of Haemophilus aegyptius, endonuclease Z.
<3>J. Virol.
<4>11
<5>508-514
<6>1973
<7>Limited digestion of simian virus 40 (SV40) DNA from both small- and
large-plaque strains with the restriction endonuclease Z from Haemophilus
aegyptius yielded 10 specific fragments.  The number of nucleotide pairs for
each fragment, determined by co-electrophoresis with PhiX174 RF fragments
produced by endonuclease Z, ranges from 2,050 to 80.  The difference in the
pattern between the large- and small-plaque strains is the disappearance of one
fragment containing approximately 255 nucleotide pairs and the appearance of a
new fragment with 145 nucleotide pairs.  This finding can be explained either
by deletions or insertions totaling 110 nucleotide pairs.  Complementary RNA
synthesized in vitro from the adeno-SV40 hybrid virus, strain ND-1, hybridized
preferentially to four of the fragments of SV40 DNA.

<>

<1>Huang, H.R., Chao, M.Y., Armstrong, B., Wang, Y., Lambowitz, A.M., Perlman, P.S.
<2>The DIVa maturase binding site in the yeast group II intron aI2 is essential for intron homing but not for in vivo splicing.
<3>Mol. Cell. Biol.
<4>23
<5>8809-8819
<6>2003
<7>Splicing of the Saccharomyces cerevisiae mitochondrial DNA group II intron aI2 depends on the
intron-encoded 62-kDa reverse transcriptase-maturase
protein (p62). In wild-type strains, p62 remains associated with the
excised intron lariat RNA in ribonucleoprotein (RNP) particles that are
essential for intron homing. Studies of a bacterial group II intron showed
that the DIVa substructure of intron domain IV is a high-affinity binding
site for its maturase. Here we first present in vitro evidence extending
that conclusion to aI2. Then, experiments with aI2 DIVa mutant strains
show that the binding of p62 to DIVa is not essential for aI2 splicing in
vivo but is essential for homing. Because aI2 splicing in the DIVa mutant
strains remains maturase dependent, splicing must rely on other
RNA-protein contacts. The p62 that accumulates in the mutant strains has
reverse transcriptase activity, but fractionation experiments at high and
low salt concentrations show that it associates more weakly than the
wild-type protein with endogenous mitochondrial RNAs, and that phenotype
probably explains the homing defect. Replacing the DIVa of aI2 with that
of the closely related intron aI1 improves in vivo splicing but not
homing, indicating that DIVa contributes to the specificity of the
maturase-RNA interaction needed for homing.

<>

<1>Huang, J., Wang, H., Liang, W., Xie, X., Guo, G.
<2>Developmental expression of Arabidopsis methyltransferase genes MET1, DRM2, and CMT3.
<3>Mol. Biol. (Mosk)
<4>48
<5>681-687
<6>2014
<7>Cytosine methylation is an epigenetic mark found in the genome of fungi, plants, and animals.
DNA methylation is catalyzed by DNA methyltransferases. The function of DNA methyltransferases
was shown to be highly conserved, but the biological role of these enzymes has not been
clearly defined. We generated transgenic plants expressing METHYLTRANSFERASES::GUS reporter
genes for three major DNA methyltransferases (MET1, DRM2 and CMT3) to gain insight into the
potential physiological relevance of the individual members of the DNA methyltransferase
family in Arabidopsis thaliana, and to investigate their expression patterns in detail. We
found that METHYLTRANSFERASE::GUS genes display unique tissue, cell-type, and temporal
patterns of expression throughout normal development, particularly in the flower. Our findings
are supported by semi-quantitative reverse-transcription PCR, as well as by analyses of
microarray databases. These data suggest that DNA methyltransferases may contribute to
morphogenesis at every developmental stage and in every plant organ.

<>

<1>Huang, J.J., Wang, H.H., Xie, X.J., Zhang, D., Liu, Y., Guo, G.Q.
<2>Roles of DNA methyltransferases in Arabidopsis development.
<3>Afr. J. Biotechnol.
<4>9
<5>8506-8514
<6>2010
<7>DNA methylation plays a vital role during development in gene expression and chromatin
organization. DNA methyl transferases catalyze the transfer of a methyl group to bases within
the DNA helix. Plants differ from animals in having methylation at the sites of CHG and CHH.
In plant, there are at least four classes of cytosine methyltransferase: MET1, CMT3, DRM and
DNMT2. They show distinct expression patterns and levels in tissues and developmental stages
and differential activity on cytosines in different sequence contexts. Mutations that cause
severe loss of DNA methylation often leads to abnormal development. In the present review, we
summarized recent findings of the three major DNA methyltransferases mutants playing vital
role in development of Arabidopsis thaliana.

<>

<1>Huang, K., Ni, J., Xu, K., Tang, H., Tao, F., Xu, P.
<2>Genome Sequence of Sporolactobacillus terrae DSM 11697, the Type Strain of the Species.
<3>Genome Announcements
<4>2
<5>e00465-14
<6>2014
<7>Sporolactobacillus terrae DSM 11697 is the type strain of S. terrae. Here, we present a 3.2-Mb
assembly of its genome sequence. As S. terrae is one of the
important lactic acid bacteria, the genome sequence may provide insights into the
molecular mechanism for its further microbial investigation.

<>

<1>Huang, L.-H., Farnet, C.M., Ehrlich, K.C., Ehrlich, M.
<2>Digestion of highly modified bacteriophage DNA by restriction endonucleases.
<3>Nucleic Acids Res.
<4>10
<5>1579-1591
<6>1982
<7>The ability of thirty Type II restriction endonucleases to cleave five
different types of highly modified DNA has been examined.  The DNA substrates
were derived from relatively large bacteriophage genomes which contain all or
most of the cytosine or thymine residues substitutes at the 5-position.  These
substituents were a proton (PBS1 DNA), a hydroxymethyl group (SP01 DNA), a
methyl group (XP12 DNA), a glucosylated 4,5-dihydroxypentyl group (SP15 DNA).
Although PBS1 DNA and SP01 DNA were digested by most of the enzymes, they were
cleaved much more slowly than was normal DNA by many of them.
5-Methyl-cytosine-rich XP12 DNA and the multiply modified T4 and SP15 DNAs were
resistant to most of these endonucleases.  The only enzyme that cleaved all
five of these DNAs was TaqI, which fragmented them extensively.

<>

<1>Huang, L.H., Farnet, C., Ehrlich, K.C., Ehrlich, M.
<2>Digestion of highly modified phage DNA by restriction endonucleases.
<3>Fed. Proc.
<4>41
<5>1199
<6>1982
<7>The ability of thirty TypeII restriction endonucleases to cleave five different
types of highly modified DNA has been examined.  The substrate DNAs were
derived from relatively large bacteriophage genomes which contain all or most
of the cytosine or thymine residues substituted at the 5-position.  These
substituents were a proton (PBS1 DNA), a hydroxymethyl group (SP01 DNA), a
methyl group (XP12 DNA), a glucosylated hydroxymethyl group (T4 DNA) or a
phosphoglucuronated, glucosylated 4,5-dihydroxypentyl group (SP15 DNA).  While
PBSI DNA and SP01 DNA were cleaved many times by most of the enzymes, they were
cleaved much more slowly than was normal DNA by many of them.
5-Methylcytosine-rich XP12 DNA and the multiply modified T4 and SP15 DNAs were
resistant to most of the enzymes.  The only enzyme which cleaved all of these
DNAs was TaqI; TaqI fragmented the five highly modified DNAs extensively.  The
TaqI fragments from XP12 DNA were susceptible to ligation catalyzed by T4 DNA
ligase.

<>

<1>Huang, N., Banavali, N.K., MacKerell, A.D. Jr.
<2>Protein-facilitated base flipping in DNA by cytosine-5-methyltransferase.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>68-73
<6>2003
<7>DNA methylation, various DNA repair mechanisms, and possibly early events in the opening of
DNA as required for transcription and replication are
initiated by flipping of a DNA base out of the DNA double helix. The
energetics and structural mechanism of base flipping in the presence of
the DNA-processing enzyme, cytosine 5-methyltransferase from HhaI
(M.HhaI), were obtained through molecular dynamics based upon free-energy
calculations. Free-energy profiles for base flipping show that, when in
the closed conformation, M.HhaI lowers the free-energy barrier to flipping
by 17 kcalmol and stabilizes the fully flipped state. Flipping is shown to
occur via the major groove of the DNA. Structural analysis indicates that
flipping is facilitated by destabilization of the DNA double-helical
structure and substitution of DNA base-pairing and base-stacking
interactions with DNA-protein interactions. The fully flipped state is
stabilized by DNA-protein interactions that are enhanced upon binding of
coenzyme. This study represents an atomic detail description of the
mechanism by which a protein facilitates specific structural distortion in
DNA.

<>

<1>Huang, N., MacKerell, A.D.
<2>Specificity in protein-DNA interactions: Energetic recognition by the (cytosine-C5)-methyltransferase from HhaI.
<3>J. Mol. Biol.
<4>345
<5>265-274
<6>2005
<7>Sequence-specific interactions between proteins and DNA are essential for a variety of
biological functions. The
(cytosine-C5)-methyltransferase from HhaI (M.HhaI) specifically
modifies the second base in GCGC sequences, employing a
base flipping mechanism to access the target base being chemically
modified. The mechanism of sequence-specific recognition of M.HhaI is
not evident based on crystallographic structures, leading to the
suggestion that recognition is linked to the flipping event itself, a
process that may be referred to as energetic recognition. Using
computational methods, it is shown that the free energy barriers to
flipping are significantly higher in non-cognate versus the cognate
sequence, supporting the energetic recognition mechanism. Energetic
recognition is imparted by two protein "selectivity filters" that
function via a "web" of protein-DNA interactions in short-lived, high
energy states present along the base flipping pathway. Other
sequence-specific DNA binding proteins whose function involves
significant distortion of DNA's conformation may use a similar
recognition mechanism.

<>

<1>Huang, N., Zou, G., Cao, X., Zhu, R.
<2>Studies on chemical modification and kinetics of restriction endonuclease Bsp78I.
<3>Wuhan Daxue Xuebao
<4>42
<5>233-236
<6>1996
<7>Restriction endonuclease Bsp78I from Bacillus sphaericus 78 has been isolated and purified.
The purified enzyme was found to be homogeneous.  Michaelis constant of the enzyme is 2.67 x
10^-8mol /L (substrate is pBR322 DNA).  The role of specific amino acid residues in Bsp78I was
assayed by chemical modifications.  Sulfhydryl groups were modified with
p-chloromercuribenzoic acid, lysine residues with pyridoxal-5'-phosphate and arginine
residues with 2,3-butanedione.  The results show that these residues are related to the
activity of Bsp78I.

<>

<1>Huang, S.L., Chen, H., Hu, A., Tuan, N.N., Yu, C.P.
<2>Draft Genome Sequence of Pseudomonas nitroreducens Strain TX1, Which Degrades Nonionic Surfactants and Estrogen-Like Alkylphenols.
<3>Genome Announcements
<4>2
<5>e01262-13
<6>2014
<7>Pseudomonas nitroreducens TX1 ATCC PTA-6168 was isolated from rice field drainage in Taiwan.
The bacterium is of special interest because of its capability to use
nonionic surfactants (alkylphenol polyethoxylates) and estrogen-like compounds
(4-t-octylphenol and 4-nonylphenol) as a sole carbon source. This is the first
report on the genome sequence of P. nitroreducens.

<>

<1>Huang, T.W., Chen, F.J., Miu, W.C., Liao, T.L., Lin, A.C., Huang, I.W., Wu, K.M., Tsai, S.F., Chen, Y.T., Lauderdale, T.L.
<2>Complete Genome Sequence of Staphylococcus aureus M013, a pvl-Positive, ST59-SCCmec Type V Strain Isolated in Taiwan.
<3>J. Bacteriol.
<4>194
<5>1256-1257
<6>2012
<7>We report the complete genome sequence of M013, a representative strain of a pvl-positive,
sequence type 59-staphylococcal cassette chromosome mec type V
(ST59-SCCmec type V) community-associated methicillin-resistant Staphylococcus
aureus (CA-MRSA) clone in Taiwan. Comparison of M013 with the genomes of two
CA-MRSA strains in the United States revealed major differences in the regions
covering several genomic islands and prophages.

<>

<1>Huang, T.W., Chen, T.L., Chen, Y.T., Lauderdale, T.L., Liao, T.L., Lee, Y.T., Chen, C.P., Liu, Y.M., Lin, A.C., Chang, Y.H., Wu, K.M., Kirby, R., Lai, J.F., Tan, M.C., Siu, L.K., Chang, C.M., Fung, C.P., Tsai, S.F.
<2>Copy Number Change of the NDM-1 Sequence in a Multidrug-Resistant Klebsiella pneumoniae Clinical Isolate.
<3>PLoS ONE
<4>8
<5>e62774
<6>2013
<7>The genetic features of the antimicrobial resistance of a multidrug resistant Klebsiella
pneumoniae strain harboring blaNDM-1
were investigated to increase our understanding of the evolution of NDM-1. The strain, KPX,
came from a Taiwanese patient with a hospitalization history in New Delhi. Complete DNA
sequencing was performed; and the genes responsible for antimicrobial resistance were
systematically examined and isolated by library screening. KPX harbored two resistance
plasmids, pKPX-1 and pKPX-2, which are 250-kb and 141-kb in size, respectively, with blaNDM-1
present on pKPX-1. The plasmid pKPX-1 contained genes associated with the IncR and IncF
groups, while pKPX-2 belonged to the IncF family. Each
plasmid carried multiple antimicrobial resistance genetic determinants. The gene responsible
for resistance to carbapenems was found on pKPX-1 and that for resistance to aztreonam was
found on pKPX-2. To our surprise, we discovered that blaNDM-1 exists on pKPX-1 as multiple
copies in the form of tandem repeats. Amplification of blaNDM-1 was found to occur by
duplication of an 8.6-kb unit, with the copy number of the repeat varying from colony to
colony. This repeat sequence is identical to that of the pNDM-MAR except for two base
substitutions. The copy number of blaNDM-1 of colonies under
different conditions was assessed by Southern blotting and quantitative PCR. The blaNDM-1
sequence was maintained in the presence of the antimicrobial selection; however, removal of
antimicrobial selection led to the emergence of susceptible bacterial populations with a
reduced copy number or even the complete loss of the blaNDM-1 sequence. The dynamic nature of
the NDM-1 sequence provides a strong argument for judicious use of the broad-spectrum
antimicrobials in order to reduce the development and spread of antimicrobial resistance among
pathogens.

<>

<1>Huang, X., Jiang, W., Zhang, F., Liang, Z.
<2>A simplified method for preparation of restriction endonuclease BamHI.
<3>Zhongguo Yi Xue Ke Xue Yuan Xue Bao
<4>4
<5>60-62
<6>1982
<7>A simplified method for preparation of restriction endonuclease BamHI was described.  With P11
cellulose column chromatography, the exonuclease and nonspecific endonuclease activity in the
crude extract could be removed and the activity of BamHI enriched by 10^5 fold.  The enzyme
preparation thus made might be used for genetic manipulation.

<>

<1>Huang, X.J., Lu, H.L., Wang, J.W., Xu, L.Q., Liu, S.Y., Sun, J.H., Gao, F.
<2>High-throughput sequencing of methylated cytosine enriched by modification-dependent restriction endonuclease MspJI.
<3>BMC Genet.
<4>14
<5>56
<6>2013
<7>Background: As a well-known epigenomic modification, DNA methylation is found to be common in
plants and plays an important role in many
biological processes. Relying on the unique feature of
methylation-dependent digestion, the family of methylation-requiring
restriction-like endonuclease, such as MspJI and its homologs, was
suggested for a potential usage in methylation detection.
Results: In this study, we combine MspJI digestion and
electrophoretic band selection with next generation high-throughput
sequencing technology to detect 5-methylcytosines in Arabidopsis
genome. By developing a bioinformatics workflow to attribute the CNNR
sites recognized by MspJI to the reference genome, we fulfilled the
systematic assessment of this method.
Conclusions: According to the assessment, here we provide the
method for generating a detailed map of plant methylome that could be
feasible, reliable and economical in methylation investigation.

<>

<1>Huang, Y., Friedman, S.
<2>The inhibition of RecA-mediated strand exchange by adducts of azacytosine-containing DNA and EcoRII methylase.
<3>FASEB J.
<4>4
<5>A2294
<6>1990
<7>Recovery of cells from treatment with 5-azacytidine is dependent on the recA,
recBC pathway.  Cells containing DNA (cytosine-5) methylases which form tight
binding complexes with azacytosine containing DNA (azaC-DNA) are more sensitive
to the drug than cells lacking them.  We therefore studied the effect of these
complexes on recA mediated strand exchange in vitro.  32P labelled 422 bp DNA
fragment containing three binding sites for the EcoRII methylase and
azacytosine in the (-) strand was prepared.  We investigated the effect of the
EcoRII methylase on recA mediated strand exchange of the fragment with
homologous M13 DNA by electrophoresis on agarose gels.  In the absence of the
methylase, azaC-DNA has the same rate and extent of strand exchange as control
DNA.  But in the presence of the methylase incorporation of duplexes into
recA-ssDNA complexes is decreased from 80-90% to 10%, and strand exchange is
completely inhibited.  Since there is no incorporation of duplexes with
heterologous acceptor ssDNA, or in the absence of ATP, the incoporation of
azaC-DNA duplexes in the presence of methylase is believed to occur by partial
pairing of duplexes with homologous ssDNA leading to the formation of an
inactive complex composed of the methylase, dsDNA and recA-ssDNA complexes.

<>

<1>Huang, Y., Friedman, S.
<2>Inhibition of recA-mediated strand exchange by adducts of Azacytosine-containing DNA and the EcoRII methylase.
<3>J. Biol. Chem.
<4>266
<5>17424-17429
<6>1991
<7>Wild type Escherichia coli cells containing elevated levels of DNA (cytosine-5)
methyltransferases have increased sensitivity to the toxic effects of
5-azacytidine.  The methyltransferases form tight binding complexes with
azacytosine in DNA which could interfere with the recA recBCD repair pathway
which is largely responsible for cell survival after treatment with the drug.
We therefore determined if these complexes interfered with recA-mediated strand
exchange in vitro, 32P-labeled DNA fragments containing a single EcoRII site,
with cytosine in the (-) strand replaced by 5-azacytosine, were prepared.  We
investigated the effect of the EcoRII methyltransferase on recA-mediated strand
exchange with homologous M13 DNA by electrophoresis in agarose gels.  In the
absence of the methylase the rate and extent of strand exchange of
azacytosine-containing DNA is the same as control DNA.  In the presence of the
methyltransferase strand exchange is inhibited, but some incorporation of
duplexes into recA-single-stranded DNA (ssDNA) complexes still occurs.  The
formation of these complexes is dependent on the length of the fragment 3' to
the methylase binding site on the strand complementary to the ssDNA.  The
greater the length the greater the number of complexes that form.
S-Adenosyl-L-methionine, which enhances binding of the methyltransferase to
azacytosine-containing DNA, causes an increase in the inhibition of strand
exchange and an increase in the number of inactive complexes formed.  The
complexes can be dissociated with guanidinium chloride which denatures the
methyltransferase and leads to release of the (+) strand.  The (-) strand
remains associated with the ssDNA.  This result implies that a plectonemic
joint is formed between recA-ssDNA complexes and azacytosine-containing
DNA-methyltransferase complexes.  However, branch migration in these complexes
is inhibited.  Denaturation of the methyltransferase allows branch migration to
proceed to completion, releasing the (+) strand.

<>

<1>Huang, Y., Jian, J., Lu, Y., Cai, S., Wang, B., Tang, J., Pang, H., Ding, Y., Wu, Z.
<2>Draft Genome Sequence of the Fish Pathogen Vibrio harveyi Strain ZJ0603.
<3>J. Bacteriol.
<4>194
<5>6644-6645
<6>2012
<7>Vibrio harveyi is an important pathogen that causes vibriosis in various aquatic  organisms.
Here, we announce the draft genome sequence of V. harveyi strain
ZJ0603, which was isolated from diseased Orange-spotted grouper (Epinephelus
coioides) in Guangdong, China.

<>

<1>Huang, Y., Li, H., Rensing, C., Zhao, K., Johnstone, L., Wang, G.
<2>Genome Sequence of the Facultative Anaerobic Arsenite-Oxidizing and Nitrate-Reducing Bacterium Acidovorax sp. Strain NO1.
<3>J. Bacteriol.
<4>194
<5>1635-1636
<6>2012
<7>Acidovorax sp. strain NO1, isolated from gold mine soil, was shown to be a facultative
anaerobic arsenite-oxidizing and nitrate-reducing bacterium. The
reported draft genome predicts the presence of genes involved in arsenic
metabolism, nitrate reduction, phosphate transport, and multiple metal
resistances and indicates putative horizontal gene transfer events.

<>

<1>Huang, Y.J., Parker, M.M., Belfort, M.
<2>Role of exonucleolytic degradation in group I intron homing in phage T4.
<3>Genetics
<4>153
<5>1501-1512
<6>1999
<7>Homing of the phage T4 td intron is initiated by the intron-encoded endonuclease I-TevI, which
cleaves the intronless allele 23 and 25 nucleotides upstream of the intron insertion site
(IS). The distance between the I-TevI cleavage site (CS) and IS implicates endo- and/or
exonuclease activities to resect the DNA segment between the IS and CS. Furthermore, 3' tails
must presumably be generated for strand invasion by 5'-3' exonuclease activity. Three
experimental approaches were used to probe for phage nucleases involved in homing: a
comparative analysis of in vivo homing levels of nuclease-deficient phage, an in vitro assay
of nuclease activity and specificity, and a coconversion analysis of flanking exon markers. It
was thereby demonstrated that T4 RNase H, a 5'-3' exonuclease, T4 DNA exonuclease A (DexA)
and the exonuclease activity of T4 DNA polymerase (43Exo), 3'-5' exonucleases, play a role
in intron homing. The absence of these functions impacts not only homing efficiency but also
the extent of degradation and flanking marker coconversion. These results underscore the
critical importance of the 3' tail in intron homing, and they provide the first direct
evidence of a role for 3' single-stranded DNA ends as intermediates in T4 recombination.
Also, the involvement of RNase H, DexA, and 43Exo in homing provides a clear example of the
harnessing of functions variously involved in phage nucleic acid metabolism for intron
propagation.

<>

<1>Hubacek, J.
<2>Functional analysis of second-step host specificity mutations in unstable escherichia coli heterozygotes.
<3>J. Gen. Microbiol.
<4>79
<5>257-264
<6>1973
<7>From an Escherichia coli strain K12 carrying a temperature-sensitive host-specific
modification (hsm) mutation, second-step mutants have been isolated that are completely
deficient in modification and restriction. Complementation analysis has revealed that one
group of these mutants is impaired in the specificity gene hss, while in the other group of
mutant strains both mutations, i.e. the first-step temperature-sensitive and the second-step
which impairs restriction and modification completely, are located in hsm. Analysis of the
heterozygotes used in the complementation experiments suggested a cis- and tandem arrangement
of the hs and leu genes in haploid, segregating exconjugants; however, the attachement of
these genes to a cryptic plasmid was not excluded.

<>

<1>Hubacek, J.
<2>Biological function of DNA methylation.
<3>Folia Microbiol. (Praha)
<4>37
<5>323-329
<6>1992
<7>Structural and functional properties of prokaryotic DNA methyltransferases are summarized. The
different aspects of the role of DNA methylation which influences DNA-protein interaction in
restriction and modification of DNA and in mismatch repair, DNA replication and gene
expression are discussed.

<>

<1>Hubacek, J., Glover, S.W.
<2>Complementation analysis of temperature-sensitive host specificity mutations in Escherichia coli.
<3>J. Mol. Biol.
<4>50
<5>111-127
<6>1970
<7>A selection procedure was devised for the isolation of temperature-sensitive
mutants of Escherichia coli K12 unable to restrict foreign DNA.  Many of the
non-restricting mutants isolated also displayed temperature sensitivity in the
modification of DNA.  The mutations were shown to map in the hs cluster of
genes which determine the host specificity of DNA in E. coli.  The kinetics of
inactivation of restriction showed that a short exposure to high temperature
was sufficient to impair restriction of phage lambda DNA and that after shift
to low temperature restriction did not return to the wild-type level until a
period of growth had occurred.  One of the mutants was used as a starting
strain from which further mutants were then selected for their inability to
host-modify DNA.  Many of the mutants thus isolated, in addition to being
impaired in modification, were found to be non-restricting at both high and low
temperatures.  A complementation analysis of the mutants was carried out using
an F' donor strain derived from E. coli B and carrying the host-specificity
genes hssB+ hsr- hsm+.  In all but one of the F' merodiploids constructed
between this F' and the temperature-sensitive host specificity mutants of E.
coli K the temperature-sensitivity of the mutant phenotype was complemented and
the merodiploids displayed K-and B-specific restriction and K- and B-specific
modification.  From these results it is concluded that all of the
temperature-sensitive mutants carry mutations in the hsm gene, and are hssK+
hsr+ hsmts.  In addition it is argued that an hsm-directed polypeptide is
required for restriction in addition to polypeptides directed by hssK and hsr.
These results are discussed in terms of models based on the interaction of
subunits to form oligomeric enzymes.

<>

<1>Hubacek, J., Holubova, I., Weiserova, M.
<2>The effect of recA mutation on the expression of EcoKI and EcoR124I hsd genes cloned in a multicopy plasmid.
<3>Folia Microbiol. (Praha)
<4>43
<5>353-359
<6>1998
<7>Type I restriction-modification endonucleases are composed of three subunits - HsdR, required
for restriction, and HsdM and HsdS which can produce a separate DNA methyltransferase.  The
HsdS subunit is required for DNA recognition.  In this paper we describe the effect of cloned
EcoKI and EcoR124I hsd genes on the resulting R-M phenotype.  The variability in the
expression of the wild type restriction phenotype after cloning of the wt hsd genes in a
multicopy plasmid in Escherichia coli recA+ background suggests that the increased production
of the restriction endonuclease from pBR322 is detrimental to the cell and this leads to the
deletion of the cloned hsd genes from the hybrid plasmid and/or inactivation of the enzyme.
The effect of a mutation in the E. coli recA gene on the expression of the R-M phenotype is
described and discussed in relation to the role of the cell surface and the localization of
the restriction endonuclease in the cell.

<>

<1>Hubacek, J., Weiserova, M.
<2>DNA restriction and modification in Escherichia coli: Functional analysis of the role of the dnaC(D) gene product.
<3>J. Gen. Microbiol.
<4>119
<5>231-238
<6>1980
<7>Escherichia coli strain PC-7 carries two independent temperature-sensitive mutations, one
affecting the restriction and modification (R-M) phenotype and the other the DNAC(D)
phenotype. The results of complementation and P1 transduction analysis of the mutation
affecting the R-M phenotype implicate a fourth gene, designated hsdX, located close to the hsd
three-gene complex. The properties of merodiploids constructed between appropriate recipients
and F' elements with different mutations in hsdS, hsdR and hsdM genes might indicate that in
strain PC-7 the temperature-sensitive products, determined by hsdR and hsdDK cistrons, are
synthesized. The role of the temperature-sensitive dnaC(D) gene product in the formation of
the restriction endonuclease was studied and no direct relation was found between the DnaC(D)
and R-M phenotypes.

<>

<1>Hubacek, J., Weiserova, M.
<2>Biological function of type I restriction enzymes.
<3>Gene Manipulation and Expression, Croom-Helm, Glass, R.E., Spizek, J., London
<4>0
<5>95-109
<6>1985
<7>Type I restriction enzymes are structurally and functionally so complicated that it was often
argued that they must perform some vital functions other than restriction and modification of
DNA. Using a genetic approach that was found to be so fruitful in the past we analyzed
temperature-sensitive (ts) mutations affecting restriction and modification. The concept of a
gene, designated hsd.X, which seems to be located outside the hsd operon, was formulated and
the role of its product in the regulation of expression of restriction and modification and in
the initiation of DNA replication is discussed. Further, we tried to establish if the
individual subunits of the type I restriction enzyme from E. coli K12 play some role in
mini-Mu and Tn9 transposition mechanisms.

<>

<1>Hubacek, J., Weiserova, M., Janscak, P., Firman, K.
<2>Restriction endonucleases R.EcoKI and R.EcoR124I are probably located in different environments within the bacterial cell.
<3>Folia Microbiol. (Praha)
<4>39
<5>162-165
<6>1994
<7>We describe the phenomenon of a transient state of R124I restriction deficiency after
long-term storage of the E. coli [pCP1005] strain at 4oC, or after growth of the culture in
synthetic M9 medium with the nonmutagenic solvent dimethyl sulfoxide. The unusual high
reversion from the R+124 to the R-124 phenotype was observed only in E. coli strain
transformed with the high-copy number plasmid pCP1005 carrying EcoR124I hsdR, M and S genes
cloned, but not with strains carrying the natural conjugative plasmid R124. The effect of both
treatments on the expression of EcoR124I phenotype in relation to the possible location of
R.EcoR124I restriction endonuclease in E. coli is discussed.

<>

<1>Hubacek, J., Zinkevich, V.E., Weiserova, M.
<2>The location of a temperature-sensitive trans-dominant mutation and its effect on restriction and modification in Escherichia coli K12.
<3>J. Gen. Microbiol.
<4>135
<5>3057-3065
<6>1989
<7>An Escherichia coli K12 chromosomal EcoRI-BamHI fragment containing a mutant
hsdS locus was cloned into plasmid pBR322.  The mcrB gene, closely linked to
hsdS, was used for selection of clones with the inserted fragment using T4alpha
gt57beta gt14 and lambda vir PvuII phages; the phage DNAs contain methylated
cytosines and hence can be used to demonstrate McrB restriction.  For the
efficient expression of the hsdS gene, a BglII fragment of phage lambda
carrying the pR promoter was inserted into the BamHI site of the hybrid
plasmid.  Under these conditions a trans-dominant effect of the hsdXts+d
mutation on restriction and modification was detected.  Inactivation of the
hsdS gene by the insertion of the lambda phage BglII fragment into the BglII
site within this gene resulted in the disappearance of the trans-dominant
effect.  When the cloned BamHI-EcoRI fragment was shortened by HpaI and EcoRI
restriction enzymes, the trans-dominant effect was fully expressed.  The
results indicate that the Xts+d mutation is located in the hsdS gene.  The
effect of gene dosage of the HsdS subunit on the expression of Xts+d mutation
was studied.  The results of complementation experiments, using F'-merodiploids
or plasmid pBR322 with an inserted Xts+d mutation, support the idea that the
HsdSts+d product competes with the wild-type HsdS product, and has a
quantitatively different effect on restriction and modification.

<>

<1>Hubscher, U., Pedrali-Noy, G., Knust-Kron, B., Doerfler, W., Spadari, S.
<2>DNA methyltransferases: Activity minigel analysis and determination with DNA covalently bound to a solid matrix.
<3>Anal. Biochem.
<4>150
<5>442-448
<6>1985
<7>We describe two methods that facilitate detection and characterization of DNA
methyltransferases: activity gel analysis and the use of DNA-cellulose or DNA-Sepharose in DNA
methylation reactions.  The first permits identification of catalytic subunits, determination
of the influence of proteolysis, and evolutionary or developmental studies.  The second allows
accurate and fast determination of DNA methyltransferase activities in crude extracts and
during purification.

<>

<1>Hue, F., Ghalyanchi, L.A., Barbour, A.G.
<2>Chromosome Sequence of Borrelia miyamotoi, an Uncultivable Tick-Borne Agent of Human Infection.
<3>Genome Announcements
<4>1
<5>e00713-13
<6>2013
<7>Borrelia miyamotoi is a newly recognized agent of human disease. B. miyamotoi strain LB-2001,
an isolate from the tick Ixodes scapularis, was propagated in
mice. The sequence of the chromosome was determined by next-generation sequencing
of DNA isolated from whole blood. The sequence established that B. miyamotoi is a
relapsing fever group species.

<>

<1>Huedo, P., Conchillo-Sole, O., Yero, D., Martinez-Servat, S., Daura, X., Gibert, I.
<2>Draft Genome Sequence of Stenotrophomonas maltophilia Strain M30, Isolated from a Chronic Pressure Ulcer in an Elderly Patient.
<3>Genome Announcements
<4>2
<5>e00576-14
<6>2014
<7>Stenotrophomonas maltophilia is an emerging opportunistic pathogen with an increasing
prevalence of multidrug-resistant strains. Here, we report the draft
genome sequence of S. maltophilia strain M30, isolated from a pressure ulcer in
an elderly patient.

<>

<1>Huggett, M.J., Hayakawa, D.H., Rappe, M.S.
<2>Genome sequence of strain HIMB624, a cultured representative from the OM43 clade  of marine Betaproteobacteria.
<3>Standards in Genomic Sciences
<4>6
<5>11-20
<6>2012
<7>Strain HIMB624 is a planktonic marine bacterium within the family Methylophilaceae of the
class Betaproteobacteria isolated from coastal seawater of Oahu, Hawaii. This strain is of
interest because it is one of few known isolates from an abundant clade of Betaproteobacteria
found in cultivation-independent studies of coastal seawater and freshwater environments
around the globe, known as OM43. Here we describe some preliminary features of the organism,
draft genome sequence and annotation, and comparative genomic analysis with one other
sequenced member of this clade (strain HTCC2181). The 1,333,209 bp genome of strain HIMB624 is
arranged in a single scaffold containing four contigs, and contains 1,381 protein encoding
genes and 39 RNA genes.

<>

<1>Hughes, C.A.N., Johnson, R.C.
<2>Methylated DNA in Borrelia species.
<3>J. Bacteriol.
<4>172
<5>6602-6604
<6>1990
<7>The DNA of Borrelia species was examined for the presence of methylated GATC
sequences.  The relapsing-fever Borrelia sp., B. coriaceae, and only 3 of 22
strains of B. burgdorferi contained adenine methylation systems.  B. anserina
lacked an adenine methylation system.  Fundamental differences in DNA
methylation exist among members of the genus Borrelia.

<>

<1>Hughes, S.G.
<2>A map of the cleavage sites for endonuclease AvaI in the chromosome of bacteriophage lambda.
<3>Biochem. J.
<4>163
<5>503-509
<6>1977
<7>The linear order of nine fragments generated by the action of endonuclease AvaI
on the DNA of bacteriophage lambda was determined from the altered
fragmentation patterns of bacteriophages containing known deletions and of
hybrids of bacteriophages lambdaand U80.  Digestion of 5'-terminally
32P-labelled bacteriophage-lambda DNA was used to identify the terminal
fragments.  Measurement of relative fragment lengths permitted rough mapping of
the endonuclease-AvaI cleavage sites relative to the ends of the
bacteriophage-lambda chromosome.  The fragment order was confirmed and the map
refined by analysis of the fragmentation of derivative phages containing single
cleavage sites for endonuclease EcoRI.

<>

<1>Hughes, S.G.
<2>Studies of plasmid encoded restriction and modification systems.
<3>Ph.D. Thesis, University of Edinburgh, Edinburgh, Scotland, , , Edinburgh
<4>
<5>1-70
<6>1977
<7>Within the general context of the distribution, role, and origin, of restriction and
modification systems among plasmids, two plasmid encoded systems have been studied.  The
EcoRII (hspII) system.  The sensitivity of the DNA of bacteriophage lambda to endo R.EcoRII in
vitro was found to depend on the presence or absence of methylation introduced by the cytosine
specific DNA methylase of E. coli K.  Subsequently, a revised minimum estimate for the number
of cleavage sites for endo R.EcoRII and a map of these sites close to the ends of the lambda
chromosome were made.  The EcoR124 (hspI) system.  The restriction and modification system
carried by the plasmid R124 was shown to be different from the EcoRI system carried by plasmid
RY5 (subsequently NTP13) with which it had previously been assumed to be identical.  Endo
R.EcoR124 was isolated and found to be dependent on ATP and S-adenosylmethionine (it is a
class I restriction endonuclease).  A derivative of R124 was isolated, by chance, which has
acquired a restriction and modification system of novel specificity.  The derivative, R124/3,
also has new cleavage sites for endo R.EcoRI and endo R.SalI in its DNA.  It is proposed that
the new system, EcoR124/3 arose by recombination between the determinants of EcoK and EcoR124.

<>

<1>Hughes, S.G., Bruce, T., Murray, K.
<2>The Isolation and Characterization of a Sequence-Specific Endonuclease from Anabaena subcylindrica.
<3>Biochem. J.
<4>185
<5>59-63
<6>1980
<7>An endonuclease, AsuI, was isolated from extracts of Anabaena subcylindrica on
the basis of gel-electrophoretic analysis of digests of bacteriophage-lambda
DNA with the partially purified extracts.  The enzyme requires Mg2+, but no
other cofactors.  Endonuclease AsuI recognizes the interrupted tetranucleotide
sequence: 5'-G-^G-N-C-C-3'   -C-C-N-G^-G- and breaks the phosphodiester bonds
indicated by the arrows to leave single-stranded trinucleotide projections at
the 5'-termini of the DNA fragments.

<>

<1>Hughes, S.G., Hattman, S.
<2>The sensitivity of bacteriophage lambda DNA to restriction endonuclease RII.
<3>J. Mol. Biol.
<4>98
<5>645-647
<6>1975
<7>Analysis of fragments of bacteriophage DNA produced by digestion with purified restriction
endonuclease RII shows that DNA propagated in the presence of the cytosine-specific DNA
methylase of Escherichia coli K is partially protected.  It is concluded that a fraction of
the recognition sequences for EcoRII are methylated by this methylase which shares an element
of sequence specificity with the RII restriction and modification enzymes.

<>

<1>Hughes, S.G., Murray, K.
<2>The Nucleotide Sequences Recognized by Endonucleases AvaI and AvaII from Anabaena variabilis.
<3>Biochem. J.
<4>185
<5>65-75
<6>1980
<7>Determination of the 5'-terminal sequences flanking all the individual cleavage
sites for endonuclease AvaI in bacteriophage-lambda DNA has shown that this
enzyme recognizes the hexanucleotide sequence: 5'-C-^Y-C-G-R-G-3'
G-R-G-C-Y-^C This sequence is cut as shown by the arrows to give
single-stranded 5'-tetranucleotide protrusions (cohesive ends).  Endonucleases
SmaI, XhoI and XmaI recognize different symmetrical subsets of this sequence
and provide independent evidence for the occurrence of these subsets at
particular endonuclease-AvaI cleavage sites in the bacteriophage-lambda genome.
Further evidence for this structure came from the demonstration that DNA
fragments generated by endonuclease AvaI can be ligated to form a discrete set
of larger molecules and from nearest-neighbour analysis which showed that
cytosine residues occurred at the 3'-side of cleavage points.  The observation
that endonuclease AvaII recognized a subset of the sites recognized by AsuI
[Hughes, Bruce & Murray (1979) Biochem. J. 185, 59-63] led to the deduction
that AvaII recognizes the pentanucleotide sequence: 5'-G-^G-A-C-C-3'
C-C-T-G-^G and breaks internucleotide bonds at the positions indicated by the
arrows.

<>

<1>Hugon, P., Mishra, A.K., Lagier, J.C., Nguyen, T.T., Couderc, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Brevibacillus massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>8
<5>1-14
<6>2013
<7>Brevibacillus massiliensis strain phR(T) sp. nov. is the type strain of B. massiliensis sp.
nov., a new species within the genus Brevibacillus. This strain
was isolated from the fecal flora of a woman suffering from morbid obesity. B.
massiliensis is a Gram-positive aerobic rod-shaped bacterium. Here we describe
the features of this organism, together with the complete genome sequence and
annotation. The 5,051,018 bp long genome (1 chromosome but no plasmid) contains
5,051 protein-coding and 84 RNA genes, and exhibits a G+C content of 53.1%.

<>

<1>Hugon, P., Mishra, A.K., Robert, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Anaerococcus vaginalis.
<3>Standards in Genomic Sciences
<4>6
<5>356-365
<6>2012
<7>We report the properties of a draft genome sequence of the bacterium Anaerococcus vaginalis
strain PH9, a species within the Anaerococcus genus. This strain, whose
genome is described here, was isolated from the fecal flora of a 26-year-old
woman suffering from morbid obesity. A. vaginalis is an obligate anaerobic
coccus. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 2,048,125-bp long (one chromosome
but no plasmid) and contains 2,095 protein-coding and 38 RNA genes, including
three rRNA genes.

<>

<1>Hugon, P., Ramasamy, D., Lagier, J.C., Rivet, R., Couderc, C., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Alistipes obesi sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>427-439
<6>2013
<7>Alistipes obesi sp. nov. strain ph8(T) is the type strain of A. obesi, a new species within
the genus Alistipes. This strain, whose genome is described here,
was isolated from the fecal flora of a 26-year-old woman suffering from morbid
obesity. A. obesi is an obligately anaerobic rod. Here we describe the features
of this organism, together with the complete genome sequence and annotation. The
3,162,233 bp long genome (1 chromosome but no plasmid) contains 2,623
protein-coding and 49 RNA genes, including three rRNA genes.

<>

<1>Hugon, P., Ramasamy, D., Robert, C., Couderc, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Kallipyga massiliensis gen. nov., sp. nov., a new member of the family Clostridiales Incertae Sedis XI.
<3>Standards in Genomic Sciences
<4>8
<5>500-515
<6>2013
<7>Kallipyga massiliensis strain ph2(T) is the type strain of Kallipyga massiliensis gen. nov.,
sp. nov., the type species of the new genus Kallipyga within the
family Clostridiales Incertae Sedis XI. This strain, whose genome is described
here, was isolated from the fecal flora of a 26-year-old woman suffering from
morbid obesity. K. massiliensis is an obligate anaerobic coccus. Here we describe
the features of this organism, together with the complete genome sequence and
annotation. The 1,770,679 bp long genome (1 chromosome but no plasmid) contains
1,575 protein-coding and 50 RNA genes, including 4 rRNA genes.

<>

<1>Huguet-Tapia, J.C., Loria, R.
<2>Draft Genome Sequence of Streptomyces acidiscabies 84-104, an Emergent Plant Pathogen.
<3>J. Bacteriol.
<4>194
<5>1847
<6>2012
<7>A draft genome sequence of the plant pathogen Streptomyces acidiscabies 84-104, an emergent
plant pathogen, is presented here. The genome is among the largest of
streptomycetes, at more than 11 Mb, and encodes a 100-kb pathogenicity island
(PAI) shared with other plant-pathogenic streptomycetes. The presence of this
conserved PAI, and the remnants of a conserved integrase/recombinase at its 3'
end, supports the hypothesis that S. acidiscabies emerged as a plant pathogen as
a result of this acquisition.

<>

<1>Hui, R.K., Chen, J.W., Chan, K.G., Leung, F.C.
<2>Complete Genome Sequence of Dyella japonica Strain A8 Isolated from Malaysian Tropical Soil.
<3>Genome Announcements
<4>2
<5>e01164-14
<6>2014
<7>We previously identified and presented the draft genome of a Xanthomonadaceae bacterial strain
Dyella japonica A8 which shows quorum-quenching activity. Here,
we report the complete, closed genome sequence of this bacterium. This complete
genome may help to further investigate the comparative quorum-quenching activity
among D. japonica strains.

<>

<1>Huletsky, A., Giroux, R.
<2>Sequences for detection and identification of methicillin-resistant Staphylococcus Aureus (MRSA) of MREJ type xiv.
<3>European Patent Office
<4>EP 2322930 A
<5>
<6>2011
<7>Described herin are novel SCCmec right extremity junction (MREJ) sequences for the detection
and/or identification of methicillin-resistant Staphylococcus aureus (MRSA). Disclosed are
methods and compositions based on DNA sequences for the specific detection of MREJ sequences
designated types xi, xii, xiii, xiv, xv, xvi, xvii, xviii, xix, and xx for diagnostic purposes
and/or epidemiological typing.

<>

<1>Huletsky, A., Rossbach, V.
<2>Sequences for detection and identification of methicillin-resistant Staphyloccocus aureus.
<3>International Patent Office
<4>WO 02099034 A
<5>
<6>2002
<7>The present invention describes novel SCCmec right extremity junction sequences for the
detection of methicillin-resistant Staphylococcus aureus (MRSA).  It relates to the use of
these DNA sequences for diagnostic purposes.

<>

<1>Huletsky, A., Rossbach, V.
<2>Methods for detection and identification of methicillin-resistant Staphylococcus aureus.
<3>US Patent Office
<4>US 7449289 B
<5>
<6>2008
<7>The present invention describes novel SCCmec right extremity junction sequences for the
detection of methicillin-resistant Staphylococcus aureus (MRSA).  It relates to the use of
these DNA sequences for diagnostic purposes.

<>

<1>Huletsky, A., Rossbach, V.
<2>Methods and sequences for detection and identification of methicillin-resistant Staphylococcus aureus.
<3>European Patent Office
<4>EP 2236621 A
<5>
<6>2010
<7>The present invention describes novel SCCmecright extremity junction sequences for the
detection of methicillin-resistant Staphylococcus aureus.  It relates to the use of these DNA
sequences for diagnostic purposes.

<>

<1>Huletsky, A., Rossbach, V.
<2>Methods and sequences for the detection and identification of methicillin-resistant Staphylococcus aureus MREJ type viii strains.
<3>European Patent Office
<4>EP 2322655 A
<5>
<6>2011
<7>The present invention describes novel SCCmec right extremity junction sequences for the
detection of methicillin-resistant Staphylococcus aureus (MRSA). It relates to the use of
these DNA sequences for diagnostic purposes.

<>

<1>Huletsky, A., Rossbach, V.
<2>Methods and sequences for the detection and identification of methicillin-resistant Staphylococcus aureus MREJ type ix strains.
<3>European Patent Office
<4>EP 2322661 A
<5>
<6>2011
<7>The present invention describes novel SCCmec right extremity junction sequences for the
detection of methicillin-resistant Staphylococcus aureus (MRSA).  It relates to the use of
these DNA sequences for diagnostic purposes.

<>

<1>Huletsky, A., Rossbach, V.
<2>Methods and sequences for the detection and identification of methicillin-resistant Staphylococcus aureus MREJ type v strains.
<3>European Patent Office
<4>EP 2322663 A
<5>
<6>2011
<7>The present invention describes novel SCCmec right extremity junction sequences for the
detection of methicillin-resistant Staphylococcus aureus (MRSA).  It relates to the use of
these DNA sequences for diagnostic purposes.

<>

<1>Huletsky, A., Rossbach, V.
<2>Methods and sequences for the detection and identification of methicillin-resistant Staphylococcus aureus MREJ type vi strains.
<3>European Patent Office
<4>EP 2322664 A
<5>
<6>2011
<7>The present invention describes novel SCCmec right extremity junction sequences for the
detection of methicillin-resistant Staphyloccus aureus (MRSA).  It relates to the use of these
DNA sequences for diagnostic purposes.

<>

<1>Hulsmann, K.-H., Quaas, R., Georgalis, Y., Saenger, W., Hahn, U.
<2>High-level expression of a semisynthetic dam gene in Escherichia coli.
<3>Gene
<4>98
<5>83-88
<6>1991
<7>We constructed a semisynthetic gene encoding a DNA-adenine-methyltransferase (Dam) that codes
for the same amino acid sequence as the wild type (wt) Escherichia coli dam gene. Since for
unknown reasons the entire wt sequence, from the start codon to the end of the gene, could not
be cloned, a gene was constructed consisting of a chemically synthesized 5' portion and a 3'
portion from the E. coli chromosome. Introduction of this semisynthetic gene into a suitable
vector allows overproduction of E. coli Dam in mg amounts per liter E. coli culture, with
optimum expression of the gene in the vector pJLA503. This plasmid places the target gene
under control of the strong, tandemly arranged PR PL promoters from bacteriophage lambda,
regulated by a temperature-sensitive lambda repressor. A rapid, two-column purification
protocol is described that allows for very fast purification of the protein. The 32-kDa
recombinant protein methylates the sequence GATC.

<>

<1>Humann, J.L., Wildung, M., Cheng, C.-H., Lee, T., Drew, J.C., Triplett, E.W., Main, D., Schroeder, B.K.
<2>Complete genome of the onion pathogen Enterobacter cloacae EcWSU1.
<3>Standards in Genomic Sciences
<4>5
<5>279-286
<6>2011
<7>
<>

<1>Humann, J.L., Wildung, M., Pouchnik, D., Bates, A.A., Drew, J.C., Zipperer, U.N., Triplett, E.W., Main, D., Schroeder, B.K.
<2>Complete genome of the switchgrass endophyte Enterobacter clocace P101.
<3>Standards in Genomic Sciences
<4>9
<5>726-734
<6>2014
<7>The Enterobacter cloacae complex is genetically very diverse. The increasing number of
complete genomic sequences of E. cloacae is helping to determine the
exact relationship among members of the complex. E. cloacae P101 is an endophyte
of switchgrass (Panicum virgatum) and is closely related to other E. cloacae
strains isolated from plants. The P101 genome consists of a 5,369,929 bp
chromosome. The chromosome has 5,164 protein-coding regions, 100 tRNA sequences,
and 8 rRNA operons.

<>

<1>Humbelin, M., Suri, B., Rao, D.N., Hornby, D.P., Eberle, H., Pripfl, T., Kenel, S., Bickle, T.A.
<2>Type III DNA restriction and modification systems EcoP1 and EcoP15.
<3>J. Mol. Biol.
<4>200
<5>23-29
<6>1988
<7>This paper presents the nucleotide sequence of the mod-res operon of phage P1,
which encodes the two structural genes for the EcoP1 type III restriction and
modification system.  We have also sequenced the mod gene of the allelic EcoP15
system.  The mod gene product is responsible for binding the system-specific
DNA recognition sequences in both restriction and modification:   it also
catalyses the modification reaction.  A comparison of the two mod gene product
sequences shows that they have conserved amino and carboxyl ends but have
completely different sequences in the middle of the molecules.  Two alleles of
the EcoP1 mod gene that are defective in modification but not in restriction
were also sequenced.  The mutations in both alleles lie within the
non-conserved regions.

<>

<1>Humbert, O., Dorer, M.S., Salama, N.R.
<2>Characterization of Helicobacter pylori factors that control transformation frequency and integration length during inter-strain DNA  recombination.
<3>Mol. Microbiol.
<4>79
<5>387-401
<6>2011
<7>P>Helicobacter pylori is a genetically diverse bacterial species, owing in part to its natural
competence for DNA uptake that facilitates
recombination between strains. Inter-strain DNA recombination occurs
during human infection and the H. pylori genome is in linkage
equilibrium worldwide. Despite this high propensity for DNA exchange,
little is known about the factors that limit the extent of
recombination during natural transformation. Here, we identify
restriction-modification (R-M) systems as a barrier to transformation
with homeologous DNA and find that R-M systems and several components
of the recombination machinery control integration length. Type II R-M
systems, the nuclease nucT and resolvase ruvC reduced integration
length whereas the helicase recG increased it. In addition, we
characterized a new factor that promotes natural transformation in H.
pylori, dprB. Although free recombination has been widely observed in
H. pylori, our study suggests that this bacterium uses multiple systems
to limit inter-strain recombination.

<>

<1>Humbert, O., Salama, N.R.
<2>Functional characterization of HP0503: An adenine methyltransferase required for the virulence of Helicobacter pylori.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>107
<5>39
<6>2007
<7>Restriction-modification systems have traditionally been implicated in the protection of the
bacterial genome from invading DNAs, but accumulating evidence suggests that they have other
cellular functions. Helicobacter pylori contains a large number of these systems with at least
23 identified. In silico genome analysis reveals that in many of these systems, the
restriction-endonuclease component is inactive but the cognate methyltransferase retains full
activity, implying that it confers a selective advantage to its host. In an effort to identify
H. pylori factors required for the colonization of the mouse stomach, we identified HP0503, a
protein of unknown function. Structure prediction algorithms revealed that HP0503 has a
similar folding pattern to the adenine methyltransferase M.TaqI from Thermus aquaticus,
leading us to hypothesize that HP0503 is a DNA methyltransferase required for the virulence of
H. pylori. The methyltransferase activity of HP0503 was demonstrated using an engineered
strain of E. coli defective in DNA methylation. Upon expression of HP0503 in this E. coli
strain, genomic DNA methylation could be detected using antibodies recognizing methylated
adenine. In addition, a point mutation in the predicted catalytic site of HP0503 completely
abolished adenine methylation, further advocating its function as an adenine
methyltransferase. To clarify the role of HP0503 in virulence, its DNA recognition site was
determined. The restriction endonuclease Rsa I could digest genomic DNA from a HP0503 mutant
strain but not WT, indicating that HP0503 acts on the same sequence as Rsa I, GTAC. This
finding was confirmed by expressing HP0503 cognate endonuclease, HP0505, in E. coli, which
cleaved DNA at GTAC sites. Interestingly, while GTAC sites are strongly avoided in the H.
pylori genome (only 115 to 184 sites/genome), 16 of these sites are found within the 2 copies
of the 3kbp DNA fragment encoding the 23S rRNA. We are currently testing the methylation
status of HP0503 cognate sites and examining their impact on gene activity to probe the
mechanism by which HP0503 confers a fitness advantage during stomach colonization.

<>

<1>Humbert, O., Salama, N.R.
<2>The Helicobacter pylori HpyAXII restriction-modification system limits exogenous DNA uptake by targeting GTAC sites but shows asymmetric  conservation of the DNA methyltransferase and restriction endonuclease  components.
<3>Nucleic Acids Res.
<4>36
<5>6893-6906
<6>2008
<7>The naturally competent organism Helicobacter pylori encodes a large number of
restriction-modification (R-M) systems that consist of a
restriction endonuclease and a DNA methyltransferase. R-M systems are not
only believed to limit DNA exchange among bacteria but may also have other
cellular functions. We report a previously uncharacterized H. pylori type
II R-M system, M.HpyAXII/R.HpyAXII. We show that this system targets GTAC
sites, which are rare in the H. pylori chromosome but numerous in
ribosomal RNA genes. As predicted, this type II R-M system showed
attributes of a selfish element. Deletion of the methyltransferase
M.HpyAXII is lethal when associated with an active endonuclease R.HpyAXII
unless compensated by adaptive mutation or gene amplification. R.HpyAXII
effectively restricted both unmethylated plasmid and chromosomal DNA
during natural transformation and was predicted to belong to the novel
'half pipe' structural family of endonucleases. Analysis of a panel of
clinical isolates revealed that R.HpyAXII was functional in a small number
of H. pylori strains (18.9%, n = 37), whereas the activity of M.HpyAXII
was highly conserved (92%, n = 50), suggesting that GTAC methylation
confers a selective advantage to H. pylori. However, M.HpyAXII activity
did not enhance H. pylori fitness during stomach colonization of a mouse
infection model.

<>

<1>Humbert, O.M.
<2>Restriction-modification systems and the regulation of genetic exchange in Helicobacter pylori.
<3>Ph.D. Thesis, University of Washington, Seattle, USA
<4>
<5>
<6>2010
<7>Helicobacter pylori is a gram negative bacterial pathogen that colonizes the stomach of over
half of the world population. Various pathologies are associated with H. pylori infection and
include gastritis, peptic ulcers, gastric cancer and MALT lymphoma. H pylori is one of the
most genetically diverse bacterial species known, a likely consequence of DNA recombination
between H. pylori strains facilitated by
their natural competence for DNA uptake. Inter-strain DNA recombination occurs in patients
infected with multiple genetically distinct H. pylori isolates but the barriers that limit DNA
exchange is this organism are not understood. Restriction-Modification (R-M) systems are made
of a restriction endonuclease and a DNA methyltransferase, both of which target the same DNA
sequence. These systems are generally believed to protect bacteria from bacteriophages
invasion but their role in limiting inter-strain recombination remains unexplored. Over the
course of my Thesis work, I first identified and characterized a novel H. pylori R-M system. I
showed that
this system targets the sequence GTAC and that it effectively restricts unmethylated DNA
during natural transformation. After designing a novel genetic assay to quickly assess
restriction endonuclease activity, I made the intriguing observation that the DNA
methyltransferase activity is highly conserved as compared to the endonuclease activity in H.
pylori, and suggested that DNA methylation has alternate roles in this
organism. In the following chapter, I quantified the size of DNA exchanged during H. pylori
inter-strain recombination and found that R-M systems reduce both recombination frequency and
size. Several components of the DNA transformation and recombination pathways were also
identified as regulators of transformation frequency and integration size during both
homologous and homeologous DNA transformation. In the last chapter, I investigated whether DNA
methylation controls gene transcription in H. pylori. Global transcriptional profiles of H.
pylori bacteria were analyzed by microarrays in response to adenine methylation of the
recognition sequences GTAC and TCGA. Transcriptional changes were observed in response to DNA
methylation, but no correlation could be established with the distribution of methylation
sites. Possible mechanisms of regulation by DNA methylation were considered.

<>

<1>Humeny, A., Beck, C., Becker, C.M., Jeltsch, A.
<2>Detection and analysis of enzymatic DNA methylation of oligonucleotide substrates by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.
<3>Anal. Biochem.
<4>313
<5>160-166
<6>2003
<7>Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF) mass
spectrometry was employed to analyze DNA
methylation carried out by the Escherichia coli dam DNA methyltransferase
using oligonucleotide substrates with molecular masses of 5000-10,000 Da
per strand. The mass spectrometry assay offers several advantages: (i) it
directly shows the methylation as the increase in the mass of the
substrate DNA, (ii) it is nonradioactive, (iii) it is quantitative, and
(iv) it can be automated for high-throughput applications. Since
unmethylated and methylated DNA are detected, the ratio of methylation can
be determined directly and accurately. Furthermore, the assay allows
detection individually of the methylation of several substrates in
competition, offering an ideal setup to analyze the specificity of DNA
interacting with enzymes. We could not identify methylation at any
noncanonical site, indicating that the dam MTase is a very specific
enzyme. Finally, MALDI-TOF mass spectrometry permitted assessment of the
number of methyl groups incorporated into each DNA strand, thereby,
allowing study of mechanistic details such as the processivity of the
methylation reaction. We provide evidence that the dam MTase modifies DNA
in a processive reaction, confirming earlier findings.

<>

<1>Humphries, P., Gordon, R.L., McConnell, D.J., Connolly, P.
<2>Endonuclease R. Hind fragments of T7 DNA.
<3>Virology
<4>58
<5>25-31
<6>1974
<7>Restriction enzyme endonuclease R. Hind of Haemophilus influenzae strain Rd,
prepared by a rapid new method, was used to cleave bacteriophage T7 DNA into a
minimum of 49 unique fragments which were separated on polyacrylamide gels.
The fragments were estimated to fall within the size range of 100-200
nucleotide pairs.

<>

<1>Hung, J.E., Mill, C.P., Clifton, S.W., Magrini, V., Bhide, K., Francois, J.A., Ransome, A.E., Fulton, L., Thimmapuram, J., Wilson, R.K., Kappock, T.J.
<2>Draft Genome Sequence of Acetobacter aceti Strain 1023, a Vinegar Factory Isolate.
<3>Genome Announcements
<4>2
<5>e00550-14
<6>2014
<7>The genome sequence of Acetobacter aceti 1023, an acetic acid bacterium adapted to traditional
vinegar fermentation, comprises 3.0 Mb (chromosome plus plasmids).
A. aceti 1023 is closely related to the cocoa fermenter Acetobacter pasteurianus
386B but possesses many additional insertion sequence elements.

<>

<1>Hung, M.S., Karthikeyan, N., Huang, B., Koo, H.C., Kiger, J., Shen, C.J.
<2>Drosophila proteins related to vertebrate DNA (5-cytosine) methyltransferases.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>11940-11945
<6>1999
<7>DNA methylation at CpG residues is closely associated with a number of biological processes
during vertebrate development. Unlike the vertebrates, however, several invertebrate species,
including the Drosophila, do not have apparent DNA methylation in their genomes. Nor have
there been reports on a DNA (5-cytosine) methyltransferase (CpG MTase) found in these
invertebrates. We now present evidence for two CpG MTase-like proteins expressed in Drosophila
cells. One of these, DmMTR1, is a protein containing peptide epitopes immunologically related
to the conserved motifs I and IV in the catalytic domain of the mammalian dnmt1. DmMTR1 has an
apparent molecular mass of 220 kDa and, similar to mammalian dnmt1, it also interacts in vivo
with the proliferating cell nuclear antigen. During interphase of the syncytial Drosophila
embryos, the DmMTR1 molecules are located outside the nuclei, as is dnmt1 in the mouse
blastocyst. However, DmMTR1 appears to be rapidly transported into, and then out of the nuclei
again, as the embryos undergo mitotic waves. Immunofluorescent data indicate that DmMTR1
molecules "paint" the whole set of condensed Drosophila chromosomes throughout the mitotic
phase, suggesting they may play an essential function in the cell-cycle regulated condensation
of the Drosophila chromosomes. Through search in the genomic database, we also have identified
a Drosophila polypeptide, DmMT2, that exhibits high sequence homology to the mammalian dnmt2
and the yeast CpG MTase homolog pmt1. The expression of DmMT2 appears to be developmentally
regulated. We discuss the evolutionary and functional implications of the discovery of these
two Drosophila proteins related to mammalian CpG MTases.

<>

<1>Huntemann, M. et al.
<2>Complete genome sequence of the thermophilic sulfur-reducer Hippea maritima type  strain (MH(2)).
<3>Standards in Genomic Sciences
<4>4
<5>303-311
<6>2011
<7>Hippea maritima (Miroshnichenko et al. 1999) is the type species of the genus Hippea, which
belongs to the family Desulfurellaceae within the class
Deltaproteobacteria. The anaerobic, moderately thermophilic marine sulfur-reducer
was first isolated from shallow-water hot vents in Matipur Harbor, Papua New
Guinea. H. maritima was of interest for genome sequencing because of its isolated
phylogenetic location, as a distant next neighbor of the genus Desulfurella.
Strain MH(2) (T) is the first type strain from the order Desulfurellales with a
completely sequenced genome. The 1,694,430 bp long linear genome with its 1,723
protein-coding and 57 RNA genes consists of one circular chromosome and is a part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Huntemann, M. et al.
<2>Complete genome sequence of the facultatively anaerobic, appendaged bacterium Muricauda ruestringensis type strain (B1(T)).
<3>Standards in Genomic Sciences
<4>6
<5>185-193
<6>2012
<7>Muricauda ruestringensis Bruns et al. 2001 is the type species of the genus Muricauda, which
belongs to the family Flavobacteriaceae in the phylum
Bacteroidetes. The species is of interest because of its isolated position in the
genomically unexplored genus Muricauda, which is located in a part of the tree of
life containing not many organisms with sequenced genomes. The genome, which
consists of a circular chromosome of 3,842,422 bp length with a total of 3,478
protein-coding and 47 RNA genes, is a part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Huntemann, M. et al.
<2>Genome sequence of the phylogenetically isolated spirochete Leptonema illini type strain (3055(T)).
<3>Standards in Genomic Sciences
<4>8
<5>177-187
<6>2013
<7>Leptonema illini Hovind-Hougen 1979 is the type species of the genus Leptonema, family
Leptospiraceae, phylum Spirochaetes. Organisms of this family have a
Gram-negative-like cell envelope consisting of a cytoplasmic membrane and an
outer membrane. The peptidoglycan layer is associated with the cytoplasmic rather
than the outer membrane. The two flagella of members of Leptospiraceae extend
from the cytoplasmic membrane at the ends of the bacteria into the periplasmic
space and are necessary for their motility. Here we describe the features of the
L. illini type strain, together with the complete genome sequence, and
annotation. This is the first genome sequence (finished at the level of Improved
High Quality Draft) to be reported from of a member of the genus Leptonema and a
representative of the third genus of the family Leptospiraceae for which complete
or draft genome sequences are now available. The three scaffolds of the 4,522,760
bp draft genome sequence reported here, and its 4,230 protein-coding and 47 RNA
genes are part of the G enomic E ncyclopedia of Bacteria and Archaea project.

<>

<1>Hunter, S.S., Yano, H., Loftie-Eaton, W., Hughes, J., De Gelder, L., Stragier, P., De Vos, P., Settles, M.L., Top, E.M.
<2>Draft Genome Sequence of Pseudomonas moraviensis R28-S.
<3>Genome Announcements
<4>2
<5>e00035-14
<6>2014
<7>We report the draft genome sequence of Pseudomonas moraviensis R28-S, isolated from the
municipal wastewater treatment plant of Moscow, ID. The strain carries a
native mercury resistance plasmid, poorly maintains introduced IncP-1 antibiotic
resistance plasmids, and has been useful for studying the evolution of plasmid
host range and stability.

<>

<1>Huntley, S., Kneip, S., Treuner-Lange, A., Sogaard-Andersen, L.
<2>Complete Genome Sequence of Myxococcus stipitatus Strain DSM 14675, a Fruiting Myxobacterium.
<3>Genome Announcements
<4>1
<5>e0010013
<6>2013
<7>Hallmarks of the myxobacteria include the formation of spore-filled fruiting bodies in
response to starvation and synthesis of secondary metabolites.
Myxococcus stipitatus forms morphologically highly distinct fruiting bodies and
produces secondary metabolites with antibiotic or cytotoxic activities. Here, we
present the 10.35-Mb genome sequence of M. stipitatus strain DSM 14675.

<>

<1>Huntley, S., Zhang, Y., Treuner-Lange, A., Kneip, S., Sensen, C.W., Sogaard-Andersen, L.
<2>Complete Genome Sequence of the Fruiting Myxobacterium Corallococcus coralloides  DSM 2259.
<3>J. Bacteriol.
<4>194
<5>3012-3013
<6>2012
<7>Corallococcus coralloides, like most other myxobacteria, undergoes a developmental program
culminating in the formation of fruiting bodies. C.
coralloides fruiting bodies are morphologically distinct from those of other
fruiting myxobacteria for which full-length genome sequences are available. The
genome sequence of the 10.0-Mb C. coralloides genome is presented herein.

<>

<1>Hunyadkurti, J., Feltoti, Z., Horvath, B., Nagymihaly, M., Voros, A., McDowell, A., Patrick, S., Urban, E., Nagy, I.
<2>Complete Genome Sequence of Propionibacterium acnes Type IB strain 6609.
<3>J. Bacteriol.
<4>193
<5>4561-4562
<6>2011
<7>Propionibacterium acnes (P. acnes) is an anaerobic Gram-positive bacterium that forms part of
the normal human cutaneous microbiota and is thought to
play central role in acne vulgaris, a chronic inflammatory disease of the
pilosebaceous unit (8). Here we present the whole genome sequence for the
P. acnes Type IB strain 6609 isolated from a patient with inflammatory
acne (15).

<>

<1>Huo, Y., Li, P.
<2>Separation and purification of restriction endonuclease MspI.
<3>Shengwu Huaxue Yu Shengwu Wuli Jinzhan
<4>16
<5>235-236
<6>1989
<7>None

<>

<1>Huo, Y.-Y., Cheng, H., Han, X.-F., Jiang, X.-W., Sun, C., Zhang, X.-Q., Zhu, X.-F., Liu, Y.-F., Li, P.-F., Ni, P.-X., Wu, M.
<2>Complete genome sequence of Pelagibacterium halotolerans B2T.
<3>J. Bacteriol.
<4>194
<5>197-198
<6>2011
<7>Pelagibacterium halotolerans B2T is a marine halotolerant bacterium that was isolated from a
seawater sample collected from the East China Sea. Here, we present the complete genome
sequence of the type strain P. halotolerans B2T, which consists of one chromosome (3,944,837
bp; 61.4% G_C content) and one plasmid (4,050 bp; 56.1% G_C content). This is the first
complete genome of a member of the Pelagibacterium genus.

<>

<1>Hupkes, M., Azevedo, R., Jansen, H., van Zoelen, E.J., Dechering, K.J.
<2>Identification of Novel Bacterial M.SssI DNA Methyltransferase Inhibitors.
<3>J. Biomol. Screen.
<4>18
<5>348-355
<6>2013
<7>DNA methylation is an important epigenetic regulator of gene expression. Abnormalities in DNA
methylation patterns have been
associated with various developmental and proliferative diseases,
particularly cancer. Targeting DNA methyltransferases (DNMTs)
represents a promising strategy for the treatment of such diseases.
Current DNMT inhibitors suffer important drawbacks with respect to
their efficacy, specificity, and toxicity. In this study, we have set
up a robust in vitro bacterial M.SssI DNMT activity assay to
systematically screen a collection of 26 240 compounds that were
predicted to compete with the S-adenosyl-L-methionine (SAM) substrate
of DNMT. This resulted in the identification of a novel set of
structurally distinct inhibitors of M.SssI DNMT activity. Although
molecular docking studies using an M.SssI homology model suggest that
these compounds might compete with SAM binding, mode of activity (MoA)
assays are still needed to confirm this hypothesis. Our set of novel
M.SssI DNMT inhibitors, once confirmed in an orthogonal DNMT assay, may
thus serve as a starting point to identify and characterize suitable
lead candidates for further drug optimization.

<>

<1>Hurd, P.J., Whitmarsh, A.J., Baldwin, G.S., Kelly, S.M., Waltho, J.P., Price, N.C., Connolly, B.A., Hornby, D.P.
<2>Mechanism-based inhibition of C5-cytosine DNA methyltransferases by 2-H pyrimidinone.
<3>J. Mol. Biol.
<4>286
<5>389-401
<6>1999
<7>DNA duplexes in which the target cytosine base is replaced by 2-H pyrimidinone have previously
been shown to bind with a significantly greater affinity to C5-cytosine DNA methyltransferases
than unmodified DNA.  Here, it is shown that 2-H pyrimidinone, when incorporated into DNA
duplexes containing the recognition sites for M.HgaI-2 and M.MspI, elicits the formation of
inhibitory covalent nucleoprotein complexes.  We have found that although covalent complexes
are formed between 2-H pyrimidinone-modified DNA and both M.HgaI-2 and M.MspI, the kinetics of
complex formation are quite distinct in each case.  Moreover, the formation of a covalent
complex is still observed between 2-H pyrimidinone DNA and M.MspI in which the active-site
cysteine residue is replaced by serine or threonine.  Covalent complex formation between
M.MspI and 2-H pyrimidinone occurs as a direct result of nucleophilic attack by the residue at
the catalytic position, which is enhanced by the absence of the 4-amino function in the base.
The substitution of the catalytic cysteine residue by tyrosine or chemical modification of the
wild-type enzyme with N-ethylmaleimide, abolishes covalent interaction.  Nevertheless the 2-H
pyrimidinone-substituted duplex still binds to M.MspI with a greater affinity than a standard
cognate duplex, since the 2-H pyrimidinone base is mis-paired with guanine.

<>

<1>Hurst, M.R., Glare, T.R., Jackson, T.A.
<2>Cloning Serratia entomophila antifeeding genes--a putative defective prophage active against the grass grub Costelytra zealandica.
<3>J. Bacteriol.
<4>186
<5>5116-5128
<6>2004
<7>Serratia entomophila and Serratia proteamaculans (Enterobacteriaceae)
cause amber disease in the grass grub Costelytra zealandica (Coleoptera:
Scarabaeidae), an important pasture pest in New Zealand. Larval disease
symptoms include cessation of feeding, clearance of the gut, amber
coloration, and eventual death. A 155-kb plasmid, pADAP, carries the genes
sepA, sepB, and sepC, which are essential for production of amber disease
symptoms. Transposon insertions in any of the sep genes in pADAP abolish
gut clearance but not cessation of feeding, indicating the presence of an
antifeeding gene(s) elsewhere on pADAP. Based on deletion analysis of
pADAP and subsequent sequence data, a 47-kb clone was constructed, which
when placed in either an Escherichia coli or a Serratia background exerted
strong antifeeding activity and often led to rapid death of the infected
grass grub larvae. Sequence data show that the antifeeding component is
part of a large gene cluster that may form a defective prophage and that
six potential members of this prophage are present in Photorhabdus
luminescens subsp. laumondii TTO1, a species which also has sep gene
homologues.

<>

<1>Hurst, S.G. IV, Ghazal, S., Morris, K., Abebe-Akele, F., Thomas, W.K., Badr, U.M., Hussein, M.A., AbouZaied, M.A., Khalil, K.M., Tisa, L.S.
<2>Draft Genome Sequence of Photorhabdus temperata Strain Meg1, an Entomopathogenic  Bacterium Isolated from Heterorhabditis megidis Nematodes.
<3>Genome Announcements
<4>2
<5>e01273-14
<6>2014
<7>Photorhabdus temperata strain Meg1 is an entomopathogenic bacterium that forms a  symbiotic
association with Heterorhabditis nematodes. We report here a 4.9-Mbp
draft genome sequence for P. temperata strain Meg1, with a G+C content of 43.18%
and containing 4,340 candidate protein-coding genes.

<>

<1>Hurst, S.G.I.V., Oshone, R., Ghodhbane-Gtari, F., Morris, K., Abebe-Akele, F., Thomas, W.K., Ktari, A., Salem, K., Mansour, S., Gtari, M., Tisa, L.S.
<2>Draft Genome Sequence of Frankia sp. Strain Thr, a Nitrogen-Fixing Actinobacterium Isolated from the Root Nodules of Casuarina cunninghamiana Grown   in Egypt.
<3>Genome Announcements
<4>2
<5>e00493-14
<6>2014
<7>Nitrogen-fixing actinobacteria of the genus Frankia are symbionts of woody dicotyledonous
plants termed actinorhizal plants. We report here a 5.3-Mbp draft
genome sequence for Frankia sp. stain Thr, a nitrogen-fixing actinobacterium
isolated from root nodules of Casuarina cunninghamiana collected in Egypt.

<>

<1>Hutchison, C.A. III, Peterson, S.N., Gill, S.R., Cline, R.T., White, O., Fraser, C.M., Smith, H.O., Venter, J.C.
<2>Global transposon mutagenesis and a minimal mycoplasma genome.
<3>Science
<4>286
<5>2165
<6>1999
<7>Mycoplasma genitalium with 517 genes has the smallest gene complement of any independently
replicating cell so far identified.  Global transposon mutagenesis was used to identify
nonessential genes in an effort to learn whether the naturally occurring gene complement is a
true minimal genome under laboratory growth conditions. The positions of 2209 transposon
insertions in the completely sequenced genomes of M. genitalium and its close relative M.
pneumoniae were determined by sequencing across the junction of the transposon and the genomic
DNA. These junctions defined 1354 distinct sites of insertion that were not lethal.  The
analysis suggests that 265 to 350 of the 480 protein-coding genes of M. genitalium are
essential under laboratory growth conditions, including about 100 genes of unknown function.

<>

<1>Hwang, D.K., Cho, J.Y., Chae, Y.K.
<2>Recombinant expression and purification of functional XorII, a restriction endonuclease from Xanthomonas oryzae pv. oryzae.
<3>J. Microbiol.
<4>45
<5>175-178
<6>2007
<7>An endonuclease from Xanthomonas oryzae pathovar oryzae KACC 10331, XorII, was recombinantly
produced in Escherichia coli using a T7
system. XorII was purified using a combination of ion exchange and
immobilized metal affinity chromatography (IMAC). An optimized washing
protocol was carried out on an IMAC in order to obtain a high purity
product. The final amount of purified XorII was approximately 2.5 mg/L
of LB medium. The purified recombinant XorII was functional and showed
the same cleavage pattern as PvuI. The enzyme activity tested the
highest at 25 degrees C in 50 mM NaCl, 10 mM Tris-HCl, 10 mM MgCl2, and
1 mM dithiothreitol at a pH of 7.9.

<>

<1>Hwang, H.-Y., Yim, J.
<2>SolI, a novel isoschizomer of BamHI isolated from Streptoverticillium olivoverticillatum.
<3>Nucleic Acids Res.
<4>22
<5>2197
<6>1994
<7>SolI, a new type II restriction endonuclease has been purified from Streptoverticillium
olivoverticillatum by chromatography on heparin-agarose and affigel-blue. The enzyme
recognizes the palindromic hexanucleotide sequence of 5'-GGATCC-3' and cleaves after the
first G to produce a 4 base 5'-extension.

<>

<1>Hwang, H.Y., Yim, J.
<2>Characterization of a new type II restriction endonuclease isolated from Streptoverticillium olivoverticillatum.
<3>Korean J. Microbiol.
<4>32
<5>208-214
<6>1994
<7>We screened many species from a wide variety of bacterial genera for a new type II restriction
endonuclease.  The purification and characterization of SolI from a soil isolate,
Streptoverticillum olivoverticillatum are described here.  The enzyme turned out to be an
isoschizomer of BamHI.  It recognized the hexanucleotide sequence 5'-G/GATCC-3' and cleaved
as shown by the arrow, generating a 4 base 5' extension.  Unlike its isoschizomer, BamHI, the
activity was sensitive to dam methylation within the recognition sequence.  Following
ammonium sulfate fractionation of the crude extract, heparin-agarose and Affi-gel Blue column
chromatography were employed to purify the enzyme.  SolI required at least 0.2 mM of MgCl2 for
the cleavage to occur.  The enzyme exhibited its maximal activity in the absence of NaCl, but
was inhibited completely in the presence of 120mM NaCl.  The pH and temperature optima for
activity were pH 8.6 and 40oC, respectively.  The molecular weight of SolI was estimated to be
43,000 Da by Superose-12 gel filtration chromatography.

<>

<1>Hwang, J.Y., Kim, S.H., Oh, H.R., Cho, Y.J., Chun, J., Chung, Y.R., Nam, D.H.
<2>Draft Genome Sequence of Kitasatospora cheerisanensis KCTC 2395, Which Produces Plecomacrolide against Phytopathogenic Fungi.
<3>Genome Announcements
<4>2
<5>e00604-14
<6>2014
<7>Kitasatospora cheerisanensis KCTC 2395, which produces antifungal metabolites with bafilomycin
derivatives, including bafilomycin C1-amide, was isolated from a
soil sample at Mt. Jiri, South Korea. Here, we report its draft genome sequence,
which contains 8.04 Mb with 73.6% G+C content and 7,810 protein-coding genes.

<>

<1>Hwang, S.B., Lee, S.H.
<2>Character and function of restriction enzyme, EcoRI inhibiting substance extracted from spinach chloroplast and Chlamydomonas.
<3>Singmul Hakhoe Chi
<4>33
<5>217-223
<6>1990
<7>Restriction enzyme inhibiting substance (REIS) extracted from spinach
chloroplast and Chlamydomonas seems not to be proteinaceous, because its
inhibiting activity was not lost by heat or trypsin treatment.  And it seems
not to be lipid or polysaccharide, because its inhibiting activity was not lost
by lipase or alpha-amylase treatment, respectively.  In Chlamydomonas,
putrescine, spermidine and spermine were present.  The amount of putrescine was
the smallest and that of spermine was the greatest.  But only spermine was
contained in REIS and the activity of REIS.  It was proportional to the amount
of spermine in REIS and it was hindered by Na+ ion.  So, the inhibiting
activity of REIS seems to be deeply related to spermine contained in REIS.  But
restriction enzyme inhibiting activity remained to the same extent although
salts and spermine were eliminated by dialysis.

<>

<1>Hyman, P., Abedon, S.T.
<2>Bacteriophage Host Range and Bacterial Resistance.
<3>Adv. Appl. Microbiol.
<4>70
<5>217-248
<6>2010
<7>Host range describes the breadth of organisms a parasite is capable of infecting, with limits
on host range stemming from parasite, host, or environmental characteristics. Parasites can
adapt to overcome host or environmental limitations, while hosts can adapt to control the
negative impact of parasites. We consider these adaptations as they occur among bacteriophages
(phages) and their bacterial hosts, since they are significant to phage use as antibacterials
(phage therapy) or to protection of industrial ferments from phage attack. Initially, we
address how phage host range can (and should) be defined plus summarize claims of host ranges
spanning multiple bacterial genera. Subsequently, we review bacterial mechanisms of phage
resistance. These include adsorption resistance, which results in reduced interaction between
phage and bacterium; what we describe as 'restriction,' where bacteria live but phages die;
and abortive infections, where both phage and bacterium die. Adsorption resistance includes
loss of phage receptor molecules on hosts as welt as physical barriers hiding receptor
molecules (e.g., capsules). Restriction mechanisms include phage-genome uptake blocks,
superinfection immunity, restriction modification, and CRISPR, all of which function postphage
adsorption but prior to terminal phage takeover of host metabolism. Standard laboratory
selection methods, involving exposure of planktonic bacteria to high phage densities, tend to
directly select for these prehost-takeover resistance mechanisms. Alternatively, resistance
mechanisms that do not prevent bacterium death are less readily artificially selected.
Contrasting especially bacteria mutation to adsorption resistance, these latter mechanisms
likely are an underappreciated avenue of bacterial resistance to phage attack.

<>

<1>Hyman, R.W., St-Onge, R.P., Allen, E.A., Miranda, M., Aparicio, A.M., Fukushima, M., Davis, R.W.
<2>Multiplex identification of microbes.
<3>Appl. Environ. Microbiol.
<4>76
<5>3904-3910
<6>2010
<7>We have adapted molecular inversion probe technology to identify
microbes in a highly multiplexed procedure. This procedure does not
require growth of the microbes. Rather, the technology employs DNA
homology twice: once for the molecular probe to hybridize to its
homologous DNA and again for the 20-mer oligonucleotide barcode on the
molecular probe to hybridize to a commercially available molecular
barcode array. As proof of concept, we have designed, tested, and
employed 192 molecular probes for 40 microbes. While these particular
molecular probes are aimed at our interest in the microbes in the human
vagina, this molecular probe method could be employed to identify the
microbes in any ecological niche.

<>

<1>Hyun, D.W., Whon, T.W., Cho, Y.J., Chun, J., Kim, M.S., Jung, M.J., Shin, N.R., Kim, J.Y., Kim, P.S., Yun, J.H., Lee, J., Oh, S.J., Bae, J.W.
<2>Genome sequence of the moderately halophilic bacterium Salinicoccus carnicancri type strain Crm(T) (= DSM 23852(T)).
<3>Standards in Genomic Sciences
<4>8
<5>255-263
<6>2013
<7>Salinicoccus carnicancri Jung et al. 2010 belongs to the genus Salinicoccus in the family
Staphylococcaceae. Members of the Salinicoccus are moderately
halophilic and originate from various salty environments. The halophilic features
of the Salinicoccus suggest their possible uses in biotechnological applications,
such as biodegradation and fermented food production. However, the genus
Salinicoccus is poorly characterized at the genome level, despite its potential
importance. This study presents the draft genome sequence of S. carnicancri
strain Crm(T) and its annotation. The 2,673,309 base pair genome contained 2,700
protein-coding genes and 78 RNA genes with an average G+C content of 47.93 mol%.
It was notable that the strain carried 72 predicted genes associated with
osmoregulation, which suggests the presence of beneficial functions that
facilitate growth in high-salt environments.

<>

<1>Iatsenko, I., Corton, C., Pickard, D.J., Dougan, G., Sommer, R.J.
<2>Draft Genome Sequence of Highly Nematicidal Bacillus thuringiensis DB27.
<3>Genome Announcements
<4>2
<5>e00101-14
<6>2014
<7>Here, we report the genome sequence of nematicidal Bacillus thuringiensis DB27, which provides
first insights into the genetic determinants of its pathogenicity
to nematodes. The genome consists of a 5.7-Mb chromosome and seven plasmids,
three of which contain genes encoding nematicidal proteins.

<>

<1>Ibanez, M., Alvarez, I., Rodriguez-Pena, J.M., Rotger, R.
<2>A ColE1-type plasmid from Salmonella enteritidis encodes a DNA cytosine methyltransferase.
<3>Gene
<4>196
<5>145-158
<6>1997
<7>The multicopy plasmid pFM366 was isolated from a virulent Salmonella enteritidis strain and
was found to code for DNA methylase activity (Ibanez and Rotger, 1993). The present work was
aimed at characterizing the genetic organization and functional of this 5.6 kb plasmid. We
found pFM366 almost identical to the plasmid P4 isolated from Shigella sonnei, that encodes
the SsoII restriction-modification system (Karyagina et al., 1993), and related to other
ColE1-type plasmids. Examination of these plasmids revealed a common organization which
suggests they were the result of similar recombinational events. The cytosine methylase of
pFM366 is nearly identical to M. SsoII, whereas the gene encoding the restrictase homologous
to R. SsoII is truncated and its product is inactive. The expression of the cytosine methylase
encoded by pFM366 is strongly affected by deletion of regions located upstream and downstream
of its ORF, and is negatively controlled by the rpoS gene in Escherichia coli. The methylase
activity encoded by pFM366 induces the SOS response, which could be responsible for the
observed delay in the growth of E. coli.

<>

<1>Ibanez, M., Rotger, R.
<2>Characterization of small cryptic plasmid from Salmonella enteritidis that affects the growth of Escherichia coli.
<3>FEMS Microbiol. Lett.
<4>109
<5>225-230
<6>1993
<7>We examined the plasmid content of 25 clinical isolates of Salmonella enteritidis, and
detected the presence of small plasmids (3-5.3 kb) in 9 of them, alone, or in addition to the
large, so-called virulence plasmid.  A 5.3-kb plasmid isolated as unique extrachromosomal DNA
from a strain responsible for a high-mortality outbreak was characterized by restriction
mapping and cloning.  The plasmid replicon was localized in a 1.7-kb fragment, that hybridized
with three of the small plasmids detected in S. enteritidis, and with another small plasmid
from Salmonella typhimurium.  A strain of Escherichia coli carrying this plasmid, or a cloned
3.7-kb PvuII restriction fragment, showed a slower growth rate, especially in minimal medium,
as well as a noticeable increase in DNA methyltransferase activity.

<>

<1>Ibarra-Caballero, J., Zerillo, M.M., Snelling, J., Cranshaw, W., Boucher, C., Tisserat, N.
<2>Genome Sequences of Strain ATCC 29281 and Pin and Northern Red Oak Isolates of Lonsdalea quercina subsp. quercina.
<3>Genome Announcements
<4>2
<5>e00584-14
<6>2014
<7>Two bacteria identified as Lonsdalea quercina subsp. quercina were isolated from  oak trees
showing symptoms of drippy blight. Here, we present their draft genome assemblies, as well as
that of the type strain of this species. To our knowledge, these are the first published
genome sequences of this subspecies of Lonsdalea quercina.

<>

<1>Ibryashkina, E.M., Sasnauskas, G., Solonin, A.S., Zakharova, M.V., Siksnys, V.
<2>Oligomeric structure diversity within the GIY-YIG nuclease family.
<3>J. Mol. Biol.
<4>387
<5>10-16
<6>2009
<7>The GIY-YIG nuclease domain has been identified in homing endonucleases, DNA repair and
recombination enzymes, and restriction endonucleases.  The Type II restriction enzyme Eco29kI
belongs to the GIY-YIG
nuclease superfamily and, like most of other family members, including the
homing endonuclease I-TevI, is a monomer. It recognizes the palindromic
sequence 5�Œ-CCGC/GG-3�Œ (�g/�h marks the cleavage position) and cuts it to
generate 3�Œ-staggered ends. The Eco29kI monomer, which contains a single
active site, either has to nick sequentially individual DNA strands or has to
form dimers or even higher-order oligomers upon DNA binding to make a
double-strand break at its target site. Here, we provide experimental evidence
that Eco29kI monomers dimerize on a single cognate DNA molecule forming the catalytically
active complex. The mechanism described here for Eco29kI differs from that of Cfr42I
isoschisomer, which also belongs to the GIY-YIG family but is functional as a tetramer. This
novel mechanism may have implications for the function of homing endonucleases and other
enzymes of the GIY-YIG family.

<>

<1>Ibryashkina, E.M., Zakharova, M.V., Baskunov, V.B., Bogdanova, E.S., Nagornykh, M.O., Den'mukhamedov, M.M., Melnik, B.S., Kolinski, A., Gront, D., Feder, M., Solonin, A.S., Bujnicki, J.M.
<2>Type II restriction endonuclease R.Eco29kI is a member of the GIY-YIG nuclease superfamily.
<3>BMC Struct. Biol.
<4>7
<5>0
<6>2007
<7>The majority of experimentally determined crystal structures of type II restriction
endonucleases exhibit a common PD-(D/E)XK fold.  Crystal structures have been also determined
for single representatives of two other folds: PLD (R.BfiI) and half-pipe (R.PabI), and
bioinformatics analyses supported by mutagenesis suggested that some REases belong to the HNH
fold.  Our previous bioinformatic analysis suggested that REase R.Eco29kl shares sequence
similarities with one more unrelated nuclease superfamily.  GIY-YIG, however so far no
experimental data were available to support this prediction.  The determination of a crystal
structure of the GIY-YIG domain of homing endonuclease I-TevI provided a template for modeling
of R.Eco29kl and prompted us to validate the model experimentally. Using protein
fold-recognition methods we generated a new alignment between R.Eco29kl and I-TevI, which
suggested a reassignment of one of the putative catalytic residues.  A theoretical model of
R.Eco29kl was constructed to illustrate its predicted three-dimensional fold and organization
of the active site, comprising amino acid residues Y49, Y76, R104, H108, E142, and N154.  A
series of mutants was constructed to generate amino acid substitutions of selected residues
(Y49A, R104A, H108F, E142A and N154L) and the mutant proteins were examined for their ability
to bind the DNA containing the Eco29kl site 5'-CCGCGG-3' and to catalyze the cleavage
reaction.  Experimental data reveal that residues Y49, R104, E142, H108, and N154 are
important for the nuclease activity of R.Eco29kl, while H108 and N154 are also important for
specific DNA binding by this enzyme.

<>

<1>Ichida, H., Maeda, K., Ichise, H., Matsuyama, T., Abe, T., Yoneyama, K., Koba, T.
<2>In silco irestiriction landmairk genome scanning analysis of Xanthomonas oryzae pathovair oryzae MAFF 311018.
<3>Biochem. Biophys. Res. Commun.
<4>363
<5>852-856
<6>2007
<7>We have developed a restriction landmark genome scanning (RLGS) system in silico, involving
two-dimensional electrophoretic analysis of DNA by
computer simulation that is based on the availability of whole-genome
sequences for specific organisms. We applied the technique to the
analysis of the Xanthomonas oryzae pathovar oryzae (Xoo) MAFF 311018,
which causes bacterial blight in rice. The coverage that was found to
be achievable using RLGS in silico, as a percentage of the genomic
regions that could be detected, ranged from 44.5% to 72.7% per image.
However, this reached a value of 96.7% using four images that were
obtained with different combinations of landmark restriction enzymes.
Interestingly, the signal intensity of some of the specific spots
obtained was significantly lower than that of other surrounding spots
when Mbol, which cleaves unmethylated 5'-GATC-3' sites, was used. DNA
gel blot analysis with both DNA adenine methylase (Dam)-sensitive and
-insensitive isoschizomers (Mbol and Sau3AI) revealed that Dam-mediated
DNA adenine methylation had indeed occurred at these particular sites.
These results suggest that a significant portion of the 5'-GATC-3'
sites within the Xoo genome is stably methylated by Dam.

<>

<1>Ichida, H., Matsuyama, T., Abe, T., Koba, T.
<2>DNA adenine methylation changes dramatically during establishment of symbiosis.
<3>FEBS J.
<4>274
<5>951-962
<6>2007
<7>The DNA adenine methylation status on specific 5'-GANTC-3' sites and its
change during the establishment of plant-microbe interactions was
demonstrated in several species of alpha-proteobacteria. Restriction
landmark genome scanning (RLGS), which is a high-resolution two
dimensional DNA electrophoresis method, was used to monitor the genomewide
change in methylation. In the case of Mesorhizobium loti MAFF303099, real
RLGS images obtained with the restriction enzyme MboI, which digests at
GATC sites, almost perfectly matched the virtual RLGS images generated
based on genome sequences. However, only a few spots were observed when
the restriction enzyme HinfI was used, suggesting that most GANTC (HinfI)
sites were tightly methylated and specific sites were unmethylated. DNA
gel blot analysis with the cloned specifically unmethylated regions (SUMs)
showed that some SUMs were methylated differentially in bacteroids
compared to free-living bacteria. SUMs have also been identified in other
symbiotic and parasitic bacteria. These results suggest that DNA adenine
methylation may contribute to the establishment and/or maintenance of
symbiotic and parasitic relationships.

<>

<1>Ichige, A., Kobayashi, I.
<2>Stability of EcoRI Restriction-Modification Enzymes In Vivo Differentiates the EcoRI Restriction-Modification System from Other Postsegregational   Cell Killing Systems.
<3>J. Bacteriol.
<4>187
<5>6612-6621
<6>2005
<7>Certain type II restriction modification gene systems can kill host cells when these gene
systems are eliminated from the host cells. Such ability
to cause postsegregational killing of host cells is the feature of
bacterial addiction modules, each of which consists of toxin and antitoxin
genes. With these addiction modules, the differential stability of toxin
and antitoxin molecules in cells plays an essential role in the execution
of postsegregational killing. We here examined in vivo stability of the
EcoRI restriction enzyme (toxin) and modification enzyme (antitoxin), the
gene system of which has previously been shown to cause postsegregational
host killing in Escherichia coli. Using two different methods, namely,
quantitative Western blot analysis and pulse-chase immunoprecipitation
analysis, we demonstrated that both the EcoRI restriction enzyme and
modification enzyme are as stable as bulk cellular proteins and that there
is no marked difference in their stability. The numbers of EcoRI
restriction and modification enzyme molecules present in a host cell
during the steady-state growth were estimated. We monitored changes in
cellular levels of the EcoRI restriction and modification enzymes during
the postsegregational killing. Results from these analyses together
suggest that the EcoRI gene system does not rely on differential stability
between the toxin and the antitoxin molecules for execution of
postsegregational cell killing. Our results provide insights into the
mechanism of postsegregational killing by restriction-modification
systems, which seems to be distinct from mechanisms of postsegregational
killing by other bacterial addiction modules.

<>

<1>Ichige, A., Kobayashi, I.
<2>Stability of EcoRI restriction modification enzymes in vivo differentiates EcoRI restriction-modification system from other postsegregational cell killing systems.
<3>Genes Genet. Syst.
<4>80
<5>454
<6>2005
<7>Certain Type II restriction modification gene systems can kill host cells when these gene
systems are eliminated from the host cells.  Such ability to cause prostsegregational killing
of host cells is the feature of bacterial addiction modules, each of which consists of toxin
and antitoxin genes.  With these addiction modules, differential stability of toxin and
antitoxin molecules in cells plays an essential role in execution of postsegregational
killing.  We here examined in vivo stability of the EcoRI restriction enzyme (toxin) and
modification enzyme (antitoxin) in Escherichia coli.  Using Western blot analysis and
pulse-chase immunoprecipitation analysis, we demonstrated that both the EcoRI restriction
enzyme and modification enzyme are as stable as bulk cellular proteins and that there is no
marked difference in their stability.  We also monitored changes in cellular levels of the
EcoRI restriction and modification enzyme during postsegregational killing.  Results from
these analyses together suggest that the EcoRI gene system does not rely on differential
stability between the toxin and the antitoxin molecules for execution of postsegregational
cell killing.

<>

<1>Ichikawa, N. et al.
<2>Genome Sequence of Kitasatospora setae NBRC 14216T: An Evolutionary Snapshot of the Family Streptomycetaceae.
<3>DNA Res.
<4>10
<5>1-10
<6>2010
<7>Kitasatospora setae NBRC 14216T (5KM-6054T) is known to produce setamycin (bafilomycin B1)
possessing antitrichomonal activity. The genus Kitasatospora is morphologically similar to the
genus Streptomyces, although they are distinguishable from each other on the basis of cell
wall composition and the 16S rDNA sequence. We have determined the complete genome sequence of
K. setae NBRC 14216T as the first Streptomycetaceae genome other than Streptomyces. The genome
is a single linear chromosome of 8 783 278 bp with terminal inverted repeats of 127 148 bp,
predicted to encode 7569 protein-coding genes, 9 rRNA operons, 1 tmRNA and 74 tRNA genes.
Although these features resemble those of Streptomyces, genome-wide comparison of orthologous
genes between K. setae and Streptomyces revealed smaller extent of synteny. Multilocus
phylogenetic analysis based on amino acid sequences unequivocally placed K. setae outside the
Streptomyces genus. Although many of the genes related to morphological differentiation
identified in Streptomyces were highly conserved in K. setae, there were some differences such
as the apparent absence of the AmfS (SapB) class of surfactant protein and differences in the
copy number and variation of paralogous components involved in cell wall synthesis.

<>

<1>Ichiyanagi, K.
<2>Inhibition of MspI cleavage activity by hydroxymethylation of the CpG site A concern for DNA modification studies using restriction endonucleases.
<3>EPIGENETICS
<4>7
<5>131-136
<6>2012
<7>In mammalian genomic DNA, cytosine methylation predominantly occurs at CpG dinucleotides and
provides epigenetic information. In some cells,
5-methyl-cytosine (5-mC) can be further converted to
5-hydroxymethyl-cytosine (5-hmC) by the ten-eleven translocation family
of proteins. MspI restriction endonuclease has been used to analyze
these modified cytosines. However, the kinetic analysis in this study
revealed that MspI activity is dramatically decreased by symmetrical
hydroxymethylation of its recognition sequence and partly inhibited by
hemi-hydroxymethylation, whereas TaqI and HaeIII are relatively
resistant to hydroxymethylation. Therefore, DNA modification studies
that use MspI, for example, reduced representation bisulfite shotgun
sequencing, quantitative analysis of 5-hmC and cleavage-sensitivity
analysis, should be carefully interpreted.

<>

<1>Ichiyanagi, K., Ishino, Y., Ariyoshi, M., Komori, K., Morikawa, K.
<2>Crystal structure of an archaeal intein-encoded homing endonuclease PI-PfuI.
<3>J. Mol. Biol.
<4>300
<5>889-901
<6>2000
<7>Inteins possess two different enzymatic activities, self-catalyzed protein splicing and
site-specific DNA cleavage. These endonucleases, which are classified as part of the homing
endonuclease family, initiate the mobility of their genetic elements into homologous alleles.
They recognize long asymmetric nucleotide sequences and cleave both DNA strands in a monomer
form. We present here the 2.1 A crystal structure of the archaeal PI-PfuI intein from
Pyrococcus furiosus. The structure reveals a unique domain, designated here as the Stirrup
domain, which is inserted between the Hint domain and an endonuclease domain. The
horseshoe-shaped Hint domain contains a catalytic center for protein splicing, which involves
both N and C-terminal residues. The endonuclease domain, which is inserted into the Hint
domain, consists of two copies of substructure related by an internal pseudo 2-fold axis. In
contrast with the I-CreI homing endonuclease, PI-PfuI possibly has two asymmetric catalytic
sites at the center of a putative DNA-binding cleft formed by a pair of four-stranded
beta-sheets. DNase I footprinting experiments showed that PI-PfuI covers more than 30 bp of
the substrate asymmetrically across the cleavage site. A docking model of the DNA-enzyme
complex suggests that the endonuclease domain covers the 20 bp DNA duplex encompassing the
cleavage site, whereas the Stirrup domain could make an additional contact with another
upstream 10 bp region. For the double-strand break, the two strands in the DNA duplex were
cleaved by PI-PfuI with different efficiencies. We suggest that the cleavage of each strand is
catalyzed by each of the two non-equivalent active sites.

<>

<1>Ihara, M.
<2>Web-based DNA sequence analysis program based on restriction enzyme cleavage and alignment.
<3>Japanese Patent Office
<4>JP 200217373
<5>
<6>2002
<7>
<>

<1>Ii, J.S.B., Rees, P.A.
<2>DNA segment coding for Hinc II restriction endonuclease.
<3>Japanese Patent Office
<4>JP 1998080284 A
<5>
<6>1998
<7>
<>

<1>Iida, S., Meyer, J., Bachi, B., Stalhammar-Carlemalm, M., Schrickel, S., Bickle, T.A., Arber, W.
<2>DNA Restriction-Modification Genes of Phage P1 and Plasmid p15B.
<3>J. Mol. Biol.
<4>165
<5>1-18
<6>1983
<7>The EcoP1 and EcoP15 DNA restriction-modification systems are coded by the related P1 prophage
and p15B plasmid.  We have examined the organization of the genes for these systems using P1
itself, "P1-P15" hybrid phages expressing the EcoP15 restriction specificity of p15B and
cloned restriction fragments derived from these phage DNAs.  The results of transposon
mutagenesis, restriction cleavage analysis, DNA heteroduplex analysis and in vitro
transcription mapping allow the following conclusions to be drawn concerning the structural
genes. (1) All of the genetic information necessary to specify either system is contained
within a contiguous DNA segment of 5000 bases which encodes two genes.  One of them, necessary
for both restriction and modification, we call mod and the other, required only for
restriction (together with mod), we call res.  (2) The res gene is about 2800 bases long and
at the heteroduplex level is largely identical for P1 and P15:  it shows a small region of
partial nonhomology and some restriction cleavage site differences.  The mod gene is about
2200 bases long and contains a 1200 base long region of non-homology between P1 and P15 toward
the N-terminus of the gene.  The rest of the gene at this level of analysis is identical for
the two systems.  (3) Each of the genes is transcribed in vitro from its own promoter.  It is
possible that the res gene is also transcribed by readthrough from the mod promoter.

<>

<1>Iida, S., Streiff, M.B., Bickle, T.A., Arber, W.
<2>Two DNA antirestriction systems of bacteriophage P1, darA, and darB:  characterization of darA- phages.
<3>Virology
<4>157
<5>156-166
<6>1987
<7>Bacteriophage P1 is only weakly restricted when it infects cells carrying type
I restriction and modification systems even though DNA purified from P1 phage
particles is a good substrate for type I restriction enzymes in vitro.  Here we
show that this protection against restriction is due to the products of two
phage genes which we call darA and darB (dar for defense against restriction).
Each of the dar gene products provides protection against a different subset of
type I restriction systems.  The darA and darB gene products are found in the
phage head and protect any DNA packaged into a phage head, including transduced
chromosomal markers, from restriction.  The proteins must, therefore, be
injected into recipient cells along with the DNA.  The proteins act strictly in
cis.  For example, upon double infection of restricting cells with dar+ and
dar- P1 phages, the dar+ genomes are protected from restriction while the
dar-genomes are efficiently restricted.

<>

<1>Iida, T., Suetake, I., Tajima, S., Morioka, H., Ohta, S., Obuse, C., Tsurimoto, T.
<2>PCNA clamp facilitates action of DNA cytosine methyltransferase 1 on hemimethylated DNA.
<3>Genes Cells
<4>7
<5>997-1008
<6>2002
<7>Proliferating cell nuclear antigen (PCNA) is a ring-shaped protein known as a processivity
factor of DNA polymerase delta.  In addition to this role, PCNA interacts with a number of
other proteins to increase their local concentration at replicated DNA sites.  DNA cytosine
methyltransferase 1 (Dnmt1), which preserves epigenetic signals by completing the methylation
of hemimethylated DNA aftr DNA replication, has been indicated as one of these PCNA binding
proteins by a previous work.  However, the molecular mechanisms and functional significance of
their association have not yet been studied.  Results: Dnmt1 can be readily isolated from
nuclear extracts by PCNA affinity chromatography.  Studies of the interactions between the two
proteins demonstrate that the N-terminal region of Dnmt1, which contains a typical PCNA
binding
motif, has core PCNA binding activity, and that the remaining portion of the protein exerts a
negative influence on the interaction of Dnmt1 with PCNA.  The affinity of Dnmt1 for DNA is
much higher for DNA bound by PCNA than for free DNA.  Furthermore, DNA methylation assays with
hemimethylated DNA as a substrate revealed that PCNA clamp-bound DNA is methylated more
efficiently by Dnmt1 than is free DNA.  Conclusion: These results provide the first
biochemical evidence that physical interactions between PCNA and Dnmt1 facilitate the
methylation of newly replicated DNA, on which PCNA remains associated as a functional clamp.

<>

<1>Ikawa, S., Shibata, T., Ando, T.
<2>Recognition sequence of endonuclease R. BamNx from Bacillus amyloliquefaciens N.
<3>Agric. Biol. Chem.
<4>43
<5>873-875
<6>1979
<7>Recently, many site-specific deoxyribonucleases including restriction
endonucleases which cleave DNA strands at unique sites have been isolated from
various kinds of microorganisms.  Recognition nucleotide sequences on DNA have
been determined for a number of them.  We have reported that Bacillus
amyloliquefaciens N and many other Bacillus strains possess site-specific
endonucleases.

<>

<1>Ikawa, S., Shibata, T., Ando, T.
<2>Host-Controlled modification and restriction in Bacillus subtilis.  Bsu168-System and BsuR-System in B. subtilis 168.
<3>Mol. Gen. Genet.
<4>170
<5>123-127
<6>1979
<7>A Bsu168-specific restriction deficient (r-1)6)8) mutant of Bacillus subtilis
Marburg 168 was transformed to be BsuR-specific restriction proficient (r+R)
with B. subtilis R DNA as efficiently as the Bsu168-specific restriction
proficient (r+1)6)8) parental strain (hsrM+, hsdR-). We constructed
r+Rm+Rr+1m6m8mm+1868 strain (ISMR4), r+Rm+Rr-1m6m8mm+1868 strain (ISR11) and
r+Rm+Rr-1m6m8mm-1868 strain (ISR6) from strain 101 (r+1868m+1868), strain 1012
(r-1868m+1868) and strain RM125 (r-1868m-1868), respectively by transformation
with B. subtilis R DNA, and tested their restriction and modification
activities on phage Phi105C.  The results show that the sites recognized by
Bsu168-specific restriction and modification enzymes and the sites recognized
by BsuR-specific ones are not overlapping. We conclude that the
Bsu168-modification and restriction system and the BsuR-modification and
restriction system are controlled independently by two distinct sets of genes
in the r_Rm+R transformant of r+1868m+1868 strain B. subtilis 168.

<>

<1>Ikawa, S., Shibata, T., Ando, T.
<2>The Site-specific Deoxyribonuclease from Bacillus pumilus (Endonuclease R.Bpu1387).
<3>J. Biochem. (Tokyo)
<4>80
<5>1457-1460
<6>1976
<7>A new site-specific endonuclease (DNase) was isolated from the cells of
Bacillus pumilus AHU 1387 strain.  This enzyme (endonuclease R.Bpu 1387)
introduced double-stranded scissions at unique sites on DNA's of coli phage
lambda, lambda dvl, coli phage T7, Bacillus phage Phi105C, Bacillus phage SP10,
and Simian Virus 40, in the presence of magnesium ion.  The activity was
stimulated by the presence of NaCl.

<>

<1>Ikawa, S., Shibata, T., Ando, T., Saito, H.
<2>Genetic studies on site-specific endodeoxyribonucleases in Bacillus subtilis:  Multiple modification and restriction systems in transformants of Bacillus subtilis 168.
<3>Mol. Gen. Genet.
<4>177
<5>359-368
<6>1980
<7>We transformed B. subtilis 168 with DNA from B. subtilis IAM1231, IAM1192 and
ATCC6633.  When we examined the restriction activities of the transformants in
vivo and in vitro using phage Phi105C we found the following: (1) Cells of
either IAM1231 or IAM1192 have two modification and restriction systems
(Bsu1231(1)-system and Bsu1231(II)-system in IAM1231, and Bsu1192(I)-system and
Bsu1192(II)-systems in IAM1192), and cells of ATCC6633 have only one system
(Bsu6633-system).  (2) The restriction enzymes of all of these five systems are
site-specific endonucleases.  (3) The nucleotide sequence specifities of the
enzymes involved in Bsu1231(I)-system, Bsu1192(I)-system and Bsu6633-system are
the same; and those of Bsu1231(II)-system and Bsu1192(II)-system are the same.
The sequence specificities of these two groups are different from each other
and also different from those of the Bsu168-system of B. subtilis 168, the
BsuR-system of B. subtilis R and the Bsu1247(I)-and Bsu1247(II)-systems which
are systems of B. subtilis IAM1247.  (4) Transformants possessing four
different modification and restriction systems (Bsu1231(I)-, Bsu1247(I)-, BsuR-
and Bsu168-systems) were constructed.  (5) Transformation of two derivatives of
168 that were mR+rR+ by DNA from IAM1231 produced 16 transformants that had the
Bsu1231(II) restriction system, but had lost the BsuR system.  Transformation
of a derivative of 168 that was m+1247(II)r+1247(II) by DNA from
m+1231(II)r+1231(II)- or mR+rR+-derivative of 168 produced about 100 each of
transformants that had the Bsu1231(II)-restriction system or the
BsuR-restriction system.  But all these transformants lost the
Bsu1247(II)-system.

<>

<1>Ikawa, S., Shibata, T., Matsumoto, K., Iijima, T., Saito, H., Ando, T.
<2>Chromosomal loci of genes controlling site-specific restriction endonucleases of Bacillus subtilis.
<3>Mol. Gen. Genet.
<4>183
<5>1-6
<6>1981
<7>We constructed transformants of B. subtilis 168 which acquired genes for
site-specific restriction endonucleases.  These endonucleases originated from
various strains of B. subtilis and were classified into five groups based on
the specificity of the sequences recognized by the enzymes.  We examined the
loci of genes for site-specific restriction endonucleases belonging to
different groups:  hsrE determined Endo.R.Bsu1231(I), hsrB Endo.R.Bsu1247(I),
hsrR Endo.R.BsuR and hsrC Endo.R.Bsu-1247(II).  One gene, hsrE, was located
between sacA and purA by transduction crosses with phage PBS1, and another
gene, hsrB, between hsrE and purA.  Genes hsrR and hsrC had been suggested to
be allelic or closely linked by previous studies with transformation.  We
located hsrR and hsrC between purB and tre.  Our previous observation and this
study show that B. subtilis 168 has at least three independent loci on the
chromosome for four genes for site-specific restriction endonucleases in
addition to the locus for the original restriction activity (Bsu168-specific
restriction) of strain 168.

<>

<1>Ikeda, H., Ishikawa, J., Hanamoto, A., Shinose, M., Kikuchi, H., Shiba, T., Sakaki, Y., Hattori, M., Omura, S.
<2>Complete genome sequence and comparative analysis of the industrial microorganism Streptomyces avermitilis.
<3>Nat. Biotechnol.
<4>21
<5>526-531
<6>2003
<7>Species of the genus Streptomyces are of major pharmaceutical interest
because they synthesize a variety of bioactive secondary metabolites. We
have determined the complete nucleotide sequence of the linear chromosome
of Streptomyces avermitilis. S. avermitilis produces avermectins, a group
of antiparasitic agents used in human and veterinary medicine. The genome
contains 9,025,608 bases (average GC content, 70.7%) and encodes at least
7,574 potential open reading frames (ORFs). Thirty-five percent of the
ORFs (2,664) constitute 721 paralogous families. Thirty gene clusters
related to secondary metabolite biosynthesis were identified,
corresponding to 6.6% of the genome. Comparison with Streptomyces
coelicolor A3(2) revealed that an internal 6.5-Mb region in the S.
avermitilis genome was highly conserved with respect to gene order and
content, and contained all known essential genes but showed perfectly
asymmetric structure at the oriC center. In contrast, the terminal regions
were not conserved and preferentially contained nonessential genes.

<>

<1>Ikeda, M., Nakagawa, S.
<2>The Corynebacterium glutamicum genome: Features and impacts on biotechnological processes.
<3>Appl. Microbiol. Biotechnol.
<4>62
<5>99-109
<6>2003
<7>Corynebacterium glutamicum has played a principal role in the progress of the amino acid
fermentation industry. The complete genome sequence of the representative wild-type strain of
C. glutamicum, ATCC 13032, has been determined and analyzed to improve our understanding of
the molecular biology and physiology of this organism, and to advance the development of more
efficient production strains. Genome annotation has helped in elucidation of the gene
repertoire defining a desired pathway, which is accelerating pathway engineering. Post genome
technologies such as DNA arrays and proteomics are currently undergoing rapid development in
C. glutamicum. Such progress has already exposed new regulatory networks and functions that
had so far been unidentified in this microbe. The next goal of these studies is to integrate
the fruits of genomics into strain development technology. A novel methodology that merges
genomics with classical strain improvement has been developed and applied for the
reconstruction of classically derived production strains. How can traditional fermentation
benefit from the C. glutamicum genomic data? The path from genomics to biotechnological
processes is presented.

<>

<1>Ikram, H.I., Catherine, R., Caroline, M., Didier, R., Hocine, H., Christelle, D.
<2>Non-contiguous finished genome sequence and description of Halopiger goleamassiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>956-959
<6>2014
<7>Halopiger goleamassiliensis strain IIH3(T) sp. nov. is a novel, extremely halophilic archaeon
within the genus Halopiger. This strain was isolated from an
evaporitic sediment in El Golea Lake, Ghardaia region (Algeria). The type strain
is strain IIH3(T). H. goleamassiliensis is moderately thermophilic, neutrophilic,
non-motile and coccus-shaped. Here we describe the features of this organism,
together with the complete genome sequence and annotation. The 3,906,923 bp long
genome contains 3,854 protein-encoding genes and 49 RNA genes (1 gene is 16S
rRNA, 1 gene is 23S rRNA, 3 genes are 5S rRNA, and 44 are tRNA genes).

<>

<1>Imagawa, T., Nakayama, H., Katunuma, N., Sakuraba, H., Ohshima, T., Itoh, T., Sako, Y., Nomura, N., Tsuge, H.
<2>Crystallization and preliminary X-ray diffraction analysis of homing endonuclease I-Tsp061I.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>60
<5>2006-2008
<6>2004
<7>Two crystal forms, rhombohedral and hexagonal, of a homing endonuclease from Thermoproteus sp.
IC-061 (I-Tsp0611) were obtained by the
hanging-drop and sitting-drop method, respectively. The hexagonal
crystals belong to space group P6(3)22, with unit-cell parameters a = b
= 111.4, c = 97.6 Angstrom, and diffract to 3.2 Angstrom resolution on
beamline BL44 at SPring-8 (Harima, Japan). The rhombohedral crystals
belong to space group R32, with unit-cell parameters a = b = 95.4, c =
192.9 Angstrom, and diffract to 2.7 Angstrom resolution using a Cu
Kalpha rotating-anode generator with an R-AXIS VII detector. The
crystal asymmetric unit contained one protein molecule and the solvent
contents of the two crystal forms were estimated to be 68.3 and 67.6%
by volume, respectively.

<>

<1>Imanaka, T., Atomi, H.
<2>Ligase of a hyperthermostable bacterium and chip using the same.
<3>European Patent Office
<4>EP 1923464 A
<5>
<6>2008
<7>It is intended to provide an efficient and sure gene targeting method embodied at an arbitrary
position in the genome of an organism and a kit therefore.  It is also intended to provide a
method for targeted-disruption of an arbitrary gene in the genome of an organism which
comprises: 1) the step of providing the whole sequencial data of the genome of the organism;
2) the step of selecting at lesat one arbitrary region in the sequence; 3) the step of
providing a vector containing a sequence homologous with the region selected above and a
marker gene; 4) the step of transforming the organism by the vector; and 5) the step of
providing the organism under such conditions as allowing homologous recombination.  Moreover,
the genome of a hyperthermostable bacterium and its array are provided.

<>

<1>Imber, R., Bickle, T.A.
<2>Purification and properties of the restriction endonuclease BglII from Bacillus globigii.
<3>Eur. J. Biochem.
<4>117
<5>395-399
<6>1981
<7>The restriction endonuclease BglII from Bacillus globigii has been purified to
homogeneity.  The enzyme is a dimer of two subunits of Mr = 27000.  The
reaction mechanism does not involve the accumulation of a DNA intermediate
nicked in one strand and the enzyme is not affected by superhelical twists in
the substrate DNA, indicating that DNA binding does not involve either winding
or unwinding of the double helix.  Antibodies were prepared against BglII.
These antibodies did not cross react with any other restriction endonucleases
tested, including other enzymes from B. globigii or from closely related
strains.  It is thus unlikely that type II restriction enzymes represent a
closely related group of proteins.

<>

<1>Imhof, P., Fischer, S., Smith, J.C.
<2>Catalytic Mechanism of DNA Backbone Cleavage by the Restriction Enzyme EcoRV: A Quantum Mechanical/Molecular Mechanical Analysis.
<3>Biochemistry
<4>48
<5>9061-9075
<6>2009
<7>Endonucleases, Such as the restriction enzyme EcoRV, cleave the DNA backbone at a specific
recognition sequence. We have investigated the
catalytic mechanism of backbone phosphodiester hydrolysis by the
restriction enzyme EcoRV by means of hybrid quantum
mechanical/molecular mechanical calculations. An exhaustive computation
of different reaction pathways is performed, thus generating a network
of pathways. Comparison of the computed (AM1d/MM) enzymatic reaction
pathways with an analogous mechanism for small-molecule model systems
[AM1/d and B3LYP/6-31 + +G(d,p)] reveals that the transition barriers
for associative hydrolysis, which is more probable in the model
systems, are not lowered by the enzyme. Instead, a reaction mechanism
which has mostly dissociative character is more likely. The protein
environment is tuned to significantly electrostatically stabilize the
transition state structures, The direct catalytic impact of essential
residues is determined: The magnesium metal Ion activates a water
molecule, thus facilitating protonation of the leaving group. A
reduction of the coordination number of the magnesium metal ion from
six to four upon the positioning of the attacking water molecule
explains why larger metal ions, such as calcium, are not catalytically
active. The nucleophile is generated by the transfer of a proton from
the attacking water molecule to a carboxylic oxygen atom of aspartate
90. The catalytic effect of lysine 92 involves proper positioning of
the scissile phosphate group and, more importantly, stabilization of
the metaphosphate intermediate in an orientation optimal for attack of
the nucleophile.

<>

<1>Inada, S., Watanabe, K.
<2>Draft Genome Sequence of Meiothermus ruber H328, Which Degrades Chicken Feathers, and Identification of Proteases and Peptidases Responsible for Degradation.
<3>Genome Announcements
<4>1
<5>e00176-13
<6>2013
<7>Meiothermus ruber H328 was isolated from Arima Hot Springs, Kobe, Japan, as a moderate
thermophile. It has a strong ability to degrade intact chicken feathers.
The enzymatic mechanism of the strain for feather degradation is unclear. The
draft genome suggests potent enzyme candidates for degradation of keratin, a
hard-to-degrade protein found in feathers.

<>

<1>Inagaki, K.
<2>Chromatographic purification of restriction enzyme of Acidiphilium.
<3>Japanese Patent Office
<4>JP 4304886
<5>
<6>1992
<7>A new restriction enzyme is obtained from the culture of a strain of the genus Acidiphilium.
The restriction enzyme recognises and cuts the nucleotide sequence (I).
USE/ADVANTAGE - The restriction enzyme has the same recognition site and digestion site as
Eco47III. It can be produced industrially.

<>

<1>Inagaki, K., Dou, D., Kita, K., Hiraoka, N., Kishimoto, N., Sugio, T., Tano, T.
<2>Isolation and characterization of restriction endonucleases from Acidiphilium sp. 16R and 22M.
<3>J. Ferment. Bioeng.
<4>69
<5>60-62
<6>1990
<7>Restriction endonucleases (Asp16RI and Asp22MI) have been identified from the acidophilic
bacteria Acidiphilium sp. 16R and 22M. The cleavage patterns with various DNAs show that both
enzymes recognize the same sequence as the PvuI restriction endonuclease (5'-CGAT^CG-3'),
which is from Proteus vulgaris ATCC 13315. Most of the catalytic properties observed for
Asp16RI and Asp22MI were similar to those observed for PvuI. However, unlike PvuI both enzymes
efficiently cleaved DNA in the absence of NaCl or KCl. The purification yield of Asp22MI is 60
times that of PvuI.

<>

<1>Inagaki, K., Hikita, T., Yanagidani, S., Nomura, Y., Kishimoto, N., Tano, T., Tanaka, H.
<2>Restriction endonuclease Aor13HI from Acidiphilium organovorum 13H, a new isoschizomer of BspMII: Purification and characterization.
<3>Biosci. Biotechnol. Biochem.
<4>57
<5>1716-1721
<6>1993
<7>A restriction endonuclease, Aor13HI, an isoschizomer of BspMII, was purified to homogeneity
from cell extracts of Acidiphilium organovorum strain 13H. The enzyme has a molecular mass of
60,000 daltons and consists of two subunits identical in molecular mass of 30,000 daltons.
Aor13HI endonuclease, like BspMII, recognizes the palindromic six-base sequence
5'-TCCGGA-3', and cleaves between the T and C to produce a four-base 5' extension. Aor13HI
is not inhibited by dam-dependent methylation. The isoelectric point of the enzyme is 5.7.
Aor13HI activity was maximum at pH 7.5, 100 mM KCI, 7.5-10mM MgC12 and 55 degrees C. The
enzyme was stable up to 60 degrees C. The N-terminal amino acid sequence (30 residues) of
Aor13HI did not show similarity with the sequence of other restriction endonuclease reported.

<>

<1>Inagaki, K., Ito, T., Sagawa, H., Kotani, H., Kishimoto, N., Sugio, T., Tano, T., Tanaka, H.
<2>AcpI, a novel isoschizomer of AsuII from Acidiphilium cryptum 25H, recognizes the sequence 5'TT^CGAA3'.
<3>Nucleic Acids Res.
<4>19
<5>6335
<6>1991
<7>AcpI, a type II restriction endonuclease, has been isolated from Acidiphilium
cryptum 25H.  AcpI, an isoschizomer of AsuII, recognizes the six base sequence
5'TTCGAA3', and cleaves between the T and the C residues to produce a two base
5' extension.

<>

<1>Inagaki, K., Kobayashi, F., Dou, D., Nomura, Y., Kotani, H., Kishimoto, N., Sugio, T., Tano, T.
<2>Isolation and identification of restriction endonuclease Asp35HI from Acidiphilium species 35H.
<3>Nucleic Acids Res.
<4>18
<5>6155
<6>1990
<7>None

<>

<1>Inano, K., Suetake, I., Ueda, T., Miyake, Y., Nakamura, M., Okada, M., Tajima, S.
<2>Maintenance-type DNA methyltransferase is highly expressed in post-mitotic neurons and localized in the cytoplasmic compartment.
<3>J. Biochem. (Tokyo)
<4>128
<5>315-321
<6>2000
<7>Maintenance-type DNA methyltransferase (Dnmt1) is usually down-regulated in nonproliferating
cells, In the present study, we
detected significant expression of Dnmt1 protein in adult mouse brain
where the majority of the cells are in a post-mitotic state. A
significant amount of Dnmt1 protein was fractionated into the
post-nuclear fraction for both cerebrum and cerebellum. The Dnmt1 in
this fraction was enzymatically active. An immunofluorescence study
revealed that Dnmt1 protein was mainly expressed in neurons and seemed
to be localized in the cytoplasmic compartment, Primary culturing of
neurons confirmed the expression and localization of Dnmt1 in the
cytoplasmic compartment, The findings that the Dnmt1 transcript in the
brain utilized the somatic-type exon and that the apparent size of the
Dnmt1 protein in the cytoplasm was identical to that in proliferating
culture cells indicate that the cytoplasmic Dnmt1 in neurons was of the
somatic-type.

<>

<1>Indiana, A., Briand, M., Arlat, M., Gagnevin, L., Koebnik, R., Noel, L.D., Portier, P., Darrasse, A., Jacques, M.A.
<2>Draft Genome Sequence of the Flagellated Xanthomonas fuscans subsp. fuscans Strain CFBP 4884.
<3>Genome Announcements
<4>2
<5>e00966-14
<6>2014
<7>We report the draft genome sequence of the flagellated strain CFBP 4884 of Xanthomonas fuscans
subsp. fuscans, which was isolated in an outbreak of common
bacterial blight of beans along with non-flagellated strains. Comparative
genomics will allow one to decipher the genomic diversity of strains cohabiting
in epidemics.

<>

<1>Inocencio, T., Vital, J., Vitor, J., Falcao, A.O.
<2>Genome inspector: A web tool for exploring bacterial genomes.
<3>Proc. InForum 2013, , Cachopo, J., Sousa Santos, B., Evora, Portugal
<4>0
<5>5-16
<6>2013
<7>Genome Inspector (GIN) is a new interactive web-based application for exploring bacterial
genomes designed to provide users with a wide choice of options to facilitate the process of
designing DNA primers for isolating genes from similar bacterial species and strains.  GIN
employs a project-based approach in which users can select any given set of available genomes
and perform a comprehensive bioinformatics analysis.  Currently, it encompasses the full set
of annotated bacterial genomes available on GenBank, a total of 4300 files corresponding to
over 740,000 annotated genes.  GIN allows new data to be added directly from GenBank as it is
being generated, as well as new genome uploading for personal analysis.  The application
interfaces were designed with potential users in mind to assist with their most common
research goals.  Users can visually explore full circular genomes, search for similar regions
in other species and strains, and visualize amplified genomic regions for several strains
simultaneously.  This interactive tool allows for dynamic graphical exploration and refinement
of search and exploration criteria.  Furthermore, it includes a multiple alignment algorithm
(MUSCLE) to help researchers in the process of designing primers.  GIN is free and publicly
available at http://gin.ul.pt/GIN2/index.php.

<>

<1>Inselburg, J.
<2>Phage P1 modification of bacterial DNA studied by generalized transduction.
<3>Virology
<4>30
<5>257-265
<6>1966
<7>Mutants of P1 that either fail to cause restriction, r-m, or neither cause
restriction nor modification, r-m-, were used to study the phage-induced
modification of the bacterial chromosome of Escherichia coli K12 and the
effects of modification on P1 generalized transduction.  The findings made were
that: (1) the bacterial chromosomal markers transduced by P1 are not sensitive
to restriction whereas the bacterial chromosomal markers transduced by P1 r-m-
are sensitive to restriction; (2) the sensitivity to restriction differed for
different markers; (3) all the regions of the bacterial chromosome tested
appear to be modified; (4) modification can interfere with the transduction of
certain markers in a way not associated with protection against restriction;
(5) the modification of bacterial DNA can probably occur without DNA
replication; (6) the sensitivity of lambda DNA to restriction is much greater
than that of bacterial DNA when both are transferred by P1 transducing
particles.  The implications of the results and limitations in interpreting
them are discussed.

<>

<1>Ioerger, T.R., Feng, Y., Chen, X., Dobos, K.M., Victor, T.C., Streicher, E.M., Warren, R.M., Gey-van-Pittius, N.C., Van Helden, P.D., Sacchettini, J.C.
<2>The non-clonality of drug resistance in Beijing-genotype isolates of Mycobacterium tuberculosis from the Western Cape of South Africa.
<3>BMC Genomics
<4>11
<5>670
<6>2010
<7>BACKGROUND: The Beijing genotype of M. tuberculosis is a virulent strain that is
disseminating worldwide and has a strong association with drug resistance. In the
Western Cape of South Africa, epidemiological studies have identified the R220
cluster of the Beijing genotype as a major contributor to a recent outbreak of
drug-resistant tuberculosis. Although the outbreak is considered to be due to
clonal transmission, the relationship among drug resistant isolates has not yet
been established. RESULTS: To better understand the evolution of drug resistance
among these strains, 14 drug-resistant clinical isolates of the Beijing genotype
were sequenced by whole-genome sequencing, including eight from R220 and six from
a more ancestral Beijing cluster, R86, for comparison. While each cluster shares
a distinct resistance mutation for isoniazid, mapping of other drug-resistance
mutations onto a phylogenetic tree constructed from single nucleotide
polymorphisms shows that resistance mutations to many drugs have arisen multiple
times independently within each cluster of isolates. Thus, drug resistance among
these isolates appears to be acquired, not clonally derived. This observation
suggests that, although the Beijing genotype as a whole might have selective
advantages enabling its rapid dissemination, the XDR isolates are relatively less
fit and do not propagate well. Although it has been hypothesized that the
increased frequency of drug resistance in some Beijing lineages might be caused
by a mutator phenotype, no significant shift in synonymous substitution patterns
is observed in the genomes. CONCLUSION: While MDR-TB is spreading by transmission
in the Western Cape, our data suggests that further drug resistance (i.e. XDR-TB)
at this stage is acquired.

<>

<1>Iordanescu, S., Surdeanu, M.
<2>Two restriction and modification systems in Staphylococcus aureus NCTC8325.
<3>J. Gen. Microbiol.
<4>96
<5>277-281
<6>1976
<7>The presence of two distinct host specificities in Staphylococcus aureus strain NCTC8325 was
revealed by the isolation of restriction- and modification-deficient mutants.  The two host
specificity systems, designated S1 and S2, are both active on phage 80 Mu Alpha but are not
additive in their restricting activity.  Restriction-deficient, modification-proficient
mutants were invariably affected in both restriction systems.  The functional relationship
between these two systems is discussed.

<>

<1>Ip, M., Wang, Z., Lam, W.Y., Zhou, H., Tsui, S.
<2>Draft Genome Sequence of Methicillin-Resistant Staphylococcus aureus CUHK_188 (ST188), a Health Care-Associated Bacteremic Isolate from Hong Kong.
<3>Genome Announcements
<4>2
<5>e00255-14
<6>2014
<7>We report the draft genome sequence of a methicillin-resistant Staphylococcus aureus strain
designated CUHK_188, isolated from a bacteremic patient undergoing treatment at a university
teaching hospital in Hong Kong. This strain belongs to sequence type 188 (ST188), with spa
type t189 and staphylococcal cassette chromosome mec type V.

<>

<1>Iraola, G., Perez, R., Naya, H., Paolicchi, F., Harris, D., Lawley, T.D., Rego, N., Hernandez, M., Calleros, L., Carretto, L., Velilla, A., Morsella, C., Mendez, A., Gioffre, A.
<2>Complete Genome Sequence of Campylobacter fetus subsp. venerealis Biovar Intermedius, Isolated from the Prepuce of a Bull.
<3>Genome Announcements
<4>1
<5>e00526-13
<6>2013
<7>Campylobacter fetus subsp. venerealis is the causative agent of bovine genital
campylobacteriosis, a sexually transmitted disease distributed worldwide. Campylobacter fetus
subsp. venerealis biovar Intermedius strains differ in their biochemical behavior and are
prevalent in some countries. We report the first genome sequence for this biovar, isolated
from bull prepuce.

<>

<1>Irisawa, T., Oshima, K., Suda, W., Kitahara, M., Sakamoto, M., Kitamura, K., Iida, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequence of Lactobacillus sucicola JCM 15457T, a Motile Lactic Acid  Bacterium Isolated from Oak Sap.
<3>Genome Announcements
<4>2
<5>e00403-14
<6>2014
<7>Here, we report the draft genome sequence of a motile lactic acid bacterium, Lactobacillus
sucicola JCM 15457(T), isolated from oak sap. Motility-related
genes and their organization in the annotated genome were broadly similar to
those in the sequenced genomes of related lactobacilli.

<>

<1>Irlinger, F., Loux, V., Bento, P., Gibrat, J.F., Straub, C., Bonnarme, P., Landaud, S., Monnet, C.
<2>Genome Sequence of Staphylococcus equorum subsp. equorum Mu2, Isolated from a French Smear-Ripened Cheese.
<3>J. Bacteriol.
<4>194
<5>5141-5142
<6>2012
<7>Staphylococcus equorum subsp. equorum is a member of the coagulase-negative staphylococcus
group and is frequently isolated from fermented food products and
from food-processing environments. It contributes to the formation of aroma
compounds during the ripening of fermented foods, especially cheeses and
sausages. Here, we report the draft genome sequence of Staphylococcus equorum
subsp. equorum Mu2 to provide insights into its physiology and compare it with
other Staphylococcus species.

<>

<1>Irma-Syakina, M.Z., Teh, L.K., Salleh, M.Z.
<2>Draft Genome Sequence of Streptococcus agalactiae PR06.
<3>Genome Announcements
<4>1
<5>e00351-13
<6>2013
<7>Streptococcus agalactiae (group B streptococcus [GBS]) is a Gram-positive bacterium that was
first recognized as a causative agent of bovine mastitis. S. agalactiae has subsequently
emerged as a significant cause of human diseases. Here, we report the draft genome sequence of
S. agalactiae PR06, which was isolated from a septicemic patient in a local hospital in
Malaysia.

<>

<1>Irschik, H., Kopp, M., Weissman, K.J., Buntin, K., Piel, J., Muller, R.
<2>Analysis of the sorangicin gene cluster reinforces the utility of a combined phylogenetic/retrobiosynthetic analysis for deciphering natural product assembly by trans-AT PKS.
<3>Chembiochem
<4>11
<5>1840-1849
<6>2010
<7>The sorangicins are a group of polyketide antibiotics produced by several strains of the
myxobacterium Sorangium cellulosum, whose activity is derived from inhibition of eubacterial
RNA polymerase.  The core structure of sorangicins A and B, the metabolites of highest
abundance, comprises a 31-membered lactone that exhibits many unusual features, including a
tetra-substituted terahydropyran, a trisubstituted dihydropyran, and a signature C31-C36
bicyclic ether moiety; sorangicin B lacks a hydroxyl group at C22 relative to sorangicin A.
The sorangicins are the most potent myxobacterial antibiotics identified to date, exhibiting
activity against both Gram-positive and Gram-negative species.  The corresponding glycosides
are only poorly active, however, suggesting that this modification might serve to protect S.
cellulosum from its own antibiotics.

<>

<1>Isakovic, L., Saavedra, O.M., Llewellyn, D.B., Claridge, S., Zhan, L.J., Bernstein, N., Vaisburg, A., Elowe, N., Petschner, A.J., Rahil, J., Beaulieu, N., Gauthier, F., MacLeod, A.R., Delorme, D., Besterman, J.M., Wahhab, A.
<2>Constrained (L-)-S-adenosyl-L-homocysteine (SAH) analogues as DNA methyltransferase inhibitors.
<3>Bioorg. Med. Chem. Lett.
<4>19
<5>2742-2746
<6>2009
<7>Potent SAH analogues with constrained homocysteine units have been designed and synthesized as
inhibitors of human DNMT enzymes. The five
membered (2S,4S)-4-mercaptopyrrolidine-2-carboxylic acid, in 1a, was a
good replacement for homocysteine, while the corresponding six-member
counterpart was less active. Further optimization of 1a, changed the
selectivity pro. le of these inhibitors. A Chloro substituent at the
2-position of 1a, compound 1d, retained potency against DNMT1, while
N-6 alkylation, compound 7a, conserved DNMT3b2 activity. The
concomitant substitutions of 1a at both 2- and N-6 positions reduced
activity against both enzymes.

<>

<1>Isalan, M., Choo, Y.
<2>Engineered Zinc Finger Proteins that Respond to DNA Modification by HaeIII and HhaI Methyltransferase Enzymes.
<3>J. Mol. Biol.
<4>295
<5>471-477
<6>2000
<7>Zinc finger modules are capable of specifically interacting with DNA that contains
5-methylcytosine (5-mC) in place of cytosine, suggesting that zinc finger-DNA binding could be
regulated by extrinsic methylation of DNA. Here, we have used phage display to engineer zinc
finger proteins that detect and discriminate DNA methylation by the prokaryotic enzymes HaeIII
and HhaI. In these systems, zinc finger-DNA complexes are induced by DNA modification using
the appropriate enzyme, which can therefore act as a switch. To further develop the
specificity of the switch, zinc finger discrimination between 5-mC and thymine in DNA
sequences is demonstrated despite the presence of the characteristic major groove methyl group
that is common to both bases. Specificity was achieved using a DNA-binding strategy involving
synergy between adjacent zinc fingers. We propose that engineered zinc fingers that recognise
particular DNA modifications, such as sequence-specific DNA methylation, could be integrated
into artificial regulatory circuits for the control of gene expression and other biological
processes.

<>

<1>Isanapong, J. et al.
<2>High-Quality Draft Genome Sequence of the Opitutaceae Bacterium Strain TAV1, a Symbiont of the Wood-Feeding Termite Reticulitermes flavipes.
<3>J. Bacteriol.
<4>194
<5>2744-2745
<6>2012
<7>Microbial communities in the termite hindgut are essential for degrading plant material. We
present the high-quality draft genome sequence of the Opitutaceae
bacterium strain TAV1, the first member of the phylum Verrucomicrobia to be
isolated from wood-feeding termites. The genomic analysis reveals genes coding
for lignocellulosic degradation and nitrogen fixation.

<>

<1>Ishaq, M., Kaji, A.
<2>Mechanism of T4 phage restriction by plasmid Rts 1. Cleavage of T4 phage DNA by Rts 1-specific enzyme.
<3>J. Biol. Chem.
<4>255
<5>4040-4047
<6>1980
<7>Rts 1 is a plasmid which confers upon host bacteria the capacity to restrict T4 phage growth
at 32°C but not at 42°C. This restriction was not dependent on cellular adenyl cyclase, while
other phenotypes of Rts 1, such as the temperature-sensitive effect on host growth, were
dependent on this enzyme. At 32"C, in Escherichia coli 2OSO/Rts 1 cells, 55 to 60% of the
parental T4 phage DNA became acid-soluble within 5 to 10 min after infection,
while in E. coli JC7623/Rts 1 cells, which lack exonuclease I and V, very little T4 DNA became
acidsoluble.  The capacity of E. coli 2OSO/Rts 1 to degrade infecting T, DNA decreased 70%
within 15 min after shifting the temperature to 42°C. Temperature shift up and down
experiments in the presence of chloramphenicol showed that the protein responsible for T, DNA
cleavage is thermosensitive, and once it is denatured it cannot regain its activity by
lowering the temperature to 32°C in vivo. Alkaline sucrose gradient centrifugation revealed
that the infecting Tq DNA was nicked in the presence of Rts 1 at 32"C, but not at 42°C in E.
coli JC7623. The cell-free extract of E. coli JC7623/Rts 1 grown at 32°C contained an enzyme
which converted Tq DNA to smaller but acid-insoluble fragments. This enzyme could not be
detected in E. coli JC7623o r E. coli JC7623/Rts 1 grown at 42°C. The enzyme was rapidly
inactivated when incubated at 42°C in vitro. Analysis of the reaction product by gel
electrophoresis and alkaline and sodium dodecyl sulfate-neutral sucrose density gradient
centrifugation, showed marked heterogeneity of the in vitro reaction product. This enzyme did
not cleave either T7 DNA or nonglucosylated T4 DNA under the same experimental conditions.
Only M$+ was required for its activity. These experiments suggest that the plasmid Rts 1 codes
for a thermosensitive enzyme which specifically degrades glucosylated T, DNA, resulting in the
restriction of T, phage growth at 32°C but not at 42°C. This enzyme, named Rts 1 endonuclease,
was purified approximately 500-fold.

<>

<1>Ishida, C., Ura, K., Hirao, A., Sasaki, H., Toyoda, A., Sakaki, Y., Niwa, H., Li, E., Kaneda, Y.
<2>Genomic organization and promoter analysis of the Dnmt3b gene.
<3>Gene
<4>310
<5>151-159
<6>2003
<7>The Dnmt3b gene encodes a de novo DNA methyltransferase that is essential for normal mouse
development. It is highly expressed in early embryos and
embryonic stem (ES) cells but downregulated in most adult somatic tissues.
To gain insight into the regulation of Dnmt3b, we have isolated a mouse
genomic bacterial artificial chromosome clone that contains the Dnmt3b
gene. Complete sequence analysis of the clone demonstrated that Dnmt3b
consists of at least 24 exons and spans 38 kilobases. S1 nuclease analysis
identified two adjacent transcriptional start sites located downstream of
a unique TATA-like element in a CpG island. There was an unknown gene
which we named mU(3) 17 kb upstream of the Dnmt3b locus, and it was
transcribed ubiquitously and in the opposite direction of Dnmt3b.
Transfection analysis revealed that the minimal promoter region containing
an Sp1 site was active even in somatic cells, and that there were several
repressor elements within 7.9 kb upstream of Dnmt3b downregulated this
gene specifically in somatic cells but not in ES cells. These findings
provide a basis for future detailed studies of the mechanisms controlling
Dnmt3b expression.

<>

<1>Ishii, S., Tago, K., Nishizawa, T., Oshima, K., Hattori, M., Senoo, K.
<2>Complete Genome Sequence of the Denitrifying and N2O-Reducing Bacterium Pseudogulbenkiania sp. Strain NH8B.
<3>J. Bacteriol.
<4>193
<5>6395-6396
<6>2011
<7>Pseudogulbenkiania sp. strain NH8B is a Neisseriales bacterium isolated from an agricultural
field. This strain has strong denitrification and
N(2)O reduction activities. Here, we report the finished and annotated
genome sequence of this organism.

<>

<1>Ishikawa, F.
<2>DNA methyltransferase.
<3>Japanese Patent Office
<4>JP 11187881
<5>
<6>1999
<7>Problem to be solved: To obtain the subject novel enzyme, having a specific amino acid
sequence, composed of a polypeptide functioning as DNA methyltransferase, capable of de novo
methylation of cytosine having a DNA strand with the aid of a mammalian cell for treatment of
tumor.  Solution: This enzyme is specified by an amino acid sequence of amino acid no. 1 to
391 shown by the formula, being a novel polypeptide functioning as DNA methyltransferase,
capable of de novo methylation of cytosine having a DNA strand with the aid of a mammalian
cell, and useful, e.g. for diagnosis and treatment of tumor.  This polypeptide is obtained by
plaque hybridization to screen a human ovarian cDNA library, using a cCNA library, as a
temperature, prepared from a cell strain PA-1 derived from human ovarian embryoma, and a DNA
fragment, as the probe, prepared by cloning by PVR using primers each having an EcoRI- or
XhoI-recognizing sequence at he 5' terminal.

<>

<1>Ishikawa, J., Yamashita, A., Mikami, Y., Hoshino, Y., Kurita, H., Hotta, K., Shiba, T., Hattori, M.
<2>The complete genomic sequence of Nocardia farcinica IFM 10152.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>14925-14930
<6>2004
<7>We determined the genomic sequence of Nocardia farcinica IFM 10152, a clinical isolate, and
revealed the molecular basis of its versatility.  The genome consists of a single circular
chromosome of 6,021,225 bp with an average G+C content of 70.8% and two plasmids of 184,027
(pNF1) and 87,093 (pNF2) bp with average G+C contents of 67.2% and 68.4%, respectively.  The
chromosome encoded 5,674, putative protein-coding sequences, including many candidate genes
for virulence and multidrug resistance as well as secondary metabolism.  Analyses of
paralogous protein families suggest that gene duplications have resulted in a bacterium that
can survive not only in soil environments but also in animal tissues, resulting in disease.

<>

<1>Ishikawa, K., Fukuda, E., Kobayashi, I.
<2>Conflicts targeting epigenetic systems and their resolution by cell death: novel concepts for methyl-specific and other restriction systems.
<3>DNA Res.
<4>17
<5>325-342
<6>2010
<7>Epigenetic modification of genomic DNA by methylation is important for defining the epigenome
and the transcriptome in eukaryotes as well as in prokaryotes. In prokaryotes, the DNA
methyltransferase genes often vary, are mobile, and are paired with the gene for a restriction
enzyme. Decrease in a certain epigenetic methylation may lead to chromosome cleavage by the
partner restriction enzyme, leading to eventual cell death. Thus, the pairing of a DNA
methyltransferase and a restriction enzyme forces an epigenetic state to be maintained within
the genome. Although restriction enzymes were originally discovered for their ability to
attack invading DNAs, it may be understood because such DNAs show deviation from this
epigenetic status. DNAs with epigenetic methylation, by a methyltransferase linked or unlinked
with a restriction enzyme, can also be the target of DNases, such as McrBC of Escherichia
coli, which was discovered because of its methyl-specific restriction. McrBC responds to
specific genome methylation systems by killing the host bacterial cell through chromosome
cleavage. Evolutionary and genomic analysis of McrBC homologues revealed their mobility and
wide distribution in prokaryotes similar to restriction-modification systems. These findings
support the hypothesis that this family of methyl-specific DNases evolved as mobile elements
competing with specific genome methylation systems through host killing. These restriction
systems clearly demonstrate the presence of conflicts between epigenetic systems.

<>

<1>Ishikawa, K., Handa, N., Kobayashi, I.
<2>Cleavage of a model DNA replication fork by a Type I restriction endonuclease.
<3>Nucleic Acids Res.
<4>37
<5>3531-3544
<6>2009
<7>Cleavage of a DNA replication fork leads to fork restoration by recombination repair. In
prokaryote cells carrying
restriction-modification systems, fork passage reduces genome methylation
by the modification enzyme and exposes the chromosome to attack by the
restriction enzyme. Various observations have suggested a relationship
between the fork and Type I restriction enzymes, which cleave DNA at a
distance from a recognition sequence. Here, we demonstrate that a Type I
restriction enzyme preparation cleaves a model replication fork at its
branch. The enzyme probably tracks along the DNA from an unmethylated
recognition site on the daughter DNA and cuts the fork upon encountering
the branch point. Our finding suggests that these restriction-modification
systems contribute to genome maintenance through cell death and indicates
that DNA replication fork cleavage represents a critical point in genome
maintenance to choose between the restoration pathway and the destruction
pathway.

<>

<1>Ishikawa, K., Handa, N., Kobayashi, I.
<2>Coupling of DNA replication and restriction: A type I restriction enzyme cleaves branched DNAs mimicing replication forks, in vitro.
<3>Genes Genet. Syst.
<4>83
<5>510
<6>2008
<7>Coupling of DNA replication and restriction: A type I restriction enzyme cleaves branched DNAs
mimicing replication forks, in vitro.  DNA cleavage by a restriction enzyme is prevented by
methylation on the recognition sequence.  It has been imagined that DNA replication and
restriction cleavage are coupled because unmethylated site will appear through replication
fork passage.  In fact, we previously observed an example suggesting such a coupling between
phage DNA replication and restriction, in vivo.  In the present study, we show that a purified
type I restriction endonuclease cleaves DNAs with long branch mimicking replication forks, in
the vicinity of the branch point dependent on an unmethylated recognition sequence.
Relationship between type I restriction enzymnes and other enzymes known that they cleave
branched DNA will be discussed.

<>

<1>Ishikawa, K., Handa, N., Sears, L., Raleigh, E.A., Kobayashi, I.
<2>Cleavage of a model DNA replication fork by a methyl-specific endonuclease.
<3>Nucleic Acids Res.
<4>39
<5>5489-5498
<6>2011
<7>Epigenetic DNA methylation is involved in many biological processes. An epigenetic status can
be altered by gain or loss of a DNA methyltransferase gene or its activity. Repair of DNA
damage can also remove DNA methylation. In response to such alterations, DNA endonucleases
that sense DNA methylation can act and may cause cell death. Here, we explored the possibility
that McrBC, a methylation-dependent DNase of Escherichia coli, cleaves DNA at a replication
fork. First, we found that in vivo restriction by McrBC of bacteriophage carrying a foreign
DNA methyltransferase gene is increased in the absence of homologous recombination. This
suggests that some cleavage events are repaired by recombination and must take place during or
after replication. Next, we demonstrated that the enzyme can cleave a model DNA replication
fork in vitro. Cleavage of a fork required methylation on both arms and removed one, the other
or both of the arms. Most cleavage events removed the methylated sites from the fork. This
result suggests that acquisition of even rarely occurring modification patterns will be
recognized and rejected efficiently by modification-dependent restriction systems that
recognize two sites. This process might serve to maintain an epigenetic status along the
genome through programmed cell death.

<>

<1>Ishikawa, K., Watanabe, M., Kuroita, T., Uchiyama, I., Bujnicki, J., Kawakami, B., Tanokura, M., Kobayashi, I.
<2>Discovery of a novel restriction endonuclease by genome comparison and application of a wheat-germ-based cell-free translation assay: PabI (5'GTA/C) from the hyperthermophilic archaeon Pyrococcus abyssi.
<3>Genes Genet. Syst.
<4>80
<5>454
<6>2005
<7>To search for restriction endonucleasees, we used a novel plant-based cell-free translation
procedure that bypasses the toxicity of these enzymes. To identify candidate genes, the
related genomes of the hyperthermophilic archaea Pyrococcus abyssi and Pyrococcus horikoshii
were compared. In line with the selfish mobile gene hypothesis for restriction-modification
systems, apparent genome rearrangement around putative restriction genes serve as a selecting
criterion.  Several candidate restriction genes were identified and expressed with a wheat
germ-based cell-free protein synthesis system.  The resulting solution could be directly
assayed for restriction activity.  We identified two deoxyribonucleases.  The novel enzymes
was denoted as PabI, purified and found to recognize 5'GTAC and leave a 3TA overhang
(5'GTA/C), a novel restriction enzyme-generated terminus.  PabI is active up to 90oC and
optimally active at a pH of around 6 and in NaCl concentrations ranging from 100 to 200 mM.
We predict that it has a novel three-dimensional structure.

<>

<1>Ishikawa, K., Watanabe, M., Kuroita, T., Uchiyama, I., Bujnicki, J.M., Kawakami, B., Tanokura, M., Kobayashi, I.
<2>Discovery of a novel restriction endonuclease by genome comparison and application of a wheat-germ-based cell-free translation assay: PabI   (5'-GTA/C) from the hyperthermophilic archaeon Pyrococcus abyssi.
<3>Nucleic Acids Res.
<4>33
<5>e112
<6>2005
<7>To search for restriction endonucleases, we used a novel plant-based cell-free translation
procedure that bypasses the toxicity of these
enzymes. To identify candidate genes, the related genomes of the
hyperthermophilic archaea Pyrococcus abyssi and Pyrococcus horikoshii were
compared. In line with the selfish mobile gene hypothesis for
restriction-modification systems, apparent genome rearrangement around
putative restriction genes served as a selecting criterion. Several
candidate restriction genes were identified and then amplified in such a
way that they were removed from their own translation signal. During their
cloning into a plasmid, the genes became connected with a plant
translation signal. After in vitro transcription by T7 RNA polymerase, the
mRNAs were separated from the template DNA and translated in a
wheat-germ-based cell-free protein synthesis system. The resulting
solution could be directly assayed for restriction activity. We identified
two deoxyribonucleases. The novel enzyme was denoted as PabI, purified and
found to recognize 5'-GTAC and leave a 3'-TA overhang (5'-GTA/C), a novel
restriction enzyme-generated terminus. PabI is active up to 90 degrees C
and optimally active at a pH of around 6 and in NaCl concentrations
ranging from 100 to 200 mM. We predict that it has a novel 3D structure.

<>

<1>Ishikawa, T., Yamada, Y.
<2>Purification of restriction endonuclease from Acetobacter pasteurianus IFO 13752 (ApaORI) and its properties.
<3>J. Gen. Appl. Microbiol.
<4>36
<5>127-135
<6>1990
<7>A restriction endonuclease, designated ApaORI, was purified from cell-free
extracts of A. pasteurianus IFO 13752 by streptomycin treatment, ammonium
sulfate fractionation, phosphocellulose treatment, hydroxylapatite and
heparin-Sepharose CL-6B column chromatography, and gel filtration using a
Superose 12 (HR10/30) column.  The purified enzyme was homogeneous on
polyacrylamide gel disc electrophoresis.  The molecular weight of the purified
enzyme was found by gel filtration to be 21,000 daltons.  The isoelectric point
of the purified enzyme was neutral.  The purified enzyme cleaved lambda,
PhiX174 RF I, M13mp7 RF I, and pBR322 DNAs at 18 or more, 2, 4 or more, and 4
or more sites respectively.  The purified enzyme worked best at 37C and pH 7.5
in a reaction mixture (50 micro liters) containing 1.0 microgram lambda DNA, 10
mM Tris-HCl, 7 mM 2-mercaptoethanol, 7 mM MgCl2, and 50 mM NaCl.  However, the
purified enzyme did not require NaCl for its reaction.  The purified enzyme
recognizes the palindromic pentanucleotides 5'-CC (A or T)GG-3' and cuts
between C and A (orT), producing 5'-cohesive mononucleotide extensions
(isoschizomer of BstNI).

<>

<1>Ishino, Y., Komori, K.
<2>DNA and protein sequences of Pyrococcus furiosus heat stable restriction endonuclease Hef and the use of the enzyme for DNA damage  repairing.
<3>Japanese Patent Office
<4>JP 2003169678 A
<5>18
<6>2003
<7>Aim.  We find protein that recognizes specific DNA structure generated by the damage.  This
protein could contribute to the process of DNA repair.  Methods.  The source of this protein
could be either (1) the thermostable protein derived from the Archaebacteria, or (2) the
nuclear protein.  The protein, which contains one or several amino-acid deletion, mutation, or
addition, has endonuclease activity that specifically cleave 3~6 base (5') upstream from the
DNA strand containing nick or gap.

<>

<1>Ishino, Y., Nomura, Y., Kato, I.
<2>Sse8647I, a new type II restriction endonuclease from a Streptomyces species cutting at 5'-AG/GWCCT-3'.
<3>Nucleic Acids Res.
<4>23
<5>742-744
<6>1995
<7>We isolated and characterized a new type II restriction endonuclease which recognizes the
palindromic heptanucleotide sequence 5'-AGGWCCT-3' and cleaves double-stranded DNA after the
first G in the sequence from a microorganism belonging to Streptomyces species. This enzyme
cleaves adenovirus 2 DNA at eight sites, but does not cleave lambda phage, pBR322, pUC18 and
19, M13mp18 and 19, SV40, ColE1 and omega-X174 DNAs.

<>

<1>Ishiwata, N., Tanimura, A., Miyahara, M., Mise, K.
<2>Studies on restriction endonucleases of bacteria causing peroral infectious diseases.  I. Characterization of Hsd plasmid in Salmonella typhi.
<3>Shokuhin Eiseigaku Zasshi
<4>36
<5>404-408
<6>1995
<7>An Hsd (host specificity for DNA) plasmid designated pSTd4 was found in Salmonella Typhi D4,
one of the standard phage typing strains.  The plasmid was introduced into E. coli K-12, and
the restriction map of this plasmid was constructed.  The efficiency of plating of lambda-O
phage was drastically reduced with pSTd4 as well as other small Hsd plasmids of
Enterobacteriaceae origin.  The results clearly indicate that pSTd4 is a type-determining
plasmid in S. Typhi D4.  We propose that rare phage types such as D4 carrying type-determining
plasmids or lysogenic phages should be eliminated from the collection of standard phage
typing strains of S. Typhi.

<>

<1>Ishmori, Y.
<2>Monitoring restriction enzyme reactions by cleaving gene with restriction enzyme, measuring amount of physico-chemical change produced as index of reaction processing.
<3>Japanese Patent Office
<4>JP 92320698
<5>
<6>1992
<7>In monitoring restriction enzyme reaction by cleaving gene with restriction enzyme, the amount
of physicochemical change produced is measured as an index of the processing state of the
reaction.
USE/ADVANTAGE - Monitoring of restriction enzyme proceeeds in a short time. Process is safe,
exact, simple and highly sensitive. When restriction enzyme is immobilised to carrier,
separationn of restriction enzyme after reaction is possible, application to automatic gene
detector becomes possible, further, running cost per reaction is decreased drastically, by
repeated use of restriction enzyme.

<>

<1>Ismail, A., Teh, L.K., Ngeow, Y.F., Norazmi, M.N., Zainul, Z.F., Tang, T.H., Najimudin, N., Salleh, M.Z.
<2>Draft Genome Sequence of a Clinical Isolate of Mycobacterium tuberculosis Strain  PR05.
<3>Genome Announcements
<4>1
<5>e00397-13
<6>2013
<7>We report the annotated genome sequence of a clinical isolate, Mycobacterium tuberculosis
strain PR05, which was isolated from the human cerebrospinal fluid
of a patient diagnosed with tuberculosis.

<>

<1>Isokpehi, R.D., Coker, A.O.
<2>In silico evidence for horizontal gene transfer of type II restriction-modification genes in Helicobacter pylori.
<3>Biochem. Soc. Trans.
<4>28
<5>A185
<6>2000
<7>Helicobacter pylori is a gram-negative bacteria associated with peptic ulcer disease and a
risk factor for gastric cancer.  The organism is the first for which the complete genome
sequences of two unrelated strains, 26695 and J99, has been published.  Analyses of these
genomes have identified a high number of genes with homology to restriction and modification
enzymes, many of which are strain specific.  Restriction-modification genes are widely
distributed in bacteria and function in defense against invasion by foreign DNA.  Recent
evidence also suggests their involvement in genomic rearrangement and evolution.  In order to
gain insight into the role of R-M genes in H. pylori evolution, this study examined G+C
content and codon bias of type II R-M genes in J99.  This strain consists of 4 paired type II
R-M systems as well as 1 unpaired restriction enzyme and 15 unpaired modification enzymes.
Compositional features of the codon positions for all 24 genes were determined with the
program CODONTREE.  Overall G+C content in paired genes is higher for modification genes than
restriction genes, suggesting a possible stepwise evolutionary process.  Three restriction and
2 modification genes significantly deviate (P<0.05) in G+C content relative to the mean G+C
content of the H. pylori genome.  Lastly, a distance matrix based on codon usage in paired R-M
system genes was generated to test the hypothesis that a restriction gene is more closely
related to its associated modification gene than to the modification genes of other paired R-M
systems.  A UPGMA distance tree generated by the program NEIGHBOUR supports this hypothesis.
These results provide in silico evidence that type II R-M genes in H. pylori J99, may have
been acquired through horizontal gene transfer.

<>

<1>Isokpehi, R.D., Coker, A.O.
<2>Possible stepwise evolution of type II restriction-modification genes in Helicobacter pylori genome.
<3>Int. J. Med. Microbiol.
<4>291S
<5>94
<6>2001
<7>Helicobacter pylori is a gram-negative bacteria associated with peptic ulcer disease.  The
complete genome sequences of strains 26695 and J99 of the organism have been published and
compared.  Analyses of these genomes have identified a high number of genes with homology to
restriction and modification enzymes, many of which are strain specific and may have been
acquired by horizontal gene transfer.  Restriction-Modification (R-M) genes are widely
distributed in bacteria and function in defense against invasion by foreign DNA.  Recent
evidence also suggests their involvement in genomic rearrangement and evolution.  In H.
pylori, the modification genes may also regulate gene expression.  Variation in base
composition of genes can provide evidence of the order of introduction of genes into a genome.
Thus, in order to gain insight into the evolution of R-M genes in H. pylori genome, this study
examined guanine and cytosine (G+C) content of the type II restriction genes (5 in 26695, 4 in
J99) and modification genes (17 in 26695, 20 in J99).  In both strains, 4 pairs of R-M genes
exist, two of which are unique to each strain.  All the type II R-M genes were retrieved from
their respective databases and compositional features of the codon position were determined
using a computer program CODONTREE.  In all the paired R-M genes, the overall G+C content of
the modification gene is greater than its associated restriction gene, suggesting a possible
stepwise evolutionary process.  Further studies such as time of introgression and amelioration
of R-M genes in the genome could confirm their stepwise evolution.

<>

<1>Ito, H., Sadaoka, A.
<2>Hinc II restriction-modification system enzyme gene prodn. - comprises cleaving plasmid vector and introducing chromosome DNA extracted from E. coli to form transformant.
<3>Japanese Patent Office
<4>JP 3247279, JP 1991247279 A
<5>
<6>1991
<7>The following are claimed: (1) Purified Hinc II restriction enzyme produced by recombinant.
(2) Hinc II restriction modification system enzyme gene. (3) Prepn. for Hinc II restriction
enzyme and/or modified enzyme by culturing gene of (2) integrated plasmid retaining microbe,
next, by collecting Hinc II restriction enzyme and/or modified enzyme from the culture.
USE/ADVANTAGE - Hinc II restriction enzyme and/or modified enzyme can be produced efficiently.
In an example, new microbe e.g. E coli was obtd. as follows: (1) Chromosome DNA was extracted
from Hinc II producing bacteria as DNA donor, partially cleavaged by restriction enzyme, and
DNA fragment was recovered. (2) Plasmid vector was cleavaged by restriction enzyme; to
cleavaged site, DNA fragment obtd. in (1) was ligated. (3) DNA obtd. in (2) was introduced in
E. coli to obtain transformant; objective DNA contg. library was prepd. and corresponding
modified enzyme gene contg. clone was isolated. (4) Restriction enzyme activity of clone obtd.
in (3) was analysed, and both restriction modification system gene integrated clone was
selected. (5) When both genes integrated clone was not selected, modified methylase gene
contg. clone's DNA fragment was prepared, hybridisation was proceeded to restriction enzyme
decomposite of chromosome DNA by using it as probe, for selection of both genes integrated DNA
fragment. (6) From DNA fragment obtd. in (5), restriction enzyme gene contg. DNA fragment and
modified enzyme gene contg. DNA fragment were prepd., each bonded to independent replication
origin and antibiotic resistant gene having plasmid respectively, introduced to host at the
same time to obtain transformant.

<>

<1>Ito, H., Sadaoka, A., Kotani, H., Hiraoka, N., Nakamura, T.
<2>Cloning, nucleotide sequence, and expression of the HincII restriction-modification system.
<3>Nucleic Acids Res.
<4>18
<5>3903-3911
<6>1990
<7>Two genes, coding for the HincII from Haemophilus influenzae Rc
restriction-modification system, were cloned and expressed in Escherichia coli
RR1.  Their DNA sequences were determined.  The HincII methylase (M.HincII)
gene was 1,506 base pairs (bp) long, corresponding to a protein of 502 amino
acid residues (Mr=55,330).  The HincII endonuclease (R.HincII) gene was 774 bp
long, corresponding to a protein of 258 amino acid residues (Mr=28,490).  The
amino acid residues predicted from the R.HincII and the N-terminal amino acid
sequence of the enzyme found by analysis were identical.  These methylase and
endonuclease genes overlapped by 1 bp on the H. influenzae Rc chromosomal DNA.
The clone, named E. coli RR1-Hinc, overproduced R.HincII.  The R.HincII
activity of this clone was 1,000-fold that from H. influenzae Rc.  The amino
acid sequence of M.HincII was compared with the sequences of four other
adenine-specific type II methylases.  Important homology was found between the
M.HincII and these other methylases.

<>

<1>Ito, H., Shimado, H., Kimizuka, F., Kato, I.
<2>HpaI restriction and modification enzyme genes: cloning and expression.
<3>Japanese Patent Office
<4>JP 03120672, JP 1993000082 A
<5>
<6>1993
<7>
<>

<1>Ito, H., Shimato, H., Sadaoka, A., Kotani, H., Kimizuka, F., Kato, I.
<2>Cloning and expression of the HpaI restriction-modification genes.
<3>Nucleic Acids Res.
<4>20
<5>705-709
<6>1992
<7>The genes from Haemophilus parainfluenzae encoding the HpaI
restriction-modification system were cloned and expressed in Escherichia coli.
From the DNA sequence, we predicted the HpaI endonuclease (R.HpaI) to have 254
amino acid residues (Mr 29,630) and the HpaI methyltransferase (M.HpaI) to have
314 amino acid residues (37,390).  The R.HpaI and M.HpaI genes overlapped by 16
base pairs on the chromosomal DNA.  The genes had the same orientation.  The
clone, named E. coli HB101-HPA2, overproduced R.HpaI.  R.HpaI activity from the
clone was 100-fold that from H. parainfluenzae.  The amino acid sequence of
M.HpaI was compared with those of other type II methyltransferases.

<>

<1>Ito, J., Kawamura, F., Duffy, J.J.
<2>Susceptibility of non-thymine containing DNA to four bacterial restriction endonucleases.
<3>FEBS Lett.
<4>55
<5>278-281
<6>1975
<7>None

<>

<1>Ito, J., Roberts, R.J.
<2>Unusual base sequence arrangement in phage Phi29 DNA.
<3>Gene
<4>5
<5>1-7
<6>1979
<7>Susceptibility of Bacillus subtilis phage Phi29 DNA to 34 different restriction
endonucleases was determined.  Three enzymes, BglI, XbaI and BstEII, were found
to cleave Phi29 DNA only once at specific sites.  The sites of these single
cleavages have been mapped.  Thirteen enzymes did not cut Phi29 DNA.  Phi29
HindIII DNA fragments inserted into pBR313 plasmid and propagated in
Escherichia coli, were resistant to these restriction endonucleases.  This
result suggests that the insusceptibility is due to the absence of the
nucleotide sequences on Phi29 recognized by the enzymes, and not to the
presence of modified nucleotides.

<>

<1>Ito, S., Shen, L., Dai, Q., Wu, S.C., Collins, L.B., Swenberg, J.A., He, C., Zhang, Y.
<2>Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine.
<3>Science
<4>333
<5>1300-1303
<6>2011
<7>5-methylcytosine (5mC) in DNA plays an important role in gene expression, genomic imprinting,
and suppression of transposable elements. 5mC can be converted to
5-hydroxymethylcytosine (5hmC) by the Tet (ten eleven translocation) proteins.
Here, we show that, in addition to 5hmC, the Tet proteins can generate
5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) from 5mC in an enzymatic
activity-dependent manner. Furthermore, we reveal the presence of 5fC and 5caC in
genomic DNA of mouse embryonic stem cells and mouse organs. The genomic content
of 5hmC, 5fC, and 5caC can be increased or reduced through overexpression or
depletion of Tet proteins. Thus, we identify two previously unknown cytosine
derivatives in genomic DNA as the products of Tet proteins. Our study raises the
possibility that DNA demethylation may occur through Tet-catalyzed oxidation
followed by decarboxylation.

<>

<1>Ito, T., Katayama, Y., Asada, K., Mori, N., Tsutsumimoto, K., Tiensasitorn, C., Hiramatsu, K.
<2>Structural comparison of three types of Staphylococcal cassette chromosome mec integrated in the chromosome in methicillin-resistant Staphylococcus aureus.
<3>Antimicrob. Agents Chemother.
<4>45
<5>1323-1336
<6>2001
<7>The beta-lactam resistance gene mecA of Staphylococcus aureus is carried by a novel mobile
genetic element, designated staphylococcal cassette chromosome mec (SCCmec), identified in the
chromosome of a Japanese methicillin-resistant S. aureus (MRSA) strain.  We now report
identification of two additional types of mecA-carrying genetic elements found in the MRSA
strains isolated in other countries of the world.  There were substantial differences in the
size and nucleotide sequences between the elements and the SCCmec.  However, new elements
shared the chromosomal integration site with the SCCmec.  Structural analysis of the new
elements revealed that they possessed all of the salient features of the SCCmec: conserved
terminal inverted repeats and direct repeats at the integration junction points, conserved
genetic organization around the mecA gene, and the presence of cassette chromosome recombinase
genes responsible for the movements of SCCmec.  The elements, therefore, were considered to
comprise the SCCmeg family of staphylococcal mobile genetic elements together with the
previously identified SCCmec.  Among 38 epidemic MRSA strains isolated in 20 countries, 34
were shown to possess one of the three typical SCCmec elements on the chromosome.  Our
findings indicated that there are at least three distinct MRSA clones in the world with
different types of SCCmec in their chromosome.

<>

<1>Ito, T., Ma, X.X., Takeuchi, F., Okuma, K., Yuzawa, H., Hiramatsu, K.
<2>Novel Type V Staphylococcal Cassette Chromosome mec Driven by a Novel Cassette Chromosome Recombinase, ccrC.
<3>Antimicrob. Agents Chemother.
<4>48
<5>2637-2651
<6>2004
<7>Staphylococcal cassette chromosome mec (SCCmec) is a mobile genetic
element composed of the mec gene complex, which encodes methicillin
resistance, and the ccr gene complex, which encodes the recombinases
responsible for its mobility. The mec gene complex has been classified
into four classes, and the ccr gene complex has been classified into three
allotypes. Different combinations of mec gene complex classes and ccr gene
complex types have so far defined four types of SCCmec elements. Now we
introduce the fifth allotype of SCCmec, which was found on the chromosome
of a community-acquired methicillin-resistant Staphylococcus aureus strain
(strain WIS [WBG8318]) isolated in Australia. The element shared the same
chromosomal integration site with the four extant types of SCCmec and the
characteristic nucleotide sequences at the chromosome-SCCmec junction
regions. The novel SCCmec carried mecA bracketed by IS431
(IS431-mecA-DeltamecR1-IS431), which is designated the class C2 mec gene
complex; and instead of ccrA and ccrB genes, it carried a single copy of a
gene homologue that encoded cassette chromosome recombinase. Since the
open reading frame (ORF) was found to encode an enzyme which catalyzes the
precise excision as well as site- and orientation-specific integration of
the element, we designated the ORF cassette chromosome recombinase C
(ccrC), and we designated the element type V SCCmec. Type V SCCmec is a
small SCCmec element (28 kb) and does not carry any antibiotic resistance
genes besides mecA. Unlike the extant SCCmec types, it carries a set of
foreign genes encoding a restriction-modification system that might play a
role in the stabilization of the element on the chromosome.

<>

<1>Ito, Y., Azuma, T., Ito, S., Suto, H., Miyaji, H., Yamazaki, Y., Kato, T., Kohli, Y., Keida, Y., Kuriyama, M.
<2>Sequence analysis and clinical significance of the iceA gene from Helicobacter pylori strains in Japan.
<3>J. Clin. Microbiol.
<4>38
<5>483-488
<6>2000
<7>The Helicobacter pylori iceA gene was recently identified as a genetic marker for the
development of peptic ulcer in a Western population.  To assess the significance of iceA
subtypes of H. pylori in relation to peptic ulcer, 140 Japanese clinical isolates (88 from
Fukui and 52 from Okinawa) were characterized.  Sequence analysis of the iceA1 gene from 25
representative Japanese strains was also carried out to identify the differences in iceA
between the ulcer group and the gastritis group.  The iceA1 genotype was not correlated with
the presence of peptide ulceration in either area.  In addition, sequence analysis led to
identification of five deletions and five point mutations (a nonsense mutation or a 1-bp
insertion) within the iceA1 open reading frame corresponding to previously published
sequences.  These mutations were identified in both clinical groups (ulcer and gastritis
groups) in each area.  Local DNA sequence analysis revealed that the endpoints of all five
deletions coincided with direct repeats.  We also found four strains that carried longer iceA1
open reading frames compared with that for strain 60190.  In conclusion, carriage of an iceA1
strain does not seem to be a risk factor for peptic ulcer in Japanese subjects.  The critical
mutations in the iceA1 gene in some isolates from patients with peptic ulcers suggested that
IceA does not participate in the pathogenesis of peptic ulcer in Japan.  We also found
deletion hot spots that were associated with direct repeats in iceA1 and that favored a
small-deletion model of slipped mispairing events during replication.  We showed that iceA1
sequence variations may be useful tools for analysis of the population genetics of H. pylori.

<>

<1>Itoh, T. et al.
<2>A 460-kb DNA sequence of the Escherichia coli K-12 genome corresponding to the 40.1-50.0 min region on the linkage map.
<3>DNA Res.
<4>3
<5>379-392
<6>1996
<7>The 465,813 base pair sequence corresponding to the 40.1-50.0 min region on the genetic map of
Escherichia coli K-12 (W3110) was determined. Analysis of the sequence revealed that this
region contained at least 466 potential open reading frames, of which 187 (40%) were
previously reported, 105 (23%) were homologous to other known genes, 103 (22%) were identical
or similar to hypothetical genes registered in databases, and the remaining 71 (15%) did not
show a significant similarity to any other gene. At the 45.2-46.0 min region, we found a very
large cluster of about 30 genes, whose functions are involved in the biosynthesis of
polysaccharides as the components of outer membranes. In addition, we identified a new
asn-tRNA gene, designated asnW, between the asnT and asnU genes and a new lysogenic phage
attachment site as the cis-element.

<>

<1>Ivancic-Bace, I., Vlasic, I., Cogelja-Cajo, G., Brcic-Kostic, K., Salaj-Smic, E.
<2>Roles of PriA protein and double-strand DNA break repair functions in UV-induced restriction alleviation in Escherichia coli.
<3>Genetics
<4>174
<5>2137-2149
<6>2006
<7>It has been widely considered that. DNA modification protects the chromosome of bacteria E.
coli K-12 against their own
restriction-modification systems. Chromosomal DNA is protected from
degradation by methylation of target sequences. However, when
unmethylated target sequences are generated in the host chromosome, the
endonuclease activity of the EcoKI restriction-modification enzyme is
inactivated by the ClpXP protease and DNA is protected. This process is
known as restriction alleviation (RA) and it can be induced by UV
irradiation (UV-induced RA). It has been proposed that chromosomal
Unmethylated target sequences, a signal for the cell to protect its own
DNA, can be generated by homologous recombination during the repair of
damaged DNA. In this study, we wanted to further investigate the
genetic requirements for recombination proteins involved in the
generation Of unmethylated target sequences. For this purpose, we
monitored the alleviation of EcoKI restriction by measuring the
survival of unmodified X in UV-irradiated cells. Our genetic analysis
showed that UV-induced RA is dependent on the excision repair protein
UvrA, the RecA-loading activity of the RecBCD enzyme, and the primosome
assembly activity of the PriA helicase and is partially dependent on
RecFOR proteins. On the basis of our results, we propose that
Unmethylated target sequences are generated at the D-loop by the strand
exchange of two hemi-methylated duplex DNAs and subsequent initiation
of DNA replication.

<>

<1>Ivancic-Jelecki, J., Baricevic, M., Santak, M., Forcic, D.
<2>Restriction enzyme cleavage of fluorescently labeled DNA fragments - Analysis of the method and its usage in examination of digestion  completeness.
<3>Anal. Biochem.
<4>349
<5>277-284
<6>2006
<7>Restriction enzymes have proven to be among the most valuable tools in molecular biology.  In
this work, we demonstrate that the cleavage of fluorescently labeled, PCR-amplified DNA can be
used as a simple and highly sensitive technique for detection of sequences present in a
percentage as low as 0.6% in a DNA pool.  Due to the fact that fluorescent labeling of DNA
fragments enables such sensitive detection and quantification of restriction enzyme cleavage,
the method was further exploited in monitoring of the enzymatic digestion completeness and in
determination of factors that influence restriction enzyme effectiveness.  We analyzed the
activity of six restriction endonucleases; the percentage of uncleaved DNA fragments
predominantly ranged between 2.0 and 2.5 and the highest value was 8.00%.  We conclude that,
since the enzymatic digestion completeness may not always be assured, each assay based on
restriction enzyme cleavage that is intended to be used in investigations of heterogeneity in
a DNA pool should be constructed so that the presence of cleaved sequences is the indication
of pool nonuniformity.  When the presence of uncleaved sequences indicates pool heterogeneity,
the results could be misleading due to possible incompleteness of enzymatic cleavage.

<>

<1>Ivanenko, T., Heitman, J., Kiss, A.
<2>Mutational analysis of the function of Met137 and Ile197, two amino acids implicated in sequence-specific DNA recognition by the EcoRI endonuclease.
<3>Biol. Chem.
<4>379
<5>459-465
<6>1998
<7>The gene encoding the EcoRI endonuclease was altered by site-directed mutagenesis to introduce
multiple substitutions of M137 and I197, two amino acids which were suggested by the revised
crystal structure to mediate recognition of the cytosines in the 5'-GAATTC-3' target
sequence.  Eight substitutions of M137 and ten substitutions of I197 were isolated.  With the
exception of M137W, M137P and M137K, all mutant enzymes retained enough activity to damage
cellular DNA in the absence of the EcoRI methyltransferase.  All M137 replacements abolished
the ability of the enzyme to restrict phage growth.  Conservative replacements at I197 (L, V)
did not impair phage restriction, whereas non-conservative changes reduced (G, W) or abolished
(D, P) restriction.  In general, substitutions at M137 were more deleterious than
substitutions at I197.  Double mutants with combinations of M137G/A and I197G/A mutations
exhibited a phenotype characteristic for the respective single M137 mutant.  Double mutants
carrying combinations of the M137G/A replacements and substitutions at R200 were viable even
in the absence of the methyltransferase, suggesting that disrupting contacts to both bases of
the GC base pair inactivates the enzyme.  None of the replacements resulted in relaxed
recognition specificity.  In summary, our findings are consistent with a role for M137 but do
not support such a role for I197 in substrate recognition by the EcoRI endonuclease.

<>

<1>Ivanova, E.P., Ng, H.J., Webb, H.K., Feng, G., Oshima, K., Hattori, M., Ohkuma, M., Sergeev, A.F., Mikhailov, V.V., Crawford, R.J., Sawabe, T.
<2>Draft Genome Sequences of Marinobacter similis A3d10T and Marinobacter salarius R9SW1T.
<3>Genome Announcements
<4>2
<5>e00442-14
<6>2014
<7>Here, we present the draft genomes of Marinobacter similis A3d10(T), a potential  plastic
biodegrader, and Marinobacter salarius R9SW1(T), isolated from
radioactive waters. This genomic information will contribute information on the
genetic basis of the metabolic pathways for the degradation of both plastic and
radionuclides.

<>

<1>Ivanova, N. et al.
<2>Genome sequence of Bacillus cereus and comparative analysis with Bacillus anthracis.
<3>Nature
<4>423
<5>87-91
<6>2003
<7>Bacillus cereus is an opportunistic pathogen causing food poisoning manifested by diarrhoeal
or emetic syndromes. It is closely related to the
animal and human pathogen Bacillus anthracis and the insect pathogen
Bacillus thuringiensis, the former being used as a biological weapon and
the latter as a pesticide. B. anthracis and B. thuringiensis are readily
distinguished from B. cereus by the presence of plasmid-borne specific
toxins (B. anthracis and B. thuringiensis) and capsule (B. anthracis). But
phylogenetic studies based on the analysis of chromosomal genes bring
controversial results, and it is unclear whether B. cereus, B. anthracis
and B. thuringiensis are varieties of the same species or different
species. Here we report the sequencing and analysis of the type strain B.
cereus ATCC 14579. The complete genome sequence of B. cereus ATCC 14579
together with the gapped genome of B. anthracis A2012 enables us to
perform comparative analysis, and hence to identify the genes that are
conserved between B. cereus and B. anthracis, and the genes that are
unique for each species. We use the former to clarify the phylogeny of the
cereus group, and the latter to determine plasmid-independent
species-specific markers.

<>

<1>Ivanova, N. et al.
<2>Complete genome sequence of the extremely halophilic Halanaerobium praevalens type strain (GSL).
<3>Standards in Genomic Sciences
<4>4
<5>312-321
<6>2011
<7>Halanaerobium praevalens Zeikus et al. 1984 is the type species of the genus Halanaerobium,
which in turn is the type genus of the family Halanaerobiaceae.
The species is of interest because it is able to reduce a variety of
nitro-substituted aromatic compounds at a high rate, and because of its ability
to degrade organic pollutants. The strain is also of interest because it
functions as a hydrolytic bacterium, fermenting complex organic matter and
producing intermediary metabolites for other trophic groups such as
sulfate-reducing and methanogenic bacteria. It is further reported as being
involved in carbon removal in the Great Salt Lake, its source of isolation. This
is the first completed genome sequence of a representative of the genus
Halanaerobium and the second genome sequence from a type strain of the family
Halanaerobiaceae. The 2,309,262 bp long genome with its 2,110 protein-coding and
70 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Ivanova, N. et al.
<2>Complete genome sequence of Truepera radiovictrix type strain (RQ-24).
<3>Standards in Genomic Sciences
<4>4
<5>91-99
<6>2011
<7>Truepera radiovictrix Albuquerque et al. 2005 is the type species of the genus Truepera within
the phylum 'Deinococcus/Thermus'. T. radiovictrix is of special
interest not only because of its isolated phylogenetic location in the order
Deinococcales, but also because of its ability to grow under multiple extreme
conditions in alkaline, moderately saline, and high temperature habitats. Of
particular interest is the fact that, T. radiovictrix is also remarkably
resistant to ionizing radiation, a feature it shares with members of the genus
Deinococcus. This is the first completed genome sequence of a member of the
family Trueperaceae and the fourth type strain genome sequence from a member of
the order Deinococcales. The 3,260,398 bp long genome with its 2,994
protein-coding and 52 RNA genes consists of one circular chromosome and is a part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Ivanova, N. et al.
<2>Complete genome sequence of Haliangium ochraceum type strain (SMP-2).
<3>Standards in Genomic Sciences
<4>2
<5>96-106
<6>2010
<7>Haliangium ochraceum Fudou et al. 2002 is the type species of the genus Haliangium in the
myxococcal family 'Haliangiaceae'. Members of the genus
Haliangium are the first halophilic myxobacterial taxa described. The cells of
the species follow a multicellular lifestyle in highly organized biofilms, called
swarms, they decompose bacterial and yeast cells as most myxobacteria do. The
fruiting bodies contain particularly small coccoid myxospores. H. ochraceum
encodes the first actin homologue identified in a bacterial genome. Here we
describe the features of this organism, together with the complete genome
sequence, and annotation. This is the first complete genome sequence of a member
of the myxococcal suborder Nannocystineae, and the 9,446,314 bp long single
replicon genome with its 6,898 protein-coding and 53 RNA genes is part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Ivanova, N. et al.
<2>Complete genome sequence of Geodermatophilus obscurus type strain (G-20).
<3>Standards in Genomic Sciences
<4>2
<5>158-167
<6>2010
<7>Geodermatophilus obscurus Luedemann 1968 is the type species of the genus, which  is the type
genus of the family Geodermatophilaceae. G. obscurus is of interest
as it has frequently been isolated from stressful environments such as rock
varnish in deserts, and as it exhibits interesting phenotypes such as lytic
capability of yeast cell walls, UV-C resistance, strong production of
extracellular functional amyloid (FuBA) and manganese oxidation. This is the
first completed genome sequence of the family Geodermatophilaceae. The 5,322,497
bp long genome with its 5,161 protein-coding and 58 RNA genes is part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Ivanova, N. et al.
<2>Complete genome sequence of Gordonia bronchialis type strain (3410).
<3>Standards in Genomic Sciences
<4>2
<5>19-28
<6>2010
<7>Gordonia bronchialis Tsukamura 1971 is the type species of the genus. G. bronchialis is a
human-pathogenic organism that has been isolated from a large
variety of human tissues. Here we describe the features of this organism,
together with the complete genome sequence and annotation. This is the first
completed genome sequence of the family Gordoniaceae. The 5,290,012 bp long
genome with its 4,944 protein-coding and 55 RNA genes is part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Ivanova, N. et al.
<2>Complete genome sequence of Leptotrichia buccalis type strain (C-1013-b).
<3>Standards in Genomic Sciences
<4>1
<5>126-132
<6>2009
<7>Leptotrichia buccalis (Robin 1853) Trevisan 1879 is the type species of the genus, and is of
phylogenetic interest because of its isolated location in the
sparsely populated and neither taxonomically nor genomically adequately accessed
family 'Leptotrichiaceae' within the phylum 'Fusobacteria'. Species of
Leptotrichia are large, fusiform, non-motile, non-sporulating rods, which often
populate the human oral flora. L. buccalis is anaerobic to aerotolerant, and
saccharolytic. Here we describe the features of this organism, together with the
complete genome sequence and annotation. This is the first complete genome
sequence of the order 'Fusobacteriales' and no more than the second sequence from
the phylum 'Fusobacteria'. The 2,465,610 bp long single replicon genome with its
2306 protein-coding and 61 RNA genes is a part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Ivanova, N. et al.
<2>Complete genome sequence of Sanguibacter keddieii type strain (ST-74).
<3>Standards in Genomic Sciences
<4>1
<5>110-118
<6>2009
<7>Sanguibacter keddieii is the type species of the genus Sanguibacter, the only genus within the
family of Sanguibacteraceae. Phylogenetically, this family is
located in the neighborhood of the genus Oerskovia and the family
Cellulomonadaceae within the actinobacterial suborder Micrococcineae. The strain
described in this report was isolated from blood of apparently healthy cows. Here
we describe the features of this organism, together with the complete genome
sequence, and annotation. This is the first complete genome sequence of a member
of the family Sanguibacteraceae, and the 4,253,413 bp long single replicon genome
with its 3735 protein-coding and 70 RNA genes is part of the Genomic Encyclopedia
of Bacteria and Archaea project.

<>

<1>Ivarie, R.
<2>Thymine methyls and DNA-protein interactions.
<3>Nucleic Acids Res.
<4>15
<5>9975-9983
<6>1987
<7>Evidence is summarized showing that thymine methyls are as important in the
recognition of specific sequences by proteins as are the more widely recognized
hydrogen bonding sites of bases in the major groove.  Strongest evidence has
come from experiments using functional group mutagenesis in which thymines in a
specific recognition sequence (e.g., promoters, operators and restriction
sites) are replaced by oligonucleotide synthesis with methyl-free uracil or
cytosine and 5-methylcytosine.  Such experiments have shown that thymine
methyls can provide contact points via van der Waals interactions with amino
acid side chains of specific DNA binding proteins.  Actual contact between a
thymine methyl and carbons of a glutamine side chain has been observed in a
cocrystal of the phage 434 repressor and its operator by X-ray analysis.  The
issue of why thymine occurs in DNA is discussed in light of these findings.

<>

<1>Iversen, P.L.
<2>Antisense Antibacterial Cell Division Composition and Method.
<3>International Patent Office
<4>WO 0149775 A
<5>
<6>2001
<7>Antisense oligomers directed to bacterial cell division and cell cycle-encoding nucleic acids
are capable of selectively modulating the biological activity thereof, and are useful in
treatment and prevention of bacterial infection.  The antisense oligomers are substantially
uncharged, and contain from 8 to 40 nucleotide subunits, including a targeting nucleic acid
sequence at leeast 10 nucleotides in length which is effective to hybridize to (i) a bacterial
tRNA or (ii) a target sequence, containing a translational start codon, within a bacterial
nucleic acid which encodes a protein associated with cell division or the cell cycle.  Such
proteins include zipA, sulA, secA, dicA, dicB, dicC, dicF, ftsZ, murC, murD, murE, murF, murG,
minC, minD, minE, mraY, mraW, mraZ, seqA, ddlB, carbamate kinase, D-ala D-ala ligase,
topoisomerase, alkyl hydroperoxide reductase, thioredoxin reductase, dihydrofolate reductase,
and cell wall enzyme.

<>

<1>Iversen, P.L.
<2>Antisense Antibacterial Cell Division Composition and Method.
<3>Japanese Patent Office
<4>JP 2003518946 A
<5>
<6>2001
<7>Antisense oligomers directed to bacterial cell division and cell cycle-encoding nucleic acids
are capable of selectively modulating the biological activity thereof, and are useful in
treatment and prevention of bacterial infection.  The antisense oligomers are substantially
uncharged, and contain from 8 to 40 nucleotide subunits, including a targeting nucleic acid
sequence at leeast 10 nucleotides in length which is effective to hybridize to (i) a bacterial
tRNA or (ii) a target sequence, containing a translational start codon, within a bacterial
nucleic acid which encodes a protein associated with cell division or the cell cycle.  Such
proteins include zipA, sulA, secA, dicA, dicB, dicC, dicF, ftsZ, murC, murD, murE, murF, murG,
minC, minD, minE, mraY, mraW, mraZ, seqA, ddlB, carbamate kinase, D-ala D-ala ligase,
topoisomerase, alkyl hydroperoxide reductase, thioredoxin reductase, dihydrofolate reductase,
and cell wall enzyme.

<>

<1>Iversen, P.L.
<2>Antisense Antibacterial Cell Division Composition and Method.
<3>Korean Patent Office
<4>KR 1020027008681 A
<5>
<6>2002
<7>
<>

<1>Iversen, P.L.
<2>Antisense antibacterial cell division composition and method.
<3>US Patent Office
<4>US 7049431 B
<5>
<6>2006
<7>Antisense oligomers directed to bacterial cell division and cell cycle-encoding nucleic acids
are capable of selectively modulating the biological activity thereof, and are useful in
treatment and prevention of bacterial infection.  The antisense oligomers are substantially
uncharged, and contain from 8 to 40 nucleotide subunits, including a targeting nucleic acid
sequence at least 10 nucleotides in length which is effective to hybridize to (i) a bacterial
tRNA or (ii) a target sequence, containing a translational start codon, within a bacterial
nucleic acid which encodes a protein associated with cell division or the cell cycle.  Such
proteins include zipA, sulA, secA, dicA, dicB, dicC, dicF, ftsA, ftsI, ftsN, ftsK, ftsL, ftsQ,
ftsW, ftsZ, murC, murD, murE, murF, murG, minC, mind, mine, mraY, mraW, mraZ, seqA, ddlB,
carbamate kinase, D-ala D-ala ligase, topoisomerase, alkyl hydroperoxide reductase,
thioredoxin reductase, dihydrofolate reductase, and cell wall enzyme.

<>

<1>Ives, C.L., Nathan, P.D., Brooks, J.E.
<2>Regulation of the BamHI restriction-modification system by a small intergenic open reading frame, bamHIC, in both Escherichia coli and Bacillus subtilis.
<3>J. Bacteriol.
<4>174
<5>7194-7201
<6>1992
<7>BamHI from Bacillus amyloliquefaciens H, is a type II restriction-modification system
recognizing and cleaving the sequence G^GATCC. The BamHI restriction-modification system
contains divergently transcribed endonuclease and methylase genes along with a small open
reading frame oriented in the direction of the endonuclease gene. The small open reading frame
has been designated bamHIC (for BamHI controlling element). It acts as both a positive
activator of endonuclease expression and a negative repressor of methylase expression of BamHI
clones in Escherichia coli. Methylase activity increased 15-fold and endonuclease activity
decreased 100-fold when bamHIC was inactivated. The normal levels of activity for both
methylase and endonuclease were restored by supplying bamHIC in trans. The BamHI
restriction-modification system was transferred into Bacillus subtilis, where bamHIC also
regulated endonuclease expression when present on multicopy plasmid vectors or integrated into
the chromosome. In B. subtilis, disruption of bamHIC caused at least a 1,000-fold decrease in
endonuclease activity; activity was partially restored by supplying bamHIC in trans.

<>

<1>Ives, C.L., Sohail, A., Brooks, J.E.
<2>The regulatory C proteins from different restriction-modification systems can cross-complement.
<3>J. Bacteriol.
<4>177
<5>6313-6315
<6>1995
<7>The BamHI restriction-modification system contains a third gene, bamHIC, which positively
regulates bamHIR.  Similar small genes from other systems were tested in vivo for their
ability to cross-complement.  C.BamHI protein was identified, purified, and used to raise
polyclonal antibodies.  Attempts to detect other C proteins in cell extracts by
cross-reactivity with C.BamHI antibodies proved unsuccessful.

<>

<1>Ivic, A., Jakeman, K.J., Penn, C.W., Brown, N.L.
<2>Type II restriction endonucleases from Helicobacter pylori include an enzyme with a novel recognition sequence.
<3>FEMS Microbiol. Lett.
<4>179
<5>175-180
<6>1999
<7>Type II restriction endonuclease activities of Helicobacter pylori strain Roberts and of the
type strain H. pylori NCTC 11637 were detected and analysed by conventional techniques. The
endonucleases were partially purified, their optima for activity and their recognition and
cleavage sites were determined. H. pylori (Roberts) contained at least two enzymes: HpyBI was
an isoschizomer of RsaI (GT/AC) and HpyBII was of a novel specificity (GTN/NAC). H. pylori
NCTC 11637 was found to contain an isoschizomer of EcoRV (HpyCI: GAT/ATC) and at least one
other enzyme which was too unstable to characterise.

<>

<1>Ivshina, I.B., Kuyukina, M.S., Krivoruchko, A.V., Barbe, V., Fischer, C.
<2>Draft Genome Sequence of Propane- and Butane-Oxidizing Actinobacterium Rhodococcus ruber IEGM 231.
<3>Genome Announcements
<4>2
<5>e01297-14
<6>2014
<7>We report a draft genome sequence of Rhodococcus ruber IEGM 231, isolated from a  water spring
near an oil-extracting enterprise (Perm region, Russian Federation).
This sequence provides important insights into the genetic mechanisms of propane
and n-butane metabolism, organic sulfide and beta-sitosterol biotransformation,
glycolipid biosurfactant production, and heavy metal resistance in
actinobacteria.

<>

<1>Iwabuchi, M., Tajima, S., Inoue, T., Shibata, T., Ando, T.
<2>A restriction enzyme, BpuI is an isoschizomer of BanII.
<3>Nucleic Acids Res.
<4>20
<5>5850
<6>1992
<7>We previously identified a type II restriction enzyme in extracts of Bacillus pumilus AHU1387
and designated it BpuI (1,2), which recognizes the sequence GRGCYC and therefore may be an
isoschizomer of HgiJII or BanII (3). In the present study, we examined the cleavage site of
BpuI as well as the effect of methylation on the cleavage reaction in order to show that BpuI
is an isoschizomer of BanII.

<>

<1>Iwai, M., Katoh, H., Katayama, M., Ikeuchi, M.
<2>Improved genetic transformation of the thermophilic cyanobacterium, Thermosynechococcus elongatus BP-1.
<3>Plant Cell Physiol.
<4>45
<5>171-175
<6>2004
<7>We improved genetic transformation of the thermophilic cyanobacterium, Thermosynechococcus
elongatus BP-1, by combining electroporation with a
top agar method. Transformation was also improved when a disruptant of a
putative type I restriction endonuclease (tll2230) was used as recipient
cells. In particular, some constructs, with which wild type has never been
transformed, were successfully integrated into the tll2230-disruptant.
Single-crossover recombination was detected more frequently than the
double-crossover recombination. In accordance with the presence of all the
homologs of pil genes in Synechocystis sp. PCC 6803, we found that T.
elongatus is naturally transformable with exogenous DNA.

<>

<1>Iwamoto, M., Hishiki, A., Shimada, T., Imasaki, T., Tsuda, J., Kita, K., Shimizu, T., Sato, M., Hashimoto, H.
<2>Crystallization and X-ray diffraction studies of DNA-free and DNA-bound forms of EcoO109I DNA methyltransferase.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>66
<5>1528-1530
<6>2010
<7>EcoO109I DNA methyltransferase (M.EcoO109I) is a type II modification enzyme from the EcoO109I
restriction-modification system identified in
Escherichia coli strain H709c. M.EcoO109I recognizes double-stranded
RGGNCCY (where R = A or G, Y = T or C and N is any base) and transfers
a methyl group to the C5 of the inner cytosines from
S-adenosylmethionine. To reveal the mechanism of substrate recognition
by M.EcoO109I, DNA-free and DNA-bound forms of M.EcoO109I were
successfully crystallized. Crystals of the DNA-free and DNA-bound forms
belonged to space groups P4(2)2(1)2, with unit-cell parameters a = b =
120.5, c = 79.8 A, and P2(1), with unit-cell parameters a = 55.8, b =
77.4, c = 117.4 A, beta = 93.5 degrees, respectively.

<>

<1>Iwaniec, L.M., Kroll, J.J., Roethel, W.M., Maybaum, J.
<2>Selective inhibition of sequence-specific protein-DNA interactions by incorporation of 6-thioguanine:  Cleavage by restriction endonucleases.
<3>Mol. Pharmacol.
<4>39
<5>299-306
<6>1991
<7>Incorporation of the antileukemic agent 6-thioguanine (TG) into cellular DNA
has been demonstrated to be a major determinant of its cytotoxicity.  We have
previously shown that complete replacement of G by TG within one DNA strand of
the SV40 origin of replication can completely inhibit sequence-specific binding
of the viral replication protein T antigen.  The aim of the present study was
to determine the effect of more selective TG substitutions on DNA-protein
interactions, by utilizing the simpler base recognition sequence motifs of
restriction endonucleases.  In the first part of our study, we replaced G with
TG in one or two of four possible sites within the duplex hexameric recognition
sequence of BamHI (5'-G^GATCC-3'), by enzymatic extension of primed
oligonucleotides.  This extension was stalled, but not completely inhibited, at
locations where insertion of consecutive TG moieties was required.  Both
strands of molecules containing a single substitution were cleaved by BamHI at
reduced rates, with the substituted strand inhibited to a greater degree.  In
molecules containing two substitutions, neither strand was cut by BamHI.  In
contrast, we found that scission of these same mono- and disubstituted
substrates by the less stringent isoschizomer MboI (5'-N^GATCN-3') was
inhibited only slightly.  In the second part of our study, we investigated the
effect of analog subsitution on scission by the type II-S enzymes AlwI and
FokI, in order to separately determine the effects of restriction site
modification versus scission site modification.  We found that the reactivity
of these enzymes was completely abolished by TG substitution within the
recognition site, whereas substitution at the scission site had no effect.  Our
results demonstrate that infrequent TG substitutions within symmetric DNA
sequences can inhibit sequence-specific interactions in an asymmetric fashion.
In addition, although previous reports have shown that TG forms a relatively
weak base pair with cytosine, it appears that the inhibition of restriction
endonuclease-mediated cleavage resulting from TG incorporation is a function of
the sequence requirements of the protein and not a general consequence of
distrupted base-pairing at the recognition locus.  These data support the idea
that the cytotoxic consequences of TG incorporation may be due to inhibition of
sequence-specific protein-DNA interactions.

<>

<1>Iyer, L.M., Abhiman, S., Aravind, L.
<2>MutL homologs in restriction-modification systems and the origin of eukaryotic MORC ATPases.
<3>Biol. Direct
<4>3
<5>8
<6>2008
<7>The provenance and biochemical roles of eukaryotic MORC proteins have remained poorly
understood since the discovery of their prototype
MORCI, which is required for meiotic nuclear division in animals. The
MORC family contains a combination of a gyrase, histidine kinase, and
MutL (GHKL) and S5 domains that together constitute a catalytically
active ATPase module. We identify the prokaryotic MORCs and establish
that the MORC family belongs to a larger radiation of several families
of GHKL proteins (paraMORCs) in prokaryotes. Using contextual
information from conserved gene neighborhoods we show that these
proteins primarily function in restriction-modification systems, in
conjunction with diverse superfamily II DNA helicases and
endonucleases. The common ancestor of these GHKL proteins, MutL and
topoisomerase ATPase modules appears to have catalyzed structural
reorganization of protein complexes and concomitant DNA-superstructure
manipulations along with fused or standalone nuclease domains.
Furthermore, contextual associations of the prokaryotic MORCs and their
relatives suggest that their eukaryotic counterparts are likely to
carry out chromatin remodeling by DNA superstructure manipulation in
response to epigenetic signals such as histone and DNA methylation.
Reviewers: This article was reviewed by Arcady Mushegian and Gaspar
Jekely.

<>

<1>Iyer, L.M., Abhiman, S., Aravind, L.
<2>Natural History of Eukaryotic DNA Methylation Systems.
<3>Prog. Mol. Biol. Transl. Sci.
<4>101
<5>25-104
<6>2011
<7>Methylation of cytosines and adenines in DNA is a widespread epigenetic mark in both
prokaryotes and eukaryotes. In eukaryotes, it has a profound influence on chromatin structure
and dynamics. Recent advances in genomics and biochemistry have considerably elucidated the
functions and provenance of these DNA modifications. DNA methylases appear to have emerged
first in bacterial restriction-modification (R-M) systems from ancient RNA-modifying enzymes,
in transitions that involved acquisition of novel catalytic residues and DNA-recognition
features. DNA adenine methylases appear to have been acquired by ciliates, heterolobosean
amoeboflagellates, and certain chlorophyte algae. Six distinct clades of cytosine methylases,
including the DNMT1, DNMT2, and DNMT3 clades, were acquired by eukaryotes through independent
lateral transfer of their precursors from bacteria or bacteriophages. In addition to these,
multiple adenine and cytosine methylases were acquired by several families of eukaryotic
transposons. In eukaryotes, the DNA-methylase module was often combined with distinct modified
and unmodified peptide recognition domains and other modules mediating specialized
interactions, for example, the RFD module of DNMT1 which contains a permuted Sm domain linked
to a helix-turn-helix domain. In eukaryotes, the evolution of DNA methylases appears to have
proceeded in parallel to the elaboration of histone-modifying enzymes and the RNAi system,
with functions related to counter-viral and counter-transposon defense, and regulation of DNA
repair and differential gene expression being their primary ancestral functions. Diverse DNA
demethylation systems that utilize base-excision repair via DNA glycosylases and cytosine
deaminases appear to have emerged in multiple eukaryotic lineages. Comparative genomics
suggests that the link between cytosine methylation and DNA glycosylases probably emerged
first in a novel R M system in bacteria. Recent studies suggest that the 5mC is not a terminal
DNA modification, with enzymes of the Tet/JBP family of 2-oxoglutarate- and iron-dependent
dioxygenases further hydroxylating it to form 5-hydroxy-methylcytosine (5hmC). These enzymes
emerged first in bacteriophages and appear to have been transferred to eukaryotes on one or
more occasions. Eukaryotes appear to have recruited three major types of DNA-binding domains
(SRA/SAD, TAM/MBD, and CXXC) in discriminating DNA with methylated or unmethylated cytosines.
Analysis of the domain architectures of these domains and the DNA methylases suggests that
early in eukaryotic evolution they developed a close functional link with SET-domain
methylases and Jumonji-related demethylases that operate on peptides in chromatin proteins. In
several eukaryotes, other functional connections were elaborated in the form of various
combinations between domains related to DNA methylation and those involved in ATP-dependent
chromatin remodeling and RNAi. In certain eukaryotes, such as mammals and angiosperms, novel
dependencies on the DNA methylation system emerged, which resulted in it affecting unexpected
aspects of the biology of these organisms such as parent offspring interactions. In genomic
terms, this was reflected in the emergence of new proteins related to methylation, such as
Stella. The well-developed methylation systems of certain heteroloboseans, stramenopiles,
chlorophytes, and haptophyte indicate that these might be new model systems to explore the
relevance of DNA modifications in eukaryotes.

<>

<1>Iyer, L.M., Tahiliani, M., Rao, A., Aravind, L.
<2>Prediction of novel families of enzymes involved in oxidative and other complex modifications of bases in nucleic acids.
<3>Cell Cycle
<4>8
<5>1698-1710
<6>2009
<7>Modified bases in nucleic acids present a layer of information that directs biological
function over and beyond the coding capacity of the
conventional bases. While a large number of modified bases have been
identified, many of the enzymes generating them still remain to be
discovered. Recently, members of the 2-oxoglutarate- and
iron(II)-dependent dioxygenase super-family, which modify diverse
substrates from small molecules to biopolymers, were predicted and
subsequently confirmed to catalyze oxidative modification of bases in
nucleic acids. Of these, two distinct families, namely the AlkB and the
kinetoplastid base J binding proteins (JBP) catalyze in situ hydroxylation
of bases in nucleic acids. Using sensitive computational analysis of
sequences, structures and contextual information from genomic structure
and protein domain architectures, we report five distinct families of
2-oxoglutarate- and iron(II)-dependent dioxygenase that we predict to be
involved in nucleic acid modifications. Among the DNA-modifying families,
we show that the dioxygenase domains of the kinetoplastid base J-binding
proteins belong to a larger family that includes the Tet proteins,
prototyped by the human oncogene Tet1, and proteins from basidiomycete
fungi, chlorophyte algae, heterolobosean amoeboflagellates and
bacteriophages. We present evidence that some of these proteins are likely
to be involved in oxidative modification of the 5-methyl group of cytosine
leading to the formation of 5-hydroxymethylcytosine. The Tet/JBP homologs
from basidiomycete fungi such as Laccaria and Coprinopsis show large
lineage-specific expansions and a tight linkage with genes encoding a
novel and distinct family of predicted transposases, and a member of the
Maelstrom-like HMG family. We propose that these fungal members are part
of a mobile transposon. To the best of our knowledge, this is the first
report of a eukaryotic transposable element that encodes its own
DNA-modification enzyme with a potential regulatory role. Through a wider
analysis of other poorly characterized DNA-modifying enzymes we also show
that the phage Mu Mom-like proteins, which catalyze the
N6-carbamoylmethylation of adenines, are also linked to diverse families
of bacterial transposases, suggesting that DNA modification by
transposable elements might have a more general presence than previously
appreciated. Among the other families of 2-oxoglutarate- and
iron(II)-dependent dioxygenases identified in this study, one which is
found in algae, is predicted to mainly comprise of RNA-modifying enzymes
and shows a striking diversity in protein domain architectures suggesting
the presence of RNA modifications with possibly unique adaptive roles. The
results presented here are likely to provide the means for future
investigation of unexpected epigenetic modifications, such as
hydroxymethyl cytosine, that could profoundly impact our understanding of
gene regulation and processes such as DNA demethylation.

<>

<1>Iyer, L.M., Zhang, D., Maxwell, B.A., Aravind, L.
<2>Computational identification of novel biochemical systems involved in oxidation,  glycosylation and other complex modifications of bases in DNA.
<3>Nucleic Acids Res.
<4>41
<5>7635-7655
<6>2013
<7>Discovery of the TET/JBP family of dioxygenases that modify bases in DNA has sparked
considerable interest in novel DNA base modifications and their
biological roles. Using sensitive sequence and structure analyses combined with
contextual information from comparative genomics, we computationally characterize
over 12 novel biochemical systems for DNA modifications. We predict previously
unidentified enzymes, such as the kinetoplastid J-base generating
glycosyltransferase (and its homolog GREB1), the catalytic specificity of
bacteriophage TET/JBP proteins and their role in complex DNA base modifications.
We also predict the enzymes involved in synthesis of hypermodified bases such as
alpha-glutamylthymine and alpha-putrescinylthymine that have remained enigmatic
for several decades. Moreover, the current analysis suggests that bacteriophages
and certain nucleo-cytoplasmic large DNA viruses contain an unexpectedly diverse
range of DNA modification systems, in addition to those using previously
characterized enzymes such as Dam, Dcm, TET/JBP, pyrimidine hydroxymethylases,
Mom and glycosyltransferases. These include enzymes generating modified bases
such as deazaguanines related to queuine and archaeosine, pyrimidines comparable
with lysidine, those derived using modified S-adenosyl methionine derivatives and
those using TET/JBP-generated hydroxymethyl pyrimidines as biosynthetic starting
points. We present evidence that some of these modification systems are also
widely dispersed across prokaryotes and certain eukaryotes such as
basidiomycetes, chlorophyte and stramenopile alga, where they could serve as
novel epigenetic marks for regulation or discrimination of self from non-self
DNA. Our study extends the role of the PUA-like fold domains in recognition of
modified nucleic acids and predicts versions of the ASCH and EVE domains to be
novel 'readers' of modified bases in DNA. These results open opportunities for
the investigation of the biology of these systems and their use in biotechnology.

<>

<1>Iyer, P.R., Geib, S.M., Catchmark, J., Kao, T.H., Tien, M.
<2>Genome Sequence of a Cellulose Producing Bacterium, Gluconacetobacter hansenii ATCC 23769.
<3>J. Bacteriol.
<4>192
<5>4256-4257
<6>2010
<7>The Gram-negative bacterium Gluconacetobacter hansenii is considered a model organism for
studying cellulose synthesis. We have determined the
genome sequence of the strain ATCC 23769.

<>

<1>Izallalen, M., Stoddard, S.
<2>Use of unmethylated DNA and selectable markers to force integration of transforming DNA in Clostridia.
<3>International Patent Office
<4>WO 2012112427 A
<5>
<6>2012
<7>The invention provides methods of preparing bacteria of interest with a stably integrated
nucleic acid sequence of interest.

<>

<1>Izquierdo, J.A. et al.
<2>Complete Genome Sequence of Clostridium clariflavum DSM 19732.
<3>Standards in Genomic Sciences
<4>6
<5>104-115
<6>2012
<7>Clostridium clariflavum is a Cluster III Clostridium within the family Clostridiaceae isolated
from thermophilic anaerobic sludge (Shiratori et al,
2009). This species is of interest because of its similarity to the model
cellulolytic organism Clostridium thermocellum and for the ability of
environmental isolates to break down cellulose and hemicellulose. Here we
describe features of the 4,897,678 bp long genome and its annotation, consisting
of 4,131 protein-coding and 98 RNA genes, for the type strain DSM 19732.

<>

<1>Izsvak, Z., Duda, E.
<2>Star activity and complete loss of specificity of CeqI endonuclease.
<3>Biochem. J.
<4>258
<5>301-303
<6>1989
<7>Restriction endonuclease CeqI, an isoschizomer of EcoRV, exhibits star activity, a relaxation
of specificity in the presence of Mn2+, dimethyl sulphoxide or glycerol. The enzyme cleaves a
set of sequences that differ from the canonical GATATC by only one nucleotide in positions
2,3,4 or 5. Two of these sequences are not cleaved if modified by dam methylase. A further
loss of specificity can be observed in circumstances less favourable for the enzyme, namely
low-ionic-strength buffers of pH values below 6.0 or above 9.4. This activity seems to cleave
DNA at any sequence, producing a smear of well-defined bands. Partial renaturation of the
denatured enzyme gives rise to a similar non-specific nuclease activity.

<>

<1>Izsvak, Z., Jobbagy, Z., Duda, E.
<2>Purification and characterization of CeqI restriction endonuclease.
<3>Z. Naturforsch. C
<4>47
<5>830-834
<6>1992
<7>CeqI, a type II restriction endonuclease, an isoschizomer of EcoRV was purified to apparent
homogeneity by a combination of salt precipitation, ion exchange, dye affinity and hydrophobic
interaction chromatographies. The crude enzyme was present in the form of large aggregates
that could be pelleted by high speed centrifugation. The enzyme was not associated with
cellular membranes, though non-ionic detergents lowered the apparent size of the aggregates.
The purified enzymes also showed a tendency to form large molecular mass (66-600 kDa)
complexes under physiological conditions, in the absence of cleavable DNA. The enzyme formed
smaller complexes in the presence of DNA and non-ionic detergents and dissociated into
subunits (and undergoes reversible loss of activity) in the presence of high concentrations of
salts. According to SDS gel electrophoresis and sedimentation analysis the molecular mass of
the monomer is 32+-2 kDa. The enzyme has a rather broad pH optimum, extending into the
alkaline range and lost specifcity and activity in buffers below pH6.

<>

<1>Izsvak, Z., Jobbagy, Z., Takacs, I., Duda, E.
<2>Cloning and characterization of the genes of the CeqI restriction-modification system.
<3>Int. J. Biochem. Cell Biol.
<4>29
<5>895-900
<6>1997
<7>Two genes from Corynebacterium equii, a Gram-positive producing the CeqI
restriction-modification enzymes were cloned and sequenced.  In vivo restriction experiments,
DNA and amino acid sequence data suggest that the two genes code for the endonuclease and the
methyltransferase enzymes.  However, when the two genes are expressed in E. coli, practically
no enzyme activity can be detected in the supernatants of sonicated cells.  Based on the DNA
sequence data CeqI restriction enodnuclease (an EcoRV isoschizomer) consists of 270 amino acid
residues with a predicted molecular mass of 31.6 kDa, in good agreement with the previously
measured 32+/- 2kDa.  The methyltansferase is 517 residues long (approx. 60 kDa).  The two
genes are in opposite orientation and overlap by 37 base pairs on the chromosome.  The deduced
amino acid sequence of the putative endonuclease gene revealed long stretches of hydrophobic
amino acids, that may form the structural basis of the unusual aggregation properties of the
restriction endonuclease.  The amino acid sequence of the methylase shows homologies with
other type II methyltransferases.

<>

<1>Izvolsky, K.I., Demidov, V.V., Nielsen, P.E., Frank-Kamenetskii, M.D.
<2>Sequence-specific protection of duplex DNA against restriction and methylation enzymes by pseudocomplementary PNAs.
<3>Biochemistry
<4>39
<5>10908-10913
<6>2000
<7>A new generation of PNAs, so-called pseudocomplementary PNAs (pcPNAs) which are able to target
the designated sites on duplex DNA with mixed
sequence of purines and pyrimidines via double-duplex invasion mode,
has recently been introduced. It has been demonstrated that appropriate
pairs of decameric pcPNAs block an access of RNA polymerase to the
corresponding promoter. Here, we show that this type of PNAs protects
selected DNA sites containing all four nucleobases from the action of
restriction enzymes and DNA methyltransferases. We have found that
pcPNAs as short as octamers form stable and sequence-specific complexes
with duplex DNA in a very salt-dependent manner. In accord with a
strand-invasion mode of complex formation, the pcPNA binding proceeds
much faster with supercoiled than with linear plasmids. The
double-duplex invasion complexes selectively shield specific DNA sites
from BclI restriction endonuclease and dam methylase. The
pcPNA-assisted protection against enzymatic methylation is more
efficient when the PNA-binding site embodies the methylase-recognition
site rather than overlaps it. We conclude that pcPNAs may provide the
robust tools allowing to sequence-specifically manipulate DNA duplexes
in a virtually sequence-unrestricted manner.

<>

<1>Jabbar, M.A., Snyder, L.
<2>Genetic and physiological studies of an Escherichia coli locus that restricts polynucleotide kinase- and RNA ligase-deficient mutants of bacteriophage T4.
<3>J. Virol.
<4>51
<5>522-529
<6>1984
<7>The RNA ligase and polynucleotide kinase of bacteriophage T4 are nonessential
enzymes in most laboratory Escherichia coli strains.  However, T4 mutants which
do not induce the enzymes are severely restricted in E. coli CTr5X, a strain
derived from a clinical E. coli isolate.  We have mapped the restricting locus
in E. coli CTr5X and have transduced it into other E. coli strains.  The
restrictive locus seems to be a gene, or genes, unique to CTr5X or to be an
altered form of a nonessential gene, since deleting the locus seems to cause
loss of the phenotypes.  In addition to restricting RNA ligase- and
polynucleotide kinase-deficient T4, the locus also restricts bacteriophages
lambda and T4 with cytosine DNA.  When lambda or T4 with cytosine DNA infect
strains with the prr locus, the phage DNA is injected, but phage genes are not
expressed and the host cells survive.  These phenotypes are unlike anything yet
described for a phage-host interaction.

<>

<1>Jack, W.E.
<2>Participation of outside DNA sequences in the EcoRI endonuclease reaction pathway.
<3>Ph.D. Thesis
<4>
<5>
<6>1983
<7>I have examined the kinetics of the interaction between EcoRI endonuclease and DNA as a model
system for the study of recognition site location by sequence-specific DNA binding proteins.
When the kinetics of interaction of nine linear DNA fragments derived from pBR322 and
containing the EcoRI site in a central position were examined, the kinetic parameters
governing both formation and decay of specific endonuclease-DNA complexes was found to
increase 8-fold with increasing DNA chain length.  In contrast, equilibrium competition
experiments demonstrated the intrinsic affinity for the recognition site was independent of
DNA chain length for these molecules.  Thus, sequences outside the EcoRI site enhance the rate
at which EcoRI endonuclease locates and leaves its recognition site without altering the
intrinsic affinity of the enzyme for that site.  These results are in accord with a
facilitated diffusion mechanism for site location, and indicate outside DNA sequences lie on
the major kinetic pathway by which EcoRI endonuclease locates and leaves its recognition site.
Competition cleavage experiments demonstrated that longer DNA molecules are preferentially
cleaved by the endonuclease, suggesting outside DNA sequences also facilitate catalysis by the
enzyme.  In accord with the view that DNHA sequences outside the EcoRI site are involved in
the reaction pathway, the enzyme processively cleaves EcoRI sites separated by short distances
on the same DNA chain.  The rate limiting step in EcoRI endonuclease catalysis has been
examined by pre-steady state measurements.  Addition of endonuclease to saturating DNA
resulted in a rapid initial burst of product formation.  This burst was stoichiometric with
the amount of enzyme added and proves that steps up to and including double strand cleavage
cannot be rate limiting in catalysis.  A similar burst was observed when EcoRI endonuclease
was incubated with DNA in the absence of Mg2+, and cleavage  initiated by Mg2+ addition.  This
latter burst quantitatively reflects the amount of site-specific enzyme-DNA complexes present
at the time of Mg2+ addition, and has been utilized to assay site-specific binding.

<>

<1>Jack, W.E., Greenough, L., Dorner, L.F., Xu, S.-Y., Strzelecka, T., Aggarwal, A.K., Schildkraut, I.
<2>Overexpression, purification and crystallization of BamHI endonuclease.
<3>Nucleic Acids Res.
<4>19
<5>1825-1829
<6>1991
<7>The type II restriction endonuclease BamHI has been expressed in E. coli,
producing 100-fold more enzyme than the wild type Bacillus amyloliquefaciens H
strain.  This high yield has facilitated purification to homogeneity of large
amounts of the enzyme, along with its crystallization in a form which diffracts
to at least 1.9 angstrom in X-ray analysis.

<>

<1>Jack, W.E., Rubin, R.A., Newman, A., Modrich, P.
<2>Structures and mechanisms of EcoRI DNA restriction and modification enzymes.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>165-179
<6>1981
<7>*
  I. Structural properties
 II. Catalytic properties
III. Enzyme-substrate interactions
 IV. Cloning and the structural genes


<>

<1>Jack, W.E., Terry, B.J., Modrich, P.
<2>Involvement of outside DNA sequences in the major kinetic path by which EcoRI endonuclease locates and leaves its recognition sequence.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>79
<5>4010-4014
<6>1982
<7>We have examined the kinetics of the interaction between endodeoxyribonuclease
EcoRI (EC 3.1.23.13) and nine linear DNA fragments that range in size between
34 and 6,200 base pairs and contain the EcoRI site of plasmid pBR322 in a
central location.  The kinetic parameters governing both formation and decay of
specific endonuclease DNA complexes increase 8-fold with increasing chain
length over this size range.  In contrast, equilibrium competition experiments
demonstrated that the intrinsic affinity of endonuclease for this recognition
sequence is independent of DNA chain length over this size range.  In contrast,
equilibrium competition experiments demonstrated that the intrinsic affinity of
endonuclease for its recognition sequence is independent of DNA chain length
over this range.  Thus, DNA sequences outside the recognition site enhance the
rate at which EcoRI endonuclease locates or leaves its recognition site without
affecting the intrinsic thermodynamic parameters of site-specific interaction.
These results are consistent with a facilitated diffusion mechanism for
specific DNA site location by this enzyme.

<>

<1>Jackson, D.A., Symons, R.H., Berg, P.
<2>Biochemical method for inserting new genetic information into DNA of simian virus 40:  circular SV40 DNA molecules containing lambda phage genes and the galactose operon of Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>2904-2909
<6>1972
<7>We have developed methods for covalently joining duplex DNA molecules to one
another and have used these techniques to construct circular dimers of SV40 DNA
and to insert a DNA segment containing lambda phage genes and the galactose
operon of E. coli into SV40 DNA.  The method involves:   (a) converting
circular SV40 DNA to a linear form, (b) adding single-stranded
homodeoxypolymeric extensions of defined composition and length to the 3' ends
of one of the DNA strands with the enzyme terminal deoxynucleotidyl transferase
(c) adding complementary homodeoxypolymeric extensions to the other DNA strand,
(d) annealing the two DNA molecules to form a circular duplex structure, and
(e) filling the gaps and sealing nicks in this structure with E. coli DNA
polymerase and DNA ligase to form a covalently closed-circular DNA molecule.

<>

<1>Jackson, S.E., Koyama, M., Ryu, J.
<2>A novel use of bacterial transformation to discover new restriction enzymes in clinical E. coli strains.
<3>J. Invest. Med.
<4>49
<5>11A
<6>2001
<7>Recent bacterial genome projects have revealed many new DNA sequences that encode restriction
endonucleases and their corresponding methylases.  These findings suggest that many
restriction enzymes have yet to be discovered.  Traditionally, restriction enzymes have been
discovered by either the classical restriction and modification phenomena of bacteriophages or
by direct enzyme assay.  To avoid the limitations of these traditional methods, we established
a simple, quantitative R-M test based on plasmid transformation efficiency (plasmid R-M test)
using DNA fragments derived from the E. coli phage Lambda.  This new plasmid R-M test works
similarly to a traditional phage efficiency of plating assay, but is defined as efficiency of
transformation.  A plasmid kit (p1-p6) was created by combining fragments of Lambda with the
vector plasmid pMECA, which confers ampicillin resistance.  We used this new EOT method to
test 250 E. coli strains obtained from 2 local hospitals.  Out of the 250 samples, 58 strains
were ampicillin sensitive and were used for further study.  The plasmid pMECA and plasmids
p1-p6 from the kit were transformed into these 58 strains.  Among those, 15 strains had a high
rate of transformability and a suspected restriction-modification system.  The restriction
results for these 15 strains were tabulated for p1-p6 by assigning a value of either (+) or
(-).  Restriction (+) indicates the presence of a DNA recognition site, and thus no bacterial
growth.  Restriction (-) indicates the absence of a recognition site, and presence of
bacterial growth.  After comparing the restriction patterns of the 15 clinical strains with
the restriction patterns of all known E. coli strains (173 listed in REBASE), we found one new
restriction sequence pattern.  This novel pattern suggests a new DNA recognition sequence.  In
order to identify this recognition sequence, we, in collaboration with UC Riverside engineers,
developed a computer program designed to analyze the +/- restriction patterns.  This series of
experiments indicates that the EOT test can be used to find new restriction enzymes in
clinical E. coli samples.  We believe that this new method can be expanded to find new
restriction enzymes and their recognition sequence in many types of bacteria.

<>

<1>Jacobs, D.
<2>Type II restriction-modification systems in Enterobacter aerogenes and Herpetosiphon giganteus.
<3>Diss. Abstr.
<4>49
<5>730B
<6>1988
<7>The Type II modification methylase M.EaeI corresponding to the restriction
endonuclease EaeI was partially purified from Enterobacter aerogenes PW201.
The methylase converts the innermost cytosine residue in each strand of the
family of related sequences recognised by EaeI (5'-PyGGCCPu-3') to
5-methylcytosine.  M.EaeI protects these sites against cleavage by HaeIII and
also protects overlapping 5'-CCGG-3 sites against cleavage by HpaII and MspI.
Such protection may be useful in genetic manipulations.  Despite using a
variety of cloning strategies, attempts to clone the EaeI
restriction-modification system in Escherichia coli were unsuccessful.  It is
now apparent that many of the problems encountered during these attempts can be
explained by the existence in most E. coli strains of restriction systems which
specifically restrict methylated DNA.  Possible evolutionary relationships
between Type II restriction-modification systems were investigated in
Herpetosiphon giganteus.  HgiJI from H. giganteus HFS101 has the recognition
specificity 5'-GG(A/T)CC-3' and is the fifth isoschizomer of AvaII to have been
identified in this genus.  The recognition specificities of HgiJII
(5'-GPuGCPyC-3') and HgiAI (5'-G(A/T)GC(A/T)C-3') from H. giganteus strains
HFS101 and HP1023 respectively are very similar and the DNAs from these strains
are resistant to cleavage by both restriction endonucleases.  HgiAI and HgiJII
may therefore be closely-related enzymes.  The potential role of Type II
restriction-modification systems in limiting gene transfer between strains in
this genus is also discussed.

<>

<1>Jacobs, D., Brown, N.L.
<2>Isolation and characterization of the M.EaeI modification methylase.
<3>Biochem. J.
<4>238
<5>613-616
<6>1986
<7>Restriction endonucleases have found wide use in the analysis and restructuring of DNA
molecules and as model systems for studying the interactions of proteins and DNA.  Many such
activities have been characterized from a large number of bacterial genera (Roberts, 1985),
and in virtually all cases examined a corresponding methylase is present that protects the
bacterial DNA from cleavage (McClelland & Nelson, 1985).  These modification methylases have
been less well-studied, presumably because they are of less importance as tools in the
manipulation of DNA in vitro.  However, such methylases can be used to protect specific sites
against cleavage by the cognate endonuclease, and they have been used to alter the number of
target sites for non-cognate restriction endonucleases on DNA to generate novel specificities
of cleavage (McClelland et al., 1984).We now report the specificity of the methylase M.EaeI
corresponding to the endonuclease EaeI from Enterobacter aerogenes PW201 (Whitehead & Brown,
1983).  We show that the methylase can be used to protect a subset of HaeIII recognition
sequences from HaeIII cleavage and HpaII/MspI sites from HpaII or MspI cleavage.  Such
protection may be useful in genetic manipulations.

<>

<1>Jacobs, M.B., Norris, S.J., Phillippi-Falkenstein, K.M., Philipp, M.T.
<2>Infectivity of the highly transformable BBE02(-) lp56(-) mutant of Borrelia burgdorferi, the Lyme disease spirochete, via ticks.
<3>Infect. Immun.
<4>74
<5>3678-3681
<6>2006
<7>Infectious Borrelia burgdorferi strains that have increased transformability with the shuttle
vector pBSV2 were recently
constructed by inactivating the gene encoding BBE02, a putative
restriction-modification gene product expressed by the linear plasmid
1p25 (Kawabata et al., Infect. Immun. 72:7147-7154, 2004). The absence
of the linear plasmid 1p56, which carries another putative
restriction-modification gene, further enhanced transformation rates.
The infectivity of these mutants was assessed previously in mice that
were inoculated with needle and syringe and was found to be equivalent
to that of wild-type spirochetes. Here we examined the infectivity of
spirochetes to ticks after capillary inoculation of Ixodes scapularis
nymphs and the subsequent spirochetal infectivity to mice via ticks by
using B. burgdorferi B31 clonal isolates lacking 1p56 and/or BBE02. The
absence of 1p56 (but not BBE02) correlated with a lower number of
spirochetes in ticks after feeding on mice; this plasmid thus may play
a role, albeit not an essential one, in supporting spirochetal survival
in the feeding tick. Importantly, however, the absence of 1p56 and
BBE02 did not detectably influence infectivity to mice via ticks.

<>

<1>Jacoby, G.A., Sutton, L.
<2>Restriction-modification systems determined by Pseudomonas plasmids.
<3>Plasmid
<4>8
<5>141-147
<6>1982
<7>Four additional Pseudomonas R plasmids determining the PaeR7
restriction-modification system have been detected.  All are transfer deficient
and appear to belong to the same incompatibility group.  The Pseudomonas
fertility plasmid FP110 determines a different restriction-modification system
and also inhibits the propagation of phage B39 by a separate mechanism.
Pseudomonas R plasmid pMG73 has a third distinct restriction-modification
specificity.  PaeFP110 and PaeR73 are proposed as designations for these new
plasmid-determined systems for restriction and modification.

<>

<1>Jacoby, G.A., Sutton, L.
<2>Restriction and modification determined by a Pseudomonas R plasmid.
<3>Plasmid
<4>1
<5>115-116
<6>1977
<7>Pseudomonas plasmid pMG7 interferes with the propagation of bacteriophages B3, D3, F116, and
G101 by determining a restriction and modification system.  This system also acts on plasmids
RP-1 and RP8 to limit transfer into a pMG7+ recipient.

<>

<1>Jacoby, K., Metzger, M., Shen, B.W., Certo, M.T., Jarjour, J., Stoddard, B.L., Scharenberg, A.M.
<2>Expanding LAGLIDADG endonuclease scaffold diversity by rapidly surveying evolutionary sequence space.
<3>Nucleic Acids Res.
<4>40
<5>4954-4964
<6>2012
<7>LAGLIDADG homing endonucleases (LHEs) are a family of highly specific DNA endonucleases
capable of recognizing target sequences approximately 20 bp in
length, thus drawing intense interest for their potential academic,
biotechnological and clinical applications. Methods for rational design of LHEs
to cleave desired target sites are presently limited by a small number of
high-quality native LHEs to serve as scaffolds for protein engineering-many are
unsatisfactory for gene targeting applications. One strategy to address such
limitations is to identify close homologs of existing LHEs possessing superior
biophysical or catalytic properties. To test this concept, we searched public
sequence databases to identify putative LHE open reading frames homologous to the
LHE I-AniI and used a DNA binding and cleavage assay using yeast surface display
to rapidly survey a subset of the predicted proteins. These proteins exhibited a
range of capacities for surface expression and also displayed locally altered
binding and cleavage specificities with a range of in vivo cleavage activities.
Of these enzymes, I-HjeMI demonstrated the greatest activity in vivo and was
readily crystallizable, allowing a comparative structural analysis. Taken
together, our results suggest that even highly homologous LHEs offer a readily
accessible resource of related scaffolds that display diverse biochemical
properties for biotechnological applications.

<>

<1>Jacques, M.A., Bolot, S., Charbit, E., Darrasse, A., Briand, M., Arlat, M., Gagnevin, L., Koebnik, R., Noel, L.D., Portier, P., Carrere, S., Boureau, T.
<2>High-Quality Draft Genome Sequence of Xanthomonas alfalfae subsp. alfalfae Strain CFBP 3836.
<3>Genome Announcements
<4>1
<5>e01035-13
<6>2013
<7>We report the high-quality draft genome sequence of Xanthomonas alfalfae subsp. alfalfae
strain CFBP 3836, the causal agent of bacterial leaf and stem spot in
lucerne (Medicago sativa). Comparative genomics will help to decipher the
mechanisms provoking disease and triggering the defense responses of this
pathogen of the model legume Medicago truncatula.

<>

<1>Jacquier, A., Dujon, B.
<2>An intron-encoded protein is active in a gene conversion process that spreads an intron into a mitochondrial gene.
<3>Cell
<4>41
<5>383-394
<6>1985
<7>The intron of the mitochondrial 21S rRNA gene of Saccharomyces cerevisiae possesses a long
internal reading frame (ORF) that is conserved in various yeast species. In crosses between
intron-plus and intron-minus variants, this intron determines a specific gene conversion
phenomenon, which results in the integration of the intron sequence within all previously
intron-minus copies of the gene. We show, from a frameshift mutant within the intron this ORF
and from the need of mitochondrial protein synthesis, that ORF encodes a protein active in the
gene conversion that spreads the intron within populations of interbreeding strains. This new
intron function is reminiscent of the transposase encoded by mobile genetic elements and is
discussed in relation to other intron functions.

<>

<1>Jacquier, A., Dujon, B.
<2>The intron of the mitochondrial 21S rRNA gene:  distribution in different yeast species and sequence comparison between Kluyveromyces thermotolerans and Saccharomyces cerevisiae.
<3>Mol. Gen. Genet.
<4>192
<5>487-499
<6>1983
<7>We have screened numerous different yeast species for the presence of sequences homologous to
the intron of the mitochondrial 21S rRNA gene of Saccharomyces cerevisiae (intron r1) and
found them in all Kluyveromyces species, some of the Saccharomyces species and none of the
other yeasts tested. We have determined the nucleotide sequence of the r1-intron in K.
thermotolerans and compared it with that of S. cerevisiae. The two introns are inserted at the
same position within the 21S rRNA gene. They contain homologous internal open reading frames
(ORFs) initiated at the same AUG codon which can be aligned over their entire length. Several
silent multi-substitutions indicate that these intronic ORFs represent selectively conserved
functional genes. Other intron segments, on the contrary, reveal short blocks of extensive
homology separated by non-homologous stretches and/or additions-deletions. Comparison of our
two yeast r1-introns with equivalent introns of N. crassa and A. nidulans mitochondria reveals
that introns with very similar RNA secondary structures can accommodate different types of
ORFs.

<>

<1>Jadhav, V.R., Ganesh, K.N.
<2>Design of a combinatorial oligonucleotide library containing all possible hexamer palindromes: PCR synthesis and application for identifying restriction cleavage sites.
<3>Biochem. Biophys. Res. Commun.
<4>242
<5>297-302
<6>1998
<7>An algorithm for designing a combinatorial library comprehensively representing all hexamer
palindrome sequences at uniquely defined sites is described.  The expected size for such a
library of 64 possible hexamer palindromes is 384 bases, which is reduced to 266 bases spread
over 8 oligonucleotides through a linear overlap of rationally selected hexamer palindromes.
The single stranded oligonucleotides of the designed sets were chemically synthesized and
converted into corresponding duplex dimers using PCR primer-dimer method.  The utility of
these duplex oligomers for identifying cleavage sites of restriction enzymes recognizing
hexamer palindromes has been demonstrated using some representative enzymes.  The library is
also useful for screening restriction enzymes with tetramer cleavage sites and identifying the
"star" sites of restriction enzymes.  The sets of oligonucleotides with high information
content, though designed for direct and unambiguous characterization of cleavage sites of
isolated restriction enzymes, have potential applications as templates for characterizing
sequence selective binding and interaction of small molecules nucleic acid.

<>

<1>Jaffe, B., Kovacs, K., Andras, C., Bodi, Z., Liu, Z., Fray, R.G.
<2>Methylation of chloroplast DNA does not affect viability and maternal inheritance in tobacco and may provide a strategy towards transgene containment.
<3>Plant Cell
<4>27
<5>1377-1384
<6>2008
<7>We report the integration of a type II restriction-methylase, mFokI, into the tobacco
chloroplast genome and we demonstrate that the
introduced enzyme effectively directs the methylation of its target
sequence in vivo and does not affect maternal inheritance. We further
report the transformation of tobacco with an E. coli dcm methylase
targeted to plastids and we demonstrate efficient cytosine methylation
of the plastid genome. Both adenosine methylation of FokI sites and
cytosine methylation of dcm sites appeared phenotypically neutral. The
ability to tolerate such plastid genome methylation is a pre-requisite
for a proposed plant transgene containment system. In such a system, a
chloroplast located, maternally inherited restriction methylase would
provide protection from a nuclear-encoded, plastid targeted restriction
endonuclease. As plastids are not paternally inherited in most crop
species, pollen from such plants would carry the endonuclease transgene
but not the corresponding methylase; the consequence of this should be
containment of all nuclear transgenes, as pollination will only be
viable in crosses to the appropriate transplastomic maternal
background.

<>

<1>Jager, K., Potts, M.
<2>Distinct fractions of genomic DNA from cyanobacterium Nostoc commune that differ in the degree of methylation.
<3>Gene
<4>74
<5>197-201
<6>1988
<7>Meeting Abstract

<>

<1>Jagoda, S.S., Tan, E., Arulkanthan, A., Kinoshita, S., Watabe, S., Asakawa, S.
<2>Draft Genome Sequence of Aeromonas hydrophila Strain Ae34, Isolated from a Septicemic and Moribund Koi Carp (Cyprinus carpio koi), a Freshwater Aquarium  Fish.
<3>Genome Announcements
<4>2
<5>e00572-14
<6>2014
<7>Aeromonas hydrophila is an important opportunistic pathogen that infects a variety of aquatic
and terrestrial animals, including humans. We report here the
draft genome sequence of A. hydrophila Ae34, a multidrug-resistant isolate from
the kidney of a moribund koi carp (Ciprinus carpio koi) with signs of hemorrhagic
septicemia.

<>

<1>Jair, K., Yen, R.-W.C., Toyota, M., Ho, C., Baylin, S.B., Issa, J.-P.J.
<2>De novo DNA methyltransferase activity in a lung cancer cell line.
<3>Proc. Amer. Assoc. Cancer Res.
<4>39
<5>95
<6>1998
<7>In mammalian cells genomic methylation patterns are important for normal embryonic development
and cell growth.  It has been proposed that distinct DNA methyltransferases are responsible
for de novo and maintenance methylation, however, only one DNA methyltransferase with
predominant maintenance activity has been identified in vertebrates so far.  In addition, the
role of DNA methyltransferases in establishing aberrant patterns of DNA methylation in cancer
remain poorly defined.  We have now tested both de novo and maintenance methylation activities
in different cancer cell lines by using a DNA methyltransferase assay with polydldC
(maintenance) and Drosophila genomic DNA (de novo) as substrates.  One lung cancer cell line,
H249, had an abnormally high de novo methyltransferase activity.  Interestingly, treatment
with 5-azadeoxycytidine reduced maintenance methylation activity in this cell, but did not
affect de novo methylation activity.  Using gel filtration chromatography, we found that the
peak activities of both maintenance and de novo methylation were detected in the same eluted
fraction, suggesting that enzymes with similar molecular size, or the same enzyme, are
responsible for maintenance and de novo methylation activities in this cell line.  Despite
this in vitro de novo methyltransferase activity, several CpG islands which are commonly
methylated in cancer remain methylation-free in this  cell line.  Thus, further investigation
of this cancer cell line H249 may provide us further insights into the mechanisms of
maintenance and de novo methylation, and their relation to aberrant methylation in cancer.

<>

<1>Jakubauskas, A.
<2>Domain organization analysis of Type II restriction endonucleases.
<3>Ph.D. Thesis, Vilnius University
<4>
<5>1-42
<6>2007
<7>CONCLUSIONS
1. Two-domain organization of Type IIS REases R.Eco31I and R.Hin4II and Type IIP REase R.SdaI
was identified by limited proteolysis.
2. The isolated N-domain of R.Eco31I was found to be responsible for the specific interaction
with DNA.
3. The single HNH nuclease-like active site was identified in the C-domain of R.Eco31I.
4. Two R-M systems were cloned in E. coli: Hin4II of novel DNA specificity 5�Œ-CCTTC(6/5) and
SdaI that recognizes and cleave eight base DNA target, 5�Œ-CCTGCA�«GG.
5. The HNH nuclease-like active site was identified in the C-domain of R.Hin4II by
bioinformatic methods.

<>

<1>Jakubauskas, A., Dauksaite, V.
<2>Construction and analysis of chimeric enzymes between type IIs restriction endonucleases Alw26I, Eco31I and Esp3I.
<3>Biologija
<4>1
<5>26-30
<6>1997
<7>Conserved regions I, III and IV were swapped between restriction endonucleases Alw26I, Eco31I
and Esp3I, and 16 chimeric enzymes were constructed.  Only the hybrid enzyme
Eco31I-(N-esp-Mva1269I) containing the N-terminus and conserved region I from Esp3I and the
remaining part from Eco31I revealed the enzymatic activity of Eco31I specificity in crude
lysate.  This indicates that the conserved region I belongs to the structural elements of
Eco31I.  The four hybrid enzymes with swapped intermediate sequences between conserved regions
III and IV and/or conserved region IV induce an SOS response in E. coli ER1992, which can be
blocked with M.Alw26I methylase.  These results suggest that DNA lesions incurred by hybrid
enzymes are site specific.

<>

<1>Jakubauskas, A., Giedriene, J., Bujnicki, J.M., Janulaitis, A.
<2>Identification of a Single HNH Active Site in Type IIS Restriction Endonuclease Eco31I.
<3>J. Mol. Biol.
<4>370
<5>157-169
<6>2007
<7>Type IIS restriction endonuclease Eco31I is a "short-distance cutter", which cleaves DNA
strands close to its recognition sequence, 5'-GGTCTC(1/5). Previously, it has been proposed
that related endonucleases recognizing a common sequence core GTCTC possess two active sites
for cleavage of both strands in the DNA substrate. Here, we present bioinformatic
identification and experimental evidence for a single nuclease active site. We identified a
short region of homology between Eco31I and HNH nucleases, constructed a three-dimensional
model of the putative catalytic domain and validated our predictions by random and
site-specific mutagenesis. The restriction mechanism of Eco31I is suggested by analogy to the
mechanisms of phage T4 endonuclease VII and homing endonuclease I-PpoI. We propose that
residues D311 and N334 coordinate the cofactor. H312 acts as a general base-activating water
molecule for the nucleophilic attack. K337 together with R340 and D345 are located in close
proximity to the active center and are essential for correct folding of catalytic motif, while
D345 together with R264 and D273 could be directly involved in DNA binding. We also predict
that the Eco31I catalytic domain contains a putative Zn-binding site, which is essential for
its structural integrity. Our results suggest that the HNH-like active site is involved in the
cleavage of both strands in the DNA substrate. On the other hand, analysis of site-specific
mutants in the region, previously suggested to harbor the second active site, revealed its
irrelevance to the nuclease activity. Thus, our data argue against the earlier prediction and
indicate the presence of a single conserved active site in type IIS restriction endonucleases
that recognize common sequence core GTCTC.

<>

<1>Jakubauskas, A., Kriukiene, E., Trinkunaite, L., Sapranauskas, R., Jurenaite-Urbanaviciene, S., Lubys, A.
<2>Bioinformatic and partial functional analysis of pEspA and pEspB, two plasmids from Exiguobacterium arabatum sp nov RFL1109.
<3>Plasmid
<4>61
<5>52-64
<6>2009
<7>The complete nucleotide sequences of two plasmids from Exiguobacterium arabatum sp. nov.
RFL1109, pEspA (4563 bp) and pEspB (.38,945 bp), have
been determined. Five ORFs were identified in the pEspA plasmid, and
putative functions were assigned to two of them. Using deletion mapping
approach, the Rep-independent replication region of pEspA, which
functions in Bacillus subtilis, was localized within a 0.6 kb DNA
region. Analysis of the pEspB sequence revealed 42 ORFs. From these,
function of two genes encoding enzymes of the Lsp11091
restriction-modification system was confirmed experimentally, while
putative functions of another 18 ORFs were suggested based on
comparative analysis. Three functional regions have been proposed for
the pEspB plasmid: the putative conjugative transfer region, the region
involved in plasmid replication and maintenance, and the region
responsible for transposition of the IS21 family-like transposable
elements.

<>

<1>Jakubauskas, A., Sasnauskas, G., Giedriene, J., Janulaitis, A.
<2>Domain organization and functional analysis of type IIS restriction endonuclease Eco31I.
<3>Biochemistry
<4>47
<5>8546-8556
<6>2008
<7>Type IIS restriction endonuclease Eco31I harbors a single HNH active site and cleaves both DNA
strands close to its recognition sequence,
5'-GGTCTC(1/5). A two-domain organization of Eco31I was determined by
limited proteolysis. Analysis of proteolytic fragments revealed that
the N-terminal domain of Eco31I is responsible for the specific DNA
binding, while the C-terminal domain contains the HNH nuclease-like
active site. Gel-shift and gel-filtration experiments revealed that a
monomer of the N-terminal domain of Eco31I is able to bind a single
copy of cognate DNA. However, in contrast to other studied type IIS
enzymes, the isolated catalytic domain of Eco31I was inactive.
Steady-state and transient kinetic analysis of Eco31I reactions was
inconsistent with dimerization of Eco31I on DNA. Thus, we propose that
Eco31I interacts with individual copies of its recognition sequence in
its monomeric form and presumably remains a monomer as it cleaves both
strands of double-stranded DNA. The domain organization and reaction
mechanism established for Eco31I should be common for a group of
evolutionary related type IIS restriction endonucleases Alw26I, BsaI,
BsmAI, BsmBI and Esp3I that recognize DNA sequences bearing the common
pentanucleotide 5'-GTCTC.

<>

<1>Jalan, N., Aritua, V., Kumar, D., Yu, F., Jones, J.B., Graham, J.H., Setubal, J.C., Wang, N.
<2>Comparative genomic analysis of Xanthomonas axonopodis pv. citrumelo F1 causing citrus bacterial spot and related strains provides insights into  virulence and host-specificity.
<3>J. Bacteriol.
<4>193
<5>6342-6357
<6>2011
<7>Xanthomonas axonopodis pv. citrumelo (Xacm) is a citrus pathogen causing citrus bacterial spot
disease that is geographically restricted within the
state of Florida. Illumina, 454 sequencing and optical mapping were used
to obtain a complete genome sequence of Xacm strain F1, 4.9Mb in size. The
strain lacks plasmids as compared to other citrus pathogens. Phylogenetic
analysis revealed that this pathogen is very close to the tomato bacterial
spot pathogen Xcv 85-10 with a completely different host range. We also
compared Xacm to the genome of citrus canker pathogen Xac 306. Comparative
genomic analysis showed differences in several gene clusters like Type 3
effectors, Type 4 secretion system, lipopolysaccharide synthesis and
others. In addition to pthA, effectors such as xopE3, xopAI and hrpW were
absent in Xacm while present in Xac. These effectors might be responsible
for survival and reduced virulence of this pathogen on citrus compared to
Xac. We also identified unique effectors in Xacm that may be related to
the different host range as compared to Xac. Xacm also lacks various genes
such as syrE1, syrE2 and RTX toxin family genes, which were present in
Xac. These may be associated with distinct virulence of Xacm and Xac.
Comparison of the complete genome sequence of Xacm to Xac and Xcv provides
valuable insights into the mechanism of bacterial virulence and
host-specificity.

<>

<1>Jalan, N., Kumar, D., Yu, F., Jones, J.B., Graham, J.H., Wang, N.
<2>Complete Genome Sequence of Xanthomonas citri subsp. citri Strain Aw12879, a Restricted-Host-Range Citrus Canker-Causing Bacterium.
<3>Genome Announcements
<4>1
<5>e00235-13
<6>2013
<7>Xanthomonas citri subsp. citri causes citrus canker. The Asiatic strain has a broad host
range, whereas the Wellington variant has a restricted host range.
Here, we present the complete genome of X. citri subsp. citri strain A(W)12879.
This study lays the foundation to further characterize the mechanisms for
virulence and host range of X. citri.

<>

<1>James, B.W., Bacon, J., Marsh, P.
<2>Mycobacterial antigens expressed under low oxygen tension.
<3>International Patent Office
<4>WO 03000721 A
<5>
<6>2003
<7>A method is provided for identifying mycobacterial genes that are induced or up-regulated
under continuous culture conditions defined by a dissolved oxygen tension of up to 10% air
saturation measured at 37oC when compared with a dissolved oxygen tension of at least 40% air
saturation measured at 37oC.  Said induced or up-regulated genes form the basis of nucleic
acid vaccines, or provide targets to allow preparation of attenuated mycobacteria for vaccines
against mycobacterial infections.  Similarly, peptides encoded by said induced or up-regulated
genes are employed in vaccines.  In a further embodiment, the identified genes/peptides
provide the means for identifying the presence of a mycobacterial infection in a clinical
sample by nucleic acid probe or antibody detection.

<>

<1>Jamoussi, K., Lazowska, J.
<2>Intragenic suppressors that restore the splicing and homing activities of the protein encoded by the second intron of the Saccharomyces capensis cyt b gene.
<3>Curr. Genet.
<4>38
<5>276-282
<6>2000
<7>The second (bi2) intron of the mitochondrial cyt b gene from Saccharomyces capensis encodes a
bifunctional protein which acts both
as a maturase, promoting intron splicing, and as a homing-endonuclease,
I-ScaI, promoting intron mobility. In this work we isolated and
characterized revertants from a respiratory-deficient mutant in which
both functions of the protein have been lost. Intragenic revertants
resulted mainly from monosubstitutions in the mutated codon and in one
case from a distant second site mutation. All novel variants of the S.
capensis bi2 intron-encoded protein are competent for the maturase
activity but only two of them can partially complement the homing
function.

<>

<1>Jancovich, J.K., Bremont, M., Touchman, J.W., Jacobs, B.L.
<2>Evidence for Multiple Recent Host Species Shifts among the Ranaviruses (Family Iridoviridae).
<3>J. Virol.
<4>84
<5>2636-2647
<6>2010
<7>Members of the genus Ranavirus (family Iridoviridae) have been recognized
as major viral pathogens of cold-blooded vertebrates. Ranaviruses have
been associated with amphibians, fish, and reptiles. At this time, the
relationships between ranavirus species are still unclear. Previous
studies suggested that ranaviruses from salamanders are more closely
related to ranaviruses from fish than they are to ranaviruses from other
amphibians, such as frogs. Therefore, to gain a better understanding of
the relationships among ranavirus isolates, the genome of epizootic
hematopoietic necrosis virus (EHNV), an Australian fish pathogen, was
sequenced. Our findings suggest that the ancestral ranavirus was a fish
virus and that several recent host shifts have taken place, with
subsequent speciation of viruses in their new hosts. The data suggesting
several recent host shifts among ranavirus species increase concern that
these pathogens of cold-blooded vertebrates may have the capacity to cross
numerous poikilothermic species barriers and the potential to cause
devastating disease in their new hosts.

<>

<1>Jancovich, J.K., Mao, J., Chinchar, V.G., Wyatt, C., Case, S.T., Kumar, S., Valente, G., Subramanian, S., Davidson, E.W., Collins, J.P., Jacobs, B.L.
<2>Genomic sequence of a ranavirus (family Iridoviridae) associated with salamander mortalities in North America.
<3>Virology
<4>316
<5>90-103
<6>2003
<7>Disease is among the suspected causes of amphibian population declines, and an iridovirus and
a chytrid fungus are the primary pathogens
associated with amphibian mortalities. Ambystoma tigrinum virus (ATV) and
a closely related strain, Regina ranavirus (RRV), are implicated in
salamander die-offs in Arizona and Canada, respectively. We report the
complete sequence of the ATV genome and partial sequence of the RRV
genome. Sequence analysis of the ATV/RRV genomes showed marked similarity
to other ranaviruses, including tiger frog virus (TFV) and frog virus 3
(FV3), the type virus of the genus Ranavirus (family Iridoviridae), as
well as more distant relationships to lymphocystis disease virus, Chilo
iridescent virus, and infectious spleen and kidney necrosis virus.
Putative open reading frames (ORFs) in the ATV sequence identified 24
genes that appear to control virus replication and block antiviral
responses. In addition, >50 other putative genes, homologous to ORFs in
other iridoviral genomes but of unknown function, were also identified.
Sequence comparison performed by dot plot analysis between ATV and itself
revealed a conserved 14-bp palindromic repeat within most intragenic
regions. Dot plot analysis of ATV vs RRV sequences identified several
polymorphisms between the two isolates. Finally, a comparison of ATV and
TFV genomic sequences identified genomic rearrangements consistent with
the high recombination frequency of iridoviruses. Given the adverse
effects that ranavirus infections have on amphibian and fish populations,
ATV/RRV sequence information will allow the design of better diagnostic
probes for identifying ranavirus infections and extend our understanding
of molecular events in ranavirus-infected cells.

<>

<1>Jang, J.Y., Lim, H.I., Park, H.W., Choi, H.J., Kim, T.W., Kang, M., Lee, J.H.
<2>Draft Genome Sequence of Lactobacillus plantarum wikim18, Isolated from Korean Kimchi.
<3>Genome Announcements
<4>2
<5>e00467-14
<6>2014
<7>This report describes the draft genome sequence of Lactobacillus plantarum strain wikim18,
isolated from the traditional Korean food kimchi. The reads generated by
Ion Torrent PGM were assembled into 327 contigs. RAST annotation of the genome
revealed 12 tRNAs and 3,316 protein-coding gene sequences.

<>

<1>Jang, K.H., Britz, M.L.
<2>Improved electrotransformation frequencies of Corynebacterium glutamicum using cell-surface mutantstransformation via electroporation using plasmid DNA.
<3>Biotechnol. Lett.
<4>22
<5>539-545
<6>2000
<7>6 Strains of Corynebacterium glutamicum were examined for electrotransformation using
heterologous or homologous DNA. These were:
AS019, a spontaneous rifampicin-resistant strain of ATCC 13059; MLB133
and MLB194, auxotrophic cell surface mutants derived from ATCC 13059 by
exposure to ethylmethane sulfonate; ATCC 13032; and RM3 and RM4,
restriction modification mutants of ATCC 13032. Heterologous DNA was
plasmid pCSL17 purified from Escherichia coli LE392. Homologous DNA was
pCSL17 from C. glutamicum AS019. Transformation by electroporation
involved a single pulse (2.5 kV, 25 uF) using a Gene-Pulser system,
with subsequent culture on LBG medium containing glycine and
isonicotinic acid hydrazide (INH). Transformation efficiency of MLB133
was up to 100-fold higher than for AS019 and, when using heterologous
derived DNA, MLB133 showed efficiencies comparable to, or better than,
RM3 and RM4, demonstrating the importance of cell surface structures in
impeding DNA uptake. MLB133 had a thinner cell wall than AS019, and
growth in glycine or INH further diminished its thickness. The impact
of glycine and INH on the mycolic composition of the strains is
discussed. (20 ref)

<>

<1>Jang, K.H., Chambers, P., Britz, M.L.
<2>Analysis of nucleotide methylation in DNA from Corynebacterium glutamicum and related species.
<3>FEMS Microbiol. Lett.
<4>136
<5>309-315
<6>1996
<7>Plasmid DNA (pCSL17) isolated from Corynebacterium glutamicum transformed recipient McrBC+
strains of Escherichia coli with lower efficiency than McrBC- strains, confirming a previous
report by Tauch et al. which inferred that C. glutamicum DNA contains methylcytidine.
Analysis of nucleotides in C. glutamicum-derived chromosomal and plasmid DNA failed to detect
significant levels of methylated adenosine, but methylated cytidine was readily detected.
Restriction enzymes which are inhibited by the presence of methylcytidine in their recognition
sequence failed to cut pCSL17 from C. glutamicum, whereas enzymes which require methylation at
adenosine in GATC sequences failed to cut.  Failure of HaeIII to cut two specific sites of C.
glutamicum-derived pCSL17 identified the first cytidine in the sequence GGCCGC as one target
of methylation in this species, which contains the methyltransferase recognition sequence.
Although Brevibacterium lactofermentum-derived DNA showed a similar methylation pattern by
HPLC analysis, HaeIII cleaved these GGCCGC sites, suggesting differences in the specificity of
methylation between these two species.  Results for all analyses of B. flavum DNA were
identical to those for C. glutamicum.

<>

<1>Jang, K.H., Chambers, P.J., Britz, M.L.
<2>Identification of a sequence containing methylated cytidine in Corynebacterium glutamicum and Brevibacterium flavum using bisulfite DNA derivatization and sequencing.
<3>J. Microbiol. Biotechnol.
<4>11
<5>819-824
<6>2001
<7>The principal DNA modification systems of the amino-acid-producing bacteria Corynebacterium
glutamicum AS019, Brevibacterium flavum BF4,
and B. lactofermentum BL1 was investigated using two approaches;
digestion of plasmid DNA isolated from these species using TseI and
Fnu4HI, and sequence analysis of the putative methyltransferase target
sites following the derivatization of DNA using metabisulfite
treatment. The C. glutamicum and B. flavum strains showed similar
digestion patterns to the two enzymes, indicating that the target for
cytidine methyltransferase recognizes 5'-GCSGC-3' (where S is either G
or C). Mapping the methylated cytidine sites by bisulfite
derivatization, followed by PCR amplification and sequencing, was only
possible when the protocol included an additional step eliminating any
underivatized DNA after PCR amplification, thereby indicating that the
derivatization was not 100% efficient. This may have been due to the
high G-C content of this genus. It was confirmed that C. glutamicum
AS019 and B. flavum BF4 methylated the cytidine in the Gm(5)CCGC
sequences, yet there were no similar patterns of methylation in B.
lactofermentum, which was consistent with the distinctive degradation
pattern seen for the above enzymes. These findings demonstrate the
successful application of a modified bisulfite derivatization method
with the Corynebacterium species for determining methylation patterns,
and showed that different species in the genus contain distinctive
restriction and modification systems.

<>

<1>Jang, K.J., Lee, H., Jin, H.L., Park, Y., Nam, J.M.
<2>Restriction-Enzyme-Coded Gold-Nanoparticle Probes for Multiplexed DNA Detection.
<3>Small
<4>5
<5>2665-2668
<6>2009
<7>The development of a multiplexed DNA detection assay has been of great interest in
gene-expression profiling, drug screening, clinical diagnostics, and the detection of
pathogens. However, it is still a challenging task to detect many different targets in one
sample while retaining high target sensitivity. Restriction enzymes that can recognize and
cleave specific double-stranded DNA (dsDNA) sequences have been widely used in molecular
biology and genetic engineering.  Over 2000 site-specific endonucleases have been identified
so far. Restriction enzymes have been used with DNA-modified gold nanoparticles (AuNPs) in
various applications, such as release of the particle from the surface, conformational effects
of nanoparticles on bioactivity, nanoparticle assembly and disassembly, and optimal conditions
of efficient biomanipulation. The availability of numerous enzymatic restriction DNA sequences
as well as the high specificity of restriction enzymes could be very useful aspects in a
multiplexed DNA detection assay.

<>

<1>Jang, S.H., Kim, J., Kim, J., Hong, S., Lee, C.
<2>Genome Sequence of Cold-Adapted Pseudomonas mandelii Strain JR-1.
<3>J. Bacteriol.
<4>194
<5>3263
<6>2012
<7>Pseudomonas mandelii is a cold-adapted bacterium that can grow at 4 degrees C but not at 37
degrees C. Here we report the draft genome sequence of P. mandelii
strain JR-1.

<>

<1>Jang, S.H., Yoon, B.H., Chang, H.I.
<2>Complete nucleotide sequence of the temperate bacteriophage LBR48, a new member of the family Myoviridae.
<3>Arch. Virol.
<4>156
<5>319-322
<6>2011
<7>The complete genomic sequence of LBR48, a temperate bacteriophage induced
from a lysogenic strain of Lactobacillus brevis, was found to be 48,211
nucleotides long and to contain 90 putative open reading frames. Based on
structural characteristics obtained from microscopic analysis and nucleic
acid sequence determination, phage LBR48 can be classified as a member of
the family Myoviridae. Analysis of the genome showed the conserved gene
order of previously reported phages of the family Siphoviridae from lactic
acid bacteria, despite low nucleotide sequence similarity. Analysis of the
attachment sites revealed 15-nucleotide-long core sequences.

<>

<1>Jang, Y., Oh, H.M., Kang, I., Lee, K., Yang, S.J., Cho, J.C.
<2>Genome sequence of strain IMCC3088, a proteorhodopsin-containing marine bacterium belonging to the OM60/NOR5 clade.
<3>J. Bacteriol.
<4>193
<5>3415-3416
<6>2011
<7>Strain IMCC3088, cultivated from the Yellow Sea, is a novel isolate belonging to the OM60/NOR5
clade and is closely related to clone OM241,
Congregibacter litoralis, and strain HTCC2080. Here the genome sequence of
strain IMCC3088 is presented, showing the absence of photosynthetic gene
clusters and the presence of proteorhodopsin.

<>

<1>Jang, Y., Oh, H.M., Kim, H., Kang, I., Cho, J.C.
<2>Genome Sequence of Strain IMCC1989, a Novel Member of the Marine Gammaproteobacteria.
<3>J. Bacteriol.
<4>193
<5>3672-3673
<6>2011
<7>Strain IMCC1989 is a novel member of the oligotrophic marine Gammaproteobateria (OMG) group,
and is closely related with a symbiont
group of genera Teredinibacter and 'Candidatus Endobugula'. Here we
present the genome sequence of strain IMCC1989 that was isolated from the
Yellow Sea by using dilution-to-extinction culturing.

<>

<1>Jangir, P.K., Singh, A., Shivaji, S., Sharma, R.
<2>Genome Sequence of the Alkaliphilic Bacterium Nitritalea halalkaliphila Type Strain LW7, Isolated from Lonar Lake, India.
<3>J. Bacteriol.
<4>194
<5>5688-5689
<6>2012
<7>An alkaliphilic bacterium, Nitritalea halalkaliphila LW7, which belongs to the family
Cyclobacteriacae in the phylum Bacteroidetes, was isolated from Lonar Lake
in Maharastra, India. Here we announce the draft genome sequence of the type
strain LW7, which contains 3,633,701 bp with a G+C content of 48.58%.

<>

<1>Jankowitsch, F., Schwarz, J., Ruckert, C., Gust, B., Szczepanowski, R., Blom, J., Pelzer, S., Kalinowski, J., Mack, M.
<2>The genome sequence of the bacterium Streptomyces davawensis JCM 4913 and heterologous production of the unique antibiotic roseoflavin.
<3>J. Bacteriol.
<4>194
<5>6818-6827
<6>2012
<7>Streptoymces davawensis JCM 4913 synthesizes the antibiotic roseoflavin, a structural
riboflavin (vitamin B(2)) analog. Here we report the 9,466,619 base
pair linear chromosome of S. davawensis JCM 4913 and a 89,331 base pair linear
plasmid. The sequence has an average G + C content of 70.58% and contains six
rRNA operons (16S-23S-5S) and 69 tRNA genes. The 8,616 predicted protein-coding
sequences include 32 clusters coding for secondary metabolites several of which
are unique to S. davawensis. The chromosome contains long terminal inverted
repeats of 33,255 bp each and atypical telomeres. Sequence analysis with regard
to riboflavin biosynthesis revealed three different patterns of gene organization
in Streptomyces species. Heterologous expression of a set of genes present on a
subgenomic fragment of S. davawensis resulted in the production of roseoflavin by
the host Streptomyces coelicolor M1152. Phylogenetic analysis revealed that S.
davawensis is a close relative to Streptomyces cinnabarinus and, much to our
surprise, we found that the latter bacterium is a roseoflavin producer as well.

<>

<1>Janosi, L., Kaji, A.
<2>Molecular cloning and expression of T-even phage specific restriction enzyme coded by drug resistance plasmid Rts1.
<3>FASEB J.
<4>6
<5>A216
<6>1992
<7>Rts1 is a drug resistance factor which restricts the growth of T-even phages at
32C but not at 42C (Ishaq & Kaji, 1980, J. Biol. Chem. 255:4040).  Using
ampicillin and E. coli bacteriophage T4 double selection, a 2.4 kilobase (kb)
EcoRI fragment of Rts1 DNA was cloned into pUC19 and pBluescript SK+ plasmids.
The determinant carried by the insert codes for restriction of multiplication
of T-even phages in E. coli 20SO.  The restriction phenomenon appeared
independently from the orientation of the insert relative to the lac promoter
of the carrier plasmids suggesting that the expression of the responsible
determinant was controlled by its own promoter.  Nucleotide squence
determination of the entire insert revealed the presence of two larger open
reading frames (ORFs), both on the same DNA strand.  One, coding for 294 amino
acids, proved to be related to phage restriction because deletions within this
ORF by various restriction endonucleases inactivated the activity.  Unusaul
feature of this ORF is that the distance between the most likely S.D. sequence
and the ATG codon spans only two nucleotides.

<>

<1>Janosi, L., Yonemitsu, H., Hong, H., Kaji, A.
<2>Molecular cloning and expression of a novel hydroxymethylcytosine-specific restriction enzyme (PvuRts1I) modulated by glucosylation of DNA.
<3>J. Mol. Biol.
<4>242
<5>45-61
<6>1994
<7>The kanamycin resistance plasmid Rts1 restricts the growth of bacteriophage T2, T4 and T6. The
DNA of these phage contains hydroxymethylcytosine (HMC) in place of regular cytosine and is
modified by glycosylation. When HMC is not glucosylated, as in the DNA of glucosyl
transferase-deficient T4 phage, this restriction becomes less apparent, a phenomenon not
observed with any other known restriction systems. On the other hand, glucosylation of HMC to
T6 phage leads to a less efficient restriction, while restriction of bacteriophage T2 remains
unchanged. The modulating effect of glucose cannot be seen when cells contain a large amount
of this enzyme, as in the case when multiple copies of its determinant are present in the
cells. T-odd phage and bacteriophage lambda are not restricted by Rts1 suggesting that the
restriction is specific to DNA containing HMC. The restriction phenotype is due to a single
gene coding for a polypeptide of 293 amino acids. This enzyme has been named PvuRts1I. A gene
with the sequence motifs similar to modification enzymes was found upstream of the gene coding
for PvuRts1I. This gene, however, neither modifies the restriction phenotype of PvuRts1I, nor
codes for detectable modification enzyme. T4 mutants with increased resistance to PvuRts1I
appear to have deficiency in their beta-glucosyl transferase enzyme.

<>

<1>Jans, C., Lacroix, C., Follador, R., Stevens, M.J.
<2>Complete Genome Sequence of the Probiotic Bifidobacterium thermophilum Strain RBL67.
<3>Genome Announcements
<4>1
<5>e00191-13
<6>2013
<7>Bifidobacterium thermophilum RBL67, an isolate from infant feces, exhibits bacteriocin-like
antimicrobial activity against Listeria spp. and Salmonella spp.
and protects HT29-MTX cells against Salmonella infection. Here, the complete
genome sequence of the probiotic B. thermophilum strain RBL67 is presented.

<>

<1>Janscak, P., Abadjieva, A., Firman, K.
<2>The type I restriction endonuclease R.EcoR124I: over-production and biochemical properties.
<3>J. Mol. Biol.
<4>257
<5>977-991
<6>1996
<7>In this paper we describe a two-plasmid system which allows over-production of
the R.EcoR124I restriction endonuclease.  The endonuclease has been purified to homogeneity in
milligram amounts and has been shown to be fully active for both restriction and modification.
Unexpectedly, the enzyme was found to require only ATP and Mg2+ for ATPase activity and
DNA cleavage; S-adenosyl methionine (SAM), which has been described as a cofactor of type I
restriction enzymes, is not required by R.EcoR124I.  However, SAM was found to stimulate the
rate of ATPase activity and DNA cleavage.  This may occur through an increase in specific DNA
binding by R.EcoR124I in the presence of SAM, as indicated by our surface plasmon resonance
experiments.  These functional differences from the well described R.EcoKI restriction
endonuclease are reflected in a possible structural difference between the two enzymes, namely
that
the stoichiometry of R.EcoR124I appears to be R1M2S1 while that of R.EcoKI is R2M2S1.
Supercoiled DNA with one or two SR124I recognition sites is cleaved by the same mechanism
inferring cooperation between specifically bound and excess enzymes.  Nicked-circle DNA is an
intermediate of cleavage reaction.  Cleavage of DNA was inhibited by an increased degree of
negative supercoiling, which may reflect an increased difficulty for the enzyme to translocate
the
DNA.  Hemi-methylated DNA was the preferred substrate for methylation.

<>

<1>Janscak, P., Bickle, T.A.
<2>The DNA recognition subunit of the type IB restriction-modification enzyme EcoAI tolerates circular permutations of its polypeptide chain.
<3>J. Mol. Biol.
<4>284
<5>937-948
<6>1998
<7>The DNA specificity subunit (HsdS) of type I restriction-modification enzymes is composed of
two independent target recognition domains and several regions whose amino acid sequence is
conserved within an enzyme family.  The conserved regions participate in intersubunit
interactions with two modification subunits (HsdM) and two restriction subunits (HsdR) to form
the complete endonuclease.  It has been proposed that the domains of the HsdS subunit have a
circular organization providing the required symmetry for their interaction with the other
subunits and with the bipartite DNA target.  To test this model, we circularly permuted the
HsdS subunit of the type IB R-M enzyme EcoAI at the DNA level by direct linkage of codons for
original termini and introduction of new termini elsewhere along the N-terminal and central
conserved regions.  By analysing the activity of mutant enzymes, two circularly permuted
variants of HsdS that had termini located at equivalent positions in the N-terminal and
central repeats, respectively, were found to fold into a functional DNA recognition subunit
with wild-type specificity, suggesting a close proximity of the N and C termini in the native
protein.  The wild-type HsdS subunit was purified to homogeneity and shown to form a stable
trimeric complex with HsdM, M2S1, which was fully active as a DNA methyltransferase.  Gel
electrophoretic mobility shift assays revealed that the HsdS protein alone was not able to
form a specific complex with a 30-mer oligoduplex containing a single EcoAI recognition site.
However, addition of stoichiometric amounts of hsdM to HsdS led to efficient specific DNA
binding.  Our data provide evidence for the circular organization of domains of the HsdS
subunit.  In addition, they suggest a possible role of hsdM subunits in the formation of this
structure.

<>

<1>Janscak, P., Bickle, T.A.
<2>DNA supercoiling during ATP-dependent DNA translocation by the type I restriction enzyme EcoAI.
<3>J. Mol. Biol.
<4>295
<5>1089-1099
<6>2000
<7>Type I restriction enzymes cleave DNA at non-specific sites far from their recognition
sequence as a consequence of ATP-dependent DNA translocation past the
enzyme. During this reaction, the enzyme remains bound to the recognition sequence and
translocates DNA towards itself simultaneously from both directions, generating
DNA loops, which appear to be supercoiled when visualised by electron microscopy. To further
investigate the mechanism of DNA translocation by type I restriction
enzymes, we have probed the reaction intermediates with DNA topoisomerases. A DNA
cleavage-deficient mutant of EcoAI, which has normal DNA translocation and
ATPase activities, was used in these DNA supercoiling assays. In the presence of eubacterial
DNA topoisomerase I, which specifically removes negative supercoils, the
EcoAI mutant introduced positive supercoils into relaxed plasmid DNA substrate in a reaction
dependent on ATP hydrolysis. The same DNA supercoiling activity followed by
DNA cleavage was observed with the wild-type EcoAI endonuclease. Positive supercoils were not
seen when eubacterial DNA topoisomerase I was replaced by eukaryotic
DNA topoisomerase I, which removes both positive and negative supercoils. Furthermore,
addition of eukaryotic DNA topoisomerase I to the product of the supercoiling
reaction resulted in its rapid relaxation. These results are consistent with a model in which
EcoAI translocation along the helical path of closed circular DNA duplex
simultaneously generates positive supercoils ahead and negative supercoils behind the moving
complex in the contracting and expanding DNA loops, respectively. In addition,
we show that the highly positively supercoiled DNA generated by the EcoAI mutant is cleaved by
EcoAI wild-type endonuclease much more slowly than relaxed DNA. This
suggests that the topological changes in the DNA substrate associated with DNA translocation
by type I restriction enzymes do not appear to be the trigger for DNA
cleavage.

<>

<1>Janscak, P., Dryden, D.T.F., Firman, K.
<2>Analysis of the subunit assembly of the type IC restriction-modification enzyme EcoR124I.
<3>Nucleic Acids Res.
<4>26
<5>4439-4445
<6>1998
<7>Type I restriction-modification enzymes are composed of three different subunits, of which
HsdS determines DNA specificity, HsdM is responsible for DNA methylation and HsdR is required
for restriction.  The HsdM and hsdS subunits can also form an independent DNA
methyltransferase with a subunit stoichiometry of M2S1.  We found that the purified EcoR124I
R-M enzyme was a mixture of two species as detected by the presence of two differently
migrating specific DNA-protein complexes in a gel retardation assay.  An analysis of protein
subunits isolated from the complexes indicated that the larger species had a stoichiometry of
R2M2S1 and the smaller species had a stoichiometry of R1M2S1.  In vitro analysis of subunit
assembly revealed that while binding of the first HsdR subunit to the M2S1 complex with an
apparent Kd of ~2.4 x 10^-7M.  Functional assays have shown that only the R2M2S1 complex is
capable of DNA cleavage, however, the R1M2S1 complex retains ATPase activity.  The relevance
of this situation is discussed in terms of the regulation of restriction activity in vivo upon
conjugative transfer of a plasmid-born R-M system into an unmodified host cell.

<>

<1>Janscak, P., MacWilliams, M.P., Sandmeier, U., Nagaraja, V., Bickle, T.A.
<2>DNA translocation blockage, a general mechanism of cleavage site selection by type I restriction enzymes.
<3>EMBO J.
<4>18
<5>2638-2647
<6>1999
<7>Type I restriction enzymes bind to a specific DNA sequence and subsequently translocate DNA
past the complex to reach a non-specific cleavage site.  We have examined several potential
blocks to DNA translocation, such as positive supercoiling or a Holliday junction, for their
ability to trigger DNA cleavage by type I restriction enzymes.  Introduction of positive
supercoiling into plasmid DNA did not have a significant effect on the rate of DNA cleavage by
EcoAI endonuclease nor on the enzyme's ability to select cleavage sites randomly throughout
the DNA molecule.  Thus, positive supercoiling does not prevent DNA translocation.  EcoR124II
endonuclease cleaved DNA at Holliday junctions present on both linear and negatively
supercoiled substrates.  The latter substrate was cleaved by a single enzyme molecule at two
sites, one on either side of the junction, consistent with a bi-directional translocation
model.  Linear DNA molecules with two recognition sites for endonucleases from different type
I families were cut between the sites when both enzymes were added simultaneously but not when
a single enzyme was added.  We propose that type I restriction enzymes can track along a DNA
substrate irrespective of its topology and cleave DNA at any barrier that is able to halt the
translocation process.

<>

<1>Janscak, P., Sandmeier, U., Bickle, T.A.
<2>Single amino acid substitutions in the HsdR subunit of the type IB restriction enzyme EcoAI uncouple the DNA translocation and DNA cleavage activities of the enzyme.
<3>Nucleic Acids Res.
<4>27
<5>2638-2643
<6>1999
<7>Type I restriction enzymes bind to specific DNA sequences but subsequently translocate
non-specific DNA past the complex in a reaction coupled to ATP hydrolysis and cleave DNA at
any barrier that can halt the translocation process. The restriction subunit of these enzymes,
HsdR, contains a cluster of seven amino acid sequence motifs typical of helicase superfamily
II, that are believed to be relevant to the ATP-dependent DNA translocation. Alignment of all
available HsdR sequences reveals an additional conserved region at the protein N-terminus with
a consensus sequence reminiscent of the P-D...(D/E)-X-K catalytic motif of many type II
restriction enzymes. To investigate the role of these conserved residues, we have produced
mutants of the type IB restriction enzyme EcoAI. We have found that single alanine
substitutions at Asp-61, Glu-76 and Lys-78 residues of the HsdR subunit abolished the
enzyme's restriction activity but had no effect on its ATPase and DNA translocation
activities, suggesting that these residues are part of the active site for DNA cleavage.

<>

<1>Janscak, P., Sandmeier, U., Szczelkun, M.D., Bickle, T.A.
<2>Subunit assembly and mode of DNA cleavage of the type III restriction endonucleases EcoP1I and EcoP15I.
<3>J. Mol. Biol.
<4>306
<5>417-431
<6>2001
<7>DNA cleavage by type III restriction endonucleases requires two inversely oriented asymmetric
recognition sequences and results from
ATP-dependent DNA translocation and collision of two enzyme molecules.
Here, we characterized the structure and mode of action of the related
EcoP1I and EcoP15I enzymes. Analytical ultracentrifugation and gel
quantification revealed a common Res(2)Mod(2) subunit stoichiometry.
Single alanine substitutions in the putative nuclease active site of
ResP1 and ResP15 abolished DNA but not ATP hydrolysis, whilst a
substitution in helicase motif VI abolished both activities. Positively
supercoiled DNA substrates containing a pair of inversely oriented
recognition sites were cleaved inefficiently, whereas the corresponding
relaxed and negatively supercoiled substrates were cleaved efficiently,
suggesting that DNA overtwisting impedes the convergence of the
translocating enzymes. EcoP1I and EcoP15I could co-operate in DNA
cleavage on a circular substrate containing several EcoP1I sites
inversely oriented to a single EcoP15I site; cleavage occurred
predominantly at the EcoP15I site. EcoP15I alone showed nicking
activity on these molecules, cutting exclusively the top DNA strand at
its recognition site. This activity was dependent on enzyme
concentration and local DNA sequence. The EcoP1I nuclease mutant
greatly stimulated the EcoP15I nicking activity, while the EcoP1I motif
VI mutant did not. Moreover, combining an EcoP15I nuclease mutant with
wild-type EcoP1I resulted in cutting the bottom DNA strand at the
EcoP15I site. These data suggest that double-strand breaks result from
top strand cleavage by a Res subunit proximal to the site of cleavage,
whilst bottom strand cleavage is catalysed by a Res subunit supplied in
trans by the distal endonuclease in the collision complex.

<>

<1>Janscak, P., Weiserova, M., Hubacek, J., Holubova, I., Dutta, C.F., Firman, K.
<2>Two temperature-sensitive mutations in the DNA binding subunit of EcoKI with differing properties.
<3>FEMS Microbiol. Lett.
<4>182
<5>99-104
<6>2000
<7>Two temperature-sensitive mutations in the hsdS gene, which encodes the DNA specificity
subunit of the type IA restriction-modification system EcoKI, designated Sts1 (Ser(340)Phe)
and Sts2 (Ala(204)Thr) had a different impact on restriction-modification functions in vitro
and in vivo. The enzyme activities of the Sts1 mutant were temperature-sensitive in vitro and
were reduced even at 30 degrees C (permissive temperature). Gel retardation assays revealed
that the Sts1 mutant had significantly decreased DNA binding, which was temperature-sensitive.
In contrast the Sts2 mutant did not show differences from the wild-type enzyme even at 42
degrees C. Unlike the HsdSts1 subunit, the HsdSts2 subunit was not able to compete with the
wild-type subunit in assembly of the restriction enzyme in vivo, suggesting that the Sts2
mutation affects subunit assembly. Thus, it appears that these two mutations map two important
regions in HsdS subunit responsible for DNA-protein and protein-protein interactions,
respectively.

<>

<1>Janssen, P.J., Morin, N., Mergeay, M., Leroy, B., Wattiez, R., Vallaeys, T., Waleron, K., Waleron, M., Wilmotte, A., Quillardet, P., de Marsac, N.T., Talla, E., Zhang, C.C., Leys, N.
<2>Genome sequence of the edible cyanobacterium Arthrospira sp. PCC 8005.
<3>J. Bacteriol.
<4>192
<5>2465-2466
<6>2010
<7>We determined the genome sequence of Arthrospira sp. PCC 8005, a cyanobacterial strain of
great interest to the European Space Agency for
its nutritive value and oxygenic properties in the Micro-Ecological Life
Support System Alternative (MELiSSA) biological life support system for
long-term manned missions into space.

<>

<1>Janto, B. et al.
<2>Genome of alkaliphilic Bacillus pseudofirmus OF4 reveals adaptations that support the ability to grow in an external pH range from 7.5 to 11.4.
<3>Environ. Microbiol.
<4>13
<5>3289-3309
<6>2011
<7>Bacillus pseudofirmus OF4 is an extreme but facultative alkaliphile that grows
non-fermentatively in a pH range from 7.5 to above 11.4 and can withstand large
sudden increases in external pH. It is a model organism for studies of
bioenergetics at high pH, at which energy demands are higher than at neutral pH
because both cytoplasmic pH homeostasis and ATP synthesis require more energy.
The alkaliphile also tolerates a cytoplasmic pH > 9.0 at external pH values at
which the pH homeostasis capacity is exceeded, and manages other stresses that
are exacerbated at alkaline pH, e.g. sodium, oxidative and cell wall stresses.
The genome of B. pseudofirmus OF4 includes two plasmids that are lost from some
mutants without viability loss. The plasmids may provide a reservoir of mobile
elements that promote adaptive chromosomal rearrangements under particular
environmental conditions. The genome also reveals a more acidic pI profile for
proteins exposed on the outer surface than found in neutralophiles. A large array
of transporters and regulatory genes are predicted to protect the alkaliphile
from its overlapping stresses. In addition, unanticipated metabolic versatility
was observed, which could ensure requisite energy for alkaliphily under diverse
conditions.

<>

<1>Janulaitis, A.
<2>Restriction Enzymes and their Applications.
<3>Trends in Science and Technology. Biotechnology series., Institute for Scientific and Technological Information of the USSR, Debabov, V.G., Moscow
<4>17
<5>1-204
<6>1989
<7>
<>

<1>Janulaitis, A.
<2>A novel approach to the study of DNA-protein interactions using related restriction endonucleases that recognize overlapping nucleotide sequences.
<3>Biologija
<4>0
<5>28-31
<6>1996
<7>Multiple attempts to change the specificity of restriction enzymes by substituting amino acids
predicted by crystallographic analysis to be involved in substrate recognition have so far
been unsuccessful.  In general, the results argue that the potential of this approach in study
of RE is limited.  A novel approach to the analysis of such structure-function relationships
is therefore suggested, which is based on the investigation of structurally related RE which
recognize overlapping (but not identical) nucleotide sequences.  In an attempt to identify
such RE, 22 genes encoding RE were cloned, sequenced, and their deduced amino acid sequences
compared with those of previously described RE by computer analysis.  For the first time a
group of 18 RE is demonstrated, which can be arranged in pairs (9 pairs) on the basis of
significant overlap in recognition sequences, and some aa sequence similarity.  Indirect
evidence is given to support the contention that members of each pair are structurally related
to provide, in turn, the basis of a new model system for investigation of the role of
structural changes in the development of changes in specificity.

<>

<1>Janulaitis, A.
<2>Restriction endonucleases.
<3>Zh. Vses. Khim.
<4>29
<5>133-138
<6>1984
<7>
<>

<1>Janulaitis, A., Bitinaite, J., Jaskeleviciene, B.
<2>A new sequence-specific endonuclease from Gluconobacter suboxydans.
<3>FEBS Lett.
<4>151
<5>243-247
<6>1983
<7>The isolation of sequence-specific endonucleases from Gluconobacter
dioxyacetonicus (IAM 1814 and IAM 1840) and Gluconobacter oxydans sub.
melanogenes (IAM 1836) has been reported.  We have examined six Gluconobacter
suboxydans strains for the presence of the enzymes of this type and discovered
in two of them restriction endonucleases of identical specificity (named GsuI
and GsbI).Here, we describe the isolation procedure of the new site-specific
endonuclease GsuI, which recognizes a hexanucleotide sequence 5' ...CTCCAG.

<>

<1>Janulaitis, A., Butkus, V.V., Petrusite, M.P., Bitinaite, J.B.
<2>Separation of type II restriction enzymes - comprises ultrasonic destruction of cells, fractionation on heparin-sepharose and chromatography on phospho-cellulose and blue sepharose.
<3>Soviet Patent Office
<4>SU 1698289 A
<5>
<6>1991
<7>The method comprises destruction of cells by ultrasonication, separation of extract on
heparin-sepharose and chromatography on phosphocellulose and blue sepharose in 0.01M potassium
phosphate buffer at pH 7.4, 0.007M 2-mercapto-ethanol, 0.001M EDTA, with elution of enzyme
from chromatographic column using NaCl gradient 0.1-0.8M for chromatography on
heparin-sepharose and phosphocellulose and NaCl gradient 0.01-0.1 M for chromatography on blue
sepharose. Tests show that the proposed method is suitable for separation of restrictases
XbaI, NotI, Eco147I, Alw26I, Alw44I, Bsp50I, Ecl136II, Eco81I, KpnI, Kpn2I, PaeI.

<>

<1>Janulaitis, A., Kazlauskiene, R., Lazareviciute, L., Gilvonauskaite, R., Steponaviciene, D., Jagelavicius, M., Petrusyte, M., Bitinaite, J., Vezeviciute, Z., Kiuduliene, E., Butkus, V.
<2>Taxonomic specificity of restriction-modification enzymes.
<3>Gene
<4>74
<5>229-232
<6>1988
<7>Meeting Abstract

<>

<1>Janulaitis, A., Klimasauskas, S., Petrusyte, M., Butkus, V.
<2>Cytosine modification in DNA by BcnI methylase yields N4-methylcytosine.
<3>FEBS Lett.
<4>161
<5>131-134
<6>1983
<7>The means by which bacteria protect their own DNA from their restriction
enzymes have not been fully investigated.  In all systems that have been
studied, cells produce a modification methylase in addition to the retriction
endonuclease.  Both enzymes recognize the same specific DNA sequence.  Not many
DNA methylases were studied in detail, but all of those studied methylate
either adenine to N6-methyladenine (m6A) or cytosine to 5-methylcytosine
(m5C).Recently a site-specific endonuclease and methylase BcnI, both of which
recognize the sequence 5'CC(C/G)GG, have been isolated from Bacillus
centrosporus strain RFLI.  We here describe the property of MBcnI to methylate
cytosine residues in DNA in vitro at the N4 position yielding N4-methylcytosine
(m4C).  The same minor base in DNA isolated from B. centrosporus was detected.
Such an unusual DNA modification is described for the first time.

<>

<1>Janulaitis, A., Marcinkeviciene, L., Petrusyte, M., Mironov, A.
<2>A new sequence-specific endonuclease from Streptococcus durans.
<3>FEBS Lett.
<4>134
<5>172-174
<6>1981
<7>Although a relatively large number of sequence-specific deoxyribonucleases
(class II restriction endonucleases) is now available, new endonucleases with
unique recognition sites are desirable, as they increase the flexibility of DNA
analysis and recombinant techniques.  We report here the isolation from
Streptococcus durans RFL 3 strain of a new enzyme (SduI) of this type, which
recognizes hexanucleotide palindromic sequence 5'-G(G/A/T)-GC(C/A/T)C
degenerated at two positions and consequently should cleave at the level of
nine possible sites.

<>

<1>Janulaitis, A., Marcinkeviciene, L.Y., Petrusyte, M.P.
<2>A specific endonuclease from Caulobacter fusiformis that cleaves only methylated DNA.
<3>Dokl. Akad. Nauk.
<4>262
<5>241-244
<6>1982
<7>None

<>

<1>Janulaitis, A., Petrusyte, M., Butkus, V.
<2>Three sequence-specific endonucleases from Escherichia coli RFL47.
<3>FEBS Lett.
<4>161
<5>213-216
<6>1983
<7>The characterization of the new restriction enzyme Eco47III recognizing a
hexanucleotide palindromic sequence 5'AGC^GCT and cleaving, as indicated by the
arrow, is reported.  It was isolated from Escherichia coli strain RFL47.
Another two specific endonuclease Eco471 (isoschizomer of AvaII and Ecoz4711
(isoschizomer of AsuI) were also found in this strain.  There are two Eco47III
recognition sites on lambda DNA at 20997 and 37060 basepairs.  The central
Eco47III fragment can be replaced by a cloned fragment in lambda vector mutant
tR2 gene; i.e. lambda gt.

<>

<1>Janulaitis, A., Petrusyte, M., Maneliene, Z., Klimasauskas, S., Butkus, V.
<2>Purification and properties of the Eco57I restriction endonuclease and methylase-prototypes of a new class (type IV).
<3>Nucleic Acids Res.
<4>20
<5>6043-6049
<6>1992
<7>The Eco57I restriction endonuclease and methylase were purified to homogeneity from the E.coli
RR1 strain carrying the Eco57IRM genes on a recombinant plasmid. The molecular weight of the
denaturated methylase is 63 kDa. The restriction endonuclease exists in a monomeric form with
an apparent molecular weight of 104-108 kDa. R.Eco57I also possesses methylase activity. The
methylation activities of both enzymes modify the outer A residue in the target sequence
5'CTGAAG yielding N6-methyladenine. M.Eco57I modifies both strands of the substrate while
R.Eco57I modifies only one. Only the methylase enzyme is stimulated by Ca2+. The restriction
endonuclease shows an absolute requirement for Mg2+ and is stimulated by AdoMet. ATP has no
influence on either activity of the enzymes. The subunit structure and enzymatic properties of
the Eco57I enzymes distinguish them from all other restriction-modification enzymes that have
been described previously. Therefore, RM.Eco57I may be regarded as a representative of a novel
class of restriction-modification systems, and we propose to classify it as type IV.

<>

<1>Janulaitis, A., Petrusyte, M.A., Jaskeleviciene, B.P., Krayev, A.S., Skryabin, K.G., Bayev, A.A.
<2>A new restriction endonuclease BcnI from Bacillus centrosporus RFL 1.
<3>FEBS Lett.
<4>137
<5>178-180
<6>1982
<7>Type II restriction endonucleases have proved to be an indispensable tool in
DNA cloning and sequencing studies.  Over 200 individual enzymes with >60
different specificities have been already described.  Here, we describe the
purification and the determination of cleavage specificity of a novel type II
restriction endonuclease from Bacillus centrosporus RFL 1.

<>

<1>Janulaitis, A., Petrusyte, M.P., Jaskeleviciene, B.P., Krayev, A.S., Skryabin, K.G., Bayev, A.A.
<2>A new restriction endonuclease BcnI from Bacillus centrosporus RFL1.
<3>Dokl. Akad. Nauk.
<4>257
<5>749-750
<6>1981
<7>Restriction endonucleases are finding wide use in the study of the structure
and function of genomes and the production of recombinant DNA molecules.  At
the present time, more than 50 restriction endonucleases with various substrate
specificities are known; however, the search for new restriction enzymes, as
before, is an urgent problem, since the detection of new restriction enzymes
expands the experimenter's possibilities in solving the indicated problems.

<>

<1>Janulaitis, A., Povilionis, P., Sasnauskas, K.
<2>Cloning of the modification methylase gene of Bacillus centrosporus in Escherichia coli.
<3>Gene
<4>20
<5>197-204
<6>1982
<7>The gene specifying a sequence-specific modification methylase of Bacillus centrosporus has
been cloned in Escherichia coli using the restriction endonuclease HindIII and the plasmid
pBR322.  The selection was based on detection of new methylation properties rendering
recombinant plasmids carrying the methylase gene nonsusceptible to BcnI endonuclease cleavage.
The presence of a 3.2-kb HindIII fragment in either orientation conferred BcnI resistance on
the recombinant plasmids.  These results suggest that the BcnI methylase gene is expressed in
E. coli under the control of a promoter located on the cloned fragment.  The relative level of
BcnI methylase enzyme in E. coli was similar to that in B. centrosporus.  The recombinant
clones do not exhibit any BcnI restriction-endonuclease activity.

<>

<1>Janulaitis, A., Rimseliene, R., Lubys, A.
<2>Nuclease.
<3>US Patent Office
<4>US 20030040614
<5>
<6>2003
<7>A process for producing a polynucleotide encoding a restriction endonuclease with an altered
specificity, which process comprises: (a) mutagenising a polynucleotide encoding a restriction
endonuclease with specificity for a recognition sequence so as to produce one or more mutated
polynucleotides; and (b) isolating therefrom a polynucleotide encoding a mutated restriction
endonuclease with specificity for an altered recognition sequence by selecting a
polynucleotide which expresses a restriction endonuclease with methylase specificity for the
altered recognition sequence.

<>

<1>Janulaitis, A., Rimseliene, R., Lubys, A.
<2>Nuclease.
<3>European Patent Office
<4>EP 1179596 A
<5>
<6>2002
<7>A process for producing a polynucleotide encoding a restriction endonuclease with an altered
specificity, which process comprises: (a) mutagenising a polynucleotide encoding a restriction
endonuclease with specificity for a recognition sequence so as to produce one or more mutated
polynucleotides; and (b) isolating therefrom a polynucleotide encoding a mutated restriction
endonuclease with specificity for an altered recognition sequence by selecting a
polynucleotide which expresses a restriction endonuclease with methylase specificity for the
altered recognition sequence.

<>

<1>Janulaitis, A., Rimseliene, R., Lubys, A.
<2>Nuclease.
<3>US Patent Office
<4>US 6893854 B
<5>
<6>2005
<7>A process for producing a polynucleotide encoding a restriction endonuclease with an altered
specificity, which process comprises:  (a) mutagenizing a polynucleotide encoding a mutated
restriction endonuclease  with specificity for a recognition sequence so as to produce one or
more mutated polynucleotides; and (b) isolating therefrom a polynucleotide encoding a mutated
restriction endonuclease with specificity for an altered recognition sequence by selecting a
polynucleotide which expresses a restriction endonuclease with methylase specificity for the
altered recognition sequence.

<>

<1>Janulaitis, A., Stakenas, P., Jaskeleviciene, B., Lebedenko, E.N., Berlin, Y.A.
<2>A new restriction endonuclease, CfrI from Citrobacter freundii.
<3>Bioorg. Khim.
<4>6
<5>1746-1748
<6>1980
<7>CfrI, a new site-specific restriction endonuclease, has been isolated from a
Citrobacter freundii strain.  The enzyme recognizes the sequence 5'
Py^G-G-C-C-Pu 3' Pu-C-C-G-G^Py in double-stranded DNA and cuts it between Py
and G residues to give 5'-protruding tetranucleotide ends G-G-C-C.

<>

<1>Janulaitis, A., Stakenas, P.S., Berlin, Y.A.
<2>A new site-specific endodeoxyribonuclease from Citrobacter freundii.
<3>FEBS Lett.
<4>161
<5>210-212
<6>1983
<7>Cfr10I, a site-specific endonuclease from Citrobacter freundii strain RFL10, was isolated. It
recognizes and cleaves the family of related sequences: 5'Pu^CCGGPy to generate DNA fragments
with 5' tetranucleotide extensions. Cfr10I may be useful in molecular cloning experiments,
especially in conjunction with other enzymes which generate the same terminal extensions. ion
have led to the detection of specific endodeoxyribonucleases (class II restriction
endonucleases), distinguished by an exceptionally high substrate specificity, which served as
the prerequisite for their use in molecular genetic and structural investigations of DNA.

<>

<1>Janulaitis, A., Stakenas, P.S., Bitinaite, J.B., Jaskeleviciene, B.P.
<2>Distribution of specific endodeoxyribonucleases in various strains of Citrobacter freundii.
<3>Dokl. Akad. Nauk.
<4>271
<5>483-485
<6>1983
<7>The discovery of the phenomenon of host specificity at the beginning of the
fifties and the investigation of its essence, realized in the discovery of
enzymes of the restriction-modification systems, have made a large contribution
to the development of modern molecular biology, genetics, and enzymology.  The
search for and study of enzymes of restriction and modification have led to the
detection of specific endodeoxyribonucleases (class II restriction
endonucleases), distinguished by an exceptionally high substrate specificity,
which served as the prerequisite for their use in molecular genetic and
structural investigations of DNA.

<>

<1>Janulaitis, A., Stakenas, P.S., Lebedenko, E.N., Berlin, Y.A.
<2>A new restriction endonuclease from Citrobacter freundii.
<3>Nucleic Acids Res.
<4>10
<5>6521-6530
<6>1982
<7>CfrI, a new restriction endonuclease of unique substrate specificity, has been
isolated from a Citrobacter freundii strain.  The enzyme recognizes a
degenerated sequence PyGGCCPu in double-strand DNA and cleaves it between Py
and G residues to yield 5'-protruding tetranucleotide ends GGCC.

<>

<1>Janulaitis, A., Stakenas, P.S., Petrusyte, M.P., Bitinaite, J.B., Klimasauskas, S.I., Butkus, V.V.
<2>Specificity of new restrictases and methylases.  Unusual modification of cytosine in position 4.
<3>Mol. Biol. (Mosk)
<4>18
<5>115-129
<6>1983
<7>Fourteen restrictases and four methylases were extracted and purified from 14 strains of
independent origin of the species Citrobacter freundii and Escherichia coli. The nucleotide
sequences recognized by the restrictases were determined by means of a comparison of the
character of DNA cleavage by the investigated and known enzymes, by study of the cleavage
frequency of substrates with a known primary structure, and by the mapping method. It was
established that Cfr10I is a new prototype recognizing the sequence 5'PuCCGGPy. The other
enzymes proved to be isoschizomers of known restrictases: Cfr5*, Cfr11I, Eco60I, and Eco61I
are isoschizomers of EcoRII; Cfr4I, Cfr8I, and Cfr12T, Sau96I; Cfr6I, PvuII; Crf9I, SmaI;
Eco6I, HgiJII; Eco32I, EcoRV; Eco52I, XmaIII; and Eco56I, NaeI. Several of these enzymes were
demonstrated for the first time in E. coli and C. freundii. A study of the specificity of the
methylases M.CfrI, M.Cfr6I, M.Cfr9I, and M.Cfr10I showed that these enzymes recognize the same
nucleotide sequence as does the restrictase extracted from the same strain. Modification of
DNA in vitro using M.CfrI and M.Cfr10I results in the production of 5-methylcytosine, and in
the case of M.Cfr6I and M.Cfr9I, N4-methylcytosine. G, where M.HpaII methylates the inner,
M.BsuFI/M.MspI the outer cytosine. Also the CCGG recognizing TRD of the multispecific B.
subtilis phage SPR Mtase is distinct from that of the host enzyme, possibly indicating
different requirements for TRDs operative in mono- and multispecific enzymes.

<>

<1>Janulaitis, A., Stankevicius, K., Lubys, A., Markauskas, A.
<2>Nuclease.
<3>European Patent Office
<4>EP 1176204 A
<5>
<6>2002
<7>A strand-specific polynucleotide nickase comprising an endonuclease which comprises a first
subunit and a second subunit and which recognizes an asymmetric nucleotide recognition
sequence, wherein the first subunit comprises a catalytic domain capable of cleaving one
strand of a DNA duplex, and the second subunit is incapable of cleaving the other strand of
the DNA duplex.

<>

<1>Janulaitis, A., Stankevicius, K., Lubys, A., Markauskas, A.
<2>Nuclease.
<3>US Patent Office
<4>US 20030148275 A
<5>
<6>2001
<7>A strand-specific polynucleotide nickase comprising an endonuclease which comprises a first
subunit and a second subunit and which recognizes an asymmetric nucleotide recognition
sequence, wherein the first subunit comprises a catalytic domain capable of cleaving one
strand of a DNA duplex, and the second subunit is incapable of cleaving the other strand of
the DNA duplex.

<>

<1>Janulaitis, A., Vaisvila, R., Timinskas, A., Klimasauskas, S., Butkus, V.
<2>Cloning and sequence analysis of the genes coding for Eco57I type IV restriction-modification enzymes.
<3>Nucleic Acids Res.
<4>20
<5>6051-6056
<6>1992
<7>A 6.3 kb fragment of E.coli RFL57 DNA coding for the type IV restriction-modification system
Eco57I was cloned and expressed in E.coli RR1. A 5775 bp region of the cloned fragment was
sequenced which contains three open reading frames (ORF). The methylase gene is 1623 bp long,
corresponding to a protein of 543 amino acids (62 kDa); the endonuclease gene is 2991 bp in
length (997 amino acids, 117 kDa). The two genes are transcribed convergently from different
strands with their 3'-ends separated by 69 bp. The third short open reading frame (186 bp, 62
amino acids) has been indentified, that precedes and overlaps by 7 nucleotides the ORF
encoding the methylase. Comparison of the deduced Eco57I endonuclease and methylase amino acid
sequences revealed three regions of significant similarity. Two of them resemble the conserved
sequence motifs characteristic of the DNA [adenine-N6] methylases. The third one shares
similarity with corresponding regions of the PaeR71, TaqI, CviBIII, PstI, BamHI and HincII.

<>

<1>Janulaitis, A., Vaitkevicius, D.P.
<2>A spectrophotometric procedure for the determination of activity of restriction.
<3>Anal. Biochem.
<4>116
<5>116-122
<6>1981
<7>A simple procedure basically applicable for the quantitative determination of
activity of all restriction endonucleases is described.  Native DNA immobilized
on cellulose is used as a substrate; after the treatment by restriction
endonucleases this DNA is released to the solution.  Changes of the optical
density of the solution containing solubilized DNA permit quantitative
determination of the restriction endonuclease activity.

<>

<1>Janulaitis, A., Vaitkevicius, D.P.
<2>New methodical approach to development of technology for production of restriction endonucleases.  Development of scheme for isolation of homogeneous preparation of restrictase MvaI.
<3>Biotekhnologiya
<4>1
<5>39-51
<6>1985
<7>Using restrictase MvaI isolated from Microccus varians RJL 19 cells a
possibility has been investigated for selection of purification schemes of
restrictases by means of study of binding of a specific enzyme with various
sorbents (DEAE-cellulose, phosphocellulose, heparinsepharose, blue sepharose,
Biorex 70 and SP-sephadex) under static conditions.  It has been illustrated
that the character of interaction of the restrictase under the static
conditions with various sorbents permits to predict its behavior under
conditions of the column chromatography and select the effective sorbents,
sequence of their application and conditions of conducting the column
chromatography.  A highly effective scheme for purification of MvaI restrictase
has been created which makes possible to produce electrophoretically
homogeneous preparation of a specific enzyme as a result of just two stages of
purification of the cell-free extract of M. varians RJL19 on phosphocellulose
and blue sepharose.  The prospects of application of the recommended methodical
approach for rapid development of a process for production on restrictases are
discussed.

<>

<1>Jaomanjaka, F., Ballestra, P., Dols-Lafargue, M., Le Marrec, C.
<2>Expanding the diversity of oenococcal bacteriophages: Insights into a novel group based on the integrase sequence.
<3>Int. J. Food Microbiol.
<4>166
<5>331-340
<6>2013
<7>Temperate bacteriophages are a contributor of the genetic diversity in the lactic
acid bacterium Oenococcus oeni. We used a classification scheme for oenococcal
prophages based on integrase gene polymorphism, to analyze a collection of
Oenococcus strains mostly isolated in the area of Bordeaux, which represented the
major lineages identified through MLST schemes in the species. Genome sequences
of oenococcal prophages were clustered into four integrase groups (A to D) which
were related to the chromosomal integration site. The prevalence of each group
was determined and we could show that members of the intB- and intC-prophage
groups were rare in our panel of strains. Our study focused on the so far
uncharacterized members of the intD-group. Various intD viruses could be easily
isolated from wine samples, while intD lysogens could be induced to produce
phages active against two permissive O. oeni isolates. These data support the
role of this prophage group in the biology of O. oeni. Global alignment of three
relevant intD-prophages revealed significant conservation and highlighted a
number of unique ORFs that may contribute to phage and lysogen fitness.

<>

<1>Jarjour, J., West-Foyle, H., Certo, M.T., Hubert, C.G., Doyle, L., Getz, M.M., Stoddard, B.L., Scharenberg, A.M.
<2>High-resolution profiling of homing endonuclease binding and catalytic specificity using yeast surface display.
<3>Nucleic Acids Res.
<4>37
<5>6871-6880
<6>2009
<7>Experimental analysis and manipulation of protein-DNA interactions pose unique biophysical
challenges arising from the structural and chemical
homogeneity of DNA polymers. We report the use of yeast surface display
for analytical and selection-based applications for the interaction
between a LAGLIDADG homing endonuclease and its DNA target. Quantitative
flow cytometry using oligonucleotide substrates facilitated a complete
profiling of specificity, both for DNA-binding and catalysis, with single
base pair resolution. These analyses revealed a comprehensive segregation
of binding specificity and affinity to one half of the pseudo-dimeric
interaction, while the entire interface contributed specificity at the
level of catalysis. A single round of targeted mutagenesis with tandem
affinity and catalytic selection steps provided mechanistic insights to
the origins of binding and catalytic specificity. These methods represent
a dynamic new approach for interrogating specificity in protein-DNA
interactions.

<>

<1>Jarman, S.N.
<2>Cleaver: software for identifying taxon specific restriction endonuclease recognition sites.
<3>Bioinformatics
<4>22
<5>2160-2161
<6>2006
<7>Cleaver is an application for identifying restriction endonuclease recognition sites that
occur in some taxa, but not in others.
Differences in DNA fragment restriction patterns among taxa are the
basis for many diagnostic assays for taxonomic identification and are
used in procedures for removing the DNA of some taxa from pools of DNA
from mixed sources. Cleaver analyses restriction digestion of groups of
orthologous DNA sequences simultaneously to allow identification of
differences in restriction pattern among the fragments derived from
different taxa.

<>

<1>Jarrell, K.F., Julseth, C., Pearson, B., Kuzio, J.
<2>Paucity of the Sau3AI recognition sequence (GATC) in the genome of Methanococcus voltae.
<3>Mol. Gen. Genet.
<4>208
<5>191-194
<6>1987
<7>High molecular weight genomic DNA isolated from the archaebacterium
Methanococcus voltae by alkaline-SDS lysis was not effectively digested with
the restriction enzyme Sau3AI, which recognizes the base sequence GATC.  M.
voltae DNA was also resistant to digestion by MboI and BamHI which recognize
sites containing the same GATC sequence.  Examination of a M. voltae genomic
library prepared in Escherichia coli JM83 with a pUC vector revealed that the
5-10 kb inserts were still resistant to Sau3AI digestion, indicating a likely
lack of the GATC sequence in M. voltae DNA.

<>

<1>Jarvik, T., Smillie, C., Groisman, E.A., Ochman, H.
<2>Short-term Signatures of Evolutionary Change in the Salmonella enterica serovar Typhimurium 14028 Genome.
<3>J. Bacteriol.
<4>192
<5>560-567
<6>2010
<7>Salmonella enterica serovar Typhimurium is a Gram-negative pathogen that causes
gastroenteritis in humans and a typhoid-like disease in mice and is often used as a model for
the disease promoted by the human-adapted S. enterica serovar Typhi. Despite its health
importance, the only S. Typhimurium strain for which the complete genomic sequence has been
determined is the avirulent LT2 strain, which is extensively used in genetic and physiologic
studies. Here, we report the complete genomic sequence of the S. Typhimurium strain 14028s, as
well as those of its progenitor and two additional derivatives. Comparison of these S.
Typhimurium genomes revealed differences in the patterns of sequence evolution and the
complete inventory of genetic alterations incurred in virulent and avirulent strains, as well
as the sequence changes accumulated during laboratory passage of pathogenic organisms.

<>

<1>Jarvis, A.W.
<2>Analysis of phage resistance mechanisms encoded by lactococcal plasmid pAJ2074.
<3>Can. J. Microbiol.
<4>39
<5>252-258
<6>1993
<7>Lactococcal plasmid pAJ2074 is a 74-kb plasmid that confers phage resistance at 30oC against
all lactococcal phages with prolate heads (referred to as prolate phage), and most small
lactococcal phages with isometric heads (referred to as small isometric phage) that have been
tested. The presence of pAJ2074 had no effect on phage adsorption or injection of phage DNA.
Replication of prolate phage c2 DNA could not be detected in bacterial cells containing the
plasmid up to 60 minutes after phage infection, whereas phage c2 DNA replication could be
demonstrated at 20 minutes in the control strain. With pAJ2074 present there was no detectable
growth of phage c2 and an 87% reduction in burst size for the small isometric phage sk1.
Infective centres were reduced in the presence of pAJ2074 by 99% for phage c2 and by 93% for
phage sk1. Plasmid pAJ2074 differed from pTR2030, in that the major effect of pAJ2074 was on
prolate phage c2, rather than on the small isometric phage sk1, and no restriction and
modification system could be detected. In addition, no DNA homology was detected between
pAJ2074 and pTRK67 (derived from pTR2030). A recombinant plasmid pAJ88 containing an 8.4kb
insert from pAJ2074 conferred an intermediate level of phage resistance. The DNA region that
encoded reduced phage sensitivity was further defined by the subcloning of a 5.6kb EcoRV
fragment that conferred resistance similar to pAJ88. The possibility of two phage-resistance
mechanisms being encoded by pAJ2074 is discussed.

<>

<1>Jarvis, A.W., Klaenhammer, T.R.
<2>Bacteriophage resistance conferred on lactic Streptococci by the conjugative plasmid pTR2030: Effects on small isometric-, large Isometric-,and prolate-headed Phages.
<3>Appl. Environ. Microbiol.
<4>51
<5>1272-1277
<6>1986
<7>A series of reactions between phages, sensitive hosts, and transconjugants where the
sensitivity of small isometric-, large isometric-, and prolate-headed phages to
pTR2030-induced phage resistance was evaluated in Streptococcus lactis and Streptococcus
cremoris strains. Phage-resistant transconjugants were constructed in the desired hosts by
conjugal transfer of lactose-fermenting ability (Lac+,pTR1040) and phage resistance (Hsp+
pTR2030) from S. lactis TEK1. S. lactis and S. cremoris transconjugants harboring pTR2030 were
resistant to all small isometric-headed phages examined. In contrast, prolate- and large
isometric-headed phages were either not inhibited in the pTR2030 transconjugants or exhibited
a reduction in plaque size without a reduction in the efficiency of plaquing. Small
isometric-headed phages subject to pTR2030 induced inhibition shared no significant DNA
homology with pTR2030, suggesting that phage immunity genes are not harbored on the plasmid or
responsible for resistance. The general effectiveness of pTR2030 against small
isometric-headed phages was highly significant since these are the phages which have been
isolated most commonly from dairy fermentation plants.

<>

<1>Jaubert, C., Danioux, C., Oberto, J., Cortez, D., Bize, A., Krupovic, M., She, Q., Forterre, P., Prangishvili, D., Sezonov, G.
<2>Genomics and genetics of Sulfolobus islandicus LAL14/1, a model hyperthermophilic archaeon.
<3>Open Biol.
<4>3
<5>130010
<6>2013
<7>The 2 465 177 bp genome of Sulfolobus islandicus LAL14/1, host of the model rudivirus SIRV2,
was sequenced. Exhaustive comparative genomic analysis of S. islandicus LAL14/1 and the nine
other completely sequenced S. islandicus strains isolated from Iceland, Russia and USA
revealed a highly syntenic common core genome of approximately 2 Mb and a long hyperplastic
region containing most of the strain-specific genes. In LAL14/1, the latter region is enriched
in insertion sequences, CRISPR (clustered regularly interspaced short palindromic repeats),
glycosyl transferase genes, toxin-antitoxin genes and MITE (miniature invertedrepeat
transposable elements). The tRNAgenes ofLAL14/1 are preferential targets for the integration
of mobile elements but clusters of atypical genes (CAG) are also integrated elsewhere in the
genome. LAL14/1 carries five CRISPR loci with 10 per
cent of spacers matching perfectly or imperfectly the genomes of archaeal viruses and plasmids
found in the Icelandic hot springs. Strikingly, theCRISPR_2 region of LAL14/1 carries an
unusually long 1.9 kb spacer interspersed between two repeat regions and displays a high
similarity to pING1-like conjugative plasmids. Finally, we have developed a genetic system for
S. islandicus LAL14/1 and created DpyrEF and DCRISPR_1 mutants using double cross-over and
pop-in/pop-out approaches, respectively. Thus,LAL14/1 is a promisingmodel to study virus-host
interactions and the CRISPR/Cas defence mechanism in Archaea.

<>

<1>Jauregui, R., Rodelas, B., Geffers, R., Boon, N., Pieper, D.H., Vilchez-Vargas, R.
<2>Draft Genome Sequence of the Naphthalene Degrader Herbaspirillum sp. Strain RV1423.
<3>Genome Announcements
<4>2
<5>e00188-14
<6>2014
<7>Herbaspirillum sp. strain RV1423 was isolated from a site contaminated with alkanes and
aromatic compounds and harbors the complete pathway for naphthalene
degradation. The new features found in RV1423 increase considerably the
versatility and the catabolic potential of a genus of bacteria previously
considered mainly to be diazotrophic endophytes to plants.

<>

<1>Javorsky, P., Pravdova, M., Vanat, I., Pristas, P., Styriak, I.
<2>Gene manipulation of Streptococcus bovis: progress and problems.
<3>Can. J. Anim. Sci.
<4>75
<5>654-655
<6>1995
<7>In general, realization of the goals to control the digestive processes of
ruminants via genetic manipulation of rumen bacteria, is limited mainly by little information
about the genome of important rumen bacteria, development of suitable cloning system,
stability of cloned DNA in rumen bacteria and in the whole rumen ecosystem as well.  In
our cloning study, the Sau 3A fragments of chromosomal DNA of S. bovis AO 24/85 were
ligated into BamHI site of shuttle vector pMX 39, as a host was used B. subtilis amy E-.
The restriction analyses of the recombinant plasmid pJK 108 isolated from B. subtilis alpha-
amylase positive clone showed that alpha-amylase gene was located within a 2.8 kb fragment of
the S. bovis chromosomal DNA.  Looking for a suitable transformation system, we have
transformed S. bovis AO 24/85 with plasmid pNZ12 by electroporation with  transformation
efficiency 1.1 x 103 ug-1 DNA.  In our laboratory we tested more S. bovis  strains for their
restriction activities.  We have developed a very simple, rapid and effective  method for
testing of RE activity of rumen bacteria.  By this method we detected and then  isolated and
characterized the restriction endonuclease SbvI, an isoschizomer of HaeIII,  from rumen
amylolytic bacterium S. bovis III/1.  Enzyme SbvI recognizes the 4-bp  palindrome,
5'-GGCC-3' and cleaves DNA after the second G in the sequence, producing blunt ends.

<>

<1>Jay, E., Wu, R.
<2>Arthrobacter luteus restriction endonuclease recognition sequence and its cleavage map of SV40 DNA.
<3>Biochemistry
<4>15
<5>3612-3620
<6>1976
<7>The nucleotide sequence at the cleavage site of the restriction endonuclease
isolated from Arthrobacter luteus (Alu) has been determined.  The endonuclease
cleaves at the center of a palindromic tetranucleotide sequence to give
even-ended duplex DNA fragments phosphorylated at the 5'-end.  The endonuclease
cleaves SV40 form I DNA into 32 fragments.  The order and sizes of these
fragments have been determined to provide an Alu cleavage map of the SV40
genome.

<>

<1>Jayapal, K.P., Lian, W., Glod, F., Sherman, D.H., Hu, W.S.
<2>Comparative genomic hybridizations reveal absence of large Streptomyces coelicolor genomic islands in Streptomyces lividans.
<3>BMC Genomics
<4>8
<5>229
<6>2007
<7>BACKGROUND: The genomes of Streptomyces coelicolor and Streptomyces lividans bear a
considerable degree of synteny. While S. coelicolor is the
model streptomycete for studying antibiotic synthesis and differentiation,
S. lividans is almost exclusively considered as the preferred host, among
actinomycetes, for cloning and expression of exogenous DNA. We used whole
genome microarrays as a comparative genomics tool for identifying the
subtle differences between these two chromosomes. RESULTS: We identified
five large S. coelicolor genomic islands (larger than 25 kb) and 18
smaller islets absent in S. lividans chromosome. Many of these regions
show anomalous GC bias and codon usage patterns. Six of them are in close
vicinity of tRNA genes while nine are flanked with near perfect repeat
sequences indicating that these are probable recent evolutionary
acquisitions into S. coelicolor. Embedded within these segments are at
least four DNA methylases and two probable methyl-sensing restriction
endonucleases. Comparison with S. coelicolor transcriptome and proteome
data revealed that some of the missing genes are active during the course
of growth and differentiation in S. coelicolor. In particular, a pair of
methylmalonyl CoA mutase (mcm) genes involved in polyketide precursor
biosynthesis, an acyl-CoA dehydrogenase implicated in timing of
actinorhodin synthesis and bldB, a developmentally significant regulator
whose mutation causes complete abrogation of antibiotic synthesis belong
to this category. CONCLUSION: Our findings provide tangible hints for
elucidating the genetic basis of important phenotypic differences between
these two streptomycetes. Importantly, absence of certain genes in S.
lividans identified here could potentially explain the relative ease of
DNA transformations and the conditional lack of actinorhodin synthesis in
S. lividans.

<>

<1>Jayaraman, K.S.
<2>Restriction venture.
<3>Nature
<4>341
<5>272
<6>1989
<7>India has launched its first venture capital company to make restriction enzymes.  Bangalore
Genei Private Ltd has been set up by Dr. P.S. Babu, a theoretical physicist-turned-molecular
biologist, and Dr. K. Prasad, an immunologist, both of whom have been associated with the
Indian Institute of Science, Bangalore.  A large part of the funding for the company comes
from the Technology Development and Information Company of India.  The new company will obtain
the know-how for the manufacture of the enzymes from Astra Research Centre, Bangalore, which
carries out research in medical biotechnology and is developing diagnostic kits for a variety
of tropical diseases.  The Centre for Genetic Engineering will also assist the new company.
Restriction enzymes are currently being imported by individual laboratories and in bulk
quantities by the CSIR Centre for Biochemicals, New Delhi.  The new company will initially
make the more important and commonly used restriction enzymes.

<>

<1>Jayaraman, R.
<2>Phase variation and adaptation in bacteria: A 'Red Queen's Race'.
<3>Curr. Sci.
<4>100
<5>1163-1171
<6>2011
<7>In nature, bacteria are constantly exposed to many stressful conditions of life. This is
particularly true of pathogens. Survival and
adaptation under stressful conditions demand multiple strategies,
genetic as well as phenotypic. Bacteria have many, pre-programmed,
phenotypic stress response systems which can handle a limited number of
stresses. Genetically, heritable as well as transient hypermutability
mechanisms have been found to facilitate bacterial adaptation to varied
and unpredictable stresses; these processes are not reviewed here.
Instead, this article will focus on processes which do not increase
global mutation rates but cause localized hypermutability in specific
loci called contingency genes which have been identified particularly
in pathogenic bacteria. These processes are collectively called phase
and antigenic variations. Most of the contingency genes are involved in
the synthesis or modification of surface-associated structures and
enzymes. Phase variation in these genes involves high frequency,
reversible, switching of their expression (on to off and off to on).
The mechanisms of this switching are reviewed. However, some phase
variable genes are not involved in the synthesis or modification of
surface structures but are components of type I and type III
restriction-modification (RM) systems. The on/off switching of these
genes (type III RM genes) leads to regulation of expression of many
unlinked genes, impacting several properties of cells. This novel type
of control of multiple gene expression by phase variation has been
named 'phasevarion'. The adaptive advantages of phase variation in
contingency genes and phasevarions in the evasion of host immunity,
virulence, niche adaptation and other phenomena are reviewed with some
illustrative examples. Phase variation and bacterial adaptation have
been likened to the 'Red Queen's Race' in Lewis Carrol's classic
Through the Looking Glass.

<>

<1>Jayashree, S., Pooja, S., Pushpanathan, M., Vishnu, U., Sankarasubramanian, J., Rajendhran, J., Gunasekaran, P.
<2>Genome Sequence of Lactobacillus fermentum Strain MTCC 8711, a Probiotic Bacterium Isolated from Yogurt.
<3>Genome Announcements
<4>1
<5>e00770-13
<6>2013
<7>Lactobacillus fermentum strain MTCC 8711 is a lactic acid bacterium isolated from yogurt.
Here, we describe the draft genome sequence and annotation of this
strain. The 2,566,297-bp-long genome consisted of a single chromosome and seven
plasmids. The genome contains 2,609 protein-coding and 74 RNA genes.

<>

<1>Jean, V., Guillot, M., Courjal, F., Savoye, C.
<2>Detection of staphylococcus aureus and identification of methicillin-resistant staphylococcus aureus.
<3>International Patent Office
<4>WO 2008140612 A
<5>
<6>2008
<7>Aspects of the present invention relate to methods and compositions for the detection and/or
quantification of S. aureus from a sample, as well as methods and compositions useful for the
detection and/or quantification of S. aureus and MRSA in a single assay.  Embodiments include
nucleic acids that hybridize to S. aureus-specific nuc sequences and MREJ sequences.

<>

<1>Jeddeloh, J.A., Lakey, N.D.
<2>Differential enzymatic fragmentation for detection of methylation at a genetic locus using methylation-sensitive and methylation-dependent  restriction enzymes.
<3>International Patent Office
<4>WO 200542704 A
<5>87
<6>2005
<7>The present invention provides methods for detecting the presence of methylation at a locus
within a population of nucleic acids.

<>

<1>Jeddeloh, J.A., Lakey, N.D.
<2>Quantifying average methylation density in a locus of genomic DNA, useful in molecular cancer research, by contacting genomic DNA with a methylation-dependent or methylation-sensitive restriction enzyme - for cancer therapy and diagnosis.
<3>International Patent Office
<4>WO 200540399
<5>
<6>2005
<7>NOVELTY - Quantifying average methylation density in a locus of genomic DNA comprises
contacting genomic DNA with a methylation-dependent restriction enzyme or a
methylation-sensitive restriction enzyme for at least some copies of potential restriction
enzyme cleavage sites in the locus to remain uncleaved. DETAILED DESCRIPTION - Quantifying
average methylation density in a locus of genomic DNA comprises contacting genomic DNA with a
methylation-dependent restriction enzyme or a methylation-sensitive restriction enzyme for at
least some copies of potential restriction
enzyme cleavage sites in the locus to remain uncleaved, quantifying
intact copies of the locus, and comparing the quantity of amplified
product to a control value representing the quantity of methylation of
control DNA, thereby quantifying the average methylation density in the
locus compared to the methylation density of the control DNA.
INDEPENDENT CLAIMS are also included for the following: (1) a method of
calculating the relative methylation density for a target locus in a
DNA sample; and (2) a kit for quantifying the average methylation
density in a locus of genomic DNA, the kit comprising a
methylation-dependent restriction enzyme or a methylation sensitive
restriction enzyme; a control DNA molecule comprising a pre-determined
number of methylated nucleotides; and control oligonucleotide primers
that hybridize to the control DNA molecule. BIOTECHNOLOGY - Preferred
Method: In the method above, the quantifying step comprises a
quantitative amplification. The quantity of the amplified product is
compared to a standard curve. The amplifying step comprises hybridizing
two oligonucleotide primers to DNA flanking the locus to produce an
amplification product corresponding to the uncleaved locus of genomic
DNA between the primers. The control value represents the quantity of
an amplification product of a DNA sample having a known or predicted
number of methylated nucleotides. The restriction enzyme is a
methylation-sensitive restriction enzyme selected from Aat II, Aci I, Acl I, Age I, Alu I, Asc
I, Ase I, AsiS I, Bbe I, BsaA I, BsaR I, BsiE I,
BsiW I, BsrF I, BssH II, BssK I, BstB I, BstN I, BstU I, Cla I, Eae I,
Eag I, Fau I, Fse I, Hha I, HinPl I, HinC II, Hpa II, Hpy99 I, HpyCH4
IV, Kas I, MIu I, MapAl I, Msp I, Nae I, Nar I, Not I, PmI I, Pst I,
Pvu I, Rsr II, Sac II, Sap I, Sau3A I, SfI I, Sfo I, SgrA I, Sma I,
SnaB I, Tsc I, Xma I, and Zra I. The restriction enzyme is a
methylation-dependent restriction enzyme or methyl-cytosine-dependent
restriction enzyme. The restriction enzyme is McrBC, McrA, or Mrr. It
may be a methyl-adenosine-dependent restriction enzyme, i.e., DpnI. The
methylation-sensitive or methylation dependent restriction enzyme is
contacted to the portion for only a partial digest of the portion. The
method comprises separating the genomic DNA into at least two equal
portions, contacting one portion with a methylation-sensitive or
methylation dependent restriction enzyme and contacting a second
portion with the isoschizomeric partner of the restriction enzyme,
amplifying the locus of genomic DNA in each portion in a step
comprising hybridizing two oligonucleotide primers to DNA flanking the
locus, quantifying the amplification product, and comparing the
quantity of amplified products from the two portions. The method
further comprises contacting the genomic DNA with an agent that
modifies unmethylated cytosine before the amplifying step, and at least
one of the two oligonucleotide primers distinguishes between modified
unmethylated and methylated DNA in the genomic DNA. The method may
further comprise contacting the DNA with at least one
methylation-sensitive restriction enzyme or methylation-sensitive
restriction enzyme before the genomic DNA is contacted with an agent
that modifies unmethylated cytosine. The genomic DNA is contacted with
a mixture of at least two different methylation-sensitive or
methylation-dependent restriction enzymes.

<>

<1>Jeddeloh, J.A., Stokes, T.L., Richards, E.J.
<2>Maintenance of genomic methylation requires a SW12/SNF2-like protein.
<3>Nat. Genet.
<4>22
<5>94-97
<6>1999
<7>Altering cytosine methylation by genetic means leads to a variety of developmental defects in
mice, plants and fungi. Deregulation of cytosine methylation also has a role in human
carcinogenesis. In some cases, these defects have been tied to the inheritance of epigenetic
alterations (such as chromatin imprints and DNA methylation patterns) that do not involve
changes in DNA sequence.  Using a forward genetic screen, we identified a gene (DDM1, decrease
in DNA methylation) from the flowering plant Arabidopsis thaliana required to maintain normal
cytosine methylation patterns. Additional ddm1 alleles (som4, 5, 6, 7, 8) were isolated in a
selection for mutations that relieved transgene silencing (E.J.R., unpublished data). Loss of
DDM1 function causes a 70% reduction of genomic cytosine methylation, with most of the
immediate hypomethylation occurring in repeated sequences. In contrast, many low-copy
sequences initially retain their methylation in ddm1 homozygotes, but lose methylation over
time as the mutants are propagated through multiple generations by self-pollination. The
progressive effect of ddm1 mutations on low-copy sequence methylation suggests that ddm1
mutations compromise the efficiency of methylation of newly incorporated cytosines after DNA
replication. In parallel with the slow decay of methylation during inbreeding, ddm1 mutants
accumulate heritable alterations (mutations or stable epialleles) at dispersed sites in the
genome that lead to morphological abnormalities. Here we report that DDM1 encodes a
SWI2/SNF2-like protein, implicating chromatin remodelling as an important process for
maintenance of DNA methylation and genome integrity.

<>

<1>Jeffery, C.J.
<2>Molecular mechanisms for multitasking: recent crystal structures of moonlighting proteins.
<3>Curr. Opin. Struct. Biol.
<4>14
<5>663-668
<6>2004
<7>Recently determined X-ray crystal structures of moonlighting proteins are helping to elucidate
how a protein can evolve two different
functions and, in some cases, switch between its two functions in
response to cellular conditions. X-ray crystal structures of the I-Anil
homing endonuclease/maturase and the PutA proline
dehydrogenase/transcription factor have provided evidence that these
proteins utilize separate protein surfaces for their multiple
functions. Also, the structure of the DegP (HtrA) protease/chaperone
has revealed information about the mechanism of its chaperone activity
and suggests how the protein regulates its protease activity. Comparing
the structure of eta-crystallin/retinal dehydrogenase with structures
of its single-function enzyme homologs provides clues to changes in the
protein structure that may have improved its ability to serve as a
crystallin, but at the same time may have adversely affected its
catalytic activity.

<>

<1>Jeganathan, L.P., Prakash, L., Sivakumar, N., Antony, A., Alqarawi, S., Prajna, L., Devarajan, B., Mohankumar, V.
<2>Draft Genome Sequence of an Invasive Multidrug-Resistant Strain, Pseudomonas aeruginosa BK1, Isolated from a Keratitis Patient.
<3>Genome Announcements
<4>2
<5>e00153-14
<6>2014
<7>Pseudomonas aeruginosa infections are difficult to treat due to the presence of a multitude of
virulence factors and antibiotic resistance. Here, we report the
draft genome sequence of P. aeruginosa BK1, an invasive and multidrug-resistant
strain, isolated from a bacterial keratitis patient in southern India.

<>

<1>Jeltsch, A.
<2>Circular permutations in the molecular evolution of DNA methyltransferase.
<3>J. Mol. Evol.
<4>49
<5>161-164
<6>1999
<7>Circular permutations of genes during molecular evolution often are regarded as elusive,
although a simple model can explain these rearrangements. The model assumes that first a gene
duplication of the precursor gene occurs in such a way that both genes become fused in frame,
leading to a tandem protein. After generation of a new start codon within the 5' part of the
tandem gene and a stop at an equivalent position in the 3' part of the gene, a protein is
encoded that represents a perfect circular permutation of the precursor gene product. The
model is illustrated here by the molecular evolution of adenine-N6 DNA methyltransferases.
Beta- and gamma-type enzymes of this family can be interconverted by a single circular
permutation event. Interestingly, tandem proteins, proposed as evolutionary intermediates
during circular permutation, can be directly observed in the case of adenine
methyltransferases, because some enzymes belonging to type IIS, like the FokI
methyltransferase, are built up by two fused enzymes, both of which are active independently
of each other. The mechanism for circular permutation illustrated here is very easy and
applicable to every protein. Thus, circular permutation can be regarded as a normal process in
molecular evolution and a changed order of conserved amino acid motifs should not be
interpreted to argue against divergent evolution.

<>

<1>Jeltsch, A.
<2>The cytosine N4-methyltransferase M.PvuII also modifies adenine residues.
<3>Biol. Chem.
<4>382
<5>707-710
<6>2001
<7>Methylation of DNA occurs at the C5 and N4 positions of cytosine and N6 of adenine. The
chemistry of methylation is similar among methyltransferases specific for cytosine-N4 and
adenine-N6. Moreover these enzymes have similar structures and active sites. Previously it has
been demonstrated that the DNA-(adenine-N6)-methyltransferases M.EcoRV, M.EcoRI, E. coli dam
and both domains of M.FokI also modify cytosine residues at the N4 position [Jeltsch et al.,
J. Biol. Chem. 274 (1999), 19538-19544]. Here we show that the cytosine-N4 methyltransferase
M.PvuII, which modifies the second cytosine in CAGCTG sequences, also methylates adenine
residues in CAGATG/CAGCTG substrates in which the target cytosine is replaced by adenine in
one strand of the recognition sequence. Therefore, adenine-N6 and cytosine-N4
methyltransferases have overlapping target base specificities. These results demonstrate that
the target base recognition by N-specific DNA methyltransferases is relaxed in many cases.
Furthermore, it shows that the catalytic mechanisms of adenine-N6 and cytosine-N4
methyltransferases are very similar.

<>

<1>Jeltsch, A.
<2>Beyond Watson and Crick: DNA methylation and molecular enzymology of DNA methyltransferases.
<3>Chembiochem
<4>3
<5>274-293
<6>2002
<7>DNA methyltransferases catalyze the transfer of a methyl group from S-adenosyl-L-methionine to
cytosine or adenine bases in DNA. These
enzymes challenge the Watson/Crick dogma in two instances: 1) They
attach inheritable information to the DNA that is not encoded in the
nucleotide sequence. This so-called epigenetic information has many
important biological functions. In prokaryotes, DNA methylation is used
to coordinate DNA replication and the cell cycle, to direct
postreplicative mismatch repair, and to distinguish self and nonself
DNA. In eukaryotes, DNA methylation contributes to the control of gene
expression, the protection of the genome against selfish DNA,
maintenance of genome integrity, parental imprinting, X-chromosome
inactivation in mammals, and regulation of development, 2) The
enzymatic mechanism of DNA methyltransferases is unusual, because these
enzymes flip their target base out of the DNA helix and, thereby,
locally disrupt the B-DNA helix. This review describes the biological
functions of DNA methylation in bacteria, fungi, plants, and mammals.
In addition, the structures and mechanisms of the DNA
methyltransferases, which enable them to specifically recognize their
DNA targets and to induce such large conformational changes of the DNA,
are discussed.

<>

<1>Jeltsch, A.
<2>Maintenance of species identity and controlling speciation of bacteria: a new function for restriction/modification systems?
<3>Gene
<4>317
<5>13-16
<6>2003
<7>Bacteria frequently exchange DNA among each other by horizontal gene transfer. However,
maintenance of species identity and in particular
speciation requires a certain barrier against an unregulated uptake of
foreign DNA. Here it is suggested that formation of such a barrier is one
important biological function of restriction/modification systems, in
addition to the classical function of protection of bacteria against
bacteriophage infection. This model explains the extreme variability and
wide distribution of restriction/modification systems among prokaryotes,
the prevalence of RM-systems in pathogenic bacteria and the existence of
several RM-systems in single bacterial strains.

<>

<1>Jeltsch, A.
<2>Molecular enzymology of mammalian DNA methyltransferases.
<3>Curr. Top. Microbiol. Immunol., Springer-Verlag, Doerfler, W., Bohm, P., Berlin, Germany
<4>301
<5>203-225
<6>2006
<7>DNA methylation is an essential modification of DNA in mammals that is involved in gene
regulation, development, genome defence and disease.
In mammals 3 families of DNA methyltransferases (MTases) comprising (so
far) 4 members have been found: Dnmt1, Dnmt2, Dnmt3A and Dnmt3B. In
addition, Dnmt3L has been identified as a stimulator of the Dnmt3A and
Dnmt3B enzymes. In this review the enzymology of the mammalian DNA
MTases is described, starting with a depiction of the catalytic
mechanism that involves covalent catalysis and base flipping.
Subsequently, important mechanistic features of the mammalian enzyme
are discussed including the specificity of Dnmt1 for hemimethylated
target sites, the target sequence specificity of Dnmt3A, Dnmt3B and
Dnmt2 and the flanking sequence preferences of Dnmt3A and Dnmt3B. In
addition, the processivity of the methylation reaction by Dnmt1, Dnmt3A
and Dnmt3B is reviewed. Finally, the control of the catalytic activity
of mammalian MTases is described that includes the regulation of the
activity of Dnmt1 by its N-terminal domain and the interaction of
Dnmt3A and Dnmt3B with Dnmt3L. The allosteric activation of Dnmt1 for
methylation at unmodified sites is described. Wherever possible,
correlations between the biochemical properties of the enzymes and
their physiological functions in the cell are indicated.

<>

<1>Jeltsch, A., Alves, J., Maass, G., Pingoud, A.
<2>On the catalytic mechanism of EcoRI and EcoRV.  A detailed proposal based on biochemical results, structural data and molecular modelling.
<3>FEBS Lett.
<4>304
<5>4-8
<6>1992
<7>EcoRI and EcoRV have a very similar active site, as is apparent from a comparison of the
structures of their respective protein-DNA complexes.  Based on structural and mechanistic
data, as well as detailed molecular modelling presented here, a mechanism for the DNA cleavage
by these enzymes is suggested in which the attacking water molecule is activated by the
phosphate group 3' to the scissile phosphodiester bond, and in which the leaving group is
protonated by a water molecule associated with the essential cofactor, Mg2+.  The mechanism
proposed may also apply to other nucleases.

<>

<1>Jeltsch, A., Alves, J., Oelgeschlager, T., Wolfes, H., Maass, G., Pingoud, A.
<2>Mutational analysis of the function of Gln115 in the EcoRI restriction endonuclease, a critical amino acid for recognition of the inner thymidine residue in the sequence -GAATTC-and for coupling specific DNA binding to catalysis.
<3>J. Mol. Biol.
<4>229
<5>221-234
<6>1993
<7>The Gln115 residue of the EcoRI restriction endonuclease has been proposed to form a
hydrophobic contact to the methyl group of the inner thymidine of the EcoRI recognition
sequence -GAATTC- and to be involved in intramolecular hydrogen bonds to the mainchain at
positions 140 and 143 as well as to the side-chain of Asn173. We have exchanged Gln115 for Ala
and Glu by site-directed mutagenesis and analysed the purified mutant protein (Q115A and
Q115E) biochemically and physico-chemically. Q115A and Q115E have the same secondary structure
composition as wild-type EcoRI but are less stable towards thermal denaturation than the
wild-type enzyme. In contrast to wild-type EcoRI the mutant proteins show a biphasic
denaturation profile under alkaline pH, presumably because the amino acid exchange labilizes
one part of the molecule, which unfolds before the rest of the protein is denatured. Q115A is
catalytically inactive under normal buffer conditions, in part due to a diminished affinity
towards DNA. At low ionic strength and alkaline pH, as well as in the presence of Mn2+, i.e.
under conditions where wild-type EcoRI shows a relaxed specificity, Q115A is active, however
not as much as wild-type EcoRI. Under these conditions it cleaves the canonical sequence
-GAATTC- with the same kcat/km value as the sequence -GAAUTC-, which differs from the former
sequence by a single methyl group, while wild-type EcoRI shows a tenfold lower kcat/Km for
cleavage of -GAAUTC- than for -GAATTC-. Binding experiments, carried out in the absence of
Mg2+, demonstrate that Q115A has a similar affinity toward -GAATTC- as to -GAAUTC-, while
wild-type EcoRI binds to -GAATTC- with a tenfold preference over -GAAUTC-. On the basis of
these thermodynamic and kinetic results it can be concluded that the hydrophobic contact
between the gamma-methylene group of Gln115 and the methyl group of the inner thymidine
contributes about 3 kJ/mol (0.7 kcal/mol) to the energy of interaction, both in the ground and
the transition state, Q115E is catalytically inactive under normal buffer conditions, but
becomes active at low ionic strength or in the presence of Mn2+. Different from Q15A, Q115E is
inactive at alkaline pH and its DNA binding affinity is highest at acidic pH. The dependence
of its DNA cleavage activity on pH, which is governed by a pKa of 7.35, can be attributed to
the protonation of the newly introduced glutamic acid residue, if it is assumed that the
carboxyl group is located in a non-polar environment and/or involved in a hydrogen bond as the
donor. Taken together, these results demonstrate that Gln115, as suggested on the basis of the
revised EcoRI-DNA co-crystal structure, interacts with the inner thymidine of the EcoRI
recognition sequence and has a structure stabilizing role. In addition, our results suggest
that Gln115 is crucial for coupling specific DNA binding to catalysis under normal buffer
conditions and we suggest that this coupling is achieved because direct and indirect
involvement of Gln115 in base recognition induces local conformational alterations of the
C-terminal region of beta-strand beta3, which contains Lys113 and Glu111 that are constituents
of the catalytic centre of EcoRI. Activation of the catalytic centre, therefore, could be
triggered by Gln115.

<>

<1>Jeltsch, A., Alves, J., Urbanke, C., Maass, G., Eickstein, H., Lianshan, Z., Bayer, E., Pingoud, A.
<2>A dodecapeptide comprising the extended chain-alpha-4 region of the restriction endonuclease EcoRI specifically binds to the EcoRI recognition site.
<3>J. Biol. Chem.
<4>270
<5>5122-5129
<6>1995
<7>The restriction endonuclease EcoRI binds and cleaves DNA containing GAATTC sequences with high
specificity. According to the crystal structure, most of the specific contacts of the enzyme
to the DNA are formed by the extended chain region and the first turn of alpha-helix alpha-4
(amino acids 137-145). Here, we demonstrate that a dodecapeptide (WDGMAAGNAIER), which is
identical in the underlined parts (M-R) of its sequence to EcoRI amino acids 137-145,
specifically binds to GAATTC sequences. The peptide inhibits DNA cleavage by EcoRI but not by
BamHI, BclI, EcoRV, HindIII, PacI, and XbaI. DNA cleavage by XbaI is slowed down at sites that
partially overlap with EcoRI sites. The peptide inhibits cleavage of GAATTC sites by ApoI,
which recognizes the sequence RAATTY. It interferes with DNA methylation by the EcoRI
methyltransferase but not by the BamHI methyltransferase. It competes with EcoRI for DNA
binding. Based on these results, the DNA binding constant of the peptide to GAATTC sequences
was calculated to be 3 x 10/4 M-1. DNA binding is not temperature-dependent, suggesting that
binding of the peptide is entropy-driven. As the peptide does not show any nonspecific binding
to DNA, its DNA binding specificity is similar to that of EcoRI, in spite of the fact that the
affinity is much smaller. These results suggest that contacts to the phosphate groups in EcoRI
mainly provide binding affinity, whereas the specificity of EcoRI is based to a large extent
on sequence-specific base contacts.

<>

<1>Jeltsch, A., Alves, J., Wolfes, H., Maass, G., Pingoud, A.
<2>Substrate-assisted catalysis in the cleavage of DNA by the EcoRI and EcoRV restriction enzymes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>8499-8503
<6>1993
<7>The crystal structure analyses of the EcoRI-DNA and EcoRV-DNA complexes do not provide clear
suggestions as to which amino acid residues are responsible for the activation of water to
carry out the DNA cleavage. Based on molecular modeling, we have proposed recently that the
attacking water molecule is activated by the negatively charged pro-Rp phosphoryl oxygen of
the phosphate group 3' to the scissile phosphodiester bond. We now present experimental
evidence to support this proposal. (i) Oligodeoxynucleotide substrates lacking this phosphate
group in one strand are cleaved only in the other strand. (ii) Oligodeoxynucleotide substrates
carrying an H-phosphonate substitution at this position in both strands and, therefore,
lacking a negatively charged oxygen at this position are cleaved at least four orders of
magnitude more slowly than the unmodified substrate. These results are supported by other
modification studies: oligodeoxynucleotide substrates with a phosphorothioate substitution at
this position in both strands are cleaved only if the negatively charged sulfur is in the Rp
configuration as shown for EcoRI [Koziolkiewics, M. and Stec, W.J. (1992) Biochemistry 31,
9460-9466] and EcoRV (B.A. Connolly, personal communication). As the phosphate residue 3' to
the scissile phosphodiester bond is not needed for strong DNA binding by both enzymes, these
findings strongly suggest that this phosphate group plays an active role during catalysis.
This proposal furthermore, give a straightforward explanation of why in the EcoRI-DNA and
EcoRV-DNA complexes the DNA is distorted differently, but in each case the 3' phosphate group
closely approaches the phosphate group that is attacked. Finally, an alternative mechanism for
DNA cleavage involving two metal ions is unlikely in the light of our findings that both EcoRI
and EcoRV and need only one Mg2+ per active site for cleavage.

<>

<1>Jeltsch, A., Alves, J., Wolfes, H., Maass, G., Pingoud, A.
<2>Pausing of the restriction endonuclease EcoRI during linear diffusion on DNA.
<3>Biochemistry
<4>33
<5>10215-10219
<6>1994
<7>Linear diffusion is a mechanism to accelerate association rates beyond their three-dimensional
diffusional limit. It is employed by the restriction endonuclease EcoRI as well as many other
proteins interacting with specific DNA sequences to locate their target sites on the
macromolecular substrate. In order to investigate biochemical and biophysical details of the
linear diffusion process, we have developed a competitive cleavage assay which allows us to
assess with great accuracy the influence of sequence, sequence context, and other structural
features on the linear diffusion of EcoRI on DNA. We show here that linear diffusion is not a
hopping but a sliding movement in which EcoRI follows the helical pitch of the DNA, because it
does not "overlook" any cleavage site. Linear diffusion is slowed when EcoRI encounters sites
on the DNA which resemble its recognition site ("star" sites). Pauses of up to 20 s are
induced, depending on sequence and orientation of the star site. These data suggest that EcoRI
can bind to DNA in two binding modes: one tight, specific, and immobile, leading to DNA
cleavage, and another one loose and nonspecific, allowing for linear diffusion. Depending on
the similarity between the recognition sequence and the DNA sequence being encountered by
EcoRI, there will be a continuous transition between these binding modes. Other proteins bound
to the DNA and irregular DNA structures such as bent DNA or a triple helix constitute a
barrier than cannot easily be passed by EcoRI.

<>

<1>Jeltsch, A., Christ, F., Fatemi, M., Roth, M.
<2>On the substrate specificity of DNA methyltransferases.
<3>J. Biol. Chem.
<4>274
<5>19538-19544
<6>1999
<7>Methylation of DNA is important in many organisms and essential in mammals. Nucleobases can be
methylated at the adenine-N6, cytosine-N4, or cytosine-C5 atoms by specific DNA
methyltransferases. We show here that the M.EcoRV, M.EcoRI, and Escherichia coli dam
methyltransferases as well as the N- and C-terminal domains of the M. FokI enzyme, which were
formerly all classified as adenine-N6 DNA methyltransferases, also methylate cytosine residues
at position N4. Kinetic analyses demonstrate that the rate of methylation of cytosine residues
by M.EcoRV and the M.FokI enzymes is reduced by only 1-2 orders of magnitude in relation to
methylation of adenines. This result shows that although these enzymes methylate DNA in a
sequence specific manner, they have a low substrate specificity with respect to the target
base. This unexpected finding has implications on the mechanism of adenine-N6 DNA
methyltransferases. Sequence comparisons suggest that adenine-N6 and cytosine-N4
methyltransferases have changed their reaction specificity at least twice during evolution, a
model that becomes much more likely given the partial functional overlap of both enzyme types.
In contrast, methylation of adenine residues by the cytosine-N4 methyltransferase M.BamHI was
not detectable. On the basis of our results, we suggest that adenine-N6 and cytosine-N4
methyltransferases should be grouped into one enzyme family.

<>

<1>Jeltsch, A., Friedrich, T., Roth, M.
<2>Kinetics of methylation and binding of DNA by the EcoRV adenine-N6 methyltransferase.
<3>J. Mol. Biol.
<4>275
<5>747-758
<6>1998
<7>The EcoRV DNA methyltransferase specifically methylates the first adenine within its
recognition sequence GATATC.  Methylation rates of DNA by this enzyme are strongly influenced
by the length of oligonucleotide substrates employed.  In substrates >20 bp compared to a
12mer substrate, the kcat/Km increases 100-fold, although the enzyme does not contact more
than 12 base-pairs on the DNA.  Single-turnover rates are higher than kcat values.  M.EcoRV
binding to DNA is fast but dissociation from the DNA is slow, demonstrating that the
multiple-turnover rate is limited by the rate of product release.  The kinetics of DNA binding
by M.EcoRV are not in accordance with the thermodynamic binding constant, suggesting that the
M.EcoRV-DNA complex is involved in a slow conformational change.  The salt dependence of DNA
binding is different for non-specific substrates (d ln(KAss)/d ln(cNaCl) = -2, indicative of
electrostatic interactions) and specific substrates (d ln(KAss)/d ln(cNaCl)=+1, indicative of
hydrophobic interactions).  This result demonstrates that the M.EcoRV-DNA complex has a
different conformation in both binding modes.  M.EcoRV does not discriminate between
hemimethylated and unmethylated substrates.  Using the 20mer we have analyzed the temperature
and pH dependence of the single-turnover rate constant of M.EcoRV-DNA methylation by M.EcoRV
which has an activation energy of 40 kJ/mol and its rate increases with increasing pH.  The pH
dependence reveals the presence of an ionizable residue with a pKa of 7.9, which must be
unprotonated for catalysis.  The rates of DNA methylation remain unchanged if an abasic site
is introduced instead of the thymidine residue that is base-paired to the target adenine,
demonstrating that flipping out the target adenine cannot contribute to the rate-limiting step
of the enzymatic reaction.

<>

<1>Jeltsch, A., Fritz, A., Alves, J., Wolfes, H., Pingoud, A.
<2>A fast and accurate enzyme-linked immunosorbent assay for the determination of the DNA cleavage activity of restriction endonucleases.
<3>Anal. Biochem.
<4>213
<5>234-240
<6>1993
<7>We have developed an assay procedure to monitor the cleavage of DNA substrates by restriction
endonucleases. This procedures uses DNA substrates that are labeled with biotin on one 5' end
and with an antigenic group, e.g., flourescein or digoxigenin, on the other 5' end. After
incubation with the restriction enzyme, the reaction is stopped with EDTA and an aliquot is
pipetted into the well of an avidin-coated microtiter plate. This imobilizes the unreacted
substrate and the biotinylated cleavage product, whereas the other cleavage product labeled
with the antigenic group is subsequently washed off. The unreacted substrate is detected by an
enzyme-linked immunosorbent assay with an appropriate enzyme-linked anitbody. To test our
assay we have measured the steady-state rate constants for cleavage of DNA by EcoRI yielding a
kcat of 8.6 min and a Km of 150 nM, which are close to values measured with other assays. The
advantage of this assay is that it is not only fast and accurate, but also very sensitive. It
allows for many samples to be analyzed in parallel and lends itself to automation.
Furthermore, this assay can be designed as a competitive assay, when two substrates carrying
different antigenic groups are used. The usefulness of such a competitive assay is
demonstrated by determining the influence of sequence context on the rate of DNA cleavage by
EcoRI.

<>

<1>Jeltsch, A., Gowhar, H.
<2>Molecular enzymology of the DNA-(adenine-N6)-methyltransferase M.EcoRV: Kinetic mechanism, kinetics of DNA binding and bending, and linear diffusion.
<3>Biochem. Soc. Trans.
<4>28
<5>A312
<6>2000
<7>Methylation of DNA is important in many organisms and essential in mammals.  Nucleobases can
be methylated at the adenine-N6, cytosine-N4 or cytosine-C5 atoms by specific DNA
methyltransferases.  The M.EcoRV adenine-N6 DNA methyltransferase specifically transfers a
methyl group from AdoMet to the first adenine within GATATC sequences.  We show here that
substrate binding to the enzyme follows an ordered bi-bi mechanism with AdoMet binding before
DNA.  After DNA binding a non-specific enzyme-DNA complex is formed that slides along the DNA
for more than 1800 bps by linear diffusion.  Upon specific complex formation the DNA is bent
in a fast process with rate constants >10s^-1.  The cofactor of the methylation reaction,
S-adenosylmethionine, but not the product of the reaction, S-adenosylhomocysteine, promotes
specific complex formation.  In the presence of cofactor, the specific complex can change into
a second conformation, in which the target sequence is more tightly contacted by the enzyme.
M.EcoRV exists in a slow equilibrium between an open and a closed conformation.  Cofactor
binding to the apoenzyme shifts the conformational equilibrium to the open conformation
thereby promoting DNA binding.  Formation of a closed enzyme-DNA complex is a slow process
(rate constant < 0.7 min^-1), that limits the rate of DNA methylation under single turnover
conditions.  Product release requires opening of the closed complex which is very slow (rate
constant < 0.05-0.1 min^-1) and limits the multiple turnover rate constant.  Slow DNA release
from the closed complex also explains the high efficiency of linear diffusion of M.EcoRV on
DNA.

<>

<1>Jeltsch, A., Gumport, R.I.
<2>DNA Methyltransferases, Bacterial.
<3>Encyclopedia of Biological Chemistry, Academic Press, Lennarz, W.J., Lane, D., 
<4>
<5>644-651
<6>2004
<7>DNA methylation has a number of important roles in bacteria including the control of gene
expression, DNA replication, and the cell cycle.  In addition, it is involved in mismatch
repair and protection of bacteria from foreign DNA in restriction modification systems.  DNA
methyltransferases are the enzymes that methylate DNA.  They deposit methyl groups on DNA at
the N6-position of adenine, or the N4- or C5-positions of cytosine in a sequence-specific
reaction using S-adenosyl-L-methionine as the methyl group donor.  Their reaction mechanism
includes rotating the target base completely out of the DNA helix in a biphasic process, where
fast flipping of the base out of the double helix is followed by a slower binding of the
flipped base into a hydrophobic pocket of the enzyme.  DNA MTases comprise two structural
domains: the larger domain contains the cofactor-binding site and the binding pocket for the
flipped base and the smaller domain is responsible for most of the sequence-specific contacts
of the enzyme to the target site.  The structures of large domains from all known DNA MTases
are similar, whereas the small domains are more heterogeneous in sequence and structure.  DNA
MTases are an attractive model system to study how proteins recognize specific sequences of
DNA and how the specificity of DNA recognition changes during molecular evolution.  In
addition, they illustrate how the biochemical properties of the enzymes are related to their
biological functions.  In this article, we shall describe the biological roles of DNA
methylation in prokaryotes, discuss the chemistry of the enzymatic methylation reaction
performed by the DNA methyltransferases (the focus of the review), and finally discuss aspects
of the enzymology of this fascinating family of enzymes that reveal how these molecular
machines perform their complicated biochemical tasks.

<>

<1>Jeltsch, A., Jurkowska, R.Z.
<2>Multimerization of the Dnmt3a DNA Methyltransferase and Its Functional Implications.
<3>Prog. Mol. Biol. Transl. Sci.
<4>117
<5>445-464
<6>2013
<7>The Dnmt3a DNA cytosine-C5 methyltransferase has been recently shown to exhibit a complex
oligomerization and multimerization potential, the
structural basis and functional implications of which will be the
subject of this contribution. The enzyme forms a linear heterotetramer
with Dnmt3L, in which the interaction of Dnmt3a and 3L stimulates the
catalytic activity of Dnmt3a. Isolated Dnmt3a forms protein filaments
that bind to several DNA molecules oriented in parallel, which plays an
essential role in the location of the enzyme to heterochromatin. Dnmt3L
disrupts Dnmt3a protein filaments and leads to a redistribution of the
enzyme in cells toward euchromatin. Finally, Dnmt3a complexes and
Dnmt3a/3L heterotetramers cooperatively multimerize on DNA forming
protein DNA filaments. This leads to a preference of the enzyme for
periodic methylation of DNA and supports its heterochromatic
localization.

<>

<1>Jeltsch, A., Jurkowska, R.Z., Jurkowski, T.P., Liebert, K., Rathert, P., Schlickenrieder, M.
<2>Application of DNA methyltransferases in targeted DNA methylation.
<3>Appl. Microbiol. Biotechnol.
<4>75
<5>1233-1240
<6>2007
<7>DNA methylation is an essential epigenetic modification. In bacteria, it is involved in gene
regulation, DNA repair, and control of cell cycle. In eukaryotes, it acts in concert with
other epigenetic modifications to regulate gene expression and chromatin structure. In
addition to these biological roles, DNA methyltransferases have several interesting
applications in biotechnology, which are the main focus of this review, namely, (1) in vivo
footprinting: as several bacterial DNA methyltransferases cannot methylate DNA bound to
histone proteins, the pattern of DNA methylation after expression of DNA methyltransferases in
the cell allows determining nucleosome positioning; (2) mapping the binding specificity of DNA
binding proteins: after fusion of a DNA methyltransferase to a DNA-binding protein and
expression of the fusion protein in a cell, the DNA methylation pattern reflects the
DNA-binding specificity of the DNA-binding protein; and (3) targeted gene silencing: after
fusion of a DNA methyltransferase to a suitable DNA-binding domain, DNA methylation can be
directed to promoter regions of target genes. Thereby, gene expression can be switched off
specifically, efficiently, and stably, which has a number of potential medical applications.

<>

<1>Jeltsch, A., Kroger, M., Pingoud, A.
<2>Evidence for an evolutionary relationship among type-II restriction endonucleases.
<3>Gene
<4>160
<5>7-16
<6>1995
<7>Type-II restriction-modification (R-M) systems comprise two enzymes, a DNA methyltransferase
(MTase) and a restriction endonuclease (ENase), each of which specifically interact with the
same 4-8-bp sequence.  All type-II MTases share several amino acid (aa) sequence motifs, which
makes an evolutionary relatedness among these enzymes probable.  The type-II ENases, in
contrast, except for some homologous isoschizomers, do not share significant aa sequence
similarity.  Therefore, ENases in general have been considered unrelated.  Here we show that
in addition to the analysis of the genotype (aa sequence), a comparison of the phenotype
(recognition sequence) of these enzymes can provide independent information regarding
evolutionary relationships, and thereby, help to analyze the significance of weak aa sequence
similarities.  Multistep Monte-Carlo analyses were employed to demonstrate that the
recognition sequences of those ENases, which were found to be related by a progressive
multiple aa sequence alignment, are more similar to each other than would be expected by
chance.  This analysis supports the notion that not only type-II MTases, but also type-II
ENases did not arise independently in evolution, but rather evolved from one or a few
primordial DNA-modifying and DNA-cleaving enzymes, respectively.

<>

<1>Jeltsch, A., Maschke, H., Selent, U., Wenz, C., Kohler, E., Connolly, B.A., Thorogood, H., Pingoud, A.
<2>DNA binding specificity of the EcoRV restriction endonuclease is increased by Mg2+ binding to a metal ion binding site distinct from the catalytic center of the enzyme.
<3>Biochemistry
<4>34
<5>6239-6246
<6>1995
<7>In contrast to many other type II restriction endonucleases, EcoRV binds specifically to DNA
only in the presence of Mg2+. According to the co-crystal structure of an EcoRV--DNA complex,
Mg2+ ion(s) bind to the active site of EcoRV liganded by Glu45, Asp74, and Asp90. Here we
present experimental evidence suggesting that the EcoRV--DNA complex also interacts with Mg2+
ions at other sites: (i) We have prepared an EcoRV triple mutant, in which all acidic amino
acids in the catalytic center are replaced by alanine. This mutant is catalytically inactive.
It binds nonspecifically to DNA in the absence of Mg2+, whereas it binds specifically to DNA
in the presence of Mg2+. This means that Mg2+ induces specific DNA binding in this mutant,
although all Mg2+ ligands in the catalytic center are removed. Therefore, additional
interactions between Mg2+ and the EcoRV--DNA complex probably occur at sites distinct from the
catalytic center. (ii) We have measured the specific and nonspecific DNA binding constants of
EcoRV and of the triple mutant in the presence and absence of Mg2+. Mg2+ reduces nonspecific
binding by 3-4 orders of magnitude, presumably because Mg2+ ions bound to the DNA have to be
released upon complex formation. In contrast, the specific binding of the wild-type enzyme and
the triple mutant is increased in the presence of Mg2+. This result can only be explained if a
Mg2+ ion binds to the specific EcoRV--DNA complex probably at a site distinct from the
catalytic center. (iii) To locate additional metal ion binding sites in the EcoRV--DNA
complex, we have determined the cleavage rates of several undecadeoxynucleotides which contain
single phosphorothioate linkages in the presence of Mg2+ and Mn2+. It turned out that an
oligodeoxynucleotide in which the first phosphate group within the GpATATC sequence (p3) is
replaced by an Rp phosphorothioate is cleaved by a factor of 50 more readily in the presence
of Mn2+ than with Mg2+. This result is interpreted to mean that p3, which is far away from the
active site of EcoRV, interacts with a Mg2+ ion. (iv) We have produced the EcoRV Y219C mutant
whose DNA cleavage activity compared to wild-type EcoRV is reduced by 3 orders of magnitude.
It binds nonspecifically to DNA in the absence of Mg2+ but not detectably in the presence of
Mg2+. Although the distinct role of Tyr219 is unclear at present, it must be pointed out that
this amino acid is far away from the active site of EcoRV but located in close proximity to
amino acid residues vis a vis p3. Hence, the behavior of this mutant also supports the
conclusion that an important interaction between Mg2+ and the EcoRV--DNA complex occurs at a
site distinct from the catalytic center.

<>

<1>Jeltsch, A., Nellen, W., Lyko, F.
<2>Two substrates are better than one: dual specificities for Dnmt2 methyltransferases.
<3>Trends Biochem. Sci.
<4>31
<5>306-308
<6>2006
<7>Dnmt2 enzymes have been widely conserved during evolution and contain all of the signature
motifs of DNA (cytosine-5)-methyltransferases;
however, the DNA methyltransferase activity of these proteins is
comparatively weak and their biochemical and functional properties
remain enigmatic. Recent evidence now shows that Dnmt2 has a novel tRNA
methyltransferase activity, raising the possibility that the biological
roles of these proteins might be broader than previously thought. This
finding has important implications for understanding the evolutionary
relationships among these enzymes.

<>

<1>Jeltsch, A., Pingoud, A.
<2>Horizontal gene transfer contributes to the wide distribution and evolution of type II restriction-modification systems.
<3>J. Mol. Evol.
<4>42
<5>91-96
<6>1996
<7>Restriction modification (RM) systems serve to protect bacteria against
bacteriophages.  They comprise a restriction endonuclease activity that specifically cleaves
DNA
and a corresponding methyltransferase activity that specifically methylates the DNA, thereby
protecting it from cleavage.  Such systems are very common in bacteria.  To find out whether
the
widespread distribution of RM systems is due to horizontal gene transfer, we have compared the
codon usages of 29 type II RM systems with the average codon usage of their respective
bacterial
hosts.  Pronounced deviations in codon usage were found in six cases: EcoRI, EcoRV, KpnI,
SinI, SmaI, and TthHB81.  They are interpreted as evidence for horizontal gene transfer in
these
cases.  As the methodology is expected to detect only one-fourth to one-third of all
horizontal gene
transfer events, this result implies that horizontal gene transfer had a considerable
influence on the
distribution and evolution of RM systems.  In all of these six cases the codon usage
deviations of
the restriction enzyme genes are much more pronounced than those of the methyltransferase
genes.
This result suggests that in these cases horizontal gene transfer had occurred sequentially
with the
gene for the methyltransferase being first acquired by the cell.  This can be explained by the
fact
that an active restriction endonuclease is highly toxic in cells whose DNA is not protected
from
cleavage by a corresponding methyltransferase.

<>

<1>Jeltsch, A., Pingoud, A.
<2>Kinetic characterization of linear diffusion of the restriction endonuclease EcoRV on DNA.
<3>Biochemistry
<4>37
<5>2160-2169
<6>1998
<7>We have examined the kinetic parameters of linear diffusion of EcoRV on DNA.  The data were
analyzed by Monte Carlo simulations in which the efficiency of recognition of EcoRV sites
during linear diffusion, the efficiency of linear diffusion, and the behavior of enzymes at
the ends of linear DNA is explicitly treated.  The analysis of the dependence of linear
diffusion on the concentrations of NaCl and MgCl2 shows that linear diffusion is maximal at 50
mM NaCl under all concentrations of MgCl2 tested and increases with increasing concentrations
of Mg2+ up to 10 mM, the highest concentration used in the test.  Under these conditions,
EcoRV scans 2 x 10^6 bp during one binding event with a velocity of about 1.7 x 10^6 bp s-1.
The enzyme tends to overlook cleavage sites at 1 mM but not at 10 mM MgCl2.  This result
confirms the thermodynamic finding that EcoRV does not bind very specifically to DNA in the
absence of Mg2+.  It demonstrates that there is a Mg2+-dependent continuous transition between
a nonspecific and a specific binding mode of EcoRV to DNA.  By comparing cleavage rates of
linear DNA whose ends are free or blocked, we have shown that EcoRV has a very low probability
to fall off at the ends of linear DNA.  The enzyme rather is "reflected" and continues linear
diffusion.  EcoRV does not cleave oligonucleotides containing two EcoRV sites processively.
Consequently, dissociation of the enzyme from the cleavage products is not preceded by a
transfer to nonspecific DNA, and linear diffusion is not involved in product dissociation in
EcoRV.

<>

<1>Jeltsch, A., Pingoud, A.M.
<2>Methods for determining activity and specificity of DNA binding and DNA cleavage by class II restriction endonucleases.
<3>Methods Mol. Biol.
<4>160
<5>287-308
<6>2001
<7>Restriction endonucleases coupled with DNA methyltransferases form the
restriction-modification systems that occur ubiquitously among bacteria.  They protect
bacterial cells against bacteriophage infection by cleaving incoming foreign DNA highly
specifically if it contains the recognition sequence.  Cellular DNA is protected from cleavage
by a specific methylation within the recognition sequence, which is introduced by the
methyltransferase.  Restriction endonucleases recognize palindromic recognition sites, 4-8
base pairs in length.  These enzymes are indispensable tools for genetic engineering.  The
biology and biochemistry of type II restriction endonucleases has been reviewed recently and
will be summarized only briefly here.  Type IIS restriction enzymes differ from type II
enzymes in that they recognize an asymmetric recognition sequence.  Monomeric in solution,
these enzymes consist of a DNA recognition domain and a catalytic domain.  Restriction
endonucleases initiate DNA cleavage by binding nonspecifically to DNA.  Association of the
enzymes to DNA is diffusion controlled, with rate constants in the order of 10^7-10^8
M^-1s^-1.  In a series of dissociation and association steps and/or sliding along the DNA, the
enzyme scans the DNA and searches for the recognition site in a one-dimensional diffusion
process.  This process permits much faster target site location than would be possible via a
three-dimensional search.  During recognition, conformational changes in the enzyme-DNA
complex lead to the activation of the catalytic centers and cleavage of the DNA.
Subsequently, the products are released, allowing for a new reaction cycle to take place.
Often product release is the rate limiting step for multiple turnover cleavage reactions.

<>

<1>Jeltsch, A., Pleckaityte, M., Selent, U., Wolfes, H., Siksnys, V., Pingoud, A.
<2>Evidence for substrate-assisted catalysis in the DNA cleavage of several restriction endonucleases.
<3>Gene
<4>157
<5>157-162
<6>1995
<7>Substrate-assisted catalysis was suggested to be involved in the DNA cleavage reaction of the
restriction endonucleases (ENases) EcoRI and EcoRV, because experimental evidence exists that
the phosphate group 3' to the scissile bond serves to deprotonate the attacking water.  Here,
we have addressed the question whether this is a general mechanistic feature of the reactions
catalyzed by ENases.  For this purpose, the cleavage rates of modified and unmodified
oligodeoxyribonucleotides (oligos), in which the phosphate group 3' to the scissile bond is
substituted by a methyl phosphonate, where measured for 17 enzymes.  Only five turned out not
to be inhibited by this modification (BglII, BstI, BstYI, Cfr10I and MunI); all others cleave
the modified substrate at a strongly reduced rate or not at all.  By employing a
hemisubstituted oligo substrate we were able to further investigate the mechanism of
inhibition of the latter group of ENases.  Some of them cleave the unmodified strand of the
modified substrate with a nearly unaltered rate, whereas the modified strand is cleaved very
slowly or not at all (BamHI, Bsp143I, Eco72I, MflI, NdeII, Sau3AI, XhoII).  The others (AluI,
Cfr9I, DpnII, MboI, PvuII) cleave the modified strand of the modified substrate with a largely
reduced rate or not at all.  These ENases, however, cleave the unmodified strand with a
reduced rate, too.  Based on these results we conclude that BamHI, Bsp143I, Cfr9I, DpnII,
Eco72I, MboI, MflI, NdeII, PvuII, Sau3AI and XhoII may possibly employ substrate assistance in
catalysis.

<>

<1>Jeltsch, A., Roth, M., Friedrich, T.
<2>Mutational analysis of target base flipping by the EcoRV adenine-N6 DNA methyltransferase.
<3>J. Mol. Biol.
<4>285
<5>1121-1130
<6>1999
<7>DNA methyltransferases flip their target base out of the DNA helix.  Here, we have
investigated base flipping by wild-type EcoRV DNA methyltransferase and five M.EcoRV variants
(D193A, Y196A, S229A, W231R and Y258A).  These variants bind to DNA and S-adenosyl-methionine
but have a severely reduced catalytic efficiency or are catalytically inactive.  To measure
base flipping three different assays were used, viz. analysis of the yields of
photocrosslinking reactions between the enzymes and a substrate in which the target base is
replaced by 5-iodouracil, analysis of the binding constants to substrates containing a
mismatch base-pair at the target position and analysis of the salt dependence of specific
complex formation.  Our data show that the Y196A, W231R and Y258A variants are not able to
stabilize a flipped target base, suggesting that the aromatic amino acid residues (Tyr196,
Trp231 and Tyr258) are involved in hydrophobic interactions with the flipped base.  The D193A
variant behaves like wild-type M.EcoRV with respect to base flipping.  The fact that this
variant is catalytically inactive indicates that Asp193 has a function in chemical catalysis.
The S229A variant can better flip modified bases but does not tightly lock the flipped base
into the adenine-binding pocket, suggesting that Ser229 could form a contact to the flipped
adenine.

<>

<1>Jeltsch, A., Sobotta, T., Pingoud, A.
<2>Structure prediction of the EcoRV DNA methyltransferase based on mutant profiling, secondary structure analysis, comparison with known structures of methyltransferases and isolation of catalytically inactive single mutants.
<3>Protein Eng.
<4>9
<5>413-423
<6>1996
<7>The EcoRV DNA methyltransferase (M.EcoRV) is an alpha-adenine methyltransferase.  We have used
two different programs to predict the secondary structure of M.EcoRV.  The resulting consensus
prediction was tested by a mutant profiling analysis.  29 neutral mutations of M.EcoRV were
generated by five cycles of random mutagenesis and selection for active variants to increase
the reliability of the prediction and to get a secondary structure prediction for some
ambiguously predicted regions.  The predicted consensus secondary structure elements could be
aligned to the common topology of the structures of the catalytic domains of M.HhaI and
M.TaqI.  In a complementary approach we have isolated nine catalytically inactive single
mutants.  Five of these mutants contain an amino acid exchange within the catalytic domain of
M.EcoRV (Val20-Ala, Lys81Arg, Cys192Arg, Asp193Gly, Trp231Arg).  The Trp231Arg mutant binds
DNA similarly to wild-type M.EcoRV, but is catalytically inactive.  Hence this mutant behaves
like a bona fide active site mutant.  According to the structure prediction, Trp231 is located
in a loop at the putative active site of M.EcoRV.  The other inactive mutants were insoluble.
They contain amino acid exchanges within the conserved amino acid motifs X, III or IV in
M.EcoRV confirming the importance of these regions.

<>

<1>Jeltsch, A., Urbanke, C.
<2>Sliding or hopping?  How restriction enzymes find their way on DNA.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>95-110
<6>2004
<7>In this contribution, we discuss target site location of restriction endonucleases, which are
extremely common among prokaryotes occurring in almost every species.  These enzymes
specifically recognize and cleave short, often palindromic, sequences on bacteriophage or
other kinds of foreign DNA.  By cleaving incoming DNA, they protect bacteria against
bacteriophage infections acting like an immune system.  In addition, restriction endonucleases
have an important role in the control of horizontal gene transfer and bacterial evolution.
The cellular DNA is protected against nucleolytic attack by an accompanying DNA
methyltransferase which recognizes the same nucleotide sequence and methylates one adenine or
cytosine residue within the site.  Since restriction endonucleases and DNA methyltransferases
are present at the same time in the cell, there is kinetic competition between them, which
necessitates that the restriction enzyme finds its target site on an invading DNA faster than
the methyltransferase.  In addition, fast target site location is also important, because the
invading bacteriophage DNA has to be degraded before it gains control over the cellular
metabolism.  Therefore, restriction enzymes were among the earliest examples where facilitated
diffusion has been shown to be effective in target site location.  However, the actual
mechanism of this process is still under debate; whereas most authors had proposed a sliding
mechanism, this has recently been disputed by Halford and colleagues who favored a hopping
process.

<>

<1>Jeltsch, A., Wenz, C., Stahl, F., Pingoud, A.
<2>Linear diffusion of the restriction endonuclease EcoRV on DNA is essential for the in vivo function of the enzyme.
<3>EMBO J.
<4>15
<5>5104-5111
<6>1996
<7>Linear diffusion along DNA is a mechanism of enhancing the association rates of proteins to
their specific recognition sites on DNA.  It has been demonstrated for several proteins in
vitro, but to date in no case in vivo.  Here we show that the restriction endonuclease EcoRV
slides along the DNA, scanning ~1000 bp in one binding event.  This process is critically
dependent on contacts between amino acid residues of the protein and the backbone of the DNA.
The disruption of single hydrogen bonds and, in particular, the alteration of electrostatic
interactions between amino acid side chains of the protein and phosphate groups of the DNA
interfere with or abolish effective sliding.  The efficiency of linear diffusion is dependent
on salt concentration, having a maximum at 50mM NaCl.  These results suggest that a
nonspecific and mobile binding mode capable of linear diffusion is dependent on a subtle
balance of forces governing the interaction of the enzyme and the DNA.  A strong correlation
between the ability of EcoRV mutants to slide along the DNA in vitro and to protect
Escherichia coli cells from phage infection demonstrates that linear diffusion occurs in vivo
and is essential for effective phage restriction.

<>

<1>Jeltsch, A., Wenz, C., Wende, W., Selent, U., Pingoud, A.
<2>Engineering novel restriction endonucleases: principles and applications.
<3>Trends Biotechnol.
<4>14
<5>235-238
<6>1996
<7>Restriction endonucleases cleave DNA with remarkable sequence specificity.  In this review, we
summarize the status of, and prospects for, engineering restriction endonucleases with new
specificities.  Such variants could be of considerable commercial value because restriction
enzymes are among the most frequently used enzymes in molecular biology, and not all the
desirable specificities are available.  While it has not yet been possible to effect
specificity changes, mutants have been described that (1) exhibit relaxed specificity, (2)
favor modified substrates over their natural substrates, (3) discriminate between cleavage
sites located in different sequences, (4) prefer metal ions other than Mg2+ as cofactors for
cleavage, or (5) possess site-specific DNA-nicking activity.

<>

<1>Jen-Jacobson, L.
<2>Structural-perturbation approaches to thermodynamics of site-specific protein-DNA interactions.
<3>Methods Enzymol.
<4>259
<5>305-345
<6>1995
<7>The solution of a large (and rapidly growing) number of crystal structures of site-specific
protein-DNA complexes has given us extremely detailed views of the positional relationships at
the interfaces between proteins and their "correct" DNA recognition sites.  This structural
information identifies interactions between particular functional groups on the protein and
particular functional groups on the DNA.  It is important to ask if the same proximity
relationships and interactions pertain in solution.  Furthermore, structural information alone
informs us about neither the quantitative contribution of each contact to the overall
stability of the complex nor the role of the contacts, singly or in combination, in enabling
the protein to discriminate between correct and incorrect DNA-binding sites.

<>

<1>Jen-Jacobson, L.
<2>Protein-DNA recognition complexes: conservation of structure and binding energy in the transition state.
<3>Biopolymers
<4>44
<5>153-180
<6>1997
<7>This paper considers how enzymes that catalyze reactions at specific DNA sites have been
engineered to overcome the problem of competitive inhibition by excess nonspecific binding
sites on DNA.  The formation of a specific protein-DNA recognition complex is discussed from
both structural and thermodynamic perspectives, and contrasted with formation of nonspecific
complexes.  Evidence (from EcoRI and BamHI endonucleases) is presented that a wide variety of
perturbations of the DNA substrate alter binding free energy but do not affect the free energy
of activation for the chemical step; that is, many energetic factors contribute equally to the
recognition complex and the transition-state complex.  This implies that the specific
recognition complex bears a close resemblance to the transition-state complex, such that very
tight binding to the recognition site on the DNA substrate does not inhibit catalysis, but
instead provides energy that is efficiently utilized along the path to the transition state.
It is suggested that this view can be usefully extended to "noncatalytic" site-specific
DNA-binding proteins like transcriptional activators and general transcription factors.

<>

<1>Jen-Jacobson, L., Engler, L.E., Ames, J.T., Kurpiewski, M.R., Grigorescu, A.
<2>Thermodynamic parameters of specific and nonspecific protein-DNA binding.
<3>Supramol. Chem.
<4>12
<5>143-160
<6>2000
<7>Proteins that bind preferentially to specific recognition sites on DNA also bind more weakly
to nonspecific DNA.  We have studied both specific and non-specific binding of the EcoRI and
BamHI restriction endonucleases, and determined enthalpic and entropic contributions to
binding free energy (delta G^o bind) using both the van't Hoff method and isothermal
titration calorimetry.  Specific binding is characterized by a strongly negative delta C^o p
and can be either enthalpy-driven or entropy-driven, depending on temperature.  Nonspecific
binding has delta C^o = 0 and is enthalpy-driven.  A strongly negative delta C^o p is the
"thermodynamic signature" of site-specific binding, because it reflects the characteristics of
a tight complementary recognition interface: the burial of previously hydrated nonpolar
surface and restriction of configurational-vibrational freedoms of protein, DNA, and water
molecules trapped at the protein-DNA interface.  These factors are absent in nonspecific
complexes.  We probed the contributions to delta C^o p by varying the sequence context
surrounding the recognition site.  As delta G^o bind improves, delta C^o p, delta H^o and
delta S^o all become more negative, and there is a linear correlation between delta H^o and
delta S^o (enthalpy-entropy compensation).  Because these context variations do not change the
protein-base or protein-phosphate contacts, the hydrophobic contribution or the number of
trapped water molecules at the interface, we conclude that a better sequence context improves
the "goodness of fit" in the interface and thus increases the magnitude of the negative
configurational-vibrational contribution to delta C^o p.

<>

<1>Jen-Jacobson, L., Engler, L.E., Jacobson, L.A.
<2>Structural and thermodynamic strategies for site-specific DNA binding proteins.
<3>Structure
<4>8
<5>1015-1023
<6>2000
<7>BACKGROUND: Site-specific protein-DNA complexes vary greatly in structural properties and in
the thermodynamic strategy for achieving an appropriate binding free energy. A better
understanding of the structural and energetic engineering principles might lead to rational
methods for modification or design of such proteins.
RESULTS: A novel analysis of ten site-specific protein-DNA complexes reveals a striking
correspondence between the degree of imposed DNA distortion and the thermodynamic parameters
of each system. For complexes with relatively undistorted DNA, favorable enthalpy change
drives unfavorable entropy change, whereas for complexes with highly distorted DNA,
unfavorable DeltaH^o is driven by favorable DeltaS^o. We show for the first time that
protein-DNA associations have isothermal enthalpy-entropy compensation, distinct from
temperature-dependent compensation, so DeltaH^o and DeltaS^o do not vary independently. All
complexes have favorable DeltaH^o from direct protein-DNA recognition interactions and
favorable DeltaS^o from water release.  Systems that strongly distort the DNA nevertheless
have net unfavorable DeltaH^o as the result of molecular strain, primarily associated with the
base pair destacking. These systems have little coupled protein folding and the strained
interface suffers less immobilization, so DeltaS^o is net favorable. By contrast, systems with
little DNA distortion have net favorable DeltaH^o, which must be counterbalanced by net
unfavorable DeltaS^o, derived from loss of vibrational entropy (a result of isothermal
enthalpy-entropy compensation) and from coupling between DNA binding and protein folding.
CONCLUSIONS: Isothermal enthalpy-entropy compensation implies that a structurally optimal,
unstrained fit is achieved only at the cost of entropically unfavorable immobilization,
whereas an enthalpically weaker, strained interface entails smaller entropic penalties.

<>

<1>Jen-Jacobson, L., Engler, L.E., Lesser, D.R., Kurpiewski, M.R., Yee, C., McVerry, B.
<2>Structural adaptations in the interaction of EcoRI endonuclease with methylated GAATTC sites.
<3>EMBO J.
<4>15
<5>2870-2882
<6>1996
<7>We have studied the interaction of EcoRI endonuclease with oligonucleotides
containing GAATTC sites bearing one or two adenine-N6-methyl groups, which would be in steric
conflict with key protein side chains involved in recognition and/or catalysis in the
canonical
complex.  Single-strand methylation of either adenine produces small penalties in binding free
energy (delta delta Gs ~ +1.4 kcal/mol), but elicits asymmetric structural adaptations in the
complex, such that cleavage rate constants are strongly inhibited and unequal in the two DNA
strands.  The dependences of cleavage rate constants on the concentration of the Mg2+ cofactor
are
unaltered.  When either adenine is methylated on both DNA strands, delta delta Gs
(~+4kcal/mol)
is larger than the expected sum of the delta delta Gs values for the single-strand
methylations,
because the asymmetric adaptations cannot occur.  Cleavage rate constants are reduced by
600,000-fold for the biologically relevant GAmATTC/CTTmAAG site, but the
GmAATTC/CTTAmAG site forms only a non-specific complex that cannot be cleaved.  These
observations provide a detailed thermodynamic and kinetic explanation of how single-strand and
double-strand methylation protect against endonuclease cleavage in vivo.  We propose that non-
additive effects on binding and structural 'adaptations' are important in understanding how
DNA
methylation modulates the biological activities of non-catalytic DNA binding proteins.

<>

<1>Jen-Jacobson, L., Kurpiewski, M., Lesser, D., Grable, J., Boyer, H.W., Rosenberg, J.M., Greene, P.J.
<2>Coordinate ion pair formation between EcoRI endonuclease and DNA.
<3>J. Biol. Chem.
<4>258
<5>14638-14646
<6>1983
<7>The free energy of the binding reaction between EcoRI restriction endonuclease
and a specific cognate dodecadeoxynucleotide (d(CGCGAATTCGCG)) has
contributions from both electrostatic and nonelectrostatic components.  These
contributions were dissected by measuring the effects of varying salt
concentration on the equilibrium binding constant and applying the
thermodynamic analyses of Record et al. (Record, M.T., Jr., Lohman, T.M., and
deHaseth, P.L. (1976) J. Mol. Biol. 107: 145-158).  Endonuclease mutation S187
(Arg 187 to Ser) (Greene, P.J., Gupta, M., Boyer, H.W., Brown, W.E., and
Rosenberg, J.M. (1981) J. Biol. Chem. 256: 2143-2153) did not significantly
affect the nonelectrostatic component but did perturb the electrostatic
contribution to the binding energy (we are numbering the amino acid residues
according to the DNA sequence).  The former was determined by extrapolating the
linear portion of the salt dependence curve (0.125 to 0.25M KCl) to 1M ionic
strength, with the same result for both wild type and S187 endonucleases at
both pH 6.0 and 7.4 (-8.5 +- 1.5 kcal/mol or greater than 50% of the total
binding free energy).  The slopes of these same curves yield estimates of eight
ionic interactions between wild type endonuclease and the DNA at both pH
values.  By contrast, binding of EcoRI-S187 to dodecanucleotide involves six
charge-charge interactions at pH 6.0.  Only two ionic interactions are observed
at pH 7.4.  This was unexpected since gel permeation chromatography
demonstrated that the recognition complex for both wild type and S187 proteins
contains an enzyme dimer and a DNA duplex.  EcoRI-S187 endonuclease retains
wild type DNA sequence specificity, and the rate of the phosphodiester
hydrolysis step is also unchanged.  Thus, electrostatic interactions are
functionally separable from sequence recognition and strand cleavage.  Our
results also establish that arginine 187 plays a key role in the electrostatic
function and suggest that it might be located at the DNA-protein interface.
The disproportionate loss of ion pairs at pH 7.4 can be rationalized by a model
which suggests that six conformationally mobile ionic groups on the protein act
in a coordinated manner during the interaction with DNA.

<>

<1>Jen-Jacobson, L., Lesser, D., Kurpiewski, M.
<2>The enfolding arms of the EcoRI Endonuclease: role in DNA binding and cleavage.
<3>Cell
<4>45
<5>619-629
<6>1986
<7>The N-terminal segments of the EcoRI endonuclease dimer form part of mobile
"arms" that encircle DNA in the recognition complex.  By treating
endonuclease-TCGCGAATTCGCG complexes with proteases, we have prepared a series
of deletion derivatives lacking defined segments of the N-terminal region.  The
5-12 segment is essential for DNA cleavage and forms one electrostatic
interaction (per subunit) with DNA phosphate.  These ionic contacts are
directly across the double helix from the scissile phosphodiester bonds; they
thus may permit the enfolding arms to immobilize DNA in apposition to the
catalytic cleft and/or contribute to the unusual "kinked" conformation of DNA
in the complex.  Sequence specificity is fully retained when 28 residues are
deleted from the N-terminus, but the complexes dissociate more rapidly.

<>

<1>Jen-Jacobson, L., Lesser, D.R., Kurpiewski, M.R.
<2>DNA sequence discrimination by EcoRI endonuclease.
<3>Nucleic Acids and Molecular Biology, Springer-Verlag, Eckstein, F., Lilley, D.M.J., 
<4>5
<5>141-170
<6>1991
<7>The EcoRI restriction endonuclease presents an especially informative case of specificity in
protein-DNA interactions, because it exhibits extremely high sequence selectivity between its
canonical recognition site GAATTC and related DNA sites that differ by as little as one base
pair.  A gene-regulatory protein (the lac repressor) also shows stringent discrimination
against single base-pair changes in Oc mutant sites.  By contrast, some bacteriophage
repressors bind to a series of related operator DNA sites in a graduated fashion ("permissive
discrimination"), yet discriminate stringently between operator and nonoperator DNA.

<>

<1>Jenkinson, E., Crickmore, N.
<2>Identification of a novel DNA methyltransferase activity from Bacillus thuringiensis.
<3>Curr. Microbiol.
<4>47
<5>144-145
<6>2003
<7>A DNA methyltransferase activity was identified in a strain of Bacillus thuringiensis that was
found to protect DNA from cleavage by the
restriction endonuclease HaeIII at overlapping sites. Site-directed
mutagenesis was used to confirm therecognition sequence of the
methyltransferase as ACGGC.

<>

<1>Jennert, K.C., Tardif, C., Young, D.I., Young, M.
<2>Gene transfer to clostridium cellulolyticum ATCC 35319.
<3>Microbiology
<4>146
<5>3071-3080
<6>2000
<7>Although much is known about the bacterial cellulosome and its various protein components,
their contributions to bacterial growth on cellulose and the process of cellulolysis in vivo
cannot currently be assessed. To remedy this, the authors have developed gene transfer
techniques for Clostridium cellulolyticum ATCC 35319. Firstly, transfer of Tn1545 has been
obtained using an Enterococcus faecalis donor. Secondly, IncP-mediated conjugative
mobilization of plasmids from Escherichia coli donors has also been achieved. The yield of
transconjugants in both cases was low and was probably limited by the suboptimal growth
conditions that must of necessity be employed for the co-culture of oligotrophic C.
cellulolyticum with copiotrophic donors. A restriction endonuclease was detected in crude
extracts of C. cellulolyticum. This enzyme, named CCE:I, is an isoschizomer of MSP:I (HPA:II).
Electro-transformation was employed to establish plasmids containing the replication functions
of pAMss1 (En. faecalis), pIM13 (Bacillus subtilis), pCB102 (Clostridium butyricum), pIP404
(Clostridium perfringens) and pWV01 (Lactococcus lactis subsp. cremoris) in C. cellulolyticum.
Transformants were only obtained if the DNA was appropriately methylated on the external C of
the sequence 5'-CCGG-3' using either BSU:FI methylase in vivo or MSP:I methylase in vitro.
Plasmids based on the pAMss1 and pIM13 replicons were more stably maintained than one based on
the pCB102 replicon. Selection of transformants on solid medium led to low apparent
transformation efficiencies (approx. 10(2) transformants per &mgr;g DNA) which might, in part,
reflect the low plating efficiency of the organism. Selection of transformants in liquid
medium led to a higher apparent yield of transformants (between 10(5) and 10(7) transformants
per &mgr;g DNA). The methods developed here will pave the way for functional analysis of the
various cellulosome components in vivo.

<>

<1>Jensen, R.V., Depasquale, S.M., Harbolick, E.A., Hong, T., Kernell, A.L., Kruchko, D.H., Modise, T., Smith, C.E., McCarter, L.L., Stevens, A.M.
<2>Complete Genome Sequence of Prepandemic Vibrio parahaemolyticus BB22OP.
<3>Genome Announcements
<4>1
<5>e00002-12
<6>2013
<7>The number of inflammatory gastroenteritis outbreaks due to the food-borne pathogen is rising
sharply worldwide and in the United States in particular. Here
we report the complete, annotated genome sequence of the prepandemic strain
BB22OP and make some initial comparisons to the complete genome sequence for
pandemic strain RIMD2210633.

<>

<1>Jensen, T.O., Kvist, T., Mikkelsen, M.J., Westermann, P.
<2>Rapid and reliable method for identification of associated endonuclease cleavage and recognition sites.
<3>Lett. Appl. Microbiol.
<4>58
<5>576-581
<6>2014
<7>One barrier to cross during genetic engineering is the restriction-modification system found
in many bacteria. In this study, we developed a fast and reliable method for mapping the
recognition and cleavage site of the restriction endonucleases. Clostridium pasteurianum, a
model organism for the study of nitrogen fixation, has been found to harbour at least two
restriction-modification systems including the restriction endonucleases CpaPI, which is an
isoschizomer of MboI and CpaAI. Dam-methylated DNA was used to isolate the activity of CpaAI.
Exposing freshly prepared cell lysate to known nucleotide fragments and directly sequencing
the pool of digested nucleotide fragments enabled identification of the cleavage sites in the
fragments. By aligning the sequences adjacent to the cleavage site, it was possible to
identify the recognition sequence. Using this method, we successfully located all CpaAI
recognition and cleavage sites within the template sequence. By modifying DNA with both Dam
and CpG methylases (M.SssI) and thereby preventing digestion by CpaPI and CpaAI, no further
endonuclease activity was detected.Significance and Impact of the
StudyRestriction-modification systems are important barriers to successful genetic
modification in many bacterial species. In this study, we demonstrate an efficient and general
applicable method for identifying endonuclease recognition and cleavage sites. For the study
and the trails, the model organism for nitrogen fixation Clostridium pasteurianum was used.
The method was proven to be reliable, and by modifying DNA at the identified sites, it is
possible to prevent digestion.

<>

<1>Jentsch, S.
<2>Restriction and modification in Bacillus subtilis:  Sequence specificities of restriction/modification systems BsuM, BsuE, and BsuF.
<3>J. Bacteriol.
<4>156
<5>800-808
<6>1983
<7>The sequence specificies of three Bacillus subtilis restriction/modification
systems were established: (i) BsuM (CTCGAG), an isoschizomer to XhoI; (ii) BsuE
(CGCG), an isoschizomer to FnuDII; and (iii) BsuF (CCGG), an isoschizomer to
MspI, HpaII.  The BsuM modification enzyme methylates the 3' cytosine of the
recognition sequence.  The BsuF modification enzyme methylates the 5' cytosine
of the sequence, rendering such sites resistant to MspI degradation and leaving
the majority of sites sensitive to HpaII degradation.

<>

<1>Jentsch, S., Gunthert, U., Trautner, T.A.
<2>DNA methyltransferases affecting the sequence 5'CCGG.
<3>Nucleic Acids Res.
<4>9
<5>2753-2759
<6>1981
<7>B. subtilis phage SPbeta and Moraxella sp. code for DNA methyl-transferases
which methylate both cytosines of the sequence 5'CCGG.  Experiments using a B.
subtilis strain whose DNA is sensitive to HpaII and resistant to MspI
degradation, indicated that methylation of the outer C of this sequence
provides protection against the restriction enzyme MspI.

<>

<1>Jentsch, S., Pawlek, B., Noyer-Weidner, M., Gunthert, U., Trautner, T.A.
<2>Restriction and modification in B. subtilis:  Temperate phages SPbeta and Phi3T for BsuR specific methyltransferases.
<3>Transformation 1980:  Proceedings of the Fifth European Meeting on Bacterial Transformation and Transfection, Cotswold Press, Polsinelli, M., Mazza, G., Oxford
<4>0
<5>363-369
<6>1980
<7>Genes for BsuR specific methyltransferases have been identified in the genomes
of SPbeta and Phi3T.  Such genes are interchangeable between these two phages.
Phages SPbeta and possibly also Phi3T code for additional methyltransferase(s)
with different sequence specificities.

<>

<1>Jeon, T.J.
<2>DNA Adenine Methylation of sams1 Gene in Symbiont-Bearing Amoeba proteus.
<3>J. Microbiol.
<4>46
<5>564-570
<6>2008
<7>The expression of amoeba sams genes is switched from sams1 to sams2 when amoebae are infected
with Legionella jeonii. To elucidate the
mechanism for the inactivation of host sams1 gene by endosymbiotic
bacteria, methylation states of the sams1 gene of D and xD amoebae was
compared in this study. The sams1 gene of amoebae was methylated at an
internal adenine residue of GATC site in symbiont-bearing xD amoebae
but not in symbiont-free D amoebae, suggesting that the modification
might have caused the inactivation of sams1 in xD amoebae. The sams1
gene of xD amoebae was inactivated at the transcriptional level.
Analysis of DNA showed that adenine residues in L. jeonii sams were
also methylated, implying that L. jeonii bacteria belong to a Dam
methylase-positive strain. In addition, both SAM and Met appeared to
act as negative regulators for the expression of sams1 whereas the
expression of sams2 was not affected in amoebae.

<>

<1>Jeong, H., Barbe, V., Lee, C.H., Vallenet, D., Yu, D.S., Choi, S.H., Couloux, A., Lee, S.W., Yoon, S.H., Cattolico, L., Hur, C.G., Park, H.S., Segurens, B., Kim, S.C., Oh, T.K., Lenski, R.E., Studier, F.W., Daegelen, P., Kim, J.F.
<2>Genome Sequences of Escherichia coli B strains REL606 and BL21(DE3).
<3>J. Mol. Biol.
<4>394
<5>644-652
<6>2009
<7>Escherichia coli K-12 and B have been the subjects of classical experiments from which much of
our understanding of molecular genetics has
emerged. We present here complete genome sequences of two E. coli B
strains, REL606, used in a long-term evolution experiment, and BL21(DE3),
widely used to express recombinant proteins. The two genomes differ in
length by 72,304 bp and have 426 single base pair differences, a seemingly
large difference for laboratory strains having a common ancestor within
the last 67 years. Transpositions by IS1 and IS150 have occurred in both
lineages. Integration of the DE3 prophage in BL21(DE3) apparently
displaced a defective prophage in the lambda attachment site of B. As
might have been anticipated from the many genetic and biochemical
experiments comparing B and K-12 over the years, the B genomes are similar
in size and organization to the genome of E. coli K-12 MG1655 and have
>99% sequence identity over approximately 92% of their genomes. E. coli B
and K-12 differ considerably in distribution of IS elements and in
location and composition of larger mobile elements. An unexpected
difference is the absence of a large cluster of flagella genes in B, due
to a 41 kbp IS1-mediated deletion. Gene clusters that specify the LPS
core, O antigen, and restriction enzymes differ substantially, presumably
because of horizontal transfer. Comparative analysis of 32 independently
isolated E. coli and Shigella genomes, both commensals and pathogenic
strains, identifies a minimal set of genes in common plus many
strain-specific genes that constitute a large E. coli pan-genome.

<>

<1>Jeong, H., Choi, S.K., Kloepper, J.W., Ryu, C.M.
<2>Genome Sequence of the Plant Endophyte Bacillus pumilus INR7, Triggering Induced  Systemic Resistance in Field Crops.
<3>Genome Announcements
<4>2
<5>e01093-14
<6>2014
<7>Bacillus pumilus INR7 is an endophytic bacterium that has been commercialized as  a biological
control product against soilborne pathogens as well as foliar
pathogens by direct antagonism and induction of systemic resistance. In the
current study, we provide the genome sequence and a possible explanation of the
function of strain INR7.

<>

<1>Jeong, H., Choi, S.K., Park, S.Y., Kim, S.H., Park, S.H.
<2>Draft Genome Sequence of Paenibacillus peoriae Strain KCTC 3763T.
<3>J. Bacteriol.
<4>194
<5>1237-1238
<6>2012
<7>Paenibacillus peoriae is a potentially plant-beneficial soil bacterium and is a close relative
to Paenibacillus polymyxa, the type species of the genus
Paenibacillus. Herein, we present the 5.77-Mb draft genome sequence of the P.
peoriae type strain with the aim of providing insight into the genomic basis of
plant growth-promoting Paenibacillus species.

<>

<1>Jeong, H., Jeong, D.E., Kim, S.H., Song, G.C., Park, S.Y., Ryu, C.M., Park, S.H., Choi, S.K.
<2>Draft Genome Sequence of the Plant Growth-Promoting Bacterium Bacillus siamensis  KCTC 13613T.
<3>J. Bacteriol.
<4>194
<5>4148-4149
<6>2012
<7>Bacillus siamensis KCTC 13613(T), a novel halophilic Bacillus species isolated from a salted
Thai food, produced antimicrobial compounds against plant pathogens
and promoted plant growth by volatile emission. We determined the 3.8-Mb genome
sequence of B. siamensis KCTC 13613(T) to reveal the plant-beneficial effect at
the genomic level.

<>

<1>Jeong, H., Jeong, D.E., Sim, Y.M., Park, S.H., Choi, S.K.
<2>Genome Sequence of Lysinibacillus sphaericus Strain KCTC 3346T.
<3>Genome Announcements
<4>1
<5>e00625-13
<6>2013
<7>Lysinibacillus sphaericus is a heterogeneous species that includes strains that produce
mosquitocidal toxin proteins. Herein, we report the 4.56-Mb draft genome
sequence of the nonpathogenic L. sphaericus strain KCTC 3346(T), which provides
clues for the phylogenetic reassessment of L. sphaericus species and an
understanding of its physiological properties.

<>

<1>Jeong, H., Park, S.H., Choi, S.K.
<2>Genome Sequence of the Acrystalliferous Bacillus thuringiensis Serovar Israelensis Strain 4Q7, Widely Used as a Recombination Host.
<3>Genome Announcements
<4>2
<5>e00231-14
<6>2014
<7>Bacillus thuringiensis serovar israelensis is well known for its mosquitocidal activity and
has long been used as a biopesticide. Herein, we present the genome sequence of B.
thuringiensis serovar israelensis strain 4Q7, a plasmid-cured derivative with higher
transformation efficiency than wild types.

<>

<1>Jeong, H., Park, S.Y., Chung, W.H., Kim, S.H., Kim, N., Park, S.H., Kim, J.F.
<2>Draft Genome Sequence of Paenibacillus polymyxa Type Strain (ATCC 842T), a Plant Growth-Promoting Bacterium.
<3>J. Bacteriol.
<4>193
<5>5026-5027
<6>2011
<7>Paenibacillus polymyxa is an endospore-forming Gram-positive soil bacterium that is well-known
for its ability to promote plant growth. Here
we report the draft genome sequence of P. polymyxa ATCC 842(T), the type
strain of the species P. polymyxa, and the family Paenibacillaceae. A
repertoire of biosynthetic genes for antibiotics and hydrolytic enzymes
accounts for its beneficial effects in rhizosphere to host plants it
associates with.

<>

<1>Jeong, H., Sim, Y.M., Kim, H.J., Lee, D.W., Lim, S.K., Lee, S.J.
<2>Genome Sequence of the Vancomycin-Producing Amycolatopsis orientalis subsp. orientalis Strain KCTC 9412T.
<3>Genome Announcements
<4>1
<5>e00408-13
<6>2013
<7>Amycolatopsis orientalis is the producer of vancomycin, a glycopeptide antibiotic that is used
for the treatment of serious infections with Gram-positive bacteria.
Here we present the next-generation sequencing (NGS)-based 9.06-Mb draft genome
sequence of the type strain Amycolatopsis orientalis subsp. orientalis KCTC 9412
(DSM 40040; ATCC 19795).

<>

<1>Jeong, H., Sim, Y.M., Kim, H.J., Lee, Y.J., Lee, D.W., Lim, S.K., Lee, S.J.
<2>Genome Sequences of Amycolatopsis orientalis subsp. orientalis Strains DSM 43388  and DSM 46075.
<3>Genome Announcements
<4>1
<5>e00545-13
<6>2013
<7>Strains of Amycolatopsis orientalis produce vancomycin or other related glycopeptide
antibiotic compounds. Here we report the draft genome sequences of
glycopeptide nonproducers Amycolatopsis orientalis subsp. orientalis DSM 43388
and DSM 46075. Their genome information will provide insights into the
acquisition and regulation of glycopeptide antibiotic resistance genes.

<>

<1>Jeong, H., Yim, J.H., Lee, C., Choi, S.H., Park, Y.K., Yoon, S.H., Hur, C.G., Kang, H.Y., Kim, D., Lee, H.H., Park, K.H., Park, S.H., Park, H.S., Lee, H.K., Oh, T.K., Kim, J.F.
<2>Genomic blueprint of Hahella chejuensis, a marine microbe producing an algicidal agent.
<3>Nucleic Acids Res.
<4>33
<5>7066-7073
<6>2005
<7>Harmful algal blooms, caused by rapid growth and accumulation of certain microalgae in the
ocean, pose considerable impacts on marine environments,
aquatic industries and even public health. Here, we present the
7.2-megabase genome of the marine bacterium Hahella chejuensis including
genes responsible for the biosynthesis of a pigment which has the lytic
activity against a red-tide dinoflagellate. H.chejuensis is the first
sequenced species in the Oceanospiralles clade, and sequence analysis
revealed its distant relationship to the Pseudomonas group. The genome was
well equipped with genes for basic metabolic capabilities and contained a
large number of genes involved in regulation or transport as well as with
characteristics as a marine heterotroph. Sequence analysis also revealed a
multitude of genes of functional equivalence or of possible foreign
origin. Functions encoded in the genomic islands include biosynthesis of
exopolysacchrides, toxins, polyketides or non-ribosomal peptides, iron
utilization, motility, type III protein secretion and pigmentation.
Molecular structure of the algicidal pigment, which was determined through
LC-ESI-MS/MS and NMR analyses, indicated that it is prodigiosin. In
conclusion, our work provides new insights into mitigating algal blooms in
addition to genetic make-up, physiology, biotic interactions and
biological roles in the community of a marine bacterium.

<>

<1>Jeong, H., Zhao, F., Igori, D., Oh, K.H., Kim, S.Y., Kang, S.G., Kim, B.K., Kwon, S.K., Lee, C.H., Song, J.Y., Yu, D.S., Park, M.S., Cho, S.H., Kim, J.F.
<2>Genome Sequence of the Hemolytic-Uremic Syndrome-Causing Strain Escherichia coli  NCCP15647.
<3>J. Bacteriol.
<4>194
<5>3747-3748
<6>2012
<7>Enterohemorrhagic Escherichia coli (EHEC) causes a disease involving diarrhea, hemorrhagic
colitis, and hemolytic-uremic syndrome (HUS). Here we present the
draft genome sequence of NCCP15647, an EHEC isolate from an HUS patient. Its
genome exhibits features of EHEC, such as genes for verotoxins, a type III
secretion system, and prophages.

<>

<1>Jeong, S., Liang, G.N., Sharma, S., Lin, J.C., Choi, S.H., Han, H., Yoo, C.B., Egger, G., Yang, A.S., Jones, P.A.
<2>Selective Anchoring of DNA Methyltransferases 3A and 3B to Nucleosomes Containing Methylated DNA.
<3>Mol. Cell. Biol.
<4>29
<5>5366-5376
<6>2009
<7>Proper DNA methylation patterns are essential for mammalian development and differentiation.
DNA methyltransferases (DNMTs) primarily establish
and maintain global DNA methylation patterns; however, the molecular
mechanisms for the generation and inheritance of methylation patterns
are still poorly understood. We used sucrose density gradients of
nucleosomes prepared by partial and maximum micrococcal nuclease
digestion, coupled with Western blot analysis to probe for the
interactions between DNMTs and native nucleosomes. This method allows
for analysis of the in vivo interactions between the chromatin
modification enzymes and their actual nucleosomal substrates in the
native state. We show that little free DNA methyltransferase 3A and 3B
(DNMT3A/3B) exist in the nucleus and that almost all of the cellular
contents of DNMT3A/3B, but not DNMT1, are strongly anchored to a subset
of nucleosomes. This binding of DNMT3A/3B does not require the presence
of other well-known chromatin-modifying enzymes or proteins, such as
proliferating cell nuclear antigen, heterochromatin protein 1,
methyl-CpG binding protein 2, Enhancer of Zeste homolog 2, histone
deacetylase 1, and UHRF1, but it does require an intact nucleosomal
structure. We also show that nucleosomes containing methylated SINE and
LINE elements and CpG islands are the main sites of DNMT3A/3B binding.
These data suggest that inheritance of DNA methylation requires cues
from the chromatin component in addition to hemimethylation.

<>

<1>Jeong, Y., Song, Y., Shin, H.S., Cho, B.K.
<2>Draft Genome Sequence of Acid-Tolerant Clostridium drakei SL1T, a Potential Chemical Producer through Syngas Fermentation.
<3>Genome Announcements
<4>2
<5>e00387-14
<6>2014
<7>Clostridium drakei SL1(T) is a strictly anaerobic, H2-utilizing, and acid-tolerant acetogen
isolated from an acidic sediment that is a potential
platform for commodity chemical production from syngas fermentation. The draft
genome sequence of this strain will enable determination of the acid resistance
and autotrophic pathway of the acetogen.

<>

<1>Jeong, Y.S., Oh, K.B., Park, J.S., Kim, J.S., Kang, Y.K.
<2>Cytoplasmic Localization of Oocyte-Specific Variant of Porcine DNA Methyltransferase-1 During Early Development.
<3>Dev. Dyn.
<4>238
<5>1666-1673
<6>2009
<7>DNA methyltransferase-1 (Dnmt1) is involved in the maintenance of genomic methylation
patterns. Rather than full-length Dnmt1, mouse
oocytes have a truncated variant called Dnmt1o. Immunofluorescence data
showed that Dnmt1o localized to the cytoplasm, but this has not been
confirmed using more direct methods. The cytoplasmic localization of
Dnmt1o has been assigned to the main cause of global DNA demethylation
in early mouse embryos. We studied localization of Dnmt1o in mouse and
pig embryos. We identified pig Dnmt1o protein and its transcript with
unique 5'-end sequence. Physically separating mouse and pig 2-cell
embryos into their nuclear and cytoplasmic components demonstrated that
Dnmt1o of both species localized to the cytoplasm. Cloned pig embryos
had Dnmt1o as the main form, with no indication of somatic Dnmt1. These
findings indicate that Dnmt1o is cytoplasmic during early development;
its presence in both pig and mouse embryos further suggests that Dnmt1o
is conserved in mammals. Developmental Dynamics 238:1666-1673, 2009.

<>

<1>Jerke, K., Nakatsu, C.H., Beasley, F., Konopka, A.
<2>Comparative analysis of eight Arthrobacter plasmids.
<3>Plasmid
<4>59
<5>73-85
<6>2008
<7>Despite the prevalence of Arthrobacter in the environment little is known
about their plasmids, or the capacity of Arthrobacter plasmids to mediate
horizontal gene transfer. In this study, we compared eight plasmids from
five Arthrobacter strains in order to identify putative core maintenance
genes for replication, segregation, and conjugation. Iteron like sequences
were identified on some of the plasmids; however, no genes with obvious
similarity to known replication sequences such as an origin of
replication, or rep genes were identified. All eight plasmids contained a
putative conjugation system. Genes with similarity to a relaxase, coupling
protein, and various components of a type IV secretion system were
identified on each plasmid; it appears that three different systems may be
present. Putative parA partitioning genes were found in all of the
plasmids. Each of the Arthrobacter strains examined contained a putative
parB gene; however, of the three plasmids in Arthrobacter strain FB24 only
one plasmid had a putative parB gene. Cluster analysis of many of the
Arthrobacter genes suggested that they often formed branches within
existing families of plasmid maintenance genes. Comparison of a
concatenation of all the maintenance genes from each plasmid suggests that
the eight Arthrobacter plasmids represent multiple evolutionary pathways.

<>

<1>Jermyn, W.S., Boyd, E.F.
<2>Characterization of a novel Vibrio pathogenicity island (VPI-2) encoding neuraminidase (nanH) among toxigenic Vibrio cholerae isolates.
<3>Microbiology
<4>148
<5>3681-3693
<6>2002
<7>Acquisition of virulence genes encoded on mobile genetic elements has played an important role
in the emergence of pathogenic isolates of
Vibrio cholerae, the causative agent of the diarrhoeal disease cholera.
The genes encoding cholera toxin (ctxAB) the main cause of profuse
secretory diarrhoea in cholera, are encoded on a filamentous
bacteriophage CTXphi. The toxin coregulated pilus (TCP), an essential
intestinal colonization factor, was originally designated as part of a
pathogenicity island named the Vibrio pathogenicity island (VPI), but
this island has more recently been proposed to be the genome of a
filamentous phage, VPIphi. In this study, it is shown that nanH, which
encodes neuraminidase, maps within a novel pathogenicity island
designated VP1-2. The 57-3 kb VP1-2 has all of the characteristic
features of a pathogenicity island, including the presence of a
bacteriophage-like integrase (int), insertion in a tRNA gene (serine)
and the presence of direct repeats at the chromosomal integration
sites. Additionally, the G+C content of VP1-2 (42 mol%) is considerably
lower than that of the entire genome (47 mol%). VPI-2 encodes several
gene clusters, such as a restriction modification system (hsdR and
hsdM) and genes required for the utilization of amino sugars (nan-nag
region) as well as neuraminidase. To determine the distribution of
VPI-2 among V. cholerae, 78 natural isolates were examined using PCR
and Southern hybridization analysis for the presence of this region.
All toxigenic V. cholerae 01 serogroup isolates examined contained
VPI-2, whereas non-toxigenic isolates lacked the island. Of 14 V.
cholerae 0139 serogroup isolates examined, only one strain, MO2,
contained the entire 57-3 kb island, whereas 13 0139 isolates contained
only a 20-0 kb region with most of the 5' region of VPI-2 which
included nanH deleted in these strains.

<>

<1>Jerome, J.P., Klahn, B.D., Bell, J.A., Barrick, J.E., Brown, C.T., Mansfield, L.S.
<2>Draft Genome Sequences of Two Campylobacter jejuni Clinical Isolates, NW and D2600.
<3>J. Bacteriol.
<4>194
<5>5707-5708
<6>2012
<7>The Campylobacter jejuni human clinical isolates NW and D2600 colonized C57BL/6
interleukin-10-deficient (IL-10(-/-)) mice without inducing a robust inflammatory
response (J. A. Bell et al., BMC Microbiol. 9:57, 2009). We announce draft genome
sequences of NW and D2600 to facilitate comparisons with strains that induce
gastrointestinal inflammation in this mouse model.

<>

<1>Jessen, W.J., Dhasarathy, A., Hoose, S.A., Carvin, C.D., Risinger, A.L., Kladde, M.P.
<2>Mapping chromatin structure in vivo using DNA methyltransferases.
<3>Methods
<4>33
<5>68-80
<6>2004
<7>Cytosine-5 DNA methyltransferases (C5 DMTases) are effective reagents for analyzing chromatin
and footprinting DNA-bound factors in vivo.
Cytosine methylation in accessible regions is assayed positively by the
PCR-based technique of bisulfite sequencing. In this article, we
outline two complementary uses for the DNA methyltransferase CviPI
(M.CviPI, GC specificity) in probing chromatin organization. First, we
describe the use of the naturally occurring, free enzyme as a
diffusible probe to map changes in nucleosome structure and to
footprint factor interactions at cis-regulatory sequences. In a second
application, termed targeted gene methylation (TAGM), the DMTase is
targeted via in-frame fusion to a DNA-binding factor. The rapid
accumulation of DNA methylation enables highly sensitive detection of
factor binding. Both strategies can be applied with any C5 DMTase, such
as M.SssI, which also possesses a short-recognition specificity (CG). A
description of methods for constructing C5 DMTase-expressing strains of
Saccharomyces cerevisiae and analyzing chromatin regions is provided.
We also include comprehensive protocols for the isolation and bisulfite
treatment of genomic DNA as well as the subsequent bisulfite sequencing
steps. Data demonstrating the efficacy of both DMTase probing
techniques, theoretical considerations, and experimental analyses are
presented at GAL1 and PHO5.

<>

<1>Jett, S.D., Bear, D.G.
<2>Snapshot blotting: transfer of nucleic acids and nucleoprotein complexes from electrophoresis gels to grids for electron microscopy.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>6870-6874
<6>1994
<7>We present a technique, "snapshot blotting", for the electrophoretic transfer of nucleic acids
and nucleoprotein complexes in gel electrophoresis bands onto highly stable carbon film-coated
grids for imaging by electron microscopy.  The method permits structural analysis of
macromolecular species that have been resolved by a gel mobility-shift assay.  To demonstrate
the efficiency and integrity of the transfer process for a multiprotein-DNA assembly, we have
imaged various species of prokaryotic transcription complex, using the cleavage-defective
EcoRI(Q111) protein as an orientation marker and as a blockade of transcription elongation.
Snapshot blotting should be of great utility in the structural characterization of nucleic
acids and protein-nucleic acid interactions.

<>

<1>Jeukens, J., Boyle, B., Bianconi, I., Kukavica-Ibrulj, I., Tummler, B., Bragonzi, A., Levesque, R.C.
<2>Complete Genome Sequence of Persistent Cystic Fibrosis Isolate Pseudomonas aeruginosa Strain RP73.
<3>Genome Announcements
<4>1
<5>e00568-13
<6>2013
<7>Pseudomonas aeruginosa can establish lifelong chronic airway infections in cystic fibrosis
(CF) patients. However, the genetic features associated with long-term
persistence in the lung are not understood. We sequenced the genome of P.
aeruginosa strain RP73, which was isolated after 16.9 years of chronic lung
infection in a CF patient.

<>

<1>Jhingan, A.K.
<2>Microwave modification of biological macromolecules.
<3>International Patent Office
<4>WO 9321344
<5>
<6>1993
<7>A novel method for the enzymatic modification of biological macromolecules is disclosed. The
procedure is rapid, efficient, and suitable for any enzyme-catalyzed reaction. It can be
utilized for restriction endonuclease digestion of DNA samples and facilitates one of the
time-consuming procedures of nucleic acid research. The method is also adaptable for automated
procedures in biological research.

<>

<1>Jhon, N.-I., Casas-Finet, J.R., Maki, A.H., Modrich, P.
<2>Investigation of the complexes of EcoRI endonuclease with decanucleotides containing canonical and modified recognition sequences using fluorescence and optical detection of magnetic resonance spectroscopy.
<3>Biochim. Biophys. Acta
<4>949
<5>189-194
<6>1988
<7>The binding of EcoRI endonuclease to the oligonucleotides d(GCGAATTCGC) and
d(GCGAA) (5BrdU) (5BrdU) d(CGC) has been investigated to determine whether
stacking interactions occur between tryptophan residues and the DNA bases.
Fluorescence binding isotherms show that the decamer containing the canonical
and that containing the modified recognition sequence bind with comparable
affinity.  Optically detected magnetic resonance spectra show limited
perturbations of the Trp zero-field splitting parameters, which are assigned to
electrical field effects.  No evidence for Trp stacking interactions has been
found.

<>

<1>Ji, B., Gimenez, G., Barbe, V., Vacherie, B., Rouy, Z., Amrani, A., Fardeau, M.L., Bertin, P., Alazard, D., Leroy, S., Talla, E., Ollivier, B., Dolla, A., Pradel, N.
<2>Complete Genome Sequence of the Piezophilic, Mesophilic, Sulfate-Reducing Bacterium Desulfovibrio hydrothermalis AM13(T.).
<3>Genome Announcements
<4>1
<5>e00226-12
<6>2013
<7>AM13 is a piezophilic, mesophilic, hydrogenotrophic sulfate-reducing bacterium collected from
a deep-sea hydrothermal chimney on the East Pacific Rise (2,600 m
depth, 13 degrees N). We report the genome sequence of this bacterium, which
includes a 3,702,934-bp chromosome and a circular plasmid of 5,328 bp.

<>

<1>Jia, D.
<2>Crystallographic and biochemical studies of mammalian de novo DNA methyltransferase Dnmt3.
<3>Ph.D. Thesis, Emory Univ., Atlanta, GA, USA
<4>
<5>1-206
<6>2007
<7>The mammalian DNA methyltransferase 3 (Dnmt3) family is responsible for de novo genomic
methylation and consists of two active methyltransferases, Dnmt3a and Dnmt3b, and their
homolog Dnmt3L. Structural and biochemical characterization have identified F261 of human
DNMT3L as a key residue important for its homo-dimerization and its hetero-dimerization with
Dnmt3a. This study provides a functional assessment for the tail-to-tail interface and key
residues observed in the crystal structure of DNMT3L. A structure of the C-terminal domain of
Dnmt3L in complex with the catalytic domain of active Dnmt3a has been determined in the
presence of methyl donor analog AdoHcy. The complex structure showed that the heterodimer of
Dnm3a-3L further dimerizes through Dnmt3a-3a interaction, forming a tetrameric enzyme complex
with two active sites. Substitution of key residues from Dnmt3a-3a or Dnmt3a-3L interfaces
eliminated the enzymatic activity of Dnmt3a. The functional implication of the molecular
architecture of the Dnmt3a-Dnmt3L tetramer with two active sites is being proposed. This
structure is the first of a genuine mammalian DNA methyltransferase as well as the first of
any methyltransferase in complex with a regulator protein. This study indicates the complexity
of mammalian Dnmt in comparison with bacterial enzymes in term of oligomeric state, substrate
recognition and functional regulation.

<>

<1>Jia, D., Jurkowska, R.Z., Zhang, X., Jeltsch, A., Cheng, X.
<2>Structure of Dnmt3a bound to Dnmt3L suggests a model for de novo DNA methylation.
<3>Nature
<4>449
<5>248-251
<6>2007
<7>Genetic imprinting, found in flowering plants and placental mammals, uses DNA methylation to
yield gene expression that is dependent on the parent of origin. DNA methyltransferase 3a
(Dnmt3a) and its regulatory factor, DNA methyltransferase 3-like protein (Dnmt3L), are both
required for the de novo DNA methylation of imprinted genes in mammalian germ cells. Dnmt3L
interacts specifically with unmethylated lysine 4 of histone H3 through its amino-terminal PHD
(plant homeodomain)-like domain. Here we show, with the use of crystallography, that the
carboxy-terminal domain of human Dnmt3L interacts with the catalytic domain of Dnmt3a,
demonstrating that Dnmt3L has dual functions of binding the unmethylated histone tail and
activating DNA methyltransferase. The complexed C-terminal domains of
Dnmt3a and Dnmt3L showed further dimerization through Dnmt3a-Dnmt3a
interaction, forming a tetrameric complex with two active sites.
Substitution of key non-catalytic residues at the Dnmt3a-Dnmt3L interface
or the Dnmt3a-Dnmt3a interface eliminated enzymatic activity. Molecular
modelling of a DNA-Dnmt3a dimer indicated that the two active sites are
separated by about one DNA helical turn. The C-terminal domain of Dnmt3a
oligomerizes on DNA to form a nucleoprotein filament. A periodicity in the
activity of Dnmt3a on long DNA revealed a correlation of methylated CpG
sites at distances of eight to ten base pairs, indicating that
oligomerization leads Dnmt3a to methylate DNA in a periodic pattern. A
similar periodicity is observed for the frequency of CpG sites in the
differentially methylated regions of 12 maternally imprinted mouse genes.
These results suggest a basis for the recognition and methylation of
differentially methylated regions in imprinted genes, involving the
detection of both nucleosome modification and CpG spacing.

<>

<1>Jiang, B., Cui, D., Li, A., Gai, Z., Ma, F., Yang, J., Ren, N.
<2>Genome Sequence of a Cold-Adaptable Sulfamethoxazole-Degrading Bacterium, Pseudomonas psychrophila HA-4.
<3>J. Bacteriol.
<4>194
<5>5721
<6>2012
<7>Pseudomonas psychrophila HA-4 is a cold-adaptable, sulfamethoxazole-degrading bacterium. The
genes related to its cold adaptation mechanism and
sulfamethoxazole metabolism were unknown. We present the draft genome of strain
HA-4. It could provide further insight into the sulfamethoxazole-degrading
mechanism of strain HA-4.

<>

<1>Jiang, B., Yao, H., Tong, Y., Yang, X., Huang, Y., Jiang, J., Cao, W.
<2>Genome Sequence of Borrelia garinii Strain NMJW1, Isolated from China.
<3>J. Bacteriol.
<4>194
<5>6660-6661
<6>2012
<7>We announce the draft genome sequence of Borrelia garinii strain NMJW1, isolated  from Ixodes
persulcatus in northeastern China. The 902,789-bp linear chromosome
(28.4% GC content) contains 813 open reading frames, 33 tRNAs, and 4 complete
rRNAs.

<>

<1>Jiang, B.H., Liu, J.L., Hu, X.M.
<2>Draft Genome Sequence of the Efficient Bioflocculant-Producing Bacterium Paenibacillus sp. Strain A9.
<3>Genome Announcements
<4>1
<5>e00131-13
<6>2013
<7>Paenibacillus sp. strain A9 is an important bioflocculant-producing bacterium, isolated from a
soil sample, and is pale pink-pigmented, aerobic, and
Gram-positive. Here, we report the draft genome sequence and the initial findings
from a preliminary analysis of strain A9, which is a novel species of
Paenibacillus.

<>

<1>Jiang, C., Yan, C.Y., Huang, C., Jiang, J.H., Yu, R.Q.
<2>A bioluminescence assay for DNA methyltransferase activity based on methylation-resistant cleavage.
<3>Anal. Biochem.
<4>423
<5>224-228
<6>2012
<7>DNA methyltransferase (MTase) is a kind of important regulatory factor in various biological
processes. Current methods to investigate DNA
MTase activity are still limited in the sensitivity and/or generality.
Therefore, developing methods with high sensitivity and improved
generality is needed. Here, we develop a new bioluminescence strategy
based on methylation-resistant cleavage and protein expression in vitro
to detect DNA MTase activity. In the strategy, Dam MTase was used as a
model enzyme and Mbol as the methylation-resistant endonuclease, and
luciferase reporter DNA (LR-DNA) was used as their action target.
Because the completely methylated LR-DNA could be expressed as
detectable luciferase, Dam MTase activity was quantified by measuring
the luminescence intensity of the expressed luciferase. The assay
provides a very low detection limit (0.08 U/ml) as well as a wide
linear range (0.2-100 U/ml). Besides, the analysis mode has improved
generality and could be extended to the detection of other DNA MTases
and the corresponding inhibitor screening.

<>

<1>Jiang, C.-Z., Heard, J.E., Ratcliffe, O., Crellman, R.A., Riechmann, J.L., Haake, V.
<2>Polynucleotides and polypeptides that confer increased biomass and tolerance to cold, water deprivation and low nitrogen to plants.
<3>US Patent Office
<4>US 7196245 B
<5>
<6>2007
<7>The invention relates to plant transcription factor polypeptides, polynucleotides that encode
them, homologs from a variety of plant species, and methods of using the polynucleotides and
polypeptides to produce transgenic plants having advantageous properties compared to a
reference plant.  These properties include increased biomass and increased tolerance to cold,
water deprivation, and low nitrogen conditions.

<>

<1>Jiang, H., Fan, H.J., Lu, C.P.
<2>Identification and distribution of putative virulent genes in strains of Streptococcus suis serotype 2.
<3>Vet. Microbiol.
<4>133
<5>309-316
<6>2009
<7>In order to identify gene sequences unique to the virulent strains,
suppression subtractive hybridization (SSH) was conducted using virulent
Streptococcus suis type 2 (SS2) strain HA9801 and avirulent S. suis type 2
strain T15. Thirty genomic regions were absent in T15, and the DNA
sequences of these regions in HA9801 were determined. These DNA fragments,
containing putative virulence genes, encoded 28 proteins that were
homologous to proteins involved in various aspects of cellular surface
structure, molecular synthesis, energy metabolism, regulation, transport
systems and others of unknown function. According to the published SS2
genomic sequence of the Chinese strain 98HAH33, PCR primers for 14
significant DNA fragments were designed and used for detection of the
distribution of these fragments in S. suis strains from different sources,
serotypes, regions, groups and times. The results showed that these 14 DNA
fragments were widely distributed in 37 detected SS2 strains, yet were
absent among the avirulent strain T15. Moreover, these fragments could be
detected in other serotypes of S. suis, but each serotype had a different
distribution of the fragments.

<>

<1>Jiang, H., Zou, G., Huang, L., Zhu, R.
<2>Purification of restriction endonuclease Bsp63I by two step affinity chromatography.
<3>Xinan Shifan Daxue Xuebao, Ziran Kexueban
<4>25
<5>74-77
<6>2000
<7>Restriction endonuclease Bsp63I has been isolated and purified by affinity chromatography on
self-made DNA Sepharose4B and Cibacron Blue F3GA Sepharose 4B from Bacillus sphaericus 63.
The purified enzyme was found to be homogenous as judged by polyacrylamide gel
electrophoresis.  The specific activity of the enzyme is greater than 61400 units per mg
protein and the yield of the purified enzyme is greater than 130 units per gram wet cells.

<>

<1>Jiang, J., Alvarez, C., Kukutla, P., Yu, W., Xu, J.
<2>Draft Genome Sequences of Enterobacter sp. Isolate Ag1 from the Midgut of the Malaria Mosquito Anopheles gambiae.
<3>J. Bacteriol.
<4>194
<5>5481
<6>2012
<7>An isolate of Enterobacter sp. was obtained from the microbial community within the gut of the
Anopheles gambiae mosquito, a major malaria vector in Africa. This
genome was sequenced and annotated. The genome sequences will facilitate
subsequent efforts to characterize the mosquito gut microbiome.

<>

<1>Jiang, K. et al.
<2>Complete genome sequence of Thauera aminoaromatica strain MZ1T.
<3>Standards in Genomic Sciences
<4>6
<5>325-335
<6>2012
<7>Thauera aminoaromatica strain MZ1T, an isolate belonging to genus Thauera, of the family
Rhodocyclaceae and the class the Betaproteobacteria, has been
characterized for its ability to produce abundant exopolysaccharide and degrade
various aromatic compounds with nitrate as an electron acceptor. These
properties, if fully understood at the genome-sequence level, can aid in
environmental processing of organic matter in anaerobic cycles by
short-circuiting a central anaerobic metabolite, acetate, from microbiological
conversion to methane, a critical greenhouse gas. Strain MZ1T is the first strain
from the genus Thauera with a completely sequenced genome. The 4,496,212 bp
chromosome and 78,374 bp plasmid contain 4,071 protein-coding and 71 RNA genes,
and were sequenced as part of the DOE Community Sequencing Program CSP_776774.

<>

<1>Jiang, K., Zheng, J., Higgins, S.B.
<2>A generic algorithm for finding restriction sites within DNA sequences.
<3>Comput. Appl. Biosci.
<4>7
<5>249-256
<6>1991
<7>This paper describes a generic algorithm for finding restriction sites within
DNA sequences.  The generality of the algorithm is made possible through the
use of set theory.  Basic elements of DNA sequences, i.e. nucleotides (bases),
are represented in sets, and DNA sequences, whether specific, ambiguous or even
protein-coding, are represented as sequences of those sets.  The set
intersection operation demonstrates its ability to perform pattern-matching
correctly on various DNA sequences.  The performance analysis showed that the
degree of complexity of the pattern matching is reduced from exponential to
linear.  An example is given to show the actual and potential restriction
sites, derived by the generic algorithm, in the DNA sequence template coding
for a synthetic calmodulin.

<>

<1>Jiang, L., Lin, M., Li, X., Cui, H., Xu, X., Li, S., Huang, H.
<2>Genome Sequence of Thermus thermophilus ATCC 33923, a Thermostable Trehalose-Producing Strain.
<3>Genome Announcements
<4>1
<5>e00493-13
<6>2013
<7>Thermus thermophilus ATCC 33923 contains a thermostable enzyme that can efficiently catalyze
the conversion of maltose into trehalose. Here we report a
2.15-Mb assembly of its genome sequence and other useful information, including
the coding sequences (CDS) responsible for biological processes such as DNA
replication, DNA repair, and RNA maturation.

<>

<1>Jiang, L., Long, M., Shao, Z.
<2>Draft Genome Sequence of Defluviimonas indica Strain 20V17T, Isolated from a Deep-Sea Hydrothermal Vent Environment in the Southwest Indian Ocean.
<3>Genome Announcements
<4>2
<5>e00479-14
<6>2014
<7>Here, we present the draft genome sequence of Defluviimonas indica 20V17(T), which was
isolated from a deep-sea hydrothermal vent chimney sample in the
southwest Indian Ocean. The draft genome sequence contains 4,268,338 bp, with a
G+C content of 66.33%.

<>

<1>Jiang, L., Zhu, L., Xu, X., Li, Y., Li, S., Huang, H.
<2>Genome Sequence of Clostridium tyrobutyricum ATCC 25755, a Butyric Acid-Overproducing Strain.
<3>Genome Announcements
<4>1
<5>e00308-13
<6>2013
<7>Clostridium tyrobutyricum ATCC 25755 is an efficient producer of butyric acid. Here we report
a 3.01-Mb assembly of its genome sequence and other useful
information, including the coding sequences (CDSs) responsible for an alternative
pathway leading to acetate synthesis as well as a series of membrane transport
systems.

<>

<1>Jiang, S., Zheng, B., Ding, W., Lv, L., Ji, J., Zhang, H., Xiao, Y., Li, L.
<2>Whole-Genome Sequence of Staphylococcus hominis, an Opportunistic Pathogen.
<3>J. Bacteriol.
<4>194
<5>4761-4762
<6>2012
<7>Staphylococcus hominis is a commensal coagulase-negative species of staphylococci. It has been
considered a presumptive and opportunistic pathogen
that causes nosocomial infections in humans. Here we present the draft genome
sequence of S. hominis ZBW5, a multidrug-resistant strain isolated from a human
skin sample, which provides opportunities to understand the mechanism and genetic
basis of its pathogenesis.

<>

<1>Jiang, T., Gao, C., Su, F., Zhang, W., Hu, C., Dou, P., Zheng, Z., Tao, F., Ma, C., Xu, P.
<2>Genome Sequence of Pseudomonas stutzeri SDM-LAC, a Typical Strain for Studying the Molecular Mechanism of Lactate Utilization.
<3>J. Bacteriol.
<4>194
<5>894-895
<6>2012
<7>Pseudomonas stutzeri SDM-LAC is an efficient lactate utilizer with various applications in
biocatalysis. Here we present a 4.2-Mb assembly of its
genome. The annotated four adjacent genes form a lactate utilization
operon, which could provide further insights into the molecular mechanism
of lactate utilization.

<>

<1>Jiang, X., Wang, S., Cheng, H., Huo, Y., Zhang, X., Zhu, X., Han, X., Ni, P., Wu, M.
<2>Genome Sequence of Halobiforma lacisalsi AJ5, an Extremely Halophilic Archaeon Which Harbors a bop Gene.
<3>J. Bacteriol.
<4>193
<5>7023-7024
<6>2011
<7>The draft genome sequence (4,398,155 bp, with 65.35% G+C content) of Halobiforma lacisalsi
AJ5, an extremely halophilic archaeon isolated from
a salt lake, is reported here. This is the first genome report for a
species of the Halobiforma genus.

<>

<1>Jiang, X., Xue, Y., Wang, L., Yu, B., Ma, Y.
<2>Genome Sequence of a Novel Polymer-Grade L-Lactate-Producing Alkaliphile, Exiguobacterium sp. Strain 8-11-1.
<3>Genome Announcements
<4>1
<5>e00616-13
<6>2013
<7>Exiguobacterium sp. strain 8-11-1 is a newly isolated alkaliphile, which was reported to
efficiently produce l-lactate using NaOH as the neutralizing agent.
Here, we present the first 2.9-Mb assembly of its genome sequence, which may
provide useful information related to its efficient lactate production and sodium
ion tolerance capacities.

<>

<1>Jiang, Y., Xiao, P., Yu, G., Sano, T., Pan, Q., Li, R.
<2>Molecular Basis and Phylogenetic Implications of Deoxycylindrospermopsin Biosynthesis in the Cyanobacterium Raphidiopsis curvata.
<3>Appl. Environ. Microbiol.
<4>78
<5>2256-2263
<6>2012
<7>New insights into the distribution and biochemistry of the cyanotoxin
cylindrospermopsin (CYN) have been provided by the recent determination of its
biosynthesis gene cluster (cyr) in several cyanobacterial species. Raphidiopsis
curvata CHAB1150 isolated from China was analyzed for CYN analogues. Only
7-deoxy-CYN was detected in the cell extracts. The cyr gene cluster of R. curvata
CHAB1150 was sequenced, and the cyr genes of this strain were found to have
extremely high similarities (96% to 100%) to those from other nostocalean
species. These species include Cylindrospermopsis raciborskii AWT205,
Aphanizomenon sp. strain 10E6, and Aphanizomenon ovalisporum ILC-146. Insertion
mutation was identified within the cyrI gene, and transcripts of cyrI and another
functional gene cyrJ were detected in R. curvata CHAB1150. General congruence
between the phylogenetic trees based on both cyr and 16S rrn was displayed.
Neutral evolution was found on the whole sequences of the cyr genes, and 0 to 89
negative selected codons were detected in each gene. Therefore, the function of
CyrI is to catalyze the oxygenation of 7-deoxy-CYN in CYN biosynthesis. The
transcripts of the mutated cyrI gene may result from polycistronic transcription.
The high conservation of the cyr genes may be ascribed to purifying selection and
horizontal gene transfer.

<>

<1>Jiang, Y., Xu, H., Li, Y., Liu, H., Yu, L., Qiao, M., Liu, G.
<2>Draft Genome Sequence of Bacillus subtilis Strain NKYL29, an Antimicrobial-Peptide-Producing Strain from Soil.
<3>Genome Announcements
<4>2
<5>e01140-14
<6>2014
<7>Bacillus subtilis strain NKYL29 is an antimicrobial-peptide-producing strain isolated from the
soil of Ranzhuang Tunnel in Hebei Province, China. Here, we
present the draft genome of this strain, which provides the genetic basis for
application of the antimicrobial peptide.

<>

<1>Jiang, Y., Yang, F., Zhang, X., Yang, J., Chen, L., Yan, Y., Nie, H., Xiong, Z., Wang, J., Dong, J., Xue, Y., Xu, X., Zhu, Y., Chen, S., Jin, Q.
<2>The complete sequence and analysis of the large virulence plasmid pSS of Shigella sonnei.
<3>Plasmid
<4>54
<5>149-159
<6>2005
<7>The complete sequence of pSS, which is the large virulence plasmid of Shigella sonnei, was
determined. The 214-kb plasmid is composed of
segments of virulence-associated genes, the O-antigen gene clusters, a
range of replication and maintenance genes, and large numbers of insertion
sequence (IS) elements. Two hundred and forty-one open reading frames
(ORFs) were identified, of which 117 are highly homologous to IS elements
or transposases, 57 are homologous to known pathogenesis-associated
proteins, and 30 are related to replication, plasmid maintenance, or other
metabolic functions. Thirty-seven ORFs have no similarity to proteins with
a known function, including two with no significant similarity to any
hypothetical proteins. Interestingly, 10 ORFs encoding O-antigen gene
clusters were identified on the plasmid and this is markedly different
from most other Shigella spp. virulent plasmids. A novel toxin-antitoxin
system, a series of stbDE homologs, was found on the plasmid immediately
downstream of the replication region; the sole segregation stability
system may be responsible for the instability of pSS. The pSS plasmid is a
mixture of genes with different origins and functions. The sequence
suggests a remarkable history of IS-mediated recombination and acquisition
of DNA across a range of bacterial species.

<>

<1>Jiang, Y., Yu, D.L., Wei, Z.Q., Shen, P., Zhou, Z.H., Yu, Y.S.
<2>Complete Nucleotide Sequence of Klebsiella pneumoniae Multidrug Resistance Plasmid pKP048, Carrying bla(KPC-2), bla(DHA-1), qnrB4, and armA.
<3>Antimicrob. Agents Chemother.
<4>54
<5>3967-3969
<6>2010
<7>The Klebsiella pneumoniae multidrug resistance plasmid pKP048 was completely sequenced. This
plasmid carries several important resistance
determinants, such as bla(KPC-2), bla(DHA-1), qnrB4, and armA, which
confer resistance to carbapenems, cephalosporins, fluoroquinolones, and
aminoglycosides, respectively. Analysis of the finished 151,188-bp
sequence data revealed 163 putative genes, 108 of which were assigned
functions such as replication, stable inheritance, antibiotic
resistance, a mobile element, conjugal transfer, and a
restriction-modification system, showing the strong phylogenetic
mosaicism and plasticity of the plasmid.

<>

<1>Jiang, Y.L., Zhang, M., Jing, H.L., Gao, L.Y.
<2>[Isolation and characterization of an iridovirus from sick giant salamander (Andrias davidianus)].
<3>Bing Du Xue Bao
<4>27
<5>274-282
<6>2011
<7>A virus was isolated from cultured sick giant salmander (Andrias davidianus ) in
a farm, Shanxi Province, China. Skin ulceration and necrosis of the distal limbs
are main clinical symptoms. Virus propagated and caused CPE at 10 degrees C to 30
degrees C in BF-2, CO, CHSE, FHM cells. The optimum condition of replication was
in BF-2 cells at 25 degrees C. The virus was proved to be senstive to chloroform,
heat, pH3 and pH10 treatment. Viral replication was inhibited by
5-Fluoro-2-deoxyuridine (FUDR). These results indicated that the virus possessed
an envelope and DNA as the genome. Electron-microscopic observation of
thin-section showed numerous hexagonal viral particles measuring 130 nm to 150 nm
in diameter orderly arranged in a lattice form in cytoplasm of BF-2 cells. The
particles showed typical iridovirus morphology. A 413 bp fragment was amplified
from the viral main capsid protein gene by PCR. The fragments was sequenced and
analysed. The results showed the isolate shared more than 96% nucleotide identity
with some Ranaviruses. We suggested that this virus was named as Andrias
davidianus iridovirus (ADIV) tentatively.

<>

<1>Jiang, Z.F., Xia, F., Johnson, K.W., Bartom, E., Tuteja, J.H., Stevens, R., Grossman, R.L., Brumin, M., White, K.P., Ghanim, M.
<2>Genome Sequences of the Primary Endosymbiont 'Candidatus Portiera aleyrodidarum'  in the Whitefly Bemisia tabaci B and Q Biotypes.
<3>J. Bacteriol.
<4>194
<5>6678-6679
<6>2012
<7>'Candidatus Portiera aleyrodidarum' is the obligate primary endosymbiotic bacterium of
whiteflies, including the sweet potato whitefly Bemisia tabaci, and
provides essential nutrients to its host. Here we report two complete genome
sequences of this bacterium from the B and Q biotypes of B. tabaci.

<>

<1>Jimenez, E., Langa, S., Martin, V., Arroyo, R., Martin, R., Fernandez, L., Rodriguez, J.M.
<2>Complete genome sequence of Lactobacillus fermentum CECT 5716, a probiotic strain isolated from human milk.
<3>J. Bacteriol.
<4>192
<5>4800
<6>2010
<7>Lactobacillus fermentum is a heterofermentative lactic acid bacterium and is frequently
isolated from mucosal surfaces of healthy humans.
Lactobacillus fermentum CECT 5716 is a well-characterized probiotic strain
isolated from human milk and, at present, is used in commercial infant
formulas. Here, we report the complete and annotated genome sequence of
this strain.

<>

<1>Jimenez, E., Martin, R., Maldonado, A., Martin, V., Gomez-de-Segura, A., Fernandez, L., Rodriguez, J.M.
<2>Complete genome sequence of Lactobacillus salivarius CECT 5713, a probiotic strain isolated from human milk and infant feces.
<3>J. Bacteriol.
<4>192
<5>5266-5267
<6>2010
<7>Lactobacillus salivarius is a homofermentative lactic acid bacterium and is frequently
isolated from mucosal surfaces of healthy humans. L. salivarius CECT 5713, a strain isolated
simultaneously from breast milk and infant feces of a healthy mother-infant pair, has
immunomodulatory, anti-inflammatory, and anti-infectious properties, as revealed by several in
vitro and in vivo assays. Here, we report its complete and annotated genome sequence.

<>

<1>Jimenez, E., Villar-Tajadura, M.A., Marin, M., Fontecha, J., Requena, T., Arroyo, R., Fernandez, L., Rodriguez, J.M.
<2>Complete Genome Sequence of Bifidobacterium breve CECT 7263, a Strain Isolated from Human Milk.
<3>J. Bacteriol.
<4>194
<5>3762-3763
<6>2012
<7>Bifidobacterium breve is an actinobacterium frequently isolated from colonic microbiota of
breastfeeding babies. Here, we report the complete and annotated
genome sequence of a B. breve strain isolated from human milk, B. breve CECT
7263. The genome sequence will provide new insights into the biology of this
potential probiotic organism and will allow the characterization of genes related
to beneficial properties.

<>

<1>Jimenez, G., Blanch, A.R., Tamames, J., Rossello-Mora, R.
<2>Complete Genome Sequence of Bacillus toyonensis BCT-7112T, the Active Ingredient  of the Feed Additive Preparation Toyocerin.
<3>Genome Announcements
<4>1
<5>e01080-13
<6>2013
<7>Strain BCT-7112, previously identified as Bacillus cereus var. toyoi, is the type strain of
the species Bacillus toyonensis, a novel species of the B. cereus
group. The complete genome of this strain, which is the active ingredient of the
feed additive preparation Toyocerin, has been sequenced and annotated to reveal
the genetic properties of this probiotic organism with a long history of safe use
in animal nutrition.

<>

<1>Jin, A., Zhang, Y., Xia, Y., Traylor, E., Nelson, M., Van Etten, J.L.
<2>New restriction endonuclease CviRI cleaves DNA at TG^CA sequences.
<3>Nucleic Acids Res.
<4>22
<5>3928-3929
<6>1994
<7>A new type II restriction endonuclease, CviRI, was isolated from virus XZ-6E infected
chlorella cells. CviRI is the first restriction endonuclease to recognize the sequence
5'-TGCA-3' and cleaves DNA between the G and C residues to produce blunt-end termini.
Methylation of the adenine or cytosine in 5'-TGCA-3' sequences prevents CviRI cleavage. Due
to its sequence specificity, CviRI may be especially useful for detecting mutant alleles of
many heritable human genetic diseases.

<>

<1>Jin, B., Robertson, K.D.
<2>DNA Methyltransferases, DNA Damage Repair, and Cancer.
<3>Adv. Exp. Med. Biol.
<4>754
<5>3-29
<6>2013
<7>The maintenance DNA methyltransferase (DNMT) 1 and the de novo methyltransferases DNMT3A and
DNMT3B are all essential for mammalian development. DNA methylation, catalyzed by the DNMTs,
plays an important role in maintaining genome stability. Aberrant expression of DNMTs and
disruption of DNA methylation patterns are closely associated with many forms of cancer,
although the exact mechanisms underlying this link remain elusive. DNA damage repair systems
have evolved to act as a genome-wide surveillance mechanism to maintain chromosome integrity
by recognizing and repairing both exogenous and endogenous DNA insults. Impairment of these
systems gives rise to mutations and directly contributes to tumorigenesis. Evidence is
mounting for a direct link between DNMTs, DNA methylation, and DNA damage repair systems,
which provide new insight into the development of cancer. Like tumor suppressor genes, an
array of DNA repair genes frequently sustain promoter hypermethylation in a variety of tumors.
In addition, DNMT1, but not the DNMT3s, appear to function coordinately with DNA damage repair
pathways to protect cells from sustaining mutagenic events, which is very likely through a DNA
methylation-independent mechanism. This chapter is focused on reviewing the links between DNA
methylation and the DNA damage response.

<>

<1>Jin, D., Chen, C., Li, L., Lu, S., Li, Z., Zhou, Z., Jing, H., Xu, Y., Du, P., Wang, H., Xiong, Y., Zheng, H., Bai, X., Sun, H., Wang, L., Ye, C., Gottschalk, M., Xu, J.
<2>Dynamics of fecal microbial communities in children with diarrhea of unknown etiology and genomic analysis of associated Streptococcus lutetiensis.
<3>BMC Microbiol.
<4>13
<5>141
<6>2013
<7>BACKGROUND: The sequences of the 16S rRNA genes extracted from fecal samples
provide insights into the dynamics of fecal microflora. This potentially gives
valuable etiological information for patients whose conditions have been ascribed
to unknown pathogens, which cannot be accomplished using routine culture methods.
We studied 33 children with diarrhea who were admitted to the Children's Hospital
in Shanxi Province during 2006. RESULTS: Nineteen of 33 children with diarrhea
could not be etiologically diagnosed by routine culture and polymerase chain
reaction methods. Eleven of 19 children with diarrhea of unknown etiology had
Streptococcus as the most dominant fecal bacterial genus at admission. Eight of
nine children whom three consecutive fecal samples were collected had
Streptococcus as the dominant fecal bacterial genus, including three in the
Streptococcus bovis group and three Streptococcus sp., which was reduced during
and after recovery. We isolated strains that were possibly from the S. bovis
group from feces sampled at admission, which were then identified as
Streptococcus lutetiensis from one child and Streptococcus gallolyticus subsp.
pasteurianus from two children. We sequenced the genome of S. lutetiensis and
identified five antibiotic islands, two pathogenicity islands, and five unique
genomic islands. The identified virulence genes included hemolytic toxin cylZ of
Streptococcus agalactiae and sortase associated with colonization of pathogenic
streptococci. CONCLUSIONS: We identified S. lutetiensis and S. gallolyticus
subsp. pasteurianus from children with diarrhea of unknown etiology, and found
pathogenic islands and virulence genes in the genome of S. lutetiensis.

<>

<1>Jin, H.M., Jeong, H., Moon, E.J., Math, R.K., Lee, K., Kim, H.J., Jeon, C.O., Oh, T.K., Kim, J.F.
<2>Complete Genome Sequence of the Polycyclic Aromatic Hydrocarbon-Degrading Bacterium Alteromonas sp. Strain SN2.
<3>J. Bacteriol.
<4>193
<5>4292-4293
<6>2011
<7>Alteromonas sp. strain SN2, able to metabolize polycyclic aromatic hydrocarbons, was isolated
from a crude oil-contaminated sea-tidal flat.
Here we report the complete 4.97-Mb genome sequence and annotation of
strain SN2. These will advance the understanding of strain SN2's
adaptation to the sea-tidal flat ecosystem and its pollutant metabolic
versatility.

<>

<1>Jin, Lu., Ye, F., Zhao, D., Chen, S., Zhu, K., Zheng, M., Jiang, R.-W., Jiang, H., Luo, C.
<2>Metadynamics Simulation Study on the Conformational Transformation of HhaI Methyltransferase: An Induced-Fit Base-Flipping Hypothesis.
<3>Biomed Res. Int.
<4>2014
<5>304563
<6>2014
<7>DNA methyltransferases play crucial roles in establishing and maintenance of DNA methylation,
which is an important epigenetic mark. Flipping the target cytosine out of the DNA helical
stack and into the active site of protein provides DNA methyltransferases with an opportunity
to access and modify the genetic information hidden in DNA. To investigate the conversion
process of base flipping in the HhaI methyltransferase (M. HhaI), we performed different
molecular simulation approaches on M.HhaI-DNA-S-adenosylhomocysteine ternary complex. The
results demonstrate that the nonspecific binding of DNA to M. HhaI is initially induced by
electrostatic interactions. Differences in chemical environment between the major and minor
grooves determine the orientation of DNA. Gln237 at the target recognition loop recognizes the
GCGC base pair from the major groove side by hydrogen bonds. In addition, catalytic loop
motion is a key factor during this process. Our study indicates that base flipping is likely
to be an 'induced-fit' process. This study provides a solid foundation for future studies on
the discovery and development of mechanism-based DNA methyltransferases regulators.

<>

<1>Jin, Q. et al.
<2>Genome sequence of Shigella flexneri 2a: insights into pathogenicity through comparison with genomes of Escherichia coli K12 and O157.
<3>Nucleic Acids Res.
<4>30
<5>4432-4441
<6>2002
<7>We have sequenced the genome of Shigella flexneri serotype 2a, the most prevalent species and
serotype that causes bacillary dysentery or shigellosis in man. The whole genome is composed
of a 4 607 203 bp chromosome and a 221 618 bp virulence plasmid, designated pCP301. While the
plasmid shows minor divergence from that sequenced in serotype 5a, striking characteristics of
the chromosome have been revealed. The S.flexneri chromosome has, astonishingly, 314 IS
elements, more than 7-fold over those possessed by its close relatives, the non-pathogenic K12
strain and enterohemorrhagic O157:H7 strain of Escherichia coli. There are 13 translocations
and inversions compared with the E.coli sequences, all involve a segment larger than 5 kb, and
most are associated with deletions or acquired DNA sequences, of which several are likely to
be bacteriophage-transmitted pathogenicity islands. Furthermore, S.flexneri, resembling
another human-restricted enteric pathogen, Salmonella typhi, also has hundreds of pseudogenes
compared with the E.coli strains. All of these could be subjected to investigations towards
novel preventative and treatment strategies against shigellosis.

<>

<1>Jin, S.G., Kadam, S., Pfeifer, G.P.
<2>Examination of the specificity of DNA methylation profiling techniques towards 5-methylcytosine and 5-hydroxymethylcytosine.
<3>Nucleic Acids Res.
<4>38
<5>e125
<6>2010
<7>DNA cytosine-5 methylation is a well-studied epigenetic pathway implicated in gene expression
control and disease pathogenesis. Different
technologies have been developed to examine the distribution of
5-methylcytosine (5mC) in specific sequences of the genome. Recently,
substantial amounts of 5-hydroxymethylcytosine (5hmC), most likely derived
from enzymatic oxidation of 5mC by TET1, have been detected in certain
mammalian tissues. Here, we have examined the ability of several commonly
used DNA methylation profiling methods to distinguish between 5mC and
5hmC. We show that techniques based on sodium bisulfite treatment of DNA
are incapable of distinguishing between the two modified bases. In
contrast, techniques based on immunoprecipitation with anti-5mC antibody
(methylated DNA immunoprecipitation, MeDIP) or those based on proteins
that bind to methylated CpG sequences (e.g. methylated-CpG island recovery
assay, MIRA) do not detect 5hmC and are specific for 5mC unless both
modified bases occur in the same DNA fragment. We also report that several
methyl-CpG binding proteins including MBD1, MBD2 and MBD4 do not bind to
sequences containing 5hmC. Selective mapping of 5hmC will require the
development of unique tools for the detection of this modified base.

<>

<1>Jin, Y., Binkowski, G., Simon, L.D., Norris, D.
<2>HO endonuclease cleaves MAT DNA in vitro by an inefficient stoichiometric reaction mechanism.
<3>J. Biol. Chem.
<4>272
<5>7352-7359
<6>1997
<7>Mating type switching in Saccharomyces cerevisiae initiates when HO endonuclease makes a
double-stranded DNA break at the yeast MAT locus.  In this report, we characterize the
fundamental biochemical properties of HO.  Using an assay that monitors cleavage of a MAT
plasmid, we define an optimal in vitro reaction, showing in particular that the enzyme has a
stringent requirement for zinc ions.  This suggests that zinc finger motifs present in HO are
important for cleavage.  The most unexpected feature of HO, however, is its extreme
inefficiency.  Maximal cleavage occurs when HO is present at a concentration of 1 molecule/3
base pairs of substrate DNA.  Even under these conditions, complete digestion requires >2h.
This inefficiency results from two characteristics of HO.  First, HO recycles slowly from
cleaved product to new substrate, in part because the enzyme has an affinity for one end of
its double strand break product.  Second, high levels of cleavage in the in vitro reaction
correlate with the appearance of large protein-DNA aggregates.  At optimal HO concentrations,
these latter aggregates, referred to as "florettes," have an ordered structure consisting of a
densely staining central region and loops of radiating DNA.  These unusual properties may
indicate that HO plays a role in other aspects of mating type switching subsequent to double
strand break formation.

<>

<1>Jindrova, E., Schmid-Nuoffer, S., Hamburger, F., Janscak, P., Bickle, T.A.
<2>On the DNA cleavage mechanism of Type I restriction enzymes.
<3>Nucleic Acids Res.
<4>33
<5>1760-1766
<6>2005
<7>Although the DNA cleavage mechanism of Type I restriction-modification enzymes has been
extensively studied, the mode of cleavage remains
elusive. In this work, DNA ends produced by EcoKI, EcoAI and EcoR124I,
members of the Type IA, IB and IC families, respectively, have been
characterized by cloning and sequencing restriction products from the
reactions with a plasmid DNA substrate containing a single recognition
site for each enzyme. Here, we show that all three enzymes cut this
substrate randomly with no preference for a particular base composition
surrounding the cleavage site, producing both 5'- and 3'-overhangs of
varying lengths. EcoAI preferentially generated 3'-overhangs of 2-3 nt,
whereas EcoKI and EcoR124I displayed some preference for the formation of
5'-overhangs of a length of approximately 6-7 and 3-5 nt, respectively. A
mutant EcoAI endonuclease assembled from wild-type and nuclease-deficient
restriction subunits generated a high proportion of nicked circular DNA,
whereas the wild-type enzyme catalyzed efficient cleavage of both DNA
strands. We conclude that Type I restriction enzymes require two
restriction subunits to introduce DNA double-strand breaks, each providing
one catalytic center for phosphodiester bond hydrolysis. Possible models
for DNA cleavage are discussed.

<>

<1>Jindrova, E., Schmid-Nuoffer, S., Hamburger, F., Janscak, P., Bickle, T.A.
<2>On the DNA cleavage mechanism of Type I restriction enzymes.
<3>FEBS J.
<4>272
<5>552
<6>2005
<7>Although the DNA-cleavage mechanism of Type I restriction-modification enzymes has been
extensively studied, the mode of how these enzymes introduce DNA double-strand breaks still
remains elusive. In this work, DNA ends produced by EcoKI, EcoAI and EcoR124I, members of the
Type IA, IB and IC families, respectively, have been characterized by cloning and sequencing
of restriction products from reactions with a plasmid DNA substrate containing a single
recognition site for each enzyme. We show that all three enzymes cut this DNA randomly with no
preference for a particular base composition surrounding the cleavage site, producing both 5'-
and 3'-overhangs of varying lengths. EcoAI preferentially generated 3''-overhangs of 2-3
nucleotides, whereas EcoKI and EcoR124I displayed some preference for formation of
5'-overhangs in a length of about 6-7 and 3-5 nucleotides, respectively. A mutant EcoAI
endonuclease assembled from wild-type and nuclease-deficient restriction subunits generated a
high proportion of nicked circular DNA, whereas the wild-type enzyme catalyzed efficient
cleavage of both DNA strands. We conclude that Type I restriction enzymes require two
restriction subunits to introduce DNA double-strand breaks, each providing one catalytic
center for phosphodiester bond hydrolysis. Possible models for DNA cleavage are further
studied and discussed.

<>

<1>Jing, X., Cao, X., Wang, Li., Lan, T., Li, Y., Xie, G.
<2>DNA-AuNPs based signal amplification for highly sensitive detection of DNA methylation, methyltransferase activity and inhibitor screening.
<3>Biosensors and Bioelectronics
<4>58
<5>40-47
<6>2014
<7>A sensitive and selective electrochemical method was developed for the detection of DNA
methylation, determination of DNA methyltransferase (MTase) activity and screening of MTase
inhibitor. Methylene blue (MB) was employed as electrochemical indicator and DNA-modified gold
nanoparticles (AuNPs) were used as signal amplification unit because the DNA strands in this
composite have strong adsorption ability for MB. First, the thiolated single-stranded DNA S1
was self-assembled on gold electrode, hybridization between the lower portion of DNA S1 and
its complementary DNA S2 formed an identical double-stranded tetranucleotide target sequence
for both DNA adenine methylation (Dam) MTase and methylation-resistant endonuclease Mbo I,
then the upper portion of DNA S1 was hybridized with its complementary DNA S3 modified on
AuNPs to bring the DNA S3-AuNPs amplification units onto the electrode. The DNA S1
/S2/S3-AuNPs bioconjugate has lots of DNA strands, and they can adsorb abundant MB. Mbo I
endounuclease could not cleave the identical target sequence after it was methylated by Dam
MTase. On the contrary, the sequence without methylation could be cleaved, which would
decrease the amount of adsorbed MB. The presence of redox-active MB was detected
electrochemically by differential pulse voltammetry (DPV). Thus, the activity of Dam MTase and
methylation status were sensitively converted to the DNA S3-AuNPs amplified DPV signals. The
DPV signal demonstrated a linear relationship with logarithm of Dam concentration ranging from
0.075 to 30 U/mL, achieving a detection limit of 0.02 U/mL (SIN=D3). Also, screening of Dam
MTase inhibitor 5-fluorouracil was successfully investigated using this fabricated sensor.

<>

<1>Jiricny, J., Martin, D.
<2>Restriction endonucleases HindII and TaqI cleave DNA with mismatched nucleotides within their recognition sequences.
<3>Nucleic Acids Res.
<4>14
<5>1943-1949
<6>1986
<7>Restriction endonucleases HindII and TaqI, but not SalI, were found to
efficiently cleave synthetic hexdecanucleotide duplexes which contained either
an A/C or a G/T mismatch within their respective restriction sites.
Double-stranded M13 DNAs with identical mismatches were also cleaved under the
assay conditions.  These results suggest that the distortion of the DNA duplex,
caused by these purine/pyrimidine mismatches is not sufficiently large so as to
interfere with the recognition and the subsequent cleavage of the DNA by these
two enzymes.  HindII and SalI, but not TaqI, were furthermore shown to
hydrolyze the two strands of the duplex with different rates.  The differences
between the mode of recognition of their respective restriction sites by these
three enzymes are discussed.

<>

<1>Jiricny, J., Wood, S.G., Martin, D., Ubasawa, A.
<2>Oligonucleotide duplexes containing inosine, 7-deazainosine, tubercidin, nebularine and 7-deazanebularine as substrates for restriction endonucleases HindII, SalI and TaqI.
<3>Nucleic Acids Res.
<4>14
<5>6579-6590
<6>1986
<7>Synthetic hexadecanucleotide duplexes containing a single purine nucleotide
analogue in the recognition sites of the restriction endonucleases HindII, SalI
and TaqI were used to investigate the restriction site determinants required by
these enzymes for sequence recognition and phosphodiester bond cleavage.  The
enzymes were, in general, unaffected by changes introduced into the minor
groove of the helix.  SalI was found to be inhibited by the major groove
modifications introduced into the fourth position of its recognition sequence
GTCGAC.  HindII and TaqI were, by contrast, able to cleave the sites containing
the analogues at this position.  TaqI and, to a lesser extent, HindII could
also be shown to tolerate "mismatch analogues" at this site.

<>

<1>Jo, K.
<2>Evolutionary relationship of NaeI restriction endonuclease with DNA topisomerases.
<3>Diss. Abstr.
<4>57
<5>889
<6>1996
<7>NaeI endonuclease must bind two DNA recognition sequences for cleavage
to occur.  Therefore, DNAs containing NaeI recognition sequences without sufficient
affinity to occupy one or the other DNA-binding sites are resistant to cleavage.  Tethering
resistant and cleavable recognition sequences together caused a switch in their relative
cleavabilities.  This switching implies that DNA cleavage occurs at the DNA-binding site
with higher affinity for the resistant sequence.  The other DNA-binding site, with higher
affinity for the cleavable sequence, acts as the effector DNA-binding site with little to no
catalytic function.  Examination of the amino acid sequence of NaeI uncovered similarity to
the active site of human ligase I, except for leucine 43 in NaeI instead of lysine essential
for
ligase activity.  Changing leucine 43 to lysine 43 (L43K) changed NaeI activity: NaeI-
L43K relaxed supercoiled DNA to yield DNA topoisomers and recombined DNA to give
dimeric molecule.  Interruption of the reactions of NaeI and NaeI-L43K with DNA
demonstrated transient protein-DNA covalent complexes.  These findings imply coupled
endonuclease and ligase domains and link NaeI endonuclease to the topoisomerase and
recombinase protein families.  Glycerol gradient sedimentation of NaeI-L43K showed that
the active conformation of NaeI-LA3K is a dimer.  The topoisomerase activity of NaeI-
LA3K changed from processive to distributive with increasing cation concentration.  NaeI-
L43K decatenated k-DNA and bonded the 5' end of the cleaved DNA.  These
characteristics mimic those of the classic type II topoisomerases.  Also, the effects of
topoisomerase drugs on NaeI-LA3K were determined.  NaeI-LA3K activity was
specifically inhibited by eukaryotic topoisomerase II drugs daunorubicin, ellipticine, and
m-AMSA.  Especially, the cleavage step of DNA relaxation by NaeI-LA3K was sensitively
inhibited by these drugs, but the cleavage activity of wild-type NaeI was not.  Therefore,
L43K amino acid change increased the sensitivity of NaeI protein to the drugs.  The
increased sensitivity implies that protein-drug interaction is enhanced by the amino acid
change and that the ligase-like active site, containing LA3K, provides a part of the drug
binding pocket.

<>

<1>Jo, K., Topal, M.D.
<2>DNA topoisomerase and recombinase activities in NaeI restriction endonuclease.
<3>Science
<4>267
<5>1817-1820
<6>1995
<7>NaeI endonuclease must bind to two DNA sequences for cleavage. Examination of the amino acid
sequence of NaeI uncovered similarity to the active site of human DNA ligase I, except for
leucine 43 in NaeI instead of the lysine essential for ligase activity. Changing leucine 43 to
lysine 43 (L43K) changed NaeI activity: NaeI-L43K relaxed supercoiled DNA to yield DNA
topisomers and recombined DNA to give dimeric molecules. Interruption of the reactions of NaeI
and NaeI-L43K with DNA demonstrated transient protein-DNA covalent complexes. These findings
imply coupled endonuclease and ligase domains and link NaeI endonuclease to the topoisomerase
and recombinase protein families.

<>

<1>Jo, K., Topal, M.D.
<2>Effects on NaeI-DNA recognition of the leucine to lysine substitution that transforms restriction endonuclease NaeI to a topoisomerase: a model for restriction endonuclease evolution.
<3>Nucleic Acids Res.
<4>24
<5>4171-4175
<6>1996
<7>Substituting lysine for leucine at position 43 (L43K) transforms NaeI from restriction
endonuclease to topoisomerase and makes NaeI hypersensitive to intercalative anticancer drugs.
Here we investigated DNA recognition by NaeI-L43K.  Using DNA competition and gel retardation
assays, NaeI-L43K showed reduced affinity for DNA substrate and the ability to bind both
single- and double-stranded DNA with a definite preference for the former.  Sedimentation
studies showed that under native conditions NaeI-L43K, like NaeI, is a dimer.  Introduction of
mismatched bases into double-stranded DNA significantly increased that DNA's ability to
inhibit NaeI-L43K.  Wild-type NaeI showed no detectable binding of either single-stranded DNA
or mismatched DNA over the concentration range studied.  These results demonstrate that the
L43K substitution caused a significant change in recognition specificity by NaeI and imply
that NaeI-L43K's topoisomerase activity is related to its ability to bind single-stranded and
distorted regions in DNA.  A mechanism is proposed for the evolution of the NaeI
restriction-modification system from a topoisomerase/ligase by a mutation that abolished
religation activity and provided a needed change in DNA recognition.

<>

<1>Jo, K., Topal, M.D.
<2>Changing a leucine to a lysine residue makes NaeI endonuclease hypersensitive to DNA intercalative drugs.
<3>Biochemistry
<4>35
<5>10014-10018
<6>1996
<7>A single amino acid change transforms restriction enzyme NaeI to a topoisomerase and
recombinase (NaeI-L43K) that shows no sequence similarity to these protein families.  This
transformation appears to result from coupled endonuclease and ligase domains.  To further
elucidate the relationship between NaeI-L43K and the topoisomerase protein family, we studied
the effect of the topoisomerase inhibitors on NaeI-L43K activity.  The intercalative drugs
amsacrine, ellipticine, and daunorubicin inhibited NaeI-L43K, whereas the nonintercalating
drugs camptothecin, VP-16, and oxolinic acid did not.  Ethidium bromide also inhibited
NaeI-L43K, implying that intercalation is responsible for its inhibition.  The effects of the
intercalative drugs on the DNA cleavage steps of NaeI and NaeI-L43K were compared.  The drugs
hardly inhibited DNA cleavage by wild type NaeI but completely inhibited DNA cleavage by
NaeI-L43K.  This difference in inhibition demonstrates that the L43K amino acid change
sensitized NaeI to these drugs.  Low concentrations of the intercalative drugs, except for
ethidium bromide, enhance production of topoisomerase--DNA covalent intermediates but
inhibited production of the NaeI-L43K-DNA covalent intermediate.  These results imply some
unique differences between DNA relaxation by NaeI-L43K and DNA topoisomerase.  Concomitant
with studying inhibition of the cleavage intermediate, NaeI-L53K was found to covalently bond
with the 5' end of the cleaved DNA strand.

<>

<1>Jo, K., Topal, M.D.
<2>Step-wise DNA relaxation and decatenation by NaeI-43K.
<3>Nucleic Acids Res.
<4>26
<5>2380-2384
<6>1998
<7>NaeI protein was originally isolated for its restriction endonuclease properties.  NaeI was
later discovered to either relax or cleave supercoiled DNA, depending upon whether NaeI
position 43 contains a lysine (43K) or leucine (43L) respectively.  NaeI-43K DNA relaxation
activity appears to be the product of coupling separate endonuclease and ligase domains within
the same polypeptide.  Whereas NaeI relaxes supercoiled DNA like a topoisomerase, even forming
a transient covalent intermediate with the substrate DNA, NaeI shows no obvious sequence
similarity to the topoisomerases.  To further characterize the topoisomerase activity of NaeI,
we report here that NaeI-43K changes the linking number of a single negatively supercoiled
topoisomer of pBR322 by units of one and therefore is a type I topoisomerase.  Positively
supercoiled pBR322 was resistant to NaeI-43K.  At low salt concentration NaeI-43K was
processive; non-saturating amounts of enzyme relaxed a fraction of the DNA. At high salt
concentration the same non-saturating amounts of NaeI-43K partially relaxed all the DNA in a
step-wise fashion to give a Gaussian distribution of topoisomers, demonstrating a switch from
a processive to a distributive mode of action.  NaeI-43K decatenated kinetoplast DNA
containing nicked circles, implying that NaeI-43K can cleave opposite a nick.  The products of
the reaction are decatenated nicked circles under both processive and distributive conditions.
The behavior of NaeI-43K is consistent with that of a prokaryotic type I topoisomerase.

<>

<1>Joardar, V. et al.
<2>Whole-genome sequence analysis of Pseudomonas syringae pv. phaseolicola 1448A reveals divergence among pathovars in genes involved in virulence and transposition.
<3>J. Bacteriol.
<4>187
<5>6488
<6>2005
<7>Pseudomonas syringae pv. phaseolicola, a gram-negative bacterial plant pathogen, is the causal
agent of halo blight of bean. In this study, we report on the genome sequence of P. syringae
pv. phaseolicola isolate 1448A, which encodes 5,353 open reading frames (ORFs) on one circular
chromosome (5,928,787 bp) and two plasmids (131,950 bp and 51,711 bp). Comparative analyses
with a phylogenetically divergent pathovar, P. syringae pv. tomato DC3000, revealed a strong
degree of conservation at the gene and genome levels. In total, 4,133 ORFs were identified as
putative orthologs in these two pathovars using a reciprocal best-hit method, with 3,941 ORFs
present in conserved, syntenic blocks. Although these two pathovars are highly similar at the
physiological level, they have distinct host ranges; 1448A causes disease in beans, and DC3000
is pathogenic on tomato and Arabidopsis. Examination of the complement of ORFs encoding
virulence, fitness, and survival factors revealed a substantial, but not complete, overlap
between these two pathovars. Another distinguishing feature between the two pathovars is their
distinctive sets of transposable elements. With access to a fifth complete pseudomonad genome
sequence, we were able to identify 3,567 ORFs that likely comprise the core Pseudomonas genome
and 365 ORFs that are P. syringae specific.

<>

<1>Jobbagy, Z., Izsvak, Z., Duda, E.
<2>Positive co-operative interaction between the subunits of CeqI restriction endonuclease.
<3>Biochem. J.
<4>286
<5>85-88
<6>1992
<7>CeqI restriction endonuclease, an isoschizomer of EcoRV, forms complexes of 2-20 subunits
under physiological conditions, in the absence of DNA. These molecules partially dissociate in
the presence of DNA sequences recognized by CeqI or in the presence of non-ionic detergents.
In solutions containing high concentrations of salts (e.g. 1M NaCl), the enzyme dissociated
into subunits, concomitantly losing its activity. According to our experiments, it is the
tetrameric form of the enzyme that binds the DNA and represents the catalytically active
molecule. Analysis of the enzyme kinetics revealed a positive co-operative interaction between
the subunits of the enzyme. Computer-assisted analysis of these data yielded a Hill
coefficient of approx. 1.35, suggesting two binding sites per tetrameric enzyme molecule, two
subunits per palindromic recognition site.

<>

<1>Jogler, C., Lin, W., Meyerdierks, A., Kube, M., Katzmann, E., Flies, C., Pan, Y., Amann, R., Reinhardt, R., Schuler, D.
<2>Toward cloning of the magnetotactic metagenome: identification of magnetosome island gene clusters in uncultivated magnetotactic bacteria from different aquatic sediments.
<3>Appl. Environ. Microbiol.
<4>75
<5>3972-3979
<6>2009
<7>In this report, we describe the selective cloning of large DNA fragments
from magnetotactic metagenomes from various aquatic habitats. This was
achieved by a two-step magnetic enrichment which allowed the mass
collection of environmental magnetotactic bacteria (MTB) virtually free of
nonmagnetic contaminants. Four fosmid libraries were constructed and
screened by end sequencing and hybridization analysis using heterologous
magnetosome gene probes. A total of 14 fosmids were fully sequenced. We
identified and characterized two fosmids, most likely originating from two
different alphaproteobacterial strains of MTB that contain several
putative operons with homology to the magnetosome island (MAI) of
cultivated MTB. This is the first evidence that uncultivated MTB exhibit
similar yet differing organizations of the MAI, which may account for the
diversity in biomineralization and magnetotaxis observed in MTB from
various environments.

<>

<1>Jogler, C., Niebler, M., Lin, W., Kube, M., Wanner, G., Kolinko, S., Stief, P., Beck, A.J., de Beer, D., Petersen, N., Pan, Y., Amann, R., Reinhardt, R., Schuler, D.
<2>Cultivation-independent characterization of 'Candidatus Magnetobacterium bavaricum' via ultrastructural, geochemical, ecological and metagenomic methods.
<3>Environ. Microbiol.
<4>12
<5>f2466-f2478
<6>2010
<7>'Candidatus Magnetobacterium bavaricum' is unusual among magnetotactic bacteria (MTB) in
terms of cell size (8-10 microm long, 1.5-2 microm in diameter), cell architecture,
magnetotactic behaviour and its distinct phylogenetic position in the deep-branching
Nitrospira phylum. In the present study, improved magnetic enrichment techniques permitted
high-resolution scanning electron microscopy and energy dispersive X-ray analysis, which
revealed the intracellular organization of the magnetosome chains. Sulfur globule accumulation
in the cytoplasm point towards a sulfur-oxidizing metabolism of 'Candidatus M. bavaricum'.
Detailed analysis of 'Candidatus M. bavaricum' microhabitats revealed more complex
distribution patterns than previously reported, with cells predominantly found in low oxygen
concentration. No correlation to other geochemical parameters could be observed. In addition,
the analysis of a metagenomic fosmid library revealed a 34 kb genomic fragment, which contains
33 genes, among them the complete rRNA gene operon of 'Candidatus M. bavaricum' as well as a
gene encoding a putative type IV RubisCO large subunit.

<>

<1>Johannssen, W.
<2>Interaction of restriction endonuclease with DNA as revealed by electron microscopy.
<3>Methods Microbiol.
<4>20
<5>325-339
<6>1988
<7>None

<>

<1>Johannssen, W.
<2>Electron microscopy studies of DNA complexes with restriction endonuclease SalGI.
<3>Z. Allg. Mikrobiol.
<4>23
<5>197-201
<6>1983
<7>The binding properties of the type II restriction endonuclease SalGI to the plasmid DNA pGW10
has been investigated by electron microscopic studies.  Samples were spread by the BAC
technique.  In the presence of magnesium, SalGI binds as dimers and tetramers to the specific
recognition site 5'-G-T-C-G-A-C-3' and with lower rate to the sequence 5'-G-T-C-A-A-C-3',
which represents the recognition site of the restriction endonucleases HindII and HincII.

<>

<1>Johannssen, W., Schutte, H., Mayer, F., Mayer, H.
<2>Quaternary structure of the isolated restriction endonuclease EndoR.BglI from Bacillus globigii as revealed by electron microscopy.
<3>J. Mol. Biol.
<4>134
<5>707-726
<6>1979
<7>The restriction endonuclease EndoR.BglI was purified nearly to homogeneity.
BglI samples, when negatively stained with 4% uranyl acetate, show two
different particle projections in the electron microscope.  Projection A has an
outer diameter of 22.5+/-0.8 nm and is composed of six intensity maxima
arranged in a ring; the centre of the ring exhibits slightly visible additional
substructures.  Projection B is also a ring; its outer diameter is 23.8+/-0.7
nm; it does not show detailed fine structure, aside from a probable 10-fold
rotational symmetry.  Variations of the negative staining technique (single
carbon layer, 2% uranyl acetate; sandwich preparation with 4% uranyl acetate)
revealed additional fine structural details for both projections.  From the
electron microscopic observations, a model of the enzyme particle was developed
containing 20 identical, biologically active monomers of molecular weight
around 61,000 arranged as a pentagonal dodecahedron.  Tilting experiments
established this structure decisively by interconversion of the different
appearances of given particles in the expected way.  By sodium dodecyl
sulphate/polyacrylamide gel electrophoresis, polyacrylamide gel electrophoresis
in a continuous molecular sieve gradient and evaluation of negatively stained
enzyme particles, a molecular weight of the monomer of 61,000 was estimated,
resulting in a total enzyme particle molecular weight of 1.2x10/6 also
determined by linear sucrose density-gradient centrifugation.

<>

<1>Johansen, S., Embley, T.M., Willassen, N.P.
<2>A family of nuclear homing endonucleases.
<3>Nucleic Acids Res.
<4>21
<5>4405
<6>1993
<7>Homing endonucleases from archaea introns, protein insertions, and the mobile group I introns
from organelles share a common motif, the LAGLI-DADG motif. Sequence motifs like the
zinc-finger and the BIY-YIG have been found in phage homing endonucleases. However, the only
described nuclear homing endonuclease, the 18 kd I-Ppo protein has none of the above consensus
sequences. This unclassified I-Ppo endonuclease is encoded by the mobile intron PpLSU3 found
in the extrachromosomal ribosomal DNA (rDNA) of the myxomycete Physarum polycephalum.
Recently, we have discovered two new group I intron elements from nuclear extrachromosomal
rDNA that bears an apparent similarity to PpLSU3. The myxomycete Didymium iridis has a mobile
group I intron that encodes a putative homing endonuclease I-Dir of 29kd, whereas the
amoeba-flagellate Naegleria andersoni ssp andersoni contains a similar intron that encodes a
putative homing endonuclease I-Naa of 28 kd. I-Dir, I-Naa and I-Ppo are all basic proteins
with a theoretical isoelectric point of 9.5, 9.9 and 8.2, respectively. Furthermore, we could
not identify any significant sequence similarity between these proteins and proteins in the
GenBank and EMBL libraries, and they do not contain any of the sequence motifs (eg. LAGLI-DADG
or GIY-YIG) found in other homing endonucleases. However, we have identified a conserved
region among the nuclear homing endonucleases which consists of several His and Cys residues.
Patterns of His and Cys residues have been associated with protein domains involved in
metal-binding and interaction with nucleic acids, suggesting a similar function of the His-Cys
Box. Database searches using the His-Cys box consensus sequence as the test sequence failed to
identify homologous regions in other proteins. Furthermore, no significant sequence similarity
is seen between I-Dir, I-Naa and I-Ppo outside this His-Cys Box (less than 10% identify). A
comparison between three homologous intron-endcoded proteins from different species of
Naegleria (identity between 85-96%;T.M.E., unpublished results) strongly support the His-Cys
Box as a highly conserved region. Thus, we propose that the nuclear homing endonucleases are
all members of the same family, identified by the His-Cys Box.

<>

<1>Johansen, S., Vogt, V.M.
<2>An intron in the nuclear ribosomal DNA of Didymium iridis codes for a group I ribozyme and a novel ribozyme that cooperate in self-splicing.
<3>Cell
<4>76
<5>725-734
<6>1994
<7>We have discovered a unique group I intron-like insertion (DiSSU) in the nuclear small subunit
ribosomal RNA gene of the myxomycete Didymium iridis.  By sequence, DiSSU consists of a group
I ribozyme at the 5' end, an open reading frame (ORF) in the middle, and novel element at the
3' end.  Intron RNA self-splices in vitro to yield ten major processed RNAs, including a
full-length circle.  The group I ribozyme can efficiently cleave at an internal processing
site, which separates the group I ribozyme from the ORF.  Surprisingly, deletions that remove
the entire group I ribozyme do not impair cleavage at the 3' splice site, implying that the
3' element itself is a catalytic RNA.  Deletions that remove portions of the 3' element
prevent utilization of the 5' splice site, suggesting that this element cooperates with the
upstream group I ribozyme in splicing.  DiSSU appears to be the first example for the
cooperative interaction of distinct ribozymes in RNA splicing.

<>

<1>Johnson, D.L., Reid, T.M., Lee, M.-S., King, C.M., Romano, L.J.
<2>Preparation and characterization of a viral DNA molecule containing a site-specific 2-aminofluorene adduct:  A new probe for mutagenesis by carcinogens.
<3>Biochemistry
<4>25
<5>449-456
<6>1986
<7>The synthetic oligonucleotide heptamer 5'-ATCCGTC-3' was reacted in vitro with
N-acetoxy-N-(trifluoroacetyl)-2-aminofluorene and the resulting product
isolated by reverse-phase high-performance liquid chromatography (HPLC).  This
purified oligonucleotide, which was shown by chemical and enzymatic analysis to
be a heptamer containing a single N-(deoxyguanin-8-yl)-2-aminofluorene adduct,
was then used to situate the putatively mutagenic aminofluorene lesion within
the genome of M13 mp9 by ligating it into a complementary single-stranded
region located at a specific site in the negative strand of the duplex M13 mp9
DNA molecule.  The presence of the adduct at the anticipated location was
confirmed by taking advantage of the facts that AF adducts inhibit many
restriction enzymes when located in or near their restriction sites and that
the AF moiety should be contained within the HincII recognition sequence on M13
mp9 DNA.  Upon attempted cleavage of the M13 DNA containing the site-specific
AF adduct with HincII, we find that the large majority of the DNA remained
circular, demonstrating the incorporation of the AF adduct in high yield into
the DNA molecule at this location.  This system should prove useful in vivo for
the study of mutagenesis by chemical carcinogens and in vitro to study the
interaction of purified DNA metabolizing proteins with a template containing a
site-specific lesion.

<>

<1>Johnson, E.A., Marshall, K.M., Bradshaw, M.
<2>Conjugative plasmids from Clostridium botulinum and methods of use thereof.
<3>International Patent Office
<4>WO 2011047274 A
<5>
<6>2011
<7>The present invention provides novel C. botulinum conjugatively transmissible plasmids and
methods of use thereof.  Specifically, described herein are novel, conjugatively transmissible
clostridial plasmids which are capable of being transferred among and between clostridial
species.  The novel plasmids of the present invention therefore permits the delivery of
heterologous clostridial genes into a clostridial host, such as C. botulinum, and the
expression of genes of interest in that host, including clostridial toxins and the
nontoxigenic components of the toxin complex, toxin fragments, or antigenic portions thereof,
in a way both that ensures abundant expression and facilitates purification.  Furthermore,
toxins with altered structures, chimeric, hybrid toxins, and other toxin derivatives valuable
in medicine could be synthesized in this system.

<>

<1>Johnson, J.G., Carpentier, S., Spurbeck, R.R., Sandhu, S.K., Dirita, V.J.
<2>Genome Sequences of Campylobacter jejuni 81-176 Variants with Enhanced Fitness Relative to the Parental Strain in the Chicken Gastrointestinal Tract.
<3>Genome Announcements
<4>2
<5>e00006-14
<6>2014
<7>Campylobacter jejuni is a major cause of food-borne infections in the United States due to its
ability to asymptomatically colonize the gastrointestinal
tracts of chickens. Using competition assays with parental C. jejuni 81-176,
variants with consistently improved fitness in chicken ceca relative to the
parental strain were identified and sequenced.

<>

<1>Johnson, J.G., Spurbeck, R.R., Sandhu, S.K., Matson, J.S.
<2>Genome Sequence of Klebsiella pneumoniae Urinary Tract Isolate Top52.
<3>Genome Announcements
<4>2
<5>e00668-14
<6>2014
<7>Klebsiella pneumoniae is a significant cause of nosocomial infections, including
ventilator-associated pneumonias and catheter-associated urinary tract
infections. K. pneumoniae strain TOP52 #1721 (Top52) was isolated from a woman
presenting with acute cystitis and subsequently characterized using various
murine models of infection. Here we present the genome sequence of K. pneumoniae
Top52.

<>

<1>Johnson, J.G., Spurbeck, R.R., Sandhu, S.K., Matson, J.S.
<2>Genome Sequence of Klebsiella pneumoniae Respiratory Isolate IA565.
<3>Genome Announcements
<4>2
<5>e00896-14
<6>2014
<7>Klebsiella pneumoniae is a clinically significant opportunistic bacterial pathogen as well as
a normal member of the human microbiota. K. pneumoniae strain
IA565 was isolated from a tracheal aspirate at the University of Iowa Hospitals
and Clinics. Here, we present the genome sequence of K. pneumoniae IA565.

<>

<1>Johnson, M., Scheinbart, L., Pike, M., Richardson, B.
<2>Procainamide inhibits DNA methyltransferase.
<3>Arthritis Rheum.
<4>32
<5>S143
<6>1989
<7>We have reported that T cells treated with procainamide (Pca) have
hypomethylated DNA.  Pca can bind DNA, so we tested whether Pca inhibits T cell
DNA methyltransferase.  DNA methyltransferase was partially purified from
nuclei isolated from the human T cell leukemia line Jurkat.  Methyltransferase
activity was measured by incubating the enzyme with Micrococcus luteus DNA and
3H-S-adenosylmethionine (3H-SAM), then precipitating DNA onto fiberglass
filters with ethanol and determining precipitated 3H.  A dose dependent
inhibition of DNA methylation was observed, with 100 micromolar Pca inhibiting
3H incorporation into DNA by 51+/-5% (mean +/1 SEM of three experiments, each
performed in triplicate) (p<0.05).  Lineweaver-Burk analysis demonstrated
competitive inhibition with DNA.  To test whether Pca is a nonspecific
inhibitor of all transmethylation reactions, Jurkat cells were treated with Pca
for four days, then incubated with 3H-SAM and 3H incorporation into membrane
phospholipids measured.  No inhibition was observed.  These results demonstrate
that Pca is a DNA methyltransferase inhibitor, and suggest that Pca inhibits
Jurkat DNA methylation by selectively inhibiting this enzyme.

<>

<1>Johnson, P.H., Lee, A.S., Sinsheimer, R.L.
<2>Production of specific fragments of PhiX174 replicative form DNA by a restriction enzyme from Haemophilus parainfluenzae, endonuclease HP.
<3>J. Virol.
<4>11
<5>596-599
<6>1973
<7>A restriction endonuclease from Haemophilus parainfluenzae degrades Phi174
replicative form DNA into eight specific fragments, ranging from 1,700 to 150
base pairs and terminated specifically by deoxycytidylic acid.

<>

<1>Johnson, T.J., Fernandez-Alarcon, C., Bojesen, A.M., Nolan, L.K., Trampel, D.W., Seemann, T.
<2>Complete Genome Sequence of Gallibacterium anatis strain UMN179, isolated from a laying hen with peritonitis.
<3>J. Bacteriol.
<4>193
<5>3676-3677
<6>2011
<7>Gallibacterium anatis is a member of the normal flora of avian hosts and an important
causative agent of peritonitis and salpingitis in laying hens. Here we report the availability
of the first completed G. anatis genome sequence of strain UMN179, isolated from an Iowa
laying hen with peritonitis.

<>

<1>Johnson, T.J., Johnson, S.J., Nolan, L.K.
<2>Complete DNA sequence of a ColBM plasmid from avian pathogenic Escherichia coli suggests that it evolved from closely related ColV virulence plasmids.
<3>J. Bacteriol.
<4>188
<5>5975-5983
<6>2006
<7>Avian pathogenic Escherichia coli (APEC), an extraintestinal pathogenic E.
coli causing colibacillosis in birds, is responsible for significant
economic losses for the poultry industry. Recently, we reported that the
APEC pathotype was characterized by possession of a set of genes contained
within a 94-kb cluster linked to a ColV plasmid, pAPEC-O2-ColV. These
included sitABCD, genes of the aerobactin operon, hlyF, iss, genes of the
salmochelin operon, and the 5' end of cvaB of the ColV operon. However,
the results of gene prevalence studies performed among APEC isolates
revealed that these traits were not always linked to ColV plasmids. Here,
we present the complete sequence of a 174-kb plasmid, pAPEC-O1-ColBM,
which contains a putative virulence cluster similar to that of
pAPEC-O2-ColV. These two F-type plasmids share remarkable similarity,
except that they encode the production of different colicins;
pAPEC-O2-ColV contains an intact ColV operon, and pAPEC-O1-ColBM encodes
the colicins B and M. Interestingly, remnants of the ColV operon exist in
pAPEC-O1-ColBM, hinting that ColBM-type plasmids may have evolved from
ColV plasmids. Among APEC isolates, the prevalence of ColBM sequences
helps account for the previously observed differences in prevalence
between genes of the "conserved" portion of the putative virulence cluster
of pAPEC-O2-ColV and those genes within its "variable" portion. These
results, in conjunction with Southern blotting and probing of
representative ColBM-positive strains, indicate that this "conserved"
cluster of putative virulence genes is primarily linked to F-type
virulence plasmids among the APEC isolates studied.

<>

<1>Johnson, T.J., Kariyawasam, S., Wannemuehler, Y., Mangiamele, P., Johnson, S.J., Doetkott, C., Skyberg, J.A., Lynne, A.M., Johnson, J.R., Nolan, L.K.
<2>The genome sequence of avian pathogenic Escherichia coli strain O1:K1:H7 shares strong similarities with human extraintestinal pathogenic E. coli  genomes.
<3>J. Bacteriol.
<4>189
<5>3228-3236
<6>2007
<7>Escherichia coli strains that cause disease outside the intestine are known as extraintestinal
pathogenic E. coli (ExPEC) and include human
uropathogenic E. coli (UPEC) and avian pathogenic E. coli (APEC).
Regardless of host of origin, ExPEC strains share many traits. It has been
suggested that these commonalities may enable APEC to cause disease in
humans. Here, we begin to test the hypothesis that certain APEC strains
possess potential to cause human urinary tract infection through virulence
genotyping of 1,000 APEC and UPEC strains, generation of the first
complete genomic sequence of an APEC (APEC O1:K1:H7) strain, and
comparison of this genome to all available human ExPEC genomic sequences.
The genomes of APEC O1 and three human UPEC strains were found to be
remarkably similar, with only 4.5% of APEC O1's genome not found in other
sequenced ExPEC genomes. Also, use of multilocus sequence typing showed
that some of the sequenced human ExPEC strains were more like APEC O1 than
other human ExPEC strains. This work provides evidence that at least some
human and avian ExPEC strains are highly similar to one another, and it
supports the possibility that a food-borne link between some APEC and UPEC
strains exists. Future studies are necessary to assess the ability of APEC
to overcome the hurdles necessary for such a food-borne transmission, and
epidemiological studies are required to confirm that such a phenomenon
actually occurs.

<>

<1>Johnston, C., Martin, B., Granadel, C., Polard, P., Claverys, J.P.
<2>Programmed Protection of Foreign DNA from Restriction Allows Pathogenicity Island Exchange during Pneumococcal Transformation.
<3>PLoS Pathog.
<4>9
<5>e1003178
<6>2013
<7>In bacteria, transformation and restriction-modification (R-M) systems play potentially
antagonistic roles. While the former, proposed as a
form of sexuality, relies on internalized foreign DNA to create genetic
diversity, the latter degrade foreign DNA to protect from bacteriophage
attack. The human pathogen Streptococcus pneumoniae is transformable
and possesses either of two R-M systems, DpnI and DpnII, which
respectively restrict methylated or unmethylated double-stranded (ds)
DNA. S. pneumoniae DpnII strains possess DpnM, which methylates dsDNA
to protect it from DpnII restriction, and a second methylase, DpnA,
which is induced during competence for genetic transformation and is
unusual in that it methylates single-stranded (ss) DNA. DpnA was
tentatively ascribed the role of protecting internalized plasmids from
DpnII restriction, but this seems unlikely in light of recent results
establishing that pneumococcal transformation was not evolved to favor
plasmid exchange. Here we validate an alternative hypothesis, showing
that DpnA plays a crucial role in the protection of internalized
foreign DNA, enabling exchange of pathogenicity islands and more
generally of variable regions between pneumococcal isolates. We show
that transformation of a 21.7 kb heterologous region is reduced by more
than 4 logs in dpnA mutant cells and provide evidence that the specific
induction of dpnA during competence is critical for full protection. We
suggest that the integration of a restrictase/ssDNA-methylase couplet
into the competence regulon maintains protection from bacteriophage
attack whilst simultaneously enabling exchange of pathogenicicy
islands. This protective role of DpnA is likely to be of particular
importance for pneumococcal virulence by allowing free variation of
capsule serotype in DpnII strains via integration of DpnI capsule loci,
contributing to the documented escape of pneumococci from capsule-based
vaccines. Generally, this finding is the first evidence for a mechanism
that actively promotes genetic diversity of S. pneumoniae through
programmed protection and incorporation of foreign DNA.

<>

<1>Johnston, C., Martin, B., Polard, P., Claverys, J.-P.
<2>Postreplication targeting of transformants by bacterial immune systems?
<3>Trends Microbiol.
<4>21
<5>516-521
<6>2013
<7>Bacteria are constantly challenged by foreign genetic elements such as bacteriophages and
plasmids. Several defense systems provide immunity against such attackers, including
restriction modification (R M) systems and clustered, regularly interspaced short palindromic
repeats (CRISPRs). These systems target attacking DNA and thus antagonize natural
transformation, which relies on uptake of exogenous DNA to promote acquisition of new genetic
traits. It is unclear how this antagonization occurs, becau e transforming DNA is single
stranded, and thus resistant to these immune systems. Here, we propose a simple model whereby
these systems limit transformation by attack of transformed chromosomes once double
strandedness is restored by chromosomal replication.

<>

<1>Jois, P.S., Madhu, N., Rao, D.N.
<2>Role of histidine residues in EcoP15I DNA methyltransferase activity as probed by chemical modification and site-directed mutagenesis.
<3>Biochem. J.
<4>410
<5>543-553
<6>2008
<7>Towards understanding the catalytic mechanism of M.EcoP15I [EcoP15I MTase (DNA
methyltransferase); an adenine methyltransferase], we
investigated the role of histidine residues in catalysis. M.EcoP15I,
when incubated with DEPC (diethyl pyrocarbonate), a histidine-specific
reagent, shows a time- and concentration-dependent inactivation of
methylation of DNA containing its recognition sequence of 5'-CAGCAG-3'.
The loss of enzyme activity was accompanied by an increase in
absorbance at 240 nm. A difference spectrum of modified versus native
enzyme shows the formation of N-carbethoxyhistidine that is diminished
by hydroxylamine. This, along with other experiments, strongly suggests
that the inactivation of the enzyme by DEPC was specific for histidine
residues. Substrate protection experiments show that pre-incubating the
methylase with DNA was able to protect the enzyme from DEPC
inactivation. Site-directed mutagenesis experiments in which the 15
histidine residues in the enzyme were replaced individually with
alanine corroborated the chemical modification studies and established
the importance of His-335 in the methylase activity. No gross
structural differences were detected between the native and H335A
mutant MTases, as evident from CD spectra, native PAGE pattern or on
gel filtration chromatography. Replacement of histidine with alanine
residue at position 335 results in a mutant enzyme that is
catalytically inactive and binds to DNA more tightly than the wild-type
enzyme. Thus we have shown in the present study, through a combination
of chemical modification and site-directed mutagenesis experiments,
that His-335 plays an essential role in DNA methylation catalysed by
M.EcoP15I.

<>

<1>Jolly, J.F.
<2>Isothermal nucleic acid amplification methods using base analogs to limit restriction endonuclease cleavage to generation of single-stranded nicks.
<3>International Patent Office
<4>WO 200028084 A
<5>
<6>2000
<7>Methods are provided for the rapid, substantially isothermal, amplification of the sequence
information of a target nucleic acid positioned within a single- or double-stranded
polynucleotide fragment.  The methods are based on the serial generation of double-stranded
DNA engineered to contain terminal nicking of at least one of those sites, and extensions from
the nick(s), thereby displacing any existing polynucleotides.  A kit combining the components
commonly used in practicing methods of the invention is also provided.

<>

<1>Jomova, K., Hegedusova, A., Vollmannova, A.
<2>Restriction endonucleases - a tool for DNA cleavage.
<3>Chem. Rozhlady
<4>4
<5>239-245
<6>2004
<7>
<>

<1>Jonczyk, P., Hines, R., Smith, D.W.
<2>The Escherichia coli dam gene is expressed as a distal gene of a new operon.
<3>Mol. Gen. Genet.
<4>217
<5>85-96
<6>1989
<7>DNA containing the Escherichia coli dam gene and sequences upstream from this gene were cloned
from the Clarke-Carbon plasmids pLC29-47 and pLC13-42. Promoter activity was localized using
pKO expression vectors and galactokinase assays to two regions, one 1650-2100 bp and the other
beyond 2400 bp upstream of the dam gene. No promoter activity was detected immediately in
front of this gene; plasmid pDam118, from which the nucleotide sequence of the dam gene was
determined, is shown to contain the pBR322 promoter for the primer RNA from the pBR322 rep
region present on a 76 bp Sau3A fragment inserted upstream of the dam gene in the correct
orientation for dam expression. The nucleotide sequence upstream of dam has been determined.
An open reading frame (ORF) is present between the nearest promoter region and the dam gene.
Codon usage and base frequency analysis indicate that this is expressed as a protein of
predicted size 46 kDa. A protein of size close to 46 kDa is expressed from this region,
detected using minicell analysis. No function has been determined for this protein, and no
significant homology exist between it and sequences in the PIR protein or GenBank DNA
databases. This unidentified reading frame (URF) is termed urf-74.3, since it is an URF
located at 74.3 min on the E. coli chromosome. Sequence comparisons between the regions
upstream of urf-74.3 and the aroB gene show that the aroB gene is located immediately upstream
of urf-74.3, and that the promoter activity nearest to dam is found within the aroB structural
gene. This activity is relatively weak (about 15% of that of the E. coli gal operon promoter).
The promoter activity detected beyond 2400 bp upstream of dam is likely to be that of the aroB
gene, and is 3 to 4 times stronger than that found within the aroB gene. Three potential DnaA
binding sites, each with homology of 8 of 9 bp, are present, two in the aroB promoter region
and one just upstream of the dam gene. Expression through the site adjacent to the dam gene is
enhanced 2- to 4-fold in dnaA mutants at 38C. Restriction site comparisons map these regions
precisely on the Clarke-Carbon plasmids pLC13-42 and pLC29-47, and show that the E. coli ponA
(mrcA) gene resides about 6 kb upstream of aroB.

<>

<1>Jones, A.C. et al.
<2>Genomic insights into the physiology and ecology of the marine filamentous cyanobacterium Lyngbya majuscula.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>8815-8820
<6>2011
<7>Filamentous cyanobacteria of the genus Lyngbya are important contributors
to coral reef ecosystems, occasionally forming dominant cover and
impacting the health of many other co-occurring organisms. Moreover, they
are extraordinarily rich sources of bioactive secondary metabolites, with
35% of all reported cyanobacterial natural products deriving from this
single pantropical genus. However, the true natural product potential and
life strategies of Lyngbya strains are poorly understood because of
phylogenetic ambiguity, lack of genomic information, and their close
associations with heterotrophic bacteria and other cyanobacteria. To gauge
the natural product potential of Lyngbya and gain insights into potential
microbial interactions, we sequenced the genome of Lyngbya majuscula 3L, a
Caribbean strain that produces the tubulin polymerization inhibitor
curacin A and the molluscicide barbamide, using a combination of Sanger
and 454 sequencing approaches. Whereas  approximately  293,000 nucleotides
of the draft genome are putatively dedicated to secondary metabolism, this
is far too few to encode a large suite of Lyngbya metabolites, suggesting
Lyngbya metabolites are strain specific and may be useful in species
delineation. Our analysis revealed a complex gene regulatory network,
including a large number of sigma factors and other regulatory proteins,
indicating an enhanced ability for environmental adaptation or microbial
associations. Although Lyngbya species are reported to fix nitrogen,
nitrogenase genes were not found in the genome or by PCR of genomic DNA.
Subsequent growth experiments confirmed that L. majuscula 3L is unable to
fix atmospheric nitrogen. These unanticipated life history characteristics
challenge current views of the genus Lyngbya.

<>

<1>Jones, M., Wagner, R., Radman, M.
<2>Mismatch repair of deaminated 5-methyl-cytosine.
<3>J. Mol. Biol.
<4>194
<5>155-159
<6>1987
<7>Deamination of 5-methyl-cytosine in double-stranded DNA produces a G.T mismatch.
Heteroduplexes of bacteriophage lambda DNA containing a G.T mismatch at the site of G.5-meC
base-pair in one of the parental phages were constructed and used to transfect Escherichia
coli cells. Genetic analysis of the progeny phages derived from such heteroduplexes suggests
that, in E. coli, mismatches resulting from the deamination of 5-methyl-cytosine are repaired
by a system requiring the E. coli dcm methylase and some, but not all, of the functions of the
E. coli methyl-directed mismatch repair system. The repair appears to act only on the G.T
mismatch and acts specifically to restore the cytosine methylation sequence.

<>

<1>Jones, P.A.
<2>DNA methylation errors and cancer.
<3>Cancer Res.
<4>56
<5>2463-2467
<6>1996
<7>It has become clear over the last few years that DNA methylation is essential for normal
embryonic development and that alterations in DNA methylation are very common in cancer cells
and are capable of directly modifying carcinogenesis.  Cytosine methylation is responsible for
the induction of a surprisingly high percentage of disease-causing point mutations in tumor
suppressor genes in somatic and germline cells.  This may either be due to the spontaneous
deamination of 5-methylcytosine or to a more active process involving side reactions during
the enzymatic modification of cytosine in DNA.  Current interest in the role of methylation
has focused on the potential for abnormal methylation events to silence tumor suppressor
genes, thus giving rise to a novel pathway to cause their progressive epigenetic inactivation.
Random methylation of CpG islands not methylated in normal cells may contribute to the
progressive inactivation of growth-inhibitory genes resulting in the clonal selection of cells
with increasingly abnormal methylation patterns.  This model for gene inactivation during
cancer development has important clinical implications, since it is possible to reactivate
these dormant genes using inhibitors of DNA methylation and potentially restore growth control
to cells.

<>

<1>Jones, P.A., Gonzalgo, M.L.
<2>Altered DNA methylation and genome instability: A new pathway to cancer?
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>2103-2105
<6>1997
<7>DNA methylation is a mechanism for changing the base sequence of DNA without altering its
coding function.  As a heritable, yet reversible, epigenetic change, it has the potential of
altering gene expression and has profound developmental and genetic consequences.  The
methylation reaction itself is mechanistically complex and involves the flipping of the target
cytosine out of the intact double helix, so that the transfer of the methyl group from
S-adenosylmethionine can occur in a cleft in the enzyme.  Cytosine methylation is inherently
mutagenic, which presumably has led to the 80% suppression of the CpG methyl acceptor site in
eukarytoic organisms, which methylate their genomes.  It contributes strongly to the
generation of polymorphisms and germ-line mutations, and to transition mutations that
inactivate tumor-suppressor genes.  Despite a 10- to 40-fold increase in the rate of
transitions at methylated versus unmethylated cytosines, methylation is not only tolerated in
several eukaryotes, but is actually required for the embryonic development of mammals.  The
reasons 5-methylcytosine is essential for development remain obscure, but most probably relate
to the well-documented ability of methylation, particularly the methylation of CpG-rich
promoters, to block transcriptional activation.  Indeed, there is growing evidence that
methylation plays a pivotal role in key developmental processes such as genomic imprinting and
stabilization of X-chromosome inactivation.  It therefore is not surprising that alterations
in this essential epigenetic system might play a role in carcinogenesis.

<>

<1>Jones, P.A., Takai, D.
<2>The role of DNA methylation in mammalian epigenetics.
<3>Science
<4>293
<5>1068-1070
<6>2001
<7>Genes constitute only a small proportion of the total mammalian genome, and the precise
control of their expression in the presence of an overwhelming background of noncoding DNA
presents a substantial problem for their regulation. Noncoding DNA, containing introns,
repetitive elements, and potentially active transposable elements, requires effective
mechanisms for its long-term silencing.  Mammals appear to have taken advantage of the
possibilities afforded by cytosine methylation to provide a heritable mechanism for altering
DNA-protein interactions to assist in such silencing.  Genes can be transcribed from
methylation-free promoters even though adjacent transcribed and nontranscribed regions are
extensively methylated. Gene promoters can be used and regulated while keeping noncoding DNA,
including transposable elements, suppressed. Methylation is also used for long-term epigenetic
silencing of X-linked and imprinted genes and can either increase or decrease the level of
transcription, depending on whether the methylation inactivates a positive or negative
regulatory element.

<>

<1>Jones, P.L., Veenstra, G.J.C., Wade, P.A., Vermaak, D., Kass, S.U., Landsberger, N., Strouboulis, J., Wolffe, A.P.
<2>Methylated DNA and MeCP2 recruit histone deacetylase to repress transcription.
<3>Nat. Genet.
<4>19
<5>187-191
<6>1998
<7>CpG methylation in vertebrates correlates with alterations in chromatin structure and gene
silencing.  Differences in DNA-methylation status are associated with imprinting phenomena and
carcinogenesis.  In Xenopus laevis oocytes, DNA methylation dominantly silences transcription
through the assembly of a repressive nucleosomal array.  Methylated DNA assembled into
chromatin binds the transcriptional repressor MeCP2 which cofractionates with Sin3 and histone
deacetylase.  Silencing conferred by MeCP2 and methylated DNA can be relieved by inhibition of
histone deacetylase, facilitating the remodelling of chromatin and transcriptional activation.
These results establish a direct casual relationship between DNA methylation-dependent
transcription silencing and the modification of chromatin.

<>

<1>Jones, W.D. Jr., Greenberg, J.
<2>Host modification and restriction with a mycobacteriophage isolated from a pseudolysogenic Mycobacterium chelonei.
<3>J. Gen. Microbiol.
<4>99
<5>389-395
<6>1977
<7>A pseudolysogenic Mycobacterium chelonei and its phage Phi630 are described.  Phage Phi630 is
the first mycobacteriophage reported to be resistant to the nonpolar solvents chloroform,
dioxan and diethyl ether.  The phage had a latent period of 75 min, a rise period of 90 min
and a burst size of 51.  Evidence is presented for host modification and restriction.  Phage
Phi630A, grown on host strain M. chelonei F-630 Rg, plated on the alternative host M.
smegmatis ATCC607 with an efficiency of plating of 10^-5 on the alternative host F-630 Rg.
Phages Phi630A and Phi630B adsorbed equally well on their alternative hosts and on their
indicator host strains.  The progeny of plaques from initial platings on the alternative host,
when grown in the alternative host, exhibited a marked reduction in e.o.p. on their original
host.

<>

<1>Joo, E.J., Park, W.S., Kim, S.K., Bae, Y.S., Moon, B.J.
<2>Restriction endonuclease EcoRV mutants that switch metal ion requirement.
<3>Bull. Korean Chem. Soc.
<4>20
<5>109-111
<6>1999
<7>Divalent metal ions, normally Mg2+, are essential for both DNA cleavage by the EcoRV
restriction endonuclease at its recognition sequence, GATATC, and also for the enzyme's
discrimination between this particular sequence and all other sequences.  In the absence of
divalent metal ions, EcoRV demonstrates no catalytic activity, though it can still bind to DNA
in a nonspecific manner with no preference in its recognition sequence.  The complex of EcoRV
and its cognate DNA, however, has a high affinity for Mg2+ due to the distortion of the bound
DNA, creating a metal-binding site between the protein and the DNA.

<>

<1>Jordan, I.K. et al.
<2>Genome Sequences for Five Strains of the Emerging Pathogen Haemophilus haemolyticus.
<3>J. Bacteriol.
<4>193
<5>5879-5880
<6>2011
<7>We report the first whole-genome sequences for five strains, two carried and three pathogenic,
of the emerging pathogen Haemophilus haemolyticus.
Preliminary analyses indicate that these genome sequences encode markers
that distinguish H. haemolyticus from its closest Haemophilus relatives
and provide clues to the identity of its virulence factors.

<>

<1>Jore, J.P.M., van Luijk, N., Luiten, R.G.M., van der Werf, M.J., Pouwels, P.H.
<2>Efficient transformation system for Propionibacterium freudenreichii based on a novel vector.
<3>Appl. Environ. Microbiol.
<4>67
<5>499-503
<6>2001
<7>A 3.6-kb endogenous plasmid was isolated from a Propionibacterium freudenreichii strain and
sequenced completely. Based on homologies with plasmids from other bacteria, notably a plasmid
from Mycobacterium, a region harboring putative replicative functions was defined. Outside
this region two restriction enzyme recognition sites were used for insertion of an Escherichia
coli-specific replicon and an erythromycin resistance gene for selection in Propionibacterium.
Hybrid vectors obtained in this way replicated in both E. coli and P. freudenreichii. Whereas
electroporation of P. freudenreichii with vector DNA isolated from an E. coli transformant
yielded 10 to 30 colonies per microgram of DNA, use of vector DNA reisolated from a
Propionibacterium transformant dramatically increased the efficiency of transformation (> or
=10(8) colonies per microgram of DNA). It could be shown that restriction-modification was
responsible for this effect. The high efficiency of the system described here permitted
successful transformation of Propionibacterium with DNA ligation mixtures.

<>

<1>Jorgensen, T.S., Xu, Z., Hansen, M.A., Sorensen, S.J., Hansen, L.H.
<2>Hundreds of circular novel plasmids and DNA elements identified in a rat cecum metamobilome.
<3>PLoS ONE
<4>9
<5>E87924
<6>2014
<7>Metagenomic approaches are widespread in microbiological research, but so far,
the knowledge on extrachromosomal DNA diversity and composition has largely
remained dependant on cultivating host organisms. Even with the emergence of
metagenomics, complete circular sequences are rarely identified, and have
required manual curation. We propose a robust in silico procedure for identifying
complete small plasmids in metagenomic datasets from whole genome shotgun
sequencing. From one very pure and exhaustively sequenced metamobilome from rat
cecum, we identified a total of 616 circular sequences, 160 of which were
carrying a gene with plasmid replication domain. Further homology analyses
indicated that the majority of these plasmid sequences are novel. We confirmed
the circularity of the complete plasmid candidates using an inverse-type PCR
approach on a subset of sequences with 95% success, confirming the existence and
length of discrete sequences. The implication of these findings is a broadened
understanding of the traits of circular elements in nature and the possibility of
massive data mining in existing metagenomic datasets to discover novel pools of
complete plasmids thus vastly expanding the current plasmid database.

<>

<1>Jose, T.J., Conlan, L.H., Dupureur, C.M.
<2>Quantitative evaluation of metal ion binding to PvuII restriction endonuclease.
<3>J. Biol. Inorg. Chem.
<4>4
<5>814-823
<6>1999
<7>Restriction enzymes are important examples of phosphodiester hydrolysis activity and as such
have been of increasing interest to structural biologists. Much of the architecture of
endonuclease active sites has been derived from X-ray crystallographic studies. These
structures implicate conserved active site acidic residues and the scissile bond of the
substrate as coordination ligands of required metal ions. Central to the development of
restriction enzyme mechanism is our understanding of the role of metal ion binding in the
reaction, an important feature of which is identifying the energetic contributions of the
enzyme and the substrate to metal ion affinity. To begin to address this issue, isothermal
titration calorimetry (ITC) and 19F NMR spectroscopy have been applied to evaluate metal ion
binding by the representative PvuII endonuclease in the absence of substrate. In separate
experiments, ITC data demonstrate that PvuII endonuclease binds 2.16 Mn(II) ions and 2.05
Ca(II) metal ions in each monomer active site with Kd values of approximately 1 mM. While
neither calorimetry nor protein NMR spectroscopy is directly sensitive to Mg(II) binding to
the enzyme, Mn(II) competes with Mg(II) for common sites(s) on PvuII endonuclease.
Substitution of the conserved active site carboxylate Glu68 with Ala resulted in a loss of
affinity for both equivalents of both Ca(II) and Mn(II). Interestingly, the active site mutant
D58A retained an affinity for Mn(II) with Kd approximately 2 mM. Mn(II) paramagnetic
broadening in 19F spectra of wild-type and mutant 3-fluorotyrosine PvuII endonucleases are
consistent with ITC results. Chemical shift analysis of 3-fluorotyrosine mutant enzymes is
consistent with a perturbed conformation for D58A. Therefore, free PvuII endonuclease binds
metal ions, and metal ion binding can precede DNA binding. Further, while Glu68 is critical to
metal ion binding, Asp58 does not appear to be critical to the binding of at least one metal
ion and appears to also have a role in structure. These findings provide impetus for exploring
the roles of multiple metal ions in the structure and function of this representative
endonuclease.

<>

<1>Joseph, B., Schneiker-Bekel, S., Schramm-Gluck, A., Blom, J., Claus, H., Linke, B., Schwarz, R.F., Becker, A., Goesmann, A., Frosch, M., Schoen, C.
<2>Comparative genome biology of a serogroup B carriage and disease strain supports a polygenic nature of meningococcal virulence.
<3>J. Bacteriol.
<4>192
<5>5363-5377
<6>2010
<7>Neisseria meningitidis serogroup B strains are responsible for most
meningococcal cases in the industrialized countries, and strains belonging
to the clonal complex ST-41/44 are among the most prevalent serogroup B
strains in carriage and disease. Here, we report the first genome and
transcriptome comparison of a serogroup B carriage strain from the clonal
complex ST-41/44 to the serogroup B disease strain MC58 from the clonal
complex ST-32. Both genomes are highly colinear, with only three major
genome rearrangements that are associated with the integration of mobile
genetic elements. They further differ in about 10% of their gene content,
with the highest variability in gene presence as well as gene sequence
found for proteins involved in host cell interactions, including Opc,
NadA, TonB-dependent receptors, RTX toxin, and two-partner secretion
system proteins. Whereas housekeeping genes coding for metabolic functions
were highly conserved, there were considerable differences in their
expression pattern upon adhesion to human nasopharyngeal cells between
both strains, including differences in energy metabolism and stress
response. In line with these genomic and transcriptomic differences, both
strains also showed marked differences in their in vitro infectivity and
in serum resistance. Taken together, these data support the concept of a
polygenic nature of meningococcal virulence comprising differences in the
repertoire of adhesins as well as in the regulation of metabolic genes and
suggest a prominent role for immune selection and genetic drift in shaping
the meningococcal genome.

<>

<1>Joseph, T.C., Baby, A., Reghunathan, D., Varghese, A.M., Murugadas, V., Lalitha, K.V.
<2>Draft Genome Sequence of the Halophilic and Highly Halotolerant Gammaproteobacteria Strain MFB021.
<3>Genome Announcements
<4>2
<5>e01156-14
<6>2014
<7>We report the 4.25-Mbp first draft sequence of Gammaproteobacteria strain MFB021, a moderate
halophile isolated from petroleum-contaminated soil in Cochin, India.
The genome of the strain MFB021 was sequenced to understand the mechanism of
hydrocarbon degradation and the halophilicity of the bacterium.

<>

<1>Joseph, T.C., Varghese, A.M., Baby, A., Reghunathan, D., Murugadas, V., Lalitha, K.V.
<2>First Draft Genome Sequence of a Member of the Genus Mangrovibacter, Isolated from an Aquaculture Farm in India.
<3>Genome Announcements
<4>2
<5>e01209-14
<6>2014
<7>Mangrovibacter sp. MFB070, a Gram-negative, facultatively anaerobic, nitrogen-fixing
bacterium, was isolated from an aquaculture farm in Cochin,
India. Here, we report the first draft genome sequence of a member of the genus
Mangrovibacter, which may help us to elucidate the evolutionary status of this
genus. The draft genome sequence of the Mangrovibacter sp. consists of 5,361,682
bp, encoding 4,971 predicted coding sequences in 57 contigs.

<>

<1>Josephsen, J., Jorgen-Jensen, B., Nyengaard, N.R.
<2>Determination of the recognition sequence of the type II restriction endonuclease, LlaCI, from Lactococcus lactis W15.
<3>FEMS Microbiol. Lett.
<4>163
<5>25-29
<6>1998
<7>A new type II restriction endonuclease, called LlaCI, was partially purified from Lactococcus
lactis subsp. Cremoris W15.  The characterization of the LlaCI endonuclease showed it to be an
isoschizomer of HindIII, recognizing the sequence 5'-A/AGCTT-3'.  The cleavage site is
indicated by the arrow.

<>

<1>Josephsen, J., Klaenhammer, T.
<2>Stacking of three different restriction and modification systems in Lactococcus lactis by cotransformation.
<3>Plasmid
<4>23
<5>71-75
<6>1990
<7>Four plasmids encoding restriction and modification (R/M) systems are described
that are different in the specificity of their restrictive activity toward the
small isometric phage p2 and prolate phage c2.  The R/M plasmids were
cotransformed into Lactococcus lactis MG1363 with pVS2, encoding resistance to
chloramphenicol and erythromycin, to indicate successful transformation events.
Analysis of cotransformants showed that three different R/M plasmids could be
combined in L. lactis MG1363.  The efficiency at which phage plaqued on the
transformants decreased as the number of R/M plasmids increased.  Some plasmid
combinations were unstable suggesting replicon incompatibility.

<>

<1>Josephsen, J., Nyengaard, N.R., Vogensen, F.K., Madsen, A.
<2>Plasmid-derived type II restriction-modification systems from Lactococcus lactis.
<3>International Patent Office
<4>WO 9625503
<5>
<6>1996
<7>A number of type II restriction-modification (R-M) systems have been derived from plasmids
found in Lactococcus lactis strains from the Danish starter culture TK5.  The R-M systems
LlaAI, LlaBI and LlaDII are claimed with their nucleotide sequences containing open reading
frames (ORFs) coding for restriction endonucleases and corresponding methylases.  Also a DNA
cassette comprising one or more of the R-M systems and fragments thereof in combination with
DNA encoding other phage resistance mechanisms is claimed as are cloning and expression
vectors including DNA selected from the group consisting of the R-M systems, fragments thereof
and DNA cassette, cells transformed with the expression vectors, a method of conferring
increased virus resistance on a cell, and the individual restriction endonucleases and
methylases of the R-M systems.

<>

<1>Josephsen, J., Vogensen, F.K.
<2>Identification of three different plasmid-encoded restriction modification systems in Streptococcus-lactis subsp cremoris W56.
<3>FEMS Microbiol. Lett.
<4>59
<5>161-166
<6>1989
<7>Streptococcus lactis subsp. cremoris W56 (S. cremoris W56) is a strain partially resistant to
phage attack. Derivatives which had lost either plasmid pJW563 or pJW566 no longer expressed
the restriction and modification systems encoded by these plasmids. Genetic evidence for the
correlation between the plasmids and the R/M systems was obtained by transformation. In
addition, a third R/M system was discovered among the transformants and was shown to be
encoded by pJW565. Thus, genetic evidence for at least 3 distinct R/M systems encoded by
plasmids in S. cremoris W56 is presented. One of the R/M-systems showed stronger restriction
of the isometric phage p2 than of the prolate phage c2. The other two systems restricted both
classes of phages with equal efficiencies.

<>

<1>Josephsen, J., Vogensen, F.K., Madsen, A.
<2>Plasmid-derived LlaDII restriction-modification system from Lactococcus lactis.
<3>US Patent Office
<4>US 6300109
<5>
<6>2001
<7>A number of type II restriction-modification (R-M) systems have been derived from plasmids
found in Lactococcus lactis strains from the
Danish starter culture TK5. The R-M systems LlaAI, LlaBI and LlaDII are
claimed with their nucleotide sequences containing open reading frames
(ORFs) coding for restriction endonucleases and corresponding
methylases. Also a DNA cassette comprising one or more of the R-M
systems and fragments thereof in combination with DNA encoding other
phage resistance mechanisms is claimed as are cloning and expression
vectors including DNA selected from the group consisting of the R-M
systems, fragments thereof and DNA cassette, cells transformed with the
expression vectors, a method of conferring increased virus resistance
on a cell, and the individual restriction endonucleases and methylases
of the R-M systems.

<>

<1>Joshi, C.P., Chiang, V.L.
<2>Conserved sequence motifs in plant S-adenosyl-L-methionine-dependent methyltransferases.
<3>Plant Mol. Biol.
<4>37
<5>663-674
<6>1998
<7>Plant S-adenosyl-L-methionine-dependent methyltransferases are the key enzymes in
phenylpropanoid, flavonoid and many other metabolic pathways of biotechnological importance.
Here we compiled the amino acid sequences of 56 SAM-Mtases from different plants and performed
a computer analysis for the conserved sequence motifs that could possibly act as SAM-binding
domains.  To date, genes or cDNAs encoding at least ten distinct groups of SAM-Mtases that
utilize SAM and a variety of substrates have been reported from higher plants.  Three amino
acid sequence motifs are conserved in most of these SAM-Mtases.  In addition, many conserved
domains have been discovered in each group of O-methyltransferases that methylate specific
substrates and may act as sites for substrate specificity in each enzyme.  Finally, a
diagrammatic representation of the relationship between different OMTs is presented.  These
SAM-Mtase sequence signatures will be useful in the identification of SAM-Mtase motifs in the
hitherto unidentified proteins as well as for designing primers in the isolation of new
SAM-Mtases from plants.

<>

<1>Joshi, H.K., Etzkorn, C., Chatwell, L., Bitinaite, J., Horton, N.C.
<2>Alteration of sequence specificity of the type II restriction endonuclease HincII through an indirect readout mechanism.
<3>J. Biol. Chem.
<4>281
<5>23852-23869
<6>2006
<7>The functional and structural consequences of a mutation of the DNA intercalating residue of
HincII, Q138F, are presented. Modeling has
suggested that the DNA intercalation by Gln-138 results in DNA
distortions potentially used by HincII in indirect readout of its
cognate DNA, GTYRAC ( Y = C or T, R = A or G) ( Horton, N. C., Dorner,
L. F., and Perona, J. J. ( 2002) Nat. Struct. Biol. 9, 42 - 47).
Kinetic data presented here indicate that the mutation of glutamine 138
to phenylalanine ( Q138F) results in a change in sequence specificity
at the center two base pairs of the cognate recognition site. We show
that the preference of HincII for cutting, but not binding, the three
cognate sites differing in the center two base pairs has been altered
by the mutation Q138F. Five new crystal structures are presented
including Q138F HincII bound to GTTAAC and GTCGAC both with and without
Ca2+ as well as the structure of wild type HincII bound to GTTAAC. The
Q138F HincII/DNA structures show conformational changes in the protein,
bound DNA, and at the protein-DNA interface, consistent with the
formation of adaptive complexes. Analysis of these structures and the
effect of Ca2+ binding on the protein-DNA interface illuminates the
origin of the altered specificity by the mutation Q138F in the HincII
enzyme.

<>

<1>Joshi, M.N., Pandit, A.S., Sharma, A., Pandya, R.V., Desai, S.M., Saxena, A.K., Bagatharia, S.B.
<2>Draft Genome Sequence of Arthrobacter crystallopoietes Strain BAB-32, Revealing Genes for Bioremediation.
<3>Genome Announcements
<4>1
<5>e00452-13
<6>2013
<7>Arthrobacter crystallopoietes strain BAB-32, a Gram-positive obligate aerobic actinobacterium
having potential application in bioremediation and bioreduction
of a few metals, was isolated from rhizosphere soil of Gandhinagar, Gujarat,
India. The draft genome (4.3 Mb) of the strain revealed a few vital gene clusters
involved in the metabolism of aromatic compounds, zinc, and sulfur.

<>

<1>Joshi, M.N., Pandit, A.S., Sharma, A., Pandya, R.V., Saxena, A.K., Bagatharia, S.B.
<2>Draft Genome Sequence of the Halophilic Bacterium Halobacillus sp. Strain BAB-2008.
<3>Genome Announcements
<4>1
<5>e00222-12
<6>2013
<7>The sp. strain BAB-2008 is a moderately halophilic, rod-shaped, Gram-positive,
orange-pigmented, carotenoid-producing bacterium isolated from saline soil near
Zazam-Solar Park Road, Gujarat, India. Here we present the 3.7-Mb genome sequence
to provide insights into its functional genomics and potential applications for
carotenoid and enzyme production.

<>

<1>Joshi, M.N., Sharma, A., Pandit, A.S., Pandya, R.V., Saxena, A.K., Bagatharia, S.B.
<2>Draft Genome Sequence of Brevibacillus sp. Strain BAB-2500, a Strain That Might Play an Important Role in Agriculture.
<3>Genome Announcements
<4>1
<5>e00021-13
<6>2013
<7>A Gram-positive bacterium, sp. strain BAB-2500, was isolated as a lab contaminant in
Gandhinagar, Gujarat, India. The draft genome (5.3 Mb) of the strain possesses
genes for the reduction of arsenate and aluminum. These findings might provide
insights into the utilization of this strain for improving crop production.

<>

<1>Joshi, M.N., Sharma, A.C., Pandya, R.V., Patel, R.P., Saiyed, Z.M., Saxena, A.K., Bagatharia, S.B.
<2>Draft Genome Sequence of Pontibacter sp. nov. BAB1700, a Halotolerant, Industrially Important Bacterium.
<3>J. Bacteriol.
<4>194
<5>6329-6330
<6>2012
<7>Pontibacter sp. nov. BAB1700 is a halotolerant, Gram-negative, rod-shaped, pink-pigmented,
menaquinone-7-producing bacterium isolated from sediments of a
drilling well. The draft genome sequence of the strain, consisting of one
chromosome of 4.5 Mb, revealed vital gene clusters involved in vitamin
biosynthesis and resistance against various metals and antibiotics.

<>

<1>Joshi, R., Ho, K.K., Tenney, K., Chen, J.H., Golden, B.L., Gimble, F.S.
<2>Evolution of I-SceI Homing Endonucleases with Increased DNA Recognition Site Specificity.
<3>J. Mol. Biol.
<4>405
<5>185-200
<6>2011
<7>Elucidating how homing endonucleases undergo changes in recognition site specificity will
facilitate efforts to engineer proteins for gene therapy
applications. I-SceI is a monomeric homing endonuclease that recognizes
and cleaves within an 18-bp target. It tolerates limited degeneracy in its
target sequence, including substitution of a C:G(+4) base pair for the
wild-type A:T(+4) base pair. Libraries encoding randomized amino acids at
I-SceI residue positions that contact or are proximal to A:T(+4) were used
in conjunction with a bacterial one-hybrid system to select I-SceI
derivatives that bind to recognition sites containing either the A:T(+4)
or the C:G(+4) base pairs. As expected, isolates encoding wild-type
residues at the randomized positions were selected using either target
sequence. All I-SceI proteins isolated using the C:G(+4) recognition site
included small side-chain substitutions at G100 and either contained
(K86R/G100T, K86R/G100S and K86R/G100C) or lacked (G100A, G100T) a K86R
substitution. Interestingly, the binding affinities of the selected
variants for the wild-type A:T(+4) target are 4- to 11-fold lower than
that of wild-type I-SceI, whereas those for the C:G(+4) target are
similar. The increased specificity of the mutant proteins is also evident
in binding experiments in vivo. These differences in binding affinities
account for the observed  approximately 36-fold difference in target
preference between the K86R/G100T and wild-type proteins in DNA cleavage
assays. An X-ray crystal structure of the K86R/G100T mutant protein bound
to a DNA duplex containing the C:G(+4) substitution suggests how sequence
specificity of a homing enzyme can increase. This biochemical and
structural analysis defines one pathway by which site specificity is
augmented for a homing endonuclease.

<>

<1>Jost, B.H., Field, A.C., Trinh, H.T., Songer, J.G., Billington, S.J.
<2>Tylosin Resistance in Arcanobacterium pyogenes Is Encoded by an Erm X Determinant.
<3>Antimicrob. Agents Chemother.
<4>47
<5>3519-3524
<6>2003
<7>Arcanobacterium pyogenes, a commensal on the mucous membranes of many economically important
animal species, is also a pathogen, causing
abscesses of the skin, joints, and visceral organs as well as mastitis and
abortion. In food animals, A. pyogenes is exposed to antimicrobial agents
used for growth promotion, prophylaxis, and therapy, notably tylosin, a
macrolide antibiotic used extensively for the prevention of liver
abscessation in feedlot cattle in the United States. Of 48 A. pyogenes
isolates, 11 (22.9%) exhibited inducible or constitutive resistance to
tylosin (MIC of > or = 128 microg/ml). These isolates also exhibited
resistance to other macrolide and lincosamide antibiotics, suggesting a
macrolide-lincosamide resistance phenotype. Of the 11 resistant isolates,
genomic DNA from nine hybridized to an erm(X)-specific probe. Cloning and
nucleotide sequencing of the A. pyogenes erm(X) gene indicated that it was
>95% similar to erm(X) genes from Corynebacterium and Propionibacterium
spp. Eight of the erm(X)-containing A. pyogenes isolates exhibited
inducible tylosin resistance, which was consistent with the presence of a
putative leader peptide upstream of the erm(X) open reading frame. For at
least one A. pyogenes isolate, 98-4277-2, erm(X) was present on a plasmid,
pAP2, and was associated with the insertion sequence IS6100. pAP2 also
carried genes encoding the repressor-regulated tetracycline efflux system
determinant Tet 33. The repA gene from pAP2 was nonfunctional in
Escherichia coli and at least one A. pyogenes isolate, suggesting that
there may be host-encoded factors required for replication of this
plasmid.

<>

<1>Jost, J.-P.
<2>Nuclear extracts of chicken embryos promote an active demethylation of DNA by excision repair of 5-methyldeoxycytidine.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>4684-4688
<6>1993
<7>Here I show that nuclear extracts of chicken embryos can promote the active demethylation of
DNA.  The evidence shows that in hemimethylated DNA (i.e., methylated on one strand only)
demethylation of 5mCpG occurs throughout nucleotide excision repair.  The first step of
demethylation is the formation of specific nicks 5' from 5-methyldeoxycytidine.  Nicks are
also observed in vitro on symmetrically methylated CpGs (i.e., methylated on both strands) but
they result in breakage of the oligonucleotide with no repair.  No specific nicks are observed
on the nonmethylated CpG. Nicks are strictly 5mCpG specific and do not occur on 5mCpC, 5mCpT,
5mCpA, or 6mApT.  The effect of nonspecific nuclease(s) has been ruled out.  The nicking of
mCpG takes place in the presence of 20 mM EDTA irrespective of the nature of the sequence
surrounding the 5mCpG.  No methylcytosine glycosylase activity could be detected.  The repair
is aphidicolin and N-ethylmaleimide resistant, suggesting a repair action by DNA polymerase
beta.  In extracts of chicken embryos, the excision repair of mCpG is highest between the 6th
and the 12th day of development, whereas it is barely detectable in nuclear extracts from
different organs of adults.  The possible implications of 5mCpG endonuclease activity in
active demethylation of DNA during differentiation is discussed.

<>

<1>Jost, J.-P.
<2>Mechanism of DNA demethylation in vertebrates and its biological significance.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, Russo, V.E.A., Martienssen, R.A., Riggs, A.D., Cold Spring Harbor
<4>0
<5>109-125
<6>1996
<7>The aim of this chapter is to present some general features of site-specific and genome-wide
demethylation.  Demethylation of DNA is part of the process responsible for the formation of
specific methylation patterns.  The establishment of a CpG methylation pattern during
embryonic development involves at least three components: DNA methyltransferase, a
demethylation system (passive or active), and sequence-specific cis-trans controlling elements
located on or near the CpG methylation sites.  The cis-trans controlling elements either
enhance or inhibit the methylation of specific sequences.  Whether a given CpG site is
methylated or not depends on the mole ratio of activities of the different factors involved
and of their respective affinities for a specified CpG site on the DNA.  This concept of
titratable factors is also valid for the cis-acting control elements.  For example, the
introduction of DNA-binding sites for specific proteins into the cell could possibly titrate
out protein factors and engender hypermethylation of the DNA.

<>

<1>Jost, J.-P., Fremont, M., Siegmann, M., Hofsteenge, J.
<2>The RNA moiety of chick embryo 5-methylcytosine-DNA glycosylase targets DNA demethylation.
<3>Nucleic Acids Res.
<4>25
<5>4545-4550
<6>1997
<7>We have previously shown that DNA demethylation by chick embryo 5-methylcytosine (5-MeC)-DNA
glycosylase needs both protein and RNA.  RNA from enzyme purified by SDS-PAGE was isolated and
cloned.  The clones have an insert ranging from 240 to 670 bp and contained on average one CpG
per 14 bases.  All six clones tested had different sequences and did not have any sequence
homology with any other known RNA.  RNase-inactivated 5-MeC-DNA glycosylase regained enzyme
activity when incubated with recombinant RNA.  However, when recombinant RNA was incubated
with the DNA substrate alone there was no demethylation activity.  Short sequences
complementary to the labeled DNA substrate are present in the recombinant RNA.  Small
synthetic oligoribonucleotides (11 bases long) complementary to the region of methylated CpGs
of the hemimethylated double-stranded DNA substrate restore the activity of the
RNase-inactivated 5-MeC-DNA glycosylase.  The corresponding oligodeoxyribonucleotide or the
oligoribonucleotide complementary to the non-methylated strand of the same DNA substrate are
inactive when incubated in the complementation test.  A minimum of 4 bases complementary of
the CpG target sequence are necessary for reactivation of RNase-treated 5-MeC-DNA glycosylase.
Complementation with double-stranded oligoribonucleotides does not restore 5-MeC-DNA
glycosylase activity.  An excess of targeting oligoribonucleotides cannot change the
preferential substrate specificity of the enzyme for hemimethylated double-stranded DNA.

<>

<1>Jost, J.-P., Jost, Y.-C.
<2>Transient DNA demethylation in differentiating mouse myoblasts correlates with higher activity of 5-methyldeoxycytidine excision repair.
<3>J. Biol. Chem.
<4>269
<5>10040-10043
<6>1994
<7>It has been recently shown that in developing chicken embryonic nuclear extracts there is a
5-methyldeoxycytidine excision repair activity.  We show that in differentiating mouse
myoblasts, a similar enzymatic reaction may be responsible for the genome-wide DNA
demethylation (up to 50% of all CmCGG) occurring between the 3rd and 5th days of
differentiation.  Furthermore, in differentiating myoblasts, there is first a 50% transient
decrease in DNA methyltransferase activity and a 90% drop in the rate of DNA synthesis,
followed by an increase in 5-methyl-CpG endonuclease and 5-methyldeoxycytidine excision repair
activities.  As tested in vitro, the maximal activity of the 5-methyldeoxycytidine excision
repair coincides with the maximal in vivo genome-wide DNA demethylation.  We also find that
3-aminobenzamide, a potent inhibitor of ADP-ribosyltransferase, blocks the differentiation of
myoblasts, the 5-methyldeoxycytidine excision repair activity, and the genome-wide
demethylation.

<>

<1>Jost, J.-P., Siegmann, M., Sun, L., Leung, R.
<2>Mechanism of DNA demethylation in chicken embryos.
<3>J. Biol. Chem.
<4>270
<5>9734-9739
<6>1995
<7>We have previously shown that in developing chicken embryos and differentiating mouse
myoblasts, the demethylation of 5-metCpGs occurs through the replacement of 5-methylcytosine
by cytosine.  We have now purified over 30,000-fold a 5-methylcytosine-DNA glycosylase from
12-day-old chicken embryos.  The enzyme copurifies with a mismatch-specific thymine-DNA
glycosylase and an apyrimidic-endonuclease.  The reaction product of the highly purified
5-methylcytosine-DNA glycosylase is 5-methylcytosine.  The copurified apyrimidic-endonuclease
activity cleaves 3' from the apyrimidic sugar.  A 52.5-kDa peptide, isolated as a single band
from preparative SDS-polyacrylamide gels, has both the 5-methylcytosine-DNA glycosylase and
the mismatch-specific thymine-DNA glycosylase activities.  5-Methylcytosine-DNA glycosylase
has an apparent pI of 5.5-7.5 and maximal activity between pH 6.5 and 7.5.  The Km for
hemimethylated oligonucleotide substrate is 8 x 10^-8 M with a Vmax of 4 x 10^-11 mol/h/ug
protein.  5-Methylcytosine-DNA glycosylase binds equally well to methylated and non-methylated
DNA.  The enzyme reacts six times faster with the hemimethylated DNA than with the same
bifilarly methylated DNA sequence, and single-stranded methylated DNA is not a substrate.  The
action of the enzyme is distributive.

<>

<1>Jost, J.-P., Siegmann, M., Thiry, S., Jost, Y.-C., Benjamin, D., Schwarz, S.
<2>A re-investigation of the ribonuclease sensitivity of a DNA demethylation reaction in chicken embryo and G8 mouse myoblasts.
<3>FEBS Lett.
<4>449
<5>251-254
<6>1999
<7>Recently published results suggest that the ribonuclease sensitivity of the DNA demethylation
reaction may be an experimental artifact due to the possible tight binding of the nucleases to
the methylated DNA substrate. Using an improved protocol we show for two different systems
that demethylation of hemi-methylated DNA is indeed sensitive to micrococcal nuclease,
requires RNA and is not an experimental artifact.  The purified 5-MeC-DNA glycosylase from
chicken embryos and G8 mouse myoblasts was first incubated for 5 min at 37 C with micrococcal
nuclease in the presence of Ca2+  in the absence of the DNA substrate.  Upon blocking the
nuclease activity by the addition of 25 mM EGTA, the DNA demethylation reaction was initiated
by adding the labeled hemimethylated DNA substrate to the reaction mixture.  Under these
conditions the DNA demethylation reaction was abolished.  In parallel controls, where the
purified 5-MeC-DNA glycosylase was pre-incubated at 37 C with the nuclease, Ca2+ and EGTA or
with the nuclease and EGTA, RNA was not degraded and no inhibition of the demethylation
reaction was obtained.  As had already been shown for chicken embryos, the loss of 5-MeC-DNA
glycosylase activity from G8 myoblasts following nuclease treatment can also be restored by
the addition of synthetic RNA complementary to the methylated strand of the substrate DNA.  No
reactivation of 5-MeC-DNA glycosylase is obtained by complementation with a random RNA
sequence, the RNA sequence complementary to the non-methylated strand or DNA, thus ruling out
a non-specific competition of the RNA for the binding of the nuclease to the labeled DNA
substrate.

<>

<1>Jothikumar, N., Kahler, A., Strockbine, N., Gladney, L., Hill, V.R.
<2>Draft Genome Sequence of Raoultella planticola, Isolated from River Water.
<3>Genome Announcements
<4>2
<5>e01061-14
<6>2014
<7>We isolated Raoultella planticola from a river water sample, which was phenotypically
indistinguishable from Escherichia coli on MI agar. The genome
sequence of R. planticola was determined to gain information about its metabolic
functions contributing to its false positive appearance of E. coli on MI agar. We
report the first whole genome sequence of Raoultella planticola.

<>

<1>Jothikumar, N., Kahler, A., Strockbine, N., Gladney, L., Hill, V.R.
<2>Draft Genome Sequence of Buttiauxella agrestis, Isolated from Surface Water.
<3>Genome Announcements
<4>2
<5>e01060-14
<6>2014
<7>MI agar is routinely used for quantifying Escherichia coli in drinking water. A suspect E.
coli colony isolated from a water sample was identified as
Buttiauxella agrestis. The whole genome sequence of B. agrestis was determined to
understand the genetic basis for its phenotypic resemblance to E. coli on MI
agar.

<>

<1>Jou, T.-W., Chen, C.-S.
<2>Purification and characterization of restriction endonuclease CstI from Clostridium sticklandii.
<3>Bot. Bull. Acad. Sinica
<4>29
<5>201-208
<6>1988
<7>A type II restriction endonuclease CstI was purified to homogeneity from
Clostridium sticklandii by phosphocellulose chromatography and preparative gel
electrophoresis.  It was found that CstI was an isoschizomer of PstI which is
widely used in molecular cloning.  Both CstI and PstI recognize and cleave the
nucleotide sequence 5'-CTGCAG-3' of double stranded DNA.  Nucleotide sequence
analysis revealed that both enzymes split the phosphodiester bond between A and
G.  The molecular weight of CstI determined by disc gel electrophoresis was
apparently 206,000.  The optimal pH, temperature, sodium chloride and magnesium
ion concentrations of CstI were shown to be 7-9, 37-40C, 50-200 mM and 5 mM,
respectively.  CstI is heat-labile.  When incubated at temperature higher than
40C for 5 minutes, the enzyme lost its activity rapidly.

<>

<1>Joung, S.M., Jeon, S.J., Lim, Y.J., Lim, J.S., Choi, B.S., Choi, I.Y., Yu, J.H., Na, K.I., Cho, E.H., Shin, S.S., Park, Y.K., Kim, C.K., Kim, H.J., Ryoo, S.W.
<2>Complete Genome Sequence of Mycobacterium bovis BCG Korea, the Korean Vaccine Strain for Substantial Production.
<3>Genome Announcements
<4>1
<5>e0006913
<6>2013
<7>Mycobacterium bovis bacillus Calmette-Guerin (BCG) is the only vaccine available  against
tuberculosis, and the strains used worldwide represent a family of
daughter strains with distinct genotypic characteristics. Here, we report the
complete genome sequence of M. bovis BCG Korea, the strain that will be actually
used in Korea for vaccine production.

<>

<1>Jousset, A., Schuldes, J., Keel, C., Maurhofer, M., Daniel, R., Scheu, S., Thuermer, A.
<2>Full-Genome Sequence of the Plant Growth-Promoting Bacterium Pseudomonas protegens CHA0.
<3>Genome Announcements
<4>2
<5>e00322-14
<6>2014
<7>We report the complete genome sequence of the free-living bacterium Pseudomonas protegens
(formerly Pseudomonas fluorescens) CHA0, a model organism used in plant-microbe interactions,
biological control of phytopathogens, and bacterial genetics.

<>

<1>Jovanovic, L., Delahunt, B., McIver, B., Eberhardt, N.L., Grebe, S.K.G.
<2>Optimising restriction enzyme cleavage of DNA derived from archival histopathological samples: an improved HUMARA assay.
<3>Pathology
<4>35
<5>70-74
<6>2003
<7>Aims: Our aim was to evaluate and optimise methylation-sensitive restriction enzyme assays for
use in formalin-fixed, paraffin-embedded tissue (FFPET) samples, in order to improve the
application of the HUMARA X-chromosome inactivation assay to FFPET samples.Methods: We
extracted DNA from normal male colon and thyroid FFPET. Several DNA clean-up procedures and
restriction enzyme buffer compositions were tested for two methylation-sensitive enzymes, HhaI
and HpaII and a non-methylation-sensitive isoschizomere, MspI.Results: By including both a
non-methylation-sensitive control enzyme and DNA from male archival specimens in our
experiments, we were able to detect even subtle degrees of incomplete digestion. We showed
that FFPET-derived DNA is a poor substrate for restriction enzymes, especially for
methylation-sensitive restriction endonucleases. An optimised DNA clean-up protocol and
restriction enzyme buffer-mix allowed us to achieve complete digestion.Conclusions: The
combination of multiple, rigorous controls, DNA clean-up and restriction buffer optimisation
increases the reliability of HUMARA-based X-chromosome inactivation analysis of FFPET samples.
Analogous approaches are likely to allow optimisation of other restriction enzyme-based assays
of FFPET samples.

<>

<1>Judge, N.A., Fields, J.A., Pajaniappan, M., Newell, D.G., Manning, J., Burns, C.M., Thompson, S.A.
<2>Mutation of a Campylobacter jejuni DNA methylase affects expression of multiple genes, motility, and epithelial cell adherence and invasion.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>105
<5>79
<6>2005
<7>Campylobacter jejuni causes over 2 million cases of severe bacterial gastroenteritis in the
U.S. every year. Poultry flocks are ubiquitously and asymptomatically colonized with C.
jejuni, and the most likely cause of human infection is via the consumption of contaminated
poultry meat products as well as other foods cross contaminated during preparation, and
contaminated water. C. jejuni is able to thrive at two different temperatures, 42ºC (the core
temperature of chickens) and 37ºC (the core temperature of humans), thus there is likely to be
temperature regulation of Campylobacter proteins for optimization of expression in each
environment, including virulence factors important for human infection. We identified an
ortholog of NCTC11168 cj1461 in C. jejuni strain 81-176 as a gene that was more highly
expressed at 37ºC than at 42ºC. cj1461 encodes a predicted DNA methylase that is not
associated with a cognate restriction endonuclease. We purified Cj1461 as a recombinant
His-tagged protein and showed that Cj1461 can bind DNA and has DNA methyltransferase activity.
We hypothesize that Cj1461 does not protect C. jejuni DNA from restriction endonucleases and
may be involved in the regulation of genes important for the virulence of the bacterium. We
inactivated the cj1461 ortholog in C. jejuni 81-176 to investigate its potential role in gene
regulation. Proteome and DNA array results comparing 81-176 with its isogenic 81-176cj1461
mutant indicated the deregulation of numerous genes, including many involved in
virulence-associated pathways such as motility, flagellar glycosylation, chemotaxis, oxidative
stress, iron uptake, and transformation competence. The 81-176cj1461 mutant was significantly
less motile than C. jejuni 81-176, and was 30 times more adherent, but 800 times less invasive
than wild-type in an in vitro INT407 tissue culture model. These results suggest that the
regulation of multiple genes in C. jejuni 81-176, including some important for virulence, can
be mediated in part by the DNA methyltransferase Cj1461.

<>

<1>Jue, K., Bestor, T., Trasler, J.M.
<2>Differential expression of DNA methyltransferase in male germ cells.
<3>Biol. Reprod.
<4>50
<5>80
<6>1994
<7>DNA methylation is involved in regulating gene expression and plays a critical role in normal
development. Patterns of methylation are believed to be established during gametogenesis and
early embryogenesis. To better understand how methylation patterns are established in the male
mouse germ line we focused on the expression of the DNA methyltransferase (DNA MTase) which
catalyzes the transfer of methyl groups from S-adenosyl-methionine to the 5' position of
cytosine in DNA. Purified populations of premeiotic, meiotic and postmeiotic germ cells were
isolated from adult and 17 day old mouse testis using cellular sedimentation at unit gravity
on a Staput apparatus. Western analysis revealed abundant DNA MTase protein in postmitotic
leptotene/zygotene spermatocyte and haploid round spermatid fractions but low levels in
pachytene spermatocyte fractions. A 6.2kb DNA MTase transcript is specific to pachytene
spermatocytes whereas in other cell types, a 5.2kb mRNA is present. These findings are
supported by immunofluorescence studies where germ cells were stained for DNA MTase.
Immunofluorescence further revealed DNA MTase staining patterns to vary within one isolated
germ cell population suggesting differential expression and localization between the
developmental steps in various cell types. We conclude that DNA MTase expression is regulated
during spermatogenesis at a time when de novo methylation is known to occur. These studies
contribute to our understanding of how methylation patterns are established in the male germ
line and their importance during early development.

<>

<1>Juez, G., Rodriguez-Valera, F., Herrero, N., Mojica, F.J.M.
<2>Evidence for salt-associated restriction pattern modifications in the Archaeobacterium Haloferax mediterranei.
<3>J. Bacteriol.
<4>172
<5>7278-7281
<6>1990
<7>DNA restriction pattern modifications were detected when Haloferax mediterranei
was grown in low (10%) salt concentrations.  After cells were grown again in
optimal (25%) salt concentrations, the original pattern was recovered.  These
salt-associated DNA modifications were revealed with 5% of the 160 DNA
fragments cloned and used as probes in hybridization experiments.  Patterns
obtained when genomic DNA was digested with different restriction enzymes
showed that these modifications are related not to insertions or deletions in
the genome but to modifications of some specific sequences.

<>

<1>Juhala, R.J., Ford, M.E., Duda, R.L., Youlton, A., Hatfull, G.F., Hendrix, R.W.
<2>Genomic sequences of bacteriophages HK97 and HK022: pervasive genetic mosaicism in the lambdoid bacteriophages.
<3>J. Mol. Biol.
<4>299
<5>27-51
<6>2000
<7>We report the complete genome DNA sequences of HK97 (39,732 bp) and HK022
(40,751 bp), double-stranded DNA bacteriophages of Escherichia coli and
members of the lambdoid or lambda-like group of phages. We provide a
comparative analysis of these sequences with each other and with two
previously determined lambdoid family genome sequences, those of E. coli
phage lambda and Salmonella typhimurium phage P22. The comparisons confirm
that these phages are genetic mosaics, with mosaic segments separated by
sharp transitions in the sequence. The mosaicism provides clear evidence
that horizontal exchange of genetic material is a major component of
evolution for these viruses. The data suggest a model for evolution in
which diversity is generated by a combination of illegitimate and
homologous recombination and mutational drift, and selection for function
produces a population in which most of the surviving mosaic boundaries are
located at gene boundaries or, in some cases, at protein domain boundaries
within genes. Comparisons of these genomes highlight a number of
differences that allow plausible inferences of specific evolutionary
scenarios for some parts of the genome. The comparative analysis also
allows some inferences about function of genes or other genetic elements.
We give examples for the generalized recombination genes of HK97, HK022
and P22, and for a putative headtail adaptor protein of HK97 and HK022. We
also use the comparative approach to identify a new class of genetic
elements, the morons, which consist of a protein-coding region flanked by
a putative delta 70 promoter and a putative factor-independent
transcription terminator, all located between two genes that may be
adjacent in a different phage. We argue that morons are autonomous genetic
modules that are expressed from the repressed prophage. Sequence
composition of the morons implies that they have entered the phages'
genomes by horizontal transfer in relatively recent evolutionary time.

<>

<1>Julio, S.M., Heithoff, D.M., Provenzano, D., Klose, K.E., Sinsheimer, R.L., Low, D.A., Mahan, M.J.
<2>DNA Adenine Methylase Is Essential for Viability and Plays a Role in the Pathogenesis of Yersinia pseudotuberculosis and Vibrio cholerae.
<3>Infect. Immun.
<4>69
<5>7610-7615
<6>2001
<7>Salmonella strains that lack or overproduce DNA adenine methylase (Dam) elicit a protective
immune response to different Salmonella species. To generate vaccines against other bacterial
pathogens, the dam genes of Yersinia pseudotuberculosis and Vibrio cholerae were disrupted but
found to be essential for viability. Overproduction of Dam significantly attenuated the
virulence of these two pathogens, leading to, in Yersinia, the ectopic secretion of virulence
proteins (Yersinia outer proteins) and a fully protective immune response in vaccinated hosts.
Dysregulation of Dam activity may provide a means for the development of vaccines against
varied bacterial pathogens.

<>

<1>Julio, S.M., Heithoff, D.M., Sinsheimer, R.L., Low, D.A., Mahan, M.J.
<2>DNA adenine methylase overproduction in Yersinia pseudotuberculosis alters YopE expression and secretion and host immune responses to  infection.
<3>Infect. Immun.
<4>70
<5>1006-1009
<6>2002
<7>Yersinia pseudotuberculosis mutants that overproduce the DNA adenine methylase (Dam) are
highly attenuated, confer fully protective immune
responses, and secrete several Yersinia virulence proteins (Yersinia
outer proteins [Yops]) under conditions that are nonpermissive for
secretion in wild-type strains. We examined here the effects of Dam
overproduction on Yersinia virulence determinant expression and
secretion, as well as the host immune response to Yersinia antigens.
Western blot analysis with convalescent antisera identified several
low-calcium-responsive antigens whose synthesis was affected by Dam
overproduction. One of these antigens was shown to be the type III
secretion effector protein, YopE, a cytotoxin involved in
antiphagocytosis. Dam overproduction disrupted both the thermal and
calcium regulation of YopE synthesis and relaxed the thermal but not
the calcium dependence of YopE secretion. Altered expression and/or
secretion of Yersinia proteins in Dam-overproducing strains may
contribute to the decreased virulence and heightened immunity observed
in vaccinated hosts and may provide a means by which to deliver
heterologous antigens and/or immune modulators of the inflammatory
response.

<>

<1>Jullien, P.E., Susaki, D., Yelagandula, R., Higashiyama, T., Berger, F.
<2>DNA methylation dynamics during sexual reproduction in Arabidopsis thaliana.
<3>Curr. Biol.
<4>22
<5>1825-1830
<6>2012
<7>DNA methylation maintains genome stability and regulates gene expression [1]. In  mammals, DNA
methylation is reprogrammed in the germline from one generation to
the next [2]. In plants, it was considered that patterns of DNA methylation are
stably maintained through sexual reproduction [3-6]. However, a recent report
showed discrete variations of DNA methylation profiles from mother to daughter
plants [7]. The mechanisms that explain these variations have remained unknown.
Here, we report that maintenance DNA methyltransferases are barely expressed
during Arabidopsis female gametogenesis. In contrast, after fertilization both
maintenance and de novo DNA methyltransferases are expressed strongly in the
embryo. Embryogenesis is marked by increased de novo DNA methylation, reaching
levels that are further maintained in the adult plant. The accumulation of these
epigenetic marks after fertilization silences a methylation-sensitive fluorescent
reporter. De novo DNA methylation in the embryo provides a mechanism that could
account for the gradual remethylation of experimentally demethylated genomes [8,
9]. In conclusion, we uncover that DNA methylation activity fluctuates during
sexual reproduction. This cycle likely explains variations of genome-wide
patterns of DNA methylation across generations in Arabidopsis [7, 10] and enables
a limited degree of reprogramming of the epigenome.

<>

<1>Jun, H.S., Kim, Y.S., Choi, K.R., Rho, H.M.
<2>Cloning and expression of the BdiI methylase gene in E. coli.
<3>Korean J. Microbiol.
<4>25
<5>40-45
<6>1987
<7>The gene for the BdiI modification enzyme, which is one of BdiI
restriction-modification system, from Brevibacterium divaricatum FERM 5948 was
cloned and expressed in E. coli.  For cloning of the BdiI methylase gene, we
have initially used three cloning site (EcoRI, BamHI and SalI) of plasmid
vector pBR322 and adopted the retransformation method after BdiI restriction
endonuclease cleavage.  Selection of transformants carrying the gene was based
on the resistance of the modified plasmid encoding the enzyme to cleavage by
BdiI restriction enzyme, and the recombinant plasmid pBDIM116 containing 5.6 kb
EcoRI insert was proved to carry the gene.  Crude cell extracts prepared from
strains carrying the plasmid pBDIM116 contained an
S-adenosylmethionine-dependent methyltransferase activity specific for the BdiI
recognition site, ATCGAT.  The restriction map was constructed with 11
restriction enzyme, and the BdiI restriction-modification system was also
discussed.

<>

<1>Jun, J.W., Kim, J.H., Choresca, C.H., Shin, S.P., Han, J.E., Park, S.C.
<2>Draft Genome Sequence of Vibrio parahaemolyticus SNUVpS-1 Isolated from Korean Seafood.
<3>Genome Announcements
<4>1
<5>e00132-12
<6>2013
<7>Vibrio parahaemolyticus is the leading cause of food-borne diseases, and several  pathogenic
strains cause global gastroenteritis outbreaks. Here, we report a
draft genome sequence of V. parahaemolyticus SNUVpS-1, which was isolated from
seafood in a fishery market in the Republic of Korea and contained TL, toxR, and
toxRS(old) genes. The current draft genome sequence will contribute to the effort
to monitor the spread of V. parahaemolyticus seafood isolates and clinical
isolates.

<>

<1>Jun, X., Lupeng, L., Minjuan, X., Oger, P., Fengping, W., Jebbar, M., Xiang, X.
<2>Complete Genome Sequence of the Obligate Piezophilic Hyperthermophilic Archaeon Pyrococcus yayanosii CH1.
<3>J. Bacteriol.
<4>193
<5>4297-4298
<6>2011
<7>Pyrococcus yayanosii CH1 is the first obligate piezophilic hyperthermophilic archaeon isolated
from the deep-sea hydrothermal site
Ashadze on the mid-Atlantic ridge at a depth of 4,100 m. This organism
grows within a temperature range of 80 to 108 degrees C and a hydrostatic
pressure range of 20 to 120 MPa, with optima at 98 degrees C and 52 MPa,
respectively. Here, we report the complete genome sequence (1,716,817 bp,
with a G+C content of 51.6%) of the type strain P. yayanosii CH1(T) (= JCM
16557). This genomic information reveals a systematic view of the
piezoadaptation strategy and evolution scenario of metabolic pathways in
Thermococcales.

<>

<1>Jung, J., Baek, J.H., Park, W.
<2>Complete genome sequence of the diesel-degrading Acinetobacter sp. strain DR1.
<3>J. Bacteriol.
<4>192
<5>4794-4795
<6>2010
<7>The genus Acinetobacter is ubiquitous in soil, aquatic, and sediment environments and includes
pathogenic strains, such as A. baumannii. Many
Acinetobacter species isolated from various environments have
biotechnological potential since they are capable of degrading a variety
of pollutants. Acinetobacter sp. strain DR1 has been identified as a
diesel degrader. Here we report the complete genome sequence of
Acinetobacter sp. DR1 isolated from the soil of a rice paddy.

<>

<1>Jung, J., Choi, S., Chun, J., Park, W.
<2>Genome Sequence of Pectin-Degrading Alishewanella aestuarii Strain B11T, Isolated from Tidal Flat Sediment.
<3>J. Bacteriol.
<4>194
<5>5476
<6>2012
<7>We present the genome sequence of Alishewanella aestuarii B11(T) (=KCTC 22051(T)=DSM
19476(T)). This species, isolated from tidal flat sediment, was
reported to be a novel species. A. aestuarii is known to degrade pectin, an
important component of plant cell wall. The presence of the genes related to
pectin metabolism in this strain indicates its capability to utilize pectin.

<>

<1>Jung, J., Chun, J., Park, W.
<2>Genome Sequence of Extracellular-Protease-Producing Alishewanella jeotgali Isolated from Traditional Korean Fermented Seafood.
<3>J. Bacteriol.
<4>194
<5>2097
<6>2012
<7>Alishewanella jeotgali MS1(T) (= KCTC 22429(T) = JCM 15561(T)) was isolated from  a
traditional Korean fermented seafood, gajami sikhae (jeotgal), and has been
reported as a novel species. A. jeotgali was proven to have extracellular
proteolytic activity, which may play an important role in the fermentation
environment of food containing fish flesh. Here, we present the genome sequence
of Alishewanella jeotgali MS1(T) as the first sequenced strain in the genus
Alishewanella and its taxonomic relatives.

<>

<1>Jung, J.H., Holden, J.F., Seo, D.H., Park, K.H., Shin, H., Ryu, S., Lee, J.H., Park, C.S.
<2>Complete Genome Sequence of the Hyperthermophilic Archaeon Thermococcus sp. Strain CL1, Isolated from a Paralvinella sp. Polychaete Worm Collected from a  Hydrothermal Vent.
<3>J. Bacteriol.
<4>194
<5>4769-4770
<6>2012
<7>Thermococcus sp. strain CL1 is a hyperthermophilic, anaerobic, and heterotrophic  archaeon
isolated from a Paralvinella sp. polychaete worm living on an active
deep-sea hydrothermal sulfide chimney on the Cleft Segment of the Juan de Fuca
Ridge. To further understand the distinct characteristics of this archaeon at the
genome level, its genome was completely sequenced and analyzed. Here, we announce
the complete genome sequence (1,950,313 bp) of Thermococcus sp. strain CL1, with
a focus on H(2)- and energy-producing capabilities and its amino acid
biosynthesis and acquisition in an extreme habitat.

<>

<1>Jung, J.H., Lee, J.H., Holden, J.F., Seo, D.H., Shin, H., Kim, H.Y., Kim, W., Ryu, S., Park, C.S.
<2>Complete Genome Sequence of the Hyperthermophilic Archaeon Pyrococcus sp. Strain  ST04, Isolated from a Deep-Sea Hydrothermal Sulfide Chimney on the Juan de Fuca  Ridge.
<3>J. Bacteriol.
<4>194
<5>4434-4435
<6>2012
<7>Pyrococcus sp. strain ST04 is a hyperthermophilic, anaerobic, and heterotrophic archaeon
isolated from a deep-sea hydrothermal sulfide chimney on the Endeavour
Segment of the Juan de Fuca Ridge in the northeastern Pacific Ocean. To further
understand the distinct characteristics of this archaeon at the genome level
(polysaccharide utilization at high temperature and ATP generation by a Na(+)
gradient), the genome of strain ST04 was completely sequenced and analyzed. Here,
we present the complete genome sequence analysis results of Pyrococcus sp. ST04
and report the major findings from the genome annotation, with a focus on its
saccharolytic and metabolite production potential.

<>

<1>Jung, J.Y., Lee, S.H., Jeon, C.O.
<2>Complete Genome Sequence of Leuconostoc carnosum Strain JB16, Isolated from Kimchi.
<3>J. Bacteriol.
<4>194
<5>6672-6673
<6>2012
<7>Leuconostoc carnosum strain JB16 was isolated from kimchi, the traditional Korean fermented
food. Here, we report the complete genome sequence of L. carnosum
strain JB16, consisting of a 1,645,096-bp circular chromosome with a G+C content
of 37.24% and four plasmids.

<>

<1>Jung, J.Y., Lee, S.H., Jeon, C.O.
<2>Complete Genome Sequence of Leuconostoc gelidum Strain JB7, Isolated from Kimchi.
<3>J. Bacteriol.
<4>194
<5>6665
<6>2012
<7>A strain of Leuconostoc gelidum, designated strain JB7, was isolated from kimchi, the
representative Korean traditional fermented food. Here we announce the
complete genome sequence of L. gelidum strain JB7, consisting of a 1,893,499-bp
circular chromosome with a G+C content of 36.68%, and provide a description of
its annotation.

<>

<1>Jung, J.Y., Lee, S.H., Lee, S.H., Jeon, C.O.
<2>Complete Genome Sequence of Leuconostoc mesenteroides subsp. mesenteroides Strain J18, Isolated from Kimchi.
<3>J. Bacteriol.
<4>194
<5>730-731
<6>2012
<7>Leuconostoc mesenteroides subsp. mesenteroides is one of the most predominant lactic acid
bacterial groups during kimchi fermentation. Here,
we report the complete genome sequence of L. mesenteroides subsp.
mesenteroides J18, which was isolated from kimchi. The genome of the
strain consists of a 1,896,561-bp chromosome and five plasmids.

<>

<1>Jung, M.J., Roh, S.W., Kim, M.S., Whon, T.W., Bae, J.W.
<2>Genome Sequence of Lentibacillus jeotgali GrbiT, Isolated from Traditional Korean Salt-Fermented Seafood.
<3>J. Bacteriol.
<4>193
<5>6414-6415
<6>2011
<7>Lentibacillus jeotgali Grbi(T), isolated from a traditional Korean salt-fermented seafood, is
a strictly aerobic, Gram-positive, nonmotile,
endospore-forming, moderately halophilic bacterium belonging to the family
Bacillaceae in the phylum Firmicutes. Here, the draft genome sequence of
L. jeotgali Grbi(T) (3,775,822 bp with a G+C content of 42.5%) is
reported. This is the first reported genome sequence from a Lentibacillus
species.

<>

<1>Jung, V., Pestka, S.B., Pestka, S.
<2>Efficient cloning of PCR generated DNA containing terminal restriction endonuclease recognition sites.
<3>Nucleic Acids Res.
<4>18
<5>6156
<6>1990
<7>None

<>

<1>Jung, V., Pestka, S.B., Pestka, S.
<2>Cloning of polymerase chain reaction-generated DNA containing terminal restriction endonuclease recognition sites.
<3>Methods Enzymol.
<4>218
<5>357-362
<6>1993
<7>Using the polymerase chain reaction (PCR) to generate DNA containing terminal restriction
endonuclease recognition sites to permit cloning usually relies on the use of unphosphorylated
primers incorporating a restriction endonuclease recognition site of choice plus three or four
extra 5' bases flanking that site. Various sites (e.g., NotI, XhoI, and XbaI) incorporated
into the termini of PCR products have proved difficult to cut with their respective
restriction endonucleases. There are several possible explanations for this difficulty; first,
Taq polymersase might be inefficient for certain terminal sequences, producing frayed ends
that cannot be cleaved by the restriction endonuclease. Alternatively, the "breathing" of
terminal sequences might prevent the stable association of restriction endonucleases with
terminal sites. Also, Taq polymerase or other contaminants might bind to the ends of the PCR
products, blocking restriction endonuclease activity.

<>

<1>Juranek, S., Wieden, H.-J., Lipps, H.J.
<2>De novo cytosine methylation in the differentiating macronucleus of the stichotrichous ciliate Stylonychia lemnae.
<3>Nucleic Acids Res.
<4>31
<5>1387-1391
<6>2003
<7>Dramatic DNA reorganization and elimination processes occur during macronuclear
differentiation in ciliates. In this study we analyzed
whether cytosine methylation of specific sequences plays a functional role
during DNA rearrangement. Three classes of sequences,
macronuclear-destined sequences (MDSs, pCE7), members from a large family
of transposon-like elements and micronuclear-specific sequences (pLJ01),
differing in their structure and future destiny during nuclear
differentiation, were studied in the micronucleus, the developing
macronucleus and, when present, in the mature macronucleus. While the MDSs
become processed to a 1.1 and 1.3 kb gene-sized macronuclear DNA molecule,
the family of transposon-like elements represented by MaA81 becomes
removed late in the course of polytene chromosome formation. The
micronuclear-specific sequence pLJ01 is eliminated together with bulk
micronuclear DNA during degradation of polytene chromosomes. No methylated
cytosine could be detected in the vegetative macronucleus and no
difference in methylation pattern was observed either between micronucleus
and developing macronucleus in MDSs or in a micronuclear-specific
sequence. However, a significant percentage of the cytosines contained in
the transposon-like element becomes methylated de novo in the course of
macronuclear differentiation. This is the first demonstration that
cytosine methylation in specific sequences occurs during macronuclear
differentiation and may provide a first step towards understanding
epigenetic factors involved in DNA processing.

<>

<1>Jurenaite-Urbanaviciene, S.
<2>Engineering of bifunctional restriction endonucleases with novel specificities.
<3>Ph.D. Thesis, Vilnius University
<4>
<5>1-46
<6>2008
<7>CONCLUSIONS 1. The BseMII restriction-modification system is composed of two enzymes, a DNA
methlytransferase and a restriction endonuclease, which are encoded by two convergently
transcribed genes. The PpiI and TstI R-M systems are each composed of a single enzyme. 2.
Regions predicted to participate in DNA target hydrolysis, methylation and recognition were
identified in the polypeptide of BseMII restriction endonuclease. From the viewpoint of the
structure-function organization, the BseMII REase is a suitable object for experiments aimed
at changing specificity. 3. The structure-function organization of PpiI and TstI resemble that
of AloI. Regions responsible for DNA target hydrolysis, methylation and recognition were
identified in polypeptides of these enzymes. 4. The construction of active hybrids between
AloI and PpiI has demonstrated that C-terminal regions of type IIB REases are involved in DNA
target recognition and revealed that proximal target recognition domains are independent
interchangeable modules. 5. Active hybrids recognizing novel DNA targets GAGN5GTG were
generated by swapping of proximal and distal target recognition domains between PpiI and TstI.
These results have demonstrated that novel type IIB REases can be engineered by recombination
of their TRDs. 6. The increase in the Tst-PpiTRD1 activity resulting from a single amino acid
substitution Gly1006Leu demonstrates that properties of hybrid REases can be improved by
rational mutagenesis. 7. Hybrid type IIB enzymes similarly to progenitor restriction
endonucleases cleave DNA on both sides of recognition sequences and display cleavage site
variability: at some positions enzymes hydrolyze DNA at two points differing from each other
by one nucleotide.

<>

<1>Jurenaite-Urbanaviciene, S., Kazlauskiene, R., Urbelyte, V., Maneliene, Z., Petrusyte, M., Lubys, A., Janulaitis, A.
<2>Characterization of BseMII, a new type IV restriction-modification system, which recognizes the pentanucleotide sequence 5'-CTCAG(N)10/8.
<3>Nucleic Acids Res.
<4>29
<5>895-903
<6>2001
<7>We report the properties of the new BseMII restriction and modification enzymes from Bacillus
stearothermophilus Isl 15-111, which recognize the sequence 5'-CTCAG, and the nucleotide
sequence of the genes encoding them. The restriction endonuclease R.BseMII makes a staggered
cut at the tenth base pair downstream of the recognition sequence on the upper strand,
producing a two base 3'-protruding end.  Magnesium ions and S-adenosyl-L-methionine (AdoMet)
are required for cleavage.  S-adenosylhomocysteine and sinefungin can replace AdoMet in the
cleavage reaction. The BseMII methyltransferase modifies unique adenine residues in both
strands of the target sequence 5'-CTCAG-3'/5'-CTGAG-3'. Monomeric R.BseMII in addition to
endonucleolytic activity also possesses methyltransferase activity that modifies the A base
only within the 5'-CTCAG strand of the target duplex. The deduced amino acid sequence of the
restriction endonuclease contains conserved motifs of DNA N6-adenine methylases involved in
S-adenosyl-L-methionine binding and catalysis. According to its structure and enzymatic
properties, R.BseMII may be regarded as a representative of the type IV restriction
endonucleases.

<>

<1>Jurenaite-Urbanaviciene, S., Serksnaite, J., Kriukiene, E., Giedriene, J., Venclovas, C., Lubys, A.
<2>Generation of DNA cleavage specificities of type II restriction endonucleases by reassortment of target recognition domains.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>10358-10363
<6>2007
<7>Type II restriction endonucleases (REases) cleave double-stranded DNA at specific sites within
or close to their recognition sequences. Shortly
after their discovery in 1970, REases have become one of the primary tools
in molecular biology. However, the list of available specificities of type
II REases is relatively short despite the extensive search for them in
natural sources and multiple attempts to artificially change their
specificity. In this study, we examined the possibility of generating
cleavage specificities of REases by swapping putative target recognition
domains (TRDs) between the type IIB enzymes AloI, PpiI, and TstI. Our
results demonstrate that individual TRDs recognize distinct parts of the
bipartite DNA targets of these enzymes and are interchangeable. Based on
these properties, we engineered a functional type IIB REase having
previously undescribed DNA specificity. Our study suggests that the
TRD-swapping approach may be used as a general technique for the
generation of type II enzymes with predetermined specificities.

<>

<1>Jurgaitis, A.P., Butkus, V.V., Klimasauskas, S.J., Janulaitis, A.
<2>The effects of N4-methylcytosine and 5-methylcytosine on the thermal stability of DNA double helix.
<3>Bioorg. Khim.
<4>14
<5>158-165
<6>1988
<7>The thermodynamic parameters (DeltaH, DeltaS) of the helix-coil transition of
self-complementary oligonucleotides d(CGCGCGCG), d(CG5mCGCGCG), d(CG4mCGCGCG),
d(GGACCCGGGTCC), d(GGA5mCCCGGGTCC), and d(GGA4mCCCGGGTCC) were determined.  The
substitution of 4mC for C was found to decrease the melting temperature of the
oligonucleotides.  The destabilization effect of the two substitutions is
equivalent to the change of an A.T for a G.C pair.  The free energy decrease of
helix-coil transition due to the introduction of two 4mC into an octanucleotide
was estimated to be 1.24 kcal/mol.

<>

<1>Jurica, M.S.
<2>I. Structures of  intron encoded homing endonucleases and II. Allosteric regulation of pyruvate kinase.
<3>Ph.D. Thesis, University of Washington, , , Seattle, USA
<4>
<5>1-146
<6>1999
<7>I.  Homing is the lateral transfer of an intervening genetic sequence,
either an intron or an intein, to a cognate allele lacking the element resulting
in the duplication of the intervening sequence.  The process of homing is
initiated by site-specific endonucleases that are encoded by open reading frames
within the mobile elements.  Several features of these proteins make them
attractive subjects for structural studies.  First, these unique endonucleases
may be contrasted with a variety of enzymes involved in nucleic acid strand
breakage and rearrangement, particularly restriction endonucleases.  Second,
because they are encoded within the intervening sequence, limitations in the
length of the open reading frames that encode them are also imposed on the
folded protein structures.  Third, these enzymes display a unique strategy of
flexible recognition of very long DNA target sites.  This strategy allows the
enzymes to minimize non-specific cleavage within the host genome, while
maximizing their ability to cleave closely related variants of the homing
recognition site.  The structural studies of homing endonucleases presented here
provide insight to their unique mechanisms of recognition and cleavage of DNA.
II.  The activation of regulated isozymes of pyruvate kinase (PK) by fructose-
1,6-bis-phosphate (FBP) directly influences cytosolic levels of ATP and GTP and
the flux of glycolytic carbon into fermentation pathways and the Krebs cycle.
This pattern of regulation may be important to cell proliferation, as
demonstrated by the re-expression of an allosterically regulated, fetal isozyme
of PK in tumor cells.  The structure of allosterically regulated pyruvate kinase
from Saccharomyces cerevisiae complexed with phosphoglycolate (a substrate
analogue), Mn2+, and K+ in the presence and absence of FBP was solved to
identify the allosteric binding site.  The location, structure, and interactions
within the allosteric site are in agreement with the pattern of alternate
genetic splicing that leads to regulated forms of the enzyme in humans.
Deregulating the allosteric control of PK in yeast by either site-directed
mutation or expression of non-regulated isozymes has profound effects on cell
growth and cell-cycle profiles.

<>

<1>Jurica, M.S., Monnat, R.J. Jr., Stoddard, B.L.
<2>DNA recognition and cleavage by the LAGLIDADG homing endonuclease I-CreI.
<3>Mol. Cell
<4>2
<5>469-476
<6>1998
<7>The structure of the LAGLIDADG intron-encoded homing endonuclease I-CreI bound to homing site
DNA has been determined. The interface is formed by an extended, concave beta sheet from each
enzyme monomer that contacts each DNA half-site, resulting in direct side-chain contacts to 18
of the 24 base pairs across the full-length homing site. The structure indicates that I-CreI
is optimized to its role in genetic transposition by exhibiting long site-recognition while
being able to cleave many closely related target sequences. DNA cleavage is mediated by a
compact pair of active sites in the I-CreI homodimer, each of which contains a separate bound
divalent cation.

<>

<1>Jurica, M.S., Stoddard, B.L.
<2>Homing endonucleases: structure, function and evolution.
<3>Cell. Mol. Life Sci.
<4>55
<5>1304-1326
<6>1999
<7>'Homing' is the lateral transfer of an intervening genetic sequence, either an intron or an
intein, to a cognate allele that lacks that element. The end result of homing is the
duplication of the intervening sequence. The process is initiated by site-specific
endonucleases that are encoded by open reading frames within the mobile elements. Several
features of these proteins make them attractive subjects for structural and functional
studies. First, these endonucleases, while unique, may be contrasted with a variety of enzymes
involved in nucleic acid strand breakage and rearrangement, particularly restriction
endonucleases. Second, because they are encoded within the intervening sequence, there are
interesting limitations on the position and length of their open reading frames, and therefore
on their structures. Third, these enzymes display a unique strategy of flexible recognition of
very long DNA target sites. This strategy allows these sequences to minimize nonspecific
cleavage within the host genome, while maximizing the ability of the endonuclease to cleave
closely related variants of the homing site. Recent studies explain a great deal about the
biochemical and genetic mechanisms of homing, and also about the structure and function of
several representative members of the homing endonuclease families.

<>

<1>Jurkowska, R.Z., Anspach, N., Urbanke, C., Jia, D., Reinhardt, R., Nellen, W., Cheng, X., Jeltsch, A.
<2>Formation of nucleoprotein filaments by mammalian DNA methyltransferase Dnmt3a in complex with regulator Dnmt3L.
<3>Nucleic Acids Res.
<4>36
<5>6656-6663
<6>2008
<7>The C-terminal domains of Dnmt3a and Dnmt3L form elongated heterotetramers (3L-3a-3a-3L).
Analytical ultracentrifugation confirmed the Dnmt3a-C/3L-C
complex exists as a 2:2 heterotetramer in solution. The 3a-3a interface is
the DNA-binding site, while both interfaces are essential for AdoMet
binding and catalytic activity. Hairpin bisulfite analysis shows
correlated methylation of two CG sites in a distance of approximately 8-10
bp in the opposite DNA strands, which corresponds to the geometry of the
two active sites in one Dnmt3a-C/3L-C tetramer. Correlated methylation was
also observed for two CG sites at similar distances in the same DNA
strand, which can be attributed to the binding of two tetramers next to
each other. DNA-binding experiments show that Dnmt3a-C/3L-C complexes
multimerize on the DNA. Scanning force microscopy demonstrates filament
formation rather than binding of single tetramers and shows that
protein-DNA filament formation leads to a 1.5-fold shortening of the DNA
length.

<>

<1>Jurkowska, R.Z., Ceccaldi, A., Zhang, Y., Arimondo, P.B., Jeltsch, A.
<2>DNA Methyltransferase Assays.
<3>Methods Mol. Biol.
<4>791
<5>157-177
<6>2011
<7>DNA methyltransferases are important enzymes and their inhibition has many potential
applications. The investigation of DNA methyltransferases as well as screening for potential
inhibitors requires specialized enzyme assays. In this chapter, we describe three DNA
methyltransferase assays, each of them based on a different method: (1) An assay using
radioactively labeled Ado Met and biotinylated DNA substrates that is ideal for enzymatic
characterization of these enzymes. (2) An assay using bisulfite conversion of in vitro
methylated DNA that is ideal to determine details of the methylation pattern introduced by
DNA-(cytosine C5)-methyltransferases. (3) A novel fluorescence-coupled, restriction-based
assay suitable for high-throughput screening of DNA methyltransferase inhibitors.

<>

<1>Jurkowska, R.Z., Jurkowski, T.P., Jeltsch, A.
<2>Structure and Function of Mammalian DNA Methyltransferases.
<3>Chembiochem
<4>12
<5>206-222
<6>2011
<7>DNA methylation plays an important role in epigenetic signalling, having an impact on gene
regulation, chromatin structure, development
and disease. Here, we review the structures and functions of the
mammalian DNA methyltransferases Dnmt1, Dnmt3a and Dnmt3b, including
their domain structures, catalytic mechanisms, localisation,
regulation, post-translational modifications and interaction with
chromatin and other proteins, summarising data obtained in genetic,
cell biology and enzymatic studies. We focus on the question of how the
molecular and enzymatic properties of these enzymes are connected to
the dynamics of DNA methylation patterns and to the roles the enzymes
play in the processes of de novo and maintenance DNA methylation.
Recent enzymatic and genome-wide methylome data have led to a new model
of genomic DNA methylation patterns based on the preservation of
average levels of DNA methylation in certain regions, rather than the
methylation states of individual CG sites.

<>

<1>Jurkowska, R.Z., Siddique, A.N., Jurkowski, T.P., Jeltsch, A.
<2>Approaches to Enzyme and Substrate Design of the Murine Dnmt3a DNA Methyltransferase.
<3>Chembiochem
<4>12
<5>1589-1594
<6>2011
<7>
<>

<1>Jurkowski, T.P., Anspach, N., Kulishova, L., Nellen, W., Jeltsch, A.
<2>The M.EcoRV DNA-(adenine N-6)-methyltransferase uses DNA bending for recognition of an expanded EcoDam recognition site.
<3>J. Biol. Chem.
<4>282
<5>36942-36952
<6>2007
<7>The M. EcoRV DNA methyltransferase recognizes GATATC sites. It is related to EcoDam, which
methylates GATC sites. The DNA binding domain
of M. EcoRV is similar to that of EcoDam suggesting a similar mechanism
of DNA recognition. We show that amino acid residue Lys(11) of M. EcoRV
is involved in recognition of Gua(1) and Arg(128) contacts the Gua in
base pair 6. These residues correspond to Lys(9) and Arg(124) in
EcoDam, which recognize the Gua residues in both strands of the Dam
recognition sequence, indicating that M. EcoRV and EcoDam make similar
contacts to outermost base pairs of their recognition sequences and M.
EcoRV recognizes its target site as an expanded GATC site. In contrast
to EcoDam, M. EcoRV considerably bends the DNA (59 +/- 4 degrees)
suggesting indirect readout of the AT-rich inner sequence. Recognition
of an expanded target site by DNA bending is a new principle for
changing DNA recognition specificity of proteins during molecular
evolution. R128A is inefficient in DNA bending and binding, whereas
K11A bends DNA with relaxed sequence specificity. These results suggest
a temporal order of the formation of protein-DNA contacts in which the
Gua(6)-Arg(128) contact forms early followed by DNA bending and,
finally, the formation of the Lys(11)-Gua(1) contact.

<>

<1>Jurkowski, T.P., Jeltsch, A.
<2>On the Evolutionary Origin of Eukaryotic DNA Methyltransferases and Dnmt2.
<3>PLoS ONE
<4>6
<5>e28104
<6>2011
<7>The Dnmt2 enzymes show strong amino acid sequence similarity with eukaryotic and prokaryotic
DNA-(cytosine C5)-methyltransferases. Yet,
Dnmt2 enzymes from several species were shown to methylate tRNA-Asp and
had been proposed that eukaryotic DNA methyltransferases evolved from a
Dnmt2-like tRNA methyltransferase ancestor [Goll et al., 2006, Science,
311, 395-8]. It was the aim of this study to investigate if this
hypothesis could be supported by evidence from sequence alignments. We
present phylogenetic analyses based on sequence alignments of the
methyltransferase catalytic domains of more than 2300 eukaryotic and
prokaryotic DNA-(cytosine C5)-methyltransferases and analyzed the
distribution of DNA methyltransferases in eukaryotic species. The Dnmt2
homologues were reliably identified by an additional conserved CFT
motif next to motif IX. All DNA methyltransferases and Dnmt2 enzymes
were clearly separated from other RNA-(cytosine-C5)-methyltransferases.
Our sequence alignments and phylogenetic analyses indicate that the
last universal eukaryotic ancestor contained at least one member of the
Dnmt1, Dnmt2 and Dnmt3 families of enzymes and additional RNA
methyltransferases. The similarity of Dnmt2 enzymes with DNA
methyltransferases and absence of similarity with RNA
methyltransferases combined with their strong RNA methylation activity
suggest that the ancestor of Dnmt2 was a DNA methyltransferase and an
early Dnmt2 enzyme changed its substrate preference to tRNA. There is
no phylogenetic evidence that Dnmt2 was the precursor of eukaryotic
Dnmts. Most likely, the eukaryotic Dnmt1 and Dnmt3 families of DNA
methyltransferases had an independent origin in the prokaryotic DNA
methyltransferase sequence space.

<>

<1>Juttermann, R., Li, E., Jaenisch, R.
<2>Toxicity of 5-aza-2'-deoxycytidine to mammalian cells is mediated primarily by covalent trapping of DNA methyltransferase rather than DNA demethylation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>11797-11801
<6>1994
<7>The deoxycytidine analog 5-aza-2'-deoxycytidine (5-azadCyd) has been widely used as a DNA
methylation inhibitor to experimentally induce gene expression and cellular differentiation.
Prior to the availability of mutant mice with altered DNA methyltransferase levels, treatment
of cells with drugs has been the only means to experimentally manipulate the level of genomic
DNA methylation in mammalian cells. Substitution of DNA with 5-azadCyd leads to covalent
trapping of the enzyme, thereby depleting the cells of enzyme activity and resulting in DNA
demethylation. 5-AzadCyd or 5-azacytidine treatment causes multiple changes in treated cells,
including activation of silent genes, decondensation of chromatin, and induction of cellular
differentiation, all of which are believed to be consequences of drug-induced demethylation.
5-AzadCyd is highly toxic in cultured cells and animals and is utilized as a potent antitumor
agent for treatment of certain human cancers. It has been postulated that the toxicity of the
drug in mammalian cells is also due to its inhibition of DNA methylation. The chemistry of the
methylation reaction is consistent, however, with an alternative mechanism: the cyto-toxic
effect of 5-azadCyd may be directly mediated through the covalent binding of DNA
methyltransferase to 5-azadCyd-substituted DNA. We have tested this possibility by using
embryonic stem cells and mice with reduced levels of DNA methyltransferase due to a targeted
mutation of the gene. When exposed to 5-azadCyd mutant embryonic stem cells or embryos were
significantly more resistant to the toxic effects of the drug than wild-type cells and
embryos, respectively. These results strongly suggest that the cellular DNA methyltransferase
itself, rather than the secondary demethylation of genomic DNA, is the primary mediator of
5-azadCyd cytotoxicity. In light of our results, some conclusions from previous studies using
5-azad-Cyd in order to experimentally manipulate cellular methylation levels may have to be
reassessed. Also, our data make clear predictions for cancer treatment: tumor cells with
elevated DNA methyltransferase levels would be expected to be susceptible to treatment with
5-azadCyd, whereas tumors with reduced levels of the enzyme would be resistant.

<>

<1>Jutur, P.P., Hoti, S.L., Reddy, A.R.
<2>Bsu2413I and Bfi2411I, two new thermophilic type II restriction endonucleases from Bacillus subtilis and Bacillus firmus: isolation and  partial purification. Thermophilic endonucleases from two Bacillus  species.
<3>Mol. Biol. Rep.
<4>31
<5>139-142
<6>2004
<7>Two new thermophilic type II restriction endonucleases, which we designated as Bsu2413I and
Bfi2411I, have been isolated from gram-positive thermophilic bacteria Bacillus subtilis strain
2413 and Bacillus firmus strain 2411 respectively and partially purified.  The restriction
endonucleases were extracted from cell extracts and purified using single step purification
through phosphocellulose column chromatography, SDS-PAGE profile showed denatured molecular
weights of 33 and 67 kDa for the Bsu2413I and 39 and 67 kDa for the Bfi2411I.  The partially
purified Bsu2413I enzyme restricted pBR322 DNA into two fragments of 3250 and 1100 bp whereas
Bfi2411I enzyme restricted pBR322 DNA into two fragments of 3500 and 800 bp.  The activity of
both endonucleases was assayed at 55oC and they required Mg+2 as cofactor like other type II
restriction endonucleases.

<>

<1>Jutur, P.P., Reddy, A.R.
<2>Isolation and partial purification of a novel type II restriction endonuclease Bsu121 I, from Bacillus subtilis - Bsu121 I, a type II  restriction endonuclease from Bacillus subtilis.
<3>Mol. Biol. Rep.
<4>29
<5>383-385
<6>2002
<7>A new type II restriction endonuclease which we designated as Bsu121I has been isolated from
the
gram-positive bacterium Bacillus subtilis strain
121 and partially purified. The restriction endonuclease was isolated
from cell extracts using step-wise purification through ammonium
sulfate precipitation, followed by phosphocellulose column
chromatography. SDS-PAGE profile showed denatured molecular weights (23
and 67 kDa) of the endonuclease. The partially purified enzyme
restricted pBR322 DNA into two fragments of 3200 and 1700 bp. The
endonuclease activity required Mg+2 as cofactor like other type II
endonucleases.

<>

<1>Jutur, P.P., Reddy, A.R.
<2>BpaI and BpnI: novel type II restriction endonucleases from Bacillus pasteurii and Bacillus pantothenticus.
<3>Biotechnol. Lett.
<4>26
<5>929-932
<6>2004
<7>Two novel type II restriction endonucleases, designated as BpaI and BpnI, were isolated from
Bacillus pasteurii strain 1761 and Bacillus
pantothenticus strain 1639, respectively. They were partially purified
and SDS-PAGE indicated Mr values of 28 and 67 kDa for BpaI, 28 and 48
kDa for BpnI. The partially purified endonucleases hydrolyzed DNA into
discrete fragments: pUC18 (2.6 kb for BpaI; 1.8 and 0.8 kb for BpnI),
pBR322 (2.5 and 1.8 kb for BpaI; 2.6 and 1.7 kb for BpnI) and Phix174
DNA (3.2 and 2.1 kb for BpaI; 4 and 1.3 kb for BpnI).

<>

<1>Jutur, P.P., Reddy, A.R.
<2>Isolation, purification and properties of new restriction endonucleases from Bacillus badius and Bacillus lentus.
<3>Microbiol. Res.
<4>162
<5>378-383
<6>2007
<7>We tentatively named two enzymes as Bbal and Blel, which were isolated and purified from
Gram-positive mesophilic bacteria Bacillus badius
1458 and Bacillus lentus 1689 respectively, by ammonium sulphate
precipitation, phosphocellulose and heparin-sepharose column
chromatography. SDS-PAGE protein profiles for Bbal and Blel showed
denatured molecular weights of 52 and 48 kDa, respectively. Bbal
hydrolyzed pUC18 DNA into 1900 and 700 bp, pBR322 DNA into two
fragments of 2800 and 1500 bp and Phi x 174 DNA into 3800 and 1600 bp.
Blel hydrolyzed pUC18 DNA into 1800 and 800 bp, pBR322 DNA into two
fragments of 2700 and 1600 bp and Phi x 174 DNA into 3700 and 1700 bp.
The effects of temperature, ionic strength, pH and Mg2+ ion
concentrations were studied to demonstrate some biochemical properties
of Bbal and Blel. Maximum activities of these enzymes were observed at
37 degrees C (pH 8.0) with 100 mM NaCl and 10 mM Mg2+ concentrations.

<>

<1>Kabisch, J., Thurmer, A., Hubel, T., Popper, L., Daniel, R., Schweder, T.
<2>Characterization and optimization of Bacillus subtilis ATCC 6051 as an expression host.
<3>J. Biotechnol.
<4>163
<5>97-104
<6>2013
<7>The genome sequence of Bacillus subtilis ATCC 6051 and its suitability as an
expression host for recombinant protein production was determined. The comparison
of this undomesticated wild type with the widely used laboratory strain B.
subtilis 168 reveals a high degree of congruency between the two strains.
Differences could only be detected on the level of point mutations or small
insertions. B. subtilis ATCC 6051 shows none of the auxotrophies known for B.
subtilis 168 and is able to produce polyketides. It exhibits better use of
complex media and higher genomic stability through reduced natural competence.
Consequently, B. subtilis ATCC 6051 was genetically modified to yield an
optimized strain for the production of heterologously expressed proteins under
control of an acetoin-inducible promoter.

<>

<1>Kachalova, G.S., Artyukh, R.I., Lavrova, N.V., Ryazanova, E.M., Karyagina, A.S., Kubareva, E.A., Bartunik, H.D.
<2>Crystallization and preliminary crystallographic analysis of the (cytosine-5)-DNA methyltransferase NlaX from Neisseria lactamica.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>61
<5>852-854
<6>2005
<7>Crystals of the (cytosine-5)-DNA methyltransferase NlaX from Neisseria lactamica (molecular
weight 36.5 kDa) have been grown at 291 K using 2.5 M NaCl as precipitant.  The crystals
diffract to 3.0 A resolution at 100 K.  The crystals belong to space group P321, with
unit-cell parameters a = 121.98, b = 121.98, c = 56.71 A.  There is one molecule in the
asymmetric unit and the solvent content is estimated to be 62.1% by volume.

<>

<1>Kachalova, G.S., Rogulin, E.A., Artyukh, R.I., Perevyazova, T.A., Zheleznaya, L.A., Matvienko, N.I., Bartunik, H.D.
<2>Crystallization and preliminary crystallographic analysis of the site-specific DNA nickase Nb.BspD6I.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>61
<5>332-334
<6>2005
<7>Crystals of site-specific DNA nickase Nb.BspD6I (of molecular weight 70.8 kDa) have been grown
at 291 K using PEG 8000 as precipitant. The
diffraction pattern of the crystal extends to 3.3 A resolution at 100 K.
The crystal belongs to space group P2(1), with unit-cell parameters a =
57.76, b = 90.67, c = 71.71, beta = 110.1 degrees. There is one molecule
in the asymmetric unit and the solvent content is estimated to be 53% by
volume.

<>

<1>Kachalova, G.S., Rogulin, E.A., Yunusova, A.K., Artyukh, R.I., Perevyazova, T.A., Matvienko, N.I., Zheleznaya, L.A., Bartunik, H.D.
<2>Structural Analysis of the Heterodimeric Type IIS Restriction Endonuclease R.BspD6I Acting as a Complex between a Monomeric Site-specific Nickase and  a Catalytic Subunit.
<3>J. Mol. Biol.
<4>384
<5>489-502
<6>2008
<7>The heterodimeric restriction endonuclease R.BspD6I from Bacillus species D6 recognizes a
pseudosymmetric sequence and cuts both DNA strands outside the recognition sequence. The large
subunit, Nt.BspD6I, acts as a type IIS site-specific monomeric nicking endonuclease. The
isolated small subunit, ss.BspD6I, does not bind DNA and is not catalytically active. We
solved the crystal structures of Nt.BspD6I and ss.BspD6I at high resolution. Nt.BspD6I
consists of three domains, two of which exhibit structural similarity to the recognition and
cleavage domains of FokI. ss.BspD6I has a fold similar to that of the cleavage domain of
Nt.BspD6I, each containing a PD-(D/E)XK motif and a histidine as an additional putative
catalytic residue. In contrast to the DNA-bound FokI structure, in which the cleavage domain
is rotated away from the DNA, the crystal structure of Nt.BspD6I shows the recognition and
cleavage domains in favorable orientations for interactions with DNA. Docking models of
complexes of Nt.BspD6I and R.BspD6I with cognate DNA were constructed on the basis of
structural similarity to individual domains of FokI, R.BpuJI and HindIII. A three-helix bundle
forming an interdomain linker in Nt.BspD6I acts as a rigid spacer adjusting the orientations
of the spatially separated domains to match the distance between the recognition and cleavage
sites accurately.

<>

<1>Kachalova, G.S., Yunusova, A.K., Artyukh, R.I., Rogulin, E.A., Perevyazova, T.A., Zheleznaya, L.A., Matvienko, N.I., Bartunik, H.D.
<2>Crystallization and preliminary x-ray diffraction analysis of the small subunit of the heterodimeric restriction endonuclease R.BspD6I.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>63
<5>795-797
<6>2007
<7>The heterodimeric restriction endonuclease R.BspD6I is composed of a small subunit with a
cleavage site and a large subunit, containing a recognition domain and a cleavage domain, that
may function separately as a monomeric nicking endonuclease.  Here, the crystallization of the
small subunit and diffraction data collection to 1.5 A resolution are reported.

<>

<1>Kaczorowski, T., Sektas, M., Skowron, P., Podhajska, A.J.
<2>The FokI methyltransferase from Flavobacterium okeanokoites - Purification and characterization of the enzyme and its truncated derivatives.
<3>Mol. Biotechnol.
<4>13
<5>1-15
<6>1999
<7>The gene encoding the FokI methyltransferase from Flavobacterium okeanokoites was cloned into
an Escherichia coli vector. The
transcriptional start sites were mapped as well as putative -10 and -35
regions of the fokIM promoter. Enzyme overproduction was ensured by
cloning the fokIM gene under the phi 10 promoter of phage T7. M.FokI
was purified using a two-step chromatography procedure. M.FokI is a
monomeric protein with an Mr = 76,000 +/- 1,500 under denaturing
conditions. It contains 21 Arg residues, at least one of which is
required for activity as shown by inhibition using 2,3-butanedione.
Deletion mutants in the N- and C-terminus of M.FokI were isolated and
characterized. The N-terminal derivative (M.FokIN) methylates the
adenine residue within the sequence 5'-GGATG-3', whereas the C-terminal
derivative (M.FokIC) modifies the adenine residue within the sequence
5'-CATCC-3'. Substrate-protection studies, utilizing chemical
modification combined with data on the effect of divalent cations and
pH on methylation activity, proved the existence of two catalytic
centers within the FokI methyltransferase molecule. M.FokI and its
truncated derivatives require S-adenosyl-L-methionine as the
methyl-group donor, and they are strongly inhibited by divalent cations
(Mg2+, Ca2+, Ba2+, Mn2+ and Zn2+) and S-adenosyl-L-homocysteine. The
Km values for the methyl donor, S-adenosyl-L-methionine are 0.6 microM
(M.FokI), 0.3 microM (M.FokIN), and 0.9 microM (M.FokIC) while the Km
values for substrate lambda DNA are 1.2 nM (M.FokI): 1.4 nM (M.FokIN),
and 1.3 nM (M.FokIC).

<>

<1>Kaczorowski, T., Skowron, P., Podhajska, A.J.
<2>Purification and characterization of the FokI restriction endonuclease.
<3>Gene
<4>80
<5>209-216
<6>1989
<7>The restriction endonuclease FokI from Flavobacterium okeanokoites was purified
to homogeneity.  Based on gel filtration, sedimentation and sodium dodecyl
sulfate-polyacrylamide-gel electrophoresis, the following properties of the
enzyme were determined:  FokI exists in one active monomeric form, and has an
Mr of 64,000-65,400.  FokI is a strongly basic protein with an isoelectric
point of 9.4.  The enzyme exhibits restriction activity in the pH range 5.0 to
10.5 (maximum level at pH 7.0-8.5) and its divalent cation requirement is
satisfied not only by Mg2+, but also by Co2+, Mn2+, Ni2+, Cd2+, Zn2+ and Fe2+.

<>

<1>Kaddurah-Daouk, R., Cho, P., Smith, H.O.
<2>Catalytic properties of the HhaII restriction endonuclease.
<3>J. Biol. Chem.
<4>260
<5>15345-15351
<6>1985
<7>The catalytic properties of the HhaII restriction endonuclease were studied
using plasmid pSK11 DNA containing a single 5'-G-A-N-T-C HhaII cleavage site as
substrate.  Reactions were followed by two methods: 1) gel electrophoretic
analysis of nicked circular and linear DNA products, or 2) release of
32P-labeled inorganic phosphate from specifically labeled HhaII sites in a
reaction coupled with bacterial alkaline phosphatase.  The enzyme is optimally
active at 37C in 10 mM Tris-HCl (pH 9.1) and 4-10 mM MgCl2 without added NaCl.
Activity is stabilized by the presence of 2-mercaptoethanol and 0.2% Triton
X-100 or 50 lg/ml bovine serum albumin.  At enzyme concentrations below 10 nM
and using pSK11 as substrate, initial kinetic rates were dependent on the order
of mixing of reactants.  A lag of 3-4 min was observed if enzyme or substrate
was added last.  Preincubation of substrate and enzyme followed by initiation
of the reaction with MgCl2 or preincubation of the enzyme with nonspecific DNA
followed by initiation with substrate eliminated or reduced the lag,
respectively, and speeded up the reactions.  Under a wide range of reaction
conditions, nicked pSK11 DNA accumulated early, while linear molecules appeared
later, suggesting that HhaII cleaves one strand at a time in separate binding
events.  The apparent Km for covalently closed pSK11 DNA molecules was
approximately 17 nM, and the turnover number for the conversion of covalent to
nicked sites was 1.1 single strand scissions/min.  Pre-steady state kinetic
analysis indicated that cleavage of the first phosphodiester bond in a site is
first order with a rate constant of about 0.8 min-1, while cleavage of the
second phosphodiester bond is first order with a rate constant of about 0.2
min-1.

<>

<1>Kaden, R., Agren, J., Ferrari, S., Lindberg, M., Backman, S., Wahab, T.
<2>Whole-Genome Sequence of Brucella canis Strain SVA13, Isolated from an Infected Dog.
<3>Genome Announcements
<4>2
<5>e00700-14
<6>2014
<7>An outbreak of canine brucellosis in Sweden was confirmed by the National Veterinary Institute
(SVA) in August 2013. The whole genome of the causative
agent was sequenced, assembled, and analyzed.

<>

<1>Kado, C.I., Rodriguez, R.L., Froman, B.E., Tait, R.C.
<2>Restriction Endonuclease XcyI.
<3>US Patent Office
<4>US 4588689
<5>877
<6>1986
<7>A novel restriction endonuclease designated XcyI recognizes and cleaves the sequence
5'-C/CCGGG-3', where the slash indicates the cleavage site. The enzyme may be obtained from
Xanthomonas cyanopsidis. Xanthomonas cyanopsidis strain 13D5 was deposited at the American
Type Culture Collection on Jan. 20, 1984, and granted accession No. 39587.

<>

<1>Kadyrov, F.A., Kaliman, A.V., Kriukov, V.M.
<2>SegE endonuclease from phage T4. I. Cloning, expression and biochemical characteristics of endonuclease activity.
<3>Mol. Biol. (Mosk)
<4>30
<5>1096-1106
<6>1996
<7>
<>

<1>Kadyrov, F.A., Kriukov, V.M., Shliapnikov, M.G., Baev, A.A.
<2>SegE-a new site-specific endodeoxyribonuclease from bacteriophage T4.
<3>Dokl. Akad. Nauk.
<4>339
<5>404-406
<6>1994
<7>
<>

<1>Kadyrov, F.A., Kryukov, V.M., Shlyapnikov, M.G., Baev, A.A.
<2>SegE--a new site-specific endodeoxyribonuclease of bacteriophage T4.
<3>Dokl. Biochem.
<4>339
<5>145-147
<6>1994
<7>Introns of group I were found in nuclear DNA of Tetrahymena, mitochondrial genomes of fungi,
genomes of cyanobacteria, archaebacteria and bacteriophages, particularly in the genome of
bacteriophage T4.  Most of these introns code for specific endonucleases that provide the
transfer of intron DNA into genes lacking introns by introducing double-strand breaks at the
sites of intron insertion.  Earlier, we determined the DNA nucleotide sequence of the region
of phage T4 hoc and uvsW genes containing an open reading frame (ORF205) designated as hoc2
gene.  The amino acid sequence of the product of this gene was shown to share a high degree of
homology with a site-specific endodeoxyribonuclease, SegA.  Subsequently, the gene was
classified with phage T4 genes coding for proteins, similar to endonucleases of group I
introns (seg), and termed segE.  The five members of the group (segA, segB, segC, segD, and
segE) are homologous to each other.

<>

<1>Kadyrov, F.A., Shlyapnikov, M.G., Kryukov, V.M.
<2>A phage T4 site-specific endonuclease, SegE, is responsible for a non-reciprocal genetic exchange between T-even-related phages.
<3>FEBS Lett.
<4>415
<5>75-80
<6>1997
<7>The bacteriophage T4 segE gene encoding site-specific endonuclease lies between the hoc.1 and
uvsW genes.  The similar region of T-even-related phage RB30 lacks the segE gene.  Here we
demonstrate that the phage T4 segE gene is inherited preferably by progeny of mixed infection
with RB30.  The preferred inheritance of the segE gene depends on its own expression and is
based on a non-reciprocal homologous recombination event providing the transfer of the gene
from the segE-containing to the segE-lacking allele.  The SegE endonuclease cleaves DNA in a
site located at the 5' end of the uvsW gene in the RB30 genome.  The T4 DNA is also cleaved
by the enzyme, but less efficiently.  The cleavage at the RB30 site appears to initiate the
observed conversion, which is stimulated by DNA homology and accompanied by co-conversion of
flanking markers.  Our findings provide a novel example of endonuclease-dependent generation
of genetic variation in prokaryotes.

<>

<1>Kaeppler, S.M., Springer, N.M., Muszynski, M.G.
<2>Class II DNA methyltransferases of Zea Mays.
<3>International Patent Office
<4>WO 0053732
<5>
<6>2000
<7>The present invention provides nucleic acids encoding polypeptides which encode a DNA
methyltransferase.  These nucleic acids can be used to stabilize transgene expression in
transgenic plants, to alter the yield or biochemical qualities of plants to silencing targeted
genes in plants in vivo.

<>

<1>Kaeppler, S.M., Springer, N.M., Muszynski, M.G.
<2>Nucleic acid and amino acid sequences encoding a de novo DNA methyltransferase.
<3>International Patent Office
<4>WO 0153470 A
<5>
<6>2001
<7>The present invention provides nucleic acids encoding polypeptides which encode a de novo DNA
methyltransferase.  These nucleic acids can be used to stabilize transgene expression in
transgenic plants, to alter the yield or biochemical qualities of plants to silencing targeted
genes in plants in vivo.

<>

<1>Kaeppler, S.M., Springer, N.M., Muszynski, M.G.
<2>New zmet3 methyltransferase protein and polynucleotide from Zea mays, useful in reducing or altering the level of DNA methylation in a plant,  or in methylating a target gene in a plant in vivo to silence or  knock-out the geneinvolving maize transgenic.
<3>International Patent Office
<4>WO 200153470
<5>
<6>2001
<7>An isolated and purified Zea mays zmet3 methyltransferase polynucleotide (I), is claimed. Also
claimed are: a zmet3
methyltransferase (II); an expression cassette (III) comprising a
promoter sequence operably linked to (I); a bacterial cell comprising
(III); a plant cell transformed with (III); a transformed plant and its
seeds; transformed plant seed containing the plant cell; and a process
for methylating a target gene in a plant by transforming a plant with a
recombinant expression cassette comprising a tissue-specific promoter
and (I). The nucleic acids may be used to reduce or alter the level of
DNA methylation in a plant, to methylate a target gene in a plant in
vivo to silence or knock-out the gene, in a marker-aided selection, and
to provide an optimal means to DNA fingerprint de novo DNA
methyltransferases in other cultivars and wild germplasm. The zmet3
methyltransferase may be used as immunogen of antigen for the
production of antibodies. Antibodies may be used to screen plants for
the expression of zmet3 methyltransferases or in affinity
chromatography in isolating zmet3 methyltransferases. (50pp)

<>

<1>Kafri, T., Hershko, A., Razin, A.
<2>Probing CpG methylation at CACGTG with BbrPI restriction enzyme.
<3>Nucleic Acids Res.
<4>21
<5>2950
<6>1993
<7>The sequence CACGTG is known to be a recognition site for basic helix-loop-helix binding
proteins such as Max and Myc. This site is a common element which is present in the upstream
region of a variety of genes that participate in developmental processes. Being a
CpG-containing sequence, it is prone to methylation in vertebrates. It is therefore of
importance to probe for the status of methylation of this site since CpG modification
frequently affects protein binding. The restriction enzyme BbrPI has been shown to be
sensitive to cytosine methylation at this site. However, since the substrate used for BbrPI
digestion has been prepared by PCR with 5-methyldCTP replacing dCTP, it was impossible to
conclude whether methylation of the outer cytosine, the inner cytosine or both is required to
inhibit digestion by BbrPI. We, therefore, use here lambda DNA methylated in vitro by M.SssI
as substrate for BbrPI digestion. The Spiroplasma methylase, M.SssI, is known to methylate
exclusively CpG sequences. Therefore, only the inner cytosine residue in CACGTG is expected to
be methylated by M.SssI. The results shown in Figure 1 clearly indicate that methylation of
the inner cytosine residue in CACGTG is sufficient to inhibit digestion by BbrPI. It is
therefore possible to probe the status of methylation of this site in mammalian genomic DNA by
BbrPI digestion followed by Southern blotting. Such an analysis with digested DNA from various
tissues of the mouse revealed that a BbrPI site in the mouse homeobox gene HoxA5 is methylated
in a tissue-specific manner.

<>

<1>Kahng, L.S., Shapiro, L.
<2>The CcrM DNA methyltransferase of Agrobacterium tumefaciens is essential, and its activity is cell cycle regulated.
<3>J. Bacteriol.
<4>183
<5>3065-3075
<6>2001
<7>DNA methylation is now recognized as a regulator of multiple bacterial cellular processes.
CcrM is a DNA adenine methyltransferase found in
the alpha subdivision of the proteobacteria. Like the Dam enzyme, which
is found primarily in Escherichia coli and other gamma proteobacteria,
it does not appear to be part of a DNA restriction-modification system.
The CcrM homolog of Agrobacterium tumefaciens was found to be essential
for viability. Overexpression of CcrM is associated with significant
abnormalities of cell morphology and DNA ploidy. Mapping of the
transcriptional start site revealed a conserved binding motif for the
global response regulator CtrA at the -35 position; this motif was
footprinted by purified Caulobacter crescentus CtrA protein in its
phosphorylated state. We have succeeded in isolating synchronized
populations of Agrobacterium cells and analyzing their progression
through the cell cycle. We demonstrate that DNA replication and cell
division can be followed in an orderly manner and that flagellin
expression is cyclic, consistent with our observation that motility
varies during the cell cycle. Using these synchronized populations, we
show that CcrM methylation of the chromosome is restricted to the late
S phase of the cell cycle. Thus, within the alpha subdivision, there is
a conserved cell cycle dependence and regulatory mechanism controlling
ccrM expression.

<>

<1>Kahramanoglou, C., Prieto, A.I., Khedkar, S., Haase, B., Gupta, A., Benes, V., Fraser, G.M., Luscombe, N.M., Seshasayee, A.S.N.
<2>Genomics of DNA cytosine methylation in Escherichia coli reveals its role in stationary phase transcription.
<3>Nat. Commun.
<4>3
<5>886
<6>2012
<7>DNA cytosine methylation regulates gene expression in mammals. In bacteria, its role in gene
expression and genome architecture is less
understood. Here we perform high-throughput sequencing of
bisulfite-treated genomic DNA from Escherichia coli K12 to describe,
for the first time, the extent of cytosine methylation of bacterial DNA
at single-base resolution. Whereas most target sites (C(m)CWGG) are
fully methylated in stationary phase cells, many sites with an extended
CC(m)CWGG motif are only partially methylated in exponentially growing
cells. We speculate that these partially methylated sites may be
selected, as these are slightly correlated with the risk of
spontaneous, non-synonymous conversion of methylated cytosines to
thymines. Microarray analysis in a cytosine methylation-deficient
mutant of E. coli shows increased expression of the stress response
sigma factor RpoS and many of its targets in stationary phase. Thus,
DNA cytosine methylation is a regulator of stationary phase gene
expression in E. coli.

<>

<1>Kajala, K., Coil, D.A., Brady, S.M.
<2>Draft Genome Sequence of Rhizobium rhizogenes Strain ATCC 15834.
<3>Genome Announcements
<4>2
<5>e01108-14
<6>2014
<7>Here, we present the draft genome of Rhizobium rhizogenes strain ATCC 15834. The  genome
contains 7,070,307 bp in 43 scaffolds. R. rhizogenes, also known as
Agrobacterium rhizogenes, is a plant pathogen that causes hairy root disease.
This hairy root induction has been used in biotechnology for the generation of
transgenic root cultures.

<>

<1>Kaji, A.
<2>Novel restriction endonuclease of plasmid Rts1 of Escherichia coli.
<3>Japanese Patent Office
<4>JP 5244946, JP 1993244946 A
<5>
<6>1993
<7>
<>

<1>Kakinuma, S., Ikeda, H., Tanaka, H., Omura, S.
<2>Isolation of restriction-reduced mutants from Streptomyces.
<3>Agric. Biol. Chem.
<4>54
<5>2611-2617
<6>1990
<7>Restriction-reduced mutants were isolated from Streptomyces rosa subsp.
notoensis KA301 and S. tanashiensis strain Kala which produce the
benzoisochromanequinone antibiotics nanaomycin and kalafungin, respectively.
The mutants of S. rosa, which can be transformed with a multi-copy plasmid and
in which the actinophage Pa16 can propagate, were selected.  They were
transformed with a single-copy plasmid propagated in S. lividans TK24, and with
its modified plasmid propagated in the mutant at higher efficiency.  The
mutants of S. tanashiensis were selected by their capability to be transformed
with a multi-copy plasmid.  The efficiency of transformation with a single-copy
plasmid propagated in S. lividans TK24 was low, but was much increased by
heating the protoplasts at 42C for 15 min prior to the transformation.  These
mutants derived from both strains probably lack at least one of their
restriction systems.

<>

<1>Kakizawa, S., Makino, A., Ishii, Y., Tamaki, H., Kamagata, Y.
<2>Draft Genome Sequence of 'Candidatus Phytoplasma asteris' Strain OY-V, an Unculturable Plant-Pathogenic Bacterium.
<3>Genome Announcements
<4>2
<5>e00944-14
<6>2014
<7>Phytoplasmas are unculturable plant-pathogenic bacteria causing devastating damage to
agricultural production worldwide. Here, we report the draft genome
sequence of 'Candidatus Phytoplasma asteris' strain OY-V. Most of the known
virulence factors and host-interacting proteins were conserved in OY-V. This
genome furthers our understanding of genetic diversity and pathogenicity of
phytoplasmas.

<>

<1>Kakutani, T., Jeddeloh, J.A., Richards, E.J.
<2>Characterization of an Arabidopsis thaliana DNA hypomethylation mutant.
<3>Nucleic Acids Res.
<4>23
<5>130-137
<6>1995
<7>We have recently isolated two Arabidopsis thaliana DNA hypomethylation mutations, identifying
the DDM1 locus, that cause a 70% reduction in genomic 5-methylcytosine levels. Here we
describe further phenotypic and biochemical characterization of the ddm1 mutants. ddm1/ddm1
homozygotes exhibited altered leaf shape, increased cauline leaf number, and a delay in the
onset of flowering when compared to non-mutant siblings in a segregating population. Our
biochemical characterization investigated two possible mechanisms for DNA hypomethylation. In
order to see if ddm1 mutations affect DNA methyltransferase function, we compared DNA
methyltransferase activities in extracts from wild-type for both the CpI and CpNpG substrates
suggesting that the DDM1 locus does not encode a DNA methyltransferase. Moreover, the ddm1
mutations did not affect the intracellular level of S-adenosylmethionine, the methyl group
donor for DNA methylation. The possibility that the DDM1 gene product functions as a modifier
of DNA methylation is discussed.

<>

<1>Kalamorz, F. et al.
<2>Draft Genome Sequence of the Thermoalkaliphilic Caldalkalibacillus thermarum Strain TA2.A1.
<3>J. Bacteriol.
<4>193
<5>4290-4291
<6>2011
<7>The genes and molecular machines that allow for a thermoalkaliphilic lifestyle have not been
defined. To address this goal, we report on the
improved high-quality draft genome sequence of Caldalkalibacillus
thermarum strain TA2.A1, an obligately aerobic bacterium that grows
optimally at pH 9.5 and 65 to 70 degrees C on a wide variety of carbon and
energy sources.

<>

<1>Kalburge, S.S., Polson, S.W., Boyd, C.K., Katz, L., Turnsek, M., Tarr, C.L., Martinez-Urtaza, J., Boyd, E.F.
<2>Complete Genome Sequence of Vibrio parahaemolyticus Environmental Strain UCM-V493.
<3>Genome Announcements
<4>2
<5>e00159-14
<6>2014
<7>Vibrio parahaemolyticus is the leading bacterial cause of seafood-related gastroenteritis in
the world. Here, we report the complete genome sequence and
annotation of an environmental strain of V. parahaemolyticus, UCM-V493, with the
aim of understanding the differences between the clinical and environmental
isolates of the bacteria. We also make some preliminary sequence comparisons with
the clinical strain RIMD2210633.

<>

<1>Kaleta, P., O'Callaghan, J., Fitzgerald, G.F., Beresford, T.P., Ross, R.P.
<2>Crucial Role for Insertion Sequence Elements in Lactobacillus helveticus Evolution as Revealed by Interstrain Genomic Comparison.
<3>Appl. Environ. Microbiol.
<4>76
<5>212-220
<6>2010
<7>Lactobacillus helveticus is a versatile dairy bacterium found to possess heterogeneous
genotypes depending on the ecosystem from which
it was isolated. The recently published genome sequence showed the
remarkable flexibility of its structure, demonstrated by a substantial
level of insertion sequence (IS) element expansion in association with
massive gene decay. To assess this diversity and examine the level of
genome plasticity within the L. helveticus species, an array-based
comparative genome hybridization (aCGH) experiment was designed in
which 10 strains were analyzed. The aCGH experiment revealed 16
clusters of open reading frames (ORFs) flanked by IS elements. Four of
these ORFs are associated with restriction/modification which may have
played a role in accelerated evolution of strains in a commercially
intensive ecosystem undoubtedly challenged through successive phage
attack. Furthermore, analysis of the IS-flanked clusters demonstrated
that the most frequently encountered ISs were also those most abundant
in the genome (IS1201, ISL2, ISLhe1, ISLhe2, ISLhe65, and ISLhe63).
These findings contribute to the overall viewpoint of the versatile
character of IS elements and the role they may play in bacterial genome
plasticity.

<>

<1>Kalhoefer, D., Thole, S., Voget, S., Lehmann, R., Liesegang, H., Wollher, A., Daniel, R., Simon, M., Brinkhoff, T.
<2>Comparative genome analysis and genome-guided physiological analysis of Roseobacter litoralis.
<3>BMC Genomics
<4>12
<5>324
<6>2011
<7>ABSTRACT: BACKGROUND: Roseobacter litoralis OCh149, the type species of
the genus, and Roseobacter denitrificans OCh114 were the first described
organisms of the Roseobacter clade, an ecologically important group of
marine bacteria. Both species were isolated from seaweed and are able to
perform aerobic anoxygenic photosynthesis. RESULTS: The genome of R.
litoralis OCh149 contains one circular chromosome of 4,505,211 bp and
three plasmids of 93,578 bp (pRLO149_94), 83,129 bp (pRLO149_83) and
63,532 bp (pRLO149_63). Of the 4537 genes predicted for R. litoralis, 1122
(24.7%) are not present in the genome of R. denitrificans. Many of the
unique genes of R. litoralis are located in genomic islands and on
plasmids. On pRLO149_83 several potential heavy metal resistance genes are
encoded which are not present in the genome of R. denitrificans. The
comparison of the heavy metal tolerance of the two organisms showed an
increased zinc tolerance of R. litoralis. In contrast to R. denitrificans,
the photosynthesis genes of R. litoralis are plasmid encoded. The activity
of the photosynthetic apparatus was confirmed by respiration rate
measurements, indicating a growth-phase dependent response to light.
Comparative genomics with other members of the Roseobacter clade revealed
several genomic regions that were only conserved in the two Roseobacter
species. One of those regions encodes a variety of genes that might play a
role in host association of the organisms. The catabolism of different
carbon and nitrogen sources was predicted from the genome and combined
with experimental data. In several cases, e.g. the degradation of some
algal osmolytes and sugars, the genome-derived predictions of the
metabolic pathways in R. litoralis differed from the phenotype.
CONCLUSIONS: The genomic differences between the two Roseobacter species
are mainly due to lateral gene transfer and genomic rearrangements.
Plasmid pRLO149_83 contains predominantly recently acquired genetic
material whereas pRLO149_94 was probably translocated from the chromosome.
Plasmid pRLO149_63 and one plasmid of R. denitrifcans (pTB2) seem to have
a common ancestor and are important for cell envelope biosynthesis.
Several new mechanisms of substrate degradation were indicated from the
combination of experimental and genomic data. The photosynthetic activity
of R. litoralis is probably regulated by nutrient availability.

<>

<1>Kaliman, A.V., Khasanova, M.A., Kryukov, V.M., Tanyashin, V.I., Baev, A.A.
<2>Cloning and study of the structural organization of the region of inh(lip)-hoc genes of T4 bacteriophage.
<3>Dokl. Akad. Nauk.
<4>303
<5>1486-1489
<6>1988
<7>The cells infected with T-even bacteriophages have been demonstrated to carry early and late
phage-specific mRNA.  It has been established that the early genes are transcribed from the
l-strand of DNA in anticlockwise direction, whereas the late genes are transcribed clockwise
in the r-strand of DNA in the genetic map of T4 phage.  A major part of late T4 phage genes is
located between genes 2 and 54, but the chromosomal segment between genes 24 and 25 contains,
besides the late genes inh and hoc, early (medium) genes uvsY and uvsW(dar).  The inh(lip)
codes for the inhibitor of proteinase, the product of gene 21.  This inhibitor, either in its
natural form or after modification with the help of genetic engineering techniques, is
particularly interesting for studying the mechanism of protein processing and for its possible
use in biotechnology analogous to the product of pin gene of T4 phage.  Gene hoc is promising
for making highly antigenic proteinous structures, which would be discussed below.

<>

<1>Kaliman, A.V., Khasanova, M.A., Kryukov, V.M., Tanyashin, V.I., Bayev, A.A.
<2>The  nucleotide sequence of the region of bacteriophage T4 inh(lip)-hoc genes.
<3>Nucleic Acids Res.
<4>18
<5>4277
<6>1990
<7>
<>

<1>Kalinin, V.N., Lapaeva, I.A., Lunin, V.G., Skripkin, E.A., Smirnov, V.D., Tikchonenko, T.I.
<2>Isolation of HindIII isoschizomeric restriction endonuclease from Bordetella bronchiseptica.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>16-19
<6>1986
<7>Site-specific restriction endonuclease BbrI has been found in bacteriophage resistant strain
B. bronchioseptica 4994.  The technique was elaborated for purification of BbrI to the stage
free of nuclease and phosphatase contamination.  The yield of purified enzyme is 6000-20,000
units per 10 g of biomass.  BbrI recognises and cleaves the same DNA sequence as HindIII with
the formation of four-nucleotide cohesive ends.  The simplicity of cultivation, security for
human, presence of the single restriction endonuclease and the high level of tis production
make B. bronchioseptica 4994 a promising producer of BbrI restriction endonuclease,
isoschizomeric to HindIII, for use in experimental practice and in industry.

<>

<1>Kalinin, V.N., Lapaeva, I.A., Lunin, V.G., Zolotova, O.M., Tikchonenko, T.I.
<2>Bordetella bronchiseptica strain 4994 produces site-specific endonuclease and is tolerant to a wide range of nutrient media.
<3>Soviet Patent Office
<4>SU 1040794 A
<5>
<6>1985
<7>Bordetella bronchiseptica strain 4994 produces site-specific endonuclease.

<>

<1>Kalinowski, J. et al.
<2>The complete Corynebacterium glutamicum ATCC 13032 genome sequence and its impact on the production of L-aspartate-derived amino acids and vitamins.
<3>J. Biotechnol.
<4>104
<5>5-25
<6>2003
<7>The complete genomic sequence of Corynebacterium glutamicum ATCC 13032, well-known in industry
for the production of amino acids, e.g. of
L-glutamate and L-lysine was determined. The C. glutamicum genome was
found to consist of a single circular chromosome comprising 3282708 base
pairs. Several DNA regions of unusual composition were identified that
were potentially acquired by horizontal gene transfer, e.g. a segment of
DNA from C. diphtheriae and a prophage-containing region. After automated
and manual annotation, 3002 protein-coding genes have been identified, and
to 2489 of these, functions were assigned by homologies to known proteins.
These analyses confirm the taxonomic position of C. glutamicum as related
to Mycobacteria and show a broad metabolic diversity as expected for a
bacterium living in the soil. As an example for biotechnological
application the complete genome sequence was used to reconstruct the
metabolic flow of carbon into a number of industrially important products
derived from the amino acid L-aspartate.

<>

<1>Kallimanis, A. et al.
<2>Complete genome sequence of Arthrobacter phenanthrenivorans type strain (Sphe3).
<3>Standards in Genomic Sciences
<4>4
<5>123-130
<6>2011
<7>Arthrobacter phenanthrenivorans is the type species of the genus, and is able to  metabolize
phenanthrene as a sole source of carbon and energy. A.
phenanthrenivorans is an aerobic, non-motile, and Gram-positive bacterium,
exhibiting a rod-coccus growth cycle which was originally isolated from a
creosote polluted site in Epirus, Greece. Here we describe the features of this
organism, together with the complete genome sequence, and annotation.

<>

<1>Kallimanis, A., Karabika, E., Mavromatis, K., Lapidus, A., Labutti, K.M., Liolios, K., Ivanova, N., Goodwin, L., Woyke, T., Velentzas, A.D., Perisynakis, A., Ouzounis, C.C., Kyrpides, N.C., Koukkou, A.I., Drainas, C.
<2>Complete genome sequence of Mycobacterium sp. strain (Spyr1) and reclassification to Mycobacterium gilvum Spyr1.
<3>Standards in Genomic Sciences
<4>5
<5>144-153
<6>2011
<7>Mycobacterium sp.Spyr1 is a newly isolated strain that occurs in a creosote contaminated site
in Greece. It was isolated by an enrichment method using pyrene
as sole carbon and energy source and is capable of degrading a wide range of PAH
substrates including pyrene, fluoranthene, fluorene, anthracene and acenapthene.
Here we describe the genomic features of this organism, together with the
complete sequence and annotation. The genome consists of a 5,547,747 bp
chromosome and two plasmids, a larger and a smaller one with sizes of 211,864 and
23,681 bp, respectively. In total, 5,588 genes were predicted and annotated.

<>

<1>Kalman, S., Mitchell, W., Marathe, R., Lammel, C., Fan, J., Hyman, R.W., Olinger, L., Grimwood, J., Davis, R.W., Stephens, R.S.
<2>Comparative genomes of Chlamydia pneumoniae and C. trachomatis.
<3>Nat. Genet.
<4>21
<5>385-389
<6>1999
<7>Chlamydia are obligate intracellular eubacteria that are phylogenetically separated from other
bacterial divisions. C. trachomatis and C. pneumoniae are both pathogens of humans but differ
in their tissue tropism and spectrum of diseases. C. pneumoniae is a newly recognized species
of Chlamydia that is a natural pathogen of humans, and causes pneumonia and bronchitis. In the
United States, approximately 10% of pneumonia cases and 5% of bronchitis cases are attributed
to C. pneumoniae infection. Chronic disease may result following respiratory-acquired
infection, such as reactive airway disease, adult-onset asthma and potentially lung cancer. In
addition, C. pneumoniae infection has been associated with atherosclerosis. C. trachomatis
infection causes trachoma, an ocular infection that leads to blindness, and sexually
transmitted diseases such as pelvic inflammatory disease, chronic pelvic pain, ectopic
pregnancy and epididymitis.  Although relatively little is known about C. trachomatis biology,
even less is known concerning C. pneumoniae. Comparison of the C. pneumoniae genome with the
C. trachomatis genome will provide an understanding of the common biological processes
required for infection and survival in mammalian cells.  Genomic differences are implicated in
the unique properties that differentiate the two species in disease spectrum. Analysis of the
1,230,230-nt C. pneumoniae genome revealed 214 protein-coding sequences not found in C.
trachomatis, most without homologues to other known sequences. Prominent comparative findings
include  expansion of a novel family of 21 sequence-variant outer-membrane proteins,
conservation of a type-III secretion virulence system, three serine/threonine protein kinases
and a pair of  parologous phospholipase-D-like proteins, additional purine and biotin
biosynthetic capability, a homologue for aromatic amino acid (tryptophan) hydroxylase and the
loss of tryptophan biosynthesis genes.

<>

<1>Kalman, T.I.
<2>Glutathione-catalyzed hydrogen isotope exchange at position 5 of uridine.  A model for enzymic carbon alkylation reactions of pyrimidines.
<3>Biochemistry
<4>10
<5>2567-2573
<6>1971
<7>The effect of glutathione on the exchange of hydrogen to deuterium at
position 5 of uridine
was studied by using proton magnetic resonance spectroscopy.  It was found that in D2O
solutions, at
80o, the rate of H-isotope exchange was enhanced in the presence of GSH and that the
enhancement of
the pseudo-first-order rate of exchange was proportional to the GSH concentration.  The
results obtained
with GSH derivatives indicated the requirement of a free SH group for catalysis.  The
GSH-catalyzed H-
isotope exchange showed a bell-shaped dependence on the OD- ion concentration,
suggesting that in
the rate-determining step the ionized SH group of GSH reacts with the nonionized species
of Urd.
Ionization of Urd causes a substantial shielding of the proton at position 6, indicating the
increased
electron density of the 5,6-double bond, which may account for the lack of reactivity
observed at high pD
values.  The results are consistent with a catalytic mechanism of H-isotope exchange
involving the
reversible addition elimination of the SH group of GSH across the 5,6-double bond of
Urd.  The
relevance of these findings to the mechanism of enzyme-catalyzed C-alkylation reactions of
pyrimidine
nucleotides is discussed.

<>

<1>Kalos, M.D., Mannion, J., Elliott, M., Algate, P.A., Fling, S.P., Henderson, R.A., Benson, D.R., Indiriasu, C.J., Secrist, H., Mohamath, R., Lodes, M.J., Reed, S.G.
<2>ompositions and methods for the therapy and diagnosis of lung.
<3>Japanese Patent Office
<4>JP 2004526401 A
<5>
<6>2004
<7>
<>

<1>Kaloss, W.D., Connaughton, J.F., Vanek, P.G., Chirikijan, J.G.
<2>Characterization of overexpressed BamHII methylase.
<3>FASEB J.
<4>6
<5>A217
<6>1992
<7>Both M.BamHI and M.BamHII recognize the palindromic sequence, 5'-GGATCC-3', and
catalyze the transfer of a methyl group from S-adenosylmethionine to the
internal cytosine within the recognition sequence.  Both enzymes have been
cloned, sequenced and expressed in E. coli.  These two methylases utilize an
unusual UUG initiation codon and are expressed at limited levels.  We wish to
report increased expression by changing the initiation codon to AUG using the
site-directed mutagenesis.  Mutants were subcloned into various expression
vectors (pSP64, pKK223-3, pPL-Lambda and Pet-11) and sequenced via double
stranded dideoxy sequencing.  These constructs were transformed into the
appropriate host and protein production was assayed by SDS-PAGE.  Constructs
yielding the highest amount of intact protein were selected for subsequent
purification procedures.  M.BamHI (in pSP64) was purified to apparent
homogeneity from SURE cells by sequential chromatography using
Phosphocellulose, Phenyl Sepharose and G-150 as previously cited, (Vanek, P.G.,
Ph.D. Thesis, Georgetown University, 1991).  M.BamHII (in Pet-11) was purified
to apparent homogeneity from BL21DE3(pLysS) by sequential chromatography using
Phosphocellulose, Heparin Sepharose, Blue Sepharose and G-50.  The BamHII
methylase has an approximate Mw of 32 Kd as predicted from sequence analysis
and confirmed by SDS-PAGE analysis of the purified protein.  This low Mw
confers a unique advantage for further biochemical and structural studies.  To
date, the BamHII methylase has been partially characterized and further
biochemical and biophysical studies, including crystallization, are in
progress.

<>

<1>Kaloss, W.D., Connaughton, J.F., Vanek, P.G., Chirikjian, J.C.
<2>Characterization of overexpressed BamHII methylase.
<3>Biophys. J.
<4>61
<5>A217
<6>1992
<7>Both M.BamHI and M.BamHII recognize the palindromic sequence, 5'-GGATCC-3', and
catalyze the transfer of a methyl group from S-adenosylmethionine to the
internal cytosine within the recognition sequence.  Both enzymes have been
cloned, sequenced and expressed in E. coli.  These two methylases utilize an
unusual UUG initiation codon and are expressed at limited levels.  We wish to
report increased expression by changing the initiation codon to AUG using the
site-directed mutagenesis.  Mutants were subcloned into various expression
vectors (pSP64, pKK223-3, pPL-Lambda and Pet-11) and sequenced via double
stranded dideoxy sequencing.  These constructs were transformed into the
appropriate host and protein production was assayed by SDS-PAGE.  Constructs
yielding the highest amount of intact protein were selected for subsequent
purification procedures.  M.BamHI (in pSP64) was purified to apparent
homogeneity from SURE cells by sequential chromatography using
Phosphocellulose, Phenyl Sepharose and G-150 as previously cited, (Vanek, P.G.,
Ph.D. Thesis, Georgetown University, 1991).  M.BamHII (in Pet-11) was purified
to apparent homogeneity from BL21DE3(pLysS) by sequential chromatography using
Phosphocellulose, Heparin Sepharose, Blue Sepharose and G-50.  The BamHII
methylase has an approximate Mw of 32 kd as predicted from sequence analysis
and confirmed by SDS-PAGE analysis of the purified protein.  This low Mw
confers a unique advantage for further biochemical and structural studies.  To
date, the BamHII methylase has been partially characterized and further
biochemical and biophysical studies, including crystallization, are in
progress.

<>

<1>Kalugin, A.A., Rina, M., Eldarov, M.A., Markaki, M., Korolev, S.V., Samko, O.T., Khoroshutina, E.B., Sikolov, N.N., Bouriotis, V.
<2>Bsp153AI and BspM39I - New isoschizomers of restriction endonuclease PvuII.
<3>Bioorg. Khim.
<4>22
<5>108-110
<6>1996
<7>New restriction endonucleases, Bsp153AI and BspM39I, were isolated from
Bacillus species strains 153A and M39, respectively.  The enzymes recognize and cleave the
nucleotide sequence (5')CAG|CTG / (3')GTC|GAC and are true isoschizomers of restriction
endonuclease PvuII.

<>

<1>Kaluza, K., Auer, J., Frey, B.
<2>Type II restriction endonuclease SexAI.
<3>European Patent Office
<4>EP 0582976 A
<5>
<6>1992
<7>A new type II restriction enzyme SexAI, recognizing A^CCWGGT, is claimed.

<>

<1>Kaluza, K., Frey, B., Jarsch, M.
<2>Type II restriction endonuclease Ssp4800I.
<3>European Patent Office
<4>EP 0480343 B
<5>
<6>1995
<7>Type II restriction endonuclease (I) recognises the following sequence and cuts it along the
line marked. (I), designated Ssp48001, is isolated from Streptomyces spec. BMTU 4800 (DSM
6181). It has temp. and pH optima 50 deg. C and 8.0, respectively.  USE/ADVANTAGE - (I) is
used for DNA analysis. It recognises and cuts a nucleotide sequence not recognised by any
known enzyme.

<>

<1>Kaluza, K., Frey, B., Schmitz, G., Jarsch, M., Kessler, C.
<2>Type II restriction endonuclease SgrAI.
<3>German Patent Office
<4>DE 3938143 A
<5>
<6>1991
<7>The new Type II restriction endonuclease SgrAI has the following recognition sequence, and
cuts at the position as marked: CPu^CCGGPyG. It is especially obtainable from the
microorganisms with the genus Streptomyces.

<>

<1>Kaluza, K., Frey, B., Schmitz-Agheguian, G., Jarsch, M., Kessler, C.
<2>Type II restriction endonuclease SgrAI.
<3>US Patent Office
<4>US 5134069
<5>
<6>1992
<7>*
The type II restriction endonuclease SgrAI has the following recognition sequence:

   5' C(A/G)^CCGG(C/T)G 3'
   3' G(C/T)GGCC^(A/G)C 5'

cleaves DNA at the cleavage site indicated by the arrows, and is preferably obtainable
from microorganisms of the genus Streptomyces. It can be used to recognize and cleave
the double-stranded DNA sequence C(A/G)CCGG(C/T)G and its complementary sequence.


<>

<1>Kaluza, K., Hoeltke, H.J., Jarsch, M., Schmitz-Agheguian, G., Kessler, C.
<2>Type II-restriction endonuclease McrI.
<3>European Patent Office
<4>EP 0428061 A
<5>
<6>1989
<7>*
The new TypeII-restriction endonuclease McrI contains the following recognition sequence,
cleavage is preferred at the marked cleavage site:

     5'-CGGC/AT CG-3'
     3'-GCCG/TA GC-5'

It is especially obtainable from the Microorganism with the genus Micrococcus.


<>

<1>Kaluza, K., Hoeltke, H.J., Schmitz-Agbeguian, G., Kessler, C.
<2>Type II restriction endonuclease McrI.
<3>US Patent Office
<4>US 5134068
<5>
<6>1992
<7>*
Type II restriction endonuclease McrI is disclosed. This endonuclease has recognition
sequence:    5' CGRY^CG 3'
             3' GC^YRGC 5'

wherein R is G or A and Y is C or T, and the cleavage site indicated by the arrows. The
endonuclease is preferably obtained from organism of genus Micrococcus. It can be used
to recognize and cleave double stranded DNA sequence: 5'CGRYCG 3' and its complementary
sequence.


<>

<1>Kaluza, K., Jarsch, M., Schmitz-Agheguian, G., Kessler, C.
<2>Type II restriction endonuclease RleAI.
<3>US Patent Office
<4>US 5134067
<5>
<6>1992
<7>*
The type II restriction endonuclease RleAI has the following recognition sequence:

   5' CCCACANNNNNNNNNNNN^N 3'
   3' GGGTGTNNNNNNNNN^NNNN 5'

cleaves DNA at the cleavage site indicated by the arrows and is preferably obtainable
from microorganisms of the genus Rhizobium. It can be used to recognize and cleave
the double-stranded DNA sequence CCCACA(N)12/9 and its complementary sequence.


<>

<1>Kaluza, K., Jarsch, M., Schmitz-Agheguian, G., Kessler, C.
<2>Type II restriction endonuclease RleAI.
<3>European Patent Office
<4>EP 0428060 A
<5>
<6>1989
<7>*
The new restriction endonuclease RleAI contains the following recognition sequence
(SEQ ID NO 1), cleavage is prefered at the marked cleavage site:

    5'-CCC ACA NNNNNNNNN NNN N-3'
    3'-GGG TGT NNNNNNNNN NNN N-5'

It is especially obtainable from Microorganisms with the genus Rhizobium.


<>

<1>Kaluza, K., Schmitz, G., Jarsch, M., Kessler, C.
<2>Type II restriction endonuclease MamI.
<3>US Patent Office
<4>US 5153122
<5>
<6>1992
<7>New type II restriction endonuclease MamI has recognition sequence:
5'-GATNN^NNATC-3'
3'-CTANN^NNTAG-5'
and a cleavage site indicated by the arrows. It is preferably obtained from microorganism
of the genus Microbacterium.

<>

<1>Kaluza, K., Schmitz, G., Jarsch, M., Kessler, C.
<2>New restriction endonuclease MamI - derived from Microbacterium ammoniaphilum.
<3>German Patent Office
<4>DE 3840358 A
<5>
<6>1990
<7>The new type-II restriction endonuclease MamI has the following recognition sequence with the
indicated cleavage site given by formula (I).  MamI has a temp. optimum of ca 37 deg. C and a
pH optimum of 8.5-9.0.  MamI is obtained by culturing Microbacterium ammoniaphilum DSM 20156,
disrupting the cells and purifying the cell supernatant by affinity chromatography on
heparin-sepharose followed by mol. sieve chromatography and anion-exchange chromatography.

<>

<1>Kamachi, K., Sota, M., Tamai, Y., Nagata, N., Konda, T., Inoue, T., Top, E.M., Arakawa, Y.
<2>Plasmid pBP136 from Bordetella pertussis represents an ancestral form of IncP-1beta plasmids without accessory mobile elements.
<3>Microbiology
<4>152
<5>3477-3484
<6>2006
<7>The complete 41 268 bp nucleotide sequence of the IncP-1beta plasmid pBP136 from the human
pathogen Bordetella pertussis, the primary aetiological agent of whooping cough, was
determined and analysed. This plasmid carried a total of 46 ORFs: 44 ORFs corresponding to the
genes in the conserved IncP-1beta backbone, and 2 ORFs similar to the XF1596 and XF1597 genes
with unknown function of the plant pathogen Xylella fastidiosa. Interestingly, pBP136 had no
accessory genes carrying genetic traits such as antibiotic or mercury resistance and/or
xenobiotic degradation. Moreover, pBP136 had only two of the kle genes (kleAE) that have been
reported to be important for the stability of IncP-1 plasmid in Pseudomonas aeruginosa.
Phylogenetic analysis of the Kle proteins revealed that the KleA and KleE of pBP136 were
phylogenetically distant from those of the present IncP-1 plasmids. In contrast, IncC1 and
KorC, encoded upstream and downstream of the kle genes respectively, and the
replication-initiation protein, TrfA, were closely related to those of the IncP-1beta 'R751
group'. These results suggest that (i) pBP136 without any apparent accessory genes diverged
early from an ancestor of the present IncP-1beta plasmids, especially those of the R751 group,
and (ii) the kle genes might be incorporated independently into the backbone region of the
IncP-1 plasmids for their stable maintenance in various host cells.

<>

<1>Kameshita, I., Sekiguchi, M., Hamasaki, D., Sugiyama, Y., Hatano, N., Suetake, I., Tajima, S., Sueyoshi, N.
<2>Cyclin-dependent kinase-like 5 binds and phosphorylates DNA methyltransferase 1.
<3>Biochem. Biophys. Res. Commun.
<4>377
<5>1162-1167
<6>2008
<7>DNA methyltransferase 1 (Dnmt1) is an enzyme that recognizes and methylates hemimethylated CpG
after DNA replication to maintain methylation patterns. Although the N-terminal region of
Dnmt1 isknown to interact with various proteins, such as methyl-CpG-binding protein 2 (MeCP2),
the associationsof protein kinases with this region have not been reported. In the present
study, we found that a 110-kDaprotein kinase in mouse brain could bind to the N-terminal
domain of Dnmt1. This 110-kDa kinase was
identified as cyclin-dependent kinase-like 5 (CDKL5) by LC-MS/MS analysis. CDKL5 and Dnmt1
werefound to colocalize in nuclei and appeared to interact with each other. Catalytically
active CDKL5,CDKL5(1-352), phosphorylated the N-terminal region of Dnmt1 in the presence of
DNA. Considering thatdefects in the MeCP2 or CDKL5 genes cause Rett syndrome, we propose that
the interaction betweenDnmt1 and CDKL5 may contribute to the pathogenic processes of Rett
syndrome.

<>

<1>Kaminska, K.H., Kawai, M., Boniecki, M., Kobayashi, I., Bujnicki, J.M.
<2>Type II restriction endonuclease R.Hpy188I belongs to the GIY-YIG nuclease superfamily, but exhibits an unusual active site.
<3>BMC Struct. Biol.
<4>8
<5>48
<6>2008
<7>Background: Catalytic domains of Type II restriction endonucleases (REases) belong to a few
unrelated three-dimensional folds. While the
PD-(D/E)XK fold is most common among these enzymes, crystal structures
have been also determined for single representatives of two other
folds: PLD (R. Bfil) and half-pipe (R. Pabl). Bioinformatics analyses
supported by mutagenesis experiments suggested that some REases belong
to the HNH fold (e. g. R. KpnI), and that a small group represented by
R. Eco29kl belongs to the GIY-YIG fold. However, for a large fraction
of REases with known sequences, the three-dimensional fold and the
architecture of the active site remain unknown, mostly due to extreme
sequence divergence that hampers detection of homology to enzymes with
known folds.
Results: R.Hpy188I is a Type II REase with unknown structure.
PSI-BLAST searches of the non-redundant protein sequence database
reveal only 1 homolog (R. HpyF17I, with nearly identical amino acid
sequence and the same DNA sequence specificity). Standard application
of state-of-the-art protein fold-recognition methods failed to predict
the relationship of R. Hpy188I to proteins with known structure or to
other protein families. In order to increase the amount of evolutionary
information in the multiple sequence alignment, we have expanded our
sequence database searches to include sequences from metagenomics
projects. This search resulted in identification of 23 further members
of R. Hpy188I family, both from metagenomics and the non-redundant
database. Moreover, fold-recognition analysis of the extended R.
Hpy188I family revealed its relationship to the GIY-YIG domain and
allowed for computational modeling of the R. Hpy188I structure.
Analysis of the R. Hpy188I model in the light of sequence conservation
among its homologs revealed an unusual variant of the active site, in
which the typical Tyr residue of the YIG half-motif had been
substituted by a Lys residue. Moreover, some of its homologs have the
otherwise invariant Arg residue in a non-homologous position in
sequence that nonetheless allows for spatial conservation of the
guanidino group potentially involved in phosphate binding.
Conclusion: The present study eliminates a significant "white spot"
on the structural map of REases. It also provides important insight
into sequence-structure-function relationships in the GIY-YIG nuclease
superfamily. Our results reveal that in the case of proteins with no or
few detectable homologs in the standard "non-redundant" database, it is
useful to expand this database by adding the metagenomic sequences,
which may provide evolutionary linkage to detect more remote homologs.

<>

<1>Kamps-Hughes, N., Quimby, A., Zhu, Z., Johnson, E.A.
<2>Massively parallel characterization of restriction endonucleases.
<3>Nucleic Acids Res.
<4>41
<5>e119
<6>2013
<7>Restriction endonucleases are highly specific in recognizing the particular DNA sequence they
act on. However, their activity is affected by sequence context, enzyme concentration and
buffer composition. Changes in these factors may lead to either ineffective cleavage at the
cognate restriction site or relaxed specificity allowing cleavage of degenerate 'star'
sites. Additionally, uncharacterized restriction endonucleases and engineered variants present
novel activities. Traditionally, restriction endonuclease activity is assayed on simple
substrates such as plasmids and synthesized oligonucleotides. We present and use
high-throughput Illumina sequencing-based strategies to assay the sequence specificity and
flanking sequence preference of restriction endonucleases. The techniques use fragmented DNA
from sequenced genomes to quantify restriction endonuclease cleavage on a complex genomic DNA
substrate in a single reaction. By mapping millions of restriction site-flanking reads back to
the Escherichia coli and Drosophila melanogaster genomes we were able to quantitatively
characterize the cognate and star site activity of EcoRI and MfeI and demonstrate genome-wide
decreases in star activity with engineered high-fidelity variants EcoRI-HF and MfeI-HF, as
well as quantify the influence on MfeI cleavage conferred by flanking nucleotides. The methods
presented are readily applicable to all type II restriction endonucleases that cleave both
strands of double-stranded DNA.

<>

<1>Kan, N.C., Lautenberger, J.A., Edgell, M.H., Hutchison, C.A. III
<2>The nucleotide sequence recognized by the Escherichia coli K12 restriction and modification enzymes.
<3>J. Mol. Biol.
<4>130
<5>191-209
<6>1979
<7>The sites recognized by the Escherichia coli K1 restriction endonuclease were
localized to defined regions on the genomes of phage PhiXsK1, PhiXsK2, and G4
by the marker rescue technique.  Methyl groups placed on the genome of plasmid
pBR322 by the E.coli K12 modification methylase were mapped in HinfI fragments
1 and 3, and HaeIII fragments 1 and 3.  A homology of seven nucleotides in the
configuration: 5'-A-A-C . . 6N . . G-T-G-C-3', where 6N represents six
unspecified nucleotides, was found among the DNA sequences containing the five
EcoK sites of PhiXsK1, PhiXsK2, G4, and pBR322.  Three lines of evidence
indicate that this sequence constitutes the recognition site of the E.coli K12
restriction enzyme.  This sequence does not occur on PhiXam3cs70, simian virus
40 (SV40), and fd DNAs which do not possess EcoK sites, and occurs only once on
PhiXsK1, PhiXsK2, and G4 DNAs, and twice on pBR322 DNA.  In order to prove that
all seven conserved nucleotides are essential for the recognition by the E.coli
K12 restriction enzyme, the nucleotide sequences of PhiX174, G4, SV40, fd, and
pBR322 were searched for sequences differing from the sequence 5'-A-A-C . . 6N
. . G-T-G-C-3' at only one of the specified positions.  It was found that
sequences differing at each of the specified positions occur on DNA sequences
that do not contain the EcoK sites.  Thus, the recognition site of the E.coli
K12 restriction enzyme has the same basic structure as that of the EcoB site
(Lautenberger et al., 1978).  In each case there are two domains, one
containing three and the other four specific nucleotides, separated by a
sequence of unspecified bases.  However, the unspecified sequence in the EcoK
site must be precisely six bases instead of the eight found in the EcoB site.
Alignment of the EcoK and EcoB sites suggests that four of the seven specified
nucleotides are conserved between the sequences recognized by these two allelic
restriction and modification systems.

<>

<1>Kan, N.C., Lautenberger, J.A., Edgell, M.H., Hutchison, C.A. III
<2>Recognition site of the Escherichia coli K restriction enzyme.
<3>Fed. Proc.
<4>37
<5>1499
<6>1978
<7>Two mutations of PhiX174 which confer sensitivity to restriction by E. coli K12
have been localized on the DNA sequence using the marker rescue technique.  The
sKI site lies between positions 817 and 953 while  sK2 lies between positions
1780 and 1900 (as numbered by Sanger et al., 1977).  The sKI mutation results
in the loss of the HhaI site between fragments 11 and 14, while the sK2
mutation leads to loss of the HhaI site between fragments 9a and 8a.  Sequence
analysis of these two mutant DNAs compared with the PhiX174am3cs70 sequence
verifies this and shows a C5T change at position 874 on the + strand of sK1
DNA, and a G5A change at position 1867 on the + strand of sK2 DNA.  The
following alignment gives the most homology between these sites: 5'
TAAAAAACGTTCTGGTGCTCGC 3' (+ strand of sK1) 3' CGGAAAACGAACAAGTGCAAGA 3' (-
strand of sK2) Since GTGC occurs 24 times in the sequence of PhiX174am3cs70 RF
DNA, some other nucleotides must be involved in the recognition site, which is
most likely to be an interrupted sequence.  The EcoK site on G4 RF DNA is being
mapped.  Comparison between the G4 DNA sequence in this region and the two
sites on PhiX174 should further define the sequence recognized by the EcoK
restriction enzyme.

<>

<1>Kan, T.N., Lin, L., Chandrasegaran, S.
<2>Cloning, sequencing, overproduction, and purification of M.CviBI (GANTC) methyltransferase from Chlorella virus NC-1A.
<3>Gene
<4>121
<5>1-7
<6>1992
<7>We have cloned and sequenced the cvibIM gene from Chlorella virus NC-1A by selecting for the
modification phenotype. The modification gene was cloned on a 7-kb BamHI fragment inserted
into the BamHI site of the pUC13 plasmid. The cvibIM gene was localized at the 3' end of this
fragment. Sequencing of this region revealed a large open reading frame that codes for
methyltransferase (MTase: symbol M) (predicting 260 amino acids). M.CviBI (GANTC) aa sequence
is homologous to M.dam (GATC), M.DpnII (GATC) and M.T4 (GATC), and not so to M.HinfI (GANTC),
M.HhaII (GANTC), and M.DpnA (GATC). We also describe the use of the polymerase chain reaction
technique to alter transcriptonal and translational signals surrounding this gene so as to
achieve overexpression in Escherichia coli. This construct yields M.CviBI at 2-3% of the total
cellular protein. The MTase was purified by phosphocellulose, DEAE, and gel filtration
chromatography. Its size by SDS-PAGE is approximately 28 kDa, in good agreement with that
predicted from the nucleotide sequence.

<>

<1>Kanaras, A.G., Wang, Z.X., Hussain, I., Brust, M., Cosstick, R., Bates, A.D.
<2>Site-specific ligation of DNA-modified gold nanoparticles activated by the restriction enzyme StyI.
<3>Small
<4>3
<5>67-70
<6>2007
<7>
<>

<1>Kandavelou, K., Mani, M., Durai, S., Chandrasegaran, S.
<2>Engineering and applications of chimeric nucleases.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>413-434
<6>2004
<7>Each human cell contains about 3x10^9 base pairs within its genome.  With the first sequence
of the human genome now available, biologists estimate that there are about 30,000-40,000
different genes within the genome.  This is fewer than originally anticipated, but still a
huge number.  These genes code for all of the human body's proteins.  Simple mutations within
the coding region of critical genes can lead to the formation of abnormal proteins, resulting
in disease phenotypes, premature death, or failure of an embryo to develop.  Furthermore,
mutations that affect the regulatory region of genes can result in aberrant gene expression
within cells, and give rise to cancer phenotypes.  The Holy Grail of the Human Genome Project
is Gene Therapy, that is, how genes might someday be used, modified, or even changed to
correct human disease.

<>

<1>Kanduri, C.
<2>Restriction enzyme BstZ17I is sensitive to cytosine methylation.
<3>FEMS Microbiol. Lett.
<4>200
<5>191-193
<6>2001
<7>The cleavage patterns of a subset of restriction enzymes are blocked or impaired when a
methylated CpG is overlapped with either the 5' or 3' end of the canonical restriction site.
BstZ17I restriction endonuclease is a blunt-end cutter, which recognises the hexanucleotide
sequence GTA^TAC. In this report, I show that the BstZ17I restriction enzyme is sensitive to
cytosine methylation. Using both in vitro-methylated episomal plasmids and lambda DNA, I
demonstrate that the BstZ17I restriction enzyme is sensitive to cytosine methylation that
occurs 3' and/or 5' of the canonical recognition sequence.

<>

<1>Kane, P.M., Yamashiro, C.T., Wolczyk, D.F., Neff, N., Goebl, M., Stevens, T.H.
<2>Protein splicing converts the yeast TFP1 gene product to the 69-kD subunit of the vacuolar H+-adenosine triphosphatase.
<3>Science
<4>250
<5>651-657
<6>1990
<7>The TFP1 gene of the yeast Saccharomyces cerevisiae encodes two proteins: the 69-kilodalton
(kD) catalytic subunit of the vacuolar proton-translocating adenosine triphosphatase
(H+-ATPase) and a 50-kD protein. The 60-kD subunit is encoded by the 5' and 3' thirds of the
TFP1 coding region, whereas the 50-kD protein is encoded by the central third. Evidence is
presented that both the 60-kD and 50-kD proteins are obtained from a single translation
product that is cleaved to release the 50-kD protein and spliced to form the 69-kD subunit.

<>

<1>Kane, S.R., Chakicherla, A.Y., Chain, P.S., Schmidt, R., Shin, M.W., Legler, T.C., Scow, K.M., Larimer, F.W., Lucas, S.M., Richardson, P.M., Hristova, K.R.
<2>Whole-Genome Analysis of the Methyl tert-Butyl Ether-Degrading Beta-Proteobacterium Methylibium petroleiphilum PM1.
<3>J. Bacteriol.
<4>189
<5>1931-1945
<6>2007
<7>Methylibium petroleiphilum PM1 is a methylotroph distinguished by its ability to completely
metabolize the fuel oxygenate methyl tert-butyl
ether (MTBE). Strain PM1 also degrades aromatic (benzene, toluene, and
xylene) and straight-chain (C(5) to C(12)) hydrocarbons present in
petroleum products. Whole-genome analysis of PM1 revealed an approximately
4-Mb circular chromosome and an approximately 600-kb megaplasmid,
containing 3,831 and 646 genes, respectively. Aromatic hydrocarbon and
alkane degradation, metal resistance, and methylotrophy are encoded on the
chromosome. The megaplasmid contains an unusual t-RNA island, numerous
insertion sequences, and large repeated elements, including a 40-kb region
also present on the chromosome and a 29-kb tandem repeat encoding
phosphonate transport and cobalamin biosynthesis. The megaplasmid also
codes for alkane degradation and was shown to play an essential role in
MTBE degradation through plasmid-curing experiments. Discrepancies between
the insertion sequence element distribution patterns, the distributions of
best BLASTP hits among major phylogenetic groups, and the G+C contents of
the chromosome (69.2%) and plasmid (66%), together with comparative genome
hybridization experiments, suggest that the plasmid was recently acquired
and apparently carries the genetic information responsible for PM1's
ability to degrade MTBE. Comparative genomic hybridization analysis with
two PM1-like MTBE-degrading environmental isolates ( approximately 99%
identical 16S rRNA gene sequences) showed that the plasmid was highly
conserved (ca. 99% identical), whereas the chromosomes were too diverse to
conduct resequencing analysis. PM1's genome sequence provides a foundation
for investigating MTBE biodegradation and exploring the genetic regulation
of multiple biodegradation pathways in M. petroleiphilum and other
MTBE-degrading beta-proteobacteria.

<>

<1>Kaneda, M., Okano, M., Hata, K., Sado, T., Tsujimoto, N., Li, E., Sasaki, H.
<2>Essential role for de novo DNA methyltransferase Dnmt3a in paternal and maternal imprinting.
<3>Nature
<4>429
<5>900-903
<6>2004
<7>Imprinted genes are epigenetically marked during gametogenesis so that they are exclusively
expressed from either the paternal or the maternal
allele in offspring. Imprinting prevents parthenogenesis in mammals and is
often disrupted in congenital malformation syndromes, tumours and cloned
animals. Although de novo DNA methyltransferases of the Dnmt3 family are
implicated in maternal imprinting, the lethality of Dnmt3a and Dnmt3b
knockout mice has precluded further studies. We here report the disruption
of Dnmt3a and Dnmt3b in germ cells, with their preservation in somatic
cells, by conditional knockout technology. Offspring from Dnmt3a
conditional mutant females die in utero and lack methylation and
allele-specific expression at all maternally imprinted loci examined.
Dnmt3a conditional mutant males show impaired spermatogenesis and lack
methylation at two of three paternally imprinted loci examined in
spermatogonia. By contrast, Dnmt3b conditional mutants and their offspring
show no apparent phenotype. The phenotype of Dnmt3a conditional mutants is
indistinguishable from that of Dnmt3L knockout mice, except for the
discrepancy in methylation at one locus. These results indicate that both
Dnmt3a and Dnmt3L are required for methylation of most imprinted loci in
germ cells, but also suggest the involvement of other factors.

<>

<1>Kaneko, K.J., Burke, J.M., Kaplan, L.J.
<2>The action of endonucleases and methylases:  Electrophoretic analysis of DNA restriction fragments.
<3>J. Chem. Educ.
<4>64
<5>274-278
<6>1987
<7>The restriction endonucleases have played an essential role in the development
of the field of recombinant DNA technology.  These enzymes catalyze the
hydrolysis of double-stranded DNA with a high level of specificity to generate
a well-defined series of polynucleotides called restriction fragments.  The
sequence specificity has been employed to generate fragments for the
construction of recombinant DNA molecules and for DNA sequence analysis.  This
paper presents the procedures used in endonuclease digestion and the
electrophoretic analysis of the restriction fragments.  In fact, two related
experiments are outlined.  One involves the use of a restriction endonuclease
to generate a set of well-defined fragments which then may be used as molecular
weight standards for gel electrophoresis.  With this as an aid, the cleavage
pattern of the DNA with other endonucleases can be analyzed.  The second
experiment employs endonucleases to monitor the protection afforded the DNA
upon methylation and illustrates the differences obtained upon in vitro and in
vivo methylation.

<>

<1>Kaneko, T. et al.
<2>Sequence analysis of the genome of the unicellular cyanobacterium Synechocystis sp. strain PCC6803.  II.  Sequence determination of the entire genome and assignment of poential protein-coding regions (supplement).
<3>DNA Res.
<4>3
<5>185-209
<6>1996
<7>none

<>

<1>Kaneko, T. et al.
<2>Sequence analysis of the genome of the unicellular cyanobacterium Synechocystis sp. Strain PCC6803.  II.  Sequence determination of the entire genome and assignment of potential protein-coding regions.
<3>DNA Res.
<4>3
<5>109-136
<6>1996
<7>The sequence determination of the entire genome of the Synechocystis sp. strain PCC6803 was
completed.  The total length of the genome finally confirmed was 3,573,470 bp, including the
previously reported sequence of 1,003,450 bp from map position 64% to 92% of the genome.  The
entire sequence was assembled from the sequences of the physical map-based contigs of cosmid
clones and of lambda clones and long PCR products which were used for gap-filling.  The
accuracy of the sequence was guaranteed by analysis of both strands of DNA through the entire
genome.  The authenticity of the assembled sequence wqs supported by restriction analysis of
long PCR products, which were directly amplified from the genomic DNA using the assembled
sequence data.  To predict the potential protein-coding regions, analysis of open reading
frames (ORFs), analysis by the GeneMark program and similarity search to databases were
performed.  As a result, a total of 3,168 potential protein genes were assigned on the genome,
in which 145 (4.6%) were identical to reported genes and 1,257 (39.6%) and 340 (10.8%) showed
similarity to reported and hypothetical genes, respectively.  The remaining 1,426 (45.0%) had
no apparent similarity to any genes in databases.  Among the potential protein genes assigned,
128 were related to the genes participating in photosynthetic reactions.  The sum of the
sequences coding for potential protein genes occupies 87% of the genome length.  By adding
rRNA and tRNA genes, therefore, the genome has a very compact arrangement of protein- and
RNA-coding regions.  A notable feature on the gene organization of the genome was that 99
ORFs, which showed similarity to transposase genes and could be classified into 6 groups, were
found spread all over the genome, and at least 26 of them appeared to remain intact.  The
result implies that rearrangement of the genome occurred frequently during and after
establishment of this species.

<>

<1>Kaneko, T. et al.
<2>Complete genome structure of the nitrogen-fixing symbiotic bacterium Mesorhizobium loti.
<3>DNA Res.
<4>7
<5>331-338
<6>2000
<7>The complete nucleotide sequence of the genome of a symbiotic bacterium Mesorhizobium loti
strain MAFF303099 was determined. The genome of M. loti consisted of a single chromosome
(7,036,071 bp) and two plasmids, designated as pMLa (351,911 bp) and pMLb (208, 315 bp). The
chromosome comprises 6752 potential protein-coding genes, two sets of rRNA genes and 50 tRNA
genes representing 47 tRNA species. Fifty-four percent of the potential protein genes showed
sequence similarity to genes of known function, 21% to hypothetical genes, and the remaining
25% had no apparent similarity to reported genes. A 611-kb DNA segment, a highly probable
candidate of a symbiotic island, was identified, and 30 genes for nitrogen fixation and 24
genes for nodulation were assigned in this region.  Codon usage analysis suggested that the
symbiotic island as well as the plasmids originated and were transmitted from other genetic
systems. The genomes of two plasmids, pMLa and pMLb, contained 320 and 209 potential
protein-coding genes, respectively, for a variety of biological functions. These include genes
for the ABC-transporter system, phosphate assimilation, two-component system, DNA replication
and conjugation, but only one gene for nodulation was identified.

<>

<1>Kaneko, T. et al.
<2>Complete genome structure of the Nitrogen-fixing symbiotic bacterium Mesorhizobium loti (Supplement).
<3>DNA Res.
<4>7
<5>381-406
<6>2000
<7>none

<>

<1>Kaneko, T. et al.
<2>Complete genomic sequence of the filamentous nitrogen-fixing cyanobacterium Anabaena sp. strain PCC 7120.
<3>DNA Res.
<4>8
<5>205-213
<6>2001
<7>The nucleotide sequence of the entire genome of a filamentous cyanobacterium, Anabaena sp.
strain PCC 7120, was determined. The genome of Anabaena consisted of a single chromosome
(6,413,771 bp) and six plasmids, designated pCC7120alpha (408,101 bp), pCC7120beta (186,614
bp), pCC7120gamma (101,965 bp), pCC7120delta (55,414 bp), pCC7120epsilon (40,340 bp), and
pCC7120zeta (5,584 bp). The chromosome bears 5368 potential protein-encoding genes, four sets
of rRNA genes, 48 tRNA genes representing 42 tRNA species, and 4 genes for small structural
RNAs. The predicted products of 45% of the potential protein-encoding genes showed sequence
similarity to known and predicted proteins of known function, and 27% to translated products
of hypothetical genes. The remaining 28% lacked significant similarity to genes for known and
predicted proteins in the public DNA databases. More than 60 genes involved in various
processes of heterocyst formation and nitrogen fixation were assigned to the chromosome based
on their similarity to the reported genes. One hundred and ninety-five genes coding for
components of two-component signal transduction systems, nearly 2.5 times as many as those in
Synechocystis sp. PCC 6803, were identified on the chromosome. Only 37% of the Anabaena genes
showed significant sequence similarity to those of Synechocystis, indicating a high degree of
divergence of the gene information between the two cyanobacterial strains.

<>

<1>Kaneko, T. et al.
<2>Complete Genomic Structure of the Bloom-forming Toxic Cyanobacterium Microcystis aeruginosa NIES-843.
<3>DNA Res.
<4>14
<5>247-256
<6>2007
<7>The nucleotide sequence of the complete genome of a cyanobacterium, Microcystis aeruginosa
NIES-843, was determined. The genome of M.
aeruginosa is a single, circular chromosome of 5 842 795 base pairs (bp)
in length, with an average GC content of 42.3%. The chromosome comprises
6312 putative protein-encoding genes, two sets of rRNA genes, 42 tRNA
genes representing 41 tRNA species, and genes for tmRNA, the B subunit of
RNase P, SRP RNA, and 6Sa RNA. Forty-five percent of the putative
protein-encoding sequences showed sequence similarity to genes of known
function, 32% were similar to hypothetical genes, and the remaining 23%
had no apparent similarity to reported genes. A total of 688 kb of the
genome, equivalent to 11.8% of the entire genome, were composed of both
insertion sequences and miniature inverted-repeat transposable elements.
This is indicative of a plasticity of the M. aeruginosa genome, through a
mechanism that involves homologous recombination mediated by repetitive
DNA elements. In addition to known gene clusters related to the synthesis
of microcystin and cyanopeptolin, novel gene clusters that may be involved
in the synthesis and modification of toxic small polypeptides were
identified. Compared with other cyanobacteria, a relatively small number
of genes for two component systems and a large number of genes for
restriction-modification systems were notable characteristics of the M.
aeruginosa genome.

<>

<1>Kaneko, T., Maita, S., Hirakawa, H., Uchiike, N., Minamisawa, K., Watanabe, A., Sato, S.
<2>Complete Genome Sequence of the Soybean Symbiont Bradyrhizobium japonicum Strain USDA6T.
<3>Genes
<4>2
<5>763-787
<6>2011
<7>The complete nucleotide sequence of the genome of the soybean symbiontBradyrhizobium japonicum
strain USDA6T was determined. The genome of USDA6T is asingle circular chromosome of 9,207,384
bp. The genome size is similar to that of thegenome of another soybean symbiont, B. japonicum
USDA110 (9,105,828 bp).Comparison of the whole-genome sequences of USDA6T and USDA110
showedcolinearity of major regions in the two genomes, although a large inversion exists
betweenthem. A significantly high level of sequence conservation was detected in three regions
oneach genome. The gene constitution and nucleotide sequence features in these three
regionsindicate that they may have been derived from a symbiosis island. An ancestral,
largesymbiosis island, approximately 860 kb in total size, appears to have been split into
thesethree regions by unknown large-scale genome rearrangements. The two integration
eventsresponsible for this appear to have taken place independently, but through
comparablemechanisms, in both genomes. 

<>

<1>Kaneko, T., Nakamura, Y., Sasamoto, S., Watanabe, A., Kohara, M., Matsumoto, M., Shimpo, S., Yamada, M., Tabata, S.
<2>Structural analysis of four large plasmids harboring in a unicellular cyanobacterium, Synechocystis sp. PCC 6803.
<3>DNA Res.
<4>10
<5>221-228
<6>2003
<7>The genome of the unicellular cyanobacterium Synechocystis sp. PCC 6803 consists of a single
chromosome and several plasmids of different sizes,
and the nucleotide sequences of the chromosome and three small plasmids
(5.2 kb, 2.4 kb, and 2.3 kb) have already been sequenced. We newly
determined the nucleotide sequences of four large plasmids, which have
been identified in our laboratory (pSYSM:120 kb, pSYSX:106 kb, pSYSA:103
kb, and pSYSG:44 kb). Computer-aided analysis was performed to explore the
genetic information carried by these plasmids. A total of 397 potential
protein-encoding genes were predicted, but little information was obtained
about the functional relationship of plasmids to host cell, as a large
portion of the predicted genes (77%) were of unknown function. The
occurrence of the potential genes on plasmids was divergent, and parA was
the only gene common to all four large plasmids. The distribution data of
a Cyanobacterium-specific sequence (HIP1: 5'-GCGATCGC-3') suggested that
respective plasmids could have originated from different cyanobacterial
strains.

<>

<1>Kaneko, T., Nakamura, Y., Sato, S., Minamisawa, K., Uchiumi, T., Sasamoto, S., Watanabe, A., Idesawa, K., Iriguchi, M., Kawashima, K., Kohara, M., Matsumoto, M., Shimpo, S., Tsuruoka, H., Wada, T., Yamada, M., Tabata, S.
<2>Complete genomic sequence of nitrogen-fixing symbiotic bacterium Bradyrhizobium japonicum USDA110.
<3>DNA Res.
<4>9
<5>189-197
<6>2002
<7>The complete nucleotide sequence of the genome of a symbiotic bacterium
Bradyrhizobium japonicum USDA110 was determined. The genome of B. japonicum was a
single circular chromosome 9,105,828 bp in length with an average GC content of
64.1%. No plasmid was detected. The chromosome comprises 8317 potential
protein-coding genes, one set of rRNA genes and 50 tRNA genes. Fifty-two percent
of the potential protein genes showed sequence similarity to genes of known
function and 30% to hypothetical genes. The remaining 18% had no apparent
similarity to reported genes. Thirty-four percent of the B. japonicum genes
showed significant sequence similarity to those of both Mesorhizobium loti and
Sinorhizobium meliloti, while 23% were unique to this species. A presumptive
symbiosis island 681 kb in length, which includes a 410-kb symbiotic region
previously reported by Gottfert et al., was identified. Six hundred fifty-five
putative protein-coding genes were assigned in this region, and the functions of
301 genes, including those related to symbiotic nitrogen fixation and DNA
transmission, were deduced. A total of 167 genes for transposases/104 copies of
insertion sequences were identified in the genome. It was remarkable that 100 out
of 167 transposase genes are located in the presumptive symbiotic island. DNA
segments of 4 to 97 kb inserted into tRNA genes were found at 14 locations in the
genome, which generates partial duplication of the target tRNA genes. These
observations suggest plasticity of the B. japonicum genome, which is probably due
to complex genome rearrangements such as horizontal transfer and insertion of
various DNA elements, and to homologous recombination.

<>

<1>Kanematsu, T., Ozawa, O.
<2>New restriction enzyme CcoI having good pH and temp. stability - recognises base sequence of double chain DNA and cuts as specified position.
<3>Japanese Patent Office
<4>JP 3244381
<5>
<6>1991
<7>New restriction enzyme recognizes the base sequence (I) of a double chain deoxyribo nucleic
acid (DNA) and cuts at the position shown by the arrows; is suited at pH8.0 at 45 deg.C; is
stabilised at pH5.0-10.0 at 70 deg.C; is stabilised in 50-100mM NaCl; and has mol.wt. 62,000,
when measured by gel filtration. The restriction enzyme is mfd. by culturing a restriction
enzyme CcoI-generative bacterium belonging to Clostridium in a medium and extracting the
restriction enzyme from the culture prod. Pref. the restriction enzyme CcoI-generative
bacterium is Clostridium coccoides B-2. The extract is purified by chromatography.
USE/ADVANTAGE - The restriction enzyme of high in pH stability and temp. stability is mfrd.
easily.

<>

<1>Kanematsu, T., Ozawa, O.
<2>Preparation of novel restriction enzyme BdiI from Bacteriodes.
<3>Japanese Patent Office
<4>JP 2283281
<5>
<6>1990
<7>
<>

<1>Kanematsu, T., Ozawa, O., Murakami, M.
<2>Manufacture of novel restriction enzyme Bbi24I with Bifidobacterium.
<3>Japanese Patent Office
<4>JP 3112484
<5>
<6>1991
<7>
<>

<1>Kanematsu, T., Ozawa, O., Murakami, M., Yoshida, N.
<2>New restriction enzyme - cleaves at specified DNA sites, it is cultured from Bacterioides and has wide range of salt stability.
<3>Japanese Patent Office
<4>JP 3216188 A
<5>
<6>1991
<7>(I) a new restriction enzyme BvuI which has enzymological properties of (a) action and
substrate specificity; it recognises the sequence C^GTACG (I) in double stranded DNA, and
cleaves as indicated. (b) optimum pH 8.0, (c) stable pH 7.0 - 9.5, (d) optimum temp. 37 deg C
(e) stable temp 50 deg C (by warming for 5 min) (f) stable salt concentration (by NaCl) ; 50 -
100 mM, and (g) MW 47,000 (by gel filtration). (2) preparation for restriction enzyme BvuI by
culturing restriction enzyme BvuI producing Bacteroides sp. in enriched medium then collecting
restriction enzyme BvuI from the culture. As Bacteroides sp., Bacteriodes vulgatus S-15 is
used.

<>

<1>Kanesaki, Y., Masutani, H., Sakanaka, M., Shiwa, Y., Fujisawa, T., Nakamura, Y., Yokota, A., Fukiya, S., Suzuki, T., Yoshikawa, H.
<2>Complete Genome Sequence of Bifidobacterium longum 105-A, a Strain with High Transformation Efficiency.
<3>Genome Announcements
<4>2
<5>e01311-14
<6>2014
<7>Bifidobacterium longum 105-A shows high transformation efficiency and allows for  the
generation of gene knockout mutants through homologous recombination. Here,
we report the complete genome sequence of strain 105-A. Genes encoding at least
four putative restriction-modification systems were found in this genome, which
might contribute to its transformation efficiency.

<>

<1>Kang, C., Wu, C.-W.
<2>Studies on SP6 promoter using a new plasmid vector that allows gene insertion at the transcription initiation site.
<3>Nucleic Acids Res.
<4>15
<5>2279-2294
<6>1987
<7>This paper documents the cleavage by HphI at a site 9 bases away from the
recognition sequence, with the production of a single base extension.  This is
in contrast to the usual 8 bases.

<>

<1>Kang, E.S., Park, C.W., Chung, J.H.
<2>Dnmt3b, de novo DNA methyltransferase, interacts with SUMO-1 and Ubc9 through its N-terminal region and is subject to modification by SUMO-1.
<3>Biochem. Biophys. Res. Commun.
<4>289
<5>862-868
<6>2001
<7>Dnmt3b, a DNA methyltransferase, is essential for mammalian development potentially through
its transcription repression activity. To
comprehend the underlying regulatory mechanism of Dnmt3b, we isolated
small ubiquitin-like modifier 1 (SUMO-1) and Ubc9 as Dnmt3b-interacting
proteins using yeast two-hybrid screens. Deletion analysis and
colocalization experiment demonstrated that Dnmt3b interacts with
SUMO-1 and Ubc9 at its N-terminal region. We also confirmed the
modification of Dnmt3b by SUMO-1 in vivo. These results suggest that
sumoylation may constitute a regulation mechanism of Dnmt3b in vivo.

<>

<1>Kang, H.L., Park, Y.H., Lee, E.J., Kim, M.H., Kim, J.S., Park, S.G., Song, J.Y., Park, J.U., Baik, S.C., Ko, G.H., Youn, H.S., Lee, W.K., Cho, M.J.
<2>Distribution of Helicobacter pylori 51-specific genes on other Korean isolates of Helicobacter pylori.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>103
<5>D-069
<6>2003
<7>Background: Many disease-related genes of H. pylori such as cagA, vacA, ure and sod were
identified and well-studied. Entire genome of H.
pylori strain 51, a Korean isolate, has been sequenced. It had 1,454
orfs. We searched disease-specific genes on Korean H. pylori isolates.
Method: The nucleotide sequences of open reading frames of three H.
pylori strains (26685, J99, 51) were compared one another and searched
the strain-specific orf. Five type strains of H. pylori and 80 Korean
isolates were screened with strain 51-specific orfs by overgo
hybridization. Result: Ninteen Korean strain 51-specific orfs were
identified. They included restriction endonuclease S subunits (hsdD,
khp538), ATPase involved in conjugal plasmid transfer (traG, khp924),
ATPase involved in pili and flagella biosynthesis (virB11, khp925),
VirB10 component of type IV secretion system (virB10, khp926),
Adenine-specific DNA methylase (khp1073), site-specific
integrase-resolvase (khp1371), transposase (khp1372), restriction
endonuclease S subunits (hsdS, khp1385), amino acid transporters (lysP,
khp1391), site-specific DNA methylase (dcm, khp1406),
membrane-associated metal-dependent hydrolase (khp1425) and seven
putative ORFs. To investigate distribution of the genes, four type
strains and 80 Korean H. pylori isolates that were isolated from
gastric cancer, gastric ulcer, duodenal ulcer and gastritis were
screened. Four orfs (khp594, khp900, khp1251, khp1391) were revealed to
be abundant in most Korean isolates. Conclusion: We found 19 Korean
strain 51-specific orfs by the comparison of the nucleotide sequence of
orfs of three sequenced H. pylori strains. But none of them seemed to
be disease-specific. But at least four orfs seemed to be specific in
korean isolates. We are expecting that we can find more reasonable
result by screening more Korean isolates.

<>

<1>Kang, I., Kang, D., Oh, H.M., Kim, H., Kim, H.J., Kang, T.W., Kim, S.Y., Cho, J.C.
<2>Genome sequence of strain IMCC2047, a novel marine member of the Gammaproteobacteria.
<3>J. Bacteriol.
<4>193
<5>3688-3689
<6>2011
<7>Strain IMCC2047 was isolated from the Yellow Sea using dilution-to-extinction culturing. The
strain was shown to occupy a
distinct phylogenetic position within the Gammaproteobacteria. Here we
present the genome sequence of strain IMCC2047 that harbors genes for
various metabolic pathways including proteorhodopsin and ribulose
bisphosphate carboxylase.

<>

<1>Kang, I., Lee, K., Yang, S.J., Choi, A., Kang, D., Lee, Y.K., Cho, J.C.
<2>Genome Sequence of 'Candidatus Aquiluna' sp. Strain IMCC13023, a Marine Member of the Actinobacteria Isolated from an Arctic Fjord.
<3>J. Bacteriol.
<4>194
<5>3550-3551
<6>2012
<7>We report the genome sequence of actinobacterial strain IMCC13023, isolated from  arctic fjord
seawater. Phylogenetic analysis of 16S rRNA gene showed that the
strain is related to 'Candidatus Aquiluna rubra.' The genome information suggests
that strain IMCC13023 is a photoheterotroph carrying actinorhodopsin, with the
smallest genome ever reported for a free-living member of the Actinobacteria.

<>

<1>Kang, I., Oh, H.M., Lim, S.I., Ferriera, S., Giovannoni, S.J., Cho, J.C.
<2>Genome sequence of Fulvimarina pelagi HTCC2506T, a Mn(II)-oxidizing alphaproteobacterium possessing an aerobic anoxygenic photosynthetic gene  cluster and Xanthorhodopsin.
<3>J. Bacteriol.
<4>192
<5>4798-4799
<6>2010
<7>Fulvimarina pelagi is a Mn(II)-oxidizing marine heterotrophic bacterium in the order
Rhizobiales. Here we announce the draft genome sequence of F.
pelagi HTCC2506(T), which was isolated from the Sargasso Sea by using
dilution-to-extinction culturing. The genome sequence contained a
xanthorhodopsin gene as well as a photosynthetic gene cluster, which
suggests the coexistence of two different phototrophic mechanisms in a
single microorganism.

<>

<1>Kang, I., Oh, H.M., Vergin, K.L., Giovannoni, S.J., Cho, J.C.
<2>Genome Sequence of the Marine Alphaproteobacterium HTCC2150, Assigned to the Roseobacter Clade.
<3>J. Bacteriol.
<4>192
<5>6315-6316
<6>2010
<7>Here we announce the genome sequence of a marine bacterium, HTCC2150, that was isolated off
the Oregon coast using dilution-to-extinction culturing
and that is affiliated with the Roseobacter clade. The 16S rRNA phylogeny
showed that the strain was closely related to members of the RCA clade.
The genome sequence suggests that strain HTCC2150 is an organoheterotroph
carrying diverse metabolic potential, including a close relationship with
phytoplankton.

<>

<1>Kang, I., Vergin, K.L., Oh, H.M., Choi, A., Giovannoni, S.J., Cho, J.C.
<2>Genome Sequence of strain HTCC2083, a Novel Member of the Marine Roseobacter Clade.
<3>J. Bacteriol.
<4>193
<5>319-320
<6>2010
<7>Strain HTCC2083 was isolated from Oregon seawater using dilution-to-extinction culturing and
represents a novel member of the Roseobacter clade. The draft genome sequence of HTCC2083 is
presented here. The genome is predicted to contain genes for aerobic anoxygenic phototrophy,
sulfite-oxidizing chemolithotrophy, anapleurotic CO2 fixation, carbon monoxide oxidation, and
dimethylsulphoniopropionate (DMSP) utilization.

<>

<1>Kang, J.Joo.S., Meier, J.L., Dervan, P.B.
<2>Design of Sequence-Specific DNA Binding Molecules for DNA Methyltransferase Inhibition.
<3>J. Am. Chem. Soc.
<4>136
<5>3687-3694
<6>2014
<7>The CpG dyad, an important genomic feature in DNA methylation and transcriptional regulation,
is an attractive target for small molecules. To assess the utility of minor groove binding
oligomers for CpG recognition, we screened a small library of hairpin pyrrole-imidazole
polyamides targeting the sequence 5'-CGCG-3' and assessed their sequence specificity using
an unbiased next-generation sequencing assay. Our findings indicate that hairpin polyamide of
sequence PyIm beta Im-gamma-PyIm beta Im (1), previously identified as a high affinity
5'-CGCG-3' binder, favors 5'-GCGC-3' in an unanticipated reverse binding orientation.
Replacement of one beta alanine with Py to afford PyImPyIm-gamma-PyIm beta Im (3) restores the
preference for 5'-CGCG-3' binding in a forward orientation. The minor groove binding hairpin
3 inhibits DNA methyltransferase activity in the major groove at its target site more
effectively than 1, providing a molecular basis for design of sequence-specific antagonists of
CpG methylation.

<>

<1>Kang, S., Lee, H., Han, J.S., Hwang, D.S.
<2>Interaction of SeqA and Dam methylase on the hemimethylated origin of Escherichia coli chromosomal DNA replication.
<3>J. Biol. Chem.
<4>274
<5>11463-11468
<6>1999
<7>Preferential binding of SeqA protein to hemimethylated oriC, the origin of Escherichia coli
chromosomal replication, delays methylation by Dam methylase.  Because the SeqA-oriC
interaction appears to be essential in timing of chromosomal replication initiation, the
biochemical functions of SeqA protein and Dam methylase at the 13-mer L, M., and R region
containing 4 GATC sequences at the left end of oriC were examined.  We found that SeqA protein
preferentially bound hemimethylated 13-mers but not fully nor unmethylated 13-mers.
Regardless of strand methylation, the binding of SeqA protein to the hemimethylated GATC
sequence of 13-mer L was followed by additional binding to other hemimethylated GATC sequences
of 13-mer M and R.  On the other hand, Dam methylase did not discriminate binding of 13-mers
in different methylation patterns and was not specific to GATC sequences.  The binding
specificity and higher affinity of SeqA protein over Dam methylase to the hemimethylated
13-mers along with the reported cellular abundance of this protein explains the dominant
action of SeqA protein over Dam methylase to the newly replicated oriC for the sequestration
of chromosomal replication.  Furthermore, SeqA protein bound to hemimethylated 13-mers was not
dissociated by Dam methylase, and most SeqA protein spontaneously dissociated 10 min after
binding.  Also, SeqA protein delayed the in vitro methylation of hemimethylated 13-mers by Dam
methylase.  These in vitro results suggest that the intrinsic binding instability of SeqA
protein results in release of sequestrated hemimethylated oriC.

<>

<1>Kang, S.C., Choi, W.S., Yoo, O.J.
<2>The effects of DNA methylation by HhaI methylase on the cleavage reactions by HaeII, AhaII and BanI endonuclease.
<3>Biochem. Biophys. Res. Commun.
<4>145
<5>482-487
<6>1987
<7>The DNA methylated by HhaI methylase was resistant against cleavage of HaeII or
AhaII endonuclease indicating that the methyl group of the C5 position of the
innermost cytosine nucleotide interferes with the interaction between the
enzyme and hexameric recognition sequence.  Considering that HaeII or AhaII
methylase has not been isolated yet, the result explained above is a useful
information for protecting a double stranded DNA from being cleaved by HaeII or
AhaII endonuclease. In contrast to HaeII or AhaII endonuclease, BanI
endonuclease which also has HhaI sequence as its tetrameric core was able to
cleave the same DNA normally.  This result suggests that the C5 position of the
inmost pyrimidine nucleotide is not an important contact point between BanI
endonuclease and its hexameric recognition sequence.

<>

<1>Kang, S.C., Yoo, O.J.
<2>Purification and characterization of AccI endonuclease.
<3>Korean J. Microbiol.
<4>23
<5>13-19
<6>1985
<7>AccI endonuclease has been isolated from 300g (wet weight) cells of
Acinetobacter calcoaceticus.  The cells were broken by using French press at
20,000 p.s.i.  After ammonium sulfate fractionation, the enzyme was further
purified by heparin agarose, DEAE-sephadex, Affi-gel Blue, phosphocellulose,
and hydroxylapatite column chromatography.  The purified AccI endonuclease has
a single polypeptide species and its subunit molecular weight was 45,000 +/-
1,000 daltons as judged by 10% SDS-polyacrylamide gel electrophoresis.  The
isolated enzyme was essentially free of contaminating nucleases as judged by
homochromatography by using a 32P-labeled oligonucleotide.  The enzyme showed
maximum activity at pH values between 8.0 and 11.0, and in the presence of
MgCl2.  AccI endonuclease was maximally active in the absence of NaCl and was
completely inhibited at 200 mM NaCl.

<>

<1>Kang, S.C., Yoon, H., Yoo, O.J.
<2>DNA protection by methylation with AluI methylase against cleavages of HindIII, SstI, PvuII and SacI endonucleases.
<3>Korean Biochem. J.
<4>19
<5>41-46
<6>1986
<7>The cleavage reactions of HindIII, SstI, PvuII and SacI endonucleases were inhibited by the
methylation of AluI methylase.  To analyze the cleavage reactions quantitatively, tritium
labeled pBR322 and pPG3282 plasmid DNAs (pBR322 DNA contains HindIII and PvuII sites, while
pPG3282 DNA which is a hog gastrin cDNA clone contains SstI and SacI sites) were used, and the
degree of the inhibition against cleavages by the four kinds of endonucleases was measured.
The results imply that we can predict the specificities of the related hexameric restriction
methylases which have not been isolated yet.  And the predictions are the followings:  (1) The
methylation sites of SstI methylase is not identical with AluI methylase since SstI
endonuclease slowly cut the sequences methylated by AluI methylase.  (2)  The methylation
sites of PvuII and SacI methylases could be identical with AluI methylase since PvuII and SacI
endonucleases could not cut the sequences methylated by AluI methylase.

<>

<1>Kang, W.Y. et al.
<2>Stationary Phase Expression, Purification, and Characterization of XorKI, a Restriction Endonuclease from Xanthomonas oryzae pv. oryzae.
<3>Protein Pept. Lett.
<4>17
<5>381-385
<6>2010
<7>An endonuclease from Xanthomonas oryzae pathovar oryzae KACC 10331, XorKI, was heterologously
produced in Escherichia coli by applying the
stationary state induction method. The yield was 5.4 mg of XorKI per
liter of LB medium. XorKI existed in multiple oligomeric forms as
evidenced by gel filtration chromatography. The specific activity of
purified XorKI was 323000 units per mg.

<>

<1>Kang, X., Ling, N., Sun, G., Zhou, Q., Zhang, L., Sheng, Q.
<2>Complete Genome Sequence of Streptococcus thermophilus Strain MN-ZLW-002.
<3>J. Bacteriol.
<4>194
<5>4428-4429
<6>2012
<7>Streptococcus thermophilus MN-ZLW-002 was originally isolated from traditionally  fermented
Chinese dairy products. One of the strain-dependent characteristics of
this bacterium is its ability to produce exopolysaccharides (EPSs). This study
determined and analyzed the genome sequence of MN-ZLW-002. Its complete genome
comprised 2,046 genes and 1,848,520 nucleotides with an average GC content of
39%. The EPS cluster of MN-ZLW-002 includes 25 open reading frames (ORFs), and
some results indicate a horizontal gene transfer between MN-ZLW-002 and other
lactic acid bacteria (LAB).

<>

<1>Kang, Y.K., Lee, H.B., Noh, M.J., Cho, N.-Y., Yoo, O.J.
<2>Different effects of base analog substitutions in BamHI restriction site on recognition by BamHI endonuclease and BamHI methylase.
<3>Biochem. Biophys. Res. Commun.
<4>206
<5>997-1002
<6>1995
<7>BamHI endonuclease and BamHI methylase were used to investigate their specific interaction
with the common recognition sequence, GGATCC. Five derivatives of the oligonucleotide,
GACGGATCCGTC, containing a variety of single-base analog substitutions within the hexameric
recognition core were synthesized. Steady-state kinetics for the reaction of the endonuclease
and the methylase showed that both enzymes recognize the sequences by contacting with
functional groups exposed in both major and minor grooves of the site but in different ways.
Removal or substitution of the 5-methyl group in thymidine blocked the endonuclease reaction
completely but still allowed the methylase reaction with less efficiency. The data also showed
that the methylase made a critical minor groove contact with the 2-amino group of the first G
but the endonuclease did with that of the second G.

<>

<1>Kang, Y.K., Ryu, J., Yoo, O.J.
<2>Effects of modified bases in sequence recognition by ClaI endonuclease and ClaI methylase.
<3>Biochem. Mol. Biol. Int.
<4>29
<5>859-865
<6>1993
<7>To understand the functional groups required for sequence recognition, dodecanucleotides
containing the ClaI sequence with the middle positions containing modified bases were
synthesized and used as substrates for ClaI endonuclease and ClaI methylase reactions. For the
modification, dU, 5-bromo-dU, and dI were used. Km values of these two enzymes were determined
for normal and modified oligonucleotides. Our data showed that 5-CH3 groups of the first and
second T residues of the hexanucleotide sequence were essential contact points for ClaI
methylase. However, ClaI endonuclease was still active without either one of those methyl
groups with elevated Km values. Bromine could compensate significantly for the loss of 5-CH3
groups at both positions. On the other hand, the 2-amino group of the G residue appeared to be
an essential contact point for both enzymes. It has been concluded that ClaI enconuclease and
ClaI methylase recognize the sequence in different ways.

<>

<1>Kania, J., Fanning, T.G.
<2>Use of a Sequence-Specific DNA-Binding Ligand to probe the Environments of EcoRI Restriction Endonuclease Cleavage Sites.
<3>Eur. J. Biochem.
<4>67
<5>367-371
<6>1976
<7>The DNAs of bacteriophage lambda and adenovirus were incubated with the
sequence-specific DNA-binding ligand 6,4'-diamidino-2-phenylindole.  Digestion
of the ligant - DNA complexes with EcoRI nuclease and subsequent agarose gel
electrophoresis demonstrated that the ligand inhibited nuclease activity at
some sites, but not at others.  The results suggest that
diamidino-2-phenylindole can be used to probe the immediate environments of the
EcoRI cleavage sites.

<>

<1>Kankel, M.W., Ramsey, D.E., Stokes, T.L., Flowers, S.K., Haag, J.R., Jeddeloh, J.A., Riddle, N.C., Verbsky, M.L., Richards, E.J.
<2>Arabidopsis MET1 cytosine methyltransferase mutants.
<3>Genetics
<4>163
<5>1109-1122
<6>2003
<7>We describe the isolation and characterization of two missense mutations in the
cytosine-DNA-methyltransferase gene, MET1, from the flowering plant
Arabidopsis thaliana. Both missense mutations, which affect the catalytic
domain of the protein, led to a global reduction of cytosine methylation
throughout the genome. Surprisingly, the met1-2 allele, with the weaker
DNA hypomethylation phenotype, alters a well-conserved residue in
methyltransferase signature motif I. The stronger met1-1 allele caused
late flowering and a heterochronic delay in the juvenile-to-adult rosette
leaf transition. The distribution of late-flowering phenotypes in a
mapping population segregating met1-1 indicates that the flowering-time
phenotype is caused by the accumulation of inherited defects at loci
unlinked to the met1 mutation. The delay in flowering time is due in part
to the formation and inheritance of hypomethylated fwa epialleles, but
inherited defects at other loci are likely to contribute as well.
Centromeric repeat arrays hypomethylated in met1-1 mutants are partially
remethylated when introduced into a wild-type background, in contrast to
genomic sequences hypomethylated in ddm1 mutants. ddm1 met1 double mutants
were constructed to further our understanding of the mechanism of DDM1
action and the interaction between two major genetic loci affecting global
cytosine methylation levels in Arabidopsis.

<>

<1>Kankia, B.I.
<2>A real-time assay for monitoring nucleic acid cleavage by quadruplex formation.
<3>Nucleic Acids Res.
<4>34
<5>e141
<6>2006
<7>Direct and straightforward methods to follow nucleic acid cleavage are needed. A
spectrophotometric quadruplex formation assay (QFA) was
developed, which allows real-time monitoring of site-specific cleavage of
nucleic acids. QFA was applied to study both protein and nucleic acid
restriction enzymes, and was demonstrated to accurately determine
Michaelis-Menten parameters for the cleavage reaction catalyzed by EcoRI.
QFA can be used to study the mechanisms of protein-nucleic acid
recognition. QFA is also a useful tool for dissecting individual nicking
rates of a double-stranded cleavage.

<>

<1>Kannan, P., Cowan, G.M., Daniel, A.S., Gann, A.A.F., Murray, N.E.
<2>Conservation of organization in the specificity polypeptides of two families of Type I restriction enzymes.
<3>J. Mol. Biol.
<4>209
<5>335-344
<6>1989
<7>We have identified the recognition sequence for the Citrobacter freundii
restriction endonuclease CfrA, a member of the A-family of type I R-M enzymes.
This bipartite target sequence differs in both its components from those of
other type I enzymes.  We determined the nucleotide sequence of its specificity
gene (hsdS) and a comparison of this with its relative EcoA identifies two
extensive variable regions, an organization analogous to that found in the
K-family of type I R-M enzymes.  The specificity polypeptides of the A-family,
unlike those of K, have an N-terminal conserved region, and this includes a
sequence repeated within the central conserved region.  A second repeat
sequence, identified at the amino acid level, coincides with the only sequence
similarity common to all type I S polypeptides.  Sequences immediately
downstream from the hsdS genes of EcoA, CfrA, EcoK, B and D are almost
identical, consistent with an allelic chromosomal location.

<>

<1>Kannan, P.R., Dharmalingam, K.
<2>Restriction alleviation and enchancement of mutagenesis of the bacteriophage T4 chromosome in recBCsbcA strains of Escherichia coli.
<3>Mol. Gen. Genet.
<4>208
<5>413-418
<6>1987
<7>The restriction of non glucosylated phage T4 DNA is reduced significantly in
host bacterial strains carrying recBCsbcA mutations even in the presence of a
functional rgl gene.  In recBCsbcA hosts a high frequency of phage mutations
are observed both in the glucosyl transferase genes and in the DNA sequences
recognised by the rgl restriction enzymes.  This hypermutagenic property of the
recBCsbcA strains is not dependent on the glucosylation of the phage DNA, and
the mutagenesis is localized to certain regions of the T4 chromosome.  However,
alleviation of rgl restriction in recBCsbcA strains is due neither to the
increased mutagenesis, nor to the absence of a functional rgl system, since
second site mutations (rra) restore rgl restriction without affecting
hypermutagenesis.

<>

<1>Kansu, F., Glaser, P.
<2>Listeria innocua, genome and applications.
<3>Japanese Patent Office
<4>JP 2004515227 A
<5>
<6>2004
<7>
<>

<1>Kant, J.A.
<2>DNA restriction enzymes and RFLPs in medicine.
<3>Methods Biochem. Anal.
<4>36
<5>129-140
<6>1992
<7>*
    1 Restriction Enzymes (Endonucleases)
  1.1 General information
  1.2 Restriction enzymes and molecular cloning
  1.3 Restriction enzymes, chromosomal mapping, and gene expression
  1.4 Technical notes
    2 Restriction fragment-length polymorphisms
  2.1 RFLPs defined
  2.2 RFLPs and alleles in human genetic disorders
2.2.1 Linkage of RFLPs to genetic diseases
2.2.2 Restriction enzymes to detect disease-causing mutations
2.2.3 Haplotypes and linkage disequilibrium
  2.3 RFLPs in neoplasia and cancer
2.3.1 Loss of RFLP heterozygosity and tumor suppressor genes
2.3.2 Clonality analysis of hematolymphoid neoplasms
2.3.3 Consistent chromosomal translocations
2.3.4 Clonality studies of nonhematolymphoid tumors
  2.4 Infectious diseases
    3 Overview


<>

<1>Kant, R. et al.
<2>Genome Sequence of 'Pedosphaera parvula' Ellin514, an Aerobic Verrucomicrobial Isolate from Pasture Soil.
<3>J. Bacteriol.
<4>193
<5>2900-2901
<6>2011
<7>"Pedosphaera parvula" Ellin514 is an aerobically grown verrucomicrobial
isolate from pasture soil. It is one of the few cultured representatives
of subdivision 3 of the phylum Verrucomicrobia. Members of this group are
widespread in terrestrial environments.

<>

<1>Kant, R., Paulin, L., Alatalo, E., de Vos, W.M., Palva, A.
<2>Genome Sequence of Lactobacillus amylovorus GRL1112.
<3>J. Bacteriol.
<4>193
<5>789-790
<6>2010
<7>Lactobacillus amylovorus is a common member of the normal gastrointestinal tract (GIT)
microbiota in pigs. Here, we report the genome sequence of L.
amylovorus GRL1112, a porcine faeces isolate displaying strong adherence
to the pig intestinal epithelial cells. The strain is of interest as it is
a potential probiotic bacterium.

<>

<1>Kant, R., Paulin, L., Alatalo, E., de Vos, W.M., Palva, A.
<2>Genome sequence of Lactobacillus amylovorus GRL1118, isolated from the pig ileum.
<3>J. Bacteriol.
<4>193
<5>3147-3148
<6>2011
<7>Lactobacillus amylovorus is a common member of the beneficial microbiota present in the pig
gastrointestinal tract (GIT). Here, we report the genome sequence of surface layer (S-layer)
protein carrying, and potential probiotic L. amylovorus GRL1118, that was isolated from
porcine ileum and showed strong adherence to the pig intestinal epithelial cells.

<>

<1>Kapatral, V. et al.
<2>Genome sequence and analysis of the oral bacterium Fusobacterium nucleatum strain ATCC 25586.
<3>J. Bacteriol.
<4>184
<5>2005-2018
<6>2002
<7>We present a complete DNA sequence and metabolic analysis of the dominant oral bacterium
Fusobacterium nucleatum. Although not considered a major dental pathogen on its own, this
anaerobe facilitates the aggregation and establishment of several other species including the
dental pathogens Porphyromonas gingivalis and Bacteroides forsythus. The F. nucleatum strain
ATCC 25586 genome was assembled from shotgun sequences and analyzed using the ERGO
bioinformatics suite (http://www.integratedgenomics.com). The genome contains 2.17 Mb encoding
2,067 open reading frames, organized on a single circular chromosome with 27% GC content.
Despite its taxonomic position among the gram-negative bacteria, several features of its core
metabolism are similar to that of gram-positive Clostridium spp., Enterococcus spp., and
Lactococcus spp. The genome analysis has revealed several key aspects of the pathways of
organic acid, amino acid, carbohydrate, and lipid metabolism. Nine very-high-molecular-weight
outer membrane proteins are predicted from the sequence, none of which has been reported in
the literature. More than 137 transporters for the uptake of a variety of substrates such as
peptides, sugars, metal ions, and cofactors have been identified. Biosynthetic pathways exist
for only three amino acids: glutamate, aspartate, and asparagine. The remaining amino acids
are imported as such or as di- or oligopeptides that are subsequently degraded in the
cytoplasm. A principal source of energy appears to be the fermentation of glutamate to
butyrate. Additionally, desulfuration of cysteine and methionine yields ammonia, H2S, methyl
mercaptan, and butyrate, which are capable of arresting fibroblast growth, thus preventing
wound healing and aiding penetration of the gingival epithelium. The metabolic capabilities of
F. nucleatum revealed by its genome are therefore consistent with its specialized niche in the
mouth.

<>

<1>Kapatral, V., Ivanova, N., Anderson, I., Reznik, G., Bhattacharyya, A., Gardner, W.L., Mikhailova, N., Lapidus, A., Larsen, N., D'Souza, M., Walunas, T., Haselkorn, R., Overbeek, R., Kyrpides, N.
<2>Genome Analysis of F. nucleatum sub spp vincentii and Its Comparison With the Genome of F. nucleatum ATCC 25586.
<3>Genome Res.
<4>13
<5>1180-1189
<6>2003
<7>We present the draft genome sequence and its analysis for Fusobacterium nucleatum sub spp.
vincentii (FNV), and compare that genome with F.
nucleatum ATCC 25586 (FN). A total of 441 FNV open reading frames (ORFs)
with no orthologs in FN have been identified. Of these, 118 ORFs have no
known function and are unique to FNV, whereas 323 ORFs have functional
orthologs in other organisms. In addition to the excretion of butyrate,
H2S and ammonia-like FN, FNV has the additional capability to excrete
lactate and aminobutyrate. Unlike FN, FNV is likely to incorporate
galactopyranose, galacturonate, and sialic acid into its O-antigen. It
appears to transport ferrous iron by an anaerobic ferrous transporter.
Genes for eukaryotic type serine/threonine kinase and phosphatase,
transpeptidase E-transglycosylase Pbp1A are found in FNV but not in FN.
Unique ABC transporters, cryptic phages, and three types of
restriction-modification systems have been identified in FNV. ORFs for
ethanolamine utilization, thermostable carboxypeptidase, gamma
glutamyl-transpeptidase, and deblocking aminopeptidases are absent from
FNV. FNV, like FN, lacks the classical catalase-peroxidase system, but
thioredoxin/glutaredoxin enzymes might alleviate oxidative stress. Genes
for resistance to antibiotics such as acriflavin, bacitracin, bleomycin,
daunorubicin, florfenicol, and other general multidrug resistance are
present. These capabilities allow Fusobacteria to survive in a mixed
culture in the mouth.

<>

<1>Kapetaniou, E.G., Kotsifaki, D., Providaki, M., Rina, M., Bouriotis, V., Kokkinidis, M.
<2>Purification, crystallization and preliminary X-ray analysis of the BseCI DNA methyltransferase from Bacillus stearothermophilus in complex with its cognate DNA.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>63
<5>12-14
<6>2007
<7>The DNA methyltransferase M. BseCI from Bacillus stearothermophilus (EC 2.1.1.72), a
579-amino-acid enzyme, methylates the N6 atom of the 30
adenine in the sequence 50-ATCGAT-3'. M.BseCI was crystallized in
complex with its cognate DNA. The crystals were found to belong to the
hexagonal space group P6, with unit-cell parameters a = b = 87.0, c =
156.1 angstrom, beta = 120.0 degrees and one molecule in the asymmetric
unit. Two complete data sets were collected at wavelengths of 1.1 and
2.0 angstrom to 2.5 and 2.8 angstrom resolution, respectively, using
synchrotron radiation at 100 K.

<>

<1>Kapfer, W., Walter, J., Trautner, T.A.
<2>Cloning, characterization and evolution of the BsuFI restriction endonuclease gene of Bacillus subtilis and purification of the enzyme.
<3>Nucleic Acids Res.
<4>19
<5>6457-6463
<6>1991
<7>The restriction endonuclease (R.BsuFI) of Bacillus subtilis recognizes the target DNA sequence
5' CCGG. The R.BsuFI gene was found in close proximity to the cognate M.BsuFI gene, which had
previously been characterized. Cloning of the R.BsuFI gene in E. coli was only possible with
the M.BsuFI Mtase gene present on a compatible plasmid. The cloned R.BsuFI gene was expressed
in E. coli and restriction activity was observed in vivo and in vitro. The R.BsuFI gene
consists of 1185 bp, coding for a protein of 395 amino acids with a calculated molecular
weight of 45.6 kD. The R.BsuFI enzyme was purified to homogeneity following overexpression. It
presumably works as a dimer and cleaves the 5' CCGG target sequence between the two cytosines
to produce sticky ends with 5' CG overhangs, like the isoschizomers R.MspI and R.HpaII. The
relatedness between R.BsuFI and R.MspI is reflected by significant similarities of the amino
acid sequences of both enzymes. This is the first case where such similarities have been
observed between isoschizomeric restriction endonucleases which belong to 5mC specific R/M
systems. This observation suggests that R.BsuFI and R.MspI genes derive from a common
ancestor. In spite of such functional and evolutionary relatedness, the R/M systems differ in
the arrangement of their R and M genes. In the BsuFI system transcription of the two genes is
convergent, whereas divergent transcription occurs in the MspI system.

<>

<1>Kapfhammer, D., Blass, J., Evers, S., Reidl, J.
<2>Vibrio cholerae phage K139: complete genome sequence and comparative genomics of related phages.
<3>J. Bacteriol.
<4>184
<5>6592-6601
<6>2002
<7>In this report, we characterize the complete genome sequence of the
temperate phage K139, which morphologically belongs to the Myoviridae
phage family (P2 and 186). The prophage genome consists of 33,106 bp, and
the overall GC content is 48.9%. Forty-four open reading frames were
identified. Homology analysis and motif search were used to assign
possible functions for the genes, revealing a close relationship to
P2-like phages. By Southern blot screening of a Vibrio cholerae strain
collection, two highly K139-related phage sequences were detected in
non-O1, non-O139 strains. Combinatorial PCR analysis revealed almost
identical genome organizations. One region of variable gene content was
identified and sequenced. Additionally, the tail fiber genes were
analyzed, leading to the identification of putative host-specific sequence
variations. Furthermore, a K139-encoded Dam methyltransferase was
characterized.

<>

<1>Kaplan, D.A., Nierlich, D.P.
<2>Cleavage of nonglucosylated bacteriophage T4 deoxyribonucleic acid by restriction endonuclease EcoRI.
<3>J. Biol. Chem.
<4>250
<5>2395-2397
<6>1975
<7>DNAs lacking the glucosyl modification (Glc-) and additionally lacking the
6-methylaminopurine (N6-methyladenine) modification (Glc-, MeAde-) were
prepared from appropriate T4 mutants.  These DNAs were cleaved by the purified
restriction endonuclease EcoRI from Escherichia coli.  Normally modified DNA
(Glc+, MeAde+) was not attacked.  The EcoRII and the Hemophilus enzymes HindII
and HindIII do not attack Glc-, MeAde- T4 DNA, possibly due to the presence of
6-hydroxymethylcytosine.  EcoRI produces approximately 40 specific fragments
from Glc- DNA ranging in molecular weights from 0.3 to 10.5x10/6.

<>

<1>Kaplan, D.J.
<2>Variation in the inhibition of restriction enzyme cleavage of lambda phage DNA produced by two covalent binding antitumor agents:  Anthramycin and Mitomycin C.
<3>Biochem. Biophys. Res. Commun.
<4>109
<5>639-648
<6>1982
<7>DNA-drug complexes containing various levels of covalently bound mitomycin C
(MC) or anthramycin were subjected to the actions of a number of restriction
enzymes.  While MC presented only a partial block to the actions of a number of
these enzymes, anthramycin, at high binding ratios, blocked enzymatic activity
very well.  The contrast seen in the restriction cleavage of these DNA-drug
complexes may be related to the different points of attachement in DNA (minor
groove vs. major groove) for these drugs.  Although similarities in
electrophoretic band patterns exist for both drug complexes, certain
differences indicative of preferences in binding sequences do not necessarily
lie immediately within the restriction cut sites but may effect the cutting of
these sites from a distance.  The results also further support anthramycin's
potential usage as a selective/reversible blocking agent for recombinant
research.

<>

<1>Kaplan, J.B., Lo, C., Xie, G., Johnson, S.L., Chain, P.S., Donnelly, R., Kachlany, S.C., Balashova, N.V.
<2>Genome Sequence of Kingella kingae Septic Arthritis Isolate PYKK081.
<3>J. Bacteriol.
<4>194
<5>3017
<6>2012
<7>Kingella kingae is a human oral bacterium that can cause infections of the skeletal system in
children. The bacterium is also a cardiovascular pathogen
causing infective endocarditis in children and adults. We report herein the draft
genome sequence of septic arthritis K. kingae strain PYKK081.

<>

<1>Kapley, A., Sagarkar, S., Tanksale, H., Sharma, N., Qureshi, A., Khardenavis, A., Purohit, H.J.
<2>Genome Sequence of Alcaligenes sp. Strain HPC1271.
<3>Genome Announcements
<4>1
<5>e00235-12
<6>2013
<7>We report a draft genome sequence of sp. strain HPC1271, which demonstrates antimicrobial
activity against multidrug-resistant bacteria. Antibiotic
production by has not been frequently reported, and hence, the availability of
the genome sequence should enable us to explore new antibiotic-producing gene
clusters.

<>

<1>Kappelman, J.R., Brady, M., Knoche, K., Murray, E., Schoenfeld, T., Williams, R., Vesselinova, N.
<2>SgfI, a new type-II restriction endonuclease that recognizes the octanucleotide sequence 5'-GCGAT/CGC-3'.
<3>Gene
<4>160
<5>55-58
<6>1995
<7>A new restriction endonuclease (ENase), SgfI, has been isolated from the bacterium
Streptomyces sp. SgfI recognizes the 8-bp palindrome 5'-GCGATCGC-3' and cleaves
double-stranded DNA after the T in this sequence, producing a two-base 3' overhang compatible
with PvuI termini.  SgfI is a rare-cutting ENase and should be useful for megabase mapping
experiments.

<>

<1>Kappelman, J.R., Williams, R.J., Murray, E.E., Vesselinova, N.I.
<2>Restriction endonuclease SgfI from Streptomyces griseoruber.
<3>US Patent Office
<4>US 5391487
<5>
<6>1995
<7>*
A restriction endonuclease is provided. The enzyme, designated SgfI, recognizes the DNA
sequence:

    5'-GCG AT^CGC-3'
    3'-CGC^TA GCG-5'

and cleaves the DNA sequence at a position indicated by the arrows and is preferably
obtainable from microorganisms of the genus Streptomyces.


<>

<1>Kappler, U. et al.
<2>Complete genome sequence of the facultatively chemolithoautotrophic and methylotrophic alpha Proteobacterium Starkeya novella type strain (ATCC 8093(T)).
<3>Standards in Genomic Sciences
<4>7
<5>44-58
<6>2012
<7>(Starkey 1934) Kelly . 2000 is a member of the family in the order , which is thus far poorly
characterized at the genome level. Cultures from this species are
most interesting due to their facultatively chemolithoautotrophic lifestyle,
which allows them to both consume carbon dioxide and to produce it. This feature
makes an interesting model organism for studying the genomic basis of regulatory
networks required for the switch between consumption and production of carbon
dioxide, a key component of the global carbon cycle. In addition, is of interest
for its ability to grow on various inorganic sulfur compounds and several
C1-compounds such as methanol. Besides , is only the second species in the family
with a completely sequenced genome of a type strain. The current taxonomic
classification of this group is in significant conflict with the 16S rRNA data.
The genomic data indicate that the physiological capabilities of the organism
might have been underestimated. The 4,765,023 bp long chromosome with its 4,511
protein-coding and 52 RNA genes was sequenced as part of the DOE Joint Genome
Institute Community Sequencing Program (CSP) 2008.

<>

<1>Kapur, V., Amonjin, A., Li, L., Zhang, K., Bannantine, J.P.
<2>Mycobacterial Diagnostics.
<3>Japanese Patent Office
<4>JP 2006514551 A
<5>
<6>2006
<7>
<>

<1>Kapur, V., Gebuhato, C.J.
<2>Nucleic acid and polypeptide sequences from Lawsonia Intracellularis and methods of using.
<3>Japanese Patent Office
<4>JP 2006501846 A
<5>
<6>2006
<7>
<>

<1>Kaput, J., Sneider, T.W.
<2>Methylation of somatic vs germ cell DNAs analyzed by restriction endonuclease digestions.
<3>Nucleic Acids Res.
<4>7
<5>2303-2322
<6>1979
<7>Bacterial restriction endonucleases containing the dinucleotide CpG in their
cleavage sequences were used to compare the methylation patterns of primarily
repeated DNA sequences in (1) bovine somatic cell native DNAs vs bovine sperm
cell native DNA and (2) native vs renatured bovine liver and sperm cell DNAs.
The restriction patterns of sperm native DNA differ markedly from those of
somatic cell native DNAs when using HpaII, HhaI, and AvaI but not when using
the enzymes EcoRI and MspI.  Digestion patterns of germ cell renatured DNA
differed significantly from those of germ cell native DNA when using HpaII but
not when using MspI or EcoRI.  The results may not be due to artifacts of
renaturation of the DNAs.  The results are consistent with the concept that
germ cell DNA may be strand asymmetrically hemimethylated.  The data also
suggest that methylation of the 5'-cytosine in the sequence CCGG renders this
site insensitive to cleavage by MspI.

<>

<1>Karabecov, B.P., Oganesian, M.G., Airumian, B.A.
<2>Susceptibility of the bacteriophage T5 to host-controlled restriction and modification in the Salmonella derby strain 456.
<3>Genetika
<4>6
<5>121-128
<6>1970
<7>It was shown that wild type of bacteriophage T5 (T5h+) was not susceptible to
host-controlled variation in a bacterial host control system, consisting of E.
coli strains B, K-12 or BB and Salmonella derby 456 strain.  The h-mutants of
T5 phage, obtained on B/1,5 strain of E. coli, became susceptible to
restriction and modification on s. derby 456.  The phage T5h, grown on E. coli
B, K-12 or BB cells showed an efficiency of plating (eop) on S. derby 456 about
10/7 - 10/9.  a small number of plaques formed by these mutants on S. derby
456, contained only modified phage T5h 456 possessing eop equal to 1.0 when
grown on E. coli strains or on S. derby 456.  Acquirement of susceptibility to
host-controlled variation by phage T5 was due to host range mutation, which
indicates that the susceptibility of the phages to this variation is an
inheritant character of phage DNA, and is subjected to genetic changes and
selection.  The data obtained allow a suggestion concerning phage genome
contribution to the processes of the host controlled restriction and
modification.  The rate of restriction of phage T5h, observed in S. derby 456
host, after maintaining on E. coli strains and T5h+ phage non-susceptibility to
the observed effect, indicate that this system of HCV (host-controlled
variation) might be used as a new experimental model for investigating
processes of mutational changes of susceptibility of DNA molecule to the
restricting and modifying mechanisms of host cells.

<>

<1>Karamov, E.V., Naroditsky, B.S., Zavizion, B.A., Tikhonenko, T.I.
<2>Transformation of EcoRI restriction endonuclease substrate specificity under the influence of glycerol.
<3>Biull. Eksp. Biol. Med.
<4>84
<5>46-48
<6>1977
<7>The restriction endonuclease EcoRI hydrolyzes DNA to a greater number of fragments in the
presence of glycerol than under normal conditions.  This enzyme begins to work by the
so-called EcoRI*-type of restriction when glycerol concentration reaches 50%.  The EcoRI*
activity appeared in experiments only when the ionic strength of the solution was decreased
and pH of the solution was increased.  However, under such extreme conditions the enzyme was
quickly inactivated and it was difficult to obtain reproducible results especially for
hydrolysis of the high-molecular DNA.  The suggested conditions for the EcoRI* activity permit
to obtain reproducible results, this being practically equivalent to discovery of the new
restriction endonuclease.

<>

<1>Karas, B.J., Jablanovic, J., Sun, L., Ma, L., Goldgof, G.M., Stam, J., Ramon, A., Manary, M.J., Winzeler, E.A., Venter, J.C., Weyman, P.D., Gibson, D.G., Glass, J.I., Hutchison, C.A.I.I.I., Smith, H.O., Suzuki, Y.
<2>Direct transfer of whole genomes from bacteria to yeast.
<3>Nat. Methods
<4>10
<5>410-412
<6>2013
<7>Transfer of genomes into yeast facilitates genome engineering for genetically intractable
organisms, but this process has been hampered by the need for cumbersome isolation of intact
genomes before transfer. Here we demonstrate direct cell-to-cell transfer of bacterial genomes
as large as 1.8 megabases (Mb)into yeast under conditions that promote cell fusion. Moreover,
we discovered that removal of restriction endonucleases from donor bacteria resulted in the
enhancement of genome transfer.

<>

<1>Karashi, H., Eda, S., Uehara, K., Nishida, K., Matsuhisa, A.
<2>PROBE FOR DIAGNOSIS OF INFECTIOUS DISEASE.
<3>Japanese Patent Office
<4>JP 1998033179 A
<5>
<6>1998
<7>
<>

<1>Karched, M., Furgang, D., Planet, P.J., Desalle, R., Fine, D.H.
<2>Genome Sequence of Aggregatibacter actinomycetemcomitans RHAA1, Isolated from a Rhesus Macaque, an Old World Primate.
<3>J. Bacteriol.
<4>194
<5>1275-1276
<6>2012
<7>Aggregatibacter actinomycetemcomitans is implicated in localized aggressive periodontitis. We
report the first genome sequence of an A. actinomycetemcomitans
strain isolated from an Old World primate.

<>

<1>Karczmarczyk, M., Wang, J., Leonard, N., Fanning, S.
<2>Complete nucleotide sequence of a conjugative IncF plasmid from an Escherichia coli isolate of equine origin containing blaCMY-2 within a novel genetic context.
<3>FEMS Microbiol. Lett.
<4>352
<5>123-127
<6>2014
<7>A blaCMY-2 -containing conjugative IncF plasmid denoted as pEQ011, previously
identified in a multidrug-resistant Escherichia coli isolate of equine origin,
was characterized. The plasmid consisted of 85 507 bp, with 118 predicted open
reading frames. This is the first known report demonstrating the association of a
blaCMY-2 gene with an IncF incompatibility-type plasmid backbone. A novel genetic
arrangement was identified wherein the blaCMY-2 resistance gene was proximally
flanked by IS1294 along with a partial blc gene located distally and within a
yacABC operon.

<>

<1>Karlin, S., Burge, C., Campbell, A.M.
<2>Statistical analyses of counts and distributions of restriction sites in DNA sequences.
<3>Nucleic Acids Res.
<4>20
<5>1363-1370
<6>1992
<7>Counts and spacings of all 4- and 6-bp palindromes in DNA sequences from a
broad range of organisms were investigated.  Both 4- and 6-bp average
palindrome counts were significantly low in all bacteriophages except one,
probably as a means of avoiding restriction enzyme cleavage.  The exception, T4
of normal 4- and 6-palindrome counts, putatively derives protection from
modification of cytosine to hydroxymethylcytosine plus glycosylation.  The
counts and distributions of 4-bp and of 6-bp restriction sites in bacterial
species are variable.  Bacterial cells with multiple restriction systems for
4-bp or 6-bp target specificities are low in aggregate 4- or 6-bp palindrome
counts/kb, respectively, but bacterial cells lacking exact 4-cutter enzymes
generally show normal or high counts of 4-bp palindromes when compared with
random control sequences of comparable nucleotide frequencies.  For example, E.
coli, apparently without an exact 4-bp target restriction endonuclease (see
text), contains normal aggregate 4-palindrome counts/kb, while B. subtilis,
which abounds with 4-bp restriction system, shows a significant
under-representation of 4-palindrome counts.  Both E. coli and B. subtilis have
many 6-bp restriction enzymes and concomitantly diminished aggregate
6-palindrome counts/kb.  Eukaryote, viral, and organelle sequences generally
have aggregate 4- and 6-palindromic counts/kb in the normal range.
Interpretations of these results are given in terms of restriction/methylation
regimes, recombination and transcription processes, and possible structural and
regulatory roles of 4- and 6-bp palindromes.

<>

<1>Karlovsky, P.
<2>Calculation of individual cleavage rates from partial digests in restriction endonuclease kinetics.
<3>J. Theor. Biol.
<4>132
<5>7-14
<6>1988
<7>A correction for results obtained by an analysis of DNA molecules partially
cleaved with restriction endonuclease was suggested.  The correction was proved
on model data.  Applications to (i) electron-microscopic analysis of singly
cleaved molecules, (ii) partial digestion of a circular molecule followed by
complete digestion with a second enzyme, (iii) systems with great cleavage
differences, and (iv) partial cleavage of end-labelled molecules were
discussed.

<>

<1>Karlovsky, P.
<2>Kinetics of circular DNA molecule digestion by restriction endonuclease A: Computation of kinetic constants from time dependence of fragment concentrations.
<3>Acta Biotheor.
<4>35
<5>279-292
<6>1986
<7>A model for kinetics of circular substrate cleavage by restriction endonuclease
was formulated.  The aim of the analysis of the model was to extract kinetic
constants for all target sites from time-dependence of fragment concentration
in reaction products.  That was proved to be possible for molecules with an odd
number of fragments only.  A symmetry of the molecules with an even number of
fragment is the cause.  A solution for molecules witih an odd number of
fragments was found and methods for dealing with the other moleculars were
suggested.

<>

<1>Karlovsky, P.
<2>A re-evaluation of evidence attributing the difference in cleavage rates of restriction endonuclease at different sites in the substrate to differences in KM values.
<3>Biochem. J.
<4>235
<5>611-612
<6>1986
<7>The only result supporting the hypothesis that the differences in restriction
endonuclease cleavage rates at various target sites are caused by differences
in KM values was reported by Forsblom, Rigler, Ehrenberg, Petterson & Philipson
[(1976) Nucl. Acids Res. 3, 3255-3269].  The present work shows that the
kinetic analysis in that paper is based on incorrect derivation and in fact
provides no support for the hypothesis mentioned.

<>

<1>Karlovsky, P.
<2>Mutual competitive inhibition between target sites during the restriction endonuclease digestion of DNA.
<3>J. Theor. Biol.
<4>132
<5>1-6
<6>1988
<7>The reaction rate of restriction endonuclease was evaluated theoretically, considering the
competition between target sites and a nonspecific DNA.  An equation for the initial cleavage
rate at a single site for a DNA substrate containing more than one recognition site was
derived.  The consequences for the study of preferential cleavage were discussed.

<>

<1>Karlyshev, A.V., Abramov, V.M.
<2>Draft Genome Sequence of Lactobacillus plantarum 2165.
<3>Genome Announcements
<4>2
<5>e01179-13
<6>2014
<7>This report describes a draft genome sequence of Lactobacillus plantarum 2165. The data
demonstrate the presence of a large number of genes responsible for
sugar metabolism and the fermentation activity of this bacterium. Different cell
surface proteins, including fibronectin and mucus-binding adhesins, may
contribute to the beneficial probiotic properties of this strain.

<>

<1>Karlyshev, A.V., Melnikov, V.G.
<2>Draft Genome Sequence of Corynebacterium pseudodiphtheriticum Strain 090104 'Sokolov'.
<3>Genome Announcements
<4>1
<5>e00921-13
<6>2013
<7>This report describes the first draft genome sequence of a Corynebacterium
pseudodiphtheriticum strain. The information on the genome organization and
putative gene products will assist in better understanding of the molecular
mechanisms involved in the beneficial probiotic effects of this bacterium.

<>

<1>Karlyshev, A.V., Melnikov, V.G., Chikindas, M.L.
<2>Draft Genome Sequence of Bacillus subtilis strain KATMIRA1933.
<3>Genome Announcements
<4>2
<5>e00619-14
<6>2014
<7>In this report, we present a draft sequence of Bacillus subtilis KATMIRA1933. Previous studies
demonstrated probiotic properties of this strain partially
attributed to production of an antibacterial compound, subtilosin. Comparative
analysis of this strain's genome with that of a commercial probiotic strain, B.
subtilis Natto, is presented.

<>

<1>Karlyshev, A.V., Melnikov, V.G., Chistyakov, V.A.
<2>Draft Genome Sequence of Bacillus amyloliquefaciens B-1895.
<3>Genome Announcements
<4>2
<5>e00633-14
<6>2014
<7>In this report, we present a draft genome sequence of Bacillus amyloliquefaciens  strain
B-1895. Comparison with the genome of a reference strain demonstrated
similar overall organization, as well as differences involving large gene
clusters.

<>

<1>Karlyshev, A.V., Melnikov, V.G., Khlebnikov, V.C., Abramov, V.M.
<2>Draft Genome Sequence of Lactobacillus crispatus 2029.
<3>Genome Announcements
<4>2
<5>e01221-13
<6>2014
<7>This report describes a draft genome sequence of Lactobacillus crispatus 2029. The reads
generated by the Ion Torrent PGM were assembled into contigs with a
total size of 2.2 Mb. The data were annotated using the NCBI GenBank and RAST
servers. A comparison with the reference strain revealed specific features of the
genome.

<>

<1>Karlyshev, A.V., Melnikov, V.G., Kosarev, I.V., Abramov, V.M.
<2>Draft Genome Sequence of Lactobacillus rhamnosus 2166.
<3>Genome Announcements
<4>2
<5>e01222-13
<6>2014
<7>In this report, we present a draft sequence of the genome of Lactobacillus rhamnosus strain
2166, a potential novel probiotic. Genome annotation and read
mapping onto a reference genome of L. rhamnosus strain GG allowed for the
identification of the differences and similarities in the genomic contents and
gene arrangements of these strains.

<>

<1>Karlyshev, A.V., Melnikov, V.G., Kosarev, I.V., Khlebnikov, V.C., Sukhikh, G.T., Abramov, V.M.
<2>Draft Genome Sequence of Lactobacillus gasseri Strain 2016.
<3>Genome Announcements
<4>1
<5>e00624-13
<6>2013
<7>Different common factors contribute to the antagonistic properties of Lactobacillus gasseri
toward various pathogens. However, there is
strain-to-strain variation in the probiotic properties of this bacterium. The
draft genome sequence of L. gasseri strain 2016 determined in this study will
assist in understanding the genetic basis for such variation.

<>

<1>Karlyshev, A.V., Nadarajah, S., Abramov, V.M.
<2>Draft Genome Sequence of Lactobacillus jensenii Strain MD IIE-70(2).
<3>Genome Announcements
<4>1
<5>e01005-13
<6>2013
<7>A draft genome sequence of Lactobacillus jensenii strain MD IIE-70(2) was determined using Ion
PGM technology. The reads were mapped to a reference strain
and assembled using a combination of tools. The genetic features revealed in this
study will assist in understanding the probiotic properties of Lactobacillus
bacteria.

<>

<1>Karlyshev, A.V., Raju, K., Abramov, V.M.
<2>Draft Genome Sequence of Lactobacillus fermentum Strain 3872.
<3>Genome Announcements
<4>1
<5>e01006-13
<6>2013
<7>This report describes a draft genome sequence of Lactobacillus fermentum strain 3872. The data
revealed remarkable similarity to and dissimilarity with the
published genome sequences of other strains of the species. The absence of and
variation in structures of some adhesins and the presence of an additional
adhesin may reflect adaptation of the bacterium to different host systems and may
contribute to specific properties of this strain as a new probiotic.

<>

<1>Karlyshev, A.V., Robyn, J., Rasschaert, G., Heyndrickx, M.
<2>Draft Genome Sequence of Enterococcus faecalis MB5259.
<3>Genome Announcements
<4>2
<5>e00634-14
<6>2014
<7>In this study, we present a draft genome sequence of Enterococcus faecalis MB5259, a promising
probiotic strain. The identified differences and common
features between this strain and reference strains will assist in better
understanding the mechanism of antibacterial action and in developing novel
probiotics.

<>

<1>Karpova, E., Kubareva, E., Petrauskene, O., Joschimiak, A., Van Kley, H.
<2>Peculiarity of the recognition and cleavage by restriction endonuclease EcoRII.
<3>FASEB J.
<4>9
<5>A1400
<6>1995
<7>The restriction endonuclease EcoRII binds with high affinity to the duplex DNA 5' CC(A/T)GG
3' and specifically cleaves it at the 5' end of the site as indicated by the arrow. It has
been shown that EcoRII requires an auxiliary site for activation, and the formation of a
four-strand supersite was suggested. To understand the enzymatic mechanism and the interaction
of EcoRII with two recognition sites, we have studied binding and cleavage of DNA duplexes
containing canonical or modified EcoRII sites by EcoRII. The reactions were analyzed by
polyacrylamide gel electrophoresis under non-denaturing conditions. We have shown, that (T to
5-fluorodeoxyuridine) or (A to N6-methyldeoxyadenosine) substitutions in the central base pair
do not affect DNA recognition, but they have a dramatic influence on cleavage. At the same
time modification or substitution of C or G nucleotides in the recognition sequence affects
both recognition and rate of cleavage of DNA. These data suggests that specific interaction
with the central base pair (A/T) contributes to the catalytic activity of the enzyme. Based on
these data we present a model for four-strand supersite formation of the enzyme-substrate
complex.

<>

<1>Karpova, E.A., Kubareva, E.A., Buryanov, Y.I., Gromova, E.S.
<2>Formation of two types of enzyme-substrate complexes at the interaction of EcoRII restriction endonuclese with synthetic DNA duplexes.
<3>Mol. Biol. (Mosk)
<4>26
<5>993-998
<6>1992
<7>Binding of EcoRII restriction endonuclease to synthetic oligodeoxyribonucleotide substrates of
11-30 base pairs long was investigated by polyacrylamide gel electrophoresis under
nondenaturing conditions in the absence of Mg2+ ions. Irrespective of the length of a
substrate, two types of specific DNA-protein complexes were shown to be formed. Their mobility
in gels was close to that of the monomer (45 kDa) and dimer (90 kDa) of marker protein
ovalbumin. The ratio of these complexes in solution depended on that of the molar
concentrations of EcoRII restriction endonuclease and DNA duplexes. The possible structure of
the complexes is discussed.

<>

<1>Karpova, E.A., Kubareva, E.A., Gromova, E.S., Buryanov, Y.I.
<2>Peculiaritites of the binding of restriction endonuclease EcoRII to synthetic DNA duplexes.
<3>Biochem. Mol. Biol. Int.
<4>29
<5>113-121
<6>1993
<7>The binding of restriction endonuclease EcoRII to synthetic oligodeoxyribonucleotide
substrates 11 to 30 bp long was investigated by gradient polyacrylamide gel electrophoresis
under nondenaturing conditions in the absence of Mg2+ ions. Irrespective of the substrate's
length, two types of specific DNA-protein complexes were shown to be formed. Their mobility in
gels was close to that of the monomer and the dimer of the marker ovalbumin. The number of
such complexes in solution depended on the ratio of the molar concentrations of restriction
endonuclease EcoRII and the DNA duplex. The possible structure of the complexes is discussed.

<>

<1>Karpova, E.A., Kubareva, E.A., Shabarova, Z.A.
<2>A model of EcoRII restriction endonuclease action: The active complex is most likely formed by one protein subunit and one DNA recognition site.
<3>Life
<4>48
<5>91-98
<6>1999
<7>To elucidate the mechanism of interaction of restriction endonuclease EcoRII with DNA, we
studied by native gel electrophoresis the binding of this endonuclease to a set of synthetic
DNA-duplexes containing the modified or canonical recognition sequence 5'-d(CCA/TGG)-3'.
All binding substrates or substrate analogues tested could be divided into two major groups:
(I) duplexes that, at the interaction with endonuclease EcoRII, form two types of stable
complexes on native gel in the absence of Mg2+ cofactor; (ii) duplexes that form only one type
of complex, observed both in the presence and absence of Mg2+.  Unlike the latter, duplexes
under the first group can be hydrolyzed by endonuclease.  Data obtained suggest that the
active complex is most likely formed by one protein subunit and one DNA recognition sequence.
A model of EcoRII endonuclease action is presented.

<>

<1>Karpova, E.A., Meehan, E., Pusey, M.L., Chen, L.
<2>Crystallization and preliminary x-ray diffraction analysis of restriction endonuclease EcoRII.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>55
<5>1604-1605
<6>1999
<7>Crystals of the restriction endonuclease EcoRII have been obtained by the vapor-diffusion
technique in the presence of ammonium sulfate or polyethylene glycol. The best crystals were
grown with ammonium sulfate as a precipitant. Crystals with dimensions of up to 0.6 x 0. 6 x
0.6 mm have been observed. The crystals diffract to about 4.0 A resolution at a
cryo-temperature of 100 K using a rotating-anode X-ray source and a Rigaku R-AXIS IV
imaging-plate detector. The space group has been determined to be either I23 or I2(1)3, with
unit-cell parameters a = b = c = 160.3 A, alpha = beta = gamma = 90 degrees. The crystal
asymmetric unit contains two protein molecules, and self-rotation function analysis shows a
pseudo-twofold symmetry relating the two monomers. Attempts to improve the resolution of
crystal diffraction and to search for heavy-atom derivatives are under way.

<>

<1>Karreman, C.
<2>The genes for two procaryotic DNA modification methylases.
<3>Ph.D. Thesis, State University of Leiden, Holland
<4>
<5>1-95
<6>1988
<7>None

<>

<1>Karreman, C., de Waard, A.
<2>Isolation and characterization of the modification methylase M.SinI.
<3>J. Bacteriol.
<4>170
<5>2533-2536
<6>1988
<7>A sequence-specific modification methylase (M.SinI) was isolated and purified from Escherichia
coli harboring a derivative of recombinant plasmid pSI4 (see accompanying manuscript: C.
Karreman and A. de Waard, J. Bacteriol. 170:2527-2532, 1988), which contains a Salmonella
infantis DNA insert. The enzyme uniquely methylates the internal deoxycytidylate residue in
the nucleotide sequence GG(A/T)MeCC, thereby protecting DNA completely against cleavage by
restriction endonuclease R.SinI or R.AvaII [GG(A/T)CC], and in part against cleavage by
R.Sau96I (GGNCC).

<>

<1>Karreman, C., de Waard, A.
<2>Agmenellum quadruplicatum M.AquI, a novel modification methylase.
<3>J. Bacteriol.
<4>172
<5>266-272
<6>1990
<7>The complete type II modification methylase of Agmenellum quadruplicatum was cloned in
Escherichia coli as an R.Sau3A fragment of approximately 4.5 kilobases. The coding sequence
was contained in a stretch of 1,156 base pairs which was organized into two parallel, partly
overlapping open reading frames of 248 and 139 codons. In vivo complementation experiments
showed that the synthesis of both predicted peptides was required for full methylase activity.
The amino acid sequences were considerably similar to regions of other deoxycytidylate
methylases.

<>

<1>Karreman, C., de Waard, A.
<2>Cloning and complete nucleotide sequences of the TypeII restriction-modification genes of Salmonella infantis.
<3>J. Bacteriol.
<4>170
<5>2527-2532
<6>1988
<7>The complete typeII restriction-modification system of Salmonella infantis was
cloned in Escherichia coli as an R.Sau3AI fragment of 3,430 base pairs.  The
clone was shown to express the restriction endonuclease as well as the
modification methylase.  The nucleotide sequence of the above fragment showed
two open reading frames of 461 and 230 codons in tail-to-tail orientation.
These were shown to represent the modification methylase M.SinI and the
restriction endonuclease R.SinI, respectively.  The methylase M.SinI amino acid
sequence revealed a considerable similarity to those of other deoxycytidylate
methylases.  In contrast, endonuclease R.SinI did not exhibit such a similarity
to other restriction enzymes.

<>

<1>Karreman, C., Tandeau de Marsac, N., de Waard, A.
<2>Isolation of a deoxycytidylate methyl transferase capable of protecting DNA uniquely against cleavage by endonuclease R. AquI (isoschizomer of AvaI).
<3>Nucleic Acids Res.
<4>14
<5>5199-5205
<6>1986
<7>A sequence-specific modification methylase (M.AquI) was isolated and purified
from Agmenellum quadruplicatum (Synechococcus PCC 7002).  This enzyme uniquely
methylates the deoxycytidylate residue in the sequence *CYCGRG indicated by the
asterisk.  It was shown to protect DNA against cleavage by restriction
endonucleases AvaI, SmaI and XhoI, which recognize the sequences CYCGRG,
CCCGGG, and CTCGAG, respectively.

<>

<1>Karrer, K.M., VanNuland, T.A.
<2>Position effect takes precedence over target sequence in determination of adenine methylation patterns in the nuclear genome of a eukaryote, Tetrahymena thermophila.
<3>Nucleic Acids Res.
<4>26
<5>4566-4573
<6>1998
<7>Approximately 0.8% of the adenine residues in the macronuclear DNA of the ciliated protozoan
Tetrahymena thermophila are modified to N6-methyladenine.  DNA methylation is site specific
and the pattern of methylation is constant between clonal cell lines.  In vivo, modification
of adenine residues appears to occur exclusively in the sequence 5'-NAT-3', but no consensus
sequence for modified sites has been found.  In this study, DNA fragments containing a site
that is uniformly methylated on the 50 copies of the macronuclear chromosome were cloned into
the extrachromosomal rDNA.  In the novel location on the rDNA minichromosome, the site was
unmethylated.  The result was the same whether the sequences were introduced in a methylated
or unmethylated state and regardless of the orientation of the sequence with respect to the
origin of DNA replication.  The data show that sequence is insufficient to account for
site-specific methylation in Tetrahymena and argue that other factors determine the pattern of
DNA methylation.

<>

<1>Karrer, K.M., VanNuland, T.A.
<2>Methylation of adenine in the nuclear DNA of Tetrahymena is internucleosomal and independent of histone H1.
<3>Nucleic Acids Res.
<4>30
<5>1364-1370
<6>2002
<7>There are about 50 copies of each chromosome in the somatic macronucleus of the ciliated
protozoan Tetrahymena. Approximately 0.8% of the adenine residues in the macronuclear DNA of
Tetrahymena are methylated to N6-methyladenine. The degree of methylation varies between sites
from a very low percentage to >90%. In this study a correlation was found between nucleosome
positioning and DNA methylation. Eight GATC sites with different levels of methylation were
examined. There was a direct correlation between the degree of methylation and proximity to
linker DNA at these sites. Although methylation occurs preferentially in linker DNA, the
patterns and extent of methylation in a histone H1 knockout strain were virtually
indistinguishable from those in wild-type cells.

<>

<1>Karska-Wysocki, B., Hua, N.M., Vzdornov, D., Szatmari, G., Mamet-Bratley, M.D.
<2>Detection and purification of a new restriction endonuclease, LlaMI, from Lactococcus lactis subsp. lactis M19.
<3>Sixth Symposium on Lactic Acid Bacteria Genetics, Metabolism and Applications, Fed. European Microbiol. Socs. and Netherlands Soc. for Microbiology, none, Veldhoven, The Netherlands
<4>0
<5>F17
<6>1999
<7>One of the multiple phage defense systems present in bacteria is the Restriction/modification
system.  Type II restriction endonucleases recognize specific nucleotide sequences in invading
phage DNA and cleave the DNA at these sites.  We describe a new restriction activity, LlaMI,
present in cell extracts of Lactococcus lactis subsp. lactis M19, a variant isolated from a
commercially available industrial starter culture.  We have characterized the strain M19 by
molecular typing using AP-PCR fingerprinting and plasmid profile analysis.  During the
purification of LlaMI, non-specific nuclease activity was detected.  The nuclease activity
could be separated from the LlaMI activity using low-pressure anion exchange and affinity
chromatography.  Partially-purified cell extracts were tested for restriction activity on the
following substrates: DNA of phages lambda, PhiX174 and T7 and plasmids pBR322 and Litmus 29.
The LlaMI DNA cleavage patterns closely resemble those of restriction enzyme ScrFI, produced
by Lactococcus lactis subsp. cremoris UC503.

<>

<1>Karska-Wysocki, B., Szatmari, G., Barrette, B., Mamet-Bratley, M.D.
<2>Characterization of a new restriction endonuclease, LlaGI, produced  by Lactococcus lactis subsp. cremoris G2.
<3>Life Sc. Confer. Ottawa
<4>0
<5>P088
<6>1996
<7>A new sequence-specific endonuclease from Lactococcus lactis
subsp. cremoris G2 was identified, by restriction maping, as the first
reported isoschizomer of NheI.  We have confirmed this identification with
a cloning test and have named this new Type II restriction endonuclease
LlaGI.  We have characterized the enzyme-producing strain G2 by molecular
typing using AP-PCR fingerprinting and plasmid profile analysis.  These
tests demonstrated the presence of amplicons of 200 to 900 bp and the
presence of five plasmid bands at appropriate size positions of 7 to 25
kbp.  Extracts containing LlaGI nuclease did not digest DNA from the
producing strain, suggesting that a modification activity is also present
in this strain.  Specific endonuclease digestion of phage DNA was detected
in diluted crude extracts corresponding to 0.9 ug (wet-weight) G2
cells/ml.  During purification of LlaGI from crude extracts, non-specific
nuclease activity was detected in fractions containing the restriction
enzyme.  This contamination could be effectively separated from LlaGI
activity using low-pressure anion exchange chromatography and elution with
a gradient of NaCl or KCl.  We are currently developing a purification
protocol for potential commercial production of this new restriction
endonuclease.

<>

<1>Karska-Wysocki, B., Szatmari, G., Beauregard, G., Mamet-Bratley, M.D.
<2>A new restriction endonuclease activity found in Lactococcus  cremoris G2.
<3>Life Sc. Confer. Ottawa, , , Ottawa
<4>0
<5>D2-D9
<6>1995
<7>Only a few strains of Lactococcus species have been demonstrated
to have type II restriction endonuclease activity.  These enzymes recognize
specific nucleotide sequence and cleave DNA at these sites.  Here we
describe a novel restriction enzyme activity present in cell extracts of a
Lactococcus cremoris G2 strain isolated from a commercial mixed-starter
culture used in the manufacture of Cheddar cheese.  Cell crude extracts
were obtained by sonification of stationary phase cultures grown in M17, or
in whey medium.  The cell extracts were tested for DNA restriction activity
on lambda, PhiX174 and pBR322 DNA substrates.  Electrophoretic analysis of
restriction digests revealed the presence of only one cleavage site in
lambda and pBR322 DNA, and the absence of cleavage of PhiX174 DNA.  The
enzymatic activity of the extract was shown to be greater than 13,000 units
per gram (wet weight) of cells.  The electrophoretic patterns of this
restriction enzyme activity was compared with known restriction
enzymes.  This analysis clearly demonstrated that this new restriction
enzyme activity is identical to that of the restriction enzyme
NheI.  Indicating that the new restriction enzyme activity is the first
known isoschizomer of NheI.  The determination of the cleavage site of this
new enzyme is currently in progress.  We propose that this non-pathogenic
food-grade microorganism will be a better source for the NheI enzyme, which
is currently isolated from a Neisseria mucosa species originally isolated
from the human pharynx.

<>

<1>Kartashova, I.M., Nikolskaya, I.I., Debov, S.S.
<2>Separation of enzymes for modification methylases and restrictases from Shigella sonnei 47.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>4
<5>31-35
<6>1985
<7>Two systems for DNA host specificity have been demonstrated for Shigella sonnei cells, SsoI
and SsoII.  The aim of the present work was to separate the modifying methylases and
restriction endonucleases from Shigella sonnei and to study the modifying functions of
methylases M.SsoI and M.SsoII.  The possibilities to separate the methylation and restriction
enzymes by column chromatography on affinity, ion exchange and hydrophobic sorbents were
analyzed.  The scheme for separation of methylases and restriction endonucleases of Shigella
sonnei was elaborated, consisting of the fractioning of total preparation on phenylsepharose
and subsequent isoelectrofocusing on ampholines.  The modification functions of M.SsoI and
M.SsoII methylases obtained by this technique and devoid of concomitant restriction
endonucleases were studied.  The in vitro experiments have shown the acceptor DNA methylated
by M.SsoI or M.SsoII to be resistant to R.SsoI or R.SsoII.

<>

<1>Karuthedath, V.S., Vir, S.A., Kumar, S.P., Garg, P., Mohan, K.V., Katoch, K., Chauhan, D.S., Scaria, V., Sivasubbu, S.
<2>Draft Genome Sequence of an Extensively Drug-Resistant Mycobacterium tuberculosis Clinical Isolate of the Ural Strain OSDD493.
<3>Genome Announcements
<4>1
<5>e00928-13
<6>2013
<7>We describe the genome sequencing and analysis of a clinical isolate of Mycobacterium
tuberculosis belonging to the Ural strain OSDD493 from India.

<>

<1>Karuthedath-Vellarikkal, S., Patowary, A., Singh, M., Periwal, V., Singh, A.V., Singh, P.K., Garg, P., Mohan, K.V., Katoch, K., Jangir, P.K., Sharma, R., Chauhan, D.S., Scaria, V., Sivasubbu, S.
<2>Draft Genome Sequence of a Clinical Isolate of Multidrug-Resistant Mycobacterium  tuberculosis East African Indian Strain OSDD271.
<3>Genome Announcements
<4>1
<5>e00541-13
<6>2013
<7>We describe the genome sequencing and analysis of a clinical isolate of Mycobacterium
tuberculosis East African Indian (EAI) strain OSDD271 from India.

<>

<1>Karyagina, A., Shilov, I., Tashlitskii, V., Khodoun, M., Vasilev, S., Lau, P.C.K., Nikolskaya, I.
<2>Specific binding of SsoII DNA methyltransferase to its promoter region provides the regulation of SsoII restriction-modification gene expression.
<3>Nucleic Acids Res.
<4>25
<5>2114-2120
<6>1997
<7>The regulation of the SsoII restriction-modification system from Shigella sonnei was studied
in vivo and in vitro.  In lacZ fusion experiments, SsoII methyltransferase (M.SsoII) was found
to repress its own synthesis but stimulate expression of the cognate restriction endonuclease.
The N-terminal 72 amino acids of M.SsoII, predicted to form a helix-turn-helix motif, was
found to be responsible for the specific DNA-binding and regulatory function of M.SsoII.
Similar HTH motifs are predicted in the N-terminus of a number of 5-methylcytosine
methyltransferases, particularly M.EcoRII, M.dcm and M.MspI, of which the ability to regulate
autogenously has been proposed.  In vitro, the binding of M.SsoII to its target DNA was
investigated using a mobility shift assay.  M.SsoII forms a specific and stable complex with a
140 bp DNA fragment containing the promoter region of SsoII R-M system.  The dissociation
constant (Kd) was determined to be 1.5 x 10^-8 M.  DNaseI footprinting experiments
demonstrated that M.SsoII protects a 48-52 bp region immediately upstream of the M.SsoII
coding sequence which includes the predicted -10 promoter sequence of M.SsoII and the -10 and
-35 sequences of R.SsoII.

<>

<1>Karyagina, A.S.
<2>Genetic determinants for the enzymes of bacterial restriction-modification systems.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>7
<5>3-11
<6>1990
<7>Recent data on the molecular arrangement and functioning of the genetic
determinants for the enzymes of restriction-modification systems are discussed.
The problems of restriction endonuclease and methylase gene localization, the
regulation of the activity of the genes for Type II restriction-modification
systems, the characteristics of the primary structure of the genes and
phylogeny of restriction endonucleases and methylases are reviewed.

<>

<1>Karyagina, A.S., Levchenko, I.Y., Nikolskaya, I.I.
<2>A new method for isolation and purification of restriction endonuclease SsoII.
<3>Biokhimiia
<4>58
<5>908-912
<6>1993
<7>A new method for isolation and purification of restriction endonuclease SsoII which results in
a homogeneous preparation suitable for all types of fine physico-chemical assays has been
elaborated. The procedure includes four chromatographic steps: fractionation on
butyl-Toyopearl, combined chromatography on SP-Toyopearl and phosphocellulose PII, and
chromatography on DEAE-Toyopearl and on QAE-Toyopearl. The use of fast flow sorbents
(Toyopearl) makes it possible to reduce the time needed for the separation of proteins and to
optimize the fractionation conditions, thus avoiding the dialysis between the chromatographic
steps which significantly decreased the enzyme activity yields in previous purification
schemes. The isolation of restriction endonuclease SsoII by the new method usually takes four
days.

<>

<1>Karyagina, A.S., Lunin, V.G., Degtyarenko, K.N., Uvarov, V.Y., Nikolskaya, I.I.
<2>Analysis of the nucleotide and derived amino acid sequences of the SsoII restriction endonuclease and methyltransferase.
<3>Gene
<4>124
<5>13-19
<6>1993
<7>A 2648-bp fragment from the P4 plasmid of Shigella sonnei strain 47 coding for the SsoII
restriction endonuclease (ENase) and methyltransferase (MTase) (recognition sequence
5'-CCNGG) was sequenced. Two divergently arranged open reading frames of 905 bp for the SsoII
ENase (R.SsoII) and 1137 bp for the MTase (M.SsoII were identified. The coding regions are
separated by 110 bp. The calculated Mr of R.SsoII (35937) and M.SsoII (42887) are in good
agreement with values previously obtained by in vitro transcription-translation experiments,
i.e., 35 and 43 kDa for the ENase and MTase, respectively. The M.SsoII amino acid (aa)
sequence revealed a considerable similarity to m5C-MTases recognizing the related sequences -
M.EcoRII, M.dcm, M.MspI, M.BsuFI, M.HpaII, and M.HhaI. Surprisingly, the greatest degree of
homology has been observed between the aa sequences of M.SsoII and M.NlaX, with an
unidentified recognition sequence. The multiple alignment of aa sequences helps to identify
the blocks of conserved aa in variable regions of MTases. These conserved aa can play a key
role in target recognition. Some aspects of evolution of m5C-MTases are discussed.

<>

<1>Karyagina, A.S., Lunin, V.G., Gruber, I.M., Polyachenko, V.M., Nikolskaya, I.I., Debov, S.S.
<2>Activity of the restriction endonuclease SsoII in Escherichia coli cells transformed by Shigella sonnei 47 plasmids.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>12
<5>26-29
<6>1987
<7>The inverse dependence of activity of restriction endonuclease SsoII
preparations on the number of low molecular mass plasmids of Shigella sonnei
transforming Escherichia coli recipient cells producing the enzyme has been
shown.  An Escherichia coli strain efficiently producing one of two Shigella
sonnei 47 restriction endonucleases, SsoII, has been isolated.  The producer
strain harbours two of the nine Shigella sonnei 47 plasmids.  One of them, P4,
codes for the SsoII+ phenotype while another, P9, determines the plasmids
conjugation transfer.  Biochemical and physiological characteristics of the
producer strain XS13 are identical to the ones of the recipient Escherichia
coli strain PS200.  XS13 is unable to induce keratoconjunctivitis in guinea
pigs in pathogenicity test.

<>

<1>Karyagina, A.S., Lunin, V.G., Levtchenko, I.Y., Labbe, D., Brousseau, R., Lau, P.C.K., Nikolskaya, I.I.
<2>The SsoII and NlaX DNA methyltransferases: overproduction and functional analysis.
<3>Gene
<4>157
<5>93-96
<6>1995
<7>Overproduction of the NlaX DNA methyltransferase (M.NlaX) in an Escherichia coli host
conferred resistance to SsoII restriction endonuclease (R.SsoII) digestion.  This suggested an
overlap of sequence specificity between M.NlaX and M.SsoII, the latter of which modifies the
internal cytosine of the target sequence 5'-CCNGG-3'.  A variant of M.NlaX (M.Sso/Nla),
containing an N-terminal extension from M.SsoII, was also enzymatically active.  Using
deletion analysis, the N-terminal 71 amino-acid residues of M.SsoII were shown to be essential
for modification activity.

<>

<1>Karyagina, A.S., Lunin, V.G., Nikolskaia, I.I., Debov, S.S.
<2>Functional characteristics of plasmids of Shigella sonnei 47 strains.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>10
<5>16-21
<6>1986
<7>The phenotypic characteristics of Shigella sonnei strain 47 containing 7 plasmids of low
molecular weight and 2 plasmids 60-100 Md large have been studied.  The strains of Escherichia
coli containing the single plasmids or plasmid groups from Shigella sonnei have been obtained
by transformation and conjugation.  The comparison of phenotypes of the strains obtained has
helped to find the plasmid location of the determinants for streptomycin resistance (P7),
genes for colicinogenicity abd colicin immunity (P5), the enzymes of host cell specificty
system Sso47I (P6), Sso47II (P4) and the genes for the conjugative DNA transfer (P9).
Escherichia coli strains producing individual restriction enzymes SsoI and SsoII have been
isolated.

<>

<1>Karyagina, A.S., Lunin, V.G., Nikolskaya, I.I.
<2>Characterization of the genetic determinants of SsoII-restriction endonuclease and modification methyltransferase.
<3>Gene
<4>87
<5>113-118
<6>1990
<7>The genes encoding SsoI and SsoII restriction endonuclease (ENase) and
methyltransferase (MTase) are located on the small plasmids P6 and P4,
respectively, of Shigella sonnei strain 47.  Functions provided by plasmids P5,
P7 and P9, which include colicinogenicity and immunity to colicin E1,
resistance to streptomycin (Sm), and conjugative DNA transfer, respectively,
have also been identified.  The genes of the SsoII restriction-modification
(R-M) system have been cloned into Escherichia coli expressing the
35-kDa(ENase) and 43-kDa(MTase) products.  A restriction map of the P4 plasmid
DNA was determined, and the approximate location of the genes encoding SsoII
ENase and MTase (ssoIIR and ssoIIM) on that have been established.  SsoI is an
isoschisomer of EcoRI and SsoII cleaves the recognition sequence 5'-CCNGG
producing 5'-protruding 5-nt long cohesive ends.

<>

<1>Karyagina, A.S., Lunin, V.G., Nikolskaya, I.I., Debov, S.S.
<2>Localization of SsoII restriction endonuclease and methylase genes on the physical map of plasmid P4.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>3
<5>16-20
<6>1989
<7>A restriction map of naturally occuring plasmid P4 from Shigella sonnei 47
strain coding for the SsoII restriction endonuclease and methylase genes has
been made.  Using the genetic engineering approach the locations of the SsoII
host cell specificity system enzyme genes have been determined.

<>

<1>Kas, E., Poljak, L., Adachi, Y., Laemmli, K.
<2>A model for chromatin opening: stimulation of topoisomerase II and restriction enzyme cleavage of chromatin by distamycin.
<3>EMBO J.
<4>12
<5>115-126
<6>1993
<7>Histone H1 preferentially and cooperatively binds scaffold-associated regions (SARS) in vitro
via specific interactions with the numerous short A+T-rich tracts (A-tracts) contained in
these sequences. Selective titration of A-tracts by the oligopeptide distamycin abolishes this
interaction and results in a redistribution of H1. Similarly, treatment of intact cells and
isolated nuclei with distamycin specifically enhances cleavage of internucleosomal linkers of
SARs by topoisomerase II and restriction enzymes. The increased accessibility of these linkers
is thought to result from the unfolding (or opening) of the chromatin fiber and to be due to a
reduced occupancy by histone H1. Chromatin extraction and H1 assembly experiments support this
view. We discuss a model whereby open, H1-depleted chromatin regions may be generated by
titration of A-tracts by putative distamycin analogues; this local opening may spread to
adjacent regions assuming highly cooperative H1-H1 interactions in chromatin.

<>

<1>Kasahara, Y., Nakai, S., Ogasawara, N., Yata, K., Sadaie, Y.
<2>Sequence analysis of the groESL-cotA region of the Bacillus subtilis genome, containing the restriction/modification system genes.
<3>DNA Res.
<4>4
<5>335-339
<6>1997
<7>We have determined a 35-kb sequence of the groESL-gutR-cotA (45o-52o) region of the Bacillus
subtilis genome.  In addition to the groESL, gutRB and cotA genes reported previously, we have
newly identified 24 ORFs including gutA and fruC genes, encoding glucitol permease and
fructokinase, respectively.  The inherent restriction/modification system genes, hsdMR and
hsdMM, were mapped between groESL and gutRB, and we have identified two open reading frames
(ORFs) encoding 5-methylcytosine forming DNA methyltransferase and an operon probably encoding
a restriction enzyme complex.  The unusual genome structure of few ORFs and lower GC content
around the restriction/modification genes strongly suggests that the region originated from a
bacteriophage integrated during evolution.

<>

<1>Kasarjian, J., Kawai, S., Ryu, J.
<2>Use of plasmid transformation to find new restriction enzymes.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>100
<5>371
<6>2000
<7>Recent genome projects have revealed many restriction enzymes and their corresponding
methylases, specifically of type I and III.  Traditionally, restriction enzymes have been
discovered by classical restriction and modification phenomenon of bacteriophages (type I, II,
and III) or direct enzyme assay (most type II enzymes).  Once genes are cloned, DNA
hybridization is also used to discover other enzymes with similar DNA sequences.  To avoid the
traditional limitation of using bacteriophages or biochemical assays, we have designed a
feasibility study using plasmid transformation to detect the restriction activities of the
cell.  Plasmids were made by subcloning six BamHI fragments from bacteriophage lambda DNA into
a pUC derivative plasmid, pMECA.  These plasmids were transformed into E. coli strains
representing each type of restriction system (type I: EcoKI, EcoAI, Eco124I; type II: HindIII;
type III: EcoPI).  As a control, plasmids were also transformed into E. coli C, which does not
contain a restriction system and efficiency of transformation values were obtained by
comparison.  Reduction of relative EOT values (10^-1 - 10^-3) were observed in each BamHI
subclone containing the corresponding recognition sites.  These results suggest that the
presence of restriction enzymes can be predicted using this plasmid transformation method as
an alternative to phage experiments or biochemical studies.  Accumulation of DNA sequence data
also makes it possible to predict the recognition sequence of a restriction enzyme with the
use of computer analysis.

<>

<1>Kasarjian, J., Valinluck, V., Ryu, J.
<2>Determination of the methylation sites of the type I restriction enzymes, KpnAI and KpnBI of Klebsiella species.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>102
<5>236
<6>2002
<7>Type I restriction endonucleases have been identified widely in bacteria since the completion
of the first genome sequence project of
Haemophilus influenzae in 1995. The enzymes recognize non-palindromic
bipartite DNA sequences made up of a 3-4 base pair (bp) 5' region, a
6-8 bp nonspecific spacer, and a 4-5 bp 3' region. The corresponding
adenine N6-methyltransferase modifies a specific adenine in each strand
within the recognition sequence to protect DNA from cleavage.
Traditionally, type I enzymes in enteric bacteria are classified into
distinguished families: IA, IB, IC and ID. Two type I restriction and
modification systems, KpnAI (type ID) and KpnBI (new family), have been
identified in Klebsiella oxytoca M5a1 (originally classified as K.
pneumoniae), and Klebsiella pneumoniae GM236, respectively. We have
previously developed a simple transformation method to identify their
recognition sequence. Using a series of plasmids with known DNA
sequence and a computer program, the recognition sequence of KpnAI and
KpnBI were determined to be GAA(N6)TGCC and CAAA(N6) RTCA,
respectively. In this study the sensitivity of the type II restriction
endonuclease, HindIII, to site-specific modification at 6-methyladenine
is used to determine the methylation sites of KpnAI and KpnBI. We
designed several synthetic DNA oligonucleotides containing a HindIII
recognition site overlapping with either a KpnAI or KpnBI site. The
oligonucleotide was then cloned into plasmid pMECA and transferred into
a KpnAI or KpnBI containing strain, M5a1 or GM236, respectively for
modification. Methylated plasmids were subjected to HindIII digestion.
When the methylated adenine (A*) of the synthetic recognition sequence
overlapped with the methylated adenine in the HindIII sequence
(A*AGCTT), the plasmids were protected from cleavage HindIII. The
methylation of the opposite strand KpnBI recognition sequence was
determined using the in vivo activity of dam+ and dam- strains.

<>

<1>Kasarjian, J.K.A., Burnett, A.S., Ryu, J.
<2>Use of plasmid transformation to find new restriction enzymes and their recognition sequences.
<3>J. Invest. Med.
<4>49
<5>12A
<6>2001
<7>Restriction-modification systems are widespread in bacteria and consist of both a restriction
endonuclease and a corresponding methylase.  Bacterial genome sequencing projects, however,
suggest that many restriction enzymes have yet to be discovered.  Restriction enzymes are
classified into three types (I, II, III) and type II restriction enzymes are useful in genetic
engineering.  Traditionally, restriction enzymes have been discovered by classical restriction
and modification tests, requiring bacteriophages or direct enzyme assay.  DNA hybridization
can find new homologous sequences, but unique DNA sequences will be overlooked and the high
cost of bacterial genome projects may limit its usefulness.  We have established a new
quantitative R-M test based on plasmid transformation efficiency using DNA fragments derived
from E. coli phage lambda.  This test is similar to traditional "efficiency of plating" assays
but measures "efficiency of transformation".  Plasmids were made by subcloning six BamHI
fragments from lambda into pMECA, a pUC derivative plasmid.  These six plasmids were each
transformed into bacterial strains representing each type of restriction system (type I;
EcoAI, EcoRI, Eco124I; type II: HindIII; type III: EcoPI).  Reduction of relative EOT values
(10-1-10-3) were observed in each BamHI subclone containing the corresponding recognition
sites.  These results confirm the effectiveness of this new method.  This method was also used
to predict the recognition sequence of KpnAI, a type I R-M system from Klebsiella pneumoniae,
M5a1.  A total of 29 lambda subclones were constructed and transformed into M5a1 to determine
the presence (+) or absence (-) of a recognition site.  DNA sequence information was then
compared using the newly developed computer program, Seth Analyzer.  Transformation of a
synthetic oligonucleotide containing the predicted recognition sequence confirmed the
recognition sequence of KpnAI to be GAA(6N)TGCC.  Because plasmid transformation methods are
available for many bacteria, this model system can be extended to many bacterial species to
search for new restriction enzymes and their recognition sequences.

<>

<1>Kasarjian, J.K.A., Burnett, A.S., Ryu, J.
<2>Identification of the recognition sequence of KpnAI, a type ID restriction enzyme, in Klebsiella pneumoniae M5a1.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>403
<6>2001
<7>Type II restriction enzymes have been studied extensively due to their usefulness in genetic
engineering, while our knowledge of type I and
III systems remains limited. Recent bacterial genome sequencing
projects suggest an abundance of all three types of
restriction-modification (R-M) systems. Finding new type I and III
restriction systems and identifying their recognition sites is
difficult, leaving many recognition sites still unknown. KpnAI, an R-M
system from Klebsiella pneumoniae, has previously been cloned and
sequenced in our laboratory. Analysis of both DNA and protein sequences
identified this system as a new member of the type ID family. Here we
have determined the recognition sequence for KpnAI using a simple in
vivo plasmid transformation method, specifically developed in our lab,
which can also be used as a tool to identify new R-M systems. Two sets
of plasmids were made by subcloning DNA fragments from bacteriophage
lambda and Escherichia coli. Each set of 30 plasmids was then
transformed into M5a1 using the restriction-minus mutant, M5a1R as a
control. Only when the plasmid contained a restriction site, a
reduction (10-2) in transformation frequency was observed when compared
to the control. These results identified the presence of a recognition
site in 24 plasmids and absence of a site in 32 plasmids. A computer
program was developed to search for a common sequence that exists in
all positive plasmids but is not found in any negative plasmids. One
unique recognition sequence was identified and a 19 base synthetic
oligonucleotide containing this sequence was cloned into pMECA.
Transformation into strain M5a1 resulted in a significant reduction in
transformation frequency confirming the recognition sequence of KpnAI
to be GAA(6N)TGCC. This model system can be extended to many bacterial
species to search for new restriction enzymes and their recognition
sequences.

<>

<1>Kasarjian, J.K.A., Hidaka, M., Horiuchi, T., Iida, M., Ryu, J.
<2>The recognition and modification sites for the bacterial type I restriction systems KpnAI, StySEAI, StySENI and StySGI.
<3>Nucleic Acids Res.
<4>32
<5>e82
<6>2004
<7>Using an in vivo plasmid transformation method, we have determined the DNA sequences
recognized by the KpnAI, StySEAI, StySENI and StySGI R-M systems from Klebsiella oxytoca
strain M5a1, Salmonella eastbourne, Salmonella enteritidis and Salmonella gelsenkirchen,
respectively. These type I restriction-modification systems were originally identified using
traditional phage assay, and described here is the plasmid transformation test and computer
program used to determine their DNA recognition sequences. For this test, we constructed two
sets of plasmids, pL and pE, that contain phage lambda and Escherichia coli K-12 chromosomal
DNA fragments, respectively. Further, using the methylation sensitivities of various known
type II restriction enzymes, we identified the target adenines for methylation (listed in bold
italics below as A or T in case of the complementary strand). The recognition sequence and
methylation sites are GAA(6N)TGCC (KpnAI), ACA(6N)TYCA (StySEAI), CGA(6N)TACC (StySENI) and
TAAC(7N)RTCG (StySGI). These DNA recognition sequences all have a typical type I bipartite
pattern and represent three novel specificities and one isoschizomer (StySENI). For
confirmation, oligonucleotides containing each of the predicted sequences were synthesized,
cloned into plasmid pMECA and transformed into each strain, resulting in a large reduction in
efficiency of transformation (EOT).

<>

<1>Kasarjian, J.K.A., Iida, M., Ryu, J.
<2>New restriction enzymes discovered from Escherichia coli clinical strains using a plasmid transformation method.
<3>Nucleic Acids Res.
<4>31
<5>e22
<6>2003
<7>The presence of restriction enzymes in bacterial cells has been predicted by either classical
phage restriction-modification (R-M) tests, direct in
vitro enzyme assays or more recently from bacterial genome sequence
analysis. We have applied phage R-M test principles to the transformation
of plasmid DNA and established a plasmid R-M test. To validate this test,
six plasmids that contain BamHI fragments of phage lambda DNA were
constructed and transformed into Escherichia coli strains containing known
R-M systems including: type I (EcoBI, EcoAI, Eco124I), type II (HindIII)
and type III (EcoP1I). Plasmid DNA with a single recognition site showed a
reduction of relative efficiency of transformation (EOT = 10(-1)-10(-2)).
When multiple recognition sites were present, greater reductions in EOT
values were observed. Once established in the cell, the plasmids were
subjected to modification (EOT = 1.0). We applied this test to screen
E.coli clinical strains and detected the presence of restriction enzymes
in 93% (14/15) of cells. Using additional subclones and the computer
program, RM Search, we identified four new restriction enzymes, Eco377I,
Eco585I, Eco646I and Eco777I, along with their recognition sequences,
GGA(8N)ATGC, GCC(6N)TGCG, CCA(7N)CTTC, and GGA(6N)TATC, respectively.
Eco1158I, an isoschizomer of EcoBI, was also found in this study.

<>

<1>Kasarjian, J.K.A., Kodama, Y., Iida, M., Matsuda, K., Ryu, J.
<2>Four new type I restriction enzymes identified in Escherichia coli clinical isolates.
<3>Nucleic Acids Res.
<4>33
<5>e114
<6>2005
<7>Using a plasmid transformation method and the RM search computer program, four type I
restriction enzymes with new recognition sites and two isoschizomers (EcoBI and Eco377I) were
identified in a collection of clinical Escherichia coli isolates. These new enzymes were
designated Eco394I, Eco826I, Eco851I and Eco912I. Their recognition sequences were determined
to be GAC(5N)RTAAY, GCA(6N)CTGA, GTCA(6N)TGAY and CAC(5N)TGGC, respectively. A methylation
sensitivity assay, using various synthetic oligonucleotides, was used to identify the adenines
that prevent cleavage when methylated (underlined). These results suggest that type I enzymes
are abundant in E.coli and many other bacteria, as has been inferred from bacterial genome
sequencing projects.

<>

<1>Kaser, M.R.
<2>Genes expressed in treated human C3A liver cell cultures.
<3>US Patent Office
<4>US 6727066 A
<5>
<6>2004
<7>The present invention relates to a composition comprising a plurality of cDNAs which are
differentially expressed in treated human C3A liver cell cultures and which may be used
entirely or in part to diagnose, to stage, to treat, or to monitor the progression or
treatment of liver disorders such as hyperlipidemia.

<>

<1>Kashyap, R.S., Bhullar, S.S., More, R.P., Puranik, S., Purohit, H.J., Taori, G.M., Daginawala, H.F.
<2>Genome Sequence of Mycobacterium tuberculosis C2, a Cerebrospinal Fluid Clinical  Isolate from Central India.
<3>Genome Announcements
<4>2
<5>e00842-14
<6>2014
<7>We report the annotated genome sequence of a Mycobacterium tuberculosis clinical  isolate from
the cerebrospinal fluid of a tuberculous meningitis patient admitted
to the Central India Institute of Medical Sciences, Nagpur, India.

<>

<1>Kaszubska, W.
<2>Studies of a DNA modification methyltransferase from Rhodobacter sphaeroides.
<3>Diss. Abstr.
<4>52
<5>1407B
<6>1991
<7>The DNA methyltransferases represent an interesting class of enzymes for the
study of protein-DNA interactions due to their high specificity, structural
simplicity, and biological importance.  RsrI methyltransferase (M.RsrI) from
Rhodobacter sphaeroides recognizes duplex d(GAATTC) and deposits methyl groups
at the N6 position of the central adenine.  M.RsrI was purified to homogeneity
from R. sphaeroides, and its gene cloned and sequenced.  The purification used
four chromatography columns, and yielded up to 100 micrograms of enzyme.
M.RsrI was overexpressed in E. coli and the yield of the purification improved
by about 100-fold with respect to that from R. sphaeroides.  Several physical
and biochemical properties of the RsrI methyltransferase were determined:
molecular weight under denaturing and nondenaturing conditions, isoelectric
point, optimal reaction conditions, the mode of methyl group transfer, and the
enzyme-DNA binding.  M.RsrI was compared to a functionally identical enzyme
from E. coli, EcoRI methyltransferase (M.EcoRI).  M.RsrI and M.EcoRI differ in
their physical properties, including a striking lack of similarity in their
deduced amino acid sequences.  The two methyltransferases might recognize the
cognate DNA sequence differently, because they do not bind to DNA with equal
efficiencies under the same conditions.  However, M.RsrI and M.EcoRI share
identical catalytic properties.  Both transfer one methyl group to the
recognition sequence per binding event.  M.RsrI and M.EcoRI represent an
opportunity to elucidate the mode of action of two structurally different but
functionally identical enzymes.  It is possible that these enzymes retain
functional similarity by having essentially the same three dimensional
configuration.  Alternatively, they might have dissimilar structures as well as
mechanistic differences.

<>

<1>Kaszubska, W., Aiken, C., O'Connor, C.D., Gumport, R.I.
<2>Purification, cloning and sequence analysis of RsrI DNA methyltransferase: lack of homology between two enzymes, RsrI and EcoRI, that methylate the same nucleotide in identical recognition sequences.
<3>Nucleic Acids Res.
<4>17
<5>10403-10425
<6>1989
<7>RsrI DNA methyltransferase (M.RsrI) from Rhodobacter sphaeroides has been
purified to homogeneity, and its gene cloned and sequenced.  This enzyme
catalyzes methylation of the same central adenine residue in the duplex
recognition sequence d(GAATTC) as does M.EcoRI.  The reduced and denatured
molecular weight of the RsrI methyltransferase (MTase) is 33,600 Da.  A
fragment of R. sphaeroides chromosomal DNA exhibited M.RsrI activity in E. coli
and was used to sequence the rsrIM gene.  The deduced amino acid sequence of
M.RsrI shows partial homology to those of the type II adenine MTases HinfI and
DpnA and N4-cytosine MTases BamHI and PvuII, and to the type III adenine MTases
EcoP1 and EcoP15.  In contrast to their corresponding isoschizomeric
endonucleases, the deduced amino acid sequences of the RsrI and EcoRI MTases
show very little homology.  Either the EcoRI and RsrI restriction modification
systems assembled independently from closely related endonuclease and more
distantly related MTase genes, or the MTase genes diverged more than their
partner endonuclease genes.  The rsrIM gene sequence has also been determined
by Stephenson and Greene (Nucl. Acids Res. (1989) 17, this issue).

<>

<1>Kaszubska, W., Aiken, C.R., Gumport, R.I.
<2>RsrI restriction-modification enzymes from Rhodobacter sphaeroides.
<3>Gene
<4>74
<5>83-84
<6>1988
<7>Meeting Abstract

<>

<1>Kaszubska, W., Webb, H.K., Gumport, R.
<2>Purification and characterization of the M.RsrI DNA methyltransferase from Escherichia coli.
<3>Gene
<4>118
<5>5-11
<6>1992
<7>The gene (rsrIM) encoding the RsrI DNA methyltransferase (M.RsrI) from Rhodobacter sphaeroides
was cloned and expressed in Escherichia coli. Under the control of a bacteriophage T7
promotor, 2% of the total protein in a crude extract was M.RsrI. This level of expression
represents an approximately 50-fold increase over that present in the natural host.
Chromatography using DNA cellulose and the S-adenosylmethionine analogue, sinefungin, was
useful in purifying the enzyme to homogeneity. The purification yielded 100 times more enzyme
than was obtained from the same quantity of R.sphaeroides cell paste. M.RsrI deposits one
methyl group per productive DNA-binding event, as does its functional but
sequence-nonhomologous analogue, M.EcoRI. Unlike M.EcoRI, the R. sphaeroides enzyme is a dimer
at micromolar concentrations.

<>

<1>Kataeva, I.A., Yang, S.J., Dam, P., Poole, F.L.I.I., Yin, Y., Zhou, F., Chou, W.C., Xu, Y., Goodwin, L., Sims, D.R., Detter, J.C., Hauser, L.G., Westpheling, J., Adams, M.W.
<2>Genome Sequence of the Anaerobic, Thermophilic and Cellulolytic Bacterium Anaerocellum thermophilum DSM 6725.
<3>J. Bacteriol.
<4>191
<5>3760-3761
<6>2009
<7>Anaerocellum thermophilum DSM 6725 is a strictly anaerobic bacterium that grows optimally at
75 degrees C. It uses a variety of polysaccharides, including crystalline cellulose and
untreated plant biomass, and has potential utility in biomass conversion. Here we report its
complete genome sequence of 2.97 Mb, which is contained within one chromosome and two plasmids
(of 8.3 and 3.6 kb). The genome encodes a broad set of cellulolytic enzymes, transporters and
pathways for sugar utilization and compared to those of other saccharolytic, anaerobic
thermophiles is most similar to that of Caldicellulosiruptor saccharolyticus DSM 8903.

<>

<1>Katano, Y., Fujinami, S., Kawakoshi, A., Nakazawa, H., Oji, S., Iino, T., Oguchi, A., Ankai, A., Fukui, S., Terui, Y., Kamata, S., Harada, T., Tanikawa, S., Suzuki, K., Fujita, N.
<2>Complete genome sequence of Oscillibacter valericigenes Sjm18-20(T) (=NBRC 101213(T)).
<3>Standards in Genomic Sciences
<4>6
<5>406-414
<6>2012
<7>Oscillibacter valericigenes is a mesophilic, strictly anaerobic bacterium belonging to the
clostridial cluster IV. Strain Sjm18-20(T) (=NBRC 101213(T) =DSM
18026(T)) is the type strain of the species and represents the genus
Oscillibacter Iino et al. 2007. It was isolated from the alimentary canal of a
Japanese corbicula clam (Corbicula japonica) collected on a seacoast in Shimane
Prefecture in Japan. Phylogenetically, strain Sjm18-20(T) is closest to
uncultured bacteria in digestive tracts, including the enriched cells thought to
represent Oscillospira guilliermondii Chatton and Perard 1913. The isolated
phylogenetic position and some distinct characteristics prompted us to determine
the complete genome sequence. The 4,410,036 bp chromosome and the 60,586 bp
plasmid were predicted to encode a total of 4,723 protein-coding genes.

<>

<1>Katiliene, Z., Katilius, E., Woodbury, N.W.
<2>Single molecule detection of DNA looping by NgoMIV restriction endonuclease.
<3>Biophys. J.
<4>84
<5>4053-4061
<6>2003
<7>Single molecule fluorescence resonance energy transfer (FRET) and fluorescence correlation
spectroscopy were used to investigate DNA looping
by NgoMIV restriction endonuclease. Using a linear double-stranded DNA
(dsDNA) molecule labeled with a fluorescence donor molecule, Cy3, and
fluorescence acceptor molecule, Cy5, and by varying the concentration of
NgoMIV endonuclease from 0 to 3 x 10(-6) M, it was possible to detect and
determine diffusion properties of looped DNA/protein complexes. FRET
efficiency distributions revealed a subpopulation of complexes with an
energy transfer efficiency of 30%, which appeared upon addition of enzyme
in the picomolar to nanomolar concentration range (using 10(-11) M dsDNA).
The concentration dependence, fluorescence burst size analysis, and
fluorescence correlation analysis were all consistent with this
subpopulation arising from a sequence specific interaction between an
individual enzyme and a DNA molecule. A 30% FRET efficiency corresponds to
a distance of approximately 65 A, which correlates well with the distance
between the ends of the dsDNA molecule when bound to NgoMIV according to
the crystal structure of this complex. Formation of the looped complexes
was also evident in measurements of the diffusion times of freely
diffusing DNA molecules with and without NgoMIV. At very high protein
concentrations compared to the DNA concentration, FRET and fluorescence
correlation spectroscopy results revealed the formation of larger
DNA/protein complexes.

<>

<1>Katna, A., Boratynski, R., Furmanek-Blaszk, B., Zolcinska, N., Sektas, M.
<2>Unbalanced Restriction Impairs SOS-induced DNA Repair Effects.
<3>J. Microbiol. Biotechnol.
<4>20
<5>30-38
<6>2010
<7>The contribution of a type II restriction-modification system (R-M system) to genome integrity
and cell viability was investigated. We
established experimental conditions that enabled the achievement of
hemimethylated and unmethylated states for the specific bases of the
recognition sequences of the host's DNA. To achieve this, we
constructed the MboII R-M system containing only one (i.e., M2.MboII)
out of two functional MboII methyltransferases found in Moraxella
bovis. Using the incomplete R-M system, we were able to perturb the
balance between methylation and restriction in an inducible manner. We
demonstrate that upon the SOS-induced DNA repair in mitomycin C treated
cells, restriction significantly reduces cell viability. Similar
results for the well-studied wild-type EcoRI R-M system, expressed
constitutively in Escherichia coli, were obtained. Our data provide
further insights into the benefits and disadvantages of maintaining of
a type II R-M system, highlighting its impact on host cell fitness.

<>

<1>Kato, F.
<2>Isolation and characterization of restriction endonuclease from amino acid- or nucleic acid-producing bacteria.
<3>Dojin News
<4>51
<5>3-9
<6>1989
<7>Restriction enzymes are endonucleases that recognize specific nucleotide
sequences in double stranded DNA and cleave both strands of the duplex.  In the
cell of origin each restriction enzyme is part of a restriction-modification
system, consisting of the restriction endonuclease and a matched modification
enzyme which recognizes and modifies the same nucleotide sequence in the DNA
strand.  Modification thus protects cellular DNA from restriction, however,
foreign DNA cleaved by the restriction endonuclease and further degraded by
other enzymes.  Such restriction modification systems, first detected by phage
restriction and modification, are widespread in bacteria and are thought to
play a role in eliminating foreign DNA that gains entrance to the cell via
viruses or as naked DNA.  The discovery of the first Class II endonuclease was
reported by Smith and Wilcox (1970).  Since that time, the number of Class II
restriction endonucleases available in purified form has been increasing at a
remarkable pace, and it is now very often possible to find one or several
restriction endonucleases having the specificity one desires.  This review is
mainly concerned with general properties and applications of Class II
restriction endonucleases including some properties of the restriction
endonucleases isolated from amino acid- or nucleic acid-producing bacteria and
related species in our laboratory.

<>

<1>Kato, F., Suetake, T., Murata, A., Mukai, T., Ueki, H.
<2>Restriction endonuclease MroI useful for genetic engineering - obtained by culturing Micrococcus roseus S.
<3>Japanese Patent Office
<4>JP 63098383
<5>
<6>1988
<7>A restriction endonuclease MroI having the following physico-chemical properties is new ;- (a)
activity and substrate specificity: Recognises base sequence (I) in a double-strand DNA and
cleaves it at the position indicated by the arrows. (where A means adenosine, G means
guanosine, T means thymidine and C means cytidine) (B) Optimum pH: 6.5-8.5. (c) Stable pH:
6.0-9.0. (d) Optimum temperature: 25-35 deg.C. Specifically, the enzyme is produced by
Micrococcus roseus S. The strain is deposited as FERM-P-8996. The strain is cultured as usual.
At log phase or early stationary phase, cells are collected and are disrupted. The extract is
subjected to precipitation with (NH4)2SO4, dialysis, ion-exchange, affinity chromatography,
gel filtration, etc.. Preferred salt concentration. is up to 300 mM for NaCl and KCl. Required
metal ion is about 10mM MgCl2 (inhibited at higher than 20 mM). MnCl can also activate the
enzyme, but this effect is lower than that of Mg (2+).

<>

<1>Kato, H., Shiwa, Y., Oshima, K., Machii, M., Araya-Kojima, T., Zendo, T., Shimizu-Kadota, M., Hattori, M., Sonomoto, K., Yoshikawa, H.
<2>Complete Genome Sequence of Lactococcus lactis IO-1, a Lactic Acid Bacterium That Utilizes Xylose and Produces High Levels of L-Lactic Acid.
<3>J. Bacteriol.
<4>194
<5>2102-2103
<6>2012
<7>We report the complete genome sequence of Lactococcus lactis IO-1 (= JCM7638). It is a
nondairy lactic acid bacterium, produces nisin Z, ferments xylose, and
produces predominantly l-lactic acid at high xylose concentrations. From ortholog
analysis with other five L. lactis strains, IO-1 was identified as L. lactis
subsp. lactis.

<>

<1>Kato, J., Endo, T., Kawamura, H., Nakashima, Y., Furukoshi, K., Oishi, K.
<2>Inhibition of restriction endonucleases by hot water extracts of spices.
<3>Bull. Coll. Agr. Vet. Med.
<4>47
<5>84-87
<6>1990
<7>Forty samples of spices were investigated for their inhibitory activities to
the cleavage of lambda DNA by restriction endonucleases.  Bayberry, clove
eucalyptus, melissa, nutmeg, oregano, peppermint and savory showed complete
inhibition to HindIII, under the condition employed.  Hot water extract of
clove showed marked inhibitory activity (MIC 2 micrograms/ml) and that of
oregano and peppermint showed potent inhibition (MIC 20 micrograms/ml).  MIC of
clove extract for five restriction endonucleases were determined.  MIC for
EcoRI and HindIII was 0.8 micrograms/ml, and 12.5 micrograms/ml for BamHI, 50
micrograms/ml for BglII and PstI.  Specificity of inhibition for restriction
endonucleases by hot water extract of clove was found.

<>

<1>Kato, J., Ooishi, K., Irie, S., Murata, K.
<2>Marine algae polysaccharide gel for restriction enzyme purification.
<3>Japanese Patent Office
<4>JP 398582
<5>
<6>1991
<7>
<>

<1>Kato, K., Toh, H., Sakamoto, N., Mori, K., Tashiro, K., Hibi, N., Sonomoto, K., Nakayama, J.
<2>Draft Genome Sequence of Lactobacillus namurensis Chizuka 01, Isolated from Nukadoko, a Pickling Bed of Fermented Rice Bran.
<3>Genome Announcements
<4>2
<5>e01263-13
<6>2014
<7>Lactobacillus namurensis Chizuka 01 was isolated from nukadoko, which is a fermented rice bran
bed traditionally used in Japan for pickling vegetables.
Here, we report the first draft of an annotated genome sequence of this organism.
This paper is the first published report of the genomic sequence of L.
namurensis.

<>

<1>Katoh, H., Miyata, S., Inoue, H., Iwanami, T.
<2>Unique features of a Japanese 'Candidatus Liberibacter asiaticus' strain revealed by whole genome sequencing.
<3>PLoS ONE
<4>9
<5>e106109
<6>2014
<7>Citrus greening (huanglongbing) is the most destructive disease of citrus worldwide.  It is
spread by citrus psyllids and is associated with phloem-limited bacteria of three species of
a-Proteobacteria, namely, 'Candidatus Liberibacter asiaticus', 'Ca.  L. americanus', and
'Ca. L. africanus'.  Recent findings suggested that some Japanese strains lack the
bacteriophage-type DNA polymerase region (DNA pol), in contrast to the Floridian psy62 strain.
The whole genome sequence of the pol-negative 'Ca. L. asiaticus' Japanese isolate lshi-1 was
determined by metagenomic analysis of DNA extracted from 'Ca. L. asiaticus'-infected
psyllids and leaf midribs.  The 1.19-Mb genome has an average 36.32% GC content.  Annotation
revealed 13 operons encoding rRNA and 44 tRNA genes, but no typical bacterial
pathogenesis-related genes were located within the genome, similar to the Floridian psy62 and
Chinese gxpsy.  In contrast to other 'Ca. L. asiaticus' strains, the genome of the Japanese
Ishi-1 strain lacks a prophage-related region.

<>

<1>Katsura, S., Harada, N., Maeda, Y., Komatsu, J., Matsuura, S., Takashima, K., Mizuno, A.
<2>Activation of restriction enzyme by electrochemically released magnesium ion.
<3>J. Biosci. Bioeng.
<4>98
<5>293-297
<6>2004
<7>Observation and cutting of DNA molecules at intended positions permit several new experimental
methods that are completely different from
conventional molecular biology methods; therefore several cutting
methods have been proposed and studied. In this paper. a new cutting
method for a DNA molecule by localizing the activity of a restriction
enzyme is presented. Since most restriction enzymes require magnesium
ions for their activation, local restriction enzyme activity can be
controlled by the local concentration of magnesium ions. Applying a
direct current (dc) voltage to a needle electrode of metallic magnesium
made it possible to control the local magnesium ion concentration at
the tip of the needle. The restriction enzyme was activated only, when
magnesium ions were electrochemically supplied.

<>

<1>Katsuragi, N., Kawakami, B., Maekawa, Y.
<2>Production method for PvuI restriction endonuclease.
<3>US Patent Office
<4>US 5049501
<5>
<6>1991
<7>A recombinant vector comprising an incorporated chromosome DNA fragment containing a PvuI
restriction endonuclease gene derived from Proteus vulgaris, a host transformed with the
recombinant vector and a method of producing PvuI restriction endonuclease characterized in
that the transformed host is cultivated and PvuI restriction endonuclease is harvested from
the resulting culture. Since the host transformed with the recombinant vector of the present
invention produces PvuI alone, it is unnecessary to remove PvuII in the purification process
for PvuI, and further, since its productivity for nonspecific DNase is lower in comparison
with conventional producer bacteria. DNase can be removed easily, making the production of
PvuI easy.

<>

<1>Katsuragi, N.T., Kawakami, B., Maekawa, Y.
<2>Production method for PvuI restriction endonuclease.
<3>European Patent Office
<4>EP 0374771 B
<5>
<6>1994
<7>A recombinant plasmid comprising an incorporated chromosome DNA fragment containing a PvuI
restriction endonuclease gene derived from Proteus vulgaris, a host transformed with the
plasmid and a method of producing PvuI restriction endonuclease characterized in that the
transformed host is cultivated and PvuI restriction endonuclease is harvested from the
resulting culture. Since the host transformed with the plasmid of the present invention
produces PvuI alone, it is unnecessary to remove PvuII in the purficiation process for PvuI,
and further, since its productivity for nonspecific DNase is lower in comparison with
conventional producer bacteria. DNase can be removed easily, making the production of PvuI
easy.

<>

<1>Katz, J.E., Dlakic, M., Clarke, S.
<2>Automated identification of putative methyltransferases from genomic open reading frames.
<3>Mol. Cell. Proteomics
<4>2
<5>525-540
<6>2003
<7>We have analyzed existing methodologies and created novel methodologies for the automatic
assignment of S-adenosylmethionine (AdoMet)-dependent
methyltransferase functionality to genomic open reading frames based on
predicted protein sequences. A large class of the AdoMet-dependent
methyltransferases shares a common binding motif for the AdoMet cofactor
in the form of a seven-strand twisted beta-sheet; this structural
similarity is mirrored in a degenerate sequence similarity that we refer
to as methyltransferase signature motifs. These motifs are the basis of
our assignments. We find that simple pattern matching based on the motif
sequence is of limited utility and that a new method of "sensitized
matrices for scoring methyltransferases" (SM(2)) produced with modified
versions of the MEME and MAST tools gives greatly improved results for the
Saccharomyces cerevisiae yeast genome. From our analysis, we conclude that
this class of methyltransferases makes up approximately 0.6-1.6% of the
genes in the yeast, human, mouse, Drosophila melanogaster, Caenorhabditis
elegans, Arabidopsis thaliana, and Escherichia coli genomes. We provide
lists of unidentified genes that we consider to have a high probability of
being methyltransferases for future biochemical analyses.

<>

<1>Katz, L.S., Turnsek, M., Kahler, A., Hill, V.R., Boyd, E.F., Tarr, C.L.
<2>Draft Genome Sequence of Environmental Vibrio cholerae 2012EL-1759 with Similarities to the V. cholerae O1 Classical Biotype.
<3>Genome Announcements
<4>2
<5>e00617-14
<6>2014
<7>Vibrio cholerae 2012EL-1759 is an environmental isolate from Haiti that was recovered in 2012
during a cholera outbreak. The genomic backbone is similar to
that of the prototypical V. cholerae O1 classical biotype strain O395, and it
carries the Vibrio pathogenicity islands (VPI-1 and VPI-2) and a cholera toxin
(CTX) prephage.

<>

<1>Kauc, L., Leszczynska, K.
<2>Identification of a new restriction endonuclease, EagI, from Enterobacter agglomerans.
<3>Acta Microbiol. Pol.
<4>35
<5>317-320
<6>1986
<7>The preliminary studies presented in this communication indicate that EndoR.
EagI and the restriction endonuclease EcoRII as well as EndoR. BstNI recognize
identical palindromic sites on the DNA substrate i.e. they are isoschizomers.
Since EndoR. EcoRII and EndoR. BstNI recognize the sequence CC(A/T)GG. EndoR.
EagI should recognize the same nucleotide sequence.

<>

<1>Kauc, L., Piekarowicz, A.
<2>Purification and properties of a new restriction endonuclease from Haemophilus influenzae Rf.
<3>Eur. J. Biochem.
<4>92
<5>417-426
<6>1978
<7>Haemophilus influenzae Rf 232, showing the phenomena of restriction and
modification, contains an endonuclease that inactivates in vitro the biological
activity of DNAs lacking the strain-specific modification.  This specific
restriction endonuclease has been purified to near homogeneity by a procedure
that includes DNA-agarose chromatography.  This highly purified enzyme requires
ATP and Mg2+ for activity and is stimulated by S-adenosylmethionine.  The
enzyme seems to cleave DNA at well-defined sites, since it produces a specific
pattern of bands upon agarose gel electrophoresis.  The enzyme has no ATPase
activity.  A methylase activity is observed in the course of the
endonucleolytic reaction, which probably protects some of the DNA sites from
cleavage.

<>

<1>Kauc, L., Piekarowicz, A.
<2>Degradation of Transforming Deoxyribonucleic Acid by the ATP Dependent Restriction Endonuclease Isolated from Haemophilus Influenzae Rf.
<3>Modern Trends in Bacterial Transformation and Transfection., Elsevier, Portoles, A., Lopez, R., Espinosa, M., Amsterdam
<4>0
<5>257-262
<6>1976
<7>The deoxyribonucleic acid of different serological types of Haemophilus
influenzae was shown to be degraded by the ATP dependent restriction
endonuclease isolated from H.influenzae Rf.

<>

<1>Kauc, L., Piekarowicz, A.
<2>Degradation of transforming and transfecting DNA by the restriction endonucleases of Type I and Type II isolated from Haemophilus influenzae.
<3>Acta Microbiol. Pol.
<4>26
<5>137-148
<6>1977
<7>The restriction endonucleases of type I and II from Haemophilus influenzae were studied for
their activity on transforming and transfecting DNA.  Type I restriction enzyme from
Haemophilus influenzae Rf, which requires adenosine 5'-triphosphate, reduced the size of
unmodified bacterial DNA from 66 x 106 daltons to approximately 18 x 106 daltons and did not
attack modified DNA.  The action of this enzyme gives only a low level of inactivation of
single and linked markers in the transforming DNA.  In contrast the HP1c1 phage DNA was
drastically inactivated by this enzyme.  The endoR.Hind III degrades the unmodified bacterial
DNA but the segments generated by this enzyme are still capable of being integrated in
transformation.  The enzyme has no activity on HP1c1 phage DNA.

<>

<1>Kaufman, D.L., Evans, G.A.
<2>Restriction endonuclease cleavage at the termini of PCR products.
<3>Biotechniques
<4>9
<5>304-305
<6>1990
<7>None

<>

<1>Kaufmann, G.
<2>Anticodon nucleases.
<3>Trends Biochem. Sci.
<4>25
<5>70-74
<6>2000
<7>A tRNALys-specific anticodon nuclease is kept in a latent form in a rare Escherichia coli
strain, complexed with a DNA restriction enzyme. A phage T4 inhibitor of DNA restriction
activates anticodon nuclease, but other T4 proteins restore tRNALys. Detection of a homologous
system in Neisseria and a different anticodon nuclease in colicin E5 suggest ubiquity and
diversity of such tRNA toxins. Analysis of these systems could reveal novel RNA recognition
and cleavage mechanisms.

<>

<1>Kaufmann, G., Amitsur, M., Chapman-Shimshoni, D.
<2>In vivo reconstitution of latent anticodon nuclease: A tRNA restriction enzyme masked by hsd restriction-modification proteins.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>174
<6>1993
<7>E. coli prr+ strains encode a latent form of phage T4-induced, tRNA Lys-specific anticodon
nuclease. The latent enzyme comprises a core factor encoded by prrC and cognate masking
elements encoded by flanking, hsd type Ic restriction modification genes. We have
reconstituted latent anticodon nuclease from separate, core and masking components in vivo by
complementing a prr cosmid carrying a null prrC mutation with PrrC provided in trans.
Expression of prrC from a low copy plasmid, insufficient to elicit detectable core activity by
itself, yielded in the presence of the hsd masking factors a full-fledged latent anticodon
nuclease phenotype. The data indicate that the Hsd proteins not only mask PrrC's activity but
also stabilize it maintaining the RNA restriction enzyme as an antiviral contingency.

<>

<1>Kaur, J., Verma, H., Tripathi, C., Khurana, J.P., Lal, R.
<2>Draft Genome Sequence of a Hexachlorocyclohexane-Degrading Bacterium, Sphingobium baderi Strain LL03T.
<3>Genome Announcements
<4>1
<5>e00751-13
<6>2013
<7>Sphingobium baderi strain LL03(T) was isolated from hexachlorocyclohexane (HCH)-contaminated
soil from Spolana, Czech Republic. Strain LL03(T) is a mutant
that is deficient in linB and linC (genes that encode hexachlorocyclohexane
haloalkane dehalogenase and dehydrogenase, respectively). The draft genome
sequence of LL03(T) (~4.85 Mb) consists of 92 contigs and 4,914 coding sequences,
with a G+C content of 63.5%.

<>

<1>Kaur, K., Rohozinski, J., Goorha, R.
<2>Identification and characterization of the frog virus 3 DNA methyltransferase gene.
<3>J. Gen. Virol.
<4>76
<5>1937-1943
<6>1995
<7>Cytosine DNA methyltransferases (MTases) first recognize specific nucleotide sequences and
then transfer a methyl group from S-adenosylmethionine to cytosine.  This division of function
is reflected in five highly conserved motifs shared by cytosine Mtases.  The region containing
the first four motifs is responsible for the catalytic function whereas the region containing
the fifth motif V provides specificity of binding to DNA.  In at least one case, two separate
proteins, one containing the first four motifs and the second containing the last motif
combine to provide full functional activity.  In the frog virus 3 (FV3) genome we have
identified an open reading frame (ORF) whose deduced amino acid (aa) sequence contains motifs
characteristic of prokaryotic as well as eukaryotic MTases.  The ORF consists of 642 bp which
codes for a protein of 214 aa with a predicted molecular mass of 24.8 kDa.  This ORF contains
the first four highly conserved motifs of cytosine MTases but the fifth motif, responsible for
DNA binding specificity, is missing.  Presumably, FV3 MTase is composed of two subunits.
Northern blot analysis showed that the putative MTase ORF is transcribed into two transcripts
belonging to the delayed-early class of FV3 messages.  These two transcripts appear to be
initiated at two different start sites but terminate in the same 3' region of the gene.  The
transcription start sites are not preceded by any known promoter sequences, but two regions of
hyphenated dyad symmetry are present at the 3' end of the message.  A protein with a
molecular mass of ~28 kDa was synthesized by a rabbit reticulocyte lysate programmed with
capped runoff transcripts from the cloned gene, suggesting that the ORF can be transcribed
into a message coding for a viral protein.  Overall, our results suggest that we have
identified a gene for a subunit of MTase in the FV3 genome.

<>

<1>Kaur, N., Kumar, S., Bala, M., Raghava, G.P., Mayilraj, S.
<2>Draft Genome Sequence of Amycolatopsis decaplanina Strain DSM 44594T.
<3>Genome Announcements
<4>1
<5>e0013813
<6>2013
<7>We report the 8.5-Mb genome sequence of Amycolatopsis decaplanina strain DSM 44594(T),
isolated from a soil sample from India. The draft genome of strain DSM
44594(T) consists of 8,533,276 bp with a 68.6% G+C content, 7,899 protein-coding
genes, and 57 RNAs.

<>

<1>Kaus-Drobek, M., Czapinska, H., Sokolowska, M., Tamulaitis, G., Szczepanowski, R.H., Urbanke, C., Siksnys, V., Bochtler, M.
<2>Restriction endonuclease MvaI is a monomer that recognizes its target sequence asymmetrically.
<3>Nucleic Acids Res.
<4>35
<5>2035-2046
<6>2007
<7>Restriction endonuclease MvaI recognizes the sequence CC/WGG (W stands for A or T, '/'
designates the cleavage site) and generates products with single nucleotide 5'-overhangs. The
enzyme has been noted for its tolerance towards DNA modifications. Here, we report a
biochemical characterization and crystal structures of MvaI in an apo-form and in a complex
with target DNA at 1.5 A resolution. Our results show that MvaI is a monomer and recognizes
its pseudosymmetric target sequence asymmetrically. The enzyme consists of two lobes. The
catalytic lobe anchors the active site residues Glu36, Asp50, Glu55 and Lys57 and contacts the
bases from the minor grove side. The recognition lobe mediates all major grove interactions
with the bases. The enzyme in the crystal is bound to the strand with T at the center of the
recognition sequence. The crystal structure with calcium ions and DNA mimics the prereactive
state. MvaI shows structural similarities to BcnI, which cleaves the related sequence CC/SGG
and to MutH enzyme, which is a component of the DNA repair machinery, and nicks one DNA strand
instead of making a double-strand break.

<>

<1>Kautharapu, K.B., Jarboe, L.R.
<2>Genome Sequence of the Psychrophilic Deep-Sea Bacterium Moritella marina MP-1 (ATCC 15381).
<3>J. Bacteriol.
<4>194
<5>6296-6297
<6>2012
<7>Moritella marina MP-1 is a bacterial species known for its production of docosahexaenoic acid.
We present the draft genome sequence of the type strain
Moritella marina MP-1 (ATCC 15381), having 4,636,778 bp with a G+C content of
40.5% and consisting of 83 contigs.

<>

<1>Kavets, A.N., Solonin, A.S.
<2>A new E. coli strain, produces the restriction endonuclease CfrBI. It is useful for the production of the restriction endonuclease, which can recognize and cleave the sequence 5'-C/CWWGG-3' in double stranded DNA.
<3>Russian Patent Office
<4>RU 2038382
<5>
<6>1995
<7>
<>

<1>Kaw, M.K., Blumenthal, R.M.
<2>Translational independence between overlapping genes for a restriction endonuclease and its transcriptional regulator.
<3>BMC Mol. Biol.
<4>11
<5>87
<6>2010
<7>Background: Most type II restriction-modification (RM) systems have two independent enzymes
that act on the same DNA sequence: a modification
methyltransferase that protects target sites, and a restriction
endonuclease that cleaves unmethylated target sites. When RM genes
enter a new cell, methylation must occur before restriction activity
appears, or the host's chromosome is digested. Transcriptional
mechanisms that delay endonuclease expression have been identified in
some RM systems. A substantial subset of those systems is controlled by
a family of small transcription activators called C proteins. In the
PvuII system, C. PvuII activates transcription of its own gene, along
with that of the downstream endonuclease gene. This regulation results
in very low R. PvuII mRNA levels early after gene entry, followed by
rapid increase due to positive feedback. However, given the lethal
consequences of premature REase accumulation, transcriptional control
alone might be insufficient. In C-controlled RM systems, there is a +/-
20 nt overlap between the C termination codon and the R (endonuclease)
initiation codon, suggesting possible translational coupling, and in
many cases predicted RNA hairpins could occlude the ribosome binding
site for the endonuclease gene.Results: Expression levels of lacZ
translational fusions to pvuIIR or pvuIIC were determined, with the
native pvuII promoter having been replaced by one not controlled by C.
PvuII. In-frame pvuIIC insertions did not substantially decrease either
pvuIIC-lacZ or pvuIIR-lacZ expression (with or without C. PvuII
provided in trans). In contrast, a frameshift mutation in pvuIIC
decreased expression markedly in both fusions, but mRNA measurements
indicated that this decrease could be explained by transcriptional
polarity. Expression of pvuIIR-lacZ was unaffected when the pvuIIC stop
codon was moved 21 nt downstream from its WT location, or 25 or 40 bp
upstream of the pvuIIR initiation codon. Disrupting the putative
hairpins had no significant effects.Conclusions: The initiation of
translation of pvuIIR appears to be independent of that for pvuIIC.
Direct tests failed to detect regulatory rules for either gene overlap
or the putative hairpins. Thus, at least during balanced growth,
transcriptional control appears to be sufficiently robust for proper
regulation of this RM system.

<>

<1>Kawabata, H., Norris, S.J., Watanabe, H.
<2>BBE02 disruption mutants of Borrelia burgdorferi B31 have a highly transformable, infectious phenotype.
<3>Infect. Immun.
<4>72
<5>7147-7154
<6>2004
<7>We constructed highly transformable and infectious Borrelia burgdorferi B31 by inactivating
BBE02, a putative restriction-modification gene on
the linear plasmid lp25. The low-passage-number B31 clones 5A4
(containing all plasmids) and 5A18 (lp28-4(-) lp56(-)) were used for
this study, and BBE02 was disrupted by homologous recombination. The
transformation efficiency with the shuttle vector pBSV2C03::gntDeltakan
was increased from <1 to similar to10 colonies per mug of DNA for 5A4
and 5A4 BBE02::Kan(r) and from 14 to approximately 600 colonies per mug
of DNA for 5A18 and 5A18 BBE02::Kanr. lp25, which is required for
infectivity in mice, was retained in BBE02 mutants transformed with
pBSV2C03::gntDeltakan, but lp25 was not detected in transformants of
the parental clones 5A4 and 5A18. BBE02 disruptants and
pBSV2C03::gntDeltakan transformants of these clones remained infectious
in C3H/HeN mice, and the 50% infective doses of the BBE02 disruptants
were <10(2) organisms per mouse. The inactivation of BBE02 thus
eliminates a transformation barrier for infectious B. burgdorferi B31
and will provide a valuable tool for studying the virulence factors of
Lyme disease.

<>

<1>Kawakami, B.
<2>Application of recombinant DNA technique to the production of N-acetylneuraminate lyase and restriction enzymes.
<3>Ph.D. Thesis, University of Kyoto University
<4>
<5>1-102
<6>1991
<7>Chapter I. N-Acetylneuraminate lyase Chapter II. 1) Cloning of BamHI RM system in B. subtilis
2) Cloning and sequencing of the AccI RM system 3) Cloning and expression of the BanI and
BanIII RM systems from Bacillus aneurinolyticus 4) Nucleotide sequence of the BanI RM genes 5)
Nucleotide sequence of the BanIII M gene.

<>

<1>Kawakami, B., Hilzheber, C., Nagatomo, M., Oka, M.
<2>Cloning and nucleotide sequences of the AccI restriction-modification genes in Acinetobacter calcoaceticus.
<3>Agric. Biol. Chem.
<4>55
<5>1553-1559
<6>1991
<7>The genes of the AccI restriction-modification system specific for
GT(A/C)(G/T)AC were cloned from the chromosomal DNA of Acinetobacter
calcoaceticus, and their nucleotides sequenced.  The restriction and
modification genes coded for polypeptides with calculated molecular weights of
42,494 and 63,078, respectively.  Both the enzymes were coded by the same DNA
strand and the restriction gene was upstream of the methylase gene, separated
by 2 bp.  The restriction gene was significantly expressed in E. coli cells, so
that the AccI restriction endonuclease could be purified to homogeneity.
Analysis by sodium dodecyl sulfate-polyacrylamide  gel electrophoresis and gel
filtration indicated that the catalytically active form of the endonuclease was
tetrameric.  Sequence comparison with related enzymes indicated that AccI
methylase contained a segment of tetra-amino acids, NPPY, characteristic of
N6-adenine methylases.  In addition, some homologous regions were found in the
sequence of HincII methylase specific for GT(C/T) (A/G)AC.

<>

<1>Kawakami, B., Katsuragi, N., Maekawa, Y., Imanaka, T.
<2>Cloning and expression of the BamHI restriction-modification system in Bacillus subtilis.
<3>J. Ferment. Bioeng.
<4>70
<5>211-214
<6>1990
<7>The genes of the GGATCC-specific BamHI restriction-modification system of
Bacillus amyloliquefaciens H have been cloned and expressed in Bacillus
subtilis MT-2.  B. subtilis MT-2 carrying the recombinant plasmid (pBamHIRM22)
produced about 10-fold more BamHI restriction endonuclease and BamHI methylase
than B. amyloliquefaciens H did.  B. subtilis MT-2(pBamHIRM22) restricted
unmodified phage.  Restriction and modification genes were stably maintained in
B. subtilis MT-2 on one plasmid and the produced BamHI endonuclease remained
stable even in the stationary phase of the culture.  BamHI endonuclease from B.
subtilis MT-2 (pBamHIRM22) had the same molecular weight and N-terminal amino
acid sequence as that from B. amyloliquefaciens H.

<>

<1>Kawakami, B., Maeda, Y.
<2>BamHI restriction endonuclease of Bacillus amyloliquefaciens.
<3>Japanese Patent Office
<4>JP 02057185
<5>
<6>1990
<7>A chromosomal DNA fragment containing Bacillus amyloliquefaciens H strain-derived BamHI
restriction endonuclease gene is inserted into a plasmid to form a recombinant plasmid capable
of being replicated in bacteria of the genus Bacillus. The recombinant plasmid is introduced
into bacteria of the genus Bacillus for transformation, and the transformant bacteria of the
genus Bacillus are incubated and the BamHI restriction endonuclease as produced from the
recombinant bacteria is isolated from the culture medium. In preparation of recombinant
plasmid: pTB53 plasmid is used as a vector DNA. A chromosomal DNA fragment as prepd. from
Bacillus amyloliquefaciens strain H is inserted into the vector DNA to obtain a new
recombinant plasmid BamHIRM 22. The recombinant plasmid BamHIRM22 is introduced into Bacillus
subtilis MT2 or MI113 for transformation. The transformed bacteria are then incubated in a
medium (e.g., L-Broth medium) and they produce the intended BamHI restriction endonuclease.

<>

<1>Kawakami, B., Maekawa, Y.
<2>Cloning and expression of restriction enzyme AccI gene.
<3>Japanese Patent Office
<4>JP 03007584
<5>
<6>1991
<7>
<>

<1>Kawakami, B., Maekawa, Y.
<2>BanI restriction endonuclease - obtained from Bacillus aneurinolyticus and replicating in Escherichia coli.
<3>Japanese Patent Office
<4>JP 1016585
<5>
<6>1989
<7>Recombinant plasmid which may be replicated with Escherichia coli, the plasmid incorporates
chromosome DNA fragment contg. BanI restriction endonuclease gene origined from Bacillus
aneurinolyticus IAM 1077. The Escherichia microorganism is transformed with recombinant
plasmid incorporated with chromosome DNA fragment contg. the BanI restriction endonuclease
gene. BanI restriction endonuclease is produced by culturing Escherichia microorganism
incorporated chromosome DNA fragment contg. BanI restriction endonuclease gene origined from
Bacillus aneurinolyticus IAM 1077.

USE - The novel recombinant plasmid incorporated chromosome DNA fragment contg. BanI
restriction endonuclease gene, microorganism transformed by introducing the plasmid and a
process for producing BanI restriction endonuclease from the micro-organism are useful in
genetic technology.


<>

<1>Kawakami, B., Sasaki, A., Oka, M., Maekawa, Y.
<2>Nucleotide sequence of the gene coding for the BanIII DNA methyltransferase in Bacillus aneurinolyticus.
<3>Agric. Biol. Chem.
<4>54
<5>3227-3233
<6>1990
<7>The gene coding for the ATCGAT specific BanIII DNA methyltransferase (M.BanIII)
of Bacillus aneurinolyticus was cloned and its nucleotides sequenced.  The
coding region was assigned on the nucleotide sequence on the basis of the
N-terminal amino acid sequence and molecular weight of the enzyme.  The
M.BanIII gene coded for a protein of 580 amino acid residues (MW 66,344).
Comparison with other methylases indicated that the M.BanIII sequence contained
a segment of tetra-amino acids, NPPY, characteristic of N6-adenine methylases.
In addition, some homologous regions were found in the sequences of type II
adenine methylases PaeR7I(CTCGAG), TaqI(TCGA) and PstI(CTGCAG), containing TCGA
within the recognition sequences.

<>

<1>Kawalec, M., Borsuk, P., Piechula, S., Stepien, P.P.
<2>A novel restriction endonuclease UnbI, a neoschizomer of Sau96I from an unidentified psychrofilic bacterium from Antarctica is inhibited by phosphate ions.
<3>Acta Biochim. Pol.
<4>44
<5>849-852
<6>1997
<7>A novel type II restriction endonuclease UnbI was isolated from an unidentified psychrofilic
bacterial strain from Antarctica.  UnbI recognizes and cleaves the sequence 5'-GGNCC-3',
producing 5 nucleotide long sticky ends.  In this respect it differs from its neoschizomer
Sau96I and all other restriction enzymes recognizing this sequence.  UnbI has a relatively low
temperature optimum of 15 C to 20 C and its activity is completely inhibited by inorganic
phosphate.

<>

<1>Kawamura, M., Sakakibara, M., Watanabe, T., Kita, K., Hiraoka, N., Obayashi, A., Takagi, M., Yano, K.
<2>A new restriction endonuclease from Spirulina platensis.
<3>Nucleic Acids Res.
<4>14
<5>1985-1989
<6>1986
<7>Three restriction endonucleases, SplI, SplII and SplIII have been purified partially from
Spirulina platensis subspecies siamese and named.  SplI cleaves bacteriophage lambda DNA at
one site, phi X 174 RF DNA at two sites, but does not cleave pBR322 DNA.  This enzyme
recognizes the sequence 5'CGTACG3'3'GCATCG5'and cuts the site indicated by the arrows.
SplIII is an isoschizomer of HaeIII.

<>

<1>Kawamura, M., Sakakibara, M., Watanabe, T., Obayashi, A., Hiraoka, N., Kita, K.
<2>New restriction endonuclease SplI from Spirulina algae recognising specific base sequence, useful in genetic engineering.
<3>United Kingdom Patent Office
<4>GB 2162852 A
<5>
<6>1984
<7>The new restriction endonuclease SplI recognises the base sequence C^GTACG and cuts it as
shown, producing fragments having a 4-base single strand extension. SplI cleaves ds DNA from
lambda phage, at only one position, and DNA from both col E1 and phiX-174RF at 2 positions. It
is activated by 5-20 mM Mg ions and is active over the NaCl concentration range 0-200 mM. The
optimum NaCl concentration is 100-120 mM, and SplI is active at pH 5.5-10 (optimum 7-7.5) and
20-65 degrees C (optimum 50-55 degrees C). It is completely inactivated after 60 min. at 75
degrees C. SplI has a mol. wt. of 370,000-430,000 (gel filtration and requires no cofactors
such as ATP or S-adenosyl-methionine.

<>

<1>Kawamura, M., Sakakibara, M., Watanabe, T., Obayashi, A., Hiraoka, N., Kita, K.
<2>Novel restriction endonuclease SplI and process for the production of the same.
<3>US Patent Office
<4>US 4886756
<5>
<6>1989
<7>A novel restriction endonuclease SplI which has the following physicochemical properties: (1)
recognizing the following base sequences in double-stranded deoxyribonucleic acid
5'-C^GTACG-3'
3'-GCATG^C-5'
and cleaving said sequences in the phosphodiester bonds between C and G as indicated with the
vertical arrows to produce DNA fragments having one strand comprising four bases at the
5'-terminal; (2) cleaving double-stranded deoxyribonucleic acid lambda-DNA in one position,
Col E1 in two positions and PhiX174 RF in two positions; (3) being activated with 5 to 20 mM
Mg2+; and (4) exhibiting an activity at a NaCl concentration of 0 to 200 mM; and a process for
the production of the restriction endonuclease SplI which comprises culturing a restriction
endonuclease SplI-producing alga belonging to the genus Spirulina, collecting the cells,
obtaining a cell-free extract therefrom the separating the purifying the restriction
endonuclease SplI.

<>

<1>Kawarabayasi, Y. et al.
<2>Complete sequence and gene organization of the genome of a hyper-thermophilic Archaebacterium, Pyrococcus horikoshii OT3 (Supplement).
<3>DNA Res.
<4>5
<5>147-155
<6>1998
<7>The circular genome of Pyrococcus horikoshii OT3, (1,738,505 bp long), was opened at the
junction of Srf I-C and Srf I-E fragments, and is represented by a linear map starting from
this site.  The nucleotide positions are indicated on the linear map by numerals in kb.  Above
and below the physical maps, the potential protein-coding regions assigned are indicated by
boxes with arrowheads, indicating the direction of transcription.  The detailed assignment
procedures are described in the main article in this issue.  The results of ORF similarity and
motif searches are shown using the following color codes: red, similarities to reported genes
with known functions; blue, similarities to hypothetical genes; green, some motifs but without
significant similarity to the registered sequences; and no color, no apparent similarity to
any reported genes and no significant protein motifs.  The ORF names defined in the main
article are given to each ORF, and the positions of the rRNA and tRNA genes are indicated by
closed box and vertical bars with a "T", respectively.  The sequence data reported here have
been deposited in DDBJ/Genbank/EMBL databases under accession numbers AP000001 (nucleotide
positions 1-287,000), AP000002 (287,001-544,000), AP000003 (544,001-777,000), AP000004
(777,001-994,000), AP000005 (994,001-1,166,000), AP000006 (1,166,001-1,485,000), and AP000007
(1,485,001-1,738,505).

<>

<1>Kawarabayasi, Y. et al.
<2>Complete sequence and gene organization of the genome of a hyper-thermophilic Archaebacterium, Pyrococcus horikoshii OT3.
<3>DNA Res.
<4>5
<5>55-76
<6>1998
<7>The complete sequence of the genome of a hyper-thermophilic Archaebacterium, Pyrococcus
horikoshii OT3, has been determined by assembling the sequences of the physical map-based
contigs of fosmid clones and of long polymerase chain reaction products which were used for
gap-filling.  The entire length of the genome was 1,738,505 bp.  The authenticity of the
entire genome sequence was supported by restriction analysis of long PCR products, which were
directly amplified from the genomic DNA.  As the potential protein-coding regions, a total of
2061 open reading frames were assigned, and by similarity search against public databases, 406
(19.7%) were related to genes with putative function and 453 (22.0%) to the sequences
registered but with unknown function.  The remaining 1202 ORFs (58.3%) did not show any
significant similarity to the sequences in the databases.  Sequence comparison among the
assigned ORFs in the genome provided evidence that a considerable number of ORFs were
generated by sequence duplication.  By similarity search, 11 ORFs were assumed to contain
intein elements.  The RNA genes identified were a single 16S-23S rRNA operon, two 5S rRNA
genes and 46 tRNA genes including two with the intron structure.  All the assigned ORFs and
RNA coding regions occupied 91.25% of the whole genome.  The data presented in this paper are
available on the internet at http://www.nite.go.jp.

<>

<1>Kawarabayasi, Y. et al.
<2>Complete genome sequence of an aerobic hyper-thermophilic crenarchaeon, Aeropyrum pernix K1.
<3>DNA Res.
<4>6
<5>83-101
<6>1999
<7>The complete sequence of the genome of an aerobic hyper-thermophilic crenarchaeon, Aeropyrum
pernix K1, which optimally grows at 95 degrees C, has been determined by the whole genome
shotgun method with some modifications. The entire length of the genome was 1,669,695 bp. The
authenticity of the entire sequence was supported by restriction analysis of long PCR
products, which were directly amplified from the genomic DNA. As the potential protein-coding
regions, a total of 2,694 open reading frames (ORFs) were assigned. By similarity search
against public databases, 633 (23.5%) of the ORFs were related to genes with putative function
and 523 (19.4%) to the sequences registered but with unknown function. All the genes in the
TCA cycle except for that of alpha-ketoglutarate dehydrogenase were included, and instead of
the alpha-ketoglutarate dehydrogenase gene, the genes coding for the two subunits of
2-oxoacid:ferredoxin oxidoreductase were identified. The remaining 1,538 ORFs (57.1%) did not
show any significant similarity to the sequences in the databases. Sequence comparison among
the assigned ORFs suggested that a considerable member of ORFs were generated by sequence
duplication. The RNA genes identified were a single 16S-23S rRNA operon, two 5S rRNA genes and
47 tRNA genes including 14 genes with intron structures. All the assigned ORFs and RNA coding
regions occupied 89.12% of the whole genome. The data presented in this paper are available on
the internet homepage (http://www.mild.nite.go.jp).

<>

<1>Kawarabayasi, Y. et al.
<2>Complete genome sequence of an aerobic thermoacidophilic crenarchaeon, Sulfolobus tokodaii strain7.
<3>DNA Res.
<4>8
<5>123-140
<6>2001
<7>The complete genomic sequence of an aerobic thermoacidophilic crenarchaeon, Sulfolobus
tokodaii strain7 which optimally grows at 80 degrees C, at low pH, and under aerobic
conditions, has been determined by the whole genome shotgun method with slight modifications.
The genomic size was 2,694,756 bp long and the G + C content was 32.8%. The following
RNA-coding genes were identified: a single 16S-23S rRNA cluster, one 5S rRNA gene and 46 tRNA
genes (including 24 intron-containing tRNA genes). The repetitive sequences identified were
SR-type repetitive sequences, long dispersed-type repetitive sequences and Tn-like repetitive
elements. The genome contained 2826 potential protein-coding regions (open reading frames,
ORFs). By similarity search against public databases, 911 (32.2%) ORFs were related to
functional assigned genes, 921 (32.6%) were related to conserved ORFs of unknown function, 145
(5.1%) contained some motifs, and the remaining 849 (30.0%) did not show any significant
similarity to the registered sequences. The ORFs with functional assignments included the
candidate genes involved in sulfide metabolism, the TCA cycle and the respiratory chain.
Sequence comparison provided evidence suggesting the integration of plasmid, rearrangement of
genomic structure, and duplication of genomic regions that may be responsible for the larger
genomic size of the S. tokodaii strain7 genome. The genome contained eukaryote-type genes
which were not identified in other archaea and lacked the CCA sequence in the tRNA genes. The
result suggests that this strain is closer to eukaryotes among the archaea strains so far
sequenced. The data presented in this paper are also available on the internet homepage
(http://www.bio.nite.go.jp/E-home/genome_list-e.html).

<>

<1>Kawasaki, K., Shibata, T.
<2>Mitochondrial HSP70 as a subunit of eukaryotic multi-site-specific endonuclease, Endo.SceI; auto-phosphorylation and heat stability.
<3>Mol. Biol. Cell
<4>6
<5>135a
<6>1995
<7>A eukaryotic site-specific endonuclease, Endo.SceI, that cuts DNA at multiple sites, is an
initiator of homologous gene conversion in yeast mitochondria.  The active form of Endo.SceI
is a heterodimer of a 75 kDa-subunit and a 50 kDa-subunit.  The 75 kDa-subunit is a
mitochondrial 70 kDa heat shock protein (HSP70) and encoded by a nuclear gene, ENS1 (identical
to SSC1).  HSP70 protein has ATP-related activities; such as ATP binding, an ATPase, and a
protein kinase.  We investigated ATP-related activities of HSP70 protein in Endo.SceI.  We
found that the 75 kDa-subunit of Endo.SceI exhibits a protein kinase activity.  The kinase
activity specifically phosphorylated the 75 kDa-subunit, but not the 50 kDa-subunit or other
proteins tested.  The phosphorylating activity requires Ca2+ and an acidic pH, and is greatly
stimulated by the presence of the 50 kDa-subunit.  Ca2+ increased the electrophoretic mobility
of Endo.SceI.  The auto-phosphorylation of the 75 kDa-subunit, by itself, does not have a
direct effect on DNase, but the phosphorylation condition stabilizes DNase activity of
Endo.SceI against heat-inactivation.  This stabilization was dependent upon Ca2+ and ATP, as
well as ATP-gamma-S or ADP.  In addition, ATP-gamma-S and ADP inhibited the
auto-phosphorylation of the 75 kDa-subunit.  These findings suggest that Ca2+ ions induced the
conformational change of Endo.SceI in which the binding of nucleotides, such as ATP and ADP,
regulated the heat-stability and autophosphorylation of Endo.SceI.

<>

<1>Kawasaki, K., Shibata, T., Ito, F.
<2>Roles of the HSP70-Subunit in a eukaryotic multi-site-specific endonuclease, Endo.SceI: Autophosphorylation and heat stability.
<3>Biosci. Biotechnol. Biochem.
<4>68
<5>2557-2564
<6>2004
<7>The 70 kDa heat shock proteins (HSP70) are a family of molecular chaperones that bind
transiently to unfolded proteins in an ATP/ADP
dependent manner. Endo.SceI comprises a unique example for mitochondria. HSP70, which exists
in a stable complex with a nucleolytic subunit as a multi-site specific DNase. The
HSP70-subunit in Endo.SceI was autophosphorylated by ATP in vitro. The
autophosphorylation was higher in the Endo.SceI complex form than in
the free form. Although the autophosphorylation had no significant
effect on the endonucleolytic activity of Endo.SceI, the factors
favoring autophosphorylation protected the endonucleolytic activity of
Endo.SceI against heat inactivation. ATP, adenosine
5'-O-(3-thiotriphosphate) (ATP-gamma-S), and ADP not only protected the
endonucleolytic activity against heat inactivation in the presence of
Ca2+ ions, but also reduced the labeling of the HSP70-subunit by
[gamma-P-32]ATP in Endo.SceI. These findings suggest that the
HSP70-subunit shields Endo.SceI from heat inactivation through ATP/ADP
binding.

<>

<1>Kawasaki, K., Takahashi, M., Ando, T., Shibata, T.
<2>Sequence-specific complex formation of DNA and a eukaryotic sequence-specific endonuclease, SceI.
<3>Eur. J. Biochem.
<4>202
<5>665-671
<6>1991
<7>Endo.SceI is a eukaryotic sequence-specific endonuclease of 120 kDa that causes
sequence-specific double-stranded scission of DNA.  Unlike results with restriction enzymes,
we found a consensus sequence around the cleavage sites for Endo.SceI instead of a common
sequence.  We searched for conditions for studying the binding of Endo.SceI to DNA other than
cutting.  Under optimized conditions including gel mobility shift assay, Endo.SceI exhibited
sequence-specific binding to a short double-stranded DNA (41 base pairs) containing a cleavage
site and the DNA reisolated from the protein-DNA complex was not cleaved.  The analysis of the
complex of Endo.SceI and DNA isolated by the gel mobility shift experiments showed that the
DNA-binding entity in the Endo.SceI preparation does have Endo.SceI activity and consists of
an equal amount of 75-kDa and 50-kDa polypeptides.  Based on this observation from previous
studies, we conclude that Endo.SceI is a heterodimer of the 75-kDa and 50-kDa subunits.  Under
the present assay conditions, Endo.SceI did not show binding to single-stranded DNA having the
same sequence of either plus or minus strand of the double-stranded DNA containing the
cleavage site (the 41-bp DNA).  Endo.SceI showed significantly higher affinity for the
consensus sequence than the major cleavage site in pBR322 DNA.  Unlike the cleavage of DNA by
Endo.SceI which requires Mg2+, this sequence-specific binding is independent of but stimulated
by Mg2+.

<>

<1>Kawasaki, K., Takahashi, M., Natori, M., Shibata, T.
<2>DNA sequence recognition by a eukaryotic sequence-specific endonuclease, Endo.SceI, from Saccharomyces cerevisiae.
<3>J. Biol. Chem.
<4>266
<5>5342-5347
<6>1991
<7>A eukaryotic sequence-specific endonuclease, Endo.SceI, causes
sequence-specific double-stranded scission of double-stranded DNA to produce
cohesive ends with four bases protruding at the 3' termini.  Unlike the case of
restriction enzymes, an asymmetric 26-base pair consensus sequence was found
around the cleavage site for Endo.SceI instead of a common sequence.  We
analyzed the base pairs that interacted with Endo.SceI on the recognition of
its cleavage sites.  A region comprising -10 through +16 base pairs from the
center of the cleavage site was shown to be essential and sufficient for the
sequence-specific cutting with Endo.SceI by experiments involving synthesized
DNAs.  Methylation interference experiments indicate that bases in the region
comprising the +7 through +14 base pairs are involved in close contact with
Endo.SceI in its recognition of the cleavage site.  This +7 through +14-base
pair region overlaps the most stringently conserved sequence in the consensus
sequence for the cleavage site, suggesting that this region constitutes the
core for the recognition by Endo.SceI.

<>

<1>Kawasaki, M., Makino, S.-I., Matsuzawa, H., Satow, Y., Ohya, Y., Anraku, Y.
<2>Folding-dependent in vitro protein splicing of the Saccharomyces cerevisiae VMA1 protozyme.
<3>Biochem. Biophys. Res. Commun.
<4>222
<5>827-832
<6>1996
<7>VMA1 translational product undergoes excision of a 50-kDa intervening segment (VDE:
VMAI-derived endonuclease) and religation of the flanking regions to create a 69-kDa catalytic
subunit of vacuolar membrane H+-ATPase.  VDEs conjugated with polypeptides at both N- and
C-terminal ends were expressed in Escherichia coli and examined for their ability to catalyze
self-splicing.  Processed VDE was found in soluble pools, while unspliced precursors
accumulated in insoluble pools, forming inclusion bodies.  We demonstrate in vitro protein
splicing by refolding of the denatured precursor molecules.  The processing reaction
efficiently occurs with the purified precursor peptide.  VDE bracketed by only 6 proximal and
4 distal amino acids is autocatalytically processed.

<>

<1>Kawashima, T., Amano, N., Koike, H., Makino, S.-i., Higuchi, S., Kawashima-Ohya, Y., Watanabe, K., Yamazaki, M., Kanehori, K., Kawamoto, T., Nunoshiba, T., Yamamoto, Y., Aramaki, H., Makino, K., Suzuki, M.
<2>Archaeal adaptation to higher temperatures revealed by genomic sequence of Thermoplasma volcanium.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>14257-14262
<6>2000
<7>The complete genomic sequence of the archaeon Thermoplasma volcanium, possessing optimum
growth temperature (OGT) of 60 degrees C, is reported. By systematically comparing this
genomic sequence with the other known genomic sequences of archaea, all possessing higher OGT,
a number of strong correlations have been identified between characteristics of genomic
organization and the OGT. With increasing OGT, in the genomic DNA, frequency of clustering
purines and pyrimidines into separate dinucleotides rises (e.g., by often forming AA and TT,
whereas avoiding TA and AT). Proteins coded in a genome are divided into two distinct
subpopulations possessing isoelectric points in different ranges (i.e., acidic and basic), and
with increasing OGT the size of the basic subpopulation becomes larger. At the metabolic
level, genes coding for enzymes mediating pathways for synthesizing some coenzymes, such as
heme, start missing. These findings provide insights into the design of individual genomic
components, as well as principles for coordinating changes in these designs for the adaptation
to new environments. Full author list: Kawashima, T., Amano, N., Koike, H., Makino, S.-i.,
Higuchi, S., Kawashima-Ohya, Y., Watanabe, K., Yamazaki, M., Kanehori, K., Kawamoto, T.,
Nunoshiba, T., Yamamoto, Y., Aramaki, H., Makino, K., Suzuki, M.

<>

<1>Kawashima, T., Yamamoto, Y., Aramaki, H., Nunoshiba, T., Kawamoto, T., Watanabe, K., Yamazaki, M., Kanehori, K., Amano, N., Ohya, Y., Makino, K., Suzuki, M.
<2>Determination of the complete genomic DNA sequence of Thermoplasma volcanium GSS1.
<3>Proc. Jpn. Acad.
<4>75
<5>213-218
<6>1999
<7>The complete genomic DNA sequence of the aero/anaero-facultative archaebacterium, Thermoplasma
volcanium GSS1, has been determined.  A number of DNA fragments were cloned by using the
phage, cosmid, and BAC systems, and sequenced.  The remaining 30 gaps were bridged by DNA
fragments constructed using the polymerase chain reaction.  The repetition in sequencing the
same base positions was 13.1 +/- 7.5 fold.  The alignment of the DNA fragments and the
completeness of the genomic sequence were confirmed by the consistency of the genomic sequence
with the lengths and partial sequences of a second set of DNA fragments that altogether
covered 88% of the genome.  The number of bases found in the genomic sequence is 1,584,799,
with a G/C content of 39.9%.  The combination of the four types of bases in the new genomic
sequence is compared with those in known genomic sequences of similar sizes.

<>

<1>Kawata, Y., Kawasaki, K., Shigeri, Y.
<2>Draft Genome Sequence of Halomonas sp. Strain KM-1, a Moderately Halophilic Bacterium That Produces the Bioplastic Poly(3-Hydroxybutyrate).
<3>J. Bacteriol.
<4>194
<5>2738-2739
<6>2012
<7>We report the draft genome sequence of Halomonas sp. strain KM-1, which was isolated in Ikeda
City, Osaka, Japan, and which produces the bioplastic
poly(3-hydroxybutyrate). The total length of the assembled genome is 4,992,811
bp, and 4,220 coding sequences were predicted within the genome. Genes encoding
proteins that are involved in the production and depolymerization of
poly(3-hydroxybutyrate) were identified. The identification of these genes might
be of use in the production of the bioplastic poly(3-hydroxybutyrate) and its
monomer 3-hydroxybutyrate.

<>

<1>Kay, E., Wren, B.W., Parkhill, J.
<2>Identification of genetic differences between strains of Campylobacter jejuni using differential genomic hybridisation and comparative  genomics.
<3>Int. J. Med. Microbiol.
<4>293
<5>130-131
<6>2003
<7>Campylobacter jejuni is the most frequently identified bacterial causative agent of diarrhoeal
disease in humans worldwide.  Strain diversity and variable host response lead to a spectrum
of outcomes from infection ranging from asymptomatic to severe inflammatory diarrhoea.  The
mode of transmission to humans is unclear but strains have been isolated from a variety of
environmental sources including poultry and water.  It is hypothesised that novel genes within
different strains may relate to different clinical outcomes for ability to survive in varied
environmental niches.  In order to test this hypothesis DNA from the sequenced strain
NCTC11168 was hybridised to whole genome macro-arrays of different strains of C. jejuni with
different characteristics to identify regions of DNA not presnt in the sequenced strain.  The
strain specific DNA was further characterized by sequencing, analysis and comparison to the
sequenced 11168 genome.  Preliminary hybridisation results using strain 81-176 show that this
method of differential hybridisation can be used to detect variable regions of DNA as well as
strain-specific DNA.  To date, 271 sequences unique to 81-176 have been assembled into 58
clusters.  These clusters contain predicted coding sequences with similarity to transporters
and a -glutamyltranspeptidase as well as several with no database similarities.  In addition,
genes with similarity to proteins that are suface associated, proteins involved in the
respiratory chain and restriction modification enzymes were shown to differ between 81-176 and
11168; these are all known to be highly variable between Campylobacter strains.

<>

<1>Kayansamruaj, P., Pirarat, N., Kondo, H., Hirono, I., Rodkhum, C.
<2>Draft Genome Sequences of Streptococcus agalactiae Strains Isolated from Nile Tilapia (Oreochromis niloticus) Farms in Thailand.
<3>Genome Announcements
<4>2
<5>e01300-14
<6>2014
<7>During 2009-2011, two clinical and one environmental strains of Streptococcus agalactiae were
isolated from Nile tilapia (Oreochromis niloticus) farms in
Thailand. Draft genome sequences of two clinical isolates comprise 2,048,343 and
2,105,006 bp, while environmental isolates comprise 2,097,115 bp, having 1,573 to
1,578 coding sequences, respectively.

<>

<1>Kazennova, E.V., Tarasov, A.P., Mileikovskaya, M.M., Semina, I.E., Tsvetkova, N.V.
<2>Isolation and purification of Bordetella pertussis restriction endonuclease BpeI.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>0
<5>56-57
<6>1982
<7>New restriction endonuclease has been isolated from Bordetella pertussis
vaccine strain 305 and purified in 1 stage on Sepharose covalently bound with
blue dextran.  The isolated restrictase has been found capable of breaking down
lambda-phage DNA into 7 fragments.  According to its specificity, BpeI is the
isoschizomer of HindIII obtained from Haemophilus influenzae strain Rd.

<>

<1>Kazlauskiene, R., Maneliene, Z., Butkus, V., Petrusyte, M., Janulaitis, A.
<2>A new specific endodeoxyribonuclease from Escherichia coli RFL 72.
<3>Bioorg. Khim.
<4>12
<5>836-838
<6>1986
<7>A restriction endonuclease Eco72I with a novel substrate specificity has been isolated from
Escherichia coli strain RFL 72.  The enzyme recognizes 5'CAG^GTG 3' 3'GTG^CAC 5'
hexanucleotide palindromic sequence and cleaves it, as indicated by the arrows, to produce
blunt ended fragments.

<>

<1>Kazrani, A.A., Kowalska, M., Czapinska, H., Bochtler, M.
<2>Crystal structure of the 5hmC specific endonuclease PvuRts1I.
<3>Nucleic Acids Res.
<4>42
<5>5929-5936
<6>2014
<7>PvuRts1I is a prototype for a larger family of restriction endonucleases that cleave DNA
containing 5-hydroxymethylcytosine (5hmC) or 5-glucosylhydroxymethylcytosine (5ghmC), but not
5-methylcytosine (5mC) or cytosine. Here, we report a crystal structure of the enzyme at 2.35
A resolution. Although the protein has been crystallized in the absence of DNA, the structure
is very informative. It shows that PvuRts1I consists of an N-terminal, atypical PD-(D/E)XK
catalytic domain and a C-terminal SRA domain that might accommodate a flipped 5hmC or 5ghmC
base. Changes to predicted catalytic residues of the PD-(D/E)XK domain or to the putative
pocket for a flipped base abolish catalytic activity. Surprisingly, fluorescence changes
indicative of base flipping are not observed when PvuRts1I is added to DNA substrates
containing pyrrolocytosine in place of 5hmC (5ghmC). Despite this caveat, the structure
suggests a model for PvuRts1I activity and presents opportunities for protein engineering to
alter the enzyme properties for biotechnological applications.

<>

<1>Ke, Y., Wang, Y., Yuan, X., Xu, J., Zhen, Q., Wang, Z., Li, T., Wang, D., Huang, L., Song, H., Chen, Z.
<2>Complete Genome Sequence of Brucella suis Field Strain BCB025 of Sequence Type ST22.
<3>J. Bacteriol.
<4>194
<5>6959
<6>2012
<7>Brucella is a genus of relatively conservative pathogenic bacteria. Brucella suis is the most
diversified Brucella species. Strains of B. suis belong to different
sequence types. Here, we report the genome sequence of B. suis strain BCB025, one
isolate of the sequence type 22 epidemic in China.

<>

<1>Ke, Y., Yuan, X., Wang, Y., Bai, Y., Xu, J., Song, H., Huang, L., Chen, Z.
<2>Genome Sequences of Brucella melitensis 16M and Its Two Derivatives 16M1w and 16M13w, Which Evolved In Vivo.
<3>J. Bacteriol.
<4>194
<5>5489
<6>2012
<7>Brucella melitensis is an intracellular pathogen that induces chronic infection in humans.
Here, we report the genome sequences of 16M and its two derivatives,
16M1w and 16M13w, which were allowed to adapt in vivo for 1 and 13 weeks,
respectively. Our findings contribute to the investigation of adaptive mutations
and mechanisms of chronic infection by B. melitensis.

<>

<1>Ke, Y., Yuan, X., Zhen, Q., Wang, Y., Li, T., Sun, Y., Song, H., Huang, L., Wang, D., Cui, B., Mao, K., Chen, Z.
<2>Genome Sequence of Brucella melitensis S66, an Isolate of Sequence Type 8, Prevalent in China.
<3>J. Bacteriol.
<4>194
<5>5451
<6>2012
<7>Brucella melitensis is the most-represented Brucella species causing human brucellosis in
China. Here we report the complete genome sequence of B.
melitensis strain S66, a representative strain of sequence type 8 (ST8), which is
prevalent in China, making it possible to compare the genome sequences of
isolates from different countries.

<>

<1>Ke, Y., Zhen, Q., Li, T., Wang, Y., Yuan, X., Xu, J., Huang, L., Wang, D., Song, H., Chen, Z.
<2>Complete Genome Sequence of Brucella melitensis 133, an Isolate of Biovar 1 of Sequence Type 32.
<3>J. Bacteriol.
<4>194
<5>6932
<6>2012
<7>Brucellosis is highly epidemic in China. Of the six classical species, Brucella melitensis and
biovar 1 are the most represented species and biovar that cause
human brucellosis in China. Here, we report the genome sequence of Brucella
melitensis strain 133, a strain of biovar 1 of sequence type 32.

<>

<1>Keatch, S.A., Leonard, P.G., Ladbury, J.E., Dryden, D.T.
<2>StpA protein from Escherichia coli condenses supercoiled DNA in preference to linear DNA and protects it from digestion by DNase I and EcoKI.
<3>Nucleic Acids Res.
<4>33
<5>6540-6546
<6>2005
<7>The nucleoid-associated protein, StpA, of Escherichia coli binds non-specifically to
double-stranded DNA (dsDNA) and apparently forms
bridges between adjacent segments of the DNA. Such a coating of protein on
the DNA would be expected to hinder the action of nucleases. We
demonstrate that StpA binding hinders dsDNA cleavage by both the
non-specific endonuclease, DNase I, and by the site-specific type I
restriction endonuclease, EcoKI. It requires approximately one StpA
molecule per 250-300 bp of supercoiled DNA and approximately one StpA
molecule per 60-100 bp on linear DNA for strong inhibition of the
nucleases. These results support the role of StpA as a
nucleoid-structuring protein which binds DNA segments together. The
inhibition of EcoKI, which cleaves DNA at a site remote from its initial
target sequence after extensive DNA translocation driven by ATP
hydrolysis, suggests that these enzymes would be unable to function on
chromosomal DNA even during times of DNA damage when potentially lethal,
unmodified target sites occur on the chromosome. This supports a role for
nucleoid-associated proteins in restriction alleviation during times of
cell stress.

<>

<1>Keatch, S.A., Su, T.J., Dryden, D.T.
<2>Alleviation of restriction by DNA condensation and non-specific DNA binding ligands.
<3>Nucleic Acids Res.
<4>32
<5>5841-5850
<6>2004
<7>During conditions of cell stress, the type I restriction and modification enzymes of bacteria
show reduced, but not zero, levels of restriction of
unmethylated foreign DNA. In such conditions, chemically identical
unmethylated recognition sequences also occur on the chromosome of the
host but restriction alleviation prevents the enzymes from destroying the
host DNA. How is this distinction between chemically identical DNA
molecules achieved? For some, but not all, type I restriction enzymes,
alleviation is partially due to proteolytic degradation of a subunit of
the enzyme. We identify that the additional alleviation factor is
attributable to the structural difference between foreign DNA entering the
cell as a random coil and host DNA, which exists in a condensed nucleoid
structure coated with many non-specific ligands. The type I restriction
enzyme is able to destroy the 'naked' DNA using a complex reaction linked
to DNA translocation, but this essential translocation process is
inhibited by DNA condensation and the presence of non-specific ligands
bound along the DNA.

<>

<1>Keeble, A.H., Mate, M.J., Kleanthous, C.
<2>HNH Endonucleases.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Berlin Heidelberg
<4>16
<5>49-65
<6>2005
<7>HNH endonucleases cleave phosphodiester bonds in many biological contexts, including intron
homing, degradation of genomic DNA, repair of genomic DNA, and restriction of viral DNA.  In
the 10 years since the HNH motif was first reported, around 500 members have been identified,
which have the following database identifiers: cd00085, SM00507 and pfam01844.  HNH enzymes
are found in all biological kingdoms, encoded by group I and group II introns, inteins, as
well as free-standing open reading frames.  In the first part of this chapter, we describe the
consensus sequence subsets that are associated with HNH enzymes and the proteins that
accommodate them.  We then present the biochemical properties of HNH enzymes and the proteins
that accommodate them.  We then present the biochemical properties of HNH enzymes as a group,
and finally describe recent structural data on HNH enzymes bound to DNA highlighting plausible
cleavage mechanisms.  Throughout, we describe how HNH enzymes are part of a wider group of
enzymes generally referred to as beta beta alpha-Me or His-Me endonucleases that also includes
His-Cys homing endonucleases and the eukaryotic apoptotic enzyme caspase-activated DNase.

<>

<1>Keeling, P.J., Roger, A.J.
<2>The selfish pursuit of sex.
<3>Nature
<4>375
<5>283
<6>1995
<7>A letter arguing that inteins are selfish elements relevant to the evolution of sex.

<>

<1>Kehrl, J.H., Patel, K.B., Kahng, L.S.
<2>Dam methylation is essential in vibrio vulnificus and influences its interactions with host cells.
<3>Gastroenterology
<4>134
<5>A713
<6>2008
<7>DNA methylation is an important epigenetic regulator in bacteria, and several pathogens
display altered virulence when their methylating
enzymes are knocked out or overexpressed. We hypothesized that
methylation by the Dam (DNA adenine methylase) enzyme plays key roles
in Vibrio vulnificus, a leading cause of lethal seafood-borne
infections. Our arms were to determine the consequences of altered Dam
levels on V. vulnificus gene expression and its interactions with host
cells, specifically its effects on infection-induced barrier function
alteration and inflammation. We sought to knock out Dam, but
chromosomal dam could only be disrupted in the presence of a
plasmid-borne copy of the gene. Thus, dam is essential for the survival
of this pathogen and so we overexpressed it using the arabinose
promoter. We then performed 2-D gel analysis of lysates and culture
supernatants, and found that a discrete subset of proteins was indeed
up- or downregulated under conditions of dam overexpression. To
determine whether these changes in protein levels resulted in altered
bacterial interactions with host cells, we assayed epithelial barrier
function and the inflammatory response. To study barrier function, we
characterized the effects of V. vulnificus on the transepithelial
electrical resistance (TER) of Caco-2 cell monolayers. Wild-type V.
vulnificus were applied to the apical surface of Caco-2 cells grown on
permeable filters, using plain medium as a negative control. We
observed a reproducible drop in TER of the infected monolayer, with a
decline of 53+/-6% after two hours, when lactate dehydrogenase assays
determined that cytotoxicity was not significantly different from
control. TER did not drop when supernatants were used or bacteria were
treated with chloramphenicol; thus, impairment of barrier function by
V. vulnificus requires new bacterial protein synthesis. We then studied
the effects of dam-overexpressing and control strains on TER, after
confirming that they grew at the same rate. The dam-overexpressing
strain yielded a delayed decrease in TER (decline of 15 +/- 3% at 2 h
and 52 +/- 5% at four hours) compared to the control strain. To study
the inflammatory response to V. vulnificus, we assayed secretion of the
pro-inflammatory cytokine, IL-8, by Caco-2 cells. incubation of
wild-type bacteria with the cells led to a reproducible, time-dependent
induction of IL-8 that was not dependent on flagella and was
consistently increased by 50% under conditions of dam overexpression.
In summary, we are the first to demonstrate that dam methylation is not
only essential for the viability of V. vulnificus but may influence its
interactions with host cells.

<>

<1>Keiichi, H., Teruyo, I., Akira, A., Hirotaka, O., Tsukasa, H.
<2>Identification method.
<3>Japanese Patent Office
<4>JP 1999056371 A
<5>
<6>1999
<7>
<>

<1>Keita, M.B., Diene, S.M., Robert, C., Raoult, D., Fournier, P.E., Bittar, F.
<2>Non-contiguous finished genome sequence and description of Bacillus massiliogorillae sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>93-105
<6>2013
<7>Strain G2(T) sp. nov. is the type strain of B. massiliogorillae, a proposed new species within
the genus Bacillus. This strain, whose genome is described here,
was isolated in France from the fecal sample of a wild western lowland gorilla
from Cameroon. B. massiliogorillae is a facultative anaerobic, Gram-variable,
rod-shaped bacterium. Here we describe the features of this organism, together
with the complete genome sequence and annotation. The 5,431,633 bp long genome (1
chromosome but no plasmid) contains 5,179 protein-coding and 98 RNA genes,
including 91 tRNA genes.

<>

<1>Keita, M.B., Padhmanabhan, R., Caputo, A., Robert, C., Delaporte, E., Raoult, D., Fournier, P.E., Bittar, F.
<2>Non-contiguous finished genome sequence and description of Gorillibacterium massiliense gen. nov, sp. nov., a new member of the family Paenibacillaceae.
<3>Standards in Genomic Sciences
<4>9
<5>807-820
<6>2014
<7>Strain G5(T) gen. nov., sp. nov. is the type strain of Gorillibacterium massiliense, a newly
proposed genus within the family Paenibacillaceae. This
strain, whose genome is described here, was isolated in France from a stool
sample of a wild Gorilla gorilla subsp. gorilla from Cameroon. G. massiliense is
a facultatively anaerobic, Gram negative rod. Here we describe the features of
this bacterium, together with the complete genome sequence and annotation. The
5,546,433 bp long genome (1 chromosome but no plasmid) contains 5,145
protein-coding and 76 RNA genes, including 69 tRNA genes.

<>

<1>Keita, M.B., Padhmananabhan, R., Caputo, A., Robert, C., Delaporte, E., Raoult, D., Fournier, P.E., Bittar, F.
<2>Non-contiguous finished genome sequence and description of Paenibacillus gorillae sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1031-1045
<6>2014
<7>Strain G1(T) sp. nov. is the type strain of Paenibacillus gorillae a newly proposed species
within the genus Paenibacillus. This strain, whose genome is
described here, was isolated in France from the fecal sample of a wild western
lowland gorilla from Cameroon. P. gorillae is a facultative anaerobic,
Gram-negative, rod-shaped bacterium. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
6,257,967 bp long genome (one chromosome but no plasmid) contains 5,856
protein-coding and 62 RNAs genes, including 60 tRNA genes.

<>

<1>Keith, T., Little, R.D., Eerdewegh, P.V., Dupuis, J., Del Mastro, R.G., Simon, J., Allen, K., Pandit, S.
<2>Nucleotide and amino acid sequences relating to respiratory diseases and obesity.
<3>US Patent Office
<4>US 7205146 A
<5>
<6>2007
<7>This invention relates to genes identified from human chromosome 12q23-qter, which are
associated with various diseases, including asthma.  The invention also relates to the
nucleotide sequences of these genes, isolated nucleic acids comprising these nucleotide
sequences, and isolated polypeptides or peptides encoded thereby.  The invention further
relates to vectors and host cells comprising the disclosed nucleotide sequences, or fragments
thereof, as well as antibodies that bind to the encoded polypeptides or peptides.  Also
related are ligands that modulate the activity of the disclosed genes or gene products.  In
addition, the invention relates to methods and compositions employing the disclosed nucleic
acids, polypeptides or peptides, antibodies, and/or ligands for use in diagnostics and
therapeutics for asthma and other diseases.

<>

<1>Kejnovsky, E., Nejedly, K., Kypr, J.
<2>Factors influencing resistance of UV-irradiated DNA to the restriction endonuclease cleavage.
<3>Int. J. Biol. Macromol.
<4>34
<5>213-222
<6>2004
<7>DNA molecules of pUC19, pBR322 and PhiX174 were irradiated by various doses of UV light and
the irradiated molecules were cleaved by about
two dozen type II restrictases. The irradiation generally blocked the
cleavage in a dose-dependent way. In accordance with previous studies,
the (A + T)-richness and the (PyPy) dimer content of the restriction
site belongs among the factors that on average, cause an increase in
the resistance of UV damaged DNA to the restrictase cleavage. However,
we observed strong effects of UV irradiation even with (G + C)-rich and
(PyPy)-poor sites. In addition, sequences flanking the restriction site
influenced the protection in some cases (e.g. HindIII), but not in
others (e.g. SalI), whereas neoschizomer couples SmaI and AvaI, or SacI
and Ecl136II, cleaved the UV-irradiated DNA similarly. Hence the
intrastrand thymine dimers located in the recognition site are not the
only photoproduct blocking the restrictases. UV irradiation of the
A-form generally made the irradiated DNA less resistant to restrictase
cleavage than irradiation in the B-form and in some cases, the A-form
completely protected the UV-irradiated DNA against the damage
recognized by the restrictases. The present results also demonstrate
that the UV irradiation approach used to generate partial digests in
genomic DNA studies, can be extended to the (G + C)-rich and
(PyPy)-poor restriction sites. The present extensive and quantitative
data can be used in genomic applications of UV damage probing by
restrictases.

<>

<1>Kelleher, J.E.
<2>Defining domains of the EcoK methylase by mutational analyses and DNA sequence comparisons.
<3>Ph.D. Thesis, University of Edinburgh
<4>
<5>1-207
<6>1990
<7>EcoK is a type I restriction and modification enzyme. It recognizes a defined DNA sequence and
may methylate specific adenine residues within this target site, on each stand of the DNA.
EcoK is composed of three different subunits encoded by the hsdR, M and S genes of E. coli
K-12. The S subunit dictates the recognition specificity and together with M forms the
methylase. All three enzyme subunits are necessary to form the restriction complex. Although
this is also capable of DNA modification its activity is selected in reponse to the presence
or absence of target site methylation. The methylase too is sensitive to the methylation state
of the target site. Efficient modification requires the imprint of a methyl group in the
complementary strand of the recognition sequence, whilst unmodified DNA, the normal substrate
for restriction, is only methylated inefficiently. (Suri et al., 1984). The experiments
described aim to identify functional domains of the EcoK methylase, from either the
comparative analyses of polypeptide sequences, or the phenotypes of mutations. Related type I
enzymes can interchange their subunits; unrelated enzymes cannot, and no general similarities
are detected when unrelated polypeptides are compared (see Bickle, 1987). Nevertheless, the
subunits of unrelated enzymes are presumed to be functionally analogous and sequence motifs
common to all type I enzymes, may identify polypeptide domains involved in common activities.
Mutational analyses have been limited to mutant derivatives of the EcoK methylase specifically
defective in only a particular aspect of their activity, and have focused on mutations that
apparently alter the substrate specificity of EcoK. The phenotypes of the new mutations
identified are consistent with enzymes that fail to differentiate between hemimethylated and
unmethylated target sites. These mutations cluster in a region of the hsdM gene, and may
identify a polypeptide domain responsible for recognition of target site methylation.

<>

<1>Kelleher, J.E., Daniel, A.S., Murray, N.E.
<2>Mutations that confer de novo activity upon a maintenance methyltransferase.
<3>J. Mol. Biol.
<4>221
<5>431-440
<6>1991
<7>DNA methyltransferases are not only sequence specific in their action, but they
also differentiate between the alternative methylation states of a target site.
Some methyltransferases are equally active on either unmethylated or
hemimethylated DNA and consequently function as de novo methyltransferases.
Others are specific for hemimethylated target sequences, consistent with the
postulated role of a maintenance methyltransferase in perpetuating a pattern of
DNA modification.  The molecular basis for the difference between de novo and
maintenance methyltransferase activity is unknown, yet fundamental to cellular
activities that are affected by different methylation states of the genome.
The methyltransferase activity of the type I restriction and modification
system, EcoK, is the only known prokaryotic methyltransferase shown to be
specific for hemimethylated target sequences.  We have isolated mutants of
Escherichia coli K-12 which are able to modify unmethylated target sequences
efficiently in a manner indicative of de novo methyltransferase activity.
Consistent with this change in specificity, some mutations shift the balance
between DNA restriction and modification as if both activities now compete at
unmethylated targets.  Two genes encode the methyltransferase and all the
mutations are loosely clustered within one of them.

<>

<1>Kelleher, J.E., Raleigh, E.A.
<2>A novel activity in Escherichia coli K-12 that directs restriction of DNA modified at CG dinucleotides.
<3>J. Bacteriol.
<4>173
<5>5220-5223
<6>1991
<7>The restriction systems McrA and McrB of Escherichia coli K-12 are known to
attack DNA containing modified cytosine.  In strains lacking both activities,
however, we observed that DNA methylated at CG dinucleotides (as is mammalian
DNA) was still significantly restricted.  We show that this substantial barrier
to the acceptance of 5-methylcytosine-containing DNA is attributable to a
hitherto unknown activity of the Mrr restriction system.  Strikingly, the
multiple systems used by the gut inhabitant to determine the fate of invading
DNA will all limit genetic exchange with its mammalian host(s), reinforcing the
idea that one role of DNA methylation is to serve as a molecular passport (E.A.
Raleigh, R. Trimarchi, and H. Revel, Genetics 122:279-296, 1989).

<>

<1>Kelleher, J.E., Raleigh, E.A.
<2>Response to UV damage by four Escherichia coli K-12 restriction systems.
<3>J. Bacteriol.
<4>176
<5>5888-5896
<6>1994
<7>To understand the role of restriction in regulating gene flow in bacterial populations, we
would like to understand the regulation of restriction enzyme activity. Several
antirestriction (restriction alleviation) systems are known that reduce the activity of type I
restriction enzymes like EcoKI in vivo. Most of these do not act on type II or type III
enzymes, but little information is available for the unclassified modification-dependent
systems, of which there are three in E. coli K-12. Of particular interest are two
physiological controls on type I enzymes: EcoKI restriction is reduced 2 to 3 orders of
magnitude following DNA damage, and a similar effect is seen constitutively in Dam- cells. We
used the behavior of EcoKI as a control for testing the response to UV treatment of the three
endogenous modification-dependent restriction systems of K-12, McrA, McrBC, and Mrr. Two of
these were also tested for response to Dam status. We find that all four resident restriction
systems show reduced activity following UV treatment, but not in a unified fashion; each
response was genetically and physiologically distinct. Possible mechanisms are discussed.

<>

<1>Kelleher, J.E., Raleigh, E.A.
<2>On the regulation and diversity of restriction in Escherichia coli.
<3>Gene
<4>157
<5>229-230
<6>1995
<7>The effect of UV irradiation on restriction mediated by four endogenous restriction systems of
E. coli K-12 was investigated using a uniform testing method.  Restriction by all four
systems was reduced when treated cells were separately challenged with lambda phage carrying
modification patterns that elicit restriction by each system.  The response of each system was
genetically and physiologically distinct.

<>

<1>Kellenberger, G., Symonds, N., Arber, W.
<2>Host specificity of DNA produced by Escherichia coli.  8.Its acquisition by phage lambda and its persistence through consecutive growth cycles.
<3>Z. Vererbungsl.
<4>98
<5>247-256
<6>1966
<7>Earlier papers in this series have demonstrated that host-controlled modification of
bacteriophage lambda consists of the donation of a specific label to the phage DNA by the
bacterial host cells.  The experiments to be described in this paper were originally designed
to extend these observations to other types of host specificity, in particular to that
produced by E. coli B.  Technically, such experiments were facilitated by the availability of
restrictionless (r-) bacterial mutants which still provide full modification (m+): Density
labelled lambda K was grown in normal medium for one cycle on a Br-m+ strain and the progeny
phages analyzes for their parental DNA content by density gradient centrifugation.  The
density fractions were then tested for both K and B host specificities.  Phage particles with
fully heavy, hence conserved, DNA molecules had both K- and B-specificity.  The fractions of
phages with densities corresponding to particles with halfheavy, supposedly semiconserved DNA
molecules also plated on both K and B indicator.  But a more careful analysis revealed that
only about half of such phages still grew on K, whereas the other half was degraded upon
infection of K bacteria.  Repeated consecutive growth cycles were also performed and allowed
firstly to confirm the findings of Arber, Hattman and Dussoix (1963) that under normal
conditions both strands of lambda DNA are carrying host specificity, and secondly to conclude
that the reduced probability of acceptance by strain K of lambda.K.B phage with semiconserved
DNA cannot find its explanation in the physico-chemical plarity of the K-modified strand.

<>

<1>Keller, K.L., Bender, K.S., Wall, J.D.
<2>A New Counterselectable Marker for Desulfovibrio vulgaris, the upp gene, Allowed for the Construction of a Marker less Deletion of a Type 1 Restriction Enzyme that Exhibits Increased Transformation Efficiency.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>109
<5>0
<6>2009
<7>In recent years, the genetic manipulation of the sulfate-reducing bacterium Desulfovibrio
vulgaris Hildenborough has seen enormous progress; however, the current method of deletion
construction via marker exchange mutagenesis does not allow for easy selection of multiple
sequential gene deletions because of the need for multiple selectable markers. To broaden the
repertoire of genetic tools for manipulation of D. vulgaris, an in-frame markerless deletion
system has been developed based on the upp-encoded uracil phosphoribosyltransferase as an
element for a counterselection strategy. In wild-type D. vulgaris, growth is inhibited by the
toxic pyrimidine analog 5-fluorouracil (5-FU), whereas a mutant bearing a deletion of the upp
gene, strain JW710, is resistant to 5-FU. The introduction of a plasmid containing the
wild-type upp gene expressed constitutively from the aph(5')-III promoter (the promoter for
the kanamycin resistance gene in Tn5) into JW710 restored sensitivity to 5-FU to wild-type
levels. This observation is the basis for the establishment of a two-step integration and
excision strategy for deleting genes of interest. Since this in-frame deletion does not leave
behind an antibiotic cassette, multiple gene deletions can be generated in a single strain.
With this method, a markerless deletion of the R-subunit (DVU1703) of a type I
restriction-modification system (hsdR), strain JW7035, was constructed. The transformation
efficiency of the JW7035 strain is greater (an approximate 2-log increase in transformants)
compared to wild-type DvH when transforming stable plasmids via electroporation.

<>

<1>Keller, K.L., Bender, K.S., Wall, J.D.
<2>Development of a markerless genetic exchange system for Desulfovibrio vulgaris Hildenborough and its use in generating a strain with increased  transformation efficiency.
<3>Appl. Environ. Microbiol.
<4>75
<5>7682-7691
<6>2009
<7>In recent years, the genetic manipulation of the sulfate-reducing bacterium Desulfovibrio
vulgaris Hildenborough has seen enormous progress.
In spite of this progress, the current marker exchange deletion method
does not allow for easy selection of multiple sequential gene deletions in
a single strain because of the limited number of selectable markers
available in D. vulgaris. To broaden the repertoire of genetic tools for
manipulation, an in-frame, markerless deletion system has been developed.
The counterselectable marker that makes this deletion system possible is
the pyrimidine salvage enzyme, uracil phosphoribosyltransferase, encoded
by upp. In wild-type D. vulgaris, growth was shown to be inhibited by the
toxic pyrimidine analog 5-fluorouracil (5-FU), whereas a mutant bearing a
deletion of the upp gene was resistant to 5-FU. When a plasmid containing
the wild-type upp gene expressed constitutively from the aph(3')-II
promoter (promoter for the kanamycin resistance gene in Tn5) was
introduced into the upp deletion strain, sensitivity to 5-FU was restored.
This observation allowed us to develop a two-step integration and excision
strategy for the deletion of genes of interest. Since this in-frame
deletion strategy does not retain an antibiotic cassette, multiple
deletions can be generated in a single strain without the accumulation of
genes conferring antibiotic resistances. We used this strategy to generate
a deletion strain lacking the endonuclease (hsdR, DVU1703) of a type I
restriction-modification system that we designated JW7035. The
transformation efficiency of the JW7035 strain was found to be 100 to
1,000 times greater than that of the wild-type strain when stable plasmids
were introduced via electroporation.

<>

<1>Keller, L.E., Thomas, J.C., Luo, X., Nahm, M.H., McDaniel, L.S., Robinson, D.A.
<2>Draft Genome Sequences of Five Multilocus Sequence Types of Nonencapsulated Streptococcus pneumoniae.
<3>Genome Announcements
<4>1
<5>e00520-13
<6>2013
<7>Nonencapsulated Streptococcus pneumoniae can colonize the human nasopharynx and cause
conjunctivitis and otitis media. Different deletions in the capsular
polysaccharide biosynthesis locus and different multilocus sequence types have
been described for nonencapsulated strains. Draft genome sequences were generated
to provide insight into the genomic diversity of these strains.

<>

<1>Kelly, S., Kaddurah-Daouk, R., Smith, H.O.
<2>Purification of the HhaII restriction endonuclease from an overproducer Escherichia coli clone.
<3>J. Biol. Chem.
<4>260
<5>15339-15344
<6>1985
<7>An Escherichia coli K12 strain carrying the HhaII methylase and restriction
genes on two separate compatible plasmids, pSK5 and pSK7, is used to
overproduce the restriction endonuclease.  Plasmid pSK5 expresses the methylase
gene constitutively from its chloramphenicol resistance gene promoter, and
plasmid pSK7 expresses the restriction endonuclease under control of the lacUV5
promoter.  Induction of the two-plasmid clone with 1 mM
isopropyl-1-thio-b-D-galactopyranoside results in a 15-fold increase in HhaII
endonuclease activity.  The enzyme has been purified to apparent homogeneity.
It migrates as a 23-kilodalton polypeptide on denaturing sodium dodecyl
sulfate-polyacrylamide electrophoretic gels and as a 51-kilodalton native
protein dimer on a high pressure liquid chromatogrpahy sizing column.

<>

<1>Kelly, T.J. Jr., Smith, H.O.
<2>A restriction enzyme from Hemophilus influenzae II.  Base sequence of the recognition site.
<3>J. Mol. Biol.
<4>51
<5>393-409
<6>1970
<7>Hemophilus influenzae strain Rd contains an enzyme, endonuclease R, which
specifically degrades foreign DNA.  With phage T7 DNA as substrate the
endonuclease introduces a limited number (about 40) double-strand breaks
(5'-phosphoryl, 3'-hydroxyl).  The limit product has an average length of about
1000 nucleotide pairs and contains no single-strand breaks.  We have explored
the nucleotide sequences at the 5'-ends of the limit product by labeling the
5'-phosphoryl groups (using polynucleotide kinase) and characterizing the
labeled fragments released by various nucleases.  Two classes of 5'-terminal
sequences were obtained: pApApCpNp...(60%) and pGpApCpNp...(40%), where N
indicates that the base in the 4th position is not unique.  The dinucleoside
monophosphates at the 3'-ends were isolated after micrococcal nuclease
digestion of the limit product and identified as TpT(60%) and TpC(40%).  We
conclude that endonuclease R of H. influenzae recognizes the following specific
nucleotide sequence: 5' . . . pGpTpPy^pPupApCp . . . 3' 3' . . .
pCpApPup^PypTpGp . . . 5' The implications of the twofold rotational symmetry
of this sequence are discussed.

<>

<1>Kelly, T.J., Smith, H.O.
<2>The nucleotide sequence of the recognition site for a restriction enzyme from H. influenzae.
<3>Fed. Proc.
<4>29
<5>405
<6>1970
<7>H. influenzae strain Rd contains an endonuclease which specifically degrades
foreign DNA (Smith & Wilcox, Fed. Proc. 28:465, 1969).  With T7 DNA as
substrate the endonuclease introduces a limited number (about 40) of
double-strand breaks (5'-phosphoryl, 3'-hydroxyl).  The limit product has an
average length of about 1000 nucleotide pairs and contains no single-strand
breaks.  We have explored the nucleotide sequences at the 5'-ends of the limit
product by labeling the 5'-phosphoryl groups (using polynucleotide kinase) and
characterising the labeled fragments released by various nucleases.  Two
classes of 5'-sequences were obtained:  pApApCpXp...(60%) and
pGpApCpXp...(40%), where X indicates that the base in the 4th position is not
unique.  The dinucleoside monophosphates at the 3'-ends were isolated after
micrococcal nuclease digestion of the limit product and identified as TpT (60%)
and TpC (40%).  We conclude that the H. influenzae endonuclease recognizes the
following specific nucleotide sequence: 5' ...pGpTpPy^pPupApCp... 3'     ^break
3' ...pCpApPup^PypTpGp... 5' The two-fold rotational symmetry of this sequence
has interesting implications concerning the mechanism of action of the H.
influenzae endonuclease.

<>

<1>Kelly, W.J., Altermann, E., Lambie, S.C., Leahy, S.C.
<2>Interaction between the genomes of Lactococcus lactis and phages of the P335 species.
<3>Front. Microbiol.
<4>4
<5>257
<6>2013
<7>Phages of the P335 species infect Lactococcus lactis and have been particularly
studied because of their association with strains of L. lactis subsp. cremoris
used as dairy starter cultures. Unlike other lactococcal phages, those of the
P335 species may have a temperate or lytic lifestyle, and are believed to
originate from the starter cultures themselves. We have sequenced the genome of
L. lactis subsp. cremoris KW2 isolated from fermented corn and found that it
contains an integrated P335 species prophage. This 41 kb prophage (Phi KW2) has a
mosaic structure with functional modules that are highly similar to several other
phages of the P335 species associated with dairy starter cultures. Comparison of
the genomes of 26 phages of the P335 species, with either a lytic or temperate
lifestyle, shows that they can be divided into three groups and that the
morphogenesis gene region is the most conserved. Analysis of these phage genomes
in conjunction with the genomes of several L. lactis strains shows that prophage
insertion is site specific and occurs at seven different chromosomal locations.
Exactly how induced or lytic phages of the P335 species interact with
carbohydrate cell surface receptors in the host cell envelope remains to be
determined. Genes for the biosynthesis of a variable cell surface polysaccharide
and for lipoteichoic acids (LTAs) are found in L. lactis and are the main
candidates for phage receptors, as the genes for other cell surface carbohydrates
have been lost from dairy starter strains. Overall, phages of the P335 species
appear to have had only a minor role in the adaptation of L. lactis subsp.
cremoris strains to the dairy environment, and instead they appear to be an
integral part of the L. lactis chromosome. There remains a great deal to be
discovered about their role, and their contribution to the evolution of the
bacterial genome.

<>

<1>Kelly, W.J., Leahy, S.C., Altermann, E., Yeoman, C.J., Dunne, J.C., Kong, Z., Pacheco, D.M., Li, D., Noel, S.J., Moon, C.D., Cookson, A.L., Attwood, G.T.
<2>The Glycobiome of the Rumen Bacterium Butyrivibrio proteoclasticus B316 Highlights Adaptation to a Polysaccharide-Rich Environment.
<3>PLoS ONE
<4>5
<5>E11942
<6>2010
<7>Determining the role of rumen microbes and their enzymes in plant
polysaccharide breakdown is fundamental to understanding digestion and
maximising productivity in ruminant animals. Butyrivibrio proteoclasticus
B316(T) is a Gram-positive, butyrate-forming rumen bacterium with a key
role in plant polysaccharide degradation. The 4.4Mb genome consists of 4
replicons; a chromosome, a chromid and two megaplasmids. The chromid is
the smallest reported for all bacteria, and the first identified from the
phylum Firmicutes. B316 devotes a large proportion of its genome to the
breakdown and reassembly of complex polysaccharides and has a highly
developed glycobiome when compared to other sequenced bacteria. The
secretion of a range of polysaccharide-degrading enzymes which initiate
the breakdown of pectin, starch and xylan, a subtilisin family protease
active against plant proteins, and diverse intracellular enzymes to break
down oligosaccharides constitute the degradative capability of this
organism. A prominent feature of the genome is the presence of multiple
gene clusters predicted to be involved in polysaccharide biosynthesis.
Metabolic reconstruction reveals the absence of an identifiable gene for
enolase, a conserved enzyme of the glycolytic pathway. To our knowledge
this is the first report of an organism lacking an enolase. Our analysis
of the B316 genome shows how one organism can contribute to the
multi-organism complex that rapidly breaks down plant material in the
rumen. It can be concluded that B316, and similar organisms with broad
polysaccharide-degrading capability, are well suited to being early
colonizers and degraders of plant polysaccharides in the rumen
environment.

<>

<1>Kennaway, C.K., Obarska-Kosinska, A., White, J.H., Tuszynska, I., Cooper, L.P., Bujnicki, J.M., Trinick, J., Dryden, D.T.
<2>The structure of M.EcoKI Type I DNA methyltransferase with a DNA mimic antirestriction protein.
<3>Nucleic Acids Res.
<4>37
<5>762-770
<6>2009
<7>Type-I DNA restriction-modification (R/M) systems are important agents in limiting the
transmission of mobile genetic elements responsible for
spreading bacterial resistance to antibiotics. EcoKI, a Type I R/M enzyme
from Escherichia coli, acts by methylation- and sequence-specific
recognition, leading to either methylation of DNA or translocation and
cutting at a random site, often hundreds of base pairs away. Consisting of
one specificity subunit, two modification subunits, and two DNA
translocase/endonuclease subunits, EcoKI is inhibited by the T7 phage
antirestriction protein ocr, a DNA mimic. We present a 3D density map
generated by negative-stain electron microscopy and single particle
analysis of the central core of the restriction complex, the M.EcoKI
M(2)S(1) methyltransferase, bound to ocr. We also present complete atomic
models of M.EcoKI in complex with ocr and its cognate DNA giving a clear
picture of the overall clamp-like operation of the enzyme. The model is
consistent with a large body of experimental data on EcoKI published over
40 years.

<>

<1>Kennaway, C.K., Taylor, J.E., Song, C.F., Potrzebowski, W., Nicholson, W., White, J.H., Swiderska, A., Obarska-Kosinska, A., Callow, P., Cooper, L.P., Roberts, G.A., Artero, J.B., Bujnicki, J.M., Trinick, J., Kneale, G.G., Dryden, D.T.F.
<2>Structure and operation of the DNA-translocating type I DNA restriction enzymes.
<3>Genes Dev.
<4>26
<5>92-104
<6>2012
<7>Type I DNA restriction/modification (RM) enzymes are molecular machines found in the majority
of bacterial species. Their early discovery paved
the way for the development of genetic engineering. They control
(restrict) the influx of foreign DNA via horizontal gene transfer into
the bacterium while maintaining sequence-specific methylation
(modification) of host DNA. The endonuclease reaction of these enzymes
on unmethylated DNA is preceded by bidirectional translocation of
thousands of base pairs of DNA toward the enzyme. We present the
structures of two type I RM enzymes, EcoKI and EcoR124I, derived using
electron microscopy (EM), small-angle scattering (neutron and X-ray),
and detailed molecular modeling. DNA binding triggers a large
contraction of the open form of the enzyme to a compact form. The path
followed by DNA through the complexes is revealed by using a DNA mimic
anti-restriction protein. The structures reveal an evolutionary link
between type I RM enzymes and type II RM enzymes.

<>

<1>Kennell, J.C., Moran, J.V., Perlman, P.S., Butow, R.A., Lambowitz, A.M.
<2>Reverse transcriptase activity associated with maturase-encoding group II introns in yeast mitochondria.
<3>Cell
<4>73
<5>133-146
<6>1993
<7>Group II introns ai1 and ai2 of the yeast mtDNA cox1 gene encode reverse transcriptase-like
proteins that function in RNA splicing and may play a role in intron mobility and excision. We
find that ribonucleoprotein particles from yeast mitochondria contain a reverse transcriptase
activity that is likely encoded by ai1 and ai2 and is highly specific for the introns and
their flanking exons.  Using a mutant strain with elevated activity, we show that the reverse
transcriptase uses either excised intron RNA or cox1 pre-mRNA as template and initiates cDNA
synthesis near the 3' end of ai2 and immediately downstream in E3.  Our results suggest that
introns ai1 and ai2 are retroelements, which encode reverse transcriptases that have adapted
to function in RNA splicing.

<>

<1>Kenri, T., Horino, A., Matsui, M., Sasaki, Y., Suzuki, S., Narita, M., Ohya, H., Okazaki, N., Shibayama, K.
<2>Complete Genome Sequence of Mycoplasma pneumoniae Type 2a Strain 309, Isolated in Japan.
<3>J. Bacteriol.
<4>194
<5>1253-1254
<6>2012
<7>Mycoplasma pneumoniae strain 309, a type 2a (subtype 2 variant) strain of this bacterium, has
variations in the P1 protein, which is responsible for attachment
of the bacterium to host cells. Here, we report the complete genome sequence of
M. pneumoniae strain 309 isolated from a pneumonia patient in Japan.

<>

<1>Kenzaka, T., Nakahara, M., Higuchi, S., Maeda, K., Tani, K.
<2>Draft Genome Sequences of Amoeba-Resistant Aeromonas spp. Isolated from Aquatic Environments.
<3>Genome Announcements
<4>2
<5>e01115-14
<6>2014
<7>Amoeba-resistant Aeromonas veronii ARB3 and Aeromonas media ARB13 and ARB20, which may be
important intracellular pathogens of eukaryotic hosts, were isolated
from pond and river waters. The draft genome sequences indicate that the strains
harbor multiple protein secretion systems and toxins that induce disruption of
the actin cytoskeleton.

<>

<1>Kenzaka, T., Yamada, Y., Tani, K.
<2>Draft Genome Sequence of an Antifungal Bacterium Isolated from the Breeding Environment of Dorcus hopei binodulosus.
<3>Genome Announcements
<4>2
<5>e00424-14
<6>2014
<7>Burkholderia sp. strain A1 was isolated from a decaying log present in the breeding
environment of a stag beetle. The draft genome sequence indicates that
strain A1 harbors many biosynthesis molecules, which have antimicrobial
properties, and thus potentially eliminates the fungi by producing antifungal
compounds, such as siderophores.

<>

<1>Kergourlay, G., Messaoudi, S., Dousset, X., Prevost, H.
<2>Genome Sequence of Lactobacillus salivarius SMXD51, a Potential Probiotic Strain  Isolated from Chicken Cecum, Showing Anti-Campylobacter Activity.
<3>J. Bacteriol.
<4>194
<5>3008-3009
<6>2012
<7>We report the draft genome sequence of Lactobacillus salivarius SMXD51, isolated  from the
cecum of healthy chickens showing an activity against Campylobacter-the
food-borne pathogen that is the most common cause of gastroenteritis in the
European Union (EU)-and potentially interesting features for a probiotic strain,
explaining our interest in it.

<>

<1>Kerr, A.L., Jeon, Y.J., Svenson, C.J., Rogers, P.L., Neilan, B.A.
<2>DNA restriction-modification systems in the ethanologen, Zymomonas mobilis ZM4.
<3>Appl. Microbiol. Biotechnol.
<4>89
<5>761-769
<6>2011
<7>To better understand the DNA restriction-modification (R-M) systems for more amenable strain
development of the alternative industrial
ethanologen, Zymomonas mobilis, three gene knockout mutants were
constructed. The gene knockout mutants were tested for their DNA
restriction activities by the determination of transformation
efficiency using methylated and unmethylated foreign plasmid DNAs.
Inactivation of a putative mrr gene encoded by ZMO0028 (zmrr) resulted
in a 60-fold increase in the transformation efficiency when
unmethylated plasmid DNA was used. This indicated that the putative mrr
gene may serve as a type IV restriction-modification system in Z.
mobilis ZM4. To assign the function of a putative type I DNA
methyltransferase encoded by ZMO1933 (putative S subunit) and ZMO1934
(putative M subunit), the putative S subunit was inactivated. The gene
inactivation of ZMO1933 resulted in a 30-fold increase in the
transformation efficiency when methylated plasmid DNA was introduced,
indicating that the putative S subunit possibly serves as a part of
functional type I R-M system(s). Growth studies performed on the mutant
strains indicate inactivation of the type I S subunit resulted in a
lower maximum specific glucose consumption rate and biomass yield,
while inactivation of the type IV Zmrr had the opposite effect, with an
increase in the maximum specific growth rate and biomass yield.

<>

<1>Kersulyte, D., Mukhopadhyay, A.K., Shirai, M., Nakazawa, T., Berg, D.E.
<2>Functional organization and insertion specificity of IS607, a chimeric element of Helicobacter pylori.
<3>J. Bacteriol.
<4>182
<5>5300-5308
<6>2000
<7>A search by subtractive hybridization for sequences present in only
certain strains of Helicobacter pylori led to the discovery of a 2-kb
transposable element to be called IS607, which further PCR and
hybridization tests indicated was present in about one-fifth of H. pylori
strains worldwide. IS607 contained two open reading frames (ORFs) of
possibly different phylogenetic origin. One ORF (orfB) exhibited
protein-level homology to one of two putative transposase genes found in
several other chimeric elements including IS605 (also of H. pylori) and
IS1535 (of Mycobacterium tuberculosis). The second IS607 gene (orfA) was
unrelated to the second gene of IS605 and might possibly be chimeric
itself: it exhibited protein-level homology to merR bacterial regulatory
genes in the first approximately 50 codons and homology to the second gene
of IS1535 (annotated as "resolvase," apparently due to a weak short
recombinase motif) in the remaining three-fourths of its length. IS607 was
found to transpose in Escherichia coli, and analyses of sequences of
IS607-target DNA junctions in H. pylori and E. coli indicated that it
inserted either next to or between adjacent GG nucleotides, and generated
either a 2-bp or a 0-bp target sequence duplication, respectively.
Mutational tests showed that its transposition in E. coli required orfA
but not orfB, suggesting that OrfA protein may represent a new, previously
unrecognized, family of bacterial transposases.

<>

<1>Kerszman, G., Glover, S.W., Aronovitch, J.
<2>The restriction of bacteriophage lambda in Escherichia coli strain W.
<3>J. Gen. Virol.
<4>1
<5>333-347
<6>1967
<7>Escherichia coli strain w adsorbs phage lambda very efficiently but the phage does not form
plaques on this strain.  In a very small fraction (10^-4) of the infected cells the phage
grows and produces small bursts of progeny phage also unable to form plaques on strain w.  E.
coli strain w is lysogenic for a temperate phage, wPhi, related to phage P2.  Non-restricting
hosts for phage lambda became restricting hosts when made lysogenic for wPhi.  When
32P-labelled lambda adsorbed to restricting wPhi lysogenic hosts, >20% of the 32P become
acid-soluble shortly after infection.  No wPhi specific modification was carried by the small
number of lambda phages which escaped this restriction process.  It is concluded that wPhi
controls a host-restriction mechanism but not a host-modification process, and in parallel
with other examples of host-controlled restriction and modification can be represented as r+m-
or r+mo.  LambdaW mutants have been isolated which escape this restriction and which form
plaques on strain w and wPhi lysogenic strains with an efficiency of I.0.  With these mutants
a w-specific host modification controlled by the genome of strain w was demonstrated.  Mixed
infection experiments with restricted lambda and unrestricted lambda w showed that that
restricted phage did not block the growth of the unrestricted mutant nor did the mutant permit
the restricted phage to grow.  In addition it was shown that lambda obtained from bacteria
mixedly infected with lambda and lambdaW was still unable to grow in restricting hosts and
lambda w similarly obtained from mixedly infected bacteria still retained its ability to grow
on restricting hosts.  It is concluded that there is a nucleotide sequence in the DNA of phage
lambda which, when lambda infects a restricting host, is specifically recognized by the
restriction mechanism controlled by the wPhi.  The mutation to lambda w involves an alteration
to this sequence such that it is no longer recognized by the restriction mechanism of the
wPhi.  Mutants of wPhi were isolated not restrictive for phage lambda.

<>

<1>Kerzhner, M.A., Shiryaev, S.A., Zheleznaya, L.A., Matvienko, N.I.
<2>Site-specific endonuclease from thermophilic Bacillus species MK strain is isoschizomer of SalI.
<3>Biokhimiia
<4>62
<5>1029-1036
<6>1997
<7>Screening of thermophilic bacterial strains revealed a strain containing site-specific
endonuclease BspMKI.  Endonuclease was purified to functional homogeneity during sequential
chromatographic steps.  The enzyme recognizes sequences 5'-G/TCGAC-3' on DNA molecule and
its isoschizomer of endonuclease SalI.  The molecular mass of BspMKI is about 45 kD.  The
enzyme is maximally active at 55oC and MRB (50 mM NaCl, 10 mM Tris-HCl, pH 7.4, 10 mM MgCl2, 1
mM dithiothreitol) is the optimal buffer.  The enzyme is highly stable and retains its
activity during two weeks at room temperature.

<>

<1>Kesel, S., Moormann, F., Gumperlein, I., Mader, A., Morikawa, M., Lieleg, O., Opitz, M.
<2>Draft Genome Sequence of the Biofilm-Producing Bacillus subtilis Strain B-1, Isolated from an Oil Field.
<3>Genome Announcements
<4>2
<5>e01163-14
<6>2014
<7>We report here the draft genome sequence of the Bacillus subtilis strain B-1, a strain known
to form biofilms. The biofilm matrix mainly consists of the
biopolymer gamma-polyglutamate (gamma-PGA). The sequence of the genome of this
strain allows the study of specific genes involved in biofilm formation.

<>

<1>Keshet, E., Cedar, H.
<2>Effect of CpG methylation on MspI.
<3>Nucleic Acids Res.
<4>11
<5>3571-3580
<6>1983
<7>The restriction enzyme MspI is inhibited by the presence of a methyl moiety at
the external cytosine of the sequence CCGG, but is generally unaffected by
methylation at the internal cytosine.  At specific subsets of this sequence
such as the hexanucleotide CCGGCC, however, methylation of the internal
cytosine strongly inhibits MspI digestion, leading to artifacts in the
interpretation of DNA methylation analyses.  Our results show, for instance,
that the CCGG site at the 5' end of the human gamma globin gene, which was
thought to be methylated at both the internal and external cytosines, is
actually methylated only at the internal CpG residue.

<>

<1>Keshi, H., Eda, S., Uehara, H., Nishida, K., Matsuhisa, A.
<2>Probes for detecting and identifying helicobacter pylori.
<3>European Patent Office
<4>EP 1375677 A
<5>
<6>2004
<7>Probes for specifically detecting and identifying Helicobacter pylori which serves as an
inflammatory bacterium in digestive diseases.  These probes contain a fragment formed by
completely digesting a DNA carried by H. pylori with a restriction enzyme HindIII.  The
information of the base sequences of these probes are useful as a structural indication of
primers to be used in the construction of probes specific to H. pylori by PCR or as standard
sequences suitable for the comparison with genomic DNAs contained in clinical specimens.

<>

<1>Keshi, H., Nishida, K., Eda, S., Matsuhisa, A., Uehara, H.
<2>Probes for detecting and identifying helicobacter pylori.
<3>European Patent Office
<4>EP 1369495 A
<5>
<6>2003
<7>
<>

<1>Kesminiene, A., Maneliene, Z., Vitkute, J., Petrusyte, M., Janulaitis, A.
<2>FspAI, a unique type II restriction endonuclease that recognizes the octanucleotide sequence 5'-RTGC^GCAY-3'.
<3>Nucleic Acids Res.
<4>29
<5>e120
<6>2001
<7>A new type II restriction endonuclease designated FspAI has been partially purified from a
Flexibacter species Tv-m21K. FspAI recognizes the octanucleotide sequence
5'-RTGCdecreaseGCAY-3' and cleaves it in the center generating blunt-ended DNA fragments.

<>

<1>Kessler, C.
<2>Class II restriction endonucleases.
<3>Cytogenetics, Springer-Verlag, Obe, G., Basler, A., 
<4>0
<5>225-279
<6>1987
<7>The availability of a large variety of class II restriction endonucleases with
different site specificities is the basis of recombinant DNA technology and
numerous analytical applications.  The importance of this enzyme class is
demonstrated by recent findings in the detailed analysis of gene structure and
the rapid succession of spectacular advances in genetic engineering (Malcolm
1981; O'Connor et al. 1984).

<>

<1>Kessler, C.
<2>Restriction enzymes.
<3>Chemie in unserer Zeit
<4>22
<5>37-49
<6>1988
<7>None

<>

<1>Kessler, C., Bolton, B.J., Comer, M.J.
<2>Screening for novel Type II restriction endonucleases.
<3>J. Cell Biochem. Suppl.
<4>10D
<5>101
<6>1986
<7>Besides various species of lactic acid bacteria (Lactobacillus, Pediococcus and
Leuconostoc) we have screened 252 different non-pathogenic species of the
genera Achromobacter, Acinetobacter, Alcaligenes, Brevibacterium, Enterobacter,
Flavobacterium and Herpetosiphon for the presence of potentially new type II
restriction endonucleases.  Among the above lactic acid bacteria screened, we
could not detect any specific activities, whereas in all the other genera we
found a high number of species producing different type II restriction
endonucleases.

<>

<1>Kessler, C., Holtke, H.J.
<2>Specificity of restriction endonucleases and methylases - a review (Edition 2).
<3>Gene
<4>47
<5>1-153
<6>1986
<7>The properties and sources of all known restriction endonucleases and
methylases are listed.  The enzymes are cross-indexed (Table I), classified
according to their recognition sequence homologies (Table II), and
characterized within Table II by the cleavage and methylation positions, the
number of recognition sites on the double-stranded DNA of the bacteriophages
lambda, phiX174 and M13mp7, the viruses Ad2 and SV40, the plasmids pBR322 and
pBR328, and the microorganisms from which they originate.  Other tabulated
properties of the restriction endonucleases included relaxed specificities
(integrated into Table II), the structure of the generated fragment ends (Table
III), and the sensitivity to different kinds of DNA methylation (Table V).  In
Table IV the conversion of two-and four-base 5'-protruding ends into new
recognition sequences is compiled which is obtained by the fill-in reaction
with Klenow fragment of the Escherichia coli DNA polymerase I or additional
nuclease S1 treatment followed by ligation of the modified fragment termini.
Interconversion of restriction sites generates novel cloning sites without the
need of linkers.  This should improve the flexibility of genetic engineering
experiments.  Table VI classifies the restriction methylases according to the
nature of the methylated base(s) within their recognition sequences.  This
table also comprises restriction endonucleases which are known to be inhibited
or activated by the modified nucleotides.  The detailed sequences of those
overlapping restriction sites are also included which become resistant to
cleavage after the sequential action of corresponding restriction methylases
and endonuclease.  By this approach large DNA fragments can be generated which
is helpful in the construction of genomic libraries.  The data given in both
Table IV and VI allow the design of novel sequence specificities.  These
procedures complement the creation of universal cleavage specificities applying
class IIS enzymes and bivalent DNA adapter molecules.

<>

<1>Kessler, C., Manta, V.
<2>Specificity of restriction endonucleases and DNA modification methyltransferases - a review (Edition 3) .
<3>Gene
<4>92
<5>1-248
<6>1990
<7>The properties and sources of all known class-I, class-II and class-III
restriction endonucleases (ENases) and DNA modification methyltransferases
(MTases) are listed and newly subclassified according to their sequence
specificity.  In addition, the enzymes are distinguished in a novel manner
according to sequence specificity, cleavage position and methylation
sensitivity.  Furthermore, new nomenclature rules are proposed for
unambiguously defined enzyme names.  In the various Tables, the enzymes are
cross-indexed alphabetically according to their names (Table I), classified
according to their recognition sequence homologies (Table II), and
characterized within Table II by the cleavage and methylation positions, the
number of recognition sites on the DNA of the bacteriophages lambda, PhiX174,
and M13mp7, the viruses Ad2 and SV40, the plasmids pBR322 and pBR328, and the
microorganisms from which they originate.  Other tabulated properties of the
ENases include relaxed specificities (integrated within Table II), the
structure of the generated fragment ends (Table III), interconversion of
restriction sites (Table IV) and the sensitivity to different kinds of DNA
methylation (Table V).  Table VI shows the influence of class-II MTases on the
activity of class-II ENases with at least partially overlapping recognition
sequences.  Table VII lists all class-II restriction endonucleases and MTases
which are commercially available.

<>

<1>Kessler, C., Nesch, G., Brack, R.
<2>SphI restriction map of bacteriophage lambda DNA.
<3>Gene
<4>16
<5>321-323
<6>1981
<7>Upon reinvestigation, the number of cleavage sites for site-specific
endonuclease SphI on lambda DNA was found to be six.  The SphI restriction map
of the lambda cI857Sam7 genome was determined.

<>

<1>Kessler, C., Neumaier, P.S., Wolf, W.
<2>Recognition sequences of restriction endonucleases and methylases - a review.
<3>Gene
<4>33
<5>1-102
<6>1985
<7>The properties and sources of all known endonucleases and methylases acting site-specifically
on DNA are listed. The enzymes are crossindexed (Table I), classified according to homologies
within their recognition sequences (Table II), and characterized within Table II by the
cleavage and methylation positions, the number of recognition sites on the DNA of the
bacteriophages lambda, PhiX174 and M13mp7, the viruses Ad2 and SV40, the plasmids pBR322 and
pBR328 and the microorganims from which they originate. Other tabulated properties of the
restriction endonucleases include relaxed specificites (Table III), the structure of the
restriction fragment ends (Table IV), and the sensitivity to different kinds of DNA
methylation (Table V). Table VI classifies the methylases according to the nature of the
methylated base(s) within their recognition sequences. This table also comprises those
restriction endonucleases, which are known to be inhibited by the modified nucleotides.
Furthermore, this review includes a restriction map of bacteriophage lambda DNA based on
sequence data. Table VII lists the exact nucleotide positions of the cleavage sites, the
length of the generated fragments ordered according to size, and the effects of the
Escherichia coli dam- and dcm-coded methylases M Eco dam and M Eco dcmI on the particular
recognition sites.

<>

<1>Ketter, P., Guentzel, M.N., Chambers, J.P., Jorgensen, J., Murray, C.K., Cap, A.P., Yu, J.J., Eppinger, M., Arulanandam, B.P.
<2>Genome Sequences of Four Acinetobacter baumannii-A. calcoaceticus Complex Isolates from Combat-Related Infections Sustained in the Middle East.
<3>Genome Announcements
<4>2
<5>e00026-14
<6>2014
<7>Acinetobacter baumannii is among the most prevalent bacterial causes of combat-related
infections on the battlefield. Antibiotic resistance and a poor
understanding of the protective host immune responses make treatment difficult.
Here, we report the genome sequences of four clinical Acinetobacter baumannii-A.
calcoaceticus complex isolates exhibiting significant differences in virulence in
a mouse sepsis model.

<>

<1>Kettler, G.C., Martiny, A.C., Huang, K., Zucker, J., Coleman, M.L., Rodrigue, S., Chen, F., Lapidus, A., Ferriera, S., Johnson, J., Steglich, C., Church, G.M., Richardson, P., Chisholm, S.W.
<2>Patterns and implications of gene gain and loss in the evolution of Prochlorococcus.
<3>PLoS Genet.
<4>3
<5>E231
<6>2007
<7>Prochlorococcus is a marine cyanobacterium that numerically dominates the
mid-latitude oceans and is the smallest known oxygenic phototroph.
Numerous isolates from diverse areas of the world's oceans have been
studied and shown to be physiologically and genetically distinct. All
isolates described thus far can be assigned to either a tightly clustered
high-light (HL)-adapted clade, or a more divergent low-light (LL)-adapted
group. The 16S rRNA sequences of the entire Prochlorococcus group differ
by at most 3%, and the four initially published genomes revealed patterns
of genetic differentiation that help explain physiological differences
among the isolates. Here we describe the genomes of eight newly sequenced
isolates and combine them with the first four genomes for a comprehensive
analysis of the core (shared by all isolates) and flexible genes of the
Prochlorococcus group, and the patterns of loss and gain of the flexible
genes over the course of evolution. There are 1,273 genes that represent
the core shared by all 12 genomes. They are apparently sufficient,
according to metabolic reconstruction, to encode a functional cell. We
describe a phylogeny for all 12 isolates by subjecting their complete
proteomes to three different phylogenetic analyses. For each non-core
gene, we used a maximum parsimony method to estimate which ancestor likely
first acquired or lost each gene. Many of the genetic differences among
isolates, especially for genes involved in outer membrane synthesis and
nutrient transport, are found within the same clade. Nevertheless, we
identified some genes defining HL and LL ecotypes, and clades within these
broad ecotypes, helping to demonstrate the basis of HL and LL adaptations
in Prochlorococcus. Furthermore, our estimates of gene gain events allow
us to identify highly variable genomic islands that are not apparent
through simple pairwise comparisons. These results emphasize the
functional roles, especially those connected to outer membrane synthesis
and transport that dominate the flexible genome and set it apart from the
core. Besides identifying islands and demonstrating their role throughout
the history of Prochlorococcus, reconstruction of past gene gains and
losses shows that much of the variability exists at the "leaves of the
tree," between the most closely related strains. Finally, the
identification of core and flexible genes from this 12-genome comparison
is largely consistent with the relative frequency of Prochlorococcus genes
found in global ocean metagenomic databases, further closing the gap
between our understanding of these organisms in the lab and the wild.

<>

<1>Kettling, U., Koltermann, A., Schwille, P., Eigen, M.
<2>Real-time enzyme kinetics monitored by dual-color fluorescence cross-correlation spectroscopy.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>1416-1420
<6>1998
<7>A method for sensitively monitoring enzyme kinetics and activities by using dual-color
fluorescence cross-correlation spectroscopy is described.  This universal method enables the
development of highly sensitive and precise assays for real-time kinetic analyses of any
catalyzed cleavage or addition reaction, where a chemical linkage is formed or cleaved through
an enzyme's action between two fluorophores that can be discriminated spectrally.  In this
work, a homogeneous assay with restriction endonuclease EcoRI and a 66-bp double-stranded DNA
containing the GAATTC recognition site and fluorophores at each 5' end is described.  The
enzyme activity can be quantified down to the low picomolar range (>1.6 pM) where the rate
constants are linearly dependent on the enzyme concentrations over two orders of magnitude.
Furthermore, the reactions were monitored online at various initial substrate concentrations
in the nanomolar range, and the reaction rates were clearly represented by the
Michaelis-Menten equation with a KM of 14+/- 1 nM and a kcat of 4.6 +/- 0.2 min-1.  In
addition to kinetic studies and activity determinations, it is proposed that enzyme assays
based on the dual-color fluorescence cross-correlation spectroscopy will be very useful for
high-throughput screening and evolutionary biotechnology.

<>

<1>Ketyi, J., Orskov, F.
<2>Host-controlled modification and restriction of foreign chromosomal and plasmid DNA in Shigella flexneri strains.
<3>Acta Path. Microbiol. Scand.
<4>78
<5>51-58
<6>1970
<7>The hsp gene (the genetic determinant for host-controlled restriction and
modification) was transferred from an E. coli K12 strain into a S. flexneri
strain, and in reverse the same gene from a Shigella strain was introduced into
E. coli K12.  The presence of the heterologous hsp gene was tested by phage T5.
It was demonstrated in interrupted mating experiments with the hybrids as
recipients that the hsp hybrids had acquired changed recipient characters
resembling those of the strains from which the hsp originated.

<>

<1>Khadem, A.F. et al.
<2>Draft Genome Sequence of the Volcano-Inhabiting Thermoacidophilic Methanotroph Methylacidiphilum fumariolicum Strain SolV.
<3>J. Bacteriol.
<4>194
<5>3729-3730
<6>2012
<7>The draft genome of Methylacidiphilum fumariolicum SolV, a thermoacidophilic methanotroph of
the phylum Verrucomicrobia, is presented. Annotation revealed
pathways for one-carbon, nitrogen, and hydrogen catabolism and respiration
together with central metabolic pathways. The genome encodes three orthologues of
particulate methane monooxygenases. Sequencing of this genome will help in the
understanding of methane cycling in volcanic environments.

<>

<1>Khalid, M.I., Teh, L.K., Lee, L.S., Zakaria, Z.A., Salleh, M.Z.
<2>Genome Sequence of Proteus mirabilis Strain PR03, Isolated from a Local Hospital  in Malaysia.
<3>Genome Announcements
<4>1
<5>e00327-13
<6>2013
<7>Proteus mirabilis is one of the pathogenic agents that commonly causes urinary tract
infections among elderly individuals and long-term catheterized patients.
Here, we report a draft genome sequence of Proteus mirabilis strain PR03
(3,932,623 bp, with a G+C content of 38.6%) isolated from a local hospital in
Malaysia.

<>

<1>Khan, F., Furuta, Y., Kawai, M., Kaminska, K.H., Ishikawa, K., Bujnicki, J.M., Kobayashi, I.
<2>A putative mobile genetic element carrying a novel type IIF restriction-modification system (PluTI).
<3>Nucleic Acids Res.
<4>38
<5>3019-3030
<6>2010
<7>Genome comparison and genome context analysis were used to find a putative mobile element in
the genome of Photorhabdus luminescens, an
entomopathogenic bacterium. The element is composed of 16-bp direct
repeats in the terminal regions, which are identical to a part of
insertion sequences (ISs), a DNA methyltransferase gene homolog, two genes
of unknown functions and an open reading frame (ORF) (plu0599) encoding a
protein with no detectable sequence similarity to any known protein. The
ORF (plu0599) product showed DNA endonuclease activity, when expressed in
a cell-free expression system. Subsequently, the protein, named R.PluTI,
was expressed in vivo, purified and found to be a novel type IIF
restriction enzyme that recognizes 5'-GGCGC/C-3' (/ indicates position of
cleavage). R.PluTI cleaves a two-site supercoiled substrate at both the
sites faster than a one-site supercoiled substrate. The modification
enzyme homolog encoded by plu0600, named M.PluTI, was expressed in
Escherichia coli and shown to protect DNA from R.PluTI cleavage in vitro,
and to suppress the lethal effects of R.PluTI expression in vivo. These
results suggested that they constitute a restriction-modification system,
present on the putative mobile element. Our approach thus allowed
detection of a previously uncharacterized family of DNA-interacting
proteins.

<>

<1>Khan, M.A., Isaacson, R.E.
<2>Identification of Escherichia coli genes that are specifically expressed in a murine model of septicemic infection.
<3>Infect. Immun.
<4>70
<5>3404-3412
<6>2002
<7>Identification and characterization of bacterial genes that are induced during the disease
process are important in understanding the molecular
mechanism of disease and can be useful in designing antimicrobial drugs to
control the disease. The identification of in vivo induced (ivi) genes of
an Escherichia coli septicemia strain by using antibiotic-based in vivo
expression technology is described. Bacterial clones resistant to
chloramphenicol in vivo were recovered from the livers of infected mice.
Most of the ivi clones were sensitive to chloramphenicol when grown in
vitro. Using reverse transcription-PCR, it was demonstrated that selected
ivi clones expressed cat in the livers of infected mice but not during in
vitro growth. A total of 750 colonies were recovered after three
successive rounds of in vivo selection, and 168 isolated ivi clones were
sequenced. The sequence analysis revealed that 37 clones encoded
hypothetical proteins found in E. coli K-12, whereas 10 clones contained
genes that had no significant homology to DNA sequences in GenBank. Two
clones were found to contain transposon-related functions. Other clones
contained genes required for amino acid metabolism, anaerobic respiration,
DNA repair, the heat shock response, and the cellular repressor of the SOS
response. In addition, one clone contained the aerobactin biosynthesis
gene iucA. Mutations were introduced in to seven of the identified ivi
genes. An in vivo mouse challenge-competition assay was used to determine
if the mutants were attenuated. The results suggested that these ivi genes
were important for survival in vivo, and three of the seven mutant ivi
clones were required for successful infection of mice.

<>

<1>Khanna, N. et al.
<2>Complete genome sequence of Enterobacter sp. IIT-BT 08: A potential microbial strain for high rate hydrogen production.
<3>Standards in Genomic Sciences
<4>9
<5>359-369
<6>2013
<7>Enterobacter sp. IIT-BT 08 belongs to Phylum: Proteobacteria, Class: Gammaproteobacteria,
Order: Enterobacteriales, Family: Enterobacteriaceae. The
organism was isolated from the leaves of a local plant near the Kharagpur railway
station, Kharagpur, West Bengal, India. It has been extensively studied for
fermentative hydrogen production because of its high hydrogen yield. For further
enhancement of hydrogen production by strain development, complete genome
sequence analysis was carried out. Sequence analysis revealed that the genome was
linear, 4.67 Mbp long and had a GC content of 56.01%. The genome properties
encode 4,393 protein-coding and 179 RNA genes. Additionally, a putative pathway
of hydrogen production was suggested based on the presence of formate hydrogen
lyase complex and other related genes identified in the genome. Thus, in the
present study we describe the specific properties of the organism and the
generation, annotation and analysis of its genome sequence as well as discuss the
putative pathway of hydrogen production by this organism.

<>

<1>Khatri, I., Dureja, C., Raychaudhuri, S., Subramanian, S.
<2>Draft Genome Sequence of the Opportunistic Human Pathogen Morganella morganii SC01.
<3>Genome Announcements
<4>1
<5>e00051-12
<6>2013
<7>We report the 4.1-Mb draft genome sequence of Morganella morganii SC01, a
gammaproteobacterium, isolated from an Indian human fecal sample.

<>

<1>Khatri, I., Kaur, S., Devi, U., Kumar, N., Sharma, D., Subramanian, S., Saini, A.K.
<2>Draft Genome Sequence of Plant Growth-Promoting Rhizobacterium Pantoea sp. Strain AS-PWVM4.
<3>Genome Announcements
<4>1
<5>e00947-13
<6>2013
<7>Nonpathogenic Pantoea spp. have been shown to confer biofertilizer and biocontrol activities,
indicating their potential for increasing crop yield. Herein, we
provide the high-quality genome sequence of Pantoea sp. strain AS-PWVM4, a
Gram-negative motile plant growth-promoting rhizobacterium isolated from a
pomegranate plant. The 4.9-Mb genome contains genes related to plant growth
promotion and the synthesis of siderophores.

<>

<1>Khatri, I., Nupur, K.S., Subramanian, S., Pinnaka, A.K.
<2>Draft Genome Sequence of Rhodovulum sp. Strain PH10, a Phototrophic Alphaproteobacterium Isolated from a Soil Sample of Mangrove of Namkhana, India.
<3>J. Bacteriol.
<4>194
<5>6363
<6>2012
<7>We report the 4.8-Mb draft genome of Rhodovulum sp. strain PH10, a phototrophic bacterium
belonging to class Alphaproteobacteria, isolated from a soil sample
collected from the mangrove forest of Namkhana in India. This genome is the first
from the genus Rhodovulum and will lead to a better understanding of the
genes/pathways involved in activities like phototrophic growth and nitrogen
fixation in this group of bacteria.

<>

<1>Khatri, I., Singh, A., Korpole, S., Pinnaka, A.K., Subramanian, S.
<2>Draft Genome Sequence of an Alphaproteobacterium, Caenispirillum salinarum AK4(T), Isolated from a Solar Saltern.
<3>Genome Announcements
<4>1
<5>e00199-12
<6>2013
<7>We report the 4.9-Mb genome sequence of Caenispirillum salinarum AK4(T) isolated  from a
sediment sample collected from a solar saltern at Kakinada, Andhra
Pradesh, India.

<>

<1>Khayi, S., Mondy, S., Beury-Cirou, A., Moumni, M., Helias, V., Faure, D.
<2>Genome Sequence of the Emerging Plant Pathogen Dickeya solani Strain RNS 08.23.3.1A.
<3>Genome Announcements
<4>2
<5>e01270-13
<6>2014
<7>Here we present the genome sequence of Dickeya solani strain RNS 08.23.3.1A (PRI3337),
isolated from Solanum tuberosum. Dickeya solani, recently described on
potato cultures in Europe, is a proposed new taxon closely related to the Dickeya
dianthicola and Dickeya dadantii species.

<>

<1>Khedkar, S., Prabhakara, S., Loganathan, R.M., SC, G.M., Arakere, G., Seshasayee, A.S.
<2>Draft Genome Sequence of Staphylococcus aureus ST672, an Emerging Disease Clone from India.
<3>J. Bacteriol.
<4>194
<5>6946-6947
<6>2012
<7>We report the draft genome sequence of methicillin-resistant Staphylococcus aureus (MRSA)
strain ST672, an emerging disease clone in India, from a septicemia
patient. The genome size is about 2.82 Mb with 2,485 open reading frames (ORFs).
The staphylococcal cassette chromosome mec (SCCmec) element (type V) and immune
evasion cluster appear to be different from those of strain ST772 on preliminary
examination.

<>

<1>Khelaifia, S., Garibal, M., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequencing of Methanobrevibacter oralis Strain JMR01, Isolated from  the Human Intestinal Microbiota.
<3>Genome Announcements
<4>2
<5>e00073-14
<6>2014
<7>Methanobrevibacter oralis, an anaerobic methanogenic archaeon, has been previously isolated
from the human oral cavity. Here, sequencing a stool isolate
(strain JMR01) yielded a 2.065-Mb genome with a 27.78% G+C content containing a
total of 2,042 open reading frames and 3 clusters of regularly interspaced short
palindromic repeat (CRISPR) loci with associated Cas proteins.

<>

<1>Khelaifia, S., Garibal, M., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of a Human-Associated Isolate of Methanobrevibacter arboriphilicus, the Lowest-G+C-Content Archaeon.
<3>Genome Announcements
<4>2
<5>e01181-13
<6>2014
<7>We report the draft genome sequence of Methanobrevibacter arboriphilicus strain ANOR1,
isolated from the human gut. Its 2.21-Mb genome exhibits a 25.46% G+C
content, the lowest value among archaea. The genome of M. arboriphilicus contains
a total of 2,111 open reading frames and three clusters of regularly interspaced
short palindromic repeat (CRISPR) loci with associated Cas proteins.

<>

<1>Khemayan, K., Prachumwat, A., Sonthayanon, B., Intaraprasong, A., Sriurairatana, S., Flegel, T.W.
<2>Complete Genome Sequence of Virulence-Enhancing Siphophage VHS1 from Vibrio harveyi.
<3>Appl. Environ. Microbiol.
<4>78
<5>2790-2796
<6>2012
<7>Vibrio harveyi siphophage 1 (VHS1) is a tailed phage with an icosahedral head of
approximately 66 nm in diameter and an unornamented, flexible tail of
approximately 153 nm in length. When Vibrio harveyi 1114GL is lysogenized with
VHS1, its virulence for the black tiger shrimp (Penaeus monodon) increases by
more than 100 times, and this coincides with production of a toxin(s) associated
with shrimp hemocyte agglutination. Curiously, the lysogen does not show
increased virulence for the whiteleg shrimp (Penaeus [Litopenaeus] vannamei).
Here we present and annotate the complete, circular genome of VHS1 (81,509 kbp;
GenBank accession number JF713456). By software analysis, the genome contains 125
putative open reading frames (ORFs), all of which appear to be located on the
same DNA strand, similar to the case for many other bacteriophages. Most of the
putative ORFs show no significant homology to known sequences in GenBank. Notable
exceptions are ORFs for a putative DNA polymerase and putative phage structural
proteins, including a portal protein, a phage tail tape measure protein, and a
phage head protein. The last protein was identified as a component of the
species-specific toxin mixture described above as being associated with
agglutination of hemocytes from P. monodon.

<>

<1>Khesin, R.B., Bogdanova, E.S.
<2>The specificity of DNA-methylase in T2-phage infected E. coli B cells.
<3>Biokhimiia
<4>31
<5>405-415
<6>1966
<7>Activity of the enzymatic system responsible for DNA methylation considerably increases after
infection of E. coli B cells with T2 phage.  The increase is inhibited by chloramphenicol.
The non-infected cells contain no inhibitor of DNA methylation, and the infected ones - no
activator of this reaction.  Hence, there is T2 phage induced production of phage DNA
methylase along with the synthesis of other enzymes.  Enzymatic preparations from the
non-infected and infected bacteria are capable of methylating heterologous (thymus, M.
lysodeikticus etc.) DNA but do not react with DNA from non-infected E. coli and from T2 and T4
phages.  However, both the enzymes methylate DNA of E. coli and T2 phage if it was
incompletely methylated in intact cells prior to isolation.  Partially methylated DNA loses
its methyl-acceptor ability with respect to the enzyme of non-infected cells after incubation
in vitro with the enzymatic system from infected cells.  Hence, the non-infected and the
infected cell methylases both react with the same combinations of nucleotides in E. coli and
T2 phage DNA, i.e., have the same specificity.  It is proposed, that methylated nucleotides in
DNA have something to do with the interaction with RNA-polymerase.  RNA prior to, as well as
after infection with T2 phage being synthesized with the same RNA-polymerase, bacterial as
well as phage DNA ought to contain the same sequences of methylated nucleotides.  This
explains the observed identity of specificities of the T2-phage induced methylase and of that
existing prior to infection.

<>

<1>Khmelenina, V.N. et al.
<2>Draft Genome Sequence of Methylomicrobium buryatense Strain 5G, a Haloalkaline-Tolerant Methanotrophic Bacterium.
<3>Genome Announcements
<4>1
<5>e00053-13
<6>2013
<7>Robust growth of the gammaproteobacterium Methylomicrobium buryatense strain 5G on methane
makes it an attractive system for CH4-based biocatalysis. Here we
present a draft genome sequence of the strain that will provide a valuable
framework for metabolic engineering of the core pathways for the production of
valuable chemicals from methane.

<>

<1>Kho, M.R., Baker, D.J., Laayoun, A., Smith, S.S.
<2>Stalling of human DNA (cytosine-5) methyltransferase at single-strand conformers from a site of dynamic mutation.
<3>J. Mol. Biol.
<4>275
<5>67-79
<6>1998
<7>Single-strand conformers from the C-rich strand of the triplet repeat at the FMR-1 locus are
rapidly and selectively methylated by the human DNA (cytosine-5) methyltransferase.  The
apparent affinity of the enzyme for the FMR-1 SSC is about tenfold higher than it is for a
control Watson-Crick paired duplex.  The de novo methylation rate for the SSC is over 150-fold
higher than the de novo rate for the control duplex.  Methylation of what is generally called
a hemi-methylated duplex occurs with a rate enhancement of over 100-fold, while methylation of
what can be viewed as a hemi-methylated FMR-1 SSC is actually slower than the de novo rate.
The pronounced inhibition of the methyltransferase by the methylated SSC sugggests that the
enzyme has a higher affinity for the methylated product of its reaction with the SSC than it
has for the unmethylated SSC substrate.  Gel retardation studies show that the
methyltransferase binds selectively to SSCs from the C-rich strand of the FMR-1 triplet
repeat.  This suggests a two-step stalling process in which the human methyltransferase first
selectively methylates and subsequently stalls at the C-rich strand SSC.  Stalling may reflect
the inability of the enzyme to release a DNA product that is fixed in a conformation
resembling its transition state by the unusual structure of the substrate.  In particular, the
data suggest that DNA methyltransferase may physically participate in biological processes
that lead to dynamic mutation at FMR-1.  In general, the data raise the possibility that a
two-step stalling process occurs at secondary structures associated with chromosome
instability, chromosome remodelling, viral replication or viral integration and may account
for the local hypermethylation and global hypomethylation associated with viral and non-viral
tumorigenesis.

<>

<1>Kholmina, G.V., Rebentish, B.A., Kozlovskii, Y.-E., Sorokin, A.V., Goldenberg, D.S., Prozorov, A.A.
<2>Purification and properties of two restriction endonucleases from Bacillus subtilis.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>23-25
<6>1986
<7>Two restriction endonucleases, Bsu1532I and Bsu1854I, have been isolated from
Bac. subtilis 1532 and 1854 cells and characterized.  The first of them,
Bsu1532I, recognizes and digests the tetranucleotide palindrome sequence GC^GC,
while Bsu1854I recognizes and digests the degenerate hexanucleotide sequence
GPuGCPy^C.  The optimum conditions were determined for both enzymes, and the
influence of various cofactors on the reactions catalyzed by them were studied.

<>

<1>Kholmina, G.V., Rebentish, B.A., Skoblov, Y.S., Mironov, A.A., Yankovskii, N.K., Kozlov, Y.I., Glatmann, L.I., Moroz, A.F., Debabov, V.G.
<2>Isolation and characterization of a new site-specific endonuclease, EcoRV.
<3>Dokl. Akad. Nauk.
<4>253
<5>495-497
<6>1980
<7>None

<>

<1>Kholodii, G.
<2>The shuffling function of resolvases.
<3>Gene
<4>269
<5>121-130
<6>2001
<7>Redistribution (shuffling) of genetic material between replicons requires
consecutive recombination at four points, two of which (X, X') are
involved in the replicon fusion and the other two (Y, Y'), in the
cointegrate resolution. The fusion of replicons by bacterial resolvases
makes the second recombination round at sites Y and Y' problematic because
of the high probability of the reverse reaction. Structural differences of
the res sites recognized by resolvase could delay the reverse reaction,
thus enhancing the probability of recombination at sites Y, Y', but the
direct reaction ensuring it (i.e. the fusion of replicons via different
res sites) has not been described yet. Here, a genetic system to test
intermolecular recombination at heterogeneous res sites has been
developed. The system was based on the res site (RS2) of the novel
resolution system, cinH-RS2, encoded by pKLH2, pKLH204 and pKLH205. As its
partner, the res site of RP4 located in the par locus, the res site of
transposon gammadelta or Tn1721, the incomplete site RS1 consisting only
of the (crossover) subsite resI also found in pKLH2/204/205 and others
were used. Except for the pairing of RS2 x gammadelta res, recombination
was observed in each case even when the homology shared by partners did
not exceed 35% (as in RS2 x par). In the latter case, the presence in cis
of an additional, enhancer-like-acting element was required. Pairing of
crossover subsites during site-specific recombination occurred in either
orientation, depending on the structure of res partners and the kind of
resolvase acting on the sites. With the complete res sites, the
antiparallel alignment resulted in the production of an unusual res having
the accessory subsites II and III at both sides of I, and a res lacking II
and III. The wide range of frequencies was observed not only in the fusion
formation but also in the dissociation of the resulting cointegrates.
Hence, the resolvase-mediated interreplicon exchange of the DNA segments
by fusion via an inefficient reaction (at sites X, X') and dissociation
via an efficient one (at sites Y, Y') become possible.

<>

<1>Khosaka, T., Kiwaki, M.
<2>Restriction endonucleases from Bifidobacterium bifidum.
<3>FEBS Lett.
<4>177
<5>57-60
<6>1984
<7>Three restriction endonucleases, BbiI, BbiII and BbiIII, have been isolated
from Bifidobacterium bifidum.  The recognition and cleavage specificity of
BbiII was determined to be 5'-GR^CGYC-3', identical to that of AcyI isolated
from a cyanobacterium Anabaena cylindrica.  The other two enzymes, BbiI and
BbiIII, were found to be isoschizomers of PstI and XhoI, respectively.

<>

<1>Khosaka, T., Kiwaki, M.
<2>BinI: A new site-specific endonuclease from Bifidobacterium infantis.
<3>Gene
<4>31
<5>251-255
<6>1984
<7>A new restriction endonuclease, BinI, from Bifidobacterium infantis 659 has
been isolated.  By mapping and sequencing of the cleavage sites on SV40 DNA, it
was deduced that BinI recognizes the asymmetric pentanucleotide sequence
5'-GGATCNNNN^-3'3'-CCTAGNNNNN^5'and cleaves at the sites indicated by the
arrows, generating mononucleotide 5'-terminal extensions.  BinI is a member of
a unique class of Type-II restriction endonucleases which recognize a specific
but asymmetric sequence and cleave at a site several bases away.

<>

<1>Khosaka, T., Kiwaki, M., Rak, B.
<2>Two site-specific endonucleases BinSI and BinSII from Bifidobacterium infantis.
<3>FEBS Lett.
<4>163
<5>170-174
<6>1983
<7>Two site-specific endonucleases, BinSI and BinSII, were isolated from
Bifidobacterium infantis S76e.  BinSI was found to be an isoschizomer of EcoRII, while
BinSII was shown to have the same sequence and cutting specificity as BbeI, 5'-
GGCGCC-3'.  Both BinSII- and BbeI-generated DNA fragments could be ligated with
HaeII-generated DNA fragments.

<>

<1>Khosaka, T., Sakurai, T., Mutai, M.
<2>A restriction enzyme and a method for production thereof.
<3>European Patent Office
<4>EP 76696 A
<5>
<6>1981
<7>Restriction enzyme has a specificity such that it can recognise a palindromic hexanucleotide
sequence GGCGC^C in double-stranded DNAs and it can cleave the phosphodiester bonds in the
DNAs between the 2 C's at the 3'-ends, as shown by the arrows. DNA fragments having
tetranucleotide long 3'-protruding ends are generated. Production of a restriction enzyme
comprises cultivation of a strain of Bifidobacterium breve and then extracting the enzyme from
the resulting culture. Enzyme (I) recognises a specific nucleotide sequence in a DNA and
cleaves the DNA at a specific site inside or close to the nucleotide sequence. It is therefore
useful for physical mapping of a DNA, for the analysis of nucleotide sequences, for the
isolation of genes from a microorganism, etc. (I) can be obtained in large yields.

<>

<1>Khosaka, T., Sakurai, T., Takahashi, H., Saito, H.
<2>A new site-specific endonuclease BbeI from Bifidobacterium breve.
<3>Gene
<4>17
<5>117-122
<6>1982
<7>A new site-specific endonuclease, BbeI, has been partially purified from the
anerobic bacterium, Bifidobacterium breve.  BbeI recognizes the hexanucleotide
sequence5'-GGCGC^C-3' 3'-C^CGCGG-5'and cleaves it at the sites indicated by the
arrows, producing 3'-cohesive termini four bases long.

<>

<1>Khosravi, Y., Rehvathy, V., Wee, W.Y., Wang, S., Baybayan, P., Singh, S., Ashby, M., Ong, J., Amoyo, A.A., Seow, S.W., Choo, S.W., Perkins, T., Chua, E.G., Tay, A., Marshall, B.J., Loke, M.F., Goh, K.L., Pettersson, S., Vadivelu, J.
<2>Comparing the genomes of Helicobacter pylori clinical strain UM032 and Mice-adapted derivatives.
<3>Gut Pathog.
<4>5
<5>25
<6>2013
<7>Background: Helicobacter pylori is a Gram-negative bacterium that persistently infects the
human stomach inducing chronic inflammation.
The exact mechanisms of pathogenesis are still not completely
understood. Although not a natural host for H. pylori, mouse infection
models play an important role in establishing the immunology and
pathogenicity of H. pylori. In this study, for the first time, the
genome sequences of clinical H. pylori strain UM032 and mice-adapted
derivatives, 298 and 299, were sequenced using the PacBio Single
Molecule, Real-Time (SMRT) technology.
Result: Here, we described the single contig which was achieved for
UM032 (1,599,441 bp), 298 (1,604,216 bp) and 299 (1,601,149 bp).
Preliminary analysis suggested that methylation of H. pylori genome
through its restriction modification system may be determinative of its
host specificity and adaptation.
Conclusion: Availability of these genomic sequences will aid in
enhancing our current level of understanding the host specificity of H.
pylori.

<>

<1>Khudyakov, I.Y., Kirnos, M.D., Aleksandrushkina, N.I., Vanyushin, B.F.
<2>2,6-Diaminopurine - a new adenine-replacing base in the DNA of cyanophage S-2.
<3>Dokl. Akad. Nauk.
<4>232
<5>965-968
<6>1977
<7>By now only some of the physicochemical properties of extremely few viruses of
blue-green algae-cyanophages-have been studied.  Unfortunately, the information
on the nucleotide composition of the DNA of cyanophages is based only on a
determination of the GC content according to the melting point and buoyant
density.  However, this does not permit a correct determination even of the
composition, if the DNA contains unusual and minor bases.  Unusual bases, which
can entirely replace cytosine or thymine residues, have been found in the DNA
of many bacterial phages.  The DNAs of cyanophages have not been studied at all
in this respect.  Moreover, it is unknown whether these DNAs contain the
modified residues of the usual bases, characterisitic of many prokaryotes and
responsible for host modification and restriction.  It is also unclear whether
host restriction and modification analogous to that in bacteria exist in
blue-green algae and cyanophages.  Therefore, a study of the intrinsic nature
of the bases in the DNA of cyanophages is of special interest.  In this work we
studied the composition and some physicochemical properties of the DNA of a new
cyanophage S-2, which lyses the blue-green alga Synechococcus sp. 698 (from the
collection of the Biological Institute of Leningrad University).  We isolated
this cyanophage from aquatic samples, taken in the environs of Leningrad.  It
consists of an icosahedral head 56 nm in diameter and a flexible,
noncontractile tail process 120 nm long.  The phage was concentrated by
chromatography on DEAE-cellulose (Whatman DE32) and purified by centrifuging in
a CsCl gradient.  The cyanophage DNA was isolated by a phenol method.

<>

<1>Khudyakov, I.Y., Kirnos, M.D., Alexandrushkina, N.I., Vanyushin, B.F.
<2>Cyanophage S-2L contains DNA with 2,6-diaminopurine substituted for adenine.
<3>Virology
<4>88
<5>8-18
<6>1978
<7>Cyanophage S-2L, which lyses some strains of unicellular cyanobacteria of the
genus Synechococcus, has a polyhedral head 56 nm in diameter and a flexible
noncontractile tail 120 nm in length.  In S-2L phage DNA, 2,6-diaminopurine is
completely substituted for adenine.  This base and the corresponding
deoxyribonucleoside have been isolated from acid and enzymatic hydrolyzates of
the phage DNA, respectively, and have been identified according to optical and
chromatographic behavior.  S-2L phage DNA is linear and double-stranded and has
a molecular weight of 26 to 28,000000.  The buoyant density of this DNA is CsCl
is 1.731 g/cm3.  2,6-Diaminopurine stabilizes the secondary structure of phage
DNA, i.e., the melting temperature of the latter (Tm is 85.6 degrees in
0.1xSSC) is 3.6 degrees higher than that of the usual adenine-containing DNA of
equivalent base composition.

<>

<1>Kiatpapan, P., Murooka, Y.
<2>Genetic manipulation system in propionibacteria.
<3>J. Biosci. Bioeng.
<4>93
<5>1-8
<6>2002
<7>Members of the genus Propionibacterium are widely used in the production of vitamin B12'
tetrapyrrole compounds, and propionic acid
as well as in the probiotic and cheese industries. Shuttle vectors were
developed in propionibacteria using replicons from endogenous plasmids
in Propionibacterium and Escherichia coli and an appropriate selection
marker. Efficient transformation was achieved using the shuttle
vector prepared from Propionibacterium freudenreichii to overcome the
high restriction modification system in propionibacteria. Expression
vectors with native promoters for use in propionibacteria were also
developed. Using this system, cholesterol oxidase, which is used as a
diagnostic enzyme, was produced in P freudenreichii. Genes involved in
5-aminolevulinic acid (ALA) and vitamin 13, biosynthesis in
propionibacteria were isolated. ALA in propionibacteria could be
synthesized via both the C4 pathway (condensation of glycine and
succinyl CoA) and the C5 pathway (from glutamate). The hemA gene
encoding ALA synthase from Rhodobacter spheroides, was overexpressed
and ALA accumulated in P. freudenreichii. Thus, the genetic manipulation
systems in propionibacteria will facilitate genetic studies of
probioties and the vitamin B-12 biosynthetic pathway.

<>

<1>Kienesberger, S., Sprenger, H., Wolfgruber, S., Halwachs, B., Thallinger, G.G., Perez-Perez, G.I., Blaser, M.J., Zechner, E.L., Gorkiewicz, G.
<2>Comparative Genome Analysis of Campylobacter fetus Subspecies Revealed Horizontally Acquired Genetic Elements Important for Virulence and Niche Specificity.
<3>PLoS ONE
<4>9
<5>E85491
<6>2014
<7>Campylobacter fetus are important animal and human pathogens and the two major
subspecies differ strikingly in pathogenicity. C. fetus subsp. venerealis is
highly niche-adapted, mainly infecting the genital tract of cattle. C. fetus
subsp. fetus has a wider host-range, colonizing the genital- and intestinal-tract
of animals and humans. We report the complete genomic sequence of C. fetus subsp.
venerealis 84-112 and comparisons to the genome of C. fetus subsp. fetus 82-40.
Functional analysis of genes predicted to be involved in C. fetus virulence was
performed. The two subspecies are highly syntenic with 92% sequence identity but
C. fetus subsp. venerealis has a larger genome and an extra-chromosomal element.
Aside from apparent gene transfer agents and hypothetical proteins, the unique
genes in both subspecies comprise two known functional groups: lipopolysaccharide
production, and type IV secretion machineries. Analyses of
lipopolysaccharide-biosynthesis genes in C. fetus isolates showed linkage to
particular pathotypes, and mutational inactivation demonstrated their roles in
regulating virulence and host range. The comparative analysis presented here
broadens knowledge of the genomic basis of C. fetus pathogenesis and host
specificity. It further highlights the importance of surface-exposed structures
to C. fetus pathogenicity and demonstrates how evolutionary forces optimize the
fitness and host-adaptation of these pathogens.

<>

<1>Kieser, T., Hopwood, D.A.
<2>Genetic manipulation of Streptomyces: integrating vectors and gene replacement.
<3>Methods Enzymol.
<4>204
<5>430-458
<6>1991
<7>The Streptomyces chromosome is a circle of about 72% G + C DNA at
least 1.5 times the size of the Escherichia coli chromosome.~ Streptomyces
coelicolor A3(2) is genetically by far the most studied streptomycete, with
a detailed chromosomal linkage map 2 and a combined physical/genetic
map based on pulsed-field gel analysis of large fragments generated by
enzymes that recognize permutations of the sequence A3T3 such as AseI,
DraI, and SspI. 3 In S. coelicolor fundamental studies of differentiation,
antibiotic production, and many other aspects of cell and molecular biology are being made by
a combination of in vivo and in vitro genetics.4,5 As
well as being amenable to a wide range of in vivo genetic procedures, the
A3(2) strain is an excellent host for the homologous cloned DNA required
for many of these studies. However, S. coelicolor shows very strong
restriction against DNA from E. coli, so the closely related Streptomyces
lividans 66 has been used for most heterologous cloning. 6-8 Streptomyces
lividans is largely nonrestricting and has the added advantage that it can
be used as an intermediate host for cloned DNA manipulated in E. coli
and destined for S. coelicolor A3(2). [Streptomyces coelicolor A3(2) has a
methylation-dependent restriction system. Bifunctional plasmids isolated
from a completely nonmethylating (Dam- Dcm- HsdS-) strain of E. coli
transform S. coelicolor. 9 This approach first succeeded for Streptomyces
avermitilis, which has methylation-dependent restriction systems. ~°]
Whether the recently discovered ability of some E. coli plasmids to promote
conjugation with Streptomyces, leading to plasmid transfer, u will
alleviate restriction barriers remains to be seen, but it clearly has some
other interesting practical implications. Differences between S. coelicolor
and S. lividans are listed in Table I.

<>

<1>Kilgore, J.A., Hoose, S.A., Gustafson, T.L., Porter, W., Kladde, M.P.
<2>Single-molecule and population probing of chromatin structure using DNA methyltransferases.
<3>Methods
<4>41
<5>320-332
<6>2007
<7>Probing chromatin structure with DNA methyltransferases offers advantages over more commonly
used nuclease-based and chromatin
immunoprecipitation methods for detection of nucleosomes and
non-histone protein DNA interactions. Here, we describe two related
methods in which the readout of MTase accessibility is obtained by
assaying 5-methylcytosine in DNA through the PCR-based technique of
bisulfite genomic sequencing. The methyltransferase accessibility
protocol (MAP) determines the relative frequency at which the enzyme
accesses each of its target sites over an entire population of PCR
amplified product. While MAP yields much quantitative information about
relative accessibility of a region of chromatin, a complementary
single-molecule view of methyltransferase accessibility, termed MAP for
individual templates (MAP-IT), is provided by analysis of cloned PCR
products. Absolute rather than relative methylation frequencies in a
region are obtained by summing the methylation status at each site over
a cohort of clones. Moreover, as the integrity of individual molecules
is maintained in MAP-IT, unique information about the distribution of
multiple footprints along continuous regions is gleaned. In principle,
the population MAP and single-molecule MAP-IT strategies can be used to
analyze chromatin structure in a variety of model systems. Here, we
describe the application of MAP in living Saccharomyces cerevisiae
cells and MAP-IT in the analysis of a mammalian tumor suppressor gene
in nuclei. This application of MAP-IT provides the first means to
simultaneously determine CpG methylation of mammalian genes and their
overlying chromatin structure in the same single DNA molecule.

<>

<1>Kim, B.-J., Choi, B.S., Lim, J.S., Choi, I.Y., Lee, J.H., Chun, J., Kook, Y.H., Kim, B.J.
<2>Complete Genome Sequence of Mycobacterium intracellulare Strain ATCC 13950T.
<3>J. Bacteriol.
<4>194
<5>2750
<6>2012
<7>Here we report the first complete genome sequence of Mycobacterium intracellulare ATCC
13950(T), a Mycobacterium avium complex (MAC) strain. This genome sequence will serve as a
valuable reference for understanding the epidemiologic, biological, and pathogenic aspects of
the disparity between MAC members.

<>

<1>Kim, B.J., Choi, B.S., Choi, I.Y., Lee, J.H., Chun, J., Hong, S.H., Kook, Y.H., Kim, B.J.
<2>Complete Genome Sequence of Mycobacterium intracellulare Clinical Strain MOTT-36Y, Belonging to the INT5 Genotype.
<3>J. Bacteriol.
<4>194
<5>4141-4142
<6>2012
<7>Here we report the complete genome sequence of the Mycobacterium intracellulare clinical
strain MOTT-36Y, previously grouped into the INT5 genotype among the 5
genotypes of M. intracellulare. This genome sequence will serve as a valuable
reference for understanding the disparity in virulence and epidemiologic traits
between M. intracellulare-related strains.

<>

<1>Kim, B.J., Choi, B.S., Lim, J.S., Choi, I.Y., Kook, Y.H., Kim, B.J.
<2>Complete Genome Sequence of Mycobacterium intracellulare Clinical Strain MOTT-64, Belonging to the INT1 Genotype.
<3>J. Bacteriol.
<4>194
<5>3268
<6>2012
<7>Here, we report the complete genome sequence of the Mycobacterium intracellulare  clinical
strain MOTT-64, previously grouped into the INT1 genotype among five
genotypes of M. intracellulare. This genome sequence will serve as a valuable
reference for understanding the disparity in the virulence and epidemiologic
traits among M. intracellulare genotypes.

<>

<1>Kim, B.J., Choi, B.S., Lim, J.S., Choi, I.Y., Lee, J.H., Chun, J., Kook, Y.H., Kim, B.J.
<2>Complete Genome Sequence of Mycobacterium intracellulare Clinical Strain MOTT-02.
<3>J. Bacteriol.
<4>194
<5>2771
<6>2012
<7>Here, we report the first complete genome sequence of the Mycobacterium intracellulare
clinical strain MOTT-02, which was previously grouped in the INT2
genotype of M. intracellulare. This genome sequence will serve as a valuable
reference for improving the understanding of the disparity in the virulence and
epidemiologic traits between M. intracellulare genotypes.

<>

<1>Kim, B.J., Kim, B.R., Hong, S.H., Seok, S.H., Kook, Y.H., Kim, B.J.
<2>Complete Genome Sequence of Mycobacterium massiliense Clinical Strain Asan 50594, Belonging to the Type II Genotype.
<3>Genome Announcements
<4>1
<5>e00429-13
<6>2013
<7>We report the complete genome sequence of the Mycobacterium massiliense clinical  strain Asan
50594, which was grouped into the M. massiliense type II genotype,
isolated from a Korean patient. This genome sequence will serve as a valuable
reference for understanding the disparity in virulence and epidemiological traits
between strains belonging to the Mycobacterium abscessus complex.

<>

<1>Kim, B.J., Kim, B.R., Lee, S.Y., Seok, S.H., Kook, Y.H., Kim, B.J.
<2>Whole-Genome Sequence of a Novel Species, Mycobacterium yongonense DSM 45126T.
<3>Genome Announcements
<4>1
<5>e00604-13
<6>2013
<7>Here, we report the complete genome sequence of Mycobacterium yongonense DSM 45126(T),
genetically closely related to the INT5 genotype of M. intracellulare.

<>

<1>Kim, B.K., Chung, J.H., Kim, S.Y., Jeong, H., Kang, S.G., Kwon, S.K., Lee, C.H., Song, J.Y., Yu, D.S., Ryu, C.M., Kim, J.F.
<2>Genome Sequence of the Leaf-Colonizing Bacterium Bacillus sp. Strain 5B6, Isolated from a Cherry Tree.
<3>J. Bacteriol.
<4>194
<5>3758-3759
<6>2012
<7>Plant growth-promoting bacteria colonize various habitats, including the phyllosphere. Here,
we present the high-quality draft genome sequence of Bacillus
sp. strain 5B6, which was isolated from the leaf of a cherry tree. The 3.9-Mb
genome uncovers its potential for understanding the nature of leaf colonization
as well as antibiosis against plant pathogens.

<>

<1>Kim, B.K., Jung, M.Y., Yu, D.S., Park, S.J., Oh, T.K., Rhee, S.K., Kim, J.F.
<2>Genome Sequence of an Ammonia-Oxidizing Soil Archaeon, 'Candidatus Nitrosoarchaeum koreensis' MY1.
<3>J. Bacteriol.
<4>193
<5>5539-5540
<6>2011
<7>Ammonia-oxidizing archaea are ubiquitous microorganisms which play important
roles in global nitrogen and carbon cycle on earth. Here we present the
high-quality draft genome sequence of an ammonia-oxidizing archaeon, "Candidatus
Nitrosopumilus koreensis" MY1, that dominated an enrichment culture of a soil
sample from the rhizosphere. Its genome contains genes for survival in the
rhizosphere environment as well as those for carbon fixation and ammonium
oxidation to nitrite.

<>

<1>Kim, B.K., Lee, S.H., Kim, S.Y., Jeong, H., Kwon, S.K., Lee, C.H., Song, J.Y., Yu, D.S., Kang, S.G., Kim, J.F.
<2>Genome Sequence of an Oligohaline Hyperthermophilic Archaeon, Thermococcus zilligii AN1, Isolated from a Terrestrial Geothermal Freshwater Spring.
<3>J. Bacteriol.
<4>194
<5>3765-3766
<6>2012
<7>Thermococcus zilligii, a thermophilic anaerobe in freshwater, is useful for physiological
research and biotechnological applications. Here we report the
high-quality draft genome sequence of T. zilligii AN1(T). The genome contains a
number of genes for an immune system and adaptation to a microbial biomass-rich
environment as well as hydrogenase genes.

<>

<1>Kim, B.K., Song, G.C., Hong, G.H., Seong, W.K., Kim, S.Y., Jeong, H., Kang, S.G., Kwon, S.K., Lee, C.H., Song, J.Y., Yu, D.S., Park, M.S., Cho, S.H., Kim, J.F.
<2>Genome Sequence of the Shiga Toxin-Producing Escherichia coli Strain NCCP15657.
<3>J. Bacteriol.
<4>194
<5>3751-3752
<6>2012
<7>Shiga toxin-producing Escherichia coli causes bloody diarrhea and hemolytic-uremic syndrome
and serious outbreaks worldwide. Here, we report the
draft genome sequence of E. coli NCCP15657 isolated from a patient. The genome
has virulence genes, many in the locus of enterocyte effacement (LEE) island,
encoding a metalloprotease, the Shiga toxin, and constituents of type III
secretion.

<>

<1>Kim, B.S., Kim, C.T., Park, B.H., Kwon, S., Cho, Y.J., Kim, N., Kim, C.J., Chun, J., Kwak, J., Maeng, J.S.
<2>Draft Genome Sequence of Staphylococcus saprophyticus subsp. saprophyticus M1-1,  Isolated from the Gills of a Korean Rockfish, Sebastes schlegeli Hilgendorf,  after High Hydrostatic Pressure Processing.
<3>J. Bacteriol.
<4>194
<5>4441-4442
<6>2012
<7>A bacterium designated M1-1 was isolated from the gills of a Korean rockfish, Sebastes
schlegeli Hilgendorf, after high hydrostatic pressure processing.
Studies of 16S rRNA phylogeny and comparative genomics demonstrated that the
isolate belongs to Staphylococcus saprophyticus subsp. saprophyticus. Here, we
report the draft genome sequence of S. saprophyticus subsp. saprophyticus M1-1
(KACC 16562).

<>

<1>Kim, B.Y., Kwon, O.S., Joo, S.A., Park, J.A., Heo, K.Y., Kim, M.S., Ahn, J.S.
<2>A column method for determination of DNA cytosine-C-5-methyltransferase activity.
<3>Anal. Biochem.
<4>326
<5>21-24
<6>2004
<7>DNA methylation at the 5th position of cytosine has been found to be correlated with
tumorigenesis. An inhibitor of DNA methylase could,
therefore, be used as an anticancer drug. However, only a few
inhibitory compounds have been discovered due to the limitations for
assaying the DNA methylation. In this study, we describe a modification
of DNA cytosine-C5-methyltransferase assay system utilizing
[H-3]-labeled S-adenosyl-methionine (SAM) and Sephadex G-25 column.
Pre-treatment of either lambda DNA or the promoter region of human
telomerase (hTERT) with HaeIII methylase greatly reduced the digestion
of the DNAs with the corresponding restriction enzyme HaeIII
endonuclease (over 100-fold), and the result was further confirmed by
agarose gel electrophoresis. Application of this column method to
another modification/restriction system, EcoRI methylase/endonuclease,
gave rise to the similar results. Our data suggest that the newly
developed column method could be effective for rapid screening of large
number of cytosine methylase inhibitors and could also be applicable to
other DNA methylases.

<>

<1>Kim, C.T., Kim, B.S., Kim, M.J., Park, B.H., Kwon, S., Maeng, H.Y., Kwak, J., Chun, J., Cho, Y.J., Kim, N., Kim, C.J., Maeng, J.S.
<2>Draft Genome Sequencing of Bacillus sp. Strain M2-6, Isolated from the Roots of Korean Ginseng, Panax ginseng C. A. Meyer, after High-Hydrostatic-Pressure  Processing.
<3>J. Bacteriol.
<4>194
<5>7003-7004
<6>2012
<7>A bacterium, designated M2-6, was isolated from Korean ginseng, Panax ginseng C.  A. Meyer,
roots after high-hydrostatic-pressure processing. On the basis of 16
rRNA gene phylogeny, the isolate was presumptively identified as a Bacillus sp.
Here we report the draft genome sequence of Bacillus sp. strain M2-6 (= KACC
16563).

<>

<1>Kim, D.-S., Jung, M.Y., Sin, Y., Kim, D.W., Paek, J., Kim, R.N., Park, I.S., Kook, J.K., Nam, S.H., Kim, A., Kang, A., Park, H.S., Choi, S.H., Chang, Y.H.
<2>Genome Sequence of the Anaerobic Bacterium Clostridium arbusti SL206T.
<3>J. Bacteriol.
<4>194
<5>2758
<6>2012
<7>A new Clostridium species has been isolated from pear orchard soil in Daejeon, Republic of
Korea. The isolate, Clostridium arbusti SL206(T) (KCTC 5449(T)), showed a nitrogenase activity
as well as an organic acid production. Here we first report the draft genome sequence of a
novel species in the genus Clostridium within the largest Gram-positive group.

<>

<1>Kim, D.H., Jiang, S., Lee, J.H., Cho, Y.J., Chun, J., Choi, S.H., Park, H.S., Hur, H.G.
<2>Draft Genome Sequence of Shewanella sp. Strain HN-41, Which Produces Arsenic-Sulfide Nanotubes.
<3>J. Bacteriol.
<4>193
<5>5039-5040
<6>2011
<7>The dissimilatory metal reducing bacterium Shewanella sp. strain HN-41 was first reported to
produce novel photoactive As-S nanotubes via reduction
of As(V) and S(2)O(3)(2-) under anaerobic conditions. Here we report the
draft genome sequence and annotation of strain HN-41.

<>

<1>Kim, D.S., Choi, S.H., Kim, D.W., Kim, R.N., Nam, S.H., Kang, A., Kim, A., Park, H.S.
<2>Genome Sequence of Leuconostoc gelidum KCTC 3527 Isolated from Kimchi.
<3>J. Bacteriol.
<4>193
<5>799-800
<6>2010
<7>Leuconostoc gelidum KCTC 3527 is found mainly in vegetables and plays an important role in
vegetable fermentation, including that of Korean traditional kimchi. Here we announce the
draft genome sequence of Leuconostoc gelidum KCTC 3527, isolated from Korean traditional
kimchi and describe major findings from its annotation.

<>

<1>Kim, D.S., Choi, S.H., Kim, D.W., Kim, R.N., Nam, S.H., Kang, A., Kim, A., Park, H.S.
<2>Genome Sequence of Leuconostoc inhae KCTC 3774 Isolated from Kimchi.
<3>J. Bacteriol.
<4>193
<5>1278-1279
<6>2010
<7>The Leuconostoc inhae KCTC 3774 strain is a gram-positive, non-spore-forming,
heterofermentative, spherical or lenticular lactic acid bacteria. Here we announce the draft
genome sequence of Leuconostoc inhae KCTC 3774, isolated from traditional Korean kimchi and
describe major findings from its annotation.

<>

<1>Kim, D.S., Choi, S.H., Kim, D.W., Kim, R.N., Nam, S.H., Kang, A., Kim, A., Park, H.S.
<2>Genome Sequence of Lactobacillus cypricasei KCTC 13900.
<3>J. Bacteriol.
<4>193
<5>5053-5054
<6>2011
<7>Lactobacillus cypricasei KCTC 13900 is important in the generation of particular flavours and
in other ripening processes associated with
specific cheeses. Here we announce the draft genome sequence of
Lactobacillus cypricasei KCTC 13900, isolated from cheeses and describe
major findings from its annotation.

<>

<1>Kim, D.S., Choi, S.H., Kim, D.W., Kim, R.N., Nam, S.H., Kang, A., Kim, A., Park, H.S.
<2>Genome Sequence of Lactobacillus versmoldensis KCTC 3814.
<3>J. Bacteriol.
<4>193
<5>5589-5590
<6>2011
<7>Lactobacillus versmoldensis KCTC 3814 was isolated from raw fermented poultry salami. The
species was present in high numbers and frequently
dominated the lactic acid bacteria (LAB) populations of the products.
Here, we announce the draft genome sequence of Lactobacillus versmoldensis
KCTC 3814, isolated from poultry salami, and describe major findings from
its annotation.

<>

<1>Kim, D.S., Choi, S.H., Kim, D.W., Nam, S.H., Kim, R.N., Kang, A., Kim, A., Park, H.S.
<2>Genome Sequence of Weissella cibaria KACC 11862.
<3>J. Bacteriol.
<4>193
<5>797-798
<6>2010
<7>The Weissella cibaria KACC 11862 is Gram-positive heterofermentative Leuconostoc-like lactic
acid bacteria and it is widely distributed in
Korean traditional foods such as kimchi. Here we report the draft genome
sequence of the type strain Weissella cibaria KACC 11862 (1,599 known
gene, 80 RNA), which consists of 72 large contigs (>100 bp in size).

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<1>Kim, D.S., Jung, M.Y., Kang, A., Cho, J., Sin, Y., Paek, J., Kim, D.W., Kim, R.N., Nam, S.H., Kim, A., Park, H.S., Choi, S.H., Chang, Y.H.
<2>Genome Sequence of Peptoniphilus rhinitidis 1-13T, an Anaerobic Coccus Strain Isolated from Clinical Specimens.
<3>J. Bacteriol.
<4>194
<5>2405-2406
<6>2012
<7>A new Peptoniphilus species has been isolated from samples from a patient who was scheduled
for endoscopic sinus surgery for chronic rhinosinusitis. The isolate,
Peptoniphilus rhinitidis 1-13(T) (KCTC 5985(T)), can use peptone as a sole carbon
source and produce butyrate as a metabolic end product. This is the first report
of the draft genome sequence of a novel species in the genus Peptoniphilus within
the group of Gram-positive anaerobic cocci.

<>

<1>Kim, D.S., Kang, Y.K., Yoo, O.J.
<2>Effects of base analog substitutions in the sequence, CCGG, on the cleavage and methylation reactions of HpaII and MspI endonucleases and their cognate methylases.
<3>Biochem. Mol. Biol. Int.
<4>32
<5>507-514
<6>1994
<7>HpaII endonuclease, HpaII methylase, MspI endonuclease, and MspI methylase were used to
investigate their specific interactions with the common recognition sequence, CCGG. Six
derivatives of the oligonucleotide, AGCCCGGGCT, containing a variety of single base analog
substitutions within the tetrameric recognition core were synthesized. Steady state kinetic
values for the reactions of all 4 enzymes with these oligonucleotide substrates were obtained.
Our data suggest that there are close contacts between the C5 positions of both cytosine
residues and the enzymes except that MspI endonuclease can accommodate a methyl group at the
C5 position of the second cytosine residue. The data also showed that minor groove
interactions between the 2-amino group of both G residues and the HpaII or MspI endonuclease
were essential for activity. However, these interactions were not essential for methylase
activity except that the oliogonucleotide substituted with inosine nucleotide at the first G
position did not react with MspI methylase.

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<1>Kim, D.S., Paek, J., Shin, J.H., Kim, D.W., Jung, M.Y., Kim, R.N., Sin, Y., Kook, J.K., Nam, S.H., Kim, A., Kang, A., Park, H.S., Choi, S.H., Chang, Y.H.
<2>Genome Sequence of Myroides injenensis M09-0166T, Isolated from Clinical Specimens.
<3>J. Bacteriol.
<4>194
<5>2748-2749
<6>2012
<7>A new Myroides species has been isolated from the urine of a patient with fever in spite of
multiple antibiotic treatments who had undergone a radical
hysterectomy for cervical cancer and percutaneous nephrostomies for
hydronephrosis in the past. The isolate, Myroides injenensis M09-0166(T) (KCTC
23367(T)), showed a high level of resistance to multiple antibiotic agents. Here
we provide the first report of the draft genome sequence of a novel species in
the genus Myroides within the nonfermenting Gram-negative group.

<>

<1>Kim, D.S., Sin, Y., Kim, D.W., Paek, J., Kim, R.N., Jung, M.Y., Park, I.S., Kim, A., Kang, A., Park, H.S., Choi, S.H., Chang, Y.H.
<2>Genome Sequence of the Probiotic Bacterium Sporolactobacillus vineae SL153T.
<3>J. Bacteriol.
<4>194
<5>3015-3016
<6>2012
<7>The novel Sporolactobacillus vineae SL153(T) strain has excellent intestinal adherence and
growth inhibitory effect on pathogenic microorganisms, including
Vibrio genus microorganisms, and therefore can be effectively used for the
prevention and treatment of disease caused by pathogenic microorganisms. Here, we
first report the draft genome sequence of a novel species in the genus
Sporolactobacillus.

<>

<1>Kim, D.W., Choi, S.H., Kang, A., Nam, S.H., Kim, D.S., Kim, R.N., Kim, A., Park, H.S.
<2>Draft Genome Sequence of Lactobacillus mali KCTC 3596.
<3>J. Bacteriol.
<4>193
<5>5037
<6>2011
<7>We announce the draft genome sequence of the type strain Lactobacillus mali KCTC 3596
(2,652,969 bp, with a G+C content of 36.0%) that one of the
most prevalent lactic acid bacteria present during the manufacturing
process of Apple juice, which consists of 122 large contigs (>100 bp in
size). All of the contigs were assembled by Newbler Assembler 2.3 (454
Life Science).

<>

<1>Kim, D.W., Choi, S.H., Kang, A., Nam, S.H., Kim, D.S., Kim, R.N., Kim, A., Park, H.S.
<2>Draft Genome Sequence of Lactobacillus zeae KCTC 3804.
<3>J. Bacteriol.
<4>193
<5>5023
<6>2011
<7>We announce the draft genome sequence of the type strain Lactobacillus zeae KCTC 3804
(3,110,326 bp, with a G+C content of 47.8%), which is one
of the most prevalent lactic acid bacteria present during the processing
of raw cow's milk. The genome consists of 113 large contigs (>100 bp). All
of the contigs were assembled by Newbler Assembler 2.3 (454 Life Science).

<>

<1>Kim, D.W., Choi, S.H., Kang, A., Nam, S.H., Kim, D.S., Kim, R.N., Kim, A., Park, H.S.
<2>Draft Genome Sequence of Lactobacillus malefermentans KCTC 3548.
<3>J. Bacteriol.
<4>193
<5>5537
<6>2011
<7>We announce the draft genome sequence of the type strain Lactobacillus malefermentans KCTC
3548 (2,003,922 bp, with a G+C content of 41.1%),
which is one of the most prevalent lactic acid bacteria present during the
manufacturing process of beer; the genome consists of 172 large contigs
(>100 bp in size). All of the contigs were assembled by using Newbler
Assembler 2.3 (454 Life Science).

<>

<1>Kim, D.W., Choi, S.H., Kang, A., Nam, S.H., Kim, R.N., Kim, A., Kim, D.S., Park, H.S.
<2>Genome Sequence of Leuconostoc pseudomesenteroides KCTC 3652.
<3>J. Bacteriol.
<4>193
<5>4299
<6>2011
<7>We announce the genome sequence of one of the most prevalent lactic acid bacteria present
during the manufacturing process of cane juice, the type
strain Leuconostoc pseudomesenteroides KCTC 3652 (3,244,985 bp, with a G+C
content of 38.3%), which consists of 1,160 large contigs (>100 bp in
size). All of the contigs were assembled by the Newbler Assembler 2.3
software program (454 Life Sciences).

<>

<1>Kim, E.B., Tyler, C.A., Kopit, L.M., Marco, M.L.
<2>Draft Genome Sequence of Fructophilic Lactobacillus florum.
<3>Genome Announcements
<4>1
<5>e00025-12
<6>2013
<7>Herein we report the first genome sequence for Lactobacillus florum. L. florum 2F was isolated
from Valencia orange leaves and is fructophilic, like other strains of this species. The draft
genome of L. florum 2F contains 1,261,842 bp with a G+C content of 41.5% in 46 contigs (>/=500
bp).

<>

<1>Kim, E.K., Kim, S.H., Nam, H.J., Choi, M.K., Lee, K.A., Choi, S.H., Seo, Y.Y., You, H., Kim, B., Lee, W.J.
<2>Draft Genome Sequence of Gluconobacter morbifer G707T, a Pathogenic Gut Bacterium Isolated from Drosophila melanogaster Intestine.
<3>J. Bacteriol.
<4>194
<5>1245
<6>2012
<7>Gluconobacter morbifer G707(T), a minor member of gut microbiota, was isolated from fruit fly
(Drosophila melanogaster). Here, the draft genome sequence of
Gluconobacter morbifer G707(T) is reported.

<>

<1>Kim, E.K., Kim, S.H., Nam, H.J., Choi, M.K., Lee, K.A., Choi, S.H., Seo, Y.Y., You, H., Kim, B., Lee, W.J.
<2>Draft Genome Sequence of Commensalibacter intestini A911T, a Symbiotic Bacterium  Isolated from Drosophila melanogaster Intestine.
<3>J. Bacteriol.
<4>194
<5>1246
<6>2012
<7>Commensalibacter intestini A911(T), a predominant symbiotic bacterium capable of  stably
colonizing gut epithelia, was isolated from the fruit fly, Drosophila
melanogaster. Here we report the draft genome sequence of Commensalibacter
intestini A911(T).

<>

<1>Kim, E.K., Park, Y.M., Lee, O.Y., Lee, W.J.
<2>Draft Genome Sequence of Lactobacillus plantarum Strain WJL, a Drosophila Gut Symbiont.
<3>Genome Announcements
<4>1
<5>e00937-13
<6>2013
<7>Lactobacillus plantarum strain WJL, a member of the symbiotic gut bacteria, was isolated from
the intestine of the fruit fly, Drosophila melanogaster. Here, we
report the draft genome sequence of L. plantarum WJL.

<>

<1>Kim, E.L., Makhovich, O.P., Maliuta, S.S.
<2>Some differences between BamHI and BnaI, the restrictases with the same specificity.
<3>Biopol. Kletka
<4>5
<5>86-89
<6>1989
<7>The restriction endonuclease BnaI, being a BamHI true isoschizomer, is isolated from B. natto
B3364 strain. Its molecular weight, stability and optimal reaction conditions are studied.
Comparative investigations have shown that according to these physicochemical characteristics
the BamHI and BnaI enzymes differ from each other in spite of their analogous specificity.

<>

<1>Kim, E.L., Maliuta, S.S.
<2>A new isoschizomer, BnaI, of the BamHI restriction endonuclease.
<3>Gene
<4>80
<5>363-368
<6>1989
<7>By assaying the yield of phage SPO1 we have identified a new
restriction-modification activity in the Bacillus natto B3364 strain.  A class
II restriction endonuclease, BnaI, isolated from the crude extract of B3364
cells was shown to be a true isoschizomer of the BamHI endonuclease.  The Mr,
stability and optimal conditions required for DNA digestion were determined for
BnaI.  Although both enzymes show the same specificity, BnaI and BamHI differ
from each other in all the properties specified above.

<>

<1>Kim, E.L., Maliuta, S.S.
<2>Purification of a DNA methyltransferase from Bacillus natto B3364.
<3>FEBS Lett.
<4>255
<5>361-364
<6>1989
<7>An S-adenosyl-L-methionine: DNA-methyltransferase, termed M.BnaI, was purified from Bacillus
natto B3364 strain by successive column chromatography. The molecular weight determined by gel
filtration was 37 kDa for M.BnaI. Analysis of methyltransferase by sodium dodecyl
sulfate-polyacrylamide gel electrophoresis showed correspondence of the M.BnaI activity with
one protein band at a molecular weight of 35 kDa. Sequencing of pUC19 DNA methylated with
M.BnaI showed the cytosine-5 methylation in the BnaI recognition sequence GGAT^CC at the
position indicated by the arrow.

<>

<1>Kim, E.L., Malyuta, S.S.
<2>BnaI a new isoshisomer of the restrictase BamHI.
<3>Biotekhnologiya
<4>4
<5>24-27
<6>1986
<7>Studies of restricted reproduction of phage SPO1 have disclosed a restriction-modification
system in the strain Bacillus natto B3364. Restriction endonuclease BnaI of class II has been
isolated and purified. BnaI has been shown to be a true isoschizomer of the restrictase BamHI.

<>

<1>Kim, G.-D., Ni, J., Kelesoglu, N., Roberts, R.J., Pradhan, S.
<2>Co-operation and communication between the human maintenance and de novo DNA (cytosine-5) methyltransferases.
<3>EMBO J.
<4>21
<5>4183-4195
<6>2002
<7>Three different families of DNA (cytosine-5) methyltransferases, DNMT1, DNMT3a and DNMT3b,
participate in establishing and maintaining genomic methylation patterns during mammalian
development. These enzymes have a large N-terminal domain fused to a catalytic domain. The
catalytic domain is homologous to prokaryotic (cytosine-5) methyltransferases and contains the
catalytic PC dipeptide, while the N-terminus acts as a transcriptional repressor by recruiting
several chromatin remodeling proteins. Here, we show that the human de novo enzymes hDNMT3a
and hDNMT3b form complexes with the major maintenance enzyme hDNMT1. Antibodies against hDNMT1
pull down both the de novo enzymes. Furthermore, the N-termini of the enzymes are involved in
protein-protein interactions. Immunocytochemical staining revealed mostly nuclear
co-localization of the fusion proteins, with the exception of hDNMT3a, which is found either
exclusively in cytoplasm or in both nucleus and cytoplasm. Pre-methylated substrate DNAs
exhibited differential methylation by de novo and maintenance enzymes. In vivo co-expression
of hDNMT1 and hDNMT3a or hDNMT3b leads to methylation spreading in the genome, suggesting
co-operation between de novo and maintenance enzymes during DNA methylation.

<>

<1>Kim, H., Jeong, W., Jeoung, H.Y., Song, J.Y., Kim, J.S., Beak, J.H., Parisutham, V., Lee, S.K., Kim, J.W., Kim, J.Y., Jung, S.C., Her, M., An, D.J.
<2>Complete Genome Sequence of Brucella abortus A13334, a New Strain Isolated from the Fetal Gastric Fluid of Dairy Cattle.
<3>J. Bacteriol.
<4>194
<5>5444
<6>2012
<7>Brucella abortus is a major pathogen that infects livestock and humans. A new strain of B.
abortus (A13334) was isolated from the fetal gastric fluid of a
dairy cow, with the aim of using it to compare genetic properties, analyze
virulence factor, and survey the epidemiological relationship to other Brucella
species. Here, we report the complete and annotated genome sequence of B. abortus
A13334.

<>

<1>Kim, H.H., Corina, L.E., Suh, J.K., Herrin, D.L.
<2>Expression, purification, and biochemical characterization of the intron-encoded endonuclease, I-CreII.
<3>Protein Expr. Purif.
<4>44
<5>162-172
<6>2005
<7>The ORF of the Cr.psbA4 intron of Chlamydomonas reinhardtii mediates efficient intron homing,
and contains an H-N-H and possibly a GIY-YIG
motif. The ORF was over-expressed in Escherichia coli without
non-native amino acids, but was mostly insoluble. However,
co-over-expression of E coli chaperonins GroEL/GroES solubilized
similar to 50% of the protein, which was purified by ion-exchange and
heparin-affinity chromatography. Biochemical characterization showed
that the protein is a double-strand-specific endonuclease that cleaves
fused psbA exon 4-exon 5 DNA, and was named I-CreII. I-CreII has a
relatively relaxed divalent metal ion requirement (Mg2+, Mn2+, Ca2+,
and Fe2+ supported cleavage), is insensitive to salt < 350 mM, and is
stabilized by DNA. Cleavage of target DNA occurs close (4 nt on the top
strand) to the intron-insertion site, and leaves 2-nt 3'-OH overhangs,
similar to GIY-YIG endonucleases. The boundaries of the recognition
sequence span similar to 30 bp, and encompass the cleavage and
intron-insertion sites. Cleavage of heterologous psbA DNAs indicates
the enzyme can tolerate multiple, but not all, substitutions in the
recognition site. This work will facilitate further study of this novel
endonuclease, which may also find use in site-specific manipulation of
chloroplast DNA.

<>

<1>Kim, H.J., Lee, J.H., Kang, B.R., Rong, X., McSpadden, G.B.B., Ji, H.J., Park, C.S., Kim, Y.C.
<2>Draft Genome Sequence of Pantoea ananatis B1-9, a Nonpathogenic Plant Growth-Promoting Bacterium.
<3>J. Bacteriol.
<4>194
<5>729
<6>2012
<7>Pantoea ananatis B1-9 is an endophytic Gram-negative rhizobacterium that was isolated for its
ability to promote plant growth and improve crop
yield in the field. Here we report the draft genome sequence of P.
ananatis B1-9. Comparison of this sequence to the sequenced genome of a
plant-pathogenic P. ananatis strain, LMG20103, indicated that the
pathogenesis-related genes were absent, but a subset of gene functions
that may be related to its plant growth promotion were present.

<>

<1>Kim, H.J., Park, J.Y., Han, S.H., Lee, J.H., Rong, X., McSpadden, G.B.B., Park, S.K., Kim, Y.C.
<2>Draft Genome Sequence of the Biocontrol Bacterium Chromobacterium sp. Strain C-61.
<3>J. Bacteriol.
<4>193
<5>6803-6804
<6>2011
<7>Chromobacterium sp. strain C-61 is a plant-associated bacterium with proven capacities to
suppress plant diseases. Here, we report the draft
genome sequence and automatic annotation of strain C-61. A comparison of
this sequence to the sequenced genome of Chromobacterium violaceum ATCC
12472 indicates the novelty of C-61 and a subset of gene functions that
may be related to its biocontrol activities.

<>

<1>Kim, H.J., Yano, A., Wada, Y., Sano, H.
<2>Properties of a tobacco DNA methyltransferase, NtMET1 and its involvement in chromatin movement during cell division.
<3>Ann. Bot.
<4>99
<5>845-856
<6>2007
<7>Background and Aims Plants possess three types of DNA methyltransferase, among which
methyltransferase type 1 (MET1) is
considered to play a major role by maintaining the CpG methylation
patterns. However, little information is available as to its enzymatic
activity, interacting proteins and spatial and temporal behaviours
during DNA replication. In the present study, one example, NtMET1 from
tobacco plants, was selected and an analysis was made of its
biochemical properties and cellular localization.
Methods NtMET1 was expressed in Sf9 insect cells, and a purified
sample was subjected to a standard in vitro methylation assay.
Intramolecular interaction was examined by the yeast two-hybrid and
pull-down assays. Transgenic tobacco plants (Nicotiana tabacum)
over-expressing NtMET1 were constructed via Agrobacterium-mediated
transformation. Cellular localization was examined by fluorescence
protein fusion, which was expressed in tobacco bright yellow 2 cells.
Key Results In vitro assays showed no detectable methylation
activity when both hemimethylated and unmethylated DNA samples were
used as the substrate. In planta assays with over-expressing transgenic
lines showed no hypermethylation but rather hypomethylation of genomc
DNA. The inability of methylation was conceivably due to a tight
intramolecular interaction between the N- and C-terminal regions with
the catalytic domain residing on the C-terminus being completely
masked. Cellular localization analyses indicated that NtMET1 localized
to the nucleus in the resting stage and migrates to the cytoplasm
during mitosis, particularly at metaphase. The pattern observed
resembled that of Ran GTPase, and in vitro pull-down assays showed a
clear interaction between NtMET1 and AtRAN3, an Arabidopsis orthologue
of tobacco Ran GTPase, NtRan-A1.
Conclusions The results suggest that enzymatic activity of NtMET1
is well adjusted by its own intra/intermolecular interaction and
perhaps by interactions with other proteins, one of which was found to
be Ran GTPase. Results also revealed that NtMET1 becomes localized to
the vicinity of chromatin with the aid of Ran GTPase during cell
division, and may play an important role in progress through mitosis
independently of methylation activity.

<>

<1>Kim, J., Jung, J., Sung, J.S., Chun, J., Park, W.
<2>Genome Sequence of Pectin-Degrading Alishewanella agri, Isolated from Landfill Soil.
<3>J. Bacteriol.
<4>194
<5>5135-5136
<6>2012
<7>Alishewanella agri BL06(T) (= KCTC 22400(T) = JCM 15597(T)) was isolated from landfill soil in
Pohang, South Korea. A. agri showed the ability to degrade
pectin, a structural heteropolysaccharide present in the cell wall of plants.
Here we report the genome sequence of Alishewanella agri BL06(T), the second
sequenced strain in the genus Alishewanella.

<>

<1>Kim, J., Kim, S.J., Kim, S.H., Kim, S.I., Moon, Y.J., Park, S.J., Kahng, H.Y., Chung, Y.H.
<2>Draft Genome Sequence of Sphingopyxis sp. Strain MWB1, a Crude-Oil-Degrading Marine Bacterium.
<3>Genome Announcements
<4>2
<5>e01256-14
<6>2014
<7>Sphingopyxis sp. strain MWB1, which is capable of degrading crude oil, diesel, and kerosene,
was isolated from crude oil-contaminated seashore in Tae-an, South
Korea. Here, we report the draft genome sequence of this strain, which comprises
3,118,428 bp with a G+C content of 62.85 mol%.

<>

<1>Kim, J., Roh, S.W., Bae, J.W.
<2>Draft Genome Sequence of Dietzia alimentaria 72T, Belonging to the Family Dietziaceae, Isolated from a Traditional Korean Food.
<3>J. Bacteriol.
<4>193
<5>6791
<6>2011
<7>Actinobacterial strain 72(T), named Dietzia alimentaria, which belongs to the family
Dietziaceae, was isolated from a traditional Korean food made
from clams. The draft genome sequence of D. alimentaria 72(T) contains
3,352,817 bp, with a G+C content of 67.34%.

<>

<1>Kim, J., Thorp, H.H.
<2>Footprinting of cytosine-5--methyltransferase by Pt2(pop)44-.
<3>ACS Abstracts
<4>212
<5>A290
<6>1996
<7>The photoreagent Pt2(pop)44- (1, pop = P2O5H22-) cleaves DNA upon visible irradiation via
abstraction of the 4' and 5' hydrogens, which provide a sequence-neutral cleavage ladder.
The development of 1 for use as an anionic DNA photocleavage agent has prompted us to apply it
to obtaining the footprint of the M.HhaI (cytosine-5) methyltransferase.  The binding of
M.HhaI to its DNA substrate causes large conformational changes with an extensive contact with
DNA around the active site.  A comparison of photocleavage with data from the crystal
structure of a reaction intermediate between the M.HhaI methyltransferase and a DNA
oligonucleotide shows a correlation between cleavage by 1 and the effects of electrostatic
forces in the binding of protein to nucleic acids.  Also underway is an anlaysis of the
photocleavage products to provide additional quantitation of the cleavage pathways.

<>

<1>Kim, J.-S., DeGiovanni, A., Jancarik, J., Adams, P.D., Yokota, H., Kim, R., Kim, S.-H.
<2>Crystal structure of DNA sequence specificity subunit of a type I restriction-modification enzyme and its functional implications.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>3248-3253
<6>2005
<7>Type I restriction-modification enzymes are differentiated from type II and type III enzymes
by their recognition of two specific dsDNA sequences separated by a given spacer and cleaving
DNA randomly away from the recognition sites. They are oligomeric proteins formed by three
subunits: a specificity subunit, a methylation subunit, and a restriction subunit. We solved
the crystal structure of a specificity subunit from Methanococcus jannaschii at 2.4 Angstroms
resolution. Two highly conserved regions (CRs) in the middle and at the C terminus form a
coiled-coil of long antiparallel alpha-helices. Two target recognition domains form globular
structures with almost identical topologies and two separate DNA binding clefts with a modeled
DNA helix axis positioned across the CR helices. The structure suggests that the coiled-coil
CRs act as a molecular ruler for the separation between two recognized DNA sequences.
Furthermore, the relative orientation of the two DNA binding clefts suggests kinking of bound
dsDNA and exposing of target adenines from the recognized DNA sequences.

<>

<1>Kim, J.F., Jeong, H., Lee, J.S., Choi, S.H., Ha, M., Hur, C.G., Kim, J.S., Lee, S., Park, H.S., Park, Y.H., Oh, T.K.
<2>The Complete Genome Sequence of Leuconostoc citreum KM20.
<3>J. Bacteriol.
<4>190
<5>3093-3094
<6>2008
<7>Leuconostoc citreum is one of the most prevalent lactic acid bacteria during the manufacturing
process of kimchi, the best-known Korean
traditional dish. Here we present the complete genome sequence of L.
citreum KM20. It consists of a 1.80-Mb chromosome and four circular
plasmids, and reveals genes likely involved in kimchi fermentation and its
probiotic effects.

<>

<1>Kim, J.F., Jeong, H., Park, S.Y., Kim, S.B., Park, Y.K., Choi, S.K., Ryu, C.M., Hur, C.G., Ghim, S.Y., Oh, T.K., Kim, J.J., Park, C.S., Park, S.H.
<2>Genome Sequence of the Polymyxin-Producing Plant-Probiotic Rhizobacterium Paenibacillus polymyxa E681.
<3>J. Bacteriol.
<4>192
<5>6103-6104
<6>2010
<7>Paenibacillus polymyxa E681, a spore-forming, low-G+C, Gram-positive bacterium isolated from
the rhizosphere of winter barley grown in South
Korea, has great potential for agricultural applications due to its
ability to promote plant growth and suppress plant diseases. Here we
present the complete genome sequence of P. polymyxa E681. Its 5.4-Mb
genome encodes functions specialized to the plant-associated lifestyle and
characteristics that are beneficial to plants, such as the production of a
plant growth hormone, antibiotics, and hydrolytic enzymes.

<>

<1>Kim, J.F., Jeong, H., Yu, D.S., Choi, S.H., Hur, C.G., Park, M.S., Yoon, S.H., Kim, D.W., Ji, G.E., Park, H.S., Oh, T.K.
<2>Genome sequence of the probiotic bacterium Bifidobacterium animalis subsp. lactis AD011.
<3>J. Bacteriol.
<4>191
<5>678-679
<6>2009
<7>Bifidobacterium animalis subsp. lactis is a probiotic bacterium that naturally inhabits the
guts of most mammals, including humans. Here we
report the complete genome sequence of B. animalis subsp. lactis AD011
that was isolated from an infant fecal sample. Biological functions
encoded in a single circular chromosome of 1,933,695 bp, smallest among
the completely sequenced bifidobacterial genomes, are suggestive of their
probiotic functions, such as utilization of bifidogenic factors and a
variety of glycosidic enzymes and biosynthesis of polysaccharides.

<>

<1>Kim, J.H., Boo, Y.B., Lee, K.Y.
<2>Preparation of restriction endonuclease-AluI from Arthrobacter Luteus.
<3>Korean J. Biochem.
<4>17
<5>149-154
<6>1985
<7>Extraction and purification of the restriction endonuclease, Alu-I, were
performed from Arthrobacter luteus.  The harvested cells were disrupted by the
treament of lysozyme and sonification, and the uspernant was precipitated with
70% saturation of ammonium sulfate.  The protein sediment was dissolved in
buffer solution and dialysed to the same buffer solution.  This solution was
successively applied to phosphocellulose, heparin-agarose, and Ultrogel AcA34
(LKB) column chromatography for the purification procedure.  The purified AluI
enzyme was revealed the singel band on SDS-polyacrylamide gel electrophoresis
and the molecular weight was shown 52 Kdal, attaining its specific activity to
312.5 units/mg or protein.  No difference was observed between our enzyme
preparation and commerical AluI (BRL) on agarose gel electropherogram.  The
enzyme sample after treatment with heparin-agarose also had multiprotein bands,
indicating the protein contaminant, however, all bands but a single band of
AluI enzyme disappeared by the subsequent column chromatography of Ultrogel
AcA34.

<>

<1>Kim, J.J., Koh, S., Kim, J.S., Lee, D.-S.
<2>Mode of action on EcoRI restriction endonuclease: EcoRI and EcoRI variant N199H have active monomeric forms.
<3>J. Biochem. Mol. Biol.
<4>31
<5>149-155
<6>1998
<7>The N199H variant of the EcoRI endonuclease has about twice the catalytic activity of the
wild-type.  A comparison of their biochemical characteristics, using synthetic
oligonucleotides 5'-dAAAACTTAAGAAAAAAAAAAA-3' (KA) and 5'-dTTTTTGAATTCTTTTTTTTTT-3' (KT),
helps to define the cleavage reaction pathway of these enzymes.  Both EcoRI and EcoRI variant
N199H were found to cleave single-stranded KA or KT about three times faster than the
double-stranded forms, although the KT oligonucleotide was more susceptible.  Using the ssDNA
substrate in kinetic analyses, lower Km values were obtained for the N199H variant than for
the wild-type at low (50 mM), as well as high (200 mM), sodium chloride concentrations.  This
difference between the endonucleases is attributed to a greater accessibility for the
substrate by the variant, and also a higher affinity for the DNA backbone.  It also appears
that the relative activities of the two enzymes, particularly at high ionic strength, are
proportional to their populations in the monomeric enzyme form.  That is, according to gel
filtration data, half of the N199H molecules exist as monomers in 200 mM NaCl, whereas those
of the wild-type are mainly dimeric.  Consequently, the Asp199 residue of the EcoRI
endonuclease may be implicated in the protein-protein interaction leading to dimerization, as
well as in coupling to DNA substrates.  In summary, it is proposed that active monomeric
endonuclease molecules, derived from the dimeric enzyme, recognize and form a complex with a
single stranded form of the DNA substrate, which then undergoes nucleophilic substitution and
cleavage.

<>

<1>Kim, J.J., Min, K.T., Kim, M.H., Augh, S.J., Kim, B.-D., Lee, D.-S.
<2>EcoRI variant N199H has enhanced specific activity.
<3>Gene
<4>171
<5>129-130
<6>1996
<7>The Asn199 residue of the EcoRI restriction endonuclease has been replaced with
other amino acids to investigate whether it mediates nucleotide recognition or catalytic
activity.
Cassette mutagenesis gave variants of EcoRI: N199D, N199H, N199L, N199R, N199S and
N199V.  Their relative cleavage rates were found to be in the following order: N199H>EcoRI
(wild type; wt)>N199L>N199V>N199S>N199R>N199D.  In particular, EcoRI variant N199H
showed about a two-fold higher specific activity than that of the wt enzyme.

<>

<1>Kim, J.K., Samaranayake, M., Pradhan, S.
<2>Epigenetic mechanisms in mammals.
<3>Cell. Mol. Life Sci.
<4>66
<5>596-612
<6>2009
<7>DNA and histone methylation are linked and subjected to mitotic inheritance in mammals. Yet
how methylation is propagated and maintained between successive cell divisions is not fully
understood. A series of enzyme families that can add methylation marks to cytosine
nucleobases, and lysine and arginine amino acid residues has been discovered. Apart from
methyltransferases, there are also histone modification enzymes and accessory proteins, which
can facilitate and/or target epigenetic marks. Several lysine and arginine demethylases have
been discovered recently, and the presence of an active DNA demethylase is speculated in
mammalian cells. A mammalian methyl DNA binding protein MBD2 and de novo DNA methyltransferase
DNMT3A and DNMT3B are shown experimentally to possess DNA demethylase activity. Thus, complex
mammalian epigenetic mechanisms appear to be dynamic yet reversible along with a
well-choreographed set of events that take place during mammalian development.

<>

<1>Kim, J.S., Jeong, W., Jeoung, H.Y., Song, J.Y., Kim, H., Beak, J.H., Parisutham, V., Lee, S.K., Kim, J.W., Kim, J.Y., Jung, S.C., Her, M., An, D.J.
<2>Complete Genome Sequence of Brucella canis Strain HSK A52141, Isolated from the Blood of an Infected Dog.
<3>J. Bacteriol.
<4>194
<5>5134
<6>2012
<7>Brucella canis infection can be clinically inapparent in dogs, and when infection goes
unnoticed, there is a chance for dog-to-human transmission. A new strain of
B. canis was isolated from the blood of an infected dog in order to analyze the
pathogenic mechanism, compare genetic properties, and develop new genetic tools
for early diagnosis of canine brucellosis. Herein, we report the complete genome
sequence of the strain B. canis HSK A52141. This is the second complete genome
sequence and biological annotation available for a member of B. canis.

<>

<1>Kim, J.S., Kim, J., Jeon, S.-E., Kim, S.-J., Kim, N.-O., Hong, S., Kang, Y.-H., Han, S., Chung, G.T.
<2>Complete nucleotide sequence of the IncI1 plasmid pSH4469 encoding CTX-M-15 extended-spectrum beta-lactamase in a clinical isolate of Shigella sonnei from an outbreak in the Republic of Korea.
<3>Int. J. Antimicrob. Agents
<4>44
<5>533-537
<6>2014
<7>An outbreak of extended-spectrum -lactamase (ESBL)-producing Shigella sonnei infections
occurred in
a school for disabled children in Gyeongbuk Province, Republic of Korea, in 2008. Five
students were
affected. Pulsed-field gel electrophoresis (PFGE) analysis revealed that all of the
ESBL-producing S. sonnei
isolates belonged to the same clone, and nucleotide sequence analysis of ESBL genes revealed
that they
harboured blaCTX-M-15. This is the first identification of blaCTX-M-15 in Shigella spp. in
South Korea. In this
study, a plasmid carrying the blaCTX-M-15 gene, designated pSH4469, recovered from a S. sonnei
isolate
responsible for the outbreak was characterised. Replicon typing and plasmid multilocus
sequence typing
(pMLST) analysis of plasmids in the outbreak strain identified that the blaCTX-M-15 gene was
located on
an IncI1 incompatibility group plasmid of sequence type 16 (ST16). The complete nucleotide
sequence
of pSH4469 revealed that this plasmid is 91109 bp and harbours 119 putative genes, including
another
antibiotic resistance gene (blaTEM-1b) that is often associated with the
ISEcp1-blaCTX-M-15-orf477delta
transposable unit. The plasmid consists of a large backbone with considerable homology to the
pEK204
plasmid isolated from Escherichia coli in the UK, except for insertion of an IS66 element
found in pEK204.
These data demonstrate that IncI1 plasmids are used as a successful platform for efficient
horizontal gene
transfer, thereby resulting in the dissemination of CTX-M-type -lactamases among
Enterobacteriaceae.

<>

<1>Kim, J.S., Li, J., Barnes, I.H.A., Thompson, S.A.
<2>Cj1461 DNA methyltransferase in Campylobacter jejuni 81-176 is involved in the regulation of virulence characteristics and bacterial cellular functions.
<3>Zoonoses Public Health
<4>54
<5>103
<6>2007
<7>DNA methylases such as Dam can regulate transcription of virulence-associated genes in
bacteria.  Cj1461 is a DNA methyltransferase conserved in Campylobacter species.  A
cj1461-insertional mutant of C. jejuni 81-176 had significantly reduced motility at 37C, and
the reduced motility was partially but significantly recovered in a strain complemented with
native cj1461 (P<0.05).  A Cj1461-overexpression strain of 81-176 was constructed by
expressing an inducible cj1461 gene on the shuttle vector pRY111.  cj1461 and cj1462 (flgI)
were about 1.5- and 1.8-fold overexpressed, respectively, in the Cj1461-overexpression strain
based on real-time PCR analysis.  In addition, the overexpression strain showed significantly
increased motility (P<0.05) compared to 81-176 containing pRY111 alone.  Proteome analysis of
the Cj1461-overexpression strain showed the altered expression of more than 20 proteins
including PEB1a, PEB3, PEB4, major outer membrane protein, gamma-glutamyl transpeptidase,
thiol peroxidase, alkyl hydroperoxide reductase, trigger factor and several TCA cycle enzymes.
These data, along with the altered regulation of numerous C. jejuni genes in the cj1461
mutant, suggest that Cj1461 DNA methyltransferase is involved in regulating virulence
characteristics such as motility and other bacterial cellular processes in C. jejuni 81-176.

<>

<1>Kim, J.S., Li, J.Q., Barnes, I.H.A., Baltzegar, D.A., Pajaniappan, M., Cullen, T.W., Trent, M.S., Burns, C.M., Thompson, S.A.
<2>Role of the Campylobacter jejuni cj1461 DNA methyltransferase in regulating virulence characteristics.
<3>J. Bacteriol.
<4>190
<5>6524-6529
<6>2008
<7>Mutation of the cj1461 predicted methyltransferase gene reduced the motility of Campylobacter
jejuni 81-176. Electron microscopy revealed
that the mutant strain had flagella but with aberrant structure. The
Delta cj1461 mutant was sevenfold more adherent to but 50-fold less
invasive of INT-407 human epithelial cells than the wild type.

<>

<1>Kim, J.W., Dutta, V., Elhanafi, D., Lee, S., Osborne, J.A., Kathariou, S.
<2>A Novel Restriction-Modification System Is Responsible for Temperature-Dependent Phage Resistance in Listeria monocytogenes ECII.
<3>Appl. Environ. Microbiol.
<4>78
<5>1995-2004
<6>2012
<7>Listeria monocytogenes epidemic clone II (ECII) strains are unusual in being completely
resistant to phage when grown at low temperatures (<=
30 degrees C). In the current study we constructed and characterized a
mariner-based mutant (J46C) of the ECII strain H7550-Cd-S that lacked
temperature-dependent resistance to phage. The transposon was localized
in LMOh7858 2753 (open reading frame [ORF] 2753), a member of a 12-ORF
genomic island unique to ECII strains. ORF 2753 and ORF 2754 exhibited
homologies to restriction endonucleases and methyltransferases
associated with type II restriction-modification (RM) systems. In
silico-based predictions of the recognition site for this putative RM
system were supported by resistance of DNA from ECII strains to
digestion by BfuI, a type II restriction enzyme specific for GTATCC
(N6/5). Similarly to J46C, a mutant harboring an in-frame deletion of
ORF 2753 was susceptible to phage regardless of temperature of growth
(25 C or 37 degrees C). Genetic complementation restored phage
resistance in 25 degrees C-grown cells of ORF 2753 mutants. Reverse
transcription (RT) and quantitative real-time PCR data suggested
enhanced transcription of ORF 2753 at low temperatures (<= 25 degrees
C) compared to 37 degrees C. In contrast, available transcriptional
data suggested that the putative methyltransferase (ORF 2754) was
constitutively expressed at all tested temperatures (4 to 37 degrees
C). Thus, temperature-dependent resistance of L. monocytogenes ECII to
phage is mediated by temperature-dependent expression of the
restriction endonuclease associated with a novel RM system (LmoH7)
unique to this epidemic clone.

<>

<1>Kim, J.Y., Wang, Y., Park, M.S., Ji, G.E.
<2>Improvement of Transformation Efficiency Through In Vitro Methylation and SacII Site Mutation of Plasmid Vector in Bifidobacterium longum MG1.
<3>J. Microbiol. Biotechnol.
<4>20
<5>1022-1026
<6>2010
<7>The different cleavage patterns of pYBamy59 plasmid isolated from E. coli DH5 alpha and B.
longum MG1 by the cell extract of B. longum MG1
suggested that the main reason for its low transformation efficiency
was related to the restriction modification (R-M) system. To confirm
the correlation between the R-M system and transformation efficiency,
in vitro methylation and site-directed mutagenesis were performed in
pYBamy59. Sequence analysis of pYBamy59 fragments digested by the cell
extract of B. longum MG1 revealed that all fragments were generated by
restriction of the sequence recognized by SacII endonuclease. When
pYBamy59 from E. coli was methylated in vitro by CpG or GpC
methyltransferase, it was protected from SacII digestion. Site-directed
mutagenesis, which removed SacII sites from pYBamy59, or in vitro
methylation of pYBamy59 showed 8- to 15-fold increases in the
transformation efficiency over intact pYBamy59. Modification of the
Sad-related R-M system in B. longum MG1 and in vitro methylation in
pYBamy 59 can improve the transformation efficiency in this strain. The
results showed that the R-M system is a factor to limit introduction of
exogenous DNA, and in vitro modification is a convenient method to
overcome the barrier of the R-M system for transformation.

<>

<1>Kim, K.K., Lee, K.C., Jeong, H., Stevens, D.A., Lee, J.S.
<2>Draft Genome Sequence of the Human Pathogen Halomonas stevensii S18214T.
<3>J. Bacteriol.
<4>194
<5>5143
<6>2012
<7>Halomonas stevensii is a Gram-negative, moderately halophilic bacterium causing environmental
contamination and infections in a dialysis center. Here we present
the 3.7-Mb draft genome sequence of the type strain (S18214(T)) of H. stevensii,
which will give insight into the pathogenic potential of H. stevensii.

<>

<1>Kim, K.K., Lee, K.C., Lee, J.S.
<2>Draft Genome Sequence of the Extremely Halophilic Archaeon Halogranum salarium B-1T.
<3>J. Bacteriol.
<4>194
<5>6659
<6>2012
<7>Halogranum salarium is an extremely halophilic archaeon isolated from evaporitic  salt
crystals and belongs to the family Halobacteriaceae. Here, we present the
4.5-Mb draft genome sequence of the type strain (B-1(T)) of H. salarium. This is
the first report of the draft genome sequence of a haloarchaeon in the genus
Halogranum.

<>

<1>Kim, M., Kim, S., Ryu, S.
<2>Complete Genome Sequence of Bacteriophage SSU5 Specific for Salmonella enterica serovar Typhimurium Rough Strains.
<3>J. Virol.
<4>86
<5>10894
<6>2012
<7>Salmonella enterica serovar Typhimurium rough strain-specific phage SSU5 was
isolated, and its whole genome was sequenced. The 103,229-bp-long double-stranded
DNA genome of SSU5 encodes 130 open reading frames with one tRNA for asparagine.
Genomic analysis revealed that SSU5 might be the phylogenetic origin of cryptic
plasmid pHCM2 harbored by Salmonella Typhi CT18.

<>

<1>Kim, M., Ryu, S.
<2>Antirepression System Associated with the Life Cycle Switch in the Temperate Podoviridae Phage SPC32H.
<3>J. Virol.
<4>87
<5>0
<6>2013
<7>Prophages switch from lysogenic to lytic mode in response to the host SOS response. The
primary factor that governs this
switch is a phage repressor, which is typically a host RecA-dependent autocleavable protein.
Here, in an effort to reveal the
mechanism underlying the phenotypic differences between the Salmonella temperate phages SPC32H
and SPC32N, whose
genome sequences differ by only two nucleotides, we identified a new class of Podoviridae
phage lytic switch antirepressor
that is structurally distinct from the previously reported Sipho- and Myoviridae phage
antirepressors. The SPC32H repressor
(Rep) is not cleaved by the SOS response but instead is inactivated by a small antirepressor
(Ant), the expression of
which is negatively controlled by host LexA. A single nucleotide mutation in the consensus
sequence of the LexA-binding
site, which overlaps with the ant promoter, results in constitutive Ant synthesis and
consequently induces SPC32N to enter
the lytic cycle. Numerous potential Ant homologues were identified in a variety of putative
prophages and temperate Podoviridae
phages, indicating that antirepressors may be widespread among temperate phages in the order
Caudovirales to
mediate a prudent prophage induction.

<>

<1>Kim, M., Yi, H., Cho, Y.J., Jang, J., Hur, H.G., Chun, J.
<2>Draft Genome Sequence of Escherichia coli W26, an Enteric Strain Isolated from Cow Feces.
<3>J. Bacteriol.
<4>194
<5>5149-5150
<6>2012
<7>An enteric bacterium, Escherichia coli W26 (KACC 16630), was isolated from feces  from a
healthy cow in South Korea. Here, we report the draft genome sequence of
the isolate, which is closely affiliated with commensal strains belonging to E.
coli phylogroup B1.

<>

<1>Kim, M.S., Whon, T.W., Roh, S.W., Shin, N.R., Bae, J.W.
<2>Draft Genome Sequence of Bacteroides faecis MAJ27T, a Strain Isolated from Human Feces.
<3>J. Bacteriol.
<4>193
<5>6801-6802
<6>2011
<7>Despite the ecological importance of the dominant gut bacteria Bacteroides, few genomes have
been defined. The Gram-negative, strictly
anaerobic intestinal bacterium Bacteroides faecis MAJ27(T) was isolated
from the feces of a healthy adult. Here, the draft genome sequence of the
type strain B. faecis MAJ27 (6.11 Mbp) is reported.

<>

<1>Kim, R., Modrich, P., Kim, S.-H.
<2>Interactive recognition in EcoRI restriction enzyme-DNA complex.
<3>Nucleic Acids Res.
<4>12
<5>7285-7292
<6>1984
<7>A solution study of interaction between DNA and EcoRI restriction enzyme shows
that there is a definite distortion of DNA in the specific recognition
complexes but no measurable DNA distortion in the non-specific interaction.

<>

<1>Kim, S., Lee, C., Lim, B., Sung, B., Yu, B., Lee, W., Lee, J., Lee, S.
<2>Linear DNA fragment for markerless deletion, novel strain having inhibited formation of biofilm and preparation method thereof.
<3>Korean Patent Office
<4>KR 1020040070017 A
<5>
<6>2004
<7>
<>

<1>Kim, S., Lee, W., Yu, B., Sung, B., Kim, J., Lee, C., Lee, J.
<2>Linear DNA fragment for developing a novel strain removed a specific region of chromosome and method for preparing the same.
<3>Korean Patent Office
<4>KR 1020020065351 A
<5>
<6>2002
<7>
<>

<1>Kim, S.C., Podhajska, A.J., Szybalski, W.
<2>Cleaving DNA at any predetermined site with adapter-primers and class-IIS restriction enzymes.
<3>Science
<4>240
<5>504-506
<6>1988
<7>A four-component system has been designed that makes it possible to prepare a
double-stranded (ds) DNA fragment; one fragment end is predesigned (by the use
of a class-IIS restriction enzyme and adapter-primer), and the other end
corresponds to any normal restriction cut.  The system is composed of the phage
M13mp7 single stranded (ss) target DNA; the Fok I restriction enzyme; an
oligodeoxynucleotide adapter-primer, which permits one to introduce Fok I cuts
at any specified site in the target DNA; and DNA polymerase, which converts the
ss target into a ds form ready for cloning.  In this system, the
oligodeoxynucleotide adapter-primer serves several purposes.  The 5' hairpin ds
domain of the adapter-primer  contains a Fok I recognition site.  Its 3' ss
domain selects a complementary site on the target ss DNA, hybridizes with it to
form the ds cleavage site, and serves as a primer to convert the ss M13mp7
target to ds DNA.

<>

<1>Kim, S.C., Posfai, G., Szybalski, W.
<2>A novel gene-fusing vector: construction of a 5'-GGmCC-specific chimeric methyltransferase, M.BspRI/M.BsuRI.
<3>Gene
<4>100
<5>45-50
<6>1991
<7>A vector was designed to allow predetermined and precise fusion between two
genes by constructing a cassette with two unique class-IIS restriction sites,
5'-ACCTGC3' (BspMI) and 5'-CCGGATG-3' (FokI overlapping with MspI), arranged
back-to-back in a divergent manner and inserted at the HincII site of a
multiple cloning site (MCS) in plasmid pUC18 or analogous vehicle.  Two DNA
fragments or genes to be precisely fused are cloned into the MCS parts located
on each side of the cassette containing the two unique class-IIS restriction
sites.  The BspMI and MspI/FokI sites are used to generate unidirectional
deletions of the genes as previously described (Hasan et al., Gene 50(1986)
55-62; Posfai and Szybalski, Nucleic Acids Res. 16(1988)6245).  The precisely
trimmed genes are ligated after the cassette containing the unique class-IIS
restriction sites are excised with BspMI + FokI and the termini were blunted
with mung-bean nuclease.  This method was used to construct a hybrid
methyltransferase (MTase) from the M.BspRI and M.BsuRI MTases, which share a
high degree of overall homology (about 65%) and have the identical sequence
specificity (5'-GGmCC-3').  A hybrid MTase composed of the N-terminal part of
M.BspRI and the C-terminal part of M.BsuRI was constructed and found to be
fully functional.

<>

<1>Kim, S.C., Skowron, P.M., Szybalski, W.
<2>Structural requirements for FokI-DNA interaction and oligodeoxyribonucleotide- instructed cleavage.
<3>J. Mol. Biol.
<4>258
<5>638-649
<6>1996
<7>The FokI restriction endonuclease recognizes the double-stranded (ds) 5'-GGATG-3'
site and cuts at the 9th and 13th nucleotides downstream from the 5'-3' and 3'-5'
strands, respectively.  To elucidate the interaction between FokI and DNA, and the effect of
Mg2+
on this interaction, we used FokI with various combinations of dsDNA, single-stranded (ss) DNA
and oligodeoxyribonucleotides (oligos) containing a double-stranded hairpin carrying the FokI
recognition site.  Oligo- and dsDNA-FokI interactions showed that for fully effective
recognition,
two or more base-pairs were required outside the 5'-GGATG-3' site.  When using FokI with
ssDNA and oligos, precise cutting with no observable byproducts was observed at the 9th or
13th
nucleotide.  This was independent of whether the region between the recognition and cut sites
was
perfectly complementary or whether there were up to four mismatches in this region, or a
single
mismatch within the cut site.  Moreover, FokI cleavage, when followed by step-wise filling-in
of
FokI cohesive ends in the dsDNA, allowed FokI to recleave such sites when two or more
nucleotides were added, releasing 2-mer, 3-mer, or 4-mer single-stranded chains.
Electrophoretic
mobility shift assays showed that the DNA helix was bent when complexed with FokI (without
Mg2+).  Such a complex, when formed in the absence of Mg2+, did not accept the subsequently
added Mg2+ for several minutes.  This suggests a tight, diffusion-resistant contact between
the
enzyme and the cognate DNA sequence.  In the presence of Mg2+, the half-life of the complex
FokI and dsDNA was 12 minutes at 22oC.  In the absence of Mg2+, such a complex, possessing a
terminally located 5'-GGATG-3' site, had a half-life of 1.5 to 2 minutes.  However, if
magnesium
ions were present, this complex had a stability similar to that of a complex formed with dsDNA
containing a centrally located 5'-GGATG-3' site.

<>

<1>Kim, S.J., Shin, S.C., Hong, S.G., Lee, Y.M., Choi, I.G., Park, H.
<2>Genome sequence of a novel member of the genus psychrobacter isolated from antarctic soil.
<3>J. Bacteriol.
<4>194
<5>2403
<6>2012
<7>Psychrobacter spp. have shown characteristics indicating remarkable capabilities  at subzero
temperatures that identify them as potential model organisms for the
study of low-temperature adaptations. Here we present the draft genome sequence
of Psychrobacter sp. PAMC 21119, which was isolated from permafrost soil of
Antarctica; this information could provide insight into adaptation and evolution
strategies under extreme environmental conditions.

<>

<1>Kim, S.J., Shin, S.C., Hong, S.G., Lee, Y.M., Lee, H., Lee, J., Choi, I.G., Park, H.
<2>Genome Sequence of Janthinobacterium sp. Strain PAMC 25724, Isolated from Alpine  Glacier Cryoconite.
<3>J. Bacteriol.
<4>194
<5>2096
<6>2012
<7>The draft genome of Janthinobacterium sp. strain PAMC 25724, which is a violacein-producing
psychrotolerant bacterium, was determined. The strain was
isolated from glacier cryoconite of the Alps mountain permafrost region. The
sequence will allow identification and characterization of the genetic
determination of its cold-adaptive properties.

<>

<1>Kim, S.K., Chung, W.H., Kim, S.H., Jung, K.H., Kim, N., Chai, Y.G.
<2>Complete Genome Sequence of Bacillus anthracis HYU01, Isolated from Soil Samples  in the Korean Peninsula.
<3>Genome Announcements
<4>2
<5>e00769-14
<6>2014
<7>Bacillus anthracis is a Gram-positive endospore-forming bacterium that causes the zoonotic
disease anthrax. We report a complete genome sequence of B. anthracis
strain HYU01, isolated from Changnyung, which belongs to the B branch (B.Br.)
001/002 canonical single nucleotide polymorphism (canSNP) group.

<>

<1>Kim, S.M., Cho, S.J., Lee, S.B.
<2>Genome Sequence of the Unclassified Marine Gammaproteobacterium BDW918.
<3>J. Bacteriol.
<4>194
<5>3753-3754
<6>2012
<7>The unclassified marine gammaproteobacterium BDW918, which utilizes volatile fatty acids but
not most common carbohydrates and amino acids, was isolated from
Dokdo seawater in South Korea. Here we present a draft genome of the strain
BDW918, which encodes many putative genes related to fatty acid metabolism and
aromatic hydrocarbon degradation.

<>

<1>Kim, S.Y., Hwang, S.M., Chang, K.S.
<2>Correlation between Sau1 restriction and modification complex type and coagulase serotype or SCCmec type of Staphylococcus aureus.
<3>J. Bacteriol. Virol.
<4>40
<5>163-170
<6>2010
<7>Staphylococcus aureus coagulase serotype I to VIII isolated from clinical samples could be
classified into two groups,
methicillin-sensitive S. aurues (MSSA) and methicilln-resistant S. aurues (MRSA), by
antibiotics susceptibility and
existence of mecA which is a gene related with methicillin resistance. Coagulase serotype I,
VI, and VIII were MSSA
which showed different antimicrobial susceptibility. Coagluase serotype II-V and VII are MRSA
in which mecA and
SCCmec are detected. To analyze Sau1 restriction and modification (R-M) complex types by
coagulase type and
SCCmec type, sau1hsdR, sau1hsdM and sau1hsdS genes involved in Sau1 R-M complex were detected
by PCR, we
found five complex types such as M1, R2M2, R2M2, R2M2S1, and R2M2S2. Coagulase serotype I, VI,
and VIII of
MSSA were M1, R2M2 and R2M2, respectively. SCCmec type II and coagulase serotype II, SCCmec
type III and
coagulase serotype III, SCCmec type IV and coagulase serotype V, and SCCmec type IV and
coagulase serotype IV,
VII of MRSA were Sau1 R-M complex type R2M2S1, R2M2, R2M2, and R2M2S2, respectively. Taken
together,
correlation between Sau1 R-M complex types and coagulase or SCCmec types of S. aureus was
found.

<>

<1>Kim, U.Y., Jon, S.C., Ra, S.R.
<2>Optimal culture condition of restriction endonuclease Bci528I producing strain by QE method.
<3>Choson Minjujuui Inmin Konghwaguk Kwahagwon Tongbo
<4>97
<5>42-43
<6>2008
<7>In this paper we determine optimal culture condition of Bacillus circulans 528 on production
of a new restriction endonuclease Bci528I by QE method.

<>

<1>Kim, W.J., Kim, Y.O., Kim, D.G., Nam, B.H., Kong, H.J., Jung, H., Lee, S.J., Kim, D.W., Kim, D.S., Chae, S.H.
<2>Genome Sequence of Marine Bacterium Idiomarina sp. Strain 28-8, Isolated from Korean Ark Shells.
<3>Genome Announcements
<4>1
<5>e00772-13
<6>2013
<7>Idiomarina sp. strain 28-8 is an aerobic, Gram-negative, flagellar bacterium isolated from the
bodies of ark shells (Scapharca broughtonii) collected from
underwater sediments in Gangjin Bay, South Korea. Here, we present the draft
genome sequence of Idiomarina sp. 28-8 (2,971,606 bp, with a G+C content of
46.9%), containing 2,795 putative coding sequences.

<>

<1>Kim, W.J., Kim, Y.O., Kim, D.S., Choi, S.H., Kim, D.W., Lee, J.S., Kong, H.J., Nam, B.H., Kim, B.S., Lee, S.J., Park, H.S., Chae, S.H.
<2>Draft Genome Sequence of Kocuria rhizophila P7-4.
<3>J. Bacteriol.
<4>193
<5>4286-4287
<6>2011
<7>We report the draft genome sequence of Kocuria rhizophila P7-4, which was isolated from the
intestine of Siganus doliatus caught in the Pacific
Ocean. The 2.83-Mb genome sequence consists of 75 large contigs (>100 bp
in size) and contains 2,462 predicted protein-coding genes.

<>

<1>Kim, Y., Chandrasegaran, S.
<2>Chimeric restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>883-887
<6>1994
<7>Fok I restriction endonuclease recognizes the nonpalindromic pentadeoxyribonucleotide
5'-GGATG-3'-5'-CATCC-3' in duplex DNA and cleaves 9 and 13 nt away from the recognition
site. Recently, we reported the presence of two distinct and separable domains within this
enzyme: one for the sequence-specific recognition of DNA (the DNA-binding domain) and the
other for the endonuclease activity (the cleavage domain). Here, we report the construction of
a chimeric restriction endonuclease by linking the Drosphila Ultrabithorax homeodomain to the
cleavage domain (F/N) of Fok I restriction endonuclease. The hybrid enzyme, Ubx-F/N, was
purified, and its cleavage properties were characterized. The hybrid enzyme show the same DNA
sequence-binding preference as that of Ubx; as expected, it cleaves the DNA away from the
recognition site. On the 5'-TTAATGGTT-3' strand the hybrid enzyme cleaves 3 nt away from the
recognition site, whereas it cuts the complementary 5'-AACCATTAA-3' strand 8, 9, or 10 nt
away from the binding site. Similarly engineered hybrid enzymes could be valuable tools in
physical mapping and sequencing of large eukaryotic genomes.

<>

<1>Kim, Y., Choi, J., Grable, J.C., Greene, P., Hager, P., Rosenberg, J.M.
<2>Studies on the canonical DNA-EcoRI endonuclease complex and the EcoRI kink.
<3>Structural Biology: The State of the Art., Adenine Press, Sarma, R.H., Sarma, M.H., New York
<4>0
<5>225-246
<6>1994
<7>The crystal structure of the complex between EcoRI endonuclease and the cognate
oligonucleotide TCGCGAATTCGCG was determined to 2.7 A resolution by multiple isomorphous
derivatives and refined to an R-factor of 0.21.  The complex includes two protein subunits
related by a two-fold axis of rotational symmetry; they have alpha/beta architecture with the
cleavage site at a "switch point" of the beta sheet.  Sequence specificity is mediated by
eighteen hydrogen bonds and numerous Van der Walls contacts between the protein and the DNA
bases.  The DNA is distorted in the complex; the "EcoRI kink" is characterized by unusual
roll, increased rise, underwinding, a "kink" or "jog" in the backbone at the ApA step and
widening of both the major and minor grooves.

<>

<1>Kim, Y., Grable, J.C., Love, R., Greene, P.J., Rosenberg, J.M.
<2>Refinement of EcoRI endonuclease crystal structure: A revised protein chain tracing.
<3>Science
<4>249
<5>1307-1309
<6>1990
<7>None

<>

<1>Kim, Y.-G., Cha, J., Chandrasegaran, S.
<2>Hybrid restriction enzymes: Zinc finger fusions to FokI cleavage domain.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>1156-1160
<6>1996
<7>A long-term goal in the field of restriction-modification enzymes has been to generate
restriction endonucleases with novel sequence specificities by mutating or engineering
existing enzymes.  This will avoid the increasingly arduous task of extensive screening of
bacteria and other microorganisms for new enzymes.  Here, we report the deliberate creation of
novel site-specific endonucleases by linking two different zinc finger proteins to the
cleavage domain of FokI endonuclease.  Both fusion proteins are active and under optimal
conditions cleave DNA in a sequence-specific manner.  Thus, the modular structure of FokI
endonuclease and the zinc finger motifs makes it possible to create "artificial" nucleases
that will cut DNA near a predetermined site.  This opens the way to generate many new enzymes
with tailor-made sequence specifities desirable for various applications.

<>

<1>Kim, Y.-G., Kim, P.S., Herbert, A., Rich, A.
<2>Construction of a Z-DNA-specific restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>12875-12879
<6>1997
<7>Novel restriction enzymes can be created by fusing the nuclease domain of FokI endonuclease
with defined DNA binding domains.  Recently, we have characterized a domain (Za) from the
N-terminal region of human double-stranded RNA adenosine deaminase, which binds the
Z-conformation with high specificity.  Here we report creation of a conformation-specific
endonuclease, Za nuclease, which is a chimera of Za and FokI nuclease.  Purified Za nuclease
cleaves negatively supercoiled plasmids only when they contain a Z-DNA forming insert, such as
(dC-dG)13.  The precise location of the cleavage sites was determined by primer extension.
Cutting has been mapped to the edge of the B-Z junction, suggesting that Za nuclease binds
within the Z-DNA insert, but cleaves in the nearby B-DNA, by using a mechanism similar to type
IIs restriction enzymes.  These data show that Za binds Z-DNA in an environment similar to
that in a cell.  Za nuclease, a structure-specific restriction enzyme, may be a useful tool
for further study of the biological role of Z-DNA.

<>

<1>Kim, Y.-G., Li, L., Chandrasegaran, S.
<2>Insertion and deletion mutants of FokI restriction endonuclease.
<3>J. Biol. Chem.
<4>269
<5>31978-31982
<6>1994
<7>FokI restriction endonuclease recognizes the nonpalindromic pentadeoxyribonucleotide,
5'-GGATG-3':5'-CATCC-3' in duplex DNA and cleaves 9 and 13 nucleotides away from the
recognition site. We have reported the presence of two distinct and separable protein domains
within this enzyme: one for the sequence-specific recognition of DNA (the DNA binding domain)
and the other for the endonuclease's activity (the cleavage domain). Our studies have
suggested that the two domains are connected by a linker region, which appears to be amenable
for repositioning of the DNA-sequence recognition domain with respect to the catalytic domain.
Here, we report the construction of several insertion (4-, 8-, 12-, 18-, 19-, or 23-amino acid
residues) and deletion (4- or 7-amino acid residues) mutants of the linker region of FokI
endonuclease. The mutant enzymes were purified, and their cleavage properties were
characterized. The mutants have the same DNA sequence specificity as the wild-type enzyme.
However, compared with the wild-type enzyme, the insertion mutants cleave predominantly one
nucleotide further away from the recognition site on both strands of the DNA substrate. The
four-codon deletion mutant shows relaxed specificity at the cut site while the seven-codon
deletion appears to inactivate the enzyme. The DNA binding and cleavage domains of FokI appear
to be linked by a relatively malleable linker. No simple linear relationship exists between
the linker length and the distance of the cut site from the recognition site. Furthermore, the
four-codon insertion mutants cleave DNA substrates containing hemi-methylated FokI sites; they
do not cleave fully methylated substrates. These results are best explained as a consequence
of protein-protein interactions between the domains.

<>

<1>Kim, Y.-G., Shi, Y., Berg, M., Chandrasegaran, S.
<2>Site-specific cleavage of DNA-RNA hybrids by zinc finger/FokI cleavage domain fusions.
<3>Gene
<4>203
<5>43-49
<6>1997
<7>Zinc-finger proteins of the Cys2His2 type bind DNA-RNA hybrids with affinities comparable to
those for DNA duplexes.  Such zinc-finger proteins were converted into site-specific cleaving
enzymes by fusing them to the FokI cleavage domain.  The  proteins are active and under
optimal conditions cleave DNA duplexes in a sequence-specific manner.  These fusions also
exhibit site-specific cleavage of the DNA strand within DNA-RNA hybrids albeit at a lower
efficiency (~/-50-fold) compared to the cleavage of the DNA duplexes.  These engineered
endonucleases represent the first of their kind in terms of their DNA-RNA cleavage properties,
and they may have important biological applications.

<>

<1>Kim, Y.-G., Smith, J., Durgesha, M., Chandrasegaran, S.
<2>Chimeric restriction enzyme: Gal4 fusion to FokI cleavage domain.
<3>Biol. Chem.
<4>379
<5>489-495
<6>1998
<7>Gal4, a yeast protein, activates transcription of genes required for metabolism of galactose
and melibiose.  It binds as a dimer to a consensus palindromic 17-base pair DNA sequence.  It
is a member of the third family of proteins that contain zinc-mediated peptide loops that
interact specifically with nucleic acids.  Gal4 has a very distinctive zinc coordination
profile and mode of DNA-binding.  Here, we report the creation of a novel site-specific
endonuclease by linking the N-terminal 147 amino acids of Gal4 to the cleavage domain of FokI
endonuclease.  The fusion protein is active and under optimal conditions, binds to a 17 bp
consensus DNA site and cleaves near this site.  As expected, the cleavage occurs on either
side of the consensus binding site(s).

<>

<1>Kim, Y.O., Kim, W.J., Choi, S.H., Kim, D.S., Kim, D.W., Lee, J.S., Kong, H.J., Nam, B.H., Kim, B.S., Lee, S.J., Park, H.S., Chae, S.H.
<2>Genome Sequence of Acinetobacter sp. Strain P8-3-8, Isolated from Fistularia commersonii in Vietnam.
<3>J. Bacteriol.
<4>193
<5>4288-4289
<6>2011
<7>Acinetobacter sp. strain P8-3-8 is an aerobic, Gram-negative marine bacterium isolated from
the intestine of the bluespotted cornetfish
(Fistularia commersonii). Here, we present the draft genome sequence of
Acinetobacter sp. P8-3-8 (3,905,565 bp, with a G+C content of 37.6%)
containing 3,621 putative coding sequences. The genome data reveal a high
density of genes encoding transcriptional regulators involved in anaerobic
respiration.

<>

<1>Kim, Y.S., Rho, H.M.
<2>Characterization of the restriction endonuclease BdiI from Brevibacterium divaricatum.
<3>Korean J. Microbiol.
<4>24
<5>18-23
<6>1986
<7>A new type II restriction endonuclease, BdiI, has been isolated from Brevibacterium
divaricatum FERM 5948 by procedures of ammonium sulfate fractionation, DEAE-cellulose
chromatography and heparin agarose chromatography. The purified BdiI restriction endonuclease
had the same cleavage patterns as ClaI whose recognition sequence is 5'ATCGAT3'. From the
result that lambda-ClaI DNA fragment could be cloned in pBR322 digested with BdiI, it has been
proven that BdiI cuts between T and C (R'AT/CGAT3') within the recognition sequence and
produces 5'pCG cohesive end. The optimal temperature for the BdiI restriction endonuclease
activity was 37C, and optimal salt (NaCl) concentration was 50-100 mM.

<>

<1>Kim, Y.S., Rho, H.M.
<2>Purification and Characterization of PstI Methylase from Providencia stuartii 164.
<3>Korean Biochem. J.
<4>17
<5>107-119
<6>1984
<7>In this report, we described the purification and characterization of PstI
methylase from Providencia stuartii 164.  PstI methylase was a site-specific
methylase and has been found to have methylation activity on single-stranded
PhiX174 DNA.  This purified PstI methylase will be valuable for the study of
the structure and expression of PstI restriction-modification gene.

<>

<1>Kimball, M., Linn, S.
<2>The release of oligonucleotides by the Escherichia coli B restriction endonuclease.
<3>Biochem. Biophys. Res. Commun.
<4>68
<5>585-591
<6>1976
<7>The Escherichia coli B (Eco B) restriction endonuclease releases approximately
75 nucleotides as acid-soluable oligonucleotides for each single-strand
endonucleolytic scission that it catalyzes.  This reaction, like the
endonucleolytic cleavage, requires ATP, Mg++, S-adenosylmethionine, and
unmodified DNA containing appropriate specificity sites.  Like the endonuclease
reaction, the release of oligonucleotides terminates after roughly 5 minutes.
The acid-soluable oligonucleotides have an average chain length of roughly 7,
and an apparently random base composition.

<>

<1>Kimbrel, J.A., Chang, J., Arp, D.J., Sayavedra-Soto, L.A.
<2>The Draft Genome Sequence of Nocardioides sp. Strain CF8 Reveals the Scope of Its Metabolic Capabilities.
<3>Genome Announcements
<4>1
<5>e00439-13
<6>2013
<7>Nocardioides sp. strain CF8 was isolated from a soil sample collected at the Hanford
Department of Energy site, Richland, WA. The strain was identified in
microcosms based on its ability to grow on butane and has been characterized for
its potential applications in the biodegradation of halogenated hydrocarbons.
Here, the draft genome sequence is reported.

<>

<1>Kimelman, A., Levy, A., Sberro, H., Kidron, S., Leavitt, A., Amitai, G., Yoder-Himes, D., Wurtzel, O., Zhu, Y., Rubin, E., Sorek, R.
<2>A vast collection of microbial genes that are toxic to bacteria.
<3>Genome Res.
<4>22
<5>802-809
<6>2012
<7>In the process of clone-based genome sequencing, initial assemblies frequently
contain cloning gaps that can be resolved using cloning-independent methods, but
the reason for their occurrence is largely unknown. By analyzing 9,328,693
sequencing clones from 393 microbial genomes, we systematically mapped more than
15,000 genes residing in cloning gaps and experimentally showed that their
expression products are toxic to the Escherichia coli host. A subset of these
toxic sequences was further evaluated through a series of functional assays
exploring the mechanisms of their toxicity. Among these genes, our assays
revealed novel toxins and restriction enzymes, and new classes of small,
non-coding toxic RNAs that reproducibly inhibit E. coli growth. Further analyses
also revealed abundant, short, toxic DNA fragments that were predicted to
suppress E. coli growth by interacting with the replication initiator DnaA. Our
results show that cloning gaps, once considered the result of technical problems,
actually serve as a rich source for the discovery of biotechnologically valuable
functions, and suggest new modes of antimicrobial interventions.

<>

<1>Kimmerly, W.J.
<2>Staphylococcus epidermidis nucleic acids and proteins.
<3>US Patent Office
<4>US 6703492 B
<5>
<6>2004
<7>S. epidermidis polypeptides and DNA (RNA) encoding such polypeptides and a procedure for
producing such polypeptides by recombinant techniques is disclosed.  Also disclosed are
methods for utilizing such polypeptides and DNA (RNA) for the treatment of infection,
particularly infections arising from S. epidermidis.  Antagonists against the function of such
polypeptides and their use as therapeutics to treat infection are also disclosed.  Also
disclosed are diagnostic assays for detecting diseases related to the presence of S.
epidermidis nucleic acid sequences and the polypeptides in a host.  Also disclosed are
diagnostic assays for detecting polynucleotides and polypeptides related to S. epidermidis.

<>

<1>Kimura, B., Takahashi, H., Hokimoto, S., Tanaka, Y., Fujii, T.
<2>Induction of the histidine decarboxylase genes of Photobacterium damselae subsp. damselae (formally P. histaminum) at low pH.
<3>J. Appl. Microbiol.
<4>107
<5>485-497
<6>2009
<7>AIMS: To elucidate the detailed mechanism of histamine production by Photobacterium damselae
subsp. damselae. METHODS AND RESULTS: Histidine decarboxylase and related genes of P. damselae
subsp. damselae were cloned, and three open reading frames named as hdcT, hdcA and hisRS were
identified. The hdcA gene encodes a polypeptide of 377 amino acids and is considered to be the
pyridoxal-P dependent histidine decarboxylase. The hdcT gene is assumed to be a
histidine/histamine antiporter, and the hisRS gene is considered to be a histidyl-tRNA
synthetase. Recombinant Escherichia coli strains harbouring plasmids carrying the P. damselae
hdc genes were shown to over-excrete histamine extracellularly. Northern blot analysis and
quantitative RT-PCR revealed high levels of mono- and bi-cistronic transcripts of hdcA, hdcT
and hisRS genes under conditions of low pH and histidine excess. CONCLUSIONS: The hdcA gene of
P. damselae was constructed as an operon with putative histidine/histamine antiporter and
histidyl-tRNA synthetase. Mono- and poly-cistronic transcripts and acid induction were
detected. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report of cloning the
histidine decarboxylase gene cluster in gram-negative bacteria. Also, these genes were induced
under acidic conditions and in the presence of excess histidine.

<>

<1>Kimura, H., Ishihara, G., Tajima, S.
<2>Isolation and expression of a Xenopus laevis DNA methyltransferase cDNA.
<3>J. Biochem. (Tokyo)
<4>120
<5>1182-1189
<6>1996
<7>A Xenopus DNA methyltransferase cDNA was isolated from a Xenopus oocyte cDNA library by
screening with the mouse DNA methyltransferase cDNA as a probe.  The elucidated nucleotide
sequence gave a 4,470 nucleotide open reading frame, and the predicted protein was composed of
1,490 amino acid residues, showing high homology to animal DNA methyltransferases, especially
in the catalytic domain in the carboxyl-terminal region.  The cysteine-rich region and the
Lys-Gly repeat which were first found in the mouse sequence were conserved in Xenopus.
However, 200 amino acid residues at the amino-terminus of Xenopus DNA methyltransferase were
quite different from those of mouse and human, but showed 70% homology with those of chicken.
The cloned Xenopus DNA methyltransferase cDNA expressed in COS1 cells showed a significant DNA
methyltransferase activity.  The size of the translation product of Xenopus DNA
methyltransferase cDNA expressed in COS1 cells was identical with that of the endogenous DNA
methyltransferase in Xenopus A6 cells and also with the size of newly synthesized DNA
methyltransferase in Xenopus oocytes.  However, a slightly larger immunoreactive band of about
205 kDa, and a small immunoreactive band of about 100 kDa, which were poorly labeled by short
incubation with radiolabeled amino acids, were the main bands in stage I-III and stage IV-VI
oocytes, respectively.

<>

<1>Kimura, H., Nakamura, T., Ogawa, T., Tanaka, S., Shiota, K.
<2>Transcription of mouse DNA methyltransferase 1 (Dnmt1) is regulated by both E2F-Rb-HDAC-dependent and -independent pathways.
<3>Nucleic Acids Res.
<4>31
<5>3101-3113
<6>2003
<7>Abnormal expression of Dnmt1 in vivo induces cellular alterations such as transformation, and
an increase in Dnmt1 mRNA plays a causal role in
c-fos-, ras- and SV40 large T antigen-induced transformation of
fibroblasts in vitro. Here, we have investigated the regulation of Dnmt1
transcription. We identified the promoter region and major transcription
start sites of mouse Dnmt1 and found two important cis-elements within the
core promoter region. One is an E2F binding site, and the other is a
binding site for an as yet unidentified factor. Point mutations in the two
cis-elements decreased promoter activity in both non-transformed and
transformed cells. Thus, both sites play a critical role in regulation of
Dnmt1 transcription in proliferating cells. Treatment with trichostatin A,
a specific inhibitor of histone deacetylase, increased Dnmt1 promoter
activity in G0/G1-arrested NIH 3T3 cells. Furthermore, the decrease in
promoter activity induced by expression of E2F-1 and Rb was reversed by
trichostatin A treatment of Saos-2 cells. Taken together, these data
indicate that transcription of Dnmt1 is regulated in a complex fashion by
E2F and other transcription factors through E2F-Rb-HDAC-dependent and
-independent pathways. These findings suggest that Dnmt1 is a target gene
of these pathways in cell proliferation, cell transformation and
tumorigenesis.

<>

<1>Kimura, H., Shiota, K.
<2>Methyl-CpG-binding Protein, MeCP2, Is a Target Molecule for Maintenance DNA Methyltransferase, Dnmt1.
<3>J. Biol. Chem.
<4>278
<5>4806-4812
<6>2003
<7>During mammalian cell division, DNA methylation patterns are transferred accurately to the
newly synthesized DNA strand. This depends on
maintenance DNA methyltransferase activity. DNA methylation can affect
chromatin organization and gene expression by recruitment of histone
deacetylases (HDACs). Here we show that the methyl-CpG binding protein,
MeCP2, interacts directly with the maintenance DNA methyltransferase,
Dnmt1. The region of MeCP2 that interacts with Dnmt1 corresponds to the
transcription repressor domain which can also recruit HDACs via a
corepressor, mSin3A. Dnmt1 can form complexes with HDACs as well as MeCP2.
Surprisingly, the MeCP2-Dnmt1 complex does not contain the histone
deacetylase, HDAC1. Thus, Dnmt1 takes the place of the mSin3A-HDAC1
complex, indicating that the MeCP2-interacting Dnmt1 does not bind to
HDAC1. Further, we demonstrate that MeCP2 can form a complex with
hemimethylated as well as fully methylated DNA. Immunoprecipitated MeCP2
complexes show DNA methyltransferase activity to hemimethylated DNA. These
results suggest that Dnmt1 associates with MeCP2 in order to perform
maintenance methylation in vivo. We propose that genome-wide and/or
-specific local DNA methylation may be maintained by the Dnmt1-MeCP2
complexes, bound to hemimethylated DNA. Dnmt1 may be recruited to targeted
regions via multiple steps that may or may not involve histone
deacetylases.

<>

<1>Kimura, H., Suetake, I., Tajima, S.
<2>Xenopus maintenance-type DNA methyltransferase is accumulated and translocated into germinal vesicles of oocytes.
<3>J. Biochem. (Tokyo)
<4>125
<5>1175-1182
<6>1999
<7>In vertebrates, DNA methylation plays an important role in the regulation of gene expression
and embryogenesis. DNA methyltransferase, which catalyzes the introduction of a methyl group
at the 5th position of cytosine in the CpG sequence, is highly accumulated in mouse oocytes
and is excluded from nuclei [Carlson et al. (1992) Genes Dev. 6, 2536-2541]. In this study, we
examined the expression level and localization of Xenopus DNA methyltransferase in oocytes
during oogenesis. The DNA methyltransferase protein was detectable in stage III oocytes and
increased thereafter, until the oocytes had matured. The rate of DNA methyltransferase
synthesis rapidly increased after stage IV oocytes. Different from in mouse oocytes, DNA
methyltransferase was equally distributed in the nuclear and post-nuclear fractions, in stage
VI oocytes. DNA methyltransferase translocated into nuclei was uniformly localized in the
nuclear matrix, and the accumulated DNA methyltransferase in stage VI nuclei had DNA
methylation activity.

<>

<1>Kimura, H., Takeda, T., Tanaka, S., Ogawa, T., Shiota, K.
<2>Expression of rat DNA (cytosine-5) methyltransferase (DNA MTase) in rodent trophoblast giant cells: Molecular cloning and characterization of rat DNA MTase.
<3>Biochem. Biophys. Res. Commun.
<4>253
<5>495-501
<6>1998
<7>Methylation of genomic DNA is involved in the basic mechanism of gene inactivation, chromatin
organization, X chromosome inactivation and genomic imprinting.  A pattern of DNA methylation
is maintained in mitotic cells by DNA (cytosine-5) methyltransferase (DNA MTase).  The DNA
MTase has been shown to be also expressed in postmitotic cells such as neurons.  In the
present report, as an approach to analyzing mechanisms underlying regulation of DNA MTase
expression, we first isolated rat DNA MTase cDNA.  The isolated cDNA encoded a protein of
1,622 amino acid residues showing 88.3% and 64.2% homology with mouse and human DNA MTase,
respectively.  Northern blot analysis showed that DNA MTase mRNA was highly expressed in
placenta during mid- to late-pregnancy.  We then analyzed the expression of DNA MTase in
Rcho-1 cells, a rat choriocarcinoma-derived cell line, which cease cell division but keep
replicating genomic DNA when differentiated in vitro.  We found that the expression of DNA
MTase protein was decreased in terminally differentiated Rcho-1 cells whereas DNA MTase mRNA
was consistently expressed.  This result suggested posttranscriptional regulation of DNA MTase
activity in Rcho-1 cells.  The Rcho-1 cells would be a valuable model for studying the
regulation of gene expression and function of DNA MTase in postmitotic, differentiated cells.

<>

<1>Kinch, L.N., Ginalski, K., Rychlewski, L., Grishin, N.V.
<2>Identification of novel restriction endonuclease-like fold families among hypothetical proteins.
<3>Nucleic Acids Res.
<4>33
<5>3598-3605
<6>2005
<7>Restriction endonucleases and other nucleic acid cleaving enzymes form a large and extremely
diverse superfamily that display little sequence similarity despite retaining a common core
fold responsible for cleavage. The lack of significant sequence similarity between protein
families makes homology inference a challenging task and hinders new family identification
with traditional sequence-based approaches. Using the consensus fold recognition method
Meta-BASIC that combines sequence profiles with predicted protein secondary structure, we
identify nine new restriction endonuclease-like fold families among previously uncharacterized
proteins and predict these proteins to cleave nucleic acid substrates. Application of
transitive searches combined with gene neighborhood analysis allow us to confidently link
these unknown families to a number of known restriction endonuclease-like structures and thus
assign folds to the uncharacterized proteins. Finally, our method identifies a novel
restriction endonuclease-like domain in the C-terminus of RecC that is not detected with
structure-based searches of the existing PDB database.

<>

<1>Kinder, S.A., Badger, J.L., Bryant, G.O., Pepe, J.C., Miller, V.L.
<2>Cloning of the YenI restriction endonuclease and methyltransferase from Yersinia enterocolitica serotype O8 and construction of a transformable R-M+ mutant.
<3>Gene
<4>136
<5>271-275
<6>1993
<7>Two different clonal groups of pathogenic Yersinia enterocolitica strains, American and
non-American, have been recognized. These are distinguished by a number of criteria, including
their virulence in a murine model of infection. However, genetic analysis of virulence in
American strains has been hampered due to the severe restriction of transformed of
electroporated DNA. Thus, we cloned the yenIMR locus from American serotype strain 8081c,
which encodes YenI, an isoschizomer pf PstI. This clone encodes both the restriction
endonuclease and methyltransferase. The location of the genes on the clone was determined and
this information was used to construct a small deletion (400 bp) that results in an R-M+
phenotype. This mutation was recombined onto the Y. enterocolitica chromosome to give an R-M+
mutant which showed at least a 1000-fold increase in electroporation frequency compared to the
wild-type strain. Southern analysis using a probe derived from yenIMR indicated that American
serotype strains have this locus whereas non-American serotype strains do not.

<>

<1>King, G., Murray, N.E.
<2>Restriction enzymes in cells, not eppendorfs.
<3>Trends Microbiol.
<4>2
<5>465-469
<6>1994
<7>Restriction enzymes are essential reagents to molecular biologists, but their relevance to
bacterial populations is less obvious. Most bacteria encode restriction and modification
systems and these are commonly considered to be a barrier to phage infection. Current evidence
also supports a more general role for them in genetic recombination.

<>

<1>King, G., Murray, N.E.
<2>Restriction alleviation and modification enhancement by the Rac prophage of Escherichia coli K-12.
<3>Mol. Microbiol.
<4>16
<5>769-777
<6>1995
<7>Bacteriophage lambda encodes an antirestriction function, Ral, which is able to modulate the
activity of the Escherichia coli K-12 restriction and modification system, EcoKI. Here we
report the characterization of an analogous function, Lar, expressed by E. coli sbcA mutants
and the hybrid phage lambda reverse. E. coli sbcA mutants and lambda reverse both express
genes of the Rac prophage, and we have located the lar gene immediately downstream of recT in
this element. The lar gene has been cloned in an expression plasmid, and a combination of
site-directed mutagenesis and labelling of plasmid-encoded proteins has enabled us to identify
a number of translational products of lar, the smallest of which is sufficient for restriction
alleviation. Lar, like Ral, is able both to alleviate restriction and to enhance modification
by EcoKI. Lar, therefore, is functionally similar to Ral and the nucleotide sequences of their
genes share 47% identity, indicating a common origin. A comparison of the predicted amino acid
sequences of Lar and Ral shows only a 25% identity, but a few short regions do align and may
indicate residues important for structure and/or function.

<>

<1>King, G., Murray, N.E.
<2>Modification enhancement and restriction alleviation by bacteriophage lambda.
<3>Gene
<4>157
<5>225
<6>1995
<7>The activity of EcoKI, and related restriction and modification (R-M) systems, is modulated by
the bacteriophage lambda ral gene product.  We have identified the coding sequence for an
analogous function in the Rac prophage of E. coli K-12.

<>

<1>King, J., Umezu, K., Ryu, J.
<2>Complementation of type ID restriction-modification systems between KpnAI and StySBLI.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>100
<5>371
<6>2000
<7>Type I restriction-modification systems are the most complex R-M systems in bacteria and
consist of three different subunits, encoded by the hsdR (restriction/cleavage of DNA), hsdM
(modification/methylation of DNA), and hsdS (recognition of DNA sequence) genes.  KpnAI, a
restriction system from Klebsiella pneumoniae strain M5a1, is a member of the Type ID family.
The predicted peptide sequences of KpnAI have a high degree of similarity with the StySBLI
system, a prototype of type ID, from Salmonella blegdam, showing 95% and 98% homology in the
HsdR and hsdM peptide sequences, respectively.  Differences in the hsdS genes (44% homology)
however, suggest that each system has its own distinct DNA recognition sequence.  A series of
complementation tests were conducted to test whether the two systems are functionally
homologous.  Classical complementation tests were conducted using chromosomal mutants for
StySBLI and plasmids containing KpnAI, and then conversely chromosomal mutants for KpnAI and
plasmids containing StySBLI.  A new complementation method was also conducted by transforming
two plasmids, containing genes from the KpnAI and StySBLI respectively, into E. coli C.  The
results were obtained using a classical R-M test with lambda and SBS bacteriophages and
determining the efficiency of plating (EOP).  The degree of restriction for the StySBLI and
KpnAI mutants was 10-5 and 10-3, respectively, and 10-3 for the two plasmid complementation.
The results show that the two different systems are functionally homologous and
complementable.

<>

<1>King, J.B., Bowen, L.M., Dupureur, C.M.
<2>Binding and conformational analysis of phosphoramidate restriction enzyme interactions.
<3>Biochemistry
<4>43
<5>8551-8559
<6>2004
<7>Phosphoramidates are modified deoxyoligonucleotides that feature nitrogen in place of the
3'-oxygen of a phosphodiester linkage. Noted
for stability against nuclease activity, these linkages are of both
mechanistic and therapeutic interest. While a number of studies
characterizing the properties of oligonucleotides composed entirely of
phosphoramidate linkages have been published, little is known about how
singly substituted phosphoramidate substitutions affect the
thermodynamics and structure of protein-oligonucleotide interactions.
We chose to investigate these interactions with PvuII endonuclease, the
DNA binding behavior of which is well-characterized. Oligonucleotide
duplexes containing a phosphoramidate substitution at the scissile
phosphates were resistant to cleavage by the enzyme, even after
extended incubations. However, the enzyme was able to cleave the native
strand in a native: phosphoramidate heteroduplex at a rate comparable
to that observed with the native substrate. Ca(II)-stimulated PvuII
binding for a phosphoramidate-substituted oligonucleotide is comparable
to that of the native duplex (K-d approximate to 200 pM). K-d values
obtained in the presence of Mg(II) are somewhat weaker (K-d approximate
to 10 nM). Under metal-free conditions, the enzyme exhibited a
remarkable approximate to50-fold greater affinity for the modified
oligonucleotide relative to the native substrate (5 vs 240 nM). While
P-31 NMR spectra indicate increased chemical shift dispersion in the
free phosphoramidate duplex, the spectrum of the enzyme-bound duplex is
similar to that of the native duplex. H-1-N-15 HSQC analysis indicates
that enzyme conformations in the presence of these oligonucleotides are
also comparable. The tight binding of the phosphoramidate duplex under
metal-free conditions and its resistance to cleavage are attributed to
local conformational adjustments propagating from the O-N substitution.

<>

<1>King, J.S., Valcarcel, E.R., Rufer, J.T., Phillps, J.W., Morgan, W.F.
<2>Noncomplementary DNA double-strand-break rejoining in bacterial and human cells.
<3>Nucleic Acids Res.
<4>21
<5>1055-1059
<6>1993
<7>We examined the rejoining of noncomplementary restriction enzyme-produced DNA double-strand
breaks in Escherichia coli and in cultured human cells. The enzymes used in this study, ClaI,
BamHI and SalI, produce double-strand breaks with 5' protruding single strands. The joining
of a ClaI-produced DNA end to a BamHI-produced end or to a SalI-produced end was examined at
the DNA sequence level. End rejoining in E.coli was studied by transforming cultures with
linear plasmid DNA that was gel purified from restriction digests, and end rejoining in
cultured human cells was studied by introducing enzymes into the cells by electroporation. The
human cells used contain an Epstein-Barr virus (EBV)-based shuttle vector, pHAZE, that was
recovered and introduced into E.coli for further analysis. The major products of DNA
end-joining processes observed in linear plasmid-transformed E.coli and in the human cells
exposed to restriction enzymes were identical. Furthermore, the deletions observed in both
systems and in the spontaneous mutant plasmid in untreated human cells had a common underlying
feature: short stretches of directly repeated DNA at the junction sites.

<>

<1>King, K., Benkovic, S.J., Modrich, P.
<2>Glu-111 is required for activation of the DNA cleavage center of EcoRI endonuclease.
<3>J. Biol. Chem.
<4>264
<5>11807-11815
<6>1989
<7>Gap repair in the presence of 2'-deoxycytosine 5'-O-(1-thiotriphosphate) has
been utilized to mutagenize the amino-terminal one-half of the structural gene
for EcoRI endonuclease.  This approach has led to identification of over 200
mutants defective in endonuclease function.  One mutant protein, which binds to
the EcoRI sequence but displays greatly reduced cleavage activity, is the
consequence of a Glu to Gly change at position 111.  This protein has been
purified to homogeneity and characterized in detail.  Subunit interactions
governing the tetramer to dimer transition of the mutant endonuclease are near
normal as are parameters governing its interaction with specific and
nonspecific DNA sequences.  However, the rate constants for first and second
strand cleavage steps are reduced by 60,000- and 30,000-fold, respectively, as
a consequence of the Glu->Gly change.  The defect in chemical cleavage steps
can be partially overcome by elevating the pH of the reaction buffer from 7.6
to 8.5, conditions which enhance the rate of EcoRI strand cleavage by wild type
enzyme to a similar degree.  We suggest that the Glu-111 mutation affects an
interface between recognition and cleavage functions of the enzyme, an idea
consistent with the suggestion that the cleavage center of the endonuclease is
subject to activation upon specific recognition of the EcoRI sequence.

<>

<1>King, K., Wright, D., Modrich, P.
<2>Mutationally altered forms of EcoRI endonuclease selectively defective in DNA cleavage.
<3>Fed. Proc.
<4>45
<5>1913
<6>1986
<7>Gap repair in the presence of dNTP[alpha-S] has been utilized to mutagenize
regions of the structural gene for EcoRI endonuclease.  This approach has led
to identification of several mutant enzymes which retain specific affinity for
the EcoRI sequence, but display greatly reduced cleavage activity.  One such
protein, involving a Glu to Gly change at position 111 (BG111), has been
purified to homogeneity and characterized in detail.  Thermodynamic and kinetic
parameters governing interaction of the mutant protein with the EcoRI sequence
are comparable to values for the wild type enzyme.  subunit interactions
governing the tetramer to dimer transition are also normal.  However, the first
order rate constants for first and second strand cleavage steps are reduced by
a factor of at least 10,000-fold relative to wild type endonuclease.  We
suggest that this mutation affects the interface between recognition and
cleavage domains of the enzyme.

<>

<1>King, K., Wright, D.J., Modrich, P.
<2>Glutamate 111 of EcoRI endonuclease is required for DNA cleavage activation.
<3>Miami BioTechnology Winter Symposium, IRL Press, Brew, K., VA
<4>8
<5>79
<6>1988
<7>The EcoRI restriction endonuclease is a well-characterized model system for
studying the interaction of enzymes with specific DNA sequences.  X-ray
crystallography has revealed major features of the interaction of this enzyme
with DNA, but has not answered important questions about the mechanism of DNA
cleavage.  We have used the cloned EcoRI endonuclease gene as a target for
site-directed mutagenesis to probe the functional role of various amino acid
residues in the enzyme.  Our studies demonstrate that glutamate 111 plays a
critical role in activation of the cleavage function of the enzyme.

<>

<1>King, K.W., Dybvig, K.
<2>Transformation of Mycoplasma capricolum and examination of DNA restriction modification in M. capricolum and Mycroplasma mycoides subsp. mycoides.
<3>Plasmid
<4>31
<5>308-311
<6>1994
<7>Plasmids pIKD and pIKD-erm have recently been developed as mycoplasmal cloning vectors. In
this report, we demonstrate that these plasmids can replicate in Mycoplasma capricolum, a
mycoplasmal species for which transformation had not previously been characterized. Both
plasmids are stably maintained at a higher copy number than in their parental species,
Mycoplasma mycoides subsp. mycoides. We have also examined the possibility of one or more
restriction-modification systems affecting transformation frequencies in both species.

<>

<1>Kingston, I.J., Gormley, N.A., Halford, S.E.
<2>DNA supercoiling enables the Type IIS restriction enzyme BspMI to recognize the relative orientation of two DNA sequences.
<3>Nucleic Acids Res.
<4>31
<5>5221-5228
<6>2003
<7>Many proteins can sense the relative orientations of two sequences at distant locations in
DNA: some require sites in inverted (head-to-head)
orientation, others in repeat (head-to-tail) orientation. Like many
restriction enzymes, the BspMI endonuclease binds two copies of its target
site before cleaving DNA. Its target is an asymmetric sequence so two
sites in repeat orientation differ from sites in inverted orientation.
When tested against supercoiled plasmids with two sites 700 bp apart in
either repeated or inverted orientations, BspMI had a higher affinity for
the plasmid with repeated sites than the plasmid with inverted sites. In
contrast, on linear DNA or on supercoiled DNA with sites 1605 bp apart,
BspMI interacted equally with repeated or inverted sites. The ability of
BspMI to detect the relative orientation of two DNA sequences thus depends
on both the topology and the length of the intervening DNA. Supercoiling
may restrain the juxtaposition of sites 700 bp apart to a particular
alignment across the superhelical axis, but the juxtaposition of sites in
linear DNA or far apart in supercoiled DNA may occur without restraint.
BspMI can therefore act as a sensor of the conformational dynamics of
supercoiled DNA.

<>

<1>Kinjo, M., Nishimura, G., Koyama, T., Mets, U., Rigler, R.
<2>Single-molecule analysis of restriction DNA fragments using fluorescence correlation spectroscopy.
<3>Anal. Biochem.
<4>260
<5>166-172
<6>1998
<7>The cleavage of fluorescence-labeled M13DNA (7250 bp) using HaeIII, HgaI, BsmAI, and BspMI was
analyzed by fluorescence correlation spectroscopy in a small volume (1.5 x 10^-15 liters).
The digestion process can be monitored by the decrease in amplitude of the fluorescence
correlation function while the original DNA molecule is divided into several fragments by the
enzymes.  To analyze this reaction by FCS, we derived a practical equation for estimating the
number of molecules in the FCS measurements.  Under standard enzymatic conditions, HaeIII and
BsmAI digested fluorescence-labeled DNA to completion in the range of 8 h, whereas HgaI and
BspMI digested the DNA after 40 h.  The comparison of recognition sequences suggested that
some tagged nucleotides could be inserted between the recognition site and the cleavage site
of the slow enzyme group.  The decrease in amplitude in the fluorescence correlation function
quantitatively monitors the hydrolysis of DNA during the digestion process.

<>

<1>Kinney, S.R.M., Pradhan, S.
<2>Regulation of Expression and Activity of DNA (Cytosine-5) Methyltransferases in Mammalian Cells.
<3>Prog. Mol. Biol. Transl. Sci.
<4>101
<5>311-333
<6>2011
<7>Three active DNA methyltransferases have been identified in mammalian cells, Dnmt1, Dnmt3a,
and Dnmt3b.  DNMT1 is primarily a maintenance methyltransferase, as it prefers to methylate
hemi-methylated DNA during DNA replication and in vitro.  DNMT3A and DNMT3B are de novo
methyltransferases and show similar activity on unmethylated and hemimethylated DNA. DNMT3L,
which lacks the catalytic domain, binds to DNMT3A and DNMT3B variants and facilitates their
chromatin targeting, presumably for de novo methylation.  There are several mechanisms by
which mammalian cells regulate DNMT levels, including varied transcriptional activation of the
respective genes and posttranslational modifications of the enzymes that can affect catalytic
activity, targeting, and enzyme degradation.  In addition, binding of miRNAs or RNA-binding
proteins can also alter the expression of DNMTs.  These regulatory processes can be disrupted
in disease or by environmental factors, resulting in altered DNMT expression and aberrant DNA
methylation patterns.

<>

<1>Kinnunen, P.K.J., Mustonen, P.K., Kere, J.K.
<2>Materials and methods for digesting DNA or RNA using restriction endonucleases.
<3>International Patent Office
<4>WO 9640899
<5>
<6>1996
<7>Disclosed are improvements for enzyme-catalyzed reactions involving DNA or RNA, including
restriction digests, which are based on conducting such reactions in the presence of lipids.

<>

<1>Kinnunen, P.K.J., Mustonen, P.K., Kere, J.K.
<2>Materials and methods for digestion of DNA or RNA using restriction endonucleases.
<3>US Patent Office
<4>US 5879950
<5>
<6>1999
<7>Disclosed are improvements for enzyme-catalyzed reactions involving DNA or RNA, including
restriction digests, which are based on conducting such reactions in the presence of lipids.

<>

<1>Kinsella, J.M., Shalaev, M.V., Ivanisevic, A.
<2>Ligation of Nanoparticle Coated DNA Cleaved with Restriction Enzymes.
<3>Chem. Mater.
<4>19
<5>3586-3588
<6>2007
<7>Material synthesis at the nanoscale frequently relies on biological molecules for inspiration,
selectivity, and specificity.  Molecular templates using biomolecules have been used to
fabricate structures with specific sizes and shapes.  Often times, the biological function of
the template molecule is utilized to connect nanoscale structures, site-specifically catalyze
reactions, or enzymatically modify templated segments.  The specificity of these biomolecular
interactions can often lead to straightforward fabrication methods of materials that are
difficult to synthesize by conventional means.  One of the simplest and most easily adapted
forms of such interactions is that involving the hybridization of single-stranded DNA to its
double-helix form.  This interaction has been used to drive the formation of complex
structures, nanoparticle conjugates, and DNA-based electronics.  More recently, the
introduction of DNA-manipulating enzymes has been investigated.  We have previously
demonstrated the ability of restriction endonucleases, which cut DNA at specific sequences, to
fragment phage DNA templated with magnetic nanoparticles.  DNA ligases, which oppose the
function of restriction enzymes by repairing cleaved DNA molecules, have recently been used to
generate specific connectivity between DNA-modified nanoparticles.

<>

<1>Kirby, R., Sangal, V., Tucker, N.P., Zakrzewska-Czerwinska, J., Wierzbicka, K., Herron, P.R., Chu, C.J., Chandra, G., Fahal, A.H., Goodfellow, M., Hoskisson, P.A.
<2>Draft Genome Sequence of the Human Pathogen Streptomyces somaliensis, a Significant Cause of Actinomycetoma.
<3>J. Bacteriol.
<4>194
<5>3544-3545
<6>2012
<7>We report the draft genome sequence of the human pathogen Streptomyces somaliensis (DSM
40738), a pathogen within a genus of largely saprophytic
organisms. S. somaliensis causes severe and debilitating deep tissue and bone
infections. The genome sequence is deposited in DDBJ/EMBL/GenBank with the
accession number AJJM01000000.

<>

<1>Kirienko, N.V., Lepikhov, K.A., Zheleznaya, L.A., Matvienko, N.I.
<2>Significance of codon usage and irregularities of rare codon distribution in genes for expression of BspLU11III methyltransferases.
<3>Biokhimiia
<4>69
<5>647-657
<6>2004
<7>Genes of adenine-specific DNA-methyltransferase M.BspLU11IIIa and cytosine-specific
DNA-methyltransferase M.BspLU11IIIb of the type IIG BspLU11III restriction-modification system
from the thermophilic strain Bacillus sp. LU11 were expressed in E. coli. They contain a large
number of codons that are rare in E. coli and are characterized by equal values of codon
adaptation index (CAI) and expression level measure (E(g)). Rare codons are either diffused
(M.BspLU11IIIa) or located in clusters (M.BspLU11IIIb). The expression level of the
cytosine-specific DNA-methyltransferase was increased by a factor of 7.3 and that of
adenine-specific DNA only by a factor of 1.25 after introduction of the plasmid pRARE
supplying tRNA genes for six rare codons in E. coli. It can be assumed that the plasmid
supplying minor tRNAs can strongly increase the expression level of only genes with cluster
distribution of rare codons. Using heparin-Sepharose and phosphocellulose chromatography and
gel filtration on Sephadex G-75 both DNA-methyltransferases were isolated as
electrophoretically homogeneous proteins (according to the results of SDS-PAGE).

<>

<1>Kirino, H., Kuwahara, R., Hamasaki, N., Oshima, T.
<2>Effect of unusual polyamines on cleavage of DNA by restriction enzymes.
<3>J. Biochem. (Tokyo)
<4>107
<5>661-665
<6>1990
<7>The effect of unusual polyamines, such as thermine, caldopentamine,
caldohexamine, tris(3-aminopropyl)amine, or tetrakis(3-aminopropyl)ammonium, on
the activities of various restriction endonucleases was investigated by using
an Escherichia coli plasmid as a substrate, which contains a high GC content
fragment from an extreme thermophile.  Restriction enzymes used were SmaI,
BanII, NaeI, RsaI, and TaqI.  Most of the polyamines tested were inhibitory to
the enzyme activities.  The larger and more branched a polyamine was, the more
the activities of nucleases were inhibited.  The inhibition was positively
correlated with the polyamine concentration.  The sites protected by a
polyamine were identical to those protected by other polyamines, and also
identical to those which were less sensitive to the restriction enzyme in the
absence of polyamines.  No sequence specificity was seen among these sites.

<>

<1>Kirsanova, O.V., Baskunov, V.B., Gromova, E.S.
<2>Type IIE and IIF restriction endonucleases interacting with two recognition sites on DNA.
<3>Mol. Biol. (Mosk)
<4>38
<5>886-900
<6>2004
<7>Recent studies have shown that restriction endonucleases (REs), which are broadly used in
genetic engineering and molecular biology, vary not
only in nucleotide sequence of the recognition site, but also in the
mechanism of their interaction with DNA. This review focuses on type
IIF and HE REs, which require simultaneous interaction with two
nucleotide sequences for efficient DNA cleavage. Crystal structures of
these REs and their complexes with DNA, stepwise interactions with DNA,
catalytic mechanisms of DNA hydrolysis, and DNA looping are considered.
Type HE REs have provided an example of a new type of DNA-protein
recognition: two copies of one recognition sequence interact
specifically with two different amino acid sequences and two different
structural motifs of one polypeptide chain.

<>

<1>Kirsanova, O.V., Cherepanova, N.A., Gromova, E.S.
<2>Inhibition of C5-cytosine-DNA-methyltransferases.
<3>Biochemistry
<4>74
<5>1175-1186
<6>2009
<7>Changes in the methylation pattern of genomic DNA, particularly hypermethylation of tumor
suppressor genes, occur at early stages of tumor development. Errors in DNA methylation
contribute to both initiation and progression of various cancers. This stimulates significant
interest in searching for inhibitors of C5-DNA-methyltransferases (MTases). Here we review the
known nucleoside mechanism-based reversible and irreversible inhibitors of the MTases, as well
as non-nucleoside ones, and discuss their inhibitory mechanisms and application for MTase
investigations and cancer therapy.

<>

<1>Kishimoto, N., Sakai, H., Jackson, J., Jacobsen, S.E., Meyerowitz, E.M., Dennis, E.S., Finnegan, E.J.
<2>Site specificity of the Arabidopsis METI DNA methyltransferase demonstrated through hypermethylation of the superman locus.
<3>Plant Mol. Biol.
<4>46
<5>171-183
<6>2001
<7>Plants with low levels of DNA methylation show a range of developmental abnormalities
including homeotic transformation of floral organs. Two independent DNA methyltransferase I
(METI) antisense transformants with low levels of DNA methylation had flowers with increased
numbers of stamens which resembled flowers seen on the loss-of-function superman (sup) mutant
plants and on transgenic plants that ectopically express APETALA3 (AP3). These METI antisense
plants have both increased and decreased methylation in and around the sup gene, compared with
untransformed controls. DNA from the antisense plants was demethylated at least 4 kb upstream
of the sup gene, while there was dense methylation around the start of transcription and
within the coding region of this gene; these regions were unmethylated in control DNA.
Methylation within the sup gene was correlated with an absence of SUP transcripts. The pattern
and density of methylation was heterogeneous among different DNA molecules from the same
plant, with some molecules being completely unmethylated. Methylcytosine occurred in
asymmetric sites and in symmetric CpA/TpG but rarely in CpG dinucleotides in the antisense
plants. In contrast, segregants lacking the METI antisense construct and epimutants with a
hypermethylated allele of sup (clark kent 3), both of which have active METI genes, showed a
higher frequency of methylation of CpG dinucleotides and of asymmetric cytosines. We conclude
that METI is the predominant CpG methyltransferase and directly or indirectly affects
asymmetric methylation.

<>

<1>Kiss, A., Baldauf, F.
<2>Molecular cloning and expression in Escherichia coli of two modification methylase genes of Bacillus subtilis.
<3>Gene
<4>21
<5>111-119
<6>1983
<7>Two modification methylase genes of Bacillus subtilis R were cloned in
Escherichia coli by using a selection procedure which is based on the
expression of these genes.  Both genes code for DNA-methyl-transferases which
render the DNA of the cloning host E. coli HB101 insensitive to the BspRI
(5'-GGCC) endonuclease of Bacillus sphaericus R.  One of the cloned genes is
part of the restriction-modification (RM) system BsuRI of B. subtilis R with
specificity for 5'-GGCC.  The other one is associated with the lysogenizing
phage SPbeta and produces the methylase M.BsuSPbetaI with specificity for
5'-GGCC.  The fragment carrying the SPbeta-derived gene also directs the
synthesis in E. coli of a third methylase activity (M.BsuSPbetaII), which
protects the host DNA against HpaII and MspI cleavage within the sequence
5'-CCGG.  Indirect evidence suggests that the two SPbeta modification
activities are encoded by the same gene.  No cross-hybridization was detected
either between the M.BsuRI and M.BsuSPbeta genes or between these and the
modification methylase gene of B. sphaericus R, which codes for the enzyme
M.BspRI with 5'-GGCC specificity.

<>

<1>Kiss, A., Finta, C., Venetianer, P.
<2>M.KpnI is an adenine-methyltransferase.
<3>Nucleic Acids Res.
<4>19
<5>3460
<6>1991
<7>The recognition sequence of the KpnI restriction-modification system is GGTACC.
There are conflicting reports in the literature concerning the nucleotide
methylated by the KpnI methyltransferase.  In one paper it was suggested that
M.KpnI produces m4-cytosine.  Other investigators obtained indirect evidence
suggesting but not proving that it methylates the adenine in the recognition
sequence.  Here we present direct evidence showing that M.KpnI is an
adenine-methyltransferase.

<>

<1>Kiss, A., Posfai, G., Keller, C.C., Venetianer, P., Roberts, R.J.
<2>Nucleotide sequence of the BsuRI restriction-modification system.
<3>Nucleic Acids Res.
<4>13
<5>6403-6420
<6>1985
<7>The genes of the 5'-GGCC specific BsuRI restriction-modification system of
Bacillus subtillis have been cloned and expressed in E. coli and their
nucleotide sequence has been determined.  The restriction and modification
genes code for polypeptides with calculated molecular weights of 66,314 and
49,642, respectively.  Both enzymes are coded by the same DNA strand.  The
restriction gene is upstream of the methylase gene and the coding regions are
separated by 780 bp.  Analysis of the RNA transcripts by S1-nuclease mapping
indicates that the restriction and modification genes are transcribed from
different promoters.  Comparison of the amino acid sequences revealed no
homology between the BsuRI restriction and modification enzymes.  There are,
however, regions of homology between the BsuRI methylase and two other GGCC
specific modification enzymes, the BspRI and SPR methylases.

<>

<1>Kiss, A., Posfai, G., Zsurka, G., Rasko, T., Venetianer, P.
<2>Role of DNA minor groove interactions in substrate recognition by the M.SinI and M.EcoRII DNA (cytosine-5) methyltransferases.
<3>Nucleic Acids Res.
<4>29
<5>3188-3194
<6>2001
<7>The SinI and EcoRII DNA methyltransferases recognize sequences (GG(A)/(T)CC and CC(A)/(T)GG,
respectively), which are characterized by an (A)/(T) ambiguity. Recognition of the A.T and T.A
base pair was studied by in vitro methyltransferase assays using oligonucleotide substrates
containing a hypoxanthine.C base pair in the central position of the recognition sequence.
Both enzymes methylated the substituted oligonucleotide with an efficiency that was comparable
to methylation of the canonical substrate. These observations indicate that M.SinI and
M.EcoRII discriminate between their canonical recognition site and the site containing a G.C
or a C.G base pair in the center of the recognition sequence (GG(G)/(C)CC and CC(G)/(C)GG,
respectively) by interaction(s) in the DNA minor groove. M.SinI mutants displaying a decreased
capacity to discriminate between the GG(A)/(T)CC and GG(G)/(C)CC sequences were isolated by
random mutagenesis and selection for the relaxed specificity phenotype. These mutations led to
amino acid substitutions outside the variable region, previously thought to be the sole
determinant of sequence specificity. These observations indicate that (A)/(T) versus (G)/(C)
discrimination is mediated by interactions between the large domain of the methyltransferase
and the minor groove surface of the DNA.

<>

<1>Kiss, A., Sain, B., Csordas-Toth, E., Venetianer, P.
<2>A new sequence-specific endonuclease (Bsp) from Bacillus sphaericus.
<3>Gene
<4>1
<5>323-329
<6>1977
<7>A new restriction endonuclease has been isolated from Bacillus sphaericus R.
The purification procedure includes Bio-Gel filtration (NH4)2SO4 fractionation
and phosphocellulose chromatography.  After the phosphocellulose step the
enzyme preparation is free of non-specific nucleases.  Bsp cleaves
double-stranded DNA with the same specificity as Bacillus subtilis (Bsu) and
Haemophilus aegyptius (HaeIII) restriction endonucleases, as concluded from
digests and double-digests of PhiX174 replicative form DNA with Bsu and Bsp.
The 5'-terminal nucleotide of the cleavage products was shown to be C.
Bacillus sphaericus R produces Bsp in extremely large quantities and the enzyme
can be easily purified in high yield.

<>

<1>Kiss, A., Weinhold, E.
<2>Functional reassembly of split enzymes on-site: a novel approach for highly sequence-specific targeted DNA methylation.
<3>Chembiochem
<4>9
<5>351-353
<6>2008
<7>In mammalian genomes, a significant fraction of the cytosine residues is methylated at the
5-position, and this modified nucleobase is found in 5'CG-3' sequences.  The methylation
patterns of the genome changes during ontogenesis, depends on the tissue and can substantially
differ in several diseases, notably cancer.  We are only at the beginning of understanding the
biological role of DNA methylation in higher organisms, but the emerging view is that
methylation of the promoter region of a substantial fraction of genes leads to transcriptional
inactivation (gene silencing).

<>

<1>Kiss, H. et al.
<2>Complete genome sequence of the filamentous gliding predatory bacterium Herpetosiphon aurantiacus type strain (114-95).
<3>Standards in Genomic Sciences
<4>5
<5>356-370
<6>2011
<7>Herpetosiphon aurantiacus Holt and Lewin 1968 is the type species of the genus Herpetosiphon,
which in turn is the type genus of the family Herpetosiphonaceae, type family of the order
Herpe-tosiphonales in the phylum Chloroflexi. H. aurantiacus cells are organized in filaments
which can rapidly glide. The species is of interest not only because of its rather isolated
position in the tree of life, but also because Herpetosiphon ssp. were identified as predators
capable of facultative pre-dation by a wolf pack strategy and of degrading the prey organisms
by excreted hydrolytic en-zymes. The genome of H. aurantiacus strain 114-95T is the first
completely sequenced genome of a member of the family Herpetosiphonaceae. The 6,346,587 bp
long chromosome and the two 339,639 bp and 99,204 bp long plasmids with a total of 5,577
protein-coding and 77 RNA genes was sequenced as part of the DOE Joint Genome Institute
Program DOEM 2005.

<>

<1>Kiss, H. et al.
<2>Complete genome sequence of 'Thermobaculum terrenum' type strain (YNP1).
<3>Standards in Genomic Sciences
<4>3
<5>153-162
<6>2010
<7>'Thermobaculum terrenum' Botero et al. 2004 is the sole species within the proposed genus
'Thermobaculum'. Strain YNP1(T) is the only cultivated member of
an acid tolerant, extremely thermophilic species belonging to a phylogenetically
isolated environmental clone group within the phylum Chloroflexi. At present, the
name 'Thermobaculum terrenum' is not yet validly published as it contravenes Rule
30 (3a) of the Bacteriological Code. The bacterium was isolated from a slightly
acidic extreme thermal soil in Yellowstone National Park, Wyoming (USA).
Depending on its final taxonomic allocation, this is likely to be the third
completed genome sequence of a member of the class Thermomicrobia and the seventh
type strain genome from the phylum Chloroflexi. The 3,101,581 bp long genome with
its 2,872 protein-coding and 58 RNA genes is a part of the Genomic Encyclopedia
of Bacteria and Archaea project.

<>

<1>Kiss, H. et al.
<2>Complete genome sequence of Denitrovibrio acetiphilus type strain (N2460).
<3>Standards in Genomic Sciences
<4>2
<5>270-279
<6>2010
<7>Denitrovibrio acetiphilus Myhr and Torsvik 2000 is the type species of the genus
Denitrovibrio in the bacterial family Deferribacteraceae. It is of phylogenetic
interest because there are only six genera described in the family
Deferribacteraceae. D. acetiphilus was isolated as a representative of a
population reducing nitrate to ammonia in a laboratory column simulating the
conditions in off-shore oil recovery fields. When nitrate was added to this
column undesirable hydrogen sulfide production was stopped because the sulfate
reducing populations were superseded by these nitrate reducing bacteria. Here we
describe the features of this marine, mesophilic, obligately anaerobic organism
respiring by nitrate reduction, together with the complete genome sequence, and
annotation. This is the second complete genome sequence of the order
Deferribacterales and the class Deferribacteres, which is the sole class in the
phylum Deferribacteres. The 3,222,077 bp genome with its 3,034 protein-coding and
51 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Kita, A., Iwasaki, Y., Sakai, S., Okuto, S., Takaoka, K., Suzuki, T., Yano, S., Sawayama, S., Tajima, T., Kato, J., Nishio, N., Murakami, K., Nakashimada, Y.
<2>Development of genetic transformation and heterologous expression system in carboxydotrophic thermophilic acetogen Moorella thermoacetica.
<3>J. Biosci. Bioeng.
<4>115
<5>347-352
<6>2013
<7>To develop a microbial production platform based on hydrogen and carbon dioxide, a genetic
transformation system for the thermophilic acetogen
Moorella thermoacetica ATCC39073 was developed. The uracil auxotrophic
strain dpyrF was constructed by disrupting pyrF for orotate
monophosphate decarboxylase. The transformation plasmids were
methylated by restriction methylases of M. thermoacetica to avoid the
decomposition of introduced plasmids by restriction-modification
system. Reintroduction of native pyrF into the mutant by homologous
recombination ensured recovery from uracil auxotrophy. To test
heterologous gene expression in dpyrF, the lactate dehydrogenase (LDH)
gene (T-ldh) from Thermoanaerobacter pseudethanolicus ATCC33223 was
electroporated into dpyrF with a promoter of the
glyceraldehyde-3-phosphate dehydrogenase (G3PD) gene of M.
thermoacetica ATCC39073. The resulting transformant (C31) successfully
transcribed T-ldh and exhibited higher LDH activity than ATCC39073 and
dpyrF, yielding 6.8 mM of lactate from fructose, whereas ATCC39073 did
not produce lactate.

<>

<1>Kita, K.
<2>Type II restriction-modification gene coded by the E. coli genome. Vestiges of horizontal transfer via phages.
<3>Kagaku To Seibutsu
<4>41
<5>348-350
<6>2003
<7>
<>

<1>Kita, K.
<2>Molecular evolution of restriction endonucleases.
<3>Baiosaiensu to Indasutori
<4>67
<5>254-257
<6>2009
<7>
<>

<1>Kita, K., Hiraoka, N., Kimizuka, F., Obayashi, A.
<2>Determination of the cleavage site of restriction enzyme, AccII, using synthetic oligonucleotide.
<3>Agric. Biol. Chem.
<4>48
<5>531-532
<6>1984
<7>None

<>

<1>Kita, K., Hiraoka, N., Kimizuka, F., Obayashi, A., Kojima, H., Takahashi, H., Saito, H.
<2>Interaction of the restriction endonuclease ScaI with its substrates.
<3>Nucleic Acids Res.
<4>13
<5>7015-7024
<6>1985
<7>The kinetic constants of the site-specific endonuclease, ScaI, for various
substrates were determined.  We estimated Vmax and Km for octa-, deca-,
dodeca-, and hexadecanucleotides and for plasmid pBR322 DNA.  Vmax for these
substrates were close, but Km were quite different (in decreasing order, octa-
> deca-, dodeca-, hexadeca- > pBR322).  The results were discussed with respect
to the tertiary structure of substrate.

<>

<1>Kita, K., Hiraoka, N., Oshima, A., Kadonishi, S., Obayashi, A.
<2>New restriction endonuclease with specificity of the DNA sequence TCCGGA produced by Acinetobacter calcoaceticus ferm BP-935.
<3>United Kingdom Patent Office
<4>GB 2183657
<5>
<6>1987
<7>A restriction enzyme having the following properties: (a) action and substrate specificity.
Recognises the base sequence in a double-stranded deoxyribonucleic acid molecule as shown
below, and cleaves it at the arrow-marked sites, T^CCGGA; (b) optimal pH: approximately 8.5;
(c) stable pH range: 6.0 to 9.5; (d) optimal working temperature: 60 to 65 deg.C.

<>

<1>Kita, K., Hiraoka, N., Oshima, A., Kadonishi, S., Obayashi, A.
<2>New restriction endonuclease with specificity of the DNA sequence TCCGGA produced by Acinetobacter calcoaceticus ferm BP-935.
<3>German Patent Office
<4>DE 3640382
<5>
<6>1987
<7>A restriction enzyme having the following properties: (a) action and substrate specificity.
Recognises the base sequence in a double-stranded deoxyribonucleic acid molecule as shown
below, and cleaves it at the arrow-marked sites, T^CCGGA; (b) optimal pH: approximately 8.5;
(c) stable pH range: 6.0 to 9.5; (d) optimal working temperature: 60 to 65 deg.C.

<>

<1>Kita, K., Hiraoka, N., Oshima, A., Kadonishi, S., Obayashi, A.
<2>AccIII, a new restriction endonuclease from Acinetobacter calcoaceticus.
<3>Nucleic Acids Res.
<4>13
<5>8685-8694
<6>1985
<7>A new site-spectific restriction endonuclease, AccIII, was isolated from Acinetobacter
calcoaceticus. AccIII recognizes T^CCGGA and cleaves at the position shown by the arrow.
AccIII activity was inhibited by adenine methylation at the overlapping dam methylase
recognition sequence.

<>

<1>Kita, K., Kawakami, H., Tanaka, H.
<2>Evidence for horizontal transfer of the EcoT38I restriction-modification gene to chromosomal DNA by the P2 phage and diversity of defective P2 prophages in Escherichia coli TH38 strains.
<3>J. Bacteriol.
<4>185
<5>2296-2305
<6>2003
<7>A DNA fragment carrying the genes coding for a novel EcoT38I restriction endonuclease
(R.EcoT38I) and EcoT38I methyltransferase (M.EcoT38I), which
recognize G(A/G)GC(C/T)C, was cloned from the chromosomal DNA of
Escherichia coli TH38. The endonuclease and methyltransferase genes were
in a head-to-head orientation and were separated by a 330-nucleotide
intergenic region. A third gene, the C.EcoT38I gene, was found in the
intergenic region, partially overlapping the R.EcoT38I gene. The gene
product, C.EcoT38I, acted as both a positive regulator of R.EcoT38I gene
expression and a negative regulator of M.EcoT38I gene expression.
M.EcoT38I purified from recombinant E. coli cells was shown to be a
monomeric protein and to methylate the inner cytosines in the recognition
sequence. R.EcoT38I was purified from E. coli HB101 expressing M.EcoT38I
and formed a homodimer. The EcoT38I restriction (R)-modification (M)
system (R-M system) was found to be inserted between the A and Q genes of
defective bacteriophage P2, which was lysogenized in the chromosome at
locI, one of the P2 phage attachment sites observed in both E. coli K-12
MG1655 and TH38 chromosomal DNAs. Ten strains of E. coli TH38 were
examined for the presence of the EcoT38I R-M gene on the P2 prophage.
Conventional PCR analysis and assaying of R activity demonstrated that all
strains carried a single copy of the EcoT38I R-M gene and expressed R
activity but that diversity of excision in the ogr, D, H, I, and J genes
in the defective P2 prophage had arisen.

<>

<1>Kita, K., Kotani, H., Hiraoka, N., Nakamura, T., Yonaha, K.
<2>Overproduction and crystallization of FokI restriction endonuclease.
<3>Nucleic Acids Res.
<4>17
<5>8741-8753
<6>1989
<7>To overproduce FokI endonuclease (R. FokI) in an Escherichia coli system, the coding region of
R.FokI predicted from the nucleotide sequence was generated from the FokI operon and joined to
the tac promoter of an expression vector, pKK223-3.  By introduction of the plasmid into E.
coli UT481 cells expressing the FokI methylase gene, the R.FokI activity was overproduced
about 30-fold, from which R. FokI was purified in amounts sufficient for crystallization.  The
removal of a stem-loop structure immediately upstream of the R. FokI coding region was
essential for overproduction.

<>

<1>Kita, K., Kotani, H., Hiraoka, S., Nakamura, T.
<2>FokI Restriction and modifying system enzymic gene and production thereof.
<3>Japanese Patent Office
<4>JP 1990142478 A, JP 2142478 A
<5>
<6>1990
<7>
<>

<1>Kita, K., Kotani, H., Ohta, H., Yanase, H., Kato, N.
<2>StsI, a new FokI isoschizomer from Streptococcus sanguis 54, cleaves 5' GGATG(N)10/14 3'.
<3>Nucleic Acids Res.
<4>20
<5>618
<6>1992
<7>StsI, an isoschizomer of FokI has been isolated from Streptococcus sanguis 54.
StsI recognizes the non-palindromic sequence 5'-GGATG-3'.  Unlike its
isoschizomer, StsI cleaves DNA 10 nucleotides to the right from the noted
recognition sequence and 14 nucleotides to the right on the opposite strand.
StsI was purified from the cell extracts by combined chromatographies on
phosphocellulose, DEAE-cellulose, heparin-Sepharose, hydroxylapatite, and
Affi-Gel Blue agarose.  The purified enzyme was homogeneous on
SDS-polyacrylamide gel electrophoresis.  The molecular mass of the enzyme was
estimated at 70 kDa.  The StsI activity was eluted at 70 kDa from a gel
filtration column.  These results indicated that the active form of StsI was a
monomer.

<>

<1>Kita, K., Kotani, H., Sugisaki, H., Takanami, M.
<2>The FokI restriction-modification system  I. Organization and nucleotide sequences of the restriction and modification genes.
<3>J. Biol. Chem.
<4>264
<5>5751-5756
<6>1989
<7>A DNA fragment that carried the genes coding for FokI endonuclease and
methylase was cloned from the chromosomal DNA of Flavobacterium okeanokoites,
and the coding regions were assigned to the nucleotide sequence by deletion
analysis.  The methylase gene was 1941 base pairs (bp) long, corresponding to a
protein of 647 amino acid residues (Mr=75622), and the endonuclease gene was
1749 bp long, corresponding to a protein of 583 amino acid residues (Mr=66216).
The assignment of the methylase gene was further confirmed by analysis of the
N-terminal amino acid sequence.  The endonuclease gene was downstream from the
methylase gene in the same orientation, separated by 69 bp.  The promoter site,
which could be recognized by Escherichia coli RNA polymerase, was upstream from
the methylase gene, and the sequences adhering to the ribosome-binding sequence
were identified in front of the respective genes.  Analysis of the gene
products expressed in E. coli cells by gel filtration and sodium dodecyl
sulfate-polyacrylamide gel electrophoresis indicated that the molecular weights
of both enzymes coincided well with the values estimated from the nucleotide
sequences, and that the monomeric forms were catalytically active.  No
significant similarity was found between the sequences of the two enzymes.
Sequence comparison with other related enzymes indicated that FokI methylase
contained two copies of a segment of tetra-amino acids which is characteristic
of adenine-specific methylase.

<>

<1>Kita, K., Suisha, M., Kotani, H., Yanase, H., Kato, N.
<2>Cloning and sequence analysis of the StsI restriction-modification gene: presence of homology to FokI restriction-modification enzymes.
<3>Nucleic Acids Res.
<4>20
<5>4167-4172
<6>1992
<7>StsI endonuclease (R.StsI), a type IIs restriction endonuclease found in Streptococcus sanguis
54, recognizes the same sequence as FokI but cleaves at different positions. A DNA fragment
that carried the genes for R.StsI and StsI methylase (M.StsI) was cloned from the chromosomal
DNA of S. sanguis 54, and its nucleotide sequence was analyzed. The endonuclease gene was
1,806 bp long, corresponding to a protein of 602 amino acid residues (Mr=68,388), and the
methylase gene was 1,959 bp long, corresponding to a protein of 653 amino acids residues (Mr
=76,064). The assignment of the endonuclease gene was confirmed by analysis of the N-terminal
amino acid sequence. Genes for the two proteins were in a tail-to-tail orientation, separated
by a 131-nucleotide intercistronic region. The predicted amino acid sequences between the StsI
system and the FokI system showed a 49% identity between the methylases and a 30% identity
between the endonucleases. The sequence comparison of M.StsI with various methylases showed
that the N-terminal half of M.StsI matches M.NlaII, and the C-terminal half matches adenine
methylases that recognize GATC and GATATC.

<>

<1>Kita, K., Suisha, M., Shintoh, M., Yanase, H., Kato, N.
<2>Overproduction and characterization of the StsI restriction endonuclease.
<3>Gene
<4>169
<5>69-73
<6>1996
<7>The StsI restriction endonuclease (R.StsI), a class-IIS restriction endonuclease, found in
Streptococcus sanguis 54, is a heteroschizomer of R.FokI, which recognizes 5M-U-GGATG-3M-U.
To overproduce R.StsI in Escherichia coli, the coding region of R.StsI was joined to the tac
promoter of an expression vector, pKK223-3.  By introduction of the plasmid into E. coli UT481
cells expressing the fokIM gene, R.StsI activity was overproduced, from which R.StsI was
purified homogeneously.  We compared the properties of R.StsI with those of R.FokI.  The
optimum reaction conditions for R.StsI were quite different from those for R.FokI.  R.StsI is
an acidic protein (pI 6.3).  Anti-R.StsI serum did not cross-react with R.FokI, indicating
three-dimensional structural dissimilarity.  The domain structure of R.StsI was elucidated by
digestion with trypsin.  In the presence of substrate DNA, R.StsI was digested to yield 45-kDa
N-terminal and 23-kDa C-terminal fragments.  The amino-acid sequences around the trypsin
cleavage sites of R.StsI and R.FokI were quite homologous.

<>

<1>Kita, K., Takasaki, Y.
<2>Enzymology of restriction enzymes.
<3>Seikagaku
<4>66
<5>1237-1245
<6>1994
<7>A review

<>

<1>Kita, K., Tsuda, J., Kato, T., Okamoto, K., Yanase, H., Tanaka, M.
<2>Evidence of horizontal transfer of the EcoO109I restriction-modification gene to Escherichia coli chromosomal DNA.
<3>J. Bacteriol.
<4>181
<5>6822-6827
<6>1999
<7>A DNA fragment carrying the genes coding for EcoO109I endonuclease and EcoO109I methylase,
which recognize the nucleotide sequence 5'-(A/G) GGNCC(C/T)-3', was cloned from the
chromosomal DNA of Escherichia coli H709c.  The EcoO109I restriction-modification system was
found to be inserted between the int and psu genes from satellite bacteriophage P4, which were
lysogenized in the chromosome at the P4 phage attachment site of the corresponding leuX gene
observed in E. coli K-12 chromosomal DNA.  The sid gene of the prophage was inactivated by
insertion of one copy of IS21.  These findings may shed light on the horizontal transfer and
stable maintenance of the R-M system.

<>

<1>Kita, K., Tsuda, J., Nakai, S.-y.
<2>C.EcoO109I, a regulatory protein for production of EcoO109I restriction endonuclease, specifically binds to and bends DNA upstream of its translational start site.
<3>Nucleic Acids Res.
<4>30
<5>3558-3565
<6>2002
<7>The EcoO109I restriction-modification system, which recognizes 5'-(A/G)GGNCC(C/T)-3', has
been cloned, and contains convergently transcribed endonuclease and methylase. The role and
action mechanism of the gene product, C.EcoO109I, of a small open reading frame located
upstream of ecoO109IR were investigated in vivo and in vitro.  The results of deletion
analysis suggested that C.EcoO109I acts as a positive regulator of ecoO109IR expression but
has little effect on ecoO109IM expression. Assaying of promoter activity showed that the
expression of ecoO109IC was regulated by its own gene product, C.EcoO109I. C.EcoO109I was
overproduced as a His-tag fusion protein in recombinant Escherichia coli HB101 and purified to
homogeneity. C.EcoO109I exists as a homodimer, and recognizes and binds to the DNA sequence
5'-CTAAG(N)(5)CTTAG-3' upstream of the ecoO109IC translational start site. It was also shown
that C.EcoO109I bent the target DNA by 54 +/- 4 degrees.

<>

<1>Kita, K., Tsuda, J., Nishigaki, R.
<2>Characterization and overproduction of EcoO109I methyltransferase.
<3>Biosci. Biotechnol. Biochem.
<4>65
<5>2512-2518
<6>2001
<7>In order to characterize EcoO109I methyltransferase, a recombinant Escherichia coli clone that
overproduces the enzyme was constructed.
The coding region of M.EcoO109I was joined to the lac promoter of an
expression vector, pUC118, and the resulting plasmid was introduced
into E. coli HB101. M.EcoO109I was purified homogeneously from
IPTG-induced cells, and was found to consist of a monomer subunit.
M.EcoO109I uniquely methylates the inner deoxycytidylate residue in the
sequence 5'-(A/G)GGNCC(C/T)-3' to produce 5-methylcytosine. The enzyme
was most active at pH 8.0-8.5 and 50 C. The enzyme activity was
not affected by the addition of Mg2+ or EDTA.

<>

<1>Kitagawa, W., Hata, M., Sekizuka, T., Kuroda, M., Ishikawa, J.
<2>Draft Genome Sequence of Rhodococcus erythropolis JCM 6824, an Aurachin RE Antibiotic Producer.
<3>Genome Announcements
<4>2
<5>e01026-14
<6>2014
<7>Rhodococcus erythropolis JCM 6824 is the producer of the quinoline antibiotic aurachin RE.
This bacterium also degrades and utilizes some aromatic compounds,
such as biphenyl and benzoate. Here, we report the draft genome sequence of this
strain.

<>

<1>Kits, K.D. et al.
<2>Genome Sequence of the Obligate Gammaproteobacterial Methanotroph Methylomicrobium album Strain BG8.
<3>Genome Announcements
<4>1
<5>e00170-13
<6>2013
<7>The complete genome sequence of Methylomicrobium album strain BG8, a methane-oxidizing
gammaproteobacterium isolated from freshwater, is reported.
Aside from a conserved inventory of genes for growth on single-carbon compounds,
M. album BG8 carries a range of gene inventories for additional carbon and
nitrogen transformations but no genes for growth on multicarbon substrates or for
N fixation.

<>

<1>Kittichotirat, W., Good, N.M., Hall, R., Bringel, F., Lajus, A., Medigue, C., Smalley, N.E., Beck, D., Bumgarner, R., Vuilleumier, S., Kalyuzhnaya, M.G.
<2>Genome Sequence of Methyloversatilis universalis FAM5T, a Methylotrophic Representative of the Order Rhodocyclales.
<3>J. Bacteriol.
<4>193
<5>4541
<6>2011
<7>Rhodocyclales are representative of versatile bacteria that are able to utilize a wide variety
of organic compounds for growth, but only few
strains have been isolated in pure culture thus far. Here we present the
genome sequence of Methyloversatilis universalis FAM5(T), the first
cultivable methylotrophic member of the order.

<>

<1>Kittler, L., Bell, A., Baguley, B.C., Lober, G.
<2>Inhibition of restriction endonucleases by DNA sequence-reading ligands.
<3>Biochem. Mol. Biol. Int.
<4>40
<5>263-272
<6>1996
<7>DNA sequence-reading bisquaternary ammonium heterocycles SN 6570, SN 6999, SN 6053, SN 6132,
SN 6131, SN 18071 and the non-specific binders SN 6113, SN 5754, SN 6324, and SN 4094
influence the enzymatic activity of restriction endonucleases in different manners.  A
prerequisite for sequence-specific ligand interaction is a dAdT run of at least four base
pairs.  The sequence-specific binders inhibit the cleavage activity of restriction
endonucleases EcoRI, SspI, and DraI with four and six dAdT base pairs in their restriction
sites, while the activity of SalI and BamHI with less than four dAdT base pairs in their
recognition motifs remains unaffected.  On the contrary, the non-specific binding DNA ligands
are incapable of suppressing the digestion for restriction nucleases under research.  These
results are in line with our footprint data.  The inhibitory effect is independent of the
number of cleavage sites in DNA and of whether the macromolecule exists in the ccc or lds
conformation.  Sequence specific binding of the ligand SN 6043 in close vicinity to the
cleavage sites of restriction endonuclease DraI also interferes with enzyme inhibition.

<>

<1>Kittler, L., Bell, A., Baguley, B.C., Loeber, G.
<2>Sequence specific modulation of DNA restriction enzyme cleavage by minor groove binders.
<3>Biol. Chem.
<4>379
<5>519-525
<6>1998
<7>The inhibition of restriction endonuclease cleavage by a series of bisquaternary ammonium
derivatives which bind to the minor groove of DNA has been studied.  The derivatives
considered included six sequence-selective binders (SN 6570, SN 6999, SN 6050, SN 6132, SN
6131 and SN 18071) and four non-specific binders (SN 6113, SN 5754, SN 6324 and SN 4094) and
can be distinguished by their activity on restriction endonucleases.  Digestion experiments
with pUC19 DNA were monitored electrophoretically using the transition of the covalently
closed circular DNA into the linear double stranded one.  Only the sequence-specific binders
inhibit the cleavage activity of restriction endonucleases EcoRI, SspI and DraI with four and
six dAdT-base pairs within their restriction sites, while the activity of SalI and BamHI with
less than four dAdT-sequences was unaffected.  In contrast, the non-specific binding ligands
were incapable of suppressing enzyme digestion.  The inhibition of the restriction
endonuclease PvuII indicates that ligand binding in close vicinity to the cleavage sites is
also involved in the enzyme inhibition.  The dAdT-content in proximity to the palindromic
sequences of three DraI cutting sites in pUC19DNA explains why the derivative SN 6053 protects
these sequences in different manners.  Gel shift experiments indicated that BQA-derivatives
inhibit the DNA-enzyme complex formation if the ligand was added to the DNA before the enzyme.
In contrast, complex formation between DNA and enzyme remained unchanged when the enzyme was
added first.

<>

<1>Kivisto, A., Larjo, A., Ciranna, A., Santala, V., Roos, C., Karp, M.
<2>Genome Sequence of Halanaerobium saccharolyticum subsp. saccharolyticum Strain DSM 6643T, a Halophilic Hydrogen-Producing Bacterium.
<3>Genome Announcements
<4>1
<5>e00187-13
<6>2013
<7>Halanaerobium saccharolyticum is a halophilic anaerobic fermentative bacterium capable of
producing hydrogen, a potential future energy carrier molecule. The
high-quality draft genome of H. saccharolyticum subsp. saccharolyticum strain DSM
6643(T) consists of 24 contigs for 2,873,865 bp with a G+C content of 32.3%.

<>

<1>Kivisto, R.I., Kovanen, S., Skarp-de-Haan, A., Schott, T., Rahkio, M., Rossi, M., Hanninen, M.-L.
<2>Evolution and Comparative Genomics of Campylobacter jejuni ST-677 Clonal Complex.
<3>Genome Biol. Evol.
<4>6
<5>2424-2438
<6>2014
<7>Campylobacter is the most common bacterial cause of gastroenteritis in the European Union with
over 200,000 laboratory-confirmed cases reported annually. This is the first study to describe
findings related to comparative genomics analyses of the sequence type (ST)-677 clonal complex
(CC), a Campylobacter jejuni lineage associated with bacteremia cases in humans. We performed
whole-genome sequencing, using Illumina HiSeq sequencing technology, on five related ST-677 CC
isolates from two chicken farms to identify microevolution taking place at the farms. Our
further aim was to identify novel putative virulence determinants from the ST-677 CC genomes.
For this purpose, clinical isolates of the same CC were included in comparative genomic
analyses against well-known reference strains of C. jejuni. Overall, the ST-677 CC was
recognized as a highly clonal lineage with relatively small differences between the genomes.
Among the farm isolates differences were identified mainly in the lengths of the homopolymeric
tracts in genes related to the capsule, lipo-oligosaccharide, and flagella. We identified
genomic features shared with C. jejuni subsp. doylei, which has also been shown to be
associated with bacteremia in humans. These included the degradation of the cytolethal
distending toxin operon and similarities between the capsular polysaccharide biosynthesis
loci. The phase-variable GDP-mannose 4,6-dehydratase (EC 4.2.1.47) (wcbK, CAMP1649),
associated with the capsular polysaccharide biosynthesis locus, may play a central role in
ST-677 CC conferring acid and serum resistance during different stages of infection.
Homology-based searches revealed several additional novel features and characteristics,
including two putative type Vb secretion systems and a novel restriction
modification/methyltransferase gene cluster, putatively associated with pathogenesis and niche
adaptation.

<>

<1>Kiwaki, M.
<2>Restriction enzymes.
<3>Bifizusukin Yakurutokabu
<4>0
<5>118-122
<6>2009
<7>
<>

<1>Kjems, J., Garrett, R.A.
<2>An intron in the 23S ribosomal RNA gene of the archaebacterium Desulfurococcus mobilis.
<3>Nature
<4>318
<5>675-677
<6>1985
<7>The archaebacteria have been defined, at a molecular level, as constituting a third primary
kingdom consisting of the methanogens, the extreme halophiles, and the sulphur-dependent
extreme thermophiles. In reaching this conclusion, Woese and colleagues used the 16S ribosomal
RNA as an approximate chronometer for evolutionary time and demonstrated that, at a nucleotide
sequence level, the archaebacteria are as different from the eubacteria and eukaryotes as the
latter kingdoms are from one another. Current research on archaebacteria is yielding valuable
insights into the evolutionary relationships between archaebacteria, eubacteria and
eukaryotes, and into the early forms of cellular life. Here, we extend this knowledge by
providing the first evidence for the occurrence of an intron within any prokaryotic ribosomal
RNA. The intron was found within the 23S rRNA gene of the sulphur-dependent and anaerobic
Thermoproteale Desulfurococcus mobilis, which was isolated from hot acidic springs in Iceland
at temperatures up to 97oC. The intron contains 622 base pairs (bp); it is very A+T-rich (65%)
compared with the 23S rRNA gene (34%), and it exhibits a large open reading frame. The
splicing site occurs in domain IV of the 23S RNA at a position close to that of an intron of
the lower eukaryote Physarum polycephalum; the intron does not readily fall into one of the
three classes of eukaryotic nuclear introns because it has features in common with those of
classes I (rRNA) and III (transfer RNA).

<>

<1>Kladde, M.P., Simpson, R.T.
<2>Rapid detection of functional expression of C-5-DNA methytransferases in yeast.
<3>Nucleic Acids Res.
<4>26
<5>1354-1355
<6>1998
<7>We have previously employed the cytosine-5-DNA methyltransferase, M.SssI, as a probe for
chromatin architecture in intact cells.  Although M.SssI offers the highest resolution of any
currently available MTase, the difficulty in establishing stable, methylation-positive strains
poses a barrier to its general utility as a chromatin probe.  We describe a single screen for
M.SssI-expressing strains that eliminates the purification of PCR products amplified from
bisulfite-treated DNA, use of radioisotopes, polyacrylamide sequencing gel electrophoresis,
and autoradiography.  The high throughput of the method now makes it feasible to introduce
M.SssI into a vareity of wild-type and mutant genetic backgrounds.

<>

<1>Kladde, M.P., Simpson, R.T.
<2>Chromatin structure mapping in vivo using methyltransferases.
<3>Methods Enzymol.
<4>274
<5>214-233
<6>1996
<7>Realization of the role of chromatin structure in the function of RNA polymerases in
eukaryotic cells has grown rapidly in the past decade.  Effects of the nucleosomal
organization of DNA on transcriptional initiation have been documented both in vivo and in
vitro.  Several studies have addressed the mechanical difficulties faced by RNA polymerase
encountering a nucleosome during transcriptional elongation, offering varying possible
resolutions to this problem.   Above the local difficulties which polymerases must face in the
basic nucleosomal organization of chromatin, there are higher order structures, including
chromatin domains, which have been implicated in more global silencing, such as position
effect variegation, X-chromosome inactivation, epigenetic inheritance, and centromeric and
telomeric repression.  All these considerations make methods for assessment of chromatin
structure of importance to those scientists interested in transcriptional mechanisms in
eukaryotic cells.  Historically, chromatin structure investigations have relied on enzymatic
(micrococcal nuclease or DNase I) or chemical (hydroxyl radical, methidium propyl-EDTA-iron,
cooper-phenanthroline) probes to degrade DNA; proteins, either histones or non-histone
regulatory proteins, left a footprint on DNA by blocking the access of the reagent to the
nucleic acid, enabling deductions about potential structures of the nucleoprotein complex.  In
general, isolation of nuclei is required prior to analysis of chromatin.  This makes the
detection of the labile chromatin constituents problematic, limiting conclusions about the
structure of transcribed or repressed genes.

<>

<1>Kladde, M.P., Simpson, R.T., Xu, M.
<2>Cloning, sequence and characterization of cytosine-5 DNA methyltransferase CviPI from Chlorella virus NYs-1 and use of CviPI in  high resolution chromatin mapping.
<3>US Patent Office
<4>US 6492168 B
<5>21
<6>2002
<7>A novel cytosine-5 DNA methyltransferase, isolated from Chlorella virus NYs-1, and its encoded
enzyme are disclosed.  The methyltransferase recognizes a GpC dinucleotide in DNA.  Methods of
using the novel methyltransferase in high resolution chromatin mapping and related techniques
are also disclosed.

<>

<1>Kladde, M.P., Simpson, R.T., Xu, M.
<2>Methyltransferase gene and enzyme.
<3>US Patent Office
<4>US 7034116 B
<5>
<6>2006
<7>A novel cytosine-5 DNA methyltransferase, isolated from Chlorella virus NYs-1, and its encoded
enzyme are disclosed.  The methyltransferase recognizes a GrpC dinucleotide in DNA.  Methods
of using the novel methyltransferase in high resolution chromatin mapping and related
techniques are also disclosed.

<>

<1>Kladde, M.P., Simpson, R.T., Xu, M.
<2>Methyltransferase gene and enzyme.
<3>US Patent Office
<4>US 20030073197
<5>
<6>2003
<7>DERWENT ABSTRACT: NOVELTY - An isolated nucleic acid molecule encoding a cytosine-5 DNA
methyltransferase that recognizes a GpC dinucleotide in DNA, is new. DETAILED DESCRIPTION -
INDEPENDENT CLAIMS are also included for: (1) a recombinant DNA molecule comprising the
nucleic acid molecule cited above, operably inserted into a vector for transforming cells; (2)
cells transformed with the recombinant DNA molecule cited above; (3) a cultured cell line of
the transformed cells; (4) a living organism comprising the transformed cells; (5) an
oligonucleotide between about 10 and 100 nucleotides in length, which specifically hybridizes
with a portion of the nucleic acid molecule cited above; (6) a polypeptide produced by
expression of the nucleic acid molecule cited above; (7) antibodies immunologically specific
for the above polypeptide; and (8) mapping DNA-protein interactions in DNA in a cell.
BIOTECHNOLOGY - Preferred Nucleic Acid: The nucleic acid molecule comprises a sequence of 1141
bp (S1) fully defined in the specification, or its natural variant. It is isolated from
Chlorella virus NYs-1. The encoded cytosine-5 DNA methyltransferase has a sequence of 362
amino acids (S2) fully defined in the specification and is catalytically active and recognizes
the GpC dinucleotide. The nucleic acid molecule may also comprise a sequence that hybridizes
with an antisense strand of S1. The oligonucleotide hybridizes with the portion of the nucleic
acid molecule that encodes a variable region of the cytosine-5 DNA methyltransferase between
motif VIII and motif IX of the methyltransferase, preferably, motif IX. The oligonucleotide
may also hybridize with a portion of a nucleic acid molecule selected from: (a) a portion of a
double-stranded nucleic acid molecule comprising S1, the portion extending from about
nucleotide 630 to a stop codon near a 3' terminus of S1, from about nucleotide 684-744 of S1,
or from about nucleotide 858-903 of S1; and (b) a portion of a double-stranded nucleic acid
molecule comprising S1, which encodes the portion of S2 extending from about residue 200 to
the carboxyl terminus of S2, from about residue 218-248 of S2, or from about residue 276-291
of S2. Preferred Cell: The cells transformed with the recombinant DNA molecule originated from
bacteria, yeast, fungi, insects, plants or mammals. Preferred Method: The method of mapping
DNA-protein interactions in DNA in a cell comprises providing a sample of cells transformed
with the above nucleic acid molecule, growing a test culture of the transformed cells under
conditions enabling production of the methyltransferase, growing a control culture of
equivalent cells that do not produce the methyltransferase, isolating the DNA from the test
culture and the control culture, exposing the DNA from the control culture to the cytosine-5
methyltransferase, and comparing the cytosine methylation of the DNA from the test culture
with the cytosine methylation of the DNA from the control culture, a decrease in extent of
methylation in the DNA of the test culture being proportional to the amount of DNA-protein
interaction occurring in the DNA in the cell. The above method further comprises comparing a
pattern of methylation in a selected region of the DNA from the test culture and the control
culture, a change in the methylation pattern in the respective DNA being indicative of a
location of a DNA-protein interaction in the DNA of the cell. The DNA-protein interaction is
an interaction between nucleosome proteins and chromosomal DNA, or between a transcriptional
regulator protein and a transcriptional response element in the DNA. Preparation: The nucleic
acid molecule was prepared using standard isolation techniques. ACTIVITY - None given.
MECHANISM OF ACTION - Gene therapy. USE - The nucleic acid molecule and polypeptide are useful
for high-resolution analysis and manipulation of protein-DNA interactions in chromatin. In
addition, the nucleic acids and polypeptides may find utility in gene therapy applications.
EXAMPLE

<>

<1>Kladde, M.P., Xu, M., Simpson, R.T.
<2>DNA methyltransferases as probes of chromatin structure in vivo.
<3>Methods Enzymol.
<4>304
<5>431-447
<6>1999
<7>A complementary strategy to conventional methods for probing chromatin organization utilizes
the expression of foreign DNA methyltransferases in intact Saccharomyces cerevisiae.  At
current levels of expression of several foreign methyltransferases in yeast, no deleterious
effects to cellular metabolism have been detected.  The innocuousness of methyltransferase
expression and the absence of endogenous adenine or cytosine methylation in the yeast genome
allow detection of de novo DNA modification in DNA that is directly isolated from engineered,
methylation-proficient cells.  The ability to bypass the isolation of nuclei, thereby avoiding
loss of labile constituents or possible reorganization of chromosome structure, has provided
the motivation for developing methyltransferase probing strategies.

<>

<1>Klaenhammer, T.R.
<2>Plasmid-directed mechanisms for bacteriophage defense in lactic streptococci.
<3>FEMS Microbiol. Lett.
<4>46
<5>313-325
<6>1987
<7>Genetic studies with lactic streptococci have identified a variety of plasmids
coding for systems that interfere with phage adsorption, direct restriction and
modification activities, and disrupt various stages in the phage lytic cycle.
This review describes mechanisms of phage defense that are plasmid-directed in
lactic streptococci, examines the physical and genetic properties of the
plasmids involved, and discusses genetic strategies for construction of
phage-insensitive starter cultures for dairy fermentations.

<>

<1>Klaenhammer, T.R.
<2>Genetic characterization of multiple mechanisms of phage defense from a prototype phage-insensitive strain, Lactococcus lactis ME2.
<3>J. Dairy Sci.
<4>72
<5>3429-3443
<6>1989
<7>Lactococci used as starter cultures in dairy fermentations are highly
susceptible to attack by bacteriophage.  Genetic studies with Lactococcus
lactis ME2, a prototype phage-insensitive strain, have identified
plasmid-encoded defenses, which interfere with phage adsorption, restrict and
modify phages, or abort lytic phage infection.  Restriction and modification
and abortion of phage infection were localized on two distinct
self-transmissible plasmids, pTN20 and pTR2030, respectively, orginating from
L. lactis ME2.  A comparison of the physical and genetic characteristics of
these two conjugative plasmids is presented.  Conjugation and cloning
strategies employed to assemble these complementary mechanisms of phage
resistance will be discussed.  The collective expression of different defense
systems provided a greater phage resistance to dairy lactococci.  Starter
cultures that are recalcitrant to phage attack can be constructed from existing
strains through application of genetic technologies, which assemble
complementary mechanisms of phage defense.

<>

<1>Klaenhammer, T.R.
<2>Development of bacteriophage-resistant strains of lactic acid bacteria.
<3>Food Biotech.
<4>19
<5>675-682
<6>1991
<7>One exciting area that has emerged through genetic analysis of the lactococci is the
definition and practical application of gene systems that provide phage resistance to these
industrially important bacteria.  Naturally occurring phage-insensitive strains have been
characterized and found to harbor multiple defense systems which can act at different points
of the lytic cycle to prevent the successful adsorption, infection, or replication of virulent
phages.  Although phage attack on lactococcal starter cultures is still a major problem for
the cultured dairy products industries, this can now be successfully addressed using genetic
approaches to construct strains which are resistant to the phages often encountered in the
industry.  In this paper I will describe the various phage defense systems that are naturally
present in lactococci, their individual and combined effects, and the genetic strategies which
are currently available to construct phage-insensitive strains for dairy fermentations.  In
addition, I will discuss the molecular responses of virulent phages which have appeared in the
industry following the introduction and use of specialized starter cultures carrying defined
mechanisms of phage resistance.  The genetic routes whereby industrial phages elicit
counter-defenses against lactococcal resistance mechanism provide new and important
information that will facilitate the development of improved starter culture systems and
phage-resistant lactic acid bacteria.

<>

<1>Klaenhammer, T.R., Fitzgerald, G.F.
<2>Bacteriophages and bacteriophage resistance.
<3>Genetics and Biotechnology of Lactic Acid Bacteria., Blackie Academic & Professional, Gasson, M.J., De Vos, W.M., 
<4>0
<5>106-168
<6>1994
<7>Food and dairy fermentations rely on the growth and acid
producing ability of the lactic acid bacteria.  Many of these have
remained as traditional fermentations, where the process is driven by
the natural microflora associated with the raw material.  Increasing
consistency, improved quality and processing efficiencies have followed
the development of controlled fermentations.  These rely on the activity
of a starter culture which is intentionally inoculated in order to drive
the primary fermentation.  However, with the increased control granted
through the repeated use of a defined starter culture comes the potential
for disruption of the fermentation by bacteriophage.

<>

<1>Klaenhammer, T.R., Romero, D., Sing, W., Hill, C.
<2>Molecular analysis of pTR2030 gene systems that confer bacteriophage resistance to Lactococci.
<3>Genetics and Molecular Biology of Streptococci, Lactococci, and Enterococci, American Society for Microbiology, Dunny, G.M., Cleary, P.P., McKay, L.L., Washington
<4>8
<5>124-130
<6>1991
<7>None

<>

<1>Klaenhammer, T.R., Sanozky-Dawes, R.B.
<2>PTN1060, a conjugal plasmid and derivatives thereof that confer phage resistance to group N streptococci.
<3>US Patent Office
<4>US 4883756
<5>
<6>1989
<7>The present invention relates to the plasmid pTN1060 and derivatives thereof which confer
phage restriction and modification activity to group N streptococci. The invention further
relates to microorganisms containing pTN1060 or a derivative thereof and to starter cultures
containing the microorganisms.

<>

<1>Klapatch, T.R., Demain, A.L., Lynd, L.R.
<2>Restriction endonuclease activity in Clostridium thermocellum and Clostridium thermosaccharolyticum.
<3>Appl. Microbiol. Biotechnol.
<4>45
<5>127-131
<6>1996
<7>Clostridium thermocellum cell extracts exhibit specific endonuclease activity with
very little non-specific exonuclease activity at 55oC.  The Dam methylation system of
Escherichia
coli offers complete protection from digestion by C. thermocellum ATCC 27405 cell extracts for
all
DNA tested (totaling >100kb, insuring that most potential restriction sequences have been
exposed).  Based on both the Dam recognition sequence and the similarity of cell extract and
MboI
DNA digests, the C. thermocellum restriction enzyme recognition sequence appears to be
5' GATC 3'.  Cell extracts made from a second thermophile, C. thermosaccarolyticum ATCC
31960 do not exhibit specific endonuclease activity under the conditions tested.  Genomic DNA
from C. thermocellum exhibits a Dam+ phenotype while genomic DNA from C.
thermosaccarolyticum exhibits a Dam- phenotype.

<>

<1>Klassen, J.L., Adams, S.M., Bramhacharya, S., Giles, S.S., Goodwin, L.A., Woyke, T., Currie, C.R.
<2>Draft Genome Sequence of Streptomyces sp. Strain Wigar10, Isolated from a Surface-Sterilized Garlic Bulb.
<3>J. Bacteriol.
<4>193
<5>6999-7000
<6>2011
<7>Streptomyces sp. strain Wigar10 was isolated from a surface-sterilized garlic bulb (Allium
sativum var. Purple Stripe). Its genome encodes
several novel secondary metabolite biosynthetic gene clusters and provides
a genetic basis for further investigation of this strain's chemical
biology and potential for interaction with its garlic host.

<>

<1>Klasson, L., Westberg, J., Sapountzis, P., Naslund, K., Lutnaes, Y., Darby, A.C., Veneti, Z., Chen, L., Braig, H.R., Garrett, R., Bourtzis, K., Andersson, S.G.
<2>The mosaic genome structure of the Wolbachia wRi strain infecting Drosophila simulans.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>5725-5730
<6>2009
<7>The obligate intracellular bacterium Wolbachia pipientis infects around 20% of all insect
species. It is maternally inherited and induces
reproductive alterations of insect populations by male killing,
feminization, parthenogenesis, or cytoplasmic incompatibility. Here, we
present the 1,445,873-bp genome of W. pipientis strain wRi that induces
very strong cytoplasmic incompatibility in its natural host Drosophila
simulans. A comparison with the previously sequenced genome of W.
pipientis strain wMel from Drosophila melanogaster identified 35
breakpoints associated with mobile elements and repeated sequences that
are stable in Drosophila lines transinfected with wRi. Additionally, 450
genes with orthologs in wRi and wMel were sequenced from the W. pipientis
strain wUni, responsible for the induction of parthenogenesis in the
parasitoid wasp Muscidifurax uniraptor. The comparison of these A-group
Wolbachia strains uncovered the most highly recombining intracellular
bacterial genomes known to date. This was manifested in a 500-fold
variation in sequence divergences at synonymous sites, with different
genes and gene segments supporting different strain relationships. The
substitution-frequency profile resembled that of Neisseria meningitidis,
which is characterized by rampant intraspecies recombination, rather than
that of Rickettsia, where genes mostly diverge by nucleotide
substitutions. The data further revealed diversification of ankyrin repeat
genes by short tandem duplications and provided examples of horizontal
gene transfer across A- and B-group strains that infect D. simulans. These
results suggest that the transmission dynamics of Wolbachia and the
opportunity for coinfections have created a freely recombining
intracellular bacterial community with mosaic genomes.

<>

<1>Klaus, S., Hartmann, M., Krugel, H., Roth, M., Walter, F., Rautenstein, Y.I., Solovyeva, N.Y.
<2>Restriction of Streptomyces phage SH5 by endonuclease ShyI from Streptomyces hygroscopicus 0477.
<3>Mol. Gen. Genet.
<4>184
<5>286-288
<6>1981
<7>DNA of the temperate Streptomyces phage SH5 (DNA molecular weight 27 x 106) is
subject to restriction-modification mediated by S. hygroscopicus 0477.  S.
levoris 1331 and 2340.  The restriction endonuclease ShyI (isoschizomeric with
SacII) isolated from S. hygroscopicus 0477 is involved in restriction of
SH5-1331 and SH5-2340 DNAs in S. hygroscopicus 0477.

<>

<1>Kleanthous, H., Garawi, A.A., Miller, C., Tomb, J.F., Oomen, R.P.
<2>Identification of polynucleotides encoding novel Helicobacter polypeptides in the Helicobacter genome.
<3>Japanese Patent Office
<4>JP 2001527393 A
<5>
<6>2001
<7>
<>

<1>Klee, S.R., Nassif, X., Kusecek, B., Merker, P., Beretti, J.L., Achtman, M., Tinsley, C.R.
<2>Molecular and biological analysis of eight genetic islands that distinguish Neisseria meningitidis from the closely related pathogen   Neisseria gonorrhoeae.
<3>Infect. Immun.
<4>68
<5>2082-2095
<6>2000
<7>The pathogenic species Neisseria meningitidis and Neisseria gonorrhoeae cause dramatically
different diseases despite strong relatedness at the
genetic and biochemical levels. N. meningitidis can cross the blood-brain
barrier to cause meningitis and has a propensity for toxic septicemia
unlike N. gonorrhoeae. We previously used subtractive hybridization to
identify DNA sequences which might encode functions specific to bacteremia
and invasion of the meninges because they are specific to N. meningitidis
and absent from N. gonorrhoeae. In this report we show that these
sequences mark eight genetic islands that range in size from 1.8 to 40 kb
and whose chromosomal location is constant. Five of these genetic islands
were conserved within a representative set of strains and/or carried genes
with homologies to known virulence factors in other species. These were
deleted, and the mutants were tested for correlates of virulence in vitro
and in vivo. This strategy identified one island, region 8, which is
needed to induce bacteremia in an infant rat model of meningococcal
infection. Region 8 encodes a putative siderophore receptor and a
disulfide oxidoreductase. None of the deleted mutants was modified in its
resistance to the bactericidal effect of serum. Neither were the mutant
strains altered in their ability to interact with endothelial cells,
suggesting that such interactions are not encoded by large genetic islands
in N. meningitidis.

<>

<1>Kleerebezem, M. et al.
<2>Complete genome sequence of Lactobacillus plantarum WCFS1.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>1990-1995
<6>2003
<7>The 3,308,274-bp sequence of the chromosome of Lactobacillus plantarum strain WCFS1, a single
colony isolate of strain NCIMB8826 that was
originally isolated from human saliva, has been determined, and contains
3,052 predicted protein-encoding genes. Putative biological functions
could be assigned to 2,120 (70%) of the predicted proteins. Consistent
with the classification of L. plantarum as a facultative
heterofermentative lactic acid bacterium, the genome encodes all enzymes
required for the glycolysis and phosphoketolase pathways, all of which
appear to belong to the class of potentially highly expressed genes in
this organism, as was evident from the codon-adaptation index of
individual genes. Moreover, L. plantarum encodes a large
pyruvate-dissipating potential, leading to various end-products of
fermentation. L. plantarum is a species that is encountered in many
different environmental niches, and this flexible and adaptive behavior is
reflected by the relatively large number of regulatory and transport
functions, including 25 complete PTS sugar transport systems. Moreover,
the chromosome encodes >200 extracellular proteins, many of which are
predicted to be bound to the cell envelope. A large proportion of the
genes encoding sugar transport and utilization, as well as genes encoding
extracellular functions, appear to be clustered in a 600-kb region near
the origin of replication. Many of these genes display deviation of
nucleotide composition, consistent with a foreign origin. These findings
suggest that these genes, which provide an important part of the
interaction of L. plantarum with its environment, form a lifestyle
adaptation region in the chromosome.

<>

<1>Kleid, D., Humayun, Z., Jeffrey, A., Ptashne, M.
<2>Novel properties of a restriction endonuclease isolated from Haemophilus parahaemolyticus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>73
<5>293-297
<6>1976
<7>The sequences in lambda DNA in and around six sites cut by HphI, a restriction
enzyme isolated from Haemophilus parahaemolyticus, are compared.  The enzyme
produces a staggered cut around an AT or TA base pair, but the sequences
immediately surrounding the cleavage sites bear no obvious relation to one
another.  Eight (in some cases nine) base pairs to one side of each cleavage
site is the common sequence TCACCAGTGC.  Two lines of evidence indicate that
these bases constitute part or all of the Hph recognition site.  First,
mutations in this sequence prevent HphI cutting.  Second,
dimethylsulfate-mediated methylation of Gs and As in this site prevent cutting,
whereas methylation of purines in the region between this sequence and the
cleavage sites has no such effect.  There is discernible 2-fold rotational
symmetry neither in the common sequence nor around the cleavage sites.

<>

<1>Kleid, D.G.
<2>Purification and properties of the HphI endonuclease.
<3>Methods Enzymol.
<4>65
<5>163-166
<6>1980
<7>5' CGGTGATACTGAGCA^CATCAG   3' GCCACTATGACTCG^TGTAGTC The nucleotide
sequence-specific endonuclease from Haemophilus parahaemolyticus has properties
that differ from the majority of type II restriction endonuclease.  This enzyme
recognizes an unsymmetric nucleotide sequence and cleaves the DNA at a site
approximately one turn of the helix from the center of the recognition
sequence.  The recognition sequence has been demonstrated to be a five base
pair sequence 5' GGTGA   3' CCACT.

<>

<1>Klein, A.
<2>Mechanisms of host-controlled modification of phage T1.
<3>Z. Vererbungsl.
<4>96
<5>346-363
<6>1965
<7>Complementation tests using T1 amber-mutants show that all genes of restricted
phage so far investigated are able to function in the restricting host.  This
ability is quickly lost when m-RNA synthesis is blocked.  The ability of
functioning of restricted phage genes depends on their location on the phage
genome.  A model is discussed explaining this polarization in terms of directed
transcription of T1 DNA.  As in host cells lysogenic for P1 restricted T1 DNA
is destroyed after infection of strains THU or BG43, which also act on T1 by
HCM.  T1 phages which do not bear P1 specific modification may in rare cases
escape from restriction in host cells lysogenic for P1 - either by infecting a
nonrestricting exceptional cell or when they are very quickly protected against
destruction, probably by rapid modification.  Multiple infection does not
increase this low probability of survival of the single phage.  T1 DNA contains
5-methylcytosine and 6-methylaminopurine.  The amount of these bases in the DNA
is independent of the host specificity controlled by P1.  However, coinfection
of P1 lysogenic cells with T1 and T3 simultaneously prevents T1 DNA from being
methylated and modified.  It may therefore be concluded that the mechanisms of
modifying T1 in this host strain consists in methylating T1 DNA in a specific
pattern.  T2gt, a mutant of T2 containing nonglucosylated DNA, is restricted by
cells lysogenic for P1 while it cannot be modified by the same host.  The
question is discussed, whether this result has some implication concerning the
role of 5 MC in the postulated HCM pattern since 5 MC cannot be formed in T2gt
DNA.

<>

<1>Klein, A.
<2>Host-controlled modification.
<3>Z. Vererbungsl.
<4>96
<5>324-345
<6>1965
<7>The phenomena and effects of host-controlled modification (HCM) as described in
the literature are comprehensively discussed.  The review includes the
occurrence of HCM, the restriction and modification caused by CM, its influence
on bacterial and phage crosses, and the function of restricted genes.  Besides,
genetic control of HCM and mutation effects of HCM and UV-irradiation are
reviewed.

<>

<1>Klein, A., Sauerbier, W.
<2>Role of methylation in host controlled modification of phage T1.
<3>Biochem. Biophys. Res. Commun.
<4>18
<5>440-445
<6>1965
<7>Non-mutational changes upon growth on certain bacterial host strains concerning
host range properties have been found in many bacteriophages.  This phenomenon
is known as host controlled modification (HCM).  Phage particles thus carry a
host specificity determined by the bacterial strain on which they were
produced.  Upon infection of a different bacterial strain the phage may be
either accepted or rejected on the basis of its specificity.  If accepted, it
multiplies in the new host cell and acquires a new specificity.  Arber and
Dussoix (1962) and Dussoix and Arber (1962) showed that host specificity of
phage lambda is carried by the phage DNA.  The chemical basis of HCM in lambda
is unknown.  It was suggested by in vitro experiments of Gold and Hurwitz
(1963) that different methylation of lambda DNA by different host cells might
confer different host specificity to the phage.  Ledinko (1964), however, found
equal amounts of 5-methylcytosine (5-MC) in DNA extracted from phage lambda
carrying the host specificities derived from the strains Escherichia coli
C,B,K, or K(P1).  Evidence will be presented here that (1) T1-DNA may in some
cases be methylated to different extents depending on its host specificity and
(2) that methylation is the chemical basis or at least an absolute requirement
for HCM of phage T1 by bacterial host cells lysogenic for P1.  In addition a
new system of HCM acting on T1 will be described which does not overlap with
the P1 system.

<>

<1>Klein, E.A., Gitai, Z.
<2>Draft Genome Sequence of Uropathogenic Escherichia coli Strain J96.
<3>Genome Announcements
<4>1
<5>e00245-12
<6>2013
<7>J96 (O4:K6) was isolated from a human pyelonephritis patient. Here, we report the draft genome
sequence of J96, which contains virulence genes, including adhesion
factors, alpha-hemolysins, and cytotoxic necrotizing factor. J96 infects the
kidney and bladder, making it an important tool for studying pathogenesis.

<>

<1>Klein, R., Baranyi, U., Rossler, N., Greineder, B., Scholz, H., Witte, A.
<2>Natrialba magadii virus phiCh1: first complete nucleotide sequence and functional organization of a virus infecting a haloalkaliphilic archaeon.
<3>Mol. Microbiol.
<4>45
<5>851-863
<6>2002
<7>The double-stranded (ds)DNA virus phiCh1 infects the haloalkaliphilic
archaeon Natrialba magadii. The complete DNA sequence of 58 498 bp of the
temperate virus was established, and the probable functions of 21 of 98
phiCh1-encoded open reading frames (ORFs) have been assigned. This
knowledge has been used to propose functional modules each required for
specific functions during virus development. The phiCh1 DNA is terminally
redundant and circularly permuted and therefore appears to be packaged by
the so-called headful mechanism. The presence of ORFs encoding homologues
of proteins involved in plasmid replication as well as experimental
evidence indicate a plasmid-mediated replication strategy of the virus.
Results from nanosequencing of virion components suggest covalent
cross-linking of monomers of at least one of the structural proteins
during virus maturation. A comparison of the phiCh1 genome with the partly
sequenced genome of Halobacterium salinarum virus phiH revealed a close
relationship between the two viruses, although their host organisms live
in distinct environments with respect to the different pH values required
for growth.

<>

<1>Kleinstiver, B.P., Berube-Janzen, W., Fernandes, A.D., Edgell, D.R.
<2>Divalent Metal Ion Differentially Regulates the Sequential Nicking Reactions of the GIY-YIG Homing Endonuclease I-BmoI.
<3>PLoS ONE
<4>6
<5>e23804
<6>2011
<7>Homing endonucleases are site-specific DNA endonucleases that function as mobile genetic
elements by introducing double-strand breaks or nicks
at defined locations. Of the major families of homing endonucleases,
the modular GIY-YIG endonucleases are least understood in terms of
mechanism. The GIY-YIG homing endonuclease I-BmoI generates a
double-strand break by sequential nicking reactions during which the
single active site of the GIY-YIG nuclease domain must undergo a
substantial reorganization. Here, we show that divalent metal ion plays
a significant role in regulating the two independent nicking reactions
by I-BmoI. Rate constant determination for each nicking reaction
revealed that limiting divalent metal ion has a greater impact on the
second strand than the first strand nicking reaction. We also show that
substrate mutations within the I-BmoI cleavage site can modulate the
first strand nicking reaction over a 314-fold range. Additionally,
in-gel DNA footprinting with mutant substrates and modeling of an
I-BmoI-substrate complex suggest that amino acid contacts to a critical
GC-2 base pair are required to induce a bottom-strand distortion that
likely directs conformational changes for reaction progress.
Collectively, our data implies mechanistic roles for divalent metal ion
and substrate bases, suggesting that divalent metal ion facilitates the
re-positioning of the GIY-YIG nuclease domain between sequential
nicking reactions.

<>

<1>Kleinstiver, B.P., Fernandes, A.D., Gloor, G.B., Edgell, D.R.
<2>A unified genetic, computational and experimental framework identifies functionally relevant residues of the homing endonuclease I-BmoI.
<3>Nucleic Acids Res.
<4>38
<5>2411-2427
<6>2010
<7>Insight into protein structure and function is best obtained through a synthesis of
experimental, structural and bioinformatic data. Here, we
outline a framework that we call MUSE (mutual information, unigenic
evolution and structure-guided elucidation), which facilitated the
identification of previously unknown residues that are relevant for
function of the GIY-YIG homing endonuclease I-BmoI. Our approach
synthesizes three types of data: mutual information analyses that identify
co-evolving residues within the GIY-YIG catalytic domain; a unigenic
evolution strategy that identifies hyper- and hypo-mutable residues of
I-BmoI; and interpretation of the unigenic and co-evolution data using a
homology model. In particular, we identify novel positions within the
GIY-YIG domain as functionally important. Proof-of-principle experiments
implicate the non-conserved I71 as functionally relevant, with an I71N
mutant accumulating a nicked cleavage intermediate. Moreover, many
additional positions within the catalytic, linker and C-terminal domains
of I-BmoI were implicated as important for function. Our results represent
a platform on which to pursue future studies of I-BmoI and other
GIY-YIG-containing proteins, and demonstrate that MUSE can successfully
identify novel functionally critical residues that would be ignored in a
traditional structure-function analysis within an extensively studied
small domain of approximately 90 amino acids.

<>

<1>Kleinstiver, B.P., Wolfs, J.M., Edgell, D.R.
<2>The monomeric GIY-YIG homing endonuclease I-BmoI uses a molecular anchor and a flexible tether to sequentially nick DNA.
<3>Nucleic Acids Res.
<4>41
<5>5413-5427
<6>2013
<7>The GIY-YIG nuclease domain is found within protein scaffolds that participate in diverse
cellular pathways and contains a single active site that hydrolyzes DNA
by a one-metal ion mechanism. GIY-YIG homing endonucleases (GIY-HEs) are
two-domain proteins with N-terminal GIY-YIG nuclease domains connected to
C-terminal DNA-binding and they are thought to function as monomers. Using I-BmoI
as a model GIY-HE, we test mechanisms by which the single active site is used to
generate a double-strand break. We show that I-BmoI is partially disordered in
the absence of substrate, and that the GIY-YIG domain alone has weak affinity for
DNA. Significantly, we show that I-BmoI functions as a monomer at all steps of
the reaction pathway and does not transiently dimerize or use sequential
transesterification reactions to cleave substrate. Our results are consistent
with the I-BmoI DNA-binding domain acting as a molecular anchor to tether the
GIY-YIG domain to substrate, permitting rotation of the GIY-YIG domain to
sequentially nick each DNA strand. These data highlight the mechanistic
differences between monomeric GIY-HEs and dimeric or tetrameric GIY-YIG
restriction enzymes, and they have implications for the use of the GIY-YIG domain
in genome-editing applications.

<>

<1>Kleinstiver, B.P., Wolfs, J.M., Kolaczyk, T., Roberts, A.K., Hu, S.X., Edgell, D.R.
<2>Monomeric site-specific nucleases for genome editing.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>109
<5>8061-8066
<6>2012
<7>Targeted manipulation of complex genomes often requires the introduction of a double-strand
break at defined locations by
site-specific DNA endonucleases. Here, we describe a monomeric nuclease
domain derived from GIY-YIG homing endonucleases for genome-editing
applications. Fusion of the GIY-YIG nuclease domain to three-member
zinc-finger DNA binding domains generated chimeric GIY-zinc finger
endonucleases (GIY-ZFEs). Significantly, the I-TevI-derived fusions
(Tev-ZFEs) function in vitro as monomers to introduce a double-strand
break, and discriminate in vitro and in bacterial and yeast assays
against substrates lacking a preferred 5'-CNNNG-3' cleavage motif. The
Tev-ZFEs function to induce recombination in a yeast-based assay with
activity on par with a homodimeric Zif268 zinc-finger nuclease. We also
fused the I-TevI nuclease domain to a catalytically inactive LADGLIDADG
homing endonuclease (LHE) scaffold. The monomeric Tev-LHEs are active
in vivo and similarly discriminate against substrates lacking the
5'-CNNNG-3' motif. The monomeric Tev-ZFEs and Tev-LHEs are distinct
from the FokI-derived zinc-finger nuclease and TAL effector nuclease
platforms as the GIY-YIG domain alleviates the requirement to design
two nuclease fusions to target a given sequence, highlighting the
diversity of nuclease domains with distinctive biochemical properties
suitable for genome-editing applications.

<>

<1>Kleiveland, C.R., Hult, L.T., Kuczkowska, K., Jacobsen, M., Lea, T., Pope, P.B.
<2>Draft Genome Sequence of the Methane-Oxidizing Bacterium Methylococcus capsulatus (Texas).
<3>J. Bacteriol.
<4>194
<5>6626
<6>2012
<7>Methanotrophic bacteria perform major roles in global carbon cycles via their unique enzymatic
activities that enable the oxidation of one-carbon compounds,
most notably methane. Here we describe the annotated draft genome sequence of the
aerobic methanotroph Methylococcus capsulatus (Texas), a type strain originally
isolated from sewer sludge.

<>

<1>Klenk, H.-P. et al.
<2>The complete genome sequence of the hyperthermophilic, sulphate-reducing archaeon Archaeoglobus fulgidus.
<3>Nature
<4>390
<5>364-370
<6>1997
<7>Archaeoglobus fulgidus is the first sulphur-metabolizing organism to have its genome sequence
determined.  Its genome of 2,178,400 base pairs contains 2,436 open reading frames.  The
information processing systems and the biosynthetic pathways for essential components
(nucleotides, amino acids and cofactors) have extensive correlation with their counterparts in
the archaeon Methanococcus jannaschii.  The genomes of these two Archaea indicate dramatic
differences in the way these organisms sense their environment, perform regulatory and
transport functions, and gain energy.  In contrast to M. jannaschii, A. fulgidus has fewer
restriction-modification systems, and none of its genes appears to contain inteins.  A quarter
(651 ORFs) of the A. fulgidus genome encodes functionally uncharacterized yet conserved
proteins, two-thirds of which are shared with M. jannaschii (428 ORFs).  Another quarter of
the genome encodes new proteins indicating substantial archaeal gene diversity.

<>

<1>Klenk, H.P. et al.
<2>Complete genome sequence of the thermophilic, hydrogen-oxidizing Bacillus tusciae type strain (T2) and reclassification in the new genus, Kyrpidia gen. nov. as  Kyrpidia tusciae comb. nov. and emendation of the family Alicyclobacillaceae da  Costa and Ra.
<3>Standards in Genomic Sciences
<4>5
<5>121-134
<6>2011
<7>Bacillus tusciae Bonjour END  DSM 2912 = NBRC 15312)
and propose its reclassification as the type strain of a new species, Kyrpidia
tusciae, and the type species of the new genus Kyrpidia, which is a sister-group
of Alicyclobacillus. The family Alicyclobacillaceae da Costa and Rainey, 2010 is
emended. The 3,384,766 bp genome with its 3,323 protein-coding and 78 RNA genes
is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Klenk, H.P., Held, B., Lucas, S., Lapidus, A., Copeland, A., Hammon, N., Pitluck, S., Goodwin, L.A., Han, C., Tapia, R., Brambilla, E.M., Potter, G., Land, M., Ivanova, N., Rohde, M., Goker, M., Detter, J.C., Kyrpides, N.C., Woyke, T.
<2>Genome sequence of the soil bacterium Saccharomonospora azurea type strain (NA-128(T)).
<3>Standards in Genomic Sciences
<4>6
<5>220-229
<6>2012
<7>Saccharomonospora azurea Runmao et al. 1987 is a member of the genus Saccharomonospora, which
is in the family Pseudonocardiaceae and thus far poorly
characterized genomically. Members of the genus Saccharomonospora are of interest
because they originate from diverse habitats, such as leaf litter, manure,
compost, the surface of peat, and moist and over-heated grain, and may play a
role in the primary degradation of plant material by attacking hemicellulose.
Next to S. viridis, S. azurea is only the second member in the genus
Saccharomonospora for which a completely sequenced type strain genome will be
published. Here we describe the features of this organism, together with the
complete genome sequence with project status 'Improved high quality draft', and
the annotation. The 4,763,832 bp long chromosome with its 4,472 protein-coding
and 58 RNA genes was sequenced as part of the DOE funded Community Sequencing
Program (CSP) 2010 at the Joint Genome Institute (JGI).

<>

<1>Klenk, H.P., Lu, M., Lucas, S., Lapidus, A., Copeland, A., Pitluck, S., Goodwin, L.A., Han, C., Tapia, R., Brambilla, E.M., Potter, G., Land, M., Ivanova, N., Rohde, M., Goker, M., Detter, J.C., Li, W.J., Kyrpides, N.C., Woyke, T.
<2>Genome sequence of the ocean sediment bacterium Saccharomonospora marina type strain (XMU15(T)).
<3>Standards in Genomic Sciences
<4>6
<5>265-275
<6>2012
<7>Saccharomonospora marina Liu et al. 2010 is a member of the genus Saccharomonospora, in the
family Pseudonocardiaceae that is poorly characterized
at the genome level thus far. Members of the genus Saccharomonospora are of
interest because they originate from diverse habitats, such as leaf litter,
manure, compost, surface of peat, moist, over-heated grain, and ocean sediment,
where they might play a role in the primary degradation of plant material by
attacking hemicellulose. Organisms belonging to the genus are usually
Gram-positive staining, non-acid fast, and classify among the actinomycetes. Here
we describe the features of this organism, together with the complete genome
sequence (permanent draft status), and annotation. The 5,965,593 bp long
chromosome with its 5,727 protein-coding and 57 RNA genes was sequenced as part
of the DOE funded Community Sequencing Program (CSP) 2010 at the Joint Genome
Institute (JGI).

<>

<1>Klima, C.L., Cook, S.R., Hahn, K.R., Amoako, K.K., Alexander, T.W., Hendrick, S., McAllister, T.A.
<2>Draft Genome Sequence of a Mannheimia haemolytica Serotype 6 Isolate Collected from the Nasopharynx of a Beef Calf with Bovine Respiratory Disease.
<3>Genome Announcements
<4>1
<5>e0005113
<6>2013
<7>The draft genome of a Mannheimia haemolytica serotype 6 isolate obtained from the nasopharynx
of a feedlot calf with bovine respiratory disease is described.

<>

<1>Klimasauskas, S., Butkus, V., Janulaitis, A.
<2>Investigation of specificity of DNA-methylases BcnI, CfrI and Cfr10I.
<3>Mol. Biol. (Mosk)
<4>21
<5>87-92
<6>1987
<7>The site specificity of three DNA methylases BcnI, CfrI and Cfr10I was
determined to be 5'Cm4C(C/G)GG, 5'PyGGm5CCPu and 5'Pum5CCGGPy, respectively.
Using the modification methylases under investigation with known restriction
endonucleases, fourteen new DNA cleavage specificities can be created.  Some
aspects of the use of restriction endonucleases in DNA methylation analysis are
discussed.

<>

<1>Klimasauskas, S., Kumar, S., Roberts, R.J., Cheng, X.
<2>Hhal methyltransferase flips its target base out of the DNA helix.
<3>Cell
<4>76
<5>357-369
<6>1994
<7>The crystal structure has been determined at 2.8 A resolution for a chemically-trapped
covalent reaction intermediate between the Hhal DNA cytosine-5-methyltransferase,
S-adenosyl-L-homocysteine, and a duplex 13-mer DNA oligonucleotide containing methylated
5-fluorocytosine at its target. The DNA is located in a cleft between the two domains of the
protein and has the characteristic conformation of B-form DNA, except for a disrupted G-C base
pair that contains the target cytosine. The cytosine residue has swung completely out of the
DNA helix and is positioned in the active site, which itself has undergone a large
conformational change. The DNA is contacted from both the major and the minor grooves, but
almost all base-specific interactions between the enzyme and the recognition bases occur in
the major groove, through two glycine-rich loops from the small domain. The structure suggests
how the active nucleophile reaches its target, directly supports the proposed mechanism for
cytosine-5 DNA methylation, and illustrates a novel mode of sequence-specific DNA regonition.

<>

<1>Klimasauskas, S., Liutkeviciute, Z., Kriukiene, E.
<2>Conversion of alpha-hydroxyalkylated residues in biomolecules using methyltransferases.
<3>International Patent Office
<4>WO 2010115847 A
<5>
<6>2010
<7>The present invention relates to targeted conversion of alpha-hydroxyalkylated residues in
biomolecules in the presence of a directing methyltransferase, namely to targeted removal of
the alpha-hydroxyalkyl moieties to give unmodified residues, or targeted derivatization of the
alpha-hydroxyalkyl groups by covalent coupling of non-cofactor compounds represented by
formula HQ-LX1 wherein X represents a functional group or a reporter group attached via a
linker moiety L, and QH is selected from HS-, HSe, HO-H2N-, HN3 or HCN in the presence of a
directing methyltransferase.  Further development of the method of targeted conversion
comprises methods for targeted labeling a biomolecule and method for detecting
hydroxymethylated target sites in a biomolecule according to the present invention.

<>

<1>Klimasauskas, S., Liutkeviciute, Z., Kriukiene, E.
<2>Derivatization of biomolecules by covalent coupling of non-cofactor compounds using methyltransferases.
<3>International Patent Office
<4>WO 2010115846 A
<5>
<6>2010
<7>The present invention relates to a use of non-cofactor compounds, represented by formulas (I)
or (II) wherein R and Z are independently selected from H, D, C1-C12-alkyl, preferably
C1-C4-alkyl, alkenyl, alkinyl, phenyl or -LX, wherein X represents a functional group or a
reporter group attached via a linker group L, and QH is selected from -SH, -SeH, -NHNH2 or
-ONH2, for a targeted modification or derivatization of a biomolecule by covalent coupling to
the biomolecule in the presence of a directing methyltransferase.  Further development of the
method of targeted modification and derivatization are the method for targeted labeling a
biomolecule and method for detecting unmethylated target sites in a biomolecule comprising
modification of the biomolecule according to the present invention.

<>

<1>Klimasauskas, S., Nelson, J.L., Roberts, R.J.
<2>The sequence specificity domain of cytosine-C5 methylases.
<3>Nucleic Acids Res.
<4>19
<5>6183-6190
<6>1991
<7>Prokaryotic DNA[cytosine-C5] methyltransferases (m5C-methylases) share a common
architectural arrangement of ten conserved sequence motifs.  A series of eleven
hybrids have been constructed between the HpaII (recognition sequence: Cm5CGG)
and HhaI (recognition sequence: Gm5CGC) DNA-methylases.  The hybrids were
over-expressed in E. coli and their in vivo methylation phenotypes
investigated.  Six were inactive by our assay while five of them retained
partial methylation activity and full specificity.  In all five cases the
specificity matched that of the parent methylase which contributed the
so-called variable region, located between conserved motifs VIII and IX.  This
was the only sequence held in common between the active hybrids and for the
first time provides unequivocal evidence that the specificity determinants of
the mono-specific m5C-methylases are located within the variable region.
Correlation of the hybrid methylase structure with the efficiency of
methylation suggests that conserved motif IX may interact with the variable
region whereas motif X most probably interacts with the N-terminal half of the
molecule.

<>

<1>Klimasauskas, S., Roberts, R.J.
<2>M.HhaI binds tightly to substrates containing mismatches at the target base.
<3>Nucleic Acids Res.
<4>23
<5>1388-1395
<6>1995
<7>The (cytosine-5) DNA methyltransferase M.HhaI causes its target cytosine base to be flipped
completely out of the DAN helix upon binding. We have investigated the effects of replacing
the target cytosine by other, mismatched bases, including adenine, guanine, thymine and
uracil. We find that M.HhaI binds more tightly to such mismatched substrates and can even
transfer a methyl group to uracil if a G:U mismatch is present. Other mismatched substrates in
which the orphan guanine is changed exhibit similar behavior. Overall, the affinity of DNA
binding correlates inversely with the stability of the target base pair, while the nature of
the target base appears irrelevant for complex formation. The presence of a cofactor analog,
S-adenosyl-L-homocysteine, greatly enhances the selectivity of the methyltransferase for
cytosine at the target site. We propose that the DNA methyltransferases have evolved from
mismatch binding proteins and that base flipping was, and still is, a key element in many
DNA-enzyme interactions.

<>

<1>Klimasauskas, S., Roberts, R.J.
<2>Disruption of the target G-C base-pair by the HhaI methyltransferase.
<3>Gene
<4>157
<5>163-164
<6>1995
<7>HhaI MTase binds DNA duplexes containing mismatches at the target base position with higher
affinity than that observed for the canonical substrate.  The stability of these MTase-DNA
complexes inversely correlates with the strength of the base pair that is disrupted upon
interaction.  This finding may offer a general tool to detect other enzymes that flip bases
out of the DNA helix.

<>

<1>Klimasauskas, S., Steponaviciene, D., Maneliene, Z., Petrusyte, M., Butkus, V., Janulaitis, A.
<2>M.SmaI is an N4-methylcytosine specific DNA-methylase.
<3>Nucleic Acids Res.
<4>18
<5>6607-6609
<6>1990
<7>An enzymatic activity rendering DNA immune to the action of the SmaI restriction endonuclease
in the presence of S-adenosyl-L-methionine has been detected in Serratia marcescens Sb. This
methylase, M.SmaI, modifies the second cytosine residue of the substrate sequence CCCGGG
yielding N4-methylcytosine.

<>

<1>Klimasauskas, S., Szyperski, T., Serva, S., Wuthrich, K.
<2>Dynamic modes of the flipped-out cytosine during HhaI methyltransferase-DNA interactions in solution.
<3>EMBO J.
<4>17
<5>317-324
<6>1998
<7>Flipping of a nucleotide out of a B-DNA helix into the active site of an enzyme has been
observed for the HhaI and HaeIII cytosine-5 methyltransferases (M.HhaI and M.HaeIII) and for
numerous DNA repair enzymes.  Here we studied the base flipping motions in the binary
M.HhaI-DNA and the ternary M.HhaI-DNA-cofactor systems in solution.  Two 5-fluorocytosines
were introduced into the DNA in the places of the target cytosine and, as an internal control,
a cytosine positioned two nucleotides upstream of the recognition sequence 5'-GCGC-3'.  The
19F NMR spectra combined with gel mobility data show that interaction with the enzyme induces
partition of the target base among three states, i.e. stacked in the B-DNA, an ensemble of
flipped-out forms and the flipped-out form locked in the enzyme active site.  Addition of the
cofactor analogue S-adenosyl-L-homocysteine greatly enhances the trapping of the target
cytosine in the catalytic site.  Distinct dynamic modes of the target cytosine have thus been
identified along the reaction pathway, which includes novel base-flipping intermediates that
were not observed in previous X-ray structures.  The new data indicate that flipping of the
target base out of the DNA helix is not dependent on binding of the cytosine in the catalytic
pocket of M.HhaI, and suggest an active role in the enzyme in the opening of the DNA duplex.

<>

<1>Klimasauskas, S., Timinskas, A., Menkevicius, S., Butkiene, D., Butkus, V., Janulaitis, A.
<2>Sequence motifs characteristic of DNA[cytosine-N4]methylases:  similarity to adenine and cytosine-C5 DNA-methylases.
<3>Nucleic Acids Res.
<4>17
<5>9823-9832
<6>1989
<7>The sequences coding for DNA[cytosine-N4]methyltransferases MvaI (from
Micrococcus varians RFL19) and Cfr9I (from Citrobacter freundii RFL9) have been
determined.  The predicted methylases are proteins of 454 and 300 amino acids,
respectively.  Primary structure comparison of M.Cfr9I and another m4C-forming
methylase, M.PvuII, revealed extended regions of homology.  The sequence
comparison of the three DNA[cytosine-N4]-methylases using originally developed
software revealed two conserved patterns, DPF-GSGT and TSPPY, which were found
similar also to those of adenine and DNA[cytosine-C5]-methylases.  These data
provided a basis for global alignment and classification of DNA-methylase
sequences.  Structural considerations led us to suggest that the first region
could be the binding site of AdoMet, while the second is thought to be directly
involved in the modification of the exocyclic amino group.

<>

<1>Klimasauskas, S., Timinskas, A., Menkevicius, S., Butkiene, D., Butkus, V., Janulaitis, A.
<2>Sequence motifs characteristic of DNA-cytosine-N4-methyltransferases:  The two domains of global similarity within DNA-methylases.
<3>Eksp. Biol.
<4>0
<5>4-12
<6>1990
<7>Sequences coding for DNA [cytosine-N4] methyltransferases MvaI (from Micrococcus varians
RFL19) and Cfr91 (from Citrobacter freundii RFL9) have been determined.  The methylases
predicted are proteins of 454 and 300 amino acids, respectively.  Primary structure comparison
to other m4C-forming methylases, M. PvuII and M. SmaI, revealed two conserved patterns
DPF-GSGTTGV and TSPPY---R; these patterns were found to have analogues within all adenine and
DNA[cytosine-C5]-methylase sequences known ever since.  The presence of the two homology
domains in all known DNA-methylases provided a basis for their global alignment and
classification.  DNA-methylases can be divided into four groups (S,D,N,G) on the basis of
substrate-structural peculiarities or into two families (S,D and N,G) based on the relative
positioning of these domains within sequences.  A subset of SD family sequences differing from
the others in reversed MTase domain configuration 21 (instead of the usual 12) forms a
structurally very compact subfamily of proteins.

<>

<1>Klimasauskas, S., Weinhold, E.
<2>A new tool for biotechnology: AdoMet-dependent methyltransferases.
<3>Trends Biotechnol.
<4>25
<5>99-104
<6>2007
<7>AdoMet-dependent methyltransferases catalyze highly specific methyl group transfers from the
ubiquitous cofactor S-adenosyl-L-methionine to a multitude of biological targets in the cell.
Recently, DNA methyltransferases have been used for the sequence-specific, covalent attachment
of larger chemical groups to plasmid and bacteriophage DNA using two classes of synthetic
Ado-Met analogs.  These synthetic cofactors, in combination with the myriad AdoMet-dependent
methyltransferases available in nature, provide new molecular tools for precise, targeted
functionalization and labeling of large natural DNAs and, in all likelihood, RNAs and
proteins.  This paves the way for numerous novel applications in the functional analysis of
biological methylation, biotechnology and medical diagnostics.

<>

<1>Klimkait, T.
<2>'Restriction-PCR' - a superior replacement for restriction endonucleases in DNA cloning applications.
<3>J. Biochem. Mol. Biol.
<4>33
<5>162-165
<6>2000
<7>Polymerase chain reaction (PCR) is well established as an indispensable tool of molecular
biology; and yet a limitation for cloning
applications continues to be that products often require subsequent
restriction digests, blunt-end ligation, or the use of special linear
vectors, Here a rapid, PCR-based system is described for the simple,
restriction enzyme-free generation of synthetic, 'restriction-like' DNA
fragments with staggered ends. Any 3'- or 5'-protruding terminus, but
also non-palindromic overhangs with an unrestricted single strand
length are specifically created. With longer overhangs,
'Restriction-PCR' does not even require a ligation step prior to
transformation. Thereby the technique presents a powerful tool e.g. for
a successive, authentic reconstitution of sub-fragments of long genes
with no need to manipulate the sequence or to introduce restriction
sites. Since restriction enzyme-free and thereby devoid of the limitations
of partial DNA digests, 'Restriction-PCR' allows a straight one-step
generation and cloning of difficult DNA fragments that internally carry
additional sites for those endonucleases involved in the cloning. Small
site-specific sequence insertions or deletions can be precisely
engineered into genes of interest. With these properties
'Restriction-PCR' has the potential to add significant speed and
versatility to a wide variety of DNA cloning applications.

<>

<1>Klinzing, D.C., Matias, R.R., Skowronski, E., Alvarez, M., Liles, V., Dimamay, M.P., Natividad, F.F.
<2>Shotgun Genome Sequence of a Yersinia enterocolitica Isolate from the Philippines.
<3>J. Bacteriol.
<4>194
<5>542-543
<6>2012
<7>The first shotgun genome sequence of a microbial pathogen from the Philippines is reported.
Yersinia enterocolitica subsp. palearctica strain
PhRBD_Ye1 is the first Y. enterocolitica strain sequenced from an animal
source, swine, which is a natural source of yersiniosis. The closest
phylogenetic match is a human clinical isolate from Germany.

<>

<1>Klippel, B. et al.
<2>Complete genome sequence of the marine cellulose- and xylan-degrading bacterium Glaciecola sp. strain 4H-3-7+YE-5.
<3>J. Bacteriol.
<4>193
<5>4547-4548
<6>2011
<7>Glaciecola sp. 4H-3-7+YE-5 was isolated from subseafloor sediments at Suruga Bay in Japan and
is capable of efficiently hydrolyzing cellulose
and xylan. The complete genome sequence of Glaciecola sp. 4H-3-7+YE-5
revealed several genes encoding putatively novel glycoside hydrolases
offering a high potential for plant biomass degradation.

<>

<1>Klippel, B. et al.
<2>Complete Genome Sequences of Krokinobacter sp. strain 4H-3-7-5 and Lacinutrix sp. strain 5H-3-7-4, polysaccharide-degrading members of the family  Flavobacteriaceae.
<3>J. Bacteriol.
<4>193
<5>4545-4546
<6>2011
<7>Two members of the family Flavobacteriaceae were isolated from subseafloor sediments using
artificial seawater with cellulose, xylan and chitin as
sole carbon and energy sources. Here, we present the complete genome
sequences of Krokinobacter sp. 4H-3-7-5 and Lacinutrix sp. 5H-3-7-4 which
both encode for putatively novel enzymes involved in cellulose,
hemicellulose and chitin metabolism.

<>

<1>Klockgether, J., Reva, O., Larbig, K., Tummler, B.
<2>Sequence Analysis of the Mobile Genome Island pKLC102 of Pseudomonas aeruginosa C.
<3>J. Bacteriol.
<4>186
<5>518-534
<6>2004
<7>The Pseudomonas aeruginosa plasmid pKLC102 coexists as a plasmid and a
genome island in clone C strains. Whereas the related plasmid pKLK106
reversibly recombines with P. aeruginosa clone K chromosomes at one of the
two tRNA(Lys) genes, pKLC102 is incorporated into the tRNA(Lys) gene only
close to the pilA locus. Targeting of the other tRNA(Lys) copy in the
chromosome is blocked by a 23,395-bp mosaic of truncated PAO open reading
frames, transposons, and pKLC102 homologs. Annotation and phylogenetic
analysis of the large 103,532-bp pKLC102 sequence revealed that pKLC102 is
a hybrid of plasmid and phage origin. The plasmid lineage conferred oriV
and genes for replication, partitioning, and conjugation, including a pil
cluster encoding type IV thin sex pili and an 8,524-bp chvB glucan
synthetase gene that is known to be a major determinant for host tropism
and virulence. The phage lineage conferred integrase, att, and a syntenic
set of conserved hypothetical genes also observed in the
tRNA(Gly)-associated genome islands of P. aeruginosa clone C chromosomes.
In subgroup C isolates from patients with cystic fibrosis, pKLC102 was
irreversibly fixed into the chromosome by the insertion of the large
23,061-bp class I transposon TNCP23, which is a composite of plasmid,
integron, and IS6100 elements. Intramolecular transposition of a copy of
IS6100 led to chromosomal inversions and disruption of plasmid synteny.
The case of pKLC102 in P. aeruginosa clone C documents the intraclonal
evolution of a genome island from a mobile ancestor via a reversibly
integrated state to irreversible incorporation and dissipation in the
chromosome.

<>

<1>Klump, H.
<2>Codification and evolution of experimentally observed specific recognition sites for restriction enzymes on DNA.
<3>Biosystems
<4>21
<5>33-49
<6>1987
<7>The list of published restriction endonucleases along with their substrates
provides an excellent data base for the evaluation of the evolution and
codification of the key elements for specific recognition sites on the DNA.  In
this paper the considerations will be limited to palindromic tetramer-,
pentamer-, and hexamer-sequences.  It is basically assumed that each base pair
within these sequences has to be recognized by directionally unique bidentate
hydrogen bonds either within the plane of the base pair or by bridging the
appropriate H-bond donor/acceptor groups of the neighbouring bases of the same
strand.  Thus sequence specificity is mediated by twelve (eight) H-bonds,
originating from the protein recognition modules.  Besides a pronounced
preference for GC base pairs expressed by their high frequency in the most
abundant sequences, serving the need of maximal thermodynamic stability of the
double helical substrates, it can also be shown that the stacking of
consecutive bases within the recognition site sequences plays a major role in
shaping the particular DNA/protein interface.  Finally it will be demonstrated
that the full set of sequences discussed in this paper can readily be derived
by stepwise expanding the vocabulary of three simple tetrameric sequences by
inserting single base pairs into the centre of a minimal sequence, thus
creating all the published pentameric restriction sites, or by inserting/adding
two GC base pairs in a palindromic way, thus creating the known multiplicity of
hexameric sites.

<>

<1>Klumpp, J., Staubli, T., Schmitter, S., Hupfeld, M., Fouts, D.E., Loessner, M.J.
<2>Genome Sequences of Three Frequently Used Listeria monocytogenes and Listeria ivanovii Strains.
<3>Genome Announcements
<4>2
<5>e00404-14
<6>2014
<7>We present the complete de novo assembled genome sequences of Listeria monocytogenes strains
WSLC 1001 (ATCC 19112) and WSLC 1042 (ATCC 23074) and
Listeria ivanovii WSLC 3009, three strains frequently used for the propagation
and study of bacteriophages because they are presumed to be free of inducible
prophages.

<>

<1>Klysik, J., Stirdivant, S.M., Singleton, C.K., Zacharias, W., Wells, R.D.
<2>Effects of 5 cytosine methylation on the B-Z transition in DNA restriction fragments and recombinant plasmids.
<3>J. Mol. Biol.
<4>168
<5>51-71
<6>1983
<7>Alternating (dC-dG)n regions in DNA restriction fragments and recombinant
plasmids were methylated at the 5 position of the cytosine residues by the HhaI
methylase.  Methylation lowers the concentration of NaCl or MgCl2 necessary to
cause the B-Z conformational transition in these sequences.  Ionic strengths
higher than physiological conditions are required to form the Z conformation
when the methylated (dC-dG)n tract is contiguous with regions that do not form
Z structures, in contrast to the results with the DNA polymer
poly(m5dC-dG).poly(m5dC-dG).  In supercoiled plasmids containing (dC-dG)n
sequences, methylation reduces the number of negative supercoils necessary to
stabilize the Z conformation.  Calculations of the observed free energy
contributions of the B-Z junction and cytosine methylation suggest that two
junctions offset the favorable effect of methylation on the Z conformation in
(dC-dG)n sequences (about 29 base-pairs in length).  Studies with individual
methylated topoisomers demonstrate that increasing Na+ concentration up to
approximately 0.2M inhibits the formation of the Z conformation in the
(m5dC-dG)n region of supercoiled plasmids.  The results suggest that
methylation may serve as a triggering mechanism for Z DNA formation in
supercoiled DNAs.

<>

<1>Knapp, B., Hundt, E., Schmidt, K.H.
<2>Proteins, in particular membrane proteins, of Helicobacter pylori,  their preparation and use.
<3>Japanese Patent Office
<4>JP 2001502886 A
<5>
<6>2001
<7>
<>

<1>Kneale, G.G.
<2>A symmetrical model for the domain structure of Type I DNA methyltransferases.
<3>J. Mol. Biol.
<4>243
<5>1-5
<6>1994
<7>Type I DNA methyltransferases are complex multisubunit enzymes that methylate a specific base
in each half of an asymmetric bipartite DNA recognition sequence. The specificity (S) subunit
contains two corresponding DNA sequence recognition domains, plus a number of conserved
regions which interact with two modification (M) subunits to form a trimeric enzyme of the
form M2S. The way in which the subunits interact with DNA in a pseudo-symmetric fashion has
long been unclear. Analysis of internal sequence repeats in the S-subunit shows the occurrence
of significant homologies between the central conserved domain and sequences near the N and C
termini. On the basis of this "split repeat", a "circular" organization of the domains of this
subunit is proposed that provides the required symmetry for interacting with the M-subunits
and with the target DNA sequence. In the proposed model, one M-subunit interacts with the N-
and C-terminal conserved regions of the S-subunit, which are thereby brought into close
proximity. The second M-subunit makes equivalent contacts with repeated sequences in the
central conserved domain. The model suggests a more general scheme for the imposition of
pseudo-dyad symmetry on protein subunits that have internal repeats by making equivalent
contacts with additional subunits.

<>

<1>Knight, A.I., Lilley, R.J., Buckland, R.M., Warner, P.J.
<2>Plasmid mediated restricted and modification in Acinetobacter sp.
<3>Heredity
<4>61
<5>284-285
<6>1988
<7>Evidence will be presented for a novel plasmid mediated restriction and
modification system active on broad host range plasmids in an alkane utilising
Acinetobacter sp.  Genetic studies using the transmissable R-factor pAV1 have
demonstrated that this system is distinct from the previously described system
specified by pAV2 in Acinetobacter sp. EBF65/65.  Evidence will also be
presented indicating that pAV1 is able to escape plasmid mediated restriction
by the mobilisation of cryptic plasmids.

<>

<1>Knizewski, L., Kinch, L.N., Grishin, N.V., Rychlewski, L., Ginalski, K.
<2>Realm of PD-(D/E)XK nuclease superfamily revisited: detection of novel families with modified transitive meta profile searches.
<3>BMC Struct. Biol.
<4>7
<5>40
<6>2007
<7>BACKGROUND: PD-(D/E)XK nucleases constitute a large and highly diverse superfamily of enzymes
that display little sequence similarity despite
retaining a common core fold and a few critical active site residues. This
makes identification of new PD-(D/E)XK nuclease families a challenging
task as they usually escape detection with standard sequence-based
methods. We developed a modified transitive meta profile search approach
and to consider the structural diversity of PD-(D/E)XK nuclease fold more
thoroughly we analyzed also lower than threshold Meta-BASIC hits to select
potentially correct predictions placed among unreliable or incorrect ones.
RESULTS: Application of a modified transitive Meta-BASIC searches on
updated PFAM families and PDB structures resulted in detection of five new
PD-(D/E)XK nuclease families encompassing hundreds of so far
uncharacterized and poorly annotated proteins. These include four families
catalogued in PFAM database as domains of unknown function (DUF506,
DUF524, DUF1626 and DUF1703) and YhgA-like family of putative
transposases. Three of these families represent extremely distant homologs
(DUF506, DUF524, and YhgA-like), while two are newly defined in updated
database (DUF1626 and DUF1703). In addition, we also confidently
identified an extended AAA-ATPase domain in the N-terminal region of
DUF1703 family proteins. CONCLUSION: Obtained results suggest that
detailed analysis of below threshold Meta-BASIC hits may push limits
further for distant homology detection in the 'midnight zone' of homology.
All identified families conserve the core evolutionary fold, secondary
structure and hydrophobic patterns common to existing PD-(D/E)XK nucleases
and maintain critical active site motifs that contribute to nucleic acid
cleavage. Further experimental investigations should address the predicted
activity and clarify potential substrates providing further insight into
detailed biological role of these newly detected nucleases.

<>

<1>Knoblich, I.M., Sellmann, E., Kaluza, K., Frey, B., Auer, J., Schmitz, G.G., Westermann, P.
<2>Ssp5230I, a novel isoschizomer of AatII from Streptomyces recognizing 5'-GACGT/C-3'.
<3>Nucleic Acids Res.
<4>20
<5>2378
<6>1992
<7>We have isolated Ssp5230I, a novel class-II restriction endonuclease from Streptomyces species
recognizing the palindromic sequence 5'-GACGT/C-3' generating 3'-protruding
ACGT-tetranucleotides. With respect to its isoschizomer AatII it can be isolated in higher
purity and stability. From the mapping and sequencing data the specificity of Ssp5230I is
concluded as 5'-GACGT/C-3' 3'-C/TGCAG-5'.

<>

<1>Knowle, D., Lintner, R.E., Touma, Y.M., Blumenthal, R.M.
<2>Nature of the Promoter Activated by C.PvuII, an Unusual Regulatory Protein Conserved among Restriction-Modification Systems.
<3>J. Bacteriol.
<4>187
<5>488-497
<6>2005
<7>A widely distributed family of small regulators, called C proteins, controls a subset of
restriction-modification systems. The C proteins
studied to date activate transcription of their own genes and that of
downstream endonuclease genes; this arrangement appears to delay
endonuclease expression relative to that of the protective
methyltransferase when the genes enter a new cell. C proteins bind to
conserved sequences called C boxes. In the PvuII system, the C boxes have
been reported to extend from -23 to +3 relative to the transcription start
for the gene for the C protein, an unexpected starting position relative
to a bound activator. This study suggests that transcript initiation
within the C boxes represents initial, C-independent transcription of
pvuIICR. The major C protein-dependent transcript appears to be a
leaderless mRNA starting farther downstream, at the initiation codon for
the pvuIIC gene. This conclusion is based on nuclease S1 transcript
mapping and the effects of a series of nested deletions in the promoter
region. Furthermore, replacing the region upstream of the pvuIIC
initiation codon with a library of random oligonucleotides, followed by
selection for C-dependent transcription, yielded clones having sequences
that resemble -10 promoter hexamers. The -35 hexamer of this promoter
would lie within the C boxes. However, the spacing between C boxes/-35 and
the apparent -10 hexamer can be varied by +/-4 bp with little effect. This
suggests that, like some other activator-dependent promoters, PpvuIICR may
not require a -35 hexamer. Features of this transcription activation
system suggest explanations for its broad host range.

<>

<1>Knox, J.D., Araujo, F.D., Bigey, P., Slack, A.D., Price, G.B., Zannis-Hadjopoulos, M., Szyf, M.
<2>Inhibition of DNA methyltransferase inhibits DNA replication.
<3>J. Biol. Chem.
<4>275
<5>17986-17990
<6>2000
<7>Ectopic expression of DNA methyltransferase transforms vertebrate cells, and inhibition of DNA
methyltransferase reverses the transformed phenotype by an unknown mechanism. We tested the
hypothesis that the presence of an active DNA methyltransferase is required for DNA
replication in human non-small cell lung carcinoma A549 cells. We show that the inhibition of
DNA methyltransferase by two novel mechanisms negatively affects DNA synthesis and progression
through the cell cycle. Competitive polymerase chain reaction of newly synthesized DNA shows
decreased origin activity at three previously characterized origins of replication following
DNA methyltransferase inhibition. We suggest that the requirement of an active DNA
methyltransferase for the functioning of the replication machinery has evolved to coordinate
DNA replication and inheritance of the DNA methylation pattern.

<>

<1>Kobayashi, E., Asada, K., Mukai, H., Yamamoto, J., Tomono, J., Uemori, T., Enoki, T., Kato, I., Sagawa, H.
<2>A stabilization method and a preservation method for a reagent for nucleic acid amplification or detection reaction.
<3>Japanese Patent Office
<4>JP 2007228977 A
<5>
<6>2007
<7>
<>

<1>Kobayashi, H., Kawasaki, K., Takeishi, K., Noda, H.
<2>Symbiont of the stink bug Plautia stali synthesizes rough-type lipopolysaccharide.
<3>Microbiol. Res.
<4>167
<5>48-54
<6>2011
<7>The structures and biosynthesis of lipopolysaccharide (LPS), a major component of
the outer membrane of Gram-negative bacteria, have been studied extensively in
cultured bacteria such as Escherichia coli. In contrast, little is known about
the structures and biosynthesis of the LPS of unculturable bacteria, including
insect symbionts, many of which are Gram-negative bacteria. A brown-winged green
bug, Plautia stali, is known to harbor a single species of gamma-proteobacterium
in the posterior mid-gut caeca. To characterize the features of its LPS, we
analyzed the genome sequence of the symbiont, and identified the putative genes
involved in LPS synthesis. Genes involved in the synthesis of lipid A and the
core oligosaccharide were found in the genome, but waaL, which encodes the
O-antigen ligase, was not. Furthermore, we characterized the LPS of this symbiont
using SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and Toll-like receptor 4
(TLR4) stimulation assays. Consistent with the genomic analysis, the SDS-PAGE
analysis suggested that the symbiont had rough-type LPS, which lacked the
O-antigen. The TLR4 stimulation assay demonstrated that LPS purified from the
symbionts activated NF-kappaB-dependent reporter expression, indicating the
existence of a bioactive lipid A portion in the LPS. These results suggest that
the P. stali symbiont produces rough-type LPS.

<>

<1>Kobayashi, I.
<2>Why are there restriction enzymes and genetic recombination?
<3>Cell Struct. Funct.
<4>19
<5>464
<6>1994
<7>If self-replication of genetic information is the essential feature of life process, why has
the process of homologous recombination, which generates variation, evolved at all? The
double-strand break repair model of homologous recombination (for which we have evidence)
proposes that generation of double-strand break on otherwise intact DNA initiates homologous
recombination. Might this be too destructive as a means of generating recombinant progeny? We
obtained several experimental results that suggest a feature of homologous interaction, a
feature that enabled evolution of homologous recombination, is destruction of genetic
information.

Why are there restriction enzymes?

Double-strand break of DNA plays an important role in homologous recombination in vivo. It
is believed that restriction/modification system serves for destruction of foreign DNA such as
viral DNA. But this theory cannot explain easily presence of rare cutters and other phenomena.
We found that restriction enzyme gene and modification methylase gene on a plasmid stabilize
it. Our results support the following post-segregational killing mechanism: When a cell that
has lost the plasmid continues dividing, the modification enzymes become dilute and cannot
modify all of the thousands of recognition sites on the chromosome. The remaining restriction
enzymes will cleave the chromosome and kill the host cells. As a result, most viable cells in
a population carry the plasmid. In brief, turning off of a pair of genes, a killer and an
anti-killer, will program cell lineage for death. A restriction modification system is a
parasite or a selfish element such as a transposon. Its benfit to the host is protection
against foreign DNA. Thus it can be called a symbiont.


<>

<1>Kobayashi, I.
<2>Selfish genes for restriction-modification systems.
<3>Saibo Kogaku
<4>14
<5>1015-1023
<6>1995
<7>
<>

<1>Kobayashi, I.
<2>Selfishness and death: raison d'etre of restriction, recombination and mitochondria.
<3>Trends Genet.
<4>14
<5>368-374
<6>1998
<7>Type II restriction-modification gene complexes, such as the EcoRI system, are not easily lost
from their host cell.  The descendants of cells that lose a restriction-modification gene
complex are unable to modify a sufficient number of recognition sites in their chromosomes to
protect them from lethal attack by the remaining molecules of the restriction enzyme.  This
capacity to act as a selfish genetic element is likely to have contributed to the spread and
maintenance of restriction-modification systems.  Homologous recombination machineries of
cells and viruses appear to be well adapted to cope with these elements.  By extrapolation,
the capacity of mitochondria to kill their host eukaryotic cell might have stabilized their
initial symbiosis.

<>

<1>Kobayashi, I.
<2>Homologous recombination and sex as a strategy against selfish genes attacking the genome.
<3>Molecular Strategies in Biological Evolution, Annals of the New York Academy of Sciences, L.H. Caporale, New York
<4>
<5>354-356
<6>1999
<7>An important issue in current biology is our failure to adequately explain the forces
underlying the evolution and maintenance of sex.  Here we refer to sex broadly as the
interaction that occurs between homologous chromosomes of often different clonal original
(outcrossing), an event that leads to recombination, frequently between distantly located
genes (crossing-over).  We propose that sex is maintained to combat the existence of
ever-changing molecular parasites that attack the organism's genome.

<>

<1>Kobayashi, I.
<2>Reshaping the genome: DNA restriction-modification systems as mobile genetic elements.
<3>Saibo Kogaku
<4>18
<5>1846-1857
<6>1999
<7>
<>

<1>Kobayashi, I.
<2>Evolution and Diversity.
<3>Plasmid
<4>45
<5>162-163
<6>2001
<7>A type II restriction enzyme gene is often linked to a modification methylase gene whose role
is to protect the recognition site from breakage in the genome.  This postsegregational
killing or postdisturbance killing can be very severe.  Block to replication of a ts plasmid
carrying an EcoRII RM gene complex leads to immediate loss of viability by several orders of
magnitude.  The cell killing was observed when an RM plasmid was challenged by an incompatible
plasmid.  Similar resistance was observed when chromosomally located RM was threatened by a
homologous stretch of DNA in E. coli.  Many of the progeny carried the donor gene as well as
the recipient gene and lost the donor gene in the absence of its selection.  Each of these
unstable diploids have experienced megabase-size genome rearrangements that include
duplication of the RM locus in question and inversion.  Precise homologous recombination at
ectopic IS3 was shown to be involved in these rearrangements.  Comparisons of closely related
bacterial genomes revealed that restriction-modification gene complexes are often linked with
gross genome polymorphism.  One comparison suggests insertion of restriction modification gene
complexes into a genome with a long (ca. 100 bp) target duplication.  Comparison of two
complete genome sequences of Pyrococcus suggested transposition of restriction modification
gene complexes with linked genes.  We hypothesize that the rearrangements in the laboratory
and in the natural environments have resulted from attack of restriction enzymes on the
chromosome.  Only those genomes that have incorporated RM genes in question and allowed their
expression would survive.  Their behavior as selfish mobile elements may explain their
competition - specificity of sequence recognition and mutual exclusion (superinfection
exclusion) - and clarify certain aspects of bacterial recombination repair.  The killing is
severe in mutants defective in XerCD-mediated site-specific recombination at dif site in E.
coli chromosome.  This result provides support to the hypothesis that this site-specific
recombination system is important in DNA damage repair.  Strong postdisturbance killing by
EcoRII RM is suppressed by Dcm, an orphan methylase.  This vaccine effect may explain why Dcm
is present at all.  The selfish self-maintenance of the restriction modification genes also
provides a unique opportunity for stable maintenance and table expression of useful genes in
bacteria.

<>

<1>Kobayashi, I.
<2>Behavior of restriction-modification systems as selfish mobile elements and their impact on genome evolution.
<3>Nucleic Acids Res.
<4>29
<5>3742-3756
<6>2001
<7>Restriction-modification (RM) systems are composed of genes that encode a restriction enzyme
and a modification methylase. RM systems sometimes behave as discrete units of life, like
viruses and transposons. RM complexes attack invading DNA that has not been properly modified
and thus may serve as a tool of defense for bacterial cells. However, any threat to their
maintenance, such as a challenge by a competing genetic element (an incompatible plasmid or an
allelic homologous stretch of DNA, for example) can lead to cell death through restriction
breakage in the genome. This post-segregational or post-disturbance cell killing may provide
the RM complexes (and any DNA linked with them) with a competitive advantage. There is
evidence that they have undergone extensive horizontal transfer between genomes, as inferred
from their sequence homology, codon usage bias and GC content difference. They are often
linked with mobile genetic elements such as plasmids, viruses, transposons and integrons. The
comparison of closely related bacterial genomes also suggests that, at times, RM genes
themselves behave as mobile elements and cause genome rearrangements. Indeed some bacterial
genomes that survived post-disturbance attack by an RM gene complex in the laboratory have
experienced genome rearrangements. The avoidance of some restriction sites by bacterial
genomes may result from selection by past restriction attacks. Both bacteriophages and
bacteria also appear to use homologous recombination to cope with the selfish behavior of RM
systems. RM systems compete with each other in several ways. One is competition for
recognition sequences in post-segregational killing. Another is super-infection exclusion,
that is, the killing of the cell carrying an RM system when it is infected with another RM
system of the same regulatory specificity but of a different sequence specificity. The
capacity of RM systems to act as selfish, mobile genetic elements may underlie the structure
and function of RM enzymes.

<>

<1>Kobayashi, I.
<2>Genome comparison: involvement of restriction modification genes in genome rearrangements.
<3>Tanpakushitsu Kakusan Koso
<4>46
<5>2393-2399
<6>2001
<7>
<>

<1>Kobayashi, I.
<2>Life cycle of restriction-modification gene complexes, powers in genome evolution.
<3>Int. Congr. Ser., Elsevier Science B.V., , 
<4>1246
<5>191-200
<6>2002
<7>Increasing lines of evidence suggest that gene complexes encoding restriction modification
enzymes may behave as selfish mobile genetic elements affecting genome stability and genome
evolution.  I here compare their hypothetical life cycles with those of temperate
bacteriophage DNAs and other mobile elements.  The restriciton modification gene complexes may
establish themselves in a new host avoiding cell killing.  Upon some environmental signals,
they would be induced to multiply.  The multiplied RM gene complex may be released to the
environment and taken up by other cells in naturally competent bacteria.  In some aspects,
they could be regarded as DNA viruses without a capsid, or DNA viroids.  They may play the
role of power giving order to the community of genes called the genome.

<>

<1>Kobayashi, I.
<2>Restriction-modification systems as minimal forms of life.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>19-62
<6>2004
<7>A restriction endonuclease recognizes a specific DNA sequence and introduces a double-strand
break.  A cognate modification enzyme methylates the same sequence and thereby protects it
from cleavage.  Together, these two enzymes form a restriction-modification system.  The genes
encoding the restriction endonuclease and the cognate modification enzyme are often tightly
linked and can be termed a restriction-modification gene complex.  Restriction enzymes will
cleave incoming DNA if it has not been modified by a cognate or another appropriate
methyltransferase.  Consequently, it is widely believed that restriction-modification systems
have been maintained by bacteria because they serve to defend the cells from infection by
viral, plasmid, and other foreign DNAs (cellular defense hypothesis).

<>

<1>Kobayashi, I.
<2>DNA modification and restriction: Selfish behavior of an epigenetic system.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, Russo, V.E.A., Martienssen, R.A., Riggs, A.D., New York
<4>0
<5>155-172
<6>1996
<7>Type II restriction endonucleases are known to make double-strand breaks within or near
specific recognition sequences on duplex DNA.  Cognate modification enzymes can methylate
these sequences and protect them from cleavage.  The tight association of a cognate
restriction enzyme gene with a modification gene has been termed the (type II)
restriction-modification system.  Although the RM systems that have been described are
individually highly specific for one or a few recognition sequences, collectively the
sequences recognized are quite diverse.  Type II RM systems are ubiquitous in prokaryotes and
in archaebacteria but are absent from eukaryotes.  Type II restriction enzymes cleave foreign
DNA such as viral and plasmid DNA when this DNA has not been modified by the appropriate
modification enzyme.  In this way, cells are protected from invasion by foreign DNA.  Thus, it
has been widely believed that the evolution and maintenance of type II restriction
modification systems have been driven by the cell's need to protect itself from infection by
foreign DNA (the cellular defense hypothesis).  However, there are several unresolved issues
that cannot be explained satisfactorily by this cellular defense hypothesis.  Here, I present
experimental evidence and theoretical arguments for an alternative hypothesis (the selfish
gene hypothesis) that the evolution of at least some RM gene pairs is driven by their
"selfishness" in the genetic and evolutionary sense of the term.

<>

<1>Kobayashi, I.
<2>Restriction modification systems as selfish mobile genetic elements maintaining and rearranging the genome.
<3>Plasmid
<4>48
<5>260-261
<6>2002
<7>A restriction enzyme gene is often linked to a modification methylase gene whose role is to
protect the recognition site from breakage by the restriction enzyme.  Attempts to eliminate
some of these restriction-modification gene complexes from bacterial cells lead to cell death
through restriction breakage in the genome.  Such post-segregational cell killing was observed
when a restriction -modification gene complex was challenged by a competitor genetic element
and likely has competitive advantage.  Comparison of closely related bacterial genomes
revealed linkage of genome polymorphisms, such as insertion and inversion, with
restriction-modification gene complexes.  Restriction site avoidance in bacterial genomes and
other observations provide further support for our hypothesis that some
restriction-modification gene complexes behave as selfish mobile elements that have attacked
and shaped the genomes.  Indeed our attempts to eliminate a restriction-modification gene
complex from a cell led to various genome rearrangements in the laboratory - genome-wide
duplication and inversion events, intra-genomic movement of a restriction-modification gene
complex, and tandem amplification of a restriction-modification gene complex.  These genome
rearrangements are likely caused and selected by the restriction attacks.

<>

<1>Kobayashi, I.
<2>Genetic addiction: A principle of gene symbiosis in a genome.
<3>Plasmid Biology, ASM Press, Barbara E. Funnell and Gregory J. Phillips, Washington, DC
<4>
<5>105-144
<6>2004
<7>One of the surprising aspects of the genome that became clear during its decoding is the
fluidity of the genes.  Genomes are full of mobile symbiotic or parasitic genetic elements.
Moreover, evolutionary analyses have suggested that many genes have joined genomes relatively
recently from distantly related organisms.  This is particularly true for the bacterial and
archaeal genomes, which contain genes that even come from eukaryotes.  In addition,
comparisons of closely related genome sequences have revealed that genomes experience frequent
rearrangements during their evolution.  These observations suggest that, rather than being a
well-designed blueprint, the genome is a community of genes that essentially act selfishly and
potentially do not have the overall order of the genome as their primary interest.

<>

<1>Kobayashi, I., Nobusato, A., Kobayashi-Takahashi, N., Uchiyama, I.
<2>Shaping the genome--restriction-modification systems as mobile genetic elements.
<3>Curr. Opin. Genet. Dev.
<4>9
<5>649-656
<6>1999
<7>A restriction enzyme gene is often linked to a modification methylase gene the role of which
is to protect a recognition site on DNA from breakage by the former. Loss of some
restriction-modification gene complexes leads to cell death through restriction breakage in
the genome. Their behavior as genomic parasites/symbionts may explain the distribution of
restriction sites and clarify certain aspects of bacterial recombination repair and
mutagenesis. A comparison of bacterial genomes supports the hypothesis that
restriction-modification gene complexes are mobile elements involved in various genome
rearrangements and evolution.

<>

<1>Kobayashi, K.
<2>Stable gene maintenance by restriction modification gene complexes.
<3>Japanese Patent Office
<4>JP 2001252074 A
<5>15
<6>2001
<7>
<>

<1>Kobayashi, M., Nomura, M., Fujita, Y., Ohmomo, S., Okamoto, T.
<2>Molecular characterization of a lactococcal plasmid reducing the growth rate of host cells.
<3>Japan Agricultural Research Quarterly
<4>37
<5>53-57
<6>2003
<7>Lactococcus lactis subsp. lactis biovar. diacetylactis DRC1 carries more than 6 plasmids,
including a 7.4 kb cryptic plasmid, which was
designated as pDR1-1. The pDR1-1 plasmid was found to significantly
affect the maximum specific growth rate (mu(max)) of the host cells
because of its limiting effect on growth. To investigate the properties
of the limiting effect, the entire nucleotide sequence of pDR1-1 was
determined. It consisted of 7412 bp, and 6 open reading frames (ORFs)
were identified. The first ORF showed a high degree of similarity to a
family of replication genes (rep) that are commonly found in
lactococcal strains. The rep gene in pDR1-1 was followed by a second
ORF of unknown function. Directly downstream of the second ORF, a third
ORF was found, that showed homology to the S subunit from type I
restriction/modification systems. No significant similarity to the
contents of the database was found for the other ORFs. PCR analysis was
carried out in order to detect pDR1-1 in the other L. lactis strains.
The mu(max) of the pDR1-1-positive strain was the same as that of DRC1.
These results suggest that the load of pDR1-1 (or pDR1-1-like plasmid)
is a major factor influencing the mu(max) of DRC1 because of its
limiting effect on growth, an effect which is much more pronounced than
that produced by the overall load of other coexisting plasmids.

<>

<1>Kobayashi, M., Nomura, M., Fujita, Y., Okamoto, T., Ohmomo, S.
<2>Influence of lactococcal plasmid on the specific growth rate of host cells.
<3>Lett. Appl. Microbiol.
<4>35
<5>403-408
<6>2002
<7>Aims: To investigate a lactococcal plasmid responsible for a reduction in growth rate of its
host cell.  Methods and Results: Lactococcus lactis subsp. lactis biovar. diacetylactis DRC1
carries a high number of plasmids.  The DRC1 wild-type strain was found to grow more slowly
than a plasmid-free derivative of DRC1.  The plasmids extracted from DRC1 together with an
indicator plasmid were cotransformed into the plasmid-free strain DRC1021.  A 7.4-kb cryptic
plasmid, designated pDR1-1, was found to significantly affect the maximum specific growth rate
(umax) of the host cell.  Polymerase chain reaction (PCR) analysis was carried out in order to
detect the presence of pDR1-1 in the other L. lactis strains.  The umax of the single
pDR1-1-positive strain was determined to be the same as that of DRC1.  Significance and Impact
of the Study: These results suggest that pDR1-1 (or a pDR1-1-like plasmid) is a critical
factor in the reduction of the umax of DRC1, and that its effect on the umax is significantly
greater than that of any other coexisting plasmid.

<>

<1>Kobayashi, M., Nomura, M., Kimoto, H.
<2>Manipulation for Plasmid Elimination by Transforming Synthetic Competitors Diversifies Lactococcus lactis Starters Applicable to Food Products.
<3>Biosci. Biotechnol. Biochem.
<4>71
<5>2647-2654
<6>2007
<7>This study was designed selectively to eliminate a theta-plasmid from
Lactococcus lactis strains by transforming synthetic competitors. A
shuttle vector for Escherichia coli and L. lactis, pDB1, was constructed
by ligating a partial replicon of pDR1-1B, which is a 7.3 kb theta-plasmid
in L. lactis DRC1, with an erythromycin resistance gene into pBluescript
II KS(+). This versatile vector was used to construct competitors to
common lactococcal theta-plasmids. pDB1 contains the 5' half of the
replication origin and the 3' region of repB of pDR1-1B, but lacks the
1.1-kb region normally found between these two segments. A set of primers,
Pv3 and Pv4, was designed to amplify the 1.1-kb middle parts of the
general theta-replicons of lactococcal plasmids. When the PCR products
were cloned into the Nru I and Xho I sites of pDB1, synthetic replicons
were constructed and replication activity was restored. A number of
theta-plasmids in L. lactis ssp. lactis and cremoris were eliminated
selectively by transforming the synthetic competitors. These competitors
were easily eliminated by subculture for a short time in the absence of
selection. The resulting variants contained no exogenous DNA and are
suitable for food products, since part of the phenotype was altered
without altering other plasmids indispensable for fermentation.

<>

<1>Koblizek, M., Janouskovec, J., Obornik, M., Johnson, J.H., Ferriera, S., Falkowski, P.G.
<2>Genome Sequence of the Marine Photoheterotrophic Bacterium Erythrobacter sp. Strain NAP1.
<3>J. Bacteriol.
<4>193
<5>5881-5882
<6>2011
<7>Here we report the full genome sequence of marine phototrophic bacterium Erythrobacter sp.
strain NAP1. The 3.3-Mb genome contains a full set of
photosynthetic genes organized in one 38.9-kb cluster; however, it does
not contain genes for CO(2) or N(2) fixation, thereby confirming that the
organism is a photoheterotroph.

<>

<1>Kochanek, S., Renz, D., Doerfler, W.
<2>Probing DNA-protein interactions in vitro with the CpG DNA methyltransferase.
<3>Nucleic Acids Res.
<4>21
<5>2339-2342
<6>1993
<7>A sensitive method was devised to monitor the in vitro binding of nuclear proteins from HeLa
cells presumably to the major groove of DNA. Upon the incubation of DNA with nuclear extracts,
the complexed DNA was incubated with the CpG DNA methyltransferase from Spiroplasma species.
Subsequently, the DNA was repurified, and the location of the methylated cytidine residues was
determined by the hydrazine reaction of the DNA sequencing method. By using as DNA substrate
the VAI (virus associated) region of human adenovirus type 2 (Ad2) DNA or specific Alu
sequences associated with a number of human genes, it was documented that those segments of
DNA that were protected by bound proteins against the reaction with DNaseI also escaped in
vitro methylation by the CpG DNA methyltransferase. This new footprinting method provides a
sensitive indicator for in vitro DNA-protein interactions which are specific for the major
groove of DNA.

<>

<1>Kodaira, K.I., Oki, M., Kakikawa, M., Watanabe, N., Hirakawa, M., Yamada, K., Taketo, A.
<2>Genome structure of the Lactobacillus temperate phage Phi g1e: the whole genome sequence and the putative promoter/repressor system.
<3>Gene
<4>187
<5>45-53
<6>1997
<7>The complete genome sequence of a Lactobacillus temperate phage phi g1e was established. The
double-stranded DNA is composed of 42,259 bp, and encodes for sixty-two possible open reading
frames (ORF) as well as several potential regulatory sequences. Based on comparative analysis
with other related proteins of the Lactobacillus and Lactococcus phages as well as the
Escherichia coli phages (such as lambda), functions were putatively assigned to several phi
g1e ORFs: cng and cpg (encoding for repressors), hel (helicase), ntp (NTPase), and several
ORFs (e.g., minor capsid proteins). An about 1000-bp DNA region of phi g1e containing cpg and
cng was inferred to function as a promoter/repressor system for the phi g1e lysogenic and
lytic pathway.

<>

<1>Koechler, S., Plewniak, F., Barbe, V., Battaglia-Brunet, F., Jost, B., Joulian, C., Philipps, M., Vicaire, S., Vincent, S., Ye, T., Bertin, P.N.
<2>Genome Sequence of Halomonas sp. Strain A3H3, Isolated from Arsenic-Rich Marine Sediments.
<3>Genome Announcements
<4>1
<5>e00819-13
<6>2013
<7>We report the genome sequence of Halomonas sp. strain A3H3, a bacterium with a high tolerance
to arsenite, isolated from multicontaminated sediments of the
l'Estaque harbor in Marseille, France. The genome is composed of a 5,489,893-bp
chromosome and a 157,085-bp plasmid.

<>

<1>Koesters, R., von Knebel-Doeberitz, M.
<2>Improved blunt-end cloning by replacing EcoRV with Eco32I.
<3>Biotechniques
<4>32
<5>1244-1246
<6>2002
<7>The restriction endonuclease EcoRV, which recognizes and cleaves DNA containing 5'-GATATC-3'
sequences, is among the best characterized and most frequently used blunt-end cutting
restriction enzymes.  Its recognition site is present in many cloning vectors currently in
use.  Compared with that of cohesive ends, blunt-end ligation is generally more difficult
because a higher number of background colonies is formed that result from self-ligation of
vector  molecules.  We (and others) have found that when using EcoRV even "white color"
selection often fails so that one ends up with a large number of apparently positive (white)
colonies.  Here we demonstrate that this problem can be reduced if one uses Eco32I, an
isoschizomer of EcoRV, instead.

<>

<1>Koh, H.Y., Jung, W., Do H, L.S.G., Lee, J.H., Kim, H.J.
<2>Draft Genome Sequence of Pseudomonas pelagia CL-AP6, a Psychrotolerant Bacterium  Isolated from Culture of Antarctic Green Alga Pyramimonas gelidicola.
<3>Genome Announcements
<4>1
<5>e00699-13
<6>2013
<7>Pseudomonas pelagia CL-AP6, isolated from a culture of the Antarctic green alga Pyramimonas
gelidicola, is a psychrotolerant bacterium. Here, we report the draft
genome sequence of this strain, which may provide insights into the mutualistic
interaction between microalgae and bacteria in sea ice, as well as the cold
adaptation mechanisms of bacteria.

<>

<1>Koh, H.Y., Lee, S.G., Lee, J.H., Doyle, S., Christner, B.C., Kim, H.J.
<2>Draft Genome Sequence of Paenisporosarcina sp. Strain TG-14, a Psychrophilic Bacterium Isolated from Sediment-Laden Stratified Basal Ice from Taylor Glacier,    McMurdo Dry Valleys, Antarctica.
<3>J. Bacteriol.
<4>194
<5>6656-6657
<6>2012
<7>The psychrophilic bacterium Paenisporosarcina sp. TG-14 was isolated from sediment-laden
stratified basal ice from Taylor Glacier, McMurdo Dry Valleys,
Antarctica. Here we report the draft genome sequence of this strain, which may
provide useful information on the cold adaptation mechanism in extremely variable
environments.

<>

<1>Kohli, P., Dua, A., Sangwan, N., Oldach, P., Khurana, J.P., Lal, R.
<2>Draft Genome Sequence of Sphingobium ummariense Strain RL-3, a Hexachlorocyclohexane-Degrading Bacterium.
<3>Genome Announcements
<4>1
<5>e00956-13
<6>2013
<7>Here, we report the draft genome sequence of the hexachlorocyclohexane (HCH)-degrading
bacterium Sphingobium ummariense strain RL-3, which was isolated
from the HCH dumpsite located in Lucknow, India (27 degrees 00'N and 81 degrees
09'E). The annotated draft genome sequence (4.75 Mb) of strain RL-3 consisted of
139 contigs, 4,645 coding sequences, and 65% G+C content.

<>

<1>Kohring, G.W., Mayer, F.
<2>In situ distribution of EcoRI methylase and restriction endonuclease in cells of Escherichia coli Bs5.
<3>FEBS Lett.
<4>216
<5>207-210
<6>1987
<7>Specific IgG antibodies were raised in rabbits against purified EcoRI methylase
and restriction endonuclease.  Post embedding labeling experiments, using the
protein A-gold technique, were made with paraformaldehyde-glutaraldehyde fixed
cells, embedded in Lowicryl K4M resin at low temperatures.  Labeling with
methylase-specific antibodies showed 60-70% of gold particles in the cytoplasm
and 30-40% at the cell envelope, whereas the use of restriction enzyme-specific
antibodies led to a distribution of 10-30% in the cytoplasm and 70-90% in the
cell envelope.  The results coincide with the proposed function of the enzymes:
in the cytoplasm methylase protects the cells' own DNA from self-destruction,
and the restriction endonuclease cuts foreign DNA when entering the cell.

<>

<1>Kohring, G.W., Mayer, F.
<2>Immunoelectron microscopic localization of the restriction endonuclease EcoRI in Escherichia coli BS5.
<3>Eur. J. Cell Biol.
<4>37
<5>1-6
<6>1985
<7>Purified restriction endonuclease EcoRI isolated fom Escherichia coli BS5 was
used for the production of enzyme-specific IgG antibodies in rabbits.  For
enzyme localization experiments, paraformaldehyde-glutaraldehyde-fixed cells
were embedded and polymerized by a low-temperature procedure using Lowicryl
K4M.  the immuno electron microscopic protein A-gold technique and an
immuno-gold method revealed that 70% of the enzyme-specific labeling were
located in the cell envelope whereas 30% were found in the cytoplasm.  In
metal-shadowed preparations, no indications for the presence of the enzme could
be found on the cell surface; however, on the surface of cell protoplasts
enzyme specific labeling could be detected.  The results indicate the presence
of major amount of EcoRI in the periplasmic space of the cell where it might be
loosely bound or even freely diffusible.

<>

<1>Koike, H., Sato, S., Morita, T., Fukuoka, T., Habe, H.
<2>Draft Genome Sequence of Acetobacter tropicalis Type Strain NBRC16470, a Producer of Optically Pure d-Glyceric Acid.
<3>Genome Announcements
<4>2
<5>e01329-14
<6>2014
<7>Here we report the 3.7-Mb draft genome sequence of Acetobacter tropicalis NBRC16470(T), which
can produce optically pure d-glyceric acid (d-GA; 99%
enantiomeric excess) from raw glycerol feedstock derived from biodiesel fuel
production processes.

<>

<1>Koike, H., Yokoyama, K., Kawashima, T., Yamasaki, T., Makino, S., Clowney, L., Suzuki, M.
<2>GATC Methylation by Dam methylase in archaea: its roles and possible transcription regulation by an FFRP.
<3>Proc. Jpn. Acad., B, Phys. Biol. Sci.
<4>81
<5>278-290
<6>2005
<7>Methylation of pairs of adenines at their N6 positions in GATC sites (GATC methylation) in
archaeal genomic DNAs has been studied. Genomic DNAs in cells of three archaeal species were
found as fully methylated, while those of four other archaeal species were free of such
methylation. Consistently with this pattern of the presence or absence of GATC methylation,
homologues of E. coli Dam methylase was found present or absent, but various other types of
DNA methylases were not. A Dam homologue from Pyrococcus sp. OT3 was expressed and its
expected function of methylating GATC was confirmed. By methylation of each adenine the DNA
duplex was destabilized by 0.56 +/- 0.10 Kcal/mol, and this effect was additive. For some
archaea, transcription regulation of Dam methylase gene appears to be needed upon cell
replication, and in regions upstream of three Dam methylase genes, nucleotide sequences close
to TTTTCTTTGAAAA were present. This arrangement, five bases each at the ends, which are
complementary to each other, sandwiching three T bases at the center, fits into a pattern the
same as those recognized by dimers of transcription factors, feast/famine regulatory proteins
(FFRPs).

<>

<1>Kokcha, S., Mishra, A.K., Lagier, J.C., Million, M., Leroy, Q., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Bacillus timonensis sp. nov.
<3>Standards in Genomic Sciences
<4>6
<5>346-355
<6>2012
<7>Bacillus timonensis strain MM10403188(T) sp. nov. is the type strain of a proposed new species
within the genus Bacillus. This strain, whose genome is
described here, was isolated from the fecal flora of a healthy patient. B.
timonensis is an aerobic Gram-negative rod shaped bacterium. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. The 4,632,049 bp long genome (1 chromosome but no plasmid) contains
4,610 protein-coding and 74 RNA genes, including 5 rRNA genes.

<>

<1>Kokcha, S., Ramasamy, D., Lagier, J.C., Robert, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Brevibacterium senegalense sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>233-245
<6>2012
<7>Brevibacterium senegalense strain JC43(T) sp. nov. is the type strain of Brevibacterium
senegalense sp. nov., a new species within the Brevibacterium
genus. This strain, whose genome is described here, was isolated from the fecal
flora of a healthy Senegalese patient. B. senegalense is an aerobic rod-shaped
Gram-positive bacterium. Here we describe the features of this organism, together
with the complete genome sequence and annotation. The 3,425,960 bp long genome (1
chromosome but no plasmid) contains 3,064 protein-coding and 49 RNA genes.

<>

<1>Kolek, J., Sedlar, K., Provaznik, I., Patakova, P.
<2>Draft Genome Sequence of Clostridium pasteurianum NRRL B-598, a Potential Butanol or Hydrogen Producer.
<3>Genome Announcements
<4>2
<5>e00192-14
<6>2014
<7>We present a draft genome sequence of Clostridium pasteurianum NRRL B-598. This strain
ferments saccharides by two-stage acetone-butanol (AB) fermentation, is
oxygen tolerant, and has high hydrogen yields.

<>

<1>Kolesnikov, V.A., Zinovev, V.V., Yashina, L.N., Karginov, V.A., Baclanov, M.M., Malygin, E.G.
<2>Relaxed specificity of endonuclease BamHI as determined by identification of recognition sites in SV40 and pBR322 DNAs.
<3>FEBS Lett.
<4>132
<5>101-104
<6>1981
<7>The specificity of restriction nucleases, which is very high under conventional
conditions, may become 'relaxed' in certain media (e.g., at decreased ionic
strength, in the presence of organic solvents).  The 'condition-relaxed'
recognition sites usually correspond to shortened or degenerate sequences
derived from the 'canonic' ones.  The reasons for relaxation are unclear,
although they are of great interest for understanding the mechanism of
restriction nuclease action in general.  Up to now, the structure of 'relaxed'
sites has been determined for only 3 endonucleases: EcoRI, Bsu1, BstI.  This
communication is concerned with the determination of the structure of 'relaxed'
restriction sites for the endonuclease BamHI.  These sites in SV40 and pBR322
DNAs appeared to be GGATC and GPuATCC (cf. canonic site GGATCC).

<>

<1>Koller, K.
<2>New primers for detecting T1-like phages, useful for early detection of contamination in Escherichia coli cultures, are derived from the adenine-N6-methyl transferase gene involving vector-mediated adenine-N-methyltransferase gene transfer for expression.
<3>German Patent Office
<4>DE 19923182
<5>1-6
<6>2001
<7>Polymerase chain reaction (PCR) primers (I) for detecting T1-like phages are derived from a
304 base sequence (7), reproduced, from the
adenine-N-methyltransferase gene of the phage. Also claimed are:
detecting phages using (I); kit for detecting phages that contains (I);
and production of (I) by solid-phase synthesis. (I) are used to detect
phages that contain sequence (7) in fermentation cultures of
recombinant Escherichia coli, where the phages are lytic and cause
failure of the fermentation. Especially they are used to test
precultures, to avoid contaminating larger-scale cultures. (I) provide
unequivocal and early detection of contamination of fermentations,
allowing appropriate hygienic measures to be taken and preventing phage
contamination of downstream products of the fermentation. The test
takes only a few hours, or less. To detect the phages that contain
sequence (7), (I) are used in an essentially conventional PCR, using
phage DNA as template, then any amplicon produced is characterized.

<>

<1>Kolocheva, T.I., Demidov, S.A., Maksakova, G.A., Nevinskii, G.A.
<2>Interaction of endonuclease EcoRI with short specific and nonspecific oligonucleotides.
<3>Mol. Biol. (Mosk)
<4>32
<5>1025-1033
<6>1998
<7>
<>

<1>Kolocheva, T.I., Maksakova, G.A., Bugreev, D.D., Nevinsky, G.A.
<2>Interaction of endonuclease EcoRI with short specific and nonspecific oligonucleotides.
<3>Life
<4>51
<5>189-195
<6>2001
<7>The interaction of EcoRI with different oligodeoxyribonucleotides (ODNs) was analyzed using
the method of the slow step-by-step simplification in their complexity. Orthophosphate (KI =
31 mM), 2-deoxyribose 5-phosphate (KI = 4.6 mM) and different dNMPs (KI = 2.1-2.5 mM) were
shown to be the minimal ligands of the enzyme. The lengthening of a nonspecific d(pN)n (n =
1-6) by one nucleotide unit resulted in the increase of their affinity by a factor of
approximately 2.0. Weak nonspecific electrostatic contacts of EcoRI with internucleotide
phosphate groups of ODNs can account for about 5 orders of magnitude in the ligand affinity,
whereas the contribution of specific interactions between EcoRI and d(pN)n is no more than 2
orders of magnitude of a total ODN's affinity.

<>

<1>Kolton, M., Green, S.J., Harel, Y.M., Sela, N., Elad, Y., Cytryn, E.
<2>Draft Genome Sequence of Flavobacterium sp. Strain F52, Isolated from the Rhizosphere of Bell Pepper (Capsicum annuum L. cv. Maccabi).
<3>J. Bacteriol.
<4>194
<5>5462-5463
<6>2012
<7>Here we report the draft genome sequence of Flavobacterium sp. strain F52, isolated from the
rhizosphere of bell pepper (Capsicum annuum L. cv. Maccabi).
Flavobacterium spp. are ubiquitous in the rhizospheres of agricultural crops;
however, little is known about their physiology. To our knowledge, this is the
first published genome of a root-associated Flavobacterium strain.

<>

<1>Kolykhalov, A.A., Repin, V.E., Fish, A.M., Rechkunova, N.I., Degtyarev, S.K.
<2>Determination of substrate specificity of restriction endonuclease Vha464I.
<3>Sib. Biol. J.
<4>19
<5>60-61
<6>1991
<7>The recognition sequence and cleavage point of restriction endonuclease Vha464I have been
determined as 5'C^TTAAG.

<>

<1>Komaki, H., Ichikawa, N., Hosoyama, A., Fujita, N., Igarashi, Y.
<2>Draft Genome Sequence of Marine-Derived Actinomycete Nocardiopsis sp. Strain TP-A0876, a Producer of Polyketide Pyrones.
<3>Genome Announcements
<4>2
<5>e00665-14
<6>2014
<7>Here we report the draft genome sequence of Nocardiopsis sp. strain TP-A0876, isolated from
marine sediment, which produces polyketide-derived pyrones called
nocapyrones. The genome contains three polyketide synthase (PKS) gene clusters,
one of which was proposed to be responsible for nocapyrone biosynthesis. This
genome sequence will facilitate the study of the potential for secondary
metabolism in Nocardiopsis strains.

<>

<1>Komori, K., Fujita, N., Ichiyanagi, K., Shinagawa, H., Morikawa, K., Ishino, Y.
<2>PI-PfuI and PI-PfuII, intein-coded homing endonucleases from Pyrococcus furiosus.  I. Purification and identification of the homing-type endonuclease activities.
<3>Nucleic Acids Res.
<4>27
<5>4167-4174
<6>1999
<7>We screened for proteins with specific binding activity to Holliday junction DNA from the
hyperthermophilic archaeon Pyrococcus furiosus and found a protein that has specific affinity
for DNA with a branched structure, like a three-way or four-way junction. The protein was
identified as one of the two inteins encoded in the gene for ribonucleotide reductase (RNR) by
gene cloning. These two inteins were spliced out from the precursor protein as polypeptides
with molecular weights of 53.078 and 43.976 kDa, respectively. The amino acid sequences of
these inteins have two copies of the LAGLIDADG motif, which is found in the site-specific DNA
endonucleases. The purified proteins actually cleaved double-stranded DNA with the sequence of
the intein(-)allele, and, therefore, they were designated PI-PfuI and PI-PfuII. They generate
a 4 bp 3'-OH overhang with a 5'-phosphate, like other known homing endonucleases originating
from inteins. The optimal conditions of the DNA cleavage reaction, including temperature, pH,
and concentrations of KCl and MgCl(2), have been determined. The high affinity for junction
DNA of PI-PfuI was confirmed using the purified protein.

<>

<1>Komori, K., Ichiyanagi, K., Morikawa, K., Ishino, Y.
<2>PI-PfuI and PI-PfuII, intein-coded homing endonucleases from Pyrococcus furiosus.  II. Characterization of the binding and cleavage abilities by site-directed mutagenesis.
<3>Nucleic Acids Res.
<4>27
<5>4175-4182
<6>1999
<7>PI-PfuI and PI-PfuII from Pyrococcus furiosus are homing endonucleases, as shown in the
accompanying paper. These two endonucleases are produced by protein splicing from the
precursor protein including ribonucleotide reductase (RNR).  We show here that both enzymes
specifically interact with their substrate DNA and distort the DNA strands by 73 degrees and
67 degrees, respectively. They have two copies of the amino acid sequence motif LAGLIDADG,
which is present in the majority of homing endonucleases and provides some of the catalytic
residues necessary for DNA cleavage activity.  Site-specific mutagenesis studies showed that
two acidic residues in the motifs, Asp149 and Glu250 in PI-PfuI, and Asp156 and Asp249 in
PI-PfuII, were critical for catalysis. The third residues of the active site triads, as
predicted from the structure of PI-SceI, were Asn225 in PI-PfuI and Lys224 in PI-PfuII.
Substitution of Asn225 in PI-PfuI by Ala did not affect catalysis. The cleavage activity of
PI-PfuII was 50-fold decreased by the substitution of Ala for Lys224.  The binding affinity of
the mutant protein for the substrate DNA also decreased 6-fold. The Lys in PI-PfuII may play a
direct or indirect role in catalysis of the endonuclease activity.

<>

<1>Konarev, P.V., Kachalova, G.S., Ryazanova, A.Yu., Kubareva, E.A., Karyagina, A.S., Bartunik, H.D., Svergun, D.I.
<2>Flexibility of the Linker between the Domains of DNA Methyltransferase SsoII Revealed by Small-Angle X-Ray Scattering: Implications for Transcription Regulation in SsoII Restriction-Modification System.
<3>PLoS ONE
<4>9
<5>e93453
<6>2014
<7>Cytosine-5)-DNA methyltransferase SsoII (M. SsoII) consists of a methyltransferase domain
(residues 72-379) and an N-terminal region (residues 1-71) which regulates transcription in
SsoII restriction-modification system. Small-angle X-ray scattering (SAXS) is employed here to
study the low resolution structure of M.SsoII and its complex with DNA containing the
methylation site. The shapes reconstructed ab initio from the SAXS data reveal two distinct
protein domains of unequal size. The larger domain matches the crystallographic structure of a
homologous DNA methyltransferase HhaI (M.HhaI), and the cleft in this domain is occupied by
DNA in the model of the complex reconstructed from the SAXS data. This larger domain can thus
be identified as the methyltransferase domain whereas the other domain represents the
N-terminal region. Homology modeling of the M.SsoII structure is performed by using the model
of M.HhaI for the methyltransferase domain and representing the N-terminal region either as a
flexible chain of dummy residues or as a rigid structure of a homologous protein (phage 434
repressor) connected to the methyltransferase domain by a short flexible linker. Both models
are compatible with the SAXS data and demonstrate high mobility of the N-terminal region. The
linker flexibility might play an important role in the function of M.SsoII as a transcription
factor.

<>

<1>Koncz, C., Kiss, A., Venetianer, P.
<2>Biochemical characterization of the restriction-modification system of Bacillus sphaericus.
<3>Eur. J. Biochem.
<4>89
<5>523-529
<6>1978
<7>A type II restriction endonuclease (endo R-Bsp) has been purified from Bacillus
sphaericus to electrophoretic homogeneity.  The enzyme appears to be a single
polypeptide chain with a molecular weight of 35000.  Its pH optimum is around
8.2, it requires 20 mM Mg2+ for optimal activity and it is inhibited by Zn2+.
The yield of the enzyme is higher than that of any type I restriction
endonuclease so far reported.  The enzyme also cleaves single-stranded DNA,
albeit at a slower rate.  It seems likely that single-stranded DNA is cleaved
at the same sequences as double-stranded DNA.  Bacillus sphaericus also
contains a modification methylase (meth M-Bsp) which completely protects the
cell's own DNA against cleavage by its restriction endonuclease.  The methylase
activity has been partially purified, it copurifies with the nuclease until the
next to the last step.  The enzyme does not require ATP or Mg2+, it transfers
the methyl group of S-adenosyl-methionine to cytosine residues of DNA.  As the
action of this methylase completely protects any DNA from endo R-Bsp cleavage,
it seems likely that the methylase recognizes and methylates the same sequence
(dG-dG-dC-dC) as the nuclease.

<>

<1>Kondo, H., Tinwongger, S., Proespraiwong, P., Mavichak, R., Unajak, S., Nozaki, R., Hirono, I.
<2>Draft Genome Sequences of Six Strains of Vibrio parahaemolyticus Isolated from Early Mortality Syndrome/Acute Hepatopancreatic Necrosis Disease Shrimp in Thailand.
<3>Genome Announcements
<4>2
<5>e00221-14
<6>2014
<7>Some strains of Vibrio parahaemolyticus cause acute hepatopancreatic necrosis disease (AHPND)
in shrimp. We sequenced 3 AHPND and 3 non-AHPND strains and found that all of them lacked the
pathogenicity island relevant to human infection. A unique sequence encoding a type IV
pilus/type IV secretion system was found in 3 AHPND strains.

<>

<1>Konforti, B.
<2>The servant with the scissors.
<3>Nat. Struct. Biol.
<4>7
<5>99-100
<6>2000
<7>In 1978, Werner Arber, (Biozentrum der Universitat, Basel, Switzerland), Dan Nathans and
Hamilton Smith (both at Johns Hopkins University School of Medicine, Baltimore, Maryland, USA)
were awarded the Nobel Prize in Physiology or Medicine for the discovery of "restriction
enzymes and their application to problems of molecular genetics".  Almost immediately, the
application of these enzymes to genetics led to "new and far reaching results".  In fact, it
is hard to imagine what the biological sciences would look like today without restriction
maps, cloning and the ability to alter genes at will, to name just a few everyday tools of the
trade.  But how did this crucial discovery come about?

<>

<1>Kong, C., Wang, L., Li, P., Qu, Y., Tang, H., Wang, J., Zhou, H., Ma, Q., Zhou, J., Xu, P.
<2>Genome Sequence of Dyella ginsengisoli Strain LA-4, an Efficient Degrader of Aromatic Compounds.
<3>Genome Announcements
<4>1
<5>e00961-13
<6>2013
<7>Dyella ginsengisoli strain LA-4 can efficiently degrade environmental pollutants  such as
biphenyl and azo dyes. Here, we present a 4.55-Mb draft genome sequence
of strain LA-4, which may provide further insights into the molecular mechanism
in environmental pollution remediation.

<>

<1>Kong, H.
<2>Characterization of a new restriction-modification system, the BcgI system of Bacillus coagulans.
<3>Ph.D. Thesis, Boston University
<4>
<5>1-130
<6>1998
<7>The BcgI and other BcgI-like restriction endonucleases differ from other types of
endonucleases in that they cleave double-stranded DNA on both sides of their recognition
sequences to excise a short fragment in the presence of Mg++ and S-adenosylmethionine.  To
understand the relationship between BcgI-like enzymes and the classic types of enzymes, the
functional organization and enzymatic properties of BcgI of Bacillus coagulans were analyzed.
The BcgI restriction-modification system is a bifunctional protein complex which can cleave or
methylate DNA.  The dual cleavage/methylation activities are dependent on the methylation
state of its DNA substrate.  BcgI cleaves unmethylated DNA only; however, it methylates
hemimethylated DNA preferentially.  The 71.6-kDa A subunit of BcgI contains two conserved
m6A-methyltransferase motifs, which form a hydrophobic pocket for AdoMet.  AdoMet serves as
allosteric activator for both cleavage and methylation reactions.  By mutational analysis, the
endonuclease activity of BcgI has been assigned to the amino-terminal half of the A subunit,
and a putative endonuclease motif, PE...EXK, has also been identified.  Substitutions in the
endonuclease motif of conserved charged residues by Ala completely abolish DNA cleavage
activity of BcgI, but have no effect on DNA methylation activity, suggesting this motif is
most likely the endonuclease active site of BcgI.  While the A subunit serves as the
restriction/methylation subunit, the 39.2-kDa B subunit is likely to be the specificity
subunit.  This suggestion is based on the required participation of the B subunit in the
methyltransferase and endonuclease reactions and its structural similarity with the S subunit
of type I R-M systems.  The stoichiometry of BcgI R-M system is one S subunit, which
determines the recognition sequence of substrate DNA, and two RM subunits which cleave (or
methylate) its substrate bilaterally.  BcgI forms a hexamer of (RM)4S2 in solution.  The
hexamer contains two functional trimer units of (RM)2S1, and may be capable of interacting
with two recognition sequences.  Indeed, the BcgI enzyme cleaves DNA substrates containing two
recognition sequences much more efficiently than substrates containing a single sequence.

<>

<1>Kong, H.
<2>Analyzing the functional organization of a novel restriction modification system, the BcgI system.
<3>J. Mol. Biol.
<4>279
<5>823-832
<6>1998
<7>BcgI is a novel, multi-subunit, restriction-modification system that differs from all the
other types of R-M system in its genetic and functional organization.  The holoenzyme contains
two diferent subunits, BcgI A and BcgI B.  Both are required for endonuclease and
methyltransferase activities.  Here, we show that the endonuclease activity is mediated by the
N-terminal portion of the A subunit.  We made this assignment by mutational analysis.  The
analytic strategy involved three steps.  First, the methyltransferase activity was inactivated
by site-directed mutagenesis of a conserved methyltransferase motif also found in the A
subunit.  One of the R+M- mutants could not methylate DNA but was still able to cleave it,
therefore expression of this mutant gene was lethal to the host.  This lethal phenotype
allowed the selective isolation of cleavage-deficient mutations in a second round of random
mutagenesis in this mutant background.  The R- mutations were all located in the N-terminal
portion of the A subunit.  There are five potential endonuclease motifs within this region.
Conserved acidic residues in each of these motifs were substituted with alanine by
site-directed mutagenesis of the wild-type A gene.  The results identified one motif,
P52E53-(X)12-E66D67K68, as the probable endonuclease active-site.  Further support for this
assignment was obtained by another round of site-directed mutagenesis directed to residues
surrounding this motif.  The results showed that DNA cleavage activity was mediated by the
predicted, conserved residues, and not any of the surrounding non-conserved residues.  One
mutant protein, BcgI-E53A, with a single amino acid substitution decreased the DNA cleavage
activity at least 700-fold.  Our present model for the functional organization of BcgI locates
both endonuclease and methyltransferase domains in the A subunit, with the target recognition
domain located in the B subunit.

<>

<1>Kong, H.
<2>Method for cloning and producing the SwaI restriction endonuclease.
<3>European Patent Office
<4>EP 1048731 B
<5>
<6>2005
<7>The present invention relates to the recombinant DNA which encodes the SwaI restriction
endonuclease and modification methylase, and the production of SwaI restriction endonuclease
from the recombinant DNA.  SwaI restriction endonuclease (see U.S. Patent No. 5,158,878) is
originally isolated from Staphlococcus warneri.  It recognizes the DNA sequence 5' ATTTAAAT 3'
and cleaves the phosphodiester bond 5' to the second A of the recognition sequence to produce
a blunt end.

<>

<1>Kong, H., Besnier, C., Xu, Y.F.
<2>Engineering nicking endonucleases from type IIs restriction endonucleases.
<3>US Patent Office
<4>US 6395523
<5>
<6>2002
<7>The present invention relates to methods to engineer nicking endonucleases from existing Type
IIs restriction endonucleases, and the
production of the engineered nicking endonucleases. Two engineering
methods are disclosed. One involves inactivating the dimerization
function of a Type IIs restriction enzyme using site-directed
mutagenesis approach. The other involves replacing the cleavage domain
of a Type IIs restriction enzyme with the cleavage domain from a
natural occurring nicking endonuclease, N.BstNBI.

<>

<1>Kong, H., Chen, Z.
<2>Isolation and identification of restriction endonuclease BstFI.
<3>Nucleic Acids Res.
<4>15
<5>7205
<6>1987
<7>BstFI, an isoschizomer of HindIII, has been purified from Bacillus stearothermophilus FH58
isolated from the soil of the campus of Fudan University. Sequencing data show that the
cleavage site of BstFI is A/AGCTT, the same as HindIII. 10,000 units BstFI can be obtained
from each gram wet w. of cells. BstFI is active over a temperature range from 37C to 65C.  The
optimal temperature for its action is 55C. The optimal pH and ionic concentration of the assay
buffer for the optimum activity of BstFI is 7.0 -7.5 and 50-100 mM NaCl, respectively.  BstFI
is very stable during incubation at 45C for as long as 10 hrs., but loses its activity easily
at 50C.

<>

<1>Kong, H., Higgins, L., Dalton, M., Kucera, R., Schildkraut, I., Wilson, G.
<2>Method of making a mutated Type IIT endonuclease which has nicking activity.
<3>European Patent Office
<4>EP 1757683 A
<5>
<6>2007
<7>The invention provides a method of making a mutated Type IIT endonuclease which has nicking
activity comprising the steps of:  (1) identifying a heterodimeric Type IIT endonuclease; (b)
identifying a conserved region within said Type IIT endonuclease; (c) generating at least one
mutation within said conserved region; and (d) analyzing the mutant endonuclease of step (c)
for nicking endonuclease activity.

<>

<1>Kong, H., Higgins, L.S.
<2>Method for cloning and expression of PleI restriction endonuclease and PleI and BstNBII methylases in E. coli.
<3>US Patent Office
<4>US 6391608
<5>No Pagination
<6>2002
<7>The present invention relates to recombinant DNA which encodes the PleI restriction
endonuclease as well as PleI and BstNBII methyltransferase
and expression of PleI restriction endonuclease and M.BstNBII in E.
coli cells containing the recombinant DNA.

<>

<1>Kong, H., Higgins, L.S.
<2>Method for cloning and expression of MlyI restriction endonuclease and MlyI methylase and BstNBII methylase in E. coli.
<3>US Patent Office
<4>US 6395531
<5>No Pagination
<6>2002
<7>The present invention relates to recombinant DNA which encodes the MlyI restriction
endonuclease as well as the MlyI and BstNBII
methyltransferases and expression of MlyI restriction endonuclease and
M.BStNBII in E. coli cells containing the recombinant DNA.

<>

<1>Kong, H., Higgins, L.S., Dalton, M., Kucera, R.B., Schildkraut, I.
<2>Cloning and producing the N.BstNBI nicking endonuclease.
<3>US Patent Office
<4>US 6191267 B
<5>
<6>2001
<7>The present invention relates to recombinant DNA which encodes a novel nicking endonuclease,
N.BstNBI, and the production of N.BstNBI restriction endonuclease from the recombinant DNA
utilizing PleI modification methylase.  Related expression vectors, as well as the application
of N.BstNBI in non-thio strand displacement amplification, is disclosed also.

<>

<1>Kong, H., Higgins, L.S., Dalton, M.A.
<2>Cloning and producing the SgrAI restriction endonuclease.
<3>US Patent Office
<4>US 6048731 A
<5>18
<6>2000
<7>The present invention relates to the recombinant DNA which encodes
the SgrAI restriction endonuclease and the MspI modification methylase, and
the production of SgrAI restriction endonuclease from the recombinant DNA.

<>

<1>Kong, H., Higgins, L.S., Dalton, M.A.
<2>Cloning and producing the DraIII restriction endonuclease.
<3>US Patent Office
<4>US 6048719 A
<5>25
<6>2000
<7>The present invention relates to the recombinant DNA which encodes the DraIII restriction
endonuclease modification methylase, and the production of DraIII restriction endonuclease
from the recombinant DNA.  Related expression vectors, pHKUV5 vector which features a strong,
constitutive UV5 promoter without the Lac repressor binding site and pHKT7 vector which
contains a powerful controllable T7 promoter and a low copy number origin of replication, are
also disclosed.

<>

<1>Kong, H., Higgins, L.S., Dalton, M.A.
<2>Method for cloning and producing the SwaI restriction endonuclease.
<3>European Patent Office
<4>EP 1048731 A
<5>
<6>2000
<7>The present invention relates to the recombinant DNA which encodes the SwaI restriction
endonuclease, modification methylase, and the production of SwaI restriction endonuclease from
the recombinant DNA.  Related expression vectors, pHKUV5 which features a strong, constitutive
UV5 promoter without the Lac repressor binding site and pHKT7 which contains a powerful
controllable T7 promoter and a low copy number origin of replication, are also disclosed.

<>

<1>Kong, H., Higgins, L.S., Dalton, M.A.
<2>Method for cloning and producing the SwaI restriction endonuclease.
<3>US Patent Office
<4>US 6245545 B
<5>
<6>2001
<7>The present invention relates to the recombinant DNA which encodes the SwaI restriction
endonuclease, modification methylase, and the production of SwaI restriction endonuclease from
the recombinant DNA.  Related expression vectors, pHKUV5 which features a strong, constitutive
UV5 promoter without the Lac repressor binding site and pHKT7 which contains a powerful
controllable T7 promoter and a low copy number origin of replication, are also disclosed.

<>

<1>Kong, H., Higgins, L.S., Dalton, M.A.
<2>Method for cloning and producing the SwaI restriction endonuclease.
<3>Japanese Patent Office
<4>JP 2000316589 A
<5>
<6>2000
<7>
<>

<1>Kong, H., Higgins, L.S., Dalton, M.A., Kucera, R.B., Schildkraut, I., Wilson, G.G.
<2>Cloning and producing the N.BstNBI nicking endonuclease and related methods for using nicking endonucleases in single-stranded displacement amplification.
<3>International Patent Office
<4>WO 0194544 A
<5>
<6>2001
<7>The present invention relates to recombinant DNA which encodes a novel nicking endonuclease,
N.BstNBI, and the production of N.BstNBI restriction endonuclease from the recombinant DNA
utilizing PleI modification methylase.  Related expression vectors, as well as the application
of N.BstNBI and other nicking enzymes in non-modified strand displacement amplifications.

<>

<1>Kong, H., Lin, L.F., Porter, N., Stickel, S., Byrd, D., Posfai, J., Roberts, R.J.
<2>Functional analysis of putative restriction-modification system genes in the Helicobacter pylori J99 genome.
<3>Nucleic Acids Res.
<4>28
<5>3216-3223
<6>2000
<7>Helicobacter pylori is a gram-negative bacterium, which colonizes the gastric mucosa of
humans and is implicated in a wide range of gastroduodenal diseases. The genomic sequences of
two H. pylori strains, 26695 and J99, have been published recently. About two dozen potential
restriction-modification (R-M) systems have been annotated in both genomes, which is far above
the average number of R-M systems in other sequenced genomes. Here we describe a functional
analysis of the 16 putative Type II R-M systems in the H. pylori J99 genome. To express
potentially toxic endonuclease genes, a unique vector was constructed, which features
repression and antisense transcription as dual control elements. To determine the methylation
activities of putative DNA methyltransferases, we developed polyclonal antibodies able to
detect DNA containing N6-methyladenine or N4-methylcytosine. We found that <30% of the
potential Type II R-M systems in H. pylori J99 strain were fully functional, displaying both
endonuclease and methyltransferase activities. Helicobacter pylori may maintain a variety of
functional R-M systems, which are believed to be a primitive bacterial 'immune' system, by
alternatively turning on/off a subset of numerous R-M systems.

<>

<1>Kong, H., Morgan, R.D., Chen, Z.
<2>Identification of a new type II restriction endonuclease BsaAI.
<3>Nucleic Acids Res.
<4>18
<5>2832
<6>1990
<7>None

<>

<1>Kong, H., Morgan, R.D., Chen, Z.
<2>A new type II restriction endonuclease, BsmAI, from Bacillus stearothermophilus.
<3>Nucleic Acids Res.
<4>18
<5>686
<6>1990
<7>None

<>

<1>Kong, H., Morgan, R.D., Maunus, R.E., Schildkraut, I.
<2>A unique restriction endonuclease, BcgI, from Bacillus coagulans.
<3>Nucleic Acids Res.
<4>21
<5>987-991
<6>1993
<7>We have purified and characterized a new restriction endonuclease, BcgI, which has properties
unlike those of the three recognized classes of restriction enzymes. BcgI was isolated from
Bacillus coagulans, and it recognizes the sequence CGAN6TGC. BcgI cleaves double stranded DNA
on both strands upstream and downstream of the recognition sequence, so that the recognition
sequence is released as a 34-base pair fragment with 2-base 3'-extensions. Mg++ and
S-adenosylmethionine are required for cleavage. Sinefungin, a structural analogue of AdoMet
which generally inhibits methylase activity, can replace AdoMet in the cleavage reaction. The
apparent binding constant Kapp for AdoMet is about 100 nM, while the Kapp for sinefungin is
about 500 nM.

<>

<1>Kong, H., Roemer, S.E., Waite-Rees, P.A., Benner, J.S., Wilson, G.G., Nwankwo, D.O.
<2>Characterization of BcgI, a new kind of restriction-modification system.
<3>J. Biol. Chem.
<4>269
<5>683-690
<6>1994
<7>The BcgI restriction enzyme from Bacillus coagulans is unusual in that it cleaves on both
sides of its recognition site, CGAN6TGC, releasing a fragment that includes the site and
several bases on each side. We report the organization and nucleotide sequences of the genes
for the BcgI restriction-modification system and the properties of the proteins that they
encode. The system comprises two adjacent, similarly oriented genes. The proximal gene, bcgIA,
codes for a 637-amino acid protein (molecular mass = 71.6 kDa) that resembles certain m6
A-specific DNA-methyltransferases, particularly those that constitute the modification subunit
of type I restriction-modification systems. The distal gene bcgIB, codes for a 341-amino acid
protein (molecular mass = 39.2 kDa) that resembles none of the sequences in the sequence data
bases. The two genes overlap by several nucleotides. Alone, neither protein restricts or
modifies DNA, but, together, they form a complex in the proportion A2B that does both. DNA
binding assays showed that the DNA-protein complex can be formed only in the presence of both
subunits, suggesting that the association of inactive subunits generates the active BcgI
enzyme that can bind DNA and then either cleaves or methylates at target site.

<>

<1>Kong, H., Schildkraut, I.
<2>A novel restriction endonuclease obtainable from Bacillus coagulans and a process for producing the same.
<3>European Patent Office
<4>EP 0466332 B
<5>
<6>1994
<7>The present invention relates to a novel restriction endonuclease, BcgI, obtainable from
Bacillus coagulans, to the process for producing the same, and to related methods of employing
this novel enzyme.

<>

<1>Kong, H., Schildkraut, I.
<2>Restriction endonuclease obtainable from Bacillus coagulans and a process for producing the same.
<3>US Patent Office
<4>US 5200336
<5>
<6>1993
<7>The present invention provides a novel Type II restriction endonuclease obtainable from
Bacillus coagulans.  The endonuclease known as BcgI, recognizes the following nucleotide
sequence and has a cleavage point at both ends outside of its recognition sequence:
5'/(N)10CGA(N)6TGC(N)12/3' 3'/(N)12GCT(N)6ACG(M)10/5' to produce a 34 base pair fragment.
Also described is a process for obtaining purified BcgI from Bacillus coagulans, as well as
processes for mapping chromosomal DNA and methods for reducing background in transformants
with enzymes such as BcgI.

<>

<1>Kong, H., Smith, C.L.
<2>Substrate DNA and cofactor regulate the activities of a multi-functional restriction-modification enzyme, BcgI.
<3>Nucleic Acids Res.
<4>25
<5>3687-3692
<6>1997
<7>The BcgI restriction-modification system consists of two subunits, A and B.  It is a
bifunctional protein complex which can cleave or methylate DNA.  The regulation of these
competing activities is determined by the DNA substrates and cofactors.  BcgI is an active
endonuclease and a poor methyltransferase on unmodified DNA substrates.  In contrast, BcgI is
an active methyltransferase and an inactive endonuclease on hemimethylated DNA substrates.
The cleavage and methylation reactions share cofactors.  While BcgI requires Mg2+ and
S-adenosyl methionine (AdoMet) for DNA cleavage, its methylation reaction requires only AdoMet
and yet is significantly stimulated by Mg2+.  Site-directed mutagenesis was carried out to
investigate the relationship between AdoMet binding and BcgI DNA cleavage/methylation
activities.  Most substitutions of conserved residues forming the AdoMet binding pocket in the
A subunit abolished both methylation and cleavage activities, indicating that AdoMet binding
is an early common step required for both cleavage and methylation.  However, one mutation
(Y439A) abolished only the methylation activity, not the DNA cleavage activity.  This mutant
protein was purified and its methylation, cleavage and AdoMet binding activities were tested
in vitro.  BcgI-Y439A had no detectable methylation activity, but it retained 40% of the
AdoMet binding and DNA cleavage activities.

<>

<1>Kong, H., Smith, C.L.
<2>Does BcgI, a unique restriction endonuclease, require two recognition sites for cleavage?
<3>Biol. Chem.
<4>379
<5>605-609
<6>1998
<7>BcgI is a multi-subunit restriction-modification complex.  BcgI prefers pBR322 DNA over pUC19
in a DNA cleavage reaction. Linearized pBR322 contains two BcgI recognition sites and pUC19
has only one site.  To test whether two target sites are required for BcgI cleavage, one of
the two sites in pBR322 was deleted, and as a result pBR322-1 became a poor substrate for
BcgI.  Conversely, adding a BcgI site to pUC19 makes it a much better substrate for BcgI
cleavage.  In addition, the BcgI (R-M) complex forms a heterohexamer in solution that is
capable of interacting with two recognition sites.  Our results suggest that BcgI requires two
recognition sites for cleavage.

<>

<1>Kong, H., Vincent, M., Xu, Y.
<2>Helicase-dependent amplification of RNA.
<3>US Patent Office
<4>US 7662594 B
<5>
<6>2010
<7>Methods and a kit are provided for selectively and exoponentially amplifying nucleic acids and
include the use of a single strand helicase preparation or a thermostable helicase in the
absence of a single strand binding protein and a DNA polymerase such that the amplification
can be performed isothermally.

<>

<1>Kong, H., Vincent, M., Xu, Y.
<2>Helicase-dependent amplification of nucleic acids.
<3>US Patent Office
<4>US 7829284 B
<5>
<6>2010
<7>Methods and a kit are provided for selectively and exponentially amplifying nucleic acids and
include the use of helicase preparation and a DNA polymerase such that the amplification can
be performed isothermally.

<>

<1>Kong, J., Gu, X., Ma, G.
<2>The characterization of pJW566 from L. lactis subsp. cremoris W56.
<3>Wei Sheng Wu Xue Bao
<4>41
<5>542-547
<6>2001
<7>The plasmid pJW566 was isolated from L. lactis subsp. cremoris W56, one strain for Danish
chadder mixed starter cultures. The strain containing
plasmid pJW566 showed resistance against three common phages species
936, c2 and P335 worldwide. It was found that pJW566 encoded for an
restriction and modification system, and showed strong resistance to
phage CHCP412 when it was introduced into the industrial strain L.
lactis CHCC2281 in milk medium. The endonuclease activity analysis
indicated that the endonuclease required Mg2+, ATP, and was stimulated
by AdoMet.

<>

<1>Kong, J., Josephsen, J.
<2>The ability of the plasmid-encoded restriction and modification system LlaBIII to protect Lactococcus lactis against bacteriophages.
<3>Lett. Appl. Microbiol.
<4>34
<5>249-253
<6>2002
<7>Aims: To investigate the potential of the plasmid-encoded restriction and modification (R/M)
system LlaBIII to protect Lactococcus lactis
against bacteriophages during milk fermentations.
Methods and Results: The R/M system LlaBIII on plasmid pJW566 was
cloned with a chloramphenicol cassette, resulting in plasmid pJK1. When
introduced into L. lactis strains, pJK1 conferred increased phage
resistance against the three most common lactococcal phage species 936,
c2, and P335 and three unclassified industrial phages. The growth of
the strains in RSM was not affected by the presence of plasmid pJK1.
Conclusions: The plasmid-encoded R/M system LlaBIII has great
ability to protect L. lactis strains against bacteriophages in milk
fermentations.
Significance and Impact of the Study: This study evaluates the
ability of the LlaBIII R/M system to function as a phage defence
mechanism which is an essential step prior to considering utilizing it
for improving starter cultures.

<>

<1>Kong, J., Josephsen, J., Ma, G.-R.
<2>The resistance conferred by the R/M system LlaBIII against bacteriophages.
<3>Wei Sheng Wu Xue Bao
<4>42
<5>246-250
<6>2002
<7>LlaBIII, isolated from the naturally occurring plasmid pJW566 from Lactococcus lactis subsp.
cremoris W56, encoded a restriction and modification (R/M) system. The plasmid pJK1 carrying
the R/M system LlaBIII was transformed into L. lactis IL1403 with type I R/M system located on
chromosome and the strain MG1614(pAW601) with AbiS gene on plasmid pAW601, respectively. The
transformants obtained showed stacking resistance against bacteriophages. The plasmid pJK1 was
transformed into industrial strains L. lactis SMQ86 and CHCC2281, the transformants showed the
EOP of the bacteriophages decreased by 10-3 and 10-5, respectively. The results indicated that
the R/M system LlaBIII could protect strains from bacteriophages in dairy fermentation.

<>

<1>Kong, J., Josephsen, J., Ma, G.R.
<2>Cloning and structure analysis of a restriction and modification system, LlaBIII from Lactococcus lactis subsp. cremoris W56.
<3>Sheng Wu Gong Cheng Xue Bao
<4>17
<5>663-668
<6>2001
<7>A 22.4 kb naturally occurring plasmid pJW566, isolated from L. lactis W56, was found to encode
an R/M system named LlaBIII. The LlaBIII R/M system was isolated on a chloramphenicol
resistant derivative of plasmid pJW566, resulting in a plasmid pJK1. Subcloning analysis
showed that the LlaBIII determinant was located on a 5 kb HindIII-Sph I fragment. The fragment
was sequenced. It contained a single open reading frame (ORF), corresponding to a protein of
1584 or 1576 aa. In the deduced amino acid sequence seven helicase motifs characteristic of
endonuclease type I and type III and a conserved catalysis motif X in the R subunits of type I
R/M systems were located in the N-terminus, followed by four conserved motifs found in DNA
N6-adenine methyltransferases. The C-terminus of the deduced amino acid sequence showed no
homology to known R/M systems. Therefore, this polypeptide encoded by LlaBIII is a
multifunctional protein possessing putative DNA recognition, methylation and restriction
activities.

<>

<1>Kong, S.L., Liu, X.Y., Fu, L.W., Yu, X.C., An, C.C.
<2>I-PfoP3I: A Novel Nicking HNH Homing Endonuclease Encoded in the Group I Intron of the DNA Polymerase Gene in Phormidium foveolarum Phage Pf-WMP3.
<3>PLoS ONE
<4>7
<5>e43738
<6>2012
<7>Homing endonucleases encoded in a group I self-splicing intron in a protein-coding gene in
cyanophage genomes have not been reported, apart
from some free-standing homing edonucleases. In this study, a nicking
DNA endonuclease, I-PfoP3I, encoded in a group IA2 intron in the DNA
polymerase gene of a T7-like cyanophage Pf-WMP3, which infects the
freshwater cyanobacterium Phormidium foveolarum is described. The
Pf-WMP3 intron splices efficiently in vivo and self-splices in vitro
simultaneously during transcription. I-PfoP3I belongs to the HNH family
with an unconventional C-terminal HNH motif. I-PfoP3I nicks the
intron-minus Pf-WMP3 DNA polymerase gene more efficiently than the
Pf-WMP4 DNA polymerase gene that lacks any intervening sequence in
vitro, indicating the variable capacity of I-PfoP3I. I-PfoP3I cleaves 4
nt upstream of the intron insertion site on the coding strand of EXON 1
on both intron-minus Pf-WMP3 and Pf-WMP4 DNA polymerase genes. Using an
in vitro cleavage assay and scanning deletion mutants of the intronless
target site, the minimal recognition site was determined to be a 14 bp
region downstream of the cut site. I-PfoP3I requires Mg2+, Ca2+ or Mn2+
for nicking activity. Phylogenetic analysis suggests that the intron
and homing endonuclease gene elements might be inserted in Pf-WMP3
genome individually after differentiation from Pf-WMP4. To our
knowledge, this is the first report of the presence of a group I
self-splicing intron encoding a functional homing endonuclease in a
protein-coding gene in a cyanophage genome.

<>

<1>Konstantinidis, K.T., DeLong, E.F.
<2>Genomic patterns of recombination, clonal divergence and environment in marine microbial populations.
<3>ISME J.
<4>2
<5>1052-1065
<6>2008
<7>Microorganisms represent the largest reservoir of biodiversity on Earth,
both in numbers and total genetic diversity, but it remains unclear
whether this biodiversity is organized in discrete units that correspond
to ecologically coherent species. To further explore this question, we
examined patterns of genomic diversity in sympatric microbial populations.
Analyses of a total of approximately 200 Mb of microbial community genomic
DNA sequence recovered from 4000 m depth in the Pacific Ocean revealed
discrete sequence-defined populations of Bacteria and Archaea, with
intrapopulation genomic sequence divergence ranging from approximately 1%
to approximately 6%. The populations appeared to be maintained, at least
in part, by intrapopulation genetic exchange (homologous recombination),
although the frequency of recombination was estimated to be about three
times lower than that observed previously in thermoacidophilic archaeal
biofilm populations. Furthermore, the genotypes of a given population were
clearly distinguishable from their closest co-occurring relatives based on
their relative abundance in situ. The genetic distinctiveness and the
matching sympatric abundances imply that these genotypes share similar
ecophysiological properties, and therefore may represent fundamental units
of microbial diversity in the deep sea. Comparisons to surface-dwelling
relatives of the Sargasso Sea revealed that distinct sequence-based
clusters were not always detectable, presumably due to environmental
variations, further underscoring the important relationship between
environmental contexts and genetic mechanisms, which together shape and
sustain microbial population structure.

<>

<1>Kontur, W.S., Schackwitz, W.S., Ivanova, N., Martin, J., Labutti, K., Deshpande, S., Tice, H.N., Pennacchio, C., Sodergren, E., Weinstock, G.M., Noguera, D.R., Donohue, T.J.
<2>Revised Sequence and Annotation of the Rhodobacter sphaeroides 2.4.1 Genome.
<3>J. Bacteriol.
<4>194
<5>7016-7017
<6>2012
<7>The DNA sequences of chromosomes I and II of Rhodobacter sphaeroides strain 2.4.1 have been
revised, and the annotation of the entire genomic sequence, including
both chromosomes and the five plasmids, has been updated. Errors in the
originally published sequence have been corrected, and approximately 11% of the
coding regions in the original sequence have been affected by the revised
annotation.

<>

<1>Koo, H., Basu, M.K., Crowley, M., Aislabie, J., Bej, A.K.
<2>Draft Genome Sequence of Pseudomonas sp. Strain Ant30-3, a Psychrotolerant Bacterium with Biodegradative Attribute Isolated from Antarctica.
<3>Genome Announcements
<4>2
<5>e00522-14
<6>2014
<7>Pseudomonas sp. strain Ant30-3, isolated from fuel-contaminated Antarctic soil, exhibited
distinctive psychrotolerant attributes and the potential for degrading
aromatic hydrocarbon compounds at cold temperatures. We report here the 6.14-Mb
draft genome of Ant30-3, which will provide insights into the genomic basis for
the psychrotolerant and biodegradative properties of this bacterium.

<>

<1>Koo, H., Ptacek, T., Crowley, M., Swain, A.K., Osborne, J.D., Bej, A.K., Andersen, D.T.
<2>Draft Genome Sequence of Hymenobacter sp. Strain IS2118, Isolated from a Freshwater Lake in Schirmacher Oasis, Antarctica, Reveals Diverse Genes for  Adaptation to Cold Ecosystems.
<3>Genome Announcements
<4>2
<5>e00739-14
<6>2014
<7>Hymenobacter sp. IS2118, isolated from a freshwater lake in Schirmacher Oasis, Antarctica,
produces extracellular polymeric substance (EPS) and manifests
tolerance to cold, UV radiation (UVR), and oxidative stress. We report the
5.26-Mb draft genome of strain IS2118, which will help us to understand its
adaptation and survival mechanisms in Antarctic extreme ecosystems.

<>

<1>Koob, M.
<2>Conferring new cleavage specificities of restriction endonucleses.
<3>Methods Enzymol.
<4>216
<5>321-329
<6>1992
<7>Detailed protocols for Achilles Heel cleavage are presented.

<>

<1>Koob, M., Burkiewicz, A., Kur, J., Szybalski, W.
<2>RecA-AC: single-site cleavage of plasmids and chromosomes at any predetermined restriction site.
<3>Nucleic Acids Res.
<4>20
<5>5831-5836
<6>1992
<7>We have developed a novel version of the Achilles' Cleavage (AC) reaction in which virtually
any restriction site on DNA of any size can be converted to a unique cleavage site. We first
polymerized RecA protein on a synthetic oligodeoxyribonucleotide (oligo) in the presence of a
nonhydrolyzable ATP analogue to generate oligo:RecA nucleoprotein filaments. These filaments
were then incubated with plasmid or intact chromosomal DNA from Saccharomyces cerevisiae to
form stable complexes in the yeast LEU2 gene at the target sequence identical (or
complementary) to that of the oligo. When HhaII (HinfI) methyltransferase (M.HhaII) was added,
all of the recognition sites for HhaII with the exception of the one protected by the RecA
filament were methylated and thus no longer cleaved by the cognate restriction endonuclease
(HinfI). After inactivation of the RecA and the M.HhaII, HinfI was used to efficiently cleave
the plasmid or chromosome specifically at the targeted restriction site. Since oligos specific
for any sequence can be easily synthesized and the other reagents necessary to perform
RecA-mediated AC (RecA-AC) reactions on both plasmids and intact chromosomes are readily
available, this procedure can be applied immediately to the precise dissection and analysis of
genomic DNA from any source and to any other research problem requiring efficient, highly
specific cleavage of DNA at predetermined sites.

<>

<1>Koob, M., Grimes, E., Szybalski, W.
<2>Conferring operator specificity on restriction endonucleases.
<3>Science
<4>241
<5>1084-1086
<6>1988
<7>Mapping and manipulation of very large genomes, including the human genome,
would be facilitated by the availability of a DNA cleavage method with very
high site specificity.  Therefore, a general method was devised that extends
the effective recognition sequences well beyond the present 8-base pair limit
by combining the specificity of the restriction endonuclease with that of
another sequence-specific protein that binds tightly to DNA.  It was shown that
the tightly binding lac or lamba repressor protects a restriction site within
the operator from specific modification methylases, M.HhaI or M.HphI, while all
other similar sites are methylated and thus rendered uncleavable.  A plasmid
containing a symmetric lac operator was specifically cleaved by HhaI, only at
the site within the operator, after M.HhaI methylation in the presence of the
lac repressor, whereas the remaining 31 HhaI sites on this plasmid were
methylated and thus not cleaved.  Analogous results were obtained with the
HaeII site within the lac operator which was similarly protected by the lac
repressor, and with the HphI site within the phage lambda OL operator, which
was protected by lambda repressor from M.HphI methylation.

<>

<1>Koob, M., Grimes, E., Szybalski, W.
<2>Conferring new specificity upon restriction endonucleases by combining repressor-operator interaction and methylation.
<3>Gene
<4>74
<5>165-167
<6>1988
<7>Meeting Abstract

<>

<1>Koob, M.D.
<2>Achilles' cleavage:  A general approach for conferring expanded specificities on restriction endonucleases.
<3>Diss. Abstr.
<4>51
<5>5161B
<6>1991
<7>Mapping and manipulation of very large genomes would be facilitated by the
availability of a DNA cleavage method with a very high site specificity.  I
have devised a general method for efficiently extending the effective
recognition sequence of a restriction endonuclease well beyond its natural
limits.  The key to this process, which I call Achilles' Cleavage (AC), is
methylation of the DNA substrate in the presence of a DNA-binding molecule that
forms sequence-specific complexes capable of excluding the methyltransferase
(MTase).  Cleavable restriction sites remain only at those sites where the
recognition sites for the cognate restriction enzyme and the DNA-binding
molecule overlap.  The lac repressor and its ideal, symmetric binding site
(lacOs), which contains the recognition sites for both HaeII (5'-AGCGCT) and
HhaI (5'-GCGC), were used as a model system for the development of this
approach.  Plasmid DNA containing lacOs was methylated by HhaI MTase (M-HhaI)
in the presence of LacI.  After inactivation of M.HhaI and lacI, HhaI and HaeII
were used to completely and specifically cleave the plasmid at the lacOs
sequence.  LacI-mediated AC of a lambda genome containing lacOs produced
similar results.  In order to test this approach on the large genomes for which
it was designed, protocols were developed for the efficient methylation and
digestion of intact chromosomal DNA embedded in agarose microbeads.  Model
genomes were generated by introducing lacOs into the 4.7-Mb circular genome of
Escherichia coli and into one of the 16 chromosomes in the 15-Mb genome of
Saccharomyces cerevisiae.  Subsequent treatment with the AC protocol resulted
in complete cleavage of these genomes exclusively at the inserted lacOs.  These
experiments demonstrate the feasibility of using the AC approach to efficiently
extend the specificity of restriction enzymes and create new tools for the
mapping precise molecular dissection of multimegabase genomes.

<>

<1>Koonin, E.V.
<2>A protein splice-junction motif in hedgehog family proteins.
<3>Trends Biochem. Sci.
<4>20
<5>141-142
<6>1995
<7>Protein splicing is a recently discovered, complex post-translational
modification that includes two concerted proteolytic cleavages and one ligation reaction,
and produces two distinct, functional proteins from a single polypeptide.  By analogy with
introns and exons in RNA precursors, the internal protein that is excised from the
translation product is called an intein, whereas the two terminal portions, when ligated,
form a fusion protein called extein.  So far, the intein-extein organization has been
observed in about ten proteins from yeast, bacteria and Archaea; in most of these cases, the
inteins are inserted within or near nucleotide-binding domains.  Protein splicing is thought
to be an autocatalytic process, since it can occur in heterologous systems; for example,
intein-containing proteins from Mycobacterium and Thermococcus have been synthesized
in Escherichia coli, and protein splicing has been demonstrated with the purified intein-
containing DNA polymerase from Pyrococcus.

<>

<1>Kopecka, H.
<2>A rapid purification method of restriction endonucleases from Haemophilus strains.
<3>Biochim. Biophys. Acta
<4>391
<5>109-120
<6>1975
<7>A simple and rapid method of purification of restriction endonucleases from
different Haemophilus strains is presented.  By this method highly purified and
stable enzymes can be obtained.  Separation of different restriction activities
present in the same strain is possible.  This method was so far successfully
used with Haemophilus influenzae, Haemophilus parainfluenzae and Haemophilus
aegyptius strains.  The main advantages over previously published procedures
reside in the simplification of certain purification steps (for instance the
BioGel A 0.5 M filtration is replaced by a hydroxyapatite batch step),
elimination of exonuclease activity by fractionation with (NH4)2SO4, separation
of different restriction activities by phosphocellulose chromatography,
application of this method to various strains and high purification degree of
enzymes.

<>

<1>Kopel, M., Helbert, W., Henrissat, B., Doniger, T., Banin, E.
<2>Draft Genome Sequence of Nonlabens ulvanivorans, an Ulvan-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e00793-14
<6>2014
<7>Here we report the draft genome sequence of the bacterium Nonlabens ulvanivorans, which was
recently isolated. To our knowledge, this is the first published genome
of a characterized ulvan-degrading bacterium. Revealing the ulvan utilization
pathways may provide access to a vast marine biomass source that has yet to be
exploited.

<>

<1>Kopel, M., Helbert, W., Henrissat, B., Doniger, T., Banin, E.
<2>Draft Genome Sequences of Two Ulvan-Degrading Isolates, Strains LTR and LOR, That Belong to the Alteromonas Genus.
<3>Genome Announcements
<4>2
<5>e01081-14
<6>2014
<7>Here, we report the draft genome sequence of two ulvan-degrading Alteromonas spp. isolated
from the feces of the sea slug, Aplysia. These sequenced genomes display
a unique ulvan degradation machinery compared with ulvanolytic enzymes previously
identified in Nonlabens ulvanivorans.

<>

<1>Kopel, M., Helbert, W., Henrissat, B., Doniger, T., Banin, E.
<2>Draft Genome Sequence of Pseudoalteromonas sp. Strain PLSV, an Ulvan-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e01257-14
<6>2014
<7>We present the draft genome sequence of Pseudoalteromonas sp. strain PLSV, isolated from the
feces of an Aplysia sea slug. The addition of the PLSV genome
to the existing genomes of three other ulvan-degrading bacterial species will
enhance our understanding of ulvan utilization.

<>

<1>Kopf, M., Klahn, S., Voss, B., Stuber, K., Huettel, B., Reinhardt, R., Hess, W.R.
<2>Finished Genome Sequence of the Unicellular Cyanobacterium Synechocystis sp. Strain PCC 6714.
<3>Genome Announcements
<4>2
<5>e00757-14
<6>2014
<7>Synechocystis sp. strain PCC 6714 is a unicellular cyanobacterium closely related to the
popular model organism Synechocystis sp. strain PCC 6803. A combination of
PacBio SMRT and Illumina GAIIx data results in a highly accurate finished genome
sequence that provides a reliable resource for further comparative analyses.

<>

<1>Kopke, M., Held, C., Hujer, S., Liesegang, H., Wiezer, A., Wollherr, A., Ehrenreich, A., Liebl, W., Gottschalk, G., Durre, P.
<2>Clostridium ljungdahlii represents a microbial production platform based on syngas.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>13087-13092
<6>2010
<7>Clostridium ljungdahlii is an anaerobic homoacetogen, able to ferment
sugars, other organic compounds, or CO(2)/H(2) and synthesis gas
(CO/H(2)). The latter feature makes it an interesting microbe for the
biotech industry, as important bulk chemicals and proteins can be produced
at the expense of CO(2), thus combining industrial needs with sustained
reduction of CO and CO(2) in the atmosphere. Sequencing the complete
genome of C. ljungdahlii revealed that it comprises 4,630,065 bp and is
one of the largest clostridial genomes known to date. Experimental data
and in silico comparisons revealed a third mode of anaerobic
homoacetogenic metabolism. Unlike other organisms such as Moorella
thermoacetica or Acetobacterium woodii, neither cytochromes nor sodium
ions are involved in energy generation. Instead, an Rnf system is present,
by which proton translocation can be performed. An electroporation
procedure has been developed to transform the organism with plasmids
bearing heterologous genes for butanol production. Successful expression
of these genes could be demonstrated, leading to formation of the biofuel.
Thus, C. ljungdahlii can be used as a unique microbial production platform
based on synthesis gas and carbon dioxide/hydrogen mixtures.

<>

<1>Korba, B.E., Hays, J.B.
<2>Partially deficient methylation of cytosine in DNA at CCA/TGG sites stimulates genetic recombination of bacteriophage lambda.
<3>Cell
<4>28
<5>531-541
<6>1982
<7>Lambda bacteriophages grown on arl mutants of Escherichia coli display
intermediate levels
of cytosine methylation: less 5-methylcytosine than phages grown on wild-type bacteria but
more than
phages grown on dcm mutants, and thus lacking the methylated sequences (Cm5CA/TGG)
characteristic of E.
coli K-12 bacteria.  Arl- phages are one twelfth as resistant to EcoRII restriction
(recognition site CCA/TGG)
as Arl+ phages, but 40-fold more resistant than Dcm- phages.  Chromatographic analyses
show the 5-
methylcytosine content of Arl- DNA to be one third that of Arl+ DNA.  Altered cytosine
methylation
frequency correlates with two previously described properties of Arl- phages, increased
genetic
recombination and unusual sensitivity of phage DNA to endonuclease S1, which are absent
in phages grown
on dcm or dcm arl bacteria.  Methylated/unmethylated heteroduplex DNA prepared in vitro
(one strand from
Eco RII-modified phages/one from Dcm- phages) is highly recombinogenic but not S1-
sensitive.  We
hypothesize that hemimethylated CCA/TGG sites in Arl- DNA are necessary and sufficient
for enhanced
recombination, and necessary but not sufficient for S1 sensitivity.

<>

<1>Korch, C.
<2>Cross index for improving cloning selectivity by partially filling in 5'-extensions of DNA produced by type II restriction endonucleases.
<3>Nucleic Acids Res.
<4>15
<5>3199-3220
<6>1987
<7>A cross index is presented for using the improved selectivity offered by the
Hung and Wensink (Nucl. Acids Res. 12, 1863-1874, 1984) method of partially
filling in 5'-extensions produced by type II restriction endonucleases.   After
this treatment, DNA fragments which normally cannot be ligated to one another,
can be joined providing that complementary cohesive ends have been generated.
The uses of this technique, which include the prevention of DNA fragments (both
vector and insert) auto-annealing, are discussed.

<>

<1>Korch, C., Hagblom, P.
<2>In-vivo-modified gonococcal plasmid pJD1: A model system for analysis of restriction enzyme sensitivity to DNA modifications.
<3>Eur. J. Biochem.
<4>161
<5>519-524
<6>1986
<7>The 4207-bp cryptic plasmid (pJD1) of Neisseria gonorrhoeae has 5-methylcytosine bases present
at several positions in the DNA sequence. Fortuitously, these modified bases lie in the
recognition sequences of many restriction enzymes.  This feature makes the cryptic plasmid a
model system for assaying the effect of these modified cytosines on the activities of the
following restriction endonucleases and their isoschizomers: R.AvaII, R.BamHI, R.BglI,
R.Fnu4HI, R.HaeII, R.HaeIII, R.HhaI, R.HpaII, R.KpnI, R.MspI, R.NaeI, R.NarI, R.NciI, R.NgoI,
R.NgoII, and R.Sau96I.  Of particular interest was the finding that methylation of one of the
external cytosines of the palindrome 5'-CCGG-3' prevented its cleavage by R.MspI, but not by
R.HpaII as had been suggested by Walder et al. [J. Biol. Chem. (1983) 158,1235-1241]. Shows
HhaI cleaves hemimethylated GCGm5C and BglI cannot cleave 5'GCCNNNNNGGm5C/3'CGGNNNNNCm5CG.

<>

<1>Korch, C., Hagblom, P., Normark, S.
<2>Type III 5-methylcytosine modification of DNA in Neisseria gonorrhoeae.
<3>J. Bacteriol.
<4>161
<5>1236-1237
<6>1985
<7>We present here the first report of a type III methyltransferase that modifies a cytosine,
Neisseria gonorrhoeae 82409/55(pJD1) modifies the first cytosine on only one strand from the
5' end of the nonpalindromic sequence: 5'-GGTGA-3' 3'-CCACT-5' m.  We have called this
modifying activity M.NgoVIII.
[ The enzyme called NgoVIII in this abstract has been renamed NgoHVIII, Jan/1998. ]

<>

<1>Korch, C., Hagblom, P., Normark, S.
<2>Sequence-specific DNA modification in Neisseria gonorrhoeae.
<3>J. Bacteriol.
<4>155
<5>1324-1332
<6>1983
<7>Neisseria gonorrhoaea 82409/55(pJD1) is postulated to possess six DNA sequence-specific
cytosine methyltransferases and one DNA sequence-specific N6-adenine methyltransferase.  From
the DNA sequencing of the plasmid pJD1 (manuscript in preparation) by a modification of the
Maxam and Gilbert chemical cleavage procedure, the cytosine methylation specificities were
demonstrated. Five of these methylating enzymes and their respective specificities are M.NgoI
(5'-PuGmCGCPy-3'), M.NgoII (5'-GGmCC-3'), M.NgoIV (5'-GmCCGGC-3'), M.NgoV
(5'-GGNNmCC-3'), and M.NgoVII (5'-GmC(C/G)GC-3').  M.NgoVI (5'-GATC-3') does not
methylate the cytosine of its recognition sequence, in agreement with a detected adenine
modification.  A biological implication of these different DNA methylating activities is
discussed.
[ The enzyme called M.NgoI in this abstract has been renamed M.NgoHIP, Jan/1998. ]
[ The enzyme called M.NgoII in this abstract has been renamed M.NgoHIIP, Jan/1998. ]
[ The enzyme called M.NgoIV in this abstract has been renamed M.NgoHIVP, Jan/1998. ]
[ The enzyme called M.NgoV in this abstract has been renamed M.NgoHVP, Jan/1998. ]
[ The enzyme called M.NgoVI in this abstract has been renamed M.NgoHVIP, Jan/1998. ]
[ The enzyme called M.NgoVII in this abstract has been renamed M.NgoHVIIP, Jan/1998. ]

<>

<1>Koren, S., Harhay, G.P., Smith, T.P., Bono, J.L., Harhay, D.M., McVey, S.D., Radune, D., Bergman, N.H., Phillippy, A.M.
<2>Reducing assembly complexity of microbial genomes with single-molecule sequencing.
<3>Genome Biol.
<4>14
<5>R101
<6>2013
<7>BACKGROUND: The short reads output by first- and second-generation DNA sequencing instruments
cannot completely reconstruct microbial chromosomes. Therefore, most
genomes have been left unfinished due to the significant resources required to
manually close gaps in draft assemblies. Third-generation, single-molecule
sequencing addresses this problem by greatly increasing sequencing read length,
which simplifies the assembly problem. RESULTS: To measure the benefit of
single-molecule sequencing on microbial genome assembly, we sequenced and
assembled the genomes of six bacteria and analyzed the repeat complexity of 2,267
complete bacteria and archaea. Our results indicate that the majority of known
bacterial and archaeal genomes can be assembled without gaps, at finished-grade
quality, using a single PacBio RS sequencing library. These single-library
assemblies are also more accurate than typical short-read assemblies and hybrid
assemblies of short and long reads. CONCLUSIONS: Automated assembly of long,
single-molecule sequencing data reduces the cost of microbial finishing to $1,000
for most genomes, and future advances in this technology are expected to drive
the cost lower. This is expected to increase the number of completed genomes,
improve the quality of microbial genome databases, and enable high-fidelity,
population-scale studies of pan-genomes and chromosomal organization.

<>

<1>Korlach, J., Turner, S.W.
<2>Going beyond five bases in DNA sequencing.
<3>Curr. Opin. Struct. Biol.
<4>22
<5>251-261
<6>2012
<7>DNA sequencing has provided a wealth of information about biological systems, but thus far has
focused on the four canonical bases, and 5-methylcytosine through
comparison of the genomic DNA sequence to a transformed four-base sequence
obtained after treatment with bisulfite. However, numerous other chemical
modifications to the nucleotides are known to control fundamental life functions,
influence virulence of pathogens, and are associated with many diseases. These
modifications cannot be accessed with traditional sequencing methods. In this
opinion, we highlight several emerging single-molecule sequencing techniques that
have the potential to directly detect many types of DNA modifications as an
integral part of the sequencing protocol.

<>

<1>Kornspan, J.D., Lysnyansky, I., Kahan, T., Herrmann, R., Rottem, S., Nir-Paz, R.
<2>Genome analysis of a Mycoplasma hyorhinis strain derived from a primary human melanoma cell line.
<3>J. Bacteriol.
<4>193
<5>4543-4544
<6>2011
<7>The complete genome of Mycoplasma hyorhinis strain MCLD has been sequenced and annotated. This
genome differs by inversion of a 14.4kb and 3.7kb
fragments and a deletion of a 9.9kb fragment from M. hyorhinis strain
HUB-1, isolated from swine respiratory tract. The genome revealed 778 CDSs
with a limited number of vlp genes encoding for variable surface
lipoproteins.

<>

<1>Korolev, S.V., Samko, O.T., Eldarov, M.A., Kalugin, A.A., Khoroshutina, E.B., Omelyanyuk, N.M., Sokolov, N.N., Skryabin, K.G.
<2>Site-specific endonuclease BcuAI from Bacillus cereus A.
<3>Bioorg. Khim.
<4>22
<5>528-531
<6>1996
<7>A new restriction endonuclease was isolated from Bacillus cereus BKM B-814 by means of the
cell disruption with ultrasonication, ammonium sulfate fractionation of the cell-free extract,
and chromatography on DEAE-Sepharose to give about 1400 U of the enzyme per gram of cells.
The enzyme revealed the maximum activity at 30-37oC, pH 7.6-8.2, and 5-10 mM MgCl2 under a
high ionic strength (50mM Tris-HCl, 100mM NaCl).  The site-specific endonuclease BcuAI was
found to recognize the 5' G/G(A/T)CC sequence in double-stranded DNA and cleave it as shown
with the arrow, thus being a true isoschisomer of the AvaII restriction endonuclease.

<>

<1>Korolev, S.V., Sokolov, N.N., Rina, M., Eldarov, M.A., Omelyanyuk, N.M., Markaki, M., Skryabin, K.G., Bouriotis, V.
<2>Isolation and characterization of the specificity of new restriction endonucleases Bsp4009I and AsiI - isoschizomers of BamHI.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>32-35
<6>1998
<7>New type II restriction endonucleases AsiI and Bsp4009I are detected in Azotobacter species
N55 and Bacillus species 4009, respectively.  Purified preparations of the restriction enzymes
free from interfering nucleases and phosphatases were obtained by column chromatography on
phosphocellulose and heparin-sepharose (AsiI) and phosphocellulose and DEAE-cellulose
(Bsp4009I).  The yield of purified AsiI and Bsp4009I was 16 x 10^3 and 8 x 10^3 units per g of
wet cells, respectively.  The above restriction endonucleases recognize the 5'-G/GATCC-3'
sequence on double-stranded DNA and cleave it as shown, thus being true isoschizomers of BamHI
restriction endonuclease.

<>

<1>Korona, R., Korona, B., Levin, B.R.
<2>Sensitivity of naturally occurring coliphages to type I and type II restriction and modification.
<3>J. Gen. Microbiol.
<4>139
<5>1283-1290
<6>1993
<7>Protection against lethal infections by bacteriophage may seem the most likely role of
restriction-modification (R-M) systems in bacteria and the reason for their evolution. There
are, however, phenomena which question this phage-mediated selection hypothesis for the
maintenance of extant R-M systems. Most prominent among these are the mechanisms phage have to
avoid or otherwise limit the effects of the restriction endonucleases produced by their host
bacteria. To evaluate the importance of these antirestriction mechanisms in Escherichia coli,
we have examined the sensitivity of coliphage from natural and laboratory sources to a series
of type I and II R-M systems. The results of our study indicate that, in vivo, restriction
endonucleases have no effect on a substantial fraction of naturally occurring coliphage. The
absence of restriction sites appears to be the most common reason why these phage are
unaffected by type II restriction endonucleases, but other antirestriction mechanisms also
operate. On the other hand, the frequency of naturally occurring coliphage sensitive to
restriction appears sufficiently great for phage-mediated selection to be a viable hypothesis
for the maintenance R-M in E. coli and its accessory elements.

<>

<1>Korona, R., Levin, B.R.
<2>Phage-mediated selection and the evolution and maintenance of restriction-modification.
<3>Evolution
<4>47
<5>556-575
<6>1993
<7>Restriction-modification (R-M) was discovered because it provides bacteria with immunity to
phage infection. But, is phage-mediated selection the sole mechanism responsible for the
evolution and maintenance of these ubiquitous and multiply evolved systems? In an effort to
answer this question, we have performed experiments with laboratory populations of E. coli and
phage and computer simulations. We consider two ecological situations whereby phage-mediated
selection could favor R-M immunity; i) when bacteria with a novel R-M system invade
communities of phage-sensitive bacteria in which there are one or more species of phage, and
ii) when bacteria colonize bacterial-free habitats in which phage are present. The results of
our experiments indicate that in established communities of bacteria and phage, the advantage
R-M provides an invading population of bacteria is ephemeral. Within short order, mutants
resistant (refractory) to the phage evolve in the dominant population and subsequently in the
invading population. The outcome of competition then depends on the relative fitness of the
resistant states of these bacterial clones, rather than R-M. As a consequence of sequential
selection for independent mutants, this rapid evolution of resistance occurs even when two and
three species of phage are present. While in our experiments resistance also evolved when
bacteria colonized new habitats in which phage were present, a novel R-M system greatly
augmented the likelihood of their becoming established. We interpret the results of this study
as support for the hypothesis that the latter, colonization selection, may play an important
role in the evolution and maintenance of restriction-modification. However, we also see these
results and other observations we discuss as questioning whether protection against phage is
the unique biological role of restriction-modification.

<>

<1>Koryszewska-Baginska, A., Aleksandrzak-Piekarczyk, T., Bardowski, J.
<2>Complete Genome Sequence of the Probiotic Strain Lactobacillus casei (Formerly Lactobacillus paracasei) LOCK919.
<3>Genome Announcements
<4>1
<5>e00758-13
<6>2013
<7>Lactobacillus casei is usually regarded as a bacterium that lives naturally in the human
intestinal tract, where it can contribute to host health and
well-being. We describe here the complete genome sequence of L. casei LOCK919, a
strain with probiotic properties isolated from child feces. The genome consists
of a 3.11-Mb chromosome and a 29,768-bp plasmid.

<>

<1>Koryszewska-Baginska, A., Bardowski, J., Aleksandrzak-Piekarczyk, T.
<2>Genome Sequence of the Probiotic Strain Lactobacillus rhamnosus (Formerly Lactobacillus casei) LOCK908.
<3>Genome Announcements
<4>2
<5>e00120-14
<6>2014
<7>Lactobacillus rhamnosus LOCK908, a patented probiotic strain (Polish patent no. 209987), was
isolated from the feces of a healthy 6-year-old girl. Here, we
present the complete genome sequence of LOCK908 and identify genes likely to be
involved in the biosynthesis of exopolysaccharides (EPSs).

<>

<1>Korzun, V., Balzer, H.J., Balzer, A., Baumlein, H., Borner, A.
<2>Chromosomal location of three wheat sequences with homology to pollen allergen encoding, DNA replication regulating, and DNA (cytosine-5)-methyltransferase genes in wheat and rye.
<3>Genome
<4>39
<5>1213-1215
<6>1996
<7>Three wheat sequences, shown to be homologous to pollen allergen encoding, DNA replication
regulating, and DNA (cytosine-5)-methyltransferase genes were localized on chromosomes using
nullisomic-tetrasomic wheat ('Chinese Spring') and wheat-rye ('Chinese
Spring'/'Imperial') addition lines.  Whereas the loci for the pollen allergen encoding
sequence (Tri a III) were shown to be located on homoeologous group 4, the DNA replication
regulating (Rep) and DNA (cytosine-5)-methyltransferase (Mtase) genes were located to
homoeologous groups 1 and 7, respectively, of Triticeae.  Chromosomal rearrangements in wheat
and rye relative to each other are discussed.

<>

<1>Kosaka, T., Toh, H., Toyoda, A.
<2>Complete Genome Sequence of a Thermophilic Hydrogenotrophic Methanogen, Methanothermobacter sp. Strain CaT2.
<3>Genome Announcements
<4>1
<5>e00672-13
<6>2013
<7>We isolated a thermophilic hydrogenotrophic methanogen, Methanothermobacter sp. strain CaT2,
which is able to aggregate and utilize formate. Here, we report the
complete genome sequence of this organism.

<>

<1>Kosaka, T., Uchiyama, T., Ishii, S., Enoki, M., Imachi, H., Kamagata, Y., Ohashi, A., Harada, H., Ikenaga, H., Watanabe, K.
<2>Reconstruction and regulation of the central catabolic pathway in the thermophilic propionate-oxidizing syntroph Pelotomaculum  thermopropionicum.
<3>J. Bacteriol.
<4>188
<5>202-210
<6>2006
<7>Obligate anaerobic bacteria fermenting volatile fatty acids in syntrophic association with
methanogenic archaea share the intermediate bottleneck
step in organic-matter decomposition. These organisms (called syntrophs)
are biologically significant in terms of their growth at the thermodynamic
limit and are considered to be the ideal model to address bioenergetic
concepts. We conducted genomic and proteomic analyses of the thermophilic
propionate-oxidizing syntroph Pelotomaculum thermopropionicum to obtain
the genetic basis for its central catabolic pathway. Draft sequencing and
subsequent targeted gap closing identified all genes necessary for
reconstructing its propionate-oxidizing pathway (i.e., methylmalonyl
coenzyme A pathway). Characteristics of this pathway include the
following. (i) The initial two steps are linked to later steps via
transferases. (ii) Each of the last three steps can be catalyzed by two
different types of enzymes. It was also revealed that many genes for the
propionate-oxidizing pathway, except for those for propionate coenzyme A
transferase and succinate dehydrogenase, were present in an operon-like
cluster and accompanied by multiple promoter sequences and a putative gene
for a transcriptional regulator. Proteomic analysis showed that enzymes in
this pathway were up-regulated when grown on propionate; of these enzymes,
regulation of fumarase was the most stringent. We discuss this tendency of
expression regulation based on the genetic organization of the open
reading frame cluster. Results suggest that fumarase is the central
metabolic switch controlling the metabolic flow and energy conservation in
this syntroph.

<>

<1>Kosinski, J., Kubareva, E., Bujnicki, J.M.
<2>A model of restriction endonuclease MvaI in complex with DNA: A template for interpretation of experimental data and a guide for specificity engineering.
<3>Proteins
<4>68
<5>324-336
<6>2007
<7>R.MvaI is a Type II restriction enzyme (REase), which specifically recognizes the
pentanucleotide DNA sequence 5'-CCWGG-3' (W indicates A
or T). It belongs to a family of enzymes, which recognize related
sequences, including 5'-CCSGG-3'(S indicates G or C) in the case of
R.BcnI, or 5'-CCNGG-3' (where N indicates any nucleoside) in the case
of R.ScrFI. REases from this family hydrolyze the phosphodiester bond
in the DNA between the 2nd and 3rd base in both strands, thereby
generating a double strand break with 5'-protruding single nucleotides.
So far, no crystal structures of REases with similar cleavage patterns
have been solved. Characterization of sequence-structure-function
relationships in this family would facilitate understanding of
evolution of sequence specificity among REases and could aid in
engineering of enzymes with new specificities. However, sequences of
R.MvaI or its homologs show no significant similarity to any proteins
with known structures, thus precluding straightforward comparative
modeling. We used a fold recognition approach to identify a remote
relationship between R.MvaI and the structure of DNA repair enzyme
MutH, which belongs to the PD-(D/E)XK superfamily together with many
other REases. We constructed a homology model of R.MvaI and used it to
predict functionally important amino acid residues and the mode of
interaction with the DNA. In particular, we predict that only one
active site of R.MvaI interacts with the DNA target at a time, and the
cleavage of both strands (5'-CCAGG-3' and 5'-CCTGG-3') is achieved by
two independent catalytic events. The model is in good agreement with
the available experimental data and will serve as a template for
further analyses of R.MvaI, R.BcnI, R.ScrFI and other related enzymes.

<>

<1>Koskinen, J.P., Laine, P., Niemi, O., Nykyri, J., Harjunpaa, H., Auvinen, P., Paulin, L., Pirhonen, M., Palva, T., Holm, L.
<2>Genome Sequence of Pectobacterium sp. Strain SCC3193.
<3>J. Bacteriol.
<4>194
<5>6004
<6>2012
<7>We report the complete and annotated genome sequence of the plant-pathogenic enterobacterium
Pectobacterium sp. strain SCC3193, a model strain isolated from
potato in Finland. The Pectobacterium sp. SCC3193 genome consists of a 516,411-bp
chromosome, with no plasmids.

<>

<1>Kossykh, V., Repyk, A., Kaliman, A., Buryanov, Y.
<2>Nucleotide sequence of the EcoRII restriction endonuclease gene.
<3>Biochim. Biophys. Acta
<4>1009
<5>290-292
<6>1989
<7>The nucleotide sequence of a 1394 basepair (bp) DNA fragment containing the EcoRII restriction
endonuclease (R.EcoRII) gene was determined.  The endonuclease gene is 1206 bp in length
(predicted 402 amino acids (aa) and Mr=45,178) and is separated by 33 bp from the EcoRII
modification methylase (M.EcoRII) gene.  The EcoRII restriction-modification system has a
tail-to-tail organization of the two genes.

<>

<1>Kossykh, V.G., Lloyd, R.S.
<2>A DNA Adenine Methyltransferase of Escherichia coli That Is Cell Cycle Regulated and Essential for Viability.
<3>J. Bacteriol.
<4>186
<5>2061-2067
<6>2004
<7>DNA sequence analysis revealed that the putative yhdJ DNA methyltransferase gene of
Escherichia coli is 55% identical to the Nostoc
sp. strain PCC7120 gene encoding DNA methyltransferase AvaIII, which
methylates adenine in the recognition sequence, ATGCAT. The yhdJ gene was
cloned, and the enzyme was overexpressed and purified. Methylation and
restriction analysis showed that the DNA methyltransferase methylates the
first adenine in the sequence ATGCAT. This DNA methylation was found to be
regulated during the cell cycle, and the DNA adenine methyltransferase was
designated M.EcoKCcrM (for "cell cycle-regulated methyltransferase"). The
CcrM DNA adenine methyltransferase is required for viability in E. coli,
as a strain lacking a functional genomic copy of ccrM can be isolated only
in the presence of an additional copy of ccrM supplied in trans. The cells
of such a knockout strain stopped growing when expression of the inducible
plasmid ccrM gene was shut off. Overexpression of M.EcoKCcrM slowed
bacterial growth, and the ATGCAT sites became fully methylated throughout
the cell cycle; a high proportion of cells with an anomalous size
distribution and DNA content was found in this population. Thus, the
temporal control of this methyltransferase may contribute to accurate cell
cycle control of cell division and cellular morphology. Homologs of
M.EcoKCcrM are present in other bacteria belonging to the gamma
subdivision of the class Proteobacteria, suggesting that methylation at
ATGCAT sites may have similar functions in other members of this group.

<>

<1>Kossykh, V.G., Repyk, A.V., Hattman, S.
<2>Sequence motifs common to the EcoRII restriction endonuclease and the proposed sequence specificity domain of three DNA-[cytosine-C5] methyltransferases.
<3>Gene
<4>125
<5>65-68
<6>1993
<7>We have compared the deduced amino acids (aa) sequences of the EcoRII restriction endonuclease
(R.EcoRII) and the proposed specificity (target recognition) domains of three
DNA-[cytosine-C5] methyltransferases (MTases). M.EcoRII, M.Dcm, and M.SPR, each of which
recognizes the same nucleotide sequence, CCWGG (where W is A or T). We have identified a
region containing sequence motifs that are partially conserved in the MTases and R.EcoRII.
This may be the first example of aa sequence homology between a MTase specificity (target
recognition) domain and its cognate restriction endonuclease (ENase). It suggests that this
region is important for DNA recognition by R.EcoRII and that the EcoRII ENase and MTase genes
may have evolved from a common progenitor.

<>

<1>Kossykh, V.G., Schlagman, S.L., Hattman, S.
<2>Conserved sequence motif DPPY in region IV of the phage T4 Dam DNA-[N6-adenine]-methyltransferase is important for S-adenosyl-L-methionine binding.
<3>Nucleic Acids Res.
<4>21
<5>3563-3566
<6>1993
<7>Comparison of the deduced amino acid sequences of DNA-[N6-adenine]-methyltransferases has
revealed several conserved regions. All of these enzymes contain a DPPY-motif, or a variant of
it. By site-directed mutagenesis of a cloned T4 dam gene, we have altered the first proline
residue in this motif (located in conserved region IV of the T4 Dam-MTase) to alanine or
threonine. The mutant enzymic forms, P172A and P172T, were overproduced and purified. Kinetic
studies showed that compared to the wild-type (wt) the two mutant enzymic forms had:(i) an
increased (6 and 23-fold, respectively) Km for substrate, S-adenosylmethionine (AdoMet) and an
increased (6 and 23-fold) Ki for product, S-adenosyl-homocysteine (AdoHcy); (ii) a slightly
reduced (1.5 and 3-fold lower) kcat; (iii) a strongly reduced kcat/Km AdoMet (10 and 80-fold);
and (iv) the same Km for substrate DNA. Equilibrium dialysis studies showed that the mutant
enzymes had a reduced (3 and 7-fold lower) Ka for AdoMet; all forms bound two molecules of
AdoMet. Taken together these data indicate that the P172A and P172T alterations resulted
primarily in a reduced affinity for AdoMet. This suggests that the DPPY-motif is important for
AdoMet-binding, and that region IV contains an AdoMet-binding site.

<>

<1>Kossykh, V.G., Schlagman, S.L., Hattman, S.
<2>Conserved sequence motif DPPY in region IV of the phage T4 Dam DNA-[N6-adenine]-methyltransferase is important for S-adenosyl-L-methionine binding.
<3>Nucleic Acids Res.
<4>21
<5>4659-4662
<6>1993
<7>Comparison of the deduced amino acid sequences of DNA-[N6-adenine]-methyltransferases has
revealed several conserved regions. All of these enzymes contain a DPPY [or closely related]
motif. By site-directed mutagenesis of a cloned T4 dam gene, we have altered the first proline
residue in this motif [located in conserved region IV of the T4 Dam-MTase] to alanine or
threonine. The mutant enzymic forms, P172A and P172T, were overproduced and purified. Kinetic
studies showed that compared to the wild-type [wt] the two mutant enzymic forms had: (i) an
increased [5 and 20-fold, respectively] Km for substrate, S-adenosyl-methionine [AdoMet]; (ii)
a slightly reduced [2 and 4-fold lower] kcat; (iii) a strongly reduced kcat/Km AdoMet [10 and
100-fold]; and (iv) almost the same Km for substrate DNA. Equilibrium dialysis studies showed
that the mutant enzymes had a reduced [4 and 9-fold lower] Ka for AdoMet. Taken together these
data indicate that the p172A and P172T alterations resulted primarily in a reduced affinity
for AdoMet. This suggests that the DPPY-motif is important for AdoMet-binding, and that region
IV contains or is part of an AdoMet-binding site.

<>

<1>Kossykh, V.G., Schlagman, S.L., Hattman, S.
<2>Phage T4 DNA [N6-adenine]Methyltransferase.
<3>J. Biol. Chem.
<4>270
<5>14389-14393
<6>1995
<7>The bacteriophage T4 dam gene, encoding the Dam DNA [N6-adenine]methyltransferase (MTase), has
been subcloned into the plasmid expression vector, pJW2. In this construct, designated
pINT4dam, transcription is from the regulatable phage lambda PR and PL promoters, arranged in
tandem. A two-step purification scheme using DEAE-cellulose and phosphocellulose columns in
series, followed by hydroxyapatite chromatography, was developed to purify the enzyme to near
homogeneity. The yield of purified protein was 2 mg/g of cell paste. The MTase has an s20,w of
3.0 S and a Stokes radius of 23 Angstroms and exists in solution as a monomer. The Km for the
methyl donor, S-adenosylmethionine, is 0.1 x 10-6M, and the Km for substrate nonglucosylated,
unmethylated T4 gt-dam- DNA is 1.1 x 10/-12M. The products of DNA methylation,
S-adenosyl-L-homocysteine and methylated DNA, are competitive inhibitors of the reaction; Ki
values of 2.4 x 10/-6M and 4.6 x 10/-12 M, respectively, were observed. T4 Dam methylates the
palindromic tetranucleotide, GATC, designated the canonical sequence. However, at high
MTase:DNA ratios, T4 Dam can methylate some noncanonical sequences belonging to GAY (where Y
represents cytosine or thymine).

<>

<1>Kossykh, V.G., Schlagman, S.L., Hattman, S.
<2>Studies on the function of conserved sequence motifs in the T4 Dam-[N6-adenine] and EcoRII [C5-cytosine] DNA methyltransferases.
<3>Gene
<4>157
<5>125-126
<6>1995
<7>We used site-directed oligodeoxyribonucleotide-mediated mutagenesis and kinetic studies with
purified wild-type (wt) and mutant proteins to evaluate the role of the conserved sequence
motifs in two prokaryotic DNA MTases.  We suggest that: (i) the main role of Pro in the
M.EcoRII PC-motif is to restrict the conformational freedom of Cys and orient it in a manner
essential for catalysis; (ii) in both M.EcoRII and T4 Dam the FXGXG-motif positions AdoMet
with respect to the catalytic site; (iii) the DPPY-motif in T4 Dam (region IV) is important
for AdoMet-binding and may be part of the binding site; and (iv) the RXNXKXXFXXPFK-motif in T4
Dam (region III) is part of the DNA binding/recognition domain.

<>

<1>Kossykh, V.G., Schlagman, S.L., Hattman, S.
<2>Function of Pro-185 in the ProCys of conserved motif IV in the EcoRII [cytosine-C5]-DNA methyltransferase.
<3>FEBS Lett.
<4>370
<5>75-77
<6>1995
<7>ProCys in the conserved sequence motif IV of [cytosine-C5]-DNA methyltransferases is known to
be part of the catalytic site.  The Cys residue is directly involved in forming a covalent
bond with the C6 of the target cytosine.  We have found that substitution of Pro-185 with
either Ala or Ser resulted in a reduced rate of methyl group transfer by the EcoRII DNA
methyltransferase.  In addition, we observed an increase in the Km for substrate
S-adenosyl-L-methionine (AdoMet), but a decrease in the Km for substrate DNA.  This is
reflected in minor changes in kcat/Km for DNA, but in 10- to 100-fold reductions in kcat/Km
for AdoMet.  This suggests that Pro-185 is important to properly orient the activated cytosine
and AdoMet for methyl group transfer by direct interaction with AdoMet and indirectly via the
Cys interaction with cytosine.

<>

<1>Kossykh, V.G., Schlagman, S.L., Hattman, S.
<2>Comparative studies of the phage T2 and T4 DNA (N6-adenine) methyltransferases: Amino acid changes that affect catalytic activity.
<3>J. Bacteriol.
<4>179
<5>3239-3243
<6>1997
<7>The bacteriophage T2 and T4 dam genes code for a DNA (N6-adenine) methyltransferase (Mtase).
Nonglucosylated, hydroxymethylcytosine-containing T2gt- virion DNA has a higher level of
methylation than T4gt- virion DNA does.  To investigate the basis for this difference, we
compared the intracellular enzyme levels following phage infection as well as the in vitro
intrinsic methylation capabilities of purified T2 and T4 Dam Mtases.  Results from Western
blotting (immunoblotting) showed that the same amounts of MTase protein were produced after
infection with T2 and T4.  Kinetic analyses with purified homogeneous enzymes showed that the
two MTases had similar Km values for the methyl donor, S-adenosyl-L-methionine, and for
substrate DNA.  In contrast, they had different kcat values (two-fold higher for T2 Dam
MTase).  We suggest that this difference can account for the ability of T2 Dam to methylate
viral DNA in vivo to a higher level than does T4 Dam.  Since the T2 and T4 MTases differ at
only three amino acid residues (at positions 20 [T4, Ser; T2, Pro], 26 [T4, Asn; T2, Asp], and
188 [T4, Asp; T2, Glu]), we have produced hybrid proteins to determine which residue(s) is
responsible for increased catalytic activity.  The results of these analyses showed that the
residues at positions 20 and 26 are responsible for the different kcat values of the two
MTases for both canonical and noncanonical sites.  Moreover, a single substitution of either
residue 20 or 26 was sufficient to increase the kcat of T4 Dam.

<>

<1>Kostiuk, G., Sasnauskas, G., Tamulaitiene, G., Siksnys, V.
<2>Degenerate sequence recognition by the monomeric restriction enzyme: single mutation converts BcnI into a strand-specific nicking endonuclease.
<3>Nucleic Acids Res.
<4>39
<5>3744-3753
<6>2011
<7>Unlike orthodox Type II restriction endonucleases that are homodimers and interact with the
palindromic 4-8-bp DNA sequences, BcnI is a monomer which has a single active site but cuts
both DNA strands within the 5'-CC downward arrowCGG-3'/3'-GGG downward arrowCC-5' target
site (' downward arrow' designates the cleavage position). Therefore, after cutting the
first strand, the BcnI monomer must re-bind to the target site in the opposite orientation;
but in this case, it runs into a different central base because of the broken symmetry of the
recognition site. Crystal-structure analysis shows that to accept both the C:G and G:C base
pairs at the center of its target site, BcnI employs two symmetrically positioned histidines
H77 and H219 that presumably change their protonation state depending on the binding mode. We
show here that a single mutation of BcnI H77 or H219 residues restricts the cleavage activity
of the enzyme to either the 5'-CCCGG-3' or the 5'-CCGGG-3' strand, thereby converting BcnI
into a strand-specific nicking endonuclease. This is a novel approach for engineering of
monomeric restriction enzymes into strand-specific nucleases.

<>

<1>Kostka, J.E., Green, S.J., Rishishwar, L., Prakash, O., Katz, L.S., Marino-Ramirez, L., Jordan, I.K., Munk, C., Ivanova, N., Mikhailova, N., Watson, D.B., Brown, S.D., Palumbo, A.V., Brooks, S.C.
<2>Genome sequences for six rhodanobacter strains, isolated from soils and the terrestrial subsurface, with variable denitrification capabilities.
<3>J. Bacteriol.
<4>194
<5>4461-4462
<6>2012
<7>We report the first genome sequences for six strains of Rhodanobacter species isolated from a
variety of soil and subsurface environments. Three of these
strains are capable of complete denitrification and three others are not.
However, all six strains contain most of the genes required for the respiration
of nitrate to gaseous nitrogen. The nondenitrifying members of the genus lack
only the gene for nitrate reduction, the first step in the full denitrification
pathway. The data suggest that the environmental role of bacteria from the genus
Rhodanobacter should be reevaluated.

<>

<1>Kostrewa, D., Winkler, F.K.
<2>Mg2+ binding to the active site of EcoRV endonuclease: A crystallographic study of complexes with substrate and product DNA at 2 Angstrom resolution.
<3>Biochemistry
<4>34
<5>683-696
<6>1995
<7>The type II restriction endonuclease EcoRV was crystallized as a complex with the substrate
DNA undecamer AAAGATATCTT. These crystals diffract to much better resolution (2 Angstrom) than
was the case for the previously reported complex with the decamer GGGATATCCC. The crystal
structure contains one dimer complex in the asymmetric unit and was solved by molecular
replacement. The same kinked DNA conformation characteristic for enzyme-bound cognate DNA is
observed. Crystals, soaked with Mg2+, show the essential cofactor bound at only one active
site of the dimer, and the DNA is not cleaved. The Mg2+ has one oxygen from the scissile
phosphodiester group and two carboxylate oxygens, one from Asp74 and one from Asp90, in its
octahedral ligand sphere. The scissile phosphodiester group is pulled by 1 Angstrom toward the
Mg2+. After substrate cleavage in solution, isomorphous crystals containing the
enzyme--product--Mg2+ complex were obtained. In this structure, each of the 5'-phosphate
groups is bound to two Mg2+. The kinked DNA conformation is essentially maintained, but the
two central adenines, 3' to the cleavage sites, form an unusual cross-strand base stacking.
The structures have been refined to R factors of 0.16 at 2.1-2.0 Angstrom resolution
maintaining very good stereochemistry. On the basis of these structures and inspired by recent
kinetic data, we have constructed a transition state model with two metals bound to the
scissile phosphorane group.

<>

<1>Kostriken, R., Strathern, J.N., Klar, A.J.S., Hicks, J.B., Heffron, F.
<2>A site-specific endonuclease essential for mating-type switching in Saccharomyces cerevisiae.
<3>Cell
<4>35
<5>167-174
<6>1983
<7>We have detected two site-specific endonucleases in strains of Saccharomyces cerevisiae. One
endonuclease, which we call YZ endo, is present only in yeast strains that are undergoing
mating-type interconversion. The site at which YZ endo cleaves corresponds to the in vivo
double-strand break occuring at the mating-type locus in yeast undergoing mating-type
interconversion. YZ endo generates a site-specific double-strand break having 4-base 3'
extensions terminating in 3' hydroxyl groups. The site of cleavage occurs in the Z1 region
near the YZ junction of the mating-type locus. Mutant mating-type loci known to decrease the
frequency of mating-type interconversion are correspondingly poor substrates for YZ endo in
vitro. In vitro analysis of a number of such altered recognition sites has delimited the
sequences required for cleavage. The molecular genetics of mating-type interconversion is
discussed in the context of this endonucleolytic activity. The second endonuclease, which we
refer to as SceII, is present in all strains of S. cerevisiae we have examined. The cleavage
site of SceII has been determined and proves to be unrelated to the cleavage site of YZ endo.

<>

<1>Kosykh, V.G.
<2>Escherichia coli K-12 plasmid R245 controlling EcoRII restriction and DNA modification.
<3>Dokl. Akad. Nauk.
<4>238
<5>1227-1230
<6>1978
<7>
<>

<1>Kosykh, V.G., Buryanov, Y.I., Baev, A.A.
<2>Strain Escherichia coli containing restrictase and methylase created by conjugationally transferring plasmid into cells of Escherichia coli to exclude ferment impurities.
<3>Soviet Patent Office
<4>SU 941423 A
<5>
<6>1982
<7>Escherichia coli microbial strain containing plasmid with genes for the EcoRII restrictase and
methylase are created by conjugationally transferring this plasmid into Escherichia coli
cells. This method is improved by using strain Escherichia coli B 234 and plasmids N 3 or R
245.   By this method, interfering ferment impurities are excluded. The Escherichia coli B 834
appears in the form of rods; dimensions (1.1-1.5)x(2.0-6.0) microns; mobile with flagellae;
gram-negative. They do not produce spores. The R 245 variety is resistant to antibiotic
tetra-cycline; the N 3 variety is resistant to tetracycline, streptomycin and sulphanilamide.

<>

<1>Kosykh, V.G., Buryanov, Y.I., Baev, A.A.
<2>Recombinant plasmid DNAs, containing restrictase and methylase genes, prepared using specified donor and recipient plasmid deoxyribonucleic acids and medium containing ampicillin and sulphanilamide antibiotics.
<3>Soviet Patent Office
<4>SU 852939 A
<5>
<6>1981
<7>Production of plasmid recombinant DNA containing genes of EcoRII restrictase and methylase,
useful in genetic engineering, includes treatment of donor's and recipients DNA with
restriction endonuclease and combining the resulting fragments by means of poly-
nucleotide-ligase enzyme. This is followed by the transformation of Escherichia coli microbial
strain cells with recombinant plasmid DNA, collecting the resulting transformation products
by means of a selective microbiological medium containing antibiotics. In order to improve
super-synthesis of restrictase and methylase enzymes, DNA fragments from the plasmids N3 and
PZ2O3 are used as donor and recipient DNA respectively and penicillin and sulphanilamide are
used as antibiotics.

<>

<1>Kosykh, V.G., Buryanov, Y.I., Baev, A.A.
<2>Escherichia coli strain culture useful in endonuclease enzyme activity evaluation during production and purification.
<3>Soviet Patent Office
<4>SU 918304 A
<5>
<6>1982
<7>Microbial strain Escherichia coli B 834 (r(b)m(b))/PBR322 contains a plasmid which is useful
as a test-substrate in the determination of endonuclease EcoRII enzymatic activity during its
separation and purification The latter is useful in molecular biology and genetic engineering.
The microbial strain is obtained from E. Coli B 834 strain containing plasmid PBR 322. This
strain does not contain DNA-cytosine methylase. The above strain can be cultured in e.g.
meat-peptone broth at pH 6.8-7.5 and 4-45 degrees C. It can utilise (as carbon source)
alcohols, organic acids, mono-saccharides etc. As nitrogen source it can utilise mineral salts
in ammonium or nitrate form or peptone, aminoacids etc. as organic source of nitrogen. It is
resistant to antibiotics such as tetracycline.

<>

<1>Kosykh, V.G., Buryanov, Y.I., Baev, A.A.
<2>Cloning the genes of EcoRII restrictase and methylase.
<3>Dokl. Akad. Nauk.
<4>247
<5>1269-1271
<6>1979
<7>
<>

<1>Kosykh, V.G., Buryanov, Y.I., Bayev, A.A.
<2>Molecular Cloning of EcoRII Endonuclease and Methylase Genes.
<3>Mol. Gen. Genet.
<4>178
<5>717-718
<6>1980
<7>The genes for restriction-modification system EcoRII have been cloned from
plasmid N3 DNA using RSF2124 as a vector plasmid.  The hybrid plasmids
designated pFK321 and pFK322 contained a 5.8 megadaltons EcoRI - fragment
derived from N3 DNA including the genes for restriction-modification system
EcoRII and a gene for resistance to sulfanilamide.

<>

<1>Kosykh, V.G., Filipchenkova, L.P., Mierinya, A.A., Buryanov, Y.I., Baev, A.A., Ansberga, S.E., Zilbere, A.M.
<2>Purification of the EcoRII restriction enzyme and DNA methylase.
<3>Soviet Patent Office
<4>SU 1262952
<5>
<6>1994
<7>
<>

<1>Kosykh, V.G., Glinskaite, I.V., Buryanov, J.I., Baev, A.A.
<2>Mapping the EcoRII restrictase and methylase genes on recombinant plasmids.
<3>Dokl. Akad. Nauk.
<4>265
<5>727-730
<6>1982
<7>None

<>

<1>Kosykh, V.G., Mantsygin, Y.A., Svyatukhina, N.V., Vitenene, I.V., Buryanov, Y.I.
<2>Purification of the restriction endonuclease EcoRII using monoclonal antibodies.
<3>Biokhimiia
<4>53
<5>1474-1478
<6>1988
<7>Hybridomas producing monoclonal antibodies to the restriction endonuclease
EcoRII were produced after immunization of two BALB/c mice with a preparation
of the homogeneous enzyme.  The IgG were obtained from ascites fluid, purified,
and covalently bonded to CNBr-activated Sepharose 4B.  The immunosorbent
obtained was used for the isolation of the restriction endonuclease EcoRII.
The isolated restriction endonuclease EcoRII gave one band in polyacrylamide
gel electrophoresis with sodium sulfate.  The enzyme was obtained in a good
yield with high specific activity.

<>

<1>Kosykh, V.G., Mierenya, A.A., Zeidaka, A.A., Buryanov, Y.I., Bayev, A.A., Shprunka, I.K.
<2>Production of EcoRII methylase.
<3>Soviet Patent Office
<4>SU 1311253
<5>
<6>1995
<7>
<>

<1>Kosykh, V.G., Puntezhis, S.A., Buryanov, Y.I., Baev, A.A.
<2>Isolation, purification and properties of restriction endonuclease EcoRII.
<3>Biokhimiia
<4>47
<5>619-625
<6>1982
<7>The restriction endonuclease EcoRII was isolated and purified to homogeneity.  The isolation
procedure involved the use of the E. coli strain B834/pSK323, containing the recombinant
plasmid pSK323 which provides for the overproduction of EcoRII enzymes.  Data from gel
filtration and SDS electrophoresis suggest that the restriction endonuclease EcoRII is a
protein made up of two subunits, each with molecular weight of 44000.

<>

<1>Kosykh, V.G., Repik, A.V., Kaliman, A.V., Buryanov, Y.I., Baev, A.A.
<2>Primary structure of the restriction endonuclease EcoRII gene.
<3>Dokl. Akad. Nauk.
<4>308
<5>1497-1499
<6>1989
<7>None

<>

<1>Kosykh, V.G., Solonin, A.S., Buryanov, Y.I., Bayev, A.A.
<2>Overproduction of the EcoRII endonuclease and methylase by Escherichia coli strains carrying recombinant plasmids constructed in vitro.
<3>Biochim. Biophys. Acta
<4>655
<5>102-106
<6>1981
<7>Recombinant DNA molecules were constructed from the plasmid pIL203 and the
EcoRI-fragment of N3 plasmid containing EcoRII endonuclease and methylase genes
and also a gene for resistance to sulfanilamide.  The pIL203 plasmid, used as a
vector, consisted of the BamHI-EcoRI-fragment of the plasmid pBR322 conferring
resistance to ampicillin and the BamHI-EcoRI-fragment of lambda phage
containing promoters, a thermosensitive mutation in the cI gene and a
suppressible amber mutation in the cro gene.
Ampicillin-sulfanilamide-resistant clones were selected and tested for their
restriction and modification phenotype.  The recombinant plasmid DNA, isolated
from ApRSuR-resistant clones, which restricted and modified phage lambda imm21
with EcoRII specificity, had the EcoRI-fragment with EcoRII genes in a single
orientation.  The recombinant plasmid pSK323 was transferred into E. coli
strains with su-, su1, su2 or su3 phenotypes.  The synthesis of products of
EcoRII genes by these strains grown at 37C is increased by 10-50-fold.

<>

<1>Kosykh, V.G., Vitenene, I.V., Buryanov, Y.A.
<2>Plasmid recombinant DNA - codes restrictase and methylase EcoRII.
<3>Soviet Patent Office
<4>SU 1453896 A
<5>
<6>1990
<7>Escherichia coli VKM CR 288D strain is used as host in producing the restrictase and methylase
EcoRII at a higher level than the known strain, for the plasmid recombinant DNA coding the
above enzymes. The recombinant DNA pVK11 containing the DNA fragment with the complete EcoRII
restrictase and methylase genes and the left promoter of phage lambda is constructed, ensuring
the efficient expression of the EcoRII genes.

<>

<1>Kotani, H., Kato, I., Kimizuka, F.
<2>New restriction enzyme.
<3>European Patent Office
<4>EP 0464990 B
<5>
<6>1995
<7>*
A new restriction capable of recognizing the nucleotide sequence in double stranded DNA

    5'-CCTGCAGG-3'
    3'-GGACGTCC-5'

and specifically cleaving it is disclosed. The enzyme may be made cultivating a strain
of the genus Streptomyces (FERM BP-3028) capable of producing it.


<>

<1>Kotani, H., Nomura, Y., Kawashima, Y., Sagawa, H., Takagi, M., Kita, A., Ito, H., Kato, I.
<2>Sse8387I, a new type-II restriction endonuclease that recognizes the octanucleotide sequence 5'-CCTGCAGG-3'.
<3>Nucleic Acids Res.
<4>18
<5>5637-5640
<6>1990
<7>A type II restriction endonuclease designated Sse8387I was partially purified
from Streptomyces sp. 8387.  This enzyme cleaved adenovirus 2 DNA at three
sites, lambda phage DNA at five sites, and pUC18 and M13mp18 RF DNA at one site
each, but did not cleave the DNAs from pBR322, SV40, or PhiX174.  Sse8387I
recognized the octanucleotide sequence 5'-CCTGCA^GG-3', cleaving where shown by
the arrow.  Sse8387I is the first restriction endonuclease to be reported that
recognizes an octanucleotide sequence consisting of all four nucleotides,
G,A,T, and C.  The frequency of occurrence of Sse83787I sites within sequenced
regions of primate genomes was 2.4 times that of NotI sites.

<>

<1>Kothari, V.V., Kothari, R.K., Kothari, C.R., Bhatt, V.D., Nathani, N.M., Koringa, P.G., Joshi, C.G., Vyas, B.R.
<2>Genome Sequence of Salt-Tolerant Bacillus safensis Strain VK, Isolated from Saline Desert Area of Gujarat, India.
<3>Genome Announcements
<4>2
<5>e00337-14
<6>2014
<7>
<>

<1>Kothari, V.V., Kothari, R.K., Kothari, C.R., Bhatt, V.D., Nathani, N.M., Koringa, P.G., Joshi, C.G., Vyas, B.R.
<2>Genome Sequence of Salt-Tolerant Bacillus safensis Strain VK, Isolated from Saline Desert Area of Gujarat, India.
<3>Genome Announcements
<4>1
<5>e00671-13
<6>2013
<7>Bacillus safensis strain VK was isolated from the rhizosphere of a cumin plant growing in the
saline desert of Radhanpar, Gujarat, India. Here, we provide the
3.68-Mb draft genome sequence of B. safensis VK, which might provide information
about the salt tolerance and genes encoding enzymes for the strain's plant
growth-promoting potential.

<>

<1>Koton, Y., Eghbaria, S., Gordon, M., Chalifa-Caspi, V., Bisharat, N.
<2>Draft Genome Sequence of Fish Pathogenic Vibrio vulnificus Biotype 2.
<3>Genome Announcements
<4>2
<5>e01224-14
<6>2014
<7>Vibrio vulnificus is a marine pathogen capable of causing severe soft tissue infections and
septicemia in humans. V. vulnificus biotype 2 is the etiological
agent of fish vibriosis. We describe here the first draft genome sequence of V.
vulnificus biotype 2, strain ES-7601, isolated from an infected eel in Japan.

<>

<1>Kotova, V.Y., Zavilgelskii, G.B., Belogurov, A.A.
<2>Weakening of the type-I restriction in the presence of plasmids of the incI group. General characteristics and molecular cloning of ard gene.
<3>Mol. Biol. (Mosk)
<4>22
<5>270-276
<6>1988
<7>Plasmids of the incI group possess the ability to weaken the effect of type-I
restrictases on unmodified DNA.  The ard locus, which is responsible for weakening of
type I restriction, is located in plasmid ColIb-P9(incIa) in the region of the Sal GI-C
fragment.  Cloning was conducted of the ard locus in multicopy vector pBR322.  The ard
gene specifically weakens type-I restriction (EcoK, EcoB, EcoA, EcoD) and does not
influence restrictase systems of type II (EcoRI) and III (EcoPI).  Activity of the ard gene
does not depend on bacterial genes recA, lexA, recBC, recF.  Product of the ard gene does
not influence the process of methylation of DNA by type-I fragments and is not a specific
methylase.

<>

<1>Koudan, E.V., Brevnov, M.G., Subach, O.M., Rechkoblit, O.A., Bujnicki, J.M., Gromova, E.S.
<2>Probing of contacts between EcoRII DNA methyltransferase and DNA with the use of substrate analogs and molecular modeling.
<3>Mol. Biol. (Mosk)
<4>41
<5>806-819
<6>2007
<7>The molecular mechanisms of DNA recognition and modification by EcoRII DNA methyltransferase
(M.EcoRII) were studied using 14-mer substrate
analogs containing 2-aminopurine or 1',2'-dideoxy-D-ribofuranose in the
M.EcoRII recognition site. The efficiency of DNA binding and
methylation depended on the position of a modified nucleoside residue
in the recognition site. A structural model of M.EcoRII in complex with
substrate DNA and the cofactor analog S-adenosyl-L-homocysteine
(AdoHcy) was constructed using the available crystal structures of
M.Hha and M.HaeIII and the recent Frankenstein's monster approach. The
amino acid residues interacting with DNA were predicted based on the
model. In addition, theoretical and experimental findings made it
possible to predict the groups of atoms of the heterocyclic bases of
the M.EcoRII recognition site that are presumably involved in the
interactions with the enzyme.

<>

<1>Koudan, E.V., Bujnicki, J.M., Gromova, E.S.
<2>Homology modeling of the CG-specific DNA methyltransferase SssI and its complexes with DNA and AdoHcy.
<3>J. Biomol. Struct. Dyn.
<4>22
<5>339-345
<6>2004
<7>Prokaryotic DNA methyltransferase M.SssI recognizes and methylates C5 position of the cytosine
residue within the CG dinucleotides in DNA. It
is an excellent model for studying the mechanism of interaction between
CG-specific eukaryotic methyltransferases and DNA. We have built a
structural model of M.SssI in complex with the substrate DNA and its
analogues as well as the cofactor analogue S-adenosyl-L-homocysteine
(AdoHcy) using the previously solved structures of M.HhaI and M.HaeIII
as templates. The model was constructed according to the recently
developed "FRankenstein's monster" approach. Based on the model, amino
acid residues taking part in cofactor binding, target recognition and
catalysis were predicted. We also modeled covalent modification of the
DNA substrate and studied its influence on protein-DNA interactions.

<>

<1>Koudan, E.V., Subach, O.M., Korshunova, G.A., Romanova, E.A., Eritja, R., Gromova, E.S.
<2>DNA duplexes containing photoactive derivatives of 2'-deoxyuridine as photocrosslinking probes for EcoRII DNA methyltransferase-substrate interaction.
<3>J. Biomol. Struct. Dyn.
<4>20
<5>421-428
<6>2002
<7>EcoRII DNA methyltransferase (M.EcoRII) recognizes the DNA sequence 5'.CC*T/AGG.3' and
catalyzes the transfer of the methyl group from S-adenosyl-L-methionine to the C5 position of
the inner cytosine residue (C*). We obtained several DNA duplexes containing photoactive
5-iodo-2'-deoxyuridine (i(5)dU) or
5-[4-(3-(trifluoromethyl)-3H-diazirin-3-yl)phenyl]-2'-deoxyuridine (Tfmdp-dU) to characterize
regions of M.EcoRII involved in DNA binding and to investigate the DNA double helix
conformational changes that take place during methylation. The efficiencies of methylation,
DNA binding affinities and M.EcoRII-DNA photocrosslinking yields strongly depend on the type
of modification and its location within the EcoRII recognition site. The data obtained agree
with the flipping of the target cytosine out of the DNA double helix for catalysis. To probe
regions of M.EcoRII involved in DNA binding, covalent conjugates M.EcoRII-DNA were cleaved by
cyanogen bromide followed by analysis of the oligonucleotide-peptides obtained. DNA duplexes
containing i(5)dU or Tfmdp-dU at the central position of the recognition site, or instead of
the target cytosine were crosslinked to the Gly(268)-Met(391) region of the EcoRII methylase.
Amino acid residues from this region may take part both in substrate recognition and
stabilization of the extrahelical target cytosine residue.

<>

<1>Koufopanou, V., Burt, A.
<2>Degeneration and domestication of a selfish gene in yeast: Molecular evolution versus site-directed mutagenesis.
<3>Mol. Biol. Evol.
<4>22
<5>1535-1538
<6>2005
<7>VDE is a homing endonuclease gene in yeasts with an unusual evolutionary history including
horizontal transmission, degeneration, and domestication into the mating-type switching locus
HO. We investigate here the effects of these features on its molecular evolution. In addition,
we correlate rates of evolution with results from site-directed mutagenesis studies.
Functional elements have lower rates of evolution than degenerate ones and higher conservation
at functionally important sites. However, functionally important and unimportant sites are
equally likely to have been involved in the evolution of new function during the domestication
of VDE into HO. The domestication event also indicates that VDE has been lost in some species
and that VDE has been present in yeasts for more than 50 Myr.

<>

<1>Koufopanou, V., Goddard, M.R., Burt, A.
<2>Adaptation for horizontal transfer in a homing endonuclease.
<3>Mol. Biol. Evol.
<4>19
<5>239-246
<6>2002
<7>Selfish genes of no function other than self-propagation are susceptible to degeneration if
they become fixed in a population, and regular transfer to new species may be the only means
for their long-term persistence. To test this idea we surveyed 24 species of yeast for VDE, a
nuclear, intein-associated homing endonuclease gene (HEG) originally discovered in
Saccharomyces cerevisiae. Phylogenetic analyses show that horizontal transmission has been a
regular occurrence in its evolutionary history. Moreover, VDE appears to be specifically
adapted for horizontal transmission. Its 31-bp recognition sequence is an unusually
well-conserved region in an unusually well-conserved gene. In addition, the nine nucleotide
sites most critical for homing are also unusually well conserved. Such adaptation for
horizontal transmission presumably arose as a consequence of selection, both among HEGs at
different locations in the genome and among variants at the same location. The frequency of
horizontal transmission must therefore be a key feature constraining the distribution and
abundance of these genes.

<>

<1>Koukalova, B., Kuhrova, V., Reich, J.
<2>Protection of nonmodified phage lambda against EcoK restriction mediated by recA protein.
<3>Folia Microbiol. (Praha)
<4>30
<5>17-24
<6>1985
<7>A study was conducted to establish whether the EcoK-specific restriction, which is alleviated
in E. coli cells after UV induction of the SOS response (Day 1977), is also alleviated under
the influence of an increased level of recA protein without induction of other SOS functions.
The host cells used were E. coli K-12, strain AB2497, and its derivatives; the nonmodified
phage lambda was a mutant b2b5(vir).  An increase of the recA protein level was induced using
the plasmid pX02, which is a recombinant of pBR322 carrying the recA gene of E. coli.
AB2497(pX02) cells were found to exhibit a lower level of restriction than those without
plasmid.  The results indicate that the recA protein protects phage DNA during the process of
restriction.  A further factor affecting restriction is the growth phase of the culture of the
restricting host: cells in the late stationary phase exhibit lower restriction than those in
the exponential phase of growth.  By a combination of these two factors (presence of plasmid
pX02 and stationary growth phase) one can reduce the restriction of nonmodified phage about
300 times.

<>

<1>Kouvelis, V.N., Davenport, K.W., Brettin, T.S., Bruce, D., Detter, C., Han, C., Nolan, M., Tapia, R., Damoulaki, A., Kyrpides, N.C., Typas, M.A., Pappas, K.M.
<2>Genome sequence of the ethanol-producing Zymomonas mobilis subsp. pomaceae lectotype strain ATCC 29192.
<3>J. Bacteriol.
<4>193
<5>5049-5050
<6>2011
<7>Zymomonas mobilis is an alphaproteobacterium studied for bioethanol production. Different
strains of this organism have been hitherto
sequenced; they all belong to the Z. mobilis subsp. mobilis taxon. Here we
report the finished and annotated genome sequence of strain ATCC 29192, a
cider spoiling agent isolated in the United Kingdom. ATCC 29192 is the
lectotype of the second best characterized subspecies of Z. mobilis, Z.
mobilis subsp. pomaceae. The nucleotide sequence of ATCC 29192 is more
deviant from that of mobilis representatives, which justifies its distinct
taxonomic positioning and proves particularly useful for comparative and
functional genomics analyses.

<>

<1>Kouvelis, V.N., Saunders, E., Brettin, T.S., Bruce, D., Detter, C., Han, C., Typas, M.A., Pappas, K.M.
<2>Complete genome sequence of ethanol producer Zymomonas mobilis NCIMB 11163.
<3>J. Bacteriol.
<4>191
<5>7140-7141
<6>2009
<7>Zymomonas mobilis is an ethanol producing alpha-proteobacterium currently considered as major
candidate organism for bioethanol production. Here we report the finished and annotated genome
sequence of the Z. mobilis subsp. mobilis strain NCIMB 11163, a British ale infecting isolate.
This is the first Z. mobilis strain whose genome, chromosomal and plasmid, is presented in its
entirety.

<>

<1>Kouvelis, V.N., Teshima, H., Bruce, D., Detter, C., Tapia, R., Han, C., Tampakopoulou, V.O., Goodwin, L., Woyke, T., Kyrpides, N.C., Typas, M.A., Pappas, K.M.
<2>Finished Genome of Zymomonas mobilis subsp. mobilis Strain CP4, an Applied Ethanol Producer.
<3>Genome Announcements
<4>2
<5>e00845-13
<6>2014
<7>Zymomonas mobilis subsp. mobilis is one of the most rigorous ethanol-producing organisms known
to date, considered by many to be the prokaryotic alternative to
yeast. The two most applied Z. mobilis subsp. mobilis strains, ZM4 and CP4,
derive from Recife, Brazil, and have been isolated from sugarcane fermentations.
Of these, ZM4 was the first Z. mobilis representative strain to be sequenced and
analyzed. Here, we report the finishing of the genome sequence of strain CP4,
which is highly similar but not identical to that of ZM4.

<>

<1>Kouzminova, E., Selker, E.U.
<2>dim-2 encodes a DNA methyltransferase responsible for all known cytosine methylation in Neurospora.
<3>EMBO J.
<4>20
<5>4309-4323
<6>2001
<7>To understand better the control of DNA methylation, we cloned and characterized the dim-2
gene of Neurospora crassa, the only eukaryotic gene currently known in which mutations appear
to eliminate DNA methylation. The dim-2 gene is responsible for methylation in both
symmetrical and asymmetrical sites. We mapped dim-2 between wc-1 and un-10 on linkage group
(LG) VIIR and identified the gene by RFLP mapping and genetic complementation. Dim-2 encodes a
1454 amino acid protein including a C-terminal domain homologous to known DNA
methyltransferases (MTases) and a novel N-terminal domain. Neither a deletion that removed the
first 186 amino acids of the protein nor a mutation in a putative nucleotide binding site
abolished function, but a single amino acid substitution in the predicted catalytic site did.
Tests for repeat-induced point mutation (RIP) indicated that dim-2 does not play a role in
this process, i.e. duplicated sequences are mutated in dim-2 strains, as usual, but the
mutated sequences are not methylated, unlike the situation in dim-2(+) strains. We conclude
that dim-2 encodes an MTase that is responsible for all DNA methylation in vegetative tissues
of Neurospora.

<>

<1>Kovalenko, N.A., Sokolov, N.N.
<2>On the restriction endonucleases immobilization.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>13-15
<6>1988
<7>The search for optimal variants of restriction endonuclease immobilization was
begun recently.  For some enzymes immobilization was successful due to the
presence of covalent bonds on CNBr-sepharose (EcoRI, BamHI, HindIII, TagI,
PaeI, SalI, PvuII).  For the enzymes EcoRI, BamHI and HindIII it was due to
hydrophobic interaction with triethyl-agarose (triethyl-triphenylmethane).  The
high yield (up to 80%) of enzymatic activity has been obtained for small number
of restriction endonucleases.  In the experiments of several aminoacid residues
modification and immobilization of restriction endonucleases the participation
of lysine, arginine, glutamic acid and SH- or S-S-groups in the catalysis and
(or) binding of these enzymes with DNA has been shown.  The restriction
endonuclease immobilization experiments and research into the enzyme's active
centre enrich each other and are very interesting for their use in molecular
biology and deepening our knowledge of protein-nucleic interactions.

<>

<1>Kovalenko, N.A., Sokolov, N.N., Chercasova, T.A., Vonsky, V.E., Leikin, Y.A.
<2>Immobilization of restriction endonucleases EcoRI, PaeI and LplI.
<3>Bioorg. Khim.
<4>18
<5>210-216
<6>1992
<7>In search for sorbents (silica gels, styrene-divinylbenzene copolymers),for immobilization of
some restriction endonucleases, derivatives of trityl-containing silochroms are shown to bind
EcoRI, Pae I and LplI endonuclease with the retention of 10-20, 60-70 and 40-60% activity,
respectively. The immobilized restriction endonucleases have unchanged substrate specificity,
can be used several times and are stable during storage. Tritylaminopropylsilochrom is
suggested to be the sorbent of choice.

<>

<1>Kovalevskaya, N.P., Ivanov, L.Y., Zheleznaya, A., Matvienko, N.I.
<2>Isolation and properties of the restriction endonuclease BstBSI from the thermophilic soil bacterium Bacillus stearothermophilus BS.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>3
<5>22-25
<6>1993
<7>The discovery at the beginning of the 1970s of site-specific endodeoxyribonucleases, known
under the name of "restriction endonucleases", which cleave DNA into strictly determined
fragments, served as the basis for the creation of a number of fundamentally new approaches to
the analysis of nucleic acid structure and was one of the decisive prerequisites for the birth
of genetic engineering. These enzymes, which interact highly specifically with DNA, are also
of great interest in the study of the mechanisms of DNA-protein interactions. At present,
despite the large assortment of site-specific endonucleases already available, the search for
them is continuing. The main stimulus for continuing investigations, as before, is the effort
to supplement the arsenal with enzymes with various substrate specificities and thereby to
expand the possibilities for experimenters.  In the course of the study of the distribution of
restriction-modification enzymes in various taxonomic groups of microorganisms, we discovered
several thermophilic strains of the genus Bacillus that produce specific endonucleases. In
particular, we reported on a new site-specific endonuclease BstBSI from a soil isolate of B.
stearothermophilus BS. This work presents data on the purification and characterization of the
endonuclease BstBSI.

<>

<1>Kovalevskaya, N.P., Ivanov, L.Y., Zheleznaya, L.A., Matvienko, N.I.
<2>BstBSI, a restriction endonuclease from Bacillus stearothermophilus BS which recognizes 5'GTATAC3'.
<3>Nucleic Acids Res.
<4>19
<5>4296
<6>1991
<7>BstBSI, a type II restriction endonuclease, has been isolated from Bacillus stearothermophilus
BS.  BstBSI, an isoschizomer of SnaI, recognizes the six base palindromic sequence and cleaves
it as indicated by arrows:
5'GTA^TAC3'
3'CAT^ATG5'.
BstBSI recognizes three sites on lambda DNA.  Double digestion mapping with BstBSI and EcoRI,
SmaI, KpnI, MluI and PvuII showed that BstBSI sites were roughly localized to the genome map
positions 15200, 18900, 19500.  Examination of the sequences of the lambda DNA showed that the
sequence GTATAC was present in all of these positions.  The cleavage of the T7 DNA results in
7 visible fragments of more than 1 kb length (upper band is a doublet) of expected molecular
weights.  To identify the cleavage site within the recognition sequence, the recombinant phage
M13tg130 with KpnI-SmaI fragment with coordinates 18556-19397 of lambda DNA was constructed.
It has the BstBSI recognition site nearby the KpnI site.  The cleavage site was determined by
cleavage of a primed-synthesis reaction.  Cleavage product resulted in a single band
comigrated with A in the middle of the recognition sequence.  An addition of T4 DNA polymerase
did not change the position of the band.  These results indicate that the BstBSI cleaves the
recognition sequence in the middle and produces blunt end fragments.  The molecular weight of
the native enzyme determined by gel filtration method is approximately 80000.  The crude
extract contained more than 50000 units per gram of wet cells.

<>

<1>Kovalevskaya, N.P., Zelinskaya, N.V., Zheleznaya, L.A., Matvienko, N.I.
<2>Isolation and properties of the site-specific endonuclease BspTS514I from thermophilic bacterium Bacillus species TS514.
<3>Bioorg. Khim.
<4>19
<5>1073-1076
<6>1993
<7>New site-specific endonucleases BspBS31I, BstBS32I, BspIS4I, BstTS5I, BspTS514I were isolated
from five thermophilic soil bacteria Bacillus sp. BS31, B. stearothermophilus BS32, Bacillus
sp. IS4, B. stearothermophilus TS5, Bacillus sp. TS514. The enzymes are isoschizomers of the
restriction endonuclease BbvII. Endonuclease BspTS514I was obtained pure from interfering
contaminations by two consecutive chromatographies on blue agarose and hydroxyapatite. The
enzyme exhibits a maximal activity at 55oC in 10 mM tris-HCl (pH 9.2), 10 mM MgCl2 and 50 mM
NaCl.

<>

<1>Kovalevskaya, N.P., Zhelenznaya, L.A., Zelinskaya, N.V., Matvienko, N.I.
<2>A new thermophilic strain Bacillus coagulans, producing the site-specific endonuclease BcoKI.
<3>Mikrobiologiia
<4>63
<5>235-238
<6>1994
<7>The strain, producing the new site-specific endonuclease BcoKI has been found during the
screening of thermophilic bacteria isolated from tobacco. A phenotype characteristic of the
strain is given. It has been identified as a new strain Bacillus coagulans. BcoKI, a class-IIS
restriction endonuclease has been obtained by three consecutive chromatographies on blue
agarose, hydroxyapatite and heparin-Sepharose. BcoKI, an isoschizomer of Ksp6321, recognizes
the six base non-palindromic sequence 5'CTCTTC3' and cleaves one nucleotide 3' of the 3'
cytosine on this strand and four nucleotides 5' of the 5' guanine on the opposite strand to
generate a three base 5' overhang.

<>

<1>Kovalevskaya, N.P., Zheleznaya, L.A., Matvienko, N.I.
<2>BspLS21, a new site-specific endonuclease from thermophilic bacterium Bacillus species LS2.
<3>Bioorg. Khim.
<4>18
<5>1473-1477
<6>1992
<7>A new restriction endonuclease BspLS2I was isolated from the thermophilic bacterium Bacillus
species LS2 and purified by blue sepharose and hydroxyapatite chromatographies. The enzyme is
an isoschizomer of SduI from Streptococcus durans. BspLS2I recognizes the sequence
5'G(G/A/T)GC(C/T/A)^C3' on double-stranded DNA and cleaves it is indicated by the arrow to
yield sticky-ended DNA fragments. Maximum catalytic activity of the endonuclease was found in
10 mM Tris-HCl (pH7.9) in the presence of 15-30 mM MgCl2 at 50oC. The phage T4 glucosylated
DNA is not cleaved by the enzyme.

<>

<1>Kovalic, D.K., Andersen, S.E., Byrum, J.R., Conner, T.W., Masucci, J.D., Zhou, Y.
<2>Nucleic acid molecules and other molecules associated with plants and uses thereof for plant improvement.
<3>US Patent Office
<4>US 7214786 A
<5>
<6>2007
<7>Recombinant polynucleotides and recombinant polypeptides useful for improvement of plants are
provided.  The disclosed recombinant polynucleotides and recombinant polypeptides find use in
production of transgenic plants to produce plants having improved properties.

<>

<1>Kovall, R.A., Matthews, B.W.
<2>Structural, functional, and evolutionary relationships between lambda-exonuclease and the type II restriction endonucleases.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>7893-7897
<6>1998
<7>Lambda-exonuclease participates in DNA recombination and repair.  It binds a free end of
double-stranded DNA and degrades one stand in the 5' to 3' direction.  The primary sequence
does not appear to be related to any other protein, but the crystal structure shows part of
lambda-exonuclease to be similar to the type II restriction endonucleases PvuII and EcoRV.
There is also a weaker correspondence with EcoRI, BamHI, and Cfr10I.  The structure
comparisons not only suggest that these enzymes all share a similar catalytic mechanism and a
common structural ancestor but also provide strong evidence that the toroidal structure of
lambda-exonuclease encircles its DNA substrate during hydrolysis.

<>

<1>Kovall, R.A., Matthews, B.W.
<2>Type II restriction endonucleases: structural, functional and evolutionary relationships.
<3>Curr. Opin. Chem. Biol.
<4>3
<5>578-583
<6>1999
<7>Type II restriction endonucleases are a paradigm for site-specific cleavage of DNA. Recent
structural analyses, in particular in the presence of various divalent metals, have shed new
insight into the mechanisms of catalysis. In addition, during this past year the crystal
structure determinations of MutH, lambda-exonuclease and FokI have revealed that these
proteins are also members of the same family.

<>

<1>Kovarik, A., Koukalova, B., Lim, L.Z., Matyasek, R., Lichtenstein, C.P., Leitch, A.R., Bezdek, M.
<2>Inhibition of tobacco DNA methyltransferase in vivo causes unequal hypomethylation of DNA repeated sequences.
<3>J. Biomol. Struct. Dyn.
<4>17
<5>1144-1145
<6>2000
<7>DNA repetitive sequences are characterized with a high content of methylated cytosine
(5-methylcytosine, 5-mC).  In tobacco genomic DNA about 30% of all cytosine residues are
methylated.  However, in our previous study it was found that two families of 5S rDNA
reiterated sequence have more than 50% of all cytosine residues methylated suggesting that
differences in DNA methylation levels between DNA repeated sequences may exist.  The aim of
this study was to compare the methylation patterns in several repetitive DNA sequences of
Nicotiana tabacum nuclear genome as well as their susceptibility to the effect of a
hypomethylating drug dihydroxypropyladenine.  This drug is thought to reduce DNA
methyltransferase activity by increasing S-adenosylhomocysteine levels.  To detect methylation
state in CG and CCG sites in DNA loci studied, methylation-sensitive restriction enzymes
(MspI, HapII, Sau3A1 and BamHI) were used.

<>

<1>Kowalski, J.C., Belfort, M., Stapleton, M.A., Holpert, M., Dansereau, J.T., Pietrokovski, S., Baxter, S.M., Derbyshire, V.
<2>Configuration of the catalytic GIY-YIG domain of intron endonuclease I-TevI: coincidence of computational and molecular findings.
<3>Nucleic Acids Res.
<4>27
<5>2115-2125
<6>1999
<7>I-TevI is a member of the GIY-YIG family of homing endonucleases.  It is folded into two
structural and functional domains, an N-terminal catalytic domain and a C-terminal DNA-binding
domain, separated by a flexible linker.  In this study we have used genetic anlayses,
computational sequence analysis and NMR spectroscopy to define the configuration of the
N-terminal domain and its relationship to the flexible linker.  The catalytic domain is an
alpha/beta structure contained within the first 92 amino acids of the 245-amino acid protein
followed by an unstructured linker.  Remarkably, this structured domain corresponds precisely
to the GIY-YIG module defined by sequence comparisons of 57 proteins including more than 30
newly reported members of the family.  Although much of the unstructured linker is not
essential for activity, residues 93-116 are required, raising the possibility that this region
may adopt an alternate conformation upon DNA binding.  Two invariant residues of the GIY-YIG
module, Arg27 and Glu75, located in alpha-helices, have properties of catalytic residues.
Furthermore, the GIY-YIG sequence elements for which the module is named form part of a
three-stranded antiparallel beta-sheet that is important for I-TevI structure and function.

<>

<1>Kowalski, J.C., Derbyshire, V.
<2>Characterization of homing endonucleases.
<3>Methods
<4>28
<5>365-373
<6>2002
<7>Homing endonucleases are a class of site-specific DNA endonucleases encoded by open reading
frames within introns and inteins. They initiate the mobility of their host element by
recognizing intronless or inteinless alleles of their host gene and making a double-strand
break. The homing endonucleases are notable for their long target sites and a tolerance for
sequence polymorphisms in their substrates. The methods used to study homing endonucleases are
similar to those used to study protein-DNA interactions in general. However, some variations
and specialized techniques are useful in characterizing homing endonucleases and these methods
are discussed.

<>

<1>Kozak, N.A., Buss, M., Lucas, C.E., Frace, M., Govil, D., Travis, T., Olsen-Rasmussen, M., Benson, R.F., Fields, B.S.
<2>Virulence factors encoded by Legionella longbeachae identified on the basis of the genome sequence analysis of clinical isolate D-4968.
<3>J. Bacteriol.
<4>192
<5>1030-1044
<6>2010
<7>Legionella longbeachae causes most cases of legionellosis in Australia and may be
underreported worldwide due to the lack of L. longbeachae-specific diagnostic tests. L.
longbeachae displays distinctive differences in intracellular trafficking, caspase 1
activation, and infection in mouse models compared to Legionella pneumophila, yet these two
species have indistinguishable clinical presentations in humans. Unlike other legionellae,
which inhabit freshwater systems, L. longbeachae is found predominantly in moist soil. In this
study, we sequenced and annotated the genome of an L. longbeachae clinical isolate from
Oregon, isolate D-4968, and compared it to the previously published genomes of L.
pneumophila. The results revealed that the D-4968 genome is larger than the L.
pneumophila genome and has a gene order that is different from that of the L.
pneumophila genome. Genes encoding structural components of type II, type IV Lvh, and type IV
Icm/Dot secretion systems are conserved. In contrast, only 42/140 homologs of genes encoding
L. pneumophila Icm/Dot substrates have been found in the D-4968 genome. L. longbeachae encodes
numerous proteins with eukaryotic motifs and eukaryote-like proteins unique to this species,
including 16 ankyrin repeat-containing proteins and a novel U-box protein. We predict that
these proteins are secreted by the L. longbeachae Icm/Dot secretion system. In contrast to the
L. pneumophila genome, the L. longbeachae D-4968 genome does not contain flagellar
biosynthesis genes, yet it contains a chemotaxis operon. The lack of a flagellum explains the
failure of L. longbeachae to activate caspase 1 and trigger pyroptosis in murine macrophages.
These unique features of L. longbeachae may reflect adaptation of this species to life in
soil.

<>

<1>Kozbial, P.Z., Mushegian, A.R.
<2>Natural history of S-adenosylmethionine-binding proteins.
<3>BMC Struct. Biol.
<4>5
<5>19
<6>2005
<7>Background.  S-adenosylmethionine is a source of diverse chemical groups used in biosynthesis
and modification of virtually every class of biomolecules. The most notable reaction requiring
S-adenosylmethionine, transfer of methyl group, is performed by a large class of enzymes,
S-adenosylmethionine-dependent methyltransferases, which have been the focus of considerable
structure-function studies. Evolutionary trajectories of these enzymes, and especially of
other classes of S-adenosylmethionine-binding proteins, nevertheless, remain poorly
understood. We addressed this issue by computational comparison of sequences and structures of
various S-adenosylmethionine-binding proteins.  Results.  Two widespread folds, Rossmann fold
and TIM barrel, have been repeatedly used in evolution for diverse types of
S-adenosylmethionine conversion. There were also cases of recruitment of other relatively
common folds for S-adenosylmethionine binding. Several classes of proteins have unique
unrelated folds, specialized for just one type of chemistry and unified by the theme of
internal domain duplications. In several cases, functional divergence is evident, when
evolutionarily related enzymes have changed the mode of binding and the type of chemical
transformation of S-adenosylmethionine. There are also instances of functional convergence,
when biochemically similar processes are performed by drastically different classes of
S-adenosylmethionine-binding proteins.  Comparison of remote sequence similarities and
analysis of phyletic patterns suggests that the last universal common ancestor of cellular
life had between 10 and 20 S-adenosylmethionine-binding proteins from at least 5 fold classes,
providing for S-adenosylmethionine formation, polyamine biosynthesis, and methylation of
several substrates, including nucleic acids and peptide chain release factor.  Conclusion.  We
have observed several novel relationships between families that were not known to be related
before, and defined 15 large superfamilies of SAM-binding proteins, at least 5 of which may
have been represented in the last common ancestor.

<>

<1>Kozdon, J.B., Melfi, M.D., Luong, K., Clark, T.A., Boitano, M., Wang, S., Zhou, B., Gonzalez, D., Collier, J., Turner, S.W., Korlach, J., Shapiro, L., McAdams, H.H.
<2>Global methylation state at base-pair resolution of the Caulobacter genome throughout the cell cycle.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>110
<5>E4658-E4667
<6>2013
<7>The Caulobacter DNA methyltransferase CcrM is one of five master cell-cycle regulators. CcrM
is transiently present near the end of DNA replication when it
rapidly methylates the adenine in hemimethylated GANTC sequences. The timing of
transcription of two master regulator genes and two cell division genes is
controlled by the methylation state of GANTC sites in their promoters. To explore
the global extent of this regulatory mechanism, we determined the methylation
state of the entire chromosome at every base pair at five time points in the cell
cycle using single-molecule, real-time sequencing. The methylation state of 4,515
GANTC sites, preferentially positioned in intergenic regions, changed
progressively from full to hemimethylation as the replication forks advanced.
However, 27 GANTC sites remained unmethylated throughout the cell cycle,
suggesting that these protected sites could participate in epigenetic regulatory
functions. An analysis of the time of activation of every cell-cycle regulatory
transcription start site, coupled to both the position of a GANTC site in their
promoter regions and the time in the cell cycle when the GANTC site transitions
from full to hemimethylation, allowed the identification of 59 genes as
candidates for epigenetic regulation. In addition, we identified two previously
unidentified N6-methyladenine motifs and showed that they maintained a constant
methylation state throughout the cell cycle. The cognate methyltransferase was
identified for one of these motifs as well as for one of two 5-methylcytosine
motifs.

<>

<1>Koziel, M., Lucid, A., Bullman, S., Corcoran, G.D., Lucey, B., Sleator, R.D.
<2>Draft Genome Sequence of Campylobacter corcagiensis Strain CIT045T, a Representative of a Novel Campylobacter Species Isolated from Lion-Tailed Macaques (Macaca silenus).
<3>Genome Announcements
<4>2
<5>e00248-14
<6>2014
<7>Campylobacter corcagiensis CIT045(T) (=CCUG 64942(T), LMG 27932(T)), a new member of the
Campylobacter genus, has recently been isolated from lion-tailed macaques in Cork, Ireland. To
further characterize this new species and its potential pathogenicity, the genome sequence of
C. corcagiensis was determined and is presented here.

<>

<1>Koziolkiewicz, M.
<2>Interactions between EcoRI restriction endonuclease and DNA.
<3>Postepy Biochem.
<4>37
<5>23-32
<6>1991
<7>*
A review in Polish arranged as:
   I. Specific interactions between proteins and DNA
  II. EcoRI endonuclease
II-1. EcoRI activity
II-2. Contact-points between EcoRI protein and DNA
II-3. Application of synthetic oligonucleotides in the studies on the mechanism of EcoRI
      endonuclease action
II-4. X-Ray studies on the EcoRI endonuclease - DNA complex


<>

<1>Koziolkiewicz, M., Stec, W.J.
<2>Application of phosphate-backbone-modified oligonucleotides in the studies on EcoRI endonuclease mechanism of action.
<3>Biochemistry
<4>31
<5>9460-9466
<6>1992
<7>Chemical synthesis of oligodeoxyribonucleotides modified at a preselected internucleotide bond
by the replacement of one of the two nonbridging oxygens by a sulfur atom or an ethoxy group
yields model substrates for studies on DNA-protein interactions. Chromatographic (RP-HPLC)
separaton of the diastereomers of oligonucleotides containing EcoRI canonical sequence
together with the assignment of the substituent of orientation in the DNA molecule allowed
study of the stereochemical aspects of DNA-EcoRI endonuclease interaction. The DNA segment
involved in interactions between EcoRI protein and phosphate groups appeared to be larger than
its canonical sequence,...GAATTC...,and was extended to the nonamer. The modification of
certain internucleotide bonds within this nonamer caused significant or complete protection
against the nucleolytic action of EcoRI and, in some cases, manifested the
diastereoselectivity of the enzyme. On the basis of the results of EcoRI-catalyzed hydrolysis
of stereodefined phosphorothioate and phosphotriester substrates, we propose a model to
explain this phenomenon at the molecular level.

<>

<1>Krabbe, M., Carlson, K.
<2>Squence and structure of endonuclease II-dependent cleavage sites in bacteriophage T4 DNA.
<3>J. Biol. Chem.
<4>266
<5>23407-23415
<6>1991
<7>Endonuclease II of bacteriophage T4 is required for in vivo restriction of
cytosine-containing DNA from its host, Escherichia coli, (as well as from phage
mutants lacking cytosine modification), normally the first step in the
reutilization of host DNA nucleotides for synthesis of phage DNA in infected
cells.  The phage cytosine-DNA is fragmented incompletely to yield genetically
defined fragments.  This restriction is different from that of type I, II, or
III restriction enzymes.  We have located seven major endonuclease II-dependent
restriction sites in the T4 genome, of which three were analyzed in detail; in
addition, abundant sites were cleaved in <5% of all molecules.  Sites I, II,
and III shared the sequence 5'-CCGNNTTGGC-3' and were cleaved in about 25% (I
and III) and 65% (II) of all molecules, predominantly staggered around the
first or second of the central unspecified base pairs to yield fragments with
one 5' base.  The less frequently cleaved sites I and III deviated from site II
in predicted helical structure when viewed from the consensus strand, and in
sequence when viewed from the opposite strand.  Thus, interaction with a
particular helical structure as well as recognition of the bases in DNA appears
important for efficient cleavage.

<>

<1>Kraev, A.S., Kravets, A.N., Chernov, B.K., Skryabin, K.G., Baev, A.A.
<2>The EcoRV restriction-modification system:  genes, enzymes, synthetic substrates.
<3>Mol. Biol. (Mosk)
<4>19
<5>278-284
<6>1985
<7>By a combination of deletion mutagenesis induced by nuclease, followed by
determination of the DNA nucleotide sequence, the organization of the genes of
the EcoRV restriction-modification system, encoded by plasmid, was established.
The genes of the restriction endonuclease (a protein with a size of 29,000)
and methylase (a protein with a size of 35,000) are read in opposite directions
from an intergene region with a size of 310 nucleotides, containing two
promoter regions.  No homology of the primary structures of the restriction
endonuclease EcoRV and the restriction endonuclease EcoRI, which recognizes a
similar nucleotide sequence and is also encoded by a plasmid, was detected.
The interaction of restriction endonuclease EcoRV with synthetic
deoxyoligonucleotides containing a phosphoamide bond in the site of the
presumed action of the restriction endonuclease was investigated.  It was shown
that this analog, in contrast to the synthetic dodecamer of the same structure,
but not containing phosphoamide bonds, is not split out by the restriction
endonuclease; in the control experiment it was found that under the action of
the restriction endonuclease EcoRV, blunt ends (GAT^ATC), rather than sticky
ends (GATAT^C) are formed, as was shown earlier.

<>

<1>Kraev, A.S., Zimin, A.A., Mironova, M.V., Janulaitis, A., Tanyashin, V.I., Skryabin, K.G., Bayev, A.A.
<2>The DNA ligase gene of bacteriophage T4.
<3>Dokl. Akad. Nauk.
<4>270
<5>1495-1500
<6>1983
<7>note: This paper shows that GAGCT^C is a site for SduI.  This is the missing
sequence from the earlier paper in FEBS Lett.

<>

<1>Kraft, C., Stack, A., Josenhans, C., Niehus, E., Dietrich, G., Correa, P., Fox, J.G., Falush, D., Suerbaum, S.
<2>Genomic Changes during Chronic Helicobacter pylori Infection.
<3>J. Bacteriol.
<4>188
<5>249-254
<6>2006
<7>The gastric pathogen Helicobacter pylori shows tremendous genetic variability within human
populations, both in gene content and at the
sequence level. We investigated how this variability arises by comparing
the genome content of 21 closely related pairs of isolates taken from the
same patient at different time points. The comparisons were performed by
hybridization with whole-genome DNA microarrays. All loci where
microarrays indicated a genomic change were sequenced to confirm the
events. The number of genomic changes was compared to the number of
homologous replacement events without loss or gain of genes that we had
previously determined by multilocus sequence analysis and mathematical
modeling based on the sequence data. Our analysis showed that the great
majority of genetic changes were due to homologous recombination, with
1/650 events leading to a net gain or loss of genes. These results suggest
that adaptation of H. pylori to the host individual may principally occur
through sequence changes rather than loss or gain of genes.

<>

<1>Kramarov, V.M.
<2>Strain of Bacillus cereus - is used as producer of restriction enzyme Bcc83I.
<3>Soviet Patent Office
<4>SU 1620484
<5>
<6>1991
<7>Bacillus cereus VKPM B-4362(83) strain is a producer of restriction enzyme Bce83I, which is
capable of recognising the nucleotide segment '...CTCAAG...' on a double-stranded DNA. The
strain was isolated as a contaminant during cultivation of another culture. The yield of crude
biomass is 3-4 g/l and the one of the enzyme is 1000 units/g of biomass.

<>

<1>Kramarov, V.M., Fomenkov, A.I., Matvienko, N.I.
<2>A new type of cleavage of the recognition sequence by a site-specific endonuclease Bst4.4I from Bacillus stearothermophilus 4.4.
<3>Bioorg. Khim.
<4>14
<5>916-920
<6>1988
<7>A site-specific endonuclease Bst4.4I was isolated from the cell extract of
Bacillus stearothermophilus 4.4 and partially purified by chromatography on
Ultragel AcA-44 and heparin-Sepharose.  It was shown that the endonuclease
cleaves lambda and M13 DNA yielding distinct fragments just as endonucleases of
Types II and III but, in contrast to them, can produce two two-strand cuts
separated with 30 to 32 nucleotides in the region of the recognition site.

<>

<1>Kramarov, V.M., Fomenkov, A.I., Matvienko, N.I., Ubieta, R.H., Smolianinov, V.V., Gorlenko, V.M.
<2>A new sequence-specific endonuclease CauB3I from Chloroflexus aurantiacus B3.
<3>Bioorg. Khim.
<4>13
<5>773-776
<6>1987
<7>A sequence-specific endonuclease CauB3I has been isolated from cell extracts of
Chloroflexus aurantiacus and partially purified by chromatography on
heparin-sepharose; the yield was 3000 units per 1 g of cells.  The final
preparation is free of non-specific nucleases.  It is shown that endonuclease
CauB3I recognizes 5' T^CCGGA 3' sequence in double-stranded DNA and cleaves it
as shown.  Methylation of adenine in the recognition sequence makes it
resistant to CauB3I.

<>

<1>Kramarov, V.M., Mazanov, A.L., Pachkunov, D.M., Smolyaninov, V.V., Matvienko, N.I.
<2>Another site-specific endonuclease from Rhodopseudomonas sphaeroides.
<3>Bioorg. Khim.
<4>10
<5>46-49
<6>1984
<7>A site-specific endonuclease RshII has been purified by chromatography on
Ultro-gel AcA-44, aminohexyl-Sepharose 4B and heparin-Sepharose 6B.  The final
preparation did not contain nonspecific nucleases or endonuclease RshI.  The
RshII endonuclease has been shown to recognize in double-stranded DNA the
following nucleotide sequence:  5'-C'C (C) GG-3'/3'-GG (G) CC-5'.

<>

<1>Kramarov, V.M., Mazanov, A.L., Smolyaninov, V.V.
<2>Isolation of a second site-specific endonuclease from Xanthomonas holcicola and its characterization.
<3>Bioorg. Khim.
<4>8
<5>220-223
<6>1982
<7>A method is proposed for purifying the site-specific endonuclease XhoII by
chromatography on phosphocellulose and aminohexyl-Sepharose.  The final product
contains no nonspecific nucleases as impurities nor the endonuclease XhoI.  It
has been shown that the endonuclease recognizes and hydrolyzes DNA in the
nucleotide sequence 5'R-G-A-T-C-Y3'.  According to the results of gel
filtration, the molecular weight of the endonuclease is 40,000 +/- 2000.  In
the hydrolysis of DNA by the endonuclease, Mg2+ can be replaced by Mn2+.

<>

<1>Kramarov, V.M., Mochalov, V.V., Smolyanino, V.V.
<2>Separation of site specific endonuclease ClaI - involves chromatography of cell-free extract of C. latum G on column containing carrier with specified immobilised dye.
<3>Soviet Patent Office
<4>SU 1306127
<5>
<6>1988
<7>The process involves preparation of a carrier by suspending 20ml of Sepharose CL-4B in 30ml
H20, adding 110mg of cibachrone (sic) blue F3GA dyestuff and then a solution of 4M NaCl up to
a final concentration of 0.4M, and making up the volume to 45ml. After stirring mechanically
for 20 mins., 0.2ml of 5M NaOH is added and the mixture is incubated at 62 deg. C. The amount
of immobilised dyestuff can be quantitatively detected on the basis of the reduction in
optical density of the supernatant at 610nm. The carrier is then washed on a glass filter to
remove unbound dye. A cell-free extract of C. latum G containing the endonuclease ClaI is then
subjected to chromatography on a column containing the precipitated carrier with the dyestuff
F3GA. The column is washed with 10mM potassium phosphate buffer containing 0.1M NaCl until the
optical absorption at 280nm is zero. Then protein is eluted with a linear gradient 0.1-1M NaCl
in 10mM potassium phosphate buffer. Endonuclease ClaI is eluted at a NaCl concentration of
0.5-0.8M. The fractions containing the enzyme are combined, dialysed for 6-10 hrs. against a
buffer containing 50% glycerine and stored at minus 20oC.

<>

<1>Kramarov, V.M., Pachkunov, D.M., Matvienko, N.I.
<2>Site-specific endonucleases BmeI and RshII.
<3>Nek. Aspekty Fiziol. Mikroorg. Akad. Nauk. SSR., Nauchn. Tsentr Biol. Issled., Gaziev, A.I., Pushchino, USSR.
<4>0
<5>22-26
<6>1983
<7>None

<>

<1>Kramarov, V.M., Prokhorova, S.D., Smolyanino, V.V.
<2>Site-specific endonuclease PvuII producer - comprises Escherichia coli strain obtained by pBp5RM plasmid transformation in E. coli strain C-600.
<3>Soviet Patent Office
<4>SU 1294824
<5>
<6>1987
<7>The recombinant plasmid pBP5RM, which codes biosynthesis of site-specific PvuII endonuclease,
has a size of 900 pairs of bases and consists of the following elements: EcoRI-SalI-fragment
of plasmid pBR322, the distance between sites 650 pairs of bases, XhoI-XhoI - fragment of DNA
of Proteus vulgaris containing the gene of PvuII endonuclease and sites ClaI, HindIII, HindIII
and EcoRI, the distances between which (from the XhoI site) are 900, 200, 1800, 1600 and 150
base pairs, respectively, Sal I - EcoR I - fragment of plasmid pBR322 containing the replicon
plasmid of pBR322 and the gene responsible for synthesis of beta-lactamase, (which determines
resistance to ampicillin). Plasmid pBP5RM is prepared by splitting the DNA of pBR322 plasmid
with SalI endonuclease, hydrolysing the DNA from Proteus vulgaris with XhoI endonuclease,
combining the obtained fragments by the action of DNA - ligase, transforming E.coli with this
mixture, separating plasmid DNA from the cells, hydrolysing with PvuII endonuclease, again
transforming E. coli cells with the obtained mixture, and selecting the ampicillin-resistant
clones, from which the recombinant plasmid is separated. The strain E. coli C600P5RM is used
to produce the site-specific PvuII endonuclease and is obtained by transformation of plasmid
pBP5RM in E. coli C 600.

<>

<1>Kramarov, V.M., Skrypina, N.A., Smolyaninov, V.V., Smirnov, V.V., Resnik, S.R., Sorokulova, I.B., Matvienko, N.I.
<2>New site-specific endonuclease producer strains of Bacillus genera.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>42-45
<6>1989
<7>52 Bacillus strains have been tested for their production of site-specific
endonucleases.  The sequence recognized by the enzymes was determined for 23
enzymes, the cleavage site inside the sequence was determined for 5 enzymes.
All the enzymes under study were found to be isoschizomers of known enzymes.
The selected strains are unusual for their high level of site-specific
endonucleases content and may be used as producers of the enzymes.

<>

<1>Kramarov, V.M., Smolyaninov, V.V.
<2>DNA methylase from Arthrobacter luteus screens DNA from the action of site-specific endonuclease AluI.
<3>Biokhimiia
<4>46
<5>1526-1529
<6>1981
<7>DNA-methylase was isolated from a cell extract of A. luteus and partially
purified by chromatography on phosphocellulose.  The purified enzyme methylates
DNA of phage lambda and plasmids pBR322, thus making them resistant to a
subsequent action of endonuclease AluI.  It has been shown that cytosine is the
object of methylation within DNA.  This modification does not screen DNA from
the action of site-specific endonucleases SalI, Bam HI, EcoRI, EcoRII, XhoI and
XhoII  It has been experimentally demonstrated that the isolated methylase is
site-specific and identifies in the DNA the nucleotide sequence 5'-AGCT-3', by
methylating cytosine in the DNA.

<>

<1>Krasilnikova, M.M., Izvolskii, K.I., Krupnik, O.V., Lazurkin, Y.S.
<2>Use of specific binding of peptide nucleic acids to DNA in the "Achilles heel" method.
<3>Dokl. Akad. Nauk.
<4>344
<5>552-555
<6>1995
<7>
<>

<1>Krasnoslobodtsev, A.V., Shlyakhtenko, L.S., Lyubchenko, Y.L.
<2>Probing Interactions within the Synaptic DNA-SfiI Complex by AFM Force Spectroscopy.
<3>J. Mol. Biol.
<4>365
<5>1407-1416
<6>2006
<7>SfiI belongs to a family of restriction enzymes that function as tetramers, binding two
recognition regions for the DNA cleavage reaction.
The SfiI protein is an attractive and convenient model for studying
synaptic complexes between DNA and proteins capable of site-specific
binding. The enzymatic action of SfiI has been very well characterized.
However, the properties of the complex before the cleavage reaction are
not clear. We used single-molecule force spectroscopy to analyze the
strength of interactions within the SfiI-DNA complex. In these
experiments, the stability of the synaptic complex formed by the enzyme
and two DNA duplexes was probed in a series of approach-retraction cycles.
In order to do this, one duplex was tethered to the surface and the other
was tethered to the probe. The complex was formed by the protein present
in the solution. An alternative setup, in which the protein was anchored
to the surface, allowed us to probe the stability of the complex formed
with only one duplex in the approach-retraction experiments, with the
duplex immobilized at the probe tip. Both types of complexes are
characterized by similar rupture forces. The stability of the complex was
determined by measuring the dependence of rupture forces on force loading
rates (dynamic force spectroscopy) and the results suggest that the
dissociation reaction of the SfiI-DNA complex has a single energy barrier
along the dissociation path. Dynamic force spectroscopy was instrumental
in revealing the role of the 5 bp spacer region within the palindromic
recognition site on DNA-SfiI in the stability of the complex. The data
show that, although the change of non-specific sequence does not alter the
position of the activation barrier, it changes values of the off rates
significantly.

<>

<1>Krause, A. et al.
<2>Complete genome of the mutualistic, N2-fixing grass endophyte Azoarcus sp. strain BH72.
<3>Nat. Biotechnol.
<4>24
<5>1385-1391
<6>2006
<7>Azoarcus sp. strain BH72, a mutualistic endophyte of rice and other
grasses, is of agrobiotechnological interest because it supplies
biologically fixed nitrogen to its host and colonizes plants in remarkably
high numbers without eliciting disease symptoms. The complete genome
sequence is 4,376,040-bp long and contains 3,992 predicted protein-coding
sequences. Genome comparison with the Azoarcus-related soil bacterium
strain EbN1 revealed a surprisingly low degree of synteny. Coding
sequences involved in the synthesis of surface components potentially
important for plant-microbe interactions were more closely related to
those of plant-associated bacteria. Strain BH72 appears to be 'disarmed'
compared to plant pathogens, having only a few enzymes that degrade plant
cell walls; it lacks type III and IV secretion systems, related toxins and
an N-acyl homoserine lactones-based communication system. The genome
contains remarkably few mobile elements, indicating a low rate of recent
gene transfer that is presumably due to adaptation to a stable, low-stress
microenvironment.

<>

<1>Krause, D.O., Little, A.C., Dowd, S.E., Bernstein, C.N.
<2>Complete genome sequence of adherent invasive Escherichia coli UM146 isolated from ileal Crohn's disease biopsy tissue.
<3>J. Bacteriol.
<4>193
<5>583
<6>2010
<7>Escherichia coli UM146 was isolated from the ileum of a Crohn's disease patient. It adheres
to and invades enterocytes and can replicate inside
macrophages. Its complete genome sequence reveals that it is most closely
related to the human urinary tract pathogen E. coli CFT073, but it has a
host of genes that are novel and for which function has not been ascribed.

<>

<1>Krause, K.L., Miller, M.D.
<2>A new engine for cleaving nucleic acid.
<3>ACS Symp. Ser.
<4>827
<5>270-293
<6>2002
<7>Rapid and accurate cleavage of nucleic acid material, such as DNA and RNA, is a fundamentally
important biochemical process in all living
organisms carried out by enzymes called nucleases. We present here a
discussion and comparison of two nucleases of widely disparate
properties but who share a newly described nuclease active site
geometry. Serratia marcescens, a pathogenic Gram negative bacterium
produces an enzyme that presents a new paradigm in nucleases. Its fold
is new, it is capable of very rapid, and relatively non sequence
dependent cleavage of several types of DNA and RNA. On the other hand,
I-PpoI is a homing endonuclease which is encoded by a group I intron in
the rRNA genes of Physarum polycephalum (1). It also possesses a new
fold, but it only slowly cleaves DNA at a very specific 15 by site.
Both enzymes possess the same active site geometry, but no other
structural homology. The structural basis for their different
properties is due to two main factors, the nature of their interaction
with substrate and the presence of a mobile metal in I-Ppol
endonuclease that must migrate into position prior to catalysis.

<>

<1>Kravets, A.N., Pertsev, A.V., Tarutina, Z.E., Solonin, A.S.
<2>High homology of plasmids carrying class II restriction modification systems, EcoRV isoschizomers, and their prevalence in natural Escherichia coli strains.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>3
<5>20-22
<6>1998
<7>Screening of 660 clinical Enterobacteriaceae strains from the collection of L.V. Gromashevsky,
Kiev Institute of Epidemiology and Infectious Diseases, for specific endonuclease activity
revealed site-specific endonucleases, EcoRV isoschizomers, in 6 E. coli strains.  Genes coding
for endonucleases and methyltransferases were localized on small (6.2 kb) multicopy Hsd+
plasmids.  All plasmids were successfully transferred in laboratory strain E. coli K802.
Restriction analysis and subcloning showed no differences in the structural and functional
organization of the plasmids studied and a previously revealed pLB1 plasmid, thus reflecting
their high homology, if not identity.  These data allow us to propose effective horizontal
transfer of EcoRV plasmids among natural E. coli isolates in the region studied.

<>

<1>Kravets, A.N., Pertsev, A.V., Tarutina, Z.Y., Krendelev, Y.D., Zakharova, M.V., Beletskaya, I.V., Solonin, A.S.
<2>New isoschizomers of restriction endonucleases in Pseudomonas aeruginosa strains.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>14-17
<6>1998
<7>Testing of Pseudomonas aeruginosa strains from the collection of the L.V. Gromashevsky
Institute of Epidemiology and Infectious Diseases (Kiev) for specific endonucleases resulted
in the isolation of five class II restriction endonucleases, which were partially purified and
their recognition targets were determined.  Two of these endonucleases, Pae2kI and Pae18kI,
are isoschizomers of BglII (5'-AGACTC-3').  Pae5kI and Pae14kI, recognize the
5'-CCGC/GG-3' sequence and are therefore true isoschizomers of SacII.  Hence, Pae17kI is an
isoschizomer of PvuII and cleaves the DNA within the recognition sequence 5'-CAG/CTG-3'.
BglII and PvuII are for the first time detected in Pseudomonas aeruginosa.

<>

<1>Kravets, A.N., Solonin, A.C., Kuzmin, N.P., Glatman, L.I., Frolov, A.F., Vasyurenko, Z.P., Tarutina, Z.E.
<2>A plasmid encoding the restriction enzyme CfrBI and its methylase.
<3>Soviet Patent Office
<4>SU 1573026 A
<5>
<6>1990
<7>The present invention describes the construction of a novel plasmid BG M3 encoding and
expressing the genes of the restriction - modification system CfrBI from Citrobacter freundii
4111 in E. coli.

<>

<1>Kravets, A.N., Solonin, A.S., Denjmuchametov, M.M., Zakharova, M.V., Beletskaya, I.V., Sineva, E.V.
<2>Plasmid pEco1831Kl carrying gene for restriction endonuclease Eco1831KI and manufacture of the enzyme with Escherichia coli.
<3>Russian Patent Office
<4>RU 2054042 C
<5>
<6>1996
<7>
<>

<1>Kravets, A.N., Solonin, A.S., Pertsev, A.V., Zakharova, M.V., Beletskaya, I.V.
<2>Plasmid pEco29KI for expression of the gene for restriction endonuclease Eco29KI and its manufacture with Escherichia coli.
<3>Russian Patent Office
<4>RU 2054037 C
<5>
<6>1996
<7>
<>

<1>Kravets, A.N., Solonin, A.S., Zakharova, M.V., Tarutina, Z.E.
<2>Plasmid localization and cloning of restriction-modification genes from Citrobacter freundii 4111 strain.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>7
<5>4-7
<6>1992
<7>Over 60 producing strains of restriction endonucleases type II have been found among 500
different strains, mostly Enterobacteriaceae. The strain Citrobacter freundii 4111 produces
restriction endonuclease CfrBI, a new isoschizomer of StyI. The genes of the
restriction-modification system CfrBI were located on the multicopy plasmid pZE8 containing
the ColE1-type replicon and cloned into E. coli K802. The deletion variant of 3.2-kb pZE8
which contains intact restriction-modification and a DNA fragement responsible for autonomous
plasmid replication was selected among the recombinant plasmids. The strain with higher R.
CfrBI production (at least 10,000,000 U/g cells, which is 500-fold higher than the wild
strain) was constructed.

<>

<1>Kravets, A.N., Zakharova, M.V., Solonin, A.S., Kuzmin, N.P., Tanyushin, V.I., Glatman, L.I., Moroz, A.F., Baev, A.A.
<2>Cloning of genes of the EcoRV restriction-modification system and regulation of their expression.
<3>Mol. Biol. (Mosk)
<4>24
<5>438-447
<6>1990
<7>A number of recombinant plasmids bearing genes of the EcoRV
restriction-modification system were constructed.  The individual genes were
inserted into plasmids belonging to different incompatibility groups.  The
regulatory regions of the genes coding for the methylase and restriction
endonuclease were cloned and studied.  It was shown using the specialized
vector pVE8 that the promoter region determining transcription of the
restriction endonuclease was comparable in terms of efficiency with the early
promoters of phage lambda and equaled approximately 70% of the efficiency of
the left-oriented promoter PL.  The promoter region of the methylase gene
provided about one half as efficient transcription as did the promoter region
of the restriction-endonuclease gene.  A recombinant plasmid was obtained in
which the EcoRV restriction-endonuclease gene found itself under the control of
the complementary regulated promoter of phage lambda, PR, and provided
30-40-fold enhancement of biosynthesis of the restriction endonuclease, given
inactivation of the phage-lambda-c1857 temperature-sensitive repressor.  Under
inducing conditions, the amount of EcoRV restriction endonuclease in the
superproducer strain amounted to approximately 10% of the total cell protein.
The factors conducive to the level of EcoRV restriction endonuclease induction
attained are discussed.

<>

<1>Kravetz, A.N., Tarutina, Z.E., Solonin, A.S.
<2>Two novel restriction endonucleases from Pseudomonas aeruginosa.
<3>Nucleic Acids Res.
<4>19
<5>4781
<6>1991
<7>PaePI and PaeHI, type II restriction endonucleases have been isolated from
clinical strain Pseudomonas aeruginosa 4148.  The enzymes were separated and
purified by chromatography on DEAE-cellulose DE52, hydroxylapatite and mono Q
column (FPLC system, Pharmacia Ltd).

<>

<1>Kravetz, A.N., Zakharova, M.V., Beletskaya, I.V., Sineva, E.V., Denjmuchametov, M.M., Petrov, S.I., Glatman, L.I., Solonin, A.S.
<2>The cleavage sites and localization of genes encoding the restriction endonuclease Eco1831I and EcoHI.
<3>Gene
<4>129
<5>153-154
<6>1993
<7>The restriction endonucleases Eco1831I and EcoHI cleave before the first 5'-cytosine in the
recognition sequence 5'CCSGG 3'/3' GGSCC 5' (where S=G or C), generate 5-base 5' cohesive
ends and are encoded by homologous plasmids that are restricted in McrA+ hosts. Thus, they
differ in their cleavage specificity from that of the BcnI isoschizomer, which cleaves after
the second 5' cytosine.

<>

<1>Kravetz, A.N., Zakharova, M.V., Beljetzkaja, I.V., Pertzev, A.V., Spivak, O.I., Solonin, A.S.
<2>Two novel restriction endonucleases from Klebsiella pneumoniae.
<3>Nucleic Acids Res.
<4>21
<5>1501
<6>1993
<7>Kpn49kI and Kpn49kII, type II restriction endonucleases have been isolated from the clinical
strain Klebsiella pneumoniae 49k.

<>

<1>Krebes, J., Morgan, R.D., Bunk, B., Sproeer, C., Luong, K., Parusel, R., Anton, B.P., Koenig, C., Josenhans, C., Overmann, J., Roberts, R.J., Korlach, J., Suerbaum, S.
<2>The complex methylome of the human gastric pathogen Helicobacter pylori.
<3>Nucleic Acids Res.
<4>42
<5>2415-2432
<6>2014
<7>The genome of Helicobacter pylori is remarkable
for its large number of restriction-modification
(R-M) systems, and strain-specific diversity in R-M
systems has been suggested to limit natural transformation,
the major driving force of genetic diversification
in H. pylori. We have determined the
comprehensive methylomes of two H. pylori
strains at single base resolution, using Single
Molecule Real-Time (SMRT) sequencing. For
strains 26695 and J99-R3, 17 and 22 methylated
sequence motifs were identified, respectively. For
most motifs, almost all sites occurring in the
genome were detected as methylated. Twelve
novel methylation patterns corresponding to nine
recognition sequences were detected (26695, 3;
J99-R3, 6). Functional inactivation, correction of
frameshifts as well as cloning and expression of
candidate methyltransferases (MTases) permitted
not only the functional characterization of multiple,
yet undescribed, MTases, but also revealed novel
features of both Type I and Type II R-M systems,
including frameshift-mediated changes of sequence
specificity and the interaction of one MTase with
two alternative specificity subunits resulting in different
methylation patterns. The methylomes of
these well-characterized H. pylori strains will
provide a valuable resource for future studies
investigating the role of H. pylori R-M systems in
limiting transformation as well as in gene regulation
and host interaction.

<>

<1>Krempl, P.M., Mairhofer, J., Striedner, G., Thallinger, G.G.
<2>Finished Genome Sequence of the Laboratory Strain Escherichia coli K-12 RV308 (ATCC 31608).
<3>Genome Announcements
<4>2
<5>e00971-14
<6>2014
<7>Escherichia coli strain K-12 substrain RV308 is an engineered descendant of the K-12 wild-type
strain. Like its ancestor, it is an important organism in
biotechnological research and is heavily used for the expression of single-chain
variable fragments. Here, we report the complete genome sequence of E. coli K-12
RV308 (ATCC 31608).

<>

<1>Kresge, N., Simoni, R.D., Hill, R.L.
<2>The Characterization of Restriction Endonucleases: the Work of Hamilton Smith.
<3>J. Biol. Chem.
<4>285
<5>e2-3
<6>2010
<7>Hamilton Othanel Smith was born in 1931 in New York City.  In 1937, he and his family moved to
Champaign-Urbana, Illinois, because his father had joined the faculty of the department of
education at the University of Illinois.  As a boy, Smith was interested in chemistry,
electricity, and electronics, and he spent many hours with his brother in their basement
laboratory, which was stocked with supplies purchased from their paper route earnings.  Smith
attended a small college preparatory school called the University Laboratory High School and
graduated in 3 years largely tue to this science teacher who allowed him to complete chemistry
and physics during the summer.

<>

<1>Kretchmar, S.A., Chiu, H.S., Srinivasan, A.
<2>Interaction of caffeine with DNA:  an evaluation with restriction endonucleases.
<3>Microbios Lett.
<4>31
<5>31-38
<6>1986
<7>The endonucleolytic action of several restriction enzymes on the lambda, pBR322
and PhiX174 (RF I, RF II and single-stranded virion DNA) were tested in the
presence of caffeine.  No aberrant restriction cleavage pattern was observed by
adding caffeine directly to the restriction reaction mixture.  DNA pretreated
with caffeine also exhibited similar results.  Ligation of DNA fragments,
havaing either protruding single-strand or flush ends, is not affected by
caffeine.  These results strongly argue against the possible interaction of
caffeine with DNA.

<>

<1>Kretz, P.L., Kohler, S.W., Short, J.M.
<2>Identification and characterization of a gene responsible for inhibiting propagation of methylated DNA sequences in mcrA mcrB1 Escherichia coli strains.
<3>J. Bacteriol.
<4>173
<5>4707-4716
<6>1991
<7>Identifying and eliminating endogenous bacterial enzyme systems can
significantly increase the efficiency of propagation of eukaryotic DNA in
Escherichia coli.  We have recently examined one such system which inhibits the
propagation of lambda DNA rescued from transgenic mouse tissues.  This rescue
procedure utilizes lambda packaging extracts for excision of the lambda DNA
from the transgenic mouse genome, as well as E. coli cells for subsequent
infection and propagation.  This assay, in combination with conjugal mating, P1
transduction, and gene cloning, was used to identify and characterize the E.
coli locus responsible for this difference in efficiency.  It was determined
that the E. coli K-12 mcrB gene when expressed on a high-copy-number plasmid
can cause a decrease in rescue efficiency despite the presence of the mcrB1
mutation, which inactivates the classic McrB restriction activity.  (This
mutation was verified by sequence analysis.)  However, this McrB1 activity is
not observed when the cloned mcrB1 gene is inserted into the E. coli genome at
one copy per chromosome.  A second locus was identified which causes a decrease
in rescue efficiency both when expressed on a high-copy-number plasmid and when
inserted into the genome.  The data presented here suggest that this locus is
mrr and that the mrr gene product can recognize and restrict
cytosine-methylated sequences.  Removal of this DNA region including the mrr
gene from E. coli K-12 strains allows high rescue efficiencies equal to those
of E. coli C strains.  These modified E. coli K-12 plating strains and lambda
packaging extract strains should also allow a significant improvement in the
efficiency and representation of eukaryotic genomic and cDNA libraries.

<>

<1>Kretz, P.L., Reid, C.H., Greener, A., Short, J.M.
<2>Effect of lambda packaging extract mcr restriction activity on DNA cloning.
<3>Nucleic Acids Res.
<4>17
<5>5409
<6>1989
<7>The negative effect of bacterial mcrA and mcrB restriction activity on the cloning of
methylated DNA has recently been demonstrated. In order to determine the effect of these
restriction systems on lambda and cosmid packaging, an mcrA-, B-, hsd-, mrr- packaging extract
strain was constructed by P1 transduction. The extract prepared from this strain, Gigapack II,
was tested against the restriction positive (mcrA+,B-) extract, Gigapack I, by comparing
efficiencies in constructing a cosmid library. The results indicate that these restriction
enzymes are active in lambda packaging extracts and can affect cloning efficiencies. The
elimination of mcrA,B restriction activity from Gigapack II allowed a 2-3 fold increase in the
number of cosmid clones obtained. Similar effects have been observed with lambda libraries.
This effect has also been shown to increase as the extent of DNA methylation increases. The
results demonstrate the importance of utilizing restriction deficient lambda packaging
extracts for improved cloning efficiency and possibly genomic representation. Optimal results
are obtained when both mcrA-,B- packaging extracts and plating strains are used.

<>

<1>Kriebardis, A., Guschlbauer, W.
<2>dam methylase from E. coli: Circular dichroism investigations of the secondary structure and influence of S-adenosylmethionine.
<3>FEBS Lett.
<4>213
<5>297-300
<6>1987
<7>The enzyme dam methylase which recognizes and methylates the adenine in the palindromic
sequence GATC in DNA was isolated and the secondary structure was determined by CD
spectroscopy and various predicting methods from the amino acid sequence. The interaction of
dam methylase with S-adenosylmethionine was studied by CD spectroscopy indicating a decrease
of the percentage of alpha-helix as the amount of S-adenosylmethionine bound to the enzyme was
increased.

<>

<1>Krishnakumar, R., Assad-Garcia, N., Benders, G.A., Phan, Q., Montague, M.G., Glass, J.I.
<2>Targeted Chromosomal Knockouts in Mycoplasma pneumoniae.
<3>Appl. Environ. Microbiol.
<4>76
<5>5297-5299
<6>2010
<7>Most gene knockouts in mycoplasmas are achieved through labor-intensive
transposon mutagenesis. Here, we describe a method for making targeted
deletions in Mycoplasma pneumoniae by use of homologous recombination. In
this method, M. pneumoniae is transformed with a plasmid carrying an
antibiotic resistance marker flanked by 1-kb regions surrounding the
target gene. Following selection for the antibiotic resistance, colonies
are screened for double crossovers which indicate complete deletion of the
target open reading frame.

<>

<1>Krishnamurthy, V., Rao, D.N.
<2>Interaction of EcoPI modification methylase with S-adenosyl-L-methionine: a UV-crosslinking study.
<3>Biochem. Mol. Biol. Int.
<4>32
<5>623-632
<6>1994
<7>EcoPI modification methylase was radioactively labeled when incubated with
S-adenosyl-L-[methyl-3H]methionine in the presence of ultraviolet light. Crosslinking of the
enzyme as detected by electrophoresis on sodium dodecyl sulfate - polyacrylamide gel followed
by fluorography and autoradiography, was shown to be specific by a number of criteria. More
importantly, EcoPI modification methylase was also radioactively labeled with
S-adenosyl-L-[carboxyl-14C]methionine demonstrating that labeling involved binding of the
entire AdoMet molecule rather than methylation of the protein. Further, c2 EcoPI mutant DNA
modification methylases which show negligible or very little methylation activity,
correspondingly formed a weak or no adduct upon crosslinking. These results suggest that
photolabeling of EcoPI DNA modification methylase occurs at the AdoMet binding site.

<>

<1>Krishnaswamy, T.D.S.
<2>Structure-based sequence alignment of type-II restriction endonucleases.
<3>Biochim. Biophys. Acta
<4>1544
<5>217-228
<6>2001
<7>The type-II restriction endonucleases generally do not share appreciable amino acid sequence
homology. The crystal structures of restriction endonucleases EcoRI and BamHI have shown these
enzymes to possess striking 3D-structural resemblance, i.e., they have a similar overall fold
and similar active sites, though they possess <23% sequence identity. Structural
superimposition of EcoRI, BamHI, EcoRV, and PvuII based on active site residues led to
sequence alignments which showed nine possible sequence motifs. EcoRV and PvuII show a more
similar pattern than EcoRI and BamHI suggesting that they belong to a different subgroup. The
motifs are characterized by charged and/or hydrophobic residues. From other studies on the
structure of these endonucleases, three of the motifs could be implicated in DNA binding,
three in forming the active site and one in dimer formation. However, the motifs were not
identifiable by regular sequence alignment methods. It is found that motif IX in BamHI is
formed by reverse sequence order and the motif IX in PvuII is formed from the symmetry related
monomer of the dimer. The inter-motif distance is also quite different in these cases. Of the
nine motifs, motif III has been earlier identified as containing the PD motif involving one of
the active site residues. These motifs were used in a modified profile analysis procedure to
identify similar regions in eight other endonuclease sequences for which structures are not
known.

<>

<1>Kriss, J., Herrin, G., Forman, L., Cotton, R.
<2>Digestion conditions resulting in altered cut site specificity for HinfI.
<3>Nucleic Acids Res.
<4>18
<5>3665
<6>1990
<7>None

<>

<1>Kristiansen, R., Dueholm, M.S., Bank, S., Nielsen, P.H., Karst, S.M., Cattoir, V., Lienhard, R., Grisold, A.J., Olsen, A.B., Reinhard, M., Soby, K.M., Christensen, J.J., Prag, J., Thomsen, T.R.
<2>Complete Genome Sequence of Actinobaculum schaalii Strain CCUG 27420.
<3>Genome Announcements
<4>2
<5>e00880-14
<6>2014
<7>Complete genome sequencing of the emerging uropathogen Actinobaculum schaalii indicates that
an important mechanism of its virulence is attachment pili, which
allow the organism to adhere to the surface of animal cells, greatly enhancing
the ability of this organism to colonize the urinary tract.

<>

<1>Kriszt, B., Tancsics, A., Cserhati, M., Toth, A., Nagy, I., Horvath, B., Nagy, I., Tamura, T., Kukolya, J., Szoboszlay, S.
<2>De Novo Genome Project for the Aromatic Degrader Rhodococcus pyridinivorans Strain AK37.
<3>J. Bacteriol.
<4>194
<5>1247-1248
<6>2012
<7>Here, we present the complete genome sequence of Rhodococcus pyridinivorans AK37  strain NCAIM
PB1376, which was isolated from an oil-polluted site in Hungary. R.
pyridinivorans AK37 is an aerobic, nonsporulating, nonmotile, Gram-positive
bacterium with remarkable aromatic-decomposing activity.

<>

<1>Kriukiene, E.
<2>Domain organization and metal ion requirement of the Type IIS restriction endonuclease MnlI.
<3>FEBS Lett.
<4>580
<5>6115-6122
<6>2006
<7>A two-domain structure of the Type US restriction endonuclease Mull has been identified by
limited proteolysis. An N-terminal domain of the
enzyme mediates the sequence-specific interaction with DNA, whereas a
monomeric C-terminal domain resembles bacterial colicin nucleases in
its requirement for alkaline earth as well as transition metal ions for
double- and single-stranded DNA cleavage activities. The results
indicate that the fusion of the non-specific HNH-type nuclease to the
DNA binding domain had transformed Mull into a Mg2+-, Ni2+-, Co2+-,
Mn2+-, Zn2+-, Ca2+-dependent sequence-specific enzyme. Nevertheless,
Mull retains a residual single-stranded DNA cleavage activity
controlled by its C-terminal colicin-like nuclease domain.

<>

<1>Kriukiene, E.
<2>Restriction endonuclease MnlI - A member of the HNH family of endonucleases.
<3>Ph.D. Thesis, Vilnius University
<4>
<5>1-45
<6>2007
<7>CONCLUSIONS
1. The Type IIS restriction-modification system MnlI is composed of N6-methyladenine and
C5-methylcytosine methyltransferases and a restriction enzyme. The methyltransferases modify
cytosine and adenine on the opposite strands of the recognition sequence, resulting in
5'-m5CCTC-3'/5'-Gm6AGG-3'. The MnlI restriction endonuclease cleaves both DNA strands 7/6
nucleotides downstream of the recognition site.
2. The active site of MnlI REase resembles those of the bacterial colicin DNases ColE7 and
ColE9, which belong to the HNH superfamily of nucleases. The motif 306Rx3ExHHx14Nx8H located
in the C-terminal part of the protein comprises the active site of MnlI.
3. MnlI is a homodimeric protein capable of binding two copies of its recognition sequence.
Simultaneous binding of two DNA target sites stimulates DNA cleavage by MnlI.
4. A two-domain structure of the Type IIS restriction endonuclease MnlI has been identified by
limited proteolysis. An N-terminal domain of the enzyme mediates the sequence-specific
interaction with DNA and in part the dimerization of MnlI. It binds the recognition sequence
as a monomer, though it displays an ability to interact with two copies of the recognition
sequence as a dimer. A C-terminal domain of MnlI is determined to be monomeric in solution.
5. The C-terminal domain of MnlI REase resembles bacterial colicin nucleases in its oligomeric
state and requirement for alkaline earth as well as transition metal ions for double- and
single-stranded DNA cleavage activities.
6. The fusion of the non-specific HNH-type nuclease to the DNA binding domain had transformed
MnlI into a Mg2+-, Ni2+-, Co2+-, Mn2+-, Zn2+-, Ca2+-dependent sequence-specific enzyme.
Nevertheless, MnlI retains a residual single-stranded DNA cleavage activity controlled by its
C-terminal colicin-like nuclease domain. The cleavage of ds- and ssDNA by MnlI with a wide
range of divalent metal ions is unparalleled among restriction endonucleases characterized to
date.

<>

<1>Kriukiene, E., Lubiene, J., Lagunavicius, A., Lubys, A.
<2>MnlI - The member of H-N-H subtype of Type IIS restriction endonucleases.
<3>Biochim. Biophys. Acta
<4>1751
<5>194-204
<6>2005
<7>The Type IIS restriction endonuclease MnlI recognizes the non-palindromic nucleotide sequence
5'-CCTC(N)7/6 down arrow and
cleaves DNA strands as indicated by the arrow. The genes encoding MnlI
restriction-modification system were cloned and sequenced. It comprises
N6-methyladenine and C5-methylcytosine methyltransferases and the
restriction endonuclease. Biochemical studies revealed that MnlI
restriction endonuclease cleaves double- and single-stranded DNA, and
that it prefers different metal ions for hydrolysis of these
substrates. Mg2+ ions were shown to be required for the specific
cleavage of double-stranded DNA, whereas Ni2+ and some other transition
metal ions were preferred for nonspecific cleavage of single-stranded
DNA. The C-terminal part of MnlI restriction endonuclease revealed an
intriguing similarity with the H-N-H type nucleolytic domain of
bacterial toxins, Colicin E7 and Colicin E9. Alanine replacements in
the conserved sequence motif 306Rx(3)ExHHx(14)Nx(8)H greatly reduced
specific activity of MnlI, and some mutations even completely
inactivated the enzyme. However, none of these mutations had effect on
MnlI binding to the specific DNA, and on its oligomerisation state as
well. We interpret the presented experimental evidence as a suggestion
that the motif 306Rx(3)ExHHx(14)Nx(8)H represents the active site of
Mnll. Consequentially, MnlI seems to be the member of Type IIS with the
active site of the H-N-H type.

<>

<1>Kriukiene, E., Lubys, A.
<2>Comparison of two restriction-modification systems recognizing the same GGATCC sequence.
<3>Biologija
<4>1
<5>55-59
<6>1998
<7>Restriction-modification system Bsp98I from Bacillus species RFL98 recognizing the sequence
GGATCC has been cloned and sequenced.  The determined sequence predicts a methyltransferase of
388 amino acids, Mr 45 kDa, and a restriction endonuclease of 199 aa, Mr 22.5 kDa.
Comparisons of the predicted aa sequences indicated that Bsp98I and isoschizomeric BamHI
restriction endonucleases share 28% identity, whereas the corresponding methyltransferases
share 42% identity.  This observation suggests that Bsp98I and BamHI genes derive from a
common ancestor.  Regardless of such evolutionary relatedness, the Bsp98I R-M system has no
equivalent of the bamHIC gene which is involved in regulation of expression of BamHI R-M
genes.  This clearly indicates that the regulation of Bsp98I R-M genes is different.  Despite
the marginal similarity among R.BamHI and R.Bsp98I, the aa residues of catalytic/Mg2+ binding
center as well as the ones making the major groove contacts in R.BamHI can be found in
appropriate positions of the aligned aa sequence of R.Bsp98I.  In contrast, some amino acids
of R.BamHI that make contacts in the minor groove are absent in R.Bsp98I.

<>

<1>Kriukiene, E., Lubys, A.
<2>Cloning and analysis of DNA regions adjacent to methyltransferase Lsp1109I of restriction-modification system Lsp1109I, type IIs.
<3>Biologija
<4>2
<5>62-65
<6>1998
<7>DNA regions flanking genes lsp11091R? were cloned and sequenced.  Two open reading frames
transcribed in the same polarity as the lsp1109IMR? were found.  One of them, lsp-inv, encodes
a protein which has 41-46% of amino acids identical with proteins belonging to the family of
Din invertases.  This suggests that lsp-inv is possibly the gene of invertase Listeria sp.
Translation product of the second ORF (lsp-tnp), shares 23-26% homology with transposases of
various mobile DNA elements.  Possible function of these genes is discussed.  Analysis of
complementary DNA strand revealed four additional ORFs whose products possess no homology with
any aa sequence from EMBL data bank.

<>

<1>Kroft, B.S., Brown, E.W., Meng, J., Gonzalez-Escalona, N.
<2>Draft Genome Sequences of Two Salmonella Strains from the SARA Collection, SARA64 (Muenchen) and SARA33 (Heidelberg), Provide Insight into Their Antibiotic  Resistance.
<3>Genome Announcements
<4>1
<5>e00806-13
<6>2013
<7>The Salmonella enterica strains that are representatives of the S. enterica serovar
Typhimurium complex in reference collection A (SARA) are closely related
but exhibit differences in antibiotic resistance, which could have public health
consequences. To better understand the mechanisms behind these resistances, we
sequenced the genomes of two multidrug-resistant strains: SARA64 (Muenchen) and
SARA33 (Heidelberg).

<>

<1>Kroger, M., Blum, E., Deppe, E., Dusterhoft, A., Erdmann, D., Kilz, S., Meyer-Rogge, S., Mostl, D.
<2>Organization and gene expression within restriction-modification systems of Herpetosiphon giganteus.
<3>Gene
<4>157
<5>43-47
<6>1995
<7>We have characterized a family of related restriction-modification (R-M) systems from the soil
bacterium Herpetosiphon giganteus (Hgi).  A comparison of their genetic organization reveals
two types of regulatory proteins, called controlling ORF C.  While one of these small reading
frames derived from RM.HgiCI seems to be an enhancer of its own promoter, evidence is provided
for a silencer function of the other ORF C derived from the closely related AvaII-type
systems RM.HgiBI/CII/EI.  The respective silencer function is detected during our various
attempts to clone three isoschizomers with unusually high differences in their specific
activity.  Sequencing and site-directed mutagenesis revealed just two amino acids as being
responsible for a massive increase in specific activity of these endonucleases.

<>

<1>Kroger, M., Hobom, G.
<2>Restriction enzyme HgiCI characterization of the 6-nucleotide staggered cut sequence and its application in mismatch cloning.
<3>Methods Enzymol.
<4>155
<5>3-10
<6>1987
<7>The gliding bacterium Herpetosiphon giganteus became one of the most
intensively screened groups of organisms in the search for new restriction
enzymes.  Among the 10 strains tested, 17 enzymes could be found with seven
different but related recognition sequences.  This led to a hypothesis
regarding the evolutionary relationship among these enzymes and could be a
basis for a better understanding of the biochemical mechanism of restriction
enzymes-DNA target interaction.  Among these enzymes HgiCI is remarkably
different from all other previously described endonucleases, since it produces
5'-hexanucleotide protruding ends.  Combined with the fact that HgiCI
recognizes a degenerate sequence, specific applications of this enzymatic
activity in gene technology are possible.  Usually, for specific base pairing
within 5'- or 3'- protruding ends, a match of 2 bp is fair, while four matching
base pairs lead to highly efficient ligase reactions.  Since a perfect match of
6 bp may not be required, we used HgiCI-restricted DNA fragments in order to
test whether DNA ligase reactions among hexanucleotide protruding ends could
proceed in spite of some mismatch positions.  Our results presented here allow
the conclusion that is is possible to obtain mismatched ligase reaction
products in considerable fractions.  A wider application of this observation
seems possible, since an isoschizomer of HgiCI BanI, is available commercially
and is obtained from an unrelated strain Bacillus aneurinolyticus (IAM 1077).
In contrast to the data given in the literature, we have determined via
cross-ligation that BanI also produces 5'-hexanucleotide protruding DNA
fragments.  In this article we intend to focus on the methodology used to
characterize recognition sequences and on the application of HgiCI (BanI)
fragment ends in mismatch cloning rather than on enzyme purification
procedures.

<>

<1>Kroger, M., Hobom, G.
<2>The nucleotide sequence recognized by the Escherichia coli A restriction and modification enzyme.
<3>Nucleic Acids Res.
<4>12
<5>887-899
<6>1984
<7>The nucleotide recognition sequence for the restriction-modification enzyme of
Escherichia coli A (EcoA) has been determined to be GAG-7N-GTCA.  This sequence
is fairly similar, but distinctly different from the two other type I
restriction enzyme recognition sites known for E. coli B and E. coli K12,
respectively.  N6-adenosine methylation has been observed at nucleotide
positions 2 and 12 within that sequence after modification by EcoA.  As a
reference point for mapping the single EcoA site in lambda, the position of
lambda point mutation Oam29 has been determined also.

<>

<1>Kroger, M., Hobom, G., Schutte, H., Mayer, H.
<2>Eight new restriction endonucleases from Herpetosiphon giganteus - divergent evolution in a family of enzymes.
<3>Nucleic Acids Res.
<4>12
<5>3127-3141
<6>1984
<7>Characterization of eight restriction endonucleases isolated from five strains
of Herpetosiphon giganteus is described.  HgiCI from strain Hpg9 recognizes and
cleaves the degenerate sequence: 7GGPyPuCC, producing 5'-hexanucleotide
protruding ends.  Endonucleases HgiBI, HgiCII and HgiEI are isoschizomers of
AvaII; HgiCIII and HgiDII are isoschizomers of SalI; and HgiDI and HgiGI are
isoschizomers of AcyI.  Based upon their closely related and in part
overlapping recognition specificities a close evolutionary relationship is
proposed for all known Hgi restriction endonucleases.

<>

<1>Krohn, K., Aapola, U., Scott, H., Antonarakis, S., Shimizu, N., Kudoh, J., Peterson, P.
<2>Structure, expression and diagnostic and therapeutic use of a human gene DNMT3L encoding a DNA methyltransferase.
<3>International Patent Office
<4>WO 0127249 A
<5>54
<6>2001
<7>The present invention relates to a novel gene, a novel protein encoded by said gene, a
corresponding mRNA of the gene and to diagnostic and therapeutic uses of the gene or a mutated
form thereof.  More specifically, the present invention relates to a novel gene which is
useful in the characterization and mutation search, and in the development of the diagnosis
and therapy of hereditary diseases linked to locus 21q22.3 and in the development of the
diagnosis and therapy of tumors.  The novel gene and protein are also useful in the diagnosis
and therapy of infertility and in the development of means for the control of reproduction.

<>

<1>Kroon, A.M., Bakker, H., Holtrop, M., Terpstra, P.
<2>The restriction endonuclease cleavage map of rat liver mitochondrial DNA.
<3>Biochim. Biophys. Acta
<4>474
<5>61-68
<6>1977
<7>Mitochondrial DNA from rat liver contains six sites for cleavage by the
restriction endonucleases HindIII and EcoRI.  A large stretch of DNA,
comprising about 40% of the mitochondrial genome is not cleaved by either of
the enzymes; eight cleavage sites are located on a DNA stretch of 35% of the
genome length suggestive of an unequal distribution of the A-T basepairs over
the molecule.  The number of HindIII and EcoRI fragments is much higher than
reported for other mammalian mitochondrial DNAs up to now.

<>

<1>Kropinski, A.M., Arutyunov, D., Foss, M., Cunningham, A., Ding, W., Singh, A., Pavlov, A.R., Henry, M., Evoy, S., Kelly, J., Szymanski, C.M.
<2>Genome and Proteome of Campylobacter jejuni Bacteriophage NCTC 12673.
<3>Appl. Environ. Microbiol.
<4>77
<5>8265-8271
<6>2011
<7>Campylobacter jejuni continues to be the leading cause of bacterial food-borne illness
worldwide, so improvements to current methods used
for bacterial detection and disease prevention are needed. We describe
here the genome and proteome of C. jejuni bacteriophage NCTC 12673 and
the exploitation of its receptor-binding protein for specific bacterial
detection. Remarkably, the 135-kb Myoviridae genome of NCTC 12673
differs greatly from any other proteobacterial phage genome described
(including C. jejuni phages CP220 and CPt10) and instead shows closest
homology to the cyanobacterial T4-related myophages. The phage genome
contains 172 putative open reading frames, including 12 homing
endonucleases, no visible means of packaging, and a putative
trans-splicing intein. The phage DNA appears to be strongly associated
with a protein that interfered with PCR amplification and estimation of
the phage genome mass by pulsed-field gel electrophoresis.
Identification and analyses of the receptor-binding protein (Gp48)
revealed features common to the Salmonella enterica P22 phage tailspike
protein, including the ability to specifically recognize a host
organism. Bacteriophage receptor-binding proteins may offer promising
alternatives for use in pathogen detection platforms.

<>

<1>Kropinski, A.M., Kovalyova, I.V., Billington, S.J., Patrick, A.N., Butts, B.D., Guichard, J.A., Pitcher, T.J., Guthrie, C.C., Sydlaske, A.D., Barnhill, L.M., Havens, K.A., Day, K.R., Falk, D.R., McConnell, M.R.
<2>The genome of epsilon 15, a serotype-converting, group E1 Salmonella enterica-specific bacteriophage.
<3>Virology
<4>369
<5>234-244
<6>2007
<7>The genome sequence of the Salmonella enterica serovar Anatum-specific, serotype-converting
bacteriophage 05 has been completed. The
nonredundant genome contains 39,671 bp and 51 putative genes. It most
closely resembles the genome of phi V10, an Escherichia coli O157:H7
specific temperate phage, with which it shares 36 related genes. More
distant relatives include the Burkholderia cepacia-specific phage,
BccpC6B (8 similar genes), the Bordetella bronchiseptica-specific
phage, BPP-1 (8 similar genes) and the Photobacterium profundum
prophage, P P phi pr1 (6 similar genes).
epsilon 15 gene identifications based on homologies with known gene
families include the terminase small and large subunits, integrase,
endolysin, two holins, two DNA methylase enzymes (one adenine-specific
and one cytosine-specific) and a RecT-like enzyme. Genes identified
experimentally include those coding for the serotype conversion
proteins, the tail fiber, the major capsid protein and the major
repressor. epsilon 15's attP site and the Salmonella attB site with
which it interacts during lysogenization have also been determined.

<>

<1>Kropp, K.A., Lucid, A., Carroll, J., Belgrudov, V., Walsh, P., Kelly, B., Smith, C., Dickinson, P., O'Driscoll, A., Templeton, K., Ghazal, P., Sleator, R.D.
<2>Draft Genome Sequence of a Serratia marcescens Strain Isolated from a Preterm Neonatal Blood Sepsis Patient at the Royal Infirmary, Edinburgh, Scotland, United  Kingdom.
<3>Genome Announcements
<4>2
<5>e00908-14
<6>2014
<7>Herein, we report the draft genome sequence for isolate ED-NGS-1015 of Serratia marcescens,
cultivated from a blood sample obtained from a neonatal sepsis
patient at the Royal Infirmary in Edinburgh, Scotland, United Kingdom.

<>

<1>Kropp, K.A., Lucid, A., Carroll, J., Belgrudov, V., Walsh, P., Kelly, B., Smith, C., Dickinson, P., O'Driscoll, A., Templeton, K., Ghazal, P., Sleator, R.D.
<2>Draft Genome Sequence of an Enterococcus faecalis Strain Isolated from a Neonatal Blood Sepsis Patient.
<3>Genome Announcements
<4>2
<5>e00907-14
<6>2014
<7>Herein, we report the draft genome sequence of Enterococcus faecalis ED-NGS-1009, cultivated
from a blood sample taken from a neonatal sepsis patient at the Royal
Infirmary in Edinburgh, Scotland, United Kingdom.

<>

<1>Kropp, K.A., Lucid, A., Carroll, J., Belgrudov, V., Walsh, P., Kelly, B., Smith, C., Dickinson, P., O'Driscoll, A., Templeton, K., Ghazal, P., Sleator, R.D.
<2>Draft Genome Sequence of a Staphylococcus warneri Strain Isolated from a Preterm  Neonate Blood Sepsis Patient at the Royal Infirmary, Edinburgh, Scotland.
<3>Genome Announcements
<4>2
<5>e00877-14
<6>2014
<7>Herein, we report the draft genome sequence of Staphylococcus warneri ED-NGS-1001, cultivated
from a blood sample taken from a preterm neonate blood
sepsis patient at the Royal Infirmary, Edinburgh, Scotland, United Kingdom.

<>

<1>Kropp, K.A., Lucid, A., Carroll, J., Belgrudov, V., Walsh, P., Kelly, B., Smith, C., Dickinson, P., O'Driscoll, A., Templeton, K., Ghazal, P., Sleator, R.D.
<2>Draft Genome Sequence of a Streptococcus agalactiae Strain Isolated from a Preterm Neonate Blood Sepsis Patient at the Royal Infirmary, Edinburgh, Scotland.
<3>Genome Announcements
<4>2
<5>e00875-14
<6>2014
<7>Herein, we report the draft genome sequence of Streptococcus agalactiae ED-NGS-1000,
cultivated from a blood sample taken from a preterm neonate blood
sepsis patient at the Royal Infirmary, Edinburgh, Scotland, United Kingdom.

<>

<1>Kropp, K.A., Lucid, A., Carroll, J., Belgrudov, V., Walsh, P., Kelly, B., Templeton, K., Smith, C., Dickinson, P., O'Driscoll, A., Ghazal, P., Sleator, R.D.
<2>Draft Genome Sequence of a Staphylococcus aureus Isolate Taken from the Blood of  a Preterm Neonatal Blood Sepsis Patient.
<3>Genome Announcements
<4>2
<5>e00906-14
<6>2014
<7>Herein, we report the draft genome sequence of Staphylococcus aureus ED-NGS-1006, cultivated
from a blood sample taken from a neonatal sepsis patient at the Royal
Infirmary in Edinburgh, Scotland, United Kingdom.

<>

<1>Kropp, K.A., Lucid, A., Carroll, J., Belgrudov, V., Walsh, P., Kelly, B., Templeton, K., Smith, C., Dickinson, P., O'Driscoll, A., Ghazal, P., Sleator, R.D.
<2>Draft Genome Sequence of a Pantoea sp. Isolated from a Preterm Neonatal Blood Sepsis Patient.
<3>Genome Announcements
<4>2
<5>e00904-14
<6>2014
<7>Herein, we report the draft genome sequence of Pantoea sp. ED-NGS-1003, cultivated from a
blood sample taken from a neonatal sepsis patient at the Royal
Infirmary, Edinburgh, Scotland, United Kingdom.

<>

<1>Krueger, D.H., Kupper, D., Meisel, A., Reuter, M., Schroeder, C.
<2>The significance of distance and orientation of restriction endonuclease recognition sites in viral DNA genomes.
<3>FEMS Microbiol. Rev.
<4>17
<5>177-184
<6>1995
<7>Studies on phage T3 and T7 have shown that these viruses avoid restriction not only by the
phage-coded Ocr (and S-adenosylmethionine hydrolase) protein functions or by the complete loss
of specific recognition sites for certain restriction endonucleases from their genomes, but
also that there are two additional modes: resistance towards EcoP15 (which recognizes a non-
symmetrical sequence) is achieved by an identical orientation of all the recognition sites in
the virus genome (strand bias) and in the case of EcoRII by the extreme reduction in number
and thereby greater distance between recognition sites in the genome.  These observations led
to the discovery that certain restriction endonucleases require the simultaneous cooperation
with two DNA sites for their function, as well as to the ongoing elucidation of the molecular
modes of action of these enzymes.  Type II and type III enzymes display fundamentally
different mechanisms of protein- DNA interaction.  For EcoRII we favor a model of simultaneous
binding of two DNA sites to a dimeric enzyme molecule (neighboring sites of the same, looping,
DNA molecule or sites located on different DNA molecules), while the action of EcoP15 seems to
conform with a tracking- collision model of two enzyme molecules bound to inversely oriented
recognition sites.  In addition to podoviruses T3 and T7, strand bias of recognition sequences
for different type III DNA modification-restriction enzymes is also observed in the inoviruses
M13, IKE and PF3.

<>

<1>Krueger, D.H., Kupper, D., Meisel, A., Tierlich, M., Reuter, M., Schroeder, C.
<2>Restriction endonucleases functionally interacting with two DNA sites.
<3>Gene
<4>157
<5>165
<6>1995
<7>Simultaneous interaction with two recognition sites was found to be a precondition for DNA
cleavage by certain type-II and type-III restriction endonucleases.  Nevertheless, the
molecular mechanisms of the protein-DNA interaction are different between members of both
classes of enzymes.

<>

<1>Krueger, D.H., Moencke-Buchner, E., Mackeldanz, P., Reuter, M.
<2>Cloning and large-scale purification of recombinant restriction endonuclease EcoP15I for use in analysis of gene expression.
<3>European Patent Office
<4>EP 1584677 A
<5>
<6>2005
<7>The present invention relates to a recombinant DNA which encodes a storable EcoP15I type III
restriction endonuclease as well as large-scale purification of EcoP15I to near homogeneity
and the use of said EcoP15I as tool for serial analysis of gene expression and/or the
identification of corresponding genes.

<>

<1>Krueger, D.H., Presber, W., Hansen, S., Rosenthal, H.A.
<2>Biological functions of the bacteriophage T3 SAMase gene.
<3>J. Virol.
<4>16
<5>453-455
<6>1975
<7>Certain differences between phage T3 on the one hand and T3sam and T7 on the other hand
indicate that the T3-coded SAMase function is responsible (i) for the development of the
pseudolysogenic state by preventing T3 DNA methylation, and (ii) for the partial protection of
the phage DNA against restriction by the P system.

<>

<1>Krueger, D.H., Reuter, M.
<2>Reliable detection of DNA cytosine methylation at CpNpG sites using the engineered restriction enzyme EcoRII-C.
<3>Biotechniques
<4>38
<5>855-856
<6>2005
<7>Methylation of cytosine to 5-methylcytosine (mC) is the most important epigenetic DNA
alteration in eukaryotes.  Cytosine methylation is involved in establishing a silenced
chromatin stage through interaction with DNA-binding proteins and recruitment of histone
deacetylases and other histone-modifying enzymes leading to chromatin remodeling.  In this
fashion, DNA methylation is connected with processes of normal and pathological gene
regulation, DNA replication, virus latency, parental imprinting embryonic development,
carcinogenesis, and genetic diseases in higher organisms.  Analogous to the terms
transcriptome and proteome, the neologism methylome has been proposed to describe the complete
set of DNA methylation in a cell, which carries information in addition to the naked DNA
sequence.  The methylome changes over time and, depending on its alterations, is linked to
aging, cancer, and polymorphic variation in populations.

<>

<1>Krueger, M., Meyerdierks, A., Gloeckner, F.O., Amann, R., Widdel, F., Kube, M., Reinhardt, R., Kahnt, J., Boecher, R., Thauer, R.K., Shima, S.
<2>A conspicuous nickel protein in microbial mats that oxidize methane anaerobically.
<3>Nature
<4>426
<5>878-881
<6>2003
<7>Anaerobic oxidation of methane (AOM) in marine sediments is an important microbial process in
the global carbon cycle and in control of greenhouse
gas emission. The responsible organisms supposedly reverse the reactions
of methanogenesis, but cultures providing biochemical proof of this have
not been isolated. Here we searched for AOM-associated cell components in
microbial mats from anoxic methane seeps in the Black Sea. These mats
catalyse AOM rather than carry out methanogenesis. We extracted a
prominent nickel compound displaying the same absorption spectrum as the
nickel cofactor F430 of methyl-coenzyme M reductase, the terminal enzyme
of methanogenesis; however, the nickel compound exhibited a higher
molecular mass than F430. The apparent variant of F(430) was part of an
abundant protein that was purified from the mat and that consists of three
different subunits. Determined amino-terminal amino acid sequences matched
a gene locus cloned from the mat. Sequence analyses revealed similarities
to methyl-coenzyme M reductase from methanogenic archaea. The abundance of
the nickel protein (7% of extracted proteins) in the mat suggests an
important role in AOM.

<>

<1>Kruger, D., Butkus, V., Reuter, M., Janulaitis, A., Petrusyte, M., Pein, C.D., Hansen, S., Cech, D.
<2>Rapid and complete degradation of DNA with restriction endonuclease - by adding second DNA sequence containing the appropriate recognition site, for cleavage of phage, vector etc.
<3>East German Patent Office
<4>DD 299440 A
<5>
<6>1992
<7>DNA molecules (I) which are incompletely cleaved by particular restriction endonucleases (RE)
when using known methods, are completely and rapidly degraded into specific fragments by
adding a second DNA species (II). (II) is a synthetic oligonucleotide duplex or a biologically
replicable DNA molecule (or their fragments) having recognition sites for the particular RE
and carrying no specific methylation.

<>

<1>Kruger, D., Butkus, V., Reuter, M., Pein, C., Hansen, S., Schroeder, C.
<2>Procedure for characterizing known DNA sequences.
<3>East German Patent Office
<4>DD 293139 A
<5>
<6>1991
<7>The finding concerns a procedure for characterizing known DNA sequences. The finding is
applicable in the fields of Molecular Biology and Genetics. The essence is to incubate a DNA
molecule, where unmethylated recognition sites will be digested by a specific restriction
endonuclease, in a reaction mixture with a second, unmethylated DNA-molecule (especially
oligonucleotide-duplexes, which includes the recognition sequence). In this way the
methylating specific restriction endonuclease will be forced to cut all unmethylated
recognition sequences, while specific recognition sequences which are methylated will not be
attacked. This method is very favorable for the detection of the Dcm-methylation with the help
of the EcoRII restriction endonuclease.

<>

<1>Kruger, D.H.
<2>Methylases as regulators of gene activity.
<3>Wiss. Fortsch.
<4>37
<5>268-270
<6>1987
<7>None

<>

<1>Kruger, D.H.
<2>Methylation of DNA as a molecular biological regulatory signal.
<3>Biol. Zentralbl.
<4>107
<5>257-266
<6>1988
<7>Methylation of adenine or cytosine adds information to the DNA double strand
which can influence the interactions between DNA and proteins.  Besides the
protection of DNA against corresponding restriction endonucleases in
prokaryotic cells, a number of new functions of DNA methylation in prokaryotic
and eukaryotic cells have been demonstrated.  These observations are mainly
related to the regulation of gene expression.  In gene technology in vitro,
artificial DNA methylation is exploited to change the number of sensitive
recognition sites for certain restriction endonucleases.  Finally, the
properties of the first natural inhibitory protein (Ocr) against DNA
methylation and the possible significance of cellular effector molecules
specifically blocking or enhancing DNA methylation are discussed.

<>

<1>Kruger, D.H.
<2>Host-controlled modification and restriction.
<3>Encyclop. Virol., Academic Press, Webster, R.G, Granoff, A, 
<4>0
<5>669-674
<6>1994
<7>The application of restriction endonucleases has revolutionized molecular biological research.
As so often was the case, studies on bacterial viruses led to the discovery of this class of
enzyme. At the beginning of the fifties, the observation was noted that phages 'remember'
the last host strain in which they reproduced. Depending on the host, the virus carries a
specific modification which usually improves its ability to grow on the same host in
subsequent rounds of infection, while impairing its growth in others. Host-controlled
modification results in a reversible phenotype, clearly distinguishable from irreversible
mutational changes in host range.

The molecular explanation of these phenomena (first demonstrated for phage lambda and P2)
was elaborated in the 60s and 70s by groups of W. Arber, H.O. Smith, M. Meselson and others.
Host-controlled modification consists of a specific DNA methylation at a defined recognition
sequence of 4-8 bp. Restriction is caused by DNA cleavage occurring at an unmethylated
recognition site. Pairs of corresponding DNA methylases and restriction endonucleases (i.e.
recognizing the same site) coded by hsd genes (host specificity of DNA) in bacterial cells are
responsible for these activities. During DNA replication, recognition sequences in nascent
daughter strands are transiently unmethylated. Such hemimethylated sites are preferential
substrates for DNA methylases, while sites unmethylated in both strands are primary targets of
restriction endonucleases and are very rarely methylated.

DNA modification consists of a C-5 or N-4 methylation of cytosine or an N-6 methylation of
adenine in the recognition sequence. S-Adenosylmethionine (AdoMet) functions as the methyl
donor. DNA restriction results in fragments with cohesive or blunt ends which are further
degraded intracellularly by other nucleases, in particular by the recBCD=exoV enzyme.

DNA modification/restriction (M/R) systems ae ubiquitous in the procaryote world but there
is no unequivocal evidence of their existence in higher eucaryotes.


<>

<1>Kruger, D.H., Barcak, G.J., Reuter, M., Smith, H.O.
<2>EcoRII can be activated to cleave refractory DNA recognition sites.
<3>Nucleic Acids Res.
<4>16
<5>3997-4008
<6>1988
<7>EcoRII restriction sites [5'-CC(A/T)GG] in phage T3 and T7 DNA are refractory
to cleavage by EcoRII, but become sensitive to cleavage in the presence of DNAs
which contain an abundance of EcoRII sensitive sites (e.g. pBR322 or lambda
DNA).  Studies using fragments of pBR322 containing different numbers of EcoRII
sites show that the susceptibility to EcoRII cleavage is proportional to the
number of sites in the individual fragment.  We postulate that EcoRII is the
prototype of restriction endonucleases which require at least 2 simultaneously
bound substrate sites for their activation.  EcoRII sites are refractory when
they occur at relatively low frequency in the DNA.  The restriction enzyme can
be activated by DNA with a higher frequency of sites.

<>

<1>Kruger, D.H., Barcak, G.J., Smith, H.O.
<2>Abolition of DNA recognition site resistance to the restriction endonuclease EcoRII.
<3>Biomed. Biochim. Acta
<4>47
<5>K1-K5
<6>1988
<7>The EcoRII recognition sites 5'-CC(A/T)GG-3' which occur three times in T3 DNA
are refractory to this restriction endonuclease.  However, these sites are
specifically cleaved by EcoRII when a second, susceptible DNA species (pBR322,
phage lambda) is present.

<>

<1>Kruger, D.H., Bickle, T.A.
<2>Bacteriophage Survival:  Multiple mechanisms for avoiding the deoxyribonucleic acid restriction systems of their hosts.
<3>Microbiol. Rev.
<4>47
<5>345-360
<6>1983
<7>None

<>

<1>Kruger, D.H., Bickle, T.A., Reuter, M., Pein, C.-D., Schroeder, C.
<2>DNA methylation and restriction processes in Escherichia coli: Insights by use of bacterial viruses T3 and T7.
<3>Nucleic Acid Methylation, Alan R. Liss, Inc., Clawson, G.A., Willis, D.B., Weissbach, A., Jones, P.A., 
<4>0
<5>113-124
<6>1990
<7>DNA modification-restriction enzymes of Escherichia coli cells can be grouped into 3 families,
called type I, II, and III. The ocr+ gene of T7 encodes the first known inhibitor protein
specifically blocking a group of methylases and endonucleases which, in particular, belong to
the type I (EcoB, EcoK) enzymes. The appearance of recognition sites for type II enzymes
(e.g., EcoRII, methylases EcoDam and EcoDcm) is strongly counterselected in the T7 genome,
furthermore, there are additional mechanisms preventing the enzymes from acting on the
remaining sites. For instance, EcoRII is a restriction enzyme which requires the coordinated
presence of at least 2 recognition sites in the substrate DNA for its activity. Sites which do
not fulfill this requirement are refractory to EcoRII but can be cleaved by this enzyme in the
presence of a second, susceptible DNA species or oligonucleotides. The resistance of T7 DNA
towards the type III enzyme EcoP15 is explained by the absolute strand bias of the
non-symmetric sites (in which only 1 strand can be methylated) in the DNA molecule. These data
allow some understanding of how the endogenous DNA in EcoP15-encoding cells could be protected
against self-restriction during replication. The frequency and polarity of recognition sites
in the DNA molecula may play a critical role in the function of certain restriction
endonucleases and methylases.

<>

<1>Kruger, D.H., Bickle, T.A., Reuter, M., Pein, C.D., Schroeder, C.
<2>Studies on DNA methylation and restriction processes by use of bacterial virus T7.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>200
<6>1989
<7>DNA modification-restriction enzymes of Escherichia coli cells can be grouped into 3 families,
called type I,II, and III. The ocr+ gene of T7 encodes the first known inhibitor protein
specifically blocking a group of methylases and endonucleases which, in particular, belong to
the type I (EcoB, EcoK) enzymes. The appearance of recognition sites for type II enzymes
(e.g., EcoRII, methylases EcoDam and EcoDcm) is strongly counterselected in the T7 genome,
futhermore, there are additional mechanisms preventing the enzymes from acting on the
remaining sites. For instance, EcoRII is a restriction enzyme which requires the coordinated
presence of at least 2 recognition sites in the substrate DNA for its activity. Sites which do
not fulfill this requirement are refractory to EcoRII but can be cleaved by this enzyme in the
presence of a second, susceptible DNA species or oligonucleotides (CDP et al., submitted). The
resistance of T7 DNA towards the type III enzyme EcoP15 is explained by the absolute strand
bias of the non-symmetric sites (in which only one strand can be methylated) in the DNA
molecule (4; CS et al., in prep.). These data allow some understanding of how the endogenous
DNA in EcoP15-encoding cells could be protected against self-restriction during replication.
The frequency and polarity of recognition sites in the DNA molecule may play a critical role
the function of certain restriction endonucleases and methylases.

<>

<1>Kruger, D.H., Chernin, L.S., Hansen, S., Rosenthal, H.A., Goldfarb, D.M.
<2>Protection of foreign DNA against host-controlled restriction in bacterial cells.
<3>Mol. Gen. Genet.
<4>159
<5>107-110
<6>1978
<7>Foreign F'lac plasmid DNA which is introduced into potentially restricting E.
coli recipient cells can be protected from restriction by preinfecting the
recipient cells with UV-inactivated T3 or T7 bacteriophages which express the
ocr gene function.  The recipient cells survive and are able to replicate
themselves as well as the newly acquired plasmid.

<>

<1>Kruger, D.H., Gola, G., Weisshuhn, I., Hansen, S.
<2>The ocr gene function of bacterial viruses T3 and T7 prevents host-controlled modification.
<3>J. Gen. Virol.
<4>41
<5>189-192
<6>1978
<7>On pre-infection of the host Escherichia coli B with u.v.-inactivated T3 or T7
phage able to express their early genes (like 0.3), B-specific modification of
super-infecting, successfully multiplying viruses does not take place.  the ocr
gene function (gene 0.3) of T3 and T7 not only prevents host-specific DNA
restriction but also modification, probably by inhibiting the same late step in
the interaction between the restriction enzyme and DNA.

<>

<1>Kruger, D.H., Hansen, S., Presber, W.
<2>Host-controlled modification and restriction of bacteriophage T7 by various Escherichia coli B strains in vivo.
<3>Z. Allg. Mikrobiol.
<4>16
<5>73-76
<6>1976
<7>It is known for more than 20 years that phages possess a "memory" for the host strain on which
they grew last.  For instance, lambda phages grown on E. coli K are restricted in E. coli B
cells, i.e. their efficiency of plating on E. coli B cells is several orders of magnitude
below their e.o.p. on E. coli K host cells.  This effect is reversible, phages grown on E.
coli B are restricted by E. coli K but grow well on E. coli B.  Therefore, the phage DNA is
not mutated but rather modified or - in absence of the right modification - restricted by the
host.  For phage lambda the molecular basis of this modification/restriction consists in a
host-specific methylation or an endonucleolytic DNA degradation at specific recognition sites
of the phage DNA.  Meanwhile, the M/R of DNA (not only of phage DNA!) has been recognized as a
general principle in molecular genetics with wide practical applications.

<>

<1>Kruger, D.H., Hansen, S., Reuter, M.
<2>The ocr+ gene function of bacteriophages T3 and T7 counteracts the Salmonella typhimurium DNA restriction systems SA and SB.
<3>J. Virol.
<4>45
<5>1147-1149
<6>1983
<7>In host cells containing the Salmonella typhimurium DNA
restriction-modification systems SA+ and SB+, replication of the ocr+
bacteriophages T3 and T7 is not impaired.  However, ocr(gene 0.3) mutants of
these phages are susceptible to DNA restriction and modification by the SA+ and
SB+ systems.

<>

<1>Kruger, D.H., Presber, W., Hansen, S., Rosenthal, H.A.
<2>Different restriction of bacteriophages T3 and T7 by P1-lysogenic cells and the role of the T3-coded SAMase.
<3>Z. Allg. Mikrobiol.
<4>17
<5>581-591
<6>1977
<7>The intracellular growth of the phages T3 and T7 is restricted in the
presence of the Escherichia coli prophage P1.  Phage T3 has a higher ability to express its
genome and to damage the host cell than T7.  This partial protection of T3 against P1
restriction is due to the T3-coded SAMase, an enzyme which degrades S-
adenosylmethionine, the cofactor of the P1 restriction endonuclease.  Since we did not
observe DNA cleavage in vivo, we conclude that the in vivo action of the P1 nuclease is
limited to a SAM-dependent repressor-like binding to T3 and T7 DNA, while further
reactions with the DNA (modification vs cleavage) are blocked.

<>

<1>Kruger, D.H., Prosch, S., Reuter, M., Goebel, W.
<2>Cloning of the resistant EcoRII recognition site of phage T7 into an EcoRII-sensitive plasmid makes the site susceptible to the restriction enzyme.
<3>J. Basic Microbiol.
<4>9
<5>679-683
<6>1990
<7>The recognition sequence 5'-CC(A/T)GG for EcoRII in the bacteriophage T7 genome
is refractory to this restriction endonuclease, despite not bearing the
specific (protective) methylation.  Following the integration of this site as
part of a 219 bp fragment (in which the recognition sequence is flanked by
about 100 bp of T7 origin) into the EcoRII-sensitive vector pUC18, the T7 site
becomes susceptible to cleavage, too.  The same is true of recombinant pBR322
plasmids containing the T7-derived recognition site.  The results show that the
flanking sequences are not immediately responsible for the refractory behaviour
of EcoRII sites and are in agreement with data according to which EcoRII
requires the coordinated presence of at least two recognition sites in its DNA
substrate.

<>

<1>Kruger, D.H., Reuter, M., Hansen, S., Schroeder, C.
<2>Influence of phage T3 and T7 gene functions on a Type III (EcoP1) DNA restriction-modification system in vivo.
<3>Mol. Gen. Genet.
<4>185
<5>457-461
<6>1982
<7>The ocr+ gene function (gp 0.3) of bacteriophages T3 and T7 not only
counteracts type I (EcoB, EcoK) but also type III restriction endonucleases
(EcoP1).  Despite the presence of recognition sites, phage DNA as well as
simultaneously introduced plasmid DNA are protected by ocr+ expression against
both the endonucleolytic and the methylating activities of the EcoP1 enzyme.
Nevertheless, the EcoP1 protein causes the exclusion of T3 and T7 in
P1-lysogenic cells, apparently by exerting a repressor-like effect on phage
gene expression.  T3 which induces an S-adenosylmethionine hydrolase is less
susceptible to the repressor effect of the SAM-stimulated EcoP1 enzyme.  The
abundance of EcoP1 recognition sites in the T7 genome is explained by their
near identity with the T7 DNA primase recognition site.

<>

<1>Kruger, D.H., Reuter, M., Schroeder, C., Glatman, L.I., Chernin, L.S.
<2>Restriction of bacteriophage T3 and T7 ocr+ strains by the type II restriction endonuclease EcoRV.
<3>Mol. Gen. Genet.
<4>190
<5>349-351
<6>1983
<7>When E. coli cells carrying the plasmid pLG13 coding for the newly discovered
type II restriction endonuclease EcoRV are infected with phage T3 or T7, only
T7 is able to replicate normally. T3 wild-type as well as its mutants are
subject to DNA restriction in vivo and in vitro.  The EcoRV enzyme cuts T3 DNA
at 5 sites.  T7 and its ocr- mutants have no EcoRV sites in their DNA.  In
contrast to the anti-restriction acitivity of the T3 and T7 ocr+ gene functon
against type I and III restriction enzymes, the ocr+ protein is unable to
inactivate the type II restriction endonuclease EcoRV.

<>

<1>Kruger, D.H., Schroeder, C., Hansen, S., Rosenthal, H.A.
<2>Active protection by bacteriophages T3 and T7 against E. coli B- and K-specific restriction of their DNA.
<3>Mol. Gen. Genet.
<4>153
<5>99-106
<6>1977
<7>The bacteriophages T3 and T7 are not modified and restricted by E. coli
strains with different host specificity (E. coli B, K, 0) in vivo.  The phages code for a gene
product with the ability to overcome classical restriction (ocr): ocr- mutants are subject to
modification and restriction via DNA methylation vs cleavage.  The T3 genome possesses
recognition sites for the restriction endonuclease R.EcoB which, unless the DNA is B-
specifically modified, trigger 5-7 DNA cleavages.  The ocr gene function of T3 and T7 is
located within the gene 0.3 region of these phages and is not identical with the sam
(SAMase) function of T3.  The mechanism of ocr protection remains unclear, while it is
certain that this protection by the gene 0.3 protein is exerted in the infected cell and not
through "over-all" modification in the preceding growth cycle of the phage.

<>

<1>Kruger, D.H., Schroeder, C., Reuter, M., Bickle, T.A., Bogdarina, I.G., Buryanov, Y.I.
<2>Use of bacterial virus T7 as a tool for the study of DNA methylation.
<3>Gene
<4>74
<5>85-87
<6>1988
<7>Meeting Abstract

<>

<1>Kruger, D.H., Schroeder, C., Reuter, M., Bogdarina, I.G., Buryanov, Y.I., Bickle, T.A.
<2>DNA methylation of bacterial viruses T3 and T7 by different DNA methylases in Escherichia coli K12 cells.
<3>Eur. J. Biochem.
<4>150
<5>323-330
<6>1985
<7>We have investigated the susceptibility of the genomes of the related bacteriophages T3 and T7
to the three major DNA methyltransferases (EcoK, dam, dcm) of their host, Escherichia coli
K12. In vivo the EcoK host specificity enzyme only methylates the DNA of ocr phages. This is
due to an inhibition of the enzyme by the phage ocr gene product, which had previously been
shown to be an inhibitor of the restriction endonuclease. EcoK-specific DNA methylation
protects the ocr viruses after one growth cycle on these host cells against the action of
corresponding restriction endonuclease EcoK. Owing to the unique S-adenosyl-L-methionine
hydrolase (sam) activity of the T3-coded ocr protein, the T3 DNA is absolutely devoid of the
methylated bases 6-methylaminopurine and 5-methylcytosine. In contrast to this, T7 derivatives
and sam- derivatives of T3 carry a small number of about 2-4 molecules 6-methylaminopurine and
5-methylcytosine per genome. The presence of 6-methylaminopurine is due to dam methylation,
though the majority of dam sites remain unmethylated. In vivo as well as in vitro the ocr
protein has no influence on the activities of the dam and dcm methylase. The experiments gave
some evidence for the existence of a second cytosine methylase in E. coli K12. Besides dam and
dcm recognition sites being undermethylated, their absolute number in T3 and T7 DNAs is far
below the expected value. Moreover, one of the two dcm sites present in T7 (Studier strain) is
missing in our T7 strain owing to a 1300-base-pair deletion in gene 0.7.

<>

<1>Kruger, D.H., Schroeder, C., Santibanez-Koref, M., Reuter, M.
<2>Avoidance of DNA methylation:  A virus-encoded methylase inhibitor and evidence for counterselection of methylase recognition sites in viral genomes.
<3>Cell Biophys.
<4>15
<5>87-95
<6>1989
<7>The ocr+ gene of bacterial virus T7 codes for the first protein recognized to
inhibit a specific group of DNA methylases.  The recognition sequences of
several other DNA methylases, not susceptible to Ocr inhibition, are
significantly suppressed in the virus genome.  The bacterial virus T3 encodes
an Ado-Met hydrolase, destroying the methyl donor and causing T3 DNA to be
totally unmethylated.  These observations could stimulate analogous
investigations into the regulation of DNA methylation patterns of eukaryotic
viruses and cells.  For instance, an underrepresentation of methylation sites
(5'-CG) is also true for animal DNA viruses.  Moreover, we were able to
disclose some novel properties of DNA restriction-modification enzymes
concerning the protection of DNA recognition sequences in which only one strand
can be methylated (e.g., type III enzyme EcoP15) and the primary resistance of
(unmethylated) DNA recognition sites towards type II restriction endonuclease
EcoRII.

<>

<1>Kruger, H., Kirch, H.
<2>A nuclear protein from HeLa cells containing a methyltransferase specific domain binds to an intragenic region of the adenovirus 12 CS-1 E1A oncogene.
<3>Biol. Chem. Hoppe Seyler
<4>373
<5>789
<6>1992
<7>We identified a binding motif (CAGCTGC) for bHLH (basic region Helix Loop Helix) proteins in
the coding part of the Adenovirus 12 E1a gene. The growth enhanced but transformation
defective Vero-cell host range mutant CS-1 of Adenovirus 12 has a 69 bp deletion in E1a
adjacent to this binding site. We examined the intragenic site in the E1a-gene for binding of
HeLa and Vero nuclear proteins using gel shift assays. We obtained comparable patterns of 3-4
retarded bands for wild type and mutant E1a sequences using DNA fragments and mutant
oligonucleotides (Cdel). Oligonucleotides bearing motifs for the transcription factors AP-4
and/or AP-1 significantly affected the binding of nuclear proteins to the E1a probes, whereas
motifs for E2F, ATF, fac-b, and NF1 had little or no effect. SDS-PAGE analysis of Cdel binding
protein, purified by different methods, revealed average molecular weights of 70 kDa and 84
kDa. Constructs containing the intragenic DNA fragments of Ad12 wild type and CS-1 mutant E1a,
the SV40 promotor, and the CAT gene did not cause any enhanced or decreased CAT-activity after
transfection of HeLa cells compared to pCAT-promotor construct alone. Screening of a HeLa
lambda-gt11 expression library for Cdel-binding proteins resulted in five identical
cDNA-clones containing a 450 bp open reading frame. We identified a 25 amino acid long domain
that has 80% homology to the conserved region X of (cytidine-5)-DNA-methyltransferases. We
currently investigate the function of the purified protein.

<>

<1>Kruger, T., Grund, C., Wild, C., Noyer-Weidner, M.
<2>Characterization of the mcrBC region of Escherichia coli K-12 wild-type and mutant strains.
<3>Gene
<4>114
<5>1-12
<6>1992
<7>We have carried out an analysis of the Escherichia coli K-12 mcrBC locus in order to (1)
elucidate its genetic organization, (2) to identify the proteins encoded by this region, and
(3) to characterize their involvement in the restriction of DNA containing methylated cytosine
residues. In vitro expression of recombinant plasmids carrying all or portions of the mcrBC
region revealed that the mcrB and mcrC genes are organized as an operon. The mcrBC operon
specifies five proteins, as evident from parallel in vitro and in in vivo expression studies.
Three proteins of 53,35 and 34 kDa originate from mcrB expression, while two proteins of 37
and 16 kDa arise from mcrC expression. Products of both the mcrB and mcrC genes are required
to restrict the methylated substrate DNA used in this study. We also determined the nature of
mutant mcrBC loci in comparison to the E. coli K-12 wild-type mcrBC locus. A major goal of
these studies was to clarify the nature of the mcrB-1 mutation, which is carried by some
strains employed in previous analyses of the E. coli K-12 McrBC system. Based on our analyses
the mutant strains investigated could be divided into different complementation groups. The
mcrB-1 mutation is a nonsense or frameshift mutation located within mcrB. It causes premature
termination of mcrB gene product synthesis and reduces the level of mcrC gene expression. This
finding helps to understand an existing conflict in the literature. We also describe
temperature-sensitive McrA activity in some of the strains analysed and its relationship to
the previously defined differences in the tolerance levels of E. coli K-12 mcrBC mutants to
cytosine methylation.

<>

<1>Kruger, T., Wild, C., Noyer-Weidner, M.
<2>McrB: a prokaryotic protein specifically recognizing DNA containing modified cytosine residues.
<3>EMBO J.
<4>14
<5>2661-2669
<6>1995
<7>Restriction of DNA by the Escherichia coli K-12 McrBC restriction endonuclease, which consists
of the two subunits McrB and McrC, depends on the presence of modified cytosine residues in a
special constellation. From previous work by others it was known that restriction of
5-methylcytosine-containing DNA requires two methylated 5'-PuC sites separated by about 40-80
non-defined base pairs. Here we show that binding of the McrBC nuclease is mediated
exclusively by the McrB subunit. McrB has a low affinity for non-methylated DNA, with which it
forms low molecular weight complexes. The affinity for DNA is significantly increased, with
variations depending on the sequence context, by hemi- or fully methylated 5'-PuC sites.
Binding to such substrates yields high molecular weight complexes, presumably involving
several McrB molecules. Methylation at unique 5'-PuC sites can be sufficient to stimulate DNA
binding by McrB. As such substrates are not cleaved by the nuclease, restriction apparently
requires the coordinated interaction of molecules bound to neighboring 5'-PumC sites. The
binding properties of McrB exhibit some similarities to recently identified eukaryotic
proteins interacting in a non-sequence-specific manner with DNA containing methylated 5'-CpG
sequences and might point to a common molecular origin of these proteins. In addition to DNA,
McrB also binds GTP, an essential cofactor in DNA restriction by McrBC. McrC neither binds to
DNA nor modulates the DNA binding potential of McrB. As McrC is essential for restriction it
appears to predominantly function in catalysis.

<>

<1>Kruger, V.D.H., Reuter, M., Chernin, L.S., Gachechiladze, K.K., Chanishvili, T.G., Schroeder, C.
<2>Biological functions of restriction endonucleases.
<3>Biol. Zentralbl.
<4>109
<5>257-266
<6>1990
<7>The biological function of restriction endonucleases in bacterial cells is
reviewed (in german).  A description of the different classes of restriction
endonucleases is followed by a discussion of their evolutionary significance in
maintaining the compromise between genetic stabilility and variability of
microorganisms.  Restriction enzymes have a role in the defence against lethal
phage infection as well as in genetic recombination.  Two groups of enzymes in
Salmonella and Escherichia strains are presented as examples of restriction
enzyme evolution.  Certain bacterial cells express defence systems which attack
methylated DNA.  These enzymes complicate the cloning of methylated eukaryotic
DNA.

<>

<1>Krukov, V.M., Shlyapnikov, M.G., Kadyrov, F.A.
<2>Endonuclease SegE phage T4.  II. Determination and characterization of recognition site.
<3>Mol. Biol. (Mosk)
<4>30
<5>1307-1315
<6>1996
<7>
<>

<1>Kruse, T. et al.
<2>Complete genome sequence of Dehalobacter restrictus PER-K23(T.).
<3>Standards in Genomic Sciences
<4>8
<5>375-388
<6>2013
<7>Dehalobacter restrictus strain PER-K23 (DSM 9455) is the type strain of the species
Dehalobacter restrictus. D. restrictus strain PER-K23 grows by
organohalide respiration, coupling the oxidation of H2 to the reductive
dechlorination of tetra- or trichloroethene. Growth has not been observed with
any other electron donor or acceptor, nor has fermentative growth been shown.
Here we introduce the first full genome of a pure culture within the genus
Dehalobacter. The 2,943,336 bp long genome contains 2,826 protein coding and 82
RNA genes, including 5 16S rRNA genes. Interestingly, the genome contains 25
predicted reductive dehalogenase genes, the majority of which appear to be full
length. The reductive dehalogenase genes are mainly located in two clusters,
suggesting a much larger potential for organohalide respiration than previously
anticipated.

<>

<1>Krut, O., Palka-Santini, M., Cleven, B., Kronke, M.
<2>Analytical device for rapid identification of pathogens.
<3>International Patent Office
<4>WO 2007039319 A
<5>
<6>2007
<7>The present invention provides an analytical device, especially a DNA microarray, for
identification and characterization of microorganisms in a sample or clinical specimen.
Furthermore, it provides for a method for rapid identification and strain profiling of
different microbial species in a sample or clinical specimen, especially in a blood culture,
utilizing said analytical device.

<>

<1>Krut, O., Palka-Santini, M., Clevin, B., Kronke, M.
<2>DNA microarray for rapid identification of Candida albicans in blood cultures.
<3>European Patent Office
<4>EP 1770171 A
<5>
<6>2007
<7>The present invention provides a DNA microarray for identification and characterization of
microorganisms in a sample or clinical specimen.  Furthermore, it provides for a method for
rapid identification and strain profiling of different microbial species in clinical
specimens, especially in blood cultures, utilizing said DNA microarray.

<>

<1>Krylov, V.N., Karapetian, A.T.
<2>Discovery of new type of restriction and modification in enterobacteriaceae.
<3>Genetika
<4>13
<5>1079-1088
<6>1977
<7>The adsorption of 23 new lamboid bacteriophages to 547 strains which were isolated from a
natural population of Enterobacteriaceae was studied.  The frequency of positive combinations
of phage-bacterium with adsorption is not more than 2%.  A study of possible causes of limited
growth of lambdoid phages in the bacterial strains revealed that neither homoimmune prophage
nor prophage P2 are single factors of the growth limitation.  It is found that in natural
populations a selection of bacterial strains with the least limitation of phage takes place.
Three cases of killing bacteria after infection with high multiplicity are found.  The reason
of the killing effect is manifestation of some functions by infecting phages.  A new
restriction-modification system is found which differs from restriction-modification system A,
B, K, 15, P1, EcoRI, EcoRII.  Most strains, which adsorb phages but do not support their
growth, are supposed to possess several mechanisms of restriction.  Thus, the search for new
restriction systems in Escherichia coli is worthwhile.

<>

<1>Krynetskaya, N.F., Kubareva, E.A., Timchenko, M.A., Belkov, V.M., Shabarova, Z.A.
<2>Transition-metal complexes as inhibitors of proteins recognizing double-stranded fragments of nucleic acids.
<3>Biokhimiia
<4>63
<5>1251-1257
<6>1998
<7>A 30-membered DNA duplex containing the recognition site of restriction endonuclease SsoII and
a 30-membered RNA-DNA hybrid duplex, a substrate of E. coli RNase H, were synthesized.  The
cleavage of the 30-membered fragments of nucleic acids catalyzed by endonucleases in the
presence of Co-phthalocyanine complex [CoPc(COONa)8 containing eight carboxyl groups at the
periphery of the ligand was studied.  It was shown that the efficiency of enzyme catalysis
decreases in the presence of the metal complex for both endonucleases.  By addition of a
100-fold excess of Co-phthalocyanine complex with respect to DNA duplex the initial rate of
substrate hydrolysis by restriction endonuclease SsoII is observed to decrease twice.  An
equimolar ratio of the metal complex and hybrid duplex leads to essentially complete
inhibition of RNA cleavage by RNase H from E. coli.  The inhibition of catalytic activity of
enzymes recognizing the double-stranded nucleic acids in the presence of Co-phthalocyanine
complex is assumed to be caused by the ability of the latter to interact with DNA, RNA, and
DNA-RNA duplexes.

<>

<1>Krzyzanowska, D.M., Iwanicki, A., Ossowicki, A., Obuchowski, M., Jafra, S.
<2>Genome Sequence of Bacillus subtilis MB73/2, a Soil Isolate Inhibiting the Growth of Plant Pathogens Dickeya spp. and Rhizoctonia solani.
<3>Genome Announcements
<4>1
<5>e00238-13
<6>2013
<7>Bacillus subilis MB73/2 is a Gram-positive bacterium isolated in Poland from a meadow soil
sample. When tested in vitro, the strain shows strong antagonism
toward plant pathogens-the soft rot-causing bacteria Dickeya spp. and the crown
rot fungus Rhizoctonia solani. Here, we present the genome sequence of MB73/2.

<>

<1>Ku, C., Lo, W.S., Chen, L.L., Kuo, C.H.
<2>Complete Genome Sequence of Spiroplasma apis B31T (ATCC 33834), a Bacterium Associated with May Disease of Honeybees (Apis mellifera).
<3>Genome Announcements
<4>2
<5>e01151-13
<6>2014
<7>Spiroplasma apis B31(T) (ATCC 33834) is a wall-less bacterium in the class Mollicutes that has
been linked to May disease of honeybees (Apis mellifera).
Here, we report the complete genome sequence of this bacterium to facilitate the
investigation of its virulence factors.

<>

<1>Ku, C., Lo, W.S., Chen, L.L., Kuo, C.H.
<2>Complete genomes of two dipteran-associated spiroplasmas provided insights into the origin, dynamics, and impacts of viral invasion in Spiroplasma.
<3>Genome Biol. Evol.
<4>5
<5>1151-1164
<6>2013
<7>Spiroplasma is a genus of wall-less, low-GC, Gram-positive bacteria with helical morphology.
As commensals or pathogens of plants, insects, ticks, or crustaceans, they are closely related
with mycoplasmas and form a monophyletic group (Spiroplasma-
Entomoplasmataceae-Mycoides) with Mycoplasma mycoides and its relatives. In this study, we
report the complete genome sequences of S. chrysopicola and S. syrphidicola from the
Chrysopicola clade. These species form the sister group to the Citri clade, which includes
several well-known pathogenic spiroplasmas. Surprisingly, these two newly available genomes
from the Chrysopicola clade contain no plectroviral genes, which were found to be
highly repetitive in the previously sequenced genomes from the Citri clade. Based on the
genome alignment and patterns of GC-skew, these two Chrysopicola genomes appear to be
relatively stable, rather than being highly rearranged as those from the Citri clade.
Phylogenetic analyses suggest that the susceptibility to plectroviral invasion probably
originated in the common ancestor of the Citri clade or one of its subclades. This
susceptibility may be attributed to the absence of antiviral systems found in the Chrysopicola
clade. Using the virus-free genomes of the Chrysopicola clade as references, we inferred the
putative viral integration sites in the Citri genomes. Comparisons of syntenic regions suggest
that the extensive viral invasion in the Citri clade promoted genome rearrangements and
expansions. More importantly, the viral invasion may have facilitated horizontal gene
transfers that contributed to adaptation in the Citri clade.

<>

<1>Kubareva, E.A., Gromova, E.S., Ortskaya, T.S., Shabarova, Z.A.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments.  X. Hydrolysis of substrates with structural anomalies.
<3>Bioorg. Khim.
<4>13
<5>1205-1211
<6>1987
<7>Interaction of the EcoRII restriction endonuclease with a set of 30-membered
substrates having structural anomalies in the recognition site (^CCT/AGG) and
in adjacent sequences has been studied.  A nick in the centre of the EcoRII
recognition site between dC and dA residues slows down hydrolysis of the
nonmodified strand, whereas the modified one is not cleaved.  Removal of the
phosphate group from the nick in this substrate does not alter the rate of the
cleavage.  The absence of one of the phosphate groups in the flanking sequence
at a twobase-pair distance from the recognition site slows down the enzymatic
hydrolysis.  Removal of dA or dT out of the EcoRII recognition site blocks the
enzymatic reaction.  It appears that EcoRII does not interact with the
phosphate group between dC and dA residues in the recognition site.
Suggestions are made concerning possible contacts of the EcoRII restriction
endonuclease with dA- and dT-residues of the recognition site and with the
sugar-phosphate backbone of the adjacent nucleotide sequences.

<>

<1>Kubareva, E.A., Gromova, E.S., Pein, C.-D., Krug, A., Oretskaya, T.S., Cech, D., Shabarova, Z.A.
<2>Oligonucleotide cleavage by restriction endonucleases MvaI and EcoRII: a comprehensive study on the influence of structural parameters on the enzyme-substrate interaction.
<3>Biochim. Biophys. Acta
<4>1088
<5>395-400
<6>1991
<7>To elucidate the mechanism of action of restriction endonucleases -
isoschizomers EcoRII and MvaI - a study was made of their interaction with a
set of synthetic oligonucleotide duplexes containing a single 5'-d(CCA/TGG)-3'
EcoRII (MvaI) recognition site.  The substrates had varying length and
structure of the nucleotide sequences flanking the recognition site.  The
structure of the flanking sequence is important for the cleavage by EcoRII and
MvaI enzymes; there is a structure which was found to speed up the EcoRII and
MvaI action.  The cleavage of oligonucleotide duplexes by EcoRII enzyme does
not go to completion.  EcoRII endonuclease cleaved extended substrates less
efficiently than short ones.  Extension of the flanking sequences, with the
same nucleotide surrounding of the recognition site, substantially altered the
whole kinetic pattern of MvaI hydrolysis.  This was not observed with EcoRII
enzyme.  The restriction endonuclease MvaI distinguished between dA and dT
residues in the recognition site, which was reflected in the higher rate of
hydrolysis of the dA-containing strand of the quasipalindromic DNA duplex.

<>

<1>Kubareva, E.A., Gromova, E.S., Romanova, E.A., Oretskaya, T.S., Shabarova, Z.A.
<2>Cleavage of substrates containing modified amino groups in heterocyclic bases by MvaI and EcoRII restriction endonucleases.
<3>Bioorg. Khim.
<4>16
<5>501-506
<6>1990
<7>14-membered DNA-duplexes containing modified nucleoside residues, viz 4-N-methyldeoxycytidine
(m4dC), 6-N-methyldeoxyadenosine (m6dA) or deoxyinosine (dI), in only one strand of the
recognition site (CCA/TGG) of MvaI and EcoRII endonucleases were synthesized.  It was shown
that MvaI and EcoRII endonucleases interact with the exocyclic amino groups of the external dC
residues and of the central dA residue of the recognition site exposed into the DNA major
groove.  These endonucleases which are isoschizomers were found to possess different
mechanisms of substrate cleavage.  The ability of MvaI endonuclease to hydrolyze only the
unmodified strand of methylated duplexes allows one to make site-directed single-strand nicks
in double-stranded DNA. Elimination of the 2-NH2-group located in the minor groove of DNA by
substituting dI for dG had little, if any, effect on the hydrolytic activity of EcoRII and
MvaI endonucleases.

<>

<1>Kubareva, E.A., Kuznetsova, S.A., Kanevsky, I.A., Karyagina, A.S., Nikolskaya, I.I., Shabarova, Z.A.
<2>DNA dumbbells as substrates for studying restriction-modification and repair enzymes.
<3>FASEB J.
<4>11
<5>A1309
<6>1997
<7>The use of the DNA dumbbells (duplexes covalently closed on both ends by single stranded
loops) for investigation of restriction-modification and repair enzymes has been demonstrated.
Their substrate properties in comparison with DNA hairpins have been analysed in the reactions
with type II restriction endonucleases (R): MvaI, EcoRII, SsoII and
DNA-(5-methylcytosine)methyltransferase (M): SsoII.  The hairpin and dumbbell, containing dU
instead of dT in the center of the restriction-modification enzyme's recognition site
(5'CCWGG3'), have been used to evaluate the uracil removal from ds DNA with high duplex
stability by uracil-DNA glycosylase.  R.EcoRII, R.MvaI and R.SsoII interact more efficiently
with more stable hairpin-like substrate than with equivalent linear DNA duplex.  Less
efficiency is observed for dumbbell cleavage by restriction endonucleases as compared with
hairpin-like duplex.  The decreased conformational mobility of the strands in the dumbbell
might affect adversely on the substrate hydrolysis by the restriction endonucleases.  Addition
of the 14-membered DNA substrate accelerates the cleavage of the dumbbell by R.EcoRII.
M.SsoII and UDG reveal 1.5-2.5-fold preference for dsDNA duplex over hairpin-like DNA, which
is 30 C more stable.  The most stable dumbbell is also a poor substrate for M.SsoII and
especially for UDG.  The M.SsoII or UDG functioning is more effective when the double helix is
less stable.

<>

<1>Kubareva, E.A., Pein, C.-D., Gromova, E.S., Kuznezova, S.A., Tashlitzki, V.N., Chech, D., Shabarova, Z.A.
<2>The role of modifications in oligonucleotides in sequence recognition by MvaI restriction endonuclease.
<3>Eur. J. Biochem.
<4>175
<5>615-618
<6>1988
<7>The interaction of MvaI restriction endonuclease with 14-membered
deoxyribonucleotide duplexes containing modifications within the recognition
site (CC[A/T]GG) has been studied.  Substitution of m5dC for the internal dC
residue, as well as substitution of fl5dU or rU for dT did not influence the
initial rate of hydrolysis (Vo) of modified strands, whereas the hydrolysis of
unmodified strands was inhibited in some cases.  Furthermore, the substitution
of a pyrophosphate bond for a scissile phosphodiester bond in one strand
completely inhibited digestion in this strand without any decrease of the rate
of hydrolysis of the unmodified strand.  In contrast to EcoRI endonuclease,
which recognizes the same DNA sequence, in the case of MvaI endonuclease
substrate recognition is possible in a wide range of conformational, electronic
and hydrophobic alterations within the recognition site.

<>

<1>Kubareva, E.A., Petrauskene, O.V., Karyagina, A.S., Nikolskaya, I.I., Gromova, E.S.
<2>Change of cleavage site of synthetic substrates by SsoII restriction endonuclease due to non-nucleotide inserts into the recognition sequence.
<3>Bioorg. Khim.
<4>18
<5>1131-1134
<6>1992
<7>The cleavage of synthetic DNA duplexes containing 1,3-propanediol, 1,2-dideoxy-D-ribofuranose
or 9-[1'-hydroxy-2'-(hydroxymethyl)ethoxy]methylguanine (glG) residues instead of one of the
dG residues or one of the nucleosides of the central base pair of the recognition site by
SsoII restriction endonuclease (CCNGG) has been studied. It is found that the non-nucleotide
insertions (except for glG) result in a change of the SsoII cleavage site and an increase of
the efficiency of the cleavage. The novel noncanonical cleavage occurs at the phosphodiester
bond adjoining the non-nucleotide insert from the 5'-end.

<>

<1>Kubareva, E.A., Petrauskene, O.V., Karyagina, A.S., Nikolskaya, I.I., Gromova, E.S.
<2>DNA duplexes with non-nucleotide inserts: interaction with restriction endonucleases.
<3>Nucleic Acids Symp. Ser.
<4>24
<5>308
<6>1991
<7>Modified DNA duplexes with trimethylene bridges or 1,2-dideoxy-D-ribofuranose instead of one
of the nucleoside residues are of interest as DNA analogs with the sugar-phosphate backbone
partially or completely preserved but with the heterocyclic base removed.  Another type of
non-nucleotide insert - 9[1'-hydroxy-2'- (hydroxymethyl)ethoxy]methylguanine (acyclic dG) -
leads to distortion of the sugar moiety of nucleoside residue while the base is preserved.
14-membered DNA duplexes have been constructed containing single non-nucleotide inserts in the
recognition sites (5'-CC(A/T)GG-3' and 5'-CCNGG-3') of a number of restriction
endonucleases (R).dG residues or one of the nucleosides of the central base-pair of the
recognition site have been modified.  It has been shown by UV spectroscopy and CD that
non-nucleotide inserts bring about some destabilization of the double helix without essential
distortion of its geometry.  Substrate properties of DNA duplexes with non-nucleotide inserts
have been studied.  R. ScrFI and R.SsoII recognize the CCNGG sequence.  R.ScrFI cleaves all
the modified duplexes, but with reduced efficiency.  Introduction of trimethylene bridge or
1,2-dideoxy-D-ribofuranose into the recognition site induces an increase in the cleavage
efficiency of modified substrates and change of specificity of their hydrolysis by R.SsoII.
Restoration of R.SsoII specificity is observed in the case of acyclic dG-containing substrate
when guanine base is returned.  To preserve canonical R.SsoII specificity the enzyme should be
able to form discrimination contacts with internal and external dG and central nucleoside
residues of the recognition site.  DNA duplexes with non-nucleotide inserts in the CC(A/T)GG
recognition sequence (with the exception of acyclic dG insert) are resistant to R.MvaI,
R.BstNI and R.EcoRII.  Probably, these enzymes interact specifically with each base of the
central A.T pair and with both dG residues.  A comparative analysis of the functioning of
restriction enodnucleases - isoschizomers - has been done on the basis of these data.

<>

<1>Kubareva, E.A., Petrauskiene, O.V., Karyagina, A.S., Tashlitsky, V.N., Nikolskaya, I.I., Gromova, E.S.
<2>Cleavage of synthetic substrates containing non-nucleotide inserts by restriction endonucleases. Change in the cleavage specificity of endonuclease SsoII.
<3>Nucleic Acids Res.
<4>20
<5>4533-4538
<6>1992
<7>A study was made of the interaction between restriction endonucleases recognizing CCNGG (SsoII
and ScrFI) or CCA/TGG (MvaI and EcoRII) DNA sequences and a set of synthetic substrates
containing 1,3-propanediol,1,2-dideoxy-D-ribofuranose or
9-[1'-hydroxy-2'-(hydroxymethyl)ethoxy] methylguanine (glG) residues replacing either one of
the central nucleosides or dG residues in the recognition site. The non-nucleotide inserts
(except for glG) introduced into the recognition site both increase the efficiency of SsoII
and change its specificity. A cleavage at the noncanonical position takes place, in some cases
in addition to the correct ones. Noncanonical hydrolysis by SsoII occurs at the phosphodiester
bond adjacent to the point of modification towards the 5'-end. With the guanine base returned
(the substrate with glG), the correct cleavage position is restored. ScrFI specifically
cleaves all the modified substrates. DNA duplexes with non-nucleotide inserts (except for the
glG-containing duplex) are resistant to hydrolysis by MvaI and EcoRII. Prompted by the data
obtained we discuss the peculiarities of recognition by restriction endonucleases of
5-membered DNA sequences which have completely or partially degenerate central base pairs. It
is sugegested that SsoII forms a complex with DNA in an "open" form.

<>

<1>Kubareva, E.A., Thole, H., Karyagina, A.S., Oretskaya, T.S., Pingoud, A., Pingoud, V.
<2>Identification of a base-specific contact between the restriction endonuclease SsoII and its recognition sequence by photocross-linking.
<3>Nucleic Acids Res.
<4>28
<5>1085-1091
<6>2000
<7>A target sequence-specific DNA binding region of the restriction endonuclease Sso II was
identified by photocross-linking with an oligodeoxynucleotide duplex which was substituted
with 5-iododeoxyuridine (5-IdU) at the central position of the Sso II recognition site
(CCNGG). For this purpose the Sso II-DNA complex was irradiated with a helium/cadmium laser
(325 nm). The cross-linking yield obtained was approximately 50%. In the presence of excess
unmodified oligodeoxynucleotide or with oligodeoxynucleotides substituted with 5-IdU
elsewhere, no cross-linking was observed, indicating the specificity of the cross-linking
reaction. The cross-linked Sso II-oligodeoxynucleotide complex was digested with chymotrypsin,
a cross-linked peptide- oligodeoxynucleotide complex isolated and the site of cross-linking
identified by Edman sequencing to be Trp61. In line with this identification is the finding
that the W61A variant cannot be cross-linked with the IdU-substituted oligodeoxynucleotide,
shows a decrease in affinity towards DNA and is inactive in cleavage. It is concluded that the
region around Trp61 is involved in specific binding of Sso II to its DNA substrate.

<>

<1>Kubareva, E.A., Walter, J., Karyagina, A.S., Vorobeva, O.V., Lau, P.C., Trautner, T.
<2>Determination of methylation site of DNA-methyltransferase NlaX by a hybrid method.
<3>Biotechniques
<4>33
<5>526-531
<6>2002
<7>Using a new method based on a combination of bisulfite reaction, the repair enzyme uracil-DNA
glycosylase, and synthetic oligodeoxyribonucleotides, the methylation site of
DNA-methyltransferase NlaX (M.NlaX) from Neisseria lactamica was established to be the inner
cytosine in the double-stranded pentanucleotide recognition sequence 5'-CCNGG-3' (where N =
any nucleoside). 5-Methylcytosine (m5C) type modification by M-N1aX was confirmed by the use
of oligonucleotide substrates that contain 5-fluoro-2'-deoxycytidine.

<>

<1>Kubareva, E.A., Walter, J., Vorobeva, O.V., Razumikhin, M.V., Karyagina, A.S., Lau, P.C.K., Trautner, T.
<2>Determination of a non-methylated deoxycytidine desidue in the decognition site of DNA-methyltransferases.
<3>Biokhimiia
<4>66
<5>1356-1360
<6>2001
<7>A method for determination of a non-methylated deoxycytidine (dC) residue in the recognition
site of 5-cytosine DNA-methyltransferases is suggested. The method is based on treatment of
methylated DNA by sodium bisulfite and successive reaction of the thus modified DNA with a
repair enzyme, uracil-DNA glycosylase. This method was successfully applied to identify NlaX
methyltransferase specificity.

<>

<1>Kube, M., Beck, A., Zinder, S.H., Kuhl, H., Reinhardt, R., Adrian, L.
<2>Genome sequence of the chlorinated compound-respiring bacterium Dehalococcoides species strain CBDB1.
<3>Nat. Biotechnol.
<4>23
<5>1269-1273
<6>2005
<7>Dehalococcoides species are strictly anaerobic bacteria, which catabolize many of the most
toxic and persistent chlorinated aromatics and aliphatics
by reductive dechlorination and are used for in situ bioremediation of
contaminated sites. Our sequencing of the complete 1,395,502 base pair
genome of Dehalococcoides strain CBDB1 has revealed the presence of 32
reductive-dehalogenase-homologous (rdh) genes, possibly conferring on the
bacteria an immense dehalogenating potential. Most rdh genes were
associated with genes encoding transcription regulators such as
two-component regulatory systems or transcription regulators of the
MarR-type. Four new paralog groups of rdh-associated genes without known
function were detected. Comparison with the recently sequenced genome of
Dehalococcoides ethenogenes strain 195 reveals a high degree of gene
context conservation (synteny) but exceptionally high plasticity in all
regions containing rdh genes, suggesting that these regions are under
intense evolutionary pressure.

<>

<1>Kube, M., Migdoll, A.M., Gehring, I., Heitmann, K., Mayer, Y., Kuhl, H., Knaust, F., Geider, K., Reinhardt, R.
<2>Genome comparison of the epiphytic bacteria Erwinia billingiae and E. tasmaniensis with the pear pathogen E. pyrifoliae.
<3>BMC Genomics
<4>11
<5>393
<6>2010
<7>ABSTRACT: BACKGROUND: The genus Erwinia includes plant-associated
pathogenic and non-pathogenic Enterobacteria. Important pathogens such as
Erwinia amylovora, the causative agent of fire blight and E. pyrifoliae
causing bacterial shoot blight of pear in Asia belong to this genus. The
species E. tasmaniensis and E. billingiae are epiphytic bacteria and may
represent antagonists for biocontrol of fire blight. The presence of genes
putative involved in virulence in E. amylovora and E. pyrifoliae is of
special interest for these species in consequence. RESULTS: Here we
provide the complete genome sequences of the pathogenic E. pyrifoliae
strain Ep1/96 with a size of 4.1 Mb and of the non-pathogenic species E.
billingiae strain Eb661 with a size of 5.4 Mb, de novo determined by
conventional Sanger sequencing and next generation sequencing techniques.
Genome comparison reveals large inversions resulting from homologous
recombination events. Furthermore, comparison of deduced proteins
highlights a relation of E. billingiae strain Eb661 to E. tasmaniensis
strain Et1/99 and a distance to E. pyrifoliae for the overall gene content
as well as for the presence of encoded proteins representing virulence
factors for the pathogenic species. Pathogenicity of E. pyrifoliae is
supposed to have evolved by accumulation of potential virulence factors.
E. pyrifoliae carries factors for type III secretion and cell invasion.
Other genes described as virulence factors for E. amylovora are involved
in the production of exopolysaccharides, the utilization of plant
metabolites such as sorbitol and sucrose. Some virulence-associated genes
of the pathogenic species are present in E. tasmaniensis but mostly absent
in E. billingiae. CONCLUSION: The data of the genome analyses correspond
to the pathogenic lifestyle of E. pyrifoliae and underlines the epiphytic
localization of E. tasmaniensis and E. billingiae as a saprophyte.

<>

<1>Kube, M., Schneider, B., Kuhl, H., Dandekar, T., Heitmann, K., Migdoll, A.M., Reinhardt, R., Seemuller, E.
<2>The linear chromosome of the plant-pathogenic mycoplasma 'Candidatus Phytoplasma mali'.
<3>BMC Genomics
<4>9
<5>306
<6>2008
<7>BACKGROUND: Phytoplasmas are insect-transmitted, uncultivable bacterial plant pathogens that
cause diseases in hundreds of economically important
plants. They represent a monophyletic group within the class Mollicutes
(trivial name mycoplasmas) and are characterized by a small genome with a
low GC content, and the lack of a firm cell wall. All mycoplasmas,
including strains of 'Candidatus (Ca.) Phytoplasma asteris' and 'Ca. P.
australiense', examined so far have circular chromosomes, as is the case
for almost all walled bacteria. RESULTS: Our work has shown that 'Ca.
Phytoplasma mali', the causative agent of apple proliferation disease, has
a linear chromosome. Linear chromosomes were also identified in the
closely related provisional species 'Ca. P. pyri' and 'Ca. P. prunorum'.
The chromosome of 'Ca. P. mali' strain AT is 601,943 bp in size and has a
GC content of 21.4%. The chromosome is further characterized by large
terminal inverted repeats and covalently closed hairpin ends. Analysis of
the protein-coding genes revealed that glycolysis, the major
energy-yielding pathway supposed for 'Ca. P. asteris', is incomplete in
'Ca. P. mali'. Due to the apparent lack of other metabolic pathways
present in mycoplasmas, it is proposed that maltose and malate are
utilized as carbon and energy sources. However, complete ATP-yielding
pathways were not identified. 'Ca. P. mali' also differs from 'Ca. P.
asteris' by a smaller genome, a lower GC content, a lower number of
paralogous genes, fewer insertions of potential mobile DNA elements, and a
strongly reduced number of ABC transporters for amino acids. In contrast,
'Ca. P. mali' has an extended set of genes for homologous recombination,
excision repair and SOS response than 'Ca. P. asteris'. CONCLUSION: The
small linear chromosome with large terminal inverted repeats and
covalently closed hairpin ends, the extremely low GC content and the
limited metabolic capabilities reflect unique features of 'Ca. P. mali',
not only within phytoplasmas, but all mycoplasmas. It is expected that the
genome information obtained here will contribute to a better understanding
of the reduced metabolism of phytoplasmas, their fastidious nutrition
requirements that prevented axenic cultivation, and the mechanisms
involved in pathogenicity.

<>

<1>Kube, M., Siewert, C., Migdoll, A.M., Duduk, B., Holz, S., Rabus, R., Seemuller, E., Mitrovic, J., Muller, I., Buttner, C., Reinhardt, R.
<2>Analysis of the Complete Genomes of Acholeplasma brassicae, A. palmae and A. laidlawii and Their Comparison to the Obligate Parasites from ' Candidatus Phytoplasma'.
<3>J. Mol. Microbiol. Biotechnol.
<4>24
<5>19-36
<6>2013
<7>Analysis of the completely determined genomes of the plant-derived Acholeplasma
brassicae strain O502 and A. palmae strain J233 revealed that the circular
chromosomes are 1,877,792 and 1,554,229 bp in size, have a G + C content of 36
and 29%, and encode 1,690 and 1,439 proteins, respectively. Comparative analysis
of these sequences and previously published genomes of A. laidlawii strain PG-8,
'Candidatus Phytoplasma asteris' strains, 'Ca. P. australiense' and 'Ca. P. mali'
show a limited shared basic genetic repertoire. The acholeplasma genomes are
characterized by a low number of rearrangements, duplication and integration
events. Exceptions are the unusual duplication of rRNA operons in A. brassicae
and an independently introduced second gene for a single-stranded binding protein
in both genera. In contrast to phytoplasmas, the acholeplasma genomes differ by
encoding the cell division protein FtsZ, a wide variety of ABC transporters, the
F0F1 ATP synthase, the Rnf-complex, SecG of the Sec-dependent secretion system, a
richly equipped repertoire for carbohydrate metabolism, fatty acid, isoprenoid
and partial amino acid metabolism. Conserved metabolic proteins encoded in
phytoplasma genomes such as the malate dehydrogenase SfcA, several transporters
and proteins involved in host-interaction, and virulence-associated effectors
were not predicted for the acholeplasmas. (c) 2013 S. Karger AG, Basel.

<>

<1>Kubicek-Sutherland, J.Z., Heithoff, D.M., Ersoy, S.C., Shimp, W.R., Mahan, M.J.
<2>Immunization with a DNA adenine methylase over-producing Yersinia pseudotuberculosis vaccine confers robust cross-protection against heterologous pathogenic serotypes.
<3>Vaccine
<4>32
<5>1451-1459
<6>2014
<7>Yersinia pseudotuberculosis is a foodborne pathogen that can cause serious human illness.
Although the source and route of transmission often remain obscure, livestock have been
implicated in some cases. The diversity of yersiniae present on farms and their widespread
distribution in animal and environmental reservoirs necessitates the use of broad prophylactic
strategies that are efficacious against many serotypes simultaneously. Herein, immunization of
mice with a modified, live attenuated Y. pseudotuberculosis vaccine that overproduces the DNA
adenine methylase (Dam P) conferred robust protection against virulent challenge (150-fold
LD50) with homologous and heterologous serotypes that have been associated with human disease
(O:1, O:1a, O:3). Further, the dam gene was shown to be essential for cell viability in all (7
of 7) Y. pseudotuberculosis strains tested. Direct selection for the inheritance of dam mutant
alleles in Y. pseudotuberculosis resulted in dam strain variants that contained compensatory
(second-site suppressor) mutations in genes encoding methyl-directed mismatch repair proteins
(mutHLS) that are involved in suppression of the non-viable cell phenotype in all (19/19)
strains tested. Such dam mutH variants exhibited a significant increase in virulence and
spontaneous mutation frequency relative to that of a Dam P vaccine strain. These studies
indicate that Y. pseudotuberculosis Dam P strains conferred potent cross-protective efficacy
as well as decreased virulence and spontaneous mutation frequency relative to those that lack
Dam, which have compensatory mutations in mutHLS loci. These data suggest that development of
yersiniae livestock vaccines based on Dam overproduction is a viable mitigation strategy to
reduce these potential foodborne contaminants. (C) 2014 Elsevier Ltd. All rights reserved.

<>

<1>Kubota, T.
<2>DNA methylase 3B (DNMT3B).
<3>Seitai Kagaku
<4>56
<5>378-379
<6>2005
<7>
<>

<1>Kucera, R.B., Dalton, M.A., Higgins, L.S., Schildkraut, I., Kong, H., Wilson, G.G.
<2>Cloning And Producing The N.BstNBI Nicking Endonuclease And Related Methods For Using Nicking Endonucleases In Single-Stranded Displacement Amplification.
<3>Japanese Patent Office
<4>JP 2003535584 A
<5>
<6>2003
<7>
<>

<1>Kuch, D., Schermelleh, L., Manetto, S., Leonhardt, H., Carell, T.
<2>Synthesis of DNA dumbbell based inhibitors for the human DNA methyltransferase Dnmt1.
<3>Angew. Chem. Int. Ed. Engl.
<4>47
<5>1515-1518
<6>2008
<7>DNA methyltransferases convert deoxycytidine nucleobases in DNA into 5-methyldeoxycytidines
using the cofactor S-adenosylmethionine as the methyl group donor.  Methylation of the
canonical dC base, particularly in gene promoter regions, induces complex processes, which
finally lead to the silencing of the corresponding gene.  This epigenetic gene silencing is of
paramount importance for cellular differentiation.  Altered methylation patterns and
corresponding changes in gene expression are found in practically all tumor cells.  The major
DNA methyltransferase Dnmt1 is a 183-kDa-large protein that preferentially methylates dC bases
in hemimethylated d(cpG) sequences after DNA replication.

<>

<1>Kuck, D., Singh, N., Lyko, F., Medina-Franco, J.L.
<2>Novel and selective DNA methyltransferase inhibitors: Docking-based virtual screening and experimental evaluation.
<3>Bioorg. Med. Chem.
<4>18
<5>822-829
<6>2010
<7>The DNA methyltransferase (DNMT) enzyme family consists of four members with diverse functions
and represents one of the most promising targets
for the development of novel anticancer drugs. However, the standard
drugs for DNMT inhibition are non-selective cytosine analogues with
considerable cytotoxic side-effects that have been developed several
decades ago. In this work, we conducted a virtual screening of more
than 65,000 lead-like compounds selected from the National Cancer
Institute collection using a multistep docking approach with a
previously validated homology model of the catalytic domain of human
DNMT1. Experimental evaluation of top-ranked molecules led to the
discovery of novel small molecule DNMT1 inhibitors. Virtual screening
hits were further evaluated for DNMT3B inhibition revealing several
compounds with selectivity towards DNMT1. These are the first small
molecules reported with biochemical selectivity towards an individual
DNMT enzyme capable of binding in the same pocket as the native
substrate cytosine, and are promising candidates for further rational
optimization and development as anticancer drugs. The availability of
enzyme-selective inhibitors will also be of great significance for
understanding the role of individual DNMT enzymes in epigenetic
regulation.

<>

<1>Kudirkiene, E., Christensen, H., Bojesen, A.M.
<2>Draft Genome Sequence of Gallibacterium anatis bv. haemolytica 12656-12 Liver, an Isolate Obtained from the Liver of a Septicemic Chicken.
<3>Genome Announcements
<4>1
<5>e00810-13
<6>2013
<7>We report the draft genome sequence of Gallibacterium anatis bv. haemolytica strain 12656-12
Liver. This strain was isolated from the liver of a septicemic
layer chicken in Denmark in 1981. The strain has been used extensively for
experimental purposes.

<>

<1>Kudirkiene, E., Hansen, M.J., Bojesen, A.M.
<2>Draft Genome Sequence of Chelonobacter oris Strain 1662T, Associated with Respiratory Disease in Hermann's Tortoises.
<3>Genome Announcements
<4>2
<5>e01322-14
<6>2014
<7>Chelonobacter oris 1662(T) is a type strain of the recently described species of  the
Pasteurellaceae family. The strain was isolated from the choanae of a captive
tortoise with signs of respiratory tract infection. The genome reported here is
approximately 2.6 Mb in size and has a G+C content of 47.1%.

<>

<1>Kudo, T. et al.
<2>Draft Genome Sequences of Psychrobacter Strains JCM 18900, JCM 18901, JCM 18902,  and JCM 18903, Isolated Preferentially from Frozen Aquatic Organisms.
<3>Genome Announcements
<4>2
<5>e00280-14
<6>2014
<7>Four Psychrobacter strains, JCM 18900, JCM 18901, JCM 18902, and JCM 18903, related to either
Psychrobacter nivimaris or Psychrobacter cibarius, were isolated from frozen marine animals.
The genome information of these four strains will be useful for studies of their physiology
and adaptation properties to frozen conditions.

<>

<1>Kudo, T., Kawauchi, A., Nakahara, T., Zhang, X., Taniyama, S., Takatani, T., Arakawa, O., Oshima, K., Suda, W., Kitamura, K., Iida, T., Iino, T., Inoue, T., Hongoh, Y., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequences of Vibrio sp. Strains Isolated from Tetrodotoxin-Bearing Scavenging Gastropod.
<3>Genome Announcements
<4>2
<5>e00623-14
<6>2014
<7>Vibrio sp. strains JCM 18905 and JCM 19053 were isolated from a tetrodotoxin (TTX)-bearing
scavenging gastropod, and Vibrio sp. strain JCM 18904 was isolated
from a sea cucumber. All these are closely related to Vibrio alginolyticus. Their
comparative genome information is useful for studies of TTX production in
bacteria.

<>

<1>Kudo, T., Nakahara, T., Zhang, X., Taniyama, S., Arakawa, O., Murase, S., Nakata, H., Oshima, K., Suda, W., Kitamura, K., Iida, T., Oshida, Y., Inoue, T., Hongoh, Y., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequences of Geomicrobium sp. Strains JCM 19037, JCM 19038, JCM 19039, and JCM 19055, Isolated from Aquatic Samples.
<3>Genome Announcements
<4>2
<5>e00622-14
<6>2014
<7>Haloalkaliphilic strains JCM 19037, JCM 19038, JCM 19039, and JCM 19055, closely  related to
Geomicrobium sediminis, were isolated from aquatic samples, and their
draft genome sequences were determined. The genome information of these four
strains will be useful for studies of their physiology and ecology.

<>

<1>Kudo, T., Sakamoto, K., Akinaga, M., Kawauchi, A., Nakahara, T., Zhang, X., Yamada, A., Oshima, K., Suda, W., Kuwahara, H., Nakamura, N., Nogi, Y., Kitamura, K., Yuki, M., Iida, T., Moriya, S., Inoue, T., Hongoh, Y., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequences of Cyclodextrin-Producing Alkaliphilic Bacillus Strains JCM 19045, JCM 19046, and JCM 19047.
<3>Genome Announcements
<4>2
<5>e00211-14
<6>2014
<7>Bacillus strains JCM 19045, JCM 19046, and JCM 19047 are alkaliphiles that produce
beta-cyclodextrin from starch. They are related to Bacillus xiaoxiensis
and Bacillus lehensis. The genome information for these three strains will be
useful for studies of the physiological role of cyclodextrin and cyclodextrin
production.

<>

<1>Kuebbing, D., Blakesley, R.J.
<2>A New Restriction Endonuclease from Streptomyces Stanford.
<3>Fed. Proc.
<4>38
<5>780
<6>1979
<7>A new site-specific endonuclease activity distinguishable from Sst I, II and
III was isolated from Streptomyces stanford. This enzyme, designated Sst IV,
cleaved SV40 DNA once, adeno-virus type-2 DNA four times, and lambda(wt) DNA
several times (incompletely), but did not cleave PhiX174 RF or pBR322 DNA.  Sst
IV was active in 15 mM Tris-HCL (pH 7.5, mM MgCl2, 90 mM NaCl and 6mM
2-mercaptoethanol at 37C.  The enzyme was purified free of specific and
non-specific nucleases by column chromatography which included
phosphocellulose, DEAE cellulose, hydroxylapatite and Blue-CNBr agarose.  The
amount of Sst IV recovered was about 2% of that for Sst II.  The number and
sizes of fragments actually generated by co-digestion of pBR322, PhiX174 RF or
SV40 DNA with Sst IV and other known restriction endonucleases were compared
with a computer generated table of predicted fragmentation patterns of these
DNAs (R. Blakesley, BRL FOCUS 1, NO. 4 (1978)) To tentatively identify the
recognition sequence of SstIV as 5'-TGATCA-3'.  Thus, SstIV has a specificity
similar to that of AtuCI, BclI and CpeI.  The recognition sequence is being
confirmed by direct sequence analysis.

<>

<1>Kuehn, J.S., Register, K.B., Phillips, G.J.
<2>Draft Genome Sequences for 10 Isolates of the Swine Pathogen Haemophilus parasuis.
<3>Genome Announcements
<4>1
<5>e00739-13
<6>2013
<7>Haemophilus parasuis colonizes the upper respiratory tract of swine and can cause a severe
systemic disease known as Glasser's disease. We report here the draft
genome sequences of 10 isolates from geographically diverse locations
representing the full virulence spectrum of the microorganism, which will aid in
understanding the pathobiology of H. parasuis.

<>

<1>Kuehn, R., Wurst, W., Meyer, M.
<2>Fusion proteins comprising a DNA-binding domain of a Tal effector protein and a non-specific cleavage domain of a restriction nuclease and their use.
<3>European Patent Office
<4>EP 2392208 A
<5>
<6>2011
<7>The present invention relates to a method of modifying a target sequence in the genome of a
eukaryotic cell, the method comprising the step: (a) introducing into the cell a fusion
protein comprising a DNA-binding domain of a Tal effector protein and a non-specific cleavage
domain of a restriction nuclease or a nucleic acid molecule encoding the fusion protein in
expressible form, wherein the fusion protein specifically binds within the target sequence and
introduces a double strand break within the target sequence.  The present invention further
relates to the method of the invention, wherein the modification of the target sequence is by
homologous recombination with a donor nucleic acid sequence further comprising the step: (b)
introducing a nucleic acid molecule into the cell, wherein the nucleic acid molecule comprises
the donor nucleic acid sequence and regions homologous to the target sequence.  The present
invention also relates to a method of producing a non-human mammal or vertebrate carrying a
modified target sequence in its genome.  Furhtermore, the present invention rlates to a fusion
protein comprising a DNA-binding domain of a Tal effector protein and a non-specific cleavage
domain of a restriction nuclease.

<>

<1>Kuehne, C., Simoncsits, A.
<2>Chimeric polypeptides and their use.
<3>International Patent Office
<4>WO 2004092194 A
<5>
<6>2004
<7>The present invention concerns chimeric molecules that contain a preferential polypeptide
region, consisting of a specific affinity for the binding to specific DNA sequences, of a
preferential polypeptidic region consisting of a DNA modifying activity, and this chimeric
molecule is capable to cross biological membranes due to the presence of a region that contins
delivery activity.  The invention contains further the isolated polynucleotides that code for
the chimeric molecules of the invention if they are as such entirely or partially of
polypeptide nature.  In another embodiment, based on the activities of the polypeptides
contained in the invention to interfere with key points of the cell-cycle regulation and the
cellular checkpoints due to their introduction of DNA double strand breaks, the invention
contains various procedures that are characterized by the use of said polypeptides of the
invention for cells in vivo and provides an activity for the modification of specific sites of
the DNA contained in a cell.  The invention also contains procedures that use the chimeric
molecules of the invention to screen for new delivery activities or combinations of delivery
activities.  The invention further provides for the therapeutic use of said compositions as
anti-prolierative, anti-neoplastic, antibiotic, antiparasitic or antiviral agents.

<>

<1>Kuester, W., Alves, J.
<2>Mutational analysis of the function of Ile197 in the EcoRI restriction endonuclease.
<3>Biol. Chem. Hoppe Seyler
<4>376
<5>S122
<6>1995
<7>The endonuclease EcoRI is one of the best studied type II restriction enzymes.  It recognizes
and cleaves the double stranded DNA sequence GAATTC in both single strands between G and A
with high specificity.  Based on the X-ray structure Ile197 has been proposed to form a van
der Waals contact with cytosine of this recognition sequence.  We have exchanged Ile197 for
Ala, Arg and Met by site directed mutagenesis and analysed the purified mutant proteins
(I197A, M and R) biochemically and physicochemically.  Secondary structure composition
measured by CD-spectroscopy was unchanged for all mutants.  They cleave DNA with approximately
the same specific activity as the wild type enzyme.  For the I197A and the I197M mutants pH
optimum is shifted to alkaline conditions resulting in a 5-10 fold higher enzymatic activity
at pH 8.8.  Furthermore, the I197M mutant shows slight changes in its dependence on sequence
flanking the recognition sequence.  We assume that a hydrophobic contact of Ile197 to cytosine
is not important for activity of the EcoRI endonuclease.  But changes at this position
influence the conformation of the enzyme leading to a higher activity at alkaline pH on
substrates with a specific sequence surrounding.

<>

<1>Kuever, J. et al.
<2>Genome analysis of Desulfotomaculum gibsoniae strain Groll(T) a highly versatile  Gram-positive sulfate-reducing bacterium.
<3>Standards in Genomic Sciences
<4>9
<5>821-839
<6>2014
<7>Desulfotomaculum gibsoniae is a mesophilic member of the polyphyletic spore-forming genus
Desulfotomaculum within the family Peptococcaceae. This
bacterium was isolated from a freshwater ditch and is of interest because it can
grow with a large variety of organic substrates, in particular several aromatic
compounds, short-chain and medium-chain fatty acids, which are degraded
completely to carbon dioxide coupled to the reduction of sulfate. It can grow
autotrophically with H2 + CO2 and sulfate and slowly acetogenically with H2 +
CO2, formate or methoxylated aromatic compounds in the absence of sulfate. It
does not require any vitamins for growth. Here, we describe the features of D.
gibsoniae strain Groll(T) together with the genome sequence and annotation. The
chromosome has 4,855,529 bp organized in one circular contig and is the largest
genome of all sequenced Desulfotomaculum spp. to date. A total of 4,666 candidate
protein-encoding genes and 96 RNA genes were identified. Genes of the acetyl-CoA
pathway, possibly involved in heterotrophic growth and in CO2 fixation during
autotrophic growth, are present. The genome contains a large set of genes for the
anaerobic transformation and degradation of aromatic compounds, which are lacking
in the other sequenced Desulfotomaculum genomes.

<>

<1>Kuhlmann, U.C., Moore, G.R., James, R., Kleanthous, C., Hemmings, A.M.
<2>Structural parsimony in endonuclease active sites: should the number of homing endonuclease families be redefined?
<3>FEBS Lett.
<4>463
<5>1-2
<6>1999
<7>Homing endonucleases are classified into four families based on active site sequence motifs.
Through structural comparisons we have found structural similarities between the endonuclease
domain of colicin E9, an H-N-H motif-containing enzyme, and both the non-specific nuclease
from Serratia and I-PpoI, a His-Cys box-containing homing endonuclease. Our comparison
identifies conservation at the heart of all three enzyme active sites and so argues for a
re-classification of H-N-H and His-Cys box homing endonucleases as a single family. We suggest
the 'betabetaalpha-Me family' of homing enzymes to reflect the three elements of secondary
structure and the metal ion that define the motif.

<>

<1>Kuhn, H., Frank-Kamenetskii, M.D., Demidov, V.V.
<2>Site-specific nicking of duplex DNA using PNAs and restriction endonucleases.
<3>J. Biomol. Struct. Dyn.
<4>20
<5>916-917
<6>2003
<7>A variety of research procedures in molecular biology and biochemistry include as a
fundamental step the selective cleavage of a designated sequence in only one strand of
double-stranded DNA (DNA nicking).  To achieve the goal, restriction endonuclease-like nicking
enzymes can be employed.  However, very few nicking enzymes (nickases) are available and they
recognize short sequences of less than or equal to 7 bp.  It is therefore highly desirable to
develop new nicking systems with much higher sequence selectivity.  We design such systems
using homopyrimidine peptide nucleic acids (PNAs).  In our design we take advantage of the
ability of homopyrimidine PNAs to sequence-specifically invade duplex DNA.  When such PNAs are
targeted to two short homopurine stretches on the same strand of duplex DNA, the opposite DNA
strand becomes accessible for hybridization with an oligonucleotide.  We demonstrate that the
thus formed secondary DNA duplex can serve as a substrate for a restriction endonuclease,
provided that it contains the recognition sequence of the enzyme.  As a result, only one
strand of the parent DNA is cleaved leading to nicked duplex DNA after removal of PNAs.  All
restriction endonucleases tested by us (AluI, BbsI, BglII, KpnI, SbfI, and SphI), have worked
well in our design leading to quantitative yield of the nicked product.  Together with the
fact that the typical target site in our protocol spans about 20-25 bp, our results indicate
that a vast class of semi-synthetic rare-cleaving DNA nickases has been generated.

<>

<1>Kuhn, H., Hu, Y., Frank-Kamenetskii, M.D., Demidov, V.V.
<2>Artificial site-specific DNA-nicking system based on common restriction enzymes assisted by PNA openers.
<3>Biochemistry
<4>42
<5>4985-4992
<6>2003
<7>We report on the peptide nucleic acid (PNA)-directed design of a DNA-nicking system that
enables selective and quantitative cleavage of one
strand of duplex DNA at a designated site, thus mimicking natural nickases
and significantly extending their potential. This system exploits the
ability of pyrimidine PNAs to serve as openers for specific DNA sites by
invading the DNA duplex and exposing one DNA strand for oligonucleotide
hybridization. The resultant secondary duplex can act as a substrate for a
restriction enzyme, which ultimately creates a nick in the parent DNA. We
demonstrate that several restriction enzymes of different types could be
successfully used in the PNA-assisted system we developed. Importantly,
the enzyme cleavage efficiency is basically not impaired on such
artificially generated substrates, compared with the efficiency on regular
DNA duplexes. Our design originates a vast class of semisynthetic
rare-cleaving DNA nickases, which are essentially absent at present. In
addition, we show that the site-specific PNA-assisted nicking of duplex
DNA can be engaged in a rolling-circle DNA amplification (RCA) reaction.
This new RCA format demonstrates the practical potential of the novel
biomolecular tool we propose for DNA technology and DNA diagnostics.

<>

<1>Kuhn, I., Stephenson, F., Boyer, H., Greene, P.
<2>Positive-selection vectors utilizing lethality of the EcoRI endonuclease.
<3>Gene
<4>42
<5>253-263
<6>1986
<7>The construction and use of a series of positive-selection vectors are
described.  These plasmids encode EcoRI endonuclease, the synthesis of which is
under the control of the lacUV5 promoter.  The pKG2 plasmid encodes a wild-type
EcoRI endonuclease.  In the absence of EcoRI methylase, the endonuclease is
lethal.  Cloning into any of the unique restriction sites within the
endonuclease-coding gene allows survival of the transformed
EcoRI-methylase-less host.  The pKGW and pKGS plasmids encode an altered EcoRI
endonuclease which, when repressed in the lacIQ host, allows survival in the
absence of the methylase.  Induction with IPTG, however, results in cell death
as a result of high-level EcoRI synthesis.  Cloning into any of the unique
restriction sites within the EcoRI gene of pKGW or pKGS allows survival of
derepressed transformed cells.  These vectors strongly select for cloning
events which inactivate the endonuclease gene.

<>

<1>Kuhnlein, U., Arber, W.
<2>Host specificity of DNA produced by Escherichia coli. XV.  The role of nucleotide methylation in in vitro B-specific modification.
<3>J. Mol. Biol.
<4>63
<5>9-19
<6>1972
<7>It is shown that in vitro Escherichia coli strain B-specific modification of
the replicative form of bacteriophage fd DNA is accompanied by the methylation
of certain adenine moieties to form N-6-methyladenine.  The reaction follows
first order kinetics and saturation is reached when about four adenines are
methylated per replicative form.  No methyl groups are transferred to
B-modified DNA.  The replicative form of a one step mutant of fd, which has a
reduced sensitivity towards B-specific restriction, has lost two of the four
methyl acceptor sites.  The replicative form of a second step mutant, which is
not subject to B-specific restriction, is completely refractory to methylation
by the modification enzyme.  It is therefore concluded that the B-modification
and the B-restriction enzyme react with the same sites on the substrate DNA and
that the replicative form of wild type fd has two such sites.  The number of
N-6-methyladenines per B-specificity site of fully modified double-stranded DNA
is two.

<>

<1>Kuhnlein, U., Linn, S., Arber, W.
<2>Host specificity of DNA produced by Escherichia coli, XI.  In vitro modification of phage fd replicative form.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>63
<5>556-562
<6>1969
<7>An enzymatic activity, having the properties expected by a B-specific
host-controlled modification enzyme, has been purified from an extract of
Escherichia coli strain B.  This activity renders the unmodified replicative
form of phage fd resistant to B-specific restriction and is only present in
strains carrying intact genes for type B modification.  In phosphate buffer,
the enzyme acts optimally at pH 6 and is dependent upon a single cofactor,
S-adenosylmethionine.

<>

<1>Kuhnlein, U., Linn, S., Arber, W.
<2>In vitro modifikation der replikativen form des bakteriophagen fd.
<3>Pathol. Microbiol. (Basel)
<4>34
<5>136
<6>1969
<7>None

<>

<1>Kukutla, P., Lindberg, B.G., Pei, D., Rayl, M., Yu, W., Steritz, M., Faye, I., Xu, J.
<2>Draft Genome Sequences of Elizabethkingia anophelis Strains R26T and Ag1 from the Midgut of the Malaria Mosquito Anopheles gambiae.
<3>Genome Announcements
<4>1
<5>e01030-13
<6>2013
<7>Elizabethkingia anophelis is a species in the family Flavobacteriaceae. It is a dominant
resident in the mosquito gut and also a human pathogen. We present the
draft genome sequences of two strains of E. anophelis, R26(T) and Ag1, which were
isolated from the midgut of the malaria mosquito Anopheles gambiae.

<>

<1>Kulakauskas, S., Barsomian, J.M., Lubys, A., Roberts, R.J., Wilson, G.G.
<2>Organization and sequence of the HpaII restriction-modification system and adjacent genes.
<3>Gene
<4>142
<5>9-15
<6>1994
<7>We report the organization of the HpaII restriction and modification (R-M) system from
Haemophilus parainfluenzae (recognition sequence: 5'...CCGG...3'), the sequence of the gene
coding for the HpaII restriction endonuclease, and the sequence of the upstream flanking DNA.
The HpaII system comprises two genes, hpaIIM, coding for the methyltransferase (MTase; 358
amino acids (aa), 40.4 kDa: product, Cm5CGG), and hpaIIR, coding for the restriction
endonuclease (ENase: 358aa, 40.9 kDa: product, C'CGG). The genes are adjacent, they have the
same orientation, and they occur in the order hpaIIM then hpaIIR. The ENase bears little aa
sequence similarity to the isoschizomeric R.BsuFI and R.MspI ENases. Upstream of, and partly
overlapping hpaIIM is the coding sequence for a 141-aa protein that resembles the
very-short-patch-repair endonuclease (Vsr) of Escherichia coli. Upstream of that is the coding
sequence for a protein that resembles valyl-tRNA synthetase (ValS).

<>

<1>Kulakauskas, S., Lubys, A., Ehrlich, S.D.
<2>DNA restriction-modification systems mediate plasmid maintenance.
<3>J. Bacteriol.
<4>177
<5>3451-3454
<6>1995
<7>Two plasmid-carried restriction-modification (R-M) systems, EcoRI (from pMB1 of Escherichia
coli) and Bsp6I (from pXH13 of Bacillus sp. strain RFL6), enhance plasmid segregational
stability in E. coli and Bacillus subtilis, respectively. Inactivation of the endonuclease or
the presence of the methylase in trans abolish the stabilizing activity of the R-M systems. We
propose that R-M systems mediate plasmid segregational stability by postsegregational killing
of plasmid-free cells. Plasmid-encoded methyltransferase modifies host DNA and thus prevents
its digestion by the restriction endonuclease. Plasmid loss entails degradation and/or
dilution of the methylase during cell growth and appearance of unmethylated sites in the
chromosome. Double-strand breaks, introduced at these sites by the endonuclease, eventually
cause the death of the plasmid-free cells. Contribution to plasmid stability is a previously
unrecognized biological role of the R-M systems.

<>

<1>Kulandai, L.T., Lakshmipathy, D., Ramasubban, G., Vetrivel, U., Rao, M.H., Rathinam, S., Narasimhan, M.
<2>Whole-Genome Sequencing and Mutation Analysis of Two Extensively Drug-Resistant Sputum Isolates of Mycobacterium tuberculosis (VRFCWCF XDRTB 232 and VRFCWCF  XDRTB 1028) from Chennai, India.
<3>Genome Announcements
<4>2
<5>e01173-14
<6>2014
<7>We announce the draft genome sequence of two extensively drug-resistant Mycobacterium
tuberculosis strains, VRFCWCF XDRTB 232 and VRFCWCF XDRTB 1028,
isolated from the sputum samples of a patient clinically suspected to have
tuberculosis, and we also report novel mutations that confer drug resistance.

<>

<1>Kulasekara, B.R. et al.
<2>Analysis of the genome of the Escherichia coli O157:H7 2006 spinach-associated outbreak isolate indicates candidate genes that may enhance virulence.
<3>Infect. Immun.
<4>77
<5>3713-3721
<6>2009
<7>In addition to causing diarrhea, Escherichia coli O157:H7 infection can
lead to hemolytic uremic syndrome (HUS), a severe disease characterized by
hemolysis and renal failure. Differences in HUS frequency among E. coli
O157:H7 outbreaks have been noted, but there is incomplete understanding
of bacterial factors that promote HUS. In 2006, an outbreak of E. coli
O157:H7, caused by consumption of contaminated spinach, occurred with a
notably high frequency of HUS. We sequenced the genome of this strain
(TW14359) with the goal of identifying candidate genetic factors that
contribute to an enhanced ability to cause HUS. The TW14359 genome
contains 70-kb of DNA segments not present in either of the two reference
O157:H7 genomes. We identified seven putative virulence determinants,
including two putative Type III secretion system effector proteins,
candidate genes that could result in increased pathogenicity or,
alternatively, adaptation to plants, and an intact anaerobic nitric oxide
reductase gene, norV. We surveyed one hundred O157:H7 isolates for the
presence of these putative virulence determinants. A norV deletion was
found in over half of strains surveyed and correlates strikingly with the
absence of stx1. The other putative virulence factors were found in eight
to thirty-five percent of O157:H7 isolates surveyed and their presence
also correlates with the presence of norV and absence of stx1 indicating
the presence of norV may serve as a marker of greater propensity for HUS
similar to the correlation between the absence of stx1 and HUS propensity.

<>

<1>Kulba, A.M., Abdel-Sabur, M.S., Butkus, V.V., Janulaitis, A., Fomichev, Y.K.
<2>New type-II restrictase from cells of Erwinia herbicola.
<3>Mol. Biol. (Mosk)
<4>21
<5>250-254
<6>1987
<7>Forty strains of the bacterial genus Erwinia were tested for the presence of
restrictases.  Type-II restrictases identical in properties were isolated and
partially purified from cells of Erw. herbicola strains 9/5 and 8608.  It was
established that restrictase EheI recognizes the sequence of nucleotides
5'-GGCGCC-3' and is a false isoschizomer of enzymes NarI, NdaI, NunII, and
BbeI, since EheI cleaves the sequence in the middle, forming blunt ends, as
opposed to the enzymes listed, which form sticky 5'- or 3'-ends when cleaving
DNA.  Optimal conditions of cleavage of DNA by EheI restrictase are determined.

<>

<1>Kulba, A.M., Elgammudi, A.A.
<2>Detection of restriction endonucleases in Aeromonas bacteria.
<3>Vestn. Beloruss. Gos. Univ.
<4>2
<5>47-49
<6>1997
<7>
<>

<1>Kuleshov, K.V., Vodop'ianov, S.O., Dedkov, V.G., Markelov, M.L., Kermanov, A.V., Kruglikov, V.D., Vodop'ianov, A.S., Pisanov, R.V., Chemisova, O.S., Mazrukho, A.B., Titova, S.V., Shipulin, G.A.
<2>Draft Genome Sequencing of Vibrio cholerae O1 El Tor Isolates Collected in the Russian Federation from Imported Cholera Cases.
<3>Genome Announcements
<4>2
<5>e00624-14
<6>2014
<7>We report the draft genome sequencing of five Vibrio cholerae O1 El Tor clinical  isolates
collected in the Russian Federation from imported cholera cases in 2006,
2010, and 2012. In the initial phylogenetic analysis, one isolate clustered with
the Haiti/Nepal-4 group.

<>

<1>Kuleshov, K.V., Vodop'ianov, S.O., Markelov, M.L., Dedkov, V.G., Kermanov, A.V., Kruglikov, V.D., Vodop'ianov, A.S., Pisanov, R.V., Mazrukho, A.B., Shipulin, G.A.
<2>Draft Genome Sequences of Vibrio cholerae O1 ElTor Strains 2011EL-301 and P-18785, Isolated in Russia.
<3>Genome Announcements
<4>1
<5>e00659-13
<6>2013
<7>We report the draft whole-genome sequences of two Vibrio cholerae O1 strains, the
environmental toxigenic strain 2011EL-301 and the clinical nontoxigenic strain
P-18785, both isolated in Russia. Some basic data comparing the two against the
GenBank repository are provided.

<>

<1>Kulik, E.M., Bickle, T.A.
<2>Regulation of the activity of the type IC EcoR124I restriction enzyme.
<3>J. Mol. Biol.
<4>264
<5>891-906
<6>1996
<7>Restriction-modification systems must regulate the expression of their genes so that the
chromosomal genome is modified at all times by the methyltransferase to protect the host cell
from the potential lethal action of the cognate restriction endonuclease.  Since type I R-M
systems can be transferred to non-modified Escherichia coli cells by conjugation or
transformation without killing the recipient, they must have some means to regulate their
restriction activity upon entering a new host cell to avoid restriction of unprotected host
DNA and cell death.  This is especially true for EcoR124I, a type IC family member, which is
coded for by a conjugative plasmid.  Control of EcoR124I restriction activity is most likely
at the post-translational level as the transfer of the EcoR124I system into a recipient cell
that already expressed the HsdR subunit of this system was not a lethal event.  Additionally,
the kinetics of restriction activity upon transfer of the genes coding for the EcoR124I RM
system to a recipient cell are the same, irrespective of the modification state of the
recipient cell or the presence or absence of the EcoR124I HsdR subunit in the new host cells.
The mechanism controlling the restriction activity of a type IC R-M system upon transfer to a
new host cell is different from that controlling the chromosomally coded type IA and IB R-M
systems.  The previously discovered hsdC mutant, which affects the establishment of the type
IA system EcoKI, was shown to affect the establishment of the type IB system EcoAI, but to
have no influence on EcoR124I.

<>

<1>Kumagai, Y., Yoshizawa, S., Oshima, K., Hattori, M., Iwasaki, W., Kogure, K.
<2>Complete Genome Sequence of Winogradskyella sp. Strain PG-2, a Proteorhodopsin-Containing Marine Flavobacterium.
<3>Genome Announcements
<4>2
<5>e00490-14
<6>2014
<7>Winogradskyella sp. strain PG-2 is a marine flavobacterium isolated from surface  seawater.
This organism contains proteorhodopsin, which can convert light energy
into available forms of biochemical energy. Here, we present its complete genome
sequence and annotation, which provide further insights into the life strategy of
proteorhodopsin-mediated phototrophy in the ocean.

<>

<1>Kumano, M., Sakakibara, M.
<2>Recombination of DNA in blue-green algae - restriction enzyme systems and transfection vectors.
<3>Baiosaiensu to Indasutori
<4>46
<5>38-41
<6>1988
<7>One table describes the presence of a number of new restriction enzymes in
seven strains of blue-green algae.  It is concluded that African and American
strains of Spirulina can be clearly distinguished from one another on the basis
of restriction enzyme content.

<>

<1>Kumar, A., Garg, N.
<2>Restriction endonucleases.
<3>Genet. Eng. (N Y)
<4>0
<5>55-74
<6>2005
<7>Restriction endonucleases bind specifically to and cleave the sugar phosphate backbone (bond
between deoxyribose and phosphate groups) in double stranded DNA at specific sites within or
adjacent to a particular sequence known as recognition sequence.  Restriction endonucleases
are of three types: Type I, Type II and Type III.  Types I and III restriction endonucleases
carry modification (like methylation) and ATP dependent cleavage activities in the same
protein.  Type III restriction endonucleases cleave the DNA at the recognition site and then
dissociate from the substrate.  However, type I restriction endonucleases bind to the
recognition sequence but cleave at random sites when the DNA loops back to the bound enzyme.
Neither type I nor type III restriction endonucleases are widely used in genetic engineering.
Type II restriction endonucleases are commonly used in genetic engineering.  They are the
invaluable molecular scissors.  These restriction endonucleases cut both strands of duplex DNA
with in a stretch of just a few bases.  Therefore, only type II restriction enonucleases are
described in this chapter.  Nomenclature of the restriction endonucleases is based on the
source of their isolation from an organism.  In case of restriction endonucleases, enzyme
activity is described in units considering one unit of the restriction endonuclease as the
amount of the enzyme required to hydrolyze one microgram of lambda DNA to completion in one
hour at the optimum temperature in a total volume of 50 ul of the assay mixture.  The lambda
DNA, due to its large size (approximately 50 Kb), is considered to have recognition sites
almost for all the restriction endonucleases.

<>

<1>Kumar, B.K., Deekshit, V.K., Rai, P., Gurtler, V., Karunasagar, I., Karunasagar, I.
<2>Draft Genome Sequence of trh+ Vibrio parahaemolyticus VP-49, Isolated from Seafood Harvested along the Mangalore Coast, India.
<3>Genome Announcements
<4>2
<5>e00607-14
<6>2014
<7>Vibrio parahaemolyticus is a seafood-borne pathogen autochthonous to the marine and estuarine
ecosystem, which is responsible for gastroenteritis due to the
consumption of contaminated raw seafood. Here, we report the draft genome
sequence of V. parahaemolyticus VP-49, isolated from seafood, to identify the
different virulence attributes and to study the mechanisms that enhance its
environmental fitness.

<>

<1>Kumar, M., Khanna, S.
<2>Diversity of 16S rRNA and dioxygenase genes detected in coal-tar-contaminated site undergoing active bioremediation.
<3>J. Appl. Microbiol.
<4>108
<5>1252-1262
<6>2010
<7>AIMS: In order to develop effective bioremediation strategies for
polyaromatic hydrocarbons (PAHs) degradation, the composition and
metabolic potential of microbial communities need to be better understood,
especially in highly PAH contaminated sites in which little information on
the cultivation-independent communities is available. METHODS AND RESULTS:
Coal-tar-contaminated soil was collected, which consisted of 122.5 mg
g(-1) total extractable PAH compounds. Biodegradation studies with this
soil indicated the presence of microbial community that is capable of
degrading the model PAH compounds viz naphthalene, phenanthrene and pyrene
at 50 ppm each. PCR clone libraries were established from the DNA of the
coal-tar-contaminated soil, targeting the 16S rRNA to characterize (i) the
microbial communities, (ii) partial gene fragment encoding the Rieske iron
sulfur center (alpha-subunit) common to all PAH dioxygenase enzymes and
(iii) beta-subunit of dioxygenase. Phylotypes related to Proteobacteria
(Alpha-, Epsilon- and Gammaproteobacteria), Acidobacteria, Actinobacteria,
Firmicutes, Gemmatimonadetes and Deinococci were detected in 16S rRNA
derived clone libraries. Many of the gene fragment sequences of
alpha-subunit and beta-subunit of dioxygenase obtained from the respective
clone libraries fell into clades that are distinct from the reference
dioxygenase gene sequences. Presence of consensus sequence of the Rieske
type [2Fe-2S] cluster binding site suggested that these gene fragments
encode for alpha-subunit of dioxygenase gene. CONCLUSIONS: Sequencing of
the cloned libraries representing alpha-subunit gene fragments (Rf1) and
beta-subunit of dioxygenase showed the presence of hitherto unidentified
dioxygenase in coal-tar-contaminated soil. SIGNIFICANCE AND IMPACT OF THE
STUDY: The combination of the Rieske primers and bacterial community
profiling represents a powerful tool for both assessing bioremediation
potential and the exploration of novel dioxygenase genes in a contaminated
environment.

<>

<1>Kumar, M.R., Hosur, R.V., Roy, K.B., Miles, H.T., Govil, G.
<2>Resonance assignment of the 500-MHz proton NMR spectrum of self-complementary dodecanucleotide d-GGATCCGGATCC:  altered conformations at BamHI cleavage sites.
<3>Biochemistry
<4>24
<5>7703-7711
<6>1985
<7>Resonance assignments of nonexchangeable base and sugar protons of the
self-complementary dodecanucleotide d-GGATCCGGATCC have been obtained by
two-dimensional NMR methods and strategies derived from interproton distance
calculations on different secondary structures of nucleic acids.
Conformational details about the glycosidic dihedral angle and sugar pucker
have been derived from the relative intensities of cross peaks in the
two-dimensional J-correlated and nuclear Overhauser enhancement correlated
spectra in D2O solution.  It is observed that d-GGATCCGGATCC assumes a
predominantly B-type conformation with sequence-dependent changes along the
chain.  The recognition site of BamHI shows a distinctly different geometrical
environment.  The sugar rings of G1 and G7 assume a C3'-endo geometry while the
rest of the sugars possess C2'-endo geometry.

<>

<1>Kumar, N., Mukhopadhyay, A.K., Patra, R., De R, B.R., Shaik, S., Alam, J., Tiruvayipati, S., Ahmed, N.
<2>Next-Generation Sequencing and De Novo Assembly, Genome Organization, and Comparative Genomic Analyses of the Genomes of Two Helicobacter pylori Isolates  from Duodenal Ulcer Patients in India.
<3>J. Bacteriol.
<4>194
<5>5963-5964
<6>2012
<7>The prevalence of different H. pylori genotypes in various geographical regions indicates
region-specific adaptations during the course of evolution. Complete
genomes of H. pylori from countries with high infection burdens, such as India,
have not yet been described. Herein we present genome sequences of two H. pylori
strains, NAB47 and NAD1, from India. In this report, we briefly mention the
sequencing and finishing approaches, genome assembly with downstream statistics,
and important features of the two draft genomes, including their phylogenetic
status. We believe that these genome sequences and the comparative genomics
emanating thereupon will help us to clearly understand the ancestry and biology
of the Indian H. pylori genotypes, and this will be helpful in solving the
so-called Indian enigma, by which high infection rates do not corroborate the
minuscule number of serious outcomes observed, including gastric cancer.

<>

<1>Kumar, R., Dwivedi, V., Negi, V., Khurana, J.P., Lal, R.
<2>Draft Genome Sequence of Sphingobium lactosutens Strain DS20T, Isolated from a Hexachlorocyclohexane Dumpsite.
<3>Genome Announcements
<4>1
<5>e00753-13
<6>2013
<7>Sphingobium lactosutens DS20(T) has been isolated from the hexachlorocyclohexane  (HCH)
dumpsite in Lucknow, India, but does not degrade any of the HCH isomers.
Here, we present the ~5.36-Mb draft genome sequence of strain DS20(T), which
consists of 110 contigs and 5,288 coding sequences, with a G+C content of 63.1%.

<>

<1>Kumar, R., Mukhopadhyay, A.K., Ghosh, P., Rao, D.N.
<2>Comparative Transcriptomics of H. pylori Strains AM5, SS1 and Their hpyAVIBM Deletion Mutants: Possible Roles of Cytosine Methylation.
<3>PLoS ONE
<4>7
<5>e42303
<6>2012
<7>Helicobacter pylori is an important human pathogen and one of the most successful chronic
colonizers of the human body. H. pylori uses diverse
mechanisms to modulate its interaction with the host in order to
promote chronic infection and overcome host immune response.
Restriction-modification genes are a major part of strain-specific
genes present in H. pylori. The role of N-6 -adenine methylation in
bacterial gene regulation and virulence is well established but not
much is known about the effect of C-5 -cytosine methylation on gene
expression in prokaryotes. In this study, it was observed by microarray
analysis and RT-PCR, that deletion of an orphan C-5 -cytosine
methyltransferase, hpyAVIBM in H. pylori strains AM5and SS1 has a
significant effect on the expression of number of genes belonging to
motility, adhesion and virulence. AM Delta DhpyAVIBM mutant strain has
a different LPS profile and is able to induce high IL-8 production
compared to wild-type. hpyAVIBM from strain 26695 is able to complement
mutant SS1 and AM5 strains. This study highlights a possible
significance of cytosine methylation in the physiology of H. pylori.

<>

<1>Kumar, R., Mukhopadhyay, A.K., Rao, D.N.
<2>Characterization of an N6 adenine methyltransferase from Helicobacter pylori strain 26695 which methylates adjacent adenines on the same strand.
<3>FEBS J.
<4>277
<5>1666-1683
<6>2010
<7>Genomic sequences of Helicobacter pylori strains 26695, J99, HPAGI and G27 have revealed an
abundance of restriction and modification genes.
hp0050, which encodes an N6 adenine DNA methyltransferase, was cloned,
overexpressed and purified to near homogeneity. It recognizes the
sequence 5'-GRRG-3' (where R is A or G) and, most intriguingly,
methylates both adenines when R is A (5'-GAAG-3'). Kinetic analysis
suggests a nonprocessive (repeated-hit) mechanism of methylation in
which HP0050 methyltransferase methylates one adenine at a time in the
sequence 5'-GAAG-3'. This is the first report of an N6 adenine DNA
methyltransferase that methylates two adjacent residues on the same
strand. Interestingly, HP0050 homologs from two clinical strains of H.
pylori (PG227 and 128) methylate only 5'-GAGG-3' compared with
5'-GRRG-3' in strain 26695. HP0050 methyltransferase is highly
conserved as it is present in more than 90% of H. pylori strains.
Inactivation of hp0050 in strain PG227 resulted in poor growth,
suggesting its role in the biology of H. pylori. Collectively, these
findings provide impetus for exploring the role(s) of this conserved
DNA methyltransferase in the cellular processes of H. pylori.

<>

<1>Kumar, R., Rao, D.N.
<2>A nucleotide insertion between two adjacent methyltransferases in Helicobacter pylori results in a bifunctional DNA methyltransferase.
<3>Biochem. J.
<4>433
<5>487-495
<6>2011
<7>Helicobacter pylori has a dynamic R-M (restriction-modification) system. It is capable of
acquiring new R-M systems from the environment
in the form of DNA released from other bacteria or other H. pylon
strains. Random mutations in RM genes can result in non-functional R-M
systems or R-M systems with new properties. hpyAVIAM and hpyAVIBM are
two solitary DNA MTase (methyltransferase) genes adjacent to each other
and lacking a cognate restriction enzyme gene in H. pylori strain
26695. Interestingly, in an Indian strain D27, hpyAVIAM hpyAVIBM
encodes a single bifunctional polypeptide clue to insertion of a
nucleotide just before the stop codon of hpyAVIBM and, when a similar
mutation was made in hpyAVIAM hpyAVIBM from strain 26695, a functional
MTase with an N-terminal C-5-cytosine MTase domain and a C-terminal
N-6-adenine MTase domain was constructed. Mutations in the AdoMet
(S-adenosylmethionine)-binding motif or in the catalytic motif of
M.HpyAVIA or M.HpyAVIB selectively abrogated the C-5-cytosine or
N-6-adenine methylation activity of M.HpyAVIA-M.HpyAVIB fusion protein.
The present study highlights the ability of H. pylori to evolve genes
with unique functions and thus generate variability. For organisms such
as H. pylori, which have a small genome, these adaptations could be
important for their survival in the hostile host environment.

<>

<1>Kumar, R., Rao, D.N.
<2>Role of DNA methyltransferases in epigenetic regulation in bacteria.
<3>Subcell. Biochem.
<4>61
<5>81-102
<6>2012
<7>In prokaryotes, alteration in gene expression was observed with the modification  of DNA,
especially DNA methylation. Such changes are inherited from generation to
generation with no alterations in the DNA sequence and represent the epigenetic
signal in prokaryotes. DNA methyltransferases are enzymes involved in DNA
modification and thus in epigenetic regulation of gene expression. DNA
methylation not only affects the thermodynamic stability of DNA, but also changes
its curvature. Methylation of specific residues on DNA can affect the protein-DNA
interactions. DNA methylation in prokaryotes regulates a number of physiological
processes in the bacterial cell including transcription, DNA mismatch repair and
replication initiation. Significantly, many reports have suggested a role of DNA
methylation in regulating the expression of a number of genes in virulence and
pathogenesis thus, making DNA methlytransferases novel targets for the designing
of therapeutics. Here, we summarize the current knowledge about the influence of
DNA methylation on gene regulation in different bacteria, and on bacterial
virulence.

<>

<1>Kumar, R., Sabareesh, V., Mukhopadhyay, A.K., Rao, D.N.
<2>Mutations in hpyAVIBM, C5 cytosine DNA methyltransferase from Helicobacter pylori result in relaxed specificity.
<3>FEBS J.
<4>279
<5>1080-1092
<6>2012
<7>The genome of Helicobacter pylori is rich in restrictionmodification (RM) systems.
Approximately 4% of the genome codes for components of RM
systems. hpyAVIBM, which codes for a phase-variable C5 cytosine
methyltransferase (MTase) from H. pylori, lacks a cognate restriction
enzyme. Over-expression of M.HpyAVIB in Escherichia coli enhances the
rate of mutations. However, when the catalytically inactive F9N or C82W
mutants of M.HpyAVIB were expressed in E. coli, mutations were not
observed. The M.HpyAVIB gene itself was mutated to give rise to
different variants of the MTase. M.HpyAVIB variants were purified and
differences in kinetic properties and specificity were observed.
Intriguingly, purified MTase variants showed relaxed substrate
specificity. Homologues of hpyAVIBM homologues amplified and sequenced
from different clinical isolates showed similar variations in sequence.
Thus, hpyAVIBM presents an interesting example of allelic variations in
H. pylori where changes in the nucleotide sequence result in proteins
with new properties.

<>

<1>Kumar, R., Srivastava, R., Singh, R.K., Surolia, A., Rao, D.N.
<2>Activation and inhibition of DNA methyltransferases by S-adenosyl-L-homocysteine analogues.
<3>Bioorg. Med. Chem.
<4>16
<5>2276-2285
<6>2008
<7>The inhibition of methyltransferases is currently of high interest, particularly in the areas
of microbial infection and cell
proliferation, as there have been serious attempts to develop novel
anti-microbial agents. In the present investigation, a series of
11S-adenosyl-L-homocysteine analogues have been synthesized and effect
of these analogues on DNA methylation catalyzed by DNA
methyltransferases was studied. It was found that, while
5'-S-(propionic acid) 5-deoxy-9-(1'-beta-D-ribofuranosyl) 1,
3-dideazaadenine was an activator of EcoP15I and HhaI DNA
methyltransferases, 5'-S-(propionic acid)
5'-deoxy-9-(1'-beta-dribofuran osyl)adenine inhibited the
methyltransferases in a non-competitive manner. An understanding of the
binding of analogues to DNA methyltransferases will greatly assist the
design of novel anti-microbial compounds.

<>

<1>Kumar, R.A., Vaze, M.B., Chandra, N.R., Vijayan, M., Muniyappa, K.
<2>Functional characterization of the precursor and spliced forms of RecA protein of Mycobacterium tuberculosis.
<3>Biochemistry
<4>35
<5>1793-1802
<6>1996
<7>The recA locus of pathogenic mycobacteria differs from that of
nonpathogenic species because it contains large intervening sequences nested in the RecA
homology region that are excised by an unusual protein-splicing reaction.  In vivo assays
indicated that Mycobacterium tuberculosis recA partially complemented Escherichia coli
recA mutants for recombination and mutagenesis.  Further, splicing of the 85 kDa
precursor to 38 kDa MtRecA protein was necessary for the display of its activity, in vivo.
To gain insights into the molecular basis for partial and lack of complementation by
MtRecA and 85 kDa proteins, respectively, we purified both of them to homogeneity.
MtRecA protein, but not the 85 kDa form, bound stoichiometrically to single-stranded
DNA in the presence of ATP.  MtRecA protein was cross-linked to 8-azidoadenosine 5'-
triphosphate with reduced efficiency, and kinetic analysis of ATPase activity suggested that
it is due to decreased affinity for ATP.  In contrast, the 85 kDa form was unable to bind
ATP, in the presence or absence of ssDNA and, consequently, was entirely devoid of
ATPase activity.  Molecular modeling studies suggested that the decreased affinity of
MtRecA protein for ATP and the reduced efficiency of its hydrolysis might be due to the
widening of the cleft which alters the hydrogen bonds and the contact area between the
enzyme and the substrate and changes in the disposition of the amino acid residues around
the magnesium ion and the gamma-phosphate.  The formation of joint molecules promoted
by MtRecA protein was stimulated by SSB when the former was added first.  The
probability of an association between the lack and partial levels of biological activity of
RecA protein(s) to that of illegitimate recombination in pathogenic mycobacteria is
considered.

<>

<1>Kumar, S., Bala, M., Raghava, G.P., Mayilraj, S.
<2>Draft Genome Sequence of Rhodococcus triatomae Strain BKS 15-14.
<3>Genome Announcements
<4>1
<5>e0012913
<6>2013
<7>We report the 5.8-Mb genome sequence of Rhodococcus triatomae BKS 15-14, isolated from an ant
hill soil sample, collected from Bhitarkanika Mangrove Reserve
Forest, Odisha, India. The draft genome of strain BKS 15-14 consists of 5,824,349
bp, with a G+C content of 69%, 5,387 protein-coding genes, and 57 RNAs.

<>

<1>Kumar, S., Cheng, X., Klimasauskas, S., Sha, M., Posfai, J., Roberts, R.J., Wilson, G.G.
<2>The DNA (cytosine-5) methyltransferases.
<3>Nucleic Acids Res.
<4>22
<5>1-10
<6>1994
<7>A review of the m5C-methyltransferases relating sequence to structure and function.

<>

<1>Kumar, S., Cheng, X., Pflugrath, J.W., Roberts, R.J.
<2>Purification, crystallization, and preliminary X-ray diffraction analysis of an M.HhaI-AdoMet complex.
<3>Biochemistry
<4>31
<5>8648-8653
<6>1992
<7>The type-II DNA-(cytosine-5)-methyltransferase M.HhaI was overexpressed in Escherichia coli
and purified to apparent homogeneity. The purification scheme exploits a unique high salt
back-extraction step to solubilize M.HhaI selectively, followed by FPLC chromatography. The
yield of purified protein was 0.75-1.0 mg per gram of bacterial paste. M.HhaI could be
isolated in two forms: bound with its cofactor S-adenosylmethionine (AdoMet) or devoid of the
cofactor. The AdoMet-bound form was capable of methylating DNA in vitro in the absence of
exogenous AdoMet. From kinetic studies of the purified enzyme, values for KmAdoMet(60 nM),
KiAdoHyc(0.4 nM), and Kcat (0.22s-1) were determined. The purified enzyme bound with its
cofactor was crystallized by the hanging drop vapor diffusion technique. Crystals were of
monoclinic space group P21 and had unit-cell dimensions of a = 55.3 A, b = 72.7 A, c = 91.0 A,
and B=102.5o, with two molecules of M.HhaI in each of the two asymmetric units. The crystals
diffract beyond 2.5 A and are suitable for structure determination.

<>

<1>Kumar, S., Horton, J.R., Jones, G.D., Walker, R.T., Roberts, R.J., Cheng, X.
<2>DNA containing 4'-thio-2'-deoxycytidine inhibits methylation by HhaI methyltransferase.
<3>Nucleic Acids Res.
<4>25
<5>2773-2783
<6>1997
<7>4'-Thio-2'-deoxycytidine was synthesized as a 5'-protected phosphoramidite compatible with
solid phase DNA synthesis.  When incorporated as the target cytosine (C*) in the GC*GC
recognition sequence for the DNA methyltransferase M.HhaI, methyl transfer was strongly
inhibited.  In contrast, these same oligonucleotides were normal substrates for the cognate
restriction endonuclease R.HhaI and its isoschizomer R.HindP1I.  M.HhaI was able to bind both
4'-thio-modified DNA and unmodified DNA to equivalent extents under equilibrium conditions.
However, the presence of 4'-thio-2'-deoxycytidine decreased the half-life of the complex by
>10-fold.  The crystal stucture of a ternary complex of M.HhaI, AdoMet and DNA containing
4'-thio-2'-deoxycytidine was solved at 2.05 A resolution with a crystallographic R-factor of
0.186 and R-free of 0.231.  The structure is not grossly different from previously solved
ternary complexes containing M.HhaI, DNA and AdoHcy.  The difference electron density suggests
partial methylation at C5 of the flipped target 4'-thio-2'-deoxycytidine.  The inhibitory
effect of the 4' sulfur atom on enzymatic activity may be traced to perturbation of a step in
the methylation reaction after DNA binding but prior to methyl transfer.  This inhibitory
effect can be partially overcome after a considerably long time in the crystal environment
where the packing prevents complex dissociation and the target is accurately positioned within
the active site.

<>

<1>Kumar, S., Johnson, W.S., Tomasz, M.
<2>Orientation isomers of the mitomycin C interstrand cross-link in non-self-complementary DNA. Differential effect of the two isomers on restriction endonuclease cleavage at a nearby site.
<3>Biochemistry
<4>32
<5>1364-1372
<6>1993
<7>Reductively activated mitomycin C (MC) forms DNA interstrand cross-links between two guanines
at CG.CG sequences. It is predictable that such cross-links should occur in two isomeric
strand orientations in duplex DNA (except when located in the center of a self-complementary
duplex). This was verified by the isolation and characterization of a pair of two isomeric
oligonucleotides in each case of five non-self-complementary duplexes of 8-bp length,
cross-linked by MC. Isomer separation was accomplished by reverse-phase HPLC. The isomers in a
pair were formed in approximately 1:1 proportion.Their structures were rigorously
characterized by a two-step cross-linking procedure: first 1-monoalkylation of each strand,
followed by conversion to a cross-linked duplex by annealing the monoalkylated strand to its
complement in the presence of a reducing agent. The resulting individual authentic orientation
isomers were used as standards for identification of the two isomers formed in the original
(one-step) cross-linking reactions. A 16-bp duplex oligonucleotide was synthesized featuring
the AluI cognate sequence, separated from a MC cross-link site by only 1 bp. Its two MC
cross-linked isomers were prepared separately, and their rate of cleavage by AluI was
determined using HPLC. Cleavage of both the unmodified and cross-linked duplexes was
nonsymmetrical. The isomer in which the 2-NH3+ of MC is oriented toward the AluI site was
cleaved essentially at the same rate as the control duplex, while cleavage of the isomer with
the MC indoloquinone group oriented toward the AluI site was inhibited 2-fold at the
faster-cleaved strand. This difference demonstrates that the two orientations of the MC
cross-link can exert differential effects on protein-DNA interactions. The modest inhibition
of AluI cleavage relative to that by CC-1065 [Hurley,L.H., Needham-VanDevanter,D.R., and
Lee,C.(1987)Proc. Natl. Acad. Sci. U.S.A. 84, 6412-6416] may indicate relatively low
distortion of DNA by the MC cross-link. This notion is supported by a comparison with psoralen
cross-links.

<>

<1>Kumar, S., Kaur, C., Kimura, K., Takeo, M., Raghava, G.P., Mayilraj, S.
<2>Draft Genome Sequence of the Type Species of the Genus Citrobacter, Citrobacter freundii MTCC 1658.
<3>Genome Announcements
<4>1
<5>e00120-12
<6>2013
<7>We report the 5.0-Mb genome sequence of the type species of the genus Citrobacter, Citrobacter
freundii strain MTCC 1658, isolated from canal water. This draft genome sequence of C.
freundii strain MTCC 1658(T) consists of 5,001,265 bp with a G+C content of 51.61%, 4,691
protein-coding genes, 70 tRNAs, and 10 rRNAs.

<>

<1>Kumar, S., Kaur, N., Singh, N.K., Raghava, G.P., Mayilraj, S.
<2>Draft Genome Sequence of Streptomyces gancidicus Strain BKS 13-15.
<3>Genome Announcements
<4>1
<5>e00150-13
<6>2013
<7>We report the 7.3-Mbp genome sequence of Streptomyces gancidicus strain BKS 13-15, isolated
from mangrove sediment samples collected from the Bhitar Kanika
Mangrove Reserve Forest, Odissha, India. The draft genome of strain Streptomyces
gancidicus strain BKS 13-15 consists of 7,300,479 bp with 72.6% G+C content,
6,631 protein-coding genes, and 71 RNAs.

<>

<1>Kumar, S., Lindquist, I.E., Sundararajan, A., Rajanna, C., Floyd, J.T., Smith, K.P., Andersen, J.L., He, G., Ayers, R.M., Johnson, J.A., Werdann, J.J., Sandoval, A.A., Mojica, N.M., Schilkey, F.D., Mudge, J., Varela, M.F.
<2>Genome Sequence of Non-O1 Vibrio cholerae PS15.
<3>Genome Announcements
<4>1
<5>e00227-12
<6>2013
<7>The draft genome sequence of a non-O1 Vibrio cholerae strain, PS15, organized into 3,512 open
reading frames within a 3.9-Mb genome, was determined. The PS15
genome sequence will allow for the study of the evolution of virulence and
environmental adaptation in V. cholerae.

<>

<1>Kumar, S., Subramanian, S., Raghava, G.P., Pinnaka, A.K.
<2>Genome Sequence of the Marine Bacterium Marinilabilia salmonicolor JCM 21150T.
<3>J. Bacteriol.
<4>194
<5>3746
<6>2012
<7>We report the 4.98-Mb genome sequence of Marinilabilia salmonicolor JCM 21150(T), which was
isolated from marine mud in the year 1961. The draft genome of strain
Marinilabilia salmonicolor JCM 21150(T) contains 4,982,627 bp with a G+C content
of 41.92% and 4,227 protein coding genes, 52 tRNAs, and 3 rRNAs.

<>

<1>Kumar, S., Vikram, S., Raghava, G.P.
<2>Genome Sequence of the Nitroaromatic Compound-Degrading Bacterium Burkholderia sp. Strain SJ98.
<3>J. Bacteriol.
<4>194
<5>3286
<6>2012
<7>We report the 7.85-Mb genome sequence of Burkholderia sp. strain SJ98, isolated from
agricultural fields of Assam, India. The draft genome of this strain will be
helpful in studying the genetic pathways involved in the degradation of aromatic
compounds.

<>

<1>Kumar, S., Vikram, S., Subramanian, S., Raghava, G.P., Pinnaka, A.K.
<2>Genome Sequence of the Halotolerant Bacterium Imtechella halotolerans K1T.
<3>J. Bacteriol.
<4>194
<5>3731
<6>2012
<7>We report the 3.087-Mb genome sequence of Imtechella halotolerans K1(T), isolated from an
estuarine water sample collected from Kochi, Kerala, India. Strain K1 was
recently reported as a novel genus of the family Flavobacteriaceae.

<>

<1>Kumar-Pinnaka, P.A., Ara, S., Singh, A., Shivaji, S.
<2>Draft Genome Sequence of Winogradskyella psychrotolerans RS-3T, Isolated from the Marine Transect of Kongsfjorden, Ny-Alesund, Svalbard, Arctic Ocean.
<3>Genome Announcements
<4>1
<5>e00630-13
<6>2013
<7>The 4.3-Mb genome of Winogradskyella psychrotolerans strain RS-3(T), isolated from a sediment
sample of a marine transect of Kongsfjorden, Ny-Alesund, Svalbard, Arctic Ocean, is reported.

<>

<1>Kumar-Pinnaka, P.A., Singh, A., Ara, S., Begum, Z., Reddy, G.S., Shivaji, S.
<2>Draft Genome Sequence of Leifsonia rubra Strain CMS 76RT, Isolated from a Cyanobacterial Mat Sample from a Pond in Wright Valley, McMurdo, Antarctica.
<3>Genome Announcements
<4>1
<5>e00633-13
<6>2013
<7>The 2.7-Mb draft genome sequence of Leifsonia rubra strain CMS 76R(T), isolated from a
cyanobacterial mat sample from a pond in Wright Valley, McMurdo, Antarctica, is reported.

<>

<1>Kunert, N.
<2>Identification and characterization of IPOD, a novel interaction partner of the DNA methyltransferase Dnmt2 from Drosophila melanogaster.
<3>Ph.D. Thesis, Germany
<4>
<5>
<6>2005
<7>
<>

<1>Kunkel, L.M., Silberklang, M., McCarthy, B.J.
<2>A third restriction endonuclease from Xanthomonas malvacearum.
<3>J. Mol. Biol.
<4>132
<5>133-139
<6>1979
<7>An additional sequence-specific endonuclease, XmaIII, has been partially
purified from Xanthomonas malvacearum.  XmaIII recognizes ten cleavage sites in
adenovirus 2 DNA, two sites in bacteriophage lambda and no site in either
simian virus 40 DNA or PhiX174 DNA.  It recognizes the sequence 5' C-^G-G-C-C-G
3' 3' G-C-C-G-G^-C 5' and cleaves at the sites indicated by the arrows.  No
other endonuclease with this particular nucleotide sequence specificity has
been reported.

<>

<1>Kunne, C., Billion, A., Mshana, S.E., Schmiedel, J., Domann, E., Hossain, H., Hain, T., Imirzalioglu, C., Chakraborty, T.
<2>Complete Sequences of Plasmids from the Hemolytic-Uremic Syndrome-Associated Escherichia coli Strain HUSEC41.
<3>J. Bacteriol.
<4>194
<5>532-533
<6>2012
<7>The complete and annotated sequences of four plasmids from a historical enteroaggregative
Shiga toxin-producing Escherichia coli (HUSEC) serotype
O104:H4 strain, HUSEC41/01-09591, isolated in 2001 in Germany are
reported.

<>

<1>Kunsch, C., Dillon, P., Barash, S.
<2>Enterococcus faecalis polynucleotides and polypeptides.
<3>Korean Patent Office
<4>KR 1019997010172 A
<5>
<6>1999
<7>
<>

<1>Kunsch, C.A., Choi, G.A., Barash, S.C., Dillon, P.J., Fannon, M.R., Rosen, C.A.
<2>Staphylococcus aureus polynucleotides and sequences.
<3>US Patent Office
<4>US 6737248 A
<5>
<6>2004
<7>
<>

<1>Kunsch, C.A., Choi, G.H., Barash, S., Dillon, P.J., Fannon, M.R., Rosen, C.A.
<2>Staphylococcus aureus polynucleotides and sequences.
<3>US Patent Office
<4>US 6593114 A
<5>
<6>2003
<7>The present invention provides polynucleotide sequences of the genome of Staphylococcus
aureus, polypeptide sequences encoded by the polynucleotide sequences, corresponding
polynucleotides and polypeptides, vectors and hosts comprising the polynucleotides, and assays
and other uses thereof.  The present invention further provides polynucleotide and polypeptide
sequence information stored on computer readable media, and computer-based systems and methods
which facilitate its use.

<>

<1>Kunsch, C.A., Choi, G.H., Dillon, P.J., Rosen, C.A., Barash, S.C., Fannon, M., Dougherty, B.A.
<2>Streptococcus pneumoniae polynucleotides and sequences.
<3>European Patent Office
<4>EP 1400592 A
<5>
<6>2004
<7>The present invention provides polynucleotide sequences of the genome of Streptococcus
pneumoniae, polypeptide sequences encoded by the polynucleotide sequences, corresponding
polynucleotides and polypeptides, vectors and hosts comprising the polynucleotides, and assays
and other uses thereof.  The present invention further provides polynucleotide and polypeptide
sequence information stored on computer readable media, and computer-based systems and methods
which facilitate its use.

<>

<1>Kunsch, C.A., Choi, G.H., Dillon, P.J., Rosen, C.A., Barash, S.C., Fannon, M.R., Dougherty, B.A.
<2>Streptococcus pneumoniae polynucleotides and sequences.
<3>US Patent Office
<4>US 7141418 A
<5>
<6>2006
<7>The present invention provides polynucleotide sequences of the genome of Streptococcus
pneumoniae, polypeptide sequences encoded by the polynucleotide sequences, corresponding
polynucleotides and polypeptides, vectors and hosts comprising the polynucleotides, and assays
and other uses thereof.  The present invention further provides polynucleotide and polypeptide
sequence information stored on computer readable media, and computer-based systems and methods
which facilitate its use.

<>

<1>Kunsch, C.A., Choi, G.H., Dillon, P.S., Rosen, C.A., Barash, S.C., Fannon, M.R., Dougherty, B.A.
<2>Streptococcus pneumoniae polynucleotides and sequences.
<3>US Patent Office
<4>US 6420135 A
<5>
<6>2002
<7>The present invention provides polynucleotide sequences of the genome of Streptococcus
pneumoniae, polypeptide sequences encoded by the polynucleotide sequences, corresponding
polynucleotides and polypeptides, vectors and hosts comprising the polynucleotides, and assays
and other uses thereof.  The present invention further provides polynucleotide and polypeptide
sequence information stored on computer readable media, and computer-based systems and methods
which facilitate its use.

<>

<1>Kunsch, C.A., Dillon, P.J., Barash, S.C.
<2>Enterococcus faecialis polynucleotides and polypeptides.
<3>Japanese Patent Office
<4>JP 2002529046 A
<5>
<6>2002
<7>
<>

<1>Kunst, F. et al.
<2>The complete genome sequence of the Gram-positive bacterium Bacillus subtilis.
<3>Nature
<4>390
<5>249-256
<6>1997
<7>Bacillus subtilis is the best-characterized member of the Gram-positive bacteria.  Its genome
of 4,214,810 base pairs comprises 4,100 protein-coding genes.  Of these protein-coding genes,
53% are represented once, while a quarter of the genome corresponds to several gene families
that have been greatly expanded by gene duplication, the largest family containing 77 putative
ATP-binding transport proeins.  In addition, a large proportion of the genetic capacity is
devoted to the utilization of a variety of carbon sources, including many plant-derived
molecules.  The identification of five signal peptidase genes, as well as several genes for
components of the secretion apparatus, is important given the capacity of Bacillus strains to
secrete large amounts of industrially important enzymes.  Many of the genes are involved in
the synthesis of secondary metabolites, including antibiotics, that are more typically
associated with Streptomyces species.  The genome contains at least ten prophages or remnants
of prophages, indicating that bacteriophage infection has played an important evolutionary
role in horizontal gene transfer, in particular in the propagation of bacterial pathogenesis.

<>

<1>Kunst, F., Glaser, P.
<2>Listeria inocua, genome and applications.
<3>International Patent Office
<4>WO 0228891 A
<5>
<6>2000
<7>The invention concerns a nucleotide sequence derived from Listeria inocua corresponding to a
sequence selected among SEQ ID NO: 1 to SEQ ID NO: 11 and the comparative analysis of said
genome with that of Listeria monocytogenes.

<>

<1>Kunz, A., Mackeldanz, P., Mucke, M., Meisel, A., Reuter, M., Schroeder, C., Kruger, D.H.
<2>Mutual activation of two restriction endonucleases: interaction of EcoPI and EcoP15.
<3>Biol. Chem.
<4>379
<5>617-620
<6>1998
<7>Type III restriction endonucleases recognize nonsymmetric nucleotide sequences.  A necessary
condition for DNA cleavage is the presence of two unmethylated recognition sites which are
inversely ('head-to-head') oriented in the DNA double strand.  A DNA substrate possessing
one EcoP1 and one EcoP15 site in the head-to-head configuration could not be cleaved by the
individual enzymes, however, it was specifically digested in the simultaneous presence of both
enzymes.  In agreement with the tracking-collision model for the DNA interaction of type III
enzymes cleavage could be abolished by Lac repressor bound between the two sites.  We conclude
that two different type III enzymes can functionally cooperate in the cleavage of DNA.

<>

<1>Kunz, A., Meisel, A., Mackeldanz, P., Reuter, M., Kruger, D.H.
<2>An experimental selection system to identify bacterial cells exhibiting a new DNA host specificity.
<3>Biol. Chem.
<4>379
<5>563-566
<6>1998
<7>Restriction-modification enzymes interact with DNA sequences in a highly specific manner.
Mutations within the DNA binding region of the enzymes could be expected to produce enzyme
variants with changed DNA sequence specificities.  We developed an efficient in vivo selection
system that enabled us to detect one cell coding for a restriction-modification system with a
new DNA sequence specificity in a background of more than 10^6 cells with the original DNA
sequence specificity.

<>

<1>Kuosmanen, M., Poso, H.
<2>Inhibition of the activity of restriction endonucleases by spermidine and spermine.
<3>FEBS Lett.
<4>179
<5>17-20
<6>1985
<7>Physiological concentrations (0.5-2.0 mM) of spermidine and spermine were
observed to inhibit the digestion in vitro of plasmid pJDB 207 by the
restriction endonucleases BamHI (EC 3.1.23.6), EcoRI (EC3.2.23.13), HindIII
(EC3.1.23.20), HpaI (EC3.1.23.23) and PstI (EC3.1.23.31).  The polyamines
protected all the tested restriction sequences of DNA, since the activity of
all endonucleases used was strongly inhibited.  These results show the need for
caution when using polyamines as experimental tools for recombinant DNA
chemistry.

<>

<1>Kupper, D., Moncke-Buchner, E., Reuter, M., Kruger, D.H.
<2>Oligonucleotide stimulators allow complete cleavage of agarose-embedded DNA by particular type II restriction endonucleases.
<3>Anal. Biochem.
<4>272
<5>275-277
<6>1999
<7>In previous work we and others discovered several restriction endonucleases requiring the
simultaneous interaction with two copies of their respective recognition sequence for enzyme
activity.  EcoRII was the first restriction enzyme for which this special property has been
described, and the mechanism underlying the cooperative interaction with two sites in the
substrate DNA was studied in more detail.  Complete cleavage of a resistant DNA site (present
alone or at a great distance to a second site in a DNA molecule) can be achieved by providing
the required second copy of the recognition site on short synthetic oligonucleotide duplexes.
This principle has been verified for at least 12 restriction endonucleases so far (e.g.,
EcoRII, HpaII, NaeI, NarI, SfiI, and others).  In addition, the practical importance of
restriction enzyme stimulation by specific oligo duplexes for the reliable detection of
prokaryotic Dcm methylation by comparative DNA digestion with EcoRII/BstNI and of eukaryotic
CpG methylation with HpaII/MspI has been demonstrated.  Now we wanted to verify that the
technique of oligo duplex-based stimulation of those restriction endonucleases is also
applicable for digesting DNA being embedded in agarose as necessary for the handling of larger
genomic DNA.

<>

<1>Kupper, D., Reuter, M., Kruger, D.H.
<2>Overproduction of His-tagged EcoRII restriction endonuclease and terminally deleted mutant proteins.
<3>Gene
<4>157
<5>97-98
<6>1995
<7>EcoRII was the first restriction endonuclease (ENase) reported requiring the cooperative
interaction with at least two DNA sites for activity.  Using two different expression systems
the enzyme could be purified and its special substrate requirements were further analyzed.  At
the present state of knowledge we suggest a model of simultaneous binding of two DNA sites to
one dimeric enzyme molecule.

<>

<1>Kupper, D., Reuter, M., Mackeldanz, P., Meisel, A., Alves, J., Schroeder, C., Kruger, D.H.
<2>Hyperexpressed EcoRII renatured from inclusion bodies and native enzyme both exhibit essential cooperativity with two DNA sites.
<3>Protein Expr. Purif.
<4>6
<5>1-9
<6>1995
<7>EcoRII was the first restriction endonuclease (ENase) reported needing the cooperative
interaction with at least two DNA sites for activity. We constructed an EcoRII-overproducing
strain of Escherichia coli by placing the coding sequence under control of the T7 gene 10
regulatory elements. The yield of EcoRII expression could be increased to about 10% of total
soluble cellular protein. Inclusion bodies are formed that mainly consist of insoluble EcoRII
molecules. After solubilization by 6M guanidine hydrochloride refolding of the enzyme was
achieved by dilution into appropriate buffer. The endonuclease was purified to homogeneity
from both the soluble protein fraction and the protein renatured from inclusion bodies. Their
identity was proven by circular dichroism and analysis of enzyme activity with respect to the
special substrate requirements of EcoRII. It is shown that EcoRII cleavage of
oligodeoxyribonucleotide duplexes (oligo duplexes) with only one recognition site follows a
sigmoidal concentration dependence, i.e., they cannot be cleaved below a distinct low DNA
concentration where simultaneous interaction with two substrate molecules is no longer
possible. We demonstrate that the restriction of oligo duplexes containing two recognition
sites does not show this concentration dependence, confirming an intramolecular site
cooperativity.

<>

<1>Kupper, D., Reuter, M., Meisel, A., Kruger, D.H.
<2>Reliable detection of DNA CpG methylation profiles by the isoschizomers MspI/HpaII using oligonucleotide stimulators.
<3>Biotechniques
<4>23
<5>843-846
<6>1997
<7>The increasing interest in methylation patterns of mammalian DNA demands a simple and reliable
method to clearly differentiate between cytosine and 5-methylcytosine within a specific DNA
region.  The simplest and most commonly used approach is the analysis with restriction
endonucleases exhibiting different methylation sensitivities, like the MspI/HpaII pair of
isoschizomeric enzymes.  Whereas MspI cleaves the recognition sequence 5'-CCGG independently
of the methylation state of the internal C, cleavage by HpaII is blocked by the presence of
5-MeC at this site.  The method is compromised by the fact that methylation can be detected
only in those CpGs that are located within the tetranucleotide recognition sequence.

<>

<1>Kupper, D., Zhou, J.-G., Kiss, A., Venetianer, P.
<2>Cloning and structure of the M.BepI modification methyltransferase.
<3>Gene
<4>74
<5>33
<6>1988
<7>Meeting abstract

<>

<1>Kupper, D., Zhou, J.-G., Venetianer, P., Kiss, A.
<2>Cloning and structure of the BepI modification methylase.
<3>Nucleic Acids Res.
<4>17
<5>1077-1088
<6>1989
<7>The gene coding for a CGCG specific DNA methylase has been cloned in E. coli from
Brevibacterium epidermidis.  The enzyme, named BepI methylase, is probably the cognate
methylase of the FnuDII isoschizomer BepI endonuclease isolated from this strain.  The
expression of BepI methylase in E. coli is dependent on the orientation of the cloned fragment
suggesting that the gene is transcribed from a promoter on the plasmid vector.  No BepI
endonuclease could be detected in the clones producing BepI methylase.  The nucleotide
sequence of the BepI methylase gene has been determined, it predicts a protein of 403 amino
acids (M:45,447).  Analysis of the amino acid sequence deduced from the nucleotide sequence
revealed similarities between the BepI methylase and other cytosine methylases.  M.BepI
methylates the external cytosine in its recognition sequence.

<>

<1>Kupriyanova, N.S., Kirilenko, P.M., Netchvolodov, K.K., Ryskov, A.P.
<2>Preferential cleavage sites for Sau3A restriction endonuclease in human ribosomal DNA.
<3>Biochem. Biophys. Res. Commun.
<4>274
<5>11-15
<6>2000
<7>Previous studies of cloned ribosomal DNA (rDNA) variants isolated from the cosmid library of
human chromosome 13 have revealed some disproportion in representativity of different rDNA
regions (N. S. Kupriyanova, K. K. Netchvolodov, P. M. Kirilenko, B. I. Kapanadze, N. K.
Yankovsky, and A. P. Ryskov, Mol. Biol. 30, 51-60, 1996). Here we show nonrandom cleavage of
human rDNA with Sau3A or its isoschizomer MboI under mild hydrolysis conditions. The
hypersensitive cleavage sites were found to be located in the ribosomal intergenic spacer
(rIGS), especially in the regions of about 5-5.5 and 11 kb upstream of the rRNA transcription
start point. This finding is based on sequence mapping of the rDNA insert ends in randomly
selected cosmid clones of human chromosome 13 and on the data of digestion kinetics of cloned
and noncloned human genomic rDNA with Sau3A and MboI. The results show that the methylation
status and superhelicity state of the rIGS have no effect on cleavage site sensitivity. It is
interesting that all primary cleavage sites are adjacent to or entering into Alu or Psi cdc 27
retroposons of the rIGS suggesting a possible role of neighboring sequences in nuclease
accessibility. The results explain nonequal representation of rDNA sequences in the human
genomic DNA library used for this study.

<>

<1>Kur, J.
<2>Integration host factor (IHF) applied for partial digestion by restriction endonucleases in large DNA molecules (IARC procedure).
<3>Acta Microbiol. Pol.
<4>42
<5>145-150
<6>1993
<7>The IHF protein of Escherichia coli was successfully used in IHF-mediated Achilles' Heel
Cleavage (IHF-AC) technique and leads to the generation of very rare restriction sites in
large DNA molecules. The first step of this procedure is methylation of DNA in the presence of
IHF, when the overlapping ihf/restriction sites are protected from methylation, and in the
second step the DNA is cut by the cognate restriction enzyme. The aim of the present study is
to develop a very exact and reproducible procedure to obtain only a few well-defined cuts with
the IHF-pre-treated DNA, depending on the variety of all parameters. This technique (IARC,
i.e., IHF-assisted rare cutters) employs the restriction enzyme and only one auxiliary protein
(IHF). The advantage of the IARC procedure is that no methylation is required (as opposed to
the IHF-AC method). Using the IARC approach, the effects of various IHF concentrations were
evaluated on the cleavage activity of the DraI, PacI, PmeI, and SwaI enzymes using DNA of
phage lambda or the entire genomic 4.7-Mb DNA of E. coli. At low IHF concentrations only a few
cut sites were eliminated by IHF binding, but a high IHF concentration, enzymes were able to
cut in only one or several specific sites.

<>

<1>Kur, J.
<2>Use of IHF mediated achilles' heel cleavage (IHF-AC) method for mapping ihf sites.
<3>Acta Microbiol. Pol.
<4>42
<5>137-144
<6>1993
<7>We have shown that Integration Host Factor of E. coli can successfully be used in the
IHF-mediated Achilles' Heel Cleavage (IHF-AC), for generating rare natural cleavage sites.
The first step of this procedure is methylation of DNA in the presence of IHF, when the
overlapping ihf/restriction sites are protected from methylation, and in the second step the
DNA is cut by the cognate restriction enzyme. The extent of cleavage could be controlled by
varying the IHF:DNA ratio and temperature. The aim of the present study is to demonstrate that
IHF-AC procedure might serve as a useful tool for finding new protein-binding sites which
overlap known restriction sites. I have used this approach in conjunction with several MTases
to find several other unknown IHF-binding sites.

<>

<1>Kur, J., Koob, M., Burkiewicz, A., Szybalski, W.
<2>A novel method for converting common restriction enzymes into rare cutters:  integration host factor-mediated Achilles' cleavage (IHF-AC).
<3>Gene
<4>110
<5>1-7
<6>1992
<7>Integration host factor (IHF)-mediated protection against enzymatic methylation at
ihf-overlapping sites provides the basis for this novel application of the Achilles' cleavage
(AC) technique [Koob et al., Science 241 (1988) 1084-1086] for generating rare natural
cleavage sites. When applying IHF-AC to plasmid, phage lambda, Escherichia coli and yeast
genomes, only a few of the EcoRI, HinfI, and MboI sites (which overlapped the ihf sites)
remained cleavable after prior methylation with the cognate M.EcoRI, M.HinfI, or Dam
methyltransferases in the presence of IHF. Thus, IHF-AC essentially converted these enzymes
into very rare cutters. The extent of cleavage could be controlled by varying the IHF:DNA
ratio and temperature. Moreover, the method permits the genomic location and strength of the
ihf sites to be determined.

<>

<1>Kurakin, A.V., Zaritskaya, L.S., Metliskaya, A.Z., Volodin, A.A., Cherny, D.I.
<2>BspRI methyltransferase as a marker for electron microscopic physical mapping.
<3>Micron and Microscopica Acta
<4>22
<5>213-221
<6>1991
<7>An approach is described in this paper for direct physical mapping of DNA by
electron microscopy.  It implies visualization of specific
DNA-methyltransferase complexes followed by computer analysis of electron
micrographs.  The BspRI methylase (recognition site GGCC) was used as a marker
owing to the large difference (at least three orders of magnitude) between its
specific and non-specific interaction with DNA, as revealed by the gel
retardation technique.  For electron microscopic mapping the optimum conditions
were established in order to produce the maps practically without non-specific
noise.  The approach was tested with well-characterized plasmid DNAs-pA03,
pUC19 and pBR322 carrying 4,11 and 22 GGCC sites respectively. The results were
analyzed and the applications of the method are discussed.

<>

<1>Kurata, A., Hirose, Y., Misawa, N., Hurunaka, K., Kishimoto, N.
<2>Draft Genome Sequence of the Ionic Liquid-Tolerant Bacterium Bacillus amyloliquefaciens CMW1.
<3>Genome Announcements
<4>2
<5>e01051-14
<6>2014
<7>Here, we report the draft genome sequence of an ionic liquid-tolerant bacterium,  Bacillus
amyloliquefaciens CMW1, which is newly isolated from a Japanese
fermented soybean paste. The genome sequence will allow for a characterization of
the molecular mechanism of its ionic liquid tolerance.

<>

<1>Kurata, A., Nishimura, M., Kishimoto, N., Kobayashi, T.
<2>Draft Genome Sequence of a Deep-Sea Bacterium, Bacillus niacini Strain JAM F8, Involved in the Degradation of Glycosaminoglycans.
<3>Genome Announcements
<4>2
<5>e00983-14
<6>2014
<7>Here, we report the draft genome sequence of Bacillus niacini JAM F8, which was newly isolated
from deep-sea sediment at a depth of 2,759 m from the
Izu-Ogasawara Trench. An array of genes related to degradation of
glycosaminoglycans in this bacterium was identified by whole-genome analysis.

<>

<1>Kurita, J., Nakajima, K.
<2>DNA encoding virus-neutralization and phylaxis-related protein of red sea bream iridovirus.
<3>Japanese Patent Office
<4>JP 2002101885 A
<5>
<6>2002
<7>
<>

<1>Kuroda, M. et al.
<2>Whole genome sequencing of meticillin-resistant Staphylococcus aureus.
<3>Lancet
<4>357
<5>1225-1240
<6>2001
<7>Staphylococcus aureus is one of the major causes of community-acquired and hospital-acquired
infections.  It produces numerous toxins including superantigens that cause unique disease
entities such as toxic-shock syndrome and staphylococcal scarlet fever, and has acquired
resistance to practically all antibiotics.  Whole genome analysis is a necessary step towards
future development of countermeasures against this organism.

<>

<1>Kuroda, M., Yamashita, A., Hirakawa, H., Kumano, M., Morikawa, K., Higashide, M., Maruyama, A., Inose, Y., Matoba, K., Toh, H., Kuhara, S., Hattori, M., Ohta, T.
<2>Whole genome sequence of Staphylococcus saprophyticus reveals the pathogenesis of uncomplicated urinary tract infection.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>13272-13277
<6>2005
<7>Staphylococcus saprophyticus is a uropathogenic Staphylococcus frequently isolated from young
female outpatients presenting with uncomplicated
urinary tract infections. We sequenced the whole genome of S.
saprophyticus type strain ATCC 15305, which harbors a circular chromosome
of 2,516,575 bp with 2,446 ORFs and two plasmids. Comparative genomic
analyses with the strains of two other species, Staphylococcus aureus and
Staphylococcus epidermidis, as well as experimental data, revealed the
following characteristics of the S. saprophyticus genome. S. saprophyticus
does not possess any virulence factors found in S. aureus, such as
coagulase, enterotoxins, exoenzymes, and extracellular matrix-binding
proteins, although it does have a remarkable paralog expansion of
transport systems related to highly variable ion contents in the urinary
environment. A further unique feature is that only a single ORF is
predictable as a cell wall-anchored protein, and it shows positive
hemagglutination and adherence to human bladder cell associated with
initial colonization in the urinary tract. It also shows significantly
high urease activity in S. saprophyticus. The uropathogenicity of S.
saprophyticus can be attributed to its genome that is needed for its
survival in the human urinary tract by means of novel cell wall-anchored
adhesin and redundant uro-adaptive transport systems, together with
urease.

<>

<1>Kuroita, T., Inoue, H., Kobayashi, K., Ishikawa, K., Watanabe, M.
<2>Novel type II restriction enzyme PabI, useful in gene recombination, genetic engineering such as gene analysis, gene modification technology, and for elucidating hereditary disease - recombinant restriction enzyme production via plasmid expression.
<3>Japanese Patent Office
<4>JP 2006042642
<5>
<6>2006
<7>
<>

<1>Kurokawa, S., Bessho, Y., Higashijima, K., Shirouzu, M., Yokoyama, S., Watanabe, K.I., Ohama, T.
<2>Adaptation of intronic homing endonuclease for successful horizontal transmission.
<3>FEBS J.
<4>272
<5>2487-2496
<6>2005
<7>Group I introns are thought to be self-propagating mobile elements, and are distributed over a
wide range of organisms through horizontal
transmission. Intron invasion is initiated through cleavage of a target
DNA by a homing endonuclease encoded in an open reading frame (ORF)
found within the intron. The intron is likely of no benefit to the host
cell and is not maintained over time, leading to the accumulation of
mutations after intron invasion. Therefore, regular invasional
transmission of the intron to a new species at least once before its
degeneration is likely essential for its evolutionary long-term
existence. In many cases, the target is in a protein-coding region
which is well conserved among organisms, but contains ambiguity at the
third nucleotide position of the codon. Consequently, the homing
endonuclease might be adapted to overcome sequence polymorphisms at the
target site. To address whether codon degeneracy affects horizontal
transmission, we investigated the recognition properties of a homing
enzyme, I-CsmI, that is encoded in the intronic ORF of a group I intron
located in the mitochondrial COB gene of the unicellular green alga
Chlamydomonas smithii. We successfully expressed and purified three
types of N-terminally truncated I-CsmI polypeptides, and assayed the
efficiency of cleavage for 81 substrates containing single nucleotide
substitutions. We found a slight but significant tendency that I-CsmI
cleaves substrates containing a silent or tolerated amino acid change
more efficiently than nonsilent or nontolerated ones. The published
recognition properties of I-SpomI, I-ScaI, and I-SceII were
reconsidered from this point of view, and we detected proficient
adaptation of I-SpomI, I-ScaI, and I-SceII for target site sequence
degeneracy. Based on the results described above, we propose that
intronic homing enzymes are adapted to cleave sequences that might
appear at the target region in various species, however, such
adaptation becomes less prominent in proportion to the time elapsed
after intron invasion into a new host.

<>

<1>Kurokawa, S., Kabayama, J., Nho, S.W., Hwang, S.D., Hikima, J., Jung, T.S., Kondo, H., Hirono, I., Takeyama, H., Aoki, T.
<2>Whole-Genome Sequence of Fish-Pathogenic Mycobacterium sp. Strain 012931, Isolated from Yellowtail (Seriola quinqueradiata).
<3>Genome Announcements
<4>1
<5>e00534-13
<6>2013
<7>The genus Mycobacterium comprises a large number of well-characterized species, several of
which are human and animal pathogens. Here, we report the whole-genome
sequence of Mycobacterium sp. strain 012931, a fish pathogen responsible for huge
losses in aquaculture farms in Japan. The strain was isolated from a marine fish,
yellowtail (Seriola quinqueradiata).

<>

<1>Kuroyedov, A.A., Grokhovsky, S.L., Zhuze, A.L., Nosikov, V.V., Polyanovsky, O.L.
<2>Distamycin A and its analogs as agents for blocking of endo R. EcoRI activity.
<3>Gene
<4>1
<5>389-395
<6>1977
<7>Distamycin A (Dst) and its analogs protect the lambda phage DNA from cleavage
with endoR. EcoRI and show selective affinity for different recognition sites
of endoR. EcoRI on this DNA producing enlarged DNA fragments of various
composition and length.  The affinity of the antibiotic for DNA is influenced
by the number of pyrrol carboxamide units in Dst molecule and does not strongly
depend on the substitution of the N-methyl group by the N-propyl one.  Since in
the complex with DNA the antibiotics of the Dst type are localized in its minor
groove a conclusion can be made that the minor groove of DNA is needed for the
interaction of the restriction endonuclease with DNA.

<>

<1>Kurpiewski, M.R., Engler, L.E., Wozniak, L.A., Kobylanska, A., Koziolkiewicz, M., Stec, W.J., Jen-Jacobson, L.
<2>Mechanisms of Coupling between DNA Recognition Specificity and Catalysis in EcoRI Endonuclease.
<3>Structure
<4>12
<5>1775-1788
<6>2004
<7>Proteins that bind to specific sites on DNA often do so in order to carry out catalysis or
specific protein-protein interaction while bound to the
recognition site. Functional specificity is enhanced if this second
function is coupled to correct DNA site recognition. To analyze the
structural and energetic basis of coupling between recognition and
catalysis in EcoRI endonuclease, we have studied stereospecific
phosphorothioate (P(S)) or methylphosphonate (P(Me)) substitutions at the
scissile phosphate GpAATTC or at the adjacent phosphate GApATTC in
combination with molecular-dynamics simulations of the catalytic center
with bound Mg(2+). The results show the roles in catalysis of individual
phosphoryl oxygens and of DNA distortion and suggest that a "crosstalk
ring" in the complex couples recognition to catalysis and couples the two
catalytic sites to each other.

<>

<1>Kurpiewski, M.R., Koziolkiewicz, M., Wilk, A., Stec, W.J., Jen-Jacobson, L.
<2>Chiral phosphorothioates as probes of protein interactions with individual DNA phosphoryl oxygens:  Essential interactions of EcoRI endonuclease with the phosphate at pGAATTC.
<3>Biochemistry
<4>35
<5>8846-8854
<6>1996
<7>The contact between EcoRI endonuclease and the "primary clamp" phosphate of its recognition
site pGAATTC is absolutely required for recognition of the canonical and all variant DNA
sites.  We have probed this contact using oligonucleotides containing the single
stereospecific (Rp)- or (Sp)- phosphorothioates.  At the GAApTTC position, where the
endonuclease interacts with only one phosphoryl oxygen at the central DNA kink, Rp-Ps inhibits
and Sp-Ps stimulates binding and cleavage; in contrast, at the pGAATTC position both
diastereomers inhibit binding.  For single-strand substitution, the penalty in binding free
energy (delta delta Go bind) is slightly greater for Sp-Ps (+0.9 kcal/mol) than for Rp-Ps
(+0.7 kcal/mol).  Binding penalties are approximately additive for double-strand substitution
(Rp,Rp-Ps or Sp,Sp-Ps).  Neither Ps diastereomer in one DNA strand affects the first-order
rate constants for cleavage in the unmodified DNA strand, and only Sp-Ps inhibits the cleavage
rate constant (3-fold) in the modified DNA strand.  Thus, the second-order cleavage rate
(including binding and catalysis) is inhibited 14-fold by Sp-Ps and 45-fold by Sp,Sp-Ps.  In
the canonical complex, the phosphate at pGAATTC is completely surrounded by protein and each
nonbridging phosphoryl oxygen receives two hydrogen bonds from the endonuclease, such that in
either orientation the increased bond length of P-S- inhibits binding.  However, the pro-Sp
oxygen interacts with residues that are connected (by proximity or inter-side-chain hydrogen
bonding) to side chains with essential roles in catalysis, so cleavage is preferentially
inhibited when these side chains are slightly displaced by the Sp-Ps diastereomer.

<>

<1>Kusano, K., Asami, Y., Fujita, A., Tanokura, M., Kobayashi, I.
<2>Type I restriction enzyme with RecA protein promotes illegitimate recombination.
<3>Plasmid
<4>50
<5>202-212
<6>2003
<7>Illegitimate (non-homologous) recombination requires little or no sequence homology between
recombining DNAs and has been regarded as being a process
distinct from homologous recombination, which requires a long stretch of
homology between recombining DNAs. However, we have found a type of
illegitimate recombination that requires an interaction between long
homologous DNA sequences. It was detected when a plasmid that carried
2-kb-long inverted repeats was subjected to type I (EcoKI) restriction in
vivo within a special mutant strain of Escherichia coli. In the present
work, we analyzed genetic requirements for this type of illegitimate
recombination in well-defined genetic backgrounds. Our analysis
demonstrated dependence on RecA function and on the presence of two EcoKI
sites on the substrate DNA. These results are in harmony with a model in
which EcoKI restriction enzyme attacks an intermediate of homologous
recombination to divert it to illegitimate recombination.

<>

<1>Kusano, K., Naito, T., Handa, N., Kobayashi, I.
<2>Restriction-modification systems as genomic parasites in competition for specific sequences.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>92
<5>11095-11099
<6>1995
<7>Restriction-modification (RM) systems are believed to have evolved to protect cells from
foreign DNA.  However, this hypothesis may not be sufficient to explain the diversity and
specificity in sequence recognition, as well as other properties, of these systems.  We report
that the EcoRI restriction endonuclease-modification methylase (rm) gene pair stabilizes
plasmids that carry it and that this stabilization is blocked by an RM of the same sequence
specificity (EcoRI or its isoschizomer, RsrI) but not by an RM of a different specificity
(PaeR7I) on another plasmid.  The PaeR7I rm likewise stabilizes plasmids, unless an rm gene
pair with identical sequence specificity is present.  Our analysis supports the following
model for stabilization and incompatibility: the descendants of cells that have lost an rm
gene pair expose the recognition sites in their chromosomes to lethal attack by any remaining
restriction enzymes unless modification by another RM system of the same specificity protects
these sites.  Competition for specific sequences among these selfish genes may have generated
the great diversity and specificity in sequence recognition among RM systems.  Such altruistic
suicide strategies, similar to those found in virus-infected cells, may have allowed selfish
RM systems to spread by effectively competing with other selfish genes.

<>

<1>Kusano, K., Sakagami, K., Yokochi, T., Naito, T., Tokinaga, Y., Euda, E., Kobayashi, I.
<2>A new type of illegitimate recombination is dependent on restriction and homologous interaction.
<3>J. Bacteriol.
<4>179
<5>5380-5390
<6>1997
<7>Illegitimate (nonhomologous) recombination requires little or no sequence homology between
recombining DNAs and has been regarded as being a process distinct from homologous
recombination, which requires a long stretch of homology between recombining DNAs.  Under
special conditions in Escherichia coli, we have found a new type of illegitimate recombination
that requires an interaction between homologous DNA sequences.  It was detected when a plasmid
that carried 2-kb-long inverted repeats was subjected to type II restriction in vitro and type
I (EcoKI) restriction in vivo within a delta-rac recBC recG ruvC strain.  Removal of one of
the repeats or its replacement with heterologous DNA resulted in a reduction in the level of
recombination.  The recombining sites themselves shared, at most a few base pairs of homology.
Many of the recombination events joined a site in one of the repeats with a site in another
repeat.  In two of the products, one of the recombining sites was at the end of one of the
repeats.  Removal of one of the EcoKI sites resulted in decreased recombination.  We discuss
the possibility that some structure made by homologous interaction between the long repeats is
used by the EcoKI restriction enzyme to promote illegitimate recombination.  The possible
roles and consequences of this type of homologous interaction are discussed.

<>

<1>Kusiak, M., Price, C., Rice, D., Hornby, D.P.
<2>The HsdS polypeptide of the Type IC restriction enzyme EcoR124 is a sequence-specific DNA-binding protein.
<3>Mol. Microbiol.
<4>6
<5>3251-3256
<6>1992
<7>The HsdS and HsdM polypeptides of the type IC restriction enzyme EcoR124 have been purified
independently and used in a set of gel retardation experiments to determine the minimum
requirements for sequence-specific recognition of DNA by this enzyme. The HsdS polypeptide
alone is able to bind to DNA in a sequence-specific manner. In addition, whilst the presence
of the HsdM polypeptide gives rise to a stimulation of DNA binding by the HsdS subunit it is
not clear whether, under the conditions of the experiments reported here, the HsdS subunit
maintains the same interactions with the HsdM subunits observed in the absence of DNA.

<>

<1>Kuspa, A., Loomis, W.F.
<2>Tagging developmental genes in Dictyostelium by restriction enzyme-mediated integration of plasmid DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>8803-8807
<6>1992
<7>Introduction of restriction enzyme along with linearized plasmid results in integration of
plasmid DNA at genomic restriction sites in a high proportion of the resulting transformants.
We have found that electroporating BamHI or EcoRI together with pyr5-6 plasmids cut with the
same enzyme stimulates the efficiency of transformation in Dictyostelium discoideum more than
20-fold over the rate seen when plasmid DNA alone is introduced. Restriction enzyme-mediated
integration generates insertions into genomic restriction sites in an apparently random
manner, some of which cause mutations. About 1 in 400 of the Dictyostelium transformants
displayed arrested or aberrant development. The integrated plasmid, along with flanking
genomic DNA, was excised from some of these mutants, cloned in Escherichia coli, and used to
transform other Dictyostelium cells. Homologous recombination within the flanking sequences
resulted in the same phenotypes displayed by the original mutants, directly demonstrating that
the affected genes were responsible for the specific morphological phenotypes. This method of
insertional mutagenesis should be useful for tagging, and subsequent cloning, of many
developmentally important genes that can be identified by their mutant phenotypes.

<>

<1>Kusumoto, M., Fukamizu, D., Ogura, Y., Yoshida, E., Yamamoto, F., Iwata, T., Ooka, T., Akiba, M., Hayashi, T.
<2>The lineage-specific distribution of IS-excision enhancer in enterotoxigenic Escherichia coli isolated from swine.
<3>Appl. Environ. Microbiol.
<4>80
<5>1394-1402
<6>2014
<7>Insertion sequences (ISs) are the simplest transposable elements and are widely distributed in
bacteria; however, they also play important roles in genome evolution. We recently identified
a protein called IS-excision enhancer (IEE) in enterohemorrhagic Escherichia coli (EHEC) O157.
IEE promotes the excision of IS elements belonging to the IS3 family, such as IS629, as well
as several other families. IEE-mediated IS excision
generates various genomic deletions that lead to the diversification of the bacterial genome.
IEE has been found in a broad range of bacterial species; however, among sequenced E. coli
strains, IEE is primarily found in EHEC isolates. In this study, we investigated non-EHEC
pathogenic E. coli strains isolated from domestic animals and found that IEE is distributed in
specific lineages of enterotoxigenic E. coli (ETEC) strains of serotypes O139
or O149 isolated from swine. The iee gene is located within integrative elements that are
similar to SpLE1 of EHEC O157. All iee-positive ETEC lineages also contained multiple copies
of IS629, a preferred substrate of IEE, and their genomic locations varied significantly
between strains, as observed in O157. These data suggest that IEE may have been transferred
among EHEC and ETEC in swine via SpLE1 or SpLE1-like integrative
elements. In addition, IS629 is actively moving in the ETEC O139 and O149 genomes, and as in
EHEC O157, is promoting the diversification of these genomes in combination with IEE.

<>

<1>Kutish, G.F., Li, Y., Lu, Z., Furuta, M., Rock, D.L., Van Etten, J.L.
<2>Analysis of 76 kb of the Chlorella virus PBCV-1 330-kb genome: Map positions 182 to 258.
<3>Virology
<4>223
<5>303-317
<6>1996
<7>Analysis of 76 kb of newly sequencing DNA, located between map positions 182 and 258 kb in the
330-kb chlorella virus PBCV-1 genome, revealed 175 open reading frames of 65 codons or longer.
One hundred and five of these 175 ORFs were considered major ORFs.  Twenty-one of the 105
major ORFs resembled proteins in databases including ribonucleotide reductase small subunit,
RNase III, thioredoxin, glutaredoxin, protein disulfide isomerase, deoxynucleoside kinase,
frog virus 3 ATPase, Acetobacter cellulose synthase, a bacteriophage encoded endonuclease, and
two C-5 cytosine DNA methyltransferases.  One of the ORFs was the PBCV-1 major capsid protein.
The 105 major ORFs were evenly distributed along the genome.  One set of ORFs was separated by
543 nucleotides whereas 75 of the ORFs were separated by fewer than 100 nucleotides.  Nineteen
of the 175 ORFs resembled other PBCV-1 ORFs, suggesting that they represent either gene
duplications or gene families.

<>

<1>Kutter, E., Rueger, W.
<2>Structure, organization, and manipulation of the genome.
<3>Bacteriophage T4, American Society for Microbiology, Mathews, C.K., Kutter, E.M., Mosig, G., Berget, P.B., Washington, D.C.
<4>0
<5>277-290
<6>1983
<7>Recent analysis of T4 transcription patterns, gene sequences, origins of replication, and
related data have taken advantage of the extensive restriction mapping of T4 and its
correlation with the genetic map.  Here we summarize the current data, including the locations
of those genes and specific transcripts that have now been mapped to within a few hundred base
pairs.  This is a continuing project; therefore, any additional sequence data, clone analyses,
and promoter mapping data will be greatly appreciated by the authors.  Sequence data should
also be submitted to the T4 sequence bank being maintained by Larry Gold, University of
Colorado, Boulder, Colo. 80309.

<>

<1>Kutter, E., Stidham, T., Guttman, B., Kutter, E., Batts, D., Peterson, S., Javakhishvili, T.D., Arisaka, F., Mesyanzhinov, V., Rueger, W., Mosig, G.
<2>Genomic map of bacteriophage T4.
<3>Molecular Biology of Bacteriophage T4, American Society for Microbiology, Karam, J.D., Drake, J.W., Kreuzer, K.N., Mosig, G., Hall, D.H., Eiserling, F.A., Washington, D.C.
<4>0
<5>491-519
<6>1994
<7>The map presented here is based on the T4 DNA sequence, integrated with information from
genetic analysis.  In general, the results agree well with those initially determined by
restriction analysis and the genetic map of Wood and Revel.

<>

<1>Kutueva, L.I., Ashapkin, V.V., Vanyushin, B.F.
<2>The methylation pattern of a cytosine DNA-methyltransferase gene in Arabidopsis thaliana plants.
<3>Biochem. Mol. Biol. Int.
<4>40
<5>347-353
<6>1996
<7>Using a PCR-amplified 5'-end proximal 600-bp fragment and two cDNA clones of cytosine
DNA-methyltransferase gene of Arabidopsis thaliana as specific probes in hybridization with
plant DNA samples, hydrolyzed by methylation-sensitive restriction endonucleases HpaII and
MspI, it has been established that CCGG sites located in the 5'-end proximal part of cytosine
DNA-methyltransferase gene are highly methylated at internal C and less but detectably
methylated at external C residues.  On the contrary, all CCGG sites in 3'-terminal half of
the coding region were found to be unmethylated at both external and internal C residues.  No
significant differences between methylation patterns of cytosine DNA-methyltransferase gene in
various organs (leaf, stem, flower) of the Arabidopsis thaliana plant were detected.

<>

<1>Kutumbaka, K.K., Han, S., Mategko, J., Nadala, C., Buser, G.L., Cassidy, M.P., Beldavs, Z.G., Weissman, S.J., Morey, K.E., Vega, R., Samadpour, M.
<2>Draft Genome Sequence of blaNDM-1-Positive Escherichia coli O25b-ST131 Clone Isolated from an Environmental Sample.
<3>Genome Announcements
<4>2
<5>e00462-14
<6>2014
<7>A multidrug-resistant NDM-1 carbapenamase-producing Escherichia coli sequence type 131 (ST131)
organism was obtained from vacuum cleaner dust collected from
the home of a case patient. Here, we report the assembly and annotation of its
genome.

<>

<1>Kuwahara, T., Yamashita, A., Hirakawa, H., Nakayama, H., Toh, H., Okada, N., Kuhara, S., Hattori, M., Hayashi, T., Ohnishi, Y.
<2>Genomic analysis of Bacteroides fragilis reveals extensive DNA inversions regulating cell surface adaptation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>14919-14924
<6>2004
<7>Bacteroides are predominant human colonic commensals, but the principal pathogenic species,
Bacteroides fragilis (BF), lives closely associated
with the mucosal surface, whereas a second major species, Bacteroides
thetaiotaomicron (BT), concentrates within the colon. We find
corresponding differences in their genomes, based on determination of the
genome sequence of BF and comparative analysis with BT. Both species have
acquired two mechanisms that contribute to their dominance among the
colonic microbiota: an exceptional capability to use a wide range of
dietary polysaccharides by gene amplification and the capacity to create
variable surface antigenicities by multiple DNA inversion systems.
However, the gene amplification for polysaccharide assimilation is more
developed in BT, in keeping with its internal localization. In contrast,
external antigenic structures can be changed more systematically in BF.
Thereby, at the mucosal surface, where microbes encounter continuous
attack by host defenses, BF evasion of the immune system is favored, and
its colonization and infectious potential are increased.

<>

<1>Kuzin, A.I., Bolesnin, M.I., Smolyaninov, V.V., Azizbekyan, R.R.
<2>Site specific restriction endonuclease BtcI from Bacillus thuringiensis var. canadensis.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>36-39
<6>1989
<7>Efficiency of bacteriophage Tp4 plating on Bacillus thuringiensis var.
canadensis H5 (Can) is decreased 10^7-fold as compared with the efficiency of
plating on Bacillus thuringiensis var. galleriae H5 (Gal).  Bacteriophage Tp4
having propagated for one cycle in Can cells can be further grown in this
strain without restriction.  The site specific restriction endonuclease BtcI
isolated from Bacillus thuringiensis var. canadensis recognises the same
nucleotide sequence GATC in DNA as recognised by restriction endonuclease
Sau3AI.

<>

<1>Kuzmin, N.P., Fodor, I., Baev, A.A.
<2>A specific endonuclease in Klebsiella pneumoniae OK8.
<3>Dokl. Akad. Nauk.
<4>236
<5>477-480
<6>1977
<7>None

<>

<1>Kuzmin, N.P., Loseva, S.P., Belyaeva, R.K., Kravets, A.N., Solonin, A.S., Tanyashin, V.I., Baev, A.A.
<2>Physical and catalytic properties of homogeneous restriction endonuclease EcoRV.
<3>Mol. Biol. (Mosk)
<4>18
<5>166-173
<6>1984
<7>The characteristics of restriction endonuclease EcoRV, puried earlier to a homogeneous state
from a superproductive strain constructed in vitro, were determined by gel filtration and
electrophoresis in polyacrylamide gel under denaturing conditions.  The enzyme has a molecular
weight of 25,000. Restriction endonuclease EcoRV differs immunologically from the enzymes
EcoRI and EcoRII.  The catalytic properties of restriction endonuclease EcoRV were determined:
the dependence of the enzymatic activity on the pH of the medium, the ionic strength of the
solution, the temperature, and the presence of divalent metal cations (Mn2+, Mg2+, Co2+, Zn2+,
Ni2+, Cd2+), and organic solvents (glycerol, dimethylsulfoxide, ethanol).  It was found that
the specificity of EcoRV decreases when divalent magnesium ions are replaced by manganese or
when organic solvents are added.  The enzyme is capable of digesting "protected" DNAs of
T-even bacteriophages:  glycosylated and containing 5-hydroxmethylcytosine.  Methylated EcoRV
DNA of bacteriophage lambda, grown on a strain of E. coli containing an EcoRV
restriction-modification system, is cleaved by EcoRV on noncanonical cleavage sites under
conditions of a decrease in the specificity of the enzyme.  The DNA fragments formed upon
cleavage by EcoRV on noncanonical sites are inserted into canonical EcoRV site of cleavage of
the vector plasmid pBR322 DNA molecule.

<>

<1>Kuzmin, N.P., Solonin, A.S., Tanyashin, V.I., Baev, A.A.
<2>Chromatographic purification of restriction enzyme EcoRV from producer strains of Escherichia coli.
<3>Soviet Patent Office
<4>SU 1218678
<5>
<6>1995
<7>
<>

<1>Kuznetsov, B.B., Ivanovsky, R.N., Keppen, O.I., Sukhacheva, M.V., Bumazhkin, B.K., Patutina, E.O., Beletsky, A.V., Mardanov, A.V., Baslerov, R.V., Panteleeva, A.N., Kolganova, T.V., Ravin, N.V., Skryabin, K.G.
<2>Draft genome sequence of the anoxygenic filamentous phototrophic bacterium Oscillochloris trichoides ssp. DG-6.
<3>J. Bacteriol.
<4>193
<5>321-322
<6>2010
<7>Oscillillochloris trichoides is a mesophilic filamentous photoautotrophic non-sulfur
diazotrophic bacterium which is capable for carbon dioxide fixation via the reductive pentose
phosphate cycle and possesses no assimilative sulfate reduction. Here we present the draft
genome sequence of Oscillillochloris trichoides ssp.DG6, the type strain of the specie, which
has permitted the prediction of genes for carbon and nitrogen metabolism and light harvesting
apparatus.

<>

<1>Kuznetsova, S.A., Kubareva, E.A., Oretskaya, T.S., Dolinnaya, N.G., Krynetskaya, N.F., Gromova, E.S., Shabarova, Z.A., Cech, D.
<2>Interaction between EcoRII restriction/modification enzymes and synthetic DNA fragments. Synthesis of substrates containing a single recognition site.
<3>Biopol. Kletka
<4>3
<5>283-289
<6>1987
<7>9-16 membered oligodeoxyribonucleotides forming DNA-duplexes with one EcoRII
site (native or modified) were synthesized by the block triester method.  The
modifications involved the replacement of one (or two) cytidine or thymidine
moieties in duplexes for m5dC or f5dU, respectively.  30-membered DNA-duplex
was obtained by enzymatic ligation of five overlapping oligonucleotides.  The
substitutions introduced neither result in any significant destabilization nor
distort the double helix geometry as is evidenced by the UV- and
CD-spectroscopy methods.

<>

<1>Kvachadze, L.I., Andriashvili, I.A., Chanishvili, T.G., Arutyunyan, E.E., Nikolskaia, I.I., Debov, S.S.
<2>Identification of the system modification-restriction in Staphylococci.
<3>Vopr. Med. Khim.
<4>31
<5>121-125
<6>1985
<7>A new system of host specificity of DNA, called Sau67 according to the available nomenclature,
was identified in Staphylococcus aureus 6782 strain by means of cross titration with
staphylophage 729 considering that the phage exhibited the highly effective absorption
properties.  A total preparation of Sau67 methylases was isolated using ammonium sulfate
fractionation.  The enzyme preparation contained methylases of cytosine and adenine, where the
activity of adenine methylases constituted only 5% of the total methylase activity.  As shown
by kinetics of methylation a low content of unspecific cellular nucleases was found in the St.
aureus 6782 strain; these reasons are important for isolation of restricting endonucleases
containing in the strain.  100 microg of protein of the total enzymatic fraction enabled the
methylation of the acceptor DNA at a maximal rate within 1.5 hr of incubation in phosphate
buffer, pH 7.9.  The fraction of cytosine methylases free of adenine methylating activity was
obtained after chromatography on Sepharose blue with NaCl concentration stepwise gradient.

<>

<1>Kwak, J., Jiang, H., Kendrick, K.E.
<2>Transformation using in vivo and in vitro methylation in Streptomyces griseus.
<3>FEMS Microbiol. Lett.
<4>209
<5>243-248
<6>2002
<7>Streptomyces griseus does not readily take up foreign DNA isolated from other Streptomyces
species or Escherichia coli, presumably due to its
unique restriction-modification systems that function as a barrier for
interspecific DNA transfer. To efficiently transform S. griseus by
avoiding the restriction barriers, we methylated incoming DNA in vivo
and in vitro and treated protoplasts with heat prior to transformation.
Whereas heat treatment of protoplasts or methylation of the E.
coli-Streptomyces shuttle vectors (pXE4 and pKK1443) did not
prominently improve the transformation efficiency, HpaII methylation of
the vectors from any E. coli strains tested in this study highly
increased the transformation efficiency. The highest transformation
efficiency was observed when the shuttle vectors were isolated from the
dam, hsd strain of E. coli (GM161) and methylated by AluI and HpaII
methyltransferases, and the efficiency was approximately the same as
that of the vectors from S. griseus. We identified several
restriction-modification systems that decrease the transformation
efficiency. This research also led us to understand methylation
profiles and restriction-modification systems in S. griseus.

<>

<1>Kwak, M.J., Jeong, H., Madhaiyan, M., Lee, Y., Sa, T.M., Oh, T.K., Kim, J.F.
<2>Genome Information of Methylobacterium oryzae, a Plant-Probiotic Methylotroph in the Phyllosphere.
<3>PLoS ONE
<4>9
<5>E106704
<6>2014
<7>Pink-pigmented facultative methylotrophs in the Rhizobiales are widespread in the
environment, and many Methylobacterium species associated with plants produce
plant growth-promoting substances. To gain insights into the life style at the
phyllosphere and the genetic bases of plant growth promotion, we determined and
analyzed the complete genome sequence of Methylobacterium oryzae CBMB20T, a
strain isolated from rice stem. The genome consists of a 6.29-Mb chromosome and
four plasmids, designated as pMOC1 to pMOC4. Among the 6,274 coding sequences in
the chromosome, the bacterium has, besides most of the genes for the central
metabolism, all of the essential genes for the assimilation and dissimilation of
methanol that are either located in methylotrophy islands or dispersed. M. oryzae
is equipped with several kinds of genes for adaptation to plant surfaces such as
defense against UV radiation, oxidative stress, desiccation, or nutrient
deficiency, as well as high proportion of genes related to motility and
signaling. Moreover, it has an array of genes involved in metabolic pathways that
may contribute to promotion of plant growth; they include auxin biosynthesis,
cytokine biosynthesis, vitamin B12 biosynthesis, urea metabolism, biosorption of
heavy metals or decrease of metal toxicity, pyrroloquinoline quinone
biosynthesis, 1-aminocyclopropane-1-carboxylate deamination, phosphate
solubilization, and thiosulfate oxidation. Through the genome analysis of M.
oryzae, we provide information on the full gene complement of M. oryzae that
resides in the aerial parts of plants and enhances plant growth. The
plant-associated lifestyle of M. oryzae pertaining to methylotrophy and plant
growth promotion, and its potential as a candidate for a bioinoculant targeted to
the phyllosphere and focused on phytostimulation are illuminated.

<>

<1>Kwak, M.J., Song, J.Y., Kim, S.Y., Jeong, H., Kang, S.G., Kim, B.K., Kwon, S.K., Lee, C.H., Yu, D.S., Park, S.H., Kim, J.F.
<2>Complete Genome Sequence of the Endophytic Bacterium Burkholderia sp. Strain KJ006.
<3>J. Bacteriol.
<4>194
<5>4432-4433
<6>2012
<7>Endophytes live inside plant tissues without causing any harm and may even benefit plants.
Here, we provide the high-quality genome sequence of Burkholderia
sp. strain KJ006, an endophytic bacterium of rice with antifungal activity. The
6.6-Mb genome, consisting of three chromosomes and a single plasmid, contains
genes related to plant growth promotion or degradation of aromatic compounds.

<>

<1>Kwan, T., Liu, J., Dubow, M., Gros, P., Pelletier, J.
<2>The complete genomes and proteomes of 27 Staphylococcus aureus bacteriophages.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>5174-5179
<6>2005
<7>Bacteriophages are the most abundant life forms in the biosphere. They
play important roles in bacterial ecology, evolution, adaptation to new
environments, and pathogenesis of human bacterial infections. Here, we
report the complete genomic sequences, and predicted proteins of 27
bacteriophages of the Gram-positive bacterium Staphylococcus aureus.
Comparative nucleotide and protein sequence analysis indicates that these
phages are a remarkable source of untapped genetic diversity, encoding
2,170 predicted protein-encoding ORFs, of which 1,402 cannot be annotated
for structure or function, and 522 are proteins with no similarity to
other phage or bacterial sequences. Based on their genome size,
organization of their gene map and comparative nucleotide and protein
sequence analysis, the S. aureus phages can be organized into three
groups. Comparison of their gene maps reveals extensive genome mosaicism,
hinting to a large reservoir of unidentified S. aureus phage genes. Among
the phages in the largest size class (178-214 kbp) that we characterized
is phage Twort, the first discovered bacteriophage (responsible for the
Twort-D'Herelle effect). These phage genomes offer an exciting opportunity
to discern molecular mechanisms of phage evolution and diversity.

<>

<1>Kwasiborski, A., Mondy, S., Beury-Cirou, A., Faure, D.
<2>Genome Sequence of the Quorum-Quenching Rhodococcus erythropolis Strain R138.
<3>Genome Announcements
<4>2
<5>e00224-14
<6>2014
<7>Rhodococcus erythropolis strain R138 was isolated from the rhizosphere of Solanum tuberosum
and selected for its capacity to degrade N-acyl-homoserine lactones,
quorum-sensing signals used as communication molecules by the potato pathogens
Pectobacterium and Dickeya. Here, we report the genome sequence of Rhodococcus
erythropolis strain R138.

<>

<1>Kwasiborski, A., Mondy, S., Beury-Cirou, A., Faure, D.
<2>Genome Sequence of the Pectobacterium atrosepticum Strain CFBP6276, Causing Blackleg and Soft Rot Diseases on Potato Plants and Tubers.
<3>Genome Announcements
<4>1
<5>e00374-13
<6>2013
<7>Pectobacterium atrosepticum strain CFBP6276 is a pectinolytic enterobacterium causing blackleg
and soft rot of the stem and tuber of Solanum tuberosum. Its
virulence is under the control of quorum sensing, with N-acylhomoserine lactones
as communication signals. Here, we report the genome sequence of P. atrosepticum
strain CFBP6276.

<>

<1>Kwiatek, A., Kobes, M., Olejnik, K., Piekarowicz, A.
<2>DNA methyltransferases from Neisseria meningitidis and Neisseria gonorrhoeae FA1090 associated with mismatch nicking endonucleases.
<3>Microbiology
<4>150
<5>1713-1722
<6>2004
<7>The genes encoding the DNA methyltransferases M.NmeDI and M.NmeAl from Neisseria meningitidis
associated with the genes encoding putative Vsr
endonucleases were overexpressed in Escherichia coli. The enzymes were
purified to apparent homogeneity on Ni-NTA agarose columns, yielding
proteins of 49  1 kDa and 39.6  1 kDa, respectively, under
denaturing conditions. M.NmeDI recognizes the degenerate sequence
5'-RCCGGB-3'. It methylates the first 5' cytosine residue on both
strands within the core sequence CCGG. The enzyme shows higher affinity
with the hemimethylated degenerate sequence than with the unmethylated
degenerate sequence. Comparison of the amino acid sequence of the
target-recognizing domain of M.NmeDI with the closest neighbours
recognizing the sequence 5'-RCCGGY-3' showed the presence of the
homologous domain and an additional domain that may be responsible for
recognizing the degenerate sequence. M.NmeAl recognizes the sequence
5'-CCGG-3' and methylates the second 5' cytosine residue on both DNA
strands. In Neisseria gonorrhoeae strain FA1090 the homologues of these
ORFs are truncated due to a variety of mutations.

<>

<1>Kwiatek, A., Luczkiewicz, M., Bandyra, K., Stein, D.C., Piekarowicz, A.
<2>Neisseria gonorrhoeae FA1090 encodes two classes of Vsr endonucleases.
<3>J. Bacteriol.
<4>192
<5>3951-3960
<6>2010
<7>Very Short Patch repair systems prevent mutations resulting from deamination of
5-methylcytosine to thymine. The Vsr endonuclease is the key enzyme of this system, providing
sequence specificity. We identified two genes encoding Vsr endonucleases from Neisseria
gonorrhoeae FA1090, V.NgoAXIII and V.NgoAXIV, based on DNA sequence similarity to genes
encoding Vsr endonucleases from other bacteria. After expression of the gonococcal genes in
Escherichia coli, the proteins were biochemically characterized and the endonucleolytic
activity and specificity of V.NgoAXIII and V.NgoAXIV were determined. V.NgoAXIII was found to
be multispecific and recognizes T:G mismatches in every tested nucleotide context whereas
V.NgoAXIV recognizes T:G mismatches in the following sequences: GTGG, CTGG, GTGC, ATGC or
CTGC. Alanine mutagenesis of conserved residues showed that Asp50 and His68 of V.NgoAXIII and
Asp51 and His69 of V.NgoAXIV are essential for hydrolytic activity. Glu25, His64 and Asp97 of
V.NgoAXIV and Glu24, Asp63 and Asp97 of V.NgoAXIII are important but not crucial for activity
V.NgoAXIII and V.NgoAXIV. However, Glu24 and Asp63 are also important for the specificity of
V.NgoAXIII. On the basis of our results, concerning features of Vsr endonucleases encoded by
N. gonorrhoeae FA1090, we postulate that at least two types of Vsr endonucleases can be
distinguished.

<>

<1>Kwiatek, A., Piekarowicz, A.
<2>The restriction endonuclease R.NmeDI from Neisseria meningitidis that recognizes a palindromic sequence and cuts the DNA on both sides of the  recognition sequence.
<3>Nucleic Acids Res.
<4>35
<5>6539-6546
<6>2007
<7>The restriction endonuclease Type II R.NmeDI from Neisseria meningitidis 2120 (serogroup C,
ST-11 complex) was characterized. The cloned nmeDIR
gene was expressed in Escherichia coli cells, and the endonucleolytic and
restriction activities of R.NmeDI were then observed in vitro and in vivo.
The nmeDIR gene consists of 1056 bp coding 351 aa protein with a
calculated molecular weight of M((r)) = 39 000 +/- 1000 Da. The R.NmeDI
enzyme was purified to apparent homogeneity following overexpression,
using metal affinity chromatography. This enzyme recognizes a palindrome
sequence and cleaves double-stranded DNA upstream and downstream of its
recognition sequence (12/7) RCCGGY (7/12) (R = A/G, Y = C/T) cutting out a
25-bp fragment. R.NmeDI cleaves in two steps. The enzyme cleaves the first
strand randomly on either side of the recognition sequence generating an
intermediate, and the second cleavage occurs more slowly and results in
the production of a final reaction product. The R.NmeDI endonuclease
requires two recognition sequences for effective cleavage. The tetramer is
an active form of the R.NmeDI enzyme.

<>

<1>Kwiatkowski, B., Boschek, B., Thiele, H., Stirm, S.
<2>Substrate specificity of two bacteriophage-associated endo-N-acetylneuraminidases.
<3>J. Virol.
<4>45
<5>367-374
<6>1983
<7>For Escherichia coli Bos12 (O16:K92:H-), a bacteriophage (phi 92) has been
isolated which carries a depolymerase active on the K92 capsular polysaccharide.
As seen under the electron microscope, phi 92 belongs to Bradley's morphology
group A and is different from the phage phi 1.2 previously described (Kwiatkowski
et al., J. Virol. 43:697-704, 1982), which grows on E. coli K235 (O1:K1:H-),
depolymerizes colominic acid, and belongs to morphology group C. The specificity
of the phi 1.2- and phi 92-associated endo-N-acetylneuraminidases has been
studied with respect to the following substrates (all alkali treated, and where
NeuNAc represents N-acetylneuraminic acid): (i) [-alpha-NeuNAc-(2 leads to 8)-]n
(colominic acid), (ii) [-alpha-NeuNAc-(2 leads to 8)-alpha-NeuNAc-(2 leads to
9)-]n (E. coli K92 polysaccharide), and (iii) [-alpha-NeuNAc-(2 leads to 9)-]n
(Neisseria meningitidis type C capsular polysaccharide). The increase in
periodate consumption of these glycans upon incubation with purified phi 1.2 or
phi 92 particles was measured, and the split products obtained from all
substrates after exhaustive degradation were analyzed by gel chromatography. It
was found that the Neisseria polysaccharide is not appreciably affected by either
virus enzyme and that phi 1.2 only depolymerizes a small fraction of the K92
glycan. Colominic acid, however, is completely degraded by both agents, phi 92
yielding smaller fragments (one to six NeuNAc residues) than phi 1.2 (two to
seven). Phage phi 92 additionally depolymerizes the K92 glycan, essentially to
oligosaccharides of two, four, and six residues. The size distribution of these
K92 oligosaccharides indicates that the phi 92 enzyme predominantly cleaves the
alpha(2 leads to 8) linkages in this polymer.

<>

<1>Kwoh, T.J., Kwoh, D.Y., McCue, A.W., Davis, G.R., Patrick, D., Gingeras, T.R.
<2>Introduction and expression of the bacterial PaeR7 methylase gene in mammalian cells.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>83
<5>7713-7717
<6>1986
<7>An approach is devised for studying the role of DNA methylation in eukaryotic
gene expression.  The approach is based on the expression of site-specific
bacterial methylase genes in animal cells.  A model system using the cloned
PaeR7 (an isoschizomer of XhoI) methylase gene was constructed to test the
feasibility of this approach.  Expression plasmids for the PaeR7 methylase gene
were introduced into mouse Ltk- cells by cotransfection with the cloned chicken
thymidine kinase (tk) gene.  Several of the cell strains derived from Tk+
colonies were found to express the PaeR7 gene as judged by four criteria:  the
cellular DNA of these strains showed increased resistance to cleavage by XhoI;
these strains contained cellular proteins that comigrated with pure PaeR7
methylase protein, as visualized by immunoblotting; PaeR7 methylase activity
was found in vitro in crude extracts of total cellular protein from these
strains; and murine adenovirus genomes grown on cells expressing PaeR7
methylase showed resistance to cleavage to PaeR7 endonuclease.  The potential
applications of this approach for the study of cellular and viral gene
regulation, DNA repair, and restriction modification are discussed.

<>

<1>Kwoh, T.J., Obermiller, P.S., McCue, A.W., Kwoh, D.Y., Sullivan, S.A., Gingeras, T.R.
<2>Introduction and expression of the bacterial PaeR7I restriction endonuclease gene in mouse cells containing the PaeR7I methylase.
<3>Nucleic Acids Res.
<4>16
<5>11489-11506
<6>1988
<7>To study the factors essential for a functional restriction system, the PaeR7I
restriction-modification system has been introduced and expressed in murine
cells.  Transfer of this system was accomplished in two steps.  First, cells
containing sufficient PaeR7I methylase to completely methylate the mouse genome
were constructed.  In the second step, the mouse metallothionein
promoter-regulated, endonuclease expression vector linked to the hygromycin B
resistance selection marker was used to transfect the high methylase-expressing
cells.  Sixty percent of the clones isolated contained PaeR7I endonuclease
enzymatic activity.  Transfected cells expressing both methylase and
endonuclease were incapable of blocking infection by DNA viruses, and possible
explanations are discussed.

<>

<1>Kwon, S.K., Kim, B.K., Song, J.Y., Kwak, M.J., Lee, C.H., Yoon, J.H., Oh, T.K., Kim, J.F.
<2>Genomic makeup of the marine flavobacterium Nonlabens (Donghaeana) dokdonensis DSW-6 and identification of a novel class of rhodopsins.
<3>Genome Biol. Evol.
<4>5
<5>187-199
<6>2013
<7>Rhodopsin-containing marine microbes such as those in the class Flavobacteria
play a pivotal role in the biogeochemical cycle of the euphotic zone .
Deciphering the genome information of flavobacteria and accessing the diversity
and ecological impact of microbial rhodopsins is important in understanding and
preserving the global ecosystems. The genome sequence of the orange-pigmented
marine flavobacterium Nonlabens dokdonensis (basonym: Donghaeana dokdonensis)
DSW-6 was determined. As a marine photoheterotroph, DSW-6 has written in its
genome physiological features that allow survival in marine oligotrophic
environments. The sequence analysis also uncovered a gene encoding an unexpected
type of microbial rhodopsin containing a unique motif in addition to a
proteorhodopsin gene and a number of photolyase or cryptochrome genes. Homologs
of the novel rhodopsin gene were found in other flavobacteria,
alphaproteobacteria, a species of cytophaga, a deinococcus, and even a eukaryote
diatom. They all contain the characteristic NQ motif and form a phylogenetically
distinct group. Expression analysis of this rhodopsin gene in DSW-6 indicated
that it is induced at high NaCl concentrations, as well as in the presence of
light and the absence of nutrients. Genomic and metagenomic surveys demonstrate
the diversity of the NQ rhodopsins in nature and the prevalent occurrence of the
encoding genes among microbial communities inhabiting hypersaline niches,
suggesting its involvement in sodium metabolism and the sodium-adapted lifestyle.

<>

<1>Kwon, Y.K., Kim, J.J., Kim, J.H., Jeon, S.M., Ye, B.R., Jang, J., Heo, S.J., Park, S.C., Kang, D.H., Oh, C.
<2>Draft genome sequence of the xylan-degrading marine bacterium strain s124, representing a novel species of the genus oceanicola.
<3>J. Bacteriol.
<4>194
<5>6325
<6>2012
<7>We isolated a xylan-degrading bacterium from seawater of Micronesia and identified it as
Oceanicola sp. strain S124. We sequenced the Oceanicola sp. S124
genome using GSFLX 454 pyrosequencing and predicted 4,433 open reading frames
(ORFs) including putative saccharification and phage-related genes.

<>

<1>Kwon, Y.T., Jun, H.S., Rho, H.M.
<2>Characterization of BmaI methylase from Bacillus macerans.
<3>Korean J. Microbiol.
<4>26
<5>88-92
<6>1988
<7>The isolation and characterization of a new type II methylase, BmaI methylase,
from Bacillus macerans ATCC 8244 were described.  BmaI methylase was isolated
by procedures of ammonium sulfate fractionation, DEAE-cellulose chromatography
and phosphocellulose chromatography.  Two types of methylases were present in
this strain and only one of the two was a site specific BmaI methylase.  The
pBR322 DNA methylated by BmaI methylase was not cleaved by BmaI endonuclease,
and pBR322 DNA cleaved by BmaI endonuclease was not methylated by BmaI
methylase.  The optimal pH for the BmaI methylase activity was 7.5, and optimal
NaCl concentration was about 50 mM.  BmaI methylase could methylate
single-stranded M13mp18 DNA.

<>

<1>Kwon, Y.T., Jun, H.S., Rho, H.M.
<2>Characterization of BmaI endonuclease from Bacillus macerans ATCC 8244.
<3>Korean J. Microbiol.
<4>26
<5>1-5
<6>1988
<7>The isolation and characterization of a new type II restriction endonuclease,
BmaI, from Bacillus macerans ATCC 8244 were described.  BmaI endonuclease was
partially purified by procedures of ammonium sulfate fractionation,
DEAE-cellulose and phosphocellulose chromatographies.  This enzyme recognized
one site on pBR322 DNA, two sites on Bluescribe DNA, three sites on lambda DNA
and no site on SV40 DNA.  The same cleavage patterns for various DNAs as PvuI
indicated that BmaI is an isoschizomer of PvuI whose recognition sequence is
5'-CGATCG-3'.  The optimal pH for the BmaI endonuclease activity was about 7.0
and optimal NaCl concentration was about 100 mM.  Manganese ion could partially
replace magnesium as a cofactor, but calcium could not at all.

<>

<1>Kwong, S.M., Yeo, C.C., Suwanto, A., Poh, C.L.
<2>Characterization of the endogenous plasmid from Pseudomonas alcaligenes NCIB 9867: DNA sequence and mechanism of transfer.
<3>J. Bacteriol.
<4>182
<5>81-90
<6>2000
<7>The endogenous plasmid pRA2 from Pseudomonas alcaligenes NCIB 9867 was determined to have
32,743 bp with a G+C content of 59.8%, Sequence
analysis predicted a total of 29 open reading frames, with
approximately half of them contributing towards the functions of
plasmid replication, mobilization, and stability. The Pac25I
restriction-modification system and two mobile elements, Tn5563 and
IS1633, were physically localized. An additional eight open reading
frames with unknown functions were also detected. pRA2 was genetically
tagged with the Omega Str(r)/Spc(r) gene cassette by homologous
recombination, Intrastrain transfer of pRA2-encoded genetic markers
between isogenic mutants of P. alcaligenes NCIB 9867,were observed at
high frequencies (2.4 x 10(-4) per donor). This transfer aas determined
to be mediated by a natural transformation process that required
cell-cell contact and was completely sensitive to DNase I (1 mg/ml),
Efficient transformation was also observed when pRA2 DNA was applied
directly onto the cells, while transformation with foreign plasmid DNAs
was not observed. pRA2 could be conjugally transferred into Pseudomonas
putida RA713 and KT2440 recipients only when plasmid RK2/RP4 transfer
functions were provided in trams, Plasmid stability analysis
demonstrated that pRA2 could be stably maintained in its original host,
P. alcaligenes NCIB 9867, as well as in P. putida RA713 after 100
generations of nonselective growth. Disruption of the pRA2 pac25I
restriction endonuclease gene did not alter plasmid stability, while
the pRA2 minireplicon exhibited only partial stability, This indicates
that other pRA2-encoded determinants could have significant roles in
influencing plasmid stability.

<>

<1>Kwong, W.K., Engel, P., Koch, H., Moran, N.A.
<2>Genomics and host specialization of honey bee and bumble bee gut symbionts.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>111
<5>11509-11514
<6>2014
<7>Gilliamella apicola and Snodgrassella alvi are dominant members of the honey bee  (Apis spp.)
and bumble bee (Bombus spp.) gut microbiota. We generated complete
genomes of the type strains G. apicola wkB1(T) and S. alvi wkB2(T) (isolated from
Apis), as well as draft genomes for four other strains from Bombus. G. apicola
and S. alvi were found to occupy very different metabolic niches: The former is a
saccharolytic fermenter, whereas the latter is an oxidizer of carboxylic acids.
Together, they may form a syntrophic network for partitioning of metabolic
resources. Both species possessed numerous genes [type 6 secretion systems,
repeats in toxin (RTX) toxins, RHS proteins, adhesins, and type IV pili] that
likely mediate cell-cell interactions and gut colonization. Variation in these
genes could account for the host fidelity of strains observed in previous
phylogenetic studies. Here, we also show the first experimental evidence, to our
knowledge, for this specificity in vivo: Strains of S. alvi were able to colonize
their native bee host but not bees of another genus. Consistent with specific,
long-term host association, comparative genomic analysis revealed a deep
divergence and little or no gene flow between Apis and Bombus gut symbionts.
However, within a host type (Apis or Bombus), we detected signs of horizontal
gene transfer between G. apicola and S. alvi, demonstrating the importance of the
broader gut community in shaping the evolution of any one member. Our results
show that host specificity is likely driven by multiple factors, including direct
host-microbe interactions, microbe-microbe interactions, and social transmission.

<>

<1>Kwong, W.K., Mancenido, A.L., Moran, N.A.
<2>Genome Sequences of Lactobacillus sp. Strains wkB8 and wkB10, Members of the Firm-5 Clade, from Honey Bee Guts.
<3>Genome Announcements
<4>2
<5>e01176-14
<6>2014
<7>We sequenced two strains from the Lactobacillus Firm-5 clade, a dominant group of symbionts in
the guts of honey bees and other social bees. The genome of strain
wkB8, comprising a 1.93-Mb chromosome and a 6.4-kb plasmid, was fully closed,
while strain wkB10 was assembled into 32 contigs. These genomes will provide
insights into how gut symbionts evolve and interact with their host species.

<>

<1>Kwun, M.J., Cheng, J., Yang, S.H., Lee, D.R., Suh, J.W., Hong, H.J.
<2>Draft Genome Sequence of Ristocetin-Producing Strain Amycolatopsis sp. Strain MJM2582 Isolated in South Korea.
<3>Genome Announcements
<4>2
<5>e01091-14
<6>2014
<7>The draft genome sequence of a ristocetin-producing Amycolatopsis strain (sp. MJM2582)
isolated in South Korea is reported here. This strain has a genome of
approximately 8.9 Mb containing 7,933 predicted genes, including the ristocetin
cluster and 32 additional predicted secondary metabolite biosynthesis clusters.

<>

<1>Kwun, M.J., Hong, H.J.
<2>Genome Sequence of Streptomyces toyocaensis NRRL 15009, Producer of the Glycopeptide Antibiotic A47934.
<3>Genome Announcements
<4>2
<5>e00749-14
<6>2014
<7>Here we report the draft genome sequence of Streptomyces toyocaensis strain NRRL  15009 which
is the producer of the glycopeptide antibiotic A47934. The genome
sequence is predicted to harbor a total of 26 secondary metabolite biosynthetic
gene clusters including the A47934 cluster.

<>

<1>Kwun, M.J., Hong, H.J.
<2>Draft Genome Sequence of Amycolatopsis lurida NRRL 2430, Producer of the Glycopeptide Family Antibiotic Ristocetin.
<3>Genome Announcements
<4>2
<5>e01050-14
<6>2014
<7>We report here the first draft genome sequence for Amycolatopsis lurida NRRL 2430, the
producer of the glycopeptide antibiotic ristocetin. The 9-Mbp genome is
predicted to harbor 8,143 genes, including those belonging to the ristocetin
biosynthesis cluster and 31 additional predicted secondary metabolite gene
clusters.

<>

<1>Kyle, J.L., Cummings, C.A., Parker, C.T., Quinones, B., Vatta, P., Newton, E., Huynh, S., Swimley, M., Degoricija, L., Barker, M., Fontanoz, S., Nguyen, K., Patel, R., Fang, R., Tebbs, R., Petrauskene, O., Furtado, M., Mandrell, R.E.
<2>Escherichia coli Serotype O55:H7 Diversity Supports Parallel Acquisition of Bacteriophage at Shiga Toxin Phage Insertion Sites during Evolution of the O157:H7 Lineage.
<3>J. Bacteriol.
<4>194
<5>1885-1896
<6>2012
<7>Enteropathogenic Escherichia coli (EPEC) continues to be a leading cause of
mortality and morbidity in children around the world. Two EPEC genomes have been
fully sequenced: those of EPEC O127:H6 strain E2348/69 (United Kingdom, 1969) and
EPEC O55:H7 strain CB9615 (Germany, 2003). The O55:H7 serotype is a recent
precursor to the virulent enterohemorrhagic E. coli O157:H7. To explore the
diversity of O55:H7 and better understand the clonal evolution of O157:H7, we
fully sequenced EPEC O55:H7 strain RM12579 (California, 1974), which was
collected 1 year before the first U.S. isolate of O157:H7 was identified in
California. Phage-related sequences accounted for nearly all differences between
the two O55:H7 strains. Additionally, O55:H7 and O157:H7 strains were tested for
the presence and insertion sites of Shiga toxin gene (stx)-containing
bacteriophages. Analysis of non-phage-associated genes supported core elements of
previous O157:H7 stepwise evolutionary models, whereas phage composition and
insertion analyses suggested a key refinement. Specifically, the placement and
presence of lambda-like bacteriophages (including those containing stx) should
not be considered stable evolutionary markers or be required in placing O55:H7
and O157:H7 strains within the stepwise evolutionary models. Additionally, we
suggest that a 10.9-kb region (block 172) previously believed unique to O55:H7
strains can be used to identify early O157:H7 strains. Finally, we defined two
subsets of O55:H7 strains that share an as-yet-unobserved or extinct common
ancestor with O157:H7 strains. Exploration of O55:H7 diversity improved our
understanding of the evolution of E. coli O157:H7 and suggested a key revision to
accommodate existing and future configurations of stx-containing bacteriophages
into current models.

<>

<1>Kyune, C., Shimonchitoshu, A.
<2>chimeric polypeptides and their use.
<3>Japanese Patent Office
<4>JP 2006523448 A
<5>
<6>2006
<7>
<>

<1>L'abee-Lund, T.M., Jorgensen, H.J., O'Sullivan, K., Bohlin, J., Ligard, G., Granum, P.E., Lindback, T.
<2>The Highly Virulent 2006 Norwegian EHEC O103:H25 Outbreak Strain Is Related to the 2011 German O104:H4 Outbreak Strain.
<3>PLoS ONE
<4>7
<5>E31413
<6>2012
<7>In 2006, a severe foodborne EHEC outbreak occured in Norway. Seventeen cases were
recorded and the HUS frequency was 60%. The causative strain, Esherichia coli
O103:H25, is considered to be particularly virulent. Sequencing of the outbreak
strain revealed resemblance to the 2011 German outbreak strain E. coli O104:H4,
both in genome and Shiga toxin 2-encoding (Stx2) phage sequence. The nucleotide
identity between the Stx2 phages from the Norwegian and German outbreak strains
was 90%. During the 2006 outbreak, stx(2)-positive O103:H25 E. coli was isolated
from two patients. All the other outbreak associated isolates, including all food
isolates, were stx-negative, and carried a different phage replacing the Stx2
phage. This phage was of similar size to the Stx2 phage, but had a distinctive
early phage region and no stx gene. The sequence of the early region of this
phage was not retrieved from the bacterial host genome, and the origin of the
phage is unknown. The contaminated food most likely contained a mixture of E.
coli O103:H25 cells with either one of the phages.

<>

<1>Laayoun, A., Baker, D.J., Riley, J., Smith, S.S.
<2>The response of M.HpaII to heteroduplexes.
<3>Gene
<4>150
<5>195-196
<6>1994
<7>Synthetic oligodeoxyribonucleotide duplexes have been used to study the methylation
specificity of M.HpaII, a bacterial DNA methyltransferase. Substrates of four types were
compared. A 30-mer containing a Watson-Crick paired CCGG recognition sequence was rapidly
methylated at the central cytosine on each strand in the recognition sequence. A 30-mer
containing an asymmetrically methylated recognition sequence, of the type transiently produced
by DNA replication, was rapidly methylated at the central cytosine on the unmethylated strand.
A heteroduplex containing an A.C mispair in the recognition sequence (CCGG/CCAG) was rapidly
methylated at the cytosine in the mispair. A heteroduplex containing an A.C and an adjacent
C.C mispair in the recognition sequence (CCGG/CCCA) was not methylated at a significant rate.
The results show that M.HpaII can tolerate a single mispair at its recognition site in a
heteroduplex without loss of activity or specificity.

<>

<1>Laayoun, A., Smith, S.S.
<2>Methylation of slipped duplexes, snapbacks and cruciforms by human DNA(cytosine-5)methyltransferase.
<3>Nucleic Acids Res.
<4>23
<5>1584-1589
<6>1995
<7>When human DNA(cytosine-5)methyltransferase was used to methylate a series of snapback
oligodeoxynucleotides of differing stem lengths, each containing a centrally located CG
dinucleotide recognition site, the enzyme required a minimum of 22 base pairs in the stem for
maximum activity. Extrahelical cytosines in slipped duplexes that were 30 base pairs in length
acted as effective methyl acceptors and were more rapidly methylated than cytosines that were
Watson-Crick paired. Duplexes containing hairpins of CCG repeats in cruciform structures in
which the enzyme recognition sequence was disrupted by a C.C mispair were also more rapidly
methylated than control Watson-Crick-paired duplexes. Since enzymes have higher affinities for
their transition states than for their substrates, the results with extrahelical and mispaired
cytosines suggest that these structures can be viewed as analogs of the transition state
intermediates produced during catalysis by methyltransferases.

<>

<1>Labahn, J., Granzin, J., Schluckebier, G., Robinson, D.P., Jack, W.E., Schildkraut, I., Saenger, W.
<2>Three-dimensional structure of the adenine-specific DNA methyltransferase M.TaqI in complex with the cofactor S-adenosylmethionine.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>10957-10961
<6>1994
<7>The Thermus aquaticus DNA methyltransferase M.TaqI (EC 2.1.1.72) methylates N6 of adenine in
the specific double-helical DNA sequence TCGA by transfer of -CH3 from the cofactor
S-adenosyl-L-methionine. The x-ray crystal structure at 2.4 Angstrom resolution of this enzyme
in complex with S-adenosylmethionine shows alpha/beta folding of the polypeptide into two
domains of about equal size. They are arranged in the form of a C with a wide cleft suitable
to accommodate the DNA substrate. The N-terminal domain is dominated by a nine-stranded
beta-sheet; it contains the two conserved segments typical for N-methyltranferases which form
a pocket for cofactor binding. The C-terminal domain is formed by four small beta-sheets and
alpha-helices. The three-dimensional folding of M.TaqI is similar to that of the
cytosine-specific HhaI methyltransferase, where the large Beta-sheet in the N-terminal domain
contains all conserved segments and the enzymatically functional parts, and the smaller
C-terminal domain is less structured.

<>

<1>Labbe, D., Holtke, H.J., Lau, P.C.K.
<2>Cloning and characterization of two tandemly arranged DNA methyltransferase genes of Neisseria lactamica: An adenine-specific M.NlaIII and a cytosine-type methylase.
<3>Mol. Gen. Genet.
<4>224
<5>101-110
<6>1990
<7>The gene encoding the Neisseria lactamica III DNA methyltransferase (M.NlaIII) which
recognizes the sequence CATG has been cloned and expressed in Escherichia coli. DNA sequencing
of a 3.125 kb EcoRI-PstI fragment localizes the M.NlaIII gene to a 334 codon open reading
frame (ORF) and identifies, 468 bp downstream, a second ORF of 313 amino acids, which is
referred to as M.NlaX. Both proteins are detectable in the E. coli coupled in vitro
transcription-translation system; they are apparently expressed from separate N. lactamica
promoters. The N-terminal half of the previously characterized M.FokI, which methylates
adenine in one of the DNA strands with its asymmetric recognition sequence (GGATG), is found
to have 41% sequence identity and a further 11.7% sequence similarity with M.NlaIII. Among the
conserved amino acids is the well known DPPY sequence motif. With one exception, analysis of
the nucleotides coding for the DP dipeptide in all known DPPY sequences shows the presence of
an inherent DNA adenine methylation (dam) recognition site of GATC. A low level of expression
of M.NlaX in E. coli prevents the elucidation of its sequence recognition specificity.
Sequence analysis of M.NlaX shows that it is closely related to the group of monospecific
5-methylcytosine DNA methyltransferases (M.EcoRII, Dcm, M.HpaII and M.HhaI) which all have a
modified cytosine at the second position of the recognition sequences. Both M.EcoRII and Dcm
amino acid sequences are about 50% identical with M.NlaX; a considerable degree of sequence
identity is found in the so-called variable region which is believed to be responsible for
sequence recognition specificity. M.NlaX is probably the counterpart to the E. coli Dcm in N.
lactamica.

<>

<1>Labbe, S., Xia, Y., Roy, P.H.
<2>BspMII and AccIII are an isoschizomer pair which differ in their sensitivity to cytosine methylation.
<3>Nucleic Acids Res.
<4>16
<5>7184
<6>1988
<7>The latest tabulation of restriction endonuclease sensitivities to
site-specific DNA methylation has shown that BspMII is able to cut the
methylated sequence TCCGGmA while AccIII is not.  We found that cellular DNA
from Acinetobacter calcoaceticus showed complete resistance to the action of
restriction endonuclease AccIII but sensitivity to the action of restriction
endonuclease BspMII, an isoschizomer of AccIII (results not shown).  This
experiment indicates that the methylation sensitivity of restriction enzymes
AccIII and BspMII are not the same.  We then tested both enzymes (AccIII and
BspMII) for sensitivity to cytosine methylation within their recognition sites
by using M.MspI (mCCGG) and M.HpaII (CmCGG) methylases on various DNA
substrates.  Adenovirus 2 DNA was used for this experiment (Fig. 1) and
additional tests were done with lambda DNA and pLQ61 DNA (which has 2 AccIII
sites and is derived from pBR328) (results not shown).  We conclude that BspMII
does not cleave the modified sequences TmCCGGA and TCmCGGA, and that
restriction by AccIII is not affected by methylation of either cytosine in the
same recognization sequence.  Thus, we suggest that the enzyme M.BspMII is a
DNA-cytosine-methyltransferase while M.AccIII is a
DNA-adenine-methyltransferase.

<>

<1>Labeots, L.A.
<2>Structure and function of DNA phosphotriester substrates of the restriction endonuclease AluI.
<3>Diss. Abstr.
<4>53
<5>2842
<6>1993
<7>The duplex d(GGAAGCTAGG).d(CCTAGCTTCC) is a substrate for AluI, which cleaves at the center of
the AGCT recognition sequence when magnesium ion is present. Large scale synthesis of the
2,2,2-trichloro-1,1-dimethylethyl phosphotriester derivative d(GGAAGp(TCDME)CTAGG) was done
using phosphite chemistry. The two diastereomers were separated on a large scale by HPLC. Both
diastereomers formed a stable heteroduplex with a complementary unmodified strand in the
hydrolysis buffer. Both modified duplexes bound to AluI with affinity similar to that of the
natural duplex, as determined by gel shift assays. When incubated with AluI under identical
conditions, one diastereomeric heteroduplex remained intact, while the other was hydrolyzed at
G-C in the unmodified strand. No cleavage of the modified strand was observed in either case,
nor was any hydrolysis detected when the single-stranded unmodified complement was treated.
The configuration at the phosphotriester sites was assigned from 2D NMR spectral data.
Resonance assignments for most nonexchangeable protons were made using DQF-COSY and NOESY.
Most resonances fell within expected ranges; however the central dG H3' and dC H5'{5'}
residues in the modified strands were shifted significantly downfield. The diastereomer
exhibiting NOE cross-relaxation between the methyl protons in the TCDME modification and the
C6 H6, C6 H4', C6 H5' H5{'}, G5 H3', and G5 H4' in the modified strand was assigned the
Rp configuration. The diastereomer showing cross-relaxation between the methyl protons of the
TCDME modification and the C6 H5'H5{'}, G5 H3', and G5 H4' in the modified strand was
assigned the Sp configuration. This assignment agrees with one based on chemical degradation
of the oligomers. The 3D structuers of the heteroduplexes were determined by integrating
resolved NOE cross-peaks and converting the volumes into distances for each mixing time. The
derived distances were applied to models and subjected to refinement. The refined structures
correspond to B-form DNA but with distortions in the region of the modified site. The triester
group of the Sp duplex projects out from the double helix into the aqueous environment,
whereas that of the Rp isomer points into the major groove.

<>

<1>Laborda, P.R., Fonseca, F.S., Angolini, C.F., Oliveira, V.M., Souza, A.P., Marsaioli, A.J.
<2>Genome Sequence of Bacillus safensis CFA06, Isolated from Biodegraded Petroleum in Brazil.
<3>Genome Announcements
<4>2
<5>e00642-14
<6>2014
<7>Bacillus safensis is a microorganism recognized for its biotechnological and industrial
potential due to its interesting enzymatic portfolio. Here, as a means
of gathering information about the importance of this species in oil
biodegradation, we report a draft genome sequence of a strain isolated from
petroleum.

<>

<1>Labrie, S.J., El Haddad, L., Tremblay, D.M., Plante, P.L., Wasserscheid, J., Dumaresq, J., Dewar, K., Corbeil, J., Moineau, S.
<2>First Complete Genome Sequence of Staphylococcus xylosus, a Meat Starter Culture  and a Host to Propagate Staphylococcus aureus Phages.
<3>Genome Announcements
<4>2
<5>e00671-14
<6>2014
<7>Staphylococcus xylosus is a bacterial species used in meat fermentation and a commensal
microorganism found on animals. We present the first complete circular
genome from this species. The genome is composed of 2,757,557 bp, with a G+C
content of 32.9%, and contains 2,514 genes and 79 structural RNAs.

<>

<1>Labrie, S.J., Samson, J.E., Moineau, S.
<2>Bacteriophage resistance mechanisms.
<3>Nat. Rev. Microbiol.
<4>8
<5>317-327
<6>2010
<7>Phages are now acknowledged as the most abundant microorganisms on the planet and are also
possibly the most diversified. This diversity is mostly driven by their dynamic adaptation
when facing selective pressure such as phage resistance mechanisms, which are widespread in
bacterial hosts. When infecting bacterial cells, phages face a range of antiviral mechanisms,
and they have evolved multiple tactics to avoid, circumvent or subvert these mechanisms in
order to thrive in most environments. In this Review, we highlight the most important
antiviral mechanisms of bacteria as well as the counter-attacks used by phages to evade these
systems.

<>

<1>Labutti, K. et al.
<2>Permanent draft genome sequence of Dethiosulfovibrio peptidovorans type strain (SEBR 4207).
<3>Standards in Genomic Sciences
<4>3
<5>85-92
<6>2010
<7>Dethiosulfovibrio peptidovorans Magot et al. 1997 is the type species of the genus
Dethiosulfovibrio of the family Synergistaceae in the recently created
phylum Synergistetes. The strictly anaerobic, vibriod, thiosulfate-reducing
bacterium utilizes peptides and amino acids, but neither sugars nor fatty acids.
It was isolated from an offshore oil well where it was been reported to be
involved in pitting corrosion of mild steel. Initially, this bacterium was
described as a distant relative of the genus Thermoanaerobacter, but was not
assigned to a genus, it was subsequently placed into the novel phylum
Synergistetes. A large number of repeats in the genome sequence prevented an
economically justifiable closure of the last gaps. This is only the third
published genome from a member of the phylum Synergistetes. The 2,576,359 bp long
genome consists of three contigs with 2,458 protein-coding and 59 RNA genes and
is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Labutti, K. et al.
<2>Complete genome sequence of Anaerococcus prevotii type strain (PC1).
<3>Standards in Genomic Sciences
<4>1
<5>159-165
<6>2009
<7>Anaerococcus prevotii (Foubert and Douglas 1948) Ezaki et al. 2001 is the type species of the
genus, and is of phylogenetic interest because of its arguable
assignment to the provisionally arranged family 'Peptostreptococcaceae'. A.
prevotii is an obligate anaerobic coccus, usually arranged in clumps or tetrads.
The strain, whose genome is described here, was originally isolated from human
plasma; other strains of the species were also isolated from clinical specimen.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. This is the first completed genome sequence of a member
of the genus. Next to Finegoldia magna, A. prevotii is only the second species
from the family 'Peptostreptococcaceae' for which a complete genome sequence is
described. The 1,998,633 bp long genome (chromosome and one plasmid) with its
1852 protein-coding and 61 RNA genes is a part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Labutti, K. et al.
<2>Complete genome sequence of Planctomyces limnophilus type strain (Mu 290).
<3>Standards in Genomic Sciences
<4>3
<5>47-56
<6>2010
<7>Planctomyces limnophilus Hirsch and Muller 1986 belongs to the order Planctomycetales, which
differs from other bacterial taxa by several distinctive
features such as internal cell compartmentalization, multiplication by forming
buds directly from the spherical, ovoid or pear-shaped mother cell and a cell
wall which is stabilized by a proteinaceous layer rather than a peptidoglycan
layer. Besides Pirellula staleyi, this is the second completed genome sequence of
the family Planctomycetaceae. P. limnophilus is of interest because it differs
from Pirellula by the presence of a stalk and its structure of fibril bundles,
its cell shape and size, the formation of multicellular rosettes, low salt
tolerance and red pigmented colonies. The 5,460,085 bp long genome with its 4,304
protein-coding and 66 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Lacap-Bugler, D.C., Jiang, J., Huo, Y.B., Chan, Y., Leung, F.C., Watt, R.M.
<2>Complete Genome Sequence of the Oral Spirochete Bacterium Treponema putidum Strain OMZ 758T (ATCC 700334T).
<3>Genome Announcements
<4>2
<5>e01076-14
<6>2014
<7>The oral spirochete bacterium Treponema putidum inhabits human periodontal niches. The
complete genome sequence of the OMZ 758(T) (ATCC 700334(T)) strain of
this species was determined, revealing a 2,796,913-bp chromosome, with a G+C
content of 37.30% and a single plasmid (pTPu1; 3,649 bp) identical to pTS1 from
Treponema denticola.

<>

<1>Lackner, G., Moebius, N., Partida-Martinez, L., Hertweck, C.
<2>Complete Genome Sequence of Burkholderia rhizoxinica, the Endosymbiont of Rhizopus microsporus.
<3>J. Bacteriol.
<4>193
<5>783-784
<6>2010
<7>Burkholderia rhizoxinica is an intracellular symbiont of the phytopathogenic fungus Rhizopus
microsporus. The vertically transmitted
endosymbiont not only delivers the antimitotic macrolide rhizoxin to its
host, but is also essential for vegetative spore formation of the fungus.
To shed light on the genetic equipment of this model organism, we
sequenced the whole genome of B. rhizoxinica HKI 0454, thus providing the
first genomic insight into of an intracellular mutualist of a fungal
species. The 3.75 Mb genome consists of a chromosome and two
strain-specific plasmids. Primary metabolism appears to be specialized for
the uptake of fungal metabolites. Besides the rhizoxin biosynthesis gene
cluster, there are 14 loci coding for nonribosomal peptide synthetase
(NRPS) assembly lines, which represent novel targets for genomic mining of
cryptic natural products. Furthermore, the endosymbionts are equipped with
a repertoire of virulence-related factors, which can now be studied to
elucidate molecular mechanisms underlying bacterial-fungal interaction.

<>

<1>Lacks, S., Greenberg, B.
<2>A deoxyribonuclease of Diplococcus pneumoniae specific for methylated DNA.
<3>J. Biol. Chem.
<4>250
<5>4060-4066
<6>1975
<7>A deoxyribonuclease specific for methylated DNA was isolated from Diplococcus
pneumoniae.  The enzyme, an endonuclease, degrades DNA from Escherichia coli to
fragments of average molecular weight about half a million; it forms discrete
fragments from phage lambda DNA.  Methyl-deficient E. coli DNA is not attacked,
neither is DNA from Micrococcus radiodurans, which contains no methylated
adenine or cytosine.  Nor is DNA from D. pneumoniae or phage T7 attacked.
However, DNA from M. radiodurans, D. pneumoniae, and T7 is attacked after
methylation with an E. coli extract.  Methylated T7 DNA is degraded to discrete
fragments.  Although the genetic transforming activity of normal DNA from D.
pneumoniae is not affected by the enzyme, transforming activity of methylated
DNA is destroyed.  The enzyme is designated endonuclease R.DpnI.  Under certain
conditions another enzyme of complementary specificity can be isolated.  This
enzyme, designated endonuclease R.DpnII, produces a similar pattern of
fragments from the DNA of T7 without prior methylation of the DNA.  It also
degrades normal DNA from D. pneumoniae.  It is suggested that this pair of
enzymes plays a role in some unknown control process, which would involve a
large fraction of the specific base sequences that are methylated in E. coli
DNA and are present but not methylated in DNA from other sources.

<>

<1>Lacks, S., Greenberg, B.
<2>Complementary specificity of restriction endonucleases of Diplococcus pneumoniae with respect to DNA methylation.
<3>J. Mol. Biol.
<4>114
<5>153-168
<6>1977
<7>Restriction endonucleases DpnI and DpnII are produced by two distinct strains of Diplococcus
pneumoniae. The two enzymes show complementary specificity with respect to methylation of
sites in DNA. From the identity of its cleavage site with that of MboI, it appears that DpnII
cleaves at the unmodified sequence 5'-G-A-T-C-3'. DpnI cleaves at the same sequence when the
adenine residue is methylated. Both enzymes produce only double-stranded breaks in susceptible
DNA. Their susceptibility to DpnI and not DpnII shows that essentially all the G-A-T-C
sequences are methylated in DNA from the pneumococcal strain that produces DpnII as well as in
DNA from Hemophilus influenzae and Escherichia coli. In the dam-3 mutant of E. coli none of
these sequences appear to be methylated. Residual adenine methylation in the dam-3 mutant DNA
most likely occurs at different sites. Different but characteristic degrees of methylation at
G-A-T-C sites are found in the DNA of bacterial viruses grown in E. coli. DNAs from mammalian
cells and viruses are not methylated at this sequence. Mitochondrial DNA from Paramecium
aurelia is not methylated, but a small proportion of G-A-T-C sequences in the macronuclear DNA
of this eukaryote appear to be methylated. Possible roles of sequence-specific methylation in
the accommodation of plasmids, in the replication of DNA, in the regulation of gene function
and in the restriction of viral infection are discussed.

<>

<1>Lacks, S., Neuberger, M.
<2>Membrane location of a deoxyribonuclease implicated in the genetic transformation of Diplococcus pneumoniae.
<3>J. Bacteriol.
<4>124
<5>1321-1329
<6>1975
<7>The cellular localization of enzymes in Diplococcus pneumoniae was examined by
fractionation of spheroplasts.  A deoxyribonuclease implicated in the entry of
deoxyribonucleic acid (DNA) into the cell during genetic transformation was
located in the cell membrane.  This enzyme, the major endonuclease of the cell
(endonuclease I), which is necessary for the conversion of donor DNA to single
strands inside the cell and oligonucleotides outside, thus could act at the
cell surface.  Another enzyme, the cell wall lysin (autolysin), was also found
in the membrane fraction.  Other enzymes, including amylomaltase, two
exonucleases, an adenosine triphosphate-dependent deoxyribonuclease, and a
restriction type endonuclease, were predominantly periplasmic in location.
Spheroplasts were obtained spontaneously on incubation of pneumococcal cells in
concentrated sugar solutions.  The autolytic enzyme appears to be involved in
this process.  Cells that were physiologically competent to take up DNA formed
osmotically sensitive spheroplasts two to three times faster than cells that
were not in the competent state.  Although some genetically incompetent mutants
also formed spheroplasts more slowly, other such mutants formed them at the
faster rate.

<>

<1>Lacks, S.A.
<2>Purification and properties of the complementary endonucleases DpnI and DpnII.
<3>Methods Enzymol.
<4>65
<5>138-146
<6>1980
<7>None

<>

<1>Lacks, S.A.
<2>Recombinant plasmids for encoding restriction enzymes DpnI and DpnII of Streptococcus pneumoniae.
<3>US Patent Office
<4>US 4960707
<5>
<6>1990
<7>Chromosomal DNA cassettes containing genes encoding either the DpnI or DpnII restriction
endonuclease from Streptococcus pneumoniae are cloned into a streptococcal vector, pLS101.
Large amounts of the restriction enzymes are produced by cells containing the multi-copy
plasmids, pLS202, and pLS207, and their derivatives pLS201, pLS211, pLS217, pLS251 and pLS252.

<>

<1>Lacks, S.A., Ayalew, S., de la Campa, A.G., Greenberg, B.
<2>Regulation of competence for genetic transformation in Streptococcus pneumoniae: expression of dpnA, a late competence gene encoding a DNA methyltransferase of the DpnII restriction system.
<3>Mol. Microbiol.
<4>35
<5>1089-1098
<6>2000
<7>The chromosomal DpnII gene cassette of Streptococcus pneumoniae encodes two methyltransferases
and an endonuclease. One methyltransferase acts on double-stranded and the other on
single-stranded DNA. Two mRNAs are transcribed from the cassette. One, a SigA promoter
transcript, includes all three genes; the other includes a truncated form of the second
methyltransferase gene (dpnA) and the endonuclease gene. The truncated dpnA, which is
translated from the second start codon in the full gene, was shown to produce active enzyme. A
promoter reporter plasmid for S. pneumoniae was devised to characterize the promoter for the
second mRNA. This transcript was found to depend on a promoter that responded to the induction
of competence for genetic transformation. The promoter contains the combox sequence recognized
by a SigH-containing RNA polymerase. As part of the competence regulon, the dpnA gene makes a
product able to methylate incoming plasmid strands to protect them from the endonuclease and
allow plasmid establishment. Its function differs from most genes in the regulon, which are
involved in DNA uptake. Comparison of R6 and Rx strains of S. pneumoniae showed the
temperature dependence of transformation in R6 to result from temperature sensitivity of the
uptake apparatus and not the development of competence.

<>

<1>Lacks, S.A., Dunn, J.J., Greenberg, B.
<2>Identification of base mismatches recognized by the heteroduplex-DNA-repair system of Streptococcus pneumoniae.
<3>Cell
<4>31
<5>327-336
<6>1982
<7>The susceptibility to repair of particular base mismatches by the hex system of
Streptococcus pneumoniae was examined by comparison of the nucleotide sequence
of the wild-type and eight mutant alleles of the malM gene.  A detailed
restriction map was constructed for pLS70, and the nucleotide sequence was
determined for its 3475 bp chromosomal insert, which contains the entire malM
gene (encoding amylomaltase), portions of malX and malP (encoding a membrane
protein and a phosphorylase, respectively) and a control region.  Transition
mismatches were highly susceptible to repair; transversion mismatches, much
less so.  A mismatch caused by a single-nucleotide deletion was reparable, but
mismatches with longer deletions were not.  The hex system also reduced
spontaneous reversion of mutations corresponding to transitions.  It is
suggested that recognition of donor or nascent DNA strands by the hex system
depends on single-strand breaks in the target strand, and that the role of DNA
methylation in mismatch repair of Escherichia coli can be accommodated to this
model.

<>

<1>Lacks, S.A., Greenberg, B.
<2>Atypical ribosome binding sites and regulation of gene expression in the DpnII restriction enzyme system of S. pneumoniae.
<3>FASEB J.
<4>7
<5>A1082
<6>1993
<7>Strains of Streptococcus pneumoniae express either the DpnI or DpnII restriction systems,
which are complementary in that DpnI cleaves methylated GATC sites, whereas DpnII cleaves
unmethylated GATC. The genes for each system are contained in a cassette located at one
particular position in the chromosome. The DpnII cassette contains three genes in an operon
that specifies two methylases, DpnM and DpnA, and the DpnII endonuclease. Translation of DpnM
and DpnA appears to depend on atypical ribosome binding sites, containing 5'-ATTTC-(5 or
6n)-TATA-3' sequences located upstream of the start codon, rather than Shine-Dalgarno
sequences. Changes within this atypical sequence, but not outside it, block translation.
Proteins appear to be synthesized from atypical sites equally well in S. pneumoniae and E.
coli. The atypical sequence, also, is complementary to an unpaired region of 16S rRNA. These
unusual ribosome binding sites may play a role in the selective translation of methylases
prior to the endonuclease when the DpnII cassette is introduced into a cell.

<>

<1>Lacks, S.A., Greenberg, B., Sabelnikov, A.G.
<2>Possible regulation of DNA methyltransferase expression by RNA processing in Streptococcus pneumoniae.
<3>Gene
<4>157
<5>209-212
<6>1995
<7>Atypical ribosome-binding sites lacking Shine-Dalgarno sequences appear to be used for
translation of the DpnM and DpnA DNA methyltransferases of the DpnII restriction system.
Preliminary results indicate that the 5'-endpoints of DpnII system mRNAs result from
degradation of the original transcript.  These tentative findings serve as the basis for a
possible regulatory model that would accommodate the DpnII cassette either as a single copy in
the chromosome or on a multicopy plasmid.

<>

<1>Lacks, S.A., Mannarelli, B.M., Springhorn, S.S., Greenberg, B.
<2>Genetic basis of the complementary DpnI and DpnII restriction systems of S. pneumoniae:  An intercellular cassette mechanism.
<3>Cell
<4>46
<5>993-1000
<6>1986
<7>Cells of S. pneumoniae contain either DpnI, a restriction endonuclease that
cleaves only the methylated DNA sequence 5'-GmeATC-3', or DpnII, which cleaves
the same sequence when not methylated.  A chromosomal DNA segment containing
DpnII genes was cloned in S. pneumoniae.  Nucleotide sequencing of this segment
revealed genes encoding the methylase and endonuclease and a third protein of
unknown function.  When the plasmid was introduced into DpnI cells,
recombination during chromosomal facilitation of its establishment substituted
genes encoding the DpnI endonuclease and another protein in place of the DpnII
genes.  DNA hybridization and sequencing showed that the DpnI and DpnII
segments share homology on either side but not between themselves or with other
regions of the chromosome.  Thus, the complementary restriction systems are
found on nonhomologous and mutally exclusive cassettes that can be inserted
into a particular point in the chromosome of S. pneumoniae on the basis of
neighboring homology.

<>

<1>Lacks, S.A., Mannarelli, B.M., Springhorn, S.S., Greenberg, B., de la Campa, A.
<2>Genetics of the complementary restriction systems DpnI and DpnII revealed by cloning and recombination in Streptococcus pneumoniae.
<3>Streptococcal Genetics, American Society for Microbiology, Ferretti, J.J., Curtiss, R., III, Washington, DC
<4>0
<5>31-41
<6>1987
<7>Restriction enzymes, because they are able to recognize and cleave specific
sequences in DNA, have had an enormous impact on genetic analysis and
engineering.  This review is concerned with some unusual restriction enzyme
systems found in Streptococcus pneumonia.

<>

<1>Lacks, S.A., Sabelnikov, A.G., Chen, J.-D., Greenberg, B.
<2>Gene expression in the DpnI and DpnII restriction enzyme systems of Streptococcus pneumoniae.
<3>DNA Transfer and Gene Expression in Microorganisms, Intercept, Balla, E., Berencsi, G., Szentirmai, A., Andover
<4>0
<5>169-178
<6>1993
<7>Although a number of bacterial species are naturally transformable, that is, their cells are
able to take up external DNA in substantial amounts and integrate it into the chromosome
without artificial manipulation of the cell surface, Streptococcus pneumoniae, the first
species in which this phenomenon was detected, remains a prototype of such transformation.
This is partly because these bacteria do not appear to use other forms of genetic exchange for
transfer of chromosomal genes, such as conjugation and transduction, but rely solely on
DNA-mediated transformation.  Cells of S. pneumoniae also contain potent restriction
endonucleases able to severely restrict DNA introduced during viral infection.  Therefore, it
should be interesting to examine the effects of the restriction enzyme systems on transforming
DNA.  Our current understanding of the genetic basis of the complementary DpnI and DpnII
restriction systems and of the biochemistry of their component enzymes will be briefly
reviewed.  The manner in which these enzymes impinge on the transfer of chromosomal genes and
of plasmids will be examined in detail.  It will be seen that far from acting against
"foreign" DNA in general, the restriction systems seem to be designed to exclude only
infecting viral DNA.  The presence of complementary restriction systems in different cells of
S. pneumoniae enhances their effectiveness in blocking viral infection and promoting species
survival.  This enhanced effectiveness requires the expression of alternative restriction
systems.  Therefore, the ability of the cells to transfer the restriction enzyme genes and to
regulate their expression are important for survival of the species.  The final part of this
paper will present currently available information on this topic.  In particular, the
localization of the restriction genes in cassettes, their transcription products, and the role
of a possibly new class of ribosome binding sites will be examined in relation to the
regulation of restriction gene expression.

<>

<1>Lacks, S.A., Springhorn, S.S.
<2>Transfer of recombinant plasmids containing the gene for DpnII DNA methylase into strains of Streptococcus pneumoniae that produce DpnI or DpnII restriction endonucleases.
<3>J. Bacteriol.
<4>158
<5>905-909
<6>1984
<7>Plasmid transfer via the transformation pathway of Streptococcus pneumoniae was
weakly restricted by the DpnI or DpnII restriction endonuclease, either of
which gave a reduction only to 0.4, compared with phage infection, which was
restricted to 10^-5.  The greater sensitivity of plasmid transfer compared with
chromosomal transformation, which was not at all restricted, can be attributed
to partially double-stranded intermediates formed from two complementary donor
fragments.  However, clustering of potential restriction sites in the plasmids
increased the probability of escape from restriction.  The recombinant plasmid
pMP10, in which the gene for the DpnII DNA methylase was cloned, can be
transferred to strains that contain neither restriction enzyme or that contain
DpnII as readily as can the vector pMP5.  Introduction of pMP10 raised the
level of methylase by five times the level normally present in DpnII strains.
Transfer of pMP10 to DpnI-containing strains was infrequent, presumably owing
to the suicidal methylation of DNA which rendered it susceptible to the host
endonuclease.  The few clones in which pMP10 was established had lost DpnI.
Loss of the plasmid after curing of the cell eliminated the methylase but did
not restore DpnI.  Although this loss of DpnI could result from spontaneous
mutation, its relatively high frequency, 0.1% suggested that the loss was due
to a regulatory shift.

<>

<1>Lacks, S.A., Springhorn, S.S.
<2>Cloning in Streptococcus pneumoniae of the gene for DpnII DNA methylase.
<3>J. Bacteriol.
<4>157
<5>934-936
<6>1984
<7>The gene coding for the pneumococcal DNA adenine methylase that recognizes the
sequence 5'-GATC-3' was cloned in a strain of Streptococcus pneumoniae that
lacked both restriction endonucleases DpnI and DpnII.  The gene was cloned as a
3.7-kilobase fragment of chromosomal DNA from a DpnII-containing strain
inserted in both possible orientations in the multicopy plasmid vector pMP5 to
give recombinant plasmids pMP8 and pMP10.  Recombinant plasmids were selected
by their resistance to DpnII cleavage.  Cells carrying the recombinant plasmids
modified phage in vivo so that it was restricted by DpnI-but not
DpnII-containing hosts.  They also showed levels of DNA methylase activity five
times higher than that in cells of the original DpnII strain.  No DpnII
activity was observed in the clones; therefore, it was concluded that the
insert did not contain an intact DpnII endonuclease gene and that methylation
of host DNA did not turn on a latent form of the gene.

<>

<1>Lacks, S.A., Springhorn, S.S., Cerritelli, S.
<2>Restriction/Modification systems of Pneumococci:  Why two methylases in the DpnII system?
<3>Genetics and Molecular Biology of Streptococci, Lactococci, and Enterococci, American Society for Microbiology, Dunny, G.M., Cleary, P.P., McKay, L.L., Washington
<4>8
<5>71-76
<6>1991
<7>None

<>

<1>Ladero, V., Alvarez-Sieiro, P., Redruello, B., Del Rio, B., Linares, D.M., Martin, M.C., Fernandez, M., Alvarez, M.A.
<2>Draft Genome Sequence of Lactobacillus plantarum Strain IPLA 88.
<3>Genome Announcements
<4>1
<5>e00524-13
<6>2013
<7>Here, we report a 3.2-Mbp draft assembly for the genome of Lactobacillus plantarum IPLA 88.
The sequence of this sourdough isolate provides insight into
the adaptation of this versatile species to different environments.

<>

<1>Ladero, V., Del Rio, B., Linares, D.M., Fernandez, M., Mayo, B., Martin, M.C., Alvarez, M.A.
<2>Genome Sequence Analysis of the Biogenic Amine-Producing Strain Lactococcus lactis subsp. cremoris CECT 8666 (Formerly GE2-14).
<3>Genome Announcements
<4>2
<5>e01088-14
<6>2014
<7>We here report a 2,801,031-bp annotated draft assembly for the Lactococcus lactis subsp.
cremoris GE2-14 genome. This dairy strain produces the biogenic amine
putrescine. This sequence may help identify the mechanisms regulating putrescine
biosynthesis and throw light on ways to reduce its presence in fermented foods.

<>

<1>Ladero, V., Herrero-Fresno, A., Martinez, N., Del Rio, B., Linares, D.M., Fernandez, M., Martin, M.C., Alvarez, M.A.
<2>Genome Sequence Analysis of the Biogenic Amine-Degrading Strain Lactobacillus casei 5b.
<3>Genome Announcements
<4>2
<5>e01199-13
<6>2014
<7>We here report a 3.02-Mbp annotated draft assembly of the Lactobacillus casei 5b  genome. The
sequence of this biogenic amine-degrading dairy isolate may help
identify the mechanisms involved in the catabolism of biogenic amines and perhaps
shed light on ways to reduce the presence of these toxic compounds in food.

<>

<1>Ladero, V., Linares, D.M., Del Rio, B., Fernandez, M., Martin, M.C., Alvarez, M.A.
<2>Draft Genome Sequence of the Tyramine Producer Enterococcus durans Strain IPLA 655.
<3>Genome Announcements
<4>1
<5>e00265-13
<6>2013
<7>We here report a 3.059-Mbp draft assembly for the genome of Enterococcus durans strain IPLA
655. This dairy isolate provides a model for studying the regulation
of the biosynthesis of tyramine (a toxic compound). These results should aid our
understanding of tyramine production and allow tyramine accumulation in food to
be reduced.

<>

<1>Ladner, J.T., Welch, T.J., Whitehouse, C.A., Palacios, G.F.
<2>Genome Sequence of Weissella ceti NC36, an Emerging Pathogen of Farmed Rainbow Trout in the United States.
<3>Genome Announcements
<4>1
<5>e00187-12
<6>2013
<7>Novel Weissella sp. bacteria have recently been reported to be associated with disease
outbreaks in cultured rainbow trout (Oncorhynchus mykiss) at commercial
farms in China, Brazil, and the United States. Here we present the first genome
sequence of this novel Weissella species, isolated from the southeastern United
States.

<>

<1>Ladner, J.T., Whitehouse, C.A., Koroleva, G.I., Palacios, G.F.
<2>Genome Sequence of Moraxella macacae 0408225, a Novel Bacterial Species Isolated  from a Cynomolgus Macaque with Epistaxis.
<3>Genome Announcements
<4>1
<5>e00188-12
<6>2013
<7>Moraxella macacae is a recently described bacterial species that has been associated with at
least two outbreaks of epistaxis in macaques. Here we present
the first genome sequence of this novel species, isolated from a symptomatic
cynomolgus macaque at the U.S. Army Medical Research Institute of Infectious
Diseases.

<>

<1>Lagace, R.E., Patterson, C., Berg, K.L.
<2>Nucleic acids encoding 3-ketoacyl-Acp reductase from Moraxella catarrahalis.
<3>US Patent Office
<4>US 6632636 B
<5>
<6>2003
<7>The present invention provides the genomic sequences of a library of purified nucleic acid
molecules, or their complements, comprising the genome of Moraxella catarrhalis.  The
invention also provides the identification of open reading frames contained within the nucleic
acid molecules of the library.  The present invention further provides for the use of the
nucleic acid molecules, their complements or fragments, and proteins or portions thereof for
identifying ligands and useful diagnostic and therapeutic compositions.  In addition the
invention provides for vectors, host cells and methods for producing M. catarrhalis proteins
or portions thereof.

<>

<1>Laganeckas, M., Margelevicius, M., Venclovas, C.
<2>Identification of new homologs of PD-(D/E)XK nucleases by support vector machines trained on data derived from profile-profile alignments.
<3>Nucleic Acids Res.
<4>39
<5>1187-1196
<6>2011
<7>PD-(D/E)XK nucleases, initially represented by only Type II restriction enzymes, now comprise
a large and extremely diverse superfamily of
proteins. They participate in many different nucleic acids transactions
including DNA degradation, recombination, repair and RNA processing.
Different PD-(D/E)XK families, although sharing a structurally conserved
core, typically display little or no detectable sequence similarity except
for the active site motifs. This makes the identification of new
superfamily members using standard homology search techniques challenging.
To tackle this problem, we developed a method for the detection of
PD-(D/E)XK families based on the binary classification of profile-profile
alignments using support vector machines (SVMs). Using a number of both
superfamily-specific and general features, SVMs were trained to identify
true positive alignments of PD-(D/E)XK representatives. With this method
we identified several PFAM families of uncharacterized proteins as
putative new members of the PD-(D/E)XK superfamily. In addition, we
assigned several unclassified restriction enzymes to the PD-(D/E)XK type.
Results show that the new method is able to make confident assignments
even for alignments that have statistically insignificant scores. We also
implemented the method as a freely accessible web server at
http://www.ibt.lt/bioinformatics/software/pdexk/.

<>

<1>Lagier, J.C., Armougom, F., Mishra, A.K., Nguyen, T.T., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Alistipes timonensis sp. nov.
<3>Standards in Genomic Sciences
<4>6
<5>315-324
<6>2012
<7>Alistipes timonensis strain JC136(T) sp. nov. is the type strain of A. timonensis sp. nov., a
new species within the genus Alistipes. This strain, whose genome is
described here, was isolated from the fecal flora of a healthy patient. A.
timonensis is an obligate anaerobic rod. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
3,497,779 bp long genome (one chromosome but no plasmid) contains 2,742
protein-coding and 50 RNA genes, including three rRNA genes.

<>

<1>Lagier, J.C., Bibi, F., Ramasamy, D., Azhar, E.I., Robert, C., Yasir, M., Jiman-Fatani, A.A., Alshali, K.Z., Fournier, P.E., Raoult, D.
<2>Non contiguous-finished genome sequence and description of Clostridium jeddahense sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1003-1019
<6>2014
<7>Clostridium jeddahense strain JCD(T) (= CSUR P693 = DSM 27834) is the type strain of C.
jeddahense sp. nov. This strain, whose genome is described here, was
isolated from the fecal flora of an obese 24 year-old Saudian male (BMI=52
kg/m(2)). Clostridium jeddahense strain JCD(T) is an obligate Gram-positive
bacillus. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 3,613,503 bp long genome (1
chromosome, no plasmid) exhibits a G+C content of 51.95% and contains 3,462
protein-coding and 53 RNA genes, including 4 rRNA genes.

<>

<1>Lagier, J.C., El Karkouri, K., Mishra, A.K., Robert, C., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Enterobacter massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>399-412
<6>2013
<7>Enterobacter massiliensis strain JC163(T) sp. nov. is the type strain of E. massiliensis sp.
nov., a new species within the genus Enterobacter. This strain,
whose genome is described here, was isolated from the fecal flora of a healthy
Senegalese patient. E. massiliensis is an aerobic rod. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. The 4,922,247 bp long genome (1 chromosome but no plasmid) exhibits a
G+C content of 55.1% and contains 4,644 protein-coding and 80 RNA genes,
including 5 rRNA genes.

<>

<1>Lagier, J.C., El Karkouri, K., Nguyen, T.T., Armougom, F., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Anaerococcus senegalensis sp. nov.
<3>Standards in Genomic Sciences
<4>6
<5>116-125
<6>2012
<7>Anaerococcus senegalensis strain JC48T sp. nov. is the type strain of A. senegalensis sp.
nov., a new species within the genus Anaerococcus. This strain, whose genome is described
here, was isolated from the fecal flora of a healthy patient. A. senegalensis is an obligate
anaerobic coccus. Here we describe the features of this organism, together with the complete
genome sequence and annotation. The 1,790,835 bp long genome (1 chromosome but no plasmid)
contains 1,721 protein-coding and 53 RNA genes, including 5 rRNA genes.

<>

<1>Lagier, J.C., Elkarkouri, K., Rivet, R., Couderc, C., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Senegalemassilia anaerobia gen. nov., sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>343-356
<6>2013
<7>Senegalemassilia anaerobia strain JC110(T) sp.nov. is the type strain of Senegalemassilia
anaerobia gen. nov., sp. nov., the type species of a new genus
within the Coriobacteriaceae family, Senegalemassilia gen. nov. This strain,
whose genome is described here, was isolated from the fecal flora of a healthy
Senegalese patient. S. anaerobia is a Gram-positive anaerobic coccobacillus. Here
we describe the features of this organism, together with the complete genome
sequence and annotation. The 2,383,131 bp long genome contains 1,932
protein-coding and 58 RNA genes.

<>

<1>Lagier, J.C., Gimenez, G., Robert, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Herbaspirillum massiliense sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>200-209
<6>2012
<7>Herbaspirillum massiliense strain JC206(T) sp. nov. is the type strain of H. massiliense sp.
nov., a new species within the genus Herbaspirillum. This strain,
whose genome is described here, was isolated from the fecal flora of a healthy
Senegalese patient. H. massiliense is an aerobic rod. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. The 4,186,486 bp long genome (one chromosome but no plasmid) contains
3,847 protein-coding and 54 RNA genes, including 3 rRNA genes.

<>

<1>Lagier, J.C., Ramasamy, D., Rivet, R., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Cellulomonas massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>258-270
<6>2012
<7>Cellulomonas massiliensis strain JC225(T) sp. nov. is the type strain of Cellulomonas
massiliensis sp., a new species within the genus Cellulomonas. This
strain, whose genome is described here, was isolated from the fecal flora of a
healthy Senegalese patient. C. massiliensis is an aerobic rod-shaped bacterium.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. The 3,407,283 bp long genome contains 3,083
protein-coding and 48 RNA genes.

<>

<1>Laging, M., Lindner, E., Fritz, H.-J., Kramer, W.
<2>Repair of hydrolytic DNA deamination damage in thermophilic bacteria: cloning and characterization of a Vsr endonuclease homolog from Bacillus stearothermophilus.
<3>Nucleic Acids Res.
<4>31
<5>1913-1920
<6>2003
<7>Hydrolytic deamination of 5-methyl cytosine in double stranded DNA results in formation of a
T/G mismatch that-if left unrepaired-leads to a C-->T
transition mutation in half of the progeny. In addition to several
mismatch-specific glycosylases that have been found in both pro- and
eukaryotes to channel this lesion into base excision repair by removing
the T from the mismatch, Vsr endonuclease from Escherichia coli has been
described which initiates repair by an endonucleolytic strand incision 5'
to the mismatched T. We have isolated a gene coding for a homolog of
E.coli Vsr endonuclease from the thermophilic bacterium Bacillus
stearothermophilus H3 (Vsr.Bst) using a method that allows PCR
amplification with degenerated primers of gene segments which code for
only one highly conserved amino acid region. Vsr.Bst was produced
heterologously in E.coli and purified to apparent homogeneity. Vsr.Bst
specifically incises heteroduplex DNA with a preference for T/G
mismatches. The selectivity of Vsr.Bst for the sequence context of the T/G
mismatch appears less pronounced than for Vsr.Eco.

<>

<1>Laguerre, S., Amari, M., Vuillemin, M., Robert, H., Loux, V., Klopp, C., Morel, S., Gabriel, B., Remaud-Simeon, M., Gabriel, V., Moulis, C., Fontagne-Faucher, C.
<2>Genome Sequences of Three Leuconostoc citreum Strains, LBAE C10, LBAE C11, and LBAE E16, Isolated from Wheat Sourdoughs.
<3>J. Bacteriol.
<4>194
<5>1610-1611
<6>2012
<7>Leuconostoc citreum is a key microorganism in fermented foods of plant origin. Here we report
the draft genome sequence for three strains of Leuconostoc
citreum, LBAE C10, LBAE C11, and LBAE E16, which have been isolated from
traditional French wheat sourdoughs.

<>

<1>Lagunavicius, A.
<2>DNA binding by MunI restriction endonuclease.
<3>Biologija
<4>1
<5>13-20
<6>1997
<7>Investigation of DNA binding properties by gel shift assay indicated that in the absence of
cofactor at pH 8.0, WT MunI binds to DNA containing or lacking the recognition sequence with
similar affinity and only at a high excess of protein.  Contrary to the WT enzyme, E98A and
D83A mutants under the same conditions bound DNA containing the recognition sequence with high
affinity.  With noncognate DNA the latter mutants exhibited only a weak binding characteristic
of the WT MunI.  The increased affinity of the mutants to the cognate DNA suggest that E98A
and D83A replacements can mimic the effect of Mg2+ ion during the development of the
specificity of MunI restriction enzyme at the catalytic step.  An increased affinity of the WT
MunI to the cognate DNA was observed also at low pH or in the presence Ca2+ ions.  Indeed, in
contrast to the binding experiments at pH 8.0, gel-shift analysis at pH 6.5 indicated a tight
sequence-specific binding of WT MunI to the cognate DNA suggesting that the protonation of the
active site carboxylate residue(s) with anomalous high pKa value control the binding
specificity.  Interestingly, Ca2+ ions that did not support DNA cleavage by MunI were able to
induce DNA binding specificity of WT MunI at pH 8.0 and probably mimic the role of Mg2+ ions
in the development of sequence specific binding.  The relief of unfavorable repulsive
constraints either by replacement of carboxylate(s) lowering the pH or metal ion binding leads
to the manifestation of binding specificity by MunI.

<>

<1>Lagunavicius, A., Grazulis, S., Balciunaite, E., Vainius, D., Siksnys, V.
<2>DNA binding specificity of MunI restriction endonuclease is controlled by pH and calcium ions: Involvement of active site carboxylate residues.
<3>Biochemistry
<4>36
<5>11093-11099
<6>1997
<7>Gel shift analysis reveals that at pH 8.3 in the absence of Mg2+, MunI restriction
endonuclease exhibits little DNA binding specificity, as compared with the D83A and E98A
mutants of MunI.  This suggests that charged carboxylate residue(s) influence the DNA binding
specificity of MunI.  In our efforts to establish the determinants of MunI binding
specificity, we investigated the possible role of the ionic milieu, and we found that lowering
pH or elevating Ca2+ levels per se induces specific DNA recognition by WT MunI.  In contrast
to the binding experiments at pH 8.3, gel shift analysis at pH 6.5 indicated tight
sequence-specific binding of WT MunI in the absence of Mg2+, suggesting that protonation of
active site carboxylate residue(s) which manifest anomalously high pKa value(s) control
binding specificity.  Interestingly, Ca2+ ion concentrations, which did not support DNA
cleavage by MunI also induced DNA binding specificity in WT MunI at pH 8.3.  To explore
possible structural changes upon DNA binding, we then used a limited proteolysis technique.
Trypsin cleavage of MunI--DNA complexes indicated that in the presence of cognate DNA the MunI
restriction endonuclease became resistant to proteolytic cleavage, suggesting that binding of
specific DNA induced a structural change.  CD measurements confirmed this observation,
suggesting minor secondary structural differences between complexes of MunI with cognate and
noncognate DNA. These results therefore suggest that binding of MunI to its recognition
sequence triggers a conformational transition that correctly juxtaposes active site
carboxylate residues, which then chelate Mg2+ ions.  In the absence of Mg2+ ions, at pH 8.3,
conditions in which carboxylate groups would be expected to be completely ionized,
electrostatic repulsion between charged carboxylates and phosphate oxygens is enhanced such as
to interfere with specific DNA binding.  Elimination of such repulsive constraints by
replacement of carboxylate residues, by lowering pH, or by metal ion binding, then promotes
MunI binding specificity.

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<1>Lagunavicius, A., Sasnauskas, G., Halford, S.E., Siksnys, V.
<2>The metal-independent type IIS restriction enzyme BfiI is a dimer that binds two DNA sites but has only one catalytic centre.
<3>J. Mol. Biol.
<4>326
<5>1051-1064
<6>2003
<7>BfiI is a novel type IIs restriction endonuclease that, unlike all other restriction enzymes
characterised to date, cleaves DNA in the absence of
Mg(2+). The amino acid sequence of the N-terminal part of BfiI has some
similarities to Nuc of Salmonella typhimurium, an EDTA-resistant nuclease
akin to phospholipase D. The dimeric form of Nuc contains a single active
site composed of residues from both subunits. To examine the roles of the
amino acid residues of BfiI that align with the catalytic residues in Nuc,
a set of alanine replacement mutants was generated by site-directed
mutagenesis. The mutationally altered forms of BfiI were all catalytically
inactive but were still able to bind DNA specifically. The active site of
BfiI is thus likely to be similar to that of Nuc. BfiI was also found by
gel-filtration to be a dimer in solution. Both gel-shift and pull-down
assays indicated that the dimeric form of BfiI binds two copies of its
recognition sequence. In reactions on plasmids with either one or two
copies of its recognition sequence, BfiI cleaved the DNA with two sites
more rapidly than that with one site. Yet, when bound to two copies of its
recognition sequence, the BfiI dimer cleaved only one phosphodiester bond
at a time. The dimer thus seems to contain two DNA-binding domains but
only one active site.

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<1>Lagunavicius, A., Siksnys, V.
<2>Mutational analysis of MunI restriction endonuclease.
<3>Biologija
<4>0
<5>35-38
<6>1996
<7>Mapping of the conserved sequence regions between MunI and EcoRI restriction endonucleases to
the known X-ray structure of the EcoRI restriction enzyme allowed us to identify a sequence
motif 82PDX14EXK as a putative catalytic site of MunI.  Site-directed mutagenesis was used to
test whether amino acids P82, D83, E98 and K100 were important for catalytic activity of MunI.
A set of MunI mutants (P82A, D83A, E98Q, E98Q, E98A, K100E, K100A) altered at the putative
catalytic site was obtained using site-directed mutagenesis, and the catalytic properties of
the mutants were studied in vivo and in vitro.  The phenotypes observed in vivo and in vitro
for the mutants of putative catalytic site residues of MunI were consistent with the proposed
active site function.  Investigation of the cleavage properties of these mutants revealed that
E98Q replacement in MunI resulted in alteration of metal ion binding.  Contrary to the wild
type enzyme, this mutant was activated by Mn2+ ions more effectively than by Mg2+ ions.

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<1>Lagunavicius, A., Siksnys, V.
<2>Site-directed mutagenesis of putative active site residues of MunI restriction endonuclease: replacement of catalytically essential carboxylate residues triggers DNA binding specificity.
<3>Biochemistry
<4>36
<5>11086-11092
<6>1997
<7>Mapping of the conserved sequence regions in the restriction endonucleases MunI (C/AATTG) and
EcoRI (G/AATTC) to the known X-ray structure of EcoRI allowed us to identify the sequence
motif 82PDX14EXK as the putative catalytic/Mg2+ ion binding site of MunI.  Site-directed
mutagenesis was then used to test whether amino acids P82, D83, E98, and K100 were important
for the catalytic activity of MunI.  Mutation P82A generated only a marginal effect on the
cleavage properties of the enzyme.  Investigation of the cleavage properties of the D83, E98,
and K100 substitution mutants, however, in vivo and in vitro, revealed either an absence of
catalytic activity or markedly reduced catalytic activity.  Interestingly, the deleterious
effect of the E98Q replacement in vitro was partially overcome by replacement of the metal
cofactor used.  Though the catalytic activity of the E98Q mutant was only 0.4% of WT under
standard conditions (in the presence of Mg2+ ions), the mutant exhibited 40% of WT catalytic
activity in buffer supplemented with Mn2+ ions.  Further, the DNA binding properties of these
substitution mutants were analyzed using the gel shift assay technique.  In the absence of
Mg2+ ions, WT MunI bound both cognate DNA and noncognate sequences with similar low
affinities.  The D83A and E98A mutants, in contrast, in the absence of Mg2+ ions, exhibited
significant specificity of binding to cognate DNA, suggesting that the substitutions made can
simulate the effect of the Mg2+ ion in conferring specificity to the MunI restriction enzyme.

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<1>Lai, C., Wu, X., Chen, C., Huang, T., Xiong, X., Wu, S., Gu, M., Deng, Z., Chen, Xi., Chen, S., Wang, L.
<2>In Vivo Mutational Characterization of DndE Involved in DNA Phosphorothioate Modification.
<3>PLoS ONE
<4>9
<5>e107981
<6>2014
<7>DNA phosphorothioate (PT) modification is a recently identified epigenetic modification that
occurs in the sugar-phosphate backbone of prokaryotic DNA. Previous studies have demonstrated
that DNA PT modification is governed by the five DndABCDE proteins in a sequence-selective and
R-P stereo-specific manner. Bacteria may have acquired this physiological modification along
with dndFGH as a restriction-modification system. However, little is known about the
biological function of Dnd proteins, especially the smallest protein, DndE, in the PT
modification pathway. DndE was reported to be a DNA-binding protein with a preference for
nicked dsDNA in vitro; the binding of DndE to DNA occurs via six positively charged lysine
residues on its surface. The substitution of these key lysine residues significantly decreased
the DNA binding affinities of DndE proteins to undetectable levels. In this study, we
conducted site-directed mutagenesis of dndE on a plasmid and measured DNA PT modifications
under physiological conditions by mass spectrometry. We observed distinctive differences from
the in vitro binding assays. Several mutants with lysine residues mutated to alanine decreased
the total frequency of PT modifications, but none of the mutants completely eliminated PT
modification. Our results suggest that the nicked dsDNA-binding capacity of DndE may not be
crucial for PT modification and/or that DndE may have other biological functions in addition
to binding to dsDNA.

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<1>Lai, J.Y., Zhang, H., Chiang, M.H., Yu, M., Zhang, R., Lau, S.C.
<2>Draft Genome Sequence of Escherichia coli E1728 Isolated from Marine Sediment in  Hong Kong.
<3>Genome Announcements
<4>2
<5>e00430-14
<6>2014
<7>Recent findings of Escherichia coli persisting autochthonously in environmental matrices
outside animal bodies have revealed largely unknown facets of the
lifestyle and ecophysiology of the species that have yet to be explored. Here, we
report the draft genome sequence of E. coli E1728 isolated from marine sediment.

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<1>Lai, J.Y., Zhang, H., Chiang, M.H., Yu, M., Zhang, R., Lau, S.C.
<2>Draft Genome Sequences of Three Escherichia coli Strains Investigated for the Effects of Lysogeny on Niche Diversification.
<3>Genome Announcements
<4>2
<5>e00955-14
<6>2014
<7>During the course of investigating the effects of lysogeny on niche diversification of
Escherichia coli, we used the temperate phages induced from
one E. coli strain to infect another and created an isogenic lysogen of the
latter. The draft genome sequences of the three E. coli strains are reported
herein.

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<1>Lai, Q., Li, C., Shao, Z.
<2>Genome Sequence of Galbibacter marinum Type Strain ck-I2-15.
<3>J. Bacteriol.
<4>194
<5>6973
<6>2012
<7>Galbibacter marinum strain ck-I2-15(T) was isolated from an arsenite-resistant consortium
enriched from the deep sea sediment of a hydrothermal vent field on
the Southwest Indian Ocean Ridge. Here, we present the draft genome of strain
ck-I2-15(T), which contains 3,572,447 bp with a G+C content of 37.04% and
contains 3,099 protein-coding genes and 38 tRNA genes.

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<1>Lai, Q., Li, G., Shao, Z.
<2>Genome Sequence of Nitratireductor pacificus Type Strain pht-3B.
<3>J. Bacteriol.
<4>194
<5>6958
<6>2012
<7>Nitratireductor pacificus strain pht-3B(T) was isolated from a pyrene-degrading consortium
enriched from the deep sea sediment of the Pacific Ocean. Here, we
present the draft genome of strain pht-3B(T), which contains 4,466,205 bp with a
G+C content of 65.51% and contains 4,197 protein-coding genes and 46 tRNA genes.

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<1>Lai, Q., Li, G., Yu, Z., Shao, Z.
<2>Genome Sequence of Nitratireductor indicus Type Strain C115.
<3>J. Bacteriol.
<4>194
<5>6990
<6>2012
<7>Nitratireductor indicus strain C115(T) was isolated from a crude-oil-degrading consortium
enriched from deep seawater of the Indian Ocean. Here, we present the
draft genome of strain C115(T), which contains 4,992,479 bp with a G+C content of
60.8% and contains 4,825 protein-coding genes and 45 tRNA genes.

<>

<1>Lai, Q., Li, W., Shao, Z.
<2>Complete Genome Sequence of Alcanivorax dieselolei Type Strain B5.
<3>J. Bacteriol.
<4>194
<5>6674
<6>2012
<7>Alcanivorax dieselolei B5(T) was isolated from oil-contaminated surface water of  the Bohai
Sea of China and characterized by the efficient degradation of alkane
(C(5)-C(36)). Here we report the complete genome of B5(T) and genes associated
with alkane degradation.

<>

<1>Lai, Q., Li, W., Wang, B., Yu, Z., Shao, Z.
<2>Complete Genome Sequence of the Pyrene-Degrading Bacterium Cycloclasticus sp. Strain P1.
<3>J. Bacteriol.
<4>194
<5>6677
<6>2012
<7>Cycloclasticus sp. strain P1 was isolated from deep-sea sediments of the Pacific  Ocean and
characterized as a unique bacterium in the degradation of pyrene, a
four-ring polycyclic aromatic hydrocarbon (PAH). Here we report the complete
genome of P1 and genes associated with PAH degradation.

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<1>Lai, Q., Liu, Y., Shao, Z.
<2>Genome Sequence of Bacillus sp. Strain HYC-10, Isolated from Intestinal Tract Contents from a Marine Fish (Mugil cephalus).
<3>J. Bacteriol.
<4>194
<5>6991
<6>2012
<7>Bacillus sp. strain HYC-10 was isolated with intestinal tract content of a fish,  Mugil
cephalus, captured from the sea close to Xiamen Island, China. Here, we
present the draft genome of strain HYC-10, which contains 3,611,918 bp with a G+C
content of 41.30% and contains 3,687 protein-coding genes and 33 tRNA genes.

<>

<1>Lai, Q., Shao, Z.
<2>Genome Sequence of Thalassospira profundimaris Type Strain WP0211.
<3>J. Bacteriol.
<4>194
<5>6956
<6>2012
<7>Thalassospira profundimaris WP0211(T) was isolated from a pyrene-degrading consortium,
enriched from deep-sea sediment collected from the West Pacific
Ocean. Here, we present the draft genome of strain WP0211(T), which contains
4,380,232 bp with a G+C content of 55.19% and contains 4,040 protein-coding genes
and 45 tRNAs.

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<1>Lai, Q., Shao, Z.
<2>Genome Sequence of Thalassospira xiamenensis Type Strain M-5.
<3>J. Bacteriol.
<4>194
<5>6957
<6>2012
<7>Thalassospira xiamenensis M-5(T) was isolated from the surface water of a waste oil pool at
the oil storage dock in the city of Xiamen, Fujian Province, China.
Here, we present the draft genome of strain M-5(T), which contains 4,705,237 bp
with a G+C content of 54.65% and contains 4,343 protein-coding genes and 46 tRNA
genes.

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<1>Lai, Q., Shao, Z.
<2>Genome Sequence of the Alkane-Degrading Bacterium Alcanivorax hongdengensis Type  Strain A-11-3.
<3>J. Bacteriol.
<4>194
<5>6972
<6>2012
<7>Alcanivorax hongdengensis A-11-3(T) was isolated from an oil-enriched consortium  enriched
from the surface seawater of Hong-Deng dock in the Straits of Malacca
and Singapore. Strain A-11-3(T) can degrade n-alkane and produce a lipopeptide
biosurfactant. Here we report the genome of A-11-3(T) and the genes associated
with alkane degradation.

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<1>Lai, Q., Shao, Z.
<2>Genome Sequence of an Alkane-Degrading Bacterium, Alcanivorax pacificus Type Strain W11-5, Isolated from Deep Sea Sediment.
<3>J. Bacteriol.
<4>194
<5>6936
<6>2012
<7>Alcanivorax pacificus W11-5(T) was isolated from a pyrene-degrading consortium, enriched from
the deep sea sediment of the Pacific Ocean. Strain W11-5(T) can
degrade various n-alkanes. Here we report the draft genome of W11-5(T) and genes
associated with alkane degradation.

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<1>Lai, Q., Shao, Z.
<2>Genome Sequence of Oceanibaculum indicum Type Strain P24.
<3>J. Bacteriol.
<4>194
<5>6942
<6>2012
<7>Oceanibaculum indicum type strain P24 was isolated from a
polycyclic-aromatic-hydrocarbon-degrading consortium enriched from a
deep-seawater sample collected from the Indian Ocean. Here we present the draft
genome of strain P24(T), which contains 3,952,792 bp with a G+C content of 65.5%
and contains 3,755 protein-coding genes and 45 tRNAs.

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<1>Lai, Q., Wang, L., Wang, W., Shao, Z.
<2>Genome Sequence of Gallaecimonas xiamenensis Type Strain 3-C-1.
<3>J. Bacteriol.
<4>194
<5>6937
<6>2012
<7>Gallaecimonas xiamenensis 3-C-1(T) was isolated from a crude-oil-degrading consortium enriched
from the surface seawater around Xiamen Island. Here, we
present the draft genome of strain 3-C-1(T), which contains 4,062,282 bp with a
G+C content of 60.58% and contains 3,798 protein-coding genes and 65 tRNAs.

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<1>Lai, Q., Wang, L., Wang, W., Shao, Z.
<2>Genome Sequence of Idiomarina xiamenensis Type Strain 10-D-4.
<3>J. Bacteriol.
<4>194
<5>6938
<6>2012
<7>Idiomarina xiamenensis strain 10-D-4(T) was isolated from an oil-degrading consortium enriched
from surface seawater around the Xiamen island. Here, we
present the draft genome of strain 10-D-4(T), which contains 2,899,282 bp with a
G+C content of 49.48% and contains 2,673 protein-coding genes and 43 tRNA genes.

<>

<1>Lai, Y.C., Yang, S.L., Peng, H.L., Chang, H.Y.
<2>Identification of genes present specifically in a virulent strain of Klebsiella pneumoniae.
<3>Infect. Immun.
<4>68
<5>7149-7151
<6>2000
<7>Klebsiella pneumoniae is a common cause of septicemia and urinary tract infections. The
PCR-supported genomic subtractive hybridization was
employed to identify genes specifically present in a virulent strain of K.
pneumoniae. Analysis of 25 subtracted DNA clones has revealed 19 distinct
nucleotide sequences. Two of the sequences were found to be the genes
encoding the transposase of Tn3926 and a capsule polysaccharide exporting
enzyme. Three sequences displayed moderate homology with bvgAS, which
encodes a two-component signal transduction system in Bordetella
pertussis. The rest of the sequences did not exhibit homology with any
known genes. The distribution of these novel sequences varied greatly in
K. pneumoniae clinical isolates, reflecting the heterogeneous nature of
the K. pneumoniae population.

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<1>Laigret, F., Gaurivaud, P., Bove, J.-M.
<2>The unique organization of the rpoB region of Spiroplasma citri: a restriction and modification system gene is adjacent to rpoB.
<3>Gene
<4>171
<5>95-98
<6>1996
<7>A 6.5-kb DNA fragment containing the gene (rpoB) encoding the RNA polymerase (RNAP) beta
subunit, from the mollicute Spiroplasma citri (Sc), was cloned and sequenced.  The classical
eubacterial organization, with the genes (rplK, A, J and L) encoding ribosomal proteins L11,
L1, L10 and L12 located immediately upstream from rpoB, was not found in the Sc DNA.  Instead,
an open reading frame (hsdS) potentially encoding a component of a type I restriction and
modification system was identified upstream from rpoB, and sequences showing similarities with
insertion elements were found between hsdS and rpoB.

<>

<1>Lail, K. et al.
<2>Complete genome sequence of Spirosoma linguale type strain (1).
<3>Standards in Genomic Sciences
<4>2
<5>176-185
<6>2010
<7>Spirosoma linguale Migula 1894 is the type species of the genus. S. linguale is a free-living
and non-pathogenic organism, known for its peculiar ringlike and
horseshoe-shaped cell morphology. Here we describe the features of this organism,
together with the complete genome sequence and annotation. This is only the third
completed genome sequence of a member of the family Cytophagaceae. The 8,491,258
bp long genome with its eight plasmids, 7,069 protein-coding and 60 RNA genes is
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Lainson, F.A., Dagleish, M.P., Fontaine, M., Bayne, C., Hodgson, J.C.
<2>Draft Genome Sequences of Strains of Pasteurella multocida Isolated from the United Kingdom and the United States.
<3>Genome Announcements
<4>1
<5>e00773-13
<6>2013
<7>Pasteurella multocida is a major pathogen of farm animals and has worldwide distribution. Here
we report the draft genome sequences of four strains that were
isolated from animals in the United Kingdom and the United States and represent
pathogenic and commensal presentation of the bacterium.

<>

<1>Lainson, F.A., Dagleish, M.P., Fontaine, M.C., Bayne, C., Hodgson, J.C.
<2>Draft Genome Sequence of Pasteurella multocida A:3 Strain 671/90.
<3>Genome Announcements
<4>1
<5>e00803-13
<6>2013
<7>Pasteurella multocida serogroup A is commonly isolated from nasal swabs of clinically healthy
calves and also from diseased lung tissue in bovine pneumonia.
Here, we report the draft genome sequence of the virulent strain P. multocida
671/90, which has been characterized previously in experimental infections of
calves and mice.

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<1>Laird, P.W., Jackson-Grusby, L., Fazeli, A., Dickinson, S.L., Jung, W.E., Li, E., Weinberg, R.A., Jaenisch, R.
<2>Suppression of intestinal neoplasia by DNA hypomethylation.
<3>Cell
<4>81
<5>197-205
<6>1995
<7>We have used a combination of genetics and pharmacology to assess the effects of reduced DNA
methyltransferase activity on ApcMin-induced intestinal neoplasia in mice. A reduction in the
DNA methyltransferase activity in Min mice due to heterozygosity of the DNA methyltransferase
gene, in conjunction with a weekly dose of the DNA methyltransferase inhibitor
5-aza-deoxycytidine, reduced the average number of intestinal adenomas from 113 in the control
mice to only 2 polyps in the treated heterozygotes. Hence, DNA methyl-transferase activity
contributes substantially to tumor developments in this mouse model of intestinal neoplasia.
Our results argue against an oncogenic effect of DNA hypomethylation. Moreover, they are
consistent with a role for DNA methyltransferase in the generation of the C to T transitions
seen at high frequency in human colorectal tumors.

<>

<1>Laird, P.W., Jaenisch, R.
<2>The role of DNA methylation in cancer genetics and epigenetics.
<3>Annu. Rev. Genet.
<4>30
<5>441-464
<6>1996
<7>The past few years have seen a wider acceptance of a role for DNA methylation in cancer.  This
can be attributed to three developments.  First, the documentation of the over-representation
of mutations at CpG dinucleotides has convincingly implicated DNA methylation in the
generation of oncogenic point mutations.  The second important advance has been the
demonstration of epigenetic silencing of tumor suppressor genes by DNA methylation.  The third
development has been the utilization of experimental methods to manipulate DNA methylation
levels.  These studies demonstrate that DNA methylation changes in cancer cells are not mere
by-products of malignant transformation, but can play an instrumental role in the cancer
process.  It seems clear that DNA methylation plays a variety of roles in different cancer
types and probably at different stages of oncogenesis.  DNA methylation is intricately
involved in a wide diversity of cellular processes.  Likewise, it appears to exert its
influence on the cancer process through a diverse array of mechanisms.  It is our task not
only to identify these mechanisms, but to determine their relative importance for each state
and type of cancer.  Our hope then will be to translate that knowledge into clinical
applications.

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<1>Lajoie, M.J., Rovner, A.J., Goodman, D.B., Aerni, H.-R., Haimovich, A.D., Kuznetsov, G., Mercer, J.A., Wang, H.H., Carr, P.A., Mosberg, J.A., Rohland, N., Schultz, P.G., Jacobson, J.M., Rinehart, J., Church, G.M., Isaacs, F.J.
<2>Genomically Recoded Organisms Expand Biological Functions.
<3>Science
<4>342
<5>357-360
<6>2013
<7>We describe the construction and characterization of a genomically recoded organism (GRO). We
replaced all known UAG stop codons in Escherichia coli MG1655 with synonymous UAA codons,
which permitted the deletion of release factor 1 and reassignment of UAG translation function.
This
GRO exhibited improved properties for incorporation of nonstandard amino acids that expand the
chemical diversity of proteins in vivo. The GRO also exhibited increased resistance to T7
bacteriophage, demonstrating that new genetic codes could enable increased viral resistance.

<>

<1>Lakey, N.D., Jeddeloh, J.A., Korshunova, Y.
<2>Amplifying unmethylated or methylated DNA fragments in a biological sample comprises digesting portions of DNA with a  methylation-sensitive, methylation-dependent or methylation-insensitive  restriction enzyme - for use in polymorphism detection, diseas.
<3>International Patent Office
<4>WO 200585477
<5>
<6>2005
<7>NOVELTY - Amplifying unmethylated or methylated DNA fragments in a biological sample comprises
digesting portions of DNA
from a sample with various restriction enzymes and selectively
amplifying unmethylated, methylated or restriction site mutated DNA
from the portions, thus, generating amplified populations of DNA.
DETAILED DESCRIPTION - Amplifying unmethylated or methylated DNA
fragments in a biological sample comprises: (a) providing randomly
fragmented DNA from the biological sample; (b) adding a sequence tag
onto at least one end of the DNA fragments;(c) digesting the DNA
fragments with a methylation-dependent or methylation-sensitive
restriction enzyme to obtain intact DNA fragments and digested DNA
fragments; and (d) amplifying the intact modified DNA fragments with at
least one primer that initiates amplification from the sequence tags,
thus, generating amplified intact DNA fragments representing the
unmethylated or methylated modified intact DNA fragments in the sample.
AN INDEPENDENT CLAIM is also included for the method for comparing the
methylation state of a specific sequence in one portion of randomly
fragmented DNA to the methylation state of the same sequence in at
least a second portion of DNA. BIOTECHNOLOGY - Preferred Method:
Amplifying unmethylated or methylated DNA fragments in a biological
sample comprises randomly fragmenting DNA from the biological sample
before the adding step. The adding step comprises ligating the sequence
tag to at least one end of the DNA fragments. The sequence tags
comprise synthetic molecules that exhibit base pairing. The synthetic
molecules are selected from peptide nucleic acids and intercalating
nucleic acids. The adding step also comprises adding a homopolymeric
sequence tag to at least one of the ends of the DNA fragments with
terminal transferase. The sequence tags are added before or after the
digesting step. The digesting step comprises digesting the fragmented
DNA with a methylation-sensitive restriction enzyme; and the amplifying
step comprises amplifying intact modified fragments having the same
sequence as the methylated DNA in the sample. The digesting step may
also comprise digesting the fragmented DNA with a methylation-dependent
restriction enzyme; and the amplifying step also comprises amplifying
intact modified fragments having the same sequence as the unmethylated
DNA in the sample. The amplifying step comprises polymerase chain
reaction, rolling circle amplification or branched chain amplification.
The amplification is linear. The above method comprises quantifying the
number of amplified intact DNA fragments comprising a particular
sequence. The quantifying step comprises hybridizing the amplified
intact DNA to a quantifying polynucleotide. The quantifying
polynucleotide comprises synthetic molecules that exhibit base pairing.
The synthetic molecules are selected from peptide nucleic acids and
intercalating nucleic acids. The quantifying step is performed after
the amplifying step and the quantifying step comprises detecting copies
of a locus with hybrid capture. The quantifying polynucleotide is used
in a quantitative amplification step, and is attached to a solid
support. Before the amplifying step, the DNA fragments are contacted
with an agent that modifies unmethylated cytosines but does not modify
methylated cytosines; and the quantifying step comprises hybridizing a
polynucleotide to amplified intact DNA where the polynucleotide
hybridizes to the converted sequence. The digesting step is performed
under conditions that allow for at least some copies of methylated
modified DNA fragments to remain intact; and the density of methylation
at a locus is determined by comparing: the number of intact methylated
modified DNA fragments that contain the locus after the digesting step;
and a control value representing the quantity or density of methylated
DNA fragments in a control DNA. The above method further comprises
sequencing the amplified intact DNA fragments. It further comp

<>

<1>Laksanalamai, P., Steyert, S.R., Burall, L.S., Datta, A.R.
<2>Genome Sequences of Listeria monocytogenes Serotype 4b Variant Strains Isolated from Clinical and Environmental Sources.
<3>Genome Announcements
<4>1
<5>e00771-13
<6>2013
<7>Listeria monocytogenes strains that show a novel PCR serotyping profile (IVb-v1)  have been
reported recently. Here, we announce the draft genome sequences of five
L. monocytogenes IVb-v1 strains isolated from the United States and Australia
that harbor a 6.3-kb DNA cassette characteristic of serotype 1/2a strains.

<>

<1>Lakshmipathy, D., Vetrivel, U., Irudayam, L.T., Ramasubban, G., Madhavan, H.N., Sridhar, R., Meenakshi, N.
<2>Draft Genome Sequence of Multidrug-Resistant Mycobacterium tuberculosis Strain CWCFVRF MDRTB 670, Isolated from the Sputum of a Patient from Chennai, India,  with Clinically Suspected Tuberculosis.
<3>Genome Announcements
<4>2
<5>e00475-14
<6>2014
<7>We announce the draft genome sequence of a multidrug-resistant Mycobacterium tuberculosis
strain (CWCFVRF MDRTB 670) isolated from sputum from a patient with
clinically suspected tuberculosis.

<>

<1>Lakshmipathy, D., Vetrivel, U., Ramasubban, G., Kulandai, L.T., Madhavan, H.N., Sridhar, R., Meenakshi, N.
<2>Whole Genome Sequence of Polyresistant Mycobacterium tuberculosis CWCFVRF PRTB 19 Sputum Isolate from Chennai, India, Closely Clustering with East African Indian 5  Genogroup.
<3>Genome Announcements
<4>2
<5>e00702-14
<6>2014
<7>We announce the draft genome sequence of a polyresistant Mycobacterium tuberculosis strain
(CWCFVRF PRTB 19) isolated from the sputum of a clinically
suspected tuberculosis patient, and it closely clusters to the East African
Indian 5 (EAI5) lineage.

<>

<1>Lal S, R.U., Zhang, X., Munir, R., Sparling, R., Levin, D.B.
<2>Draft Genome Sequence of the Cellulolytic, Mesophilic, Anaerobic Bacterium Clostridium termitidis Strain CT1112 (DSM 5398).
<3>Genome Announcements
<4>1
<5>e00281-13
<6>2013
<7>Here, we report the draft genome sequence of Clostridium termitidis strain CT1112 (DSM 5398),
a mesophilic, cellulolytic bacterium that can utilize a variety of
sugars, as well as pure cellulose, as a sole carbon source; it also synthesizes
fermentation end products with potential industrial applications.

<>

<1>Lal S, R.U., Zhang, X., Sparling, R., Levin, D.B.
<2>Draft Genome Sequence of the Hydrogen- and Ethanol-Producing Bacterium Clostridium intestinale Strain URNW.
<3>Genome Announcements
<4>1
<5>e00871-13
<6>2013
<7>Here, we report the draft genome sequence of Clostridium intestinale strain URNW, which can
convert biomass to useful products such as biofuels (hydrogen or
ethanol) and other soluble end products.

<>

<1>Lam, M.C., Seemann, T., Bulach, D.M., Gladman, S.L., Chen, H., Haring, V., Moore, R.J., Ballard, S., Grayson, M.L., Johnson, P.D., Howden, B.P., Stinear, T.P.
<2>Comparative Analysis of the First Complete Enterococcus faecium Genome.
<3>J. Bacteriol.
<4>194
<5>2334
<6>2012
<7>Vancomycin-resistant enterococci (VRE) are one of the leading causes of nosocomial infections
in healthcare facilities around the globe. In particular, infections caused by
vancomycin-resistant Enterococcus faecium are becoming increasingly common. Comparative and
functional genomic studies of E. faecium isolates have so far been limited owing to the lack
of a fully assembled E. faecium genome sequence. Here we address this issue and report the
complete 3.0 Mb genome sequence of the multi-locus sequence type 17 vancomycin-resistant
Enterococcus faecium strain Aus0004, isolated from the bloodstream of a patient in Melbourne,
Australia in 1998. The genome comprises a 2.9 Mb circular chromosome and three circular
plasmids. The chromosome harbours putative E. faecium virulence factors such as enterococcal
surface protein, hemolysin and collagen-binding adhesion. Aus0004 has a very large accessory
genome (38%) that includes, three prophage and two genomic islands absent among 22 other E.
faecium genomes. One of the prophage was present as inverted 50kb repeats that appear to have
facilitated a 683 kb chromosomal inversion across the replication terminus, resulting in a
striking replichore imbalance. Other distinctive features include 95 insertion sequence
elements and a single chromosomal copy of Tn1549 containing the vanB vancomycin-resistance
element. A complete E. faecium genome will be a useful resource to assist our understanding of
this emerging nosocomial pathogen.

<>

<1>Lam, W.C., Tsao, D.H.H., Maki, A.H., Maegley, K.A., Reich, N.O.
<2>Spectroscopic studies of arsenic(III) binding to Escherichia coli RI methyltransferase and to two mutants, C223S and W183F.
<3>Biochemistry
<4>31
<5>10438-10442
<6>1992
<7>The interactions of an arsenic(III) reagent, (CH3)2AsSCH2CONH2, with two Escherichia coli RI
methyltransferase mutants, W183F and C223S, have been studied by phosphorescence, optically
detected magnetic resonance, and fluorescence spectroscopy. The phosphorescence specrum of the
W183F mutant containing only one tryptophan at position 225 reveals a single 0,0-band that is
red-shifted by 9.8 nm upon binding of As(III). Fluorescence titration of W183F with
(CH3)2AsSCH2CONH2 produces a large tryptophan fluorescence quenching. Analysis of the
quenching data points to a single high-affinity As(III) binding site that is associated with
the fluorescence quenching. Triplet-state kinetic measurements performed on the perturbed
tryptophan show large reductions in the lifetimes of the triplet sublevels, especially that of
the Tx sublevel. As(III) binding to the enzymes at a site very close to the Trp225 residue
induces an external heavy-atom effect, showing that the perturber atom is in van der Waals
contact with the indole chromophore. In the case of the C223S mutant, a single tryptophan
0,0-band also is observed in the phosphorescence spectrum but no change occurs upon addition
of the As(III) reagent. Fluorescence titration of C223S with As(III) shows essentially no
quenching of tryptophan fluorescence, in contrast with W183F. These results along with
previous triplet-state and biochemical studies on the wild-type enzyme [Tsao,D.H>H., & Maki,
A.H. (1991) Biochemistry 30, 4565-4572], show that As(III) binds with high affinity to the
Cys223 residue and that the Trp225 side chain is located close enough to that of Cys223 to
produce a heavy-atom perturbation when As(III) is bound.

<>

<1>Lambert, A.R., Sussman, D., Shen, B., Maunus, R., Nix, J., Samuelson, J., Xu, S.Y., Stoddard, B.L.
<2>Structures of the Rare-Cutting Restriction Endonuclease NotI Reveal a Unique Metal Binding Fold Involved in DNA Binding.
<3>Structure
<4>16
<5>558-569
<6>2008
<7>The structure of the rare-cutting restriction endonuclease NotI, which recognizes the 8 bp
target 5'-GCGGCCGC-3', has been solved with and
without bound DNA. Because of its specificity (recognizing a site that
occurs once per 65 kb), NotI is used to generate large genomic fragments
and to map DNA methylation status. NotI contains a unique metal binding
fold, found in a variety of putative endonucleases, occupied by an iron
atom coordinated within a tetrahedral Cys4 motif. This domain positions
nearby protein elements for DNA recognition, and serves a structural role.
While recognition of the central six base pairs of the target is
accomplished via a saturated hydrogen bond network typical of restriction
enzymes, the most peripheral base pairs are engaged in a single direct
contact in the major groove, reflecting reduced pressure to recognize
those positions. NotI may represent an evolutionary intermediate between
mobile endonucleases (which recognize longer target sites) and canonical
restriction endonucleases.

<>

<1>Lambert, G.R., Carr, N.G.
<2>Resistance of DNA from filamentous and unicellular cyanobacteria to restriction endonuclease cleavage.
<3>Biochim. Biophys. Acta
<4>781
<5>45-55
<6>1984
<7>Chromosomal DNA from nine species of filamentous cyanobacteria as
diverse as Nostoc, Gloeotrichia and Plectonema is suggested to be extensively modified
(methylated) by its resistance to cleavage by a number of restriction endonucleases.  A
remarkably similar pattern of DNA modification in these species contrasts with the known
heterogeneity of their type II restriction endonuclease content.  In particular, Nostoc PCC
73102, which lacks detectable sequence-specific endonucleases, is shown to possess
extensive DNA modification.  The use of isoschizomers demonstrates the presence of a
methylase in the filamentous strains analogous to the dam enzyme of Escherichia coli.  As a
preliminary to assessing the significance of the DNA modification, a study of susceptibility
to restriction endonuclease cleavage of the genomes of five unicellular cyanobacteria
revealed considerable variation between the different strains.  The significance of the DNA
modification patterns elucidated is discussed in terms of the restriction endonuclease
content and cellular differentiation of the relevant cyanobacterial strains.

<>

<1>Lambert, J.N.
<2>The chemistry of biology: Molecular evolution, the total synthesis of proteins, and DNA restriction and modification.
<3>Aust. J. Chem.
<4>49
<5>1179-1196
<6>1996
<7>Three topics of current interest in modern biological organic chemistry are presented:
molecular evolution, the total synthesis of proteins, and the study of DNA restriction and
modification systems.  These topics are used to illustrate the application of chemical
understanding to the study of chemistry in a biological context.

<>

<1>Lambie, S.C., Altermann, E., Leahy, S.C., Kelly, W.J.
<2>Draft Genome Sequence of Lactococcus lactis subsp. cremoris HPT, the First Defined-Strain Dairy Starter Culture Bacterium.
<3>Genome Announcements
<4>2
<5>e00107-14
<6>2014
<7>Lactococcus lactis subsp. cremoris HP(T) has been widely used in studies of the metabolism of
lactococcal dairy starter cultures. A comparison of the draft HP(T)
genome with those from other strains of L. lactis subsp. cremoris will aid our
understanding of the domestication and evolution of these important industrial
cultures.

<>

<1>Lambowitz, A.M.
<2>Infectious introns.
<3>Cell
<4>56
<5>323-326
<6>1989
<7>Among the most vigorously debated questions in modern biology is whether introns were present
in primordial genes and subsequently lost from most present-day prokaryotes or whether they
are more recent additions to eukaryotic genes. While it is not necessarily true that all
introns originated in the same way from a common ancestor, recent studies, represented by four
papers in this issue of Cell, provide definitive evidence that a number of group I introns can
propagate themselves by insertion of group II introns and for relationships between these
introns and autonomous elements.

<>

<1>Lambowitz, A.M., Belfort, M.
<2>Introns as mobile genetic elements.
<3>Annu. Rev. Biochem.
<4>62
<5>587-622
<6>1993
<7>*
Introduction
Intron Structure and Splicing Pathway
Group I introns
Group II introns
Archaeal introns
Intron Mobility
Intron distribution
Intron-encoded proteins
Intron acquisition or loss
Autonomous introns and intronlike elements
An autonomous Group II intron -- a-senDNA
Mitochondrial plasmids of neurospora
Intron evaluation
Introns early and late
ORF acquisition and evolution of group I intron mobility
Acquisition of group II intron ORFs
Regulation of intron mobility
Horizontal transmission?
Concluding remarks


<>

<1>Lambowitz, A.M., Caprara, M.G., Zimmerly, S., Perlman, P.S.
<2>Group I and Group II ribozymes as RNPs: Clues to the past and guides to the future.
<3>The RNA World, Cold Spring Harbor Laboratory Press, Gesteland, R.F., Cech, T.R., Atkins, J.F., Cold Spring Harbor
<4>
<5>451-485
<6>1999
<7>Group I and group II introns are not only catalytic RNAs, but also mobile genetic elements.
The success of these introns as mobile elements almost certainly relates to their innate
self-splicing capability, which enables them to propagate by inserting into host genes while
only minimally impairing gene expression.  Nevertheless, both types of introns have become
dependent on proteins for efficient splicing in vivo to help fold the intron RNA into the
catalytically active structure.  A review.

<>

<1>Lambowitz, A.M., Mohr, G., Zimmerly, S.
<2>Group II intron homing endonucleases: ribonucleoprotein complexes with programmable target specificity.
<3>Nucleic Acids Mol. Biol.
<4>16
<5>121-145
<6>2005
<7>Group II intron homing endonucleases are ribonucleoproteins consisting of a catalytically
active intron RNA and an intron-encoded protein, with reverse transcriptase and/or DNA
endonuclease activity.  The RNP is formed when the IEP binds to the intron in unspliced RNA
and promotes its splicing by stabilizing the catalytically active RNA structure.  Afterwards,
the IEP remains tightly bound to the excised intron RNA to constitute the homing endonuclease.
The homing endonuclease promotes intron mobility by a remarkable mechanism in which the intron
RNA reverse splices directly into a target DNA and is then reverse-transcribed by the IEP.
Importantly, the target site for intron insertion is determined mainly by base pairing between
short sequence elements in the intron RNA and target DNA, making it straightforward to change
the target specificity of the homing endonuclease simply by modifying the intron RNA.  This
feature combined with their very high specificity and insertion frequencies have made it
possible to develop mobile group II introns into gene targeting vectors, called "targetrons",
with programmable target specificity.

<>

<1>Lamontanara, A., Orru, L., Cattivelli, L., Russo, P., Spano, G., Capozzi, V.
<2>Genome Sequence of Oenococcus oeni OM27, the First Fully Assembled Genome of a Strain Isolated from an Italian Wine.
<3>Genome Announcements
<4>2
<5>e00658-14
<6>2014
<7>Oenococcus oeni OM27 is a strain selected from 'Nero di Troia' wine undergoing spontaneous
malolactic fermentation. 'Nero di Troia' is a wine made from 'Uva di
Troia' grapes, an autochthonous black grape variety from the Apulian region
(south of Italy). In this paper we present a 1.78-Mb assembly of the O. oeni OM27
genome, the first fully assembled genome of an O. oeni strain from an Italian
wine.

<>

<1>Lan, J., Hua, S., He, X.N., Zhang, Y.
<2>DNA methyltransferases and methyl-binding proteins of mammals.
<3>Acta Biochim. Biophys. Sin.
<4>42
<5>243-252
<6>2010
<7>In mammals, DNA methylation, characterized by the transfer of the methyl group from
S-adenosylmethionines to a base (mainly referred to cytosine), acts as a major epigenetic
modification. In parallel to DNA sequences arrangement, modification of methylation to DNA
sequences has far-reaching influence on biological functions and activities, for it involves
controlling gene transcription, regulating chromatin structure, sustaining genome stability
and integrity, maintaining parental imprinting and X-chromosome inactivation, suppressing
homologous recombination as well as limiting transposable elements, during which DNA
methyltransferases (DNMTs) and methyl-binding proteins play important roles. Their aberrance
can give rise to dysregulation of gene expression, cell maltransformation and so on. Hence, it
is necessary to gain a good understanding of these two important kinds of proteins, which will
help to better investigate the epigenetic mechanisms and manipulate the modifications
according to our will based on its reversibility. Here we briefly review our current
understanding of DNMTs and methyl-binding proteins in mammals.

<>

<1>Lan, J., Hua, S., Liu, J., Zhang, Y.
<2>cDNA Cloning of Goat DNA Methyltransferase 1, Screening of shRNA Vectors and Influences to Development of Nuclear Transfer Embryos.
<3>Agric. Sci. China
<4>9
<5>1035-1040
<6>2010
<7>This study was designed to clone cDNA of goat DNA methyltransferase 1 (DNMT1) gene, to screen
an effective shRNA-producing vector targeting
goat DNA methyltransferase 1 and to improve the developmental
competence of goat nuclear transfer embryos by decreasing the DNMT1
expression in donor cells. In this study, PCR primers were designed
against regions of high homology between bovine and sheep sequences and
then used to amplify the larger portions of the coding regions. Next, 3
RNAi oligonucleotides were designed based on the cloned sequences and
inserted into pRNAT-U6.1/Neo vector, acquiring 3 new vectors,
respectively termed pRNAD1, pRNAD2 and pRNAD3. Then the positive cells
were sorted by flow cytometry after transfection and detected by
real-time PCR analysis and sodium bisulfite genomic sequencing.
Finally, the developmental rates of nuclear transfer (NT) embryos
generated using donor cells with and without the effective shRNA vector
respectively, as well as in vitro fertilization (IVF) embryos were
observed and recorded. The results showed that the coding regions of
goat DNA methyltransferase 1 gene was successfully cloned (GenBank no.
FJ617538). Furthermore, an effective interfering shRNA (pRNAD2) was
obtained, with its interference effect being 47.88%. Finally, NT
embryos with shRNA vector harbored better developmental competence
during morula and blastocyst stage compared to controls (P<0.05),
reaching the similar rates to IVF embryos (P>0.05). In conclusion, goat
DNA methyltransferase 1 gene cDNA was cloned and sequenced, an
effective shRNA vector responsible for inhibiting DNA methyltransferase
1 expression was developed and the developmental competence of goat
nuclear transfer morulae and blastcysts was significantly improved,
which provided a feasible pathway for improving goat nuclear transfer
embryo development competence by decreasing the methylation level in
donor cells through RNAi-mediated manner.

<>

<1>Lan, S.F., Huang, C.H., Chang, C.H., Liao, W.C., Lin, I.H., Jian, W.N., Wu, Y.G., Chen, S.Y., Wong, H.C.
<2>Characterization of a New Plasmid-Like Prophage in a Pandemic Vibrio parahaemolyticus O3:K6 Strain.
<3>Appl. Environ. Microbiol.
<4>75
<5>2659-2667
<6>2009
<7>Vibrio parahaemolyticus is a common food-borne pathogen that is normally associated with
seafood. In 1996, a pandemic O3:K6 strain
abruptly appeared and caused the first pandemic of this pathogen to
spread throughout many Asian countries, America, Europe, and Africa.
The role of temperate bacteriophages in the evolution of this pathogen
is of great interest. In this work, a new temperate phage, VP882, from
a pandemic O3: K6 strain of V. parahaemolyticus was purified and
characterized after mitomycin C induction. VP882 was a Myoviridae
bacteriophage with a polyhedral head and a long rigid tail with a
sheath-like structure. It infected and lysed high proportions of V.
parahaemolyticus, Vibrio vulnificus, and Vibrio cholerae strains. The
genome of phage VP882 was sequenced and was 38,197 bp long, and 71
putative open reading frames were identified, of which 27 were putative
functional phage or bacterial genes. VP882 had a linear plasmid-like
genome with a putative protelomerase gene and cohesive ends. The genome
does not integrate into the host chromosome but was maintained as a
plasmid in the lysogen. Analysis of the reaction sites of the
protelomerases in different plasmid-like phages revealed that VP882 and
Phi HAP-1 were highly similar, while N15, Phi KO2, and PY54 made up
another closely related group. The presence of DNA adenine methylase
and quorum-sensing transcriptional regulators in VP882 may play a
specific role in this phage or regulate physiological or
virulence-associated traits of the hosts. These genes may also be
remnants from the bacterial chromosome following transduction.

<>

<1>Land, M. et al.
<2>Complete genome sequence of Beutenbergia cavernae type strain (HKI 0122).
<3>Standards in Genomic Sciences
<4>1
<5>21-28
<6>2009
<7>Beutenbergia cavernae (Groth et al. 1999) is the type species of the genus and is of
phylogenetic interest because of its isolated location in the actinobacterial
suborder Micrococcineae. B. cavernae HKI 0122(T) is a Gram-positive, non-motile,
non-spore-forming bacterium isolated from a cave in Guangxi (China). B. cavernae
grows best under aerobic conditions and shows a rod-coccus growth cycle. Its cell
wall peptidoglycan contains the diagnostic L-lysine <-- L-glutamate interpeptide
bridge. Here we describe the features of this organism, together with the
complete genome sequence, and annotation. This is the first completed genome
sequence from the poorly populated micrococcineal family Beutenbergiaceae, and
this 4,669,183 bp long single replicon genome with its 4225 protein-coding and 53
RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Land, M. et al.
<2>Non-contiguous finished genome sequence of Bacteroides coprosuis type strain (PC139).
<3>Standards in Genomic Sciences
<4>4
<5>233-243
<6>2011
<7>Bacteroides coprosuis Whitehead et al. 2005 belongs to the genus Bacteroides, which is a
member of the family Bacteroidaceae. Members of the genus Bacteroides
in general are known as beneficial protectors of animal guts against pathogenic
microorganisms, and as contributors to the degradation of complex molecules such
as polysaccharides. B. coprosuis itself was isolated from a manure storage pit of
a swine facility, but has not yet been found in an animal host. The species is of
interest solely because of its isolated phylogenetic location. The genome of B.
coprosuis is already the 5(th) sequenced type strain genome from the genus
Bacteroides. The 2,991,798 bp long genome with its 2,461 protein-coding and 78
RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Land, M. et al.
<2>Complete genome sequence of Actinosynnema mirum type strain (101).
<3>Standards in Genomic Sciences
<4>1
<5>46-53
<6>2009
<7>Actinosynnema mirum Hasegawa et al. 1978 is the type species of the genus, and is of
phylogenetic interest because of its central phylogenetic location in the
Actino-synnemataceae, a rapidly growing family within the actinobacterial
suborder Pseudo-nocardineae. A. mirum is characterized by its motile spores borne
on synnemata and as a producer of nocardicin antibiotics. It is capable of
growing aerobically and under a moderate CO(2) atmosphere. The strain is a
Gram-positive, aerial and substrate mycelium producing bacterium, originally
isolated from a grass blade collected from the Raritan River, New Jersey. Here we
describe the features of this organism, together with the complete genome
sequence and annotation. This is the first complete genome sequence of a member
of the family Actinosynnemataceae, and only the second sequence from the
actinobacterial suborder Pseudonocardineae. The 8,248,144 bp long single replicon
genome with its 7100 protein-coding and 77 RNA genes is part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Landry, D., Barsomian, J.M., Feehery, G.R., Wilson, G.G.
<2>Characterization of Type II DNA-Methyltransferases.
<3>Methods Enzymol.
<4>216
<5>244-259
<6>1992
<7>
<>

<1>Landry, D., Looney, M.C., Feehery, G.R., Slatko, B.E., Jack, W.E., Schildkraut, I., Wilson, G.G.
<2>M.FokI methylates adenine in both strands of its asymmetric recognition sequence.
<3>Gene
<4>77
<5>1-10
<6>1989
<7>M.FokI, a type-IIS modification enzyme from Flavobacterium okeanokoites, was purified, and its
activity was characterized in vitro.  The enzyme was found to be a DNA-adenine
methyltransferase and to methylate both strands of the asymmetric FokI recognition sequence:
M.FokI
5'-GGATG / CCTAC-5' M.FokI-> 5'-GGm6ATG/CCTm6AC-5.  M.FokI does not methylate
single-stranded DNA, nor does it methylate double-stranded DNA at sequences other than FokI
sites.

<>

<1>Landthaler, M., Begley, U., Lau, N.C., Shub, D.A.
<2>Two self-splicing group I introns in the ribonucleotide reductase large subunit gene of Staphylococcus aureus phage Twort.
<3>Nucleic Acids Res.
<4>30
<5>1935-1943
<6>2002
<7>We have recently described three group I introns inserted into a single gene, orf142, of the
staphylococcal bacteriophage Twort and suggested the presence of at least two additional
self-splicing introns in this phage genome. Here we report that two previously uncharacterized
introns, 429 and 1087 nt in length, interrupt the Twort gene coding for the large subunit of
ribonucleotide reductase (nrdE). Reverse transcription-polymerase chain reaction (RT-PCR) of
RNA isolated from Staphylococcus aureus after phage infection indicates that the introns are
removed from the primary transcript in vivo. Both nrdE introns show sequence similarity to the
Twort orf142 introns I2 and I3, suggesting either a common origin of these introns or
shuffling of intron structural elements. Intron 2 encodes a DNA endonuclease, I-TwoI, with
similarity to homing endonucleases of the HNH family. Like I-HmuI and I-HmuII, intron-encoded
HNH endonucleases in Bacillus subtilis phages SPO1 and SP82, I-TwoI nicks only one strand of
its DNA recognition sequence. However, whereas I-HmuI and I-HmuII cleave the template strand
in exon 2, I-TwoI cleaves the coding strand in exon 1. In each case, the 3' OH created on the
cut strand is positioned to prime DNA synthesis towards the intron, suggesting that this
reaction contributes to the mechanism of intron homing. Both nrdE introns are inserted in
highly conserved regions of the ribonucleotide reductase gene, next to codons for functionally
important residues.

<>

<1>Landthaler, M., Lau, N.C., Shub, D.A.
<2>Group I intron homing in Bacillus phages SPO1 and SP82: A gene conversion event initiated by a nicking homing endonuclease.
<3>J. Bacteriol.
<4>186
<5>4307-4314
<6>2004
<7>Many group I introns encode endonucleases that promote intron homing by initiating a
double-stranded break-mediated homologous recombination event. In this work we describe intron
homing in Bacillus subtilis phages SPO1 and SP82. The introns encode the DNA endonucleases
I-HmuI and I-HmuII, respectively, which belong to the H-N-H endonuclease family and possess
nicking activity in vitro. Coinfections of B. subtilis with intron-minus and intron-plus
phages indicate that I-HmuI and I-HmuII are required for homing of the SPO1 and SP82 introns,
respectively. The homing process is a gene conversion event that does not require the major B.
subtilis recombination pathways, suggesting that the necessary functions are provided by
phage-encoded factors. Our results provide the first examples of H-N-H endonuclease-mediated
intron homing and the first demonstration of intron homing initiated by a nicking
endonuclease.

<>

<1>Landthaler, M., Shen, B.W., Stoddard, B.L., Shub, D.A.
<2>I-Basl and I-Hmul: Two phage intron-encoded endonucleases with homologous DNA recognition sequences but distinct DNA Specificities.
<3>J. Mol. Biol.
<4>358
<5>1137-1151
<6>2006
<7>I-HmuI and I-BasI are two highly similar nicking DNA endonucleases, which are each encoded by
a group I intron inserted into homologous sites within
the DNA polymerase genes of Bacillus phages SPO1 and Bastille,
respectively. Here, we present a comparison of the DNA specificities and
cleavage activities of these enconucleases with homologous target sites.
I-BasI has properties that are typical of homing endonucleases, nicking
the intron-minus polymerase genes in either host genome, three nucleotides
downstream of the intron insertion site. In contrast, I-HmuI nicks both
the intron-plus and intron-minus site in its own host genome, but does not
act on the target from Bastille phage. Although the enzymes have distinct
DNA substrate specificities, both bind to an identical 25bp region of
their respective intron-minus DNA polymerase genes surrounding the intron
insertion site. The endonucleases appear to interact with the DNA
substrates in the downstream exon 2 in a similar manner. However, whereas
I-HmuI is known to make its only base-specific contacts within this exon
region, structural modeling analyses predict that I-BasI might make
specific base contacts both upstream and downstream of the site of intron
insertion. The predicted requirement for base-specific contacts in exon 1
for cleavage by I-BasI was confirmed experimentally. This explains the
difference in substrate specificities between the two enzymes, including
the observation that the former enzyme is relatively insensitive to the
presence of an intron upstream of exon 2. These differences are likely a
consequence of divergent evolutionary constraints.

<>

<1>Landthaler, M., Shub, D.A.
<2>The nicking homing endonuclease I-BasI is encoded by a group I intron in the DNA polymerase gene of the Bacillus thuringiensis phage Bastille.
<3>Nucleic Acids Res.
<4>31
<5>3071-3077
<6>2003
<7>Here we describe the discovery of a group I intron in the DNA polymerase gene of Bacillus
thuringiensis phage Bastille. Although the intron
insertion site is identical to that of the Bacillus subtilis phages SPO1
and SP82 introns, the Bastille intron differs from them substantially in
primary and secondary structure. Like the SPO1 and SP82 introns, the
Bastille intron encodes a nicking DNA endonuclease of the H-N-H family,
I-BasI, with a cleavage site identical to that of the SPO1-encoded enzyme
I-HmuI. Unlike I-HmuI, which nicks both intron-minus and intron-plus DNA,
I-BasI cleaves only intron-minus alleles, which is a characteristic of
typical homing endonucleases. Interestingly, the C-terminal portions of
these H-N-H phage endonucleases contain a conserved sequence motif, the
intron-encoded endonuclease repeat motif (IENR1) that also has been found
in endonucleases of the GIY-YIG family, and which likely comprises a small
DNA-binding module with a globular betabetaalphaalphabeta fold, suggestive
of module shuffling between different homing endonuclease families.

<>

<1>Landy, A., Foeller, C., Ross, W.
<2>DNA fragments carrying genes for tRNATyrI.
<3>Nature
<4>249
<5>738-742
<6>1974
<7>The duplicated genes coding for tRNA TyrI have been isolated using
site-specific nucleases and gel electrophoresis.  An unduplicated sequence of
4-120 base pairs separates the two genes.  Specific DNA fragments can be
obtained in sizes appropriate for both functional and structural analysis of
the tRNA TyrI region.

<>

<1>Landy, A., Olchowski, E., Ross, W., Reiness, G.
<2>Isolation of a functional lac regulatory region.
<3>Mol. Gen. Genet.
<4>133
<5>273-281
<6>1974
<7>A DNA fragment containing the lac regulatory region has been isolated by
digestion of lambda plac 5 DNA with restriction endonuclease from Hemophilus
influenzae Rd followed by gel electrophoresis.  This DNA fragment, which is
approximately 660 base pairs, is identified by its ability to bind purified lac
repressor, and has been shown to be functional in a coupled
transcription-translation system.  A smaller repressor binding fragment of
approximately 174 base pairs has been derived from the larger fragment
utilizing a secondary digestion with restriction endonuclease from Hemophilus
aegyptius.

<>

<1>Landy, A., Ruedisueli, E., Robinson, L., Foeller, C., Ross, W.
<2>Digestion of deoxyribonucleic acids from bacteriophage T7, lambda, and Phi80h with site-specific nucleases from Hemophilus influenzae strain Rc and strain Rd.
<3>Biochemistry
<4>13
<5>2134-2142
<6>1974
<7>The digestion profiles of DNA from bacteriophage T7, lambda, and Phi80h by the
site-specific nucleases from Hemophilus influenzae strain Rc and strain Rd have
been analyzed.  The DNA fragments range in molecular weight from approximately
0.02 to 3 Md and are resolved by polyacrylamide-agarose gel electrophoresis.
The digestion profiles thus produced are unique and characteristic for both the
DNA and the nuclease.  The T7 DNA digestion products produced by the Rc and Rd
nucleases are identical.  With Phi80h and lambda DNAs only 87 and 71%
respectively, of the total number of fragments produced by the Rd nuclease are
common to both nuclease digestions, the remaining DNA fragments being unique to
either the Rc or the Rd digestions.  Strain Rc possesses a site-specific
nuclease activity which is identical to an activity found in strain Rd.  In
addition strain Rd carries a second activity which is not found in strain Rc
and which elutes at slightly higher salt on phosphocellulose.  Protection
experiments with individual modification methylases of strain Rd (Roy, P.H. and
Smith, H.O. (1973), J. Mol. Biol. 81: 427) establish that the nuclease which is
common to both Rd and Rc is associated with methylase II.  This nuclease in
strain Rd is called HindII and in strain Rc is called HincII according to the
nomenclature proposed by Smith and Nathans (Smith, H.O., and Nathans, D.
(1973), J. Mol. Biol. 81: 419-423.  The additional activity which elutes second
from phosphocellulose is HindIII.  The HincII-HindII activity (i.e., HincII or
HindII) is best prepared from strain Rc.  It makes 34 cuts in lambda DNA, 43
cuts in Phi80h DNA, and 51 cuts in T7 DNA.  The HindIII nuclease makes no cuts
in T7 DNA and 6 and 4 cuts in lambda and Phi80h DNA, respectively.  A
nomenclature system for the DNA digestion products is proposed and all the DNA
fragments produced by Rc and Rd digestions of the three related phages, Phi80h
ambl, Phi80h psuIII+-, and Phi80h psuIII+ are identified and their molecular
weights are presented.

<>

<1>Lang, E. et al.
<2>Complete genome sequence of Weeksella virosa type strain (9751).
<3>Standards in Genomic Sciences
<4>4
<5>81-90
<6>2011
<7>Weeksella virosa Holmes et al. 1987 is the sole member and type species of the genus Weeksella
which belongs to the family Flavobacteriaceae of the phylum
Bacteroidetes. Twenty-nine isolates, collected from clinical specimens provided
the basis for the taxon description. While the species seems to be a saprophyte
of the mucous membranes of healthy man and warm-blooded animals a causal
relationship with disease has been reported in a few instances. Except for the
ability to produce indole and to hydrolyze Tween and proteins such as casein and
gelatin, this aerobic, non-motile, non-pigmented bacterial species is
metabolically inert in most traditional biochemical tests. The 2,272,954 bp long
genome with its 2,105 protein-coding and 76 RNA genes consists of one circular
chromosome and is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Lang, E. et al.
<2>Complete genome sequence of Dyadobacter fermentans type strain (NS114).
<3>Standards in Genomic Sciences
<4>1
<5>133-140
<6>2009
<7>Dyadobacter fermentans (Chelius and Triplett, 2000) is the type species of the genus
Dyadobacter. It is of phylogenetic interest because of its location in the
Cytophagaceae, a very diverse family within the order 'Sphingobacteriales'. D.
fermentans has a mainly respiratory metabolism, stains Gram-negative, is
non-motile and oxidase and catalase positive. It is characterized by the
production of cell filaments in aging cultures, a flexirubin-like pigment and its
ability to ferment glucose, which is almost unique in the aerobically living
members of this taxonomically difficult family. Here we describe the features of
this organism, together with the complete genome sequence, and its annotation.
This is the first complete genome sequence of the sphingobacterial genus
Dyadobacter, and this 6,967,790 bp long single replicon genome with its 5804
protein-coding and 50 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Lange, C., Jugel, A., Walter, J., Noyer-Weidner, M., Trautner, T.A.
<2>Pseudo domains in phage-encoded DNA methyltransferases.
<3>Nature
<4>352
<5>645-648
<6>1991
<7>5-Cytosine-DNA-methyltransferases, which are found in many organisms ranging
from bacteriophages to mammals, transfer a methyl group from
S-adenosylmethionine to the carbon-5 of a cytosine residue in specific DNA
target sequences.  Some phage-encoded methyltransferases methylate more than
one sequence:  these enzymes contain several independent target-recognizing
domains each responsible for recognizing a different site.  The amino-acid
sequences of these multispecific methyltransferases reveal that some enzymes in
addition carry domains that do not contribute to the enzymes' methylation
potential, but strongly resemble previously identified target-recognizing
domains.  Here we show that introducing defined amino-acid alterations into
these inactive domains endows these enzymes with additional methylation
specificities.  Gel retardation analysis demonstrates that these novel
methylation specificities correlate with the acquisition of additional
DNA-binding potential of the proteins.

<>

<1>Lange, C., Noyer-Weidner, M., Trautner, T.A., Weiner, M., Zahler, S.A.
<2>M.H2I, a multispecific 5C-DNA methyltransferase encoded by Bacillus amyloliquefaciens phage H2.
<3>Gene
<4>100
<5>213-218
<6>1991
<7>Bacillus amyloliquefaciens phage H2 codes for a multispecific
cytosine-5-DNA-methyltransferase (MTase), M.H2I, which methylates GGCC, GCNGC
and G(A/G/T)GC(T/C/A)C target sequences.  The gene coding for M.H2I was cloned
in Escherichia coli and its nucleotide (nt) sequence was determined.  It
consists of 1509 bp, corresponding to a protein of 503 amino acids (aa) with a
calculated Mr of 57,166.  A comparison of the aa sequence of M.H2I with those
of the multispecific MTases encoded by Bacillus subtilis phages SPR, Phi3T and
Rho11S, revealed that M.H2I is closely related to these enzymes.  A very high
degree of homology was observed between M.H2I and M.Rho11S, with 96.2% aa
identity and 97.8% nt identity of the corresponding genes.

<>

<1>Lange, C., Wild, C., Trautner, T.A.
<2>Altered sequence recognition specificity of a C5-DNA methyltransferase carrying a chimeric 'target recognizing domain'.
<3>Gene
<4>157
<5>127-128
<6>1995
<7>A MTase with a chimeric TRD with the N-terminal half derived from a TRD recognizing GCNGC, the
C-terminal half from one with CCWGG recognition, was constructed.  Its target specificity is
reported (hemimethylation of the first C in GCTGGC).

<>

<1>Lange, C., Wild, C., Trautner, T.A.
<2>Identification of a subdomain within DNA-(cytosine-C5)-methyltransferases responsible for the recognition of the 5' part of their DNA target.
<3>EMBO J.
<4>15
<5>1443-1450
<6>1996
<7>In previous work on DNA-(cytosine-C5)-methyltransferases (C5-MTases), domains had been
identified which are responsible for the sequence specificity of the different enzymes
(target-recognizing domains, TRDs).  Here we have analyzed the DNA methylation patterns of two
C5-MTases containing reciprocal chimeric TRDs, consisting of the N- and C-terminal parts
derived from two different parental TRDs specifying the recognition of 5'-CC(A/T)GG-3' and
5'-GCNGC-3'.  Sequences recognized by these engineered MTases were non-symmetrical and
degenerate, but contained at their 5' part a consensus sequence which was very similar to the
5' part of the target recognized by the parental TRD which contributed the N-terminal moiety
of the chimeric TRD.  The results are discussed in connection with the present understanding
of the mechanism of DNA target recognition of C5-MTases.  They demonstrate the possibility of
designing C5-MTases with novel DNA methylation specificities.

<>

<1>Lange, C., Wild, C., Trautner, T.A.
<2>Colinearity between amino acids of the target recognizing domains of a C5-DNA-methyltransferase and the nucleotide sequence of its target.
<3>FASEB J.
<4>9
<5>A1391
<6>1995
<7>Cytosine (C5)-DNA methyltransferases (C5-MTases) are composed of several highly conserved
domains involved in general steps of the methylation reaction and one variable target
recognizing domain (TRD), which is responsible for the recognition of the enzymes' specific
DNA targets.  TRDs have between about 30 and 50 amino acids of different composition.  We have
constructed two MTases with recombinant TRDs combining the amino- and carboxy terminal halves
of two (parental) TRDs, which recognize the pentameric sequences 5'-GCNGC (I) and
5'-CC(A/T)GG (II).  DNA methylated by the recombinant enzymes was treated with bisulfite to
distinguish cytosines from methyl-cytosines and then sequenced.  We observe that the TRD whose
amino terminus was derived from enzyme (I) recognized hexameric, non-symmetrical sequences
with the consensus 5'-GCTGGC.  The reciprocal TRD was inactive, but could be activated by
site specific mutagenesis to recognize the target 5'-PyNCCPyPy.  Neither of the enzymes
methylated the parental sequence.  The results are compatible with the three dimensional
structure of M.HhaI complexed with its DNA target, where two sequential recognition loops of
the TRD interact with contiguous parts of the DNA target.

<>

<1>Langella, P., Chopin, A.
<2>Effect of restriction-modification systems on transfer of foreign DNA into Lactococcus lactis subsp. lactis.
<3>FEMS Microbiol. Lett.
<4>59
<5>301-306
<6>1989
<7>The efficiency of transfer of plasmid pIP501 DNA into Restriction/Modification
(R/M) proficient strains of Lactococcus lactis subsp. lactis strongly depends
on the mode of transfer.  In two strains, each one containing a different R/M
system, the efficiency of protoplast transformation with unmodified pIP501 DNA
was restriction by more than 3 orders of magnitude when compared to pIP501
carrying the host modification.  By contrast, no difference was observed when
modified or unmodified DNA was introduced into the same hosts by
electroporation.  The pIP501 conjugation frequency between a R-/M- donor an an
isogenic recipient were also unaffected whether the recipient possesses a R/M
system or not.

<>

<1>Langhans, M.T., Palladino, M.J.
<2>Cleavage of mispaired heteroduplex DNA substrates by numerous restriction enzymes.
<3>Curr. Issues Mol. Biol.
<4>11
<5>1-12
<6>2008
<7>The utility of restriction endonucleases as a tool in molecular biology is in large part due
to the high degree of specificity with which they
cleave well-characterized DNA recognition sequences. The specificity of
restriction endonucleases is not absolute, yet many commonly used
assays of biological phenomena and contemporary molecular biology
techniques rely on the premise that restriction enzymes will cleave
only perfect cognate recognition sites. In vitro, mispaired
heteroduplex DNAs are commonly formed, especially subsequent to
polymerase chain reaction amplification. We investigated a panel of
restriction endonucleases to determine their ability to cleave
mispaired heteroduplex DNA substrates. Two straightforward,
non-radioactive assays are used to evaluate mispaired heteroduplex DNA
cleavage: a PCR amplification method and an oligonucleotide-based
assay. These assays demonstrated that most restriction endonucleases
are capable of site-specific double-strand cleavage with heteroduplex
mispaired DNA substrates, however, certain mispaired substrates do
effectively abrogate cleavage to undetectable levels. These data are
consistent with mispaired substrate cleavage previously reported for
Eco RI and, importantly, extend our knowledge of mispaired heteroduplex
substrate cleavage to 13 additional enzymes.

<>

<1>Langowski, J., Alves, J., Pingoud, A., Maass, G.
<2>Does the specific recognition of DNA by the restriction endonuclease EcoRI involve a linear diffusion step?  Investigation of the processivity of the EcoRI endonuclease.
<3>Nucleic Acids Res.
<4>11
<5>501-513
<6>1983
<7>The time course of the EcoRI endonuclease catalysed cleavage of three
substrates, two plasmid DNAs and one oligonucleotide, each with two EcoRI
sites, was measured.  The two plasmid DNAs with the EcoRI sites 318 and 96 base
pairs apart are cut in a distributive fashion, while the oligonucleotide with
the EcoRI sites 8 base pairs apart is cut in a partially processive manner.  It
is concluded that a linear diffusion of the EcoRI endonuclease on its substrate
across long stretches of DNA is not likely to be operative during the
recognition process.  Microscopic dissociation-reassociation processes,
however, increase the probability of the enzyme to attack further sites located
in the immediate vicinity of a given site.

<>

<1>Langowski, J., Pingoud, A., Goppelt, M., Maass, G.
<2>Inhibition of EcoRI action by polynucleotides.  A characterization of the non-specific binding of the enzyme to DNA.
<3>Nucleic Acids Res.
<4>8
<5>4727-4736
<6>1980
<7>The cleavage of the plasmid pBR322 by the restriction endonuclease EcoRI has
been studied in the presence of various polynucleotides and the double stranded
octanucleotide d-(GGAATTCC) in order to clarify whether there is a preferential
interation of EcoRI with DNA sequences other than -GAATTC-.  The steady state
kinetic analysis shows that all polynucleotides investigated with the possible
exception of poly-dG.poly-dC inhibit the cleavage competitively with Ki values
in the range of 10-4 to 10-5 [M nucleotides].  The Ki of d-(GGAATTCC) is
1.5.10-6 [M nucleotides], indicating that the specific binding is approx. 2
orders of magnitude stronger than non-specific binding.

<>

<1>Langowski, J., Urbanke, C., Pingoud, A., Maass, G.
<2>Transient cleavage kinetics of the EcoRI restriction endonuclease measured in a pulsed quench-flow apparatus:  enzyme concentration-dependent activity change.
<3>Nucleic Acids Res.
<4>9
<5>3483-3490
<6>1981
<7>We report measurements of the cleavage rate of pBR322 plasmid DNA by
restriction endonuclease EcoRI as a function of enzyme and DNA concentration.
The reaction, which at high excess of enzyme over DNA occurs between 0.2 and 5
seconds, was studied by the means of a microprocessor controlled pulsed
quench-flow apparatus.  Enzyme concentrations were between 1 and 100 nM with
DNA concentrations being 3 to 6 nM (specific EcoRI sites).  The catalytic
constants for cleavage of the first and second phosphodiester bonds as measured
at high enzyme concentration both have the same value of 0.35 sec-1 at 21C.  At
enzyme concentrations comparable to or less than DNA concentration, the rate of
the first cleavage is proportional to enzyme concentration, while the second
step is independent of concentration.  At approx. 10 nM EcoRI endonuclease
concentration, a rate increase shows up in both the first and the second
cleavage.  We suggest that this increase is due to the tetramerization reported
by Modrich & Zabel, which occurs in this concentration range.

<>

<1>Lanie, J.A., Ng, W.L., Kazmierczak, K.M., Andrzejewski, T.M., Davidsen, T.M., Wayne, K.J., Tettelin, H., Glass, J.I., Winkler, M.E.
<2>Genome Sequence of Avery's Virulent Serotype 2 Strain D39 of Streptococcus pneumoniae and Comparison with That of Unencapsulated Laboratory Strain  R6.
<3>J. Bacteriol.
<4>189
<5>38-51
<6>2007
<7>Streptococcus pneumoniae (pneumococcus) is a leading human respiratory pathogen that causes a
variety of serious mucosal and invasive diseases.
D39 is an historically important serotype 2 strain that was used in
experiments by Avery and coworkers to demonstrate that DNA is the genetic
material. Although isolated nearly a century ago, D39 remains extremely
virulent in murine infection models and is perhaps the strain used most
frequently in current studies of pneumococcal pathogenesis. To date, the
complete genome sequences have been reported for only two S. pneumoniae
strains: TIGR4, a recent serotype 4 clinical isolate, and laboratory
strain R6, an avirulent, unencapsulated derivative of strain D39. We
report here the genome sequences and new annotation of two different
isolates of strain D39 and the corrected sequence of strain R6.
Comparisons of these three related sequences allowed deduction of the
likely sequence of the D39 progenitor and mutations that arose in each
isolate. Despite its numerous repeated sequences and IS elements, the
serotype 2 genome has remained remarkably stable during cultivation, and
one of the D39 isolates contains only five relatively minor mutations
compared to the deduced D39 progenitor. In contrast, laboratory strain R6
contains 71 single-base-pair changes, six deletions, and four insertions
and has lost the cryptic pDP1 plasmid compared to the D39 progenitor
strain. Many of these mutations are in or affect the expression of genes
that play important roles in regulation, metabolism, and virulence. The
nature of the mutations that arose spontaneously in these three strains,
the relative global transcription patterns determined by microarray
analyses, and the implications of the D39 genome sequences to studies of
pneumococcal physiology and pathogenesis are presented and discussed.

<>

<1>Lanio, T., Jeltsch, A.
<2>PCR-based random mutagenesis method using spiked oligonucleotides to randomize selected parts of a gene without any wild-type background.
<3>Biotechniques
<4>25
<5>958-965
<6>1998
<7>Site-directed mutagenesis is a common tool to analyze protein structure and function.  In
polymerase chain reaction (PCR)-based SDM methods, the mutation is introduced by a PCR primer.
Also, a restriction-enzyme-marker site is often introduced to allow fast and convenient
screening for the presence of the mutation.  Usually, each primer encodes only one particular
amino acid exchange.  In principle then, it is sufficient to identify one marker-positive
clone.  Under these circumstances, the level of background of nonmutated genes is not
important considering that more than approximately 20% of the clones carry the mutation.  If
only 20% of the transformants contained a mutated gene, then screening 10-20 of the clones
would be sufficient to identify at least one mutant, with a probability of 90%-99%.  However,
the approach of introducing nucleotide exchanges into a gene by a PCR primer can be easily
extended to randomize larger parts of the gene if spiked oligonucleotides, which contain some
degree of a mixture of all 4 nucleotides, are used.  Then, one usually intends to isolate as
many mutant clones as possible to obtain as large a library of mutated genes as possible.
Because it is impossible with respect to time and cost to screen more than a few hundred
clones, a method for mutagenesis that excludes the wild-type genes from the transformants with
high confidence is required.

<>

<1>Lanio, T., Jeltsch, A., Pingoud, A.
<2>Towards the design of rare cutting restriction endonucleases: using directed evolution to generate variants of EcoRV differing in their substrate specificity by two orders of magnitude.
<3>J. Mol. Biol.
<4>283
<5>59-69
<6>1998
<7>The restriction endonuclease EcoRV cleaves DNA highly specifically within GATATC sequences. In
order to create EcoRV variants that have an extended recognition site we have employed a
semi-rational random mutagenesis/selection procedure. Twenty-two amino acid residues were
subjected to random mutagenesis and about 500 EcoRV variants representing three generations of
mutants were screened. Among these some highly active variants that strongly prefer AT-flanked
cleavage sites (e.g. S183A/Q224R, T93S/I103F/S183A/T222S or N97T/S183A/T222S) and others that
prefer GC flanks (e.g. K104N/A181T) were identified. As wild-type EcoRV does not discriminate
between these cleavage sites, the generation of these variants represents a significant first
step towards redesigning EcoRV to become an 8 or 10 bp cutter. Such enzymes, only very rarely
found in nature, could be extremely helpful for the manipulation of large DNA fragments.

<>

<1>Lanio, T., Jeltsch, A., Pingoud, A.
<2>On the possibilities and limitations of rational protein design to expand the specificity of restriction enzymes: a case study employing EcoRV as the target.
<3>Protein Eng.
<4>13
<5>275-281
<6>2000
<7>The restriction endonuclease EcoRV has been characterized in structural and functional terms
in great detail. Based on this detailed information we employed a structure-guided approach to
engineer variants of EcoRV that should be able to discriminate between differently flanked
EcoRV recognition sites. In crystal structures of EcoRV complexed with d(CGGGATATCCC)(2) and
d(AAAGATATCTT)(2), Lys104 and Ala181 closely approach the two base pairs flanking the GATATC
recognition site and thus were proposed to be a reasonable starting point for the rational
extension of site specificity in EcoRV [Horton,N.C. and Perona,J.J. (1998) J. Biol. Chem.,
273, 21721-21729]. To test this proposal, several single (K104R, A181E, A181K) and double
mutants of EcoRV (K104R/A181E, K104R/A181K) were generated. A detailed characterization of all
variants examined shows that only the substitution of Ala181 by Glu leads to a considerably
altered selectivity with both oligodeoxynucleotide and macromolecular DNA substrates, but not
the predicted one, as these variants prefer cleavage of a TA flanked site over all other
sites, under all conditions tested. The substitution of Lys104 by Arg, in contrast, which
appeared to be very promising on the basis of the crystallographic analysis, does not lead to
variants which differ very much from the EcoRV wild-type enzyme with respect to the flanking
sequence preferences. The K104R/A181E and K104R/A181K double mutants show nearly the same
preferences as the A181E and A181K single mutants. We conclude that even for the very well
characterized restriction enzyme EcoRV, properties that determine specificity and selectivity
are difficult to model on the basis of the available structural information.

<>

<1>Lanio, T., Jeltsch, A., Pingoud, A.
<2>Automated purification of His6-tagged proteins allows exhaustive screening of libraries generated by random mutagenesis.
<3>Biotechniques
<4>29
<5>338-342
<6>2000
<7>In the course of site-directed mutagenesis or directed evolution
experiments, large numbers of
protein variants are often generated. To characterize functional properties of
individual
mutant proteins in vitro, a rapid and reliable protein purification system is
required. We
have developed an automated method for the parallel purification of 96 different
protein
variants that takes about two hours. Using a 96-well format, the whole process
can be
performed automatically by a pipetting robot. Coupled with a suitable assay,
again using a
96-well format, all variants can be functionally characterized within a few
hours. The protein
purification procedure described here is based on the interaction between His6-
tagged proteins
and Ni-NTA-coated microplates. Typical yields are 3-8 pmol purified
protein/well, which is
sufficient to analyze most enzymatic activities. Using this procedure, we have
purified and
characterized variants of the restriction endonuclease EcoRV, which were
produced in an effort
to enhance the selectivity of this enzyme. For this purpose, three amino acid
residues were
randomized in a region known from the co-crystal structure to be located at the
protein-DNA
interface. From a library of about 1200 variants, predominantly single and
double mutants,
more than 1000 variants were purified and characterized in parallel, which
corresponds to an
almost complete screening of the library.

<>

<1>Lanio, T., Selent, U., Wenz, C., Wende, W., Schulz, A., Adiraj, M., Katti, S.B., Pingoud, A.
<2>EcoRV-T94V: a mutant restriction endonucleaase with an altered substrate specificity towards modified oligodeoxynucleotides.
<3>Protein Eng.
<4>9
<5>1005-1010
<6>1996
<7>Synthetic oligodeoxynucleotides with single methyl phosphonate substitutions were used for an
analysis of the contribution of phosphate contacts to the recognition of the cleavage site by
the restriction endonuclease EcoRV.  Only in the last position within the recognition
sequence, is the methyl phosphonate substitution tolerated by the enzyme.  The wild type
enzyme cleaves the SP diastereomer of the oligodeoxynucleotide GACGATATmpCGTC and the
unmodified sequence with equal rates, whereas the RP diastereomer is cleaved much more slowly.
Inspection of the crystal structure of an EcoRV-DNA complex revealed that the non-bridging
oxygen atoms of the phosphodiester bond between the T and C bases are in hydrogen bonding
distance of the hydroxyl group of the amino acid Thr94.  We therefore tried to engineer a
variant of EcoRV that would prefer a methyl phosphonate linkage over a normal phophodiester
bond and produced mutants with amino acid exchanges at position 94.  One of them, Thr94Val,
shows a dramatically reduced activity towards the unmodified DNA and does not accept the RP
diastereomer, but cleaves the SP diastereomer with the same rate as wild type EcoRV.  Its
selectivity, i.e. the ratio of cleavage rates determined for the unmodified and modified
substrates, differs by three orders of magnitude from that of the wild type enzyme.

<>

<1>Lannoy, N.N., Hoet, P.P., Cocito, C.G.
<2>Cloning of DNA segments of phage 2C, which allows autonomous plasmid replication in Bacillus subtilis.
<3>Eur. J. Biochem.
<4>152
<5>137-142
<6>1985
<7>The chromosome of Bacillus subtilis phage 2C is a 100-MDa double-stranded DNA molecule,
containing hydroxymethyluracil in place of thymine and carrying redundant ends each
encompassing 10% of the genome. 2C DNA was cleaved with EcoRI and HindIII, and cloned in the
shuttle plasmids pSC 540 and pCP 115, both containing segments originating from B. subtilis
and Escherichia coli plasmids. These chimaerical plasmids, carrying the chloramphenicol
resistance gene, were unable to replicate in B. subtilis; this ability was restored, however,
after the insertion of viral DNA segments. Physical maps of the recombinant plasmids were
made; a large deletion of the E. coli-derived segment of pSC 540 was observed (which
paralleled a loss of replication in this host), whereas addition of 2C DNA segments in pCP 115
was not accompanied by deletion (replication in E. coli was conserved in this case). Cloned
viral segments mapped mostly, but not exclusively, within the redundant ends of 2C DNA. It is
suggested that the thirteen recombinant clones carried the replication origin region of phage
2C DNA, and that these sequences originated within or close to the redundant extremities of
the viral chromosome.

<>

<1>Lanois, A., Ogier, J.C., Gouzy, J., Laroui, C., Rouy, Z., Givaudan, A., Gaudriault, S.
<2>Draft Genome Sequence and Annotation of the Entomopathogenic Bacterium Xenorhabdus nematophila Strain F1.
<3>Genome Announcements
<4>1
<5>e00342-13
<6>2013
<7>We report the 4.3-Mb genome sequence of Xenorhabdus nematophila strain F1, a Gram-negative
bacterium that is a symbiont of the entomopathogenic nematode
Steinernema carpocapsae and pathogenic by direct injection for a wide variety of
insects.

<>

<1>Lanzarotti, E., Pellizza, L., Bercovich, A., Foti, M., Coria, S.H., Vazquez, S.C., Ruberto, L., Hernandez, E.A., Dias, R.L., Mac, C.W.P., Cicero, D.O., Smal, C., Nicolas, M.F., Vasconcelos, A.T., Marti, M.A., Turjanski, A.G.
<2>Draft Genome Sequence of Bizionia argentinensis, Isolated from Antarctic Surface Water.
<3>J. Bacteriol.
<4>193
<5>6797-6798
<6>2011
<7>A psychrotolerant marine bacterial strain, designated JUB59(T), was isolated from Antarctic
surface seawater and classified as a new species
of the genus Bizionia. Here, we present the first draft genome sequence
for this genus, which suggests interesting features such as UV resistance,
hydrolytic exoenzymes, and nitrogen metabolism.

<>

<1>Lao, W., Chen, S.
<2>HsaI:  A restriction enzyme from human being.
<3>Sci. Sin.
<4>29
<5>947-953
<6>1986
<7>The HsaI restriction enzyme from the embryos of human, Homo sapiens, has been
isolated with both the tissue extract and nuclear extract.  It proves to be an
unusual enzyme, clearly related functionally to Type II endonuclease.  HsaI
seems to be an isoschizomer of EcoRI, but it has a distinctive property of
elution, differing from EcoRI.  Upon SDS-polyacrylamide gel electrophoresis,
the enzyme preparation showed Coomassie blue staining bands, having the
molecular weight of 65,000 and 22,000 daltons in size, respectively.

<>

<1>Lapidus, A. et al.
<2>Complete genome sequence of Brachybacterium faecium type strain (Schefferle 6-10).
<3>Standards in Genomic Sciences
<4>1
<5>3-11
<6>2009
<7>Brachybacterium faecium Collins et al. 1988 is the type species of the genus, and is of
phylogenetic interest because of its location in the Dermabacteraceae, a
rather isolated family within the actinobacterial suborder Micrococcineae. B.
faecium is known for its rod-coccus growth cycle and the ability to degrade uric
acid. It grows aerobically or weakly anaerobically. The strain described in this
report is a free-living, nonmotile, Gram-positive bacterium, originally isolated
from poultry deep litter. Here we describe the features of this organism,
together with the complete genome sequence, and annotation. This is the first
complete genome sequence of a member of the actinobacterial family
Dermabacteraceae, and the 3,614,992 bp long single replicon genome with its 3129
protein-coding and 69 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Lapidus, A. et al.
<2>Genome sequence of the filamentous, gliding Thiothrix nivea neotype strain (JP2(T)).
<3>Standards in Genomic Sciences
<4>5
<5>398-406
<6>2011
<7>Thiothrix nivea (Rabenhorst 1865) Winogradsky 1888 (Approved Lists 1980) emend. Larkin and
Shinabarger 1983 is the type species of the genus Thiothrix in the
family Thiotrichaceae. The species is of interest not only because of its
isolated location in the yet to be genomically characterized region of the tree
of life, but also because of its life-style with gliding gonidia, the multilayer
sheath, rosettes, and the embedded sulfur granules. Strain JP2(T) is the neotype
strain of the species which was first observed by Rabenhorst in 1865 and later
reclassified by Winogradsky in 1888 into the then novel genus Thiothrix. This is
the first completed (improved-high-quality-draft) genome sequence to be published
of a member of the family Thiotrichaceae. The genome in its current assembly
consists of 15 contigs in four scaffolds with a total of 4,691,711 bp bearing
4,542 protein-coding and 52 RNA genes and is a part of the Genomic Encyclopedia
of Bacteria and Archaea project.

<>

<1>Lapidus, A. et al.
<2>Genome sequence of the moderately thermophilic halophile Flexistipes sinusarabici strain (MAS10).
<3>Standards in Genomic Sciences
<4>5
<5>86-96
<6>2011
<7>Flexistipes sinusarabici Fiala et al. 2000 is the type species of the genus Flexistipes in the
family Deferribacteraceae. The species is of interest because
of its isolated phylogenetic location in a genomically under-characterized region
of the tree of life, and because of its origin from a multiply extreme
environment; the Atlantis Deep brines of the Red Sea, where it had to struggle
with high temperatures, high salinity, and a high concentrations of heavy metals.
This is the fourth completed genome sequence to be published of a type strain of
the family Deferribacteraceae. The 2,526,590 bp long genome with its 2,346
protein-coding and 53 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Lapidus, A., Clum, A., Labutti, K., Kaluzhnaya, M.G., Lim, S., Beck, D.A., Glavina del Rio, T., Nolan, M., Mavromatis, K., Huntemann, M., Lucas, S., Lidstrom, M.E., Ivanova, N., Chistoserdova, L.
<2>Genomes of three methylotrophs from a single niche reveal the genetic and metabolic divergence of the Methylophilaceae.
<3>J. Bacteriol.
<4>193
<5>3757-3764
<6>2011
<7>The genomes of three representatives of the family Methylophilaceae, Methylotenera mobilis
JLW8, Methylotenera versatilis 301 and Methylovorus glucosetrophus SIP3-4, all isolated from a
single study site, Lake Washington in Seattle, were completely sequenced. These were compared
to each other and to the previously published genomes of Methylobacillus flagellatus KT and an
unclassified Methylophilales strain HTCC2181. Comparative analysis revealed that the core
genome of Methylophilaceae may be as small as approximately 600 genes while the pangenome may
be as large as approximately 6000 genes. Significant divergence between the genomes was
uncovered in terms of both gene content and gene and protein conservation, including varied
presence of certain genes involved in methylotrophy. Overall, our data demonstrate that
metabolic potentials can vary significantly between different species of Methylophilaceae,
including organisms inhabiting the very same environment. These data suggest that genetic
divergence among the members of this family may be responsible for their specialized and
non-redundant functions in C1 cycling and in turn suggest means for their successful
co-existence in their specific ecological niches.

<>

<1>Lapkouski, M., Panjikar, S., Janscak, P., Smatanova, I.K., Carey, J., Ettrich, R., Csefalvay, E.
<2>Structure of the motor subunit of type I restriction-modification complex EcoR124I.
<3>Nat. Struct. Mol. Biol.
<4>16
<5>94-95
<6>2009
<7>Type I restriction-modification enzymes act as conventional adenine methylases on
hemimethylated DNAs, but unmethylated recognition targets
induce them to translocate thousands of base pairs before cleaving
distant sites nonspecifically. The first crystal structure of a type I
motor subunit responsible for translocation and cleavage suggests how
the pentameric translocating complex is assembled and provides a
structural framework for translocation of duplex DNA by RecA-like
ATPase motors.

<>

<1>Lapkouski, M., Panjikar, S., Smatanova, I.K., Csefalvay, E.
<2>Purification, crystallization and preliminary X-ray analysis of the HsdR subunit of the EcoR124I endonuclease from Escherichia coli.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>63
<5>582-585
<6>2007
<7>EcoR124I is a multicomplex enzyme belonging to the type I restriction-modification system from
Escherichia coli. Although
EcoR124I has been extensively characterized biochemically, there is no
direct structural information available about particular subunits. HsdR
is a motor subunit that is responsible for ATP hydrolysis, DNA
translocation and cleavage of the DNA substrate recognized by the
complex. Recombinant HsdR subunit was crystallized using the
sitting-drop vapour-diffusion method. Crystals belong to the primitive
monoclinic space group, with unit-cell parameters a = 85.75, b =
124.71, c = 128.37 angstrom, beta = 108.14 degrees. Native data were
collected to 2.6 angstrom resolution at the X12 beamline of EMBL
Hamburg.

<>

<1>Lappalainen, I., Vihinen, M.
<2>Structural basis of ICF-causing mutations in the methyltransferase domain of DNMT3B.
<3>Protein Eng.
<4>15
<5>1005-1014
<6>2002
<7>Mutations in the gene encoding for a de novo methyltransferase, DNMT3B, lead to an autosomal
recessive Immunodeficiency, Centromeric instability and Facial anomalies (ICF) syndrome. To
analyse the protein structure and consequences of ICF-causing mutations, we modelled the
structure of the DNMT3B methyltransferase domain based on Haemophilus haemolyticus protein in
complex with the cofactor AdoMet and the target DNA sequence. The structural model has a
two-subdomain fold where the DNA-binding region is situated between the subdomains on a
surface cleft having positive electrostatic potential. The smaller subdomains of the
methyltransferases differ in length and sequences and therefore only the target recognition
domain loop was modelled to show the location of an ICF-causing mutation. Based on the model,
the DNMT3B recognizes the GC sequence and flips the cytosine from the double-stranded DNA to
the catalytic pocket. The amino acids in the cofactor and target cytosine binding sites and
also the electrostatic properties of the binding pockets are conserved. In addition, a
registry of all known ICF-causing mutations, DNMT3Bbase, was constructed. The structural
principles of the pathogenic mutations based on the modelled structure and the analysis of chi
angle rotation changes of mutated side chains are discussed.

<>

<1>Larbi, D., Decaris, B., Simonet, J.M.
<2>Different bacteriophage resistance mechanisms in Streptococcus salivarius subsp. thermophilus.
<3>J. Dairy Res.
<4>59
<5>349-357
<6>1992
<7>Streptococcus salivarius subsp. thermophilus strain NST5 exhibited a temperature-dependent
defence mechanism against the virulent bacteriophages phiB1.2 and phiA1.1. It was active at 42
degrees C but not at 30 degrees C as demonstrated by a significant increase of both plaque
size and efficiency of plaquing. This defence mechanism did not affect host-dependent phage
replication and did not interfere with phage adsorption to NST5. These results suggest that it
interfered with phage development. The phages phiT33, phiT58, phiD1, phiT21 and phiT9,
belonging to the same phage type as phiB1.2, were examined for their ability to infect NST3
and NST5. Restriction modification systems of different specificity were detected in NST3 and
NST5; host-dependent phage replication was detected at 30 and 42 degrees C; an abortive
defence mechanism was detected in NST5 which was active at 42 degrees C, but not 30 degreesC,
and was independent of restriction modification action or interference with phage adsorption.
Our investigations of phage-host interactions showed that the two Str. salivarius subsp.
thermophilus strains studied avoided attack by related bacteriophages by evolving at least
three different resistance systems.

<>

<1>Larimer, F.W.
<2>Cleavage by ApaI is inhibited by overlapping dcm methylation.
<3>Nucleic Acids Res.
<4>15
<5>9087
<6>1987
<7>
<>

<1>Lark, C., Arber, W.
<2>Host specificity of DNA produced by Escherichia coli.  XIII.  Breakdown of cellular DNA upon growth in ethionine of strains with r15+, r+P1 or r+N3 restriction phenotypes.
<3>J. Mol. Biol.
<4>52
<5>337-348
<6>1970
<7>All methionine-requiring strains of Escherichia coli which have been tested show a two-fold
increase in DNA content when grown in the presence of ethionine or norleucine. In addition, E.
coli strains with the restriction phenotype r15+, rP1+ or rN3+ will degrade their
intracellular DNA when these methionine analogs are substituted for required methionine in the
growth medium. Degradation does not occur in restriction-deficient mutants and in strains
carrying the rK+, rA+ or rB+ phenotypes alone. Breakdown of DNA appears to be a characteristic
of the restriction gene products rather than of the location of these genes on plasmid or
chromosomal DNA molecules.

<>

<1>Larner-Svensson, H., Worning, P., Bartels, M.D., Hestbjerg, H.L., Boye, K., Westh, H.
<2>Complete Genome Sequence of Staphylococcus aureus Strain M1, a Unique t024-ST8-IVa Danish Methicillin-Resistant S. aureus Clone.
<3>Genome Announcements
<4>1
<5>e00336-13
<6>2013
<7>We report the genome sequence, in five contigs, of a methicillin-resistant Staphylococcus
aureus isolate designated M1. This clinical isolate was from the
index patient of a methicillin-resistant Staphylococcus aureus (MRSA) outbreak in
Copenhagen, Denmark, that started in 2003. This strain is sequence type 8 (ST8),
spa type t024, and staphylococcal cassette chromosome mec element (SCCmec) type
IVa.

<>

<1>Larsson, P., Elfsmark, D., Svensson, K., Wikstrom, P., Forsman, M., Brettin, T., Keim, P., Johansson, A.
<2>Molecular evolutionary consequences of niche restriction in Francisella tularensis, a facultative intracellular pathogen.
<3>PLoS Pathog.
<4>5
<5>e1000472
<6>2009
<7>Francisella tularensis is a potent mammalian pathogen well adapted to intracellular habitats,
whereas F. novicida and F. philomiragia are less virulent
in mammals and appear to have less specialized lifecycles. We explored
adaptations within the genus that may be linked to increased host association, as
follows. First, we determined the genome sequence of F. tularensis subsp.
mediasiatica, the only subspecies that had not been previously sequenced. This
genome, and those of 12 other F. tularensis isolates, were then compared to the
genomes of F. novicida (three isolates) and F. philomiragia (one isolate). Signs
of homologous recombination were found in approximately 19.2% of F. novicida and
F. philomiragia genes, but none among F. tularensis genomes. In addition, random
insertions of insertion sequence elements appear to have provided raw materials
for secondary adaptive mutations in F. tularensis, e.g. for duplication of the
Francisella Pathogenicity Island and multiplication of a putative glycosyl
transferase gene. Further, the five major genetic branches of F. tularensis seem
to have converged along independent routes towards a common gene set via
independent losses of gene functions. Our observations suggest that despite an
average nucleotide identity of >97%, F. tularensis and F. novicida have evolved
as two distinct population lineages, the former characterized by clonal structure
with weak purifying selection, the latter by more frequent recombination and
strong purifying selection. F. tularensis and F. novicida could be considered the
same bacterial species, given their high similarity, but based on the
evolutionary analyses described in this work we propose retaining separate
species names.

<>

<1>Lartigue, C., Vashee, S., Algire, M.A., Chuang, R.Y., Benders, G.A., Ma, L., Noskov, V.N., Denisova, E.A., Gibson, D.G., Assad-Garcia, N., Alperovich, N., Thomas, D.W., Merryman, C., Hutchison, C.A.I.I.I., Smith, H.O., Venter, J.C., Glass, J.I.
<2>Creating bacterial strains from genomes that have been cloned and engineered in yeast.
<3>Science
<4>325
<5>1693-1696
<6>2009
<7>We recently reported the chemical synthesis, assembly, and cloning of a bacterial genome in
yeast. To produce a synthetic cell, the genome must be transferred from yeast to a receptive
cytoplasm. Here we describe methods to accomplish this. We cloned a Mycoplasma mycoides genome
as a yeast centromeric plasmid and then transplanted it into Mycoplasma capricolum to produce
a viable M. mycoides cell. While in yeast, the genome was altered by using yeast genetic
systems and then transplanted to produce a new strain of M. mycoides. These methods allow the
construction of strains that could not be produced with genetic tools available for this
bacterium.

<>

<1>Lasek, R., Dziewit, L., Bartosik, D.
<2>Plasmid pP62BP1 isolated from an Arctic Psychrobacter sp. strain carries two highly homologous type II restriction-modification systems and a putative organic sulfate metabolism operon.
<3>Extremophiles
<4>16
<5>363-376
<6>2012
<7>The complete nucleotide sequence of plasmid pP62BP1 (34,467 bp), isolated from Arctic sp. DAB
AL62B, was determined and annotated. The
conserved plasmid backbone is composed of several genetic modules,
including a replication system (REP) with similarities to the REP
region of the iteron-containing plasmid pPS10 of . The additional
genetic load of pP62BP1 includes two highly related type II
restriction-modification systems and a set of genes () encoding enzymes
engaged in the metabolism of organic sulfates, plus a putative
transcriptional regulator (SlfR) of the AraC family. The pP62BP1 has a
compact and unique structure. It is predicted that the enzymes SlfC,
SlfH, SlfS and SlfL carry out a chain of reactions leading to the
transformation of alkyl sulfates into acyl-CoA, with dodecyl sulfate
(SDS) as a possible starting substrate. Comparative analysis of the
nucleotide sequences of pP62BP1 and other spp. plasmids revealed their
structural diversity. However, the presence of a few highly conserved
DNA segments in pP62BP1, plasmid 1 of K5 and pRWF-101 of sp. PRwf-1 is
indicative of recombinational shuffling of genetic information, and is
evidence of lateral gene transfer in the Arctic environment.

<>

<1>Lasko, D.D., Basu, A.K., Kadlubar, F.F., Evans, F.E., Lay, J.O., Essigmann, J.M.
<2>A probe for the mutagenic activity of the carcinogen 4-aminobiphenyl:  synthesis and characterization of an M13mp10 genome containing the major carcinogen-DNA adduct at a unique site.
<3>Biochemistry
<4>26
<5>3072-3081
<6>1987
<7>The duplex genome of Escherichia coli virus M13mp10 was modified at a unique
site to contain N-(deoxyguanosin-8-yl)-4-aminobiphenyl (dG8-ABP), the major
carcinogen-DNA adduct of the human bladder carcinogen 4-aminobiphenyl.  A
tetradeoxynucleotide containing a single dG8-ABP residue was synthesized by
reacting 5'-d(TpGpCpA)-3' with N-acetoxy-N-(trifluoroacetyl)-4-aminobiphenyl,
followed by high-performance liquid chromatography purification of the
principal reaction product 5'-d(TpG8-ABPpCpA)-3' (yield 15-30%).
Characterization by fast atom bombardment mass spectrometry confirmed the
structure as an intact 4-aminobiphenyl-modified tetranucleotide, while 1H
nuclear magnetic resonance spectroscopy established the site of substitution
and the existence of ring stacking between the carcinogen residue and DNA
bases.  Both 5'-d(TpG8-ABPpCpA)-3' and 5'-d(TpGpCpA)-3' were 5'-phosphorylated
by use of bacteriophage T4 polynucleotide kinase and were incorporated into a
four-base gap uniquely positioned in the center of the recognition site for the
restriction endonuclease PstI, in an otherwise duplex genome of M13mp10.  In
the case of the adducted tetranucleotide, dG8-ABP was located in the minus
strand at genome position 6270.  Experiments in which the tetranucleotides were
5' end labeled with [32P] phosphate revealed the following: (i) the adducted
oligomer, when incubated in a 1000-fold molar excess in the presence of T4 DNA
ligase and ATP, was found to be incorporated into the gapped DNA molecules with
an efficiency of approximately 30%, as compared to the unadducted d(pTpGpCpA),
which was incorporated with 60% ligation efficiency; (ii) radioactivity from
the 5' end of each tetranucleotide was physically mapped to a restriction
fragment that contained the PstI site and represented 0.2% of the genome; (iii)
the presence of the lesion within the PstI recognition site inhibited the
ability of PstI to cleave the genome at this site; (iv) in genomes in which
ligation occurred, T4 DNA ligase was capable of covalently joining both
modified and unmodified tetranucleotides to the gapped structures on both the
5' and the 3' ends with at least 90% efficiency.  Evidence also is presented
showing that the dG8-ABP-modified tetranucleotide was stable to the conditions
of the recombinant DNA techniques used to insert it into the viral genome.  On
the basis of these and other data, the dG8-ABP-modified genome was judged to be
a useful probe for investigation of site-specific mutagenesis in E. coli.

<>

<1>Laszlo, V.G., Rimanoczy, I.
<2>Restriction and modification of Shigella flexneri phages by R factors.
<3>Acta Microbiol. Acad. Sci. Hung.
<4>23
<5>259-270
<6>1976
<7>Out of 420 R factors derived from Shigella flexneri strains, 50.8% restricted Escherichia coli
and S. flexneri phages.  Phage restriction was produced both by fi- and fi+ R factors.  The R
factors were divided into nine groups on the basis of the efficiency of plating of S. flexneri
phages.  Changes of phage types were produced by transferring R factors of different
restrictive types.  The changes offered some information concerning the evolution of phage
types.  Studies on phage modification supported the grouping of R factors determined on the
basis of restriction.  R factors of different restrictive types were type-specific except for
types VIII and IX.  Modified phages proved to be highly practical for epidemiological
purposes.  The use of modified phages, as an additional phage-set besides the basic phage-set,
was suggested to trace the source of strains which changed their phage types as an effect of R
factors.

<>

<1>Latif, H., Lerman, J.A., Portnoy, V.A., Tarasova, Y., Nagarajan, H., Schrimpe-Rutledge, A.C., Smith, R.D., Adkins, J.N., Lee, D.H., Qiu, Y., Zengler, K.
<2>The Genome Organization of Thermotoga maritima Reflects Its Lifestyle.
<3>PLoS Genet.
<4>9
<5>E1003485
<6>2013
<7>The generation of genome-scale data is becoming more routine, yet the subsequent
analysis of omics data remains a significant challenge. Here, an approach that
integrates multiple omics datasets with bioinformatics tools was developed that
produces a detailed annotation of several microbial genomic features. This
methodology was used to characterize the genome of Thermotoga maritima-a
phylogenetically deep-branching, hyperthermophilic bacterium. Experimental data
were generated for whole-genome resequencing, transcription start site (TSS)
determination, transcriptome profiling, and proteome profiling. These datasets,
analyzed in combination with bioinformatics tools, served as a basis for the
improvement of gene annotation, the elucidation of transcription units (TUs), the
identification of putative non-coding RNAs (ncRNAs), and the determination of
promoters and ribosome binding sites. This revealed many distinctive properties
of the T. maritima genome organization relative to other bacteria. This genome
has a high number of genes per TU (3.3), a paucity of putative ncRNAs (12), and
few TUs with multiple TSSs (3.7%). Quantitative analysis of promoters and
ribosome binding sites showed increased sequence conservation relative to other
bacteria. The 5'UTRs follow an atypical bimodal length distribution comprised of
"Short" 5'UTRs (11-17 nt) and "Common" 5'UTRs (26-32 nt). Transcriptional
regulation is limited by a lack of intergenic space for the majority of TUs.
Lastly, a high fraction of annotated genes are expressed independent of growth
state and a linear correlation of mRNA/protein is observed (Pearson r = 0.63,
p<2.2x10(-16) t-test). These distinctive properties are hypothesized to be a
reflection of this organism's hyperthermophilic lifestyle and could yield novel
insights into the evolutionary trajectory of microbial life on earth.

<>

<1>Latif, H., Li, H.J., Charusanti, P., Palsson, B.O., Aziz, R.K.
<2>A Gapless, Unambiguous Genome Sequence of the Enterohemorrhagic Escherichia coli  O157:H7 Strain EDL933.
<3>Genome Announcements
<4>2
<5>e00821-14
<6>2014
<7>Escherichia coli EDL933 is the prototypic strain for enterohemorrhagic E. coli serotype
O157:H7, associated with deadly food-borne outbreaks. Because the
publicly available sequence of the EDL933 genome has gaps and >6,000 ambiguous
base calls, we here present an updated high-quality, unambiguous genome sequence
with no assembly gaps.

<>

<1>Lau, E.Y., Bruice, T.C.
<2>Active site dynamics of the HhaI methyltransferase: Insights from computer simulation.
<3>J. Mol. Biol.
<4>293
<5>9-18
<6>1999
<7>A molecular dynamics study was performed on the DNA methyltransferase M.HhaI in a ternary
complex with DNA and AdoMet in solution. Methylation involves addition of the Cys81 sulfhydryl
anion to the 6-position of Cyt18, followed by a nucleophilic attack of the resultant carbanion
at C5 on the AdoMet methyl group. It was found in this simulation that the distances between
the sulfhydryl group (SG) of Cys81 to the C6 of Cyt18 (SG-C6) and methyl carbon (CH3) of
AdoMet to the C5 of cytosine (CH3-C5) are dependent on the dihedral angle chi
(O4'-C1'-N1-C2) of the nucleotide. When the chi angle of Cyt18 is low (<-80 degrees ), the
SG-C6 and CH3-C5 distances are large. A high chi angle (>-80 degrees ) for the target cytosine
residue reduces the distances for both SG-C6 and CH3-C5, and the angles formed between the
cytosine ring and AdoMet correspond well to values for the transition state structures formed
during methylation of cytosine from ab initio calculations. Two possible proton sources for
protonation of N3 of the cytosine residue upon formation of the covalent intermediate were
found in the simulation. The protonated amine group of AdoMet could provide a proton via a
water bridge, or Arg163 could also be the source of the proton for N3 via a water bridge. The
simulation provides insights into how the H5 of cytosine could go from the active site into
solvent. Conserved residues Asn304 and Gln82 stabilize a water network within the active site
of M.HhaI which provides a route for H5 to diffuse into bulk solvent. An initially distant
water molecule was able to diffuse into the active site of the enzyme and replace a position
of a crystallographic water molecule in close proximity to the C5 of cytosine. The movement of
this water molecule showed that a channel exists between Gln82 and the AdoMet in M.HhaI which
allows both water and protons to easily gain access to the active site of the enzyme.

<>

<1>Lau, P.C.K., Forghani, F., Labbe, D., Bergeron, H., Brousseau, R., Holtke, H.J.
<2>The NlaIV restriction and modification genes of Neisseria lactamica are flanked by leucine biosynthesis genes.
<3>Mol. Gen. Genet.
<4>243
<5>24-31
<6>1994
<7>The genes encoding the Neisseria lactamica restriction endonuclease IV (R.NlaIV) and its
cognate DNA methyltransferase (M.NlaIV), both of which recognize the sequence GGNNCC, have
been cloned in Escherichia coli and overexpressed using the T7 polymerase/promoter system.
Analysis of a sequenced 3.58kb fragment established the gene order, leuD-M.NlaIV-R.NlaIV-leuB.
The predicted primary sequence of M.NlaIV (423 amino acids) shows the highest degree of
identity to a pair of cytosine-specific methyltransferases, M.BanI (44.9%) and M.HgiCI
(44.3%), which recognize the sequence GGYRCC (Y, pyrimidines; R, purines). In contrast, the
R.NlaIV protein sequence (243 amino acids) is unique in the existing database, a situation
that holds for most endonucleases. Flanking the NlaIV modification and restriction genes are
homologues of the leuD and leuB genes of enteric bacteria, which code for enzymes in the
leucine biosynthesis pathway. This gene context implies a possible new mode of gene regulation
for the RM.NlaIV system, which would involve a mechanism similar to the recently discovered
leucine/Lrp regulon in E. coli.

<>

<1>Lau, R.H., Doolittle, W.F.
<2>AquI: a more easily purified isoschizomer of AvaI.
<3>FEBS Lett.
<4>121
<5>200-202
<6>1980
<7>Since its first description by Murray and coworkers the restriction
endonuclease AvaI, which recognizes and cleaves at the sequence C^PyCGPuG, has
found wide use in DNA sequence studies.  There are, however, three difficulties
associated with preparation of this enzyme from its usual source, Anabaena
variabilis (Kutzing).  This sheathed filamentous cyanobacterium does not
readily yield single colonies on agar plates and is thus not easily rid of
contaminating bacteria.  It grows slowly (generation times of the order of
24h).  Furthermore, it contains two additional restriction endonucleases, AvaII
and AvaIII.  We frequently find commercial preparations of AvaI to be
contaminated with one of these additional enzymes, and Roizes et al. were
unable to separate AvaI and AvaIII.  We report here that Agmenellum
quadruplicatum (strain PR-6) produces an isoschizomer of AvaI (AquI).  This
unicellular cyanobacterium is readily maintained in axenic condition, grows
rapidly (4-5h generation time at 37C) and contains only this single restriction
endonuclease activity.

<>

<1>Lau, R.H., Visentin, L.P., Martin, S.M., Hofman, J.D., Doolittle, W.F.
<2>Site-specific restriction endonuclease from the filamentous cyanobacterium Nostoc sp. MAC PCC 8009.
<3>FEBS Lett.
<4>179
<5>129-132
<6>1985
<7>We report here the presence of a type-II restriction endonuclease in the
filamentous cyanobacterium Nostoc sp. MAC PCC 8009.  This restriction enzyme,
Nsp MACI, is the first reported isoschizomer of BglII and is very readily
purified from non-specific deoxyribonuclease activity in the crude lysate by
one round of phosphocellulose column chromatography.

<>

<1>Laue, F., Ankenbauer, W., Schmitz, G.
<2>Reducing star activity of restriction enzymes - by cleaving DNA in presence of antibiotic that binds to start sequences.
<3>European Patent Office
<4>EP 449178
<5>
<6>1991
<7>Nonspecific 'star activity' of restriction endonucleases used to cleave DNA molecules by
incubation in a buffer solution, i.e. is reduced by adding an antibiotic which binds to the
DNA at or near the star sequences but not within the SRS. A footprint assay (Biochem.,
28,6259) may be performed to determine if an antibiotic binds to the SRS, and a comparison
between DNA cleavage in the presence and absence of the antibiotic may be performed to
determine if the antibiotic binds to star sequences. The antibiotic is (a) actinomycin D for
BamHI, BfrI, EcoRI, EcoRV, HpaI and XbaI; (b) mithramycin for BamHI and EcoRV; (c) netropsin
for HaeIII, HhaI, PaeR7I, PstI, PvuII, SgrAI, SalI and SstI; (d) distamycin for HaeIII and
HhaI; or (e) Echinomycin for HindII and XbaI. The antibiotic is used at a concentration of
5-200 micromolar.

<>

<1>Laue, F., Ankenbauer, W., Schmitz, G.G., Kessler, C.
<2>The selective inhibitory effect of netropsin on relaxation of sequence specificity of restriction endonuclease SgrAI recognizing 5'-CR^CCGGYG-3'.
<3>Nucleic Acids Res.
<4>18
<5>3421
<6>1990
<7>None

<>

<1>Laue, F., Evans, L.R., Jarsch, M., Brown, N.L., Kessler, C.
<2>A complex family of class-II restriction endonucleases, DsaI-VI, in Dactylococcopsis salina.
<3>Gene
<4>97
<5>87-95
<6>1991
<7>A series of class-II restriction endonucleases (ENases) was discovered in the halophilic,
phototrophic, gas-vacuolated cyanobacterium Dactylococcopsis salina sp. nov. The six novel
enzymes are characterized by the following recognition sequences and cut positions: 5'
-C^CRYGG-3' (DsaI); 5' -GG^CC-3' (DsaII); 5' -R^GATCY-3' (DsaIII); 5' -G^GWCC-3'
(DsaIV); 5' -^CCNGG-3' (DsaV); and 5' -GTMKAC-3' (DsaVI), where W = A or T, M = A or C, K
= G or T, and N = A, G, C or T. In addition, traces of further possible activity were
detected. DsaI has a novel sequence specificity and DsaV is an isoschizomer of ScrFI, but with
a novel cut specificity. A purification procedure was established to separate all six ENases,
resulting in their isolation free of contaminating nuclease activities. DsaI cleavage is
influenced by N6-methyladenine residues [derived from the Escherichia coli-encoded DNA
methyltransferase (MTase) M.Eco damI] within the overlapping sequence 5'-CCRYM/GGATC-3';
DsaV hydrolysis is inhibited by a C-5 methylcytosine residue in its recognition sequence
(5'-CM/CNGG-3'), generated in some DsaV sites by the E. coli-encoded MTase, M.Eco dcmI.

<>

<1>Laurens, N., Bellamy, S.R., Harms, A.F., Kovacheva, Y.S., Halford, S.E., Wuite, G.J.
<2>Dissecting protein-induced DNA looping dynamics in real time.
<3>Nucleic Acids Res.
<4>37
<5>5454-5464
<6>2009
<7>Many proteins that interact with DNA perform or enhance their specific functions by binding
simultaneously to multiple target sites, thereby
inducing a loop in the DNA. The dynamics and energies involved in this
loop formation influence the reaction mechanism. Tethered particle motion
has proven a powerful technique to study in real time protein-induced DNA
looping dynamics while minimally perturbing the DNA-protein interactions.
In addition, it permits many single-molecule experiments to be performed
in parallel. Using as a model system the tetrameric Type II restriction
enzyme SfiI, that binds two copies of its recognition site, we show here
that we can determine the DNA-protein association and dissociation steps
as well as the actual process of protein-induced loop capture and release
on a single DNA molecule. The result of these experiments is a
quantitative reaction scheme for DNA looping by SfiI that is rigorously
compared to detailed biochemical studies of SfiI looping dynamics. We also
present novel methods for data analysis and compare and discuss these with
existing methods. The general applicability of the introduced techniques
will further enhance tethered particle motion as a tool to follow
DNA-protein dynamics in real time.

<>

<1>Laurens, N., Rusling, D.A., Pernstich, C., Brouwer, I., Halford, S.E., Wuite, G.J.
<2>DNA looping by FokI: the impact of twisting and bending rigidity on protein-induced looping dynamics.
<3>Nucleic Acids Res.
<4>40
<5>4988-4997
<6>2012
<7>Protein-induced DNA looping is crucial for many genetic processes such as transcription, gene
regulation and DNA replication. Here, we use
tethered-particle motion to examine the impact of DNA bending and twisting
rigidity on loop capture and release, using the restriction endonuclease FokI as
a test system. To cleave DNA efficiently, FokI bridges two copies of an
asymmetric sequence, invariably aligning the sites in parallel. On account of the
fixed alignment, the topology of the DNA loop is set by the orientation of the
sites along the DNA. We show that both the separation of the FokI sites and their
orientation, altering, respectively, the twisting and the bending of the DNA
needed to juxtapose the sites, have profound effects on the dynamics of the
looping interaction. Surprisingly, the presence of a nick within the loop does
not affect the observed rigidity of the DNA. In contrast, the introduction of a
4-nt gap fully relaxes all of the torque present in the system but does not
necessarily enhance loop stability. FokI therefore employs torque to stabilise
its DNA-looping interaction by acting as a 'torsional' catch bond.

<>

<1>Laurent, J.P., Faske, S., Cangelosi, G.A.
<2>Characterization of IS999, an unstable genetic element in Mycobacterium avium.
<3>Gene
<4>294
<5>249-257
<6>2002
<7>An IS3-family insertion element, IS999, was identified in the opportunistic pathogen
Mycobacterium avium. The 1347 bp element has 29 bp
inverted repeats and two overlapping open reading frames coding for
putative transposases. It was detected in the genomes of ten of 12 M.
avium isolates examined. Copy numbers ranged from four to 16. IS999 is
less stable than IS1245, the most commonly-used marker for typing M. avium
isolates. Among 60 colonies picked from a single patient isolate, there
were two distinct IS1245 restriction fragment length polymorphism banding
patterns compared to eight distinct IS999 patterns (five in one IS1245
group and three in the other). In view of its instability, we asked
whether transposition of IS999 might have phenotypic consequences.
Nucleotide sequence analysis of insertion sites in four isolates revealed
16 putative structural genes that were variably disrupted by IS999.
Insertions into hdhA, a gene that codes for a putative short chain alcohol
dehydrogenase, were distributed non-randomly between colony type variants,
consistent with phenotypic consequences that exert selective pressure.
These observations illustrate the genetic heterogeneity that can exist
within populations of M. avium that appear to be homogeneous by IS1245
analysis. IS999 may be a useful marker for tracking, at the sub-strain
level, the rapid genetic drift that M. avium isolates undergo in nature
and in the laboratory.

<>

<1>Laurie, C.C.
<2>Maize genomic marker set.
<3>US Patent Office
<4>US 7939708 B
<5>
<6>2011
<7>Maize markers useful for genotyping and association studies, e.g. association with oil content
QTLs in populations derived from the Illinois High Oil and Illinois Low Oil maize lines.
Primers and hybridization probes for Taqman assays are provided for 488 SNP markers in 484
loci.

<>

<1>Laurino, P., Tawfik, D.
<2>Engineering DNA methyltransferases for a novel cofactor.
<3>FEBS J.
<4>280
<5>173-174
<6>2013
<7>Cofactors are metals, or organic compounds, which play fundamental roles in enzymes.  Cofactor
engineering has only been partially pursued and rarely have natural cofactors been substituted
with synthetic organic molecules.  Herein, we have designed a synthetic compound presenting in
its structure few key modifications that, in turn, an engineered enzyme can exploit to achieve
cofactor specificity, tight binding and orthogonal recognition of it instead of the natural
cofactor.  The cofactor was designed considering aspects like cell permeability, which allows
extracellular administration of the cofactor, and the possibility to track the enzyme's
product.  The key candidate of our investigation is DNA methyltransferases, that use
S-adenosyl methionine as methyl donor to methylate specific DNA target sequences.  Although
these enzymes are key epigenetic mediators, their genomic targets are often unknown and their
cellular remain poorly understood.  To remodel the catalytic site for the new cofactor, a
protein engineering study ahs been carried out, using computational design as well as directed
evolution.  We believe that the evolved mammalian DNA methylases which will have acquired
orthogonality for the synthetic cofactor, as well as the ability to modify DNA with tractable
groups, will provide new insights regarding the role of this enzyme in epigentics, including
in dictating the genomic methylation patterns in cancer.

<>

<1>Lauro, F.M., Chastain, R.A., Ferriera, S., Johnson, J., Yayanos, A.A., Bartlett, D.H.
<2>Draft Genome Sequence of the Deep-Sea Bacterium Shewanella benthica Strain KT99.
<3>Genome Announcements
<4>1
<5>e00210-13
<6>2013
<7>We report the draft genome sequence of the obligately piezophilic Shewanella benthica strain
KT99 isolated from the abyssal South Pacific Ocean. Strain KT99
is the first piezophilic isolate from the Tonga-Kermadec trench, and its genome
provides many clues on high-pressure adaptation and the evolution of deep-sea
piezophilic bacteria.

<>

<1>Lauro, F.M., Stratton, T.K., Chastain, R.A., Ferriera, S., Johnson, J., Goldberg, S.M., Yayanos, A.A., Bartlett, D.H.
<2>Complete Genome Sequence of the Deep-Sea Bacterium Psychromonas Strain CNPT3.
<3>Genome Announcements
<4>1
<5>e00304-13
<6>2013
<7>Members of the genus Psychromonas are commonly found in polar and deep-sea environments. Here
we present the genome of Psychromonas strain CNPT3.
Historically, it was the first bacterium shown to piezoregulate the composition
of its membrane lipids and to have a higher growth rate at 57 megapascals (MPa)
than at 0.1 MPa.

<>

<1>Lauster, R.
<2>Duplication and variation as a phylogenetic principle of Type II DNA methyltransferases.
<3>Gene
<4>74
<5>243
<6>1988
<7>Meeting Abstract

<>

<1>Lauster, R.
<2>Evolution of type II DNA methyltransferases:  A gene duplication model.
<3>J. Mol. Biol.
<4>206
<5>313-321
<6>1989
<7>On the basis of consensus sequences, which had previously been defined for two
groups of closely related cytosine-specific and adenine-specific DNA
methyltransferases, homologies can be detected that indicate a common origin
for these proteins.  Intramolecular comparisons of several of these enzymes
reveal homology relationships, which suggests that gene duplication is a
phylogenetic principle in the evolution of the Mtases.  One or two duplications
of an ancestral gene encoding a 12,000 to 16,000 Mr protein, followed by
divergent evolution, may have led to very different protein structures and
could explain the differences in amino acid sequences, molecular weights and
biochemical properties.  Intermolecular and intramolecular homologies were also
recognized in type II restriction endonucleases, suggesting a very similar
evolutionary pathway.

<>

<1>Lauster, R.
<2>Close relationship between the HinfI and DpnA DNA-methyltransferase.
<3>Nucleic Acids Res.
<4>17
<5>4402
<6>1989
<7>None

<>

<1>Lauster, R., Kriebardis, A., Guschlbauer, W.
<2>The GATATC-modification enzyme EcoRV is closely related to the GATC-recognizing methyltransferases DpnII and dam from E. coli and phage T4.
<3>FEBS Lett.
<4>220
<5>167-176
<6>1987
<7>The amino acid sequence of EcoRV DNA methyltransferase which methylates the amino group of the
5'-adenine residue of the target sequence GATATC has been found to be closely related to that
of three other adenine methyltransferases, DpnII, dam and damT4, the target sequence of which
is GATC. Despite large differences on the DNA level, the four sequences show four blocks of
homologies. One of these blocks has the sequence DVYXDPPY and is found with little
modification in numerous other DNA methyltransferases. It is speculated that it could be the
binding site of the methyl donor, S-adenosylmethionine. On the other hand, the identification
of a DNA-binding region is more tenuous. As expected, no analogies with (dimeric) repressors
and cro proteins which have the characteristic helix-turn-helix motif have been observed.

<>

<1>Lauster, R., Trautner, T.A., Noyer-Weidner, M.
<2>Cytosine-specific type II DNA methyltransferases:  A conserved enzyme core with variable target-recognizing domains.
<3>J. Mol. Biol.
<4>206
<5>305-312
<6>1989
<7>Comparisons of the amino acid sequences of m5C DNA methyltransferases (Mtases)
from 11 prokaryotes and one eukaryote reveal a very similar organization.
Among all the enzymes one can distinguish highly conserved core sequences and
variable regions.  The core sequences apparently mediate steps of the
methylation reaction that are common to all the enzymes.  The major variable
region has been shown in our previous studies on multispecific phage Mtases to
contain the target-recognizing domains (TRDs) of these enzymes.  Here we have
compared the amino acid sequences of various TRDs from phage Mtases.  This has
revealed the presence of both highly conserved and variable amino acids.  We
postulate that the conserved residues represent a consensus sequence defining a
TRD, whereas the specificity of the TRD is determined by the variable residues.
We have observed similarity between this consensus sequence and sequences in
the variable region of the monospecific Mtases.  We predict that the regions
thus identified represent part of the TRDs of monospecific Mtases.

<>

<1>Lautenberger, J., Eskin, B., Linn, S.
<2>The DNA modification methylase and restriction endonuclease of Escherichia coli B.
<3>Fed. Proc.
<4>31
<5>474
<6>1972
<7>The modification and restriction enzymes from E. coli B have been extensively
purified.  As judged by SDS gel electrophoresis, the methylase contains two
subunits, beta and gamma, of molecular weights 60,000 and 55,000, respectively.
The endonuclease contains beta, gamma and a third subunit, alpha, of molecular
weight 135,000.  These results are consistent with the observations of Hubacek
and Glover [J. Mol. Biol. 50, 11, 1970] showing that the hss and hsm genes are
required for both activities, and in addition, the hsr gene is required for
restriction.  The methylase is isolated in an active 6S form, beta 1 gamma 1,
but it is able to disproportionate into an active 11S form, beta 3 gamma 1.
The endonuclease activity sediments in a broad peak around 15S and contains
alpha, beta, and gamma in the approximate ratio 2.5:2.5:1.  This material is a
mixture of active enzyme species, each containing the three subunits, but in
different proportions.  The methylase forms 6-methylaminopurine using
S-adenosylmethionine (SAM) as the methyl donor.  It acts optimally at pH 5.8,
and is stimulated 3- to 5-fold by Mg++, Mn++, or Ca++, and 30% by ATP.  The
endonuclease requires Mg++, SAM, and ATP, and unlike the methylase, it degrades
massive amounts of ATP to ADP and Pi during its action on DNA.  Both enzymes
are inhibited by S-adenosylethionine or 5'-methylthioadenosine, but not by
S-adenosylhomocysteine.

<>

<1>Lautenberger, J.A., Edgell, M.H., Hutchison, C.A. III
<2>The nucleotide sequence recognized by the BstEII restriction endonuclease.
<3>Gene
<4>12
<5>171-174
<6>1980
<7>The restriction site for the BstEII endonuclease is characterized by the
heptamer sequence: 5'-G-^G-T-N-A-C-C-3' 3'-C-C-A-N-T-G-^G-5' with five
nucleotide long cohesive termini.

<>

<1>Lautenberger, J.A., Edgell, M.H., Hutchison, C.A. III
<2>The DNA sequence on bacteriophage G4 recognized by the Escherichia coli B restriction enzyme.
<3>J. Mol. Biol.
<4>131
<5>871-875
<6>1979
<7>Bacteriophage G4 possesses a single EcoB site located in the overlap between restriction
fragments HinfI-12 and HaeIII-6.  The sequence 5'-T-G-A...8N...T-G-C-T occurs once in this
segment and nowhere else in the DNA sequence of G4.  Four independent G4 mutants that were not
restricted by Escherichia coli B possessed the sequence 5'-T-G-A...8N...T-G-C-C.  The common
sequence shared by the previously mapped EcoB sites on PhiXsB1, simian virus 40, f1, and fd
DNAs is 5'-T-G-A...8N...T-G-C-T...9N...T.  However, the sequence in the region of the G4 EcoB
site contains an A instead of the final T conserved in these other examples.  When the G4 EcoB
site is aligned with the other EcoB sites, there are no conserved residues within 50 bases of
the common sequence, 5'-T-G-A...8N....T-G-C-T, except for those seven residues.  The analysis
of the EcoB site on G4 provides further evidence that only those seven bases are recognized by
the E. coli B restriction enzyme.

<>

<1>Lautenberger, J.A., Kan, N.C., Lackey, D., Linn, S., Edgell, M.H., Hutchison, C.A. III
<2>Recognition site of Escherichia coli B restriction enzyme on PhiXsB1 and simian virus 40 DNAs: An interrupted sequence.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>75
<5>2271-2275
<6>1978
<7>Methyl groups placed on PhiXsB1 replicative form DNA by the Escherichia coli B
modification enzyme are located in the overlap between fragments MboII-3 and
AluI-2, a 61-base-pair DNA segment.  Mutations that led to loss of
susceptibility to restriction by E. coli B occurred within this segment at
three positions spanning 14 nucleotides.  A sequence difference between PhiXsB1
and PhiXam3cs70, a PhiX174 strain not restricted by E. coli B, occurs at one of
these positions.  The site on simian virus 40 DNA methylated by the
modification enzyme is located in the 115-base-pair overlap between fragments
HaeIII-I and AluI-G.  The sequences of these segments of PhiXsB1 and simian
virus 40 DNA and two regions of phage f1 DNA recognized by the E. coli B
restriction enzyme [Ravetch, J.V., Horiuchi, K. & Zinder, N.D. (1978) Proc.
Natl. Acad. Sci. USA 75, 2266-2270] contain a homology of nine bases in the
configuration:  5'-T-G-A...8N...T-G-C-G...9N...T-N-N-T-3'. The sequence
5'-T-G-A...8N...T-G-C-T-3' may constitute the restriction enzyme recognition
site since it does not occur in PhiXam3cs70 DNA and occurs only once in simian
virus 40 DNA, and since all observed mutations leading to loss of the site
occur at one of the bases specified by this sequence.  Analysis of the sequence
of PhiXam3cs70 showed that if no other residues are recognizd, all seven of
these bases are essential for recognition and the interval between the two
groups of specified bases must be precisely eight.

<>

<1>Lautenberger, J.A., Linn, S.
<2>The deoxyribonucleic acid modification and restriction enzymes of Escherichia coli B. I. Purification, subunit structure, and catalytic properties of the modification methylase.
<3>J. Biol. Chem.
<4>247
<5>6176-6182
<6>1972
<7>The modification methylase of Escherichia coli B has been purified to apparent
homogeneity.  The enzyme can exist in several forms, each possessing two
nonidentical polypeptides, b and c, of molecular weights of 60,000 and 55,000,
respectively.  Freshly isolated enzyme has the structure b1c1, but upon storage
at neutral pH and low salt, it disproportionates, producing another form, b3c1.
Treatment at pH 5 converts the mixture of b3c1 and b1c1 to a mixture of b1c1
and another form b2c1, whereas exposure to high salt breaks down the b3c1
structure.  The enzyme is inhibited by S-adenosylethionine and
5'-methylthioadenosine, but not by S-adenosylhomocysteine.  None of these
compounds can replace the required cofactor, S-adenosylmethionine.  The enzyme
can methylate a wide variety of DNAs, but not DNA produced by E. coli B.

<>

<1>Lautenberger, J.A., White, C.T., Haigwood, N.L., Edgell, M.H., Hutchison, C.A.
<2>The recognition site of Type II restriction enzyme BglI is interrupted.
<3>Gene
<4>9
<5>213-231
<6>1980
<7>The Type II restriction endonuclease BglI recognizes the interrupted DNA
sequence 5'-G-C-C-N-N-N-N-N-G-G-C-.  This sequence occurs at all locations in
over 33,000 base pairs of DNA sequence where the enzyme was found to cut DNA
and nowhere else.  All six of the specified bases are essential parts of the
site since all groups of five of the six bases occur in the DNA sequences
tested and none of them are cut by BglI.  The length of the block of
intervening unspecified positions must be exactly five since all other sizes
between zero and 15 occur in the DNA sequences searched and none are cut by
BglI.  The 5'-terminal nucleotides of BglI cleaved phage G4 replicative form
DNA and plasmid pER18 DNA were compared with the DNA sequences near the BglI
sites on these DNAs.  These results indicated that BglI cuts within the
intervening unspecified region and produces single-stranded 3' termini that are
three bases long.  The BglI recognition site and cleavage points can thus be
represented as follows: 5'-G-C-C-N-N-N-N-^-N-G-G-C-3'   This study of the BglI
recognition site was facilitated by the use of inexpensive microcomputers.  A
system of programs was developed that allowed analysis of over 33 kb of DNA
sequences stored on flexible magnetic disks or audio cassettes.  While these
programs were generally written in the higher level language BASIC, some
assembly language subroutines were utilized to reduce executive time.

<>

<1>Laverde-Gomez, J.A., van Schaik, W., Freitas, A.R., Coque, T.M., Weaver, K.E., Francia, M.V., Witte, W., Werner, G.
<2>A multiresistance megaplasmid pLG1 bearing a hylEfm genomic island in hospital Enterococcus faecium isolates.
<3>Int. J. Med. Microbiol.
<4>301
<5>165-175
<6>2011
<7>Enterococcus faecium is considered to be a nosocomial pathogen with
increasing medical importance. The putative virulence factor, hyl(Efm),
encoding a putative hyaluronidase, is enriched among the
hospital-associated polyclonal subpopulation of E. faecium.. The hyl(Efm)
gene is described to be part of a genomic island and was recently
identified to be plasmid-located. Here, we present a description of the
structure, localization, and distribution of the putative pathogenicity
factor hyl(Efm) and its putative island among 39 clinical isolates and
elucidate the composition and host range of pLG1, a hyl(Efm)
multiresistance plasmid of approximately 281.02kb. The hyl(Efm) gene was
located within a 17,824-bp element highly similar to the putative genomic
island (GI) structure that had been previously described. This genomic
region was conserved among 39 hyl(Efm)-positive strains with variation in
a specific region downstream of hyl(Efm) in 18 strains. The putative
hyl(Efm) was located on large plasmids (150-350kb) in 37 strains. pLG1
could be horizontally transferred into four different E. faecium recipient
strains (n=4) but not into E. faecalis (n=3). Sequencing of pLG1 resolved
putative plasmid replication, conjugation, and maintenance determinants as
well as a pilin gene cluster, carbon uptake and utilization genes, heavy
metal and antibiotic resistance clusters. The hyl(Efm) transferable
plasmid pLG1 bears additional putative pathogenicity factors and
antibiotic resistance genes. These findings suggest horizontal gene
transfer of virulence factors and antibiotic resistance gene clusters by a
single genetic event (conjugative transfer) which might be triggered by
heavy antibiotic use common in health care units where E. faecium is
increasingly prevalent.

<>

<1>Lavezzo, E., Toppo, S., Barzon, L., Cobelli, C., Di Camillo, B., Finotello, F., Franchin, E., Peruzzo, D., Toffolo, G.M., Trevisan, M., Palu, G.
<2>Draft genome sequences of two Neisseria meningitidis serogroup C clinical isolates.
<3>J. Bacteriol.
<4>192
<5>5270-5271
<6>2010
<7>Neisseria meningitidis is a human-specific pathogen known for its capability to cause sepsis
and meningitis. Here we report the availability
of 2 draft genome sequences obtained from patients infected during the
same epidemic outbreak. Both bacterial isolates belong to serogroup C, but
their genome sequences show local and remarkable differences compared with
each other or with the reference genome of strain FAM18.

<>

<1>Law, J.A., Jacobsen, S.E.
<2>Establishing, maintaining and modifying DNA methylation patterns in plants and animals.
<3>Nat. Rev. Genet.
<4>11
<5>204-220
<6>2010
<7>Cytosine DNA methylation is a stable epigenetic mark that is crucial for diverse  biological
processes, including gene and transposon silencing, imprinting and X chromosome inactivation.
Recent findings in plants and animals have greatly increased our understanding of the pathways
used to accurately target, maintain and modify patterns of DNA methylation and have revealed
unanticipated mechanistic similarities between these organisms. Key roles have emerged for
small RNAs, proteins with domains that bind methylated DNA and DNA glycosylases in these
processes. Drawing on insights from both plants and animals should deepen our understanding of
the regulation and biological significance of DNA methylation.

<>

<1>Lawrence, M.L., Paulsen, D.B., Scruggs, D.W.
<2>Methods of preparation of live attenuated bacterial vaccines by alteration of DNA adenine methylase (DAM) activity in those bacteria.
<3>International Patent Office
<4>WO 200464776 A
<5>39
<6>2004
<7>Live attenuated bacterial vaccines are provided.  Also provided are methods by which such
vaccines can be obtained, including: a method by which a copy of the dam gene from a
pathogenic bacteria is cloned into a plasmid capable of replication in the same bacterial
species such that it is overexpressed from either a lac promoter, tac promoter, araBAD
promoter, trc promoter, trp promoter, T7, Sp6, or T5 bacteriophage promoters, a native
promoter from that species, or other appropriate promoter.  The plasmid containing the dam
gene is then transferred into the pathogens to cause increased expression of Dam resulting in
the formation of a live attenuated bacterial vaccines.  Alternative methods for producing the
vaccine are also provided including altering or replacing the chromosomal promoter for the
native dam gene so as to alter Dam expression or mutating or replacing the native dam gene so
as to alter the expression of Dam in a pathogenic bacteria.

<>

<1>Lawrence, P.K., Bey, R.F., Wiener, B., Kittichotirat, W., Bumgarner, R.E.
<2>Genome Sequence of a Presumptive Mannheimia haemolytica Strain with an A1/A6-Cross-Reactive Serotype from a White-Tailed Deer (Odocoileus virginianus).
<3>Genome Announcements
<4>2
<5>e00114-14
<6>2014
<7>Mannheimia haemolytica is a Gram-negative bacterium and the principal etiological agent
associated mostly with bovine respiratory disease complex. However, we
report here the sequence of a strain with the novel A1/A6-cross-reactive
serotype, strain PKL10, isolated from white-tailed deer. PKL10 was isolated from
the spleen of farmed white-tailed deer showing clinical signs of pneumonia. The
genome structure of PKL10 is dramatically different from that of previously
sequenced isolates, which was demonstrated by genome alignments. In addition, the
coding sequences in PKL10 share approximately 86% sequence identity with the
coding sequences in other fully sequenced M. haemolytica strains. This suggests
that PKL10 is a novel Mannheimia species.

<>

<1>Lawrence, P.K., Kittichotirat, W., Bumgarner, R.E., McDermott, J.E., Herndon, D.R., Knowles, D.P., Srikumaran, S.
<2>Genome sequences of Mannheimia haemolytica serotype A2: ovine and bovine isolates.
<3>J. Bacteriol.
<4>192
<5>1167-1168
<6>2010
<7>This report describes the genome sequences of Mannheimia haemolytica serotype A2
isolated from pneumonic lungs of two different ruminant species, one from Ovis
aries, designated ovine (O), and the other from Bos taurus, designated bovine
(B).

<>

<1>Lawrence, P.K., Wiener, B.L., Kolander-Bremer, T., Bey, R.F., Stine, D.L., Kittichotirat, W., Bumgarner, R.E.
<2>Genome-Wide Association Studies of Virulent and Avirulent Haemophilus parasuis Serotype 4 Strains.
<3>Genome Announcements
<4>2
<5>e00884-14
<6>2014
<7>Haemophilus parasuis is a normal commensal of the upper respiratory tract of healthy pigs.
However, in conjunction with stress and/or viral infections, or in
immunocompromised animals, H. parasuis can transform into a pathogen causing
Glasser's disease, which is typically characterized by fibrinous polyserositis,
polyarthritis, meningitis, and sometimes acute pneumonia and septicemia. H.
parasuis serotype 5 is highly virulent and more frequently isolated from
respiratory and systemic infection in pigs. Recently Newport Laboratories
isolated highly virulent H. parasuis serotype 4 strains from the tissues of
diseased pigs. This study was undertaken to identify the genes responsible for H.
parasuis serotype 4 virulence. To achieve this objective we performed genome-wide
association studies (GWAS) across two virulent and three avirulent H. parasuis
serotype 4 strains.

<>

<1>Lawrenz, M.B., Kawabata, H., Norris, S.J.
<2>Plasmid content determines the ability to transform Borrelia burgdorferi.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>102
<5>179
<6>2002
<7>The progress of research on Borrelia burgdorferi, the causative agent of Lyme disease, has
been hampered by the lack of genetic techniques that can
be used in the laboratory. Recent advancements in this field have included
the development of a reliable selection marker and the creation of both
stable shuttle vectors and a suicide vector to truncate linear plasmids.
To date, many researchers have published successful genetic manipulation
of high passage isolates of B. burgdorferi, but only very limited success
has been reported with low passage clones. Along with increased
transformation efficiency, high passage clones have also lost the ability
to infect the mammalian host, due to loss of plasmids during in vitro
passage. In this study, we have utilized the pBSV2 shuttle vector to
transform a library of low passage clones of B. burgdorferi B31 to
determine if the increased transformation efficiency seen in high passage
clones can be attributed to loss of one of the plasmids. Three
transformation phenotypes were identified that correlate with the presence
or absence of lp25 and/or lp56. Clones that lacked both plasmids yielded
greater than 1000 transformants per mug of DNA, whereas isolates that
possessed both plasmids yielded 0 to 5 transformants per mug of DNA. B.
burgdorferi that lacked either lp25 or lp56 had an intermediate
transformation efficiency. To date, all transformants isolated from
lp25-positive clones electroporated with pBSV2 lack lp25, consistent with
selective transformation of individual cells in which lp25 was missing.
This information indicates that lp25 and lp56 represent important barriers
against successful B. burgdorferi transformation, which may be related to
plasmid-encoded restriction modification systems.

<>

<1>Lawrenz, M.B., Kawabata, H., Purser, J.E., Norris, S.J.
<2>Decreased electroporation efficiency in Borrelia burgdorferi containing linear plasmids lp25 and lp56: impact on transformation of infectious  B. burgdorferi.
<3>Infect. Immun.
<4>70
<5>4798-4804
<6>2002
<7>The presence of the linear plasmids lp25 and lp56 of Borrelia burgdorferi B31 was found to
dramatically decrease the rate of transformation by electroporation with the shuttle vector
pBSV2, an autonomously replicating plasmid that confers kanamycin resistance (P. E. Stewart,
R. Thalken, J. L. Bono, and P. Rosa, Mol. Microbiol. 39:714-721, 2001). B. burgdorferi B31
clones had transformation efficiencies that were either low, intermediate, or high, and this
phenotype correlated with the presence or absence of lp25 and lp56. Under the conditions
utilized in this study, no transformants were detected in clones that contained both lp25 and
lp56; the few kanamycin-resistant colonies isolated did not contain pBSV2, indicating that the
resistance was due to mutation. Intermediate electroporation rates (10 to 200 colonies per
micro g of DNA) were obtained with B31 clones that were either lp25(-) and lp56(+) or lp25(+)
and lp56(-). Clones in this group that initially contained lp25 lacked this plasmid in pBSV2
transformants, a finding consistent with selective transformation of lp25(-) variants. High
transformation rates (>1,000 colonies per micro g of DNA) occurred in clones that lacked both
lp25 and lp56. Sequence analysis indicated that lp25 and lp56 contain genes that may encode
restriction and/or modification systems that could result in the low transformation rates
obtained with strains containing these plasmids. The previously reported correlation between
lp25 and infectivity in mice, coupled with the barrier lp25 presents to transformation, may
explain the difficulty in obtaining virulent transformants of B. burgdorferi.

<>

<1>Lazarev, V.N. et al.
<2>Complete Genome and Proteome of Acholeplasma laidlawii.
<3>J. Bacteriol.
<4>193
<5>4943-4953
<6>2011
<7>We present the complete genome sequence and proteogenomic map for Acholeplasma laidlawii
PG-8A: class Mollicutes, order Acholeplasmatales,
family Acholeplasmataceae. The genome of A. laidlawii is represented by a
single 1 496 992 b.p. circular chromosome with the average G+C content of
31 mol%. This is the longest genome among the Mollicutes with a known
nucleotide sequence. It contains genes of polymerase type I, SOS-response,
and signal transduction systems, as well as RNA regulatory elements,
riboswitches and T-boxes. This demonstrates a significant capability for
the regulation of gene expression and mutagenic response to stress.
Acholeplasma laidlawii and phytoplasmas are the only Mollicutes known to
use the universal genetic code, in which UGA is a stop codon. Within the
Mollicutes group, only the sterol-nonrequiring Acholeplasma has the
capacity to synthesize saturated fatty acids de novo. Proteomic data was
used in the primary annotation of the genome, validating expression of
many predicted proteins. We also detected post-translational modifications
of A.laidlawii proteins: phosphorylation and acylation. Seventy four
candidate phosphorylated proteins were found: sixteen candidates are
proteins unique to A. laidlawii, and eleven of them are surface-anchored
or integral membrane proteins, which implies the presence of active
signaling pathways. Among twenty acylated proteins, fourteen contained
palmitic chains, and six, stearic chains. No residue of linoleic or oleic
acid was observed. Acylated proteins were mainly components of sugar and
inorganic ion transport systems, and were surface-anchored proteins with
unknown functions.

<>

<1>Lazarevic, V., Dusterhoft, A., Soldo, B., Hilbert, H., Mauel, C., Karamata, D.
<2>Nucleotide sequence of the Bacillus subtilis temperate bacteriophage SPbetaetac2.
<3>Microbiology
<4>145
<5>1055-1067
<6>1999
<7>The Bacillus subtilis 168 chromosomal region extending from 184 degrees to 195 degrees,
corresponding to prophage SPbeta, has been completely sequenced using DNA of the
thermoinducible SPbetac2 mutant.  This 134416 bp segment comprises 187 putative ORFs which,
according to their orientation, were grouped into three clusters.  Compared to its host,
SPbetac2 is characterized by a lower G&C content, shorter mean ORF length, as well as a
different usage of start codons.  Nearly 75% of predicted ORFs do not share significant
homologies to sequences in available databases.  The only highly similar proteins to
SPbetac2-encoded ones are host paralogues.  SPbetac2 promoter regions contain SOS box
consensus sequences and a repeated motif, designated SPbeta repeated element, that is absent
from the host genome.  Gene sspC, encoding the small acid-soluble protein C, that has been
previously sequenced and mapped to the vicinity of the SPbeta region, was found to be part of
the prophage.

<>

<1>Lazarevic, V., Soldo, B., Dusterhoft, A., Hilbert, H., Mauel, C., Karamata, D.
<2>Introns and intein coding sequence in the ribonucleotide reductase genes of Bacillus subtilis temperate bacteriophage SPbeta.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>1692-1697
<6>1998
<7>The two putative ribonucleotide reductase subunits of the Bacillus subtilis bacteriophage
SPbeta are encoded by the bnrdE and bnrdF genes that are highly similar to corresponding host
paralogs, located on the opposite replication arm.  In contrast to their bacterial
counterparts, bnrdE and bnrdF each are interrupted by a group I intron, efficiently removed in
vivo by mRNA processing.  The bnrdF intron contains an ORF encoding a polypeptide similar to
homing endonucleases responsible for intron mobility, whereas the bnrdE intron has no obvious
trace of coding sequence.  The downstream bnrdE exon harbors an intervening sequence not
excised at the level of the primary transcript, which encodes an in-frame polypeptide
displaying all the features of an intein.  Presently, this is the only intein identified in
bacteriophages.  In addition, bnrdE provides an example of a group I intron and an intein
coding sequence within the same gene.

<>

<1>Lazareviciute, L., Maneliene, Z., Padegimiene, A., Kiuduliene, L., Laucys, V., Bitinaite, J., Gruber, I.M., Polyachenko, V.M., Butkus, V., Janulaitus, A.
<2>Isolation and characteristics of new restriction endonucleases from Haemophilus influenzae.
<3>Bioorg. Khim.
<4>16
<5>889-897
<6>1990
<7>Various strains of Haemophilus influenzae have been examined for the presence
of site-specific endonuclease activities, and eleven restriction endonucleases
have been isolated from seven strains.  For all the endonucleases recognition
sequences were determined, for three of them cleavage sites being identified.
The enzymes proved to be isoschizomers of known endonucleases, viz. Hin1I,
Hin8I - AcyI; Hin1II, Hin8II-NlaIII; Hin2I, Hin5I-HpaII; Hin3I-CauII;
Hin5II-AsuI; Hin5III-HindIII; Hin6I, Hin7I-HhaI.  Restriction endonucleases
Hin1I, Hin1II and Hin6I recognize nucleotide sequences 5'GR^CGYC, 5'CATG^,
5'G^CGC, respectively, and cleave them as indicated by the arrows.

<>

<1>Lazareviciute, L.G., Butkus, V.V., Maneliene, Z.P., Kiuduliene, L.J., Bitinaite, J.B., Laucys, V.S., Gruber, I.M., Polyachenko, V.M., Janulaitis, A.
<2>Preparation of restriction endonuclease - capable of recognising and splitting specified sequences of nucleotide(s) in DNA, using an Haemophilus influenzae strain.
<3>Soviet Patent Office
<4>SU 1458388
<5>
<6>1989
<7>Restriction endonucleases Hin1I and Hin1II capable or recognising and splitting the nucleotide
sequences 5'-GPuGGYyG-3' and 5'-CATG-3' respectively, are produced by the strain of
Haemophilus influenzae RFL I (VKPM B-4035). The strain is grown with aeration until the
optical density of the cell suspension reaches 1.8-2.5. Disintegration of the cells with
ultrasound and centrifuging is followed by chromatography of the extract on phosphocellulose,
and chromatographic purification of the restrictases Hin1I and Hin1II on prescribed sorbents.
The yields are 15000 and 1000 units/g. of biomass respectively.

<>

<1>Lazareviciute, L.G., Padegimiene, A.M., Kiuduliene, L.J., Laucys, V.C., Bitinaite, J.B., Gruder, I.M., Polyachenko, V.M., Gordachev, I.D., Butkus, V.V., Janulaitis, A.
<2>Restriction endonuclease splitting nucleotide sequence - is obtained by using Haemophilus influenzae as producer.
<3>Soviet Patent Office
<4>SU 1576565 A
<5>
<6>1990
<7>Use of Haemophilus influenzae (RFL 6) VKPM B-4275 as producer of restriction endonuclease
Hin61 which splits the nucleotide sequence 5'-G^CGC-3', increases the efficiency of the
method. Growing in specified medium is followed by disintegrating the cells, centrifuging, and
purifying chromatographically in three stages, on phosphocellulose, heparin-sepharose and
blue sepharose in K phosphate buffer, pH 7.4-7.5. The first stage elution uses 0.1-0.8M NaCl
gradient, and the second and third stage uses the 0.0-1.0M NaCl gradient.

<>

<1>Lazim, H., Josephsen, J., Ben Hassen, A., Belhadj, O., Limam, R.
<2>Eco1524I, a type II restriction endonuclease - Isolation, partial purification, and characterization.
<3>Appl. Biochem. Biotechnol.
<4>125
<5>189-199
<6>2005
<7>Various strains of Escherichia coli, isolated from different patients, were screened for type
II restriction endonuclease activity. In 1 out
of 23 patients, a type II restriction endonuclease activity was found.
The restriction endonuclease designated Eco1524I was purified to near
homogeneity, based on hydroxyapatite and heparin sepharose
chromatography. Eco1524I exhibited endonuclease restriction activity in
the pH range from 6.0 to 10.0 (maximum level at pH 8.0) and required
Mg2+ as divalent cation. The enzyme was stable till temperature 55
degrees C and pH range from 6.0 to 10.0. Eco1524I recognized the
sequence 6-bp palindromic 5'AGG|CCT 3', producing blunt end
and is found to be an isoschizomer of StuI.

<>

<1>Lazowska, J., Meunier, B., Macadre, C.
<2>Homing of a group II intron in yeast mitochondrial DNA is accompanied by unidirectional co-conversion of upstream-located markers.
<3>EMBO J.
<4>13
<5>4963-4972
<6>1994
<7>Group II introns ai1 and ai2 of the Saccharomyces cerevisiae mitochondrial COX1 gene encode
proteins having a dual function (maturase and reverse transcriptase) and are mobile genetic
elements.  By construction of adequate donor genomes, we demonstrate that each of them is
self-sufficient and practices homing in the absence of homing-type endonucleases encoded by
either group I introns or the ENS2 gene.  Each of the S. cerevisiae group II self-mobile
introns was tested for its ability to invade mitochondrial DNA (mtDNA) from two related
Saccharomyces species.  Surprisingly, only ai2 was observed to integrate into both genomes.
The non-mobility of ai1 was clearly correlated with some polymorphic changes occurring in
sequences flanking its insertion sites in the recipient mtDNAs.  Importantly, studies of the
behavior of these introns in interspecific crosses demonstrate that flanking marker
co-conversion accompanying group II intron homing is unidirectional and efficient only in the
3' to 5' direction towards the upstream exon.  Thus, the polar co-conversion and dependence
of the splicing proficiency of the intron reported previously by us are hallmarks of group II
intron homing, which significantly distinguish it from the strictly DNA-based group I intron
homing and strictly RNA-based group II intron transposition.

<>

<1>Lazowska, J., Szczepanek, T., Macadre, C., Dakova, M.
<2>Two homologous mitochondrial introns from closely related Saccharomyces species differ by only a few amino acid replacements in their open reading frames: one is mobile, the other is not.
<3>C.R. Acad. Sci. III
<4>315
<5>37-41
<6>1992
<7>We have undertaken a comprehensive study of the gene conversion of all the mitochondrial
introns of Saccharomyces capensis.  The approach used involved the measurement of intron
transmission amongst the progeny of crosses between a recipient strain (Saccharomyces
cerevisiae intronless mitochondria) and various donor strains (Saccharomyces capensis, with
various combinations of mitochondrial introns).  We have shown that the S. capensis second
intron (bi2 of cytochrome b gene) is extremely active as a donor in gene conversion whereas
its homologous S. cerevisiae intron is not.  Determination of the sequence of the S. capensis
intron demonstrates that it differs from that of the homologous S. cerevisiae intron (bi2) by
a very small number of nucleotide substitutions.

<>

<1>Le Bars, H., Bousarghin, L., Bonnaure-Mallet, M., Jolivet-Gougeon, A., Barloy-Hubler, F.
<2>Complete Genome Sequence of the Strong Mutator Salmonella enterica subsp. enterica Serotype Heidelberg Strain B182.
<3>J. Bacteriol.
<4>194
<5>3537-3538
<6>2012
<7>In bacteria, normal mutation frequencies are mostly around 10(-10) per base pair. However,
there exists natural isolates, called 'mutators,' that exhibit permanent
mutation occurrences up to 1,000-fold greater than usual. As mutations play
essential roles, particularly in the evolution of antibiotic resistance, bacteria
showing elevated mutation rates could have an important responsibility in the
emergence of antibiotic resistance, especially in the clinical background. In
this announcement, we report the first complete genome sequence of the Salmonella
enterica subsp. enterica serotype Heidelberg B182 mutator strain, isolated from
bovine feces (France), which consists of a 4,750,465-bp circular chromosome
(cB182_4750; GC, 52.2%) and one circular plasmid of 37,581 bp (pB182_37; GC,
42.8%).

<>

<1>Le Gac, G., Esteve, P.-O., Ferec, C., Pradhan, S.
<2>DNA damage-induced down-regulation of human Cdc25C and Cdc2 is mediated by cooperation between p53 and maintenance DNA (Cytosine-5) methyltransferase 1.
<3>J. Biol. Chem.
<4>281
<5>24161-24170
<6>2006
<7>The Cdc25C phosphatase mediates cellular entry into mitosis in mammalian cells. Cdc25C
activates Cdc2 for entry into mitosis by dephosphorylating Thr and Tyr at the site of
inhibitory phosphorylation. The Cdc25C gene contains tumor suppressor p53 binding sites and is
demonstrated to contribute to the p53-dependent cell cycle arrest upon DNA damage. Here we
show that both Cdc25C and Cdc2 were down-regulated in wild-type HCT116 cells but not in
p53-null, DNMT1-null or DNMT1and DNMT3b-null cells, upon p53 stabilization following
doxorubicin-mediated DNA damage. Furthermore, zebularine, a drug that selectively traps and
depletes nuclear DNMT1 and DNMT3b, relieved p53-mediated repression of endogenous Cdc25C and
Cdc2. Methylation analysis of the Cdc25C and Cdc2 promoter displayed internal CG methylation
proximal to the p53 binding site upon DNA damage in a p53-dependent manner. Chromatin
immunoprecipitation of doxorubicin treated wild-type HCT116 cells showed the presence of
DNMT1, p53, H3K9me2, and the transcriptional repressor HDAC1 on the Cdc25C and Cdc2 promoters,
suggesting their involvement as repressive complexes in Cdc25C and Cdc2 gene silencing. Thus,
the general mechanism of p53-mediated gene repression may involve recruitment of other
repressive factors.

<>

<1>Le Guen, L., Santos, R., Camadro, J.-M.
<2>Functional analaysis of the hemK gene product involvement in protoporphyrinogen oxidase activity in yeast.
<3>FEMS Microbiol. Lett.
<4>173
<5>175-182
<6>1999
<7>The Escherichia coli hemK gene has been described as being involved in protoporphyrinogen
oxidase activity; however, there is no biochemical evidence for this. In the context of
characterizing the mechanisms of protoporphyrinogen oxidation in the yeast Saccharomyces
cerevisiae, we investigated the yeast homolog of HemK, which is encoded by the ORF YNL063w, to
find out whether it has any protoporphyrinogen oxidase activity and/or whether it modulates
protoporphyrinogen oxidase activity. Phenotype analysis and enzyme activity measurements
indicated that the yeast HemK homolog is not involved in protoporphyrinogen oxidase activity.
Complementation assays in which the yeast HemK homolog is overproduced do not restore
wild-type phenotypes in a yeast strain with deficient protoporphyrinogen oxidase activity.
Protein sequence analysis of HemK-related proteins revealed consensus motif for
S-adenosyl-methionine-dependent methyltransferase.

<>

<1>Le Marechal, C., Hernandez, D., Schrenzel, J., Even, S., Berkova, N., Thiery, R., Vautor, E., Fitzgerald, J.R., Francois, P., Le Loir, Y.
<2>Genome Sequence of two Staphylococcus aureus ovine strains that induce severe (strain O11) and mild (strain O46) mastitis.
<3>J. Bacteriol.
<4>193
<5>2353-2354
<6>2011
<7>Staphylococcus aureus is a major etiological agent of mastitis in ruminants. We report here
the genome sequences of two ovine strains that were isolated from gangrenous (strain O11) and
subclinical (strain O46) ewe mastitis. Both strains belong to the same clonal complex. Despite
this close genotypic relationship, the two isolates were shown to reproducibly induce highly
divergent type of infections, either severe (O11) or mild (O46) mastitis in experimental ewe
model.

<>

<1>le Roux, W.J., Chan, W.Y., De Maayer, P., Venter, S.N.
<2>Genome Sequence of Vibrio cholerae G4222, a South African Clinical Isolate.
<3>Genome Announcements
<4>1
<5>e0004013
<6>2013
<7>Vibrio cholerae, a Gram-negative pathogen autochthonous to the aquatic environment, is the
causative agent of cholera. Here, we report the complete
genome sequence of V. cholerae G4222, a clinical isolate from South Africa.

<>

<1>Le, Y., Zhu, S.
<2>Purification of restriction endonuclease MspI.
<3>Huaxue Shiji
<4>10
<5>35-37
<6>1988
<7>A new procedure has been developed for the purification of restriction
endonuclease Msp I.  The procedure uses Sephadex G-200 gel filtration,
chromatography on phosphocellulose and heparin sepharose, and gives product
with sufficient purity to permit its use in genetic engineering.

<>

<1>Leahy, S.C., Kelly, W.J., Altermann, E., Ronimus, R.S., Yeoman, C.J., Pacheco, D.M., Li, D., Kong, Z., McTavish, S., Sang, C., Lambie, S.C., Janssen, P.H., Attwood, G.T.
<2>The genome sequence of the rumen methanogen Methanobrevibacter ruminantium M1.
<3>PLoS ONE
<4>5
<5>e8926
<6>2010
<7>Background: Methane (CH4) is a potent greenhouse gas (GHG), having a global warming potential
21 times that of carbon dioxide (CO2). Methane emissions from agriculture represent around 40%
of the emissions produced by human-related activities, the single largest source being enteric
fermentation, mainly in ruminant livestock. Technologies to reduce these emissions are
lacking. Ruminant methane is formed by the action of methanogenic archaea typified by
Methanobrevibacter ruminantium, which is present in ruminants fed a wide variety of diets
worldwide. To gain more insight into the lifestyle of a rumen methanogen, and to identify
genes and proteins that can be targeted to reduce methane production, we have
sequenced the 2.93 Mb genome of M. ruminantium M1, the first rumen methanogen genome to be
completed.  Methodology/Principal Findings: The M1 genome was sequenced, annotated and
subjected to comparative genomic and
metabolic pathway analyses. Conserved and methanogen-specific gene sets suitable as targets
for vaccine development or chemogenomic-based inhibition of rumen methanogens were identified.
The feasibility of using a synthetic peptidedirected vaccinology approach to target epitopes
of methanogen surface proteins was demonstrated. A prophage genome was described and its lytic
enzyme, endoisopeptidase PeiR, was shown to lyse M1 cells in pure culture. A predicted
stimulation of M1 growth by alcohols was demonstrated and microarray analyses indicated
up-regulation of methanogenesis genes during co-culture with a hydrogen (H2) producing rumen
bacterium. We also report the discovery of non-ribosomal peptide synthetases in M. ruminantium
M1, the first reported in archaeal species.  Conclusions/Significance: The M1 genome sequence
provides new insights into the lifestyle and cellular processes of this important rumen
methanogen. It also defines vaccine and chemogenomic targets for broad inhibition of rumen
methanogens and represents a significant contribution to worldwide efforts to mitigate
ruminant methane emissions and reduce production of anthropogenic greenhouse gases.

<>

<1>Leahy, S.C., Kelly, W.J., Li, D., Li, Y., Altermann, E., Lambie, S.C., Cox, F., Attwood, G.T.
<2>The Complete genome sequence of Methanobrevibacter sp. AbM4.
<3>Standards in Genomic Sciences
<4>8
<5>215-227
<6>2013
<7>Methanobrevibacter sp. AbM4 was originally isolated from the abomasal contents of a sheep and
was chosen as a representative of the
Methanobrevibacter wolinii clade for genome sequencing. The AbM4 genome
is smaller than that of the rumen methanogen M. ruminantium M1 (2.0 Mb
versus 2.93 Mb), encodes fewer open reading frames (ORFs) (1,671 versus
2,217) and has a lower G+C percentage (29% versus 33%). Overall, the
composition of the AbM4 genome is very similar to that of M1 suggesting
that the methanogenesis pathway and central metabolism of these strains
are highly similar, and both organisms are likely to be amenable to
inhibition by small molecule inhibitors and vaccine-based methane
mitigation technologies targeting these conserved features. The main
differences compared to M1 are that AbM4 has a complete coenzyme M
biosynthesis pathway and does not contain a prophage or non-ribosomal
peptide synthase genes. However, AbM4 has a large CRISPR region and
several type I and type II restriction-modification system components.
Unusually, DNA-directed RNA polymerase beta' and beta ' subunits of
AbM4 are joined, a feature only previously observed in some
thermophilic archaea. AbM4 has a much reduced complement of genes
encoding adhesin-like proteins which suggests it occupies a ruminal
niche different from that of M1.

<>

<1>Leao, S.C., Matsumoto, C.K., Viana-Niero, C., Ramos, R.T., Carneiro, A.R., Barbosa, M.S., Lima, K.V., Lopes, M.L., Azevedo, V., Silva, A.
<2>Draft Genome Sequence of Mycobacterium abscessus subsp. bolletii INCQS 00594.
<3>Genome Announcements
<4>1
<5>e00896-13
<6>2013
<7>An epidemic of surgical-site infections by a single strain of Mycobacterium abscessus subsp.
bolletii affected >1,700 patients in Brazil from 2004 to 2008.
The genome of the epidemic prototype strain M. abscessus subsp. bolletii INCQS
00594, deposited in the collection of the National Institute for Health Quality
Control (INCQS), was sequenced.

<>

<1>Lebedev, L.R., Afinogenova, G.N., Andreeva, I.S., Pustoshilova, N.M., Pozdnyakov, S.G., Chizhikov, V.E.
<2>Determination of substrate specificity of a site-specific deoxyribonuclease RtrI.
<3>Bioorg. Khim.
<4>17
<5>277-279
<6>1991
<7>A new site-specific deoxyribonuclease RtrI from Rhizobium trifolii has been
shown to recognize the sequence 5'-G^TCGAC-3' in double-stranded DNA and to
cleave it at the point indicated by an arrow.  Therefore the enzyme is a true
isoschizomer of the restriction endonuclease SalI.

<>

<1>Lebedev, L.R., Andreeva, I.S., Gordienko, N.Y., Maystrenko, G.G., Repin, V.E., Pustoshilova, N.M.
<2>The bacterial strain Rhizobium leguminosarum is a producer of the restriction enzyme Rle69I.
<3>Russian Patent Office
<4>RU 2001952 C
<5>
<6>1993
<7>Isolation, characterization and purification of Rle69I

<>

<1>Lebedev, L.R., Pustoshilova, N.M., Repin, V.E., Degtyarev, S.K.
<2>Isolation of restriction endonuclease RsaI from Rhodopseudomonas sphaeroides and study of its properties.
<3>Prikl. Biokhim. Mikrobiol.
<4>27
<5>330-337
<6>1991
<7>A technique is proposed for the isolation of the restrictase RsaI from
Rhodopseudomonas sphaeroides, which involves cultivation of the bacteria under
aerobic conditions, ultrasonic disruption of the cells, fractionation in the
PEG-dextran two-phase system, chromatography on phosphocellulose P-11 and
heparin-sepharose.  The enzyme yield from 1 g of wet cells is 35000 U.
Gel-filtration through Sephadex G-200 and electrophoresis in polyacrylamide gel
under denaturing conditions showed that RsaI in solution was a monomer with a
molecular weight of 24000 +/- 2000.  The optimal conditions for the highest
enzymatic activity are the following: NaCl concentration 0-20 mM; Mg2+
concentration 10-12 mM, pH 8.0-8.5; 37-50C.  Mg2+ ions cannot be replaced with
Mn2+, Zn2+, Ca2+ and Cu2+ ions.  Glycerol and ethanol at concentrations up to
10%, and p-chloromercuric benzoate at concentrations up to 0.3 mM have no
inhibitory effect.

<>

<1>Lebenka, A.I., Melvidas, V.I.
<2>The sensitivity of E. coli and C. freundii strains to 5-azacytidine.
<3>Vopr. Med. Khim.
<4>47
<5>477-482
<6>2001
<7>The sensitivity of E. coli and C. freundii strains to 5-azacytidine and restrictase activity
of partially purified cell-free extracts were investigated.  Restrictase activity was found
only in 5-azacytidine-sensitive strains.  In the 5-azacytidine-resistant strains restrictase
activity was not detected.

<>

<1>Lebenka, A.Y.
<2>Method of producing the restriction endonuclease CfuI.
<3>Soviet Patent Office
<4>SU 1546485
<5>
<6>1990
<7>The present invention is directed to a method for producing the restriction endonuclease CfuI
by 1) fermenting Caulobacter fusiformis in media containing peptone, yeast extract and MgSO4;
2) disruption of cells by sonication; 3) chromatography on phophocellulose with a salt
gradient of 0.1-1.0M NaCl; 4) chromatography on DEAE-Sephacell and heparin-sepharose.

<>

<1>Lebenka, A.Y., Chitavichyus, D.B.
<2>Enzymes of DNA restriction-modification from Caulobacter fusiformis BC-25.
<3>Biokhimiia
<4>53
<5>1895-1899
<6>1988
<7>The CfuII system of restriction-modification enzymes, as well as the methylase
CfuIII, was detected in cells of Caulobacter fusiformis BC-25.  R.CfuII
recognizes and cleaves the sequence 5'-CTGCA^G-3' (PstI).  This system has the
corresponding methylase CfuII, which protects the host DNA.  The manifestation
of R.CfuII activity requires Mg2+ or Mn2+.  M.CfuIII protects DNA from cleavage
by the restriction endonuclease EcoRI.  Modification of DNA by a Cfu of type
III is a generic characteristic of Caulobacter.

<>

<1>Lebenka, A.Y., Kanopkaite, S.I., Buryanov, Y.I.
<2>DNA methylation in the cells of Escherichia coli MRE 600 in the presence of S-methylmethionine.
<3>Biokhimiia
<4>46
<5>2160-2163
<6>1981
<7>The effect of S-methylmethionine, a methyl group donor, on enzymatic methylation of DNA in E.
coli MRE 600 cells was studied.  It was found that SMM can be used as a donor of methyl groups
during bacterial DNA methylation in vivo without changing the specificity of DNA methylation.

<>

<1>Lebenka, A.Y., Rackus, Y.A.
<2>DNA methylase Sau3AI:  isolation and properties.
<3>Biokhimiia
<4>54
<5>1009-1014
<6>1989
<7>DNA-methylase Sau3AI has been isolated for the first time from Staphylococcus aureus 3A cells
and purified by column chromatography on phosphocellulose PII, heparin-Sepharose and blue
Sepharose. The purified enzyme methylates the GATC sequence with the formation of GATm5C as
can be evidenced from the protection of DNA from digestion with restrictases Sau3AI and BamHI,
the lack of the C3H3-group incorporation into Sau3AI DNA-restricts and the formation of a
single methylated base m5C. Sau3AI methylase modifies only double-stranded (but not
single-stranded) DNA. Thus, methylase Sau3AI modifies both DNA chains in the recognition site
during a single binding act. The 5-azacytidine-containing DNA inhibits by 95% the activity of
methylase Sau3AI. Ado-met is the single methyl group donor for methylase Sau3AI. The presence
of m6A in the recognition site does not affect the activity of methylase Sau3AI. The practical
recommendations for the use of M.Sau3AI, alongside with M.Eco dam, for the study of dam
methylation by additional methylation of the DNA in vitro in the presence of
[methyl-3H]-S-adenosyl-methionine are given.

<>

<1>Lebionka, A.Y., Melvidas, V.I.
<2>The ability of bacterial DNA methyltransferases to use methylcobalamine as a cofactor in DNA methylation reactions.
<3>Bioorg. Khim.
<4>53
<5>159-163
<6>2007
<7>The ability of bacterial DNA methyltransferases Alu I, Cfr I, Cfr 6, Cfr 10, Eco RI, Eco RII,
Msp I, Mva I, Pvu I, Pvu II, and Sau 3A to use
methyl-cobalamine and methyl-methionine as cofactors of DNA methylation
in vitro. These bacterial DNA methyl transferase used
methyl-cobalamine, but not methylmethionine for DNA methylation.

<>

<1>LeBon, J.M., Kado, C.I., Rosenthal, L.J., Chirikjian, J.G.
<2>DNA modifying enzymes of Agrobacterium tumefaciens: Effect of DNA topoisomerase, restriction endonuclease, and unique DNA endonuclease on plasmid and plant DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>75
<5>4097-4101
<6>1978
<7>Extracts from Agrobacterium tumefaciens strain 1D135 contain three enzymes that
have been characterized and partially purified.  The first enzyme, a DNA
topoisomerase, appeared to relax only negatively twisted DNA.  The second
enzyme, AtuI, a type II restriction endonuclease, generated the identical DNA
digestion pattern as EcoRII when several DNAs were used.  The third enzyme,
endonuclease A, showed a preference for superhelical DNAs as substrates.  When
plasmid pCK135DNA, obtained from the virulent strain 1D135 of A. tumefaciens,
or plant DNA was exposed to the three enzymes, changes in DNA patterns were
observed due to either conformational changes or digestion of the DNAs.  These
enzymes may function in vivo in the processing and incorporation of bacterial
DNA in plant cells.

<>

<1>Lebreton, F., Valentino, M.D., Duncan, L.B., Zeng, Q., Manson, Mc.G.A., Earl, A.M., Gilmore, M.S.
<2>High-Quality Draft Genome Sequence of Vagococcus lutrae Strain LBD1, Isolated from the Largemouth Bass Micropterus salmoides.
<3>Genome Announcements
<4>1
<5>e01087-13
<6>2013
<7>Vagococci are usually isolated from marine hosts and occasionally from endodontic infections.
Using 16S rRNA gene comparison, the closest relatives are members of
the genera Enterococcus and Carnobacterium. A draft sequence of Vagococcus lutrae
was generated to clarify the relationship of Vagococcus to these and other
related low-G+C Gram-positive bacteria.

<>

<1>Lechner, M., Frey, B., Laue, F., Ankenbauer, W., Schmitz, G.
<2>SwaI, a unique restriction endonuclease from Staphylococcus warneri, which recognizes 5'-ATTTAAAT-3'.
<3>Fresenius Z. Anal. Chem.
<4>343
<5>121-122
<6>1992
<7>Restriction endonucleases with low cutting frequencies are important tools in molecular
genetics. Analysis of complex DNA molecules e.g. chromosomes requires cutting of DNA into
fragments of megabase size range. Nucleases with low cutting frequencies either recognize
sequences which are under represented in chromosomal DNA or are characterized by recognition
sequences of more than six base pairs. We discovered and isolated SwaI a novel class-II
restriction endonuclease from Staphylococcus warneri whose recognition specificity requires
eight nucleotides.

<>

<1>Lechner, M., Frey, B., Laue, F., Anton-Botella, J., Smith, C.L., Ankenbauer, W., Schmitz, G.G.
<2>SwaI, a unique restriction endonuclease from Staphylococcus warneri, which recognizes 5'-ATTTAAAT-3'.
<3>Nucleic Acids Res.
<4>20
<5>2293-2296
<6>1992
<7>A novel class-II restriction endonuclease designated SwaI was purified from Staphyloccus
warneri. This enzyme cleaves adenovirus 2 DNA, SV40 DNA and M13mp7 at one site each, but does
not cleave lambda, PhiX174, pBR322 or pBR328 DNA. SwaI recognizes the octanucleotide sequence
5'-ATTTAAAT-3', cleaving in the center of the recognition sequence creating blunt ended DNA
fragments. SwaI was used to digest chromosomal DNA from various microorganisms and human
cells.

<>

<1>Leclercq, S., Dittmer, J., Bouchon, D., Cordaux, R.
<2>Phylogenomics of 'Candidatus Hepatoplasma crinochetorum,' a Lineage of Mollicutes Associated with Noninsect Arthropods.
<3>Genome Biol. Evol.
<4>6
<5>407-415
<6>2014
<7>Bacterial gut communities of arthropods are highly diverse and tightly related to
host feeding habits. However, our understanding of the origin and role of the
symbionts is often hindered by the lack of genetic information. "Candidatus
Hepatoplasma crinochetorum" is a Mollicutes symbiont found in the midgut glands
of terrestrial isopods. The only available nucleotide sequence for this symbiont
is a partial 16S rRNA gene sequence. Here, we present the 657,101 bp assembled
genome of Candidatus Hepatoplasma crinochetorum isolated from the terrestrial
isopod Armadillidium vulgare. While previous 16S rRNA gene-based analyses have
provided inconclusive results regarding the phylogenetic position of Candidatus
Hepatoplasma crinochetorum within Mollicutes, we performed a phylogenomic
analysis of 127 Mollicutes orthologous genes which confidently branches the
species as a sister group to the Hominis group of Mycoplasma. Several genome
properties of Candidatus Hepatoplasma crinochetorum are also highlighted compared
with other Mollicutes genomes, including adjacent tryptophan tRNA genes, which
further our understanding of the evolutionary dynamics of these genes in
Mollicutes, and the presence of a probably inactivated CRISPR/Cas system, which
constitutes a testimony of past interactions between Candidatus Hepatoplasma
crinochetorum and mobile genetic elements, despite their current lack in this
streamlined genome. Overall, the availability of the complete genome sequence of
Candidatus Hepatoplasma crinochetorum paves the way for further investigation of
its ecology and evolution.

<>

<1>Lederberg, S.
<2>Genetics of host-controlled restriction anad modification of deoxyribonucleic acid in Escherichia coli.
<3>J. Bacteriol.
<4>91
<5>1029-1036
<6>1966
<7>The locus for the host specific restriction and modification of
deoxyribonucleic acid in Escherichia coli has been mapped by matings between
mutants for these characters in strains K-12, C600, and B. Linkage analysis and
kinetics of marker transfer indicate that a single or closely linked multiple
chromosomal site located about 4 min counterclockwise to leucine is responsible
for these activities.  Secondary factors which affect the quantitative level of
restriction also were detected.  Wild-type recombinants were isolated in
crosses between rm- (restriction or modification, or both) mutants.  The
expression in zygotes of the restrictionless character of a rm- donor is masked
by a separate, physiological impairment of restriction, which results from
mating and is independent of the modification state of the donor.  The
relevance of the restriction character to mating incompatibilities in these and
other bacterial strains is considered.

<>

<1>Lederberg, S.
<2>Host-controlled restriction and modification of deoxyribonucleic acid in Eschericia coli.
<3>Virology
<4>27
<5>378-387
<6>1965
<7>In Escherichia coli strain C600 (which has the host specificity of K12),
restrictions against infection by phage lambda of host specificities B and C
have the same sensitivity to heat inactivation.  Likewise, in strain B,
restrictions against phage lambda of host specificities C600 and C have an
identical heat sensitivity.  Strain C600(P1) has a heat sensitivity for loss of
its P1-directed restriction different from that of its C600-controlled
restriction.  Mutants of K12-type strains and of B which are impaired in their
restriction and modification activities have been isolated.  In K12 strains,
restrictions toward phage lambda of host specificities C and B are impaired by
the same mutation.  In B, restrictions toward phage lambda of host
specificities C and C600 also are lost simultaneously by mutation.  The
coordinate changes in restriction by heat treatment and by mutation indicate
that a common mechanism for restriction of lambda of host specificities C and B
operates in K12 strains, and that a single mechanism for restriction of phage
lambda of host specificities C and K12 occurs in strain B.  Restrictionless
mutants of B, unlike their parent, act as fertile F- in crosses with K12 Hfr
strains.  Modificationless mutants of K12 Hfr, unlike their parent, are no
longer fertile with restricting C600 strains.  Thus, the same mutations affect
the modification and/or restriction of the DNA of bacteria as well as phage.
Models are proposed for host restriction and modfication.  The models visualize
restriction as either a screening of new DNA by a DNA-site specific degrading
activity-successful passage permitting the DNA to operate in that cell-or a
seavenging of new DNA by a nonspecific degrading activity when such DNA fails
to be complexed with a site-specific protecting agent.

<>

<1>Ledwaba, B., Mafofo, J., van Heerden, H.
<2>Genome Sequences of Brucella abortus and Brucella suis Strains Isolated from Bovine in Zimbabwe.
<3>Genome Announcements
<4>2
<5>e01063-14
<6>2014
<7>This is a report of whole-genome sequences of a Brucella abortus strain and two Brucella suis
strains isolated from bovine in Zimbabwe. These strains were
selected based on their origin and data obtained when using multiplex PCR assays,
then sequenced using next-generation sequencing technologies.

<>

<1>Lee, A.S., Sinsheimer, R.L.
<2>A cleavage map of bacteriophage PhiX174 genome.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>71
<5>2882-2886
<6>1974
<7>Restriction endonucleases isolated from Hemophilus influenzae, Hemophilus
parainfluenzae, and Hemophilus aegyptius were used to cleave PhiX174
replicative form DNA into three sets of specific DNA fragments.  The order of
these fragments in the PhiX replicative form molecule was determined by (1)
analysis of partial digest products, (2) analysis of overlapping sets of
fragments produced by two different restrictive enzymes.  On the basis of these
results, a detailed physical map of the PhiX174 genome has been constructed
with respect to the cleavage sites of all three enzymes.

<>

<1>Lee, B., Muller, M.T.
<2>SUMOylation enhances DNA methyltransferase 1 activity.
<3>Biochem. J.
<4>421
<5>449-461
<6>2009
<7>DNA methylation regulates gene expression through Complex network of protein-protein and
protein-DNA interactions in chromatin. The
maintenance methylase, DNMT1 (DNA methyltransferase 1), is a prominent
enzyme in the process that is linked to DNA replication and drives the
heritable nature of epigenetic modifications. The mechanistic details
that explain how DNMT1 catalytic action is directed and regulated in
chromatin are important ill our overall understanding of gene control.
In this work, we show that DNMT1 is modified by SUMOylation and we have
mapped these SUMOylation sites by defined mutations. SUMOylated DNMT1
is catalytically active on genomic DNA in vivo and we find that
SUMOylation significantly enhances the methylase activity of DNMT1 both
in vitro and in chromatin. These data suggest that SUMOylation
modulates the endogenous activity of a prominent epigenetic maintenance
pathway in somatic cells.

<>

<1>Lee, B.H., Yegnasubramanian, S., Lin, X.H., Nelson, W.G.
<2>Procainamide is a specific inhibitor of DNA methyltransferase 1.
<3>J. Biol. Chem.
<4>280
<5>40749-40756
<6>2005
<7>CpG island hypermethylation occurs in most cases of cancer, typically resulting in the
transcriptional silencing of critical cancer genes.
Procainamide has been shown to inhibit DNA methyltransferase activity
and reactivate silenced gene expression in cancer cells by reversing
CpG island hypermethylation. We report here that procainamide
specifically inhibits the hemimethylase activity of DNA
methyltransferase 1 (DNMT1), the mammalian enzyme thought to be
responsible for maintaining DNA methylation patterns during
replication. At micromolar concentrations, procainamide was found to be
a partial competitive inhibitor of DNMT1, reducing the affinity of the
enzyme for its two substrates, hemimethylated DNA and
S-adenosyl-L-methionine. By doing so, procainamide significantly
decreased the processivity of DNMT1 on hemimethylated DNA. Procainamide
was not a potent inhibitor of the de novo methyltransferases DNMT3a and
DNMT3b2. As further evidence of the specificity of procainamide for
DNMT1, procainamide failed to lower genomic 5-methyl-2'-deoxycytidine
levels in HCT116 colorectal cancer cells when DNMT1 was genetically
deleted but significantly reduced genomic 5-methyl-2'-deoxycytidine
content in parental HCT116 cells and in HCT116 cells where DNMT3b was
genetically deleted. Because many reports have strongly linked DNMT1
with epigenetic alterations in carcinogenesis, procainamide may be a
useful drug in the prevention of cancer.

<>

<1>Lee, B.M., Park, Y.J., Park, D.S., Kang, H.W., Kim, J.G., Song, E.S., Park, I.C., Yoon, U.H., Hahn, J.H., Koo, B.S., Lee, G.B., Kim, H., Park, H.S., Yoon, K.O., Kim, J.H., Jung, C.H., Koh, N.H., Seo, J.S., Go, S.J.
<2>The genome sequence of Xanthomonas oryzae pathovar oryzae KACC10331, the bacterial blight pathogen of rice.
<3>Nucleic Acids Res.
<4>33
<5>577-586
<6>2005
<7>The nucleotide sequence was determined for the genome of Xanthomonas oryzae pathovar oryzae
(Xoo) KACC10331, a bacterium that causes bacterial
blight in rice (Oryza sativa L.). The genome is comprised of a single, 4
941 439 bp, circular chromosome that is G + C rich (63.7%). The genome
includes 4637 open reading frames (ORFs) of which 3340 (72.0%) could be
assigned putative function. Orthologs for 80% of the predicted Xoo genes
were found in the previously reported X.axonopodis pv. citri (Xac) and
X.campestris pv. campestris (Xcc) genomes, but 245 genes apparently
specific to Xoo were identified. Xoo genes likely to be associated with
pathogenesis include eight with similarity to Xanthomonas avirulence (avr)
genes, a set of hypersensitive reaction and pathogenicity (hrp) genes,
genes for exopolysaccharide production, and genes encoding extracellular
plant cell wall-degrading enzymes. The presence of these genes provides
insights into the interactions of this pathogen with its gramineous host.

<>

<1>Lee, C., Peters, V., Melefors, O., Romling, U.
<2>Draft Genome Sequence of Pseudomonas aeruginosa SG17M, an Environmental Isolate Belonging to Clone C, Prevalent in Patients and Aquatic Habitats.
<3>Genome Announcements
<4>2
<5>e00186-14
<6>2014
<7>Pseudomonas aeruginosa SG17M is an environmental isolate recovered from river water in the
city of Mulheim, Germany. SG17M belongs to clone C, which is
distributed worldwide. This is the first clone C strain whose genome sequence has
been determined.

<>

<1>Lee, D., Seo, H., Park, C., Park, K.
<2>WeGAS: a web-based microbial genome annotation system.
<3>Biosci. Biotechnol. Biochem.
<4>73
<5>213-216
<6>2009
<7>We have developed WeGAS, a Web based microbial Genome
Annotation System, which provides features that include gene prediction,
homology search, promoter/motif analysis, genome browsing, gene ontology
analysis based on the COGs and GO, and metabolic pathway analysis with
web-based interfaces. Most raw data and intermediate data from genome
projects can be managed with the WeGAS database system, and analysis
results, including information on each gene and final genome maps, are
provided by its visualization modules. Especially, a pie-view browser
displaying circular maps of contigs and a COG-GO combination browser are
very helpful for an overview of projects. Major public microbial genome
databases can be imported, searched, and browsed through the WeGAS
modules. WeGAS is freely accessible via web site
http://ns.smallsoft.co.kr:8051.

<>

<1>Lee, D.G., Urbach, J.M., Wu, G., Liberati, N.T., Feinbaum, R.L., Miyata, S., Diggins, L.T., He, J., Saucier, M., Deziel, E., Friedman, L., Li, L., Grills, G., Montgomery, K., Kucherlapati, R., Rahme, L.G., Ausubel, F.M.
<2>Genomic analysis reveals that Pseudomonas aeruginosa virulence is combinatorial.
<3>Genome Biol.
<4>7
<5>R90
<6>2006
<7>ABSTRACT: BACKGROUND: Pseudomonas aeruginosa is a ubiquitous environmental bacterium and an
important opportunistic human pathogen. Generally, the
acquisition of genes in the form of pathogenicity islands distinguishes
pathogenic isolates from non-pathogens. We therefore sequenced a highly
virulent strain of P. aeruginosa, PA14, and compared it to a previously
sequenced (and less pathogenic) strain, PAO1, to identify novel virulence
genes. RESULTS: The PA14 and PAO1 genomes are remarkably similar, although
PA14 has a slightly larger genome (6.5 MB) than PAO1 (6.3 MB). We
identified 58 PA14 gene clusters that are absent in PAO1 to determine
which of these genes, if any, contribute to its enhanced virulence in a
Caenorhabditis elegans pathogenicity model. First, we tested 18 additional
diverse strains in the C. elegans model and observed a wide range of
pathogenic potential; however, genotyping these strains using a custom
microarray showed that the presence of PA14 genes absent in PAO1 did not
correlate with the virulence of these strains. Second we utilized a
full-genome non-redundant mutant library of PA14 to identify five genes
(absent in PAO1) required for C. elegans killing. Surprisingly, although
these five genes are present in many other P. aeruginosa strains, they do
not correlate with virulence in C. elegans. CONCLUSIONS: Genes required
for pathogenicity in one strain of P. aeruginosa are neither required for
nor predictive of virulence in other strains. We therefore propose that
virulence in this organism is multifactorial and combinatorial, the result
of a pool of pathogenicity-related genes that interact in various
combinations in different genetic backgrounds.

<>

<1>Lee, D.H., Jin, S.G., Cai, S., Chen, Y., Pfeifer, G.P., O'Connor, T.R.
<2>Repair of methylation damage in DNA and RNA by mammalian AlkB homologues.
<3>J. Biol. Chem.
<4>280
<5>39448-39459
<6>2005
<7>Human and Escherichia coli derivatives of AlkB enzymes remove methyl groups from
1-methyladenine and 3-methylcytosine in nucleic acids via an
oxidative mechanism that releases the methyl group as formaldehyde. In
this report, we demonstrate that the mouse homologues of the
alpha-ketoglutarate Fe(II) oxygen-dependent enzymes mAbh2 and Abh3 have
activities comparable to those of their human counterparts. The mAbh2 and
mAbh3 release modified bases from both DNA and RNA. Comparison of the
activities of the homogenous ABH2 and ABH3 enzymes demonstrate that these
activities are shared by both sets of enzymes. An assay for the detection
of alpha-ketoglutarate Fe(II) dioxygenase activity using an
oligodeoxyribonucleotide with a unique modification shows activity for all
four enzymes studied and a loss of activity for eight mutant proteins.
Steady-state kinetics for removal of methyl groups from DNA substrates
indicates that the reactions of the proteins are close to the diffusion
limit. Moreover, mAbh2 or mAbh3 activity increases survival in a strain
defective in alkB. The mRNAs of AHB2 and ABH3 are expressed most in testis
for ABH2 and ABH3, whereas expression of the homologous mouse genes is
different. The mAbh3 is strongly expressed in testis, whereas highest
expression of mAbh2 is in heart. Other purified human AlkB homologue
proteins ABH4, ABH6, and ABH7 do not manifest activity. The demonstration
of mAbh2 and mAbh3 activities and their distributions provide data on
these mammalian homologues of AlkB that can be used in animal studies.

<>

<1>Lee, G.H., Kumar, S., Lee, J.H., Chang, D.H., Kim, D.S., Choi, S.H., Rhee, M.S., Lee, D.W., Yoon, M.H., Kim, B.C.
<2>Genome Sequence of Oscillibacter ruminantium Strain GH1, Isolated from Rumen of Korean Native Cattle.
<3>J. Bacteriol.
<4>194
<5>6362
<6>2012
<7>Oscillibacter ruminantium strain GH1 was isolated from the rumen of Korean native cattle
(HanWoo; Bos taurus coreanae). Here, we present the 3.07-Mb draft genome
of this strain, which could reveal the presence of certain fiber-specific
glycoside hydrolases and butyric acid-producing genes.

<>

<1>Lee, G.W., Lee, K.J., Chae, J.C.
<2>Genome Sequence of Herbaspirillum sp. Strain GW103, a Plant Growth-Promoting Bacterium.
<3>J. Bacteriol.
<4>194
<5>4150
<6>2012
<7>Herbaspirillum sp. strain GW103 was isolated from rhizosphere soil of the reed Phragmites
australis on reclaimed land. Here we report the 5.05-Mb draft genome
sequence of the strain, providing bioinformation about the agronomic benefits of
this strain, such as multiple traits relevant to plant root colonization and
plant growth promotion.

<>

<1>Lee, H., Kang, S., Yim, J.-B., Hwang, D.S.
<2>Identification of hemimethylated DNA binding activity in the seqA mutant.
<3>Korean J. Biol. Sci.
<4>2
<5>351-353
<6>1998
<7>A 245 bp segment of E. coli chromosomal replication origin, oriC, contains 11 repeats of the
GATC sequence in which adenine is methylated by Dam methylase.  Newly replicated oriC is
hemimethylated.  The parental strand of the newly replicated oriC is methylated, but the
nascent strand is not yet methylated until methylated by Dam methylase.  The hemimethylated
oriC plays an important role in the regulation of chromosomal replication.  Activity in the
seqA mutant was identified to bind preferentially to hemimethylated DNA, but not to
fully-methylated DNA.  This activity may participate in the sequestration of initiation of
chromosomal replication.

<>

<1>Lee, H., Kim, B.J., Kim, K., Hong, S.H., Kook, Y.H., Kim, B.J.
<2>Whole-Genome Sequence of Mycobacterium intracellulare Clinical Strain MOTT-H4Y, Belonging to INT5 Genotype.
<3>Genome Announcements
<4>1
<5>e00006-13
<6>2013
<7>Here, we report the draft genome sequence of the clinical strain MOTT-H4Y, grouped previously
into the INT5 genotype of the 5 genotypes of .

<>

<1>Lee, H., Shin, S.C., Lee, J., Kim, S.J., Kim, B.K., Hong, S.G., Kim, E.H., Park, H.
<2>Genome Sequence of Sphingomonas sp. Strain PAMC 26621, an Arctic-Lichen-Associated Bacterium Isolated from a Cetraria sp.
<3>J. Bacteriol.
<4>194
<5>3030
<6>2012
<7>The lichen-associated bacterial strain Sphingomonas sp. PAMC 26621 was isolated from an Arctic
lichen Cetraria sp. on Svalbard Islands. Here we report the draft genome sequence of this
strain, which could provide novel insights into the molecular principles of lichen-microbe
interactions.

<>

<1>Lee, H.J., Nishi, K., Song, J.M., Kim, J.S.
<2>Expression, crystallization and preliminary X-ray diffraction analysis of a modification subunit of a putative type I restriction enzyme from Vibrio vulnificus YJ016.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>65
<5>1271-1273
<6>2009
<7>Modification (HsdM) and specificity (HsdS) subunits are constituents of an active
methyltransferase (MTase) of multifunctional type I
restriction enzymes. To provide a molecular background on HsdM, a
putative hsdM gene from Vibrio vulnificus YJ016 (HsdM Vv) was cloned
and the expressed protein was purified and crystallized from 22%(w/v)
polyethylene glycol 8000, 0.02 M imidazole pH 7.5 and 5 mM
beta-mercaptoethanol. Diffraction data were collected to 1.86 angstrom
resolution using synchrotron radiation. The crystal belonged to the
tetragonal space group P4(1)2(1)2 or P4(3)2(1)2, with unit-cell
parameters a = b = 78.9, c = 165.8 angstrom. With one molecule in the
asymmetric unit, the crystal volume per unit protein weight was 2.12
angstrom(3) Da(-1), with a solvent content of 42%.

<>

<1>Lee, H.S., Bae, S.S., Kim, M.S., Kwon, K.K., Kang, S.G., Lee, J.H.
<2>Complete Genome Sequence of Hyperthermophilic Pyrococcus sp. Strain NA2, Isolated from Deep-Sea Hydrothermal Vent Area.
<3>J. Bacteriol.
<4>193
<5>3666-3667
<6>2011
<7>Pyrococcus sp. NA2, isolated from a deep-sea hydrothermal vent sample, is a novel marine
hyperthermophilic archaeon to grow optimally at 93 degrees C. The complete genome sequence of
the strain contains all genes for
tricarboxylic acid cycle except for succinate dehydrogease/fumarate reductase, but it doesn't
encode proteins involved in polysaccharide utilization.

<>

<1>Lee, H.S., Kang, S.G., Bae, S.S., Lim, J.K., Cho, Y., Kim, Y.J., Jeon, J.H., Cha, S.S., Kwon, K.K., Kim, H.T., Park, C.J., Lee, H.W., Kim, S.I., Chun, J., Colwell, R.R., Kim, S.J., Lee, J.H.
<2>The complete genome sequence of Thermococcus onnurineus NA1 reveals a mixed heterotrophic and carboxydotrophic metabolism.
<3>J. Bacteriol.
<4>190
<5>7491-7499
<6>2008
<7>Members of the genus Thermococcus, sulfur-reducing
hyperthermophilic archaea, are ubiquitously present in various deep-sea hydrothermal vent
systems and are considered to play a significant role in the microbial consortia. We present
the complete genome sequence and feature analysis of Thermococcus onnurineus NA1 isolated from
a deep-sea hydrothermal vent area, which reveal clues to its physiology. Based on results of
genomic analysis, T. onnurineus NA1 possesses the metabolic pathways for organotrophic growth
on peptides, amino acids, or sugars. More interesting was the discovery that the genome
encoded unique proteins that are involved in carboxydotrophy to generate energy by oxidation
of CO to CO(2), thereby providing a mechanistic basis for growth with CO as a substrate. This
lithotrophic feature in combination with carbon fixation via RuBisCO (ribulose
1,5-bisphosphate carboxylase/oxygenase) introduces a new strategy with a complementing energy
supply for T. onnurineus NA1 potentially allowing it to cope with nutrient stress in the
surrounding of hydrothermal vents, providing the first genomic evidence for the
carboxydotrophy in Thermococcus.

<>

<1>Lee, H.S., Kang, S.G., Kwon, K.K., Lee, J.H., Kim, S.J.
<2>Genome Sequence of the Algicidal Bacterium Kordia algicida OT-1.
<3>J. Bacteriol.
<4>193
<5>4031-4032
<6>2011
<7>Kordia algicida OT-1 is an algicidal bacterium against the bloom-forming microalgae. The
genome sequence of K. algicida revealed a number of
interesting features, including degradation of macromolecules, the
biosynthesis of carotenoid pigment and secondary metabolites, and the
capacity for gliding motility, which might facilitate the understanding of
algicidal mechanisms.

<>

<1>Lee, J., Roh, S.W., Whon, T.W., Shin, N.R., Kim, Y.O., Bae, J.W.
<2>Genome Sequence of Strain TW15, a Novel Member of the Genus Ruegeria Belonging to the Marine Roseobacter Clade.
<3>J. Bacteriol.
<4>193
<5>3401-3402
<6>2011
<7>Ruegeria sp. TW15, which belongs to the family Rhodobacteraceae, was isolated from an ark clam
in the South Sea of Korea. Here is presented the
draft genome sequence of Ruegeria sp. TW15 (4,490,771 bp, with a G+C
content of 55.7%), a member of the marine Roseobacter clade, which
comprises up to 20% of bacterioplankton in the coastal and oceanic mixed
layer.

<>

<1>Lee, J., Shin, S.C., Kim, S.J., Kim, B.K., Hong, S.G., Kim, E.H., Park, H., Lee, H.
<2>Draft Genome Sequence of a Sphingomonas sp., an Endosymbiotic Bacterium Isolated  from an Arctic Lichen Umbilicaria sp.
<3>J. Bacteriol.
<4>194
<5>3010-3011
<6>2012
<7>Sphingomonas sp. strain PAMC 26617 has been isolated from an Arctic lichen Umbilicaria sp. on
the Svalbard Islands. Here we present the draft genome
sequence of this strain, which represents a valuable resource for understanding
the symbiotic mechanisms between endosymbiotic bacteria and lichens surviving in
extreme environments.

<>

<1>Lee, J.-H., Shin, H., Kim, H., Ryu, S.
<2>Complete Genome Sequence of Salmonella Bacteriophage SPN3US.
<3>J. Virol.
<4>85
<5>13470-13471
<6>2011
<7>Salmonella bacteriophage SPN3US was isolated from a chicken fecal sample. It is a virulent
phage belonging
to the Myoviridae family and showing effective inhibition of Salmonella enterica and a few
Escherichia coli
O157:H7 strains. Here we announce the completely sequenced first genome of a Salmonella phage
using flagella
as receptors. It is the largest genome among Salmonella phages sequenced to date, and major
findings from its
annotation are described.

<>

<1>Lee, J.-I., Yang, C.-H.
<2>The purification and characterization of PvuII, restriction endonuclease.
<3>Korean Biochem. J.
<4>17
<5>357-364
<6>1984
<7>Restriction endonuclease PvuII has been purified from Proteus vulgaris, aerobic
bacterium, by streptomycin sulfate fractionation, (NH4)SO4 precipitation, gel
filtration on Sephadex G-150, Phosphocellulose (P11) chromatography, and
DEAE-cellulose chromatography.

<>

<1>Lee, J.-T., Cho, T.-J., Lim, J.-Y.
<2>Characterization of a restriction endonuclease AspJI from Alcaligenes sp. J-482.
<3>Korean J. Microbiol.
<4>32
<5>285-290
<6>1994
<7>About 500 bacterial and fungal strains from a wide variety of natural habitats were screened
for a new type II restriction endonuclease.  Among the 500 species, we selected one species
that produced a new restriction endonuclease.  This strain has an optimum temperature of 30oC
for growth.  Morphological, cultural, and physiological characteristics were examined for
identification of the isolated strain J-482.  This strain was found to belong to the genus
Alcaligenes.  The restriction endonuclease was named as AspJI and partially purified from
Alcaligenes sp. J-482 by DEAE-Sephadex A-50 column chromatography and gel filtration.  Most of
other nucleases were removed by the purification steps.  The AspJI has a substrate
specificity to lambda DNA, pBR322 and Adenovirus-2 DNA.  For its maximal activity, the
isolated enzyme requires MgCl2, which should be at least 12.5  mM and it does not need any
other cofactors.  It is maximally active in the absence of NaCl and is completely inactivated
at 100 mM NaCl.  The pH and temperature optima for activity were pH 7.5 and 37oC,
respectively.  The DNA fragments generated by digesting lambda DNA, pBR322, and Adenovirus-2
DNA with AspJI were the same as that produced by AatII.  This suggests that AspJI is an
isoschizomer of AatII.

<>

<1>Lee, J.C., Kim, D.S., Moon, D.C., Lee, J.H., Kim, M.J., Lee, S.M., Lee, Y.S., Kang, S.W., Lee, E.J., Kang, S.S., Lee, E., Hyun, S.H.
<2>Prediction of bacterial proteins carrying a nuclear localization signal and nuclear targeting of HsdM from Klebsiella pneumoniae.
<3>J. Microbiol.
<4>47
<5>641-645
<6>2009
<7>Nuclear targeting of bacterial proteins is an emerging pathogenic mechanism whereby bacterial
proteins can interact with nuclear
molecules and alter the physiology of host cells. The fully sequenced
bacterial genome can predict proteins that target the nuclei of host
cells based on the presence of nuclear localization signal (NLS). In
the present study, we predicted bacterial proteins with the NLS
sequences from Klebsiella pneumoniae by bioinformatic analysis, and 13
proteins were identified as carrying putative NLS sequences. Among
them, HsdM, a subunit of KpnAl that is a type I
restriction-modification system found in K. pneumoniae, was selected
for the experimental proof of nuclear targeting in host cells. HsdM
carried the NLS sequences, (7)KKAKAKK(13), in the N-terminus. A
transient expression of HsdM-EGFP in COS-1 cells exhibited exclusively
a nuclear localization of the fusion proteins, whereas the fusion
proteins of HsdM with substitutions in residues lysine to alanine in
the NLS sequences, (7)AAAKAAA(13), were localized in the cytoplasm.
HsdM was co-localized with importin o in the nuclei of host cells.
Recombinant HsdM alone methylated the eukaryotic DNA in vitro assay.
Although HsdM tested in this study has not been considered to be a
virulence factor, the prediction of NLS motifs from the full sequenced
genome of bacteria extends our knowledge of functional genomics to
understand subcellular targeting of bacterial proteins.

<>

<1>Lee, J.H., Bae, J.W., Chun, J.
<2>Draft Genome Sequence of Weissella koreensis KCTC 3621T.
<3>J. Bacteriol.
<4>194
<5>5711-5712
<6>2012
<7>Weissella koreensis is a Gram-positive, rod-shaped, nonmotile, and facultative anaerobic
species belonging to the lactic acid bacteria (LAB). The members of
this species have been repeatedly isolated from kimchi (a traditional Korean
fermented food) and are known for their beneficial effects on human and animal
intestinal microflora through producing various clinically important amino acids
such as gamma-aminobutyric acid and ornithine. Here we report the genome sequence
of the type strain of W. koreensis (KCTC 3621(T)) to provide taxonomic and
functional insights into the species.

<>

<1>Lee, J.H., Chae, J.P., Lee, J.Y., Lim, J.S., Kim, G.B., Ham, J.S., Chun, J., Kang, D.K.
<2>Genome Sequence of Lactobacillus johnsonii PF01, Isolated from Piglet Feces.
<3>J. Bacteriol.
<4>193
<5>5030-5031
<6>2011
<7>Lactobacillus johnsonii PF01, an autochthonous bacterium of the gastrointestinal tract, was
isolated from a fecal sample of piglet. The
strain adhered specifically to the duodenal and jejunal epithelial cells
of the piglet and had high bile resistance activity. Here, we report a
genomic sequence of L. johnsonii PF01.

<>

<1>Lee, J.H., Cheon, I.S., Shim, B.S., Kim, D.W., Kim, S.W., Chun, J., Song, M.
<2>Draft Genome Sequence of Klebsiella pneumoniae subsp. pneumoniae DSM 30104T.
<3>J. Bacteriol.
<4>194
<5>5722-5723
<6>2012
<7>Klebsiella pneumoniae is a Gram-negative, rod-shaped, nonmotile, and opportunistic pathogenic
species with clinical importance. It is a part of
natural flora of humans and animals. Here we report the draft genome sequence of
the type strain of Klebsiella pneumoniae subsp. pneumoniae (DSM 30104(T)) to
provide taxonomic and functional insights into the species.

<>

<1>Lee, J.H., Choi, Y., Shin, H., Lee, J., Ryu, S.
<2>Complete Genome Sequence of Cronobacter sakazakii Temperate Bacteriophage phiES15.
<3>J. Virol.
<4>86
<5>7713-7714
<6>2012
<7>While most phage genome studies have been focused on the virulent phages, the
inducible temperate bacteriophage genome study provides more detailed information
about the interaction between the host strain and the phage. To study this
interaction in detail, UV-induced phiES15 bacteriophage was isolated from the
host strain Cronobacter sakazakii ES15 and its genome was completely sequenced.
Here we announce the genome sequence of phiES15 and report major findings from
the annotation.

<>

<1>Lee, J.H., Karamychev, V.N., Kozyavkin, S.A., Mills, D., Pavlov, A.R., Pavlova, N.V., Polouchine, N.N., Richardson, P.M., Shakhova, V.V., Slesarev, A.I., Weimer, B., O'Sullivan, D.J.
<2>Comparative genomic analysis of the gut bacterium Bifidobacterium longum reveals loci susceptible to deletion during pure culture growth.
<3>BMC Genomics
<4>9
<5>247
<6>2008
<7>BACKGROUND: Bifidobacteria are frequently proposed to be associated with good intestinal
health primarily because of their overriding dominance in
the feces of breast fed infants. However, clinical feeding studies with
exogenous bifidobacteria show they don't remain in the intestine,
suggesting they may lose competitive fitness when grown outside the gut.
RESULTS: To further the understanding of genetic attenuation that may be
occurring in bifidobacteria cultures, we obtained the complete genome
sequence of an intestinal isolate, Bifidobacterium longum DJO10A that was
minimally cultured in the laboratory, and compared it to that of a culture
collection strain, B. longum NCC2705. This comparison revealed colinear
genomes that exhibited high sequence identity, except for the presence of
17 unique DNA regions in strain DJO10A and six in strain NCC2705. While
the majority of these unique regions encoded proteins of diverse function,
eight from the DJO10A genome and one from NCC2705, encoded gene clusters
predicted to be involved in diverse traits pertinent to the human
intestinal environment, specifically oligosaccharide and polyol
utilization, arsenic resistance and lantibiotic production. Seven of these
unique regions were suggested by a base deviation index analysis to have
been precisely deleted from strain NCC2705 and this is substantiated by a
DNA remnant from within one of the regions still remaining in the genome
of NCC2705 at the same locus. This targeted loss of genomic regions was
experimentally validated when growth of the intestinal B. longum in the
laboratory for 1,000 generations resulted in two large deletions, one in a
lantibiotic encoding region, analogous to a predicted deletion event for
NCC2705. A simulated fecal growth study showed a significant reduced
competitive ability of this deletion strain against Clostridium difficile
and E. coli. The deleted region was between two IS30 elements which were
experimentally demonstrated to be hyperactive within the genome. The other
deleted region bordered a novel class of mobile elements, termed mobile
integrase cassettes (MIC) substantiating the likely role of these elements
in genome deletion events. CONCLUSION: Deletion of genomic regions, often
facilitated by mobile elements, allows bifidobacteria to adapt to
fermentation environments in a very rapid manner (2 genome deletions per
1,000 generations) and the concomitant loss of possible competitive
abilities in the gut.

<>

<1>Lee, J.H., Koh, H.Y., Lee, S.G., Doyle, S., Christner, B.C., Kim, H.J.
<2>Draft Genome Sequence of Paenisporosarcina sp. Strain TG-20, a Psychrophilic Bacterium Isolated from the Basal Ice of Taylor Glacier.
<3>J. Bacteriol.
<4>194
<5>6636
<6>2012
<7>We report the draft genome sequence of Paenisporosarcina sp. strain TG-20, which  is 4.12 Mb
in size and consists of 4,071 protein-coding genes and 76 RNA genes.
The genome sequence of Paenisporosarcina sp. TG-20 may provide useful information
about molecular adaptations that enhance survival in icy subsurface environments.

<>

<1>Lee, J.H., Rhee, M.S., Kumar, S., Lee, G.H., Chang, D.H., Kim, D.S., Choi, S.H., Lee, D.W., Yoon, M.H., Kim, B.C.
<2>Genome Sequence of Methanobrevibacter sp. Strain JH1, Isolated from Rumen of Korean Native Cattle.
<3>Genome Announcements
<4>1
<5>e00002-13
<6>2013
<7>The sp. strain JH1 was isolated from the rumen of Korean native cattle (HanWoo; ). Here, we
provide a 2.06-Mb draft genome sequence of strain JH1 that might
provide more information about the lifestyle of rumen methanogens and about the
genes and proteins that can be targeted to curb methane emissions.

<>

<1>Lee, J.H., Shin, H., Choi, Y., Ryu, S.
<2>Complete genome sequence analysis of bacterial-flagellum-targeting bacteriophage chi.
<3>Arch. Virol.
<4>158
<5>2179-2183
<6>2013
<7>Bacteriophage chi is a well-known phage that infects pathogens such as E. coli,
Salmonella, and Serratia via bacterial flagella. To further understand its
host-phage interaction and infection mechanism via host flagella, the genome was
completely sequenced and analyzed. The phage genome contains 59,407-bp-length DNA
with a GC content of 56.51 %, containing 75 open reading frames (ORFs) with no
tRNA genes. Its annotation and functional analysis revealed that chi is
evolutionarily very closely related to Enterobacter phage Enc34 and Providencia
phage Redjac. However, most of the annotated genes encode hypothetical proteins,
indicating that further genomic study of phage chi is required to elucidate the
bacterial-flagellum-targeting infection mechanism of phage chi.

<>

<1>Lee, J.H., Valeriano, V.D., Shin, Y.R., Chae, J.P., Kim, G.B., Ham, J.S., Chun, J., Kang, D.K.
<2>Genome Sequence of Lactobacillus mucosae LM1, Isolated from Piglet Feces.
<3>J. Bacteriol.
<4>194
<5>4766
<6>2012
<7>Lactobacillus mucosae LM1, isolated from stool samples of a healthy piglet, displays good in
vitro mucin adhesion and antimicrobial activity against
pathogenic bacteria. To elucidate its antimicrobial effects and to find its
epithelial cell and mucin adhesion genes, the genomic sequence of L. mucosae LM1
was investigated.

<>

<1>Lee, K., Yi, H., Cho, Y.J., Jang, J., Hur, H.G., Chun, J.
<2>Draft Genome Sequence of Escherichia coli AI27, a Porcine Isolate Belonging to Phylogenetic Group B1.
<3>J. Bacteriol.
<4>194
<5>6640-6641
<6>2012
<7>Escherichia coli AI27 is a putatively commensal strain isolated from feces of a pig. Here we
report the draft genome sequence of E. coli AI27. This is the first
porcine strain in the phylogenetic group B1 whose genome sequence has been
determined.

<>

<1>Lee, K.-F., Kam, K.-M., Shaw, P.-C.
<2>A bacterial methyltransferase M.EcoHK31I requires two proteins for in vitro methylation.
<3>Nucleic Acids Res.
<4>23
<5>103-108
<6>1995
<7>The genes encoding the EcoHK31I restriction-modification (R-M) system were isolated from a
clinically-isolated Escherichia coli strain HK31. The entire R-M system of EcoHK31I is located
in a 2.1 kb fragment. R.EcoHK31I is an isoschizomer of EaeI which recognizes and cleaves
Y/GGCCR. M.EcoHK31I consists of two polypeptides alpha and beta with sizes 309 and 176 aa,
respectively. Polypeptide beta is encoded within an alternative reading frame of polypeptide
alpha. All the conserved motifs in mC5-MTases can be found in polypeptide alpha except motif
IX which is present in polypeptide beta. Polypeptides alpha and beta were separately
synthesized in a T7 promoter controlled over-expression system and in vitro methylation
occurred only when the two extracts were mixed and thus confirms that two polypeptides are
required for methylation.

<>

<1>Lee, K.-F., Liaw, Y.-C., Shaw, P.-C.
<2>Overproduction, purification and characterization of M.EcoHK31I, a bacterial methyltransferase with two polypeptides.
<3>Biochem. J.
<4>314
<5>321-326
<6>1996
<7>The two overlapping genes coding for EcoHK31I methyltransferase have previously been cloned,
sequenced and expressed.  Here we describe protocols developed to purify polypeptides alpha
and beta together or separately, to apparent homogeneity by various chromatographic media.
M.EcoHK31I is a heterodimer with a native molecular mass of 61 kDa.  Its specific activity
towards non-methylated lambda DNA was 3.0 x 105 units per mg of protein.  The respective
denatured molecular masses of polypeptides alpha and beta were 38 and 23 kDa, and their pI
values were 8.7 and 6.8.  Initial rate kinetic parameters of the native enzyme were 2.0 nM,
0.58 uM and 3 min-1 for Km DNA, Km AdoMet and kcat, respectively, where AdoMet stands for
S-adenosyl-L-methionine.  Fully active enzyme was reconstituted by co-purifying the two
separately synthesized polypeptides, and activity assays confirmed our previous finding that
two polypeptides were needed to methylate substrate DNA.

<>

<1>Lee, K.-F., Shaw, P.-C., Picone, S.J., Wilson, G.G., Lunnen, K.D.
<2>Sequence comparison of the EcoHK31I and EaeI restriction-modification systems suggests an intergenic transfer of genetic material.
<3>Biol. Chem.
<4>379
<5>437-441
<6>1998
<7>The genes coding for the EcoHK31I and EaeI restriction-modification systems from Escherichia
coli strain HK31 and Enterobacter aerogenes, respectively, have been cloned and sequenced.
Both ENases recognize and cleave Y/GGCCR leaving 4 nucleotide 5'-protruding ends, while the
MTases modify the internal cytosine.  The systems were isolated on a 2.3 kb AseI fragment for
EcoHK31I, and a 4.6 kb HindIII fragment for EaeI.  The R and M genes of both systems converge
and overlap by 14 nucleotides.  Previously, we found that M.EcoHK31I consisted of two
subunits, (alpha and beta), with the beta subunit being translated from an alternative open
reading frame within the gene encoding the alpha subunit.  Sequence comparison between the
EcoHK31I and EaeI systems reveals striking similarity.  The aeaIM gene also encodes alpha and
beta polypeptides of 309 and 176 amino acids which share 95% and 97% identity, respectively,
with those of ecoHK31IM.  eCoHK31IR and aeaIR encode proteins of 318 and 315 aa, respectively,
which share 92% identity but are otherwise unique in the GenBank database.  The EaeI and the
EcoHK31I R-M systems were found to be flanked by genes coding for integrases.  It is possible
that these integrases have facilitated the transfer of this system among different bacterial
species.

<>

<1>Lee, K.-F., Shi, S.-D., Kam, K.-M., Shaw, P.-C.
<2>Restriction endonuclease from thermophilic bacterial species III.  Isolation and characterization of BsiHKAI.
<3>Nucleic Acids Res.
<4>20
<5>921
<6>1992
<7>BsiHKAI is a type II restriction endonuclease from a Bacillus
stearothermophilus strain isolated from soil.  BsiHKAI in 1l culture (4.7 g wet
cells) was purified by a DEAE-sephacel column (30 ml bed volume).  Enzyme
eluted at about 0.3 M NaCl was dialysed against buffer A (1) and loaded onto a
heparin column (8 ml bed volume).  Enzyme eluted at 0.4 - 0.5 M NaCl was
dialysed against buffer B (1) and loaded onto an FPLC Mono Q anion exchange
column.  Enzyme was eluted at 0.3 - 0.4 M NaCl. The purified enzyme was used to
digest various DNA with known sequences (Fig. 1).  The sizes and numbers of
fragments produced are identical to those cleaved by mesophilic enzyme HgiAI
which recognizes 5'G(AT)GC(AT)C3' (2).  The cleavage site of BsiHKA I was
determined according to (3) using pUC18 DNA as the template and a 17 mer
oligonucleotide with sequence 5'CAGCACTGACCCTTTTG 3' as the primer.  The primer
was annealed to position 359-375 of the denatured pUC18 DNA and was extended
through the BsiHKAI site at position 444.  Figure 2 shows that the cleavage
product of reaction I comigrates with the band corresponding to nucleotide T in
the sequence GAGCTC and reaction II comigrates with the band corresponding to
the first G nucleotide.  Therefore, BsiHKAI recognizes and cleaves
5'G(AT)GC(AT)!C3'.  The optimal buffer for this enzyme was medium salt (4) and
optimal reaction temperature 60C.

<>

<1>Lee, K.F., Ling, J.M.-L., Kam, K.-M., Clark, D.R., Shaw, P.-C.
<2>Restriction endonucleases in clinical isolates of Shigella spp.
<3>J. Med. Microbiol.
<4>46
<5>949-952
<6>1997
<7>Thirteen restriction endonuclease-containing strains were isolated from a collection of 186
clinical isolates of Shigella spp.  Among these, eight and five isolates carried isoschizomers
of EcoRII and NciI, respectively.  The former restriction-modification system was homologous
to that of EcoRII and was located on plasmids with sizes of 46.6 or 55.6 kb.  Isolates
producing NciI isoschizomers contained a 5.7-kb nontransferable plasmid.  Together with
antimicrobial susceptibility tests and plasmid profile studies, it is concluded that these two
R-M systems are not widely spread but confined to strains with similar antibiotic resistance
and plasmid profile.

<>

<1>Lee, K.S.
<2>Cytotoxicity of patulin and its effect on lambda DNA cleavage by restriction endonuclease.
<3>Korean J. Toxicol.
<4>7
<5>157-163
<6>1991
<7>The effect of patulin, a mycotoxin, on the growth of Escherichia coli cells was investigated.
E. coli cell elongation usually shown during SOS-response for DNA repair was induced by 20 ug
of patulin per ml. After staining the E. coli chromosome with a fluorescent dye (DAPI, 4',
6-diamino-2-phenyl-indole), chromosomal DNA partitioning was not affected by patulin. This
observation indicates that patulin acts as a DNA damaging agent which is effective for E. coli
cell elongation introduced by the inhibition of septum formation. Moreover, patulin inhibits
the cleavage reaction of some restriction endonucleases (StyI, AsnI, BamHI, HindIII) on lambda
phage DNA. The restriction endonucleolytic fragments of lambda phage DNA preincubated with
patulin were larger than standard digestion products. The lambda DNA recognition site
(cleavage site) for StyI (C^CAAGG) was very sensitive to patulin, while the site for AsnI had
a very low reactivity with patulin. It seems likely that the cytosine of StyI recognition
sequences in lambda phage DNA would be a highly possible target nucleotide for patulin.

<>

<1>Lee, K.S., Seo, S.Y., Lee, S.W., Rho, H.M.
<2>Cloning and Expression of the Pst I Restriction Endonuclease Gene.
<3>Korean Biochem. J.
<4>15
<5>315-329
<6>1982
<7>We report the cloning of the PstI restriction-modification system of
Providencia stuartii 164.  The DNA isolated from the native PstI producing
strain, P. stuartii 164, was completely digested with HindIII and inserted into
the HindIII target site of pBR322.  The cloned strain of E. coli was selected
on the basis of acquired resistance to bacteriophage Phi80 infection and PstI
endonuclease productivity.  Plasmid and host DNA from the cloned E. coli were
not digested by PstI, and the strain produced PstI restriction endonuclease.
These results indicated that both the restriction enzyme and modification
(methylase) genes had been cloned and were being expressed.  The plasmid (pRL
829) isolated from the cloned strain contains an insert of approximately 3,700
base pairs.  The cloned E. coli produces approximately 15 times more PstI
endonuclease activity than does the native strain under similar conditions of
culture.

<>

<1>Lee, K.W., Arumugam, K., Purbojati, R.W., Tay, Q.X., Williams, R.B., Kjelleberg, S., Rice, S.A.
<2>Draft Genome Sequence of Klebsiella pneumoniae Strain KP-1.
<3>Genome Announcements
<4>1
<5>e01082-13
<6>2013
<7>Klebsiella pneumoniae is ubiquitous in the environment and is a member of a three-species
biofilm model. We compared the genome sequence of an environmental
isolate, K. pneumoniae strain KP-1, to those of two clinical strains (NTUH-K2044
and MGH 78578). KP-1 possesses strain-specific prophage sequences that
distinguish it from the clinical strains.

<>

<1>Lee, L., Teh, L., Zainuddin, Z., Salleh, M.
<2>The Genome Sequence of a Type ST239 Methicillin-Resistant Staphylococcus aureus Isolate from a Malaysian Hospital.
<3>Standards in Genomic Sciences
<4>9
<5>933-939
<6>2014
<7>We report the genome sequence of a healthcare-associated MRSA type ST239 clone isolated from a
patient with septicemia in Malaysia. This clone typifies the
characteristics of ST239 lineage, including resistance to multiple antibiotics
and antiseptics.

<>

<1>Lee, M., Roh, S.W., Lee, H.W., Yim, K.J., Kim, K.N., Bae, J.W., Choi, K.S., Jeon, Y.J., Jung, W.K., Kang, H., Hyun, C.G., Kim, D.
<2>Draft Genome Sequence of Pedobacter agri PB92T, Which Belongs to the Family Sphingobacteriaceae.
<3>J. Bacteriol.
<4>194
<5>3738
<6>2012
<7>Strain PB92(T) of Pedobacter agri, which belongs to the family Sphingobacteriaceae, was
isolated from soil in the Republic of Korea. The draft
genome of strain PB92(T) contains 5,141,552 bp, with a G+C content of 38.0%. This
is the third genome sequencing project of the type strains among the Pedobacter
species.

<>

<1>Lee, N., Valinluck, B., Randriamahefa, A., Rutebuka, O., Ryu, J.
<2>Cloning and mapping of the hsdR genes of KpnAI and KpnBI restriction-modification systems of Klebsiella pneumoniae.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>93
<5>199
<6>1993
<7>Two possible type I restriction-modification systems, KpnAI and KpnBI have been identified in
K. pneumoniae strains M5a1 and GM236 respectively (ASM Abstracts 1989, p180). The hsdR genes
from Sau3AI partially digested chromosomes of both strains were cloned into the BamHI site of
either pBR322 or pUC18. The clones expressing an r+ phenotype in r-m- recipients were
identified among the transformants resistant to non-modified phage SBS. A total of four clones
(8.8 to 14.8 kb) of KpnAI and two (5.8 and 6.2 kb) of KpnBI were obtained. The common portion
of these clones, which supposedly code for hsdR genes, were inferred as a 3.3 kb HindIII
fragment of KpnAI and a 3.9 kb EcoRI fragment for KpnBI respectively. Complementation results
with r-m- mutants suggest that the 6.2 kb fragment of the KpnBI clone contains a second gene
(either hsdM or hsdS) as well.

Clones of each type were hybridized to a membrane which contains pulsed-field gel
electrophoresed fragments of the chromosome of the corresponding strain. The KpnAI clone was
mapped on a 19 Kb XbaI fragment, located at about 40 min on the recently constructed physical
map of the M5a1 chromosome (ASM Abstracts 1992, p212). Similarly, the KpnBI clone was mapped
on a 40 kb XbaI fragment of the GM236 chromosome.


<>

<1>Lee, N.S., Rutebuka, O., Arakawa, T., Bickle, T.A., Ryu, J.
<2>KpnAI, a new type I restriction-modification system in Klebsiella pneumoniae.
<3>J. Mol. Biol.
<4>271
<5>342-348
<6>1997
<7>The KpnAI restriction-modification system has been identified in Klebsiella pneumoniae strain
M5a1.  The restriction gene of KpnAI was first cloned into pBR322 using an r-m+ M5a1
derivative and phage SBS for screening.  Subsequently, an adjacent DNA fragment showing
modification activity was cloned into pUC19.  A total of 7.2 kb DNA sequencing data revealed
three open reading frames, corresponding to hsdR, hsdM and hsdS genes of type I R-M systems.
The predicted hsdR, hsdM and hsdS-coded peptides shared 95%, 98% and 44% identity,
respectively, with the corresponding peptides of the recently identified StySBLI system, a
prototype of the type ID family.  This high homology suggests that KpnAI is also a member of
the type ID family.  The KpnAI system seems to be the first type I system identified in
Klebsiella species.

<>

<1>Lee, N.S., Ryu, J.
<2>DNA sequence of the hsdR region of KpnBI R-M system of Klebsiella pneumoniae GM236.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>237
<6>1994
<7>The hsdR gene of the KpnBI restriction-modification (R-M) system of Klebsiella pneumoniae
GM236 was cloned (ASM Abstracts 1993, p199). The DNA sequence of a 3.5 kb BamHI-EcoRI fragment
that includes the hsdR gene was determined by the Sanger's dideoxy method. An open reading
frame (ORF) of 3,039 bp (nucleotides 219 to 3,257) is the coding region for the HsdR
polypeptide. The 1,013 amino acid product deduced from the nucleotide sequence would have a
molecular weight of 116,312 daltons, in excellent agreement with the 116 kD size estimated
from the electrophoretic mobility of the putative HsdR protein in SDS polyacrylamide gels.

The nucleotide sequence of the hsdR gene showed no significant similarity to any other
sequence in the GenBank database. However, the amino acid sequence scored highly with
EcoR124/3I, a type I restriction enzyme. After alignment of the two proteins, two conserved
domains, an ATP binding and a DNA binding domain, were found.

Seven small ORFs were identified within a few kb on either side of the hsdR gene. When
subcloned, none of these ORFs showed any modification activity. This suggests that the KpnBI
system differs from other R-M systems where the hsdR and hsdM genes are adjacent. However,
complementation with an r-KpnBIm-KpnBI mutant suggests that one downstream ORF is a positive
effector for expression of the KpnBI modification activity. Therefore, a control gene for the
KpnBI modification genes may be located next to the hsdR gene.


<>

<1>Lee, R.D., Jospin, G., Coil, D.A., Eisen, J.A.
<2>Draft Genome Sequence of the Endosymbiont 'Candidatus Ruthia magnifica' UCD-CM (Phylum Proteobacteria).
<3>Genome Announcements
<4>2
<5>e00717-14
<6>2014
<7>Here, we present the draft genome of the endosymbiont 'Candidatus Ruthia magnifica' UCD-CM,
a member of the phylum Proteobacteria, found from the gills of
a deep-sea giant clam, Calyptogena magnifica. The assembly consists of 1,160,249
bp contained in 18 contigs.

<>

<1>Lee, S., Cho, Y.J., Lee, A.H., Chun, J., Ha, N.J., Ko, G.
<2>Genomic sequence of Lactobacillus ruminis SPM0211, isolated from a fecal sample from a healthy Korean.
<3>J. Bacteriol.
<4>193
<5>5034
<6>2011
<7>Lactobacillus ruminis SPM0211 is a potential probiotic strain that shows antimicrobial
activity against emerging pathogens. Here, we present the
draft genomic sequence of L. ruminis SPM0211, isolated from a fecal sample
from a healthy Korean, and we describe both the common and unique features
of this strain.

<>

<1>Lee, S., Ward, T.J., Siletzky, R.M., Kathariou, S.
<2>Two Novel Type II Restriction-Modification Systems Occupying Genomically Equivalent Locations on the Chromosomes of Listeria monocytogenes Strains.
<3>Appl. Environ. Microbiol.
<4>78
<5>2623-2630
<6>2012
<7>Listeria monocytogenes is responsible for the potentially life-threatening food-borne disease
listeriosis. One
epidemic-associated clonal group of L. monocytogenes, epidemic clone I
(ECI), harbors a Sau3AI-like restriction-modification (RM) system also
present in the same genomic region in certain strains of other
lineages. In this study, we identified and characterized two other,
novel type II RM systems, LmoJ2 and LmoJ3, at this same locus. LmoJ2
and LmoJ3 appeared to recognize GCWGC (W = A or T) and GCNGC,
respectively. Both RM systems consisted of genes with GC content below
the genome average and were in the same genomic region in strains of
different serotypes and lineages, suggesting site-specific horizontal
gene transfer. Genomic DNA from the LmoJ2 and LmoJ3 strains grown at
various temperatures (4 to 42 degrees C) was resistant to digestion
with restriction enzymes recognizing GCWGC or GCNGC, indicating that
the methyltransferases were expressed under these conditions. Phages
propagated in an LmoJ2-harboring strain exhibited moderately increased
infectivity for this strain at 4 and 8 degrees C but not at higher
temperatures, while phages propagated in an LmoJ3 strain had
dramatically increased infectivity for this strain at all temperatures.
Among the sequenced Listeria phages, lytic phages possessed
significantly fewer recognition sites for these RM systems than
lysogenic phages, suggesting that in lytic phages sequence content
evolved toward reduced susceptibility to such RM systems. The ability
of LmoJ2 and LmoJ3 to protect against phages may affect the efficiency
of phages as biocontrol agents for L. monocytogenes strains harboring
these RM systems.

<>

<1>Lee, S.F., Forsberg, C.W., Gibbins, A.M.
<2>Type II DNA restriction-modification system and an endonuclease from the ruminal bacterium Fibrobacter succinogenes S85.
<3>J. Bacteriol.
<4>174
<5>5275-5283
<6>1992
<7>Fibrobacter succinogenes is an important cellulolytic bacterium found in the rumen and cecum
of herbivores.  Numerous attempts to introduce foreign DNA into F. succinogenes S85 have
failed, suggesting the presence of genetic barriers in this organism.  Results from this study
clearly demonstrate that F. succinogenes S85 possesses a type II restriction endonuclease,
FsuI, which recognizes the sequence 5'-GG(A/T)CC-3'.  Analysis of the restriction products
on sequencing gels showed that FsuI cleaves between the two deoxyguanosine residues, yielding
a 3-base 5' protruding end.  These data demonstrate that FsuI is an isoschizomer of AvaII.  A
methyltransferase activity has been identified in the cell extract of F. succinogenes S85.
This activity modified DNA in vitro and protected the DNA from restriction by FsuI and AvaII.
DNA modified in vivo by a cloned methylase gene, which codes for M.Eco47II, also protected the
DNA from restriction by FsuI, suggesting that FsuI is inhibited by methylation at one or both
deoxycytosine residues of the recognition sequence.  The methyltransferase activity in F.
succinogenes S85 is likely modifying the same deoxycytosine residues, but the exact site(s) is
unknown.  A highly active DNase (DNase A) was also isolated from the cell extract of this
organism.  DNase A is an endonuclease which showed high activity to all forms of DNA
(single-stranded, double-stranded, linear, and circular) but no activity on RNA.  In vitro,
the DNase A hydrolyzed F. succinogenes S85 DNA extensively, indicating the lack of protection
against hydrolysis by this enzyme.  In the presence of Mg2+, DNA was hydrolyzed to fragments
of 8 to 10 nucleotides in length.  The presence of DNase A and the type II
restriction-modification system of F. succinogenes S85 may be the barriers preventing the
introduction of foreign DNA into this bacterium.  Author agreed to change name to FssI to
avoid confusion with previous enzyme named FsuI.

<>

<1>Lee, S.G., Koh, H.Y., Lee, J.H., Kang, S.H., Kim, H.J.
<2>Draft Genome Sequence of Moritella dasanensis Strain ArB 0140, a Psychrophilic Bacterium Isolated from the Arctic Ocean.
<3>J. Bacteriol.
<4>194
<5>5452-5453
<6>2012
<7>The psychrophilic bacterium Moritella dasanensis strain ArB 0140 was isolated near a glacier
in Kongsfjorden, Svalbard Archipelago, Norway. Here we report a
4.89-Mb draft genome sequence of Moritella dasanensis ArB 0140, which could
provide comprehensive information on a psychrophilic mechanism in extreme
environments.

<>

<1>Lee, S.H., Jin, H.M., Lee, H.J., Kim, J.M., Jeon, C.O.
<2>Complete Genome Sequence of the BTEX-Degrading Bacterium Pseudoxanthomonas spadix BD-a59.
<3>J. Bacteriol.
<4>194
<5>544
<6>2012
<7>Pseudoxanthomonas spadix BD-a59, able to metabolize all six BTEX (benzene, toluene,
ethylbenzene, and o-, m-, and p-xylene) compounds, was isolated
from gasoline-contaminated sediment. Here, we report the complete 3.45-Mb
genome sequence and annotation of strain BD-a59. These advance the
understanding of strain BD-a59's genomic properties and pollutant
metabolic versatility.

<>

<1>Lee, S.H., Jung, J.Y., Jeon, C.O.
<2>Draft Genome Sequence of Salimicrobium sp. Strain MJ3, Isolated from Myulchi-Jeot, Korean Fermented Seafood.
<3>J. Bacteriol.
<4>194
<5>6695
<6>2012
<7>Salimicrobium sp. strain MJ3 was isolated from myulchi-jeot, traditional fermented seafood
made from anchovy in South Korea. Here we announce the draft
genome sequence of Salimicrobium sp. MJ3 with 2,717,782 bp, which consists of 45
contigs (>500 bp in size), and provide a description of their annotation.

<>

<1>Lee, S.H., Jung, J.Y., Lee, S.H., Jeon, C.O.
<2>Complete Genome Sequence of Leuconostoc kimchii Strain C2, Isolated from Kimchi.
<3>J. Bacteriol.
<4>193
<5>5548
<6>2011
<7>Leuconostoc kimchii strain C2 was isolated from fermented kimchi in Korea. Here we announce
the complete genome sequence of Leuconostoc kimchii
strain C2, consisting of a 1,877,174-bp chromosome with a G+C content of
37.9% and no plasmid and describe major findings from its annotation.

<>

<1>Lee, S.H., Rho, H.M.
<2>Cloning of PstI methylase gene.
<3>Korean J. Genet.
<4>7
<5>42-48
<6>1985
<7>The clonings of PstI restriction-modification system were reported in two cases.  One was by
Walder et al. (1981) and the other was by Lee et al. (1982). pRL829 constructed by Lee et al.
was a recombinant plasmid containing 3.7kb insert fragment at HindIII site of pBR322, which
encoded whole PstI restriction-modification system.  In this research, a new recombinant
plasmid, pRL830, containing only PstI methylase gene was constructed from pRL829 by partial
digestion with HincII.  pRL830 contained 2.2kb insert DNA out of 3.7kb insert of pRL829.  The
host DNA and plasmid from the cells transformed with pRL830 were not cleaved by PstI,
suggesting that PstI methylase gene was cloned and expressed in the cells.  However, E. coli
transformed with pRL830 was easily infected and lysed by bacteriophage Phi80, indicating that
PstI endonuclease gene was at least inactivated or removed from pRL830.  Soluble fractions
from the E. coli containing pRL830 were passed through the DEAE-cellulose and phosphocellulose
columns.  Eluted enzyme fraction with salt gradient were assayed for the PstI endonuclease and
methylase activities. Results showed that PstI methylase activity was detected, whereas PstI
endonuclease activity was not.  It was convinced pRL830 encoded PstI methylase gene, but not
endonuclease gene.

<>

<1>Lee, S.J., Lee, Y.J., Jeong, H., Lee, S.J., Lee, H.S., Pan, J.G., Kim, B.C., Lee, D.W.
<2>Draft Genome Sequence of Virgibacillus halodenitrificans 1806.
<3>J. Bacteriol.
<4>194
<5>6332-6333
<6>2012
<7>Virgibacillus halodenitrificans 1806 is an endospore-forming halophilic bacterium isolated
from salterns in Korea. Here, we report the draft genome sequence of V.
halodenitrificans 1806, which may reveal the molecular basis of osmoadaptation
and insights into carbon and anaerobic metabolism in moderate halophiles.

<>

<1>Lee, S.J., Lee, Y.J., Park, G.S., Kim, B.C., Lee, S.J., Shin, J.H., Lee, D.W.
<2>Draft Genome Sequence of an Anaerobic and Extremophilic Bacterium, Caldanaerobacter yonseiensis, Isolated from a Geothermal Hot Stream.
<3>Genome Announcements
<4>1
<5>e00923-13
<6>2013
<7>Caldanaerobacter yonseiensis is a strictly anaerobic, thermophilic, spore-forming bacterium,
which was isolated from a geothermal hot stream in Indonesia. This
bacterium utilizes xylose and produces a variety of proteases. Here, we report
the draft genome sequence of C. yonseiensis, which reveals insights into the
pentose phosphate pathway and protein degradation metabolism in thermophilic
microorganisms.

<>

<1>Lee, S.J., Lee, Y.J., Ryu, N., Park, S., Jeong, H., Lee, S.J., Kim, B.C., Lee, D.W., Lee, H.S.
<2>Draft Genome Sequence of the Thermophilic Bacterium Anoxybacillus kamchatkensis G10.
<3>J. Bacteriol.
<4>194
<5>6684-6685
<6>2012
<7>Anoxybacillus kamchatkensis G10 is a spore-forming thermophilic bacterium isolated from a hot
spring in Indonesia. Here, we report the draft genome
sequence of A. kamchatkensis G10 that may reveal insights into aerobic/anaerobic
metabolisms and carbon utilization in moderate thermophiles.

<>

<1>Lee, S.P., Porter, D., Chirikjian, J.G., Knutson, J.R., Han, M.K.
<2>A fluorometric assay for DNA cleavage reactions characterized with BamHI restriction endonuclease.
<3>Anal. Biochem.
<4>220
<5>377-383
<6>1994
<7>Fluorescently labeled oligonucleotides and DNA fragments have promise in nucleic acid research
with applications that include DNA hybridization, automated DNA sequencing, fluorescence
anisotropy, and resonance energy transfer studies. Past concerns with fluorescent-labeled DNA
arose from interactions between fluorophores and DNA that result in quenched fluorescence.
This quenching phenomenon is most problematic in fluorescence resonance energy transfer
studies because quenching of the donor fluorescence could result from either resonance energy
transfer or nontransfer effects. In the present study, relief of nontransfer quenching of a
14-mer fluorescein 5-isothiocyanate (FITC)-labeled oliognucleotide containing the BamHI
restriction site was characterized with both steady-state and time-resolved fluorescence
techniques. The FITC-labeled single strand was best fit by a triexponential decay with
lifetimes of 0.5, 2.7, and 4.2 ns. The 4.2-ns component was found to contribute more than 80%
of the total steady-state intensity. Upon annealing with an unmodified complementary strand,
the contribution from the 4.2-ns component was significantly decreased, resulting in twofold
quenching of total fluorescence. We reasoned that this quenching phenomenon should be a
reversible process and could be employed to study strand separation processes in molecular
biology. Hence, cleavage of the fluorescently labeled substrate was examined using DNase I and
BamHI restriction endonuclease. Our results show that the quenched fluorescence is totally
recovered upon cleavage (compared to that of the single strand). The extent of cleavage
measured by fluorescence was confirmed by nondenaturing polyacrylamide gel electrophoresis
analysis. We believe this fluorescence dequenching technique may be used to quantify the
kinetics of other DNA strand separation and cleavage processes in molecular biology.

<>

<1>Lee, S.P., Porter, D.K., Chirikjian, J.G., Knutson, J.R., Han, M.K.
<2>The development of a fluorescence assay for the BamHI restriction endonuclease utilizing modified oligonucleotides.
<3>Biophys. J.
<4>66
<5>A163
<6>1994
<7>Fluorescent labeled oigonucleotides and DNA fragments have promise in nucleic acid research
including DNA hybridization, automated DNA sequencing, and fluorescence anisotropy or
resonance energy transfer studies. One concern with fluorescent-labeled DNA is that the
interactions between fluorophores and DNA may result in quenching of the probe fluorescence.
This quenching phenomenon is most problematic in fluorescence enegy transfer studies because
donor quenching can occur as a result of both resonance transfer and these non-transfer
effects. In the present studies, the nontransfer quenching of a 14-mer FITC-labeled
oligonucleotide with the BamHI restriction site was characterized with both steady-state and
time-resolved fluoescence techniques. The FITC-labeled single strand was best fit by
triexponential decay with lifetimes of 0.5, 2.7, and 4.2 ns. The 4.2 ns component was found to
contribute more than 80% of total steady-state intensity. Upon annealing with an unmodified
complementary strand, the 4.2 ns component was significantly decreased, resulting in 2-fold
quenching of total fluorescence. We examined the reversible quenching process and applied our
findings to quantify DNA strand separation and cleavage processes. Utilizing this information,
we developed a continuous fluorescence assay for the restriction endonucleases reaction for
BamHI.

<>

<1>Lee, S.Y., Kim, B.Y., Ahn, J.H., Song, J., Seol, Y.J., Kim, W.G., Weon, H.Y.
<2>Draft Genome Sequence of the Biocontrol Bacterium Bacillus amyloliquefaciens Strain M27.
<3>J. Bacteriol.
<4>194
<5>6934-6935
<6>2012
<7>Bacillus amyloliquefaciens strain M27 is a biocontrol agent with antagonistic activities
against a wide range of fungal pathogens. Here we present the 3.86-Mb
draft genome sequence of the bacterium with the aims of providing insights into
the genomic basis of its antifungal mechanism and facilitating its application in
the biocontrol of plant diseases.

<>

<1>Lee, S.Y., Kim, S.H., Lee, D.G., Shin, S., Yun, S.H., Choi, C.W., Chung, Y.H., Choi, J.S., Kahng, H.Y., Kim, S.I.
<2>Draft Genome Sequence of Petroleum Oil-Degrading Marine Bacterium Pseudomonas taeanensis Strain MS-3, Isolated from a Crude Oil-Contaminated Seashore.
<3>Genome Announcements
<4>2
<5>e00818-13
<6>2014
<7>Pseudomonas taeanensis MS-3(T), isolated from a crude oil-contaminated seashore in South
Korea, is capable of degrading petroleum oils, such as gasoline, diesel,
and kerosene. Here, we report the draft genome sequence of this strain, which
consists of 5,477,045 bp, with a G+C content of 60.72%.

<>

<1>Lee, S.Y., Mermelstein, L.D., Bennett, G.N., Papoutsakis, E.T.
<2>Vector construction, transformation, and gene amplification in Clostridium acetobutylicum ATCC 824.
<3>Ann. NY Acad. Sci.
<4>665
<5>39-51
<6>1992
<7>The acetone-butanol-ethanol fermentation by the gram-positive, endo-
spore-forming, obligate
anaerobe Clostridium acetobutylicum has received renewed interest because recombinant
DNA technology
offers the possibility of creating genetically engineered strains as potential producers of
these solvents from
renewable biomass.  The ability to express homologous and heterologous genes in C.
acetobutylicum and
alter the cellular metabolism in a beneficial way is critical to the creation of overproducing
strains for
industrial applications.  It is also necessary to understand the molecular mechanisms
involved in metabolic
regulation in order to rationally manipulate metabolic pathways using recDNA technology.

<>

<1>Lee, S.Y., Yun, S.H., Choi, C.W., Lee, D.G., Choi, J.S., Kahng, H.Y., Kim, S.I.
<2>Draft Genome Sequence of an Aniline-Degrading Bacterium, Burkholderia sp. K24.
<3>Genome Announcements
<4>2
<5>e01250-14
<6>2014
<7>Burkholderia sp. K24 is an aniline-degrading soil bacterium that utilizes aniline and its
analogues as sole carbon and nitrogen sources. Here, we report the draft
genome sequence of this strain that consists of 8,344,181 bp, with a G+C content
of 61.7%.

<>

<1>Lee, W.-K., An, Y.-S., Kim, K.-H., Kim, S.-H., Song, J.-Y., Ryu, B.-D., Choi, Y.-J., Yoon, Y.-H., Baik, S.-C., Rhee, K.-H., Cho, M.-J.
<2>Construction of a Helicobacter pylori-Escherichia coli shuttle vector for gene transfer in Helicobacter pylori.
<3>Appl. Environ. Microbiol.
<4>63
<5>4866-4871
<6>1997
<7>In this study, a Helicobacter pylori-Escherichia coli shuttle vector was constructed for
transferring DNA into H. pylori.  The smallest cryptic plasmid (1.2 kb), pHP489, among those
harbored by 77 H. pylori isolates was selected as a base replicon for constructing vectors.
HindIII-digested pHP-489 was ligated with a kanamycin resistance gene [aph(3')-III], which
originated from Campylobacter jejuni, to produce the recombinant plasmid pHP489K.  pHP489K was
efficiently transformed into and stably maintained in H. pylori strains.  The shuttle vector
pBHP489K (3.6kb) was constructed by the recombination of pHP489, ColE1, and aph(3')-III
sequences.  PBHP489K was reciprocally transformed into and maintained in both H. pylori and E.
coli.  Introduction of the shuttle vector clone DNA (pBHP489K/AB; 6.7 kb), containing the ureA
and ureB genes of H. pylori, into urease-negative mutants of H. pylori led to the restoration
of their urease activity.  The transformants were confirmed to contain the incoming plasmid
DNA.  PBHP489K satisfied the requirements for an H. pylori-E. coli shuttle vector, implying
that it might be a useful vector for investigating pathogenicity and restriction-modification
systems of H. pylori.

<>

<1>Lee, Y.-F., Tawfik, D.S., Griffiths, A.D.
<2>Investigating the target recognition of DNA cytosine-5 methyltransferase HhaI by library selection using in vitro compartmentalization.
<3>Nucleic Acids Res.
<4>30
<5>4937-4944
<6>2002
<7>In vitro compartmentalisation (IVC), a technique for selecting genes encoding enzymes based on
compartmentalising gene translation and enzymatic reactions in emulsions, was used to
investigate the interaction of the DNA cytosine-5 methyltransferase M.HhaI with its target DNA
(5'-GCGC-3'). Crystallography shows that the active site loop from the large domain of
M.HhaI interacts with a flipped-out cytosine (the target for methylation) and two target
recognition loops (loops I and II) from the small domain make almost all the other
base-specific interactions. A library of M.HhaI genes was created by randomising all the loop
II residues thought to make base-specific interactions and directly determine target
specificity. The library was selected for 5'-GCGC-3' methylation. Interestingly, in 11
selected active clones, 10 different sequences were found and none were wild-type. At two of
the positions mutated (Ser252 and Tyr254) a number of different amino acids could be
tolerated. At the third position, however, all active mutants had a glycine, as in wild-type
M.HhaI, suggesting that Gly257 is crucial for DNA recognition and enzyme activity. Our results
suggest that recognition of base pairs 3 and 4 of the target site either relies entirely on
main chain interactions or that different residues from those identified in the crystal
structure contribute to DNA recognition.

<>

<1>Lee, Y.-H., Blakesley, R.W., Smith, L.A., Chirikjian, J.G.
<2>Preparation and properties of insolubilized restriction endonucleases.
<3>Nucleic Acids Res.
<4>5
<5>679-689
<6>1978
<7>Type II restriction endonucleases BamHI and EcoRI were covalently coupled to
Sepharose.  These insolubilized enzymes generated fragment patterns for several
viral DNAs identical to those produced by the respective free enzymes.
Conditions for optimal activity were similar for both bound and unbound form of
the enzymes.  Insolubilization improved thermal stability of BamHI and EcoRI.
The bound enzyme can be recovered from reaction mixtures and reused several
times.  Upon storage at 4C, coupled endonucleases remained stable for several
months.

<>

<1>Lee, Y.-W., Broday, L., Costa, M.
<2>Effects of nickel on DNA methyltransferase activity and genomic DNA methylation levels.
<3>Mutat. Res.
<4>415
<5>213-218
<6>1998
<7>Methylation of DNA plays an important role in organizing the genome into transcriptionally
active and inactive zones.  Nickel compounds cause chromatin condensation and DNA methylation
in the transgenic gpt+ Chinese hamster cell line (G12).  Here we show that nickel is an
inhibitor of cytosine 5-methyltransferase activity in vivo and in vitro.  In living cells,
this inhibition is transient and following a recovery period after nickel treatment, Mtase
activity slightly rebounds.  Genomic DNA methylation levels are also somewhat decreased
following nickel treatment, but with time, there is an elevation of total DNA methylation
above basal levels and before any rebound of methyltransferase activity.  These results
suggest that nickel exposure can elevate total genomic DNA methylation levels even when DNA
methyltransferase activity is depressed.  These findings may explain the hypermethylation of
senescence and tumor suppressor genes found during nickel carcinogenesis and support the model
of a direct effect of Ni2+ on chromatin leading to de novo DNA methylation.

<>

<1>Lee, Y.D., Chang, H.I., Park, J.H.
<2>Complete genomic sequence of virulent Cronobacter sakazakii phage ESSI-2 isolated from swine feces.
<3>Arch. Virol.
<4>156
<5>721-724
<6>2011
<7>A newly identified virulent Cronobacter sakazakii phage, ESSI-2, was
isolated from fecal samples from swine. The morphological characteristics
evident under a transmission electron microscope indicated that phage
ESSI-2 belonged to the family Myoviridae. The genome of phage ESSI-2
comprised a double-stranded DNA of 28,765 bp with a G+C content of 55.17%.
Bioinformatic analysis of the phage genome identified 36 putative open
reading frames (ORFs). The genome of phage ESSI-2 was not significantly
similar to that of a previously reported bacteriophage of the members of
Enterobacteriaceae. A lysogeny module was found within the genome of this
virulent phage.

<>

<1>Lee, Y.H., Blakesley, R., Smith, L.A., Chirikjian, G.
<2>Preparation and properties of immobilized sequence specific endonucleases.
<3>Methods Enzymol.
<4>65
<5>173-182
<6>1980
<7>The type II restriction endonucleases have become valuable reagents for research in molecular
biology.  This is primarily due to their unique property of cleaving DNAs at a limited number
of specific sites.  These enzymes have been employed for use in physical DNA mapping, plasmid
construction, and gene cloning.  Most of these enzymes recognize and cleave DNA at specific
sequences, which are usually in the form of a palindrome.  Since there is evidence implying
that these endonucleases are membrane bound, one approach to studying them is to determine
what effect insolubilization (i.e., mimicking membrane binding) has on their activity.

<>

<1>Lee, Y.H., Chirikjian, J.G.
<2>Sequence-specific endonuclease BglI.
<3>J. Biol. Chem.
<4>254
<5>6838-6841
<6>1979
<7>The sequence-specific endonuclease BglI from Bacillus globigii (RUB561) has
been purified to homogeneity as determined by denaturing polyacrylamide gel
analysis.  The active form of the enzyme is a single polypeptide with a
molecular weight of 32,000.  The enzyme requires Mg2+ in the reaction mixture
and displays a broad pH and monovalent cation requirement.  BglI is not
sensitive to sulfhydryl reagents but was affected by reagents that modify
lysine and arginine residues.  When lysine residues were modified by pyridoxal
5'-phosphate, both binding and catalysis were diminished while modification of
arginine residues by 2,3-butanedione inhibited the enzyme activity but had no
effect on its binding properties.

<>

<1>Lee, Y.H., Smith, L.A., Blakesley, R.W.
<2>Preparation and properties of matrix-bound restriction endonucleases.
<3>Fed. Proc.
<4>37
<5>1414
<6>1978
<7>Properties of these specific endonucleases have made them valuable reagents for
several areas of research.  These enzymes are present in bacterial cells
although functions for most such enzymes are not known.  In as much as several
restriction endonucleases are potentially membrane bound, we have undertaken to
investigate the effect of insolubilization on their stability and reaction
characteristics.  Using cyanogen bromide activated sepharose we have covalently
coupled several enzymes including the widely used BamHI, EcoRI and HindIII.
Matrix bound ezymes digest DNAs, generating an identical pattern to those
produced by the respective free enzyme, indicating no change in specificity.
Specific activities for bound enzymes were 100-4000 units/ml of sepharose,
where a unit digests 1 microgram DNA at 37C in 1 hr.  Coupled enzymes display
high thermal stability making them useful in probing DNA structure at high
temperatures.  Coupled enzymes are reusable, can be removed rapidly from
reaction mixtures, and adapted to a flow-through system.

<>

<1>Lee, Y.J., Lee, S.J., Jeong, H., Kim, H.J., Ryu, N., Kim, B.C., Lee, H.S., Lee, D.W., Lee, S.J.
<2>Draft Genome Sequence of Bacillus oceanisediminis 2691.
<3>J. Bacteriol.
<4>194
<5>6351-6352
<6>2012
<7>Bacillus oceanisediminis 2691 is an aerobic, Gram-positive, spore-forming, and moderately
halophilic bacterium that was isolated from marine sediment of the
Yellow Sea coast of South Korea. Here, we report the draft genome sequence of B.
oceanisediminis 2691 that may have an important role in the bioremediation of
marine sediment.

<>

<1>Lee, Y.J., Lee, S.J., Kim, S.H., Lee, S.J., Kim, B.C., Lee, H.S., Jeong, H., Lee, D.W.
<2>Draft Genome Sequence of Bacillus endophyticus 2102.
<3>J. Bacteriol.
<4>194
<5>5705-5706
<6>2012
<7>Bacillus endophyticus 2102 is an endospore-forming, plant growth-promoting rhizobacterium
isolated from a hypersaline pond in South Korea. Here we present
the draft sequence of B. endophyticus 2102, which is of interest because of its
potential use in the industrial production of algaecides and bioplastics and for
the treatment of industrial textile effluents.

<>

<1>Lee, Y.W., Clanton, D., Chirikjian, J.G.
<2>Subpalindromic sequences as inhibitors for restriction endonucleases.
<3>Fed. Proc.
<4>38
<5>294
<6>1979
<7>BglI from B. globigii was purified to homogeneity using several chromatographic
steps including phenyl-sepharose.  The enzyme is a single polypeptide chain of
32,000 daltons as determined by calibrated techniques.  BglI activity required
Mg++ and was stimulated by NaCl.  When Mn++ replaced Mg++ in the reaction
mixture, no change in specificity was observed.  Arg, and lys, but not
sulfhydryl groups were necessary for activity.  The effect of subpalindromic
sequences to inhibit restriction endonuclease activity was examined using
purified BglI and BamHI.  The degree of inhibition on the BamHI (G7GATCC) was
found to be d(pG)2>d(pC)2>d(pApT) while d(pA)2, d(pGpA) and d(pCpT)2 had no
effect.  In addition, d(pTpC) > d(pG)2=d(pC)2 inhibited BglI GGCCG
AGGCGGCCCT^CGGCC CCGGC^TCCGCCCGGA GCCGG Application of this method in studies
on the mechanism of binding and catalysis for BamHI, BglI and other restriction
endonucleases as well as cognate methylases will be presented.

<>

<1>Lees, N.D., Welker, N.E.
<2>Restriction and modification of bacteriophage in Bacillus stearothermophilus.
<3>J. Virol.
<4>11
<5>606-609
<6>1973
<7>Host-controlled restriction and modification of TP-1C phage and infectious
phage DNA occurs in Bacillus stearothermophilus and is subject to control by
TP-8 or TP-12 prophage.

<>

<1>Lees-Murdock, D.J., McLoughlin, G.A., McDaid, J.R., Quinn, L.M., O'Doherty, A., Hiripi, L., Hack, C.J., Walsh, C.P.
<2>Identification of 11 pseudogenes in the DNA methyltransferase gene family in rodents and humans and implications for the functional loci.
<3>Genomics
<4>84
<5>193-204
<6>2004
<7>DNA (cytosine-5-)-methyltransferase genes are important for normal development in mice and
humans. We describe here 11 pseudogenes spread
among human, mouse, and rat belonging to this gene family, ranging from
1 pseudogene in humans to 7 in rat, all belonging to the Dnmt3
subfamily. All except 1 rat Dnmt3b pseudogene appear to be
transcriptionally silent. Dnmt3a2, a transcript variant of Dnmt3a
starting at an alternative promoter, had the highest number of
processed pseudogenes, while none were found for the canonical Dnmt3a,
suggesting the former transcript is more highly expressed in germ
cells. Comparison of human, mouse, and rat Dnmt3a2 sequences also
suggests that human exon 8 is a recent acquisition. Alignment of the
3'UTR of Dnmt3a2 among the functional genes and the processed
pseudogenes suggested that a second polyadenylation site downstream of
the RefSeq poly(A) was being used in mice, resulting in a longer 3'UTR,
a finding confirmed by RT-PCR in mouse tissues. We also found conserved
cytoplasmic polyadenylation elements, usually implicated in regulating
translation in oocytes, in Dnmt3b and Dnmt1. Expression of DNMT3B in
the mouse oocyte was confirmed by immunocytochemistry. These results
clarify the structure of a number of loci in the three species examined
and provide some useful insights into the structure and evolution of
this gene family.

<>

<1>Lefebure, T., Richards, V.P., Lang, P., Pavinski-Bitar, P., Stanhope, M.J.
<2>Gene Repertoire Evolution of Streptococcus pyogenes Inferred from Phylogenomic Analysis with Streptococcus canis and Streptococcus dysgalactiae.
<3>PLoS ONE
<4>7
<5>E37607
<6>2012
<7>Streptococcus pyogenes, is an important human pathogen classified within the
pyogenic group of streptococci, exclusively adapted to the human host. Our goal
was to employ a comparative evolutionary approach to better understand the
genomic events concomitant with S. pyogenes human adaptation. As part of
ascertaining these events, we sequenced the genome of one of the potential sister
species, the agricultural pathogen S. canis, and combined it in a comparative
genomics reconciliation analysis with two other closely related species,
Streptococcus dysgalactiae and Streptococcus equi, to determine the genes that
were gained and lost during S. pyogenes evolution. Genome wide phylogenetic
analyses involving 15 Streptococcus species provided convincing support for a
clade of S. equi, S. pyogenes, S. dysgalactiae, and S. canis and suggested that
the most likely S. pyogenes sister species was S. dysgalactiae. The
reconciliation analysis identified 113 genes that were gained on the lineage
leading to S. pyogenes. Almost half (46%) of these gained genes were phage
associated and 14 showed significant matches to experimentally verified bacteria
virulence factors. Subsequent to the origin of S. pyogenes, over half of the
phage associated genes were involved in 90 different LGT events, mostly involving
different strains of S. pyogenes, but with a high proportion involving the horse
specific pathogen S. equi subsp. equi, with the directionality almost exclusively
(86%) in the S. pyogenes to S. equi direction. Streptococcus agalactiae appears
to have played an important role in the evolution of S. pyogenes with a high
proportion of LGTs originating from this species. Overall the analysis suggests
that S. pyogenes adaptation to the human host was achieved in part by (i) the
integration of new virulence factors (e.g. speB, and the sal locus) and (ii) the
construction of new regulation networks (e.g. rgg, and to some extent speB).

<>

<1>Lefort, F., Calmin, G., Crovadore, J., Falquet, J., Hurni, J.P., Osteras, M., Haldemann, F., Farinelli, L.
<2>Whole-Genome Shotgun Sequence of Arthrospira platensis Strain Paraca, a Cultivated and Edible Cyanobacterium.
<3>Genome Announcements
<4>2
<5>e00751-14
<6>2014
<7>Here we report the whole-genome shotgun sequence of a Peruvian strain of Arthrospira platensis
(Paraca), a cultivated and edible haloalkaliphilic
cyanobacterium of great scientific, technical, and economic potential.

<>

<1>Lefort, F., Calmin, G., Crovadore, J., Osteras, M., Farinelli, L.
<2>Whole-Genome Shotgun Sequence of Pseudomonas viridiflava, a Bacterium Species Pathogenic to Ararabidopsis thaliana.
<3>Genome Announcements
<4>1
<5>e00116-12
<6>2013
<7>We report here the first whole-genome shotgun sequence of Pseudomonas viridiflava strain
UASWS38, a bacterium species pathogenic to the biological model plant Ararabidopsis thaliana
but also usable as a biological control agent and thus of great scientific interest for
understanding the genetics of plant-microbe interactions.

<>

<1>Lefort, F., Calmin, G., Pelleteret, P., Farinelli, L., Osteras, M., Crovadore, J.
<2>Whole-Genome Shotgun Sequence of Bacillus amyloliquefaciens Strain UASWS BA1, a Bacterium Antagonistic to Plant Pathogenic Fungi.
<3>Genome Announcements
<4>2
<5>e00016-14
<6>2014
<7>We report here the whole-genome shotgun sequence of Bacillus amyloliquefaciens strain UASWS
BA1, isolated from inner wood tissues of a decaying Platanus x
acerifolia tree. This strain proved to be antagonistic to several plant
pathogenic fungi and oomycetes and can be developed as a biological control agent
in agriculture.

<>

<1>LeFrancois, J., Gasc, A.-M., Sicard, M.
<2>Electrotransformation of Streptococcus pneumoniae: Evidence for restriction of the DNA on entry.
<3>Microb. Drug Resist.
<4>3
<5>101-104
<6>1997
<7>Electrotransformation is a method generally used in biotechnology to introduce recombinant DNA
into a wide range of bacteria.  However the mechanism of DNA entry is poorly understood.  We
report that in Streptococcus pneumoniae, a naturally transformable species,
electrotransformation efficiently introduces a plasmid replicon.  DNA is strongly restricted
by the restriction-modification systems DpnI and DpnII which degrade methylated and
nonmethylated DNA respectively at GATC sequences.  This suggests that in electrotransformation
double-strand DNA penetrates into these bacteria without a single-strand step in contrast to
natural transformation.  Single-strand DNA by itself is able to electrotransform very weakly
and linearized double-stranded plasmid DNA yields barely detectable levels of transformants.

<>

<1>Lefrancois, J., Sicard, A.M.
<2>Electrotransformation of Streptococcus pneumoniae: evidence for restriction of DNA on entry.
<3>Microbiology
<4>143
<5>523-526
<6>1997
<7>Electrotransformation is a method generally used in biotechnology to introduce recombinant DNA
into a wide range of bacteria.  However, the mechanism of DNA entry is poorly understood.  We
report that in Streptococcus pneumoniae, a naturally transformable species,
electrotransformation efficiently introduces a plasmid replicon.  DNA is strongly restricted
by the restriction-modification systems DpnI and DpnII which degrade methylated and
non-methylated DNA, respectively, at GATC sequences.  This suggests that in
electrotransformation double-stranded DNA penetrates into these bacteria without a
single-stranded DNA step in contrast to natural transformation.  Single-stranded DNA by itself
is able to electrotransform very weakly and linearized double-stranded plasmid DNA yields
barely detectable levels of transformants.

<>

<1>Legrain, P., Rain, J.C., Colland, F., de Reuse, H., Labigne, A.
<2>Protein-protein interactions in Helicobacter pylori.
<3>International Patent Office
<4>WO 02066501 A
<5>
<6>2002
<7>The present invention relates to protein-protein interactions of Helicobacter pylori.  More
specifically, the present invention relates to complexes of polypeptides or polynucleotides
encoding the polypeptides, fragments of the polypeptides, antibodies to the complexes,
Selected Interacting Domains (SID) which are identified due to the protein-protein
interactions, methods for screening drugs for agents which modulate the interaction of
proteins and pharmaceutical compositions that are capable of modulating the protein-protein
interactions.

<>

<1>Legrain, P., Selig, L., Rain, J.-C.
<2>Collection of prokaryotic DNA for two hybrid systems Helicobacter pylori protein-protein interactions and application thereof.
<3>US Patent Office
<4>US 6916615 A
<5>
<6>2005
<7>The present invention concerns collections of recombinant cell clones derived from a
prokaryotic genome, more particularly from Helicobacter pylori genome, usable for two-hybrid
systems and methods to produce such collections.  The invention further relates to then
identification of H. pylori protein-protein interactions and to the application of said
collections of recombinant cell clones and said identified proteins interactions to the
pharmaceutical and diagnostic field.

<>

<1>Legrain, P., Selig, L., Rain, J.C.
<2>Prokaryotic DNA collection for two-hybrid system, protein-protein interaction of Helicobacter Pylori and their uses.
<3>Japanese Patent Office
<4>JP 2002542821 A
<5>
<6>2002
<7>
<>

<1>Legrain, P., Selig, L., Rain, J.C.
<2>Collection of prokaryotic dna for two hybrid systems helicobacter pylori protein-protein interactions and application thereof.
<3>International Patent Office
<4>WO 0066722 A
<5>
<6>2000
<7>The present invention concerns collections of recombinant cell clones derived from a
prokaryotic genome, more particularly from Helicobacter pylori genome, usable for two-hybrid
systems and methods to produce such collections.  The invention further relates to the
identification of H. pylori protein-protein interactions and to the application of said
collections of recombinant cell clones and said identified proteins interactions to the
pharmaceutical and diagnostic field.

<>

<1>Lehours, P., Chaineux, J., Dupouy, S., Menard, A., Morgner, A., Megraud, F.
<2>Identification of a new restriction modification system in Helicobacter pylori.
<3>Int. J. Antimicrob. Agents
<4>24
<5>S211
<6>2004
<7>Restriction modification systems are among the largest categories of specific genes in
Helicobacter pylori.  In an H. pylori strain isolated from a patient with gastric MALT
lymphoma, a new RMS was identified by subtractive hydridisation and chromosome walking.  This
RMS is present in a highly conserved area of the H. pylori genome of J99 and 26,695 reference
strains, but harbouring an ORF with unknown function and without homologies to a methylase or
an endonuclease.  This RMS was also described in parallel by Taiwanese researchers and named
HpyCII.  Its restriction site is identical to BccI (RMS type 1 of Bacteroides cacae).  Our
goal was to study the prevalence and the genetic evolution of this RMS in H. pylori.  The
prevalence of this new RMS was tested by PCR and dot blot hybridisation on a collection of
62II.pylori strains isolated from patients with gastric MALT lymphoma.  The restriciton
profiles obtained with BccI on the chromosomal DNA of these strains were then compared to the
amplicon size obtained with primers designed on the ORF flanking HpyCII.  The prevalence of
HpyCII in our collection of H. pylori strains from gastric MALT lymphoma, was 66.1% (41/62).
For 35 strains (85.4%) the amplicons obtained had the expected size of 3831 bp indicating that
the RMS was complete.  In 30 out of 35 of these strains (85.7%), the DNA obtained was
protected from digestion by BccI as expected while in five cases a digestion was observed.
For six additional strains, a 1636 bp amplicon was obtained indicating that probably a
truncated RMS was prsent.  DNA from these six strains were digested by BccI.  For most H.
pylori strains harbouring HpyCII, this RMS is present in an active form.  The genetic
evolution from an active to an inactive form, then to its complete deletion as in reference
strains, remains to be determined.  To further investigate the question, amplicons from
strains with discrepant results will be sequenced.

<>

<1>Lehours, P., Dupouy, S., Chaineux, J., Ruskone-Fourmestraux, A., Delchier, J.C., Morgner, A., Megraud, F., Menard, A.
<2>Genetic diversity of the HpyC1I restriction modification system in Helicobacter pylori.
<3>Res. Microbiol.
<4>158
<5>265-271
<6>2007
<7>Helicobacter pylori is unique because of the unusually high number and diversity of its
restriction modification (R-M) systems. HpyC1I R-M was
recently characterized and contains an endonuclease which is an
isoschizomer of the endonuclease BccI. This R-M is involved in
adherence to gastric epithelial cells, a crucial step in bacterial
pathogenesis. This observation illustrates the fact that R-M systems
have other putative biological functions in addition to protecting the
bacterial genome from external DNA. The genomic diversity of HpyC1I R-M
was evaluated more precisely on a large collection of H. pylori strains
by PCR, susceptibility to BccI digestion and sequencing. The results
obtained support the mechanism of gain and loss of this R-M system in
the H. pylori genome, and suggest that it is an ancestral system which
gradually disappears during H. pylori evolution, following successive
steps: (1) inactivation of the endonuclease gene, followed or
accompanied by: (2) inactivation of the methyltransferase genes, and
then: (3) definitive loss, leaving only short endonuclease remnant
sequences.

<>

<1>Lehours, P., Vale, F.F., Bjursell, M.K., Melefors, O., Advani, R., Glavas, S., Guegueniat, J., Gontier, E., Lacomme, S., Alves, M.A., Menard, A., Megraud, F., Engstrand, L., Andersson, A.F.
<2>Genome sequencing reveals a phage in Helicobacter pylori.
<3>MBio
<4>2
<5>e00239-11
<6>2011
<7>Helicobacter pylori chronically infects the gastric mucosa in more than half of the human
population; in a subset of this population, its presence
is associated with development of severe disease, such as gastric cancer.
Genomic analysis of several strains has revealed an extensive H. pylori
pan-genome, likely to grow as more genomes are sampled. Here we describe
the draft genome sequence (63 contigs; 26x mean coverage) of H. pylori
strain B45, isolated from a patient with gastric mucosa-associated
lymphoid tissue (MALT) lymphoma. The major finding was a 24.6-kb prophage
integrated in the bacterial genome. The prophage shares most of its genes
(22/27) with prophage region II of Helicobacter acinonychis strain Sheeba.
After UV treatment of liquid cultures, circular DNA carrying the prophage
integrase gene could be detected, and intracellular tailed phage-like
particles were observed in H. pylori cells by transmission electron
microscopy, indicating that phage production can be induced from the
prophage. PCR amplification and sequencing of the integrase gene from 341
H. pylori strains from different geographic regions revealed a high
prevalence of the prophage (21.4%). Phylogenetic reconstruction showed
four distinct clusters in the integrase gene, three of which tended to be
specific for geographic regions. Our study implies that phages may play
important roles in the ecology and evolution of H. pylori. IMPORTANCE:
Helicobacter pylori chronically infects the gastric mucosa in more than
half of the human population, and while most of the infected individuals
do not develop disease, H. pylori infection doubles the risk of developing
gastric cancer. An abundance and diversity of viruses (phages) infect
microbial populations in most environments and are important mediators of
microbial diversity. Our finding of a 24.6-kb prophage integrated inside
an H. pylori genome and the observation of circular integrase
gene-containing DNA and phage-like particles inside cells upon UV
treatment demonstrate that we have discovered a viable H. pylori phage.
The additional finding of integrase genes in a large proportion of
screened isolates of diverse geographic origins indicates that the
prevalence of prophages may have been underestimated in H. pylori. Since
phages are important drivers of microbial evolution, the discovery should
be important for understanding and predicting genetic diversity in H.
pylori.

<>

<1>Lei, H., Oh, S.P., Okano, M., Juttermann, R., Goss, K.A., Jaenisch, R., Li, E.
<2>De novo DNA cytosine methyltransferase activities in mouse embryonic stem cells.
<3>Development
<4>122
<5>3195-3205
<6>1996
<7>It has been a controversial issue as to how many DNA cytosine methyltransferase mammalian
cells have and whether de novo methylation and maintenance methylation activities are encoded
by a single gene or two different genes.  To address these questions, we have generated a null
mutation of the only known mammalian DNA methyltransferase gene through homologous
recombination in mouse embryonic stem cells and found that the development of the homozygous
embryos is arrested prior to the 8-somite stage.  Surprisingly, the null mutant embryonic stem
cells are viable and contain low but stable levels of methyl cytosine and methyltransferase
activity, suggesting the existence of a second DNA methyltransferase in mammalian cells.
Further studies indicate that de novo methylation activity is not impaired by the mutation as
integrated provirus DNA in MoMuLV-infected homozygous embryonic stem cells become methylated
at a similar rate as in wild-type cells.  Differentiation of mutant cells results in further
reduction of methyl cytosine levels, consistent with the de novo methylation activity being
down regulated in differentiated cells.  These results provide the first evidence that an
independently encoded DNA methyltransferase is present in mammalian cells which is capable of
de novo methylating cellular and viral DNA in vivo.

<>

<1>Lei, X.Y., Ou, T., Zhu, R.L., Zhang, Q.Y.
<2>Sequencing and analysis of the complete genome of Rana grylio virus (RGV).
<3>Arch. Virol.
<4>157
<5>1559-1564
<6>2012
<7>Infection with Rana grylio virus (RGV), an iridovirus isolated in China in 1995,
resulted in a high mortality rate in frogs. The complete genome sequence of RGV
was determined and analyzed. The genomic DNA was 105,791 bp long, with 106 open
reading frames (ORFs). Dot plot analysis showed that the gene order of RGV shared
colinearity with three completely sequenced ranaviruses. A phylogenetic tree was
constructed based on concatenated sequences of iridovirus 26 core-gene-encoded
proteins, and the result showed high bootstrap support for RGV being a member of
the genus Ranavirus and that iridoviruses of other genera also clustered closely.
A microRNA (miRNA) prediction revealed that RGV could encode 18 mature miRNAs,
many of which were located near genes associated with virus replication.
Thirty-three repeated sequences were found in the RGV genome. These results
provide insight into the genetic nature of RGV and are useful for laboratory
diagnosis for vertebrate iridoviruses.

<>

<1>Leismann, O., Roth, M., Friedrich, T., Wende, W., Jeltsch, A.
<2>The Flavobacterium okeanokoites adenine-N6-specific DNA-methyltransferase M.FokI is a tandem enzyme of two independent domains with very different kinetic properties.
<3>Eur. J. Biochem.
<4>251
<5>899-906
<6>1998
<7>The Flavobacterium okeanokoites adenine-N6-specific DNA-methyltransferase, M.FokI, modifies
both adenine residues within its asymmetric recognition sequence 5'-GGATG/CATCC-3'.  It is a
fusion protein comprising two independent enzymes.  We have cloned, overexpressed and purified
full-length M.FokI as well as both individual domains and analyzed their kinetics of DNA
methylation using unmethylated and hemimethylated oligodeoxynucleotide substrates.  Our data
show that both domains of M.FokI methylate DNA independently of each other but cooperate in
DNA binding.  In agreement to former studies, the N-terminal domain of M.FokI modifies the
upper strand of the recognition sequence (GGATG).  It strongly prefers hemimethylated
(5'-GGATG/CmATCC-3'; mA=N6-methyladenosine) over unmethylated substrates.  In contrast, the
terminal domain prefers unmethylated DNA substrates.  Surprisingly, in addition to methylating
the lower strand of the recognition sequence (CATCC), M.FokI-(355-647) also modifies the upper
strand (GGATG), albeit with a lower activity.  In addition methylation was detected at CACCC
sites, but not at sites in which a central AT dinucleotide is flanked by at least one A.T or
T.A base pair.  These results suggests that M.FokI-(335-647) either has a very degenerate
specificity for (G/C)AT(G/C) and sites similar to CATCC or GGATG or, alternatively, that it
has a dual specificity for CATCC and GGATG.

<>

<1>Lemieux, C., Lee, R.W.
<2>Nonreciprocal recombination between alleles of the chloroplast 23S rRNA gene in interspecific Chlamydomonas crosses.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>84
<5>4166-4170
<6>1987
<7>The inheritance of six polymorphic loci mapping in the rRNA-encoding (rDNA) region of the
inverted repeat sequence of chloroplast DNA (cpDNA) was scored in hybrid subclones derived
from reciprocal interspecific crosses between the green algae Chlamydomonas eugametos and
Chlamydomonas moewusii. In order to enhance the detection of cells that had undergone
recombination between parental cpDNAs, hybrids were selected that inherited a chloroplast
antiboiotic-resistance marker contributed by the mating-type-minus (mt-) parent, the parent
that normally contributes fewer cpDNA molecules. The major findings of this study can be
summarized as follows. (i) The majority of the hybrids (14/17) were recombinant for cpDNA
markers in the 10-kilobase-pair rDNA region under study. (ii) Only one allele of each
polymorphic cpDNA locus was ever detected in the hybrids, thus suggesting that newly
recombined rDNA sequences in one copy of the inverted repeat are rapidly spread to the other
by a copy-correction mechanism. (iii) Chloroplast streptomycin-resistance (sr-2) and
erythromycin-resistance (er-nM1) loci, although showing litle or no genetic linkage, were
mapped to the 16S and 23S rRNA gene regions of the cpDNA, respectively, by virtue of their
perfect coinheritance with polymorphic markers within these genes. (iv) cpDNA markers
associated with a putative intron of the C. eugametos 23S rRNA gene were inherited by all 17
hybrids. Such a result is similar to that observed for certain alleles of the large rRNA gene
of yeast mitochrondria in crosses between Omega+ and Omega- strains.

<>

<1>Lemriss, H., Lemriss, S., Butin, M., Ibrahimi, A., El Kabbaj, S., Rasigade, J., Laurent, F.
<2>Non-contiguous finished genome sequence of Staphylococcus capitis CR01 (pulsetype NRCS-A).
<3>Standards in Genomic Sciences
<4>9
<5>1118-1127
<6>2014
<7>Staphylococcus capitis is a coagulase-negative staphylococcus (CoNS) commonly found in the
human microflora. Recently, a clonal population of Staphylococcus
capitis (denominated NRCS-A) was found to be a major cause of late-onset sepsis
(LOS) in several neonatal intensive care units in France. Here, we report the
complete genome sequence and annotation of the prototype Staphylococcus capitis
NCRS-A strain CR01. The 2,504,472 bp long genome (1 chromosome and no plasmids)
exhibits a G+C content of 32.81%, and contains 2,468 protein-coding and 59 tRNA
genes and 4 rRNA genes.

<>

<1>Lenneman, E.M., Barney, B.M.
<2>Draft Genome Sequences of the Alga-Degrading Bacteria Aeromonas hydrophila Strain AD9 and Pseudomonas pseudoalcaligenes Strain AD6.
<3>Genome Announcements
<4>2
<5>e00709-14
<6>2014
<7>Aeromonas hydrophila AD9 and Pseudomonas pseudoalcaligenes AD6 have been linked to algal cell
degradation. Here we report the draft genomes of A. hydrophila AD9
and P. pseudoalcaligenes AD6 for the investigation of causative agents for algal
cell degradation.

<>

<1>Lenz, T., Bonnist, E.Y.M., Pljevaljcic, G., Neely, R.K., Dryden, D.T.F., Scheidig, A.J., Jones, A.C., Weinhold, E.
<2>2-Aminopurine Flipped into the Active Site of the Adenine-Specific DNA Methyltransferase M.TaqI: Crystal Structures and Time-Resolved Fluorescence.
<3>J. Am. Chem. Soc.
<4>129
<5>6240-6248
<6>2007
<7>We report the crystal structure of the DNA adenine-N6 methyltransferase, M.TaqI, complexed
with DNA, showing the fluorescent adenine analog, 2-aminopurine, flipped out of the DNA helix
and occupying virtually the same position in the active site as the natural target adenine.
Time-resolved fluorescence spectroscopy of the crystalline complex faithfully reports this
state: base flipping is accompanied by the loss of the very short ( approximately 50 ps)
lifetime component associated with fully base-stacked 2-aminopurine in DNA, and 2-aminopurine
is subject to considerable quenching by pi-stacking interactions with Tyr108 in the catalytic
motif IV (NPPY). This proves 2-aminopurine to be an excellent probe for studying base flipping
by M.TaqI and suggests similar quenching in the active sites of DNA and RNA adenine-N6 as well
as DNA cytosine-N4 methyltransferases sharing the conserved motif IV. In solution, the same
distinctive fluorescence response confirms complete destacking from DNA and is also observed
when the proposed key residue for base flipping by M.TaqI, the target base partner thymine, is
substituted by an abasic site analog. The corresponding cocrystal structure shows
2-aminopurine in the active site of M.TaqI, demonstrating that the partner thymine is not
essential for base flipping. However, in this structure, a shift of the 3' neighbor of the
target base into the vacancy left after base flipping is observed, apparently replicating a
stabilizing role of the missing partner thymine. Time-resolved fluorescence and acrylamide
quenching measurements of M.TaqI complexes in solution provide evidence for an alternative
binding site for the extra-helical target base within M.TaqI and suggest that the partner
thymine assists in delivering the target base into the active site.

<>

<1>Leon, M.J., Ghai, R., Fernandez, A.B., Sanchez-Porro, C., Rodriguez-Valera, F., Ventosa, A.
<2>Draft Genome of Spiribacter salinus M19-40, an Abundant Gammaproteobacterium in Aquatic Hypersaline Environments.
<3>Genome Announcements
<4>1
<5>e00179-12
<6>2013
<7>We have previously used a de novo metagenomic assembly approach to describe the presence of an
abundant gammaproteobacterium comprising nearly 15% of the
microbial community in an intermediate salinity solar saltern pond. We have
obtained this microbe in pure culture and describe the genome sequencing of the
halophilic photoheterotrophic microbe, Spiribacter salinus M19-40.

<>

<1>Leonard, M.T., Davis-Richardson, A.G., Ardissone, A.N., Kemppainen, K.M., Drew, J.C., Ilonen, J., Knip, M., Simell, O., Toppari, J., Veijola, R., Hyoty, H., Triplett, E.W.
<2>The methylome of the gut microbiome: disparate Dam methylation patterns in intestinal Bacteroides dorei.
<3>Front. Microbiol.
<4>5
<5>361
<6>2014
<7>Despite the large interest in the human microbiome in recent years, there are no reports of
bacterial DNA methylation in the microbiome.  Here metagenomic sequencing using the Pacific
Biosciences platform allowed for rapid identification of bacterial GATC methylation status of
a bacterial species in human stool samples.  For this work, two stool samples were chosen that
were dominated by a single species, Bacteroides dorei.  Based on 16S rRNA analysis, this
species represented over 45% of the bacteria present in these two samples.  The B. dorei
genome sequence from these samples was determined and the GATC methylation sites mapped.  The
Bacteroides dorei genome from one subject lacked any GATC methylation and lacked the DNA
adenine methyltransferase genes.  In contrast, B. dorei from another subject contained 20,551
methylated GATC sites.  Of the 4970 open reading frames identified in the GATC methylated B.
dorei genome, 3184 genes were methylated as well as 1735 GATC methylations in intergenic
regions.  These results suggest that DNA methylation patterns are important to consider in
multi-omic analyses of microbiome samples seeking to discover the diversity of bacterial
functions and may differ between disease states.

<>

<1>Leonard, M.T., Fagen, J.R., Davis-Richardson, A.G., Davis, M.J., Triplett, E.W.
<2>Complete genome sequence of Liberibacter crescens BT-1.
<3>Standards in Genomic Sciences
<4>7
<5>0
<6>2012
<7>
<>

<1>Leonard, M.T., Panayotova, N., Farmerie, W.G., Triplett, E.W., Nicholson, W.L.
<2>Complete Genome Sequence of Carnobacterium gilichinskyi Strain WN1359T (DSM 27470T).
<3>Genome Announcements
<4>1
<5>e00985-13
<6>2013
<7>We report the complete genome sequence of Carnobacterium gilichinskyi strain WN1359,
previously isolated from Siberian permafrost and capable of growth under
cold (0 degrees C), anoxic, CO2-dominated, low-pressure (0.7-kPa) conditions in a
simulation of the Mars atmosphere.

<>

<1>Leonard, M.T., Valladares, R.B., Ardissone, A., Gonzalez, C.F., Lorca, G.L., Triplett, E.W.
<2>Complete Genome Sequences of Lactobacillus johnsonii Strain N6.2 and Lactobacillus reuteri Strain TD1.
<3>Genome Announcements
<4>2
<5>e00397-14
<6>2014
<7>We report here the complete genome sequences of Lactobacillus johnsonii strain N6.2, a
homofermentative lactic acid intestinal bacterium, and Lactobacillus
reuteri strain TD1, a heterofermentative lactic acid intestinal bacterium, both
isolated from a type 1 diabetes-resistant rat model.

<>

<1>Leonard, S.R., Lacher, D.W., Elkins, C.A., Jung, C.M.
<2>Draft Genome Sequence of the Multidrug-Resistant Escherichia coli Strain LR09, Isolated from a Wastewater Treatment Plant.
<3>Genome Announcements
<4>2
<5>e00272-14
<6>2014
<7>We report the draft genome sequence of Escherichia coli O1:H6 strain LR09, which  was isolated
from a wastewater treatment plant and displays high resistance to five fluoroquinolone
antimicrobials. The assembled data determine that the strain clusters with E. coli phylogroup
F and harbors a plasmid conferring resistance to a broad spectrum of antibiotics.

<>

<1>Leonard, W.J., Wolf, J.B., Halden, N.F.
<2>Novel restriction endonuclease.
<3>US Patent Office
<4>US 4935367
<5>
<6>1990
<7>A new restriction enzyme, MfeI, has been discovered. MfeI recognizes the sequence CAATTG and
cuts at the recognition sequence C^AATTG and generates compatible cohesive ends with EcoRI
cleaved fragments. Various utilities of the enzyme have been described.

<>

<1>Leonhardt, H., Page, A.W., Weier, H.U., Bestor, T.H.
<2>A targeting sequence directs DNA methyltransferase to sites of DNA replication in mammalian nuclei.
<3>Cell
<4>71
<5>865-873
<6>1992
<7>Tissue-specific patterns of methylated deoxycytidine residues in the mammalian genome are
preserved by postreplicative methylation of newly synthesized DNA. DNA methyltransferase
(MTase) is here shown to associate with replication foci during S phase but to display a
difuse nucleoplasmic distribution in non-S phase cells. Analysis of DNA MTase-B-galactosidase
fusion proteins has shown that association with replication foci is mediated by a novel
targeting sequence located near the N-terminus of DNA MTase. This sequence has the properties
expected of a targeting sequence in that it is not required for enzymatic activity, prevents
proper targeting when deleted and, when fused to B-galactosidase, causes the fusion protein to
associate with replication foci in a cell cycle-dependent manner.

<>

<1>Leopold, S.R., Magrini, V., Holt, N.J., Shaikh, N., Mardis, E.R., Cagno, J., Ogura, Y., Iguchi, A., Hayashi, T., Mellmann, A., Karch, H., Besser, T.E., Sawyer, S.A., Whittam, T.S., Tarr, P.I.
<2>A precise reconstruction of the emergence and constrained radiations of Escherichia coli O157 portrayed by backbone concatenomic analysis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>8713-8718
<6>2009
<7>Single nucleotide polymorphisms (SNPs) in stable genome regions provide
durable measurements of species evolution. We systematically identified
each SNP in concatenations of all backbone ORFs in 7 newly or previously
sequenced evolutionarily instructive pathogenic Escherichia coli O157:H7,
O157:H(-), and O55:H7. The 1,113 synonymous SNPs demonstrate emergence of
the largest cluster of this pathogen only in the last millennium.
Unexpectedly, shared SNPs within circumscribed clusters of organisms
suggest severely restricted survival and limited effective population
sizes of pathogenic O157:H7, tenuous survival of these organisms in
nature, source-sink evolutionary dynamics, or, possibly, a limited number
of mutations that confer selective advantage. A single large segment
spanning the rfb-gnd gene cluster is the only backbone region convincingly
acquired by recombination as O157 emerged from O55. This concatenomic
analysis also supports using SNPs to differentiate closely related
pathogens for infection control and forensic purposes. However,
constrained radiations raise the possibility of making false associations
between isolates.

<>

<1>Lepikhov, K., Tchernov, A., Zheleznaja, L., Matvienko, N., Walter, J., Trautner, T.A.
<2>Characterization of the type IV restriction modification system BspLU11III from Bacillus sp. LU11.
<3>Nucleic Acids Res.
<4>29
<5>4691-4698
<6>2001
<7>We report the characterization and cloning of the genes for an unusual type IV
restriction-modification system, BspLU11III, from Bacillus sp. LU11. The system consists of
two methyltransferases and one endonuclease, which also possesses methyltransferase activity.
The three genes of the restriction-modification system, bsplu11IIIMa, bsplu11IIIMb and
bsplu11IIIR, are closely linked and tandemly arranged. The corresponding enzymes recognize the
dsDNA sequence 5'-GGGAC-3'/5'-GTCCC-3', with M.BspLU11IIIa modifying the A (underlined) of
one strand and M.BspLU11IIIb the inner C (underlined) of the other strand. R.BspLU11III has
both endonuclease and adenine-specific methyltransferase activities and is able to protect the
DNA against cleavage by itself. In contrast to all type IV restriction-modification systems
described so far, which have only one adenine-specific methyltransferase, BspLU11III is the
first type IV restriction-modification system that includes two methyltransferases, one of
them being cytosine specific.

<>

<1>Lepp, D., Roxas, B., Parreira, V.R., Marri, P.R., Rosey, E.L., Gong, J., Songer, J.G., Vedantam, G., Prescott, J.F.
<2>Identification of novel pathogenicity loci in Clostridium perfringens strains that cause avian necrotic enteritis.
<3>PLoS ONE
<4>5
<5>E10795
<6>2010
<7>Type A Clostridium perfringens causes poultry necrotic enteritis (NE), an
enteric disease of considerable economic importance, yet can also exist as
a member of the normal intestinal microbiota. A recently discovered
pore-forming toxin, NetB, is associated with pathogenesis in most, but not
all, NE isolates. This finding suggested that NE-causing strains may
possess other virulence gene(s) not present in commensal type A isolates.
We used high-throughput sequencing (HTS) technologies to generate draft
genome sequences of seven unrelated C. perfringens poultry NE isolates and
one isolate from a healthy bird, and identified additional novel
NE-associated genes by comparison with nine publicly available reference
genomes. Thirty-one open reading frames (ORFs) were unique to all NE
strains and formed the basis for three highly conserved NE-associated loci
that we designated NELoc-1 (42 kb), NELoc-2 (11.2 kb) and NELoc-3 (5.6
kb). The largest locus, NELoc-1, consisted of netB and 36 additional
genes, including those predicted to encode two leukocidins, an
internalin-like protein and a ricin-domain protein. Pulsed-field gel
electrophoresis (PFGE) and Southern blotting revealed that the NE strains
each carried 2 to 5 large plasmids, and that NELoc-1 and -3 were localized
on distinct plasmids of sizes approximately 85 and approximately 70 kb,
respectively. Sequencing of the regions flanking these loci revealed
similarity to previously characterized conjugative plasmids of C.
perfringens. These results provide significant insight into the
pathogenetic basis of poultry NE and are the first to demonstrate that
netB resides in a large, plasmid-encoded locus. Our findings strongly
suggest that poultry NE is caused by several novel virulence factors,
whose genes are clustered on discrete pathogenicity loci, some of which
are plasmid-borne.

<>

<1>Lesaulnier, C., Papamichail, D., McCorkle, S., Ollivier, B., Skiena, S., Taghavi, S., Zak, D., van der Lelie, D.
<2>Elevated atmospheric CO2 affects soil microbial diversity associated with trembling aspen.
<3>Environ. Microbiol.
<4>10
<5>926-941
<6>2008
<7>The effects of elevated atmospheric CO(2) (560 p.p.m.) and subsequent
plant responses on the soil microbial community composition associated
with trembling aspen was assessed through the classification of 6996
complete ribosomal DNA sequences amplified from the Rhinelander WI
free-air CO(2) and O(3) enrichment (FACE) experiments microbial community
metagenome. This in-depth comparative analysis provides an unprecedented,
detailed and deep branching profile of population changes incurred as a
response to this environmental perturbation. Total bacterial and
eukaryotic abundance does not change; however, an increase in
heterotrophic decomposers and ectomycorrhizal fungi is observed. Nitrate
reducers of the domain bacteria and archaea, of the phylum Crenarchaea,
potentially implicated in ammonium oxidation, significantly decreased with
elevated CO(2). These changes in soil biota are evidence for altered
interactions between trembling aspen and the microorganisms in its
surrounding soil, and support the theory that greater plant detritus
production under elevated CO(2) significantly alters soil microbial
community composition.

<>

<1>Lescot, M., Audic, S., Robert, C., Nguyen, T.T., Blanc, G., Cutler, S.J., Wincker, P., Couloux, A., Claverie, J.M., Raoult, D., Drancourt, M.
<2>The genome of Borrelia recurrentis, the agent of deadly louse-borne relapsing fever, is a degraded subset of tick-borne Borrelia duttonii.
<3>PLoS Genet.
<4>4
<5>e1000185
<6>2008
<7>In an effort to understand how a tick-borne pathogen adapts to the body louse, we sequenced
and compared the genomes of the recurrent fever agents
Borrelia recurrentis and B. duttonii. The 1,242,163-1,574,910-bp
fragmented genomes of B. recurrentis and B. duttonii contain a unique
23-kb linear plasmid. This linear plasmid exhibits a large polyT track
within the promoter region of an intact variable large protein gene and a
telomere resolvase that is unique to Borrelia. The genome content is
characterized by several repeat families, including antigenic
lipoproteins. B. recurrentis exhibited a 20.4% genome size reduction and
appeared to be a strain of B. duttonii, with a decaying genome, possibly
due to the accumulation of genomic errors induced by the loss of recA and
mutS. Accompanying this were increases in the number of impaired genes and
a reduction in coding capacity, including surface-exposed lipoproteins and
putative virulence factors. Analysis of the reconstructed ancestral
sequence compared to B. duttonii and B. recurrentis was consistent with
the accelerated evolution observed in B. recurrentis. Vector
specialization of louse-borne pathogens responsible for major epidemics
was associated with rapid genome reduction. The correlation between gene
loss and increased virulence of B. recurrentis parallels that of
Rickettsia prowazekii, with both species being genomic subsets of
less-virulent strains.

<>

<1>Lesiak, J.M., Liebl, W., Ehrenreich, A.
<2>Development of an in vivo methylation system for the solventogen Clostridium saccharobutylicum NCP 262 and analysis of two endonuclease mutants.
<3>J. Biotechnol.
<4>188
<5>97-99
<6>2014
<7>The genome of the biotechnologically important solventogenic Clostridium saccharobutylicum NCP
262 contains two operons coding for genes of presumed type I RM systems belonging to the
families A and C. They represent a limiting factor for the development of transformation and
conjugation protocols. We established an efficient triparental mating system to transfer DNA
to C. saccharobutylicum by conjugation, which includes an in vivo methylation of the donor
DNA. Furthermore we describe increased rates of conjugation in knock-out mutants of the
restrictase subunits of both RM systems.

<>

<1>Lesnik, E.A., Beschetnikova, Z.A., Maslova, R.N., Varshavsky, J.M.
<2>The inhibition of restriction endonucleases due to Z-DNA in negatively supercoiled plasmids.
<3>FEBS Lett.
<4>280
<5>91-93
<6>1991
<7>Plasmid pGC20 containing the (dGC) insert in the SmaI recognition site has been
used to study the inhibition of cleavage by different restriction endonucleases
due to Z-DNA formation in the (dCG) sequence of the negatively supercoiled
plasmid.  Data obtained indicate the different sensitivity of restriction
endonucleases to DNA conformational perturbations resulted from Z-DNA
formation.  Therefore, the inhibition of DNA cleavage by a particular
restriction endonuclease cannot serve as a criterion for the estimation of the
length of B-Z junctions in circular supercoiled DNAs.

<>

<1>Lesnik, E.A., Beschetnikova, Z.A., Maslova, R.N., Varshavsky, J.M.
<2>The influence of Z-DNA formation on the activity of restriction endonucleases.
<3>J. Biomol. Struct. Dyn.
<4>8
<5>A118
<6>1991
<7>Plasmid pGC20 containing the (dGC)9 insert in SmaI recognition site has been
used to study the inhibition of cleavage by six different restriction
endonucleases due to Z-DNA formation in negatively supercoiled plasmid.  The
recognition sites of the KpnI, SacI and EcoRI were located correspondingly at
0, 6 and 12 b.p. away from Z-DNA.  The recognition sites of the BamHI, XbaI and
SalI were 1, 7 and 13 b.p. away from Z-DNA on the other side of it.  Formation
of Z-DNA in the (CG)10 sequence inhibited cleavage by restriction endonucleases
to different extents.  In the KpnI, SacI and EcoRI series the degree of enzyme
inhibition decreased as the distance between their recognition sites and the
insert increased.  Such a correlation was not observed in the BamHI, XbaI and
SalI series.  All three endonucleases were rather strongly inhibited.  These
data indicate on the different sensitivity of restriction endonucleases to DNA
conformational perturbations resulted from Z-DNA formation in the negatively
supercoiled plasmid.  Therefore, the inhibition of DNA cleavage by a particular
restriction endonuclease cannot serve as a criterion for the estimation of the
length of B-Z junctions in circular supercoiled DNAs.

<>

<1>Lesnik, E.A., Khalimullina, Z.A.
<2>The influence of Z-conformation of the enzyme activity of restriction endonucleases.
<3>Mol. Biol. (Mosk)
<4>23
<5>1638-1644
<6>1989
<7>Recombinant plasmid pGC20 containing (GC)9-insert into the SmaI site of pUC19
has been used to study the inhibition of cleavage by six restriction
endonucleases; KpnI, SacI, EcoRI and also BamHI, XbaI and SalI, due to Z-DNA
formation in negatively supercoiled plasmids.  The recognition sites of these
enzymes were located at different distances on both side of the (CG)
10-sequence.  It was shown that the inhibition of cleavage by KpnI, SacI and
EcoRI decreased in this series as fast as the distance between recognition site
and B-Z junction was increased, and no inhibition of cleavage by EcoRI was
found.  However, such a correlation was not found in the series of BamHI, XbaI
and SalI.  In contrast with EcoRI the cleavage by SalI was inhibited
completely.  These results indicate the difference in sensitivity of
restriction endonucleases to the structural perturbations of DNA associated
with B-Z junctions.  It seems to depend on features of the enzyme-substrate
interaction mechanisms and also on recognition and flanking sequences of DNA.
Consequently, experiments with the inhibition of the cleavage by any enzyme
cannot help to determine the dimension of the region of DNA with altered
structure.

<>

<1>Lesser, D.R., Grajkowski, A., Kurpiewski, M.R., Koziolkiewicz, M., Stec, W.J., Jacobson, L.J.
<2>Steroselective interaction with chiral phosphorothioates at the central DNA kink of the EcoRI endonuclease-GAATTC Complex.
<3>J. Biol. Chem.
<4>267
<5>24810-24818
<6>1992
<7>We have probed the contacts between EcoRI endonuclease and the central phosphate of its
recognition site GAApTTC, using synthetic oligonucleotides containing single sterospecific Rp-
or Sp-phosphorothioates (ps). These substitutions produce subtle sterospecific effects on
EcoRI endonuclease binding and cleavage. An Sp-Ps substitution in one strand of the DNA duplex
improves binding free energy by -1.5 kcal/mol, whereas the Rp-Ps substitution has an
unfavorable effect (+0.3 kcal/mol) on binding free energy. These effects derive principally
from changes in the first order rate constants for dissociation of the enzyme-DNA commplexes.
The first order rate constants for strand scission are also affected, in that a strand
containing Sp-Ps substitution is cleaved 2 to 3 times more rapidly than a strand containing a
normal prochiral phosphate, whereas a strand containing Rp-Ps substitution is cleaved about 3
times slower than normal. As a result, single-strand substitutions produce pronounced
asymmetry in the rates of cleavage of the two DNA strands, and this effect is exaggerated in
an Rp,Sp-heteroduplex. Ethylation-interference foot-printing indicates that none of the Ps
substitutions cause any major change in contacts between endonuclease and DNA phosphates. When
an Sp-Ps localizes P=O in the DNA major groove, a hydrogen -bonding interaction with the
backbone amide-NH of Gly116 of the endonuclease is improved relative to that with a prochiral
phosphate having intermediate P-O bond order and delocalized charge.

<>

<1>Lesser, D.R., Kurpiewski, M.R., Jen-Jacobson, L.
<2>The energetic basis of specificity in the EcoRI endonuclease-DNA interaction.
<3>Science
<4>250
<5>776-786
<6>1990
<7>High sequence selectivity in DNA-protein interactions was analyzed by measuring
discrimination by EcoRI endonuclease between the recognition site GAATTC and
systematically altered DNA sites.  Base analogue substitutions that preserve
the sequence-dependent conformational motif of the GAATTC site permit deletion
of single sites of protein-base contact at a cost of +1 to +2 kcal/mol.
However, the introduction of any one incorrect natural base pair costs +6 to
+13 kcal/mol in transition state interaction energy, the resultant of the
following interdependent factors:  deletion of one or two hydrogen bonds
between the protein and a purine base; unfavorable steric apposition between a
group on the protein and an incorrectly placed functional group on a base;
disruption of a pyrimidine contact with the protein; loss of some crucial
interactions between protein and DNA phosphates; and an increased energetic
cost of attaining the required DNA conformation in the transition state
complex.  EcoRI endonuclease thus achieves stringent discrimination by both
direct readout (protein-base contacts) and indirect readout (protein-phosphate
contacts and DNA conformation) of the DNA sequence.

<>

<1>Lesser, D.R., Kurpiewski, M.R., Waters, T., Connolly, B.A., Jen-Jacobson, L.
<2>Facilitated distortion of the DNA site enhances EcoRI endonuclease-DNA recognition.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>7548-7552
<6>1993
<7>We have measured the binding of EcoRI endonuclease to a complete set of purine-base analogue
sites, each of which deletes one functional group that forms a hydrogen bond with the
endonuclease in the canonical GAATTC complex. For five of six functional group deletions, the
observed penalty in binding free energy is +1.3 to +1.7 kcal/mol. For two of these cases
(replacement of adenine N7 with carbon) a single protein-base hydrogen bond is removed without
deleting an interstrand Watson-Crick hydrogen bond or causing structural adaptation in the
complex. This observation establishes that the incremental energetic contribution of one
protein-base hydrogen bond is about -1.5 kcal.mol. By contrast, deletion of the N6-amino group
of the inner adenine in the site improves binding by -1.0 kcal/mol because the penalty for
deleting a protein-base hydrogen bond is outweighed by facilitation of the required DNA
distortion (kinking) in the complex. This result provides direct evidence that the energetic
cost of distorting a DNA site can make an unfavorable contribution to protein-DNA binding.

<>

<1>Letek, M. et al.
<2>The genome of a pathogenic rhodococcus: cooptive virulence underpinned by key gene acquisitions.
<3>PLoS Genet.
<4>6
<5>e1001145
<6>2010
<7>We report the genome of the facultative intracellular parasite Rhodococcus equi, the only
animal pathogen within the biotechnologically important actinobacterial genus Rhodococcus. The
5.0-Mb R. equi 103S genome is significantly smaller than those of environmental rhodococci.
This is due to genome expansion in nonpathogenic species, via a linear gain of paralogous
genes and an accelerated genetic flux, rather than reductive evolution in R. equi. The 103S
genome lacks the extensive catabolic and secondary metabolic complement of environmental
rhodococci, and it displays unique adaptations for host colonization and competition in the
short-chain fatty acid-rich intestine and manure of herbivores--two main R. equi reservoirs.
Except for a few horizontally acquired (HGT) pathogenicity loci, including a cytoadhesive
pilus determinant (rpl) and the virulence plasmid vap pathogenicity island (PAI) required for
intramacrophage survival, most of the potential virulence-associated genes identified in R.
equi are conserved in environmental rhodococci or have homologs in nonpathogenic
Actinobacteria. This suggests a mechanism of virulence evolution based on the cooption of
existing core actinobacterial traits, triggered by key host niche-adaptive HGT events. We
tested this hypothesis by investigating R. equi virulence plasmid-chromosome crosstalk, by
global transcription profiling and expression network analysis. Two chromosomal genes
conserved in environmental rhodococci, encoding putative chorismate mutase and anthranilate
synthase enzymes involved in aromatic amino acid biosynthesis, were strongly coregulated with
vap PAI virulence genes and required for optimal proliferation in macrophages. The regulatory
integration of chromosomal metabolic genes under the control of the HGT-acquired plasmid PAI
is thus an important element in the cooptive virulence of R. equi.

<>

<1>Leung, D.W., Lui, A.C.P., Merilees, H., McBride, B.C., Smith, M.
<2>A restriction enzyme from Fusobacterium nucleatum 4H which recognizes GCNGC.
<3>Nucleic Acids Res.
<4>6
<5>17-25
<6>1979
<7>A site-specific restriction endonuclease Fnu4HI isolated from Fusobacterium
nucleatum 4H recognizes the DNA nucleotide sequence 5'-G C^N G C-3' 3'-C G N^C
G-5' and cleaves as indicated by the arrows.

<>

<1>Leung, S.M., Chan, K.Y., Suen, Y.K., Shaw, P.C.
<2>Screening and characterization of restriction endonucleases from a bacterial culture collection in Hong Kong.
<3>Nucleic Acids Res.
<4>17
<5>10133
<6>1989
<7>None

<>

<1>Leung, S.M., Kam, K.M., Chan, K.Y., Shaw, P.C.
<2>Purification and characterization of restriction endonuclease BcoI from a soil isolate of Bacillus coagulans.
<3>FEMS Microbiol. Lett.
<4>66
<5>153-156
<6>1990
<7>A soil isolate identified as Bacillus coagulans is found to produce BcoI, an
isoschizomer of AvaI and with the same cleavage site.  This thermostable
enzyme, BcoI, is produced at high level and can be isolated by passing the
crude bacterial lysate through a DEAE-cellulose column.

<>

<1>Leveau, J.H., Gerards, S., de Boer, W., van Veen, J.A.
<2>Phylogeny-function analysis of (meta)genomic libraries: screening for expression of ribosomal RNA genes by large-insert library fluorescent in situ hybridization (LIL-FISH).
<3>Environ. Microbiol.
<4>6
<5>990-998
<6>2004
<7>We assessed the utility of fluorescent in situ hybridization (FISH) in the
screening of clone libraries of (meta)genomic or environmental DNA for the
presence and expression of bacterial ribosomal RNA (rRNA) genes. To
establish proof-of-principle, we constructed a fosmid-based library in
Escherichia coli of large-sized genomic DNA fragments of the mycophagous
soil bacterium Collimonas fungivorans, and hybridized 768 library clones
with the Collimonas-specific fluorescent probe CTE998-1015. Critical to
the success of this approach (which we refer to as large-insert library
FISH or LIL-FISH) was the ability to induce fosmid copy number, the
exponential growth status of library clones in the FISH assay and the use
of a simple pooling strategy to reduce the number of hybridizations.
Twelve out of 768 E. coli clones were suspected to harbour and express
Collimonas 16S rRNA genes based on their hybridization to CTE998-1015.
This was confirmed by the finding that all 12 clones were also identified
in an independent polymerase chain reaction-based screening of the same
768 clones using a primer set for the specific detection of Collimonas 16S
ribosomal DNA (rDNA). Fosmids isolated from these clones were grouped by
restriction analysis into two distinct contigs, confirming that C.
fungivorans harbours at least two 16S rRNA genes. For one contig,
representing 1-2% of the genome, the nucleotide sequence was determined,
providing us with a narrow but informative view of Collimonas genome
structure and content.

<>

<1>Levin, B.R.
<2>Restriction-modification immunity and the maintenance of genetic diversity in bacterial populations.
<3>Evolutionary Processes and Theory, Academic Press, S. Karlin, E. Nero, New York
<4>0
<5>669-688
<6>1986
<7>A simple model is presented of the population dynamics of phage and bacteria
with restriction-modification immunity.  The analysis of this model suggests
that: i) in phage-limited communities of bacteria, there are broad conditions
under which this type of immune system will maintain clonal diversity via
frequency-dependent selection that favors rare restriction-modification types;
and ii) when phage-resistant bacteria are present and the community is limited
by resources, the conditions for this mechanism to maintain clonal diversity
are much narrower.  Reviewing that which is known about the genetic structure
of natural populations of E. coli and the population biology of bacteriophage,
the proposition is evaluated that this type of phage-mediated,
frequency-dependent selection is responsible for maintaining the considerable
chromosomal gene and plasmid diversity found in this bacterial species.
Finally, the analogy is considered between this type of phage-mediated
selection on restriction-modification diversity and that postulated earlier for
parasite-mediated, frequency-dependent selection maintaining antigenic
diversity in vertebrates.

<>

<1>Levine, J.D., Cech, C.L.
<2>Low frequency restriction enzymes in pulsed field electrophoresis.
<3>Biotechnology
<4>7
<5>1033-1036
<6>1989
<7>None

<>

<1>Levine, S.M., Lin, E.A., Emara, W., Kang, J., DiBenedetto, M., Ando, T., Falush, D., Blaser, M.J.
<2>Plastic cells and populations: DNA substrate characteristics in Helicobacter pylori transformation define a flexible but conservative system for genomic variation.
<3>FASEB J.
<4>21
<5>3458-3467
<6>2007
<7>Helicobacter pylori, bacteria that colonize the human gastric mucosa, are naturally competent
for transformation by exogenous DNA, and show a
panmictic population structure. To understand the mechanisms involved
in its horizontal gene transfer, we sought to define the interval
required from exposure to substrate DNA until DNA uptake and expression
of a selectable phenotype, as well as the relationship of transforming
fragment length, concentration, homology, symmetry, and strandedness,
to the transformation frequency. We provide evidence that natural
transformation in H. pylori differs in efficiency among wild-type
strains but is saturable and varies with substrate DNA length,
symmetry, strandedness, and species origin. We show that H. pylori
cells can be transformed within one minute of contact with DNA, by DNA
fragments as small as 50 bp, and as few as 5 bp on one flank of a
selectable single nucleotide mutation is sufficient substrate for
recombination of a transforming fragment, and that double-stranded DNA
is the preferred (1000-fold > single-stranded) substrate. The high
efficiency of double-stranded DNA as transformation substrate, in
conjunction with strain-specific restriction endonucleases suggests a
model of short-fragment recombination favoring closest relatives,
consistent with the observed H. pylori population biology.

<>

<1>Levitz, R., Chapman, D., Amitsur, M., Green, R., Snyder, L., Kaufmann, G.
<2>The optional E. coli prr locus encodes a latent form of phage T4-induced anticodon nuclease.
<3>EMBO J.
<4>9
<5>1383-1389
<6>1990
<7>The optional Escherichia coli prr locus restricts phage T4 mutants lacking polynucleotide
kinase or RNA ligase.  Underlying this restriction is the specific manifestation of the
T4-induced anticodon nuclease, an enzyme which triggers the cleavage-ligation of the host
tRNALys.  We report here the molecular cloning, nucleotide sequence and mutational analysis of
prr-associated DNA.  The results indicate that prr encodes a latent form of anticodon nuclease
consisting of a core enzyme and cognate masking agents.  They suggest that the T4-encoded
factors of anticodon nuclease counteract the prr-encoded masking agents, thus activating the
latent enzyme.  The encoding of a tRNA cleavage-ligation pathway by two separate genetic
systems which cohabitate E. coli may provide a clue to the evolution of RNA splicing
mechanisms mediated by proteins.

<>

<1>Levy, W.P., Welker, N.E.
<2>Deoxyribonucleic acid modification methylase from Bacillus stearothermophilus.
<3>Biochemistry
<4>20
<5>1120-1127
<6>1981
<7>A modification methylase was isolated from Bacillus stearothermophilus 1503-4R
(Bst153I) and purified to homogeneity.  The enzyme is an acidic protein and
composed of a subunit with a molecular weight of 105000, and only the
tetrameric form was detected in solution.  The methylase exhibited maximal
activity between 54 and 61C and between pH 8.1 and 9.3.  In contrast to
Bst1503I endonuclease [Catterall, J.F., & Welker, N.E. (1977) J. Bacteriol.
129: 1110-1120], the methylase is completely inactivated when exposed to
temperatures near the optimal growth temperature (63-67C).  The methylase was
also inactivated when exposed to temperatures below the minimal growth
temperature (48-53C).  The thermostability of the methylase is significantly
enhanced by Na+, K+, or NH4+.  Membrane-bound methylase is resistant to heat
inactivation at temperatures near the maximum growth temperature (73-75C).  The
methylase functions as a tetramer.  The initial rates of methyl transfer are
first order in methylase concentration, and the enzyme obeys Michaelis-Menten
kinetics with respect to DNA but not to S-adenosyl-L-methionine.

<>

<1>Levy, W.P., Welker, N.E.
<2>Purification and properties of a modification methylase from Bacillus stearothermophilus (Meth M.Bst 1503).
<3>Fed. Proc.
<4>37
<5>1414
<6>1978
<7>A modification methylase has been isolated from B. stearothermophilus 1503-4R
Res+Mod+ (Math M.Bst 1503) and purified 500 fold from sonicated protoplasts
using ammonium sulfate fractionation, Bio Gel A5m gel filtration and
DEAE-cellulose ion exhange column chromatography.  Incurbation of Meth M.Pst
1503 with various DNA substrates in the presence of S-adenosyl methionine
protects these substrates from the action of Endo R.Bst 1503.  Meth M.Bst 1503
has an apparent molecular weight between 400,000-450,000 daltons.  A single
protein staining band with a molecular weight of between 90,000-100,000 daltons
is obtained when the enzyme is analyzed by SDS polyacrylamide gel
electrohoresis.  Meth M.Bst 1503 does not exhibit restriction endonuclease
activity.  The enzyme in 50% glycerol can be stored at -25C for at least 3
weeks with no detectable loss of activity.  Data are presented which indicate
that Endo R.Bst 1503 and Meth M.Bst 1503 do not share a common polypeptide.
Temperature and pH profiles of enzymatic activity were determined.

<>

<1>Lewin, J.
<2>Method for detecting cytosine methylation of DNA using hemi-methylation restriction enzymes for diagnosis of cancer and identification of  undesirable drug effects.
<3>International Patent Office
<4>WO 200545069 A
<5>24
<6>2005
<7>The following invention conerns an enzymatic method for investigating cytosine methylation in
DNA sequences.  The DNA to be investigated is hybridized to oligonucleotides.  The hybrids are
reacted with restriction enzymes, which are able to distinguish hemi-methylated DNA double
strands either from unmethylated or from homogenously methylated DNA double strands.  The
occurrence or non-occurrence of restriction (and thereby methylation status of the cytosine
positions to be investigated) can be determined by various techniques.  The method is
particularly suitable for the diagnosis of cell proliferative disorders (including cancer) and
other diseases associated with a change of the methylation status as well as for the prognosis
of undesired drug effects.

<>

<1>Lewis, L.K., Lobachev, K., Westmoreland, J.W., Karthikeyan, G., Williamson, K.M.J.J.J., Resnick, M.A.
<2>Use of a restriction endonuclease cytotoxicity assay to identify inducible GAL1 promoter variants with reduced basal activity - for use  in fucntional genomics.
<3>Gene
<4>363
<5>183-192
<6>2005
<7>Inducible promoter fusions are commonly employed to study the biological functions of genes as
well as to investigate mechanisms of transcription regulation. A concern for many studies of
heterologous gene expression is that steady state transcription may be too high under
non-inducing conditions, producing undesired phenotypes prior to induction. Fusions containing
the galactose-inducible GAL1 promoter joined to PvuII, a bacterial DNA endonuclease gene, are
toxic to yeast cells even under non-inducing conditions, i.e., in glucose media. This toxicity
was utilized in conjunction with PCR-based mutagenesis of the GAL1 regulatory region to
isolate mutant promoters that retained high inducibility but exhibited reduced basal level
expression. The Mig1 repressor binding and putative TATA box regions were unchanged among four
mutant promoters examined in detail. However, each promoter contained one or more mutations
within previously identified binding sites for the Gal4 activator protein. Genetic assays
developed to monitor GAL1p::I-SceI endonuclease-induced recombination demonstrated that basal
expression from two of the new promoters (designated GAL1-V4 and GAL1-V10) was strongly
reduced. These experiments and additional quantitative luciferase reporter gene assays
demonstrate the utility of the approach for identifying promoters that permit more tightly
controlled gene expression.

<>

<1>Lewis, Z.A., Adhvaryu, K.K., Honda, S., Shiver, A.L., Knip, M., Sack, R., Selker, E.U.
<2>DNA Methylation and Normal Chromosome Behavior in Neurospora Depend on Five Components of a Histone Methyltransferase Complex, DCDC.
<3>PLoS Genet.
<4>6
<5>e1001196
<6>2010
<7>Methylation of DNA and of Lysine 9 on histone H3 (H3K9) is associated with gene silencing in
many animals, plants, and fungi. In Neurospora
crassa, methylation of H3K9 by DIM-5 directs cytosine methylation by
recruiting a complex containing Heterochromatin Protein-1 (HP1) and the
DIM-2 DNA methyltransferase. We report genetic, proteomic, and
biochemical investigations into how DIM-5 is controlled. These studies
revealed DCDC, a previously unknown protein complex including DIM-5,
DIM-7, DIM-9, CUL4, and DDB1. Components of DCDC are required for
H3K9me3, proper chromosome segregation, and DNA methylation.
DCDC-defective strains, but not HP1-defective strains, are
hypersensitive to MMS, revealing an HP1-independent function of H3K9
methylation. In addition to DDB1, DIM-7, and the WD40 domain protein
DIM-9, other presumptive DCAFs (DDB1/CUL4 associated factors)
co-purified with CUL4, suggesting that CUL4/DDB1 forms multiple
complexes with distinct functions. This conclusion was supported by
results of drug sensitivity tests. CUL4, DDB1, and DIM-9 are not
required for localization of DIM-5 to incipient heterochromatin
domains, indicating that recruitment of DIM-5 to chromatin is not
sufficient to direct H3K9me3. DIM-7 is required for DIM-5 localization
and mediates interaction of DIM-5 with DDB1/CUL4 through DIM-9. These
data support a two-step mechanism for H3K9 methylation in Neurospora.

<>

<1>Li, A., Gai, Z., Cui, D., Ma, F., Yang, J., Zhang, X., Sun, Y., Ren, N.
<2>Genome Sequence of a Highly Efficient Aerobic Denitrifying Bacterium, Pseudomonas stutzeri T13.
<3>J. Bacteriol.
<4>194
<5>5720
<6>2012
<7>Pseudomonas stutzeri T13 is a highly efficient aerobic denitrifying bacterium. Information
about the genome of this aerobic denitrifying bacterium has been
limited until now. We present the draft genome of P. stutzeri T13. The results
could provide further insight into the aerobic denitrification mechanism in
strain T13.

<>

<1>Li, A., Geng, J., Cui, D., Shu, C., Zhang, S., Yang, J., Xing, J., Wang, J., Ma, F., Hu, S.
<2>Genome Sequence of Agrobacterium tumefaciens Strain F2, a Bioflocculant-Producing Bacterium.
<3>J. Bacteriol.
<4>193
<5>5531
<6>2011
<7>Agrobacterium tumefaciens F2 is an efficient bioflocculant-producing bacterium. But the genes
related to the metabolic pathway of bioflocculant
biosynthesis in strain F2 are unknown. We present the draft genome of A.
tumefaciens F2. It could provide further insight into the biosynthetic
mechanism of polysaccharide-like bioflocculant in strain F2.

<>

<1>Li, B., Chen, Y., Liu, Q., Hu, S., Chen, X.
<2>Complete Genome Analysis of Sulfobacillus acidophilus Strain TPY, Isolated from a Hydrothermal Vent in the Pacific Ocean.
<3>J. Bacteriol.
<4>193
<5>5555-5556
<6>2011
<7>Sulfobacillus acidophilus strain TPY is a moderately thermoacidophilic bacterium originally
isolated from a hydrothermal vent in the Pacific
Ocean. Ferrous iron and sulfur oxidation in acidic environments in strain
TPY have been confirmed. Here we report the genome sequence and annotation
of the strain TPY, which is the first complete genome of Sulfobacillus
acidophilus.

<>

<1>Li, B., Shi, Y., Ibrahim, M., Liu, H., Shan, C., Wang, Y., Kube, M., Xie, G.L., Sun, G.
<2>Genome Sequence of the Rice Pathogen Dickeya zeae Strain ZJU1202.
<3>J. Bacteriol.
<4>194
<5>4452-4453
<6>2012
<7>Dickeya zeae is a phytopathogenic bacterium causing soft rot diseases in a wide range of
economically important crops. Here we present the draft genome sequence
of strain ZJU1202, which is the causal agent of rice foot rot in China. The draft
genome will contribute to epidemiological and comparative genomic studies and the
quarantine of this devastating phytopathogen.

<>

<1>Li, C., Li, Y.H., Zhang, X.M., Stafford, P., Dinu, V.
<2>ICRPfinder: a fast pattern design algorithm for coding sequences and its application in finding potential restriction enzyme recognition sites.
<3>BMC Bioinformatics
<4>10
<5>286
<6>2009
<7>Background: Restriction enzymes can produce easily definable segments from DNA sequences by
using a variety of cut patterns. There are,
however, no software tools that can aid in gene building - that is,
modifying wild-type DNA sequences to express the same wild-type amino
acid sequences but with enhanced codons, specific cut sites, unique
post-translational modifications, and other engineered-in components
for recombinant applications. A fast DNA pattern design algorithm,
ICRPfinder, is provided in this paper and applied to find or create
potential recognition sites in target coding sequences.
Results: ICRPfinder is applied to find or create restriction enzyme
recognition sites by introducing silent mutations. The algorithm is
shown capable of mapping existing cut-sites but importantly it also can
generate specified new unique cut-sites within a specified region that
are guaranteed not to be present elsewhere in the DNA sequence.
Conclusion: ICRPfinder is a powerful tool for finding or creating
specific DNA patterns in a given target coding sequence. ICRPfinder
finds or creates patterns, which can include restriction enzyme
recognition sites, without changing the translated protein sequence.
ICRPfinder is a browser-based JavaScript application and it can run on
any platform, in on-line or off-line mode.

<>

<1>Li, D., Greenfield, P., Rosewarne, C.P., Midgley, D.J.
<2>Draft Genome Sequence of Thermoanaerobacter sp. Strain A7A, Reconstructed from a  Metagenome Obtained from a High-Temperature Hydrocarbon Reservoir in the Bass  Strait, Australia.
<3>Genome Announcements
<4>1
<5>e00701-13
<6>2013
<7>The draft genome sequence of Thermoanaerobacter sp. strain A7A was reconstructed  from a
metagenome of a microbial consortium obtained from the Tuna oil field in
the Gippsland Basin, Australia. The organism is a strict anaerobe that is
predicted to ferment a range of simple sugars and undertake sulfur reduction.

<>

<1>Li, E., Beard, C., Jaenisch, R.
<2>Role for DNA methylation in genomic imprinting.
<3>Nature
<4>366
<5>362-366
<6>1993
<7>The paternal and maternal genomes are not equivalent and both are required for mammalian
development. The difference between the parental genomes is believed to be due to
gametespecific differential modification, a process known as genomic imprinting. The study of
transgene methylation has shown that methylation patterns can be inherited in a
parent-of-origin-specific manner, suggesting that DNA mthylation may play a role in genomic
imprinting. The functional significance of DNA methylation in genomic imprinting was
strengthened by the recent finding that CpG islands (or sites) in three imprinted genes, H19,
insulin-like growth factor 2 (Igf-2), and Igf-2 receptor (Igf-2r), are differentially
methylated depending on their parental origin. We have examined the expression of these three
imprinted genes in mutant mice that are deficient in DNA methyltransferase activity. We report
here that expression of all three genes was affected in mutant embryos: the normally silent
paternal allele of the H19 gene was activated, whereas the normally active paternal allele of
the Igf-2 gene and the active maternal allele of the Igf-2r gene were repressed. Our results
demonstrate that a normal level of DNA methylation is required for controlling differential
expression of the paternal and maternal alleles of imprinted genes.

<>

<1>Li, E., Bestor, T.H., Jaenisch, R.
<2>Targeted mutation of the DNA methyltransferase gene results in embryonic lethality.
<3>Cell
<4>69
<5>915-926
<6>1992
<7>Gene targeting in embryonic stem (ES) cells has been used to mutate the murine DNA
methyltransferase gene. ES cell lines homozygous for the mutation were generated by
consecutive targeting of both wild-type alleles; the mutant cells were viable and showed no
obvious abnormalities with respect to growth rate or morphology, and had only trace levels of
DNA methyltransferase activity. A quantitative end-labeling assay showed that the level of m5C
in the DNA of homozygous mutant cells was about one-third that of wild-type cells, and
Southern blot analysis after cleavage of the DNA with a methylation-sensitive restriction
endonuclease revealed substantial demethylation of endogenous retroviral DNA. The mutation was
introduced into the germline of mice and found to cause a recessive lethal phenotype.
Homozygous embryos were stunted, delayed in development, and did not survive past
mid-gestation. The DNA of homozygous embryos showed a reduction of the level of m5C similar to
that of homozygous ES cells. These results indicate that while a 3-fold reduction in levels of
genomic m5C has no detectable effect on the viability or proliferation of ES cells in culture,
a similar reduction of DNA methylation in embryos causes abnormal development and embryonic
lethality.

<>

<1>Li, E., Okano, M., Xie, S.
<2>De novo DNA cytosine methyltransferase genes, polypeptides and uses thereof.
<3>US Patent Office
<4>US 7342108 A
<5>
<6>2008
<7>De novo DNA cytosine methyltransferase polynucleotides and polypeptides and methods for
producing said polypeptides are disclosed.  Also disclosed are methods for utilizing de novo
DNA cytosine methyltransferase polynucleotides and polypeptides in diagnostic assays, for an
in vitro DNA methylation application and therapeutic applications such as the treatment of
neoplastic disorders.

<>

<1>Li, E., Okano, M., Xie, S., Chen, T.
<2>De novo DNA cytosine methyltransferase genes, polypeptides and uses thereof.
<3>US Patent Office
<4>US 7368551 A
<5>
<6>2008
<7>De novo DNA cytosine methyltransferase polynucleotides and polypeptides and methods for
producing said polypeptides are disclosed.  Also disclosed are methods for utilizing de novo
DNA cytosine methyltransferase polynucleotides and polypeptides in diagnostic assays, in vitro
DNA methylation assays for screening agonists and antagonists, and therapeutic applications
such as the treatment of neoplastic disorders.

<>

<1>Li, F., Jiang, P., Zheng, H., Wang, S., Zhao, G., Qin, S., Liu, Z.
<2>Draft Genome Sequence of Marine bacterium Streptomyces griseoaurantiacus M045, which produces Novel Manumycin-type Antibiotics with a pABA Core Component.
<3>J. Bacteriol.
<4>193
<5>3417-3418
<6>2011
<7>Streptomyces griseoaurantiacus M045 isolated from marine sediment produces manumycin and
chinikomycin antibiotics. Here we present a high-quality
draft genome sequence of S. griseoaurantiacus M045, the first marine
Streptomyces species to be sequenced and annotated. The genome encodes
several gene clusters for biosynthesis of secondary metabolites and has
provided insight into genomic islands linking secondary metabolism to
functional adaptation in marine S. griseoaurantiacus M045.

<>

<1>Li, F., Papworth, M., Minczuk, M., Rohde, C., Zhang, Y., Ragozin, S., Jeltsch, A.
<2>Chimeric DNA methyltransferases target DNA methylation to specific DNA sequences and repress expression of target genes.
<3>Nucleic Acids Res.
<4>35
<5>100-112
<6>2007
<7>Gene silencing by targeted DNA methylation has potential applications in basic research and
therapy. To establish targeted methylation in human
cell lines, the catalytic domains (CDs) of mouse Dnmt3a and Dnmt3b DNA
methyltransferases (MTases) were fused to different DNA binding domains
(DBD) of GAL4 and an engineered Cys2His2 zinc finger domain. We
demonstrated that (i) Dense DNA methylation can be targeted to specific
regions in gene promoters using chimeric DNA MTases. (ii) Site-specific
methylation leads to repression of genes controlled by various cellular or
viral promoters. (iii) Mutations affecting any of the DBD, MTase or target
DNA sequences reduce targeted methylation and gene silencing. (iv)
Targeted DNA methylation is effective in repressing Herpes Simplex Virus
type 1 (HSV-1) infection in cell culture with the viral titer reduced by
at least 18-fold in the presence of an MTase fused to an engineered zinc
finger DBD, which binds a single site in the promoter of HSV-1 gene
IE175k. In short, we show here that it is possible to direct DNA MTase
activity to predetermined sites in DNA, achieve targeted gene silencing in
mammalian cell lines and interfere with HSV-1 propagation.

<>

<1>Li, G., Hu, F.Z., Yang, X., Cui, Y., Yang, J., Qu, F., Gao, G.F., Zhang, J.R.
<2>Complete Genome Sequence of Streptococcus pneumoniae Strain ST556, a Multidrug-Resistant Isolate from an Otitis Media Patient.
<3>J. Bacteriol.
<4>194
<5>3294-3295
<6>2012
<7>Streptococcus pneumoniae is a major pathogen causing bacterial infection in the middle ear of
humans. We previously used S. pneumoniae strain ST556, a
low-passage 19F isolate from an otitis media patient, to perform a whole-genome
screen for ear infection-associated genes in a chinchilla model. This report
presents the complete genome sequence of ST556. The genome sequence will provide
information complementary to the experimental data from our genetic study of this
strain.

<>

<1>Li, G., Mo, Z., Li, J., Xiao, P., Hao, B.
<2>Complete Genome Sequence of Vibrio anguillarum M3, a Serotype O1 Strain Isolated  from Japanese Flounder in China.
<3>Genome Announcements
<4>1
<5>e00769-13
<6>2013
<7>Vibrio anguillarum is an important bacterial pathogen that causes vibriosis in marine fish. We
present the complete genome sequence of V. anguillarum M3, a
serotype O1 clinical strain isolated from Japanese flounder (Paralichthys
olivaceus) in Shandong, China.

<>

<1>Li, G., Xie, F., Zhang, Y., Wang, C.
<2>Draft Genome Sequence of Actinobacillus pleuropneumoniae Serotype 7 Strain S-8.
<3>J. Bacteriol.
<4>194
<5>6606-6607
<6>2012
<7>The Gram-negative bacterium Actinobacillus pleuropneumoniae is the etiological agent of
porcine pleuropneumonia, a respiratory disease that leads to severe
economic losses in the swine industry. For years, scientists working with it have
lacked a reliable genome sequence for comparison with other Actinobacillus
species. Here, we report the draft genome sequence of A. pleuropneumoniae
serotype 7 (strain S-8), isolated from swine lung in China in 1992.

<>

<1>Li, H., Pellenz, S., Ulge, U., Stoddard, B.L., Monnat, R.J. Jr.
<2>Generation of single-chain LAGLIDADG homing endonucleases from native homodimeric precursor proteins.
<3>Nucleic Acids Res.
<4>37
<5>1650-1662
<6>2009
<7>Homing endonucleases (HEs) cut long DNA target sites with high specificity to initiate and
target the lateral transfer of mobile introns or inteins.
This high site specificity of HEs makes them attractive reagents for gene
targeting to promote DNA modification or repair. We have generated several
hundred catalytically active, monomerized versions of the
well-characterized homodimeric I-CreI and I-MsoI LAGLIDADG family homing
endonuclease (LHE) proteins. Representative monomerized I-CreI and I-MsoI
proteins (collectively termed mCreIs or mMsoIs) were characterized in
detail by using a combination of biochemical, biophysical and structural
approaches. We also demonstrated that both mCreI and mMsoI proteins can
promote cleavage-dependent recombination in human cells. The use of single
chain LHEs should simplify gene modification and targeting by requiring
the expression of a single small protein in cells, rather than the
coordinate expression of two separate protein coding genes as is required
when using engineered heterodimeric zinc finger or homing endonuclease
proteins.

<>

<1>Li, H., Tong, Y., Huang, Y., Bai, J., Yang, H., Liu, W., Cao, W.
<2>Complete Genome Sequence of Bartonella quintana, a Bacterium Isolated from Rhesus Macaques.
<3>J. Bacteriol.
<4>194
<5>6347
<6>2012
<7>Bartonella quintana is a re-emerging pathogen and the causative agent of a broad  spectrum of
disease manifestations in humans. The present study reports the
complete genome of B. quintana strain RM_11, which was isolated from rhesus
macaques.

<>

<1>Li, H., Ulge, U.Y., Hovde, B.T., Doyle, L.A., Monnat, R.J.Jr.
<2>Comprehensive homing endonuclease target site specificity profiling reveals evolutionary constraints and enables genome engineering applications.
<3>Nucleic Acids Res.
<4>40
<5>2587-2598
<6>2012
<7>Homing endonucleases (HEs) promote the evolutionary persistence of selfish DNA elements by
catalyzing element lateral transfer into new host organisms. The high
site specificity of this lateral transfer reaction, termed homing, reflects both
the length (14-40 bp) and the limited tolerance of target or homing sites for
base pair changes. In order to better understand molecular determinants of
homing, we systematically determined the binding and cleavage properties of all
single base pair variant target sites of the canonical LAGLIDADG homing
endonucleases I-CreI and I-MsoI. These Chlorophyta algal HEs have very similar
three-dimensional folds and recognize nearly identical 22 bp target sites, but
use substantially different sets of DNA-protein contacts to mediate site-specific
recognition and cleavage. The site specificity differences between I-CreI and
I-MsoI suggest different evolutionary strategies for HE persistence. These
differences also provide practical guidance in target site finding, and in the
generation of HE variants with high site specificity and cleavage activity, to
enable genome engineering applications.

<>

<1>Li, H., Zhou, S., Johnson, T., Vercruysse, K., Ropelewski, A.J., Thannhauser, T.W.
<2>Draft Genome Sequence of New Bacillus cereus Strain tsu1.
<3>Genome Announcements
<4>2
<5>e01294-14
<6>2014
<7>This paper reports the draft genome sequence of new Bacillus cereus strain tsu1,  isolated on
an agar-cellulose plate. The draft genome sequence is 5.81 Mb,
revealing 5,673 coding sequences. It contains genes for cellulose-degradation and
biosynthesis pathways of polyhydroxybutyrate (PHB) and 8 rRNA genes (5S, 16S, and
23S).

<>

<1>Li, J., Lan, R., Xiong, Y., Ye, C., Yuan, M., Liu, X., Chen, X., Yu, D., Liu, B., Lin, W., Bai, X., Wang, Y., Sun, Q., Wang, Y., Zhao, H., Meng, Q., Chen, Q., Zhao, A., Xu, J.
<2>Sequential Isolation in a Patient of Raoultella planticola and Escherichia coli Bearing a Novel ISCR1 Element Carrying blaNDM-1.
<3>PLoS ONE
<4>9
<5>E89893
<6>2014
<7>BACKGROUND: The gene for New Delhi metallo-beta-lactamase 1 (NDM-1) has been
reported to be transmitted via plasmids which are easily transferable and capable
of wide distribution. We report the isolation of two NDM-1 producing strains and
possible in vivo transfer of blaNDM-1 in a patient. METHODS: Clinical samples
were collected for bacterial culture and antibiotic susceptibility testing from a
patient during a 34-day hospitalization. The presence of blaNDM-1 was detected by
PCR and sequencing. Plasmids of interest were sequenced. Medical records were
reviewed for evidence of association between the administration of antibiotics
and the acquisition of the NDM-1 resistance. RESULTS: A NDM-1 positive Raoultella
planticola was isolated from blood on the ninth day of hospitalization without
administration of any carbapenem antibiotics and a NDM-1 positive Escherichia
coli was isolated from feces on the 29th day of hospitalization and eight days
after imipenem administration. The blaNDM-1 was carried by a 280 kb plasmid
pRpNDM1-1 in R. planticola and a 58 kb plasmid pEcNDM1-4 in E. coli. The two
plasmids shared a 4812 bp NDM-1-ISCR1 element which was found to be excisable
from the plasmid as a free form and transferrable in vitro to a NDM-1 negative
plasmid from E. coli. CONCLUSION: blaNDM-1 was embedded in an ISCR1 complex class
1 integron as a novel 4812 bp NDM-1-ISCR1 element. The element was found to be
able to self excise to become a free form, which may provide a new vehicle for
NDM-1 dissemination. This mechanism could greatly accelerate the spread of NDM-1
mediated broad spectrum beta-lactam resistance.

<>

<1>Li, J., Peng, H., Xu, L.G., Xie, Y.Z., Xuan, X.B., Ma, C.X., Hu, S., Chen, Z.X., Yang, W., Xie, Y.P., Pan, Y., Tao, L.
<2>Draft Genome Sequence of Haemophilus parasuis gx033, a Serotype 4 Strain Isolated from the Swine Lower Respiratory Tract.
<3>Genome Announcements
<4>1
<5>e00224-13
<6>2013
<7>Haemophilus parasuis serotype 4 is a Gram-negative pathogen that is the most prevalent H.
parasuis serovar in the world, but its genome sequence information
has not yet been reported. Thus, we determined the genome of H. parasuis strain
gx033, a serovar 4 strain isolated from a lung specimen of a diseased piglet in
southwestern China. Here, we present the first draft genome sequence of this
species.

<>

<1>Li, J., Yan, H.F., Wang, K.M., Tan, W.H., Zhou, X.W.
<2>Hairpin fluorescence DNA probe for real-time monitoring of DNA methylation.
<3>Anal. Chem.
<4>79
<5>1050-1056
<6>2007
<7>DNA methylation catalyzed by methylase plays an important role in many biological events.
However, traditional methods of methylase activity
analysis by gel electrophoresis were laborious and discontinuous. In
this paper, we report a new strategy to study methylase activity using
fluorescent probes coupled with enzyme-linkage reactions. A hairpin DNA
probe is prepared with a fluorophore and a quencher linked at the 5'-
and 3'-terminus of the probe. A disturbance of the stem sequence by DNA
methylation would cause the separation of the fluorophore and the
quencher, resulting in the restoration of the fluorescence. We used DNA
adenine methylation (Dam) methyltransferase (MTase) and Dpn I
endonuclease, both having a 5'-G-A-T-C-3' recognition sequence. Dam
MTase catalyzed the methylation of the sequence of 5'-GATC-3', and Dpn
I cut the sequence of 5'-G-Am-T-C-3'. The fluorescence of the hairpin
probe was restored when it was cleaved by Dpn I endonuclease during the
course of methylation. Unlike traditional methods, this assay was done
in real time and could be used to monitor the dynamic process of
methylation. Our method is easy, simple, and nonradioactive, yet as
efficient as gel electrophoresis in detecting the activity of
methylase. It also had the potential to screen suitable inhibitor drugs
for Dam methylase.

<>

<1>Li, J.-K., Tu, J.
<2>Ssp RFI, a novel class-II restriction endonuclease from Synechococcus RF-1 recognizing 5'TT/CGAA-3' .
<3>Nucleic Acids Res.
<4>19
<5>4770
<6>1991
<7>A novel class II restriction endonuclease has been isolated from a
cyanobacterium Synechococcus RF-1, which shows a circadian rhythm in its
nitrogen fixation activity, and designated as SspRFI.  The enzyme recognizes
the palindrome 5'-TT/CGAA-3' and generates 5'-protruding CG-dinucleotides.  Its
isoschizomers have also been isolated from Streptomyces, Bacillus,
Lactobacillus and other cyanobacteria.  SspRFI favors low salt conditions and
can be inhibited by a methylation of the intter adenine residue.

<>

<1>Li, J.Y., Pu, M.T., Hirasawa, R., Li, B.Z., Huang, Y.N., Zeng, R., Jing, N.H., Chen, T., Li, E., Sasaki, H., Xu, G.L.
<2>Synergistic function of DNA Methyltransferases dnmt3a and dnrnt3b in the methylation of Oct4 and Nanog.
<3>Mol. Cell. Biol.
<4>27
<5>8748-8759
<6>2007
<7>DNA methylation plays an important role in gene silencing in mammals. Two de novo
methyltransferases, Dnmt3a and Dnmt3b, are required for the
establishment of genomic methylation patterns in development. However,
little is known about their coordinate function in the silencing of
genes critical for embryonic development and how their activity is
regulated. Here we show that Dnmt3a and Dnmt3b are the major components
of a native complex purified from embryonic stem cells. The two enzymes
directly interact and mutually stimulate each other both in vitro and
in vivo. The stimulatory effect is independent of the catalytic
activity of the enzyme. In differentiating embryonic carcinoma or
embryonic stem cells and mouse postimplantation embryos, they function
synergistically to methylate the promoters of the Oct4 and Nanog genes.
Inadequate methylation caused by ablating Dnmt3a and Dnmt3b is
associated with dysregulated expression of Oct4 and Nanog during the
differentiation of pluripotent cells and mouse embryonic development.
These results suggest that Dnmt3a and Dnmt3b form a complex through
direct contact in living cells and cooperate in the methylation of the
promoters of Oct4 and Nanog during cell differentiation. The physical
and functional interaction between Dnmt3a and Dnmt3b represents a novel
regulatory mechanism to ensure the proper establishment of genomic
methylation patterns for gene silencing in development.

<>

<1>Li, K., Wang, S., Shi, Y., Qu, J., Zhai, Y., Xu, L., Xu, Y., Song, J., Liu, L., Rahman, M.A., Yan, Y.
<2>Genome Sequence of Paracoccus sp.TRP, a Chlorpyrifos Biodegrader.
<3>J. Bacteriol.
<4>193
<5>1786-1787
<6>2011
<7>A Paracoccus sp. strain TRP, isolated from activated sludge, could completely biodegrade
chlorpyrifos and 3,5,6-trichloro-2-pyridinol. Here, we report the draft genome sequence of
Paracoccus sp.strain TRP which could be used to predict genes for xenobiotics biodegradation
and metabolism.

<>

<1>Li, L.
<2>Studies on FokI (a type IIS) restriction endonuclease.
<3>Diss. Abstr.
<4>55
<5>877B-878B
<6>1994
<7>FokI restriction endonuclease recognizes the nonpalindromic pentadeoxyribonucleotide
5'-GGATG-3'/5'-CATCC-3' in duplex DNA and cleaves 9 and 13 nucleotides away from the
recognition site. This implies that this enzyme has separate domains for DNA recognition and
cleavage. The fokIM and fokIR were subcloned into Escherichia coli. FokI endonuclease was
overexpressed and purified to homogeneity. The recognition and cleavage domains of FokI were
analyzed by trypsin digestion. FokI in the presence of a DNA substrate is cleaved into a 41
kDa amino-terminal fragment and a 25 kDA carboxyl-terminal fragment. Gel-mobility-shift assays
indicate that all the protein contacts necessary for the sequence-specific recognition of DNA
substrates are encoded within the 41 kDa fragment. On further digestion, the 41 kDa fragment
degrades into 30 kDa amino-terminal and 11 kDa carboxyl-terminal fragments which together bind
DNA substrates. Thus, the 41 kDa amino-terminal fragment constitutes the FokI recognition
domain. This was confirmed by the characterization of two carboxyl-terminal deletion mutant
proteins (41 kDa and 30 kDa, respectively) of FokI. The 41 kDa mutant protein binds the DNA
sequence 5'-GGATG-3':5' - CATCC-3' specifically like the wild-type FokI. The 30 kDa mutant
protein does not bind DNA. Addition of the HPLC-purified 11 kDa fragment to the 30 kDa mutant
protein restores its sequence specific DNA-binding property. The affinity of the 41 kDa mutant
protein for the DNA substrate is comparable to that of the wild-type FokI. The 41 kDa mutant
protein interacts with its substrate in a similar way compared to the wild-type enzyme as
revealed by hydroxyl radical footprinting experiments. The 25 kDa carboxyl-terminal fragment
of FokI cleaves DNA substrates non-specifically in the presence of MgCl2. Thus, the 25 kDa
fragment constitutes the FokI cleavage domain. Additional amino acids have been introduced
between the two domains of FokI by insertion mutagenesis. The mutant enzymes have the same DNA
sequence specificity as the wild-type FokI. However, compared with the wild-type enzyme, they
cleave one nucleotide further away from the recognition site on both strands of the DNA
substrates. Thus, it is possible to alter the cleavage distance of FokI by protein
engineering.

<>

<1>Li, L., Bannantine, J.P., Zhang, Q., Amonsin, A., May, B.J., Alt, D., Banerji, N., Kanjilal, S., Kapur, V.
<2>The complete genome sequence of Mycobacterium avium subspecies paratuberculosis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>12344-12349
<6>2005
<7>We describe here the complete genome sequence of a common clone of Mycobacterium avium
subspecies paratuberculosis (Map) strain K-10, the
causative agent of Johne's disease in cattle and other ruminants. The K-10
genome is a single circular chromosome of 4,829,781 base pairs and encodes
4,350 predicted ORFs, 45 tRNAs, and one rRNA operon. In silico analysis
identified >3,000 genes with homologs to the human pathogen, M.
tuberculosis (Mtb), and 161 unique genomic regions that encode 39
previously unknown Map genes. Analysis of nucleotide substitution rates
with Mtb homologs suggest overall strong selection for a vast majority of
these shared mycobacterial genes, with only 68 ORFs with a synonymous to
nonsynonymous substitution ratio of >2. Comparative sequence analysis
reveals several noteworthy features of the K-10 genome including: a
relative paucity of the PE/PPE family of sequences that are implicated as
virulence factors and known to be immunostimulatory during Mtb infection;
truncation in the EntE domain of a salicyl-AMP ligase (MbtA), the first
gene in the mycobactin biosynthesis gene cluster, providing a possible
explanation for mycobactin dependence of Map; and Map-specific sequences
that are likely to serve as potential targets for sensitive and specific
molecular and immunologic diagnostic tests. Taken together, the
availability of the complete genome sequence offers a foundation for the
study of the genetic basis for virulence and physiology in Map and enables
the development of new generations of diagnostic tests for bovine Johne's
disease.

<>

<1>Li, L., Chandrasegaran, S.
<2>Alteration of the cleavage distance of Fok I restriction endonuclease by insertion mutagenesis.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>2764-2768
<6>1993
<7>FokI restriction endonuclease recognizes the nonpalindromic pentadeoxyribonucleotide
5'-GGATC-3'-5'CATCC-3' in duplex DNA and cleaves 9 and 13 nucleotides away from the
recognition site. Recently, we reported the presence of two distinct and separable protein
domains within this enzyme-one for the sequence-specific recognition and the other for
endonuclease activity. Here, we report the construction of two insertion mutants of FokI
endonuclease. The mutant enzymes were purified, and their cleavage properties were
characterized. The mutants have the same DNA sequence specificity as the wild-type enzyme.
However compared with the wild-type enzyme, they cleave one nucleotide further away from the
recognition site on both strands of the DNA substrates. Thus, it is possible to alter the
cleavage distance of FokI by protein engineering.

<>

<1>Li, L., Cheng, C.K., Cheung, M.K., Law, P.T., Ling, J.M., Kam, K.M., Cheung, W.M., Kwan, H.S.
<2>Draft Genome Sequence of Salmonella enterica Serovar Typhimurium ST1660/06, a Multidrug-Resistant Clinical Strain Isolated from a Diarrheic Patient.
<3>J. Bacteriol.
<4>194
<5>6319-6320
<6>2012
<7>Salmonella enterica serovar Typhimurium is one of the most prevalent serovars of  Salmonella
that causes human gastroenteritis. Here, we report the draft genome
sequence of the S. Typhimurium multidrug-resistant strain ST1660/06. Comparative
genomic analysis unveiled three strain-specific genomic islands that potentially
confer the multidrug resistance and virulence of the strain.

<>

<1>Li, L., Su, F., Wang, Y., Zhang, L., Liu, C., Li, J., Ma, C., Xu, P.
<2>Genome Sequences of Two Thermophilic Bacillus licheniformis Strains, Efficient Producers of Platform Chemical 2,3-Butanediol.
<3>J. Bacteriol.
<4>194
<5>4133-4134
<6>2012
<7>Both Bacillus licheniformis strains 10-1-A and 5-2-D are efficient producers of
2,3-butanediol. Here we present 4.3-Mb and 4.2-Mb assemblies of their genomes.
The key genes for the regulation and metabolism of 2,3-butanediol production were
annotated, which may provide further insights into the molecular mechanism for
the production of 2,3-butanediol with high yield and productivity.

<>

<1>Li, L., Wang, Y., Li, K., Su, F., Ma, C., Xu, P.
<2>Genome Sequence of meso-2,3-Butanediol-Producing Strain Serratia marcescens ATCC  14041.
<3>Genome Announcements
<4>2
<5>e00590-14
<6>2014
<7>Serratia marcescens strain ATCC 14041 was found to be an efficient meso-2,3-butanediol
(meso-2,3-BD) producer from glucose and sucrose. Here we
present a 5.0-Mb assembly of its genome. We have annotated 4 coding sequences
(CDSs) for meso-2,3-BD fermentation and 2 complete operons including 6 CDSs for
sucrose utilization.

<>

<1>Li, L., Wang, Y., Wang, K., Li, K., Ma, C., Xu, P.
<2>Genome Sequence of Thermophilic Bacillus licheniformis Strain 3F-3, an Efficient  Pentose-Utilizing Producer of 2,3-Butanediol.
<3>Genome Announcements
<4>2
<5>e00615-14
<6>2014
<7>Bacillus licheniformis strain 3F-3 is an efficient pentose-utilizing producer of  platform
chemical, 2,3-butanediol. Here we present a 4.1-Mb assembly of its
genome. The key genes for pentose utilization, regulation, and metabolism of
2,3-butanediol were annotated, which may provide further insights into the
molecular mechanism of 2,3-butanediol production from biomass pentose.

<>

<1>Li, L., Wu, L.P., Chandrasegaran, S.
<2>Functional domains in FokI restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>4275-4279
<6>1992
<7>The PCR was used to alter transcriptional and translational signals surround the
Flavobacterium okeanokoites restriction endonuclease (fokIR) gene, so as to achieve high
expression in Escherichia coli. By changing the ribosome-binding site sequence preceding the
fokIR gene to match the consensus E. coli signal and by placing a positive retroregulator
stem-loop sequence downstream of the gene, FokI yield was increased to 5-8% of total cellular
protein. FokI was purified to homogeneity with phosphocellulose, DEAE-Sephadex, and gel
chromatography, yielding 50 mg of pure FokI endonuclease per liter of culture medium. The
recognition and cleavage domains of FokI were analyzed by trypsin digestion. FokI in the
absence of a DNA substrate cleaves into a 58-kDa carboxyl-terminal and 8-kDa amino-terminal
fragment. The 58-kDa fragment does not bind the DNA substrate. FokI in the presence of a DNA
substrate cleaves into a 41-kDa amino-terminal fragment and a 25-kDa carboxyl-terminal
fragment. On further digestion, the 41-kDa fragment degrades into 30-kDa amino terminal and
11-kDa carboxyl-terminal fragments. The cleaved fragments both bind DNA substrates, as does
the 41-kDa fragment. Gel-mobility-shift assays indicate that all the protein contacts
necessary for the sequence-specific recognition of DNA substrates are encoded within the
41-kDa fragment. Thus, the 41-kDa amino-terminal fragment constitutes the FokI recognition
domain. The 25-kDa fragment purified by using a DEAE-Sephadex column, cleaves nonspecifically
both methylated (pACYCfokIM) and nonmethylated (pTZ19R) DNA substrates in the presence of
MgCl2. Thus, the 25-kDa carboxyl-terminal fragment constitutes the FokI cleavage domain.

<>

<1>Li, L., Wu, L.P., Clarke, R., Chandrasegaran, S.
<2>C-terminal deletion mutants of the FokI restriction endonuclease.
<3>Gene
<4>133
<5>79-84
<6>1993
<7>*
We have constructed two C-terminal deletion mutants of the FokI restriction endonuclease by
using the polymerase-chain-reaction technique and expressed them in Escherichia coli. The two
mutant proteins (MP) of 41 and 30 kDa, were purified to homogeneity and their DNA-binding
properties were characterized. The 41-kDa MP specifically binds the DNA sequence:

   5'GGATG
   3'CCTAC

like the wild-type (wt) FokI, but does not cleave DNA. The 30-kDa MP does not bind DNA. The
affinity of the 41-kDa MP for the DNA substrate is comparable to that of wt FokI. The 41-kDa
MP interacts with its substrate like the wt FokI, as revealed by hydroxyl radical footprinting
experiments. In the presence of a DNA substrate, the 41-kDa MP is cleaved by trypsin into a
30-kDa N-terminal fragment and an 11-kDa C-terminal fragment. Addition of the HPLC-purified
11-kDa C-terminal fragment to the 30-kDa MP restores its sequence-specific DNA binding
property.
These results confirm that the N-terminal 41-kDa fragment of the FokI ENase constitutes the
DNA recognition domain of the ENase.


<>

<1>Li, L.G., Cai, L., Zhang, T.
<2>Genome of Cupriavidus sp. HMR-1, a Heavy Metal-Resistant Bacterium.
<3>Genome Announcements
<4>1
<5>e00202-12
<6>2013
<7>Cupriavidus sp. HMR-1 was isolated from a heavy metal-enriched culture of activated sludge
from a wastewater treatment plant in Hong Kong. Here, we release
the HMR-1 genome to provide basic genetic characteristics for a better
understanding of its multiple heavy metal resistance properties.

<>

<1>Li, L.I., Tanyashin, V.I., Matvienko, N.I., Bayev, A.A.
<2>Production of Specific Fragments of T4 DNA by Endonuclease EcoRI.
<3>Dokl. Akad. Nauk.
<4>223
<5>1262-1265
<6>1975
<7>Restriction enzyme EcoRI was used to cleave nonglucosylated bacteriophage T4
DNA in a minimum 41 fragments, which were separated on agarose gels.  The
fragments were estimated to fall within the size range of 8.2-0.38 Mdalton. It
was shown that either alpha- or beta-glucosylation protects fully T4 DNA
against EcoRI function. Recombinant cells E. coli K12 r 2,4-, r 6-(RI)/6
harbouring the RI plasmid were isolated.  The efficiency of plating of
different T4 mutants with nonglucosylated and partly glucosylated DNA on them
was not lower then on the strain E. coli K 12 r 2,4-,r6-.

<>

<1>Li, L.L., Taghavi, S., Izquierdo, J.A., van der Lelie, D.
<2>Complete Genome Sequence of Clostridium sp. Strain BNL1100, a Cellulolytic Mesophile Isolated from Corn Stover.
<3>J. Bacteriol.
<4>194
<5>6982-6983
<6>2012
<7>We present the full genome sequence of Clostridium sp. strain BNL1100, a Gram-positive,
endospore-forming, lignocellulolytic bacterium isolated from a
corn stover enrichment culture. The 4,613,747-bp genome of strain BNL1100
contains 4,025 putative protein-coding genes, of which 103 are glycoside
hydrolases, the highest detected number in cluster III clostridia.

<>

<1>Li, M.Z., Elledge, S.J.
<2>MAGIC, an in vivo genetic method for the rapid construction of recombinant DNA molecules.
<3>Nat. Genet.
<4>37
<5>311-319
<6>2005
<7>We describe a highly engineered in vivo cloning method, mating-assisted genetically integrated
cloning (MAGIC), that facilitates the rapid
construction of recombinant DNA molecules. MAGIC uses bacterial mating, in
vivo site-specific endonuclease cleavage and homologous recombination to
catalyze the transfer of a DNA fragment between a donor vector in one
bacterial strain and a recipient plasmid in a separate bacterial strain.
Recombination events are genetically selected and result in placement of
the gene of interest under the control of new regulatory elements with
high efficiency. MAGIC eliminates the need for restriction enzymes, DNA
ligases, preparation of DNA and all in vitro manipulations required for
subcloning and allows the rapid construction of multiple constructs with
minimal effort. We show that MAGIC can generate constructs for expression
in multiple organisms. As this new method requires only the simple mixing
of bacterial strains, it represents a substantial advance in
high-throughput recombinant DNA production that will save time, effort and
expense in functional genomics studies.

<>

<1>Li, N., Zhang, J., Zhang, L.Q., Nie, P.
<2>Difference in genes between a high virulence strain G4 and a low virulence strain G18 of Flavobacterium columnare by using suppression subtractive hybridization.
<3>J. Fish Dis.
<4>33
<5>403-412
<6>2010
<7>Flavobacterium columnare is the causative agent of columnaris disease. Different genetic
groups of F. columnare show to some extent different degrees of virulence. To identify genetic
differences between
the high virulence strain G4 and the low virulence strain G18 of F. columnare, suppression
subtractive hybridization was used. A total of 46 genes were identified from the virulent
strain G4, 35 of which
showed some degree of homology with known proteins and can be classified into 11 categories:
DNA replication or recombination proteins, inorganic
ion transport proteins, outer membrane proteins, enterotoxin, binding proteins, YD repeat
proteins, transposase, chaperon, signal transduction-
related proteins, regulatory proteins, metabolism-
related proteins. Several putative virulence factors identified in other bacteria could also
be identified in the virulent strain G4, such as ferrous
iron transport protein, TonB-dependent receptor, transposases, as well as ABC transporter
permease protein. The flanking region of a putative transposase ISFclI was analysed, and a
putative Rhs element was located at the downstream of the putative transposase. The analysis
of isfcl I gene in
24 strains of F. columnare isolated in China revealed that 11 strains have isfcl I, and all
the strains from Zhaoqing, Anhui and Qingjiang have
isfclI.

<>

<1>Li, N., Zhang, L.Q., Zhang, J., Liu, Z.X., Huang, B., Zhang, S.H., Nie, P.
<2>Type I restriction-modification system and its resistance in electroporation efficiency in Flavobacterium columnare.
<3>Vet. Microbiol.
<4>160
<5>61-68
<6>2012
<7>Flavobacterium columnare, the causative agent of columnaris disease, infects freshwater fish
worldwide. However, the pathogenicity of this
bacterium is poorly understood due possibly to the lack of an efficient
in-frame knockout technique. In order to improve electroporation
efficiency, the type I restriction-modification system (R-M system) was
cloned and its role in electroporation was examined in F. columnare
G(4) strain. The complete sequence of type I R-M system in the
bacterium, designated as Fcl, contains all three subunits of type I R-M
system, named as fclM, fclS, fclR, respectively, with the
identification of a hypothetical gene, fclX. Constitutive transcription
of the three genes was observed in F. columnare G(4) by RT-PCR. The ORF
of fclM and fclS was cloned into the plasmid pACYC184 and transformed
into Escherichia colt TOP10. The resultant E. coli strain, designated
as E. coli TOPmt, was transformed with the integrative plasmid pGL006
constructed for F. columnare G(4). The integrative plasmid was
re-isolated from TOPmt and incubated with the lysate of F. columnare
G(4). The re-isolated integrative plasmid, designated as pGL006',
showed higher resistance than pGL006. With pGL006', the electroporation
efficiency of the strain G(4) increased 2.6 times, while that of F.
columnare G(18) was not obviously improved. Furthermore, a method to
improve the electroporation efficiency of F. columnare G(4) was
developed using the integrative plasmid methylated by E. coil TOPmt
which contains the fclM and fclS gene of F. columnare G(4). Further
analyses showed that the fcl gene cluster may be a unique type I R-M
system in F. columnare G(4). It will be of significant interest to
examine the composition and diversity of R-M systems in strains of F.
columnare in order to set up a suitable genetic manipulation system for
the bacterium.

<>

<1>Li, N.Z., Xia, T., Xu, Y.L., Qiu, R.R., Xiang, H., He, D., Peng, Y.Y.
<2>Genome Sequence of Paenibacillus sp. Strain Aloe-11, an Endophytic Bacterium with Broad Antimicrobial Activity and Intestinal Colonization Ability.
<3>J. Bacteriol.
<4>194
<5>2117-2118
<6>2012
<7>Paenibacillus sp. strain Aloe-11, a Gram-positive, spore-forming, facultatively anaerobic
bacterium isolated from the root of Aloe chinensis in the southwest
region of China, has excellent antibiotic activity and intestine colonization
ability. Here, we present the 5.8-Mb draft genome sequence of Paenibacillus sp.
strain Aloe-11.

<>

<1>Li, P., Yang, C., Wang, J., Yi, S., Li, H., Wang, Y., Qiu, S., Song, H.
<2>Draft Genome Sequence of a New Shigella flexneri Subserotype, 4S BJ10610.
<3>Genome Announcements
<4>2
<5>e00715-14
<6>2014
<7>Shigella flexneri is of great concern in the prevalence of shigellosis and resistance to many
antibiotics in developing countries. Here, we report the draft
genome sequence of a new S. flexneri subserotype, 4S BJ10610, isolated from the
stool specimens of a patient in Beijing, China.

<>

<1>Li, P.Y., Xie, B.B., Zhang, X.Y., Qin, Q.L., Dang, H.Y., Wang, X.M., Chen, X.L., Yu, J., Zhang, Y.Z.
<2>Genetic structure of three fosmid-fragments encoding 16S rRNA genes of the Miscellaneous Crenarchaeotic Group (MCG): implications for physiology and evolution of marine sedimentary archaea.
<3>Environ. Microbiol.
<4>14
<5>467-479
<6>2012
<7>Archaea of the Miscellaneous Crenarchaeotic Group (MCG) exist widely in soil,
freshwater and marine sediments of both surface and subsurface. However, current
knowledge about this group is limited to its phylogenetic diversity. An archaeal
16S library was constructed from a sediment sample from the South China Sea,
which was dominated by MCG and Marine Group I (MG-I). A metagenomic library was
constructed from the same sediment sample, and three MCG fosmids (E6-3G, E37-7F
and E48-1C) containing 16S rRNA genes were screened. Annotation showed that the
three genomic fragments encode a variety of open reading frames (ORFs) that are
potentially homologous to important functional genes related to lipid
biosynthesis, energy metabolism, and resistance to oxidants. No colinear regions
were found between MCG fosmids and reported archaeal genomic fragments or
genomes, suggesting that the MCG archaea are quite different from the sequenced
archaea in gene arrangement. Analyses of both the phylogenies of 16S rRNA genes
and several informational processing genes and nucleotide frequencies showed that
MCG archaea are distinct from MG-I plus relatives. In addition, tetranucleotide
frequency analysis in combination with phylogenetic analysis suggested that some
fragments in the MCG fosmids are probably derived from non-MCG or non-archaeal
genomes.

<>

<1>Li, Q., Hu, Y., Xu, L., Xie, X., Tao, M., Jiao, X.
<2>Complete Genome Sequence of Salmonella enterica Serovar Choleraesuis Vaccine Strain C500 Attenuated by Chemical Mutation.
<3>Genome Announcements
<4>2
<5>e01022-14
<6>2014
<7>Salmonella enterica serovar Choleraesuis strain C500 is a live vaccine attenuated by chemical
methods. Here, we report the complete genome sequence of the strain,
which may be helpful for elucidating the attenuation mechanism of the vaccine
strain.

<>

<1>Li, S., Skov, R.L., Han, X., Larsen, A.R., Larsen, J., Sorum, M., Wulf, M., Voss, A., Hiramatsu, K., Ito, T.
<2>Novel types of staphylococcal cassette chromosome mec elements identified in clonal complex 398 methicillin-resistant Staphylococcus aureus strains.
<3>Antimicrob. Agents Chemother.
<4>55
<5>3046-3050
<6>2011
<7>The structures of staphylococcal cassette chromosome mec (SCCmec) elements
carried by 31 clonal complex 398 (CC398) methicillin-resistant
Staphylococcus aureus (MRSA) strains isolated from the participants at a
conference were analyzed. The SCCmecs were classified into novel types,
namely, IX, X, V(5C2&5) subtype c, and IVa. Type V(5C2&5) subtype c, IX,
and X SCCmecs carried genes conferring resistance to metals. The
structures of SCCmecs from CC398 strains were distinct from those normally
found in humans, adding to the evidence that humans are not the original
host for CC398.

<>

<1>Li, S., Yang, D., Qiu, M., Shao, J., Guo, R., Shen, B., Yin, X., Zhang, R., Zhang, N., Shen, Q.
<2>Complete Genome Sequence of Paenibacillus polymyxa SQR-21, a Plant Growth-Promoting Rhizobacterium with Antifungal Activity and Rhizosphere Colonization Ability.
<3>Genome Announcements
<4>2
<5>e00281-14
<6>2014
<7>Here we report the complete genome sequence of a plant growth-promoting
rhizobacterium (PGPR), Paenibacillus polymyxa SQR-21, which consists of one
circular chromosome of 5,828,438 bp with 5,024 coding sequences (CDS). The data
presented highlight multiple sets of functional genes associated with its
plant-beneficial characteristics.

<>

<1>Li, S., Zhao, H., Li, Y., Niu, S., Cai, B.
<2>Complete Genome Sequence of the Naphthalene-Degrading Pseudomonas putida Strain ND6.
<3>J. Bacteriol.
<4>194
<5>5154-5155
<6>2012
<7>Pseudomonas putida strain ND6 is an efficient naphthalene-degrading bacterium. The complete
genome of strain ND6 was sequenced and annotated. The genes encoding
the enzymes involved in catechol degradation by the ortho-cleavage pathway were
found in the chromosomal sequence, which indicated that strain ND6 is able to
metabolize naphthalene by the catechol meta- and ortho-cleavage pathways.

<>

<1>Li, T., Chen, J., Chang, D., Fang, X., Wang, J., Guo, Y., Su, L., Xu, G., Wang, Y., Chen, Z., Liu, C.
<2>Draft Genome Sequence of Escherichia coli Strain LCT-EC59.
<3>Genome Announcements
<4>1
<5>e00242-12
<6>2013
<7>The space environment is a very special condition under which many organisms change many
features. is employed widely as a prokaryotic model organism in the
fields of biotechnology and microbiology. Here, we present the draft genome
sequence of strain LCT-EC59 exposed to space conditions.

<>

<1>Li, T., Huang, S., Jiang, W.Z., Wright, D., Spalding, M.H., Weeks, D.P., Yang, B.
<2>TAL nucleases (TALNs): hybrid proteins composed of TAL effectors and FokI DNA-cleavage domain.
<3>Nucleic Acids Res.
<4>39
<5>359-372
<6>2010
<7>DNA double-strand breaks enhance homologous recombination in cells and have been exploited for
targeted genome editing through use of engineered
endonucleases. Here we report the creation and initial characterization of
a group of rare-cutting, site-specific DNA nucleases produced by fusion of
the restriction enzyme FokI endonuclease domain (FN) with the
high-specificity DNA-binding domains of AvrXa7 and PthXo1. AvrXa7 and
PthXo1 are members of the transcription activator-like (TAL) effector
family whose central repeat units dictate target DNA recognition and can
be modularly constructed to create novel DNA specificity. The hybrid
FN-AvrXa7, AvrXa7-FN and PthXo1-FN proteins retain both recognition
specificity for their target DNA (a 26 bp sequence for AvrXa7 and 24 bp
for PthXo1) and the double-stranded DNA cleaving activity of FokI and,
thus, are called TAL nucleases (TALNs). With all three TALNs, DNA is
cleaved adjacent to the TAL-binding site under optimal conditions in
vitro. When expressed in yeast, the TALNs promote DNA homologous
recombination of a LacZ gene containing paired AvrXa7 or asymmetric
AvrXa7/PthXo1 target sequences. Our results demonstrate the feasibility of
creating a tool box of novel TALNs with potential for targeted genome
modification in organisms lacking facile mechanisms for targeted gene
knockout and homologous recombination.

<>

<1>Li, T., Pu, F., Yang, R., Fang, X., Wang, J., Guo, Y., Chang, D., Su, L., Guo, N., Jiang, X., Zhao, J., Liu, C.
<2>Draft Genome Sequence of Escherichia coli LCT-EC106.
<3>J. Bacteriol.
<4>194
<5>4443-4444
<6>2012
<7>Escherichia coli is a Gram-negative, rod-shaped bacterium that is commonly found  in the
intestine of warm-blooded organisms. Most E. coli strains are harmless,
but some serotypes can cause serious food poisoning in humans. Here, we present
the complete genome sequence of Escherichia coli LCT-EC106, which was isolated
from CGMCC 1.2385.

<>

<1>Li, W., Shi, J., Wang, X., Han, Y., Tong, W., Ma, L., Liu, B., Cai, B.
<2>Complete nucleotide sequence and organization of the naphthalene catabolic plasmid pND6-1 from Pseudomonas sp. strain ND6.
<3>Gene
<4>336
<5>231-240
<6>2004
<7>Pseudomonas sp. strain ND6, which was isolated from industrial wastewater in
Tianjin, China, was capable of dissimilating naphthalene as sole carbon and
energy sources. We identified one plasmid, pND6-1, which was associated with the
metabolism of naphthalene and determined the complete nucleotide sequence of
pND6-1 (101,858 bp) using a whole-genome-shotgun approach. Computational analyses
indicated that the naphthalene metabolism of the strain ND6 is associated with
this plasmid. This is the first report of a complete sequence of naphthalene
catabolic plasmid. pND6-1 encodes 102 putative coding sequences (CDSs). Among
them, 23 CDSs were predicted to be involved in naphthalene catabolism, 14 were
predicted to be involved in transposition and integration, 2 encoded putative
transporters, 3 were putative transcriptional regulators, and 9 were proteins
necessary for plasmid replication and partitioning. Most of the naphthalene
catabolic genes of pND6-1 have 99-100% identity in amino acid sequences
homologous to their nearest counterparts found in plasmid pDTG1, NAH7 and in a
chromosome region in Pseudomonas stutzeri AN10 except for two duplicated genes
(ND013 and ND016). Results of this study indicated that globally distributed
naphthalene catabolic genes are highly conserved among different bacterial
species.

<>

<1>Li, W., Wu, P., Zhang, H., Cai, C.X.
<2>Signal Amplification of Graphene Oxide Combining with Restriction Endonuclease for Site-Specific Determination of DNA Methylation and Assay of Methyltransferase Activity.
<3>Anal. Chem.
<4>84
<5>7583-7590
<6>2012
<7>Site-specific identification of DNA methylation and assay of MTase activity are important in
determining specific cancer types, providing
insights into the mechanism of gene repression, and developing novel
drugs to treat methylation-related diseases. This work reports an
electrochemical method for gene-specific methylation detection and
MTase activity assay using HpaII endonuclease to improve selectivity
and employing signal amplification of graphene oxide (GO) to enhance
the assay sensitivity. The method was developed by designing a probe
DNA, which was immobilized on electrode surface, to hybridize with
target DNA (one 137 mer DNA from exon 8 promoter region of the Homo
sapiens p53 gene, was extracted from HCT116 cells). The assay is based
on the electrochemical responses of the reporter (thionine), which was
conjugated to 3'-terminus of the probe DNA via GO, after the DNA hybrid
was methylated (under catalysis of M.SssI MTase) and cleaved by HpaII
endonuclease (a site-specific endonuclease recognizing the duplex
symmetrical sequence of 5'-CCGG-3' and catalyzing cleavage between the
cytosines). This model can determine DNA methylation at the site of CpG
and has an ability to discriminate the target DNA sequence from even
single-base mismatched sequence. The electrochemical signal has a
linear relationship with M.SssI activities ranging from 0.1 to 450 U/mL
with a detection limit of similar to(0.05 +/- 0.02) U/mL at a
signal/noise of 3. The advantages of this assay are ease of performance
having a good specificity and selectivity. In addition, we also
demonstrate the method can be used for rapid evaluation and screening
of the inhibitors of MTase and has a potential application in discovery
of new anticancer drugs.

<>

<1>Li, X., Gong, J., Hu, Y., Cai, L., Johnstone, L., Grass, G., Rensing, C., Wang, G.
<2>Genome Sequence of the Moderately Halotolerant, Arsenite-Oxidizing Bacterium Pseudomonas stutzeri TS44.
<3>J. Bacteriol.
<4>194
<5>4473-4474
<6>2012
<7>We present the draft genome sequence of Pseudomonas stutzeri TS44, a moderately halotolerant,
arsenite-oxidizing bacterium isolated from arsenic-contaminated
soil. The genome contains genes for arsenite oxidation, arsenic resistance, and
ectoine/hydroxyectoine biosynthesis. The genome information will be useful for
exploring adaptation of P. stutzeri TS44 to an arsenic-contaminated environment.

<>

<1>Li, X., Gu, Q., Lou, X., Zhang, X., Song, D., Shen, L., Zhao, Y.
<2>Complete Genome Sequence of the Probiotic Lactobacillus plantarum Strain ZJ316.
<3>Genome Announcements
<4>1
<5>e0009413
<6>2013
<7>Lactobacillus plantarum strain ZJ316, a probiotic strain with several functions,  was isolated
from healthy newborn infant fecal samples. Here we report the
finished and annotated genome sequence of this organism.

<>

<1>Li, X., Hu, Y., Gong, J., Lin, Y., Johnstone, L., Rensiing, C., Wang, G.
<2>Genome sequence of the highly efficient arsenite-oxidizing bacterium Achromobacter arsenitoxydans SY8.
<3>J. Bacteriol.
<4>194
<5>1243-1244
<6>2012
<7>We report the draft genome sequence of Achromobacter arsenitoxydans SY8, the first reported
arsenite-oxidizing bacterium belonging to the genus Achromobacter and containing a genomic
arsenic island, an intact type III secretion system, and multiple metal(loid) transporters.
The genome may be helpful to explore the mechanisms intertwining metal(loid) resistance and
pathogenicity.

<>

<1>Li, X., Hu, Y., Gong, J., Lin, Y., Johnstone, L., Rensing, C., Wang, G.
<2>Genome Sequence of the Highly Efficient Arsenite-Oxidizing Bacterium Achromobacter arsenitoxydans SY8.
<3>J. Bacteriol.
<4>194
<5>1243-1244
<6>2012
<7>We report the draft genome sequence of Achromobacter arsenitoxydans SY8, the first reported
arsenite-oxidizing bacterium belonging to the genus Achromobacter
and containing a genomic arsenic island, an intact type III secretion system, and
multiple metal(loid) transporters. The genome may be helpful to explore the
mechanisms intertwining metal(loid) resistance and pathogenicity.

<>

<1>Li, X., Koblizek, M., Feng, F., Li, Y., Jian, J., Zeng, Y.
<2>Whole-Genome Sequence of a Freshwater Aerobic Anoxygenic Phototroph, Porphyrobacter sp. Strain AAP82, Isolated from the Huguangyan Maar Lake in  Southern China.
<3>Genome Announcements
<4>1
<5>e0007213
<6>2013
<7>The Porphyrobacter genus (of the class Alphaproteobacteria) contains aerobic anoxygenic
phototrophic species. Here we report a draft genome sequence of a
freshwater bacterium, Porphyrobacter sp. strain AAP82. It contains a 38-kb-long
photosynthesis gene cluster, but carbon-fixation genes are absent. The presence
of respiratory enzymes, tricarboxylic acid (TCA) cycle, and the Entner-Doudoroff
pathway demonstrates its aerobic photoorganotrophic character.

<>

<1>Li, X., Liu, F., Hu, Y., Mi, K.
<2>Draft Genome Sequence of mc251, a Highly Hydrogen Peroxide-Resistant Mycobacterium smegmatis Mutant Strain.
<3>Genome Announcements
<4>2
<5>e00092-14
<6>2014
<7>Here, we report the draft genome sequence of the Mycobacterium tuberculosis-like
Mycobacterium smegmatis mutant strain, mc(2)51, compared to that of wild-type
strain M. smegmatis mc(2)155. The draft genome sequence comprises 6,823,739 bp,
revealing 6,569 coding DNA sequences (CDSs) and 50 RNA genes (4 rRNA genes and 46
tRNA genes).

<>

<1>Li, X., Song, C., Zhao, M., Li, Y.
<2>Continuous monitoring of restriction endonuclease cleavage activity by universal molecular beacon light quenching coupled with real-time polymerase chain reaction.
<3>Anal. Biochem.
<4>381
<5>1-7
<6>2008
<7>We describe a method for sensitive monitoring of restriction endonuclease kinetics and
activity by use of a universal molecular beacon (U-MB) coupled with real-time polymerase chain
reaction (PCR). The method is used to monitor the progress of DNA cleavage in a sealed
reaction tube and offers more accurate and high-throughput detection. The template has a
universal tail hybridized with the U-MB and the remaining sequence is complementary to one of
the restriction endonuclease digestion products. The U-MB is replaced by the extension of
digested product and the fluorescence quenches. With this concept, one universal fluorescence
probe can be used in different enzyme analytical systems. In the work described here,
homogenous assays were performed with the restriction endonucleases AluI, EcoRI, XhoI, and
SacI at smoothly controlled temperature. Cleavage efficiencies were determined, and the
potential applications of this method were discussed. Furthermore, the AluI and EcoRI cleavage
reactions were monitored online at varying substrate concentrations at the molecular level,
and K(m), V(max), and K(cat) values were calculated. The results suggest that U-MB monitoring
of restriction endonuclease assays based on real-time PCR will be very useful for
high-throughput, sensitive, and precise assays for enzyme activity screening and evolutionary
biotechnology analysis.

<>

<1>Li, X., Yuan, H., Yang, J., Li, B.
<2>Genome Sequence of the Polyphosphate-Accumulating Organism Arthrobacter sp. Strain PAO19 Isolated from Maize Rhizosphere Soil.
<3>Genome Announcements
<4>1
<5>e00566-13
<6>2013
<7>Arthrobacter sp. strain PAO19 is a polyphosphate-accumulating organism isolated from maize
rhizosphere soil. Here we report its genome sequence, which may shed
light on its role in phosphate removal from water bodies. To our knowledge, this
is the first genome announcement of a polyphosphate-accumulating strain of the
genus Arthrobacter.

<>

<1>Li, X.F., Liao, X.Y., Liu, Y.F., Guo, L.Q., Ye, Z.W., Lin, J.F.
<2>Complete Genome Sequence of Probiotic Lactobacillus plantarum Strain FMNP01, Isolated from Mango Fruit.
<3>Genome Announcements
<4>2
<5>e01207-14
<6>2014
<7>Lactobacillus plantarum strain FMNP01 is a new strain with probiotic properties that was
isolated from fresh mango from Guangzhou, China. Here, we report the
complete genome of this organism.

<>

<1>Li, X.R., Gong, F.M., Zheng, H.J., Zhang, Z.H., Luo, Y.Y., Liu, C.J.
<2>Draft Genome Sequence of Lactobacillus plantarum Strain AY01, Isolated from the Raw Material of Fermented Goat Milk Cheese.
<3>Genome Announcements
<4>1
<5>e00737-13
<6>2013
<7>Lactobacillus plantarum is an important probiotic that is isolated mostly from fermented
foods. Here, we report the first draft genome sequence of L. plantarum
strain AY01, isolated from the raw material of fermented goat milk cheese. This
bacterium, with optimum growth at 30 degrees C, has a G+C content of 43.68%.

<>

<1>Li, Y., Cao, B., Zhang, Y., Zhou, J., Yang, B., Wang, L.
<2>Complete Genome Sequence of Staphylococcus aureus T0131, an ST239-MRSA-SCCmec type III Clone isolated in China.
<3>J. Bacteriol.
<4>193
<5>3411-3412
<6>2011
<7>We report here the complete genome sequence of Staphylococcus aureu T0131, which is a
multi-resistant clinical isolate recovered in China and the
first sequenced epidemic ST239-MRSA-SCCmec type III strain obtained in
Asia. Comparison with two published genomes of ST239 reveals the
polymorphism among strains of this type from different continents.

<>

<1>Li, Y., Fu, P., Shi, L.-Y.
<2>Study on a type II restriction endonuclease NcrI.
<3>Chinese Biochem. J.
<4>6
<5>309-313
<6>1990
<7>NcrI isolated from Nocardia carnea C-212 strain is a type II restriction endonuclease.  By
comparing the NcrI digests of lambda DNA with that of the BglII and characterization of the
recognition specificity and its cleavage site, NcrI is identified as an isoschizomer of BglII,
with cleavage site the same as that of BglII at:
5'...A^GATCT...3'
3'...TCTAG^A...5'.

<>

<1>Li, Y., Huang, C.C., Zheng, J.B., Qi, H.L.
<2>Electrogenerated chemiluminescence biosensing method for the detection of DNA methylation and assay of the methyltransferase activity.
<3>Biosensors and Bioelectronics
<4>38
<5>407-410
<6>2012
<7>A novel electrogenerated chemiluminescence (ECL) biosensing method for highly sensitive
detection of DNA methylation and assay of the CpG
methyltransferase (M. Sssl) activity was developed on basis of
enzyme-linkage reactions and ruthenium complex served as an ECL tag.
The ECL biosensing electrode was fabricated by self-assembling 5'-thiol
modified 32-mer single-strand DNA (ss-DNA)-tagged with ruthenium bis
(2,2'-bipyridine) (2,2'-bipyridine-4,4'-dicarboxylic
acid)-ethylenediamine on the surface of a gold electrode, and then
hybridized with complementary ss-DNA to form duplex DNA (ds-DNA). When
M. Sssl and S-adenosylmethionine were introduced, all cytosine residues
within 5'-CG-3' of ds-DNA on the biosensing electrode were methylated.
After the methylated biosensing electrode was treated by HpaII
endonuclease, the un-methylated cytosines were cleaved, thus led to
decrease ECL signal. The ECL intensity of ECL biosensing electrode is
related to the methylation level and M. Sssl activity in a fixed
concentration HpaII endonuclease. The increased ECL intensity was
direct proportion to M. Sssl activity in the range from 0.05 to 100
U/mL with a detection limit of 0.02 U/mL. This work demonstrates that
the combination of the enzyme-linkage reactions with a highly sensitive
ECL technique is a great promising approach for the detection of DNA
methylation level, assay of the activity of MTase, and evaluation of
the capability of inhibitors for the methyltransferase.

<>

<1>Li, Y., Liu, Y., Zhou, Z., Huang, H., Ren, Y., Zhang, Y., Li, G., Zhou, Z., Wang, L.
<2>Complete Genome Sequence of Aeromonas Veronii Strain B565.
<3>J. Bacteriol.
<4>193
<5>3389-3390
<6>2011
<7>Aeromonas veronii strain B565 was isolated from aquaculture pond sediment in China. We present
here the complete genome sequence of B565 and compare
it with 2 published genome sequences of pathogenic strains in Aeromonas
genus. The result represents an independent step-wise acquisition of
virulence factors of pathogenic strains in this genus.

<>

<1>Li, Y., Lu, Z., Sun, L., Ropp, S., Kutish, G.F., Rock, D.L., Van Etten, J.L.
<2>Analysis of 74 kb of DNA located at the right end of the 330-kb chlorella virus PBCV-1 genome.
<3>Virology
<4>237
<5>360-377
<6>1997
<7>This report completes a preliminary analysis of the sequence of the 330,740-bp chlorella virus
PBCV-1 genome, the largest virus genome to be sequenced to date.  The PBCV-1 genome is 57% the
size of the genome from the smallest self-replicating organism, Mycoplasma genitalium,
Analysis of 74 kb of newly sequenced DNA, from the right terminus of the PBCV-1 genome,
revealed 153 open reading frames of 65 codons or longer.  Eighty-five of these ORFs, which are
evenly distributed on both strands of the DNA, were considered major ORFs.  Fifty-nine of the
major ORFs were separated by less than 100 bp.  The largest intergenic distance was 729 bp,
which occurred between two ORFs located in the 2.2-kb inverted terminal repeat region of the
PBCV-1 genome.  Twenty-seven of the 85 major ORFs resemble proteins in databases, including
the large subunit of ribonucleotide diphosphate reductase, ATP-dependent DNA ligase, type II
DNA topoisomerase, a helicase, histidine decarboxylase, dCMP deaminase, dUTP pyrophosphatase,
proliferating cell nuclear antigen, a transposase, fungal translation elongation factor 3, UDP
glucose dehydrogenase, a protein kinase, and an adeneine DNA methyltransferase and its
corresponding DNA site-specific endonuclease.  Seventeen of the 153 ORFs resembled other
PBCV-1 ORFs, suggesting that they represent either gene duplications or gene families.

<>

<1>Li, Y., Ng, I.S., Zhang, X., Wang, N.
<2>Draft Genome Sequence of the Dye-Decolorizing and Nanowire-Producing Bacterium Shewanella xiamenensis BC01.
<3>Genome Announcements
<4>2
<5>e00721-14
<6>2014
<7>Shewanella xiamenensis BC01 is an important biodecolorizing and nanowire-producing bacterium
which was found in Xiamen, China. Here, we present
the draft genome sequence consisting of 4,677,169 bp (GC content, 46.21%) and
3,999 coded proteins. This information boosts insight into and understanding of
the genetic evolution of Shewanella species.

<>

<1>Li, Y., Yan, Z., Zheng, J., Qi, H.
<2>Label-free and amplified electrogenerated chemiluminescence biosensing method for the determination of DNA methyltransferase activity using signal reagent-assembled graphene oxide.
<3>Electrochim. Acta.
<4>137
<5>454-461
<6>2014
<7>A novel label-free electrogenerated chemiluminescence (ECL) biosensing method for the
determination of DNA methyltransferase (MTase) activity was developed on base of
enzyme-linkage reactions and tris(1, 10-phenanthroline) ruthenium-assembled graphene oxide
(GO) served as an ECL signal compound. The ECL biosensing electrode was fabricated by
self-assembling 5'-thiol modified hairpin-capture DNA probe containing methylation
recognition site 5'-GATC-3' on the surface of a gold electrode. When DNA adenine methylation
(Dam) MTase and S-adenosyl-L-methionine were introduced, all adenine residues within
5'-GATC-3' of hairpin-capture DNA probe on the biosensing electrode were methylated. After
the methylated biosensing electrode was treated by the methylation-sensitive restriction
endonuclease Dpn I, the methylated adenines were cleaved, methylation-induced scission of
hairpin-capture DNA probe would displace the hairpin section and remain the 'capture DNA
probe' section on the gold electrode, then a long ssDNA was immobilized via the partial
hybridization reaction between long ssDNA and hairpin-capture DNA probe remained section, the
more binding site allow tris(1, 10-phenanthroline) ruthenium-assembled GO to be more bound to
the long ssDNA on the electrode surface through both hydrophobic and pi-pi stacking
interaction, in conjunction with the generation of a increased ECL signal. The ECL intensity
versus the concentration of Dam MTase was linear in the range from 0.02 unit/mL to 10 unit/mL.
The detection limit was 0.01 unit/mL. This work demonstrates that using the different
affinities of GO for ssDNA and dsDNA for the fabrication of the label-free ECL biosensing
method for DNA MTase activity is promising approach.

<>

<1>Li, Y., Zheng, H., Liu, Y., Jiang, Y., Xin, J., Chen, W., Song, Z.
<2>The Complete Genome Sequence of Mycoplasma bovis Strain Hubei-1.
<3>PLoS ONE
<4>6
<5>e20999
<6>2011
<7>Infection by Mycoplasma bovis (M. bovis) can induce diseases, such as pneumonia and otitis
media in young calves and
mastitis and arthritis in older animals. Here, we report the finished and annotated genome
sequence of M. bovis strain
Hubei-1, a strain isolated in 2008 that caused calf pneumonia on a Chinese farm. The genome of
M. bovis strain Hubei-1
contains a single circular chromosome of 953,114 bp with a 29.37% GC content. We identified
803 open reading frames
(ORFs) that occupy 89.5% of the genome. While 34 ORFs were Hubei-1 specific, 662 ORFs had
orthologs in the M. bovis type
strain PG45 genome. Genome analysis validated lateral gene transfer between M. bovis and the
Mycoplasma mycoides
subspecies mycoides, while phylogenetic analysis found that the closest M. bovis neighbor is
Mycoplasma agalactiae.
Glycerol may be the main carbon and energy source of M. bovis, and most of the biosynthesis
pathways were incomplete.
We report that 47 lipoproteins, 12 extracellular proteins and 18 transmembrane proteins are
phase-variable and may help M.
bovis escape the immune response. Besides lipoproteins and phase-variable proteins, genomic
analysis found two possible
pathogenicity islands, which consist of four genes and 11 genes each, and several other
virulence factors including
hemolysin, lipoate protein ligase, dihydrolipoamide dehydrogenase, extracellular cysteine
protease and 59-nucleotidase.

<>

<1>Li, Y., Zhu, H., Li, C., Zhang, H., Chen, Z., Zheng, W., Xu, H., Zheng, T.
<2>Draft Genome Sequence of the Algicidal Bacterium Mangrovimonas yunxiaonensis Strain LY01.
<3>Genome Announcements
<4>2
<5>e01234-14
<6>2014
<7>Mangrovimonas yunxiaonensis LY01, a novel bacterium isolated from mangrove sediment, showed
high algicidal effects on harmful algal blooms of Alexandrium
tamarense. Here, we present the first draft genome sequence of this strain to
further understanding of the functional genes related to algicidal activity.

<>

<1>Li, Y.Q., Zhou, P.Z., Zheng, X.D., Walsh, C.P., Xu, G.L.
<2>Association of Dnmt3a and thymine DNA glycosylase links DNA methylation with base-excision repair.
<3>Nucleic Acids Res.
<4>35
<5>390-400
<6>2007
<7>While methylcytosines serve as the fifth base encoding epigenetic information, they are also a
dangerous endogenous mutagen due to their
intrinsic instability. Methylcytosine undergoes spontaneous deamination,
at a rate much higher than cytosine, to generate thymine. In mammals, two
repair enzymes, thymine DNA glycosylase (TDG) and methyl-CpG binding
domain 4 (MBD4), have evolved to counteract the mutagenic effect of
methylcytosines. Both recognize G/T mismatches arising from methylcytosine
deamination and initiate base-excision repair that corrects them to G/C
pairs. However, the mechanism by which the methylation status of the
repaired cytosines is restored has remained unknown. We show here that the
DNA methyltransferase Dnmt3a interacts with TDG. Both the PWWP domain and
the catalytic domain of Dnmt3a are able to mediate the interaction with
TDG at its N-terminus. The interaction affects the enzymatic activity of
both proteins: Dnmt3a positively regulates the glycosylase activity of
TDG, while TDG inhibits the methylation activity of Dnmt3a in vitro. These
data suggest a mechanistic link between DNA repair and remethylation at
sites affected by methylcytosine deamination.

<>

<1>Li, Z., Chen, H., Chen, X., Zhou, T., Zhao, L., Zhang, C., Jin, W.
<2>Genome Sequence of the Human-Pathogenic Bacterium Vibrio vulnificus Type Strain ATCC 27562.
<3>J. Bacteriol.
<4>194
<5>6954-6955
<6>2012
<7>Vibrio vulnificus, which is the causative agent of cholera, is a Gram-negative, curved,
motile, and rod-shaped bacterium. Here, we present the draft genome
sequence of the type strain, ATCC 27562, which was the first isolated Vibrio
vulnificus strain.

<>

<1>Li, Z., Ma, Z., Hao, X., Wei, G.
<2>Draft Genome Sequence of Sinorhizobium meliloti CCNWSX0020, a Nitrogen-Fixing Symbiont with Copper Tolerance Capability Isolated from Lead-Zinc Mine Tailings.
<3>J. Bacteriol.
<4>194
<5>1267-1268
<6>2012
<7>Sinorhizobium meliloti CCNWSX0020 was isolated from Medicago lupulina plants growing in
lead-zinc mine tailings, which can establish a symbiotic relationship
with Medicago species. Also, the genome of this bacterium contains a number of
protein-coding sequences related to metal tolerance. We anticipate that the
genomic sequence provides valuable information to explore environmental
bioremediation.

<>

<1>Li, Z., Wu, S., Bai, X., Liu, Y., Lu, J., Liu, Y., Xiao, B., Lu, X., Fan, L.
<2>Genome Sequence of the Tobacco Bacterial Wilt Pathogen Ralstonia solanacearum.
<3>J. Bacteriol.
<4>193
<5>6088-6089
<6>2011
<7>Ralstonia solanacearum is a causal agent of plant bacterial wilt with thousands of distinct
strains in a heterogeneous species complex. Here we
report the genome sequence of a phylotype IB strain, Y45, isolated from
tobacco (Nicotiana tabacum) in China. Compared with the published genomes
of eight strains which were isolated from other hosts and habitats, 794
specific genes and many rearrangements/inversion events were identified in
the tobacco strain, demonstrating that this strain represents an important
node within the R. solanacearum complex.

<>

<1>Li, Z.F., Li, X., Liu, H., Liu, X., Han, K., Wu, Z.H., Hu, W., Li, F.F., Li, Y.Z.
<2>Genome Sequence of the Halotolerant Marine Bacterium Myxococcus fulvus HW-1.
<3>J. Bacteriol.
<4>193
<5>5015-5016
<6>2011
<7>Myxococcus fulvus HW-1 (ATCC BAA-855) is a halotolerant marine myxobacterium. This strain
exhibits complex social behaviors in the
presence of low concentrations of seawater but adopts an asocial living
pattern under oceanic conditions. The whole genome of M. fulvus HW-1 will
enable us to further investigate the details of its evolution.

<>

<1>Liang, G., Chan, M.F., Tomigahara, Y., Tsai, Y.C., Gonzales, F.A., Li, E., Laird, P.W., Jones, P.A.
<2>Cooperativity between DNA methyltransferases in the maintenance methylation of repetitive elements.
<3>Mol. Cell. Biol.
<4>22
<5>480-491
<6>2002
<7>We used mouse embryonic stem (ES) cells with systematic gene knockouts for DNA
methyltransferases to delineate the roles of DNA methyltransferase 1 (Dnmt1) and Dnmt3a and
Dnmt3b in maintaining methylation patterns in the mouse genome. Dnmt1 alone was able to
maintain
methylation of most CpG-poor regions analyzed. In contrast, both Dnmt1 and Dnmt3a and/or
Dnmt3b were required for methylation of a select class of sequences which included abundant
murine LINE-1 promoters. We used a novel hemimethylation assay to show that even in wild-type
cells these sequences contain high levels of hemimethylated DNA, suggestive of poor
maintenance methylation. We showed that Dnmt3a and/or -3b could restore methylation of these
sequences to pretreatment levels following transient exposure of cells to 5-aza-CdR, whereas
Dnmt1 by itself could not. We conclude that ongoing de novo methylation by Dnmt3a and/or
Dnmt3b compensates for inefficient maintenance methylation by Dnmt1 of these endogenous
repetitive sequences. Our results reveal a previously unrecognized degree of cooperativity
among mammalian DNA methyltransferases in ES cells.

<>

<1>Liang, J., Wang, Z., He, X., Li, J., Zhou, X., Deng, Z.
<2>DNA modification by sulfur: analysis of the sequence recognition specificity surrounding the modification sites.
<3>Nucleic Acids Res.
<4>35
<5>2944-2954
<6>2007
<7>The Dnd (DNA degradation) phenotype, reflecting a novel DNA modification by sulfur in
Streptomyces lividans 1326, was strongly aggravated when one (dndB) of
the five genes (dndABCDE) controlling it was mutated. Electrophoretic banding
patterns of a plasmid (pHZ209), reflecting DNA degradation, displayed a clear
change from a preferential modification site in strain 1326 to more random
modifications in the mutant. Fourteen randomly modifiable sites on pHZ209 were
localized, and each seemed to be able to be modified only once. Residues in a
region (5'-c-cGGCCgccg-3') including a highly conserved 4-bp central core
(5'-GGCC-3') in a well-documented preferential modification site were assessed
for their necessity by site-directed mutagenesis. While the central core (GGCC)
was found to be stringently required in 1326 and in the mutant, 'gccg' flanking
its right could either abolish or reduce the modification frequency only in the
mutant, and two separate nucleotides to the left had no dramatic effect. The lack
of essentiality of DndB for S-modification suggests that it might only be
required for enhancing or stabilizing the activity of a protein complex at the
required preferential modification site, or resolving secondary structures
flanking the modifiable site(s), known to constitute an obstacle for efficient
modification.

<>

<1>Liang, J.X., Blumenthal, R.M.
<2>Naturally-occurring, dually-functional fusions between restriction endonucleases and regulatory proteins.
<3>BMC Evol. Biol.
<4>13
<5>0
<6>2013
<7>Background: Restriction-modification (RM) systems appear to play key roles in modulating gene
flow among bacteria and archaea. Because the
restriction endonuclease (REase) is potentially lethal to unmethylated
new host cells, regulation to ensure pre-expression of the protective
DNA methyltransferase (MTase) is essential to the spread of RM genes.
This is particularly true for Type IIP RM systems, in which the REase
and MTase are separate, independently-active proteins. A substantial
subset of Type IIP RM systems are controlled by an activator-repressor
called C protein. In these systems, C controls the promoter for its own
gene, and for the downstream REase gene that lacks its own promoter.
Thus MTase is expressed immediately after the RM genes enter a new
cell, while expression of REase is delayed until sufficient C protein
accumulates. To study the variation in and evolution of this regulatory
mechanism, we searched for RM systems closely related to the
well-studied C protein-dependent PvuII RM system. Unexpectedly, among
those found were several in which the C protein and REase genes were
fused.
Results: The gene for CR. NsoJS138I fusion protein (nsoJS138ICR,
from the bacterium Niabella soli) was cloned, and the fusion protein
produced and partially purified. Western blots provided no evidence
that, under the conditions tested, anything other than full-length
fusion protein is produced. This protein had REase activity in vitro
and, as expected from the sequence similarity, its specificity was
indistinguishable from that for PvuII REase, though the optimal
reaction conditions were different. Furthermore, the fusion was active
as a C protein, as revealed by in vivo activation of a lacZ reporter
fusion to the promoter region for the nsoJS138ICR gene.
Conclusions: Fusions between C proteins and REases have not
previously been characterized, though other fusions have (such as
between REases and MTases). These results reinforce the evidence for
impressive modularity among RM system proteins, and raise important
questions about the implications of the C-REase fusions on expression
kinetics of these RM systems.

<>

<1>Liang, R., Liu, H., Tao, F., Liu, Y., Ma, C., Liu, X., Liu, J.
<2>Genome Sequence of Pseudomonas putida Strain SJTE-1, a Bacterium Capable of Degrading Estrogens and Persistent Organic Pollutants.
<3>J. Bacteriol.
<4>194
<5>4781-4782
<6>2012
<7>Pseudomonas putida strain SJTE-1 can utilize 17beta-estradiol and other environmental
estrogens/toxicants, such as estrone, and naphthalene as sole
carbon sources. We report the draft genome sequence of strain SJTE-1 (5,551,505
bp, with a GC content of 62.25%) and major findings from its annotation, which
could provide insights into its biodegradation mechanisms.

<>

<1>Liang, X., Jensen, K., Frank-Kamenetskii, M.D.
<2>Acceleration of de novo DNA synthesis by restriction enzymes.
<3>J. Biomol. Struct. Dyn.
<4>22
<5>766-767
<6>2005
<7>We have found that in the presence of a restriction endonuclease, DNA polymerase efficiently
synthesizes and amplifies DNA in the absence of any added template and primer nucleic acid
under isothermal conditions.  In the presence of restriction endonuclease Tsp509I (recognition
sequence /AATT) or TspRI (recognition sequence: NNCAG(C)TGNN/), more than 90% of the available
dNTPs can be consumed by Vent (exo-) DNA polymerase in 1 h at 70oC.  The synthesized DNA has a
highly repetitive palindromic sequence containing recognition sites for the restriction
enzyme, e.g. (AAAAATTTTT)n and (ATACACTGTATATACAGTG-TAT)n.  Our data show that the high
efficiency of the restriction-endonuclease-DNA-polymerase (RE-pol) DNA synthesis results from
an efficient exponential amplification involving digestion-elongation cycles.

<>

<1>Liang, X.G., Jensen, K., Frank-Kamenetskii, M.D.
<2>Very efficient template/primer-independent DNA synthesis by thermophilic DNA polymerase in the presence of a thermophilic  restriction endonuclease.
<3>Biochemistry
<4>43
<5>13459-13466
<6>2004
<7>We have found that, in the presence of a thermophilic restriction endonuclease, thermophilic
DNA polymerase efficiently synthesizes and
amplifies DNA in the absence of any added template and primer nucleic
acid under isothermal conditions. More than 10 micrograms of DNA can be
synthesized by 1 unit of DNA polymerase in 1 h, and the reaction
proceeds until available dNTPs are consumed. We used mostly the Tsp509I
restriction endonuclease (recognition sequence: ^AATT), the
TspRI restriction endonuclease (recognition sequence: NNCA(G/C)TGNN^), and Vent (exo(-)) and
Vent DNA polymerase. The synthesized
double-stranded DNA has a highly repetitive palindromic sequence, e.g.
(AAAAATTTTT)(n) and (ATACACTGTATATACAGTGTAT)(n). In every repeating
unit, there are one or two recognition sites for the restriction
enzyme. Our data show that the high efficiency of the
restriction-endonuclease-DNA-polymerase (RE-pol) DNA synthesis results
from an efficient exponential amplification involving
digestion-elongation cycles: a longer DNA with numerous recognition
sites for the restriction enzyme is digested to short fragments, and
the short fragments are used as seeds for elongation to synthesize
longer DNA. A possible role of RE-pol DNA synthesis in the evolutionary
development of genetic materials is briefly discussed.

<>

<1>Liao, T.L., Lin, A.C., Chen, E., Huang, T.W., Liu, Y.M., Chang, Y.H., Lai, J.F., Lauderdale, T.L., Wang, J.T., Chang, S.C., Tsai, S.F., Chen, Y.T.
<2>Complete Genome Sequence of Klebsiella oxytoca E718, a New Delhi Metallo-beta-Lactamase-1-Producing Nosocomial Strain.
<3>J. Bacteriol.
<4>194
<5>5454
<6>2012
<7>We report the complete genome sequence of Klebsiella oxytoca E718, a New Delhi
metallo-beta-lactamase-1 (NDM-1)-producing strain isolated from a renal
transplant patient. The genome contains a 6,097,032-bp chromosome and two
multidrug resistance plasmids with sizes of 324,906 bp and 110,781 bp.

<>

<1>Liao, Y.C., Chen, Y.H., Lin, H.H., Mu, J.J., Wu, H.S., Chen, F.C., Hsiung, C.A.
<2>Draft Genome Sequence of NDM-1-Producing Klebsiella pneumoniae Clinical Isolate 303K.
<3>Genome Announcements
<4>1
<5>e01069-13
<6>2013
<7>Multidrug-resistant New Delhi metallo-beta-lactamase 1 (NDM-1)-producing bacteria have spread
globally and become a major clinical and public health threat. We
report here the draft genome sequence of the Klebsiella pneumoniae clinical
isolate 303K, harboring an NDM-1 coding sequence.

<>

<1>Liao, Y.C., Chen, Y.Y., Lin, H.H., Chang, J.R., Su, I.J., Huang, T.S., Dou, H.Y.
<2>Draft Genome Sequence of the Mycobacterium tuberculosis Clinical Isolate C2, Belonging to the Latin American-Mediterranean Family.
<3>Genome Announcements
<4>2
<5>e00536-14
<6>2014
<7>Tuberculosis remains a major infectious disease in Taiwan. Here we present the draft genome
sequence of the Mycobacterium tuberculosis C2 strain, belonging to
the Latin American-Mediterranean lineage. The draft genome sequence comprises
4,453,307 bp with a G+C content of 65.6%, revealing 4,390 coding genes and 45
tRNA genes.

<>

<1>Liao, Y.C., Chen, Y.Y., Lin, H.H., Chang, J.R., Su, I.J., Huang, T.S., Dou, H.Y.
<2>Draft Genome Sequences of the Mycobacterium tuberculosis Clinical Strains A2 and  A4, Isolated from a Relapse Patient in Taiwan.
<3>Genome Announcements
<4>2
<5>e00672-14
<6>2014
<7>The recurrence rate of Mycobacterium tuberculosis in Taiwan is 3%. Here, we present the draft
genome sequences of M. tuberculosis strains A2 and A4 from a
relapse patient. The draft genome sequences comprise 4,443,031 bp and 4,487,096
bp, revealing 4,220 and 4,143 coding sequences for A2 and A4, respectively, as
well as 49 tRNA genes for the both isolates.

<>

<1>Lieb, M.
<2>Bacterial genes mutL, mutS, and dcm participate in repair of mismatches at 5-methylcytosine sites.
<3>J. Bacteriol.
<4>169
<5>5241-5246
<6>1987
<7>Certain amber mutationsn in the cI gene of bacteriophage lambda appear to recombine very
frequently with nearby mutations. The aberrant mutations included C-to-T transitiions at the
second cytosine in 5'CC(A/T)GG sequences (which are subject to methylation by bacterial
cytosine methylase) and in 5'CCAG and 5'CAGG sequences. Excess cI+ recombinants arising in
crosses that utilize these mutations are attributable to the correction of mismatches by a
bacterial very-short-patch (VSP) mismatch repair system. In the present study I found that two
genes required for methyladenine-directed (long-patch) mismatch repair, mutL and mutS, also
functioned in VSP mismatch repair; mutH and mutU (uvrD) were dispensable. VSP mismatch repair
was greatly reduced in a dcm Escherichia coli mutant, in which 5-methylcytosine was not
methylated. However, mismatches in heterduplexes prepared from lambda DNA lacking
5-methylcytosine were repaired in dcm+ bacteria. These results indicate that the product of
gene dcm has a repair function in addition to its methylase activity.

<>

<1>Lieb, M.
<2>Specific mismatch correction in bacteriophage lambda crosses by very short patch repair.
<3>Mol. Gen. Genet.
<4>191
<5>118-125
<6>1983
<7>In crosses under rec+, red+, gam+ conditions, mutation am6 in the cI
(repressor) gene of bacteriophage lambda recombines with other cI mutations
much more frequently than predicted by the physical distances involved.  In
four-factor crosses of am6 with mutations located 22-60 base pairs to the left,
cI+ recombinants that are expected to require three crossovers (triple
recombinants) are more frequent than recombinants that require only one
crossover.  However, when am6 is crossed with large insertions in cI, which may
be expected to interfere with the formation of heteroduplexes by branch
migration, the frequency of cI+ triple recombinants is very low.  In addition,
cI+ recombinants in crosses between am6 and adjacent mutations have a high
probability of retaining the flanking markers of the am6 parent.  These
findings suggest that am6 is particularly susceptible to mismatch repair in
heteroduplexes spanning cI.  A large fraction of such heteroduplexes are
presumed to be the result of branch migration from crossovers occuring at some
distance from am6.  The absence of co-repair when am6 is crossed with adjacent
cI mutations indicates that most repair tracts extend no farther than about 20
bp to either side of the mismatch.  The am6 mutation arose in the glutamine
codon in a CCAGG sequence, in which the central cytosines are methylated in K12
strains.  Their location in methylated sequences may make certain amber
mutations susceptible to a specific very short patch (VSP) repair.

<>

<1>Lieb, M.
<2>Spontaneous Mutation at a 5-methylcytosine hotspot is prevented by very short patch (VSP) mismatch repair.
<3>Genetics
<4>128
<5>23-27
<6>1991
<7>In many strains of Escherichia coli, the product of gene dcm methylates the internal cytosines
in the sequence 5'CC(A or T)GG. Spontaneous deamination of 5-methylcytosine produces thymine
which, if not corrected, can result in a transition mutation. 5-Methylcytosines in the lacI
gene are hotspots for spontaneous C to T mutations. dcm is linked to vsr, a gene required for
very short patch (VSP) repair. VSP respir corrects T.G mispairs in the following contexts:
CTAGG/GGTCC, CTTGG/GGACC, TAGG/GTCC and CTAG/GGTC. I have investigated the relationships
between cytosine methylation, mutation, and VSP repair. Spontaneous mutations in the repressor
(cI) gene of lambda prophage were isolated in wild-type and mutant lysogens. A hotspot for
spontaneous mutation that corresponds with a 5-methylcytosine was observed in wild-type
lysogens but was not present in bacteria lacking both methylase and VSP repair activity.
Introduction of a plasmid containg dcm+ and vsr+ restored the mutation hotspot. If the added
plasmid carried only dcm+, the frequency of spontaneous mutations at the 5-methylcytosine was
over 10-fold higher than in Dcm+Vsr+ lysogens. The addition of vsr on a plasmid to a wild-type
lysogen resulted in a 4-fold reduciton in mutation at the hotspot. These findings support the
previously untested hypothesis that VSP repair prevents mutations resulting from deamination
of 5-methylcytosine.

<>

<1>Lieb, M., Allen, E., Read, D.
<2>Very short patch mismatch repair in phage lambda: repair sites and length of repair tracts.
<3>Genetics
<4>114
<5>1041-1060
<6>1986
<7>Five amber mutations in the repressor (cI) gene of bacteriophage lambda recombine anomalously
with nearby cI mutations.  When any of these markers is used in four-factor crosses, cI+
recombinants that are expected to require three crossovers occur at high frequencies.  These
recombinants are attributable to very-short-patch (VSP) repair of specific mismatches in DNA
heteroduplexes formed during recombination between the markers flanking cI.  The sites of the
repair-prone mutations and the lengths of repair tracts have now been determined.  Amber
mutations subject to VSP repair are C to T transitions in 5'CCA/TGG, the sequence methylated
by the product of gene dcm, and also in the related 5'CAGG or 5'CCAG sequences.  Ambers
arising in CAG sequences found in other contexts, or in codons other than CAG, were not
subject to VSP repair.  Rapair tracts rarely, if ever, exceed ten nucleotides in length, and
can be as short as two nucleotides.  A repair-prone mutation does not stimulate recombination
between flanking cI markers.

<>

<1>Lieb, M., Bhagwat, A.S.
<2>Very short patch mismatch repair activity associated with gene dcm is not conferred by a plasmid coding for EcoRII methylase.
<3>J. Bacteriol.
<4>170
<5>4967-4968
<6>1988
<7>The only cytosine methylase in Escherichia coli K-12 methylates the second cytosine in the
sequence CC(A/T)GG and is encoded by gene dcm. Methylation and very short patch mismatch
repair activities lacking in a dcm mutant of E. coli were restored by a plasmid containing the
cloned dcm gene. In contrast, plasmids with the gene for EcoRII methylase, which is a homolog
of dcm, restored only cytosine methylase activity and not mismatch repair.

<>

<1>Lieb, M., Bhagwat, A.S.
<2>Very short patch repair: reducing the cost of cytosine methylation.
<3>Mol. Microbiol.
<4>20
<5>467-473
<6>1996
<7>In Escherichia coli and related bacteria, the product of gene dcm methylates the second
cytosine of 5'-CCWGG sequences (where W is A or T).  Deamination of 5-methylcytosine results
in C to T mutations.  The mutagenic potential of 5meC is reduced by a system called very short
patch repair, which replaces T with C.  T:G and U:G mispairs in the methylatable sequence and
in related sequences are recognized by the product of vsr, a gene adjacent to dcm.  Vsr
creates a nick just 5' of the mispaired pyrimidine to initiate the repair.  Additional
products known to be required for VSP repair are DNA polymerase I and DNA ligase.  MutS and
MutL have a stimulatory role but are not required.  The ability of Vsr to recognize T:G
mispairs in sequences related to CCWGG is probably responsible for over- and
under-representation of certain tetranucleotides in the E. coli genome.  Although VSP repair
reduces spontaneous mutations at 5meCs in replicating bacteria, mutation hot-spots persist at
these sites. Under conditions that more accurately mimic the natural environment of E. coli,
VSP repair appears to be effective in preventing mutation at 5meC.

<>

<1>Lieb, M., Rehmat, S.
<2>Very short patch repair of T:G mismatches in vivo: importance of context and accessory proteins.
<3>J. Bacteriol.
<4>177
<5>660-666
<6>1995
<7>In Escherichia coli, T:G mismatches in specific contexts are corrected by a very short patch
(VSP) repair system. Previous studies have shown that the product of gene vsr mediates
correction of T:G to C:G in the 5'CTAGG/3'GGTCC context and in some related contexts. Amber
mutations that arose in CAG sequences in gene cI of bacteriophage lambda were used to
determine the effect of flanking bases on the repair of T:G mispairs arising during phage
recombination. The experimental findings were combined with published data on mismatch repair
of mutations in lambda gene P and E. coli gene lacI. While VSP repair was most efficient in
the context 5'CTAGG, there was very significant correction when either the 5' C or the 3' G
was replaced by another base. Some mismatch repair of TAG to CAG occurred in all contexts
tested. Reduction in VSP repair caused by the lack of MutL or MutS was fully complemented by
the addition of vsr+ plasmids when the T:G mispair was in the 5'CTAGG/3'GGTCC context. VSP
repair was decreased in bacteria containing mutS+ on a multicopy plasmid. It is suggested that
VSP repair maintains sequences such as the repetitive extragenic palindromic (REP) and Chi
sequences, which have important roles in E. coli and closely related bacteria.

<>

<1>Lieb, M., Rehmat, S.
<2>5-Methylcytosine is not a mutation hot spot in nondividing Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>940-945
<6>1997
<7>Spontaneous deamination of 5-methylcytosine (5meC) causes hot spots of C.G-T.A mutations in
Escherichia coli and in human cells.  In E. coli, the resulting T.G mispairs can be corrected
to C.G by very short patch (VSP) repair, which requires the product of gene vsr.  Mutation hot
spots in genes of replicating vsr+ bacteria are attributable to low Vsr activity.  To
determine the rate of deamination of 5meC and the efficiency of VSP repair in nondividing
bacteria, we used kanamycin-sensitive (KanS) lysogens containing a lambda kan- prophage.
Deamination of a 5meC in the kan- gene resulted in mutation to kanamycin resistance (KanR).
Lysogens containing a single lambda kan- prophage per bacterial genome were grown in synthetic
medium with limiting amino acids and stored at 15oC or 37oC.  In the absence of VSP repair,
KanR mutants accumulated at the rate of approximately 1.3 x 10^-7 per bacterium per day at
37oC.  This is similar to the 5meC--T mutation rate reported for DNA in solution.  In vsr+
bacteria, the KanR accumulation rate was 3 x 10^-9 per bacterium per day, which is not
significantly higher than the rate observed when the target cytosine was unmethylated.  The
increase in KanR mutants was barely detectable in vsr+ cultures stored at 15oC for 4 months.
It is likely that mutation hot spots at 5meC in rapidly dividing cells are attributable to
insufficient time for T.G correction in the interval between deamination of 5meC and
subsequent DNA replication.  DNA synthesis occurred in bacteria starved for amino acids and
this synthesis was not highly mutagenic.

<>

<1>Lieb, M., Rehmat, S., Bhagwat, A.S.
<2>Interaction of MutS and Vsr: Some Dominant-Negative mutS Mutations That Disable Methyladenine-Directed Mismatch Repair Are Active in Very-Short-Patch Repair.
<3>J. Bacteriol.
<4>183
<5>6487-6490
<6>2001
<7>In Escherichia coli and related bacteria, the very-short-patch (VSP) repair pathway uses an
endonuclease, Vsr, to correct T.G mismatches that result from the deamination of
5-methylcytosines in DNA to C.G. The products of mutS and mutL, which are required for
adenine methylation-directed mismatch repair (MMR), enhance VSP repair. Multicopy plasmids
carrying mutS alleles that are dominant negative for MMR were tested for their effects on VSP
repair. Some mutS mutations (class I) did not lower VSP repair in a mutS(+) background, and
most class I mutations increased VSP repair in mutS cells more than plasmids containing
mutS(+). Other plasmid-borne mutS mutations (class II) and mutS(+) decreased VSP repair in the
mutS(+) background. Thus, MutS protein lacking functions required for MMR can still
participate in VSP repair, and our results are consistent with a model in which MutS binds
transiently to the mispair and then translocates away from the mispair to create a specialized
structure that enhances the binding of Vsr.

<>

<1>Liebert, K., Hermann, A., Schlickenrieder, M., Jeltsch, A.
<2>Stopped-flow and mutational analysis of base flipping by the Escherichia coli dam DNA-(adenine-N6)-methyltransferase.
<3>J. Mol. Biol.
<4>341
<5>443-454
<6>2004
<7>By stopped-flow kinetics using 2-aminopurine as a probe to detect base flipping, we show here
that base flipping by the Escherichia coli Dam
DNA-(adenine-N6)-methyltransferase (MTase) is a biphasic process:
target base flipping is very fast (k(flip) > 240 s(-1)), but binding of
the flipped base into the active site pocket of the enzyme is slow (k =
0.1-2 s(-1)). Whereas base flipping occurs in the absence of
S-adenosyl-L-methionine (AdoMet), binding of the target base in the
active site pocket requires AdoMet. Our data suggest that the tyrosine
residue in the DPPY motif conserved in the active site of
DNA-(adenine-N6)-MTases stacks to the flipped target base. Substitution
of the aspartic acid residue of the DPPY motif by alanine abolished
base flipping, suggesting that this residue contacts and stabilizes the
flipped base. The exchange of Ser188 located in a loop next to the
active center by alanine led to a seven- to eightfold reduction of
k(flip), which was also reduced with substrates having altered GATC
recognition sites and in the absence of AdoMet. These findings provide
evidence that the enzyme actively initiates base flipping by
stabilizing the transition state of the process. Reduced rates of base
flipping in substrates containing the target base in a non-canonical
sequence demonstrate that DNA recognition by the MTase starts before
base flipping. DNA recognition, cofactor binding and base flipping are
correlated and efficient base flipping takes place only if the enzyme
has bound to a cognate target site and AdoMet is available.

<>

<1>Liebert, K., Horton, J.R., Chahar, S., Orwick, M., Cheng, X.D., Jeltsch, A.
<2>Two alternative conformations of S-Adenosyl-L-homocysteine bound to Escherichia coli DNA adenine methyltransferase and the implication of conformational changes in regulating the catalytic cycle.
<3>J. Biol. Chem.
<4>282
<5>22848-22855
<6>2007
<7>The crystal structure of the Escherichia coli DNA adenine methyltransferase( EcoDam) in a
binary complex with the cofactor
product S-adenosyl-L-homocysteine ( AdoHcy) unexpectedly showed the
bound AdoHcy in two alternative conformations, extended or folded. The
extended conformation represents the catalytically competent
conformation, identical to that of EcoDam-DNA-AdoHcy ternary complex.
The folded conformation prevents catalysis, because the homocysteine
moiety occupies the target Ade binding pocket. The largest difference
between the binary and ternary structures is in the conformation of the
N-terminal hexapeptide ( (9)KWAGGK(14)). Cofactor binding leads to a
strong change in the fluorescence of Trp(10), whose indole ring
approaches the cofactor by 3.3 angstrom. Stopped-flow kinetics
andAdoMetcross-linking studies indicate that the cofactor prefers
binding to the enzyme after preincubation with DNA. In the presence of
DNA, AdoMet binding is similar to 2-fold stronger than AdoHcy binding.
In the binary complex the side chain of Lys(14) is disordered, whereas
Lys(14) stabilizes the active site in the ternary complex. Fluorescence
stopped-flow experiments indicate that Lys(14) is important for EcoDam
binding of the extrahelical target base into the active site pocket.
This suggests that the hexapeptide couples specific DNA binding
(Lys(9)), AdoMet binding (Trp(10)), and insertion of the flipped target
base into the active site pocket (Lys(14)).

<>

<1>Lienau, E.K., Strain, E., Wang, C., Zheng, J., Ottesen, A.R., Keys, C.E., Hammack, T.S., Musser, S.M., Brown, E.W., Allard, M.W., Cao, G., Meng, J., Stones, R.
<2>Identification of a salmonellosis outbreak by means of molecular sequencing.
<3>N. Engl. J. Med.
<4>364
<5>981-982
<6>2011
<7>
<>

<1>Liesegang, H., Kaster, A.K., Wiezer, A., Goenrich, M., Wollherr, A., Seedorf, H., Gottschalk, G., Thauer, R.K.
<2>Complete Genome Sequence of Methanothermobacter marburgensis, a Methanoarchaeon Model Organism.
<3>J. Bacteriol.
<4>192
<5>5850-5851
<6>2010
<7>The circular genome sequence of the chemolithoautotrophic euryarchaeon Methanothermobacter
marburgensis, with 1,639,135 bp, was determined and
compared with that of Methanothermobacter thermautotrophicus. The genomes
of the two model methanogens differ substantially in protein coding
sequences, in insertion sequence (IS)-like elements, and in clustered
regularly interspaced short palindromic repeats (CRISPR) loci.

<>

<1>Liljeqvist, M., Valdes, J., Holmes, D.S., Dopson, M.
<2>Draft Genome of the Psychrotolerant Acidophile Acidithiobacillus ferrivorans SS3.
<3>J. Bacteriol.
<4>193
<5>4304-4305
<6>2011
<7>Acidithiobacillus ferrivorans SS3 is a psychrotolerant acidophile capable of growth in the
range of 5 degrees  to 30 degrees C (optimum,
approximately 25 degrees C). It gains energy from the oxidation of ferrous
iron and inorganic sulfur compounds and obtains organic carbon from carbon
dioxide. Here, we present the draft genome sequence of A. ferrivorans SS3
that will permit investigation of genes involved in growth in acidic
environments at low temperatures.

<>

<1>Lilley, D.M.J.
<2>Cisplatin adducts in DNA: distortion and recognition.
<3>J. Biol. Inorg. Chem.
<4>1
<5>189-191
<6>1996
<7>cis-Diamminedichloroplatinum (II) (cisplatin) and derivatives are very successful anticancer
chemotherapeutic agents.  They crosslink cellular DNA, forming bifunctional adducts with the
N7 of guanine bases.  In this review, recent structures of cisplatin adducts are summarized,
and the significance for the recognition of DNA structure by proteins is discussed.  Two new
structures of intrastrand GpG adducts have been presented, showing a significant kinking of
the helix axis and a novel hybrid A-B helical geometry.  The relevance of this structure to
the recognition of HMG-box and related proteins is discussed.  A new structure of a
cross-strand cisplatin adduct reveals a major disruption of the local DNA structure.  The
basepairs containing the modified guanine bases are broken, with extrusion of the cytosine
bases into the solvent.  The backbone reverses direction locally, with the result that the
platinum adduct is located in what is the minor groove of the DNA overall.  The extrusion of
single bases out of the helix is strongly reminiscent of the effect of certain methylases on
their DNA targets.

<>

<1>Lim, H.I., Lee, J., Jang, J.Y., Park, H.W., Choi, H.J., Kim, T.W., Kang, M.R., Lee, J.H.
<2>Draft Genome Sequence of Lactobacillus sakei Strain wikim 22, Isolated from Kimchi in Chungcheong Province, South Korea.
<3>Genome Announcements
<4>2
<5>e01296-14
<6>2014
<7>We report the draft genome sequence of Lactobacillus sakei strain wikim 22, a Lactobacillus
species isolated from kimchi in North Chungcheong Province, South
Korea, having 155 contigs with 2,447 genes and an average G+C content of 40.61%.

<>

<1>Lim, J., Lee, T.H., Nahm, B.H., Choi, Y.D., Kim, M., Hwang, I.
<2>Complete Genome Sequence of Burkholderia glumae BGR1.
<3>J. Bacteriol.
<4>191
<5>3758-3759
<6>2009
<7>Burkholderia glumae is the causative agent of grain and seedling rot in rice and of bacterial
wilt in many field crops. Here, we report the complete genome sequence of B. glumae BGR1
isolated from a diseased rice panicle in Korea.

<>

<1>Lim, J.B., Hwang, H.Y.
<2>A novel restriction enzyme SolI from Streptoverticillium olivoverticillatum.
<3>Korean Patent Office
<4>KR 147778 B
<5>
<6>1998
<7>
<>

<1>Lim, J.S., Choi, B.S., Choi, A.Y., Kim, K.D., Kim, D.I., Choi, I.Y., Ka, J.O.
<2>Complete Genome Sequence of the Fenitrothion-Degrading Burkholderia sp. Strain YI23.
<3>J. Bacteriol.
<4>194
<5>896
<6>2012
<7>Burkholderia species are ubiquitous in soil environments. Many Burkholderia species isolated
from various environments have the potential
to biodegrade man-made chemicals. Burkholderia sp. strain YI23 was
isolated from a golf course soil and identified as a
fenitrothion-degrading bacterium. In this study, we report the complete
genome sequence of Burkholderia sp. strain YI23.

<>

<1>Lim, S.K., Kim, S.J., Cha, S.H., Oh, Y.K., Rhee, H.J., Kim, M.S., Lee, J.K.
<2>Complete genome sequence of Rhodobacter sphaeroides KD131.
<3>J. Bacteriol.
<4>191
<5>1118-1119
<6>2009
<7>Rhodobacter sphaeroides is a purple nonsulfur photosynthetic bacterium that is considered a
possible source of H(2) production. R. sphaeroides
KD131, which was isolated from sea mud in South Korea, was found to
produce high levels of H(2). Here we report the complete and annotated
genome sequence of R. sphaeroides KD131.

<>

<1>Lima, A.O., Cabral, A., Andreote, F.D., Cavalett, A., Pessatti, M.L., Dini-Andreote, F., da Silva, M.A.
<2>Draft Genome Sequence of Bacillus stratosphericus LAMA 585, Isolated from the Atlantic Deep Sea.
<3>Genome Announcements
<4>1
<5>e00204-13
<6>2013
<7>Bacillus stratosphericus LAMA 585 was isolated from the Mid-Atlantic-Ridge seafloor (5,500-m
depth). This bacterium presents the capacity for cellulase,
xylanase, and lipase production when growing aerobically in marine-broth media.
Genes involved in the tolerance of oligotrophic and extreme conditions and
prospection of biotechnological products were annotated in the draft genome (3.7
Mb).

<>

<1>Lima, A.R., Siqueira, A.S., Dos Santos, B.G., da Silva, F.D., Inada, D.T., Lima, C.P., Cardoso, J.F., Vianez-Junior, J.L., Nunes, M.R., Goncalves, E.C.
<2>Draft Genome Sequence of Rhodobacter sp. Strain CACIA 14H1, a Heterotrophic Bacterium Obtained from a Nonaxenic Culture of a Cyanobium Species.
<3>Genome Announcements
<4>2
<5>e01116-13
<6>2014
<7>Despite their prominent importance, few efforts have been paid to the genomic analysis of
heterotrophic bacteria associated with cyanobacteria. Thus, this work
presents the draft genome sequence (~3.9 Mbp) of a heterotrophic bacterium
(Rhodobacter sp. strain CACIA 14H1) recovered from a nonaxenic culture of a
Cyanobium species.

<>

<1>Lima, A.R., Siqueira, A.S., Dos Santos, B.G., da Silva, F.D., Lima, C.P., Cardoso, J.F., Vianez, J.J.L., Dall'Agnol, L.T., McCulloch, J.A., Nunes, M.R., Goncalves, E.C.
<2>Draft Genome Sequence of the Brazilian Cyanobium sp. Strain CACIAM 14.
<3>Genome Announcements
<4>2
<5>e00669-14
<6>2014
<7>Given the scarcity of data pertaining to whole-genome sequences of cyanobacterial strains
isolated in Brazil, we hereby present the draft genome sequence of the
Cyanobium sp. strain CACIAM 14, isolated in southeastern Amazonia.

<>

<1>Lima, A.R., Siqueira, A.S., Dos Santos, B.G., da Silva, F.D., Lima, C.P., Cardoso, J.F., Vianez-Junior, J.L., Nunes, M.R., Goncalves, E.C.
<2>Draft Genome Sequence of Blastomonas sp. Strain CACIA 14H2, a Heterotrophic Bacterium Associated with Cyanobacteria.
<3>Genome Announcements
<4>2
<5>e01200-13
<6>2014
<7>With the new methods for assembling sequence data from metagenomic samples, the genomic study
of heterotrophic bacterium-cyanobacterium associations can now be
improved. In this work, the draft genome sequence of Blastomonas sp. strain CACIA
14H2, obtained from a nonaxenic culture of a Cyanobium sp., is presented.

<>

<1>Limsowtin, G.K.Y., Heap, H.A., Lawrence, R.C.
<2>Heterogeneity among strains of lactic streptococci.
<3>New Zealand J. Dairy Sci. Techn.
<4>13
<5>1-8
<6>1978
<7>Heterogeneity within lactic streptococcal cultures was studied with respect to
acid production and phage sensitivity.  All strains were found to accumulate
slow coagulating variants at different rates, depending upon the growth media
used.  Maintenance of cultures in M17 broth resulted in a significant
accumulation of slow coagulating variants for most strains studied.  In all but
one strain investigated, variants were detected which exhibited different phage
sensitivies.  When isolates from these cultures were examined four phenotypes
were observed:  1) Variants exhibiting host modification and restriction of
phages; 2) Variants which adsorbed but did not form plaques (with one or more
particular phages); 3) Variants resistant against one or more particular
phages; 4) Variants sensitive to different phages.  Generally only one or two
of these four phenotypes were recovered from any one strain but a detailed
investigation of strain 108 showed that all four phenotypes were present.  The
appearance of these variants was dependent upon the nature of subculturing
medium, being much more rapid in M17 broth than in autoclaved reconstituted
skim milk.  The commercial importance of these findings is discussed.

<>

<1>Lin, A.C., Liao, T.L., Lin, Y.C., Lai, Y.C., Lu, M.C., Chen, Y.T.
<2>Complete Genome Sequence of Klebsiella pneumoniae 1084, a Hypermucoviscosity-Negative K1 Clinical Strain.
<3>J. Bacteriol.
<4>194
<5>6316
<6>2012
<7>We report the complete genome sequence of Klebsiella pneumoniae 1084, a
hypermucoviscosity-negative K1 clinical strain. Sequencing and annotation
revealed a 5,386,705-bp circular chromosome (57.4% G+C content), which contains
4,962 protein-coding genes, 80 tRNA genes, and 25 rRNA genes.

<>

<1>Lin, B., Lu, G., Li, S., Hu, Z., Chen, H.
<2>Draft Genome Sequence of the Novel Agarolytic Bacterium Aquimarina agarilytica ZC1.
<3>J. Bacteriol.
<4>194
<5>2769
<6>2012
<7>The marine bacterium ZC1 is the type strain of the recently identified novel species
Aquimarina agarilytica. It can produce multiple agarases. Here we report
the draft genome sequence of strain ZC1 (4,253,672 bp, with a GC content of
32.8%) and major findings from its annotation. It is the first reported genome in
the genus Aquimarina.

<>

<1>Lin, B.-C., Chien, M.-C., Lou, S.-Y.
<2>A sequence-specific endonuclease (XmnI) from Xanthomonas manihotis.
<3>Nucleic Acids Res.
<4>8
<5>6189-6198
<6>1980
<7>A type II restriction endonuclease XmnI with a novel site specificity has been
isolated from Xanthomonas manihotis.  XmnI does not cleave SV40 DNA, but
cleaves PhiX174 DNA into three fragments, which constitute 76.61%, 18.08% and
5.31% of the total length of 5386 base pairs, and cleaves pBR322 DNA into two
fragments of 55.71% and 44.29% of the entire 4362 base pairs.  The nucleotide
sequences around the cleavage sites made by XmnI are not exactly homologous,
but they have a common sequence of 5' GAANNNNTTC 3' according to a simple
computer program analysis on nucleotide sequences of PhiX174 DNA, pBR322 DNA
and SV40 DNA. The results suggest that the cleavage site of XmnI is located
within its recognition sequence of 5' GAANNNNTTC 3'.

<>

<1>Lin, C., den Bakker, H.C., Suzuki, H., Lefebure, T., Ponnala, L., Sun, Q., Stanhope, M.J., Wiedmann, M., Duhamel, G.E.
<2>Complete Genome Sequence of the Porcine Strain Brachyspira pilosicoli P43/6/78(T.).
<3>Genome Announcements
<4>1
<5>e00215-12
<6>2013
<7>Reported herein is the complete genome sequence of strain P43/6/78, isolated from a pig with
clinical disease. This sequence will aid in the study of genome-wide
comparison among species.

<>

<1>Lin, D., Guo, Y., Chen, C., Fuzhu, Y., Xiao, S., Zhu, D., Wang, M., Xu, X.
<2>Draft Genome Sequence of Linezolid-Resistant Enterococcus faecalis Clinical Isolate HS0914.
<3>Genome Announcements
<4>2
<5>e00782-14
<6>2014
<7>We report the draft genome sequence of linezolid-resistant Enterococcus faecalis  strain
HS-0914 isolated from a teaching hospital in Shanghai, China. The draft
genome sequence is composed of 61 contigs for 2,816,079 bp. Ribosomal RNA
mutations and cfr, which mediates linezolid resistance, are not present.

<>

<1>Lin, H., Coletta-Filho, H.D., Han, C.S., Lou, B., Civerolo, E.L., Machado, M.A., Gupta, G.
<2>Draft Genome Sequence of 'Candidatus Liberibacter americanus' Bacterium Associated with Citrus Huanglongbing in Brazil.
<3>Genome Announcements
<4>1
<5>e00275-13
<6>2013
<7>We report here the draft genome sequence of 'Candidatus Liberibacter americanus'  strain
PW_SP. The 1,176,071-bp genome, with 31.6% G+C content, comprises 948 open
reading frames, 38 tRNAs, and three complete rRNAs.

<>

<1>Lin, H., Han, C.S., Liu, B., Lou, B., Bai, X., Deng, C., Civerolo, E.L., Gupta, G.
<2>Complete Genome Sequence of a Chinese Strain of 'Candidatus Liberibacter asiaticus'.
<3>Genome Announcements
<4>1
<5>e00184-13
<6>2013
<7>We report here the complete genome sequence of 'Candidatus Liberibacter asiaticus' (strain
Guangxi-1). The 1,268,237-bp genome with a 36.5% G+C content
comprises 1,141 open reading frames, 44 tRNAs, and 3 complete rRNAs in a circular
chromosome.

<>

<1>Lin, I.-H., Liu, T.-T., Teng, Y.-T., Wu, H.-L., Liu, Y.-M., Wu, K.-M., Chang, C.-H., Hsu, M.-T.
<2>Sequencing and Comparative Genome Analysis of Two Pathogenic Streptococcus gallolyticus Subspecies: Genome Plasticity, Adaptation and Virulence.
<3>PLoS ONE
<4>6
<5>e20519
<6>2011
<7>Streptococcus gallolyticus infections in humans are often associated with bacteremia,
infective endocarditis and colon cancers. The disease manifestations are different depending
on the subspecies of S. gallolyticus causing the infection. Here, we present the complete
genomes of S. gallolyticus ATCC 43143 (biotype I) and S. pasteurianus ATCC 43144 (biotype
II.2).  The genomic differences between the two biotypes were characterized with comparative
genomic analyses. The chromosome of ATCC 43143 and ATCC 43144 are 2,36 and 2,10 Mb in length
and encode 2246 and 1869 CDS respectively.  The organization and genomic contents of both
genomes were most similar to the recently published S. gallolyticus UCN34, where 2073 (92%)
and 1607 (86%) of the ATCC 43143 and ATCC 43144 CDS were conserved in UCN34 respectively.
There are around 600 CDS conserved in all Streptococcus genomes, indicating the Streptococcus
genus has a small core-genome (constitute around 30% of total CDS) and substantial
evolutionary plasticity. We identified eight and five regions of genome plasticity in ATCC
43143 and ATCC 43144 respectively. Within these regions, several proteins were recognized to
contribute to the fitness and virulence of each of the two subspecies. We have also predicted
putative cell-surface associated proteins that could play a role in adherence to host tissues,
leading to persistent infections causing sub-acute and chronic diseases in
humans. This study showed evidence that the S. gallolyticus still possesses genes making it
suitable in a rumen environment, whereas the ability for S. pasteurianus to live in rumen is
reduced. The genome heterogeneity and genetic diversity among the two biotypes, especially
membrane and lipoproteins, most likely contribute to the differences in the pathogenesis of
the two S. gallolyticus biotypes and the type of disease an infected patient eventually
develops.

<>

<1>Lin, I.G., Han, L., Taghva, A., O'Brien, L.E., Hsieh, C.L.
<2>Murine de novo methyltransferase Dnmt3a demonstrates strand asymmetry and site preference in the methylation of DNA in vitro.
<3>Mol. Cell. Biol.
<4>22
<5>704-723
<6>2002
<7>CpG methylation is involved in a wide range of biological processes in vertebrates as well as
in plants and fungi. To date, three enzymes, Dnmt1, Dnmt3a, and Dnmt3b, are known to have DNA
methyltransferase activity in mouse and human. It has been proposed that de novo methylation
observed in early embryos is predominantly carried out by the Dnmt3a and Dnmt3b
methyltransferases, while Dntm1 is believed to be responsible for maintaining the established
methylation patterns upon replication. Analysis of the sites methylated in vivo using the
bisulfite genomic sequencing method confirms the previous finding that some regions of the
plasmid are much more methylated by Dnmt3a than other regions on the same plasmid. However,
the preferred targets of the enzyme cannot be determined due to the presence of other
methylases, DNA binding proteins, and chromatin structure. To discern the DNA targets of
Dnmt3a without these compounding factors, sites methylated by Dnmt3a in vitro were analyzed.
These analyses revealed that the two cDNA strands have distinctly different methylation
patterns. Dnmt3a prefers CpG sites on a strand in which it is flanked by pyrimidines over CpG
sites flanked by purines in vitro. These findings indicate that, unlike Dnmt1, Dnmt3a most
likely methylates one strand of DNA without concurrent methylation of the CpG site on the
complementary strand. These findings also indicate that Dnmt3a may methylate some CpG sites
more frequently than others, depending on the sequence context. Methylation of each DNA strand
independently and with possible sequence preference is a novel feature among the known DNA
methyltransferases.

<>

<1>Lin, J., Vogt, V.M.
<2>I-PpoI, the endonuclease encoded by the group I intron PpLSU3, is expressed from an RNA polymerase I transcript.
<3>Mol. Cell. Biol.
<4>18
<5>5809-5817
<6>1998
<7>PpLSU3, a mobile group I intron in the rRNA genes of Physarum polycephalum, also can home into
yeast chromosomal ribosomal DNA (rDNA). By integrating PpLSU3 into the rDNA copies of a yeast
strain temperature sensitive for RNA polymerase I, we have shown that the I-PpoI homing
endonuclease encoded by PpLSU3 is expressed from an RNA polymerase I transcript. We have also
developed a method to integrate mutant forms of PpLSU3 as well as the Tetrahymena intron
TtLSU1 into rDNA, by expressing I-PpoI in trans. Analysis of I-PpoI expression levels in these
mutants, along with subcellular fractionation of intron RNA, strongly suggests that the
full-length excised intron RNA, but not RNAs that are further cleaved, serves as or gives rise
to the mRNA.

<>

<1>Lin, L., Xu, Q., Zhang, Y., Yin, B., Fan, Y., Luo, Y., Han, S., Zhang, Z., Wu, G., Shen, Y.
<2>Alternative splicing of de novo methyltransferase gene 3b in adult and newborn mice.
<3>Zhongguo Yixue Kexueyuan Xuebao
<4>22
<5>312-316
<6>2000
<7>Objective. To unravel the biological significance of the alternative splicing of de novo
methyltransferase 3b, expression of the Dnmt3b gene in various tissues and developmental
stages was investigated in postnatal mice.  Methods. RT-PCR and capillary electrophoresis were
employed to analyze the alternative splicing pattern of Dnmt3b in tissues of newborn and adult
mice.  The results had been further reaffirmed by repeating and statistics analysis.
Bioinformatics tools were used to predict the structure and hydrophobicity of the Dnmt3b
exon10 coding sequence.  Results. The isoform with Dnmt3b exon10 was the abundant form in the
lung of newborn mice and in the liver of both newborn and adult mice, while in other tissues
of newborn and adult mice, the spliced isoform was present as the predominant one.  The
peptide encoded by Dnmt3b exon10 was mainly random coil on the surface of Dnmt3b protein.
Conclusions. The data demonstrate that the specific expression of Dnmt3b exists in tissues and
developmental stages of postnatal mice.  The alternative splicing of the exon 10 of Dnmt3b is
possibly involved in the regulation of Dnmt3b's catalytic function.  These results provide an
insight into the developmental regulation and physiological function of the alternative
splicing of the Dnmt3b gene.

<>

<1>Lin, L.F., Posfai, J., Roberts, R.J., Kong, H.M.
<2>Comparative genomics of the restriction-modification systems in Helicobacter pylori.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>2740-2745
<6>2001
<7>Helicpbacter pylori is a Gram-negative bacterial pathogen with a small genome of 1.64-1.67 Mb.
More than 20 putative DNA
restriction-modification (R-M) systems, comprising more than 4% of the
total genome, have been identified in the two completely sequenced H.
pylori strains, 26695 and J99, based on sequence similarities. In this
study, we have investigated the biochemical activities of 14 Type II
R-M systems in H. pylori 26695. Less than 30% of the Type II R-M
systems in 26695 are fully functional, similar to the results obtained
from strain J99. Although nearly 90% of the R-M genes are shared by the
two H. pylori strains, different sets of these R-M genes are
functionally active in each strain. Interestingly, all strain-specific
R-M genes are active, whereas most shared genes are inactive. This
agrees with the notion that strain-specific genes have been acquired
more recently through horizontal transfer from other bacteria and
selected far function. Thus, they are less likely to he impaired by
random mutations. Our results also show that H. pylori has extremely
diversified R-M systems in different strains, and that the diversity
may be maintained by constantly acquiring new R-M systems and by
inactivating and deleting the old ones.

<>

<1>Lin, M.J., Lee, T.L., Hsu, D.W., Shen, C.K.
<2>One-codon alternative splicing of the CpG MTase dnmt1 transcript in mouse somatic cells.
<3>FEBS Lett.
<4>469
<5>101-104
<6>2000
<7>The genomic methylation patterns in the mammalian somatic cells are presumably maintained by a
single enzyme, dnmt1. In mouse, this DNA
(cytosine-5)-methyltransferase, or CpG MTase, is encoded by the Dnmt1 gene. We now present
evidence that in different tissues and cell types, the primary transcript of
mouse dnmt1 is alternatively spliced to generate two poly-(A) RNAs of approximately similar
abundance. This alternative splicing most likely originates from the existence
of two tandemly arranged acceptor sites separated by only 3 nt. The two Dnmt1 mRNAs thus
encode two CpG MTases differing by two amino acids. We discuss the
implications of the discovery of two dnmt1 isozymes, instead of one enzyme as previously
thought, in the somatic cells of both mouse and human.

<>

<1>Lin, M.J., Tang, L.Y., Reddy, M.N., Shen, C.K.J.
<2>DNA methyltransferase gene dDnmt2 and longevity of Drosophila.
<3>J. Biol. Chem.
<4>280
<5>861-864
<6>2005
<7>The DNA methylation program of the fruit fly Drosophila melanogaster is carried out by the
single DNA methyltransferase gene dDnmt2, the
function of which is unknown before. We present evidence that
intactness of the gene is required for maintenance of the normal life
span of the fruit flies. In contrast, overexpression of dDnmt2 could
extend Drosophila life span. The study links the Drosophila DNA
methylation program with the small heatshock proteins and longevity/
aging and has interesting implication on the eukaryotic DNA methylation
programs in general.

<>

<1>Lin, N., Liu, Z., Zhou, J., Wang, S., Fleming, J.
<2>Draft Genome Sequences of Two Super-XDR Isolates of M. tuberculosis from China.
<3>FEMS Microbiol. Lett.
<4>347
<5>93-96
<6>2013
<7>The prevalence of drug-resistance in Mycobacterium tuberculosis is already having a negative
impact on the control of TB. We report the draft genome sequences of
two super-extensively drug-resistant (S-XDR) Mycobacterium tuberculosis isolates
from China, FJ05194 (lineage 2) and GuangZ0019 (lineage 4), and compare them to
the H37Rv reference strain to identify possible sources of genetic variation
associated with their extensive drug resistance. Our results suggest that their
extensive drug resistance most likely results from the stepwise accumulation of
resistances to individual drugs. This article is protected by copyright. All
rights reserved.

<>

<1>Lin, P.M., Lee, C.H., Roberts, R.J.
<2>Cloning and characterization of the genes encoding the MspI restriction modification system.
<3>Nucleic Acids Res.
<4>17
<5>3001-3011
<6>1989
<7>The genes encoding the MspI restriction modification system, which recognizes
the sequence 5' CCGG, have been cloned into pUC9.  Selection was based on
expression of the cloned methylase gene which renders plasmid DNA insensitive
to MspI cleavage in vitro.  Initially, an insert of 15 kb was obtained which,
upon subcloning, yielded a 3 kb EcoRI to HindIII insert, carrying the genes for
both the methylase and the restriction enzyme.  This insert has been sequenced.
Based upon the sequence, together with appropriate subclones, it is shown that
the two genes are transcribed divergently with the methylase gene encoding a
polypeptide of 418 amino acids, while the restriction enzyme is composed of 262
amino acids.  Comparison of the sequence of the MspI methylase with other
cytosine methylases shows a striking degree of similarity.  Especially
noteworthy is the high degree of similarity with the HhaI and EcoRII
methylases.

<>

<1>Lin, S.Y., Shieh, W.Y., Chen, J.S., Tang, S.L.
<2>Complete Genome Sequence of Simiduia agarivorans SA1(T), a Marine Bacterium Able  To Degrade a Variety of Polysaccharides.
<3>Genome Announcements
<4>1
<5>e00039-12
<6>2013
<7>Simiduia agarivorans strain SA1(T) is able to degrade a variety of polysaccharides found in
marine algae, plants, and animals. The genome of S. agarivorans SA1(T) consists of a single
chromosome (4,309,711 bp), and its information may provide insights into the
polysaccharide-degrading capability, cell division, flagellar motility, and chemotaxis of this
bacterium.

<>

<1>Lin, T.L., Shun, C.T., Chang, K.C., Wang, J.T.
<2>Isolation and characterization of a HpyC1I restriction-modification system in Helicobacter pylori.
<3>J. Biol. Chem.
<4>279
<5>11156-11162
<6>2004
<7>Using transposon shuttle mutagenesis, we identified six Helicobacter pylori mutants from the
NTUH-C1 strain that exhibited decreased adherence and cell elongation. Inverse polymerase
chain reaction and DNA sequencing revealed that the same locus was interrupted in these six
mutants. Nucleotide and amino acid sequences showed no homologies with H. pylori 26695 and J99
strains. This novel open reading frame contained 1617 base pairs. The amino acid sequence
shared 24% identity with a putative nicking enzyme in Bacillus halodurans and 23 and 20%
identity with type IIS restriction endonucleases PleI and MlyI, respectively. The purified
protein, HpyC1I, showed endonuclease activity with the recognition and cleavage site
5'-CCATC( 4/5)-3'. Two open reading frames were located upstream of the gene encoding
HpyC1I. Together, HpyC1I and these two putative methyltransferases (M1.HpyC1I and M2.HpyC1I)
function as a restriction-modification (R-M) system. The HpyC1I R-M genes were found in 9 of
the 15 H. pylori strains tested. When compared with the full genome, significantly lower G + C
content of HpyC1I R-M genes implied that these genes might have been acquired by horizontal
gene transfer. Plasmid DNA transformation efficiencies and chromosomal DNA digestion assays
demonstrated protection from HpyC1I digestion by the R-M system. In conclusion, we have
identified a novel R-M system present in similar to 60% of H. pylori strains. Disruption of
this R-M system results in cell elongation and susceptibility to HpyC1I digestion.

<>

<1>Lin, X., Wang, Z., Li, Y.
<2>Draft Genome Sequence of Pseudoalteromonas sp. Strain A2, an Isolate with High Antioxidative Activity from Arctic Seawater.
<3>Genome Announcements
<4>2
<5>e00885-14
<6>2014
<7>Here we report the draft genome sequence of Pseudoalteromonas strain A2, isolated from Arctic
seawater in the pack-ice zone, which has high antioxidative activity
against H2O2. The genomics information of this strain will facilitate the study
of antioxidative mechanisms, cold adaptation properties, and evolution of this
genus.

<>

<1>Lin, Y., Fan, H., Hao, X., Johnstone, L., Hu, Y., Wei, G., Alwathnani, H.A., Wang, G., Rensing, C.
<2>Draft genome sequence of Halomonas sp. Strain HAL1, a moderately halophilic arsenite-oxidizing bacterium isolated from gold-mine soil.
<3>J. Bacteriol.
<4>194
<5>199-200
<6>2011
<7>We report the draft genome sequence of arsenite-oxidizing Halomonas sp. strain HAL1, isolated
from the soil of a gold mine. Genes encoding proteins involved in arsenic resistance and
transformation, phosphate utilization and uptake, and betaine biosynthesis were identified.
Their identification might help in understanding how arsenic and phosphate metabolism are
intertwined.

<>

<1>Lin, Y., Hao, X., Johnstone, L., Miller, S.J., Baltrus, D.A., Rensing, C., Wei, G.
<2>Draft Genome of Streptomyces zinciresistens K42, a Novel Metal-Resistant Species Isolated from Copper-Zinc Mine Tailings.
<3>J. Bacteriol.
<4>193
<5>6408-6409
<6>2011
<7>A draft genome sequence of Streptomyces zinciresistens K42, a novel Streptomyces species
displaying a high level of resistance to zinc and
cadmium, is presented here. The genome contains a large number of genes
encoding proteins predicted to be involved in conferring metal resistance.
Many of these genes appear to have been acquired through horizontal gene
transfer.

<>

<1>Lin, Y.T., Tsai, J.C., Chen, H.J., Hung, W.C., Hsueh, P.R., Teng, L.J.
<2>A Novel Staphylococcal Cassette Chromosomal Element, SCCfusC, Carrying fusC and speG in Fusidic Acid-Resistant Methicillin-Resistant Staphylococcus aureus.
<3>Antimicrob. Agents Chemother.
<4>58
<5>1224-1227
<6>2014
<7>A high prevalence of fusC (16/46, 59%) was found in fusidic acid-resistant
methicillin-resistant Staphylococcus aureus isolates collected from 2008 to 2010.
Nucleotide sequencing of fusC and flanking regions revealed a novel
staphylococcal cassette chromosome (SCC) structure, SCCfusC, which was integrated
into rlmH and located upstream from SCCmec. The SCCfusC element contained speG,
which may contribute to the polyamine resistance.

<>

<1>Lin, Z., Liu, Z., Yang, R., Zou, Y., Wan, D., Chen, J., Guo, M., Zhao, J., Fang, C., Yang, R., Liu, F.
<2>Whole-Genome Sequencing of Lactobacillus shenzhenensis Strain LY-73T.
<3>Genome Announcements
<4>1
<5>e00972-13
<6>2013
<7>Lactobacillus shenzhenensis strain LY-73T is a novel species which was first isolated from
fermented goods. Here, we report the draft genome sequence of
Lactobacillus shenzhenensis LY-73T.

<>

<1>Linares, D.M., Kok, J., Poolman, B.
<2>Genome Sequences of Lactococcus lactis MG1363 (Revised) and NZ9000 and Comparative Physiological Studies.
<3>J. Bacteriol.
<4>192
<5>5806-5812
<6>2010
<7>Lactococcus lactis NZ9000 and its parent MG1363 are the most commonly used lactic acid
bacteria for expression and physiological studies. We noted
unexpected but significant differences in the growth behaviors of both
strains. We sequenced the entire genomes of the original NZ9000 and MG1363
strains using an ultradeep sequencing strategy. The analysis of the L.
lactis NZ9000 genome yielded 79 differences, mostly point mutations, with
the annotated genome sequence of L. lactis MG1363. Resequencing of the
MG1363 strain revealed that 73 out of the 79 differences were due to
errors in the published sequence. Comparative transcriptomic studies
revealed several differences in the regulation of genes involved in sugar
fermentation, which can be explained by two specific mutations in a region
of the ptcC promoter with a key role in the regulation of cellobiose and
glucose uptake.

<>

<1>Lindahl, T.
<2>Enzymatic methylation of DNA - Roles and prospects.
<3>Biol. Chem.
<4>379
<5>375-376
<6>1998
<7>The occurrence of a fifth base in the DNA of mammalian cells and plant cells has been known
for close to fifty years.  However, the difficulties in relating 5-methylcytosine residues to
specific functional roles, and the apparent absence of such DNA residues in the intensely
studied eukaryotes Drosophila melanogaster and Saccharomyces cerevisiae, meant there were
relatively few investigations carried out on DNA methylation until recently.  New data on DNA
methylation as a mechanism for silencing of gene expression and its involvement in other
aspects of epigenetic control in mammalian cells, as well as the embryonic lethal phenotype of
gene knockout mouse constructs lacking the methyltransferase required for maintenance
methylation, have greatly increased the amount of activity in the field.  Nevertheless,
several potentially important aspects, such as the complicated perturbations of DNA
methylation patterns in cancer cells involving both hypermethylation and hypomethylation, and
the mechanisms of de novo methylation and demethylation, are still at very early stages of
investigation.  The merging data on eukaryotes may be contrasted with the, by now, well
established roles of DNA methylation in bacteria.  This occurs at DNA adenine as well as
cytosine residues, and serves to protect the genome from endogenous restriction endonucleases
as well as providing a strategy for strand discrimination in mismatch repair immediately after
DNA replication.

<>

<1>Linder, P., Doelz, R., Gubler, M., Bickle, T.A.
<2>An anticodon nuclease gene inserted into a hsd region encoding a type I DNA restriction system.
<3>Nucleic Acids Res.
<4>18
<5>7170
<6>1990
<7>None

<>

<1>Lindler, L.E., Plano, G.V., Burland, V., Mayhew, G.F., Blattner, F.R.
<2>Complete DNA sequence and detailed analysis of the Yersinia pestis KIM5 plasmid encoding murine toxin and capsular antigen.
<3>Infect. Immun.
<4>66
<5>5731-5742
<6>1998
<7>Yersinia pestis, the causative agent of plague, harbors at least three plasmids necessary for
full virulence of the organism, two of which are
species specific. One of the Y. pestis-specific plasmids, pMT1, is thought
to promote deep tissue invasion, resulting in more acute onset of symptoms
and death. We determined the entire nucleotide sequence of Y. pestis KIM5
pMT1 and identified potential open reading frames (ORFs) encoded by the
100,990-bp molecule. Based on codon usage for known yersinial genes,
homology with known proteins in the databases, and potential ribosome
binding sites, we determined that 115 of the potential ORFs which we
considered could encode polypeptides in Y. pestis. Five of these ORFs were
genes previously identified as being necessary for production of the
classic virulence factors, murine toxin (MT), and the fraction 1 (F1)
capsule antigen. The regions of pMT1 encoding MT and F1 were surrounded by
remnants of multiple transposition events and bacteriophage, respectively,
suggesting horizontal gene transfer of these virulence factors. We
identified seven new potential virulence factors that might interact with
the mammalian host or flea vector. Forty-three of the remaining 115
putative ORFs did not display any significant homology with proteins in
the current databases. Furthermore, DNA sequence analysis allowed the
determination of the putative replication and partitioning regions of
pMT1. We identified a single 2,450-bp region within pMT1 that could
function as the origin of replication, including a RepA-like protein
similar to RepFIB, RepHI1B, and P1 and P7 replicons. Plasmid partitioning
function was located ca. 36 kb from the putative origin of replication and
was most similar to the parABS bacteriophage P1 and P7 system. Y. pestis
pMT1 encoded potential genes with a high degree of similarity to a wide
variety of organisms, plasmids, and bacteriophage. Accordingly, our
analysis of the pMT1 DNA sequence emphasized the mosaic nature of this
large bacterial virulence plasmid and provided implications as to its
evolution.

<>

<1>Lindroth, A.M., Cao, X., Jackson, J.P., Zilberman, D., McCallum, C.M., Henikoff, S., Jacobsen, S.E.
<2>Requirement of CHROMOMETHYLASE3 for maintenance of CpXpG methylation.
<3>Science
<4>292
<5>2077-2080
<6>2001
<7>Epigenetic silenced alleles of the Arabidopsis SUPERMAN locus (the clark kent alleles) are
associated with dense hypermethylation at noncanonical cytosines (CpXpG and asymmetric sites,
where X = A, T, C, or G). A genetic screen for suppressors of a hypermethylated clark kent
mutant identified nine loss-of-function alleles of CHROMOMETHYLASE3 (CMT3), a novel cytosine
methyltransferase homolog. These cmt3 mutants display a wild-type morphology but exhibit
decreased CpXpG methylation of the SUP gene and of other sequences throughout the genome. They
also show reactivated expression of endogenous retrotransposon sequences. These results show
that a non-CpG DNA methyltransferase is responsible for maintaining epigenetic gene silencing.

<>

<1>Lindsay, J.A.
<2>Genomic variation and evolution of Staphylococcus aureus.
<3>Int. J. Med. Microbiol.
<4>300
<5>98-103
<6>2010
<7>The evolution of new human and animal pathogenic strains of Staphylococcus aureus has been due
to the accumulation of mobile genetic elements (MGE) encoding methicillin resistance and
virulence factors into successful lineages. These include epidemic methicillin-resistant S.
aureus in hospitals (EMRSA), community-associated MRSA (CA-MRSA), fully vancomycin-resistant
MRSA (VRSA) and livestock-associated MRSA (LA-MRSA). The S. aureus population in humans is
dominated by about ten S. aureus lineages while animals generally have different lineages.
Individual isolates within each lineage have unique combination of MGE often encoding
virulence and resistance genes. S. aureus evolves due to point mutation and selection, but
also dramatically due to the horizontal transfer of these MGE between strains or from other
species or genera. Horizontal transfer, by conjugation or transduction, can be blocked by S.
aureus restriction modification systems which are lineage specific. Because of the mobility of
MGE, there are prospects for increasingly Virulent and resistant Strains to emerge that could
severely affect healthcare and agriculture more effectively than the current pathogens.

<>

<1>Lindsay, J.A.
<2>Staphylococcus aureus genomics and the impact of horizontal gene transfer.
<3>Int. J. Med. Microbiol.
<4>304
<5>103-109
<6>2014
<7>Whole genome sequencing and microarrays have revealed the population structure of
Staphylococcus aureus, and identified epidemiological shifts, transmission routes, and
adaptation of major clones. S. aureus genomes are highly diverse. This is partly due to a
population structure of conserved lineages, each with unique combinations of genes encoding
surface proteins, regulators, immune evasion and virulence pathways. Even more variable are
the mobile genetic elements (MGE), which encode key proteins for antibiotic resistance,
virulence and host-adaptation. MGEs can transfer at high frequency between isolates of the
same lineage by horizontal gene transfer (HGT). There is increasing evidence that HGT is key
to bacterial adaptation and success. Recent studies have shed light on new mechanisms of DNA
transfer such as transformation, the identification of receptors for transduction, on
integration of DNA pathways, mechanisms blocking transfer including CRISPR and new restriction
systems, strategies for evasion of restriction barriers, as well as factors influencing MGE
selection and stability. These studies have also lead to new tools enabling construction of
genetically modified clinical S. aureus isolates. This review will focus on HGT mechanisms and
their importance in shaping the evolution of new clones adapted to antibiotic resistance,
healthcare, communities and livestock.

<>

<1>Lindsey, R.L., Trees, E., Sammons, S., Loparev, V., Frace, M., Strockbine, N., Sabol, A.L., Sowers, E., Stripling, D., Martin, H., Knipe, K., Rowe, L., Gerner-Smidt, P.
<2>Draft Whole-Genome Sequences of Nine Non-O157 Shiga Toxin-Producing Escherichia coli Strains.
<3>Genome Announcements
<4>2
<5>e00501-14
<6>2014
<7>Shiga toxin-producing Escherichia coli (STEC) is an important food-borne pathogen. Here, we
report the draft whole-genome sequences of nine STEC strains
isolated from clinical cases in the United States. This is the first report of
such information for STEC of serotypes O69, H11, O145:H25, O118:H16, O91:H21,
O146:H21, O45:H2, O128:H2, and O121:H19.

<>

<1>Lindstrom, W.M. Jr.
<2>The enzymology of bacterial dna methyltransferases.
<3>Ph.D. Thesis, Univ. of California, Santa Barbara
<4>
<5>109
<6>2000
<7>DNA methyltransferases are excellent prototypes for investigating DNA distortion and enzymne
specificity because catalysis requires the extrahelical stabilization of the target base
within the enzyme active site.  Equilibrium dissociation constants were determined for the
binding of the DNA (adenine-N6-)-methyltransferase M.EcoRI to DNA containing abasic sites and
base analogs incorporated at the target base.  Tighter binding to oligonucleotides containing
destabilized target base pairs was observed, corroborating spectroscopic evidence for
nucleoside flipping by that enzyme.

<>

<1>Lindstrom, W.M. Jr., Flynn, J., Reich, N.O.
<2>HhaI DNA cytosine-C5 methyltransferase: Reconciling function and structure.
<3>FASEB J.
<4>13
<5>A1443
<6>1999
<7>HhaI DNA methyltransferase utilizes S-adenosyl-L-methionine to modify the 5-carbon of the
inner cytosine in the cognate sequence 5'-GCGC-3'.  We directly demonstrate the catalytic
competence of the enzyme-DNA complex, and the catalytic incompetence of the enzyme-AdoMet
complex.  Formation of the dead-end enzyme-AdoMet complex is consistent with the AdoMet
binding observed by x-ray crystallographic analysis of the binary complex.  The enzyme has a
two-fold preference for unmethylated over hemimethylated DNA based on kcat/KmDNA.  The methyl
transfer step contributes little to this discrimination, in contrast to the mammalian DNA
cytosine methyltransferase.  Surprisingly, in the absence of any cofactor, the enzyme binds
hemimethylated DNA seven-fold more tightly than unmethylated DNA.  This overall DNA binding
affinity is increased 10,000-fold and the preference for hemimethylated DNA is enhanced to
greater than 140-fold in the ternary enzyme-DNA-cofactor complex.  The differential binding
affinities of hemi- and unmethylated substrates directly alter how the enzyme processes its
substrate during the catalytic cycle.  Our functional characterization is presented in the
context of the available co-crystal structures of the enzyme-hemimethylated DNA-cofactor,
enzyme-unmethylated DNA-cofactor, and enzyme-AdoMet complexes.  The combined functional and
structural analyses provide insight into the discrimination of hemi- and unmethylated DNA, and
the methyl transfer step, and comparisons with the mammalian DNA methyltransferase.

<>

<1>Lindstrom, W.M. Jr., Malygin, E.G., Ovechkina, L.G., Zinoviv, V.V., Reich, N.O.
<2>Functional analysis of BamHI DNA cytosine-N4 methyltransferase.
<3>J. Mol. Biol.
<4>325
<5>711-720
<6>2003
<7>We show that the kinetic mechanism of the DNA (cytosine-N(4)-)-methyltransferase M.BamHI,
which modifies the underlined
cytosine (GGATCC), differs from cytosine C(5) methyltransferases, and is
similar to that observed with adenine N(6) methyltransferases. This
suggests that the obligate order of ternary complex assembly and
disassembly depends on the type of methylation reaction. In contrast, the
single-turnover rate of catalysis for M.BamHI (0.10s(-1)) is closer to the
DNA (cytosine-C(5)-)-methyltransferases (0.14s(-1)) than the DNA
(adenine-N(6)-)-methyltransferases (>200s(-1)). The nucleotide flipping
transition dominates the single-turnover constant for adenine N(6)
methyltransferases, and, since the disruption of the guanine-cytosine
base-pair is essential for both types of cytosine DNA methyltransferases,
this transition may be a common, rate-limiting step for methylation for
these two enzyme subclasses. The similar overall rate of catalysis by
M.BamHI and other DNA methyltransferases is consistent with a common
rate-limiting catalytic step of product dissociation. Our analyses of
M.BamHI provide functional insights into the relationship between the
three different classes of DNA methyltransferases that complement both
prior structural and evolutionary insights.

<>

<1>Lindstrom, W.M., Flynn, J., Reich, N.O.
<2>Reconciling structure and function in HhaI DNA cytosine-C-5 methyltransferase.
<3>J. Biol. Chem.
<4>275
<5>4912-4919
<6>2000
<7>Pre-steady state partitioning analysis of the HhaI DNA methyltransferase directly demonstrates
the catalytic competence of the
enzyme DNA complex and the lack of catalytic competence of the
enzyme-S-adenosyl-L-methionine (AdoMet) complex. The enzyme.AdoMet
complex does form, albeit with a 50-fold decrease in affinity compared
with the ternary enzyme.AdoMet.DNA complex. These findings reconcile
the distinct binding orientations previously observed within the binary
enzyme.AdoMet and ternary enzyme.S-adenosyl-L-homocysteine.DNA crystal
structures. The affinity of the enzyme for DNA is increased 900-fold in
the presence of its cofactor, and the preference for hemimethylated DNA
is increased to 12-fold over unmethylated DNA We suggest that this
preference is partially due to the energetic cost of retaining a cavity
in place of the B-methyl moiety in the ternary complex with the
unmethylated DNA, as revealed by the corresponding crystal structures.
The hemi- and unmethylated substrates alter the fates and lifetimes of
discrete enzyme substrate intermediates during the catalytic cycle.
Hemimethylated substrates partition toward product formation versus
dissociation significantly more than unmethylated substrates. The
mammalian DNA cytosine-C-5 methyltransferase Dnmt1 shows an even more
pronounced partitioning toward product formation.

<>

<1>Ling, Y., Sankpal, U.T., Robertson, A.K., McNally, J.G., Karpova, T., Robertson, K.D.
<2>Modification of de novo DNA methyltransferase 3a (Dnmt3a) by SUMO-1 modulates its interaction with histone deacetylases (HDACs) and its capacity to repress transcription.
<3>Nucleic Acids Res.
<4>32
<5>598-610
<6>2004
<7>The de novo DNA methyltransferase Dnmt3a is one of three mammalian DNA methyltransferases that
has been shown to play crucial roles in embryonic development, genomic imprinting and
transcriptional silencing. Despite its importance, very little is known about how the
enzymatic activity and transcriptional repression functions of Dnmt3a are regulated. Here we
show that Dnmt3a interacts with multiple components of the sumoylation machinery, namely the
E2 sumo conjugating enzyme Ubc9 and the E3 sumo ligases PIAS1 and PIASx , all of which are
involved in conjugating the small ubiquitin-like modifier polypeptide, SUMO-1, to its target
proteins. Dnmt3a is modified by SUMO-1 in vivo and in vitro and the region of Dnmt3a
responsible for interaction maps to the N-terminal regulatory domain. Functionally,
sumoylation of Dnmt3a disrupts its ability to interact with histone deacetylases (HDAC1/2),
but not with another interaction partner, Dnmt3b. Conditions that enhance the sumoylation of
Dnmt3a in vivo abolish its capacity to repress transcription. These studies reveal a new level
of regulation governing Dnmt3a whereby a post-translational modification can dramatically
regulate its interaction with specific protein partners and alter its ability to repress
transcription.

<>

<1>Ling, Y.H., Nelson, J.A., Chan, J.Y., Beattie, K.L.
<2>6-Thioguanine substituted DNA as a substrate for restriction endonucleases and ligases.
<3>Proc. Amer. Assoc. Cancer Res.
<4>33
<5>543
<6>1992
<7>
<>

<1>Linn, C.P., Walker, G.T., Spears, P.A.
<2>Fluorescence polarization detection of nucleic acids.
<3>US Patent Office
<4>US 5641633
<5>
<6>1997
<7>The present invention provides methods for detecting amplified or unamplified nucleic acid
target sequences at increased temperatures by changes in fluorescence polarization.  The
decrease in fluorescence polarization associated with hybridization of oligonucleotides at
higher, more stringent, temperatures is overcome by including a double-stranded DNA binding
protein in the assay.  At elevated temperatures, the double-stranded DNA binding protein
restores, and often enhances, the magnitude of the change in fluorescence polarization
associated with single- to double-stranded conversion of an oligonucleotide probe or primer.

<>

<1>Linn, S.
<2>The 1978 Nobel prize in physiology or medicine.
<3>Science
<4>202
<5>1069-1071
<6>1978
<7>The 1978 Nobel Prize in Physiology or Medicine was awarded to Werner Arber, 49, of the
Biozentrum in Basel, Switzerland; Hamilton O. Smith, 47 of Johns Hopkins University; and to
Daniel Nathans, 50, also of Johns Hopkins University.  The awards recognize the development of
restriction endonucleases, enzymes that can be used to study genetic organization and to
manipulate DNA for "genetic engineering".  Arber is credited with having first predicted the
existence of the enzymes, Smith with having isolated the first such enzyme and describing its
specific reaction, and Nathans with having first applied these enzymes to the study of gene
organization and regulation.

<>

<1>Linn, S., Arber, W.
<2>Host specificity of DNA produced by Escherichia coli, X.  In vitro restriction of phage fd replicative form.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>59
<5>1300-1306
<6>1968
<7>An activity has been found in fractionated extracts from Escherichia coli which
reduces the infectivity of the replicative form of phage fd DNA.  It is
correlated with the in vivo restriction phenomenon by (1) its presence only in
fractions from restricting strains of bacteria and (2) its specificity for
nonmodified DNA.  The inactivation requires S-adenosylmethionine, ATP, Mg++,
and the products of at least two gene functions; it seems to be accompanied by
double-strand cleavage of the DNA.

<>

<1>Linn, S., Lautenberger, J.A., Eskin, B., Lackey, D.
<2>Host-controlled restriction and modification enzymes of Escherichia coli B1,2.
<3>Fed. Proc.
<4>33
<5>1128-1134
<6>1974
<7>The Escherichia coli B modification methylase transfers a methyl group from
S-adenosylmethionine to DNA in a reaction wherein the enzyme may not turn over.
The corresponding restriction endonuclease puts one nick into the DNA, or
converts a previously placed restriction nick into a double-strand break in a
reaction which requires ATP and S-adenosylmethionine.  The DNA is not cleaved
at the specific recognition site, as demonstrated by centrifugational and
electrophoretic analyses and the fact that restricted DNA can be methylated by
the modification enzyme.  After the DNase event, the enzyme no longer cleaves
DNA, but forms a complex with S-adenosylmethionine and the DNA molecule which
it has restricted.  This complex actively hydrolyzes ATP to ADP and Pi.  The
modification enzyme exists in several forms, each having in some ratio the
polypeptides b and c, and a (molecular weight, 135,000).  In vitro
complementation analyses demonstrate that b and c from modification enzyme are
active in a restriction reaction when mixed with a factor that appears to be
the alpha-subunit.  These results are compatible with the genetic analysis of
the system.  The nature of the recognition site and the complexity of the
restriction reaction are discussed.

<>

<1>Linz, B., Windsor, H.M., Gajewski, J.P., Hake, C.M., Drautz, D.I., Schuster, S.C., Marshall, B.J.
<2>Helicobacter pylori genomic microevolution during naturally occurring transmission between adults.
<3>PLoS ONE
<4>8
<5>e82187
<6>2013
<7>The human gastric pathogen Helicobacter pylori is usually acquired during childhood and, in
the absence of treatment, chronic infection persists through most of the host's life.
However, the frequency and importance of H. pylori transmission between adults is
underestimated. Here we sequenced the complete genomes of H. pylori strains
that were transmitted between spouses and analysed the genomic changes. Similar to H. pylori
from chronic infection, a significantly high proportion of the determined 31 SNPs and 10
recombinant DNA fragments affected
genes of the hop family of outer membrane proteins, some of which are known to be adhesins. In
addition, changes in a fucosyltransferase gene modified the LPS component of the bacterial
cell surface, suggesting strong diversifying selection. In contrast, virulence factor genes
were not affected by the genomic changes. We propose a model of the genomic changes that are
associated with the transmission and adaptation of H. pylori to a new human host.

<>

<1>Linz, B., Windsor, H.M., McGraw, J.J., Hansen, L.M., Gajewski, J.P., Tomsho, L.P., Hake, C.M., Solnick, J.V., Schuster, S.C., Marshall, B.J.
<2>A mutation burst during the acute phase of Helicobacter pylori infection in humans and rhesus macaques.
<3>Nat. Commun.
<4>5
<5>4165
<6>2014
<7>The evolution rate and genetic changes that occur during chronic infection with Helicobacter
pylori have been analysed, but little is known about the genomic changes during the initial,
acute bacterial infection phase.  Here we analyse the rate and pattern of genome evolution in
H. pylori from the genomes of two input strains isolated from human volunteers with
asymptomatic infection, and the genomes of two output strains collected 20 and 44 days after
re-infection.  Similarly, we analyse genome evolution in bacteria from the genome sequences of
input and output strains sequentially taken after experimental infection of a rhesus macaque.
The estimated mutation rate reveals a mutation burst during the acute infection phase that is
over 10 times faster than the mutation rate during chronic infection, and orders of magnitude
faster than mutation rates in any other bacteria.  The elevated frequency of mutations in
outer membrane protein genes suggests that the mutation burst facilitates rapid host
adaptation of the bacteria.

<>

<1>Lio, P., Vannucci, M.
<2>Finding pathogenicity islands and gene transfer events in genome data.
<3>Bioinformatics
<4>16
<5>932-940
<6>2000
<7>Motivation: There is a growing literature on wavelet theory and wavelet methods showing
improvements on more classical techniques, especially in the contexts of smoothing and
extraction of fundamental components of signals. G+C patterns occur at different lengths
(scales) and, for this reason, G+C plots are usually difficult to interpret. Current methods
for genome analysis choose a window size and compute a Chi-squared statistics of the average
value for each window with respect to the whole genome. Results: Firstly, wavelets are used to
smooth G+C profiles to locate characteristic patterns in genome sequences. The method we use
is based on performing a Chi-squared statistics on the wavelet coefficients of a profile; thus
we do not need to choose a fixed window size, in that the smoothing occurs at a set of
different scales. Secondly, a wavelet scalogram is used as a measure for sequence profile
comparison; this tool is very general and can be applied to other sequence profiles commonly
used in genome analysis. We show applications to the analysis of Deinococcus radiodurans
chromosome I, of two strains of Helicobacter pylori (26695, J99) and two of Neisseria
meningitidis (serogroup B strain MC58 and serogroup A strain Z2491).  We report a list of loci
that have different G+C content with respect to the nearby regions; the analysis of N.
meningitidis serogroup B shows two new large regions with low G+C content that are putative
pathogenicity islands. Availability: Software and numerical results (profiles, scalograms,
high and low frequency components) for all the genome sequences analyzed are available upon
request from the authors.

<>

<1>Liolios, K. et al.
<2>Complete genome sequence of the gliding, heparinolytic Pedobacter saltans type strain (113).
<3>Standards in Genomic Sciences
<4>5
<5>30-40
<6>2011
<7>Pedobacter saltans Steyn et al. 1998 is one of currently 32 species in the genus  Pedobacter
within the family Sphingobacteriaceae. The species is of interest for
its isolated location in the tree of life. Like other members of the genus P.
saltans is heparinolytic. Cells of P. saltans show a peculiar gliding, dancing
motility and can be distinguished from other Pedobacter strains by their ability
to utilize glycerol and the inability to assimilate D-cellobiose. The genome
presented here is only the second completed genome sequence of a type strain from
a member of the family Sphingobacteriaceae to be published. The 4,635,236 bp long
genome with its 3,854 protein-coding and 67 RNA genes consists of one chromosome,
and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Liolios, K. et al.
<2>Complete genome sequence of Thermobispora bispora type strain (R51).
<3>Standards in Genomic Sciences
<4>2
<5>318-326
<6>2010
<7>Thermobispora bispora (Henssen 1957) Wang et al. 1996 is the type species of the  genus
Thermobispora. This genus is of great interest because it is strictly
thermophilic and because it has been shown for several of its members that the
genome contains substantially distinct (6.4% sequence difference) and
transcriptionally active 16S rRNA genes. Here we describe the features of this
organism, together with the complete genome sequence and annotation. This is the
second completed genome sequence of a member from the suborder
Streptosporangineae and the first genome sequence of a member of the genus
Thermobispora. The 4,189,976 bp long genome with its 3,596 protein-coding and 63
RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Liolos, K. et al.
<2>Complete genome sequence of the halophilic bacterium Spirochaeta africana type strain (Z-7692(T)) from the alkaline Lake Magadi in the East African Rift.
<3>Standards in Genomic Sciences
<4>8
<5>165-176
<6>2013
<7>Spirochaeta africana Zhilina et al. 1996 is an anaerobic, aerotolerant, spiral-shaped
bacterium that is motile via periplasmic flagella. The type strain
of the species, Z-7692(T), was isolated in 1993 or earlier from a bacterial bloom
in the brine under the trona layer in a shallow lagoon of the alkaline equatorial
Lake Magadi in Kenya. Here we describe the features of this organism, together
with the complete genome sequence, and annotation. Considering the pending
reclassification of S. caldaria to the genus Treponema, S. africana is only the
second 'true' member of the genus Spirochaeta with a genome-sequenced type strain
to be published. The 3,285,855 bp long genome of strain Z-7692(T) with its 2,817
protein-coding and 57 RNA genes is a part of the G enomic E ncyclopedia of B
acteria and A rchaea project.

<>

<1>Liou, M.L., Liu, C.C., Lu, C.W., Hsieh, M.F., Chang, K.C., Kuo, H.Y., Lee, C.C., Chang, C.T., Yang, C.Y., Tang, C.Y.
<2>Genome Sequence of Acinetobacter baumannii TYTH-1.
<3>J. Bacteriol.
<4>194
<5>6974
<6>2012
<7>Acinetobacter baumannii has emerged recently as a major cause of health care-associated
infections due to the extent of its antimicrobial resistance and
its propensity to cause large nosocomial outbreaks. Here we report the genome
sequence of Acinetobacter baumannii TYTH-1 isolated in Taiwan during 2008.

<>

<1>Lippmann, J.E., Wu, H., Fives-Taylor, P.M.
<2>Directed mutagenesis of the DNA-adenine-methyltransferase (DAM) gene in Actinobacillus actinomycetemcomitans (Aa) by conjugation.
<3>J. Dent. Res.
<4>81
<5>A-145
<6>2002
<7>Actinobacillus actinomycetemcomitans (Aa) is a slow growing, fastidious, Gram-negative,
capnophillic bacterium.  Of the over 500 different organisms in the oral cavity, Aa is one
which has been strongly implicated in juvenile and adult periodontis.  Aa manifests disease by
its ability to produce toxins, adhere to and invade epithelial cells.
DNA-adenine-methyltransferase (DAM) is generally associated with DNA modification, but it may
also act as a global regulator of gene expression in many organisms.  By adding methyl groups
to various sites along the genome, DAM alters the interactions of regulatory proteins with
their target genes.  Recently, virulence genes in Salmonella strains have been shown to be
modulated by DAM methylation.  Objectives:  To create a DAM minus strain of Aa SUNY465 for use
in evaluating the role of DAM in Aa virulence.  Methods: The dam gene was cloned and
sequenced.  The PCR-amplified gene was disrupted by insertion of an antibiotic cassette into a
unique blunt ended restriction site.  The disrupted gene was cloned into a conjugative plasmid
and transferred from E. coli SM10(lpir) to Aa.  Results: The insertional mutation resulted in
the loss of Dam methylation of the Aa genome as shown by restriction analysis using DpnI &
DpnII endonucleases, which cleave DNA at methylated and non-methylated sites, respectively.
Confirmation of the genetic mutation was confirmed by colony PCR and Southern blot analysis.
Conclusion: The chromosomal gene coding for DNA-adenine-methyltransferase of A.
actinomycetemcomitans has been cloned, sequenced, and disrupted.

<>

<1>Lippow, S., Lipovsek, D., Aha, P.M.
<2>Methods for engineering sequence-specific restriction endonucleases and their uses for nucleic acid assembly.
<3>International Patent Office
<4>WO 2008130629 A
<5>
<6>2008
<7>Aspects of the invention provide engineered endonucleases that are characterized by both a
long recognition sequence and specific cleavage outside of the recognition site.  Engineered
endonucleases of the invention are useful for manipulating long pieces of DNA.

<>

<1>Lippow, S.M., Aha, P.M., Parker, M.H., Blake, W.J., Baynes, B.M., Lipovsek, D.
<2>Creation of a type IIS restriction endonuclease with a long recognition sequence.
<3>Nucleic Acids Res.
<4>37
<5>3061-3073
<6>2009
<7>Type IIS restriction endonucleases cleave DNA outside their recognition sequences, and are
therefore particularly useful in the assembly of DNA
from smaller fragments. A limitation of type IIS restriction endonucleases
in assembly of long DNA sequences is the relative abundance of their
target sites. To facilitate ligation-based assembly of extremely long
pieces of DNA, we have engineered a new type IIS restriction endonuclease
that combines the specificity of the homing endonuclease I-SceI with the
type IIS cleavage pattern of FokI. We linked a non-cleaving mutant of
I-SceI, which conveys to the chimeric enzyme its specificity for an 18-bp
DNA sequence, to the catalytic domain of FokI, which cuts DNA at a defined
site outside the target site. Whereas previously described chimeric
endonucleases do not produce type IIS-like precise DNA overhangs suitable
for ligation, our chimeric endonuclease cleaves double-stranded DNA
exactly 2 and 6 nt from the target site to generate homogeneous, 5',
four-base overhangs, which can be ligated with 90% fidelity. We anticipate
that these enzymes will be particularly useful in manipulation of DNA
fragments larger than a thousand bases, which are very likely to contain
target sites for all natural type IIS restriction endonucleases.

<>

<1>Lippow, S.M., Aha, P.M., Parker, M.H., Blake, W.J., Baynes, B.M., Lipovsek, D.
<2>BIOT 7-Creation of a type IIS restriction endonuclease with a long recognition sequence.
<3>ACS Abstracts
<4>236
<5>0
<6>2008
<7>We have engineered a novel family of type IIS restriction endonucleases that combines the high
specificity of the homing endonuclease I-SceI with the type-IIS cleavage of FokI. Our hybrid
endonucleases feature a non-cleaving mutant of I-SceI linked to the catalytic domain of FokI
through a series of peptide linkers. We find that length and composition of the linker affect
the cleavage specificity of the hybrid enzymes. The endonucleases containing the FokI native
linker or a 20-residue synthetic linker are the most specific, cutting double-stranded DNA
exactly two and seven nucleotides from the recognition sequence to generate homogeneous, 5',
five-base overhangs. These two hybrid endonucleases generate DNA cleavage products that can be
ligated with greater than 80% fidelity.
We anticipate that these novel enzymes will be particularly useful for manipulating or
assembling kilobase and longer DNA fragments, which are likely to contain recognition sites
for all natural type IIS restriction endonucleases.

<>

<1>Lira, F., Hernandez, A., Belda, E., Sanchez, M.B., Moya, A., Silva, F.J., Martinez, J.L.
<2>Whole-Genome Sequence of Stenotrophomonas maltophilia D457, a Clinical Isolate and a Model Strain.
<3>J. Bacteriol.
<4>194
<5>3563-3564
<6>2012
<7>Stenotrophomonas maltophilia is an opportunistic pathogen with an environmental origin, and it
is an increasingly relevant cause of nosocomial infections. Here
we present the whole-genome sequence of S. maltophilia strain D457, a clinical
isolate that is being used as a model for studying antibiotic resistance in this
bacterial species.

<>

<1>Lisle, W., Dutta, C.F., Penner, M., Amitsur, M., Kaufmann, G., Firman, K.
<2>Phage T4-ancoded Stp alleviates the DNA restriction activity of EcoR124I endonuclease by affecting a critical step in the subunit assembly pathway.
<3>Mol. Biol. Today
<4>1
<5>57-64
<6>2000
<7>The tRNA(Lys)-specific anticodon nuclease is kept latent by virtue of a physical
association with the type IC DNA
restriction-modification enzyme EcoprrI. Previous work in vivo has
indicated that the phage T4-encoded polypeptide Stp inhibits EcoprrI
DNA restriction and activates anticodon nuclease. These studies also
suggested that EcoprrI is the immediate target of Stp. The presumptive
interaction was investigated here in vitro using the synthetic Stp-like
polypeptide Stp sub(2-26). Stp sub(2-26) inhibited the DNA restriction
activity of both EcoprrI and the related type IC DNA restriction enzyme
EcoR124I in crude cell extracts more effectively than with purified
EcoR124I. However, complementation with an EcoR124I-free crude extract
fully restored Stp function, suggesting the existence of any
co-operating host factor(s). Stp sub(2-26) also enhanced limited
proteolysis of the purified EcoR124I holoenzyme (HsdR sub(2)M sub(2)S),
but not the HsdM sub(2)S subassembly indicating that the R subunit is
important for the interaction and, perhaps, serves as the immediate
target of Stp. Stp sub(2-26) also produced a subtle shift of the
equilibrium between the Hsd R2 M2 S1 and Hsd R1 M2
S1 subassembly towards the latter form. Since only the
holoenzyme is active as a restriction endonuclease, this effect can
account for the ability of Stp to inhibit the DNA restriction enzyme
and activate the appended anticodon nuclease.

<>

<1>Little, E.J., Babic, A.C., Horton, N.C.
<2>Early interrogation and recognition of DNA sequence by indirect readout.
<3>Structure
<4>16
<5>1828-1837
<6>2008
<7>Control of replication, transcription, recombination and repair requires proteins capable of
finding particular DNA sequences in a background of a large excess of nonspecific sequences.
Such recognition can involve direct readout, with direct contacts to the bases of DNA, or in
some cases through the less well characterized indirect readout mechanisms. In order to
measure the relative contributions of direct and indirect readout by a sequence specific
endonuclease,  HincII, a mutant enzyme deficient in a direct contact, was characterized, and
surprisingly showed no loss of sequence specificity. The three dimensional crystal structure
shows the loss of most of the direct readout contacts to the DNA, possibly capturing an
early stage in target site recognition using predominately indirect readout to prescreen sites
before full sequence interrogation.

<>

<1>Little, E.J., Horton, N.C.
<2>DNA-induced conformational changes in type II restriction endonucleases: The structure of unliganded HincII.
<3>J. Mol. Biol.
<4>351
<5>76-88
<6>2005
<7>The 2.1 angstrom crystal structure of the unliganded type II restriction endonuclease, HincII,
is described. Although the asymmetric
unit contains only a single monomer, crystal lattice contacts bring two
monomers together to form a dimer very similar to that found in the DNA
bound form. Comparison with the published DNA bound structure reveals
that the DNA binding pocket is expanded in the unliganded structure.
Comparison of the unliganded and DNA liganded structures reveals a
simple rotation of subunits by 11 degrees each, or 22 degrees total, to
a more closed structure around the bound DNA. Comparison of this
conformational change to that observed in the other type II restriction
endonucleases where DNA bound and unliganded forms have both been
characterized, shows considerable variation in the types of
conformational changes that can occur. The conformational changes in
three can be described by a simple rotation of subunits, while in two
others both rotation and translation of subunits relative to one
another occurs. In addition, the endonucleases having subunits that
undergo the greatest amount of rotation upon DNA binding are found to
be those that distort the bound DNA the least, suggesting that DNA
bending may be less facile in dimers possessing greater flexibility (c)
2005 Elsevier Ltd. All rights reserved.

<>

<1>Little, J.W., Mount, D.W.
<2>Creating new restriction sites by silent changes in coding sequences.
<3>Gene
<4>32
<5>67-73
<6>1984
<7>We present methods for identifying a useful type of DNA site - one that can be
mutated to create a new restriction site within a coding region without
changing the amino acid sequence.  These "latent sites" are abundant - silent
mutations creating one of 44 different 6-bp or 8-bp recognition sites were
found at relatively high density, roughly one latent site per 9 bp, in the
eleven genes tested.  Our analysis suggests that site-directed mutagenesis can
be used to refashion coding sequences at will for flexible analysis.

<>

<1>Liu, B., Frostegard, A., Shapleigh, J.P.
<2>Draft genome sequences of five strains in the genus thauera.
<3>Genome Announcements
<4>1
<5>e00052-12
<6>2013
<7>Thauera species are members of the betaproteobacteria and are most noted for their ability to
metabolize aromatic compounds under anoxic conditions. Here, we
announce the draft genome sequences of five Thauera strains in an effort to
provide further genetic information as a resource for understanding the
ecological function of this environmentally important genus.

<>

<1>Liu, B., Hu, B., Zhou, Z., Guo, D., Guo, X., Ding, P., Feng, L., Wang, L.
<2>A novel non-homologous recombination-mediated mechanism for Escherichia coli unilateral flagellar phase variation.
<3>Nucleic Acids Res.
<4>40
<5>4530-4538
<6>2012
<7>Flagella contribute to the virulence of bacteria through chemotaxis, adhesion to
and invasion of host surfaces. Flagellar phase variation is believed to
facilitate bacterial evasion of the host immune response. In this study, the flnA
gene that encodes Escherichia coli H17 flagellin was examined by whole genome
sequencing and genetic deletion analysis. Unilateral flagellar phase variation
has been reported in E. coli H3, H47 and H17 strains, although the mechanism for
phase variation in the H17 strain has not been previously understood. Analysis of
phase variants indicated that the flagellar phase variation in the H17 strain was
caused by the deletion of an approximately 35 kb DNA region containing the flnA
gene from diverse excision sites. The presence of covalently closed
extrachromosomal circular forms of this excised 35 kb region was confirmed by the
two-step polymerase chain reaction. The deletion and complementation test
revealed that the Int1157 integrase, a tyrosine recombinase, mediates the
excision of this region. Unlike most tyrosine recombinases, Int1157 is suggested
to recognize diverse sites and mediate recombination between non-homologous DNA
sequences. This is the first report of non-homologous recombination mediating
flagellar phase variation.

<>

<1>Liu, C., Hu, J., Fang, X., Zhang, D., Chang, D., Wang, J., Li, T., Guo, Y., Dai, W., Liu, C.
<2>Genome Sequence of Pseudomonas aeruginosa Strain LCT-PA41, with Changed Metabolism after Space Flight.
<3>Genome Announcements
<4>2
<5>e01124-13
<6>2014
<7>To explore the effects of space flight on microorganisms, Pseudomonas aeruginosa  ATCC 27853
was sent into orbit for 398 h on the spacecraft ShenZhou VIII. Here,
we present the draft genome sequence of the P. aeruginosa strain LCT-PA41,
determined after space flight.

<>

<1>Liu, D., Zhou, Y., Naito, M., Yumoto, H., Li, Q., Miyake, Y., Liang, J., Shu, R.
<2>Draft Genome Sequence of Porphyromonas gingivalis Strain SJD2, Isolated from the  Periodontal Pocket of a Patient with Periodontitis in China.
<3>Genome Announcements
<4>2
<5>e01091-13
<6>2014
<7>Porphyromonas gingivalis strain SJD2 was isolated from subgingival plaque of a patient in
China with chronic periodontitis. Here, we report the draft genome of
this strain, with a size of 2,328,850 bp, average G+C content of 48.3%, and 2,020
predicted protein-coding sequences.

<>

<1>Liu, G., Deng, C., Song, L., Peng, Q., Zhang, J., Lereclus, D., Song, F.
<2>Draft Genome Sequence of Bacillus thuringiensis Strain LM1212, Isolated from the  Cadaver of an Oryctes gigas Larva in Madagascar.
<3>Genome Announcements
<4>2
<5>e00516-14
<6>2014
<7>We report the draft genome sequence of Bacillus thuringiensis strain LM1212, which
differentiates into crystal producers or spore formers during the
stationary phase. Availability of this genome sequence will facilitate the study
of spore formation, crystal formation, cell differentiation, and evolution of B.
thuringiensis.

<>

<1>Liu, G., Liu, B., Lin, N., Tang, W., Tang, J., Lin, Y.
<2>Genome Sequence of the Aerobic Bacterium Bacillus sp. Strain FJAT-13831.
<3>J. Bacteriol.
<4>194
<5>6633
<6>2012
<7>Bacillus sp. strain FJAT-13831 was isolated from the no. 1 pit soil of Emperor Qin's
Terracotta Warriors in Xi'an City, People's Republic of China. The isolate
showed a close relationship to the Bacillus cereus group. The draft genome
sequence of Bacillus sp. FJAT-13831 was 4,425,198 bp in size and consisted of
5,567 genes (protein-coding sequences [CDS]) with an average length of 782 bp and
a G+C value of 36.36%.

<>

<1>Liu, G., Liu, B., Tang, W., Che, J., Lin, Y., Zhu, Y., Su, M., Tang, J.
<2>Genome Sequence of Bacillus sp. Strain FJAT-14515.
<3>Genome Announcements
<4>2
<5>e01123-13
<6>2014
<7>We report the draft genome sequence of Bacillus sp. strain FJAT-14515. The genome is 5.44 Mb
in length. It covers 5,263 genes with an average length of 791 bp, has
a G+C value of 37.06%, and contains 67 tRNAs, 31 small RNAs, and 5 rRNA loci.

<>

<1>Liu, G., Ou, H.Y., Wang, T., Li, L., Tan, H., Zhou, X., Rajakumar, K., Deng, Z., He, X.
<2>Cleavage of Phosphorothioated DNA and Methylated DNA by the Type IV Restriction Endonuclease ScoMcrA.
<3>PLoS Genet.
<4>6
<5>e1001253
<6>2010
<7>Many taxonomically diverse prokaryotes enzymatically modify their DNA by replacing a
non-bridging oxygen with a sulfur atom at specific sequences.
The biological implications of this DNA S-modification
(phosphorothioation) were unknown. We observed that simultaneous
expression of the dndA-E gene cluster from Streptomyces lividans 66, which
is responsible for the DNA S-modification, and the putative Streptomyces
coelicolor A(3)2 Type IV methyl-dependent restriction endonuclease
ScoA3McrA (Sco4631) leads to cell death in the same host. A His-tagged
derivative of ScoA3McrA cleaved S-modified DNA and also Dcm-methylated DNA
in vitro near the respective modification sites. Double-strand cleavage
occurred 16-28 nucleotides away from the phosphorothioate links. DNase I
footprinting demonstrated binding of ScoA3McrA to the Dcm methylation
site, but no clear binding could be detected at the S-modified site under
cleavage conditions. This is the first report of in vitro endonuclease
activity of a McrA homologue and also the first demonstration of an enzyme
that specifically cleaves S-modified DNA.

<>

<1>Liu, G., Song, L., Shu, C., Wang, P., Deng, C., Peng, Q., Lereclus, D., Wang, X., Huang, D., Zhang, J., Song, F.
<2>Complete Genome Sequence of Bacillus thuringiensis subsp. kurstaki Strain HD73.
<3>Genome Announcements
<4>1
<5>e0008013
<6>2013
<7>Bacillus thuringiensis is a Gram-positive bacterium that produces intracellular protein
crystals toxic to a wide variety of insect larvae. We report the complete
genome sequence of Bacillus thuringiensis subsp. kurstaki strain HD73 from the
Centre OILB (Institut Pasteur, France), which belongs to serotype 3ab and is
toxic to lepidopteran larvae.

<>

<1>Liu, G., Zhang, W., Lu, C.
<2>Complete Genome Sequence of Streptococcus agalactiae GD201008-001, Isolated in China from Tilapia with Meningoencephalitis.
<3>J. Bacteriol.
<4>194
<5>6653
<6>2012
<7>This work describes a whole-genome sequence of Streptococcus agalactiae strain GD201008-001, a
pathogen causing meningoencephalitis in cultural tilapia in
China. The genome sequence provides opportunities to understand the piscine GBS
pathogenicity and its genetic basis associated with host tropism.

<>

<1>Liu, G.H., Liu, B., Tang, J.Y., Che, J.M., Zhu, Y.J., Su, M.X., Wang, J.P., Chen, Q.Q.
<2>Draft Genome Sequence of Bacillus fengqiuensis FJAT-14578, Isolated from a Soil Sample in China.
<3>Genome Announcements
<4>2
<5>e01126-14
<6>2014
<7>Here, we report the first high-quality draft genome sequence of Bacillus fengqiuensis
FJAT-14578, isolated from a soil sample collected from China. The
genome size was 5,569,389 bp, with a 40.93 mol% G+C content. The number of tRNAs
was 69 and of rRNAs was 10 (5S, 16S, and 23S).

<>

<1>Liu, H., Qiu, H., Zhao, W., Cui, Z., Ibrahim, M., Jin, G., Li, B., Zhu, B., Xie, G.L.
<2>Genome Sequence of the Plant Pathogen Pseudomonas syringae pv. panici LMG 2367.
<3>J. Bacteriol.
<4>194
<5>5693-5694
<6>2012
<7>Pseudomonas syringae pv. panici is a phytopathogenic bacterium causing brown stripe disease in
economically important crops worldwide. Here, we announce the
draft genome sequence of Pseudomonas syringae pv. panici LMG2367 to provide
further valuable insights for comparison of the pathovars among species
Pseudomonas syringae.

<>

<1>Liu, H., Wu, Z., Li, M., Zhang, F., Zheng, H., Han, J., Liu, J., Zhou, J., Wang, S., Xiang, H.
<2>Complete Genome Sequence of Haloarcula hispanica, a Model Haloarchaeon for Studying Genetics, Metabolism, and Virus-Host Interaction.
<3>J. Bacteriol.
<4>193
<5>6086-6087
<6>2011
<7>Haloarcula hispanica is an extremely halophilic archaeon that has an unusually low restriction
barrier and is therefore significant for
studying archaeal genetics, metabolism, and virus-host interactions. Here
we report the complete genome sequence (3,890,005 bp) of H. hispanica
strain CGMCC 1.2049, consisting of two chromosomes and one megaplasmid.

<>

<1>Liu, J., Cheng, A., Bangayan, N.J., Barnard, E., Curd, E., Craft, N., Li, H.
<2>Draft Genome Sequences of Propionibacterium acnes Type Strain ATCC6919 and Antibiotic-Resistant Strain HL411PA1.
<3>Genome Announcements
<4>2
<5>e00740-14
<6>2014
<7>Propionibacterium acnes is a major skin commensal and is associated with acne vulgaris, the
most common skin disease. Here we report the draft genome sequences
of two P. acnes strains, the type strain ATCC6919 and an antibiotic-resistant
strain, HL411PA1.

<>

<1>Liu, J., Xue, Y., Meng, Y., Zhao, X., Cai, Y.
<2>Expression detection and location analysis of BstNI isoschizomer restriction-modification system gene.
<3>Wei Sheng Wu Xue Bao
<4>39
<5>209-214
<6>1999
<7>Some genetic markers of E. coli HB101 and JM110 were identified, two bacterial strains were
used as recipients respectively to detect the expression of a restriction endonuclease gene
and a methylase gene of BstNI isoschizomer restriction-modification system.  DNA fragment
containing the R-M genes was deleted unidirectionally with exoIII and 23 deletion subclones
were obtained.  According to the enzyme activity of each subclone, R and M gene were located
respectively at the regions of 0.2 to 1.4 kb and 1.5 to 3.3 kb from cloning site PstI.
Analysis showed that the R-M system belongs to type II, two genes are controlled by the
different promoters; the recognition sequence of this system is the same as that of
DNA-cytosine methyltransferase (Dcm), the latter's methylation function can resist the R
enzyme.  It was interesting that the recombinant plasmid with an R+M- genotype appeared to be
lethal to dcm+ hosts yet.  This indicated that the M gene closely linking to R gene is of
critical importance for the existence of the R-M system in process of evolution.

<>

<1>Liu, J., Zhao, X., Meng, Y., Shen, J., Xue, Y., Shi, S., Cai, Y.
<2>Expression and deletion analysis of EcoRII endonuclease and methylase gene.
<3>Chin. Med. Sci. J.
<4>16
<5>200-203
<6>2001
<7>Objective. To clone complete EcoRII restriction endonuclease gene (ecoRIIR) and
methyltransferase gene (ecoRIIM) in one vector and to analyze the coordinated expression of
this whole R-M system.  Methods. Unidirectional deletion subclones were constructed with
ExoIII. ecoRIIR/M genes were preliminarily located in the cloned fragment according to the
enzyme activities of subclones.  Exact deletion sites were determined by sequencing, and
transcriptional start sites were determined by S1 mapping.  Results. The DNA fragment which
was cloned into pBluescript SK + contained intact ecoRIIR gene and ecoRIIM gene, and two
transcriptional start sites of ecoRIIIR gene were determined.  132bp to 458bp from 3' end of
ecoRIIR gene are indispensable to enzyme activities and deletion of 202bp from 3' end of
ecoRIIM gene made enzyme lose the capability in DNA protection to resist specific cut with
EcoRII endonuclease (EcoRII.R).  Deletion of the coding and flanking sequences of one gene did
not affect the expression of the other gene, and the recombinants only containing ecoRIIR gene
appeared to be lethal to dcm+ host.  Conclusion. ecoRIIIM gene linking closely to ecoRIIR gene
is very important for the existence of the R-M system in process of evolution, but the key to
control EcoRII R-M order may not exist in transcriptional level.

<>

<1>Liu, J., Zhao, X., Meng, Y., Shen, J., Xue, Y., Shi, S., Cai, Y.
<2>Expression and deletion analysis of EcoRII endonuclease and methylase gene.
<3>Zhongguo Yi Xue Ke Xue Yuan Xue Bao
<4>22
<5>111-114
<6>2000
<7>Objective. To clone the complete EcoRII restriction endonuclease gene (ecoRIIR) and
methyltransferase (ecoRIIM) gene into one vector and to analyze the expression of the whole
system.  Methods. Unidirectional deletion subclones, constructed with ExoIII, of the ecoRIIR/M
genes were preliminarily located in the cloned fragments according to the enzyme activities of
each subclone, exact deletion sites were determined by sequencing, and transcriptional start
sites were mapped by S1 mapping.  Results.  The DNA fragment which was cloned into pBluescript
SK+ contained the complete ecoRIIR gene and ecoRIIM gene, there are two transcriptional start
sites in the ecoRIIR gene, 132 bp to 458 bp from 3' end of the ecoRIIR gene are indispensable
to enzyme activities and deletion of 202 bp from the 3' end of the ecoRIIM gene made it lose
the capability to resist specific cleavage by EcoRIIR enzyme, deletion of coding region and
flanking sequence of one gene did not affect the expression of the other gene, the recombinant
only containing ecoRIIR gene appeared to be lethal to dcm+ host.  Conclusions. ecoRIIM gene
closely linked to the ecoRIIR gene was very important for the existence of the R-M system in
the process of evolution, but the key to make sure that the EcoRIIR enzyme is produced later
than the EcoRIIM enzyme does not exist at the transcriptional level.

<>

<1>Liu, J., Zhou, Q., Ibrahim, M., Liu, H., Jin, G., Zhu, B., Xie, G.
<2>Genome Sequence of the Biocontrol Agent Microbacterium barkeri Strain 2011-R4.
<3>J. Bacteriol.
<4>194
<5>6666-6667
<6>2012
<7>Microbacterium barkeri strain 2011-R4 is a Gram-positive epiphyte which has been  confirmed as
a biocontrol agent against several plant pathogens in our previous
studies. Here, we present the draft genome sequence of this strain, which was
isolated from the rice rhizosphere in Tonglu city, Zhejiang province, China.

<>

<1>Liu, J.-Y., Zhao, X.-J., Cai, Y.Y., Shen, J., Meng, Y.
<2>High expression and purification of EcoRII restriction endonuclease.
<3>Chin. J. Biochem. Mol. Biol.
<4>16
<5>417-420
<6>2000
<7>EcoRII was the first restriction endonuclease ever found
requiring the cooperative interaction with the least two DNA sites for
digestion activity.  To study the specific activity, EcoRII was purified
from hyperexpression engineering bacteria in which the specific
expression products increased to about 20% of total cellular protein.
By using chromatography on DEAE-cellulose column, phosphocellulose
column and FPLC of Resource Q, the enzyme was purified from soluble
protein fraction.  The inclusion bodies were solved and renatured, and
the enzyme was purified from this part of protein with higher specific
activity by using FPLC of Resource Q.  Detection showed that the enzyme
was purified to homogeneity and was free of detectable contamination by
other DNase (exo and endo).

<>

<1>Liu, K., Wang, Y.F., Cantemir, C., Muller, M.T.
<2>Endogenous assays of DNA methyltransferases: Evidence for differential activities of DNMT1, DNMT2, and DNMT3 in mammalian cells in vivo.
<3>Mol. Cell. Biol.
<4>23
<5>2709-2719
<6>2003
<7>While CpG methylation can be readily analyzed at the DNA sequence level in wild-type and
mutant cells, the actual DNA (cytosine-5) methyltransferases
(DNMTs) responsible for in vivo methylation on genomic DNA are less
tractable. We used an antibody-based method to identify specific
endogenous DNMTs (DNMT1, DNMT1b, DNMT2, DNMT3a, and DNMT3b) that stably
and selectively bind to genomic DNA containing 5-aza-2'-deoxycytidine
(aza-dC) in vivo. Selective binding to aza-dC-containing DNA suggests that
the engaged DNMT is catalytically active in the cell. DNMT1b is a splice
variant of the predominant maintenance activity DNMT1, while DNMT2 is a
well-conserved protein with homologs in plants, yeast, Drosophila, humans,
and mice. Despite the presence of motifs essential for transmethylation
activity, catalytic activity of DNMT2 has never been reported. The data
here suggest that DNMT2 is active in vivo when the endogenous genome is
the target, both in human and mouse cell lines. We quantified relative
global genomic activity of DNMT1, -2, -3a, and -3b in a mouse
teratocarcinoma cell line. DNMT1 and -3b displayed the greatest in vivo
binding avidity for aza-dC-containing genomic DNA in these cells. This
study demonstrates that individual DNMTs can be tracked and that their
binding to genomic DNA can be quantified in mammalian cells in vivo. The
different DNMTs display a wide spectrum of genomic DNA-directed activity.
The use of an antibody-based tracking method will allow specific DNMTs and
their DNA targets to be recovered and analyzed in a physiological setting
in chromatin.

<>

<1>Liu, L., Gao, P., Chen, G., Wang, L.
<2>Draft Genome Sequence of Cellulose-Digesting Bacterium Sporocytophaga myxococcoides PG-01.
<3>Genome Announcements
<4>2
<5>e01154-14
<6>2014
<7>Sporocytophaga myxococcoides, a Gram-negative bacterium isolated from soil, is an efficient
hydrolyzer of crystalline cellulose. Here, we report its draft genome
sequence, which may provide important genetic information regarding the
cellulolytic and hemicellulolytic enzymes that contribute to the
cellulose-degrading abilities of this bacterium.

<>

<1>Liu, L., Li, Y., Zhang, J., Zhou, Z., Liu, J., Li, X., Zhou, J., Du, G., Wang, L., Chen, J.
<2>Complete Genome Sequence of the Industrial Strain Ketogulonicigenium vulgare WSH-001.
<3>J. Bacteriol.
<4>193
<5>6108-6109
<6>2011
<7>Ketogulonicigenium vulgare is an industrial organism commonly used in the vitamin C industry.
Here, we report the finished, annotated, and compared
3.28-Mbp high-quality genome sequence of Ketogulonicigenium vulgare
WSH-001, a 2-keto-l-gulonic acid-producing industrial strain stocked in
our laboratory.

<>

<1>Liu, L., Li, Y., Zhang, J., Zou, W., Zhou, Z., Liu, J., Li, X., Wang, L., Chen, J.
<2>Complete Genome Sequence of the Industrial Strain Bacillus megaterium WSH-002.
<3>J. Bacteriol.
<4>193
<5>6389-6390
<6>2011
<7>Bacillus megaterium, an industrial strain, has been widely used in protein production and the
vitamin C industry. Here we reported a finished,
annotated, and compared 4.14-Mbp high-quality genome sequence of B.
megaterium WSH-002, which is the companion strain for Ketogulonicigenium
vulgare in the vitamin C industry and is stocked in our laboratory.

<>

<1>Liu, L., Santi, D.V.
<2>Mutation of asparagine 229 to aspartate in thymidylate synthase converts the enzyme to a deoxycytidylate methylase.
<3>Biochemistry
<4>31
<5>5100-5104
<6>1992
<7>The conserved Asn 229 of thymidylate synthase (TS) forms a cyclic hydrogen bond network with
the 3-NH and 4-O of the nucleotide substrate dUMP. The Asn 229 to Asp mutant of Lactobacillus
casei thymidylate synthase (TS N229D) has been prepared, purified, and investigated.
Steady-state kinetic parameters of TS N229D show 3.5- and 10-fold increases in the Km values
of CH2H4 folate and dUMP, respectively, and a 1000-fold decrease in kcat. Most important, the
Asp 229 mutation changes the substrate specificity of TS to an enzyme which recognizes and
methylates dCMP in preference to dUMP. With TS N229D the Km for dCMP is about 3-fold higher
than for dUMP, and the Km for CH2H4 folate is increased about 5-fold; however, the kcat for
dCMP methylation is 120-fold higher than that for dUMP methylation. Specificity for dCMP
versus dUMP, as measured by Kcat/Km, changes from negligible with wild-type TS to about a
40-fold increase with TS N229D. TS N229D reacts with CH2H4 folate and FdUMP or FdCMP to form
ternary complexes which are analogous to the TS-FdUMP-CH2H4 folate complex. From what is known
of the mechanism and structure of TS, the dramatic change in substrate specificity of TS N229D
is proposed to involve a hydrogen bond network between Asp 229 and the 3-N and 4-NH2 of the
cytosine mechanism is proposed for related enzymes which catalyze one-carbone transfers to
cytosine heterocycles.

<>

<1>Liu, L.J., You, X.Y., Zheng, H.J., Wang, S.Y., Jiang, C.Y., Liu, S.J.
<2>Complete genome sequence of Metallosphaera cuprina, a metal sulfide-oxidizing archaeon from hot spring.
<3>J. Bacteriol.
<4>193
<5>3387-3388
<6>2011
<7>The genome of metal sulfide oxidizing, thermoacidiphilic Metallosphaera cuprina Ar-4 has been
completely sequenced and annotated. Originally
isolated from a sulfuric hotspring, strain Ar-4 grows optimally at 65
degrees C and pH of 3.5. The M. cuprina genome has a 1,840,348 bp circular
chromosome (2029 ORFs) and is 16% smaller than the previously sequenced
Metallosphaera sedula genome. Compared to the M. sedula genome, there are
no counterpart genes in the M. cuprina genome for about 480 ORFs in the M.
sedula genome, of which 243 ORFs are annotated as hypothetical proteins.
Still, there are 235 ORFs uniquely occurring in M. cuprina. Genome
annotation supports that M. cuprina lives a facultative life on CO(2) and
organics, and obtains energy from oxidation of sulfidic ores and reduced
inorganic sulfuric compounds.

<>

<1>Liu, M., Chen, S.
<2>Draft Genome Sequence of Vibrio parahaemolyticus V110, Isolated from Shrimp in Hong Kong.
<3>Genome Announcements
<4>1
<5>e00300-13
<6>2013
<7>We report the whole-genome sequence of a tdh- and trh-negative Vibrio parahaemolyticus strain,
V110, from shrimp. The major difference of V110 from
clinical strains was its lack of the type III secretion system T3SS2, a key
component of virulence. Further sequence comparison can shed light on the
pathogenesis of V. parahaemolyticus.

<>

<1>Liu, M.S., Gingery, M., Doulatov, S.R., Liu, Y.C., Hodes, A., Baker, S., Davis, P., Simmonds, M., Churcher, C., Mungall, K., Quail, M.A., Preston, A., Harvill, E.T., Maskell, D.J., Eiserling, F.A., Parkhill, J., Miller, J.F.
<2>Genomic and genetic analysis of Bordetella bacteriophages encoding reverse transcriptase-mediated tropism-switching cassettes.
<3>J. Bacteriol.
<4>186
<5>1503-1517
<6>2004
<7>Liu et al. recently described a group of related temperate bacteriophages that infect
Bordetella subspecies and undergo a unique
template-dependent, reverse transcriptase-mediated tropism switching
phenomenon (Liu et al., Science 295:2091-2094, 2002). Tropism switching
results from the introduction of single nucleotide substitutions at
defined locations in the VR1 (variable region 1) segment of the mtd
(major tropism determinant) gene, which determines specificity for
receptors on host bacteria. In this report, we describe the complete
nucleotide sequences of the 42.5- to 42.7-kb double-stranded DNA
genomes of three related phage isolates and characterize two additional
regions of variability. Forty-nine coding sequences were identified. Of
these coding sequences, bbp36 contained VR2 (variable region 2), which
is highly dynamic and consists of a variable number of identical 19-bp
repeats separated by one of three 5-bp spacers, and bpm encodes a DNA
adenine methylase with unusual site specificity and a homopolymer tract
that functions as a hotspot for frameshift mutations. Morphological and
sequence analysis suggests that these Bordetella phage are genetic
hybrids of P22 and T7 family genomes, lending further support to the
idea that regions encoding protein domains, single genes, or blocks of
genes are readily exchanged between bacterial and phage genomes.
Bordetella bacteriophages are capable of transducing genetic markers in
vitro, and by using animal models, we demonstrated that lysogenic
conversion can take place in the mouse respiratory tract during
infection.

<>

<1>Liu, P., Li, P., Jiang, X., Bi, D., Xie, Y., Tai, C., Deng, Z., Rajakumar, K., Ou, H.Y.
<2>Complete Genome Sequence of Klebsiella pneumoniae subsp. pneumoniae HS11286, a Multidrug-Resistant Strain Isolated from Human Sputum.
<3>J. Bacteriol.
<4>194
<5>1841-1842
<6>2012
<7>Klebsiella pneumoniae is an important pathogen commonly associated with opportunistic
infections. Here we report the genome sequence of a strain,
HS11286, isolated from human sputum in 2011 in Shanghai, China. It contains one
chromosome (5.3 Mb), three multidrug resistance plasmids ( approximately 110 kb),
including a carbapenemase producer, and three small plasmids ( approximately 3
kb).

<>

<1>Liu, P., Yang, X.-Hai., Wang, Q., Huang, J., Liu, J.-Bo., Zhu, Y., He, L.-L., Wang, Ke.-Min.
<2>Sensitive detection of DNA methyltransferase activity based on rolling circle amplification technology.
<3>Chin. Chem. Lett.
<4>25
<5>1047-1051
<6>2014
<7>This work develops a fluorescence approach for sensitive detection of DNA methyltransferase
activity based on endonuclease and rolling circle amplification (RCA) technique. In the
presence of DNA adenine methylation (Dam) MTase, the methylation-responsive sequence of
hairpin probe is methylated and cleaved by the methylation-sensitive restriction endonuclease
Dpn I. The products cleaved by restriction endonuclease Dpn I then function as a signal primer
to initiate RCA reaction by hybridizing with the circular DNA template. Each RCA product
containing thousands of repeated sequences might hybridize with a large number of molecular
beacons (detection probes), resulting in an enhanced fluorescence signal. In the absence of
Dam MTase, neither methylation/cleavage nor RCA reaction can be initiated and no fluorescence
signal is observed. The proposed method exhibits a dynamic range from 0.5 U/mL to 30 U/mL and
a detection limit of 0.18 U/mL. This method can be used for the screening of antimicrobial
drugs and has a great potential to be further applied in early clinical diagnosis.

<>

<1>Liu, P.Y., Huang, Y.T., Lin, S.Y., Chang, G.C., Chen, J.W.
<2>Draft Genome Sequence of the Serratia marcescens Strain VGH107, a Taiwanese Clinical Isolate.
<3>Genome Announcements
<4>1
<5>e00249-13
<6>2013
<7>Serratia marcescens, a member of the family Enterobacteriaceae, is the causative  agent of
various types of wound infections. We report a high-quality draft genome
sequence of S. marcescens strain VGH107, which was isolated from a patient with
an infection from a snakebite wound.

<>

<1>Liu, Q., Chen, X., Zhao, X., Chen, Y., Chen, D.
<2>The effect of methylation outside the recognition sequence of restriction endonuclease PvuII on its cleavage efficiency.
<3>Gene
<4>113
<5>89-93
<6>1992
<7>This study is to extend our earlier observation that Dam and Dcm methylation outside the PvuII
recognition sequence inhibited PvuII cleavage in one of the three PvuII sites of pGEM4Z-ras
DNA. In this paper, a new recombinant plasmid DNA, pGEM4-SV40ori-anti-ras, was constructed
which has only two PvuII sites, I and II. The Dam and Dcm-methylated and unmethylated DNAs
were produced in Escherichia coli and linearized by ScaI. The DNA molecules were digested with
different amounts of PvuII. The results show that by comparing the DNA fragment number and
intensity of the partial and final products in agarose gels, PvuII site I on the methylated
DNA molecule was digested four- to eight-fold more slowly than site II. In the unmethylated
plasmid DNA, the two PvuII sites were cleaved at about the same rate. The difference was
caused only by methylation of Dam and Dcm sites outside the PvuII recognition sequence. A
methylated Dam site immediately adjacent to the less efficiently cut PvuII site I may be
responsible for the inhibitory effect. We suggest that a new parameter, involving methylation
of sites outside the recognition sequence, be considered in kinetic experiments on cleavage.

<>

<1>Liu, Q., Derbyshire, V., Balfort, M., Edgell, D.R.
<2>Distance determination by GIY-YIG intron endonucleases: Discrimination between repression and cleavage functions.
<3>Nucleic Acids Res.
<4>34
<5>1755
<6>2006
<7>GIY-YIG homing endonucleases are modular proteins, with conserved N-terminal catalytic domains
connected by linkers to C-terminal DNA-binding domains. I-TevI, the T4 phage GIY-YIG intron
endonuclease, functions both in promoting td intron homing, and in acting as a transcriptional
autorepressor. Repression is achieved by binding to an operator, which is cleaved at 100-fold
reduced efficiency relative to the intronless homing site. The linker includes a zinc finger,
which functions in distance determination, to constrain the catalytic domain to cleave the
homing site at a fixed position. Here we show that I-BmoI, a related GIY-YIG endonuclease
lacking a zinc finger, also possesses some cleavage distance discrimination. Furthermore,
hybrid endonucleases constructed by swapping the domains of I-BmoI and I-TevI are active,
precise and demonstrate that features other than the zinc finger facilitate distance
determination. Most importantly, I-TevI zinc finger mutants cleave the operator more
efficiently than the homing site, the converse of wild-type protein. These results are
consistent with the zinc finger acting as a measuring device, directing efficient cleavage of
the homing site to promote intron mobility, while reducing cleavage at the operator to ensure
transcriptional autorepression and phage viability.

<>

<1>Liu, Q.Q., Dansereau, J.T., Puttamadappa, S.S., Shekhtman, A., Derbyshire, V., Belfort, M.
<2>Role of the interdomain linker in distance determination for remote cleavage by homing endonuclease I-TevI.
<3>J. Mol. Biol.
<4>379
<5>1094-1106
<6>2008
<7>I-TevI is a modular intron-encoded endonuclease, consisting of an N-terminal catalytic domain
and a C-terminal DNA-binding domain, joined
by a 75 amino acid linker. This linker can be divided into three
regions, starting at the N terminus: the deletion-intolerant (DI)
region; the deletion-tolerant (DT) region; and a zinc finger, which
acts as a distance determinant for cleavage. To further explore linker
function, we generated deletion and substitution mutants that were
tested for their preference to cleave at a particular distance or at
the correct sequence. Our results demonstrate that the I-TevI linker is
multi-functional, a property that sets it apart from junction sequences
in most other proteins. First, the linker DI region has a role in
I-TevI cleavage activity. Second, the DT linker region participates in
distance determination, as evident from DT mutants that display a
phenotype similar to that of the zinc-finger mutants in their selection
of a cleavage site. Finally, NMR analysis of a freestanding 56 residue
linker segment showed an unstructured stretch corresponding to the DI
region and a portion of the DT region, followed by a beta-strand
corresponding to the remainder of the DT region and containing a key
distance-determining arginine, R129. Mutation of this arginine to
alanine abolished distance determination and disrupted the beta-strand,
indicating that the structure of the DT linker region has a role in
cleavage at a fixed distance.

<>

<1>Liu, S., Wang, Y., Xu, J., Li, Y., Guo, J., Ke, Y., Yuan, X., Wang, L., Du X, W.Z., Huang, L., Zhang, N., Chen, Z.
<2>Genome Sequence of an OXA23-Producing, Carbapenem-Resistant Acinetobacter baumannii Strain of Sequence Type ST75.
<3>J. Bacteriol.
<4>194
<5>6000-6001
<6>2012
<7>The increase of Acinetobacter baumannii resistance to carbapenems is of great concern. OXA23
is one of the most prevalent carbapenemases of A. baumannii that
causes outbreaks. Here, we announce the genome sequence of an OXA23-producing A.
baumannii strain assigned ST75, a newly emerged sequence type harboring
carbapenemase.

<>

<1>Liu, S.L., Hessel, A., Sanderson, K.E.
<2>Genomic mapping with I-CeuI, an intron-encoded endonuclease specific for genes for ribosomal RNA, in Salmonella ssp., Escherichia coli, and other bacteria.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>6874-6878
<6>1993
<7>Construction of physical maps of genomes by pulsed-field gel electrophoresis requires enxymes
which cut the genome into an analyzable number of fragments; most produce too many fragments.
The enzyme I-CeuI, encoded by a mobile intron in the chloroplast 23S ribosomal RNA (rrl) gene
of Chlamydomonas eugametos, cuts a 26-bp site in the rrl gene. This enzyme digests DNA of
Salmonella typhimurium at seven sites, each corresponding to one of the rrl genes of the rrn
operons, but at no other site. These seven fragments were located on the previously determined
XbaI physical map, and the I-CeuI sites, and thus the rrn genes of S. typhimurium, were mapped
on the 4800-kb chromosome. Escherichia coli K-12 also yields seven fragments of sizes similar
to those of S. typhimurium, indicating conservation of rrn genes and their location, and a
chromosome size of 4600 kb. The sizes of the E. coli fragments are close to the size predicted
from restriction maps and nucleotide sequence. The I-CeuI maps of Salmonella typhi were
deduced after digesting genomic DNA with I-CeuI and probing with DNA of S. typhimurium; the
data indicated strong conservation of rrn gene number and position and genome sizes up to 4950
kb. Digestion of DNA of other bacteria (species of Haemophilus, Neisseria, Proteus, and
Pasteurella) suggested that only rrn genes are cut in all these species. I-CeuI digestion
followed by pulsed-field gel electrophoresis is a powerful tool for determining genome
structure and evolution.

<>

<1>Liu, T.-T., Liu, J.-F., Wang, W.-X., Wang, H., Wang, Z.-L., Zeng, Z.-J., Yan, W.-Y.
<2>Cloning and expression profiling of the DNA methyltransferase Dnmt3 gene in the Chinese honeybee, Apis cerana cerana (Hymenoptera: Apidae).
<3>Acta Entomologica Sinica
<4>55
<5>284-290
<6>2012
<7>To explore the pattern of methylation in the Chinese honeybee, Apis cerana cerana, the DNA
methyltransferase 3 (Dnmt3) gene in A. cerana
cerana was cloned by using RT-PCR, and the quantitative analysis of the
expression level of Dnmt3 mRNA in different developmental stages of
worker (4 d-old pupa, 1, 7 and 30 d-old workers, and laying worker) and
queen (4 d-old pupa, 1 d-old queen and laying queen) were conducted
using real-time qPCR. The full-length cDNA of Dnmt3 gene (GenBank
accession no. JQ740768) is 2 277 bp, encoding 758 amino acids, and the
predicted MW and pI are 88. 24 kD and 7. 85, respectively. Based on the
comparison of the domain and the phylogenetic tree of the amino acids
of Dnmt3 in A. cerana cerana and other species, the sequence obtained
has up to 99% identity with that of A. mellifera. The Dnmt3 transcript
was clearly detected in different developmental stages of worker and
queen, and it was expressed significantly higher in 30 d-old worker
than in 1 d- and 7 d-old worker (P < 0.05), while no difference existed
between 1 d- and 7 d-old worker (P > 0. 05). The Dnmt3 transcript was
expressed higher in queen pupae than in worker pupae (P < 0.05), and
was higher in 1 d-old queen than in 1 d-old worker (P <0. 05). The
expression level of Dnmt3 gene between laying worker and laying queen
had no significant difference. The results suggested that Dnmt3 may be
involved in the division of labor in workers and ovary development in
honeybees.

<>

<1>Liu, W.
<2>Fluorescence studies of EcoRI restriction endonuclease structure and dynamics.
<3>Diss. Abstr.
<4>58
<5>681
<6>1997
<7>Sequence specific protein-DNA interaction involves changes in protein conformation and
dynamics.  Research work involved in this thesis is the study of structure and dynamics of
EcoRI restriction endonuclease N-termini.  The N-termini of EcoRI endonuclease has been shown
to be essential for DNA cleavage and also stabilize EcoRI-DNA complex.  But they are not
resolved in the X-ray crystal structure.  An Asn3Cys mutation was made at the N-termini by
site-directed mutagenesis and the mutant was subjected to cysteine crosslinking, pyrene
labeling and fluorescence studies.  Chemical crosslinking of Asn3Cys mutant and steady-state
fluorescence studies of pyrene-labeled mutant indicated that the N-termini are in close
proximity and probably involved in the dimer interface.  Time resolved fluorescence
measurements revealed dynamics of the N-termini by examining the dissociation and reformation
of pyrene excimers as well as the monomer spectral shifts caused by N-terminal segment
movements.  Fluorescence anisotropy decay analysis indicated the N-termini are on the protein
surface and not totally disoriented in solution.  Substrate DNA binding, however, causes the
N-termini to be more mobile without affecting the proximity relationship.  Fluorescence energy
transfer experiments were carried out using a double-mutant Asn3Cys,Trp104Tyr.  The through
space distance between the fluorescent labels, MIANS or 1,5 -IAEDANS, and Trp246 is around 30
Angstroms.  Substrate DNA binding decreased the distance to 26 Angstroms but cofactor Mg2+ ion
did not cause any distance change.  Single Trp246 fluorescence in the double mutant also
revealed possible conformational changes upon substrate DNA or cofactor binding.  Experimental
evidence indicates that the N-termini are located at the dimer interface but distal to the DNA
binding site.  The findings provide further insight into the function of the N-termini of
EcoRI endonuclease.

<>

<1>Liu, W., Chen, Y., Watrob, H., Bartlett, S.G., Jen-Jacobson, L., Barkley, M.D.
<2>N-termini of EcoRI restriction endonuclease dimer are in close proximity on the protein surface.
<3>Biochemistry
<4>37
<5>15457-15465
<6>1998
<7>The N-terminal region of EcoRI endonuclease is essential for cleavage yet is invisible in the
2.5 A crystal structure of endonuclease-DNA. We used site-directed fluorescence spectroscopy
and chemical cross-linking to locate the N-terminal region and assess its flexibility in the
absence and presence of DNA substrate. The second amino acid in each subunit of the homodimer
was replaced with cysteine and labeled with pyrene or reacted with bifunctional cross-linkers.
The broad absorption spectra and characteristic excimer emission bands of pyrene-labeled
muteins indicated stacking of the two pyrene rings in the homodimer. Proximity of N-terminal
cysteines was confirmed by disulfide bond formation and chemical cross-linking. The dynamics
of the N-terminal region were determined from time-resolved emission anisotropy measurements.
The anisotropy decay had two components: a fast component with rotational correlation time of
0.3-3 ns representing probe internal motions and a slow component with 50-100 ns correlation
time representing overall tumbling of the protein conjugate. We conclude that the N-termini
are close together at the dimer interface with limited flexibility. Binding of Mg2+ cofactor
or DNA substrate did not affect the location or flexibility of the N-terminal region as sensed
by pyrene fluorescence and cross-linking, indicating that substrate binding is not accompanied
by folding or unfolding of the N-terminus.

<>

<1>Liu, W., Fang, L., Li, S., Li, Q., Zhou, Z., Feng, Z., Luo, R., Shao, G., Wang, L., Chen, H., Xiao, S.
<2>Complete Genome Sequence of Mycoplasma hyorhinis Strain HUB-1.
<3>J. Bacteriol.
<4>192
<5>5844-5845
<6>2010
<7>Mycoplasma hyorhinis is generally considered a swine pathogen yet is most commonly found
infecting laboratory cell lines. An increasing body of
evidence suggests that chronic infections with M. hyorhinis may cause
oncogenic transformation. Here, we announce the complete genome sequence
of M. hyorhinis strain HUB-1.

<>

<1>Liu, W., Feng, Z., Fang, L., Zhou, Z., Li, Q., Li, S., Luo, R., Wang, L., Chen, H., Shao, G., Xiao, S.
<2>Complete Genome Sequence of Mycoplasma hyopneumoniae Strain 168.
<3>J. Bacteriol.
<4>193
<5>1016-1017
<6>2011
<7>Mycoplasma hyopneumoniae strain 168, a pathogenic strain prevalent in China, was isolated in
1974. Although this strain has been widespread for
a long time, the genome sequence had not been determined. Here, we
announce the complete genome sequence of M. hyopneumoniae strain 168.

<>

<1>Liu, W., Jing, Z., Ou, Q., Cui, B., He, Y., Wu, Q.
<2>Complete Genome Sequence of Brucella melitensis Biovar 3 Strain NI, Isolated from an Aborted Bovine Fetus.
<3>J. Bacteriol.
<4>194
<5>6321
<6>2012
<7>From an aborted bovine fetus in China, a bacterial strain named NI was isolated and identified
as Brucella melitensis by a PCR assay. Strain NI was further
characterized as B. melitensis biovar 3 using biochemical assays. Here we report
the complete genome sequence of strain NI.

<>

<1>Liu, W., Xiao, S., Li, M., Guo, S., Li, S., Luo, R., Feng, Z., Li, B., Zhou, Z., Shao, G., Chen, H., Fang, L.
<2>Comparative genomic analyses of Mycoplasma hyopneumoniae pathogenic 168 strain and its high-passaged attenuated strain.
<3>BMC Genomics
<4>14
<5>80
<6>2013
<7>BACKGROUND: Mycoplasma hyopneumoniae is the causative agent of porcine enzootic
pneumonia (EP), a mild, chronic pneumonia of swine. Despite presenting with low
direct mortality, EP is responsible for major economic losses in the pig
industry. To identify the virulence-associated determinants of M. hyopneumoniae,
we determined the whole genome sequence of M. hyopneumoniae strain 168 and its
attenuated high-passage strain 168-L and carried out comparative genomic
analyses. RESULTS: We performed the first comprehensive analysis of M.
hyopneumoniae strain 168 and its attenuated strain and made a preliminary survey
of coding sequences (CDSs) that may be related to virulence. The 168-L genome has
a highly similar gene content and order to that of 168, but is 4,483 bp smaller
because there are 60 insertions and 43 deletions in 168-L. Besides these indels,
227 single nucleotide variations (SNVs) were identified. We further investigated
the variants that affected CDSs, and compared them to reported virulence
determinants. Notably, almost all of the reported virulence determinants are
included in these variants affected CDSs. In addition to variations previously
described in mycoplasma adhesins (P97, P102, P146, P159, P216, and LppT), cell
envelope proteins (P95), cell surface antigens (P36), secreted proteins and
chaperone protein (DnaK), mutations in genes related to metabolism and growth may
also contribute to the attenuated virulence in 168-L. Furthermore, many mutations
were located in the previously described repeat motif, which may be of primary
importance for virulence. CONCLUSIONS: We studied the virulence attenuation
mechanism of M. hyopneumoniae by comparative genomic analysis of virulent strain
168 and its attenuated high-passage strain 168-L. Our findings provide a
preliminary survey of CDSs that may be related to virulence. While these include
reported virulence-related genes, other novel virulence determinants were also
detected. This new information will form the foundation of future investigations
into the pathogenesis of M. hyopneumoniae and facilitate the design of new
vaccines.

<>

<1>Liu, W., Yang, M., Xu, Z., Zheng, H., Liang, W., Zhou, R., Wu, B., Chen, H.
<2>Complete Genome Sequence of Pasteurella multocida HN06, a Toxigenic Strain of Serogroup D.
<3>J. Bacteriol.
<4>194
<5>3292-3293
<6>2012
<7>Pasteurella multocida is an important etiological agent that can cause many economically
important diseases in a wide range of mammals and birds. Here, we
report the complete genome sequence of P. multocida HN06, a toxigenic serogroup D
strain isolated from a diseased pig in China.

<>

<1>Liu, W.B., Yu, W.B., Gao, S.H., Ye, B.C.
<2>Genome Sequence of Saccharopolyspora erythraea D, a Hyperproducer of Erythromycin.
<3>Genome Announcements
<4>1
<5>e00718-13
<6>2013
<7>Saccharopolyspora erythraea is a Gram-positive bacterium that can produce antibiotics.
However, this microorganism must often be genetically improved for
higher production before it can be used in an industrial setting. Here, we report
the whole-genome sequence of the industrial hyperproducer strong mutator
Saccharopolyspora erythraea strain D.

<>

<1>Liu, W.Q., Feng, Y., Wang, Y., Zou, Q.H., Chen, F., Guo, J.T., Peng, Y.H., Jin, Y., Li, Y.G., Hu, S.N., Johnston, R.N., Liu, G.R., Liu, S.L.
<2>Salmonella paratyphi C: genetic divergence from Salmonella choleraesuis and pathogenic convergence with Salmonella typhi.
<3>PLoS ONE
<4>4
<5>E4510
<6>2009
<7>BACKGROUND: Although over 1400 Salmonella serovars cause usually
self-limited gastroenteritis in humans, a few, e.g., Salmonella typhi and
S. paratyphi C, cause typhoid, a potentially fatal systemic infection. It
is not known whether the typhoid agents have evolved from a common
ancestor (by divergent processes) or acquired similar pathogenic traits
independently (by convergent processes). Comparison of different typhoid
agents with non-typhoidal Salmonella lineages will provide excellent
models for studies on how similar pathogens might have evolved.
METHODOLOGIES/PRINCIPAL FINDINGS: We sequenced a strain of S. paratyphi C,
RKS4594, and compared it with previously sequenced Salmonella strains.
RKS4594 contains a chromosome of 4,833,080 bp and a plasmid of 55,414 bp.
We predicted 4,640 intact coding sequences (4,578 in the chromosome and 62
in the plasmid) and 152 pseudogenes (149 in the chromosome and 3 in the
plasmid). RKS4594 shares as many as 4346 of the 4,640 genes with a strain
of S. choleraesuis, which is primarily a swine pathogen, but only 4008
genes with another human-adapted typhoid agent, S. typhi. Comparison of
3691 genes shared by all six sequenced Salmonella strains placed S.
paratyphi C and S. choleraesuis together at one end, and S. typhi at the
opposite end, of the phylogenetic tree, demonstrating separate ancestries
of the human-adapted typhoid agents. S. paratyphi C seemed to have
suffered enormous selection pressures during its adaptation to man as
suggested by the differential nucleotide substitutions and different sets
of pseudogenes, between S. paratyphi C and S. choleraesuis. CONCLUSIONS:
S. paratyphi C does not share a common ancestor with other human-adapted
typhoid agents, supporting the convergent evolution model of the typhoid
agents. S. paratyphi C has diverged from a common ancestor with S.
choleraesuis by accumulating genomic novelty during adaptation to man.

<>

<1>Liu, W.Y., Chung, K.M., Wong, C.F., Jiang, J.W., Hui, R.K., Leung, F.C.
<2>Complete Genome Sequence of the Endophytic Enterobacter cloacae subsp. cloacae Strain ENHKU01.
<3>J. Bacteriol.
<4>194
<5>5965
<6>2012
<7>Enterobacter cloacae subsp. cloacae strain ENHKU01 is a Gram-negative endophyte isolated from
a diseased pepper (Capsicum annuum) plant in Hong Kong. This is the
first complete genome sequence report of a plant-endophytic strain of E. cloacae
subsp. cloacae.

<>

<1>Liu, X., Gai, Z., Tao, F., Yu, H., Tang, H., Xu, P.
<2>Genome Sequences of Pseudomonas luteola XLDN4-9 and Pseudomonas stutzeri XLDN-R,  Two Efficient Carbazole-Degrading Strains.
<3>J. Bacteriol.
<4>194
<5>5701-5702
<6>2012
<7>Pseudomonas luteola XLDN4-9 and Pseudomonas stutzeri XLDN-R are two efficient
carbazole-degrading pseudomonad strains. Here we present 4.63- and 4.70-Mb
assemblies of their genomes. Their annotated key genes for carbazole catabolism
are similar, which may provide further insights into the molecular mechanism of
carbazole degradation in Pseudomonas.

<>

<1>Liu, X., Zhou, R., Fu, G., Zhang, W., Min, Y., Tian, Y., Huang, D., Wang, K., Wan, Z., Yao, J., Yang, Z.
<2>Draft Genome Sequence of Bacillus thuringiensis NBIN-866 with High Nematocidal Activity.
<3>Genome Announcements
<4>2
<5>e00429-14
<6>2014
<7>Bacillus thuringiensis NBIN-866, a Gram-positive bacterium, was isolated from soil in China.
We announce here the draft genome sequence of strain B.
thuringiensis NBIN-866, which possesses highly nematocidal factors, such as
proteins and small molecular peptides.

<>

<1>Liu, X.-Q.
<2>Protein-splicing intein: Genetic mobility, origin, and evolution.
<3>Annu. Rev. Genet.
<4>34
<5>61-76
<6>2000
<7>Intein is the protein equivalent of intron and has been discovered in increasing numbers of
organisms and host proteins.  A self-splicing intein catalyzes its own removal from the host
protein through a posttranslational process of protein splicing.  A mobile intein displays a
site-specific endonuclease activity that confers genetic mobility to the intein through intein
homing.  Recent findings of intein structure and the mechanism of protein splicing illuminated
how inteins work and yielded clues regarding intein's origin, spread, and evolution.  Inteins
can evolve into new structures and new functions, such as split inteins that do
trans-splicing.  The structural basis of intein function needs to be identified for a full
understanding of the origin and evolution of this marvelous genetic element.

<>

<1>Liu, X.-Q., Hu, Z.
<2>A DnaB intein in Rhodothermus marinus: Indication of recent intein homing across remotely related organisms.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>7851-7856
<6>1997
<7>A dnaB gene encoding a homologue of the Escherichia coli DNA helicase DnaB was cloned and
sequenced in the thermophilic eubacterium Rhodothermus marinus, predicting a DnaB protein that
harbors an intein.  This DnaB intein is 428 amino acid residues long, has several putative
intein sequence motifs (including two putative endonuclease motifs), and is capable of protein
splicing when produced in E. coli cells.  The R. marinus DnaB intein is a close homologue of a
DnaB intein in the cyanobacterium Synechocystis sp. strain PCC6803.  The two inteins are
positioned identically in their respective DnaB proteins.  They also share a 54% sequence
identity (74% sequence similarity) that is markedly higher than the 37% sequence identity
shared by the extein sequences of the two DnaB proteins.  Horizontal intein transfer (homing)
is therefore invoked to relate these two DnaB inteins.  The codon usage of R. marinus DnaB
intein coding sequence differs markedly from the codon usage of its flanking extein coding
sequences and other genes in the same genome, suggesting more recent acquisition of the DnaB
intein in this organism.

<>

<1>Liu, X.Y., Luo, X.J., Li, C.X., Lai, Q.L., Xu, J.H.
<2>Draft Genome Sequence of Burkholderia sp. Strain MP-1, a Methyl Parathion (MP)-Degrading Bacterium from MP-Contaminated Soil.
<3>Genome Announcements
<4>2
<5>e00344-14
<6>2014
<7>Burkholderia sp. strain MP-1 was isolated from pesticide-contaminated soil. Herein, we report
the draft genome sequence of strain MP-1, which contains 168
contigs of 8,611,053 bp, with a G+C content of 62.55% and 7,631 protein-coding
genes.

<>

<1>Liu, X.Y., Min, Y., Wang, K.M., Wan, Z.Y., Zhang, Z.G., Cao, C.X., Zhou, R.H., Jiang, A.B., Liu, C.J., Zhang, G.Y., Cheng, X.L., Zhang, W., Yang, Z.W.
<2>Draft genome sequence of Bacillus amyloliquefaciens HB-26.
<3>Standards in Genomic Sciences
<4>9
<5>775-782
<6>2014
<7>Bacillus amyloliquefaciens HB-26, a Gram-positive bacterium was isolated from soil in China.
SDS-PAGE analysis showed this strain secreted six major protein
bands of 65, 60, 55, 34, 25 and 20 kDa. A bioassay of this strain reveals that it
shows specific activity against P. brassicae and nematode. Here we describe the
features of this organism, together with the draft genome sequence and
annotation. The 3,989,358 bp long genome (39 contigs) contains 4,001
protein-coding genes and 80 RNA genes.

<>

<1>Liu, Y., Hu, A., Shen, L., Yao, T., Jiao, N., Wang, N., Xu, B.
<2>Draft genome sequence of Dyadobacter tibetensis type strain (Y620-1) isolated from glacial ice.
<3>Standards in Genomic Sciences
<4>9
<5>883-892
<6>2014
<7>Dyadobacter tibetensis Y620-1 is the type strain of the species Dyadobacter tibetensis,
isolated from ice at a depth of 59 m from a high altitude glacier in
China (5670 m above sea level). It is psychrotolerant with growth temperature
ranges of 4 to 35 degrees C. Here we describe the features of this organism,
together with the draft genome sequence and annotation. The 5,313,963 bp long
genome contains 4,828 protein-coding genes and 39 RNA genes. To the best of our
knowledge, this is the first Dyadobacter strain that was isolated from glacial
ice. This study provides genetic information of this organism to identify the
genes linked to its specific mechanisms for adaption to extreme glacial
environment.

<>

<1>Liu, Y., Ichige, A., Kobayashi, I.
<2>Regulation of the EcoRI restriction-modification system: Identification of ecoRIM gene promoters and their upstream negative regulators in the ecoRIR gene.
<3>Gene
<4>400
<5>140-149
<6>2007
<7>Type II restriction-modification (R-M) systems are composed of linked restriction endonuclease
and modification methyltransferase genes and serve as barriers to horizontal gene transfer
even though they are mobile in themselves.  Their products kill host bacterial cells that have
lost the R-M genes, a process that helps to maintain the frequency of the R-M systems in the
viable cell population.  Their establishment and maintenance in a bacterial host are expected
to involve fine regulation of their gene expression.  In the present study, we analyzed
transcription of the modification gene and its regulation within the EcoRI R-M system.
Northern blotting revealed that the downstream ecoRIM gene is transcribed as a monocistronic
mRNA and as part of a larger bicistronic mRNA together with the upstream ecoRIR gene.  Primer
extension, RNase protection, and mutational analysis using lacZ gene fusions identified two
overlapping promoters for ecoRIM gene transcription within the ecoRIR gene.  Further
mutational analysis revealed that two upstream AT-rich elements within the ecoRIR gene,
"AATAAA" and "ATTATAAATATA", function as negative regulators of these promoters.  Simultaneous
substitution of these two elements resulted in a four-fold increase in beta-galactosidase
activity and a five-fold increase in transcript levels as measured by RNase protection assay.
RNA measurements of the ecoRIM transcript suggested that these elements decreased ecoRIM
expression by interfering with transcription initiation of the ecoRIM promoters.  Possible
roles for these ecoRIM promoters and their negative regulators in the EcoRI R-M system are
discussed.

<>

<1>Liu, Y., Lu, S.E., Baird, S.M., Qiao, J., Du, Y.
<2>Draft Genome Sequence of Pseudomonas chlororaphis YL-1, a Biocontrol Strain Suppressing Plant Microbial Pathogens.
<3>Genome Announcements
<4>2
<5>e01225-13
<6>2014
<7>Pseudomonas chlororaphis YL-1 was isolated from soybean root tips and showed a broad range of
antagonistic activities to microbial plant pathogens. Here, we
report the high-quality draft genome sequence of YL-1, which consists of a
chromosome with an estimated size of 6.8 Mb with a G+C value of 63.09%.

<>

<1>Liu, Y., Oakeley, E.J., Sun, L., Jost, J.-P.
<2>Multiple domains are involved in the targeting of the mouse DNA methyltransferase to the DNA replication foci.
<3>Nucleic Acids Res.
<4>26
<5>1038-1045
<6>1998
<7>It has been shown that, during the S-phase of the cell cycle, the mouse DNA methyltransferase
is targeted to sites of DNA replication by an amino acid sequence (aa 207-455) lying in the
N-terminal domain of the enzyme.  In this paper it is shown, by using enhanced green
fluorescent protein fusions, that peptide sequences of DNA MTase are also involved in this
targeting.  The work focuses on a sequence, downstream of the reported targeting sequence,
which is homologous to the Polybromo-1 protein.  This motif (designated as PBHD) is separated
from the reported targeting sequence by a zinc-binding motif.  Primed in situ extension using
centromeric-specific primers was used to show that both the host DNA MTase and EGFP fusion
proteins containing the targeting sequences were localized to centromeric, but not telomeric,
regions during late S-phase and mitosis.  Also found was that, in ~10% of the S-phase cells,
the EGFP fusions did not co-localize with the centromeric regions.  Mutants containing either,
or both, of these targeting sequences could act as dominant negative mutants against the host
DNA MTase.  EGFP fusion proteins, containing the reported TS (aa 207-455), were targeted to
centromeric regions throughout the mitotic stage which lead to the discovery of similar
behavior of the endogenous DNA MTase although the host MTase showed much less intense staining
than in S-phase cells.  The biological role of the centromeric localization of DNA MTase
during mitosis is currently unknown.

<>

<1>Liu, Y., Santi, D.V.
<2>m5C RNA and m5C DNA methyl transferases use different cysteine residues as catalysts.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>8263-8265
<6>2000
<7>A family of RNA m(5)C methyl transferases (MTases) containing over 55 members in eight
subfamilies has been identified recently by an iterative search of the genomic sequence
databases by using the known 16S rRNA m(5)C 967 MTase, Fmu, as an initial probe. The RNA m(5)C
MTase family contained sequence motifs that were highly homologous to motifs in the DNA m(5)C
MTases, including the ProCys sequence that contains the essential Cys catalyst of the
functionally similar DNA-modifying enzymes; it was reasonable to assign the Cys nucleophile to
be that in the conserved ProCys. The family also contained an additional conserved Cys residue
that aligns with the nucleophilic catalyst in m(5)U54 tRNA MTase. Surprisingly, the mutant of
the putative Cys catalyst in the ProCys sequence was active and formed a covalent complex with
5-fluorocytosine-containing RNA, whereas the mutant at the other conserved Cys was inactive
and unable to form the complex. Thus, notwithstanding the highly homologous sequences and
similar functions, the RNA m(5)C MTase uses a different Cys as a catalytic nucleophile than
the DNA m(5)C MTases. The catalytic Cys seems to be determined, not by the target base that is
modified, but by whether the substrate is DNA or RNA. The function of the conserved ProCys
sequence in the RNA m(5)C MTases remains unknown.

<>

<1>Liu, Y., Shen, W., Shi, G., Wang, Z.
<2>Cloning, expression and nucleotide sequence analysis of gene encoding for BamHI methyltransferase from Bacillus amyloliquefaciens CICIM B2125.
<3>Shengwu Jishu Tongbao
<4>11
<5>65-68
<6>2009
<7>A gene encoding for BamHI methyltransferase was cloned from Bacillus amyloliquefaciens CICIM
B2125.  The bamHIM was expressed under its own promoter in E. coli JM109.  The gene contained
an open reading frame of 1,271 bp, which encoded for 423 amino acid residues.  The molecular
weight of mature protein was 49 kD.  The BamHI site could be methylased by the enzyme M.BamHI.
The amino acid sequence analysis revealed that the enzyme contained a domain of Rossmann-fold
(NAD(P)(+)-binding proteins.

<>

<1>Liu, Y., Sun, L., Jost, J.-P.
<2>In differentiating mouse myoblasts DNA methyltransferase is posttranscriptionally and posttranslationally regulated.
<3>Nucleic Acids Res.
<4>24
<5>2718-2722
<6>1996
<7>Upon the onset of mouse myoblast differentiation there is a rapid drop in DNA
methyltransferase activity followed by a genome wide demethylation.  Here we show by using
specific antibodies directed against DNA methyltransferase that upon differentiation there was
a rapid drop in nuclear DNA methyltransferase whilst the internal control histone H1 remained
constant.  The loss of nuclear methyltransferase was not due to a translocation of the enzyme
from the nucleus to the cytoplasm where there was an increase in creatine phosphokinase
protein.  In vitro run on experiments carried out with growing and differentiating myoblast
nuclei showed no difference in the rate of DNA methyltransferase mRNA synthesis.  As measured
by Northern blot hybridization the relative half life of DNA methyltransferase mRNA in growing
and differentiating cells in the presence of Actinomycin D was 5h and 1h 30min, respectively,
whereas in the same cells the half life of histone H4 mRNA was in both cases 80 min.  As
measured by a combination of pulse chase experiments with labeled leucine and
immunoprecipitation, the relative half-life of DNA methyltransferase in growing and
differentiating cells was ~18h and 4h 30min, respectively.

<>

<1>Liu, Y., Sun, L., Jost, J.-P.
<2>In differentiating mouse myoblasts DNA methyltransferase is posttranscriptionally and posttranslationally regulated.
<3>Mol. Biol. Cell
<4>7
<5>8a
<6>1996
<7>Upon the onset of mouse myoblast differentiation there is a rapid drop in DNA
methyltransferase activity followed by a genome wide demethylation.  Here we show by using
specific antibodies directed against DNA methyltransferase that upon differentiation there was
a rapid drop in nuclear DNA methyltransferase whilst the internal control Histone H1 remained
constant.  The loss of nuclear methyltransferase was not due to a translocation of the enzyme
from the nucleus to the cytoplasm where there was an increase in creatine phosphokinase
protein.  In vitro run on experiments carried out with growing and differentiating myoblast
nuclei showed no difference in the rate of DNA methyltransferase mRNA synthesis.  As measured
by Northern blot hybridization the relative half life of DNA methyltransferase mRNA in growing
and differentiating cells in the presence of Actinomycin D was 5 h and 1 h 30 min
respectively, whereas in the same cells the half life of Histone H4 mRNA was in both cases 80
minutes.  As measured by a combination of pulse chase experiments with labeled leucine and
immunoprecipitation, the relative half life of DNA methyltransferase in growing and
differentiating cells was approximately 18 h and 4 h 30 min respectively.

<>

<1>Liu, Y., Wang, Y., Schwarz, S., Wang, S., Chen, L., Wu, C., Shen, J.
<2>Investigation of a multiresistance gene cfr that fails to mediate resistance to phenicols and oxazolidinones in Enterococcus faecalis.
<3>J. Antimicrob. Chemother.
<4>69
<5>892-898
<6>2014
<7>OBJECTIVES: To investigate the basis of susceptibility to phenicols and
oxazolidinones of the porcine Enterococcus faecalis CPPF5 despite the presence of
the multiresistance gene cfr. METHODS: Southern blotting, conjugation and
transformation analyses were conducted to confirm the plasmid location and
transferability of cfr in CPPF5. The genetic environment of cfr was determined by
sequence analysis. Transcription and translation of cfr were examined by RT-PCR
and western blotting, respectively, and modifications at A2503 within the 23S
rRNA sequence were identified by primer extension. RESULTS: Electrotransformation
and Southern blotting indicated that CPPF5 and its transformant 5B2-3 contained
two cfr-carrying plasmids approximately 50 and approximately 12 kb in size. The
complete 12,270 bp sequence of the smaller plasmid, pCPPF5, was determined and
shared 99.9% (12,269/12,270 bp) identity with the corresponding region of the
cfr-carrying plasmid pEF-01 in E. faecalis of cattle origin. Moreover, the
genetic environment of cfr in the approximately 50 kb plasmid was the same as
that in pCPPF5 according to sequencing results. Although cfr mRNA, Cfr protein
and a modification at the A2503 site were detected, the cfr-carrying transformant
5B2-3 did not have elevated MICs of chloramphenicol, florfenicol and linezolid,
indicating that cfr fails to mediate resistance to the respective antibiotics in
E. faecalis. CONCLUSIONS: This is the first report of the cfr gene failing to
elevate MICs of the corresponding antibiotics. Although the genetic basis for the
apparent 'no resistance' phenotype remains to be determined, this finding may
have implications for surveillance studies that target the cfr gene.

<>

<1>Liu, Y., Yao, S., Liu, Y., Xu, Y., Cheng, C.
<2>Genome Sequence of Luteimonas huabeiensis HB-2, a Novel Species of Luteimonas with High Oil Displacement Efficiency.
<3>Genome Announcements
<4>2
<5>e00152-14
<6>2014
<7>Luteimonas huabeiensis HB-2 is a novel and newly isolated strain, which shows a superior
property of oil displacement. Here, we present a 4.3-Mb assembly of its
genome. The key genes for phospholipid and fatty acid metabolism were annotated,
which are crucial for crude oil emulsification and recovery.

<>

<1>Liu, Y., Ye, W., Zheng, J., Fang, L., Peng, D., Ruan, L., Sun, M.
<2>High-quality draft genome sequence of nematocidal Bacillus thuringiensis Sbt003.
<3>Standards in Genomic Sciences
<4>9
<5>624-631
<6>2014
<7>Bacillus thuringiensis represents one of the six species of 'Bacillus cereus group' in the
genus Bacillus within the family Bacillaceae. Strain Sbt003 was
isolated from soil and identified as B. thuringiensis. It harbors at least seven
plasmids and produces three shapes of parasporal crystals including oval,
bipyramidal and rice. SDS-PAGE analysis of spore-crystal suspension of this
strain reveals six major protein bands, which implies the presence of multiple
parasporal crystal genes. Bioassay of this strain reveals that it shows specific
activity against nematodes and human cancer cells. In this study, we report the
whole genomic shotgun sequences of Sbt003. The high-quality draft of the genome
is 6,175,670 bp long (including chromosome and plasmids) with 6,372
protein-coding and 80 RNA genes.

<>

<1>Liu, Y., Yi, Z., Zeng, R.
<2>Draft Genome Sequence of a Symbiotic Bacterium, Rhizobium vignae CCBAU 05176T.
<3>Genome Announcements
<4>2
<5>e00657-14
<6>2014
<7>The Rhizobium vignae strain CCBAU 05176(T) was isolated from a root nodule of Astragalus
dahuricus grown in Hebei Province, China. It grows on yeast mannitol
agar (YMA) supplemented with 0 to 2% (wt/vol) NaCl. We report the annotated
genome sequence of this strain in a 6.34-Mb scaffold.

<>

<1>Liu, Y., Zheng, H., Zhan, G.Hua., Qin, W., Li, T., Li, W.Li.
<2>Establishment of an efficient transformation protocol and its application in marine-derived Bacillus strain.
<3>Sci. China C Life Sci.
<4>57
<5>627-635
<6>2014
<7>Marine-derived Bacillus strains have been proved to be a very promising source for natural
product leads. However, transformation of environmental strains is much more difficult than
that of domesticated strains. Here, we report the development of an efficient and robust
electroporation-based transformation system for marine-derived Bacillus marinus B-9987, which
is a macrolactin antibiotics producer and a very promising biological control agent against
fungal plant diseases. The transformation efficiency was greatly enhanced 10(3)-fold by using
unmethylated plasmid to bypass modification-restriction barrier, and using glycine betaine to
protect cells from electrical damages during electroporation. Addition of HEPES and 2 mmol L-1
MgCl2 further improved the efficiency by additional 2-fold, with a maximum value of 7.1x10(4)
cfu/mu g pHT3101. To demonstrate the feasibility and efficiency of the protocol, a green
fluorescent protein reporter system was constructed; furthermore, phosphopantetheinyl
transferase gene sfp, which is essential to the biosynthesis of polyketides and nonribosomal
peptides, was overexpressed in B-9987, leading to increased production of macrolactin A by
about 1.6-fold. In addition, this protocol is also applicable to marine-derived Bacillus
licheniforms EI-34-6, indicating it could be a reference for other undomesticated Bacillus
strains. To our knowledge, this is the first report regarding the transformation of
marine-derived Bacillus strain.

<>

<1>Liu, Y.P., Kobayashi, I.
<2>Negative regulation of the EcoRI restriction enzyme gene is associated with intragenic reverse promoters.
<3>J. Bacteriol.
<4>189
<5>6928-6935
<6>2007
<7>Type 11 restriction-modification systems are expected to possess mechanisms for tight
regulation of their expression to suppress the
potential of lethal attack on their host bacteria when they establish
and maintain themselves within them. Although the EcoRI restriction
enzyme has been well characterized, regulation of its expression is
still poorly understood. In this study, mutational analysis with lacZ
gene fusion and primer extension assay identified a promoter for the
transcription of the ecoRIR gene. Further analyses revealed that an
intragenic region containing two overlapping reverse promoter-like
elements acted as a negative regulator for ecoRIR gene expression. The
activity of these putative reverse promoters was verified by
transcriptional gene fusion, primer extension and in vitro
transcription. Mutations in these reverse promoters resulted in
increased gene expression in both translational and transcriptional
gene fusions. An RNase protection assay revealed that the transcript
level of the wild type relative to that of the reverse promoter mutant
at the downstream regions was much lower than the level at the upstream
regions. This suggests that these reverse promoter-like elements affect
their downstream transcript level. The possible mechanisms of this kind
of negative regulation, in addition to their possible biological roles,
are discussed.

<>

<1>Liutkeviciute, Z., Kriukiene, E., Grigaityte, I., Masevicius, V., Klimasauskas, S.
<2>Methyltransferase-Directed Derivatization of 5-Hydroxymethylcytosine in DNA.
<3>Angew. Chem. Int. Ed. Engl.
<4>50
<5>2090-2093
<6>2011
<7>The modification of cytosine by S-adenosylmethionine-dependent DNA methyltransferases is part
of an intricate epigenetic regulation mechanism in vertebrates.  DNA cytosine-5
methyltransferases (catalyze the transfer of a methyl group from S-adenosyl-L-methionine to
the cytosine residue in CpG dinucleotides.  Recent studies of genomic DNA from mouse embryonic
stem cells, neurons, and the brain found that a substantial fraction of 5-methylcytosine in CG
sequences is converted into 5-hydroxymethylcytosine by the action of 2-oxoglutarate- and
Fe2+-dependent oxygenases of the TET family.  As interactions of the 5-methyl- and
5-hydroxymethyl groups with cellular proteins in DNA are distinct, hmC residues may play an
independent role in yet unknown epigenetic pathways during embryonic development, brain
functioning, and cancer progression.  However, further studies of these intriguing phenomena
are hampered by the lack of efficient analytical techniques for mapping hmC residues in the
genome.  Herein we show that C5-MTases can direct the condensation of exogenous thiols and
selenols with hmC in DNA to yield the corresponding 5-chalcogenomethyl derivatives.  These
transformations open new possibilities for the sequence-specific derivatization and analysis
of this newly discovered epigenetic mark in mammalian DNA.

<>

<1>Liutkeviciute, Z., Kriukiene, E., Grigaityte, I., Vainorius, G., Masevicius, V., Klimasauskas, S.
<2>Catalytic versatility of DNA cytosine-5 methyltransferases: reactions involving non-cofactor-like substrates.
<3>FEBS J.
<4>279
<5>540
<6>2012
<7>DNA cytosine-5 methyltransferases catalyze site-specific transfers of a methyl group from the
cofactor S-adenosyl-L-methionine (SAM) onto the 5-position of their target cytosine residues
in DNA.  Recently we have shown that, in the absence of SAM, methyltranferases are able to add
formaldehyde to their target cytosines yielding 5-hydroxymethylcytosine.  This reaction can be
reversed yielding unmodified cytosine, or can be further extended by condensation with thiols
or selenols.  Lately hmC was discovered in mammals DNA but the biological role of this new
base is unclear because further studies are restricted by the lack of efficient analytical
techniques for mapping hmC residues in the genome.  These atypical reactions of DNA cytosine-5
methyltransferase open new ways for analysis of hnmC in genomic DNA and provide inroads into
active demethylation of 5-methylcytosine residues in the genome.

<>

<1>Liutkeviciute, Z., Lukinavicius, G., Masevicius, V., Daujotyte, D., Klimasauskas, S.
<2>Cytosine-5-methyltransferases add aldehydes to DNA.
<3>Nat. Chem. Biol.
<4>5
<5>400-402
<6>2009
<7>Targeted methylation of cytosine residues by S-adenosylmethionine-dependent DNA
methyltransferases modulates gene
expression in vertebrates. Here we show that
cytosine-5-methyltransferases catalyze reversible covalent addition of
exogenous aliphatic aldehydes to their target residues in DNA, thus
yielding corresponding 5-alpha-hydroxyalkylcytosines. Such atypical
enzymatic reactions with non-cofactor-like substrates open new ways for
sequence-specific derivatization of DNA and demonstrate enzymatic
exchange of 5-hydroxymethyl groups on cytosine in support of an
oxidative mechanism of DNA demethylation.

<>

<1>Llado, S., Xu, Z., Sorensen, S.J., Baldrian, P.
<2>Draft Genome Sequence of Burkholderia sordidicola S170, a Potential Plant Growth  Promoter Isolated from Coniferous Forest Soil in the Czech Republic.
<3>Genome Announcements
<4>2
<5>e00810-14
<6>2014
<7>Burkholderia species are key players in the accumulation of carbon from cellulose
decomposition in coniferous forest ecosystems. We report here the draft genome of
Burkholderia sordidicola strain S170, containing features associated with known
genes involved in plant growth promotion, the biological control of plant
diseases, and green remediation technologies.

<>

<1>Lloyd, K.G., Schreiber, L., Petersen, D.G., Kjeldsen, K.U., Lever, M.A., Steen, A.D., Stepanauskas, R., Richter, M., Kleindienst, S., Lenk, S., Schramm, A., Jorgensen, B.B.
<2>Predominant archaea in marine sediments degrade detrital proteins.
<3>Nature
<4>496
<5>215-218
<6>2013
<7>Half of the microbial cells in the Earth's oceans are found in sediments. Many of these cells
are members of the Archaea, single-celled prokaryotes in a domain of
life separate from Bacteria and Eukaryota. However, most of these archaea lack
cultured representatives, leaving their physiologies and placement on the tree of
life uncertain. Here we show that the uncultured miscellaneous crenarchaeotal
group (MCG) and marine benthic group-D (MBG-D) are among the most numerous
archaea in the marine sub-sea floor. Single-cell genomic sequencing of one cell
of MCG and three cells of MBG-D indicated that they form new branches basal to
the archaeal phyla Thaumarchaeota and Aigarchaeota, for MCG, and the order
Thermoplasmatales, for MBG-D. All four cells encoded extracellular
protein-degrading enzymes such as gingipain and clostripain that are known to be
effective in environments chemically similar to marine sediments. Furthermore, we
found these two types of peptidase to be abundant and active in marine sediments,
indicating that uncultured archaea may have a previously undiscovered role in
protein remineralization in anoxic marine sediments.

<>

<1>Lloyd, R.S., Cheng, X.
<2>Mechanistic link between DNA methyltransferases and DNA repair enzymes by base flipping.
<3>Biopolymers
<4>44
<5>139-151
<6>1997
<7>Rotation of a DNA nucleotide out of the double helix and into a protein binding pocket ("base
flipping") was first observed in the structure of a DNA methyltransferase.  There is now
evidence that a variety of proteins, particularly DNA repair enzymes, use base flipping in
their interactions with DNA.  Though the mechanisms for base movement into extrahelical
positions are still unclear, the focus of this review is how base recognition is modulated by
the stringency of binding to the extrahelical base(s) or sugar moiety.

<>

<1>Lluch-Senar, M., Luong, K., Llorens-Rico, V., Delgado, J., Fang, G., Spittle, K., Clark, T.A., Schadt, E., Turner, S.W., Korlach, J., Serrano, L.
<2>Comprehensive Methylome Characterization of Mycoplasma genitalium and Mycoplasma  pneumoniae at Single-Base Resolution.
<3>PLoS Genet.
<4>9
<5>e1003191
<6>2013
<7>In the bacterial world, methylation is most commonly associated with restriction-modification
systems that provide a defense mechanism against
invading foreign genomes. In addition, it is known that methylation plays
functionally important roles, including timing of DNA replication, chromosome
partitioning, DNA repair, and regulation of gene expression. However, full DNA
methylome analyses are scarce due to a lack of a simple methodology for rapid and
sensitive detection of common epigenetic marks (ie N(6)-methyladenine (6 mA) and
N(4)-methylcytosine (4 mC)), in these organisms. Here, we use Single-Molecule
Real-Time (SMRT) sequencing to determine the methylomes of two related human
pathogen species, Mycoplasma genitalium G-37 and Mycoplasma pneumoniae M129, with
single-base resolution. Our analysis identified two new methylation motifs not
previously described in bacteria: a widespread 6 mA methylation motif common to
both bacteria (5'-CTAT-3'), as well as a more complex Type I m6A sequence motif
in M. pneumoniae (5'-GAN(7)TAY-3'/3'-CTN(7)ATR-5'). We identify the
methyltransferase responsible for the common motif and suggest the one involved
in M. pneumoniae only. Analysis of the distribution of methylation sites across
the genome of M. pneumoniae suggests a potential role for methylation in
regulating the cell cycle, as well as in regulation of gene expression. To our
knowledge, this is one of the first direct methylome profiling studies with
single-base resolution from a bacterial organism.

<>

<1>Lo, C.I., Mishra, A.K., Padhmanabhan, R., Samb, B., Sow, A.G., Robert, C., Couderc, C., Faye, N., Raoult, D., Fournier, P.E., Fenollar, F.
<2>Non-contiguous finished genome sequence and description of Clostridium dakarense  sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>14-27
<6>2013
<7>Clostridium dakarense strain FF1(T), is the type strain of Clostridium dakarense  sp. nov., a
new species within the genus Clostridium. This strain, whose genome
is described here, was isolated from the fecal flora of a 4-month-old Senegalese
child suffering from gastroenteritis. C. dakarense sp. nov. strain FF1(T) is an
obligate anaerobic Gram-positive bacillus. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
3,735,762 bp long genome (1 chromosome but no plasmid) exhibits a G+C content of
27.98% and contains 3,843 protein-coding and 73 RNA genes, including 8 rRNA
genes.

<>

<1>Lo, I., Denef, V.J., Verberkmoes, N.C., Shah, M.B., Goltsman, D., Dibartolo, G., Tyson, G.W., Allen, E.E., Ram, R.J., Detter, J.C., Richardson, P., Thelen, M.P., Hettich, R.L., Banfield, J.F.
<2>Strain-resolved community proteomics reveals recombining genomes of acidophilic bacteria.
<3>Nature
<4>446
<5>537-541
<6>2007
<7>Microbes comprise the majority of extant organisms, yet much remains to be
learned about the nature and driving forces of microbial diversification.
Our understanding of how microorganisms adapt and evolve can be advanced
by genome-wide documentation of the patterns of genetic exchange,
particularly if analyses target coexisting members of natural communities.
Here we use community genomic data sets to identify, with strain
specificity, expressed proteins from the dominant member of a genomically
uncharacterized, natural, acidophilic biofilm. Proteomics results reveal a
genome shaped by recombination involving chromosomal regions of tens to
hundreds of kilobases long that are derived from two closely related
bacterial populations. Inter-population genetic exchange was confirmed by
multilocus sequence typing of isolates and of uncultivated natural
consortia. The findings suggest that exchange of large blocks of gene
variants is crucial for the adaptation to specific ecological niches
within the very acidic, metal-rich environment. Mass-spectrometry-based
discrimination of expressed protein products that differ by as little as a
single amino acid enables us to distinguish the behaviour of closely
related coexisting organisms. This is important, given that microorganisms
grouped together as a single species may have quite distinct roles in
natural systems and their interactions might be key to ecosystem
optimization. Because proteomic data simultaneously convey information
about genome type and activity, strain-resolved community proteomics is an
important complement to cultivation-independent genomic (metagenomic)
analysis of microorganisms in the natural environment.

<>

<1>Lo, J.R., Lang, J.M., Darling, A.E., Eisen, J.A., Coil, D.A.
<2>Draft Genome Sequence of an Actinobacterium, Brachybacterium muris Strain UCD-AY4.
<3>Genome Announcements
<4>1
<5>e0008613
<6>2013
<7>Here we present the draft genome of an actinobacterium, Brachybacterium muris UCD-AY4. The
assembly contains 3,257,338 bp and has a GC content of 70%. This
strain was isolated from a residential bath towel and has a 16S rRNA gene 99.7%
identical to that of the original B. muris strain, C3H-21.

<>

<1>Lo, W.-S., Ku, C., Chen, L.-L., Chang, T.-H., Kuo, C.-H.
<2>Comparison of metabolic capacities and inference of gene content evolution in mosquito-associated Spiroplasma diminutum and S. taiwanense.
<3>Genome Biol. Evol.
<4>5
<5>1512-1523
<6>2013
<7>Mosquitoes are hosts of several Spiroplasma species that belong to different serogroups. To
investigate the genetic mechanisms that may be involved in the utilization of similar hosts in
these phylogenetically distinct bacteria, we determined the complete genome sequences of S.
diminutum and S. taiwanense for comparative analysis. The genome alignment indicates that
their chromosomal organization is highly conserved,
which is in sharp contrast to the elevated genome instabilities observed in other Spiroplasma
lineages. Examination of the substrate utilization strategies revealed that S. diminutum can
use a wide range of carbohydrates, suggesting that it is well suited to living in the gut (and
possibly the circulatory system) of its mosquito hosts. In comparison, S. taiwanense has lost
several carbohydrate utilization genes and acquired additional sets of oligopeptide
transporter genes through tandem duplications, suggesting
that proteins from digested blood meal or lysed host cells may be an important nutrient
source. Moreover, one glycerol-3-phosphate oxidase gene (glpO) was found in S. taiwanense but
not S. diminutum. This gene is linked to the production of reactive oxygen species and has
been shown to be a major virulence factor in Mycoplasma mycoides. This finding may explain the
pathogenicity of S. taiwanense observed in previous artificial infection experiments, while no
apparent effect was found for S. diminutum. To infer the gene content evolution at deeper
divergence levels, we incorporated other Mollicutes genomes for comparative analyses. The
results suggest that the losses of biosynthetic pathways are a recurrent theme in these
host-associated bacteria.

<>

<1>Lo, W.S., Chen, H., Chen, C.Y., Kuo, C.H.
<2>Complete Genome Sequence of Vibrio vulnificus 93U204, a Bacterium Isolated from Diseased Tilapia in Taiwan.
<3>Genome Announcements
<4>2
<5>e01005-14
<6>2014
<7>Vibrio vulnificus 93U204 is a bacterium isolated from a moribund tilapia collected in
Kaohsiung, Taiwan. Here, we report the complete genome sequence of
this bacterium to facilitate the investigation of its pathogenicity and for
comparative analyses with human-pathogenic strains within the same species.

<>

<1>Lobner-Olesen, A., Boye, E., Marinus, M.G.
<2>Expression of the Escherichia coli dam gene.
<3>Mol. Microbiol.
<4>6
<5>1841-1851
<6>1992
<7>The Escherichia coli dam gene and upstream sequences were cloned from the Kohara phage 4D4.
Five promoters were found to contribute to dam gene transcription.  P1 and P2 (the major
promoter) were situated approximately 3.5 kb upstream of the structural gene, P3 was within
the aroB gene, P4 was within the urf74.3 gene, and P5 was in the urf74.3-dam intergenic
region.  The nucleotide sequence of 2280 bp of DNA containing P1 and P2 was determined and
shown to have the potential to encode a protein of approximately 16 kDa between P1, P2 and the
aroB gene.  This 16 kDa open reading frame has been identified as aroK, the gene for shikimic
acid kinase I.  Thus the dam gene is part of an operon containing aroK, aroB, urf74.3, and
dam.  The transcriptional start points of the promoters were determined.  A comparison of
their nucleotide sequences suggested that P1-P4 were all recognized by the sigma 70 subunit of
the RNA polymerase.

<>

<1>Lobner-Olesen, A., Skovgaard, O., Marinus, M.G.
<2>Dam methylation: coordinating cellular processes.
<3>Curr. Opin. Microbiol.
<4>8
<5>154-160
<6>2005
<7>GATC sequences in Escherichia coli DNA are methylated at the adenine residue by DNA adenine
methyltransferase (DamMT). These methylated
residues and/or the level of DamMT can influence cellular functions
such as gene transcription, DNA mismatch repair, initiation of
chromosome replication and nucleoid structure. In certain bacteria,
unlike E coli, DamMT is essential for viability perhaps owing to its
role in chromosome replication. DamMT has also been implicated as a
virulence factor in bacterial pathogenesis. The origin and phylogeny of
DamMT, based on sequenced genomes, has been deduced.

<>

<1>Lobner-Olesen, A., von Freiesleben, U.
<2>Chromosomal replication incompatibility in Dam methyltransferase deficient Escherichia coli cells.
<3>EMBO J.
<4>15
<5>5999-6008
<6>1996
<7>Dam methyltransferase deficient Escherichia coli cells containing minichromosomes were
constructed.  Free plasmid DNA could not be detected in these cells and the minichromosomes
were found to be integrated in multiple copies in the origin of replication (oriC) region of
the host chromosome.  The absence of the initiation cascade in Dam- cells is proposed to
account for this observation of apparent incompatibility between plasmid and chromosomal
copies of oriC.  Studies using oriC-pBR322 chimeric plasmids and their deletion derivatives
indicated that the incompatibility determinant is an intact and functional oriC sequence.  The
seqA2 mutation was found to overcome the incompatibility phenotype by increasing the cellular
oriC copy number 3-fold thereby allowing minichromosomes to coexist with the chromosome.  The
replication pattern of a wild-type strain with multiple integrated minichromosomes in the oriC
region of the chromosome, led to the conclusion that initiation of DNA replication commences
at a fixed cell mass, irrespective of the number of origins contained on the chromosome.

<>

<1>Lobocka, M.B., Rose, D.J., Rusin, M., Plunkett, G. III, Samojedny, A., Lehnherr, H., Yarmolinsky, M.B., Blattner, F.R.
<2>Genome of bacteriophage P1.
<3>J. Bacteriol.
<4>186
<5>7032-7068
<6>2004
<7>P1 is a bacteriophage of Escherichia coli and other enteric bacteria.
It lysogenizes its hosts as a circular, low-copy-number plasmid. We have
determined the complete nucleotide sequences of two strains of a P1
thermoinducible mutant, P1 c1-100. The P1 genome (93,601 bp) contains at
least 117 genes, of which almost two-thirds had not been sequenced
previously and 49 have no homologs in other organisms. Protein-coding
genes occupy 92% of the genome and are organized in 45 operons, of which
four are decisive for the choice between lysis and lysogeny. Four others
ensure plasmid maintenance. The majority of the remaining 37 operons are
involved in lytic development. Seventeen operons are transcribed from
sigma(70) promoters directly controlled by the master phage repressor C1.
Late operons are transcribed from promoters recognized by the E. coli RNA
polymerase holoenzyme in the presence of the Lpa protein, the product of a
C1-controlled P1 gene. Three species of P1-encoded tRNAs provide
differential controls of translation, and a P1-encoded DNA
methyltransferase with putative bifunctionality influences transcription,
replication, and DNA packaging. The genome is particularly rich in Chi
recombinogenic sites. The base content and distribution in P1 DNA indicate
that replication of P1 from its plasmid origin had more impact on the base
compositional asymmetries of the P1 genome than replication from the lytic
origin of replication.

<>

<1>Lobos, C., Vasquez, C.
<2>Purification and characterisation of BstLVI restriction endonuclease, a thermostable isoschizomer of ClaI from Bacillus stearothermophilus LV.
<3>Biochim. Biophys. Acta
<4>1171
<5>295-298
<6>1993
<7>This work describes the purification and biochemical characterization of BstLVI restriction
endonuclease, a thermostable isoschizomer of ClaI, from Bacillus stearothermophilus LV. The
enzyme was purified by successive DEAE-cellulose, Affi-Gel Blue and Heparin-Sepharose CL-6B
column chromatography. A molecular weight of 37000 was determined for BstLVI by gel
filtration. As expected for thermophilic proteins, the enzyme showed a high stability towards
heat and also to other known protein-denaturing agents.

<>

<1>Lodes, M.J., Mohamath, R., Henderson, R.A., Benson, D.R., Secrist, H.
<2>Compositions and methods for the therapy and diagnosis of lung cancer.
<3>US Patent Office
<4>US 6759508 A
<5>
<6>2004
<7>Compositions and methods for the therapy and diagnosis of cancer, particularly lung cancer,
are disclosed.  Illustrative compositions comprise one or more lung tumor polypeptides,
immunogenic portions thereof, polynucleotides that encode such polypeptides, antigen
presenting cell that expresses such polypeptides, and T cells that are specific for cells
expressing such polypeptides.  The disclosed compositions are useful, for example, in the
diagnosis, prevention and/or treatment of diseases, particularly lung cancer.

<>

<1>Lodwick, D., Ross, H.N.M., Harris, J.E., Almond, J.W., Grant, W.D.
<2>dam Methylation in the archaebacteria.
<3>J. Gen. Microbiol.
<4>132
<5>3055-3059
<6>1986
<7>The DNA of certain species of halophilic and methanogenic archaebacteria is dam methylated, as
shown by restriction endonuclease sensitivities. The Dam+ phenotype appears to be confined to
particular taxonomic groupings defined by DNA:rRNA hybridization or 16S RNA oligonucleotide
cataloging.

<>

<1>Loeffler, J.M., Fischetti, V.A.
<2>Lysogeny of Streptococcus pneumoniae with MM1 Phage: Improved Adherence and Other Phenotypic Changes.
<3>Infect. Immun.
<4>74
<5>4486-4495
<6>2006
<7>Pneumococcal prophages are extremely frequent, but no role in pathogenesis has so far been
attributed to them. We isolated a variant of phage MM1, named MM1-1998, from a serotype 24
strain of Streptococcus pneumoniae. We created three isogenic strain pairs (serotypes 3, 4,
and 24) that differed only by the lysogenic presence of the MM1-1998 phage and did a
phenotypic comparison. Lysogeny led to improved adherence to inert surfaces and pharyngeal
cells compared to that with the cured variants of the strains. We found that lysogeny with
MM1-1998 coincided with a more transparent phenotype and phage curing with more opaque
colonies in all strain pairs, and we discovered that transparency was associated with more
successful and stable lysogeny. Since transparency alone was possibly responsible for the
adherence difference, we further compared the TIGR4 lysogen with an equally transparent
variant of TIGR4 in order to reassess the role of phage or transparency separately. The
results revealed that improved adherence was independently associated with lysogeny with the
MM1-1998 phage. Other phenotypic differences such as faster growth, increased autolysis, and
decreased intracellular hemolytic activity were more likely due to transparency. By improving
the adherence of pneumococci, this prophage may contribute to their fitness and possibly to
their persistence in humans.

<>

<1>Loenen, W., Murray, N.
<2>Modification Enhancement by Restriction Alleviation Protein (Ral) of Bacteriophage lambda.
<3>J. Mol. Biol.
<4>190
<5>11-22
<6>1986
<7>The product of the lambda ral gene alleviates restriction and enhances
modification by the Escherichia coli K-12 restriction and modification system.
An open reading frame (orf) located between genes N and Ealpha10 has been
assigned to the ral gene.  We have cloned this orf in a plasmid where its
transcription is controlled by a thermolabile lambda repressor.  Inactivation
of the lambda repressor caused a 1000-fold reduction in K-specific restriction
of unmodified lambda phage and a 100-fold increase in modification.  In
minicells transformed with ral+ plasmids, derepression resulted in the
appearance of a polypeptide with a lower mobility than that predicted for a
protein encoded by the orf attributed to ral; in a transcription and
translation system in vitro DNA from a ral+ plasmid encoded a polypeptide with
the same mobility.  This polypeptide was absent when the plasmid DNA carried a
mutant ral gene.  The nucleotide sequence of this mutant gene defined two base
changes, one of which inactivates the initiation codon of the orf.  The K
restriction endonuclease, which is also a K-specific methylase, is encoded by
three genes designated hsdR, hsdM and hsdS, although the hsdR polypeptide is
not essential for the methylase activity.  We show that Ral enchances
modification in a host strain lacking the entire hsdR gene, and lambda phages
carrying the hsdM and S genes modify their their own DNA inefficiently in the
absence of Ral, despite the fact that derivatives of these phages provide
efficient amplification of the K-specific methylase.  Our data support a model
in which, as a consequence of the interaction of Ral with either the hsdM or
the hsdS polypeptide, the conformation of the enzyme is changed and the
efficiency of methylation of unmodified target sites is enhanced.  It has been
postulated that Ral counteracts Rho, but in our experiments Ral did not relieve
transcriptional polarity.

<>

<1>Loenen, W.A., Dryden, D.T., Raleigh, E.A., Wilson, G.G.
<2>Type I restriction enzymes and their relatives.
<3>Nucleic Acids Res.
<4>42
<5>20-44
<6>2014
<7>Type I restriction enzymes (REases) are large pentameric proteins with separate restriction
(R), methylation (M) and DNA sequence-recognition (S) subunits. They
were the first REases to be discovered and purified, but unlike the enormously
useful Type II REases, they have yet to find a place in the enzymatic toolbox of
molecular biologists. Type I enzymes have been difficult to characterize, but
this is changing as genome analysis reveals their genes, and methylome analysis
reveals their recognition sequences. Several Type I REases have been studied in
detail and what has been learned about them invites greater attention. In this
article, we discuss aspects of the biochemistry, biology and regulation of Type I
REases, and of the mechanisms that bacteriophages and plasmids have evolved to
evade them. Type I REases have a remarkable ability to change sequence
specificity by domain shuffling and rearrangements. We summarize the classic
experiments and observations that led to this discovery, and we discuss how this
ability depends on the modular organizations of the enzymes and of their S
subunits. Finally, we describe examples of Type II restriction-modification
systems that have features in common with Type I enzymes, with emphasis on the
varied Type IIG enzymes.

<>

<1>Loenen, W.A., Dryden, D.T., Raleigh, E.A., Wilson, G.G., Murray, N.E.
<2>Highlights of the DNA cutters: a short history of the restriction enzymes.
<3>Nucleic Acids Res.
<4>42
<5>3-19
<6>2014
<7>In the early 1950's, 'host-controlled variation in bacterial viruses' was reported as a
non-hereditary phenomenon: one cycle of viral growth on certain bacterial hosts affected the
ability of progeny virus to grow on other hosts by either restricting or enlarging their host
range. Unlike mutation, this change was reversible, and one cycle of growth in the previous
host returned the virus to its original form. These simple observations heralded the discovery
of the endonuclease and methyltransferase activities of what are now termed Type I, II, III
and IV DNA restriction-modification systems. The Type II restriction enzymes (e.g. EcoRI) gave
rise to recombinant DNA technology that has transformed molecular biology and medicine. This
review traces the discovery of restriction enzymes and their continuing impact on molecular
biology and medicine.

<>

<1>Loenen, W.A., Raleigh, E.A.
<2>The other face of restriction: modification-dependent enzymes.
<3>Nucleic Acids Res.
<4>42
<5>56-69
<6>2014
<7>The 1952 observation of host-induced non-hereditary variation in bacteriophages by Salvador
Luria and Mary Human led to the discovery in the 1960s of modifying enzymes that glucosylate
hydroxymethylcytosine in T-even phages and of genes encoding corresponding host activities
that restrict non-glucosylated phage DNA: rglA and rglB (restricts glucoseless phage). In the
1980's, appreciation of the biological scope of these activities was dramatically expanded
with the demonstration that plant and animal DNA was also sensitive to restriction in cloning
experiments. The rgl genes were renamed mcrA and mcrBC (modified cytosine restriction). The
new class of modification-dependent restriction enzymes was named Type IV, as distinct from
the familiar modification-blocked Types I-III. A third Escherichia coli enzyme, mrr (modified
DNA rejection and restriction) recognizes both methylcytosine and methyladenine. In recent
years, the universe of modification-dependent enzymes has expanded greatly. Technical advances
allow use of Type IV enzymes to study epigenetic mechanisms in mammals and plants. Type IV
enzymes recognize modified DNA with low sequence selectivity and have emerged many times
independently during evolution. Here, we review biochemical and structural data on these
proteins, the resurgent interest in Type IV enzymes as tools for epigenetic research and the
evolutionary pressures on these systems.

<>

<1>Loenen, W.A.M.
<2>Tracking EcoKI and DNA fifty years on: a golden story full of surprises.
<3>Nucleic Acids Res.
<4>31
<5>7059-7069
<6>2003
<7>1953 was a historical year for biology, as it marked the birth of the DNA helix, but also a
report by Bertani and Weigle on 'a barrier to infection'
of bacteriophage lambda in its natural host, Escherichia coli K-12, that
could be lifted by 'host-controlled variation' of the virus. This paper
lay dormant till Nobel laureate Arber and PhD student Dussoix showed that
the lambda DNA was rejected and degraded upon infection of different
bacterial hosts, unless it carried host-specific modification of that DNA,
thus laying the foundations for the phenomenon of restriction and
modification (R-M). The restriction enzyme of E.coli K-12, EcoKI, was
purified in 1968 and required S-adenosylmethionine (AdoMet) and ATP as
cofactors. By the end of the decade there was substantial evidence for a
chromosomal locus hsdK with three genes encoding restriction (R),
modification (M) and specificity (S) subunits that assembled into a large
complex of >400 kDa. The 1970s brought the message that EcoKI cut away
from its DNA recognition target, to which site the enzyme remained bound
while translocating the DNA past itself, with concomitant ATP hydrolysis
and subsequent double-strand nicks. This translocation event created
clearly visible DNA loops in the electron microscope. EcoKI became the
archetypal Type I R-M enzyme with curious DNA translocating properties
reminiscent of helicases, recognizing the bipartite asymmetric site
AAC(N6)GTGC. Cloning of the hsdK locus in 1976 facilitated molecular
understanding of this sophisticated R-M complex and in an elegant 'pas de
deux' Murray and Dryden constructed the present model based on a large
body of experimental data plus bioinformatics. This review celebrates the
golden anniversary of EcoKI and ends with the exciting progress on the
vital issue of restriction alleviation after DNA damage, also first
reported in 1953, which involves intricate control of R subunit activity
by the bacterial proteasome ClpXP, important results that will keep
scientists on the EcoKI track for another 50 years to come.

<>

<1>Loenen, W.A.M., Daniel, A.S., Braymer, H.D., Murray, N.E.
<2>Organization and sequence of the hsd genes of Escherichia coli K12.
<3>J. Mol. Biol.
<4>198
<5>159-170
<6>1987
<7>The nucleotide sequence of the hsdR and M genes, together with that for hsdS
comprises an 8400 base segment spanning the entire hsd region of Escherichia
coli K-12.  The three hsd genes are transcribed in the same direction, but from
two promoters.  hsdR and hsdM are separated by 492 base-pairs, whereas the
termination codon of hsdM overlaps the initiation codon of hsdS.  pres precedes
hsdR, and our data indicate a transcription termination signal in the interval
between hsdR and -mod, as expected if transcription of hsdM and S is dependent
on pmod.  Transcription from pres is not influenced by the products of the hsdM
and S genes, and the mechanism whereby restriction is prevented when the hsd
region is transferred to a modification-deficient cell remains to be
elucidated.  A segment of the predicted amino acid sequence of the M
polypeptide shares homology with a variety of adenine methylases and may
identify part of the active site for methylation of specific adenine residues.
The R polypeptide shows homology with a variety of ATPases, and pronounced
regions of alpha-helical structure are predicted, one of which is amphipathic.

<>

<1>Loessner, M.J., Wendlinger, G., Scherer, S.
<2>Heterogeneous endolysins in Listeria monocytogenes bacteriophages: a new class of enzymes and evidence for conserved holin genes within the siphoviral lysis cassettes.
<3>Mol. Microbiol.
<4>16
<5>1231-1241
<6>1995
<7>Listeria monocytogenes bacteriophages A118, A500 and A511 are members of three distinct phage
groups with characteristic host ranges. Their endolysin (ply) genes were cloned and expressed
in Escherichia coli as demonstrated by the conferred lytic phenotype when colonies of
recombinant cells were overlaid with a lawn of Listeria cells. The nucleotide sequences of the
cloned DNA fragments were determined and the individual enzymes (PLY118, 30.8 kDa; PLY500,
33.4 kDa; PLY511, 36.5 kDa) were shown to have varying degrees of homology within their
N-terminal or C-terminal domains. Transcriptional analysis revealed them to be 'late' genes
with transcription beginning 15-20 min post-infection. The enzymes were overexpressed and
partially purified and their individual specificities examined. When applied exogenously, the
lysins induced rapid lysis of Listeria strains from all species but generally did not affect
other bacteria. Using hydrolysis of purified listerial cell walls, PLY511 was characterized as
an N-acetylmuramoyl-L-alanine amidase (EC 3.5.1.28) and shows homology in its N-terminal
domain to other enzymes of this type. In contrast, PLY118 and PLY500 were shown to represent a
new class of cell wall lytic enzymes which cleave between the L-alanine and D-glutamate
residues of listerial peptidoglycan; these were designated as L-alanoyl-D-glutamate
peptidases. These two enzymes share homology in the N-terminal domain which we propose
determines hydrolytic specificity. Highly conserved holin (hol) gene sequences are present
upstream of ply118 and ply500. They encode proteins of structural similarity to the product of
phage lambda gene S, and are predicted to be membrane proteins which form pores to allow
access of the lysins to their peptidoglycan substrates. This arrangement of conserved holin
genes with downstream lysin genes among the siphoviral lysis cassettes explains why the
cytoplasmic endolysins alone are not lethal, since they require a specific transport function
across the cell membrane.

<>

<1>Loewen, P.C., Alsaadi, Y., Fernando, D., Kumar, A.
<2>Genome Sequence of a Tigecycline-Resistant Clinical Isolate of Acinetobacter baumannii Strain AB031 Obtained from a Bloodstream Infection.
<3>Genome Announcements
<4>2
<5>e01036-14
<6>2014
<7>We report here the 3.8-Mbp genome sequence of a blood isolate of Acinetobacter baumannii
strain AB031.

<>

<1>Loewen, P.C., Alsaadi, Y., Fernando, D., Kumar, A.
<2>Genome Sequence of an Extremely Drug-Resistant Clinical Isolate of Acinetobacter  baumannii Strain AB030.
<3>Genome Announcements
<4>2
<5>e01035-14
<6>2014
<7>We report the 4.3-Mbp genome sequence of a blood isolate of Acinetobacter baumannii strain
AB030.

<>

<1>Loewen, P.C., Switala, J., Fernando, W.G., de Kievit, T.
<2>Genome Sequence of Pseudomonas brassicacearum DF41.
<3>Genome Announcements
<4>2
<5>e00390-14
<6>2014
<7>Pseudomonas brassicacearum DF41, a Gram-negative soil bacterium, is able to suppress the
fungal pathogen Sclerotinia sclerotiorum through a process known as
biological control. Here, we present a 6.8-Mb assembly of its genome, which is
the second fully assembled genome of a P. brassicacearum strain.

<>

<1>Loewen, P.C., Villenueva, J., Fernando, W.G., de Kievit, T.
<2>Genome Sequence of Pseudomonas chlororaphis Strain PA23.
<3>Genome Announcements
<4>2
<5>e00689-14
<6>2014
<7>Pseudomonas chlororaphis strain PA23 is a plant-beneficial bacterium that is able to suppress
disease caused by the fungal pathogen Sclerotinia sclerotiorum
through a process known as biological control. Here we present a 7.1-Mb assembly
of the PA23 genome.

<>

<1>Loftus, B. et al.
<2>The genome of the protist parasite Entamoeba histolytica.
<3>Nature
<4>433
<5>865-868
<6>2005
<7>Entamoeba histolytica is an intestinal parasite and the causative agent of amoebiasis, which
is a significant source of morbidity and mortality in developing countries. Here we present
the genome of E. histolytica, which reveals a variety of metabolic adaptations shared with two
other amitochondrial protist pathogens: Giardia lamblia and Trichomonas vaginalis. These
adaptations include reduction or elimination of most mitochondrial metabolic pathways and the
use of oxidative stress enzymes generally associated with anaerobic prokaryotes. Phylogenomic
analysis identifies evidence for lateral gene transfer of bacterial genes into the E.
histolytica genome, the effects of which centre on expanding aspects of E. histolytica's
metabolic repertoire. The presence of these genes and the potential for novel metabolic
pathways in E. histolytica may allow for the development of new chemotherapeutic agents. The
genome encodes a large number of novel receptor kinases and contains expansions of a variety
of gene families, including those associated with virulence. Additional genome features
include an abundance of tandemly repeated transfer-RNA-containing arrays, which may have a
structural function in the genome. Analysis of the genome provides new insights into the
workings and genome evolution of a major human pathogen.

<>

<1>Loizos, N., Silva, G.H., Belfort, M.
<2>Intron-encoded endonuclease I-TevII binds across the minor groove and induces two distinct conformational changes in its DNA substrate.
<3>J. Mol. Biol.
<4>255
<5>412-424
<6>1996
<7>I-TevII is the homing endonuclease encoded by the sunY intron of bacteriophage T4.  The enzyme
cleaves an intronless sunY gene near the exon I-exon II junction, thereby initiating intron
homing into its cognate intronless allele.  Specifically, I-TevII cleaves its DNA target 13 to
15 nucleotides downstream of the sunY intron insertion site, generating 2-nt 3'-OH
extensions.  Here, we present evidence that I-TevII makes predominantly minor groove contacts
in two regions of its recognition sequence, as does I-TevI, the other homing endonuclease
encoded by phage T4.  Following cleavage, I-TevII was shown to remain bound to one of its DNA
products, suggesting possible additional roles for the endonuclease in the mobility process.
Interestingly, two distinct conformational changes were detected by gel analysis in the DNA
substrate following binding by I-TevII, one occurring in the absence of Mg2+, the second being
dependent on the presence of Mg2+.  The Mg2+-induced distortion accompanies a nick in one
strand, and may serve to bring the cleavage site on the other strand into proximity with the
catalytic domain of the protein.

<>

<1>Loizos, N., Tillier, E.R.M., Belfort, M.
<2>Evolution of mobile group I introns: recognition of intron sequences by an intron-encoded endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>11983-11987
<6>1994
<7>Mobile group I introns are hypothesized to have arisen after invasion by endonuclease-encoding
open reading frames (ORFs) which mediate their mobility. Consistent with an endonuclease-ORF
invasion event, we report similarity between exon junction sequences (the recognition site for
the mobility endonuclease) and intron sequences flanking the endonuclease ORF in the sunY gene
of phage T4. Furthermore, we have demonstrated the ability of the intron-encoded endonuclease
to recognize and cleave these intron sequences when present in fused form in synthetic
constructs. These observations and accompanying splicing data are consistent with models in
which the invading endonuclease ORF is provided safe haven within a splicing element. In turn
the intron is afforded immunity to the endonuclease product, which imparts mobility to the
intron.

<>

<1>Loman, N.J., Snyder, L.A., Linton, J.D., Langdon, R., Lawson, A.J., Weinstock, G.M., Wren, B.W., Pallen, M.J.
<2>Genome sequence of the emerging pathogen Helicobacter canadensis.
<3>J. Bacteriol.
<4>191
<5>5566-5567
<6>2009
<7>We determined the genome sequence of the type strain of Helicobacter
canadensis, an emerging human pathogen with diverse animal reservoirs.
Potential virulence determinants carried by the genome include systems for
N-linked glycosylation and capsular export. A protein-based phylogenetic
analysis places H. canadensis close to Wolinella succinogenes.

<>

<1>Lomovskaya, N.D., Chater, K.F., Mkrtumian, N.M.
<2>Genetics and Molecular Biology of Streptomyces Bacteriophages.
<3>Microbiol. Rev.
<4>44
<5>206-229
<6>1980
<7>The streptomycetes are genetically among the most tractable of bacteria.  In
the best-studied strain, Streptomyces coelicolor A3, chromosome mapping is
straightforward, and a single circular linkage map of more than 100 markers has
been established.  Extremely efficient protoplast fusion can be induced by
polyethylene glycol.  Plasmids have been demonstrated both physically and
genetically, and some of them are self-transmissible sex factors having a
variety of interactions with the host chromosome.  Reintroduction
(transformation) of isolated plasmid deoxyribonucleic acid (DNA) into
protoplasts occurs at high frequency in the presence of polyethylene glycol.
These features are important in the study of several special characteristics of
streptomycetes, in particular, their remarkable status as antibiotic producers
and their morphological complexity.  Much greater progress will be possible in
these investigations when the techniques already available are supplemented by
gene cloning and transposon genetics and by systems for fine-structure mapping.
Among the important requirements for the development and optimum exploitation
of such techniques is an understanding of Streptomyces phages, particularly of
their genetics and DNA. About a decade ago it was possible for major reviews of
S. coelicolor genetics and Streptomyces phages to be completely independent of
each other; the potential benefits of using a genetically studied host in phage
studies, and vice versa, were unfulfilled.  More recently, considerable
advances have been made in the genetic and physical analysis of these phages
and their interactions with their hosts.  Thus, when temperate phages acting on
S. coelicolor A3 were isolated, it became possible to initiate studies of such
problems as the relationship between phages and differentiated bacteria,
comparison with model eubacterial phage-host systems, the genetic basis of
lysogenization in streptomycetes, the structure and function of the genomes of
Streptomyces phages, and the development of Streptomyces phages as DNA-cloning
vectors.  Both temperate and virulent phages have been instrumental in
understanding Streptomyces host-controlled restriction and modification (RM)
systems, and they have also helped in various aspects of the genetic analysis
of streptomycetes. The most extensive studies have been of the temperate phage
UC31, and we will focus on this phage as a main theme, with occasional
variations provided by observations on some other phages.  We shall then
consider these and other results in the context of the use of phages as vectors
for the introduction of particular DNA segments into streptomycetes, and
finally review interactions between the phages and host-controlled RM systems.

<>

<1>Long, M., Ruan, L., Yu, Z., Xu, X.
<2>Genome sequence of Pseudomonas sp. S9, an extracellular arylsulfatase producing bacterium isolated from the mangrove soil.
<3>J. Bacteriol.
<4>193
<5>4041
<6>2011
<7>Pseudomonas sp. S9 was originally isolated from the mangrove soil in Xiamen, China. It is an
aerobic bacterium which shows extracellular
arylsulfatase activity. Here, we describe the 4.8 Mb draft genome sequence
of Pseudomonas sp. S9 which exhibits novel cysteine-type sulfatases.

<>

<1>Longley, M.J., Mosbaugh, D.W.
<2>Characterization of DNA metabolizing enzymes in situ following polyacrylamide gel electrophoresis.
<3>Biochemistry
<4>30
<5>2655-2664
<6>1991
<7>We have detected the in situ activities of DNA glycosylase, endonuclease, DNA
polymerase, and DNA ligase using a novel polyacrylamide activity gel
electrophoresis procedure.  DNA metabolizing enzymes were resolved through
either native or SDS-polyacrylamide gels containing defined 32P-labeled
oligonucleotides annealed to M13 DNA.  After electrophoresis, these enzymes
catalyzed in situ reactions and their [32P]DNA products were resolved from the
gel by a second dimension of electrophoresis through a denaturing DNA
sequencing gel.  Detection of modified (degraded or elongated) oligonucleotide
chains was used to locate various enzyme activities.  The catalytic and
physical properties of Novikoff hepatoma DNA polymerase beta were found to be
similar under both in vitro and in situ conditions.  With 3'-terminally matched
and mismatched [32P]DNA substrates in the same activity gel, DNA polymerase
and/or 3' to 5' exonuclease activities of Escherichia coli DNA polymerase I
(large fragment), DNA polymerase III (holoenzyme), and exonuclease III were
detected and characterized.  In addition, use of matched and mismatched DNA
primers permitted the uncoupling of mismatch excision and chain extension
steps.  Activities first detected in nondenaturing activity gels as either
multifunctional or multimeric enzymes were also identified in denaturing
activity gels, and assignment of activities to specific polypeptides suggested
subunit composition.  Furthermore, DNA substrates cast within polyacrylamide
gels were successfully modified by the exogenous enzymes polynucleotide kinase
and alkaline phosphatase before and after in situ detection of E. coli DNA
ligase activity, respectively.  Several restriction endonucleases and the
tripeptide (Lys-Trp-Lys), which acts as an apurinic/apyrimidinic endonuclease,
were able to diffuse into gels and modify DNA.  This ability to create
intermediate substrates within activity gels could prove extremely useful in
delineating the steps of DNA replication and repair pathways.

<>

<1>Longo, M.C., Smith, M.D.
<2>Cloned NsiI restriction-modification system.
<3>US Patent Office
<4>US 5470740
<5>
<6>1995
<7>This present invention discloses the cloning and expression in Escherichia coli of the NsiI
restriction-modification system from Neisseria sicca, utilizing a two step protocol.  Initial
protection of E. coli host with methylase expressed on a plasmid was required to stabilize a
compatible plasmid carrying the endonuclease gene on a single DNA fragment.  A chromosomal map
was generated localizing the genes for NsiI methylase and endonuclease.  An E. coli strain was
constructed which produced high levels of NsiI endonuclease.

<>

<1>Longo, M.C., Smith, M.D.
<2>Cloned NsiI restriction-modification system.
<3>US Patent Office
<4>US 5670359
<5>
<6>1997
<7>The present invention discloses the cloning and expression in Escherichia coli of the NsiI
restriction-modification system from Neisseria sicca, utilizing a two step protocol.  Initial
protection of the E. coli host with methylase expressed on a plasmid was required to stabilize
a compatible plasmid carrying the endonuclease gene on a single DNA fragment.  A chromosomal
map was generated localizing the genes for NsiI methylase and endonuclease.  An E. coli strain
was constructed which produced high levels of NsiI endonuclease.

<>

<1>Longo, M.C., Smith, M.D., Chatterjee, D.K.
<2>Cloning and expressing restriction endonucleases and modification methylases from Caryophanon.
<3>US Patent Office
<4>US 5312746
<5>
<6>1994
<7>The present invention is directed to recombinant hosts which contain and express the ClaI
Type-II restriction endonuclease and/or modification methylase genes. The present invention is
also directed to vectors or DNA molecules which contain these genes, and to methods of
producing the enzymes. One source of these enzymes is Caryophanon latum, although other
microorganisms may be used to isolate the restriction endonuclease isoschizomers and
modification methylase isoschizomers of the invention.

<>

<1>Longo, M.C., Smith, M.D., Chatterjee, D.K.
<2>Cloning and expressing restriction endonucleases and modification methylases from caryophanon.
<3>European Patent Office
<4>EP 0605854 B
<5>
<6>2002
<7>The present invention is directed to an isolated DNA sequence coding for ClaI restriction
endonuclease obtainable from Caryophanon and an isolated DNA sequence coding for ClaI
modification methylase obtainable from Caryophanon.  The invention is further directed to
vectors comprising an isolated DNA sequence coding for ClaI restriction
endonuclease/modification methylase obtainable from Caryophanon.  The invention is further
directed to recombinant host cells comprising an isolated DNA sequence/vector coding for ClaI
restriction endonuclease/Modification methylase obtainable from Caryophanon.  The invention is
further directed to processes for obtaining ClaI restriction endonuclease/modification
methylase.  According to the process of this invention, the restriction endonuclease and/or
modification methylase is produced by culturing a recombinant host comprising an isolated DNA
sequence coding for ClaI restriction endonuclease/modification methylase and isolating enzymes
from the recombinant host.

<>

<1>Longo, M.C., Smith, M.D., Chatterjee, D.K.
<2>Cloning and expressing restriction endonucleases and modification  methylases from caryophanon.
<3>European Patent Office
<4>EP 1170362 A
<5>
<6>2002
<7>The present invention is directed to recombinant hosts which
contain and express the ClaI Type-II restriction endonuclease and/or
modification methylase genes.  The present invention is also directed to
vectors or DNA molecules which contain this gene, and to methods of
producing the enzymes.  One source of this enzyme is Caryophanon latum,
although other microorganisms may be used to isolate the restriction
endonuclease isoschizomers and modification methylase isoschizomers of the
invention.

<>

<1>Longo, M.C., Smith, M.D., Harris, R.D.
<2>Cloned SstI/SacI restriction-modification system.
<3>International Patent Office
<4>WO 9607736
<5>
<6>1996
<7>The present invention discloses the cloning and expression in Escherichia coli of the
SstI/SacI restriction-modification system.  The SstI methylase gene was cloned by methylase
selection (the "Hungarian Trick") and the orientation and boundaries of the gene were
established.  The level of methylation of the host was improved significantly by expressing
the SstI methylase from the strong RPSL promoter.  Using the SstI methylase clone as a probe,
a chromosomal map was generated in the area of the gene for the SstI methylase.  DNA
downstream from the methylase was cloned and found to contain the gene for the SstI
endonuclease.  The orientation of the SstI endonuclease was established and a strong promoter
was placed close to the SstI endonuclease gene by generating nested deletions.  By these
methods an E. coli strain was constructed which produced high levels of SstI endonuclease.
The SacI locus in the S. achromogenes chromosome was found to be identical to the SstI locus
in S. stanford in all measured respects.

<>

<1>Longo, M.C., Smith, M.D., Harris, R.D.
<2>Cloned SstI/SacI restriction-modification system.
<3>US Patent Office
<4>US 5534428
<5>
<6>1996
<7>The present invention discloses the cloning and expression in Escherichia coli of the
SstI/SacI restriction-modification system.  The SstI methylase gene was cloned by methylase
selection (the "Hungarian Trick" and the orientation and boundaries of the gene were
established.  The level of methylation of the host was improved significantly by expressing
the SstI methylase from the strong RPSL promoter.  Using the SstI methylase clone as a probe,
a chromosomal map was generated in the area of the gene for the SstI methylase.  DNA
downstream from the methylase was cloned and found to contain the gene for the SstI
endonuclease.  The orientation of the SstI endonuclease was established and a strong promoter
was placed close to the SstI endonuclease gene by generating nested deletions.  By these
methods, an E. coli strain was constructed which produced high levels of SstI endonuclease.
The SacI locus in the S. achromogenes chromosome was found to be identical to the SstI locus
in S. stanford in all measured respects.

<>

<1>Longo, M.C., Smith, M.D., Harris, R.D.
<2>Cloned SstI/SacI restriction-modification system.
<3>US Patent Office
<4>US 5759839
<5>
<6>1998
<7>The present invention discloses the cloning and expression in Escherichia coli of the
SstI/SacI restriction-modification system.  The SstI methylase gene was cloned by methylase
selection (the "Hungarian Trick") and the orientation and boundaries of the gene were
established.  The level of methylation of the host was improved significantly by expressing
the SstI methylase from the strong RPSL promoter.  Using the SstI methylase clone as a probe,
a chromosomal map was generated in the area of the gene for the SstI methylase.  DNA
downstream from the methylase was cloned and found to contain the gene for the SstI
endonuclease.  The orientation of the SstI endonuclease was established and a strong promoter
was placed close to the SstI endonuclease gene by generating nested deletions.  By these
methods, an E. coli strain was constructed which produced high levels of SstI endonuclease.
The SacI locus in the S. achromogenes chromosome was found to be identical to the SstI locus
in S. Stanford in all measured respects.

<>

<1>Looney, M., Wilson, G.
<2>Method for producing the FokI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0321266 B
<5>
<6>1994
<7>The present invention relates to clones for the FokI restriction endonuclease and modification
methylase, and to the production of these enzymes from the clones.

<>

<1>Looney, M.C., Moran, L.S., Jack, W.E., Feehery, G.R., Benner, J.S., Slatko, B.E., Wilson, G.G.
<2>Nucleotide sequence of the FokI restriction-modification system:  separate strand-specificity domains in the methyltransferase.
<3>Gene
<4>80
<5>193-208
<6>1989
<7>The genes for FokI, a type-IIS restriction-modification system from
Flavobacterium okeanokoites (asymmetric recognition sequence:
5'-GGATG/3'-CCTAC), were cloned into Escherichia coli.  Recombinants carrying
the fokIR and fokIM genes were found to modify their DNA completely, and to
restrict lambdoid phages weakly.  The nucleotide sequences of the genes were
determined, and the probable start codons were confirmed by amino acid
sequencing.  The FokI endonuclease (R.FokI) and methyltransferase (M.FokI) are
encoded by single, adjacent genes, aligned in the same orientation, in the
order M then R.  The genes are large by the standards of type-II systems, 1.9
kb for the M gene, and 1.7 kb for the R gene.  Preceding each gene is a pair of
FokI recognition sites; it is conceivable that interactions between the sites
and the FokI proteins could regulate expression of the genes.

<>

<1>Looney, M.E.C., Wilson, G.G.
<2>Method for producing the FokI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 999294
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the FokI restriction
endonuclease by (1) introducing the restriction endonuclease gene from Flavobacterium
okeanokoites IFO 12536 into a host whereby the restriction gene is expressed (2) fermenting
the host which contains the vector encoding and expressing the FokI restriction endonuclease
and (3) purifying the FokI restriction endonuclease from the fermented host which contains the
vector encoding and expressing the FokI restriction endonuclease activity.

<>

<1>Lopatina, N., Haskell, J.F., Andrews, L.G., Poole, J.C., Saldanha, S., Tollefsbol, T.
<2>Differential maintenance and de novo methylating activity by three DNA methyltransferases in aging and immortalized fibroblasts.
<3>J. Cell. Biochem.
<4>84
<5>324-334
<6>2002
<7>Genomic methylation, which influences many cellular processes such as gene expression and
chromatin organization, generally declines with
cellular senescence although some genes undergo paradoxical
hypermethylation during cellular aging and immortalization. To explore
potential mechanisms for this process, we analyzed the methylating
activity of three DNA methyltransferases (Dnmts) in aging and
immortalized WI-38 fibroblasts. Overall maintenance methylating
activity by the Dnmts greatly decreased during cellular senescence. In
immortalized WI-38 cells, maintenance methylating activity was similar
to that of normal young cells. Combined de novo methylation activity of
the Dnmts initially decreased but later increased as WI-38 cells aged
and was strikingly elevated in immortalized cells. To further elucidate
the mechanisms for changes in DNA methylation in aging and immortalized
cells, the individual Dnmts were separated and individually assessed
for maintenance and de novo methylating activity. We resolved three
Dnmt fractions, one of which was the major maintenance
methyltransferase, Dnmt1, which declined steadily in activity with
cellular senescence and immortalization. However, a more basic Dnmt,
which has significant de novo methylating activity, increased markedly
in activity in aging and immortalized cells. We have identified this
methyltransferase as Dnmt3b which has an important role in neoplastic
transformation but its role in cellular senescence and immortalization
has not previously been reported. An acidic Dnmt we isolated also had
increased de novo methylating activity in senescent and immortalized
WI-38 cells. These studies indicate that reduced genome-wide
methylation in aging cells may be attributed to attenuated Dnmt1
activity but that regional or gene-localized hypermethylation in aging
and immortalized cells may be linked to increased de novo methylation
by Dnmts other than the maintenance methyltransferase.

<>

<1>Lopatina, N.G., Kirnos, M.D., Suchkov, S.V., Vanyushin, B.F., Nikolskaya, I.I., Debov, S.S.
<2>Determination of the recognition site for adenine-specific methylase of Shigella sonnei 47 by hydrazinolysis of DNA, followed by separation of the purine oligonucleotides by thin-layer chromatography on DEAE-cellulose.
<3>Biokhimiia
<4>50
<5>495-502
<6>1985
<7>A method has been developed for the separation of oligopurine units according
to length and composition by two-dimensional thin-layer chromatography on
plates with DEAE-cellulose, permitting a comparative analysis of the content of
various purine isopliths in DNA of different origin.  In the case of the
analysis of methylated DNA, the method permits a comparison of the substrate
specificity of various enzymes of methylation of the adenine residues in DNA.
In conjunction with enzymatic treatment of labeled methylated isopliths, the
method permits determination of the methylatable sequence and in a number of
cases an ascertainment of the recognition site for adenine-specific methylase
as a whole.  The proposed method was used to establish the fact that the
methylase SsoI recognizes the sequence 5' ...G-*A-A-T-T-C...3' and methylates
the adenine residue closest to its 5'-end.

<>

<1>Lopatina, N.G., Lopareva, E.N., Posypanova, A.M., Nikolskaya, A., Debov, S.S.
<2>Cytosine DNA-methylase SsoII from Shigella sonnei 47 cells.
<3>Biokhimiia
<4>53
<5>1265-1269
<6>1988
<7>Shigella sonnei 47 cells were found to contain DNA-methylase SsoII which is a
modifying component of the system of host specificity of SsoII.  The
recognition sequence (RS) of methylase SsoII is represented by a five-member
palindromic structure -5'...CCNGG...3'- with a degenerate central nucleotide.
Modification of SsoII affords protection of acceptor DNA not only from SsoII
type restriction, but also from other restrictases, e.g., EcoRII having an
analogous RS but with a less degenerate central nucleotide pair.  A simple and
rapid procedure for isolation and purification of DNA-methylase SsoII, which
employs hydrophobic chromatography on phenyl-Sepharose, has been developed.
The enzyme preparation does not contain trace amounts of specific and
nonspecific endonucleases and keeps stable on storage in 30% glycerol over a
period of one year.

<>

<1>Lopatina, N.G., Nikolskaya, I.I., Buryanov, Y.I., Debov, S.S.
<2>Presence of 2 DNA-methylases of cytosine in Escherichia coli CK cells.
<3>Dokl. Akad. Nauk.
<4>240
<5>1475-1477
<6>1978
<7>
<>

<1>Lopatina, N.G., Nikolskaya, I.I., Sverdiov, E.D., Debov, S.S.
<2>Determination of the recognition sites of adenine DNA-methylases from Escherichia coli CK.
<3>Vopr. Med. Khim.
<4>27
<5>220-223
<6>1981
<7>DNA-methylases from a strain of E. coli CK were studied.  Three adenine methylases were found
in the strain studied, which were eluted by 0.16M, 0.23M and 0.7M NaCl in phosphocellulose
P-11 chromatography.  According to this, the enzymes were designated as DM-A16, DM-A23 and
DM-A-70.  Indirect data on the presence of adenine specific methylases dissimilar in their
recognition sites in cells of E. coli CK were obtained using the test of additional
methylation modified by I.I. Nikolskaya.  These conclusions were confirmed, when the
dinucleotide sequence was determined in the recognition site using DM-A23 and DM-A70.  The
methylase DM-A23 was shown to recognize the dinucleotide sequence 5'...A-G...3'.

<>

<1>Lopatina, N.G., Suchkov, S.V., Kulikov, S.M., Nikolskaya, I.I., Debov, S.S.
<2>Site-specificity of DNA methylases from Shigella sonnei 47 cells.
<3>Biokhimiia
<4>50
<5>1694-1701
<6>1985
<7>A complex approach involving isoplith analysis, enzymatic treatment of methylated isopliths
and computer analysis of experimental data has been used to determine site specificity of six
methylases from Shigella sonnei 47 cells termed according to their base specificity and pI as
MC4.2, MC5.3, MC6.2, MC7.4, MC8.4 and MA9.5.  It has been found that the recognition site of
MA9.5 is a palindrome of the 5' ...GAATTC...3' type and that this enzyme is an isomethylomer
of M.EcoRI.  It has been demonstrated for the first time for methylases that the recognition
site of MC4.2 is represented by a non-symmetrical four-member sequence, 5'...NCCCCN...3'
characterized by unique blocking of cytosines.  MC8.4 possesses a broad specificity of
substrate recognition and methylates the cytosine residue within the non-symmetrical unique
sequence 5'...N(C/Pu) CCN...3', whose 5'-terminal base is depleted in three nucleotides.
MC5.3 methylates the 3'-terminal cytosine residue within the composition of the
pentanucleotide palindrome recognition site, 5'...CCNGG...3' MC6.2 and MC7.4 possess
identical pentanucleotide recognition sites, 5'...(Py)CNG(Pu)...3', but are distinguished by
pI.  The latter finding has been shown for the first time for different methylases within one
strain.

<>

<1>Loper, J.E. et al.
<2>Comparative Genomics of Plant-Associated Pseudomonas spp.: Insights into Diversity and Inheritance of Traits Involved in Multitrophic Interactions.
<3>PLoS Genet.
<4>8
<5>E1002784
<6>2012
<7>We provide here a comparative genome analysis of ten strains within the
Pseudomonas fluorescens group including seven new genomic sequences. These
strains exhibit a diverse spectrum of traits involved in biological control and
other multitrophic interactions with plants, microbes, and insects. Multilocus
sequence analysis placed the strains in three sub-clades, which was reinforced by
high levels of synteny, size of core genomes, and relatedness of orthologous
genes between strains within a sub-clade. The heterogeneity of the P. fluorescens
group was reflected in the large size of its pan-genome, which makes up
approximately 54% of the pan-genome of the genus as a whole, and a core genome
representing only 45-52% of the genome of any individual strain. We discovered
genes for traits that were not known previously in the strains, including genes
for the biosynthesis of the siderophores achromobactin and pseudomonine and the
antibiotic 2-hexyl-5-propyl-alkylresorcinol; novel bacteriocins; type II, III,
and VI secretion systems; and insect toxins. Certain gene clusters, such as those
for two type III secretion systems, are present only in specific sub-clades,
suggesting vertical inheritance. Almost all of the genes associated with
multitrophic interactions map to genomic regions present in only a subset of the
strains or unique to a specific strain. To explore the evolutionary origin of
these genes, we mapped their distributions relative to the locations of mobile
genetic elements and repetitive extragenic palindromic (REP) elements in each
genome. The mobile genetic elements and many strain-specific genes fall into
regions devoid of REP elements (i.e., REP deserts) and regions displaying
atypical tri-nucleotide composition, possibly indicating relatively recent
acquisition of these loci. Collectively, the results of this study highlight the
enormous heterogeneity of the P. fluorescens group and the importance of the
variable genome in tailoring individual strains to their specific lifestyles and
functional repertoire.

<>

<1>Lopes, T. et al.
<2>Complete Genome Sequence of Corynebacterium pseudotuberculosis Strain Cp267, Isolated from a Llama.
<3>J. Bacteriol.
<4>194
<5>3567-3568
<6>2012
<7>In this work we report the genome of Corynebacterium pseudotuberculosis strain 267, isolated
from a llama. This pathogen is of great veterinary and economic
importance, as it is the cause of caseous lymphadenitis in several livestock
species around the world and causes significant losses due to the high cost of
treatment.

<>

<1>Lopez, O.J., Nelson, R.M.
<2>DNA methyltransferase genotyping.
<3>International Patent Office
<4>WO 9910540
<5>
<6>1999
<7>DNA methyltransferases can be utilized in methods for quickly and accurately: determining
variations, mutations or polymorphisms in DNA sequences; identifying specific alleles in
single copy genes; creating genomic fingerprints; creating DNA paints; and generating ordered
maps.

<>

<1>Lopez, O.J., Nelson, R.M.
<2>DNA methyltransferase genotyping.
<3>US Patent Office
<4>US 6514698
<5>
<6>2003
<7>DNA Methyltransferases can be utilized in methods for quickly and accurately: determining
variations, mutations or polymorphisms in DNA
sequences; identifying specific alleles in single copy genes; creating
genomic fingerprints; creating DNA Paints; and generating ordered maps.

<>

<1>Lopez, O.J., Quintanar, A., Padhye, N.V., Nelson, M.
<2>Genotyping of DNA using sequence-specific methyltransferases followed by immunochemical detection.
<3>J. Immunoassay Immunochem.
<4>24
<5>11-28
<6>2003
<7>Modern molecular genetics relies on the ability to map the positions of genes on chromosomes,
relative to known DNA markers. The first such DNA
markers described were Restriction Fragment Length Polymorphisms, but any
restriction endonuclease used for RFLP mapping is just one member of a
restriction-modification pair. For each restriction endonuclease, there is
a companion methyltransferase (MTase) that has the same DNA sequence
specificity. Therefore, in principle, it should be possible to use MTases
rather than restriction enzymes to detect polymorphic sites in DNA. We
have used sequence-specific DNA MTases to detect polym orphisms in closely
related viral pathogens. If at least one MTase recognition site is present
in PCR-amplified DNA, then methyl groups are incorporated; if no MTase
site is present, then methyl groups are not incorporated. When several
different sequence-specific DNA MTase reactions are carried out, the
pattern of methyl incorporation defines a DNA MTase genotype. DNA MTase
Genotyping (DMG) can be used to rapidly diagnose heritable or infectious
diseases, to immunochemically detect DNA at defined 2 to 8 base pair
sites, or to characterize the amplicons by constructing ordered maps.

<>

<1>Lopez-Garrido, J., Casadesus, J.
<2>Regulation of Salmonella enterica Pathogenicity Island 1 by DNA Adenine Methylation.
<3>Genetics
<4>184
<5>637-649
<6>2010
<7>DNA adenine methylase (Dam ) mutants of Salmonella enterica are attenuated in the mouse model
and present multiple virulence-related
defects. Impaired interaction of Salmonella Dam mutants with the
intestinal epithelium has been tentatively correlated with reduced
secretion of pathogenicity island 1 (SPI-1) effectors. In this study,
we show that S. enterica Dam mutants contain lowered levels of the
SPI-1 transcriptional regulators HilA, HilC, HilD, and InvF. Epistasis
analysis indicates that Dam-dependent regulation of SPI-1 requires
HilD, while HilA, HilC, and InvF are dispensable. A transcriptional
hilD::lac fusion is expressed at similar levels in Dam(+) and Dam
hosts. However, lower levels of hilD mRNA are found in a Dam
background, thus providing unsuspected evidence that Dam methylation
might exert post-transcriptional regulation of hilD expression. This
hypothesis is supported by the following lines of evidence: (i) lowered
levels of hilD mRNA are found in Salmonella Dam mutants when hilD is
transcribed from a heterologous promoter; (ii) increased hilD mRNA
turnover is observed in Dam mutants; (iii) lack of the Hfq RNA
chaperone enhances hilD mRNA instability in Dam mutants; and (iv) lack
of the RNA degradosome components polynucleotide phosphorylase and
ribonuclease E suppresses hilD mRNA instability in a Dam background.
Our report of Dam-dependent control of hilD mRNA stability suggests
that DNA adenine methylation plays hitherto unknown roles in
post-transcriptional control of gene expression.

<>

<1>Lopez-Lopez, A., Richter, M., Pena, A., Tamames, J., Rossello-Mora, R.
<2>New insights into the archaeal diversity of a hypersaline microbial mat obtained by a metagenomic approach.
<3>Syst. Appl. Microbiol.
<4>36
<5>205-214
<6>2013
<7>A metagenomic approach was carried out in order to study the genetic pool of a
hypersaline microbial mat, paying more attention to the archaeal community and,
specifically, to the putatively methanogenic members. The main aim of the work
was to expand the knowledge of a likely ecologically important archaeal lineage,
candidate division MSBL1, which is probably involved in methanogenesis at very
high salinities. The results obtained in this study were in accordance with our
previous report on the bacterial diversity encountered by using a number of
molecular techniques, but remarkable differences were found in the archaeal
diversity retrieval by each of the procedures used (metagenomics and 16S
rRNA-based methods). The lack of synteny for most of the metagenomic fragments
with known genomes, together with the low degree of similarity of the annotated
open reading frames (ORFs) with the sequences in the databases, reflected the
high degree of novelty in the mat community studied. Linking the sequenced clones
with representatives of division MSBL1 was not possible because of the lack of
additional information concerning this archaeal group in the public gene
repositories. However, given the high abundance of representatives of this
division in the 16S rRNA clone libraries and the low identity of the archaeal
clones with known genomes, it was hypothesized that some of them could arise from
MSBL1 genomes. In addition, other prokaryotic groups known to be relevant in
organic matter mineralization at high salinities were detected.

<>

<1>Lopez-Perez, M., Gonzaga, A., Rodriguez-Valera, F.
<2>Genomic Diversity of 'Deep Ecotype' Alteromonas macleodii Isolates: Evidence for Pan-Mediterranean Clonal Frames.
<3>Genome Biol. Evol.
<4>5
<5>1220-1232
<6>2013
<7>We have compared genomes of Alteromonas macleodii "deep ecotype" isolates from
two deep Mediterranean sites and two surface samples from the Aegean and the
English Channel. A total of nine different genomes were analyzed. They belong to
five clonal frames (CFs) that differ among them by approximately 30,000
single-nucleotide polymorphisms (SNPs) over their core genomes. Two of the CFs
contain three strains each with nearly identical genomes ( approximately 100 SNPs
over the core genome). One of the CFs had representatives that were isolated from
samples taken more than 1,000 km away, 2,500 m deeper, and 5 years apart. These
data mark the longest proven persistence of a CF in nature (outside of clinical
settings). We have found evidence for frequent recombination events between or
within CFs and even with the distantly related A. macleodii surface ecotype. The
different CFs had different flexible genomic islands. They can be classified into
two groups; one type is additive, that is, containing different numbers of gene
cassettes, and is very variable in short time periods (they often varied even
within a single CF). The other type was more stable and produced the complete
replacement of a genomic fragment by another with different genes. Although this
type was more conserved within each CF, we found examples of recombination among
distantly related CFs including English Channel and Mediterranean isolates.

<>

<1>Lopez-Perez, M., Rodriguez-Valera, F., Webb, H.K., Crawford, R.J., Ivanova, E.P.
<2>Genome Sequence of 'Thalassospira australica' NP3b2T Isolated from St. Kilda Beach, Tasman Sea.
<3>Genome Announcements
<4>2
<5>e01139-14
<6>2014
<7>Here, we present the draft genome of 'Thalassospira australica' NP3b2(T), a potential
poly(ethylene terephthalate) (PET) plastic biodegrader. This genomic
information will enhance information on the genetic basis of metabolic pathways
for the degradation of PET plastic.

<>

<1>Lorincz, M.C., Groudine, M.
<2>CmC(a/t)GG methylation: a new epigenetic mark in mammalian DNA?
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>10034-10036
<6>2001
<7>Twenty-five years ago, based on the knowledge of cytosine methylation in higher organisms and
the newly discovered bacterial adenine methyltransferase, Riggs and Holliday and Pugh
independently proposed that the covalent modification of DNA by methylation might serve as a
means to propagate heritable expression states in eukaryotes.  In the years since, the
association between cytosine methylation and transcriptional silencing in mammalian cells has
become well established, and a number of proteins that catalyze the transfer of a methyl group
to the 5-carbon of the cytosine pyrimidine ring have been cloned and characterized.  These DNA
methyltransferases (m5C-MTases) are encoded by a diverse family of genes found in prokaryotes
as well as all four groups of eukaryotes.

<>

<1>Lorincz, M.C., Schubeler, D., Hutchinson, S.R., Dickerson, D.R., Groudine, M.
<2>DNA methylation density influences the stability of an epigenetic imprint and Dnmt3a/b-independent de novo methylation.
<3>Mol. Cell. Biol.
<4>22
<5>7572-7580
<6>2002
<7>DNA methylation plays an important role in transcriptional repression. To gain insight into
the dynamics of demethylation and de novo methylation, we introduced a proviral reporter,
premethylated at different densities, into a defined chromosomal site in murine
erythroleukemia cells and monitored the stability of the introduced methylation and reporter
gene expression. A high density of methylation was faithfully propagated in vivo. In contrast,
a low level of methylation was not stable, with complete demethylation and associated
transcriptional activation or maintenance-coupled de novo methylation and associated silencing
occurring with equal probability. Deletion of the proviral enhancer increased the probability
of maintenance-coupled de novo methylation, suggesting that this enhancer functions in part to
antagonize such methylation. The DNA methyltransferases (MTases) Dnmt3a and Dnmt3b are thought
to be the sole de novo MTases in the mammalian genome. To determine whether these enzymes are
responsible for maintenance-coupled de novo methylation, the unmethylated or premethylated
proviral reporter was introduced into DNA MTase-deficient embryonic stem cells. These studies
revealed the presence of a Dnmt3a/Dnmt3b-independent de novo methyltransferase activity that
is stimulated by the presence of preexisting methylation.

<>

<1>Lou, S., Wu, L.
<2>Isolation of thermostable restriction endonuclease from thermophilic Synechococcus elongatus T3.
<3>J. Xiamen Univ.
<4>36
<5>272-275
<6>1997
<7>A restriction endonuclease was isolated from thermophilic cyanobacterium Synechococcus
elongatus T3, which was named SelI.  This restriction endonuclease was characterized as an
isoschizomer of Sau96I, its recognition sequences is GGNCC.  Bu the reaction conditions of
SelI are different from that of Sau96I.  SalI is a thermostable restriction endonuclease with
optimal reaction temperature 50-70oC.

<>

<1>Low, D.A., Casadesus, J.
<2>Clocks and switches: bacterial gene regulation by DNA adenine methylation.
<3>Curr. Opin. Microbiol.
<4>11
<5>106-112
<6>2008
<7>N(6) methylation in adenosine moieties causes changes in DNA structure and can modulate
DNA-protein interactions. In both alpha-Proteobacteria and gamma-Proteobacteria,
postreplicative formation of N(6)-methyl-adenine regulates transcription of specific genes and
provides two general types of controls: (i) clock-like controls that permit transient gene
transcription during a specific stage of DNA replication; (ii) switch-like controls in which
transcription is regulated by a DNA methylation pattern. DNA adenine methylation may also
regulate gene expression by affecting nucleoid topology. Recent transcriptomic studies have
unveiled novel cases of genes regulated by DNA adenine methylation, including virulence genes
of bacterial pathogens.

<>

<1>Low, D.A., Weyand, N.J., Mahan, M.J.
<2>Roles of DNA adenine methylation in regulating bacterial gene expression and virulence.
<3>Infect. Immun.
<4>69
<5>7197-7204
<6>2001
<7>DNA methylation provides a mechanism by which additional information is imparted to DNA, and
such epigenetic information can alter the timing and targeting of cellular events.  DNA
methylation occurs throughout the living world, including bacteria, plants, and mammals. Until
recently, methylated DNA sequences were not detected in the fruit fly, in brewer's yeast, or
in the nematode.  However, analysis by Lyko and colleagues showed that Drosophila melanogaster
does contain methylated DNA, and thus it is possible that yeast and worms may also have it.
In this review, we focus our attention on the roles of DNA methylation in regulating bacterial
gene expression and virulence.  Although some background information about DNA methylation is
presented, we refer the reader to excellent reviews on the subject.  DNA methylation occurs at
the C-5 or N-4 positions of cytosine and at the N-6 position of adenine and is catalyzed by
enzymes known as DNA methyltransferases (Mtases).  All MTases use S-adenosyl methionine as a
methyl donor.  DNA methylation has historically been associated with DNA
restriction-modification systems thought to be important in protecting cells from foreign DNAs
such as transposons and viral DNAs.  Restriction-modification systems contain a DNA methylase
that protects host DNA sequences from restriction with their cognate restriction enzymes,
which digest unmodified foreign DNAs.  Certain MTases, including DNA cytosine MTase (Dcm),
which methylates the C-5 position of cytosine in CC(A/T)GG sequences, DNA adenine methylase
(Dam), which methylates N-6 of adenine in GATC sequences, and cell cycle-regulated methylase
(CcrM), which methylates the N-6 adenine of GAnTC, do not have cognate restriction enzymes
associated with them.  These methylases participate in cellular regulatory events, including
those that control bacterial virulence, which are the primary focus of this review.

<>

<1>Lower, M.
<2>Methods and probes for preparation of restriction endonucleases with desired specifity using in vitro cell-free compartmentalization systems and FRET and FACS selection.
<3>International Patent Office
<4>WO 201137485 A
<5>
<6>2011
<7>Method of preparation of restriction endonucleases, particularly those exhibiting the desired
sequential specificity consists in that a fluorescence-marked DNA probe is used for screening
a library of mutants, preferably in IVC format, and/or using other high-performance screening
technique, which is attained through expression of proteins included in the library of mutants
in a cell-free system in the presence and by means of the DNA probe, and proteins thus
obtained, resulting from expression of clones from the library, degrade the DNA probe, if
their substrate specificity matches the searched one, the degradation of the DNA probe being
detected as a disapparance of the FRET phenomenon between fluorescence markers included in the
probe, and then microcompartments in which the FRET phenomenon ceases to occur, are separated
from the remaining ones using Fluorescence Activated Cell Sorter and/or other equipment for
HTS analysis, and then DNA coding clones capable of degrading the probe are amplified using
polymerase chain reaction technique and are used as a basis for construction of the subsequent
library of mutants, which is searched during the subsequent round of screening, according to
the scheme mentioned above, and the subsequent rounds of screening are carried out until the
enzyme having the desired properties is obtained.  The fluorescence-marked DNA probe is
characterized in that the markers of the DNA probe are located in a direct vicinity of
recognizable sequence by searched restriciotn enzyme and/or in the vicinity of DNA restriciton
sites, and between the markers the FRET (Free Radiationless Energy Transfer) phenomenon
occurs.

<>

<1>Lowery, D.E., Fuller, T.E., Kennedy, M.J.
<2>Anti-bacterial vaccine compositions.
<3>Japanese Patent Office
<4>JP 2002541790 A
<5>
<6>2002
<7>
<>

<1>Lowery, D.E., Fuller, T.E., Kennedy, M.J.
<2>Anti-bacterial vaccine compositions.
<3>US Patent Office
<4>US 7476391 B
<5>
<6>2009
<7>Gram negative bacterial virulence genes are identified, thereby allowing the identification of
novel anti-bacterial agents that target these virulence genes and their products, and the
provision of novel gram negative bacterial mutants useful in vaccines.

<>

<1>Lowery, D.E., Fuller, T.E., Kennedy, M.J.
<2>Anti-Bacterial Vaccine Compositions.
<3>Korean Patent Office
<4>KR 1020067025082 A
<5>
<6>2006
<7>
<>

<1>Lowery, D.E., Fuller, T.E., Kennedy, M.J.
<2>Anti-Bacterial Vaccine Compositions.
<3>Korean Patent Office
<4>KR 1020067027580 A
<5>
<6>2006
<7>
<>

<1>Lowery, D.E., Fuller, T.E., Kennedy, M.J.
<2>Anti-bacterial vaccine compositions.
<3>European Patent Office
<4>EP 1860117 A
<5>
<6>2007
<7>Gram negative bacterial virulence genes are identified, thereby allowing the identification of
novel-bacterial agents that target these virulence genes and their products, and the provision
of novel gram negative bacterial mutants useful in vaccines.

<>

<1>Lowery, D.E., Fuller, T.E., Kennedy, M.J.
<2>Anti-Bacterial Vaccine Compositions.
<3>Korean Patent Office
<4>KR 1020067022704 A
<5>
<6>2006
<7>
<>

<1>Loyola, C., Saavedra, C., Gomez, I., Vasquez, C.
<2>The amino acidic substitution of cysteine 167 by serine (C167S) in BstVI restriction endonuclease of Bacillus stearothermophilus V affects its conformation and thermostability.
<3>Biochimie
<4>81
<5>261-266
<6>1999
<7>The restriction endonuclease BstVI from Bacillus stearothermophilus V contains three cysteine
residues at positions 134, 167 and 180. Titration of Cys residues with DTNB showed that none
of them are involved in disulphide bond formation. Cysteine triplets 134 and 167 were modified
by recombinant PCR to introduce a serine residue in each case. The mutated genes were cloned
into pGEM-T vector and transformed into E. coli JM109. Even though pGEM-T is not designed for
expression, the mutant proteins were efficiently expressed in E. coli. The endonuclease
carrying the mutation C134S was purified to homogeneity but appeared to be very unstable. In
contrast, the C167S mutant enzyme was stable when pure and was studied biochemically. This
mutant enzyme was as stable and resistant to protein-denaturing agents as the wild type
enzyme. The activity of both enzymes was not affected by preincubations of 2 h at 80 degrees
C. A short preincubation at 95 degrees C caused a complete inactivation of the mutant enzyme
while the wild type endonuclease retained 30% of its activity. Moreover, the C167S BstVI was
more susceptible to be hydrolyzed by proteinase K and trypsin compared to the wild type
endonuclease. These results show that the substitution Cys --> Ser at position 167 affects the
configuration and thermostability of BstVI restriction endonuclease.

<>

<1>Lu, A.-L., Jack, W.E., Modrich, P.
<2>DNA determinants important in sequence recognition by EcoRI endonuclease.
<3>J. Biol. Chem.
<4>256
<5>13200-13206
<6>1981
<7>Alkylation interference and protection methods (Siebenlist, U., and Gilbert,
W., (1980) Proc. Natl. Acad. Sci. USA 77, 122-126) have been utilized to deduce
potential DNA contracts involved in specific complex formation between EcoRI
endonuclease and its recognition sequence.  The endonuclease protected the N7
position (major groove) of the dG and the N3 position (minor groove) of both dA
residues within the EcoRI sequence against alkylation by dimethylsulfate,
d(7Gp7Ap7ApTpTpC), suggesting the presence of polypeptide in both grooves in
the vicinity of affected nitrogens.  Results of methylation interference
analysis suggest that the N7 of the EcoRI site dG and the N3 of the central dA,
d(7GpAp7ApTpTpC), are utilized as contacts by the enzyme.  The failure to
observe interference upon methylation of the 5'-penultimate dA within the
sequence implies that the endonuclease does not bond to the N3 of this residue,
despite the fact that it is protected against alkylation by the protein.
Ethylation interference patterns suggest four major phosphate contacts between
endonuclease and each DNA strand.  Two of these phosphates are 5'-external to
the EcoRI sequence, d(7pN7pG7pApA7pTpTpC), suggesting involvement of outside
phosphates in electrostatic interactions.  Moreover, alkylation protection and
interference effects on the two DNA strands display perfect 2-fold symmetry.
Thus, the endonuclease interacts with a minimum of 10 nucleotide pairs to yield
a DNA-protein complex characterized by elements of symmetry.  In contrast,
specific alkylation effects were not observed in comparable experiments with
the endonuclease and a DNA which had been previously methylated by the EcoRI
modification enzyme.

<>

<1>Lu, A.L., Cheng, S.C., Jack, W.E., Modrich, P.
<2>DNA contacts and mechanism of specific site location of EcoRI endonuclease, and effects of modification methylation on DNA helix conformation.
<3>Fed. Proc.
<4>41
<5>1199
<6>1982
<7>Alkylation interference and protection methods have been utilized to deduce
potential DNA contacts involved in specific complex formation between EcoRI
endonuclease and its recognition sequence.  The endonuclease interacts
symmetrically with a minimum of 10 nucleotide pairs, including nitrogens in
both major and minor grooves and phosphate groups along the DNA backbone.
Dissociation and association rates for site-specific EcoRI endonuclease-DNA
complexes increase with increasing DNA chain length up to a plateau.  Since
intrinsic EcoRI site Keq are identical for the different length DNA fragments,
outside DNA sequences must form the basis for observed kinetic differences.
Thus, outside DNA sequences lie on the major kinetic pathway by which EcoRI
endonuclease locates and leaves its recognition site.  Conformational changes
associated with DNA biological methylation was investigated (using the
electrophoresis band shift method).  The DNA helix was found to be unwound
about 0.5 degree for each methyl group introduced to the N6 position of
adenine.  No detectable effect (< 0.2 degree) was observed when methyl group
was introduced to DNA on C5 of cytosine.

<>

<1>Lu, H., Graham, D., Xiao, S.D., Gutierrez, O., Yamaoka, Y.
<2>Relationship between Helicobacter pylori restriction endonuclease-replacing gene, Hrga and clinical outcome.
<3>Gastroenterology
<4>124
<5>A270
<6>2003
<7>Background: Recently, polymorphisms in the hpyIII locus in Helicobacter pylori have been
identified and the presence of a new gene, hrgA, that had replaced hpyIIIR, was reported to be
related to gastric cancer in Asian strains.  However, this relation was data-derived and the
hypothesis has not yet been confirmed.  Aims: To confirm the relationship between hrgA allelic
type and clinical outcome in strains from both Asian and Western countries.  Methods: We
examined H. pylori strains from Asia (Korea, Hong Kong, Thailand, Taiwan, and Vietnam) and
Western countries (Columbia, Canada, France, and Italy).  hrgA and hpyIIIR status were
determined by polymerase chain reaction.  Results: There were 332 strains including as 172
Asian (49 cancer, 123 non-cancer) and 160 Western strains (50 gastric cancer, 90 non-cancer).
The prevalence of hrgA gene was significantly higher in Western strains 46.8% than Asian
strains (33.7%) irrespective of the clinical outcome (P = 0.01).  In Asian, the prevalence of
hrgA gene was 34.78% in strains from cancer and 33.3% from non-cancer.  In western countries,
the prevalence of hrgA gene was 50% in strains from cancer and 45% from non-cancer.
Discussion: We were unable to confirm the hypothesis that the hrgA gene was related to gastric
cancer.  We found no relationship between the prevalence of the hrgA gene and clinical outcome
in either Asian or Western countries.  The prevalence of hrgA gene was higher in Western
strains compared with Asian strains which are the opposite of other important putative
virulence factors such as cag pathogenicity island, vacA s1 genotype, babA2 genotypes and oipA
"on" types.

<>

<1>Lu, J.J., Wang, J.F., Hu, X.F.
<2>Genome Sequence of Growth-Improving Paenibacillus mucilaginosus Strain KNP414.
<3>Genome Announcements
<4>1
<5>e00881-13
<6>2013
<7>Paenibacillus mucilaginosus is a critical growth-improving silicate bacterium. Here, we report
the complete genome sequence of P. mucilaginosus strain KNP414.
This information will provide us with the opportunity to understand its molecular
mechanisms and develop more effective utilization of the strain.

<>

<1>Lu, L., Patel, H., Bissler, J.J.
<2>Optimizing DpnI digestion conditions to detect replicated DNA.
<3>Biotechniques
<4>33
<5>316-318
<6>2002
<7>In vitro replication systems are powerful tools for studying DNA replication and repair.
Replication competent cellular extracts can be used with reporter plasmids that contain the
simian virus 40 (SV40) DNA replication origin.  Replication is initiated by the addition of
the large T antigen.  By using plasmid DNA prepared in bacteria expressing adenosine methylase
as substrate, processive eukaryotic replication involving polymerase alpha, beta and epsilon
can be distinguished from gap-filling reactions mediated by other DNA polymerases such as
polymerase beta by the methylation status of the resulting DNA (Figure 1).  The bacterially
derived plasmid will contain methyladenosine at the DpnI restriction site, GATC, making the
plasmid susceptible to DpnI cleavage.  The complete absence of methylation profoundly
suppresses DNA cleavage by DpnI.  However, after a single round of semi-conservative
replication, the DNA is only hemimethylated.  The susceptibility of hemimethylated DNA has not
been well studied, though previous work suggests that replicated DNA also may be susceptible
to cleavage.  We demonstrate that under conditions of enzyme excess, long duration digestion,
or small DNA amounts, the hemimethylated DNA is susceptible to DpnI cleavage.  Other
investigators have identified that DpnI digestions can be difficult to interpret. For the
analysis of DNA replication, we optimized the digestion conditions so that only fully
methylated DNA would be digested, preserving hemimethylated DNA for analyses.

<>

<1>Lu, L., Zhou, S.Y., Chen, C., Weng, S.P., Chan, S.M., He, J.G.
<2>Complete genome sequence analysis of an iridovirus isolated from the orange-spotted grouper, Epinephelus coioides.
<3>Virology
<4>339
<5>81-100
<6>2005
<7>Orange-spotted grouper iridovirus (OSGIV) was the causative agent of
serious systemic diseases with high mortality in the cultured
orange-spotted grouper, Epinephelus coioides. Here we report the complete
genome sequence of OSGIV. The OSGIV genome consists of 112,636 bp with a
G+C content of 54%. 121 putative open reading frames (ORF) were identified
with coding capacities for polypeptides varying from 40 to 1168 amino
acids. The majority of OSGIV shared homologies to other iridovirus genes.
Phylogenetic analysis of the major capsid protein, ATPase, cytosine DNA
methyl transferase and DNA polymerase indicated that OSGIV was closely
related to infectious spleen and kidney necrosis virus (ISKNV) and rock
bream iridovirus (RBIV), but differed from lymphocytisvirus and ranavirus.
The determination of the genome of OSGIV will facilitate a better
understanding of the molecular mechanism underlying the pathogenesis of
the OSGIV and may provide useful information to develop diagnosis method
and strategies to control outbreak of OSGIV.

<>

<1>Lu, S., Zhang, X., Zhu, Y., Kim, K.S., Yang, J., Jin, Q.
<2>Complete Genome Sequence of the Neonatal-Meningitis-Associated Escherichia coli Strain CE10.
<3>J. Bacteriol.
<4>193
<5>7005
<6>2011
<7>Neonatal bacterial meningitis continues to be an important cause of mortality and morbidity
worldwide. Escherichia coli possessing the K1
capsular polysaccharide is the most common Gram-negative pathogen causing
neonatal meningitis. Here we present the complete genome sequence of
neonatal meningitis-associated E. coli strain CE10, a unique K1 strain
with a functional type III secretion system. Functional analysis of the
genome should enhance our knowledge of the pathogenesis of neonatal E.
coli K1 meningitis.

<>

<1>Lu, W., Wise, M.J., Tay, C.Y., Windsor, H.M., Marshall, B.J., Peacock, C., Perkins, T.
<2>Comparative analysis of the full genome of the Helicobacter pylori isolate, Sahul64, identifies genes of high divergence.
<3>J. Bacteriol.
<4>196
<5>1073-1083
<6>2013
<7>Isolates of Helicobacter pylori can be classified phylogeographically. High genetic diversity
and rapid microevolution are a hallmark of H. pylori genomes, a phenomenon that is proposed to
play a functional role in persistence and colonisation of diverse human populations. To
provide further genomic evidence in the lineage of H. pylori and to further characterise
diverse strains of this pathogen in different human populations, we report the finished genome
sequence of Sahul64, an H. pylori strain isolated from an Indigenous Australian. Our analysis
identified highly divergent genes, when compared to the 38 publically available genomes, which
include genes involved in the biosynthesis and modification of lipopolysaccharide, putative
prophage genes, restriction modification components and hypothetical genes. Furthermore, the
virulence associated vacA locus is a pseudogene and the cagPAI is not present. However, the
genome does contain a gene cluster associated with pathogenicity including dupA. Our analysis
found that with the addition of Sahul64 to the 38 genomes, the core genome content of H.
pylori is reduced by approximately 14% ( approximately 170 genes) and the pan-genome has
expanded from 2070 to 2238 genes. We have identified three putative horizontally acquired
regions, including one that is likely to have been acquired prior to speciation from the
closely related Helicobacter cetorum. Our results suggest that Sahul64, with the absence of
cagPAI, highly divergent cell envelope proteins and a predicted non-transportable VacA, could
be more highly adapted to ancient indigenous Australian people but with lower virulence
potential compared to other sequenced and cagPAI positive H. pylori strains.

<>

<1>Lu, W.D., Pang, H.Q., Guo, L.Z.
<2>Genome Sequence of the Fructan-Degrading Organism Marinimicrobium sp. Strain LS-A18, Isolated from a Marine Solar Saltern.
<3>Genome Announcements
<4>1
<5>e00776-13
<6>2013
<7>Marinimicrobium sp. strain LS-A18 is a fructan-degrading organism isolated from a brine sample
from a marine solar saltern in Jiaozhou Bay, China. The draft genome
sequence of this bacterium is 3,815,107 bp in length, with a G+C content of
59.03%. To our knowledge, this is the first genome announcement of a
fructan-degrading strain of the genus Marinimicrobium.

<>

<1>Lu, Z., Li, Y., Que, Q., Kutish, G.F., Rock, D.L., Van Etten, J.L.
<2>Analysis of 94 kb of the chlorella virus PBCV-1 330-kb genome: Map positions 88 to 182.
<3>Virology
<4>216
<5>102-123
<6>1996
<7>Analysis of 94 kb of DNA, located between map positions 88 and 182 kb in the 330-kb chlorella
virus PBCV-1 genome, revealed 195 open reading frames 65 codons or longer.  One hundred and
five of the 195 ORFs were considered major ORFs.  Twenty-six of the 105 major ORFs resembled
genes in the databases including three chitinases, a chitosanase, three serine/threonine
protein kinases, two additional protein kinases, a tyrosine protein phosphatase, two ankyrins,
an ornithine decarboxylase, a copper/zinc-superoxide dismutase, a proliferating cell nuclear
antigen, a DNA polymerase, a fibronectin-binding protein, the yeast Ski2 protein, an adenine
DNA methyltransferase and its corresponding DNA site-specific endonuclease, and an amidase.
The genes for the 105 major ORFs were evenly distributed along the genome and, except for one
noncoding 1788-nucleotide stretch, the genes were close together.  Unexpectedly, a 900-bp
region in the 1788-bp noncoding sequence resembled a CpG island.

<>

<1>Lu, Z., Lu, Y.
<2>Complete Genome Sequence of a Thermophilic Methanogen, Methanocella conradii HZ254, Isolated from Chinese Rice Field Soil.
<3>J. Bacteriol.
<4>194
<5>2398-2399
<6>2012
<7>Members of the order Methanocellales play a key role in methane emissions in paddy fields.
Because of their slow growth and fastidious culture conditions,
pure cultures are difficult to isolate and have been unavailable until recently.
Here we report the complete genome sequence of a novel isolate in this group,
Methanocella conradii strain HZ254.

<>

<1>Luan, X., Cui, Z., Gao, W., Li, Q., Yin, X., Zheng, L.
<2>Genome Sequence of the Petroleum Hydrocarbon-Degrading Bacterium Alcanivorax sp.  Strain 97CO-5.
<3>Genome Announcements
<4>2
<5>e01277-14
<6>2014
<7>Alcanivorax sp. strain 97CO-5 was isolated from a crude-oil-degrading consortium, enriched
from Yellow Sea sediment of China. Here, we present the draft genome of
strain 97CO-5, which comprises 3,251,558 bp with a G+C content of 54.54% and
contains 2,962 protein-coding genes and 42 tRNAs.

<>

<1>Lubys, A., Janulaitis, A.
<2>Cloning and analysis of the plasmid-borne genes encoding the Bsp6I restriction and modification enzymes.
<3>Gene
<4>157
<5>25-29
<6>1995
<7>The Bsp6I restriction and modification (R-M) system has been localized on the plasmid pXH13,
naturally occurring in the Bacillus sp. strain RFL6.  The genes coding for the Bsp6I-R-M
system, an Fnu4HI isoschizomer recognizing the sequence GCNGC, have been cloned in Escherichia
coli by two steps.  The nucleotide sequence of a 2126-bp region containing the genes for
restriction endonuclease (ENase; bsp6IR) and DNA methyltransferase (MTase; bsp6IM) has been
determined.  The genes are separated by 99 bp and are arranged tandemly with bsp6IR preceding
bsp6IM.  The DNA sequence predicts an ENase of 174 amino acids (19.9 kDa) and a MTase of 315
aa (36.3 kDa).  M.Bsp6I contains all the conserved aa sequence motifs characteristic for
m5C-MTases.  In addition, its variable region exhibits a slight similarity to the
5'-GCNGC-3'-specific target-recognition domain from M.Phi3T.  No aa sequence similarity was
found between R.Bsp6I and M.Bsp6I, nor among R.Bsp6I and other known ENases.  We have tested
recombinant plasmids carrying the complete R-M system for their ability to transform native
and pre-methylated Escherichia coli hosts.  The results indicate that pre-methylation
increases the efficiency of establishment of the complete R-M system.  In addition, we have
obtained orientation-dependent differences in transformation efficiency.

<>

<1>Lubys, A., Jurenaite, S., Janulaitis, A.
<2>Structural organization and regulation of the plasmid-borne type II restriction-modification system Kpn21 from Klebsiella pneumoniae RFL2.
<3>Nucleic Acids Res.
<4>27
<5>4228-4234
<6>1999
<7>Kpn2I enzymes of a type II restriction-modification (R-M) system from the bacterium Klebsiella
pneumoniae strain RFL2 recognize the sequence 5'-TCCGGA-3'. The Kpn2I R-M genes have been
cloned and expressed in Escherichia coli. DNA sequence analysis revealed the presence of two
convergently transcribed open reading frames (ORFs) coding for a restriction endonuclease
(Enase) of 301 amino acids (34.8 kDa) and methyltransferase (Mtase) of 375 amino acids (42.1
kDa). The 3'-terminal ends of these genes ( kpn2IR and kpn2IM, respectively) overlap by 11
bp. In addition, a small ORF (gene kpn2IC ) capable of coding for a protein of 96 amino acids
in length (10.6 kDa) was found upstream of kpn2IM. The direction of kpn2IC transcription is
opposite to that of kpn2IM. The predicted amino acid sequence of this ORF includes a probable
helix-turn-helix motif. We show that the product of kpn2IC represses expression of the Kpn2I
Mtase but has no influence on expression of the Enase gene. Such a mode of regulation is
unique among R-M systems analyzed so far. The Kpn2I R-M is located on the K.pneumoniae RFL2
plasmid pKp4.3, which is able to replicate in E.coli cells.

<>

<1>Lubys, A., Lubiene, J.
<2>Cloning of the genes encoding type IIS restriction-modification system HphI from Haemophilus parahaemolyticus.
<3>Biologija
<4>0
<5>39-41
<6>1996
<7>The restriction-modification system HphI from Haemophilus parahaemolyticus was cloned into E.
coli and sequenced.  Five open reading frames aligned in the same orientation were found.  The
first ORF, orfX, is incomplete and encodes an unknown protein.  The next is the gene for
C5-methylcytosine forming HphI methylase (hphIMC).  The third gene codes for the
N6-methyladenine forming HphI methylase (hphIMA).  The fourth gene encodes HphI restriction
endonuclease.  The expression of hphIR was obtained only after the subcloning of this gene
under the control of T7 RNA polymerase promoter.  The last gene, menB, codes for the
dihydroxynaphthoate synthase.

<>

<1>Lubys, A., Lubiene, J., Kulakauskas, S., Stankevicius, K., Timinskas, A., Janulaitis, A.
<2>Cloning and analysis of the genes encoding the type IIS restriction-modification system HphI from Haemophilus parahaemolyticus.
<3>Nucleic Acids Res.
<4>24
<5>2760-2766
<6>1996
<7>The genomic region encoding the type IIS restriction-modification (R-M) system HphI (enzymes
recognizing the asymmetric sequence 5'-GGTGA-3'/5'-TCACC-3') from Haemophilus
parahaemolyticus were cloned into Escherichia coli and sequenced.  Sequence analysis of the
R-M HphI system revealed three adjacent genes aligned in the same orientation: a cytosine 5
methyltransferase (gene hphIMC), an adenine N6 methyltransferase (hphIMA) and the HphI
restriction endonuclease (gene hphIR).  Either methyltransferase is capable of protecting
plasmid DNA in vivo against the action of the cognate restriction endonuclease.  hphIMA
methylation renders plasmid DNA resistant to R.HindIII at overlapping sites, suggesting that
the adenine methyltransferase modifies the 3'-terminal A residue on the GGTGA strand.  Strong
homology was found between the N-terminal part of the m6A methyltransferase and an
unidentified reading frame interrupted by an incomplete galE gene of Neisseria meningitidis.
The HphI R-M genes are flanked by a copy of a 56 bp direct nucleotide repeat on each side.
Similar sequences have also been identified in the non-coding regions of H.influenzae Rd DNA.
Possible involvement of the repeat sequences in the mobility of the HphI R-M system is
discussed.

<>

<1>Lubys, A., Menkevicius, S., Timinskas, A., Butkus, V., Janulaitis, A.
<2>Cloning and analysis of translational control for genes encoding the Cfr9I restriction-modification system.
<3>Gene
<4>141
<5>85-89
<6>1994
<7>The complete type-II Cfr9I restriction-modification (R-M) system of Citrobacter freundii
strain RFL9, recognizing the DNA sequence CCCGGG, has been cloned and expressed, and
functionally active enzymes have been produced in Escherichia coli. Both the methyltransferase
(MTase; M.Cfr9I) and restriction endonuclease (ENase; R.Cfr9I) were found to be encoded on a
2.3-kb cloned fragment in the same transcriptional orientation, but differing in translational
phases. The last codon (underlined) (ATGA) of the MTase-encoding gene (Cf9IM) overlaps with
the start codon for the ENase-encoding gene (overlined) Cfr9IR). A nucleotide sequence
complementary to a predicted Shine-Dalgarno sequence preceeding cfr9IR is within this gene.
Predicted free energy (deltaG) for formation of the mRNA secondary structure involving these
complementary sequences was found to be - 16.1 kcal mol. Amino-acid sequence homology of 80%
was found between R.Cfr9I and R.XcyI.

<>

<1>Lubys, A., Vitkute, J., Lubiene, J., Janulaitis, A.
<2>Methylation-specific restriction endonucleases and their applications.
<3>US Patent Office
<4>US 20110207139 A
<5>
<6>2011
<7>Provided is a methylation-specific restriction endonuclease for a DNA duplex substrate, which
endonuclease recognizes in a strand of the duplex a 2 to 6 nucleotide recognition sequence
comprising a 5-methylcytosine, and cleaves each strand of the duplex at a fixed position
outside the recognition sequence.

<>

<1>Lubys, A., Vitkute, J., Lubiene, J., Janulaitis, A.
<2>Restriction endonucleases and their applications.
<3>European Patent Office
<4>EP 2365067 A
<5>
<6>2011
<7>Provided is a methylation-specific restriction endonuclease for a DNA duplex substrate, which
endonuclease recognizes in a strand of the duplex a 2 to 6 nucleotide recognition sequence
comprising a 5-methylcytosine, and cleaves each strand of the duplex at a fixed position
outside the recognition sequence.

<>

<1>Lucas, P., Otis, C., Mercier, J.-P., Turmel, M., Lemieux, C.
<2>Rapid evolution of the DNA-binding site in LAGLIDADG homing endonucleases.
<3>Nucleic Acids Res.
<4>29
<5>960-969
<6>2001
<7>Sequence analysis of chloroplast and mitochondrial large subunit rRNA genes from over 75 green
algae disclosed 28 new group I intron-encoded proteins carrying a single LAGLIDADG motif.
These putative homing endonucleases form four subfamilies of homologous enzymes, with the
members of each subfamily being encoded by introns sharing the same insertion site. We showed
that four divergent endonucleases from the I-CreI subfamily cleave the same DNA substrates.
Mapping of the 66 amino acids that are conserved among the members of this subfamily on the
3-dimensional structure of I-CreI bound to its recognition sequence revealed that these
residues participate in protein folding, homodimerization, DNA recognition and catalysis.
Surprisingly, only seven of the 21 I-CreI amino acids interacting with DNA are conserved,
suggesting that I-CreI and its homologs use different subsets of residues to recognize the
same DNA sequence. Our sequence comparison of all 45 single-LAGLIDADG proteins identified so
far suggests that these proteins share related structures and that there is a weak pressure in
each subfamily to maintain identical protein-DNA contacts. The high sequence variability we
observed in the DNA-binding site of homologous LAGLIDADG endonucleases provides insight into
how these proteins evolve new DNA specificity.

<>

<1>Lucas, S. et al.
<2>Complete Genome Sequence of the Thermophilic, Piezophilic, Heterotrophic Bacterium Marinitoga piezophila KA3.
<3>J. Bacteriol.
<4>194
<5>5974-5975
<6>2012
<7>Marinitoga piezophila KA3 is a thermophilic, anaerobic, chemoorganotrophic, sulfur-reducing
bacterium isolated from the Grandbonum deep-sea hydrothermal vent
site at the East Pacific Rise (13 degrees N, 2,630-m depth). The genome of M.
piezophila KA3 comprises a 2,231,407-bp circular chromosome and a 13,386-bp
circular plasmid. This genome was sequenced within Department of Energy Joint
Genome Institute CSP 2010.

<>

<1>Lucas-Elio, P., Goodwin, L., Woyke, T., Pitluck, S., Nolan, M., Kyrpides, N.C., Detter, J.C., Copeland, A., Lu, M., Bruce, D., Detter, C., Tapia, R., Han, S., Land, M.L., Ivanova, N., Mikhailova, N., Johnston, A.W., Sanchez-Amat, A.
<2>Complete genome sequence of Marinomonas posidonica type strain (IVIA-Po-181(T)).
<3>Standards in Genomic Sciences
<4>7
<5>31-43
<6>2012
<7>IVIA-Po-181 Lucas-Elio 2011 belongs to the family within the phylum . Different species of the
genus can be readily isolated from the seagrass . is among the
most abundant species of the genus detected in the cultured microbiota of
suggesting a close relationship with this plant, which has a great ecological
value in the Mediterranean Sea, covering an estimated surface of 38,000 Km. Here
we describe the genomic features of . The 3,899,940 bp long genome harbors 3,544
protein-coding genes and 107 RNA genes and is a part of the project.

<>

<1>Lucas-Elio, P., Goodwin, L., Woyke, T., Pitluck, S., Nolan, M., Kyrpides, N.C., Detter, J.C., Copeland, A., Teshima, H., Bruce, D., Detter, C., Tapia, R., Han, S., Land, M.L., Ivanova, N., Mikhailova, N., Johnston, A.W., Sanchez-Amat, A.
<2>Complete genome sequence of the melanogenic marine bacterium Marinomonas mediterranea type strain (MMB-1(T)).
<3>Standards in Genomic Sciences
<4>6
<5>63-73
<6>2012
<7>Marinomonas mediterranea MMB-1(T) Solano & Sanchez-Amat 1999 belongs to the family
Oceanospirillaceae within the phylum Proteobacteria. This species is of
interest because it is the only species described in the genus Marinomonas to
date that can synthesize melanin pigments, which is mediated by the activity of a
tyrosinase. M. mediterranea expresses other oxidases of biotechnological
interest, such as a multicopper oxidase with laccase activity and a novel
L-lysine-epsilon-oxidase. The 4,684,316 bp long genome harbors 4,228
protein-coding genes and 98 RNA genes and is a part of the Genomic Encyclopedia
of Bacteria and Archaea project.

<>

<1>Lucas-Elio, P., Goodwin, L., Woyke, T., Pitluck, S., Nolan, M., Kyrpides, N.C., Detter, J.C., Copeland, A., Teshima, H., Bruce, D., Detter, C., Tapia, R., Han, S., Land, M.L., Ivanova, N., Mikhailova, N., Johnston, A.W.B., Sanchez-Amat, A.
<2>Complete genome sequence of the melanogenic marine bacterium Marinomonas mediterranea type strain (MMB-1T).
<3>Standards in Genomic Sciences
<4>5
<5>63-73
<6>2012
<7>
<>

<1>Lucchini, S., Sidoti, J., Brussow, H.
<2>Broad-range bacteriophage resistance in Streptococcus thermophilus by insertional mutagenesis.
<3>Virology
<4>275
<5>267-277
<6>2000
<7>Streptococcus thermophilus is a lactic acid bacterium used in industrial milk fermentation. To
obtain phage-resistant starters, S. thermophilus strain Sfi1 was submitted to mutagenesis with
the thermolabile insertional vector pG(+)host9:ISS1 followed by a challenge with the lytic S.
thermophilus phage Sfi19.  Vector insertions into four distinct sites led to a
phage-resistance phenotype. Three mutants were characterized further. They were protected
against the homologous challenging phage and 14 heterologous phages. All three mutants
adsorbed phages. No intracellular phage DNA synthesis was observed in mutants R7 and R71,
while mutant R24 showed a delayed and diminished phage DNA synthesis compared to the parental
Sfi1 strain. In mutant R7 a short deletion occurred next to the insertion site which removed
the upstream sequences and the 15 initial codons from orf 394, encoding a likely transmembrane
protein.  Analogy with other phage systems suggests an involvement of this protein in the
phage DNA injection process. In mutant R24 the vector was inserted into orf 269 predicting an
oxido-reductase. When the vector sequence was removed via homologous recombination across the
duplicated insertion elements, mutant R24 returned to the phage susceptibility of the parental
strain. This observation suggested that inactivation of orf  269 was not crucial for the
resistance phenotype. A gene encoding a likely restriction subunit of a type I
restriction-modification system was located directly downstream of the insertion site in
mutant R24. HsdM and hsdS genes encoding the modification and specificity subunits of a type I
R-M system and biological evidence for an active R-M system were detected in strain Sfi1,
suggesting involvement of a type I R-M system in the resistance phenotype of R24.

<>

<1>Lucid, A., Bullman, S., Koziel, M., Corcoran, G.D., Cotter, P.D., Sleator, R.D., Lucey, B.
<2>Draft Genome Sequence of Campylobacter ureolyticus Strain CIT007, the First Whole-Genome Sequence of a Clinical Isolate.
<3>Genome Announcements
<4>2
<5>e00262-14
<6>2014
<7>Herein, we present the draft genome sequence of Campylobacter ureolyticus. Strain CIT007 was
isolated from a stool sample from an elderly female presenting with diarrheal illness and
end-stage chronic renal disease.

<>

<1>Lucker, S., Wagner, M., Maixner, F., Pelletier, E., Koch, H., Vacherie, B., Rattei, T., Damste, J.S., Spieck, E., Le Paslier, D., Daims, H.
<2>A Nitrospira metagenome illuminates the physiology and evolution of globally important nitrite-oxidizing bacteria.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>13479-13484
<6>2010
<7>Nitrospira are barely studied and mostly uncultured nitrite-oxidizing bacteria, which are,
according to molecular data, among the most diverse
and widespread nitrifiers in natural ecosystems and biological wastewater
treatment. Here, environmental genomics was used to reconstruct the
complete genome of 'Candidatus Nitrospira defluvii' from an activated
sludge enrichment culture. On the basis of this first-deciphered
Nitrospira genome and of experimental data, we show that Ca. N. defluvii
differs dramatically from other known nitrite oxidizers in the key enzyme
nitrite oxidoreductase (NXR), in the composition of the respiratory chain,
and in the pathway used for autotrophic carbon fixation, suggesting
multiple independent evolution of chemolithoautotrophic nitrite oxidation.
Adaptations of Ca. N. defluvii to substrate-limited conditions include an
unusual periplasmic NXR, which is constitutively expressed, and pathways
for the transport, oxidation, and assimilation of simple organic compounds
that allow a mixotrophic lifestyle. The reverse tricarboxylic acid cycle
as the pathway for CO(2) fixation and the lack of most classical defense
mechanisms against oxidative stress suggest that Nitrospira evolved from
microaerophilic or even anaerobic ancestors. Unexpectedly, comparative
genomic analyses indicate functionally significant lateral gene-transfer
events between the genus Nitrospira and anaerobic ammonium-oxidizing
planctomycetes, which share highly similar forms of NXR and other proteins
reflecting that two key processes of the nitrogen cycle are evolutionarily
connected.

<>

<1>Luckwu-de-Lucena, B.T., Silva, G.G., Manoel-Dos-Santos, B., Dias, G.M., Amaral, G.R., Moreira, A.P., de Morais, J.M.A., Dutilh, B.E., Edwards, R.A., Balbino, V., Thompson, C.C., Thompson, F.L.
<2>Genome Sequences of the Ethanol-Tolerant Lactobacillus vini Strains LMG 23202T and JP7.8.9.
<3>J. Bacteriol.
<4>194
<5>3018
<6>2012
<7>We report on the genome sequences of Lactobacillus vini type strain LMG 23202(T)(DSM
20605)isolated from fermenting grape musts in Spain) and the industrial strain L. vini JP7.8.9
(isolated from a bioethanol plant in northeast Brazil.  All contigs were assembled using
gsAssembler, and genes were predicted and annotated using Rapid Annotation using Subsystem
Technology (RAST). The identified genome sequence of LMG 23202(T) had 2.201.333 bp, 37.6% G+C,
and 1,833 genes, whereas the identified genome sequence of JP7.8.9 had 2.301.037 bp, 37.8%
G+C, and 1,739 genes. The gene repertoire of the species L. vini offers promising
opportunities for biotechnological applications.

<>

<1>Lugli, G.A., Duranti, S., Milani, C., Turroni, F., Viappiani, A., Mangifesta, M., van Sinderen, D., Ventura, M.
<2>The Genome Sequence of Bifidobacterium moukalabense DSM 27321 Highlights the Close Phylogenetic Relatedness with the Bifidobacterium dentium Taxon.
<3>Genome Announcements
<4>2
<5>e00048-14
<6>2014
<7>Bifidobacterium moukalabense DSM 27321 is the reference strain for a recently described new
bifidobacterial species that has been isolated from a wild west
lowland gorilla. Here, we report the whole-genome sequence of DSM 27321, which
supports very close phylogenetic relatedness with members of the Bifidobacterium
adolescentis phylogenetic group and, in particular, the Bifidobacterium dentium
taxon.

<>

<1>Luhtanen, A.M., Eronen-Rasimus, E., Kaartokallio, H., Rintala, J.M., Autio, R., Roine, E.
<2>Isolation and characterization of phage-host systems from the Baltic Sea ice.
<3>Extremophiles
<4>18
<5>121-130
<6>2014
<7>In search for sea ice bacteria and their phages from the Baltic Sea ice, two ice
samples were collected from land-fast ice in a south-west Finland coastal site in
February and March 2011. Bacteria were isolated from the melted sea ice samples
and phages were screened from the same samples for 43 purified isolates.
Plaque-producing phages were found for 15 bacterial isolates at 3 degrees C. Ten
phage isolates were successfully plaque purified and eight of them were chosen
for particle purification to analyze their morphology and structural proteins.
Phage 1/32 infecting an isolate affiliated to phylum Bacteroidetes
(Flavobacterium sp.) is a siphovirus and six phages infecting isolates affiliated
to gamma-Proteobacteria (Shewanella sp.) hosts were myoviruses. Cross titrations
between the hosts showed that all studied phages are host specific. Phage
solutions, host growth and phage infection were tested in different temperatures
revealing phage temperature tolerance up to 45 degrees C, whereas phage infection
was in most of the cases retarded above 15 degrees C. This study is the first to
report isolation and cultivation of ice bacteria and cold-active phages from the
Baltic Sea ice.

<>

<1>Lui, A.C.P., McBride, B.C., Vovis, G.F., Smith, M.
<2>Site specific endonuclease from Fusobacterium nucleatum.
<3>Nucleic Acids Res.
<4>6
<5>1-15
<6>1979
<7>Four different isolates of Fusobacterium nucleatum (A,C,D and E) contain
restriction endonucleases of differing specificity.  Whilst many of the
endonucleases are isochizomers of known enzymes, two novel activities are Fnu
DII which recognizes and cleaves the sequence 5'-CG^CG-3' 3'-GC^GC-5' and Fnu
EI which recognizes and cleaves the sequence 5'-^GATC-3' 3'-CTAG^-5'
irrespective of the extent of methylation of the adenine residues.

<>

<1>Lukacs, C.M., Aggarwal, A.K.
<2>BglII and MunI: what a difference a base makes.
<3>Curr. Opin. Struct. Biol.
<4>11
<5>14-18
<6>2001
<7>Restriction endonucleases are resilient to alterations in their DNA-binding specificities.
Structures of the BglII and MunI endonucleases bound to their palindromic DNA sites, which
differ by only their outer base pairs from the recognition sequences of BamHI and EcoRI,
respectively, have recently been determined. A comparison of these complexes reveals
surprising differences and similarities in structure, and provides a basis for understanding
the immutability of restriction endonucleases.

<>

<1>Lukacs, C.M., Kucera, R., Schildkraut, I., Aggarwal, A.K.
<2>Understanding the immutability of restriction enzymes: crystal structure of BglII and its DNA substrate at 1.5A resolution.
<3>Nat. Struct. Biol.
<4>7
<5>134-140
<6>2000
<7>Restriction endonucleases are remarkably resilient to alterations in their DNA binding
specificity. To understand the basis of this immutability, we have determined the crystal
structure of endonuclease BglII bound to its recognition sequence (AGATCT), at 1. 5 A
resolution. We compare the structure of BglII to endonuclease BamHI, which recognizes a
closely related DNA site (GGATCC). We show that both enzymes share a similar alpha/beta core,
but in BglII, the core is augmented by a beta-sandwich domain that encircles the DNA to
provide extra specificity.  Remarkably, the DNA is contorted differently in the two
structures, leading to different protein-DNA contacts for even the common base pairs.
Furthermore, the BglII active site contains  a glutamine in place of the glutamate at the
general base position in BamHI, and only a single metal is found coordinated to the putative
nucleophilic water and the phosphate oxygens. This surprising diversity in structures shows
that different strategies can be successful in achieving site-specific recognition and
catalysis in restriction endonucleases.

<>

<1>Luke, P.A., Halford, S.E.
<2>Solubility of the EcoRI restriction endonuclease and its purification from an over-producing strain.
<3>Gene
<4>37
<5>241-246
<6>1985
<7>The solubility of the EcoRI restriction endonuclease was measured in solutions
of varying NaCl concentrations, at different temperatures and in the presence
of DNA.  The precipitation of the protein was enhanced by low NaCl
concentrations, by elevated temperatures, and by the addition of DNA.  These
observations are discussed in relationship to the interaction of this protein
with DNA.  The purification of the EcoRI restriction enzyme from a strain of
Escherichia coli that over-produces this enzyme was hampered by the
insolubility of the protein, and hence the purification procedure was modified
to optimize the recovery of active enzyme.

<>

<1>Luke, P.A., Halford, S.E.
<2>Fluorescent labelling of EcoRI and EcoRV.
<3>Biochem. Soc. Trans.
<4>14
<5>259-260
<6>1986
<7>Fluorescent chromophores, covalently attached to proteins, have been widely used as reporter
groups to monitor associations of proteins with ligands, either by observing perturbations to
fluorescence spectra or by fluorescence polarization methods.  In the latter, given excitation
with light polarized in the vertical plane and measurements of emission in both horizontal and
vertical planes, the binding of a ligand that increases the rotational correlation time of the
protein (relative to the excited state lifetime of the fluorophore) may reduce the H/V ratio.

<>

<1>Luke, P.A., McCallum, S.A., Halford, S.E.
<2>The EcoRV restriction endonuclease.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>185-207
<6>1987
<7>Type II restriction endonucleases have attracted attention for two main
reasons:  firstly, their many applications in the dissection of DNA and in the
construction of novel DNA molecules; secondly, as systems for studying the
interactions of proteins with specific DNA sequences.  With respect to the
latter, the EcoRI restriction endonuclease has been examined in greater depth
than any other type II enzyme.  However, the EcoRI enzyme has a major
disadvantage as a system for studying DNA-protein interactions:  the protein
has a remarkably low solubility.  The solutions in which EcoRI shows maximal
activity, and also affinity for its recognition site, are saturated at less
than 0.5 microM of this protein.  Consequently, many techniques that have been
developed to study protein-ligand interactions but which require high
concentrations of the protein in solution, such as NMR spectroscopy, cannot be
used on EcoRI.  But this drawback does not apply to all type II restriction
enzymes.  A different enzyme, the EcoRV restriction endonuclease, has special
advantages as a system for studying DNA-protein interactions.  In particular,
this is the only type II restriction enzyme (apart from EcoRI) for which
crystals of the protein have been reported.

<>

<1>Lukinavicius, G., Klimasauskas, S., Dalhoff, C., Weinhold, E.
<2>Conversion of DNA methyltransferases to alkyltransferases via cofactor engineering.
<3>FEBS Lett.
<4>273
<5>336-337
<6>2006
<7>S-Adenosyl-L-methionine (AdoMet) is a biological sulfonium compound known as the major methyl
donor in the cell.  AdoMet-dependent methyltransferases (MTases) catalyze the transfer of the
methyl group from the cofactor S-adenosyl-L-methionine (AdoMet) to defined positions within
various substrates like DNA, RNA, proteins and other biomolecules.  A series of new AdoMet
analogs with extended linear carbon chains replacing the methyl group was obtained by chemical
synthesis.  Remarkably, we find that extended groups containing a double or triple
carbon-carbon bond one unit away from the sulfonium center, was opposed to saturated carbon
chains, are readily transferred onto DNA owing to conjugative stabilization of the SN2-like
transition state.  The MTase-assisted transalkylations of DNA are truly catalytic, efficient
and proceed in a sequence-specific manner yielding corresponding adenine-N-6, cytosine-N4 or
cytosine-5 derivatives.

<>

<1>Lukinavicius, G., Lapiene, V., Stasevskij, Z., Dalhoff, C., Weinhold, E., Klimasauskas, S.
<2>Targeted labeling of DNA by methyltransferase-directed transfer of activated groups (mTAG).
<3>J. Am. Chem. Soc.
<4>129
<5>2758-2759
<6>2007
<7>Methyltransferases catalyze highly specific transfers of methyl groups from the ubiquitous
cofactor S-adenosyl-L-methionine (AdoMet) to
various biopolymers like DNA, RNA, and proteins. Here we describe the
first synthetic analogue of AdoMet with an activated side chain
carrying a primary amino group that permits efficient
methyltransferase-directed functionalization of DNA and subsequent
amine-specific chemoligations with various reporter groups. The
demonstrated two-step sequence-specific labeling of natural DNA offers
a facile way to query the methylation status of the target sites and
envisions numerous applications in functional studies and medical
diagnostics.

<>

<1>Lukinavicius, G., Lapinaite, A., Urbanaviciute, G., Gerasimaite, R., Klimasauskas, S.
<2>Engineering the DNA cytosine-5 methyltransferase reaction for sequence-specific labeling of DNA.
<3>Nucleic Acids Res.
<4>40
<5>11594-11602
<6>2012
<7>DNA methyltransferases catalyse the transfer of a methyl group from the ubiquitous cofactor
S-adenosyl-L-methionine (AdoMet) onto specific target sites
on DNA and play important roles in organisms from bacteria to humans. AdoMet
analogs with extended propargylic side chains have been chemically produced for
methyltransferase-directed transfer of activated groups (mTAG) onto DNA, although
the efficiency of reactions with synthetic analogs remained low. We performed
steric engineering of the cofactor pocket in a model DNA cytosine-5
methyltransferase (C5-MTase), M.HhaI, by systematic replacement of three
non-essential positions, located in two conserved sequence motifs and in a
variable region, with smaller residues. We found that double and triple
replacements lead to a substantial improvement of the transalkylation activity,
which manifests itself in a mild increase of cofactor binding affinity and a
larger increase of the rate of alkyl transfer. These effects are accompanied with
reduction of both the stability of the product DNA-M.HhaI-AdoHcy complex and the
rate of methylation, permitting competitive mTAG labeling in the presence of
AdoMet. Analogous replacements of two conserved residues in M.HpaII and M2.Eco31I
also resulted in improved transalkylation activity attesting a general
applicability of the homology-guided engineering to the C5-MTase family and
expanding the repertoire of sequence-specific tools for covalent in vitro and ex
vivo labeling of DNA.

<>

<1>Lukinavicius, G., Tomkuviene, M., Masevicius, V., Klimasauskas, S.
<2>Enhanced Chemical Stability of AdoMet Analogues for Improved Methyltransferase-Directed Labeling of DNA.
<3>ACS Chem. Biol.
<4>8
<5>1134-1139
<6>2013
<7>Methyltransferases catalyze specific transfers of methyl groups from the ubiquitous cofactor
S-adenosyl-L-methionine (AdoMet) to various
nucleophilic positions in biopolymers like DNA, RNA, and proteins. We
had previously described synthesis and application of AdoMet analogues
carrying sulfonium-bound 4-substituted but-2-ynyl side chains for
transfer by methyltransferases. Although useful in certain
applications, these cofactor analogues exhibited short lifetimes in
physiological buffers. Examination of the reaction kinetics and
products showed that their fast inactivation followed a different
pathway than observed for AdoMet and rather involved a pH-dependent
addition of a water molecule to the side chain. This side reaction was
eradicated by synthesis of a series of cofactor analogues in which the
separation between an electronegative group and the triple bond was
increased from one to three carbon units. The designed hex-2-ynyl
moiety-based cofactor analogues with terminal amino, azide, or alkyne
groups showed a markedly improved enzymatic transalkylation activity
and proved well suitable for methyltransferase-directed
sequence-specific labeling of DNA in vitro and in bacterial cell
lysates.

<>

<1>Lukyanchuk, V., Reva, O., Polishchuk, L.
<2>Restriction endonucleases of type II from two endophytic strains of Bacillus.
<3>Biochem. Soc. Trans.
<4>28
<5>A181
<6>2000
<7>Restriction modification systems were found in more than 3000 bacterial species.  It is
generally accepted that their primary role is to protect host bacteria against phage
infection.  We used a simple technique proposed by Belavin with our modification, for
detection of bacterial restriction endonucleases.  Endonucleases activity of 24 Bacillus
strains isolated from inner tissues of cotton plants was tested by this method.  Enzyme of
Bacillus licheniformis IMB B-55O8B (Bli55O8B) cuts DNA of lambda phage at 2 sites and DNA of
T7 phage at 29 sites.  We found that computer calculated numbers of DNAs fragments and their
molecular sizes that would be generated at the sequence GGTCTC, correlated with the observed
cleavage frequency of the both mentioned substrates.  We suppose a new isoschisomer of type II
endonuclease of restriction Eco31I was isolated from Bacillus licheniformis IMB B-55O8B.  A
restriction endonuclease of Bacillus subtilis IMB 5044B (Bsu5044B) cut DNA at M13mp18 at four
sites, DNA of pUC19 at six sites and DNA of pBR322 at fifteen sites.  We found that computer
calculated number of fragments that would be generated the sequence 5'-GGNCC-3' correlated
with the observed cleavage frequency of the above mentioned substrates.  Recently this
sequence was determined for restrictase Asu1.  Similarity of digestion profiles of gel
electrophoresis of T7 and lambda DNA fragments obtained after treating by Cfr13I (isoschizomer
of AsuI) and Bsu5O44B was evidence of Bsu5044B could be new isoschizomer of AsuI.  Thus two
endophytic strains of Bacillus were found to produce endonucleases of restriction.  Found
enzymes were isoschizomers of well known enzymes AsuI and EcoR31I.

<>

<1>Lukyanchuk, V.V., Polishchuk, L.V., Strizhkova, G.M., Kopieiko, O.P., Matselyukh, B.P.
<2>Determination of site-specificity of the II type restriction endonuclease in Streptomyces sp. 48.
<3>Mikrobiol. Zh.
<4>61
<5>80-83
<6>1999
<7>The activity of a restriction endonuclease which splits DNA of lambda and T7 phages into 8
fragments has been detected in the mycelium lysates of the 2-days culture of Streptomyces sp.
48.  The fragments dimensions according to the data of agarose gel electrophoresis, coincide
with those of AsuII fragments of DNA of lambda and T7 phages.  This allows the studied Ssp48
restrictase to be considered the isoschizomer of AsuII restriction endonuclease.

<>

<1>Lukyanchuk, V.V., Reva, O.M., Polishchuk, L.V.
<2>Type-II site-specific restriction endonucleases in endophytic strains of the genus Bacillus.
<3>Mikrobiol. Zh.
<4>61
<5>28-30
<6>1999
<7>Using a modified Belavin's method the site-specific II type restriction endonucleases were
determined for two endophytic strains of the Bacillus genus from the inner tissues of cotton
plants.

<>

<1>Lukyanchuk, V.V., Reva, O.N., Polishchuk, L.V.
<2>Endophytic bacilli producing type II restriction endonucleases.
<3>Mikrobiologiia
<4>71
<5>491-493
<6>2002
<7>Two of thirteen bacillar strains isolated from the inner tissues of cotton plants were found
to produce type II restriction endonucleases. The investigation of the site specificity of
these enzymes showed that they are AsuI and Eco31I isoschizomers.

<>

<1>Lukyanchuk, V.V., Strizhkova, G.M., Polishchuk, L.V., Kopeyko, O.P., Matselyukh, B.P.
<2>Search for site-specific endonucleases of the II type restriction among Streptomycetes.
<3>Mikrobiol. Zh.
<4>60
<5>33-35
<6>1998
<7>Three Streptomycetes strains producing endonucleases of restriction of II type have been found
using the modified method of Belavin.

<>

<1>Lukyanchuk, V.V., Suchanoff, S.M., Polishchuk, L.V., Matselyukh, B.P.
<2>Isolation of site-specific endonuclease of Streptomyces sp. 48.
<3>Biopol. Kletka
<4>16
<5>559-561
<6>2000
<7>A method of isolation and purification of site-specific endonuclease of the II type S. sp. 48
has been worked out.  It has been determined that the S. sp. 48 lysate contains only one
enzyme with site-specific activity.  Maximum elution of the endonuclease was made by buffer
with ionic strength of 350 mM NaCl.  This method provides sufficient purification and
concentration of this enzyme with preservation of its specific activity.

<>

<1>Lulitanond, A., Ito, T., Li, S., Han, X., Ma, X.X., Engchanil, C., Chanawong, A., Wilailuckana, C., Jiwakanon, N., Hiramatsu, K.
<2>ST9 MRSA strains carrying a variant of type IX SCCmec identified in the Thai community.
<3>BMC Infect. Dis.
<4>13
<5>214
<6>2013
<7>BACKGROUND: Infections caused by methicillin-resistant Staphylococcus aureus
(MRSA) in Thailand occur most frequently in healthcare facilities. However,
reports of community-associated MRSA are limited. METHODS: We characterized 14
MRSA isolates from outpatients (O-1 to O-14) by phenotypic and genotypic methods
and compared them with 5 isolates from inpatients (I-1 to I-5). Thai MRSA
isolates from a healthcare worker (N-1) and a pig (P-1) were also included as ST9
MRSA strains from other sources. RESULTS: All MRSA isolates from the outpatients
and inpatients were multidrug-resistant (resistant to >/=3 classes of
antimicrobials). All of them except strains O-2 and I-3 carried type III SCCmec
and belonged to agrI, coagulase IV, spa type t037 or t233, which related to
ST239. The strain O-2 (JCSC6690) carried type IX SCCmec and belonged to agrII,
coagulaseXIc, spa type t337 and ST9, whereas the strain I-3 carried a type III
SCCmec and belonged to ST1429. Nucleotide sequence determination revealed that
the type IX SCCmec element in strain O-2 was distinct from that in a Thai ST398
strain (JCSC6943) previously identified in 2011; nucleotide identities of ccrA
and ccrB were 93 and 91%, respectively and several open reading frames (ORFs) at
the joining regions were different. PCR experiments suggested that strain O-2 and
N-1 carried similar SCCmec element, whereas that of strain P-1 was different,
suggesting that distinct ST9-MRSA-IX clones might be spreading in this province.
CONCLUSIONS: The SCCmecIX-ST9 MRSA clones of distinct SCCmec subtypes might have
emerged in the Thai community and might also have disseminated into the hospital.

<>

<1>Lundberg, U., Meinke, A., Nagy, E., von Gabain, A., Noiges, B., Gelbmann, D., Poljak, A., Triska, C.
<2>Borrelia antigens.
<3>European Patent Office
<4>EP 2287176 A
<5>
<6>2011
<7>The present invention relates to isolated nucleic acid molecules which encode a protein,
isolated nucleic acid molecules which encode a hyperimmune serum reactive antigen, a vector
which comprises such nucleic acid molecule, a host cell comprising such vector, a hyperimmune
reactive antigen from Borrelia species, proteins which are preferably hyperimmune serum
reactive antigens, hyperimmune serum reactive antigens, antigens, a process for producing such
proteins, hyperimmune serum reactive antigens or antigens, a process for producing a cell
which expresses such proteins, hyperimmune serum reactive antigens or antigens, a process for
producing a cell which expresses such protein, hyperimmune serum reactive antigen or antigen,
an antibody binding to such protein, hyperimmune serum reactive antigen or antigen, a
hybridoma cell producing such antibody, methods for producing such antibody, a pharmaceutical
composition comprising such nucleic acid molecule, protein, hyperimmune serum reactive
antigen, antigen or antibody, the use of such nucleic acid molecule, protein, hyperimmune
serum reactive antigen, antigen or antibody for the manufacture of a medicament, methods for
identifying an antagonist capable of reducing or inhibiting the interaction activity of such
protein, hyperimmune serum reactive antigen or antigen, methods for diagnosing an infection
and methods for the treatment of an infection.  More specifically such proteins, hyperimmune
serum reactive antigens or antigens are produced by or associated with bacterial pathogens
causing Lyme disease or bacterial infections caused by Borrelia burgdorferi s.1.

<>

<1>Lundberg, U., Meinke, A., Nagy, E., von Gabain, A., Noiges, B., Gelbmann, D., Poljak, A., Triska, C.
<2>Borrelia antigens.
<3>European Patent Office
<4>EP 2289906 A
<5>
<6>2011
<7>The present invention relates to isolated nucleic acid molecules which encode a protein,
isolated nucleic acid molecules which encode a hyperimmune serum reactive antigen, a vector
which comprises such nucleic acid molecule, a host cell comprising such vector, a hyperimmune
reactive antigen from Borrelia species, proteins which are preferably hyperimmune serum
reactive antigens, hyperimmune serum reactive antigens, antigens, a process for producing such
proteins, hyperimmune serum reactive antigens or antigens, a process for producing a cell
which expresses such protein, hyperimmune serum reactive antigen or antigen, an antibody
binding to such protein, hyperimmune serum reactive antigen or antigen, a hybridoma cell
producing such antibody, methods for producing such antibody, a pharmceutical composition
comprising such nucleic acid molecule, protein, hyperimmune serum reactive antigen, antigen or
antibody, the use of such nucleic acid molecule, protein, hyperimmune serum ractive antigen,
antigen or antibody for the manufacture of a medicament, methods for identifying an antagonist
capable of reducing or inhibiting the interaction activity of such protein, hyperimmune serum
reactive antigen or antigen, methods for diagnosing an infection and methods for the treatment
of an infection.  More specifically such proteins, hyperimmune serum reactive antigens or
antigens are produced by or associated with bacterial pathogens causing Lyme disease or
bacterial infections caused by Borrelia burgdorferi s.1.

<>

<1>Lundin, A., Bjorkholm, B., Kupershmidt, I., Unemo, M., Nilsson, P., Andersson, D.I., Engstrand, L.
<2>Slow genetic divergence of Helicobacter pylori strains during long-term colonization.
<3>Infect. Immun.
<4>73
<5>4818-4822
<6>2005
<7>The genetic variability of Helicobacter pylori is known to be high compared to that of many
other bacterial species. H. pylori is adapted to
the human stomach, where it persists for decades, and adaptation to each
host results in every individual harboring a distinctive bacterial
population. Although clonal variants may exist within such a population,
all isolates are generally genetically related and thus derived from a
common ancestor. We sought to determine the rate of genetic change of H.
pylori over 9 years in two asymptomatic adult patients. Arbitrary primed
PCR confirmed the relatedness of individual subclones within a patient.
Furthermore, sequencing of 10 loci ( approximately 6,000 bp) in three
subclones per time and patient revealed only two base pair changes among
the subclones from patient I. All sequences were identical among the
patient II subclones. However, PCR amplification of the highly divergent
gene amiA revealed great variation in the size of the gene between the
subclones within each patient. Thus, both patients harbored a single
strain with clonal variants at both times. We also studied genetic changes
in culture- and mouse-passaged strains, and under both conditions no
genetic divergence was found. These results suggest that previous
estimates of the rate of genetic change in H. pylori within an individual
might be overestimates.

<>

<1>Lunnen, K., Davis, T., Wilson, G.
<2>Method for cloning and expression of Streptomyces SbfI restriction endonuclease and SbfI methylase in E. coli.
<3>European Patent Office
<4>EP 1516927 A
<5>
<6>2005
<7>The present invention relates to: recombinant DNA encoding the SbfI restriction endonuclease
as well as the SbfI methylase, and expression of the SbfI restriction endonuclease and SbfI
methylase in E. coli cells containing the recombinant DNA; and methods for cloning the SbfI
restriction gene (sbfIR) from Streptomyces species Bf-61 into E. coli by PCR.  The method
relied on primers based on DNA sequences predicted from amino acid sequences of the purified
SbfI restriction endonuclease.

<>

<1>Lunnen, K., Greci, J., Wilson, G.
<2>Sequence and expression of acc65I restriction endonuclease and acc65I methylase from Acinetobacter.
<3>US Patent Office
<4>US 20080009036 A
<5>
<6>2008
<7>An isolated DNA is provided which encodes a protein that is capable of binding to
5'GGTACC-3', the isolated DNA being capable of hybridizing to SEQ ID NO:3 under stringent
hybridization conditions.  The isolated DNA may be alternatively characterized as coding for a
protein having an amino acid sequence comprising SEQ ID NO:5 or by an amino acid sequence with
an expectation value of less than E=e-02 in a BLAST search using SEQ ID NO:5.  Vectors
containing the isolated DNA and host cells expressing the vectors as well as a method for
making recombinant Acc65I having the above properties are also provided.

<>

<1>Lunnen, K., Greci, J., Wilson, G.
<2>Method for cloning and expression of Acc65I restriction endonuclease and Acc65I methylase in E. Coli.
<3>US Patent Office
<4>US 07795408
<5>
<6>2010
<7>An isolated DNA is provided which encodes a protein that is capable of binding to 5 '
GGTACC-3 ', the isolated DNA being capable of
hybridizing to SEQ ID NO:3 under stringent hybridization conditions.
The isolated DNA may be alternatively characterized as coding for a
protein having an amino acid sequence comprising SEQ ID NO:5 or by an
amino acid sequence with an expectation value of less than E=e(-02 )in
a BLAST search using SEQ ID NO:5. Vectors containing the isolated DNA
and host cells expressing the vectors as well as a method for making
recombinant Acc65I having the above properties are also provided.

<>

<1>Lunnen, K., Greci, J., Wilson, G.
<2>Method for cloning and expression of Acc65I restriction endonuclease and Acc65I methylase in E. Coli.
<3>US Patent Office
<4>US 7795408 B
<5>
<6>2010
<7>An isolated DNA is provided which encodes a protein that is capable of binding to
5'GGTACC-3', the isolated DNA being capable of hybridizing to SEQ ID No: 3 under stringent
hybridization conditions.  The isolated DNA may be alternatively characterized as coding for a
protein having an amino acid sequence comprising SEQ ID NO: 5 or by an amino acid sequence
with an expectation value fo less than E=e-02 in  a BLAST search using SEQ ID NO: 5.  Vectors
containing the isolated DNA and host cells expressing the vectors as well as a method for
making recombinant Acc65I having the above properties are also provided.

<>

<1>Lunnen, K., Wilson, G.
<2>Method for producing the HgiAI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0321271 B
<5>
<6>1994
<7>The present invention relates to clones for the HgiAI restriction endonuclease and
modification methylase, and to the production of these enzymes from the clones.

<>

<1>Lunnen, K.D., Barsomian, J.M., Camp, R.R., Card, C.O., Chen, S.-Z., Croft, R., Looney, M.C., Meda, M.M., Moran, L.S., Nwankwo, D.O., Slatko, B.E., Van Cott, E.M., Wilson, G.G.
<2>Cloning Type II restriction and modification genes.
<3>Gene
<4>74
<5>25-32
<6>1988
<7>We have cloned into Escherichia coli the genes for 38 type-II bacterial
modification methyltransferases.  The clones were isolated by selecting in
vitro for protectively modified recombinants.  Most of the clones modify their
DNA fully but a substantial number modify only partially.  In approximately
one-half of the clones, the genes for the corresponding endonucleases are also
present.  Some of these clones restrict infecting phages and others do not.
Clones carrying endonuclease genes but lacking methyltransferase genes have
been found, in several instances, to be viable.

<>

<1>Lunnen, K.D., Dalton, M.A., Wilson, G.G., Xu, S.-Y.
<2>Method for cloning and producing the AvaI restriction endonuclease in E. coli and purification of the recombinant AvaI restriction endonuclease.
<3>US Patent Office
<4>US 6004793
<5>
<6>1999
<7>A recombinant restriction endonuclease from Anabaena variabilis is provided, as well as the
isolated gene which encodes it and methods for the production of the recombinant AvaI
restriction endonuclease.  An isolated gene encoding a modification methylase from A.
variabilis is also provided.

<>

<1>Lunnen, K.D., Davis, T., Wilson, G.G.
<2>Method for cloning and expression of SbfI restriction endonuclease and SbfI methylase in E. coli.
<3>US Patent Office
<4>US 20050064433 A
<5>
<6>2005
<7>The present invention relates to: recombinant DNA encoding the SbfI restriction endonuclease
as well as the SbfI methylase, and expression of the SbfI restriction endonuclease and SbfI
methylase in E. coli cells containing the recombinant DNA; and methods for cloning the SbfI
restriction gene (sbfIR) from Streptomyces species Bf-61 into E. coli by PCR.  The method
relied on primers based on DNA sequences predicted from amino acid sequences of the purified
SbfI restriction endonuclease.

<>

<1>Lunnen, K.D., Davis, T., Wilson, G.G.
<2>Method for cloning and expression of SbfI restriction endonuclease  and SbfI methylase in E. coli.
<3>US Patent Office
<4>US 6958230 B
<5>
<6>2005
<7>The present invention relates to: recombinant DNA encoding the
SbfI restriction endonuclease as well as the SbfI methylase, and expression
of the SbfI restriction endonuclease and SbfI methylase in E. coli cells
containing the recombinant DNA; and methods for cloning the SbfI
restriction gene (sbfIR) from Streptomyces species Bf-61 into E. coli by
PCR.  The method relied on primers based on DNA sequences predicted from
amino acid sequences of the purified SbfI restriction endonuclease.

<>

<1>Lunnen, K.D., Kong, H., Wilson, G.G.
<2>Method for cloning and producing the SnaBI restriction endonuclease and purification of the recombinant SnaBI restriction endonuclease.
<3>US Patent Office
<4>US 6025179
<5>
<6>2000
<7>The methylase selection method was used to clone the SnaBI methylase gene from Sphaerotilus
natans (ATCC 15291).  An active SnaBI methylase was cloned in E. coli using pSNaBI-2, a pUC19
derivative containing two SnaBI sites.  Because methylase and restriction genes are usually
located alongside each other in restriction-modification systems, efforts were made to clone
the downstream DNA by inverse PCR.  Inverse PCR cloned the missing portion of the SnaBI
endonuclease and also identified a control, or C, protein.  The two open reading frames snaBIR
(ORF1) and snaBIC (ORF2) converged toward the SnaBI methylase gene (ORF).  Attempts to
establish a snaBIR-recombinant plasmid expressing the SnaBI endonuclease in E. coli modified
with SnaBI methylase failed.  Overexpression of the SnaBI endonuclease in E. coli required the
use of the heterospecific BsaAI methylase.

<>

<1>Lunnen, K.D., Kong, H., Wilson, G.G.
<2>Method for cloning and producing the SnaBI restriction endonuclease and purification thereof.
<3>European Patent Office
<4>EP 0985730 A
<5>
<6>2000
<7>The methylase selection method was used to clone the SnaBI methylase gene (snaBIM) from
Sphaerotilus natans (ATCC 15291).  An active SnaBI methylase was cloned in E. coli using
pSnaBI-2, a pUC19 derivative containing two SnaBI sites.  Because methylase and restriction
genes are usually located alongside each other in restriction-modification systems, efforts
were made to clone the downstream DNA by inverse PCR.  The missing portion of the SnaBI
endonuclease was cloned by inverse PCR.  A control, or C, protein was identified using the
same technique.  The two open reading frames snaBIR (ORF1) and snaBIC (ORF2) were orientated
towards the SnaBI methylase gene (ORF).  Attempts to establish a snaBIR-recombinant plasmid
expressing the SnaBI endonuclease in E. coli modified with SnaBI methylase failed.
Overexpression of the SnaBI endonuclease in E. coli required the use of the heterospecific
BsaAI methylase.

<>

<1>Lunnen, K.D., Kong, H., Wilson, G.G.
<2>Method for cloning and producing the SnaBI restriction endonuclease and purification thereof.
<3>European Patent Office
<4>EP 0985730 B
<5>
<6>2006
<7>
<>

<1>Lunnen, K.D., Kong, H., Wilson, G.G.
<2>Cloning and production of SnaBI restricted endonuclease and method for the purification of recombinant SnaBI restricted endonuclease.
<3>Japanese Patent Office
<4>JP 2000083686 A
<5>
<6>2000
<7>The methylase selection method was used to clone the SnaBI methylase gene (snaBIM) from
Sphaerotilus natans (ATCC15291).  An active SnaBI methylase was cloned in E. coli using
pSnaBI-2, a pUC19 derivative containing two SnaBI sites.  Because methylase and restriction
genes are usually located alongside each other in a restriction-modification system, efforts
were made to clone the downstream DNA by inverse PCR.  Inverse PCR cloned the missing portion
of the SnaBI endonuclease and also identified a control, or C, protein.  The two open reading
frames snaBIR (ORF1) and snaBIC (ORF2) converged toward the snaBI methylase gene (ORF).
Attempts to establish a snaBIR-recombinant plasmid expressing the SnaBI endonulcease in E.
coli modified with SnaBI methylase failed.  Overexpression of the SnaBI endonulocease in E.
coli required the use of the heterospecific BsaAI methylase.

<>

<1>Lunnen, K.D., Meixsell, T., Morgan, R.D., Wilson, G.G.
<2>Method for cloning and producing the RsaI restriction endonuclease in E. coli.
<3>European Patent Office
<4>EP 1164189 A
<5>
<6>2001
<7>RsaI, a restriction enzyme from the bacterium Rhodopseudomonas sphaeroides, recognizes the DNA
sequence 5'-GTAC-3'.  Because RsaI is commercially valuable, we sought to overproduce it by
cloning the genes for RsaI and its accompanying, modification, enzyme.  The
'methylase-selection' method, the customary procedure for cloning restriction and
modification genes, was applied to RsaI.  The method yielded clones containing the methylase
gene (rsaIM), but none containing both the methylase gene and the restriction gene (rsaIR).
Inverse-PCR was then used to recover sections of the DNA downstream of rsaIM.  These sections
were sequenced, and the sequences were joined in silico to reveal the gene organization of the
RsaI R-M system.  By comparing the coding potential of the DNA with the N-terminal amino acid
sequence of the purified RsaI restriction enzyme, we discovered that the RsaI R and M genes,
rather than being adjacent-the situation that pertains in most R-M systems-are separated by an
intervening gene of unknown function.  Based on this information, the rsaIR gene was cloned by
PCR instead of methylase-selection.  These new clones provided to be highly unstable, however,
even in the presence of the rsaM gene.  Various attempts were made to stabilize the gene, but
most met with failure.  Stability was finally achieved by introducing a second methylase gene,
mjaVM, to augment the protection provided by rsaIM, and by tightly controlling the expression
of rsaIR using a special two-promoter, anti-sense transcription, expression vector.

<>

<1>Lunnen, K.D., Morgan, R.D., Meixsell, T., Wilson, G.G.
<2>Method for cloning and producing the RsaI restriction endonuclease in E. coli and purification of the recombinant RsaI restriction endonuclease.
<3>US Patent Office
<4>US 6210945 B
<5>
<6>2001
<7>RsaI, a restriction enzyme from the bacterium Rhodopseudomonas sphaeroides, recognizes the DNA
sequence 5'-GTAC-3'.  Because RsaI is commercially valuable, we sought to overproduce it by
cloning the genes for RsaI and its accompanying, modification enzyme.  The
'methylase-selection' method, the customary procedure for cloning restriction and
modification genes, was applied to RsaI.  The method yielded clones containing the methylase
gene (rsaIM), but none containing both the methylase gene and the restriction gene (rsaIR).
Inverse-PCR was then used to recover sections of the DNA downstream of rsaIM.  These sections
were sequenced, and the sequences were joined in silico to reveal the gene organization of the
RsaI R-M system.  By comparing the coding potential of the DNA with the N-terminal amino acid
sequence of the purified RsaI restriction enzyme, we discovered that the RsaI R and M genes,
rather than being adjacent-the situation that pertains in most R-M systems-are separated by an
intervening gene of unknown function.  Based on this information, the rsaIR gene was cloned by
PCR instead of methylase-selection.  These new clones proved to be highly unstable, however,
even in the presence of the rsaIM gene.  Various attempts were made to stabilize the gene, but
most met with failure.  Stability was finally achieved by introducing a second methylase gene,
mjaVM, to augment the protection provided by rsaIM, and by tightly controlling the expression
of rsaIR using a special two-promoter, anti-sense transcription, expression vector.

<>

<1>Lunnen, K.D., Morgan, R.D., Meixsell, T., Wilson, J.G.
<2>Process for cloning and producing RsaI-restricted endonuclease in Escherichia coli.
<3>Japanese Patent Office
<4>JP 2002306181 A
<5>
<6>2002
<7>
<>

<1>Lunnen, K.D., Morgan, R.D., Timan, C.J., Krzycki, J.A., Reeve, J.N., Wilson, G.G.
<2>Characterization and cloning of MwoI (GCN7GC), a new type-II restriction-modification system from Methanobacterium wolfei.
<3>Gene
<4>77
<5>11-19
<6>1989
<7>R.MwoI, a type-II restriction enzyme with the new specificity 5'-GCN7GC-3', was
found in extracts of the thermophilic archaebacterium, Methanobacterium wolfei.
R.MwoI cleaves duplex DNA producing fragments with 3-nt, 3'-terminal
extensions, thus: GCN5/N2GC.  The genes coding for the MwoI restriction and
modification enzymes were cloned into Escherichia coli on the plasmid vector
pBR322.  The clones synthesize a low level of R.MwoI endonuclease.  The
plasmids display incomplete MwoI-specific modification, suggesting that the
clones synthesize a low level of the M.MwoI methyltransferases, too.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the MwoI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5053330
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the MwoI restriction
endonuclease by 1) introducing the restriction endonuclease gene from M. wolfei into a host
whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid
encoding and expressing the MwoI restriction endonuclease activity, and 3) purifying the MwoI
restriction endonuclease from the fermented host which contains the plasmids encoding and
expressing the MwoI restriction endonuclease activity.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the XmaI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5002882
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the XmaI restriction
endonuclease (1) introducing the restriction endonuclease gene from X. malvacearum into a host
whereby the restriction gene is expressed; (2) fermenting the host which contains the plasmid
encoding and expressing the XmaI restriction endonuclease activity, and (3) purifying the XmaI
restriction endonuclease from the fermented host which contains the plasmid encoding and
expressing the XmaI restriction endonuclease activity.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the HgiAI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 4987074
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the HgiAI restriction
endonuclease by (1) introducing the restriction endonuclease gene from Herpetosiphon giganteus
into a host whereby the restriction gene is expressed; (2) fermenting the host which contains
the vector encoding and expressing the HgiAI restriction endonuclease, and (3) purifying the
HgiAI restriction endonuclease from the fermented host which contains the vector encoding and
expressing the HgiAI restriction endonuclease activity.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the AflII restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5030569
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the AflII restriction
endonuclease by 1) introducing the restriction endonuclease gene from Anabaena flos-aquae into
a host whereby the restriction gene is expressed; 2) fermenting the host which contains the
plasmid encoding and expressing the AflII restriction endonuclease activity, and 3) purifying
the AflII restriction endonuclease from the fermented host which contains the plasmid encoding
and expressing the AflII restriction endonuclease activity.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the XmaI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0395367 B
<5>
<6>1998
<7>The present invention is directed to a method for cloning and producing the XmaI restriction
endonuclease by 1) introducing the restriction endonuclease gene from X. malvacaerum into a
host whereby the restriction gene is expressed; 2) fermenting the host which contains the
plasmid encoding and expressing the XmaI restriction endonuclease activity, and 3) purifying
the XmaI restriction endonuclease from the fermented host which contains the plasmid encoding
and expressing the XmaI restriction endonuclease activity.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the MwoI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0388136 B
<5>
<6>1995
<7>The present invention relates to clones for the MwoI restriction endonuclease and modification
methylase, and the production of these enzymes from the clones.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the BglI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5366882
<5>
<6>1994
<7>In accordance with the present invention there is provided an isolated DNA coding for the BglI
restriction endonuclease and modification methylase derived from Bacillus globigii RUB561
strain, as well as related methods for cloning said recombinant DNA. The present invention
also relates to clones which express recombinant BglI restriction endonuclease and recombinant
modification methylase produced from BglI recombinant DNA and to methods for producing said
enzymes.

<>

<1>Lunnen, K.D., Wilson, G.G.
<2>Method for producing the AflII restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0314346 B
<5>
<6>1995
<7>The present invention relates to a method for cloning DNA coding for the AflII restriction
endonuclease, and the production of the enzyme from the clone.

<>

<1>Luo, C., Walk, S.T., Gordon, D.M., Feldgarden, M., Tiedje, J.M., Konstantinidis, K.T.
<2>Genome sequencing of environmental Escherichia coli expands understanding of the ecology and speciation of the model bacterial species.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>7200-7205
<6>2011
<7>Defining bacterial species remains a challenging problem even for the model
bacterium Escherichia coli and has major practical consequences for reliable
diagnosis of infectious disease agents and regulations for transport and
possession of organisms of economic importance. E. coli traditionally is thought
to live within the gastrointestinal tract of humans and other warm-blooded
animals and not to survive for extended periods outside its host; this
understanding is the basis for its widespread use as a fecal contamination
indicator. Here, we report the genome sequences of nine environmentally adapted
strains that are phenotypically and taxonomically indistinguishable from typical
E. coli (commensal or pathogenic). We find, however, that the commensal genomes
encode for more functions that are important for fitness in the human gut, do not
exchange genetic material with their environmental counterparts, and hence do not
evolve according to the recently proposed fragmented speciation model. These
findings are consistent with a more stringent and ecologic definition for
bacterial species than the current definition and provide means to start
replacing traditional approaches of defining distinctive phenotypes for new
species with omics-based procedures. They also have important implications for
reliable diagnosis and regulation of pathogenic E. coli and for the coliform
cell-counting test.

<>

<1>Luo, J., Bruice, T.C.
<2>Low-frequency normal mode in DNA HhaI methyltransferase and motions of residues involved in the base flipping.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>16194-16198
<6>2005
<7>The results of normal-mode analyses are in accord with the proposal that a low-frequency
motion of the HhaI methyltransferase enzyme is responsible for base flipping in bound DNA. The
vectors of the low-frequency normal mode of residues Ser-85 and Ile-86 point directly to the
phosphate and ribose moieties of the DNA backbone near the target base in position to rotate
the dihedral angles and flip the base out of the DNA duplex. The vector of residue Gln-237 on
the major groove is in the proper orientation to assist base separation. Our results favor the
major groove pathway and the protein active process in base flipping.

<>

<1>Luo, W.Y., Tu, A.H.T., Cao, Z.H., Yu, H.L., Dybvig, K.
<2>Identification of an isoschizomer of the HhaI DNA methyltransferase in Mycoplasma arthritidis.
<3>FEMS Microbiol. Lett.
<4>290
<5>195-198
<6>2009
<7>The genome of Mycoplasma arthritidis strain 158 has modified cytosine residues at AGCT
sequences that render the DNA resistant to digestion
with the AluI restriction endonuclease. The DNA methyltransferase
responsible for the base modification has previously been designated
MarI. From the complete genome sequence of M. arthritidis, we identify
Marth orf138 as a candidate marI gene. Marth orf138 was cloned in
Escherichia coli and its TGA codons converted to TGG. DNA isolated from
E. coli cells expressing the modified Marth orf138 gene was degraded by
the AluI nuclease, indicating that Marth orf138 does not code for MarI.
However, the DNA from E. coli was found to have acquired resistance to
the restriction endonuclease HhaI. Genomic DNA from M. arthritidis was
also found to be resistant to HhaI (recognizes GCGC). The M.
arthritidis isoschizomer of the HhaI DNA methyltransferase, coded by
Marth orf138, is designated MarII. Transformation of M. arthritidis was
not significantly affected by modification of plasmid at HhaI sites,
indicating that the mycoplasma lacks a restriction endonuclease that
recognizes GCGC sites.

<>

<1>Luo, Y., Kong, Q., Yang, J., Golden, G., Wanda, S.Y., Jensen, R.V., Ernst, P.B., Curtiss, R.I.I.I.
<2>Complete Genome Sequence of the Universal Killer, Salmonella enterica serovar Typhimurium UK-1 (ATCC 68169).
<3>J. Bacteriol.
<4>193
<5>4035-4036
<6>2011
<7>The Salmonella enterica serovar Typhimurium strain UK-1 presents the highest invasion and
virulence attributes among the most frequently
studied strains. S. Typhimurium UK-1 has been used as the foundation for
developing recombinant vaccines and been extensively used on virulence and
colonization studies in chickens and mice. We describe here the complete
genome sequence of S. Typhimurium UK-1. Comparative genomics of Salmonella
Typhimurium will provide insight into factors that determine virulence and
invasion.

<>

<1>Luo, Y., Wang, C., Allard, S., Strain, E., Allard, M.W., Brown, E.W., Zheng, J.
<2>Draft Genome Sequences of Paenibacillus alvei A6-6i and TS-15.
<3>Genome Announcements
<4>1
<5>e00673-13
<6>2013
<7>Here, we report draft genomes of Paenibacillus alvei strains A6-6i and TS-15, which were
isolated, respectively, from plant material and soil in the Virginia
Eastern Shore (VES) tomato growing area. An array of genes related to
antimicrobial biosynthetic pathways have been identified with whole-genome
analyses of these strains.

<>

<1>Luo, Y., Xu, X., Ding, Z., Liu, Z., Zhang, B., Yan, Z., Sun, J., Hu, S., Hu, X.
<2>Complete genome of Phenylobacterium zucineum - a novel facultative intracellular bacterium isolated from human erythroleukemia cell line  K562.
<3>BMC Genomics
<4>9
<5>386
<6>2008
<7>BACKGROUND: Phenylobacterium zucineum is a recently identified facultative intracellular
species isolated from the human leukemia cell line K562.
Unlike the known intracellular pathogens, P. zucineum maintains a stable
association with its host cell without affecting the growth and morphology
of the latter. RESULTS: Here, we report the whole genome sequence of the
type strain HLK1T. The genome consists of a circular chromosome (3,996,255
bp) and a circular plasmid (382,976 bp). It encodes 3,861 putative
proteins, 42 tRNAs, and a 16S-23S-5S rRNA operon. Comparative genomic
analysis revealed that it is phylogenetically closest to Caulobacter
crescentus, a model species for cell cycle research. Notably, P. zucineum
has a gene that is strikingly similar, both structurally and functionally,
to the cell cycle master regulator CtrA of C. crescentus, and most of the
genes directly regulated by CtrA in the latter have orthologs in the
former. CONCLUSION: This work presents the first complete bacterial genome
in the genus Phenylobacterium. Comparative genomic analysis indicated that
the CtrA regulon is well conserved between C. crescentus and P. zucineum.

<>

<1>Luo, Y.R., Kang, S.G., Kim, S.J., Kim, M.R., Li, N., Lee, J.H., Kwon, K.K.
<2>Genome Sequence of Benzo(a)pyrene-Degrading Bacterium Novosphingobium pentaromativorans US6-1.
<3>J. Bacteriol.
<4>194
<5>907
<6>2012
<7>Novosphingobium pentaromativorans US6-1 showed a good ability to degrade high-molecular-weight
polycyclic aromatic hydrocarbons. We report the
draft genome sequence of strain US6-1, which contains a main chromosome
(5,096,413 bp, G+C content of 63.1%) and two plasmids (188,476 and 60,085
bp). The majority of the aromatic-hydrocarbon-degrading genes are encoded
in the larger plasmid.

<>

<1>Lupker, H.S.C., Dekker, B.M.M.
<2>Purification of the sequence-specific endonuclease SinI from Salmonella infantis.
<3>Biochim. Biophys. Acta
<4>654
<5>297-299
<6>1981
<7>A sequence-specific endonuclease was isolated from a strain of Salmonella
infantis.  Its recognition sequence proved to be identical to that of
restriction endonuclease AvaII (an endonuclease from the blue-green alga
Anabaena variabilis), G^G(A/T)CC.

<>

<1>Luria, S.E.
<2>Host-induced modifications of viruses.
<3>Cold Spring Harb. Symp. Quant. Biol.
<4>18
<5>237-244
<6>1953
<7>Viruses exhibit extensive adaptability to growth in various hosts or tissues.
It was widely held in the past that virus adaptability reflected a peculiar
plasticity of virus heredity, which allowed it to be directly influenced by its
host cells.  The alternative interpretation of virus adaptation to new host
cells as due to spontaneous mutations, which provide a range of genotypes for
the new hosts to select from, always had authoritative proponents (see Findlay,
1939).  This viewpoint finally gained wide recognition (see Burnet, 1946),
partly as a consequence of the development of phage genetics and of the
interpenetration of various branches of virology.  It is now recognized that
most variation in virus properties is caused by viral mutations, and that virus
plasticity results from the variety of genotypes present in the large viral
populations.  It was, therefore, an unexpected development when a new type of
virus variation was discovered in bacteriophages.  This has been called
host-induced or host-controlled variation (Luria and Human, 1952; Ralston and
Krueger, 1952; Anderson and Felix, 1952; Bertani and Weigle, 1953).  Its
outstanding characteristics are that is is strictly phenotypic, non-hereditary,
and that it is determined by the host cell in which a virus has been produced.
In this paper, I shall summarize the features of this phenomenon; I shall
compare it with other instances of nonhereditary phage variation; and I shall
attempt to assess its possible bearing on certain problems in other areas of
virology.

<>

<1>Luria, S.E.
<2>The recognition of DNA in Bacteria.
<3>Sci. Am.
<4>222
<5>88-102
<6>1970
<7>Some bacteria have enzyme systems that scan invading DNA molecules injected by
viruses and break them unless they are chemically marked at specific
recognition sites.

<>

<1>Luria, S.E., Human, M.L.
<2>A nonhereditary, host-induced variation of bacterial viruses.
<3>J. Bacteriol.
<4>64
<5>557-569
<6>1952
<7>One of virology's most generally valid rules is that the properties of virus particles are
unaffected by the host in which they grow.  Host adaptation and tissue adaptation, the
apparent exceptions, are explained today by selective reproduction of virus mutants in new
hosts or in new tissues.  In analyzing the relation between certain phages and certain mutants
of their bacterial hosts, we have encountered a novel situation: the genotype of the host in
which a virus reproduces affects the phenotype of the new virus.  The phenotypic change
suppresses the ability of the virus to reproduce in certain hosts but not in others.  It is a
transient change, in the sense that one cycle of growth in a suitable host returns the virus
to its original form.  Both the growth ability of the modified virus and, in some cases, its
production from normal virus are controlled in part by the physiological state of the host
cell.  The present paper describes these findings and discusses their general implications.
Other instances of host-controlled phenotypic changes in bacteriophages have recently been
discovered (Bertani and Weigle, 1952).

<>

<1>Lushnikov, A.Y., Potaman, V.N., Oussatcheva, E.A., Sinden, R.R., Lyubchenko, Y.L.
<2>DNA strand arrangement within the SfiI-DNA complex: atomic force microscopy analysis.
<3>Biochemistry
<4>45
<5>152-158
<6>2006
<7>The SfiI restriction enzyme binds to DNA as a tetramer holding two usually distant DNA
recognition sites together before cleavage of the four DNA strands. To elucidate structural
properties of the SfiI-DNA complex, atomic force microscopy (AFM) imaging of the complexes
under noncleaving conditions (Ca2+ instead of Mg2+ in the reaction buffer) was performed.
Intramolecular complexes formed by protein interaction between two binding sites in one DNA
molecule (cis interaction) as well as complexes formed by the interaction of two sites in
different molecules (trans interaction) were analyzed. Complexes were identified unambiguously
by the presence of a tall spherical blob at the DNA intersections. To characterize the path of
DNA within the complex, the angles between the DNA helices in the proximity of the complex
were systematically analyzed. All the data show clear-cut bimodal distributions centered
around peak values corresponding to 60 degrees and 120 degrees. To unambiguously distinguish
between the crossed and bent models for the DNA orientation within the complex, DNA molecules
with different arm lengths flanking the SfiI binding site were designed. The analysis of the
AFM images for complexes of this type led to the conclusion that the DNA recognition sites
within the complex are crossed. The angles of 60 degrees or 120 degrees between the DNA
helices correspond to a complex in which one of the helices is flipped with respect to the
orientation of the other. Complexes formed by five different recognition sequences
(5'-GGCCNNNNNGGCC-3'), with different central base pairs, were also analyzed. Our results
showed that complexes containing the two possible orientations of the helices were formed
almost equally. This suggests no preferential orientation of the DNA cognate site within the
complex, suggesting that the central part of the DNA binding site does not form strong
sequence specific contacts with the protein.

<>

<1>Lutsenko, E., Bhagwat, A.S.
<2>Principal causes of hot spots for cytosine to thymine mutations at sites of cytosine methylation in growing cells.  A model, its experimental support and implications.
<3>Mutat. Res.
<4>437
<5>11-20
<6>1999
<7>In Escherichia coli and human cells, many sites of cytosine methylation in DNA are hot spots
for C to T mutations. It is generally believed that T.G mismatches created by the hydrolytic
deamination of 5-methylcytosines (5meC) are intermediates in the mutagenic pathway. A number
of hypotheses have been proposed regarding the source of the mispaired thymine and how the
cells deal with the mispairs. We have constructed a genetic reversion assay that utilizes a
gene on a mini-F to compare the frequency of occurrence of C to T mutations in different
genetic backgrounds in exponentially growing E. coli. The results identify at least two causes
for the hot spot at a 5meC: (1) the higher rate of deamination of 5meC compared to C generates
more T.G than uracil.G (U.G) mismatches, and (2) inefficient repair of T.G mismatches by the
very short-patch (VSP) repair system compared to the repair of U. G mismatches by the
uracil-DNA glycosylase (Ung). This combination of increased DNA damage when the cytosines are
methylated coupled with the relative inefficiency in the post-replicative repair of T.G
mismatches can be quantitatively modeled to explain the occurrence of the hot spot at 5meC.
This model has implications for mutational hot and cold spots in all organisms.

<>

<1>Lux, T.M., Lee, R., Love, J.
<2>Complete Genome Sequence of a Free-Living Vibrio furnissii sp. nov. Strain (NCTC 11218).
<3>J. Bacteriol.
<4>193
<5>1487-1488
<6>2011
<7>The Vibrionales are widespread, free-living, Gram-negative proteobacteria that have been
linked to pathogenicity in animals and gastroenteric infection in humans.  We report the
annotated genome sequence of a free-living strain of Vibrio furnissii (NCTC 11218) harvested
from an estuarine environment.  It consists of two circular chromosomes (3.2 Mb and 1.6 Mb)
and reveals novel genes likely to be involved in pathogenicity.

<>

<1>Lv, Y., Liao, J., Wu, Z., Han, S., Lin, Y., Zheng, S.
<2>Genome Sequence of Corynebacterium glutamicum ATCC 14067, Which Provides Insight into Amino Acid Biosynthesis in Coryneform Bacteria.
<3>J. Bacteriol.
<4>194
<5>742-743
<6>2012
<7>We report the genome sequence of Corynebacterium glutamicum ATCC 14067 (once named
Brevibacterium flavum), which is useful for taxonomy research
and further molecular breeding in amino acid production. Preliminary
comparison with those of the reported coryneform strains revealed some
notable differences that might be related to the difficulties in molecular
manipulation.

<>

<1>Lv, Y., Wu, Z., Han, S., Lin, Y., Zheng, S.
<2>Genome Sequence of Corynebacterium glutamicum S9114, a Strain for Industrial Production of Glutamate.
<3>J. Bacteriol.
<4>193
<5>6096-6097
<6>2011
<7>Here we report the genome sequence of Corynebacterium glutamicum S9114, an industrial producer
widely used in production of glutamate in China.
Preliminary comparison with the sequences of the Corynebacterium
glutamicum strains ATCC 13032 and R revealed some notable mutagenesis that
might be related to the high yield of glutamate.

<>

<1>Lycksell, P.O., Graslund, A., Claesens, F., McLaughlin, L.W., Larsson, U., Rigler, R.
<2>Base pair opening dynamics of a 2-aminopurine substituted EcoRI restriction sequence and its unsubstituted counterpart in oligonucleotides.
<3>Nucleic Acids Res.
<4>15
<5>9011-9025
<6>1987
<7>Studies of 1H NMR selective recovery were performed to determine the imino
proton exchange with solvent water of the base pairs in the EcoRI endonuclease
recognition sequence GAATTC, placed at the center of self-complementary decamer
and dodecamer oligonucleotides.  In one oligonucleotide the innermost adenine
was replaced by the fluorescent base analogue 2-aminopurine (2AP).  From the
measurements at different concentrations of TRIS buffer acting as proton
exchange catalyst, base pair lifetimes were evaluated.  The results at 25C show
that the AT base pairs have lifetimes of the order of a few ms, whereas the
surrounding GC base pairs in a dodecamer have lifetimes of about 100 ms.  The
(2AP)T base pair has a shorter lifetime than the corresponding AT base pair.
The temperature dependent optical absorption, and for the 2AP containing
oligonucleotide fluorescence, were used to study the single strand-duplex
equilibrium of the decamers.  The results indicate that NMR and the optical
techniques, although applied at very different concentrations, monitor the same
conformational transition of the oligonucleotide.

<>

<1>Lykidis, A., Mavromatis, K., Ivanova, N., Anderson, I., Land, M., DiBartolo, G., Martinez, M., Lapidus, A., Lucas, S., Copeland, A., Richardson, P., Wilson, D.B., Kyrpides, N.
<2>Genome sequence and analysis of the soil cellulolytic actinomycete Thermobifida fusca YX.
<3>J. Bacteriol.
<4>189
<5>2477-2486
<6>2007
<7>Thermobifida fusca is a moderately thermophilic soil bacterium that belongs to Actinobacteria.
It is a major degrader of plant cell walls and
has been used as a model organism for the study of secreted, thermostable
cellulases. The complete genome sequence showed that T. fusca has a single
circular chromosome of 3,642,249 bp predicted to encode 3,117 proteins and
65 RNA species with a coding density of 85%. Genome analysis revealed the
existence of 29 putative glycoside hydrolases in addition to the
previously identified cellulases and xylanases. The glycosyl hydrolases
include enzymes predicted to exhibit mainly dextran/starch- and
xylan-degrading functions. T. fusca possesses two protein secretion
systems: the sec general secretion system and the twin-arginine
translocation system. Several of the secreted cellulases have sequence
signatures indicating their secretion may be mediated by the twin-arginine
translocation system. T. fusca has extensive transport systems for import
of carbohydrates coupled to transcriptional regulators controlling the
expression of the transporters and glycosylhydrolases. In addition to
providing an overview of the physiology of a soil actinomycete, this study
presents insights on the transcriptional regulation and secretion of
cellulases which may facilitate the industrial exploitation of these
systems.

<>

<1>Lykke-Andersen, J., Garrett, R.A., Kjems, J.
<2>Protein footprinting approach to mapping DNA binding sites of two archaeal homing enzymes: evidence for a two-domain protein structure.
<3>Nucleic Acids Res.
<4>24
<5>3982-3989
<6>1996
<7>The archaeal intron-encoded homing enzymes I-PorI and I-DmoI belong to a family of
endonucleases that contain two copies of a characteristic LAGLIDADG motif.  These
endonucleases cleave their intron- or intein- alleles site-specifically, and thereby
facilitate homing of the introns or inteins which encode them.  The protein structure and the
mechanism of DNA recognition of these homing enzymes is largely unknown.  Therefore, we
examined these properties of I-PorI and I-DmoI by protein footprinting.  Both proteins were
susceptible to proteolytic cleavage within regions that are equidistant from each of the two
LAGLIDADG motifs.  When complexed with their DNA substrates, a characteristic subset of the
exposed sites, located in regions immediately after and 40-60 amino acids after each of the
LAGLIDADG motifs, were protected.  Our data suggest that the enzymes are structured into two,
tandemly repeated, domains, each containing both the LAGLIDADG motif and two putative DNA
binding regions.  The latter contains a potentially novel DNA binding motif conserved in
archaeal homing enzymes.  The results are consistent with a model where the LAGLIDADG
endonucleases bind to their non-palindromic substrates as monomeric enzymes, with each of the
two domains recognizing one half of the DNA substrate.

<>

<1>Lykke-Andersen, J., Garrett, R.A., Kjems, J.
<2>Mapping metal ions at the catalytic centres of two intron-encoded endonucleases.
<3>EMBO J.
<4>16
<5>3272-3281
<6>1997
<7>Divalent metal ions play a crucial role in forming the catalytic centres of DNA endonucleases.
Substitution of Mg2+ ions by Fe2+ ions in two archaeal intron-encoded homing endonucleases,
I-DmoI and I-PorI, yielded functional enzymes and enabled the generation of reactive hydroxyl
radicals within the metal ion binding sites.  Specific hydroxyl radical-induced cleavage was
observed within, and immediately after, two conserved LAGLIDADG motifs in both proteins and at
sites at, and near, the scissile phosphates of the corresponding DNA substrates.  Titration of
Fe2+-containing protein-DNA complexes with Ca2+ ions, which are unable to support
endonucleolytic activity, was performed to distinguish between the individual metal ions in
the complex.  Mutations of single amino acids in this region impaired catalytic activity and
caused the preferential loss of a subset of hydroxyl radical cleavages in both the protein and
the DNA substrate, suggesting an active role in metal ion coordination for these amino acids.
The data indicate that the endonucleases cleave their DNA substrates as monomeric enzymes, and
contain a minimum of four divalent metal ions located at or near the catalytic centres of each
endonuclease.  The metal ions involved in cleaving the coding and the non-coding strand are
positioned immediately after the N- and C-terminally located LAGLIDADG motifs, respectively.
The dual protein/nucleic acid footprinting approach described here is generally applicable to
other protein-nucleic acid complexes when the natural metal ion can be replaced by Fe2+.

<>

<1>Lykke-Andersen, J., Thi-Ngoc, H.P., Garrett, R.A.
<2>DNA substrate specificity and cleavage kinetics of an archaeal homing-type endonuclease from Pyrobaculum organotrophum.
<3>Nucleic Acids Res.
<4>22
<5>4583-4590
<6>1994
<7>The protein encoded by intron 1 of the single 23S rRNA gene of the archaeal hyperthermophile
Pyrobaculum organotrophum was isolated and shown to constitute a homing-type DNA endonuclease,
I-PorI. It cleaves the intron- 23S rDNA of the closely related organism Pyrobaculum islandicum
near the site of intron insertion in Pb. organotrophum. In contrast, no endonuclease activity
was detected for the protein product of intron 2 of the same gene of Pb. organotrophum which,
like I-PorI, carries the LAGLI-DADG motif, common to group I intron-encoded homing enzymes.
I-PorI cleaves optimally at 80oC, with kcat and km values of about 2 min-1 and 4 nM,
respectively, and generates four nucleotide 3'-overhangs and 5'-phosphates. It can cleave a
25 base pair DNA fragment encompassing the intron insertion site. A mutation-selection study
established the base pair specificity of the endonuclease within a 17 bp region, from
positions -6 to +11 with respect to the intron-insertion site. Four of the essential base
pairs encode the sequence involved in the intron-exon interaction in the pre-rRNA that is
required for recognition by the RNA splicing enzymes. Properties of the enzyme are compared
and contrasted with those of eucaryotic homing endonucleases.

<>

<1>Lyko, F.
<2>Screening method for the identification and characterization of DNA methyltransferase inhibitors.
<3>European Patent Office
<4>EP 1384787 A
<5>11
<6>2004
<7>The present invention therefore provides a screening method for the identification and
characterization of inhibitors of DNA methyltransferases comprising the following steps: a)
generation of at least one Drosophila culture which culture comprises at least one offspring
that is derived from a cross between flies of the Drosophila strains GawB(69B) and UAS-Dnmt3a
and in an appropriate developmental stage, a sufficient amount of Drosophila medium and an
appropriate concentration of at least one candidate compound for a DNA methyltransferase
inhibitor, b) incubation of the Drosophila culture of step a) under incubation conditions
which allow normal Drosophila development for an appropriate incubation period, c)
identification of those Drosophila cultures of step b) that show the presence of mobile larvae
or pupae.  In another aspect, the invention concerns the offsprings derived from a cross
between the transgenic Drosophila strains GawB(69B) and UAS-Dnmt3a, as well as their use for
the identification and characterization of inhibitors of DNA methyltransferases, particularly
of inhibitors of Dnmt3a.

<>

<1>Lyko, F., Brown, R.
<2>DNA methyltransferase inhibitors and the development of epigenetic cancer therapies.
<3>J. Natl. Cancer Inst.
<4>97
<5>1498-1506
<6>2005
<7>Epimutations, such as the hypermethylation and epigenetic silencing of tumor suppressor genes,
play a role in the etiology of human cancers.
In contrast to DNA mutations, which are passively inherited through DNA
replication, epimutations must be actively maintained because they are
reversible. In fact, the reversibility of epimutations by
small-molecule inhibitors provides the foundation for the use of such
inhibitors in novel cancer therapy strategies. Among the compounds that
inhibit epigenetic processes, the most extensively studied are DNA
methyltransferase inhibitors. In this review, we examine the literature
on DNA methyltransferase inhibitors and discuss the efficacy of such
compounds as antitumor agents, as evaluated in phase I-III clinical
trials. We also discuss future areas of research, including the
development of nonnucleoside inhibitors, the application of novel
bioanalytical tools for DNA methylation analysis (which will be
important for the clinical application of these compounds by allowing
rational approaches to trial design), the need to optimize treatment
schedules for maximal biologic effectiveness, and the need to define
molecular endpoints so that changes induced by demethylating drugs in
patients can be monitored during treatment. Assays for genome-wide and
tumor-specific DNA methylation also need to be further developed to
establish the pharmacodynamic parameters of DNA methyltransferase
inhibitors in patients and to provide rational approaches to maximizing
the therapeutic efficacy of these compounds.

<>

<1>Lyko, F., Ramsahoye, B.H., Jaenisch, R.
<2>DNA methylation in Drosophila melanogaster.
<3>Nature
<4>408
<5>538-540
<6>2000
<7>Certain cytosine residues of eukaryotic DNA are methylated in inactive regions of the genome.
For a long time the fruitfly Drosophila was thought to be an exception, but now the evidence
points to the existence of a functional DNA-methylation system in Drosophila as well.  Here we
show that DNA is methylated, but that Drosophila genomic methylation is restriction to the
early stages of development.

<>

<1>Lyko, F., Ramsahoye, B.H., Kashevsky, H., Tudor, M., Mastrangelo, M.A., Orr-Weaver, T.L., Jaenisch, R.
<2>Mammalian (cytosine-5) methyltransferases cause genomic DNA methylation and lethality in drosophila.
<3>Nat. Genet.
<4>23
<5>363-366
<6>1999
<7>CpG methylation is essential for mouse development as well as gene regulation and genome
stability. Many features of mammalian DNA methylation are consistent with the action of a de
novo methyltransferase that establishes methylation patterns during early development and the
post-replicative maintenance of these patterns by a maintenance methyltransferase. The mouse
methyltransferase Dnmt1 (encoded by Dnmt) shows a preference for hemimethylated substrates in
vitro, making the enzyme a candidate for a maintenance methyltransferase. Dnmt1 also has de
novo methylation activity in vitro, but the significance of this finding is unclear, because
mouse embryonic stem (ES) cells contain a de novo methylating activity unrelated to Dnmt1
(ref. 10). Recently, the Dnmt3 family of methyltransferases has been identified and shown in
vitro to catalyse de novo methylation. To analyse the function of these enzymes, we expressed
Dnmt and Dnmt3a in transgenic Drosophila melanogaster. The absence of endogenous methylation
in Drosophila facilitates detection of experimentally induced methylation changes. In this
system, Dnmt3a functioned as a de novo methyltransferase, whereas Dnmt1 had no detectable de
novo methylation activity. When co-expressed, Dnmt1 and Dnmt3a cooperated to establish and
maintain methylation patterns. Genomic DNA methylation impaired the viability of transgenic
flies, suggesting that cytosine methylation has functional consequences for Drosophila
development.

<>

<1>Lyko, F., Whittaker, A.J., Orr-Weaver, T.L., Jaenisch, R.
<2>The putative Drosophila methyltransferase gene dDnmt2 is contained in a transposon-like element and is expressed specifically in ovaries.
<3>Mech. Dev.
<4>95
<5>215-217
<6>2000
<7>Several organisms, including Drosophila melanogaster, are apparently devoid of DNA
methylation. This might reflect a highly restricted activity of DNA methyltransferases, a loss
of methyltransferase activity during evolution or the dispensability of DNA methylation due to
an efficient substitute mechanism. Vestiges of a Drosophila DNA methylation system have been
identified recently. We show here that the putative DNA methyltransferase gene, dDnmt2, is the
component of a transposon-like element. This element also contains a second, novel open
reading frame with homologies to a yeast protein involved in RNA processing. Both open reading
frames are coordinately expressed and transcripts are present specifically in ovarian nurse
cells as well as during early stages of embryonic development.

<>

<1>Lynch, K.H., Stothard, P., Dennis, J.J.
<2>Genomic analysis and relatedness of P2-like phages of the Burkholderia cepacia complex.
<3>BMC Genomics
<4>11
<5>599
<6>2010
<7>Background: The Burkholderia cepacia complex (BCC) is comprised of at least seventeen
Gram-negative species that cause infections in cystic
fibrosis patients. Because BCC bacteria are broadly antibiotic
resistant, phage therapy is currently being investigated as a possible
alternative treatment for these infections. The purpose of our study
was to sequence and characterize three novel BCC-specific phages: KS5
(vB BceM-KS5 or vB BmuZ-ATCC 17616), KS14 (vB BceM-KS14) and KL3 (vB
BamM-KL3 or vB BceZ-CEP511).
Results: KS5, KS14 and KL3 are myoviruses with the A1 morphotype.
The genomes of these phages are between 32317 and 40555 base pairs in
length and are predicted to encode between 44 and 52 proteins. These
phages have over 50% of their proteins in common with enterobacteria
phage P2 and so can be classified as members of the Peduovirinae
subfamily and the 'P2-like viruses' genus. The BCC phage proteins
similar to those encoded by P2 are predominantly structural components
involved in virion morphogenesis. As prophages, KS5 and KL3 integrate
into an AMP nucleosidase gene and a threonine tRNA gene, respectively.
Unlike other P2-like viruses, the KS14 prophage is maintained as a
plasmid. The P2 E+E' translational frameshift site is conserved among
these three phages and so they are predicted to use frameshifting for
expression of two of their tail proteins. The lysBC genes of KS14 and
KL3 are similar to those of P2, but in KS5 the organization of these
genes suggests that they may have been acquired via horizontal transfer
from a phage similar to l. KS5 contains two sequence elements that are
unique among these three phages: an ISBmu2-like insertion sequence and
a reverse transcriptase gene. KL3 encodes an EcoRII-C
endonuclease/methylase pair and Vsr endonuclease that are predicted to
function during the lytic cycle to cleave non-self DNA, protect the
phage genome and repair methylation-induced mutations.
Conclusions: KS5, KS14 and KL3 are the first BCC-specific phages to
be identified as P2-like. As KS14 has previously been shown to be
active against Burkholderia cenocepacia in vivo, genomic
characterization of these phages is a crucial first step in the
development of these and similar phages for clinical use against the
BCC.

<>

<1>Lynch, K.H., Stothard, P., Dennis, J.J.
<2>Comparative analysis of two phenotypically-similar but genomically-distinct Burkholderia cenocepacia-specific bacteriophages.
<3>BMC Genomics
<4>13
<5>223
<6>2012
<7>ABSTRACT: BACKGROUND: Genomic analysis of bacteriophages infecting the
Burkholderia cepacia complex (BCC) is an important preliminary step in the
development of a phage therapy protocol for these opportunistic pathogens. The
objective of this study was to characterize KL1 (vB_BceS_KL1) and AH2
(vB_BceS_AH2), two novel Burkholderia cenocepacia-specific siphoviruses isolated
from environmental samples. RESULTS: KL1 and AH2 exhibit several unique
phenotypic similarities: they infect the same B. cenocepacia strains, they
require prolonged incubation at 30degreesC for the formation of plaques at low
titres, and they do not form plaques at similar titres following incubation at
37degreesC. However, despite these similarities, we have determined using
whole-genome pyrosequencing that these phages show minimal relatedness to one
another. The KL1 genome is 42,832 base pairs (bp) in length and is most closely
related to Pseudomonas phage 73 (PA73). In contrast, the AH2 genome is 58,065 bp
in length and is most closely related to Burkholderia phage BcepNazgul. Using
both BLASTP and HHpred analysis, we have identified and analyzed the putative
virion morphogenesis, lysis, DNA binding, and MazG proteins of these two phages.
Notably, MazG homologs identified in cyanophages have been predicted to
facilitate infection of stationary phase cells and may contribute to the unique
lysis phenotype of KL1 and AH2. CONCLUSIONS: The nearly indistinguishable
phenotypes but distinct genomes of KL1 and AH2 provide further evidence of both
vast diversity and convergent evolution in the BCC-specific phage population.

<>

<1>Lynch, K.H., Stothard, P., Dennis, J.J.
<2>Characterization of DC1, a broad-host-range Bcep22-like podovirus.
<3>Appl. Environ. Microbiol.
<4>78
<5>889-891
<6>2012
<7>Bcep22-like phages are a recently described group of podoviruses that infect
strains of Burkholderia cenocepacia. We have isolated and characterized a novel
member of this group named DC1. This podovirus shows many genomic similarities to
BcepIL02 and Bcep22, but it infects strains belonging to multiple Burkholderia
cepacia complex (BCC) species.

<>

<1>Lynch, T.W., Sligar, S.G.
<2>Macromolecular Hydration Changes Associated with BamHI Binding and Catalysis.
<3>J. Biol. Chem.
<4>275
<5>30561-30565
<6>2000
<7>In this report, the effects of osmotic pressure on BamHI cognate binding and catalysis were
investigated and compared with a previous study on EcoRI (Robinson, C. R. and Sligar, S. G.
(1998) Proc. Natl. Acad. Sci. U.S.A. 95, 2186-2191). Our observation of the dependence of
binding and catalytic parameters on osmotic pressure has allowed for the comparison of
hydration changes associated with site-specific DNA recognition for both endonucleases. Over a
large range of osmotic pressures (pi), the dependence of BamHI on osmotic stress during
cognate binding and catalysis was very different from that of the related endonuclease EcoRI.
The binding of EcoRI to cognate DNA was dominated by a dehydration of the endonuclease-DNA
complex, whereas binding by BamHI to its cognate sequence was accompanied by a solvent release
corresponding to some 125 fewer waters. Catalytic analysis at elevated osmotic pressures
indicated that both endonucleases had undergone a net hydration of the complex with BamHI
displaying a much greater dependence on osmotic stress than EcoRI. Although the enzymes shared
core structural motifs, comparisons of high resolution x-ray structures revealed many
different secondary structural features of the complexed endonucleases. The large difference
in hydration changes by both BamHI and EcoRI could be attributed to these dissimilar secondary
structural features, as well as the functional differences of the two endonucleases during
site-specific DNA recognition.

<>

<1>Lyngstadaas, A., Lobner-Olesen, A., Boye, E.
<2>Characterization of three genes in the dam-containing operon of Escherichia coli.
<3>Mol. Gen. Genet.
<4>247
<5>546-554
<6>1995
<7>The dam-containing operon in Escherichia coli is located at 74 min on the chromosomal map and
contains the genes aroK, aroB, a gene called urf74.3,
dam and trpS. We have determined the nucleotide sequence between the dam
and trpS genes and show that it encodes two proteins with molecular
weights of 24 and 27 kDa. Furthermore, we characterize the three genes
urf74.3, 24kDa, 27kDa and the proteins they encode. The predicted amino
acid sequences of the 24 and 27 kDa proteins are similar to those of the
CbbE and CbbZ proteins, respectively, of the Alcaligenes eutrophus cbb
operon, which encodes enzymes involved in the Calvin cycle. In separate
experiments, we have shown that the 24 kDa protein has
d-ribulose-5-phosphate epimerase activity (similar to CbbE), and we call
the gene rpe. Similarly, the 27 kDa protein has 2-phosphoglycolate
phosphatase activity (similar to CbbZ), and we name the gene gph. The
Urf74.3 protein, with a predicted molecular weight of 46 kDa, migrated as
a 70 kDa product under denaturing conditions. Overexpression of Urf74.3
induced cell filamentation, indicating that Urf74.3 directly or indirectly
interferes with cell division. We present evidence for translational
coupling between aroB and urf74.3 and also between rpe and gph. Proteins
encoded in the dam superoperon appear to be largely unrelated: Dam, and
perhaps Urf74.3, are involved in cell cycle regulation, AroK, AroB, and
TrpS function in aromatic amino acid biosynthesis, whereas Rpe and Gph are
involved in carbohydrate metabolism.

<>

<1>Lynn, S.P., Cohen, L.K., Gardner, J.F., Kaplan, S.
<2>Characterization of a site-specific restriction endonuclease from Rhodopseudomonas sphaeroides.
<3>J. Bacteriol.
<4>138
<5>505-509
<6>1979
<7>A type II restriction endonuclease, RshI, has been partially purified from
photoheterotrophically grown Rhodopseudomonas sphaeroides strain 2.4.1.  The
enzyme preparation, after a single DE-52 column fractionation, is free of 5'
exonuclease and phospatase activities but contains a trace of 3' exonuclease
activity.  Based upon deoxyribonucleic acid (DNA) sequencing data in the
vicinity of the enzyme-promoted cleavage of pBR322 DNA, we have concluded that
RshI probably recognizes the palinodromic hexanucleotide sequence 5'-CGATCG-3'
and cleaves between the T and C.  Lambda cI857 DNA contains three RshI sites,
two of which lie in the replaceable region.  The plasmid pBR322, which carries
resistances to ampicillin and tetracycline, contains a single RshI site in the
ampicillin resistance determinant.  Insertion of DNA into the RshI site of
pBR322 results in loss of ampicillin resistance but retention of tetracycline
resistance, thereby providing a convenient screening procedure for recombinant
plasmids.

<>

<1>Lynn, S.P., Cohen, L.K., Kaplan, S., Gardner, J.F.
<2>RsaI: a new sequence-specific endonuclease activity from Rhodopseudomonas sphaeroides.
<3>J. Bacteriol.
<4>142
<5>380-383
<6>1980
<7>A new type II sequence-specific endonuclease, RsaI, has been identified from
Rhodopseudomonas sphaeroides strain 28/5.  An RsaI purification scheme that
yields enzyme which is free of contaminating exonuclease and phosphatase
activities after a single column fractionation has been developed.  The enzyme
recognized the tetranucleotide sequence 5'-GTAC-3' and cleaved between the T
and A, thereby generating flush ends.  RsaI should be extremely useful in
deoxyribonucleic acid sequencing experiments.

<>

<1>Lyra, C., Halme, T., Torsti, A.-M., Tenkanen, T., Sivonen, K.
<2>Site-specific restriction endonucleases in cyanobacteria.
<3>J. Appl. Microbiol.
<4>89
<5>979-991
<6>2000
<7>Aim: Planktic cyanobacteria were screened for endodeoxyribonucleases. Principal component
analysis (PCA) was employed to demonstrate a
potential relationship between certain enzymes and a group of
cyanobacteria. The data were obtained from a data bank and this study.
Methods and Results: Enzymes were partially purified using column
chromatography. Anabaena strains contained Asp83/1I(5'-TTCGAA-3'),
Asp83/1II (5'-GGCC-3'), Asp90I (5'-ACRYGT-3') and five isoschizomeric
enzymes (5'-ATCGAT-3'). Aphanizomenon and Microcystis strains contained
ApcTR183I (5'-TGCGCA-3') and Msp199I (5'-CCGG-3'), respectively.
Planktothrix strains possessed Psc2I (5'-GAANNNNTTC-3'), Psc27I and
Psc28I (5'-TTCGAA-3'). PCA showed that the most common cyanobacterial
endonuclease types were AvaII, AvaI and AsuII.
Conclusions: All planktic cyanobacteria studied contained
restriction endonucleases. The defined restriction endonucleases were
isoschizomers of known enzymes. The Nostoc and the Spirulina genera had
an association, while the majority of the genera had no association
with certain endonuclease type(s).
Significance and Impact of the Study: The defined enzymes in this
study and the estimated trend in the endonuclease type distribution
allow more efficient avoidance of cynobacterial restriction barriers.

<>

<1>Lyumkis, D., Talley, H., Stewart, A., Shah, S., Park, C.K., Tama, F., Potter, C.S., Carragher, B., Horton, N.C.
<2>Allosteric Regulation of DNA Cleavage and Sequence-Specificity through Run-On Oligomerization.
<3>Structure
<4>21
<5>1848-1858
<6>2013
<7>SgrAI is a sequence specific DNA endonuclease that functions through an unusual enzymatic
mechanism that is allosterically activated 200- to 500-fold by effector
DNA, with a concomitant expansion of its DNA sequence specificity. Using
single-particle transmission electron microscopy to reconstruct distinct
populations of SgrAI oligomers, we show that in the presence of allosteric,
activating DNA, the enzyme forms regular, repeating helical structures
characterized by the addition of DNA-binding dimeric SgrAI subunits in a run-on
manner. We also present the structure of oligomeric SgrAI at 8.6 A resolution,
demonstrating the conformational state of SgrAI in its activated form. Activated
and oligomeric SgrAI displays key protein-protein interactions near the helix
axis between its N termini, as well as allosteric protein-DNA interactions that
are required for enzymatic activation. The hybrid approach reveals an unusual
mechanism of enzyme activation that explains SgrAI's oligomerization and
allosteric behavior.

<>

<1>Lyutzkanova, D., Stoilova-Disheva, M., Peltekova, V.
<2>The restriction-modification system in Streptomyces flavopersicus.
<3>Folia Microbiol. (Praha)
<4>46
<5>119-122
<6>2001
<7>To clone bifunctional vectors in streptomycetes, it was necessary to define the
restriction-modification system of Streptomyces
flavopersicus. Plasmid DNA from bifunctional vectors pIJ699 and
pXED3-13, isolated from E. coli strains with different methylation
systems: E. coli DH5 alpha (dam(+) dcm(+)), E. coli MB5386 (dam dcm), E.
coli CB51 (dam dcm(+)), E. coli NM544 (dam(+) dcm), was used for
transformation of protoplasts from strain S. flavopersicus. Restriction
of dcm-methylated DNA from S. flavopersicus was established. As a host
in the intermediate cloning strain E.coli NM544 (dam(+) dcm) should be
used, as the dcm-transmethylase-dependent strain S. flavopersicus does
not process DNA from this strain.

<>

<1>Ma, A.P., Jiang, J., Tun, H.M., Mauroo, N.F., Yuen, C.S., Leung, F.C.
<2>Complete Genome Sequence of Staphylococcus xylosus HKUOPL8, a Potential Opportunistic Pathogen of Mammals.
<3>Genome Announcements
<4>2
<5>e00653-14
<6>2014
<7>We report here the first complete genome sequence of Staphylococcus xylosus strain HKUOPL8,
isolated from giant panda feces. The whole genome sequence of
this strain will provide an important framework for investigating the genes
responsible for potential opportunistic infections with this species, as well as
its survival in various environments.

<>

<1>Ma, B.C., Wang, J., Fang, X.H.
<2>Fluorescence study of DNA binding and bending by EcoRI DNA methyltransferase.
<3>J. Phys. Chem. B
<4>110
<5>19647-19651
<6>2006
<7>We have applied fluorescence anisotropy and fluorescence resonance energy transfer (FRET)
techniques to study the interaction between
EcoRI DNA methyltransferase (M.EcoRI) and its target DNA in solution.
Upon binding with M.EcoRI, the dsDNA containing GAATTC bends to flip
out the second adenine for methylation. The binding affinity of M.
EcoRI to two dsDNA fragments (20 and 38 bp) was studied with
fluorescence anisotropy. Their binding constants at different
temperatures from 20 to 40 degrees C were obtained, and the
thermodynamic parameters of binding were derived. The results showed
that M.EcoRI had a higher binding affinity to the short dsDNA strand
than to the long one, and its binding to DNA was primarily
entropy-driven. By labeling the 5' ends of the 20-bp dsDNA with two
fluorescent dyes, fluorescein (FAM) and tetramethylrhodamine (TMR), we
were able to monitor the enhanced TMR fluorescence in the presence of
M.EcoRI. The end-to-end distance of the dsDNA determined from the FRET
efficiency was changed from 72.4 to 63.4 angstrom, and the DNA bending
angle was estimated as 57.8 degrees.

<>

<1>Ma, C.B., Tang, Z.W., Wang, K.M., Tan, W.H., Yang, X.H., Li, W., Li, Z.H., Lv, X.Y.
<2>Real-time monitoring of restriction endonuclease activity using molecular beacon.
<3>Anal. Biochem.
<4>363
<5>294-296
<6>2007
<7>Restriction endonucleases are one of the most important enzymes in molecular biology.  These
enzymes are essential in recombinant technology, genotyping, mapping, and sequencing of large
strands of DNA.  Because of the biological importance of restriction endonucleases and their
wide use, restriction endonuclease activity assays are essential.  Traditional methods for
assaying activity of restriciton endonucleases, such as gel electrophoresis, filter binding,
high-performance liquid chromatography, and enzyme-linked immunosorbent assay, have been used.
All of these methods, however, are discontinuous, time-consuming, and laborious, and they
usually require isotope labeling.  Several spectroscopic methods using fluorescence
measurements have been developed.  These assays are continuous.  However, up to now all of
them have been designed only for doubly labeled DNA substrates with very large fluorophores
that may interfere with recognition and reactivity with certain restriction endonucleases.
Furthermore, the potential for achieving high sensitivity through fluorescent resonance energy
transfer has not been fully realized.  Therefore, it is necessary to develop a sensitive,
convenient, and non-isotope-labeled method for assaying restriction endonuclease activity.

<>

<1>Ma, D., Bai, J., Deng, G., Li, S.
<2>Sequence analysis of DNA methyltransferase gene from Largemouth bass ulcerative syndrome virus and its expression in prokaryote.
<3>Nong Ye Sheng Wu Ji Shu Xue Bao
<4>19
<5>342-349
<6>2011
<7>In order to further reveal the characteristics of Largemouth bass ulcerative syndrome virus
(LBUSV) isolated recently, a further study the group of LBUSV, DFV (Doctor fish virus) and
LMBV (Largemouth bass virus), the full-length DNA methyltransferase (DNA MTase) gene was
analyzed and expressed using prokaryotic system.  A about 200 bp core fragment was amplified
and sequenced, and the full-length of DNA MTase was identified by genome walking.  The open
reading frame (ORF) of DNA MTase gene was 663 bp encoding 220 amino acids (GenBank accession
No. GU256634).  Motif analysis indicated that LBUSV DNA MTase protein contained blocks I, IV,
VI and VIII, and cofactor binding sites, substrate interacting sites and DNA binding sites
were also found in LBUSV DNA MTase, and the proline-proline-cysteine tripeptide was proposed
catalytic site.  Additionally, the primers were designed according to DNA MTase ORF, and the
PCR products were inserted in vector pBV220 and transformed to host Escherichia coli DH5a.
After temperature inducement and SDS-PAGE analysis, the recombinant expression bacteria
produced a special proein about 25 kD in molecular weight.  The analysis, the recombinant
expression bacteria produced a special protein about 25 kD in molecular weight.  The
proportion of recombinant protein in total bacterial protein was about 30%.  Characteristic
analysis of DNA MTase gene showed that the predicted protein may play a role of DNA
methyltransferase, and confirmed that the taxonomic status of LBUSV is genus Ranavirus of the
family Iridoviridae.

<>

<1>Ma, D.P., King, Y.T., Kim, Y., Luckett, S.
<2>The group I intron of apocytochrome b gene from Chlamydomonas smithii encodes a site-specific endonuclease.
<3>Plant Mol. Biol.
<4>18
<5>1001-1004
<6>1992
<7>The mitochondrial DNA molecules of two interfertile algal species, Chlamydomonas smithii and
C. reinhardtii, are co-linear except for a 1075 bp intron (the alpha-insert) that is present
in the cob gene of C. smithii. The alpha-insert, a group I intron (Cs cob.1) containing an
open reading frame (ORF) which encodes a basic, hydrophilic protein of 237 amino acids, is
unidirectionally transmitted to all diploid progeny during interspecific crosses. In this
report, we show that the Cs cob.1-encoded protein is a site-specific endonuclease (I-Csm I)
which could mediate intron transfer via the gene conversion mechanism. The Cs cob.1 ORF was
cloned into the vector pMALcr1 and over-expressed as a hybrid protein fused to maltose-binding
protein (MBP). This fusion protein exhibited an in vivo endonuclease activity which
specifically cleaved the intron homing site within the intronless cob gene.

<>

<1>Ma, J., He, Y., Hu, B., Luo, Z.Q.
<2>Genome Sequence of an Environmental Isolate of the Bacterial Pathogen Legionella  pneumophila.
<3>Genome Announcements
<4>1
<5>e00320-13
<6>2013
<7>We report here the genomic sequence of Legionella pneumophila strain LPE509 from  the water
distribution system of a hospital in Shanghai, China. This is the first
complete genome sequence of an environmental L. pneumophila isolate. Genomic
analyses identified approximately 600 genes unique to LPE509 compared to those of
the 7 available L. pneumophila genomes.

<>

<1>Ma, L., Chen, K., Clarke, D.J., Nortcliffe, C.P., Wilson, G.G., Edwardson, J.M., Morton, A.J., Jones, A.C., Dryden, D.T.F.
<2>Resriction endonuclease TseI cleaves A:A and T:T mismatches in CAG and CTG repeats.
<3>Nucleic Acids Res.
<4>41
<5>4999-5009
<6>2013
<7>The type II restriction endonuclease TseI recognizes the DNA target sequence 5'-G^CWGC-3'
(where W=A or T) and cleaves after the first G to produce fragments with three-base
5'-overhangs.  We have determined that it is a dimeric protein capable of cleaving not only
its target sequence but also one containing A:A or T:T mismatches at the central base pair in
the target sequence.  The cleavage of targets containing these mismatches is as efficient as
cleavage of the correct target sequence containing a central A:T base pair.  The cleavage
mechanism does not apparently use a base flipping mechanism as found for some other type II
restriction endonuclease recognizing similarly degenerate target sequences.  The ability of
TseI to cleave targets with mismatches means that it can cleave the unusual DNA hairpin
structures containing A:A or T:T mismatches formed by the repetitive DNA sequences associated
with Huntington's disease (CAG repeats) and myotonic dystrophy type 1 (CTG repeats).

<>

<1>Ma, L., Wu, X., Wilson, G.G., Jones, A.C., Dryden, D.T.F.
<2>Time-resolved fluorescence of 2-aminopurine in DNA duplexes in the presence of the EcoP15I Type III restriction-modification enzyme.
<3>Biochem. Biophys. Res. Commun.
<4>449
<5>120-125
<6>2014
<7>EcoP15I is a Type III DNA restriction and modification enzyme of Escherichia coli. We show
that it contains two modification (Mod) subunits for sequence-specific methylation of DNA and
one copy of a restriction endonuclease (Res) subunit for cleavage of DNA containing
unmethylated target sequences. Previously the Mod(2) dimer in the presence of cofactors was
shown to use nucleotide flipping to gain access to the adenine base targeted for methylation
(Reddy and Rao, J. Mol. Biol. 298 (2000) 597-610.). Surprisingly the Mod(2) enzyme also
appeared to flip a second adenine in the target sequence, one which was not subject to
methylation. We show using fluorescence lifetime measurements of the adenine analogue,
2-aminopurine, that only the methylatable adenine undergoes flipping by the complete
Res(1)Mod(2) enzyme and that this occurs even in the absence of cofactors. We suggest that
this is due to activation of the Mod(2) core by the Res subunit.

<>

<1>Ma, L., Zhu, Z., Li, T., Wang, Z.
<2>Assaying multiple restriction endonucleases functionalities and inhibitions on DNA microarray with multifunctional gold nanoparticle probes.
<3>Biosensors and Bioelectronics
<4>52
<5>118
<6>2014
<7>Herein, a double-stranded (ds) DNA microarray-based resonance light scattering (RLS) assay
with multifunctional gold nanoparticle (GNP) probes has been developed for studying
restriction endonuclease functionality and inhibition. Because of decreasing significantly
melting temperature, the enzyme-cleaved dsDNAs easily unwind to form single-stranded (ss)
DNAs. The ssDNAs are hybridized with multiplex complementary ssDNAs functionalized GNP probes
followed by silver enhancement and RLS detection. Thre restriction endonucleases (EcoRI, BamHI
and EcoRV) and three potential inhibitors (doxorubicin hydrochloride (DOX), ethidium bromide
(EB) and an EcoRI-derived helical peptide (alpha 4)) were selected to demonstrate capability
of the assay. Enzyme activities of restriction endonucleases are detected simultaneously with
high specificity down to the limits of 2.0 x 10(-2) U/mL for EcoRI, 1.1 x 10(-2) U/mL for
BamHI and 1.6 x 10(-2) U/mL for EcoRV, respectively. More importantly, the inhibitory
potencies of t hree inhibitors are showed quantitatively, indicating that our approach has
great promise for high-throughput screening of restriction endonuclease inhibitors.

<>

<1>Ma, M., Wang, C., Ding, Y., Li, L., Shen, D., Jiang, X., Guan, D., Cao, F., Chen, H., Feng, R., Wang, X., Ge, Y., Yao, L., Bing, X., Yang, X., Li, J., Du, B.
<2>Complete Genome Sequence of Paenibacillus polymyxa SC2, a Strain of Plant Growth Promoting Rhizobacteria with Broad-Spectrum Antimicrobial Activity.
<3>J. Bacteriol.
<4>193
<5>311-312
<6>2010
<7>Paenibacillus polymyxa SC2 is an important plant growth promoting rhizobacterium (PGPR). Here
we report the complete genome sequence of P. polymyxa SC2. Multiple sets of functional genes
have been found in the genome. As far as we know, this is the first complete genome sequence
of Paenibacillus polymyxa.

<>

<1>Ma, Q., Qu, Y., Tang, H., Yu, H., Ma, F., Shi, S., Zhang, X., Zhou, H., Zhou, J., Xu, P.
<2>Genome Sequence of a Novel Indigo-Producing Strain, Pseudomonas monteilii QM.
<3>J. Bacteriol.
<4>194
<5>4459-4460
<6>2012
<7>Pseudomonas monteilii is a versatile bacterium found in various niches. A newly isolated
strain, P. monteilii QM, can effectively produce indigoids from indoles.
Here we present a 5.76-Mb assembly of the P. monteilii genome for the first time.
It may provide abundant molecular information for the transformation of
aromatics.

<>

<1>Ma, X.X., Ito, T., Tiensasitorn, C., Jamklang, M., Chongtrakool, P., Boyle-Vavra, S., Daum, R.S., Hiramatsu, K.
<2>Novel Type of Staphylococcal Cassette Chromosome mec Identified in Community-Acquired Methicillin-Resistant Staphylococcus aureus Strains.
<3>Antimicrob. Agents Chemother.
<4>46
<5>1147-1152
<6>2002
<7>We identified a new type of staphylococcal cassette chromosome mec
(SCCmec) from two community-acquired methicillin-resistant Staphylococcus
aureus (MRSA) strains. The novel element, designated type IV SCCmec, had a
unique combination of the class B mec gene complex and the type 2 ccr gene
complex and was much smaller in size (21 to 24 kb) than previously
identified SCCmec elements of hospital-acquired MRSA. Consistent with the
strains' susceptibilities to various non-beta-lactam antibiotics, the type
IV SCCmec was devoid of any antibiotic resistance genes other than the
mecA gene.

<>

<1>Ma, Y.F., Zhang, Y., Zhang, J.Y., Chen, D.W., Zhu, Y., Zheng, H., Wang, S.Y., Jiang, C.Y., Zhao, G.P., Liu, S.J.
<2>The Complete Genome of Comamonas testosteroni Reveals Its Genetic Adaptations to Changing Environments.
<3>Appl. Environ. Microbiol.
<4>75
<5>6812-6819
<6>2009
<7>Members of the gram-negative, strictly aerobic genus Comamonas occur in
various environments. Here we report the complete genome of Comamonas
testosteroni strain CNB-2. Strain CNB-2 has a circular chromosome that is
5,373,643 bp long and has a G+C content of 61.4%. A total of 4,803 open
reading frames (ORFs) were identified; 3,514 of these ORFs are
functionally assigned to energy production, cell growth, signal
transduction, or transportation, while 866 ORFs encode hypothetical
proteins and 423 ORFs encode purely hypothetical proteins. The CNB-2
genome has many genes for transportation (22%) and signal transduction
(6%), which allows the cells to respond and adapt to changing
environments. Strain CNB-2 does not assimilate carbohydrates due to the
lack of genes encoding proteins involved in glycolysis and pentose
phosphate pathways, and it contains many genes encoding proteins involved
in degradation of aromatic compounds. We identified 66 Tct and nine TRAP-T
systems and a complete tricarboxylic acid cycle, which may allow CNB-2 to
take up and metabolize a range of carboxylic acids. This nutritional bias
for carboxylic acids and aromatic compounds enables strain CNB-2 to occupy
unique niches in environments. Four different sets of terminal oxidases
for the respiratory system were identified, and they putatively functioned
at different oxygen concentrations. This study conclusively revealed at
the genomic level that the genetic versatility of C. testosteroni is vital
for competition with other bacteria in its special niches.

<>

<1>Ma, Z., Geng, J., Zhang, H., Yu, H., Yi, L., Lei, M., Lu, C.P., Fan, H.J., Hu, S.
<2>Complete Genome Sequence of Streptococcus equi subsp. zooepidemicus Strain ATCC 35246.
<3>J. Bacteriol.
<4>193
<5>5583-5584
<6>2011
<7>Streptococcus equi subsp. zooepidemicus is an opportunistic pathogen. It has caused a very
large economic loss in the swine industry of China and
has become a threat to human health. We announce the complete genome
sequence of S. equi subsp. zooepidemicus strain ATCC 35246, which provides
opportunities to understand its pathogenesis mechanism and genetic basis.

<>

<1>Ma, Z., Geng, J.N., Yi, L., Xu, B., Jia, R.Y., Li, Y., Meng, Q.S., Fan, H.J., Hu, S.N.
<2>Insight into the specific virulence related genes and toxin-antitoxin virulent pathogenicity islands in swine streptococcosis pathogen Streptococcus equi ssp zooepidemicus strain ATCC35246.
<3>BMC Genomics
<4>14
<5>377
<6>2013
<7>Background: Streptococcus equi ssp. zooepidemicus (S. zooepidemicus) is an important pathogen
causing swine streptococcosis in China. Pathogenicity islands (PAIs) of S. zooepidemicus have
been transferred among bacteria through horizontal gene transfer (HGT) and play important
roles in the adaptation and increased virulence of S. zooepidemicus. The present study used
comparative genomics to examine the different pathogenicities of S. zooepidemicus. Results:
Genome of S. zooepidemicus ATCC35246 (Sz35246) comprises 2,167,264-bp of a single circular
chromosome, with a GC content of 41.65%. Comparative genome analysis of Sz35246, S.
zooepidemicus MGCS10565 (Sz10565), Streptococcus equi. ssp. equi. 4047 (Se4047) and S.
zooepidemicus H70 (Sz70) identified 320 Sz35246-specific genes, clustered into three
toxin-antitoxin (TA) systems PAIs and one restriction modification system (RM system) PAI.
These four acquired PAIs encode proteins that may contribute to the overall pathogenic
capacity and fitness of this bacterium to adapt to different hosts. Analysis of the in vivo
and in vitro transcriptomes of this bacterium revealed differentially expressed PAI genes and
non-PAI genes, suggesting that Sz35246 possess mechanisms for infecting animals and adapting
to a wide range of host environments. Analysis of the genome identified potential Sz35246
virulence genes. Genes of the Fim III operon were presumed to be involved in breaking the
host-restriction of Sz35246. Conclusion: Genome wide comparisons of Sz35246 with three other
strains and transcriptome analysis revealed novel genes related to bacterial virulence and
breaking the host-restriction. Four specific PAIs, which were judged to have been transferred
into Sz35246 genome through HGT, were identified for the first time. Further analysis of the
TA and RM systems in the PAIs will improve our understanding of the pathogenicity of this
bacterium and could lead to the development of diagnostics and vaccines.

<>

<1>Ma, Z., Shen, X., Hu, H., Wang, W., Peng, H., Xu, P., Zhang, X.
<2>Genome Sequence of Sphingomonas wittichii DP58, the First Reported Phenazine-1-Carboxylic Acid-Degrading Strain.
<3>J. Bacteriol.
<4>194
<5>3535-3536
<6>2012
<7>Sphingomonas wittichii DP58 (CCTCC M 2012027), the first reported phenazine-1-carboxylic acid
(PCA)-degrading strain, was isolated from pimiento
rhizosphere soils. Here we present a 5.6-Mb assembly of its genome. This sequence
would contribute to the elucidation of the molecular mechanism of PCA degradation
to improve the antifungal's effectiveness or remove superfluous PCA.

<>

<1>Maas, R.
<2>Change in plasmid DNA structure, hypermethylation, and lon-proteolysis as steps in a replicative cascade.
<3>Cell
<4>105
<5>945-955
<6>2001
<7>I have defined conditions under which RepFIC plasmid DNA can be maintained in a state of
lowered helical density.  In exponentially growing cultures, the DNA of lowered helical
density is present in small amounts but never totally absent, suggesting that it is a normal
variant of plasmid maintenance.  It is fully methylated at frequent sites by
dam-methyltransferase, some not previously recognized, further suggesting that the variant is
a precursor of replication.  The low-helical density plasmid is present in dam hosts,
indicating that methylation is not essential for the change in helical density.  The lowered
helical density is stabilized in lon hosts, suggesting that Lon-protease may remove both the
protein(s) that lower the helical density and the dam-methyltransferase after each round of
replication.

<>

<1>Maass, G.
<2>Recognition of DNA sequences by restriction endonucleases.
<3>NATO ASI Ser.
<4>137
<5>225-237
<6>1987
<7>Type II restriction endonucleases recognize DNA sequences 4 or 6 base pairs
long with very high specificity in a vast excess of non-specific DNA binding
sites.  They cleave the DNA within or in the immediate vicinity of the
recognition sequence.  Because of the immense importance of these enzymes in
analysis, preparation and in vitro recombination of DNA many investigations
have been carried out to understand the structural and mechanistic features
underlying their mode of recognition.  Among approximately 500 type II
restriction endonucleases described, EcoRI is the so far best studied enzyme.
EcoRI is a dimeric protein with two identical subunits, the aminoacid sequence
of which is known.  It recognizes the palindromic sequence -G^AATTC- -CTTAA^G-
and cleaves the DNA double strand at the marked positions.  For its enzymatic
activity it needs Mg2+ as a co-factor.  It was the first DNA-binding protein
that was co-crystallized with its substrate TCGCGAATTCGCG by Frederick et al.
in the absence of Mg2+.  The 3A X-ray analysis revealed a complementary
interface between the enzyme and the major groove of the DNA.  The binding of
the enzyme introduces distortions of the DNA, so called neo-kinks, and thereby
widens the major groove.  In a more recent though preliminary presentation, a
more detailed interpretation of the 3A-structure was given by J. Rosenberg, who
suggested that EcoRI interacts with its canonical sequence via 12 hydrogen
bonds between glutamic acid and arginine residues and the guanine and adenine
residues of GAATTC.  However, the data available at present are not yet
sufficient to understand the molecular basis of the EcoRI action.  A more
detailed knowledge of the thermodynamic, kinetic and structural aspects of the
EcoRI-DNA interaction is needed.  This contribution intends to present
physicochemical studies on the binding and catalytic properties of EcoRI and to
discuss structural aspects of the interaction.

<>

<1>Maass, G., Alves, J., Fliess, A., Pingoud, A., Wolfes, H.
<2>Recognition of DNA-sequences by restriction endonucleases.
<3>Hoppe Seylers Z. Physiol. Chem.
<4>367
<5>47
<6>1986
<7>Class II restriction endonucleases cleave DNA specifically at defined sites.
EcoRI, e.g., which recognizes the sequence GAATTC, cleaves at sites differing
in one base pair from this sequence by a factor of 103 to 105 more slowly than
at its canonical site.  This specificity is a complex function of thermodynamic
and kinetic parameters which govern the non-specific and specific binding of
EcoRI to DNA, the linear diffusion of the protein along DNA and conformational
changes of the enzyme and substrate.  We have been interested in the
thermodynamics, kinetics as well as structural aspects of these processes in
order to understand how DNA recognizing proteins interacting with DNA select
their specific site in a huge excess of nonspecific sites.  The initial contact
between EcoRI and its substrate is to nonspecific sites, binding to these sites
in the presence of Mg2+ is by a least a factor of 100 weaker than to the
specific site.  After nonspecific association EcoRI "slides" along the DNA in a
random fashion:  at physiological Mg2+ concentrations EcoRI cleave several
EcoRI sites in close proximity processively.  At high Mg2+ or at high ionic
strength dissociation is favoured over linear diffusion, and consequently,
processivity is suppressed.  Discrimination between specific and non-specific
sites resides in several unique contacts that can only be formed in the
specific complex.  Many studies have been carried out to define structural
elements on the DNA important for recognition, few studies, however, were
concerned with the identification of the regions of the protein important for
specific binding.  Using bromodeoxyuridine containing oligodeoxynucleotides
EcoRI was photo-crosslinked to its substrate via Met 157 demonstrating the
involvement of this region in the recognition complex.  This identification was
confirmed by the interference of antibodies against this region of EcoRI with
the catalytic action of EcoRI.  Met 157 is at the end of a sequence which shows
a striking homology to the sequence of the recognition helix of gene regulatory
proteins.  We are currently probing the importance of individual amino acid
residues for the catalytic action of EcoRI by site directed mutagenesis.

<>

<1>Maass, G., Ruter, T., Oelgeschlager, T., Alves, J., Wolfes, H., Pingoud, A.
<2>Site-directed mutagenesis studies suggest that Asp91 is part of the catalytic center of the restriction endonuclease EcoRI.
<3>J. Biomol. Struct. Dyn.
<4>8
<5>A126
<6>1991
<7>The comparison of the structures of EcoRI and EcoRV has revealed an interesting
homology which presumably concerns the active center of the two enzymes:
EcoRI:  90 - P D....... 111 E A K
EcoRV:  73 - P D.......  90 D I K
In the crystal structure of EcoRI Glu111 is near the bond cleaved.

<>

<1>Macaluso, A., Mettus, A.-M.
<2>Efficient transformation of Bacillus thuringiensis requires nonmethylated plasmid DNA.
<3>J. Bacteriol.
<4>173
<5>1353-1356
<6>1991
<7>The transformation efficiency of Bacillus thuringiensis depends upon the source of plasmid
DNA. DNA isolated from B. thuringiensis, Bacillus megaterium, or a Dam-Dcm- Escherichia coli
strain efficiently transformed several B. thuringiensis strains. B. thuringiensis strains were
grouped according to which B. thuringiensis backgrounds were suitable sources of DNA for
transformation of other B. thuringiensis strains, suggesting that B. thuringiensis strains
differ in DNA modification and restriction. Efficient transformation allowed the demonstration
of developmental regulation of cloned crystal protein genes in B. thuringiensis.

<>

<1>MacAogain, M., Johari, B.M., Bower, J.E., O'Toole, R.F.
<2>Draft Genome Sequence of a Multidrug-Resistant New Zealand Isolate of Mycobacterium tuberculosis Lineage 3.
<3>Genome Announcements
<4>2
<5>e01017-14
<6>2014
<7>Multidrug resistance constitutes a threat worldwide to the management of tuberculosis (TB). We
report the draft whole-genome sequence of a lineage 3 (East-African Indian) isolate of
Mycobacterium tuberculosis which presented as multidrug resistant in New Zealand, and describe
a number of single-nucleotide polymorphisms in genes relating to drug resistance.

<>

<1>Macdonald, P.M., Mosig, G.
<2>Regulation of a new bacteriophage T4 gene, 69, that spans an origin of DNA replication.
<3>EMBO J.
<4>3
<5>2863-2871
<6>1984
<7>We have determined the DNA sequence and transcription patterns in a 3-kb segment (between 15
and 18 kb on the standard phage T4 map) spanning an origin of DNA replication. A new gene, 69,
spans this origin. Gene 69 codes for two overlapping proteins that share a common C-terminal
segment. Defective DNA replication in an appropriate amber mutant shows that at least the
larger of the two proteins is required for efficient T4 DNA replication. The two proteins
coded by gene 69 are expressed from different transcripts that are under different regulation.
The smaller protein, gp69*, can be expressed immediately from an Escherichia coli-like
promoter, whereas expression of the larger protein, gp69, must be delayed since its middle
promoter requires T4 coded proteins, most likely gp mot, for activation. We discuss the
possible significance of two overlapping proteins in the assembly of replisomes. Gene 69 is
bracketed by the nonessential early gene dam (DNA adenine methylase) and the late gene soc
(small outer capsid protein). Transcripts through this region are interdigitated in a complex
pattern, which reveals all elements that are thought to be important in regulation of
pre-replicative and post-replicative T4 genes.

<>

<1>MacGregor, B.J., Biddle, J.F., Siebert, J.R., Staunton, E., Hegg, E.L., Matthysse, A.G., Teske, A.
<2>Why Orange Guaymas Basin Beggiatoa spp. Are Orange: Single-Filament-Genome-Enabled Identification of an Abundant Octaheme Cytochrome    with Hydroxylamine Oxidase, Hydrazine Oxidase, and Nitrite Reductase Activities.
<3>Appl. Environ. Microbiol.
<4>79
<5>1183-1190
<6>2013
<7>Orange, white, and yellow vacuolated Beggiatoaceae filaments are visually dominant members of
microbial mats found near sea floor hydrothermal vents and
cold seeps, with orange filaments typically concentrated toward the mat centers.
No marine vacuolate Beggiatoaceae are yet in pure culture, but evidence to date
suggests they are nitrate-reducing, sulfide-oxidizing bacteria. The nearly
complete genome sequence of a single orange Beggiatoa (â€œCandidatus
Maribeggiatoaâ€�) filament from a microbial mat sample collected in 2008 at a
hydrothermal site in Guaymas Basin (Gulf of California, Mexico) was recently
obtained. From this sequence, the gene encoding an abundant soluble
orange-pigmented protein in Guaymas Basin mat samples (collected in 2009) was
identified by microcapillary reverse-phase high-performance liquid chromatography
(HPLC) nano-electrospray tandem mass spectrometry (Î¼LCâ€"MS-MS) of a pigmented band
excised from a denaturing polyacrylamide gel. The predicted protein sequence is
related to a large group of octaheme cytochromes whose few characterized
representatives are hydroxylamine or hydrazine oxidases. The protein was
partially purified and shown by in vitro assays to have hydroxylamine oxidase,
hydrazine oxidase, and nitrite reductase activities. From what is known of
Beggiatoaceae physiology, nitrite reduction is the most likely in vivo role of
the octaheme protein, but future experiments are required to confirm this
tentative conclusion. Thus, while present-day genomic and proteomic techniques
have allowed precise identification of an abundant mat protein, and its potential
activities could be assayed, proof of its physiological role remains elusive in
the absence of a pure culture that can be genetically manipulated.

<>

<1>MacGregor, B.J., Biddle, J.F., Teske, A.
<2>Mobile Elements in a Single-Filament Orange Guaymas Basin Beggiatoa ('Candidatus Maribeggiatoa') sp Draft Genome: Evidence for Genetic Exchange with Cyanobacteria.
<3>Appl. Environ. Microbiol.
<4>79
<5>3974-3985
<6>2013
<7>The draft genome sequence of a single orange Beggiatoa ('Candidatus Maribeggiatoa') filament
collected from a microbial mat at a
hydrothermal site in Guaymas Basin (Gulf of California, Mexico) shows
evidence of extensive genetic exchange with cyanobacteria, in
particular for sensory and signal transduction genes. A putative homing
endonuclease gene and group I intron within the 23S rRNA gene; several
group II catalytic introns; GyrB and DnaE inteins, also encoding homing
endonucleases; multiple copies of sequences similar to the fdxN
excision elements XisH and XisI (required for heterocyst
differentiation in some cyanobacteria); and multiple sequences related
to an open reading frame (ORF) (00024 0693) of unknown function all
have close non-Beggiatoaceae matches with cyanobacterial sequences.
Sequences similar to the uncharacterized ORF and Xis elements are found
in other Beggiatoaceae genomes, a variety of cyanobacteria, and a few
phylogenetically dispersed pleiomorphic or filamentous bacteria. We
speculate that elements shared among filamentous bacterial species may
have been exchanged in microbial mats and that some of them may be
involved in cell differentiation.

<>

<1>Macgregor, R.B.
<2>Reversible inhibition of EcoRI with elevated pressure.
<3>Biochem. Biophys. Res. Commun.
<4>170
<5>775-778
<6>1990
<7>The endonuclease activity of EcoRI is completely inhibited at 200 MPa, 37C
using the plasmid pBR322 as a substrate.  When assayed at 133 MPa approximately
half the activity at atmospheric pressure was observed; from these data the
standard molar volume change is estimated to be -80 cm3  mol-1.  Upon return to
atmospheric pressure the enzyme reacted in a standard manner with its
restriction site in pBR322.  Pressurization did not decrease the specificity of
the endonuclease activity of the enzyme for its canonical site.  These results
are discussed in terms of the role of ionic interactions in protein-DNA
interactions.

<>

<1>Machado, H., Mansson, M., Gram, L.
<2>Draft Genome Sequence of Photobacterium halotolerans S2753, Producer of Bioactive Secondary Metabolites.
<3>Genome Announcements
<4>2
<5>e00535-14
<6>2014
<7>We report here the whole draft genome sequence of marine isolate Photobacterium halotolerans
S2753, which produces the known antibiotic holomycin and also
ngercheumicins and solonamides A and B, which interfere with virulence of
methicillin-resistant Staphylococcus aureus strains by interacting with the
quorum-sensing system.

<>

<1>Machado, V.S., Bicalho, R.C.
<2>Complete Genome Sequence of Trueperella pyogenes, an Important Opportunistic Pathogen of Livestock.
<3>Genome Announcements
<4>2
<5>e00400-14
<6>2014
<7>Here, we report the complete genome sequence of Trueperella pyogenes TP6375, a strain isolated
from the uterus of a dairy cow affected with metritis. The
complete circular genome is 2,338,390 bp and contains several genes needed for
pathogenicity.

<>

<1>Macickova-Cahova, H., Hocek, M.
<2>Cleavage of adenine-modified functionalized DNA by type II restriction endonucleases.
<3>Nucleic Acids Res.
<4>37
<5>7612-7622
<6>2009
<7>A set of 6 base-modified 2'-deoxyadenosine derivatives was incorporated to diverse DNA
sequences by primer extension using Vent (exo-) polymerase and
the influence of the modification on cleavage by diverse restriction
endonucleases was studied. While 8-substituted (Br or methyl) adenine
derivatives were well tolerated by the restriction enzymes and the
corresponding sequences were cleaved, the presence of 7-substituted
7-deazaadenine in the recognition sequence resulted in blocking of
cleavage by some enzymes depending on the nature and size of the
7-substituent. All sequences with modifications outside of the recognition
sequence were perfectly cleaved by all the restriction enzymes. The
results are useful both for protection of some sequences from cleavage and
for manipulation of functionalized DNA by restriction cleavage.

<>

<1>Macickova-Cahova, H., Pohl, R., Hocek, M.
<2>Cleavage of Functionalized DNA Containing 5-Modified Pyrimidines by Type II Restriction Endonucleases.
<3>Chembiochem
<4>12
<5>431-438
<6>2011
<7>A series of six pyrimidine-modified dNTPs-5-ethynyl-, 5-phenyl-, and
5-(3-nitrophenyl)deoxycitidine and -deoxyuridine triphosphates-were
prepared and incorporated by primer extension with Vent
(exo-)polymerase to specific DNA sequences within or next to the
recognition sequences of selected restriction endonucleases. The
cleavage of these pyrimidine-modified DNA sequences by 13 restriction
enzymes was then studied. Whereas the presence of any modified C within
the target sequence completely prevented any restriction cleavage, most
enzymes tolerated the presence of 5-ethynylU and two of them even the
presence of 5-phenyl- and 5-(3-nitrophenyl)U. Modifications outside the
recognition sequence were tolerated except in the case of phenyl
derivatives with the PvuII enzyme. 5-EthynylC was used for protection
of the recognition sequence from cleavage in the presence of the second
unmodified copy of the same sequence that was cleaved.

<>

<1>Macinnes, J.I., Mackinnon, J., Bujold, A.R., Ziebell, K., Kropinski, A.M., Nash, J.H.
<2>Complete Genome Sequence of Actinobacillus suis H91-0380, a Virulent Serotype O2  Strain.
<3>J. Bacteriol.
<4>194
<5>6686-6687
<6>2012
<7>Here, we report the first complete genome sequence of Actinobacillus suis, an important
opportunistic pathogen of swine. By comparing the genome sequence of A.
suis with those of other members of the family Pasteurellaceae, we hope to better
understand the role of these organisms in health and disease in swine.

<>

<1>Macintyre, G., Atwood, C.V., Cupples, C.G.
<2>Lowering S-adenosylmethionine levels in Escherichia coli modulates C-to-T transition mutations.
<3>J. Bacteriol.
<4>183
<5>921-927
<6>2001
<7>Deoxycytosine methylase (Dcm) enzyme activity causes mutagenesis in vitro either directly by
enzyme-induced deamination of cytosine to uracil in the absence of the methyl donor,
S-adenosylmethionine (SAM), or indirectly through spontaneous deamination of
[5-methyl]cytosine to thymine. Using a Lac reversion assay, we investigated the contribution
of the first mechanism to Dcm mutagenesis in vivo by lowering the levels of SAM. Escherichia
coli SAM levels were lowered by reducing SAM synthetase activity via the introduction of a
metK84 allele or by hydrolyzing SAM using the bacteriophage T3 SAM hydrolase.  The metK84
strains exhibited increased C-to-T mutagenesis. Expression of the T3 SAM hydrolase gene, under
the control of the arabinose-inducible P(BAD) promoter, effectively reduced Dcm-mediated
genomic DNA methylation. However, increased mutagenesis was not observed until extremely high
arabinose concentrations were used, and genome methylation at Dcm sites was negligible.

<>

<1>Macintyre, G., Doiron, K., Cupples, G.
<2>Analysis of the levels of Dcm and Vsr in E. coli.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>97
<5>294
<6>1997
<7>In Escherichia coli, T/G mismatches at C(T/G)WGG are cleaved by Vsr endonuclease, this is the
initial step in the very short patch repair process which repairs the T/G mismatch to C/G.
Since Dcm cytosine methylase methylates the second C in CCWGG, the function of VSP repair is
probably the prevention of CG to TA mutations caused by deamination of 5-methylcytosine to
thymine.  The dcm and vsr genes overlap, and are expressed from a single promoter situated
upstream of dcm.  The cellular levels of Dcm and Vsr are unknown.  We are investigating the
consequences of altering the levels of these two proteins.  Overexpression of Vsr in E. coli
stimulates transition and frameshift mutations.  Overexpression of dcm does not stimulate
mutations in cells with a functional vsr; in vsr-deleted cells, high levels of Dcm stimulate C
to T transitions at 5-methylcytosines.  Thus the expression of a single copy of vsr on the
chromosome is sufficient to counteract the mutagenic effect of even high levels of Dcm.  Since
Vsr is mutagenic, the organization of these two genes may reflect the need to produce low
levels of Vsr.  This would allow Vsr to counteract the mutagenic effects of Dcm, while
minimizing the mutagenic effect of Vsr.  We have constructed plasmids which allow the
purification of 6his-Dcm and 6his-Vsr.  The purified Dcm and Vsr will be used to produce
antibodies, to analyse the relative levels of Dcm and Vsr in E. coli by Western blot analysis.

<>

<1>Macintyre, G., Doiron, K.M.J., Cupples, C.G.
<2>The Vsr endonuclease of Escherichia coli: an efficient DNA repair enzyme and a potent mutagen.
<3>J. Bacteriol.
<4>179
<5>6048-6052
<6>1997
<7>The Vsr endonuclease of Escherichia coli initiates the repair of T/G mismatches caused by
deamination of 5-methylcytosine to thymine.  In this paper, we examine the capacity of Vsr to
prevent CG-to-TA mutations in cells with increased transcription of the cytosine methylase
gene (dcm).  We find that sufficient Vsr is produced by a single chromosomal copy of vsr to
prevent mutagenesis.  We also investigate the cause of the transition and frameshift mutations
in cells overproducing Vsr.  Neither the absence of the dcm methylase nor its overproduction
affects Vsr-stimulated mutagenesis.  However, addition of mutS, mutL, or mutH on multicopy
plasmids has a significant effect: mutL or mutH decreases the number of mutations, while mutS
stimulates mutagenesis.  The mut-containing plasmids have the same effect in cells treated
with 2-aminopurine and in cells made defective in DNA proofreading, two experimental
situations known to cause transition and frameshift mutations by saturating mismatch repair.

<>

<1>Macintyre, G., Pitsikas, P., Cupples, C.G.
<2>Growth phase-dependent regulation of Vsr endonuclease may contribute to 5-methylcytosine mutational hot spots in Escherichia coli.
<3>J. Bacteriol.
<4>181
<5>4435-4436
<6>1999
<7>Using rabbit polyclonal antibodies, we have shown that the Dcm cytosine methylase of
Escherichia coli is maintained at a constant level during cell growth, while Vsr endonuclease
levels are growth phase dependent. Decreased production of Vsr relative to Dcm during the log
phase may contribute substantially to the mutability of 5-methylcytosine.

<>

<1>Mackeldanz, P., Alves, J., Moncke-Buchner, E., Wyszomirski, K.H., Kruger, D.H., Reuter, M.
<2>Functional consequences of mutating conserved SF2 helicase motifs in the Type III restriction endonuclease EcoP15I translocase domain.
<3>Biochimie
<4>95
<5>817-823
<6>2013
<7>For efficient DNA hydrolysis, Type III restriction endonuclease EcoP15I interacts with two
inversely oriented recognition sites in an
ATP-dependent process. EcoP15I consists of two methylation (Mod)
subunits and a single restriction (Res) subunit yielding a
multifunctional enzyme complex able to methylate or to hydrolyse DNA.
Comprehensive sequence alignments, limited proteolysis and mass
spectroscopy suggested that the Res subunit is a fusion of a motor or
translocase (Tr) domain of superfamily II helicases and an endonuclease
domain with a catalytic PD...EXK motif. In the Tr domain, seven
predicted helicase motifs (I, la, II VI), a recently discovered Q-tip
motif and three additional regions (Ilia, IVa, Va) conserved among Type
III restriction enzymes have been identified that are predicted to be
involved in DNA binding and ATP hydrolysis. Because DNA unwinding
activity for EcoP15I (as for bona fide helicases) has never been found
and EcoP15I ATPase rates are only low, the functional importance of the
helicase motifs and regions was questionable and has never been probed
systematically. Therefore, we mutated all helicase motifs and conserved
regions predicted in Type III restriction enzyme EcoP15I and examined
the functional consequences on EcoP15I enzyme activity and the
structural integrity of the variants by CD spectroscopy. The resulting
eleven enzyme variants all, except variant IVa, are properly folded
showing the same secondary structure distribution as the wild-type
enzyme. Classical helicase motifs I VI are important for ATP and DNA
cleavage by EcoP15I and mutations therein led to complete loss of
ATPase and cleavage activity. Among the catalytically inactive enzyme
variants three preserved the ability to bind ATP. In contrast, newly
assigned motifs Q-tip, Ia and Va are not essential for EcoP15I activity
and the corresponding enzyme variants were still catalytically active.
DNA binding was only marginally reduced (2-7 fold) in all enzyme
variants tested.

<>

<1>Mackenzie, D.A., McLay, K., Roos, S., Walter, J., Swarbreck, D., Drou, N., Crossman, L.C., Juge, N.
<2>Draft Genome Sequence of a Novel Lactobacillus salivarius Strain Isolated from Piglet.
<3>Genome Announcements
<4>2
<5>e01231-13
<6>2014
<7>Lactobacillus salivarius is part of the vertebrate indigenous microbiota of the
gastrointestinal tract, oral cavity, and milk. The properties associated with
some L. salivarius strains have led to their use as probiotics. Here we describe
the draft genome of the pig isolate L. salivarius cp400, providing insights into
host-niche specialization.

<>

<1>Macklaim, J.M., Gloor, G.B., Anukam, K.C., Cribby, S., Reid, G.
<2>At the crossroads of vaginal health and disease, the genome sequence of Lactobacillus iners AB-1.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>4688-4695
<6>2011
<7>Lactobacilli have long been regarded as important constituents of the healthy
human vagina. Lactobacillus iners is the most frequently detected bacterial
species in the vagina, but little is known about its characteristics. We report a
description of the whole-genome sequence of L. iners AB-1 along with comparative
analysis of published genomes of closely related strains of lactobacilli. The
genome is the smallest Lactobacillus reported to date, with a 1.3-Mbp single
chromosome. The genome seems to have undergone one or more rapid evolution events
that resulted in large-scale gene loss and horizontal acquisition of a number of
genes for survival in the vagina. L. iners may exhibit specialized adaptation
mechanisms to the vaginal environment, such as an iron-sulfur cluster assembly
system, and several unique sigma factors to regulate gene transcription in this
fluctuating environment. A potentially highly expressed homolog of a
cholesterol-binding lysin may also contribute to host cell adhesion or act as a
defense mechanism against other microbes. Notably, there is a lack of apparent
adhesion proteins, but several cell-anchor proteins were identified and may be
important for interaction with the host mucosal tissues. L. iners is widely
present in healthy females as well as those suffering from bacterial vaginosis or
who have undergone antimicrobial therapy, suggesting that it is an important
indigenous species of the vagina.

<>

<1>Mackova, M., Pohl, R., Hocek, M.
<2>Polymerase Synthesis of DNAs Bearing Vinyl Groups in the Major Groove and their Cleavage by Restriction Endonucleases.
<3>Chembiochem
<4>15
<5>2306-2312
<6>2014
<7>DNA molecules containing 5-vinyluracil, 5-vinylcytosine, or 7-deaza-7-vinyladenine were
prepared by polymerase incorporation of the corresponding vinyl-modified 2-deoxyribonucleoside
triphosphates, and the influence of the vinyl group in the major groove of DNA on the cleavage
by diverse type II restriction endonucleases (REs) was studied. The presence of 5-vinyluracil
was tolerated by most of the REs, whereas only some REs were able to cleave sequences
containing 7-deaza-7-vinyladenine. The enzyme ScaI was found to cleave DNA containing
5-vinylcytosine efficiently but not DNA containing the related 5-ethynylcytosine. All other
REs failed to cleave sequences containing any cytosine modifications.

<>

<1>MacLeod, A.R., Szyf, M.
<2>Expression of antisense to DNA methyltransferase mRNA induces DNA demethylation and inhibits tumorigenesis.
<3>J. Biol. Chem.
<4>270
<5>8037-8043
<6>1995
<7>Many tumor cell lines overexpress DNA methyltransferase (MeTase) activity; however it is still
unclear whether this increase in DNA MeTase activity plays a casual role in naturally
occurring tumors and cell lines, whether it is critical for the maintenance of transformed
phenotypes, and whether inhibition of the DNA MeTase in tumor cells can reverse
transformation. To address these basic questions, we transfected a murine adrenocortical tumor
cell line Y1 with a chimeric construct expressing 600 base pairs from the 5' of the DNA
MeTase cDNA in the antisense orientation. The antisense transfectants show DNA demethylation,
distinct morphological alterations, are inhibited in their ability to grow in an
anchorage-independent manner, and exhibit decreased tumorigenicity in syngeneic mice. Ex vivo,
cells expressing the antisense construct show increased serum requirements, decreased rate of
growth, and induction of an apoptotic death program upon serum deprivation.
5-Azadeoxycytidine-treated cells exhibit a similar dose-dependent reversal of the transformed
phenotype. These results support the hypothesis that the DNA MeTase is actively involved in
oncogenic transformation.

<>

<1>MacLeod, R.A.
<2>Optimized antisense oligonucleotides complementary to DNA methyltransferase sequences.
<3>International Patent Office
<4>WO 9940186
<5>
<6>1999
<7>The invention provides optimized antisense oligonucleotides complementary to the DNA
methyltransferase gene or its RNA transcript.  The invention further provides methods for
using such antisense oligonucleotides as analytical and diagnostic tools, as potentiators of
transgenic plant and animal studies and gene therapy approaches, and as potential therapeutic
agents.

<>

<1>MacMillan, A.M., Chen, L., Verdine, G.L.
<2>Synthesis of an oligonucleotide suicide substrate for DNA methyltransferases.
<3>J. Org. Chem.
<4>57
<5>2989-2991
<6>1992
<7>The large-scale chemical synthesis of an oligodeoxynucleotide containing
5-fluoro-2'-deoxycytidine (FdC)and its characterization are described. The FdC residue is
introduced via the corresponding 4-O-(2,4,6-trimethylphenyl)-2'-deoxyuridine derivative,
which undergoes clean conversion to FdC during removal of the oligonucleotide protecting
groups with ammonia. A double-stranded oligodeoxynucleotide containing FdC inactivated the DNA
methyltransferase enzyme M.HaeIII by irreversible formation of a covalent protein-DNA complex.

<>

<1>MacNeil, D.
<2>Characterization of a unique methyl-specific restriction system in Streptomyces avermitilis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>88
<5>156
<6>1988
<7>Streptomyces avermitilis contains a restriction system that restricts plasmid DNA containing
N6-methyladenine or 5-methylcytosine. This system restricts DNA isolated from strains with DNA
methylases. Shuttle vectors iolated from Escherichia coli, or plasmids isolated from
modification proficient Streptomyces cannot be directly introduced into S. avermitilis. S.
lividans appears to lack the ability to modify DNA. Plasmids isolated from S. lividans which
have been modified in vitro with DNA methylases are restricted by S. avermitilis. The
transformation frequency is reduced 1000-fold when plasmid DNA is modified by dam or TaqI
methylases to contain N6-methyladenine, or by AluI, HhaI, or HphI methylases to contain
5-methylcytosine. Methyl-specific restriction appears to be common in Streptomyces, since
either N6-methyladenine-specific restriction, or 5-methylcytosine-specific restriction was
observed in 7 of 9 strains tested. S. avermitilis is unique in that it restricts both
N6-methyladenine and 5-methylcytosine containing DNA.

<>

<1>MacNeil, D.J.
<2>Characterization of a unique methyl-specific restriction system in Streptomyces avermitilis.
<3>J. Bacteriol.
<4>170
<5>5607-5612
<6>1988
<7>Streptomyces avermitilis contains a unique restriction system that restricts plasmid DNA
containing N6-methyladenine or 5-methylcytosine. Shuttle vectors isolated from Escherichia
coli RR1 or plasmids isolated from modification-proficient Streptomyces spp. cannot be
directly introduced into S. avermitilis. This restriction barrier can be overcome by first
transferring plasmids into Streptomyces lividans or a modification-deficient E. coli strain
and then into S. avermitilis. The transformation frequency was reduced ->1,000-fold when
plasmid DNA was modified by dam or TaqI methylases to contain N6-methyladenine or by AluI,
HhaI, or HphI methylases to contain 5-methylcytosine. Methyl-specific restriction appears to
be common in Streptomyces spp., since either N6-methyladenine-specific or
5-methylcytosine-specific restriction was observed in seven of nine strains tested.

<>

<1>Macreadie, I.G., Scott, R.M., Zinn, A.R., Butow, R.A.
<2>Transposition of an intron in yeast mitochondria requires a protein encoded by that intron.
<3>Cell
<4>41
<5>395-402
<6>1985
<7>The optional 1143 bp intron in the yeast mitochondrial 21S rRNA gene (Omega+) is nearly
quantitatively inserted in genetic crosses into 21S rRNA alleles that lack it (Omega-). The
intron contains an open reading frame that can encode a protein of 235 amino acids, but no
function has been ascribed to this sequence. We previously found an in vivo double-strand
break in Omega- DNA at or close to the intron insertion site only in zygotes of Omega+ x
Omega- crosses that appears with the same kinetics as intron insertion. We now show that
mutations in the intron open reading frame that would alter the translation product
simulataneously inhibit nonreciprocal Omega recombination and the in vivo double-strand break
in Omega- DNA. These results provide evidence that the open reading frame encodes a protein
required for intron transposition and support the role of the double strand break in the
process.

<>

<1>MacWilliams, M., Meister, J., Jutte, H., Bickle, T.
<2>Generation of a new type-I restriction-modification specificity by transposition.
<3>J. Cell Biochem. Suppl.
<4>18C
<5>136
<6>1994
<7>*
We have characterised a novel mutant of EcoDXXI, a type IC DNA restriction and modification
system, in which the specificity has been altered due to a Tn5 insertion into the middle of
hsdS, the gene which codes the polypepide that confers DNA sequence specificity to both the
restriction and modification reactions. With the type IR-M systems, both the DNA restriction
and modification functions are carried out by a single enzyme composed of 3 subunits: hsdS
(DNA binding specificity), hsdM (modification/methylation), and hsdR (restriction). A complex
of only hsdS and hsdM can catalyse methylation but not restriction. Type I restriction of
unmodified DNA occurs a great distance from the enzyme's recognition site and is accompanied
by large amounts of ATP hydrolysis. It is proposed that the ATP hydrolysis fuels the "pumping"
of the DNA past the bound enzyme to reach the cleavage site.

Like other type I enzymes, the wild type EcoDXXI recognises a sequence composed of two
asymmetrical half sites separated by a spacer region: TCA(N7)RTTC. Purification of the EcoDXXI
mutant methylase and subsequent in vitro DNA methylation assays, identified the mutant
recognition sequence as an interrupted palindrome. TCA(N8)TGA, in which the 5' half site of
the wild type site is repeated in inverse orientation. The additional base pair in the
non-specific spacer of the mutant recognition sequence maintains the proper spacing between
the two methylatable adenine groups.

Sequencing of both the wild type and mutant EcoDXXI hsdS genes showed that the Tn5 insertion
occurred at nucleotide 673 of the 1221 bp gene. This effectively deletes the entire carboxyl
DNA binding domain which recognises the 3' half of the EcoDXXI binding site. Subsequent
deletion analysis demonstrated that the Tn5 element as well as the distal hsdS3' sequence are
dispensible; an additional 65 bp of the hsdS sequence could also be removed without loss of
methylation or restriction activity. The minimum characterised hsdS fragment encodes both the
amino terminal DNA binding domain as well as the conserved repeated sequence that defines the
length of the recognition site spacer region. The predicted 205 amino acid peptide must
contain all the information necessary for DNA binding and subunit interactions. We propose
that the EcoDXXI mutant methylase utilises two truncated hsdS subunits to recognise its
binding site. The implications of this finding in terms of subunit interactions and the
malleability of the type I R-M system will be discussed.


<>

<1>MacWilliams, M.P., Bickle, T.A.
<2>Generation of new DNA binding specificity by truncation of the type IC EcoDXXI hsdS gene.
<3>EMBO J.
<4>15
<5>4775-4783
<6>1996
<7>The hsdS subunit of a type IC restriction-modification enzyme is responsible for the enzyme's
DNA binding specificity.  Type I recognition sites are characterized by two defined half-sites
separated by a non-specific spacer of defined length.  The hsdS subunit contains two
independent DNA binding domains, each targeted towards one DNA half-site.  We have shown
previously that the 5' half of hsdS can code for a functional substitute of the full-length
hsdS.  Here we demonstrate that the 3' half of the gene, when fused to the appropriate
transcriptional and translational start signals, also codes for a peptide which imparts DNA
binding specificity to the enzyme.  About half the natural hsdS size, the mutant peptide
contains a single DNA recognition domain flanked by one copy of each internal repeat found in
the full-length hsdS.  Deletion of either repeat sequence results in loss of activity.  Like
the 5' hsdS mutant, the 3' mutant recognizes an interrupted palindrome, GAAYN5RTTC,
suggesting that two truncated subunits participate in DNA recognition.  Coexpression of the
5' hsdS mutant and the 3' hsdS mutant along with hsdM regenerates the wild-type methylation
specificity.  Thus, there is a free assortment of subunits in the cell.

<>

<1>Madhaiyan, M., Chan, K.L., Ji, L.
<2>Draft Genome Sequence of Methylobacterium sp. Strain L2-4, a Leaf-Associated Endophytic N-Fixing Bacterium Isolated from Jatropha curcas L.
<3>Genome Announcements
<4>2
<5>e01306-14
<6>2014
<7>Methylobacterium sp. strain L2-4 is an efficient nitrogen-fixing leaf colonizer of biofuel
crop Jatropha curcas. This strain is able to greatly improve the
growth and seed yield of Jatropha curcas and is the second reported genome
sequence of plant growth-promoting bacteria isolated from Jatropha curcas.

<>

<1>Madhaiyan, M., Peng, N., Ji, L.
<2>Complete Genome Sequence of Enterobacter sp. Strain R4-368, an Endophytic N-Fixing Gammaproteobacterium Isolated from Surface-Sterilized Roots of Jatropha   curcas L.
<3>Genome Announcements
<4>1
<5>e00544-13
<6>2013
<7>Enterobacter sp. strain R4-368 is one of the few characterized Jatropha endophytic
diazotrophic bacteria and was isolated from surface-sterilized roots.
This bacterium shows strong growth-promoting effects, being able to increase
plant biomass and seed yields. Enterobacter sp. R4-368 is the second fully
sequenced diazotrophic Enterobacter species. The sequence information shall
facilitate the elucidation of the molecular mechanisms of plant growth promotion,
nitrogen fixation in nonlegume plant species, and evolution of biological
nitrogen fixation systems.

<>

<1>Madhavan, T.P., Steen, J.A., Hugenholtz, P., Sakellaris, H.
<2>Genome Sequence of Enterotoxigenic Escherichia coli Strain B2C.
<3>Genome Announcements
<4>2
<5>e00247-14
<6>2014
<7>Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrheal disease around the
globe, causing an estimated 380,000 deaths annually. The disease is caused by a wide variety
of strains. Here, we report the genome sequence of ETEC strain B2C, which was isolated from an
American soldier in Vietnam.

<>

<1>Madhavilatha, G.K., Joseph, B.V., Paul, L.K., Kumar, R.A., Hariharan, R., Mundayoor, S.
<2>Whole-Genome Sequences of Two Clinical Isolates of Mycobacterium tuberculosis from Kerala, South India.
<3>J. Bacteriol.
<4>194
<5>4430
<6>2012
<7>We report the annotated genome sequence of two clinical isolates of Mycobacterium tuberculosis
isolated from Kerala, India.

<>

<1>Madhusoodanan, J., Seo, K.S., Remortel, B., Park, J.Y., Hwang, S.Y., Fox, L.K., Park, Y.H., Deobald, C.F., Wang, D., Liu, S., Daugherty, S.C., Gill, A.L., Bohach, G.A., Gill, S.R.
<2>An Enterotoxin-Bearing Pathogenicity Island in Staphylococcus epidermidis.
<3>J. Bacteriol.
<4>193
<5>1854-1862
<6>2011
<7>Cocolonization of human mucosal surfaces causes frequent encounters
between various staphylococcal species, creating opportunities for the
horizontal acquisition of mobile genetic elements. The majority of
Staphylococcus aureus toxins and virulence factors are encoded on S.
aureus pathogenicity islands (SaPIs). Horizontal movement of SaPIs between
S. aureus strains plays a role in the evolution of virulent clinical
isolates. Although there have been reports of the production of toxic
shock syndrome toxin 1 (TSST-1), enterotoxin, and other superantigens by
coagulase-negative staphylococci, no associated pathogenicity islands have
been found in the genome of Staphylococcus epidermidis, a generally less
virulent relative of S. aureus. We show here the first evidence of a
composite S. epidermidis pathogenicity island (SePI), the product of
multiple insertions in the genome of a clinical isolate. The taxonomic
placement of S. epidermidis strain FRI909 was confirmed by a number of
biochemical tests and multilocus sequence typing. The genome sequence of
this strain was analyzed for other unique gene clusters and their
locations. This pathogenicity island encodes and expresses staphylococcal
enterotoxin C3 (SEC3) and staphylococcal enterotoxin-like toxin L (SElL),
as confirmed by quantitative reverse transcription-PCR (qRT-PCR) and
immunoblotting. We present here an initial characterization of this novel
pathogenicity island, and we establish that it is stable, expresses
enterotoxins, and is not obviously transmissible by phage transduction. We
also describe the genome sequence, excision, replication, and packaging of
a novel bacteriophage in S. epidermidis FRI909, as well as attempts to
mobilize the SePI element by this phage.

<>

<1>Madhusoodanan, U.K., Rao, D.N.
<2>Diversity of DNA methyltransferases that recognize asymmetric target sequences.
<3>Crit. Rev. Biochem. Mol. Biol.
<4>45
<5>125-145
<6>2010
<7>DNA methyltransferases (MTases) are a group of enzymes that catalyze the methyl group transfer
from S-adenosyl-L-methionine in a sequence-specific manner. Orthodox Type II DNA MTases
usually recognize palindromic DNA sequences and add a methyl group to the target base (either
adenine or cytosine) on both strands. However, there are a number of MTases that recognize
asymmetric target sequences and differ in their subunit organization. In a bacterial cell,
after each round of replication, the substrate for any MTase is hemimethylated DNA, and it
therefore needs only a single methylation event to restore the fully methylated state. This is
in consistent with the fact that most of the DNA MTases studied exist as monomers in solution.
Multiple lines of evidence suggest that some DNA MTases function as dimers. Further,
functional analysis of many restriction-modification systems showed the presence of more than
one or fused MTase genes. It was proposed that presence of two MTases responsible for the
recognition and methylation of asymmetric sequences would protect the nascent strands
generated during DNA replication from cognate restriction endonuclease. In this review, MTases
recognizing asymmetric sequences have been grouped into different subgroups based on their
unique properties. Detailed characterization of these unusual MTases would help in better
understanding of their specific biological roles and mechanisms of action. The rapid progress
made by the genome sequencing of bacteria and archaea may accelerate the identification and
study of species- and strain-specific MTases of host-adapted bacteria and their roles in
pathogenic mechanisms.</.

<>

<1>Madsen, A., Josephsen, J.
<2>Characterization of LlaCI, a new restriction-modification system from Lactococcus lactis subsp. cremoris W15.
<3>Biol. Chem.
<4>379
<5>443-449
<6>1998
<7>The genes encoding the restriction-modification system LlaCI have been found on the naturally
occurring 7.0 kb plasmid pAW153 in L. lactis subsp. cremoris W15.  The R/M system was isolated
on a chloramphenicol resistant derivative of the wild type plasmid (pAW153cat).  Plasmid
pAW153cat and a 2.4 kb HincII-SphI fragment cloned into a high- and a low-copy vector
conferred decreased sensitivity in L. lactis LM2301 and L. lactis SMQ86 against small
isometric-headed phages of the 936 or P335 species, respectively.  Increased plasmid copy
number enhanced the level of phage restriction.  Sequencing the 2.4 kb HincII-SphI fragment
revealed two open reading frames arranged convergently with a 94 bp separation.  IIaCIM showed
66% identity to hindIIIM, and IlaCIR showed 45% identity to hindIIIR.  The organization of the
LlaCI operon differs from the HindIII operon, where the endonuclease and methylase genes
overlap and are transcribed in the same direction.  The LlaCI methylase is predicted to be 296
amino acids long, with 63% identity to the HindIII methylase, while the LlaCI endonuclease is
predicted to consist of  324 or 332 amino acids, depending on the position of the start codon.
It shows 24% identity to the HindIII endonuclease.

<>

<1>Madsen, A., Josephsen, J.
<2>Cloning and characterization of the lactococcal plasmid-encoded type II restriction/modification system, LlaDII.
<3>Appl. Environ. Microbiol.
<4>64
<5>2424-2431
<6>1998
<7>The LlaDII restriction/modification system was found on the naturally occurring 8.9-kb plasmid
pHW393 in Lactococcus lactis subsp. cremoris W39.  A 2.4-kb PstI-EcoRI fragment inserted into
the Escherichia coli-L.lactis shuttle vector pCI3340 conferred to L. lactis LM2301 and L.
lactis SMQ86 resistance against representatives of the three most common lactococcal phage
species: 936, P335, and c2.  The LlaDII endonuclease was partially purified and found to
recognize and cleave the sequence 5'-GC/NGC-3', where the arrow indicates the cleavage site.
It is thus an isoschizomer of the commercially available restriction endonuclease Fnu4HI.
Sequencing of the 2.4-kb PstI-EcoRI fragment revealed two open reading frames-arranged
tandemly and separated by a 105-bp intergenic region.  The endonuclease gene of 543 bp
preceded the methylase gene of 954 bop.  The deduced amino acid sequence of the LlaDII R/M
system showed high homology to that of its only sequence isoschizomer, Bsp6I from Bacillus sp.
strain RFL6, with 41% identity between the endonucleases and 60% identity between the
methylases.  The genetic organizations of the LlaDII and Bsp6I R/M systems are identical.
Both methylases have two recognition sites (5'-GCGGC-3' and 5'-GCCGC-3') forming a
putative stem-loop structure spanning part of the presumed -35 sequence and part of the
intervening region between the -35 and -10 sequences.  Alignment of the LlaDII and Bsp6I
methylases with other m5C methylases showed that the protein primary structures possessed the
same organization.

<>

<1>Madsen, A., Josephsen, J.
<2>The LlaGI restriction and modification system of Lactococcus lactis W10 consists of only one single polypeptide.
<3>FEMS Microbiol. Lett.
<4>200
<5>91-96
<6>2001
<7>The naturally occurring 12.1-kb plasmid, pEW104, in Lactococcus lactis ssp. cremoris W10 was
found to confer decreased bacteriophage sensitivity to its host. Plasmid pEW104 encodes a
non-classic restriction and modification (R/M) system, named LlaGI, consisting of only one
single polypeptide. Analysis of the amino acid sequence revealed the presence of a catalytic
motif and seven helicase-like motifs (DEAD-box motifs) characteristic of type I and III
endonucleases, followed by four conserved methylase motifs characteristic of
adenine-methylases. A comparison between LlaGI and the very similar R/M system, LlaBIII,
suggests that the C-terminal region of LlaGI, apparently containing no known motifs, could
possibly specify target DNA recognition. Conceivably, the LlaGI gene is included in the operon
of the plasmid replication machinery. Finally, it is proposed that LlaGI represents a variant
of the type I R/M systems.

<>

<1>Madsen, A., Nellemann, L.J., Josephsen, J.
<2>The restriction and modification system LlaCI and its use in the development of phage resistant lactococcal starter cultures.
<3>FASEB J.
<4>11
<5>A1035
<6>1997
<7>The mixed Cheddar starter culture, TK5, consisting of Lactococcus lactis strains, is highly
bacteriophage resistant, and has been used for cheese production for 12 years.  One of the
major problems encountered in the dairy industry is infection of the starter cultures by lytic
phages.  It is possible to obtain plasmids encoding phage defense mechanisms from bacterial
isolates of TK5.  The 7 kb plasmid, pAW153, from isolate W15, encodes the type II R/M system,
LlaCI with recognition sequence AAGCTT, an isoschizomer of HindIII.  LlaCI restricts the small
isometric headed lactococcal phage p2 with an EOP of 10^-2, this effect, together with other
bacteriophage resistance mechanisms, will be utilized in experiments for the development of
phage resistant starter cultures.  A 2.4 kb HincII-SphI fragment containing LlaCI was
subcloned from pAW153 into pC13340 to generate pCAC1.  The nucleotide sequence of this
fragment encoding the LlaCI activity has been determined.  Analysis of the DNA sequence
predicts; A, a methylase of 296 amino acids showing 62% identity to M.HindIII, and B, an
endonuclease of 332 amino acids showing 24% identity to R.HindIII.  The methylase is an
adenine methylase containing the I-D-PY, LK, T-KP-L, and LD-F-GSGTT-A motifs characteristic of
this class of adenine methylases.  Upstream of the two genes, putative ribosome binding sites
and promoter regions are situated.

<>

<1>Madsen, A., Westphal, C., Josephsen, J.
<2>Characterization of a novel plasmid-encoded HsdS subunit, S.LlaW12I, from Lactococcus lactis W12.
<3>Plasmid
<4>44
<5>196-200
<6>2000
<7>A novel type I restriction-modification specificity subunit, S.LlaW12I, has been identified on
the naturally occurring 8.0-kb plasmid pAW122 in the lactic acid bacterium Lactococcus lactis
subsp. cremoris W12. Presence of the HsdS protein together with a complete type I
restriction-modification system conferred increased phage restriction to the host, indicating
exchange of specificity subunits. Sequence analysis showed that the S.LlaW12I subunit is most
probably of type IC. Presumably, the hsdS gene is organized together with the repB gene on one
transcriptional unit.

<>

<1>Maeda, A.H., Nishi, S., Ozeki, Y., Ohta, Y., Hatada, Y., Kanaly, R.A.
<2>Draft Genome Sequence of Sphingobium sp. Strain KK22, a High-Molecular-Weight Polycyclic Aromatic Hydrocarbon-Degrading Bacterium Isolated from Cattle Pasture   Soil.
<3>Genome Announcements
<4>1
<5>e00911-13
<6>2013
<7>Sphingobium sp. strain KK22 was isolated from a bacterial consortium that originated from
cattle pasture soil from Texas. Strain KK22 grows on phenanthrene
and has been shown to biotransform the high-molecular-weight (HMW) polycyclic
aromatic hydrocarbon (PAH) benz[a]anthracene. The genome of strain KK22 was
sequenced to investigate the genes involved in aromatic pollutant
biotransformation.

<>

<1>Maeder, D.L., Anderson, I., Brettin, T.S., Bruce, D.C., Gilna, P., Han, C.S., Lapidus, A., Metcalf, W.W., Saunders, E., Tapia, R., Sowers, K.R.
<2>The Methanosarcina barkeri genome: comparative analysis with Methanosarcina acetivorans and Methanosarcina mazei reveals extensive  rearrangement within methanosarcinal genomes.
<3>J. Bacteriol.
<4>188
<5>7922-7931
<6>2006
<7>We report here a comparative analysis of the genome sequence of Methanosarcina barkeri with
those of Methanosarcina acetivorans and Methanosarcina mazei. The genome of M. barkeri is
distinguished by having an organization that is well conserved with respect to the other
Methanosarcina spp. in the region proximal to the origin of replication, with interspecies
gene similarities as high as 95%. However, it is disordered and marked by increased
transposase frequency and decreased gene synteny and gene density in the distal semigenome. Of
the 3,680 open reading frames (ORFs) in M. barkeri, 746 had homologs with better than 80%
identity to both M. acetivorans and M. mazei, while 128 nonhypothetical ORFs were unique
(nonorthologous) among these species, including a complete formate dehydrogenase operon, genes
required for N-acetylmuramic acid synthesis, a 14-gene gas vesicle cluster, and a
bacterial-like P450-specific ferredoxin reductase cluster not previously observed or
characterized for this genus. A cryptic 36-kbp plasmid sequence that contains an orc1 gene
flanked by a presumptive origin of replication consisting of 38 tandem repeats of a
143-nucleotide motif was detected in M. barkeri. Three-way comparison of these genomes reveals
differing mechanisms for the accrual of changes. Elongation of the relatively large M.
acetivorans genome is the result of uniformly distributed multiple gene scale insertions and
duplications, while the M. barkeri genome is characterized by localized inversions associated
with the loss of gene content. In contrast, the short M. mazei genome most closely
approximates the putative ancestral organizational state of these species.

<>

<1>Maeder, D.L., Weiss, R.B., Dunn, D.M., Cherry, J.L., Gonzalez, J.M., DiRuggiero, J., Robb, F.T.
<2>Divergence of the hyperthermophilic archaea Pyrococcus furiosus and P. horikoshii inferred from complete genomic sequences.
<3>Genetics
<4>152
<5>1299-1305
<6>1999
<7>Divergence of the hyperthermophilic Archaea, Pyrococcus furiosus and Pyrococcus horikoshii,
was assessed by analysis of complete genomic sequences of both
species. The average nucleotide identity between the genomic sequences is 70-75%
within ORFs. The P. furiosus genome (1.908 mbp) is 170 kbp larger than the P.
horikoshii genome (1.738 mbp) and the latter displays significant deletions in
coding regions, including the trp, his, aro, leu-ile-val, arg, pro, cys, thr, and
mal operons. P. horikoshii is auxotrophic for tryptophan and histidine and is
unable to utilize maltose, unlike P. furiosus. In addition, the genomes differ
considerably in gene order, displaying displacements and inversions. Six allelic
intein sites are common to both Pyrococcus genomes, and two intein insertions
occur in each species and not the other. The bacteria-like methylated chemotaxis
proteins form a functional group in P. horikoshii, but are absent in P. furiosus.
Two paralogous families of ferredoxin oxidoreductases provide evidence of gene
duplication preceding the divergence of the Pyrococcus species.

<>

<1>Maegley, K., Reich, N.O.
<2>Enhanced substrate specificity observed in EcoRI DNA methyltransferase upon mutagenesis of cysteine 223.
<3>FASEB J.
<4>6
<5>A61
<6>1992
<7>The EcoRI DNA methyltransferase is part of a type II restriction modification
system and methylates the second adenine in the recognition sequence GAATTC.
Cys223 was implicated as a critical residue by our previous protein chemical
data ((1990) J. Biol. Chem. 265,17713-17719) and by homology to other DNA
methyltransferases.  Substitution of Cys223 with Ser, Ala, and Gly by site
directed methods produced three mutant enzymes all with altered substrate
specificity.  Kinetic analysis of the mutant enzymes yields kcat and kmDNA
values for the canonical site, that do not significantly differ from those of
the wild type enzyme.  However, a significant decrease in the methylation of
non-canonical sequences is observed with the mutant enzymes both in vitro and
in vivo.  Our results suggest that either Cys223 is near the DNA binding region
or that flexibility in the region of Cys223 may be important for specificity.

<>

<1>Maegley, K., Reich, N.O.
<2>Enhanced substrate specificity observed in EcoRI DNA methyltransferase upon mutagenesis of cysteine 223.
<3>ACS Abstracts
<4>203
<5>57-BIOL
<6>1992
<7>The EcoRI DNA methyltransferase is part of a type II restriction-modification system and
methylates the second adenine in the recognition sequence GAATTC. Cys223 was implicated as a
critical residue by our previous protein chemical data (JBC, 1990, 265: 17713-17719) and by
homology to other DNA methyltransferases. Substitution of Cys223 with Ser, Ala and Gly by
site-directed methods produced three mutant enzymes all with altered substrate specificity.
Kinetic analysis of the mutant enzymes yields Kcat and KmDNA values for the cannonical site
that do not significantly differ from those of the wild type enzyme. However, a significant
decrease in the methylation of non-canonical sequences is observed with the mutant enzymes
both in vitro and in vivo. Our results suggest that either Cys223 is near the DNA binding
region or that flexibility in the region of Cys223 may be important for specificity.

<>

<1>Maegley, K., Reich, N.O.
<2>Enhanced substrate discriminiation and identification of an essential catalytic residue of the EcoRI DNA methyltransferase by site directed mutagenesis.
<3>FASEB J.
<4>7
<5>A1197
<6>1993
<7>The EcoRI DNA methyltransferase (Mtase) is part of a type II restriction modification system
and catalyzes the AdoMet dependent methylation of the ds-DNA sequence GAATTC. Cys223 was
implicated as a critical functional residue by prevous protein chemical data [(1990) J. Biol.
Chem. 265, 17713-17719] and by sequence homology. Substitution of Cys223 with Ser, Ala, and
Gly by site directed mutagenesis produced three mutant Mtases with enhanced sequence
specificity. Further characterization of the mutants has indicated possible mechanistic
reasons for the enhanced specificity. Chemical modification with DEPC has identified one
histidine residue with a pKa of approx 6, which is critical for catalysis. Site directed
mutagenesis was used to identify which histidine is critical and to better understand the role
of histidine in the chemical mechanism of the Mtase. Limited proteolysis of the Mtase in the
presence of DNA and the AdoMet analog sinefungin produced a 26 kDa (p26) and a 12 kDa (p12)
fragment [(1991) Biochemistry 30, 2940-2946]. Preliminary results suggested that the p26
fragment retained significant activity. Therefore, p26 (aa's 105-end) was genetically
produced, expressed and characterized.

<>

<1>Maegley, K., Reich, N.O.
<2>Enhanced substrate specificity observed in EcoRI DNA methyltransferase upon mutagenesis of cysteine 223.
<3>Biochemistry
<4>31
<5>2197
<6>1992
<7>The EcoRI DNA methyltransferase is part of a type-II restriction modification
system and methylates the second adenine in the recognition sequence GAATTC.
Cys223 was implicated as a critical residue by our previous protein chemical
data [1990] J. Biol. Chem. 265, 17713-17719] and by homology to other DNA
methyltransferases.  Substitution of Cys223 with Ser, Ala, and Gly by
site-directed methods produced three mutant enzymes, all with altered substrate
specificity.  Kinetic analysis of the mutant enzymes yields Kcat and KmDNA
values for the canonical site that do not significantly differ from those of
the wild type enzyme.  However, a significant decrease in the methylation of
noncanonical sequences is observed with the mutant enzymes both in vitro and in
vivo.  Our results suggest that either Cys223 is near the DNA binding region or
that flexibility in the region of Cys223 may be important for specificity.

<>

<1>Maegley, K., Reich, N.O.
<2>Enhanced substrate specificity observed in EcoRI DNA methyltransferase upon mutagenesis of cysteine 223.
<3>Biophys. J.
<4>61
<5>A61
<6>1992
<7>The EcoRI DNA methyltransferase is part of a type II restriction modification
system and methylates the second adenine in the recognition sequence GAATTC.
Cys223 was implicated as a critical residue by our previous protein chemical
data [1990] J. Biol. Chem. 265, 17713-17719) and by homology to other DNA
methyltransferases.  Substitution of Cys223 with Ser, Ala, and Gly by site
directed methods produced three mutant enzymes all with altered substrate
specificity.  Kinetic analysis of the mutant enzymes yields kcat and kmDNA
values for the canonical site, that do not significantly differ from those of
the wild type enzyme.  However, a significant decrease in the methylation of
non-canonical sequences is observed with the mutant enzymes both in vitro and
in vivo.  Our results suggest that either Cys223 is near the DNA binding region
or that flexibility in the region of Cys223 may be important for specificity.

<>

<1>Maegley, K., Shoemaker, D., Everett, B., Reich, N.O.
<2>Limited proteolysis of EcoRI DNA methylase.
<3>FASEB J.
<4>4
<5>A1793
<6>1990
<7>Limited proteolysis by soybean trpsin in the presence of various combinations
of ligands was used to probe the domain structure of the EcoRI DNA methylase
and its interactions with DNA, S-adenosyl methionine (AdoMet), S-Adenosyl
homocysteine (AdoHcy) and sinefungin (an AdoMet analog).  Proteolysis at 37C
initially results in cleavage at Lys-14 and Lys-16 followed by cleaving at
Arg-217.  The resultant fragments (14,22,22.5 Kilodaltons) are relatively
resistant to further cleavage.  This cleavage pattern was also found with
proteases of different specificities.  This data suggests a tertiary structure
consisting of two domains connected by a tether region.  In the presence of
AdoMet, AdoHcy, or DNA the 22K and 14K fragments are formed, but the rate of
proteolysis is slowed by as much as 100-fold.  Although AdoMet and sinefungin
have comparable affinities for the methylase, sinefungin shows no ability to
slow the rate of proteolysis.  Thus the methylase-sinefungin complex is
conformationally distinct from the methylase-AdoMet complex.  In the presence
of both DNA and sinefungin, proteolysis results in the cleavage of the first
105 amino acids generating fragments of 12 and 26K.  These fragments show the
greatest resistance to further proteolysis and initial results suggest that the
26K fragment is active.

<>

<1>Maegley, K., Shoemaker, D., Reich, N.O.
<2>Covalent intermediate in EcoRI DNA methyltransferase investigated by site directed mutagenesis.
<3>J. Cell. Biochem.
<4>15G
<5>175
<6>1991
<7>EcoRI DNA methyltransferase catalyses the methylation of the second adenine in
the recognition sequence GAATTC, at the N6 position.  Our protein modification
studies have implicated cysteine 223 as an important residue in the chemical
mechanism of the methyltransferase (JBC 265, 17713-17719).  Cytosine N4 and
adenine N6 methyltransferases are two unrelated classes of enzymes that
catalyze similar reactions and our analyses show homology between both types of
methyltransferases in the region of cysteine 223.  Based on these results and
the fact that the N6 of adenine is a poor nucleophile, we have proposed a
chemical mechanism involving a covalent enzyme-DNA intermediate.  Three mutants
have been constructed to test the proposed mechanism; ser223, ala223 and
gly223.  Characterization of these mutants will be described.

<>

<1>Maegley, K.A.
<2>Structural and functional characterization of the EcoRI DNA methyltransferase.
<3>Diss. Abstr.
<4>55
<5>3293B
<6>1995
<7>*
This dissertation describes continued structural and functional characterization of the EcoRI
DNA Methyltransferase (MTase). The MTase catalyzes the transfer of a methyl group from
S-adenosyl-L-methionine (AdoMet) to the N6 position of the second adenine in the DNA sequence
5' GAATTC 3'. Previously, little structural information was available on this MTase. Only
recently has a crystal structure been solved for an AdoMet-dependent enzyme. Structural
characterization of the MTase was performed using the techniques of limited proteolysis and
fluorescence spectroscopy. Site-directed mutagenesis was used to investigate the role of
Cys223 in the chemical mechanism of the MTase.

        Limited proteolysis of the MTase with trypsin suggests that the MTase is a two domain
protein consisting of a large N-terminal domain connected to a smaller C-terminal domain by a
flexible, solvent-exposed region. This "hinge" region contains part of the AdoMet binding site
and is close to two residues discussed below, Trp225 and Cys223. Structural dynamics of the
MTase were probed by performing proteolysis in the presence of the following ligands: AdoMet,
S-adenosyl-L-homocysteine (AdoHcy), sinefungin (an AdoMet analog) and DNA. All of the ligands
except sinefungin protected the Mtase from digestion, with AdoMet affording the greatest
protection (approx. 1000-fold). These results imply that there are structural differences
between the MTase-AdoMet, MTase-AdoHcy and MTase-sinefungin binary complexes.

        Fluorescence spectroscopy of the wild-type and tryptophan 183 to phenylalanine (WF183)
mutant MTases suggests that the two tryptophans within the MTase (W183 and W225) are partially
buried within the protein. Changes in the MTase emission spectrum upon AdoMet, AdoHcy,
adenosine, or adenine binding are not consistent with a ligand induced conformational change.
Therefore, the protection from proteolysis seen upon AdoMet binding is likely due to steric
obstruction of the cleavage site by the bound cofactor or by increaased rigidity of the hinge
region when AdoMet is bound.

        The fluorescence intensity of the wild-type MTase is substantially increased by the
formation of the MTase-DNA complex, whereas the fluorescence intensity of the WF183 MTase-DNA
complex decreases slightly upon DNA binding. This indicates that the fluorescence of
tryptophan 225 is greatly enhanced upon DNA binding. DNA binding may cause a conformational
change which results in the movement of tryptophan 225 away from a positively charged residue
such as arginine or lysine. Alternatively, the negatively charged phosphate backbone of the
DNA may form salt bridges with positively charged residues near tryptophan 225, thereby
causing the fluorescence intensity to increase.

        Protein modification studies with the MTase implicated Cys223 as being critical for
catalysis. Therefore, three mutant MTases were constructed, cysteine 223 to serine (CS223),
alanine (CA223), and glycine (CG223). Kinetic analysis of the mutants indicates that their
ability to methylate the canonical site remains unchanged, while their ability to methylate
closely related non-canonical sites is significantly decreased. The mutant MTases are enhanced
in their specificity. Detailed analysis of the CG223 mutant suggests that Cys223 does not
directly contact the DNA. Initial velocity and single turnover kinetics showed that a
significant portion of the enhanced specificity is due to a 5-fold decrease in the rate of
methyl transfer (kmeth). For both the wild-type and CG223 MTases kmeth is substantially faster
than kcat for canonical site methylation, whereas kmeth determines kcat for non-canonical
methylation.


<>

<1>Maegley, K.A., Gonzalez, L., Smith, D.W., Reich, N.O.
<2>Cofactor and DNA interactions in EcoRI DNA methyltransferase.
<3>J. Biol. Chem.
<4>267
<5>18527-18532
<6>1992
<7>EcoRI DNA methyltransferase contains tryptophans at positions 183 and 225. Tryptophan 225 is
adjacent to residues previously implicated in S-adenosylmethionine (AdoMet) binding and to
cysteine 223, previously shown to be the site of N-ethyl maleimide-mediated inactivation of
the enyzme (Reich,N.O, and Everett,E. (1990) J. Biol Chem. 265, 8929-8934; Everett,E.A.,
Falick, A.M., and Reich, N.O. (1990) J. Biol Chem 265, 17713-17719). The fluorescence spectra
of the wild-type enzyme is centered at 338 nm indicating partial tryptophan solvent
accessibility. Substitution of tryptophan 183 with phenylalanine results n a 45% drop in
fluorescence intensity, but no shift in lambdamax. DNA binding to the wild-type
methyltransferase caused an increase in the fluorescence intensity, while binding to the
tryptophan 183 mutant had a quenching effect, suggesting that DNA binding induces a
conformational change near both tryptophans. Binding of AdoMet and various AdoMet analogs to
the wild-type methyltransferase results in no change in the fluorescence spectrum when
excitation occurs at 295 nm, suggesting that no conformational change occurs, and AdoMet does
not interact with either tryptophan. In contrast, quenching was observed when excitation
occurred at 280 nm, suggesting that AdoMet and its analogs may be quenching tyrosine to
tryptophan energy transfer. Protein-ligand complexes were titrated with acrylamide, and the
data also implicate conformational changes upon DNA binding but not upon AdoMet binding,
consistent with previous limited proteolysis results (Reich, N.O., Maegley, K.A., Shoemaker,
D.D. and Everett, E. (1991) Biochemistry 30 2940-2946).

<>

<1>Maehara, T., Itaya, M., Ogura, M., Tanaka, T.
<2>Effect of Bacillus subtilis BsuM restriction-modification on plasmid transfer by polyethylene glycol-induced protoplast fusion.
<3>FEMS Microbiol. Lett.
<4>325
<5>49-55
<6>2011
<7>Polyethylene glycol (PEG)-induced cell fusion is a promising method to transfer larger DNA
from one cell to another than conventional genetic
DNA transfer systems. The laboratory strain Bacillus subtilis 168
contains a restriction (R) and modification (M) system, BsuM, which
recognizes the sequence 5'-CTCGAG-3'. To study whether the BsuM system
affects DNA transfer by the PEG-induced cell fusion between R+M+ and
R-M- strains, we examined transfer of plasmids pHV33 and pLS32neo
carrying no and eight BsuM sites, respectively. It was shown that
although the transfer of pLS32neo but not pHV33 from the R-M- to R+M+
cells was severely restricted, significant levels of transfer of both
plasmids from the R+M+ to R-M- cells were observed. The latter result
shows that the chromosomal DNA in the R-M- cell used as the recipient
partially survived restriction from the donor R+M+ cell, indicating
that the BsuM R-M- strain is useful as a host for accepting DNA from
cells carrying a restriction system(s). Two such examples were
manifested for plasmid transfer from Bacillus circulans and Bacillus
stearothermophilus strains to a BsuM-deficient mutant, B. subtilis
RM125.

<>

<1>Maekawa, Y., Kawakami, B.
<2>The relaxation of specificity of BanI restriction endonuclease from Bacillus aneurinolyticus IAM 1077.
<3>J. Ferment. Bioeng.
<4>69
<5>57-59
<6>1990
<7>The restriction endonuclease BanI from Bacillus aneurinolyticus IAM 1077, which
recognizes 5'-GGPyPuCC-3' and cleaves between G and G within this sequence, has
decreased substrate specificity at high nuclease concentrations.  The
relaxation of its specificity was enhanced during modified reactions:
digestion of pBR322 DNA or lambda DNA in the presence of high glycerol and
dimethyl-sulfoxide (DMSO) produced additional fragments in addition to the
inherent fragments.  Therefore, it is required to check the reaction conditions
carefully for generation of inherent fragments.

<>

<1>Maekawa, Y., Kawakami, B.
<2>Cloning and expression in Escherichia coli of the BanI and BanIII restriction-modification systems from Bacillus aneurinolyticus.
<3>J. Ferment. Bioeng.
<4>69
<5>195-198
<6>1990
<7>The genes coding for the GGPyPuCC-specific (BanI) and ATCGAT-specific (BanIII)
restriction-modification systems of Bacillus aneurinolyticus IAM1077 were
cloned and expressed in Escherichia coli using pBR322 as a vector.  The
plasmids carrying the BanI and BanIII restriction-modification genes were
designated pBanIRM8 and pBanIIIRM12, respectively.  The restriction maps of
these recombinant plasmids were constructed.  These two plasmids were stably
maintained in E. coli HB101.  However, when E. coli JM109 was used as a host,
pBanIIIRM12 was efficiently propagated but pBanIRM8 was not. The HB101 cells
carrying only the restriction gene of BanIII were viable, but the BanI
restriction gene carrier could not form colonies on agar plates.  The growth of
bacteriophage lambda was strongly restricted only in the E. coli HB101 cells
harboring pBanIRM8.  These facts indicate that the BanI restriction enzyme is
expressed and functions more efficiently than BanI modification enzyme in E.
coli.

<>

<1>Maekawa, Y., Kawakami, F., Yasukawa, H.
<2>Cloning and expression of the BanI restriction endonuclease and DNA methylase genes of Bacillus aneurinolyticus.
<3>Japanese Patent Office
<4>JP 9130679 A, JP 1991030679 A
<5>
<6>1991
<7>A DNA fragment which codes the BanI restriction endonuclease derived from Bacillus
aneurinolyticus is new. Also claimed is a DNA fragment which codes BanI methylase derived from
B. aneurinolyticus, an expression vector containing the DNA fragments, a transformant
transformed by the expression vector, prepn. of Ban I methylase, by transformation of
biological cells or microbe with the expression vector, culturing the transformant and
collecting Ban I methylase from the culture, and Ban I methylase derived from B.
aneurinolyticus and has a MW of 44000 (SDS-PAGE).

<>

<1>Maekawa, Y., Yasukawa, H., Kawakami, B.
<2>Cloning and nucleotide sequences of the BanI restriction-modification genes in Bacillus aneurinolyticus.
<3>J. Biochem. (Tokyo)
<4>107
<5>645-649
<6>1990
<7>The genes of the BanI restriction-modification system specific for GGPyPuCC were cloned from
the chromosomal DNA of Bacillus aneurinolyticus IAM107, and the coding regions were assigned
on the nucleotide sequence on the basis of the N-terminal amino acid sequences and molecular
weights of the enzymes. The restriction and modification genes coded for polypeptides with
calculated molecular weights of 39,841 and 42,637, respectively. Both the enzymes were coded
by the same DNA strand. The restriction gene was located upstream of the methylase gene,
separated by 21 bp. The cloned genes were significantly expressed in E. coli cells, so that
the respective enzymes could be purified to homogeneity. Analysis by sodium dodecyl sulfate
polyacrylamide gel electrophoresis and gel filtration indicated that the catalytically active
form of the endonuclease was dimeric and that of the methylase was monomeric. Comparison of
the amino acid sequences revealed no significant homology between the endonuclease and
methylase, though both enzymes recognize the same target sequence. Sequence comparison with
other related enzymes indicated that BanI methylase contains sequences common to
cytosine-specific methylases.

<>

<1>Magni, C., Espeche, C., Repizo, G.D., Saavedra, L., Suarez, C.A., Blancato, V.S., Espariz, M., Esteban, L., Raya, R.R., Font-de-Valdez, G., Vignolo, G., Mozzi, F., Taranto, M.P., Hebert, E.M., Nader-Macias, M.E., Sesma, F.
<2>Draft Genome Sequence of Enterococcus mundtii CRL1656.
<3>J. Bacteriol.
<4>194
<5>550
<6>2012
<7>We report the draft genome sequence of Enterococcus mundtii CRL1656, which was isolated from
the stripping milk of a clinically healthy adult
Holstein dairy cow from a dairy farm of the northwestern region of Tucuman
(Argentina). The 3.10-Mb genome sequence consists of 450 large contigs and
contains 2,741 predicted protein-coding genes.

<>

<1>Magrini, V., Salmi, D., Thomas, D., Herbert, S.K., Hartzell, P.L., Youderian, P.
<2>Temperate Myxococcus xanthus phage Mx8 encodes a DNA adenine methylase, Mox.
<3>J. Bacteriol.
<4>179
<5>4254-4263
<6>1997
<7>Temperate bacteriophage Mx8 of Myxococcus xanthus encapsidates terminally repetitious DNA,
packaged as circular permutations of its 49-kbp genome.  During both lytic and lysogenic
development, Mx8 expresses a nonessential DNA methylase, Mox, which modifies adenine residues
in occurrences of XhoI and PstI recognition sites, CTCGAG and CTGCAG, respectively, on both
phage DNA and the host chromosome.  The mox gene is necessary for methylase activity in vivo,
because an amber mutation in the mox gene abolishes activity.  The mox gene is the only phage
gene required for methylase activity in vivo, because ectopic expression of mox as part of the
M. xanthus mglBA operon results in partial methylation of the host chromosome.  The predicted
amino acid sequence of Mox is related most closely to that of the methylase involved in the
cell cycle control of Caulobacter crescentus.  We speculate that Mox acts to protect Mx8 phage
DNA against restriction upon infection of a subset of natural M. xanthus hosts.  One natural
isolate of M. xanthus, the lysogenic source of related phage Mx81, produces a restriction
endonuclease with the cleavage specificity of endonuclease BstBI.

<>

<1>Mahan, M.J., Heithoff, D.M., Low, D.A., Sinsheimer, R.L.
<2>Attenuated bacteria with altered DNA adenine methylase activity.
<3>US Patent Office
<4>US 20020076417 A
<5>
<6>2002
<7>The present invention is directed towards an attenuated strain of pathogenic bacteria (e.g.
Haemophilus, E. Coli, and/or Salmonella) having non-reverting genetic mutations relative to
the wild-type organism which alter activity of DNA adenine methylase (Dam).  The invention
further includes compositions comprised of the attenuated bacteria and methods using these
compositions to elicit an immune response to produce highly specific antibodies.  The
invention also provides methods for preparing vaccines as well as screening methods to
identify agents which may have antibacterial activity.

<>

<1>Mahan, M.J., Heithoff, D.M., Low, D.A., Sinsheimer, R.L.
<2>Reducing bacterial virulence.
<3>US Patent Office
<4>US 20020077272 A
<5>
<6>2002
<7>The virulence of bacterial strains and in particular pathogenic bacteria which infect humans
is reduced by an agent which alters the bacteria's native level or activity of DNA
methyltransferase (Dam).  The agent causes an alteration in the bacteria's native level of
methylation of adenine in a GATC tetranucleotide which inhibits virulence of the bacteria.
Thus, compounds and formulations thereof which reduce bacterial virulence inhibit
proliferation of bacteria and are useful in treating bacterial infections, particularly in
humans.

<>

<1>Mahan, M.J., Heithoff, D.M., Low, D.A., Sinsheimer, R.L.
<2>Attenuated strain of bacteria useful for eliciting immune response in an individual, has altered DNA adenine methylase activity - attenuated  bacterium adenine-methylase useful as a recombinant vaccine.
<3>US Patent Office
<4>US 20020086032
<5>
<6>2002
<7>NOVELTY - An attenuated strain (I) of bacteria, comprising altered DNA adenine methylase
(Dam)activity, such that the
bacteria are attenuated, is new. DETAILED DESCRIPTION - INDEPENDENT
CLAIMS are included for the following: (1) administering (M1) to a
subject capable of generating an immune response, a composition
comprising a pharmaceutically acceptable excipient and an immunogenic
dose of attenuated bacteria comprising altered Dam activity relative to
a Wild-type bacteria, and allowing the composition to remain in the
subject for a time and under conditions to allow the subject to
generate an immune response to the attenuated bacteria and produce
antibodies specific to the attenuated bacteria; and (2) an immunogenic
composition (II), comprising a pharmaceutically acceptable excipient,
and live bacteria comprising altered Dam activity, where the altered
activity reduces virulence relative to the bacteria with wild-type Dam
activity. WIDER DISCLOSURE - The following are also disclosed as new:
(1) immunogenic compositions comprising killed pathogenic bacteria
containing a mutation which alters Dam activity; (2) screening methods
for identifying an agent which has antibacterial activity; (3)
preparing vaccines, immunogenic compositions and mutant strains of (I);
(4) preparing attenuated bacteria or live vaccine from a virulent
pathogenic bacteria; (5) reducing bacterial virulence by contacting the
bacteria with an agent that alters Dam activity; (6) compositions for
controlling bacterial pathogenicity, comprising a compound that alters
Dam activity; (7) vaccine for provoking an immunological response in a
host to be vaccinated; and (8) kits containing one or more of the
strains and/or vaccines formulations described above, in suitable
packaging. BIOTECHNOLOGY - Preferred Method: In M1, the antibodies
generated are IgG type antibodies that are highly specific for an
antigen of the attenuated bacteria. The attenuated bacteria remain the
subject under conditions and for a period of time sufficient to allow
for B cells of the subject to undergo isotope switching and further for
the B cells to undergo cleanly expansion. The amount of antibodies
produced by the subject exceeds 150% of the amount of antibodies which
would be produced by the subject administered with the wild-type
bacteria in amount equivalent to the immunogenic dose of attenuated
bacteria. The attenuated bacteria are a genetically altered form of the
wild-type bacteria and the genetic alteration is a non-lethal,
non-reverting mutation of the pathogenic bacteria which renders the
pathogenic bacteria non-pathogenic. The bacteria are selected from
Escherichia, Vibrio, Yersinia, Salmonella and Haemophilus. The
attenuated bacteria further comprises a nucleotide sequence operatively
inserted to express a heterologous antigen e.g. antigen of a pathogenic
virus or bacteria, a mammalian tumor antigen, mammalian immune disease
antigen or a microorganism which causes an enteric infection,
respiratory infection, sexually transmitted disease, hepatitis virus
infection, fungal infection, parasitic infection or vector borne
infection, or an antigen of a microorganism selected from Leptospira
spp, Staphylococcus saprophyticus and uropathogenic E.coli. The
microorganism is selected from any one of the bacterial, fungal, viral
or parasitic organism given in the specification e.g. Enterotoxigenic
E.coli, Helicobacter pylori, Astrovirus, Campylobacter, Aspergillus
fumigatus, Ascaris lumbricoides, Blastomyces dermatitidis and Entamoeba
histolytica. Preferred Composition: The Dam activity in (II) is altered
by a heterologous nucleotide or by a mutation in Dam by reduced
expression, no expression, over expression, or expression of a form of
Dam altered from Dam native to the bacteria. Preferred Strain: (I) has
reduced or eliminated Dam activity, altered by a deletion in a dam gene
or an increase in expression of Dam. (I) is an attenuated form of
Haemophilus. ACTIVITY - None given. MECHANISM OF ACTION - Vaccine.

<>

<1>Mahan, M.J., Low, D.A.
<2>DNA methylation regulates bacterial gene expression and virulence.
<3>ASM News
<4>67
<5>356-361
<6>2001
<7>DNA adenine methylase (Dam) plays a pivotal role in bacteria such as Escherichia coli and
Salmonella - acting as a global regulator of gene expression and affecting a wide range of
critical cellular functions, including DNA replication, DNA repair, transposition, and
segregation of chromosomal DNA.  This extraordinary versatility stems from the inherent
biochemical activity of Dam.  Thus, by adding methyl groups to various sites along the
cellular DNA, Dam alters interactions of a variety of regulatory proteins with their
designated gene targets and, in the process, effectively controls expression of those genes.
In some cases, such changes modulate bacterial virulence and also serve to elicit protective
immune responses in host organisms that Salmonella or other bacterial pathogens may infect.

<>

<1>Mahato, N.K., Tripathi, C., Verma, H., Singh, N., Lal, R.
<2>Draft Genome Sequence of Deinococcus sp. Strain RL Isolated from Sediments of a Hot Water Spring.
<3>Genome Announcements
<4>2
<5>e00703-14
<6>2014
<7>Deinococcus sp. strain RL, a moderately thermophilic bacterium, was isolated from sediments of
a hot water spring in Manikaran, India. Here, we report the draft
genome (2.79 Mbp) of this strain, which contains 62 contigs and 2,614 coding DNA
sequences, with an average G+C content of 69.4%.

<>

<1>Maher, L.J. III
<2>DNA triple-helix formation: an approach to artificial gene repressors?
<3>Bioessays
<4>14
<5>807-815
<6>1992
<7>Certain sequences of double-helical DNA can be recognized and tightly bound by
oligonucleotides. The effects of such triple-helical structures
on DNA binding proteins have been studied. Stabilities of DNA
triple-helices at or near physiological conditions are sufficient to
inhibit DNA binding proteins directed to overlapping sites. Such proteins
include restriction endonucleases, methylases, transcription factors, and
RNA polymerases. These and other results suggest that
oligonucleotide-directed triple-helix formation could provide the basis
for designing artificial gene repressors. The general question of whether
biological systems employ RNA molecules for recognition and regulation of
double-helical DNA is discussed.

<>

<1>Maher, L.J., Wold, B., Dervan, P.B.
<2>Inhibition of DNA binding proteins by oligonucleotide-directed triple helix formation.
<3>Science
<4>245
<5>725-733
<6>1989
<7>Oligonucleotides that bind to duplex DNA in a sequence-specific manner by
triple helix formation offer an approach to the experimental manipulation of
sequence-specific protein binding.  Micromolar concentrations of pyrimidine
oligodeoxyribonucleotides are shown to block recognition of double helical DNA
by prokaryotic modifying enzymes and a eukaryotic transcription factor at a
homopurine target site.  Inhibition is sequence-specific.  Oligonucleotides
containing 5-methylcytosine provide substantially more efficient inhibition
than oligonucleotides containing cytosine.  The results have implications for
gene-specific repression by oligonucleotides or their analogs.

<>

<1>Maheshuwari, J.K., Buramachari, S.K., Dasshu, D., Sharma, R.
<2>A COMPUTER BASED VERSATILE METHOD FOR IDENTIFYING PROTEIN CODING DNA SEQUENCES USEFUL AS DRUG TARGETS.
<3>Japanese Patent Office
<4>JP 2007512829 A
<5>
<6>2007
<7>
<>

<1>Mahowald, M.A. et al.
<2>Characterizing a model human gut microbiota composed of members of its two dominant bacterial phyla.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>5859-5864
<6>2009
<7>The adult human distal gut microbial community is typically dominated by 2
bacterial phyla (divisions), the Firmicutes and the Bacteroidetes. Little
is known about the factors that govern the interactions between their
members. Here, we examine the niches of representatives of both phyla in
vivo. Finished genome sequences were generated from Eubacterium rectale
and E. eligens, which belong to Clostridium Cluster XIVa, one of the most
common gut Firmicute clades. Comparison of these and 25 other gut
Firmicutes and Bacteroidetes indicated that the Firmicutes possess smaller
genomes and a disproportionately smaller number of glycan-degrading
enzymes. Germ-free mice were then colonized with E. rectale and/or a
prominent human gut Bacteroidetes, Bacteroides thetaiotaomicron, followed
by whole-genome transcriptional profiling, high-resolution proteomic
analysis, and biochemical assays of microbial-microbial and microbial-host
interactions. B. thetaiotaomicron adapts to E. rectale by up-regulating
expression of a variety of polysaccharide utilization loci encoding
numerous glycoside hydrolases, and by signaling the host to produce
mucosal glycans that it, but not E. rectale, can access. E. rectale adapts
to B. thetaiotaomicron by decreasing production of its glycan-degrading
enzymes, increasing expression of selected amino acid and sugar
transporters, and facilitating glycolysis by reducing levels of NADH, in
part via generation of butyrate from acetate, which in turn is used by the
gut epithelium. This simplified model of the human gut microbiota
illustrates niche specialization and functional redundancy within members
of its major bacterial phyla, and the importance of host glycans as a
nutrient foundation that ensures ecosystem stability.

<>

<1>Maida, I., Bosi, E., Perrin, E., Papaleo, M.C., Orlandini, V., Fondi, M., Fani, R., Wiegel, J., Bianconi, G., Canganella, F.
<2>Draft Genome Sequence of the Fast-Growing Bacterium Vibrio natriegens Strain DSMZ 759.
<3>Genome Announcements
<4>1
<5>e00648-13
<6>2013
<7>Vibrio natriegens is a Gram-negative bacterium known for its extremely short doubling time.
Here we present the annotated draft genome sequence of Vibrio
natriegens strain DSMZ 759, with the aim of providing insights about its high
growth rate.

<>

<1>Mairhofer, J., Krempl, P.M., Thallinger, G.G., Striedner, G.
<2>Finished Genome Sequence of Escherichia coli K-12 Strain HMS174 (ATCC 47011).
<3>Genome Announcements
<4>2
<5>e00975-14
<6>2014
<7>Escherichia coli strain K-12 substrain HMS174 is an engineered descendant of the  E. coli K-12
wild-type strain. Like its ancestor, it is an important organism in
biotechnological research and is used in fermentation processes for heterologous
protein production. Here, we report the complete genome sequence of E. coli
HMS174 (ATCC 47011).

<>

<1>Maitra, N., Whitman, W.B., Ayyampalayam, S., Samanta, S., Sarkar, K., Bandopadhyay, C., Aftabuddin, M., Sharma, A.P., Manna, S.K.
<2>Draft Genome Sequence of the Aquatic Phosphorus-Solubilizing and -Mineralizing Bacterium Bacillus sp. Strain CPSM8.
<3>Genome Announcements
<4>2
<5>e01265-13
<6>2014
<7>Bacillus sp. strain CPSM8 is an efficient solubilizer and mineralizer of phosphorus. Here, we
present the 4.39-Mb draft genome sequence of the strain,
providing insight into the phosphorus-releasing genes related to productivity in
aquatic habitats.

<>

<1>Majid, M., Kumar, N., Qureshi, A., Yerra, P., Kumar, A., Kumar, M.K., Tiruvayipati, S., Baddam, R., Shaik, S., Srikantam, A., Ahmed, N.
<2>Genomes of Two Clinical Isolates of Mycobacterium tuberculosis from Odisha, India.
<3>Genome Announcements
<4>2
<5>e00199-14
<6>2014
<7>We report whole-genome sequences of two clinical isolates of Mycobacterium tuberculosis
isolated from patients in Odisha, India. The sequence analysis
revealed that these isolates are of an ancestral type and might represent some of
the 'pristine' isolates in India that have not admixed with other lineages.

<>

<1>Majima, T., Endo, M.
<2>Protein variants with photo-reactivity and therapeutic products containing the protein variants.
<3>Japanese Patent Office
<4>JP 2006169258 A
<5>
<6>2006
<7>
<>

<1>Majumder, K.
<2>The importance of flanking sequences in the sequence specific cleavage by the restriction endonucleases RsaI, AluI and HaeIII.
<3>Biophys. J.
<4>55
<5>48a
<6>1989
<7>Recent discoveries on the dependence in the sensitivity of some of restriction
enzymes (REs) on conformational changes in DNA indicate a complex diversity in
the nature of RE-DNA interaction.  Although the bases around these recognition
sequences are believed to be of considerable importance, not much data is
available in this regard.  We have studied the importance of flanking sequences
in the sequence specific cleavage by the REs - RsaI, AluI and HaeIII.  The
oligomers d(ACGTACGT), d(CGTACGTACG), poly d(ACGT), d(AGCTAGCT), poly d(AGCT),
d(GGCC) and d(CCGGCCGG) were used as the substrates for the relevant enzymes.
We have found that in the absence of the flanking bases the REs - RsaI, AluI
and HaeIII fail to cleave the corresponding recognition sites.  The minimum
number of such flanking bases (termed as flanking number or FN) has been found
to be two (i.e., FN=2) for RsaI.  Preliminary indications showed that the
flanking bases play an important role in the case of several other REs also.
It appears to us that in the sequence specific cleavage of DNA by RE the
flanking bases play two roles viz. (i) provide better binding and improved
kinetic stability to the enzyme-DNA complex formed prior to cleavage and (ii)
stabilization of the local active conformation by buffering out the
neighbouring influences like fraying or altered structure.

<>

<1>Mak, A.N., Lambert, A.R., Stoddard, B.L.
<2>Folding, DNA Recognition, and Function of GIY-YIG Endonucleases: Crystal Structures of R.Eco29kI.
<3>Structure
<4>18
<5>1321-1331
<6>2010
<7>The GIY-YIG endonuclease family comprises hundreds of diverse proteins and a multitude of
functions; none have been visualized bound to DNA. The
structure of the GIY-YIG restriction endonuclease R.Eco29kI has been
solved both alone and bound to its target site. The protein displays a
domain-swapped homodimeric structure with several extended surface loops
encircling the DNA. Only three side chains from each protein subunit
contact DNA bases, two directly and one via a bridging solvent molecule.
Both tyrosine residues within the GIY-YIG motif are positioned in the
catalytic center near a putative nucleophilic water; the remainder of the
active site resembles the HNH endonuclease family. The structure
illustrates how the GIY-YIG scaffold has been adapted for the highly
specific recognition of a DNA restriction site, in contrast to nonspecific
DNA cleavage by GIY-YIG domains in homing endonucleases or
structure-specific cleavage by DNA repair enzymes such as UvrC.

<>

<1>Mak, A.N.S., Fung, W.T., Kong, K.P.S., Poon, A.W.S., Ngai, S.M., Shaw, P.C.
<2>Characterization of the large subunit of EcoHK31I methyltransferase by structural modeling and mutagenesis.
<3>Biol. Chem.
<4>388
<5>265-271
<6>2007
<7>M.EcoHK31I is a naturally occurring mC5-methyltransferase with a large alpha polypeptide and a
small beta polypeptide. Polypeptide alpha
contains conserved motifs I-VIII and X, and polypeptide beta contains
motif IX. To understand how polypeptide alpha carries out its function,
a molecular model of the large domain of polypeptide a was generated
using M.Hhal and M.Haelll as templates. The large domain is a mixed
alpha/beta structure. Residues 15-19 in motif I (Phe-Naa-Gly-Naa) are
conserved for cofactor binding. The key catalytic residue Cys-79 in
motif IV is also conserved in comparison with other C-5 MTases.
Comparing polypeptide a with M.Hhal and M.Haelll revealed a unique
region upstream of motif X. To understand the role of this region, 14
charged residues between R224 and E271 in the putative small domain
were mutated. Activity assays indicated that most of these charges can
be eliminated or changed conservatively. Among these charged residues,
R224, E240, D245 and D251 may take part in proper interaction with DNA
in the presence of polypeptide beta.

<>

<1>Mak, T.N., Sfanos, K.S., Bruggemann, H.
<2>Draft Genome Sequences of Two Strains of Propionibacterium acnes Isolated from Radical Prostatectomy Specimens.
<3>Genome Announcements
<4>1
<5>e01071-13
<6>2013
<7>Propionibacterium acnes is a Gram-positive bacterium that is closely associated with various
parts of the human body, in particular with sebaceous follicles of
the skin. It has also been frequently isolated from diseased human prostates.
Here, we report draft genome sequences of two P. acnes strains, P6 and PA2,
isolated from radical prostatectomy specimens.

<>

<1>Makarova, K. et al.
<2>Comparative genomics of the lactic acid bacteria.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>15611-15616
<6>2006
<7>Lactic acid-producing bacteria are associated with various plant and animal niches and play a
key role in the production of fermented foods and
beverages. We report nine genome sequences representing the phylogenetic
and functional diversity of these bacteria. The small genomes of lactic
acid bacteria encode a broad repertoire of transporters for efficient
carbon and nitrogen acquisition from the nutritionally rich environments
they inhabit and reflect a limited range of biosynthetic capabilities that
indicate both prototrophic and auxotrophic strains. Phylogenetic analyses,
comparison of gene content across the group, and reconstruction of
ancestral gene sets indicate a combination of extensive gene loss and key
gene acquisitions via horizontal gene transfer during the coevolution of
lactic acid bacteria with their habitats.

<>

<1>Makarova, K.S., Wolf, Y.I., Koonin, E.V.
<2>Comparative genomics of defense systems in archaea and bacteria.
<3>Nucleic Acids Res.
<4>41
<5>4360-4377
<6>2013
<7>Our knowledge of prokaryotic defense systems has vastly expanded as the result of comparative
genomic analysis, followed by experimental validation. This expansion is both quantitative,
including the discovery of diverse new examples of known types of defense systems, such as
restriction-modification or toxin-antitoxin systems, and qualitative, including the discovery
of fundamentally new defense mechanisms, such as the CRISPR-Cas immunity system. Large-scale
statistical analysis reveals that the distribution of different defense systems in bacterial
and archaeal taxa is non-uniform, with four groups of organisms distinguishable with respect
to the overall abundance and the balance between specific types of defense systems. The genes
encoding defense system components in bacterial and archaea typically cluster in defense
islands. In addition to genes encoding known defense systems, these islands contain numerous
uncharacterized genes, which are candidates for new types of defense systems. The tight
association of the genes encoding immunity systems and dormancy- or cell death-inducing
defense systems in prokaryotic genomes suggests that these two major types of defense are
functionally coupled, providing for effective protection at the population level.

<>

<1>Makarova, K.S., Wolf, Y.I., Snir, S., Koonin, E.V.
<2>Defense islands in bacterial and archaeal genomes and prediction of novel defense systems.
<3>J. Bacteriol.
<4>193
<5>6039-6056
<6>2011
<7>The arms race between cellular life forms and viruses is a major driving force of evolution. A
substantial fraction of bacterial and archaeal
genomes is dedicated to antivirus defense. We analyzed the distribution of
defense genes and typical mobilome components (such as viral and
transposon genes) in bacterial and archaeal genomes, and demonstrated
statistically significant clustering of antivirus defense systems and
mobile genes and elements in genomic islands. The defense islands are
enriched in putative operons and contain numerous over-represented gene
families. A detailed sequence analysis of the proteins encoded by genes in
these families shows that many of them are diverged variants of known
defense system components, whereas others show features, such as
characteristic operonic organization, that are suggestive of novel defense
systems. Thus, genomic islands provide abundant material for experimental
study of bacterial and archaeal antivirus defense. Except for the
CRISPR-Cas systems, different classes of defense systems, in particular
toxin-antitoxin and restriction-modification systems, show non-random
clustering in defense islands. It remains unclear to what extant these
associations reflect functional cooperation between different defense
systems and to what extent the islands are genomic 'sinks' that accumulate
diverse non-essential genes, particularly those acquired via HGT. The
characteristics of defense islands resemble those of mobilome islands.
Defense and mobilome genes are non-randomly associated in islands,
suggesting non-adaptive evolution of the islands via a preferential
attachment-like mechanism underpinned by the addictive properties of
defense systems such as toxins-antitoxins and an important role of
horizontal mobility in the evolution of these islands.

<>

<1>Maki, A.H., Tsao, D.H.H.
<2>Microwave-optical study of an As(III) derivative of EcoRI methylase.
<3>Biomol. Spectroscopy II
<4>1432
<5>119-128
<6>1991
<7>We report on the formation of an unusually stable As(III)-thiolate with a
single high-affinity cysteine (Cys) of E. coli RI methylase, monitored via its
influence on a neighboring tryptophan (Trp) residue in the enzyme structure.
The binding was studied by Trp fluorescence quenching, low temperature
phosphorescence and triplet state optically detected magnetic resonance (ODMR)
of the intrinsic Trp residue(s).  The affected Trp is subject to an external
heavy atom effect (HAE) from arsenic, quenching its fluorescence and reducing
its phosphorescence lifetime from 6 sec to ca. 70 msec.  The enzyme high
affinity binding site has at least 27 times the affinity for As(III) as does a
typical sulfhydryl reagent, HSCH2CONH2.  The accessibility of the arsenical to
this Cys site was reduced upon formation of the ternary complex
methylase-DNA-sinefungin, suggesting a local conformational change in the
enzyme when DNA is bound.  The enzymatic activity assay of methylase is not
affected by the addition of a 1:1 molar ratio of the arsenical to the
methylase, but incubation with an excess of As(III) causes complete loss of
enzymatic activity.  This suggests that the high-affinity Cys residue is not a
part of the active site of the enzyme, but the addition of a molar excess of
arsenical to the enzyme derivatizes the Cys residue known to be located in the
active site.

<>

<1>Makino, K., Ishii, K., Yasunaga, T., Hattori, M., Yokoyama, K., Yutsudo, H.C., Kubota, Y., Yamaichi, Y., Iida, T., Yamamoto, K., Honda, T., Han, C.G., Ohtsubo, E., Kasamatsu, M., Hayashi, T., Kuhara, S., Shinagawa, H.
<2>Complete nucleotide sequences of 93-kb and 3.3-kb plasmids of an enterohemorrhagic Escherichia coli O157:H7 derived from Sakai outbreak.
<3>DNA Res.
<4>5
<5>1-9
<6>1998
<7>Enterohemorrhagic Escherichia coli (EHEC) O157:H7, derived from an
outbreak in Sakai city, Japan in 1996, possesses two kinds of plasmids: a
93-kb plasmid termed pO157, found in clinical EHEC isolates world-wide and
a 3.3-kb plasmid termed pOSAK1, prevalent in EHEC strains isolated in
Japan. Complete nucleotide sequences of both plasmids have been
determined, and the putative functions of the encoded proteins and the
cis-acting DNA sequences have been analyzed. pO157 shares strikingly
similar genes and DNA sequences with F-factor and the transmissible
drug-resistant plasmid R100 for DNA replication, copy number control,
plasmid segregation, conjugative functions and stable maintenance in the
host, although it is defective in DNA transfer by conjugation due to the
truncation and deletion of the required genes and DNA sequences. In
addition, it encodes several proteins implicated in EHEC pathogenicity
such as an EHEC hemolysin (HlyA), a catalase-peroxidase (KatP), a serine
protease (EspP) and type II secretion system. pOSAK1 possesses a
ColE1-like replication system, and the DNA sequence is extremely similar
to that of a drug-resistant plasmid, NTP16, derived from Salmonella
typhimurium except that it lacks drug resistance transposons.

<>

<1>Makino, K., Oshima, K., Kurokawa, K., Yokoyama, K., Uda, T., Tagomori, K., Iijima, Y., Najima, M., Nakano, M., Yamashita, A., Kubota, Y., Kimura, S., Yasunaga, T., Honda, T., Shinagawa, H., Hattori, M., Iida, T.
<2>Genome sequence of Vibrio parahaemolyticus: a pathogenic mechanism distinct from that of V cholerae.
<3>Lancet
<4>361
<5>743-749
<6>2003
<7>BACKGROUND: Vibrio parahaemolyticus, a gram-negative marine bacterium, is a worldwide cause of
food-borne gastroenteritis. V parahaemolyticus
strains of a few specific serotypes, probably derived from a common clonal
ancestor, have lately caused a pandemic of gastroenteritis. The organism
is phylogenetically close to V cholerae, the causative agent of cholera.
METHODS: The whole genome sequence of a clinical V parahaemolyticus strain
RIMD2210633 was established by shotgun sequencing. The coding sequences
were identified by use of Gambler and Glimmer programs. Comparative
analysis with the V cholerae genome was undertaken with MUMmer. FINDINGS:
The genome consisted of two circular chromosomes of 3288558 bp and 1877212
bp; it contained 4832 genes. Comparison of the V parahaemolyticus genome
with that of V cholerae showed many rearrangements within and between the
two chromosomes. Genes for the type III secretion system (TTSS) were
identified in the genome of V parahaemolyticus; V cholerae does not have
these genes. INTERPRETATION: The TTSS is a central virulence factor of
diarrhoea-causing bacteria such as shigella, salmonella, and
enteropathogenic Escherichia coli, which cause gastroenteritis by invading
or intimately interacting with intestinal epithelial cells. Our results
suggest that V parahaemolyticus and V cholerae use distinct mechanisms to
establish infection. This finding explains clinical features of V
parahaemolyticus infections, which commonly include inflammatory diarrhoea
and in some cases systemic manifestations such as septicaemia, distinct
from those of V cholerae infections, which are generally associated with
non-inflammatory diarrhoea.

<>

<1>Makino, K., Yokoyama, K., Kubota, Y., Watanabe, M., Kimura, S., Yutsudo, C.H., Kurokawa, K., Ishii, K., Hattori, M., Abe, H., Yamamoto, K., Hayashi, T., Yasunaga, T., Honda, T., Sasakawa, C., Shinagawa, H.
<2>Complete nucleotide sequence of the prophage VT2-Sakai carrying the verotoxin 2 genes of the enterohemorrhagic Escherichia coli O157:H7 derived from the Sakai outbreak.
<3>Genes Genet. Syst.
<4>74
<5>227-239
<6>1999
<7>The enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain RIMD 0509952, derived from an
outbreak in Sakai city, Japan, in 1996, produces two kinds of verotoxins, VT1 and VT2, encoded
by the stx1 and stx2 genes. In the EHEC strains, as well as in other VT-producing E. coli
strains, the toxins are encoded by lysogenic bacteriophages. The EHEC O157:H7 strain RIMD
0509952 did not produce plaque-forming phage particles upon inducing treatments. We have
determined the complete nucleotide sequence of a prophage, VT2-Sakai, carrying the stx2A and
stx2B genes on the chromosome, and presumed the putative functions of the encoded proteins and
the cis-acting DNA elements based on sequence homology data. To our surprise, the sequences in
the regions of VT2-Sakai corresponding to the early gene regulators and replication proteins,
and the DNA sequences recognized by the regulators share very limited homology to those of the
VT2-encoding 933W phage carried by the EHEC O157:H7 strain EDL933 reported by Plunkett et al.
(J. Bacteriol., p1767-1778, 181, 1999), although the sequences corresponding to the structural
components are almost identical. These data suggest that these two phages were derived from a
common ancestral phage and that either or both of them underwent multiple genetic
rearrangements. An IS629 insertion was found downstream of the stx2B gene and upstream of the
lysis gene S, and this might be responsible for the absence of plaque-forming activity in the
lysate obtained after inducing treatments.

<>

<1>Makino, O., Kawamura, F., Saito, H., Ikeda, Y.
<2>Inactivation of restriction endonuclease BamNx after infection with phage UNR2.
<3>Nature
<4>277
<5>64-65
<6>1979
<7>A host-controlled restriction-modification system is a means of protection by a
host against attacks of bacteriophages.  Recently, coliphage T3 and T7 have
been found to overcome the host-controlled restriction-modification systems of
their host strains Escherichia coli B and K, but details of the mechanism are
not well understood at present.  We have found that a Bacillus subtilis phage
UNR2rH, a mutant of UNR2, also has the ability to overcome the
restriction-modification system of B. amyloliquefaciens strain N, and detected
a BamNx inhibitor in cells infected with UNR2rH.

<>

<1>Makino, O., Saito, H., Ando, T.
<2>Bacillus subtilis-Phage Phi1 overcomes host-controlled restriction by producing BamNx inhibitor protein.
<3>Mol. Gen. Genet.
<4>179
<5>463-468
<6>1980
<7>Bacillus amyloliquefaciens N produces two restriction enzymes, BamNI and BamNX.
Subtilis-phage Phi1 is strongly restricted by BamNx.  We isolated Phi1rH, a
mutant of phage Phi1, which overcame the BamNx-restriction by producing
inhibitor.  This inhibitor inactivated BamNx specifically and reversibly.  The
inhibitor directly interacted with BamNx and the inactivation might be the
result of formation of a binary complex.  The inhibitory activity was sensitive
to treatment with trypsin.  The molecular weight of the inhibitor protein was
estimated to be approximately 20,000 daltons by gel filtration.

<>

<1>Makovets, S., Doronina, V.A., Murray, N.E.
<2>Regulation of endonuclease activity by proteolysis prevents breakage of unmodified bacterial chromosomes by type I restriction enzymes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>9757-9762
<6>1999
<7>ClpXP-dependent proteolysis has been implicated in the delayed detection of restriction
activity after the acquisition of the genes (hsdR, hsdM, and hsdS) that specify EcoKI and
EcoAI, representatives of two families of type I restriction and modification (R-M) systems.
Modification, once established, has been assumed to provide adequate protection against a
resident restriction system. However, unmodified targets may be generated in the DNA of an
hsd(+) bacterium as the result of replication errors or recombination-dependent repair. We
show that ClpXP-dependent regulation of the endonuclease activity enables bacteria that
acquire unmodified chromosomal target sequences to survive. In such bacteria, HsdR, the
polypeptide of the R-M complex essential for restriction but not modification, is degraded in
the presence of ClpXP. A mutation that blocks only the modification activity of EcoKI, leaving
the cell with approximately 600 unmodified targets, is not lethal provided that ClpXP is
present. Our data support a model in which the HsdR component of a type I restriction
endonuclease becomes a substrate for proteolysis after the endonuclease has bound to
unmodified target sequences, but before completion of the pathway that would result in DNA
breakage.

<>

<1>Makovets, S., Powell, L.M., Titheradge, A.J.B., Blakely, G.W., Murray, N.E.
<2>Is modification sufficient to protect a bacterial chromosome from a resident restriction endonuclease?
<3>Mol. Microbiol.
<4>51
<5>135-147
<6>2004
<7>It has been generally accepted that DNA modification protects the chromosome of a bacterium
encoding a restriction and modification
system. But, when target sequences within the chromosome of one such
bacterium (Escherichia coli K-12) are unmodified, the cell does not
destroy its own DNA; instead, ClpXP inactivates the nuclease, and
restriction is said to be alleviated. Thus, the resident chromosome is
recognized as 'self' rather than 'foreign' even in the absence of
modification. We now provide evidence that restriction alleviation may
be a characteristic of Type I restriction-modification systems, and
that it can be achieved by different mechanisms. Our experiments
support disassembly of active endonuclease complexes as a potential
mechanism. We identify amino acid substitutions in a restriction
endonuclease, which impair restriction alleviation in response to
treatment with a mutagen, and demonstrate that restriction alleviation
serves to protect the chromosome even in the absence of mutagenic
treatment. In the absence of efficient restriction alleviation, a Type
I restriction enzyme cleaves host DNA and, under these conditions,
homologous recombination maintains the integrity of the bacterial
chromosome.

<>

<1>Makovets, S., Titheradge, A.J.B., Murray, N.E.
<2>ClpX and ClpP are essential for the efficient acquisition of genes specifying type IA and IB restriction systems.
<3>Mol. Microbiol.
<4>28
<5>25-35
<6>1998
<7>Efficient acquisition of genes that encode a restriction and modification system with
specificities different from any already present in the recipient bacterium requires the
sequential production of the new modification enzyme followed by the restriction activity in
order that the chromosome of the recipient bacterium is protected against attack by the
restriction endonuclease.  We show that ClpX and ClpP, the components of ClpXP protease, are
necessary for the efficient transmission of the genes encoding EcoKI and EcoAI,
representatives of two families of type I R-M systems, thus implicating ClpXP in the
modulation of restriction activity.  Loss of ClpX imposed a bigger barrier than loss of ClpP,
consistent with a dual role for ClpX, possibly as a chaperone and as a component of the ClpXP
protease.  Transmission of genes specifying EcoKI was more dependent on ClpX and ClpP than
transmission of the genes for EcoAI.  Sensitivity to absence of the protease was also
influenced by the mode of gene transfer; conjugative transfer and transformation were more
dependent on ClpXP than transduction.  In the absence of either ClpX or ClpP transfer of the
EcoKI genes by P1-mediated transduction was impaired, transfer of the EcoAI genes was not.

<>

<1>Maksimenko, A., Gottikh, M.B., Kubareva, E.A., Fedorova, O.A., Shabarova, Z.A.
<2>Restriction endonucleases can be used to confirm a structure of unusual DNA duplexes.
<3>Biol. Chem.
<4>379
<5>625-630
<6>1998
<7>Cyclic and polycyclic oligonucleotides were synthesized using chemical ligation.  Two types of
catenanes with one and several intertwinings were produced.  The yield of these molecules
depended on the ligation conditions and nucleotide sequence of the ligated oligonucleotide and
its template.  Structure of ligation products was investigated and confirmed using restriction
endonuclease MvaI.  Interaction of the synthesized molecules with restriction endonucleases
SsoII, EcoRII and HindIII was also studied.

<>

<1>Makula, R.A., Meagher, R.B.
<2>A new restriction endonuclease from the anaerobic bacterium, Desulfovibrio desulfuricans, Norway.
<3>Nucleic Acids Res.
<4>8
<5>3125-3131
<6>1980
<7>The purification and characterization of a new restriction endonuclease, DdeI,
from a sulfate-reducing, anaerobic bacterium, Desulfovibrio desulfuricans,
Norway, is reported.  The enzyme recognizes the sequence 5'-C-^T-N-A-G-3'
3'-G-A-N-T-^C-5' and cleaves at the position indicated by the arrows.  The
enzyme preparation obtained is suitable for restriction mapping and ligation.

<>

<1>Malagnac, F., Gregoire, A., Goyon, C., Rossignol, J.-L., Faugeron, G.
<2>Masc2, a gene from Ascobolus encoding a protein with a DNA-methyltransferase activity in vitro, is dispensable for in vivo methylation.
<3>Mol. Microbiol.
<4>31
<5>331-338
<6>1999
<7>We have shown previously that masc1, a gene encoding a putative C5-DNA-methyltransferase, was
necessary for the de novo 'Methylation Induced Premeiotically' (MIP) process and sexual
reproduction in Ascobolus, whereas it was dispensable for maintenance methylation.  A second
MTase gene from Asobolus, masc2, encodes a protein, Masc2, which possesses the large
amino-terminal part characteristic of eukaryotic maintenance MTases.  In vitro assays have
shown that Masc2 displays a methylation activity, suggesting that it might be the MTase
responsible for maintenance methylation.  To check its function in vivo, we engineered a
disruption of the masc2 gene.  The resulting mutant strains did not exhibit any particular
phenotype during either vegetative growth or sexual reproduction.  Neither the masc2 mutation
nor the double masc1 masc2 mutation had any detectable effect upon the maintenance of the
pre-existing methylation of single gene copies previously subjected to MIP, natural
retroelement-like repeats and tandemly repeated rDNA.  The masc2 mutation did not alter either
MIP or the other de novo methylation process that operates in vegetative cells.  Nor did it
impair the meiotic process of methylation transfer.  These results suggest that at least a
third MTase gene responsible for maintenance and vegetative de novo methylation is present in
Asobolus.

<>

<1>Malagnac, F., Wendel, B., Goyon, C., Faugeron, G., Zickler, D., Rossignol, J.-L., Noyer-Weidner, M., Vollmayr, P., Trautner, T.A., Walter, J.
<2>A gene essential for de novo methylation and development in Ascobolus reveals a novel type of eukaryotic DNA methyltransferase structure.
<3>Cell
<4>91
<5>281-290
<6>1997
<7>Molecular mechanisms determining methylation patterns in eukaryotic genomes still remain
unresolved.  We have characterized, in Ascobolus, a gene for de novo methylation.  This novel
eukaryotic gene, masc1, encodes a protein that has all motifs of the catalytic domain of
eukaryotic C5-DNA-methyltransferases but is unique in that it lacks a regulatory N-terminal
domain.  The disruption of masc1 has no effect on viability or methylation maintenance but
prevents the de novo methylation of DNA repeats, which takes place after fertilization,
through the methylation induced premeiotically (MIP) process.  Crosses between parents
harboring the masc1 disruption are arrested at an early stage of sexual reproduction,
indicating that the activity of Masc1, the product of the gene, is crucial in this
developmental process.

<>

<1>Malathi, J., Murugan, N., Umashankar, V., Bagyalakshmi, R., Madhavan, H.N.
<2>Draft Genome Sequence of Multidrug-Resistant Pseudomonas aeruginosa Strain VRFPA02, Isolated from a Septicemic Patient in India.
<3>Genome Announcements
<4>1
<5>e00425-13
<6>2013
<7>Multidrug-resistant Pseudomonas aeruginosa strains, which are notable nosocomial  pathogens,
have greatly increased the mortality rate of septicemic patients due
to treatment failure. Here, we report the draft genome sequence of P. aeruginosa
strain VRFPA02, a human bloodstream isolate that has phenotypically proven to be
resistant to a broad spectrum of antibiotics.

<>

<1>Malcolm, A.D.B.
<2>Binding sites of restriction endonucleases.
<3>Biochem. Soc. Trans.
<4>14
<5>202-204
<6>1986
<7>There are now more than 600 known restriction endonucleases and these enzymes recognize more
than 100 different base sequences in their DNA substrates.  When considering binding sites we
must, of course, consider both the protein and the nucleic acid.  Many, but by no means all,
of the nucleic acid sequences recognized show a two-fold rotational axis of symmetry, for
example, BamHI recognizes GGATCCCCTAGG and HindIII cleaves at AAGCTTTTCGAA.   One might
therefore expect this symmetry to be reflected in the protein's subunit structure and
disposition of active sites.  While this probably holds in most cases, it is certainly not
universally true because some of the nucleases are active as monomers (Koncz et al., 1978). 	
Another feature worthy of comment is the predominance of G-C base-pairs in the recognition
sites.  This can be illustrated by the fact that, although many enzymes (e.g. ApaI, SacII,
BsePI, NarI, NaeI, SmaI) which recognize sequences composed entirely of GC's have been known
for some time, the first all AT-recognizing enzyme (AhaIII) was only discovered very recently
(Whitehead & Brown, 1982).  It is, of course, possible that the reasons for this bias are
evolutionary rather then mechanistic, but to date there are no useful suggestions under either
heading to account for this.

<>

<1>Malcolm, A.D.B.
<2>The use of restriction enzymes in genetic engineering.
<3>Genet. Eng. (N Y), Academic Press, R. Williamson, New York
<4>2
<5>129-173
<6>1981
<7>When the coliphage lambda is grown on E. coli strain C and then attempts are
made to grow it in E. coli K12, it grows very poorly (Bertani and Weigle, 1953.
It was shown that this was caused by a response of the host to the phage.
Since the growth of the phage was restricted, the mechanism used by the E. coli
was called a "restriction system".  Phage grown originally on K12 could be
regrown efficiently on K12, and this implies that the host also possessed a
means of protecting the phage from this restriction and the mechanism became a
"restriction-modification system".  Meselson and Yuan (1968) incubated lambda
DNA, which had been grown in E. coli K12, and lambda DNA from E. coli C with an
extract from K12.  Analysis by sedimentation through a sucrose gradient showed
that the k.K DNA was unaffected but that k.C DNA was degraded.  This
degradation was limited in extent and suggested that the restriction system
(enzyme) only hydrolysed the DNA at a small number of sites.  As it happens the
restriction enzyme from E. coli K does not cut the DNA at specific sequences
and hence has not been greatly exploited for genetic engineering. 	The first
site specific restriction enzyme was discovered by Smith and Wilcox (1970)
during their studies on recombination in Haemophilus influenzae.  They showed
by viscometry that a Haemophilus cell extract degraded DNA from the
bacteriophage P22 but had no effect on DNA from Haemophilus itself.  Using the
purified enzyme Kelly and Smith (1970) showed that the breaks produced in T7
DNA all occurred at the symmetrical sequence       5' ....G p T p Py^p Pu p A p
C....3'	       3' ....C p A p Pu p^ Py p T p G...5' where the hydrolysis occurs
at the phosphodiester bonds indicated by the arrows.  A catalogue of
restriction endonucleases isolated since then now numbers over 250 examples,
although it is not certain that all these are in fact different enzymes (Table
1).  Excellent earlier reviews are by Roberts (1976) and Modrich (1979).

<>

<1>Malcolm, A.D.B., Mofatt, J.R.
<2>Measurement of differential reactivities at restriction endonuclease sites.
<3>Biochem. Soc. Trans.
<4>8
<5>734-735
<6>1980
<7>Type II restriction endonucleases recognize and hydrolyse DNA at specific sites, often a
symmetrical four- or six-base-pair sequence.  Although the presence of the unmethylated
recognition sequence is both a necessary and sufficient condition for cleavage, it has been
recognized for some time that the rate of cleavage is not constant and presumably depends on
neighboring sequences.  Only a few attempts have so far been made to quantify these
observations, but the methods used, involving either the use of deletion mutants or analysis
of the data by a comparatively sophisticated computer analysis, may not be suitable for
routine use.

<>

<1>Malcolm, A.D.B., Moffat, J.R.
<2>Differential reactivities at restriction enzyme sites.
<3>Biochim. Biophys. Acta
<4>655
<5>128-135
<6>1981
<7>A method has been developed to measure the rates of digestion by restriction
enzymes at individual sites.  This involves a simple arithmetical treatment of
the integrated areas from a densitometer scan of an ethidium bromide stained
gel.  We have used this method to study the digestion by HpaI, HincII and SalI
of pBR322 and PhiX174 DNA, and the effect of various DNA binding ligands.  One
of the two HpaI sites in PhiX174 DNA is much more sensitive to inhibition by
ligands such as netropsin, which display a preference for AT base pairs, than
is the other site.  Inspection of the sequences flanking the restriction sites
shows that the former contains a much higher proportion of AT base-pairs than
does the latter.  The opposite phenomenon is observed with the two HincII sites
in pBR322.  This illustrates the importance of neighbouring sequences in the
interaction between restriction enzymes and their cleavage sites in DNA.

<>

<1>Malcolm, A.D.B., Moffatt, J.R., Fox, K.R., Waring, M.J.
<2>Differential inhibition of a restriction enzyme by quinoxaline antibiotics.
<3>Biochim. Biophys. Acta
<4>699
<5>211-216
<6>1982
<7>The inhibition of cleavage by HpaI at two well-defined restriction sites in
linearised PhiX174-RF DNA by quinoxaline antibiotics has been investigated.
Echinomycin, which displays a certain preference for binding to GC basepairs,
inhibits cleavage at one site much more than the other, whereas triostin A,
which displays less pronounced sequence-selectivity, inhibits both sites about
equally.  Other congeners inhibit reaction at the two sites with varying
effectiveness.  The results demonstrate the usefulness of studying inhibition
of cleavage at specific sites by restriction enzymes as a means of exploring
the specificity of DNA-ligand interactions.

<>

<1>Malcolm, A.D.B., Snounou, G.
<2>Restriction enzymes and DNA.
<3>Top. Mol. Struct. Biol.
<4>9
<5>193-222
<6>1987
<7>The discovery of restriction and modification enzymes, which proved to be a
major turning point in the progress of molecular biology, was a consequence of
a bacteriological observation in the early 1950s (Luria and Human, 1952;
Bertani and Weigle, 1953).  The two groups reported the curious behaviour of
phage grown on two different strains of bacteria.  Phages propagated on one
strain were found to grow poorly on the second and vice versa.  However, the
few phages that escaped restriction could then grow well on the new host, thus
being modifified in a way that afforded them protection from the restriction
imposed by the host.

<>

<1>Maldonado-Barragan, A., Caballero-Guerrero, B., Lucena-Padros, H., Ruiz-Barba, J.L.
<2>Genome Sequence of Lactobacillus pentosus IG1, a Strain Isolated from Spanish-Style Green Olive Fermentations.
<3>J. Bacteriol.
<4>193
<5>5605
<6>2011
<7>Lactobacillus pentosus is the most prevalent lactic acid bacterium in Spanish-style green
olive fermentations. Here we present the draft genome
sequence of L. pentosus IG1, a bacteriocin-producing strain with
biotechnological and probiotic properties isolated from this food
fermentations.

<>

<1>Maldonado-Contreras, A., Mane, S.P., Zhang, X.-S., Pericchi, L., Alarcon, T., Contreras, M., Linz, B., Blaser, M.J., Dominguez-Bello, M.G.
<2>Phylogeographic evidence of cognate recognition site patterns and transformation efficiency differences in H. pylori: theory of strain dominance.
<3>BMC Microbiol.
<4>13
<5>14
<6>2013
<7>Background: Helicobacter pylori has diverged in parallel to its human host, leading to
distinct phylogeographic populations. Recent evidence suggests that in the current human
mixing in Latin America, European H. pylori (hpEurope) are increasingly dominant at the
expense of Amerindian haplotypes (hspAmerind). This phenomenon might occur via DNA
recombination, modulated by restriction-modification systems (RMS), in which differences in
cognate recognition sites (CRS) and in active methylases will det rmine direction and
frequency of gene flow. We hypothesized that genomes from hspAmerind strains that evolved from
a small founder population have lost CRS for RMS and active methylases, promoting hpEurope's
DNA invasion. We determined the observed and expected frequencies of CRS for RMS in DNA from 7
H. pylori whole genomes and 110 multilocus sequences. We also measured the number of active
methylases by resistance to in vitro digestion by 16 restriction enzymes of genomic DNA from 9
hpEurope and 9 hspAme rind strains, and determined the direction of DNA uptake in co-culture
experiments of hspAmerind and hpEurope strains.Results: Most of the CRS were underrepresented
with consistency between whole genomes and multilocus sequences. Although neither the
frequency of CRS nor the number of active methylases differ among the bacterial populations
(average 8.6 +/- 2.6), hspAmerind strains had a restriction profile distinct from that in
hpEurope strains, with 15 recognition sites accounting for the differences. Am erindians
strains also exhibited higher transformation rates than European strains, and were more
susceptible to be subverted by larger DNA hpEurope-fragments than vice versa.Conclusions: The
geographical variation in the pattern of CRS provides evidence for ancestral differences in
RMS representation and function, and the transformation findings support the hypothesis of
Europeanization of the Amerindian strains in Latin America via DNA recombination.

<>

<1>Maldonado-Contreras, A.L., Olagibel, L., Dominguez-Bello, M.G.
<2>Geographical differences in restriction-modification system cognate recognition sites in Helicobacter pylori.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>107
<5>656
<6>2007
<7>Restriction-modification systems provide a barrier to horizontal gene transfer. Restriction
Endonucleases (RE) identify cognate DNA sequences and cleave the DNA, unless it is methylated.
H. pylori, an obligate bacterium of the human stomach, is highly recombinant. But in which
direction does recombination occurs? As human hosts mix, H. pylori strains (showing geographic
clusters) also mix, but there seems to be a shift towards predominance of European alleles in
cultured strains from Latin America. Why are Amerindian strains being lost? Are they being
subverted by recombination? We hypothesize that European strains would have lower number of
restriction sites than Amerindian strains, and an increased capability to subvert them. To
test this hypothesis, we compared the number of cognate restriction sites in the DNA from H.
pylori strains from Asian, European, African and Amerindian hosts. We analyzed in silico the
restriction profiles of DNA sequences corresponding to 7 concatenated housekeeping gene
sequences. DNA was digested in silico with 16 restriction enzymes (RE) previously reported to
be present in H. pylori strains. Phylogenetic analysis showed 4 clusters by host origin, with
Amerindian strains close to Asian strains. There was high variability in the number of cognate
RE sites within and between the phylogroups. All strains had cognate sequences for at least 13
of the 16 tested RE, and there were no significant differences in the number of cognate sites
by phylogroup (p>0.05). The average number of restriction sites was 5.3 for European strains,
5.2 for Amerindian, 5.5 for Asian and 4.9 for African strains. The results show little
differences based in the number of cognate restriction sites between the phylogroups. However
the possibility of RM differences among strain phylogroups given by the number of active
methylases cannot be assessed in silico. In vitro assays need to be performed to test for the
real susceptibility to RE digestion, which assesses the methylase functions.

<>

<1>Malfatti, S. et al.
<2>Complete genome sequence of Halogeometricum borinquense type strain (PR3).
<3>Standards in Genomic Sciences
<4>1
<5>150-159
<6>2009
<7>Halogeometricum borinquense Montalvo-Rodriguez et al. 1998 is the type species of the genus,
and is of phylogenetic interest because of its distinct location
between the halobacterial genera Haloquadratum and Halosarcina. H. borinquense
requires extremely high salt (NaCl) concentrations for growth. It can not only
grow aerobically but also anaerobically using nitrate as electron acceptor. The
strain described in this report is a free-living, motile, pleomorphic,
euryarchaeon, which was originally isolated from the solar salterns of Cabo Rojo,
Puerto Rico. Here we describe the features of this organism, together with the
complete genome sequence, and annotation. This is the first complete genome
sequence of the halobacterial genus Halogeometricum, and this 3,944,467 bp long
six replicon genome with its 3937 protein-coding and 57 RNA genes is part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Malhotra, J., Dua, A., Saxena, A., Sangwan, N., Mukherjee, U., Pandey, N., Rajagopal, R., Khurana, P., Khurana, J.P., Lal, R.
<2>Genome Sequence of Acinetobacter sp. Strain HA, Isolated from the Gut of the Polyphagous Insect Pest Helicoverpa armigera.
<3>J. Bacteriol.
<4>194
<5>5156
<6>2012
<7>In this study, Acinetobacter sp. strain HA was isolated from the midgut of a fifth-instar
larva of Helicoverpa armigera. Here, we report the draft genome
sequence (3,125,085 bp) of this strain that consists of 102 contigs, 2,911
predicted coding sequences, and a G+C content of 41%.

<>

<1>Malik, G., Dangwal, M., Kapoor, S., Kapoor, M.
<2>Role of DNA methylation in growth and differentiation in Physcomitrella patens and characterization of cytosine DNA methyltransferases.
<3>FEBS J.
<4>279
<5>4081-4094
<6>2012
<7>Epigenetic mechanisms such as DNA methylation are known to regulate important developmental
processes in higher eukaryotes. However, little
is known about the necessity and role of this process in early land
plants. Using the methyltransferase (MTase) inhibitor zebularine
(1-(beta-d-ribofuranosyl)-1,2-dihydropyrimidine-2-one), the impact of
loss of genome-wide methylation on the overall development in
Physcomitrellapatens was analyzed. It is observed that various aspects
of growth and differentiation during gametophyte development become
aberrant. A search for the core molecular components of methylation
machinery, cytosine DNA MTases, revealed the presence of seven loci in
the P.patens genome. Five of the loci code for MTases that are similar
to corresponding proteins in higher plants, while two MTases appear
specific to P.patens and are closely related to human DNMT3a and
DNMT3b, respectively. These proteins possess all the conserved
catalytic motifs characteristic of MTases and a domain of unknown
function, DUF3444. Association of these highly conserved motifs with a
DUF has not been reported in any of the MTases known so far. All the
seven genes are differentially but ubiquitously expressed in
gametophytes at low levels. Subcellular localization of GFP-fused
proteins shows patterns of distribution that can be correlated with
their putative cellular functions. This work bridges the knowledge of
MTases in P.patens and makes this simple model plant accessible for
studies on epigenetic aspects that remain intractable in higher plants.

<>

<1>Malik, S., Siezen, R.J., Renckens, B., Vaneechoutte, M., Vanderleyden, J., Lebeer, S.
<2>Draft Genome Sequence of Lactobacillus plantarum CMPG5300, a Human Vaginal Isolate.
<3>Genome Announcements
<4>2
<5>e01149-14
<6>2014
<7>The draft genome of a highly auto-aggregating Lactobacillus plantarum strain isolated from a
human vagina is reported. The peculiar phenotype also provides an
adhesive and co-aggregative potential with various pathogens, which could be of
significance in the vaginal niche. Detailed genome analysis could aid in
identifying the adhesins of the strain.

<>

<1>Malin, G., Iakobashvili, R., Lapidot, A.
<2>Effect of tetrahydropyrimidine derivatives on protein-nucleic acids interaction.
<3>J. Biol. Chem.
<4>274
<5>6920-6929
<6>1999
<7>2-Methyl-4-carboxy,5-hydroxy-3,4,5,5-tetrahydropyrimidine (THP(A) or hydroxyectoine) and
2-methyl,4-carboxy-3,4,5,6-tetrahydropyrimidine (THP(B) or ectoine) are now recognized as
ubiquitous bacterial osmoprotectants.  To evaluate the impact of tetrahydropyrimidine
derivatives on protein-DNA interaction and on restriction-modification systems, we have
examined their effect on the cleavage of plasmid DNA by 10 type II restriction endonucleases.
THP(A) completely arrested the cleavage of plasmid and bacteriophage lambda DNA by EcoRI
endonuclease at 0.4 mM and the oligonucleotide (d(CGCGAATTCGCG))2 at about 4.0 mM.  THP(B) was
10-fold less effective than THP(A), whereas for betaine and proline, a notable inhibition was
observed only at 100 mM.  Similar effects of THP(A) were observed for all tested restriction
endonucleases, except for SmaI and PvuII, which were inhibited only partially at 50mM THP(A).
No effect of THP(A) on the activity of DNase I, RNase A, and Taq DNA polymerase was noticed.
Gel shift assays showed that THP(A) inhibited the EcoRI-(d-(CGCGAATTCGCG))2 complex formation,
whereas facilitated diffusion of EcoRI along the DNA was not affected.  Methylation of the
carboxy group significantly decreased the activity of THPs, suggesting that their zwitterionic
character is essential for the inhibition effect.  Possible mechanisms of inhibition, role of
THPs in the modulation of the protein-DNA interaction, and the in vivo relevance of the
observed phenomena are discussed.

<>

<1>Mallamaci, M.A., Bascoy, M.L., Brown, J., Combates, N.J., Winkle, S.A.
<2>Locating binding sites for the carcinogen N-acetoxy-N-acetyl-2-aminofluorene using restriction enzyme inhibition assays.
<3>J. Biomol. Struct. Dyn.
<4>10
<5>83-96
<6>1992
<7>Restriction enzyme inhibition studies have been employed to map the locations of high affinity
binding sites of the carcinogen N-acetoxy-N-acetyl-2-aminofluorene (acetoxyAAF) on pBR322,
phiX174 and SV40 DNAs. Bound carcinogen levels were kept low (less than 20 bound AAF moieties
per DNA molecule) in order to observe only the binding to the high affinity sites. Inhibition
of certain restriction enzymes was observed in a limited number of locations on these DNAs.
Inhibition increased as bound AAF increased and the particular restriction enzymes inhibited
varied with location. On all three DNAs, activities of these enzymes was not affected in other
locations. Comparison of the sequences at the sites of inhibition on the three DNAs indicates
that all sites have common sequence elements: the presence of either the sequence
T(C/G)TT(G/C) or the sequence T(G/C)CTT(G/C).

<>

<1>Mallonee, R.L.
<2>Method for the determination of mutant restriction enzymes.
<3>US Patent Office
<4>US 5955369
<5>
<6>1999
<7>The present invention is directed to a method for the determination of mutant restriction
enzymes which comprises incubating restriction enzymes under non-native conditions with a
labeled double stranded oligonucleotide to a solid support to form an enzyme-oligonucleotide
complex and detecting the label to determine cleavage of the oligonucleotide by mutant
enzymes.

<>

<1>Malone, C.S., Miner, M.D., Doerr, J.R., Jackson, J.P., Jacobsen, S.E., Wall, R., Teitell, M.
<2>CmC(A/T)GG DNA methylation in mature B cell lymphoma gene silencing.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>10404-10409
<6>2001
<7>DNA methylation has been linked to gene silencing in cancer. Primary effusion lymphoma (PEL)
and myeloma are lymphoid malignancies that arise
from terminally differentiated B cells. Interestingly, PEL do not express
immunoglobulins or most B lineage-specific genes. The B cell-specific B29
(Igbeta/CD79b) gene is silenced in PEL and some myelomas but is expressed
in other normal and malignant B cells. B29 expression was reactivated in
PEL by demethylating and histone deacetylase inhibiting treatments.
Bisulfite sequencing revealed two types of DNA methylation in silenced B29
promoters: at conventional CpG and at CC(A/T)GG B29 promoter sites. The
pattern of methylated CpG ((m)CpG) and C(m)C(A/T)GG B29 promoter
methylation observed was similar to that recently reported for epigenetic
silencing of an integrated retrovirus. Methylation of C(m)C(A/T)GG sites
in the B29 promoter significantly repressed in vivo transcriptional
activity. Also, methylation of a central conserved C(m)CTGG B29 promoter
site blocked the binding of early B cell factor. This methylated motif
formed DNA-protein complexes with nuclear extracts from all cell types
examined. Therefore, C(m)C(A/T)GG methylation may represent an important
type of epigenetic marker on mammalian DNA that impacts transcription by
altering DNA-protein complex formation.

<>

<1>Malone, T., Blumenthal, R.M., Cheng, X.
<2>Structure-guided analysis reveals nine sequence motifs conserved among DNA amino-methyl-transferases, and suggests a catalytic mechanism for these enzymes.
<3>J. Mol. Biol.
<4>253
<5>618-632
<6>1995
<7>Previous X-ray crystallographic studies have revealed that the catalytic domain of a DNA
methyltransferase (Mtase) generating C5-methylcytosine bears a striking structural similarity
to that of a Mtase generating N6-methyladenine.  Guided by this common structure, we performed
a multiple sequence alignment of 42 amino-Mtases (N6-adenine and N4-cytosine).  This
comparison revealed nine conserved motifs, corresponding to the motifs I to VIII and X
previously defined in C5-cytosine Mtases.  The amino and C5-cytosine Mtases thus appear to be
more closely related than has been appreciated.  The amino Mtases could be divided into three
groups, based on the sequential order of motifs, and this variation in order may explain why
only two motifs were previously recognized in the amino Mtases.  The amino and C5-cytosine
Mtases thus appear to be more closely related than has been appreciated.  The amino Mtases
could be divided into three groups, based on the sequential order of motifs, and this
variation in order may explain why only two motifs were previously recognized in the amino
Mtases.  The Mtases grouped in this way show several other group-specific properties,
including differences in amino acid sequence, molecular mass and DNA sequence specificity.
Surprisingly, the N4-cytosine and N6-adenine Mtases do not form separate groups.  These
results have implications for the catalytic mechanisms, evolution and diversification of ths
family of enzymes.  Furthermore, a comparative analysis of the S-adenosyl-L-methionine and
adenine/cytosine binding pockets suggests that, stucturally and functionally, they are
remarkably similar to one another.

<>

<1>Maltchenko, S.
<2>The target recognition domains from the methyltransferases of II class have common subdomains.
<3>Protein Engineering Suppl.
<4>6
<5>46
<6>1993
<7>The proteins of the class II restriction-modification systems are very interesting objects for
biotechnology. Both endonuclease and methyltransferase enzymes recognize identical DNA target
sites, but they are not significantly similar in their amino acid sequences. Recently,
alignment for m5C-methyltransferases (m5C-Mets) was done and common domains (from 1 to 10) for
these proteins were identified. The target recognition domain (TRDs) for BsuPhi3T and SprI
m5C-Mets which recognize a specific DNA site were determined by site-directed mutagenesis. The
location of TRDs is between domains 8 and 9, which were determined from the alignment of
m5C-Met sequences. The size of TRDs varies from 86 to 274 amino acids. It was shown that some
of the homologous sequences are within the TRD. We analyzed TRDs from 18 m5C-Met sequences for
the presence of common subsequences and compared the subsequences with the restriction and
methyltransferase enzyme sequences.

<>

<1>Maltseva, D.V., Baykov, A.A., Jeltsch, A., Gromova, E.S.
<2>Impact of 7,8-Dihydro-8-oxoguanine on Methylation of the CpG Site by Dnmt3a (dagger).
<3>Biochemistry
<4>48
<5>1361-1368
<6>2009
<7>7,8-Dihydro-8-oxoguanine (8-oxoG) is a ubiquitous oxidative DNA lesion resulting from injury
to DNA via reactive oxygen species. 8-oxoG lesions
may play a role in the formation of aberrant DNA methylation patterns
during carcinogenesis. In this study, we assessed the effects of 8-oxoG on
methylation and complex formation of nine 30-mer oligodeoxynucleotide
duplexes by the catalytic domain of murine Dnmt3a DNA methyltransferase
(Dnmt3a-CD). The effects of 8-oxoG on the methylation rate of
hemimethylated duplexes varied from a 25-fold decrease to a 1.8-fold
increase, depending on the position of the lesion relative to the
Dnmt3a-CD recognition site (CpG) and target cytosine (C). The most
significant effect was observed when 8-oxoG replaced guanine within the
recognition site immediately downstream of the target cytosine.
Fluorescence polarization experiments with fluorescein-labeled duplexes
revealed that two molecules of Dnmt3a-CD bind per duplex, generating
sigmoid binding curves. Duplexes exhibiting the highest apparent binding
cooperativity formed the least stable 1:2 complexes with Dnmt3a-CD and
were methylated at the lowest rate. Kinetic analyses disclosed the
formation of very stable nonproductive enzyme-substrate complexes with
hemimethylated duplexes that act as suicide substrates of Dnmt3a-CD. The
presence of 8-oxoG within the CpG site downstream of the target cytosine
markedly diminished productive versus nonproductive binding. We propose
that 8-oxoG located adjacent to the target cytosine interferes with
methylation by weakening the affinity of DNA for Dnmt3a-CD, thereby
favoring a nonproductive binding mode.

<>

<1>Malygin, E.G., Evdokimov, A.A., Hattman, S.
<2>Dimeric/oligomeric DNA methyltransferases: an unfinished story.
<3>Biol. Chem.
<4>390
<5>835-844
<6>2009
<7>DNA methyltransferases (MTases) are enzymes that carry out post-replicative sequence-specific
modifications. The initial
experimental data on the structure and kinetic characteristics of the
EcoRI MTase led to the paradigm that type II systems comprise dimeric
endonucleases and monomeric MTases. In retrospect, this was logical
because, while the biological substrate of the restriction endonuclease
is two-fold symmetrical, the in vivo substrate for the MTase is
generally hemi-methylated and, hence, inherently asymmetric. Thus, the
paradigm was extended to include all DNA MTases except the more complex
bifunctional type I and type III enzymes. Nevertheless, a gradual
enlightenment grew over the last decade that has changed the accepted
view on the structure of DNA MTases. These results necessitate a more
complex view of the structure and function of these important enzymes.

<>

<1>Malygin, E.G., Evdokimov, A.A., Zinoviev, V.V., Ovechkina, L.G., Lindstrom, W.M., Reich, N.O., Schlagman, S.L., Hattman, S.
<2>A dual role for substrate S-adenosyl-L-methionine in the methylation reaction with bacteriophage T4 Dam DNA-[N6-adenine]-methyltransferase.
<3>Nucleic Acids Res.
<4>29
<5>2361-2369
<6>2001
<7>The fluorescence of 2-aminopurine (2A)-substituted duplexes (contained in the GATC target
site) was investigated by titration with T4 Dam DNA-(N6-adenine)-methyltransferase. With an
unmethylated target (2A/A duplex) or its methylated derivative (2A/mA duplex), T4 Dam produced
up to a 50-fold increase in fluorescence, consistent with 2A being flipped out of the DNA
helix. Though neither S-adenosyl-L-homocysteine nor sinefungin had any significant effect,
addition of substrate S-adenosyl-L-methionine (AdoMet) sharply reduced the Dam-induced
fluorescence with these complexes. In contrast, AdoMet had no effect on the fluorescence
increase produced with an 2A/2A double-substituted duplex. Since the 2A/mA duplex cannot be
methylated, the AdoMet-induced decrease in fluorescence cannot be due to methylation per se.
We propose that T4 Dam alone randomly binds to the asymmetric 2A/A and 2A/mA duplexes, and
that AdoMet induces an allosteric T4 Dam conformational change that promotes reorientation of
the enzyme to the strand containing the native base. Thus, AdoMet increases enzyme
binding-specificity, in addition to serving as the methyl donor. The results of
pre-steady-state methylation kinetics are consistent with this model.

<>

<1>Malygin, E.G., Hattman, S.
<2>DNA methyltransferases: Mechanistic models derived from kinetic analysis.
<3>Crit. Rev. Biochem. Mol. Biol.
<4>47
<5>97-193
<6>2012
<7>The sequence-specific transfer of methyl groups from donor S-adenosyl-L-methionine (AdoMet) to
certain positions of DNA-adenine or
-cytosine residues by DNA methyltransferases (MTases) is a major form
of epigenetic modification. It is virtually ubiquitous, except for some
notable exceptions. Site-specific methylation can be regarded as a
means to increase DNA information capacity and is involved in a large
spectrum of biological processes. The importance of these functions
necessitates a deeper understanding of the enzymatic mechanism(s) of
DNA methylation. DNA MTases fall into one of two general classes; viz.
amino-MTases and [C5-cytosine]-MTases. Amino-MTases, common in
prokaryotes and lower eukaryotes, catalyze methylation of the exocyclic
amino group of adenine ([N6-adenine]-MTase) or cytosine
([N4-cytosine]-MTase). In contrast, [C5-cytosine]-MTases methylate the
cyclic carbon-5 atom of cytosine. Characteristics of DNA MTases are
highly variable, differing in their affinity to their substrates or
reaction products, their kinetic parameters, or other characteristics
(order of substrate binding, rate limiting step in the overall
reaction). It is not possible to present a unifying account of the
published kinetic analyses of DNA methylation because different authors
have used different substrate DNAs and/or reaction conditions.
Nevertheless, it would be useful to describe those kinetic data and the
mechanistic models that have been derived from them. Thus, this review
considers in turn studies carried out with the most consistently and
extensively investigated [N6-adenine]-, [N4-cytosine]- and
[C5-cytosine]-DNA MTases.

<>

<1>Malygin, E.G., Lindstrom, W.M. Jr., Schlagman, S.L., Hattman, S., Reich, N.O.
<2>Pre-steady state kinetics of bacteriophage T4 Dam DNA-[N6-adenine] methyltransferase: interaction with native (GATC) or modified sites.
<3>Nucleic Acids Res.
<4>28
<5>4207-4211
<6>2000
<7>The DNA methyltransferase of bacteriophage T4 (T4 Dam MTase) recognizes the palindromic
sequence GATC, and catalyzes transfer of the methyl group from S-adenosyl-L-methionine
(AdoMet) to the N(6)-position of adenine [generating N(6)-methyladenine and
S-adenosyl-L-homocysteine (AdoHcy)].  Pre-steady state kinetic analysis revealed that the
methylation rate constant k(meth) for unmethylated and hemimethylated substrates (0.56 and
0.47 s(-1), respectively) was at least 20-fold larger than the overall reaction rate constant
k(cat) (0.023 s(-1)). This indicates that the release of products is the rate-limiting step in
the reaction. Destabilization of the target-base pair did not alter the methylation rate,
indicating that the rate of target nucleoside flipping does not limit k(meth). Preformed T4
Dam MTase-DNA complexes are less efficient than preformed T4 Dam MTase-AdoMet complexes in the
first round of catalysis. Thus, this data is consistent with a preferred route of reaction for
T4 Dam MTase in which AdoMet is bound first; this preferred reaction route is not observed
with the DNA-[C5-cytosine]-MTases.

<>

<1>Malygin, E.G., Lindstrom, W.M., Zinoviev, V.V., Evdokimov, A.A., Schlagman, S.L., Reich, N.O., Hattman, S.
<2>Bacteriophage T4Dam (DNA-(adenine-N6)-methyltransferase). Evidence for two distinct stages of methylation under single turnover conditions.
<3>J. Biol. Chem.
<4>278
<5>41749-41755
<6>2003
<7>We compared the (pre)steady-state and single turnover methylation kinetics of bacteriophage
T4Dam (DNA-(adenine-N6)-methyltransferase)-mediated
methyl group transfer from S-adenosyl-l-methionine (AdoMet) to
oligodeoxynucleotide duplexes containing a single recognition site
(palindrome 5'-GATC/5'-GATC) or some modified variant. T4Dam-AdoMet
functions as a monomer under steady-state conditions (enzyme/DNA << 1),
whereas under single turnover conditions (enzyme/DNA > 1), a catalytically
active complex containing two Dam-AdoMet molecules is formed initially,
and two methyl groups are transferred per duplex (to produce a methylated
duplex and S-adenosyl-l-homocysteine (AdoHcy)). We propose that the single
turnover reaction proceeds in two stages. First, two preformed
T4Dam-AdoMet complexes bind opposite strands of the unmodified target
site, and one enzyme molecule catalyzes the rapid transfer of the
AdoMet-methyl group (kmeth1 = 0.21 s-1); this is 2.5-fold slower than the
rate observed with monomeric T4Dam-AdoMet bound under pre-steady-state
conditions for burst determination. In the second stage, methyl transfer
to adenine in GATC on the complementary strand occurs at a rate that is 1
order of magnitude slower (kmeth2 = 0.023 s-1). We suggest that under
single turnover conditions, methylation of the second strand is
rate-limited by Dam-AdoHcy dissociation or its clearance from the
methylated complementary strand. The hemimethylated duplex 5'-GATC/5'-GMTC
also interacts with T4Dam-AdoMet complexes in two stages under single
turnover reaction conditions. The first stage (kmeth1) reflects
methylation by dimeric T4Dam-AdoMet productively oriented to the strand
with the adenine residue capable of methylation. The slower second stage
(kmeth2) reflects methylation by enzyme molecules non-productively
oriented to the GMTC chain, which then have to re-orient to the opposite
productive chain. Substitutions of bases and deletions in the recognition
site affect the kinetic parameters in different fashions. When the GAT
portion of GATC was disrupted, the proportion of the initial productive
enzyme-substrate complexes was sharply reduced.

<>

<1>Malygin, E.G., Ovechkina, L.G., Evdokimov, A.A., Zinoviev, V.V.
<2>Single turnover kinetics of methylation by T4 DNA-(N6-adenine)-methyltransferase.
<3>Mol. Biol. (Mosk)
<4>35
<5>65-78
<6>2001
<7>Interaction of T4 DNA-(N6-adenine)-methyltransferase was studied with a variety of synthetic
oligonucleotide substrates containing the native
recognition site GATC or its modified variants. The data obtained in
the decisecond and second intervals of the reaction course allowed for
the first time the substrate methylation rates to be compared with the
parameters of the steady-state reaction. It was established that the
substrate reaction proceeds in two stages. Because it is shown that in
steady-state conditions T4 MTase forms a dimeric structure, the
following sequence of events is assumed. Upon collision of a T4 MTase
monomer with an oligonucleotide duplex, an asymmetrical complex forms
in which the enzyme randomly oriented relative to one of the strands of
the specific recognition site catalyzes a fast transfer of the methyl
group from S-adenosylmethionine to the adenosine residue (k(1) = 0.21
s(-1)). Simultaneously, a second T4 MTase subunit is added to the
complex, providing for the continuation of the reaction. In the course
of a second stage, which is by an order of magnitude slower (k(2) =
0.023 s(-1) for duplex with the native site), the dimeric T4 MTase
switches over to the second strand and the methylation of the second
residue, target. The rate of the methyl group transfer from donor,
S-adenosylmethionine, to DNA is much higher than the overall rate of
the T4 MTase-catalyzed steady-state reaction, although this difference
is considerably less than that shown for EcoRI MTase. Base
substitutions and deletions in the recognition site affect the
substrate parameters in different fashions. When the GAT sequence is
disrupted, the proportion of the initial productive enzyme-substrate
complexes is usually sharply reduced. The flipping of the adenosine
residue to be modified in the recognition site upon interaction with
the enzyme, revealed by fluorescence titration, supports the existing
notions about the involvement of such a DNA deformation in reactions
catalyzed by various DNA-MTases.

<>

<1>Malygin, E.G., Ovechkina, L.G., Zinoviev, V.V., Linstrem, U.M., Reich, N.O.
<2>DNA-(N4-cytosine)-methyltransferase from Bacillus amyloliquefaciens: kinetic and substrate-binding properties.
<3>Mol. Biol. (Mosk)
<4>35
<5>42-51
<6>2001
<7>Interaction of DNA-(N4-cytosine)-methyltransferase from Bacillus amyloliquefaciens (BamHI
MTase, 49 kDa) with a 20-mer duplex containing
a palindromic recognition site GGATCC was studied by methods of
steady-state and pre-steady-state kinetics of the methyl group
transfer, gel retardation, and crosslinking of the enzyme subunits with
glutaraldehyde. In steady-state conditions, BamHI MTase displays a
simple kinetic behavior toward the 20-mer substrate. A linear
dependence was observed for the reaction rate on the enzyme
concentration and a Michaelis dependence of the reaction rate on the
concentration of both substrates: S-adenosyl-L-methionine (SAM), the
methyl group donor, and DNA, the methyl group acceptor. In independent
experiments, the concentration of the 20-mer duplex or SAM was changed,
the enzyme concentration being substantially lower than the
concentrations of substrates. The k(cat) values determined in these
conditions are in good agreement with one another and approximately
equal to 0.05 s(-1). The K-M values for the duplex and SAM are 0.35 and
1.6 nM, respectively. An analysis of single turnover kinetics (at
limiting concentration of the 20-mer duplex) revealed the following
characteristics of the BamHI MTase-dependent methylation of DNA. The
value of the rate constant of the DNA methylation step at the enzyme
saturating concentration is on average 0.085 s(-1), which is only 1.6
times higher than the value determined in steady-state conditions. Only
one of two target cytidine residues was methylated in a single turnover
of the enzyme, which coincides with the earlier data on EcoRI MTase.
Regardless of the order of enzyme preincubation with SAM and DNA, both
curves for the single turnover methylation are comparable. These
results are consistent with the model of the random order of the
productive ternary enzyme-substrate complex formation. In contrast to
the relatively simple kinetic behavior of BamHI MTase in the
steady-state reaction are the data on the enzyme binding with DNA. In
gel retardation experiments, there was no stoichiometrically simple
complex with the oligonucleotide duplex even at low enzyme
concentrations. The molecular mass of the complexes was so high that
they did not enter 12% PAG. In experiments on crosslinking of the BamHI
MTase subunits, it was shown that the enzyme in a free state exists as
a dimer. Introduction of substoichiometric amounts of DNA into the
reaction mixture results in pronounced multimerization of the enzyme.
However, addition of SAM in saturating concentration at an excess of
the oligonucleotide duplex over BamHI MTase converts most of the enzyme
into a monomeric state.

<>

<1>Malygin, E.G., Petrov, N.A., Gorbunov, Y.A., Kossykh, V.G., Hattman, S.
<2>Interaction of the phage T4 Dam DNA-[N6-adenine] methyltransferase with oligonucleotides containing native or modified (defective) recognition sites.
<3>Nucleic Acids Res.
<4>25
<5>4393-4399
<6>1997
<7>The DNA-[N6-adenine]-methyltransferase of phage T4 catalyzes methyl group transfer from
S-adenosyl-L-methionine to the N6-position of adenine in the palindromic sequence, GATC.  We
have used a gel shift assay to monitor complex formation between T4 Dam and various synthetic
duplex oligonucleotides, either native or modified/defective.  The results are summarized as
follows.  (i) T4 Dam bound with ~100-fold higher affinity to a 20mer specific
(GATC-containing) duplex containing the canonical palindromic methylation sequence, GATC, than
to a non-specific duplex containing another palindrome, GTAC.  (ii) Compared with the
unmethylated duplex, the hemimethylated 20mer specific duplex had a slightly increased
(~2-fold) ability to form complexes with T4 Dam.  (iii) No stable complex was formed with a
synthetic 12mer specific (GATC-containing) duplex, although T4 Dam can methylate it.  This
indicates that there is no relation between formation of a catalytically competent 12mer-Dam
complex and one stable to gel electrophoresis.  (iv) Formation of a stable complex did not
require that both strands be contiguous or completely complementary.  Absence of a single
internucleotide phosphate strongly reduced complex formation only when missing between the T
and C residues.  This suggests that if T4 Dam makes critical contact(s) with a backbone
phosphate(s), then the one between T and C is the only likely candidate.  Having only one half
of the recognition site intact on one strand was sufficient for stable complex formation
provided that the 5' G.C base-pairs be present at both ends of the palindromic, GATC.  Since
absence of either a G or C abolished T4 Dam binding, we conclude that both strands are
recognized by T4 Dam.

<>

<1>Malygin, E.G., Sclavi, B., Zinoviev, V.V., Evdokimov, A.A., Hattman, S., Buckle, M.
<2>Bacteriophage T4Dam DNA-(adenine-N-6)-methyltransferase - Comparison of pre-steady state and single turnover methylation of 40-mer duplexes  containing two (un)modified target sites.
<3>J. Biol. Chem.
<4>279
<5>50012-50018
<6>2004
<7>We analyzed pre-steady state and single turnover kinetics of bacteriophage T4Dam
DNA-(adenine-N-6)methyltransferase- mediated methyl
group transfer from S-adenosyl-L-methionine ( AdoMet) to 40-mer
duplexes containing native recognition sites (5'-GATC/5'- GATC) or some
modified variant(s). The results extend a model from studies with
single-site 20-mer duplexes. Under pre-steady state conditions,
monomeric T4Dam methyltransferase- AdoMet complexes were capable of
rapid methylation of adenine residues in 40-mer duplexes containing two
sites. During processive movement of T4Dam to the next site, the
rate-limiting step was the exchange of the product
S-adenosyl-L-homocysteine ( AdoHcy) for AdoMet without T4Dam
dissociating from the duplex. Consequently, instead of a single
exponential rate dependence, complex methylation curves were obtained
with at least two pre-steady state steps. With 40-mer duplexes
containing a single target site, the kinetics were simpler, fitting a
single exponential followed by a linear steady state phase. Single
turnover methylation of 40-mer duplexes also proceeded in two stages.
First, two dimeric T4Dam-AdoMet molecules bound, and each catalyzed a
two-step methylation. Instead of processive movement of T4Dam, a
conformational adaptation occurred. We propose that following methyl
transfer to one strand, dimeric ( T4Dam-AdoMet)-(T4Dam-AdoHcy) was
capable of rapidly reorienting itself and catalyzing methyl transfer to
the target adenine on the complementary, unmethylated strand. This
second stage methyl transfer occurred at a rate about 25-fold slower
than in the first step; it was rate-limited by Dam-AdoHcy dissociation
or its clearance from the methylated complementary strand. Under single
turnover conditions, there was complete methylation of all target
adenine residues with each of the two-site 40-mer duplexes.

<>

<1>Malygin, E.G., Zinovev, V.V.
<2>Studies of the role of symmetry in the specific recognition of natural and synthetic DNA by type II restriction and modification enzymes.
<3>Sov. Sci. Rev. D. Physiochem. Biol.
<4>9
<5>87-142
<6>1989
<7>Type II restriction endonucleases and methylases are site-specific enzymes acting on synthetic
and natural DNAs. In prokaryotic cells they make up a unique system of
restriction-modification which destroys any foreign DNA penetrating the cell. The overwhelming
majority of restrictases and methylases recognize palindromic sites with a second-order
symmetry axis. Two alternative mechanisms for this recognition exist: (i) symmetric
recognition by way of double-sided symmetrical contacts of the enzyme with non-overlapping
nucleotide groups of the duplex site; (ii) asymmetric recognition is brought about through
asymmetrical contact between the DNA chain and the enzyme molecule. The majority of
restrictases have a subunit structure, the number of subunits being even. All type II
methylases which have been isolated up to now consist of a single polypeptide chain, a monomer
being their stable form in solution. A central theme that has emerged from the data is that
multiple mechanisms exist for discrimination of base pairs. Hence, the elucidation of such
enzyme-DNA binding schemes as may be employed in sequence discrimination requires the analysis
of multiple systems. This review deals with our data on the interaction of restriction
endonucleases and Ecodam methylase with various natural and synthetic substrates containing
either completely symmetrical recognition sites or their asymmetrical variations. Our results
concerning the properties of different substrates confirm a symetrical model of interaction
between DNA and type II enzymes and demonstrate the probable interdependence of the active
centers in the cases of the BamHI and Sau3AI endonuclease just as there is with Ecodam
methylase. Dimerization of monomeric Ecodam methylase and endonuclease MvaI induced by the
oligonucleotide substrate has been demonstrated by gel filtration, ultracentrifugation and
small angle X-ray scattering methods. Thus, the catalytically active form of Ecodam methylase
and other monomeric type II enzymes seems to be a dimer. In discussing the results, attention
has mainly been paid to the significance of symmetry in the structures of both substrate DNs
and type II enzymes with respect to providing a productive interaction.

<>

<1>Malygin, E.G., Zinovev, V.V., Gorbunov, Y.A., Popov, S.G., Rechkunova, N.I., Buryanov, Y.I., Nesternko, V.F., Baev, A.A.
<2>Influence of the structure of oligonucleotide substrates on the interaction with Eco dam methylase.
<3>Biokhimiia
<4>53
<5>1639-1647
<6>1988
<7>The interaction of Eco dam methylase with synthetic substrates containing various damages in
the recognition site of the enzyme was investigated. The imperfect oligonucleotide complexes
contained a complete GATC recognition seuqence in one strand, but various defects in the
complementary strand in the recognition site: omission of one or several nucleotides, the
presence of an S-methylthiophosphate residue at the site of a break in the oligonucleotide
strand. The presence of the S-methyl-thiophosphate residue has no significant effect on
methylation in comparison with analogous substrates that did not contain an internucleotide
phosphate. A productive enzyme-substrate interaction occurred only with oligonucleotide
complexes containing both GA pairs in the recognition site of Eco dam methylase. The data
obtained suggest that Eco dam methylase, like type II restriction endonucleases, can form a
symmetrical structure in the enzyme-substrate complex.

<>

<1>Malygin, E.G., Zinoviev, V.V., Evdokimov, A.A., Lindstrom, W.M. Jr., Reich, N.O., Hattman, S.
<2>DNA (cytosine-N4-)- and -(adenine-N6-)-methyltransferases have different kinetic mechanisms but the same reaction route. A comparison of M.BamHI and T4 Dam.
<3>J. Biol. Chem.
<4>278
<5>15713-15719
<6>2003
<7>We studied the kinetics of methyl group transfer by the BamHI
DNA-(cytosine-N(4)-)-methyltransferase (MTase) from Bacillus
amyloliquefaciens to a 20-mer oligodeoxynucleotide duplex containing the
palindromic recognition site GGATCC. Under steady state conditions the
BamHI MTase displayed a simple kinetic behavior toward the 20-mer duplex.
There was no apparent substrate inhibition at concentrations much higher
than the K(m) for either DNA (100-fold higher) or S-adenosyl-l-methionine
(AdoMet) (20-fold higher); this indicates that dead-end complexes did not
form in the course of the methylation reaction. The DNA methylation rate
was analyzed as a function of both substrate and product concentrations.
It was found to exhibit product inhibition patterns consistent with a
steady state random bi-bi mechanism in which the dominant order of
substrate binding and product release (methylated DNA, DNA(Me), and
S-adenosyl-l-homocysteine, AdoHcy) was Ado-Met DNA DNA(Me) AdoHcy. The
M.BamHI kinetic scheme was compared with that for the T4 Dam
(adenine-N(6)-)-MTase. The two differed with respect to an effector action
of substrates and in the rate-limiting step of the reaction (product
inhibition patterns are the same for the both MTases). From this we
conclude that the common chemical step in the methylation reaction, methyl
transfer from AdoMet to a free exocyclic amino group, is not sufficient to
dictate a common kinetic scheme even though both MTases follow the same
reaction route.

<>

<1>Malygin, E.G., Zinoviev, V.V., Petrov, N.A., Evdokimov, A.A., Jen-Jacobson, L., Kossykh, V.G., Hattman, S.
<2>Effect of base analog substitutions in the specific GATC site on binding and methylation of oligonucleotide duplexes by the bacteriophage T4 Dam DNA-[N6-adenine] methyltransferase.
<3>Nucleic Acids Res.
<4>27
<5>1135-1144
<6>1999
<7>The interaction of the phage T4 Dam DNA-[N6-adenine] methyltransferase with 24mer synthetic
oligonucleotide duplexes having different purine base substitutions in the palindromic
recognition sequence, GATC, was investigated by means of gel shift and methyl transfer assays.
The substitutions were introduced in either the upper or lower strand: guanine by
7-deazaguanine (G-D) or 2-aminopurine (G-N) and target adenine by purine (A-P) or
2-aminopurine (A-N).  The effects of each base modification on binding/methylation were
approximately equivalent for both strands.  G-D and G-N substitutions resulted in a sharp
decrease in binary complex formation.  This suggests that T4 Dam makes hydrogen bonds with
either the N7- or O6-keto groups (or both) in forming the complex.  In contrast, A-P and A-N
substitutions were much more tolerant for complex formation.  This confirms our earlier
observations that the presence of intact 5'-G:C base pairs at both ends of the methylation
site is critical, but that base substitutions within the central A:T base pairs show less
inhibition of complex formation.  Addition of T4 Dam to a complete substrate mixture resulted
in a burst of [3H]methylated product.  In all cases the substrate dependencies of bursts and
methylation rates were proportional to each other.  For the perfect 24mer kcat=0.014/s and
Km=7.7 nM was obtained.  In contrast to binary complex formation the two guanine substitutions
exerted relatively minor effects on catalytic turnover (the kcat was reduced at most
2.5-fold), while the two adenine substitutions showed stronger effects (5- to 15-fold
reduction in kcat).  The effects of base analog substitutions on Km(DNA) were more variable:
A-P (decreased); A-N and G-D (unchanged); G-N (increased).

<>

<1>Malyguine, E., Vannier, P., Yot, P.
<2>Alteration of the specificity of restriction endonucleases in the presence of organic solvents.
<3>Gene
<4>8
<5>163-177
<6>1980
<7>The specificity of XbaI, SalI, HhaI, PstI, BamHI and SstI is relaxed in the
presence of an organic solvent, such as 20% glycerol or 8% dimethylsulfoxide
(DMSO).  This alteration, very pronounced in some cases, requires an excess of
enzyme, varies from one kind of enzyme to another and is highly dependent on
the pH, the ionic strength, the nature of the metallic cofactor and/or the
presence of a second organic solvent.  Preliminary data concerning XbaI and
BamHI used under conditions where the relaxation of specificity is moderate,
suggest that some of the new ("pseudo") sites correspond to shortened sequences
derived from the normal recognition sequence cleaved under the standard
conditions of the reaction.

<>

<1>Mamelak, L., Boyer, H.W.
<2>Genetic control of the secondary modification of deoxyribonucleic acid in Escherichia coli.
<3>J. Bacteriol.
<4>104
<5>57-62
<6>1970
<7>The wild-type restriction and modification alleles of Escherichia coli K-12 and
B were found to have no measurable effect on the patterns of methylated bases
in the deoxyribonucleic acid (DNA) of these strains.  The genetic region
controlling the methylation of cytosine in E. coli K-12 was mapped close to
his, and the presence or absence of this gene in E. coli B or E. coli K had no
effect on the restriction and modification properties of these strains.  Thus,
only a few of the methylated bases in the DNA of these strains are involved in
host modification, and the biological role of the remainder remains obscure.

<>

<1>Manachini, P.L., Parini, C., Fortina, M.G., Benazzi, L.
<2>BliI, a restriction endonuclease from Bacillus licheniformis.
<3>FEBS Lett.
<4>214
<5>305-307
<6>1987
<7>From Bacillus licheniformis a site-specific restriction endonuclease, named
BliI, has been purified and characterized.  BliI was able to digest lambda DNA
at pH 9.1 over a wide temperature range (25-65C).  Digestion of lambda and
PhiX174 DNAs with BliI produced banding patterns identical to those seen with
HaeIII.  Therefore, BliI and HaeIII endonucleases are isoschizomers.

<>

<1>Manakova, E., Golovenko, D., Grazulis, S., Zaremba, M., Siksnys, V.
<2>Structural mechanism of cognate DNA recognition by the BfiI restriction enzyme.
<3>FEBS J.
<4>279
<5>446-447
<6>2012
<7>Restriction endonuclease BfiI recognizes and cleaves the asymmetric DNA sequence
5'-ACTGGG-3' downstream of the site independently of Mg-ions.  BfiI is evolved as a fusion
of two domains: the C-terminal DNA binding domain similar to DNHA binding domans of EcoRII and
B3 family of plant transcription factors, and the N-terminal catalytic domain, which is
similar to the Nuc nuclease of S. thyphimurium.  We have solved the crystal structure of the
BfiI DNA binding domjain in complex with the specific DNA to reveal the mechanism of the
specific DNA recognition. Superposition of the apoBfiI structure with the DNA bound C-domain
suggests that BfiI must change its conformation to accommopdate the scissile phosphate in the
active site.  To get a glimpse on the structural changes occurring upon BfiI binding to
cognate DNA we have performed the X-ray small angle scatterinhg measurements of apo and DNA
bound BfiI.  Ab initio shape determination as well as rigid body modeling using the
crystallographic data suggest that apo BfiI retains the similar conformation in the solution
as in a crystal, whereas DNA bound BfiI shows a conformational flexibility.  The truncated
heterodimar of BfiI which lacks one of the two DNHA-bidning domains was constructed in order
to simplify the system for SAXS experiments.

<>

<1>Manakova, E., Grazulis, S., Zaremba, M., Tamulaitiene, G., Golovenko, D., Siksnys, V.
<2>Structural mechanisms of the degenerate sequence recognition by Bse634I restriction endonuclease.
<3>Nucleic Acids Res.
<4>40
<5>6741-6751
<6>2012
<7>Restriction endonuclease Bse634I recognizes and cleaves the degenerate DNA sequence
5'-R/CCGGY-3' (R stands for A or G; Y for T or C, '/' indicates a
cleavage position). Here, we report the crystal structures of the Bse634I R226A
mutant complexed with cognate oligoduplexes containing ACCGGT and GCCGGC sites,
respectively. In the crystal, all potential H-bond donor and acceptor atoms on
the base edges of the conserved CCGG core are engaged in the interactions with
Bse634I amino acid residues located on the alpha6 helix. In contrast, direct
contacts between the protein and outer base pairs are limited to van der Waals
contact between the purine nucleobase and Pro203 residue in the major groove and
a single H-bond between the O2 atom of the outer pyrimidine and the side chain of
the Asn73 residue in the minor groove. Structural data coupled with biochemical
experiments suggest that both van der Waals interactions and indirect readout
contribute to the discrimination of the degenerate base pair by Bse634I.
Structure comparison between related enzymes Bse634I (R/CCGGY), NgoMIV (G/CCGGC)
and SgrAI (CR/CCGGYG) reveals how different specificities are achieved within a
conserved structural core.

<>

<1>Manavalan, P., Johnson, W.C. Jr., Modrich, P.
<2>Prediction of secondary structure for EcoRI endonuclease.
<3>J. Biol. Chem.
<4>259
<5>11666-11667
<6>1984
<7>The circular dichroism of EcoRI restriction endonuclease was measured to 178 nm
and analyzed for secondary structure.  The results (33% alpha-helix, 25%
beta-sheet, 17% turns, and 25% other structures) compare well with our joint
prediction from sequence data.

<>

<1>Mancini, S., Abicht, H.K., Karnachuk, O.V., Solioz, M.
<2>Genome Sequence of Desulfovibrio sp. A2, a Highly Copper Resistant, Sulfate-Reducing Bacterium Isolated from Effluents of a Zinc Smelter at  the Urals.
<3>J. Bacteriol.
<4>193
<5>6793-6794
<6>2011
<7>Desulfovibrio sp. A2 is an anaerobic Gram-negative sulfate-reducing bacterium with remarkable
tolerance to copper. It was isolated from
wastewater effluents of a zinc smelter at the Urals. Here, we report the
4.2-Mb draft genome sequence of Desulfovibrio sp. A2 and identify
potential copper resistance mechanisms.

<>

<1>Mandape, S.N., Marshall, D.R., Dent, L.L., Pratap, S.
<2>Draft Genome Sequence of Multidrug-Resistant Acinetobacter baumannii Strain MMC4, Isolated from a Patient in Tennessee.
<3>Genome Announcements
<4>2
<5>e00051-14
<6>2014
<7>Acinetobacter baumannii multidrug-resistant strain MMC4 was isolated from a bronchoalveolar
lavage fluid sample from a patient in Nashville, TN, USA. Here,
we report a draft genome sequence with a size of 3,985,367 bp, an average G+C
content of 39.8%, and 3,863 predicted protein-coding sequences.

<>

<1>Mane, S.P. et al.
<2>Host-interactive genes in amerindian Helicobacter pylori diverge from their old world homologs and mediate inflammatory responses.
<3>J. Bacteriol.
<4>192
<5>3078-3092
<6>2010
<7>Helicobacter pylori is the dominant member of the gastric microbiota and
has been associated with an increased risk of gastric cancer and peptic
ulcers in adults. H. pylori populations have migrated and diverged with
human populations, and health effects vary. Here, we describe the whole
genome of the cag-positive strain V225d, cultured from a Venezuelan Piaroa
Amerindian subject. To gain insight into the evolution and host adaptation
of this bacterium, we undertook comparative H. pylori genomic analyses. A
robust multiprotein phylogenetic tree reflects the major human migration
out of Africa, across Europe, through Asia, and into the New World,
placing Amerindian H. pylori as a particularly close sister group to East
Asian H. pylori. In contrast, phylogenetic analysis of the
host-interactive genes vacA and cagA shows substantial divergence of
Amerindian from Old World forms and indicates new genotypes (e.g., VacA
m3) involving these loci. Despite deletions in CagA EPIYA and CRPIA
domains, V225d stimulates interleukin-8 secretion and the hummingbird
phenotype in AGS cells. However, following a 33-week passage in the mouse
stomach, these phenotypes were lost in isolate V225-RE, which had a 15-kb
deletion in the cag pathogenicity island that truncated CagA and
eliminated some of the type IV secretion system genes. Thus, the unusual
V225d cag architecture was fully functional via conserved elements, but
the natural deletion of 13 cag pathogenicity island genes and the
truncation of CagA impaired the ability to induce inflammation.

<>

<1>Manfredi, P., Pagni, M., Cornelis, G.R.
<2>Complete Genome Sequence of the Dog Commensal and Human Pathogen Capnocytophaga canimorsus Strain 5.
<3>J. Bacteriol.
<4>193
<5>5558-5559
<6>2011
<7>Capnocytophaga canimorsus is a commensal Gram-negative bacterium, originally isolated from a
dog's mouth, that causes septicemia in humans.
C. canimorsus has the unusual ability to feed on host cells, including
phagocytes. This capacity depends on surface-exposed glycan-foraging
systems. Here we present the first complete genome sequence of a C.
canimorsus strain (Cc5).

<>

<1>Mangiamele, P., Nicholson, B., Wannemuehler, Y., Seemann, T., Logue, C.M., Li, G., Tivendale, K.A., Nolan, L.K.
<2>Complete Genome Sequence of the Avian Pathogenic Escherichia coli Strain APEC O78.
<3>Genome Announcements
<4>1
<5>e0002613
<6>2013
<7>Colibacillosis, caused by avian pathogenic Escherichia coli (APEC), is a significant disease,
causing extensive animal and financial losses globally.
Because of the significance of this disease, more knowledge is needed regarding
APEC's mechanisms of virulence. Here, we present the fully closed genome sequence
of a typical avian pathogenic E. coli strain belonging to the serogroup O78.

<>

<1>Manichanh, C., Chapple, C.E., Frangeul, L., Gloux, K., Guigo, R., Dore, J.
<2>A comparison of random sequence reads versus 16S rDNA sequences for estimating the biodiversity of a metagenomic library.
<3>Nucleic Acids Res.
<4>36
<5>5180-5188
<6>2008
<7>The construction of metagenomic libraries has permitted the study of
microorganisms resistant to isolation and the analysis of 16S rDNA
sequences has been used for over two decades to examine bacterial
biodiversity. Here, we show that the analysis of random sequence reads
(RSRs) instead of 16S is a suitable shortcut to estimate the biodiversity
of a bacterial community from metagenomic libraries. We generated 10,010
RSRs from a metagenomic library of microorganisms found in human faecal
samples. Then searched them using the program BLASTN against a prokaryotic
sequence database to assign a taxon to each RSR. The results were compared
with those obtained by screening and analysing the clones containing 16S
rDNA sequences in the whole library. We found that the biodiversity
observed by RSR analysis is consistent with that obtained by 16S rDNA. We
also show that RSRs are suitable to compare the biodiversity between
different metagenomic libraries. RSRs can thus provide a good estimate of
the biodiversity of a metagenomic library and, as an alternative to 16S,
this approach is both faster and cheaper.

<>

<1>Maniloff, J., Dybvig, K., Sladek, T.L.
<2>Mycoplasma DNA Restriction and Modification.
<3>Mycoplasmas: Molecular Biology and Pathogenesis, American Society for Microbiology, Maniloff, J., Rochester, NY
<4>0
<5>325-330
<6>1992
<7>*
Mycoplasma DNA Restriction and Modification
Introduction
Restriction and Modification in Eubacteria
Restriction-Modification Systems
Restriction and Modification in Mycoplasmas
Mycoplasma Restriction-modification systems
A. laidlawii K2
A. laidlawii JA1
A. laidlawii JA1 Restriction and recA Mutants
S. citri ASP2
M. fermentans
U. urealyticum 960
Mycoplasma methylases
Restriction-modification systems and Mycoplasma Evolution
Acknowledgments
References


<>

<1>Manivasakam, P., Aubrecht, J., Sidhom, S., Schiestl, R.H.
<2>Restriction enzymes increase efficiencies of illegitimate DNA integration but decrease homologous integration in mammalian cells.
<3>Nucleic Acids Res.
<4>29
<5>4826-4833
<6>2001
<7>Mammalian cells repair DNA double-strand breaks by illegitimate end-joining or by homologous
recombination. We investigated the effects
of restriction enzymes on illegitimate and homologous DNA integration
in mammalian cells. A plasmid containing the neoR expression cassette,
which confers G418 resistance, was used to select for illegitimate
integration events in CHO wild-type and xrcc5 mutant cells.
Co-transfection with the restriction enzymes BamHI, Bglll, EcoRI and
Kpnl increased the efficiency of linearized plasmid integration up to
5-fold in CHO cells. In contrast, the restriction enzymes did not
increase the integration efficiency in xrcc5 mutant cells. Effects of
restriction enzymes on illegitimate and homologous integration were
also studied in mouse embryonic stem (ES) cells using a plasmid
containing the neoR gene flanked by exon 3 of Hprt. The enzymes BamHI,
Bglll and EcoRI increased the illegitimate integration efficiency of
transforming DNA several-fold, similar to the results for CHO cells.
However, all three enzymes decreased the absolute frequency of
homologous integration apprx2-fold, and the percentage of homologous
integration decreased >10-fold. This suggests that random DNA breaks
attract illegitimate recombination (IR) events that compete with
homology search.

<>

<1>Manivasakam, P., Schiestl, R.H.
<2>Nonhomologous end joining during restriction enzyme-mediated DNA integration in Saccharomyces cerevisiae.
<3>Mol. Cell. Biol.
<4>18
<5>1736-1745
<6>1998
<7>The BamHI restriction enzyme mediates integration of nonhomologous DNA into the Saccharomyces
cerevisiae genome (R. H. Schiestl and T. D. Petes,
Proc. Natl. Acad. Sci. USA 88:7585-7589, 1991). The present study
investigates the mechanism of such events: in particular, the mediating
activity of various restriction enzymes and the processing of resultant
fragment ends. Our results show that in addition to BamHI, BglII and KpnI
increase DNA integration efficiencies severalfold, while Asp718, HindIII,
EcoRI, SalI, SmaI, HpaI, MscI, and SnaBI do not. Secondly, the three
active enzymes stimulated integrations only of fragments containing 5' or
3' overhangs but not of blunt-ended fragments. Thirdly, integrations
mediated by one enzyme and utilizing a substrate created by another
required at least 2 bp of homology. Furthermore, an Asp718 fragment
possessing a 5' overhang integrated into a KpnI (isoschizomer) site
possessing a 3' overhang, most likely by filling of the 5' overhang
followed by 5' exonuclease digestion to produce a 3' end. We classified
and analyzed the restriction enzyme-mediated integration events in the
context of their genomic positions. The majority of events integrated into
single sites. In the remaining 6 of 19 cases each end of the plasmid
inserted into a different sequence, producing rearrangements such as
duplications, deletions, and translocations.

<>

<1>Mann, A.J., Hahnke, R.L., Huang, S., Werner, J., Xing, P., Barbeyron, T., Huettel, B., Stueber, K., Reinhardt, R., Harder, J., Gloeckner, F.O., Amann, R.I., Teeling, H.
<2>The genome of the algae-associated marine flavobacterium Formosa agariphila KMM 3901T reveals a broad potential for the degradation of algal polysaccharides.
<3>Appl. Environ. Microbiol.
<4>79
<5>6813
<6>2013
<7>In recent years, representatives of the Bacteroidetes have been increasingly recognized as
specialists for the degradation of macromolecules. Formosa constitutes a Bacteroidetes genus
within the class Flavobacteria, whose members have been found in marine habitats with high
levels of organic matter, such as an association with algae, invertebrates and fecal pellets.
Here we report on the generation and analysis of the genome of the type strain of Formosa
agariphila(KMM 3901T) - an isolate from the green algae Acrosiphonia sonderi.  F. agariphila
is a facultative anaerobe with the capacity for mixed acid fermentation and denitrification.
Its genome harbors 129 proteases and 88 glycoside hydrolases, indicating a pronounced
specialization on the degradation of protein, polysaccharides and glycoproteins.  65 of the
glycoside hydrolases are organized in at least 13 distinct polysaccharide utilization loci,
where they are clustered with TonB-dependent receptors, SusD-like proteins,
sensors/transcription factors, transporters and oftentimes sulfatases.  These loci play a
pivotal role in bacteroidetal polysaccharide biodegradation, and in the case of F. agariphila
revealed the capacity to degrade a wide range of algal polysaccharides from green, red and
brown algae and thus a strong specialization of towards an algae-associated lifestyle.  This
was corroborated by growth experiments, which confirmed usage of particularly those
monosaccharides that constitute the building blocks of abundant agal polysaccharides as well
as of distinct algal polysaccharides such as laminarins, xylans and k-carrageenans.

<>

<1>Mann, M.B.
<2>The cloned HhaII restriction-modification system.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>229-237
<6>1981
<7>I. Procedure for cloning restriction-modification (R-M) systems.  II. A clone that
overproduces the HhaII R-M gene products.
III. Biologic properties of the cloned HhaII R-M system.
IV. The problem of site-specific DNA-protein interaction.
V. HhaII*, a form of HhaII with grossly altered site specificity.

<>

<1>Mann, M.B., Rao, R.N., Smith, H.O.
<2>Cloning of restriction and modification genes in E. coli: the HhaII system from Haemophilus haemolyticus.
<3>Gene
<4>3
<5>97-112
<6>1978
<7>The genes for a Class II restriction-modification system (HhaII) from
Haemophilus haemolyticus have been cloned in Escherichia coli.  The vector used
for cloning was plasmid pBR322 which confers resistance to tetracycline and
ampicillin and contains a single endonuclease R-PstI site,
(5')C-T-G-C-A7-G(3'), in the ampicillin gene.  The procedure developed by
Bolivar et al. (1977) was used to form DNA recombinants.  H. haemolyticus DNA
was cleaved with PstI endonuclease and poly(dC) extensions were added to the
3'-OH termini using terminal deoxynucleotidyl transferase.  Circular pBR322 DNA
was cleaved to linear molecules with PstI endonuclease and poly(dG) extensions
were added to the 3'-OH termini, thus regenerating the PstI cleavage site
sequence.  Recombinant molecules, formed by annealing the two DNAs, were used
to transfect a restriction and modification-deficient strain of E. coli (HB101
r-m-recA).  Tetracycline-resistant clones were tested for acquisition of
restriction phenotype (as measured by growth on plates seeded with phage
lambdacI-0).  A single phage-resistant clone was found.  The recombinant
plasmid, pDI10, isolated from this clone, had acquired 3 kilobases of
additional DNA which could be excised with PstI endonuclease.  In addition to
the restriction function, cells carrying the plasmid expressed the HhaII
modification function.  Both activities have been partially purified by
single-stranded DNA-agarose chromatography.  The cloned HhaII restriction
activity yields cleavage patterns identical to HinfI. A restriction map of the
cloned DNA segment is presented.

<>

<1>Mann, M.B., Smith, H.O.
<2>Specificity of DNA Methylases from Haemophilus Sp.
<3>Proceedings of the Conference on Transmethylation, Elsevier North Holland, Usdin, E., Borchardt, R.T., Creveling, C.R., New York
<4>0
<5>483-492
<6>1979
<7>We have studied the specificity of three DNA cytosine methylases found in
Haemophilus sp., which have the common property of acting at tetranucleotide
sites containing cytosine and guanine residues.  The methylases belong to the
three restriction and modification systems:  HhaI, HpaII, and HaeIII, for which
the restriction endonuclease recognition sites are (5') pG-C-G-C, (5')
pC-C-G-G, and (5') pG-G-C-C.  The sequence specificity and position of cytosine
methylation within the recognition sequence for M.HpaII and M.HaeIII have been
previously reported as (5') pC-CmC.1  Here we present comparable data for
M.HhaI.  In addition, we determine the ability of each of the three methylases
to utilize as substrates a variety of polymers containing base analogues.
Judging from their behaviour on these substrates, we conclude that the
mechanism for discriminating a given base pair varies from one enzyme to
another.

<>

<1>Mann, M.B., Smith, H.O.
<2>Size of 5'-terminal fragments cleaved from poly(dG-dC) by EndoR.HhaI.
<3>Nucleic Acid - Protein Recognition, Academic Press, Vogel, H.J., New York
<4>0
<5>269-271
<6>1977
<7>None

<>

<1>Mann, M.B., Smith, H.O.
<2>Specificity of HpaII and HaeIII DNA methylases.
<3>Nucleic Acids Res.
<4>4
<5>4211-4221
<6>1977
<7>The methylases M HaeIII and M HpaII recognize the tetranucleotide sequences
(5') G/C-G/C*-C*/G-C/G (5') and (5') C/G-C*/G-G/C*-G/C (5') respectively, in
DNA, and transfer a methyl group from S-adenosylmethionine to the 5-position of
cytosine on each strand as indicated by the asterisks.  Restriction
endonuclease R HaeIII does not cleave the methylated sequence G/C-G/Cm-Cm/G-C/G
but can cleave G/Cm-G/C-C/G-Cm/G in which methylation is introduced on the
unnatural external cytosine positions.  Similarly, R HpaII does not cleave
C/G-Cm/G-G/Cm-G/C but can cleave Cm/G-C/G-G/C-G/C.

<>

<1>Mannarelli, B.M., Balganesh, T.S., Greenberg, B., Springhorn, S.S., Lacks, S.A.
<2>Nucleotide sequence of the DpnII DNA methylase gene of Streptococcus pneumoniae and its relationship to the dam gene of Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>82
<5>4468-4472
<6>1985
<7>The structural gene (dpnM) for the DpnII DNA methylase of Streptococcus pneumoniae, which is
part of the DpnII restriction system and methylates adenine in the sequence 5'-G-A-T-C-3',
was identified by subcloning fragments of a chromosomal segment from a DpnII-producing strain
in an S. pneumoniae host/vector cloning system and demonstrating function of the gene also in
Bacillus subtilis. Determination of the nucleotide sequence of the gene and adjacent DNA
indicates that it encodes a polypeptide of 32,903 daltons. A putative promoter for
transcription of the gene lies within a hundred nucleotides of the polypeptide start codon.
Comparison of the coding sequence to that of the dam gene of Escherichia coli, which encodes a
similar methylase, revealed 30% of the amino acid residues in the two enzymes to be identical.
This homology presumably reflects a common origin of the two genes prior to the divergence of
Gram-positive and Gram-negative bacteria. it is suggested that the restriction function of the
gene is primitive, and that the homologous restriction system in E. coli has evolved to play
an accessory role in heteroduplex DNA base mismatch repair.

<>

<1>Mannino, S.J., Jenkins, C.L., Raines, R.T.
<2>Chemical mechanism of DNA cleavage by the homing endonuclease I-PpoI.
<3>Biochemistry
<4>38
<5>16178-16186
<6>1999
<7>Homing endonucleases are distinguished by their ability to catalyze the cleavage of
double-stranded DNA with extremely high specificity. I-PpoI endonuclease, a homing
endonuclease from the slime mold Physarum polycephalum, is a small enzyme (2 x 20 kDa) of
known three-dimensional structure that catalyzes the cleavage of a long target DNA sequence
(15 base pairs). Here, a detailed chemical mechanism for catalysis of DNA cleavage by I-PpoI
endonuclease is proposed and tested by creating six variants in which active-site residues are
replaced with alanine. The side chains of three residues (Arg61, His98, and Asn119) are found
to be important for efficient catalysis of DNA cleavage. This finding is consistent with the
proposed mechanism in which His98 abstracts a proton from an attacking water molecule bound by
an adjacent phosphoryl oxygen, Arg61 and Asn119 stabilize the pentavalent transition state,
and Asn119 also binds to the essential divalent metal cation (e.g., Mg(2+) ion), which
interacts with the 3'-oxygen leaving group. Because Mg(2+) is required for cleavage of a
substrate with a good leaving group (p-nitrophenolate), Mg(2+) likely stabilizes the
pentavalent transition state. The pH-dependence of k(cat) for catalysis by I-PpoI reveals a
macroscopic pK(a) of 8.4 for titratable groups that modulate product release. I-PpoI appears
to be unique among known restriction endonucleases and homing endonucleases in its use of a
histidine residue to activate the attacking water molecule for in-line displacement of the
3'-leaving group.

<>

<1>Mansilla, S., Garcia-Ferrer, I., Mendez, C., Salas, J.A., Portugal, J.
<2>Differential inhibition of restriction enzyme cleavage by chromophore-modified analogues of the antitumour antibiotics mithramycin and chromomycin reveals structure-activity relationships.
<3>Biochem. Pharmacol.
<4>79
<5>1418-1427
<6>2010
<7>Differential cleavage at three restriction enzyme sites was used to determine the specific
binding to DNA of the antitumour antibiotics
mithramycin A (MTA), chromomycin A(3) (CRO) and six
chromophore-modified analogues bearing shorter side chains attached at
C-3, instead of the pentyl chain All these antibiotics were obtained
through combinatorial biosynthesis in the producer organisms. MTA, CRO
and their six analogues showed differences in their capacity for
inhibiting the rate of cleavage by restriction enzymes that recognize
C/G-rich tracts. Changes in DNA melting temperature produced by these
molecules were also analyzed, as well as their antiproliferative
activities against a panel of colon, ovarian and prostate human
carcinoma cell lines. Moreover, the cellular uptake of several
analogues was examined to identify whether intracellular retention was
related to cytotoxicity These experimental approaches provided mutually
consistent evidence of a seeming correlation between the strength of
binding to DNA and the antiproliferative activity of the
chromophore-modified molecules Four of the analogues (mithramycin SK,
mithramycin SDK, chromomycin SK and chromomycin SDK) showed promising
biological profiles.

<>

<1>Manso, A.S., Chai, M.H., Atack, J.M., Furi, L., De Ste, C.M., Haigh, R., Trappetti, C., Ogunniyi, A.D., Shewell, L.K., Boitano, M., Clark, T.A., Korlach, J., Blades, M., Mirkes, E., Gorban, A.N., Paton, J.C., Jennings, M.P., Oggioni, M.R.
<2>A random six-phase switch regulates pneumococcal virulence via global epigenetic  changes.
<3>Nat. Commun.
<4>5
<5>5055
<6>2014
<7>Streptococcus pneumoniae (the pneumococcus) is the world's foremost bacterial pathogen in
both morbidity and mortality. Switching between phenotypic forms (or
'phases') that favour asymptomatic carriage or invasive disease was first
reported in 1933. Here, we show that the underlying mechanism for such phase
variation consists of genetic rearrangements in a Type I restriction-modification
system (SpnD39III). The rearrangements generate six alternative specificities
with distinct methylation patterns, as defined by single-molecule, real-time
(SMRT) methylomics. The SpnD39III variants have distinct gene expression
profiles. We demonstrate distinct virulence in experimental infection and in vivo
selection for switching between SpnD39III variants. SpnD39III is ubiquitous in
pneumococci, indicating an essential role in its biology. Future studies must
recognize the potential for switching between these heretofore undetectable,
differentiated pneumococcal subpopulations in vitro and in vivo. Similar systems
exist in other bacterial genera, indicating the potential for broad exploitation
of epigenetic gene regulation.

<>

<1>Manso-Silvan, L. et al.
<2>Draft Genome Sequences of Mycoplasma alkalescens, Mycoplasma arginini, and Mycoplasma bovigenitalium, Three Species with Equivocal Pathogenic Status for  Cattle.
<3>Genome Announcements
<4>1
<5>e00348-13
<6>2013
<7>We report here the draft genome sequences of Mycoplasma alkalescens, Mycoplasma arginini, and
Mycoplasma bovigenitalium. These three species are regularly
isolated from bovine clinical specimens, although their role in disease is
unclear.

<>

<1>Mansour, C.A., Doiron, K.M., Cupples, C.G.
<2>Characterization of functional interactions among the Escherichia coli mismatch repair proteins using a bacterial two-hybrid assay.
<3>Mutat. Res.
<4>485
<5>331-338
<6>2001
<7>Vsr mediates very short patch repair in Escherichia coli, correcting T/G mismatches caused by
deamination of 5-methylcytosine to thymine. MutS and
MutL, part of the post-replication mismatch repair system, stimulate VSP
repair. In this study, we use a bacterial two-hybrid assay to show that
MutL interacts with Vsr. We also show that interaction between Vsr and
MutL inhibits the ability of MutL to dimerize, to interact with MutS and
MutH and to mediate a previously unknown interaction between MutS and
MutH. This inhibition may explain why high levels of Vsr are mutagenic in
vivo. In addition, we show that the Mut fusion proteins are repair
proficient in the bacterial two-hybrid assay, making it possible to study
their interactions in various genetic backgrounds, or in the presence of
DNA damaging agents.

<>

<1>Mansour, S.R., Oshone, R., Hurst, S.G.I.V., Morris, K., Thomas, W.K., Tisa, L.S.
<2>Draft Genome Sequence of Frankia sp. Strain CcI6, a Salt-Tolerant Nitrogen-Fixing Actinobacterium Isolated from the Root Nodule of Casuarina cunninghamiana.
<3>Genome Announcements
<4>2
<5>e01205-13
<6>2014
<7>Members of the actinomycete genus Frankia form a nitrogen-fixing symbiosis with 8 different
families of actinorhizal plants. We report a 5.57-Mbp draft genome
sequence for Frankia sp. strain CcI6, a salt-tolerant nitrogen-fixing
actinobacterium isolated from root nodules of Casurina cunninghamiana grown in
Egyptian soils.

<>

<1>Manwaring, N.P., Skurray, R.A., Firth, N.
<2>Nucleotide sequence of the F plasmid leading region.
<3>Plasmid
<4>41
<5>219-225
<6>1999
<7>The entire nucleotide sequence of the first DNA segment of the conjugative
F plasmid to enter the recipient cell, the leading region, is described.
Analysis of the sequence provides further evidence that products encoded
within the 13.2-kb leading region are likely to be expressed and perform
functions associated with the transferred strand in the recipient cell.

<>

<1>Manzanera, M., Santa-Cruz-Calvo, L., Vilchez, J.I., Garcia-Fontana, C., Silva-Castro, G.A., Calvo, C., Gonzalez-Lopez, J.
<2>Genome Sequence of Arthrobacter siccitolerans 4J27, a Xeroprotectant-Producing Desiccation-Tolerant Microorganism.
<3>Genome Announcements
<4>2
<5>e00526-14
<6>2014
<7>We report the first genome sequence for Arthrobacter siccitolerans 4J27, a newly  described
desiccation-tolerant species. The complete genome of A. siccitolerans
4J27 has been sequenced and is estimated to be around 5.3 Mb in size, with an
average GC content of 65.13%. We predict 4,480 protein-coding sequences (CDSs).

<>

<1>Manzoor, S., Bongcam-Rudloff, E., Schnurer, A., Muller, B.
<2>First Genome Sequence of a Syntrophic Acetate-Oxidizing Bacterium, Tepidanaerobacter acetatoxydans Strain Re1.
<3>Genome Announcements
<4>1
<5>e00213-12
<6>2013
<7>Syntrophic acetate-oxidizing bacteria (SAOB) have been identified as key organisms for
efficient biogas production from protein-rich materials. is the
first reported SAOB for which the genome has been sequenced. Genome analysis will
aid us in understanding the mechanisms regulating syntrophy, particularly
energy-conserving and electron transfer mechanisms.

<>

<1>Manzoor, S., Muller, B., Niazi, A., Bongcam-Rudloff, E., Schnurer, A.
<2>Draft Genome Sequence of Clostridium ultunense Strain Esp, a Syntrophic Acetate-Oxidizing Bacterium.
<3>Genome Announcements
<4>1
<5>e0010713
<6>2013
<7>Clostridium ultunense strain Esp belongs to the functional group of syntrophic
acetate-oxidizing bacteria (SAOB), which have been identified as key organisms
for efficient biogas production from protein-rich materials. Genome analysis and
comparative genomics might aid us to define physiological features that are
essential for maintaining this particular syntrophic lifestyle.

<>

<1>Manzoor, S., Niazi, A., Bejai, S., Meijer, J., Bongcam-Rudloff, E.
<2>Genome Sequence of a Plant-Associated Bacterium, Bacillus amyloliquefaciens Strain UCMB5036.
<3>Genome Announcements
<4>1
<5>e0011113
<6>2013
<7>We announce here the genome sequence of Bacillus amyloliquefaciens strain UCMB5036, a plant
growth-promoting bacterium isolated from a cotton plant. Its
genome contains gene clusters involved in nonribosomal synthesis of secondary
metabolites known for their antimicrobial activities. The availability of this
genome will provide novel insights into plant-bacterium-associated activities.

<>

<1>Mao, D., Grogan, D.
<2>Genomic evidence of rapid, global-scale gene flow in a Sulfolobus species.
<3>ISME J.
<4>6
<5>1613-1616
<6>2012
<7>Local populations of Sulfolobus islandicus diverge genetically with geographical
separation, and this has been attributed to restricted transfer of propagules
imposed by the unfavorable spatial distribution of acidic geothermal habitat. We
tested the generality of genetic divergence with distance in Sulfolobus species
by analyzing genomes of Sulfolobus acidocaldarius drawn from three populations
separated by more than 8000 km. In sharp contrast to S. islandicus, the
geographically diverse S. acidocaldarius genomes proved to be nearly identical.
We could not link the difference in genome conservation between the two species
to a corresponding difference in genome stability or ecological factors affecting
propagule dispersal. The results provide the first evidence that genetic
isolation of local populations does not result primarily from properties
intrinsic to Sulfolobus and the severe discontinuity of its geothermal habitat,
but varies with species, and thus may reflect biotic interactions that act after
propagule dispersal.

<>

<1>Mao, X.H., Wei, M., Zhu, C.F., Lu, J.X., Gao, J.M., Simon, A.J., Shi, J.Y., Huang, Q., Fan, C.H.
<2>Real Time in Vitro Regulation of DNA Methylation Using a 5-Fluorouracil Conjugated DNA-Based Stimuli-Responsive Platform.
<3>ACS Appl. Mater. Inter.
<4>5
<5>2604-2609
<6>2013
<7>DNA methylation, catalyzed by methylases, plays a critical role in many biological processes,
and many methylases have been regarded as
promising targets for antimicrobial drugs. In this work, we report a
stimulus responsive, self-regulating anticancer drug release platform,
comprising a multifunctional DNA that upon methylation by
methyltransferase (MTase) releases 5-fluorouracil (5-Fu) and in turn
inhibits subsequent expression of MTase. The multifunctional DNA with
anticancer drug are first methylated by DNA adenine methylation (DAM)
methyltransferase (MTase) and then cut by the methylation-sensitive
restriction endonuclease Dpn I. Removal of duplex from the functional
DNA by the methylation/cleavage process will release the anticancer
drug, resulting in inhibition of the activity of DAM in turn.
Consequently, the enzyme activity of DAM MTase can be self-regulated.
Furthermore, we found that the inhibition efficiency of 5-Fu
significantly increase as it is functionalized with DNA.

<>

<1>Mao, Z., Li, M., Chen, J.
<2>Draft Genome Sequence of Pseudomonas plecoglossicida Strain NB2011, the Causative Agent of White Nodules in Large Yellow Croaker (Larimichthys crocea).
<3>Genome Announcements
<4>1
<5>e00586-13
<6>2013
<7>We describe the draft genome sequence of Pseudomonas plecoglossicida strain NB2011, the
causative agent of white nodules in cultured large yellow croaker
(Larimichthys crocea) in China. The draft genome sequence of the bacterium
consists of 5.41 million bp, with a G+C content of 62.8%. A total of 4,952 genes
were identified.

<>

<1>Mappley, L.J., Black, M.L., Abuoun, M., Darby, A.C., Woodward, M.J., Parkhill, J., Turner, A.K., Bellgard, M.I., La, T., Phillips, N.D., La Ragione, R.M., Hampson, D.J.
<2>Comparative genomics of Brachyspira pilosicoli strains: genome rearrangements, reductions and correlation of genetic compliment with phenotypic diversity.
<3>BMC Genomics
<4>13
<5>454
<6>2012
<7>ABSTRACT: BACKGROUND: The anaerobic spirochaete Brachyspira pilosicoli causes
enteric disease in avian, porcine and human hosts, amongst others. To date, the
only available genome sequence of B. pilosicoli is that of strain 95/1000, a
porcine isolate. In the first intra-species genome comparison within the
Brachyspira genus, we report the whole genome sequence of B. pilosicoli B2904, an
avian isolate, the incomplete genome sequence of B. pilosicoli WesB, a human
isolate, and the comparisons with B. pilosicoli 95/1000. We also draw on
incomplete genome sequences from three other Brachyspira species. Finally we
report the first application of the high-throughput Biolog phenotype screening
tool on the B. pilosicoli strains for detailed comparisons between genotype and
phenotype. RESULTS: Feature and sequence genome comparisons revealed a high
degree of similarity between the three B. pilosicoli strains, although the
genomes of B2904 and WesB were larger than that of 95/1000 (~2,765, 2.890 and
2.596 Mb, respectively). Genome rearrangements were observed which correlated
largely with the positions of mobile genetic elements. Through comparison of the
B2904 and WesB genomes with the 95/1000 genome, features that we propose are
non-essential due to their absence from 95/1000 include a peptidase, glycine
reductase complex components and transposases. Novel bacteriophages were detected
in the newly-sequenced genomes, which appeared to have involvement in intra- and
inter-species horizontal gene transfer. Phenotypic differences predicted from
genome analysis, such as the lack of genes for glucuronate catabolism in 95/1000,
were confirmed by phenotyping. CONCLUSIONS: The availability of multiple B.
pilosicoli genome sequences has allowed us to demonstrate the substantial genomic
variation that exists between these strains, and provides an insight into genetic
events that are shaping the species. In addition, phenotype screening allowed
determination of how genotypic differences translated to phenotype. Further
application of such comparisons will improve understanding of the metabolic
capabilities of Brachyspira species.

<>

<1>Marasa, B.S., Revollo, J., Iram, S., Sung, K., Xu, J., Khan, S.
<2>Draft Genome Sequence of a Methicillin-Resistant Staphylococcus aureus ST1413 Strain for Studying Genetic Mechanisms of Antibiotic Resistance.
<3>Genome Announcements
<4>2
<5>e00162-14
<6>2014
<7>Here we report the whole draft genome sequence of a methicillin-resistant Staphylococcus
aureus ST1413 strain. Determining the distribution and arrangement
of various genes associated with drug resistance, toxicity, and diseases will
enhance our understanding about its adaptability to thrive in different
ecological niches and help in the development of effective treatments for
enterotoxigenic staphylococcal infections.

<>

<1>Marcaida, M.J. et al.
<2>Homing endonucleases: from basics to therapeutic applications.
<3>Cell. Mol. Life Sci.
<4>67
<5>727-748
<6>2010
<7>Homing endonucleases (HE) are double-stranded DNAses that target large recognition sites
(12-40 bp). HE-encoding sequences are usually embedded in either introns or inteins. Their
recognition sites are extremely rare, with none or only a few of these sites present in a
mammalian-sized genome. However, these enzymes, unlike standard restriction endonucleases,
tolerate some sequence degeneracy within their recognition sequence. Several members of this
enzyme family have been used as templates to engineer tools to cleave DNA sequences that
differ from their original wild-type targets. These custom HEs can be used to stimulate
double-strand break homologous recombination in cells, to induce the repair of defective genes
with very low toxicity levels. The use of tailored HEs opens up new possibilities for gene
therapy in patients with monogenic diseases that can be treated ex vivo. This review provides
an overview of recent advances in this field.

<>

<1>Marcaida, M.J., Prieto, J., Redondo, P., Nadra, A.D., Alibes, A., Serrano, L., Grizot, S., Duchateau, P., Paques, F., Blanco, F.J., Montoya, G.
<2>Crystal structure of I-DmoI in complex with its target DNA provides new insights into meganuclease engineering.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>16888-16893
<6>2008
<7>Homing endonucleases, also known as meganucleases, are sequence-specific enzymes with large
DNA recognition sites. These enzymes can be used to
induce efficient homologous gene targeting in cells and plants, opening
perspectives for genome engineering with applications in a wide series of
fields, ranging from biotechnology to gene therapy. Here, we report the
crystal structures at 2.0 and 2.1 A resolution of the I-DmoI meganuclease
in complex with its substrate DNA before and after cleavage, providing
snapshots of the catalytic process. Our study suggests that I-DmoI
requires only 2 cations instead of 3 for DNA cleavage. The structure sheds
light onto the basis of DNA binding, indicating key residues responsible
for nonpalindromic target DNA recognition. In silico and in vivo analysis
of the I-DmoI DNA cleavage specificity suggests that despite the
relatively few protein-base contacts, I-DmoI is highly specific when
compared with other meganucleases. Our data open the door toward the
generation of custom endonucleases for targeted genome engineering using
the monomeric I-DmoI scaffold.

<>

<1>Marcaida-Lopez, M.J., Prieto-Lugo, F.J., Montoya-Blanco, G.
<2>The crystal structure of homing endonuclease I-DmoI in complex with its DNA target and its use to model improved chimeric meganucleases and new DNA targets.
<3>International Patent Office
<4>WO 201001189 A
<5>
<6>2010
<7>The present invention relates to the three-dimensional structure of the meganuclease I-DmoI in
combination with its DNA target and from this the prediction of residues in the I-DmoI enzyme
which affect the binding, catalytic and other properties of this enzyme.  The present
invention also relates to I-DmoI enzymes with altered characteristics such as altered target
half sites or altered catalysis properties and to chimeric meganucleases comprising portion of
I-DmoI and to the use of the three-dimensional structure of the meganuclease I-DmoI in
combination with its DNA target in an in silico screening method.

<>

<1>March, J.B., Clark, J.
<2>Enzymes by post-restriction enzyme stability.
<3>Nat. Biotechnol.
<4>18
<5>243
<6>2000
<7>The authors suggest that many restriction enzymes are sufficiently stable at room temperature
that they could be transported abroad without expensive ice-packs or dry ice.

<>

<1>Marchionni, M.A., Roufa, D.J.
<2>Digestion of 5-bromodeoxyuridine-substituted lambda-DNA by restriction endonucleases.
<3>J. Biol. Chem.
<4>253
<5>9075-9081
<6>1978
<7>5-bromodeoxyuridine (BrdUrd) DNA from bacteriophage lambda affects both the
rates and sites of cleavage by Endo R.EcoRI, HindIII, and SmaI.  Endonucleases
EcoRI and HindIII, both of which recognize nucleotide sequences that contain 4
thymidine residues, cleaved fully substituted BrdUrd-DNA at the same sites as
unsubstituted DNA.  However, when treated with limiting amounts of either of
the two endonucleases, BrdUrd-DNA was digested more slowly than was the
unsubstituted DNA.  Endo R.SmaI, which recognizes a nucleotide sequence that
does not contain thymidine digested BrdUrd-DNA at approximately the same rate
as unsubstituted DNA.  Surprisingly, one of the three SmaI sites in lambda-DNA
(located at 0.656 on the genome's physical map) appeared to be highly resistant
to cleavage by SmaI in substituted DNA.  Hence, cleavage of BrdUrd-DNA by SmaI
generated three restriction products instead of the expected four products
derived from unsubstituted DNA.  These BrdUrd-DNA products were identified as
the characteristic SmaI.A and C fragments as well as an unusual, large product
that contained the fused B and D fragments.  Therefore, the SmaI cleavage site
at the junction of the B-D fragments appears to differ from the other two sites
by aspects of DNA structure determined outside of the canonical SmaI
recognition sequence.  This finding indicates that site-specific DNA enzymes
can be influenced by DNA determinants that reside outside of the accepted
recognition sequences of the enyzmes.

<>

<1>Marcoleta, A., Gutierrez-Cortez, S., Maturana, D., Monasterio, O., Lagos, R.
<2>Whole-Genome Sequence of the Microcin E492-Producing Strain Klebsiella pneumoniae RYC492.
<3>Genome Announcements
<4>1
<5>e00178-13
<6>2013
<7>Here, we report the draft genome sequence of the Gram-negative strain Klebsiella  pneumoniae
RYC492, which produces the amyloid-forming and antibacterial peptide
microcin E492. The sequenced genome consists of a 5,095,761-bp assembled open
chromosome where the gene cluster for microcin production is located in a
putative 31-kb genomic island flanked by sequence repeats and containing a
putative integrase-coding gene.

<>

<1>Marcon, J., Taketani, R.G., Dini-Andreote, F., Mazzero, G.I., Soares, F.L.J., Melo, I.S., Azevedo, J.L., Andreote, F.D.
<2>Draft Genome Sequence of Bacillus thuringiensis Strain BrMgv02-JM63, a Chitinolytic Bacterium Isolated from Oil-Contaminated Mangrove Soil in Brazil.
<3>Genome Announcements
<4>2
<5>e01264-13
<6>2014
<7>Here, we report the draft genome sequence and the automatic annotation of Bacillus
thuringiensis strain BrMgv02-JM63. This genome comprises a set of genes
involved in the metabolism of chitin and N-acetylglucosamine utilization, thus
suggesting the possible role of this strain in the cycling of organic matter in
mangrove soils.

<>

<1>Marczynski, G.T.
<2>Chromosome methylation and measurement of faithful, once and only once per cell cycle chromosome replication in Caulobacter crescentus.
<3>J. Bacteriol.
<4>181
<5>1984-1993
<6>1999
<7>Caulobacter crescentus exhibits cell-type-specific control of chromosome replication and DNA
methylation.  Asymmetric cell division yields a replicating stalked cell and a nonreplicating
swarmer cell.  The motile swarmer cell must differentiate into a sessile stalked cell in order
to replicate and execute asymmetric cell division.  This program of cell division implies that
chromosome replication initiates in the stalked cell only once per cell cycle.  DNA
methylation is restricted to the predivisional cell stage, and since DNA synthesis produces an
unmethylated nascent strand, late DNA methylation also implies that DNA near the replication
origin remains hemimethylated longer than DNA located further away.  In this report, both
assumptions are tested with an engineered Tn5-based transposon, Tn5omega-MP.  This allows a
sensitive Southern blot assay that measures fully methylated, hemimethylated, and unmethylated
DNA duplexes.  Tn5omega-MP DNA near the replication origin remained hemimethylated longer than
DNA located further away.  One Tn5omega-MP placed near the replication origin revealed small
but detectable amounts of unmethylated duplex DNA in replicating stalked cells.  Extra DNA
synthesis produces a second unmethylated nascent strand.  Therefore, measurement of
unmethylated DNA is a critical test of the "once and only once per cell cycle" rule of
chromosome replication in C. crescentus.  Fewer than 1 in 1,000 stalked cells prematurely
initiate a second round of chromosome replication.  The implications for very precise negative
control of chromosome replication are discussed with respect to the bacterial cell cycle.

<>

<1>Mardanov, A.V., Beletskii, A.V., Gumerov, V.M., Karbysheva, E.A., Mikheeva, L.E.
<2>New low-copy plasmid in cyanobacterium Anabaena variabilis.
<3>Genetika
<4>49
<5>798-805
<6>2013
<7>Complete genome sequencing was performed for Anabaena variabilis ATCC 29413 from the
collection of the Chair of Genetics, Department of
Biology, Moscow State University, Russia. In addition to known plasmids
A, B, and C, a new circular low-copy plasmid was detected and named D.
It was also sequenced completely and found to have 27051 bp. The
plasmid contained the parA and parB genes of the partition system, two
genes that encode replication proteins, a gene for site-specific
recombinase, a type-I restriction-modification system, and several
genes with unknown functions. Analysis by PCR revealed the presence of
plasmid D in two epiphytic strains from Vietnam, i.e., Anabaena sp. 182
and Anabaena sp. 281, as well as in Anabaena sp. V5 and A. azollae
(Newton's isolate).

<>

<1>Mardanov, A.V., Eldarov, M.A., Sklyarenko, A.V., Dumina, M.V., Beletsky, A.V., Yarotsky, S.V., Ravin, N.V.
<2>Draft Genome Sequence of Escherichia coli Strain VKPM B-10182, Producing the Enzyme for Synthesis of Cephalosporin Acids.
<3>Genome Announcements
<4>2
<5>e01222-14
<6>2014
<7>Escherichia coli strain VKPM B-10182, obtained by chemical mutagenesis from E. coli strain
ATCC 9637, produces cephalosporin acid synthetase employed in the
synthesis of beta-lactam antibiotics, such as cefazolin. The draft genome
sequence of strain VKPM B-10182 revealed 32 indels and 1,780 point mutations that
might account for the improvement in antibiotic synthesis that we observed.

<>

<1>Mardanov, A.V., Gumerov, V.M., Beletsky, A.V., Prokofeva, M.I., Bonch-Osmolovskaya, E.A., Ravin, N.V., Skryabin, K.G.
<2>Complete genome sequence of the thermoacidophilic crenarchaeon Thermoproteus uzoniensis 768-20.
<3>J. Bacteriol.
<4>193
<5>3156-3157
<6>2011
<7>Thermoproteus uzoniensis 768-20 is a thermoacidophilic anaerobic crenarchaeon isolated from a
solfataric field in Kamchatka, Russia. The
complete genome sequence reveals genes for protein and carbohydrate-active
enzymes, beta-oxidation of fatty acids, the Embden-Meyerhof and
Entner-Doudoroff pathways for glucose metabolism, the tricarboxylic acid
cycle, the dicarboxylate/4-hydroxybutyrate cycle, hydrogenase and sulfur
reductase.

<>

<1>Mardanov, A.V., Gumerov, V.M., Slobodkina, G.B., Beletsky, A.V., Bonch-Osmolovskaya, E.A., Ravin, N.V., Skryabin, K.G.
<2>Complete genome sequence of strain 1860, a crenarchaeon of the genus pyrobaculum able to grow with various electron acceptors.
<3>J. Bacteriol.
<4>194
<5>727-728
<6>2012
<7>Strain 1860, a novel member of the genus Pyrobaculum, is a hyperthermophilic organotrophic
crenarchaeon growing anaerobically with
various electron acceptors. The complete genome sequence reveals genes for
several membrane-bound oxidoreductases, the Embden-Meyerhof and
Entner-Doudoroff pathways for glucose metabolism, the tricarboxylic acid
cycle, the glyoxylate cycle, and the dicarboxylate/4-hydroxybutyrate
cycle.

<>

<1>Mardanov, A.V., Kochetkova, T.V., Beletsky, A.V., Bonch-Osmolovskaya, E.A., Ravin, N.V., Skryabin, K.G.
<2>Complete Genome Sequence of the Hyperthermophilic Cellulolytic Crenarchaeon 'Thermogladius cellulolyticus' 1633.
<3>J. Bacteriol.
<4>194
<5>4446-4447
<6>2012
<7>Strain 1633, a novel member of the genus Thermogladius, isolated from a freshwater hot spring,
is an anaerobic hyperthermophilic crenarchaeon capable of
fermenting proteinaceous and cellulose substrates. The complete genome sequence
reveals genes for protein and carbohydrate-active enzymes, the Embden-Meyerhof
pathway for glucose metabolism, cytoplasmic NADP-dependent hydrogenase, and
several energy-coupling membrane-bound oxidoreductases.

<>

<1>Mardanov, A.V., Ravin, N.V., Svetlitchnyi, V.A., Beletsky, A.V., Miroshnichenko, M.L., Bonch-Osmolovskaya, E.A., Skryabin, K.G.
<2>Metabolic versatility and indigenous origin of the archaeon Thermococcus sibiricus, isolated from a siberian oil reservoir, as revealed by genome analysis.
<3>Appl. Environ. Microbiol.
<4>75
<5>4580-4588
<6>2009
<7>Thermococcus species are widely distributed in terrestrial and marine
hydrothermal areas, as well as in deep subsurface oil reservoirs.
Thermococcus sibiricus is a hyperthermophilic anaerobic archaeon isolated
from a well of the never flooded oil-bearing Jurassic horizon of a
high-temperature oil reservoir. To obtain insight into the genome of an
archaeon inhabiting the oil reservoir, we have determined and annotated
the complete 1,845,800-base genome of T. sibiricus. A total of 2,061
protein-coding genes have been identified, 387 of which are absent in
other members of the order Thermococcales. Physiological features and
genomic data reveal numerous hydrolytic enzymes (e.g., cellulolytic
enzymes, agarase, laminarinase, and lipases) and metabolic pathways,
support the proposal of the indigenous origin of T. sibiricus in the oil
reservoir, and explain its survival over geologic time and its
proliferation in this habitat. Indeed, in addition to proteinaceous
compounds known previously to be present in oil reservoirs at limiting
concentrations, its growth was stimulated by cellulose, agarose, and
triacylglycerides, as well as by alkanes. Two polysaccharide degradation
loci were probably acquired by T. sibiricus from thermophilic bacteria
following lateral gene transfer events. The first, a "saccharolytic gene
island" absent in the genomes of other members of the order
Thermococcales, contains the complete set of genes responsible for the
hydrolysis of cellulose and beta-linked polysaccharides. The second
harbors genes for maltose and trehalose degradation. Considering that
agarose and laminarin are components of algae, the encoded enzymes and the
substrate spectrum of T. sibiricus indicate the ability to metabolize the
buried organic matter from the original oceanic sediment.

<>

<1>Mardanov, A.V., Slododkina, G.B., Slobodkin, A.I., Beletsky, A.V., Gavrilov, S.N., Kublanov, I.V., Bonch-Osmolovskaya, E.A., Skryabin, K.G., Ravin, N.V.
<2>The genome of Geoglobus acetivorans: Fe(III) reduction, acetate utilization, autotrophic growth and degradation of aromatic compounds in a hyperthermophilic archaeon.
<3>Appl. Environ. Microbiol.
<4>0
<5>0
<6>2014
<7>Geoglobus acetivorans is a hyperthermophilic anaerobic euryarchaeon of the order
Archaeoglobales isolated from deep-sea hydrothermal vents. A unique physiological
feature of the members of the genus Geoglobus is their obligate dependence on
Fe(III) reduction, which plays an important role in the geochemistry of
hydrothermal systems. The features of this organism and its complete 1,860,815-bp
genome sequence are described in this report. Genome analysis revealed pathways
enabling oxidation of molecular hydrogen, proteinaceous substrates, fatty acids,
aromatic compounds, n-alkanes and organic acids including acetate, through
anaerobic respiration linked to Fe(III) reduction. Consistent with the inability
of G. acetivorans to grow on carbohydrates, the modified Embden-Meyerhof pathway
encoded by the genome is incomplete. Autotrophic CO2 fixation is enabled by the
Wood-Ljungdahl pathway. Reduction of insoluble poorly crystalline Fe(III) oxide
depends on the transfer of electrons from the quinone pool to multiheme c-type
cytochromes exposed on the cell surface. Direct contact of the cells and Fe(III)
oxide particles could be facilitated by pili-like appendages. Genome analysis
indicated the presence of metabolic pathways for anaerobic degradation of
aromatic compounds and n-alkanes, although the ability of G. acetivorans to grow
on these substrates was not observed in laboratory experiments. Overall, our
results suggest that Geoglobus species could play an important role in microbial
communities of deep-sea hydrothermal vents as lithoautotrophic producers. An
additional role as decomposers would close the biogeochemical cycle of carbon
through complete mineralization of various organic compounds via Fe(III)
respiration.

<>

<1>Mardanov, A.V., Svetlitchnyi, V.A., Beletsky, A.V., Prokofeva, M.I., Bonch-Osmolovskaya, E.A., Ravin, N.V., Skryabin, K.G.
<2>The Genome Sequence of the Crenarchaeon Acidilobus saccharovorans Supports a New Order, Acidilobales, and Suggests an Important Ecological Role in Terrestrial Acidic Hot Springs.
<3>Appl. Environ. Microbiol.
<4>76
<5>5652-5657
<6>2010
<7>Acidilobus saccharovorans is an anaerobic, organotrophic,
thermoacidophilic crenarchaeon isolated from a terrestrial hot spring. We
report the complete genome sequence of A. saccharovorans, which has
permitted the prediction of genes for Embden-Meyerhof and Entner-Doudoroff
pathways and genes associated with the oxidative tricarboxylic acid cycle.
The electron transfer chain is branched with two sites of proton
translocation and is linked to the reduction of elemental sulfur and
thiosulfate. The genomic data suggest an important role of the order
Acidilobales in thermoacidophilic ecosystems whereby its members can
perform a complete oxidation of organic substrates, closing the anaerobic
carbon cycle.

<>

<1>Mardanova, A.M., Toymentseva, A.A., Gilyazeva, A.G., Kazakov, S.V., Shagimardanova, E.I., Khaitlina, S.Y., Sharipova, M.R.
<2>Draft Genome Sequence of Serratia grimesii Strain A2.
<3>Genome Announcements
<4>2
<5>e00937-14
<6>2014
<7>We report the first draft genome assembly of Serratia grimesii strain A2, previously
identified as Escherichia coli strain A2, which produces protease
ECP32 with a high specificity toward actin. S. grimesii strain A2 has multidrug
resistance associated with a number of efflux pump genes.

<>

<1>Marenda, M.S., Sagne, E., Poumarat, F., Citti, C.
<2>Suppression subtractive hybridization as a basis to assess Mycoplasma agalactiae and Mycoplasma bovis genomic diversity and species-specific sequences.
<3>Microbiology
<4>151
<5>475-489
<6>2005
<7>The phylogenically related Mycoplasma agalactiae and Mycoplasma bovis
species are two ruminant pathogens difficult to differentiate and for
which a limited amount of sequence data are available. To assess the
degree of genomic diversity existing between and within these mycoplasma
species, sets of DNA fragments specific for M. bovis type-strain PG45 or
for M. agalactiae type-strain PG2 were isolated by suppression subtractive
hybridization and used as probes on a panel of M. agalactiae and M. bovis
field isolates. Results indicated that approximately 70 % of the DNA
fragments specific to one or the other type strain are represented in all
field isolates of the corresponding species. Only one M. bovis isolate,
which was first classified as M. agalactiae, reacted with 15 % of the
PG2-specific probes, while several M. agalactiae isolates reacted with 15
% of the PG45-specific probes. Sequence analyses indicated that most of
the genomic diversity observed within one species is related to ORFs with
(i) no homologies to proteins recorded in the databases or (ii) homologies
to proteins encoded by restriction modification systems. Reminiscent of
gene transfer as a means for genomic diversity, a PG45-specific DNA
fragment with significant homologies to a central protein of an
integrative conjugative element of Mycoplasma fermentans (ICEF) was found
in most M. bovis field isolates and in a few M. agalactiae isolates.
Finally, sequences encoding part of DNA polymerase III were found in both
sets of M. agalactiae- and M. bovis-specific DNA fragments and were used
to design a species-specific PCR assay for the identification and
differentiation of M. agalactiae and M. bovis.

<>

<1>Margolles, A., Gueimonde, M., Sanchez, B.
<2>Genome Sequence of the Antarctic Psychrophile Bacterium Planococcus antarcticus DSM 14505.
<3>J. Bacteriol.
<4>194
<5>4465
<6>2012
<7>Planococcus antarcticus DSM 14505 is a psychrophile bacterium that was isolated from
cyanobacterial mat samples, originally collected from ponds in McMurdo,
Antarctica. This orange-pigmented bacterium grows at 4 degrees C and may possess
interesting enzymatic activities at low temperatures. Here we report the first
genomic sequence of P. antarcticus DSM 14505.

<>

<1>Margot, J.B., Aguirre-Arteta, A.M., Di Giacco, B.V., Pradhan, S., Roberts, R.J., Cardoso, M.C., Leonhardt, H.
<2>Structure and Function of the Mouse DNA Methyltransferase Gene: Dnmt1 shows a Tripartite Structure.
<3>J. Mol. Biol.
<4>297
<5>293-300
<6>2000
<7>Dnmt1 is the predominant DNA methyltransferase (MTase) in mammals. The C-terminal domain of
Dnmt1 clearly shares sequence similarity with many
prokaryotic 5mC methyltransferases, and had been proposed to be sufficient for catalytic
activity. We show here by deletion analysis that the C-terminal domain alone is not
sufficient for methylating activity, but that a large part of the N-terminal domain is
required in addition. Since this complex structure of Dnmt1 raises issues about its
evolutionary origin, we have compared several eukaryotic MTases and have determined the
genomic organization of the mouse Dnmt1 gene. The 5' most part of the
N-terminal domain is dispensible for enzyme activity, includes the major nuclear import signal
and comprises tissue-specific exons. Interestingly, the functional subdivision
of Dnmt1 correlates well with the structure of the Dnmt1 gene in terms of intron/exon size
distribution as well as sequence conservation. Our results, based on functional,
structural and sequence comparison data, suggest that the gene has evolved from the fusion of
at least three genes.

<>

<1>Margot, J.B., Cardoso, M.C., Leonhardt, H.
<2>Mammalian DNA methyltransferases show different subnuclear distributions.
<3>J. Cell. Biochem.
<4>83
<5>373-379
<6>2001
<7>In mammalian cells, DNA methylation patterns are precisely maintained after DNA replication
with defined changes occurring during development. The major DNA methyltransferase (Dnmt1) is
associated with nuclear replication sites during S-phase, which is consistent with a role in
maintenance methylation. The subcellular distribution of the recently discovered de novo DNA
methyltransferases, Dnmt3a and Dnmt3b, was investigated by immunofluorescence and by epitope
tagging. We now show that both Dnmt3a and Dnmt3b are distributed throughout the nucleoplasm
but are not associated with nuclear DNA replication sites during S-phase. These results
suggest that de novo methylation by Dnmt3a and Dnmt3b occurs independently of the replication
process and might involve an alternative mechanism for accessing the target DNA. The different
subcellular distribution of mammalian DNA methyltransferases might thus contribute to the
regulation of DNA methylation.

<>

<1>Margot, J.B., Ehrenhofer-Murray, A.E., Leonhardt, H.
<2>Interactions within the mammalian DNA methyltransferase family.
<3>BMC Mol. Biol.
<4>4
<5>7
<6>2003
<7>BACKGROUND: In mammals, epigenetic information is established and maintained via the
postreplicative methylation of cytosine residues by the
DNA methyltransferases Dnmt1, Dnmt3a and Dnmt3b. Dnmt1 is required for
maintenance methylation whereas Dnmt3a and Dnmt3b are responsible for de
novo methylation. Contrary to Dnmt3a or Dnmt3b, the isolated C-terminal
region of Dnmt1 is catalytically inactive, despite the presence of the
sequence motifs typical of active DNA methyltransferases. Deletion
analysis has revealed that a large part of the N-terminal domain is
required for enzymatic activity. RESULTS: The role played by the
N-terminal domain in this regulation has been investigated using the yeast
two-hybrid system. We show here the presence of an intra-molecular
interaction in Dnmt1 but not in Dnmt3a or Dnmt3b. This interaction was
confirmed by immunoprecipitation and was localized by deletion mapping.
Furthermore, a systematic analysis of interactions among the Dnmt family
members has revealed that DNMT3L interacts with the C-terminal domain of
Dnmt3a and Dnmt3b. CONCLUSIONS: The lack of methylating ability of the
isolated C-terminal domain of Dnmt1 could be explained in part by a
physical interaction between N- and C-terminal domains that apparently is
required for activation of the catalytic domain. Our deletion analysis
suggests that the tertiary structure of Dnmt1 is important in this process
rather than a particular sequence motif. Furthermore, the interaction
between DNMT3L and the C-terminal domains of Dnmt3a and Dnmt3b suggests a
mechanism whereby the enzymatically inactive DNMT3L brings about the
methylation of its substrate by recruiting an active methylase.

<>

<1>Margulies, M. et al.
<2>Genome sequencing in microfabricated high-density picolitre reactors.
<3>Nature
<4>437
<5>376-380
<6>2005
<7>The proliferation of large-scale DNA-sequencing projects in recent years has
driven a search for alternative methods to reduce time and cost. Here we describe
a scalable, highly parallel sequencing system with raw throughput significantly
greater than that of state-of-the-art capillary electrophoresis instruments. The
apparatus uses a novel fibre-optic slide of individual wells and is able to
sequence 25 million bases, at 99% or better accuracy, in one four-hour run. To
achieve an approximately 100-fold increase in throughput over current Sanger
sequencing technology, we have developed an emulsion method for DNA amplification
and an instrument for sequencing by synthesis using a pyrosequencing protocol
optimized for solid support and picolitre-scale volumes. Here we show the
utility, throughput, accuracy and robustness of this system by shotgun sequencing
and de novo assembly of the Mycoplasma genitalium genome with 96% coverage at
99.96% accuracy in one run of the machine.

<>

<1>Marin, M.A., Fonseca, E.L., Andrade, B.N., Cabral, A.C., Vicente, A.C.
<2>Worldwide Occurrence of Integrative Conjugative Element Encoding Multidrug Resistance Determinants in Epidemic Vibrio cholerae O1.
<3>PLoS ONE
<4>9
<5>E108728
<6>2014
<7>In the last decades, there has been an increase of cholera epidemics caused by
multidrug resistant strains. Particularly, the integrative and conjugative
element (ICE) seems to play a major role in the emergence of multidrug resistant
Vibrio cholerae. This study fully characterized, by whole genome sequencing, new
ICEs carried by multidrug resistant V. cholerae O1 strains from Nigeria (2010)
(ICEVchNig1) and Nepal (1994) (ICEVchNep1). The gene content and gene order of
these two ICEs are the same, and identical to ICEVchInd5, ICEVchBan5 and
ICEVchHai1 previously identified in multidrug resistant V. cholerae O1. This ICE
is characterized by dfrA1, sul2, strAB and floR antimicrobial resistance genes,
and by unique gene content in HS4 and HS5 ICE regions. Screening for ICEs, in
publicly available V. cholerae genomes, revealed the occurrence and widespread
distribution of this ICE among V. cholerae O1. Metagenomic analysis found
segments of this ICE in marine environments far from the direct influence of the
cholera epidemic. Therefore, this study revealed the epidemiology of a
spatio-temporal prevalent ICE in V. cholerae O1. Its occurrence and dispersion in
V. cholerae O1 strains from different continents throughout more than two decades
can be indicative of its role in the fitness of the current pandemic lineage.

<>

<1>Marinho-Almeida, D., Dini-Andreote, F., Camargo-Neves, A.A., Juca-Ramos, R.T., Andreote, F.D., Carneiro, A.R., Oliveira-de-Souza, L.A., Caracciolo-Gomes-de-Sa, P.H., Ribeiro-Barbosa, M.S., Araujo, W.L., Silva, A.
<2>Draft Genome Sequence of Methylobacterium mesophilicum Strain SR1.6/6, Isolated from Citrus sinensis.
<3>Genome Announcements
<4>1
<5>e00356-13
<6>2013
<7>Methylobacterium mesophilicum strain SR1.6/6 is an endophytic bacterium isolated  from a
surface-sterilized Citrus sinensis branch. Ecological and biotechnological
aspects of this bacterium, such as the genes involved in its association with the
host plant and the primary oxidation of methanol, were annotated in the draft
genome.

<>

<1>Marinus, M.G.
<2>DNA methylation in Escherichia coli.
<3>Annu. Rev. Genet.
<4>21
<5>113-131
<6>1987
<7>None

<>

<1>Marinus, M.G.
<2>Methylation of DNA.
<3>Escherichia coli and Salmonella typhimurium: Cellular and molecular biology., ASM Publications, Neidhardt, F.C., Washington, D.C.
<4>0
<5>697-702
<6>1987
<7>None

<>

<1>Marinus, M.G.
<2>Methylation of prokaryotic DNA.
<3>DNA Methylation. Biochemistry and Biological Significance., Springer-Verlag, Razin, A., Cedar, H., Riggs, A.D., New York
<4>0
<5>81-109
<6>1984
<7>The biological function of methylated bases in DNA of prokaryotes appears to be
quite different than that of eukaryotes.  In this chapter, most of the
information presented is derived fom studies with Escherichia coli K-12 simply
because more is known about DNA methylation in this organism than in any other
one.  Some data from certain E. coli bacteriophages also will be reviewed, in
addition to selected aspects about DNA methylation in certain other
prokaryotes.  Other recent reviews that complement this one are by Razin and
Friedman (1981) and Hattman (1981).

<>

<1>Marinus, M.G.
<2>DNA methylation influences trpR promoter activity in Escherichia coli K-12.
<3>Mol. Gen. Genet.
<4>200
<5>185-186
<6>1985
<7>Methylation of adenine in the GATC-sequence of the -35 region of the trpR
promoter decreases activity by 2-3 fold.

<>

<1>Marinus, M.G.
<2>Location of DNA methylase genes on the Escherichia coli K-12 genetic map.
<3>Mol. Gen. Genet.
<4>127
<5>47-55
<6>1973
<7>The genes responsible for DNA adenine methylation (dam) and DNA cytosine methylation (dcm)
have been mapped on the E. coli K-12 genetic map. The dam gene is situated at min 65 and the
gene order cysG-(trpS, dam)-aroB inferred. The dcm gene is located at min 37.5 and the gene
order is supD-dcm-flaA1. In F' merodiploids, the dam and dcm alleles are recessive.

<>

<1>Marinus, M.G.
<2>Adenine methylation of Okazaki fragments in Escherichia coli.
<3>J. Bacteriol.
<4>128
<5>853-854
<6>1976
<7>In Escherichia coli polA lig-4 bacteria, the moles percent 6-methyladenine
content of 10S deoxyribonucleic acid (Okazaki fragments) is 0.96 compared with
1.4 for bulk desoxyribonucleic acid.

<>

<1>Marinus, M.G.
<2>Influence of uvrD3, uvrE502, and recL152 mutations on the phenotypes of Escherichia coli K-12 dam mutants.
<3>J. Bacteriol.
<4>141
<5>223-226
<6>1980
<7>The recF143 allele did not alter the phenotypes of dam mutants of Escherichia coli. The uvrD3,
uvrE502, and recL152 mutations did alter some of the phenotypes of dam bacteria. It was
concluded that the uvrD, uvrE, and recL gene products are involved in the same
deoxyribonucleic acid repair pathway as the dam gene product.

<>

<1>Marinus, M.G.
<2>Recombination is essential for viability of an Escherichia coli dam (DNA adenine methyltransferase) mutant.
<3>J. Bacteriol.
<4>182
<5>463-468
<6>2000
<7>Double mutants of Escherichia coli dam (DNA adenine methyltransferase) strains with ruvA,
ruvB, or ruvC could not be constructed, whereas dam derivatives with recD, recF, recJ, and
recR were viable.  The ruv gene products are required for Holliday junction translocation and
resolution of recombination intermediates.  A dam recG (Holliday junction translocation)
mutant strain was isolated but at a very much lower frequency than expected.  The inviability
of a dam lexA (Ind-) host was abrogated by the simultaneous presence of plasmids encoding both
recA and ruvAB.  This result indicates that of more than 20 SOS genes, only recA and ruvAB
need to be derepressed to allow for dam mutant survival.  The presence of mutS or mutL
mutations allowed the construction of dam lexA (Ind-) derivatives.  The requirement for recA,
recB, recC, ruvA, ruvB, ruvC, and possibly recG gene expression indicates that recombination
is essential for viability of dam bacteria probably to repair DNA double-strand breaks.  The
effect of mutS and mutL mutations indicates that DNA mismatch repair is the ultimate source of
most of these DNA breaks.  The requirement for recombination also suggests an explanation for
the sensitivity of dam cells to certain DNA-damaging agents.

<>

<1>Marinus, M.G.
<2>DNA Methylation.
<3>Methylation of DNA in Escherichia coli and Salmonella typhimurium: Cellular and Molecular Biology, Am. Soc. Microbiol., Neidhardt, F.C., Ingraham, J.L., Low, K.B., Magasanik, B., Schaechter, M., Umbarger, H.E., Washington, DC
<4>0
<5>782-791
<6>1996
<7>DNA methylation in bacteria is most often thought of in its
role to protect DNA from restriction endonucleases.  In addition to this
role, however, studies in Escherichia coli have shown that methylated
bases have other biological functions.  As described below, DNA adenine
methylation is frequently used to control the rate at which these
functions exert their effects.  Thus it is primarily used for regulatory
purposes.

<>

<1>Marinus, M.G., Carraway, M., Frey, A.Z., Brown, L., Arraj, J.A.
<2>Insertion mutations in the dam gene of Escherichia coli K-12.
<3>Mol. Gen. Genet.
<4>192
<5>288-289
<6>1983
<7>The dam gene of E. coli can be inactivated by insertion of Tn9 or Mud phage. Strains bearing
these mutations are viable indicating that the dam gene product is dispensable.

<>

<1>Marinus, M.G., Casadesus, J.
<2>Roles of DNA adenine methylation in host-pathogen interactions: mismatch repair, transcriptional regulation, and more.
<3>FEMS Microbiol. Rev.
<4>33
<5>488-503
<6>2009
<7>The DNA adenine methyltransferase (Dam methylase) of Gammaproteobacteria and the cell
cycle-regulated methyltransferase (CcrM) methylase of Alphaproteobacteria catalyze an
identical reaction (methylation of adenosine moieties using S-adenosyl-methionine as a methyl
donor) at similar DNA targets (GATC and GANTC, respectively). Dam and CcrM are of independent
evolutionary origin. Each may have evolved from an ancestral restriction-modification system
that lost its restriction component, leaving an 'orphan' methylase devoted solely to
epigenetic genome modification. The formation of 6-methyladenine reduces the thermodynamic
stability of DNA and changes DNA curvature. As a consequence, the methylation state of
specific adenosine moieties can affect DNA-protein interactions. Well-known examples include
binding of the replication initiation complex to the methylated oriC, recognition of
hemimethylated GATCs in newly replicated DNA by the MutHLS mismatch repair complex, and
discrimination of methylation states in promoters and regulatory DNA motifs by RNA polymerase
and transcription factors. In recent years, Dam and CcrM have been shown to play roles in
host-pathogen interactions. These roles are diverse and have only partially been understood.
Especially intriguing is the evidence that Dam methylation regulates virulence genes in
Escherichia coli, Salmonella, and Yersinia at the posttranscriptional level.

<>

<1>Marinus, M.G., Konrad, E.B.
<2>Hyper-recombination in dam mutants of Escherichia coli K-12.
<3>Mol. Gen. Genet.
<4>149
<5>273-277
<6>1976
<7>F-prime heterogenotes of dam-3 bacteria segregate F-prime homogenotes at a frequency 30-200
times higher than the isogenic dam+ strain. A hyperrecombination mutant which shows increased
recombination between chromosomal duplications was characterized as a dam mutant. The dam-3
allele causes a reduction in linkage of proximal unselected markers in transconjugants and
increases the recombination frequency between a pair of closely linked markers. It is
concluded that dam mutations confer a hyperrecombination phenotype to the cell.

<>

<1>Marinus, M.G., Lobner-Olesen, A.
<2>DNA Methylation.
<3>EcoSal-Escherichia coli and Salmonella:   Cellular and Molecular Biology., ASM Press, Boeck, A., Curtiss, R. III, Kaper, J.B., Karp, P.D., Neidhardt, F.C., Nystroem, T., Slauch, J.M., Squires, C.L., Ussery, D., Washington, DC
<4>
<5>1-51
<6>2009
<7>DNA methylation in bacteria is most often thought of in its role to protect DNA from
restriction endonucleases.  In addition o this role, however, studies in Escherichia coli,
Salmonella enteric serovar Typhimurium (referred to as serovar Typhimurium hereafter), and
Caulobacter crescentus have shown that methylated bases have other biological functions.  In
these cases, the methylated bases are not part of a restriction/modification system and the
enzymes that produce them are often referred to as orphan or solitary DNA methyltransferases. 
The postreplicative DNA methylation produced by this enzyme  superimposes on the primary DNA
sequence secondary information that has significance for DNA transactions such as
transcription, transposition, initiation of chromosome replication, mRNA utilization, and
prevention of mutations by DNA repair.  These alterations are brought about in two ways, the
first being simply a change in the steady-state level of the methyltransferase either up or
down from normal.  The second mechanism is through the configuration of the nucleotide
sequence subject to methylation; it can exist as symmetrically methylated, unmethylated, or
two possible hemi-methylated arrangements.  The details about the changes in DNA transactions
through alteration of methyltransferase levels or state of methylation sequences form the bulk
of this review.

<>

<1>Marinus, M.G., Morris, N.R.
<2>Biological function for 6-methyladenine residues in the DNA of Escherichia coli K12.
<3>J. Mol. Biol.
<4>85
<5>309-322
<6>1974
<7>A strain of Escherichia coli K12 mutant at the dam-3 site contains 0.08 mole % 6-methyl
adenine as compared to 0.5 mole % in the wild type, and the residual DNA methylation is not
due to the K12 modification methylase specified by the hsp genes. The dam-3 mutant is more
sensitive to ultraviolet irradiation and to mitomycin C than the wild type and also shows a
higher mutability. DNA isolated from the dam-3 mutant contains single stranded breaks that are
amplified in dam-3 polA12 and dam-3 lig-7 double mutants. A function of dam-specified 6-methyl
adenine residues in DNA would, therefore, appear to be the protection of DNA from nuclease(s)
that causes the development of breaks. Combination of dam-3 with polA,recA,recB and recC is
lethal.

<>

<1>Marinus, M.G., Morris, N.R.
<2>Pleiotropic effects of a DNA adenine methylation mutation (dam-3) in Escherichia coli K12.
<3>Mutat. Res.
<4>28
<5>15-26
<6>1975
<7>The dam-3 mutation results in a five-fold reduction in the number of 6-methyladenine (6-meA)
residues in the DNA of E. coli K12 or phage lambda. The DNA of phage fd appears to be devoid
of 6-meA when propagated on dam-3 bacteria. The phenotypic differences between dam-3 and dam+
bacteria include: (1) increased free phage in lysogenic dam-3 cultures, (2) increased
sensitivity to methyl methanesulfonate (MMS), (3) inviability of dam-3 lex-I strains, (4)
lower molecular weight of DNA in dam-3 bacteria in the absence of DNA ligase and (5) increased
rate of DNA degradation in dam-3 recA strains.

<>

<1>Marinus, M.G., Morris, N.R.
<2>Isolation of deoxyribonucleic acid methylase mutants of Escherichia coli K-12.
<3>J. Bacteriol.
<4>114
<5>1143-1150
<6>1973
<7>Fourteen deoxyribonucleic acid (DNA) and 10 ribonucleic acid (RNA) methylation mutants were
isolated from Escherichia coli K-12 by examining the ability of nucleic acids prepared from
clones of unselected mutagenized cells to accept methyl groups from wild-type crude extract.
Eleven of the DNA methylation mutants were deficient in 5-methylcytosine (5-MeC) and were
designated Dcm. Three DNA methylation mutants were deficient in N6-methyladenine (N6-MeA) and
were designated Dam. Extracts of the mutants were tested for DNA-cytosine:S-adenosylmethione
and DNA-adenine:S-adenosylmethionine methyltransferase activities. With one exception, all of
the mutants had reduced or absent activity. A representative Dcm mutation was located at 36 to
37 min and a representative Dam mutation was located in the 60- to 66-min region on the
genetic map. The Dcm mutants had no obvious associated phenotypic abnormality. The Dam mutants
were defective in their ability to restrict lambda. None of the mutations had the effect of
being lethal.

<>

<1>Marinus, M.G., Poteete, A., Arraj, J.A.
<2>Correlation of DNA adenine methylase activity with spontaneous mutability in Escherichia coli K-12.
<3>Gene
<4>28
<5>123-125
<6>1984
<7>Using a multicopy plasmid in which the tac promoter has been placed in front of the dam gene
of Escherichia coli K-12, we show that levels of DNA adenine methylase activity are correlated
with the spontaneous mutation frequency.

<>

<1>Mark, K.-K., Studier, F.W.
<2>Purification of the gene 0.3 protein of bacteriophage T7, an inhibitor of the DNA restriction system of Escherichia coli.
<3>J. Biol. Chem.
<4>256
<5>2573-2578
<6>1981
<7>The gene 0.3 protein of bacteriophage T7 prevents the DNA restriction system of
Escherichia coli from interfering with T7 infection.  A mutant strain of T7
that greatly overproduces the 0.3 protein has been constructed and used for
purification of this protein.  The 0.3 protein was found to be extremely acidic
and can be separated from virtually all other proteins of the infected cell by
chromatography on DEAE-cellulose.  Residual contaminating proteins and nucleic
acids can be removed by gel filtration, but an even simpler final purification
is possible, because under appropriate conditions the 0.3 protein is soluble in
high concentrations of ethanol.  Thus, a simple, essentially two-step
purification can produce about 50 mg of pure 0.3 protein from 30 liters of
culture.  The purified protein appears to be a dimer of identical subunits.  As
expected from its known function during infection, the purified 0.3 protein
inhibits the nuclease and ATPase activities of partially purified EcoB, the DNA
restriction enzyme of E. coli B, but it does not interfere with several
different type II restriction endonucleases tested.  The inhibition of EcoB
appears to require stoichiometric rather than catalytic amounts of 0.3 protein.

<>

<1>Markie, D., Tvrdeich, G., Hill, D.F., Poulter, R.
<2>The response of Type II restriction endonucleases to single base pair mismatches in heteroduplex DNA.
<3>Proc. Univ. Otago Med. Sch.
<4>64
<5>13-14
<6>1986
<7>The Type II restriction endonucleases cleave double-stranded DNA at specific sites within
short recognition sequences and have proven invaluable in the analysis and manipulation of
genetic material.  In contrast, the Type I endonucleases, which cleave DNA non-specifically at
varying distances from their recognition sequences, have played only a minor role in modern
molecular genetics.  Factors affecting DNA recognition and cleavage have been carefully
defined.  The resistance of mismatched heteroduplex DNA to digestion was initially reported
for a Type I enzyme (EcoBI) and this has since been confirmed for a single Type II enzyme
(EcoRI).  In this paper we extend this finding to four more Type II restriction enzymes,
BamHI, AccI, KpnI and SmaI, using a novel assay for digestion of heteroduplex DNA.

<>

<1>Marks, P., McGeehan, J., Kneale, G.
<2>A novel strategy for the expression and purification of the DNA methyltransferase, M.AhdI.
<3>Protein Expr. Purif.
<4>37
<5>236-242
<6>2004
<7>Biochemical and structural studies of the methylase from the type 1 1/2 R-M system AhdI
require the ability to purify this multisubunit enzyme
in significant quantities in a soluble and active form. Several
Escherichia coli expression systems were tested for their ability to
produce the intact methylase but this could not be achieved in a simple
co-expression system. Expression experiments were optimised to produce
high yields of soluble M and S subunits as individual proteins.
Temperature and conditions of induction proved to be the most useful
factors and although purification of the S subunit was successful, an
efficient strategy for the M subunit remained elusive. A novel strategy
was developed in which individual subunits are expressed separately and
the bacterial cells mixed before lysis. This method produced a high
yield of the multi-subunit methylase when purified to homogeneity by
means of heparin and size-exclusion chromatography. It was found to be
essential, however, to remove tightly bound DNA by ammonium sulphate
precipitation in 1 M NaCl. The intact methylase can now be consistently
produced, avoiding the use of fusion proteins. The purified enzyme is
stable over long time periods, unlike the individual subunits. This
method may be of general application where the expression of
multi-subunit proteins, or indeed their individual components, is
problematic.

<>

<1>Marks, P., McGeehan, J., Wilson, G., Errington, N., Kneale, G.
<2>Purification and characterization of a novel DNA methyltransferase, M.AhdI.
<3>Nucleic Acids Res.
<4>31
<5>2803-2810
<6>2003
<7>We have cloned the M and S genes of the restriction-modification (R-M) system AhdI and have
purified the resulting methyltransferase to
homogeneity. M.AhdI is found to form a 170 kDa tetrameric enzyme having a
subunit stoichiometry M2S2 (where the M and S subunits are responsible for
methylation and DNA sequence specificity, respectively). Sedimentation
equilibrium experiments show that the tetrameric enzyme dissociates to
form a heterodimer at low concentration, with K(d) approximately 2 microM.
The intact (tetrameric) enzyme binds specifically to a 30 bp DNA duplex
containing the AhdI recognition sequence GACN5GTC with high affinity (K(d)
approximately 50 nM), but at low enzyme concentration the DNA binding
activity is governed by the dissociation of the tetramer into dimers,
leading to a sigmoidal DNA binding curve. In contrast, only non-specific
binding is observed if the duplex lacks the recognition sequence.
Methylation activity of the purified enzyme was assessed by its ability to
prevent restriction by the cognate endonuclease. The subunit structure of
the M.AhdI methyltransferase resembles that of type I MTases, in contrast
to the R.AhdI endonuclease which is typical of type II systems. AhdI
appears to be a novel R-M system with properties intermediate between
simple type II systems and more complex type I systems, and may represent
an intermediate in the evolution of R-M systems.

<>

<1>Marquez, V.E., Eritja, R., Kelley, J.A., Vanbemmel, D., Christman, J.
<2>Potent inhibition of HhaI DNA methylase by the aglycon of 2-(1H)-pyrimidinone riboside (Zebularine) at the G(C)under-barGC recognition domain.
<3>Therapeutic Oligonucleotides
<4>1002
<5>154-164
<6>2003
<7>A short oligodeoxynucleotide (ODN) with 2-(1H)-pyrimidinone at the HhaI DNA methyltransferase
target site (GCGC) is shown to induce a level of inhibition of methyl transfer and thermal
stability of the complex with the enzyme identical to that achieved with a similar ODN
substituted with 5-azacytosine. The drugs responsible for these effects-zebularine and
5-azacytidine/2'-deoxy-5-azacytidine-are contrasted in terms of chemical stability and
possible metabolic activation by a brief structure-activit analysis.

<>

<1>Marquez, V.E., Eritja, R., Kelley, J.A., Vanbemmel, D., Christman, J.K.
<2>Potent inhibition of HhaI DNA methylase by the aglycon of 2-(1H)-pyrimidinone riboside (Zebularine) at the GCGC recognition domain.
<3>Ann. NY Acad. Sci.
<4>1002
<5>154-164
<6>2003
<7>A short oligodeoxynucleotide (ODN) with 2-(1H)-pyrimidinone at the HhaI DNA methyltransferase
target site (GCGC) is shown to induce a level of
inhibition of methyl transfer and thermal stability of the complex with
the enzyme identical to that achieved with a similar ODN substituted with
5-azacytosine. The drugs responsible for these effects-zebularine and
5-azacytidine/2'-deoxy-5-azacytidine-are contrasted in terms of chemical
stability and possible metabolic activation by a brief structure-activity
analysis.

<>

<1>Marquez, V.E., Goddard, A., Alvarez, E., Ford, H. Jr., Christman, J.K., Sheikhnejad, G., Brank, A., Marasco, C.J., Suffrin, J.R., O'Gara, M., Cheng, X.
<2>Oligonucleotides containing 5,6-dihydro-5-azacytosine at CpG sites can produce potent inhibition of DNA cytosine-C5-methyltransferase without covalently binding to the enzyme.
<3>Antisense Nucleic Acid Drug Dev.
<4>9
<5>415-421
<6>1999
<7>C5-cytosine methylation of DNA at CpG sites is catalyzed by DNA cytosine-C5-methyltransferase
in both prokaryotes and eukaryotes.  The mechanism of transfer of the methyl group from the
cofactor S-adenosyl-L-methionine to the target cytosine occurs in a similar fashion in most
(if not all) C5-MTases, which share highly conserved sequence motifs in the catalytic and
AdoMet binding regions.  In mammals, the enzyme is responsible for the maintenance of
methylation patterns in the genome.  However, aberrant DNA methylation patterns are associated
with tumorigenesis and are common in cancer.

<>

<1>Marquez, V.E., Wang, P.Y., Nicklaus, M.C., Maier, M., Manoharan, M., Christman, J.K., Banavali, N.K., Mackerell, A.D.
<2>Inhibition of (cytosine C5)-methyltransferase by oligonucleotides containing flexible (cyclopentane) and conformationally constrained  (bicyclo[3.1.0]hexane) abasic sites.
<3>Nucleosides Nucleotides Nucleic Acids
<4>20
<5>451-459
<6>2001
<7>Pseudorotationally locked sugar analogues based on bicyclo[3.1.0]-hexane templates were placed
in DNA duplexes as abasic target sites in the M.HhaI recognition sequence. The binding
affinity
of the enzyme increases when the abasic site is constrained to the
South conformation and decreases when it is constrained to the North
conformation. A structural understanding of these differences is
provided.

<>

<1>Marra, A., Shuman, H.A.
<2>Isolation of a Legionella pneumophila restriction mutant with increased ability to act as a recipient in heterospecific matings.
<3>J. Bacteriol.
<4>171
<5>2238-2240
<6>1989
<7>The ability of Legionella pneumophila to act as a recipient of IncP and IncQ
plasmids in matings with Escherichia coli varies widely from strain to strain.
We found that the low efficiency of mating of the Philadelphia-1 strain is due
to a type II restriction-modification system, and we isolated and characterized
a Philadelphia-1 mutant that lacks the restriction enzyme activity.

<>

<1>Marrero, R., Welkos, S.L.
<2>The transformation frequency of plasmids into Bacillus anthracis is affected by adenine methylation.
<3>Gene
<4>152
<5>75-78
<6>1995
<7>Plasmids pLTV1 and pHV33, capable of replicating in both Gram+ and Gram- bacterial hosts
(shuttle vectors), when derived from the Escherichia coli strain HB101, were inactive in an
electro-transformation assay employing the Bacillus anthracis strains delta Ames-1 and delta
V1B-1 as recipients.  The same plasmids isolated from the DNA methyltransferase
(MTase)-deficient E. coli strain GM2929 (dam,dcm), were able to transform the B. anthracis
strains at a frequency of 10/2-10/3 transformants/ug of plasmid DNA.  Efficient transformation
was also obtained when the plasmids were propagated in strains of B. subtilis 168 (10/2-10/4
transformants/ug of plasmid DNA).  The B. subtilis strains used are known to harbor
restriction/modification systems that recognize cytosine as a target for methylation.  In
contrast, no adenine methylation activities have been reported for these strains.  The data
presented indicate that DNA containing methylated adenine residues is restricted in the B.
anthracis strains studied here, resulting in decreased plasmid DNA-mediated transformation
frequencies.  This inhibition could be alleviated by propagating plasmid species in
MTase-deficient (dam) strains of E. coli or B. subtilis 168, before their introduction into
strains of B. anthracis.

<>

<1>Marsan, D., Wommack, K.E., Ravel, J., Chen, F.
<2>Draft Genome Sequence of Synechococcus sp. Strain CB0101, Isolated From the Chesapeake Bay Estuary.
<3>Genome Announcements
<4>2
<5>e01111-13
<6>2014
<7>Here, we report the draft genome sequence of the estuarine Synechococcus sp. strain CB0101.
The genomics information of this strain will facilitate the study
of the poorly understood Synechococcus subcluster 5.2 and how this strain is
capable of thriving in a dynamic estuarine system, such as the Chesapeake Bay.

<>

<1>Marshall, J.J., Gowers, D.M., Halford, S.E.
<2>Restriction Endonucleases that Bridge and Excise Two Recognition Sites from DNA.
<3>J. Mol. Biol.
<4>367
<5>419-431
<6>2007
<7>Most restriction endonucleases bridge two target sites before cleaving DNA: examples include
all of the translocating Type I and Type III
systems, and many Type II nucleases acting at their sites. A subset of
Type II enzymes, the IIB systems, recognise bipartite sequences, like Type
I sites, but cut specified phosphodiester bonds near their sites, like
Type IIS enzymes. However, they make two double-strand breaks, one either
side of the site, to release the recognition sequence on a short DNA
fragment; 34 bp long in the case of the archetype, BcgI. It has been
suggested that BcgI needs to interact with two recognition sites to cleave
DNA but whether this is a general requirement for Type IIB enzymes had yet
to be established. Ten Type IIB nucleases were tested against DNA
substrates with one or two copies of the requisite sequences. With one
exception, they all bridged two sites before cutting the DNA, usually in
concerted reactions at both sites. The sites were ideally positioned in
cis rather than in trans and were bridged through 3-D space, like Type II
enzymes, rather than along the 1-D contour of the DNA, as seen with Type I
enzymes. The standard mode of action for the restriction enzymes that
excise their recognition sites from DNA thus involves concurrent action at
two DNA sites.

<>

<1>Marshall, J.J., Smith, R.M., Ganguly, S., Halford, S.E.
<2>Concerted action at eight phosphodiester bonds by the BcgI restriction endonuclease.
<3>Nucleic Acids Res.
<4>39
<5>7630-7640
<6>2011
<7>The BcgI endonuclease exemplifies a subset of restriction enzymes, the Type IIB class, which
make two double-strand breaks (DSBs) at each copy of
their recognition sequence, one either side of the site, to excise the
sequence from the remainder of the DNA. In this study, we show that BcgI
is essentially inactive when bound to a single site and that to cleave a
DNA with one copy of its recognition sequence, it has to act in trans,
bridging two separate DNA molecules. We also show that BcgI makes the two
DSBs at an individual site in a highly concerted manner. Intermediates cut
on one side of the site do not accumulate during the course of the
reaction: instead, the DNA is converted straight to the final products cut
on both sides. On DNA with two sites, BcgI bridges the sites in cis and
then generally proceeds to cut both strands on both sides of both sites
without leaving the DNA. The BcgI restriction enzyme can thus excise two
DNA segments together, by cleaving eight phosphodiester bonds within a
single-DNA binding event.

<>

<1>Marshall, J.J.T., Halford, S.E.
<2>The Type IIB restriction endonucleases.
<3>Biochem. Soc. Trans.
<4>38
<5>410-416
<6>2010
<7>The endonucleases from the Type IIB restriction-modification systems differ from all other
restriction enzymes. The Type IIB enzymes cleave
both DNA strands at specified locations distant from their recognition
sequences, like Type IIS nucleases, but they are unique in that they do
so on both sides of the site, to liberate the site from the remainder
of the DNA on a short duplex. The fact that these enzymes cut DNA at
specific locations mark them as Type II systems, as opposed to the Type
I enzymes that cut DNA randomly, but in terms of gene organization and
protein assembly, most Type IIB restriction-modification systems have
more in common with Type I than with other Type II systems. Our current
knowledge of the Type IIB systems is reviewed in the present paper.

<>

<1>Marshall, P., Davis, T.B., Lemieux, C.
<2>The I-CeuI endonuclease: purification and potential role in the evolution of Chlamydomonas group I introns.
<3>Eur. J. Biochem.
<4>220
<5>855-859
<6>1994
<7>During genetic crosses between the interfertile green algae Chlamydomonas eugametos and
Chlamydomonas moewusii, the I-CeuI endonuclease encoded by the fifth group I intron (CeLSU.5)
in the C. eugametos chloroplast large subunit rRNA gene mediates the mobility of this intron
by introducing a double-strand break near the insertion site of the intron in the
corresponding C. moewusii intronless allele.  To characterize the biochemical properties of
this endonuclease, we have purified I-CeuI and determined the optimal reaction conditions for
cleavage.  I-CeuI activity is maximal at 50oC, pH 10.0, 2.5 mM MgCl2 and in the absence of
NaCl.  Unlike the well-characterized I-SceI endonuclease, I-CeuI remains stable following
preincubation in the absence of substrate.  We discuss the role that homing endonucleases may
have played in the evolution of Chlamydomonas chloroplast group I introns.

<>

<1>Marshall, P., Lemieux, C.
<2>The I-CeuI endonuclease recognizes a sequence of 19 base pairs and preferentially cleaves the coding strand of the Chlamydomonas moewusii chloroplast large subunit rRNA gene.
<3>Nucleic Acids Res.
<4>20
<5>6401-6407
<6>1992
<7>The I-CeuI endonuclease is a member of the growing family of homing endonucleases that
catalyse mobility of group I introns by making a double-strand break at the homing site of
these introns in cognate intronless alleles during genetics crosses. In a previous study, we
have shown that a short DNA fragment of 26 bp, encompassing the homing site of the fifth
intron in the Chlamydomonas eugametos chloroplast large subunit rRNA gene (Ce LSU.5), was
sufficient for I-CeuI recognition and cleavage. Here, we report the recognition sequence of
the I-CeuI endonuclease, as determined by random mutagenesis of nucleotide positions adjacent
to the I-CeuI cleavage site. Single-base substitutions that completely abolish endonuclease
activity delimit a 15-bp sequence whereas those that reduce the cleavage rate define a 19-bp
sequence that extends from position -7 to position +12 with respect to the Ce LSU.5 intron
insertion site. As the other homing endonucleases that have been studied so far, the I-CeuI
endonuclease recognizes a non-symmetric degenerate sequence. The top strand of the recognition
sequence is preferred for I-CeuI cleavage and the bottom strand most likely determines the
rate of double-strand breaks.

<>

<1>Marshall, P., Lemieux, C.
<2>Cleavage pattern of the homing endonuclease encoded by the fifth intron in the chloroplast large subunit rRNA-encoding gene of Chlamydomonas eugametos.
<3>Gene
<4>104
<5>241-245
<6>1991
<7>The fifth group-I intron in the chloroplast large subunit rRNA-encoding gene of Chlamydomonas
eugametos (CeLSU.5) is mobile during interspecific crosses between C. eugametos and
Chlamydomonas moewusii. Like the six other mobile introns that have been well characterized so
far, CeLSU.5 contains a long open reading frame (ceuIR) coding for a site-specific
endonuclease (I-CeuI) that cleaves the C. moewsuii intronless gene in the vicinity of the
intron-insertion site. This stimulates gap repair and mediates efficient transfer of the
intron at its cognate site. By expressing the ceuIR gene in the Escherichia coli vectors
pKK233-2 and pTRC-99A, we recently demonstrated that the endonuclease is highly toxic to E.
coli. To eliminate this problem and characterize the cleavage pattern and recognition sequence
of the I-CeuI endonuclease, we have expressed the ceuIR gene in E. coli under the control of a
bacteriophage T7 promoter in a tightly regulated M13 system, and developed an in vitro system
to assay partially purified I-CeuI activity. This allowed us to determine that I-CeuI
recognizes a sequence of less than 26 bp centered around the insertion site and produces a
staggered cut 5 bp downstream from this site, yielding 4-nucleotide (CTAA), 3'-OH overhangs.

<>

<1>Marshall, P., Lemieux, C., Gauthier, A., Turmel, M.
<2>Homing endonuclease which originates from Chlamydomonas eugametos and recognizes and cleaves A 15, 17 or 19 degenerate double stranded nucleotide sequence.
<3>US Patent Office
<4>US 5420032
<5>
<6>1995
<7>The present invention relates to a homing endonuclease which originates from Chlamydomonas
eugametos, and was overproduced in E. coli, purified and characterized.  The homing
endonuclease of the present invention recognizes and cleaves degenerate double-stranded DNA at
a specific recognition site; it particularly recognizes and cleaves 15, 17 and 19 nucleotide
sequences.  The cleavage of target DNA by this endonuclease produces a 4 nucleotide extension
with a 3' OH overhang.  A method to use the endonuclease of the present invention to cleave
DNA fragments useful for gene mapping is also disclosed.

<>

<1>Marston, M.F., Pierciey, F.J. Jr., Shepard, A., Gearin, G., Qi, J., Yandava, C., Schuster, S.C., Henn, M.R., Martiny, J.B.
<2>Rapid diversification of coevolving marine Synechococcus and a virus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>109
<5>4544-4549
<6>2012
<7>Marine viruses impose a heavy mortality on their host bacteria, whereas at the
same time the degree of viral resistance in marine bacteria appears to be high.
Antagonistic coevolution-the reciprocal evolutionary change of interacting
species-might reconcile these observations, if it leads to rapid and dynamic
levels of viral resistance. Here we demonstrate the potential for extensive
antagonistic coevolution between the ecologically important marine cyanobacterium
Synechococcus and a lytic virus. In a 6-mo-long replicated chemostat experiment,
Synechococcus sp. WH7803 and the virus (RIM8) underwent multiple coevolutionary
cycles, leading to the rapid diversification of both host and virus. Over the
course of the experiment, we detected between 4 and 13 newly evolved viral
phenotypes (differing in host range) and between 4 and 11 newly evolved
Synechococcus phenotypes (differing in viral resistance) in each chemostat.
Genomic analysis of isolates identified several candidate genes in both the host
and virus that might influence their interactions. Notably, none of the viral
candidates were tail fiber genes, thought to be the primary determinants of host
range in tailed bacteriophages, highlighting the difficulty in generalizing
results from bacteriophage infecting gamma-Proteobacteria. Finally, we show that
pairwise virus-host coevolution may have broader community consequences;
coevolution in the chemostat altered the sensitivity of Synechoccocus to a
diverse suite of viruses, as well as the virus' ability to infect additional
Synechococcus strains. Our results indicate that rapid coevolution may contribute
to the generation and maintenance of Synechococcus and virus diversity and
thereby influence viral-mediated mortality of these key marine bacteria.

<>

<1>Marti, R., Hagens, S., Loessner, M.J., Klumpp, J.
<2>Genome Sequence of Salmonella bongori Strain N268-08.
<3>Genome Announcements
<4>1
<5>e01018-13
<6>2013
<7>Volume 1, no. 4, e00580-13, 2013. Page 1: The article title should read as given above. After
the publication of this article, we were
made aware of the fact that strain N268-08 was not isolated from a clinical sample.

<>

<1>Marti, R., Hagens, S., Loessner, M.J., Klumpp, J.
<2>Genome Sequence of Salmonella bongori Strain N268-08, a Rare Clinical Isolate.
<3>Genome Announcements
<4>1
<5>e00580-13
<6>2013
<7>Salmonella bongori is a close relative of the highly virulent members of S. enterica
subspecies enterica, encompassing more than 2,500 serovars, most of
which cause human salmonellosis, one of the leading food-borne illnesses. S.
bongori is only very rarely implicated in infections. We here present the
sequence of a clinical isolate from Switzerland, S. bongori strain N268-08.

<>

<1>Martienssen, R.
<2>Chipping away at chromatin.
<3>Nat. Genet.
<4>27
<5>240-241
<6>2001
<7>When DNA-binding proteins are tethered to dam methylase from Escherichia coli. adenine
methylation is directed to eukaryotic target
sites in vivo. Hybridization of methylated DNA to microarrays allows
binding sites to be displayed genome-wide, providing a versatile
alternative to chromatin immunoprecipitation.

<>

<1>Martienssen, R.A., Colot, V.
<2>DNA methylation and epigenetic inheritance in plants and filamentous fungi.
<3>Science
<4>293
<5>1070-1074
<6>2001
<7>Plants and filamentous fungi share with mammals enzymes responsible for DNA methylation. In
these organisms, DNA methylation is associated with gene silencing and transposon control.
However, plants and fungi differ from mammals in the genomic distribution, sequence
specificity, and heritability of methylation. We consider the role that transposons play in
establishing methylation patterns and the epigenetic consequences of their perturbation.

<>

<1>Martienssen, R.A., Richards, E.J.
<2>DNA methylation in eukaryotes.
<3>Curr. Opin. Genet. Dev.
<4>5
<5>234-242
<6>1995
<7>Recent advances have expanded our understanding of the processes underlying the establishment,
maintenance, and elaboration of DNA methylation patterns in eukaryotes.  The functional
significance of DNA methylation is sought in a comparison of results on a variety of
epigenetic phenomena in different eukaryotes.  The recent development of DNA methylation
mutants in mice, Neurospora, and Arabadopsis will allow traditional genetic dissection to be
applied to long-standing problems regarding the function and regulation of eukaryotic DNA
methylation.  Although methylation appears to be important for maintenance of different
epigenetic states, the mechanism that establishes these states is likely to involve additional
processes.

<>

<1>Martin, A.M., Horton, N.C., Luseti, S., Reich, N.O., Perona, J.J.
<2>Divalent metal dependence of site-specific DNA binding by EcoRV endonuclease.
<3>Biochemistry
<4>38
<5>8430-8439
<6>1999
<7>Measurements of binding equilibria of EcoRV endonuclease to DNA, for a series of base-analogue
substrates, demonstrate that expression of sequence selectivity is strongly enhanced by the
presence of Ca2+ ions. Binding constants were determined for short duplex
oligodeoxynucleotides containing the cognate DNA site, three cleavable noncognate sites, and a
fully nonspecific site. At pH 7.5 and 100 mM NaCl, the full range of specificity from the
specific (tightest binding) to nonspecific (weakest binding) sites is 0.9 kcal/mol in the
absence of metal ions and 5.8 kcal/mol in the presence of Ca2+. Precise determination of
binding affinities in the presence of the active Mg2+ cofactor was found to be possible for
substrates retaining up to 1.6% of wild-type activity, as determined by the rate of phosphoryl
transfer. These measurements show that Ca2+ is a near-perfect analogue for Mg2+ in binding
reactions of the wild-type enzyme with DNA base-analogue substrates, as it provides identical
DeltaDeltaG degrees bind values among the cleavable noncognate sites. Equilibrium dissociation
constants of wild-type and base-analogue sites were also measured for the weakly active EcoRV
mutant K38A, in the presence of either Mg2+ or Ca2+. In this case, Ca2+ allows expression of a
greater degree of specificity than does Mg2+. DeltaDeltaG degrees/bind values of K38A toward
specific versus nonspecific sites are 6.1 kcal/mol with Ca2+ and 3.9 kcal/mol with Mg2+,
perhaps reflecting metal-specific conformational changes in the ground-state ternary
complexes. The enhancement of binding specificity provided by divalent metal ions is likely to
be general to many restriction endonucleases and other metal-dependent nucleic acid-modifying
enzymes. These results strongly suggest that measurements of DNA binding affinities for EcoRV,
and likely for many other restriction endonucleases, should be performed in the presence of
divalent metal ions.

<>

<1>Martin, A.M., Sam, M.D., Reich, N.O., Perona, J.J.
<2>Structural and energetic origins of indirect readout in site-specific DNA cleavage by a restriction endonuclease.
<3>Nat. Struct. Biol.
<4>6
<5>269-277
<6>1999
<7>Specific recognition by EcoRV endonuclease of its cognate, sharply bent GATATC site at the
center TA step occurs solely via hydrophobic interaction with thymine methyl groups.
Mechanistic kinetic analyses of base analog-substituted DNAs at this position reveal that
direct readout provides 5 kcal mol(-1) toward specificity, with an additional 6-10 kcal
mol(-1) arising from indirect readout. Crystal structures of several base analog complexes
show that the major-groove hydrophobic contacts are crucial to forming required divalent
metal-binding sites, and that indirect readout operates in part through the sequence-dependent
free-energy cost of unstacking the center base-pair step of the DNA.

<>

<1>Martin, J.L., McMillan, F.M.
<2>SAM (dependent) I AM: the S-adenosylmethionine-dependent methyltransferase fold.
<3>Curr. Opin. Struct. Biol.
<4>12
<5>783-793
<6>2002
<7>The S-adenosylmethionine-dependent methyltransferase enzymes share little sequence identity,
but incorporate a highly conserved structural
fold. Surprisingly, residues that bind the common cofactor are poorly
conserved, although the binding site is localised to the same region of
the fold. The substrate-binding region of the fold varies enormously.
Over the past two years, there has been a significant increase in the
number of structures that are known to incorporate this fold, including
several uncharacterised proteins and two proteins that lack
methyltransferase activity.

<>

<1>Martin, P., Boulukos, K.E., Pognonec, P.
<2>REtools: a laboratory program for restriction enzyme work: enzyme selection and reaction condition assistance.
<3>BMC Bioinformatics
<4>7
<5>98
<6>2006
<7>ABSTRACT : BACKGROUND : Restriction enzymes are one of the everyday tools used in molecular
biology. The continuously expanding panel of known
restriction enzymes (several thousands) renders their optimal use
virtually impossible without computerized assistance. Several
manufacturers propose on-line sites that assist scientists in their
restriction enzyme work, however, none of these sites meet all the actual
needs of laboratory workers, and they do not take into account the enzymes
actually present in one's own laboratory. RESULTS : Using FileMaker Pro,
we developed a stand-alone application which can run on both PCs and
Macintoshes. We called it REtools, for Restriction Enzyme tools. This
program, which references all currently known enzymes (>3500), permits the
creation and update of a personalized list of restriction enzymes actually
available in one's own laboratory. Upon opening the program, scientists
will be presented with a user friendly interface that will direct them to
different menus, each one corresponding to different situations that
restriction enzyme users commonly encounter. We particularly emphasized
the ease of use to make REtools a solution that laboratory members would
actually want to use. CONCLUSION : REtools, a user friendly and easily
customized program to organize any laboratory enzyme stock, brings a
software solution that will make restriction enzyme use and reaction
condition determination straightforward and efficient. The usually
unexplored potential of isoschizomers also becomes accessible to all,
since REtools proposes all possible enzymes similar to the one(s) chosen
by the user. Finally, many of the commonly overlooked subtleties of
restriction enzyme work, such as methylation requirement, unusual reaction
conditions, or the number of flanking bases required for cleavage, are
automatically provided by REtools.

<>

<1>Martin, R.N.A., McGeehan, J.E., Ball, N.J., Streeter, S.D., Thresh, S.J., Kneale, G.G.
<2>Structural analysis of DNA-protein complexes regulating the restriction-modification system Esp1396I.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>69
<5>962-966
<6>2013
<7>The controller protein of the type II restriction-modification (RM) system Esp1396I binds to
three distinct DNA operator sequences upstream
of the methyltransferase and endonuclease genes in order to regulate
their expression. Previous biophysical and crystallographic studies
have shown molecular details of how the controller protein binds to the
operator sites with very different affinities. Here, two protein-DNA
co-crystal structures containing portions of unbound DNA from native
operator sites are reported. The DNA in both complexes shows
significant distortion in the region between the conserved symmetric
sequences, similar to that of a DNA duplex when bound by the controller
protein (C-protein), indicating that the naked DNA has an intrinsic
tendency to bend when not bound to the C-protein. Moreover, the width
of the major groove of the DNA adjacent to a bound C-protein dimer is
observed to be significantly increased, supporting the idea that this
DNA distortion contributes to the substantial cooperativity found when
a second C-protein dimer binds to the operator to form the tetrameric
repression complex.

<>

<1>Martin, R.N.A., McGeehan, J.E., Kneale, G.
<2>Structural and Mutagenic Analysis of the RM Controller Protein C.Esp1396I.
<3>PLoS ONE
<4>9
<5>e98365
<6>2014
<7>Bacterial restriction-modification (RM) systems are comprised of two complementary enzymatic
activities that prevent the establishment of foreign DNA in a bacterial cell: DNA methylation
and DNA restriction. These two activities are tightly regulated to prevent over-methylation or
auto-restriction. Many Type II RM systems employ a controller (C) protein as a transcriptional
regulator for the endonuclease gene (and in some cases, the methyltransferase gene also). All
high-resolution structures of C-protein/DNA-protein complexes solved to date relate to
C.Esp1396I, from which the interactions of specific amino acid residues with DNA bases and/or
the phosphate backbone could be observed. Here we present both structural and DNA binding data
for a series of mutations to the key DNA binding residues of C.Esp1396I. Our results indicate
that mutations to the backbone binding residues (Y37, S52) had a lesser affect on DNA binding
affinity than mutations to those residues that bind directly to the bases (T36, R46), and the
contributions of each side chain to the binding energies are compared. High-resolution X-ray
crystal structures of the mutant and native proteins showed that the fold of the proteins was
unaffected by the mutations, but also revealed variation in the flexible loop conformations
associated with DNA sequence recognition. Since the tyrosine residue Y37 contributes to DNA
bending in the native complex, we have solved the structure of the Y37F mutant protein/DNA
complex by X-ray crystallography to allow us to directly compare the structure of the DNA in
the mutant and native complexes.

<>

<1>Martin, V., Cardenas, N., Jimenez, E., Maldonado, A., Rodriguez, J.M., Fernandez, L.
<2>Genome Sequence of Lactobacillus gastricus PS3, a Strain Isolated from Human Milk.
<3>Genome Announcements
<4>1
<5>e00489-13
<6>2013
<7>Lactobacillus gastricus is a mostly unknown lactobacilli species associated with  mucosal
surfaces. We present the draft annotated genome sequence of L. gastricus
strain PS3, isolated from a human milk sample, to provide new insights into its
biology and to characterize those genes related to advantageous technological and
beneficial properties.

<>

<1>Martin, V., Maldonado-Barragan, A., Jimenez, E., Ruas-Madiedo, P., Fernandez, L., Rodriguez, J.M.
<2>Complete Genome Sequence of Streptococcus salivarius PS4, a Strain Isolated from  Human Milk.
<3>J. Bacteriol.
<4>194
<5>4466-4467
<6>2012
<7>Streptococcus salivarius is a commensal species commonly found in the human oropharyngeal
tract. Some strains of this species have been developed for use as
oral probiotics, while others have been associated with a variety of
opportunistic human infections. Here, we report the complete sequence of strain
PS4, which was isolated from breast milk of a healthy woman.

<>

<1>Martin-Cuadrado, A.B., Lopez-Garcia, P., Alba, J.C., Moreira, D., Monticelli, L., Strittmatter, A., Gottschalk, G., Rodriguez-Valera, F.
<2>Metagenomics of the deep mediterranean, a warm bathypelagic habitat.
<3>PLoS ONE
<4>2
<5>e914
<6>2007
<7>BACKGROUND: Metagenomics is emerging as a powerful method to study the
function and physiology of the unexplored microbial biosphere, and is
causing us to re-evaluate basic precepts of microbial ecology and
evolution. Most marine metagenomic analyses have been nearly exclusively
devoted to photic waters. METHODOLOGY/PRINCIPAL FINDINGS: We constructed a
metagenomic fosmid library from 3,000 m-deep Mediterranean plankton, which
is much warmer (approximately 14 degrees C) than waters of similar depth
in open oceans (approximately 2 degrees C). We analyzed the library both
by phylogenetic screening based on 16S rRNA gene amplification from clone
pools and by sequencing both insert extremities of ca. 5,000 fosmids.
Genome recruitment strategies showed that the majority of high scoring
pairs corresponded to genomes from Rhizobiales within the
Alphaproteobacteria, Cenarchaeum symbiosum, Planctomycetes, Acidobacteria,
Chloroflexi and Gammaproteobacteria. We have found a community structure
similar to that found in the aphotic zone of the Pacific. However, the
similarities were significantly higher to the mesopelagic (500-700 m deep)
in the Pacific than to the single 4000 m deep sample studied at this
location. Metabolic genes were mostly related to catabolism, transport and
degradation of complex organic molecules, in agreement with a prevalent
heterotrophic lifestyle for deep-sea microbes. However, we observed a high
percentage of genes encoding dehydrogenases and, among them, cox genes,
suggesting that aerobic carbon monoxide oxidation may be important in the
deep ocean as an additional energy source. CONCLUSIONS/SIGNIFICANCE: The
comparison of metagenomic libraries from the deep Mediterranean and the
Pacific ALOHA water column showed that bathypelagic Mediterranean
communities resemble more mesopelagic communities in the Pacific, and
suggests that, in the absence of light, temperature is a major stratifying
factor in the oceanic water column, overriding pressure at least over 4000
m deep. Several chemolithotrophic metabolic pathways could supplement
organic matter degradation in this most depleted habitat.

<>

<1>Martin-Cuadrado, A.B., Rodriguez-Valera, F., Moreira, D., Alba, J.C., Ivars-Martinez, E., Henn, M.R., Talla, E., Lopez-Garcia, P.
<2>Hindsight in the relative abundance, metabolic potential and genome dynamics of uncultivated marine archaea from comparative metagenomic analyses of bathypelagic plankton of different oceanic regions.
<3>ISME J.
<4>2
<5>865-886
<6>2008
<7>Marine planktonic archaea are widespread and abundant in deep oceanic waters but, despite
their obvious ecological importance, little is known about them. Metagenomic analyses of large
genome fragments allow access to both gene content and genome structure from single
individuals of these cultivation-reluctant organisms. We present the comparative analysis of
22 archaeal genomic clones containing 16S rRNA genes that were selected from four metagenomic
libraries constructed from meso- and bathypelagic plankton of different oceanic regions (South
Atlantic, Antarctic Polar Front,
Adriatic and Ionian Sea; depths from 500 to 3000 m). We sequenced clones of the divergent
archaeal lineages Group 1A (Crenarchaeota) and Group III (Euryarchaeota) as well as clones
from the more frequent Group I Crenarchaeota and Group II Euryarchaeota. Whenever possible, we
analysed
clones that had identical or nearly identical 16S rRNA genes and that were retrieved from
distant geographical locations, that is, that defined pan-oceanic operational taxonomic units
(OTUs). We detected genes involved in nitrogen fixation in Group 1A Crenarchaeota, and genes
involved in
carbon fixation pathways and oligopeptide importers in Group I Crenarchaeota, which could
confirm the idea that these are mixotrophic. A two-component system resembling that found in
ammoniaoxidizing bacteria was found in Group III Euryarchaeota, while genes for anaerobic
respiratory
chains were detected in Group II Euryarchaeota. Whereas gene sequence conservation was high,
and recombination and gene shuffling extensive within and between OTUs in Group I
Crenarchaeota, gene sequence conservation was low and global synteny maintained in Group II
Euryarchaeota. This implies remarkable differences in genome dynamics in Group I Crenarchaeota
and Group II Euryarchaeota with recombination and mutation being, respectively, the dominant
genome-shaping forces. These observations, along with variations in GC content, led us to
hypothesize that the two groups of organisms have fundamentally different lifestyles.

<>

<1>Martin-Laurent, F., Marti, R., Waglechner, N., Wright, G.D., Topp, E.
<2>Draft Genome Sequence of the Sulfonamide Antibiotic-Degrading Microbacterium sp.  Strain C448.
<3>Genome Announcements
<4>2
<5>e01113-13
<6>2014
<7>We report the draft genome sequence of Microbacterium sp. strain C448, isolated from
agricultural soil with a decade of exposure to veterinary antibiotics on the
basis of using sulfamethazine and other antibiotics as the sole sources of
carbon. The genome sequence revealed that strain C448 harbors several antibiotic
resistance genes, including sulI.

<>

<1>Martineau, C., Villeneuve, C., Mauffrey, F., Villemur, R.
<2>Complete Genome Sequence of Hyphomicrobium nitrativorans Strain NL23, a Denitrifying Bacterium Isolated from Biofilm of a Methanol-Fed Denitrification  System Treating Seawater at the Montreal Biodome.
<3>Genome Announcements
<4>2
<5>e01165-13
<6>2014
<7>Hyphomicrobium nitrativorans strain NL23 has been isolated from the biofilm of a
denitrification system treating seawater. This strain has the capacity to
denitrify using methanol as a carbon source. Here, we report the complete genome
sequence of this strain in an effort to increase understanding of the function of
this bacterium within the biofilm.

<>

<1>Martinet, N., Michel, B.Y., Bertrand, P., Benhida, R.
<2>Small molecules DNA methyltransferases inhibitors.
<3>MedChemComm
<4>3
<5>263-273
<6>2012
<7>Methylation catalyzed by the DNA methyltransferases affects the C5 position of cytosine
residues in DNA. This physiological process is
active from the embryo conception, throughout all its developmental
steps, and also later for the maintenance of the adult organism. Excess
methylated cytosine in tumor suppressor genes is a consistent hallmark
of human cancers. However, DNA methylation variation is now
acknowledged to significantly contribute to genetic and common
diseases. DNA methyltransferases became attractive therapeutic targets
as DNA demethylation, in vitro, brought cancer cell differentiation and
apoptosis. Inhibitors are already in use, alone or in combination, to
treat myeloid malignancies, while clinical assays are ongoing for other
diseases. DNA methylation and histone modifications are intimately
correlated with epigenetic heritable modifications of gene expression
that are independent of changes in the genetic sequence. Common
initiatives for epigenetic research have built public databases with
useful resources. The recent discovery of 5-hydroxymethyl cytosine has
added new questions and challenges for the epigenome community. We
review here knowledge about DNA methylation to provide researchers with
the information needed to make more active inhibitors for the benefit
of patients. Because of space limitations, many important works cannot
be cited. We refer the reader to reviews containing these references.

<>

<1>Martinez, R.J., Bruce, D., Detter, C., Goodwin, L.A., Han, J., Han, C.S., Held, B., Land, M.L., Mikhailova, N., Nolan, M., Pennacchio, L., Pitluck, S., Tapia, R., Woyke, T., Sobecky, P.A.
<2>Complete Genome Sequence of Rahnella sp. Strain Y9602, a Gammaproteobacterium Isolate from Metal- and Radionuclide-Contaminated Soil.
<3>J. Bacteriol.
<4>194
<5>2113-2114
<6>2012
<7>Rahnella sp. strain Y9602 is a gammaproteobacterium isolated from contaminated subsurface
soils that is capable of promoting uranium phosphate mineralization as
a result of constitutive phosphatase activity. Here we report the first complete
genome sequence of an isolate belonging to the genus Rahnella.

<>

<1>Martinez, R.J., Bruce, D., Detter, C., Goodwin, L.A., Han, J., Han, C.S., Held, B., Land, M.L., Mikhailova, N., Nolan, M., Pennacchio, L., Pitluck, S., Tapia, R., Woyke, T., Sobecky, P.A.
<2>Complete Genome Sequence of Rahnella aquatilis CIP 78.65.
<3>J. Bacteriol.
<4>194
<5>3020-3021
<6>2012
<7>Rahnella aquatilis CIP 78.65 is a gammaproteobacterium isolated from a drinking water source
in Lille, France. Here we report the complete genome sequence of Rahnella aquatilis CIP 78.65,
the type strain of R. aquatilis.

<>

<1>Martinez, V., Hormigo, D., Del Cerro, C., Gomez-de-Santos, P., Garcia-Hidalgo, J., Arroyo, M., Prieto, A., Garcia, J.L., de la Mata, I.
<2>Genome Sequence of Streptomyces exfoliatus DSMZ 41693, a Source of Poly(3-Hydroxyalkanoate)-Degrading Enzymes.
<3>Genome Announcements
<4>2
<5>e01272-13
<6>2014
<7>Here we report the draft genome sequence of Streptomyces exfoliatus DSMZ 41693, which includes
a gene encoding a poly(3-hydroxyoctanoate) depolymerase, an enzyme
which can be used for the industrial synthesis of chiral (R)-3-hydroxyalkanoic
acids. In addition, the genome carries numerous genes involved in the
biosynthesis of secondary metabolites, including polyketides and terpenes.

<>

<1>Martinez-Abarca, F., Garcia-Rodriguez, F.M., Toro, N.
<2>Homing of a bacterial group II intron with an intron-encoded protein lacking a recognizable endonuclease domain.
<3>Mol. Microbiol.
<4>35
<5>1405-1412
<6>2000
<7>RmInt1 is a functional group II intron found in Sinorhizobium meliloti where it interrupts a
group of IS elements of the IS630-Tc1 family. In contrast to many other group II introns, the
intron-encoded protein (IEP) of RmInt1 lacks the characteristic conserved part of the Zn
domain associated with the IEP endonuclease activity. Nevertheless, in this study, we show
that RmInt1 is capable of inserting into a vector containing the DNA spanning the RmInt1
target site from the genome of S. meliloti. Efficient homing was also observed in the absence
of homologous recombination (RecA- strains). In addition, it is shown that RmInt1 is able to
move to its target in a heterologous host (S. medicae). Homing of RmInt1 occurs very
efficiently upon DNA target uptake (conjugation/electroporation) by the host cell resulting in
a proportion of invaded target of 11-30%. Afterwards, the remaining intronless target DNA is
protected from intron invasion.

<>

<1>Martinez-Abarca, F., Martinez-Rodriguez, L., Lopez-Contreras, J.A., Jimenez-Zurdo, J.I., Toro, N.
<2>Complete Genome Sequence of the Alfalfa Symbiont Sinorhizobium/Ensifer meliloti Strain GR4.
<3>Genome Announcements
<4>1
<5>e00174-12
<6>2013
<7>We present the complete nucleotide sequence of the multipartite genome of
Sinorhizobium/Ensifer meliloti GR4, a predominant rhizobial strain in an
agricultural field site. The genome (total size, 7.14 Mb) consists of five
replicons: one chromosome, two expected symbiotic megaplasmids (pRmeGR4c and
pRmeGR4d), and two accessory plasmids (pRmeGR4a and pRmeGR4b).

<>

<1>Martinez-Penafiel, E., Fernandez-Ramirez, F., Ishida, C., Reyes-Cortes, R., Sepulveda-Robles, O., Guarneros-Pena, G., Bermudez-Cruz, R.M., Kameyama, L.
<2>Overexpression of Ipe protein from the coliphage mEp021 induces pleiotropic effects involving hemolysis by HlyE-containing vesicles and cell death.
<3>Biochimie
<4>94
<5>1262-1273
<6>2012
<7>Lysogenic Escherichia coli K-12 harbouring the prophage mEp021 displays haemolytic activity.
From a genomic library of mEp021, we identified an open reading frame (ORF 4) that was
responsible for the haemolytic activity. However, the ORF 4 sequence contains four initiation
codons in the same frame: ORF 4.1eORF 4.4, coding for 83-a.a., 82-a.a., 77-a.a. and 72-a.a.
products, respectively. The expression of the cloned ORF 4.3, or inducer of pleiotropic
effects (ipe), reproduced the haemolytic phenotype in a native strain carrying the gene hlyEþ,
but not in the mutant hlyE strain. The overexpression of Ipe induced several pleiotropic
effects, such as the inhibition of cell growth and the deregulation of cell division, which
resulted in a mixture of normal and desiccated-like cells: normal-filamentous,
desiccated-likefilamentous bacilli, minicells etc. Other effects included abnormalities of the
cell membrane, the production of vesicles containing HlyE, and finally, cell death. These
events were analysed at the molecular level by microarray assays. The global transcription
profile of E. coli K-12 strain MC4100, which expressed Ipe after 4 h, revealed differential
expression of various genes, most of which were related either to cell membrane and murein
biosynthesis or to cell division. The up-regulation of some of these transcripts was confirmed
by qRT-PCR. Additional research is needed to determine whether these effects are directly
related to Ipe activity or are consequences of the cellular responses to putative structural
damage induced by Ipe.

<>

<1>Maruyama, F., Kobata, M., Kurokawa, K., Nishida, K., Sakurai, A., Nakano, K., Nomura, R., Kawabata, S., Ooshima, T., Nakai, K., Hattori, M., Hamada, S., Nakagawa, I.
<2>Comparative genomic analyses of Streptococcus mutans provide insights into chromosomal shuffling and species-specific content.
<3>BMC Genomics
<4>10
<5>358
<6>2009
<7>Background: Streptococcus mutans is the major pathogen of dental caries, and it occasionally
causes infective endocarditis. While the
pathogenicity of this species is distinct from other human pathogenic
streptococci, the species-specific evolution of the genus Streptococcus
and its genomic diversity are poorly understood.Results: We have
sequenced the complete genome of S. mutans serotype c strain NN2025,
and compared it with the genome of UA159. The NN2025 genome is composed
of 2,013,587 bp, and the two strains show highly conserved core-genome.
However, comparison of the two S. mutans strains showed a large genomic
inversion across the replication axis producing an X-shaped symmetrical
DNA dot plot. This phenomenon was also observed between other
streptococcal species, indicating that streptococcal genetic
rearrangements across the replication axis play an important role in
Streptococcus genetic shuffling. We further confirmed the genomic
diversity among 95 clinical isolates using long-PCR analysis. Genomic
diversity in S. mutans appears to occur frequently between insertion
sequence (IS) elements and transposons, and these diversity regions
consist of restriction/modification systems, antimicrobial peptide
synthesis systems, and transporters. S. mutans may preferentially
reject the phage infection by clustered regularly interspaced short
palindromic repeats (CRISPRs). In particular, the CRISPR-2 region,
which is highly divergent between strains, in NN2025 has long repeated
spacer sequences corresponding to the streptococcal phage
genome.Conclusion: These observations suggest that S. mutans strains
evolve through chromosomal shuffling and that phage infection is not
needed for gene acquisition. In contrast, S. pyogenes tolerates phage
infection for acquisition of virulence determinants for niche
adaptation.

<>

<1>Marvig, R.L., Jochumsen, N., Johansen, H.K., Hoiby, N., Molin, S., Sommer, M.O., Jelsbak, L., Folkesson, A.
<2>Draft Genome Sequences of Pseudomonas aeruginosa B3 Strains Isolated from a Cystic Fibrosis Patient Undergoing Antibiotic Chemotherapy.
<3>Genome Announcements
<4>1
<5>e00804-13
<6>2013
<7>Pseudomonas aeruginosa frequently establishes chronic infections in the airways of patients
suffering from cystic fibrosis (CF). Here, we report the draft genome
sequences of four P. aeruginosa B3 strains isolated from a chronically infected
CF patient undergoing antibiotic chemotherapy.

<>

<1>Marx, C.J. et al.
<2>Complete genome sequences of six strains of the genus methylobacterium.
<3>J. Bacteriol.
<4>194
<5>4746-4748
<6>2012
<7>The complete and assembled genome sequences were determined for six strains of the
alphaproteobacterial genus Methylobacterium, chosen for their key adaptations
to different plant-associated niches and environmental constraints.

<>

<1>Marx, H., Graf, A.B., Tatto, N.E., Thallinger, G.G., Mattanovich, D., Sauer, M.
<2>Genome Sequence of the Ruminal Bacterium Megasphaera elsdenii.
<3>J. Bacteriol.
<4>193
<5>5578-5579
<6>2011
<7>Megasphaera elsdenii is a Gram-negative ruminal bacterium. It is being investigated as a
probiotic supplement for ruminants as it may provide
benefits for energy balance and animal productivity. Furthermore, it is of
biotechnological interest due to its capability of producing various
volatile fatty acids. Here we report the complete genome sequence of M.
elsdenii DSM 20460, the type strain for the species.

<>

<1>Marx, J.L.
<2>Restriction enzymes:  new tools for studying DNA.
<3>Science
<4>180
<5>482-485
<6>1973
<7>None

<>

<1>Marzabal, S., DuBois, S., Thielking, V., Cano, A., Eritja, R., Guschlbauer, W.
<2>Dam methylase from Escherichia coli: kinetic studies using modified DNA oligomers: hemimethylated substrates.
<3>Nucleic Acids Res.
<4>23
<5>3648-3655
<6>1995
<7>We have measured steady-state kinetics of the N6-adenine methyltransferase Dam Mtase using as
substrates non-selfcomplementary tetradecamer duplexes (d[GCCGGATCTAGACG].d[CGTCTAGATCCGGC])
containing the hemimethylated GATC target sequence in one or the other strand and
modifications in the GATC target sequence of the complementary strands.  Modifications
included substitution of guanine by hypoxanthine (I), thymine by uracil (U) or 5-ethyl-uracil
(E) and adenine by 2,6-diamino-purine (D).  Thermodynamic parameters were obtained from the
concentration dependence of the melting temperature (Tm) of the duplexes.  Large differences
in DNA methylation of duplexes containing single dl for dG substitution of the Dam recognition
site were observed compared with the canonical substrate, if the substitution involved the
top strand (on the G.C rich side).  Substitution in either strand by uracil (dU) or
5-ethyl-uracil (dE) resulted in small perturbation of the methylation patterns.  When
2,5-diamino-purine (dD) replaced the adenine to be methylated, small, but significant
methylation was observed.  The kinetic parameters of the methylation reaction were compared
with the thermodynamic free energies and significant correlation was observed.

<>

<1>Mas-Llado, M., Pina-Villalonga, J.M., Brunet-Galmes, I., Nogales, B., Bosch, R.
<2>Draft Genome Sequences of Two Isolates of the Roseobacter Group, Sulfitobacter sp. Strains 3SOLIMAR09 and 1FIGIMAR09, from Harbors of Mallorca Island  (Mediterranean Sea).
<3>Genome Announcements
<4>2
<5>e00350-14
<6>2014
<7>We present the draft genome sequences of two isolates of the Roseobacter lineage, 3SOLIMAR09
and 1FIGIMAR09, which were obtained from harbors of Mallorca Island,
Spain, and are affiliated with the Sulfitobacter genus. Both isolates harbor the
complete gene set for protocatechuate catabolism and incomplete pathways for
several additional monoaromatic compounds.

<>

<1>Masai, E., Kamimura, N., Kasai, D., Oguchi, A., Ankai, A., Fukui, S., Takahashi, M., Yashiro, I., Sasaki, H., Harada, T., Nakamura, S., Katano, Y., Narita-Yamada, S., Nakazawa, H., Hara, H., Katayama, Y., Fukuda, M., Yamazaki, S., Fujita, N.
<2>Complete Genome Sequence of Sphingobium sp. Strain SYK-6, a Degrader of Lignin-Derived Biaryls and Monoaryls.
<3>J. Bacteriol.
<4>194
<5>534-535
<6>2012
<7>Sphingobium sp. strain SYK-6 is able to grow on an extensive variety of lignin-derived biaryls
and monoaryls, and the catabolic genes for these
compounds are useful for the production of industrially valuable
metabolites from lignin. Here we report the complete nucleotide sequence
of the SYK-6 genome which consists of the 4,199,332-bp-long chromosome and
the 148,801-bp-long plasmid.

<>

<1>Mashhoon, N.
<2>Kinetic and chemical mechanisms of the EcoRI DNA (N6-adenine) methyltransferase.
<3>Diss. Abstr.
<4>54
<5>5132B-5133B
<6>1994
<7>A kinetic analysis of the EcoRI DNA N6-adenine methyltransferase (Mtase) is presented. The
enzyme catalyzes the S-adenosylmethionine (AdoMet)-dependent methylation of a short, synthetic
14 base pair DNA substrate and plasmid pBR322 DNA substrate with kcat/Km values of 0.51 x 10/8
and 4.1 x 10/8 s-1 M-1, respectively. The Mtase is thus one of the most efficient biocatalysts
known. The steady state kinetic data are consistent with an ordered bi-bi steady-state
mechanism in which AdoMet binds first, followed by (AdoHcy), is an uncompetitive inhibitor
with respect to DNA and a competitive inhibitor with respect to AdoMet. Thus, initial DNA
binding followed by AdoHcy binding leads to formation of a ternary deadend complex
(Mtase-AdoHcy binding leads to formation of a ternary dead-end complex (Mtase-DNA-AdoHcy). The
product inhibition patterns and apparent order of substrate binding can be reconciled by a
mechanism in which the Mtase binds AdoMet and noncanonical DNA randomly but that recognition
of the canonical site requires AdoMet to be bound. Presteady-state and isotope partition
analyses starting with the binary Mtase-AdoMet complex confirm its catalytic competence. In
summary, it was determined that the EcoRI DNA Mtase binds AdoMet and noncanonical DNA randomly
but recognition of the canonical site requires AdoMet to be bound. It was shown that step(s)
after methyl transfer limit the overall reaction rate and that methyl transfer at the N6 amino
moiety of adenine on each strand requires a single binding orientation. Furthermore, it was
demonstrated that no critical protein residues undergo ionization state changes in the pH
range 5.5-8.0. However, above pH 8.5 at least four residues in the free enzyme and AdoMet
bound enzyme and two residues in the central complex (Mtase-DNA-AdoMet) are implicated to be
critical in binding and/or catalysis; candidate amino acid residues are cysteine (other than
cysteine 223), lysine, tyrosine, and/or arginine.

<>

<1>Mashhoon, N., Carroll, M., Pruss, C., Eberhard, J., Ishikawa, S., Estabrook, R.A., Reich, N.
<2>Functional Characterization of Escherichia coli DNA Adenine Methyltransferase, a Novel Target for Antibiotics.
<3>J. Biol. Chem.
<4>279
<5>52075-52081
<6>2004
<7>We have characterized Escherichia coli DNA adenine methyltransferase, a critical regulator of
bacterial virulence. Steady-state kinetics, product
inhibition, and isotope exchange studies are consistent with a kinetic
mechanism in which the cofactor S-adenosylmethionine binds first, followed
by sequence-specific DNA binding and catalysis. The enzyme has a fast
methyl transfer step followed by slower product release steps, and we
directly demonstrate the competence of the enzyme cofactor complex.
Methylation of adjacent GATC sites is distributive with DNA derived from a
genetic element that controls the transcription of the adjacent genes.
This indicates that the first methylation event is followed by enzyme
release. The affinity of the enzyme for both DNA and S-adenosylmethionine
was determined. Our studies provide a basis for further structural and
functional analysis of this important enzyme and for the identification of
inhibitors for potential therapeutic applications.

<>

<1>Mashhoon, N., Pruss, C., Carroll, M., Johnson, P.H., Reich, N.O.
<2>Selective inhibitors of bacterial DNA adenine methyltransferases.
<3>J. Biomol. Screen.
<4>11
<5>497-510
<6>2006
<7>The authors describe the discovery and characterization of several structural classes of
small-molecule inhibitors of bacterial DNA
adenine methyltransferases. These enzymes are essential for bacterial
virulence (DNA adenine methyltransferase [DAM]) and cell viability
(cell cycle-regulated methyltransferase [CcrM]). Using a novel
high-throughput fluorescence-based assay and recombinant DAM and CcrM,
the authors screened a diverse chemical library. They identified 5
major structural classes of inhibitors composed of more than 350
compounds: cyclopentaquinolines, phenyl vinyl furans,
pyrimidine-diones, thiazolidine-4-ones, and phenyl-pyrroles. DNA
binding assays were used to identify compounds that interact directly
with DNA. Potent compounds selective for the bacterial target were
identified, whereas other compounds showed greater selectivity for the
mammalian DNA cytosine methyltransferase, Dnmt1. Enzyme inhibition
analysis identified mechanistically distinct compounds that interfered
with DNA or cofactor binding. Selected compounds demonstrated
cell-based efficacy. These small-molecule DNA methyltransferase
inhibitors provide useful reagents to probe the role of DNA methylation
and may form the basis of developing novel antibiotics.

<>

<1>Mashhoon, N., Reich, N.O.
<2>Analysis of the kinetic and chemical mechanisms of EcoRI methylase.
<3>FASEB J.
<4>4
<5>A1979
<6>1990
<7>Our goal is to elucidate the kinetic and chemical mechanisms of the EcoRI DNA
(adenine-N6)-methyltransferase.  Steady state and pre-steady state kinetic
experiments suggest an ordered bi bi mechanism in which S-Adenosylmethionine
(AdoMet) binds first, followed by DNA substrate.  After methyl transfer,
S-Adenosylhomocysteine (AdoHcy) dissociates followed by methylated DNA.  The
methyl transfer step is at least nine fold faster than the catalytic turnover
(kcat).  The kcat and Michaelis constants (Km) for AdoMet with a short 14 base
pair (14mer) and a long 4363 base pair plasmid DNA substrates are similar (150
nM).  In contrast, the Km for the plasmid DNA is fifty fold lower than 14mer.
The specificity constant (kcat/Km) for plasmid DNA is about 50 fold greater
than for the 14mer, suggesting that EcoRI methylase is a processive enzyme.
With a kcat/Km ratio of 2.9x10/8 M-1 sec-1 for plasmid DNA, EcoRI methylase is
among the most efficient enzymes reported.  pH studies implicating specific
amino acid residues in catalysis and (or) substrate binding will be presented.

<>

<1>Mashhoon, N., Reich, N.O.
<2>A kinetic study of DNA methylation by EcoRI methylase.
<3>ACS Abstracts
<4>196
<5>96
<6>1988
<7>The ability of a protein to recognize and subsequently modify a specific
sequence of bases in DNA is a fundamental biochemical question that is being
addressed in our laboratory by studying the EcoRI DNA methylase.  EcoRI
methylase, a 38 kilodalton bacterial protein, requires S-adenosyl methionine
(AdoMet) to methylate the double stranded DNA sequence GAATTC at the second
adenine.  Methylation of DNA serves to protect the organism's DNA against
cleavage by the corresponding endonuclease.  Little is known about the enzyme's
kinetic mechanism, the order of substrate binding and product release and the
rate limiting step in the reaction.  Steady state and pre-steady state analysis
were used to determine the kinetic mechanism.  Product inhibition with
S-adenosyl homocysteine is competitive with respect to AdoMet, whereas it is
non-competitive with respect to DNA.  Data will be presented on the steady
state kinetic parameters for AdoMet and DNA, the rate limiting step in the
reaction, and the order of substrate binding and product release.

<>

<1>Mashhoon, N., Reich, N.O.
<2>Investigation of ionizable residues critical for sequence-specific enzymatic DNA modification: Protein modification and steady-state and pre-steady-state kinetic pH analyses of EcoRI DNA methyltransferase.
<3>Biochemistry
<4>33
<5>7113-7119
<6>1994
<7>Steady- and pre-steady-state pH kinetic analyses are widely used methods to investigate
important ionizable groups in enzyme-catalyzed reactions. The first such analysis to identify
ionizable residues critical for sequence-specific modification of DNA is presented. EcoRI DNA
methyltransferase uses S-adenosyl-L-methionine (AdoMet) to catalyze the N6 methylation of the
second adenine in the double-stranded DNA sequence GAATTC. The kinetic mechanism was
previously shown to be steady-state-ordered bi bi in which AdoMet binds first followed by DNA
addition (Reich, N.O. & Mashhoon, N. (1991) Biochemistry 30, 2933-2939). Steady-state
parameters are strongly dependent on pH and implicate at least four residues with pKa values
between 8.2 and 8.9 in the free enzyme and AdoMet-bound enzyme and one residue with an
apparent pKa of 6.0. The data obtained aer consistent with the enzyme binding the form of
AdoMet in which the alpha amino group is protonated. Two protein residues with an apparent pKa
between 8.9 and 9.2 were implicated within the central complex (enzyme-DNA-AdoMet). The
general insensitivity of all steady-state parameters to pH changes between pH 6.0 and 8.0
suggests that no critical protein residues undergo ionization-state changes in this range. The
lack of significant pH-dependent changes in protein fluorescence and DNA thermal stability
suggests minimal structural changes in either macromolecule. In support of the steady-state
results single-turnover experiments reveal minimal pH dependence of the methylation rate
constant between pH 5.53 and 8.6. Thus, no amino acids critical for catalysis undergo
ionization-state changes in this range. The previously identified site of NEM-mediated
inactivation of the methyltransferase, cysteine 223 (Everett et al. (1990) J. Biol. Chem. 265,
17713-17719), has a pKa of 7.9; on the basis of the steady- and pre-steady-state pH
dependences this cysteine does not directly contribute to catalysis and substrate binding. The
results are discussed in the context of the requirement for significant enzymatic activation
of the poorly nucleophilic N6 position.

<>

<1>Mashhoon, N., Reich, N.O.
<2>A kinetic study of DNA methylation by EcoRI methylase.
<3>J. Cell Biol.
<4>107
<5>838a
<6>1988
<7>The ability of a protein to recognize and subsequently modify a specific
sequence of bases in DNA is a fundamental biochemical question that is being
addressed in our laboratory by studying the EcoRI DNA methylase.  EcoRI
methylase, a 38 kilodalton bacterial protein, requires S-adenosyl methionine
(AdoMet) to methylate the double stranded DNA sequence GAATTC at the second
adenine.  Methylation of DNA serves to protect the organism's DNA against
cleavage by the corresponding endonuclease.  Little is known about the enzyme's
kinetic mechanism, the order of substrate binding and product release, and the
rate limiting step in the reaction.  Steady state and pre-steady state analysis
were used to determine the kinetic mechanism.  Product inhibition with
S-adenosyl homocysteine is competitive with respect to AdoMet, whereas it is
non-competititve with respect to DNA.  Data will be presented on the steady
state kinetic parameters for AdoMet and DNA, the rate limiting step in the
reaction, and the order of substrate binding and product release.

<>

<1>Mashhoon, N., Reich, N.O.
<2>An investigation of the chemical mechanism of the EcoRI DNA methyltransferase.
<3>FASEB J.
<4>6
<5>A333
<6>1992
<7>The pH dependence of the EcoRI DNA methyltransferase activity was studied under
steady state and pre-steady state conditions, and with chemical modification
analysis.  Steady state pH analysis suggests that a residue with a pKa of
8.5-9.0 is necessary in its protonated form for enzyme activity.  KmDNA
increases over 250 fold as pH increases from 5.5 to 9.5 and kcat decreases over
20 fold.  Pre-steady state experiments were performed to measure the
methylation rate constant under single turnover conditions (>1 s-1 compared to
the kcat of .124 s-1) and to determine how pH affects this methylation rate
constant.  Chemical modification of the methylase with N-ethylmaleimide was
carried out to substantiate the steady state experimental findings.

<>

<1>Masignani, V.
<2>Staphylococcus aureus proteins and nucleic acids.
<3>European Patent Office
<4>EP 18290892 A
<5>
<6>2007
<7>The invention provides proteins from Staphylococcus aureus including amino acid sequences and
the corresponding nucleotide sequences.  The proteins are useful for vaccines, immunogenic
compositions, diagnostics, enzymatic studies and also as targets for antibiotics.

<>

<1>Masignani, V., Arico, M.
<2>Polypeptides from non-typeable haemophilus influenzae.
<3>International Patent Office
<4>WO 2005111066 A
<5>
<6>2005
<7>Polypeptides comprising non-typeable Haemophilus influenzae (NTHi) amino acid sequences.  Over
2500 specific NTHi proteins are disclosed.  The invention also provides related polypeptides,
nucleic acids, antibodies and methods.  These can all be used in medicine for treating or
preventing disease and/or infection caused by H. influenzae, such as otitis media.

<>

<1>Masignani, V., Arico, M.
<2>POLYPEPTIDES FROM NON-TYPEABLE HAEMOPHILUS INFLUENZAE.
<3>Japanese Patent Office
<4>JP 2008506364 A
<5>
<6>2008
<7>
<>

<1>Masignani, V., Arico, M.
<2>POLYPEPTIDES FROM NON-TYPEABLE HAEMOPHILUS INFLUENZAE.
<3>Japanese Patent Office
<4>JP 2012000112 A
<5>
<6>2012
<7>
<>

<1>Masignani, V., Mora, M., Scarselli, M.
<2>Staphylococcus aureus proteins and nucleic acids.
<3>International Patent Office
<4>WO 02094868 A
<5>
<6>2002
<7>The invention provides proteins from Staphylococcus aureus including amino acid sequences and
the corresponding nucleotide sequences.  The proteins are useful for vaccines, immunogenic
compositions, diagnostics, enzymatic studies and also as targets for antibiotics.

<>

<1>Masignani, V., Rappuoli, R., Tettelin, H.
<2>HAEMOPHILUS INFLUENZAE TYPE B.
<3>Japanese Patent Office
<4>JP 2008538183 A
<5>
<6>2008
<7>
<>

<1>Masignani, V., Tettelin, H., Fraser, C.
<2>Streptococcus pneumoniae proteins and nucleic acids.
<3>International Patent Office
<4>WO 02077021 A
<5>
<6>2002
<7>The invention provides proteins and nucleic acid sequences from Streptococcus pneumoniae,
together with a genome sequence.  These are useful for the development of vaccines,
diagnostics, and antibiotics.

<>

<1>Masignani, V., Tettelin, H., Fraser, C.
<2>Streptococcus pneumoniae proteins and nucleic acids.
<3>European Patent Office
<4>EP 1630230 A
<5>
<6>2006
<7>The invention provides proteins and nucleic acid sequences from Streptococcus pneumoniae,
together with a genome sequence.  These are useful for the development of vaccines,
diagnostics, and antibodies.

<>

<1>Masignani, V., Tettelin, H., Fraser, C.
<2>Streptococcus pneumoniae proteins and nucleic acids.
<3>European Patent Office
<4>EP 2278008 A
<5>
<6>2011
<7>The invention provides proteins and nucleic acid sequences from Streptococcus pneumoniae,
together with a genome sequence.  These are useful for the development of vaccines,
diagnostics, and antibiotics.

<>

<1>Maslov, A., Metrikin, M., Vijg, J.
<2>A dual-activation, adenoviral-based system for the controlled induction of DNa double-strand breaks by the restriction endonuclease SacI.
<3>Biotechniques
<4>47
<5>847-854
<6>2009
<7>Spontaneous damage to DNA is frequent and may lead to cell death, cell senescence, or
mutations.  DNA double-strand breaks are of special interest because they are highly toxic and
have been implicated in neurodegeneration, cancer, and aging.  Until now, there has not been a
reliable system allowing tunable induction of random DSBs without affecting other
macromolecules or cell functions.  Here, we describe an adenoviral-based,
doxycycline-mediated, and tamoxifen-dependent system for quantitative introduction of DSBs in
mammalian cells.  We generated a single adenoviral vector containing a tet-inducible,
composite SalI restriction endonuclease/estrogen receptor gene, and a constitutively expressed
reverse transactivator gene.  Transduced mouse embryonic fibroblasts - as well as mouse liver
cells in vivo - demonstrated a high level of DSBs in response to treatment with doxycycline
and tamoxifen.  We show that the amount of induced DSBs can be titrated by doxycycline dose
and duration of treatment.  This system should be useful for studying the processing of
randomly induced DSBs and their effects on cell fate, without the side effects normally
associated with radiation or chemical treatment.

<>

<1>Masood, N., Jackson, E., Moore, K., Farbos, A., Paszkiewicz, K., Dickins, B., McNally, A., Forsythe, S.
<2>Draft Genome Sequence of 'Candidatus Cronobacter colletis' NCTC 14934T, a New Species in the Genus Cronobacter.
<3>Genome Announcements
<4>2
<5>e00585-14
<6>2014
<7>Members of the Cronobacter genus are associated with serious infections in neonates. This is
the first report of the draft genome sequence for the newly
proposed species Cronobacter colletis.

<>

<1>Masood, N., Moore, K., Farbos, A., Hariri, S., Block, C., Paszkiewicz, K., Dickins, B., McNally, A., Forsythe, S.
<2>Draft Genome Sequence of a Meningitic Isolate of Cronobacter sakazakii Clonal Complex 4, Strain 8399.
<3>Genome Announcements
<4>1
<5>e00833-13
<6>2013
<7>The Cronobacter sakazakii clonal lineage defined as clonal complex 4 (CC4), composed of nine
sequence types, is associated with severe cases of neonatal
meningitis. To date, only closely related C. sakazakii sequence type 4 (ST4)
strains have been sequenced. C. sakazakii strain 8399, isolated from a case of
neonatal meningitis, was sequenced as the first non-ST4 C. sakazakii strain.

<>

<1>Masood, N., Moore, K., Farbos, A., Hariri, S., Paszkiewicz, K., Dickins, B., McNally, A., Forsythe, S.
<2>Draft Genome Sequences of Three Newly Identified Species in the Genus Cronobacter, C. helveticus LMG23732T, C. pulveris LMG24059, and C. zurichensis  LMG23730T.
<3>Genome Announcements
<4>1
<5>e00783-13
<6>2013
<7>Cronobacter helveticus, Cronobacter pulveris, and Cronobacter zurichensis are newly described
species in the Cronobacter genus, which is associated with
serious infections of neonates. This is the first report of draft genome
sequences for these species.

<>

<1>Masood, N., Moore, K., Farbos, A., Hariri, S., Paszkiewicz, K., Dickins, B., McNally, A., Forsythe, S.
<2>Draft Genome Sequence of the Earliest Cronobacter sakazakii Sequence Type 4 Strain, NCIMB 8272.
<3>Genome Announcements
<4>1
<5>e00782-13
<6>2013
<7>The Cronobacter sakazakii clonal lineage defined as sequence type 4 (ST4) is associated with
severe cases of neonatal meningitis and persistence in powdered
infant formula. For genome sequencing of the earliest deposited culture
collection strain of Cronobacter sakazakii ST4, we used the strain NCIMB 8272,
originally isolated from milk powder in 1950.

<>

<1>Masson, J.M., Lefevre, F., Saves, I., Laneelle, M.A., Daffe, M.
<2>Method for detecting a mycobacterium tuberculosis specific intein and use in diagnosis of tuberculosis.
<3>International Patent Office
<4>WO 0161035 A
<5>
<6>2001
<7>The invention concerns a method for detecting and/or quantifying Mycobacterium tuberculosis in
a sample, characterised in that it consists in detecting in said sample the presence of an
intein inserted at a site whereof the location is Mycobacterium tuberculosis specific using a
reagent specific to said location, and optionally in quantifying the detected signal.

<>

<1>Master, S.S.
<2>What is the origin and role of the large variable region in the M-AluI DNA-(cytosine c5) methyltransferase?
<3>Diss. Abstr.
<4>57
<5>1626
<6>1996
<7>AluI DNA-(cytosine C5)-methyltransferase is part of a restriction-modification system, and
methylates the 5-carbon of cytosine in the sequence 5'-AGCT-3'.  The amino acid sequences of
all known 5mC MTases contain ten conserved motifs, and between Motifs VIII and IX have a
variable region containing one or more Target Recognizing Domains.  TRDs are responsible for
DNA sequence specificity: monospecific MTases appear to have only one, while multispecific
MTases have as many as five.  M.AluI has the second largest variable region of all known 5mC
MTases, and sequence comparisons reveal four candidate TRDs in its variable region.  What is
the origin and role of this large variable region?  One possibility is that M.AluI evolved
from a multispecific MTase, or imported its variable region from a multispecific MTase.
Alternatively, it may have evolved from a monospecific MTase, expanding its variable region by
partial duplications.  Phylogenetic parsimony analysis and FASTA followed by Spearman's rank
order correlation coefficient suggested that neither M.AluI nor its variable region is closely
related to the known multispecific MTases.  The M.AluI pattern, however, is consistent with
importation of the variable region.  Methylation of AGCT sites accounted for 85-90% of the
methylating activity, but M.HhaI gave no unexplained methylating activity.  Various insertions
or deletion mutants failed to identify dispensable portions of the variable region, but a
sensitive in vivo assay based on McrBC restriction did reveal that the central portion is
particularly important for activity.  If M.AluI is like the multispecific MTases, which it
resembles in size, then it might be expected to accommodate a TRD from a multispecific MTase.
When such a TRD was moved into two locations in the M.AluI variable region, neither construct
gained a new specificity and both had greatly reduced activity.  Hence, M.AluI behaves in most
respects like a monospecific MTase, and the remarkable size of its variable region remains to
be explained.

<>

<1>Master, S.S., Blumenthal, R.M.
<2>Is the M.AluI DNA-(cytosine C5)-methyltransferase monospecific or multispecific?
<3>FASEB J.
<4>9
<5>A1399
<6>1995
<7>The AluI DNA-(cytosine C5)-methyltransferase (M.AluI) is part of a type II
restriction-modification system and methylates cytosine at the 5-carbon position in the
sequence 5'-AGCT-3'. The predicted amino acid sequences of all known 5-methylcytosine (5mC)
MTases contain ten conserved motifs, and a variable region which lies between motifs VIII and
IX. This variable region contains one or more Target Recognition Domains (TRDs) responsible
for the DNA sequence specificity of the MTase. Monospecific 5mC MTases are believed to have
only one TRD, while multispecific 5mC methylases have as many as four. M.AluI has the
second-largest variable region of all known 5mC MTases, and sequence comparisons with the TRDs
of multispecific MTases reveal four candidate TRDs in the M.AluI variable region. We have
confirmed that M.AluI is monospecific. Furthermore, we have constructed a series of insertion
and deletion mutants in order to identify a single contiguous region as the candidate TRD
responsible for M.AluI 5'-AGCT-3' specificity. In multispecific MTases a TRD is necessary
and sufficient to confer the cognate specificity, as shown by moving the TRD into another
multispecific MTase. In monospecific MTases, however, the TRD alone is not sufficient to
confer the required specificity but requires the presence of at least motif IX from the parent
MTase. Is M.AluI functionally more closely related to the multispecifics than to other
monospecifics. We are testing this in collaboration with another laboratory. If M.AluI is like
the multispecific MTases, than it should accommodate a TRD from a multispecific MTase. When a
TRD was moved from a multispecific MTase into two different locations in the variable region
of M.AluI, neither construct gained a new specificity and both had greatly reduced activity.
To test if the candidate M.AluI TRD is sufficient to confer specificity we are moving it into
the variable region of a multispecific MTase. From the existing data, it seems as if M.AluI
behaves in all respects as a monospecific MTase, and the remarkable size of its variable
region remains to be explained.

<>

<1>Master, S.S., Blumenthal, R.M.
<2>A genetic and functional analysis of the unusually large variable region in the M.AluI DNA-(cytosine C5)-methyltransferase.
<3>Mol. Gen. Genet.
<4>257
<5>14-22
<6>1997
<7>The M.AluI DNA-(cytosine C5)-methyltransferase (5mC methylase) acts on the sequence
5'-AGCT-3'.  The amino acid sequences of known 5mC methylases contain ten conserved motifs,
with a variable region between Motifs VIII and IX that contains one or more
"target-recognizing domains" responsible for DNA sequence specificity.  Monospecific 5mC
methylases are believed to have only one TRD, while multi-specific 5mC methylases have as many
as five.  M.AluI has the second-largest variable region of all known 5mC methylase, and
sequence analysis reveals five candidate TRDs.  In testing whether M.AluI is in fact
monospecific it was found that AGCT methylation represents only 80-90% of the methylating
activity of this enzyme, while control experiments with the enzyme M.HhaI gave no unexplained
activity.  Because individual TRDs can be deleted from multispecific methylases without
general loss of activity, a series of insertion and deletion mutants of the M.AluI variable
region were prepared.  All deletions that removed more than single amino acids from the
variable region caused significant loss of activity; a sensitive in vivo assay for methylase
activity based on McrBC restriction suggested that the central portion of the variable region
is particularly important.  In some cases, multispecific methylases can accommodate a TRD from
another multispecific methylase, thereby acquiring an additional specificity.  When TRDs were
removed from a multispecific methylase into two different locations in the variable region of
M.AluI, all hybrid enzymes had greatly reduced activity and no new specificities.  M.AluI thus
behaves in most respects as a monospecific methylase despite the remarkable sizeof its
variable region.

<>

<1>Masuda, H., Shiwa, Y., Yoshikawa, H., Zylstra, G.J.
<2>Draft Genome Sequence of the Versatile Alkane-Degrading Bacterium Aquabacterium sp. Strain NJ1.
<3>Genome Announcements
<4>2
<5>e01271-14
<6>2014
<7>The draft genome sequence of a soil bacterium, Aquabacterium sp. strain NJ1, capable of
utilizing both liquid and solid alkanes, was deciphered. This is the
first report of an Aquabacterium genome sequence.

<>

<1>Masuda, S., Hori, K., Maruyama, F., Ren, S., Sugimoto, S., Yamamoto, N., Mori, H., Yamada, T., Sato, S., Tabata, S., Ohta, H., Kurokawa, K.
<2>Whole-Genome Sequence of the Purple Photosynthetic Bacterium Rhodovulum sulfidophilum Strain W4.
<3>Genome Announcements
<4>1
<5>e00577-13
<6>2013
<7>We report the draft genome sequence of the purple photosynthetic bacterium Rhodovulum
sulfidophilum. The photosynthesis gene cluster comprises two
segments-a unique feature among photosynthesis gene clusters of purple bacteria.
The genome information will be useful for further analysis of bacterial
photosynthesis.

<>

<1>Matejkova, P., Strouhal, M., Smajs, D., Norris, S.J., Palzkill, T., Petrosino, J.F., Sodergren, E., Norton JE, S.J., Richmond, T.A., Molla, M.N., Albert, T.J., Weinstock, G.M.
<2>Complete genome sequence of Treponema pallidum ssp. pallidum strain SS14 determined with oligonucleotide arrays.
<3>BMC Microbiol.
<4>8
<5>76
<6>2008
<7>BACKGROUND: Syphilis spirochete Treponema pallidum ssp. pallidum remains the enigmatic
pathogen, since no virulence factors have been identified
and the pathogenesis of the disease is poorly understood. Increasing rates
of new syphilis cases per year have been observed recently. RESULTS: The
genome of the SS14 strain was sequenced to high accuracy by an
oligonucleotide array strategy requiring hybridization to only three
arrays (Comparative Genome Sequencing, CGS). Gaps in the resulting
sequence were filled with targeted dideoxy-terminators (DDT) sequencing
and the sequence was confirmed by whole genome fingerprinting (WGF). When
compared to the Nichols strain, 327 single nucleotide substitutions (224
transitions, 103 transversions), 14 deletions, and 18 insertions were
found. On the proteome level, the highest frequency of amino acid-altering
substitution polymorphisms was in novel genes, while the lowest was in
housekeeping genes, as expected by their evolutionary conservation.
Evidence was also found for hypervariable regions and multiple regions
showing intrastrain heterogeneity in the T. pallidum chromosome.
CONCLUSION: The observed genetic changes do not have influence on the
ability of Treponema pallidum to cause syphilitic infection, since both
SS14 and Nichols are virulent in rabbit. However, this is the first
assessment of the degree of variation between the two syphilis pathogens
and paves the way for phylogenetic studies of this fascinating organism.

<>

<1>Mather, A.E. et al.
<2>Distinguishable epidemics of multidrug-resistant Salmonella Typhimurium DT104 in different hosts.
<3>Science
<4>341
<5>1514-1517
<6>2013
<7>The global epidemic of multidrug-resistant Salmonella Typhimurium DT104 provides
an important example, both in terms of the agent and its resistance, of a widely
disseminated zoonotic pathogen. Here, with an unprecedented national collection
of isolates collected contemporaneously from humans and animals and including a
sample of internationally derived isolates, we have used whole-genome sequencing
to dissect the phylogenetic associations of the bacterium and its antimicrobial
resistance genes through the course of an epidemic. Contrary to current tenets
supporting a single homogeneous epidemic, we demonstrate that the bacterium and
its resistance genes were largely maintained within animal and human populations
separately and that there was limited transmission, in either direction. We also
show considerable variation in the resistance profiles, in contrast to the
largely stable bacterial core genome, which emphasizes the critical importance of
integrated genotypic data sets in understanding the ecology of bacterial zoonoses
and antimicrobial resistance.

<>

<1>Mathew, M.J., Subramanian, G., Nguyen, T.T., Robert, C., Mediannikov, O., Fournier, P.E., Raoult, D.
<2>Genome Sequence of Diplorickettsia massiliensis, an Emerging Ixodes ricinus-Associated Human Pathogen.
<3>J. Bacteriol.
<4>194
<5>3287
<6>2012
<7>Diplorickettsia massiliensis is a gammaproteobacterium in the order Legionellales and an agent
of tick-borne infection. We sequenced the genome from strain 20B,
isolated from an Ixodes ricinus tick. The genome consists of a 1,727,973-bp
chromosome but no plasmid and includes 2,269 protein-coding genes and 42 RNA
genes, including 3 rRNA genes.

<>

<1>Mathimaran, N., Srivastava, R., Wiemken, A., Sharma, A.K., Boller, T.
<2>Genome sequences of two plant growth-promoting fluorescent pseudomonas strains, r62 and r81.
<3>J. Bacteriol.
<4>194
<5>3272-3273
<6>2012
<7>Plant growth-promoting rhizobacterial (PGPR) strains R62 and R81 have previously  been
isolated and characterized as part of the Indo-Swiss Collaboration in
Biotechnology. Here we present the draft genome sequences of these two PGPR
strains, with the aim of unraveling the mechanisms behind their ability to
promote wheat growth.

<>

<1>Matic, I., Taddei, F., Radman, M.
<2>Genetic barriers among bacteria.
<3>Trends Microbiol.
<4>4
<5>69-73
<6>1996
<7>Barriers to chromosomal gene transfer between bacterial species control their genetic
isolation.  These barriers, such as different microhabitats, the host ranges of genetic
exchange vectors and restriction-modification systems, limit gene exchange, but the major
limitation is genomic sequence divergence.  The mismatch-repair system inhibits interspecies
recombination, the inducible SOS system stimulates interspecies recombination, while natural
selection determines the effective recombination frequencies.

<>

<1>Matilla, M.A., Pizarro-Tobias, P., Roca, A., Fernandez, M., Duque, E., Molina, L., Wu, X., van der Lelie, D., Gomez, M.J., Segura, A., Ramos, J.L.
<2>Complete genome of the plant-growth promoting rhizobacterium Pseudomonas putida BIRD-1.
<3>J. Bacteriol.
<4>193
<5>1290
<6>2010
<7>We report the complete sequence of the 5.7-Mbp of Pseudomonas putida BIRD-1, a
metabolically-versatile plant growth-promoting rhizobacterium that is highly tolerant to
desiccation, capable of solubilizing inorganic phosphate and iron, and of synthesizing
phytohormones that stimulate seed germination and plant growth.

<>

<1>Matin, M.M., Hornby, D.P.
<2>Exploring the structural flexibility of 5m-cytosine-DNA methyltransferases.
<3>Biochem. Soc. Trans.
<4>26
<5>S394
<6>1998
<7>DNA methyltransferases are found in organisms ranging from bacteria to mammals.  They
recognize specific DNA sequences and transfer a methyl group to adenine or cytosine residues.
5m-cytosine methyltransferases form the basis of the work considered here, they belong to type
II restriction-modification systems, which use S-adenosyl-L-methionine as a cofactor to add
methyl groups to the 5 position of cytosine.  Two families of 5mC Mtases are known;
mono-specific methyltransferases which recognize and methylate a single DNA sequence, and
multi-specific Mtases that recognize and modify more than one DNA sequence.

<>

<1>Matin, M.M., Hornby, D.P.
<2>A positive selection vector combining tetracycline resistance that eliminates the need for bacterial plating comprising a modified version of the gene encoding the cytosine-specific DNA-methyltransferase and a modified form of the plasmid pBR322 tetA(C).
<3>Anal. Biochem.
<4>278
<5>46-51
<6>2000
<7>The construction of plasmid pMTet1 that combined positive selection with
tetracycline-resistance was described. The vector comprised a
modified version of the gene encoding the cytosine(C-5)-specific
DNA-methyltransferase (C5-Mtase) MspI and a modified form of the
plasmid pBR322 tetA(C) gene. This combination of a C5-Mtase gene and
the tetA(C) derived from plasmid pBR322 permitted the isolation of
recombinant plasmids in liquid culture which for the first time
eliminated the need to isolate single, antibiotic-resistant colonies
and therefore significantly accelerated recombinant plasmid isolation.
Furthermore, the application of this novel cloning vector, in
conjunction with chromatographic DNA fractionation, for the
construction of size-selected recombinant molecules was reported. The
construction of the plasmid pMTet1 was followed by the DNA
amplification reaction and nucleic acid chromatography. The plasmid
pMTet1 facilitated the rapid cloning of DNA molecules generated by
proof-reading DNA-polymerases and restriction digests using enzymes
that produced noncohesive termini.

<>

<1>Matje, D.M., Coughlin, D.F., Connolly, B.A., Dahlquist, F.W., Reich, N.O.
<2>Determinants of Precatalytic Conformational Transitions in the DNA Cytosine Methyltransferase M.HhaI.
<3>Biochemistry
<4>50
<5>1465-1473
<6>2011
<7>The DNA methyltransferase M.HhaI is an excellent model for understanding how recognition of a
nucleic acid substrate is translated
into site-specific modification. In this study, we utilize direct,
real-time monitoring of the catalytic loop position via engineered
tryptophan fluorescence reporters to dissect the conformational
transitions that occur in both enzyme and DNA substrate prior to
methylation of the target cytosine. Using nucleobase analogues in place
of the target and orphan bases, the kinetics of the base flipping and
catalytic loop closure rates were determined, revealing that base
flipping precedes loop closure as the rate-determining step prior to
methyl transfer. To determine the mechanism by which individual
specific hydrogen bond contacts at the enzyme DNA interface mediate
these conformational transitions, nucleobase analogues lacking hydrogen
bonding groups were incorporated into the recognition sequence to
disrupt the major groove recognition elements. The consequences of
binding, loop closure, and catalysis were determined for four contacts,
revealing large differences in the contribution of individual hydrogen
bonds to DNA recognition and conformational transitions on the path to
catalysis. Our results describe how M.HhaI utilizes direct readout
contacts to accelerate extrication of the target base that offer new
insights into the evolutionary history of this important class of
enzymes.

<>

<1>Matje, D.M., Zhou, H., Smith, D.A., Neely, R.K., Dryden, D.T., Jones, A.C., Dahlquist, F.W., Reich, N.O.
<2>Enzyme-Promoted Base Flipping Controls DNA Methylation Fidelity.
<3>Biochemistry
<4>52
<5>1677-1685
<6>2013
<7>A quantitative understanding of how conformational transitions contribute to enzyme catalysis
and specificity remains a fundamental challenge. A suite of
biophysical approaches was used to reveal several transient states of the
enzyme-substrate complexes of the model DNA cytosine methyltransferase M.HhaI.
Multidimensional, transverse relaxation-optimized nuclear magnetic resonance
(NMR) experiments show that M.HhaI has the same conformation with noncognate and
cognate DNA sequences. The high-affinity cognatelike mode requires the formation
of a subset of protein-DNA interactions that drive the flipping of the target
base from the helix to the active site. Noncognate substrates lacking these
interactions undergo slow base flipping, and fluorescence tracking of the
catalytic loop corroborates the NMR evidence of a loose, nonspecific binding mode
prior to base flipping and subsequent closure of the catalytic loop. This slow
flipping transition defines the rate-limiting step for the methylation of
noncognate sequences. Additionally, we present spectroscopic evidence of an
intermediate along the base flipping pathway that has been predicted but never
previously observed. These findings provide important details of how
conformational rearrangements are used to balance specificity with catalytic
efficiency.

<>

<1>Matney, T.S., MacDougall, N.L.T., Butler, M.A., Suit, J.C.
<2>Host-induced modification/restriction and the utilization of 5-bromouracil by thymineless mutants of Escherichia coli.
<3>J. Bacteriol.
<4>104
<5>606-607
<6>1970
<7>An mk-rk- mutation exerted no measurable effect on 5-bromouracil incorporation
by a thymineless derivative of Escherichia coli K.

<>

<1>Matselyukh, A.B.
<2>Genetic transformation of Streptomyces globisporus 1912 strains: restriction barrier and plasmid compatibility.
<3>Mikrobiol. Zh.
<4>63
<5>15-21
<6>2001
<7>Low efficiency of genetic transformation of protoplasts of different strains of Streptomyces
globisporus 1912 by means of DNA preparations
of three vectors pIJ487, pGM160 and pWHM4 is explained by the presence of
a restriction barrier in the recipients. This obstacle can be
overcome by the use of modified DNA of the same vectors, isolated from
not numerous transformants. The frequency of transformation by such
modified vector DNA was increased by two-three orders in comparison
with initial DNA, isolated from Streptomyces lividans TK24, losing
restriction-modification system. The vector pCNB4001, containing the
replicon of endogeneous plasmid pSG1912-1, effectively transformed the
protoplasts of all S. globisporus 1912 strains. Compatibility of pIJ487
and pSG1912-1 plasmids and incompatibility of the latter and pWHM4 was
shown.

<>

<1>Matson, E.G., Zhang, X., Leadbetter, J.R.
<2>Selenium controls transcription of paralogous formate dehydrogenase genes in the termite gut acetogen, Treponema primitia.
<3>Environ. Microbiol.
<4>12
<5>2245-2258
<6>2010
<7>Summary The termite gut spirochete, Treponema primitia, is a CO(2)-reductive acetogen that is
phylogenetically distinct from other distantly related and more extensively studied acetogens
such as Moorella thermoacetica. Research on T. primitia has revealed details about the role of
spirochetes in CO(2)-reductive acetogenesis, a process important to the mutualism occurring
between termites and their gut microbial communities. Here, a locus of the T. primitia genome
containing Wood-Ljungdahl pathway genes for CO(2)-reductive acetogenesis was sequenced. This
locus contained methyl-branch genes of the pathway (i.e. for the reduction of CO(2) to the
level of methyl-tetrahydrofolate) including paralogous genes for cysteine and selenocysteine
(Sec) variants of formate dehydrogenase (FDH) and genes for Sec incorporation. The FDH
variants affiliated phylogenetically with hydrogenase-linked FDH enzymes, suggesting that T.
primitia FDH enzymes utilize electrons derived directly from molecular H(2). Sub-nanomolar
concentrations of selenium decreased transcript levels of the cysteine variant FDH gene.
Selenium concentration did not markedly influence the level of mRNA upstream of the Sec-codon
in the Sec variant FDH; however, the level of transcript extending downstream of the Sec-codon
increased incrementally with increasing selenium concentrations. The features and regulation
of these FDH genes are an indication that T. primitia may experience dynamic selenium
availability in its H(2)-rich gut environment.

<>

<1>Matsuhisa, A., Eda, S., Uehara, H., Nishida, K., Keshi, H.
<2>Probe HP-34 for detecting Helicobacter pylori.
<3>Japanese Patent Office
<4>JP 2005198657 A
<5>
<6>2005
<7>
<>

<1>Matsui, M., Mise, K., Yoshida, Y., Ishidate, M.
<2>Production of restriction endonucleases from various Salmonella strains of human origin.
<3>Eisei Shikenjo Hokoku
<4>104
<5>92-96
<6>1986
<7>Using a safe procedure for the detection of restriction endonuclease-producing
strains, 21 restriction-positive strains were found among 120 Salmonella
strains of human origin.  The designation of the restriction endonucleases and
their producers was SinI and SinII in Salmonella infantis (11 strains), SblI in
Salmonella blockley (3 strains), StmI in Salmonella typhimurium, SbaI in
Salmonella bareilly, SscI in Salmonella schwarzengrund, SthI in Salmonella
thompson, SanI in Salmonella anatum, SisI in Salmonella Isangi and SbrI in
Salmonella bredeney.  Activity of all the endonucleases was very high.  No Hsd
plasmids has been isolated from these restriction endonuclease-producing
strains in spite of several trials, indicating that the hsd+ gene might be
carried on chromosomal DNA.

<>

<1>Matsumoto, A., Igo, M.M.
<2>Species-Specific Type II Restriction-Modification System of Xylella fastidiosa Temecula1.
<3>Appl. Environ. Microbiol.
<4>76
<5>4092-4095
<6>2010
<7>The transformation efficiency of Xylella fastidiosa can be increased by interfering with
restriction by the strain-specific type II system
encoded by the PD1607 and PD1608 genes. Here, we report results for two
strategies: in vitro methylation using M. SssI and isolation of DNA
from an Escherichia coli strain expressing the methylase PD1607.

<>

<1>Matsumura, H., Takahashi, H., Inoue, T., Yamamoto, T., Hashimoto, H., Nishioka, M., Fujiwara, S., Takagi, M., Imanaka, T., Kai, Y.
<2>Crystal structure of intein homing endonuclease II encoded in DNA polymerase gene from hyperthermophilic archaeon Thermococcus kodakaraensis strain KOD1.
<3>Proteins
<4>63
<5>711-715
<6>2006
<7>
<>

<1>Matsunaga, T., Okamura, Y., Fukuda, Y., Wahyudi, A.T., Murase, Y., Takeyama, H.
<2>Complete Genome Sequence of the Facultative Anaerobic Magnetotactic Bacterium Magnetospirillum sp. strain AMB-1.
<3>DNA Res.
<4>12
<5>157-166
<6>2005
<7>Magnetospirillum sp. strain AMB-1 is a Gram-negative alpha-proteobacterium that synthesizes
nano-sized magnetites, referred to as magnetosomes,
aligned intracellularly in a chain. The potential of this nano-sized
material is growing and will be applicable to broad research areas. It has
been expected that genome analysis would elucidate the mechanism of
magnetosome formation by magnetic bacteria. Here we describe the genome of
Magnetospirillum sp. AMB-1 wild type, which consists of a single circular
chromosome of 4967148 bp. For identification of genes required for
magnetosome formation, transposon mutagenesis and determination of
magnetosome membrane proteins were performed. Analysis of a non-magnetic
transposon mutant library focused on three unknown genes from 2752 unknown
genes and three genes from 205 signal transduction genes. Partial proteome
analysis of the magnetosome membrane revealed that the membrane contains
numerous oxidation/reduction proteins and a signal response regulator that
may function in magnetotaxis. Thus, oxidation/reduction proteins and
elaborate multidomain signaling proteins were analyzed. This comprehensive
genome analysis will enable resolution of the mechanisms of magnetosome
formation and provide a template to determine how magnetic bacteria
maintain a species-specific, nano-sized, magnetic single domain and
paramagnetic morphology.

<>

<1>Matsuo, K., Silke, J., Gramatikoff, K., Schaffner, W.
<2>The CpG-specific methylase SssI has topoisomerase activity in the presence of Mg2+.
<3>Nucleic Acids Res.
<4>22
<5>5354-5359
<6>1994
<7>A prokaryotic CpG-specific methylase from Spiroplasma, SssI methylase, is now widely used to
study the effect of CpG methylation in mammalian cells, and can processively modify cytosines
in CpG dinucleotides in the absence of Mg2+. In the presence of Mg2+, we found (i) that the
methylation reaction is distributive rather than processive as a result of the decreased
affinity of SssI methylase for DNA, and (ii) that a type I-like topoisomerase activity is
present in SssI methylase preparations. This topoisomerase activity was still present in SssI
methylase further purified by either SDS-polyacrylamide or isoelectric focusing gel
electrophoresis. We show that methylase and topoisomerase activities are not functionally
interdependent, since conditions exist where only one or the other enzymatic activity is
detectable. The catalytic domains of SssI methylase and prokaryotic topoisomerases show
similarity at the amino acid level, further supporting the idea that the topoisomerase
activity is a genuine activity of SssI methylase. Mycoplasmas, including Spiroplasma, have the
smallest genomes of all living organisms; thus, this condensation of two enzymatic activities
into the same protein may be a result of genome economy, and may also have functional
implications for the mechanism of methylation.

<>

<1>Matsuoka, S., Asai, K., Sadaie, Y.
<2>Restriction and modification of SP10 phage by BsuM of Bacillus subtilis Marburg.
<3>FEMS Microbiol. Lett.
<4>244
<5>335-339
<6>2005
<7>Bacillus subtilis Marburg has only one intrinsic restriction and modification system BsuM that
recognizes the CTCGAG (XhoI site)
sequence. It consists of two operons, BsuMM operon for two cytosine DNA
methyltransferases, and BsuMR operon for a restriction nuclease and two
associated proteins of unknown function. In this communication, we
analyzed the BsuM system by utilizing phage SPIO that possesses more
than twenty BsuM target sequences on the phage genome. SPIO phages
grown in the restriction and modification-deficient strain could not
make plaques on the restriction-proficient BsuMR(+) indicator strain.
An enforced expression of the wild type BsuMM operon in the BsuMR(+)
indicator strain, however, allowed more than thousand times more
plaques. DNA extracted from SPIO phages, thus, propagated became more
but not completely refractory to XhoI digestion in vitro. Thus, the
SPIO phage genome DNA is able to be nearly full-methylated but some
BsuM sites are considered to be unmethylated.

<>

<1>Matsushima, P., Baltz, R.H.
<2>Streptomyces lipmanii expresses two restriction systems that inhibit plasmid transformation and bacteriophage plaque formation.
<3>J. Bacteriol.
<4>171
<5>3128-3132
<6>1989
<7>Bacteriophage host range studies suggested that several beta-lactam-producing streptomycetes
express similar restriction-modification systems.  Streptomyces lipmanii LE32 expressed two
restriction-modification systems, designated SliI and SliII.  A mutant strain, PM87, was
defective only in SliI restriction but expressed both SliI and SliII modification.
Streptomyces sp. strain A57986, a natural isolate partially deficient in the expression of
SliI and SliII restriction, nevertheless modified bacteriophage DNA for both SliI and SliII
specificities.  Protoplasts of PM87 and A57986 were transformed by several plasmids, and the
modified plasmids isolated from these strains transformed wild-type S. lipmanii efficiently.

<>

<1>Matsushima, P., Baltz, R.H.
<2>Restriction and modification in Streptomyces lipmanii.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>89
<5>361
<6>1989
<7>Steptomyces lipmanii and several other beta-lactam antibiotic-producing
streptomycetes express restriction systems that inhibit plasmid transformation
and bacteriophage plaque formation.  Bacteriophage host range studies suggested
that many beta-lactam producers express some common restriction system(s).  We
isolated a mutant, S. lipmanii PM87, defective in one restriction system (SliI)
and analyzed restriction and modification of several different bacteriophages
in PM87 and its parent strain, LE32.  PM87 appears to be proficient in SliI
modification and expresses a second restriction/modification system, designated
SliII.  Another beta-lactam producing streptomycete, strain A57986, which was
naturally less restricting than S. lipmanii, expressed only SliI restriction
and modification.  PM87 and A57986 were readily transformed by many unmodified
plasmids; once modified in these hosts the same plasmids were efficiently
introduced into more restricting strains by transformation.

<>

<1>Matsushima, P., Cox, K.L., Baltz, R.H.
<2>Highly transformable mutants of Streptomyces fradiae defective in several restriction systems.
<3>Mol. Gen. Genet.
<4>206
<5>393-400
<6>1987
<7>Streptomyces fradiae JS85 is a mutant defective in tylosin production and an
efficient recipient for conjugal transfer of tylosin genes.  JS85 was
mutagenized with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and derivatives
defective in restriction were isolated by sequential selection for increased
transformability by several plasmid DNAs.  From the number of mutation and
selection cycles required to eliminate most restriction, it was estimated that
wild type S. fradiae expressed at least five restriction systems.  From the
patterns of restriction enzyme digestion of chromosomal DNA observed in the
series of mutants that became progressively less restricting, it was suggested
that wild type S. fradiae normally expresses modification (and presumably
restriction) systems similar or analogous to PstI, XhoI, ScaI and EcoRI.  The
least restricting mutant of S. fradiae was readily transformable by many
plasmids, including a bifunctional cosmid vector containing a large insert of
Streptomyces DNA.

<>

<1>Matsushima, P., McHenney, M.A., Baltz, R.H.
<2>Efficient transformation of Amycolatopsis orientalis (Nocardia orientalis) protoplasts by Streptomyces plasmids.
<3>J. Bacteriol.
<4>169
<5>2298-2300
<6>1987
<7>Conditions for efficient transformation of Amycolatopsis orientalis (Nocardia
orientalis) protoplasts by Streptomyces plasmid cloning vectors were
identified.  Three streptomycete plasmid origins of replication function in A.
orientalis, as do the apramycin resistance gene from Escherichia coli, the
thiostrepton resistance gene from Streptomyces azureus, and the tyrosinase gene
from Streptomyces antibioticus.  A. orientalis appears to express some
restriction and modification, because highest transformation frequencies
(1000000/microgram DNA) were obtained when plasmid pIJ702 was modified in A.

<>

<1>Matsushima, P., McHenney, M.A., Baltz, R.H.
<2>Transduction and transformation of plasmid DNA in Streptomyces fradiae strains that express different levels of restriction.
<3>J. Bacteriol.
<4>171
<5>3080-3084
<6>1989
<7>We constructed nonrestricting strains of Streptomyces fradiae blocked in
different steps in tylosin biosynthesis.  Plasmid transformation frequencies
were 10/3 to 10/4-fold higher and bacteriophage plating efficiences were 10/4
to 10/8-fold higher in the nonrestricting strains than in the restricting
strains.  The efficiences of transduction of plasmid pRHB101 in S. fradiae
strains varied by over 1,000-fold, depending on growth conditions, and optimum
transduction frequencies were obtained when cells were grown to mid-exponential
phase at 39C.  Under these conditions, restricting and nonrestricting strains
were transduced at frequencies that differed by only two- to fivefold.

<>

<1>Matsutani, M., Hirakawa, H., Nishikura, M., Soemphol, W., Ali, I.A., Yakushi, T., Matsushita, K.
<2>Increased number of Arginine-based salt bridges contributes to the thermotolerance of thermotolerant acetic acid bacteria, Acetobacter tropicalis SKU1100.
<3>Biochem. Biophys. Res. Commun.
<4>409
<5>120-124
<6>2011
<7>Thermotolerant acetic acid bacteria (AAB), Acetobacter tropicalis SKU1100,
can grow above 40 degrees C. To investigate the basis of its
thermotolerance, we compared the genome of A. tropicalis SKU1100 with that
of mesophilic AAB strain Acetobacter pasteurianus IFO3283-01. The
comparative genomic study showed that amino acid substitutions from large
to small residue and Lys to Arg occur in many orthologous genes.
Furthermore, comparative modeling study was carried out with the
orthologous proteins between SKU1100 and IFO3283-01 strains, indicating
that the number of Arg-based salt bridges increased in protein models.
Since it has been reported that Arg-based salt bridges are important
factor for thermo-stability of protein structure, our results strongly
suggest that the increased number of Arg-based salt bridges may
contributes to the thermotolerance of A. tropicalis SKU1100 (the
thermo-stability of proteins in A. tropicalis SKU1100).

<>

<1>Matsutani, M., Kawajiri, E., Yakushi, T., Adachi, O., Matsushita, K.
<2>Draft Genome Sequence of Dihydroxyacetone-Producing Gluconobacter thailandicus Strain NBRC 3255.
<3>Genome Announcements
<4>1
<5>e00118-13
<6>2013
<7>Here, we report the draft genome sequence of the acetic acid bacterium Glucnobacter
thailandicus strain NBRC 3255. The draft genome sequence is composed
of 109 contigs in 3,305,227 bp and contains 3,225 protein-coding genes. Two
paralogous sets of sldAB operons, which are responsible for dihydroxyacetone
production from glycerol, were identified.

<>

<1>Matsutani, M., Shirakihara, Y., Imada, K., Yakushi, T., Matsushita, K.
<2>Draft Genome Sequence of a Thermophilic Member of the Bacillaceae, Anoxybacillus  flavithermus Strain Kn10, Isolated from the Kan-nawa Hot Spring in Japan.
<3>Genome Announcements
<4>1
<5>e00311-13
<6>2013
<7>Here, we report the draft genome sequence of the Anoxybacillus flavithermus Kn10  strain (NBRC
109594), isolated from a water drain of the Kan-nawa Hot Spring in
Japan. The draft genome sequence is composed of 90 contigs for 2,772,624 bp with
41.6% G+C content and contains 2,883 protein-coding genes and 80 tRNA genes.

<>

<1>Matsutani, M., Suzuki, H., Yakushi, T., Matsushita, K.
<2>Draft genome sequence of Gluconobacter thailandicus NBRC 3257.
<3>Standards in Genomic Sciences
<4>9
<5>614-623
<6>2014
<7>Gluconobacter thailandicus strain NBRC 3257, isolated from downy cherry (Prunus tomentosa), is
a strict aerobic rod-shaped Gram-negative bacterium. Here, we
report the features of this organism, together with the draft genome sequence and
annotation. The draft genome sequence is composed of 107 contigs for 3,446,046 bp
with 56.17% G+C content and contains 3,360 protein-coding genes and 54 RNA genes.

<>

<1>Matsuura, M., Saldanha, R., Ma, H., Wank, H., Yang, J., Mohr, G., Cavangh, S., Dunny, G.M., Belfort, M., Lambowitz, A.M.
<2>A bacterial group II intron encoding reverse transcriptase, maturase, and DNA endonuclease activities: biochemical demonstration of maturase activity and insertion of new genetic information within the intron.
<3>Genes Dev.
<4>11
<5>2910-2924
<6>1997
<7>The Lactococcus lactis group II intron Ll.ltrB is similar to mobile yeast mtDNA group II
introns, which encode reverse transcriptase, RNA maturase, and DNA endonuclease activities for
site-specific DNA insertion. Here, we show that the Lactococcal intron can be expressed and
spliced efficiently in Escherichia coli. The intron-encoded protein LtrA has reverse
transcriptase and RNA maturase activities, with the latter activity shown both in vivo and in
vitro, a first for any group II intron-encoded protein. As for the yeast mtDNA introns, the
DNA endonuclease activity of the Lactococcal intron is associated with RNP particles
containing both the intron-encoded protein and the excised intron RNA. Also, the intron RNA
cleaves the sense-strand of the recipient DNA by a reverse splicing reaction, whereas the
intron-encoded protein cleaves the antisense strand. The Lactococcal intron endonuclease can
be obtained in large quantities by coexpression of the LtrA protein with the intron RNA in E.
coli or reconstituted in vitro by incubating the expressed LtrA protein with in
vitro-synthesized intron RNA. Furthermore, the specificity of the endonuclease and reverse
splicing reactions can be changed predictably by modifying the RNA component. Expression in E.
coli facilitates the use of group II introns for the targeting of specific foreign sequences
to a desired site in DNA.

<>

<1>Matsuura, S.-I., Hirano, K., Zako, T., Katsura, S., Nagamune, T., Mizuno, A.
<2>Direct observation of sliding of restriction endonuclease EcoRI on a single DNA molecule.
<3>Eur. Biophys. J.
<4>29
<5>255
<6>2000
<7>In recent years, a fluorescence microscopy technique has been used to image the dynamics of
individual DNA and protein molecules.  For the advanced investigation of the molecular
mechanism in DNA-protein interactions such as sliding of restriction endonucleases on DNA
molecules, direct observation of a single protein molecule will be significant.  To observe
dynamics of individual proteins under a fluorescence microscopy on real-time, it requires
labeling proteins with a fluorescent dye.  In this study, therefore, we developed fluorescent
labeling system for a restriction endonuclease EcoRI as a model to label DNA binding proteins.
EcoRI bound on DNA molecules was treated with amine-reactive dye Oregon-Green500.
Consequently, we found that restriction endonuclease activity of labeled EcoRI was retained,
even though EcoRI was fluorescently labeled.  Moreover, when DNA-staining dye YOYO-1
concentration was YOYO-1 : nucleotide pair = 1:100 in molar ratio, EcoRI digested the DNA
molecules as unstained DNA.  Finally, we observed that fluorescent labeled EcoRI slid on
stained DNA straightening on 3-APTES-treated cover glass in the absence of Mg2+ using a
fluorescence microscopy.

<>

<1>Matsuzaki, S., Inoue, T., Tanaka, S.
<2>Evidence for the existence of a restriction-modification system common to several species of the family Vibronaceae.
<3>FEMS Microbiol. Lett.
<4>94
<5>191-194
<6>1992
<7>A broad-host-range vibriophage KVP40 originally isolated on Vibrio parahaemolyticus 1010 was
restricted and modified by strains of at least five Vibrio and one Photobacterium species.
1010 was a non-restricting host. An anti-restriction mutant KVP40 aar1 was isolated after
propagating the phage on a restricting host, V. anguillarum VIB36, as well as the parental
phage grown on VIB36, showed much higher efficiencies of plating on all the restricting hosts
as compared with the parental phage grown on 1010, indicating that these restricting hosts
probably share a common restriction-modification system active in vivo on KVP40.

<>

<1>Matsuzawa, T., Mori, K., Kadowaki, T., Shimada, M., Tashiro, K., Kuhara, S., Inagawa, H., Soma, G., Takegawa, K.
<2>Genome Sequence of Pantoea agglomerans Strain IG1.
<3>J. Bacteriol.
<4>194
<5>1258-1259
<6>2012
<7>Pantoea agglomerans is a Gram-negative bacterium that grows symbiotically with various plants.
Here we report the 4.8-Mb genome sequence of P. agglomerans
strain IG1. The lipopolysaccharides derived from P. agglomerans IG1 have been
shown to be effective in the prevention of various diseases, such as bacterial or
viral infection, lifestyle-related diseases. This genome sequence represents a
substantial step toward the elucidation of pathways for production of
lipopolysaccharides.

<>

<1>Matveyev, A.V., Young, K.T., Meng, A., Elhai, J.
<2>DNA methyltransferases of the cyanobacterium Anabaena PCC 7120.
<3>Nucleic Acids Res.
<4>29
<5>1491-1506
<6>2001
<7>From the characterization of enzyme activities and the analysis of genomic sequences, the
complement of DNA methyltransferases (MTases) possessed by the cyanobacterium Anabaena PCC
7120 has been deduced. Anabaena has nine DNA MTases. Four are associated with Type II
restriction enzymes (AvaI, AvaII, AvaIII and the newly recognized inactive AvaIV), and five
are not. Of the latter, four may be classified as solitary MTases, those whose function lies
outside of a restriction/modification system. The group is defined here based on biochemical
and genetic characteristics. The four solitary MTases, DmtA/M.AvaVI, DmtB/M.AvaVII,
DmtC/M.AvaVIII and DmtD/M.AvaIX, methylate at GATC, GGCC, CGATCG and RCCGGY, respectively.
DmtB methylates cytosines at the N4 position, but its sequence is more similar to N6-adenine
MTases than to cytosine-specific enzymes, indicating that it may have evolved from the former.
The solitary MTases, appear to be of ancient origin within cyanobacteria, while the
restriction MTases appear to have arrived by recent horizontal transfer as did five now
inactive Type I restriction systems. One Mtase, M.AvaV, cannot reliably be classified as
either a solitary or restriction MTase. It is structurally unusual and along with a few
proteins of prokaryotic and eukaryotic origin defines a structural class of MTases distinct
from all previously described.

<>

<1>Matvienko, N.I., Kramarov, V.M., Irismetov, A.A.
<2>Isolation and characterization of a DNA-methylase from Flavobacterium okeanokoites.
<3>Bioorg. Khim.
<4>11
<5>953-956
<6>1985
<7>FokI DNA methylase has been isolated from a cell extract of Flavobacterium
okeanokoites by gel filtration followed by gel chromatography on
hydroxylapatite.  The purified enzyme methylated the DNA of plasmid pBR322,
making it resistant to the subsequent action of the site-specific FokI
endonuclease.  It has been shown that it is the cytosine residues that are
methylated.  This modification does not protect the DNA from hydrolysis by
BbvI, BmeI, and EcoRII endonucleases, the recognition sites of which contain
cytosine, which shows the specific nature of the methylation of DNA by the FokI
methylase.

<>

<1>Matvienko, N.I., Kramarov, V.M., Pachkunov, D.M.
<2>Isolation and some properties of the site-specific endonuclease and methylase Bme216I from Bacillus megaterium 216.
<3>Eur. J. Biochem.
<4>165
<5>565-570
<6>1987
<7>The site-specific endonuclease Bme216I was isolated as a homogeneous
preparation by chromatography on phosphocellulose, hydroxyapatite and
heparin-agarose.  The molecular mass of the enzyme, determined by gel
filtration and by electrophoresis under denaturing conditions, was found to be
60 kDa and 30 kDa respectively.  These data indicate that the native enzyme
consists of two identical subunits.  The enzyme recognized the pentanucleotide
sequence 5'-G^GACC-3' . 3'-CCTG^G-5' and cleaves the sequence as indicated by
arrows.  The optimal concentration for endonuclease reaction is 6-7 mM Mg2+.
The endonuclease relaxes its specificity in the presence of glycerol or
dimethyl sulfoxide at low Mg2+ concentration (1-3 mM).  Methylase Bme216I,
which protects DNA against endonuclease Bme216I action by DNA methylation, was
isolated from the same bacterial strain.

<>

<1>Matvienko, N.I., Pachkunov, D.M., Kramarov, V.M.
<2>The recognition sequence of site-specific endonuclease BbvII from Bacillus brevis 80.
<3>FEBS Lett.
<4>177
<5>23-26
<6>1984
<7>Site-specific endonuclease BbvII from Bacillus brevis 80 recognizes the
non-symmetrical hexanucleotide and cleaves DNA at distances of 2 and 6
nucleotides from the recognition site:5'-GAAGACNN^ 3'-CTTCTGNNNNNN^This enzyme
may be used in molecular cloning for vectors with multiple restriction sites.

<>

<1>Matvienko, N.N., Kramarov, V.M., Ivanov, L.Y., Matvienko, N.I.
<2>Bce83I, a restriction endonuclease from Bacillus cereus 83 which recognizes novel nonpalindromic sequence 5'-CTTGAG-3' and is stimulated by S-adenosylmethionine.
<3>Nucleic Acids Res.
<4>20
<5>1803
<6>1992
<7>Restriction endonuclease Bce83I has been purified by chromatography on blue-sepharose and
hydroxyapatite. It recognizes 4,6,5,13 and more than 20 sites on pUC18, pBR322, M13mp18,
lambda and T7 DNA, respectively.

<>

<1>Matvienko, N.N., Kramarov, V.M., Zheleznaya, L.A., Matvienko, N.I.
<2>New site-specific endonuclease and methylase from Bacillus licheniformis 736.
<3>Biokhimiia
<4>58
<5>1139-1153
<6>1993
<7>*
The site-specific endonuclease R.Bli736I and methylase M.Bli736I have been isolated from the
Bacilus licheniformis strain 736 by blue-agarose, hydroxyapatite-Ultragel and
heparin-Sepharose
chromatography. The enzymes are free from interfering impurities. R.Bli736I recognizes the
sequence:

   5'-GGTCTCN^-3'
   3'-CCAGAGNNNNN^-5'

on the DNA and cleaves the DNA as indicated by arrows to form single-stranded 4-nucleotide
5'-protruding termini. This enzyme is an isoschizomer of Eco31I isolated from E.coli.


<>

<1>Matvienko, N.N., Kramarov, V.M., Zheleznaya, L.A., Matvienko, N.I.
<2>Isolation of site-specific endonuclease and methylase from Bacillus cereus 83.
<3>Biokhimiia
<4>58
<5>1845-1860
<6>1993
<7>
 The site-specific endonuclease R.Bce83I and methylase M.Bce83I were isolated from Bacillus
 cereus 83 by three consecutive chromatographies on blue agarose, hydroxyapatite and
 heparin-Sepharose. R.Bce83I recognizes the
 

     5'-CTTGAG16N^-3'
     3'-GAACTC14N^-5'
 

 sequences on the DNA and cleaves the DNA as indicated by the arrows. The endonuclease is
 stimulated by S-adenosyl-L-methionine and may consequently be referred to as a type IV
 restriction enzyme.


<>

<1>Matvienko, N.N., Zeleznaja, L.A., Matvienko, N.I.
<2>BspLU11I, a novel site specific endonuclease which cleaves 5'-ACATGT-3'.
<3>Nucleic Acids Res.
<4>21
<5>1495
<6>1993
<7>
<>

<1>Matvienko, N.N., Zheleznaya, L.A., Chernyshova, E.E., Buryanov, Y.I., Matvienko, N.I.
<2>Peculiarities of gene expression of the EcoRII modification-restriction system.
<3>Biokhimiia
<4>62
<5>1314-1318
<6>1997
<7>The restriction-modification genes of the EcoRII system have been cloned into plasmids under
control of phage-specific promoters T7 and SP6.  The transcription was induced by cell
infection with the recombinant M13 phages with the corresponding genes of phage
RNA-polymerases under control of the Plac-promoter in the presence of IPTG.  The induction
yields significant amounts of EcoRII DNA-methylase for both phage-specific promoters.  In both
cases no increase in EcoRII endonuclease expression could be achieved.  We hypothesize that
the expression of the endonuclease gene is regulated on the translational level.

<>

<1>Matvienko, N.N., Zheleznaya, L.A., Zelinskaya, N.V., Matvienko, N.I.
<2>Site-specific DNA-methylase M.BspST5I methylates only one strand of the recognized site.
<3>Biokhimiia
<4>62
<5>304-306
<6>1997
<7>We recently isolated a site-specific adenine DNA-methylase, M.BspST5I, which methylates only
one strand of the recognized site GCA*TC.  The methylated base is indicated by an asterisk.

<>

<1>Matyi, S.A., Hoyt, P.R., Hosoyama, A., Yamazoe, A., Fujita, N., Gustafson, J.E.
<2>Draft Genome Sequences of Elizabethkingia meningoseptica.
<3>Genome Announcements
<4>1
<5>e00444-13
<6>2013
<7>Elizabethkingia meningoseptica is ubiquitous in nature, exhibits a multiple-antibiotic
resistance phenotype, and causes rare opportunistic
infections. We now report two draft genome sequences of E. meningoseptica type
strains that were sequenced independently in two laboratories.

<>

<1>Matyi, S.A., Ramaraj, T., Sundararajan, A., Lindquist, I.E., Devitt, N.P., Schilkey, F.D., Lamichhane-Khadka, R., Hoyt, P.R., Mudge, J., Gustafson, J.E.
<2>Draft Genomes of Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Strain MM66 and MM66 Derivatives with Altered Vancomycin Resistance Levels.
<3>Genome Announcements
<4>2
<5>e00688-14
<6>2014
<7>The draft genomes of heterogeneous vancomycin-intermediate Staphylococcus aureus  (hVISA)
strain MM66 and MM66 isolates demonstrating altered vancomycin resistance
levels were produced in an effort to provide information on mutations
contributing to the vancomycin resistance levels observed in these strains.

<>

<1>Maus, I., Wibberg, D., Stantscheff, R., Eikmeyer, F.G., Seffner, A., Boelter, J., Szczepanowski, R., Blom, J., Jaenicke, S., Konig, H., Puhler, A., Schluter, A.
<2>Complete Genome Sequence of the Hydrogenotrophic, Methanogenic Archaeon Methanoculleus bourgensis Strain MS2T, Isolated from a Sewage Sludge Digester.
<3>J. Bacteriol.
<4>194
<5>5487-5488
<6>2012
<7>Methanoculleus bourgensis, of the order Methanomicrobiales, is a dominant methanogenic
archaeon in many biogas-producing reactor systems fed with renewable
primary products. It is capable of synthesizing methane via the hydrogenotrophic
pathway utilizing hydrogen and carbon dioxide or formate as the substrates. Here
we report the complete and finished genome sequence of M. bourgensis strain
MS2(T), isolated from a sewage sludge digester.

<>

<1>Mavian, C., Lopez-Bueno, A., Balseiro, A., Casais, R., Alcami, A., Alejo, A.
<2>The Genome Sequence of the Emerging Common Midwife Toad Virus Identifies an Evolutionary Intermediate within Ranaviruses.
<3>J. Virol.
<4>86
<5>3617-3625
<6>2012
<7>Worldwide amphibian population declines have been ascribed to global warming,
increasing pollution levels, and other factors directly related to human
activities. These factors may additionally be favoring the emergence of novel
pathogens. In this report, we have determined the complete genome sequence of the
emerging common midwife toad ranavirus (CMTV), which has caused fatal disease in
several amphibian species across Europe. Phylogenetic and gene content analyses
of the first complete genomic sequence from a ranavirus isolated in Europe show
that CMTV is an amphibian-like ranavirus (ALRV). However, the CMTV genome
structure is novel and represents an intermediate evolutionary stage between the
two previously described ALRV groups. We find that CMTV clusters with several
other ranaviruses isolated from different hosts and locations which might also be
included in this novel ranavirus group. This work sheds light on the phylogenetic
relationships within this complex group of emerging, disease-causing viruses.

<>

<1>Mavian, C., Lopez-Bueno, A., Fernandez, S., A, L.-B., Alcami, A., Alejo, A.
<2>Complete genome sequence of European sheatfish virus.
<3>J. Virol.
<4>86
<5>6365-6366
<6>2012
<7>Viral diseases are an increasing threat to the thriving aquaculture industry worldwide. An
emerging group of fish pathogens is formed by several ranaviruses, which have been isolated at
different locations from freshwater and seawater fish species since 1985.  We report the
complete genome sequence of European sheatfish ranavirus (ESV), the first ranavirus isolated
in Europe, which causes high mortality rates in infected sheatfish (Silurus glanis) and in
other species. Analysis of the genome sequence shows that ESV belongs to the amphibian-like
ranaviruses and is closely related to the epizootic hematopoietic necrosis virus (EHNV), a
disease agent geographically confined to the Australian continent and notifiable to the World
Organization for Animal Health.

<>

<1>Mavrodi, D.V., Mavrodi, O.V., McSpadden-Gardener, B.B., Landa, B.B., Weller, D.M., Thomashow, L.S.
<2>Identification of Differences in Genome Content among phlD-Positive Pseudomonas fluorescens Strains by Using PCR-Based Subtractive Hybridization.
<3>Appl. Environ. Microbiol.
<4>68
<5>5170-5176
<6>2002
<7>Certain 2,4-diacetylphloroglucinol-producing strains of Pseudomonas
fluorescens colonize roots and suppress soilborne diseases more
effectively than others from which they are otherwise phenotypically
almost indistinguishable. We recovered DNA fragments present in the
superior colonizer P. fluorescens Q8r1-96 but not in the less
rhizosphere-competent strain Q2-87. Of the open reading frames in 32
independent Q8r1-96-specific clones, 1 was similar to colicin M from
Escherichia coli, 3 resembled known regulatory proteins, and 28 had no
significant match with sequences of known function. Seven clones
hybridized preferentially to DNA from strains with superior rhizosphere
competence, and sequences in two others were highly expressed in vitro and
in the rhizosphere.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of Riemerella anatipestifer type strain (ATCC 11845).
<3>Standards in Genomic Sciences
<4>4
<5>144-153
<6>2011
<7>Riemerella anatipestifer (Hendrickson and Hilbert 1932) Segers et al. 1993 is the type species
of the genus Riemerella, which belongs to the family
Flavobacteriaceae. The species is of interest because of the position of the
genus in the phylogenetic tree and because of its role as a pathogen of
commercially important avian species worldwide. This is the first completed
genome sequence of a member of the genus Riemerella. The 2,155,121 bp long genome
with its 2,001 protein-coding and 51 RNA genes consists of one circular
chromosome and is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of Spirochaeta smaragdinae type strain (SEBR 4228).
<3>Standards in Genomic Sciences
<4>3
<5>136-144
<6>2010
<7>Spirochaeta smaragdinae Magot et al. 1998 belongs to the family Spirochaetaceae.  The species
is Gram-negative, motile, obligately halophilic and strictly
anaerobic and is of interest because it is able to ferment numerous
polysaccharides. S. smaragdinae is the only species of the family Spirochaetaceae
known to reduce thiosulfate or element sulfur to sulfide. This is the first
complete genome sequence in the family Spirochaetaceae. The 4,653,970 bp long
genome with its 4,363 protein-coding and 57 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Mavromatis, K. et al.
<2>Permanent draft genome sequence of the gliding predator Saprospira grandis strain Sa g1 (= HR1).
<3>Standards in Genomic Sciences
<4>6
<5>210-219
<6>2012
<7>Saprospira grandis Gross 1911 is a member of the Saprospiraceae, a family in the  class
'Sphingobacteria' that remains poorly characterized at the genomic level.
The species is known for preying on other marine bacteria via 'ixotrophy'. S.
grandis strain Sa g1 was isolated from decaying crab carapace in France and was
selected for genome sequencing because of its isolated location in the tree of
life. Only one type strain genome has been published so far from the
Saprospiraceae, while the sequence of strain Sa g1 represents the second genome
to be published from a non-type strain of S. grandis. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. The 4,495,250 bp long Improved-High-Quality draft of the genome with
its 3,536 protein-coding and 62 RNA genes is a part of the Genomic Encyclopedia
of Bacteria and Archaea project.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of Alicyclobacillus acidocaldarius type strain (104-IA).
<3>Standards in Genomic Sciences
<4>2
<5>9-18
<6>2010
<7>Alicyclobacillus acidocaldarius (Darland and Brock 1971) is the type species of the larger of
the two genera in the bacillal family 'Alicyclobacillaceae'. A.
acidocaldarius is a free-living and non-pathogenic organism, but may also be
associated with food and fruit spoilage. Due to its acidophilic nature, several
enzymes from this species have since long been subjected to detailed molecular
and biochemical studies. Here we describe the features of this organism, together
with the complete genome sequence and annotation. This is the first completed
genome sequence of the family 'Alicyclobacillaceae'. The 3,205,686 bp long genome
(chromosome and three plasmids) with its 3,153 protein-coding and 82 RNA genes is
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of Cryptobacterium curtum type strain (12-3).
<3>Standards in Genomic Sciences
<4>1
<5>93-100
<6>2009
<7>Cryptobacterium curtum Nakazawa etal. 1999 is the type species of the genus, and  is of
phylogenetic interest because of its very distant and isolated position within the family
Coriobacteriaceae. C. curtum is an asaccharolytic, opportunistic pathogen with a typical
occurrence in the oral cavity, involved in dental and oral infections like periodontitis,
inflammations and abscesses. Here we describe the features of this organism, together with the
complete genome sequence, and annotation. This is the first complete genome sequence of the
actinobacterial family Coriobacteriaceae, and this 1,617,804 bp long single replicon genome
with its 1364 protein-coding and 58 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of Vulcanisaeta distributa type strain (IC-017).
<3>Standards in Genomic Sciences
<4>3
<5>117-125
<6>2010
<7>Vulcanisaeta distributa Itoh et al. 2002 belongs to the family Thermoproteaceae in the phylum
Crenarchaeota. The genus Vulcanisaeta is characterized by a global
distribution in hot and acidic springs. This is the first genome sequence from a
member of the genus Vulcanisaeta and seventh genome sequence in the family
Thermoproteaceae. The 2,374,137 bp long genome with its 2,544 protein-coding and
49 RNA genes is a part of the Genomic Encyclopedia of Bacteriaand Archaea
project.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of Coraliomargarita akajimensis type strain (04OKA010-24).
<3>Standards in Genomic Sciences
<4>2
<5>290-299
<6>2010
<7>Coraliomargarita akajimensis Yoon et al. 2007 is the type species of the genus
Coraliomargarita. C. akajimensis is an obligately aerobic, Gram-negative,
non-spore-forming, non-motile, spherical bacterium that was isolated from
seawater surrounding the hard coral Galaxea fascicularis. C. akajimensis is of
special interest because of its phylogenetic position in a genomically
under-studied area of the bacterial diversity. Here we describe the features of
this organism, together with the complete genome sequence, and annotation. This
is the first complete genome sequence of a member of the family Puniceicoccaceae.
The 3,750,771 bp long genome with its 3,137 protein-coding and 55 RNA genes is a
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of the moderate thermophile Anaerobaculum mobile type strain (NGA(T)).
<3>Standards in Genomic Sciences
<4>8
<5>47-57
<6>2013
<7>Anaerobaculum mobile Menes and Muxi 2002 is one of three described species of the genus
Anaerobaculum, family Synergistaceae, phylum Synergistetes. This anaerobic
and motile bacterium ferments a range of carbohydrates and mono- and dicarboxylic
acids with acetate, hydrogen and CO2 as end products. A. mobile NGA(T) is the
first member of the genus Anaerobaculum and the sixth member of the phylum
Synergistetes with a completely sequenced genome. Here we describe the features
of this bacterium, together with the complete genome sequence, and annotation.
The 2,160,700 bp long single replicon genome with its 2,053 protein-coding and 56
RNA genes is part of the G enomic E ncyclopedia of Bacteria and Archaea project.

<>

<1>Mavromatis, K. et al.
<2>Complete genome sequence of the bile-resistant pigment-producing anaerobe Alistipes finegoldii type strain (AHN2437(T)).
<3>Standards in Genomic Sciences
<4>8
<5>26-36
<6>2013
<7>Alistipes finegoldii Rautio et al. 2003 is one of five species of Alistipes with  a validly
published name: family Rikenellaceae, order Bacteroidetes, class
Bacteroidia, phylum Bacteroidetes. This rod-shaped and strictly anaerobic
organism has been isolated mostly from human tissues. Here we describe the
features of the type strain of this species, together with the complete genome
sequence, and annotation. A. finegoldii is the first member of the genus
Alistipes for which the complete genome sequence of its type strain is now
available. The 3,734,239 bp long single replicon genome with its 3,302
protein-coding and 68 RNA genes is part of the G enomic E ncyclopedia of Bacteria
and Archaea project.

<>

<1>Mavromatis, K., Doyle, C.K., Lykidis, A., Ivanova, N., Francino, M.P., Chain, P., Shin, M., Malfatti, S., Larimer, F., Copeland, A., Detter, J.C., Land, M., Richardson, P.M., Yu, X.J., Walker, D.H., McBride, J.W., Kyrpides, N.C.
<2>The genome of the obligately intracellular bacterium Ehrlichia canis reveals themes of complex membrane structure and immune evasion  strategies.
<3>J. Bacteriol.
<4>188
<5>4015-4023
<6>2006
<7>Ehrlichia canis, a small obligately intracellular, tick-transmitted, gram-negative,
alpha-proteobacterium, is the primary etiologic agent of
globally distributed canine monocytic ehrlichiosis. Complete genome
sequencing revealed that the E. canis genome consists of a single circular
chromosome of 1,315,030 bp predicted to encode 925 proteins, 40 stable RNA
species, 17 putative pseudogenes, and a substantial proportion of
noncoding sequence (27%). Interesting genome features include a large set
of proteins with transmembrane helices and/or signal sequences and a
unique serine-threonine bias associated with the potential for O
glycosylation that was prominent in proteins associated with pathogen-host
interactions. Furthermore, two paralogous protein families associated with
immune evasion were identified, one of which contains poly(G-C) tracts,
suggesting that they may play a role in phase variation and facilitation
of persistent infections. Genes associated with pathogen-host interactions
were identified, including a small group encoding proteins (n = 12) with
tandem repeats and another group encoding proteins with eukaryote-like
ankyrin domains (n = 7).

<>

<1>Mavrommatis, K. et al.
<2>Complete genome sequence of Capnocytophaga ochracea type strain (VPI 2845).
<3>Standards in Genomic Sciences
<4>1
<5>101-109
<6>2009
<7>Capnocytophaga ochracea (Prevot et al. 1956) Leadbetter et al. 1982 is the type species of the
genus Capnocytophaga. It is of interest because of its location in
the Flavobacteriaceae, a genomically not yet charted family within the order
Flavobacteriales. The species grows as fusiform to rod shaped cells which tend to
form clumps and are able to move by gliding. C. ochracea is known as a
capnophilic (CO(2)-requiring) organism with the ability to grow under anaerobic
as well as aerobic conditions (oxygen concentration larger than 15%), here only
in the presence of 5% CO(2). Strain VPI 2845(T), the type strain of the species,
is portrayed in this report as a gliding, Gram-negative bacterium, originally
isolated from a human oral cavity. Here we describe the features of this
organism, together with the complete genome sequence, and annotation. This is the
first completed genome sequence from the flavobacterial genus Capnocytophaga, and
the 2,612,925 bp long single replicon genome with its 2193 protein-coding and 59
RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Maxwell, A., Halford, S.E.
<2>The SalGI restriction endonuclease.  (Enzyme specificity).
<3>Biochem. J.
<4>203
<5>93-98
<6>1982
<7>We have analysed the kinetics of DNA cleavage in the reaction between the SalGI
restriction endonuclease and plasmid pMB9.  This reaction is subject to
competitive inhibiton by DNA sequences outside the SalGI recognition site; we
have determined the Km and Vmax. for the reaction of this enzyme at its
recognition site and the KI for its interaction at other DNA sequences.  We
conclude that the specificity of DNA cleavage by the enzyme is only partly
determined by the discrimination it shows for binding at its recognition
sequence compared with binding to other DNA sequences.

<>

<1>Maxwell, A., Halford, S.E.
<2>The SalGI restriction endonuclease.  (Purification and properties).
<3>Biochem. J.
<4>203
<5>77-84
<6>1982
<7>The type II restriction endonuclease SalGI has been purified to near
homogeneity.  At least 80% of the protein remaining after the final stage of
the preparation is SalGI restriction endonuclease; no contaminating nucleases
remain detectable.  The principal form of the protein under both native and
denaturing conditions is a monomer of Mr about 29000. The optimal conditions
for both enzyme stability and enzyme activity have been determined.

<>

<1>Maxwell, A., Halford, S.E.
<2>The mechanism of DNA cleavage by restriction endonuclease SalGI.
<3>Biochem. Soc. Trans.
<4>9
<5>227P
<6>1981
<7>The SalGI restriction endonuclease cleaves duplex DNA only at its recognition
site.  Under optimal conditions (pH 8, 10 mM MgCl2), both strands of the DNA
are cleaved in one concerted reaction:  a covalently closed DNA molecule with
one SalGI recognition site is converted directly to linear DNA.  But under
other conditions (viz 1 mM MgCl2), each reaction of this enzyme cleaves either
one or both strands of the DNA; the covalently closed DNA is now converted into
either the open-circle or the linear forms, the two being produced
simultaneously rather than consecutively.  The enzyme will also cleave the DNA
nicked at the SalGI recognition site but this second reaction is much slower
than the first.  The SalGI restriction enzyme therefore interacts with DNA by a
fundamentally different mechanism from some other restriction enzymes such as
EcoRI.

<>

<1>Maxwell, A., Halford, S.E.
<2>The SalGI restriction endonuclease.  Mechanism of DNA cleavage.
<3>Biochem. J.
<4>203
<5>85-92
<6>1982
<7>The cleavage of supercoiled DNA of plasmid pMB9 by restriction endonuclease
SalGI has been studied.  Under the optimal conditions for this reaction, the
only product is the linear form of the DNA, in which both strands of the duplex
have been cleaved at the SalGI recognition site.  DNA molecules cleaved in one
strand at this site were found to be poor substrates for the SalGI enzyme.
Thus, both strands of the DNA appear to be cleaved in a concerted reaction.
However, under other conditions, the enzyme cleaves either one or both strands
of the DNA; the supercoiled substrate is then converted to either open-circle
or linear forms, the two being produced simultaneously rather than
consecutively.  We propose a mechanism for the SalGI restriction endonuclease
which accounts fo the reactions of this enzyme under both optimal and other
conditions.  These reactions were unaffected by the tertiary structure of the
DNA.

<>

<1>May, B.J., Zhang, Q., Li, L.L., Paustian, M.L., Whittam, T.S., Kapur, V.
<2>Complete genomic sequence of Pasteurella multocida, Pm70.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>3460-3465
<6>2001
<7>We present here the complete genome sequence of a common avian clone of Pasteurella multocida,
Pm70. The genome of Pm70 is a single circular chromosome 2,257,487 base pairs in length and
contains 2,014 predicted coding regions, 6 ribosomal RNA operons, and 57 tRNAs. Genome-scale
evolutionary analyses based on pairwise comparisons of 1,197 orthologous sequences between P.
multocida, Haemophilus influenzae, and Escherichia coli suggest that P. multocida and H.
influenzae diverged approximately 270 million years ago and the gamma subdivision of the
proteobacteria radiated about 680 million years ago. Two previously undescribed open reading
frames, accounting for approximately 1% of the genome, encode large proteins with homology to
the virulence-associated filamentous hemagglutinin of Bordetella pertussis. Consistent with
the critical role of iron in the survival of many microbial pathogens, in silico and
whole-genome microarray analyses identified more than 50 Pm70 genes with a potential role in
iron acquisition and metabolism. Overall, the complete genomic sequence and preliminary
functional analyses provide a foundation for future research into the mechanisms of
pathogenesis and host specificity of this important multispecies pathogen.

<>

<1>May, M.S., Hattman, S.
<2>Analysis of bacteriophage deoxyribonucleic acid sequences methylated by host- and R-factor-controlled enzymes.
<3>J. Bacteriol.
<4>123
<5>768-770
<6>1975
<7>Phages lambda and fd were propagated in Escherichia coli strains that have
either host K-12 or the N-3 R-factor deoxyribonucleic acid-cytosine methylase
activity.  Pyrimidine tracts containing 3H-labeled 5-methylcytosine (MeC) were
analyzed; in all cases, the major methylated sequence was 5'...C-MeC-T ...3'.

<>

<1>May, M.S., Hattman, S.
<2>Deoxyribonucleic acid-cytosine methylation by host- and plasmid-controlled enzymes.
<3>J. Bacteriol.
<4>122
<5>129-138
<6>1975
<7>Deoxyribonucleic acid (DNA)-cytosine methylation specified by the wild-type Escherichia coli
K12 mec+ gene and by the N-3 drug resistance (R) factor was studied in vivo and in vitro.
Phage lambda and fd were propagated in the presence of L-[methyl-3H]methionine in various host
bacteria. The in vivo labeled DNA was isolated from purified phage and depurinated by formic
acid-diphenylamine treatment. The resulting pyrimidine oligonucleotide tracts were separated
according to size and base composition by chromatography on diethylaminoethyl-cellulose in 7 M
urea at pH 5.5 and 3.5, respectively. The distribution of labeled 5-methylcytosine in DNA
pyrimidine tracts was identical for phage grown in mec+ and mec- (N-3) cells. For phage lambda
the major 5-methylcytosine-containing tract was the tripyrimidine, C2T; for both fd.mec- (N-3)
DNA and fd.mec+ DNA, C2T was the sole 5-methylcytosine-containing tract. When various lambda
DNAs were methylated to saturation in vitro by crude extracts from mec+ and mec- (N-3) cells,
the extent of cytosine methylation was the same. This is in contrast to in vivo methylation
where k.mec- (N-3) DNA contains twice as many 5-methylcytosines per genome as k.mec+ DNA.
Therefore, we suggest that the K12 met+ cytosine methylase and the N-3 plasmid modification
methylases are capable of recognizing the same nucleotide sequences, but that the in vivo
methylation rate is lower in mec+ cells.

<>

<1>Mayer, A., Barany, F.
<2>DNA photoaffinity crosslinking to TaqI endonuclease by a phosphorothioate-linked aryl azide.
<3>FASEB J.
<4>6
<5>A487
<6>1992
<7>To identify amino acid residues required for the function of TaqI endonuclease
(cleaves T^CGA), a new type of DNA photoaffinity crosslinking reagent was
designed.  DNA (16-mer) was chemically synthesized in which a sulphur atom
replaced a non-bridging phosphate oxygen at the position 5' to the C (scissile
phosphate).  The two resulting phosphorothioate diasteriomers were separated
using HPLC (C-18 column) and then alkylated with p-azidophenacyl bromide.  The
endonuclease bound the modified substrates in a sequence-nonspecific manner and
was crosslinked in the presence of UV light (366nm) with an efficiency of 25%.
The TaqI-DNA crosslink was purified from unreacted material by FPLC (MonoQ).
This species has proven highly resistant to a variety of proteases, and efforts
are underway to effectively fragment the protein and identify the crosslinked
residue.  The approach described here should be generally applicable to the
study of other DNA binding proteins, given the facility of reagent synthesis
and the flexibility of photoactive crosslinker placement.

<>

<1>Mayer, A.N.
<2>Defining the contacts between Taq endonuclease and the DNA phosphate backbone.
<3>Diss. Abstr.
<4>57
<5>2359B
<6>1996
<7>The restriction endonuclease TaqI exhibits extreme specificity for its cognate sequence, TCGA,
but a direct readout model fails to account for this property.  This study examines the role
of phosphate contacts in the enzyme-substrate and transition-state complexes.  An S-methyl
group was introduced into each of the pTpCpGpApNpN internucleotide linkages using a hybrid
chemical-enzymatic synthesis, in which sulfur substitutions of non-bridging phosphate oxygens
directed the placement of methyl groups.  The resulting twelve diastereomerically pure
phosphate-modified substrates were tested for binding and cleavage by TaqI endonuclease.  The
largest binding effects were produced by pro-Sp methylations at the pTpCpGA phosphates, which
destablized the enzyme-substrate complex by 1.0-1.6 kcal/mol.  Cleavage of the modified strand
was inhibited completely by modifications at the TpCpGpA phosphates, and inhibited
significantly at the TCGApNp phosphates.  Cleavage of both strands was completely inhibited by
modification of the TCGpA linkage.  Effects on the cleavage of the unmodified strand
implicated phosphate modifications which caused global perturbations in the structure of the
transition-state complex.  These results support a model to account for the specificity of
TaqI, in which sequence-specific phosphate contacts are formed in the transition state, thus
amplifying the apparent contribution of base contacts to the transition-state complex.  To
identify amino acid residues which are in contact with the DNA, a sequence-specific
photoaffinity reagent was designed which exploits the finding that modification of the Rp
oxygen of the scissile phosphate does not interfere with substrate binding.  Accordingly, the
scissile phosphate was substituted with an Rp phosphorothioate group to direct the placement
of the bifunctional reagent, p-azidophenacyl bromide.  TaqI bound the photoaffinity reagent
specifically and formed a covalent adduct with the enzyme in the presence of UV light.  Upon
digestion of the covalent complex and isolation of a labeled peptide, the modified amino acid
was identified as Tyr161.  This residue was changed to phenylalanine by site-directed
mutagenesis, and the resulting Y161F mutant was characterized.  Removal of the Tyr161 hydroxyl
group lowered both the kcat and the KM 5-fold, indicating that while this residue may be near
the scissile phosphate, it is not critically required for catalysis.

<>

<1>Mayer, A.N., Barany, F.
<2>Interaction of TaqI endonuclease with the phosphate backbone.
<3>J. Biol. Chem.
<4>269
<5>29067-29076
<6>1994
<7>The restriction endonuclease TaqI exhibits extreme specificity for its cognate sequence, TCGA,
but a direct hydrogen bond readout model fails to account for this property. The present study
examines the role of phosphate contacts in the enzyme-substrate and transition state
complexes. An S-methyl group was introduced into each of the pTpCpGpApNpN internucleotide
linkages using a hybrid chemical-enzymatic synthesis, in which sulfur substitutions of
nonbridging phosphate oxygens directed the placement of methyl groups. The resulting 12
diastereomerically pure phosphate-modified substrates were tested for binding and cleavage by
TaqI. The largest binding effects were induced by pro-Sp methylations at the pTpCpGA
phosphates, which destabilized the enzyme-substrate complex by 1.0-1.6 kcal/mol. Cleavage of
the modified strand was inhibited completely by modifications at the TpCpGpA phosphates and
inhibited significantly at the TCGApNp phosphates. Cleavage of both strands was completely
inhibited by modification of the TCGpA linkage. Effects on the cleavage of the unmodified
strand were used to implicate phosphate modifications that caused global perturbations in the
structure of the transition state complex. These results lend support for a model for the
specificity of TaqI, in which sequence-specific phosphate contacts are formed in the
transition state, thus amplifying the apparent contribution of base contacts to transition
state stabilization.

<>

<1>Mayer, A.N., Barany, F.
<2>Photoaffinity cross-linking of TaqI restriction endonuclease using an aryl azide linked to the phosphate backbone.
<3>Gene
<4>153
<5>1-8
<6>1995
<7>In an effort to identify amino acid (aa) residues near the active site of TaqI restriction
endonuclease (ENase), a sequence-specific photoaffinity reagent was designed. This reagent
exploits the finding that modification of the Rp oxygen of the scissile phosphate does not
interfere with substrate binding. The TpCGA phosphate was substituted with an Rp
phosphorothioate group to direct the placement of the heterobifunctional reagent
p-azidophenacyl bromide. TaqI bound the photoaffinity reagent specifically and formed a
covalent adduct with the ENase in the presence of UV light. The modified aa was identified as
Tyr161. This aa was changed to Phe by site-directed mutagenesis, and the resulting Y161F
mutant was characterized. Removal of the Tyr161 hydroxyl group lowered both the kcat and the
Km five-fold, indicating that, while this aa may be near the scissile phosphate, it is not
critically rquired for catalysis.

<>

<1>Mayer, H.
<2>Optimization of the EcoRI* activity of EcoRI endonuclease.
<3>FEBS Lett.
<4>90
<5>341-344
<6>1978
<7>None

<>

<1>Mayer, H., Goebel, W.
<2>Isolierung der Restriktionsendonuclease EcoRI aus einem Antibiotka-Resistenz-freien Stamm von Escherichia coli.
<3>Hoppe Seylers Z. Physiol. Chem.
<4>356
<5>253-254
<6>1975
<7>Der Antibiotikaresistenz-(R-Faktor R1drd16 ist ein durch Deletionsmutation
entstandenes Derivat des ursprunglichen R1-Faktors, der das Restriktionsenzym
EcoRI kodiert.  Der R1drd16-Faktor determiniert nur noch Resistenz gegen
Kanamycin.  Durch weitere Mutation konnte ein Plasmid erhalten werden, das
keine Antibiotikaresistenz, wohl aber noch die Synthese von EcoRI determiniert.
Im Grobfermentationsannsatz konnte der E.-coli-Stamm, der diesen mutierten
R-Faktor tragt, mit einer Ausbeute von 35 g Bakterienfeuchtmasse/Igezuchtet
werden, ohne dab ein erkennbarer Verlust des Plasmids zu beobachten wr.  Nach
Zellaufschlub wurden in einem einzigen Fallungsschritt mit
Cetyltrimetylam-moniumbromid Zellmembran und DNA entfernt.  Bei der
Chromatographie an  Phosphocellulose und Hydroxyl-apatit verhalt sich das
Exonuclease-freie Enzym wie EcoRI des ursprunglichen Plasmids.  Auch die
Spaltungs-produkte von verschiedenen bakteriellen DNAs mit beiden
Enzympraparationen sind identisch.

<>

<1>Mayer, H., Grosschedl, R., Schutte, H., Hobom, G.
<2>ClaI, a new restriction endonuclease from Caryophanon latum L.
<3>Nucleic Acids Res.
<4>9
<5>4833-4845
<6>1981
<7>From Caryophanon latum L a site specific restriction endonuclease (ClaI) has been purified,
which recognises the DNA hexanucleotide palindrome 5'-A-T-^C-G-A-T-3'. Staggered cleavage
generates DNA restriction fragments with 5'-terminal pCG extensions. A ClaI map of
bacteriophage lambda has been determined, which indicates cleavage inhibition due to adenine
methylation at overlapping ClaI-GATC recognition sequences. Plasmid pBR322 is cut only once,
in the tetracycline promoter region, and can, therefore, be used as a vector system for
cloning fragments derived from ClaI digestions, and in addition for fragments generated by
TaqI, HpaII, and several other enzymes.

<>

<1>Mayer, H., Reichenbach, H.
<2>Restriction Endonucleases: General Survey Procedure and Survey of Gliding Bacteria.
<3>J. Bacteriol.
<4>136
<5>708-713
<6>1978
<7>Among 120 strains of gliding bacteria which were screened for restriction
endonucleases, 27 were found positive.  Additionally, three strains carried
enzymes able to release the supercoiled state of closed circular DNA.  By using
a new rapid method, restriction endonuclease activity was released by stirring
about 0.5 g of cells (fresh weight) in a motor-driven glass homogenizer in
buffer containing Triton X-101, ethylenediaminetetraacetic acid, and
mercaptoethanol.  A yield from 60 to 80% of the total activity present in the
cells was obtained with minimal destruction of the cells.  The enzyme activity
in the crude extract was measured semi-quantitatively be digestion of DNA and
subsequent separation of the fragments on an agarose slab gel.  The method
appears to be generally applicable for the extraction of restriction
endonucleases from gram-negative bacteria on an analytical scale and in a
modified form for large-scale preparation of restriction enzymes.

<>

<1>Mayer, M.J., Narbad, A., Gasson, M.J.
<2>Molecular characterization of a Clostridium difficile bacteriophage and its cloned biologically active endolysin.
<3>J. Bacteriol.
<4>190
<5>6734-6740
<6>2008
<7>Clostridium difficile infection is increasing in both frequency and
severity, with the emergence of new highly virulent strains highlighting
the need for more rapid and effective methods of control. Here, we show
that bacteriophage endolysin can be used to inhibit and kill C. difficile.
The genome sequence of a novel bacteriophage that is active against C.
difficile was determined, and the bacteriophage endolysin gene was
subcloned and expressed in Escherichia coli. The partially purified
endolysin was active against 30 diverse strains of C. difficile, and
importantly, this group included strains of the major epidemic ribotype
027 (B1/NAP1). In contrast, a range of commensal species that inhabit the
gastrointestinal tract, including several representatives of the
clostridium-like Firmicutes, were insensitive to the endolysin. This
endolysin provides a platform for the generation of both therapeutic and
detection systems to combat the C. difficile problem. To investigate a
method for the protected delivery and production of the lysin in the
gastrointestinal tract, we demonstrated the expression of active CD27L
endolysin in the lactic acid bacterium Lactococcus lactis MG1363.

<>

<1>Mayer, M.J., Payne, J., Gasson, M.J., Narbad, A.
<2>Genomic Sequence and Characterization of the Virulent Bacteriophage {phi}CTP1 from Clostridium tyrobutyricum and Heterologous Expression of Its Endolysin.
<3>Appl. Environ. Microbiol.
<4>76
<5>5415-5422
<6>2010
<7>The growth of Clostridium tyrobutyricum in developing cheese leads to
spoilage and cheese blowing. Bacteriophages or their specific lytic
enzymes may provide a biological control method for eliminating such
undesirable organisms without affecting other microflora. We isolated the
virulent bacteriophage phiCTP1 belonging to the Siphoviridae and have
shown that it is effective in causing lysis of sensitive strains. The
double-stranded DNA genome of phiCTP1 is 59,199 bp, and sequence analysis
indicated that it has 86 open reading frames. orf29 was identified as the
gene coding for the phage endolysin responsible for cell wall degradation
prior to virion release. We cloned and expressed the ctp1l gene in E. coli
and demonstrated that the partially purified protein induced lysis of C.
tyrobutyricum cells and reduced viable counts both in buffer and in milk.
The endolysin was inactive against a range of clostridial species but did
show lysis of Clostridium sporogenes, another potential spoilage organism.
Removal of the C-terminal portion of the endolysin completely abolished
lytic activity.

<>

<1>Mayer, W., Niveleau, A., Walter, J., Fundele, R., Haaf, T.
<2>Demethylation of the zygotic paternal genome.
<3>Nature
<4>403
<5>501-502
<6>2000
<7>In mammals, both parental genomes undergo dramatic epigenetic changes after fertilization to
form the diploid somatic genome.  Here we show that the paternal genome in the mouse is
significantly and actively demethylated within 6-8 hours of fertilization, before the onset of
DNA replication, whereas the maternal genome is demethylated after several cleavage divisions.
This active demethylation of the paternal genome may be associated with epigenetic remodelling
of sperm chromatin, in order to establish parent-specific developmental programmes during
early embryogenesis.

<>

<1>Maynard-Smith, M.D., McKelvie, J.C., Wood, R.J., Harmer, J.E., Ranasinghe, R.T., Williams, C.L., Coomber, D.M., Stares, A.F., Roach, P.L.
<2>Direct and continuous fluorescence-based measurements of Pyrococcus horikoshii DNA N-6 adenine methyltransferase activity.
<3>Anal. Biochem.
<4>418
<5>204-212
<6>2011
<7>N-6 methylation of adenine destabilises duplex DNA and this can increase the proportion of DNA
that dissociates into single strands. We
have investigated utilising this property to measure the DNA adenine
methyltransferase-catalyzed conversion of hemimethylated to fully
methylated DNA through a simple, direct, fluorescence-based assay. The
effects of methylation on the kinetics and thermodynamics of
hybridisation were measured by comparing a fully methylated
oligonucleotide product and a hemimethylated oligonucleotide substrate
using a 13-bp duplex labeled on adjacent strands with a fluorophore
(fluorescein) and quencher (dabcyl). Enzymatic methylation of the
hemimethylated GATC site resulted in destabilisation of the duplex,
increasing the proportion of dissociated DNA, and producing an
observable increase in fluorescence. The assay provides a direct
measurement of methylation rate in real time and is highly
reproducible, with a coefficient of variance over 48 independent
measurements of 3.6%. DNA methylation rates can be measured as low as
3.55 +/- 1.84 fmol s(-1) in a 96-well plate format, and the assay has
been used to kinetically characterise the Pyrococcus horikoshii DNA
adenine methyltransferase.

<>

<1>Mayo, B., Hardisson, C., Brana, A.F.
<2>Nucleolytic activities in Lactococcus lactis subsp. lactis NCDO 497.
<3>FEMS Microbiol. Lett.
<4>79
<5>195-198
<6>1991
<7>Two nucleolytic activities were detected in Lactococcus lactis subsp. lactis NCDO 497. One of
them was a specific endonuclease, located in the cytoplasm, with the typical characteristics
of type II restriction endonucleases. The second activity was a non-specific deoxyribonuclease
with exocytoplasmic location.

<>

<1>Maze, A. et al.
<2>Complete genome sequence of the probiotic Lactobacillus casei strain BL23.
<3>J. Bacteriol.
<4>192
<5>2647-2648
<6>2010
<7>The entire genome of Lactobacillus casei BL23, a strain with probiotic properties, has been
sequenced. The genomes of BL23 and the industrially
used probiotic strain Shirota YIT 9029 (Yakult) seem to be very similar.

<>

<1>Mazin, A.L., Vanyushin, B.F.
<2>Possible origin and evolution of enzymatic methylation of eukaryotic DNA.  Methylation of cytosine residues in three families of palindromes: RYRY, YYRR, and YYRYRR.
<3>Mol. Biol. (Mosk)
<4>24
<5>16-35
<6>1990
<7>On the basis of an analysis of the experimental data on the closest neighbors of the
5-methylcytosine residues in eukaryotic DNA it was established that the regions of methylation
CG and CNG may be included in three families of palindromes: RYRY, YYRR and YYRYRR.  It was
shown that the entire variety of methylated sequences detectable in their DNA can arise as a
result of mutational substitutions 5-MeC -> T, which occur in the deamination of 5-MeC
residues in prototype portions of each of these families: GCGC, CCGG and CCGCGG.  The question
of the multiplicity of DNA-methyltransferases in eukaryotes and their evolutionary origin from
prokaryotic methylases of the second type, which recognize analogous sequences in DNA, is
discussed.

<>

<1>Mazurek, M., Sowers, L.C.
<2>The paradoxical influence of thymine analogues on restriction endonuclease cleavage of oligodeoxynucleotides.
<3>Biochemistry
<4>35
<5>11522-11528
<6>1996
<7>Thymine residues in the DNA of eukaryotes may be replaced occasionally by uracil (U) or
5-(hydroxymethyl)uracil (H) as consequences of dUMP misincorporation or thymine oxidation,
respectively.  In this study, we constructed a series of 44-base oligonucleotides containing
site-specific U or H residues and 5'-fluorescein labels in order to probe the influence of
such modifications on sequence-specific DNA-protein interactions using several type II
restriction endonucleases.  We find that substitution within the recognition sites of several
restriction endonucleases increases initial cleavage velocity by up to an order of magnitude.
These results contrast dramatically with several previous studies which demonstrated that U
substitution in short oligonucleotides inhibits or prevents nuclease cleavage.  We propose
that this apparent paradox results because the rate-limiting step in the cleavage of longer
oligonucleotides is product release whereas for shorter oligonucelotides substrate binding is
most probably rate-limiting.  For longer oligonucleotides and DNA, more rapid release of the
cleaved, substituted oligonucleotides results in more rapid turnover and a faster apparent
cleavage rate.  The sequence length at which the transition is rate-limiting step occurs
likely corresponds to the size of th enzyme footprint on its DNA recognition site.  We
conclude that both U and H do perturb sequence-specific DNA - protein interactions, and the
magnitude of this effect is site-dependent.

<>

<1>Mazzarelli, J., Scholtissek, S., McLaughlin, L.W.
<2>Effects of functional group changes in the EcoRV recognition site on the cleavage reaction catalyzed by the endonuclease.
<3>Biochemistry
<4>28
<5>4616-4622
<6>1989
<7>Oligodeoxynucleotides have been prepared which contain changes in the
functional group pattern present in the EcoRV recognition site d(GATATC).
These modifications involve the deletion of specific functional groups or the
reversal of the relative positions of functional groups within the canonical
six base pair recognition site.  The duplex stability of these modified
oligodeoxynucleotides has been assessed by determining the thermodynamic
parameters characterizing helix formation.  Steady-state kinetic parameters
have been used to characterize the interaction of the modified
oligodeoxynucleotides with the EcoRV endonuclease.  The enzyme is very
sensitive to the deletion of either of the adenine amino or thymine methyl
groups, or the reversal of the relative positions of the adenine amino group
and thymine carboxy group which form an interstrand hydrogen bond in the major
groove of the B-DNA helix.  Conversely, deletion of the guanine amino group had
only minimal effects upon the measured kinetic parameters.  Deletion of the
exocyclic amino group from the inner dA-dT base pair resulted in the fragment
which interacted with the enzyme on the basis of observed inhibition
experiments but was not cleaved.  The results suggest that the endonuclease
interacts with its recognition sequence via contacts in the major groove of the
B-DNA helix and that both hydrogen bonding to the adenine amino groups and also
hydrophobic interactions with the thymine methyl groups are involved.

<>

<1>McCallum, C.M.
<2>TILLING for Arabidopsis chromomethylase function.
<3>Ph.D. Thesis, University of Washington, Seattle, USA
<4>
<5>
<6>2005
<7>In contrast to prokaryotic methyltransferases, comparatively little is known about the
detailed structure and function of eukaryotic cytosine-5-methyltransferase enzymes.  During
the course of my thesis work, I identified two DNA methyltransferase homologs that also
contained a chromodomain (termed "chromomethylases") in Arabidopsis thaliana.  It is thought
that chromodomains target proteins to interact with specific chromatin determinants; thus
chromomethylases might be involved in epigenetic silencing by linking chromatin structure to
DNA methylation.  In addition, the unique structure of the chromomethylases suggest they may
not play a general role in maintaining CpG DNA methylation patterns, as may be true for the
Arabidopsis methylase MET1 but, instead, may play a more specialized role in either
establishing methylation or maintaining methylation on CpNpG or other nonsymmetrical sites.
The two new genes, CMT2 and CMT3, were studied to determine their biological functions.  The
goal of my thesis work was to characterize the chromomethylase genes specifically in hopes of
determining the function of DNA methylation in plants.  Along the way I developed the reverse
genetics method TILLING (Targeting Induced Local Lesions IN Genomes) and explored genome wide
methylation targets of CMT3 using methylation profiling.  This work revealed that cmt3 mutants
appear to have decreased CpNpG methylation while CpG methylation is unaffected.

<>

<1>McCallum, C.M., Comai, L., Greene, E.A., Henikoff, S.
<2>Targeted screening for induced mutations.
<3>Nat. Biotechnol.
<4>18
<5>455-459
<6>2000
<7>With the accumulation of large-scale sequence data, emphasis in genomics has shifted from
determining gene structure to testing gene function, and this relies on reverse genetic
methodology. Here we explore the feasibility of screening for chemically induced mutations in
target sequences in Arabidopsis thaliana. Our TILLING (Targeting Induced Local Lesions IN
Genomes) method combines the efficiency of ethyl methanesulfonate (EMS)-induced mutagenesis
with the ability of denaturing high-performance liquid chromatography (DHPLC) to detect base
pair changes by heteroduplex analysis. Importantly, this method generates a wide range of
mutant alleles, is fast and automatable, and is applicable to any organism that can be
chemically mutagenized.

<>

<1>McCarren, J., DeLong, E.F.
<2>Proteorhodopsin photosystem gene clusters exhibit co-evolutionary trends and shared ancestry among diverse marine microbial phyla.
<3>Environ. Microbiol.
<4>9
<5>846-858
<6>2007
<7>Since the recent discovery of retinylidene proteins in marine bacteria (proteorhodopsins), the
estimated abundance and diversity of this gene
family has expanded rapidly. To explore proteorhodopsin photosystem
evolutionary and distributional trends, we identified and compared 16
different proteorhodopsin-containing genome fragments recovered from
naturally occurring bacterioplankton populations. In addition to finding
several deep-branching proteorhodopsin sequences, proteorhodopsins were
found in novel taxonomic contexts, including a betaproteobacterium and a
planctomycete. Approximately one-third of the proteorhodopsin-containing
genome fragments analysed, as well as a number of recently reported marine
bacterial whole genome sequences, contained a linked set of genes required
for biosynthesis of the rhodopsin chromophore, retinal. Phylogenetic
analyses of the retinal biosynthetic genes suggested their co-evolution
and probable coordinated lateral gene transfer into disparate lineages,
including Euryarchaeota, Planctomycetales, and three different
proteobacterial lineages. The lateral transfer and retention of genes
required to assemble a functional proteorhodopsin photosystem appears to
be a coordinated and relatively frequent evolutionary event. Strong
selection pressure apparently acts to preserve these light-dependent
photosystems in diverse marine microbial lineages.

<>

<1>McCarthy, A.J., Lindsay, J.A.
<2>The distribution of plasmids that carry virulence and resistance genes in Staphylococcus aureus is lineage associated.
<3>BMC Microbiol.
<4>12
<5>104
<6>2012
<7>Background: Staphylococcus aureus is major human and animal pathogen. Plasmids often carry
resistance genes and virulence genes that can
disseminate through S. aureus populations by horizontal gene transfer
(HGT) mechanisms. Sequences of S. aureus plasmids in the public domain
and data from multi-strain microarrays were analysed to investigate (i)
the distribution of resistance genes and virulence genes on S. aureus
plasmids, and (ii) the distribution of plasmids between S. aureus
lineages.
Results: A total of 21 plasmid rep gene families, of which 13 were
novel to this study, were characterised using a previously proposed
classification system. 243 sequenced plasmids were assigned to 39
plasmid groups that each possessed a unique combination of rep genes.
We show some resistance genes (including ermC and cat) and virulence
genes (including entA, entG, entJ, entP) were associated with specific
plasmid groups suggesting there are genetic pressures preventing
recombination of these genes into novel plasmid groups. Whole genome
microarray analysis revealed that plasmid rep, resistance and virulence
genes were associated with S. aureus lineages, suggesting
restriction-modification (RM) barriers to HGT of plasmids between
strains exist. Conjugation transfer (tra) complex genes were rare.
Conclusion: This study argues that genetic pressures are
restraining the spread of resistance and virulence genes amongst S.
aureus plasmids, and amongst S. aureus populations, delaying the
emergence of fully virulent and resistant strains.

<>

<1>McCarthy, C.B., Romanowski, V.
<2>Digestion of I-PpoI recognition sites in unfavorable sequence contexts achieved by changing the reaction conditions.
<3>Biochem. Genet.
<4>44
<5>61-67
<6>2006
<7>
<>

<1>McClain, M.S., Shaffer, C.L., Israel, D.A., Peek, R.M. Jr., Cover, T.L.
<2>Genome sequence analysis of Helicobacter pylori strains associated with gastric ulceration and gastric cancer.
<3>BMC Genomics
<4>10
<5>3
<6>2009
<7>BACKGROUND: Persistent colonization of the human stomach by Helicobacter
pylori is associated with asymptomatic gastric inflammation (gastritis)
and an increased risk of duodenal ulceration, gastric ulceration, and
non-cardia gastric cancer. In previous studies, the genome sequences of H.
pylori strains from patients with gastritis or duodenal ulcer disease have
been analyzed. In this study, we analyzed the genome sequences of an H.
pylori strain (98-10) isolated from a patient with gastric cancer and an
H. pylori strain (B128) isolated from a patient with gastric ulcer
disease. RESULTS: Based on multilocus sequence typing, strain 98-10 was
most closely related to H. pylori strains of East Asian origin and strain
B128 was most closely related to strains of European origin. Strain 98-10
contained multiple features characteristic of East Asian strains,
including a type s1c vacA allele and a cagA allele encoding an EPIYA-D
tyrosine phosphorylation motif. A core genome of 1237 genes was present in
all five strains for which genome sequences were available. Among the 1237
core genes, a subset of alleles was highly divergent in the East Asian
strain 98-10, encoding proteins that exhibited <90% amino acid sequence
identity compared to corresponding proteins in the other four strains.
Unique strain-specific genes were identified in each of the newly
sequenced strains, and a set of strain-specific genes was shared among H.
pylori strains associated with gastric cancer or premalignant gastric
lesions. CONCLUSION: These data provide insight into the diversity that
exists among H. pylori strains from diverse clinical and geographic
origins. Highly divergent alleles and strain-specific genes identified in
this study may represent useful biomarkers for analyzing geographic
partitioning of H. pylori and for identifying strains capable of inducing
malignant or premalignant gastric lesions.

<>

<1>McClarin, J.A., Frederick, C.A., Grable, J., Samudzi, C.T., Wang, B.-C., Greene, P., Boyer, H.W., Rosenberg, J.M.
<2>Structural studies on a DNA-EcoRI endonuclease recognition complex.
<3>Biomolecular Sterodynamics, Adenine Press, Sarma, R.H., Sarma, M.H., New York
<4>3
<5>45-68
<6>1986
<7>The 3 angstrom structure of a co-crystalline recognition complex between EcoRI
endonuclease and the cognate oligonucleotide TCGCGAATTCGCG was solved by the
ISIRT method using a platinum isomorphous derivative.  The complex possesses a
common two-fold symmetry axis which relates both strands of the
self-complementary oligonucleotide and the two identical subunits of the
protein dimer.  Each subunit is organized into an alpha/beta domain based on a
five stranded beta-sheet and an extension, called the arm, which wraps around
the DNA.  The primary beta-sheet consists of anti-parallel and parallel
segments which, respectively, contain the sites of DNA strand scission and
sequence recognition.  (DNA hydrolysis was inhibited via omission of
magnesium).  The DNA departs significantly from the conformations seen in the
absence of protein, suggesting that binding of the enzyme is required for their
stability.  These are termed neo-conformations to distinguish them from those
which are intrinsically stable in the absence of protein and include the
torsional type-1 neo-kink which unwinds the DNA by approximately 25 degrees.
This separates the DNA backbones by approximately 3.5 angstrom without
unstacking the bases, and is a structural requirement for the recognition
modules of the protein to gain access to the edges of the bases exposed at the
bottom of the major groove.  We suspect that there are a finite number of
structurally feasible neo-conformations which are important for DNA-protein
interactions in general.

<>

<1>McClarin, J.A., Frederick, C.A., Wang, B.C., Greene, P., Boyer, H.W., Grable, J., Rosenberg, J.M.
<2>Structure of the DNA-EcoRI endonuclease recognition complex at 3 angstrom resolution.
<3>Science
<4>234
<5>1526-1541
<6>1986
<7>The crystal structure of the complex between EcoRI endonuclease and the cognate
oligonucleotide TCGC-GAATTCGCG provides a detailed example of the structural basis of
sequence-specific DNA-protein interactions.  The structure was determined, to 3 angstrom
resolution, by the ISIR (iterative single isomorphous replacement) method with a platinum
isomorphous derivative.  The complex has twofold symmetry.  Each subunit of the endonuclease
is organized into an a/b domain consisting of a five-stranded beta sheet, alpha helices, and
an extension, called the "arm," which wraps around the DNA.  The large beta sheet consists of
antiparallel and parallel motifs that form the foundations for loops and alpha helices
responsible for DNA strand scission and sequence-specific recognition, respectively.  The DNA
cleavage site is located in a cleft that binds the DNA backbone in the vicinity of the
scissile bond. Sequence specificity is mediated by 12 hydrogen bonds originating from alpha
helical recognition modules.  Arg200 forms two hydrogen bonds with guanine while Glu144 and
Arg145 form four hydrogen bonds to adjacent adenine residues. These interactions discriminate
the EcoRI hexanucleotide GAATTC from all other hexanucleotides because any base substitution
would require rupture of at least one of these hydrogen bonds.

<>

<1>McClelland, M.
<2>The effect of sequence specific DNA methylation on restriction endonuclease cleavage.
<3>Nucleic Acids Res.
<4>9
<5>5859-5866
<6>1981
<7>Sequence specific DNA methylation sometimes results in the protection of some
or all of a restriction endonucleases' cleavage sites.  This is usually, but
not always, the result of methylation of one or both strands of DNA at the site
characteristic of the corresponding "cognate" modification methylase.  The
known effects of sequence methylation on restriction endonucleases are
compiled.

<>

<1>McClelland, M.
<2>Purification and characterization of two new modification methylases: M.ClaI from Caryophanon latum and M.TaqI from Thermus aquaticus YTI.
<3>Nucleic Acids Res.
<4>9
<5>6795-6804
<6>1981
<7>A method for detecting Type II modification methylases and determining their methylation site
by assaying the ability of methylated DNA to be cleaved by heterologous restriction enzymes is
described and applied to the isolation of the restriction modification methylases from Thermus
thermophilus HB8, Thermus aquaticus YTI and Caryophanon latum L.  M.TaqI is shown to have a
methylation specificity identical to M.TthI (TCGmeA). M.ClaI methylates at adenine and
protects a subset of TthI sites indicating that it methylates the sequence ATCGmeAT.
Methylation by M.TthI also protects against cleavage by SalI, XhoI and at some HindII, AccI
and MboI sites.

<>

<1>McClelland, M.
<2>The effect of site specific methylation on restriction endonuclease cleavage (update).
<3>Nucleic Acids Res.
<4>11
<5>r169-r173
<6>1983
<7>I present here an update of the compilation published in 1981 in this journal
on the effects of site specific methylation on restriction endonuclease
cleavage.  The amount of information available has increased by nearly 50%
since that time.  Table 1 is organized by length of restriction endonuclease
recognition sequence and then alphabetically by sequence.  Isoschizomers are
listed alphabetically by name.  Only the effects of methylation at the N6
position of adenine and C5 of cytosine are considered.  References to the
purification of restriction enzymes and the determination of their recognition
sequences can be found in R.J. Roberts review.

<>

<1>McClelland, M.
<2>The frequency and distribution of methylatable DNA sequences in leguminous plant protein coding genes.
<3>J. Mol. Evol.
<4>19
<5>346-354
<6>1983
<7>Methylation of higher plant DNA occurs at up to 25% of all cytosines, primarily
in the sequences CpG and CpNpG, both of which are over 80% methylated in wheat
and tobacco (Gruenbaum et al. 1981).  CpG and CpNpG frequencies and
distributions in the known sequences of cloned genes of leguminous plants were
analyzed.  In this sample CpG occurred at only 49% of the frequency expected if
the bases were distributed at random.  This lower frequency may be attributed
to the fixation of mutations generated by a high rate of deamination of
5-methylcytosine to thymine (Salser 1977).  Consistent with this hypothesis,
the product of CpG transitions, TpG and CpA, were significantly above their
expected frequency.  However CpNpG occurred at approximately expected levels
and there was no significant increase in its transition products CpNpA and
TpNpG.  Possible explanations for this phenomenon are discussed.  An analysis
of the distribution of di- and trinucleotides across functionally classified
regions of genes showed CpG to be asymmetrically distributed.  CpG was on
average significantly enriched in the 3' flanking regions compared to other
regions.  This may reflect a methylation-mediated regulatory role for this
region in some legume genes.

<>

<1>McClelland, M.
<2>Site-specific cleavage of DNA at 8-, 9-, and 10-bp sequences.
<3>Methods Enzymol.
<4>155
<5>22-33
<6>1987
<7>Site-specific cleavage of DNA at 8-, 9-, and 10-bp sequences had been reported.
This technique relies on a restriction enzyme, DpnI, which only cuts the sequence
GATC when both strands are methylated at adenine 3,4;
    5'...GmA T C...3'
    3'...C TmA G...5'
DpnI does not cut the DNA of most species because they lack this methylated sequence.

<>

<1>McClelland, M.
<2>Selection against dam methylation sites in the genomes of DNA of enterobacteriophages.
<3>J. Mol. Evol.
<4>21
<5>317-322
<6>1984
<7>Post replicative methylation of adenine in Escherichia coli DNA to produce G6mATC (where 6mA
is 6-methyladenine) has been associated with preferential daughter-strand repair and possibly
regulation of replication. An analysis was undertaken to determine if these, or other, as yet
unknown roles of GATC, have had an effect on the frequency of GATC in E. coli or bacteriophage
DNA. It was first ascertained that the most accurate predictions of GATC frequency were based
on the observed frequencies of GAT and ATC, which would be expected since these predictors
take into account preferences in codon usage. The predicted frequencies were compared with
observed GATC frequencies in all available bacterial and phage nucleotide sequences. The
frequency of GATC was close to the predicted frequency in most genes of E. coli and its RNA
bacteriophages and in the genes of nonenteric bacteria and their bacteriophages. However, for
DNA enterobacteriophages the observed frequency of GATC was generally significantly lower than
predicted when assessed by the chi square test. No elevation in the rate of mutation of 6mA in
GATC relative to other bases was found when pairs of DNA sequences from closely related phages
or pairs of homologous genes from enterobacteria were compared, nor was any preferred pathway
for mutation of 6mA evident in the E. coli DNA bacteriophages. This situation contrasts with
that of 5-methylcytosine, which is hypermutable, with a preferred pathway to thymine. Thus,
the low level of GATC in enterobacteriophages is probably due not to 6mA hypermutability, but
to selection against GATC in order to bypass a GATC mediated host function.

<>

<1>McClelland, M.
<2>Recognition sequences of Type II restriction systems are constrained by the G+C content of host genomes.
<3>Nucleic Acids Res.
<4>16
<5>2283-2294
<6>1988
<7>I show that the recognition sequences of Type II restriction systems are
correlated with the G+C content of the host bacterial DNA.  Almost all
restriction systems with G+C rich tetranucleotide recognition sequences are
found in species with A+T rich genomes, whereas G+C rich hexanucleotide and
octanucleotide recognition sequences are found almost exclusively in speciies
with G+C rich genomes.  Most hexanucleotide recognition sequences found in
species with A+T rich genomes are A+T rich.  This distribution eliminates a
substantial proportion of the potential variance in the frequency of
restriction recognition sequences in the host genomes.  As a consequence,
almost all restriction recognition sequences, including those eight base pairs
in length (NotI and SfiI), are predicted to occur with a frequency ranging from
once every 300 to once every 5,000 base pairs in the host genome.  Since the
G+C content of bacteriophage DNA and of the host genome are also correlated,
the data presented is evidence that most TypeII "restriction systems" are
indeed involved in phage restriction.

<>

<1>McClelland, M. et al.
<2>Complete genome sequence of Salmonella enterica serovar typhimurium LT2.
<3>Nature
<4>413
<5>852-856
<6>2001
<7>Salmonella enterica subspecies I, serovar Typhimurium (S. typhimurium), is a leading cause of
human gastroenteritis, and is used as a mouse model of human typhoid fever. The incidence of
non-typhoid salmonellosis is increasing worldwide, causing millions of infections and many
deaths in the human population each year. Here we sequenced the 4,857-kilobase (kb) chromosome
and 94-kb virulence plasmid of S. typhimurium strain LT2. The distribution of close homologues
of S. typhimurium LT2 genes in eight related enterobacteria was determined using previously
completed genomes of three related bacteria, sample sequencing of both S. enterica serovar
Paratyphi A (S. paratyphi A) and Klebsiella pneumoniae, and hybridization of three unsequenced
genomes to a microarray of S. typhimurium LT2 genes. Lateral transfer of genes is frequent,
with 11% of the S. typhimurium LT2 genes missing from S. enterica serovar Typhi (S. typhi),
and 29% missing from Escherichia coli K12. The 352 gene homologues of S. typhimurium LT2
confined to subspecies I of S. enterica-containing most mammalian and bird pathogens-are
useful for studies of epidemiology, host specificity and pathogenesis. Most of these
homologues were previously unknown, and 50 may be exported to the periplasm or outer membrane,
rendering them accessible as therapeutic or vaccine targets.

<>

<1>McClelland, M. et al.
<2>Comparison of genome degradation in Paratyphi A and Typhi, human-restricted serovars of Salmonella enterica that cause typhoid.
<3>Nat. Genet.
<4>36
<5>1268-1274
<6>2004
<7>Salmonella enterica serovars often have a broad host range, and some cause both
gastrointestinal and systemic disease. But the serovars Paratyphi A
and Typhi are restricted to humans and cause only systemic disease. It has
been estimated that Typhi arose in the last few thousand years. The
sequence and microarray analysis of the Paratyphi A genome indicates that
it is similar to the Typhi genome but suggests that it has a more recent
evolutionary origin. Both genomes have independently accumulated many
pseudogenes among their approximately 4,400 protein coding sequences: 173
in Paratyphi A and approximately 210 in Typhi. The recent convergence of
these two similar genomes on a similar phenotype is subtly reflected in
their genotypes: only 30 genes are degraded in both serovars.
Nevertheless, these 30 genes include three known to be important in
gastroenteritis, which does not occur in these serovars, and four for
Salmonella-translocated effectors, which are normally secreted into host
cells to subvert host functions. Loss of function also occurs by mutation
in different genes in the same pathway (e.g., in chemotaxis and in the
production of fimbriae).

<>

<1>McClelland, M., Bhagwat, A.S.
<2>Biased DNA repair.
<3>Nature
<4>355
<5>595-596
<6>1992
<7>Hennecke et al. report the first in vitro characterization of a DNA mismatch
endonuclease.  This enzyme, the vsr gene product, initiates very short patch
repair of E. coli by nicking one strand of the duplex next to the T at a
mismatched T:G in the sequence CTWG or TWGG.

<>

<1>McClelland, M., Hanish, J., Nelson, M., Patel, Y.
<2>KGB: a single buffer for all restriction endonucleases.
<3>Nucleic Acids Res.
<4>16
<5>364
<6>1988
<7>Most recommended restriction buffers contain Na+ and Cl-.  However, in bacteria
the most abundant intracellular cation and anion are usually potassium and
glutamate, respectively.  Furthermore, restriction endonucleases cleave DNA in
potassium glutamate (KGlu) over a much broader concentration range than they do
in NaCl.  These facts encouraged us to investigate the possibility that we
could use KGlu in a chloride-free buffer and achieve normal levels of activity
for all restriction endonucleases.  We have tested fifty-five restriction
endonucleases for their ability to cleave DNA in a series of KGlu buffers (KGB,
see Table 1) and compared the level of activity with that found under
conditions recommended by the vendors (New England Biolabs, Boehringer Mannheim
Biochem. and International Biotech. Inc.).  Assays were performed as partial
digests (0.2 units per ug of DNA in 30 ul for 30 min.) and as overnight
digestions with excess enzyme to ensure that no loss of specificity (start
activity) occurred.  Most restriction endonucleases, polymerases and ligase
showed broad KGlu concentration optima and all enzymes functioned in 100 mM
KGlu (1X KGB).  Reducing agent was not normally required.  Some enzymes worked
well in concentrations of KGlu over 400 mM (data not presented).  KGB can be
used to simplify laboratory procedures including double digests, DNA cleavage
followed by end-labeling, or the digestion of DNA embedded in agarose prior to
pulsed field gel electrophoresis.  DNA in KGB can be phenol extracted and
ethanol precipitated using standard protocols.

<>

<1>McClelland, M., Jones, R., Patel, Y., Nelson, M.
<2>Restriction endonucleases for pulsed field mapping of bacterial genomes.
<3>Nucleic Acids Res.
<4>15
<5>5985-6005
<6>1987
<7>Fundamental to many bacterial genome mapping strategies currently under
development is the need to cleave the genome into a few large DNA fragments
that can be resolved by pulsed field gel electrophoresis.  Identification of
endonucleases that infrequently cut a genome is of key importance in this
process.  We show that the tetranucleotide CTAG is extremely rare in most
bacterial genomes with G+C contents above 45%.  As a consequence, most of the
sixteen bacterial genomes we have tested are cleaved less than once every
100,000 base pairs by one or more endonucleases that have CTAG in their
recognition sequences:  XbaI (TCTAGA), SpeI (ACTAGT), AvrII (CCTAGG) and NheI
(GCTAGC).  Similarily, CCG and CGG are the rarest trinucleotides in many
genomes with G+C content of less than 45%.  Thus, SmaI (CCCGGG), RsrII
(CGGWCCG), NaeI (GCCGGC) and SacII (CCGCGG) are often suitable endonucleases
for producing fragments that average over 100,000 base pairs from such genomes.
Pulsed field gel electrophoresis of the fragments that result from cleavage
witih endonucleases that cleave only a few times per genome should assist in
the physical mapping of many prokaryotic genomes.

<>

<1>McClelland, M., Kessler, L.G.
<2>Site specific cleavage of DNA - by creating DNA specific methylation dependent restriction endonuclease recognition sequence.
<3>US Patent Office
<4>US 4808525
<5>
<6>1989
<7>The following site specific clearage of a double-stranded (d-s) DNA molecule (I) is claimed.
(I) which has at least two methylase recognition sequences, is contacted with at least one
sequence specific methylase to produce a methylated d-s molecule with a newly created DNA
sepcific methylation dependent restriction endonulcease recognition equence. The latter partly
overlaps with at least two of the methylase recognition sequences and the overlapping
sequences have a combined length of not less than 8 bp. The prod. is then contacted with a
methylation dependent DNA specific restriction endonuclease. In an embodiment . (I) has at
least one methylase recognition sequence. Also claimed is a DNA vector with a d-s DNA sequence
of not less than 8 bp long. N = A, C, G or T. 16 Other sequences are defined in the claims.

<>

<1>McClelland, M., Kessler, L.G., Bittner, M.
<2>Site-specific cleavage of DNA at 8- and 10-base pair sequences.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>81
<5>983-987
<6>1984
<7>A method is described for cutting DNA at specific sites that are 8 and 10 base
pairs long.  The DNA is first treated with a specific methylase, either the
restriction-modification enzyme M. TaqI, which converts the 4-base sequence
T-C-G-A to T-C-G-mA, or the similar enzyme M. ClaI, which converts the 6-base
sequence A-T-C-G-A-T to A-T-C-G-mA-T.  The DNA is then cleaved with DpnI, a
restriction endonuclease that recognizes the sequence G-mA-T-C.  DpnI is unique
in that it cuts only DNA that is methylated at adenine in both strands of its
recognition sequence.  In DNAs that are not otherwise methylated at adenine in
both strands of the sequence G-A-T-C, cleavage by DpnI occurs only at the
following sequences: 	in the case of M. TaqI methylation, 5' T-C-G-mA- T-C-G-mA
3'3'mA-G-C- T-mA-G-C- T 5'; in the case of M. ClaI methylation, 5' A- T-C-G-mA-
T-C-G-mA-T 3'3' T-mA-G-C -T-mA-G-C -T-A 5'.  Specific cutting and cloning at
these methylase/DpnI-generated sites is shown experimentally.  Further, we
describe how the above technique can be extended to generate DpnI cleavage
sites of up to 12 base pairs.  In DNA that contains equal amounts of each base
distributed at random, 8- and 10-base pair recognition sequences occur, on the
average, approximately once every 65,000 and 1,000,000 base pairs,
respectively.  Potential applications, including the development of cloning
vectors and a rapid method for chromosome walking, are discussed.

<>

<1>McClelland, M., Nelson, M.
<2>Enhancement of the apparent cleavage specificities of restriction endonucleases:  applications to megabase mapping of chromosomes.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>257-282
<6>1987
<7>Restriction endonucleases generally recognize DNA sequences of four to six base
pairs.  Therefore, these enzymes cut DNA that contains equal amounts of A,C,G
and T, distributed at random, into fragments with an average size of 4/4 to 4/6
base pairs, (256 to 4096 base pairs).  Accordingly, restriction endonuclease
mapping techniques are appropriate for the analysis of gene-sized DNA
molecules.  For example, a restriction endonuclease with a six base specificity
will cut the human genome into approximately 750,000 fragments averaging 4
kilobase pairs.  However, more specific DNA cutting methods would yield fewer
and larger fragments, suitable for the analysis of large, complex genomes.
Techniques have been described for the preparation and separation of DNA
molecules of one megabase pair or more.  To produce fragments of this size from
random DNA one must have a recognition sequence which is the log (base 4) of
1,000,000; approximately ten base pairs.  The genome sizes of various organisms
and the number of fragments generated by cleavage systems of varying
specificities are presented in Table 1.  We describe here a number of
strategies which can be used to increase the apparent specificity of
restriction endonucleases to produce Average Fragment Sizes (AFS) in the range
fo 6,000 to 270,000,000 base pairs.  These are:  (1) protection of restriction
endonuclease recognition sites from cleavage by sequence-specific DNA
methylation. (2) generation of cleavage sites using sequence-specific DNA
methylation and a methylation-dependent restriction endonuclease, DpnI.  (3)
blocking DNA methylases by other methylases.  (4) prediction of rare
restriction recognition sites for particular genomes based on the non-random
sequence arrangement of natural DNAs.  We believe that combinations of the
above-mentioned three methods will provide a framework for the eventual
molecular dissection of large, complex, DNA molecules.

<>

<1>McClelland, M., Nelson, M.
<2>The effect of site-specific DNA methylation on restriction endonucleases and DNA modification methyltransferases - a review.
<3>Gene
<4>74
<5>291-304
<6>1988
<7>Review of methylation sensitivity.

<>

<1>McClelland, M., Nelson, M.
<2>The 5'-GGATCC-3' cleavage specificity of BamHI is increased to 5'-CCGGATCCGG-3' by sequential double methylation with M.HpaII and M.BamHI.
<3>Gene
<4>74
<5>169-176
<6>1988
<7>Site-specific DNA methylation is known to block cleavage by a number of
restriction endonucleases.  We show that methylation at non-canonical DNA
modification sites can also block methylation by five of 13 DNA
methyltransferases (MTases) tested.  Furthermore, MTases and endonucleases that
recognize the same nucleotide sequence can differ in their sensitivity to
non-canonical methylation.  In particular, BamHI endonuclease can cut
5'-GGATCm5C efficiently, whereas M.BamHI cannot methylate this modified
sequence.  Methyltransferase/endonuclease pairs which differ in their
sensitivity to non-canonical methylation can be exploited to generate rare DNA
cleavage sites.  For example, we show that M.HpaII, M.BamHI, and BamHI can be
used sequentially in a three-step procedure to specifically cleave DNA at the
10-bp sequence 5'-CCGGATCCGG.  Several highly selective DNA cutting strategies
are made possible by these sequential double methylation-blocking reactions.

<>

<1>McClelland, M., Nelson, M.
<2>The effect of site specific methylation on restriction endonuclease digestion.
<3>Nucleic Acids Res.
<4>13
<5>r201-r207
<6>1985
<7>None

<>

<1>McClelland, M., Nelson, M.
<2>Effect of site-specific methylation on DNA modification methyltransferases and restriction endonucleases.
<3>Nucleic Acids Res.
<4>20
<5>2145-2157
<6>1992
<7>We present in Table I an updated list of the sensitivities of over 280 restriction
endonucleases to the site-specific DNA modifications m4C, m5C, hm5C, and m6A, four
modifications that are common in DNA prokaryotes, eukaryotes, and their viruses. Table II is a
list of over 190 characterized DNA methyltransferases. A detailed list of cloned
restriction-modification genes has been made by Wilson. Table III lists the sensitivities of
over 20 Type II DNA methyltransferases to m4C, m5C, hm5C, and m6A modification. Most DNA
methyltransferases are sensitive to non-canonical modifications within their recognition
sequences, and this sensitivity may differ from that of their restriction endonuclease
partners. Finally, several restriction endonuclease isoschizomers are known to differ in their
ability to cleave DNA which has been methylated. Table IV lists over 20 known isoschizomer
pairs and one isomethylator pair, along with the modified recognition sites at which they
differ.

<>

<1>McClelland, M., Nelson, M., Cantor, C.R.
<2>Purification of MboII methylase (GAAGmA) from Moraxella bovis: site specific cleavage of DNA at nine and ten base pair sequences.
<3>Nucleic Acids Res.
<4>13
<5>7171-7182
<6>1985
<7>The restriction modification methylase M.MboII has been purified using a sensitive
oligonucleotide linker assay.  The enzyme methylates the MboII recognition sequence GAAGA at
adenine to produce GAAGmA.  M.MboII can be used in conjunction with the methylation dependent
restriction endonuclease DpnI (GmATC) to produce cleavage at the 10 base sequence GAAGATCTTC.
When M.MboII is used in combination with M.ClaI (ATCGATCGAT), GAAGATCTTC, GAAGATCGAT,
ATCGATCTTC and ATCGATCGAT, which is equivalent to a nine base recognition site. The use of
combinations of adenine methylases and DpnI to generate highly selective DNA cleavages at a
variety of sequences up to fourteen base pairs is discussed.

<>

<1>McClelland, M., Nelson, M., Raschke, E.
<2>Effect of site-specific modification on restriction endonucleases and DNA modification methyltransferases.
<3>Nucleic Acids Res.
<4>22
<5>3640-3659
<6>1994
<7>Restriction endonucleases have site-specific interactions with DNA that can often be inhibited
by site-specific DNA methylation and other site-specific DNA modifications. However, such
inhibition cannot generally be predicted. The empirically acquired data on these effects are
tabulated for over 320 restriction endonucleases. In addition, a table of known site-specific
DNA modification methyltransferases and their specificities is presented along with EMBL
database accession numbers for cloned genes.

<>

<1>McClelland, S.E., Dryden, D.T.F., Szczelkun, M.D.
<2>Continuous assays for DNA translocation using fluorescent triplex dissociation: Application to type I restriction endonucleases.
<3>J. Mol. Biol.
<4>348
<5>895-915
<6>2005
<7>Fluorescent assays and accompanying kinetic models are described for the analysis of DNA
translocation independent of duplex unwinding. A
triplex binding site (TBS) was introduced into DNA substrates at
precise loci downstream of recognition sequences for type IA, IB and IC
restriction endonucleases (EcoKI, EcoAI and EcoR1241, respectively).
Each endonuclease was incubated (without ATP) with substrates on which
a hexachlorofluoroscein-labelled triplex-forming oligonucleotide
(HEX-TFO) was pre-bound. Following addition of ATP, 1-D enzyme motion
resulted in collision with, and displacement of, the HEX-TFO, producing
a > twofold increase in fluorescent intensity. Alternatively, a
decrease in anisotropy following displacement of a rhodamine-labelled
TFO was monitored. Using rapid mixing in a stopped-flow fluorimeter,
continuous kinetic profiles were produced in which displacement is
preceded by a lag-phase, directly proportional to the distance moved.
For each enzyme, we obtained not only the translocation. rate but also
information on slow isomerisation step(s) at initiation. Furthermore,
we demonstrated that enzymes deficient in DNA cleavage but with maximal
ATPase activity showed initiation and translocation rates identical to
wild-type, confirming that DNA strand breaks are not a pre-requisite of
motion.

<>

<1>McClelland, S.E., Szczelkun, M.D.
<2>The type I and III restriction endonucleases: Structural elements in the molecular motors that process DNA.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>111-135
<6>2004
<7>The Type I and III restriction endonucleases are larger, multimeric protein complexes with
four enzyme activities; DNA methyltransferase, DNA endonuclease, ATPase and DNA translocase.
It has been demonstrated that ATP-dependent protein motion along DNA is necessary for
endonuclease activity.  Studies have shown that Type I enzymes remain bound to their
recognition sites whilst simultaneously translocating adjacent non-specific dsDNA past a
stationary complex.  This occurs bi-directionally so that two DNA loops are extruded.  An
equivalent unidirectional mechanism has been suggested for the Type III enzymes.  DNA cleavage
generally results when the enzymes stall against another restriction enzyme complex.  Both the
HsdR subunits of the Type I enzymes and the Res subunits of the Type III enzymes carry amino
acid motifs characteristic of superfamily 2 helicases.  In this review, the structural and
mechanistic implications of this relationship are discussed and models s!
uggested for how the ATP-dependent restriction enzymes might couple chemical energy to
mechanical motion on DNA.

<>

<1>McConnell, D.J., Searcy, D.G., Sutcliffe, J.G.
<2>A restriction enzyme ThaI from the thermophilic mycoplasma Thermoplasma acidophilum.
<3>Nucleic Acids Res.
<4>5
<5>1729-1739
<6>1978
<7>A type II restriction enzyme (ThaI) has been isolated from the thermophilic
mycoplasma Thermoplasma acidophilum.  A new method of general application was
used to determine the DNA sequence cleaved by the enzyme.  ThaI cuts DNA in the
centre of the sequence CGCG.  Single-stranded DNA is not a substrate.  ThaI
does not cut T. acidophilum DNA which is presumably modified.  This is the
first description of a restriction enzyme from a mycoplasma.  Because ThaI is
easily prepared in large amounts of approximately 105 units per gram of cells,
it will be a valuable addition to the battery of restriction enzymes used in
studies of DNA sequences.  It is active at high temperatures and may therefore
be useful for special purposes requiring more extreme conditions.

<>

<1>McConnell, S.A., Takeuchi, R., Pellenz, S., Davis, L., Maizels, N., Monnat, R.J. Jr., Stoddard, B.L.
<2>Generation of a nicking enzyme that stimulates site-specific gene conversion from the I-AniI LAGLIDADG homing endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>5099-5104
<6>2009
<7>Homing endonucleases stimulate gene conversion by generating site-specific DNA double-strand
breaks that are repaired by homologous recombination.
These enzymes are potentially valuable tools for targeted gene correction
and genome engineering. We have engineered a variant of the I-AniI homing
endonuclease that nicks its cognate target site. This variant contains a
mutation of a basic residue essential for proton transfer and solvent
activation in one active site. The cleavage mechanism, DNA-binding
affinity, and substrate specificity profile of the nickase are similar to
the wild-type enzyme. I-AniI nickase stimulates targeted gene correction
in human cells, in cis and in trans, at approximately 1/4 the efficiency
of the wild-type enzyme. The development of sequence-specific nicking
enzymes like the I-AniI nickase will facilitate comparative analyses of
DNA repair and mutagenesis induced by single- or double-strand breaks.

<>

<1>McConnell-Smith, A., Stoddard, B.L.
<2>Generation of a DNA nicking enzyme that stimulates site-specific gene conversion from a homing endonuclease.
<3>International Patent Office
<4>WO 201093966 A
<5>
<6>2010
<7>An engineered highly specific DNA-cleavage enzyme delivers a site-specific nick in a double
stranded DNA, to cleave one DNA strand within its target site while leaving the opposing DNA
strand intact.  The engineered enzyme provides the ability to induce a gene conversion event
in a mammalian cell.  An engineered sequence-specific nickase derived from a LAGLIDADG homing
endonuclease is altered by a single amino acid residue, wherein the amino acid residue is
involved in the polarization of solvent molecules and acid-base catalysis in the active site
without affecting direct contacts between the enzyme and either the bound DNA or bound metal
ions.  Engineered, site-specific nickase variants, such as of I-AniI and other homing
endonucleases, are particularly useful in targeted genome engineering as well as therapeutic,
targeted gene repair.

<>

<1>McCulloch, J.A., de Oliveira, V.M., de Almeida, P.A.V., Perez-Chaparro, P.J., de Almeida, L.M., de Vasconcelos, J.M., de Oliveira, L.F., da Silva, D.E., Rogez, H.L., Cretenet, M., Mamizuka, E.M., Nunes, M.R.
<2>Complete Genome Sequence of Lactococcus lactis Strain AI06, an Endophyte of the Amazonian Acai Palm.
<3>Genome Announcements
<4>2
<5>e01225-14
<6>2014
<7>We report the genome, in a single chromosome, of Lactococcus lactis strain AI06,  isolated
from the mesocarp of the acai fruit (Euterpe oleracea) in eastern
Amazonia, Brazil. This strain is an endophyte of the acai palm and also a
component of the microbiota of the edible food product.

<>

<1>McCusker, M.P., Hokamp, K., Buckley, J.F., Wall, P.G., Martins, M., Fanning, S.
<2>Complete Genome Sequence of Salmonella enterica Serovar Agona Pulsed-Field Type SAGOXB.0066, Cause of a 2008 Pan-European Outbreak.
<3>Genome Announcements
<4>2
<5>e01219-13
<6>2014
<7>Salmonella enterica serovar Agona is in the top 10 most common nontyphoidal serovars reported
in humans in the European Union. Here we report the complete
genome sequence of an S. enterica serovar Agona isolate, designated 24249, that
was the cause of a pan-European outbreak in 2008 with 163 confirmed cases
reported.

<>

<1>McDermott, P.F., Crawford, J.T.
<2>A plasmid-encoded restriction-modification system in Mycobacterium avium.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>89
<5>167
<6>1989
<7>We previously described an R-M system in M. avium strain LR25.  This system was
demonstrated by restriction and host-induced modification of phage JF2.  Curing
of the plasmids of LR25 produced a strain, LR163, that lacks the R-M system.
We have now demonstrated the presence of a restriction endonuclease, designated
MavI, in extracts of LR25 and other M. avium strains.  Cells were grown in
7H9-Av broth and treated with cycloserine for 16 hr.  Cells were harvested and
then lysed by sonication.  Debris was removed by centrifugation, and the DNA
was precipitated with streptomycin sulfate.  The crude lysates were assayed for
endonuclease activity using various test DNAs.  The digest patterns obtained
with the extract of LR25 were the same as those produced by XhoI.  Further
studies showed that MavI cleaves at the same site as XhoI, C^TCGAG.  The enzyme
was demonstrated in several plasmid-containing strains but was absent from
LR163.  DNA isolated from the strains having MavI activity ws found to be
resistant to cleavage by XhoI but sensitive to other endonucleases indicating
the expected modification.  Modification of some SalI sites was also observed,
but no corresponding endonuclease activity was detected.

<>

<1>McDonald, R.R., Golding, G.R., Irvine, J., Graham, M.R., Tyler, S., Mulvey, M.R., Levett, P.N.
<2>Draft Genome Sequence of Methicillin-Susceptible Staphylococcus aureus Strain 06BA18369, a Pathogen Associated with Skin and Soft Tissue Infections in Northern  Saskatchewan, Canada.
<3>Genome Announcements
<4>1
<5>e00389-13
<6>2013
<7>Here, we announce the draft sequence of a representative methicillin-susceptible
Staphylococcus aureus (MSSA) isolate (06BA18369) whose strain type (spa type
t311) was commonly isolated from skin and soft tissue coinfections with
Streptococcus pyogenes. This strain sequence provides insight into a highly
successful community-associated MSSA strain type.

<>

<1>McDonald, R.R., Golding, G.R., Irvine, J., Graham, M.R., Tyler, S., Mulvey, M.R., Levett, P.N.
<2>Draft Genome Sequence of Streptococcus pyogenes Strain 06BA18369, a Human Pathogen Associated with Skin and Soft Tissue Infections in Northern Canada.
<3>Genome Announcements
<4>1
<5>e00387-13
<6>2013
<7>We report the draft sequence of Streptococcus pyogenes 06BA18369 (emm type 41.2,  sequence
type 579 [ST579]), isolated from a skin and soft tissue infection (SSTI)
mixed with Staphylococcus aureus. This genome provides insight into the genetic
composition of S. pyogenes strains associated with mixed SSTIs.

<>

<1>McDowell, A., Hunyadkurti, J., Horvath, B., Voros, A., Barnard, E., Patrick, S., Nagy, I.
<2>Draft Genome Sequence of an Antibiotic-Resistant Propionibacterium acnes Strain,  PRP-38, from the Novel Type IC Cluster.
<3>J. Bacteriol.
<4>194
<5>3260-3261
<6>2012
<7>Propionibacterium acnes, a non-spore-forming, anaerobic Gram-positive bacterium,  is most
notably recognized for its association with acne vulgaris (I. Kurokawa et
al., Exp. Dermatol. 18:821-832, 2009). We now present the draft genome sequence
of an antibiotic-resistant P. acnes strain, PRP-38, isolated from an acne patient
in the United Kingdom and belonging to the novel type IC cluster.

<>

<1>Mcdowell, P., Foster, S., Clark, S., Bramel, C.
<2>Antigenic Polypeptides.
<3>Japanese Patent Office
<4>JP 2004500883 A
<5>
<6>2004
<7>
<>

<1>McGann, P., Hang, J., Clifford, R.J., Yang, Y., Kwak, Y.I., Kuschner, R.A., Lesho, E.P., Waterman, P.E.
<2>Complete Sequence of a Novel 178-Kilobase Plasmid Carrying blaNDM-1 in a Providencia stuartii Strain Isolated in Afghanistan.
<3>Antimicrob. Agents Chemother.
<4>56
<5>1673-1679
<6>2012
<7>In response to global concerns over the spread of the New Delhi metallo-beta-lactamase gene 1,
bla(NDM-1), a monthly surveillance program was initiated in September 2010. All
carbapenem-resistant Gram-negative strains forwarded to our facility are screened for this
gene. To date, 321 carbapenem-resistant isolates, encompassing 11 bacterial species, have been
tested. In February 2011, two strains of Providencia stuartii, submitted from a military
hospital in Afghanistan, tested positive for bla(NDM-1). Both strains were identical by
pulsed-field gel electrophoresis (PFGE). bla(NDM-1) was carried on a large plasmid, pMR0211,
which was sequenced by emulsion PCR and pyrosequencing. pMR0211 is 178,277 bp in size and
belongs to incompatibility group A/C. The plasmid consists of a backbone with considerable
homology to pAR060302 from Escherichia coli, and it retains many of the antibiotic resistance
genes associated with it. The plasmid also shares common elements with the pNDM-HK plasmid,
including bla(NDM-1), armA, and sul1. However, gene orientation is reversed, and a 3-kb
fragment from this region is absent from pMR0211. pMR0211 also contains additional genes,
including the aminoglycoside-modifying enzyme loci aadA and aac(6'), the quinolone resistance
gene qnrA, a gene with highest homology to a U32 family peptidase from Shewanella amazonensis,
and the bla(OXA-10) gene. The finding of this gene in an intrinsically colistin-resistant
species such as Providencia stuartii is especially worrisome, as it renders the organism
resistant to nearly every available antibiotic. The presence of multiple insertion sequences
and transposons flanking the region containing the bla(NDM-1) gene further highlights the
potential mobility associated with this gene.

<>

<1>McGeehan, J.E., Ball, N.J., Streeter, S.D., Thresh, S.J., Kneale, G.G.
<2>Recognition of dual symmetry by the controller protein C.Esp1396I based on the structure of the transcriptional activation complex.
<3>Nucleic Acids Res.
<4>40
<5>4158-4167
<6>2012
<7>The controller protein C.Esp1396I regulates the timing of gene expression of the
restriction-modification (RM) genes of the RM system Esp1396I. The molecular
recognition of promoter sequences by such transcriptional regulators is poorly
understood, in part because the DNA sequence motifs do not conform to a
well-defined symmetry. We report here the crystal structure of the controller
protein bound to a DNA operator site. The structure reveals how two different
symmetries within the operator are simultaneously recognized by the homo-dimeric
protein, underpinned by a conformational change in one of the protein subunits.
The recognition of two different DNA symmetries through movement of a flexible
loop in one of the protein subunits may represent a general mechanism for the
recognition of pseudo-symmetric DNA sequences.

<>

<1>McGeehan, J.E., Papapanagiotou, I., Streeter, S.D., Kneale, G.G.
<2>Cooperative Binding of the C.AhdI Controller Protein to the C/R Promoter and its Role in Endonuclease Gene Expression.
<3>J. Mol. Biol.
<4>358
<5>523-531
<6>2006
<7>The controller (C) proteins of a wide variety of restriction-modification (R-M) systems are
thought to regulate expression of the endonuclease (R) gene by a genetic switch that ensures
that methylation precedes endonuclease expression. Previous DNA footprinting experiments with
C.AhdI have located the binding site upstream of the C and R genes in the AhdI R-M system, and
the structure of C.AhdI has recently been determined. Here, we provide evidence that the
binding site can accommodate either one or two dimers of C.AhdI in a concentration-dependent
manner The dimer binding site is adjacent to the -35 hexamer site required for the interaction
with RNA polymerase (RNAP); however, co-operative binding of a second dimer blocks this site.
Optimum DNA binding site sizes for dimer and tetramer formation were determined to be ca 21 bp
and 34 bp, respectively. The stoichiometry and affinities of relevant DNA-protein complexes
have been characterised by sedimentation velocity and EMSA using native and mutant promoter
sequences. Molecular models of the dimer and tetramer complexes have been constructed that are
consistent with the hydrodynamic data. Our results suggest a mechanism for both positive and
negative regulation of endonuclease expression, whereby at moderate levels of C.AhdI, the
protein binds to the promoter as a dimer and stimulates transcription by the interaction with
RNAP. As the levels of C.AhdI increase further, binding of the second dimer competes with
RNAP, thus down-regulating transcription of its own gene, and hence that of the endonuclease.

<>

<1>McGeehan, J.E., Streeter, S., Cooper, J.B., Mohammed, F., Fox, G.C., Kneale, G.G.
<2>Crystallization and preliminary X-ray analysis of the controller protein C.Ahdl from Aeromonas hydrophilia.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>60
<5>323-325
<6>2004
<7>Single crystals of purified homodimeric controller protein from Aeromonas hydrophilia (C.AhdI)
have been grown under several different
conditions using vapour diffusion. X-ray diffraction data have been
collected using synchrotron radiation from crystals of both the native
and a selenomethionine (SeMet) derivative of the protein. The native
crystal form belongs to space group P2(1) and data were collected to a
resolution of 2.2 Angstrom. Two crystal forms of the SeMet protein have
been obtained and were found to belong to space groups P1 and P2(1);
data have been recorded to 2.0 and 1.7 Angstrom resolution,
respectively, for the two crystal forms. Three-wavelength MAD data were
collected to 1.7 Angstrom for the SeMet derivative crystal, which is
isomorphous with the native P2(1) crystal.

<>

<1>McGeehan, J.E., Streeter, S., Papapanagiotou, I., Fox, G.C., Kneale, G.G.
<2>High-resolution crystal structure of the restriction-modification controller protein C.AhdI from Aeromonas hydrophila.
<3>J. Mol. Biol.
<4>346
<5>689-701
<6>2005
<7>Restriction/modification (R/M) systems serve to protect the host bacterium from invading
bacteriophage. The multi-component system includes a methyltransferase, which recognizes and
methylates a specific DNA sequence, and an endonuclease which recognises the same sequence and
cleaves within or close to this site. The endonuclease will only cleave DNA that is
unmethylated at the specific site, thus host DNA is protected while non-host DNA is cleaved.
However, following DNA replication, expression of the endonuclease must be delayed until the
host DNA is appropriately methylated. In many R/M systems, this regulation is achieved at the
transcriptional level via the controller protein, or C-protein.  We have solved the first
X-ray structure of an R/M controller protein, C.AhdI, to 1.69  resolution using
selenomethionine MAD. C.AhdI is part of a Type IIH R/M system from the pathogen Aeromonas
hydrophila. The structure reveals an all-B protein that contains a classical helix-turn-helix
(HTH) domain and can be assigned to the Xre family of transcriptional regulators. Unlike its
monomeric structural homologues, an extended helix generates an interface that results in
dimerisation of the free protein. The dimer is electrostatically polarised and a positively
charged surface corresponds to the position of the DNA recognition helices of the HTH domain.
Comparison with the structure of the cI ternary complex suggests that C.AhdI activates
transcription through direct contact with the 70 subunit of RNA polymerase.

<>

<1>McGeehan, J.E., Streeter, S.D., Thresh, S.J., Ball, N., Ravelli, R.B., Kneale, G.G.
<2>Structural analysis of the genetic switch that regulates the expression of restriction-modification genes.
<3>Nucleic Acids Res.
<4>36
<5>4778-4787
<6>2008
<7>Controller (C) proteins regulate the timing of the expression of restriction and modification
(R-M) genes through a combination of positive and negative feedback circuits. A single dimer
bound to the operator switches on transcription of the C-gene and the endonuclease gene; at
higher concentrations, a second dimer bound adjacently switches off these genes. Here we
report the first structure of a C protein-DNA operator complex, consisting of two C protein
dimers bound to the native 35 bp operator sequence of the R-M system Esp1396I. The structure
reveals a role for both direct and indirect DNA sequence recognition. The structure of the DNA
in the complex is highly distorted, with severe compression of the minor groove resulting in a
50 degrees bend within each operator site, together with a large expansion of the major groove
in the centre of the DNA sequence. Cooperative binding between dimers governs the
concentration-dependent activation-repression switch and arises, in part, from the interaction
of Glu25 and Arg35 side chains at the dimer-dimer interface. Competition between Arg35 and an
equivalent residue of the sigma(70) subunit of RNA polymerase for the Glu25 site underpins the
switch from activation to repression of the endonuclease gene.

<>

<1>McGeehan, J.E., Streeter, S.D., Thresh, S.J., Taylor, J.E., Shevtsov, M.B., Kneale, G.G.
<2>Structural Analysis of a Novel Class of R-M Controller Proteins: C.Csp231I from Citrobacter sp. RFL231.
<3>J. Mol. Biol.
<4>409
<5>177-188
<6>2011
<7>Controller proteins play a key role in the temporal regulation of gene expression in bacterial
restriction-modification (R-M) systems and are
important mediators of horizontal gene transfer. They form the basis of a
highly cooperative, concentration-dependent genetic switch involved in
both activation and repression of R-M genes. Here we present biophysical,
biochemical, and high-resolution structural analysis of a novel class of
controller proteins, exemplified by C.Csp231I. In contrast to all
previously solved C-protein structures, each protein subunit has two extra
helices at the C-terminus, which play a large part in maintaining the
dimer interface. The DNA binding site of the protein is also novel, having
largely AAAA tracts between the palindromic recognition half-sites,
suggesting tight bending of the DNA. The protein structure shows an
unusual positively charged surface that could form the basis for wrapping
the DNA completely around the C-protein dimer.

<>

<1>McGillivary, G., Tomaras, A.P., Rhodes, E.R., Actis, L.A.
<2>Cloning and sequencing of a genomic island found in the Brazilian purpuric fever clone of Haemophilus influenzae biogroup aegyptius.
<3>Infect. Immun.
<4>73
<5>1927-1938
<6>2005
<7>A genomic island was identified in the Haemophilus influenzae biogroup
aegyptius Brazilian purpuric fever (BPF) strain F3031. This island, which
was also found in other BPF isolates, could not be detected in non-BPF
biogroup aegyptius strains or in nontypeable or typeable H. influenzae
strains, with the exception of a region present in the type b Eagan
strain. This 34,378-bp island is inserted, in reference to H. influenzae
Rd KW20, within a choline transport gene and contains a mosaic structure
of Mu-like prophage genes, several hypothetical genes, and genes
potentially encoding an Erwinia carotovora carotovoricin Er-like
bacteriocin. The product of the tail fiber ORF in the bacteriocin-like
region shows a hybrid structure where the C terminus is similar to an H.
influenzae phage HP1 tail protein implicating this open reading frame in
altering host specificity for a putative bacteriocin. Significant synteny
is seen in the entire genomic island with genomic regions from Salmonella
enterica subsp. enterica serovar Typhi CT18, Photorhabdus luminescens
subsp. laumondii TT01, Chromobacterium violaceum, and to a lesser extent
Haemophilus ducreyi 35000HP. In a previous work, we isolated several
BPF-specific DNA fragments through a genome subtraction procedure, and we
have found that a majority of these fragments map to this locus. In
addition, several subtracted fragments generated from an independent
laboratory by using different but related strains also map to this island.
These findings underscore the importance of this BPF-specific chromosomal
region in explaining some of the genomic differences between highly
invasive BPF strains and non-BPF isolates of biogroup aegyptius.

<>

<1>McGrath, K.C., Thomas-Hall, S.R., Cheng, C.T., Leo, L., Alexa, A., Schmidt, S., Schenk, P.M.
<2>Isolation and analysis of mRNA from environmental microbial communities.
<3>J. Microbiol. Methods
<4>75
<5>172-176
<6>2008
<7>The advent of metagenomics has revealed that our planet harbors millions
of previously undiscovered microbial species. However, functional insights
into the activities of microbial communities cannot easily be obtained
using metagenomics. Using transcriptional analyses to study microbial gene
functions is currently problematic due to difficulties working with
unstable microbial mRNA as a small fraction of total cellular RNA. Current
techniques can be expensive and time consuming, and still result in
significant levels of rRNA contamination. We have adapted techniques to
rapidly isolate high high-quality RNA from environmental samples and
developed a simple method for specific isolation of mRNA by size
separation. This new technique was evaluated by constructing cDNA
libraries directly from uncultured environmental microbial communities,
including agricultural soil samples, aquatic flocculants, organic
composts, mammalian oral and faecal samples, and wastewater sludge. The
sequencing of a fraction of these cDNA clones revealed a high degree of
novelty, demonstrating the potential of this approach to capture a large
number of unique transcripts directly from the environment. To our
knowledge, this is the first study that uses gel electrophoresis to
isolate mRNA from microbial communities. We conclude that this method
could be used to provide insights into the microbial 'metatranscriptome'
of entire microbial communities. Coupled with high-throughput sequencing
or the construction of cDNA microarrays, this approach will provide a
useful tool to study the transcriptional activities of microorganisms,
including those of entire microbial communities and of non-culturable
microorganisms.

<>

<1>McGrath, S., Seegers, J.F.M.L., Fitzgerald, G.F., van Sinderen, D.
<2>Molecular characterization of a phage-encoded resistance system in Lactococcus lactis.
<3>Appl. Environ. Microbiol.
<4>65
<5>1891-1899
<6>1999
<7>A specific fragment of the genome of Tuc2009, a temperate lactococcal bacteriophage, was shown
to contain several open reading frames, whose deduced protein products exhibited similarities
to proteins known to be involved in DNA replication and modification.  In this way, a putative
single-stranded binding protein, replisome organizer protein, topoisomerase I, and a methylase
were identified.  When the genetic information coding for the putative replisome organizer
protein of Tuc2009, Rep2009, was supplied on a high-copy-number plasmid vector, it was shown
to confer a phage-encoded resistance (Per) phenotype on its lactococcal host UC509.9.  The
presence of this recombinant plasmid was shown to cause a marked reduction in Tuc2009 DNA
replication, suggesting that the observed phage resistance was due to titration of a factor,
or factors, required for Tuc2009 DNA replication.  Further experiments delineated the phage
resistance-conferring region to a 160-bp fragment rich in direct repeats.  Gel retardation
experiments, which indicated a protein-DNA interaction between this 160-bp fragment and the
Rep2009 protein, were performed.  UC509.9 strains harboring plasmids with randomly mutated
versions of this fragment were shown to display a variable phage resistance phenotype,
depending on the position of the mutations.

<>

<1>McGraw, B.R., Marinus, M.G.
<2>Isolation and characterization of Dam+ revertants and suppressor mutations that modify secondary phenotypes of dam-3 strains of Escherichia coli K-12.
<3>Mol. Gen. Genet.
<4>178
<5>309-315
<6>1980
<7>Bacteria mutant in the dam (DNA adenine methylation) gene and in either recA or recB or recC
genes are inviable (Virm- phenotype). From crosses between dam-3 bacteria and recA1 or recB21
recC22 strains, Vrm+ recombinants were recovered. Among these recombinants, Dam+ revertants
were present which did not show the phenotypes normally associated with dam-3 bacteria. Three
classes of indirectly suppressed strains (dam 3 genotype) were also recovered which showed
alterations in the secondary phenotypes normally associated with dam-3 bacteria. These strains
contained a second unlinked mutation in either mutL or mutS or sin. In addition, mutation in
either sbcA or sbcB supresses the Vrm-phenotype of dam-3 recB21 recC22 strains.

<>

<1>McInerney, M.J., Rohlin, L., Mouttaki, H., Kim, U., Krupp, R.S., Rios-Hernandez, L., Sieber, J., Struchtemeyer, C.G., Bhattacharyya, A., Campbell, J.W., Gunsalus, R.P.
<2>The genome of Syntrophus aciditrophicus: Life at the thermodynamic limit of microbial growth.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>7600-7605
<6>2007
<7>Biochemically, the syntrophic bacteria constitute the missing link in our understanding of
anaerobic flow of carbon in the biosphere. The completed genome sequence of Syntrophus
aciditrophicus SB, a model fatty acid- and aromatic acid-degrading syntrophic bacterium,
provides a glimpse of the composition and architecture of the electron transfer and
energy-transducing systems needed to exist on marginal energy economies of a syntrophic
lifestyle. The genome contains 3,179,300 base pairs and 3,169 genes where 1,618 genes were
assigned putative functions. Metabolic reconstruction of the gene inventory revealed that most
biosynthetic pathways of a typical Gram-negative microbe were present. A distinctive feature
of syntrophic metabolism is the need for reverse electron transport; the presence of a unique
Rnf-type ion-translocating electron transfer complex, menaquinone, and membrane-bound Fe-S
proteins with associated heterodisulfide reductase domains suggests mechanisms to accomplish
this task. Previously undescribed approaches to degrade fatty and aromatic acids, including
multiple AMP-forming CoA ligases and acyl-CoA synthetases seem to be present as ways to form
and dissipate ion gradients by using a sodium-based energy strategy. Thus, S. aciditrophicus,
although nutritionally self-sufficient, seems to be a syntrophic specialist with limited
fermentative and respiratory metabolism. Genomic analysis confirms the S. aciditrophicus
metabolic and regulatory commitment to a nonconventional mode of life compared with our
prevailing understanding of microbiology.

<>

<1>McKane, M., Milkman, R.
<2>Transduction, restriction and recombination patterns in Escherichia coli.
<3>Genetics
<4>139
<5>35-43
<6>1995
<7>Chromosomal DNA from several Escherichia coli reference (ECOR) strains was transducted by
bacteriophage P1 into E. coli strain K12 W3110 trpA33. Recombination patterns of the
transductants were determined by restriction fragment length polymorphism over a 40-kb region
centering on a single marker (trpA+) in the tryptophan operon. These experiments demonstrate
that transduction between different strains of E. coli can result in recombinational
replacements that are small in comparison to the entrant molecule (replacements average 8-14
kb, whereas P1 packages approximately 100 kb) often in a series of discrete segments. The
transduction patterns generated resemble the natural mosaic sequence patterns of the ECOR
strains described in previous work. Extensive polymorphisms in the restriction-modification
systems of the ECOR strains are a possible explanation for the sequence patterns in nature. To
test this possibility, two transductants were back-transduced into strain K12 W3110 trpA33.
The resulting patterns were strikingly different from the original transductions. The size of
the replacements was greater, and no multiple replacements were observed, suggesting a role
for restriction-modification systems in the transduction patterns and perhaps for the mosaic
sequence patterns in nature.

<>

<1>McKay, L.L., Bohanon, M.J., Polzin, K.M., Rule, P.L., Baldwin, K.A.
<2>Localization of separate genetic loci for reduced sensitivity towards small isometric-headed bacteriophage sk1 and prolate-headed bacteriophage c2 on pGBK17 from Lactococcus lactis subsp. lactis KR2.
<3>Appl. Environ. Microbiol.
<4>55
<5>2702-2709
<6>1989
<7>The mechanism of reduced sensitivity to the small isometric-headed bacteriophage sk1 encoded
on a 19-kilobase (kb) HpaII fragment subcloned from pKR223 of Lactococcus lactis subsp. lactis
KR2 was examined. The reduced sensitivity to phage sk1 was due to a modest
restriction/modification (R/M) system that was not active against prolate-headed phage c2. The
genetic loci for the R/M system against sk1 and the abortive phage infection (Abi) mechanism
effective against phage c2 were then localized by restriction mapping, subcloning, and
deletion analysis. The restriction gene was localized to a region of a 2.7-kb EcoRV fragment
and included an EcoRI site within that fragment. The modification gene was found to be
physically separate from the restriction gene and was present on a 1.75-kb BstEII-XbaI
fragment. The genetic locus for the Abi phenotype against phage c2 was localized to a region
containing a 1.3-kb EcoRI fragment. Attempts to clone the c2 Abi mechanism independent of the
sk1 R/M system were unsuccessful, suggesting that expression of the abi genes required
sequences upstream of the modification gene. Some pGBK17 (vector pGB301 plus a 19-kb HpaII
insert fragment) transformants exhibited the R/M system against phage sk1 but lost the Abi
mechanism against phage c2. These transformants contained a 1.2-1.3-kb insertion in the Abi
region. The data identified genetic loci on a cloned 19-kb HpaII fragment responsible for
restriction activity and for modification activity against a small isometric-headed phage and
for Abi activity against prolate-headed phage c2. A putative insertion element was also found
to inactivate the abi gene(s).

<>

<1>McKelvie, J.C., Richards, M.I., Harmer, J.E., Milne, T.S., Roach, P.L., Oyston, P.C.F.
<2>Inhibition of Yersinia pestis DNA adenine methyltransferase in vitro by a stibonic acid compound: identification of a potential novel class of antimicrobial agents.
<3>Br. J. Pharmacol.
<4>168
<5>172-188
<6>2013
<7>Background and Purpose Multiple antibiotic resistant strains of plague are emerging, driving a
need for the development of novel antibiotics
effective against Yersinia pestis. DNA adenine methylation regulates
numerous fundamental processes in bacteria and alteration of DNA
adenine methlytransferase (Dam) expression is attenuating for several
pathogens, including Y. pestis. The lack of a functionally similar
enzyme in humans makes Dam a suitable target for development of novel
therapeutics for plague. Experimental Approach Compounds were evaluated
for their ability to inhibit Dam activity in a high-throughput
screening assay. DNA was isolated from Yersinia grown in the presence
of lead compounds and restricted to determine the effect of inhibitors
on DNA methylation. Transcriptional analysis was undertaken to
determine the effect of an active inhibitor on virulence-associated
phenotypes. Key Results We have identified a series of aryl stibonic
acids which inhibit Dam in vitro. The most active,
4-stibonobenzenesulfonic acid, exhibited a competitive mode of
inhibition with respect to DNA and a Ki of 6.46 nM. One compound was
found to inhibit DNA methylation in cultured Y. pestis. The effects of
this inhibition on the physiology of the cell were widespread, and
included altered expression of known virulence traits, including iron
acquisition and Type III secretion. Conclusions and Implications We
have identified a novel class of potent Dam inhibitors. Treatment of
bacterial cell cultures with these inhibitors resulted in a decrease in
DNA methylation. Expression of virulence factors was affected,
suggesting these inhibitors may attenuate bacterial infectivity and
function as antibiotics.

<>

<1>McKenna, W.G., Brown, F.L., Musich, P.R., Maio, J.J.
<2>Cleavage of mammalian repetitive deoxyribonucleic acids by a mammalian site-specific endodeoxyribonuclease.
<3>J. Mol. Biol.
<4>154
<5>379-384
<6>1982
<7>We probed the structure of mammalian repetitive DNAs with a site-specific
mammalian endodeoxyribonuclease, which we recently identified, and which
apparently represents a common enzyme activity among the mammals (McKenna et
al., 1981).  With several of the DNAs (e.g. mouse satellite, guinea pig
beta-satellite, variable repeated spacer DNA from mouse ribosomal genes and
primate alphoid sequences), the endonuclease activity gave highly specific
cleavage patterns when the digestion products were analyzed by gel
electrophoresis.  These patterns were not always identical to those produced by
microbial restriction enzymes.  However, in other cases (e.g. bovid and caprid
satellites and guinea pig alpha-satellite) the repetitive DNAs appeared to be
degraded randomly.  Thus, the mammalian enzyme reveals structural features of
the repetitive sequences that are not rendered immediately obvious by microbial
restriction enzyme analysis.  Evidence from mapping data presented here
suggests that the mammalian site-specific endonucleases are not sequence
specific but have special affinity for imperfect or hyphenated palindromic
sequences in repetitive DNAs and in other eukaryrotic DNA sequences.

<>

<1>McKenna, W.G., Maio, J.J., Brown, F.L.
<2>Purification and properties of a mammalian endonuclease showing site-specific cleavage of DNA.
<3>J. Biol. Chem.
<4>256
<5>6435-6443
<6>1981
<7>An endonuclease activity has been identified from a variety of mammalian
sources which cleaves native viral DNAs (adenovirus-2, SV40) and mammalian
repetitive DNA to yield specific, double-stranded DNA segments when the
cleavage products are analyzed by gel electrophoresis.  The activity has been
purified 750-fold from bull testes by ammonium sulfate precipitations and ion
exchange chromatography.  The enzyme has a pH optimum of 7.5 and shows an
absolute requirement for Mg2+ or Mn2+ and reducing agents in the reaction
buffer.  It is strongly inhibited by salt and stimulated by glycerol or
dimethyl sulfoxide.  By glycerol gradient analysis it has a sedimentation
coefficient of 4.5 S (65,000 daltons if globular) and it may exist as a dimer
of 6.4 S (130,000 daltons).  The enzyme liberates 3'-OH and 5'-P termini in its
cleavage of mammalian viral and repetitive DNAs and there is a clear
nonrandomness in the 5'-terminal nucleotides: purines and 5'-pG in particular
predominate.  The similarities between the mammalian endonuclease activity and
the bacterial restriction enzymes may be superficial.  By gel electrophoresis
the mammalian endonuclease produces its most characteristic series of segments
with viral DNAs in the molecular weight range of 150,000 to 1,500.000  These
are true double-stranded intermediate products in the cleavage of high
molecular weight DNA but they are not end products in the reaction as they
would be in the case of the bacterial restriction enzymes.  With time, the
endonuclease degrades viral DNA to shorter and shorter segments without
releasing acid-soluble nucleotides at any stage in the digestion and the final
end products are about 120 base pairs long.  The enzyme degrades both double-
and single-stranded DNA endonucleolytically though it shows site specificity
(production of discrete segments upon gel electrophoresis) only with the
double-stranded DNA substrates.

<>

<1>McKernan, K.J., Spangler, J., Helbert, Y., Zhang, L., Tadigotla, V.
<2>DREAMing of a patent-free human genome for clinical sequencing.
<3>Nat. Biotechnol.
<4>31
<5>884-887
<6>2013
<7>Can methylation be the key to challenging the interpretation of existing gene patent claims?
And can a novel PCR method be used to enable the sequencing of hundreds of genes?  The case
for the perils of gene patents and the negative impact of a profit motive in scientific
endeavors has been made.  Often, gene patent discussions will conflate economic and ethical
concerns while failing to properly define the scope of property or the influence of profit
motivations.  In an attempt to untangle these two topics, we review the impact of methylation
on the scope of gene patent property rights and suggest a simple PCR strategy that may
challenge the interpretation of many patent claims still in force after the US Supreme
Court's decision in Associaton for Moleuclar Pathology v. Mayriad Genetics.  We also offer an
alternatie economic perspective on profit motivation and its impact on the ethics of gene
patents.

<>

<1>McLaughlin, L.W., Benseler, F., Graeser, E., Piel, N., Scholtissek, S.
<2>Effects of functional group changes in the EcoRI recognition site on the cleavage reaction catalyzed by the endonuclease.
<3>Biochemistry
<4>26
<5>7238-7245
<6>1987
<7>Oligodeoxynucleotides have been prepared that contain changes in the functional
group pattern present in the EcoRI recognition site.  These changes involve
functional group deletions, functional group reversals, and displaced
functional groups.  Steady-state kinetic parameters have been used to
characterize the interaction of these modified recognition sites with the EcoRI
endonuclease.  Changes in the functional group pattern have varying effects
upon the cleavage reaction.  Both the exocyclic amino groups of the two adenine
residues and the methyl groups of the thymine residues appear to interact with
the endonuclease quite differently.  In both cases efficient catalysis was
observed when these functional groups were present at the outer dA-dT base
pair.  Selectivity was decreased by over an order of magnitude largely via
increases in Km when these functional groups were deleted.  Similar
modifications at the inner dA-dT base pair did not alter the kinetic parameters
significantly from those observed with the native sequence.  Addition of an
amino group to the minor groove at the outer dA-dT base pair resulted in a
modified recognition site that interacted with the enzyme, on the basis of
observed competitive inhibition kinetics, but was not cleaved.

<>

<1>McLaughlin, R.W. et al.
<2>Draft Genome Sequence of Clostridium mangenotii TR, Isolated from the Fecal Material of a Timber Rattlesnake.
<3>Genome Announcements
<4>2
<5>e01107-13
<6>2014
<7>Here, we report the draft genome sequence of Clostridium mangenotii strain TR, which was
isolated from the fecal material of a timber rattlesnake. This
bacterium is nonpathogenic but contains 68 genes involved in virulence, disease,
and defense.

<>

<1>McLeod, A., Brede, D.A., Rud, I., Axelsson, L.
<2>Genome Sequence of Lactobacillus sakei subsp. sakei LS25, a Commercial Starter Culture Strain for Fermented Sausage.
<3>Genome Announcements
<4>1
<5>e00475-13
<6>2013
<7>Lactobacillus sakei is a lactic acid bacterium associated primarily with fermented meat and
fish. Here, we present the draft genome sequence of L. sakei
subsp. sakei strain LS25, a commercial starter culture strain for fermented
sausage.

<>

<1>McLeod, M.P. et al.
<2>The complete genome of Rhodococcus sp. RHA1 provides insights into a catabolic powerhouse.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>15582-15587
<6>2006
<7>Rhodococcus sp. RHA1 (RHA1) is a potent polychlorinated biphenyl-degrading soil actinomycete
that catabolizes a wide range of compounds and
represents a genus of considerable industrial interest. RHA1 has one of
the largest bacterial genomes sequenced to date, comprising 9,702,737 bp
(67% G+C) arranged in a linear chromosome and three linear plasmids. A
targeted insertion methodology was developed to determine the telomeric
sequences. RHA1's 9,145 predicted protein-encoding genes are exceptionally
rich in oxygenases (203) and ligases (192). Many of the oxygenases occur
in the numerous pathways predicted to degrade aromatic compounds (30) or
steroids (4). RHA1 also contains 24 nonribosomal peptide synthase genes,
six of which exceed 25 kbp, and seven polyketide synthase genes, providing
evidence that rhodococci harbor an extensive secondary metabolism. Among
sequenced genomes, RHA1 is most similar to those of nocardial and
mycobacterial strains. The genome contains few recent gene duplications.
Moreover, three different analyses indicate that RHA1 has acquired fewer
genes by recent horizontal transfer than most bacteria characterized to
date and far fewer than Burkholderia xenovorans LB400, whose genome size
and catabolic versatility rival those of RHA1. RHA1 and LB400 thus appear
to demonstrate that ecologically similar bacteria can evolve large genomes
by different means. Overall, RHA1 appears to have evolved to
simultaneously catabolize a diverse range of plant-derived compounds in an
O(2)-rich environment. In addition to establishing RHA1 as an important
model for studying actinomycete physiology, this study provides critical
insights that facilitate the exploitation of these industrially important
microorganisms.

<>

<1>McMahon, S.A., Roberts, G.A., Johnson, K.A., Cooper, L.P., Liu, H., White, J.H., Carter, L.G., Sanghvi, B., Oke, M., Walkinshaw, M.D., Blakely, G.W., Naismith, J.H., Dryden, D.T.
<2>Extensive DNA mimicry by the ArdA anti-restriction protein and its role in the spread of antibiotic resistance.
<3>Nucleic Acids Res.
<4>37
<5>4887-4897
<6>2009
<7>The ardA gene, found in many prokaryotes including important pathogenic species, allows
associated mobile genetic elements to evade the ubiquitous
Type I DNA restriction systems and thereby assist the spread of resistance
genes in bacterial populations. As such, ardA contributes to a major
healthcare problem. We have solved the structure of the ArdA protein from
the conjugative transposon Tn916 and find that it has a novel extremely
elongated curved cylindrical structure with defined helical grooves. The
high density of aspartate and glutamate residues on the surface follow a
helical pattern and the whole protein mimics a 42-base pair stretch of
B-form DNA making ArdA by far the largest DNA mimic known. Each monomer of
this dimeric structure comprises three alpha-beta domains, each with a
different fold. These domains have the same fold as previously determined
proteins possessing entirely different functions. This DNA mimicry
explains how ArdA can bind and inhibit the Type I restriction enzymes and
we demonstrate that 6 different ardA from pathogenic bacteria can function
in Escherichia coli hosting a range of different Type I restriction
systems.

<>

<1>McMullen, P.D., Gillaspy, A.F., Gipson, J., Bobo, L.D., Skiest, D.J., Freitag, N.E.
<2>Genome Sequence of Listeria monocytogenes 07PF0776, a Cardiotropic Serovar 4b Strain.
<3>J. Bacteriol.
<4>194
<5>3552
<6>2012
<7>Listeria monocytogenes is a food-borne bacterial pathogen commonly associated with serious
invasive infections of the central nervous system or of the
developing fetus. We present the genome sequence of Listeria monocytogenes
07PF0776, a serovar 4b isolate from a human myocardial abscess that exhibits
enhanced invasion of cardiac tissue.

<>

<1>McMurdie, P.J., Behrens, S.F., Muller, J.A., Goke, J., Ritalahti, K.M., Wagner, R., Goltsman, E., Lapidus, A., Holmes, S., Loffler, F.E., Spormann, A.M.
<2>Localized plasticity in the streamlined genomes of vinyl chloride respiring Dehalococcoides.
<3>PLoS Genet.
<4>5
<5>E1000714
<6>2009
<7>Vinyl chloride (VC) is a human carcinogen and widespread priority
pollutant. Here we report the first, to our knowledge, complete genome
sequences of microorganisms able to respire VC, Dehalococcoides sp.
strains VS and BAV1. Notably, the respective VC reductase encoding genes,
vcrAB and bvcAB, were found embedded in distinct genomic islands (GEIs)
with different predicted integration sites, suggesting that these genes
were acquired horizontally and independently by distinct mechanisms. A
comparative analysis that included two previously sequenced
Dehalococcoides genomes revealed a contextually conserved core that is
interrupted by two high plasticity regions (HPRs) near the Ori. These HPRs
contain the majority of GEIs and strain-specific genes identified in the
four Dehalococcoides genomes, an elevated number of repeated elements
including insertion sequences (IS), as well as 91 of 96 rdhAB, genes that
putatively encode terminal reductases in organohalide respiration. Only
three core rdhA orthologous groups were identified, and only one of these
groups is supported by synteny. The low number of core rdhAB, contrasted
with the high rdhAB numbers per genome (up to 36 in strain VS), as well as
their colocalization with GEIs and other signatures for horizontal
transfer, suggests that niche adaptation via organohalide respiration is a
fundamental ecological strategy in Dehalococccoides. This adaptation has
been exacted through multiple mechanisms of recombination that are mainly
confined within HPRs of an otherwise remarkably stable, syntenic,
streamlined genome among the smallest of any free-living microorganism.

<>

<1>McMurrough, T.A., Dickson, R.J., Thibert, S.M.F., Gloor, G.B., Edgell, D.R.
<2>Control of catalytic efficiency by a coevolving network of catalytic and noncatalytic residues.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>111
<5>E2376-E2383
<6>2014
<7>The active sites of enzymes consist of residues necessary for catalysis and structurally
important noncatalytic residues that together maintain the architecture and function of the
active site. Examples of evolutionary interactions between catalytic and non-catalytic
residues have been difficult to define and experimentally validate due to a general
intolerance of these residues to substitution. Here, using computational methods to predict
coevolving residues, we identify a network of positions consisting of two catalytic
metal-binding residues and two adjacent noncatalytic residues in LAGLIDADG homing
endonucleases (LHEs). Distinct combinations of the four residues in the network map to
distinct LHE subfamilies, with a striking distribution of the metal-binding Asp (D) and Glu
(E) residues. Mutation of these four positions in three LHEs-I-LtrI, I-OnuI, and
I-HjeMI-indicate that the combinations of residues tolerated are specific to each enzyme.
Kinetic analyses under single-turnover conditions revealed that I-LtrI activity could be
modulated over an similar to 100-fold range by mutation of residues in the coevolving network.
I-LtrI catalytic site variants with low activity could be rescued by compensatory mutations at
adjacent noncatalytic sites that restore an optimal coevolving network and vice versa. Our
results demonstrate that LHE activity is constrained by an evolutionary barrier of residues
with strong context-dependent effects. Creation of optimal coevolving active-site networks is
therefore an important consideration in engineering of LHEs and other enzymes.

<>

<1>McNamara, A.R., Hurd, P.J., Smith, A.E.F., Ford, K.G.
<2>Characterisation of site-biased DNA methyltransferases: specificity, affinity and subsite relationships.
<3>Nucleic Acids Res.
<4>30
<5>3818-3830
<6>2002
<7>DNA methylation is now seen as a primary signal in the cell for mediating transcriptional
repression through chromatin formation. The construction and evaluation of enzymes capable of
influencing this process in vivo is therefore of significant interest. We have fused the
C5-cytosine DNA methyltransferases, M.HhaI and M.HpaII, which both methylate 4 bp sequences
containing a CpG dinucleotide, to a three zinc finger protein recognising a 9 bp DNA sequence.
DNA methylation analyses demonstrate specific DNA methylation by both enzymes at target sites
comprising adjacent methyltransferase and zinc finger subsites, targeted M.HpaII being the
most specific. Binding analysis of the targeted M.HpaII enzyme reveals an 8-fold preference
for binding to its target site, compared to binding to a zinc finger site alone, and an
18-fold preference over binding to a methyltransferase site alone, thereby demonstrating
enhanced binding by the fusion protein, compared to its component proteins. Both DNA binding
and methylation are specific for the target site up to separations of approximately 40 bp
between the zinc finger and methyltransferase subsites. Ex vivo plasmid methylation
experiments are also described that demonstrate targeted methylation. These targeted enzymes,
however, are shown to be not fully mono-functional, retaining a significant non-targeted
activity most evident at elevated protein concentrations.

<>

<1>McNulty, N.P. et al.
<2>The impact of a consortium of fermented milk strains on the gut microbiome of gnotobiotic mice and monozygotic twins.
<3>Sci. Transl. Med.
<4>3
<5>106RA106
<6>2011
<7>Understanding how the human gut microbiota and host are affected by probiotic bacterial
strains requires carefully controlled studies in humans and in mouse models of the gut
ecosystem where potentially confounding variables that are difficult to control in humans can
be constrained. Therefore, we characterized the fecal microbiomes and metatranscriptomes of
adult female monozygotic twin pairs through repeated sampling 4 weeks before, 7 weeks during,
and 4 weeks after consumption of a commercially available fermented milk product (FMP)
containing a consortium of Bifidobacterium animalis subsp. lactis, two strains of
Lactobacillus delbrueckii subsp. bulgaricus, Lactococcus lactis subsp. cremoris, and
Streptococcus thermophilus. In addition, gnotobiotic mice harboring a 15-species model human
gut microbiota whose genomes contain 58,399 known or predicted protein-coding genes were
studied before and after gavage with all five sequenced FMP strains. No significant changes in
bacterial species composition or in the proportional representation of genes encoding known
enzymes were observed in the feces of humans consuming the FMP. Only minimal changes in
microbiota configuration were noted in mice after single or repeated gavage with the FMP
consortium. However, RNA-Seq analysis of fecal samples and follow-up mass spectrometry of
urinary metabolites disclosed that introducing the FMP strains into mice results in
significant changes in expression of microbiome-encoded enzymes involved in numerous metabolic
pathways, most prominently those related to carbohydrate metabolism. B. animalis subsp.
lactis, the dominant persistent member of the FMP consortium in gnotobiotic mice, up-regulates
a locus in vivo that is involved in the catabolism of xylooligosaccharides, a class of glycans
widely distributed in fruits, vegetables, and other foods, underscoring the importance of
these sugars to this bacterial species. The human fecal metatranscriptome exhibited
significant changes, confined to the period of FMP consumption, that mirror changes in
gnotobiotic mice, including those related to plant polysaccharide metabolism. These
experiments illustrate a translational research pipeline for characterizing the effects of
FMPs on the human gut microbiome.

<>

<1>McShan, W.M., Ferretti, J.J., Karasawa, T., Suvorov, A.N., Lin, S., Qin, B., Jia, H., Kenton, S., Najar, F., Wu, H., Scott, J., Roe, B.A., Savic, D.J.
<2>Genome Sequence of a Nephritogenic and Highly Transformable M49 Strain of Streptococcus pyogenes.
<3>J. Bacteriol.
<4>190
<5>7773-7785
<6>2008
<7>The 1,815,783-bp genome of a serotype M49 strain of Streptococcus pyogenes
(group A streptococcus [GAS]), strain NZ131, has been determined. This GAS
strain (FCT type 3; emm pattern E), originally isolated from a case of
acute post-streptococcal glomerulonephritis, is unusually competent for
electrotransformation and has been used extensively as a model organism
for both basic genetic and pathogenesis investigations. As with the
previously sequenced S. pyogenes genomes, three unique prophages are a
major source of genetic diversity. Two clustered regularly interspaced
short palindromic repeat (CRISPR) regions were present in the genome,
providing genetic information on previous prophage encounters. A unique
cluster of genes was found in the pathogenicity island-like emm region
that included a novel Nudix hydrolase, and, further, this cluster appears
to be specific for serotype M49 and M82 strains. Nudix hydrolases
eliminate potentially hazardous materials or prevent the unbalanced
accumulation of normal metabolites; in bacteria, these enzymes may play a
role in host cell invasion. Since M49 S. pyogenes strains have been known
to be associated with skin infections, the Nudix hydrolase and its
associated genes may have a role in facilitating survival in an
environment that is more variable and unpredictable than the uniform
warmth and moisture of the throat. The genome of NZ131 continues to shed
light upon the evolutionary history of this human pathogen. Apparent
horizontal transfer of genetic material has led to the existence of highly
variable virulence-associated regions that are marked by multiple
rearrangements and genetic diversification while other regions, even those
associated with virulence, vary little between genomes. The genome regions
that encode surface gene products that will interact with host targets or
aid in immune avoidance are the ones that display the most sequence
diversity. Thus, while natural selection favors stability in much of the
genome, it favors diversity in these regions.

<>

<1>Mead, D., Swaminathan, N., Van Etten, J., Skowron, P.
<2>Dinucleotide restriction endonuclease preparations and methods of use.
<3>International Patent Office
<4>WO 9421663 A
<5>
<6>1994
<7>The present invention is directed to materials and methods for the quasi-random and complete
fragmentation of DNA using restriction endonuclease reagents capable of cutting DNA at a
dinucleotide sequence. The invention is also directed to methods for labeling DNA, for shotgun
cloning, for sequencing of DNA, for epitope mapping and for anonymous primer cloning, all
using fragments of DNA generated by the method of the present invention. In addition, the
present invention is directed to DNA sequences encoding a novel restriction endonuclease
(designated R.CviJI) and variants thereof as well as to methods and materils for production of
the same by recombinant methods. A bacterial host cell transformed with DNA encoding R.CviJI
is also disclosed as well as methods for expressing R.CviJI in the bacterial host system and
subsequent materials and methods for purifying the enzyme.

<>

<1>Mead, D., Swaminathan, N., Van Etten, J., Skowron, P.
<2>Recombinant CviJI restriction endonuclease.
<3>US Patent Office
<4>US 5472872
<5>
<6>1995
<7>DNA sequences encoding a novel restriction endonuclease (designated R.CviJI) and variants
thereof are disclosed along with method and materials for production of the same by
recombinant methods.  A bacterial host cell line transformed with DNA encoding R.CviJI is also
disclosed, as well as methods for expressing R.CviJI in the bacterial host system and
subsequent materials and methods for purification of the enzyme.

<>

<1>Mead, D.A. et al.
<2>Complete Genome Sequence of Paenibacillus strain Y4.12MC10, a Novel Paenibacillus lautus strain Isolated from Obsidian Hot Spring in Yellowstone National Park.
<3>Standards in Genomic Sciences
<4>6
<5>381-400
<6>2012
<7>Paenibacillus sp.Y412MC10 was one of a number of organisms isolated from Obsidian Hot Spring,
Yellowstone National Park, Montana, USA under permit from the
National Park Service. The isolate was initially classified as a Geobacillus sp.
Y412MC10 based on its isolation conditions and similarity to other organisms
isolated from hot springs at Yellowstone National Park. Comparison of 16 S rRNA
sequences within the Bacillales indicated that Geobacillus sp.Y412MC10 clustered
with Paenibacillus species, and the organism was most closely related to
Paenibacillus lautus. Lucigen Corp. prepared genomic DNA and the genome was
sequenced, assembled, and annotated by the DOE Joint Genome Institute. The genome
sequence was deposited at the NCBI in October 2009 (NC_013406). The genome of
Paenibacillus sp. Y412MC10 consists of one circular chromosome of 7,121,665 bp
with an average G+C content of 51.2%. Comparison to other Paenibacillus species
shows the organism lacks nitrogen fixation, antibiotic production and social
interaction genes reported in other paenibacilli. The Y412MC10 genome shows a
high level of synteny and homology to the draft sequence of Paenibacillus sp.
HGF5, an organism from the Human Microbiome Project (HMP) Reference Genomes.
This, combined with genomic CAZyme analysis, suggests an intestinal, rather than
environmental origin for Y412MC10.

<>

<1>Meda, M.M., Menin, J.F.
<2>Isolated DNA encoding the HphI restriction endonuclease and related methods for producing the same.
<3>US Patent Office
<4>US 5731185
<5>
<6>1998
<7>The present invention is directed to a method for cloning and producing the HphI restriction
endonuclease by 1)linking up the restriction gene to transcription elements that can be
processed in E. coli and 2) expressing DNA modification enzymes in the host that protect
against HphI digestion.

<>

<1>Medema, M.H., Trefzer, A., Kovalchuk, A., van den Berg, M., Muller, U., Heijne, W., Wu, L., Alam, M.T., Ronning, C.M., Nierman, W.C., Bovenberg, R.A., Breitling, R., Takano, E.
<2>The sequence of a 1.8-mb bacterial linear plasmid reveals a rich evolutionary reservoir of secondary metabolic pathways.
<3>Genome Biol. Evol.
<4>2
<5>212-224
<6>2010
<7>Plasmids are mobile genetic elements that play a key role in the evolution
of bacteria by mediating genome plasticity and lateral transfer of useful
genetic information. Although originally considered to be exclusively
circular, linear plasmids have also been identified in certain bacterial
phyla, notably the actinomycetes. In some cases, linear plasmids engage
with chromosomes in an intricate evolutionary interplay, facilitating the
emergence of new genome configurations by transfer and recombination or
plasmid integration. Genome sequencing of Streptomyces clavuligerus ATCC
27064, a Gram-positive soil bacterium known for its production of a
diverse array of biotechnologically important secondary metabolites,
revealed a giant linear plasmid of 1.8 Mb in length. This megaplasmid
(pSCL4) is one of the largest plasmids ever identified and the largest
linear plasmid to be sequenced. It contains more than 20% of the putative
protein-coding genes of the species, but none of these is predicted to be
essential for primary metabolism. Instead, the plasmid is densely packed
with an exceptionally large number of gene clusters for the potential
production of secondary metabolites, including a large number of putative
antibiotics, such as staurosporine, moenomycin, beta-lactams, and
enediynes. Interestingly, cross-regulation occurs between chromosomal and
plasmid-encoded genes. Several factors suggest that the megaplasmid came
into existence through recombination of a smaller plasmid with the arms of
the main chromosome. Phylogenetic analysis indicates that heavy traffic of
genetic information between Streptomyces plasmids and chromosomes may
facilitate the rapid evolution of secondary metabolite repertoires in
these bacteria.

<>

<1>Mediannikov, O., El Karkouri, K., Diatta, G., Robert, C., Fournier, P.E., Raoult, D.
<2>Non-contiguous finished genome sequence and description of Bartonella senegalensis sp. nov.
<3>Standards in Genomic Sciences
<4>8
<5>279-289
<6>2013
<7>Bartonella senegalensis sp. nov. strain OS02(T) is the type strain of B. senegalensis sp.
nov., a new species within the genus Bartonella. This strain,
whose genome is described here, was isolated in Senegal from the soft tick
Ornithodoros sonrai, the vector of relapsing fever. B. senegalensis is an
aerobic, rod-shaped, Gram-negative bacterium. Here we describe the features of
this organism, together with the complete genome sequence and its annotation. The
1,966,996 bp-long genome contains 1,710 protein-coding and 46 RNA genes,
including 6 rRNA genes.

<>

<1>Mediannikov, O., El Karkouri, K., Robert, C., Fournier, P.E., Raoult, D.
<2>Non-contiguous finished genome sequence and description of Bartonella florenciae  sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>185-196
<6>2013
<7>Bartonella florenciae sp. nov. strain R4(T) is the type strain of B. florenciae sp. nov., a
new species within the genus Bartonella. This strain, whose genome is
described here, was isolated in France from the spleen of the shrew Crocidura
russula. B. florenciae is an aerobic, rod-shaped, Gram-negative bacterium. Here
we describe the features of this organism, together with the complete genome
sequence and its annotation. The 2,010,844 bp-long genome contains 1,909
protein-coding and 46 RNA genes, including two rRNA operons.

<>

<1>Medigue, C. et al.
<2>Coping with cold: the genome of the versatile marine Antarctica bacterium Pseudoalteromonas haloplanktis TAC125.
<3>Genome Res.
<4>15
<5>1325-1335
<6>2005
<7>A considerable fraction of life develops in the sea at temperatures lower than 15 degrees C.
Little is known about the adaptive features selected
under those conditions. We present the analysis of the genome sequence of
the fast growing Antarctica bacterium Pseudoalteromonas haloplanktis
TAC125. We find that it copes with the increased solubility of oxygen at
low temperature by multiplying dioxygen scavenging while deleting whole
pathways producing reactive oxygen species. Dioxygen-consuming lipid
desaturases achieve both protection against oxygen and synthesis of lipids
making the membrane fluid. A remarkable strategy for avoidance of reactive
oxygen species generation is developed by P. haloplanktis, with
elimination of the ubiquitous molybdopterin-dependent metabolism. The P.
haloplanktis proteome reveals a concerted amino acid usage bias specific
to psychrophiles, consistently appearing apt to accommodate asparagine, a
residue prone to make proteins age. Adding to its originality, P.
haloplanktis further differs from its marine counterparts with recruitment
of a plasmid origin of replication for its second chromosome.

<>

<1>Medina-Franco, J.L., Caulfield, T.
<2>Advances in the computational development of DNA methyltransferase inhibitors.
<3>Drug Discovery Today
<4>16
<5>418-425
<6>2011
<7>DNA methylation is an epigenetic change that results in the addition of a methyl group at the
carbon-5 position of cytosine residues. The process is mediated by DNA methyltransferases
(DNMTs), a family of enzymes for which inhibition is a promising strategy for the treatment of
cancer and other diseases. Here, we review the current status of the computational studies
directed to rationalize, at the molecular level, the enzymatic activity of DNMT inhibitors. We
also review successful virtual screenings to identify inhibitors with novel scaffolds as well
as the emerging efforts to characterize the dynamic behavior of DNMTs. Thus, computational
approaches form part of multidisciplinary efforts to further advance epigenetic therapies.

<>

<1>Medina-Franco, J.L., Yoo, J.
<2>Docking of a novel DNA methyltransferase inhibitor identified from high-throughput screening: insights to unveil inhibitors in chemical databases.
<3>Mol. Diversity
<4>17
<5>337-344
<6>2013
<7>Inhibitors of DNA methyltransferase (DNMT) are attractive compounds not only as potential
therapeutic agents for the treatment of cancer and
other diseases, but also as research tools to investigate the role of
DNMTs in epigenetic events. Recent advances in high-throughput
screening (HTS) for epigenetic targets and the availability of the
first crystallographic structure of human DNMT1 encourage the
integration of research strategies to uncover and optimize the activity
of DNMT inhibitors. Herein, we present a binding model of a novel
small-molecule DNMT1 inhibitor obtained by HTS, recently released in a
public database. The docking model is in agreement with key
interactions previously identified for established inhibitors using
extensive computational studies including molecular dynamics and
structure-based pharmacophore modeling. Based on the chemical structure
of the novel inhibitor, a sequential computational screening of five
chemical databases was performed to identify candidate compounds for
testing. Similarity searching followed by molecular docking of chemical
databases such as approved drugs, natural products, a DNMT-focused
library, and a general screening collection, identified at least 108
molecules with promising DNMT inhibitory activity. The chemical
structures of all hit compounds are disclosed to encourage the research
community working on epigenetics to test experimentally the enzymatic
and demethylating activity in vivo. Five candidate hits are drugs
approved for other indications and represent potential starting points
of a drug repurposing strategy.

<>

<1>Medrano, E.G., Bell, A.A.
<2>Genome Sequence of Pantoea sp. Strain Sc 1, an Opportunistic Cotton Pathogen.
<3>J. Bacteriol.
<4>194
<5>3019
<6>2012
<7>Pantoea is comprised of a broad spectrum of species, including plant pathogens. Here, we
provide an annotated genome sequence of Pantoea sp. strain Sc 1, which
was isolated from a diseased cotton boll. This research provides the first genome
sequence of a bona fide Pantoea sp. insect-vectored cotton pathogen.

<>

<1>Medrano, E.G., Forray, M.M., Bell, A.A.
<2>Complete Genome Sequence of a Klebsiella pneumoniae Strain Isolated from a Known  Cotton Insect Boll Vector.
<3>Genome Announcements
<4>2
<5>e00850-14
<6>2014
<7>Klebsiella pneumoniae (associated with bacterial pneumonia) was previously isolated from
Nezara viridula, a significant vector of cotton boll-rot pathogens.
We provide the first annotated genome sequence of the cotton opportunistic strain
K. pneumoniae 5-1. This data provides guidance to study the bases of cotton
pathogenesis by bacteria associated with vectors.

<>

<1>Medrano-Felix, A., Estrada-Acosta, M., Jimenez, M., Gomez-Gil, B., Leon-Felix, J., Amarillas, L., Chaidez, C.
<2>Draft Genome Sequence of Salmonella enterica subsp. enterica Serotype Oranienburg Strain S-76, Isolated from an Aquatic Environment.
<3>Genome Announcements
<4>1
<5>e01017-13
<6>2013
<7>Salmonella is a widespread microorganism and a common causative agent of food-borne illnesses.
Salmonella enterica subsp. enterica serotype Oranienburg is
highly prevalent in surface water from tropical ecosystems and is not commonly
related to illnesses. Here, we report the first genome sequence of Salmonella
Oranienburg strain S-76, isolated from an aquatic environment.

<>

<1>Meehan, R.R., Ulrich, E., Bird, A.P.
<2>Restriction endonuclease NciI is not blocked by CpG methylation.
<3>Nucleic Acids Res.
<4>21
<5>5517-5518
<6>1993
<7>Vertebrate DNA is highly methylated at cytosines in the dinucleotide sequence CpG. Many
restriction enzymes are unable to cleave DNA if their recognition sequences contain methylated
CpG, and this property has been used to study the pattern of DNA methylation in higher
eukaryotes. An enzyme that has recently been used in this way is NciI (recognition sequence
CCSGG, where S is C or G). It was reported that this enzyme cannot cleave a region of the
mouse actin promoter when the CpG in its recognition site is methylated on both strands, but
can cleave when either one or both strands are unmethylated. As a result, NciI has been used
to monitor the conversion of symmetrically methylated DNA to hemi-methylated DNA in a number
of systems. Its general use for this purpose depends upon the idea that NciI cannot cleave any
symmetrically methylated site. We have digested symmetrically methylated and non-methylated
DNA with NciI, and find that it can cleave both forms to completion, though at markedly
different rates.

<>

<1>Meessen-Pinard, M., Sekulovic, O., Fortier, L.C.
<2>Evidence of in vivo prophage induction during Clostridium difficile infection.
<3>Appl. Environ. Microbiol.
<4>78
<5>7662-7670
<6>2012
<7>Prophages contribute to the evolution and virulence of most bacterial pathogens,
but their role in Clostridium difficile is unclear. Here we describe the
isolation of four Myoviridae phages, MMP01, MMP02, MMP03, and MMP04, that were
recovered as free viral particles in the filter-sterilized stool supernatants of
patients suffering from C. difficile infection (CDI). Furthermore, identical
prophages were found in the chromosomes of C. difficile isolated from the
corresponding fecal samples. We therefore provide, for the first time, evidence
of in vivo prophage induction during CDI. We completely sequenced the genomes of
MMP02 and MMP04, and bioinformatics analyses did not reveal the presence of
virulence factors but underlined the unique character of MMP04. We also studied
the mobility of MMP02 and MMP04 prophages in vitro. Both prophages were
spontaneously induced, with 4 to 5 log PFU/ml detected in the culture
supernatants of the corresponding lysogens. When lysogens were grown in the
presence of subinhibitory concentrations of ciprofloxacin, moxifloxacin,
levofloxacin, or mitomycin C, the phage titers further increased, reaching 8 to 9
log PFU/ml in the case of MMP04. In summary, our study highlights the extensive
genetic diversity and mobility of C. difficile prophages. Moreover, antibiotics
known to represent risk factors for CDI, such as quinolones, can stimulate
prophage mobility in vitro and probably in vivo as well, which underscores their
potential impact on phage-mediated horizontal gene transfer events and the
evolution of C. difficile.

<>

<1>Mehling, J.S., Lavender, H., Clegg, S.
<2>A Dam methylation mutant of Klebsiella pneumoniae is partially attenuated.
<3>FEMS Microbiol. Lett.
<4>268
<5>187-193
<6>2007
<7>In Klebsiella pneumoniae, a chromosomal insertion mutation was constructed in the dam gene,
which encodes DNA adenine methylase (Dam),
resulting in a mutant unable to methylate specific nucleotides. In some
bacteria, the Dam methylase has been shown to play an important role in
virulence gene regulation as well as in methyl-directed mismatch repair
and the regulation of replication initiation. Disruption of the normal
Dam function by either eliminating or greatly increasing expression in
several organisms has been shown to cause attenuation of virulence in
murine models of infection. In K. pneumoniae, a mutation-eliminating
Dam function is shown here to result in only partial attenuation
following intranasal and intraperitoneal infection of Balb/C mice.

<>

<1>Mehnaz, S., Bauer, J.S., Gross, H.
<2>Complete Genome Sequence of the Sugar Cane Endophyte Pseudomonas aurantiaca PB-St2, a Disease-Suppressive Bacterium with Antifungal Activity toward the Plant  Pathogen Colletotrichum falcatum.
<3>Genome Announcements
<4>2
<5>e01108-13
<6>2014
<7>The endophytic bacterium Pseudomonas aurantiaca PB-St2 exhibits antifungal activity and
represents a biocontrol agent to suppress red rot disease of sugar
cane. Here, we report the completely sequenced 6.6-Mb genome of P. aurantiaca
PB-St2. The sequence contains a repertoire of biosynthetic genes for secondary
metabolites that putatively contribute to its antagonistic activity and its
plant-microbe interactions.

<>

<1>Mehra, R.S., Malhotra, V.P., Rembhotkar, G.W.
<2>Rapid purification of a restriction endonuclease from Escherichia coli RY13.
<3>Biotechnol. Tech.
<4>7
<5>411-414
<6>1993
<7>A two step method for the purification of the restriction endonuclease EcoRI was developed.
The first step involved the purification of the enzyme on a Cibacron Blue-F3GA-agarose column,
followed by a hydroxyapatite column. The enzyme was homogeneous on SDS-PAGE and completely
free from contaminating nucleases and phosphatases, and can be used for direct DNA hydrolysis.

<>

<1>Mei, Y., Sun, Y., He, J., Wang, Q., Sun, Y., Shao, W.
<2>Genome Sequences of Pseudomonas fragi Strains A22 and B25.
<3>J. Bacteriol.
<4>194
<5>3276-3277
<6>2012
<7>Pseudomonas fragi A22 is a novel isolate that produces bead-like particles (A22B) in its cell
wall. To explore the genetic basis for the formation of A22B, P.
fragi A22 and the type strain of the species, P. fragi B25, were subjected to
genome sequence analysis. Here, we report the draft genome sequences and
automatic annotation of both strains. These data offer a solid base for related
studies of P. fragi, including comparative genomics, proteomics, and gene mining.

<>

<1>Meidler, R., Morad, I., Amitsur, M., Inokuchi, H., Kaufmann, G.
<2>Detection of anticodon nuclease residues involved in tRNALys cleavage specificity.
<3>J. Mol. Biol.
<4>287
<5>499-510
<6>1999
<7>The tRNALys-specific anticodon
nuclease exists in latent form in Escherichia coli strains containing
the optional prr locus. The latency is a result of a masking
interaction between the anticodon nuclease core-polypeptide PrrC and
the Type IC DNA restriction-modification enzyme EcoprrI. Activation of
the latent enzyme by phage T4-infection elicits cleavage of tRNALys 5'
to the wobble base, yielding 5'-OH and 2', 3'-cyclic phosphate termini.
The N-proximal half of PrrC has been implicated with (A/G) TPase and
EcoprrI interfacing activities. Therefore, residues involved in
recognition and cleavage of tRNALys were searched for at the C-half.
Random mutagenesis of the low-G+C portion encoding PrrC residues
200-313 was performed, followed by selection for loss of anticodon
nuclease-dependent lethality and production of full-sized PrrC-like
protein. This process yielded a cluster of missense mutations mapping
to a region highly conserved between PrrC and two putative Neisseria
meningitidis MC58 homologues. This cluster included two adjacent
members that relaxed the inherent enzyme's cleavage specificity. We
also describe another mode of relaxed specificity, due to mere
overexpression of PrrC. This mode was shared by wild-type PrrC and the
other mutant alleles. The additional substrates recognised under the
promiscuous conditions had, in general, anticodons resembling that of
tRNALys. Taken together, the data suggest that the anticodon of tRNALys
harbours anticodon nuclease identity elements and implicates a
conserved region in PrrC in their recognition.

<>

<1>Meier-Kolthoff, J.P., Lu, M., Huntemann, M., Lucas, S., Lapidus, A., Copeland, A., Pitluck, S., Goodwin, L.A., Han, C., Tapia, R., Potter, G., Land, M., Ivanova, N., Rohde, M., Goker, M., Detter, J.C., Woyke, T., Kyrpides, N.C., Klenk, H.P.
<2>Genome sequence of the chemoheterotrophic soil bacterium Saccharomonospora cyanea type strain (NA-134(T)).
<3>Standards in Genomic Sciences
<4>9
<5>28-41
<6>2013
<7>Saccharomonospora cyanea Runmao et al. 1988 is a member of the genus Saccharomonospora in the
family Pseudonocardiaceae that is moderately well
characterized at the genome level thus far. Members of the genus
Saccharomonospora are of interest because they originate from diverse habitats,
such as soil, leaf litter, manure, compost, surface of peat, moist, over-heated
grain, and ocean sediment, where they probably play a role in the primary
degradation of plant material by attacking hemicellulose. Species of the genus
Saccharomonospora are usually Gram-positive, non-acid fast, and are classified
among the actinomycetes. S. cyanea is characterized by a dark blue (= cyan blue)
aerial mycelium. After S. viridis, S. azurea, and S. marina, S. cyanea is only
the fourth member in the genus for which a completely sequenced (non-contiguous
finished draft status) type strain genome will be published. Here we describe the
features of this organism, together with the draft genome sequence, and
annotation. The 5,408,301 bp long chromosome with its 5,139 protein-coding and 57
RNA genes was sequenced as part of the DOE funded Community Sequencing Program
(CSP) 2010 at the Joint Genome Institute (JGI).

<>

<1>Meincke, L. et al.
<2>Complete genome sequence of Polynucleobacter necessarius subsp. asymbioticus type strain (QLW-P1DMWA-1(T)).
<3>Standards in Genomic Sciences
<4>6
<5>74-83
<6>2012
<7>Polynucleobacter necessarius subsp. asymbioticus strain QLW-P1DMWA-1(T) is a planktonic
freshwater bacterium affiliated with the family Burkholderiaceae
(class Betaproteobacteria). This strain is of interest because it represents a
subspecies with cosmopolitan and ubiquitous distribution in standing freshwater
systems. The 16S-23S ITS genotype represented by the sequenced strain comprised
on average more than 10% of bacterioplankton in its home habitat. While all
strains of the subspecies P. necessarius asymbioticus are free-living freshwater
bacteria, strains belonging to the only other subspecies, P. necessarius subsp.
necessarius are obligate endosymbionts of the ciliate Euplotes aediculatus. The
two subspecies of P. necessarius are the instances of two closely related
subspecies that differ in their lifestyle (free-living vs. obligate
endosymbiont), and they are the only members of the genus Polynucleobacter with
completely sequenced genomes. Here we describe the features of P. necessarius
subsp. asymbioticus, together with the complete genome sequence and annotation.
The 2,159,490 bp long chromosome with a total of 2,088 protein-coding and 48 RNA
genes is the first completed genome sequence of the genus Polynucleobacter to be
published and was sequenced as part of the DOE Joint Genome Institute Community
Sequencing Program 2006.

<>

<1>Meinke, A., Bui, D.M., Nagy, E., Henics, T.
<2>H. pylori antigens.
<3>Japanese Patent Office
<4>JP 2007524366 A
<5>
<6>2007
<7>
<>

<1>Meinke, A., Triska, C., Henics, T., Minh-Bui, D., Nagy, E., Prustomersky, S.
<2>
<3>International Patent Office
<4>WO 2005103073 A
<5>
<6>2005
<7>The present invention discloses isolated nucleic acid molecules encoding a hyperimmune serum
ractive antigen or a fragment thereof as well as hyperimmune serum reactive antigens or
fragments thereof from enteroaggregative E. coli, entertoxicgenic E. coli, S. flexneri and C.
jejuni, methods for isolating such antigens and specific uses thereof.

<>

<1>Meints, G.A., Drobny, G.P.
<2>Dynamic impact of methylation at the m. hhai target site: a solid-state deuterium nmr study.
<3>Biochemistry
<4>40
<5>12436-12443
<6>2001
<7>Base methylation plays an important role in numerous biological functions of DNA, from
inhibition of cleavage by endonucleases to inhibition of transcription factor binding. Studies
of nucleic acid structure have shown little differences in unmethylated DNAs and the identical
sequence containing methylated analogues. We have investigated changes in the local dynamics
of DNA upon substitution of a methylated cytosine analogue for cytosine using solid-state
deuterium NMR. In particular, we have observed changes in the local dynamics at the target
site of the M.HhaI restriction system. These studies observe changes in the amplitudes of the
local backbone dynamics at the actual target site of the HhaI methyltransferase. This
conclusion is another indication that the significant result of base methylation is to perturb
the local dynamics, and therefore the local conformational flexibility, of the DNA helix,
inhibiting or restricting the protein's ability to manipulate the DNA helix in order to
perform its chemical alterations.

<>

<1>Meints, R.H., Schuster, A.M., Van Etten, J.L.
<2>Chlorella Viruses.
<3>Plant Mol. Biol. Rep.
<4>3
<5>180-187
<6>1985
<7>Although viruses that infect blue-green algae (cyanobacteria) have been
extensively studied (Sherman and Brown, 1978), little is known about viruses of
eukaryotic algae (see reviews, Lemke, 1976, Sherman and Brown, 1978, Dodds,
1979, and Dodds, 1983).  Most viruses or virus-like particles (VLP) in
eukaryotic algae have been detected by ultrastructural studies, and only a few
attempts have been made to characterize these particles, primarily because they
are difficult to obtain in reasonable quantities.  Several factors contribute
to this lack of material: (i) usually only a few algal cells contained
particles, (ii) usually the cells only contained particles at one stage of the
algal life cycle, (iii) usually the cells that had particles did not lyse, and
(iv) in most cases the particles were not infectious.  These factors, plus the
fact that some of these particles were present in multicellular filamentous
algae, have hindered the development of a biological assay for them.  Thus it
is interesting that we have recently discovered a group of large (negatively
stained particles are 150 to 190 nm in diameter) polyhedral, dsDNA-containing
viruses which infect and replicate in certain strains of unicellular,
eukaryotic, exsymbiont Chlorella-like green algae.  These viruses can be
produced in large quantities and, most importantly, the viruses can be assayed
by plaque formation on lawns of the host Chlorella.  These viruses, therefore,
have the potential to serve as excellent model systems for studying gene
regulation in a photosynthetic eukaryote in the manner that bacterio phages
served as model systems for studying gene regulation in bacteria.  This review
will briefly describe some of the pertinent properties of these viruses,
focusing primarily on the most studied virus PBCV-1.  A more comprehensive
review of these viruses is in press (Van Etten et al., 1986).

<>

<1>Meiron, H., Nahon, E., Raveh, D.
<2>Identification of the heterothallic mutation in HO-endonuclease of S. cerevisiae using HO/ho chimeric genes.
<3>Curr. Genet.
<4>28
<5>367-373
<6>1995
<7>HO-endonuclease initiates a mating-type switch in the yeast S. cerevisiae by making a
double-strand cleavage in the DNA of the mating-type gene, MAT.  Heterothallic strains of
yeast have a stable mating type and contain a recessive ho allele.  Here we report the
sequence of the ho allele; ho has four point mutations all of which encode for substitute
amino acids.  The fourth mutation is a leucine to histidine substitution within a presumptive
zinc finger.  Chimeric HO/ho genes were constructed in vivo by converting different parts of
the sequence of the genomic ho allele to the HO sequence by gene conversion.  HO activity was
assessed by three bioassays: a mating-type switch, extinction of expression of an a-specific
reporter gene, and the appearance of Canr Ade- papillae resulting from excision of an
engineered Ty element containing the HO-endonuclease target site and a SUP4o gene.  We found
that the replacement of the fourth point mutation in ho to the HO sequence restored HO
activity to the chimeric endonuclease.

<>

<1>Meisel, A., Bickle, T.A., Kruger, D.H., Schroeder, C.
<2>Type III restriction enzymes need two inversely oriented recognition sites for DNA cleavage.
<3>Nature
<4>355
<5>467-469
<6>1992
<7>Type III restriction/modification enzymes recognize short, nonpalindromic
sequences that can be methylated on only one strand, with the paradoxical
consequence that during replication of what is in effect hemimethylated DNA
totally unmodified sites arise.  Why the unmodified sites are not subject to
suicidal restriction was not clear.  Here we show that restriction requires two
unmodified recognition sites that can be separated by different distances but
which must be in inverse orientation.  All of the unmodified sites in newly
replicated DNA are of course in the same orientation, which explains why they
are not restricted.  This result may be of relevance to other manifestations of
anisotropy in double-stranded DNA, such as genetic imprinting.

<>

<1>Meisel, A., Kruger, D.H., Bickle, T.A.
<2>M.EcoP15I methylates the second adenine in its recognition sequence.
<3>Nucleic Acids Res.
<4>19
<5>3997
<6>1991
<7>The type III restriction/modification system EcoP15I recognizes the
non-palindromic sequence 5'-CAGCAG-3'.  The restriction enzyme cleaves the DNA
25-27 bp to the right of the sequence as written.  The modification methylase
methylates one of the two adenine residues in the recognition sequence.
Attempts to determine which of the adenines is methylated by the enzyme were
foiled by the internal repeated symmetry of the recognition sequence.

<>

<1>Meisel, A., Mackeldanz, P., Bickle, T.A., Kruger, D.H., Schroeder, C.
<2>Type III restriction endonucleases translocate DNA in a reaction driven by recognition site-specific ATP hydrolysis.
<3>EMBO J.
<4>14
<5>2958-2966
<6>1995
<7>Type III restriction/modification systems recognize short non-palindromic sequences, only one
strand of which can be methylated. Replication of type III-modified DNA produces completely
unmethylated recognition sites which, according to classical mechanisms of restriction, should
be signals for restriction. We have shown previously that suicidal restriction by the type III
enzyme EcoP15I is prevented if all the unmodified sites are in the same orientation:
restriction by EcoP15I requires a pair of unmethylated, inversely oriented recognition sites.
We have now addressed the molecular mechanism of site orientation-specific DNA restriction.
EcoP15I is demonstrated to possess an intrinsic ATPase activity, the potential driving force
of DNA translocation. The ATPase activity is uniquely recognition site-specific, but
EcoP15I-modified sites also support the reaction. EcoP15I DNA restriction patterns are shown
to be predetermined by the enzyme-to-site ratio, in that site-saturating enzyme levels elicit
cleavage exclusively between the closest pair of head-to-head oriented sites. DNA restriction
is blocked by Lac repressor bound in the intervening sequence between the two EcoP15I sites.
These results rule out DNA looping and strongly suggest that cleavage is triggered by the
close proximity of two convergently tracking EcoP15I-DNA complexes.

<>

<1>Meister, G.E.
<2>Non-associating heterodimeric DNA methyltransferases as a platform for developing designer methyltransferases.
<3>Ph.D. Thesis, Johns Hopkins University, USA
<4>
<5>
<6>2009
<7>The ability to site-specifically methylate a unique DNA sequence within a genome has numerous
potential uses including 1) a tool for the study of DNA methylation patterns, 2) a tool to
silence genes of interest, and 3) a potential gene therapy device to correct conditions caused
by hypomethylation. Current approaches include linking methyltransferases to DNA binding
domains to localize enzymes next to a target site. This approach has achieved site-biased
methylation, however the engineered methyltransferases are still active in the absence of
binding their intended target and methylate non-target sites. We demonstrate an alternative
strategy in which fragments of a DNA methyltransferase, compromised in their ability to
methylate DNA, are fused to two zinc fingers designed to bind 9 bp sites flanking a
methylation site. Using the naturally heterodimeric methyltransferase M.EcoHK31I, we have
demonstrated that this strategy can yield a methyltransferase capable of a high level of
methylation at the target site with undetectable levels of methylation at the non-target sites
in E. coli. The two zinc fingers acted synergistically to target methylation to the desired
site. However, some nontarget methylation could be detected at higher expression levels of the
zinc fingermethyltransferase indicating that further improvements will be necessary to attain
the desired exclusive target specificity.

<>

<1>Meister, G.E., Chandrasegaran, S., Ostermeier, M.
<2>BIOT 149-Heterodimeric DNA methyltransferases.
<3>ACS Abstracts
<4>236
<5>0
<6>2008
<7>DNA methylation patterns play an important role in determining gene expression patterns. These
patterns are of particular interest in embryonic development and in cancer cells, which often
exhibit abnormal methylation patterns. The ability to control the activity and specificity of
DNA methyltransferases would have applications in the study of DNA methylation in cells, would
offer an avenue to control gene expression epigenetically and potentially would allow the
correction of abnormal methylation patterns for therapeutic purposes. Most known DNA
methyltransferases are encoded in a single polypeptide chain.  DNA methyltransferases in which
the activity is encoded by heterodimerizing peptides, offer unique platform for engineering
DNA methyltransferases with novel properties. The C5-methylcytosine methyltransferases M. AquI
and M. EcoHK31I each have alpha and beta peptide chains that associate to create a functional
enzyme. Methylation is not possible without the association of the two fragments.   Truncated
version of these fragments exhibit decreased association in vitro. We have used an in vivo
method to determine if the fragment's association is more or less sensitive to these
truncations in the cellular environment.  Select fragments formed the basis for designed
methyltransferase libraries that will methylate unique sites.

<>

<1>Meister, G.E., Chandrasegaran, S., Ostermeier, M.
<2>An engineered split M.Hhal-zinc finger fusion lacks the intended methyltransferase specificity.
<3>Biochem. Biophys. Res. Commun.
<4>377
<5>226-230
<6>2008
<7>The ability to site-specifically methylate DNA in vivo would have wide applicability to the
study of basic biomedical problems as well as
enable studies on the potential of site-specific DNA methylation as a
therapeutic strategy for the treatment of diseases. Natural DNA
methyltransferases lack the specificity required for these
applications. Nomura and Barbas [W. Nomura, C.F. Barbas 3rd, In vivo
site-specific DNA methylation with a designed sequence-enabled DNA
methylase, J. Am. Chem. Soc. 129 (2007) 8676-8677] have reported that
an engineered DNA methyltransferase comprised of fragments of M.Hhal
methyltransferase and zinc finger proteins has very high specificity
for the chosen target site. Our analysis of this engineered enzyme
shows that the fusion protein methylates target and non-target sites
with similar efficiency.

<>

<1>Meister, G.E., Chandrasegaran, S., Ostermeier, M.
<2>Heterodimeric DNA methyltransferases as a platform for creating designer zinc finger methyltransferases for targeted DNA methylation in cells.
<3>Nucleic Acids Res.
<4>38
<5>1749-1759
<6>2010
<7>The ability to target methylation to specific genomic sites would further the study of DNA
methylation's biological role and potentially offer a
tool for silencing gene expression and for treating diseases involving
abnormal hypomethylation. The end-to-end fusion of DNA methyltransferases
to zinc fingers has been shown to bias methylation to desired regions.
However, the strategy is inherently limited because the methyltransferase
domain remains active regardless of whether the zinc finger domain is
bound at its cognate site and can methylate non-target sites. We
demonstrate an alternative strategy in which fragments of a DNA
methyltransferase, compromised in their ability to methylate DNA, are
fused to two zinc fingers designed to bind 9 bp sites flanking a
methylation target site. Using the naturally heterodimeric DNA
methyltransferase M.EcoHK31I, which methylates the inner cytosine of
5'-YGGCCR-3', we demonstrate that this strategy can yield a
methyltransferase capable of significant levels of methylation at the
target site with undetectable levels of methylation at non-target sites in
Escherichia coli. However, some non-target methylation could be detected
at higher expression levels of the zinc finger methyltransferase
indicating that further improvements will be necessary to attain the
desired exclusive target specificity.

<>

<1>Meister, J., MacWilliams, M., Hubner, P., Jutte, H., Skrzypek, E., Piekarowicz, A., Bickle, T.A.
<2>Macroevolution by transposition: drastic modification of DNA recognition by the type I restriction enzyme following Tn5 transposition.
<3>EMBO J.
<4>12
<5>4585-4591
<6>1993
<7>We have characterized a novel mutant of EcoDXXI, a type IC DNA restriction and modification
(R-M) system, in which the specificity has been altered due to a Tn5 insertion into the middle
of hsdS, the gene which encodes the polypeptide that confers DNA sequence specificity to both
the restriction and the modification reactions. Like other type I enzymes, the wild type
EcoDXXI recognizes a sequence composed of two asymmetrical half sites separated by a spacer
region: TCA(N7)RTTC. Purification of the EcoDXXI mutant methylase and subsequent in vitro DNA
methylation assays identified the mutant recognition sequence as an interrupted palindrome,
TCA(N8)TGA, in which the 5' half site of the wild type site is repeated in inverse
orientation. The additional base pair in the non-specific spacer of the mutant recognition
sequence maintains the proper spacing between the two methylatable adenine groups. Sequencing
of both the wild type and mutant EcoDXXI hsdS genes showed that the Tn5 insertion occurred at
nucleotide 673 of the 1221 bp gene. This effectively deletes the entire carboxyl-terminal DNA
binding domain which recognizes the 3' half of the EcoDXXI binding site. The truncated hsdS
gene still encodes both the amino-terminal DNA binding domain and the conserved repeated
sequence that defines the length of the recognition site spacer region. We propose that the
EcoDXXI mutant methylase utilizes two truncated hsdS subunits to recognize its binding site.
The implications of this finding in terms of subunit interactions and the malleability of the
type I R-M systems will be discussed.

<>

<1>Mejean, A., Mazmouz, R., Mann, S., Calteau, A., Medigue, C., Ploux, O.
<2>The genome sequence of the cyanobacterium Oscillatoria sp. PCC 6506 reveals several gene clusters responsible for the biosynthesis of toxins  and secondary metabolites.
<3>J. Bacteriol.
<4>192
<5>5264-5265
<6>2010
<7>We report a draft sequence of the genome of Oscillatoria sp. PCC 6506, a cyanobacterium that
produces anatoxin-a and homoanatoxin-a, two
neurotoxins, and cylindrospermopsin, a cytotoxin. Beside the clusters of
genes responsible for the biosynthesis of these toxins, we have found
other clusters of genes likely involved in the biosynthesis of
not-yet-identified secondary metabolites.

<>

<1>Melcher, U., Fletcher, J.
<2>Inactivation of a Spiroplasma citri DNA modification methylase by a virus-like insertion element suggested by DNA sequence.
<3>J. Plant Pathol.
<4>82
<5>71
<6>2000
<7>A partial SpV1-like viral DNA sequence in the Spiroplasma citri chromosome, the D3 progenitor,
was previously implicated as the donor of viral DNA sequences causing recombinational
instability of insert-containing virus vectors.  Re-examination of the D3 progenitor sequence
suggested that the virus-like sequence had inserted into a DNA adenine modification methylase
gene, inactivating it.  Comparison of the D3 progenitor sequence with SpV1-C74 revealed that
it was more closely related to this virus than to SpV1-R8A2 B and that the point of insertion
corresponded to an inverted terminal repeat similar to those terminating elements of the IS3
family of insertion sequences.  The transposases most similar to the SpV1-C74 is probably an
encapsilated insertion element.

<>

<1>Melcher, U., Sha, Y., Ye, F., Fletcher, J.
<2>Mechanisms of Spiroplasma genome variation associated with SpV1-like viral DNA inferred from sequence comparisons.
<3>Microb. Comp. Genomics
<4>4
<5>29-46
<6>1999
<7>Genomes of Spiroplasma citri strains have rearranged frequently during their evolution, partly
due to multiple integrated sequences of spiroplasma viruses. To understand better the role of
viral sequences in genome evolution, we examined available nucleotide sequences of viruslike
elements in the S. citri chromosome. Comparison of integrated and nonintegrated sequences of
spiroplasma virus SpV1-C74 DNA suggested that it is an encapsidated form of the circular
transposition intermediate belonging to an insertion sequence (IS3) family member. One
SpV1-C74 viral DNA fragment was identified as interrupting the remains of a DNA adenine
modification methylase gene. A viral DNA insertion of SpV1-R8A2 B DNA had hallmarks of having
suffered an internal deletion by a site-specific recombination system. Homologous
recombination likely was responsible for several deletions within viral DNA. A homologous
recombination event was inferred between part of a viral DNA insertion and a similar
chromosomal sequence. Dispersed sequences from SpV1-like C4 open reading frames (ORFs) were
identified as involved in a complex deletion-inversion event. Thus, SpV1-like sequences likely
have altered spiroplasma genomes by inserting within active genes, destroying their function,
by providing targets for site-specific recombination, by mediating deletions of sequences
adjacent to their integration sites, and by providing targets for homologous recombination,
leading to inversions.

<>

<1>Mell, J.C., Sinha, S., Balashov, S., Viadas, C., Grassa, C.J., Ehrlich, G.D., Nislow, C., Redfield, R.J., Garmendia, J.
<2>Complete Genome Sequence of Haemophilus influenzae Strain 375 from the Middle Ear of a Pediatric Patient with Otitis Media.
<3>Genome Announcements
<4>2
<5>e01245-14
<6>2014
<7>Originally isolated from a pediatric patient with otitis media, Haemophilus influenzae strain
375 (Hi375) has been extensively studied as a model system for
intracellular invasion of airway epithelial cells and other pathogenesis traits.
Here, we report its complete genome sequence and methylome.

<>

<1>Melnik, A.I., Rebentish, B.A., Bolotin, A.V., Mendzhul, M.I.
<2>Two site-specific endonucleases of the Cyanobacterium Nostoc linckia.
<3>Mikrobiol. Zh.
<4>53
<5>24-28
<6>1991
<7>Two restrictases Nli387/7I and Nli387/7II have been isolated from
cyanobacterium Nostoc linckia using chromatography on phosphocellulose, <Mono
Q> column, and heparin sepharose 4B.  The preparations are described by the
method of electrophoresis in polyacrylamide gels under denaturating conditions.
The catalytic properties of the restrictases have been determined:  optimal
pH 9.0 - 9.5, optimal concentration of Na+ - 5 mM, that of Mg2+ - 6 mM, optimal
temperature - 37C.  The isolated enzymes are isoschizomers of the restrictases
AvaI and AvaII.  The point of cutting is determined for enzyme Nli387/7 I.  It
is shown that restrictase Nli387/7 I is a false isoschizomer of AvaI.

<>

<1>Mendez-Tenorio, A., Larios-Serrato, V., Olguin-Ruiz, G.E., Sanchez-Vallejo, C.J., Torres-Lopez, R.C., Aviles-Jimenez, F., Camorlinga-Ponce, M., Torres, J.
<2>Genome Sequence of a Helicobacter pylori Strain Isolated from a Mexican Patient with Intestinal Gastric Cancer.
<3>Genome Announcements
<4>2
<5>e01214-13
<6>2014
<7>Helicobacter pylori strains are the major risk factor for gastric cancer. Strains vary in
their content of disease-associated genes, so genome-wide analysis of
cancer-isolated strains will help elucidate their pathogenesis and genetic
diversity. We present the draft genome sequence of H. pylori isolated from a
Mexican patient with intestinal gastric cancer.

<>

<1>Mendzhul, M.I., Moshinsky, I.D., Syrchin, S.A.
<2>Endonuclease activity in cyanobacteria Anabaena variabilis and Plectonema boryanum involving restriction endonuclease and nuclease.
<3>Mikrobiol. Zh.
<4>61
<5>10-14
<6>1999
<7>Site-specific endodeoxyribonuclease activity was found in crude cell-free extracts of
cyanobacteria Plectonema boryanum CALU 465 and
Plectonema edaphycum CALU 262. Cyanobacterium Anabaena variabilis CALU
458 was shown to possess restriction site-specific endonuclease Ava458I
which was probably an isoschizomer of CfrI with the recognition site
PyGGCCPu.

<>

<1>Mendzhul, M.I., Syrchin, S.A., Averkiev, A.A., Rebentish, B.A.
<2>The way of defense of cyanophage LPP-3 DNA against restriction-modification systems in the cells of the cyanobacterium Plectonema boryanum.
<3>Biopol. Kletka
<4>9
<5>54-61
<6>1993
<7>The HPLC method was used to study some peculiarities of the DNA composition of the
cyanobacterium Plectonema boryanum and cyanophage LPP-3. Analysis of the elution profile of
the products of DNA acid hydrolysis allow the identification in P. boryanum DNA, in addition
to the canonical bases, of the presence of 4.5% N-6-methyladenine and 1.2% 5-methyl-cytosine;
DNA from LPP-3 has 0.8% 5-methycytosine and no N-6-methyladenine. The presence of
5-methylcytosine was detected only by the application of a modified hydrolysis method using
HF. Restriction endonuclease isoschizomers, whose hydrolysis of DNA depends on the presence of
methylated bases in the recognition sites, were used to detect site-specific methylation. The
comparison of the products of the DNA LPP-3 and P. boryanum fermentative hydrolysis by MspI
and HpaII; Sau3AI, MboI and DpnI; Apyl and MvaI restrictases enabled the determination that
DNA from LPP-3 and P. boryanum has a high degree of methylation of the inner cytosine in
CC(A/T)GG sequences and in P. boryanum the adenine in the GATC site; CCGG sites were not
methylated in either DNA. It is concluded that dam- and dcm-like modification systems are in
P. boryanum. The defense of viral DNA against host R-M systems occurs by cytosine methylation
in the sequence CmC(A/T)GG and counter-selection at BamHI sites.

<>

<1>Mendzhul, M.I., Syrchin, S.A., Rebentish, B.A., Averkiev, A.A., Busakhina, I.V.
<2>The resistance of the DNA of cyanophage LPP-3 to the action of different restriction endonucleases.
<3>Mikrobiol. Zh.
<4>55
<5>47-53
<6>1993
<7>Data on the study of structural peculiarities of cyanophage LPP-3 DNA are presented in the
work. The length of cyanophage DNA calculated by means of the enzymatic hydrolysis by
restrictases is 40+/-3.5 thousand base pairs. Cyanophage LPP-3 DNA was hydrolysed by more than
50 different restrictases. As a result of screening it was found out that the great number of
restrictases, which recognized hexanucleotide sequences did not hydrolyse DNA of cyanophage
LPP-3. A considerable deviation of the number of the observed sites of restriction from their
theoretically expected number for restrictases, which recognized hexanucleotide sequences did
not hydrolyse DNA of cyanophage LPP-3. A considerable deviation of the number of the observed
sites of restriction from their theoretically expected number for restrictases HaeIII and
Cfr131 was established. Restrictases-isoschizomers with different sensitivity to the
methylation of the recognition sites -- MspI, HpaII and Sau3A, MboI and DpnI were used to
check the availability of methylated bases in LPP-3 DNA. Absence of methylated adenine in the
site GATC and methylated cytosine in the second position of the site CCGG were established.
The results obtained permit supposing that the expressed counterselection by the sites of
recognition of many restriction endonucleases takes place in cyanophage LPP-3 DNA. It is
supposed that apparently, this method of protection of its genome in LPP-3 is one of most
important but the inconsiderable percentage of site-specific methylation of the virus DNA
cannot be completely excluded.

<>

<1>Menendez, C., Bournat, J.C., Ramirez, J.L.
<2>Cloning and overexpression of EcoRI methylase.
<3>Interciencia
<4>14
<5>37-40
<6>1989
<7>The author cloned a HindIII fragment of pMB3 plasmid containing part of the coding region for
EcoRI endonuclease, and the entire sequence for EcoRI methylase downstream of the PL promoter
of the expression vector pcp 40.  The recombinant was introduced into a lysogenic E. coli
strain, harboring a lambda phage with a temperature sensitive mutation for the repressor gene
(c1857). After heat induction, a protein band of apparent molecular mass of 39 Kd, increased
continuously during 8 hours.  After four hours of thermal induction, the band represented
nearly 50% of the soluble proteins observed by gel electrophoresis.  At the maximum induction
time, the methylase activity registered a 15 fold increment over the non-induced state.

<>

<1>Meng, X., Cai, W., Schwartz, D.C.
<2>Inhibition of restriction endonuclease activity by DNA binding fluorochromes.
<3>J. Biomol. Struct. Dyn.
<4>13
<5>945-951
<6>1996
<7>Activity of type II restriction endonucleases is affected by many common factors including
buffer composition and sequences flanking the recognition site.  The successful development of
Optical Mapping relied on optimization of light microscope-based imaging of fluorescently
labeled DNA molecules during restriction endonuclease digestion.  Little was known about the
effects of commonly used DNA-fluorochromes on restriction endonuclease activity.  Thus, we
developed an enzyme activity assay using lambda bacteriophage DNA or adenovirus-2 DNA to
evaluate the effects of five DNA binding fluorochromes (4'-6-daimidine-2-phenylindole (DAPI),
ethidium bromide (EtdBr), ethidium bromide homodimer (EthD-1), bis-benzimide (H33258) and
benzothiazolium-4-quinolinium dimer (TOTO-1)) on the enzymatic activities of eleven type II
restriction endonucleases (AscI, CspI, DraI, EcoRI, HhaI, HindIII, NotI, RsrII, SfiI, SgrAI
and SmaI).  We found that the minor groove binding fluorochrome, DAPI, did not measurably
inhibit activity of this group, with the exception of DraI.  Similarly, another minor groove
binding fluorochrome H33258 inhibited DraI and NotI (slightly).  The three intercalating
fluorochromes EtdBr, EthD-1 and TOTO-1, however, variably inhibited the other enzymes.  Since
beta-mercaptoethanol (BME) is used to discourage photodamage of stained DNA molecules, we also
assessed its effect on restriction endonuclease activity.  Interestingly, DraI, EcoRI, HhaI,
HindIII, SfiI and SmaI retained full activities at high concentration of BME (5%), but AscI,
CspI, NotI, RsrII and SgrAI showed varying sensitivities to the BME.  Isoschizomers CspI and
RsrII behaved differently to both fluorochromes and BME.  The results presented here should
provide a basis for further development of new Optical Mapping-based techniques requiring
fluorescence labeling of other actively imaged enzymatic reactions.

<>

<1>Meramveliotaki, C., Kotsifaki, D., Androulaki, M., Hountas, A., Eliopoulos, E., Kokkinidis, M.
<2>Purification, crystallization, x-ray diffraction analysis and phasing of an engineered single-chain PvuII restriction endonuclease.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>63
<5>836-838
<6>2007
<7>The restriction endonuclease PvuII from Proteus vulgaris has been converted from its wild-type
homodimeric form into the enzymatically
active single-chain variant scPvuII by tandemly joining the two
subunits through the peptide linker Gly-Ser-Gly-Gly. scPvuII, which is
suitable for the development of programmed restriction endonucleases
for highly specific DNA cleavage, was purified and crystallized. The
crystals diffract to a resolution of 2.35 angstrom and belong to space
group P4(2), with unit-cell parameters a = b = 101.92, c = 100.28
angstrom and two molecules per asymmetric unit. Phasing was
successfully performed by molecular replacement.

<>

<1>Merga, J.Y., Winstanley, C., Williams, N.J., Yee, E., Miller, W.G.
<2>Complete Genome Sequence of the Arcobacter butzleri Cattle Isolate 7h1h.
<3>Genome Announcements
<4>1
<5>e00655-13
<6>2013
<7>Arcobacter butzleri strain 7h1h was isolated in the United Kingdom from the feces of a
clinically healthy dairy cow. The genome of this isolate was sequenced to
completion. Here, we present the annotation and analysis of the completed 7h1h
genome, along with a comparison of this genome to the existing A. butzleri
genomes.

<>

<1>Merhej, V., Armougom, F., Robert, C., Raoult, D.
<2>Genome Sequence of Lactobacillus ingluviei, a Bacterium Associated with Weight Gain in Animals.
<3>J. Bacteriol.
<4>194
<5>5697
<6>2012
<7>We report the draft genome sequence of Lactobacillus ingluviei strain Autruche 4  (CSURP209)
isolated from an ostrich. L. ingluviei is associated with weight gain
in mice. This genome sequence may help us understand the obesity-induced
mechanisms of intestinal bacteria.

<>

<1>Merhej, V., Croce, O., Robert, C., Rolain, J.M., Raoult, D.
<2>Genome Sequence of Bartonella rattaustraliani, a Bacterium Isolated from an Australian Rat.
<3>J. Bacteriol.
<4>194
<5>7012
<6>2012
<7>Bartonella rattaustraliani is a facultative intracellular bacterium isolated from the blood of
a Rattus sp. in Australia. The present study reports the draft
genome of B. rattaustraliani strain AUST/NH4 (CSUR B609(T)).

<>

<1>Merhej, V., Croce, O., Robert, C., Rolain, J.M., Raoult, D.
<2>Genome Sequence of Bartonella rattimassiliensis, a Bacterium Isolated from European Rattus norvegicus.
<3>J. Bacteriol.
<4>194
<5>7013
<6>2012
<7>Bartonella rattimassiliensis is a facultative intracellular bacterium isolated from the blood
of Rattus norvegicus in Marseille. The present study reports the
draft genome of B. rattimassiliensis strain 15908 (CIP 107705(T)).

<>

<1>Merino, A., Gamba, G.
<2>Biologia molecular en medicina.  II. Enzimas de restriccion.
<3>Rev. Invest. Clin.
<4>48
<5>159-163
<6>1996
<7>
<>

<1>Merino, M., Alvarez-Fraga, L., Gomez, M.J., Aransay, A.M., Lavin, J.L., Chaves, F., Bou, G., Poza, M.
<2>Complete Genome Sequence of the Multiresistant Acinetobacter baumannii Strain AbH12O-A2, Isolated during a Large Outbreak in Spain.
<3>Genome Announcements
<4>2
<5>e01182-14
<6>2014
<7>We report the complete genome sequence of Acinetobacter baumannii strain AbH12O-A2, isolated
during a large outbreak in Spain. The genome has 3,875,775 bp
and 3,526 coding sequences, with 39.4% G+C content. The availability of this
genome will facilitate the study of the pathogenicity of the Acinetobacter
species.

<>

<1>Merkiene, E., Klimasauskas, S.
<2>Probing a rate-limiting step by mutational perturbation of AdoMet binding in the HhaI methyltransferase.
<3>Nucleic Acids Res.
<4>33
<5>307-315
<6>2005
<7>DNA methylation plays important roles via regulation of numerous cellular mechanisms in
diverse organisms, including humans. The paradigm bacterial methyltransferase (MTase) HhaI
(M.HhaI) catalyzes the transfer of a methyl group from the cofactor S-adenosyl-L-methionine
(AdoMet) onto the target cytosine in DNA, yielding 5-methylcytosine and
S-adenosyl-L-homocysteine (AdoHcy). The turnover rate (kcat) of M.HhaI, and the other two
cytosine-5 MTases examined, is limited by a step subsequent to methyl transfer; however, no
such step has so far been identified. To elucidate the role of cofactor interactions during
catalysis, eight mutants of Trp41, which is located in the cofactor binding pocket, were
constructed and characterized. The mutants show full proficiency in DNA binding and
base-flipping, and little variation is observed in the apparent methyl transfer rate kchem as
determined by rapid-quench experiments using immobilized fluorescent-labeled DNA. However, the
Trp41 replacements with short side chains substantially perturb cofactor binding (100-fold
higher KD(AdoMet)and KM(AdoMet))leading to a faster turnover of the enzyme (10-fold higher
kcat). Our analysis indicates that the rate-limiting breakdown of a long-lived ternary product
complex is initiated by the dissociation of AdoHcy or the opening of the catalytic loop in the
enzyme.

<>

<1>Merkiene, E., Lukinavicius, G., Klimasauskas, S.
<2>Synergy of substrate binding, base flipping and catalytic loop motions in a DNA methyltransferase.
<3>Eur. Biophys. J.
<4>34
<5>617
<6>2005
<7>The HhaI methyltransferase transfers a methyl group from cofactor AdoMet onto its target
cytosine residue in DNA.  Crystal structures revealed that M.HhaI flips its target base out of
the DNA helix.  This transition is accompanied by a massive motion of a loop in the protein
which locks the flipped out base in the catalytic site.  We used fluorescence of a unique
tryptophan residue to specifically monitor cofactor binding.  Equilibrium binding studies
revealed a highly improved binding of the cofactor AdoMet and the reaction product AdoHcy in
the presence of specific DNA.  No such effect was observed with non-specific DNA in the case
of AdoMet, but surprisingly, led to a substantial drop in binding affinity in the case of
AdoHcy!  To elucidate the role of the catalytic loop in substrate binding we constructed two
variants of M.HhaI in which large segments of the catalytic loop were entirely removed.
Although the binary interactions with the substrates and base flipping was almost unaffected
by the deletions, the synergy of substrate binding in the ternary complexes were completely
lost.  To 'visualize' the loop motions directly during the reaction cycle we prepared a
series of double mutants in which a unique tryptophan residue was placed in selected positions
on the mobile catalytic loop.  Single turnover stopped flow kinetic studies of the mutants
revealed two conformational transitions of the loop which coincide with the formation of the
tight ternary complex and the release of products.

<>

<1>Merkiene, E., Vilkaitis, G., Klimasauskas, S.
<2>Coexistence of single-strand and double-strand DNA cytosine-N4-methyltransferases in the BcnI restriction-modification system.
<3>Biologija
<4>1
<5>5-8
<6>1997
<7>Sequence analysis of the BcnI RM system, besides the previously characterized restriction
endonuclease (bcnIR) and cytosine-N4-methyltransferase (bcnIB), revealed the presence of a
large open reading frame potentially encoding a second cytosine-N4-methyltransferase (bcnIA).
The bcnIA gene, when subcloned in E. coli on the pUC19 vector, rendered protection of the
5'CC(C/G)GG-3' sites against cleavage by the cognate BcnI endonuclease.  The two
methyltransferases were partially purified, and their activities in vitro were compared using
various DNA substrates.  Both enzymes act on 5'-CC(C/G)GG-3' sites in double-stranded DNA,
however, BcnIA can also, at a comparable rate, modify the specific targets in single-stranded
DNA.  Biological significance of the presence of two distinct methyltransferases in the BcnI
RM system is discussed.

<>

<1>Merkiene, E., Vilkaitis, G., Klimasauskas, S.
<2>A pair of single-strand and double-strand DNA cytosine-N4 methyltransferases from Bacillus centrosporus.
<3>Biol. Chem.
<4>379
<5>569-571
<6>1998
<7>Sequence analysis of the BcnI restriction-modification system revealed the presence of an open
reading frame encoding a second cytosine-N4 methyltransferase, M.BcnIA, in the vicinity of the
genes specifying the previously characterized cytosine-N4 methyltransferase M.BcnIB and
restriction endonuclease R.BcnI.  Both methyltransferases were purified from the E. coli cells
expressing the individual genes, and their enzymatic efficiencies in vitro were compared with
a variety of DNA substrates.  Both enzymes act on 5'-CC(C/G)GG-3' sites in double-stranded
DNA, however, M.BcnIA can also, with a comparable efficiency, modify the specific targets in
single-stranded DNA.  The biological significance of the presence of the tandem
methyltransferases in the BcnI system is discussed.

<>

<1>Merkiene, E., Weinhold, E., Klimasauskas, S.
<2>Kinetics of cofactor binding and catalytic loop movements of HhaI methyltransferase.
<3>Biochem. Soc. Trans.
<4>28
<5>A464
<6>2000
<7>HhaI methyltransferase (M.HhaI) flips its target cytosine out of the DNA helix and into a
catalytic site in the enzyme.  This conformational transition of bound DNA is accompanied by a
large movement of the catalytic loop (over 20 Angstrom) in the enzyme itself.  Our studies
concentrated on the motions of the catalytic loop and interaction between M.HhaI and its
cofactor S-adenosyl-L-methionine (AdoMet) which serves as the methyl group donor in the MTase
reaction.  We found that the intrinsic fluorescence of a unique tryptophan residue (W41) is
significantly quenched upon binding of cofactor, but is unaffected upon binding of DNA.  A
series of kinetic association and equilibrium binding studies in the presence of various DNA
substrates permitted direct determination of kinetic parameters for cofactor binding.  We find
that non-specific DNA causes a substantial decrease in the affinity of the enzyme toward the
product AdoHcy but not toward cofactor AdoMet, which suggests a mechanism for cofactor
reloading during diffusion of MTase over regions of non-specific DNA.  Single-turnover
experiments revealed a concentration independent transient (2 s^-1) which we tentatively
attribute to the aforementioned conformational rearrangement of the catalytic loop.  To
'visualize' the loop motions directly we prepared a series of double mutants in which the
unique Trp was placed in selected positions on the mobile catalytic loop.  Preliminary
characterization of the mutant proteins indicates that they are suitable for direct kinetic
measurement of conformational transitions in the catalytic loop.  Kinetic analysis of these
transitions using global fitting and simulation is now underway.

<>

<1>Merkl, R., Fritz, H.J.
<2>Statistical evidence for a biochemical pathway of natural, sequence-targeted G/C to C/G transversion mutagenesis in Haemophilus influenzae Rd.
<3>Nucleic Acids Res.
<4>24
<5>4146-4151
<6>1996
<7>Markov chain analysis of the Haemophilus influenzae Rd genome reveals striking
under-representation of three palindromic tetranucleotide strings (CCGG, GGCC and CATG),
accompanied by over-representation of six tetranucleotide strings that are derived from the
former by exchanging strand location of the two residues making up a G/C nucleotide pair at
the terminal palindrome position.  Constraints are outlined for a molecular model able to
explain the phenomenon as the result of sequence-targeted, enzyme-driven G/C to C/G
transversion mutagenesis.  Possible participation in the process by components of known DNA
mismatch repair or restriction/modification systems (in particular, cytosine methylation) is
discussed.  The effect widens the spectrum of enzyme-driven, specific mutagenesis beyond the
formerly described C/G to T/A transition (VSP repair of Escherichia coli).  Potential
evolutionary benefits of enzymatic pathways of specific mutagenesis can be envisioned.

<>

<1>Merkl, R., Kroeger, M., Rice, P., Fritz, H.-J.
<2>Statistical evaluation and biological interpretation of non-random abundance in the E. coli K-12 genome of tetra- and pentanucleotide sequences related to VSP DNA mismatch repair.
<3>Nucleic Acids Res.
<4>20
<5>1657-1662
<6>1992
<7>The abundance of all tetra- and pentanucleotide sequences is calculated for a set of DNA
sequence data comprising 767,393 nucleotides of the E. coli K-12 genome.  Observed frequencies
are compared to those expected from a Markov chain prediction algorithm.  Systematic and
extreme non-random representations are found for special sets of sequences.  These are
interpreted as arising from incorporation of a 2'-deoxyguanosine residue opposite thymidine
during replication which, in special sequence contexts, leads to a T/G mismatch that is
simultaneously substrate for two competing DNA mismatch repair systems: the mutHLS and the VSP
pathway.  Processing by the former leads to error correction, by the latter to mutation
fixation.  The significance of the latter process, as demonstrated here, makes it unlikely
that VSP repair has evolved mainly as a mutation avoidance mechanism.  It is proposed that in
E. coli K-12, VSP repair, together with DNA cytosine methylation, constitutes a
mutagenesis/recombination system capable of promoting gene-conversion-like unidirectional
transfer of short stretches of DNA sequence.

<>

<1>Merlo, D.J., Thompson, D.V.
<2>In vitro sodium bisulfite mutagenesis of restriction endonuclease recognition sites.
<3>Anal. Biochem.
<4>163
<5>79-87
<6>1987
<7>Sodium bisulfite treatment of single-stranded DNA deaminates exposed cytosine
residues to form uracil, resulting in cytosine-to-thymidine transition
mutations following DNA replication.  We have used this reaction in vitro to
destroy the recognition sequences for the restriction endonucleases HindIII and
XmaI in the aminoglycoside 3'-phosphotransferase I coding region of plasmid
pUC4K.  This procedure should be applicable to the mutation of any recognition
sequence of restriction endonucleases which generate cytosine-containing
single-stranded ends.  The possibility of mutagenesis of restriction sites to
generate stop codons in coding regions is discussed.

<>

<1>Mermelstein, L.D., Popoutsakis, E.T.
<2>In vivo methylation in Escherichia coli by the Bacillus subtilis phage Phi3T methyltransferase to protect plasmids from restriction upon transformation of Clostridium acetobutylicum ATCC 824.
<3>Appl. Environ. Microbiol.
<4>59
<5>1077-1081
<6>1993
<7>The restriction endonuclease Cac824I has been shown to be a major barrier to
electrotransformation of Clostridium acetobutylicum ATCC 824 (L.D. Mermelstein, N.E. Welker,
G.N. Bennett, and E.T. Popoutsakis, BioTechnology 10:190-195, 1992). Methylation by the Phi3T
I methyltransferase encoded by Bacillus subtilis phage Phi3T was shown to protect plasmid DNA
from restriction by Cac824I. Expression in Escherichia coli of the phi3tI gene (which encodes
the Phi3TI methyltransferase) from pAN1, which replicates via the p15A origin of replication,
was sufficient to completely methylate coresident E. coli-C. acetobutylicum shuttle vectors
with ColE1 origins of replication. Three shuttle vectors (pIMP1, pSYL2, and pSYL7) methylated
in this manner were used to efficiently electrotransform strain ATCC 824. These vectors could
not be introduced into strain ATCC 824 when unmethylated because the E. coli portions of the
plasmids contain a large number of Cac824I sites. This method obviates the need to use B.
subtilis-C. acetobutylicum shuttle vectors with few Cac824I sites to introduce DNA into C.
acetobutylicum ATCC 824.

<>

<1>Mermelstein, L.D., Welker, N.E., Bennett, G.N., Papoutsakis, E.T.
<2>Expression of cloned homologous genes in Clostridium acetobutylicum ATCC 824.
<3>Biotechnology
<4>10
<5>190-195
<6>1992
<7>We have previously cloned the acetone-formation pathway gene, encoding acetoacetate
deacarboxylase (adc), and butyrate-formation pathway gene, encoding phosphtransbutyrylase
(ptb), of Clostridium acetobutylicum ATCC 824 in Escherichia coli. Here we report their
subcloning in Bacillus subtilis and transfer to strain ATCC 824 via electrotransformation,
where the corresponding enzyme activities were expressed at elevated levels, using pFNK1, a
new B. subtilis/C. acetobutylicum shuttle vector. Plasmid pFNK1 was used because shuttle
vectors that function in E. coli were unable to electrotransform ATCC 824 unless they became
deleted in the E. coli-plasmid regions. The difficulties with shuttle vectors that function in
E. coli are probably due to the presence of a restriction endonuclease in ATCC 824. This
endonuclease recognizes the sequence 5'-GCNGC-3', which is prevalent in E. coli plasmids,
but occurs infrequently in pFNK1 and C. acetobutylicum genes. Cloning of genes in C.
acetobutylicum is critical for redirecting the cellular metabolism (metabolic engineering) as
well as for genetic studies of this industrial organism.

<>

<1>Mernagh, D., Marks, P., Kneale, G.
<2>AhdI, a new class of restriction-modification system?
<3>Biochem. Soc. Trans.
<4>27
<5>A126
<6>1999
<7>Bacterial restriction-modification systems have traditionally been divided into three distinct
classes, types I, II and III.  Here we report a study of what appears to be a new class of R-M
systems, exemplified by AhdI.  Analysis of the DNA sequence and gene organization of AhdI has
revealed that the methyltransferase more closely resembles a type I MTase than a type II.  In
contrast, the AhdI endonuclease is more characteristic of a type II R-M system.  It is
possible that the AhdI R-M system is an evolutionary intermediate made up of a type II-like
endonuclease and a methyltransferase that resembles those found in the multisubunit type I
systems.  This new class of R-M system has been named type 1-1/2.  The AhdI methyltransferase
is a multisubunit enzyme containing two subunits, one that shares some degree of sequence
homology with the type I HsdM subunit and the other with the HsdS subunit found in type I
MTases.  Whereas the M subunit of AhdI closely resembles a type I HsdM subunit in the sequence
and size of the protein, the S subunit is only half the size of the type I counterpart.  We
report here the cloning and characterization of the AhdI methyltransferase.  With experimental
evidence obtained from gel electrophoresis and analytical gel filtration we have determined
the stoichiometry of the enzyme and with the use of gel retardation and surface plasmon
resonance we will address the relationship of the subunit composition to DNA binding by the
AhdI MTase.

<>

<1>Mernagh, D.R., Janscak, P., Firman, K., Kneale, G.G.
<2>Protein-protein and protein-DNA interactions in the type I restriction endonuclease R.EcoR124I.
<3>Biol. Chem.
<4>379
<5>497-503
<6>1998
<7>The type I restriction-modification system EcoR124I recognizes and binds to the split DNA
recognition sequence 5'-GAAN6RTCG-3'.  The methyltransferase, consisting of HsdM and HsdS
subunits with the composition M2S, can interact with one or more subunits of the HsdR subunit
to form the endonuclease.  The interaction of the methyltransferase with HsdR has been
investigated by surface plasmon resonance, showing that there are two non-equivalent binding
sites for hsdR which differ in binding affinity by at least two orders of magnitude.  DNA
footprinting experiments using exonuclease III suggest that the addition of HsdR to the
methyltransferase (at a stoichiometry of either 1:1 or 2:1) increases the stability of the
resulting DNA-protein complex but does not increase the size of the footprint.  More extensive
in situ footprinting experiments using copper-phenanthroline on the DNA-protein complexes
formed by M2S, R1M2S and R2M2S also show no difference in the detailed cleavage pattern, with
~18 nucleotides protected on both strands in each complex.  Thus the HsdR subunit(s) of the
endonuclease stabilize the interaction of the M2S complex with DNA, but do not directly
contribute to DNA binding.  In addition, the thymidine nucleotide in the tetranucleotide
recognition sequence GTCG is hyper-reactive to cleavage in each case, suggesting that the DNA
structure in this region is altered in these complexes.

<>

<1>Mernagh, D.R., Kneale, G.G.
<2>High resolution footprinting of a type I methyltransferase reveals a large structural distortion within the DNA recognition site.
<3>Nucleic Acids Res.
<4>24
<5>4853-4858
<6>1996
<7>The type I DNA methyltransferase M.EcoR124I is a multi-subunit enzyme that binds to the
sequence GAAN6RTCG, transferring a methyl group from S-adenosylmethionine to a specific
adenine on each DNA strand.  We have investigated the protein-DNA interactions in the complex
by DNase I and hydroxyl radical footprinting.  The DNase I footprint is unusually large: the
protein protects the DNA on both strands for at least two complete turns of the helix,
indicating that the enzyme completely encloses the DNA in the complex.  The higher resolution
hydroxyl radical probe shows a smaller, but still extensive, 18 bp footprint encompassing the
recognition site.  Within this region, however, there is a remarkably hyper-reactive site on
each strand.  The two sites of enhanced cleavage are co-incident with the two adenines that
are the target bases for methylation, showing that the DNA is both accessible and highly
distorted at these sites.  The hydroxyl radical footprint is unaffected by the presence of the
cofactor S-adenosylmethionine, showing that the distorted DNA structure induced by M.EcoR124I
is formed during the initial DNA binding reaction and not as a transient intermediate in the
reaction pathway.

<>

<1>Mernagh, D.R., Reynolds, L.A., Kneale, G.G.
<2>DNA binding and subunit interactions in the type I methyltransferase M.EcoR124I.
<3>Nucleic Acids Res.
<4>25
<5>987-991
<6>1997
<7>The type I DNA methyltransferase M.EcoR124I consists of two methylation subunits (hsdM) and
one DNA recognition subunit (HsdS).  When expressed independently, HsdS is insoluble, but this
subunit can be obtained in soluble form as a GST fusion protein.  We show that the HsdS
subunit, even as a fusion protein, is unable to form a discrete complex with its DNA
recognition sequence.  When HsdM is added to the HsdS fusion protein, discrete complexes are
formed but these are unable to methylate DNA.  The two complexes formed correspond to species
with one or two copies of the HsdM subunit, indicating that blocking the N-terminus of hsdS
affects one of the HsdM binding sites.  However, removal of the GST moiety from such complexes
results in tight and specific DNA binding and restores full methylation activity.  The results
clearly demonstrate the importance of the HsdM subunit for DNA binding, in addition to its
catalytic role in the methyltransferase reaction.

<>

<1>Mernagh, D.R., Taylor, I.A., Kneale, G.G.
<2>Interaction of the type I methyltransferase M.EcoR124I with modified DNA substrates: sequence discrimination and base flipping.
<3>Biochem. J.
<4>336
<5>719-725
<6>1998
<7>We have analyzed the DNA-protein contacts made between the type I DNA methyltransferase
M.EcoR124I and its recognition sequence.  The effects of base modifications have been probed
by measuring the affinity of M.EcoR124I for the modified sequences relative to that for the
wild-type sequence by using gel-retardation competition assays.  These results, along with
those from methylation interference footprinting and photo-affinity cross-linking have
identified the location of potential DNA contacts within the DNA recognition site.
Substitution of 6-thioguanosine for each of the three specific guanines in the recognition
sequence leads to a large (10-20-fold) decrease in the strength of DNA binding, indicating the
importance of hydrogen-bonding interactions in the major groove of DNA.  In contrast,
replacement of either (or both) of the adenine at the target site for methylation by the
enzyme, to produce either a base pair mismatch or loss of the base, leads to a marked increase
in DNA-binding affinity.  The results strongly support the proposal that type I
methyltransferases employ a base-flipping mechanism to methylate their target base.

<>

<1>Merritt, J., Qi, F.X., Shi, W.Y.
<2>A unique nine-gene comY operon in Streptococcus mutans.
<3>Microbiology
<4>151
<5>157-166
<6>2005
<7>Many Gram-positive and Gram-negative bacteria possess natural competence mechanisms for DNA
capture and internalization. In Bacillus
subtilis, natural competence is absolutely dependent upon the presence
of a seven-gene operon known as the comG operon (comGA-G). In species
of Streptococcus, this function has been described for a four-gene
operon (comYA-D in Streptococcus gordonii and cglA-D in Streptococcus
pneumoniae). In this study, a nine-orf operon (named comYA-I) required
for natural competence in Streptococcus mutans was identified and
characterized. Orf analysis of this operon indicates that the first
four Orfs (ComYA-D) share strong homology with ComYA-D of S. gordonii
and CgIA-D of S. pneumoniae, the fifth to seventh Orfs (ComYE-G) match
conserved hypothetical proteins from various species of Streptococcus
with ComYF possessing a predicted ComGF domain, the eighth Orf (ComYH)
shows a strong homology to numerous DNA methyltransferases from
restriction/modification systems, and the ninth Orf (ComYI) is
homologous to acetate kinase (AckA). RT-PCR analysis of the orf
junctions confirmed that all nine orfs were present in a single
transcript, while real-time RT-PCR analysis demonstrated that these
orfs were expressed at a level very similar to that of the first orf in
the operon. Mutations were constructed in all nine putative orfs. The
first seven genes (comYA-G) were found to be essential for natural
competence, while comYH and comYI had reduced and normal natural
competence ability, respectively. Analyses of S. mutans comY-luciferase
reporter fusions indicated that comY expression is growth-phase
dependent, with maximal expression at an OD600 of about 0(.)2, while
mutations in ciaH, comC and luxS reduced the level of comY expression.
In addition, comY operon expression appears to be correlated with
natural competence ability.

<>

<1>Mertineit, C., Yoder, J.A., Taketo, T., Laird, D.W., Trasler, J.M., Bestor, T.H.
<2>Sex-specific exons control DNA methyltransferase in mammalian germ cells.
<3>Development
<4>125
<5>889-897
<6>1998
<7>The spermatozoon and oocyte genomes bear sex-specific methylation patterns that are
established during gametogenesis and are required for the allele-specific expression of
imprinted genes in somatic tissues.  The mRNA for Dnmt1, the predominant maintenance and de
novo DNA (cytosine-5)-methyl transferase in mammals, is present at high levels in postmitotic
murine germ cells but undergoes alternative splicing of sex-specific 5' exons, which controls
the production and localization of enzyme during specific stages of gametogenesis.  An
oocyte-specific 5' exon is associated with the production of very large amounts of active
Dnmt1 protein, which is truncated at the N terminus and sequestered in the cytoplasm during
the later stages of oocyte growth, while a spermatocyte-specific 5' exon interferes with
translation and prevents production of Dnmt1 during the prolonged crossing-over stages of male
meiosis.  During the course of postnatal oogenesis, Dnmt1 is present at high levels in nuclei
only in growing dictyate oocytes, a stage during which gynogenetic developmental potential is
lost and biparental developmental potential is gained.

<>

<1>Mertz, J.E., Davis, R.W.
<2>Cleavage of DNA by EcoRI restriction endonuclease generates cohesive ends.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>3370-3374
<6>1972
<7>EcoRI restriction endonuclease cleaves duplex DNA at a specific sequence,
probably 6 nucleotide pairs in length, by making two single-strand staggered
cleavages, generating 5'-phosphoryl and 3'-hydroxyl termini.  The single-strand
ends produced at each break have identical and complementary sequences of 4 or
6 nucleotides in length.  Therefore, the cleavage site possesses a 2-fold
rotational axis of symmetry perpendicular to the helix axis.  The ends of
full-length linear SV40 DNA, generated by EcoRI endonuclease cleavage, can be
joined by Escherichia coli ligase to regenerate duplex, fully infectious,
covalently-closed circular molecules.  It was further found that all EcoRI
endonuclease-generated ends are identical and complementary.  Therefore, any
two DNA molecules with EcoRI sites can be "recombined" at their restriction
sites by the sequential action of EcoRI endonuclease and DNA ligase to generate
hybrid DNA molecules.

<>

<1>Meselson, M., Yuan, R.
<2>DNA-restriction enzyme from E. coli K.
<3>Procedures in Nucleic Acids Research, Harper and Row, Cantoni, G.I., Davies, R., New York
<4>2
<5>889-895
<6>1972
<7>Most strains of E. coli are able to recognize and degrade DNA from foreign E.
coli strains.  Whether a foreign DNA molecule is degraded depends on certain
nonheritable properties imparted by the cell from which it is obtained.  These
properties are called host-controlled modifications.  For example, the result
of infecting E. coli strain K with phage lambda depends on the host in which
the phages were last grown.  Phages grown in bacteria possessing the
modification character mk multiply successfully, but phages from bacteria
lacking mk do not.  Instead, their DNA is quickly degraded on entering cells of
strain K.  The ability of strain K to degrade or "restrict" DNA from cells
lacking mk is itself under genetic control, the responsible character being
designated rk.  An endonuclease has been shown to be responsible for
restriction in E. coli K.  It is specifically active against lambda DNA from
strains lacking mk and is called endonuclease R.K.

<>

<1>Meselson, M., Yuan, R.
<2>DNA restriction enzyme from E. coli.
<3>Nature
<4>217
<5>1110-1114
<6>1968
<7>An endonuclease which degrades foreign DNA has been isolated.  The enzyme
requires S-adenosylmethionine, ATP and Mg++.

<>

<1>Meselson, M., Yuan, R., Heywood, J.
<2>Restriction and modification of DNA.
<3>Annu. Rev. Biochem.
<4>41
<5>447-466
<6>1972
<7>None

<>

<1>Meslier, V., Loux, V., Renault, P.
<2>Genome Sequence of Lactococcus raffinolactis Strain 4877, Isolated from Natural Dairy Starter Culture.
<3>J. Bacteriol.
<4>194
<5>6364
<6>2012
<7>The nonstarter lactic acid bacterium Lactococcus raffinolactis is prevalent in a  wide range
of environments, such as the dairy environment, but little is known
about this species. Here, we present the draft genome of Lactococcus
raffinolactis strain 4877, isolated from a natural mesophilic dairy starter
culture.

<>

<1>Meslier, V., Loux, V., Renault, P.
<2>Genome Sequence of Leuconostoc pseudomesenteroides Strain 4882, Isolated from a Dairy Starter Culture.
<3>J. Bacteriol.
<4>194
<5>6637
<6>2012
<7>The nonstarter lactic acid bacterium Leuconostoc pseudomesenteroides is a species widely found
in the dairy industry and plays a key role in the formation of
aromatic compounds. Here, we report the first genome sequence of a dairy strain
of Leuconostoc pseudomesenteroides, which is 2 Mb.

<>

<1>Messer, W., Noyer-Weidner, M.
<2>Timing and targeting: the biological functions of Dam methylation in E. coli.
<3>Cell
<4>54
<5>735-737
<6>1988
<7>Postreplicative DNA methylation superimposes on the fixed primary information of the DNA
sequence secondary information that has significance in the regulation of a variety of
cellular processes in prokaryotes and eukaryotes.  In many eukaryotes, methylation of cytosine
residues in defined regions of the genome is involved in a global control of gene activity.
The methylation pattern may change during development, but once established, it can be stably
transmitted to daughter cells by the activity of maintenance methyltransferases.  Highly
methylated regions are often transcriptionally inactive.  In contrast to the situation in
eukaryotes, the regulatory effects of Dam methylation in Escherichia coli do not arise from
selective methylation of defined regions of the genome. The Dam DNA methyltransferase
methylates the N6 position of adenine at all GATC sites.  However, because there is a lag
between replication and methylation, newly replicated DNA is hemimethylated and therefore
distinct from the rest of the genome.  The hemimethylated status of new DNA provides a time
window during which certain cellular processes are activated or suppressed, thus linking these
processes to the cell cycle.  Moreover, the methylation asymmetry in this window allows
distinction between parental (methylated) and daughter (unmethylated) DNA strands.  Here we
discuss the regulatory potential of these two forms of superimposed information.

<>

<1>Messick, J.B., Santos, A.P., Guimaraes, A.M.
<2>Complete Genome Sequences of Two Hemotropic Mycoplasmas, Mycoplasma haemofelis Strain Ohio2 and Mycoplasma suis Strain Illinois.
<3>J. Bacteriol.
<4>193
<5>2068-2069
<6>2011
<7>We report the complete and fully assembled genomes of Mycoplasma haemofelis strain Ohio2 and
Mycoplasma suis strain Illinois, which are the
first available genomes of these uncultivatable hemoplasma species. The
single circular chromosomes of 1,152,484 bp and 742,431 bp for M.
haemofelis and M. suis, respectively, are typical of mycoplasma species,
having reduced size and low G+C content (38.8% for M. haemofelis and 31.1%
for M. suis). Their metabolic pathways are reduced, with evidence of
adaption to the blood environment.

<>

<1>Metcalf, W.W.
<2>Genetic analysis in the domain Archaea.
<3>Methods Microbiol.
<4>29
<5>277-326
<6>1999
<7>The recognition of the Archaea as a phylogenetic domain at the same hierarchical level as the
Bacteria or Eukarya is a relatively recent development in the biological sciences.  This
revision of the classical prokaryotic vs. eukaryotic phylogeny stems largely from the work of
Carl Woese and his collaborators, and originated in the late 1970's from the study of what
were then known as methane-producing bacteria.  Utilizing 16S RNA as a molecular marker for
determining phylogenetic relationships between organisms, Woese and his collaborators showed
that these organisms were vastly different from the known bacterial species and proposed they
should be considered as a separate group, designated the Archaebacteria.  Although this
assertion was hotly contested for many years, its validity has been gradually substantiated by
a variety of methods and is now widely accepted.  The term Archaebacteria has been replaced by
Archaea to emphasize the important point that these are not Bacteria.

<>

<1>Methe, B.A. et al.
<2>Genome of Geobacter sulfurreducens: metal reduction in subsurface environments.
<3>Science
<4>302
<5>1967-1969
<6>2003
<7>The complete genome sequence of Geobacter sulfurreducens, a delta-proteobacterium, reveals
unsuspected capabilities, including
evidence of aerobic metabolism, one-carbon and complex carbon metabolism,
motility, and chemotactic behavior. These characteristics, coupled with
the possession of many two-component sensors and many c-type cytochromes,
reveal an ability to create alternative, redundant, electron transport
networks and offer insights into the process of metal ion reduction in
subsurface environments. As well as playing roles in the global cycling of
metals and carbon, this organism clearly has the potential for use in
bioremediation of radioactive metals and in the generation of electricity.

<>

<1>Methe, B.A. et al.
<2>The psychrophilic lifestyle as revealed by the genome sequence of Colwellia psychrerythraea 34H through genomic and proteomic analyses.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>10913-10918
<6>2005
<7>The completion of the 5,373,180-bp genome sequence of the marine psychrophilic bacterium
Colwellia psychrerythraea 34H, a model for the study of life in permanently cold environments,
reveals capabilities important to carbon and nutrient cycling, bioremediation, production of
secondary metabolites, and cold-adapted enzymes. From a genomic perspective, cold adaptation
is suggested in several broad categories involving changes to the cell membrane fluidity,
uptake and synthesis of compounds conferring cryotolerance, and strategies to overcome
temperature-dependent barriers to carbon uptake. Modeling of three-dimensional protein
homology from bacteria representing a range of optimal growth temperatures suggests changes to
proteome composition that may enhance enzyme effectiveness at low temperatures. Comparative
genome analyses suggest that the psychrophilic lifestyle is most likely conferred not by a
unique set of genes but by a collection of synergistic changes in overall genome content and
amino acid composition.

<>

<1>Meunier, B., Tian, G.-L., Macadre, C., Slonimski, P.P., Lazowska, J.
<2>Group II introns transpose in yeast mitochondria.
<3>Structure, function and biogenesis of energy transfer systems., Elsevier Scientific Publishing Company, Quagliariello, E., Papa, S., Palmieri, F., Saccone, C., Amsterdam
<4>0
<5>169-174
<6>1990
<7>There is a set of genetic phenomena which, first discovered in yeast mitochondrial genetics
twenty years ago, is still of considerable interest for molecular biologists.  Variously
described as polarity of transmission and recombination of genetic markers, unidirectional
gene conversion with coconversion of flanking markers, intron gene conversion, duplicative
transposition, homing or intron infectivity these phenomena have one feature in common: a
specific portion of the donor mitochondrial genome, is preferentially transmitted to the
progeny when compared to other parts of the same genome.  The leading element of this gene
conversion is an intron and the process is initiated by an intron-encoded-protein which has a
specific double strand DNA endonuclease activity that cleaves an intronless recipient
mitochondrial genome at (or in the vicinity of) the site of intron insertion.  For many years
the phenomenon was restricted to the omega+ intron located in the large rRNA gene but more
recently it was observed for the ai4 intron of the COX1 gene.  Both omega+ and ai4 belong to
the group I of intronic RNA structure and code for proteins which share several polypeptide
motifs in common with RNA maturases.

<>

<1>Meyer, P.
<2>DNA methylation of flower color transgenes in Petunia hybrida.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, Russo, V.E.A., Martienssen, R.A., Riggs, A.D., Cold Spring Harbor
<4>0
<5>305-317
<6>1996
<7>Inactivation of transgene constructs that have been introduced into plants is a frequently
reported phenomenon.  Many such silencing events are associated with DNA methylation, although
it is still a matter of debate whether de novo methylation is the cause or the consequence of
gene inactivation.  A correlation between DNA methylation and gene inactivation is not limited
to transgenes but is also observed for transposable elements and for some, but not all,
endogenous genes.  Interestingly, changes in methylation patterns that correlate with changes
in tissue-specific expression have been found in distant upstream regions of some genes,
indicating that methylation-mediated control of gene expression does not exclusively affect
promoter regions.

<>

<1>Meyerdierks, A., Kube, M., Kostadinov, I., Teeling, H., Glockner, F.O., Reinhardt, R., Amann, R.
<2>Metagenome and mRNA expression analyses of anaerobic methanotrophic archaea of the ANME-1 group.
<3>Environ. Microbiol.
<4>12
<5>422-439
<6>2010
<7>Microbial consortia mediating the anaerobic oxidation of methane with sulfate are composed of
methanotrophic Archaea (ANME) and Bacteria related
to sulfate-reducing Deltaproteobacteria. Cultured representatives are not
available for any of the three ANME clades. Therefore, a metagenomic
approach was applied to assess the genetic potential of ANME-1 archaea. In
total, 3.4 Mbp sequence information was generated based on metagenomic
fosmid libraries constructed directly from a methanotrophic microbial mat
in the Black Sea. These sequence data represent, in 30 contigs, about
82-90% of a composite ANME-1 genome. The dataset supports the hypothesis
of a reversal of the methanogenesis pathway. Indications for an
assimilatory, but not for a dissimilatory sulfate reduction pathway in
ANME-1, were found. Draft genome and expression analyses are consistent
with acetate and formate as putative electron shuttles. Moreover, the
dataset points towards downstream electron-accepting redox components
different from the ones known from methanogenic archaea. Whereas catalytic
subunits of [NiFe]-hydrogenases are lacking in the dataset, genes for an
[FeFe]-hydrogenase homologue were identified, not yet described to be
present in methanogenic archaea. Clustered genes annotated as secreted
multiheme c-type cytochromes were identified, which have not yet been
correlated with methanogenesis-related steps. The genes were shown to be
expressed, suggesting direct electron transfer as an additional possible
mode to shuttle electrons from ANME-1 to the bacterial sulfate-reducing
partner.

<>

<1>Meyerink, J.H., Retel, J.
<2>Topographical analysis of yeast ribosomal DNA by cleavage with restriction endonucleases.  II. The physical map of EcoRI fragments.
<3>Nucleic Acids Res.
<4>3
<5>2697-2707
<6>1976
<7>Yeast ribosomal DNA (rDNA) was digested with the restriction endonuclease
EcoRI.  Eight distinct fragments were obtained with a molecular weight of 4.35
(1), 1.75 (2), 1.45 (3), 1.07 (4), 0.42 (5), 0.37 (6), 0.26 (7) and 0.22 (8)
Mdaltons, respectively.  Except for fragment 1 with a molecular wieght of 4.35
Mdaltons, all fragments are derived from the multiple ribosomal transcription
units.  The 'spacer' sequences, on the other hand, gave rise to digestion
products which are very heterogeneous in size.  By analysis of the partial
digestion products, together with the data obtained by digestion with a
combination of two restriction enzymes (EcoRI and HindII or HindIII) and
redigation of the HindII- and HindIII-fragments with EcoRI, the physical map of
the EcoRI cleavage sites in the ribosomal transcription unit could be
established.

<>

<1>Meyerink, J.H., Retel, J., Planta, R.J., Heidekamp, F.
<2>Topographical analysis of yeast ribosomal DNA by cleavage with restriction endonucleases.
<3>Mol. Biol. Rep.
<4>2
<5>393-400
<6>1976
<7>Yeast ribosomal DNA (rDNA) was digested with the restriction enzymes HindIII,
HindII and a mixture of HindII and HindIII.  The cleavage products were
analyzed by electrophoresis on 1.5% agarose gels.  Several distinct bands could
be observed, which are derived from the redundant ribosomal transcription unit.
They are superimposed on a rather broad smear of background DNA, representing
the heterogenous 'spacer' sequences.  From the restriction maps, together with
data obtained by partial digestion, a physical map for the ribosomal
transcription unit in yeast could be constructed.

<>

<1>Meyertons, J.L.
<2>Actinophages and restriction endonucleases from Micromonospora species (Actinomycetales).
<3>Diss. Abstr.
<4>48
<5>1888
<6>1988
<7>In the course of developing a screening procedure for the detection of
restriction endonucleases in micromonosporae, an economically important group
of actinomycetes, previously undiscovered actinophages were isolated from soils
enriched with Micromonospora species. The majority of the actinophages belonged
to Ackermann's type B1 since they had isometric heads, hexagonal in shape, and
long, noncontractile tails. The tails were often flexible and striated, and
some had terminal bulbs. One actinophage was classified as type C1 because of
the very short, noncontractile tail and the isometric head. The actinophages
all contained double-stranded DNA and had genome sizes ranging from
approximately 40 to 60 kilobases. All actinophages were polyvalent except for
one monovalent isolate specific for M. coerulea (NRRL B16092). The host-ranges
of the actinophages were determined on thirty species of Micromonospora.
Certain actinophages were also capable of infecting strains of
Amorphosporangium, Ampullariella and Catellatospora which are Actinomycetes of
the same cell wall chemotype as Micromonosporae (type II). Whole-cell sugar
analyses were performed on the Micromonospora species using a newly developed
thin-layer chromatography (TLC) method. The TLC method used an
acetonitrile:water development system, and N-naphthylethylenediamine
hydrochloride or acid aniline phthalate to visualize hexoses or pentoses,
respectively. A comparison of host-range studies and whole-cell sugar analyses
suggested that xylose may be involved in actinophage receptors for
Micromonospora species. The actinophages were used in efficiency of plating
studies as biological indicators of host-controlled restriction-modification
activity. Restriction enzymes were detected and isolated from three strains of
Micromonospora. Type II restriction endonucleases were isolated from
Micromonospora echinospora ssp. echinospora (ATCC 15837), M. purpurea (ATCC
15835) and M. zionensis (LL-100-125), and were designed MecI, MpuI and MziI,
respectively. Restriction enzymes MecI and MpuI are isoschizomers of XhoI (from
Xanthomonas holcicola), while MziI is an isoschizomer of PvuII (from Proteus
vulgaris). One can speculate that a number of R-M systems remain to be
identified in Micromonospora species and that the discovery of an enzyme with a
novel recognition sequence is possible.

<>

<1>Meyertons, J.L.
<2>Actinophages and restriction endonucleases from Micromonospora species (Actinomycetales).
<3>Ph.D. Thesis, Rutgers University, USA
<4>
<5>1-117
<6>1987
<7>In the course of developing a screening procedure for the detection of
restriction endonucleases in Micromonosporae, an economically important group
of actinomycetes, previously undiscovered actinophages were isolated from soils
enriched with Micromonospora species.  The majority of the actinophages
belonged to Ackermann's type B1 since they had isometric heads, hexagonal in
shape, and long, noncontractile tails.  The tails were often flexible and
striated, and some had terminal bulbs.  One actinophage was classified as type
C1 because of the very short, noncontractile tail and the isometric head.  The
actinophages all contained double-stranded DNA and had genome sizes ranging
from approximately 40 to 60 kilobases.  All actinophages were polyvalent except
for one monovalent isolate specific for M. coerulea (NRRL B16093).  The
host-ranges of the actinophages were determined on thirty species of
Micromonospora.  Certain actinophages were also capable of infecting strains of
Amorphosporangium, Ampullariella and Catellatospora which are actinomycetes of
the same cell wall chemotype as Micromonosporae (type II).  Whole-cell sugar
analyses were performed on the Micromonospora species using a newly developed
thin-layer chromatography (TLC) method.  The TLC method used an
acetonitrile:water development system, and N-naphthylethylenediamine
hydrochloride or acid aniline phthalate to visualize hexoses or pentoses,
respectively.  A comparison of host-range studies and whole-cell sugar analyses
suggested that xylose may be involved in actinophage receptors for
Micromonospora species.  The actinophages were used in efficiency of plating
studies as biological indicators of host-controlled restriction-modification
activity.  Restriction enzymes were detected and isolated from three strains of
Micromonospora.  Type II restriction endonucleases werre isolated from
Micromonospora echinospora ssp. echinospora (ATCC 15837), M. purpurea (ATCC
15835) and M. zionensis (LL-100-125), and were designated MecI, MpuI and MziI,
respectively.  Restriction enzymes MecI and MpuI are isoschizomers of XhoI
(from Xanthomonas holcicola), while MziI is an isoschizomer of PvuII (from
Proteus vulgaris).  One can speculate that a number of R-M systems remain to be
identified in Micromonospora species and that the discovery of an enzyme with a
novel recognition sequence is possible.

<>

<1>Meyertons, J.L., Tilley, B.C., Lechevalier, M.P., Lechevalier, H.A.
<2>Actinophages and restriction enzymes from Micromonospora species (Actinomycetales).
<3>J. Ind. Microbiol.
<4>2
<5>293-303
<6>1987
<7>To develop a screening procedure for the detection of restriction endonucleases
in Micromonosporae and Catellatosporae based on efficiency of plating, eight
different actinophages were isolated from soils enriched with Micromonospora
species and one from Catellatospora-enriched soil.  The lytic actinophages all
contained double-stranded DNA and the majority appeared, when examined by
electron microscopy, to belong to Ackermann's type B1 since they had isometric
heads and noncontractile tails.  One actinophage was classified as type C1
because of its isometric head and very short noncontractile tail.  The host
ranges of the actinophages were determined on strains of Micromonospora and
selected species from other actinomycete genera of cell wall chemotype II.
Type II restriction enzymes were isolated from M. echinospora ssp. echinospora
(ATCC 15837), M. purpurea (ATCC 15835) and M. zionensis (LL-100-125) and were
designated MecI, MpuI, and MziI, respectively.  Restriction enzymes MecI and
MpuI are isoschizomers of XhoI, while MziI is an isoschizomer of PvuII.

<>

<1>Mhanni, A.A., Yoder, J.A., Dubesky, C., McGowan, R.A.
<2>Cloning and sequence analysis of a zebrafish cDNA encoding DNA (cytosine-5)-methyltransferase-1.
<3>Genesis
<4>30
<5>213-219
<6>2001
<7>Summary: The zebrafish has become a well-established animal model for the analysis of
development and of several disease phenotypes. Several of the favorable traits that make it a
popular model organism would also be beneficial for the study of normal and abnormal
vertebrate development in which DNA methylation may play a role. We report the determination
of the full-length cDNA sequence corresponding to the zebrafish DNA (cytosine-5-)
methyltransferase gene, Dnmt1. It is 4,907 bases long and has an open reading frame predicted
to encode a 1,499 amino acid protein that is similar in size and sequence to a number of other
methyltransferases identified in other organisms.

<>

<1>Mi, S., Alonso, D., Roberts, R.J.
<2>Functional analysis of Gln-237 mutants of HhaI methyltransferase.
<3>Nucleic Acids Res.
<4>23
<5>620-627
<6>1995
<7>When the HhaI (cytosine-5) methyltransferase (M.HhaI) binds DNA it causes the target cytosine
to be flipped 180 degrees out of the helix. The space becomes occupied by two amino acids,
Ser-87 and Gln-237, which enter the helix from opposite sides and form a hydrogen bond to each
other. Gln-237 may be involved in specific sequence recognition since it forms three hydrogen
bonds to the orphan guanosine, which is the partner of the target cytosine. We have prepared
all 19 mutants of Gln-237 and tested their biochemical properties. We find that mutations of
this residue greatly affect the stability of the M.HhaI-DNA complex without affecting the
enzyme's specificity for the target sequence. Surprisingly, all mutants retain detectable
levels of enzymatic activity.

<>

<1>Mi, S., Roberts, R.J.
<2>How M.MspI and M.HpaII decide which base to methylate.
<3>Nucleic Acids Res.
<4>20
<5>4811-4816
<6>1992
<7>The HpaII methylase (M.HpaII) recognizes the sequence CCGG and methylates the inner cytosine
residue. The MspI methylase (MspI) recognizes the same sequence but methylates the outer
cytosine residue. Both methylases have the usual architecture of 10 well-conserved motifs
surrounding a variable region, responsible for sequence specific recognition, that is quite
different in the two methylases. We hav constructed hybrids between these two methylases and
studied their methylation properties. A hybrid containing the variable region and C-terminal
sequences from M.MspI methylates the outer cytosine residue. A second hybrid identical to the
first except that the variable region derives from the M.HpaII methylates the inner cytosine
residue. Thus the choice of base to be methylated within the recognition sequence is
determined by the variable region.

<>

<1>Mi, S., Roberts, R.J.
<2>The DNA binding affinity of HhaI methylase is increased by a single amino acid substitution in the catalytic center.
<3>Nucleic Acids Res.
<4>21
<5>2459-2464
<6>1993
<7>The HhaI methyltransferase recognizes the sequence GCGC and transfers a methyl group to C5 of
the first cytosine residue. All m5C-methyltransferases contain a highly conserved sequence
motif called the P-C motif. The cysteine residue of this motif is involved in catalysis by
forming a covalent bond with the 6-position of cytosine prior to methyl group transfer. For
the EcoRII methyltransferase, it has been shown that substitution of this catalytic cysteine
by glycine is cytotoxic to E.coli cells expressing the mutant methyltransferase (Wyszynski et
al. Nucl. Acids. Res. 20:319,1992). We now show that this observation can be extended to the
HhaI system and suggest that the cytotoxicity is due to abnormally tight DNA binding by the
mutant methyltransferase, which probably interferes with replication or transcription.

<>

<1>Mi, S., Song, J., Lin, J., Che, Y., Zheng, H., Lin, J.
<2>Complete genome of Leptospirillum ferriphilum ML-04 provides insight into its physiology and environmental adaptation.
<3>J. Microbiol.
<4>49
<5>890-901
<6>2011
<7>Leptospirillum ferriphilum has been identified as the dominant, moderately
thermophilic, bioleaching microorganism in bioleaching processes. It is an acidic
and chemolithoautrophic bacterium that gains electrons from ferrous iron
oxidation for energy production and cell growth. Genetic information about this
microorganism has been limited until now, which has hindered its further
exploration. In this study, the complete genome of L. ferripilum ML-04 is
sequenced and annotated. The bacterium has a single circular chromosome of
2,406,157 bp containing 2,471 coding sequences (CDS), 2 rRNA operons, 48 tRNA
genes, a large number of mobile genetic elements and 2 genomic islands. In silico
analysis shows L. ferriphilum ML-04 fixes carbon through a reductive citric acid
(rTCA) cycle, and obtains nitrogen through ammonium assimilation. The genes
related to "cell envelope biogenesis, outer membrane" (6.9%) and "DNA
replication, recombination and repair" (5.6%) are abundant, and a large number of
genes related to heavy metal detoxification, oxidative and acidic stress defense,
and signal transduction pathways were detected. The genomic plasticity, plentiful
cell envelope components, inorganic element metabolic abilities and stress
response mechanisms found the base for this organism's survival in the
bioleaching niche.

<>

<1>Miao, S., Li, F., Zhang, M., Wang, X., He, J., Jiang, J.
<2>Molecular cloning and sequence analysis of Rana tigrina ranavirus (RTV) DNA methyltransferase gene.
<3>Shuichan Xuebao
<4>26
<5>157-160
<6>2002
<7>The complete gene of RTV DNA methyltransferase (MTase) was amplified, cloned and sequenced.
The ORF consists of 642 bp, which codes for a protein of 214 aa with a predicted molecular
mass
of 24.8kD.  Sequence analysis of MTase gene shows much higher identity to the species of genus
Ranavirus (96%-97%) than to lymphocystis disease virus (56%), the type species of the genus
Lymphocystivirus, indicating again that RTV was the member of the genus Ranavirus; just as the
other vertebrate iridoviruses, the MTase gene of RTV contains the first four highly conserved
motifs of cytosine MTases but the fifth motif, responsible for DNA binding specificity, is
missing; among the virus of RTV, doctor fish iridovirus and largemouth bass ranavirus, the
very
different identity between MTase and MCP gene suggests that the gene used as target to
estimate the evolution of the iridoviruses should be suitable.

<>

<1>Michael, G.B., Kadlec, K., Sweeney, M.T., Brzuszkiewicz, E., Liesegang, H., Daniel, R., Murray, R.W., Watts, J.L., Schwarz, S.
<2>ICEPmu1, an integrative conjugative element (ICE) of Pasteurella multocida: structure and transfer.
<3>J. Antimicrob. Chemother.
<4>67
<5>91-100
<6>2012
<7>BackgroundIntegrative and conjugative elements (ICEs) have not been
detected in Pasteurella multocida. In this study the multiresistance
ICEPmu1 from bovine P. multocida was analysed for its core genes and its
ability to conjugatively transfer into strains of the same and different
genera.MethodsICEPmu1 was identified during whole genome sequencing.
Coding sequences were predicted by bioinformatic tools and manually
curated using the annotation software ERGO. Conjugation into P. multocida,
Mannheimia haemolytica and Escherichia coli recipients was performed by
mating assays. The presence of ICEPmu1 and its circular intermediate in
the recipient strains was confirmed by PCR and sequence analysis.
Integration sites were sequenced. Susceptibility testing of the
ICEPmu1-carrying recipients was conducted by broth
microdilution.ResultsThe 82 214 bp ICEPmu1 harbours 88 genes. The core
genes of ICEPmu1, which are involved in excision/integration and
conjugative transfer, resemble those found in a 66 641 bp ICE from
Histophilus somni. ICEPmu1 integrates into a tRNA(Leu) and is flanked by
13 bp direct repeats. It is able to conjugatively transfer to P.
multocida, M. haemolytica and E. coli, where it also uses a tRNA(Leu) for
integration and produces closely related 13 bp direct repeats. PCR assays
and susceptibility testing confirmed the presence and the functional
activity of the ICEPmu1-associated resistance genes in the recipient
strains.ConclusionsThe observation that the multiresistance ICEPmu1 is
present in a bovine P. multocida and can easily spread across strain and
genus boundaries underlines the risk of a rapid dissemination of multiple
resistance genes, which will distinctly decrease the therapeutic options.

<>

<1>Michel, F., Dujon, B.
<2>Genetic exchanges between bacteriophage T4 and filamentous fungi?
<3>Cell
<4>46
<5>323
<6>1986
<7>Note the sequence similarity between I-TevI and a protein encoded by a class 1 intron in the
NADH-dehydrogenase gene of Neurospora crassa.

<>

<1>Mick, J.M., Beck, D.J.
<2>Diamminedichloroplatinum(II) modification of DNA inhibits cutting by restriction endonucleases in a sequence specific manner.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>88
<5>164
<6>1988
<7>cis-Diamminedichloroplatinum(II), cis-DDP, is a potent antitumor agent while
the trans-isomer is ineffective.  Both have been shown to bind to DNA
preferentially at guanine.  In this study, [32P]-end-labeled DNA modified with
cis-DDP or trans-DDP was digested with restriction endonucleases having
recognition sequences at which DDP is capable of reacting to form bifunctional
adducts:  BstUI, GG^CG; HinPI, G^CGC; HhaI, GCG^C; and BamHI, G^GATCC.
Inhibition of cutting by these enzymes was compared to that for ClaI, AT^CGAT,
as DDP reacts with bases of its recognition site in a monofunctional manner.
Restriction fragments were subjected to electrophoresis and the amounts of
radioactivity in each were quantitated by densitometry and liquid scintillation
spectrophotometry.  Inhibition by trans-DDP was greatest when the base sequence
GXG was adjacent to, or occluding the cut site.  Thus, two guanines separated
by a third base appears to be a biologically inhibitory adduct of trans-DDP
while cis-DDP forms such adducts at lower frequency.  Inhibition by cis-DDP was
greatest at the BamHI site where it can form crosslinks between adjacent
guanines.

<>

<1>Middleton, J.H., Edgell, M.H., Hutchison, C.A. III
<2>Specific fragments of PhiX174 deoxyribonucleic acid produced by a restriction enzyme from Haemophilus aegyptius, endonuclease Z.
<3>J. Virol.
<4>10
<5>42-50
<6>1972
<7>A restriction-like enzyme has been purified from Haemophilus aegyptius.  This
nuclease, endonuclease Z, produces a rapid decrease in the viscosity of native
calf thymus and H. influenzae deoxyribonucleic acids (DNA), but does not
degrade homologous DNA.  The specificity of endonuclease Z is different from
that of the similar endonuclease isolated from H. influenzae (endonuclease R).
The purified enzyme cleaves the double-stranded replicative form DNA of
bacteriophage PhiX174 (PhiX174 RF DNA) into at least 11 specific limit
fragments whose molecular sizes have been estimated by gel electrophoresis.
The position of these fragments with respect to the genetic map of PhiX174 can
be determined by using the genetic assay for small fragments of PhiX174 DNA.

<>

<1>Midha, S., Ranjan, M., Sharma, V., Kumari, A., Singh, P.K., Korpole, S., Patil, P.B.
<2>Genome Sequence of Pediococcus pentosaceus Strain IE-3.
<3>J. Bacteriol.
<4>194
<5>4468
<6>2012
<7>We report the 1.8-Mb genome sequence of Pediococcus pentosaceus strain IE-3, isolated from a
dairy effluent sample. The whole-genome sequence of this strain
will aid in comparative genomics of Pediococcus pentosaceus strains of diverse
ecological origins and their biotechnological applications.

<>

<1>Midha, S., Ranjan, M., Sharma, V., Pinnaka, A.K., Patil, P.B.
<2>Genome Sequence of Xanthomonas citri pv. mangiferaeindicae Strain LMG 941.
<3>J. Bacteriol.
<4>194
<5>3031
<6>2012
<7>We report the 5.1-Mb genome sequence of Xanthomonas citri pv. mangiferaeindicae strain LMG
941, the causal agent of bacterial black spot in mango. Apart from evolutionary studies, the
draft genome will be a valuable resource for the epidemiological studies and quarantine of
this phytopathogen.

<>

<1>Mierzejewska, K., Siwek, W., Czapinska, H., Kaus-Drobek, M., Radlinska, M., Skowronek, K., Bujnicki, J.M., Dadlez, M., Bochtler, M.
<2>Structural basis of the methylation specificity of R.DpnI.
<3>Nucleic Acids Res.
<4>42
<5>8745-8754
<6>2014
<7>R.DpnI consists of N-terminal catalytic and C-terminal winged helix domains that  are
separately specific for the Gm6ATC sequences in Dam-methylated DNA. Here we
present a crystal structure of R.DpnI with oligoduplexes bound to the catalytic
and winged helix domains and identify the catalytic domain residues that are
involved in interactions with the substrate methyl groups. We show that these
methyl groups in the Gm6ATC target sequence are positioned very close to each
other. We further show that the presence of the two methyl groups requires a
deviation from B-DNA conformation to avoid steric conflict. The methylation
compatible DNA conformation is complementary with binding sites of both R.DpnI
domains. This indirect readout of methylation adds to the specificity mediated by
direct favorable interactions with the methyl groups and solvation/desolvation
effects. We also present hydrogen/deuterium exchange data that support
'crosstalk' between the two domains in the identification of methylated DNA,
which should further enhance R.DpnI methylation specificity.

<>

<1>Mierzejewska, K., Siwek, W., Czapinska, H., Skowronek, K., Bujnicki, J., Bochtler, M.
<2>Molecular basis of 6-methyladenine recognition by R.DpnI restriction endonuclease.
<3>FEBS J.
<4>280
<5>122-123
<6>2013
<7>The 6-methyladenine is one of the key epigenetic modifications in prokaryotes.  R.DpnI is the
best known 6 mA-dependent restriction endonuclease, specific for Dam methylated G6 mATC sites
and widely used for site-directed mutagenesis.  Our previous studies have shown that R.DpnI
consists of two domains, an N-terminal catalytic PD-D/E_XK domain and a C-terminal winged
helix domain and that both independently read out DNA sequence and methylation status.  In our
first structure of R.DpnI-DNA complex, there is only one substrate oligoduplex per enzyme
molecule, which is specifically bound to the wH domain and distant from the calalytic domain.
Hence, the question remained open how the catalytic domain of R.DpnI interacts with its
target, and how it specifically recognizes the methyl groups which license DNA cleavage.  Such
a process is difficult to realize with stringency, because attractive van der Waals
interactions with the small hydrophobic group are relatively weak when compared to repulsive
ones.  The structures depicting the phenomenon are rare and there is still relatively little
known about the mechanism of recognition by modification dependent enzymes.  Here, we present
a high resolution structure of R.DpnI which features both protein domains bound to the target
DNA.  Recognition of the 6mAs by the catalytic domain is carried out without flipping bases
out of the helix.  A previously disordered loop wraps around the major groove of the target
DNA, where both methyl groups are located in close proximity to each other.  The PD-D/E)XK
domain places the 6mAs in a hydrophobic pocket formed by residues Leu129 and Trp 138.  To the
best of our knowledge this is the first structure showing how a protein can efficiently detect
two 6mA modified sites together.

<>

<1>Mikhailova, V.K.
<2>Restriction endonuclease XbaI preparation - involves disintegrating biomass cells on Xanthomonas badrii, chromatographic purification and dialysis fractionating.
<3>Soviet Patent Office
<4>SU 1634713 A
<5>
<6>1991
<7>The restriction endonuclease XbaI is obtained from the biomass produced by Xanthomonas badrii
VKPM B-2754 (ATCC 11672). The cells are disintegrated ultrasonically and the homogenate is
fractionated in a two phase system, containing 7% polyethylene glycol and 2% dextran, in the
presence of 0.35-0.42M NaCl. Chromatographic purification and dialysis completes the process.
Tests show that the purification of the product on heparin-sepharose, followed by dialysis
against 50% solution of glycerin, produces the enzyme in yields of 12-15k units/g of biomass.
The enzyme is free of non-specific endonucleases and phosphatases. All purification stages are
carried out at 4 +/- 4 degrees C.

<>

<1>Mikhailova, V.K., Puchkova, L.I., Kuvshinov, V.N., Ushakova, T.A., Repin, V.E.
<2>New method for the isolation of endonuclease restriction Bse21 l, comprises recognizing and cleaving the nucleotide sequence 5'-GCTNAGG-3' and purifying - restriction endonuclease isolation and purification using fractionation.
<3>Russian Patent Office
<4>RU 2184777
<5>
<6>2002
<7>NOVELTY - New method for the isolation of endonuclease restriction Bse21 I comprises: (1)
recognizing and cleaving the nucleotide sequence 5'-GCTNAGG-3' to ensure that the isolated
enzyme contains no impurities comprising of nonspecific nucleases, phosphatases and/or protein
to give an enzyme yield of 3 000 000 U (sic) per 10 g of wetted biomass; and (2) purifying.
BIOTECHNOLOGY - Preferred Method: Isolation of endonuclease restriction
Bse21 I is carried out at the room temperature and pH = 7.5 by fractionation of cellular
homogenate in a two-phase system of polyethylene glycol/dextran in the presence of sodium
chloride (0.2-0.3 M). Purification of the enzyme is carried out by chromatography on
phosphocellulose P-11 and hydroxylapatite. USE - To prepare endonuclease restriction Bse21 I.
ADVANTAGE - Method produces an isolation yield of up to 30 000 U of enzyme/g of biomass within
40 hours.

<>

<1>Miki, T., Okada, N.
<2>Draft Genome Sequence of Chromobacterium haemolyticum Causing Human Bacteremia Infection in Japan.
<3>Genome Announcements
<4>2
<5>e01047-14
<6>2014
<7>Chromobacterium haemolyticum is a Gram-negative bacterium displaying remarkable hemolysis
against human and sheep erythrocytes. In addition, C. haemolyticum
infects humans, in which the infection mechanism remains unknown. We report here
the draft genome sequence of C. haemolyticum strain T124, isolated from a young
patient with sepsis in Japan.

<>

<1>Milani, C., Duranti, S., Lugli, G.A., Bottacini, F., Strati, F., Arioli, S., Foroni, E., Turroni, F., van Sinderen, D., Ventura, M.
<2>Comparative Genomics of Bifidobacterium animalis subsp. lactis Reveals a Strict Monophyletic Bifidobacterial Taxon.
<3>Appl. Environ. Microbiol.
<4>79
<5>4304-4315
<6>2013
<7>Strains of Bifidobacterium animalis subsp. lactis are extensively exploited by
the food industry as health-promoting bacteria, although the genetic variability
of members belonging to this taxon has so far not received much scientific
attention. In this article, we describe the complete genetic makeup of the B.
animalis subsp. lactis Bl12 genome and discuss the genetic relatedness of this
strain with other sequenced strains belonging to this taxon. Moreover, a detailed
comparative genomic analysis of B. animalis subsp. lactis genomes was performed,
which revealed a closely related and isogenic nature of all currently available
B. animalis subsp. lactis strains, thus strongly suggesting a closed pan-genome
structure of this bacterial group.

<>

<1>Milani, C., Lugli, G.A., Duranti, S., Turroni, F., Bottacini, F., Mangifesta, M., Sanchez, B., Viappiani, A., Mancabelli, L., Taminiau, B., Delcenserie, V., Barrangou, R., Margolles, A., van Sinderen, D., Ventura, M.
<2>Genome encyclopaedia of type strains of the genus Bifidobacterium.
<3>Appl. Environ. Microbiol.
<4>80
<5>6290-6302
<6>2014
<7>Bifidobacteria represent one of the dominant microbial groups that are present in the gut of
various animals, being particularly prevalent during the suckling stage of life of humans and
other mammals.  However, the overall genome structure of this group of microorganisms remains
largely unexplored.  Here, we sequenced the genomes of 42 representative (sub)species across
the Bifidobacterium genus, and used this information to explore the overall genetic picture of
this bacterial group.  Furthermore, the here described genomic data were used to reconstruct
the evolutionary development of the Bifidobacterium genus.  This reconstruction suggests that
its evolution was substantially influenced by genetic adaptations to obtain access to glycans,
thereby representing a common and potent evolutionary force in shaping bifidobacterial
genomes.

<>

<1>Militello, K.T., Simon, R.D., Qureshi, M., Maines, R., VanHorne, M.L., Hennick, S.M., Jayakar, S.K., Pounder, S.
<2>Conservation of Dcm-mediated cytosine DNA methylation in Escherichia coli.
<3>FEMS Microbiol. Lett.
<4>328
<5>78-85
<6>2012
<7>In Escherichia coli, cytosine DNA methylation is catalyzed by the DNA cytosine
methyltransferase (Dcm) protein and occurs at the second
cytosine in the sequence 5'CCWGG3'. Although the presence of cytosine
DNA methylation was reported over 35years ago, the biological role of
5-methylcytosine in E.coli remains unclear. To gain insight into the
role of cytosine DNA methylation in E.coli, we (1) screened the 72
strains of the ECOR collection and 90 recently isolated environmental
samples for the presence of the full-length dcm gene using the
polymerase chain reaction; (2) examined the same strains for the
presence of 5-methylcytosine at 5'CCWGG3' sites using a restriction
enzyme isoschizomer digestion assay; and (3) quantified the levels of
5-methyl-2'-deoxycytidine in selected strains using liquid
chromatography tandem mass spectrometry. Dcm-mediated cytosine DNA
methylation is conserved in all 162 strains examined, and the level of
5-methylcytosine ranges from 0.86% to 1.30% of the cytosines. We also
demonstrate that Dcm reduces the expression of ribosomal protein genes
during stationary phase, and this may explain the highly conserved
nature of this DNA modification pathway.

<>

<1>Militello, K.T., Simon, R.D., Qureshi, M., Maines, R., VanHorne, M.L., Hennick, S.M., Jayakar, S.K., Pounder, S.
<2>Dcm-mediated cytosine DNA methylation is conserved in Escherichia coli and influences the expression of ribosomal protein genes.
<3>FASEB J.
<4>24
<5>0
<6>2010
<7>In Escherichia coli, cytosine DNA methylation is catalyzed by the Dcm (DNA cytosine methylase)
protein and occurs at the second cytosine in
the sequence 5'CCWGG3'. Although the presence of cytosine DNA
methylation was reported over 35 years ago, the biological role of
5-methylcytosine in E. coli remains unclear. There is data indicating
that Dcm can protect the genome against attack by a restriction enzyme
that cleaves the same sequence, yet DNA methyltransferases in
eukaryotes often influence gene expression. In order to gain insight
into the potential roles of cytosine DNA methylation in E. coli, we:
(a) screened 162 strains including laboratory strains, pathogens, and
recently isolated environmental samples for the presence of the
full-length dcm gene using the polymerase chain reaction; (b) examined
the same strains for the presence of 5-methylcytosine at 5'CCWGG3'
sites using a restriction enzyme isoschizomer digestion assay; and (c)
quantified the levels of 5-methyl-2'-deoxycytidine in selected strains
using liquid chromatography tandem mass spectrometry. Dcm-mediated
cytosine DNA methylation is conserved in all 162 strains examined, and
the level of 5-methylcytosine ranges from 0.8-1.2% of the cytosines. We
also tested the hypothesis that Dcm influences gene expression. Gene
expression in wild-type and dcm knockout cells was compared via qPCR.
We focused on ribosomal protein genes, as they have been previously
demonstrated to contain numerous 5'CCWGG3' sites. We demonstrate that
Dcm represses expression of ribosomal protein genes during stationary
phase, and this may explain the ubiquitous presence of this DNA
modification pathway. Support for this work was provided by the Geneseo
Foundation and NIH grant R15AI074035-01.

<>

<1>Milkman, R.
<2>Recombination and population structure in Escherichia coli.
<3>Genetics
<4>146
<5>745-750
<6>1997
<7>A major focus of population genetics, and now of molecular evolution, is the study of gene
lineages.  Population genetic parameters usually apply to small chromosomal regions rather
than to the genome as a whole, although genes do not always descend independently of their
surroundings.  The genome of Escherichia coli strikes me as an unusually favorable theater in
which to observe the lineages of genes and their relationships with the evolutionary processes
that operate at the population level.  I should like to trace the developing understanding of
E. coli's population genetics in  the coexistence of clonality and recombination, the
recognition of restriction as important to recombination, and the emerging features of its
genomic structure.

<>

<1>Milkman, R., Jaeger, E., McBride, R.D.
<2>Molecular evolution of the Escherichia coli chromosome. VI. Two regions of high effective recombination.
<3>Genetics
<4>163
<5>475-483
<6>2003
<7>Two 6- to 8-min regions, centered respectively near 45 min (O-antigen region) and 99 min
(restriction-modification region) on the Escherichia
coli chromosome, display unusually high variability among 11 otherwise
very similar strains. This variation, revealed by restriction fragment
length polymorphism (RFLP) and nucleotide sequence comparisons, appears to
be due to a great local increase in the retention frequency of recombinant
replacements. We infer a two-step mechanism. The first step is the
acquisition of a small stretch of DNA from a phylogenetically distant
source. The second is the successful retransmission of the imported DNA,
together with flanking native DNA, to other strains of E. coli. Each cell
containing the newly transferred DNA has a very high selective advantage
until it reaches a high frequency and (in the O-antigen case) is
recognized by the new host's immune system. A high selective advantage
increases the probability of retention greatly; the effective
recombination rate is the product of the basic recombination rate and the
probability of retention. Nearby nucleotide sequences clockwise from the
O-antigen (rfb) region are correlated with specific O antigens, confirming
local hitchhiking. Comparable selection involving imported restriction
endonuclease genes is proposed for the region near 99 min.

<>

<1>Milkman, R., Raleigh, E.A., McKane, M., Cryderman, D., Bilodeau, P., McWeeny, K.
<2>Molecular evolution of the escherichia coli chromosome. V. Recombination patterns among strains of diverse origin.
<3>Genetics
<4>153
<5>539-554
<6>1999
<7>Incorporation patterns of donor DNA into recipient chromosomes following transduction or
conjugation have been studied in the progeny of a variety of Escherichia coli crosses in which
donor and recipient nucleotide sequences differ by 1-3%. Series of contiguous or variously
spaced PCR fragments have been amplified from each recombinant chromosome and digested with a
commercial restriction endonuclease previously shown to distinguish the respective parents in
a given fragment. We conclude that entering donor DNA fragments are frequently abridged (cut
and shortened) before incorporation, the cutting being due to restriction systems, and the
shortening presumably due to exonuclease activity. Analysis of several backcrosses confirms,
and extends to conjugation, the importance of restriction in E. coli recombination in nature.
The transmission patterns in conjugation are similar to those of transduction, but (as
expected) on a much larger scale. Asymmetric results of reciprocal crosses imply that mismatch
frequency is not a major factor. Marked differences among the results of simple crosses
according to parental strain combinations are consistent with observations that E. coli
strains in nature vary dramatically in their restriction-modification systems.

<>

<1>Milla, M.A., Spears, P.A., Pearson, R.E., Walker, G.T.
<2>Use of the restriction enzyme AvaI and Exo- Bst polymerase in strand displacement amplification.
<3>Biotechniques
<4>24
<5>392-396
<6>1998
<7>Strand displacement amplification is an isothermal in vitro method of amplifying DNA.  This
technique is based on the ability of a restriction enzyme to nick one strand of a
hemiphosphorothioated form of its double-stranded recognition site and the ability of a
polymerase to initiate synthesis at the nick and displace the downstream DNA strand during
replication.  The method consists of two parts: (i) a target generation process that makes
copies of the target sequence flanked by nickable restriction sites and (ii) the exponential
amplification of these modified target sequences by repeated nicking, strand displacement and
priming of displaced strands, as depicted in Figure 1.  The first SDA system we developed used
the restriction enzyme HincII and the 3'-5' exonuclease-deficient Klenow fragment of E. coli
polymerase I.  This system achieves 10^8-fold amplification in 2h at 37-40 C.  More recently,
we have developed a thermophilic SDA system that operates at 60 C and uses a restriction
enzyme from Bacillus stearothermophilus and a 3'-5' exonuclease-deficient Klenow fragment of
a DNA polymerase from B. caldotenax.  This system is faster and more powerful, achieving a
10^10-fold amplification in 15 min.  It is also more specific, with a significant reduction in
background amplification.

<>

<1>Millard, A., Clokie, M.R., Shub, D.A., Mann, N.H.
<2>Genetic organization of the psbAD region in phages infecting marine Synechococcus strains.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>11007-11012
<6>2004
<7>The discovery of the genes psbA and psbD, encoding the D1 and D2 core components  of the
photosynthetic reaction center PSII (photosystem II), in the genome of the
bacteriophage S-PM2 (a cyanomyovirus) that infects marine cyanobacteria begs the
question as to how these genes were acquired. In an attempt to answer this
question, it was established that the occurrence of the genes is widespread among
marine cyanomyovirus isolates and may even extend to podoviruses. The phage psbA
genes fall into a clade that includes the psbA genes from their potential
Synechococcus and Prochlorococcus hosts, and thus, this phylogenetic analysis
provides evidence to support the idea of the acquisition of these genes by
horizontal gene transfer from their cyanobacterial hosts. However, the phage psbA
genes form distinct subclades within this lineage, which suggests that their
acquisition was not very recent. The psbA genes of two phages contain identical
212-bp insertions that exhibit all of the canonical structural features of a
group I self-splicing intron. The different patterns of genetic organization of
the psbAD region are consistent with the idea that the psbA and psbD genes were
acquired more than once by cyanomyoviruses and that their horizontal transfer
between phages via a common phage gene pool, as part of mobile genetic modules,
may be a continuing process. In addition, genes were discovered encoding a
high-light inducible protein and a putative key enzyme of dark metabolism,
transaldolase, extending the areas of host-cell metabolism that may be affected
by phage infection.

<>

<1>Millard, J.T., Beachy, T.M.
<2>Cytosine methylation enhances mitomycin C cross-linking.
<3>Biochemistry
<4>32
<5>12850-12856
<6>1993
<7>MitomycinC (M) is a powerful antitumor agent that targets the DNA sequence CpG. Because it is
likely that this dinucleotide will contain 5-methylcytosine in vivo, we have compared the
cross-linking efficiency of MC for DNA containing either 5-methylcytosine or normal cytosine
embedded in random sequence DNA oligomers. We have found that mitomycin C displays a small but
significant preference for methylated DNA. Recognition of an abnormal methylation pattern in
the DNA of transformed cells may therefore be one mechanism by which MC exerts its
chemotherapeutic effects.

<>

<1>Miller, C.A., Cohen, S.N.
<2>Phenotypically cryptic EcoRI endonuclease specified by the ColE1 plasmid.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>75
<5>1265-1269
<6>1978
<7>An endonuclease having EcoRI specificity is produced by bacteria containing the
ColE1 plasmid.  Such bacterial cells fail to express restriction or
modification functions in vivo, and phage or plasmid DNA obtained from
ColE1-containing cells has unmodified EcoRI sites that are extracted from
bacteria that carry ColE1.  No EcoRI DNA methylase activity associated with
ColE1 has been detected.  The finding of phenotypically cryptic ColE1-dependent
EcoRI endonuclease activity and the absence of any detectable EcoRI
modification system in ColE1-containing cells suggest a control mechanism that
appears to prevent functional expression of the ColE1-determined enzyme in
vivo.

<>

<1>Miller, D.A. et al.
<2>Complete Genome Sequence of the Cellulose-Degrading Bacterium Cellulosilyticum lentocellum.
<3>J. Bacteriol.
<4>193
<5>2357-2358
<6>2011
<7>Cellulosilyticum lentocellum DSM 5427 is an anaerobic, endospore-forming member of the
Firmicutes. We describe the complete genome sequence of this cellulose-degrading bacterium;
originally isolated from estuarine sediment of a river that received both domestic and paper
mill waste. Comparative genomics of cellulolytic clostridia will provide insight into factors
that influence degradation rates.

<>

<1>Miller, G.G., Peak, R.M. Jr., Tompson, S.A., Braza, M.J.
<2>IceA gene and method relating thereto.
<3>Japanese Patent Office
<4>JP 1999511032 A
<5>
<6>1999
<7>
<>

<1>Miller, G.G., Peek, R.M. Jr., Thompson, S.A., Blaser, M.J.
<2>IceA gene and related methods.
<3>US Patent Office
<4>US 5780278
<5>
<6>1998
<7>A purified IceA protein of Helicobacter pylori is provided.  The protein is expressed as
either an IceA 1 or IceA2 variant.  A purified polypeptide fragment of the IceA protein is
also provided.  An antigenic fragment of IceA is provided.  An isolated nucleic acid that
encodes an IceA protein of H. pylori is provided.  A nucleic acid that encodes an IceA 1
variant and a nucleic acid that encodes an IceA 2variant are also provided.  Fragments of the
iceA gene are provided.  A method of detecting the presence of an antibody against H. pylori
in a sample is provided.  The method comprises the following steps: a) contacting the sample
with a purified IceA protein of H. pylori or a H. pylori-specific fragment thereof; and b)
detecting the binding of the antibody in the sample to the protein or fragment, the detection
of binding indicating the presence in the sample of antibodies against H. pylori.  A method of
detecting the presence of an antibody against an ulcerative Helicobacter pylori strain in a
sample is also provided.

<>

<1>Miller, G.G., Peek, R.M. Jr., Thompson, S.A., Blaser, M.J.
<2>IceA gene and related methods.
<3>US Patent Office
<4>US 6004354
<5>
<6>1999
<7>A purified IceA protein of Helicobacter pylori is provided.  The protein is expressed as
either an IceA 1 or IceA2 variant.  A purified polypeptide fragment of the IceA protein is
also provided.  An antigenic fragment of IceA is provided.  An isolated nucleic acid that
encodes an IceA protein of H. pylori is provided.  A nucleic acid that encodes an IceA 1
variant and a nucleic acid that encodes an IceA 2 variant is also provided.  Fragments of the
iceA gene are provided.  A method of detecting the presence of an antibody against H. pylori
in a sample is provided.  The method comprises the following steps: a) contacting the sample
with a purified IceA protein of H. pylori or an H. pylori-specific fragment thereof; and b)
detecting the binding of the antibody in the sample to the protein or fragment, the detection
of binding indicating the presence in the sample of antibodies against H. pylori.  A method of
detecting the presence of an antibody against an ulcerative Helicobacter pylori strain in a
sample is also provided.

<>

<1>Miller, G.G., Peek, R.M. Jr., Thompson, S.A., Blaser, M.J.
<2>IceA gene and related methods.
<3>US Patent Office
<4>US 6107464
<5>
<6>2000
<7>A purified IceA protein of Helicobacter pylori is provided.  The protein is expressed as
either an IceA 1 or IceA 2 variant.  A purified polypeptide fragment of the IceA protein is
also provided.  An antigenic fragment of IceA is provided.  An isolated nucleic acid that
encodes an IceA protein of H. pylori is provided.  A nucleic acid that encodes an IceA 1
variant and a nucleic acid that encodes an IceA 2 variant are also provided.  Fragments of the
iceA gene are provided.  A method of detecting the presence of an antibody against H. pylori
in a sample is provided.  The method comprises the following steps: a) contacting the sample
with a purified IceA protein of H. pylori or a H. pylori-specific fragment thereof; and b)
detecting the binding of the antibody in the sample to the protein or fragment, the detection
of binding indicating the presence in the sample of antibodies against H. pylori.  A method of
detecting the presence of an antibody against an ulcerative Helicobacter pylori strain in a
sample is also provided.

<>

<1>Miller, J.C., Holmes, M.C., Wang, J., Guschin, D.Y., Lee, Y.L., Rupniewski, I., Beausejour, C.M., Waite, A.J., Wang, N.S., Kim, K.A., Gregory, P.D., Pabo, C.O., Rebar, E.J.
<2>An improved zinc-finger nuclease architecture for highly specific genome editing.
<3>Nat. Biotechnol.
<4>25
<5>778-785
<6>2007
<7>Genome editing driven by zinc-finger nucleases (ZFNs) yields high gene-modification
efficiencies (>10%) by introducing a recombinogenic
double-strand break into the targeted gene. The cleavage event is induced
using two custom-designed ZFNs that heterodimerize upon binding DNA to
form a catalytically active nuclease complex. Using the current ZFN
architecture, however, cleavage-competent homodimers may also form that
can limit safety or efficacy via off-target cleavage. Here we develop an
improved ZFN architecture that eliminates this problem. Using
structure-based design, we engineer two variant ZFNs that efficiently
cleave DNA only when paired as a heterodimer. These ZFNs modify a native
endogenous locus as efficiently as the parental architecture, but with a
>40-fold reduction in homodimer function and much lower levels of
genome-wide cleavage. This architecture provides a general means for
improving the specificity of ZFNs as gene modification reagents.

<>

<1>Miller, J.F., Dower, W.J., Tompkins, L.S.
<2>High-voltage electroporation of bacteria:  Genetic transformation of Campylobacter jejuni with plasmid DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>85
<5>856-860
<6>1988
<7>Electroporation permits the uptake of DNA by mammalian cells and plant
protoplasts because it induces transient permeability of the cell membrane.  We
investigated the utility of high-voltage electroporation as a method for
genetic transformation of intact bacterial cells by using the enteric pathogen
Campylobacter jejuni as a model system.  This report demonstrates that the
application of high-voltage discharges to bacterial cells permits genetic
transformation.  Our method involves exposure of a Campylobacter cell
suspension to a high-voltage exponential decay discharge (5-13 kV/cm) for a
brief period of time (resistance-capacitance time constant = 2.4-26 msec) in
the presence of plasmid DNA.  Electrical transformation of C. jejuni results in
frequencies as high as 1.2 x 10/6 transformants per microgram of DNA.  We have
investigated the effects of pulse amplitude and duration, cell growth
conditions, divalent cations, and DNA concentration on the efficiency of
transformation.  Transformants of C. jejuni obtained by electroporation
contained structurally intact plasmid molecules.  In addition, evidence is
presented that indicates that C. jejuni possesses DNA restriction and
modification systems.  The use of electroporation as a method for transforming
other bacterial species and guidelines for its implementation are also
discussed.

<>

<1>Miller, P.A., Shajani, Z., Meints, G.A., Caplow, D., Goobes, G., Varani, G., Drobny, G.P.
<2>Contrasting Views of the Internal Dynamics of the HhaI Methyltransferase Target DNA Reported by Solution and Solid-State NMR Spectroscopy.
<3>J. Am. Chem. Soc.
<4>128
<5>15970-15971
<6>2006
<7>The enzymatic methylation of deoxyribonucleic acid is essential for many biological processes
and has therefore been studied in considerable detail.  The structure of the ternary complex
of the HhaI methyltransferase with its DNA target [5'-(dGATAGCGCTATC)-3']2 and the
methyl-donating cofactor S-adenosyl L-methionine demonstrated that the substrate cytosine is
extruded from its normally base-paired position.  Through this confirmational change, the
carbon at the C5 position on the base of the underlined cytosine becomes accessible in the
enzyme's catalytic pocket.  The interactions observed between the protein and the extruded
base explain the stabilization of this highly distorted structure but do not provide a
mechanism or pathway to flip the base outside of the double helix.  What are the energetically
favorable pathways that allow base extrusion?  Are they sequence dependent?

<>

<1>Miller, P.B., Wakarchuk, W.W., Warren, R.A.J.
<2>Alpha-putrescinylthymine and the sensitivity of bacteriophage Phi W-14 DNA to restriction endonucleases.
<3>Nucleic Acids Res.
<4>13
<5>2559-2568
<6>1985
<7>The modified base alpha-putrescinylthymine (putT) in Phi W-14 DNA blocks
cleavage of the DNA by 17 of 32 Type II restriction endonucleases.  The enzymes
cleaving the DNA do so to widely varying extents.  The frequencies of cleavage
of three altered forms of the DNA show that putT blocks recognition sites
either when it occurs within the site or when it is in a sequence flanking the
site.  The blocking is dependent on both charge and steric factors.  The charge
effects can be greater than the steric effects for some of the enzymes tested.
All the enzymes cleaving Phi W-14 DNA release discrete fragments, showing that
the distribution of putT is ordered.  The cleavage frequencies for different
enzymes suggest that the sequence CAputTG occurs frequently in DNA.  Only TaqI
of the enzymes tested appeared not to be blocked by putT, but it was slowed
down.  TaqI generated fragments are joinable by T4 DNA ligase.

<>

<1>Miller, T.R., Delcher, A.L., Salzberg, S.L., Saunders, E., Detter, J.C., Halden, R.U.
<2>Genome Sequence of the Dioxin-Mineralizing Bacterium Sphingomonas wittichii RW1.
<3>J. Bacteriol.
<4>192
<5>6101-6102
<6>2010
<7>Pollutants such as polychlorinated biphenyls and dioxins pose a serious threat to human and
environmental health. Natural attenuation of these
compounds by microorganisms provides one promising avenue for their
removal from contaminated areas. Over the past 2 decades, studies of the
bacterium Sphingomonas wittichii RW1 have provided a wealth of knowledge
about how bacteria metabolize chlorinated aromatic hydrocarbons. Here we
describe the finished genome sequence of S. wittichii RW1 and major
findings from its annotation.

<>

<1>Miller, W.G.
<2>Characterization of multiple Campylobacter jejuni type I restriction-modification loci.
<3>Int. J. Med. Microbiol.
<4>291
<5>79
<6>2001
<7>Type I restriction-modification systems have been found in many bacterial taxa.  Type I
enzymes are composed of three subunits, encoded by the hsdR, hsdS, and hsdM genes.  All three
gene products are necessary for restriction, whereas the hsdM and hsdS gene products are
sufficient for methylation.  To characterize the Campylobacter jejuni type I
restriction-modification systems, the hsd loci from eight C. jejuni strains were cloned and
sequence.  The enteric hsd genes and the H. pylori hsd genes are contiguous; however, in C.
jejuni, intervening ORFs are present between hsdR and hsdS and between hsdS and hsdM.  The
function of the "RS"  ORF is unknown and the "SM" ORF probably serves no function since it is
absent in at least two loci.  Based on DNA homology and complementation, the enteric hsd loci
have been subdivided into five families (IA, IB, IC, ID, and IE).  The hsd loci of C. jejuni
can also be divided into families: BLAST analysis suggests that the hsd locus of NCTC 81116 is
homologous to the IB family and the hsd loci of NCTC 11168 and 81-176 are homologous to the ID
family.  The hsdM gene of C. jejuni RM1221 ("IB") is more homologous to its enteric
counterparts (E-e-74) than to the hsdM genes of NCTC 11168 (E=e-24) and H. pylori 26695
(E=e-22).  To determine if additional families exist or if some strains have multiple loci, 39
C. jejuni strains were amplified with "IB" - and "ID"-specific probes.  Twelve strains could
not be assigned to either the "IB" or "ID" family, suggesting that at least one additional
family may exist.  Also, no strain amplified with both the "IB" and "ID" probes; however, the
presence of one or more strains that contain multiple loci of the same family cannot be
eliminated.

<>

<1>Miller, W.G., Keech, A.M., Pearson, B.M., Wells, J.M., Kapitonov, V.V., Konkel, M.E., Mandrell, R.E.
<2>Diversity of Campylobacter type I restriction-modification loci: Induction of hsdS by exogenous DNA.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>104
<5>210
<6>2004
<7>Restriction-modification systems are found in many bacterial taxa and are believed to provide
a barrier against foreign DNA and bacteriophages.  R-M systems have been classified into three
types, type I, II, and III.  The type I enzyme is a bifunctional, multi-subunit complex
containing HsrR, HsdS, and HsdM.  Restriction and modification are represented by the HsdR and
HsdM proteins, respectively.  HsdS interacts with the target sequence as part of both the
restriction and modification complexes.  The type I restriction-modification (hsd) genes of 62
Campylobacter jejuni strains were characterized by DNA sequencing and amplification.  No
evidence was found that C. jejuni strains contain multiple hsd loci.  Unlike many
characterized hsd loci from other taxa, intervening open reading frames are present between
hsdS and the hsdR and hsdM genes.  These ORFs, designated as rlo (R-linked ORF) and mlo
(M-linked ORF), have no defined function but association of rlo genes with particular hsdS
alleles may suggest a role in R-M function.  Based on parsimony analysis of amino-acid
sequences, the C. jejuni hsd loci were assigned to one of three families: 'IAB', 'IC', or
'IF'.  HsdM proteins within a family are strongly conserved but share little homology with
HsdM proteins from the other two families.  Also, there is significant diversity within the
'IAB' hsd family: 8 different 'IAB' hsdS alleles and 7 different 'IAB' rlo alleles have
been characterized.  Finally, RT-PCR analysis using hsdS-specific primer sets was used to
assess whether the unique hsdS alleles were expressed.  Expression of only two hsdS alleles
was detected by RT-PCR from cultures grown on rich media; however, expression of 7 additional
hsdS alleles was detected after the addition of exogenous DNA to the C. jejuni cells.

<>

<1>Miller, W.G., Pearson, B.M., Wells, J.M., Parker, C.T., Kapitonov, V.V., Mandrell, R.E.
<2>Diversity within the Campylobacter jejuni type I restriction - modification loci.
<3>Microbiology
<4>151
<5>337-351
<6>2005
<7>The type I restriction-modification (hsd) systems of 73 Campylobacter jejuni strains were
characterized according to their DNA and amino acid
sequences, and/or gene organization. A number of new genes were
identified which are not present in the sequenced strain NCTC 11168.
The closely related organism Helicobacter pylori has three type I
systems; however, no evidence was found that C. jejuni strains contain
multiple type I systems, although hsd loci are present in at least two
different chromosomal locations. Also, unlike H. pylori, intervening
ORFs are present, in some strains, between hsdR and hsdS and between
hsdS and hsdM. No definitive function can be ascribed to these ORFs,
designated here as rloA-H ((R) under bar-(l) under bar inked (O) under
bar RF) and mloA-B ((M) under bar-(l) under bar inked (O) under bar
RF). Based on parsimony analysis of amino acid sequences to assess
character relatedness, the C. jejuni type I R-M systems are assigned to
one of three families: 'IAB', 'IC' or 'IF'. This study confirms that
HsdM proteins within a family are highly conserved but share little
homology with HsdM proteins from other families. The 'IC' hsd loci are
> 99% identical at the nucleoticle level, as are the 'IF' hsd loci.
Additionally, whereas the nucleoticle sequences of the 'IAB' hsdR and
hsdM genes show a high degree of similarity, the nucleoticle sequences
of the 'IAB' hsdS and no genes vary considerably. This diversity
suggests that recombination between 'IAB' hsd loci would lead not only
to new hsdS alleles but also to the exchange of no genes; five C.
jejuni hsd loci are presumably the result of such recombination. The
importance of these findings with regard to the evolution of C. jejuni
type I R-M systems is discussed.

<>

<1>Miller, W.G., Wang, G., Binnewies, T.T., Parker, C.T.
<2>The complete genome sequence and analysis of the human pathogen Campylobacter lari.
<3>Foodborne Pathog. Dis.
<4>5
<5>371-386
<6>2008
<7>Campylobacter lari is a member of the epsilon subdivision of the Proteobacteria and is part of
the thermotolerant Campylobacter group, a
clade that includes the human pathogen C. jejuni. Here we present the
complete genome sequence of the human clinical isolate, C. lari RM2100.
The genome of strain RM2100 is approximately 1.53 Mb and includes the 46
kb megaplasmid pCL2100. Also present within the strain RM2100 genome is a
36 kb putative prophage, termed CLIE1, which is similar to CJIE4, a
putative prophage present within the C. jejuni RM1221 genome. Nearly all
(90%) of the gene content in strain RM2100 is similar to genes present in
the genomes of other characterized thermotolerant campylobacters. However,
several genes involved in amino acid biosynthesis and energy metabolism,
identified previously in other Campylobacter genomes, are absent from the
C. lari RM2100 genome. Therefore, C. lari RM2100 is predicted to be
multiply auxotrophic, unable to synthesize eight different amino acids,
acetyl-coA, and pantothenate. Additionally, strain RM2100 does not contain
a complete TCA cycle and is missing the CydAB terminal oxidase of the
respiratory chain. Defects in the amino acid biosynthetic pathways in this
organism could be potentially compensated by the large number of encoded
peptidases. Nevertheless, the apparent absence of certain key enzymatic
functions in strain RM2100 would be expected to have an impact on C. lari
biology. It is also possible that the reduction in the C. lari metabolic
machinery is related to its environmental range and host preference.

<>

<1>Mills, D.A., Manias, D.A., McKay, L.L., Dunny, G.M.
<2>Homing of a group II intron from Lactococcus lactis subsp. lactis ML3.
<3>J. Bacteriol.
<4>179
<5>6107-6111
<6>1997
<7>Ll.ltrB is a functional group II intron located within a gene (ltrB) encoding a conjugative
relaxase essential for transfer of the lactococcal element pRS01.  In this work, the Ll.ltrB
intron was shown to be an independent mobile element capable of inserting into an intronless
allele of the ltrB gene.  Ll.ltrB was not observed to insert into a deletion derivative of the
ltrB gene in which the intron splice site was removed. In contrast, a second vector containing
a 271-nucleotide segment of ltrB spanning the Ll.ltrB splice site was shown to be a proficient
recipient of intron insertion.  Efficient homing was observed in the absence of a functional
host homologous recombination system.  This work demonstrates that the Ll.ltrB intron is a
novel site-specific mobile element in lactococci and that group II intron self-transfer is a
mechanism for intron dissemination among bacteria.

<>

<1>Mills, D.A., McKay, L.L., Dunny, G.M.
<2>Splicing of a group II intron involved in the conjugative transfer of pRS01 in Lactococci.
<3>J. Bacteriol.
<4>178
<5>3531-3538
<6>1996
<7>Analysis of a region involved in the conjugative transfer of the lactococcal conjugative
element pRS01 has revealed a bacterial group II intron.  Splicing of this lactococcal intron
(designated Ll.ltrB) in vivo resulted in the ligation of two exon messages (ltrBE1 and ltrBE2)
which encoded a putative conjugative relaxase essential for the transfer of pRS01.  Like many
group II introns, the Ll.ltrB intron possessed an open reading frame (ltrA) with homology to
reverse transcriptases.  Remarkably, sequence analysis of ltrA suggested a greater similarity
to open reading frames encoded by eukaryotic mitochondrial group II introns than to those
identified to date from other bacteria.  Several insertional mutations within ltrA resulted in
plasmids exhibiting a conjugative transfer-deficient phenotype.  These results provide the
first direct evidence for splicing of a prokaryotic group II intron in vivo and suggest that
conjugative transfer is a mechanism for group II intron dissemination in bacteria.

<>

<1>Mills, H.J., Eisen, J., Sobecky, P.A.
<2>Insights into the role of cryptic marine plasmids based on sequence analysis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>103
<5>N-147
<6>2003
<7>Knowledge has been gained from the intensive study of a limited group of bacterial plasmids
particularly those from clinical settings and
terrestrial environments. The molecular characterization of plasmid
populations occurring in a wider range of habitats, especially marine,
is necessary to provide knowledge on plasmid ecology and their
contributions to the genetic plasticity of microbial communities. DNA
sequencing and analysis have greatly facilitated the determination of
putative functions to otherwise 'cryptic' marine plasmids. In
collaboration with TIGR, we have undertaken whole plasmid sequencing to
elucidate putative gene functions to gain a better understanding of
plasmid diversity and ecological roles. A random shotgun method was
used to obtain DNA sequences from five 'cryptic' marine plasmids
ranging in size from 9-kb to 80-kb from Vibrio and Shewanella. A
comparative and systematic analysis of the marine plasmid sequences
have indicated that p172, a 28.8-kb plasmid from marine Vibrio sp. 172
encodes partitioning elements, colicin-mediated cell killing,
restriction modification systems, and natural competence. A co-resident
replicon in Vibrio sp. 172, p172-A, is a 9.0-kb element that appears to
be the replicative form of a previously identified Vibrio
parahaemolyticus phage. A second marine Vibrio strain designated 09022,
contains a 31.0-kb plasmid, p09022, that remains mostly cryptic,
however p09022 does encode partitioning elements and a restriction
modification system related to similar genes on p172. A majority of the
putative genes from p23023 (52.1-kb) resident in a third Vibrio sp.,
and p0908 (81.4-kb) isolated from a Shewenella sp., possesses no
homology to any known ORFs. However, p23023 encodes a putative cell
adhesion operon, previously reported in non-marine bacterial hosts.
Sequence analysis demonstrates the capacity to assign putative function
to cryptic plasmids, but also exemplifies the need for additional
sequencing and analyzing of marine genes. Such plasmid-encoded
adhesion, competency and colicin production traits likely provide hosts
with competitive advantages in marine ecosystems.

<>

<1>Mills, K.I., Ramsahoye, B.H.
<2>Methods in Molecular Biology. DNA methylation protocols.
<3>Methods Mol. Biol.
<4>200
<5>1-7
<6>2002
<7>This book provides a set of reproducible protocols for the analysis of DNA methylation and
methylases. Each technique includes a summary of the basic
theory, a materials list, step-by-step instructions, and notes for
avoiding pitfalls. It was written for all researchers investigating
replication, transcription, growth, differentiation, and carcinogenesis.
Bibliographical references, illustrations, and an index are included.

<>

<1>Mills, K.V., Paulus, H.
<2>Biochemical mechanisms of intein-mediated protein splicing.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M. et al., Berlin Heidelberg
<4>16
<5>233-255
<6>2005
<7>This chapter discusses the mechanism of the self-catalyzed process by which inteins promote
both their own excision from a host protein and the direct linkage of the flanking host
protein segments, the N- and C-exteins, by a peptide bond.  The majority of inteins have a
nucleophilic amino acid at their N-terminus and asparagine at their C-terminus and are linked
to a C-extein with an N-terminal nucleophilic amino acid.  These canonical inteins promote
protein splicing by a four-step mechanism of sequential acyl rearrangements.  Non-canonical
inteins, which lack either the N-terminal nucleophile or the C-terminal asparagine, promote
protein splicing by a variant of this mechanism or promote protein cleavage rather than
splicing.  A remarkable feature of the protein splicing process is that it involves multiple
steps that are chemically autonomous yet proceed in a highly coordinated manner without side
reactions unless perturbed by mutation, unnatural exteins, or non-physiological conditions.
The factors that may serve to integrate protein splicing into a system that ordinarily
operates efficiently without side reactions are discussed.

<>

<1>Mills, S., Griffin, C., O'Sullivan, O., Coffey, A., McAuliffe, O.E., Meijer, W.C., Serrano, L.M., Ross, R.P.
<2>A new phage on the 'Mozzarella' block: Bacteriophage 5093 shares a low level of homology with other Streptococcus thermophilus phages.
<3>Int. Dairy Journal
<4>21
<5>963-969
<6>2011
<7>Streptococcus thermophilus bacteriophage 5093 is a virulent phage that infects the industrial
Mozzarella starter CSK939. The genome of phage
5093 is 37,184 base pairs (bps) containing 50 open reading frames
(orfs). Genetic analysis revealed that the genome of phage 5093 is
highly mosaic when compared with other sequenced S. thermophilus
phages. This is particularly apparent in the late gene cluster with
regions displaying high homology to prophage sequences of non-dairy
streptococci and limited homology to either pac or cos-type S.
thermophilus phages. In addition, a definitive antireceptor gene was
not observed - suggesting that phage 5093 may have developed a
different system for host recognition. Interestingly, the phage does
contain a methylase domain that probably evolved as a phage
counter-defence mechanism. These findings suggest that phage 5093 may
represent a third group of S. thermophilus phage and provide the link
between phages that infect S. thermophilus and its non-dairy ancestors.

<>

<1>Mills, S., McAuliffe, O.E., Coffey, A., Fitzgerald, G.F., Ross, R.P.
<2>Plasmids of lactococci - genetic accessories or genetic necessities?
<3>FEMS Microbiol. Rev.
<4>30
<5>243-273
<6>2006
<7>Lactococci are one of the most exploited microorganisms used in the manufacture of food.
These intensively used cultures are generally characterized by having a rich plasmid
complement.  It could be argued that it is the plasmid complement of commercially utilized
cultures that gives them their technical superiority and individuality.  Consequently, it is
timely to reflect on the desirable characteristics encoded on lactococcal plasmids.  It is
argued that plasmids play a key role in the evolution of modern starter strains and are a lot
more than just selfish replicosomes but more essential necessities of intensively used
commercial starters.  Moreover, the study of plasmid biology provides a genetic blueprint that
has proved essential for the generation of molecular tools for the genetic improvement of
Lactococcus lactis.

<>

<1>Milsom, S.E., Halford, S.E., Embleton, M.L., Szczelkun, M.D.
<2>Analysis of DNA looping interactions by type II restriction enzymes that require two copies of their recognition sites.
<3>J. Mol. Biol.
<4>311
<5>515-527
<6>2001
<7>Before cleaving DNA substrates with two recognition sites, the Cfr10I, NgoMIV, NaeI and SfiI
restriction endonucleases bridge the two sites
through 3D space, looping out the intervening DNA. To characterise
their looping interactions, the enzymes were added to plasmids with two
recognition sites interspersed with two res sites for site-specific
recombination by Tn21 resolvase, in buffers that contained either EDTA
or CaCl2 so as to preclude DNA cleavage by the endonuclease; the extent
to which the res sites were sequestered into separate loops was
evaluated from the degree of inhibition of resolvase. With Cfr10I, a
looped complex was detected in the presence but not in the absence of
Ca2+; it had a lifetime of about 90 seconds. Neither NgoMIV nor NaeI
gave looped complexes of sufficient stability to be detected by this
method. In contrast, SfiI with Ca++ produced a looped complex that survived for more than
seven hours, whereas its looping interaction in
EDTA lasts for about four minutes. When resolvase was added to a SfiI
binding reaction in EDTA followed immediately by CaCl2, the looped DNA
was blocked from recombination while the unlooped DNA underwent
recombination. By measuring the distribution between looped and
unlooped DNA at various SfiI concentrations, and by fitting the data to
a model for DNA binding by a tetrameric protein to two sites in cis, an
equilibrium constant for the looping interaction was determined. The
equilibrium constant was essentially independent of the length of DNA
between the SfiI sites.

<>

<1>Minarovits, J.
<2>MICROBE-INDUCED EPIGENETIC ALTERATIONS IN HOST CELLS: THE COMING ERA OF PATHO-EPIGENETICS OF MICROBIAL INFECTIONS A REVIEW.
<3>Acta Microbiol. Immunol. Hung.
<4>56
<5>1-19
<6>2009
<7>It is well documented that the double-stranded DNA (dsDNA) genomes of certain viruses and the
proviral genomes of retroviruses are regularly
targeted by epigenetic regulatory mechanisms (DNA methylation, histone
modifications, binding of regulatory proteins) in infected cells. In
parallel, proteins encoded by viral genomes may affect the activity of
a set of cellular promoters by interacting with the very same
epigenetic regulatory machinery. This may result in epigenetic
dysregulation and subsequent cellular dysfunctions that may manifest in
or contribute to the development of pathological changes (e. g.
initiation and progression of malignant neoplasms; immunodeficiency).
Bacteria infecting mammals may cause diseases in a similar manner, by
causing hypermethylation of key cellular promoters at CpG dinucleotides
(promoter silencing, e. g. by Campylobacter rectus in the placenta or
by Helicobacter pylori in gastric mucosa). I suggest that in addition
to viruses and bacteria, other microparasites (protozoa) as well as
macroparasites (helminths, arthropods, fungi) may induce pathological
changes by epigenetic reprogramming of host cells they are interacting
with. Elucidation of the epigenetic consequences of microbe-host
interactions (the emerging new field of patho-epigenetics) may have
important therapeutic implications because epigenetic processes can be
reverted and elimination of microbes inducing patho-epigenetic changes
may prevent disease development.

<>

<1>Minczuk, M., Papworth, M.A., Kolasinska, P., Murphy, M.P., Klug, A.
<2>Sequence-specific modification of mitochondrial DNA using a chimeric zinc finger methylase.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>19689-19694
<6>2006
<7>We used engineered zinc finger peptides (ZFPs) to bind selectively to predetermined sequences
in human mtDNA. Surprisingly, we found that
engineered ZFPs cannot be reliably routed to mitochondria by using only
conventional mitochondrial targeting sequences. We here show that addition
of a nuclear export signal allows zinc finger chimeric enzymes to be
imported into human mitochondria. The selective binding of
mitochondria-specific ZFPs to mtDNA was exemplified by targeting the
T8993G mutation, which causes two mitochondrial diseases, neurogenic
muscle weakness, ataxia, and retinitis pigmentosa (NARP) and also
maternally inherited Leigh's syndrome. To develop a system that allows the
monitoring of site-specific alteration of mtDNA we combined a ZFP with the
easily assayed DNA-modifying activity of hDNMT3a methylase. Expression of
the mutation-specific chimeric methylase resulted in the selective
methylation of cytosines adjacent to the mutation site. This is a proof of
principle that it is possible to target and alter mtDNA in a
sequence-specific manner by using zinc finger technology.

<>

<1>Minegishi, K., Aikawa, C., Furukawa, A., Watanabe, T., Nakano, T., Ogura, Y., Ohtsubo, Y., Kurokawa, K., Hayashi, T., Maruyama, F., Nakagawa, I., Eishi, Y.
<2>Complete Genome Sequence of a Propionibacterium acnes Isolate from a Sarcoidosis  Patient.
<3>Genome Announcements
<4>1
<5>e00016-12
<6>2013
<7>Propionibacterium acnes is a human skin commensal that resides preferentially within sebaceous
follicles and is the only microorganism that has been isolated from sarcoid lesions. We report
the complete genome sequence of P. acnes, which was isolated from a Japanese patient with
sarcoidosis.

<>

<1>Miner, Z.
<2>Comparative analysis of the T2 and T4 DNA [N6-adenine] methyltransferase (dam) genes and characterization of single amino acid substitutions that alter the sequence specificity of their encoded proteins.
<3>Diss. Abstr.
<4>51
<5>588B
<6>1990
<7>Bacteriophage T4 encodes a DNA-[N6-adenine]methyltransferase (Dam) which recognizes primarily
the sequence GATC in both cytosine- and hydroxymethylcytosine-containing DNA. The
corresponding enzyme encoded by the related bacteriophage T2 is able to methylate both
substrates to a greater extent than T4 Dam. The T2 dam gene has been cloned and sequenced, and
the sequence compared with that of the T4 dam gene. From the coding region, I infer there are
three amino acid differences, two of which are located in a region of homology (I) that is
shared by T2 and T4 Dam, Escherichia coli Dam, and the modification enzyme of Streptococcus
pneumoniae, all of which methylate the sequence 5' GATC 3'. Although the T2 and T4 dam
promoters are not identical, gene fusion experiments indicate that the T4 promoter produces
about two-fold more Beta-galactosidase activity than does the T2 promoter. T2 and T4 dam give
rise to hypermethylating mutants (damh) which exhibit an alteration in sequence specificity;
that is, they are readily able to methylate non-canonical sites. I have determined that the
damh mutation in T4 produces a single amino acid change (Pro126 to Ser126) in another region
of homology (III) shared by the four DNA-adenine methyltransferases. Another mutant, damc, is
described which methylates GATC in cytosine-containing DNA, but not in
hydroxymethylcytosine-containing DNA. This mutation alters a single amino acid (Phe127 to
Val127). The effect of several different amino acids at residue 126 was examined by creating
an amber codon at that position and comparing the methylation capability of partially purified
enzymes produced in the presence of various suppressors. No enzyme activity is observed for
proteins containing phenylalanine, glutamic acid, or histidine at position 126. However,
insertion of alanine, cysteine, or glycine at residue 126 produces enzymatic activity similar
to that of Damh. These results suggest that at least two regions of the Dam protein are
involved in sequence recognition. These regions may be in close proximity to one another in
the native protein to form a domain that is involved in nucleotide recognition and protein-DNA
interation.

<>

<1>Miner, Z., Hattman, S.
<2>Molecular cloning, sequencing, and mapping of the bacteriophage T2 dam gene.
<3>J. Bacteriol.
<4>170
<5>5177-5184
<6>1988
<7>Bacteriophage T2 codes for a DNA-(adenine-N6) methyltransferase (Dam), which is able to
methylate both cytosine- and hydroxymethylcytosine-containing DNAs to a greater extent than
the corresponding methyltransferase encoded by bacteriophage T4. We have cloned and sequenced
the T2 dam gene and compared it with the T4 dam gene. In the Dam coding region, there are 22
nucleotide differences, 4 of which result in three coding differences (2 are in the same
codon). Two of the amino acid alterations are located in a region of homology that is shared
by T2 and T4 Dam, Escherichia coli Dam, and the modification enzyme of Streptococcus
pneumoniae, all of which methylate the sequence 5'GATC3'. The T2 dam and T4 dam promoters
are not identical and appear to have slightly different efficiencies; when fused to the E.
coli lacZ gene, the T4 promoter produces about twofold more beta-galactosidase activity than
does the T2 promoter. In our first attempt to isolate T2 dam, a truncated gene was cloned on a
1.67-kilobase XbaI fragment. This construct produces a chimeric protein composed of the first
163 amino acids of T2 Dam followed by 83 amino acids coded by the pUC18 vector. Surprisingly,
the chimera has Dam activity, but only on cytosine-containing DNA. Genetic and physical
analyses place the T2 dam gene at the same respective map location as the T4 dam gene.
However, relative to T4, T2 contains an insertion of 536 base pairs 5' to the dam gene.
Southern blot hybridization and computer analysis failed to reveal any homology between this
insert and either T4 or E. coli DNA.

<>

<1>Miner, Z., Schlagman, S., Hattman, S.
<2>Single amino acid changes which alter the sequence specificity of the T4 and T2 (Dam) DNA-adenine methyltransferases.
<3>Gene
<4>74
<5>275-276
<6>1988
<7>Meeting Abstract

<>

<1>Miner, Z., Schlagman, S., Hattman, S.
<2>Single amino acid changes which alter the sequence specificity of the T4 (dam) DNA-adenine methyltransferase.
<3>Biochem. Pharmacol.
<4>37
<5>1811-1812
<6>1988
<7>Many enteric bacteria contain a DNA adenine methyltransferase (Dam) that methylates the A
residue in the sequence, GATC. The related T-even phages, T2 and T4 (but not T6), also encode
Dam methylases; the normal substrate for these enzymes is 5-hydroxymethylcytosine
(hmC)-containing DNA, since these viruses contain this base in place of C, and the hmC is
modified further by glucosylation. Nonglucosylating mutants (gt-) have been isolated that are
different from their gt+ parents in that they are unable to grow on certain strains, such as
P1 lysogenic hosts. Derivatives of T2 gt- and T4 gt- phage capable of growth on P1 lysogens
have been isolated; these are designated damh because they exhibit hypermethylation of their
DNA. Thus, Damh, but not Dam+, methylation protects against P1 restriction of the asymmetric
sequence, AGACC.

<>

<1>Miner, Z., Schlagman, S., Hattman, S.
<2>The dam DNA-methyltransferases of E. coli and its phages T2 and T4.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>199
<6>1989
<7>DNA (adenine-N6) methyltransferases (MTases) recognizing the palindromic tetranucleotide
sequence, 5'-GATC-3', are encoded by (dam) genes of various bacteriophages (e.g.
T2,T4,T1,P1) and bacteria (e.g. Escherichia, Salmonella, Neisseria, Streptococcus).  Cloning
and nucleotide sequence analysis has revealed that these Dam polypeptides contain several
regions of considerable amino acid sequence homology; and one of these regions (IV) contains a
sequence motif, (Asp/Asn)-Pro-Pro-(Phe/Tyr), also found in MTases that methylate adenine in
sequences other than GATC.  The conservation of amino acid sequences among these enzymes
suggests that they are in domains important for MTase function and specificity; i.e. substrate
(S-adenosylmethionine, AdoMet) and DNA-nucleotide sequence recognition/interaction.  We have
been focusing on the Dam MTases of phages T2 and T4, particularly because mutants (dam^h) are
known that methylate DNA to higher extents than the enzyme from the wild-type (dam+) parent.
In addition, second site mutants (dam^h dam-x) have been isolated that abolish all DNA
methylation ability.  The normal substrate for the phage DNA MTases is DNA containing
5-hydroxymethylcytosine (hmC), because the viruses contain this base in place of C; however,
C-DNA is still a good substrate for these enzymes.  The two wild-type phage MTases differ in
that the T2 enzyme adds about twice as many methyl groups per unit DNA than does the T4
enzyme.  A comparison of the wild-type T2 and T4 Dam+ encoded polypeptides revealed three
amino acid differences; viz., at positions 20,26,188.  The latter is a conserved changed
(Asp->Glu) and does not appear to be involved in sequence specificity.  The other two changes
are located in homology region I, implicating this domain in nucleotide sequence recognition.
Current efforts to prove this include production of chimeric enzymes and site-directed
mutagenesis.  We have shown that the dam+ -> dam^h mutation produces a Pro ->Ser change at
amino acid residue 126.  A Phe -> Val change at residue 127 prevents the MTase from
methylating hmC-DNA, but not C-DNA.  These two residues are contained in homology region III,
also implicating this domain in nucleotide sequence recognition.

<>

<1>Miner, Z., Schlagman, S.L., Hattman, S.
<2>Single amino acid changes that alter the DNA sequence specificity of the DNA-[N/6-adenine] methyltransferase (Dam) of bacteriophage T4.
<3>Nucleic Acids Res.
<4>17
<5>8149-8157
<6>1989
<7>Bacteriophage T4 codes for a DNA-[N/6-adenine] methyltransferase (Dam) which
recognizes primarily the sequence GATC in both cytosine-and
hydroxymethylcytosine-containing DNA.  Hypermethylating mutants, dam/h, exhibit
a relaxation in sequence specificity, that is, they are readily able to
methylate non-canonical sites.  We have determined that the dam/h mutation
produces a single amino acid change (Pro/126 to Ser/126) in a region of
homology (III) shared by three DNA-adenine methyltransferases; viz, T4 Dam,
Escherichia coli Dam, and the DpnII modification enzyme of Streptococcus
pneumoniae.  We also describe another mutant, dam/c, which methylates GATC in
cytosine-containing DNA, but not in hydroxymethylcytosine-containing DNA.  This
mutation also alters a single amino acid (Phe/127 to Val/127).  These results
implicate homology region III as a domain involved in DNA sequence recognition.
The effect of several different amino acids at residue 126 was examined by
creating a polypeptide chain terminating codon at that position and comparing
the methylation capability of partially purified enzymes produced in the
presence of various suppressors.  No enzyme activity is detected when
phenylalanine, glutamic acid, or histidine is inserted at position 126.
However, insertion of alanine, cysteine, or glycine at residue 126 produces
enzymatic activity similar to Dam/h.

<>

<1>Minion, F.C., Lefkowitz, E.J., Madsen, M.L., Cleary, B.J., Swartzell, S.M., Mahairas, G.G.
<2>The genome sequence of Mycoplasma hyopneumoniae strain 232, the agent of swine mycoplasmosis.
<3>J. Bacteriol.
<4>186
<5>7123-7133
<6>2004
<7>We present the complete genome sequence of Mycoplasma hyopneumoniae, an important member of
the porcine respiratory disease complex. The genome is
composed of 892,758 bp and has an average G+C content of 28.6 mol%. There
are 692 predicted protein coding sequences, the average protein size is
388 amino acids, and the mean coding density is 91%. Functions have been
assigned to 304 (44%) of the predicted protein coding sequences, while 261
(38%) of the proteins are conserved hypothetical proteins and 127 (18%)
are unique hypothetical proteins. There is a single 16S-23S rRNA operon,
and there are 30 tRNA coding sequences. The cilium adhesin gene has six
paralogs in the genome, only one of which contains the cilium binding
site. The companion gene, P102, also has six paralogs. Gene families
constitute 26.3% of the total coding sequences, and the largest family is
the 34-member ABC transporter family. Protein secretion occurs through a
truncated pathway consisting of SecA, SecY, SecD, PrsA, DnaK, Tig, and
LepA. Some highly conserved eubacterial proteins, such as GroEL and GroES,
are notably absent. The DnaK-DnaJ-GrpR complex is intact, providing the
only control over protein folding. There are several proteases that might
serve as virulence factors, and there are 53 coding sequences with
prokaryotic lipoprotein lipid attachment sites. Unlike other mycoplasmas,
M. hyopneumoniae contains few genes with tandem repeat sequences that
could be involved in phase switching or antigenic variation. Thus, it is
not clear how M. hyopneumoniae evades the immune response and establishes
a chronic infection.

<>

<1>Minko, I., Hattman, S., Lloyd, R.S., Kossykh, V.
<2>Methylation by a mutant T2 DNA [N6-adenine] methyltransferase expands the usage of RecA-assisted endonuclease (RARE) cleavage.
<3>Nucleic Acids Res.
<4>29
<5>1484-1490
<6>2001
<7>Properties of a mutant bacteriophage T2 DNA [N6-adenine] methyltransferase (T2 Dam MTase) have
been investigated for its potential utilization in RecA-assisted restriction endonuclease
(RARE) cleavage. Steady-state kinetic analyses with oligonucleotide duplexes revealed that,
compared to wild-type T4 Dam, both wild-type T2 Dam and mutant T2 Dam P126S had a 1.5-fold
higher kcat in methylating canonical GATC sites. Additionally, T2 Dam P126S showed increased
efficiencies in methylation of non-canonical GAY sites relative to the wild-type enzymes. In
agreement with these steady-state kinetic data, when bacteriophage  DNA was used as a
substrate, maximal protection from restriction nuclease cleavage in vitro was achieved on the
sequences GATC, GATN and GACY, while protection of GACR sequences was less efficient.
Collectively, our data suggest that T2 Dam P126S can modify 28 recognition sequences. The
feasibility of using the mutant enzyme in RARE cleavage with BclI and EcoRV endonucleases has
been shown on phage  DNA and with BclI and DpnII endonucleases on yeast chromosomal DNA
embedded in agarose.

<>

<1>Minogue, T.D. et al.
<2>Whole-genome sequences of 24 Brucella strains.
<3>Genome Announcements
<4>2
<5>e00915-14
<6>2014
<7>Brucella species are intracellular zoonotic pathogens which cause, among other pathologies,
increased rates of abortion in ruminants. Human infections are
generally associated with exposure to contaminated and unpasteurized dairy
products; however Brucellae have been developed as bioweapons. Here we present 17
complete and 7 scaffolded genome assemblies of Brucella strains.

<>

<1>Minogue, T.D., Daligault, H.A., Davenport, K.W., Bishop-Lilly, K.A., Broomall, S.M., Bruce, D.C., Chain, P.S., Chertkov, O., Coyne, S.R., Freitas, T., Frey, K.G., Gibbons, H.S., Jaissle, J., Redden, C.L., Rosenzweig, C.N., Xu, Y., Johnson, S.L.
<2>Complete Genome Assembly of Streptococcus pyogenes ATCC 19615, a Group A beta-Hemolytic Reference Strain.
<3>Genome Announcements
<4>2
<5>e00976-14
<6>2014
<7>We present the complete genome assembly of Streptococcus pyogenes ATCC 19615 (Rosenbach) as
submitted to GenBank under accession number CP008926. This group A
nonmotile beta-hemolytic clinical isolate is used for quality control in a
variety of commercially available tests. The assembled genome is 1.84 Mb (38.5%
G+C content) and contains 1,788 coding regions.

<>

<1>Minogue, T.D., Daligault, H.A., Davenport, K.W., Bishop-Lilly, K.A., Broomall, S.M., Bruce, D.C., Chain, P.S., Chertkov, O., Coyne, S.R., Freitas, T., Frey, K.G., Gibbons, H.S., Jaissle, J., Redden, C.L., Rosenzweig, C.N., Xu, Y., Johnson, S.L.
<2>Complete Genome Assembly of Escherichia coli ATCC 25922, a Serotype O6 Reference  Strain.
<3>Genome Announcements
<4>2
<5>e00969-14
<6>2014
<7>We present the complete genome assembly of Escherichia coli ATCC 25922 as submitted to NCBI
under accession no. CP009072. This strain was originally
isolated from a clinical sample in Seattle, Washington (1946), and is often used
in quality control testing. The assembled genome is 5.20 Mb (50.4% G+C content)
and includes two plasmids.

<>

<1>Minogue, T.D., Daligault, H.A., Davenport, K.W., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Chertkov, O., Coyne, S.R., Freitas, T., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Palacios, G.F., Redden, C.L., Xu, Y., Johnson, S.L.
<2>Complete Genome Assembly of Reference Strain Ochrobactrum anthropi ATCC 49687.
<3>Genome Announcements
<4>2
<5>e00962-14
<6>2014
<7>Ochrobactrum anthropi is an occasional cause of nosocomial infections; however, interest in
the organism lies in its phylogenetic proximity to the genus
Brucella. Here, we present the 4.9-Mb finished genome of Ochrobactrum anthropi
ATCC 49687, most commonly used as an exclusionary reference organism.

<>

<1>Minogue, T.D., Daligault, H.A., Davenport, K.W., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Chertkov, O., Coyne, S.R., Freitas, T., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Palacios, G.F., Redden, C.L., Xu, Y., Johnson, S.L.
<2>Draft Genome Assembly of Neisseria lactamica Type Strain A7515.
<3>Genome Announcements
<4>2
<5>e00951-14
<6>2014
<7>We present the scaffolded genome assembly of Neisseria lactamica type strain A7515 (ATCC
23970) as submitted to NCBI under accession no. JOVI00000000. This
type strain of the lactose-fermenting Neisseria species is often used in quality
control testing and intra-genus phylogenetic analyses. The assembly includes four
contigs placed into a single scaffold.

<>

<1>Minogue, T.D., Daligault, H.E., Davenport, K.W., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Chertkov, O., Freitas, T., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Palacios, G.F., Redden, C.L., Xu, Y., Johnson, S.L.
<2>Draft Genome Assemblies of Enterobacter aerogenes CDC 6003-71, Enterobacter cloacae CDC 442-68, and Pantoea agglomerans UA 0804-01.
<3>Genome Announcements
<4>2
<5>e01073-14
<6>2014
<7>The Enterobacteriaceae are environmental and enteric microbes. We sequenced the genomes of two
Enterobacter reference strains, E. aerogenes CDC 6003-71 and E.
cloacae CDC 442-68, as well as one near neighbor used as an exclusionary
reference for diagnostics, Pantoea agglomerans CDC UA0804-01. The genome sizes
range from 4.72 to 5.55 Mbp and have G+C contents from 54.6 to 55.1%.

<>

<1>Minogue, T.D., Daligault, H.E., Davenport, K.W., Bishop-Lilly, K.A., Bruce, D.C., Chain, P.S., Coyne, S.R., Chertkov, O., Freitas, T., Frey, K.G., Jaissle, J., Koroleva, G.I., Ladner, J.T., Palacios, G.F., Redden, C.L., Xu, Y., Johnson, S.L.
<2>Draft Genome Assemblies of Proteus mirabilis ATCC 7002 and Proteus vulgaris ATCC  49132.
<3>Genome Announcements
<4>2
<5>e01064-14
<6>2014
<7>The pleomorphic swarming bacilli of the genus Proteus are common human gut commensal organisms
but also the causative agents of recurrent urinary tract
infections and bacteremia. We sequenced and assembled the 3.99-Mbp genome of
Proteus mirabilis ATCC 7002 (accession no. JOVJ00000000) and the 3.97-Mbp genome
of Proteus vulgaris ATCC 49132 (accession no. JPIX00000000), both of which are
commonly used reference strains.

<>

<1>Minogue, T.D., Daligault, H.E., Davenport, K.W., Broomall, S.M., Bruce, D.C., Chain, P.S., Coyne, S.R., Chertkov, O., Freitas, T., Gibbons, H.S., Jaissle, J., Koroleva, G.I., Ladner, J.T., Palacios, G.F., Rosenzweig, C.N., Xu, Y., Johnson, S.L.
<2>Complete Genome Assembly of Enterococcus faecalis 29212, a Laboratory Reference Strain.
<3>Genome Announcements
<4>2
<5>e00968-14
<6>2014
<7>Enterococcus faecalis is a nonmotile Gram-positive coccus, found both as a commensal organism
in healthy humans and animals and as a causative agent of
multiple diseases, in particular endocarditis. We sequenced the genome of E.
faecalis ATCC 29212, a commonly used reference strain in laboratory studies, to
complete 'finished' annotated assembly (3 Mb).

<>

<1>Minogue, T.D., Daligault, H.E., Davenport, K.W., Broomall, S.M., Bruce, D.C., Chain, P.S., Coyne, S.R., Gibbons, H.S., Jaissle, J., Chertkov, O., Freitas, T., Rosenzweig, C.N., Xu, Y., Johnson, S.L.
<2>Draft Genome Assembly of Pseudomonas aeruginosa Quality Control Reference Strain  Boston 41501.
<3>Genome Announcements
<4>2
<5>e00960-14
<6>2014
<7>We present the scaffolded genome assembly of Pseudomonas aeruginosa Boston 41501, now publicly
available in GenBank (JOVK00000000) in 10 contigs placed into a
single scaffold. The 6.82-Mbp genome contains 66.1% G+C content and 6,295 coding
sequences, including type 4 pilus and type 3 secretion system production genes.

<>

<1>Minot, S., Grunberg, S., Wu, G., Lewis, J., Bushman, F.
<2>Hypervariable loci in the human gut virome.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>109
<5>3962-3966
<6>2012
<7>Genetic variation is critical in microbial immune evasion and drug
resistance, but variation has rarely been studied in complex heterogeneous communities such as
the human microbiome. To begin to study natural variation, we analyzed DNA viruses present in
the lower gastrointestinal tract of 12 human volunteers by determining 48 billion bases of
viral DNA sequence. Viral genomes mostly showed low variation, but 51 loci of ¡­100 bp showed
extremely high variation, so that up to 96% of the viral genomes
encoded unique amino acid sequences. Some hotspots of hypervariation
were in genes homologous to the bacteriophage BPP-1 viral tail-fiber gene, which is known to
be hypermutagenized by a unique reverse-transcriptase (RT)-based mechanism. Unexpectedly,
other hypervariable loci in our data were in previously undescribed gene types, including
genes encoding predicted Ig-superfamily proteins. Most of the hypervariable loci were linked
to genes encoding RTs of a single clade, which we find is the most abundant clade
among gut viruses but only a minor component of bacterial RT populations. Hypervariation was
targeted to 5¡Ç-AAY-3¡Ç asparagine codons, which allows maximal chemical diversification of
the encoded amino acids while avoiding formation of stop codons. These findings document
widespread targeted hypervariation in the human gut virome, identify previously undescribed
types of genes targeted for hypervariation, clarify association with RT gene clades, and
motivate studies of hypervariation in the full human microbiome.

<>

<1>Minton, N., Carter, G., Herbert, M., O'Keeffe, T., Purdy, D., Elmore, M., Ostrowski, A., Pennington, O., Davis, I.
<2>The development of Clostridium difficile genetic systems.
<3>Anaerobe
<4>10
<5>75-84
<6>2004
<7>Clostridum difficile is a major cause of healthcare-associated disease in the western world,
and is particularly prominent in the elderly. Its
incidence is rising concomitant with increasing longevity. More
effective countermeasures are required. However, the pathogenesis of C.
difficile infection is poorly understood. The lack of effective genetic
tools is a principal reason for this ignorance. For many years. the
only tools available for the transfer of genes into C. difficile have
been conjugative transposons, such as Tn916, delivered via filter
mating from Bacillus subtilis donors. They insert into a preferred site
within the genome. Therefore, they may not be employed for classical
mutagenesis studies, but can be employed to modulate gene function
through the delivery of antisense RNA. Attempts to develop
transformation procedures have so far met with little success. However,
in recent years the situation has been dramatically improved through
the demonstration of efficient conjugative transfer of both
replication-proficient and replication-deficient plasmids from
Escherichia coli donors. This efficient transfer can only be achieved
in certain strains through negation of the indigenous restriction
barrier, and is generally most effective when the plasmid employed is
based on the replicon of the C. difficile plasmid. pCD6.

<>

<1>Minton, N.P., Purdy, D.A., Elmore, M.J., O'Keeffe, K.M.T.
<2>New plasmid, designated pCD6 useful for transformation of Clostridium difficile and for expressing gene in Clostridium difficile plasmid pCD25EC-mediated chloramphenicol-acetyltransferase gene transfer and expression in Clostridium botulum or Clostridium.
<3>International Patent Office
<4>WO 200177319
<5>
<6>2001
<7>Plasmid pCD6 for the transformation of Clostridium difficile NCIMB-8052 or C. difficile 630,
Clostridium botulum or Clostridium beijerinckii is claimed. Also claimed are: expressing a DNA
sequence in C. difficile by introducing pCD6; identification of a C. difficile virulence
factor by culturing C. difficile in the absence and presence of a regulating factor that
promotes expression of virulence factors and by administering an antisense sequence to the
regulating factor; a DNA whose differentiation expression regulates expression of a virulence
factor; identification of vector that integrates into a Gram-positive bacterial genome; and
identification of a C. difficile methylase gene. In an example, nucleotide sequence analysis
revealed that the vector was composed of 6,829 bp. Derivative of pCD6, plasmid pCD25EC, was
constructed expressing chloramphenicol-acetyltransferase (EC-2.3.1.28) and transformed into
630 with erythromycin as selective marker. Other selective markers were also employed such as
Enterococcus faecalis or Staphylococcus aureus spectinomycin.

<>

<1>Miriagou, V., Papagiannitsis, C.C., Kotsakis, S.D., Loli, A., Tzelepi, E., Legakis, N.J., Tzouvelekis, L.S.
<2>Sequence of pNL194, a 79.3-Kilobase IncN Plasmid Carrying the blaVIM-1 Metallo-{beta}-Lactamase Gene in Klebsiella pneumoniae.
<3>Antimicrob. Agents Chemother.
<4>54
<5>4497-4502
<6>2010
<7>The nucleotide sequence of pNL194, a VIM-1-encoding plasmid, is described.
pNL194 (79,307-bp) comprised an IncN characteristic segment (38,940-bp)
and a mosaic structure (40,367-bp) including blaVIM-1, aacA7, aadA1,
dfrA1, dfrA12, aphA1, strA, strB, and sul1. Tn1000 or Tn5501 insertion
within fipA probably facilitated recruitment of additional mobile elements
carrying resistance genes.

<>

<1>Miroshnichenko, B.A., Naroditskii, B.S., Khilko, S.N., Platonova, N., Gruber, I.M., Tikhonenko, T.I.
<2>Purification of specific endonucleases EcoRI and BglII by affinity chromatography.
<3>Biokhimiia
<4>47
<5>686-694
<6>1982
<7>Highly active preparations of specific endonucleases EcoRI from E. coli cells and BglII from
Bacillus globigii cells were obtained by affinity chromatography.  The isolation and
purification procedures included ultrasonic disintegration of the cells, high-speed
centrifugation, and chromatography. Dextran blue-Sepharose, folate-Sepharose, and
phenyl-Sepharose were used as affinity sorbents.  The optimum conditions for sorption and
elution of the endonucleases without intermediate steps of dialysis and concentration were
selected.  The consecutive combining of the above-mentioned sorbents with various ligands made
it possible to achieve highly efficient purification.  The purified enzyme preparations do not
contain nonspecific nucleases or phosphatases, they are fairly concentrated, and can be used
for the specific hydrolysis of DNA.

<>

<1>Misaki, W.
<2>A recombinant lactobacillus strain expressing genes coding for restriction enzymes cleaving the HIV genomes for use as a live  microbicide strategy against heterosexual transmission of HIV.
<3>Afr. J. Biotechnol.
<4>6
<5>1750-1756
<6>2007
<7>Using genetically engineered endogenous lactobacillus strains colonizing the vagina mucosa to
express heterogenous proteins has of
late joined the novel strategies aimed at developing a microbicides
against HIV. Using the lactobacillus metabolic genome pathway, we found
that these bacteria do not naturally produce restriction enzymes, but
rather, have a number of putative alien genes of the type. In view of
the antiviral defence role of restriction modification systems (RMS),
we searched for enzymes that cleave HIV-1, 2 and other SIV genomes
using theoretical computational methods. With over 200 such enzymes
identified, we present herein a plasmid vector mediated strategy for
modifying lactobacillus strains to express RMS islands as an approach
to developing a live HIV microbicide. This model is transferable to
other viral infections that find their way into humans through mucosal
orifices.

<>

<1>Misaki, W.
<2>Why bacteria derived R-M nucleic enzymatic peptides are likely efficient therapeutic molecules for use in the design and development of novel HIV inhibitory strategies.
<3>Afr. J. Biotechnol.
<4>7
<5>1791-1796
<6>2008
<7>In the past, we have identified, described and isolated over 200 bacteria derived Restriction
Modification (R-M) nucleic enzymatic peptides as efficient therapeutic molecules for use in
the development of novel HIV inhibitory strategies. In the issuing months of our publications,
3 questions have been directed to our work; (1) HIV is an RNA virus, thus restriction peptides
are impotent as defense peptides. (2) HIV genome is encapsulated in nuclear capsid and viral
envelope, making access impossible. (3) Human genome contains several palindromes recognizable
by R-M peptides, making safety delineation critical. This paper serves to provide succinct
responses to these issues, and highlight critical strategies being employed in ensuring the
development of safe Microbides and therapeutic vaccines based on this approach.

<>

<1>Misaki, W., Wilson, B., Henry, K.
<2>Frequency and site mapping of HIV-1/SIVcpz, HIV2/SIVsmm and other SIV gene sequence cleavage by various bacteria restriction enzymes:  Precursors for a novel HIV inhibitory product.
<3>Afr. J. Biotechnol.
<4>6
<5>1225-1232
<6>2007
<7>Resistance, toxicity and virologic failure have underlined the need to develop new HIV
inhibitory products. Base on the natural bacteria
"restriction modification system" antiviral immune model, we set out to
analyze the effects of various restriction enzymes on the HIV genome. A
computer simulated model using Web cutter Version 2.0, and cytogenetic
analysis. 339 restriction enzymes from Promega database, 10
HIV-1/SIVcpz genes, 10 HIV-2/SIVsmm genes and 10 other SIV genes. Gene
sequences were fed into Web cutter 2.0 set to search enzymes with at
least 6 recognition base pairs ( palindromes). A background in vitro
cytogenetic control analysis using HIV-1/ SIVcpz GAG, POL and ENV genes
was done. Of the 339 enzymes used, 238 ( 70.2%) cleaved the HIV-1/
SIVcpz A1. BY. 97.97BL006  AF193275 genome with 9037 bp compared to 225
( 66.4%) and 219 ( 64.6%) for the HIV-2/SIVsmm genome ( 9713 bp) and
other SIV B. FR. 83. HXB2  LAI  IIIB  BRU  K03455 genome ( 9719 bp),
respectively. Individual genes had differing but potent susceptibility
to the enzymes, with a 98.9% Web cutter PPV ( 95% CI, 97.2%99.6%) for
in vitro cytogenetics. The natural bacteria RMS antiviral immune model
offers precursors for developing novel HIV and other viral therapeutic
molecules.

<>

<1>Mise, K., Miyahara, M.
<2>Restriction endonucleases: Their characteristics and distribution in pathogenic bacteria.
<3>Eisei Shikenjo Hokoku
<4>111
<5>1-12
<6>1993
<7>Restriction endonucleases have been widely employed in almost all fields of genetic
engineering including DNA mapping, cloning, sequencing, hybridization, amplification and
diagnosis. The general characteristics of restriction endonucleases and their reactions are
reviewed in this paper, together with their distribution in pathogenic bacteria. Many
restriction endonucleases with novel specificity have been found in these bacteria in our
laboratory. Some of them are commercially available and have been employed for the molecular
biologist. Rapid method for detection of restriction endonucleases in pathogenic bacteria is
also described.

<>

<1>Mise, K., Miyahara, M., Maruyama, T., Kudoh, Y., Ohashi, M.
<2>Usefulness in the epidemiology of food poisoning cases of detection of specific restriction endonucleases in some serotypes of Salmonella and Yersinia.
<3>Microbial Toxins in Foods and Feeds: Cell. Mol. Modes Action, Plenum Press, Pohland, A.E., Dowell, V.R., Richard, J.L., New York
<4>0
<5>127-129
<6>1990
<7>Restriction endonucleases have been employed as an extremely important tool in
recombinant DNA technology.  To date, the occurrence of more than 100
restriction endonucleases with different specificities has been reported.
Restriction endonucleases have been screened in this laboratory for pathogenic
bacteria belonging to the Enterobacteriaceae in the hope that: (i) the
detection of a specific restriction endonuclease is found at a high frequency
in the species; and (ii) new restriction endonucleases with novel specificities
might be found in pathogenic bacteria, since screening for restriction
endonucleases has rarely been carried out.  Here, we report that four
clinically important food-poisoning bacteria produce specific restriction
endonucleases at high frequencies.  Some of these are expected to be useful for
recombinant DNA technology after cloning of their gene into Escherichia coli
K-12.

<>

<1>Mise, K., Nakajima, K.
<2>Purification of a new restriction endonuclease, StyI, from Escherichia coli carrying the hsd+ miniplasmid.
<3>Gene
<4>33
<5>357-361
<6>1985
<7>A new restriction endonuclease, StyI, free of contaminating nuclease
activities, has been isolated from Escherichia coli carrying the hsd+
miniplasmid of Salmonella typhi origin.  In the presence of 10 mM Mg2+, it
recognizes and cleaves a hexanucleotide sequence of 5'-C^C(A/T)(A/T)GG.  The
advantages of the StyI endonuclease include its stability, high yield (more
than 2 x 103 units/g of wet cells), easy handling of producer cells, and the
ability to recognize new sequences, CCAAGG and CCTTGG.

<>

<1>Mise, K., Nakajima, K.
<2>Isolation of restriction enzyme EcoVIII, an isoschizomer of HindIII, produced by Escherichia coli E1585-68.
<3>Gene
<4>30
<5>79-85
<6>1984
<7>A restriction endonuclease designated EcoVIII, an isoschizomer of HindIII, was
isolated from Escherichia coli E1585-68 and purified by dextran-polyethylene
glycol (DPG) phase partitiion, ammonium sulfate precipitation, phospho- and
DEAE-cellulose chromatography, and hydroxylapatite chromatography.  The
purified EcoVIII was stable during the purification procedure and its high
specific activity required 10 mM Mg2+.  Unlike HindIII, EcoVIII exhibited a
high specific activity even at low pH (pH 6.3) and showed the highest activity
at 48C.  Transformation of purified plasmid DNA from E. coli E1585-68 into K-12
indicated that the EcoVIII gene was carried on a multicopy 4.4-kb miniplasmid.
EcoVIII seems to be preferable to HindIII for its production and use because of
easier handling of producer cells and a wider range of activity.

<>

<1>Mise, K., Nakajima, K.
<2>Restriction endonuclease EcoO109 from Escherichia coli H709c with heptanucleotide recognition site 5'-PuG^GNCCPy.
<3>Gene
<4>36
<5>363-367
<6>1985
<7>A new restriction endonuclease, EcoO109, has been isolated from Escherichia
coli H709c by polyethylenemine (PEI) precipitation, DEAE-cellulose
chromatography and heparin agarose chromatography.  The yield was high, more
than 3000 units/g of wet cells.  The EcoO109 endonuclease recognizes and
cleaves a nucleotide sequence of 5'-PuG^GN C CPy-3' 3'-PyC CN'G^GPu-5'in the
presence of 10 mM Mg2+.  The enzyme will be useful for structural analysis and
molecular cloning of DNA because of the stability, high yield and easy handling
of the producer strain.

<>

<1>Mise, K., Nakajima, K., Terakado, N., Ishidate, M.
<2>Production of restriction endonucleases using multicopy Hsd plasmids occurring naturally in pathogenic Escherichia coli and Shigella boydii.
<3>Gene
<4>44
<5>165-169
<6>1986
<7>A convenient procedure has been devised for detection of restriction
endonucleases in the Escherichia coli-Shigella group.  With this procedure, two
restriction endonucleases, designated Sbo13 and EcoT22, were found and later
were identified as isoschizomers of NruI and AvaIII, respectively.  These
endonucleases were shown to be produced from small multicopy plasmids.  They
were isolated from nonpathogenic E. coli into which the plasmids had been
introduced by transformation, and purified from contaminating nuclease
activity.  The yield was high, 1000 units/g of wet cells for Sbo13 and 500
units/g for EcoT22.  Sbo13 and EcoT22 should be preferable to NruI and AvaIII
because of the high yield and ease in handling the producer cells.

<>

<1>Mishra, A.K., Edouard, S., Dangui, N.P., Lagier, J.C., Caputo, A., Blanc-Tailleur, C., Ravaux, I., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Nosocomiicoccus massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>205-219
<6>2013
<7>Nosocomiicoccus massiliensis strain NP2(T) sp. nov. is the type strain of a new species within
the genus Nosocomiicoccus. This strain, whose genome is described
here, was isolated from the fecal flora of an AIDS-infected patient living in
Marseille, France. N. massiliensis is a Gram-positive aerobic coccus. Here we
describe the features of this organism, together with the complete genome
sequence and annotation. The 1,645,244 bp long genome (one chromosome but no
plasmid) contains 1,738 protein-coding and 45 RNA genes, including 3 rRNA genes.

<>

<1>Mishra, A.K., Gimenez, G., Lagier, J.C., Robert, C., Raoult, D., Fournier, P.E.
<2>Genome sequence and description of Alistipes senegalensis sp. nov.
<3>Standards in Genomic Sciences
<4>6
<5>1-16
<6>2012
<7>Alistipes senegalensis strain JC50(T) is the type strain of A. senegalensis sp. nov., a new
species within the Alistipes genus. This strain, whose genome is
described here, was isolated from the fecal flora of an asymptomatic patient. A.
senegalensis is an anaerobic Gram-negative rod-shaped bacterium. Here we describe
the features of this organism, together with the complete genome sequence and
annotation. The 4,017,609 bp long genome (1 chromosome, but no plasmid) contains
3,113 protein-coding and 50 RNA genes, including 5 rRNA genes.

<>

<1>Mishra, A.K., Hugon, P., Lagier, J.C., Nguyen, T.T., Couderc, C., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Enorma massiliensis gen. nov., sp. nov., a new member of the Family Coriobacteriaceae.
<3>Standards in Genomic Sciences
<4>8
<5>290-305
<6>2013
<7>Enorma massiliensis strain phI(T) is the type strain of E. massiliensis gen. nov., sp. nov.,
the type species of a new genus within the family
Coriobacteriaceae, Enorma gen. nov. This strain, whose genome is described here,
was isolated from the fecal flora of a 26-year-old woman suffering from morbid
obesity. E. massiliensis strain phI(T) is a Gram-positive, obligately anaerobic
bacillus. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 2,280,571 bp long genome (1
chromosome but no plasmid) exhibits a G+C content of 62.0% and contains 1,901
protein-coding and 51 RNA genes, including 3 rRNA genes.

<>

<1>Mishra, A.K., Hugon, P., Lagier, J.C., Nguyen, T.T., Robert, C., Couderc, C., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Peptoniphilus obesi sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>357-369
<6>2013
<7>Peptoniphilus obesi strain ph1(T) sp. nov., is the type strain of P. obesi sp. nov., a new
species within the genus Peptoniphilus. This strain, whose genome is
described here, was isolated from the fecal flora of a 26-year-old woman
suffering from morbid obesity. P. obesi strain ph1(T) is a Gram-positive,
obligate anaerobic coccus. Here we describe the features of this organism,
together with the complete genome sequence and annotation. The 1,774,150 bp long
genome (1 chromosome but no plasmid) contains 1,689 protein-coding and 29 RNA
genes, including 5 rRNA genes.

<>

<1>Mishra, A.K., Hugon, P., Robert, C., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Peptoniphilus grossensis sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>320-330
<6>2012
<7>Peptoniphilus grossensis strain ph5(T) sp. nov., is the type strain of Peptoniphilus
grossensis sp. nov., a new species within the Peptoniphilus genus.
This strain, whose genome is described here, was isolated from the fecal flora of
a 26-year-old woman suffering from morbid obesity. P. grossensis strain ph5 is a
Gram-positive obligate anaerobic coccus. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
2,101,866-bp long genome (1 chromosome but no plasmid) exhibits a G+C content of
33.9% and contains 2,041 protein-coding and 29 RNA genes, including 3 rRNA genes.

<>

<1>Mishra, A.K., Lagier, J.C., Nguyen, T.T., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Peptoniphilus senegalensis sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>370-381
<6>2013
<7>Peptoniphilus senegalensis strain JC140(T) sp. nov., is the type strain of P. senegalensis sp.
nov., a new species within the genus Peptoniphilus. This strain,
whose genome is described here, was isolated from the fecal flora of a healthy
patient. P. senegalensis strain JC140(T) is an obligate Gram-positive anaerobic
coccus. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 1,840,641 bp long genome (1
chromosome but no plasmid) exhibits a G+C content of 32.2% and contains 1,744
protein-coding and 23 RNA genes, including 3 rRNA genes.

<>

<1>Mishra, A.K., Lagier, J.C., Pfleiderer, A., Nguyen, T.T., Caputo, A., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Holdemania massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>395-409
<6>2013
<7>Holdemania massiliensis strain AP2(T) sp. nov. is the type strain of H. massiliensis sp. nov.,
a new species within the genus Holdemania. This strain,
whose genome is described here, was isolated from the fecal flora of a
21-year-old French Caucasian female suffering from severe restrictive anorexia
nervosa. H. massiliensis is a Gram-positive, anaerobic bacillus. Here we describe
the features of this organism, together with the complete genome sequence and
annotation. The 3,795,625 bp-long genome (one chromosome but no plasmid) contains
3,461 protein-coding and 49 RNA genes, including 3 rRNA genes.

<>

<1>Mishra, A.K., Lagier, J.C., Rivet, R., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Paenibacillus senegalensis sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>70-81
<6>2012
<7>strain JC66, is the type strain of sp. nov., a new species within the genus . This strain,
whose genome is described here, was isolated from the fecal flora of
a healthy patient. strain JC66 is a facultative Gram-negative anaerobic
rod-shaped bacterium. Here we describe the features of this organism, together
with the complete genome sequence and annotation. The 5,581,254 bp long genome (1
chromosome but no plasmid) exhibits a G+C content of 48.2% and contains 5,008
protein-coding and 51 RNA genes, including 9 rRNA genes.

<>

<1>Mishra, A.K., Lagier, J.C., Robert, C., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Peptoniphilus timonensis sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>1-11
<6>2012
<7>strain JC401 sp. nov. is the type strain of sp. nov., a new species within the genus. This
strain, whose genome is described here, was isolated from the fecal
flora of a healthy patient. is an obligate Gram-positive anaerobic coccus. Here
we describe the features of this organism, together with the complete genome
sequence and annotation. The 1,758,598 bp long genome (1 chromosome, no plasmid)
contains 1,922 protein-coding and 22 RNA genes, including 5 rRNA genes.

<>

<1>Mishra, A.K., Lagier, J.C., Robert, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Clostridium senegalense sp. nov.
<3>Standards in Genomic Sciences
<4>6
<5>386-395
<6>2012
<7>Clostridium senegalense strain JC122(T), is the type strain of Clostridium senegalense sp.
nov., a new species within the genus Clostridium. This strain,
whose genome is described here, was isolated from the fecal flora of a healthy
patient. C. senegalense strain JC122(T) is an obligate anaerobic Gram-positive
rod-shaped bacterium. Here we describe the features of this organism, together
with the complete genome sequence and annotation. The 3,893,008 bp long genome (1
chromosome but no plasmid) exhibits a G+C content of 26.8% and contains 3,704
protein-coding and 57 RNA genes, including 6 rRNA genes.

<>

<1>Mishra, A.K., Lagier, J.C., Robert, C., Raoult, D., Fournier, P.E.
<2>Genome sequence and description of Timonella senegalensis gen. nov., sp. nov., a  new member of the suborder Micrococcinae.
<3>Standards in Genomic Sciences
<4>8
<5>318-335
<6>2013
<7>Timonella senegalensis strain JC301(T) gen. nov., sp. nov. is the type strain of  T.
senegalensis gen. nov., sp. nov., a new species within the newly proposed
genus Timonella. This bacterial strain was isolated from the fecal flora of a
healthy Senegalese patient. In this report, we detail the features of this
organism, together with the complete genome sequence and annotation. Timonella
senegalensis strain JC301(T) exhibits the highest 16S rRNA similarity (95%) with
Sanguibacter marinus, the closest validly published bacterial species. The genome
of T. senegalensis strain JC301(T) is 3,010,102-bp long, with one chromosome and
no plasmid. The genome contains 2,721 protein-coding genes and 72 RNA genes,
including 5 rRNA genes. The genomic annotation revealed that T. senegalensis
strain JC301(T) possesses the complete complement of enzymes necessary for the de
novo biosynthesis of amino acids and vitamins (except for riboflavin and biotin),
as well as the enzymes involved in the metabolism of various carbon sources,
chaperone genes, and genes involved in the regulation of polyphosphate and
glycogen levels.

<>

<1>Mishra, A.K., Pfleiderer, A., Lagier, J.C., Robert, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Bacillus massilioanorexius sp. nov.
<3>Standards in Genomic Sciences
<4>8
<5>465-479
<6>2013
<7>Bacillus massilioanorexius strain AP8(T) sp. nov. is the type strain of B. massilioanorexius
sp. nov., a new species within the genus Bacillus. This strain,
whose genome is described here, was isolated from the fecal flora of a
21-year-old Caucasian French female suffering from a severe form of anorexia
nervosa since the age of 12 years. B. massilioanorexius is a Gram-positive
aerobic bacillus. Here we describe the features of this organism, together with
the complete genome sequence and annotation. The 4,616,135 bp long genome (one
chromosome but no plasmid) contains 4,432 protein-coding and 87 RNA genes,
including 8 rRNA genes.

<>

<1>Mishra, N.C.
<2>Restriction Endonucleases.
<3>Nucleases. Molecular Biology and Applications., John Wiley & Sons, Inc., none, Hoboken, NJ
<4>
<5>62-80
<6>2002
<7>Luria and Human first described the phenomenon of host-controlled specificity in T-even
phages.  This was immediately confirmed by Bertani and Weigle in lambda and P2 phages.  In
these studies it was shown that a particular bacteriophage possessed different efficiencies of
infection on several closely related strains of bacteria.  However, the progeny of
bacteriophage which initially plated with a low efficiency was later found to plate
efficiently after one generation of plating on the same bacterium.  This phenomenon of
host-controlled specificity was first discovered in T-even phages by Luria and Human  and in
lambda and P2 phages by Bertani and Weigle.  The epigenetic nature of such host range
specificity was indicated by the fact that the newly acquired high efficiency of plating on a
particular bacterial host was lost by the progeny of the same bacteriophage when plated on
different hosts.  The first host modification unique to T-even phages involved the
glycosylation of hydroxymethyl cytosine residue.  However, the modification introduced in
lambda phages was found to be of universal occurrence.  The molecular basis of such host range
specificity depended on the restriction or modification of certain DNA sequences by
restriction and modification enzymes (Arber and Dussoix, 1962; Arbor and Linn, 1969).  Foreign
DNA molecules (lacking appropriate modification) were cleaved by the host restriction
endonuclease upon entry into the bacterial cell.  During this process some of the phage DNA
may escape restriction by host endonucleases and instead get modified by methylation of
adenine or cytosine moieties in DNA and thus acquire the ability to infect the same host
efficiently in subsequent rounds of infection.  In these series of events, bacterial DNAs
remain protected because of previous modification of the DNA sequences by methylase.  This
molecular scenario for the host range specificity by Arber and Dussoix was confirmed by the
discovery of bacterial restriction-modification enzymes.  The restriction endonucleases widely
differ in terms of subunit composition, cofactor requirements, and interaction with DNA
substrate in addition to their specificity for nucleotide sequence as sites of recognition and
of cleavage or modification.  They are classified under three categories.  These are: type I
restriction endonuclease, type II restriction endonuclease, and type III restriction
endonuclease.  Among these three types of restriction endonuclease, type I and type III
enzymes can be referred to as ATP-dependent restriction endonucleases and the type II enzymes
can be referred to as ATP-independent restriction endonucleases.  The physiological roles of
the ATP-dependent restriction endonucleases in biological restriction and modification have
been genetically identified.  The physiological role of the ATP-independent endonucleases
remains to be elucidated.  However, the discovery of this group of restriction endonucleases
has revolutionized molecular biology by facilitating the physical mapping and nucleotide
sequencing of DNA segments and their amplification by molecular cloning.  The properties of
the three types of restriction enzymes are compared in Table 4.1.

<>

<1>Misra, V.K., Hecht, J.L., Sharp, K.A., Friedman, R.A., Honig, B.
<2>Salt effects on protein-DNA interactions.  The lambdacI repressor and EcoRI endonuclease.
<3>J. Mol. Biol.
<4>238
<5>264-280
<6>1994
<7>In this paper, finite-difference solutions to the nonlinear Poisson-Boltzmann (NLPB) equation
are used to calculate the salt dependent contribution to the electrostatic DNA binding free
energy for both the lambdacI repressor and the EcoRI endonuclease. For the protein-DNA systems
studied, the NLPB method describes nonspecific univalent salt dependent effects on the binding
free energy which are in excellent agreement with experimental results. In these systems, the
contribution of the ion atmosphere to the binding free energy substantially destabilizes the
protein-DNA complexes. The magnitude of this effect involves a macromolecular structure
dependent redistribution of both cations and anions around the protein and the DNA which is
dominated by long range electrostatic interactions. We find that the free energy associated
with global ion redistribution upon binding is more important than changes associated with
local protein-DNA interactions (ion-pairs) in determining salt effects. The NLPB model reveals
how long range salt effects can play a significant role on the relative stability of
protein-DNA complexes with different structures.

<>

<1>Mitra, A., Higgins, D.W.
<2>The Chlorella virus adenine methyltransferase gene is a strong promoter in plants.
<3>Plant Mol. Biol.
<4>26
<5>85-93
<6>1994
<7>An upstream region isolated from a eukaryotic algal virus adenine methyltransferase gene was
tested for promoter function in plants. Fusion of this region to the chloramphenicol
acetyltransferase reporter gene resulted in significantly higher expression than fusion with
the cauliflower mosaic virus 35S promoter. Strong levels of expression were also found in
electroporated monocot plant cells. The promoter activity in transgenic tobacco plants showed
tissue-specific expression. Leaves had the highest expression followed by stems and flowers.
The promoter activity was not detected in root tissue. Environmental cues, such as light, and
the phytohormones auxin and cytokinines had no effect on the promoter's expression. This
promoter might be utilized to achieve high levels of expression of introduced genes in higher
plants.

<>

<1>Mitra, A., Que, Q.
<2>Ectopic expression of a viral adenine methyltransferase gene in tobacco.
<3>Biochim. Biophys. Acta
<4>1219
<5>244-249
<6>1994
<7>Plant genomes contain both methylated adenine and cytosine residues although the roles of
these methylations are not well understood. A Chlorella virus adenine methyltransferase gene
under the control of cauliflower mosaic virus 35S promoter in a binary plant transformation
vector was expressed both in transgenic tobacco plants and transformed tobacco calli. The
transgenic plants as well as transformed calli produced functional adenine methyltransferase
enzyme, but the level of expression was higher in tobacco calli. A transgenic tobacco cell
line that expressed the methyltransferase enzyme and carried an Arabidopsis cab3 gene
containing a single target site for the adenine methyltransferase enzyme showed that the
adenine residue was not methylated. HPLC analysis of genomic DNA from transgenic calli also
showed no detectable levels of methylated adenine residues.

<>

<1>Mitra, A., van Etten, J.
<2>Promoters from Chlorella virus genes providing for expression of genes in prokaryotic and eukaryotic hosts.
<3>US Patent Office
<4>US 6252140
<5>
<6>2001
<7>An expression cassette (I) is claimed. (I) contains a promoter or its mutant substantially
corresponding to a promoter from a Chlorella virus
DNA-methyltransferase gene linked to a first DNA sequence encoding a
gene that is different from the Chlorella virus gene. Also claimed are:
a plasmid (II) containing (I); a transformed plant cell (II) that
contains an expression cassette containing a promoter or its mutant
corresponding to a promoter from a Chlorella DNA-methyltransferase; a
transgenic plant (IV) containing plant cells having an expression
cassette stably integrated into the genome; a seed (V) comprising a
transformed plant cell; a transformed bacterial (VI); an isolated
promoter; and a method of producing a recombinant gene product in a
cell. The promoters are useful for highly level gene expression in
prokaryotic or eukaryotic cells. (21pp)

<>

<1>Mitrovic, J., Siewert, C., Duduk, B., Hecht, J., Moelling, K., Broecker, F., Beyerlein, P., Buettner, C., Bertaccini, A., Kube, M.
<2>Generation and Analysis of Draft Sequences of 'Stolbur' Phytoplasma from Multiple Displacement Amplification Templates.
<3>J. Mol. Microbiol. Biotechnol.
<4>24
<5>1-11
<6>2014
<7>Phytoplasma-associated diseases are reported for more than 1,000 plant species worldwide.
Only a few genome sequences are available in contrast to the economical importance of these
bacterial pathogens.  A new strategy was used to retrieve phytoplasma strain-specifric genome
data.  Multiple displacement amplification was performed on DNA obtained from <3 g of plant
tissue from tobacco and parsley samples infected with 'stolbur' strains.  Random hexamers
and Phi29 polymerase were evaluated with and without supplementation by group-assigned
oligonucleotides providing templates for Illumina's sequencing approach.  Metagenomic drafts
derived from individual and pooled strain-specific de novo assembles were analyzed.
Supplementation of the Phi29 reaction with the group-assigned oligonucleotides resulted in an
about 2-fold enrichment of the percentage of phytoplasma-assigned reads and thereby improved
assembly results.  The obtained genomic drafts represent the largest datasets available from
'stolbur' phytoplasmas.  Sequences of the two strains (558 kb, 448 proeins and 516 kb, 346
proteins, respectively) were annotated allowing the identification of prominent membrane
proteins and reconstruction of core pathways.  Analysis of a putative truncated sucrose
phosphorylase provides hints on sugar degradation.  Furthermore, it is shown that drafts
obtained from repetitive-rich genomes allow only limited analysis on multicopy regions and
genome completeness.

<>

<1>Miura, T., Kusada, H., Kamagata, Y., Hanada, S., Kimura, N.
<2>Genome Sequence of the Multiple-beta-Lactam-Antibiotic-Resistant Bacterium Acidovorax sp. Strain MR-S7.
<3>Genome Announcements
<4>1
<5>e00412-13
<6>2013
<7>Acidovorax sp. strain MR-S7 was isolated from activated sludge in a treatment system for
wastewater containing beta-lactam antibiotic pollutants. Strain MR-S7
demonstrates multidrug resistance for various types of beta-lactam antibiotics at
high levels of MIC. The draft genome sequence clarified that strain MR-S7 harbors
unique beta-lactamase genes.

<>

<1>Miura, T., Tsuchikane, K., Numata, M., Hashimoto, M., Hosoyama, A., Ohji, S., Yamazoe, A., Fujita, N.
<2>Complete Genome Sequence of an Alkane Degrader, Alcanivorax sp. Strain NBRC 101098.
<3>Genome Announcements
<4>2
<5>e00766-14
<6>2014
<7>Alcanivorax sp. strain NBRC 101098 was isolated from seawater in Japan. Strain NBRC 101098 is
able to degrade various types of n-alkanes. Here, we report the
complete genome of strain NBRC 101098.

<>

<1>Miyada, C.G., Born, T.L.
<2>Support-bound nucleotide probe for Neisseria gonorrhoeae.
<3>US Patent Office
<4>US 5525717
<5>
<6>1996
<7>A nucleotide sequence characteristic of Neisseria gonorrhoeae is disclosed.  The sequence can
be the basis for hybridization type, nucleic acid-based, rapid, in vitro diagnostic assays.
The unique nature of the sequence makes it possible to clearly discriminate N. gonorrhoeae
from other Neisseria species thus eliminating or substantially reducing the number of false
positive readings.  A 350 base pair N. gonorrhoeae DNA restriction fragment was cloned after
subtractive hybridization to Neisseria meningitidis DNA.  In further cloning experiments the
sequences adjacent to the original 350 base pair fragment were determined.  A portion of this
sequence was shown to detect 105 of 106 N. gonorrhoeae strains and no other Neisseria species.
In addition to use as detection probes, all or portions of the nucleotide sequence can be used
as a ligand for the sandwich capture of N. gonorrhoeae sequences and as primers for in vitro
amplification of N. gonorrhoeae sequences.  The polypeptides encoded by the presently
disclosed sequence, including antibodies thereto, are also disclosed as are their uses.

<>

<1>Miyada, C.G., Born, T.L.
<2>A DNA sequence for the discrimination of Neisseria gonorrhoeae from other Neisseria species.
<3>Mol. Cell. Probes
<4>5
<5>327-335
<6>1991
<7>A 350 base pair Neisseria gonorrhoeae DNA restriction fragment was cloned after subtractive
hybridization to Neisseria meningitidis DNA.  This restriction fragment hybridized to 105 out
of 106 N. gonorrhoeae strains tested.  While three N. meningitidis strains did not hybridize
to this probe, Neisseria mucosa DNA exhibited cross-hybridization.  This particular clone was
used to screen a N. gonorrhoeae genomic DNA library.  A positive 2-4 kilobase pair clone was
shown by DNA sequencing to contain two long open reading frames.  One open reading frame did
not hybridize to N. mucosa and other Neisseria species, while it retained specificity for the
original 105 N. gonorrhoeae strains.  This open reading frame also showed significant homology
to cytosine DNA methyltransferases.

<>

<1>Miyagi, T., Javorsky, P., Pristas, P., Karita, S., Sakka, K., Ohmiya, K.
<2>Partial purification and characterization of RalF40I, a class II restriction endonuclease from Ruminococcus albus F-40, which recognizes and cleaves 5'-/GATC-3'.
<3>FEMS Microbiol. Lett.
<4>164
<5>215-218
<6>1998
<7>Restriction endonuclease RalF40I was purified from cell-free extracts of the rumen
cellulolytic bacterium Ruminococcus albus F-40 by heparin-Sepharose chromatography.  The
preparation was active only on DNA substrates that were not Dam-methylated.  RalF40I
recognizes the 4-bp palindrome, 5'-/GATC-3', and cleaves DNA at the 5' side of G in the
sequence, producing 5' tetranucleotide protruding ends.  RalF40I is a class II restriction
endonuclease and an isoschizomer of MboI and DpnII.

<>

<1>Miyahara, M., Fujiwara, R., Mise, K., Shimada, T., Matsushita, S., Kudoh, Y., Ishiwata, N., Tanimura, A.
<2>Characterization of restriction endonucleases from Vibrio parahaemolyticus.
<3>J. Food. Hyg. Sci. Japan
<4>35
<5>605-609
<6>1994
<7>Forty-six restriction endonuclease (ENase)-positive strains were found among 270 strains of
Vibrio parahaemolyticus tested. Thirty-six ENases were purified and 7 different specificities
were identified. All are isoschizomers of already-known ENases, i.e., isoschizomers of AvaII,
AsuI, PmaCI, PstI, Eco31I, EarI, and SapI. It is noteworthy that the majority of V.
parahaemolyticus ENases recognize non-palindromic or interrupted palindromic sequences.
Specific ENases with the same specificity were found at a high frequency in some serotypes of
V. parahaemolyticus.

<>

<1>Miyahara, M., Ishiwata, N., Yoshida, Y.
<2>StyD4I restriction-modification system of Salmonella typhi D4: Cloning and sequence analysis.
<3>Biol. Pharm. Bull.
<4>20
<5>201-203
<6>1997
<7>A plasmid (5.4 kbp) from Salmonella Typhi D4 has been identified as encoding a restriction and
modification (R-M) system.  DNA fragments (2537 bp) that carried the genes for the restriction
endonuclease and methyltransferase encoded on the plasmid were sequenced.  Two divergently
arranged open reading frames of 957 bp for the restriction endonuclease consisting of 318 aa
(amino acids) and 1140 bp for the DNA methyltransferase consisting of 379 aa were identified.
These sequences were similar to the sequences of the SsoII R-M system, including the
interspace between the two genes.

<>

<1>Miyahara, M., Kimizuka, F., Kita, A., Matsushita, S., Kudo, Y., Shimada, T., Mise, K.
<2>Isolation and characterization of restriction endonuclease in Plesiomonas shigelloides and Aeromonas species.
<3>Biol. Pharm. Bull.
<4>19
<5>1506-1507
<6>1996
<7>Five restriction endonucleases (ENases) and one ENase were found in a screen of 196 strains of
Plesiomonas shigelloides and 147 strains of Aeromonas species.  Plesiomonas and Aeromonas
species are classified as Vibrionaceae, identified as food-poisoning bacteria, are closely
genetically related to each other, and their ENases producing abilities have not been
reported.  ENases were detected at relatively low frequencies in these species as compared to
those in other species, such as Salmonella species and Vibrio parahaemolyticus.  All ENases
were shown to be isoschizomers of already known ENases.  One of the Plesiomonas ENases,
designated PshBI, recognizing the sequence 5'-AT/TAAT-3' should be useful, since PshBI ENase
is produced at a high yield of 7000 units/g of wet cells.  The specificities of other ENases
are also described in this paper.

<>

<1>Miyahara, M., Kudoh, Y., Mise, K.
<2>Widespread occurrence of specific restriction endonucleases in Salmonella infantis, Salmonella thompson, and Salmonella blockley isolated from humans in Japan.
<3>Appl. Environ. Microbiol.
<4>56
<5>2248-2250
<6>1990
<7>Specific restriction endonucleases were detected in three serotypes of
Salmonella spp. isolated from humans in Japan from 1970 to 1987: an
isoschizomer of AvaII endonuclease at a frequency of 0.91 in Salmonella
infantis, an isoschizomer of KpnI at a frequency of 0.34 in Salmonella
thompson, and an isoschziomer of StyI at a frequency of 0.30 in Salmonella
blockley.  Of interest is that restriction endonuclease-producing S. thompson
was detected at high frequencies in the 1970s but at low frequencies in the
1980s.

<>

<1>Miyahara, M., Maruyama, T., Wake, A., Mise, K.
<2>Widespread occurrence of the restriction endonuclease YenI, an isoschizomer of PstI, in Yersinia enterocolitica serotype 08.
<3>Appl. Environ. Microbiol.
<4>54
<5>577-580
<6>1988
<7>The cold-active restriction endonuclease YenI, an isoschizomer of PstI, was
found in 12 of 14 Yersinia enterocolitica serotype 08 strains of different
origins, but not in other serotypes of Y. enterocolitica, Yersinia
pseudotuberculosis, or Yersinia pestis.  In spite of the limited number of
strains tested, the result suggests that the detection of YenI endonuclease or
the gene might result in more rapid determination of the prominently pathogenic
serotype of Y. enterocolitica.

<>

<1>Miyahara, M., Mise, K.
<2>Rapid method for detection of restriction endonuclease-producing strains in enteropathogenic bacteria.
<3>Anal. Chim. Acta
<4>213
<5>273-277
<6>1988
<7>An improved rapid method is described for the detection of restriction
endonucleases in enteropathogenic bacteria including Salmonella and Escherichia
coli.  With the improved method, at least six restriction endonucleases with
different specificity were found in 415 strains tested.  Five of them were
shown to be isoschizomers of known restriction endonucleases, while one seemed
to be novel.  Among the isoschizomers, SthI endonuclease (isoschizomer of KpnI)
in Salmonella thompson appears to be useful; unlike KpnI, SthI generates DNA
fragments with a 5'-protruding end.

<>

<1>Miyahara, M., Mise, K.
<2>Isolation and characterization of the StyD4I restriction endonuclease a neoschizomer of ScrFI, from Escherichia coli K-12 carrying a small multicopy Hsd Plasmid of Salmonella typhi origin.
<3>Gene
<4>129
<5>83-86
<6>1993
<7>A restriction endonuclease designated StyD4I, a neoschizomer of ScrFI, has been isolated from
Escherichia coli K-12 carrying a small multicopy host specificity for DNA (Hsd) plasmid of
Salmonella typhi D4 origin. In the presence of 10 mM Mg2+, StyD4I cleaves the sequence
5'-/CCNGG-3' and generates a 5-nucleotide cohesive end. StyD4I should be useful for
recombinant DNA technology, because of the stability and ease in handling the producer cells.

<>

<1>Miyahara, M., Mise, K., Kimizuka, F., Matsumoto, H., Terawaki, Y.
<2>Purification of restriction endonuclease EcoO128I produced by an enteropathogenic Escherichia coli O128Ly3.
<3>Gene
<4>113
<5>135-136
<6>1992
<7>Restriction endonuclease EcoO128I, an isoschizomer of BstEII, was purified from a rough mutant
of Escherichia coli O128Ly3. EcoO128I should be more convenient for recombinant DNA
applications than BstEII, because of its improved cleavage activity at 37C.

<>

<1>Miyahara, M., Nakajima, K., Kawanishi, T., Mise, K.
<2>SshAI restriction endonuclease from Salmonella shikmonah.
<3>FEMS Microbiol. Lett.
<4>66
<5>245-248
<6>1990
<7>A new type II restriction endonuclease, SshAI, was purified from Salmonella shikmonah TK139 of
kangaroo origin.  The recognition and cleavage specificity of SshAI was determined to be
5'-CC/TNAGG-3', identical to that of SauI from Streptomyces aureofaciens and Bsu36I from
Bacillus subtilis.  Based on closely related and in part overlapping recognition specificities
of SshAI and other restriction endonucleases, a close evolutionary relationship is proposed
for all known Salmonella restriction endonucleases.

<>

<1>Miyahara, M., Nakajima, K., Shimada, T., Mise, K.
<2>Restriction endonuclease PshAI from Plesiomonas shigelloides with the novel recognition site 5'-GACNN/NNGTC.
<3>Gene
<4>87
<5>119-122
<6>1990
<7>A new restriction endonuclease (ENase), PshAI, has been isolated from
Plesiomonas shigelloides 319-73, an organism that causes food poisoning in
humans.  The enzyme was stable and produced a yield of 410 units/g of cells.
In the presence of 10 mM MgCl2, PshAI recognizes and cleaves the nucleotide
sequence 5'-GACNN/NNGTC, producing blunt ends.  PshAI will be useful for
structural analysis and molecular cloning of DNA, because no ENases recognizing
the sequence GACNNNNGTC have been previously described.

<>

<1>Miyahara, M., Nakamura, A., Mise, K.
<2>Characterization of two restriction endonucleases, SenPT14bI and SenPT16I, in standard phage-type strains of Salmonella enteritidis.
<3>Biol. Pharm. Bull.
<4>20
<5>1212-1214
<6>1997
<7>Two restriction endonucleases were found by screening 38 standard phage strains of Salmonella
enteritidis.  An isoschizomer of SacII Enase that recognizes the sequence 5'-CCGC/GG-3' was
identified in S. enteritidis PT14b, and an isoschizomer of XmaIII ENase (5'-C/GGCCG-3') was
found in S. enteritidis PT16.  It is of special interest that the recognition specifities of
all known ENases in Salmonella, including those of the S. enteritidis ENases, are very similar
to each other.

<>

<1>Miyahara, M., Shimada, T., Kotani, H., Mise, K.
<2>Isolation and characterization of new restriction endonucleases from Vibrio parahaemolyticus: VpaK32I enzyme with the class-IIS heptanucleotide specificity, GCTCTTCN1/N4.
<3>Gene
<4>117
<5>103-106
<6>1992
<7>Six restriction endonucleases (ENases), classified into four different specificities, were
found in a screen among 68 reference strains of Vibrio parahaemolyticus of human origin. Five
of these ENases are isoschizomers of well-know ENases, while the remaining one, designated
VpaK321, is a novel and highly efficient class-IIS ENase with the heptanucleotide recognition
site, 5'-GCTCTTC(1/4)-3'.

<>

<1>Miyahara, M., Shimada, T., Mise, K.
<2>Characterization of restriction endonucleases from Vibrio cholerae non 01.
<3>J. Food. Hyg. Sci. Japan
<4>35
<5>599-604
<6>1994
<7>Fourteen restriction endonucleases (ENases) were found by screening of 118 O antigen reference
strains of Vibrio cholerae. Ten of these ENases were partially purified and classified into
eight specificity categories. All of them were isoschizomers of already-known ENases. It is of
interest that the isoschizomer of EcoRI ENase designated VchO2I was found in V. cholerae O2,
since VchO2I, as well as EcoRI, shows high activity at pH 8-9, and most V. cholerae strains
can grow well in this pH range.

<>

<1>Miyahara, M., Shinohara, N., Mise, K.
<2>Purification of EcoO44I restriction endonuclease in Escherichia coli O44 isolated from an affected human.
<3>Eisei Shikenjo Hokoku
<4>114
<5>13-15
<6>1996
<7>A restriction endonuclease (Enase) designated EcoO44I  was purified without non-specific
nucleases from enteropathogenic Escherichia coli O44 Hiromi strain of affected human origin.
The yield was 1,100 units/g of wet cells.  The EcoO44I Enase recognized and cleaved the
specific sequence 5'-GGTCTC-3' (1/5) as was the case with Eco31I or BsaI Enase.  Because of
the stability and high yield, EcoO44I would be useful for recombinant DNA technology after
isolation of EcoO44-positive, avirulent mutant strains of E. coli O44 Hiromi.

<>

<1>Miyake, M., Asada, Y., Shiraki, M.
<2>Thermostable restriction endonuclease SelI - Synechococcus elongatus thermostable enzyme production, purification and characterization for use in DNA cleavage.
<3>Japanese Patent Office
<4>JP 6153937
<5>
<6>1994
<7>
<>

<1>Miyake, M., Asada, Y., Shiraki, M., Kato, S., Kotani, H.
<2>Thermostable restriction endonuclease SelI - and its preparation with Synechococcus elongatus.
<3>Japanese Patent Office
<4>JP 94153937 A
<5>
<6>1994
<7>
<>

<1>Miyake, M., Kotani, H., Asada, Y.
<2>Isolation and identification of restriction endonuclease, SelI from a cyanobacterium, Synechococcus elongatus.
<3>Nucleic Acids Res.
<4>20
<5>2605
<6>1992
<7>A restriction endonuclease, SelI has been isolated from a thermophilic and unicellular
cyanobacterium, Synechococcus elongatus. SelI, an isoschizomer of FnuDII, recognizes and
cleaves at the sequence, 5'-|CGCG-3'.

<>

<1>Miyake, T., Hiraishi, H., Sammoto, H., Ono, B.-I.
<2>Involvement of the VDE homing endonuclease and rapamycin in regulation of the Saccharomyces cerevisiae GSH11 gene encoding the high affinity  glutathione transporter.
<3>J. Biol. Chem.
<4>278
<5>39632-39636
<6>2003
<7>The Saccharomyces cerevisiae gene HGT1/GSH11 encodes the high affinity glutathione transporter
and is repressed by cysteine added to the culture
medium. It has been found previously that a 5'-upstream cis-element,
CCGCCACAC, is responsible for regulating GSH11 expression and that several
proteins bind to this element (Miyake, T., Kanayama, M., Sammoto, H., and
Ono, B. (2002) Mol. Genet. Genomics 266, 1004-1011). In this report we
present evidence that the most prominent of these proteins is VDE, known
previously as the homing endonuclease encoded by VMA1. We show also that
GSH11 is not expressed in a VDE-deleted strain and that inability to
express the GSH11 of this strain is overcome by introduction of the coding
region of VDE or the entire VMA1 gene. It is also found that VDE does not
cut DNA in the vicinity of the GSH11 cis-element. Rapamycin, an inhibitor
of the target of rapamycin (TOR) signal-transduction system, is found to
enhance expression of GSH11 in a VDE-dependent manner under conditions of
sulfur starvation. These results indicate that GSH11 is regulated by a
system sensitive to sulfur starvation (presumably via cysteine depletion)
and a more general system involving the nutritional starvation signal
mediated by the TOR system. Both systems need to be operational
(inhibition of TOR and sulfur starvation) for full expression of GSH11.

<>

<1>Miyamoto, H., Nakai, W., Yajima, N., Fujibayashi, A., Higuchi, T., Sato, K., Matsushiro, A.
<2>Sequence analysis of Stx2-converting phage VT2-Sa shows a great divergence in early regulation and replication regions.
<3>DNA Res.
<4>6
<5>235-240
<6>1999
<7>In enterohemorrhagic Escherichia coli, Shiga toxin is produced by lysogenic prophages. We have
isolated the prophage VT2-Sa that is responsible for production of Shiga toxin type 2 protein,
and determined the complete nucleotide sequence of this phage DNA. The entire DNA sequence
consisted of 60,942 bp, exhibiting marked similarity to the 933W phage genome. However,
several differences were observed in the immunity and replication regions, where cI, cII,
cIII, N, cro, O, and P genes were present: Predicted amino acid sequences of N, cI, cro, O and
P in the VT2-Sa genome did not show significant similarity to the counterparts of the 933W
genome; however its cI showed higher similarity to lambda. Furthermore, O and P closely
resembled those of phage HK022. These observations suggest that the various degrees of
homology observed in the immunity and replication regions of VT2-Sa could have resulted from
frequent recombination events among the lambdoid phages, and that these regions play a key
role as a functional unit for phage propagation in competition with other lambdoid phages.

<>

<1>Miyamoto, K., Chakrabarti, G., Morino, Y., McClane, B.A.
<2>Organization of the plasmid cpe locus in Clostridium perfringens type A isolates.
<3>Infect. Immun.
<4>70
<5>4261-4272
<6>2002
<7>Clostridium perfringens type A isolates causing food poisoning have a chromosomal enterotoxin
gene (cpe), while C. perfringens type A isolates
responsible for non-food-borne human gastrointestinal diseases carry a
plasmid cpe gene. In the present study, the plasmid cpe locus of the
type A non-food-borne-disease isolate F4969 was sequenced to design
primers and probes for comparative PCR and Southern blot studies of the
cpe locus in other type A isolates. Those analyses determined that the
region upstream of the plasmid cpe gene is highly conserved among type
A isolates carrying a cpe plasmid. The organization of the type A
plasmid cpe locus was also found to be unique, as it contains IS1469
sequences located similarly to those in the chromosomal cpe locus but
lacks the IS1470 sequences found upstream of IS1469 in the chromosomal
cpe locus. Instead of those upstream IS1470 sequences, a partial open
reading frame potentially encoding cytosine methylase (dcm) was
identified upstream of IS1469 in the plasmid cpe locus of all type A
isolates tested. Similar dcm sequences were also detected in several
cpe-negative C. perfringens isolates carrying plasmids but not in type
A isolates carrying a chromosomal cpe gene. Contrary to previous
reports, sequences homologous to IS1470, rather than IS1151, were found downstream of the
plasmid cpe gene in most type A isolates
tested. Those IS1470-like sequences reside in about the same position
but are oppositely oriented and defective relative to the IS1470
sequences found downstream of the chromosomal cpe gene. Collectively,
these and previous results suggest that the cpe plasmid of many type A
isolates originated from integration of a cpe-containing genetic
element near the dcm sequences of a C. perfringens plasmid. The
similarity of the plasmid cpe locus in many type A isolates is
consistent with horizontal transfer of a common cpe plasmid among C.
perfringens type A strains.

<>

<1>Miyamoto, K., Fisher, D.J., Li, J., Sayeed, S., Akimoto, S., McClane, B.A.
<2>Complete Sequencing and Diversity Analysis of the Enterotoxin-Encoding Plasmids in Clostridium perfringens Type A Non-Food-Borne Human Gastrointestinal Disease Isolates.
<3>J. Bacteriol.
<4>188
<5>1585-1598
<6>2006
<7>Enterotoxin-producing Clostridium perfringens type A isolates are an
important cause of food poisoning and non-food-borne human
gastrointestinal diseases, e.g., sporadic diarrhea (SPOR) and
antibiotic-associated diarrhea (AAD). The enterotoxin gene (cpe) is
usually chromosomal in food poisoning isolates but plasmid-borne in
AAD/SPOR isolates. Previous studies determined that type A SPOR isolate
F5603 has a plasmid (pCPF5603) carrying cpe, IS1151, and the beta2 toxin
gene (cpb2), while type A SPOR isolate F4969 has a plasmid (pCPF4969)
lacking cpb2 and IS1151 but carrying cpe and IS1470-like sequences. By
completely sequencing these two cpe plasmids, the current study identified
pCPF5603 as a 75.3-kb plasmid carrying 73 open reading frames (ORFs) and
pCPF4969 as a 70.5-kb plasmid carrying 62 ORFs. These plasmids share an
approximately 35-kb conserved region that potentially encodes virulence
factors and carries ORFs found on the conjugative transposon Tn916. The
34.5-kb pCPF4969 variable region contains ORFs that putatively encode two
bacteriocins and a two-component regulator similar to VirR/VirS, while the
approximately 43.6-kb pCPF5603 variable region contains a functional cpb2
gene and several metabolic genes. Diversity studies indicated that other
type A plasmid cpe(+)/IS1151 SPOR/AAD isolates carry a pCPF5603-like
plasmid, while other type A plasmid cpe(+)/IS1470-like SPOR/AAD isolates
carry a pCPF4969-like plasmid. Tn916-related ORFs similar to those in
pCPF4969 (known to transfer conjugatively) were detected in the cpe
plasmids of other type A SPOR/AAD isolates, as well as in representative
C. perfringens type B to D isolates carrying other virulence plasmids,
possibly suggesting that most or all C. perfringens virulence plasmids
transfer conjugatively.

<>

<1>Miyamoto, K., Yumine, N., Mimura, K., Nagahama, M., Li, J., McClane, B.A., Akimoto, S.
<2>Identification of Novel Clostridium perfringens Type E Strains That Carry an Iota Toxin Plasmid with a Functional Enterotoxin Gene.
<3>PLoS ONE
<4>6
<5>E20376
<6>2011
<7>Clostridium perfringens enterotoxin (CPE) is a major virulence factor for
human gastrointestinal diseases, such as food poisoning and antibiotic
associated diarrhea. The CPE-encoding gene (cpe) can be chromosomal or
plasmid-borne. Recent development of conventional PCR cpe-genotyping
assays makes it possible to identify cpe location (chromosomal or plasmid)
in type A isolates. Initial studies for developing cpe genotyping assays
indicated that all cpe-positive strains isolated from sickened patients
were typable by cpe-genotypes, but surveys of C. perfringens environmental
strains or strains from feces of healthy people suggested that this assay
might not be useful for some cpe-carrying type A isolates. In the current
study, a pulsed-field gel electrophoresis Southern blot assay showed that
four cpe-genotype untypable isolates carried their cpe gene on a plasmid
of  approximately 65 kb. Complete sequence analysis of the  approximately
65 kb variant cpe-carrying plasmid revealed no intact IS elements and a
disrupted cytosine methyltransferase (dcm) gene. More importantly, this
plasmid contains a conjugative transfer region, a variant cpe gene and
variant iota toxin genes. The toxin genes encoded by this plasmid are
expressed based upon the results of RT-PCR assays. The  approximately 65
kb plasmid is closely related to the pCPF4969 cpe plasmid of type A
isolates. MLST analyses indicated these isolates belong to a unique
cluster of C. perfringens. Overall, these isolates carrying a variant
functional cpe gene and iota toxin genes represent unique type E strains.

<>

<1>Miyamoto, S., Mizutani, R., Satow, Y., Kawasaki, M., Ohya, Y., Anraku, Y.
<2>Recognition and cleavage of double-stranded DNA by yeast VMA1-derived endonuclease.
<3>Nucleic Acids Symp. Ser.
<4>42
<5>197-198
<6>1999
<7>DNA endonuclease derived from the yeast VMA1-gene product recognizes and cleaves 31 base-pairs
of double-stranded DNA (dsDNA). Mixtures of the endonuclease (VDE) with a full DNA substrate
consisting of 34 base-pairs, with nicked substrates each having a nick in either DNA chain,
and with cleaved substrates each having a cleaved-off chain are prepared. Molecular weights
(MWs) of eluted peaks from gel filtration columns were estimated from elution profiles in the
presence of Mg2+ ions. Each mixture exhibited an eluted peak at about 63k MW, larger than the
MW of VDE unbound to dsDNA. This indicates that VDE and dsDNA substrates form stable
complexes. The mixture of VDE either with the full substrate or with the nicked substrate
having a nick in the anti-sense chain eluted an additional 25k-MW peak, which presumably
corresponds to a cleaved product. The complex of VDE with the full substrate was eluted at
62k-MW location in the absence of Mg2+ ions and yielded a single crystal. Stable complexes of
VDE either with the dsDNA substrates or with the cleaved products are obtainable.

<>

<1>Miyazaki, R., Sato, Y., Ito, M., Ohtsubo, Y., Nagata, Y., Tsuda, M.
<2>Complete nucleotide sequence of an exogenously isolated plasmid, pLB1, involved in .gamma.-hexachlorocyclohexane degradation.
<3>Appl. Environ. Microbiol.
<4>72
<5>6923-6933
<6>2006
<7>The alpha-proteobacterial strain Sphingobium japonicum UT26 utilizes a highly chlorinated
pesticide, gamma-hexachlorocyclohexane (gamma-HCH), as a sole source of carbon and energy, and
haloalkane dehalogenase LinB catalyzes the second step of gamma-HCH degradation in UT26.
Functional complementation of a linB mutant of UT26, UT26DB, was performed by the exogenous
plasmid isolation technique using HCH-contaminated soil, leading to our successful
identification of a plasmid, pLB1, carrying the linB gene. Complete sequencing analysis of
pLB1, with a size of 65,998 bp, revealed that it carries (i) 50 totally annotated coding
sequences, (ii) an IS6100 composite transposon containing two copies of linB, and (iii)
potential genes for replication, maintenance, and conjugative transfer with low levels of
similarity to other homologues. A minireplicon assay demonstrated that a 2-kb region
containing the predicted repA gene and its upstream region of pLB1 functions as an
autonomously replicating unit in UT26. Furthermore, pLB1 was conjugally transferred from
UT26DB to other alpha-proteobacterial strains but not to any of the beta- or
gamma-proteobacterial strains examined to date. These results suggest that this exogenously
isolated novel plasmid contributes to the dissemination of at least some genes for gamma-HCH
degradation in the natural environment. To the best of our knowledge, this is the first
detailed report of a plasmid involved in gamma-HCH degradation.

<>

<1>Miyazono, K., Furuta, Y., Watanabe-Matsui, M., Miyakawa, T., Ito, T., Kobayashi, I., Tanokura, M.
<2>A sequence-specific DNA glycosylase mediates restriction-modification in Pyrococcus abyssi.
<3>Nat. Commun.
<4>5
<5>1-8
<6>2014
<7>Restriction-modification systems consist of genes that encode a restriction enzyme and a
cognate methyltransferase. Thus far, it was believed that restriction enzymes are
sequence-specific endonucleases that introduce double-strand breaks at specific sites by
catalysing the cleavages of phosphodiester bonds. Here we report that based on the crystal
structure and enzymatic activity, one of the restriction enzymes, R.PabI, is not an
endonuclease but a sequence-specific adenine DNA glycosylase. The structure of the R.PabI-DNA
complex shows that R.PabI unwinds DNA at a 5'-GTAC-3' site and flips the guanine and adenine
bases out of the DNA helix to recognize the sequence. R.PabI catalyses the hydrolysis of the
N-glycosidic bond between the adenine base and the sugar in the DNA and produces two opposing
apurinic/apyrimidinic (AP) sites. The opposing AP sites are cleaved by heat-promoted beta
elimination and/or by endogenous AP endonucleases of host cells to introduce a double-strand
break.

<>

<1>Miyazono, K., Watanabe, M., Kosinski, J., Ishikawa, K., Kamo, M., Sawasaki, T., Nagata, K., Bujnicki, J.M., Endo, Y., Tanokura, M., Kobayashi, I.
<2>Novel protein fold discovered in the PabI family of restriction enzymes.
<3>Nucleic Acids Res.
<4>35
<5>1908-1918
<6>2007
<7>Although structures of many DNA-binding proteins have been solved, they fall into a limited
number of folds. Here, we describe an approach that
led to the finding of a novel DNA-binding fold. Based on the behavior of
Type II restriction-modification gene complexes as mobile elements, our
earlier work identified a restriction enzyme, R.PabI, and its cognate
modification enzyme in Pyrococcus abyssi through comparison of closely
related genomes. While the modification methyltransferase was easily
recognized, R.PabI was predicted to have a novel 3D structure. We
expressed cytotoxic R.PabI in a wheat-germ-based cell-free translation
system and determined its crystal structure. R.PabI turned out to adopt a
novel protein fold. Homodimeric R.PabI has a curved anti-parallel
beta-sheet that forms a 'half pipe'. Mutational and in silico DNA-binding
analyses have assigned it as the double-strand DNA-binding site. Unlike
most restriction enzymes analyzed, R.PabI is able to cleave DNA in the
absence of Mg(2+). These results demonstrate the value of genome
comparison and the wheat-germ-based system in finding a novel DNA-binding
motif in mobile DNases and, in general, a novel protein fold in
horizontally transferred genes.

<>

<1>Miyoshi-Akiyama, T., Kuwahara, T., Tada, T., Kitao, T., Kirikae, T.
<2>Complete Genome Sequence of Highly Multidrug-Resistant Pseudomonas aeruginosa NCGM2.S1, a Representative Strain of a Cluster Endemic to  Japan.
<3>J. Bacteriol.
<4>193
<5>7010
<6>2011
<7>We report the completely annotated genome sequence of Pseudomonas aeruginosa NCGM2.S1, a
representative strain of a cluster endemic to Japan
with a high level of resistance to carbapenem (MIC >/= 128 mug/ml),
amikacin (MIC >/= 128 mug/ml), and fluoroquinolone (MIC >/= 128 mug/ml).

<>

<1>Miyoshi-Akiyama, T., Matsumura, K., Iwai, H., Funatogawa, K., Kirikae, T.
<2>Complete Annotated Genome Sequence of Mycobacterium tuberculosis Erdman.
<3>J. Bacteriol.
<4>194
<5>2770
<6>2012
<7>We report the completely annotated genome sequence of Mycobacterium tuberculosis  Erdman (TMC
107; ATCC 35801), which is a well-known laboratory strain of M.
tuberculosis.

<>

<1>Miyoshi-Akiyama, T., Matsumura, K., Kobayashi, N., Maeda, S., Kirikae, T.
<2>Genome Sequence of Clinical Isolate Mycobacterium tuberculosis NCGM2209.
<3>J. Bacteriol.
<4>193
<5>6792
<6>2011
<7>We report the annotated genome sequence of a clinical isolate, Mycobacterium tuberculosis
strain NCGM2209, which belongs to the 'Beijing
family' and was isolated in Japan.

<>

<1>Miyoshi-Akiyama, T., Satou, K., Kato, M., Shiroma, A., Matsumura, K., Tamotsu, H., Iwai, H., Teruya, K., Funatogawa, K., Hirano, T., Kirikae, T.
<2>Complete annotated genome sequence of Mycobacterium tuberculosis (Zopf) Lehmann and Neumann (ATCC35812) (Kurono).
<3>Tuberculosis
<4>95
<5>37-39
<6>2014
<7>We report the completely annotated genome sequence of Mycobacterium tuberculosis
(Zopf) Lehmann and Neumann (ATCC35812) (Kurono), which is a used for virulence
and/or immunization studies. The complete genome sequence of M. tuberculosis
Kurono was determined with a length of 4,415,078 bp and a G+C content of 65.60%.
The chromosome was shown to contain a total of 4,340 protein-coding genes, 53
tRNA genes, one transfer messenger RNA for all amino acids, and 1 rrn operon.
Lineage analysis based on large sequence polymorphisms indicated that M.
tuberculosis Kurono belongs to the Euro-American lineage (lineage 4).
Phylogenetic analysis using whole genome sequences of M. tuberculosis Kurono in
addition to 22 M. tuberculosis complex strains indicated that H37Rv is the
closest relative of Kurono based on the results of phylogenetic analysis. These
findings provide a basis for research using M. tuberculosis Kurono, especially in
animal models.

<>

<1>Miyoshi-Akiyama, T., Takeshita, N., Ohmagari, N., Kirikae, T.
<2>Complete Genome Sequence of Helicobacter cinaedi Type Strain ATCC BAA-847.
<3>J. Bacteriol.
<4>194
<5>5692
<6>2012
<7>Here we report the completely annotated genome sequence of the Helicobacter cinaedi type
strain (ATCC BAA-847), which is an emerging pathogen that causes
cellulitis and bacteremia. The genome sequence will provide new insights into the
diagnosis, pathogenic mechanisms, and drug resistance of H. cinaedi.

<>

<1>Miyoshi-Akiyama, T., Watanabe, S., Kirikae, T.
<2>Complete Genome Sequence of Streptococcus pyogenes M1 476, Isolated from a Patient with Streptococcal Toxic Shock Syndrome.
<3>J. Bacteriol.
<4>194
<5>5466
<6>2012
<7>Here, we report the completely annotated genome sequence of Streptococcus pyogenes M1 476
isolated from a patient with streptococcal toxic shock syndrome
(STSS) during pregnancy. The genome sequence will provide new insights into the
mechanisms underlying STSS.

<>

<1>Mizoguchi, H., Mori, H., Hara, K.
<2>Useful microorganisms for industry.
<3>International Patent Office
<4>WO 2006057341 A
<5>
<6>2006
<7>After culturing for a definite period of time, a mutant of wild type Escherichia coli, which
has a chromosomal DNA shorter by 470 kbp or more than the chromosomal DNA of the wild type,
shows a larger cell count than the wild type.  Therefore, a useful substance such as a
protein, a peptide, an amino acid, a nucleic acid, a vitamin, a saccharide, an organic acid or
a lipid can be efficiently produced by culturing the above-described mutant, a medium thus
allowing it to produce and accumulate the useful substance in the culture medium.

<>

<1>Mizumura, H., Shibata, T., Morishima, N.
<2>Stable association of 70-kDa heat shock protein induces latent multisite specificity of a unisite-specific endonuclease in yeast mitochondria.
<3>J. Biol. Chem.
<4>274
<5>25682-25690
<6>1999
<7>The multisite-specific endonuclease Endo.SceI of yeast mitochondria is unique among
endonucleases because its 50-kDa subunit forms a stable dimer with the mitochondrial 70-kDa
heat shock protein (mtHSP70), which otherwise fulfills a chaperone function by binding
transiently to unfolded proteins. Here we show that the mtHSP70 subunit confers broader
sequence specificity, greater stability, and higher activity on the 50-kDa subunit. The 50-kDa
subunit alone displayed weaker activity and highly sequence-specific endonuclease activity.
The 50-kDa protein exists as a heterodimer with mtHSP70 in vivo, allowing Endo.SceI to cleave
specifically at multiple sites on mitochondrial DNA. Endo.SceI may have evolved from a highly
specific endonuclease that gained broader sequence specificity after becoming a stable partner
of mtHSP70.

<>

<1>Mizumura, H., Shibata, T., Morishima, N.
<2>Association of HSP70 with endonucleases allows the expression of otherwise silent mutations.
<3>FEBS Lett.
<4>522
<5>177-182
<6>2002
<7>A subpopulation of the 70 kDa heat shock protein (HSP70) found within the mitochondria of
Saccharomyces cerevisiae functions as a stable
binding partner of the endonuclease SceI. We have previously found that
the SceI endonuclease monomer recognizes and cleaves a unique, 26 bp
sequence in vitro. Dimerization with HSP70 changes the specificity of
SceI, allowing it to cleave at multiple sequences. This study shows
that SuvI, an ortholog of SceI isolated from a different yeast strain,
contains two amino acid substitutions, yet it shows the same uni-site
specificity in its monomeric form. Binding of HSP70 to the SuvI monomer
confers multi-site specificity that is different from that exhibited by
the HSP70/SceI heterodimer. Mutation of single residues of SceI to the
corresponding residue in SuvI provides enzymes with specificities
intermediate between SceI and SuvI when complexed with HSP70. These
results suggest that HSP70 interaction with certain endonucleases
allows the expression of otherwise silent mutations in them, causing a
change in enzyme cleavage specificity.

<>

<1>Mizuno, A., Nagamune, T., Mioda, M.
<2>Control of enzyme reaction by cold mixing of restriction endonuclease and sample, e.g. DNA, freezing and laser irradiation of selected regions for selective DNA cleavage reaction.
<3>Japanese Patent Office
<4>JP 7184665
<5>
<6>1995
<7>
<>

<1>Mizuno, C.M., Rodriguez-Valera, F., Garcia-Heredia, I., Martin-Cuadrado, A.B., Ghai, R.
<2>Reconstruction of Novel Cyanobacterial Siphovirus Genomes from Mediterranean Metagenomic Fosmids.
<3>Appl. Environ. Microbiol.
<4>79
<5>688-695
<6>2013
<7>Cellular metagenomes are primarily used for investigating microbial community
structure and function. However, cloned fosmids from such metagenomes capture
phage genome fragments that can be used as a source of phage genomes. We show
that fosmid cloning from cellular metagenomes and sequencing at a high coverage
is a credible alternative to constructing metaviriomes and allows capturing and
assembling novel, complete phage genomes. It is likely that phages recovered from
cellular metagenomes are those replicating within cells during sample collection
and represent "active" phages, naturally amplifying their genomic DNA and
increasing chances for cloning. We describe five sets of siphoviral contigs
(MEDS1, MEDS2, MEDS3, MEDS4 and MEDS5), obtained by sequencing fosmids from the
cellular metagenome of the deep chlorophyll maximum in the Mediterranean. Three
of these represent complete siphoviral genomes and two partial ones. These are
the first set of phage genomes assembled directly from cellular metagenomic
fosmid libraries. They exhibit low sequence similarities to one another and to
known siphoviruses, but are remarkably similar in overall genome architecture. We
present evidence suggesting they infect picocyanobacteria, likely Synechococcus.
Four of these sets also define a novel branch in the phylogenetic tree of phage
large subunit terminases. Moreover, some of these siphoviral groups are globally
distributed and abundant in the oceans, comparable to some known myoviruses and
podoviruses. This suggests that as more siphoviral genomes become available, we
will be better able to assess the abundance and influence of this diverse and
polyphyletic group in the marine habitat.

<>

<1>Mizuno, C.M., Rodriguez-Valera, F., Kimes, N.E., Ghai, R.
<2>Expanding the marine virosphere using metagenomics.
<3>PLoS Genet.
<4>9
<5>e1003987
<6>2013
<7>Viruses infecting prokaryotic cells (phages) are the most abundant entities of the biosphere
and contain a largely uncharted
wealth of genomic diversity. They play a critical role in the biology of their hosts and in
ecosystem functioning at large. The
classical approaches studying phages require isolation from a pure culture of the host. Direct
sequencing approaches have
been hampered by the small amounts of phage DNA present in most natural habitats and the
difficulty in applying metaomic
approaches, such as annotation of small reads and assembly. Serendipitously, it has been
discovered that cellular
metagenomes of highly productive ocean waters (the deep chlorophyll maximum) contain
significant amounts of viral DNA
derived from cells undergoing the lytic cycle. We have taken advantage of this phenomenon to
retrieve metagenomic
fosmids containing viral DNA from a Mediterranean deep chlorophyll maximum sample. This method
allowed description of
complete genomes of 208 new marine phages. The diversity of these genomes was remarkable,
contributing 21 genomic
groups of tailed bacteriophages of which 10 are completely new. Sequence based methods have
allowed host assignment
to many of them. These predicted hosts represent a wide variety of important marine
prokaryotic microbes like members of
SAR11 and SAR116 clades, Cyanobacteria and also the newly described low GC Actinobacteria. A
metavirome constructed
from the same habitat showed that many of the new phage genomes were abundantly represented.
Furthermore, other
available metaviromes also indicated that some of the new phages are globally distributed in
low to medium latitude ocean
waters. The availability of many genomes from the same sample allows a direct approach to
viral population genomics
confirming the remarkable mosaicism of phage genomes.

<>

<1>Mizuno, H., Suzuki, T., Akagawa, M., Yamasato, K., Yamada, Y.
<2>Purification, properties and determination of recognition sequence and cleavage site of restriction endonuclease from Agrobacterium gelatinovorum IAM 12617, a marine bacterium (AgeI).
<3>Agric. Biol. Chem.
<4>54
<5>1797-1802
<6>1990
<7>A new restriction endonuclease, designated as AgeI, was purified from cell-free
extracts of a marine bacterium, Agrobacterium gelatinovorum IAM 12617 by
streptomycin treatment, ammonium sulfate fractionation, combined column
chromatographies on heparin-Sepharose CL-6B and DEAE-Sepharose CL-6B and FPLC
on Mono Q (HR 5/5) and Superose 12 (HR 10/30).  The purified enzyme was
homogenous on SDS-polyacrylamide gel disc electrophoresis and free from other
phosphatase and exonuclease activities on ligation-recutting test.  The
relative molecular mass of the enzyme was 24,000 daltons by SDS-polyacrylamide
gel disc electrophoresis.  The gel filtration using Superose 12 (HR 10/30) gave
the same calculation (23,000 daltons).  These data indicated that the enzyme is
a monomer.  The isoelectric point of the enzyme was 6.5.  The purified enzyme
cleaved lambda and Ad2 DNAs at 10 or more and 5 sites, respectively.  However,
the purified enzyme did not cleave SV40, PhiX174 RF I, M13mp18 RF I or pBR322
DNAs.  pBR328 DNA was cleaved at 1 site by the purified enzyme.  The purified
enzyme worked best at 37C and pH 7.5 in a reaction mixture (50 microliters)
containing 1.0 micrograms lambda DNA, 10 mM Tris-HCl, 7 mM 2-mercaptoethanol, 7
mM MgCl2 and 50 mM NaCl.  The purified enzyme did not require monovalent
cations necessarily for the enzyme reaction.  The enzyme recognized the
palindromic hexanucleotide DNA sequence 5'-ACCGGT-3' and cut between A and C,
producing a 5'-cohesive tetranucleotide extension.

<>

<1>Mizuno, H., Suzuki, T., Yamada, Y., Akagawa, M., Yamasato, K.
<2>Purification and properties of restriction endonuclease from Deleya marina IAM 14114, a marine bacterium (DmaI).
<3>Agric. Biol. Chem.
<4>54
<5>2863-2867
<6>1990
<7>A restriction endonuclease, designated as DmaI, was purified from cell-free
extracts of Deleya marina IAM 14114 by streptomycin treatment, ammonium sulfate
fractionation and two steps of chromatographicy on Heparin-Sepharose CL-6B and
Mono Q (HR 5/5, FPLC).  The purified enzyme was homogeneous on
SDS-polyacrylamide gel disk electrophoresis and a ligation-recutting test.  The
relative molecular mass measurements of the purified enzyme gave 28,000 daltons
by SDS-polyacrylamide gel disk electrophoresis and 56,000 daltons by gel
filtration.  These data indicated that the purified enzyme (56,000 daltons) has
a dimeric structure composed of two 28,000-dalton subunits.  The isoelectric
point was 5.5.  The purified enzyme worked best at 37C in a reaction mixture
(50 microliters) containing 1.0 microgram lambda DNA, 10 mM Tris-HCI, 7 mM
2-mercaptoethanol, 7 mM MgCl2 and 100 mM NaCl (pH 7.5).  The enzyme was stable
up to 55C and between pH 7.0 and 9.0.  The purified enzyme recognizes the
palindromic hexanucleotide DNA sequence 5'-CAGCTG-3', cuts between G and C and
produces a flush end (isoschizomer of PvuII).

<>

<1>Mizutani, R., Anraku, Y., Satow, Y.
<2>Protein splicing of yeast VMA1-derived endonuclease via thiazolidine intermediates.
<3>J. Synchrotron Radiat.
<4>11
<5>109-112
<6>2004
<7>Protein splicing precisely excises out an internal intein segment from a protein precursor,
and concomitantly ligates the N- and C-terminal extein polypeptides flanking the intein.  A
recombinant X10SNS bearing N- and C-extein polypeptides has been prepared for the intein
endonuclease derived from the Saccharomyces cerevisiae VMA1 gene.  X10SNS has replacements of
C284S, H362N, and C738S, and forms the intein and extein segments in the crystal lattice.  The
crystal structure of X10SNS revealed a linkage between the N- and C-extein segments, and
showed that the C284 amino group of the resultant intein segment is in interaction with the
G283 O atom of the N-extein segment.  A mechanism for the final S-N acyl shift step proposes
that a tetrahedral intermediate involves a five-membered thiazolidine ring at G283-C738
junction.  An oxyanion of the thiazolidine intermediate is to be stabilized by the C284 N
atom.

<>

<1>Mizutani, R., Nogami, S., Kawasaki, M., Ohya, Y., Anraku, Y., Satow, Y.
<2>Protein-splicing reaction via a thiazolidine intermediate: Crystal structure of the VMA1-derived endonuclease bearing the N and C-terminal  propeptides.
<3>J. Mol. Biol.
<4>316
<5>919-929
<6>2002
<7>Protein splicing excises an internal intein segment from a protein precursor precisely, and
concomitantly ligates flanking N and C-extein
polypeptides at the respective sides of the precursor. Here, a series
of precursor recombinants bearing 11 N-extein and ten C-extein residues
is prepared for the intein of the Saccharomyces cerevisiae VMAI-derived
homing endonuclease referred to as VIDE and as PI-SceI. The recombinant
with replacements of C284S, H362N, N737S, and C738S is chosen as a
splice-able precursor model and is then subjected to a 2.1 Angstrom
resolution crystallographic analysis. The crystal structure shows that
the introduced extein polypeptides are located in the vicinity of the
splicing site, and that each of their peptide bonds is in the trans
conformation. The S284 O-gamma atom located at a distance of 3.1
Angstrom from the G283 C atom in the N-terminal junction suggests that
a nucleophilic attack of the C284 S-gamma atom on the G283 C atom forms
a tetrahedral intermediate containing a five-membered thiazolidine
ring. The tetrahedral intermediate is supposedly resolved into a
thioester acyl group upon the cleavage of the linkage between the G283
C and C284 N atoms, and this thioester acyl formation completes the
initial steps of N --> S acyl shift at the junction between the
N-extein and intein. The S738 O-gamma atom in the C-terminal junction
is placed in close proximity to the S284 O-gamma atom at a distance of
3.6 Angstrom, and is well suited for another nucleophilic attack on the
resultant thioester acyl group that is then subjected to the
transesterification in the next step. The reaction steps proposed for
the acyl shift are driven entirely by protonation and deprotonation, in
which proton ingress and egress is balanced within the splicing site.

<>

<1>Mizuuchi, K., Nobbs, T.J., Halford, S.E., Adzuma, K., Qin, J.
<2>A new method for determining the Stereochemistry of DNA cleavage reactions: Application to the SfiI and HpaII restriction endonucleases and to the MuA transposase.
<3>Biochemistry
<4>38
<5>4640-4648
<6>1999
<7>A new method was developed for tracking the stereochemical path of enzymatic cleavage of DNA.
DNA with a phosphorothioate of known chirality at the scissile bond is cleaved by the enzyme
in H218O. The cleavage produces a DNA molecule with the 5'-[16O,18O, S]-thiophosphoryl group,
whose chirality depends on whether the cleavage reaction proceeds by a single-step hydrolysis
mechanism or by a two-step mechanism involving a protein-DNA covalent intermediate. To
determine this chirality, the cleaved DNA is joined to an oligonucleotide by DNA ligase. Given
the strict stereochemistry of the DNA ligase reaction, determined here, the original chirality
of the phosphorothioate dictates whether the 18O is retained or lost in the ligation product,
which can be determined by mass spectrometry. This method has advantages over previous methods
in that it is not restricted to particular DNA sequences, requires substantially less
material, and avoids purification of the products at intermediate stages in the procedure. The
method was validated by confirming that DNA cleavage by the EcoRI restriction endonuclease
causes inversion of configuration at the scissile phosphate. It was then applied to the
reactions of the SfiI and HpaII endonucleases and the MuA transposase. In all three cases, DNA
cleavage proceeded with inversion of configuration, indicating direct hydrolysis of the
phosphodiester bond by water as opposed to a reaction involving a covalent enzyme-DNA
intermediate.

<>

<1>Mo, D., Wu, L., Xu, Y., Ren, J., Wang, L., Huang, L., Wu, Q.-J., Bao, P., Xie, M.-H., Yin, P., Liu, B.-F., Liang, Y., Zhang, Y.
<2>A maturase that specifically stabilizes and activates its cognate group  I intron at high temperatures.
<3>Biochimie
<4>93
<5>533-541
<6>2011
<7>Folding of large structured RNAs into their functional tertiary structures at high
temperatures is challenging. Here we show that
I-Tnal protein, a small LAGLIDADG homing endonuclease encoded by a
group I intron from a hyperthermophilic bacterium, acts as a maturase
that is essential for the catalytic activity of this intron at high
temperatures and physiological cationic conditions. I-Tnal specifically
binds to and induces tertiary packing of the P4-P6 domain of the
intron: this RNA protein complex might serve as a thermostable platform
for active folding of the entire intron. Interestingly, the binding
affinity of I-Tnal to its cognate intron RNA largely increases with
temperature; over 30-fold stronger binding at higher temperatures
relative to 37 degrees C correlates with a switch from an
entropy-driven (37 degrees C) to an enthalpydriven (55-60 degrees C)
interaction mode. This binding mode may represent a novel strategy how
an RNA binding protein can promote the function of its target RNA
specifically at high temperatures.

<>

<1>Mobberley, J.M., Authement, R.N., Segall, A.M., Paul, J.H.
<2>The temperate marine phage PhiHAP-1 of Halomonas aquamarina possesses a linear plasmid-like prophage genome.
<3>J. Virol.
<4>82
<5>6618-6630
<6>2008
<7>A myovirus-like temperate phage, PhiHAP-1, was induced with mitomycin C
from a Halomonas aquamarina strain isolated from surface waters in the
Gulf of Mexico. The induced cultures produced significantly more
virus-like particles (VLPs) (3.73 x 10(10) VLP ml(-1)) than control
cultures (3.83 x 10(7) VLP ml(-1)) when observed with epifluorescence
microscopy. The induced phage was sequenced by using linker-amplified
shotgun libraries and contained a genome 39,245 nucleotides in length with
a G+C content of 59%. The PhiHAP-1 genome contained 46 putative open
reading frames (ORFs), with 76% sharing significant similarity (E value of
<10(-3)) at the protein level with other sequences in GenBank. Putative
functional gene assignments included small and large terminase subunits,
capsid and tail genes, an N6-DNA adenine methyltransferase, and
lysogeny-related genes. Although no integrase was found, the PhiHAP-1
genome contained ORFs similar to protelomerase and parA genes found in
linear plasmid-like phages with telomeric ends. Southern probing and PCR
analysis of host genomic, plasmid, and PhiHAP-1 DNA indicated a lack of
integration of the prophage with the host chromosome and a difference in
genome arrangement between the prophage and virion forms. The linear
plasmid prophage form of PhiHAP-1 begins with the protelomerase gene,
presumably due to the activity of the protelomerase, while the induced
phage particle has a circularly permuted genome that begins with the
terminase genes. The PhiHAP-1 genome shares synteny and gene similarity
with coliphage N15 and vibriophages VP882 and VHML, suggesting an
evolutionary heritage from an N15-like linear plasmid prophage ancestor.

<>

<1>Mochizuki, A., Yahara, K., Kobayashi, I., Iwasa, Y.
<2>Genetic addiction: selfish gene's strategy for symbiosis in the genome.
<3>Genetics
<4>172
<5>1309-1323
<6>2006
<7>The evolution and maintenance of the phenomenon of postsegregational host killing or genetic
addiction are paradoxical. In this phenomenon, a gene complex, once
established in a genome, programs death of a host cell that has eliminated it.
The intact form of the gene complex would survive in other members of the host
population. It is controversial as to why these genetic elements are maintained,
due to the lethal effects of host killing, or perhaps some other properties are
beneficial to the host. We analyzed their population dynamics by analytical
methods and computer simulations. Genetic addiction turned out to be advantageous
to the gene complex in the presence of a competitor genetic element. The
advantage is, however, limited in a population without spatial structure, such as
that in a well-mixed liquid culture. In contrast, in a structured habitat, such
as the surface of a solid medium, the addiction gene complex can increase in
frequency, irrespective of its initial density. Our demonstration that genomes
can evolve through acquisition of addiction genes has implications for the
general question of how a genome can evolve as a community of potentially selfish
genes.

<>

<1>Modise, T., Ryder, C., Mane, S.P., Bandara, A.B., Jensen, R.V., Inzana, T.J.
<2>Genomic Comparison between a Virulent Type A1 Strain of Francisella tularensis and Its Attenuated O-Antigen Mutant.
<3>J. Bacteriol.
<4>194
<5>2775-2776
<6>2012
<7>We report the complete genome sequences of TI0902, a highly virulent type A1 strain, and
TIGB03, a related, attenuated chemical mutant strain. Compared to the
wild type, the mutant strain had 45 point mutations and a 75.9-kb duplicated
region that had not been previously observed in Francisella species.

<>

<1>Modrich, P.
<2>Studies on sequence recognition by type II restriction and modification enzymes.
<3>CRC Crit. Rev. Biochem.
<4>13
<5>287-323
<6>1982
<7>DNA restriction endonucleases and modification methylases are strain-specific
enzymes responsible for the host-specific barriers to interstrain transfer of
DNA that have been identified in numerous prokaryotic cell types.  Foreign DNA
entering a bacterial cell is subject to rapid endonucleolytic hydrolysis if it
is devoid of the modification characteristic of the particular bacterial
strain.  The strain specific modification enzyme catalyzes methyl transfer from
S-adenosyl-L-methionine (AdoMet) to a specific DNA sequence which is
characteristic of the particular host specificity system.  Thus, cellular DNA
is rendered resistant to attack by the endogenous restriction enzyme by virtue
of being methylated at DNA sites that are also recognized by the endonuclease.

<>

<1>Modrich, P.
<2>Structures and mechanisms of DNA restriction and modification enzymes.
<3>Q. Rev. Biophys.
<4>12
<5>315-369
<6>1979
<7>Although the phenomenon of host specificity was initially observed in the early
1950s (Luria & Human, 1952; Bertani & Weigle, 1953), it was nearly a decade
later that Arber and his colleagues accurately predicted the molecular basis of
the phenomenon.  Their experiments with bacteriophage lambda demonstrated that
a given host-specificity system imparts a specific modification of the viral
DNA, and further, that restriction of DNA lacking the appropriate modificaton
is a consequence of nucleolytic hydrolysis upon entry into the host cell (Arber
& Dussoix, 1962; Dussoix & Arber, 1962; Arber, Hattman & Dussoix, 1963).  These
observations led to their proposal that host specificity is based on a
two-enzyme system.  They suggested that cells of a given host specificity
possess a restriction endodeoxyribonuclease that recognizes a unique sequence
of nucleotide pairs and introduces double strand breaks into unmodified DNA.
The second component of the system, a modification enzyme, was proposed to
recognize the same nucleotide sequence and to modify the DNA, yielding a
species which is no longer subject to hydrolysis by the restriction
endonuclease.  Moreover, a variety of biological experiments suggested a
correlation between the phenomenon of modifications and polynucleotide
methylation (Arber, 1965; Klein & Sauerbier, 1965; Arber & Smith, 1966).  Thus,
cellular DNA would be resistant to restriction cleavage by virtue of being
appropriately methylated.  DNA foreign to the cell and lacking the appropriate
modification would, however, be specifically recognized and hydrolysed by the
restriction endonuclease.  Such systems then could account for the observed
barriers to transfer of unmodified DNA elements between prokaryotic cell types.

<>

<1>Modrich, P., Roberts, R.J.
<2>Type-II restriction and modification enzymes.
<3>Nucleases, Cold Spring Harbor Laboratory Press, Linn, S.M., Roberts, R.J., Cold Spring Harbor, New York
<4>0
<5>109-154
<6>1982
<7>I. IntroductionII. Structure of type-II restriction and modification enzymes.III. Methyl
transfer by type-II modification enzymes.IV. Fidelity of type-II restriction and modification
enzymes.V. Type-II enzyme-DNA interaction: thermodynamic and Kinetic parameters.VI. Type-II
enzyme-DNA interaction: DNA determinants important in specific recognition.VII. Concluding
remarks.

<>

<1>Modrich, P., Rubin, R.A.
<2>Role of the 2-amino group of deoxyguanosine in sequence recognition by EcoRI restriction and modifcation enzymes.
<3>J. Biol. Chem.
<4>252
<5>7273-7278
<6>1977
<7>The dG residues within the EcoRI recognition sequence of ColE1 DNA have been
selectively replaced with dI.  Methylation of the altered sequence by the EcoRI
modification enzyme is extremely slow as compared with methyl transfer to the
natural recognition site.  Since the affinity of the modification enzyme for
the dI-containing sequence is considerably less than that for the nature
sequence, we have concluded that the 2-amino group of dG has an important role
in DNA site recognition by this enzyme.  In contrast, the altered site is
subject to cleavage by EcoRI endonuclease at rates essentially identical with
those observed with the natural sequence.  These results strongly suggest that
the two enzymes utilize different contacts within the EcoRI site and are
consistent with our conclusion (Rubin, R.A., and Modrich, P. (1977) J. Biol.
Chem. 252, 7265-7272) that the two proteins interact with their common
recognition sequence in different ways.

<>

<1>Modrich, P., Zabel, D.
<2>EcoRI endonuclease.  Physical and catalytic properties of the homogeneous enzyme.
<3>J. Biol. Chem.
<4>251
<5>5866-5874
<6>1976
<7>A procedure for large scale isolation of Escherichia coli RI endonuclease in
high yield has been developed.  The purified enzyme is homogenous as judged by
polyacrylamide gel electrophoresis and analytical sedimentation.  The denatured
and reduced form of the enzyme has a molecular weight of 28,500 -/+ 500.  In
solution the enzyme exists as a mixture of dimers and tetramers of molecular
weights 57,000 and 114,000, respectively.  We estimate the dissociation
constant for tetramer to dimer transition to be less than or approximately
equal to 1 x 10-7 M.  Steady state kinetic analysis of the endonuclease with
ColE1 DNA as substrate showed that the enzyme obeys Michaelis-Menten kinetics.
At 37C the turnover number is four double strand scissions per min, and the Km
for ColE1 molecules is 8 x 10-9 M.  At 0C the major product of endonuclease
action contains only one single strand break in the RI site, and such molecules
can dissociate from the enzyme.  In contrast, at 30C or 37C, two single strand
breaks are introduced into the RI sequence prior to dissociation of the enzyme.
A transient enzyme-bound intermediate containing only one break in the RI site
was observed in studies of a single turnover at 30C.  Kinetic analysis of this
reaction indicates that the first break is introduced into the RI site with a
first order rate constant of at elast 40 min-1, while the second cleavage
occurs with a rate constant of 14 min-1.  Since the turnover number of the
enzyme at 30C is only 0.72 min-1, these results indicate that the rate-limiting
step is release of endonuclease from its DNA product.

<>

<1>Moencke-Buchner, E., Mackeldanz, P., Krueger, D.H., Reuter, M.
<2>Overexpression and affinity chromatography purification of the Type III restriction endonuclease EcoP15I for use in transcriptome analysis.
<3>J. Biotechnol.
<4>114
<5>99-106
<6>2004
<7>The Type III restriction endonuclease EcoP15I is a multifunctional hetero-oligomeric enzyme
that recognizes the non-symmetric DNA sequence 5'-CAGCAG. For efficient cleavage, EcoP15I
needs the interaction with two copies of the recognition sequence that have to be inversely
oriented in the DNA double strand. The enzyme cuts the upper DNA strand 25-26bp and the lower
DNA strand 27-28bp, respectively, downstream of the recognition sequence-a distinct feature
that makes the enzyme particularly valuable for gene expression profiling methods relying on
the SAGE procedure (Matsumura et al., PNAS 100, 15718, 2003). Because the broader use of this
transcriptome analysis method requires the availability of larger amounts of restriction
endonuclease EcoP15I and the enzyme is not commercially available, we have cloned the genes
coding for the EcoP15I restriction endonuclease into pQE-16 plasmid vector that provides the
enzyme with a C-terminal 6xHis-tag. After Ni-NTA affinity chromatography and ion exchange
chromatography on heparin sepharose, we obtained 5mg homogeneous EcoP15I per gram cell pellet
within 1-2 day(s). Moreover, the C-terminally 6xHis-tagged EcoP15I restriction endonuclease
shows comparable enzymatic activity as the untagged enzyme.

<>

<1>Moffatt, B.A., Studier, F.W.
<2>Entry of bacteriophage T7 DNA into the cell and escape from host restriction.
<3>J. Bacteriol.
<4>170
<5>2095-2105
<6>1988
<7>T7 DNA did not become susceptible to degradation by the host restriction
enzymes EcoB, EcoK, or EcoP1 until 6 to 7 min. after infection (at 30C).
During this period, T7 gene 0.3 protein is made and inactivates EcoB and EcoK,
allowing wild-type T7, or even a mutant that has recognition sites flanking
gene 0.3, to escape restriction by these enzymes.  However, T7 failed to escape
restriction by EcoP1 even though 0.3 protein was made, evidently because 0.3
protein is unable to inactivate EcoP1.  How T7 DNA can be accessible to
transcription but not restriction in the first few minutes of infection is not
yet understood, but we favor the idea that the entering DNA is initially
segregated in a special place.  Entry of T7 DNA into the cell is normally
coupled to transcription.  Tests of degradation of DNAs having their first
restriction sites different distances from the end of the DNA indicated that
only the first 1,000 or so base pairs (2.5%) of the molecule enter the cell
without transcription.  An exception was the only mutant tested that lacks base
pairs 343 to 393 of T7 DNA; most or all of this DNA entered the cell without
being transcribed, apparently because it lacks a sequence that normally arrests
entry.  This block to DNA entry would normally be relieved by the host RNA
polymerase transcribing from an appropriately situated promoter, but the block
can also be relieved by T7 RNA polymerase, if supplied by the host cell.  T7
mutants that lack all three strong early promoters A1, A2, and A3 could grow by
using a secondary promoter.

<>

<1>Mogila, V.A., Bellaiche, Y., Perrimon, N.
<2>Expression of I-SceI in Drosophila to induce DNA double-strand breaks.
<3>Methods Mol. Biol.
<4>113
<5>439-445
<6>1999
<7>Generation of double-strand breaks in chromosomal DNA induces repair machinery of a cell, and
is also a necessary step for recombination events.  A system for the directed introduction of
DSBs into a genome could substantially facilitate progress in understanding DSB repair
mechanisms and could be used for efficient gene targeting.  The most successful attempts
toward this goal in Drosophila have utilized the P element transposition system.  However,
directed introduction of DSBs is still neither highly precise nor efficient, probably in part
owing to the innate properties of the P element transposase, which although being a
site-specific DNA binding protein, also has an affinity for nonspecific DNA sequences in
vitro.  As a result, DSBs generated by P element transposase are distributed randomly in the
Drosophila genome with the highest frequency close to or at the P element ends.  Site-specific
endonucleases with sufficiently long recognition sequences potentially could provide a
solution to this problem.  Among the most specific is the I-SceI endonuclease.  It recognizes
an 18-bp nonpalindromic sequence and has very low tolerance to nucleotide substitution.
Theoretically, this recognition site should appear only once in every 6.87 x 10^10 bp, which
exceeds the size of the Drosophila genome by about 400 times.

<>

<1>Mohan, K.N., Chaillet, J.R.
<2>Cell and Molecular Biology of DNA Methyltransferase 1.
<3>Int. Rev. Cell Mol. Biol.
<4>306
<5>1-42
<6>2013
<7>The DNA cytosine methyltransferase 1 (DNMT1) is a ubiquitous nuclear enzyme that catalyzes the
well-established reaction of placing methyl
groups on the unmethylated cytosines in methyl-CpG:CpG base pairs in
the hemimethylated DNA formed by methylated parent and unmethylated
daughter strands. This activity regenerates fully methylated
methyl-CpG:methyl-CpG pairs. Despite the straightforward nature of its
catalytic activity, detailed biochemical, genetic, and developmental
studies revealed intricate details of the central regulatory role of
DNMT1 in governing the epigenetic makeup of the nuclear genome. DNMT1
mediates demethylation and also participates in seemingly wide cellular
functions unrelated to maintenance DNA methylation. This review brings
together mechanistic details of maintenance methylation by DNMT1, its
regulation at transcriptional and posttranscriptional levels, and the
seemingly unexpected functions of DNMT1 in the context of DNA
methylation which is central to epigenetic changes that occur during
development and the process of cell differentiation.

<>

<1>Mohn, W.W., Teather, R.M.
<2>Partial purification and characterisation of Bfi57I and Bfi89I, restriction endonucleases from different strains of Butyrivibrio fibrisolvens.
<3>Gene
<4>155
<5>131-132
<6>1995
<7>Two class-II restriction endonucleases (ENases), Bfi57I and Bfi89I, were partially purified
from Butyrivibrio fibrisolvens OB157 and OB189, respectively. Bfi57I (isoschizomer Sau3AI) had
the DNA recognition/cleavage sequence 5'-/GATC-3'; it is not inhibited by Dam methylation,
but is partially inhibited by M.BamHI methylation. Bfi89I (isoschizomer EaeI) had the
recognition/cleavage sequence 5'Y/GGCCR-3'; unlike the EaeI isoschizomer it is not fully
inhibited by M.HaeIII methylation.

<>

<1>Mohr, G., Smith, D., Belfort, M., Lambowitz, A.M.
<2>Rules for DNA target-site recognition by a lactococcal group II intron enable retargeting of the intron to specific DNA sequences.
<3>Genes Dev.
<4>14
<5>559-573
<6>2000
<7>Group II intron homing occurs primarily by a mechanism in which the intron RNA reverse splices
into a DNA target site and is then reverse transcribed by the intron-encoded protein. The DNA
target site is recognized by an RNP complex containing the intron-encoded protein and the
excised intron RNA. Here, we analyzed DNA target-site requirements for the Lactococcus lactis
Ll.LtrB group II intron in vitro and in vivo. Our results suggest a model similar to yeast
mtDNA introns, in which the intron-encoded protein first recognizes a small number of
nucleotide residues in double-stranded DNA and causes DNA unwinding, enabling the intron RNA
to base-pair with the DNA for reverse splicing. Antisense-strand cleavage requires additional
interactions between the protein and 3' exon. Key nucleotide residues are recognized directly
by the intron-encoded protein independent of sequence context, and there is a stringent
requirement for fixed spacing between target site elements recognized by the protein and RNA
components of the endonuclease. Experiments with DNA substrates containing GC-clamps or
"bubbles" indicate a requirement for DNA unwinding in the 3' exon but not the distal 5' exon
region. Finally, by applying the target-site recognition rules, we show that the L1.LtrB
intron can be modified to insert at new sites in a plasmid-borne thyA gene in Escherichia
coli. This strategy should be generally applicable to retargeting group II introns and to
delivering foreign sequences to specific sites in heterologous genomes.

<>

<1>Moineau, S., D'Amelio, G.E., Pandian, S., Klaenhammer, T.R.
<2>The susceptibility of evolving dairy bacteriophages to restriction enzymes and lactococcal R/M system.
<3>J. Dairy Sci.
<4>75
<5>114
<6>1992
<7>One approach to limit phage development during milk fermentation is to use strains insensitive
to predominant phage species present in cheese plants, particularly prolate (c2) and small
isometric-headed (P008) species. However this may lead to the emergence of other phage
species. Recently, we have isolated 7 industrial phages (phi48, phi50,al,bl, cs, d1 from USA
and UL36 from Canada) able to propagate on Lactococcus lactis LMA12, and except for UL36, also
on its pTR2030 transconjugants. Electron microscopy and DNA homology studies have placed these
phages within the P335 species (composed of lytic and temperate types). Molecular analyses
have shown a relatively high number of restriction sites in their genomes for many
endonucleases, including ScrFI. The industrial phages, compared to phages sk1, p2, jj50 (P008
species) and c2, were highly sensitive to 4 plasmid-encoded R/M systems (pTN20, pTRK12,
pTRK30, pTRK68). The paucity of restriction sites in many phage genomes has been proposed as
an antirestriction response which has evolved in some lactococcal phages. Since the 7 phages
studied herein are more sensitive to R/M and their DNA cuts more frequently, this group may
represent a younger generation of phages that have just recently evolved to lactococci.

<>

<1>Moineau, S., Pandian, S., Klaenhammer, T.R.
<2>Restriction/modification systems and restriction endonucleases are more effective on lactococcal bacteriophages that have emerged recently in the dairy industry.
<3>Appl. Environ. Microbiol.
<4>59
<5>197-202
<6>1993
<7>Recently, eight lytic small isometric-headed bacteriophages were isolated from
cheese-manufacturing plants throughout North America. The eight phages were different, but all
propagated on one strain, Lactococcus lactis NCK203. On the basis of DNA homology, they were
classified in the P335 species. Digestion of their genomes in vitro with restriction enzymes
resulted in an unusually high number of type II endonuclease sites compared with the more
common lytic phages of the 936 (small isometric-headed) and c2 (prolate-headed) species. In
vivo, the P335 phages were more sensitive to four distinct lactococcal restriction and
modification (R/M) systems than phages belonging to the 936 and c2 species. A significant
correlation was found between the number of restriction sites for endonucleases (purified from
other bacterial genera) and the relative susceptibility of phages to lactococcal R/M systems.
Comparisons among these three phage species indicate that the P335 species may have emerged
most recently in the dairy industry.

<>

<1>Moineau, S., Walker, S.A., Holler, B.J., Vedamuthu, E.R., Vandenbergh, P.A.
<2>Expression of a Lactococcus lactis phage resistance mechanism by Streptococcus thermophilus.
<3>Appl. Environ. Microbiol.
<4>61
<5>2461-2466
<6>1995
<7>The 7.8-kb lactococcal plasmid pSRQ700 encodes the LlaII restriction/modification system which
recognizes and cleaves the sequence 3'-GATC-5'. When the plasmid pSRQ700 is introduced into
a phage-sensitive Lactococcus lactis strain, strong phage resistance is conferred by the LlaII
system. In this report, we show that pSRQ700 cannot replicate in Streptococcus thermophilus.
However, if cloned into the vector pNZ123, the native LlaII system is expressed and strong
phage resistance is conferred to various industrial S. thermophilus strains. Resistance
against phages isolated from yogurt and mozzarella wheys was observed. To our knowledge, this
is the first report of increased phage resistance in S. thermophilus.

<>

<1>Moineau, S., Walker, S.A., Vedamuthu, E.R., Vandenbergh, P.A.
<2>Isolated DNA encoding enzyme for phage resistance.
<3>International Patent Office
<4>WO 9621017
<5>
<6>1996
<7>An isolated DNA of a Lactococcus lactis showing a SEQ ID NO: 1 encoding a restriction and two
modification enzymes (R/M SEQ ID NO:2, 3 and 4).  The isolated DNA is used to transform
sensitive dairy cultures, such as Lactococcus lactis and Streptococcus thermophilus, to
provide phage resistance.  Escherichia coli can be used to produce endonucleases.

<>

<1>Moineau, S., Walker, S.A., Vedamuthu, E.R., Vandenbergh, P.A.
<2>Cloning and sequencing of LlaII restriction/modification genes from Lactococcus lactis and relatedness of this system to the Streptococcus pneumoniae DpnII system.
<3>Appl. Environ. Microbiol.
<4>61
<5>2193-2202
<6>1995
<7>The natural 7.8-kb plasmid pSRQ700 was isolated from Lactococcus lactis subsp. cremoris DCH-4.
It encodes a restriction/modification system named LlaII. When introduced into a
phage-sensitive L. lactis strain, pSRQ700 confers strong phage resistance against the three
most common lactococcal phage species, namely, 936, c2, and P335. The LlaII endonuclease was
purified and found to cleave the palindromic sequence 5'-GATC-3'. It is an isoschizomer of
Streptococcus pneumoniae DpnII. The plasmid pSRQ700 was mapped, and the genetic organization
of LlaII was localized. Cloning and sequencing of the entire LlaII system allowed the
identification of three open reading frames. The three genes (llaIIA, llaIIB, and llaIIC)
overlapped and are under one putative promoter. A putative terminator was found at the end of
llaIIC. The genes llaIIA and llaIIB coded for m6A methyltransferases, and llaIIC coded for an
endonuclease. The LlaII system shares strong genetic similarities with the DpnII system. The
deduced amino acid sequence of M.LlaIIA was 75% identical with that of M.DpnII, whereas
M.LlaIIB was 88% identical with M.DpnA. However, R.LlaII shared only 31% identity with
R.DpnII.

<>

<1>Moineau, S., Walker, S.A., Vedamuthu, E.R., Vandenburgh, P.A.
<2>Isolated DNA encoding enzyme for phage resistance.
<3>US Patent Office
<4>US 5824523
<5>
<6>1998
<7>An isolated DNA of a Lactococcus lactis showing a SEQ ID No: 1 encoding a restriction and two
modification enzymes (R/M SEQ ID NO: 2, 3, and 4).  The isolated DNA is used to transform
sensitive dairy cultures, such as Lactococcus lactis and Streptococcus thermophilus, to
provide phage resistance.  Escherichia coli can be used to produce endonucleases.

<>

<1>Moissidou, A., Rina, M., Bouriotis, V.
<2>Isolation and identification of restriction BshKI.
<3>Nucleic Acids Res.
<4>17
<5>8884
<6>1989
<7>None

<>

<1>Mok, Y.K., Clark, D.R., Kam, K.M., Shaw, P.C.
<2>BsiYI, a novel thermophilic restriction endonuclease that recognizes 5' CCNNNNNNNGG3' and the discovery of a wrongly sequenced site in pACYC177 .
<3>Nucleic Acids Res.
<4>19
<5>2321-2323
<6>1991
<7>A new type II restriction endonuclease designated BsiYI has been purified from
a thermophilic soil Bacillus stearothermophilus strain.  This enzyme recognizes
and cleaves the highly degenerate sequence 5'CCNNNNN^NNGG3'.  During the
identification of the recognition sequence of BsiYI, we discovered that there
should be five G nucleotides instead of four at position 1227 - 1230 of the
plasmid pACYC177.

<>

<1>Mok, Y.K., Clark, D.R., Kam, K.M., Shaw, P.C.
<2>Restriction endonuclease from thermophilic bacterial species I.  Isolation and characterization of BsiEI.
<3>Nucleic Acids Res.
<4>18
<5>4954
<6>1990
<7>
<>

<1>Mok, Y.K., Clark, D.R., Kam, K.M., Shaw, P.C.
<2>Restriction endonuclease from thermophilic bacterial species II. Isolation and characterization of BsiBl.
<3>Nucleic Acids Res.
<4>18
<5>6740
<6>1990
<7>None

<>

<1>Mokrishcheva, M.L., Solonin, A.S., Nikitin, D.V.
<2>Fused eco29kIR- and M genes coding for a fully functional hybrid polypeptide as a model of molecular evolution of restriction-modification systems.
<3>BMC Evol. Biol.
<4>11
<5>35
<6>2011
<7>Background: The discovery of restriction endonucleases and modification DNA
methyltransferases, key instruments of genetic engineering, opened
a new era of molecular biology through development of the recombinant
DNA technology. Today, the number of potential proteins assigned to
type II restriction enzymes alone is beyond 6000, which probably
reflects the high diversity of evolutionary pathways. Here we present
experimental evidence that a new type IIC restriction and modification
enzymes carrying both activities in a single polypeptide could result
from fusion of the appropriate genes from preexisting bipartite
restriction-modification systems.
Results: Fusion of eco29kIR and M ORFs gave a novel gene encoding
for a fully functional hybrid polypeptide that carried both restriction
endonuclease and DNA methyltransferase activities. It has been placed
into a subclass of type II restriction and modification enzymes - type
IIC. Its MTase activity, 80% that of the M.Eco29kI enzyme, remained
almost unchanged, while its REase activity decreased by three times,
concurrently with changed reaction optima, which presumably can be
caused by increased steric hindrance in interaction with the substrate.
In vitro the enzyme preferentially cuts DNA, with only a low level of
DNA modification detected. In vivo new RMS can provide a 10(2)-fold
less protection of host cells against phage invasion.
Conclusions: We propose a molecular mechanism of appearing of type
IIC restriction-modification and M.SsoII-related enzymes, as well as
other multifunctional proteins. As shown, gene fusion could play an
important role in evolution of restriction-modification systems and be
responsible for the enzyme subclass interconversion. Based on the
proposed approach, hundreds of new type IIC enzymes can be generated
using head-to-tail oriented type I, II, and III restriction and
modification genes. These bifunctional polypeptides can serve a basis
for enzymes with altered recognition specificities. Lastly, this study
demonstrates that protein fusion may change biochemical properties of
the involved enzymes, thus giving a starting point for their further
evolutionary divergence.

<>

<1>Mol, C.D., Arvai, A.S., Begley, T.J., Cunningham, R.P., Tainer, J.A.
<2>Structure and activity of a thermostable thymine-DNA glycosylase: Evidence for base twisting to remove mismatched normal DNA bases.
<3>J. Mol. Biol.
<4>315
<5>373-384
<6>2002
<7>The repair of T:G mismatches in DNA is key for maintaining bacterial restriction/modification
systems and gene silencing in higher eukaryotes. T:G mismatch repair can be initiated by a
specific mismatch glycosylase (MIG) that is homologous to the helix-hairpin-helix (HhH) DNA
repair enzymes. Here, we present a 2.0 A resolution crystal structure and complementary
mutagenesis results for this thermophilic HhH MIG enzyme. The results suggest that MIG
distorts the target thymine nucleotide by twisting the thymine base approximately 90 degrees
away from its normal anti position within DNA. We propose that functionally significant
differences exist in DNA repair enzyme extrahelical nucleotide binding and catalysis that are
characteristic of whether the target base is damaged or is a normal base within a mispair.
These results explain why pure HhH DNA glycosylases and combined glycosylase/AP lyases cannot
be interconverted by simply altering their functional group chemistry, and how
broad-specificity DNA glycosylase enzymes may weaken the glycosylic linkage to allow a variety
of damaged DNA bases to be excised.

<>

<1>Molemans, F., van Emmelo, J., Fiers, W.
<2>The sequence specificity of endonucleases CauI and CauII isolated from Chloroflexus aurantiacus.
<3>Gene
<4>18
<5>93-96
<6>1982
<7>The type II restriction enzymes CauI and CauII, isolated from Chloroflexus
aurantiacus, recognize and cleave (at the position indicated by an arrow) the
sequences G^GA/TCC and CC^G/CGG, respectively.  These conclusions are supported
by the results from restriction site mapping, sequence analysis by partial
chemical degradation, end-group analysis after lambda exonuclease treatment and
computer-assisted comparison of DNA sequence data.

<>

<1>Molholt, B., Fraser, D.
<2>Host-controlled restriction of T-even bacteriophages:  relation of endonuclease I and T-even-induced nucleases to restriction.
<3>J. Virol.
<4>2
<5>313-319
<6>1968
<7>Restriction of nonglucosylated T2 phage (T*2) as a function of bacterial growth
state was the same for endonuclease I-containing and endonuclease I-deficient
strains of Escherichia coli B.  Furthermore, E. coli strains with various
levels of restriction for T2 had comparable endonuclease I activities.  It was
also found that a T4 mutant temperature-sensitive for gene 46 and 47 functions
was fully restricted at 42 C.  It therefore appears that neither endonuclease I
nor the phage-induced nucleases whose activities are blocked by mutations in
genes 46 and 47 catalyze the initial event in restriction of nonglucosylated
T-even phages.

<>

<1>Molina, L., Bernal, P., Udaondo, Z., Segura, A., Ramos, J.L.
<2>Complete Genome Sequence of a Pseudomonas putida Clinical Isolate, Strain H8234.
<3>Genome Announcements
<4>1
<5>e00496-13
<6>2013
<7>We report the complete genome sequence of Pseudomonas putida strain H8234, which  was isolated
from a hospital patient presenting with bacteremia. This strain has
a single chromosome (6,870,827 bp) that contains 6,305 open reading frames. The
strain is not a pathogen but exhibits multidrug resistance associated with 40
genomic islands.

<>

<1>Molina, R., Redondo, P., Stella, S., Marenchino, M., D'Abramo, M., Gervasio, F.L., Charles, E.J., Valton, J., Grizot, S., Duchateau, P., Prieto, J., Montoya, G.
<2>Non-specific protein-DNA interactions control I-CreI target binding and cleavage.
<3>Nucleic Acids Res.
<4>40
<5>6936-6945
<6>2012
<7>Homing endonucleases represent protein scaffolds that provide powerful tools for  genome
manipulation, as these enzymes possess a very low frequency of DNA
cleavage in eukaryotic genomes due to their high specificity. The basis of
protein-DNA recognition must be understood to generate tailored enzymes that
target the DNA at sites of interest. Protein-DNA interaction engineering of
homing endonucleases has demonstrated the potential of these approaches to create
new specific instruments to target genes for inactivation or repair. Protein-DNA
interface studies have been focused mostly on specific contacts between amino
acid side chains and bases to redesign the binding interface. However, it has
been shown that 4 bp in the central DNA sequence of the 22-bp substrate of a
homing endonuclease (I-CreI), which do not show specific protein-DNA
interactions, is not devoid of content information. Here, we analyze the
mechanism of target discrimination in this substrate region by the I-CreI
protein, determining how it can occur independently of the specific protein-DNA
interactions. Our data suggest the important role of indirect readout in this
substrate region, opening the possibility for a fully rational search of new
target sequences, thus improving the development of redesigned enzymes for
therapeutic and biotechnological applications.

<>

<1>Mollet, B., Zwahlen, M., Paul, T., Schell, M.A., Pridomore, R.D., Arigoni, F., Delley, M.
<2>NCC2705 - The genome of a Bifidobacterium.
<3>Japanese Patent Office
<4>JP 2004531245 A
<5>
<6>2004
<7>
<>

<1>Mollmann, S., Albersmeier, A., Ruckert, C., Tauch, A.
<2>Complete Genome Sequence of Corynebacterium imitans DSM 44264, Isolated from a Five-Month-Old Boy with Suspected Pharyngeal Diphtheria.
<3>Genome Announcements
<4>2
<5>e01210-14
<6>2014
<7>The complete genome sequence of the type strain Corynebacterium imitans DSM 44264 comprises
2,565,321 bp with a mean G+C content of 64.26%. The detection of the
antibiotic resistance genes erm(X), aphA1-IAB, strA-strB, and cmx is fully
consistent with the previously observed multidrug-resistant pattern of C. imitans
isolates.

<>

<1>Molloy, P.L., Symons, R.H.
<2>Cleavage of DNA.RNA hybrids by Type II restriction enzymes.
<3>Nucleic Acids Res.
<4>8
<5>2939-2946
<6>1980
<7>The action of a number of restriction enzymes on DNA.RNA hybrids has been examined using
hybrids synthesised with RNAs of cucumber mosaic virus as templates. The enzymes EcoRI,
HindII, SalI, MspI, HhaI, AluI, TaqI and HaeIII cleaved the DNA strand of the hybrids (and
possibly also the RNA strand) into specific fragments. For four of these enzymes, HhaI, AluI,
TaqI and HaeIII, comparison of the restriction fragments produced with the known sequences of
the viral RNAs confirmed that they were recognising and cleaving the DNA strand of the hybrids
at their correct recognition sequences. It is likely that the ability to utilise DNA.RNA
hybrids as substrates is a general property of Type II restriction enzymes.

<>

<1>Molloy, P.L., Watt, F.
<2>Effect of cytosine methylation on cutting by the restriction enzyme MaeII.
<3>Nucleic Acids Res.
<4>16
<5>2335
<6>1988
<7>The restriction enzyme MaeII, isolated from Methanococcus aeolicus, recognizes
and cuts within the sequence ACGT.  The susceptibility of MaeII to inhibition
by cytosine methylation of interest for studies of gene expression as its
recognition sequence contains a CG dinucleotide which has the potential to be
methylated in vertebrate DNA.  In particular its recognition sequence is found
within the binding sites for at least three mammalian transcription factors -
the adenovirus major late gene upstream element, CCACGTGA, the cAMP responsive
element, TGACGTCA, and an adenovirus E2 factor binding site, (T/A)CGTCA.

<>

<1>Molnar, A., Geck, P., Orosz, A., Kulcsar, P., Nasz, I.
<2>Purification of a new restriction endonuclese from Streptococcus mutans and identification of its recognition sequence.
<3>Acta Microbiol. Hung.
<4>38
<5>55-60
<6>1991
<7>SmuEI, a type II restriction endonuclease, has been isolated from Streptococcus mutans
serotype E, which is an isoschizomer of AvaII recognizes the palindromic pentanucleotide
sequence 5' GGWCC 3'. Similarly to AvaII, SmuEI cleaves the sequence, G^GWCC generating 5'
protruding fragment termini.

<>

<1>Molnarova, V., Javorsky, P., Pristas, P.
<2>Occurrence of restriction and modification systems in ruminal Selenomonades.
<3>Chemical Papers-Chemicke Zvesti
<4>52
<5>284
<6>1998
<7>Bacterial restriction and modification systems consist of a restriction endonuclease plus a
"cognate" modification methyltransferase having the same specificity. The biological function
of restriction and modification systems is to protect bacteria from invading phage and plasmid
DNA. The rumen is one of the most complex and best studied of all microbial ecosystems. In
addition to protozoa, bacteria and fungi the rumen is known to contain a large and diverse
population of bacteriophages. These phages probably play a significant role in the population
dynamics of ruminal bacteria. Thus under ruminal condition possession of restriction and
modification system can provide bacteria with a selective advantage.

<>

<1>Molnarova, V., Pristas, P., Javorsky, P.
<2>Prevalence of CTGCAG recognizing restriction and modification systems in ruminal selenomonades.
<3>Anaerobe
<4>5
<5>37-41
<6>1999
<7>Analysis of restriction and modification activities in natural population of Selenomonas
ruminantium have revealed the prevalence of CTGCAG (PstI isoschizomers) recognizing
restriction and/or modification systems in these bacteria.  PstI isoschizomeric restriction
endonucleases were detected in 4 out of 15 strains tested.  In one strain, the PstI
isoschizomeric restriction system was accompanied by another restriction and modification
system recognizing the sequence GAATTC (EcoRI isoschizomer).  Four other strains contained
CTGCAG specific methylases which lacked cognate endonuclease activities.  The presence of
identical restriction and modification systems in both subspecies of S. ruminantium, as well
as the occurrence of PstI isoschizomers in various combinations, indicate the possibility of
horizontal transfer of genes coding for these systems.

<>

<1>Momparler, R.L., Bovenzi, V.
<2>DNA methylation and cancer.
<3>J. Cell. Physiol.
<4>183
<5>145-154
<6>2000
<7>The methylation of DNA is an epigenetic modification that can play an important role in the
control of gene expression in mammalian cells. The enzyme involved in this process is DNA
methyltransferase, which catalyzes the transfer of a methyl group from S-adenosyl-methionine
to cytosine residues to form 5-methylcytosine, a modified base that is found mostly at CpG
sites in the genome. The presence of methylated CpG islands in the promoter region of genes
can suppress their expression. This process may be due to the presence of 5-methylcytosine
that apparently interferes with the binding of transcription factors or other DNA-binding
proteins to block transcription. In different types of tumors, aberrant or accidental
methylation of CpG islands in the promoter region has been observed for many cancer-related
genes resulting in the silencing of their expression. How this aberrant hypermethylation takes
place is not known. The genes involved include tumor suppressor genes, genes that suppress
metastasis and angiogenesis, and genes that repair DNA suggesting that epigenetics plays an
important role in tumorigenesis. The potent and specific inhibitor of DNA methylation,
5-aza-2'-deoxycytidine (5-AZA-CdR) has been demonstrated to reactivate the expression most of
these "malignancy" suppressor genes in human tumor cell lines. These genes may be interesting
targets for chemotherapy with inhibitors of DNA methylation in patients with cancer and this
may help clarify the importance of this epigenetic mechanism in tumorigenesis.

<>

<1>Momynaliev, K., Chelysheva, V., Selezneva, O., Akopian, T., Alexeev, D., Govorun, V.
<2>Complete Genome Sequences of Helicobacter pylori Rifampin-Resistant Strains.
<3>Genome Announcements
<4>1
<5>e00446-13
<6>2013
<7>Here we present the complete genome sequences of two Helicobacter pylori rifampin-resistant
(Rif(r)) strains (Rif1 and Rif2). Rif(r) strains were obtained
by in vitro selection of H. pylori 26695 on agar plates with 20 microg/ml
rifampin. The genome data provide insights on the genomic diversity of H. pylori
under selection by rifampin.

<>

<1>Momynaliev, K.T., Rogov, S.I., Govorun, V.M.
<2>Nucleotide correspondence between protein-coding sequences of Helicobacter pylori 26695 and J99 strains.
<3>Mol. Biol. (Mosk)
<4>39
<5>945-951
<6>2005
<7>Comparison of open-reading frames (ORFs) H. pylori 26695 and J99 strains has been revealed
prevalence of nucleotide replacements as transitions (more than 3%) above transversions (less
than 1%). Prevalence of nucleotide transitions is caused by high speed of C : G to T : A
transitions in a coding strand of DNA (3.5-5.3%) and not coding strand (2.9-3.9%). The
correspondence rate of transversion (A --> C, A --> T, C --> A, C --> G, G --> C, G --> T, T
--> A and T --> G) did not exceed 0.84%. The highest correspondence frequency between C and T
was detected in ACGT-ATGT (28.3%) - the site of methylation by active methyltransferase
M.Hpy99XI in H. pylori 26695 and J99. Thus one can speculate that predominant transition
taking place in H. pylori is mutation of C into T, which is realized through cytosine
methylation-deamination mechanism.

<>

<1>Momynaliev, K.T., Smirnova, O.V., Kudryavtseva, L.V., Govorun, V.M.
<2>Comparative genome analysis of Helicobacter pylori strains.
<3>Mol. Biol. (Mosk)
<4>37
<5>529-536
<6>2003
<7>DNA macroarrays were used to characterize 17 Helicobacter pylori strains isolated in four
geographic regions of Russia (Moscow, St.
Petersburg, Kazan, and Novosibirsk). Of all genes, 1272 (81%) proved to
occur in all strains and to constitute a functional core of the genome,
and 293 (18.7%) were strain-specific and greatly varied among the H.
pylori strains. Most (71%) of the latter had unknown functions; the
remainder included restriction-modification genes (3-9%), transposition
genes (2-4%), and genes coding for outer membrane proteins (2-4%). The
Russian H. pylori strains did not differ in genome organization or in
the number and distribution of strain-specific genes from strains
isolated in other countries.

<>

<1>Monaci, E., Masignani, V., Pizza, M., Fontana, M.R.
<2>Gonococcal proteins and nucleic acids journal.
<3>Japanese Patent Office
<4>JP 2004537977
<5>
<6>2004
<7>
<>

<1>Monastiriakos, S.K., Doiron, K.M., Siponen, M.I., Cupples, C.G.
<2>Functional interactions between the MutL and Vsr proteins of Escherichia coli are dependent on the N-terminus of Vsr.
<3>DNA Repair
<4>3
<5>639-647
<6>2004
<7>The crystal structure of the Escherichia coli Vsr endonuclease bound to a C(T/G)AGG substrate
revealed that the DNA is held by a pincer composed of
a trio of aromatic residues which intercalate into the major groove, and
an N-terminus alpha helix which lies across the minor groove. We have
constructed an N-terminus truncation (Delta14) which removes most of the
alpha helix. The mutant is still fairly proficient in mediating very short
patch repair. However, its endonuclease activity is considerably reduced
and, in contrast to that of the wild type protein, cannot be stimulated by
MutL. We had shown previously that excess Vsr in vivo causes mutagenesis,
probably by inhibiting the participation of MutL in mismatch repair. The
Delta14 mutant has diminished mutagenicity. In contrast, four
enzymatically inactive mutants, with intact N-termini, are as mutagenic as
the wild type protein. On the basis of these results we suggest that MutL
causes a conformational change in the N-terminus of Vsr which enhances Vsr
activity, and that this functional interaction between Vsr and MutL
decreases the ability of MutL to carry out mismatch repair.

<>

<1>Monastyrskaya, G.S., Fushan, A.A., Abaev, I.V., Filyukova, O.B., Kostina, M.B., Pecherskih, E., Sverdlov, E.D.
<2>Genome-wide comparison reveals great inter- and intraspecies variability in B. pseudomallei and B. mallei pathogens.
<3>Res. Microbiol.
<4>155
<5>781-793
<6>2004
<7>Burkholderia mallei and B. pseudomallei, closely related Gram-negative
bacteria, are causative agents of serious infectious diseases of humans
and animals: glanders and melioidosis, respectively. Despite numerous
studies of these pathogens, the detailed mechanism of their pathogenesis
is still unknown. The problem is even more complicated due to natural
variability of B. pseudomallei and B. mallei strains, the understanding of
which is a prerequisite for rational design of tools for diagnostics,
prophylaxis and therapy of the diseases. Using a subtractive hybridization
technique, we compared the genomes of B. pseudomallei C-141 and B. mallei
C-5 strains. A subtracted library of DNA fragments specific for B.
pseudomallei C-141 and absent from B. mallei C-5 was obtained and
analyzed. A variety of differences have been detected and mapped on the
recently sequenced genome of B. pseudomallei K96243. A comparative
sequence analysis also revealed considerable genomic differences between
B. pseudomallei C-141 and B. mallei ATCC 23344 strains sequenced at The
Institute for Genomic Research (TIGR). We also observed significant
genomic differences between B. pseudomallei C-141 and B. pseudomallei
K96243. Some of the differential DNA fragments displayed similarity to
different mobile elements which have not yet been described for B.
pseudomallei, whereas the others matched various prophage components,
components of active transport systems, different enzymes and
transcription regulators. A substantial proportion of the differential
clones had no database matches either at the nucleotide or protein level.
The results provide evidence for great genome-wide variability of B.
pseudomallei, further confirmed by Southern blot analysis of various B.
pseudomallei strains. The data obtained can be useful for future
development of efficient diagnostic tools allowing rapid identification of
species, strains and isolates of B. mallei and B. pseudomallei.

<>

<1>Moncke-Buchner, E., Rothenberg, M., Reich, S., Wagenfuhr, K., Matsumura, H., Terauchi, R., Kruger, D.H., Reuter, M.
<2>Functional Characterization and Modulation of the DNA Cleavage Efficiency of Type III Restriction Endonuclease EcoP15I in Its Interaction with Two  Sites in the DNA Target.
<3>J. Mol. Biol.
<4>387
<5>1309-1319
<6>2009
<7>EcoP15I is a Type III restriction endonuclease requiring the interaction with two inversely
oriented 5'-CAGCAG recognition sites for efficient DNA
cleavage. Diverse models have been developed to explain how enzyme
complexes bound to both sites move toward each other, DNA translocation,
DNA looping and simple diffusion along the DNA. Conflicting data also
exist about the impact of cofactor S-adenosyl-l-methionine (AdoMet), the
AdoMet analogue sinefungin and the bases flanking the DNA recognition
sequence on EcoP15I enzyme activity. To clarify the functional role of
these questionable parameters on EcoP15I activity and to optimize the
enzymatic reaction, we investigated the influence of cofactors, ionic
conditions, bases flanking the recognition sequence and enzyme
concentration. We found that AdoMet is not necessary for DNA cleavage.
Moreover, the presence of AdoMet dramatically impaired DNA cleavage due to
competing DNA methylation. Sinefungin neither had an appreciable effect on
DNA cleavage by EcoP15I nor compensated for the second recognition site.
Moreover, we discovered that adenine stretches on the 5' or 3' side of
CAGCAG led to preferred cleavage of this site. The length of the adenine
stretch was pivotal and had to be different on the two sides for most
efficient cleavage. In the absence of AdoMet and with enzyme in molar
excess over recognition sites, we observed minor cleavage at two
communicating DNA sites simultaneously. These results could also be
exploited in the high-throughput, quantitative transcriptome analysis
method SuperSAGE to optimize the crucial EcoP15I digestion step.

<>

<1>Mondy, S., Lalouche, O., Dessaux, Y., Faure, D.
<2>Genome Sequence of the Quorum-Sensing-Signal-Producing Nonpathogen Agrobacterium  tumefaciens Strain P4.
<3>Genome Announcements
<4>1
<5>e00798-13
<6>2013
<7>Agrobacterium tumefaciens P4 is a quorum-sensing-signal-producing bacterium that  has been
isolated from the tobacco rhizosphere. This strain belongs to
genomospecies 1 of the A. tumefaciens complex; it is avirulent on various
putative host plants, devoid of the Ti plasmid, and contains a luxI homolog on
the At plasmid.

<>

<1>Mondy, S., Lalouche, O., Dessaux, Y., Faure, D.
<2>Genome Sequence of the Quorum-Quenching Agrobacterium tumefaciens Strain WRT31.
<3>Genome Announcements
<4>1
<5>e00653-13
<6>2013
<7>Agrobacterium tumefaciens strain WRT31 is a quorum-sensing signal-degrading bacterium that has
been isolated from the rhizosphere of tobacco plants. This
strain belongs to A. tumefaciens genomovar G1, is avirulent on various putative
host plants, devoid of Ti plasmid, and contains the blcC gene encoding a
gamma-butyrolactonase.

<>

<1>Mones, L., Kulhanek, P., Florian, J., Simon, I., Fuxreiter, M.
<2>Probing the two-metal ion mechanism in the restriction endonuclease BamHI.
<3>Biochemistry
<4>46
<5>14514-14523
<6>2007
<7>The choreography of restriction endonuclease catalysis is a long-standing paradigm in
molecular biology. Bivalent metal ions are
required almost for all PD..D/ExK type enzymes, but the number of
cofactors essential for the DNA backbone scission remained ambiguous.
On the basis of crystal structures and biochemical data for various
restriction enzymes, three models have been developed that assign
critical roles for one, two, or three metal ions during the
phosphodiester hydrolysis. To resolve this apparent controversy, we
investigated the mechanism of BamHI catalysis using quantum
mechanical/molecular mechanical simulation techniques and determined
the activation barriers of three possible pathways that involve a
Glu-113 or a neighboring water molecule as a general base or an
external nucleophile that penetrated from bulk solution. The extrinsic
mechanism was found to be the most favorable with an activation free
energy of 23.4 kcal/mol, in reasonable agreement with the experimental
data. On the basis of the effect of the individual metal ions on the
activation barrier, metal ion A was concluded to be pivotal for the
reaction, while the enzyme lacking metal ion B still has moderate
efficiency. Thus, we propose that the catalytic scheme of BamHI does
not involve a general base for nucleophile generation and requires one
obligatory metal ion for catalysis that stabilizes the attacking
nucleophile and coordinates it throughout the nucleophilic attack. Such
a model may also explain the variation in the number of metal ions in
the crystal structures and thus could serve as a framework for a
unified catalytic scheme of type II restriction endonucleases.

<>

<1>Mones, L., Simon, I., Fuxreiter, M.
<2>Metal-binding sites at the active site of restriction endonuclease BamHI can conform to a one-ion mechanism.
<3>Biol. Chem.
<4>388
<5>73-78
<6>2007
<7>The number of metal ions required for phosphoryl transfer in restriction endonucleases is
still an unresolved question in molecular
biology. The two Ca2+ and Mn2+ ions observed in the pre- and
post-reactive complexes of BamHI conform to the classical two-metal ion
choreography. We probed the Mg2+ cofactor positions at the active site
of BamHI by molecular dynamics simulations with one and two metal ions
present and identified several catalytically relevant sites. These can
mark the pathway of a single ion during catalysis, suggesting its
critical role, while a regulatory function is proposed for a possible
second ion.

<>

<1>Mongodin, E.F. et al.
<2>The genome of Salinibacter ruber: Convergence and gene exchange among hyperhalophilic bacteria and archaea.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>18147-18152
<6>2005
<7>Saturated thalassic brines are among the most physically demanding habitats on Earth: few
microbes survive in them. Salinibacter ruber is among these organisms and has been found
repeatedly in significant numbers in climax saltern crystallizer communities. The phenotype of
this bacterium is remarkably similar to that of the hyperhalophilic Archaea (Haloarchaea). The
genome sequence suggests that this resemblance has arisen through convergence at the
physiological level (different genes producing similar overall phenotype) and the molecular
level (independent mutations yielding similar sequences or structures). Several genes and gene
clusters also derive by lateral transfer from (or may have been laterally transferred to)
haloarchaea. S. ruber encodes four rhodopsins. One resembles bacterial proteorhodopsins and
three are of the haloarchaeal type, previously uncharacterized in a bacterial genome. The
impact of these modular adaptive elements on the cell biology and ecology of S. ruber is
substantial, affecting salt adaptation, bioenergetics, and photobiology.

<>

<1>Mongodin, E.F., Shapir, N., Daugherty, S.C., DeBoy, R.T., Emerson, J.B., Shvartsbeyn, A., Radune, D., Vamathevan, J., Riggs, F., Grinberg, V., Khouri, H.M., Wackett, L.P., Nelson, K.E., Sadowsky, M.J.
<2>Secrets of Soil Survival Revealed by the Genome Sequence of Arthrobacter aurescens TC1.
<3>PLoS Genet.
<4>2
<5>e214
<6>2006
<7>Arthrobacter sp. strains are among the most frequently isolated, indigenous, aerobic bacterial
genera found in soils. Member of the genus are metabolically and ecologically diverse and have
the ability to survive in environmentally harsh conditions for extended periods of time. The
genome of Arthrobacter aurescens strain TC1, which was originally isolated from soil at an
atrazine spill site, is composed of a single 4,597,686 basepair (bp) circular chromosome and
two circular plasmids, pTC1 and pTC2, which are 408,237 bp and 300,725 bp, respectively. Over
66% of the 4,702 open reading frames (ORFs) present in the TC1 genome could be assigned a
putative function, and 13.2% (623 genes) appear to be unique to this bacterium, suggesting
niche specialization. The genome of TC1 is most similar to that of Tropheryma, Leifsonia,
Streptomyces, and Corynebacterium glutamicum, and analyses suggest that A. aurescens TC1 has
expanded its metabolic abilities by relying on the duplication of catabolic genes and by
funneling metabolic intermediates generated by plasmid-borne genes to chromosomally encoded
pathways. The data presented here suggest that Arthrobacter's environmental prevalence may be
due to its ability to survive under stressful conditions induced by starvation, ionizing
radiation, oxygen radicals, and toxic chemicals.

<>

<1>Monk, I.R., Foster, T.J.
<2>Genetic manipulation of Staphylococci-breaking through the barrier.
<3>Front Cell Infect Microbiol
<4>2
<5>49
<6>2012
<7>Most strains of Staphylococcus aureus and Staphylococcus epidermidis possess a strong
restriction barrier that hinders exchange of DNA. Recently, major advances have been made in
identifying and characterizing the restriction-modification (RM) systems involved. In
particular a novel type IV restriction enzyme that recognizes cytosine methylated DNA has been
shown to be the major barrier to transfer of plasmid DNA from Escherichia coli into S. aureus
and S. epidermidis. While the conserved type I RM system provides a further barrier. Here we
review the recent advances in understanding of restriction systems in staphylococci and
highlight how this has been exploited to improve our ability to manipulate genetically
previously untransformable strains.

<>

<1>Monk, I.R., Shah, I.M., Xu, M., Tan, M.W., Foster, T.J.
<2>Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically Staphylococcus aureus and Staphylococcus epidermidis.
<3>MBio
<4>3
<5>e00277
<6>2012
<7>The strong restriction barrier present in Staphylococcus aureus and Staphylococcus epidermidis
has limited functional genomic analysis to a
small subset of strains that are amenable to genetic manipulation.
Recently, a conserved type IV restriction system termed SauUSI (which
specifically recognizes cytosine methylated DNA) was identified as the
major barrier to transformation with foreign DNA. Here we have
independently corroborated these findings in a widely used laboratory
strain of S. aureus. Additionally, we have constructed a DNA cytosine
methyltransferase mutant in the high-efficiency Escherichia coli
cloning strain DH10B (called DC10B). Plasmids isolated from DC10B can
be directly transformed into clinical isolates of S. aureus and S.
epidermidis. We also show that the loss of restriction (both type I and
IV) in an S. aureus USA300 strain does not have an impact on virulence.
Circumventing the SauUSI restriction barrier, combined with an improved
deletion and transformation protocol, has allowed the genetic
manipulation of previously untransformable strains of these important
opportunistic pathogens.
IMPORTANCE Staphylococcal infections place a huge burden on the
health care sector due both to their severity and also to the economic
impact of treating the infections because of prolonged hospitalization.
To improve the understanding of Staphylococcus aureus and
Staphylococcus epidermidis infections, we have developed a series of
improved techniques that allow the genetic manipulation of strains that
were previously refractory to transformation. These developments will
speed up the process of mutant construction and increase our
understanding of these species as a whole, rather than just a small
subset of strains that could previously be manipulated.

<>

<1>Monnat, R.J. Jr., Hackmann, A.F.M., Cantrell, M.A.
<2>Generation of highly site-specific DNA double-strand breaks in human cells by the homing endonucleases I-PpoI and I-CreI.
<3>Biochem. Biophys. Res. Commun.
<4>255
<5>88-93
<6>1999
<7>We have determined the ability of two well-characterized eukaryotic homing endonucleases,
I-PpoI from the myxomycete Physarum polycephalum and I-CreI from the green alga Chlamydomonas
reinhardtii, to generate site-specific DNA double-strand breaks in human cells. These 18-kDa
proteins cleave highly conserved 15- or 24-bp rDNA homing sites in their respective hosts to
generate homogeneous 4-base, 3' ends that initiate target intron transposition or "homing."
We show that both endonucleases can be expressed in human cells and can generate site-specific
DNA double-strand breaks in 28S rDNA and homing site plasmids. These endonuclease-induced
breaks can be repaired in vivo, although break repair is mutagenic with the frequent
generation of short deletions or insertions. I-PpoI and I-CreI should be useful for analyzing
DNA double-strand break repair in human cells and rDNA.

<>

<1>Monnerat, M.-P., Thiaucourt, F., Nicolet, J., Frey, J.
<2>Comparative analysis of the lppA locus in Mycoplasma capricolum subsp. capricolum and Mycoplasma capricolum subsp. capripneumoniae.
<3>Vet. Microbiol.
<4>69
<5>157-172
<6>1999
<7>The lppA gene, encoding the lipoprotein named LppA[Mcaca] was characterised in Mycoplasma
capricolum subsp. capricolum.  It encodes a lipoprotein with an apparent molecular mass of 57
kDa as determined by SDS-PAGE. Using antibodies directed against recombinant LppA[Mcaca], we
showed the expression of this lipoprotein in all M. capricolum subsp. capricolum by immunoblot
analysis. The serum did not cross-react with other members of the Mycoplasma mycoides cluster,
hence showing that LppA[Mcaca] was antigenically specific to M. capricolum subsp. capricolum.
The lppA gene was conserved within the subspecies and was used for the development of a
specific PCR assay for the identification of M. capricolum subsp. capricolum.
The taxonomically related Mycoplasma capricolum subsp. capripneumoniae (F38) was found to
contain an lppA-pseudo-gene.  It showed high similarity to functional lppA genes of other
mycoplasmas in the M. mycoides cluster. However, it contained interrupted open reading frames.
Moreover, the nucleotide sequence of the lppA pseudo-genes in different strains of M.
capricolum subsp. capripneumoniae were quite variable. Interestingly, the lppA pseudo-gene had
a size similar to that of the functional lppA genes of other mycoplasmas of the M. mycoides
cluster and occupied the same genomic location as the latter ones in the vicinity of the mtlD
genes. This study showed that all members of the M. mycoides cluster contain each a species-,
subspecies- respectively type- specific lppA gene analogue which encodes a lipoprotein that
has structural and functional relationship to the surface lipoprotein LppA [MmymySC],
previously named P72, of M. mycoides subsp mycoides SC, with the exceptionof M. capricolum
subsp. capripneumoniae which seems not to express an LppA analogue.

<>

<1>Monnerat, M.P., Thiaucourt, F., Poveda, J.B., Nicolet, J., Frey, J.
<2>Genetic and serological analysis of lipoprotein LppA in Mycoplasma mycoides subsp. mycoides LC and Mycoplasma mycoides subsp. capri.
<3>Clin. Diagn. Lab. Immunol.
<4>6
<5>224-230
<6>1999
<7>The genes encoding the 62-kDa lipoproteins from the Mycoplasma mycoides subsp. mycoides
large-colony type (LC) strain Y-goat and the M. mycoides subsp. capri strain PG3 were cloned
and analyzed by sequencing. These two lipoproteins have been named LppA[MmymyLC] and
LppA[Mmyca], and their corresponding genes have been named lppA[MmymyLC] and lppA[Mmyca],
respectively. The nucleotide and deduced amino acid sequences of these two lipoproteins showed
a very high degree of similarity between these two mycoplasmas. Given the sequence data,
LppA seems to fulfill the same structural functions as the previously described major
lipoproteins P72 of M. mycoides subsp. mycoides small-colony type and P67 of the Mycoplasma
species bovine group 7. Based on lppA gene sequences of M. mycoides subsp. mycoides LC and M.
mycoides subsp. capri type strains, a specific PCR assay was developed so that it amplified
this gene in all field strains of the two species analyzed in this study but not in the other
members of the M. mycoides cluster. Analysis of the PCR-amplified lppA genes with frequently
cutting restriction
enzymes showed a certain degree of genetic variability which, however, did not cluster the two
subspecies. This PCR therefore allows a rapid identification of M. mycoides subsp. mycoides LC
and M. mycoides subsp. capri but does not distinguish between these two closely related
subspecies. LppA was expressed in Escherichia coli K-12 and used for the production of
polyclonal mouse antiserum. Antibodies against recombinant LppA[MmymyLC] reacted with a 62 kDa
protein in all M. mycoides subsp. mycoides LC and M. mycoides subsp. capri type strains and
field strains tested but not with the other members of the M. mycoides cluster, thus showing
the antigenic specificity of LppA and further supporting the concept that a close relationship
exists between these two mycoplasmas.

<>

<1>Monnet, C., Loux, V., Bento, P., Gibrat, J.F., Straub, C., Bonnarme, P., Landaud, S., Irlinger, F.
<2>Genome Sequence of Corynebacterium casei UCMA 3821, Isolated from a Smear-Ripened Cheese.
<3>J. Bacteriol.
<4>194
<5>738-739
<6>2012
<7>Corynebacterium casei is one of the most prevalent species present on the surfaces of
smear-ripened cheeses, where it contributes to the production
of the desired organoleptic properties. Here, we report the draft genome
sequence of Corynebacterium casei UCMA 3821 to provide insights into its
physiology.

<>

<1>Monsieurs, P., Mijnendonckx, K., Provoost, A., Venkateswaran, K., Ott, C.M., Leys, N., Van Houdt, R.
<2>Genome Sequences of Cupriavidus metallidurans Strains NA1, NA4, and NE12, Isolated from Space Equipment.
<3>Genome Announcements
<4>2
<5>e00719-14
<6>2014
<7>Cupriavidus metallidurans NA1, NA4, and NE12 were isolated from space and
spacecraft-associated environments. Here, we report their draft genome sequences
with the aim of gaining insight into their potential to adapt to these
environments.

<>

<1>Monsieurs, P., Mijnendonckx, K., Provoost, A., Venkateswaran, K., Ott, C.M., Leys, N., Van Houdt, R.
<2>Draft Genome Sequences of Ralstonia pickettii Strains SSH4 and CW2, Isolated from Space Equipment.
<3>Genome Announcements
<4>2
<5>e00887-14
<6>2014
<7>Ralstonia pickettii SSH4 and CW2 were isolated from space equipment. Here, we report their
draft genome sequences with the aim of gaining insight into their
potential to adapt to these environments.

<>

<1>Monsieurs, P., Provoost, A., Mijnendonckx, K., Leys, N., Gaudreau, C., Van Houdt, R.
<2>Genome Sequence of Cupriavidus metallidurans Strain H1130, Isolated from an Invasive Human Infection.
<3>Genome Announcements
<4>1
<5>e01051-13
<6>2013
<7>Cupriavidus metallidurans H1130 was repeatedly isolated from different blood culture sets
taken from a patient suffering from significant nosocomial
septicemia. Here, we announce the H1130 genome sequence for use in comparative
analyses and for exploring the adaptation and pathogenic potential of this
bacterium.

<>

<1>Monteilhet, C., Dziadkowiec, D., Szczepanek, T., Lazowska, J.
<2>Purification and characterization of the DNA cleavage and recognition site of I-ScaI mitochondrial group I intron encoded endonuclease produced in Escherichia coli.
<3>Nucleic Acids Res.
<4>28
<5>1245-1251
<6>2000
<7>The second intron in the mitochondrial cytb gene of Saccharomyces capensis, belonging to group
I, encodes a 280 amino acid protein containing two LAGLIDADG motifs. Genetic and molecular
studies have previously shown that this protein has a dual function in the wild-type strain.
It acts as a specific homing endonuclease I-ScaI promoting intron mobility and as a maturase
promoting intron splicing. Here we describe the synthesis of a universal code equivalent to
the mitochondrial sequence coding for this protein and the in vitro characterization of I-ScaI
endonuclease activity, using a truncated mutant form of the protein p28bi2 produced in
Escherichia coli. We have also determined the cleavage pattern as well as the recognition site
of p28bi2. It was found that p28bi2 generates a double-strand cleavage downstream from the
intron insertion site with 4 nt long 3'-overhangs. Mutational analysis of the DNA target site
shows that p28bi2 recognizes a 16-19 bp sequence from positions -11 to +8 with respect to the
intron insertion site.

<>

<1>Monteilhet, C., Perrin, A., Thierry, A., Colleaux, L., Dujon, B.
<2>Purification and characterization of the in vitro activity of I-Sce I, a novel and highly specific endonuclease encoded by a group I intron.
<3>Nucleic Acids Res.
<4>18
<5>1407-1413
<6>1990
<7>Group I intron encoded proteins represent a novel class of site specific double
strand endonucleases.  The endonuclease activity of this class of proteins has
been first demonstrated in vivo for I-Sce I which is encoded  by a
mitochondrial intron of Saccharomyces cerevisiae.  Assays using crude cell
extracts have shown that I-Sce I can be used in vitro as a restriction
endonuclease potentially useful for recombinant DNA technology owing to its
large recognition sequence (18 nucleotides).  We report here the purification
and the first detailed analysis of the in vitro activity and properties of
I-Sce I.

<>

<1>Monteiro-Vitorello, C.B. et al.
<2>The genome sequence of the gram-positive sugarcane pathogen Leifsonia xyli subsp. xyli.
<3>Mol. Plant Microbe Interact.
<4>17
<5>827-836
<6>2004
<7>The genome sequence of Leifsonia xyli subsp. xyli, which causes ratoon stunting disease and
affects sugarcane worldwide, was determined. The
single circular chromosome of Leifsonia xyli subsp. xyli CTCB07 was 2.6 Mb
in length with a GC content of 68% and 2,044 predicted open reading
frames. The analysis also revealed 307 predicted pseudogenes, which is
more than any bacterial plant pathogen sequenced to date. Many of these
pseudogenes, if functional, would likely be involved in the degradation of
plant heteropolysaccharides, uptake of free sugars, and synthesis of amino
acids. Although L. xyli subsp. xyli has only been identified colonizing
the xylem vessels of sugarcane, the numbers of predicted regulatory genes
and sugar transporters are similar to those in free-living organisms. Some
of the predicted pathogenicity genes appear to have been acquired by
lateral transfer and include genes for cellulase, pectinase, wilt-inducing
protein, lysozyme, and desaturase. The presence of the latter may
contribute to stunting, since it is likely involved in the synthesis of
abscisic acid, a hormone that arrests growth. Our findings are consistent
with the nutritionally fastidious behavior exhibited by L. xyli subsp.
xyli and suggest an ongoing adaptation to the restricted ecological niche
it inhabits.

<>

<1>Monteiro-Vitorello, C.B., Zerillo, M.M., Van Sluys, M.A., Camargo, L.E., Kitajima, J.P.
<2>Complete Genome Sequence of Leifsonia xyli subsp. cynodontis Strain DSM46306, a Gram-Positive Bacterial Pathogen of Grasses.
<3>Genome Announcements
<4>1
<5>e00915-13
<6>2013
<7>We announce the complete genome sequence of Leifsonia xyli subsp. cynodontis, a vascular
pathogen of Bermuda grass. The species also comprises Leifsonia xyli
subsp. xyli, a sugarcane pathogen. Since these two subspecies have genome
sequences available, a comparative analysis will contribute to our understanding
of the differences in their biology and host specificity.

<>

<1>Montenegro, M.A., Pawlek, B., Behrens, B., Trautner, T.A.
<2>Restriction and modification in Bacillus subtilis:  expression of the cloned methyltransferase gene from B. subtilis phage SPR in E. coli and B. subtilis.
<3>Mol. Gen. Genet.
<4>189
<5>17-20
<6>1983
<7>Expression of the SPR methyltransferase gene from B. subtilis phage SPR cloned
into lambda and SPP1 was studied by analyzing the sensitivity of the hybrid
phage DNAs to restriction by the enzymes HaeIII, MspI, and HpaII.  The
following results were obtained:  (1) The genes were expressed both in the
homologous (B. subtilis) and heterologous (E. coli) host. (2) The specificity
of the expression of the cloned gene was identical to that of the gene in SPR.
(3) Expression depended on the orientation of the cloned segment within the
vector DNAs suggesting that vector promoters were involved in transcription.
The coding strand of the cloned DNA was identified through hybridization with
SPR mRNA.

<>

<1>Montoya, G., Blanco, F., Prieto, J.
<2>Engineering a LAGLIDADG homing endonuclease variant having novel substrate specificity, by mutating an amino acid residue of a parent LAGLIDADG homing endonuclease, and selecting variants having a pattern of cleaved DNA targets - recombinant protein.
<3>International Patent Office
<4>WO 2008102198
<5>
<6>2008
<7>DERWENT ABSTRACT: NOVELTY - Engineering a LAGLIDADG homing endonuclease variant having novel
substrate specificity comprises: (a) the mutation
of at least one amino acid residue of the final C-terminal loop of a
parent LAGLIDADG homing endonuclease, with the exclusion of the
threonine 140 of I-CreI; and (b) the selection and/or screening of the
variants from (a) having a pattern of cleaved DNA targets that is
different from that of the parent LAGLIDADG homing endonuclease.
DETAILED DESCRIPTION - INDEPENDENT CLAIMS are: (1) a homodimeric or
heterodimeric LAGLIDADG homing endonuclease variant, which is
obtainable by the method above, with the exclusion of the homodimeric
variants of SEQ ID NO. 3 and 4, not defined in the specification, and
the homodimeric and heterodimeric variants comprising a monomer of SEQ
ID NO. 5, not defined in the specification; (2) a single-chain chimeric
meganuclease comprising two monomers or core domains of one or two
variants above, or its combination of both;(3) a polynucleotide
fragment encoding one monomer of the variant or the single-chain
meganuclease; (4) a recombinant vector comprising the polynucleotide
fragment; (5) an expression vector comprising two polynucleotide
fragments each encoding one of the two monomers of the heterodimeric
variant, the fragments are operatively linked to regulatory sequences
allowing the production of the two monomers; (6) an expression vector
comprising a polynucleotide fragment encoding the single-chain
meganuclease, the fragment is operatively linked to regulatory
sequences allowing the production of the single-chain meganuclease; (7)
a host cell comprising the polynucleotide fragments or the vector; (8)
a non-human transgenic animal comprising the polynucleotide fragments;
(9) a transgenic plant comprising the polynucleotide fragments; and
(10) a method for decreasing the toxicity of a parent LAGLIDADG homing
endonuclease comprising the mutation of at least one amino acid of the
final C-terminal loop of the parent LAGLIDADG homing endonuclease.
BIOTECHNOLOGY - Preferred Method: In engineering a LAGLIDADG homing
endonuclease variant having novel substrate specificity, the mutations
are in positions of amino acid residues of the final C-terminal loop,
which are contacting the phosphate backbone of the parent LAGLIDADG
homing endonuclease DNA cleavage site. The mutations modify the
interaction between the amino acid residues of the final C-terminal
loop and the phosphate backbone of the parent LAGLIDADG homing
endonuclease DNA cleavage site. The mutations are in positions 138,
139, 142, and/or 143, by reference to I-CreI amino acid sequence
numbering. The residues in positions 138 and/or 139 are substituted by
a hydrophobic amino acid and/or the residues in positions 142 and/or
143 are substituted by a small amino acid. The residue in position 138
is substituted by an alanine, the residue in position 139 is
substituted by a methionine, and/or the residues in positions 142
and/or 143 are substituted by glycines. In the method, step (a)
comprises the mutation of two residues, each one from a different pair
selected from the residues in positions 138 and 139 and the residues in
positions 142 and 143. The parent LAGLIDADG homing endonuclease is a
homodimeric LAGLIDADG homing endonuclease, which is I-CreI. The
homodimeric LAGLIDADG homing endonuclease is an I-CreI variant having
mutations in positions 26-40 and 44-77 of I-CreI and cleaving a
palindromic DNA sequence, where at least the nucleotides in positions
+3 to +5 and +8 to +10 or -10 to -8 and -5 to -3 of one-half of the DNA
sequence correspond to the nucleotides in positions +3 to +5 and +8 to
+10 or -10 to -8 and -5to -3 of one half of a genomic DNA target from a
gene of interest. Step (a) also comprises, simultaneously or
subsequently, the mutation of at least one amino acid residue in a
first functional subdomain corresponding to that situated from
positions 26-40 of I-CreI amino acid sequence, that alter the
specificity towards the nucleotide in positions 8-10 of the DNA target,
and/or the mutation of at least amino acid residue in a second
functional subdomain corresponding to that situated from positions
44-77 of I-CreI amino acid sequence, that alter the specificity towards
the nucleotide in positions + 3 to 5 of the DNA target. Step (a) also
comprises, simultaneously or subsequently, the random mutation of the
whole or the C-terminal half of the LAGLIDADG homing
endonuclease/variant amino acid sequence. In the method, step (b)
comprises the selection and/or screening of the variants from (a),
which are able to cleave at least one DNA target sequence that is not
cleaved by the parent LAGLIDADG homing endonuclease, the DNA target
sequence is derived from the parent LAGLIDADG homing endonuclease
cleavage site, by the replacement of at least one nucleotide of
one-half of the cleavage site, with a different nucleotide. The DNA
target sequence is derived from the I-CreI palindromic site having the
sequence SEQ ID NO. 1, not defined in the specification. The DNA target
has mutations in the nucleotides in positions 1-2, 6-7, 8-10, and/or
11-12. The DNA target sequence is a genomic sequence which is present
in a gene of interest. In decreasing the toxicity of a parent LAGLIDADG
homing endonuclease, the mutation is Lys139Met and/or Thr143Gly.
Preferred Variant: The variant is a heterodimer comprising the monomers
of two different variants obtainable by the method. It is an I-CreI
variant having one or two mutations, each one from a different pair of
mutations selected from the pair S138A and K139M and the pair K142G and
T143G. The variant comprises SEQ ID NO. 6-9, not defined in the
specification. The variant is a heterodimeric I-CreI variant comprising
two monomers, each monomer further comprising different mutations in
positions 26-40 and 44-77 of I-CreI, the variant is able to cleave a
genomic DNA target from a gene of interest. The variant, single-chain
meganuclease, polynucleotides, vector, cell, transgenic plant, or
non-human transgenic mammal are associated with the targeting DNA
construct. Preferred Vector: The vector includes a targeting DNA
construct comprising sequences sharing homologies with the region
surrounding the genomic DNA target sequence. The targeting DNA
construct comprises: (a) sequences sharing homologies with the region
surrounding the genomic DNA target sequence, and (b) sequences to be
introduced flanked by sequence as in (a). ACTIVITY - Virucide. No
biological data given. MECHANISM OF ACTION - Gene Therapy. USE - The
methods are useful for engineering a LAGLIDADG homing endonuclease
variant having novel substrate specificity and for decreasing the
toxicity of a parent LAGLIDADG homing endonuclease. The variant,
single-chain meganuclease, polynucleotide fragments, vector, host cell,
transgenic plant, or non-human transgenic mammal is useful for
molecular biology, for in vivo or in vitro genetic engineering, and for
in vivo or in vitro genome engineering, for non-therapeutic purposes.
It can be used for the preparation of a medicament for preventing,
improving, or curing a genetic disease in an individual, where the
medicament is intended to be administrated by any means to the
individual. It can also be used for the preparation of a medicament for
preventing, improving, or curing a disease caused by an infectious
agent that presents a DNA intermediate, in an individual, where the
infectious agent is a virus. It can also be used for inhibiting the
propagation, inactivating, or deleting an infectious agent that
presents a DNA intermediate, in biological derived products or products
intended for biological uses or for disinfecting an object in vitro.
The variant, single-chain meganuclease, polynucleotide fragments, or
vector can be used as a scaffold for engineering other meganucleases
(all claimed). EXAMPLE - No suitable example given.

<>

<1>Moodley, A., Riley, M.C., Kania, S.A., Guardabassi, L.
<2>Genome Sequence of Staphylococcus pseudintermedius Strain E140, an ST71 European-Associated Methicillin-Resistant Isolate.
<3>Genome Announcements
<4>1
<5>e0020712
<6>2013
<7>We report the first genome sequence of the methicillin-resistant Staphylococcus
pseudintermedius (MRSP) strain E140, isolated from a canine bite wound infection
in Denmark. This strain represents the dominant clonal lineage associated with
canine MRSP infections in Europe.

<>

<1>Moon, B.J., Kim, S.K., Kim, N.H., Kwon, O.S.
<2>Synthesis and characterization of dodecanucleotides containing the XhoI recognition sequence with a phosphorothioate group at the cleavage site.
<3>Bull. Korean Chem. Soc.
<4>17
<5>1031-1036
<6>1996
<7>The synthesis and characterization of diastereomeric dodecanucleotides, d[GATCp(s)TCGAGATC],
containing the recognition sequence of the XhoI restriction endonuclease with a
phosphorothioate internucleotidic linkage at the cleavage sites are described.  Rp and Sp
forms of each diastereomerically pure dinucleoside phosphorothioates d[Cp(S)T] were
presynthesized and used for the addition to the growing oligonucleotide chain as a block.  The
stereochemistry of dinucleoside phosphorothioate was assigned by 31P NMR spectroscopy, enzyme
digestion, and reverse-phase HPLC.  XhoI restriction endonuclease cuts only theRp diastereomer
d[GATCp(s)TCGAGATC].  The rate of hydrolysis is slower than that of the unmodified dodecamer
d[GATCTCGAGATC].  The phosphorothioate nucleotide is used for determination of the
stereochemical course of the XhoI catalyzed reaction.

<>

<1>Moon, B.J., Vipond, I.B., Halford, S.E.
<2>Site-directed mutagenesis of Ile91 of restriction endonuclease EcoRV: Dramatic consequences on the activity and the properties of the enzyme.
<3>J. Biochem. Mol. Biol.
<4>29
<5>17-21
<6>1996
<7>Ile91 of restriction endonuclease EcoRV, which has not been known to take part directly in
catalytic activity, was substituted with Leu by site-directed mutagenesis.  The Ile91Leu
mutant shows over 1000-fold less activity than the wild type EcoRV under standard reaction
condition.  The metal ion dependency of the reaction was altered.  In contrast to the wild
type EcoRV, the mutant prefers Mn2+ to Mg2+ as the cofactor.  In Mn2+ buffer the mutant is as
active as the wild type enzyme in Mg2+ buffer.  Like the wild type enzyme, the mutant shows an
unspecific binding of DNA in gel shift experiments.  In contrast to the wild type enzyme, the
mutant did not cleave at noncognate sites of DNA under star condition.  Note: this paper was
published without Dr. S. Halford's knowledge.

<>

<1>Moon, B.J., Vipond, I.B., Halford, S.E.
<2>Site-directed mutagenesis studies with restriction endonuclease EcoRV to identify the role of Ile91 in recognition and catalysis.
<3>J. Biochem. Mol. Biol.
<4>29
<5>99-104
<6>1996
<7>Site-directed substitutions were made to change the Ile91 of restriction endonuclease EcoRV to
either Val, Ala or Gly to identify the role of Ile91 in recognition and catalysis, since
substitution of Ile91 with Leu afforded dramatic effects on the activity and properties of
restriction endonuclease EcoRV.  These changes alter the size of the hydrophobic side chain at
position 91 and thus might have revealed the reason for the altered phenotype of Ile91 Leu.
However, the properties of Ile91Val and Ile91Ala mutants were much like wild type EcoRV, in
both activity and metal ion preference.  Ile91Gly had very little activity with either Mg2+ or
Mn2- as cofactors.  To try to understand the unusual Mn2+ profile of the Ile91Leu mutant, two
double mutants, Ile91Leu;Asp90Asn and Ile91Leu;Glu45Met were created.  Both double mutants
were seriously disabled by the second amino acid change. Ile91Leu;Glu45Met had some residual
activity in the Mn2+ reaction buffer, whereas the Ile91Leu;Asp90Asn displayed no detectable
activity.

<>

<1>Moon, C.D., Janssen, P.H., Altermann, E.H., Sang, C., Tootill, C.M., Dey, D., Schofield, L.R., Kong, Z., Dong, L., Ronimus, R.S., Kelly, W.J., Leahy, S.C., Attwood, G.T.
<2>Phage phiMRU polynucleotides and polypeptides and uses thereof.
<3>Japanese Patent Office
<4>JP 2010539927 A
<5>
<6>2010
<7>
<>

<1>Moon, W.J., Cho, J.-Y., Chae, Y.K.
<2>Recombinant expression, purification, and characterization of XorKII: A restriction endonuclease from Xanthomonas oryzae pv. oryzae.
<3>Protein Expr. Purif.
<4>62
<5>230-234
<6>2008
<7>An endonuclease from Xanthomonas oryzae pathovar oryzae (Xoo) KACC10331, XorKII, was
recombinantlyproduced in Escherichia coli by applying the stationary state induction method,
which was necessary toprevent the unwanted lysis of E. coli cells. XorKII was purified by
immobilized metal affinity chromatographyon an FPLC system. The yield was 3.5 mg of XorKII per
liter of LB medium. The purified recombinantXorKII showed that it recognized and cleaved to
the same site as PstI. It behaved as a dimer as evidenced
by the size exclusion chromatography. The specific activity of the purified XorKII was
determined to be31,300 U/mg. The enzyme activity was monitored by cleaving lambda DNA or YEp24
plasmid as substrates. The enzyme was the most active at 10 mM Tris-HCl pH 7.0, 10 mM MgCl2, 1
mM dithiothreitol
at 37 oC. XorKII was easily inactivated by heating at 65 oC for 5 min, but retained most of
the original activity after incubation at 37oC for 24 h.

<>

<1>Moore, A.M., Patel, S., Forsberg, K.J., Wang, B., Bentley, G., Razia, Y., Qin, X., Tarr, P.I., Dantas, G.
<2>Pediatric fecal microbiota harbor diverse and novel antibiotic resistance genes.
<3>PLoS ONE
<4>8
<5>E78822
<6>2013
<7>Emerging antibiotic resistance threatens human health. Gut microbes are an
epidemiologically important reservoir of resistance genes (resistome), yet prior
studies indicate that the true diversity of gut-associated resistomes has been
underestimated. To deeply characterize the pediatric gut-associated resistome, we
created metagenomic recombinant libraries in an Escherichia coli host using fecal
DNA from 22 healthy infants and children (most without recent antibiotic
exposure), and performed functional selections for resistance to 18 antibiotics
from eight drug classes. Resistance-conferring DNA fragments were sequenced
(Illumina HiSeq 2000), and reads assembled and annotated with the PARFuMS
computational pipeline. Resistance to 14 of the 18 antibiotics was found in
stools of infants and children. Recovered genes included chloramphenicol
acetyltransferases, drug-resistant dihydrofolate reductases, rRNA
methyltransferases, transcriptional regulators, multidrug efflux pumps, and every
major class of beta-lactamase, aminoglycoside-modifying enzyme, and tetracycline
resistance protein. Many resistance-conferring sequences were mobilizable; some
had low identity to any known organism, emphasizing cryptic organisms as
potentially important resistance reservoirs. We functionally confirmed three
novel resistance genes, including a 16S rRNA methylase conferring aminoglycoside
resistance, and two tetracycline-resistance proteins nearly identical to a
bifidobacterial MFS transporter (B. longum s. longum JDM301). We provide the
first report to our knowledge of resistance to folate-synthesis inhibitors
conferred by a predicted Nudix hydrolase (part of the folate synthesis pathway).
This functional metagenomic survey of gut-associated resistomes, the largest of
its kind to date, demonstrates that fecal resistomes of healthy children are far
more diverse than previously suspected, that clinically relevant resistance genes
are present even without recent selective antibiotic pressure in the human host,
and that cryptic gut microbes are an important resistance reservoir. The observed
transferability of gut-associated resistance genes to a gram-negative (E. coli)
host also suggests that the potential for gut-associated resistomes to threaten
human health by mediating antibiotic resistance in pathogens warrants further
investigation.

<>

<1>Moore, G.P., Moore, A.R.
<2>The average spacing of restriction enzyme recognition sites in DNA.
<3>J. Theor. Biol.
<4>98
<5>165-169
<6>1982
<7>The discovery of naturally occurring enzymes which cleave DNA at sites specific to particular
nucleotide sequences has had a great impact on molecular biology.  The function of these
enzymes in vivo is to protect bacterial cells from viral invasion by degradation of foreign
DNA.  Several hundred of these "restriction" enzymes are known and they are a very common tool
both for analysis and manipulation of DNA.  The overwhelming majority of restriction enzyme
recognition sites are four to six nucleotides in length and have the remarkable property of
internal symmetry with respect to nucleotide sequence (i.e. diad symmetry).  A major practical
offshoot of the characterization of restriction enzymes has been their use in DNA cloning.  In
addition, comparison of restriction enzyme cleavage patterns has been used analytically to
assess the similarity of DNA from related organisms-particularly mitochondrial DNA.  As an aid
in these studies, a number of statistical methods have been devised to analyze data generated
by comparison of restriction enzyme digestion patterns.  The first of these studies was
carried out by Upholt and Upholt & Dawid.  This work was revised by Nei & Li and by Gotoh et
al.  The intent of this note is to add practical detail to these analyses.

<>

<1>Moore, K.H., Johnson, P.H., Chandler, E.W., Grossman, L.I.
<2>A restriction endonuclease cleavage map of mouse mitochondrial DNA.
<3>Nucleic Acids Res.
<4>4
<5>1273-1289
<6>1977
<7>A restriction endonuclease cleavage map is presented for mouse mitochrondrial DNA.  This map
was constructed by electron microscopic measurements on partial digests containing fixed
D-loops, and by electrophoretic analysis of partial and complete single enzyme digests, and of
double digests.  No map differences were detected between mitochrondrial DNA from cultured LA9
cells and an inbred mouse line for the six endonucleases used.  Three cleavage sites
recognized by HpaI, five sites recognized by HincII, two sites recognized by PstI and four
sites recognized by BamI were located with respect to the origin of replication and the EcoRI
and HinIII sites previously determined by others.  No cleavages were produced by KpnI or SalI.
The migration of linear DNA with a molecular weight greater than 1 x 10^6 was not a linear
function of log molecular weight in 1% agarose gels run at 6.6 volts/cm.

<>

<1>Moore, S.P., McAleer, M.A., Moss, S.H.
<2>Restriction enzyme cleavage of UV-irradiated DNA:  A comparison of far-UV and mid-UV wavelengths.
<3>Photochem. Photobiol.
<4>45
<5>253-263
<6>1987
<7>UV-irradiated DNA is less susceptible to restriction by Type II endonucleases
than unirradiated DNA presumably due to photolesions formed in the recognition
sites.  Previous reported studies have used 254 nm radiation or 313 nm plus
acetophenone, both treatments which introduce pyrimidine dimers in preference
to other photolesions.  To assess the effect of a longer wavelength, at which
the ratio of pyrimidine dimer formation to the formation of other photolesions
is reduced, two different DNAs were irradiated with UV of either 254 or 313 nm
and restricted with suitable restriction endonucleases.  Restriction patterns
were analysed for novel fragments resulting from UV-induced alteration of
enzyme recognition sites.  EcoRI restriction of 254 nm irradiated lambda DNA
produced six novel bands, only three of which were observed following
restriction of 313 nm irradiated lambda.  These three represented the largest
fragments resulting from single site blocks.  Novel fragments involving
adjacent site blocks observed at 254 nm were not found with 313 nm radiation.
Comparison of 254 nm irradiated pSVgpt to that irradiated at 313 nm, both
restricted with DraI, revealed a more complex pattern.  Although all sites were
singly blocked by radiation of both wavelengths, multiple site blocks produced
by 313 nm radiation did not occur in the order predicted by the 254 nm
radiation dose response.  These data suggest that certain sites in pSVgpt may
be more refractory to multiple site blocks than others when irradiated at 313
nm.

<>

<1>Morad, I., Chapman-Shimshoni, D., Amitsur, M., Kaufmann, G.
<2>Functional expression and properties of tRNALys-specific core anticodon nuclease encoded by Escherichia coli prrC.
<3>J. Biol. Chem.
<4>268
<5>26842-26849
<6>1993
<7>Escherichia coli carrying the optional locus prr harbor a latent, tRNALys-specific anticodon
nuclease, activated by the product of phage T4 stp.  Anticodon nuclease latency is ascribed to
the masking of prrC, implicated with the enzymatic activity, by flanking, type Ic DNA
restriction modification genes (prrA, B&D-hsdM, S&R).  Overexpression of plasmid-borne prrC
elicited anticodon nuclease activity in uninfected E. coli.  In vitro, the prrC-coded core
activity was indifferent to a synthetic Stp polypeptide, GTP, ATP, and endogenous DNA,
effectors that synergistically activate the latent enzyme.  Several facts suggested that PrrC
is highly labile in the absence of the masking proteins.  The core activity decayed with t1/2
below 1 min at 30oC, and the PrrC portion of a fusion protein was unstable.  Moreover,
expression of prrC from its own promoter at low plasmid copy number did not allow detection of
core activity.  Yet, it sufficed for establishment of a latent, T4-inducible enzyme when
complemented by the masking Hsd proteins, which were provided by another replicon.
Interaction between the antagonistic components of latent anticodon nuclease was also
demonstrated immunochemically.  The coupling of anticodon nuclease with a DNA restriction
modification system may serve to ward off its inadvertent toxicity and maintain it as an
antiviral contingency.

<>

<1>Morais, L.L., Garza, D.R., Loureiro, E.C., Nunes, K.N., Vellasco, R.S., da Silva, C.P., Nunes, M.R., Thompson, C.C., Vicente, A.C., Santos, E.C.
<2>Complete Genome Sequence of a Sucrose-Nonfermenting Epidemic Strain of Vibrio cholerae O1 from Brazil.
<3>J. Bacteriol.
<4>194
<5>2772
<6>2012
<7>We report the genome sequence of Vibrio cholerae strain IEC224, which fails to ferment
sucrose. It was isolated from a cholera outbreak in the Amazon. The
defective sucrose phenotype was determined to be due to a frameshift mutation,
and a molecular marker of the Latin American main epidemic lineage was
identified.

<>

<1>Morais-Silva, F.O., Santos, C.I., Rodrigues, R., Pereira, I.A., Rodrigues-Pousada, C.
<2>Role of HynAB and Ech, the only two hydrogenases found in the model sulfate reducer Desulfovibrio gigas.
<3>J. Bacteriol.
<4>195
<5>4753
<6>2013
<7>Sulfate-reducing bacteria are characterized by a high number of hydrogenases, which have been
proposed to contribute to the overall energy metabolism of the cell, but exactly in what role
is not clear. Desulfovibrio spp. can produce or consume H2 when growing on organic or
inorganic substrates in the presence or absence of sulfate. Because of the presence of only
two hydrogenases encoded in its genome, the periplasmic HynAB and cytoplasmic Ech
hydrogenases, Desulfovibrio gigas is an excellent model organism for investigation of the
specific function of each of these enzymes during growth. In this study, we analyzed the
physiological response to the deletion of the genes that encode the two hydrogenases in D.
gigas, through the generation of DeltaechBC and DeltahynAB single mutant strains. These
strains were analyzed for the ability to grow on different substrates, such as lactate,
pyruvate, and hydrogen, under respiratory and fermentative conditions. Furthermore, the
expression of both hydrogenase genes in the three strains studied was assessed through
quantitative reverse transcription-PCR. The results demonstrate that neither hydrogenase is
essential for growth on lactate-sulfate, indicating that hydrogen cycling is not
indispensable. In addition, the periplasmic HynAB enzyme has a bifunctional activity and is
required for growth on H2 or by fermentation of pyruvate. Therefore, this enzyme seems to play
a dominant role in D. gigas hydrogen metabolism.

<>

<1>Moran, J.V., Mecklenburg, K.L., Sass, P., Belcher, S.M., Mahnke, D., Lewin, A., Perlman, P.
<2>Splicing defective mutants of the COXI gene of yeast mitochondrial DNA: initial definition of the maturase domain of the group II intron AI2.
<3>Nucleic Acids Res.
<4>22
<5>2057-2064
<6>1994
<7>Six mutations blocking the function of a seven intron form of the mitochondrial gene encoding
subunit I of cytochrome c oxidase (COXI) and mapping upstream of exon 3 were isolated and
characterized.  A cis-dominant mutant of the group IIA intron 1 defines a helical portion of
the C1 substructure of domain 1 as essential for splicing.  A trans-recessive mutant confirms
that the intron 1 reading frame encodes a maturase function.  A cis-dominant mutant in exon 2
was found to have no effect on the splicing of intron 1 or 2.  A trans-recessive mutant,
located in the group IIA intron 2, demonstrates for the first time that intron 2 encodes a
maturase.  A genetic dissection of the five missense mutations present in the intron 2 reading
frame of that strain demonstrates that the maturase defect results from one or both of the
missense mutations in a newly-recognized conserved sequence called domain X.

<>

<1>Moran, J.V., Wernette, C.M., Mecklenburg, K.L., Butow, R.A., Perlman, P.S.
<2>Intron 5a of the COXI gene of yeast mitochondrial DNA is a mobile group I intron.
<3>Nucleic Acids Res.
<4>20
<5>4069-4076
<6>1992
<7>We have found that intron 5a of the COXI gene (aI5a) of yeast mtDNA is a mobile group I intron
in crosses between strains having or lacking the intron. We have demonstrated the following
hallmarks of that process: 1) co-conversion of flanking optional intron markers; 2) mutations
that truncate the intron open reading frame block intron mobility; and 3) the intron open
reading frame encodes an endonuclease activity that is required for intron movement. The
endonuclease activity, termed I-SceIV, cleaves the COXI allele lacking aI5a near the site of
intron insertion, making a four-base staggered cut with 3' OH overhangs. Three cloned DNAs
derived from different forms of the COXI gene, which differ in primary sequence at up to seven
nucleotides around the cleavage site, are all good substrates for in vitro I-SceIV cleavage
activity. Two of the strains from which these substrates derived were tested in crosses and
are comparably efficient as aI5a recipients. When compared with omega mobility occurring
simultaneously in one cross, aI5a is less efficient as a mobile element.

<>

<1>Moran, J.V., Zimmerly, S., Eskes, R., Kennell, J.C., Lambowitz, A.M., Butow, R.A., Perlman, P.S.
<2>Mobile group II introns of yeast mitochrondrial DNA are novel site-specific retroelements.
<3>Mol. Cell. Biol.
<4>15
<5>2828-2838
<6>1995
<7>Group II introns aI1 and aI2 of the yeast mitochondrial COX1 gene are mobile elements that
encode an intron-specific reverse transcriptase (RT) activity.  We show here that the introns
of Saccharomyces cerevisiae ID41-6/161 insert site specifically into intronless alleles.  The
mobility is accompanied by efficient, but highly asymmetric, coconversion of nearby flanking
exon sequences.  Analysis of mutants shows that the aI2 protein is required for the mobility
of both aI1 and aI2.  Efficient mobility is dependent on both the RT activity of the
aI2-encoded protein and a separate function, a putative DNA endonuclease, that is associated
with the Zn2+ finger-like region of the intron reading frame.  Surprisingly, there appear to
be two  mobility modes: the major one involves cDNAs reverse transcribed from unspliced
precursor RNA; the minor one, observed in two mutants lacking detectable RT activity, appears
to involve DNA level recombination.  A cis-dominant splicing-defective mutant of aI2 continues
to synthesize cDNAs containing the introns but is completely defective in both mobility modes,
indicating that the splicing or the structure of the intron is required.  Our results
demonstrate that the yeast group II intron aI2 is a retroelement that uses novel mobility
mechanisms.

<>

<1>Moran, M.A. et al.
<2>Genome sequence of Silicibacter pomeroyi reveals adaptations to the marine environment.
<3>Nature
<4>432
<5>910-913
<6>2004
<7>Since the recognition of prokaryotes as essential components of the oceanic food web,
bacterioplankton have been acknowledged as catalysts of
most major biogeochemical processes in the sea. Studying heterotrophic
bacterioplankton has been challenging, however, as most major clades have
never been cultured or have only been grown to low densities in sea water.
Here we describe the genome sequence of Silicibacter pomeroyi, a member of
the marine Roseobacter clade (Fig. 1), the relatives of which comprise
approximately 10-20% of coastal and oceanic mixed-layer bacterioplankton.
This first genome sequence from any major heterotrophic clade consists of
a chromosome (4,109,442 base pairs) and megaplasmid (491,611 base pairs).
Genome analysis indicates that this organism relies upon a
lithoheterotrophic strategy that uses inorganic compounds (carbon monoxide
and sulphide) to supplement heterotrophy. Silicibacter pomeroyi also has
genes advantageous for associations with plankton and suspended particles,
including genes for uptake of algal-derived compounds, use of metabolites
from reducing microzones, rapid growth and cell-density-dependent
regulation. This bacterium has a physiology distinct from that of marine
oligotrophs, adding a new strategy to the recognized repertoire for coping
with a nutrient-poor ocean.

<>

<1>Moran, N.A., Degnan, P.H., Santos, S.R., Dunbar, H.E., Ochman, H.
<2>The players in a mutualistic symbiosis: Insects, bacteria, viruses, and virulence genes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>16919-16926
<6>2005
<7>Aphids maintain mutualistic symbioses involving consortia of coinherited
organisms. All possess a primary endosymbiont, Buchnera, which compensates
for dietary deficiencies; many also contain secondary symbionts, such as
Hamiltonella defensa, which confers defense against natural enemies.
Genome sequences of uncultivable secondary symbionts have been refractory
to analysis due to the difficulties of isolating adequate DNA samples. By
amplifying DNA from hemolymph of infected pea aphids, we obtained a set of
genomic sequences of H. defensa and an associated bacteriophage. H.
defensa harbors two type III secretion systems, related to those that
mediate host cell entry by enteric pathogens. The phage, called APSE-2, is
a close relative of the previously sequenced APSE-1 but contains intact
homologs of the gene encoding cytolethal distending toxin (cdtB), which
interrupts the eukaryotic cell cycle and which is known from a variety of
mammalian pathogens. The cdtB homolog is highly expressed, and its genomic
position corresponds to that of a homolog of stx (encoding Shiga-toxin)
within APSE-1. APSE-2 genomes were consistently abundant in infected pea
aphids, and related phages were found in all tested isolates of H.
defensa, from numerous insect species. Based on their ubiquity and
abundance, these phages appear to be an obligate component of the H.
defensa life cycle. We propose that, in these mutualistic symbionts,
phage-borne toxin genes provide defense to the aphid host and are a basis
for the observed protection against eukaryotic parasites.

<>

<1>Mordasini, T., Curioni, A., Andreoni, W.
<2>Why do divalent metal ions either promote or inhibit enzymatic reactions? The case of BamHI restriction endonuclease from combined quantum-classical simulations.
<3>J. Biol. Chem.
<4>278
<5>4381-4384
<6>2003
<7>Divalent metal ions are essential to many enzymatic reactions involving nucleic acids, but
their critical and specific role still needs to be uncovered. Restriction endonucleases are a
prominent group of such metal-requiring enzymes. Large-scale accurate simulations of Mg- and
Ca-BamHI elucidate the mechanism of the catalytic reaction leading to DNA cleavage and show
that it involves the concerted action of two metal ions and water molecules. It is also
established that what is decisive for the dramatically different behavior of magnesium (a
cocatalyst) and calcium (an inhibitor) are kinetic factors and not the properties of the
pre-reactive states of the enzymes. A new perspective is opened for the understanding of the
functional role of metal ions in biological processes.

<>

<1>Moreau, H., Piganeau, G., Desdevises, Y., Cooke, R., Derelle, E., Grimsley, N.
<2>Marine prasinovirus genomes show low evolutionary divergence and acquisition of protein metabolism genes by horizontal gene transfer.
<3>J. Virol.
<4>84
<5>12555-12563
<6>2010
<7>Although marine picophytoplankton are at the base of the global food
chain, accounting for half of the planetary primary production, they are
outnumbered 10 to 1 and are largely controlled by hugely diverse
populations of viruses. Eukaryotic microalgae form a ubiquitous and
particularly dynamic fraction of such plankton, with environmental clone
libraries from coastal regions sometimes being dominated by one or more of
the three genera Bathycoccus, Micromonas, and Ostreococcus (class
Prasinophyceae). The complete sequences of two double-stranded (dsDNA)
Bathycoccus, one dsDNA Micromonas, and one new dsDNA Ostreococcus virus
genomes are described. Genome comparison of these giant viruses revealed a
high degree of conservation, both for orthologous genes and for synteny,
except for one 36-kb inversion in the Ostreococcus lucimarinus virus and
two very large predicted proteins in Bathycoccus prasinos viruses. These
viruses encode a gene repertoire of certain amino acid biosynthesis
pathways never previously observed in viruses that are likely to have been
acquired from lateral gene transfer from their host or from bacteria.
Pairwise comparisons of whole genomes using all coding sequences with
homologous counterparts, either between viruses or between their
corresponding hosts, revealed that the evolutionary divergences between
viruses are lower than those between their hosts, suggesting either
multiple recent host transfers or lower viral evolution rates.

<>

<1>Moreira, R.F., Noren, C.J.
<2>Minimum duplex requirements for restriction enzyme cleavage near the termini of linear DNA fragments.
<3>Biotechniques
<4>19
<5>56-59
<6>1995
<7>Restriction endonucleases differ in their ability to cleave close to the ends of linear DNA
fragments, each requiring a characteristic minimum number of base pairs adjacent to their
recognition sites for efficient cleavage. Quantitative cleavage close to either end of a
linear DNA fragment is crucial when carrying out digests at adjacent restriction sites within
cloning vector polylinkers or when trimming the ends of polymerase chain reaction (PCR)
products prior to cloning. In the former case, incomplete digestion by the second restriction
enzyme (following linearization by the first) would be undetectable by gel electrophoresis,
yet could result in an unacceptably high background level of transformants following ligation
of the insert. In the latter case, recovery of a cloned PCR product is dependent upon
generation of the desired overhangs at either end of the amplified product prior to cloning.
Incorporation of restriction sites within PCR primers for future cloning steps has become an
important factor in primer design: If not enough bases are added upstream from each site, then
the corresponding enzyme will not cut the PCR product.

<>

<1>Moretti, C., Cortese, C., Passos-da-Silva, D., Venturi, V., Firrao, G., Buonaurio, R.
<2>Draft Genome Sequence of Erwinia oleae, a Bacterium Associated with Olive Knots Caused by Pseudomonas savastanoi pv. savastanoi.
<3>Genome Announcements
<4>2
<5>e01308-14
<6>2014
<7>Erwinia oleae is a nonpathogenic bacterial species isolated from olive knots caused by
Pseudomonas savastanoi pv. savastanoi. Since the presence of E. oleae
in the knots increases disease severity, interspecies interactions with the
pathogen are hypothesized. Here, we report the first draft genome sequence of the
E. oleae type strain.

<>

<1>Moretti, C., Cortese, C., Passos-da-Silva, D., Venturi, V., Ramos, C., Firrao, G., Buonaurio, R.
<2>Draft Genome Sequence of Pseudomonas savastanoi pv. savastanoi Strain DAPP-PG 722, Isolated in Italy from an Olive Plant Affected by Knot Disease.
<3>Genome Announcements
<4>2
<5>e00864-14
<6>2014
<7>Olive knot disease, caused by the bacterium Pseudomonas savastanoi pv. savastanoi, seriously
affects olive trees in the Mediterranean basin. Here, we
report the draft genome sequence of P. savastanoi pv. savastanoi DAPP-PG 722, a
strain isolated in Italy from an olive plant affected by knot disease.

<>

<1>Moretti, C., Cortese, C., Passos-da-Silva, D., Venturi, V., Torelli, E., Firrao, G., Buonaurio, R.
<2>Draft Genome Sequence of a Hypersensitive Reaction-Inducing Pantoea agglomerans Strain Isolated from Olive Knots Caused by Pseudomonas savastanoi pv. savastanoi.
<3>Genome Announcements
<4>2
<5>e00774-14
<6>2014
<7>Pantoea agglomerans strains inducing a hypersensitive reaction in tobacco leaves  are
frequently isolated inside olive knots caused by Pseudomonas savastanoi pv.
savastanoi. Here, we report the draft genome sequence of the Italian P.
agglomerans strain, which is able to increase olive knot disease severity when
coinoculated with P. savastanoi pv. savastanoi.

<>

<1>Morgan, R.
<2>A type II restriction endonuclease obtainable from Arthrobacter species and a process for producing the same.
<3>European Patent Office
<4>EP 0498404 B
<5>
<6>1992
<7>The present invention relates to a new Type II restriction endonuclease, AscI, obtainable from
Arthrobacter species, and to the process for producing the same.

<>

<1>Morgan, R.
<2>Method for producing the AseI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0332406 B
<5>
<6>1994
<7>The present invention relates to clones for the AseI restriction endonuclease and modification
methylase, and to the production of these enzymes from the clones.

<>

<1>Morgan, R.
<2>A novel Type II restriction endonuclease, PmeI, obtainable from Pseudomonas mendocina and a process for producing the same.
<3>European Patent Office
<4>EP 0517111 B
<5>
<6>1994
<7>The present invention relates to a new Type II restriction endonuclease, PmeI, obtainable from
Pseudomonas mendocina, and to the process for producing the same

<>

<1>Morgan, R., Heiter, D., Lunnen, K., Wilson, G., Benner, J., Nkenfou, C.N., Picone, S.
<2>A Novel Modular Type II Restriction Endonuclease, CspCI, and the Use of Modular Endonucleases for Generating Endonucleases with New.
<3>Japanese Patent Office
<4>JP 2007530046 A
<5>
<6>2007
<7>
<>

<1>Morgan, R., Nkenfou, C.N.
<2>Novel type II restriction endonuclease, CspCI, obtainable from Citrobacter species 2144 and a process for producing the same.
<3>US Patent Office
<4>US 20050233433 A
<5>
<6>2005
<7>A novel restriction endonuclease has been identified from Citrobacter species 2144 (NEB#1398)
which can cleave at nt sequence 5'-CAANNNNNGTGG-3' (SEQ ID NO:13) in double-stranded DNA
molecules.

<>

<1>Morgan, R., Nkenfou, C.N.
<2>CspCI restriction endonuclease.
<3>US Patent Office
<4>US 7247464 B
<5>
<6>2007
<7>A  novel restriction endonuclease has been identified from Citrobacter species 2144 (NEB#1398)
which can cleave at nt sequence 5'-CAANNNNNGTGG-3' (SEQ ID NO:13) in double-stranded DNA
molecules.

<>

<1>Morgan, R., Wilson, G., Lunnen, K., Heiter, D., Benner, J., Nkenfou, C.N., Picone, S.
<2>Novel modular type II restriction endonuclease CspCI, and the use of modular endonucleases for generating endonucleases with new specificities.
<3>International Patent Office
<4>WO 200594516 A
<5>
<6>2005
<7>A novel restriciton endonuclease and methods of making the same are obtianable from either
Citrobacter species 2144 (NEB#1398) or the recombinant strain Escherichia coli (NEB#1554)
which cleaves at nt sequence 5'-CAANNNNNGTGG-3' (SEQ ID NO:14) in double-stranded DNA
molecules.  The novel restriction endonuclease is a modular protein in which the specificity
moiety is an independent module from the restriciton-modification module.

<>

<1>Morgan, R., Wilson, G., Lunnen, K., Heiter, D., Benner, J., Nkenfou, C.N., Picone, S.
<2>Novel modular type II restriction endonuclease, CspCI, and the use of modular endonucleases for generating endonucleases with new specificities.
<3>US Patent Office
<4>US 20070166719 A
<5>
<6>2007
<7>A novel restriction endonuclease and methods of making the same are obtainable from either
Citrobacter species 2144 (NEB#1398) or the recombinant strain Escherichia coli (NEB#1554)
which cleaves at nt sequence 5'-CAANNNNNGTGG-3' (SEQ ID NO:14) in double-stranded DNA
molecules.  The novel restriction endonuclease is a modular protein in which the specificity
moiety is an independent module from the restriction-modification module.

<>

<1>Morgan, R., Xiao, J.-P., Xu, S.-Y.
<2>Characterization of an extremely thermostable restriction enzyme, PspGI, from a Pyrococcus strain and cloning of the PspGI restriction-modification system in Escherichia coli.
<3>Appl. Environ. Microbiol.
<4>64
<5>3669-3673
<6>1998
<7>An extremely thermostable restriction endonuclease, PspGI, was purified from Pyrococcus sp.
strain GI-H.  PspGI is an isoschizomer of EcoRII and cleaves DNA before the first C in the
sequence 5' ^CCWGG 3' (W is A or T).  PspGI digestion can be carried out at 65 to 85 C.  To
express PspGI at high levels, the PspGI restriction-modification genes (pspGIR and pspGIM)
were cloned in Escherichia coli.  M.PspGI contains the conserved sequence motifs of
alpha-aminomethyltransferases; therefore, it must be an N4-cytosine methylase.  M.PspGI shows
53% similarity to (44% identity with) its isoschizomer, M.MvaI from Micrococcus variabilis.
In a segment of 87 amino acid residues, PspGI shows significant sequence similarity to EcoRII
and to regions of SsoII and StyD4I which have a closely related recognition sequence (5'
^CCNGG 3').  PspGI was expressed in E. coli via a T7 expression system.  Recombinant PspGI
was purified to near homogeneity and had a half-life of 2 h at 95 C.  PspGI remained active
following 30 cycles of thermocycling; thus, it can be used in DNA-based diagnostic
applications.

<>

<1>Morgan, R., Zhou, B.
<2>Type II restriction endonuclease, Pme I, obtainable from Pseudomonas mendocina and a process for producting the same.
<3>US Patent Office
<4>US 5196330
<5>
<6>1993
<7>*
The present invention provides a novel type II restriction endonuclease obtainable from
Pseudomonas mendocina. The endonuclease known as PmeI, recognizes the following nucleotide
sequence and has a cleavage point indicated by the arrows:   5'GTTT^AAAC3'
                                                             3'CGGG^TTTG5'
Also described is a process for obtaining PmeI from Pseudomonas mendocina.


<>

<1>Morgan, R.D.
<2>Type II restriction endonuclease AscI, obtainable from Arthrobacter species and a process for producting the same.
<3>US Patent Office
<4>US 5192676
<5>
<6>1993
<7>*
The present invention provides a novel type II restriction endonuclease obtainable from
Arthrobacter species. The endonuclease known as AscI, recognizes the following nucleotide
sequence and has a cleavage point indicated by the arrows:   5' GG^CGCGCC 3'
                                                             3' CCGCGC^GG 5'
Also described is a process for obtaining AscI from Arthrobacter species.


<>

<1>Morgan, R.D.
<2>Method for producing the NlaIII restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0477532 B
<5>
<6>1995
<7>Recombinant DNA encoding the NlaIII restriction endonuclease and modification methylase, and
method for the production of these enzymes from said recombinant DNA.

<>

<1>Morgan, R.D.
<2>Isolated DNA encoding the FseI restriction endonuclease and related methods for producing the same.
<3>European Patent Office
<4>EP 0712933 A
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the FseI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Frankia species into a
host whereby the restriction gene is expressed; 2) fermenting the host which contains the
plasmid encoding and expressing the FseI restriction endonuclease activity, and 3) purifying
the FseI restriction endonuclease from the fermented host which contains the plasmid encoding
and expressing the FseI restriction endonuclease activity.

<>

<1>Morgan, R.D.
<2>Method for producing that NlaIII restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5278060
<5>
<6>1994
<7>Recombinant DNA encoding the NlaIII restriction endonuclease and modification methylase, and
methods for the production of these enzymes from said recombinant DNA.

<>

<1>Morgan, R.D.
<2>Method for producing the NlaIV restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5075232
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the NlaIV restriction
endonuclease by 1) introducing the restriction endonuclease gene from N. lactamica NRCC 2118
into a host whereby the restriction gene is expressed; 2) fermenting the host which contains
the vector encoding and expressing the NlaIV restriction endonuclease, and 3) purifying the
NlaIV restriction endonuclease from the fermented host which contains the vector encoding and
expressing the NlaIV restriction endonuclease activity.

<>

<1>Morgan, R.D.
<2>Rational design of binding proteins that recognize desired specific sequences.
<3>International Patent Office
<4>WO 2008157789 A
<5>
<6>2008
<7>Methods and compositions are provided for creating a binding protein that recognizes a
rationally chosen recognition sequence in which a first amino acid has been substituted for a
second amino acid using site-directed mutagenesis of a member protein of a set of proteins at
an identified position or positions correlated with recognition of a chosen specified target
module in the recognition sequence.  A system is provided for automating the storage and
manipulation of the correlations between positions and types of amino acid residues in the
binding protein with specific modules at specified positions in the target recognition
sequence and for designing and creating proteins with novel specificities.

<>

<1>Morgan, R.D.
<2>MseI, a unique restriction endonuclease from Micrococcus species which recognizes 5'T^TAA 3'.
<3>Nucleic Acids Res.
<4>16
<5>3104
<6>1988
<7>A new typeII restriction endonuclease, MseI, has been isolated from Micrococcus
species (NEB446).  MseI recognizes the palindromic sequence 5'TTAA3' and
cleaves between the two T residues to produce a 2 base 5' extension: T/TAA.

<>

<1>Morgan, R.D.
<2>Type II restriction endonuclease NmeAIII from Neisseria meningitidis and its recombinant production and use in genetic technologies.
<3>International Patent Office
<4>WO 2008083065 A
<5>
<6>2008
<7>A protein is described that has an amino acid sequence characterized by at least 90% sequence
identity with SEQ ID NO: 24, the protein being capable of recognizing a sequence consisting of
5'-GCCGAG-3' within the double-stranded DNA and cleaving the substrate predominantly at
21/19 nucleotides from the recognition site.  A method is also described that utilizes the
protein for creating a DNA tag for use as a unique identifier for paired and sequencing of DNA
or serial analysis of gene expression.

<>

<1>Morgan, R.D.
<2>Rational engineering of DNA binding and cleavage specificity in a family of Type II restriction endonucleases.
<3>Ph.D. Thesis, Boston University
<4>
<5>
<6>2009
<7>Type II restriction endonucleases serve as important tools for molecular biology. Although
enzymes recognizing 274 unique sequences are known, it would be desirable to be able to create
'designer endonucleases' to cut at sequences of choice rather than rely on the limited
number of natural isolates. However, these endonucleases have proved remarkably resistant to
all attempts to engineer changes in their recognition specificity. The Type II endonucleases
typically exhibit little sequence similarity. This study has identified a new family of
unusual Type II endonucleases through genome mining that do share a great deal of sequence
conservation. Twenty individual members of this family were biochemically characterized and
all recognize unique DNA sequences. These enzymes also employ a unique mode of modification,
in that they modify only one DNA strand for host protection against the action of the
endonuclease. This family of single-strand-modifying enzymes serves as the archetype for a new
restriction endonuclease subclass, the Type IIL group. Because these enzymes encode
endonuclease, methyltransferase and specificity functions in the same polypeptide, alterations
to the recognition domain result in a coordinated change in specificity for both host
protection and endonuclease activity. To alter recognition specificity, multiple sequence
alignments of the recognition sequences and protein sequences were created and interrogated to
identify correlations between position-specific amino acid residues and position-specific DNA
base recognition. Correlations were identified between pairs of amino acid positions and the
DNA bases recognized at three separate recognition positions. Enzymes having new recognition
specificity were then created by altering the amino acid residues at the correlating positions
to residues correlated with recognition of a desired new DNA base. Using this approach three
positions in the recognition sequences have been altered singly and in combination to create a
dozen novel Type II endonucleases having predictable new recognition sequences. From this work
it is now possible to rationally engineer hundreds of new Type II restriction endonucleases
specificities. These results move us closer to the goal of creating 'designer restriction
endonucleases' that recognize and cleave at any desired DNA sequence.

<>

<1>Morgan, R.D.
<2>Isolated DNA encoding the FseI restriction endonuclease and related methods for producing the same.
<3>US Patent Office
<4>US 5543308
<5>
<6>1996
<7>The present invention is directed to a method for cloning and producing the FseI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Frankia species into a
host whereby the restriction gene is expressed; 2) fermenting the host which contains the
plasmid encoding and expressing the FseI restriction endonuclease activity, and 3) purifying
the FseI restriction endonuclease from the fermented host which contains the plasmid encoding
and expressing the FseI restriction endonuclease activity.

<>

<1>Morgan, R.D.
<2>Method for producing the AseI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5100793
<5>
<6>1992
<7>The present invention is directed to a method for cloning and producing the AseI restriction
endonuclease by 1) introducing the restriction endonuclease gene from A. serpens ATCC 12638
into a host whereby the restriction gene is expressed; 2) fermenting the host which contains
the vector encoding and expressing the AseI restriction endonuclease, and 3) purifying the
AseI restriction endonuclease from the fermented host which contains the vector encoding and
expressing the AseI restriction endonuclease activity.

<>

<1>Morgan, R.D.
<2>Method for producing the NlaIV restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0355981 A
<5>
<6>1988
<7>The present invention is directed to a method for cloning and producing the NlaIV restriction
endonuclease by 1) introducing the restriction endonuclease gene obtainable from N. lactamica
into a host whereby the restriction gene is expressed; 2) fermenting the host which contains
the vector encoding and expressing the NlaIV restriction endonuclease, and 3) purifying the
NlaIV restriction endonuclease from the fermented host which contains the vector encoding and
expressing the NlaIV restriction endonuclease activity. The preferred source of DNA is N.
lactamica NRCC 2118.

<>

<1>Morgan, R.D.
<2>Recombinant Type II restriction endonuclease, NmeAIII, and a process for producing the same.
<3>US Patent Office
<4>US 20100159534
<5>
<6>2010
<7>A protein is described that has an amino acid sequence characterized by at least 90% sequence
identity with SEQ ID NO: 24, the protein being capable of recognizing a sequence consisting of
5'-GCCGAG-3' within the double-stranded DNA and cleaving the substrate predominantly at
21/190 nucleotides from the recognition site.  A method is also described that utilizes the
protein for creating a DNA tag for use as a unique identifier for paired end sequencing of DNA
or serial analysis of gene expression.

<>

<1>Morgan, R.D.
<2>Recombinant type II restriction endonuclease, NmeAIII, and a process for producing the same.
<3>US Patent Office
<4>US 08361774
<5>
<6>2013
<7>A protein is described that has an amino acid sequence characterized by at least 90% sequence
identity with SEQ ID NO: 24, the protein being
capable of recognizing a sequence consisting of 5 '-GCCGAG-3 ' within
the double-stranded DNA and cleaving the substrate predominantly at
21/19 nucleotides from the recognition site. A method is also described
that utilizes the protein for creating a DNA tag for use as a unique
identifier for paired end sequencing of DNA or serial analysis of gene
expression.

<>

<1>Morgan, R.D., Benner, J.S., Claus, T.E.
<2>Isolated DNA encoding the NotI restriction endonuclease and related methods for producing the same.
<3>US Patent Office
<4>US 5371006
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the NotI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Nocardia
otitidis-caviarum into a host whereby the restriction gene is expressed; 2) fermenting the
host which contains the plasmid encoding and expressing the NotI restriction endonuclease
activity, and 3) purifying the NotI restriction endonuclease from the fermented host which
contains the plasmid encoding and expressing the NotI restriction endonuclease activity.

<>

<1>Morgan, R.D., Benner, J.S., Claus, T.E.
<2>Isolated DNA encoding the NotI restriction endonuclease and related methods for producing the same.
<3>European Patent Office
<4>EP 0607005 B
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the NotI restriction
endonuclease by 1) introducing the restriction endonuclease gene from Nocardia
otitidis-caviarum into a host whereby the restriction gene is expressed; 2) fermenting the
host which contains the plasmid encoding and expressing the NotI restriction endonuclease
activity, and 3) purifying the NotI restriction endonuclease from the fermented host which
contains the plasmid encoding and expressing the NotI restriction endonuclease activity.

<>

<1>Morgan, R.D., Bhatia, T.
<2>Novel type II restriction endonuclease, CstMI, obtainable from corynebacterium striatum m82b.
<3>US Patent Office
<4>US 20050009034 A
<5>28
<6>2005
<7>In accordance with the present invention, there is provided a novel type II restriction
endonuclease, obtainable from Corynebacterium striatum M82B, hereinafter referred to as
"CstMI", which endonuclease:  (1) recognizes the nucleotide sequence 5'-AAGGAG-3' in a
double-stranded DNA molecule as shown below,
5'-AAGGAGN20/-3'
3'-TTCCTCN18/-5'
(wherein G represents guanine, C represents cytosine, A represents adenine, T represents
thymine and N represents either G, C, A, or T); (2) cleaves said sequence at the
phosphodiester bonds between the 20th and the 21st nucleotides 3' to the recognition sequence
in the 5'-AAGGAG-3 strand of the DNA, and between the 18th and 19th nucleotides 5' to the
recognition sequence in the complement stand, 5'-CTCCTT-3', to produce a 2 base 3'
extension; and (3) possesses a second enzymatic activity that recognizes the same DNA
sequence, 5'-AAGGAG-3', but modifies this sequence by the addition of a methyl group to
prevent cleavage by the CstMI endonuclease activity.

<>

<1>Morgan, R.D., Bhatia, T.
<2>Type II restriction endonuclease, CstMI, obtainable from Corynebacterium striatum M82B and a process for producing the same.
<3>US Patent Office
<4>US 07186538
<5>
<6>2007
<7>In accordance with the present invention, there is provided a novel type II restriction
endonuclease, obtainable from Corynebacterium
striatum M82B, hereinafter referred to as "CstMI", which endonuclease:
(1) recognizes the nucleotide sequence 5 '-AAGGAG-3 ' in a
double-stranded DNA molecule as shown below, 5 '-AAGGAGN20 down arrow-3
' 3 '-TTCCTCN18 up arrow-5 ' (wherein G represents guanine, C
represents cytosine, A represents adenine, T represents thymine and N
represents either G, C, A, or T); (2) cleaves said sequence at the
phosphodiester bonds between the 20th and the 21th nucleotides 3 ' to
the recognition sequence in the 5 '-AAGGAG-3 strand of the DNA, and
between the 18th and 19th nucleotides 5 ' to the recognition sequence
in the complement stand, 5 '-CTCCTT-3 ', to produce a 2 base 3 '
extension; and (3) possesses a second enzymatic activity that
recognizes the same DNA sequence, 5 '-AAGGAG-3 ', but modifies this
sequence by the addition of a methyl group to prevent cleavage by the
CstMI endonuclease activity.

<>

<1>Morgan, R.D., Bhatia, T., Davis, T., Lovasco, L.
<2>Recombinant type II restriction endonucleases, MmeI and related endonucleases and methods for producing the same.
<3>International Patent Office
<4>WO 2004007670
<5>
<6>2004
<7>In accordance with the present invention, there is provided a DNA (deoxyribonucleic acid)
fragment which encodes the MmeI type II restriction endonuclease enzyme.  This one polypeptide
possesses two related enzymatic functions; namely an endonuclease activity which recognizes
the DNA sequence 5'-TCC(Pu)AC-3' and cleaves as indicated by the arrows: 5'-TCCRAC(N20)
/-3' 3'-AGGYTG(N18)/-5' and a second enzymatic activity that recognizes the same DNA
sequence, 5'-TCC(Pu)AC-3', but modifies this sequence by the addition of a methyl group to
prevent cleavage by the MmeI endonuclease activity.

<>

<1>Morgan, R.D., Bhatia, T., Davis, T., Lovasco, L.
<2>Recombinant type II restriction endonucleases, MmeI and related endonucleases and methods for producing the same.
<3>US Patent Office
<4>US 7115407 B
<5>
<6>2006
<7>In accordance with the present invention, there is provided a DNA (deoxyribonucleic acid)
fragment which encodes the MmeI type II restriction endonuclease enzyme.  This one polypeptide
possesses two related enzymatic functions; namely an endonuclease activity which recognizes
the DNA sequence 5'-TCC(Pu)AC-3' and cleaves as indicated by the arrows:
5'-TCCRAC(N20)/-3' 3'-AGGYTG(N18)/-5' and a second enzymatic activity that recognizes the
same DNA sequence, 5'-TCC(Pu)AC-3', but modifies this sequence by the addition of a methyl
group to prevent cleavage by the MmeI endonuclease activity.

<>

<1>Morgan, R.D., Bhatia, T., Davis, T., Lovasco, L.
<2>Recombinant type II restriction endonucleases, Mmel and related endonucleases and methods for producing the same.
<3>European Patent Office
<4>EP 2228442 A
<5>
<6>2010
<7>In accordance with the present invention, there is provided a DNA fragment which encodes the
MmeI type II restriction endonuclease enzyme.  This one polypeptide possesses two related
enzymatic functions; namely an endonuclease activity which recognizes the DNA sequence
5'-TCC(Pu)AC-3' and cleaves as indicated by the arrows:
5'-TCCRAC(N2O)/-3'3'-AGGYTG(N18)/-5' and a second enzymatic activity that recognizes the
same DNA sequence, 5'-TCC(Pu)AC-3', but modifies this sequence by the addition of a methyl
group to prevent cleavage by the MmeI endonuclease activity.

<>

<1>Morgan, R.D., Bhatia, T.K., Lovasco, L., Davis, T.B.
<2>MmeI: a minimal Type II restriction-modification system that only modifies one DNA strand for host protection.
<3>Nucleic Acids Res.
<4>36
<5>6558-6570
<6>2008
<7>MmeI is an unusual Type II restriction enzyme that is useful for generating long sequence
tags. We have cloned the MmeI
restriction-modification (R-M) system and found it to consist of a single
protein having both endonuclease and DNA methyltransferase activities. The
protein comprises an amino-terminal endonuclease domain, a central DNA
methyltransferase domain and C-terminal DNA recognition domain. The
endonuclease cuts the two DNA strands at one site simultaneously, with
enzyme bound at two sites interacting to accomplish scission. Cleavage
occurs more rapidly than methyl transfer on unmodified DNA. MmeI modifies
only the adenine in the top strand, 5'-TCCRAC-3'. MmeI endonuclease
activity is blocked by this top strand adenine methylation and is
unaffected by methylation of the adenine in the complementary strand,
5'-GTYGGA-3'. There is no additional DNA modification associated with the
MmeI R-M system, as is required for previously characterized Type IIG R-M
systems. The MmeI R-M system thus uses modification on only one of the two
DNA strands for host protection. The MmeI architecture represents a
minimal approach to assembling a restriction-modification system wherein a
single DNA recognition domain targets both the endonuclease and DNA
methyltransferase activities.

<>

<1>Morgan, R.D., Calvet, C., Demeter, M., Agra, R., Kong, H.M.
<2>Characterization of the specific DNA nicking activity of restriction endonuclease N.BstNBI.
<3>Biol. Chem.
<4>381
<5>1123-1125
<6>2000
<7>N.BstNBI is a unique restriction endonuclease isolated from Bacillus stearothermophilus, We
have characterized the recognition sequence and
the cleavage site of N.BstNBI. Mapping of cleavage sites of N.BstNBI
showed that it recognizes an asymmetric sequence, 5' GAGTC 3', and
cleaves only on the top strand 4 base pairs away from its recognition
sequence. To verify the nicking activity of N.BstNBI, we have
constructed two plasmids containing a single recognition sequence
(pNB1) or no recognition site (pNB0). When pNB1 and pNB0 were incubated
with the enzyme, N.BstNBI nicked only the plasmid pNB1, suggesting that
N.BstNBI is a specific nicking endonuclease.

<>

<1>Morgan, R.D., Camp, R.R., Wilson, G.G., Xu, S.-Y.
<2>Molecular cloning and expression of NlaIII restriction-modification system in E. coli.
<3>Gene
<4>183
<5>215-218
<6>1996
<7>The NlaIII restriction enzyme isolated from Neisseria lactamica recognizes the sequence
5'-CATG-3', cleaving after the G to generate a four base 3' overhang.  The NlaIII methylase
and a portion of the NlaIII endonuclease gene were cloned into E. coli by the methylase
selection method, and the remaining portion of the NlaIII endonuclease gene was cloned by
inverse PCR.  The nucleotide sequence of the endonuclease gene and the methylase gene were
determined.  The NlaIII endonuclease gene is 693 bp, encoding a protein with a predicted
molecular weight of 26,487.  The NlaIII methylase gene was identical with that previously
reported [Labbe, D., Joltke, H.J. and Lau, P.C. (1990)  Cloning and characterization of two
tandemly arranged DNA methyltransferase genes of Neisseria lactamica: an adenine-specific
M.NlaIII and a cytosine-type methylase.  Mol. Gen. genet. 224, 101-110].  The endonuclease and
methylase genes overlap by four bases and are transcribed in the same orientation.  The
endonuclease gene was cloned into an improved T7 vector, and a high level of NlaIII
endonuclease expression was achieved in E. coli.

<>

<1>Morgan, R.D., Chang, Z.
<2>Discovery of and method for cloning and producing the PspGI restriction endonuclease.
<3>US Patent Office
<4>US 5849558
<5>
<6>1998
<7>The present invention relates to the type II restriction endonuclease PspGI, obtainable from
Pyrococcus species G-I-H (NEB #906), an enzyme which recognizes the DNA sequence 5' CC(A/T)GG
3' and cleaves before the first C of the recognition sequence to produce a 5 base 5'
extension:  5'-/CC(A/T)GG-3' 3'-GG(T/A)CC/-5' (wherein G represents guanine, C represents
cytosine, A represents adenine and T represents thymine and (A/T) represents either A or T in
that one position).

<>

<1>Morgan, R.D., Chang, Z.
<2>Discovery of and method for cloning and producing the PspGI restriction endonuclease.
<3>International Patent Office
<4>WO 9851783
<5>
<6>1998
<7>The present invention relates to the type II restriction endonuclease PspGI, obtainable from
Pyrococcus species G-I-H (NEB #906), an enzyme which recognizes the DNA sequence
5'CC(A/T)GG3' and cleaves before the first C of the recognition sequence to produce a 5 base
5' extension: 5'-/CC(A/T)GG-3', 3'-GG(T/A)CC/-5', (wherein G represents guanine, C
represents cytosine, A represents adenine and T represents thymine and (A/T) represents either
A or T in that one position).

<>

<1>Morgan, R.D., Chang, Z., Mersha, F.B.
<2>Method for cloning and producing the SpeI restriction endonuclease.
<3>European Patent Office
<4>EP 0869174 A
<5>
<6>1998
<7>The present invention relates to the recombinant DNA which encodes the SpeI restriction
endonuclease and modification methylase, and the production of SpeI restriction endonuclease
from the recombinant DNA.

<>

<1>Morgan, R.D., Chang, Z., Mersha, F.B.
<2>Method for cloning and producing the SpeI restriction endonuclease.
<3>US Patent Office
<4>US 5945326
<5>
<6>1999
<7>The present invention relates to the recombinant DNA which encodes the SpeI restriction
endonuclease and modification methylase, and the production of SpeI restriction endonuclease
from the recombinant DNA.

<>

<1>Morgan, R.D., Dalton, M., Stote, R.
<2>A unique type II restriction endonuclease from Acinetobacter lwoffi N.
<3>Nucleic Acids Res.
<4>15
<5>7201
<6>1987
<7>A new type II restriction endonuclease, AlwN I, has been isolated from
Acinetobacter lwoffi N (NEB 419).  AlwN I recognizes the interrupted
palindromic sequence 5' CAGNNNCTG 3' and cleaves between the 3' N and C to
produce a 3 base 3' extension.

<>

<1>Morgan, R.D., Dwinell, E.A., Bhatia, T.K., Lang, E.M., Luyten, Y.A.
<2>The MmeI family: type II restriction-modification enzymes that employ single-strand modification for host protection.
<3>Nucleic Acids Res.
<4>37
<5>5208-5221
<6>2009
<7>The type II restriction endonucleases form one of the largest families of
biochemically-characterized proteins. These endonucleases typically share
little sequence similarity, except among isoschizomers that recognize the
same sequence. MmeI is an unusual type II restriction endonuclease that
combines endonuclease and methyltransferase activities in a single
polypeptide. MmeI cuts DNA 20 bases from its recognition sequence and
modifies just one DNA strand for host protection. Using MmeI as query we
have identified numerous putative genes highly similar to MmeI in database
sequences. We have cloned and characterized 20 of these MmeI homologs.
Each cuts DNA at the same distance as MmeI and each modifies a conserved
adenine on only one DNA strand for host protection. However each enzyme
recognizes a unique DNA sequence, suggesting these enzymes are undergoing
rapid evolution of DNA specificity. The MmeI family thus provides a rich
source of novel endonucleases while affording an opportunity to observe
the evolution of DNA specificity. Because the MmeI family enzymes employ
modification of only one DNA strand for host protection, unlike previously
described type II systems, we propose that such single-strand modification
systems be classified as a new subgroup, the type IIL enzymes, for Lone
strand DNA modification.

<>

<1>Morgan, R.D., Luyten, Y.A.
<2>Rational engineering of type II restriction endonuclease DNA binding and cleavage specificity.
<3>Nucleic Acids Res.
<4>37
<5>5222-5233
<6>2009
<7>The type II restriction endonucleases are indispensible tools for molecular biology. Although
enzymes recognizing nearly 300 unique
sequences are known, the ability to engineer enzymes to recognize any
sequence of choice would be valuable. However, previous attempts to
engineer new recognition specificity have met limited success. Here we
report the rational engineering of multiple new type II specificities. We
recently identified a family of MmeI-like type II endonucleases that have
highly similar protein sequences but different recognition specificity. We
identified the amino-acid positions within these enzymes that determine
position specific DNA base recognition at three positions within their
recognition sequences through correlations between their aligned
amino-acid residues and aligned recognition sequences. We then altered the
amino acids at the identified positions to those correlated with
recognition of a desired new base to create enzymes that recognize and cut
at predictable new DNA sequences. The enzymes so altered have similar
levels of endonuclease activity compared to the wild-type enzymes. Using
simple and predictable mutagenesis in this family it is now possible to
create hundreds of unique new type II restriction endonuclease
specificities. The findings suggest a simple mechanism for the evolution
of new DNA specificity in Nature.

<>

<1>Morgan, R.D., Roberts, R.J.
<2>Novel restriction endonucleases, DNA encoding these endonucleases and methods for identifying new endonucleases with the same or varied specificity.
<3>International Patent Office
<4>WO 200797778 A
<5>
<6>2007
<7>Specified restriction endonucleases have been characterized for the first time by their amino
acid and DNA sequences.  These sequences and those with at least 90% identity thereto have
been used as probes in sequence similarity analyses to identify sequence matches in a sequence
database that corresponds to novel restriction endonucleases or isoschizomers.  The sequence
similarity analyses includes selecting a positive sequence match from any sequence producing
an expectation value of less than or equal to e-02.

<>

<1>Morgan, R.D., Roberts, R.J.
<2>Restriction endonucleases, DNA encoding these endonucleases and methods for indentifying new endonucleases with the same or varied specificity.
<3>US Patent Office
<4>US 8685689 B
<5>
<6>2014
<7>Specified restriction endonucleases have been characterized for the first time by their amino
acid and DNA sequences. These sequences and those with at least 90% identity thereto have been
used as probes in sequence similarity analyses to identify sequence matches in a sequence
database that corresponds to novel restriction endonucleases or isoschizomers. The sequence
similarity analyses includes selecting a positive sequence match from any sequence producing
an expectation value of less than or equal to e-02.

<>

<1>Morgan, R.D., Roberts, R.J.
<2>Restriction endonucleases, DNA encoding these endonucleases and methods for identifying new endonucleases with the same or varied specificity.
<3>US Patent Office
<4>US 08227231
<5>
<6>2012
<7>Specified restriction endonucleases have been characterized for the first time by their amino
acid and DNA sequences. These sequences and
those with at least 90% identity thereto have been used as probes in
sequence similarity analyses to identify sequence matches in a sequence
database that corresponds to novel restriction endonucleases or
isoschizomers. The sequence similarity analyses includes selecting a
positive sequence match from any sequence producing an expectation
value of less than or equal to e-02.

<>

<1>Morgan, R.D., Walsh, P.
<2>New substantially pure Type II restriction endonuclease obtainable from Bacillus thermoglucosidasius 36A, useful for cleaving DNA at new  positions within the DNA molecule - isolation and purification of a  restriction endonuclease from Bacillus thermogl.
<3>US Patent Office
<4>US 20050233432
<5>
<6>2005
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
obtainable from Bacillus thermoglucosidasius 36A (NEB#1384), hereinafter referred to as BtgZI,
which endonuclease: (1) recognizes the nucleotide sequence 5'-GCGATG-3' in a double-stranded
DNA molecule as shown below, 5'-GCGATGNNNNNNNNNN/-3' (SEQ ID NO:9)
3'-CGCTACNNNNNNNNNNNNNN/-5' (wherein G represents guanine, C represents cytosine, A
represents adenine, T represents thymine and N represents either G, C, A, or T); (2) cleaves
said sequence in the phosphodiester bonds between the 10th and the 11th nucleotides 3' to the
recognition sequence in the 5'-GCGATG-3 strand of the DNA, and between the 14th and 15th
nucleotides 5' to the recognition sequence in the complement stand, 5'-CATCGC-3', to
produce a 4 base 5' extension; and (3) cleaves double-stranded pBR322 DNA to produce 3
fragments of 2892, 1181 and 288 base pairs.

<>

<1>Morgan, R.D., Walsh, P.
<2>Type II restriction endonuclease BtgZI, obtainable from Bacillus thermoglucosidasius 36A and a process for producing the same.
<3>US Patent Office
<4>US 7029900 B
<5>
<6>2006
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
obtainable from Bacillus thermoglucosidasius 36A (NEB#1384), hereinafter referred to as
"BtgZI", which endonuclease: (1) recognizes the nucleotide sequence 5'-GCGATG-3' in a
double-stranded DNA molecule as shown below.  5'-GCGATGNNNNNNNNNN/-3' (SEQ ID NO: 9)
3'-CGCTACNNNNNNNNNNNNNN/-5' (wherein G represents guanine, C represents cytosine, A
represents adenine, T represents thymine and N represents either G, C, A, or T); (2) cleaves
said sequence in the phosphodiester bonds between the 10th and the 11th nucleotides 3' to the
recognition sequence in the 5'-GCGATG-3 strand of the DNA, and between the 14th and 15th
nucleotides 5' to the recognition sequence in the complement stand, 5'-CATCGC-3', to
produce a 4 base 5' extension; and (3) cleaves double-stranded pBR322 DNA to produce 3
fragments of 2892, 1181 and 288 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>Type II restriction endonuclease, HpyCH4V, obtainable from Helicobacter pylori CH4 and a process for producing the same.
<3>US Patent Office
<4>US 6133009
<5>
<6>2000
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori CH4 (NEB#1236), hereinafter referred to as
"HpyCH4V", which endonuclease: (1) recognizes the nucleotide sequence 5'-TGCA-3' in a
double-stranded DNA molecule as shown below,
5'-TC/CA-3'
3'-AC/GT-5'
(wherein G represents guanine, C represents cytosine, A represents adenine, T represents
thymine and N represents either G, C, A, or T); (2) cleaves said sequence in the
phosphodiester bonds between the G and C as indicated with the arrows; and (3) cleaves
double-stranded pBR322 DNA to produce 21 fragments, including fragments of 576, 498, 441, 335,
315, 312, 296, 244 and 205 base pairs, and 12 fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>A novel type II restriction endonuclease, HpyCH4III, obtainable from Helicobacter pylori CH4 and a process for producing the same.
<3>International Patent Office
<4>WO 0121777 A
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori CH4 (NEB#1236), hereinafter referred to as
"HpyCH4III", which endonuclease: (1) recognizes the nucleotide sequence 5'-ACNGT-3' in a
double-stranded DNA molecule: 5'-CAN/GT-3' and 3'-TG/NCA-5' (wherein G represents guanine,
C represents cytosine, A represents adenine, T represents thymine and N represents either G,
C, A, or T); (2) cleaves said sequence in the phosphodiester bonds between the N and G as
indicated with the arrows; and (3) cleaves double-stranded PhiX174 DNA to produce 15
fragments, including fragments of 1284, 814, 536, 517, 454, 404, 302, 292, 270 and 222 base
pairs, and 5 fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>Type II restriction endonuclease, Hpy188III, obtainable from Helicobacter pylori J188 and a process for producing the same.
<3>US Patent Office
<4>US 6238901 B
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori J188 (NEB#1174), hereinafter referred to as
"Hpy188III", which endonuclease:  (1) recognizes the nucleotide sequence 5'-TCNNGA-3' in a
double-stranded DNA molecule as shown below, 5'-TC/NNGA-3' 3'-AGNN/CT-5' (wherein G
represents guanine, C represents cytosine, A represents adenine, T represents thymine and N
represents either G, C, A, or T); (2) cleaves said sequence in the phosphodiester bonds
between the C and first unspecified nucleotide N as indicated with the arrows; and (3) cleaves
double-stranded PhiX174 DNA to produce 26 fragments, including fragments of 704, 560, 497,
477, 423, 309, 308, 304, 261, 225 base pairs, and 16 fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>Type II restriction endonuclease, HpyCH4III, obtainable from Helicobacter pylori CH4 and a process for producing the same.
<3>US Patent Office
<4>US 6238904 B
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori CH4 (NEB#1236), hereinafter referred to as
"HpyCH4III", which endonuclease:  (1) recognizes the nucleotide sequence 5'-ACNGT-3' in a
double-stranded DNA molecule as shown below, 5'-Can/GT-3' 3'-TG/NCA-5' (wherein G
represents guanine, C represents cytosine, A represents adenine, T represents thymine and N
represents either G, C, A, or T); (2) cleaves said sequence in the phosphodiester bonds
between the N and G as indicated with the arrows; and (3) cleaves double-stranded PhiX174 DNA
to produce 15 fragments, including fragments of 1284, 814, 536, 517, 454, 404, 302, 292, 270
and 222 base pairs, and 5 fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>A novel type II restriction endonuclease, Hpy188I, obtainable from Helicobacter pylori J188 and a process for producing the same.
<3>International Patent Office
<4>WO 0118186 A
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori J188 (NEB#1174), hereinafter referred to as
"Hpy188I", which endonuclease: (1) recognizes the nucleotide sequence 5'-TCNGA-3' in a
double-stranded DNA molecule as shown below, 5'-TCN/GA-3' 3'-AG/NCT-3' (wherein G
represents guanine, C represents cytosine, A represents adenine, T represents thymine and N
represents either G, C, A; or T); (2) cleaves said sequence in the phosphodiester bonds
between the N and G as indicated with the arrows; and (3) cleaves double-stranded PhiX174 DNA
to produce 18 fragments, including fragments of 1331, 813, 572, 525, 396, 302, 293, 219 base
pairs, and 10 fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>A type II restriction endonuclease from Helicobacter pylori CH4.
<3>International Patent Office
<4>WO 0121638 A
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori CH4 (NEB#1236), hereinafter referred to as
"HpyCH4IV", which endonuclease: (1) recognizes the nucleotide sequence 5'-ACGT-3' in a
double-stranded DNA molecule: 5'-A/CGT-3' 3'-TGC/A-5' (wherein G represents guanine, C
represents cytosine, A represents adenine, T represents thymine and N represents either G, C,
A, or T): (2) cleaves said sequence in the phosphodiester bonds between the A and C as
indicated with the arrows; and (3) cleaves double-stranded PhiX174 DNA to produce 19
fragments, including fragments of 1036, 749, 397, 379, 371, 365, 362, 343, 282, 282, 268 base
pairs, and 8 fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>A novel type II restriction endonuclease, HpyCH4V, obtainable from Helicobacter pylori CH4 and a process for producing the same.
<3>International Patent Office
<4>WO 0121776 A
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori CH4 (NEB#1236), hereinafter referred to as
"HpyCH4V", which endonuclease: (1) recognizes the nucleotide sequence 5'-TGCA-3' in a
double-stranded DNA molecule: 5'-TG/CA-3'; 3'-AC/GT-3' (wherein G represents guanine, C
represents cytosine, A represents adenine, T represents thymine and N represents either G, C,
A, or T); (2) cleaves said sequence in the phosphodiester bonds between the G and C as
indicated with the arrows; and (3) cleaves double-stranded pBR322 DNA to produce 21 fragments,
including fragments of 576, 498, 441, 335, 315, 312, 296, 244 and 205 base pairs, and 12
fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>A novel type II restriction endonuclease, Hpy99I, obtainable from Helicobacter pylori J99 and a process for producing the same.
<3>International Patent Office
<4>WO 0121778 A
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori (NEB#1237), hereinafter referred to as
"Hpy99I", which endonuclease: (1) recognizes the nucleotide sequence 5'-CGWCG-3' in a
double-stranded DNA (wherein G represents guanine, C represents cytosine, A represents
adenine, T represents thymine and W represents either A, or T); (2) cleaves double-stranded
PhiX174 DNA to produce 8 fragments, including fragments of 3063, 629, 602, 447, 389, and 176
base pairs, and 2 fragments smaller than 100 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>Type II restriction endonuclease, Hpy188I, obtainable from Helicobacter pylori J188 and a process for producing the same.
<3>US Patent Office
<4>US 6258583
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter
pylori J188 (NEB#1174), hereinafter referred to as "Hpy188I", which
endonuclease: (1) recognizes the nucleotide sequence 5'-TCNGA-3' in a
double-stranded DNA molecule as shown below, 5'-TCN|GA-3'
3'-AG^NCT-5' (wherein G represents guanine, C represents
cytosine, A represents adenine, T represents thymine and N represents
either G, C, A, or T); (2) cleaves said sequence in the phosphodiester
bonds between the N and G as indicated with the arrows; and (3) cleaves
double-stranded PhiX174 DNA to produce 18 fragments, including
fragments of 1331, 813, 572, 525, 396, 302, 293, 219 base pairs, and 10
fragments smaller than 200 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>Type II restriction endonuclease, Hpy99I, obtainable from Helicobacter pylori J99 and a process for producing the same.
<3>US Patent Office
<4>US 6280992 B
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori J99 (NEB#1237), hereinafter referred to as
"Hpy99I", which endonuclease: (1) recognizes the nucleotide sequence 5'-CGWCG-3' in a
double-stranded DNA (wherein G represents guanine, C represents cytosine, A represents
adenine, T represents thymine and W represents either A, or T); (2) cleaves double-stranded
PhiX174 DNA to produce 8 fragments, including fragments of 3063, 629, 602, 447, 389, and 176
base pairs, and 2 fragments smaller than 100 base pairs.

<>

<1>Morgan, R.D., Xu, Q.
<2>Type II restriction endonuclease, HpyCH4IV, obtainable from Helicobacter pylori CH4 and a process for producing the same.
<3>US Patent Office
<4>US 6194188 B
<5>
<6>2001
<7>In accordance with the present invention, there is provided a novel restriction endonuclease
and its DNA obtainable from Helicobacter pylori CH4 (NEB#1236), hereinafter referred to as
"HpyCH4IV", which endonuclease: (1) recognizes the nucleotide sequence 5'-ACGT-3'  in a
double-stranded DNA molecule as shown below,
5'-A/CGT-3'
3'-TGC/A-5'
(wherein G represents guanine, C represents cytosine, A represents adenine, T represents
thymine and N represents either G, C, A, or T0: (2) cleaves said sequence in the
phosphodiester bonds between the A and C as indicated with the arrows; and (3) cleaves
double-stranded PhiX174 DNA to produce 19 fragments, including fragments of 1036, 749, 397,
379, 371, 365, 362, 343, 282, 282, 268 base pairs, and 8 fragments smaller than 200 base
pairs.

<>

<1>Morgan, W.F., Fero, M.L., Land, M.C., Winegar, R.A.
<2>Inducible expression and cytogenic effects of the EcoRI restriction endonuclease in chinese hamster ovary cells.
<3>Mol. Cell. Biol.
<4>8
<5>4204-4211
<6>1988
<7>The cytogenic endpoints of sister chromatid exchange (SCE) and chromosome
aberrations are widely used as indicators of DNA damage induced by mutagenic
carcinogens.  Chromosome aberrations appear to result directly from DNA
double-strand breaks, but the lesion(s) giving rise to SCE formation remains
unknown.  Most compounds that induce SCEs induce a spectrum of lesions in DNA.
To investigate the role of double-strand breakage in SCE formation, we
constructed a plasmid that gives rise to one specific lesion, a staggered-end
("cohesive") DNA double-strand break.  This plasmid, designated pMENs, contains
a selectable marker, neo, which is a bacterial gene for neomycin resistance,
and the coding sequence for the bacterial restriction endonuclease EcoRI
attached to the mouse metallothionein gene promoter.  EcoRI recognizes G^AATTC
sequences in DNA and makes DNA double-strand breaks with four nucleotides
overhanging as staggered ends.  Cells transfected with pMENS were resistant to
the antibiotic G418 and contained an integrated copy of the EcoRI gene,
detectable by DNA filter hybridization.  The addition of the heavy metal CdSO4
resulted in the intracellular production of EcoRI, as measured by an anti-EcoRI
antibody.  Cytogenetic analysis after the addition of CdSO4 indicated a
dramatic increase in the frequency of chromosome aberrations but very little
effect on SCE frequency.  Although there was some intercellular heterogeneity,
these results confirm that DNA double-strand breaks do result in chromosome
aberrations but that these breaks are not sufficient to give rise to SCE
formation.

<>

<1>Mori, K., Yamazoe, A., Hosoyama, A., Ohji, S., Fujita, N., Ishibashi, J., Kimura, H., Suzuki, K.
<2>Thermotoga profunda sp. nov. and Thermotoga caldifontis sp. nov., anaerobic thermophilic bacteria isolated from terrestrial hot springs.
<3>Int. J. Syst. Evol. Microbiol.
<4>64
<5>2128-2136
<6>2014
<7>Two thermophilic, strictly anaerobic, Gram-negative bacteria, designated strains
AZM34c06(T) and AZM44c09(T), were isolated from terrestrial hot springs in Japan.
The optimum growth conditions for strain AZM34c06(T) were 60 degrees C, pH 7.4 and 0 %
additional NaCl, and those for strain AZM44c09(T) were 70 degrees C, pH
7.4 and 0 % additional NaCl. Complete genome sequencing was performed for both strains,
revealing genome sizes of 2.19 Mbp (AZM34c06(T)) and 2.01 Mbp (AZM44c09(T)). Phylogenetic
analyses based on 16S rRNA gene sequences and the concatenated predicted amino acid sequences
of 33 ribosomal proteins showed that both strains belonged to the genus Thermotoga. The
closest relatives of strains
AZM34c06(T) and AZM44c09(T) were the type strains of Thermotoga lettingae (96.0 % similarity
based on the 16S rRNA gene and 84.1 % similarity based on ribosomal
proteins) and Thermotoga hypogea (98.6 and 92.7 % similarity), respectively.
Using blast, the average nucleotide identity was 70.4-70.5 % when comparing strain AZM34c06(T)
and T. lettingae TMO(T) and 76.6 % when comparing strain
AZM44c09(T) and T. hypogea NBRC 106472(T). Both values are far below the 95 % threshold value
for species delineation. In view of these data, we propose the inclusion of the two isolates
in the genus Thermotoga within two novel species, Thermotoga profunda sp. nov. (type strain
AZM34c06(T) = NBRC 106115(T) = DSM
23275(T)) and Thermotoga caldifontis sp. nov. (type strain AZM44c09(T) = NBRC
106116(T) = DSM 23272(T)).

<>

<1>Mori, T., Takahashi, M., Tanaka, R., Shibata, T., Kuroda, K., Ueda, M., Takeyama, H.
<2>Draft Genome Sequence of Falsirhodobacter sp. Strain alg1, an Alginate-Degrading  Bacterium Isolated from Fermented Brown Algae.
<3>Genome Announcements
<4>2
<5>e00826-14
<6>2014
<7>Falsirhodobacter sp. alg1 is an alginate-degrading bacterium, the second species  from the
nonphototrophic bacterial genus Falsirhodobacter. We report the first
draft genome of a bacterium from this genus and point out possible important
features related to alginate assimilation and its evolutionary aspects.

<>

<1>Moriel, D.G. et al.
<2>Identification of protective and broadly conserved vaccine antigens from the genome of Extraintestinal Pathogenic Escherichia coli (ExPEC).
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>9072-9077
<6>2010
<7>Extraintestinal pathogenic Escherichia coli (ExPEC) are a common cause of disease in both
mammals and birds. A vaccine to prevent such infections would be desirable given the
increasing antibiotic resistance of these bacteria. We have determined the genome
sequence of ExPEC IHE3034 (ST95) isolated from a case of neonatal
meningitis and compared this to available genome sequences of
other ExPEC strains and a few nonpathogenic E. coli. We found
19 genomic islands present in the genome of IHE3034, which are
absent in the nonpathogenic E. coli isolates. By using subtractive
reverse vaccinology we identified 230 antigens present in ExPEC
but absent (or present with low similarity) in nonpathogenic
strains. Nine antigens were protective in a mouse challenge model.
Some of them were also present in other pathogenic non-ExPEC
strains, suggesting that a broadly protective E. coli vaccine may
be possible. The gene encoding the most protective antigen was
detected in most of the E. coli isolates, highly conserved in
sequence and found to be exported by a type II secretion system
which seems to be nonfunctional in nonpathogenic strains.

<>

<1>Morikawa, K., Shirakawa, M.
<2>Three-dimensional structural views of damaged-DNA recognition: T4 endonuclease V, E. coli Vsr protein, and human nucleotide excision repair factor XPA.
<3>Mutat. Res.
<4>460
<5>257-275
<6>2000
<7>Genetic information is frequently disturbed by introduction of modified or mismatch bases into
duplex DNA, and hence all organisms contain DNA repair systems to restore normal genetic
information by removing such damaged bases or nucleotides and replacing them by correct ones.
The understanding of this repair mechanism is a central subject in cell biology. This review
focuses on the three-dimensional structural views of damaged DNA recognition by three
proteins. The first protein is T4 endonuclease V (T4 endo V), which catalyzes the first
reaction step of the excision repair pathway to remove pyrimidine-dimers (PD) produced within
duplex DNA by UV irradiation. The crystal structure of this enzyme complexed with DNA
containing a thymidine-dimer provided the first direct view of DNA lesion recognition by a
repair enzyme, indicating that the DNA kink coupled with base flipping-out is important for
damaged DNA recognition. The second is very short patch repair (Vsr) endonuclease, which
recognizes a TG mismatch within the five base pair consensus sequence. The crystal structure
of this enzyme in complex with duplex DNA containing a TG mismatch revealed a novel mismatch
base pair recognition scheme, where three aromatic residues intercalate from the major groove
into the DNA to strikingly deform the base pair stacking but the base flipping-out does not
occur. The third is human nucleotide excision repair (NER) factor XPA, which is a major
component of a large protein complex. This protein has been shown to bind preferentially to
UV- or chemical carcinogen-damaged DNA. The solution structure of the XPA central domain,
essential for the interaction of damaged DNA, was determined by NMR. This domain was found to
be divided mainly into a (Cys)4-type zinc-finger motif subdomain for replication protein A
(RPA) recognition and the carboxyl terminal subdomain responsible for DNA binding.

<>

<1>Morishima, N., Mizumura, H., Shibata, T.
<2>Endonuclease.
<3>US Patent Office
<4>US 6528296 A
<5>
<6>2003
<7>The present invention relates to a site-specific endonuclease which recognizes a specific
nucleotide sequence, to a gene coding for the endonuclease, to a recombinant vector containing
the gene, to a transformant containing the vector, and to a process for producing the
endonuclease.

<>

<1>Morishima, N., Nakagawa, K.-I., Shibata, T.
<2>A sequence-specific endonuclease, Endo.SceI, can efficiently induce gene conversion in yeast mitochondria lacking a major exonuclease.
<3>Curr. Genet.
<4>23
<5>537-541
<6>1993
<7>Endo.SceI most likely initiates homologous recombination of the yeast mitochondrial genome
through sequence-specific double-strand scission of DNA.  According to the double-strand
break-repair model for the mechanism of homologous recombination, DNA ends created by
sequence-specific endonucleases have to be processed by exonucleases.  The major mitochondrial
exonuclease (encoded by NUC1) has been shown to greatly affect the length of conversion tracts
at the 21S rRNA locus when site-specific gene conversion is induced by omega endonuclease.  In
order to examine the role of the NUC1 nuclease in the Endo.SceI-induced recombination,
recombination frequencies were measured after crossing of parental strains either in the
presence or absence of NUC1 nuclease activity.  The frequency of gene conversion in the oli2
locus induced by Endo.SceI was not reduced by disruption of the NUC1 gene.  This result
strongly implicates the presence of multiple exonucleases for the processing of the DNA ends
created by sequence-specific endonucleases.

<>

<1>Morishima, N., Nakagawa, K.-I., Yamamoto, E., Shibata, T.
<2>A subunit of yeast site-specific endonuclease SceI is a mitochondrial version of the 70-kDa heat shock protein.
<3>J. Biol. Chem.
<4>265
<5>15189-15197
<6>1990
<7>Endo.SceI of Saccharomyces cerevisiae is a heterodimeric site-specific
endonuclease, which is distinguishable from prokaryotic restriction
endonucleases in the mode of recognition of its cleavage site.  We have used
monoclonal antibodies specific to the larger subunit (75 kDa) of Endo.SceI to
isolate the gene for the subunit (ENS1) from S. cerevisiae.  Unexpectedly ENS1
was found to encode a 70-kDa heat shock protein-related polypeptide and to be
identical to recently cloned SSC1.  Subcellular fractionation experiments on
yeast cells revealed that the primary target site of the larger subunit is
mitochondria, where almost all the Endo.SceI activity is localized.  Molecular
genetic analysis of ENS1 demonstrated its indispensability for growth and the
requirement of a high level of its expression at the sporulation and
germination stages.  The data suggest that ENS1 plays an important role,
especially at these differentiation stages.

<>

<1>Morishima, N., Shibata, T.
<2>SceI: an endonuclease with multiple cutting sites induces homologous genetic recombination.
<3>Seikagaku
<4>64
<5>1420-1431
<6>1992
<7>A review.

<>

<1>Morishima, N., Shibata, T.
<2>Sequence-specific endonucleases involved in genetic recombination.
<3>Tanpakushitsu Kakusan Koso
<4>36
<5>1716-1720
<6>1991
<7>
<>

<1>Morishima, N., Shibata, T., Mizumura, H.
<2>Endonuclease.
<3>European Patent Office
<4>EP 0972836 A
<5>
<6>2000
<7>The present invention relates to a site-specific endonuclease which recognizes a specific
nucleotide sequence, to a gene coding for the endonuclease, to a recombinant vector containing
the gene, to a transformant containing the vector, and to a process for producing the
endonuclease.

<>

<1>Morita, H., Toh, H., Fukuda, S., Horikawa, H., Oshima, K., Suzuki, T., Murakami, M., Hisamatsu, S., Kato, Y., Takizawa, T., Fukuoka, H., Yoshimura, T., Itoh, K., O'Sullivan, D.J., McKay, L.L., Ohno, H., Kikuchi, J., Masaoka, T., Hattori, M.
<2>Comparative Genome Analysis of Lactobacillus reuteri and Lactobacillus fermentum Reveal a Genomic Island for Reuterin and Cobalamin Production.
<3>DNA Res.
<4>15
<5>151-161
<6>2008
<7>Lactobacillus reuteri is a heterofermentative lactic acid bacterium that naturally inhabits
the gut of humans and other animals. The probiotic
effects of L. reuteri have been proposed to be largely associated with the
production of the broad-spectrum antimicrobial compound reuterin during
anaerobic metabolism of glycerol. We determined the complete genome
sequences of the reuterin-producing L. reuteri JCM 1112(T) and its closely
related species Lactobacillus fermentum IFO 3956. Both are in the same
phylogenetic group within the genus Lactobacillus. Comparative genome
analysis revealed that L. reuteri JCM 1112(T) has a unique cluster of 58
genes for the biosynthesis of reuterin and cobalamin (vitamin B(12)). The
58-gene cluster has a lower GC content and is apparently inserted into the
conserved region, suggesting that the cluster represents a genomic island
acquired from an anomalous source. Two-dimensional nuclear magnetic
resonance (2D-NMR) with (13)C(3)-glycerol demonstrated that L. reuteri JCM
1112(T) could convert glycerol to reuterin in vivo, substantiating the
potential of L. reuteri JCM 1112(T) to produce reuterin in the intestine.
Given that glycerol is shown to be naturally present in feces, the
acquired ability to produce reuterin and cobalamin is an adaptive
evolutionary response that likely contributes to the probiotic properties
of L. reuteri.

<>

<1>Morita, H., Toh, H., Oshima, K., Murakami, M., Taylor, T.D., Igimi, S., Hattori, M.
<2>Complete genome sequence of probiotic Lactobacillus rhamnosus ATCC 53103.
<3>J. Bacteriol.
<4>191
<5>7630-7631
<6>2009
<7>Lactobacillus rhamnosus is a facultatively heterofermentative lactic acid bacterium and is
frequently isolated from human gastrointestinal mucosa of healthy individuals. L. rhamnosus
ATCC 53103 isolated from a healthy human intestinal flora is one of the most widely used and
well-documented probiotics. Here we report the finished and annotated genome sequence of this
organism.

<>

<1>Morita, H., Toh, H., Oshima, K., Yoshizaki, M., Kawanishi, M., Nakaya, K., Suzuki, T., Miyauchi, E., Ishii, Y., Tanabe, S., Murakami, M., Hattori, M.
<2>Complete Genome Sequence and Comparative Analysis of the Fish Pathogen Lactococcus garvieae.
<3>PLoS ONE
<4>6
<5>E23184
<6>2011
<7>Lactococcus garvieae causes fatal haemorrhagic septicaemia in fish such as
yellowtail. The comparative analysis of genomes of a virulent strain Lg2
and a non-virulent strain ATCC 49156 of L. garvieae revealed that the two
strains shared a high degree of sequence identity, but Lg2 had a 16.5-kb
capsule gene cluster that is absent in ATCC 49156. The capsule gene
cluster was composed of 15 genes, of which eight genes are highly
conserved with those in exopolysaccharide biosynthesis gene cluster often
found in Lactococcus lactis strains. Sequence analysis of the capsule gene
cluster in the less virulent strain L. garvieae Lg2-S, Lg2-derived strain,
showed that two conserved genes were disrupted by a single base pair
deletion, respectively. These results strongly suggest that the capsule is
crucial for virulence of Lg2. The capsule gene cluster of Lg2 may be a
genomic island from several features such as the presence of insertion
sequences flanked on both ends, different GC content from the chromosomal
average, integration into the locus syntenic to other lactococcal genome
sequences, and distribution in human gut microbiomes. The analysis also
predicted other potential virulence factors such as haemolysin. The
present study provides new insights into understanding of the virulence
mechanisms of L. garvieae in fish.

<>

<1>Morita, R., Ishikawa, H., Nakagawa, N., Kuramitsu, S., Masui, R.
<2>Crystal structure of a putative DNA methylase TTHA0409 from Thermus thermophilus HB8.
<3>Proteins
<4>73
<5>259-264
<6>2008
<7>To protect the cell from exogenous DNA, most species have DNA modification methylase.  It is
also reported that DNA methylation is essentially involved in bacterial virulence.  Many
restriction-modification systems have been researched, and several three-dimensional
structures of R-M enzymes have been determined.  DNA modification methylases catalyze the
transfer of a methyl group to DNA from S-adenosyl-L-methionine, which is consequently
converted to S-adenosyl-L-homocysteine.  These enzymes are categorized into the following two
classes based on the position of the transferred methyl group on the DNA bases: endocyclic
MTases and exocyclic amino MTases.  Endocyclic MTases methylate the C5 position of a cytosine
base, whereas exocyclic amino MTases methylate the N6 position of an adenine base or N4
position of a cytosine base.  Furtermore, the exocyclic amino MTases are subdivided into six
classes, namely, alpha, beta, gamma, delta and epsilon, based on their amino acid sequences.

<>

<1>Moriya, S., Yanagawa, S., Aoki, N., Iwabuchi, M., Inoue, T., Ando, T.
<2>Isolation and characterization of a restriction enzyme BspO4I from an alkalophilic bacterium.
<3>Nucleic Acids Res.
<4>20
<5>3781
<6>1992
<7>A type II restriction enzyme, BspO4I, has been isolated from Bacillus sp 0-4 (ATCC 21536), an
alkalophilic bacterium. The enzyme was an isoschizomer of PvuII, recognizing the six base
palindromic sequence of 5'CAGCTG3' and cleaves between G and C residues to produce
blunt-ended cleavage products. Bsp04I was partially purified from a cell-free extract using
column chromatography on DEAE-cellulose, heparin-cellulofine and phosphocellulose. Optimal
conditions for the enzyme activity were pH 8.5, 20 mM MgCl2, 250 mM monovalent cation (NaCl or
KCl), 10 mM 2-mercaptoethanol at 40oC. The cleavage site of BspO4I was determined by primer
extension experiments and indicated that BspO4I recognizes and cleaves the following sequence:
5'CAG^CTG3'
3'GTC^GAC5'.

<>

<1>Morohoshi, T., Kato, T., Someya, N., Ikeda, T.
<2>Complete Genome Sequence of N-Acylhomoserine Lactone-Producing Pseudomonas sp. Strain StFLB209, Isolated from Potato Phyllosphere.
<3>Genome Announcements
<4>2
<5>e01037-14
<6>2014
<7>Pseudomonas sp. strain StFLB209 is isolated from the potato leaf and produces N-acylhomoserine
lactone quorum-sensing signal compounds. Here, we present the
6,332,373-bp complete genome sequence of StFLB209, with a G+C content of 60.7%,
which carries 5,598 protein-coding genes, 6 rRNA operons, and 69 tRNA genes.

<>

<1>Morohoshi, T., Wang, W.Z., Someya, N., Ikeda, T.
<2>Genome Sequence of Microbacterium testaceum StLB037, an N-Acylhomoserine Lactone-Degrading Bacterium Isolated from Potato Leaves.
<3>J. Bacteriol.
<4>193
<5>2072-2073
<6>2011
<7>Microbacterium testaceum is an endophytic Gram-positive bacterium that resides within plant
hosts. M. testaceum StLB037 was isolated from potato
leaves and shows N-acylhomoserine lactone-degrading activity. Here, we
present the 3.98-Mb complete genome sequence of StLB037, with an average
GC content of 70.28%.

<>

<1>Morohoshi, T., Wang, W.Z., Someya, N., Ikeda, T.
<2>Complete Genome Sequence of Chryseobacterium sp. Strain StRB126, an N-Acylhomoserine Lactone-Degrading Bacterium Isolated from Potato Root.
<3>Genome Announcements
<4>2
<5>e00952-14
<6>2014
<7>Chryseobacterium sp. strain StRB126 was isolated from a potato root and showed
N-acylhomoserine lactone-degrading activity. Here, we present the complete
5,503,743-bp genome sequence of StRB126, which has a G+C content of 35.6% and
carries 4,828 protein-coding genes, six rRNA operons, and 80 tRNA genes.

<>

<1>Morris, D.W., Parish, J.H.
<2>Restriction in Myxococcus virescens.
<3>Arch. Microbiol.
<4>108
<5>227-230
<6>1976
<7>1.  The plating efficiency of bacteriophage MX-1 on Myococcus xanthus strains A
and B and M. virescens V2 were compared.  Comparison of strains V2 and A
suggest that V2 is restrictive and A is not (restriction coefficient was
approximately 8).  A derivative of M. virescens V2 (strain V2-9) was obtained
by repeated exposure of strain V2 to ultraviolet radiation.  Strain V2-9 was
also unrestrictive.  Strain B is apparently unrestrictive too but analysis of
phenotypic changes in phage derived from hosts V2, B and A suggested that some
of the host-cell processes differ from orthodox restriction and modification.
2.  Cell-free extracts from M. virescens V2 were fractionated by ion-exchange
chromatography and two restriction endonucleases, R. MviV2I and R. MviV2II were
identified.  Nuclease I was found to hydrolyse coliphage lambda DNA at
apparently one site only and MX-1 DNA at approximately 10 sites; nuclease II
was found to hydrolyse MX-1 DNA at a very large number of sites and its
restriction sequence was of comparable frequency with that of R.EcoRII.
"Modified MX-1 DNA", obtained from phage whose last host was M. virescens V2
was hydroxlysed by nuclease II but not by nuclease I.  The significance of
these findings for restriction in myxococci is discussed.

<>

<1>Morris, P., Marinelli, L.J., Jacobs-Sera, D., Hendrix, R.W., Hatfull, G.F.
<2>Genomic characterization of mycobacteriophage giles: evidence for phage acquisition of host DNA by illegitimate recombination.
<3>J. Bacteriol.
<4>190
<5>2172-2182
<6>2008
<7>A characteristic feature of bacteriophage genomes is that they are
architecturally mosaic, with each individual genome representing a unique
assemblage of individual exchangeable modules. Plausible mechanisms for
generating mosaicism include homologous recombination at shared boundary
sequences of module junctions, illegitimate recombination in a
non-sequence-directed process, and site-specific recombination. Analysis
of the novel mycobacteriophage Giles genome not only extends our current
perspective on bacteriophage genetic diversity, with more than 60% of the
genes unrelated to other mycobacteriophages, but offers novel insights
into how mosaic genomes are created. In one example, the
integration/excision cassette is atypically situated within the structural
gene operon and could have moved there either by illegitimate
recombination or more plausibly via integrase-mediated site-specific
recombination. In a second example, a DNA segment has been recently
acquired from the host bacterial chromosome by illegitimate recombination,
providing further evidence that phage genomic mosaicism is generated by
nontargeted recombination processes.

<>

<1>Morrison, E.A., Garner, S., Echaubard, P., Lesbarreres, D., Kyle, C.J., Brunetti, C.R.
<2>Complete genome analysis of a frog virus 3 (FV3) isolate and sequence comparison with isolates of differing levels of virulence.
<3>Virol. J.
<4>11
<5>46
<6>2014
<7>BACKGROUND: Frog virus 3 (FV3) is the type species of the genus Ranavirus, and in
the past few decades, FV3 infections have resulted in considerable morbidity and
mortality in a range of wild and cultivated amphibian species in the Americas,
Europe, and Asia. The reasons for the pathogenicity of FV3 are not well
understood. FINDINGS: We investigated three FV3 isolates designated SSME, wt-FV3,
and aza-Cr, and reported that our wt-FV3 and aza-Cr strains showed similar levels
of virulence, while SSME was the least virulent in an in vivo study with
Lithiobates pipiens tadpoles. Using 454 GS-FLX sequencing technology, we
sequenced SSME and compared it to the published wt-FV3 genome. SSME had multiple
amino acid deletions in ORFs 49/50L, 65L, 66L, and 87L, which may explain its
reduced virulence. We also investigated repeat regions and found that repeat copy
number differed between isolates, with only one group of 3 isolates and 1 pair of
isolates being identical at all 3 locations. CONCLUSIONS: In this study we have
shown that genetic variability is present between closely related FV3 isolates,
both in terms of deletions/insertions, and even more so at select repeat
locations. These genomic areas with deletions/insertions may represent regions
that affect virulence, and therefore require investigation. Furthermore, we have
identified repeat regions that may prove useful in future phylogeographical
tracking and identification of ranaviral strains across different environmental
regions.

<>

<1>Morrison, H.A., Seligman, L.M.
<2>Homing endonuclease I-CreI mutants with substitutions at residues 30, 32 and 38 include altered specificity derivatives.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>104
<5>313
<6>2004
<7>The homing endonuclease I-CreI recognizes and cleaves a specific 22 base pair DNA sequence.
The amino acid contacts responsible for DNA recognition have been identified; three such
residues that cooperate to interact with a particular target site base pair are N30, S32, and
Q38.  In order to study how these residues function in DNA recognition and cleavage, as well
as the interactions bertween the residues themselves, we employed site-directed mutagenesis to
alter the amino acids at these positions.  Resulting I-CreI derivatives were then assayed in
vivo in an Escherichia coli based system for cleavage activity against appropriate DNA
targets.  A number of active I-CreI derivatives altered at these positions have been isolated
and characterized both in vivo and in vitro.  Our results demonstrate that it is possible to
isolate derivatives of I-CreI altered at residues 30, 32 and 38 that exhibit novel DNA
recognition properties.

<>

<1>Morrison, M.
<2>Deoxyribonucleic acid restriction/modification systems and gene transfer strategies in Ruminococcus albus 8 and Ruminococcus flavofaciens FD-1.
<3>Ph.D. Thesis, University of Illinois, USA
<4>
<5>1-134
<6>1991
<7>One goal critical to the use of recombinant DNA technologies with rumen bacteria is the
establishment of a stable DNA transfer system. Research to date has shown that Ruminococcus
can be classified as a genus resistant to transformation, and that electroporation may offer
the only means to introduce foreign DNA. This thesis aims to elucidate and solve limitations
to the utilization of electroporation with Ruminococcus albus 8 and Ruminococcus flavefaciens
FD-1. The limited degradation of DNA by restriction enzymes and confirmation of plasmid uptake
were given priority, although the consequences of incompatible plasmid replicons and failure
in the expression of selectable markers cannot be overlooked. Fluorescent labelled dextrans
were used in place of plasmid DNA and indicated that electroporation resulted in the uptake of
macromolecules. Type-II endonuclease activities were present in most strains of Ruminococcus
tested. However, the cell-free extract of R. flavefaciens FD-1 did not provide a simple DNA
protection strategy, so the restriction/modification systems of R. albus 8 and R. flavefaciens
FD-1 were studied in more detail. Plasmids derived from a dam proficient Escherichia coli
background are protected against the Type-IIS restriction enzyme of R. albus 8. Adenine
methylation by M.TaqI and a methylase from Chlorella will inhibit DNA cleavage by RflFI and
RFlFII, respectively. The initial electroporation experiments utilizing methylated DNA were
unsuccessful in yielding phenotypic transformants of R. albus 8. Plasmid DNA isolated directly
from Ruminococcus would be a valuable tool in addressing some of the problems still affecting
gene transfer. The plasmid pBAW301, present in R. flavefaciens strain R13c2, was isolated and
is sufficiently small to facilitate construction of potential shuttle vectors as well as broad
host range, chimeric plasmids. Knowledge of DNA modification in Ruminococcus has been
obtained. Other species possessing stable plasmids and gene/s encoding antibiotic resistance
have also been identified. Greater emphasis can now be placed on the electroporation technique
itself, as well as a wider range of selective markers. Ruminococcus ssp. required further
investigation if model genetic systems are to be developed and some of the proposed goals of
molecular biology research for this genus are to be fully realized.

<>

<1>Morrison, M., Mackie, R.I., White, B.A.
<2>Partial characterization of a DNA restriction endonuclease from Ruminococcus flavefaciens FD-1 and its inhibition by site-specific adenine methylation.
<3>Appl. Environ. Microbiol.
<4>58
<5>66-69
<6>1992
<7>The principal DNA restriction-modification system of the cellulolytic ruminal
bacterium Ruminococcus flavefaciens FD-1 is described.  The restriction
endonuclease RflFI could be separated from cell extracts by phosphocellulose
and heparin-sepharose chromatography.  Restriction enzyme digests utilizing
RflFI alone or in combination with SalI, a restriction enzyme isolated from
Streptomyces albus G, showed that the DNA sequence recognized by RflFI either
overlapped or was the same as that recognized by SalI.  DNA sequence analysis
confirmed that RflFI was identical in activity to SalI, with the recognition
sequence being 5'-GTCGAC-3' and cleavage occurring between G and T.  Adenine
methylation within this sequence can be catalyzed in vitro by TaqI methylase,
and this inhibited the cleavage of plasmid DNA molecules by RflFI and SalI.
Chromosomal DNA from R. flavefaciens FD-1 is also methylated within this DNA
sequence because neither restriction endonuclease could degrade this DNA
substrate.  These findings provide a means to protect plasmid molecules from
degradation prior to gene transfer experiments with R. flavefaciens FD-1.

<>

<1>Morrison, M., Mackie, R.I., White, B.A.
<2>Partial purification and characterization of Ral8I, a class-IIS restriction endonuclease from Ruminococcus albus 8 which recognizes 5'-GGATC .
<3>Gene
<4>111
<5>105-108
<6>1992
<7>Heparin-agarose chromatography was used to isolate a restriction endonuclease (ENase) from the
cellulolytic Gram + anaerobe, Ruminococcus albus 8. The enzyme, Ral8I, was eluted from the
column using 230-310 mM Na+. However, the preparation was active only with DNA substrates that
were not Dam-methylated. Moreover the restriction fragment pattern generated from simian virus
40 (SV40) DNA was not consistent with the expected number of Dam-methylation sites. Alignment
of the Dam-methylation sites in SV40 DNA indicated that Ral8I may actually recognize the
asymmetric sequence, GGATC. This was confirmed by nucleotide (nt) sequence analysis and,
further, Ral8I was found to cause cleavage of the DNA approx. 5 nt downstream from the
recognition sequence. Ral8I can therefore be classified as a type-IIS restriction endonuclease
and is an isoschizomer of AlwI, BinI and BthII.

<>

<1>Morrison, M., Mackie, R.I., White, B.A.
<2>Restriction-modification systems in Ruminococcus and development of a gene transfer system.
<3>Aust. Microbiol.
<4>0
<5>301-303
<6>1992
<7>
<>

<1>Morrison, M., Mackie, R.I., White, B.A.
<2>The restriction endonuclease RflFII, isolated from Ruminococcus flavefaciens FD-1, recognizes the sequence 5'-AGTACT-3', and is inhibited by site-specific adenine methylation.
<3>FEMS Microbiol. Lett.
<4>122
<5>181-185
<6>1994
<7>Molecular studies of the rumen bacterium Ruminococcus flavefaciens are constrained by the lack
of stable gene transfer system. We report here on the characterization of RflFII, a
restriction endonuclease isolated from R. flavefaciens FD-1. The enzyme is an isoschizomer of
ScaI, and cleavage of the DNA is blunt-ended, between the internal TA dinucleotide sequence of
5'-AGTACT-3'. Chromosomal DNA preparations were used to demonstrate that adenine methylation
of DNA within the sequence 5'-GTAC-3' inhibits both RflFII and the restriction endonucleases
RsaI and ScaI. Chromosomal DNA from R. flavefaciens FD-1 is also host modified to protect
against cleavage by ScaI.

<>

<1>Morrow, J.F., Berg, P.
<2>Cleavage of Simian Virus 40 DNA at a unique site by a bacterial restriction enzyme.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>3365-3369
<6>1972
<7>The RI restriction endonuclease of Escherichia coli converts covalently-closed
circular Simian Virus 40 (SV40) DNA to unit-length linear duplex molecules.
Cleavage occurs at a unique site, since denaturation and renaturation of these
linear molecules yield linear but no circular molecules.  The distance from the
cleavage site to the SV40 DNA sequence contained in the adenovirus-SV40 hybrid,
Ad2+ND1, is 0.11 of the length of SV40 DNA.  T4 gene 32 protein binds to SV40
DNA in a region 0.45 of the length of SV40 DNA from the RI cleavage site.  E.
coli B restriction endonuclease can cleave SV40 DNA at several sites.

<>

<1>Mortusewicz, O., Schermelleh, L., Walter, J., Cardoso, M.C., Leonhardt, H.
<2>Recruitment of DNA methyltransferase I to DNA repair sites.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>8905-8909
<6>2005
<7>In mammalian cells, the replication of genetic and epigenetic information is directly coupled;
however, little is known about the maintenance of epigenetic information in DNA repair. Using
a laser microirradiation system to introduce DNA lesions at defined subnuclear sites, we
tested whether the major DNA methyltransferase (Dnmt1) or one of the two de novo
methyltransferases (Dnmt3a, Dnmt3b) are recruited to sites of DNA repair in vivo. Time lapse
microscopy of microirradiated mammalian cells expressing GFP-tagged Dnmt1, Dnmt3a, or Dnmt3b1
together with red fluorescent protein-tagged proliferating cell nuclear antigen (PCNA)
revealed that Dnmt1 and PCNA accumulate at DNA damage sites as early as 1 min after
irradiation in S and non-S phase cells, whereas recruitment of Dnmt3a and Dnmt3b was not
observed. Deletion analysis showed that Dnmt1 recruitment was mediated by the PCNA-binding
domain. These data point to a direct role of Dnmt1 in the restoration of epigenetic
information during DNA repair.

<>

<1>Moser, D., Kallas, T.
<2>Characteristics of a restriction endonuclease from the Cyanobacterium Nostoc PCC 7121 and protection of DNA with Eco 47 II methylase.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>155
<6>1991
<7>In order to investigate the structure and function of the cytochrome b6-f
complex, we are trying to develop procedures for gene transfer into and gene
replacement in Nostoc PCC 7121, a heterotrophic cyanobacterium from which the
cytochrome b6-f genes have been cloned and sequenced.  Restriction
endonucleases in Nostoc are being investigated as possible barriers to
transformation.  A restriction endonuclease which we designated Nsp7121I has
been isolated and partially purified.  The DNA recognition sequence (GGNCC) for
this endonuclease was established by comparison of Nsp7171I restriction digests
of plasmid and bacteriophage DNAs with computer generated restriction fragment
profiles.  Plasmid encoded Eco47II methylase protected DNA against restriction
by the Nostoc endonuclease.  Unmodified plasmids previously used in
transformation attempts were cleaved at multiple sites.  Thus one barrier to
transformation has been identified and a modification methylase is available
for its circumvention.  Work is in progress to test transformation of Nostoc
PCC 7121 with Eco47II protected DNA.

<>

<1>Moser, D.P., Zarka, D., Kallas, T.
<2>Characterization of a restriction barrier and electrotransformation of the cyanobacterium Nostoc PCC-7121.
<3>Arch. Microbiol.
<4>160
<5>229-237
<6>1993
<7>We have investigated host restriction as a barrier to transformation and developed a method
for gene transfer into the previously untransformable, heterotrophic cyanobacterium Nostoc PCC
7121. A restriction endonuclease, designed Nsp7121I, has been partialy purified by
phosphocellulose chromatography of Nostoc cell extracts. Comparisons of Nsp7121I digests of
bacteriophage lambda and plasmid DNAs with computer-generated restriction fragment profiles
showed that Nsp7121I is an isoschizomer of restriction endonucleases, such as AsuI, Nsp75241V,
Sau96I, and Eco4711, that recognize the sequence GGNCC. Cleavage by Nsp71211 within this
sequence was confirmed by sequence analysis of DNA fragments cleaved at a unique Nsp7121I
site. These date further suggested that cleavage occurs after the first G (5'-G/GNCC-3') in
this site to generate a three base 5' overhang. Nsp7121I degraded all plasmids used in
previous transformation attempts but modification of these DNA molecules by Eco47II methylase
effectively prevented digestion by Nsp7121I. Plasmids premethylated by passage through
Escherichia coli carrying a plasmid-encoded Eco47II methylase have now been used in an
electroporation procedure to transform Nostoc PCC 7121 to neomycin resistance at frequencies
as high as one transformant per 1000 viable cells. Transformation and stable replication
within Nostoc of one of the transforming plasmids (pRL25), was confirmed by recovery of pRL25,
in its original form, from transformants. Conjugal transfer of pRL25 from E. coli into Nostoc
was also possible but at much lower efficiency than by electroporation. These findings
establish the basis for genetic analysis of Nostoc PCC 7121, from which genes for
photosynthetic electron transport have been cloned.

<>

<1>Moses, P., Horiuchi, K.
<2>Specific Recombination in Vitro Promoted by the Restriction Endonuclease HgaI.
<3>J. Mol. Biol.
<4>135
<5>517-524
<6>1979
<7>We describe the use of the restriction endonuclease HgaI from Haemophilus
gallinarum for the efficient construction in vitro of recombinant molecules.
Using bacteriophage f1 DNA, we show that only HgaI fragments that were
orginally adjacent on the genome are ligated, that upon ligation infectious
molecules are reassembled with high efficiency, and that recombinant genomes
can thus be easily constructed.  The method relies upon the unique properties
of HgaI and is applicable to any viral or plasmid DNA that contains several
HgaI recognition sites.

<>

<1>Mosier, A.C., Allen, E.E., Kim, M., Ferriera, S., Francis, C.A.
<2>Genome Sequence of 'Candidatus Nitrosopumilus salaria' BD31, an Ammonia-Oxidizing Archaeon from the San Francisco Bay Estuary.
<3>J. Bacteriol.
<4>194
<5>2121-2122
<6>2012
<7>Ammonia-oxidizing archaea (AOA) play important roles in nitrogen and carbon cycling in marine
and terrestrial ecosystems. Here, we present the draft genome
sequence for the ammonia-oxidizing archaeon 'Candidatus Nitrosopumilus salaria'
BD31, which was enriched in culture from sediments of the San Francisco Bay
estuary. The genome sequences revealed many similarities to the genome of
Nitrosopumilus maritimus.

<>

<1>Mosier, A.C., Allen, E.E., Kim, M., Ferriera, S., Francis, C.A.
<2>Genome Sequence of 'Candidatus Nitrosoarchaeum limnia' BG20, a Low-Salinity Ammonia-Oxidizing Archaeon from the San Francisco Bay Estuary.
<3>J. Bacteriol.
<4>194
<5>2119-2120
<6>2012
<7>Here, we present the draft genome sequence of 'Candidatus Nitrosoarchaeum limnia' BG20, an
ammonia-oxidizing archaeon enriched in culture from low-salinity
sediments of the San Francisco Bay estuary. The genome sequence revealed many
similarities to the previously sequenced genome of 'Ca. Nitrosoarchaeum limnia'
SFB1 (enriched from a nearby site in San Francisco Bay) and is representative of
a clade of ammonia-oxidizing archaea (AOA) found in low-salinity habitats
worldwide.

<>

<1>Motamedchaboki, K.
<2>Characterization of a restriction endonuclease isolated from Bacillus subtilis F2.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>457
<6>2001
<7>Usefulness of restriction system with their high substrate specificities for ds-DNA, and the
processing of the free or packaged
DNA during uptake into Bacillus subtilis cells made us study about the
restriction enzyme activity of one of our laboratory isolate, Bacillus
subtilis F2 which have been used for the propagation of the newly
isolated PAK phage. During the life cycle of PAK phage in Bacillus
subtilis F2 host, it has been noticed that DNA of phage get fragmented
in vivo, therefore, it becomes important to explore the endonuclease
activity in this strain. A cell bound enzyme was isolated from Bacillus
subtilis F2, and was characterized in crude extract of bacterial cell
pellet. The enzyme showed to have endonuclease activity on
bacteriophage lambda DNA and other DNA as well. Interestingly, lambda
DNA seems to have very few cut sites. Another important characteristic
of this enzyme is to have optimum pH shifted toward alkaline range,
which is 8.5 among of 182 known restriction enzymes. The optimum
temperature found to be 40 C. It seems to work at high salt
concentration. Lysates extracted at pH 6 and 6.5 have shown
endonuclease activity with a new profile of lambda DNA fragmentation of
lysate extracted at pH 7.5. Hence BsuF2 is a type II restriction enzyme
it seems to have high potential to be used as a genetic tool in
molecular biology.

<>

<1>Motoshima, K., Ishikawa, M., Hashimoto, Y., Sugita, K.
<2>Inhibition of Restriction Enzymes EcoRI, BamHI and HindIII by Phenethylphenylphthalimides Derived from Thalidomide.
<3>Chem. Pharm. Bull. (Tokyo)
<4>59
<5>880-884
<6>2011
<7>We discovered inhibitors of the restriction enzymes EcoRI, BamHI and HindIII by screening our
library of compounds with a
phenethylphenylphthalimide skeleton, based on alpha-glucosidase
inhibitors and liver X receptor antagonists derived from thalidomide.
Structural development afforded the potent restriction enzyme
inhibitors 25 and 26.

<>

<1>Moulin, L. et al.
<2>Complete Genome sequence of Burkholderia phymatum STM815(T), a broad host range and efficient nitrogen-fixing symbiont of Mimosa species.
<3>Standards in Genomic Sciences
<4>9
<5>763-774
<6>2014
<7>Burkholderia phymatum is a soil bacterium able to develop a nitrogen-fixing symbiosis with
species of the legume genus Mimosa, and is frequently found
associated specifically with Mimosa pudica. The type strain of the species, STM
815(T), was isolated from a root nodule in French Guiana in 2000. The strain is
an aerobic, motile, non-spore forming, Gram-negative rod, and is a highly
competitive strain for nodulation compared to other Mimosa symbionts, as it also
nodulates a broad range of other legume genera and species. The 8,676,562 bp
genome is composed of two chromosomes (3,479,187 and 2,697,374 bp), a megaplasmid
(1,904,893 bp) and a plasmid hosting the symbiotic functions (595,108 bp).

<>

<1>Moulin, L., Mornico, D., Melkonian, R., Klonowska, A.
<2>Draft Genome Sequence of Rhizobium mesoamericanum STM3625, a Nitrogen-Fixing Symbiont of Mimosa pudica Isolated in French Guiana (South America).
<3>Genome Announcements
<4>1
<5>e00066-12
<6>2013
<7>Rhizobium mesoamericanum STM3625 is a Mimosa pudica symbiont isolated in French Guiana. This
strain serves as a model bacterium for comparison of adaptation to mutualism (symbiotic
traits, bacterial genetic programs for plant infection) between alpha and beta rhizobial
symbionts of Mimosa pudica.

<>

<1>Mounts, W.M., Whitley, M.Z., Murphy, E.
<2>Nucleic acid arrays for detecting multiple strains of a non-viral species.
<3>International Patent Office
<4>WO 2005014857 A
<5>
<6>2005
<7>Nucleic acid arrays and methods of using the same for concurrent or discriminable detection of
different strains of a non-viral species. In many embodiments, the nucleic acid arrays of the
present invention include probes that are specific to different respective strains of a
non-viral species. In many other embodiments, the nucleic acid arrays of the present invention
include probes that are common to two or more different strains of the non-viral species. In
one embodiment, the non-viral species is Staphylococcus aureus, and the different
Staphylococcus aureus strains include COL, N315, MOO, EMRSA-16, MSSA-476, and 8325 strains.
In another embodiment, a nucleic acid array of the present invention includes polynucleotide
probes capable of hybridizing under stringent or nucleic acid array hybridization conditions
to respective sequences selected from SEQ ID NOs: 1 to 7,852, or the complements thereof.

<>

<1>Moure, C.M., Gimble, F.S., Quiocho, F.A.
<2>Crystal structure of the intein homing endonuclease PI-SceI bound to its recognition sequence.
<3>Nat. Struct. Biol.
<4>9
<5>764-770
<6>2002
<7>The first X-ray structures of an intein-DNA complex, that of the two-domain homing
endonuclease PI-SceI bound to its 36-base pair DNA substrate, have been determined in the
presence and absence of Ca(2+). The DNA shows an asymmetric bending pattern, with a major 50
degrees bend in the endonuclease domain and a minor 22 degrees bend in the splicing domain
region. Distortions of the DNA bound to the endonuclease domain cause the insertion of the two
cleavage sites in the catalytic center. DNA binding induces changes in the protein
conformation. The two overlapping non-identical active sites in the endonucleolytic center
contain two Ca(+2) ions that coordinate to the catalytic Asp residues. Structure analysis
indicates that the top strand may be cleaved first.

<>

<1>Moure, C.M., Gimble, F.S., Quiocho, F.A.
<2>The crystal structure of the gene targeting homing endonuclease I-SceI reveals the origins of its target site specificity.
<3>J. Mol. Biol.
<4>334
<5>685-695
<6>2003
<7>The I-SceI homing endonuclease enhances gene targeting by introducing double-strand breaks at
specific chromosomal loci, thereby increasing the
recombination frequency. Here, we report the crystal structure of the
enzyme complexed to its DNA substrate and Ca(2+) determined at 2.25A
resolution. The structure shows the prototypical beta-saddle of LAGLIDADG
homing endonucleases that is contributed by two pseudo-symmetric domains.
The high specificity of I-SceI is explained by the large number of
protein-DNA contacts, many that are made by a long beta-hairpin loop that
reaches into the major groove of the DNA. The DNA minor groove is
compressed at the catalytic center, bringing the two scissile
phosphodiester bonds into close proximity. The protein-Ca(2+)-DNA
structure shows the protein bound to its DNA substrate in a pre-reactive
state that is defined by the presence of two asymmetric active sites, one
of which appears poised to first cleave the DNA bottom strand.

<>

<1>Moure, C.M., Gimble, F.S., Quiocho, F.A.
<2>Crystal structures of I-SceI complexed to nicked DNA substrates: snapshots of intermediates along the DNA cleavage reaction pathway.
<3>Nucleic Acids Res.
<4>36
<5>3287-3296
<6>2008
<7>I-SceI is a homing endonuclease that specifically cleaves an 18-bp double-stranded DNA. I-SceI
exhibits a strong preference for cleaving the bottom strand DNA. The published structure of
I-SceI bound to an uncleaved DNA substrate provided a mechanism for bottom strand cleavage but
not for top strand cleavage. To more fully elucidate the I-SceI catalytic mechanism, we
determined the X-ray structures of I-SceI in complex with DNA substrates that are nicked in
either the top or bottom strands. The structures resemble intermediates along the DNA cleavage
reaction. In a structure containing a nick in the top strand, the spatial arrangement of metal
ions is similar to that observed in the structure that contains uncleaved DNA, suggesting that
cleavage of the bottom strand occurs by a common mechanism regardless of whether this strand
is cleaved first or second. In the structure containing a nick in the bottom strand, a new
metal binding site is present in the active site that cleaves the top strand. This new metal
and a candidate nucleophilic water molecule are correctly positioned to cleave the top strand
following bottom strand cleavage, providing a plausible mechanism for top strand cleavage.

<>

<1>Moure, C.M., Quiocho, F.A.
<2>The structure and function of intein-associated homing endonucleases.
<3>Nucleic Acids Mol. Biol.
<4>16
<5>257-271
<6>2005
<7>Homing endonucleases are a large group of proteins that are characterized by their ability to
recognize long (14-40 base pairs, bp) asymmetric or pseudo-palindromic double-stranded DNA
sequences as cleavage sites.  By cleaving DNA, they assist in the homing process of their
encoding genes into other genes.  Homing endonucleases include both intron-encoded and intein-
(for intervening protein) associated members that self-splice at the mRNA and protein level,
respectively.  Those inteins that contain associated endonuclease domains are especially
interesting due to their bifunctionality and their exceptionally long DNA recognition
abilities.  With two exceptions, all of the approximately 160 inteins that have been
characterized to date are associated with the LAGLIDADG homing endonuclease subfamily, which
also includes numerous intron-encoded endonucleases.  This chapter is devoted mainly to the
structural studies of intein-associated LAGLIDADG homing endonucleases that have revealed the
domain organization and architecture of this type of protein, providing insights into the
combination of protein splicing and site-specific DNA recognition and cleavage across a single
peptide chain.  The knowledge gained in these structure-function studies has been successfully
exploited in biotechnology to devise systems for protein purification and to study
protein-protein interactions using the splicing capability of inteins, and in gene targeting
studies, which use their rare-cutting endonuclease properties.  These applications are
discussed in other chapters in this volume.

<>

<1>Moxon, E.R., Rainey, P.B., Nowak, M.A., Lenski, R.E.
<2>Adaptive evolution of highly mutable loci in pathogenic bacteria.
<3>Curr. Biol.
<4>4
<5>24-33
<6>1994
<7>Bacteria have specific loci that are highly mutable.  We argue that the coexistence within
bacterial genomes of such 'contingency' genes with high mutation rates, and 'housekeeping'
genes with low mutation rates, is the result of adaptive evolution, and facilitates the
efficient exploration of phenotypic solutions to unpredictable aspects of the host environment
while minimizing deleterious effects on fitness.

<>

<1>Moya-Beltran, A., Cardenas, J.P., Covarrubias, P.C., Issotta, F., Ossandon, F.J., Grail, B.M., Holmes, D.S., Quatrini, R., Johnson, D.B.
<2>Draft Genome Sequence of the Nominated Type Strain of 'Ferrovum myxofaciens,' an  Acidophilic, Iron-Oxidizing Betaproteobacterium.
<3>Genome Announcements
<4>2
<5>e00834-14
<6>2014
<7>'Ferrovum myxofaciens' is an iron-oxidizing betaproteobacterium with widespread distribution
in acidic low-temperature environments, such as acid mine drainage
streams. Here, we describe the genomic features of this novel acidophile and
investigate the relevant metabolic pathways that enable its survival in these
environments.

<>

<1>Mrazek, J., Karlin, S.
<2>Detecting alien genes in bacterial genomes.
<3>Ann. NY Acad. Sci.
<4>870
<5>314-329
<6>1999
<7>We present new methods for calculating codon bias of a group of genes or an individual gene
relative to a standard gene class.  This method is suitable for identifying alien (e.g.,
horizonatally transferred) and highly expressed genes.  In yeast and several bacterial
genomes, highly expressed genes typically include ribosomal protein genes, elongation factors,
chaperonins (heat shock proteins), and a subset of genes involved in glycolysis generally
essential in exponential growth.  Highly expressed genes of the Synechocystis genome feature
several photosystem II genes, and highly expressed genes in several methanogens (Methanococcus
jannaschii, M. thermoautotrophicum) are essential for methanogenesis.  Alien genes mostly
consist of ORFs of unknown function, transposases, prophage genes, and
restriction/modification enzymes.  Notably, nuclear ribosomal proteins of yeast are highly
expressed, whereas mitochondrial ribosomal protein genes appear to be alien genes.  Alien
genes often occur in clusters, suggesting in these cases that transfer events entail several
genes.

<>

<1>Mrazek, J., Piknova, M., Pristas, P., Kopecny, J.
<2>Occurrence of restriction-modification systems in ruminal butyrate-producing bacteria.
<3>Anaerobe
<4>11
<5>280-284
<6>2005
<7>Thirty-five strains of ruminal bacteria belonging to the former Butyrivibrio fibrisolvens
species were screened for the presence of
site-specific restriction endonuclease and modification
methyltransferase activities. Seven strains possessed endonuclease
activities detectable in crude cell extracts. The recognition sequences
and optimal reaction conditions for seven of them were determined. Five
enzymes were found to be isoschizomers of type II endonucleases (EcoRV,
Nsil, Asel (2x) and Saul), one was type IIS (FokI) and two remained
unknown. The optimal reaction buffer was found to be a low ionic
strength buffer and all enzymes possessed sufficient activity at 39
degrees C. The presence of DNA modification among all strains was also
determined. Most of the methylation activities correlated with
restriction activities, yet some strains possessed unaccompanied
modification methyltransferases.

<>

<1>Mruk, I., Blumenthal, R.M.
<2>Tuning the relative affinities for activating and repressing operators of a temporally regulated restriction-modification system.
<3>Nucleic Acids Res.
<4>37
<5>983-998
<6>2009
<7>Most type II restriction-modification (R-M) systems produce separate endonuclease (REase) and
methyltransferase (MTase) proteins. After R-M
genes enter a new cell, MTase activity must appear before REase or the
host chromosome will be cleaved. Temporal control of these genes thus has
life-or-death consequences. PvuII and some other R-M systems delay
endonuclease expression by cotranscribing the REase gene with the upstream
gene for an autogenous activator/repressor (C protein). C.PvuII was
previously shown to have low levels early, but positive feedback later
boosts transcription of the C and REase genes. The MTase is expressed
without delay, and protects the host DNA. C.PvuII binds to two sites
upstream of its gene: O(L), associated with activation, and O(R),
associated with repression. Even when symmetry elements of each operator
are made identical, C.PvuII binds preferentially to O(L). In this study,
the intra-operator spacers are shown to modulate relative C.PvuII
affinity. In light of a recently reported C.Esp1396I-DNA co-crystal
structure, in vitro and in vivo effects of altering O(L) and O(R) spacers
were determined. The results suggest that the GACTnnnAGTC consensus is the
primary determinant of C.PvuII binding affinity, with intra-operator
spacers playing a fine-tuning role that affects mobility of this R-M
system.

<>

<1>Mruk, I., Blumenthal, R.M.
<2>Real-time kinetics of restriction-modification gene expression after entry into a new host cell.
<3>Nucleic Acids Res.
<4>36
<5>2581-2593
<6>2008
<7>Most type II restriction-modification (R-M) systems produce separate restriction endonuclease
(REase) and methyltransferase (MTase) proteins.
After R-M system genes enter a new cell, protective MTase must appear
before REase to avoid host chromosome cleavage. The basis for this
apparent temporal regulation is not well understood. PvuII and some other
R-M systems appear to achieve this delay by cotranscribing the REase gene
with the gene for an autogenous transcription activator/repressor (the 'C'
protein C.PvuII). To test this model, bacteriophage M13 was used to
introduce the PvuII genes into a bacterial population in a relatively
synchronous manner. REase mRNA and activity appeared approximately 10 min
after those of the MTase, but never rose if there was an inactivating
pvuIIC mutation. Infection with recombinant M13pvuII phage had little
effect on cell growth, relative to infection with parental M13. However,
infection of cells pre-expressing C.PvuII led to cessation of growth. This
study presents the first direct demonstration of delayed REase expression,
relative to MTase, when type II R-M genes enter a new host cell.
Surprisingly, though the C and REase genes are cotranscribed, the pvuIIC
portion of the mRNA was more abundant than the pvuIIR portion after stable
establishment of the R-M system.

<>

<1>Mruk, I., Cichowicz, M., Kaczorowski, T.
<2>Characterization of the LlaCl methyltransferase from Lactococcus lactis subsp cremoris W15 provides new insights into the biology of type II  restriction-modification systems.
<3>Microbiology
<4>149
<5>3331-3341
<6>2003
<7>The gene encoding the LlaCI methyltransferase (M.LlaCI) from Lactococcus lactis subsp.
cremoris W15 was overexpressed in Escherichia coli. The
enzyme was purified to apparent homogeneity using three consecutive steps
of chromatography on phosphocellulose, blue-agarose and Superose 12HR,
yielding a protein of M(r) 31 300+/-1000 under denaturing conditions. The
exact position of the start codon AUG was determined by protein
microsequencing. This enzyme recognizes the specific palindromic sequence
5'-AAGCTT-3'. Purified M.LlaCI was characterized. Unlike many other
methyltransferases, M.LlaCI exists in solution predominantly as a dimer.
It modifies the first adenine residue at the 5' end of the specific
sequence to N(6)-methyladenine and thus is functionally identical to the
corresponding methyltransferases of the HindIII (Haemophilus influenzae
Rd) and EcoVIII (Escherichia coli E1585-68) restriction-modification
systems. This is reflected in the identity of M.LlaCI with M.HindIII and
M.EcoVIII noted at the amino acid sequence level (50 % and 62 %,
respectively) and in the presence of nine sequence motifs conserved among
N(6)-adenine beta-class methyltransferases. However, polyclonal antibodies
raised against M.EcoVIII cross-reacted with M.LlaCI but not with
M.HindIII. Restriction endonucleases require Mg(2+) for phosphodiester
bond cleavage. Mg(2+) was shown to be a strong inhibitor of the M.LlaCI
enzyme and its isospecific homologues. This observation suggests that
sensitivity of the M.LlaCI to Mg(2+) may strengthen the restriction
activity of the cognate endonuclease in the bacterial cell. Other
biological implications of this finding are also discussed.

<>

<1>Mruk, I., Kaczorowski, T.
<2>Genetic organization and molecular analysis of the EcoVIII restriction-modification system of Escherichia coli E1585-68 and its comparison with isospecific homologs.
<3>Appl. Environ. Microbiol.
<4>69
<5>2638-2650
<6>2003
<7>The EcoVIII restriction-modification (R-M) system is carried by the Escherichia coli E1585-68
natural plasmid pEC156 (4,312 bp). The two genes
were cloned and characterized. The G+C content of the EcoVIII R-M system
is 36.1%, which is significantly lower than the average G+C content of
either plasmid pEC156 (43.6%) or E. coli genomic DNA (50.8%). The
difference suggests that there is a possibility that the EcoVIII R-M
system was recently acquired by the genome. The 921-bp EcoVIII
endonuclease (R. EcoVIII) gene (ecoVIIIR) encodes a 307-amino-acid protein
with an M(r) of 35,554. The convergently oriented EcoVIII
methyltransferase (M. EcoVIII) gene (ecoVIIIM) consists of 912 bp that
code for a 304-amino-acid protein with an M(r) of 33,930. The exact
positions of the start codon AUG were determined by protein
microsequencing. Both enzymes recognize the specific palindromic sequence
5'-AAGCTT-3'. Preparations of EcoVIII R-M enzymes purified to homogeneity
were characterized. R. EcoVIII acts as a dimer and cleaves a specific
sequence between two adenine residues, leaving 4-nucleotide 5' protruding
ends. M. EcoVIII functions as a monomer and modifies the first adenine
residue at the 5' end of the specific sequence to N(6)-methyladenine.
These enzymes are thus functionally identical to the corresponding enzymes
of the HindIII (Haemophilus influenzae Rd) and LlaCI (Lactococcus lactis
subsp. cremoris W15) R-M systems. This finding is reflected by the levels
of homology of M. EcoVIII with M. HindIII and M. LlaCI at the amino acid
sequence level (50 and 62%, respectively) and by the presence of nine
sequence motifs conserved among m(6) N-adenine beta-class
methyltransferases. The deduced amino acid sequence of R. EcoVIII shows
weak homology with its two isoschizomers, R. HindIII (26%) and R. LlaCI
(17%). A catalytic sequence motif characteristic of restriction
endonucleases was found in the primary structure of R. EcoVIII
(D(108)X(12)DXK(123)), as well as in the primary structures of R. LlaCI
and R. HindIII. Polyclonal antibodies raised against R. EcoVIII did not
react with R. HindIII, while anti-M. EcoVIII antibodies cross-reacted with
M. LlaCI but not with M. HindIII. R. EcoVIII requires Mg(II) ions for
phosphodiester bond cleavage. We found that the same ions are strong
inhibitors of the M. EcoVIII enzyme. The biological implications of this
finding are discussed.

<>

<1>Mruk, I., Kaczorowski, T.
<2>A rapid and efficient method for cloning genes of type II restriction-modification systems by use of a killer plasmid.
<3>Appl. Environ. Microbiol.
<4>73
<5>4286-4293
<6>2007
<7>We present a method for cloning restriction-modification (R-M) systems that is based on the
use of a lethal plasmid (pKILLER). The plasmid
carries a functional gene for a restriction endonuclease having the
same DNA specificity as the R-M system of interest. The first step is
the standard preparation of a representative, plasmid-borne genomic
library. Then this library is transformed with the killer plasmid. The
only surviving bacteria are those which carry the gene specifying a
protective DNA methyltransferase. Conceptually, this in vivo selection
approach resembles earlier methods in which a plasmid library was
selected in vitro by digestion with a suitable restriction
endonuclease, but it is much more efficient than those methods. The new
method was successfully used to clone two R-M systems, BstZ1II from
Bacillus stearothermophilus 14P and Csp231I from Citrobacter sp. strain
RFL231, both isospecific to the prototype HindIII R-M system.

<>

<1>Mruk, I., Kobayashi, I.
<2>To be or not to be: regulation of restriction-modification systems and other toxin-antitoxin systems.
<3>Nucleic Acids Res.
<4>42
<5>70-86
<6>2013
<7>One of the simplest classes of genes involved in programmed death is that containing the
toxin-antitoxin (TA) systems of prokaryotes. These systems are composed of an intracellular
toxin and an antitoxin that neutralizes its effect. These systems, now classified into five
types, were initially discovered because some of them allow the stable maintenance of mobile
genetic elements in a microbial population through postsegregational killing or the death of
cells that have lost these systems. Here, we demonstrate parallels between some TA systems and
restriction-modification systems (RM systems). RM systems are composed of a restriction enzyme
(toxin) and a modification enzyme (antitoxin) and limit the genetic flux between lineages with
different epigenetic identities, as defined by sequence-specific DNA methylation. The
similarities between these systems include their postsegregational killing and their effects
on global gene expression. Both require the finely regulated expression of a toxin and
antitoxin. The antitoxin (modification enzyme) or linked protein may act as a transcriptional
regulator. A regulatory antisense RNA recently identified in an RM system can be compared with
those RNAs in TA systems. This review is intended to generalize the concept of TA systems in
studies of stress responses, programmed death, genetic conflict and epigenetics.

<>

<1>Mruk, I., Liu, Y., Ge, L., Kobayashi, I.
<2>Antisense RNA associated with biological regulation of a restriction-modification system.
<3>Nucleic Acids Res.
<4>39
<5>5622-5632
<6>2011
<7>Restriction-modification systems consist of a modification enzyme that methylates a specific
DNA sequence and a restriction endonuclease that cleaves DNA lacking this epigenetic
signature. Their gene expression should be finely regulated because their potential to attack
the host bacterial genome needs to be controlled. In the EcoRI system, where the restriction
gene is located upstream of the modification gene in the same orientation, we previously
identified intragenic reverse promoters affecting gene expression. In the present work, we
identified a small (88 nt) antisense RNA (Rna0) transcribed from a reverse promoter (P(REV0))
at the 3' end of the restriction gene. Its antisense transcription, as measured by
transcriptional gene fusion, appeared to be terminated by the P(M1,M2) promoter. P(M1,M2)
promoter-initiated transcription, in turn, appeared to be inhibited by P(REV0). Mutational
inactivation of P(REV0) increased expression of the restriction gene. The biological
significance of this antisense transcription is 2-fold. First, a mutation in P(REV0) increased
restriction of incoming DNA. Second, the presence of the antisense RNA gene (ecoRIA) in trans
alleviated cell killing after loss of the EcoRI plasmid (post-segregational killing). Taken
together, these results strongly suggested the involvement of an antisense RNA in the
biological regulation of this restriction-modification system.

<>

<1>Mruk, I., Rajesh, P., Blumenthal, R.M.
<2>Regulatory circuit based on autogenous activation-repression: roles of C-boxes and spacer sequences in control of the PvuII  restriction-modification system.
<3>Nucleic Acids Res.
<4>35
<5>6935-6952
<6>2007
<7>Type II restriction-modification (R-M) systems comprise a restriction endonuclease (REase) and
a protective methyltransferase (MTase). After R-M
genes enter a new cell, MTase must appear before REase or the chromosome
will be cleaved. PvuII and some other R-M systems achieve this delay by
cotranscribing the REase gene with the gene for an autogenous
transcription activator (the controlling or 'C' protein C.PvuII). This
study reveals, through in vivo titration, that C.PvuII is not only an
activator but also a repressor for its own gene. In other systems, this
type of circuit can result in oscillatory behavior. Despite the use of
identical, symmetrical C protein-binding sequences (C-boxes) in the left
and right operators, C.PvuII showed higher in vitro affinity for O(L) than
for O(R), implicating the spacer sequences in this difference. Mutational
analysis associated the repression with O(R), which overlaps the promoter
-35 hexamer but is otherwise dispensable for activation. A nonrepressing
mutant exhibited poor establishment in new cells. Comparing
promoter-operator regions from PvuII and 29 R-M systems controlled by C
proteins revealed that the most-highly conserved sequence is the
tetranucleotide spacer separating O(L) from O(R). Any changes in that
spacer reduced the stability of C.PvuII-operator complexes and abolished
activation.

<>

<1>Mruk, I., Sektas, M., Kaczorowski, T.
<2>Characterization of pEC156, a ColE1-Type Plasmid from Escherichia coli E1585-68 That Carries Genes of the EcoVIII Restriction-Modification System.
<3>Plasmid
<4>46
<5>128-139
<6>2001
<7>The complete 4312-bp sequence of the pEC156 plasmid from Escherichia coli E1585-68, which
carries genes encoding the EcoVIII restriction-modification (R-M) system, an isoschizomer of
HindIII from Haemophilus influenzae, has been determined. Two clustered and convergently
oriented open reading frames, large enough to encode genes of the EcoVIII R-M system, were
found. The transcriptional start points were mapped by the primer extension method. The
relative molecular masses of the EcoVIII endonuclease and EcoVIII methyltransferase deduced
from the nucleotide sequence are 35,554 and 33,910, respectively. Nucleotide sequence analysis
of pEC156 suggests that this plasmid is a ColE1-type replicon. It consists of an origin of
replication and two untranslated genes encoding RNA I and RNA II, both involved in the
regulation of plasmid DNA replication. The replication region also contains the gene encoding
a 64-aa Rom-like protein. Inactivation of the putative rom gene by insertion of a
kanamycin-resistance cassette resulted in 4.5-fold increase in pEC156-derived plasmid copy
number in E. coli cells. All of these elements (RNA I, RNA II, and rom) reveal a high level of
similarity to ColE1 homologs. The replication of all ColE1-type plasmids is dependent on the
activity of E. coli DNA polymerase I. It was shown that a pEC156 derivative (pIB8) carrying an
antibiotic resistance gene indeed failed to replicate in an E. coli polA12(ts) mutant at 43
degrees C, and its copy number was reduced in the E. coli pcnB80 mutant. These results prove
that pEC156 is a ColE1-type replicon. Copyright 2001 Academic Press.

<>

<1>Muchova, J., Lacova, B., Godany, A., Sevcikova, B.
<2>High transformable mutants of Streptomyces aureofaciens containing restriction-modification systems.
<3>J. Basic Microbiol.
<4>31
<5>141-147
<6>1991
<7>Streptomyces aureofaciens 13 is a mutant defective in chlortetracycline
production. It was chosen as a potentially useful host for gene cloning in
investigations of the organization of the biosynthetic genes for the
tetracycline antibiotic pathway.  From the Streptomyces aureofaciens 13 strain,
three suitable clones were used for our work.  The conditions for optimal
formation and efficient transformation of protoplasts with plasmid DNAs have
been determined.  Transformation frequencies of about 10/4 to 10/5 per
microgram of plasmid DNA were obtained when plasmids were isolated from
Streptomyces strains.  From the patterns of restriction enzyme digestion of
plasmid DNA isolated from Streptomyces aureofaciens transformants, it was
observed that the clones express modification systems which render plasmid DNAs
resistant to cleavage by HindIII and EcoRI.  Additionally, one of the clones
produces the restriction endonuclease Sau13I (isoschizomer of SauI).  The
presence of the restriction-modification system of Sau13I does not reduce the
efficiency of plasmid transformation.  Note: Sau13I is already used for an
isoschizomer of AsuI from Staphylococcus aureus.

<>

<1>Mucke, M., Grelle, G., Behlke, J., Kraft, R., Kruger, D.H., Reuter, M.
<2>EcoRII: a restriction enzyme evolving recombination functions?
<3>EMBO J.
<4>21
<5>5262-5268
<6>2002
<7>The restriction endonuclease EcoRII requires the cooperative interaction with two copies of
the sequence 5'CCWGG for DNA cleavage. We found by limited proteolysis that EcoRII has a
two-domain structure that enables this particular mode of protein-DNA interaction. The
C-terminal domain is a new restriction endonuclease, EcoRII-C. In contrast to the wild-type
enzyme, EcoRII-C cleaves DNA specifically at single 5'CCWGG sites. Moreover, substrates
containing two or more cooperative 5'CCWGG sites are cleaved much more efficiently by
EcoRII-C than by EcoRII. The N-terminal domain binds DNA specifically and attenuates the
activity of EcoRII by making the enzyme dependent on a second 5'CCWGG site. Therefore, we
suggest that a precursor EcoRII endonuclease acquired an additional DNA-binding domain to
enable the interaction with two 5'CCWGG sites. The current EcoRII molecule could be an
evolutionary intermediate between a site-specific endonuclease and a protein that functions
specifically with two DNA sites such as recombinases and transposases. The combination of
these functions may enable EcoRII to accomplish its own propagation similarly to transposons.

<>

<1>Mucke, M., Kruger, D.H., Reuter, M.
<2>Diversity of Type II restriction endonucleases that require two DNA recognition sites.
<3>Nucleic Acids Res.
<4>31
<5>6079-6084
<6>2003
<7>Orthodox Type IIP restriction endonucleases, which are commonly used in molecular biological
work, recognize a single palindromic DNA recognition
sequence and cleave within or near this sequence. Several new studies have
reported on structural and biochemical peculiarities of restriction
endonucleases that differ from the orthodox in that they require two
copies of a particular DNA recognition sequence to cleave the DNA. These
two sites requiring restriction endonucleases belong to different subtypes
of Type II restriction endonucleases, namely Types IIE, IIF and IIS. We
compare enzymes of these three types with regard to their DNA recognition
and cleavage properties. The simultaneous recognition of two identical DNA
sites by these restriction endonucleases ensures that single unmethylated
recognition sites do not lead to chromosomal DNA cleavage, and might
reflect evolutionary connections to other DNA processing proteins that
specifically function with two sites.

<>

<1>Mucke, M., Lurz, R., Mackeldanz, P., Behlke, J., Kruger, D.H., Reuter, M.
<2>Imaging DNA loops induced by restriction endonuclease EcoRII. A single amino acid substitution uncouples target recognition from cooperative DNA interaction and cleavage.
<3>J. Biol. Chem.
<4>275
<5>30631-30637
<6>2000
<7>EcoRII is a type IIE restriction endonuclease characterized by a highly cooperative reaction
mechanism that depends on simultaneous binding of the dimeric enzyme molecule to two copies of
its DNA recognition site. Transmission electron microscopy provided direct evidence that
EcoRII mediates loop formation of linear DNA containing two EcoRII recognition sites. Specific
DNA binding of EcoRII revealed a symmetrical DNase I footprint occupying 16-18 bases. Single
amino acid replacement of Val(258) by Asn yielded a mutant enzyme that was unaffected in
substrate affinity and DNase I footprinting properties, but exhibited a profound decrease in
cooperative DNA binding and cleavage activity. Because the electrophoretic mobility of the
mutant enzyme-DNA complexes was significantly higher than that of the wild-type, we
investigated if mutant V258N binds as a monomer to the substrate DNA. Analysis of the
molecular mass of mutant V258N showed a high percentage of protein monomers in solution. The
dissociation constant of mutant V258N confirmed a 350-fold decrease of the enzyme dimerization
capability. We conclude that Val(258) is located in a region of EcoRII involved in
homodimerization. This is the first report of a specific amino acid replacement in a
restriction endonuclease leading to the loss of dimerization and DNA cleavage while retaining
specific DNA binding.

<>

<1>Mucke, M., Pingoud, V., Grelle, G., Kraft, R., Kruger, D.H., Reuter, M.
<2>Asymmetric photocross-linking pattern of restriction endonuclease EcoRII to the DNA recognition sequence.
<3>J. Biol. Chem.
<4>277
<5>14288-14293
<6>2002
<7>The EcoRII homodimer engages two of its recognition sequences (5'-CCWGG) simultaneously and
is therefore a type IIE restriction endonuclease. To identify the amino acids of EcoRII that
interact specifically with the recognition sequence, we photocross-linked EcoRII with
oligonucleotide substrates that contained only one recognition sequence for EcoRII. In this
recognition sequence, we substituted either 5-iododeoxycytidine for each C or
5-iododeoxyuridine for A, G, or T. These iodopyrimidine bases were excited using a UV laser to
result in covalent cross-linking products. The yield of EcoRII photocross-linked to the 5'-C
of the 5'-CCAGG strand of the recognition sequence was 45%. However, we could not
photocross-link EcoRII to the 5'-C of the 5'-CCTGG strand. Thus, the contact of EcoRII to
the bases of the recognition sequence appears to be asymmetric, unlike that expected for most
type II restriction endonucleases. Tryptic digestion of free and of cross-linked EcoRII,
followed by high performance liquid chromatography (HPLC) separation of the individual
peptides and Edman degradation, identified amino acids 25-49 of EcoRII as the cross-linking
peptide. Mutational analysis of the electron-rich amino acids His(36) and Tyr(41) of this
peptide indicates that Tyr(41) is the amino acid involved in the cross-link and that it
therefore contributes to specific DNA recognition by EcoRII.

<>

<1>Mucke, M., Reich, S., Moncke-Buchner, E., Reuter, M., Kruger, D.H.
<2>DNA cleavage by type II restriction-modification enzyme EcoP15I is independent of spacer distance between two head to head oriented recognition sites.
<3>J. Mol. Biol.
<4>312
<5>687-698
<6>2001
<7>The type III restriction-modification enzyme EcoP15I requires the interaction of two
unmethylated, inversely oriented recognition sites 5'-CAGCAG in head to head configuration to
allow an efficient DNA cleavage. It has been hypothesized that two convergent
DNA-translocating enzyme-substrate complexes interact to form the active cleavage complex and
that translocation is driven by ATP hydrolysis. Using a half-automated, fluorescence-based
detection method, we investigated how the distance between two inversely oriented recognition
sites affects DNA cleavage efficiency. We determined that EcoP15I cleaves DNA efficiently even
for two adjacent head to head or tail to tail oriented target sites. Hence, DNA translocation
appears not to be required for initiating DNA cleavage in these cases. Furthermore, we report
here that EcoP15I is able to cleave single-site substrates. When we analyzed the interaction
of EcoP15I with DNA substrates containing adjacent target sites in the presence of
non-hydrolyzable ATP analogues, we found that cleavage depended on the hydrolysis of ATP.
Moreover, we show that cleavage occurs at only one of the two possible cleavage positions of
an interacting pair of target sequences. When EcoP15I bound to a DNA substrate containing one
recognition site in the absence of ATP, we observed a 36 nucleotide DNaseI-footprint that is
asymmetric on both strands. All of our footprinting experiments showed that the enzyme did not
cover the region around the cleavage site. Analyzing a DNA fragment with two head to head
oriented recognition sites, EcoP15I protected 27-33 nucleotides around the recognition
sequence, including an additional region of 26 bp between both cleavage sites. For all DNA
substrates examined, the presence of ATP caused altered footprinting patterns. We assume that
the altered patterns are most likely due to a conformational change of the enzyme. Overall,
our data further refine the tracking-collision model for type III restriction enzymes.

<>

<1>Mucke, M., Reuter, M., Kruger, D.
<2>Restriction endonucleases, method of synthesis and use thereof.
<3>US Patent Office
<4>US 07101697
<5>
<6>2006
<7>A restriction endonuclease having one DNA binding site is proposed, synthesized from a
restriction endonuclease that has one C-terminal
domain and one N-terminal domain and two DNA binding sites, by
proteolytic cleavage into the two domains or by cloning the gene
segment that codes for the domains and expression of the domains and
selection of the endonucleolytic domains having one DNA binding site.
In addition, a method of synthesis of the restriction endonuclease and
its use are claimed.

<>

<1>Muckerman, C.C., Springhorn, S.S., Greenberg, B., Lacks, S.A.
<2>Transformation of restriction endonuclease phenotype in Streptococcus pneumoniae.
<3>J. Bacteriol.
<4>152
<5>183-190
<6>1982
<7>The genetic basis of the unique restriction endonuclease DpnI, that cleaves
only at a methylated sequence, 5'-GmeATC-3', and of the complementary
endonuclease DpnII, which cleaves at the same sequence when it is not
methylated, was investigated.  Different strains of Streptococcus pneumoniae
isolated from patients contained either DpnI (two isolates) or DpnII (six
isolates).  The latter strains also contained DNA methylated at the 5'-GATC-3'
sequence.  A restrictable bacteriophage, HB-3, was used to characterize the
various strains and to select for transformants.  One laboratory strain
contained neither DpnI nor DpnII.  It was probably derived from a
DpnI-containing strain, and its DNA was not methylated at 5'-GATC-3'.  Cells of
this strain were transformed to the DpnI restriction phenotype by DNA from a
DpnI-containing strain and to the DpnII restriction phenotype by DNA from
DpnII-containing strain.  Neither cross-transformation, that is, transformation
to one phenotype by DNA from a strain of the other phenotype, nor spontaneous
conversion was observed.  Extracts of transformants to the new restriction
phenotype were shown to contain the corresponding endonuclease.

<>

<1>Mueller, J.E., Bryk, M., Loizos, N., Belfort, M.
<2>Homing endonucleases.
<3>Nucleases, Cold Spring Harbor Laboratory Press, Linn, S.M., Lloyd, R.S., Roberts, R.J., Cold Spring Harbor
<4>0
<5>111-143
<6>1993
<7>*
I. Introduction
II. Historic review
III. General characteristics of homing endonucleases
        A. Endonuclease-mediated homing
        B. Endonuclease ORF location
        C. Endonuclease expression
        D. Sequence motifs
                1. The LAGLI-DADG motif
                2. The GIY-YIG motif
                3. The zinc finger motif
                4. Unclassified
        E. Target recognition and cleavage specificity
                1. General properties
                2. Genetic studies
                3. Physical studies
IV. Evolutionary considerations
        A. Evidence for mobility of endonuclease genes
        B. Invasion and its aftermath
        C. Endonuclease properties that potentiate invasiveness
        D. Down-regulation of endonuclease expression
        E. Cross-species transfer


<>

<1>Mueller, J.E., Clyman, J., Huang, Y.-J., Parker, M.M., Belfort, M.
<2>Intron mobility in phage T4 occurs in the context of recombination-dependent DNA replication by way of multiple pathways.
<3>Genes Dev.
<4>10
<5>351-364
<6>1996
<7>Numerous group I introns in both prokaryotes and euykaryotes behave as mobile genetic
elements.  The functional requirements for intron mobility were determined in the T4 phage
system using an in vivo assay to measure intron homing with wild-type and mutant derivatives.
Thus, it was demonstrated that intron mobility occurs in the context of phage
recombination-dependent replication, a pathway that uses overlapping subsets of replication
and recombination functions.  The functional requirements for intron homing and the nature of
recombinant products are only partially consistent with the accepted double-strand-break
repair model for intron inheritance, and implicate additional homing pathways.  Whereas
ambiguities in resolvase requirements and underrepresentation of crossover recombination
products are difficult to rationalize strictly by DSBR, these properties are most readily
consistent with a synthesis-dependent strand annealing pathway.  The pathways share common
features in the strand invasion steps, but differ in subsequent repair synthesis and
resolution steps, influencing the genetic consequences of the intron transfer event.

<>

<1>Mueller, J.E., Smith, D., Belfort, M.
<2>Exon coconversion biases accompanying intron homing: battle of the nucleases.
<3>Genes Dev.
<4>10
<5>2158-2166
<6>1996
<7>Intron homing in phage T4 occurs in the context of recombination-dependent replication, by
virtue of intron-encoded endonucleolytic activity.  After the td intron endonuclease I-TevI
cleaves the intronless recipient 23 and 25 nucleotides upstream of the intron insertion site,
exonucleolytic degradation is required for recombination to proceed.  This resection process
results in coconversion of exon sequences flanking the intron.  In a genetic system designed
to study coconversion of flanking markers, we demonstrate that although there is a
bidirectional polarity gradient, coconversion can be highly asymmetric.  Furthermore, we show
that the coconversion of flanking markers favors exon I sequences, upstream of the I-TevI
cleavage site.  These data are consistent with the asymmetric features of the homing pathways
that have been invoked for intron mobility in phage T4.  Moreover, these results are in accord
with the finding that once the td homing-site substrate is cleaved, I-TevI remains bound to
the downstream cleavage product, protecting against exonucleolytic degradation, and thereby
limiting the extent of coconversion into exon II.  The results suggest that recombination
events are influenced by a competition between the homing endonuclease and exonucleases for
sequences downstream of the I-TevI cleavage site, thereby implying a role for the homing
endonuclease in the repair process.

<>

<1>Mueller, J.E., Smith, D., Bryk, M., Belfort, M.
<2>Intron-encoded endonuclease I-TevI binds as a monomer to effect sequential cleavage via conformational changes in the td homing site.
<3>EMBO J.
<4>14
<5>5724-5735
<6>1995
<7>I-TevI, the intron-encoded endonuclease from the thymidylate synthase (td) gene of
bacteriophage T4, binds its DNA substrate across the minor groove in a sequence-tolerant
fashion.  We demonstrate here that the 28 kDa I-TevI binds the extensive 37 bp td homing site
as a monomer and significantly distorts its substrate.  In situ cleavage assays and phasing
analyses indicate that upon nicking the bottom strand of the td homing site, I-TevI induces a
directed bend of 38o towards the major groove near the cleavage site.  Formation of the bent
I-TevI-DNA complex is proposed to promote top-strand cleavage of the homing site.
Furthermore, reductions in the degree of distortion and in the efficiency of binding
base-substitution variants of the td homing site indicate that sequences flanking the cleavage
site contribute to the I-TevI-induced conformational change.  These results, combined with
genetic, physical and computer-modeling studies, form the basis of a model, wherein I-TevI
acts as a hinged monomer to induce a distortion that widens the minor groove, facilitating
access to the top-strand cleavage site.  The model is compatible with both unmodified DNA and
glucosylated hydroxymethylcytosine-containing DNA, as exists in the T-even phages.

<>

<1>Muhd, S.M.K., Abdul, R.A.Y., Saito, J.A., Hou, S., Alam, M.
<2>Complete Genome Sequence of the Thermophilic Bacterium Geobacillus thermoleovorans CCB_US3_UF5.
<3>J. Bacteriol.
<4>194
<5>1239
<6>2012
<7>Geobacillus thermoleovorans CCB_US3_UF5 is a thermophilic bacterium isolated from a hot spring
in Malaysia. Here, we report the complete genome of G.
thermoleovorans CCB_US3_UF5, which shows high similarity to the genome of
Geobacillus kaustophilus HTA 426 in terms of synteny and orthologous genes.

<>

<1>Muir, R.S., Flores, H., Zinder, N.D., Model, P., Soberon, X., Heitman, J.
<2>Temperature-sensitive mutants of the EcoRI endonuclease.
<3>J. Mol. Biol.
<4>274
<5>722-737
<6>1997
<7>The EcoRI endonuclease is an important recombinant DNA tool and a paradigm of
sequence-specific DNA-protein interactions.  We have isolated temperature sensitive EcoRI
endonuclease mutants (R56Q, G78D, P90S, V97I, R105K, M157I, C218Y, A235E, M255I, T261I and
L263F) and characterized activity in vivo and in vitro.  Although the majority were TS for
function in vivo, all of the mutant enzymes were stably expressed and largely soluble at both
30oC and 42oC in vivo and none of the mutants was found to be TS in vitro.  These findings
suggest that these mutations may affect folding of the enzyme at elevated temperature in vivo.
Both non-conservative and conservative substitutions occurred but were not correlated with
severity of the mutation.  Of the 12 residues identified, 11 are conserved between EcoRI and
the isoschizomer RsrI (which shares 50% identity), a further indication that these residues
are critical for EcoRI structure and function.  Inspection of the 2.8 A resolution X-ray
crystal structure of the wild-type EcoRI endonuclease-DNA complex revealed that: (1) the TS
mutations cluster in one half of the globular enzyme; (2) several of the substituted residues
interact with each other; (3) most mutations would be predicted to disrupt local structures;
(4) two mutations may affect the dimer interface (G78D and A235E); (5) one mutation (P90S)
occurred in a residue that is part of, or immediately adjacent to, the EcoRI active site and
which is conserved in the distantly related EcoRV endonuclease.  Finally, one class of mutants
restricted phage in vivo and was active in vitro, whereas a second class did not restrict and
was inactive in vitro.  The two classes of mutants may differ in kinetic properties or
cleavage mechanism.  In summary, these mutations provide insights into EcoRI structure and
function, and complement previous genetic, biochemical, and structural analyses.

<>

<1>Mukherjee, U., Kumar, R., Mahato, N.K., Khurana, J.P., Lal, R.
<2>Draft Genome Sequence of Sphingobium sp. Strain HDIPO4, an Avid Degrader of Hexachlorocyclohexane.
<3>Genome Announcements
<4>1
<5>e00749-13
<6>2013
<7>Sphingobium sp. strain HDIPO4 was isolated from a hexachlorocyclohexane (HCH) dumpsite and
degraded HCH isomers rapidly. The draft genome sequence of HDIPO4
(~4.7 Mbp) contains 143 contigs and 4,646 coding sequences with a G+C content of
65%.

<>

<1>Mukherjee, U., Saxena, A., Kumari, R., Singh, P., Lal, R.
<2>Draft Genome Sequence of Amycolatopsis mediterranei DSM 40773, a Tangible Antibiotic Producer.
<3>Genome Announcements
<4>2
<5>e00752-14
<6>2014
<7>Amycolatopsis mediterranei DSM 40773 has been of special interest as successors of this strain
are in use for the commercial production of rifamycin B. Here we
present the draft genome sequence (~10 Mb) of this strain, which contains 108
contigs, 9,198 genes, and has a G+C content of 71.3%.

<>

<1>Mukhopadhyay, A.K., Kersulyte, D., Jeong, J.Y., Datta, S., Ito, Y., Chowdhury, A., Chowdhury, S., Santra, A., Bhattacharya, S.K., Azuma, T., Nair, G.B., Berg, D.E.
<2>Distinctiveness of genotypes of Helicobacter pylori in Calcutta India.
<3>J. Bacteriol.
<4>182
<5>3219-3227
<6>2000
<7>The genotypes of 78 strains of Helicobacter pylori from Calcutta, India (55 from ulcer
patients and 23 from more-benign infections), were studied, with a focus on putative virulence
genes and neutral DNA markers that were likely to be phylogenetically informative. PCR tests
indicated that 80 to 90% of Calcutta strains carried the cag pathogenicity island (PAI) and
potentially toxigenic vacAs1 alleles of the vacuolating cytotoxin gene (vacA), independent of
disease status. This was higher than in the West (where cag PAI(+) vacAs1 genotypes are
disease associated) but lower than in east Asia. The iceA2 gene was weakly disease associated
in Calcutta, whereas in the West the alternative but unrelated iceA1 gene at the same locus is
weakly disease associated. DNA sequence motifs of vacAm1 (middle region) alleles formed a
cluster that was distinct from those of east Asia and the West, whereas the cagA sequences of
Calcutta and Western strains were closely related. An internal deletion found in 20% of
Calcutta iceA1 genes was not seen in any of approximately 200 strains studied from other
geographic regions and thus seemed to be unique to this H. pylori population. Two mobile DNAs
that were rare in east Asian strains were also common in Calcutta. About 90% of Calcutta
strains were metronidazole resistant. These findings support the idea that H. pylori gene
pools differ regionally and emphasize the potential importance of studies of Indian and other
non-Western H. pylori populations in developing a global understanding of this gastric
pathogen and associated disease.

<>

<1>Mukhopadhyay, P., Roy, K.B.
<2>Protein engineering of BamHI restriction endonuclease: replacement of Cys54 by Ala enhances catalytic activity.
<3>Protein Eng.
<4>11
<5>931-935
<6>1998
<7>Chemical modification studies of BamHI endonuclease indicated the importance of the cysteine
residue in catalysis.  Of the three cysteine residues at position 34, 54 and 64 in the BamHI
endonuclease Cys54 and Cys64 are at the DNA-protein interface.  The co-crystal structure of
the BamHI-DNA complex, however, does not indicate any role of cysteines either in binding or
catalysis.  In the context of strong biochemical evidence, Cys54 in BamHI was changed to Ala54
to investigate its role in catalysis.  The mutation was carried out by PCR overlap extension,
the mutant gene was cloned and characterized by sequencing.  The mutant BamHI was expressed
and purified to homogeneity and the kinetic parameters (KM and kcat) of the wild type and the
C54A mutant were determined.  The mutation results in up to ~40% enhancement of kcat and some
increase in KM.  These in vitro results were also supported by in vivo SOS induction assays:
the C54A mutant gene under the T7 promoter caused complete lysis in JH139 in absence of T7 RNA
polymerase whereas the wild-type gene gave deep blue colonies under the same conditions.  The
results suggest no direct role in Cys54 in catalysis, but it can influence the catalytic
activity through Val57 backbone contact seen in the co-crystal structure.

<>

<1>Mukhopadhyay, S., Thomason, M.K., Lentz, S., Nolan, N., Willner, K., Gee, J.E., Glass, M.B., Inglis, T.J., Merritt, A., Levy, A., Sozhamannan, S., Mateczun, A., Read, T.D.
<2>High-redundancy draft sequencing of 15 clinical and environmental burkholderia strains.
<3>J. Bacteriol.
<4>192
<5>6313-6314
<6>2010
<7>The Gram-negative Burkholderia genus includes several species of intracellular bacterial
pathogens that pose substantial risk to humans. In
this study, we have generated draft genome sequences of 15 strains of B.
oklahomensis, B. pseudomallei, B. thailandensis, and B. ubonensis to an
average sequence read coverage of 25- to 40-fold.

<>

<1>Mukund, M.A.
<2>Restriction and modification systems: unrestricted frontiers.
<3>Curr. Sci.
<4>65
<5>509-511
<6>1993
<7>Report on Saxton's River Meeting, July 1993

<>

<1>Mulder, C., Delius, H.
<2>Specificity of the break produced by restricting endonuclease R in Simian virus 40 DNA, as revealed by partial denaturation mapping.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>3215-3219
<6>1972
<7>Superhelical circular (form 1) SV40 DNA was converted to linear molecules by
the action of a partially purified restriction enzyme of resistance transfer
factor-R of Escherichia coli.  The resulting linear DNA molecules are full
length, as judged by their sedimentation through alkaline sucrose gradient and
by direct observation in an electron microscope.  Nicked circular (form II) DNA
was found as an intermediate in the conversion of form I DNA to linear DNA.
Analysis of partial denaturation maps obtained by alkaline denaturation of the
unit-length linear molecules showed that the break in SV40 DNA occurred at a
specific site on the DNA.

<>

<1>Muller, D. et al.
<2>A tale of two oxidation states: bacterial colonization of arsenic-rich environments.
<3>PLoS Genet.
<4>3
<5>e53
<6>2007
<7>Microbial biotransformations have a major impact on contamination by toxic elements, which
threatens public health in developing and industrial
countries. Finding a means of preserving natural environments-including
ground and surface waters-from arsenic constitutes a major challenge
facing modern society. Although this metalloid is ubiquitous on Earth,
thus far no bacterium thriving in arsenic-contaminated environments has
been fully characterized. In-depth exploration of the genome of the
beta-proteobacterium Herminiimonas arsenicoxydans with regard to
physiology, genetics, and proteomics, revealed that it possesses
heretofore unsuspected mechanisms for coping with arsenic. Aside from
multiple biochemical processes such as arsenic oxidation, reduction, and
efflux, H. arsenicoxydans also exhibits positive chemotaxis and motility
towards arsenic and metalloid scavenging by exopolysaccharides. These
observations demonstrate the existence of a novel strategy to efficiently
colonize arsenic-rich environments, which extends beyond oxidoreduction
reactions. Such a microbial mechanism of detoxification, which is possibly
exploitable for bioremediation applications of contaminated sites, may
have played a crucial role in the occupation of ancient ecological niches
on earth.

<>

<1>Muller, E.E., Pinel, N., Gillece, J.D., Schupp, J.M., Price, L.B., Engelthaler, D.M., Levantesi, C., Tandoi, V., Luong, K., Baliga, N.S., Korlach, J., Keim, P.S., Wilmes, P.
<2>Genome Sequence of 'Candidatus Microthrix parvicella' Bio17-1, a Long-Chain-Fatty-Acid-Accumulating Filamentous Actinobacterium from a Biological    Wastewater Treatment Plant.
<3>J. Bacteriol.
<4>194
<5>6670-6671
<6>2012
<7>'Candidatus Microthrix' bacteria are deeply branching filamentous actinobacteria  which
occur at the water-air interface of biological wastewater treatment plants,
where they are often responsible for foaming and bulking. Here, we report the
first draft genome sequence of a strain from this genus: 'Candidatus Microthrix
parvicella' strain Bio17-1.

<>

<1>Muller, H., Furnkranz, M., Grube, M., Berg, G.
<2>Genome Sequence of Serratia plymuthica Strain S13, an Endophyte with Germination- and Plant-Growth-Promoting Activity from the Flower of Styrian Oil Pumpkin.
<3>Genome Announcements
<4>1
<5>e00594-13
<6>2013
<7>The bacterium Serratia plymuthica strain S13 was demonstrated to colonize various
plant-associated microhabitats and to suppress damping-off diseases. The
completed genome sequence has a size of 5.5 Mb, containing 4,957 putative
protein-encoding regions, and will be used to identify genetic determinants
enabling the bacterium to escort a plant's entire life cycle.

<>

<1>Muller, H., Zachow, C., Alavi, M., Tilcher, R., Krempl, P.M., Thallinger, G.G., Berg, G.
<2>Complete Genome Sequence of the Sugar Beet Endophyte Pseudomonas poae RE*1-1-14,  a Disease-Suppressive Bacterium.
<3>Genome Announcements
<4>1
<5>e0002013
<6>2013
<7>The endophytic bacterium Pseudomonas poae RE*1-1-14 shows broad antagonistic activity and is
applied to seeds as a biocontrol agent to suppress late root rot
in the sugar beet. The completely sequenced 5.5-Mb genome reveals genes that
putatively contribute to this antagonistic activity and the intimate
plant-microbe interaction.

<>

<1>Muller, I., Kube, M., Reinhardt, R., Jelkmann, W., Geider, K.
<2>The Complete Genome Sequences of three Erwinia amylovora Phages Isolated in North America and a Bacteriophage Induced from an Erwinia tasmaniensis  Strain.
<3>J. Bacteriol.
<4>193
<5>795-796
<6>2010
<7>Fire blight, a plant disease of economical importance caused by Erwinia amylovora, may be
controlled by application of bacteriophages. Here we
provide the complete genome sequences and the annotation of three E.
amylovora-specific phages isolated in North America and genomic
information about a bacteriophage induced by mitomycin C-treatment of an
E. tasmaniensis strain, antagonistic for E. amylovora. The American phages
resemble two already described viral genomes, whereas the E. tasmaniensis
phage displays a singular genomic sequence in BLAST searches.

<>

<1>Muller, R., Wenzel, S., Garcia, R.
<2>Synthetic pathway enzymes for the production of Argyrins.
<3>European Patent Office
<4>EP 2141242 A
<5>
<6>2010
<7>The invention provides the amino acid sequences comprised in or constituting the synthetic
pathway enzymes participating in the production of Argyrins, as well as the nucleic acid
sequences encoding the synthetic pathway enzymes participating in the production of Argyrins,
as well as genetically manipulated micro-organisms containing nucleic acid sequences encoding
the synthetic pathway enzymes for the production of Argyrins, e.g. for inserting one or more
of these coding sequences, mutating in a targeted manner one or more of these nucleic acid
sequences, in a wild type producer micro-organism or in a heterologous micro-organism, for the
production of Argyrins.

<>

<1>Muller, S., Willett, J.W., Bahr, S.M., Darnell, C.L., Hummels, K.R., Dong, C.K., Vlamakis, H.C., Kirby, J.R.
<2>Draft Genome Sequence of Myxococcus xanthus Wild-Type Strain DZ2, a Model Organism for Predation and Development.
<3>Genome Announcements
<4>1
<5>e00217-13
<6>2013
<7>Myxococcus xanthus is a member of the Myxococcales order within the Deltaproteobacteria
subdivision. The myxobacteria reside in soil, have relatively
large genomes, and display complex life cycles. Here, we report the whole-genome
shotgun sequence of strain DZ2, which includes unique genes not found previously
in strain DK1622.

<>

<1>Muller, S., Willett, J.W., Bahr, S.M., Scott, J.C., Wilson, J.M., Darnell, C.L., Vlamakis, H.C., Kirby, J.R.
<2>Draft Genome of a Type 4 Pilus Defective Myxococcus xanthus Strain, DZF1.
<3>Genome Announcements
<4>1
<5>e00392-13
<6>2013
<7>Myxococcus xanthus is a member of the Myxococcales order within the deltaproteobacterial
subdivision. Here, we report the whole-genome shotgun
sequence of the type IV pilus (T4P) defective strain DZF1, which includes many
genes found in strain DZ2 but absent from strain DK1622.

<>

<1>Mulligan, E.A., Dunn, J.J.
<2>Cloning, purification and initial characterization of E. coli McrA, a putative 5-methylcytosine-specific nuclease.
<3>Protein Expr. Purif.
<4>62
<5>98-103
<6>2008
<7>Expression strains of Escherichia coli BL21(DE3) overproducing the E. coli m(5)C McrA
restriction protein were produced by cloning the mcrA coding
sequence behind a T7 promoter. The recombinant mcrA minus BL21(DE3) host
produces active McrA as evidenced by its acquired ability to selectively
restrict the growth of T7 phage containing DNA methylated in vitro by
HpaII methylase. The mcrA coding region contains several non-optimal E.
coli triplets. Addition of the pACYC-RIL tRNA encoding plasmid to the
BL21(DE3) host increased the yield of recombinant McrA (rMcrA) upon
induction about 5- to 10-fold. McrA protein expressed at 37 degrees C is
insoluble but a significant fraction is recovered as soluble protein after
autoinduction at 20 degrees C. rMcrA protein, which is predicted to
contain a Cys(4)-Zn(2+) finger and a catalytically important histidine
triad in its putative nuclease domain, binds to several metal chelate
resins without addition of a poly-histidine affinity tag. This feature was
used to develop an efficient protocol for the rapid purification of nearly
homogeneous rMcrA. The native protein is a dimer with a high alpha-helical
content as measured by circular dichroism analysis. Under all conditions
tested purified rMcrA does not have measurable nuclease activity on HpaII
methylated (Cm(5)CGG) DNA, although the purified protein does specifically
bind HpaII methylated DNA. These results have implications for
understanding the in vivo activity of McrA in "restricting"
m(5)C-containing DNA and suggest that rMcrA may have utility as a reagent
for affinity purification of DNA fragments containing m(5)C residues.

<>

<1>Mulligan, E.A., Hatchwell, E., McCorkle, S.R., Dunn, J.J.
<2>Differential binding of Escherichia coli McrA protein to DNA sequences that contain the dinucleotide m5CpG.
<3>Nucleic Acids Res.
<4>38
<5>1997-2005
<6>2010
<7>The Escherichia coli McrA protein, a putative C(5)-methylcytosine/C(5)-hydroxyl
methylcytosine-specific nuclease, binds
DNA with symmetrically methylated HpaII sequences (Cm5CGG), but its
precise recognition sequence remains undefined. To determine McrA's
binding specificity, we cloned and expressed recombinant McrA with a
C-terminal StrepII tag (rMcrA-S) to facilitate protein purification and
affinity capture of human DNA fragments with m5C residues. Sequence
analysis of a subset of these fragments and electrophoretic mobility shift
assays with model methylated and unmethylated oligonucleotides suggest
that N(Y > R) m5CGR is the canonical binding site for rMcrA-S. In addition
to binding HpaII-methylated double-stranded DNA, rMcrA-S binds DNA
containing a single, hemimethylated HpaII site; however, it does not bind
if A, C, T or U is placed across from the m5C residue, but does if I is
opposite the m5C. These results provide the first systematic analysis of
McrA's in vitro binding specificity.

<>

<1>Mullineux, S.T., Costa, M., Bassi, G.S., Michel, F., Hausner, G.
<2>A group II intron encodes a functional LAGLIDADG homing endonuclease and self-splices under moderate temperature and ionic conditions.
<3>RNA
<4>16
<5>1818-1831
<6>2010
<7>A group II intron encoding a protein belonging to the LAGLIDADG family of homing endonucleases
was identified in the mitochondrial rns gene of
the filamentous fungus Leptographium truncatum, and the catalytic
activities of both the intron and its encoded protein were
characterized. A model of the RNA secondary structure indicates that
the intron is a member of the IIB1 subclass and the open reading frame
is inserted in ribozyme domain III. In vitro assays carried out with
two versions of the intron, one in which the open reading frame was
removed and the other in which it was present, demonstrate that both
versions of the intron readily self-splice at 37 degrees C and at a
concentration of MgCl2 as low as 6 mM. The open reading frame encodes a
functional LAGLIDADG homing endonuclease that cleaves 2 (top strand)
and 6 (bottom strand) nucleotides (nt) upstream of the intron insertion
site, generating 4 nt 3' OH overhangs. In vitro splicing assays carried
out in the absence and presence of the intron-encoded protein indicate
that the protein does not enhance intron splicing, and RNA-binding
assays show that the protein does not appear to bind to the intron RNA
precursor transcript. These findings raise intriguing questions
concerning the functional and evolutionary relationships of the two
components of this unique composite element.

<>

<1>Mullings, R., Bennett, S.P., Brown, N.L.
<2>Investigation of sequence homology in a group of type-II restriction/modification isoschizomers.
<3>Gene
<4>74
<5>245-251
<6>1988
<7>We have dissected the cloned PstI M and R genes to make DNA hybridization
probes spanning most of the sequence.  These subclones, and also the intact
sequence, were used to search for nucleic acid homology by Southern blot in the
DNA from twelve organisms which produce PstI isoschizomers.  One of these
probes, a 206-bp fragment from the N-terminal domain of the endonuclease,
showed significant hybridisation in four strains (Escherichia coli strains
RFL48, RFL49 and RFL83, and Streptomyces albus P).  No significant
hybridisation was detected with other parts  of the PstI proteins and the known
sequences of other type-II systems that recognise different sites.  We
postulate a possible recognition domain within the M.PstI methyltransferase
based on similarity to the M.PaeR7 and M.TaqI methyltransferases.

<>

<1>Mullings, R., Evans, L.R., Brown, N.L.
<2>Type II restriction endonucleases from Bacillus sphaericus.
<3>FEMS Microbiol. Lett.
<4>37
<5>237-240
<6>1986
<7>We report the isolation and characterisation of 3 Type II restriction
endonucleases from Bacillus sphaericus.  These are BspAI (an isoschizomer of
Sau3AI) in strain JL4B; BspBI and BspBII (isoschizomers of PstI and AsuI) in
strain JL14.  These are the first reports of these activities in B. sphaericus
and the first citation of an AsuI isoschizomer in a member of the genus
Bacillus.  We briefly discuss the possible uses in vitro and significance in
vivo of these enzymes.

<>

<1>Mullins, M.A., Register, K.B., Bayles, D.O., Dyer, D.W., Kuehn, J.S., Phillips, G.J.
<2>Genome sequence of Haemophilus parasuis strain 29755.
<3>Standards in Genomic Sciences
<4>5
<5>61-68
<6>2011
<7>Haemophilus parasuis is a member of the family Pasteurellaceae and is the etiologic agent of
Glasser's disease in pigs, a systemic syndrome associated with
only a subset of isolates. The genetic basis for virulence and systemic spread of
particular H. parasuis isolates is currently unknown. Strain 29755 is an invasive
isolate that has long been used in the study of Glasser's disease. Accordingly,
the genome sequence of strain 29755 is of considerable importance to
investigators endeavoring to understand the molecular pathogenesis of H.
parasuis. Here we describe the features of the 2,224,137 bp draft genome sequence
of strain 29755 generated from 454-FLX pyrosequencing. These data comprise the
first publicly available genome sequence for this bacterium.

<>

<1>Mun, B.J.
<2>Mutant restriction enzyme EcoRV.
<3>Korean Patent Office
<4>KR 192599 B
<5>
<6>1999
<7>
<>

<1>Mund, C., Musch, T., Strodicke, M., Assmann, B., Li, E., Lyko, F.
<2>Comparative analysis of DNA methylation patterns in transgenic Drosophila overexpressing mouse DNA methyltransferases.
<3>Biochem. J.
<4>378
<5>763-768
<6>2003
<7>DNA methyltransferases (Dnmts) mediate the epigenetic modification of eukaryotic genomes.
Mammalian DNA methylation patterns are established and
maintained by co-operative interactions among the Dnmt proteins Dnmt1,
Dnmt3a and Dnmt3b. Owing to their simultaneous presence in mammalian
cells, the activities of individual Dnmt have not yet been determined.
This includes a fourth putative Dnmt, namely Dnmt2, which has failed to
reveal any activity in previous assays. We have now established transgenic
Drosophila strains that allow for individual overexpression of all known
mouse Dnmts. Quantitative analysis of genomic cytosine methylation levels
demonstrated a robust Dnmt activity for the de novo methyltransferases
Dnmt3a and Dnmt3b. In addition, we also detected a weak but significant
activity for Dnmt2. Subsequent methylation tract analysis by genomic
bisulphite sequencing revealed that Dnmt3 enzymes preferentially
methylated CpG dinucleotides in a processive manner, whereas Dnmt2
methylated isolated cytosine residues in a non-CpG dinucleotide context.
Our results allow a direct comparison of the activities of mammalian Dnmts
and suggest a significant functional specialization of these enzymes.

<>

<1>Mundo, S.L., Gilardoni, L.R., Hoffman, F.J., Lopez, O.J.
<2>Rapid and Sensitive Method To Identify Mycobacterium avium subsp paratuberculosis in Cow's Milk by DNA Methylase Genotyping.
<3>Appl. Environ. Microbiol.
<4>79
<5>1612-1618
<6>2013
<7>Paratuberculosis is an infectious, chronic, and incurable disease that affects ruminants,
caused by Mycobacterium avium subsp. paratuberculosis. This bacterium is shed primarily
through feces of infected cows but can be also excreted in colostrum and milk and might
survive pasteurization. Since an association of genomic sequences of M. avium subsp.
paratuberculosis in patients with Crohn's disease has been described; it is of interest to
rapidly detect M. avium subsp. paratuberculosis in milk for human consumption. IS900 insertion
is used as a target for PCR amplification to identify the presence of M. avium subsp.
paratuberculosis in biological samples. Two target sequences were selected: IS1 (155 bp) and
IS2 (94 bp). These fragments have a 100% identity among all M. avium subsp. paratuberculosis
strains sequenced. M. avium subsp. paratuberculosis was specifically concentrated from milk
samples by immunomagnetic separation prior to performing PCR. The amplicons were characterized
using DNA methylase Genotyping, i.e., the amplicons were methylated with 6-methyl-adenine and
digested with restriction enzymes to confirm their identity. The methylated amplicons from 100
CFU of M. avium subsp. paratuberculosis can be visualized in a Western blot format using an
anti-6-methyl-adenine monoclonal antibody. The use of DNA methyltransferase genotyping coupled
to a scintillation proximity assay allows for the detection of up to 10 CFU of M. avium subsp.
paratuberculosis per ml of milk. This test is rapid and sensitive and allows for automation
and thus multiple samples can be tested at the same time.

<>

<1>Muniesa, M., Recktenwald, J., Bielaszewska, M., Karch, H., Schmidt, H.
<2>Characterization of a shiga toxin 2e-converting bacteriophage from an Escherichia coli strain of human origin.
<3>Infect. Immun.
<4>68
<5>4850-4855
<6>2000
<7>An infectious Shiga toxin (Stx) 2e-converting bacteriophage (phiP27) was
isolated from Stx2e-producing Escherichia coli ONT:H(-) isolate 2771/97
originating from a patient with diarrhea. The phage could be transduced to
E. coli laboratory strain DH5alpha, and we could show that lysogens were
able to produce biologically active toxin in a recA-dependent manner. By
DNA sequence analysis of a 6,388-bp HindIII restriction fragment of
phiP27, we demonstrated that the stx(2e) gene was located directly
downstream of ileZ and argO tRNA genes. Although no analogue of an
antiterminator Q encoding gene was present on this fragment, a lysis
cassette comprising two holin genes which are related to the holin genes
of Pseudomonas aeruginosa phage phiCTX and a gene homologous to the
endolysin gene gp19 of phage PS3 were detected. The results of our study
demonstrated for the first time that Stx2e can be encoded in the genome of
an infectious bacteriophage.

<>

<1>Munk, A.C. et al.
<2>Complete genome sequence of Tsukamurella paurometabola type strain (no. 33).
<3>Standards in Genomic Sciences
<4>4
<5>342-351
<6>2011
<7>Tsukamurella paurometabola corrig. (Steinhaus 1941) Collins et al. 1988 is the type species of
the genus Tsukamurella, which is the type genus to the family
Tsukamurellaceae. The species is not only of interest because of its isolated
phylogenetic location, but also because it is a human opportunistic pathogen with
some strains of the species reported to cause lung infection, lethal meningitis,
and necrotizing tenosynovitis. This is the first completed genome sequence of a
member of the genus Tsukamurella and the first genome sequence of a member of the
family Tsukamurellaceae. The 4,479,724 bp long genome contains a 99,806 bp long
plasmid and a total of 4,335 protein-coding and 56 RNA genes, and is a part of
the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Munk, A.C. et al.
<2>Complete genome sequence of Rhodospirillum rubrum type strain (S1).
<3>Standards in Genomic Sciences
<4>4
<5>293-302
<6>2011
<7>Rhodospirillum rubrum (Esmarch 1887) Molisch 1907 is the type species of the genus
Rhodospirillum, which is the type genus of the family Rhodospirillaceae in
the class Alphaproteobacteria. The species is of special interest because it is
an anoxygenic phototroph that produces extracellular elemental sulfur (instead of
oxygen) while harvesting light. It contains one of the most simple photosynthetic
systems currently known, lacking light harvesting complex 2. Strain S1(T) can
grow on carbon monoxide as sole energy source. With currently over 1,750 PubMed
entries, R. rubrum is one of the most intensively studied microbial species, in
particular for physiological and genetic studies. Next to R. centenum strain SW,
the genome sequence of strain S1(T) is only the second genome of a member of the
genus Rhodospirillum to be published, but the first type strain genome from the
genus. The 4,352,825 bp long chromosome and 53,732 bp plasmid with a total of
3,850 protein-coding and 83 RNA genes were sequenced as part of the DOE Joint
Genome Institute Program DOEM 2002.

<>

<1>Munk, C. et al.
<2>Complete genome sequence of Stackebrandtia nassauensis type strain (LLR-40K-21).
<3>Standards in Genomic Sciences
<4>1
<5>234-241
<6>2009
<7>Stackebrandtia nassauensis Labeda and Kroppenstedt (2005) is the type species of  the genus
Stackebrandtia, and a member of the actinobacterial family
Glycomycetaceae. Stackebrandtia currently contains two species, which are
differentiated from Glycomyces spp. by cellular fatty acid and menaquinone
composition. Strain LLR-40K-21(T) is Gram-positive, aerobic, and nonmotile, with
a branched substrate mycelium and on some media an aerial mycelium. The strain
was originally isolated from a soil sample collected from a road side in Nassau,
Bahamas. Here we describe the features of this organism, together with the
complete genome sequence and annotation. This is the first complete genome
sequence of the actinobacterial suborder Glycomycineae. The 6,841,557 bp long
single replicon genome with its 6487 protein-coding and 53 RNA genes is part of
the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Munoz, I.G. et al.
<2>Molecular basis of engineered meganuclease targeting of the endogenous human RAG1 locus.
<3>Nucleic Acids Res.
<4>39
<5>729-743
<6>2011
<7>Homing endonucleases recognize long target DNA sequences generating an accurate double-strand
break that promotes gene targeting through
homologous recombination. We have modified the homodimeric I-CreI
endonuclease through protein engineering to target a specific DNA sequence
within the human RAG1 gene. Mutations in RAG1 produce severe combined
immunodeficiency (SCID), a monogenic disease leading to defective immune
response in the individuals, leaving them vulnerable to infectious
diseases. The structures of two engineered heterodimeric variants and one
single-chain variant of I-CreI, in complex with a 24-bp oligonucleotide of
the human RAG1 gene sequence, show how the DNA binding is achieved through
interactions in the major groove. In addition, the introduction of the
G19S mutation in the neighborhood of the catalytic site lowers the
reaction energy barrier for DNA cleavage without compromising DNA
recognition. Gene-targeting experiments in human cell lines show that the
designed single-chain molecule preserves its in vivo activity with higher
specificity, further enhanced by the G19S mutation. This is the first time
that an engineered meganuclease variant targets the human RAG1 locus by
stimulating homologous recombination in human cell lines up to 265 bp away
from the cleavage site. Our analysis illustrates the key features for a la
carte procedure in protein-DNA recognition design, opening new
possibilities for SCID patients whose illness can be treated ex vivo.

<>

<1>Munson, R.S. Jr., Bakaletz, L.O., Dyer, D.W.
<2>GENES OF AN OTITIS MEDIA ISOLATE OF NONTYPEABLE HAEMOPHILUS INFLUENZAE.
<3>Japanese Patent Office
<4>JP 2010279359 A
<5>
<6>2010
<7>
<>

<1>Munson, R.S., Harrison, A., Gillaspy, A., Ray, W.C., Carson, M., Armbruster, D., Gipson, J., Gipson, M., Johnson, L., Lewis, L., Dyer, D.W., Bakaletz, L.O.
<2>Partial analysis of the genomes of two nontypeable Haemophilus influenzae otitis media isolates.
<3>Infect. Immun.
<4>72
<5>3002-3010
<6>2004
<7>In 1995, The Institute for Genomic Research completed the genomic sequence of a rough
derivative of Haemophilus influenzae serotype d,
strain KW20. This sequence, though extremely useful in understanding
the basic biology of H. influenzae, has yet to provide significant
insight into our understanding of disease caused by nontypeable H.
influenzae (NTHI), because serotype d strains are not generally
pathogens. In contrast, NTHI strains are frequently mucosal pathogens
and are the primary pathogens of chronic otitis media as well as a
significant cause of acute otitis media in children. Thus, it is of
great importance to further understand their biology. We used a
DNA-based microarray approach to identify genes present in a clinical
isolate of NTHI that were absent from strain Rd. We also sequenced the
genome of a second NTHI isolate from a child with chronic otitis media
to threefold coverage and then used an array of bioinformatics tools to
identify genes present in this NTHI strain but absent from strain Rd.
These methods were complementary in approach and results. We
identified, in both strains, homologues of H. influenzae lav, an
autotransported protein of unknown function; tna4, which encodes
tryptophanase; as well as a homologue of Pasteurella multocida tsaA,
which encodes an alkyl peroxidase that may play a role in protection
against reactive oxygen species. We also identified a number of
putative restriction-modification systems, bacteriophage genes and
transposon-related genes. These data provide new insight that
complements and extends our ongoing analysis of NTHI virulence
determinants.

<>

<1>Murakami, A., Yamamoto, Y., Namba, M., Iwase, R., Yamaoka, T.
<2>Photo-cross-linked oligonucleotide duplex as a decoy-DNA for inhibition of restriction endonuclease activity.
<3>Bioorg. Chem.
<4>29
<5>223-233
<6>2001
<7>As a novel type of regulator molecule for DNA-recognizing proteins, a photo-cross-linked
oligonucleotide duplex was designed and synthesized,
The molecule regulated the activity of a restriction endonuclease by
being recognized as a substrate. This type of regulating molecule is
regarded as a decoy-DNA. 4,5',8-[4-Aminoethylaminomethyl]-trioxaten
(aeAMT) was conjugated with art oligodeoxyribonucleotide (ODN) at the 5'-end and the aeAMT
was cross-linked with the thymine residue of the
complementary oligonucleotide upon UVA irradiation. The terminally
cross-linked oligonucleotides, singly clipped (SC) decoy-DNA, acquired
thermal stability, An oligonucleoside phosphorothioate (OPT) was also
introduced as one or both components, yielding three types of
decoy-DNAs, SC-ODN-ODN (SC.DD), SC-OPT-ODN (SC.SD). and SC-OPT-OPT
(SC.SS). The SC decoy-DNAs inhibited the function of the restriction
endonuclease, AatII, in a sequence-specific and concentration-dependent
manner with an appreciable IC50 value (1.3 microM for SC.DD, 0.016 microM for
SC.SD. 0.002 microM for SC.SS). The SC decoy-DNAs were found to be
effective for regulating the DNA recognizing proteins.

<>

<1>Murakami, M., Mizuno, H., Yamada, Y.
<2>The restriction endonuclease GinI of Gluconobacter cerinus IFO 3260, an isoschizomer of BamHI, has a monomeric structure.
<3>Agric. Biol. Chem.
<4>54
<5>2747-2749
<6>1990
<7>Type II restriction endonucleases, which are indispensable for gene
manipulation and gene analysis, are widely found in the Procaryotae such as
bacteria and cyanobacteria.  During the course of our studies, we reported a
new restriction endonuclease ApaLI from cells of Acetobacter pasteurianus IFO
13753.  We have found that the restriction endonuclease GceinI, an isoschizomer
of BamHI, has a monomeric structure different from the BamHI endonuclease.
This paper is concerned with the purification and properties of the restriction
endonuclease GceinI of Gluconobacter cerinus.  Note this enzyme is incorrectly
called GceinI in this paper.  It should be GinI.

<>

<1>Murakami, M., Ozawa, O., Kanematsu, T., Yamada, Y.
<2>A new restriction endonuclease from Bacteroides distasonis (BdiI).
<3>Agric. Biol. Chem.
<4>54
<5>275-277
<6>1990
<7>Authors have changed the name to BdiSI to avoid confusion.

<>

<1>Murakami, M., Ozawa, O., Kanematsu, T., Yamada, Y.
<2>Characterization of restriction endonuclease CbiI, an isoschizomer of AsuII, from Clostridium bifermentans strain B-4.
<3>Nucleic Acids Res.
<4>19
<5>3458
<6>1991
<7>We have found that an extremely large amount of a restriction endonuclease, designated CbiI,
is produced within the cells of Clostridium bifermentans strain B-4.  CbiI was purified from
cell extracts by combined high performance liquid chromatographies on DEAE-Toyopearlpak 650M.
TSKgel DEAE-5PW, TSKgel HA-1000 and TSKgel G3000SW.  The purified enzyme (0.4 mg protein from
1.7 g cell extract) was homogeneous on polyacrylamide gel disc electrophoresis (PAGDE).  The
relative molecular mass of the enzyme was calculated as 49,000 daltons by gel filtration and
by SDS-PAGE.  These data indicated that CbiI has a monomeric structure.

<>

<1>Murakami, M., Ozawa, O., Kanematsu, T., Yamada, Y.
<2>Characterization of restriction endonuclease BdiSI, an isoschizomer of SfeI, from Bacteroides distasonis strain S-7.
<3>Agric. Biol. Chem.
<4>55
<5>261-263
<6>1991
<7>This paper describes the purification and properties of the restriction
endonuclease BdiSI, an isoschizomer of SfeI (CTRYAG).

<>

<1>Murakami, M., Yamada, Y.
<2>Purification, properties and recognition sequence and cleavage site determinations of restriction endonuclease from Acetobacter pasteurianus IFO 13753 (ApaLI).
<3>Agric. Biol. Chem.
<4>54
<5>1791-1796
<6>1990
<7>A new restriction endonuclease, designated as ApaLI, was purified from
cell-free extracts of Acetobacter pasteurianus IFO 13753 by streptomycin
treatment, ammonium sulfate fractionation, combined column chromatographies on
heparin-Sepharose CL-6B and DEAE-Sepharose CL-6B and Fast Protein Liquid
Chromatography on Mono Q HR 5/5.  The purified enzyme was homogenous on
polyacrylamide gel disc electrophoresis.  The molecular weight of the purified
enzyme was calculated as 26,000 daltons by gel filtration using Sephadex G-200,
and the isoelectric point of the purified enzyme was 4.8 by ampholine
sucrose-density gradient isoelectric focusing.  The purified enzyme cleaves
lambda, Ad2, SV40, M13mp18 RF I, PhiX174 RF I and pBR322 DNAs at 4, 7, 0, 0, 1
and 3 sites, respectively.  The purified enzyme worked best at 37C and pH 8.0
in a reaction mixture (50 microliters) containing 1.0 micrograms lambda DNA, 10
mM Tris-HCI, 7 mM 2-mercaptoethanol, 7 mM MgCl2 and 25 mM NaCl.  However, the
purified enzyme did not require NaCl necessarily for the enzyme reaction.  The
purified enzyme recognized the palindromic hexanucleotide DNA sequence,
5'-GTGCAC-3' and cut between G and T, producing a 5'-cohesive tetranucleotide
extension.

<>

<1>Mural, R.J.
<2>Cleavage by the restriction endonuclease Asp718, an isoschizomer of KpnI, is sensitive to Escherichia coli Dcm methylation.
<3>Nucleic Acids Res.
<4>15
<5>9085
<6>1987
<7>While attempting to use Asp718, a KpnI isoschizomer, to subclone KpnI fragments from pAn602, a
plasmid known to have two KpnI sites, I found that one of these sites was cut very poorly by
this enzyme. DNA sequence analysis showed that the poorly cut site overlapped the sequence
CCAGG, an Escherichia coli Dcm methylation site. When this plasmid DNA was propagated in E.
coli GM31 (dcm-6), a host which does not methylate this site, and digested with either Asp718
or KpnI both sites were cut to completion.

<>

<1>Murali, T.S., Paul, B., Parikh, H., Singh, R.P., Kavitha, S., Bhat, M.K., Satyamoorthy, K.
<2>Genome Sequences of Four Clinical Staphylococcus aureus Strains with Diverse Drug Resistance Profiles Isolated from Diabetic Foot Ulcers.
<3>Genome Announcements
<4>2
<5>e00204-14
<6>2014
<7>Staphylococcus aureus is a major pathogen associated with diabetic foot ulcer infections. To
gain insight into their pathogenicity and virulence potential, we
report draft genome sequences of four strains of Staphylococcus aureus, isolated
from diabetic foot ulcers, showing profiles with various degrees of resistance to
common antibiotics.

<>

<1>Murata, T., Ohnishi, M., Ara, T., Kaneko, J., Han, C.G., Li, Y.F., Takashima, K., Nojima, H., Nakayama, K., Kaji, A., Kamio, Y., Miki, T., Mori, H., Ohtsubo, E., Terawaki, Y., Hayashi, T.
<2>Complete nucleotide sequence of plasmid Rts1: implications for evolution of large plasmid genomes.
<3>J. Bacteriol.
<4>184
<5>3194-3202
<6>2002
<7>Rts1, a large conjugative plasmid originally isolated from Proteus vulgaris, is a prototype
for the IncT plasmids and exhibits pleiotropic thermosensitive phenotypes. Here we report the
complete nucleotide sequence of Rts1. The genome is 217,182 bp in length and contains 300
potential open reading frames                        (ORFs). Among these, the products of 141
ORFs, including 9 previously identified genes, displayed significant sequence similarity to
known proteins. The set of genes responsible for the conjugation function of Rts1 has been
identified. A broad array of genes related to diverse processes of DNA metabolism were also
identified. Of particular interest was the presence of tus-like genes that could be involved
in replication termination. Inspection of the overall genome organization revealed that the
Rts1 genome is composed of four large modules, providing an example of modular evolution of
plasmid genomes.

<>

<1>Murawska, E., Fiedoruk, K., Bideshi, D.K., Swiecicka, I.
<2>Complete Genome Sequence of Bacillus thuringiensis subsp. thuringiensis Strain IS5056, an Isolate Highly Toxic to Trichoplusia ni.
<3>Genome Announcements
<4>1
<5>e0010813
<6>2013
<7>The genome sequence of the entomopathogen Bacillus thuringiensis subsp. thuringiensis strain
IS5056 was determined. The chromosome is composed of
5,491,935 bp. In addition, IS5056 harbors 14 plasmids ranging from 6,880 to
328,151 bp, four of which contain nine insecticidal protein genes, cry1Aa3,
cry1Ab21, cry1Ba1, cry1Ia14, cry2Aa9, cry2Ab1, vip1, vip2, and vip3Aa10.

<>

<1>Murchie, A.I.H., Portugal, J., Lilley, D.M.J.
<2>Cleavage of a four-way DNA function by a restriction enzyme spanning the point of strand exchange.
<3>EMBO J.
<4>10
<5>713-718
<6>1991
<7>The four-way DNA junction is believed to fold in the presence of metal ions
into an X-shaped structure, in which there is pairwise coaxial stacking of
helical arms.  A restriction enzyme MboII has been used to probe this
structure.  A junction was constructed containing a recognition site for MboII
in one helical arm, positioned such that stacking of arms would result in
cleavage in a neighbouring arm.  Strong cleavage was observed, at the sites
expected on the basis of coaxial stacking.  An additional cleavage was seen
corresponding to the formation of an alternative stacking isomer, suggesting
that the two isomeric forms are in dynamic equilibrium in solution.

<>

<1>Muro-Pastor, A.M., Herrero, A., Flores, E.
<2>Sequence-specific endonucleases from the cyanobacterium Nostoc sp. ATCC 29132.
<3>FEMS Microbiol. Lett.
<4>77
<5>1-4
<6>1991
<7>The nitrogen-fixing cyanobacterium Nostoc sp. ATCC 29132 was shown to contain
two sequence-specific endonucleases.  Nsp(29132) I was an isoschizomer of
AsuII, and Nsp(29132) II was an isoschizomer of BamHI.  Nsp(29132) II was shown
to generate ends that could be ligated to those generated by BamHI.

<>

<1>Murphy, E., Mounts, W.M.
<2>Microarray for monitoring gene expression in multiple strains of Streptococcus pneumonia.
<3>International Patent Office
<4>WO 2007106407 A
<5>
<6>2007
<7>The present invention features an array capable of monitoring gene expression patterns of
multiple strains of Streptococcus pneumonia including a substrate having a plurality of
addresses, each of which has a probe disposed.

<>

<1>Murphy, J., Klumpp, J., Mahony, J., O'Connell-Motherway, M., Nauta, A., van Sinderen, D.
<2>Methyltransferases acquired by lactococcal 936-type phage provide protection against restriction endonuclease activity.
<3>BMC Genomics
<4>15
<5>831
<6>2014
<7>Background: So-called 936-type phages are among the most frequently isolated phages in dairy
facilities utilising Lactococcus lactis starter cultures. Despite extensive efforts to control
phage proliferation and decades of research, these phages continue to negatively impact cheese
production in terms of the final product quality and consequently, monetary return.Results:
Whole genome sequencing and in silico analysis of three 936-type phage genomes identified
several putative (orphan) methyltransferase (MTase)-encoding genes located within the
packaging and replication regions of the genome. Utilising SMRT sequencing, methylome analysis
was performed on all three phages, allowing the identification of adenine modifications
consistent with N-6 methyladenine sequence methylation, which in some cases could be
attributed to these phage-encoded MTases. Heterologous gene expression revealed that M.
Phi145I/M. Phi93I and M. Phi93DAM, encoded by genes located within the packaging module,
provide protection against the restriction enzymes HphI and DpnII, respectively, representing
the first functional MTases identified in members of 936-type phages.Conclusions: SMRT
sequencing technology enabled the identification of the target motifs of MTases encoded by the
genomes of three lytic 936-type phages and these MTases represent the first functional MTases
identified in this species of phage. The presence of these MTase-encoding genes on 936-type
phage genomes is assumed to represent an adaptive response to circumvent host encoded
restriction-modification systems thereby increasing the fitness of the phages in a dynamic
dairy environment.

<>

<1>Murphy, J., Mahony, J., Ainsworth, S., Nauta, A., van Sinderen, D.
<2>Bacteriophage Orphan DNA Methyltransferases: Insights from Their Bacterial Origin, Function, and Occurrence.
<3>Appl. Environ. Microbiol.
<4>79
<5>7547
<6>2013
<7>Type II DNA methyltransferases (MTases) are enzymes found ubiquitously in the prokaryotic
world, where they play important roles in several cellular processes, such as host protection
and epigenetic regulation. Three classes of type II MTases have been identified thus far in
bacteria which function in transferring a methyl group from S-adenosyl-L-methionine (SAM) to a
target nucleotide base, forming N-6-methyladenine (class I), N-4-methylcytosine (class II), or
C-5-methylcytosine (class III). Often, these MTases are associated with a cognate restriction
endonuclease (REase) to form a restriction-modification (R-M) system protecting bacterial
cells from invasion by foreign DNA. When MTases exist alone, which are then termed orphan
MTases, they are believed to be mainly involved in regulatory activities in the bacterial
cell. Genomes of various lytic and lysogenic phages have been shown to encode multi-and
mono-specific orphan MTases that have the ability to confer protection from restriction
endonuclea ses of their bacterial host(s). The ability of a phage to overcome R-M and other
phage-targeting resistance systems can be detrimental to particular biotechnological processes
such as dairy fermentations. Conversely, as phages may also be beneficial in certain areas
such as phage therapy, phages with additional resistance to host defenses may prolong the
effectiveness of the therapy. This minireview will focus on bacteriophage-encoded MTases,
their prevalence and diversity, as well as their potential origi n and function.

<>

<1>Murphy, K.C., Ritchie, J.M., Waldor, M.K., Lobner-Olesen, A., Marinus, M.G.
<2>Dam methyltransferase is required for stable lysogeny of the Shiga toxin (Stx2)-encoding bacteriophage 933W of enterohemorrhagic Escherichia coli O157 : H7.
<3>J. Bacteriol.
<4>190
<5>438-441
<6>2008
<7>Shiga toxin 2 (Stx2), one of the principal virulence factors of enterohemorrhagic Escherichia
coli, is encoded by 933W, a lambda-like
prophage. 933W prophage induction contributes to Stx2 production, and
here, we provide evidence that Dam methyltransferase is essential for
maintenance of 933W lysogeny. Our findings are consistent with the idea
that the 933W prophage has a relatively low threshold for induction,
which may promote Stx2 production during infection.

<>

<1>Murphy, M., Schmid, N.S., Bickle, T.A.
<2>Lack of regulation of the modification-dependent restriction enzyme McrBC in Escherichia coli.
<3>J. Bacteriol.
<4>184
<5>1794-1795
<6>2002
<7>Restriction alleviation (RA) by the type I restriction enzyme EcoKI is caused by treatments
that damage DNA. RA is due to proteolysis of the EcoKI HsdR subunit by the ClpXP ATP-dependent
protease. Here we show that the modification-dependent enzyme McrBC is not subject to RA,
although it is moderately sensitive to ClpAP.

<>

<1>Murphy, R.A., Boyd, E.F.
<2>Three pathogenicity islands of Vibrio cholerae can excise from the chromosome and form circular intermediates.
<3>J. Bacteriol.
<4>190
<5>636-647
<6>2008
<7>Vbrio pathogenicity island-2 (VPI-2) is a 57-kb region integrated at a transfer RNA
(tRNA)-serine locus that encompasses VC1758 to VC1809 on
the V. cholerae N16961 genome and is present in pandemic isolates.
VPI-2 encodes a P4-like integrase, a restriction modification system, a
Mu phage-like region, and a sialic acid metabolism region, as well as
neuraminidase (VC1784), which is a glycosylhydrolase known to release
sialic acid from sialoglycoconjugates to unmask GM1 gangliosides, the
receptor for cholera toxin. We examined the tRNA-serine locus among the
sequenced V. cholerae genomes and identified five variant VPI-2
regions, four of which retained the sialometabolism region. Three
variant VPI-2 regions contained a type three secretion system. By using
an inverse nested PCR approach, we found that the VPI-2 region can form
an extrachromosomal circular intermediate (CI) molecule after precise
excision from its tRNA-serine attachment site. We constructed a
knockout mutant of VC1758 (int) with V. cholerae strain N16961 and
found that no excision PCR product was produced, indicating that a
functional cognate, VPI-2 integrase, is required for excision. The
Vibrio seventh pandemic island-I (VSP-I) and VSP-II regions are present
in V. cholerae 01 El Tor and 0139 serogroup isolates. Novel regions are
present at the VSP-I insertion site in strain MZO-3 and at the VSP-II
insertion site in strain 623-39. VSP-II is a 27-kb region that
integrates at a tRNA-methionine locus, is flanked by direct repeats,
and encodes a P4-like integrase. We show that VSP-II can excise and
form a CI and that the cognate VSP-II integrase is required for
excision. Interestingly, VSP-I is not inserted at a tRNA locus and does
encode a XerDC-like recombinase, but similar to VPI-2 and VSP-II, VSP-I
does excise from the genome to form a CI. These results show that all
three pathogenicity islands can excise from the chromosome, which is
likely a first step in their horizontal transfer.

<>

<1>Murray, I.A., Clark, T.A., Morgan, R.D., Boitano, M., Anton, B.P., Luong, K., Fomenkov, A., Turner, S.W., Korlach, J., Roberts, R.J.
<2>The methylomes of six bacteria.
<3>Nucleic Acids Res.
<4>40
<5>11450-11462
<6>2012
<7>Six bacterial genomes, Geobacter metallireducens GS-15, Chromohalobacter salexigens, Vibrio
breoganii 1C-10, Bacillus cereus ATCC 10987, Campylobacter jejuni subsp. jejuni 81-176 and C.
jejuni NCTC 11168, all of which had previously been sequenced using other platforms were
re-sequenced using single-molecule, real-time (SMRT) sequencing specifically to analyze their
methylomes. In every case a number of new N6-methyladenine (m6A) and N4-methylcytosine (m4C)
methylation patterns were discovered and the DNA methyltransferases (MTases) responsible for
those methylation patterns were assigned. In 15 cases, it was possible to match MTase genes
with MTase recognition sequences without further sub-cloning. Two Type I restriction systems
required sub-cloning to differentiate their recognition sequences, while four MTase genes that
were not expressed in the native organism were sub-cloned to test for viability and
recognition sequences. Two of these proved active. No attempt was made to detect 5-
methylcytosine (m5C) recognition motifs from the SMRT sequencing data because this
modification produces weaker signals using current methods.  However, all predicted m6A and
m4C MTases were detected unambiguously. This study shows that the addition of SMRT sequencing
to traditional sequencing approaches gives a wealth of useful functional information about a
genome showing not only which MTase genes are active but also revealing their recognition
sequences.

<>

<1>Murray, I.A., Stickel, S.K., Roberts, R.J.
<2>Sequence-specific cleavage of RNA by Type II restriction enzymes.
<3>Nucleic Acids Res.
<4>38
<5>8257-8268
<6>2010
<7>The ability of 223 Type II restriction endonucleases to hydrolyze RNA-DNA heteroduplex
oligonucleotide substrates was assessed. Despite the
significant topological and sequence asymmetry introduced when one strand
of a DNA duplex is substituted by RNA we find that six restriction enzymes
(AvaII, AvrII, BanI, HaeIII, HinfI and TaqI), exclusively of the Type IIP
class that recognize palindromic or interrupted-palindromic DNA sequences,
catalyze robust and specific cleavage of both RNA and DNA strands of such
a substrate. Time-course analyses indicate that some endonucleases
hydrolyze phosphodiester bonds in both strands simultaneously whereas
others appear to catalyze sequential reactions in which either the DNA or
RNA product accumulates more rapidly. Such strand-specific variation in
cleavage susceptibility is both significant (up to orders of magnitude
difference) and somewhat sequence dependent, notably in relation to the
presence or absence of uracil residues in the RNA strand. Hybridization to
DNA oligonucleotides that contain endonuclease recognition sites can be
used to achieve targeted hydrolysis of extended RNA substrates produced by
in vitro transcription. The ability to 'restrict' an RNA-DNA hybrid,
albeit with a limited number of restriction endonucleases, provides a
method whereby individual RNA molecules can be targeted for site-specific
cleavage in vitro.

<>

<1>Murray, K., Hughes, S.G., Brown, J.S., Bruce, S.A.
<2>Isolation and characterization of two sequence-specific endonucleases from Anabaena variabilis.
<3>Biochem. J.
<4>159
<5>317-322
<6>1976
<7>Two endonucleases, AvaI and AvaII, were isolated from Anabaena variabilis on
the basis of their ability to make a limited number of breaks at specific
points in bacteriophage lambda DNA.  Neither enzyme has cofactor requirements
beyond Mg2+.  Endonuclease AvaI makes eight breaks in the phage lambda
chromosome at which the 5'-terminal sequence is pPy-C-G-N.  AvaII endonuclease
cuts phage lambda DNA more extensively, yielding fragments with the 5'-terminal
sequence G-T-C-N or G-A-C-N.  Neither enzyme generates cohesive ends.

<>

<1>Murray, K., Murray, N.E., Brammar, W.J.
<2>Restriction enzymes and the cloning of eukaryotic DNA.
<3>FEBS J.
<4>10
<5>193-207
<6>1975
<7>If bacteriophage lambda that has been grown on Escherichia coli strain C is transferred to E.
coli strain K, its efficiency of growth is impaired by several orders of magnitude, but those
phage that survive on strain K subsequently grow normally upon it.  If the surviving phage are
then transferred back to strain C they grow with normal efficiency, but if after this growth
on strain C they are again transferred to strain K, the reduced efficiency of growth is once
more observed.  This is an example of the general phenomenon of host-controlled restriction.
Experiments with 32P-labelled phage showed that the impairment of phage growth was correlated
with the ability of the restricting host strain to degrade the phage DNA to acid-soluble
products, a property that the original host strain lacked and which was attributed to a
host-specific endonuclease.  It is clearly necessary for the bacterial cell to protect its own
DNA from the action of this enzyme and this it does by methylation of certain bases (adenine
to 6-methylamino purine or, less commonly, cytosine to 5-methyl cytosine) as was demonstrated
by the analysis of DNA from phage f1 that had been propagated on restricting and
non-restricting strains of E. coli.  This process of host-controlled modification explains why
those phage that survive restriction subsequently grow normally upon the restricting strain:
their DNA is methylated by the host modification methylase and therefore survives attack by
the restriction endonuclease.

<>

<1>Murray, K., Old, R.W.
<2>The primary structure of DNA.
<3>Prog. Nucleic Acid Res. Mol. Biol.
<4>14
<5>117-185
<6>1974
<7>Brown and Todd, in 1952, summarized the evidence then available and proposed
the now generally accepted structure for deoxyribonucleic acid, in which the
deoxynucleoside units are linked by 3',5'-phosphodiester bonds.

<>

<1>Murray, N.E.
<2>Type I restriction systems: sophisticated molecular machines.
<3>Microbiol. Mol. Biol. Rev.
<4>64
<5>412-434
<6>2000
<7>Restriction enzymes are well known as reagents widely used by molecular biologists for genetic
manipulation and analysis, but these reagents represent only one class (type II) of a wider
range of enzymes that recognize specific nucleotide sequences in DNA molecules and detect the
provenance of the DNA on the basis of specific modifications to their target sequence. Type I
restriction and modification (R-M) systems are complex; a single multifunctional enzyme can
respond to the modification state of its target sequence with the alternative activities of
modification or restriction. In the absence of DNA modification, a type I R-M enzyme behaves
like a molecular motor, translocating vast stretches of DNA towards itself before eventually
breaking the DNA molecule. These sophisticated enzymes are the focus of this review, which
will emphasize those aspects that give insights into more general problems of molecular and
microbial biology. Current molecular experiments explore target recognition, intramolecular
communication, and enzyme activities, including DNA translocation. Type I R-M systems are
notable for their ability to evolve new specificities, even in laboratory cultures. This
observation raises the important question of how bacteria protect their chromosomes from
destruction by newly acquired restriction specifities. Recent experiments demonstrate
proteolytic mechanisms by which cells avoid DNA breakage by a type I R-M system whenever their
chromosomal DNA acquires unmodified target sequences. Finally, the review will reflect the
present impact of genomic sequences on a field that has previously derived information almost
exclusively from the analysis of bacteria commonly studied in the laboratory.

<>

<1>Murray, N.E.
<2>Immigration control of DNA in bacteria: self versus non-self.
<3>Microbiology
<4>148
<5>3-20
<6>2002
<7>Bacteria commonly endow their DNA with an identity mark.  When DNA is transferred from one
bacterium to another strain of the same species, DNA that lacks the identification mark of the
recipient strain is recognized as 'foreign' rather than 'self'.  Foreign DNA is commonly
degraded.  The first evidence for this discriminatory process was the demonstration of a
barrier, albeit incomplete, to the productive infection of Escherichia coli strain K-12 by
bacteriophage lambda previously propagated in either E. coli strain C or E. coli strain B.
Much later it was proven that the growth of phages in E. coli K-12 can be 'restricted' by an
endonuclease, a restriction enzyme (EcoKI), which attacks foreign DNA.  Occasionally phages
escape restriction and they, like the resident bacterial chromosome, acquire a protective
identification mark from a strain-specific modification enzyme that methylates defined bases
within a specific target sequence.  This sequence-specific modification identifies the
immediate provenance of bacterial, or phage, DNA.

<>

<1>Murray, N.E.
<2>The impact of phage lambda: from restriction to recombineering.
<3>Biochem. Soc. Trans.
<4>34
<5>203-207
<6>2006
<7>Experiments using phage lambda provided early insights into important molecular mechanisms,
including genetic recombination and the control of
gene expression. Before recombinant DNA technology, the use of lambda,
most particularly lambda transducing phages, illustrated the importance of
cloning bacterial genes, already providing some insight into how to use
cloned genes to advantage. Subsequently, lambda made significant
contributions to recombinant DNA technology, including the early
generation of genomic and cDNA libraries. More recently, lambda genes
associated with recombination have enabled techniques referred to as
'recombineering' to be developed. These techniques permit the refined
manipulation, including mutation, of foreign genes in Escherichia coli and
their subsequent return to the donor organism.

<>

<1>Murray, N.E., Barcus, V.A., Campbell, A.J.B., Dryden, D.T.F., Kelleher, J.E., Powell, L.M., Willcock, D., Sharp, P.M.
<2>Type I restriction enzymes of enteric bacteria.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>152
<6>1993
<7>The multifunctional type I restriction and modification (R/M) enzymes are encoded by three
genes (hsdR, hsdM and hsdS). These enzymes recognize asymmetric, bipartite target sequences
and their alternative responses of either restriction or modification are determined by the
methylation state of the target sequence. Three discrete families of type I R/M systems have
been described (A,B and C), and we now present evidence for a fourth. This new system and the
enzymes of families IA and IB are all encoded by allelic genes. Members of a family are highly
conserved, but differ in their specificity (A) subunits; each of two recognition domains of
the S subunit confers specificity for one of the two components of the target sequence.
Members of different families are dissimilar in all of their three subunits. Even when encoded
by allelic genes, amino acid identity may be only about 20%. The diversity of type I R/M
systems will be reviewed in relation to their natural distribution and their evolutionary
relationships. Horizontal transfer of hsd genes between species and frequency-dependent
selection for alternative specificities are invoked to explain the high levels of variability.
Comparative analyses of polypeptides identify conserved sequences within the M polypeptides
common to methyltransferases (mtases) in general and adenine mtases in particular, and within
the R polypeptides features in common with ATP-dependent helicases. We have made mutations in
some of these conserved regions and have isolated other mutants on the basis of a change in
response to the methylation state of the target sequence. The latter mutations mimic the
effect of a phage encoded polypeptide (Ral) that can enhance modification and ameliorate
restriction. The mtase component of EcoK, the type I R/M system found in E.coli K-12, has been
purified and characterized, particularly in terms of binding to target sequences and to the
cofactor S-adenosylmethionine. The properties of the wild-type mtase and some mutant
derivatives will be reported.

<>

<1>Murray, N.E., Batten, P.L., Murray, K.
<2>Restriction of bacteriophage lambda by Escherichia coli K.
<3>J. Mol. Biol.
<4>81
<5>395-407
<6>1973
<7>Derivatives of phage lambda, for which the numbers and positions of the
recognition sites for endonuclease R, EcoK are known, were used as substrates
for the Escherichia coli K restriction system in vivo and in vitro.  A single
unmodified recognition site was sufficient for a DNA molecule to be bound and
broken by the K restriction enzyme.  Although discrete fragments of DNA were
not produced, the breaks were made preferentially in the proximity of the
recognition site.  Breakage of a DNA molecule with only one recognition site
required a 10 to 40-fold higher concentration of restriction enzyme than
breakage of a DNA molecule with two or more recognition sites, but these
substrates were all equally effective in a binding assay for the enzyme.  The
polynucleotide kinase reaction provided no evidence for new 5'-terminal
sequences generated by restriction in vitro; the 5' termini were either
refractory to the polynucleotide kinase reaction or had no sequence
specificity.

<>

<1>Murray, N.E., Brammar, W.J.
<2>The trpE gene of Escherichia coli K contains a recognition sequence for the K-restriction system.
<3>J. Mol. Biol.
<4>77
<5>615-624
<6>1973
<7>A recognition sequence for the K-restriction system has been localized within
the trpE gene of Escherichia coli.  Mutations conferring resistance to
restriction do not always inactivate the E gene.  By selecting for phage
mutants which have simultaneously lost two restriction-recognition sites, one
in trpE and the other near gene N of phage lambda, we have isolated deletions
which localize a site of action of the N protein betwen cIII and N itself.

<>

<1>Murray, N.E., Daniel, A.S., Cowan, G.M., Sharp, P.M.
<2>Conservation of motifs within the unusually variable polypeptide sequences of type I restriction and modification enzymes.
<3>Mol. Microbiol.
<4>9
<5>133-143
<6>1993
<7>Type I restriction enzymes comprise three subunits encoded by genes designated hsdR, hsdM, and
hsdS; S confers sequence specificity. Three families of enzymes are known and within families,
but not between, hsdM and hsdR are conserved. Consequently, interfamily comparisons of M and R
sequences focus on regions of putative functional significance, , while both inter- and
intrafamily comparisons address the origin, nature and role of diversity of type I restriction
systems. We have determined the sequence of the hsdR gene for EcoAI, thus making available
sequences of all three hsd genes of one representative from each family. The predicted R
polypeptide sequences share conserved regions with one superfamily of putative helicases,
so-called DEAD box proteins; these conserved sequences may be associated with the
ATP-dependent translocation of DNA that precedes restriction. We also present hsdM and hsdR
sequences for EcoEI, a member of the same family as EcoAI. The sequences of the M and R genes
of EcoAI and EcoEI are at least as divergent as typical genes from Escherichia coli and
Salmonella, perhaps as the result of selection favouring diversity of restriction
specificities combined with lateral transfer among different species.

<>

<1>Murray, N.E., de Ritis, P.M., Foster, L.A.
<2>DNA targets for the Escherichia coli K restriction system analysed genetically in recombinants between phages Phi80 and lambda.
<3>Mol. Gen. Genet.
<4>120
<5>261-281
<6>1973
<7>Genetic analyses demonstrate the segregation of three targets for the K
restriction system in h80-lambda hybrid phages.  Mutations in each of these
three targets have been isolated and shown to confer resistance in cis but not
in trans.  Two of the three targets (sk-1 and sk-2) have been located on the
lambda genome:  sk-1 is right of gene R and sk-2 is between genes cIII and N.
The third target is in the phi80 genome right of, but close to, att.  Phage
lambda lacking both sk-1 and sk-2 retains at least 3 targets for the K
restriction system.

<>

<1>Murray, N.E., Gough, J.A., Suri, B., Bickle, T.A.
<2>Structural homologies among type I restriction-modification systems.
<3>EMBO J.
<4>1
<5>535-539
<6>1982
<7>Structural homologies among different restriction systems of Escherichia coli
and several Salmonella species have been investigated by immunological methods
using antibodies prepared against two subunits of the E. coli K12 restriction
enzyme, and by DNA hybridization experiments using different fragments of the
E. coli K12 hsd genes as probes.  The results with both techniques show a
strong homology between the E. coli K12 and B restriction-modification systems,
weaker but nevertheless marked homology between E. coli K12 and the Salmonella
systems SB, SP, and SQ and, surprisingly, no homology between the E. coli K12
and A systems.

<>

<1>Murray, N.E., Murray, K.
<2>Manipulation of restriction targets in phage lambda to form receptor chromosomes for DNA fragments.
<3>Nature
<4>251
<5>476-481
<6>1974
<7>Fragments of DNA from derivatives of phage lambda having either one or two
targets for R.EcoRI have been joined in new combinations to give biologically
active phage genomes.  These include two classes of deletion mutants; in one,
only the DNA between targets is deleted, but in the second the deletions are
more extensive.  Other fragments of DNA have been inserted into these phage
chromosomes.

<>

<1>Murthy, T., Rolfs, A., Hu, Y., Shi, Z., Raphael, J., Moreira, D., Kelley, F., McCarron, S., Jepson, D., Taycher, E., Zuo, D., Mohr, S.E., Fernandez, M., Brizuela, L., Labaer, J.
<2>A Full-Genomic Sequence-Verified Protein-Coding Gene Collection for Francisella tularensis.
<3>PLoS ONE
<4>2
<5>E577
<6>2007
<7>The rapid development of new technologies for the high throughput (HT)
study of proteins has increased the demand for comprehensive plasmid clone
resources that support protein expression. These clones must be
full-length, sequence-verified and in a flexible format. The generation of
these resources requires automated pipelines supported by software
management systems. Although the availability of clone resources is
growing, current collections are either not complete or not fully
sequence-verified. We report an automated pipeline, supported by several
software applications that enabled the construction of the first
comprehensive sequence-verified plasmid clone resource for more than 96%
of protein coding sequences of the genome of F. tularensis, a highly
virulent human pathogen and the causative agent of tularemia. This clone
resource was applied to a HT protein purification pipeline successfully
producing recombinant proteins for 72% of the genes. These methods and
resources represent significant technological steps towards exploiting the
genomic information of F. tularensis in discovery applications.

<>

<1>Murugan, N., Malathi, J., Umashankar, V., Madhavan, H.N.
<2>Comparative Genomic Analysis of Multidrug-Resistant Pseudomonas aeruginosa Clinical Isolates VRFPA06 and VRFPA08 with VRFPA07.
<3>Genome Announcements
<4>2
<5>e00140-14
<6>2014
<7>Pseudomonas aeruginosa isolates harboring acquired drug-resistant genes lead to increased
mortality. Here, we have sequenced and annotated the genomes of two
multidrug-resistant (MDR) P. aeruginosa isolates and a susceptible P. aeruginosa
clinical isolate evidencing divergent antibiotic susceptibilities. Genomic
analysis showed insight on the different genomic strategies adapted by P.
aeruginosa to combat antimicrobial effects.

<>

<1>Murugapiran, S.K. et al.
<2>Thermus oshimai JL-2 and T. thermophilus JL-18 genome analysis illuminates pathways for carbon, nitrogen, and sulfur cycling.
<3>Standards in Genomic Sciences
<4>7
<5>449-468
<6>2013
<7>The complete genomes of Thermus oshimai JL-2 and T. thermophilus JL-18 each consist of a
circular chromosome, 2.07 Mb and 1.9 Mb, respectively, and two
plasmids ranging from 0.27 Mb to 57.2 kb. Comparison of the T. thermophilus JL-18
chromosome with those from other strains of T. thermophilus revealed a high
degree of synteny, whereas the megaplasmids from the same strains were highly
plastic. The T. oshimai JL-2 chromosome and megaplasmids shared little or no
synteny with other sequenced Thermus strains. Phylogenomic analyses using a
concatenated set of conserved proteins confirmed the phylogenetic and taxonomic
assignments based on 16S rRNA phylogenetics. Both chromosomes encode a complete
glycolysis, tricarboxylic acid (TCA) cycle, and pentose phosphate pathway plus
glucosidases, glycosidases, proteases, and peptidases, highlighting highly
versatile heterotrophic capabilities. Megaplasmids of both strains contained a
gene cluster encoding enzymes predicted to catalyze the sequential reduction of
nitrate to nitrous oxide; however, the nitrous oxide reductase required for the
terminal step in denitrification was absent, consistent with their incomplete
denitrification phenotypes. A sox gene cluster was identified in both
chromosomes, suggesting a mode of chemolithotrophy. In addition, nrf and psr gene
clusters in T. oshmai JL-2 suggest respiratory nitrite ammonification and
polysulfide reduction as possible modes of anaerobic respiration.

<>

<1>Murugapiran, S.K. et al.
<2>Whole Genome Sequencing of Thermus oshimai JL-2 and Thermus thermophilus JL-18, Incomplete Denitrifiers from the United States Great Basin.
<3>Genome Announcements
<4>1
<5>e00106-12
<6>2013
<7>The strains Thermus oshimai JL-2 and Thermus thermophilus JL-18 each have a circular
chromosome, 2.07 Mb and 1.9 Mb in size, respectively, and each has two plasmids ranging from
0.27 Mb to 57.2 kb. The megaplasmid of each strain contains a gene cluster for the reduction
of nitrate to nitrous oxide, consistent with their incomplete denitrification phenotypes.

<>

<1>Muscarella, D.E., Ellison, E.L., Ruoff, B.M., Vogt, V.M.
<2>Characterization of I-Ppo, an intron-encoded endonuclease that mediates homing of a group I intron in the ribosomal DNA of Physarum polycephalum.
<3>Mol. Cell. Biol.
<4>10
<5>3386-3396
<6>1990
<7>A novel and only recently recognized class of enzymes is composed of the site-specific
endonucleases encoded by some group I introns. We have characterized several aspects of I-Ppo,
the endonuclease that mediates the mobility of intron 3 in the ribosomal DNA of Physarum
polycephalum. This intron is unique among mobile group I introns in that it is located in
nuclear DNA. We found that I-Ppo is encoded by an open reading frame in the 5' half of intron
3, upstream of the sequences required for self-splicing of group I introns. Either of two AUG
initiation codons could start this reading frame, one near the beginning of the intron and the
other in the upstream exon, leading to predicted polypeptides of 138 and 160 amino acid
residues. The longer polypeptide was the major form translated in vitro in a reticulocyte
extract. From nuclease assays of proteins synthesized in vitro with partially deleted DNAs, we
conclude that both polypeptides possess endonuclease activity. We also have expressed I-Ppo in
Escherichia coli, using a bacteriophage T7 RNA polymerase expression system. The longer
polypeptide also was the predominant form made in this system. It showed enzymatic activity in
bacteria in vivo, as demonstrated by the cleavage of a plasmid carrying the target site. Like
several other intron-encoded endonucleases, I-Ppo makes a four-base staggered cut in its
ribosomal DNA target sequence, very near the site where intron 3 becomes integrated in crosses
of intron 3-containing and intron 3-lacking Physarum strains.

<>

<1>Muscarella, D.E., Vogt, V.M.
<2>A mobile group I intron in the nuclear DNA of Physarum polycephalum.
<3>Cell
<4>56
<5>443-454
<6>1989
<7>We have shown that a strain-specific group I intron (intron 3) in the nuclear extrachromosomal
rDNA of Physarum polycephalum is a mobile element. Shortly after mating of amoebae from
intron-lacking and intron-containing strains, intron 3 transposes in a site-specific manner
into all available recipient molecules. The transposition appears to occur by gene conversion,
as evidenced by the co-conversion of adjacent sequences and by double strand breakage observed
in some of the recipient rDNA molecules. We infer that the double strand break is induced by
an endonuclease encoded by intron 3, since in vitro transcription and translation of the
cloned intron leads to the synthesis of an enzymatically active, site-specific nuclease. This
is the first demonstration of the transposition of a nuclear intron in an experimental
setting, and provides a rare example of a protein encoded by an RNA polymerase I transcript.

<>

<1>Mushtaq, R., Naeem, S., Sohail, A., Riazuddin, S.
<2>BseRI a novel restriction endonuclease from a Bacillus species which recognizes the sequence 5'...GAGGAG...3'.
<3>Nucleic Acids Res.
<4>21
<5>3585
<6>1993
<7>BseRI, a unique Type IIS restriction endonuclease, has been isolated from Bacillus species R
(CAMB 2669). BseRI recognizes a six base pair non-palindromic sequence 5'-GAGGAG-3' and
cleaves double stranded DNA ten nucleotides beyond this sequence on the top strand and eight
nucleotides beyond the sequence on the complementary strand to produce a two nucleotide 3'
extension.

<>

<1>Musich, P.R.
<2>Effects of cytosine methylation on the activity of restriction enzymes.
<3>J. Cell Biol.
<4>105
<5>155a
<6>1987
<7>Restriction cleavage of DNA is a major tool in the characterization eukaryotic
genes.  Cleavage by a restriction enzyme requires only that the DNA contain the
recognition site for the enzyme and that it not be modified so as to prohibit
the hydrolytic reaction.  In mammals, the DNA is modified in some of the
5'-CG-3' doublets by C5 methylation of cytosine (5mC).  The 5mC inhibits the
cleavage activity of some nucleases, but does not affect others.  However, the
effect of 5mC on most restriction enzyme activities has not been thoroughly
documented.  The studies presented address this need.  Single stranded pTZ19U
and pTZ19R plasmid DNAs were hybridized with the universal and reverse M13
sequencing primers, respectively.  These primed templates were extended by DNA
polI with dATP, dGTP, dTTP and either dCTP or d5mCTP, generating double strand
substrates in which one strand was completely methylated at all cytosine
positions.  These substrates were treated with 12 restriction enzymes, each
with a single cleavage site in the polyclonal region of these vectors.  AccI,
Asp718, SacI, SalI and SmaI were inactive on the hemimethylated substrates.
HindIII, KpnI, PstI and XbaI were active but exhibited modified specificities.
BamHI, EcoRI and HincII activitiies were not affected by 5mC.  These results
indicate that cytosine methylation can affect restriction cleavage either by
inhibition or by alteration of the selected cleavage site.  In addition, the
data confirm that isoschizomers are affected differently by methylation of the
same site.

<>

<1>Muyzer, G., Sorokin, D.Y., Mavromatis, K., Lapidus, A., Clum, A., Ivanova, N., Pati, A., d'Haeseleer, P., Woyke, T., Kyrpides, N.C.
<2>Complete genome sequence of 'Thioalkalivibrio sulfidophilus' HL-EbGr7.
<3>Standards in Genomic Sciences
<4>4
<5>23-35
<6>2011
<7>'Thioalkalivibrio sulfidophilus' HL-EbGr7 is an obligately chemolithoautotrophic,
haloalkaliphilic sulfur-oxidizing bacterium (SOB) belonging to the
Gammaproteobacteria. The strain was found to predominate a full-scale bioreactor,
removing sulfide from biogas. Here we report the complete genome sequence of
strain HL-EbGr7 and its annotation. The genome was sequenced within the Joint
Genome Institute Community Sequencing Program, because of its relevance to the
sustainable removal of sulfide from bio- and industrial waste gases.

<>

<1>Muyzer, G., Sorokin, D.Y., Mavromatis, K., Lapidus, A., Foster, B., Sun, H., Ivanova, N., Pati, A., D'haeseleer, P., Woyke, T., Kyripides, N.C.
<2>Complete genome sequence of Thioalkalivibrio sp. K90mix.
<3>Standards in Genomic Sciences
<4>5
<5>341-355
<6>2011
<7>Thioalkalivibrio sp. K90mix is an obligately chemolithoautotrophic, natronophilic
sulfur-oxidizing bacterium (SOxB) belonging to the family Ectothiorhodospiraceae within the
Gammaproteobacteria. The strain was isolated from a mixture of sediment samples obtained from
different soda lakes located in the Kulunda Steppe (Altai, Russia) based on its extreme
potassium carbonate tolerance as an enrichment method. Here we report the complete ge-nome
sequence of strain K90mix and its annotation. The genome was sequenced within the Joint Genome
Institute Community Sequencing Program, because of its relevance to the sus-tainable removal
of sulfide from wastewater and gas streams.

<>

<1>Myers, G.S. et al.
<2>Skewed genomic variability in strains of the toxigenic bacterial pathogen, Clostridium perfringens.
<3>Genome Res.
<4>16
<5>1031-1040
<6>2006
<7>Clostridium perfringens is a Gram-positive, anaerobic spore-forming bacterium commonly found
in soil, sediments, and the human
gastrointestinal tract. C. perfringens is responsible for a wide spectrum
of disease, including food poisoning, gas gangrene (clostridial
myonecrosis), enteritis necroticans, and non-foodborne gastrointestinal
infections. The complete genome sequences of Clostridium perfringens
strain ATCC 13124, a gas gangrene isolate and the species type strain, and
the enterotoxin-producing food poisoning strain SM101, were determined and
compared with the published C. perfringens strain 13 genome. Comparison of
the three genomes revealed considerable genomic diversity with >300 unique
"genomic islands" identified, with the majority of these islands unusually
clustered on one replichore. PCR-based analysis indicated that the large
genomic islands are widely variable across a large collection of C.
perfringens strains. These islands encode genes that correlate to
differences in virulence and phenotypic characteristics of these strains.
Significant differences between the strains include numerous novel mobile
elements and genes encoding metabolic capabilities, strain-specific
extracellular polysaccharide capsule, sporulation factors, toxins, and
other secreted enzymes, providing substantial insight into this medically
important bacterial pathogen.

<>

<1>Nadeev, A.N., Chernukhin, V.A., Sevastyanova, O.O., Tomilova, Y.E., Shinkarenko, N.M., Evdokimov, A.A., Degtyarev, S.K.
<2>BstKTI, a new dam-sensitive neoschizomer of restriction endonuclease MboI, which is able to cleave 5 hemimethylated substrate.
<3>Biotekhnologiya
<4>0
<5>5-10
<6>2006
<7>The strain Bacillus stearothermophilus KT, a producer of a new restriction endonuclease
BstKTI, which is the neoschizomer of restrictase MboI that recognizes the sequence
5'-GATC-3', has been found. The preparation of the enzyme was obtained, and its properties
were studied including substrate specificity and cleavage position in DNA. It was shown that
DNA hydrolysis occurred in position 5'-GAT^C-3'. Unlike the other isoschizomers of the
restriction endonuclease MboI, BstKTI is sensitive to dam-methylation, but it is able to
cleave the adenine-hemimethylated substrate. Moreover, BstKTI is capable of cleaving the
sequence 5'-GATC-3' that contains the modified cytosine in position C5.

<>

<1>Naderer, M., Brust, J.R., Knowle, D., Blumenthal, R.M.
<2>Mobility of a restriction-modification system revealed by its genetic context in three hosts.
<3>J. Bacteriol.
<4>184
<5>2411-2419
<6>2002
<7>The flow of genes among prokaryotes plays a fundamental role in shaping bacterial evolution,
and restriction-modification systems can modulate this flow. However, relatively little is
known about the distribution and movement of restriction-modification systems themselves. We
have isolated and characterized the genes for restriction-modification systems from two
species of Salmonella, S. enterica serovar Paratyphi A and S. enterica serovar Bareilly. Both
systems are closely related to the PvuII restriction-modification system and share its target
specificity. In the case of S. enterica serovar Paratyphi A, the restriction endonuclease is
inactive, apparently due to a mutation in the subunit interface region. Unlike the
chromosomally located Salmonella systems, the PvuII system is plasmid borne. We have completed
the sequence characterization of the PvuII plasmid pPvu1, originally from Proteus vulgaris,
making this the first completely sequenced plasmid from the genus Proteus. Despite the
pronounced similarity of the three restriction-modification systems, the flanking sequences in
Proteus and Salmonella are completely different. The SptAI and SbaI genes lie between an
equivalent pair of bacteriophage P4-related open reading frames, one of which is a putative
integrase gene, while the PvuII genes are adjacent to a mob operon and a XerCD recombination
(cer) site.

<>

<1>Nadig, S., Velusamy, N., Lalitha, P., Kar, S., Sharma, S., Arakere, G.
<2>Staphylococcus aureus eye infections in two Indian hospitals: emergence of ST772 as a major clone.
<3>Clinical Ophthalmology
<4>6
<5>165-173
<6>2012
<7>PURPOSE: The purpose of this study was to perform molecular characterization of Staphylococcus
aureus isolates causing a variety of eye infections from two major eye care hospitals in
India. METHODS: Twenty-four isolates from Aravind Eye Hospital, Madurai, India, and nine
isolates from LV Prasad Eye Institute, Bhubaneswar, India, representing severe to nonsevere
eye infections like microbial keratitis to lacrimal sac abscess, were characterized.
Staphylococcal cassette chromosome mec typing, multilocus sequence typing, accessory gene
regulator typing, staphylococcal protein A typing, and pulsed field gel electrophoresis were
used, along with determination of the presence of Panton-Valentine leucocidin toxin and
endotoxin gene cluster among each sequence type. RESULTS: The majority of eye infections, both
severe and nonsevere, were caused by sequence type (ST)772, positive for the Panton-Valentine
leucocidin gene, and carrying methicillin-resistant staphylococcal cassette chromosome mec
type V cassette (22/33, 67%). Some of the other sequence types that caused severe eye
infections were ST1 (9%), 5 (3%), 72 (6%), 88 (3%), 121 (3%), and 672 (3%).
This is the first report of the presence of ST1 and 88 in India. CONCLUSION:
Although the number of isolates included in this study was small, most of the eye infections
were caused by community-associated S. aureus where patients had no history of hospitalization
or treatment in the past year. In the case of six severe infections, patients were admitted
for surgeries and there is probability of hospital infection. In addition, only
methicillin-resistant S. aureus isolates carrying staphylococcal cassette chromosome mec type
V were detected. Epidemic methicillin-resistant Staphylococcus aureus 15 (ST22) is a major ST
found in health care as well as community settings in non-eye infections in India, but only
one methicillin-sensitive S. aureus isolate belonging to ST22 was detected.
Predominantly ST772, along with a few other STs, caused the 33 eye infections studied.

<>

<1>Nagai, Y., Nogami, S., Kumagai-Sano, F., Ohya, Y.
<2>Karyopherin-mediated nuclear import of the homing endonuclease VMA1-derived endonuclease is required for self-propagation of the  coding region.
<3>Mol. Cell. Biol.
<4>23
<5>1726-1736
<6>2003
<7>VMA1-derived endonuclease (VDE), a site-specific endonuclease in Saccharomyces cerevisiae,
enters the nucleus to generate a double-strand
break in the VDE-negative allelic locus, mediating the self-propagating
gene conversion called homing. Although VDE is excluded from the nucleus
in mitotic cells, it relocalizes at premeiosis, becoming localized in both
the nucleus and the cytoplasm in meiosis. The nuclear localization of VDE
is induced by inactivation of TOR kinases, which constitute central
regulators of cell differentiation in S. cerevisiae, and by nutrient
depletion. A functional genomic approach revealed that at least two
karyopherins, Srp1p and Kap142p, are required for the nuclear localization
pattern. Genetic and physical interactions between Srp1p and VDE imply
direct involvement of karyopherin-mediated nuclear transport in this
process. Inactivation of TOR signaling or acquisition of an extra nuclear
localization signal in the VDE coding region leads to artificial nuclear
localization of VDE and thereby induces homing even during mitosis. These
results serve as evidence that VDE utilizes the host systems of nutrient
signal transduction and nucleocytoplasmic transport to ensure the
propagation of its coding region.

<>

<1>Nagamalleswari, E., Vasu, K., Nagaraja, V.
<2>Ca(2+) binding to the ExDxD motif regulates the DNA cleavage specificity of a promiscuous endonuclease.
<3>Biochemistry
<4>51
<5>8939-8949
<6>2012
<7>Most of the restriction endonucleases (REases) are dependent on Mg(2+) for DNA cleavage, and
in general, Ca(2+) inhibits their activity. R.KpnI, an HNH active
site containing betabetaalpha-Me finger nuclease, is an exception. In presence of
Ca(2+), the enzyme exhibits high-fidelity DNA cleavage and complete suppression
of Mg(2+)-induced promiscuous activity. To elucidate the mechanism of unusual
Ca(2+)-mediated activity, we generated alanine variants in the putative Ca(2+)
binding motif, E(132)xD(134)xD(136), of the enzyme. Mutants showed decreased
levels of DNA cleavage in the presence of Ca(2+). We demonstrate that ExDxD
residues are involved in Ca(2+) coordination; however, the invariant His of the
catalytic HNH motif acts as a general base for nucleophile activation, and the
other two active site residues, D148 and Q175, also participate in
Ca(2+)-mediated cleavage. Insertion of a 10-amino acid linker to disrupt the
spatial organization of the ExDxD and HNH motifs impairs Ca(2+) binding and
affects DNA cleavage by the enzyme. Although ExDxD mutant enzymes retained
efficient cleavage at the canonical sites in the presence of Mg(2+), the
promiscuous activity was greatly reduced, indicating that the carboxyl residues
of the acidic triad play an important role in sequence recognition by the enzyme.
Thus, the distinct Ca(2+) binding motif that confers site specific cleavage upon
Ca(2+) binding is also critical for the promiscuous activity of the Mg(2+)-bound
enzyme, revealing its role in metal ion-mediated modulation of DNA cleavage.

<>

<1>Nagao, K., Suzuki, K., Hamada, S., Yahara, S., Yamamura, R., Uyeda, M.
<2>1513-DMIa and 1513-DMIb, DNA methyltransferase inhibitors produced by Streptomyces sp. strain no. 1513.
<3>J. Enzym. Inhib.
<4>13
<5>135-146
<6>1998
<7>Two new methyltransferase inhibitors were isolated from the culture filtrate of Streptomyces
sp. strain No. 1513 and named 1513-DMIa and 1513-DMIb.  1513-DMIa and 1513-DMIb were
distinguished in certain properties from DMI-1, DMI-2, DMI-3 and DMI-4 previously reported.
The molecular weight of 1513-DMIa and 1513-DMIb were estimated to be 576 and 8400 from the
results of FAB-MS and gel filtration, respectively.  The inhibitory activities of 1513-DMIa
and 1513-DMIb were shown to be pH- and temperature-dependent and both inhibited M.EcoRI in an
uncompetitive manner with respect to DNA or S-adenosylmethionine (SAM).

<>

<1>Nagao, K., Suzuki, K., Tokunaga, J., Miyazaki, H., Katayama, N., Mitsuyama, R., Uyeda, M.
<2>DMI-2 and DMI-3, DNA methyltransferase inhibitors produced by Streptomyces sp. strain no. 560.
<3>J. Enzym. Inhib.
<4>10
<5>115-124
<6>1996
<7>Streptomyces sp. strain no. 560 produces several types of DNA methyltransferase inhibitors in
the culture filtrate.  Two of them, DMI-2 and DMI-3, were distinguished from the previously
reported DMI-1 by their inhibitory spectrum and inhibition characteristics against DNA
methyltransferase.  The molecular weights of DMI-2 and DMI-3 were 854 and 435, respectively.
The structure of DMI-2 was determined to be
4R,6aR,10S,10aS-8-acetyl-6a.10a-d:hydroxy-2-methoxy-12-methyl-10-[4-[3-hydroxy-3,5-dimethy
tetrahydropyran-1-yloxy)-5-methylcyclohexan-1-yloxyl-1,4,6,7,9-pentaoxo-1,4,6,6a,7,8,9,10,10a
The chemical structure of DMI-2 was established as a tautomer of duiomycin which is an
antitumor antibiotic produced by Streptomyces sp. 1725.  DMI-2 and DMI-3 showed strong
inhibition against N6-methyladenine-DNA methyltransferase (M.EcoRI).  DMI-2 inhibited M.EcoRI
in a competitive manner with respect to plasmid pUC19 used as DNA substrate and in an
uncompetitive manner with respect to S-adenosylmethionine (SAM) used as methyl donor.  DMI-3
inhibited M.EcoRI in a competitive manner with respect to plasmid pUC19 and SAM.  The
inhibitory activities of both inhibitors depended upon the pH and temperature in the assay
media.

<>

<1>Nagaraja, V., Saravanan, M., Zhu, Z.
<2>KpnI restriction endonucleases with reduced star activity.
<3>US Patent Office
<4>US 20080227133
<5>
<6>2008
<7>Methods are provided for making restriction endonucleases with reduced star activity by one or
more targeted mutations to a catalytic site within the restriction endonuclease.  Examples of
modifications to restriction endonucleases with significant sequence identity with KpnI are
provided and reduced star activity demonstrated.

<>

<1>Nagaraja, V., Saravanan, M., Zhu, Z.
<2>Variant of KpnI restriction endonuclease.
<3>US Patent Office
<4>US 08119382
<5>
<6>2012
<7>Methods are provided for making restriction endonucleases with reduced star activity by one or
more targeted mutations to a catalytic site within the restriction endonuclease. Examples of
modifications to restriction endonucleases with significant sequence identity with KpnI are
provided and reduced star activity demonstrated.

<>

<1>Nagaraja, V., Shepherd, J.C.W., Bickle, T.A.
<2>A hybrid recognition sequence in a recombinant restriction enzyme and the evolution of DNA sequence specificity.
<3>Nature
<4>316
<5>371-372
<6>1985
<7>Early attempts to generate new restriction specificities by recombination
between allelic restriction-modification systems have been unsuccessful.
Bullas et al. succeeded in isolating a new specificity, SQ, in Salmonella that
they interpreted as being the result of a recombination event betwen the
parental strains, Salmonella typhimurium and S. potsdam, which encode the SB
and SP restriction systems, respectively.  This interpretation has recently
been confirmed by DNA heteroduplex studies with the SB, SP and SQ structural
genes.  We have determined the DNA sequences recognized by the SB and SP
enzymes and found that, like all type I restriction sequences, they are split
into two specific domains by a spacer of nonspecific sequence that, for both SB
and SP, is 6 base pairs (bp) long.  We have now determined the sequence
recognized by the recombinant SQ enzyme and find that it is a hybrid between
the SB and SP sequences, containing one specific domain from each parental
strain.  This result implies that each of the two specific domains is
recognized by a physically distinct part of the enzyme.

<>

<1>Nagaraja, V., Shepherd, J.C.W., Pripfl, T., Bickle, T.A.
<2>Two Type I restriction enzymes from Salmonella species.  Purification and DNA recognition sequences.
<3>J. Mol. Biol.
<4>182
<5>579-587
<6>1985
<7>We have purified the type I restriction enzymes SB and SP from Salmonella typhimurium and S.
potsdam, respectively, and determined the DNA sequences that they recognize.  These sequences
resemble those previously determined for the type I enzymes, EcoB, EcoK and EcoA, in that the
specific part of the sequence is divided into two domains by a spacer of non-specific sequence
that has a fixed length for each enzyme.  Two main differences from the previously determined
sequences are seen.  Both of the new sequences are degenerate and one of them, SB, has one
trinucleotide and one pentanucleotide-specific domain rather than the trinucleotide and
tetranucleotide domains seen for all of the other enzymes.  The only conserved features of the
recognition sequences are the adenosyl residues that are methylated in the modification
reaction.  For all of the enzymes these are situated ten or 11 base-pairs apart, one on each
strand of the DNA.  This suggests that the enzymes bind to DNA along one face of the double
helix making protein-DNA interaction in two successive major grooves with most of the
non-specific spacer sequence in the intervening minor groove.

<>

<1>Nagaraja, V., Stieger, M., Nager, C., Hadi, S.M., Bickle, T.A.
<2>The nucleotide sequence recognized by the Escherichia coli D type I restriction and modification enzyme.
<3>Nucleic Acids Res.
<4>13
<5>389-399
<6>1985
<7>A type I restriction endonuclease from a new isolate of Escherichia coli (E.
coli E166) has been purified and characterized.  The enzyme, EcoD, has a
recognition sequence similar in overall structure to the previously determined
type I enzyme sequences, an exception being that it is degenerate.  The
sequence is 5'-T-T-A-N-N-N-N-N-N-N-N-G-T-C-Y-3'
3'-A-A-T-N-N-N-N-N-N-N-N-C-A-G-R-5'where Y is a pyrimidine, R is a purine and N
can be any nucleotide.  The enzyme methylates adenosyl residues in both strands
of the DNA that are separated by ten base pairs, suggesting that the enzyme
interacts with DNA along one face of the helix making contacts in two
successive major grooves.

<>

<1>Nagaraja, V., Suri, B., Shepherd, J.C.W., Bickle, T.A.
<2>New variations of an old theme:  Type I restriction enzymes and their recognition sequences.
<3>Gene Manipulation and Expression, CroomHelm, Glass, R.E., Spizek, J., 
<4>0
<5>62-78
<6>1985
<7>None

<>

<1>Nagarajan, H., Butler, J.E., Klimes, A., Qiu, Y., Zengler, K., Ward, J., Young, N.D., Methe, B.A., Palsson, B.O., Lovley, D.R., Barrett, C.L.
<2>De Novo assembly of the complete genome of an enhanced electricity-producing variant of Geobacter sulfurreducens using only short reads.
<3>PLoS ONE
<4>5
<5>E10922
<6>2010
<7>State-of-the-art DNA sequencing technologies are transforming the life
sciences due to their ability to generate nucleotide sequence information
with a speed and quantity that is unapproachable with traditional Sanger
sequencing. Genome sequencing is a principal application of this
technology, where the ultimate goal is the full and complete sequence of
the organism of interest. Due to the nature of the raw data produced by
these technologies, a full genomic sequence attained without the aid of
Sanger sequencing has yet to be demonstrated.We have successfully
developed a four-phase strategy for using only next-generation sequencing
technologies (Illumina and 454) to assemble a complete microbial genome de
novo. We applied this approach to completely assemble the 3.7 Mb genome of
a rare Geobacter variant (KN400) that is capable of unprecedented current
production at an electrode. Two key components of our strategy enabled us
to achieve this result. First, we integrated the two data types early in
the process to maximally leverage their complementary characteristics. And
second, we used the output of different short read assembly programs in
such a way so as to leverage the complementary nature of their different
underlying algorithms or of their different implementations of the same
underlying algorithm.The significance of our result is that it
demonstrates a general approach for maximizing the efficiency and success
of genome assembly projects as new sequencing technologies and new
assembly algorithms are introduced. The general approach is a meta
strategy, wherein sequencing data are integrated as early as possible and
in particular ways and wherein multiple assembly algorithms are
judiciously applied such that the deficiencies in one are complemented by
another.

<>

<1>Nagashima, S., Kamimura, A., Shimizu, T., Nakamura-Isaki, S., Aono, E., Sakamoto, K., Ichikawa, N., Nakazawa, H., Sekine, M., Yamazaki, S., Fujita, N., Shimada, K., Hanada, S., Nagashima, K.V.
<2>Complete Genome Sequence of Phototrophic Betaproteobacterium Rubrivivax gelatinosus IL144.
<3>J. Bacteriol.
<4>194
<5>3541-3542
<6>2012
<7>Rubrivivax gelatinosus is a facultative photoheterotrophic betaproteobacterium living in
freshwater ponds, sewage ditches, activated sludge, and food processing
wastewater. There have not been many studies on photosynthetic
betaproteobacteria. Here we announce the complete genome sequence of the
best-studied phototrophic betaproteobacterium, R. gelatinosus IL-144 (NBRC
100245).

<>

<1>Nagata, Y., Ohtsubo, Y., Endo, R., Ichikawa, N., Ankai, A., Oguchi, A., Fukui, S., Fujita, N., Tsuda, M.
<2>Complete Genome Sequence of the Representative {gamma}-Hexachlorocyclohexane-Degrading Bacterium Sphingobium japonicum  UT26.
<3>J. Bacteriol.
<4>192
<5>5852-5853
<6>2010
<7>Sphingobium japonicum strain UT26 utilizes gamma-hexachlorocyclohexane (gamma-HCH), a man-made
chlorinated pesticide that causes serious
environmental problems due to its toxicity and long persistence, as a sole
source of carbon and energy. Here, we report the complete genome sequence
of UT26, which consists of two chromosomes and three plasmids. The 15 lin
genes involved in gamma-HCH degradation are dispersed on the two
chromosomes and one of the three plasmids.

<>

<1>Nagornykh, M., Zakharova, M., Protsenko, A., Bogdanova, E., Solonin, A.S., Severinov, K.
<2>Regulation of gene expression in restriction-modification system Eco29kI.
<3>Nucleic Acids Res.
<4>39
<5>4653-4663
<6>2011
<7>The Eco29kI restriction-modification (R-M) system consists of two partially overlapping genes,
eco29kIR, encoding a restriction endonuclease
and eco29kIM, encoding methyltransferase. The two genes are thought to
form an operon with the eco29kIR gene preceding the eco29kIM gene. Such an
organization is expected to complicate establishment of plasmids
containing this R-M system in naive hosts, since common logic dictates
that methyltransferase should be synthesized first to protect the DNA from
cleavage by the endonuclease. Here, we characterize the Eco29kI gene
transcription. We show that a separate promoter located within the
eco29kIR gene is sufficient to synthesize enough methyltransferase to
completely modify host DNA. We further show that transcription from two
intragenic antisense promoters strongly decreases the levels of eco29kIR
gene transcripts. The antisense transcripts act by preventing translation
initiation from the bicistronic eco29kIR-eco29kIM mRNA and causing its
degradation. Both eco29kIM and antisense promoters are necessary for
Eco29kI genes establishment and/or stable maintenance, indicating that
they jointly contribute to coordinated expression of Eco29kI genes.

<>

<1>Nagornykh, M.O., Bogdanova, E.S., Protsenko, A.S., Solonin, A.S., Zakharova, M.V., Severinov, K.V.
<2>Regulation of gene expression in a type II restriction-modification system.
<3>Genetika
<4>44
<5>606-615
<6>2008
<7>Type II restriction-modification systems are comprised of a restriction endonuclease and
methyltransferase. The enzymes are coded by individual genes and recognize the same DNA
sequence. Endonuclease makes a double-stranded break in the recognition site, and
methyltransferase covalently modifies DNA bases within the recognition site, thereby
preventing cleavage by the endonuclease. The concerted action of these enzymes plays the role
of a primitive immune system and protects the bacterial host cell from invasion by foreign
(for example, viral) DNA. However, uncontrolled expression of restriction-modification system
genes can result in the death of a bacterial host cell because of endonuclease cleavage of the
host DNA. In the present review, data on the regulation of expression of the type II
restriction-modification enzymes genes are discussed.

<>

<1>Nahon, E., Raveh, D.
<2>Targeting a truncated Ho-endonuclease of yeast to novel DNA sites with foreign zinc fingers.
<3>Nucleic Acids Res.
<4>26
<5>1233-1239
<6>1998
<7>Ho-endonuclease of the yeast, Saccharomyces cerevisiae, initiates a mating type switch by
making a site-specific double strand break in the mating type gene, MAT.  Ho is a dodecamer
endonuclease and shares six of the seven intein motifs with PI-SceI endonuclease, an intein
encoded by the VMAI gene.  We show that a 113 residue truncated Ho-endonuclease starting at
intein motif C initiates a mating type switch in yeast.  Ho is the only dodecamer endonuclease
with zinc finers.  To see whether they have a role in determining site specificity we
exchanged them for zinc fingers of the yeast transcription factor, Swi5.  A chimeric
endonuclease comprising the dodecamer motifs of Ho (C-E) and the zinc finger domain of Swi5
cleaves a Swi5 substrate plasmid in vivo.  A similar chimera with the zinc fingers of Sp1
cleaves a GC box rich substrate plasmid.  These experiments delineate a catalytic fragment of
Ho-endonuclease that can be fused to various DNA binding moieties in the design of chimeric
endonucleases with new site specificities.

<>

<1>Nair, D., Memmi, G., Hernandez, D., Bard, J., Beaume, M., Gill, S., Francois, P., Cheung, A.L.
<2>Whole genome sequencing of Staphylococcus aureus strain RN4220, a key laboratory strain used in virulence research, identifies mutations that  affect not only virulence factors but also the fitness of the strain.
<3>J. Bacteriol.
<4>193
<5>2332-2335
<6>2011
<7>S. aureus RN4220, a cloning intermediate, is sometimes used in virulence, resistance and
metabolic studies. Using whole genome sequencing, we showed that RN4220 differs from NCTC8325
and contains a number of genetic polymorphisms that affect both virulence and general fitness,
thus implying caution in using this strain for these studies.

<>

<1>Naito, M., Hirakawa, H., Yamashita, A., Ohara, N., Shoji, M., Yukitake, H., Nakayama, K., Toh, H., Yoshimura, F., Kuhara, S., Hattori, M., Hayashi, T., Nakayama, K.
<2>Genome sequence determination of Porphyromonas gingivalis strain ATCC 33277 and genomic comparison with strain W83 revealed extensive genome rearrangements in P. gingivalis.
<3>DNA Res.
<4>15
<5>215-225
<6>2008
<7>The gram-negative anaerobic bacterium Porphyromonas gingivalis is a major causative agent of
chronic periodontitis. Porphyromonas gingivalis strains
have been classified into virulent and less-virulent strains by mouse
subcutaneous soft tissue abscess model analysis. Here, we present the
whole genome sequence of P. gingivalis ATCC 33277, which is classified as
a less-virulent strain. We identified 2090 protein-coding sequences
(CDSs), 4 RNA operons, and 53 tRNA genes in the ATCC 33277 genome. By
genomic comparison with the virulent strain W83, we identified 461 ATCC
33277-specific and 415 W83-specific CDSs. Extensive genomic rearrangements
were observed between the two strains: 175 regions in which genomic
rearrangements have occurred were identified. Thirty-five of those genomic
rearrangements were inversion or translocation and 140 were simple
insertion, deletion, or replacement. Both strains contained large numbers
of mobile elements, such as insertion sequences, miniature inverted-repeat
transposable elements (MITEs), and conjugative transposons, which are
frequently associated with genomic rearrangements. These findings indicate
that the mobile genetic elements have been deeply involved in the
extensive genome rearrangement of P. gingivalis and the occurrence of many
of the strain-specific CDSs. We also describe here a very unique feature
of MITE400, which we renamed MITEPgRS (MITE of P. gingivalis with
Repeating Sequences).

<>

<1>Naito, T., Kobayashi, I.
<2>Cell death programmed by selfish genes -- or, why are there restriction enzymes?
<3>Jikken Igaku
<4>13
<5>1444-1447
<6>1995
<7>
<>

<1>Naito, T., Kusano, K., Kobayashi, I.
<2>Selfish behavior of restriction-modification systems.
<3>Science
<4>267
<5>897-899
<6>1995
<7>Plasmids carrying gene pairs encoding type II DNA restriction endonucleases and their cognate
modification enzymes were shown to have increased stability in Escherichia coli. The
descendants of cells that had lost these genes appeared unable to modify a sufficient number
of recognition sites in their chromosomes to protect them from lethal attack by the remaining
restriction enzyme molecules. The capacity of these genes to act as a selfish symbiont is
likely to have contributed to the evolution of restriction-modification gene pairs.

<>

<1>Naito, Y., Naito, T., Kobayashi, I.
<2>Selfish restriction modification genes: Resistance of a resident R/M plasmid to displacement by an incompatible plasmid mediated by host killing.
<3>Biol. Chem.
<4>379
<5>429-436
<6>1998
<7>Previous work from this laboratory demonstrated that plasmids carrying a type II
restriction-modification gene complex are not easily lost from their bacterial host because
plasmid-free segregant cells are killed through chromosome cleavage.  Here, we have followed
the course of events that takes place when an Escherichia coli recBC sbcA strain carrying a
plasmid coding for the PaeR7I restriction-modification gene complex is transformed by a
plasmid with an identical origin of replication.  The number of transformants that appeared
was far fewer than with the restriction-minus (r-) control.  Most of the transformants were
very small.  After prolonged incubation, the number and the size of the colonies increased,
but this increase never attained the level of the r- control.  Most of the transformed
colonies retained the drug-resistance of the resident, r+m+ plasmid.  These results indicate
that post-segregational host killing occurs when a plasmid bearing an R/M gene complex is
displaced by an incompatible plasmid.  Such cell killing eliminates the competitor plasmid
along with the host and, thus, would allow persistence of the R/M plasmid in the neighboring,
clonal host cells in nature.  This phenomenon is reminiscent of mammalian apoptosis and other
forms of altruistic cell death strategy against infection.  This type of resistance to
displacement was also studied in a wild type Escherichia coli strain that was normal for
homologous recombination.  A number of differences between the recBC sbcA strain and the rec+
strain were observed and these will be discussed.

<>

<1>Naka, H., Dias, G.M., Thompson, C.C., Dubay, C., Thompson, F.L., Crossa, J.H.
<2>Complete genome sequence of the marine fish pathogen Vibrio anguillarum harboring the pJM1 virulence plasmid and genomic comparison with other virulent strans of V. anguillarum and V. ordalii.
<3>Infect. Immun.
<4>79
<5>2889-2900
<6>2011
<7>We dissected the complete genome sequence of the O1 serotype strain Vibrio anguillarum
775(pJM1) and determined the draft genomic sequences of plasmidless strains of serotype O1
(strain 96F) and O2 (strain
RV22) and V. ordalii. All strains harbor two chromosomes, but 775 also harbors the virulence
plasmid pJM1, which carries the anguibactin-producing and cognate transport genes, one of the
main virulence factors of V.
anguillarum. Genomic analysis identified eight genomic islands in chromosome 1 of V.
anguillarum 775(pJM1) and two in chromosome 2. Some of them carried potential virulence genes
for the biosynthesis of O antigens, hemolysins, and exonucleases as well as others for sugar
transport and metabolism. The majority of genes for essential cell functions and pathogenicity
are located on chromosome 1. In contrast, chromosome 2 contains a larger fraction (59%) of
hypothetical genes than does chromosome 1 (42%). Chromosome 2 also harbors a superintegron, as
well as host "addiction" genes that are typically found on plasmids. Unique distinctive
properties include homologues of type III secretion system genes in 96F, homologues of V.
cholerae zot and ace toxin genes in RV22, and the biofilm formation syp genes in V. ordalii.
Mobile genetic elements, some of them possibly originated in the pJM1 plasmid, were very
abundant in 775, resulting in the silencing of specific genes, with only few insertions in the
96F and RV22 chromosomes.

<>

<1>Nakagawa, K.-I., Hashikawa, J.-I., Makino, O., Ando, T., Shibata, T.
<2>Subunit structure of a yeast site-specific endodeoxyribonuclease, endoSceI.  A study using monoclonal antibodies.
<3>Eur. J. Biochem.
<4>171
<5>23
<6>1988
<7>EndoSceI is a eucaryotic site-specific endoDNase of 120 kDa that causes
double-stranded scission at well-defined sites, but is distinguishable from
procaryotic restriction endonucleases by its mode of sequence recognition and
lack of related specific DNA modification.  In purified preparations of
endoSceI, only two polypeptide species of 75 kDa (75-kDa peptide) and 50 kDa
(50-kDa peptide) are detected in apparently equal amounts.  We prepared mouse
monoclonal IgGs that bound specifically to the 75-kDa peptide (but not the
50-kDa peptide) without inhibiting the endoSceI activity.  Immunoprecipitation
experiments with these IgGs revealed that the 75-kDa peptide and the 50-kDa
peptide are physically associated with each other and with the endonucleolytic
activity.  Full endoSceI activity was recovered by mixing the purified 75-kDa
peptide and the partially purified 50-kDa peptide, each of which exhibited
little or no endonuclease activity alone.  These observations indicate that
endoSceI consists of two non-identical subunits of 75 kDa and 50 kDa, and that
both subunits are required for full enzyme activity.

<>

<1>Nakagawa, K.-I., Morishima, N., Shibata, T.
<2>A maturase-like subunit of the sequence-specific endonuclease Endo.SceI from yeast mitochondria.
<3>J. Biol. Chem.
<4>266
<5>1977-1984
<6>1991
<7>Some yeast strains possess a sequence-specific endonuclease, Endo.SceI, which is a
heterodimeric enzyme localized in mitochondria.  The larger subunit (75 kDa) of Endo.SceI,
encoded by a nuclear gene (ENS1), is transported from the cytosol into the mitochondria.  In
this study, we determined the partial amino acid sequence of the smaller subunit (50 kDa) of
Endo.SceI.  The determined sequence matched well the partial sequence deduced from a
mitochondrial open reading frame (RF3).  The RF3 locus is known to exhibit polymorphism since
this reading frame in some yeast strains is supposed to encode a maturase-like protein,
whereas in other strains, the frame is interrupted by GC clusters, which thus break the frame.
Southern blot analysis of various yeast strains showed that the continuity of RF3 is
correlated with the presence of Endo.SceI activity.  These data indicate that the continuous
RF3 sequence is a functional gene (ENS2) coding for the smaller subunit of Endo.SceI.  The
results of cytoduction, by which the continuous RF3 sequence was transferred into a yeast
strain lacking mitochondrial DNA, confirmed this conclusion.  This study suggests the
involvement of Endo.SceI in genetic recombination of mitochondrial DNA.

<>

<1>Nakagawa, K.-I., Morishima, N., Shibata, T.
<2>An endonuclease with multiple cutting sites, Endo.SceI, initiates genetic recombination at its cutting site in yeast mitochondria.
<3>EMBO J.
<4>11
<5>2707-2715
<6>1992
<7>Endo.SceI is a mitochondrial sequence-specific endonuclease which has multiple cutting sites.
In order to examine the possible role of Endo.SceI in homologus recombination, we analyzed the
mode of recombination upon mating using antibiotic resistance markers on the mitochondrial
genome. The segregation of a marker located very close to one of the Endo.SceI cutting sites
showed a disparity (polarized segregation, i.e. gene conversion). This gene conversion
depended on the presence of the functional Endo.SceI gene. In vivo cutting of mitochondrial
DNA upon mating was detected at the cutting site in the antibiotic marker region, which also
depended on the Endo.SceI activity. These results suggest that mitochrondrial recombination is
induced by cleavage of mitochondrial DNA by this sequence-specific endonuclease. This is the
first demonstration that a sequence-specific endonuclease with multiple cutting sites induces
genetic recombination.

<>

<1>Nakagawa, S., Mizoguchi, H., Ando, S., Hayashi, M., Ochiai, K., Yokoi, H., Tateishi, N., Senoh, A., Ikeda, M., Ozaki, A.
<2>Novel polynucleotides.
<3>European Patent Office
<4>EP 2107128 A
<5>
<6>2009
<7>Novel polynucleotides derived from microorganisms belonging to coryneform bacteria and
fragments thereof, polypeptides encoded by the polynucleotides and fragments thereof,
polynucleotide arrays comprising the polynucleotides and fragments thereof, recording media in
which the nucleotide sequences of the polynucleotide and fragments thereof have been recorded
which are readable in a computer, and use of them.

<>

<1>Nakagawa, S., Mizoguchi, H., Ando, S., Hayashi, M., Ochiai, K., Yokoi, H., Tateishi, N., Senoh, A., Ikeda, M., Ozaki, A.
<2>Novel polynucleotides.
<3>Korean Patent Office
<4>KR 1020000077439 A
<5>
<6>2000
<7>Provided are novel polynucleotides derived from microorganisms belonging to Coryneform
bacteria and fragments thereof, polypeptides encoded by the polynucleotides and fragments
thereof, polynucleotide arrays comprising the polynucleotides and fragments thereof, recording
media in which the nucleotide sequences of the polynucleotide and fragments thereof have been
recorded and are readable in a computer, and their use.  The method for at least one of the
following: (A) identifying a mutation point of a gene derived from a mutant of a Coryneform
bacterium; (B) measuring an expression amount of a gene derived from a Coryneform, bacterium;
(C) analyzing an expression profile of a gene derived from a Coryneform bacterium; (D)
analyzing expression patterns of genes derived from a Coryneform bacterium; or (E) identifying
a gene homoologous to a gene derived from a Coryneform bacterium comprising the following
steps: (a) producing a polynucleotide array by adhering to a solid support of at least two
polynucleotides selected from the group consisting of (i) polynucleotides comprising the
nucleotide sequence represented by any one of SEQ ID NOS: 1 to 3501, (ii) polynucleotides
which hybridize with the (i) polynucleotides under stringent conditions, and (iii)
polynucleotides comprising a sequence of 10 to 200 continuous bases of the (i) or (ii)
polynucleotides; (b) incubating the polynucleotide array with at least one of a labeled
polynucleotide derived from a Coryneform bacterium, a labeled polynucleotide derived from a
mutant of the Coryneform bacterium or a labeled polynucleotide to be examined under
hybridization conditions; (c) detecting any hybridization; and (d) analyzing the result of the
hybridization.

<>

<1>Nakagawa, S., Mizoguchi, H., Ando, S., Hayashi, M., Ochiai, K., Yokoi, H., Tateishi, N., Senoh, A., Ikeda, M., Ozaki, A.
<2>Mutant of homoserine dehydrogenase from Corynebacterium and DNA encoding thereof.
<3>US Patent Office
<4>US 7332310 A
<5>
<6>2008
<7>Novel polynucleotides derived from microorganisms belonging to coryneform bacteria and
fragments thereof, polypeptides encoded by the polynucleotides and fragments thereof,
polynucleotide arrays comprising the polynucleotides and fragments thereof, recording media in
which the nucleotide sequences of the polynucleotide and fragments thereof have been recorded
which are readable in a computer, and use of them.

<>

<1>Nakagawa, S., Mizoguchi, H., Ando, S., Hayashi, M., Ochiai, K., Yokoi, H., Tateishi, N., Senoo, A., Ikeda, M., Ozaki, A.
<2>Novel polynucleotide.
<3>Japanese Patent Office
<4>JP 2002191370 A
<5>
<6>2002
<7>
<>

<1>Nakagawa, S., Shimamura, S., Takaki, Y., Suzuki, Y., Murakami, S., Watanabe, T., Fujiyoshi, S., Mino, S., Sawabe, T., Maeda, T., Makita, H., Nemoto, S., Nishimura, S., Watanabe, H., Watsuji, T., Takai, K.
<2>Allying with armored snails: the complete genome of gammaproteobacterial endosymbiont.
<3>ISME J.
<4>8
<5>40-51
<6>2014
<7>Deep-sea vents harbor dense populations of various animals that have their specific symbiotic
bacteria. Scaly-foot gastropods, which are snails with mineralized scales covering the sides
of its foot, have a gammaproteobacterial endosymbiont in their enlarged esophageal glands and
diverse epibionts on the surface of their scales.  In this study, we report the complete
genome sequencing of gammaproteobacterial endosymbiont.  The endosymbiont genome displays
features consistent with ongoing genome reduction such as large proportions of pseudogenes and
insertion elements.  The genome encodes functions commonly found in deep-sea vent
chemoautotrophs such as sulfur oxidation and carbon fixation.  Stable carbon isotope
(13C)-labeling experiments confirmed the endosymbiont chemoautotrophy.  The genome also
includes an intact hydrogenase gene cluster that potentially has been horizontally transferred
from phylogenetically distant bacteria.  Notable findings include the presence and
transcription of genes for flagellar assembly, through which proteins are potentially exported
from bacterium to the host.  Symbionts of snail individuals exhibited extreme genetic
homogeneity, showing only two synonymous changes in 19 different genes (13810 positions in
total) determined for 32 individual gastropods collected from a single colony at one time.
The extremely low genetic individuality in endosymbionts probably reflects that the stringent
symbiont selection by host prevents the random genetic drift in the small population of
horizontally transmitted symbiont.  This study is the first complete genome analysis of
gastropod endosymbiont and offers an opportunity to study genome evolution in a recently
evolved endosymbiont.

<>

<1>Nakagawa, S., Takaki, Y., Shimamura, S., Reysenbach, A.L., Takai, K., Horikoshi, K.
<2>Deep-sea vent epsilon-proteobacterial genomes provide insights into emergence of pathogens.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>12146-12150
<6>2007
<7>Deep-sea vents are the light-independent, highly productive ecosystems driven primarily by
chemolithoautotrophic microorganisms, in particular by
epsilon-Proteobacteria phylogenetically related to important pathogens. We
analyzed genomes of two deep-sea vent epsilon-Proteobacteria strains,
Sulfurovum sp. NBC37-1 and Nitratiruptor sp. SB155-2, which provide
insights not only into their unusual niche on the seafloor, but also into
the origins of virulence in their pathogenic relatives, Helicobacter and
Campylobacter species. The deep-sea vent epsilon-proteobacterial genomes
encode for multiple systems for respiration, sensing and responding to
environment, and detoxifying heavy metals, reflecting their adaptation to
the deep-sea vent environment. Although they are nonpathogenic, both
deep-sea vent epsilon-Proteobacteria share many virulence genes with
pathogenic epsilon-Proteobacteria, including genes for virulence factor
MviN, hemolysin, invasion antigen CiaB, and the N-linked glycosylation
gene cluster. In addition, some virulence determinants (such as the
H(2)-uptake hydrogenase) and genomic plasticity of the pathogenic
descendants appear to have roots in deep-sea vent epsilon-Proteobacteria.
These provide ecological advantages for hydrothermal vent
epsilon-Proteobacteria who thrive in their deep-sea habitat and are
essential for both the efficient colonization and persistent infections of
their pathogenic relatives. Our comparative genomic analysis suggests that
there are previously unrecognized evolutionary links between important
human/animal pathogens and their nonpathogenic, symbiotic,
chemolithoautotrophic deep-sea relatives.

<>

<1>Nakagawahara, K., Mori, M., Mioka, C.
<2>Types and uses of restriction enzymes.
<3>Rinsho Kensa
<4>40
<5>826-835
<6>1996
<7>A review (in japanese).

<>

<1>Nakahigashi, K., Kubo, N., Narita, S., Shimaoka, T., Goto, S., Oshima, T., Mori, H., Maeda, M., Wada, C., Inokuchi, H.
<2>HemK, a class of protein methyl transferase with similarity to DNA methyl transferases, methylates polypeptide chain release factors, and hemK knockout induces defects in translational termination.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>1473-1478
<6>2002
<7>HemK, a universally conserved protein of unknown function, has high amino acid similarity with
DNA-(adenine-N6) methyl transferases (MTases). A certain mutation in hemK gene rescues the
photosensitive phenotype of a ferrochelatase-deficient (hemH) mutant in Escherichia coli. A
hemK knockout strain of E. coli not only suffered severe growth defects, but also showed a
global shift in gene expression to anaerobic respiration, as determined by microarray
analysis, and this shift may lead to the abrogation of photosensitivity by reducing the
oxidative stress. Suppressor mutations that abrogated the growth defects of the hemK knockout
strain were isolated and shown to be caused by a threonine to alanine change at codon 246 of
polypeptide chain release factor (RF) 2, indicating that hemK plays a role in translational
termination. Consistent with such a role, the hemK knockout strain showed an enhanced rate of
read-through of nonsense codons and induction of transfer-mRNA-mediated tagging of proteins
within the cell. By analysis of the methylation of RF1 and RF2 in vivo and in vitro, we showed
that HemK methylates RF1 and RF2 in vitro within the tryptic fragment containing the conserved
GGQ motif, and that hemK is required for the methylation within the same fragment of, at
least, RF1 in vivo. This is an example of a protein MTase containing the DNA MTase motif and
also a protein-(glutamine-N5) MTase.

<>

<1>Nakajima, T., Matsubara, K., Ueno, H., Kagawa, S., Moore, J.E., Millar, B.C., Matsuda, M.
<2>Molecular identification and characterization of type III restriction-modification (R-M) gene cluster in Campylobacter lari.
<3>Ann. Microbiol. (Paris)
<4>63
<5>1629-1637
<6>2013
<7>Although the human clinical strain of Campylobacter lari (RM2100) has been shown not to carry
any type III restriction-modification (R-M) systems, an R-M genes cluster was identified
downstream of the full-length cytolethal distending toxin gene operon in the urease-positive
thermophilic Campylobacter (UPTC) CF89-12 strain. Two possible open reading frames (ORFs) for
restriction endonuclease and methyltransferase were predicted to encode peptides of 947 and
613 amino acid residues with calculated mo ecular weights of 111 and 70.8 kDa, respectively.
Two putative promoters consisting of the consensus sequences and two probable ribosome binding
sites for the two ORFs were also identified. Reverse transcription PCR identified
co-transcription of the R-M genes in the cells. The existence of an S-adenosyl
methionine-binding motif in the N-terminal conserved region of the possible ORF for the M
gene, and seven conserved helicase motifs in the R gene were also identified. PCR and Southern
blot hybridization a nalyses for type III R-M enzyme genes with some of the C. lari isolates
including UPTC gave positive signals. UPTC isolates were shown to carry type III R-M enzyme
genes, with a relatively high frequency.

<>

<1>Nakamaye, K.L., Eckstein, F.
<2>Inhibition of restriction endonuclease NciI cleavage by phosphorothioate groups and its application to oligonucleotide-directed mutagenesis.
<3>Nucleic Acids Res.
<4>14
<5>9679-9698
<6>1986
<7>M13 RF IV DNA where phosphorothioate groups are incorporated at restriction
endonuclease NciI recognition sites in the (-)strand is efficiently nicked by
the action of this enzyme.  Incubation of such nicked DNA with exonuclease III
produces gapped DNA.  The gap can be filled by reaction with deoxynucleoside
triphosphates and DNA polymerase I.  When this sequence of reactions is
performed with DNA containing a mismatch oligonucleotide primer in the
(-)-strand mutational frequencies of 70 - 90% can be obtained upon
transformation.  The general nature of this methodology has been further shown
to be applicable to other restriction enzymes such as HindII, PstI and FspI.
The mutational frequency obtained using these enzymes is between 40 - 80%
mainly because of less efficient nicking and gapping.  Studies on inhibition of
NciI cleavage show that in addition to a phosphorothioate group at the position
of cleavage an additional group in the 5'-neighbouring position is necessary
for complete inhibition.

<>

<1>Nakamura, S., Ikehata, H., Ono, T.
<2>Characteristics of mutations generated through digestion with restriction enzyme and ligation in plasmid DNA.
<3>Environ. Mol. Mutagen.
<4>38
<5>46-54
<6>2001
<7>Recently, the use of restriction enzymes has been extended to studies in which rare events
such as mutation and mistakes in DNA repair are
examined. In these studies, the specificity of restriction enzymes
becomes critical. To clarify the nature of the rare unexpected events
occurring in the process of cutting of DNA with restriction enzymes
then ligating it, we studied the molecular characteristics of
unexpected plasmid DNAs that were retrieved as mutants of the plasmid
after transfection to E. coli. The plasmid used was pUR288, containing
lacZ as a marker of mutation. It was digested with restriction enzymes
under the conditions recommended by the supplier of the enzymes and
under the presence of DMSO, which is known to induce star activity of
the enzymes. Comparisons of mutant frequencies and of nucleotide
sequences of the mutants found in the different conditions indicated
that nonspecific endonucleolytic activity similar to that found under
star activity was present under the recommended conditions and,
further, was responsible for the creation of deletion-type mutations.
The frequency of these events ranged from 10^-5 to 10^-3, depending
on the kind of restriction enzymes analyzed. Although the levels of the
nonspecificity were not high, they should be considered in assays such
as mutation and mistakes in DNA repair, where rare events are examined.

<>

<1>Nakamura, T., Maeda, Y., Oka, T., Tabata, H., Futai, M., Kawai, T.
<2>Atomic force microscope observation of plasmid deoxyribose nucleic acid with restriction enzyme.
<3>J. Vac. Sci. Technol. B
<4>17
<5>288-293
<6>1999
<7>We have observed plasmid deoxyribose nucleic acid, in real space images, before and after
treating with restriction enzyme (PvuII or HincII) using an atomic force microscope.  The
enzyme is recognized on DNA even in the absence of Mg2+ ions.  In the presence of Mg2+, on the
other hand, direct evidence was obtained that the enzyme could bind to circular plasmid DNA
without cutting and cleaved it at the site corresponding to the specificity.  Lengths of the
DNA fragments observed by AFM were consistent with the values estimated by agarose gel
electrophoresis.  In addition, as substrates for the AFM observation, rutile TiO2(110) single
crystal surface was found to be effective in expanding and fixing the DNA molecules as
straight chains along the stepped surface.

<>

<1>Nakamura, Y. et al.
<2>Complete genome structure of the thermophilic cyanobacterium Thermosynechococcus elongatus BP-1.
<3>DNA Res.
<4>9
<5>123-130
<6>2002
<7>The entire genome of a thermophilic unicellular cyanobacterium, Thermosynechococcus elongatus
BP-1, was sequenced. The genome consisted of a circular chromosome 2,593,857 bp long, and no
plasmid was detected. A total of 2475 potential protein-encoding genes, one set of rRNA genes,
42 tRNA genes representing 42 tRNA species and 4 genes for small structural RNAs were assigned
to the chromosome by similarity search and computer prediction. The translated products of 56%
of the potential protein-encoding genes showed sequence similarity to experimentally
identified and predicted proteins of known function, and the products of 34% of these genes
showed sequence similarity to the translated products of hypothetical genes. The remaining 10%
lacked significant similarity to genes for predicted proteins in the public DNA databases.
Sixty-three percent of the T. elongatus genes showed significant sequence similarity to those
of both Synechocystis sp. PCC 6803 and Anabaena sp. PCC 7120, while 22% of the genes were
unique to this species, indicating a high degree of divergence of the gene information among
cyanobacterial strains. The lack of genes for typical fatty acid desaturases and the presence
of more genes for heat-shock proteins in comparison with other mesophilic cyanobacteria may be
genomic features of thermophilic strains. A remarkable feature of the genome is the presence
of 28 copies of group II introns, 8 of which contained a presumptive gene for maturase/reverse
transcriptase. A trace of genome rearrangement mediated by the group II introns was also
observed.

<>

<1>Nakamura, Y. et al.
<2>Complete genome structure of the thermophilic cyanobacterium Thermosynechococcus elongatus BP-1 (supplement).
<3>DNA Res.
<4>9
<5>135-148
<6>2002
<7>none

<>

<1>Nakamura, Y. et al.
<2>Complete genome structure of Gloeobacter violaceus PCC 7421, a cyanobacterium that lacks thylakoids.
<3>DNA Res.
<4>10
<5>137-145
<6>2003
<7>The nucleotide sequence of the entire genome of a cyanobacterium Gloeobacter violaceus PCC
7421 was determined. The genome of G. violaceus
was a single circular chromosome 4,659,019 bp long with an average GC
content of 62%. No plasmid was detected. The chromosome comprises 4430
potential protein-encoding genes, one set of rRNA genes, 45 tRNA genes
representing 44 tRNA species and genes for tmRNA, B subunit of RNase P,
SRP RNA and 6Sa RNA. Forty-one percent of the potential protein-encoding
genes showed sequence similarity to genes of known function, 37% to
hypothetical genes, and the remaining 22% had no apparent similarity to
reported genes. Comparison of the assigned gene components with those of
other cyanobacteria has unveiled distinctive features of the G. violaceus
genome. Genes for PsaI, PsaJ, PsaK, and PsaX for Photosystem I and PsbY,
PsbZ and Psb27 for Photosystem II were missing, and those for PsaF, PsbO,
PsbU, and PsbV were poorly conserved. cpcG for a rod core linker peptide
for phycobilisomes and nblA related to the degradation of phycobilisomes
were also missing. Potential signal peptides of the presumptive products
of petJ and petE for soluble electron transfer catalysts were less
conserved than the remaining portions. These observations may be related
to the fact that photosynthesis in G. violaceus takes place not in
thylakoid membranes but in the cytoplasmic membrane. A large number of
genes for sigma factors and transcription factors in the LuxR, LysR, PadR,
TetR, and MarR families could be identified, while those for major
elements for circadian clock, kaiABC were not found. These differences may
reflect the phylogenetic distance between G. violaceus and other
cyanobacteria.

<>

<1>Nakanishi, M., Meirelles, P., Suzuki, R., Takatani, N., Mino, S., Suda, W., Oshima, K., Hattori, M., Ohkuma, M., Hosokawa, M., Miyashita, K., Thompson, F.L., Niwa, A., Sawabe, T., Sawabe, T.
<2>Draft Genome Sequences of Marine Flavobacterium Nonlabens Strains NR17, NR24, NR27, NR32, NR33, and Ara13.
<3>Genome Announcements
<4>2
<5>e01165-14
<6>2014
<7>Here, we present the draft genome sequences of six carotenoid producers affiliated with
Nonlabens spp. isolated from marine environments in both the
northern and southern parts of Japan. The genomic information will help to
elucidate the function and evolution of carotenoid synthetic gene clusters not
only in the genus Nonlabens but also in the family Flavobacteriaceae.

<>

<1>Nakanishi, S., Tazumi, A., Moore, J.E., Millar, B.C., Matsuda, M.
<2>Molecular and comparative analyses of the full-length cytolethal distending toxin (cdt) gene operon and its adjacent genetic loci from urease-positive thermophilic  Campylobacter (UPTC) organisms.
<3>Br. J. Biomed. Sci.
<4>67
<5>208-215
<6>2010
<7>Molecular and comparative analyses of the full-length cytolethal distending toxin (cdt) gene
operon and its adjacent genetic loci (2.7-9.4 kilo base pairs in
length) are carried out with 12 urease-positive thermophilic Campylobacter (UPTC)
isolates using several polymerase chain reaction (PCR) primer pairs. Three
putative open reading frames (ORFs) for cdtA, cdtB and cdtC, two putative
promoters and a hypothetically intrinsic rho-independent transcription terminator
were identified in all the operons of the 12 UPTC isolates examined. Although the
number of amino acid residues slightly varied for the putative cdtA and cdtC
ORFs, those for the cdtB were similar among all the UPTC isolates, as well as the
six urease-negative (UN) C. lari examined previously. Regarding the cdt genes in
UPTC CF89-12, each ORF commenced with an ATG start codon and terminated with a
TAG stop codon for cdtA and cdtB and a TAA for cdtC. Start and stop codons of the
three ORFs for the other 11 UPTC isolates were identical to those from the UPTC
CF89-12 isolate except for the TTG start codon for cdtC in the two isolates
(NCTC12892 and 12893) and the TGA stop codon for cdtA in five isolates (A1, A2,
A3, 89049 and 92251). Two putative promoter structures, consisting of sequences
at the -35-like (TTAATA) and -10-like (TATTAA) regions, as well as the start
codon (ATG), were identified for the transcriptional promoter, immediately
upstream of the cdtA gene in all the 12 isolates, Although the genetic
heterogeneity of the cdtB gene locus occurred in all 28 C. lari isolates (n = 16
UN C. lari; n = 12 UPTC) examined, all nine amino acid-specific DNase residues
were completely conserved in all their cdtB genes. Variable gene insertions with
heterogeneous order and combinations occurred between cdtC and lpxB genes in the
all UPTC organisms examined.

<>

<1>Nakano, Y., Steward, N., Sekine, M., Kusano, T., Sano, H.
<2>A tobacco NtMET1 cDNA encoding a DNA methyltransferase: Molecular characterization and abnormal phenotypes of transgenic tobacco plants.
<3>Plant Cell Physiol.
<4>41
<5>448-457
<6>2000
<7>A cDNA encoding a DNA methyltransferase, with a predicted polypeptide of 1556 amino acid
residues containing all motifs conserved in this
enzyme family, was isolated from tobacco plants, and the corresponding
gene was designated as NtMET1, RNA blot analysis indicated NtMET1
transcripts to accumulate in dividing tissues of tobacco plants, and
they could be detected during the S phase in synchronized dividing BY2
cells. In situ hybridization revealed the transcripts to be localized
exclusively in actively proliferating tissues around axillary apical
meristem. In order to ascertain physiological roles, transgenic tobacco
plants that had the antisense construct were made and examined for
phenotypes. Methylation levels of genomic DNA from transgenic plants
significantly decreased in comparison with wild-type levels, and
distinct phenotypic changes including small leaves, short internodes
and abnormal flower morphology were noted. Microscopic observation
revealed that leaf structure differed between transgenic and wild-type
plants. These results suggest that NtMET1 functions during DNA
replication, and that DNA methylation plays an important role in plant
morphogenesis.

<>

<1>Nakao, K., Chinen, A., Nobusato, A., Fujitani, Y., Uchiyama, I., Kobayashi, I.
<2>Relation between restriction modification genes and genome rearrangements suggested from genome sequence comparison within genus  Neisseria.
<3>Genome Inf. Ser.
<4>12
<5>398-399
<6>2001
<7>Restriction-modification gene complexes, such as EcoRI, encode two enzymatic functions,
restriction and modification.  A restriction enzyme will recognize a specific sequence in DNA
and cut the DNA unless it is methylated by a cognate modification enzyme.  RM systems will
defend bacterial cells by attacking incoming foreign DNA.  It is widely held that bacteria
have evolved RM systems and maintain them in order to protect their genome from invasion by
foreign DNA such as bacteriophages and plasmids.

<>

<1>Nakatsu, C.H., Barabote, R., Thompson, S., Bruce, D., Detter, C., Brettin, T., Han, C., Beasley, F., Chen, W., Konopka, A., Xie, G.
<2>Complete genome sequence of Arthrobacter sp. strain FB24.
<3>Standards in Genomic Sciences
<4>9
<5>106-116
<6>2013
<7>Arthrobacter sp. strain FB24 is a species in the genus Arthrobacter Conn and Dimmick 1947, in
the family Micrococcaceae and class Actinobacteria. A number of
Arthrobacter genome sequences have been completed because of their important role
in soil, especially bioremediation. This isolate is of special interest because
it is tolerant to multiple metals and it is extremely resistant to elevated
concentrations of chromate. The genome consists of a 4,698,945 bp circular
chromosome and three plasmids (96,488, 115,507, and 159,536 bp, a total of
5,070,478 bp), coding 4,536 proteins of which 1,257 are without known function.
This genome was sequenced as part of the DOE Joint Genome Institute Program.

<>

<1>Nakayama, H., Morinaga, Y., Nomura, N., Nunoura, T., Sako, Y., Uchida, A.
<2>An archaeal homing endonuclease I-PogI cleaves at the insertion site of the neighboring intron, which has no nested open reading frame.
<3>FEBS Lett.
<4>544
<5>165-170
<6>2003
<7>Homing endonucleases (HEs) of the LAGLIDADG family cleave intron/inteinless cognate DNA at, or
near, the insertion site (IS) of
their own intron/intein. Here, we describe a notable exception to this
rule. Two introns, Pog.S1205 (length 32 bp) and Pog.S1213 (664 bp), whose
ISs are 8 bp apart, exist within the 16S rRNA gene of the archaeon
Pyrobaculum oguniense. Pog.S1213 harbors a nested open reading frame (ORF)
encoding a 22 kDa monomeric protein, I-PogI, which contains two LAGLIDADG
motifs and has optimal DNA cleavage activity at 90 degrees C.
Intriguingly, I-PogI cleaves the Pog.S1205-less substrate DNA in the
presence or absence of Pog.S1213. The cleavage site (CS) of I-PogI does
not coincide with the IS of Pog.S1213 but with that of Pog.S1205. Thus,
I-PogI activity both promotes the homing of its own intron, Pog.S1213, and
guarantees co-conversion of the ORF-less intron Pog.S1205.

<>

<1>Nakayama, H., Shimamura, T., Imagawa, T., Shirai, N., Itoh, T., Sako, Y., Miyano, M., Sakuraba, H., Ohshima, T., Nomura, N., Tsuge, H.
<2>Structure of a hyperthermophilic archaeal homing endonuclease, I-Tsp061I: Contribution of cross-domain polar networks to thermostability.
<3>J. Mol. Biol.
<4>365
<5>362-378
<6>2007
<7>A novel LAGLIDADG-type homing endonuclease (HEase), I-TspO61I, from the hyperthermophilic
archaeon Thermoproteus sp. IC-061 16 S rRNA gene
(rDNA) intron was characterized with respect to its structure,
catalytic properties and thermostability. It was found that I-Tsp061I
is a HEase isoschizomer of the previously described I-PogI and exhibits
the highest thermostability among the known LAGLIDADG-type HEases.
Determination of the crystal structure of I-Tsp061I at 2.1 A resolution
using the multiple isomorphous replacement and anomalous scattering
method revealed that the overall fold is similar to that of other known
LAGLIDADG-type HEases, despite little sequence similarity between
I-TspO61I and those HEases. However, I-Tsp061I contains important
cross-domain polar networks, unlike its mesophilic counterparts.
Notably, the polar network Tyr6-Asp104-His180-107O-HOH12-104O-Asn177
exists across the two packed a-helices containing both the LAGLIDADG
catalytic motif and the GxxxG hydrophobic helix bundle motif. Another
important structural feature is the salt-bridge network
Asp29-Arg31-GIu182 across N and C-terminal domain interface, which
appears to contribute to the stability of the domain/domain packing. On
the basis of these structural analyses and extensive mutational
studies, we conclude that such cross-domain polar networks play key
roles in stabilizing the catalytic center and domain packing, and
underlie the hyperthermostability of T-Tsp061I.

<>

<1>Nakayama, K., Endo, M., Fujitsuka, M., Majima, T.
<2>Monitoring of three distinct structures of restriction enzyme complexes using characteristic fluorescence from site-selectively incorporated solvatochromic probe.
<3>Photochem. Photobiolog.
<4>6
<5>836-841
<6>2007
<7>The local change in the three different structures of restriction enzyme BamHI, which include
DNA-free dimer and non-specific and
specific complexes with DNA, were detected by the fluorescence from a
site-selectively introduced solvatochromic fluorophore
N-beta-L-alanyl-5-(N,N-dimethylamino)naphthalene-1-sulfonamide
(DanAla). According to the crystal structure, alpha-helices of the
non-specific complex containing Ile82, Glu86 and Trp206 residues are
converted into random coil by the formation of specific complex with a
substrate. To understand the microenvironmental change caused by the
structural transition around these positions, the DanAla probe was
site-specifically introduced into the positions, and steady-state and
time-resolved fluorescence was observed. The steady-state fluorescence
gave us information that the rigidity of the polypeptide chains would
be enhanced by the formation of the specific complex. The time-resolved
fluorescence supported that the change in a water molecule-accessible
space was induced by DNA-binding. We revealed that the change in
rigidity and solvation around the specific positions was detected by
the characteristic fluorescence using the combination of steady-state
and time-resolved fluorescence techniques.

<>

<1>Nakayama, K., Endo, M., Majima, T.
<2>A hydrophilic azobenzene-bearing amino acid for photochemical control of a restriction enzyme BamHI.
<3>Bioconjugate Chem.
<4>16
<5>1360-1366
<6>2005
<7>A novel hydrophilic and negatively charged azobenzene-bearing amino acid,
4'-carboxyphenylazophenylalanine (azoAla 1), has been designed
and synthesized for investigation of the photochemical regulation of
the enzyme activity. The properties of photoisomerization and thermal
stability of the cis-isomer were similar to those of a commonly used
phenylazophenylalanine (azoAla 2). For photochemical control of the
enzyme, these two azobenzene-bearing amino acids were incorporated into
the specific position at the dimer interface of a restriction enzyme
BamHI. These trans-azobenzene derivatives in the BamHI suppressed the
enzymatic activity, and the following photoirradiation at 366 nm
induced the recovery of its activity. Although the activities of both
azoAla-BamHI mutants were same level after a long time irradiation, the
recovery of the activity of azoAla 1-BamHI was faster than that of
azoAla 2-BamHI with a short time irradiation. This result suggests that
the negatively charged carboxylate group introduced into an azobenzene
moiety affects the behavior of azoAla in the protein scaffold during
the trans-cis photoisomerization.

<>

<1>Nakayama, K., Endo, M., Majima, T.
<2>Photochemical regulation of the activity of a restriction enzyme BamHI using an azobenzene moiety incorporated into the dimer interface.
<3>ACS Abstracts
<4>229
<5>U399-U400
<6>2005
<7>We describe the control of enzymatic activity by photochemical regulation of protein-protein
interaction.  Restriction enzyme BamHI has a typical dimer interface with salt-bridge network
and need the dimer formation to show the activity.  Using this enzyme, we designed the
photochemically controllable BamHI, which has a photofunctional molecule in the dimer
interface for inactivation, and initiates the activity with photoirradiation (366 nm).
Photoisomerizable trans-phenylazophenylalanine (trans-azoAla) was site-selectively introduced
at 132 position in the dimer interface.  The photofunctional BamHI showed no activity, and the
following photoisomerization induced the activity.  These results suggest that, the bulky
trans-azoAla may induce the misalignment of the two BamHI monomers as an inactive dimer form,
while the compact cis-azoAla may allow the specific hydrogen bondings in the dimer interface
as similar to the wild-type BamHI.  By employing the phosoisomerization of azoAla residue, we
have successfully constructed photofunctional BamHI which is activated by photoirradiation.

<>

<1>Nakayama, K., Yamashita, A., Kurokawa, K., Morimoto, T., Ogawa, M., Fukuhara, M., Urakami, H., Ohnishi, M., Uchiyama, I., Ogura, Y., Ooka, T., Oshima, K., Tamura, A., Hattori, M., Hayashi, T.
<2>The Whole-genome Sequencing of the Obligate Intracellular Bacterium Orientia tsutsugamushi Revealed Massive Gene Amplification During  Reductive Genome Evolution.
<3>DNA Res.
<4>15
<5>185-199
<6>2008
<7>Scrub typhus ('Tsutsugamushi' disease in Japanese) is a mite-borne infectious disease. The
causative agent is Orientia tsutsugamushi, an
obligate intracellular bacterium belonging to the family Rickettsiaceae of
the subdivision alpha-Proteobacteria. In this study, we determined the
complete genome sequence of O. tsutsugamushi strain Ikeda, which comprises
a single chromosome of 2 008 987 bp and contains 1967 protein coding
sequences (CDSs). The chromosome is much larger than those of other
members of Rickettsiaceae, and 46.7% of the sequence was occupied by
repetitive sequences derived from an integrative and conjugative element,
10 types of transposable elements, and seven types of short repeats of
unknown origins. The massive amplification and degradation of these
elements have generated a huge number of repeated genes (1196 CDSs,
categorized into 85 families), many of which are pseudogenes (766 CDSs),
and also induced intensive genome shuffling. By comparing the gene content
with those of other family members of Rickettsiacea, we identified the
core gene set of the family Rickettsiaceae and found that, while much more
extensive gene loss has taken place among the housekeeping genes of
Orientia than those of Rickettsia, O. tsutsugamushi has acquired a large
number of foreign genes. The O. tsutsugamushi genome sequence is thus a
prominent example of the high plasticity of bacterial genomes, and
provides the genetic basis for a better understanding of the biology of O.
tsutsugamushi and the pathogenesis of 'Tsutsugamushi' disease.

<>

<1>Nakayama, T., Kamikawa, R., Tanifuji, G., Kashiyama, Y., Ohkouchi, N., Archibald, J.M., Inagaki, Y.
<2>Complete genome of a nonphotosynthetic cyanobacterium in a diatom reveals recent adaptations to an intracellular lifestyle.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>111
<5>11407-11412
<6>2014
<7>The evolution of mitochondria and plastids from bacterial endosymbionts were key
events in the origin and diversification of eukaryotic cells. Although the
ancient nature of these organelles makes it difficult to understand the earliest
events that led to their establishment, the study of eukaryotic cells with
recently evolved obligate endosymbiotic bacteria has the potential to provide
important insight into the transformation of endosymbionts into organelles.
Diatoms belonging to the family Rhopalodiaceae and their endosymbionts of
cyanobacterial origin (i.e., "spheroid bodies") are emerging as a useful model
system in this regard. The spheroid bodies, which appear to enable rhopalodiacean
diatoms to use gaseous nitrogen, became established after the divergence of
extant diatom families. Here we report what is, to our knowledge, the first
complete genome sequence of a spheroid body, that of the rhopalodiacean diatom
Epithemia turgida. The E. turgida spheroid body (EtSB) genome was found to
possess a gene set for nitrogen fixation, as anticipated, but is reduced in size
and gene repertoire compared with the genomes of their closest known free-living
relatives. The presence of numerous pseudogenes in the EtSB genome suggests that
genome reduction is ongoing. Most strikingly, our genomic data convincingly show
that the EtSB has lost photosynthetic ability and is metabolically dependent on
its host cell, unprecedented characteristics among cyanobacteria, and
cyanobacterial symbionts. The diatom-spheroid body endosymbiosis is thus a unique
system for investigating the processes underlying the integration of a bacterial
endosymbiont into eukaryotic cells.

<>

<1>Nakayama, Y., Kobayashi, I.
<2>Restriction-modification gene complexes as selfish gene entities: roles of a regulatory system in their establishment, maintenance, and apoptotic mutual exclusion.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>6442-6447
<6>1998
<7>We have reported some type II restriction-modification gene complexes on plasmids resist
displacement by an incompatible plasmid through postsegregational host killing.  Such selfish
behavior may have contributed to the spread and maintenance of RM systems.  Here we analyze
the role of regulatory genes, often found linked to RM gene complexes, in their interaction
with the host and the other RM gene complexes.  We identified the C gene of EcoRV as a
positive regulator of restriction.  A C mutation eliminated postsegregational killing by
EcoRV.  The C system has been proposed to allow establishment of RM systems in new hosts by
delaying the appearance of restriction activity.  Consistent with this proposal, bacteria
pre-expressing ecoRVC were transformed at a reduced efficiency by plasmids carrying the EcoRV
RM gene complex.  Cells carrying the BamHI RM gene complex were transformed at a reduced
efficiency by a plasmid carrying a PvuII RM gene complex, which shares the same C specificity.
The reduction most likely was caused by chromosome cleavage at unmodified PvuII sites by
prematurely expressed PvuII restriction enzyme.  Therefore, association of the C genes of the
same specificity with RM gene complexes of different sequence specificities can confer on a
resident RM gene complex the capacity to abort establishment of a second, incoming RM gene
complex.  This phenomenon, termed "apoptotic mutual exclusion," is reminiscent of suicidal
defense against virus infection programmed by other selfish elements.  PvuIIC and bamHIC genes
define one incompatibility group of exclusion whereas ecoRVC gene defines another.

<>

<1>Nakayashiki, T., Nishimura, K., Inokuchi, H.
<2>Cloning and sequencing of a previously unidentified gene that is involved in the biosynthesis of heme in Escherichia coli.
<3>Gene
<4>153
<5>67-70
<6>1995
<7>We have isolated a number of porphyrin (Por)-synthesis mutants as light-resistant revertants
of a light-sensitive strain delta visA (hemH) of Escherichia coli that accumulates protoPor IX
in the cell.  Among such mutants, we found a double mutant (H103) with mutations in hemA and
in a new gene downstream of hemA. This new gene, designated hemK, was located at 27 min on the
linkage map of the E. coli chromosome. By nucleotide (nt) sequencing, it was demonstrated that
hemK forms part of the hemA-prfA-hemK operon and encodes 225 amino acids that show no
significant homology to any protein in the standard databases. The mutant strain H103 formed
small colonies and showed no catalase activity even in the presence of 5-aminolevulinic acid
(ALA), indicating its inability to catalyze a step in the biosynthesis of heme from ALA. An
extract of H103 cells has readily detectable ALA dehydratase and porphobilinogen deaminase
activities. H103 cells carrying a plasmid that included only hemA as an insert accumulated
protoPor and coproPor, but showed no sensitivity to light, a result that suggests that it may
be deficient in protoporphyrinogen oxidase activity.

<>

<1>Nakonieczna, J., Kaczorowski, T., Obarska-Kosinska, A., Bujnicki, J.M.
<2>Functional Analysis of MmeI from Methanol Utilizer Methylophilus methylotrophus, a Subtype IIC Restriction-Modification Enzyme Related to Type I Enzymes.
<3>Appl. Environ. Microbiol.
<4>75
<5>212-223
<6>2009
<7>MmeI from Methylophilus methylotrophus belongs to the type II restriction-modification
enzymes. It recognizes an asymmetric DNA
sequence, 5'-TCCRAC-3' (R indicates G or A), and cuts both strands at
fixed positions downstream of the specific site. This particular
feature has been exploited in transcript profiling of complex genomes
(using serial analysis of gene expression technology). We have shown
previously that the endonucleolytic activity of MmeI is strongly
dependent on the presence of S-adenosyl-L-methionine (J. Nakonieczna,
J. W. Zmijewski, B. Banecki, and A. J. Podhajska, Mol. Biotechnol. 37:
127-135, 2007), which puts MmeI in subtype IIG. The same cofactor is
used by MmeI as a methyl group donor for modification of an adenine in
the upper strand of the recognition site to N-6-methyladenine. Both
enzymatic activities reside in a single polypeptide (919 amino acids
[aa]), which puts MmeI also in subtype IIC of the
restriction-modification systems. Based on a molecular model, generated
with the use of bioinformatic tools and validated by site-directed
mutagenesis, we were able to localize three functional domains in the
structure of the MmeI enzyme: (i) the N-terminal portion containing the
endonucleolytic domain with the catalytic Mg2+-binding motif
D-70-X-9-EXK82, characteristic for the PD-(D/E)XK superfamily of
nucleases; (ii) a central portion (aa 310 to 610) containing nine
sequence motifs conserved among N-6-adenine gamma-class DNA
methyltransferases; (iii) the C-terminal portion (aa 610 to 919)
containing a putative target recognition domain. Interestingly, all
three domains showed highest similarity to the corresponding elements
of type I enzymes rather than to classical type II enzymes. We have
found that MmeI variants deficient in restriction activity (D70A, E80A,
and K82A) can bind and methylate specific nucleotide sequence. This
suggests that domains of MmeI responsible for DNA restriction and
modification can act independently. Moreover, we have shown that a
single amino acid residue substitution within the putative target
recognition domain (S807A) resulted in a MmeI variant with a higher
endonucleolytic activity than the wild-type enzyme.

<>

<1>Nakonieczna, J., Zmijewski, J.W., Banecki, B., Podhajska, A.J.
<2>Binding of MmeI restriction-modification enzyme to its specific recognition sequence is stimulated by S-adenosyl-L-methionine.
<3>Mol. Biotechnol.
<4>37
<5>127-135
<6>2007
<7>Restriction endonucleases serve as a very good model for studying specific protein-DNA
interaction. MmeI is a very interesting
restriction endonuclease, but although it is useful in Serial Analysis
of Gene Expression, still very little is known about the mechanism of
its interaction with DNA. MmeI is a unique enzyme as besides cleaving
DNA it also methylates specific sequence. For endonucleolytic activity
MmeI requires Mg(II) and S-adenosyl-L-methionine (AdoMet). AdoMet is a
methyl donor in the methylation reaction, but its requirement for DNA
cleavage remains unclear. In the present article we investigated MmeI
interaction with DNA with the use of numerous methods. Our
electrophoretic mobility shift assay revealed formation of two types of
specific protein-DNA complexes. We speculate that faster migrating
complex consists of one protein molecule and one DNA fragment whereas,
slower migrating complex, which appears in the presence of AdoMet, may
be a dimer or multimer form of MmeI interacting with specific DNA.
Additionally, using spectrophotometric measurements we showed that in
the presence of AdoMet, MmeI protein undergoes conformational changes.
We think that such change in the enzyme structure, upon addition of
AdoMet, may enhance its specific binding to DNA. In the absence of
AdoMet MmeI binds DNA to the much lower extent.

<>

<1>Nam, S.H., Choi, S.H., Kang, A., Kim, D.W., Kim, D.S., Kim, R.N., Kim, A., Park, H.S.
<2>Genome Sequence of Leuconostoc fallax KCTC 3537.
<3>J. Bacteriol.
<4>193
<5>588-589
<6>2010
<7>Leuconostoc fallax is known to be present during the manufacturing process of kimchi, the
best-known traditional Korean dish. Here, we present the
draft genome sequence of the type strain Leuconostoc fallax KCTC 3537
(1,638,971 bp, with a G+C content of 37.5%), which consists of 30 large
contigs (>100 bp in size).

<>

<1>Nam, S.H., Choi, S.H., Kang, A., Kim, D.W., Kim, D.S., Kim, R.N., Kim, A., Park, H.S.
<2>Genome Sequence of Lactobacillus coryniformis subsp. coryniformis KCTC 3167.
<3>J. Bacteriol.
<4>193
<5>1014-1015
<6>2011
<7>Lactobacillus coryniformis subsp. coryniformis is known to be present during the manufacturing
process of kimchi, the best-known traditional Korean dish. Here, we present the draft genome
sequence of Lactobacillus coryniformis subsp. coryniformis type strain KCTC 3167 (2,964,752
bp, with a G+C content of 42.8%), which consists of 55 scaffolds.

<>

<1>Nam, S.H., Choi, S.H., Kang, A., Kim, D.W., Kim, R.N., Kim, A., Kim, D.S., Park, H.S.
<2>Genome Sequence of Lactobacillus animalis KCTC 3501.
<3>J. Bacteriol.
<4>193
<5>1280-1281
<6>2010
<7>Lactobacillus animalis is one of the most prevalent lactic acid bacteria present during the
manufacturing process of kimchi, the best-known traditional Korean dish. Here, we present the
draft genome sequence of the type strain Lactobacillus animalis KCTC 3501 (1,882,795 bp, with
a G+C content of 41.1%), which consists of 7 scaffolds.

<>

<1>Nam, S.H., Choi, S.H., Kang, A., Kim, D.W., Kim, R.N., Kim, A., Kim, D.S., Park, H.S.
<2>Genome Sequence of Lactobacillus farciminis KCTC 3681.
<3>J. Bacteriol.
<4>193
<5>1790-1791
<6>2011
<7>Lactobacillus farciminis is one of the most prevalent lactic acid bacteria present during the
manufacturing process of kimchi, the best-known traditional Korean dish. Here, we present the
draft genome sequence of the type strain Lactobacillus farciminis KCTC 3681 (2,498,309 bp,
with a G+C content of 36.4%), which consists of 5 scaffolds.

<>

<1>Nam, S.H., Choi, S.H., Kang, A., Kim, D.W., Kim, R.N., Kim, A., Park, H.S.
<2>Genome Sequence of Leuconostoc argentinum KCTC 3773.
<3>J. Bacteriol.
<4>192
<5>6490-6491
<6>2010
<7>Leuconostoc argentinum is one of the most prevalent lactic acid bacteria present during the
manufacturing process of kimchi, the best-known
traditional Korean dish. Here, we present the draft genome sequence of
type strain KCTC 3773 of Leuconostoc argentinum (1,720,683 bp, with a G+C
content of 42.9%), which consists of 98 large contigs (>100 bp in size).

<>

<1>Nam, S.H., Choi, S.H., Kang, A., Kim, D.W., Kim, R.N., Kim, D.S., Kim, A., Park, H.S.
<2>Genome Sequence of Lactobacillus suebicus KCTC 3549.
<3>J. Bacteriol.
<4>193
<5>5532-5533
<6>2011
<7>Lactobacillus suebicus is important in the generation of particular flavors and in other
ripening processes associated with apple mash. Here,
we present the draft genome sequence of the type strain Lactobacillus
suebicus KCTC 3549 (2,656,936 bp, with a G+C content of 39.0%), which
consists of 143 large contigs (>100 bp).

<>

<1>Nam, S.H., Choi, S.H., Kang, A., Lee, K.S., Kim, D.W., Kim, R.N., Kim, D.S., Park, H.S.
<2>Genome Sequence of Lactobacillus fructivorans KCTC 3543.
<3>J. Bacteriol.
<4>194
<5>2111-2112
<6>2012
<7>Lactobacillus fructivorans is important in the generation of particular flavors and in other
ripening processes associated with fermented food. Here, we present
the draft genome sequence of the type strain Lactobacillus fructivorans KCTC 3543
(1,373,326 bp, with a G+C content of 38.9%), which consists of 5 scaffolds. The
genome sequence was obtained by using a whole-genome shotgun strategy with Roche
454 GS (FLX Titanium) pyrosequencing, and all of the reads were assembled using
Newbler Assembler 2.3.

<>

<1>Nam, S.H., Kim, A., Choi, S.H., Kang, A., Kim, D.W., Kim, R.N., Kim, D.S., Park, H.S.
<2>Genome Sequence of Leuconostoc carnosum KCTC 3525.
<3>J. Bacteriol.
<4>193
<5>6100-6101
<6>2011
<7>We announce the draft genome sequence of the type strain Leuconostoc carnosum KCTC 3525
(3,234,408 bp with a G+C content of 40.9%), one of the
most prevalent lactic acid bacteria present during the manufacturing
process of vacuum-packaged meats, which consists of 2,407 large contigs
(>500 bp in size). The genome sequence was obtained by a whole-genome
shotgun strategy using Roche 454 GS (FLX Titanium) pyrosequencing, and all
of the reads were assembled using Newbler Assembler 2.3.

<>

<1>Nam, Y.D., Chung, W.H., Seo, M.J., Lim, S.I.
<2>Draft Genome Sequence of Staphylococcus vitulinus F1028, a Strain Isolated from a Block of Fermented Soybean.
<3>J. Bacteriol.
<4>194
<5>5961-5962
<6>2012
<7>Staphylococcus vitulinus is a coagulase-negative staphylococcus in the family
Staphylococcaceae. This report describes the draft genome sequence of S.
vitulinus F1028, which was isolated from a traditional Korean soybean food
(meju). This 2.56-Mbp genome sequence is the first S. vitulinus genome of a
strain isolated from a fermented soybean product.

<>

<1>Nam, Y.D., Chung, W.H., Seo, M.J., Lim, S.I., Yi, S.H.
<2>Genome Sequence of Staphylococcus lentus F1142, a Strain Isolated from Korean Soybean Paste.
<3>J. Bacteriol.
<4>194
<5>5987
<6>2012
<7>This report describes the draft genome sequence of Staphylococcus lentus F1142, which was
isolated from a Korean fermented soybean paste (doenjang). The draft
genome sequence contained 2.79 Mbp with a G+C content of 31.8%; this is the first
S. lentus genome to be reported.

<>

<1>Nam, Y.D., Lee, H.W., Lee, M., Yim, K.J., Kim, K.N., Roh, S.W., Kim, D.
<2>Draft Genome Sequence of Gillisia sp. Strain CBA3202, a Novel Member of the Genus Gillisia, Which Belongs to the Family Flavobacteriaceae.
<3>J. Bacteriol.
<4>194
<5>3739
<6>2012
<7>Gillisia sp. strain CBA3202, which belongs to the family Flavobacteriaceae, was isolated from
sand of the seashore on Jeju Island, Republic of Korea. The draft
genome of Gillisia sp. CBA3202 contains 2,981,404 bp with a G+C content of 34.9%.
This is the second genome sequence of the Gillisia strains.

<>

<1>Nam, Y.D., Seo, M.J., Lim, S.I., Lee, S.Y.
<2>Genome Sequence of Lysinibacillus boronitolerans F1182, Isolated from a Traditional Korean Fermented Soybean Product.
<3>J. Bacteriol.
<4>194
<5>5988
<6>2012
<7>Lysinibacillus is a Gram-positive, rod-shaped, and round-spore-forming bacterial  genus of the
family Bacillaceae. We analyzed the genome sequence of
Lysinibacillus boronitolerans F1182, isolated from a traditional Korean fermented
soybean product. The genome sequence contained 4.46 Mbp with a G+C content of
37.5%. This is the first report of an L. boronitolerans genome.

<>

<1>Nam, Y.D., Seo, M.J., Lim, S.I., Park, S.L.
<2>Genome Sequence of Kocuria atrinae C3-8, Isolated from Jeotgal, a Traditional Korean Fermented Seafood.
<3>J. Bacteriol.
<4>194
<5>5996
<6>2012
<7>Kocuria is a Gram-positive coccus, catalase-positive, coagulase-negative, strictly aerobic
bacterial genus in the family Micrococcaceae. Kocuria atrinae
C3-8 was isolated from a traditional Korean fermented seafood. This study
describes the first genome sequence of K. atrinae strain C3-8, which has a
3.19-Mbp genome and a G+C content of 63.8%.

<>

<1>Nan, X., Ng, H.-H., Johnson, C.A., Laherty, C.D., Turner, B.M., Eisenman, R.N., Bird, A.
<2>Transcriptional repression by the methyl-CpG-binding protein MeCP2 involves a histone deacetylase complex.
<3>Nature
<4>393
<5>386-389
<6>1998
<7>Cytosine residues in the sequence 5'CpG (cytosine-guanine) are often postsynthetically
methylated in animal genomes.  CpG methylation is involved in long-term silencing of certain
genes during mammalian development and in repression of viral genomes.  The methyl-CpG-binding
proteins MeCP1 and MeCP2 interact specifically with methylated DNA and mediate transcriptional
repression.  Here we study the mechanism of repression by MeCP2, an abundant nuclear protein
that is essential for mouse embryogenesis.  MeCP2 binds tightly to chromosomes in a
methylation-dependent manner.  It contains a transcriptional-repression domain that can
function at a distance in vitro and in vivo.  We show that a region of MeCP2 that localizes
with the TRD associates with a corepressor complex containing the transcriptional repressor
mSin3A and histone deacetylases.  Transcriptional repression in vivo is relieved by the
deacetylase inhibitor trichostatin A, indicating the deacetylation of histones (and/or of
other proteins) is an essential component of this repression mechanism.  The data suggest that
two global mechanisms of gene regulation, DNA methylation and histone deacetylation, can be
linked by MeCP2.

<>

<1>Nandasena, K. et al.
<2>Complete genome sequence of Mesorhizobium ciceri bv. biserrulae type strain (WSM1271(T)).
<3>Standards in Genomic Sciences
<4>9
<5>462-472
<6>2014
<7>Mesorhizobium ciceri bv. biserrulae strain WSM1271(T) was isolated from root nodules of the
pasture legume Biserrula pelecinus growing in the Mediterranean
basin. Previous studies have shown this aerobic, motile, Gram negative,
non-spore-forming rod preferably nodulates B. pelecinus - a legume with many
beneficial agronomic attributes for sustainable agriculture in Australia. We
describe the genome of Mesorhizobium ciceri bv. biserrulae strain WSM1271(T)
consisting of a 6,264,489 bp chromosome and a 425,539 bp plasmid that together
encode 6,470 protein-coding genes and 61 RNA-only encoding genes.

<>

<1>Naor, A., Lazary, R., Barzel, A., Papke, R.T., Gophna, U.
<2>In vivo characterization of the homing endonuclease within the polB gene in the halophilic archaeon Haloferax volcanii.
<3>PLoS ONE
<4>6
<5>e15833
<6>2011
<7>Inteins are parasitic genetic elements, analogous to introns that excise themselves at the
protein level by self-splicing, allowing the formation
of functional non-disrupted proteins. Many inteins contain a homing
endonuclease (HEN) gene, and rely on its activity for horizontal
propagation. In the halophilic archaeon, Haloferax volcanii, the gene
encoding DNA polymerase B (polB) contains an intein with an annotated but
uncharacterized HEN. Here we examine the activity of the polB HEN in vivo,
within its natural archaeal host. We show that this HEN is highly active,
and able to insert the intein into both a chromosomal target and an
extra-chromosomal plasmid target, by gene conversion. We also demonstrate
that the frequency of its incorporation depends on the length of the
flanking homologous sequences around the target site, reflecting its
dependence on the homologous recombination machinery. Although several
evolutionary models predict that the presence of an intein involves a
change in the fitness of the host organism, our results show that a strain
deleted for the intein sequence shows no significant changes in growth
rate compared to the wild type.

<>

<1>Narayanan, S., Deshpande, U.
<2>Whole-Genome Sequences of Four Clinical Isolates of Mycobacterium tuberculosis from Tamil Nadu, South India.
<3>Genome Announcements
<4>1
<5>e00186-13
<6>2013
<7>We report the annotated genome sequences of four clinical isolates of Mycobacterium
tuberculosis from Tamil Nadu, India.

<>

<1>Nardone, G., Blakesley, R.
<2>A new type II restriction enzyme from Haemophilus influenzae.
<3>Fed. Proc.
<4>40
<5>1848
<6>1981
<7>We have previously reported a new Type II restriction activity (Hin GUII) from
Haemophilus influenzae (Fed. Proc. 27: 1415, 1978).  These cells also possess
an isoschizomer of HhaI.  These two enzyme activities can be separated by
chromatography of the cell extract on phosphocellulose and single-stranded DNA
agarose.  Two possible recognition sequences were predicted for Hin GUII by the
computer program RESITE (Tolstoshev, C. and Blakesley, R., manuscript in prep.)
from the lengths of the eight fragments generated by digestion of PhiX174 RF
DNA.  By comparison of digestion products of SV40 (11 fragments) and pBR322 (12
fragments) DNAs, the sequence GGATG/CATCC was predicted.  This was confirmed by
mapping each on the Hin GUII sites PhiX174 RF DNA and direct DNA sequencing of
3 cleavage sites on PhiX174 RF and one site on pBR322 DNAs.  Hin GUII is a
member of a unique group of Type II restriction enzymes which bind to a
specific sequence and cleave at another site.  The Hin GUII cleavage site is
situated 9-11 bases 3' to the recognition sequence GGATG/CATCC.

<>

<1>Nardone, G., Chirikjian, J.G.
<2>The enzymes of the BamHI restriction-modification system.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>147-184
<6>1987
<7>The remarkable sequence specificity of Type II restriction endonucleases and
their cognate methyl transferases has recently placed them under the scrutiny
of nucleic acid enzymology.  From a biochemical perspective, their
comparatively uncomplicated reaction requirements, intermediate size and
concise, well-defined recognition sites has made them attractive models for the
investigation of specific protein-nucleic acid interactions.  The solution of
the same DNA sequence recognition problem by the enzymes of a
restriction-modification system has led to interesting questions concerning
structure-function relationships between these genetically distinct proteins.
Further study of restriction-modification enzymes should promote a better
understanding of the patterns and principles of protein-nucleic acid
interactions.  The type II restriction-modification enzymes of Bacillus
amyloliquefaciens H recognizes the duplex, symmetrical sequence 5'-GGATCC-3'.
In the presence of Mg+2 the endonuclease catalyzes double stranded cleavage
between the guanines, generating 5'-phophoryl and 3'-hydroxyl staggered
termini.  The methylase catalyzes methyl group transfer from
S-adenosyl-L-methionine to the C5 position of the internal cytosines.
Methylation prevents cleavage by the endonuclease and is the presumed host
controlled mechanism for the protection of endogenous DNA.  The specificity of
the methyl acceptor must be as stringent as the position of strand scission
since methylation of the external cytosines or the 6-amino groups of the
adenines does not prevent cleavage.  We have been involved with the
purification and characterization of these enzymes.  Emphasis has been placed
on their catalytic properties and mechanisms of sequence discrimination.

<>

<1>Nardone, G., Connaughton, J.F., Kaloss, W., Chirikjian, J.G.
<2>Changes in the kinetics of the BamHI restriction/modification enzymes with systematic alterations of the bases flanking the recognition site.
<3>FASEB J.
<4>4
<5>A1794
<6>1990
<7>Several type II restriction enzymes exhibit different reaction kinetics with
identical recognition sites in a given DNA molecule.  Kinetic preferences have
been ascribed to different sequences flanking each recognition site.  Using
site-specific mutagenesis, we have made 30 M13mp8 linear DNA substrates having
mutations within the first 3 nucleotides on both flanks of the single BamHI
site.  The advantages of these substrates are the constant position of the
BamHI site relative to the DNA termini, the elimination of changes in secondary
structure induced by supercoiling and the retention of all the flanking
sequences except for the specified base changes.  Initial reaction velocities
were determined in the presence of saturating DNA.  The greater difference in
cleavage rates was 5 fold.  No correlation was found between flanking G/C
content and reaction kinetics.  The endonuclease was sensitive to the position
of base substitution.  C and T in flanking positions 1 and 2 produced the
greatest reduction in velocites.  A in the second position of either flank
produced the greatest increase in velocity.  Double or triple substitutions of
A or T did not generate additive kinetic effects.  The methylase prefers
substrates containing A in flanking positions 1 and 2 but is inhibited by this
base in position 3.  T substitutions reduced methylation rates.

<>

<1>Nardone, G., George, J., Chirikjian, J.G.
<2>Differences in the Kinetic Properties of BamHI Endonuclease and Methylase with Linear DNA Substrates.
<3>J. Biol. Chem.
<4>261
<5>12128-12133
<6>1986
<7>BamHI endonuclease and methylase were found to exhibit a kinetic preference for linear pBR322
DNA substrates containing the recognition site in a central location.  The Km values for
substrates having the recognition site in a terminal location were approximately 3-fold
greater than those with a centrally located site.  This phenomenon may be partially due to
facilitated transfer of the enzymes to the recognition site over nonspecific flanking
sequences.  The exploitation of facilitated transfer by these enzymes has been inferred from
studies demonstrating kinetic preferences for longer DNA substrates.  The reaction rates of
the endonuclease were 9-fold greater with full-length pBR322 DNA than with a 74-base pair
derivative.  The methylase exhibits a kinetic preference for longer substrates but only under
conditions of comparatively higher DNA concentrations.  In addition, the methylase has the
property of increasing long chain preference with increasing salt concentrations up to 120 mM.
Increasing salt concentrations decreased the endonuclease's preference for longer substrates.
Nonspecific inhibition studies revealed qualitative and quantitative differences between the
two enzymes under catalytic conditions. These studies suggest that BamHI endonuclease and
methylase interact with nonspecific DNA in different ways.

<>

<1>Nardone, G., George, J., Chirikjian, J.G.
<2>Sequence-specific BamHI methylase.
<3>J. Biol. Chem.
<4>259
<5>10357-10362
<6>1984
<7>BamHI methylase has been purified to apparent homogeneity.  The isolated form
of the enzyme is a single polypeptide with a molecular weight of 56,000 as
determined by sodium dodecyl sulfate-polyacrylamide electrophoresis.  Unlike
BamHI endonuclease, which is isolated as a dimer and higher aggregates, the
methylase has no apparent higher form.  The methylase requires
S-adenosyl-L-methionine as the methyl-group donor and is inhibited by Mg2+.
The enzyme is also inhibited by 2,3-butanedione and reagents specific for
sulfhydryl groups, such as N-ethylmaleimide, which suggests a role for arginine
and cysteine residues, respectively.  DNA efficiently protects the enzyme
against the butanedione modification while S-adeno-sylmethionine has no effect.
In contrast, S-adenosyl-methionine protects against cysteine modification
while DNA produces only small amounts of protection.  Studies on the mechanism
of methylation indicate that both strands of the recognition sequence are
modified in a single binding event.  The sequence specificity of the methylase
is relaxed upon the addition of glycerol in the reaction mixture.  In the
presence of 30% glycerol the enzyme methylates sequences that are also
recognized by BamHI endonuclease when acting under conditions of relaxed
specificity.

<>

<1>Nardone, G., Wastney, M., Hensley, P., Chirikjian, J.G.
<2>Kinetics of BamHI endonuclease cleavage of form I SV40 DNA.
<3>Fed. Proc.
<4>45
<5>1504
<6>1986
<7>BamHI endonuclease recognizes the symmetrical, duplex sequence 5'-GGATCC-3' and cleaves
between the guanines on both strands.  The kinetic mechanism of the enzyme was investigated
with form [3H]-SV40 DNA which contains a single BamHI site.  The time course data was analyzed
by numerical integration of the rate equation and a coordinated non-linear least squares
analysis in terms of the rate constants.  The kinetic model used was that postulated for the
EcoRI endonuclease.  	k1	k2	k3	k4E + 1 - EI 5 EII 5 EIII5 E + III	k1k-567 k5E + II	To simplify
the analysis it was assumed that k1 and k-1 were large with respect to k2 and that k4 were not
distinguished.  In reactions containing 12 nM DNA 2 nM endonuclease an excellent fit to the
model was obtained with k2 = 0.192 min-1 +/- 0.0019, k3 (k4) = 1.190 +/- 0.042 and k5/k-5 =
1.07 +- 0.067.  Unlike EcoRI endonuclease, the cleavage of the first strand is slower with
this substrate. This analysis is useful in substantiating the model as well as providing
self-consistent values for the rate constants.  These studies wre being extended to other DNA
substrates (supported by USPS Grant GM 27701).

<>

<1>Nardone, G., Wastney, M.E., Hensley, P.
<2>DNA structural polymorphism modulates the kinetics of superhelical DNA cleavage by BamHI restriction endonucleases.
<3>J. Biol. Chem.
<4>265
<5>15308-15315
<6>1990
<7>A compartmental model developed by Hensley (Hensley, P., Nardone, G.,
Chirikjian, J.G., and Wastney, M.E., (1990) J. Biol. Chem. 265, 15300-15307)
for analysis of the time courses of the cleavage of superhelical DNA substrates
by the restriction endonuclease, BamHI, has been used to quantify the effects
of changes in temperature, ionic strength, superhelical density, and the DNA
substrate on the binding and strand cleavage processes.  Studies reported here
indicate that changes in topology may be introduced into the DNA substrate
solely as a result of the plasmid preparation process and in the absence of
covalent bond cleavage and ligation.  These changes in topology have
qualitatively different effects on the kinetics than those promoted by changes
in the superhelical density.  The former are removed by briefly warming the DNA
prior to assay, suggesting that they are only kinetically stable, while the
latter changes are not affected by heating.  Increasing the [NaCl] from 0.0 M
to 0.1 M increases the overall rate of plasmid cleavage by increasing both the
rates of cleavage and enzyme DNA association.  To describe the decrease in the
overall cleavage rate observed in 0.15 M NaCl, an ionic strength-dependent
rate-determining structural transition in the DNA substrate was incorporated
into the model.  The largest changes in the rate of the cleavage process
resulted from changes in the DNA substrate.  For the SV40 substrate compared to
pBR322, the rate constants describing the two association processes and the
first bond cleavage event were increased 6- to 7-fold.  The rate of the second
bond cleavage process was not affected.  These changes may be due to
differences in the flanking sequences.

<>

<1>Naroditsky, B.S., Khilko, S.N., Loparev, V.N.
<2>Current methods for the isolation of specific endonucleases.
<3>Vestn. Akad. Med. Nauk SSSR
<4>2
<5>26-28
<6>1981
<7>Various schemes and methods used to isolate specific endonucleases from bacteria are
discussed.  The advantages of employing affinity chromatography to purify restrictases are
shown with special reference to the isolation of a set of enzymes from B. globigii.  In
particular, the utilization, in the first stage of purification, of Sepharose 4 B with the
group-specific ligand Cibacron blue "sewed on" to it, made it possible to eliminate nucleic
acids and the bulk of protein and to obtain, already after this stage, an enzyme suitable for
physical mapping.  Additional purification of the enzyme activity maintaining fractions made
it possible to achieve mutual separation of the enzymes BglI and BglII and BglI and BglIII and
at the same time to ride of admixtures of nonspecific activities (the purification was done on
heparin Sepharose).  It is concluded that using affinity chromatography enzymes suitable for
genetic engineering work can be obtained rapidly and with a high recovery rate.

<>

<1>Narva, K.E., Skrdla, M.P., Wendell, D.L., Van Etten, J.L.
<2>Isolation of a DNA methyltransferase gene, M.CviBIII, from Chlorella virus NC-1A.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>87
<5>314
<6>1987
<7>As an initial step in the study of DNA restriction-modification systems of
viruses which infect a eucaryotic green alga, Chlorella, we have isolated the
gene for one methyltransferase, M.CviBIII, from virus NC-1A in Escherichia coli
plasmid pUC8.  M.CviBIII methylates A in TCGA sequences.  DNA from E. coli
strains harboring the recombinant plasmid, pNC-1A.14, is resistant to digestion
by SalI (GTCGAC) and TaqI (TCGA) as well as methylation by the bacterial
methylase, M.TaqI.  M.CviBIII activity in crude extracts from these E. coli
strains methylates SalI sites in lambda DNA in an in vitro protection assay.
Transposon Tn5 mutagenesis localized the M.CviBIII gene to a 1.5 kbp region on
pNC-1A.14.  Nucleic acid hybridization studies have produced several findings:
(i) Five of the 29 other Chlorella viruses described in the literature contain
a gene homologous to M.CviBIII. (ii) Spontaneous mutants of NC-1A, the DNA from
which is sensitive to TaqI and SalI, have the M.CviBIII gene deleted.  (iii)
Transcription of the M.CviBIII gene is under temporal control; a 1.4 kb mRNA
species can be detected with pNC-1A.14 probes only during the early stages of
NC-1A infection of Chlorella cells.

<>

<1>Narva, K.E., Van Etten, J.L., Slatko, B.E., Benner, J.S.
<2>The amino acid sequence of the eukaryotic DNA [N6-adenine]methyltransferase, M.CviBIII, has regions of similarity with the prokaryotic isoschizomer M.TaqI and other DNA [N6-adenine] methyltransferases.
<3>Gene
<4>74
<5>253-259
<6>1988
<7>the sequences of the genes encoding M.CviBIII (from virus NC-1A which infects a
eukaryotic alga) [Narva et al., Nucleic Acids Res. 15 (1987) 9807-9823] and
M.TaqI (from the bacterium Thermus aquaticus) [Slatko et al., Nucleic Acids
Res. 15 (1987) 9781-9796] have been determined recently.  Both enzymes
methylate adenine in the sequence TCGA.  We have compared the predicted amino
acid sequences of these two methyltransferases (MTases), with each other and
with ten other N6A-MTases and find regions of similarity.  M.CviBIII and M.TaqI
were most closely related followed by M.PaeR7, whose recognition sequence
(CTCGAG) contains the M.TaqI/M.CviBIII recognition sequence TCGA, and M.PstI,
whose recognition sequence is CTGCAG.  All of the N6-MTases contain the
sequence Asp/Asn-Pro-Pro-Tyr (B-P-P-Y) referred to by Hattman et al. [J.
Bacteriol. 164 (1985) 932-937] as region IV.  The predicted secondary structure
of this region forms a finger-like structure (Beta finger) containing a
Beta-pleated sheet (...XXXB), two Beta-turns (P-P) followed by another
Beta-pleated sheet [Y/FXXX...].

<>

<1>Narva, K.E., Wendell, D.L., Skrdla, M.P., Van Etten, J.L.
<2>Molecular cloning and characterization of the gene encoding the DNA methyltransferase, M.CviBIII, from Chlorella virus NC-1A.
<3>Nucleic Acids Res.
<4>15
<5>9807-9823
<6>1987
<7>The gene encoding the DNA methyltransferase, M.CviBIII, from Chlorella virus NC-1A was cloned
and expressed in E. coli plasmid pUC8. Plasmid (pNC-1A.14.8) encoded M.CviBIII methylates
adenine in TCGA sequences both in vivo in E. coli and in vitro. Transposon Tn5 mutagenesis
localized the M.CviBIII functional domain to a 1.5 kbp region of pNC-1A.14.8 and also
indicated that a virus promoter directs transcription of the gene in E. coli. The 2.1 kbp
insert containing the M.CviBIII gene was sequenced and a single open reading frame of 1131 bp
was identified within the domain determined by Tn5 mutagenesis. When the M.CviBIII gene was
fused in-frame with the 19 amino-terminal codons of lacZ a 45 kD polypeptide was identified in
maxicells as predicted by the DNA sequence. The M.CviBIII gene was not essential for virus
replication since a virus M.CviBIII deletion mutant also replicated in Chlorella.

<>

<1>Nascimento, A.L. et al.
<2>Comparative Genomics of Two Leptospira interrogans Serovars Reveals Novel Insights into Physiology and Pathogenesis.
<3>J. Bacteriol.
<4>186
<5>2164-2172
<6>2004
<7>Leptospira species colonize a significant proportion of rodent populations
worldwide and produce life-threatening infections in accidental hosts,
including humans. Complete genome sequencing of Leptospira interrogans
serovar Copenhageni and comparative analysis with the available Leptospira
interrogans serovar Lai genome reveal that despite overall genetic
similarity there are significant structural differences, including a large
chromosomal inversion and extensive variation in the number and
distribution of insertion sequence elements. Genome sequence analysis
elucidates many of the novel aspects of leptospiral physiology relating to
energy metabolism, oxygen tolerance, two-component signal transduction
systems, and mechanisms of pathogenesis. A broad array of transcriptional
regulation proteins and two new families of afimbrial adhesins which
contribute to host tissue colonization in the early steps of infection
were identified. Differences in genes involved in the biosynthesis of
lipopolysaccharide O side chains between the Copenhageni and Lai serovars
were identified, offering an important starting point for the elucidation
of the organism's complex polysaccharide surface antigens. Differences in
adhesins and in lipopolysaccharide might be associated with the adaptation
of serovars Copenhageni and Lai to different animal hosts. Hundreds of
genes encoding surface-exposed lipoproteins and transmembrane outer
membrane proteins were identified as candidates for development of
vaccines for the prevention of leptospirosis.

<>

<1>Nash, J.H., Villegas, A., Kropinski, A.M., Aguilar-Valenzuela, R., Konczy, P., Mascarenhas, M., Ziebell, K., Torres, A.G., Karmali, M.A., Coombes, B.K.
<2>Genome sequence of adherent-invasive Escherichia coli and comparative genomic analysis with other E. coli pathotypes.
<3>BMC Genomics
<4>11
<5>667
<6>2010
<7>ABSTRACT: BACKGROUND: Adherent and invasive Escherichia coli (AIEC) are
commonly found in ileal lesions of Crohn's Disease (CD) patients, where
they adhere to intestinal epithelial cells and invade into and survive in
epithelial cells and macrophages, thereby gaining access to a typically
restricted host niche. Colonization leads to strong inflammatory responses
in the gut suggesting that AIEC could play a role in CD immunopathology.
Despite extensive investigation, the genetic determinants accounting for
the AIEC phenotype remain poorly defined. To address this, we present the
complete genome sequence of an AIEC, revealing the genetic blueprint for
this disease-associated E. coli pathotype. Results - We sequenced the
complete genome of E. coli NRG857c (O83:H1), a clinical isolate of AIEC
from the ileum of a Crohn's Disease patient. Our sequence data confirmed a
phylogenetic linkage between AIEC and extraintestinal pathogenic E. coli
causing urinary tract infections and neonatal meningitis. The comparison
of the NRG857c AIEC genome with other pathogenic and commensal E. coli
allowed for the identification of unique genetic features of the AIEC
pathotype, including 41 genomic islands, and unique genes that are found
only in strains exhibiting the adherent and invasive phenotype.
CONCLUSIONS: Up to now, the virulence-like features associated with AIEC
are detectable only phenotypically. AIEC genome sequence data will
facilitate the identification of genetic determinants implicated in
invasion and intracellular growth, as well as enable functional genomic
studies of AIEC gene expression during health and disease.

<>

<1>Nasri, M., Sayadi, S., Thomas, D.
<2>Relaxation of PvuII recognition sequence.
<3>FEBS Lett.
<4>185
<5>101-104
<6>1985
<7>The substrate specificity of PvuII endonuclease is relaxed in the presence of
dimethyl sulfoxide.  The new recognition sequences cleaved in pBR322 DNA have
been found to be CCGCTG, CATCTG, CAGATG, CAGGTG and CAGCGG.

<>

<1>Nasri, M., Thomas, D.
<2>Alteration of the specificity of PvuII restriction endonuclease.
<3>Nucleic Acids Res.
<4>15
<5>7677-7687
<6>1987
<7>The restriction endonuclease PvuII which cleaves the sequence CAG^CTG, at the
position indicated by the arrow, was found to decrease its substrate
specificity in the presence of organic solvents.  Thirty-three sites, that we
have named PvuII* sites, were identified on the nucleotide sequence of pBR322
DNA.  The new recognition sequences cleaved in pBR322 DNA, at the positions
indicated by the arrows, were shown to be AAGCTG, GAGCTG, CNGCTG, CANCTG,
CAGNTG, CAGCNG, CAGCTC and CAGCTT.  (TAGCTG and the complementary sequence
CAGCTA are not present in pBR322 DNA).  From these recognition sequences, we
deduced that PvuII* activity recognizes and cleaves degenerate sequences which
differ from the standard PvuII sequence CAGCTG at only one of the recognition
site.  Any substitution can occur at any one of the six positions in the
hexanucleotide sequence.  The optimum incubation medium for PvuII* activity was
found to be:  10-50 mM Tris-HCl, pH 8.5, 12-15 mM MgCl2, 50 mM NaCl, 10%
ethanol + 10% dimethylsulfoxide (DMSO).

<>

<1>Nasri, M., Thomas, D.
<2>Increase of the potentialities of restriction endonucleases by specificity relaxation in the presence of organic solvents.
<3>Ann. NY Acad. Sci.
<4>542
<5>255-265
<6>1988
<7>None

<>

<1>Nasri, M., Thomas, D.
<2>Relaxation of recognition sequence of specific endonuclease HindIII.
<3>Nucleic Acids Res.
<4>14
<5>811-821
<6>1986
<7>Under the standard reaction conditions, the restriction endonuclease HindIII
cleaves double-stranded DNA, within the recognition sequence -A^AGCTT- at the
position indicated by the arrow.  In the presence of Dimethyl sulfoxide the
substrate specificity of this enzyme is reduced and cleavages occur at
additional sites.  We have determined the secondary sites in pBR322 DNA
recognized by HindIII endonuclease under relaxed conditions and found that it
cleaves the hexanucleotides:  G^AGCTT, A^GGCTT, A^TGCTT, A^ATCTT, A^AGCAT,
A^AGCGT, A^AGCTC,  at the positions indicated by the arrows, producing
fragments with cohesive ends.

<>

<1>Nasri, M., Thomas, D.
<2>The reactions of SacI, PvuII and EcoRI endonucleases.
<3>Biomed. Biochim. Acta
<4>45
<5>997-1005
<6>1986
<7>A new approach to the mechanism study of DNA restriction is suggested.  It is
based on enzymatic hydrolysis of DNA in the presence of organic solvent.  We
found that in the presence of DMSO (10% v/v), the superhelical cleavage of
pBR322 DNA by restriction endonucleases SacI and EcoRI proceeds with extensive
accumulation of an intermediate, the single-nicked circular DNA, while with
PvuII endonuclease the superhelical form is converted directly to the linear
form.

<>

<1>Nasri, M., Thomas, D.
<2>Immobilization of the restriction endonucleases PvuII and HindIII.
<3>Appl. Biochem. Biotechnol.
<4>15
<5>119-130
<6>1987
<7>The effects of several chemical reagents on the activity of the restriction
endonucleases PvuII and HindIII were investigated.  Carbodiimide, which reacts
preferentially with carboxyl groups, was found to inactivate these enzymes.
This specific effect could be prevented by Mg2+ cation.  pBR322 DNA, which
contains PvuII and PvuII sites and HindIII and HindIII sites, did not protect
the enzymes from the carbodiimide.  On the other hand, glutaraldehyde, which
reacts primarily with lysine residues, inactivates PvuII and HindIII enzymes.
This specific effect could not be prevented by pBR322 DNA.  Preincubation with
high concentrations of N-ethylmaleimide, which reacts with sulfhydryl groups,
caused slight inhibition of PvuII activity, but had no effect on the activity
of HindIII enzyme.  The effects of glutaraldehyde , carbodiimide, and
N-ethylmaleimide on other restriction endonucleases were also investigated.
Restriction endonucleases PvuII and HindIII were immobilized by covalent
coupling to various insoluble carriers.  Both immobilized enzymes retained
partial enzyme activities, when immobilized through phenolic groups and were
stable for at least two months.

<>

<1>Nassif, X., Tinsley, C.
<2>DNAs and proteins or peptides specific to bacteria of the species Neisseria meningitidis, processes for obtaining them and their biological uses.
<3>US Patent Office
<4>US 7029845 A
<5>
<6>2006
<7>The DNA of the invention are characterised in that they concern the whole or part of genes,
with their reading frame, to be found in Neisseria meningitidis, but not in Neisseria
gonorrhoeae, or in Neisseria lactamica except the genes involved in the biosynthesis of the
polysaccharide capsule, frp A, frpC, opc, por A, rotamase the sequence IC1106, IgA protease,
pilline, pilC, transferrin binding proteins and opacity proteins.  The invention also concerns
the polypeptides corresponding to these DNA and the antibodies directed against these
polypeptides.  It is applicable in the prevention and the detection of meningococcus induced
infections and meningitis.

<>

<1>Nassif, X., Tinsley, C.
<2>DNA and proteins or peptides specific of bacteria of the Neisseria meningitidis species, methods for obtaining them and biological applications thereof.
<3>US Patent Office
<4>US 7368556 A
<5>
<6>2008
<7>The DNA of the invention are characterised in that they concern the whole or part of genes,
with their reading frame, to be found in Neisseria meningitidis, but not in Neisseria
gonorrhoeae, or in Neisseria lactamica except the genes involved in the biosynthesis of the
polysaccharide capsule, frp A, frp C, opc, por A, rotamase the sequence IC1106, IgA protease,
pilline, pilC, transferrin binding proteins and opacity proteins.  The invention also concerns
the polypeptides corresponding to these DNA and the antibodies directed against these
polypeptides.  It is applicable in the prevention and the detection of meningococcus induced
infections and meningitis.

<>

<1>Nassif, X., Tinsley, C., Achtman, M., Ruelle, J.L., Vinals, C., Merker, P.
<2>DNA and specific proteins or peptides of the Neisseria meningitidis species bacteria, method for obtaining them and their biological applications.
<3>Japanese Patent Office
<4>JP 2001504684 A
<5>
<6>2001
<7>
<>

<1>Nassif, X., Tinsley, C., Aujame, L., Perrin, A., Rokbi, B., Bouchardon, A., Renauld, M.G.
<2>
<3>French Patent Office
<4>FR 2785293 A
<5>
<6>2000
<7>
<>

<1>Nastri, H.G., Evans, P.D., Walker, I.H., Riggs, P.D.
<2>Catalytic and DNA binding properties of PvuII restriction endonuclease mutants.
<3>J. Biol. Chem.
<4>272
<5>25761-25767
<6>1997
<7>The role of particular residues of the PvuII endonuclease in DNA binding and cleavage was
studied by mutational analysis using a number of in vivo and in vitro approaches.  While
confirming the importance of residues predicted to be involved directly in function by the
crystal structure, the analysis led to several striking results. Aspartate 34, which contacts
the central base pair of the PvuII site (5'-CAGCTG-3') through the minor groove, plays a
critical role in binding specificity.  A D34G mutant binds with high affinity to any of the
sequences in the set CANNTG, although its low level of cleavage activity acts only on the
wild-type site.  In addition, a His to Ala mutation at the residue that contacts the central
G, and is predicted to be blocked by PvuII methylation, still requires the PvuII methylase to
be maintained in vivo, arguing against this hypothesis as the only mechanism for methylation
protection.  Finally, four of the five mutations that reduce cleavage activity while still
exhibiting binding in the gel shift assay are at residues that form DNA- or subunit-subunit
contacts rather than in the catalytic center.  This provides further evidence for a strong
linkage between specific binding and catalysis.

<>

<1>Nath, K.
<2>Effect of sulfhydryl group inhibitors on restriction endonuclease activities.
<3>Arch. Biochem. Biophys.
<4>212
<5>611-617
<6>1981
<7>The activity of restriction endonuclease BamHI was abolished by
p-mercuribenzoate and 5,5'-dithiobis(2-nitrobenzoic acid).  The activity of
restriction endonuclease PvuI was abolished by p-mercuribenzoate.  The activity
of none of the eight other restriction endonucleases tested could be abolished
by the sulfydryl group inhibitors.  Despite the general practice of inclusion
of sulfhydryl reducing agents in reaction mixtures containing restriction
endonucleases it appears that most of these enzymes function without the active
participation of a -SH moiety.

<>

<1>Nath, K., Azzolina, B.A.
<2>Cleavage properties of site-specific restriction endonucleases.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>113-130
<6>1981
<7>*
  I. Introduction
 II. Site preference by restriction endonucleases
III. Perturbations in restriction-endonuclease cleavage properties
 IV. Application of manipulated restriction endonuclease cleavage
  V. Summary


<>

<1>Nath, K., Bollon, A.P.
<2>Generation of discrete yeast DNA fragments by endonuclease RI.
<3>Nature
<4>257
<5>155-157
<6>1975
<7>Fine structure genetic analysis has shown that the ilv1 gene of yeast,
Saccharomyces cerevisiae, is multifunctional.  The ilv1 gene product, threonine
deaminase, shows catalytic activity, as well as participating in multivalent
repression of other enzymes involved in isoleucine-valine biosynthetic
pathways.  A better understanding of the regulatory role of the ilv1 gene
product and other proteins, such as isoleucyl-tRNA synthetase, which in
addition to the ilv1 gene product seems to regulate ilv2 and ilv3 gene
expression, could be obtained if a pure preparation of the various ilv genes
were isolated in large quantities.  One way of isolating the genes would be to
use restriction enzymes to cleave yeast DNA which contains normal ilv genes and
join the fragments to a bacterial plasmid.  Because of the similarity of the
isoleucine-valine pathways in yeast and Escherichia coli, the fused DNA could
then be transformed into a strain of E. coli with a deletion in the particular
ilv gene.  Restriction enzyme-cleaved DNA from various sources has been
amplified by growth in E. coli.  We have found that restriction enzyme EcoRI
treatment of yeast DNA results in not only a reduction in the size of total DNA
but also the generation of several distinct species of homogeneous size DNAs
some of which are derived from ribosomal genes and others from mitochondrial
genes.

<>

<1>Nathan, D., Crothers, D.M.
<2>Bending and flexibility of methylated and unmethylated EcoRI DNA.
<3>J. Mol. Biol.
<4>316
<5>7-17
<6>2002
<7>We used cyclization kinetics experiments and Monte Carlo simulations to determine a structural
model for a DNA decamer containing the EcoRI restriction site. Our findings agree well with
recent crystal and NMR structures of the EcoRI dodecamer, where an overall bend of seven
degrees is distributed symmetrically over the molecule. Monte Carlo simulations indicate that
the sequence has a higher flexibility, assumed to be isotropic, compared to that of a
"generic" DNA sequence. This model was used as a starting point for the investigation of the
effect of cytosine methylation on DNA bending and flexibility. While methylation did not
affect bend magnitude or direction, it resulted in a reduction in bending flexibility and
under-winding of the methylated nucleotides. We demonstrate that our approach can augment the
understanding of DNA structure and dynamics by adding information about the global structure
and flexibility of the sequence. We also show that cyclization kinetics can be used to study
the properties of modified nucleotides.

<>

<1>Nathan, P.D., Brooks, J.E.
<2>Characterization of clones of the BamHI methyltransferase gene.
<3>Gene
<4>74
<5>35-36
<6>1988
<7>Meeting Abstract

<>

<1>Nathan, P.D., Brooks, J.E.
<2>Characterization of clones of the BamHI methyltransferase gene.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>212
<6>1989
<7>The BamHI type II restriction-modification system from Bacillus
amyloliquefaciensH, recognizes the sequence 5'-GGATCC.  We are characterizing
different subclones of the BamHI methyltransferase (MTase) gene.  One clone
carries a 2.2-kb HindIII fragment (pBamM2.2) that contains the MTase gene and
the N-terminal end of the endonuclease gene.  A second clone carries a 1.8-kb
XmnI-HindIII fragment (pBamM1.8) that contains the MTase gene and a portion of
the ORF located in the intergenic region.  These two clones differ in their
compatibility with McrB (modified cytosine restriction).  Only the plasmid
pBamM1.8 is restricted by McrB+ hosts.  Two approaches are being used to
investigate the different McrB compatibilites.  First, we have found that cells
containing pBamM1.8 produce significantly greater amounts of BamHI MTase than
cells containing pBamM2.2.  Second, we have found that a disruption of plasmid
pBamM2.2 in the intergenic ORF results in the loss of McrB compatibility.
Together, these results suggest that the higher level of MTase from pBamM1.8 is
responsible for the McrB restriction, and that the region located between the
two genes affects the level of MTase produced.

<>

<1>Nathan, P.D., Brooks, J.E.
<2>Regulation of the BamHI restriction-modification system.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>89
<5>180
<6>1989
<7>The BamHI restriction-modification (RM) system recognizes the sequence
5'-GGATCC.  The endonuclease cleaves to leave a 5' GATC extension.  The
methyltransferase (MTase) modifies the internal cytosine residue to give
GGATmCC.  We have investigated clones of the BamHI MTase gene, pBamM1.8 and
pBamM2.2, that differ in their restriction by the McrB (modified cytosine
restriction) system of Escherichia coli, only pBamM1.8 is restricted by McrB+
cells.  We have found that cells containing pBamM1.8 produce more MTase than
cells containing pBamM2.2.  We suggest that the higher expression of MTase by
cells containing pBamM1.8 is responsible for the McrB restriction.  By using
linkers to disrupt the plasmid pBamM2.2, we have identified and subcloned a
genetic region involved in the McrB effect.  This region contains a small open
reading frame (ORF) that lies between the BamHI RM genes.  Recently we have
investigated the effect of the ORF region on the endonuclease gene.  Cells that
contain a plasmid with the BamHI RM system disrupted in the ORF region produce
a lower level of endonuclease.  Moreover, endonuclease levels are restored when
the cells are cotransformed with the ORF containing plasmid.  These results
syggest that the region located between the two genes is important in
controlling the expression of both BamHI MTase and endonuclease.

<>

<1>Nathans, D., Adler, S.P., Brockman, W.W., Danna, K.J., Lee, T.N.H., Sack, G.H. Jr.
<2>Use of restriction endonucleases in analyzing the genome of simian virus 40.
<3>Fed. Proc.
<4>33
<5>1135-1138
<6>1974
<7>Bacterial restriction endonucleases have been used to cleave SV40 DNA at
specific sites.  The resulting fragments have been ordered in the molecule,
resulting in a physical cleavage map of the SV40 chromosome.  With this map as
a reference, sites of initiation and termination of DNA replication have been
located, as have regions of the genome expressed early and late in productively
infected cells.  The 5' - 3' orientation of each strand of SV40 DNA has also
been determined; from this information and prior identification of the early
and late template strands, the direction of early and late transcription could
be deduced.

<>

<1>Nathans, D., Smith, H.O.
<2>Restriction endonucleases in the analysis and restructuring of DNA molecules.
<3>Annu. Rev. Biochem.
<4>44
<5>273-293
<6>1975
<7>Restriction enzymes are endodeoxyribonucleases that recognize specific
nucleotide sequences in double stranded DNA and cleave both strands of the
duplex.  In the cell of origin each restriction enzyme is part of a
restriction-modification (R-M) system, consisting of the restriction
endonuclease and a matched modification enzyme which recognizes and modifies
(generally by methylation) the same nucleotide sequence in DNA recognized by
the restriction enzyme.  Modification thus protects cellular DNA from
restriction; however, foreign (unmodified) DNA is cleaved by the restriction
endonuclease and further degraded by other enzymes.  Such R-M systems, first
detected by phage restriction and modification, are widespread in bacteria and
are thought to play a role in eliminating foreign DNA that gains entrance to
the cell via viruses or as naked DNA.  The biochemistry and genetics of R-M
systems have recently been reviewed.  The usefulness of restriction
endonucleases in the analysis and restructuring of DNA, which is the topic of
this review, rests on the fact that some of the restriction enzymes cleave DNA
at specific nucleotide sequences.  These cleavage site-specific endonucleases
are thus analogous to specific proteolytic enzymes and are proving as useful in
the study of DNA structure and function as trypsin and chymotrypsin have been
in protein analysis.  After the first characterization of a restriction
endonuclease from Escherichia coli strain K in 1968, there was a curious lag in
the application of restriction enzymes as analytical tools.  In a sense this
delay was fortunate, since the enzymes isolated initially are in the class now
known to be nonspecific in their cleavage sites.  However, after the discovery
of cleavage site-specific endonucleases, there was immediate applicaton of
these enzymes to the analysis of viral genomes.  In the past three years there
has been an almost explosive rate of discovery of new site-specific restriction
enzymes and rapid application to physical mapping of chromosomes, nucleotide
sequence analysis of DNA, isolation of genes, and restructuring of DNA
molecules.  Our purpose is to review these recent developments.

<>

<1>Naumann, T.A., Tavassoli, A., Benkovic, S.J.
<2>Genetic selection of cyclic peptide dam methyltransferase inhibitors.
<3>Chembiochem
<4>9
<5>194-197
<6>2008
<7>Enzymatic methylation of specific DNA bases is fundamental to the survival and propagation of
a variety of organisms, including humans.  DNA methyltransferases control and regulate a
variety of cellular processes, and due to the central role these processes play in the
organism's lifecycle, bacterial methyltransferases are attractive targets for the development
of new antibiotics.  The Escherichia coli dam methyltransferase protein is an N-6 adenine
methyltransferase that methylates the GATC palindrome by transfer of a methyl group from
S-adenosyl-L-methionine.  EcoDam functions in a number of diverse and important cellular
processes, the most well studied being postreplicative DNA mismatch repair and control of DNA
replication.  In uropathogenic E. coli, EcoDam activity is required for conversion to and
maintenance of the virulent phenotype.

<>

<1>Naumann, U., Daxinger, L., Kanno, T., Eun, C., Long, Q.A., Lorkovic, Z.J., Matzke, M., Matzke, A.J.M.
<2>Genetic Evidence That DNA Methyltransferase DRM2 Has a Direct Catalytic Role in RNA-Directed DNA Methylation in Arabidopsis thaliana.
<3>Genetics
<4>187
<5>977-979
<6>2011
<7>RNA-directed DNA methylation (RdDM) is a small RNA-mediated epigenetic modification in plants.
We report here the identification of DOMAINS
REARRANGED METHYLTRANSFERASE 2 (DRM2) in a forward screen for mutants
defective in RdDM in Arabidopsis thaliana. The finding of a mutation in
the presumptive active site argues in favor of direct catalytic
activity for DRM2.

<>

<1>Navas, E., Bohle, H., Henriquez, P., Grothusen, H., Bustamante, F., Bustos, P., Mancilla, M.
<2>Draft Genome Sequence of the Fish Pathogen Yersinia ruckeri Strain 37551, Serotype O1b, Isolated from Diseased, Vaccinated Atlantic Salmon (Salmo salar) in  Chile.
<3>Genome Announcements
<4>2
<5>e00858-14
<6>2014
<7>We sequenced the genome of a motile O1b Yersinia ruckeri field isolate from Chile, which is
causing enteric redmouth disease (ERM) in vaccinated Atlantic
salmon (Salmo salar). The draft genome has 3,775,486 bp, a G+C content of 47.1%,
and is predicted to contain 3,406 coding sequences.

<>

<1>Naz, S., Tareb, R., Bernardeau, M., Vaisse, M., Lucchetti-Miganeh, C., Rechenmann, M., Vernoux, J.P.
<2>Genome Sequence of Lactobacillus plantarum Strain UCMA 3037.
<3>Genome Announcements
<4>1
<5>e00251-13
<6>2013
<7>Nucleic acid of the strain Lactobacillus plantarum UCMA 3037, isolated from raw milk camembert
cheese in our laboratory, was sequenced. We present its draft
genome sequence with the aim of studying its functional properties and
relationship to the cheese ecosystem.

<>

<1>Nazarenko, I.A., Gorbunov, Y.A., Malysgin, E.G.
<2>Cleavage of synthetic RNA-DNA hybrids with restriction endonucleases BamHI and Sau3AI.
<3>Bioorg. Khim.
<4>13
<5>928-933
<6>1987
<7>Synthetic oligodeoxyribonucleotides, containing one or two ribonucleotides in
the recognition sequence, and RNA-DNA hybrids were tested for their activity in
cleavage with BamHI and Sau3AI endonucleases.  The replacement of dG with G in
the first position of BamHI-site (GGATCC) of one of the chains does not affect
the rate of the BamHI hydrolysis.  The similar heteroduplex, containing G
residue in the second position, displays a decreased rate of the BamHI
hydrolysis of the modified strand and to a lesser extent, of the unmodified
complementary strand.  Oligodeoxyribonucleotides in complex with
oligoribonucleotides can be cleaved with the excess of BamHI and Sau3AI
oligoribonucleotides remaining intact.

<>

<1>Nazir, R., Hansen, M.A., Sorensen, S., van Elsas, J.D.
<2>Draft Genome Sequence of the Soil Bacterium Burkholderia terrae Strain BS001, Which Interacts with Fungal Surface Structures.
<3>J. Bacteriol.
<4>194
<5>4480-4481
<6>2012
<7>Burkholderia terrae BS001 is a soil bacterium which was originally isolated from  the
mycosphere of the ectomycorrhizal fungus Laccaria proxima. It exhibits a
range of fungus-interacting traits which reveal its propensity to actively
interact at fungal interfaces. Here, we present the approximately 11.5-Mb (G+C
content, 61.52%) draft genome sequence of B. terrae BS001 with the aim of
providing insight into the genomic basis of its ecological success in
fungus-affected soil settings.

<>

<1>Neave, M.J., Michell, C.T., Apprill, A., Voolstra, C.R.
<2>Whole-genome sequences of three symbiotic endozoicomonas strains.
<3>Genome Announcements
<4>2
<5>e00802-14
<6>2014
<7>Members of the genus Endozoicomonas associate with a wide range of marine organisms. Here, we
report on the whole-genome sequencing, assembly, and
annotation of three Endozoicomonas type strains. These data will assist in
exploring interactions between Endozoicomonas organisms and their hosts, and it
will aid in the assembly of genomes from uncultivated Endozoicomonas spp.

<>

<1>Neaves, K.J., Cooper, L.P., White, J.H., Carnally, S.M., Dryden, D.T., Edwardson, J.M., Henderson, R.M.
<2>Atomic force microscopy of the EcoKI Type I DNA restriction enzyme bound to DNA shows enzyme dimerization and DNA looping.
<3>Nucleic Acids Res.
<4>37
<5>2053-2063
<6>2009
<7>Atomic force microscopy (AFM) allows the study of single protein-DNA interactions such as
those observed with the Type I
Restriction-Modification systems. The mechanisms employed by these systems
are complicated and understanding them has proved problematic. It has been
known for years that these enzymes translocate DNA during the restriction
reaction, but more recent AFM work suggested that the archetypal EcoKI
protein went through an additional dimerization stage before the onset of
translocation. The results presented here extend earlier findings
confirming the dimerization. Dimerization is particularly common if the
DNA molecule contains two EcoKI recognition sites. DNA loops with dimers
at their apex form if the DNA is sufficiently long, and also form in the
presence of ATPgammaS, a non-hydrolysable analogue of the ATP required for
translocation, indicating that the looping is on the reaction pathway of
the enzyme. Visualization of specific DNA loops in the protein-DNA
constructs was achieved by improved sample preparation and analysis
techniques. The reported dimerization and looping mechanism is unlikely to
be exclusive to EcoKI, and offers greater insight into the detailed
functioning of this and other higher order assemblies of proteins
operating by bringing distant sites on DNA into close proximity via DNA
looping.

<>

<1>Nebendahl, A., Baumlein, H.
<2>Analysis of overlapping cDNA clones specific for a putative second DNA methyltransferase-encoding gene in Arabidopsis thaliana.
<3>Gene
<4>157
<5>269-272
<6>1995
<7>We have isolated and sequenced overlapping cDNA clones specific for a putative second DNA
methyltransferase (MTase)-encoding gene (MTase11) from Arabidopsis thaliana (At) recently
described as a genomic DNA fragment.  The gene seems to be present as a single copy in the At
genome and is transcribed into a 2.2-kb messenger detectable only in young seedlings.  Using
sequence comparison we found structural differences between the cDNA clones and the previously
reported genomic fragment.  The amino-acid sequence deduced from the 1.8-kb cDNA sequence
shows the occurrence of the conserved motif number VI characteristic for m5C-MTases.  Northern
and Southern analysis detects no cross-hybridization with the originally described
MTase-encoding At gene (MTase1).

<>

<1>Nechaeva, A.A., Kalinina, N.A., Sukhodolets, V.V.
<2>Plasmid profiles and stability of industrial strains of lactococci.
<3>Genetika
<4>31
<5>1210-1217
<6>1995
<7>Industrial lactococci strains, usually cultivated on agar with hydrolized milk, are
characterized by an increased instability with respect to their ability to utilize lactose on
an enriched M21 medium.  Plasmid profiles of 14 industrial strains of Lactococcus lactis subs.
lactis and of a large number of segregants phenotypically differing in sugar-fermenting
ability and phage sensitivity were examined.  Four out of 14 strains segregated Lac-Gal-
clones, i.e., simultaneously lost the ability to utilize lactose and galactose.  All
phage-sensitive segregants retained systems of DNA restriction and modification.  Segregants
sensitive to the phage lethal effect were found.  These segregants possibly retained the
defense mechanism against phages manifested as an abortive infection.  In the majority of lac-
segregants, large plasmids of 40 to 55 kb were lost, although changes in plasmid profiles of
many Lac- segregants were not detected.

<>

<1>Needels, M.C., Fried, S.R., Love, R., Rosenberg, J.M., Boyer, H.W., Greene, P.J.
<2>Determinants of EcoRI endonuclease sequence discrimination.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>86
<5>3579-3583
<6>1989
<7>The arginine at position 200 of EcoRI endonuclease is thought to make two
hydrogen bonds to the guanine of the sequence GAATTC and thus be an important
determinant of sequence discrimination.  Arg-200 was replaced by each of the
other 29 naturally occurring amino acids, and the mutant endonucleases were
assessed for activities in vivo and in vitro.  The mutant endonuclease with
lysine at position 200 exhibits the most in vivo activity of all the position
200 mutants, although the in vitro activity is less than 1/100th of wild-type
activity.  Five other mutants show more drastically reduced levels of in vivo
activity (Cys, Pro, Val, Ser, and Trp).  The Cys, Val, and Ser mutant enzymes
appear to have in vivo activity which is specific for the wild-type canonical
site despite the loss of hydrogen bonding potential at position 200.  The Pro
and Trp mutants retain in vivo activity which is independent of the presence of
the EcoRI methylase.  In crude cell lysates, only the Cys mutant shows a very
low level of in vitro activity.  None of the mutant enzymes show a preference
for alternative sites in assays in vitro.  The implications of these results
are discussed.

<>

<1>Needels, M.C., Reich, N.O., Boyer, H.W., Greene, P.
<2>Cassette mutagenesis of EcoRI endonuclease.
<3>J. Cell Biochem. Suppl.
<4>11C
<5>209
<6>1987
<7>From inspection of the x-ray crystal structure of the DNA-EcoRI endonuclease
recognition complex, regions of the protein which are responsible for binding
specificity and catalysis were inferred.  Based on this structural information,
we have subjected the EcoRI endonuclease gene to site directed mutagenesis.  In
the first round of mutagenesis we used mismatched oligonucleotides to introduce
a small number of conservative substitutions into the putative recognition
regions.  In order to extend this analysis, we have now engineered several
unique restriction sites into the EcoRI endonuclease gene bracketing
potentially important regions.  For example, we have introduced sites which
bracket the N-terminal residues of each of the recognition helices.  Also, the
introduction of flanking BstEII and SpeI sites around residue 130 has allowed
us to test a model for the mechanism of phosphodiester hydrolysis.  Based on
the crystal structure of the protein-DNA complex, it is possible that Lys 130
participates in general acid catalysis of this reaction.  Employing cassette
mutagenesis, we have generated mutants of the EcoRI endonuclease gene which
encode a variety of different amino acids at position 130.  The mutant gene
products are being characterized and the effect on catalysis of different
substitutions at position 130 is being assessed.

<>

<1>Neely, L., Hackett, M.
<2>Methods for analyzing nucleic acids by binding of detectably labeled type II restriction endonucleases.
<3>International Patent Office
<4>WO 200512575 A
<5>39
<6>2005
<7>The invention relates to methods, products and systems for analyzing nucleic acid molecules
using a nucleic acid binding protein such as a sequence-specific endonuclease.  The methods
can be used to obtain sequence information about the nucleic acid molecules.

<>

<1>Neely, L., Hackett, M.
<2>Methods and compositions related to the use of sequence-specific endonucleases for analyzing nucleic acids under non-cleaving conditions.
<3>US Patent Office
<4>US 20050123944 A
<5>
<6>2005
<7>The invention relates to methods, products and systems for analyzing nucleic acid molecules
using a nucleic acid binding protein such as a sequence-specific endonuclease.  The methods
can be used to obtain sequence information about the nucleic acid molecules.

<>

<1>Neely, R.K., Daujotyte, D., Grazulis, S., Magennis, S.W., Dryden, D.T.F., Klimasauskas, S., Jones, A.C.
<2>Time-resolved fluorescence of 2-aminopurine as a probe of base flipping in M.HhaI-DNA complexes.
<3>Nucleic Acids Res.
<4>33
<5>6953-6960
<6>2005
<7>DNA base flipping is an important mechanism in molecular enzymology, but its study is limited
by the lack of an accessible and reliable diagnostic technique. A series of crystalline
complexes of a DNA methyltransferase, M.HhaI, and its cognate DNA, in which a fluorescent
nucleobase analogue, 2-aminopurine (AP), occupies defined positions with respect the target
flipped base, have been prepared and their structures determined at higher than 2 A
resolution. From time-resolved fluorescence measurements of these single crystals, we have
established that the fluorescence decay function of AP shows a pronounced, characteristic
response to base flipping: the loss of the very short (approximately 100 ps) decay component
and the large increase in the amplitude of the long (approximately 10 ns) component. When AP
is positioned at sites other than the target site, this response is not seen. Most
significantly, we have shown that the same clear response is apparent when M.HhaI complexes
with DNA in solution, giving an unambiguous signal of base flipping. Analysis of the AP
fluorescence decay function reveals conformational heterogeneity in the DNA-enzyme complexes
that cannot be discerned from the present X-ray structures.

<>

<1>Neely, R.K., Roberts, R.J.
<2>The BsaHI Restriction-Modification System: Cloning, Sequencing and Analysis of Conserved Motifs.
<3>BMC Mol. Biol.
<4>9
<5>48
<6>2008
<7>ABSTRACT: BACKGROUND: Restriction and modification enzymes typically recognise short DNA
sequences of between two and eight bases in length.
Understanding the mechanism of this recognition represents a significant
challenge that we begin to address for the BsaHI restriction-modification
system, which recognises the six base sequence GRCGYC. RESULTS: The DNA
sequences of the genes for the BsaHI methyltransferase, bsaHIM, and
restriction endonuclease, bsaHIR, have been determined (GenBank accession
#EU386360), cloned and expressed in E.coli. Both the restriction
endonuclease and methyltransferase enzymes share significant homology with
a group of 6 other enzymes comprising the RM systems HgiDI and HgiGI and
the putative HindVP, NlaCORFDP, NpuORFC228P and SplZORFNP RM systems. A
sequence alignment of these homologues shows that their amino acid
sequences are largely conserved and highlights several motifs of interest.
One such conserved motif was identified at the C-terminal end of the
bsaHIR gene as a potential DNA-recognising region of the enzyme. A
mutational analysis of these amino acids is consistent with this
assignment. Sequence alignment of the methyltransferase gene reveals a
motif that may be used as a diagnostic tool to define the recognition
sequences of the cytosine C5 methyltransferases. CONCLUSIONS: We have
identified a region of the R.BsaHI enzyme that is likely involved in DNA
recognition. Analysis of the amino acid sequence of the BsaHI
methyltransferase enzyme led us to propose two new motifs that can be used
in the diagnosis of the recognition sequence of the cytosine
C5-methyltransferases.

<>

<1>Neely, R.K., Tamulaitis, G., Chen, K., Kubala, M., Siksnys, V., Jones, A.C.
<2>Time-resolved fluorescence studies of nucleotide flipping by restriction enzymes.
<3>Nucleic Acids Res.
<4>37
<5>6859-6870
<6>2009
<7>Restriction enzymes Ecl18kI, PspGI and EcoRII-C, specific for interrupted 5-bp target
sequences, flip the central base pair of these sequences into
their protein pockets to facilitate sequence recognition and adjust the
DNA cleavage pattern. We have used time-resolved fluorescence spectroscopy
of 2-aminopurine-labelled DNA in complex with each of these enzymes in
solution to explore the nucleotide flipping mechanism and to obtain a
detailed picture of the molecular environment of the extrahelical bases.
We also report the first study of the 7-bp cutter, PfoI, whose recognition
sequence (T/CCNGGA) overlaps with that of the Ecl18kI-type enzymes, and
for which the crystal structure is unknown. The time-resolved fluorescence
experiments reveal that PfoI also uses base flipping as part of its DNA
recognition mechanism and that the extrahelical bases are captured by PfoI
in binding pockets whose structures are quite different to those of the
structurally characterized enzymes Ecl18kI, PspGI and EcoRII-C. The
fluorescence decay parameters of all the enzyme-DNA complexes are
interpreted to provide insight into the mechanisms used by these four
restriction enzymes to flip and recognize bases and the relationship
between nucleotide flipping and DNA cleavage.

<>

<1>Neesen, K., Volckaert, G.
<2>Construction and shuttling of novel bifunctional vectors for Streptomyces spp. and Escherichia coli.
<3>J. Bacteriol.
<4>171
<5>1569-1573
<6>1989
<7>Shuttle vectors for gene transfer between Streptomyces spp. and Escherichia
coli have been constructed by fusion of an artificial multicopy E. coli
replicon and DNA fragments of pIJ702.  Stable transfer to Streptomyces lividans
was obtained.  Marked differences in transformation efficiency were observed
when plasmid DNA isolated from E. coli GM119 was used instead of that from
strain HB101.

<>

<1>Neff, N.F.
<2>Protein splicing: selfish genes invade cellular proteins.
<3>Curr. Opin. Cell Biol.
<4>5
<5>971-976
<6>1993
<7>Protein splicing is a series of enzymatic events involving intramolecular
protein breakage, rejoining and intron homing, in which introns are able to promote the
recombinative transposition of their own coding sequences.  Eukaryotic and prokaryotic
spliced proteins have conserved similar gene structure, but little amino acid identity.  The
genes coding for these spliced proteins contain internal in-frame introns that encode
polypeptides that apparently self-excise from the resulting host protein sequences.
Excision of the 'protein intron' is coupled with joining of the two flanking protein regions
encoded by exons of the host gene.  Some introns of this type encode DNA endonucleases,
related to Group I RNA intron gene products, that stimulate gene conversion and self-
transmission.

<>

<1>Negi, V., Lata, P., Sangwan, N., Kumar-Gupta, S., Das, S., Rao, D.L., Lal, R.
<2>Draft Genome Sequence of Hexachlorohexane (HCH)-Degrading Sphingobium lucknowense Strain F2T, Isolated from an HCH Dumpsite.
<3>Genome Announcements
<4>2
<5>e00788-14
<6>2014
<7>Sphingobium lucknowense F2(T), isolated from the hexachlorocylcohexane (HCH) dumpsite located
in Ummari village, Lucknow, India, rapidly degrades HCH isomers.
Here we report the draft genome of strain F2 (4.4 Mbp), consisting of 4,910
protein coding genes with an average G+C content of 64.3%.

<>

<1>Nejedly, K., Matyasek, R., Palecek, E.
<2>Site-specific chemical modification of B-Z junctions in supercoiled DNA as detected by nuclease S1 digestion, inhibition of restriction cleavage and nucleotide sequencing.
<3>J. Biomol. Struct. Dyn.
<4>6
<5>261-273
<6>1988
<7>Structural distortions on the boundary between right-handed and left-
handed segments in the superhelical plasmid pPK2 (a derivative of pUC19 containing (dC-
dG)n segments cloned into polylinker) were studied by means of chemical probes.  Strong
osmium tetroxide, pyridine (Os,py) modification of DNA at native superhelical density (sigma)
was found in four thymines surrounding the (dC-dG)13 segment.  These results correlated  with
restriction cleavage inhibition (due to modification): BamHI cleavage was strongly  inhibited,
unlike the neighboring XbaI and SalI (weak or no inhibition).  In the (dC-dG)8  segment
considerably weaker modification of the B-Z junctions was observed,
accompanied by weak inhibition of BamHI cleavage, while the neighboring SmaI and KpnI
were not affected.  Os,py modification of DNA at native sigma was not detected by nuclease S1
cleavage at and (dC-dG)n segment.  However, this enzyme recognized and cleaved at the  B-Z
junction, osmium modified at more negative sigma.  The results obtained with the glyoxal  and
diethyl pyrocarbonate modification support the idea of very narrow B-Z junctions at  native
sigma.

<>

<1>Nekrasov, S.V., Agafonova, O.V., Belogurova, N.G., Delver, E.P., Belogurov, A.A.
<2>Plasmid-encoded Antirestriction Protein ArdA Can Discriminate between Type I Methyltransferase and Complete Restriction-Modification System.
<3>J. Mol. Biol.
<4>365
<5>284-297
<6>2007
<7>Many promiscuous plasmids encode the antirestriction proteins ArdA (alleviation of restriction
of DNA) that specifically affect the
restriction activity of heterooligomeric type I restriction-modification
(R-M) systems in Escherichia coli cells. In addition, a lot of the
putative ardA genes encoded by plasmids and bacterial chromosomes are
found as a result of sequencing of complete genomic sequences, suggesting
that ArdA proteins and type I R-M systems that seem to be widespread among
bacteria may be involved in the regulation of gene transfer among
bacterial genomes. Here, the mechanism of antirestriction action of ArdA
encoded by IncI plasmid ColIb-P9 has been investigated in comparison with
that of well-studied T7 phage-encoded antirestriction protein Ocr using
the mutational analysis, retardation assay and His-tag affinity
chromatography. Like Ocr, ArdA protein was shown to be able to efficiently
interact with EcoKI R-M complex and affect its in vivo and in vitro
restriction activity by preventing its interaction with specific DNA.
However, unlike Ocr, ArdA protein has a low binding affinity to EcoKI
Mtase and the additional C-terminal tail region (VF-motif) is needed for
ArdA to efficiently interact with the type I R-M enzymes. It seems likely
that this ArdA feature is a basis for its ability to discriminate between
activities of EcoKI Mtase (modification) and complete R-M system
(restriction) which may interact with unmodified DNA in the cells
independently. These findings suggest that ArdA may provide a very
effective and delicate control for the restriction and modification
activities of type I systems and its ability to discriminate against DNA
restriction in favour of the specific modification of DNA may give some
advantage for efficient transmission of the ardA-encoding promiscuous
plasmids among different bacterial populations.

<>

<1>Nellemann, L.J., Aamand, J.L., Madsen, A., Josephsen, J.
<2>Characterization of a non-type II restriction/modification system on plasmid PJW565 from Lactococcus lactis subsp. cremoris W56.
<3>FASEB J.
<4>11
<5>A1035
<6>1997
<7>Lactococcus lactis subsp. cremoris W56 contains several plasmid encoded
restriction-modification systems.  The 12.826 kb plasmid pJW565 exhibits a non-type II
activity, designated LlaBII, and displays resistance against small-isometric-headed phages p2
and SK1.  The entire plasmid has been sequenced.  Sequence analysis revealed that the R/M
system was encoded by at least two genes, carrying code for the 231 amino acid protein
designated llabiiM, and the 1376 amino acid protein designated llabiiR.  Both genes were
preceded by putative promoter and RBS sequences.  The two genes were separated by 1
nucleotide, indicating polycistronic transcription.  The predicted proteins (M.LlaBII and
R.LlaBII) showed homology to several adenine-specific methyltransferases and an E. coli type I
restriction enzyme, respectively.  Apart from the putative genes involved in the R/M system
activity and the genes involved in replication, pJW565 contained several open reading frames
with no known function or homology to existing sequencing data in the EMBL or Genbank
databases.  In vitro studies of LlaBII activity revealed that the cofactor ATP was essential,
and the methyl-group donor Adomet stimulated LlaBII endonuclease activity.

<>

<1>Nelson, B.A., Ramaiya, P., Lopez-de-Leon, A., Kumar, R., Crinklaw, A., Jolkovsky, E., Crane, J.M., Bergstrom, G.C., Rey, M.W.
<2>Complete Genome Sequence for the Fusarium Head Blight Antagonist Bacillus amyloliquefaciens Strain TrigoCor 1448.
<3>Genome Announcements
<4>2
<5>e00219-14
<6>2014
<7>We present the complete genome sequence for Bacillus amyloliquefaciens TrigoCor 1448 (ATCC
202152), a bacterial biological control agent for Fusarium head blight
in wheat. We compare it to its closest relative, B. amyloliquefaciens strain
AS43.3.

<>

<1>Nelson, H.C.M., Bestor, T.H.
<2>Base eversion and shuffling by DNA methyltransferases.
<3>Chem. Biol.
<4>3
<5>419-423
<6>1996
<7>The structures of two DNA cytosine methyltransferases reveal two novel methods of gaining
access to the substrate cytosine residue, both of which involve eversion of the cytosine in a
process that may require DNA bending.  In one instance there is also widespread base shuffling
and distortion of the DNA.

<>

<1>Nelson, J.M., Miceli, S.M., Lechevalier, M.P., Roberts, R.J.
<2>FseI, a new type II restriction endonuclease that recognizes the octanucleotide sequence 5' GGCCGGCC 3' .
<3>Nucleic Acids Res.
<4>18
<5>2061-2064
<6>1990
<7>A Type II restriction endonuclease, designated FseI, has been partially purified from a
Frankia species (NRRL 18528). This enzyme cleaves Adenovirus 2 DNA at three sites, but does
not cleave the DNAs from bacteriophages lambda, T7 and PhiX174, the animal virus SV40, pUC18
and pBR322. FseI recognizes the octanucleotide sequence 5' GGCCGG^CC 3' and cleaves as
indicated by the arrow. The frequency of occurrence of FseI sites within sequenced regions of
the human genome is similar to that for NotI sites.

<>

<1>Nelson, K.E. et al.
<2>Evidence for lateral gene transfer between Archaea and bacteria from genome sequence of Thermotoga maritima.
<3>Nature
<4>399
<5>323-329
<6>1999
<7>The 1,860,725-base pair genome of Thermotoga maritima MSB8 contains 1,877 predicted coding
regions, 1,014 (54%) of which have functional assignments and 863 (46%) of which are of
unknown function. Genome analysis reveals numerous pathways involved in degradation of sugars
and plant polysaccharides, and 108 genes that have orthologues only in the genomes of other
thermophilic Eubacteria and Archaea.  Of the Eubacteria sequenced to date, T. maritima has the
highest percentage (24%) of genes that are most similar to archaeal genes.  Eighty-one
archaeal-like genes are clustered in 15 regions of the T. maritima genome that range in size
from 4 to 20 kilobases.  Conservation of gene order between T. maritima and Archaea in many of
the clustered regions suggests that lateral gene transfer may have occurred between
thermophilic Eubacteria and Archaea.

<>

<1>Nelson, K.E. et al.
<2>Complete genome sequence of the oral pathogenic bacterium Porphyromonas gingivalis strain W83.
<3>J. Bacteriol.
<4>185
<5>5591-5601
<6>2003
<7>The complete 2,343,479-bp genome sequence of the gram-negative, pathogenic oral bacterium
Porphyromonas gingivalis strain W83, a major contributor to
periodontal disease, was determined. Whole-genome comparative analysis
with other available complete genome sequences confirms the close
relationship between the Cytophaga-Flavobacteria-Bacteroides (CFB) phylum
and the green-sulfur bacteria. Within the CFB phyla, the genomes most
similar to that of P. gingivalis are those of Bacteroides thetaiotaomicron
and B. fragilis. Outside of the CFB phyla the most similar genome to P.
gingivalis is that of Chlorobium tepidum, supporting the previous
phylogenetic studies that indicated that the Chlorobia and CFB phyla are
related, albeit distantly. Genome analysis of strain W83 reveals a range
of pathways and virulence determinants that relate to the novel biology of
this oral pathogen. Among these determinants are at least six putative
hemagglutinin-like genes and 36 previously unidentified peptidases. Genome
analysis also reveals that P. gingivalis can metabolize a range of amino
acids and generate a number of metabolic end products that are toxic to
the human host or human gingival tissue and contribute to the development
of periodontal disease.

<>

<1>Nelson, K.E. et al.
<2>Whole genome comparisons of serotype 4b and 1/2a strains of the food-borne pathogen Listeria monocytogenes reveal new insights into the core genome   components of this species.
<3>Nucleic Acids Res.
<4>32
<5>2386-2395
<6>2004
<7>The genomes of three strains of Listeria monocytogenes that have been associated with
food-borne illness in the USA were subjected to whole
genome comparative analysis. A total of 51, 97 and 69 strain-specific
genes were identified in L.monocytogenes strains F2365 (serotype 4b,
cheese isolate), F6854 (serotype 1/2a, frankfurter isolate) and H7858
(serotype 4b, meat isolate), respectively. Eighty-three genes were
restricted to serotype 1/2a and 51 to serotype 4b strains. These strain-
and serotype-specific genes probably contribute to observed differences in
pathogenicity, and the ability of the organisms to survive and grow in
their respective environmental niches. The serotype 1/2a-specific genes
include an operon that encodes the rhamnose biosynthetic pathway that is
associated with teichoic acid biosynthesis, as well as operons for five
glycosyl transferases and an adenine-specific DNA methyltransferase. A
total of 8603 and 105 050 high quality single nucleotide polymorphisms
(SNPs) were found on the draft genome sequences of strain H7858 and strain
F6854, respectively, when compared with strain F2365. Whole genome
comparative analyses revealed that the L.monocytogenes genomes are
essentially syntenic, with the majority of genomic differences consisting
of phage insertions, transposable elements and SNPs.

<>

<1>Nelson, M., Burbank, D.E., Van Etten, J.L.
<2>Chlorella viruses encode multiple DNA methyltransferases.
<3>Biol. Chem.
<4>379
<5>423-428
<6>1998
<7>The >320 kb dsDNA genomes of 16 viruses which infect Chlorella strain NC64A and 5 viruses
infecting Chlorella strain Pbi were tested for their sensitivity/resistance to more than 80
DNA restriction endonucleases.  From the known methylation sensitivities of these enzymes to
site-specific 5-methylcytosine and N6-methyladenine DNA modifications, we deduce that the 16
NC64A viruses encode at least 13 different sequence-specific DNA methyltransferases and the 5
Pbi viruses encode at least 7 sequence-specific DNA methyltransferases.  Each DNA
methyltransferase has a 2 to 4 base pair DNA recognition sequence.  Some individual viruses
encode as many as ten different DNA methyltransferases, making these chlorella virus genomes
among the most concentrated sources of DNA methyltransferase genes known.

<>

<1>Nelson, M., Christ, C., Schildkraut, I.
<2>Alteration of apparent restriction endonuclease recognition specificities by DNA methylases.
<3>Nucleic Acids Res.
<4>12
<5>5165-5173
<6>1984
<7>An in vitro method of altering the apparent cleavage specificities of
restriction endonucleases was developed using DNA modification methylases.
This method was used to reduce the number of cleavage sites for 34 restriction
endonucleases.  In particular, single-site cleavages were achieved for NheI in
Adeno-2 DNA and for AccI and HincII in pBR322 DNA by specifically methylating
all but one recognition sequence.

<>

<1>Nelson, M., McClelland, M.
<2>The effect of site-specific methylation on restriction-modification enzymes.
<3>Nucleic Acids Res.
<4>15
<5>r219-r230
<6>1987
<7>Previous tabulations of restriction endonuclease sensitivities to site-specific
DNA methylation have shown that these endonucleases cannot cut particular DNA
recognition sequences which have been methylated at 4mC, 5mC or 6mA.  Since our
previous tabulation in this journal the major new additions are extensive data
on 4mC.  We have altered our notation to incorporate the 4mC data and added a
number of footnotes.  Fine structural details of cleavage reactions, rate
differences on hemi- and bi-methylated substrates, and experimental
discrepancies are noted where such data is available.

<>

<1>Nelson, M., McClelland, M.
<2>Purification and assay of Type II DNA methylases.
<3>Methods Enzymol.
<4>155
<5>32-41
<6>1987
<7>Site-specific DNA methylases (S-adenosylmethionine:  DNA methyltransferases)
have a variety of uses in DNA analytical procedures.  Methylases may be used as
probes for DNA conformation, in recombinant DNA cloning strategies, or in
combination with restriction endonucleases to generate rare or novel DNA
cleavage specificities.  DNA methylases have been purified from a number of
different sources.

<>

<1>Nelson, M., McClelland, M.
<2>Effect of site-specific methylation on DNA modification methyltransferases and restriction endonucleases.
<3>Nucleic Acids Res.
<4>17
<5>r389-r415
<6>1989
<7>Review of the effects of methylation.

<>

<1>Nelson, M., McClelland, M.
<2>Site-specific methylation: effect on DNA modification methyltransferases and restriction endonucleases.
<3>Nucleic Acids Res.
<4>19
<5>2045-2071
<6>1991
<7>We present in Table I an updated list of the sensitivities of over 240
restriction endonucleases to the site-specific DNA modifications m4C, m5C,
hm5C, and m6A, four modifications that are common in DNA prokaryotes,
eukaryotes, and their viruses (Mc2, Mc5, Mc8, Mc11, Ne3, Ne4).  Table II is a
list of over 130 characterized DNA methyltransferases.  A detailed list of
cloned restriction-modification genes has been made Wilson (Wi4).  Table III
lists the sensitivities of over 20 Type II DNA methyltransferases to m4C, m5C,
hm5C, and m6A modification.  Most DNA methyltransferases are sensitive to
non-canonical modifications within their recognition sequences (Bu5, Mc10, Ne3,
Po4), and this sensitivity may differ from that of their restriction
endonuclease partners.  Finally, several restriction endonuclease isoschizomers
are known to differ in their ability to cleave DNA which has been methylated.
Table IV lists over 20 known isoschizomer pairs and one isomethylator pair,
along with the modified recognition sites at which they differ.

<>

<1>Nelson, M., McClelland, M.
<2>Chromosome mapping at the megabase level: Guidelines for choosing restriction endonucleases for pulsed field gel mapping.
<3>Promega Notes
<4>18
<5>1-3
<6>1989
<7>A review.

<>

<1>Nelson, M., McClelland, M.
<2>Use of DNA methyltransferase/endonuclease enzyme combinations for megabase mapping of chromosomes.
<3>Methods Enzymol.
<4>216
<5>279-303
<6>1992
<7>Based on a knowledge of the methylation sensitivities of restriction-modification enzymes, and
the specificities of bacterial DNA methyltransferases, it is possible to mix and match
site-specific DNA methylation and restriction endonuclease cleavage reactions to produce rare
or novel DNA cleavages (Dobritsa and Dobritsa, 1980); Nelson et al., 1984; McClelland et al.,
1985) Detailed procedures for the purification, assay and use of methylase/endonuclease
combinations in sequential two-step reactions were described in a previous volume of this
series (McClelland, 1987, Nelson and McClelland, 1987; Nelson and Shildkraut, 1987).
Three-step procedures have also been described, based upon methylase/endonuclease combinations
and sequence-specific masking by bacterial repressor proteins (Koob and Szybalski, 1980,
polypyrimidine triplexes (Maher et al, 1989; Hanvey et al 1989), or other DNA
methyltransferases (McClelland and Nelson, 1987; McClelland and Nelson, 1988a; Posfai and
Szybalski, 1988). This article describes four selected DNA methylase/ endonuclease
combinations which may be used for megabase mapping of chromosome fragments in the size range
from 100 kb-2000 kb, paying special attention to features of these reactions which have
sometimes proven problematic.

<>

<1>Nelson, M., Raschke, E., McClelland, M.
<2>Effect of site-specific methylation on restriction endonucleases and DNA modification methyltransferases.
<3>Nucleic Acids Res.
<4>21
<5>3139-3154
<6>1993
<7>We present in Table I an updated list of the sensitivities of 298 restriction endonucleases
and 20 DNA methyltransferases to site-specific modification at 4-methylcytosine (m4C),
5-methylcytosine (m5C), 5-hydroxymethylcytosine (hm5C), and 6-methyladenine (m6A), four
modifications that are common in the DNA of prokaryotes, eukaryotes, and their viruses. In
addition, new information is included on restriction endonuclease cleavage at sites modified
with 5-hydroxymethyluracil (hm5U). Knowledge of the sensitivity of restriction endonucleases
to site-specific modification can be used to study cellular DNA methylation. Several
restriction-modification enzymes share the same recognition sequence specificity, but have
different sensitivities to site-specific methylation. Table II lists 32 known isoschizomer
pairs and one isomethylator pair, along with the modified recognition sites at which they
differ.

<>

<1>Nelson, M., Schildkraut, I.
<2>The use of DNA methylases to alter the apparent recognition specificities of restriction endonucleases.
<3>Methods Enzymol.
<4>155
<5>41-48
<6>1987
<7>DNA methylases can be used to alter the apparent recognition specificity of
restriction endonucleases.  These altered specificities are unique and increase
the list of cleavage sequences which can be utilized by molecular biologists.

<>

<1>Nelson, M., Zhang, Y., Van Etten, J.L.
<2>DNA methyltransferases and DNA-site-specific endonucleases encoded by chlorella viruses.
<3>DNA Methylation: Molecular Biology and Biological Significance, Birkhauser Verlag, Jost, J.P., Saluz, H.P., Basel, Switzerland
<4>0
<5>186-211
<6>1993
<7>Large polyhedral (diameter of 150 to 190 nm) dsDNA-containing (>30 kbp) viruses which infect
certain unicellular, eukaryotic, chlorella-like green algae are common in fresh water
collected throught the world (Van Etten et al, 1985; Schuster et al. 1986; Zhang et al., 1988;
Reisser et al., 1988; Yamada et al., 1991). The hosts for these lytic chlorella viruses are
exsymbiotic Chlorella strains NC64A and Pbi, originally isolated from the protozoan Paramecium
bursaria. Chlorella viruses, which can be produced in large quantities, are the first viruses
infecting a photosynthetic eukaryotic organism which can be plaque assayed (Van Etten et al.,
1983) and have been given family status with the name Phycodnaviridae (Francki et al., 1991).
A comprehensive review on the chlorella viruses has recently been published (Van Etten et al.,
1991).

<>

<1>Nelson, P.S., Papas, T.S., Schweinfest, C.W.
<2>Restriction endonuclease cleavage of 5-methyl-deoxycytosine hemimethylated DNA at high enzyme-to-substrate ratios.
<3>Nucleic Acids Res.
<4>21
<5>681-686
<6>1993
<7>We have investigated the ability of a large number of restriction enzymes to digest
non-canonically hemimethylated DNA at high enzyme-to-substrate ratios. A single-stranded
unmethylated phagemid was used as a template to complete synthesis of the second strand using
5-methyl-dCTP to substitute for all the deoxycytosine residues. A fragment of this
double-stranded hemimethylated DNA which contains the multiple cloning site region was used as
a substrate. For all the enzymes tested, at least some degree of protection from digestion is
observed. Sites completely protected from digestion by their cognate enzymes are SalI, BstXI,
SacI, SacII, SmaI, SstI, XhoI, PstI, HinfI, BamHI and AccI. Sites partially protected from
digestion by their cognate enzymes are XbaI, HindIII, KpnI, SpeI, ClaI, EcoRI and PvuII.
Knowledge of the sensitivity of commonly used restriction enzymes to hemimethylated substrates
is useful for several applications, which will be discussed.

<>

<1>Neoh, H.M., Mohamed-Hussein, Z.A., Tan, X.E., B, R.Abd.R.R.M., Hussin, S., Mohamad, Z.N., Jamal, R.
<2>Draft Genome Sequences of Four Nosocomial Methicillin-Resistant Staphylococcus aureus (MRSA) Strains (PPUKM-261-2009, PPUKM-332-2009, PPUKM-377-2009, and  PPUKM-775-2009) Representative of Dominant MRSA Pulsotypes Circulating in a  Malaysian University T.
<3>Genome Announcements
<4>1
<5>e00103-12
<6>2013
<7>Here, we report the draft genome sequences of four nosocomial methicillin-resistant
Staphylococcus aureus strains (PPUKM-261-2009, PPUKM-332-2009, PPUKM-377-2009, and
PPUKM-775-2009) isolated from a university teaching hospital in Malaysia. Three of the strains
belong to sequence type 239 (ST239), which has been associated with sustained hospital
epidemics worldwide.

<>

<1>Nerdal, W., Hare, D.R., Reid, B.R.
<2>Solution structure of the EcoRI DNA sequence: Refinement of NMR-derived distance geometry structures by NOESY spectrum back-calculations.
<3>Biochemistry
<4>28
<5>10008-10021
<6>1989
<7>The solution structure of the self-complementary DNA duplex [d(CGCGAATTCGCG)]2,
which contains the EcoRI restriction site sequence GAATTC at the center, has
been studied by two-dimensional nuclear magnetic resonance spectroscopy.
Time-dependent nuclear Overhauser effect spectra were used to obtain the
initial cross-relaxation rates between 155 pairs of protons.  These initial
cross-relaxation rates were converted into interproton distances and entered
into a distance (bounds) matrix.  A distance geometry algorithm (DSPACE) was
used to create embedded starting structures and to refine these structures
until they showed good agreement with the distance matrix; symmetry constraints
were included in the refinement procedure, making the two strands in the
refined distanced geometry structures virtually identical and significantly
improving the agreement with the distance matrix.  The NOESY spectrum for one
of these distance geometry was then calculated from the explicit coordinates by
numerically integrating all the z-magnetization transfer pathways among
neighboring protons within a specified radius. Distances in this distance
geometry structure that did not agree with the experimental NOESY time course
were then adjusted accordingly.  This process was iterated until a good
agreement between calculated and experimental NOESY spectra was reached.  The
final structure, which generates good agreement with the experimental NOESY
spectrum, display kinks at the C3-G4 base step and at the A6-T7 base step that
appear to be similar to those reported for the EcoRI restriction site DNA bound
to its endonuclease.  The solution structure is not the same as the crystal
structure of this DNA duplex.

<>

<1>Neri, A., Fazio, C., Ciammaruconi, A., Anselmo, A., Fortunato, A., Palozzi, A., Vacca, P., Fillo, S., Lista, F., Stefanelli, P.
<2>Draft Genome Sequence of C:P1.5-1,10-8:F3-6:ST-11 Meningococcal Clinical Isolate  Associated with a Cluster on a Cruise Ship.
<3>Genome Announcements
<4>2
<5>e01263-14
<6>2014
<7>Meningococcal serogroup C strains, in particular those belonging to the ST-11 clonal complex,
are known to cause invasive diseases worldwide. We report the
genome sequence of a Neisseria meningitidis strain linked to a cluster of cases
of invasive meningococcal disease on a cruise ship that was described in 2012.

<>

<1>Neri, F., Krepelova, A., Incarnato, D., Maldotti, M., Parlato, C., Galvagni, F., Matarese, F., Stunnenberg, H.G., Oliviero, S.
<2>Dnmt3L Antagonizes DNA Methylation at Bivalent Promoters and Favors DNA Methylation at Gene Bodies in ESCs.
<3>Cell
<4>155
<5>121-134
<6>2013
<7>The de novo DNA methyltransferase 3-like (Dnmt3L) is a catalytically inactive DNA
methyltransferase that cooperates with Dnmt3a and Dnmt3b to methylate DNA. Dnmt3L
is highly expressed in mouse embryonic stem cells (ESCs), but its function in
these cells is unknown. Through genome-wide analysis of Dnmt3L knockdown in ESCs,
we found that Dnmt3L is a positive regulator of methylation at the gene bodies of
housekeeping genes and, more surprisingly, is also a negative regulator of
methylation at promoters of bivalent genes. Dnmt3L is required for the
differentiation of ESCs into primordial germ cells (PGCs) through the activation
of the homeotic gene Rhox5. We demonstrate that Dnmt3L interacts with the
Polycomb PRC2 complex in competition with the DNA methyltransferases Dnmt3a and
Dnmt3b to maintain low methylation levels at the H3K27me3 regions. Thus, in ESCs,
Dnmt3L counteracts the activity of de novo DNA methylases to maintain
hypomethylation at promoters of bivalent developmental genes.

<>

<1>Nesbo, C.L., Boucher, Y., Dlutek, M., Doolittle, W.F.
<2>Lateral gene transfer and phylogenetic assignment of environmental fosmid clones.
<3>Environ. Microbiol.
<4>7
<5>2011-2026
<6>2005
<7>Metagenomic data, especially sequence data from large insert clones, are
most useful when reasonable inferences about phylogenetic origins of
inserts can be made. Often, clones that bear phylotypic markers (usually
ribosomal RNA genes) are sought, but sometimes phylogenetic assignments
have been based on the preponderance of blast hits obtained with predicted
protein coding sequences (CDSs). Here we use a cloning method which
greatly enriches for ribosomal RNA-bearing fosmid clones to ask two
questions: (i) how reliably can we judge the phylogenetic origin of a
clone (that is, its RNA phylotype) from the sequences of its CDSs? and
(ii) how much lateral gene transfer (LGT) do we see, as assessed by CDSs
of different phylogenetic origins on the same fosmid? We sequenced 12 rRNA
containing fosmid clones, obtained from libraries constructed using DNA
isolated from Baltimore harbour sediments. Three of the clones are from
bacterial candidate divisions for which no cultured representatives are
available, and thus represent the first protein coding sequences from
these major bacterial lineages. The amount of LGT was assessed by making
phylogenetic trees of all the CDSs in the fosmid clones and comparing the
phylogenetic position of the CDS to the rRNA phylotype. We find that the
majority of CDSs in each fosmid, 57-96%, agree with their respective rRNA
genes. However, we also find that a significant fraction of the CDSs in
each fosmid, 7-44%, has been acquired by LGT. In several cases, we can
infer co-transfer of functionally related genes, and generate hypotheses
about mechanism and ecological significance of transfer.

<>

<1>Nesterenko, V.F., Buryanov, Y.I., Baev, A.A.
<2>Isolation and properties of DNA-cytosine-methylase I from Escherichia coli MRE 600.
<3>Biokhimiia
<4>44
<5>130-140
<6>1979
<7>DNA-cytosine-methylase I was isolated and purified to homogeneity.  The yield
made up to about 30% of total activity.  The enzyme molecular weight as
determined by centrifugation in a sucrose gradient, by gel filtration and by
electrophoresis in polyacrylamide gel in the presence of sodium dodecyl sulfate
was found to be 45000.  The Michaelis constant was 1,8.10-6 M for SAM and
2.10-4 M for DNA.  DNA-cytosine-methylase I modifies phage lambda DNA in 60
sites.  This modification does not protect DNA from the effects of restriction
endonucleases HpaII and BsuRI.  The enzyme methylates DNA in the nucleotide
sequence: 5'...Pu-MC-C-G-G-Py...3'.

<>

<1>Nesterenko, V.F., Buryanov, Y.I., Baev, A.A.
<2>Determination of DNA-cytosine methylase I (M. Eco MRE600I) in plasmid CoLA.
<3>Dokl. Akad. Nauk.
<4>250
<5>1265-1267
<6>1980
<7>None

<>

<1>Nesterenko, V.F., Buryanov, Y.I., Baev, A.A.
<2>Bacterial DNA-cytosine methylase extracted from cells of Escherichia coli MRE600 strain with subsequent chromatography and dialysis.
<3>Soviet Patent Office
<4>SU 737443 A
<5>
<6>1980
<7>Enzyme DNA-cytosine-methylase 1 is obtained from cells of Escherichia coli MRE600 by:
disintegrating the biomass; centrifuging the resulting homogenate at 10000-30000 G;
precipitating nucleic acids from the cell-free extract with protamine sulphate; fractionating
the remaining protein solution with (NH4)2SO4; desalting the enzyme-containing fraction by gel
filtration and chromatography using Sephadex.  The protein solution is then chromatographed
twice on DNA-agarose and then on carboxy-methyl cellulose. The resulting solution is passed
down a series column through cellulose and is then finally stabilised by dialysis against a
glycerol solution. The resulting enzyme product is homogeneous and is obtained in good yield.

<>

<1>Nestor, C., Ruzov, A., Meehan, R.R., Dunican, D.S.
<2>Enzymatic approaches and bisulfite sequencing cannot distinguish between 5-methylcytosine and 5-hydroxymethylcytosine in DNA.
<3>Biotechniques
<4>48
<5>317-319
<6>2010
<7>DNA cytosine methylation (5mC) is highly abundant in mammalian cells and is associated with
transcriptional repression. Recently, hydroxymethylcytosine
(hmC) has been detected at high levels in certain human cell types; however,
its roles are unknown. Due to the structural similarity between 5mC and hmC,
it is unclear whether 5mC analyses can discriminate between these nucleotides.
Here we show that 5mC and hmC are experimentally indistinguishable using
established 5mC mapping methods, thereby implying that existing 5mC data
sets will require careful re-evaluation in the context of the possible presence of
hmC. Potential differential enrichment of 5mC and hmC DNA sequences
may be facilitated using a 5mC monoclonal antibody.

<>

<1>Netesov, S.V., Grachev, S.A.
<2>An effective method for isolation of restriction endonucleases.
<3>Bioorg. Khim.
<4>7
<5>790-791
<6>1981
<7>A modification of the published method for isolating the restriction
endonucleases is proposed.  The modification involves the use of Triton X-100
containing buffers at all steps of the isolation procedure which results in a
2-12-fold increase in the yield of enzymes depending on the strain of bacteria.

<>

<1>Netesova, N.A., Golikova, L.N., Ovetchkina, L.G., Evdokimov, A.A., Malygin, E.G., Gololobova, N.S., Gonchar, D.A., Degtyarev, S.K.
<2>Comparative study of the M.BstF5I-1 and M.BstF5I-3 DNA methyltransferases from the Bacillus stearothermophilus F5 restriction-modification system.
<3>Mol. Biol. (Mosk)
<4>36
<5>136-143
<6>2002
<7>The BstF5I restriction-modification system from Bacillus stearothermophilus F5, unlike all
known restriction-modification
systems, contains three genes encoding DNA methyltransferases. In
addition to revealing two DNA methylases responsible for modification
of adenine in different DNA strands, it has been first shown that one
bacterial cell has two DNA methylases, M.BstF5I-1 and M.BstF5I-3, with
similar substrate specificity. The boundaries of the gene for DNA
methyltransferase M.BstF5I-1 have been verified. The bstF5IM-1 gene was
cloned in pJW and expressed in Escherichia coli. Homogeneous samples of
M.BstF5I-1 and M.BstF5I-3 were obtained by chromatography with
different sorbents. The main kinetic parameters have been determined
for M.BstF5I-1 and M.BstF5I-3, both modifying adenine in the
recognition site 5'-GGATG-3'.

<>

<1>Neupane, S. et al.
<2>Complete genome sequence of Serratia plymuthica strain AS12.
<3>Standards in Genomic Sciences
<4>6
<5>165-173
<6>2012
<7>A plant-associated member of the family Enterobacteriaceae, Serratia plymuthica strain AS12
was isolated from rapeseed roots. It is of scientific interest
because it promotes plant growth and inhibits plant pathogens. The genome of S.
plymuthica AS12 comprises a 5,443,009 bp long circular chromosome, which consists
of 4,952 protein-coding genes, 87 tRNA genes and 7 rRNA operons. This genome was
sequenced within the 2010 DOE-JGI Community Sequencing Program (CSP2010) as part
of the project entitled 'Genomics of four rapeseed plant growth promoting
bacteria with antagonistic effect on plant pathogens'.

<>

<1>Neupane, S. et al.
<2>Complete genome sequence of the rapeseed plant-growth promoting Serratia plymuthica strain AS9.
<3>Standards in Genomic Sciences
<4>6
<5>54-62
<6>2012
<7>Serratia plymuthica are plant-associated, plant beneficial species belonging to the family
Enterobacteriaceae. The members of the genus Serratia are ubiquitous
in nature and their life style varies from endophytic to free-living. S.
plymuthica AS9 is of special interest for its ability to inhibit fungal pathogens
of rapeseed and to promote plant growth. The genome of S. plymuthica AS9
comprises a 5,442,880 bp long circular chromosome that consists of 4,952
protein-coding genes, 87 tRNA genes and 7 rRNA operons. This genome is part of
the project entitled 'Genomics of four rapeseed plant growth promoting bacteria
with antagonistic effect on plant pathogens' awarded through the 2010 DOE-JGI
Community Sequencing Program (CSP2010).

<>

<1>Neupane, S. et al.
<2>Complete genome sequence of the plant-associated Serratia plymuthica strain AS13.
<3>Standards in Genomic Sciences
<4>7
<5>22-30
<6>2012
<7>AS13 is a plant-associated , isolated from rapeseed roots. It is of special interest because
of its ability to inhibit fungal pathogens of rapeseed and to
promote plant growth. The complete genome of AS13 consists of a 5,442,549 bp
circular chromosome. The chromosome contains 4,951 protein-coding genes, 87 tRNA
genes and 7 rRNA operons. This genome was sequenced as part of the project
entitled 'Genomics of four rapeseed plant growth promoting bacteria with
antagonistic effect on plant pathogens' within the 2010 DOE-JGI Community
Sequencing Program (CSP2010).

<>

<1>Neupane, S. et al.
<2>Non-contiguous finished genome sequence of plant-growth promoting Serratia proteamaculans S4.
<3>Standards in Genomic Sciences
<4>8
<5>441-449
<6>2013
<7>Serratia proteamaculans S4 (previously Serratia sp. S4), isolated from the rhizosphere of wild
Equisetum sp., has the ability to stimulate plant growth and
to suppress the growth of several soil-borne fungal pathogens of economically
important crops. Here we present the non-contiguous, finished genome sequence of
S. proteamaculans S4, which consists of a 5,324,944 bp circular chromosome and a
129,797 bp circular plasmid. The chromosome contains 5,008 predicted genes while
the plasmid comprises 134 predicted genes. In total, 4,993 genes are assigned as
protein-coding genes. The genome consists of 22 rRNA genes, 82 tRNA genes and 58
pseudogenes. This genome is a part of the project 'Genomics of four rapeseed
plant growth-promoting bacteria with antagonistic effect on plant pathogens'
awarded through the 2010 DOE-JGI's Community Sequencing Program.

<>

<1>Newby, A.E.R., Lau, E.Y., Bruice, T.C.
<2>A theoretical examination of the factors controlling the catalytic efficiency of the DNA-(adenine-N6)-methyltransferase from Thermus aquaticus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>7922-7927
<6>2002
<7>Ab initio and density functional calculations have been carried out to more fully understand
the factors controlling the catalytic activity of the Thermus aquaticus DNA methyltransferase
(M.TaqI) in the N-methylation at the N6 of an adenine nucleobase. The noncatalyzed reaction
was modeled as a methyl transfer from trimethylsulfonium to the N6 of adenine. Activation
barriers of 32.0 kcal/mol and 24.0 kcal/mol were predicted for the noncatalyzed reaction in
the gas phase by MP2/6-31+G(d,p)//HF/6-31+G(d,p) and B3LYP/6-31+G(d,p) calculations,
respectively. Calculations performed to evaluate the effect of substrate positioning in the
active site of MTaqI on the reaction determine the barrier to be 23.4 kcal/mol and 17.3
kcal/mol for the MP2/6-31+G(d,p)//HF/6-31+G(d,p) and B3LYP/6-31+G(d,p) gas phase calculations,
respectively. The effect of hydrogen bonding between the N6 of adenine and the terminal oxygen
of Asn-105 on the activation barrier was also studied. A formamide molecule was modeled into
the system to mimic the function of active site residue Asn-105. The activation barrier for
this reaction was found to be 21.8 kcal/mol and 15.8 kcal/mol as determined from the
MP2/6-31+G(d,p)//HF/6-31+G(d,p) and B3LYP/6-31+G(d,p) calculations, respectively. This result
predicts a contribution of less than 2 kcal/mol to the lowering of the activation barrier from
amide hydrogen bonding between formamide and N6 of adenine. Comparison of the reaction
coordinates suggest that it is not the hydrogen bonding of the Asn-105 that lends to the
catalytic prowess of the enzyme since the organization of the substrates in the active site of
the enzyme has a far greater effect on reducing the activation barrier. The results also
suggest a stepwise mechanism for the removal of the hydrogen from the N6 of adenine as opposed
to a concerted reaction in which a proton is abstracted simultaneously with the transfer of
the methyl group. The hydrogen on the N6 of the intermediate methyl adenine product is far
more acidic than in the reactant complex and may be subsequently abstracted by basic groups in
the active site that are too weak to abstract the proton before the full sp3 hybridization of
the attacking nitrogen.

<>

<1>Newman, A.K., Rubin, R.A., Kim, S.H., Modrich, P.
<2>DNA Sequences of Structural Genes for EcoRI DNA Restriction and Modified Enzymes.
<3>J. Biol. Chem.
<4>256
<5>2131-2139
<6>1981
<7>We have determined the sequence of a 2210-base pair DNA segment containing genes required for
expression of EcoRI DNA restriction and modification phenotypes.  Polypeptide sequences
encoded within the two longest open reading frames in this region are in agreement with NH2-
and C00H-terminal sequences of the two EcoRI enzymes (Rubin, R.A., Modrich, P., and Vanaman,
T.C. (1981) J. Biol. Chem. 256, 2140-2142), indicating that these are structural genes for the
two proteins.  The DNA sequence encoding EcoRI endonuclease specifies a 277-residue
polypeoptide (Mr = 31,063) while the methylase gene encodes a 326-residue protein (Mr =
38,048).  Genes for the two proteins are nonoverlapping, being separated by a 29-nucleotide
intercistronic region. Analysis of each polypeptide and its gene sequence for internal regions
of homology led to identification of a striking 4-fold repeat of Leu-Ile-Lys within the
methylase and a tandem repeat within the endonuclease, both of which are unlikely on the basis
of chance.  Comparison of DNA and polypeptide sequences also demonstrated a limited, but
statistically significant, region of homology between EcoRI endonuclease and methylase primary
structures. However, the general lack of homology between the two polypeptides suggests
different evolutionary origins for the two proteins.  The two enzymes also differ markedly at
higher levels of structure as judged by circular dichroism and prediction of secondary
structure by the probabilistic method of Chou and Fasman (Chou, P.Y., and Fasman, G.D. (1978)
Adv. Enzymol. 47, 45-148).  Thus, despite their ability to interact with the same DNA
sequence, the EcoRI enzymes have quite distinct structural properties.

<>

<1>Newman, M., Lunnen, K., Wilson, G., Greci, J., Schildkraut, I., Phillips, S.E.V.
<2>Crystal structure of restriction endonuclease BglI bound to its interrupted DNA recognition sequence.
<3>EMBO J.
<4>17
<5>5466-5476
<6>1998
<7>The crystal structure of the type II restriction endonuclease BglI bound to DNA containing its
specific recognition sequence has been determined at 2.2 A resolution.  This is the first
structure of a restriction endonuclease that recognizes and cleaves an interrupted DNA
sequence, producing 3' overhanging ends.  BglI is a homodimer that binds its specific DNA
sequence with the minor groove facing the protein.  Parts of the enzyme reach into both the
major and minor grooves to contact the edges of the bases within the recognition half-sites.
The arrangement of active site residues is strikingly similar to other restriction
endonucleases, but the coordination of two calcium ions at the active site gives new insight
into the catalytic mechanism.  Surprisingly, the core of a BglI subunit displays a striking
similarity to subunits of EcoRV and PvuII, but the dimer structure is dramatically different.
The BglI DNA complex demonstrates, for the first time, that a conserved subunit fold can
dimerize in more than one way, resulting in different DNA cleavage patterns.

<>

<1>Newman, M., Strzelecka, T., Dorner, L.F., Schildkraut, I., Aggarwal, A.K.
<2>Structure of restriction endonuclease BamHI and its relationship to EcoRI.
<3>Nature
<4>368
<5>660-664
<6>1994
<7>Type II restriction endonucleases are characterized by the remarkable specificity with which
they cleave specific DNA sequences. Surprisingly, their protein sequences are in most cases
unrelated, and no recurring structural motif has yet been identified. We have determined the
structure of restriction endonuclease BamHI at 1.95 angstrom resolution. BamHI shows striking
resemblance to the structure of endonuclease EcoRI, despite the lack of sequence similarity
between them. We also observe some curious differences between the two structures, and propose
an evolutionary scheme that may explain them. The active site of BamHI is structurally similar
to the active sites of EcoRI and EcoRV, but the mechanism by which BamHI activates a water
molecule for nucleophilic attack may be different.

<>

<1>Newman, M., Strzelecka, T., Dorner, L.F., Schildkraut, I., Aggarwal, A.K.
<2>Structure of restriction endonuclease BamHI phased at 1.95 A resolution by MAD analysis.
<3>Structure
<4>2
<5>439-452
<6>1994
<7>Type II restriction endonucleases recognize DNA sequences that vary between four to eight base
pairs, and require only Mg2+ as a cofactor to catalyze the hydrolysis of DNA. Their protein
sequences display a surprising lack of similarity, and no recurring structural motif analogous
to the helix-turn-helix or the zinc finger of transcription factors, has yet been discovered.
We have determined the crystal structure of restriction endonuclease BamHI at 1.95 A
resolution. The structure was solved by combining phase information derived from
multi-wavelength X-ray data by algebraic and maximum likelihood methods. The BamHI subunit
consists of a central beta-sheet with alpha-helices on both sides. The dimer configuration
reveals a large cleft which could accommodate B-form DNA. Mutants of the enzyme that are
deficient in cleavage are located at or near the putative DNA-binding cleft. BamHI and
endonuclease EcoRI share a common core motif (CCM) consisting of five beta-strands and two
helices. It remains to be determined if other restriction enzymes also contain the CCM. The
structure of BamHI provides the first clear evidence that there may be substantial structural
homology amongst restriction enzymes, even though it is undetectable at the sequence level.

<>

<1>Newman, M., Strzelecka, T., Dorner, L.F., Schildkraut, I., Aggarwal, A.K.
<2>Structure of BamHI endonuclease bound to DNA: partial folding and unfolding on DNA binding.
<3>Science
<4>269
<5>656-663
<6>1995
<7>The crystal structure of restriction endonuclease BamHI complexed to DNA has been determined
at 2.2 angstrom resolution.  The DNA binds in the cleft and retains a B-DNA type of
conformation.  The enzyme, however, undergoes a series of conformational changes, including
rotation of subunits and folding of disordered regions.  The most striking conformational
change is the unraveling of carboxyl-terminal alpha helices to form partially disordered
"arms".  The arm from one subunit fits into the minor groove while the arm from the symmetry
related subunit follows the DNA sugar-phosphate backbone.  Recognition of DNA base pairs
occurs primarily in the major groove, with a few interactions occurring in the minor groove.
Tightly bound water molecules play an equally important role as side chain and main chain
atoms in the recognition of base pairs.  The complex also provides new insights into the
mechanism by which the enzyme catalyzes the hydrolysis of DNA phosphodiester groups.

<>

<1>Newman, P.C.
<2>Investigation of the sequence-specific DNA/protein interactions of the EcoRV restriction enzyme.
<3>Diss. Abstr.
<4>51
<5>3366B
<6>1991
<7>The EcoRV restriction enzyme from Escherichia coli recognises the sequence GATATC on double
stranded DNA and cuts between the central residues with very high specificity.  The self
complementary dodecadeoxynucleotide dGACGATATCGTC which contains the EcoRV recognition site
(underlined) is also a substrate for the endonuclease.  The thesis describes the synthesis of
fourteen dodecamers of the above parent sequence in which the functional groups of the central
ATAT residues accessible to the protein via the DNA major and minor grooves have been
systematically and sequentially deleted.  Conservative contact deletions were achieved by the
substitution of the two deoxyadenosine residues in turn with the base analogues purine
deoxyriboside (dP), 7-deazadeoxyadenosine (d7CA) and 3-deazadeoxyadenosine (d3CA).  Similarly
the two thymidine residues were substituted with deoxyuridine (dU), 5-methyl-2-pyrimidinone
deoxyriboside (d4HT), 4-thiothymidine (d4ST) and 2-thiothymidine (d2ST).  To obtain the
complete set of analogues, efficient synthetic routes for the formation of derivatives of
d4HT, d4ST and d2ST suitable for oligodeoxynucleotide synthesis using the cyanoethyl
phosphoramidite approach are described (also see Connolly

<>

<1>Newman, P.C., Nwosu, V.U., Williams, D.M., Cosstick, R., Seela, F., Connolly, B.A.
<2>Incorporation of a complete set of deoxyadenosine and thymidine analogues suitable for the study of protein nucleic acid interactions into oligodeoxynucleotides.  Application to the EcoRV restriction endonuclease and modification methylase.
<3>Biochemistry
<4>29
<5>9891-9901
<6>1990
<7>A complete set of dA and T analogues designed for the study of protein DNA
interactions has been prepared.  These modified bases have been designed by
considering the groups on the dA and T bases that are accessible to proteins
when these bases are incorporated into double-helical B-DNA [Seeman, N.C.,
Rosenberg, J.M. and Rich, A. (1976) Proc. Natl. Acad. Sci. USA 73, 804-808].
Each of the positions on the two bases, having the potential to interact with
proteins, have been subject to nondisruptive, conservative change.  Typically a
particular group (e.g., the 6-amino group of dA or the 5-methyl of T) has been
replaced with a hydrogen atom.  Occasionally keto groups (the 2- and 4-keto
oxygen atoms of T) have been replaced with sulfur.  The base set has been
incorporated into the self-complementary dodecamer d(GACGATATCGTC) at the
central d(ATAT) sequence.  Melting temperature determination shows that the
modified bases do not destabilize the double helix.  Additionally, circular
dichroism spectroscoy shows that almost all the altered bases have very little
effect on overall oligodeoxynucleotide conformation and that most of the
modified oligomers have a B-DNA type structure.  d(GATATC) is the recognition
sequence for the EcoRV restriction modification system.  Initial rate
measurements (at a single oligodeoxynucleotide concentration of 20 microM) have
been carried out with both the EcoRV restriction endonuclease and modification
methylase.  This has enabled a preliminary identification of the groups of the
dA and T bases within the d(GATATC) sequence that make important contacts to
both proteins.

<>

<1>Newman, P.C., Williams, D.M., Cosstick, R., Seela, F., Connolly, B.A.
<2>Interaction of the EcoRV restriction endonuclease with the deoxyadenosine and thymidine bases in its recognition hexamer d(GATATC).
<3>Biochemistry
<4>29
<5>9902-9910
<6>1990
<7>A set of dA and T analogues suitable for the study of protein DNA interactions
have been incorporated into the central d(ATAT) sequence within
d(GACGATATCGTC).  The individual analogues have one potential protein contact
(either a hydrogen-bonding group or a CH3 group capable of a van der Waals
interaction) deleted.  In general, the modified bases do not perturb the
overall structure of the dodecamer, enabling results obtained to be simply
interpreted in terms of loss of protein DNA contacts.  We have used the
modified oligodeoxynucleotide set to study the recognition of DNA by the EcoRV
restriction endonuclease [recognition sequence d(GATATC)].  The kcat and Km
values for the set have been determined, and a comparison with results seen
with the parent oligodeoxynucleotide (containing no modified bases) has been
carried out.  Three classes of results are seen.  First, some analogues lead to
no change in kinetic parameters, meaning no enzyme contact at the altered site.
Second (this is seen for most of the modified oligodeoxynucleotides), a drop
in the kcat/Km ratio relative to the parent is observed.  This comes mainly
from a decrease in kcat, implying that the endonuclease uses the interaction
under study to lower the transition-state barrier rather than to bind the
substrate.  Analyses of these results show that the drop in kcat/Km is what
would be expected for the simple loss of a hydrogen bond or a CH3 contact
between the enzyme and the oligodeoxynucleotide.  This implies a contact of
these types at these sites.  Third, some analogue-containing
oligodeoxynucleotides are not substrates; i.e., the kcat/Km drop is much
greater than would be expected for loss of a single hydrogen bond or CH3
contact.  These results are interpreted in terms of a cooperative mechanism
whereby the loss of one interaction causes a rearrangement at the enzyme active
site leading to a consequent loss of further protein substrate contacts.
However, in these cases gross structural changes in the oligodeoxynucleotide
conformation cannot be excluded.  It is found that the endonuclease makes very
many interactions to the d(ATAT) sequence within its d(GATATC) recognition
site, and these occur in both the major and minor grooves.  The results
obtained have been used to explain how the enzyme achieves the high degree of
cleavage specificity for d(GATATC) as compared to all other sequences.

<>

<1>Ng, E.K.O., Tsang, W.P., Ng, S.S.M., Jin, H.C., Yu, J., Li, J.J., Roecken, C., Ebert, M.P.A., Kwok, T.T., Sung, J.J.Y.
<2>MicroRNA-143 targets DNA methyltransferases 3A in colorectal cancer.
<3>Br. J. Cancer
<4>101
<5>699-706
<6>2009
<7>BACKGROUND: MicroRNAs (miRNAs) are 19-25-nucleotides regulatory non-protein-coding RNA
molecules that regulate the expressions of a
wide variety of genes, including some involved in cancer development.
In this study, we investigated the possible role of miR-143 in
colorectal cancer (CRC).METHODS: Expression levels of human mature
miRNAs were examined using real-time PCR-based expression arrays on
paired colorectal carcinomas and adjacent non-cancerous colonic
tissues. The downregulation of miR-143 was further evaluated in colon
cancer cell lines and in paired CRC and adjacent non-cancerous colonic
tissues by qRT-PCR. Potential targets of miR-143 were defined. The
functional effect of miR-143 and its targets was investigated in human
colon cancer cell lines to confirm miRNA-target association.RESULTS:
Both real-time PCR-based expression arrays and qRT-PCR showed that
miR-143 was frequently downregulated in 87.5% (35 of 40) of colorectal
carcinoma tissues compared with their adjacent non-cancerous colonic
tissues. Using in silico predictions, DNA methyltranferase 3A (DNMT3A)
was defined as a potential target of miR-143. Restoration of the
miR-143 expression in colon cell lines decreased tumour cell growth and
soft-agar colony formation, and downregulated the DNMT3A expression in
both mRNA and protein levels. DNMT3A was shown to be a direct target of
miR-143 by luciferase reporter assay. Furthermore, the miR-143
expression was observed to be inversely correlated with DNMT3A mRNA and
protein expression in CRC tissues.CONCLUSION: Our findings suggest that
miR-143 regulates DNMT3A in CRC. These findings elucidated a
tumour-suppressive role of miR-143 in the epigenetic aberration of CRC,
providing a potential development of miRNA-based targeted approaches
for CRC therapy. British Journal of Cancer (2009) 101, 699-706. doi:
10.1038/sj.bjc.6605195 www.bjcancer.com Published online 28 July 2009

<>

<1>Ng, K.P., Yew, S.M., Chan, C.L., Chong, J., Tang, S.N., Soo-Hoo, T.S., Na, S.L., Hassan, H., Ngeow, Y.F., Hoh, C.C., Lee, K.W., Yee, W.Y.
<2>Draft Genome Sequence of the First Isolate of Extensively Drug-Resistant (XDR) Mycobacterium tuberculosis in Malaysia.
<3>Genome Announcements
<4>1
<5>e00056-12
<6>2013
<7>The emergence of the global threat of extensively drug-resistant (XDR) Mycobacterium
tuberculosis reveals weaknesses in tuberculosis management and diagnostic services. We report
the draft genome sequence of the first extensively drug-resistant Mycobacterium tuberculosis
strain isolated in Malaysia. The sequence was also compared against a reference strain to
elucidate the polymorphism that is related to their extensive resistance.

<>

<1>Ng, W.V., Ciufo, S.A., Smith, T.M., Bumgarner, R.E., Baskin, D., Faust, J., Hall, B., Loretz, C., Seto, J., Slagel, J., Hood, L., DasSarma, S.
<2>Snapshot of a large dynamic replicon in a halophilic archaeon: Megaplasmid or minichromosome?
<3>Genome Res.
<4>8
<5>1131-1141
<6>1998
<7>Extremely halophilic archaea, which flourish in hypersaline environments, are known to contain
a variety of large dynamic replicons. Previously, the analysis of one such replicon, pNRC100,
in Halobacterium sp. strain NRC-1, showed that it undergoes high-frequency insertion sequence
element-mediated insertions and deletions, as well as inversions via recombination between
39-kb-long inverted repeats. Now, the complete sequencing of pNRC100, a 191,346-bp circle, has
shown the presence of 27 IS elements representing eight families.  A total of 176 ORFs or
likely genes of 850-bp average size were found, 39 of which were repeated within the large
IRs. More than one-half of the ORFs are likely to represent novel genes that have no known
homologs in the databases. Among ORFs with previously characterized homologs, three different
copies of putative plasmid replication and four copies of partitioning genes were found,
suggesting that pNRC100 evolved from IS element-mediated fusions of several smaller plasmids.
Consistent with this idea, putative genes typically found on plasmids, including those
encoding a restriction-modification system and arsenic resistance, as well as buoyant
gas-filled vesicles and a two-component regulatory system, were found on pNRC100.  However,
additional putative genes not expected on an extrachromosomal element, such as those encoding
an electron transport chain cytochrome d oxidase, DNA nucleotide synthesis enzymes thioredoxin
and thioredoxin reductase, and eukaryotic-like TATA-binding protein transcription factors and
a chromosomal replication initiator protein were also found.  A multi-step IS element-mediated
process is proposed to account for the acquisition of these chromosomal genes.  The finding of
essential genes on pNRC100 and its property of resistance to curing suggest that this replicon
may be evolving into a new chromosome.

<>

<1>Ngeow, Y.F., Leong, M.L., Wong, Y.L., Wong, G.J., Tan, J.L., Wee, W.Y., Ong, C.S., Pang, Y.K., Choo, S.W.
<2>Draft Genome Sequence of Mycobacterium massiliense Strain M159, Showing Phenotypic Resistance to beta-Lactam and Tetracycline Antibiotics.
<3>Genome Announcements
<4>1
<5>e00669-13
<6>2013
<7>Mycobacterium massiliense is a nontuberculous mycobacterium associated with human infections.
We report here the draft genome sequence of M. massiliense strain
M159, isolated from the bronchial aspirate of a patient who had a pulmonary
infection. This strain showed genotypic and in vitro resistance to a number of
tetracyclines and beta-lactam antibiotics.

<>

<1>Ngeow, Y.F., Wee, W.Y., Wong, Y.L., Tan, J.L., Ongi, C.S., Ng, K.P., Choo, S.W.
<2>Genomic Analysis of Mycobacterium abscessus Strain M139, Which Has an Ambiguous Subspecies Taxonomic Position.
<3>J. Bacteriol.
<4>194
<5>6002-6003
<6>2012
<7>Mycobacterium abscessus is a ubiquitous, rapidly growing species of nontuberculous
mycobacteria that colonizes organic surfaces and is frequently
associated with opportunistic infections in humans. We report here the draft
genome sequence of Mycobacterium abscessus strain M139, which shows genomic
features reported to be characteristic of both Mycobacterium abscessus subsp.
abscessus and Mycobacterium abscessus subsp. massiliense.

<>

<1>Ngeow, Y.F., Wong, Y.L., Lokanathan, N., Wong, G.J., Ong, C.S., Ng, K.P., Choo, S.W.
<2>Genomic Analysis of Mycobacterium massiliense Strain M115, an Isolate from Human  Sputum.
<3>J. Bacteriol.
<4>194
<5>4786
<6>2012
<7>We report the draft genome sequence of a clinical isolate, strain M115, identified as
Mycobacterium massiliense, a member of the newly created taxon of
Mycobacterium abscessus subspecies bolletii comb. nov.

<>

<1>Ngeow, Y.F., Wong, Y.L., Tan, J.L., Arumugam, R., Wong, G.J., Ong, C.S., Ng, K.P., Choo, S.W.
<2>Genome Sequence of Mycobacterium massiliense M18, Isolated from a Lymph Node Biopsy Specimen.
<3>J. Bacteriol.
<4>194
<5>4125
<6>2012
<7>Mycobacterium massiliense is a rapidly growing mycobacterial species. The pathogenicity of
this subspecies is not well known. We report here the annotated
genome sequence of M. massiliense strain M18, which was isolated from a lymph
node biopsy specimen from a Malaysian patient suspected of having tuberculous
cervical lymphadenitis.

<>

<1>Ngeow, Y.F., Wong, Y.L., Tan, J.L., Ong, C.S., Ng, K.P., Choo, S.W.
<2>Genome Sequence of Mycobacterium abscessus Strain M152.
<3>J. Bacteriol.
<4>194
<5>6662
<6>2012
<7>Mycobacterium abscessus is an environmental bacterium with increasing clinical relevance.
Here, we report the annotated whole-genome sequence of M. abscessus
strain M152.

<>

<1>Nguyen, L.D., Cajthamlova, K., Nguyen, H.T., Weiser, J., Holubova, I., Weiserova, M.
<2>Identification of the EcoKI and EcoR124I type I restriction-modification enzyme subunits by non-equilibrium pH gradient two-dimensional gel electrophoresis.
<3>Folia Microbiol. (Praha)
<4>47
<5>641-648
<6>2002
<7>Effectively optimized and reproducible procedure for monitoring the composition of type I
restriction-modification endonucleases EcoKI and
EcoR124I by non-equilibrium pH gradient two-dimensional (2-D) gel
electrophoresis is described. Three subunits of the enzyme complex,
which widely differ from one another in their isoelectric points and
molar mass, were identified in crude cell extracts of E. coli. For the
first time all three subunits of both EcoKI and EcoR124I were detected
as distinct spots on a single 2-D gel. A sensitive immunoblotting
procedure was suggested suitable for routine use in determining the
identity of individual subunits. Potential application of this method
for detailed studies of regulation of the function and stoichiometry of
the enzyme complexes is discussed.

<>

<1>Nho, S.W., Hikima, J.I., Cha, I.S., Park, S.B., Jang, H.B., Del Castillo, C.S., Kondo, H., Hirono, I., Aoki, T., Jung, T.S.
<2>Complete genome sequence and immunoproteomic analyses of the fish bacterial pathogen Streptococcus parauberis.
<3>J. Bacteriol.
<4>193
<5>3356-3366
<6>2011
<7>Although Streptococcus parauberis is known as a bacterial pathogen associated with bovine
udder mastitis, it has recently become one of the
major causative agents of olive flounder (Paralichthys olivaceus)
streptococcosis in northeast Asia, causing massive mortality resulting in
severe economic losses. S. parauberis contains two serotypes, and it is
likely that capsular polysaccharide antigens serve to differentiate the
serotypes. In the present study, the complete genome sequence of S.
parauberis (serotype I) was determined using the GS-FLX system to
investigate its phylogeny, virulence factors, and antigenic proteins. S.
parauberis possesses a single chromosome of 2,143,887 bp containing 1,868
predicted coding sequences (CDSs) with an average GC content of 35.6%.
Whole-genome dot plot analysis and phylogenetic analysis of a 60 kDa
chaperonin-encoding gene and the GAPDH-encoding gene showed that the
strain was evolutionarily closely related to S. uberis. S. parauberis
antigenic proteins were analyzed using an immunoproteomic technique.
Twenty-one antigenic protein spots were identified in S. parauberis, by
reaction with an antiserum obtained from S. parauberis-challenged olive
flounder. This work provides the foundation needed to understand more
clearly the relationship between pathogen and host, and develops new
approaches toward prophylactic and therapeutic strategies to deal with
streptococcosis in fish. The work also provides a better understanding of
the physiology and evolution of a significant representative of the
Streptococcaceae.

<>

<1>Niazi, A., Manzoor, S., Bejai, S., Meijer, J., Bongcam-Rudloff, E.
<2>Complete genome sequence of a plant associated bacterium Bacillus amyloliquefaciens subsp. plantarum UCMB5033.
<3>Standards in Genomic Sciences
<4>9
<5>718-725
<6>2014
<7>Bacillus amyloliquefaciens subsp. plantarum UCMB5033 is of special interest for its ability to
promote host plant growth through production of stimulating
compounds and suppression of soil borne pathogens by synthesizing antibacterial
and antifungal metabolites or priming plant defense as induced systemic
resistance. The genome of B. amyloliquefaciens UCMB5033 comprises a 4,071,167 bp
long circular chromosome that consists of 3,912 protein-coding genes, 86 tRNA
genes and 10 rRNA operons.

<>

<1>Nicholson, T.L., Shore, S.M., Bayles, D.O., Register, K.B., Kingsley, R.A.
<2>Draft Genome Sequence of the Bordetella bronchiseptica Swine Isolate KM22.
<3>Genome Announcements
<4>2
<5>e00670-14
<6>2014
<7>Bordetella bronchiseptica swine isolate KM22 has been used in experimental infections of swine
as a model of clinical B. bronchiseptica infections within
swine herds and to study host-to-host transmission. Here we report the draft
genome sequence of KM22.

<>

<1>Nicholson, W.L., Leonard, M.T., Fajardo-Cavazos, P., Panayotova, N., Farmerie, W.G., Triplett, E.W., Schuerger, A.C.
<2>Complete Genome Sequence of Serratia liquefaciens Strain ATCC 27592.
<3>Genome Announcements
<4>1
<5>e00548-13
<6>2013
<7>We report the complete genome sequence of Serratia liquefaciens strain ATCC 27592, which was
previously identified as capable of growth under low-pressure
conditions. To the best of our knowledge, this is the first announcement of the
complete genome sequence of an S. liquefaciens strain.

<>

<1>Nickoloff, J.A.
<2>Converting restriction sites by filling in 5' extensions.
<3>Biotechniques
<4>12
<5>512-514
<6>1992
<7>None

<>

<1>Nickoloff, J.A., Chen, E.Y., Heffron, F.
<2>A 23-base-pair DNA sequence from the MAT locus stimulates intergenic recombination in yeast.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>83
<5>7831-7835
<6>1986
<7>HO nuclease is a site-specific double-strand endonuclease present in haploid Saccharomyces
cerevisiae undergoing mating type interconversion. HO nuclease initiates mating type
interconversion by making a double-strand break within the MAT locus. To define the
recognition site for the enzyme in vitro, we have constructed a number of point mutations and
deletions within or adjacent to the HO recognition site. Digestion of these substrates with HO
in vitro reveals that the minimal recognition site is 18 base pairs long, although several
shorter substrates and substrates containing point mutations are cleaved at low levels in
vitro. A 24-base-pair HO recognition site stimulates homologous recombination when present in
a region unrelated to MAT. Recombinants arise from both gene conversion and crossover events.
The identification of the HO recognition site provides a way of introducing a defined
initiation site for recombination.

<>

<1>Nicolskaya-Sanovich, I.I., Uporova, T.M., Gromova, E.S., Levchenko, I.Y., Arutyunyan, E.E., Gruber, I.M., Kubareva, E.A.
<2>SsoII restrictase producing process - restriction endonuclease production by Escherichia coli B834 carrying plasmid d24.
<3>Soviet Patent Office
<4>SU 1822877
<5>
<6>1993
<7>
<>

<1>Nie, H., Yang, F., Zhang, X., Yang, J., Chen, L., Wang, J., Xiong, Z., Peng, J., Sun, L., Dong, J., Xue, Y., Xu, X., Chen, S., Yao, Z., Shen, Y., Jin, Q.
<2>Complete genome sequence of Shigella flexneri 5b and comparison with Shigella flexneri 2a.
<3>BMC Genomics
<4>7
<5>173
<6>2006
<7>ABSTRACT: BACKGROUND: Shigella bacteria cause dysentery, which remains a significant threat to
public health. Shigella flexneri is the most common
species in both developing and developed countries. Five Shigella genomes
have been sequenced, revealing dynamic and diverse features. To
investigate the intra-species diversity of S. flexneri genomes further, we
have sequenced the complete genome of S. flexneri 5b strain 8401
(abbreviated Sf8401) and compared it with S. flexneri 2a (Sf301). RESULTS:
The Sf8401 chromosome is 4.5-Mb in size, a little smaller than that of
Sf301, mainly because the former lacks the SHI-1 pathogenicity island
(PAI). Compared with Sf301, there are 6 inversions and one translocation
in Sf8401, which are probably mediated by insertion sequences (IS). There
are clear differences in the known PAIs between these two genomes. The
bacteriophage SfV segment remaining in SHI-O of Sf8401 is clearly larger
than the remnants of bacteriophage SfII in Sf301. SHI-1 is absent from
Sf8401 but a specific related protein is found next to the pheV locus.
SHI-2 is involved in one intra-replichore inversion near the origin of
replication, which may change the expression of iut/iuc genes. Moreover,
genes related to the glycine-betaine biosynthesis pathway are present only
in Sf8401 among the known Shigella genomes. CONCLUSIONS: Our data show
that the two S. flexneri genomes are very similar, which suggests a high
level of structural and functional conservation between the two serotypes.
The differences reflect different selection pressures during evolution.
The ancestor of S. flexneri probably acquired SHI-1 and SHI-2 before SHI-O
was integrated and the serotypes diverged. SHI-1 was subsequently deleted
from the S. flexneri 5b genome by recombination, but stabilized in the S.
flexneri 2a genome. These events may have contributed to the differences
in pathogenicity and epidemicity between the two serotypes of S. flexneri.

<>

<1>Nielsen, H., Einvik, C., Lentz, T.E., Hedegaard, M.M., Johansen, S.D.
<2>A conformational switch in the DiGIR1 ribozyme involved in release and folding of the downstream I-DirI mRNA.
<3>RNA
<4>15
<5>958-967
<6>2009
<7>DiGIR1 is a group I-like cleavage ribozyme found as a structural domain within a nuclear
twin-ribozyme group I intron. DiGIR1 catalyzes
cleavage by branching at an Internal Processing Site (IPS) leading to
formation of a lariat cap at the 5'-end of the 3'-cleavage product. The
3'-cleavage product is subsequently processed into an mRNA encoding a
homing endonuclease. By analysis of combinations of 5'- and
3'-deletions, we identify a hairpin in the 5'-UTR of the mRNA (HEG P1)
that is formed by conformational switching following cleavage. The
formation of HEG P1 inhibits the reversal of the branching reaction,
thus giving it directionality. Furthermore, the release of the mRNA is
a consequence of branching rather than hydrolytic cleavage. A model is
put forward that explains the release of the I-DirI mRNA with a lariat
cap and a structured 5'-UTR as a direct consequence of the DiGIR1
branching reaction. The role of HEG P1 in GIR1 branching is reminiscent
of that of hairpin P-1 in splicing of the Tetrahymena rRNA group I
intron and illustrates a general principle in RNA-directed RNA
processing.

<>

<1>Nielsen, H., Westhof, E., Johansen, S.
<2>An mRNA is capped by a 2', 5' lariat catalyzed by a group I-like ribozyme.
<3>Science
<4>309
<5>1584-1587
<6>2005
<7>Twin-ribozyme introns are formed by two ribozymes belonging to the group I family and occur in
some ribosomal RNA transcripts. The group I-like
ribozyme, GIR1, liberates the 5' end of a homing endonuclease messenger
RNA in the slime mold Didymium iridis. We demonstrate that this cleavage
occurs by a transesterification reaction with the joining of the first and
the third nucleotide of the messenger by a 2',5'-phosphodiester linkage.
Thus, a group I-like ribozyme catalyzes an RNA branching reaction similar
to the first step of splicing in group II introns and spliceosomal
introns. The resulting short lariat, by forming a protective 5' cap, might
have been useful in a primitive RNA world.

<>

<1>Nielsen, P.E., Egholm, M., Berg, R.H., Buchardt, O.
<2>Sequence-selective recognition of DNA by strand displacement with a Thymine-substituted polyamide.
<3>Science
<4>254
<5>1497-1500
<6>1991
<7>A polyamide nucleic acid (PNA) was designed by detaching the deoxyribose
phosphate backbone of DNA in a computer model and replacing it with an achiral
polyamide backbone.  On the basis of this model, oligomers consisting of
thymine-linked aminoethylglycyl units were prepared.  These oligomers recognize
their complementary target in double-stranded DNA by strand displacement.  The
displacement is made possible by the extraordinarily high stability of the
PNA-DNA hybrids.  The results show that the backbone of DNA can be replaced by
a polyamide, with the resulting oligomer retaining base-specificity
hybridization.

<>

<1>Nielsen, P.E., Egholm, M., Berg, R.H., Buchardt, O.
<2>Sequence specific inhibition of DNA restriction enzyme cleavage by PNA.
<3>Nucleic Acids Res.
<4>21
<5>197-200
<6>1993
<7>Plasmids containing double-stranded 10-mer PNA (peptide nucleic acid chimera) targets
proximally flanked by two restriction enzyme sites were challenged with the complementary PNA
or PNAs having one or two mismatches, and the effect on the restriction enzyme cleavage of the
flanking sites was assayed. The following PNAs were used: T10-LysNH2, T5CT4-LysNH2 and
T2CT2CT4-LysNH2 and the corresponding targets cloned into pUC19 were flanked by BamHI, SalI or
PstI sites, respectively. In all cases it was found that complete inhibition of restriction
enzyme cleavage was obtained with the complementary PNA, a significantly reduced effect was
seen with a PNA having one mismatch, and no effect was seen with a PNA having two mismatches.
These results show that PNA can be used as sequence specific blockers of DNA recognizing
proteins.

<>

<1>Nierman, W.C. et al.
<2>Complete genome sequence of Caulobacter crescentus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>4136-4141
<6>2001
<7>The complete genome sequence of Caulobacter crescentus was determined to be 4,016,942 base
pairs in a single circular chromosome encoding 3,767 genes. This organism, which grows in a
dilute aquatic environment, coordinates the cell division cycle and multiple cell
differentiation events. With the annotated genome sequence, a full description of the genetic
network that controls bacterial differentiation, cell growth, and cell cycle progression is
within reach. Two-component signal transduction proteins are known to play a significant role
in cell cycle progression. Genome analysis revealed that the C. crescentus genome encodes a
significantly higher number of these signaling proteins (105) than any bacterial genome
sequenced thus far. Another regulatory mechanism involved in cell cycle progression is DNA
methylation. The occurrence of the recognition sequence for an essential DNA methylating
enzyme that is required for cell cycle regulation is severely limited and shows a bias to
intergenic regions. The genome contains multiple clusters of genes encoding proteins essential
for survival in a nutrient poor habitat. Included are those involved in chemotaxis, outer
membrane channel function, degradation of aromatic ring compounds, and the breakdown of
plant-derived carbon sources, in addition to many extracytoplasmic function sigma factors,
providing the organism with the ability to respond to a wide range of environmental
fluctuations. C. crescentus is, to our knowledge, the first free-living alpha-class
proteobacterium to be sequenced and will serve as a foundation for exploring the biology of
this group of bacteria, which includes the obligate endosymbiont and human pathogen Rickettsia
prowazekii, the plant pathogen Agrobacterium tumefaciens, and the bovine and human pathogen
Brucella abortus.

<>

<1>Nierman, W.C. et al.
<2>Structural flexibility in the Burkholderia mallei genome.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>14246-14251
<6>2004
<7>The complete genome sequence of Burkholderia mallei ATCC 23344 provides
insight into this highly infectious bacterium's pathogenicity and
evolutionary history. B. mallei, the etiologic agent of glanders, has come
under renewed scientific investigation as a result of recent concerns
about its past and potential future use as a biological weapon. Genome
analysis identified a number of putative virulence factors whose function
was supported by comparative genome hybridization and expression profiling
of the bacterium in hamster liver in vivo. The genome contains numerous
insertion sequence elements that have mediated extensive deletions and
rearrangements of the genome relative to Burkholderia pseudomallei. The
genome also contains a vast number (>12,000) of simple sequence repeats.
Variation in simple sequence repeats in key genes can provide a mechanism
for generating antigenic variation that may account for the mammalian
host's inability to mount a durable adaptive immune response to a B.
mallei infection.

<>

<1>Niharika, N., Sangwan, N., Ahmad, S., Singh, P., Khurana, J.P., Lal, R.
<2>Draft Genome Sequence of Sphingobium chinhatense Strain IP26T, Isolated from a Hexachlorocyclohexane Dumpsite.
<3>Genome Announcements
<4>1
<5>e00680-13
<6>2013
<7>Sphingobium chinhatense strain IP26(T) is a conducive hexachlorocyclohexane (HCH) degrader
isolated from a heavily contaminated (450 mg HCH/g soil) HCH dumpsite.
IP26(T) degrades alpha-, beta-, gamma-, and delta-HCH, which are highly
persistent in the environment. Here we report the draft genome sequence (~5.8
Mbp) of this strain.

<>

<1>Nikiforov, T.T., Connolly, B.A.
<2>Oligodeoxynucleotides containing 4-thiothymidine and 6-thiodeoxyguanosine as affinity labels for the Eco RV restriction endonuclease and modification methylase.
<3>Nucleic Acids Res.
<4>20
<5>1209-1214
<6>1992
<7>4-Thiothymidine and 6-thiodeoxyguanosine were incorporated into synthetic
dodecamers containing the recognition site d(GATATC) of the enzymes EcoRV
endonuclease and EcoRV methyltransferase.  Upon irradiation with long
wavelength UV light (340- 360 nm), these oligodeoxynucleotides were
photochemically crosslinked to both enzymes.  The yields were up to 35% with
the methyltransferase, but lower (up to 6%) with the endonuclease.
Oligodeoxynucleotides containing 4-thiothymidine generally gave higher yields
of crosslinking than those containing 6-thiodeoxyguanosine.  Although both
specific (i.e. those containing the d(GATATC) sequence) and non-specific
(lacking this sequence) photoreactive oligodeoxynucleotides gave rise to
crosslinked products, the use of a non-reactive, competitive substrate
oligodeoxynucleotide suppressed the crosslinking, indicating that the reaction
takes place at the enzymes' active sites.  Oligodeoxynucleotides containing
4-thiocyanatothymidine or 6-thiocyanatodeoxyguanosine were also prepared by
treatment of the title oligomers with CNBr and KCN.  The dodecamers containing
4-thiocyanatothymidine were found to covalently modify both enzymes under
study, with levels of crosslinking reaching up to 42% with the endonuclease and
up to 12% with the methyltransferase.  No crosslinking was observed with
oligodeoxynucleotides containing 6-thiocyanatodeoxyguanosine.

<>

<1>Nikitin, D., Mokrishcheva, M., Denjmukhametov, M., Pertzev, A., Zakharova, M., Solonin, A.
<2>Construction of an overproducing strain, purification, and biochemical characterization of the 6His-Eco29kI restriction endonuclease.
<3>Protein Expr. Purif.
<4>30
<5>26-31
<6>2003
<7>We constructed a strain of Escherichia coli overproducing 6His-tagged Eco29kI by placing the
coding sequence under control of a strong
bacteriophage T5 promoter. The yield of 6His-Eco29kI restriction
endonuclease expression could be increased to about 20% of the total
cellular protein, but inclusion bodies formed consisting of insoluble
6His-Eco29kI protein. We developed a fast and effective protocol for
purification of the homogeneous enzyme from both soluble and insoluble
fractions and established their identity by catalytic activity assay. The
isolated enzymes were tested for recognition specificity and optimal
reaction conditions as a function of NaCl and KCl concentrations,
temperature, and pH compared with the native Eco29kI restriction
endonuclease. The 6His-tagged enzyme retained the specificity of the
native protein but had an altered optimum of its catalytic reaction.

<>

<1>Nikitin, D., Mokrishcheva, M., Solonin, A.
<2>6His-Eco29kI methyltransferase methylation site and kinetic mechanism characterization.
<3>Biochim. Biophys. Acta
<4>1774
<5>1014-1019
<6>2007
<7>A new type 11 6His-Eco29kI DNA methyltransferase was tested for methylation site (CC(Me)GCGG)
and catalytic reaction optimal
conditions. With high substrate concentrations, an inhibitory effect of
DNA, but not AdoMet, on its activity was observed. Isotope partitioning
and substrate preincubation assays showed that the enzyme-AdoMet
complex is catalytically active. Considering effect of different
concentrations of DNA and AdoMet on initial velocity, ping-pong
mechanisms were ruled out. According to data obtained, the enzyme
appears to work by preferred ordered bi-bi mechanism with AdoMet as
leading substrate.

<>

<1>Nikitin, D.V., Mokrishcheva, M.L., Solonin, A.S.
<2>Binding of DNA methyltransferase M.Ecl18 to operator-promoter region decreases its methylating activity.
<3>Biokhimiia
<4>77
<5>392-397
<6>2012
<7>The type II bifunctional DNA methyltransferase (MTase) Ecl18 that is able to control
transcription of its own gene was studied kinetically. Based on initial velocity dependences
from S-adenosyl-L-methionine (AdoMet) and target DNA and substrate preincubation assays, it is
proposed that the enzyme apparently works by a rapid equilibrium ordered bi-bi mechanism with
DNA binding first. By measuring the enzyme activity depending on DNA and AdoMet at different
fixed concentrations of the operator sequence oligonucleotide, it was found that its binding
has noncompetitive inhibitory effect on Ecl18 MTase activity.

<>

<1>Nikitin, D.V., Mokrishcheva, M.L., Solonin, A.S.
<2>Binding of DNA methyltransferase M.Ecl18 to operator-promoter region decreases its methylating activity.
<3>Biochemistry
<4>77
<5>307-311
<6>2012
<7>The type II bifunctional DNA methyltransferase (MTase) Ecl18 that is able to control
transcription of its own gene was studied kinetically.
Based on initial velocity dependences from S-adenosyl-L-methionine
(AdoMet) and target DNA and substrate preincubation assays, it is
proposed that the enzyme apparently works by a rapid equilibrium
ordered bi-bi mechanism with DNA binding first. By measuring the enzyme
activity depending on DNA and AdoMet at different fixed concentrations
of the operator sequence oligonucleotide, it was found that its binding
has noncompetitive inhibitory effect on Ecl18 MTase activity.

<>

<1>Nikolaichik, Y., Gorshkov, V., Gogolev, Y., Valentovich, L., Evtushenkov, A.
<2>Genome Sequence of Pectobacterium atrosepticum Strain 21A.
<3>Genome Announcements
<4>2
<5>e00935-14
<6>2014
<7>We report the annotated genome sequence of the enterobacterial plant pathogen Pectobacterium
atrosepticum strain 21A, isolated in Belarus from potato stem with
blackleg symptoms.

<>

<1>Nikolajewa, S., Byer, A., Friedel, M., Hollunder, J., Wilhelm, T.
<2>Common patterns in type II restriction enzyme binding sites.
<3>Nucleic Acids Res.
<4>33
<5>2726-2733
<6>2005
<7>Restriction enzymes are among the best studied examples of DNA binding proteins. In order to
find general patterns in DNA recognition sites, which may reflect important properties of
protein-DNA interaction, we analyse the binding sites of all known type II restriction
endonucleases. We find a significantly enhanced GC content and discuss three explanations for
this phenomenon. Moreover, we study patterns of nucleotide order in recognition sites. Our
analysis reveals a striking accumulation of adjacent purines (R) or pyrimidines (Y). We
discuss three possible reasons: RR/YY dinucleotides are characterized by (i) stronger H-bond
donor and acceptor clusters, (ii) specific geometrical properties and (iii) a low stacking
energy. These features make RR/YY steps particularly accessible for specific protein-DNA
interactions. Finally, we show that the recognition sites of type II restriction enzymes are
underrepresented in host genomes and in phage genomes.

<>

<1>Nikolskaya, I., Lopatina, N., Suchkov, S., Kartashova, I., Debov, S.
<2>Sequence Specificity of Isolated DNA-Cytosine Methylases from Shigella Sonnei 47 Cells.
<3>Biochem. Int.
<4>9
<5>771-781
<6>1984
<7>Five individual DNA-cytosine methylases differing in pI (isoelectric point) values are present
in Shigella sonnei 47 cells.  The sequence specificity of each of those was determined 'in
vitro' by a highly efficient combined approach that included pyrimidine tract (isostic)
analysis, identification of the immediate neighborhood of the methylated base within the
recognition sequence and the calculation method.  The enzyme with pI 5.3 (Msso 5,3) is the
counterpart of the RSso 47II in the Sso 47II restriction-modification system and methylates
the internal cytosine residue of the 'palindromic' 5'-C-C-N-G-G-3' sequence.  The enzymes
with pI 6.2 (Msso 6,2) and 7.4 (MSso 7,4) exhibit identical specificity upon methylation of
the 'palindromic' 5'-Py-C-N-G-Pu-3' sequence, but differ in the pI values of the proteins.
The enzyme with pI 4.2 (MSso 4,2) recognizes the unique tetranucleotide 5'-C-C-C-C-3'
sequence and methylates the second cytosine residue at the 5'-end of the sequence.  The
enzyme with pI 8.4 (MSso 8,4) methylates the centrol cytosine residue within the degenerative
trinucleotide 5'-(Pu)-C-C-3' sequence. MSso5,3', MSso6,2', and MSso7,4 are presumed to
belong to the 'family' of sequence-specific (EcoRII-like) enzymes. These DNA-cytosine
methylases are likely to be evolutionarily related to EcoRII and to have undergone a
sufficient genetic drift so as to recognize similar (but more degenerative) nucleotide
sequences.

<>

<1>Nikolskaya, I., Tediashvili, M., Lopatina, N., Chanishvili, T.G., Debov, S.
<2>Specificity and functions of guanine methylase of Shigella sonnei DDVI phage.
<3>Biochim. Biophys. Acta
<4>561
<5>232-239
<6>1979
<7>DNA methylase methylating adenine with formation of 6-methylamino-purine has
been identified in Shigella sonnei 1188 cells which are the natural host of
DDVI phage.  At the same time, in DNA of DDVI phage replicating both in Sh.
sonnei 1188 cells and Escherichia coli B cells 7-methylguanine was found as the
only minor base in amounts of 0.25 and 0.27 mol per 100 mos of nucleotides,
respectively.  The extract of the infected cells was found to contain both
kinds of DNA methylases; virus-specific guanine methylase and cellular adenine
methylase.  The place of 6-methylaminopurine in DNA of this phage is explained
by reversible inhibition of the cell enzyme in the infected cells.  The amount
of methyl groups transferred by DDVI-specific methylase on DNA does not depend
on the specifics of the infected cells and is similar in the case of unmodified
SD phage DNA and DNA of T2 phase methylated by E. coli B enzyme.  Guanine
methylase has been shown to be a DDVI-induced modification enzyme and to
protect against restriction of B-type. It methylates double-stranded DNAs only
and is inhibited by S-adenosylhomocysteine.

<>

<1>Nikolskaya, I.I., Aleksandrova, S.S., Lopatina, N.G., Debov, S.S.
<2>Fractionation and purification of DNA methylases and specific endonucleases from cells of Escherichia coli SK.
<3>Biokhimiia
<4>42
<5>598-608
<6>1977
<7>Fractionation and purification of DNA methylases and specific endonucleases from cells of
Escherichia coli SK responsible for DNA specificity to host prokaryotic cells were studied.
The most efficient purification was achieved by precipitation of proteins by 60% saturated
ammonium sulfate with subsequent chromatography on CM-cellulose and concentration of fractions
by dialysis against glycerol.  Under these conditions the methylase activity produced 4
discrete fractions.  Due to purification the specific activity of methylases increased
11-20-fold in various fractions.  Methylase from the first (A) and fourth (BII) peaks
catalyzed the methylation of cytosine to produce 5-methylcytosine; methylase from the third
peak (BI) methylated adenine to produce 6-methylaminopurine.  The chemical specificity of the
second peak (B) methylase could not be established due to very high lability of the enzyme in
this fraction.  Specific endonuclease was found in the gradient zones eluted by 0.1-0.2M and
0.65-0.75M NaCl.   It is assumed that those enzymes providing for DNA hydrolysis up to the
formation of high-molecular discrete fragments, are restriction endonucleases of the SK
system.  The results obtained strongly suggest the existence of several types of methylases
and restricting endonucleases in E. coli SK cells.

<>

<1>Nikolskaya, I.I., Aleksandrova, S.S., Lopatina, N.G., Debov, S.S.
<2>Fractionation and specificity of DNA methylases from the Escherichia coli SK cells.
<3>Mol. Cell. Biochem.
<4>20
<5>17-24
<6>1978
<7>Specificity of DNA methylation enzymes from the E. coli SK cells and conditions for their
separation have been investigated.  Column chromatography on carboxymethylcellulose permits
fractionation of methylase activity into six discrete peaks whose specificity to the
methylated base has been determined in vitro with H3-SAM as precursor.  All methylases
specific for adenine produced 6'-methylaminopurine, all methylases specific for cytosine
yielded 5'-methylcytosine.  The first enzymatic activity peak containing cytosine methylase
free of traces of adenine-methylating activity (E1), and the second peak containing both the
enzymes (E2) were not adsorbed on the ion exchanger and went off the column with the effluent
(column buffer).  Adenine specific methylase E2 is retarded to a small extent during the
passage through the column.  The second adenine methylase (W) was characterized by weak bonds
with the ion exchanger and was removed when washing the column with column buffer.  The
elution with NaCl gradient produced successively three enzymatic activity peaks: adenine
methylase (GI), cytosine methylase (GII), and one more adenine methylase (GIII) removed from
the column by 0.16M, 0.24M and 0.43M NaCl respectively.  Using a new modification of the
complementary methylation test, the specificity with regard to recognition site was examined
for all enzymes, except for W and GIII, which were extremely unstable.  The adenine methylases
E2 and GI and the cytosine methylases E1 and GII were shown to recognize different sites and
to be different enzymes.  In view of the drastic differences in their chromatographic behavior
and physical stability, the adenine methylases W and GIII are probably also different enzymes.

<>

<1>Nikolskaya, I.I., Debov, S.S.
<2>Molecular and medical aspects of DNA modification.
<3>Vestn. Akad. Med. Nauk SSSR
<4>0
<5>23-29
<6>1987
<7>The role of methylation in the cell's life cycle is considered with special
reference to regulation of transcription in pro- and eukaryotes.  In
eukaryotes, an inverse correlation exists between levels of gene expression and
methylation, as is illustrated by experiments with eukaryotic viruses.  In the
case of bacteriophages, the expression of viral genes may be regulated through
methylation both negatively and positively.  The modifying role of methylation
enzymes in prokaryotes, based on the identity or overlap of recognition sites
in methylases and restriction endonucleses, is analyzed at length.

<>

<1>Nikolskaya, I.I., Karpetz, L.Z., Kartashova, M., Lopatina, N.G., Skripkin, E.A., Suchkov, S.V., Uporova, T.M., Gruber, I.M., Debov, S.S.
<2>Enzymes of the new system of the host specificity Sso47II.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>5-10
<6>1983
<7>The DNA host specificity systems SsoI and SsoII are present in Sh. sonnei 47 cells.  The
R.SsoII and M.SsoII were isolated from these cells and purified by means of affinity,
ion-exchange, hydrophobic chromatography and isoelectric focusing.  The fine structure of
R.SsoII and M.SsoII recognition sites was the following: ^CC*NGG.  The pI of M.SsoII was found
to be 5.3 as determined by isoelectric focusing.  M.SsoII transfers methyl groups to the inner
cytosine in the recognition sequence CC*NGG.  R.SsoII and M.SsoII are isoshiso- and
isomethymeric of EcoRII type correspondingly with the SsoII recognition sequence being more
degenerative.

<>

<1>Nikolskaya, I.I., Lopatina, N.G., Anikeicheva, N.V., Debov, S.S.
<2>Determination of the recognition sites of cytosine DNA-methylases from Escherichia coli SK.
<3>Nucleic Acids Res.
<4>7
<5>517-528
<6>1979
<7>Two different cytosine DNA-methylases, NI and GII, are present in Escherichia
coli SK.  The GII methylase recognizes the five-member symmetric sequence:
5'...NpCpCpApGpGpN...3'.  This sequence is identical with the recognition site
of the hsp II type determined by RII plasmid but, in contrast to RII methylase,
the GII enzyme methylates cytosine located on the 5' side of the site.  By
analogy with the isoshizomery of the restricting endonucleases, RII and GII DNA
methylaeses may be called isomethymers which recognize the same site but
methylate different bases.  Since the phage of the SK and hsp II phenotypes is
effectively restricted in respective cells it may be assumed that the
isomethymeric modification does not provide any protection against the
corresponding restrictases.  NI methylase recognizes the five-member symmetric
site which represents an inverted sequence of the GII site:
5'...NpGpGpApCpCpN...3'.  In this case cytosine at the 3'-end of the
recognition site is methylated.

<>

<1>Nikolskaya, I.I., Lopatina, N.G., Chaplygina, N.M., Debov, S.S.
<2>The host specificity system in Escherichia coli SK.
<3>Mol. Cell. Biochem.
<4>13
<5>79-87
<6>1976
<7>E. coli SK has its own enzyme system providing DNA host specificity which differs from the
known types of specificity in E. coli K12 and E. coli B.  Modification and restriction are
observed when the PBVI or PBV3 phages are transferred from E. coli SK to E. coli B or K12 (and
back).  A methylase has been isolated from E. coli SK cells and partly purified.  This
methylase catalyzes in vitro transfer of the labeled methyl groups from S-adenosylmethionine
to DNA of both phage and tissue origin which gives rise to 5'-methylcytosine and
6'-methylaminopurine.  The methylase preparations isolated from the cells at stationary phase
have proved to be 1.5-1.7 times as active as the enzyme from cells at the logarithmic growth
stage.  The extract of E. coli SK cells infected with the phage SD cannot methylate DNA in
vitro.  This fact is due to de novo synthesis of the enzyme which degrades SAM to
5'-methylthioadenosine and homoserine.  This enzyme is not found in cells infected with the
SD phage in the presence of chloroamphenicol.  The activity of the enzyme which degrades SAM
is highest between the 4th and the 5th minutes of infection.  Thus it may be assumed that this
enzyme, most probably, is an early virus specific protein and prevents in vivo methylation of
the phage DNA.

<>

<1>Nikolskaya, I.I., Lopatina, N.G., Debov, S.S.
<2>On heterogeneity of DNA methylases from Escherichia coli SK cells.
<3>Mol. Cell. Biochem.
<4>35
<5>3-10
<6>1981
<7>The presence in E. coli SK cells of five different DNA-methylases differing in specificity to
the methylated sequence is documented has been proven.  Two enzymes methylate cytosine with
the formation of 5'-methylcytosine and three enzymes methylate adenine with formation of
6'-methylaminopurine.  A method for simultaneous isolation of the five individual enzymes
including gel filtration on Biogel A-0.5 M is proposed.  The direct evidence has been
presented showing that the additional methylation test in our method modification actually can
discriminate between enzymes differing in sensitive sites.

<>

<1>Nikolskaya, I.I., Lopatina, N.G., Dedov, S.S.
<2>Some peculiarities of phage DDVI-specific methylases.
<3>Biokhimiia
<4>42
<5>828-832
<6>1977
<7>Two types of methylases are found in the cellular extract of Escherichia coli B, infected with
phage DDVI.  One of them is a cellular enzyme, which methylates adenine to form
6-methylaminopurine and is repressed in the infected cell in vivo.  The second type, which is
not found in the non-infected cell, is specific for phage DDVI and induces the formation of
7-methylguanine.  Both enzymes recognize various sites, which accounts for variation in the
ratio 6-MAP/7-MG in heterologous DNAs between 2.07 in phage Sd DNA and 0.40 in phage DDII DNA.
During in vitro incubation with homologous methylases phage DDVI DNA and especially phage T2
DNA are subjected to further methylation, which is probably indicative of their
undermethylation in vivo.  The DDVI-specific enzyme, similar to B-specific type, methylates
DNA with a normal set of nitrogenous bases (phages Sd and DDII0, as well as DNAs containing
5-oxymethylcytosine and glucose (phages T2 and DDVI).  Both methylases under study use only
native double-helical DNA as substrate and are strongly inhibited by S-adenosylhomocysteine.
Phage DDVI methylase is characterized by low stability.

<>

<1>Nikolskaya, I.I., Lopatina, N.G., Lopareva, E.N., Posypanova, A.M., Debov, S.S.
<2>Modifying methylase SsoI from Shigella sonnei 47 cells.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>10
<5>26-30
<6>1988
<7>A simple and fast method for isolation and purification of SsoI methylase from
the bacterial strain Shigella sonnei 47 has been proposed.  The enzyme is a
modifying component of the host cell specificity system and protects the
acceptor DNA from hydrolysis by restriction endonuclease SsoI and EcoRI.  The
method is based on hydrophobic chromatography of ammonium sulphate fraction on
phenylsepharose.  The enzyme preparation obtained is devoid of specific and
nonspecific endonucleases and is stable at storage in 30% glycerol during a
year.  The conditions of manifestation of "star" activity by the enzyme were
studied.

<>

<1>Nikolskaya, I.I., Lopatina, N.G., Rekunova, V.N., Yurkevich, A.M., Debov, S.S.
<2>On the effect of S-nucleosyl homocysteines on activity of several DNA methylases.
<3>Vopr. Med. Khim.
<4>24
<5>252-255
<6>1978
<7>The effect was studied of S-adenosyl, -uridyl, -cytidyl and -inosyl homocysteines on the
activity of bacterial adenine and cytosine methylases from E. coli CK as well as on guanine
methylase specific for DDVI phage.  S-adenosyl homocysteine was shown to be a strong inhibitor
of methylation; 10 microM of the substance inhibited all the enzymes studied by 98-99%.  Use
of total enzymatic preparations did not show any difference in the affinity of S-uridyl,
-cytidyl and -inosyl homocysteines to various DNA methylases studied.  All these preparations
inhibited DNA methylases by 55-65%.  Increase in the concentration of inhibitor up to 20
microM did not elevate the inhibitory effect.  Action of S-nucleosyl homocysteines did not
depend on the type of acceptor DNA.

<>

<1>Nikolskaya, I.I., Lopatina, N.G., Sharkova, E.V., Suchkov, S.V., Somodi, P., Foldes, I., Debov, S.S.
<2>Sequence specificity of isolated DNA-adenine methylases from Mycobacterium smegmatis (butyricum) and Shigella sonnei 47 cells.
<3>Biochem. Int.
<4>10
<5>405-413
<6>1985
<7>A set of four individual DNA-adenine methylases differing in pI (isoelectric point) values
(MMbu4.2', MMbu6.4', MMbu7.3' and MMbu8.7), and a sole methylating enzyme with the same
base specificity (MSso9.5) are present in M. smegmatis (butyricum) and S. sonnei 47 cells,
respectively.  The sequence specificity of each of those was studied in vitro by a combined
approach that comprised isostich (purine tract) analysis and identification of the immediate
neighbourhood of the methylated base within the sequence methylated.  The MSso9.5 recognition
site has been established as the hexanucleotide 'palindromic' 5'-G-A*-A-T-T-C-3' sequence
which is structurally similar to the analogous M.EcoRI recognition site.  However, in contrast
to M.EcoRI, MSso 9.5 methylates the 5'-end adenine residue in the sequence and thus it
appears to be an isometimer of M.EcoRI.  By means of the same approach, the partial nucleotide
sequences methylated by each of the four individual M. butyricum enzymes were determined.
MMbu7.3 and MMbu8.7 exhibit the identical sequence specificity upon methylation of the
degenerative trinucleotide 5'-Py-A*-Py-3' sequence and thus these enzymes are assumed to
represent the different molecular forms of the methylase.  MMbu4.2 methylates the
5'-G-G-A*-3' sequence and thus it is of a great value as the tool for negating effects of
the R.BamHI and R.AvaII-type restriction.  MMbu6.4 is of a particular interest on account of
its unique DNA methylation pattern which is distinguished in the pronounced clustering of
purine bases in the 5'-Pu-Pu-Pu-Pu-Pu-3' sequence methylated.

<>

<1>Nikolskaya, I.I., Sharkova, E.V., Suchkov, S.V., Karpets, L.Z., Debov, S.S., Somodi, P., Foldes, I.
<2>Isoelectric focusing of bacterial DNA methylases.
<3>Biochem. Int.
<4>7
<5>605-615
<6>1983
<7>The multiplicity of bacterial DNA methylases has been shown for new microorganisms,
Mycobacteria and Shigella, by a double-step procedure including column chromatography followed
by isoelectric focusing of the total methylase fraction.  The profiles of the DNA methylating
activity of Sh.sonnei 47 and M. butyricum strains were studied.  Sh. Sonnei 47 cells were
found to contain five different proteins responsible for DNA methylation and having pI 4.2,
5.3, 6.2, 8.4 and 9.2.  Four M. butyricum methylases were represented by proteins with pI 4.2,
6.0, 8.0 and 9.0.

<>

<1>Nikolskaya, I.I., Sharkova, E.V., Suchkov, S.V., Lopatina, N.G., Habar, K., Somodi, P., Foldes, I., Debov, S.S.
<2>Factors of activation and stabilization of DNA-methylases from Shigella Sonnei 47 and Mycobacterium smegmatis butyricum cells.
<3>Biochem. Int.
<4>15
<5>127-138
<6>1987
<7>A comparative study of factors of activation and stabilization of individual
DNA-methylases from two bacterial strains - Shigella sonnei 47 and
Mycobacterium smegmatis butyricum - isolated by isoelectrofocusing in a pH
gradient has been carried out.  Storage of enzymes at +4C (pH 7.5) is
accompanied by periodic changes in the methylating activity.  No such changes
are observed when the enzymes are stored at pI of the protein.  In this case
the methylases with alkaline or close to neutral values of pI remain stable
over a period of at least two weeks, whereas acidic proteins are irreversibly
inactivated by the end of a two-week period.  A stabilizing effect of BSA on
DNA-methylases of Sso47 and Mbu strains has been demonstrated.  A direct
correlation between the stabilizing effect of BSA and the degree of enzyme
purity has been established.  Ca2+ appear to be a universal activator of
methylases of the above strains; these cations produce a potent, although a
short-term effect and can be used in the production of enzyme preparations with
a high specific activity in DNA recombinant technology.  Protease inhibitors do
not exert any appreciable effect on the methylase activity upon storage.
Storage at -20C and at neutral pH leads to complete inactivation of all
DNA-methylases within 24 hours.  In this case glycerol is fairly ineffective as
a stabiliziing agent.

<>

<1>Nikolskaya, I.I., Tediashvili, M.G., Vasileva, M.B., Chanishvili, T.G., Debov, S.S.
<2>System of host specificity and the DNA methylases of Shigellae and their phages.
<3>Vopr. Virusol.
<4>6
<5>724-731
<6>1978
<7>In Shigella sonnei cells there is a host DNA specificity system responsible for modification
and restriction of DDII phage.  DNA methylase from Shigella stutzeri cells is specific for
adenine and catalyses the appearance of 6'-methylaminopurine in the acceptory DNA.
Methylases from Shigella sonnei cells are specific for adenine and cytosine and provide for
the presence of 6'-methylaminopurine and 5'-methylcytosine in DNA.  The modifying activity
of these cells may be equally likely associated with both the enzymes.  A simplified version
of the additional methylation test has been developed for the study of enzyme specificity.
The results of additional and cross methylation suggest that several adenine methylases are
present in the cells of these Shigella, one of these enzymes being shared by Shigella stutzeri
and Shigella sonnei.  The DNA's isolated from Shigella sonnei and Shigella stutzeri cells are
undermethylated and in vitro undergo additional methylation upon incubation with the
appropriate enzyme.

<>

<1>Nikolskaya, I.I., Tediashvili, M.I., Chanishvili, T.G., Debov, S.S.
<2>Investigation of methylation character and DNA methylases specificity in Shigella.
<3>Biokhimiia
<4>43
<5>1228-1232
<6>1978
<7>The nature and content of minor bases in DNA of 3 Shigella strains are investigated.  DNA's
from Shigella stutzeri 2, S. sonnei 1188 and S. sonnei 311 are found to contain 0.43, 0.56 and
0.45 mol. % of N6-methyladenine respectively.  5-methylcytosine (0.16%) is discovered in S.
sonnei 311.  Substrate specificity of adenine methylase from S. sonnei 1188 with respect to
phage DNA's of different host modification is investigated.  Recognition sites for guanine
methylase of DDVI phage and for adenine methylase of S. sonnei 1188 turned out to be
different.  DNA of DDII phage grown in S. stutzeri 2 cells does not accept methyl groups under
the treatment with S. sonnei 1188 extracts, but it is methylated by Escherichia coli extract.
Adenine methylases of S. sonnei 1188 and S. stutzeri 2 are suggested to be either the same
enzyme, or enzymes, which recognition sites are partially overlapped.

<>

<1>Nikolskaya, I.I., Uporova, T.M., Tereshina, E.V., Gruber, I.M., Zhdanova, L.G., Debov, S.S.
<2>Specificity and properties of DNA methylases in Shigella.
<3>Vestn. Akad. Med. Nauk SSSR
<4>2
<5>16-21
<6>1981
<7>The specificity of and several properties of whole DNA methylase preparations from two
Shigella strains, S. sonnei 47843 and S. flexneri, were studied.  Both kinds of preparation
contained adenine and cytosine methylases which led to the presence of 6-methylaminopurine and
5-methylcytosine in the bacterial DNAs.  The concentrations of these in the S. sonnei DNA were
0.47 and 0.43 mole per 100 moles of nucleotides, respectively; the corresponding figures for
the S. flexneri DNAs were 0.42 and 0.38.  In vitro study of substrate specificity showed both
DNAs to be undermethylated in vivo and to be sensitive substrates for their "own" enzyme.  The
best acceptors for methyl groups were the thymus and phage Cd DNAs.  The denatured DNA
specimens did not undergo methylation.  The methylase activities of both strains were
inhibited by 95-97% by S-adenosylhomocysteine present in an amount of 10 micromoles.

<>

<1>Nikolskaya, I.I., Vanyushin, B.F., Mardashev, S.R.
<2>Identification of DNA methylases in cells of Escherichia coli CK.
<3>Biokhimiia
<4>40
<5>1081-1086
<6>1975
<7>It was established that E. coli CK cells contain a methylase which catalyzes
the incorporation of CH3 groups into tissue and phage DNAs in vitro in the
presence of the methyl group donor S-adenosyl-L-methionine.  During isolation
and purification the enzyme was found in the fraction of 30-60% saturation by
ammonium sulfate and its specific activity increased 1.9-fold.  The methylase
was active in both phosphate and tris-HCl buffers at pH 6.5-7.5 and did not
require Mg ions, EDTA, or dithiothreitol.  The enzyme recognized specific
nucleotide sequences in all the DNAs investigated and had broader substrate
specificity than the analogous enzyme from E. coli B.  The methylase of E. coli
CK was most active with thymus DNA.  The methylase from rat liver nuclei was
inactive in relation to the DNA of bacteriophages.

<>

<1>Nikolskaya-Sanovich, I.I., Arutyunyan, E.E., Gonchar, N.A., Gruber, I.M., Levchenko, I.Y.
<2>Purification of restriction endonuclease Sau 6782.
<3>Soviet Patent Office
<4>SU 1796676
<5>
<6>1993
<7>
<>

<1>Nilsson, M.-G., Skarped, C., Magnusson, G.
<2>Structure at restriction endonuclease MboI cleavage sites protected by actinomycin D or distamycin A.
<3>FEBS Lett.
<4>145
<5>360-364
<6>1982
<7>Restriction endonuclease MboI cleavage of DNA was inhibited by actinomycin D
and distamycin A.  The two inhibitors protected different subsets of the 8
cleavage sites in polyoma DNA.  The cleavage reactions were analyzed both in
the presence of minimal inhibitory concentrations of the compounds and at
higher concentrations, allowing cleavage at only 1 site/DNA molecule.  The
experiments showed that cleavage sites most efficiently protected by
actinomycin D had putative inhibitor binding sites as a distance of 1-2 base
pairs from the MboI recognition sequence.  Distamycin A, in contrast,
apparently has to bind immediately adjacent to the MboI recognition sequence to
protect from cleavage.

<>

<1>Nishi, S., Tsubouchi, T., Takaki, Y., Koyanagi, R., Satoh, N., Maruyama, T., Hatada, Y.
<2>Draft Genome Sequence of Loktanella cinnabarina LL-001T, Isolated from Deep-Sea Floor Sediment.
<3>Genome Announcements
<4>1
<5>e00927-13
<6>2013
<7>This report describes the draft genome sequence of Loktanella cinnabarina LL-001(T), which was
the first isolated strain from deep-sea floor sediment of
the genus Loktanella. The draft genome sequence contains 3,896,245 bp, with a G+C
content of 66.7%.

<>

<1>Nishibe, C., Canevari, C.A.B., Dalla, C.R., Pinto, B.J., Varuzza, L., Cataldi, A.A., Bernardelli, A., Bigi, F., Blanco, F.C., Zumarraga, M.J., Almeida, N.F., Araujo, F.R.
<2>Draft Genome Sequence of Mycobacterium bovis 04-303, a Highly Virulent Strain from Argentina.
<3>Genome Announcements
<4>1
<5>e00931-13
<6>2013
<7>Mycobacterium bovis strain 04-303 was isolated from a wild boar living in a free-ranging field
in Argentina. This work reports the draft genome sequence of
this highly virulent strain and the genomic comparison of its major
virulence-related genes with those of M. bovis strain AF2122/97 and Mycobacterium
tuberculosis strain H37Rv.

<>

<1>Nishida, H.
<2>Genome DNA Sequence Variation, Evolution, and Function in Bacteria and Archaea.
<3>Curr. Issues Mol. Biol.
<4>15
<5>19-24
<6>2013
<7>Comparative genomics has revealed that variations in bacterial and archaeal genome DNA
sequences cannot be explained by only neutral mutations. Virus resistance and plasmid
distribution systems have resulted in changes in bacterial and archaeal genome sequences
during evolution. The restriction-modification system, a virus resistance system, leads to
avoidance of palindromic DNA sequences in genomes. Clustered, regularly interspaced, short
palindromic repeats (CRISPRs) found in genomes represent yet another virus resistance system.
Comparative genomics has shown that bacteria and archaea have failed to gain any DNA with GC
content higher than the GC content of their chromosomes. Thus, horizontally transferred DNA
regions have lower GC content than the host chromosomal DNA does. Some nucleoid-associated
proteins bind DNA regions with low GC content and inhibit the expression of genes contained in
those regions. This form of gene repression is another type of virus resistance system. On the
other hand, bacteria and archaea have used plasmids to gain additional genes. Virus resistance
systems influence plasmid distribution. Interestingly, the restriction-modification system and
nucleoid-associated protein genes have been distributed via plasmids. Thus, GC content and
genomic signatures do not reflect bacterial and archaeal evolutionary relationships.

<>

<1>Nishigaki, K., Kaneko, Y., Wakuda, H., Husimi, Y., Tanaka, T.
<2>Type II restriction endonucleases cleave single-stranded DNAs in general.
<3>Nucleic Acids Res.
<4>13
<5>5747-5759
<6>1985
<7>Restriction endonucleases (13 out of 18 species used for the test) were
certified to cleave single-stranded(ss)DNA.  Such enzymes as AvaII, HaeII,
DdeI, AluI, Sau3AI, AccII, TthHB81 and HapII were newly reported to cleave
ssDNA.  A model to account for the cleavage of ssDNA by restriction enzyes was
proposed with supportive data.  The essential part of the model was that
restriction enzymes preferentially cleave transiently formed secondary
structures (called canonical structures) in ssDNA composed of two recognition
sequences with two fold rotational symmetry.  This means that a restriction
enzyme can cleave ssDNAs in general so far as the DNAs have the sequences of
restriction sites for the enzyme, and that the rate of cleavage depends on the
stabilities of canonical structures.

<>

<1>Nishimura, Y., Tsuboi, M.
<2>A possible correlation between DNA conformation and the mode of action of restriction enzymes.
<3>J. Biochem. (Tokyo)
<4>96
<5>1807-1811
<6>1984
<7>The cutting modes of restriction endonucleases which recognize
tetradeoxyribonucleotide sequences are classified into two groups.  d(GGCC) and
d(CGCG), for example, are cut to produce blunt ends, while d(CCGG) and d(GCGC)
are cut to produce two-base-long cohesive ends.  A conformational analysis by
the Calladine-Dickerson method indicates that d(GGCC) and d(CGCG) should have a
roll angle of successive base-pairs open towards the major groove at the
central (second) base-pair step.  On the other hand, d(CCGG) and d(GCGC) have
such open roll angles at the first and third base-pair steps.  It is suggested
that, in general, the cutting mode of a tetramer-specific enzyme depends
primarily upon the substrate conformation, rather than upon the enzyme.
Similar correlations between the mode of action and substrate conformation are
also suggested for hexamer-specific enzymes.

<>

<1>Nishio, Y., Nakamura, Y., Kawarabayasi, Y., Usuda, Y., Kimura, E., Sugimoto, S., Matsui, K., Yamagishi, A., Kikuchi, H., Ikeo, K., Gojobori, T.
<2>Comparative complete genome sequence analysis of the amino acid replacements responsible for the thermostability of Corynebacterium efficiens.
<3>Genome Res.
<4>13
<5>1572-1579
<6>2003
<7>Corynebacterium efficiens is the closest relative of Corynebacterium glutamicum,  a species
widely used for the industrial production of amino acids. C. efficiens
but not C. glutamicum can grow above 40 degrees C. We sequenced the complete C.
efficiens genome to investigate the basis of its thermostability by comparing its
genome with that of C. glutamicum. The difference in GC content between the
species was reflected in codon usage and nucleotide substitutions. Our
comparative genomic study clearly showed that there was tremendous bias in amino
acid substitutions in all orthologous ORFs. Analysis of the direction of the
amino acid substitutions suggested that three substitutions are important for the
stability of the C. efficiens proteins: from lysine to arginine, serine to
alanine, and serine to threonine. Our results strongly suggest that the
accumulation of these three types of amino acid substitutions correlates with the
acquisition of thermostability and is responsible for the greater GC content of
C. efficiens.

<>

<1>Nishioka, M., Fujiwara, S., Takagi, M., Imanaka, T.
<2>Characterization of two intein homing endonucleases encoded in the DNA polymerase gene of Pyrococcus kodakaraensis strain KOD1.
<3>Nucleic Acids Res.
<4>26
<5>4409-4412
<6>1998
<7>Two intein endonucleases, denoted PI-PkoI and PI-PkoII, in the DNA polymerase gene of the
hyperthermophilic archaeon Pyrococcus kodakaraensis KOD1 were expressed in Escherichia coli
and the recombinant endonucleases were characterized.  Both endonucleases were thermostable
and cleaved their inteinless DNA sequences leaving four base 3'-hydroxyl overhangs.  PI-PkoII
and the activity of PI-PkoII was enhanced at higher potassium ion concentrations (1M).
Recognition sequences were also determined using synthetic oligonucleotides inserted into
plasmid pUC19.  It was shown that DNA sequences of 19 and 16 bp are needed for cleavage by
PI-PkoI and PI-PkoII, respectively.  PI-PkoII could cleave the downstream junction region
between intein-encoding and mature DNA polymerase regions and cleavage by PI-PkoII could be
detected even when chromosomal DNA of P. kodakaraensis KOD1 was used as substrate.  Therefore,
it is suggested that these endonucleases are switching endonucleases whose function lies in
the rearrangement of chromosomal DNA.

<>

<1>Nishiyama, R., Ito, M., Yamaguchi, Y., Koizumi, N., Sano, H.
<2>Correlation between maternal inheritance of chloroplast genes and a chloroplast-resident DNA methyltransferase in the green alga, Chlamydomonas reinhardtii.
<3>Plant Cell Physiol.
<4>43
<5>s32
<6>2002
<7>Chloroplast DNA of Chlamydomonas reinhardtii is maternally inherited.  Methylation mapping and
indirect immunofluorescence analyses revealed that chloroplast DNA of mating type plus (mt+)
gametes is heavily methylated while that of mating type minus (mt-) gametes is not.  To
clarify the relationship between methylation and maternal inheritance of chloroplast DNA, we
isolated a cDNA encoding a DNA methyltransferase.  The deduced protein, CrMET1 was transferred
to chloroplasts, shown by GFP analyses.  Upon gametogenesis, CrMET1 transcripts clearly
increased in mt+ but not in mt- cells.  These experiments suggest that the CrMET1 protein is
located in chloroplasts and that it specifically methylates chloroplast DNA in mt+ gametes.
This conclusion was further strengthened by the observation that, during gametogenesis, CrMET1
is expressed in a mt- mutant, mat-1, whose chloroplast DNA is heavily methylated in gametes
and paternally inherited.  The results provide evidence that cytosine methylation plays a
critical role in maternal inheritance of chloroplast genes.

<>

<1>Nishiyama, R., Ito, M., Yamaguchi, Y., Koizumi, N., Sano, H.
<2>A chloroplast-resident DNA methyltransferase is responsible for hypermethylation of chloroplast genes in Chlamydomonas maternal gametes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>5925-5930
<6>2002
<7>Chloroplast DNA of the green alga Chlamydomonas reinhardtii is maternally inherited.
Methylation mapping directly revealed that,
before mating, chloroplast DNA of maternal (mating type plus; mt(+))
gametes is heavily methylated whereas that of paternal (mating type
minus; mt(-)) gametes is not. Indirect immunofluorescence analyses with
anti-5-methylcytosine mAbs visually showed methylation to occur
exclusively in chloroplast DNA of mt+ gametes, and not in mt- gametes
or nuclear DNA of either mt. To clarify the relationship between
methylation and maternal inheritance of chloroplast DNA, we have
isolated and characterized a cDNA encoding a DNA methyltransferase. The
deduced protein, CrMET1, consists of 1,344 aa and contains a conserved
catalytic domain at the C terminal and a nonconserved N-terminal
region. The predicted N-terminal region has an arginine-rich domain,
suggesting CrMET1 is transferred to chloroplasts. This finding could be
directly shown by green fluorescent protein epifluorescence microscopy
analyses. CrMET1 transcripts were found to be absent in both mt(+) and
mt(-) vegetative cells. Upon gametogenesis, however, transcript levels
clearly increased in mt(+) but not mt(-) cells. These experiments
suggest that the CrMET1 protein is located in chloroplasts and that it
specifically methylates cytosine residues of chloroplast DNA in mt+
gametes. This conclusion was further strengthened by the observation
that, during gametogenesis, CrMET1 is expressed in a mt(-) mutant,
mat-1, whose chloroplast DNA is heavily methylated in gametes and
paternally inherited. The results provide evidence that cytosine
methylation plays a critical role in maternal inheritance of
chloroplast genes in C. reinhardtii.

<>

<1>Nishiyama, R., Wada, Y., Mibu, M., Yamaguchi, Y., Shimogawara, K., Sano, H.
<2>Role of a nonselective de novo DNA methyltransferase in maternal inheritance of chloroplast genes in the green alga, Chlamydomonas  reinhardtii.
<3>Genetics
<4>168
<5>809-816
<6>2004
<7>In the green alga, Chlamydomonas, chloroplast DNA is maternally transmitted to the offspring.
We previously hypothesized that the
underlying molecular mechanism involves specific methylation of
maternal gamete DNA before mating, protecting against degradation. To
obtain direct evidence for this, we focused on a DNA methyltransferase,
DMT1, which was previously shown to be localized in chloroplasts. The
full-length DMT1 protein with a molecular mass of 150 kD was expressed
in insect cells, and its catalytic activity was determined. In vitro
assays using synthetic DNA indicated methylation of all cytosine
residues, with no clear selectivity in terms of the neighboring
nucleotides. Subsequently, transgenic paternal cells constitutively
expressing DMTI were constructed and direct methylation mapping assays
of their DNA showed a clear nonselective methylation of chloroplast
DNA. When transgenic paternal cells were crossed with wild-type
maternal cells, the frequency of biparental and paternal offspring of
chloroplasts increased up to 23% while between wild-type strains it was
similar to3%. The results indicate that DMT1 is a novel type of DNA
methyltransferase with a nonselective cytosine methylation activity,
and that chloroplast DNA methylation by DMT1 is one of factors
influencing maternal inheritance of chloroplast genes.

<>

<1>Nishizawa, A., Arshad, A.B., Nishizawa, T., Asayama, M., Fujii, K., Nakano, T., Harada, K., Shirai, M.
<2>Cloning and characterization of a new hetero-gene cluster of nonribosomal peptide synthetase and polyketide synthase from the cyanobacterium Microcystis aeruginosa K-139.
<3>J. Gen. Appl. Microbiol.
<4>53
<5>17-27
<6>2007
<7>Two nonribosomal peptide synthetase genes responsible for the biosynthesis
of microcystin and micropeptin in Microcystis aeruginosa K-139 have been
identified. A new nonribosomal peptide synthetase gene, psm3, was
identified in M. aeruginosa K-139. The gene is a cluster extending 30 kb
and comprising 13 bidirectionally transcribed open reading frames arranged
in two putative operons. psm3 encodes four adenylation proteins, one
polyketide synthase, and several unique proteins, especially Psm3L
consisting of halogenase, acyl-CoA binding protein-like protein, and acyl
carrier protein. Alignment of the binding pocket of the adenylation domain
and an ATP-PPi exchange analysis using a recombinant protein with the
adenylation domain of Psm3B showed that Psm3G and Psm3B activate aspartic
acid and tyrosine, respectively. Although disruption of psm3 did not
reveal the product produced by Psm3, we identified microviridin B and
aeruginosin K139 in the cells of M. aeruginosa K-139. The above-mentioned
results indicated that M. aeruginosa possesses at least five nonribosomal
peptide synthetase gene clusters.

<>

<1>Nishizawa, T., Hanami, T., Hirano, E., Miura, T., Watanabe, Y., Takanezawa, A., Komatsuzaki, M., Ohta, H., Shirai, M., Asayama, M.
<2>Isolation and Molecular Characterization of a Multicellular Cyanobacterium, Limnothrix/Pseudanabaena sp Strain ABRG5-3.
<3>Biosci. Biotechnol. Biochem.
<4>74
<5>1827-1835
<6>2010
<7>A cyanobacterium, semi-filamentous multicellular strain ABRG5-3, was isolated and its unique
nature was characterized. This axenic strain
formal colonies and was motile on an agarose plate. The 16S rRNA gene
of ABRG5-3 exhibited similarities to those of the Limnothrix and
Pseudanabaena strains, which are known as filamentous and
nonheterocystous cyanobacteria. Peaks in absorbance for the
accumulation of chlorophyll a, phycocyanin, and phycoerythrin were
observed in the cell extract. Natural separation of the pigments
occurred in the supernatant of the autolysed cells. The cell lysis was
promoted by osmotic shocks and lysozyme treatments. Chlorophyll a and
total DNA were abundantly recovered from the cells. Analysis of the
restriction-modification system for genomic DNA revealed novel
diversity. Moreover, we made a successful attempt to create
antibiotic-resistant strains by conjugation with a foreign plasmid,
which indicates that strain ABRG5-3 is transformable.

<>

<1>Nishizawa, T., Tago, K., Oshima, K., Hattori, M., Ishii, S., Otsuka, S., Senoo, K.
<2>Complete Genome Sequence of the Denitrifying and N2O-Reducing Bacterium Azoarcus  sp. Strain KH32C.
<3>J. Bacteriol.
<4>194
<5>1255
<6>2012
<7>We report the finished and annotated genome sequence of a denitrifying and N(2)O-reducing
betaproteobacterium, Azoarcus sp. strain KH32C. The genome is
composed of one chromosome and one megaplasmid and contains genes for
plant-microbe interactions and the gene clusters for aromatic-compound
degradations.

<>

<1>Niu, B., Rueckert, C., Blom, J., Wang, Q., Borriss, R.
<2>The Genome of the Plant Growth-Promoting Rhizobacterium Paenibacillus polymyxa M-1 Contains Nine Sites Dedicated to Nonribosomal Synthesis of  Lipopeptides and Polyketides.
<3>J. Bacteriol.
<4>193
<5>5862-5863
<6>2011
<7>The genome of Paenibacillus polymyxa M-1 consisted of a 5.8-Mb chromosome and a 360-kb
plasmid. Nine sites were dedicated to nonribosomal synthesis
of lipopeptides and polyketides. Eight of them were located at the
chromosome, while one gene cluster predicted to encode an unknown
secondary metabolite was present on the plasmid.

<>

<1>Niu, Y., Tenney, K., Li, H., Gimble, F.S.
<2>Engineering Variants of the I-SceI Homing Endonuclease with Strand-specific and Site-specific DNA-nicking Activity.
<3>J. Mol. Biol.
<4>382
<5>188-202
<6>2008
<7>The number of strand-specific nicking endonucleases that are currently available for
laboratory procedures and applications in vivo is limited,
and none is sufficiently specific to nick single target sites within
complex genomes. The extreme target specificity of homing endonucleases
makes them attractive candidates for engineering high-specificity nicking
endonucleases. I-SceI is a monomeric homing enzyme that recognizes an 18
bp asymmetric target sequence, and cleaves both DNA strands to leave
3'-overhangs of 4 bp. In single turnover experiments using plasmid
substrates, I-SceI generates transient open circle intermediates during
the conversion of supercoiled to linear DNA, indicating that the enzyme
cleaves the two DNA strands sequentially. A novel hairpin substrate was
used to demonstrate that although wild-type I-SceI cleaves either the top
or bottom DNA strand first to generate two nicked DNA intermediates, the
enzyme has a preference for cleaving the bottom strand. The kinetics data
are consistent with a parallel sequential reaction mechanism. Substitution
of two pseudo-symmetric residues, Lys122 and Lys223, markedly reduces top
and bottom-strand cleavage, respectively, to generate enzymes with
significant strand- and sequence-specific nicking activity. The two active
sites are partially interdependent, since alterations to one site affect
the second. The kinetics analysis is consistent with X-ray crystal
structures of I-SceI/DNA complexes that reveal a role for the lysines in
establishing important solvent networks that include nucleophilic water
molecules thought to attack the scissile phosphodiester bonds.

<>

<1>Niv, M.Y., Ripoll, D.R., Vila, J.A., Liwo, A., Vanamee, E.S., Aggarwal, A.K., Weinstein, H., Scheraga, H.A.
<2>Topology of Type II REases revisited; structural classes and the common conserved core.
<3>Nucleic Acids Res.
<4>35
<5>2227-2237
<6>2007
<7>Type II restriction endonucleases (REases) are deoxyribonucleases that cleave DNA sequences
with remarkable specificity. Type II REases are highly divergent in sequence as well as in
topology, i.e. the connectivity of secondary structure elements. A widely held assumption is
that a structural core of five beta-strands flanked by two alpha-helices is common to these
enzymes. We introduce a systematic procedure to enumerate secondary structure elements in an
unambiguous and reproducible way, and use it to analyze the currently available X-ray
structures of Type II REases. Based on this analysis, we propose an alternative definition of
the core, which we term the alphabetaalpha-core. The alphabetaalpha-core includes the most
frequently observed secondary structure elements and is not a sandwich, as it consists of a
five-strand beta-sheet and two alpha-helices on the same face of the beta-sheet. We use the
alphabetaalpha-core connectivity as a basis for grouping the Type II REases into distinct
structural classes. In these new structural classes, the connectivity correlates with the
angles between the secondary structure elements and with the cleavage patterns of the REases.
We show that there exists a substructure of the alphabetaalpha-core, namely a common conserved
core, ccc, defined here as one alpha-helix and four beta-strands common to all Type II REase
of known structure.

<>

<1>Niv, M.Y., Skrabanek, L., Roberts, R.J., Scheraga, H.A., Weinstein, H.
<2>Identification of GATC- and CCGG-recognizing Type II REases and their putative specificity-determining positions using Scan2S--a novel motif scan algorithm with  optional secondary structure constraints.
<3>Proteins
<4>71
<5>631-640
<6>2008
<7>Restriction endonucleases (REases) are DNA-cleaving enzymes that have become indispensable
tools in molecular biology. Type II REases are highly divergent in
sequence despite their common structural core, function and, in some cases,
common specificities towards DNA sequences. This makes it difficult to identify
and classify them functionally based on sequence, and has hampered the efforts of
specificity-engineering. Here, we define novel REase sequence motifs, which
extend beyond the PD-(D/E)XK hallmark, and incorporate secondary structure
information. The automated search using these motifs is carried out with a newly
developed fast regular expression matching algorithm that accommodates long
patterns with optional secondary structure constraints. Using this new tool,
named Scan2S, motifs derived from REases with specificity towards GATC- and
CGGG-containing DNA sequences successfully identify REases of the same
specificity. Notably, some of these sequences are not identified by standard
sequence detection tools. The new motifs highlight potential
specificity-determining positions that do not fully overlap for the GATC- and the
CCGG-recognizing REases and are candidates for specificity re-engineering.

<>

<1>Nkenfou, C.
<2>Cloning and studying of the cleavage flexibility of some type IIs  restriction endonucleases.
<3>Ph.D. Thesis, University of Yaounde I, Cameroon, , , Yaounde
<4>
<5>1-226
<6>2002
<7>More than 3000 bacterial samples were screened for restriction
enzymes.  Three new specificities were found and a few isoschizomers with
interesting properties.  Two of these isoschizomers: BceAI, an isoschizomer
of BcefI (ACGGC) and BspCNI, an isoschizomer of BseMII (CTCAG), belonging
to the subgroup of Type IIs restriction enzymes were cloned as well as the
prototype of BceAI, BcefI.  These two cloned isoschizomers and one other,
BpuCI, a neoschizomer of Ecil (GGCGGA), were studied for their cleavage
flexibility alongside that of their prototypes.  These studies were carried
out on different DNA substrates.  The conformation as well as the bending
mode of these DNA substrates were predicted using computer programs.  The
flexibility of the 3 pairs of enzymes studied was as follows:  For BceAI
and BcefI, their preferred site was 12/14 and their alternative site was
11/13.  For BspCNI and BseMII, for some sequences their preferred site was
10/8, for other sequences the preferred site was 9/7 and some sequences
were cleaved at both positions with the same rate.  For BpuCI and Ecil, the
preferred site for BpuCI was 12/10 and its alternative site was 13/11.  The
preferred site for EciI was 11/9 and its alternative site was 12/10.  These
cleavage positions and rates were determined to be sequence-dependent as
well as influenced by the conformation of the protein (enzyme).  Based on
the prediction of the conformation and bending mode of the different
substrates together with the results of the cleavage positions and rates,
allowed the following conclusions.  For sequences presenting an anisotropic
bending mode, the cleavage position tended to be further away from the
recognition sequence.  For sequences presenting an isotropic bending mode,
the cleavage tended to occur closer to the recognition sequence.

<>

<1>Noar, J., Makwana, S.T., Bruno-Barcena, J.M.
<2>Complete Genome Sequence of Solvent-Tolerant Clostridium beijerinckii Strain SA-1.
<3>Genome Announcements
<4>2
<5>e01310-14
<6>2014
<7>We report the complete genome sequence of Clostridium beijerinckii SA-1, derived  by directed
evolution from C. beijerinckii NCIMB 8052, selecting for enhanced
solvent tolerance. This sequence allows for accurate placement of SA-1 as C.
beijerinckii, permits functional analyses of mutant phenotypes, and suggests
methods for distinguishing SA-1 from its parent.

<>

<1>Noar, J.D., Bruno-Barcena, J.M.
<2>Complete Genome Sequences of Azotobacter vinelandii Wild-Type Strain CA and Tungsten-Tolerant Mutant Strain CA6.
<3>Genome Announcements
<4>1
<5>e00313-13
<6>2013
<7>We report the complete genome sequences of Azotobacter vinelandii mutant strain CA6 and its
parent wild-type strain, CA. When fixing nitrogen, strain CA6
displays slow growth and impaired molybdate uptake, tolerance to tungstates, and
production of hydrogen gas, compared to results for strain CA. Comparing these
genome sequences may provide a genetic basis for these mutant phenotypes.

<>

<1>Nobbs, T.F., Wentzell, L.M., Szczelkum, M.D., Halford, S.E.
<2>SfiI: An unconventional restriction enzyme.
<3>The NEB Transcript
<4>8
<5>10-11
<6>1997
<7>Abrief review of SfiI

<>

<1>Nobbs, T.J., Halford, S.E.
<2>DNA cleavage at two recognition sites by the SfiI restriction endonuclease: Salt dependence of Cis and Trans interactions between distant DNA sites.
<3>J. Mol. Biol.
<4>252
<5>399-411
<6>1995
<7>At low ionic strength, the SfiI restriction enzyme cleaved at similar rates both supercoiled
and linear DNA with two SfiI sites and linear DNA with one SfiI site.  For the substrates with
two sites, the majority of the DNA was converted directly to products cut at both sites; the
enzyme appears to bind to two sites before catalyzing its reactions, looping out the
intervening DNA.  At high ionic strength, linear DNA with one SfiI site was not cut at all,
linear DNA with two sites was cleaved slowly while supercoiled DNA with two sites was cleaved
rapidly, though only half of the DNA with two sites was cut at both sites; the DNA that had
been cut at one site was not cleaved again at the remaining site.  The singly cut product must
therefore have been generated by a reaction incorporating both sites.  All DNA cleavage
reactions by SfiI thus involve the tetrameric enzyme bound to two copies of its recognition
sequence, but weakened DNA-protein interactions at high ionic strength can cause this complex
to dissociate before cleaving both sites.  Intramolecular interactions between distant DNA
sites are generally thought to be enhanced by supercoiling and to be more stable than
intermolecular interactions.  The preference of SfiI at high ionic strength for substrates
with two sites over substrates with one site and, in the former case, for supercoiled over
linear DNA, validates this view.  At low ionic strength, the similar rates with the different
substrates may be due to rate-limiting product dissociation.

<>

<1>Nobbs, T.J., Szczelkun, M.D., Wentzell, L.M., Halford, S.E.
<2>DNA excision by the SfiI restriction endonuclease.
<3>J. Mol. Biol.
<4>281
<5>419-432
<6>1998
<7>A mechanism for the precise excision of DNA between two target sites was elucidated by
analyzing the individual steps during the reactions of the SfiI endonuclease on a plasmid with
two SfiI sites.  Previous studies had indicated that SfiI is a tetrameric protein that binds
to two copies of its recognition site before cleaving both sites in both strands.  In this
study, the concerted cleavage of four phosphodiester bonds was shown to arise from four
consecutive reactions that had similar values for their intrinsic rate constants.  Each
reaction is presumably mediated by one of the four active sites in the tetramer and all four
were generally completed within the life-time of the complex between the protein and two
recognition sites, though products cleaved in one or two phosphodiester bonds were also
detected following premature dissociation of the enzyme-substrate complex at elevated
temperatures.  At the physiological temperature for this enzyme, all four bonds were cleaved
within one minute but the subsequent dissociation of the enzyme-product complex, liberating
the excised segment of DNA, took about one hour.  The tetrameric structure of SfiI was
confirmed by equilibrium centrifugation.

<>

<1>Nobbs, T.J., Williams, S.A., Connolly, B.A., Halford, S.E.
<2>Phosphorothioate substrates for the SfiI restriction endonuclease.
<3>Biol. Chem.
<4>379
<5>599-604
<6>1998
<7>Oligodeoxynucleotides carrying the recognition sequence for the SfiI endonuclease were
synthesized with phosphorothioates at the cleavage site.  The Rp and Sp diasteroisomers of the
oligonucleotides were separated by HPLC using a mobile phase containing L-cysteine.  The
duplex with Rp phosphorothioates was cleaved very slowly in the presence of Mg2+, though
virtually complete cleavage was obtained with Mn2+.  No significant cleavage of the duplex
with Sp phosphorothioates occurred with either Mg2+ or Mn2+.  When added to a plasmid with one
SfiI site, the duplexes with either Rp or Sp phosphorothioates inhibited the rate at which
SfiI cleaved the plasmid: a control duplex with oxyester linkages enhanced the rate of plasmid
cleavage.  In contrast to type IIe nucleases such as EcoRII and NaeI, which can be activated
by non-hydrolysable analogues of their substrates, SfiI reactions require four susceptible
phosphodiester bonds.

<>

<1>Nobile, C.
<2>An improved method for partial restriction digestion of ultraviolet irradiated DNA.
<3>Nucleic Acids Res.
<4>18
<5>4288
<6>1990
<7>None

<>

<1>Nobu, M.K., Narihiro, T., Tamaki, H., Qiu, Y.L., Sekiguchi, Y., Woyke, T., Goodwin, L., Davenport, K.W., Kamagata, Y., Liu, W.T.
<2>Draft Genome Sequence of Syntrophorhabdus aromaticivorans Strain UI, a Mesophilic Aromatic Compound-Degrading Syntroph.
<3>Genome Announcements
<4>2
<5>e01064-13
<6>2014
<7>Syntrophorhabdus aromaticivorans strain UI is a mesophilic bacterium capable of degrading
aromatic substrates in syntrophic cooperation with a partner
methanogen. The draft genome sequence is 3.7 Mb, with a G+C content of 52.0%.

<>

<1>Nobusato, A., Uchiyama, I., Kobayashi, I.
<2>Diversity of restriction-modification gene homologues in Helicobacter pylori.
<3>Gene
<4>259
<5>89-98
<6>2000
<7>The complete genome sequences of two Helicobacter pylori strains have recently become
available. We have searched them for homologues of restriction-modification genes. One strain
(26695) carried 52 such homologues, and the other (J99) carried 53. Their sequence alignments
were arranged in the form of a phylogenetic tree and compared with the tree based on rRNA. The
trees showed that the homologues are scattered among diverse groups of bacteria. They also
revealed high polymorphism within the species - there are 42 pairs with high homology, 10
specific to 26695, and 11 specific to J99. Many of the restriction-modification homologues
were characterized by a GC content lower than that of the average gene in the genome. Some of
the restriction-modification homologues showed a different codon use bias from the average
genes. These observations are interpreted in terms of horizontal transfer of the
restriction-modification genes.

<>

<1>Nobusato, A., Uchiyama, I., Ohashi, S., Kobayashi, I.
<2>Insertion with long target duplication: a mechanism for gene mobility suggested from comparison of two related bacterial genomes.
<3>Gene
<4>259
<5>99-108
<6>2000
<7>The complete genome sequences of two closely related organisms - two Helicobacter pylori
strains - have recently become available. Comparison of these genomes at single base pair
level has suggested the presence of a mechanism for bacterial gene mobility - insertion with
long target duplications. This mechanism is formally similar to classical transposon
insertion, but the duplication is much longer, often in the range of 100bp.  Restriction
and/or modification enzyme genes are often within or adjacent to the insertion. A similar
process may have mediated insertion of the cag+ pathogenicity island in H. pylori. A similar
structure was identified in comparisons between Neisseria meningitidis and Neisseria
gonorrhoeae genomes. We hypothesize that this mechanism, as well as two other types of
polymorphism linked with restriction-modification genes (insertion accompanied by target
deletion and a tripartite structure composed of  substitution/inversion/deletion), have
resulted from attack by restriction enzymes on the chromosome.

<>

<1>Noel, A.J., Wende, W., Pingoud, A.
<2>DNA recognition by the homing endonuclease PI-SceI involves a divalent metal ion cofactor-induced conformational change.
<3>J. Biol. Chem.
<4>279
<5>6794-6804
<6>2004
<7>PI-SceI, a homing endonuclease of the IAGLIDADG family, consists of two domains involved in
DNA cleavage and protein splicing, respectively.
Both domains cooperate in binding the recognition sequence. Comparison
of the structures of PI-SceI in the absence and presence of substrate
reveals major conformational changes in both the protein and DNA.
Notably, in the protein-splicing domain the loop comprising residues
53-70 and adopts a "closed" conformation, thus enabling it to interact
with the DNA. We have studied the dynamics of DNA binding and
subsequent loop movement by fluorescence techniques. Six amino acids in
loop53-70 were individually replaced by cysteine and modified by
fluorescein. The interaction of the modified PI-SceI variants with the
substrate, unlabeled or labeled with tetramethylrhodamine, was analyzed
in equilibrium and stopped-flow experiments. A kinetic scheme was
established describing the interaction between PI-SceI and DNA. It is
noteworthy that the apparent hinge-flap motion of loop53-70 is only
observed in the presence of a divalent metal ion cofactor. Substitution
of the major Mg2+-binding ligands in PI-SceI, Asp-218 and Asp-326, by
Asn or "nicking" PI-SceI with trypsin at Arg-277, which interferes with
formation of an active enzyme-substrate complex, both prevent the
conformational change of loop53-70. Deletion of the loop inactivates
the enzyme. We conclude that loop53-70 is an important structural
element that couples DNA recognition by the splicing domain with DNA
cleavage by the catalytic domain and as such "communicates" with the
Mg2+ binding sites at the catalytic centers.

<>

<1>Nogami, S., Fukuda, T., Nagai, Y., Yabe, S., Sugiura, M., Mizutani, R., Satow, Y., Anraku, Y., Ohya, Y.
<2>Homing at an extragenic locus mediated by VDE (PI-Scel) in  Saccharomyces cerevisiae.
<3>Yeast
<4>19
<5>773-782
<6>2002
<7>PI-SceI (VDE), a homing endonuclease with protein splicing activity,  is a genomic parasite in
the VMA1 gene of Saccharomyces cerevisiae. In a
heterozygous diploid of the VDE-less VMA1 allele and a VDE-containing VMA1
allele, VDE specifically cleaves its recognition sequence (VRS) in the
VDE-less VMA1 allele at meiosis, followed by 'homing', i.e. a conversion
to a VDE-containing allele. We found that upon VDE expression, homing of a
marker gene at an extragenic locus occurs only when a 45 bp element
containing the VRS is inserted at its allelic site, while mutants of VDE
with no endonuclease activity lack authentic extragenic homing activity.
Thus, both the VRS and VDE are required for homing. Insertion of the VRS
in a homozygous diploid significantly lowered the spore germination
ability, indicating that a template for gene repair at its allelic locus
is essential for efficient homing and survival of yeast cells. Copyright
(C) 2002 John Wiley Sons, Ltd.

<>

<1>Nolan, M. et al.
<2>Complete genome sequence of Ferrimonas balearica type strain (PAT).
<3>Standards in Genomic Sciences
<4>3
<5>174-182
<6>2010
<7>Ferrimonas balearica Rossello-Mora et al. 1996 is the type species of the genus Ferrimonas,
which belongs to the family Ferrimonadaceae within the
Gammaproteobacteria. The species is a Gram-negative, motile, facultatively
anaerobic, non spore-forming bacterium, which is of special interest because it
is a chemoorganotroph and has a strictly respiratory metabolism with oxygen,
nitrate, Fe(III)-oxyhydroxide, Fe(III)-citrate, MnO(2), selenate, selenite and
thiosulfate as electron acceptors. This is the first completed genome sequence of
a member of the genus Ferrimonas and also the first sequence from a member of the
family Ferrimonadaceae. The 4,279,159 bp long genome with its 3,803
protein-coding and 144 RNA genes is a part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Nolan, M. et al.
<2>Complete genome sequence of Streptobacillus moniliformis type strain (9901).
<3>Standards in Genomic Sciences
<4>1
<5>300-307
<6>2009
<7>Streptobacillus moniliformis Levaditi et al. 1925 is the type and sole species of the genus
Streptobacillus, and is of phylogenetic interest because of its
isolated location in the sparsely populated and neither taxonomically nor
genomically much accessed family 'Leptotrichiaceae' within the phylum
Fusobacteria. The 'Leptotrichiaceae' have not been well characterized,
genomically or taxonomically. S. moniliformis,is a Gram-negative, non-motile,
pleomorphic bacterium and is the etiologic agent of rat bite fever and Haverhill
fever. Strain 9901(T), the type strain of the species, was isolated from a
patient with rat bite fever. Here we describe the features of this organism,
together with the complete genome sequence and annotation. This is only the
second completed genome sequence of the order Fusobacteriales and no more than
the third sequence from the phylum Fusobacteria. The 1,662,578 bp long chromosome
and the 10,702 bp plasmid with a total of 1511 protein-coding and 55 RNA genes
are part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Nolan, M. et al.
<2>Complete genome sequence of Rhodothermus marinus type strain (R-10).
<3>Standards in Genomic Sciences
<4>1
<5>283-290
<6>2009
<7>Rhodothermus marinus Alfredsson et al. 1995 is the type species of the genus and  is of
phylogenetic interest because the Rhodothermaceae represent the deepest
lineage in the phylum Bacteroidetes. R. marinus R-10(T) is a Gram-negative,
non-motile, non-spore-forming bacterium isolated from marine hot springs off the
coast of Iceland. Strain R-10(T) is strictly aerobic and requires slightly
halophilic conditions for growth. Here we describe the features of this organism,
together with the complete genome sequence, and annotation. This is the first
complete genome sequence of the genus Rhodothermus, and only the second sequence
from members of the family Rhodothermaceae. The 3,386,737 bp genome (including a
125 kb plasmid) with its 2914 protein-coding and 48 RNA genes is part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Nolan, M. et al.
<2>Complete genome sequence of Streptosporangium roseum type strain (NI 9100).
<3>Standards in Genomic Sciences
<4>2
<5>29-37
<6>2010
<7>Streptosporangium roseum Crauch 1955 is the type strain of the species which is the type
species of the genus Streptosporangium. The 'pinkish coiled
Streptomyces-like organism with a spore case' was isolated from vegetable garden
soil in 1955. Here we describe the features of this organism, together with the
complete genome sequence and annotation. This is the first completed genome
sequence of a member of the family Streptosporangiaceae, and the second largest
microbial genome sequence ever deciphered. The 10,369,518 bp long genome with its
9421 protein-coding and 80 RNA genes is a part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Nolling, J. et al.
<2>Genome sequencing and comparative analysis of the solvent-producing bacterium Clostridium acetobutylicum.
<3>J. Bacteriol.
<4>183
<5>4823-4838
<6>2001
<7>The genome sequence of the solvent-producing bacterium Clostridium acetobutylicum ATCC 824 has
been determined by the shotgun approach.  The genome consists of a 3.94-Mb chromosome and a
192-kb megaplasmid that contains the majority of genes responsible for solvent production.
Comparison of C. acetobutylicum to Bacillus subtilis reveals significant local conservation of
gene order, which has not been seen in comparisons of other genomes with similar, or, in some
cases closer, phylogenetic proximity.  This conservation allows the prediction of many
previously undetected operons in both bacteria.  However, the C. acetobutylicum genome also
contains a significant number of predicted operons that are shared with distantly related
bacteria and archaea but not with B. subtilis.  Phylogenetic analysis is compatible with the
dissemination of such operons by horizontal transfer.  The enzymes of the solventogenesis
pathway and of the cellulosome of C. acetobutylicum comprise a new set of metabolic capacities
not previously represented in the collection of complete genomes.  These enzymes show a
complex pattern of evolutionary affinities, emphasizing the role of lateral gene exchange in
the evolution of the unique metabolic profile of the bacterium.  Many of the sporulation genes
identified in B. subtilis are missing in C. acetobutylicum, which suggests major differences
in the sporulation process.  Thus, comparative analysis reveals both significant conservation
of the genome organization and pronounced differences in many systems that reflect unique
adaptive strategies of the two gram-positive bacteria.

<>

<1>Nolling, J., de Vos, W.M.
<2>Characterization of the archaeal, plasmid-encoded type II restriction-modification system MthTI from Methanobacterium thermoformicicum THF: Homology to the bacterial NgoPII system from Neisseria gonorrhoeae.
<3>J. Bacteriol.
<4>174
<5>5719-5726
<6>1992
<7>A restriction-modification system, designated MthTI, was localized on plasmid pFV1 from the
thermophilic archaeon Methanobacterium thermoformicicum THF. The MthTI system is a new member
of the family of GGCC-recognizing restriction-modification systems. Functional expression of
the archaeal MthTI genes was obtained in Escherichia coli. The mthTIR and mthTIM genes are 843
and 990 bp in size and code for proteins of 281 (32,102 Da) and 330 (37,360 Da) amino acids,
respectively. The deduced amino acids sequences of M.MthTI showed high similarity with that of
the isospecific methyltransferases M.NgoPII and M.HaeIII. In addition, extensive sequence
similarity on the amino acid level was observed for the endonucleases R.MthTI and R.NgoPII.
Moreover, the endonuclease and methyltransferase genes of the thermophilic MthTI system and
those of Neisseria gonorrhoeae NgoPII system show identical organizations and high (54.5%)
nucleotide identity. This finding suggests horizontal transfer of restriction-modification
systems between members of the domains Bacteria and Archaea.

<>

<1>Nolling, J., de Vos, W.M.
<2>Identification of the CTAG-recognizing restriction-modification systems MthZI and MthFI from Methanobacterium thermoformicicum and characterization of the plasmid-encoded mthZIM gene.
<3>Nucleic Acids Res.
<4>20
<5>5047-5052
<6>1992
<7>Two CTAG-recognizing restriction and modification (R/M) systems, designated MthZI and MthFI,
were identified in the thermophilic archaeon Methanobacterium thermoformicicum strains Z-245
and FTF, respectively. Further analysis revealed that the methyltransferase (MTase) genes are
plasmid-located in both strains. The plasmid pFZ1-encoded mthZIM gene of strain Z-245 was
further characterized by subcloning and expression studies in Escherichia coli followed by
nucleotide sequence analysis. The mthZIM gene is 1065 bp in size and may code for a protein of
355 amino acids (Mr 42,476 Da). The deduced amino acid sequence of the M.MthZI enzyme shares
substantial similarity with four distinct regions from several m4C and m6A-MTases, and
contains the TSPPY motif that is so far only found in m4C-MTases. Partially overlapping with
the mthZIM gene and in reverse orientation, an additional ORF was identified with a size of
606 bp potentially coding for a protein of 202 amino acids (mr 23.710 Da). This ORF is
suggested to encode the corresponding endonuclease R.MthZI.

<>

<1>Nolling, J., Groffen, A., De Vos, W.M.
<2>PhiF1 and PhiF3, two novel virulent, archaeal phages infecting different thermophilic strains of the genus Methanobacterium.
<3>J. Gen. Microbiol.
<4>139
<5>2511-2516
<6>1993
<7>Two virulent archaeal phages, PhiF1 and PhiF3, were isolated that were capable of infecting
different thermophilic members of the genus Methanobacterium. Both phages exhibited a similar
morphology consisting of a polyhedral head and tail, but differed considerably in their host
specificities and the size and topology of their genomes. Phage PhiF1 contained a linear,
double-stranded DNA genome of 85+/-5 kb in size and showed a broad host range including M.
thermoformicicum strains Z-245, FTF FF1, FF3 and CSM3, and M. thermoautotrophicum strain H. In
contrast, PhiF3 phage particles contained a circular or terminally redundant linear genome,
comprising approximately 36+/-2 kb double-stranded DNA, and could only be propagated on M.
thermoformicicum strain FF3. Hybridization experiments did not reveal similarity between the
genomes of PhiF1 and PhiF3 nor between both phages and genomic DNA from different thermophilic
members of the genus Methanobacterium or DNA from phage M1 of M. thermoautotrophicum Marburg.
A physical map of both phage genomes was constructed. The DNA of phage phiF1 was found to
contain multiple GGCC sites which form the target for the restriction-modification (R/M)
system MthTI of M. thermoformicicum THF. In contrast, the DNA of PhiF1 contained only a single
CTAG site recognized by the R/M systems MthZI and MthFI of M. thermoformicicum Z-245 and FTF,
respectively. The distribution of these sites correlates well with the capacity of PhiF1 to
infect M. thermoformicicum strains Z-245 and FTF but not strain THF.

<>

<1>Nolling, J., Van Eeden, J.M., Eggen, R.I.L., de Vos, W.M.
<2>Modular organization of related Archaeal plasmids encoding different restriction-modification systems in Methanobacterium thermoformicicum.
<3>Nucleic Acids Res.
<4>20
<5>6501-6507
<6>1992
<7>Nucleotide sequence comparison of the related 13513-bp plasmid pFV1 and the 11014-bp plasmid
pFZ1 from the thermophilic Archaeon Methanobacterium thermoformicicum THF and Z-245,
respectively, revealed a homologous, approximately 8.2 kb backbone structure that is
interrupted by plasmid-specific elements. Various highly conserved palindromic structures and
an ORF that could code for an NTP-binding protein were identified within the backbone
structure and may be involved in plasmid maintenance and replication. Each plasmid contains at
comparable locations a module which specifies components of different restriction-modificaton
(R/M) systems. The R/M module of pFV1 contained, in addition to the genes of the
GGCC-recognizing R/M system MthTI, an ORF which may be involved in repair of G-T mismatches
generated by deamination of m5C at high temperatures.

<>

<1>Nomura, N., Morinaga, Y., Shirai, N., Sako, Y.
<2>I-Apel: A novel intron-encoded LAGLIDADG homing endonuclease from the archaeon, Aeropyrum pernix K1.
<3>Nucleic Acids Res.
<4>33
<5>5017
<6>2005
<7>Over 50 introns have been reported in archaeal rRNA genes (rDNAs), a subset of which nests
putative homing endonuclease (HEase) genes. Here, we report the identification and
characterization of a novel archaeal LAGLIDADG-type HEase, I-ApeI, encoded by the ApeK1.S908
intron within the 16S rDNA of Aeropyrum pernix K1. I-ApeI consists of 222 amino acids and
harbors two LAGLIDADG-like sequences. It recognizes the 20 bp non-palindromic sequence
5'-GCAAGGCTGAAAC TTAAAGG and cleaves target DNA to produce protruding tetranucleotide 3'
ends. Either Mn2+ or Co2+ can be substituted for Mg2+ as a cofactor in the cleavage reaction.
Of the 20 bases within the minimal recognition site, 7 are essential for cleavage and are
located at positions proximal to the cleavage sites.

<>

<1>Nomura, N., Nomura, Y., Sussman, D., Klein, D., Stoddard, B.L.
<2>Recognition of a common rDNA target site in archaea and eukarya by analogous LAGLIDADG and His-Cys box homing endonucleases.
<3>Nucleic Acids Res.
<4>36
<5>6988-6998
<6>2008
<7>The presence of a homing endonuclease gene (HEG) within a microbial intron or intein empowers
the entire element with the ability to invade
genomic targets. The persistence of a homing endonuclease lineage
depends in part on conservation of its DNA target site. One such rDNA
sequence has been invaded both in archaea and in eukarya, by LAGLIDADG
and His-Cys box homing endonucleases, respectively. The bases encoded
by this target include a universally conserved ribosomal structure,
termed helix 69 (H69) in the large ribosomal subunit. This region forms
the 'B2a' intersubunit bridge to the small ribosomal subunit, contacts
bound tRNA in the A-and P-sites, and acts as a trigger for ribosome
disassembly through its interactions with ribosome recycling factor. We
have determined the DNA-bound structure and specificity profile of an
archaeal LAGLIDADG homing endonuclease (I-Vdi141I) that recognizes this
target site, and compared its specificity with the analogous eukaryal
His-Cys box endonuclease I-PpoI. These homodimeric endonuclease
scaffolds have arrived at similar specificity profiles across their
common biological target and analogous solutions to the problem of
accommodating conserved asymmetries within the DNA sequence, but with
differences at individual base pairs that are fine-tuned to the
sequence conservation of archaeal versus eukaryal ribosomes.

<>

<1>Nomura, W., Barbas, C.F. III
<2>In Vivo Site-Specific DNA Methylation with a Designed Sequence-Enabled DNA Methylase.
<3>J. Am. Chem. Soc.
<4>129
<5>8676-8677
<6>2007
<7>As an alternative to the continual expression of transcriptional repressors to turn off genes
after they have served their purpose, nature has developed epigenetic strategies that result
in the covalent modification of DNA itself to induce heritable gene silencing.  Mounting
evidence supports the notion that once a genomic region has been targeted for silencing by
acquisition of one or more covalent epigenetic marks, mark can be propagated and may influence
acquisition of others.  If epigenetic modifications can be made specifically by the addition
of targeted exogenous agents, new approaches to transcriptional therapy should result.

<>

<1>Nomura, W., Masuda, A., Tamamura, H.
<2>Development of site-specific DNA methylase for epigenetic regulation of gene expression.
<3>Seikagaku
<4>82
<5>393-397
<6>2010
<7>
<>

<1>Nomura, W., Tamamura, H., Barbas, C.F. III
<2>Site-selective cytosine methylation by a split DNA methylase.
<3>Pept. Sci.
<4>45
<5>491-492
<6>2009
<7>Cytosine methylation plays pivotal roles in gene expression.  Methylation pattern in genome is
heritable, then the effect of methylation is largely influenced across generation.  Truly
specific methylation has been a challenging problem because fusion methylases with zinc finger
domain still methylate the native sites as background.  To avoid this, split methylase domains
were constructed.  These domains were designed to assemble only on the zinc finger target site
with high specificity.  The split methylase showed it works as specific methylase to the
target sites.

<>

<1>Nomura, Y., Ishino, Y., Kato, I.
<2>A novel restriction endonuclease.
<3>European Patent Office
<4>EP 0698663 A
<5>
<6>1994
<7>A restriction endonuclease which recognizes the nucleotide sequence of the
following chemical formula 1 in double stranded DNA and strictly cleaves the said nucleotide
sequence at the arrow-marked sites.  Chemical formula 1:  5'-AG^GA/TCCT-3' 3'-
TCCT/AG^GA-5'.  The endonuclease may be made by cultivating a strain of the genus
Streptomyces capable of producing it.

<>

<1>Nomura, Y., Ishino, Y., Kato, I.
<2>Restriction Endonuclease.
<3>US Patent Office
<4>US 5726052
<5>
<6>1998
<7>A restriction endonuclease which recognizes the nucleotide sequence of the following chemical
formula 1 in double stranded DNA and strictly cleaves the nucleotide sequence at the sites
marked by arrows.  5'-AGGA/TCCT-3' 3'-TCCT/AGGA-5'.  The endonuclease may be made
cultivating a strain of the genus Streptomyces such as Streptomyces sp. YH 8647 (FERM BP-5022)
capable of producing it.

<>

<1>Nomura, Y., Ishino, Y., Kimizuka, F., Kato, I.
<2>A novel Streptomyces restriction endonuclease, Sse1825I, cleaving at 5'-GG/GWCCC-3'.
<3>Gene
<4>157
<5>323-324
<6>1995
<7>We isolated and characterized from a Streptomyces species a new class-II restriction
endonuclease, which recognizes the palindromic heptanucleotide sequence: 5'-GG/GWCCC
3'-CCCWG/GG (where W=A or T) and cleaves double-stranded DNA after the second G in this
sequence.  This Sse1825I enzyme cleaves phage lambda DNA at one site, adenovirus type 2 DNA at
eight sites, but does not cleave pBR322, SV40, ColE1, pUC18 and pUC19, and replicative forms
of M13mp18 and M13mp19, and PhiX174 DNAs.

<>

<1>Nomura, Y., Ishizaki, I., Oshima, K., Kato, I.
<2>New restriction enzyme, Sse2321-restriction endonuclease extracted from Streptomyces sp.
<3>Japanese Patent Office
<4>JP 10127279
<5>
<6>1998
<7>
<>

<1>Nomura, Y., Kimizuka, F., Ishino, Y., Kato, I.
<2>Restriction endonuclease Sse1825I.
<3>European Patent Office
<4>EP 0655496 A
<5>
<6>1993
<7>A new restriction endonuclease capable of recognizing the nucleotide sequence of the following
formula 1 in double stranded DNA.  5'-GTGGA/TCCC-3' 3'-CCCT/AGGG-5' and specifically
cleaving it is disclosed.  The endonuclease may be made cultivating a strain of the genus
Streptomyces (FERM BP-4836) capable of producing it.

<>

<1>Nomura, Y., Kimizuka, F., Ishino, Y., Kato, I.
<2>Restriction endonuclease.
<3>US Patent Office
<4>US 5496717
<5>
<6>1996
<7>A new restriction endonuclease capable of recognizing the nucleotide sequence of the following
formula 1 in double stranded DNA.  5'-GGGA/TCCC-3' 3'-CCCT/AGGG-5' and specifically
cleaving it is disclosed.  The endonuclease may be made cultivating a strain of the genus
Streptomyces (FERM BP-4836) capable of producing it.

<>

<1>Nomura, Y., Kotani, H., Kita, K., Sadaoka, A., Hiraoka, N., Nakamura, T., Abe, N., Izaki, K.
<2>Purification and properties of a new restriction endonuclease, MxaI, from Myxococcus xanthus F18E.
<3>Agric. Biol. Chem.
<4>54
<5>3011-3012
<6>1990
<7>Gliding bacteria have an unusual cell cycle, which consists of vegetative
growth, the formation of a swarm, and the development of fruiting bodies and
microcysts.  Only one restriction endonuclease of these bacteria has been
reported.  Here, we screened 103 strains of Myxococcus species isolated from
soil and detected endonuclease activity in nine strains.  One strain,
Myxococcus xanthus F18E, produced a restriction endonuclease, MxaI, that is a
new isoschizomer of SacI.  We identified the recognition and cleavage sites of
MxaI and compared the properties of this enzyme with those of SacI.

<>

<1>Nonaka, H., Keresztes, G., Shinoda, Y., Ikenaga, Y., Abe, M., Naito, K., Inatomi, K.F.K., Inui, M., Yukawa, H.
<2>Complete genome sequence of the dehalorespiring bacterium Desulfitobacterium hafniense Y51 and comparison with Dehalococcoides ethenogenes 195 - bacterium genomic comparison and genome sequencing for use in bioremediation.
<3>J. Bacteriol.
<4>188
<5>2262-2274
<6>2006
<7>AUTHOR ABSTRACT - Desulfitobacterium strains have the ability to dechlorinate halogenated
compounds under anaerobic conditions by
dehalorespiration. The complete genome of the tetrachloroethene
(PCE)-dechlorinating strain Desulfitobacterium hafniense Y51 is a
5,727,534-bp circular chromosome harboring 5,060 predicted protein
coding sequences. This genome contains only two reductive dehalogenase
genes, a lower number than reported in most other dehalorespiring
strains. More than 50 members of the dimethyl sulfoxide reductase
superfamily and 30 paralogs of the flavoprotein subunit of the fumarate
reductase are encoded as well. A remarkable feature of the genome is
the large number of O-demethylase paralogs, which allow utilization of
lignin-derived phenyl methyl ethers as electron donors. The large
genome reveals a more versatile microorganism that can utilize a larger
set of specialized electron donors and acceptors than previously
thought. This is in sharp contrast to the PCE-dechlorinating strain
Dehalococcoides ethenogenes 195, which has a relatively small genome
with a narrow metabolic repertoire. A genomic comparison of these two
very different strains allowed us to narrow down the potential
candidates implicated in the dechlorination process. Our results
provide further impetus to the use of desulfitobacteria as tools for
bioremediation.

<>

<1>Noom, M.C., van den Broek, B., Wuite, G.J.L.
<2>Direct observation of the searching mechanism of restriction enzymes using multiple optical traps.
<3>Biophys. J.
<4>84
<5>304a
<6>2003
<7>Restriction enzymes recognize specific sequences in dsDNA.  Once they find their target
sequence, they cut the DNA at that site.  This cutting is evolved in bacteria to protect
itself against foreign DNA.  The searching mechanism for finding the target sequence is
important, because the number of non-specific sites is usually larger than the number of
specific sites.  Therefore, it's unlikely that the initial encounter of the enzyme with the
DNA will be at the target sequence.  To visualize the possible sliding, hopping and jumping of
a restriction enzyme during the searching process, the translocation of EcoRI enzymes is
analyzed using a three-bead-assay created with multiple optical traps.  In this assay
biotinylated EcoRI enzymes are bound to streptavidin-coated beads.  Such coated beads are
offered to a DNA molecule held between two beads kept in optical tweezers.  Movement of a
coated bead is tracked with video microscopy and the forces exerted by the enzyme on the DNA
are measured using the traps.  Here we report our first results on the movement by EcoRI
during the searching process for the target sequence.  Such direct observation of restriction
enzymes during the searching for and recognition of the target sequence provides us with new
insights into the dynamics of these enzyme-DNA interactions.

<>

<1>Noor, Y.M., Samsulrizal, N.H., Jema'on, N.A., Low, K.O., Ramli, A.N., Alias, N.I., Damis, S.I., Fuzi, S.F., Isa, M.N., Murad, A.M., Raih, M.F., Bakar, F.D., Najimudin, N., Mahadi, N.M., Illias, R.M.
<2>A comparative genomic analysis of the alkalitolerant soil bacterium Bacillus lehensis G1.
<3>Gene
<4>545
<5>253-261
<6>2014
<7>Bacillus lehensis G1 is a Gram-positive, moderately alkalitolerant bacterium
isolated from soil samples. B. lehensis produces cyclodextrin glucanotransferase
(CGTase), an enzyme that has enabled the extensive use of cyclodextrin in
foodstuffs, chemicals, and pharmaceuticals. The genome sequence of B. lehensis G1
consists of a single circular 3.99Mb chromosome containing 4017 protein-coding
sequences (CDSs), of which 2818 (70.15%) have assigned biological roles, 936
(23.30%) have conserved domains with unknown functions, and 263 (6.55%) have no
match with any protein database. Bacillus clausii KSM-K16 was established as the
closest relative to B. lehensis G1 based on gene content similarity and 16S rRNA
phylogenetic analysis. A total of 2820 proteins from B. lehensis G1 were found to
have orthologues in B. clausii, including sodium-proton antiporters, transport
proteins, and proteins involved in ATP synthesis. A comparative analysis of these
proteins and those in B. clausii and other alkaliphilic Bacillus species was
carried out to investigate their contributions towards the alkalitolerance of the
microorganism. The similarities and differences in alkalitolerance-related genes
among alkalitolerant/alkaliphilic Bacillus species highlight the complex
mechanism of pH homeostasis. The B. lehensis G1 genome was also mined for
proteins and enzymes with potential viability for industrial and commercial
purposes.

<>

<1>Nord, D., Sjoberg, B.M.
<2>Unconventional GIY-YIG homing endonuclease encoded in group I introns in closely related strains of the Bacillus cereus group.
<3>Nucleic Acids Res.
<4>36
<5>300-310
<6>2008
<7>Several group I introns have been previously found in strains of the Bacillus cereus group at
three different insertion sites in the nrdE gene
of the essential nrdIEF operon coding for ribonucleotide reductase. Here,
we identify an uncharacterized group IA intron in the nrdF gene in 12
strains of the B. cereus group and show that the pre-mRNA is efficiently
spliced. The Bacillus thuringiensis ssp. pakistani nrdF intron encodes a
homing endonuclease, denoted I-BthII, with an unconventional GIY-(X)8-YIG
motif that cleaves an intronless nrdF gene 7 nt upstream of the intron
insertion site, producing 2-nt 3' extensions. We also found four
additional occurrences of two of the previously reported group I introns
in the nrdE gene of 25 sequenced B. thuringiensis and one B. cereus
strains, and one non-annotated group I intron at a fourth nrdE insertion
site in the B. thuringiensis ssp. Al Hakam sequenced genome. Two strains
contain introns in both the nrdE and the nrdF genes. Phylogenetic studies
of the nrdIEF operon from 39 strains of the B. cereus group suggest
several events of horizontal gene transfer for two of the introns found in
this operon.

<>

<1>Nord, D., Torrents, E., Sjoberg, B.M.
<2>A Functional Homing Endonuclease in the Bacillus anthracis nrdE Group I Intron.
<3>J. Bacteriol.
<4>189
<5>5293-5301
<6>2007
<7>The essential Bacillus anthracis nrdE gene carries a self-splicing group I intron with a
putative homing endonuclease belonging to the GIY-YIG
family. Here, we show that the nrdE pre-mRNA is spliced and that the
homing endonuclease cleaves an intronless nrdE gene 5 nucleotides (nt)
upstream of the intron insertion site, producing 2-nt 3' extensions. We
also show that the sequence required for efficient cleavage spans at least
4 bp upstream and 31 bp downstream of the cleaved coding strand. The
position of the recognition sequence in relation to the cleavage position
is as expected for a GIY-YIG homing endonuclease. Interestingly, nrdE
genes from several other Bacillaceae were also susceptible to cleavage,
with those of Bacillus cereus, Staphylococcus epidermidis (nrdE1), B.
anthracis, and Bacillus thuringiensis serovar konkukian being better
substrates than those of Bacillus subtilis, Bacillus lichenformis, and S.
epidermidis (nrdE2). On the other hand, nrdE genes from Lactococcus
lactis, Escherichia coli, Salmonella enterica serovar Typhimurium, and
Corynebacterium ammoniagenes were not cleaved. Intervening sequences
(IVSs) residing in protein-coding genes are often found in enzymes
involved in DNA metabolism, and the ribonucleotide reductase nrdE gene is
a frequent target for self-splicing IVSs. A comparison of nrdE genes from
seven gram-positive low-G+C bacteria, two bacteriophages, and Nocardia
farcinica showed five different insertion sites for self-splicing IVSs
within the coding region of the nrdE gene.

<>

<1>Nordlund, T.M., Andersson, S., Nilsson, L., Rigler, R.
<2>Structure and dynamics of a fluorescent DNA oligomer containing the EcoRI recognition sequence: fluorescence, molecular dynamics, and NMR studies.
<3>Biochemistry
<4>28
<5>9095-9103
<6>1989
<7>The self-complementary DNA decamer duplex d(CTGAATTCAG)2 and its modified counterpart
d(CTGA[2AP]TTCAG)2, where the innermost adenine (6-aminopurine) has been replaced with the
fluorescent analogue 2-aminopurine (2AP), have been studied by fluorescence and NMR
spectroscopy and simulated by molecular dynamics.  Both decamers are recognized and cleaved by
the EcoRI restriction endonuclease.  2D NMR results show that both decamers have a standard
B-type conformation below 20oC, though a disturbance exists to the 5' side of the 2AP site
which may originate from increased local mobility.  The fluorescence and fluorescence
anisotropy decays of both decamers, as well as the one containing 2AP in only one chain, were
studied as a function of temperature.  The data show that the 2AP base exists in a
temperature-dependent distribution of states and shows rapid motions, suggesting
interconversion among these states on a time scale of about 10^-10s.  The integrated
fluorescence of the decamer with 2AP in both chains shows a large increase around the helix
melting temperature whereas the decamer with one 2AP shows only a mild increase, showing that
the mixed helix has a different structural transition as sensed by the 2AP base.  The data
suggest a model of conformational states which have distinct fluorescence decay times.  The
various states may differ in the degree of base stacking.  Fluctuations in the degree of
stacking of the A or 2AP base are supported by molecular dynamics simulations, which
additionally show that the 2AP-T or A-T base pair hydrogen bonds remain intact during these
large motions.

<>

<1>Norlander, L., Davies, J.K., Hagblom, P., Normark, S.
<2>Deoxyribonucleic acid modifications and restriction endonuclease production in Neisseria gonorrhoeae.
<3>J. Bacteriol.
<4>145
<5>788-795
<6>1981
<7>Modification of gonococcal deoxyribonucleic acid (DNA) was investigated, and the relationship
with endonuclease production was explored.  Both chromosomal and plasmid DNA from different
gonococcal strains, irrespective of their plasmid content, was poorly cleaved by the
restriction endonucleases HaeII, HaeIII, SacII, and BamHI.  The fragment pattern of the Tn3
segment present on the 7.2-kilobase gonococcal resistance plasmid, when compared to its known
DNA sequence, allowed us to conclude that the HaeIII and BamHI resistance was due to
modification of these sites.  A comparison of the fragment pattern of the resistance plasmid,
when isolated from Escherichia coli or Neisseria gonorrhoeae, revealed that the resistance of
HaeII must also be due to modification of its recognition sequence.  Isoschizomers of HaeII
and HaeIII can be found in isolates of N. gonorrhoeae (NgoI and NgoII, respectively).  A new
restriction endonuclease in gonococci, NgoIII, with a specificity similar to SacII, is
reported here.  High-pressure liquid chromatography of gonococcal DNA showed the presence of
5-methylcytosine.  It is suggested that the methylation of cytosine residues in the HaeII
(NgoI), HaeIII (NgoII), and SacII (NgoIII) recognition sites is the basis for the resistance
of gonococcal DNA to cleavage by these enzymes.  This methylation may be part of a
host-restriction modification system.  In two out of five gonococcal strains the sequence
-GATC- was modified.  One strain unable to modify this sequence was a spontaneous mutant of a
strain carrying such a modifying function. [ The enzyme called NgoI in this abstract has been
renamed NgoHI, Jan/1998. ] [ The enzyme called NgoII in this abstract has been renamed NgoHII,
Jan/1998. ] [ The enzyme called NgoIII in this abstract has been renamed NgoKIII, Jan/1998. ]

<>

<1>Normand, P., Gury, J., Pujic, P., Chouaia, B., Crotti, E., Brusetti, L., Daffonchio, D., Vacherie, B., Barbe, V., Medigue, C., Calteau, A., Ghodhbane-Gtari, F., Essoussi, I., Nouioui, I., Abbassi-Ghozzi, I., Gtari, M.
<2>Genome Sequence of Radiation-Resistant Modestobacter marinus Strain BC501, a Representative Actinobacterium That Thrives on Calcareous Stone Surfaces.
<3>J. Bacteriol.
<4>194
<5>4773-4774
<6>2012
<7>Here we report the full genome sequence of Modestobacter marinus strain BC501, an
actinobacterial isolate that thrives on stone surfaces. The generated chromosome
is circular, with a length of 5.57 Mb and a G+C content of 74.13%, containing
5,445 protein-coding genes, 48 tRNAs, and 3 ribosomal operons.

<>

<1>Norton Hughes, C.A., Johnson, R.C.
<2>Identification of a DNA methylase gene homologue in Borrelia burgdorferi.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>125
<6>1994
<7>DNA methylation may play an important role in DNA mismatch repair, DNA replication and
recombination, and gene expression. DNA methylases are enzymes that transfer methyl groups
from donor S-adenosylmethionine to the adenine or cytosine residues within a recognizable DNA
sequence, thereby preventing digestion at the site and protecting host DNA. The dcm gene
product of Escherichia coli methylates the C5 position (m5C) of the internal cytosine in the
sequence CC(A/T)GG. Cytosine (dcm) methylation is prevalent in Borrelia burgdorferi. In this
study, we examine the B. burgdorferi, 297 genome for the presence of the dcm gene homologue.

An oligonucleotide (21 nucleotides) was synthesized to target a consensus sequence for the
prokaryotic cytosine methyltransferases. The oligonucleotide was labeled at the 3' end by the
addition of digoxigenin-11-ddUTP using terminal transferase. The labeled oligonucleotide was
used to probe genome B. burgdorferi, 297 DNA. The probe hybridized strongly to chromosomal DNA
in addition to the ospA/ospB plasmid and a 28.4-kb plasmid. These results suggest that a dcm
homologue is present in B. burgdorferi.


<>

<1>Nosikov, V.V., Braga, E.A., Karlishev, A.V., Zhuze, A.L., Polyanovsky, O.L.
<2>Protection of particular cleavage sites of restriction endonucleases by distamycin A and actinomycin D.
<3>Nucleic Acids Res.
<4>3
<5>2293-2301
<6>1976
<7>It is shown here that distamycin A and actinomycin D can protect the
recognition sites of endo R.EcoRI, EcoRII, HindII, HindIII, HpaI and HpaII from
the attack of these restriction endonucleases.  At proper distamycin
concentrations only two endo R.EcoRI sites of phage lambda DNA are available
for the restriction enzyme - sRI1 and sRI4.  This phenomenon results in the
appearance of large DNA fragments comprising several consecutive fragments of
endo R.EcoRI complete cleavage.  The distamycin fragments isolated from the
agarose gels can be subsequently cleaved by endo R.EcoRI with the yield of the
fragments of complete digestion.  We have compared the effect of distamycin A
and actinomycin D on a number of restriction endonucleases having different
nucleotide sequences in the recogniton sites and established that antibiotic
action depends on the nucleotide sequences of the recognition sites and their
closest environment.

<>

<1>Nosikov, V.V., Sain, B.
<2>Protection of particular endonuclease R.HindIII cleavage sites by distamycin A, propyl-distamycin and netropsin.
<3>Nucleic Acids Res.
<4>4
<5>2263-2273
<6>1977
<7>It is shown that three related antibiotics, distamycin A, propyl-distamycin and netropsin, can
protect certain endo R.HindIII cleavage sites from attack by endonuclease, giving rise, after
endo R.HindIII digestion, to larger DNA fragments.  Bacteriophage lambda DNA has six
recognition sites for HindIII enzyme.  Three of these sites: shindIII 2, 3 and 6 can be
protected from nuclease action by all the antibiotics used.  Propyl-distamycin protects partly
shindIII 5, too.  Netropsin protects partly sites shindIII5 and 4, while distamycin A protects
all the sites but shindIII 1 so the HindII digestion produces only two large fragments of
lambda DNA.

<>

<1>Notani, N.K., Setlow, J.K.
<2>Molecular events accompanying the fixation of genetic information in Haemophilus heterospecific transformation.
<3>J. Bacteriol.
<4>112
<5>751-760
<6>1972
<7>Heterospecific transformation between Haemophilus influenzae and H.
parainfluenzae was investigated by isopycnic analysis of deoxyribonucleic acid
(DNA) extracts of 3H-labeled transforming cells that had been exposed to
32P-labeled, heavy transforming DNA.  The density distribution of genetic
markers from the resident DNA and from the donor DNA was determined by
transformation assay of fractions from CsCl gradients, both species being used
as recipients.  About 50% of the 32P atoms in H. parainfluenzae donor DNA taken
up by H. influenzae cells were transferred to resident DNA, and only a small
amount of the label was lost under conditions of little cell growth.  There was
less transfer in the reciprocal cross, and almost half of the donor label was
lost.  In both crosses,the transferred donor material transformed for the donor
marker considerably more efficiently when assayed on the donor species than on
the recipient species, indicating that at least some of the associated 32P
atoms are contained in relatively long stretches of donor DNA.  When the
transformed cultures were incubated under growth conditions, the donor marker
associated with recipient DNA transformed the donor species with progressively
decreasing efficiency.  The data indicate that the low heterospecific
transformation between H. influenzae and H. parainfluenzae  may be due partly
to events occurring before association of donor and resident DNA but results
mostly from events that occur after the association of the two DNA
preparations.

<>

<1>Noto, M.J., Kreiswirth, B.N., Monk, A.B., Archer, G.L.
<2>Gene acquisition at the insertion site for SCCmec, the genomic island conferring methicillin resistance in Staphylococcus aureus.
<3>J. Bacteriol.
<4>190
<5>1276-1283
<6>2008
<7>Staphylococcus aureus becomes resistant to methicillin by acquiring a
genomic island, known as staphylococcal chromosome cassette mec (SCCmec),
which contains the methicillin resistance determinant, mecA. SCCmec is
site-specifically integrated into the staphylococcal chromosome at a locus
known as the SCCmec attachment site (attB). In an effort to gain a better
understanding of the potential that methicillin-sensitive S. aureus (MSSA)
isolates have for acquiring SCCmec, the nucleotide sequences of attB and
surrounding DNA regions were examined in a diverse collection of 42 MSSA
isolates. The chromosomal region surrounding attB varied among the
isolates studied and appears to be a common insertion point for acquired
foreign DNA. Insertions of up to 15.1 kb were found containing open
reading frames with homology to enterotoxin genes,
restriction-modification systems, transposases, and several sequences that
have not been previously described in staphylococci. Two groups,
containing eight and four isolates, had sequences found in known SCCmec
elements, suggesting SCCmec elements may have evolved through repeated DNA
insertions at this locus. In addition, the attB sequences of the majority
of MSSA isolates in this collection differ from the attB sequences of
strains for which integrase-mediated SCCmec insertion or excision has been
demonstrated, suggesting that some S. aureus isolates may lack the ability
to site-specifically integrate SCCmec into their chromosomes.

<>

<1>Nou, X., Skinner, B., Braaten, B., Blyn, L., Hirsch, D., Low, D.
<2>Regulation of pyelonephritis-associated pili phase-variation in Excherichia coli: binding of the PapI and the Lrp regulatory proteins is controlled by DNA methylation.
<3>Mol. Microbiol.
<4>7
<5>545-553
<6>1993
<7>Expression of pyelonephritis-associated pili (Pap) in Escherichia coli is under a
phase-variation control mechanism in which individual cells alternate between pili+ (ON) and
pili- (OFF) states through a process involving DNA methylation by deoxyadenosine methylase
(Dam). Methylation of two GATC sites (GATC1028 and GATC1130) within the pap regulatory region
is differentially inhibited in phase ON and phase OFF cells. The GATC1028 site of phase ON
cells is non-methylated and the GATC1130 site is fully methylated. Conversely, in phase OFF
cells the GATC1028 site is fully methylated whereas the GATC1130 site is non-methylated. Two
transcriptional activators, PapI and Lrp (leucine-responsive regulatory protein), are required
for this specific methylation inhibition. DNA footprint analysis using non-methylated pap DNAs
indicates that Lrp binds to a region surrounding the GATC1130 site, whereas PapI does not
appear to bind to pap regulatory DNA. However, addition of Lrp and PapI together results in an
additional DNaseI footprint around the GATC1028 site. Moreover, Dam methylation inhibits
binding of Lrp/PapI near the GATC1028 site and alters binding of Lrp at the GATC1130 site. Our
results support a model in which Dam and Lrp/PapI compete for binding near the GATC1028 site,
regulating the methylation state of this GATC site and consequently, the pap transcription
state.

<>

<1>Nouioui, I. et al.
<2>Draft Genome Sequence of Frankia sp. Strain BMG5.12, a Nitrogen-Fixing Actinobacterium Isolated from Tunisian Soils.
<3>Genome Announcements
<4>1
<5>e00468-13
<6>2013
<7>Members of the actinomycete genus Frankia form a nitrogen-fixing symbiosis with 8 different
families of actinorhizal plants. We report a draft genome sequence for
Frankia sp. strain BMG5.12, a nitrogen-fixing actinobacterium isolated from
Tunisian soils with the ability to infect Elaeagnus angustifolia and Myrica gale.

<>

<1>Nowak, A., Kur, J., Gospodarek, E., Bielawski, K.
<2>Characterization of restriction endonuclease AjoI from Acinetobacter johnsonii.
<3>FEMS Microbiol. Lett.
<4>117
<5>97-102
<6>1994
<7>A new type II restriction endonuclease, named AjoI, was detected in Acinetobacter johnsonii.
The enzyme AjoI, an isoschizomer of PstI, recognized the hexanucleotide sequence
[5'-CTGCA/G-3'], with a cleavage site generating fragments of DNA with protruding cohesive
3' termini.

<>

<1>Noy-Malka, C., Yaari, R., Itzhaki, R., Mosquna, A., Gershovitz, N.A., Katz, A., Ohad, N.
<2>A single CMT methyltransferase homolog is involved in CHG DNA methylation and development of Physcomitrella patens.
<3>Plant Mol. Biol.
<4>84
<5>719-735
<6>2014
<7>C-5 DNA methylation is an essential mechanism controlling gene expression and developmental
programs in a variety of organisms. Though the role of DNA methylation has been intensively
studied in mammals and Arabidopsis, little is known about the evolution of this mechanism. The
chromomethylase (CMT) methyltransferase family is unique to plants and was found to be
involved in DNA methylation in Arabidopsis, maize and tobacco. The moss Physcomitrella patens,
a model for early terrestrial plants, harbors a single homolog of the CMT protein family
designated as PpCMT. Our phylogenetic analysis suggested that the CMT family is unique to
embryophytes and its earliest known member PpCMT belongs to the CMT3 subfamily. Thus, P.
patens may serve as a model to study the ancient functions of the CMT3 family. We have
generated a Delta Ppcmt deletion mutant which demonstrated that PpCMT is essential for P.
patens protonema and gametophore development and is involved in CHG methylation as
demonstrated at four distinct genomic loci. PpCMT protein accumulation pattern correlated with
proliferating cells and was sub-localized to the nucleus as predicted from its function. Taken
together, our results suggested that CHG DNA methylation mediated by CMT has been employed
early in land plant evolution to control developmental programs during both the vegetative and
reproductive haploid phases along the plant life cycle.

<>

<1>Noyer-Weidner, M.
<2>A novel mcrB-based Escherichia coli K-12 vector system and its use in analyzing the genetic determinants for the McrB nuclease.
<3>Gene
<4>74
<5>177-178
<6>1988
<7>Meeting Abstract

<>

<1>Noyer-Weidner, M.
<2>System for positive selection of recombinant DNA molecules.
<3>German Patent Office
<4>DE 3802040 A
<5>
<6>1989
<7>Describes a positive selection system for recombinant DNA molecules using the McrA/B systems
of E. coli.

<>

<1>Noyer-Weidner, M., Diaz, R., Reiners, L.
<2>Cytosine-specific DNA modification interferes with plasmid establishement in Escherichia coli K12:  Involvement of rgl B.
<3>Mol. Gen. Genet.
<4>205
<5>469-475
<6>1986
<7>Several chimeric pBR322/328 derivatives containing genes for cytosine-specific
DNA methyltransferases (Mtases) can be transformed into the Escherichia coli
K12/E. coli B hybrid strains HB101 and RR1 but not into other commonly used E.
coli K12 strains.  In vitro methylation of cytosine residues in pBR328 and
other unrelated plasmids also reduces their potential to transform such
methylation sensitive strains, albeit to a lesser degree than observed with
plasmids containing Mtase genes.  The extent of reduced transformability
depends on the target specificity of the enzyme used for in vitro modification.
The role of a host function in the discrimination against methylated plasmids
was verified by the isolation of K12 mutants which tolerate cytosine methylated
DNA.  The mutations map in the vicinity of the serB locus.  This and other data
indicate that the host rg/B function is involved in the discrimination against
modified DNA.

<>

<1>Noyer-Weidner, M., Jentsch, S., Kupsch, J., Bergbauer, M., Trautner, T.A.
<2>DNA methyltransferase genes of Bacillus subtilis phages: structural relatedness and gene expression.
<3>Gene
<4>35
<5>143-150
<6>1985
<7>The DNA methyltransferase (Mtase) genes of temperate Bacillus subtilis phages
Phi3T, Rho11 and SPbeta were cloned and expressed in Escherichia coli.  Each
gene specifies a 47-kDal protein, which modifies BsuR (GGCC) and Fnu4HI (GCNGC)
target sequences.  Transcription is controlled by phage promoters located on
the cloned fragments.  The direction of transcription and the approximate
position of the Mtase genes were determined.  DNA/DNA hybridization experiments
revealed close structural relatedness of the Phi3T, Rho11 and SPbeta genes.  A
significant degree of homology was also found among these genes and the Mtase
gene of related phage SPR, which codes for an enzyme with different
modification specificity.  These results suggest a common ancestor of the
different phage Mtase genes.  Phage Z, the only BsuR-sensitive member of this
phage group, lacks a modification gene, but contains regions homologous to
sequences flanking the SPR, Phi3T, Rho11 and SPbeta Mtase genes.

<>

<1>Noyer-Weidner, M., Jentsch, S., Pawlek, B., Gunthert, U., Trautner, T.A.
<2>Restriction and modification in Bacillus subtilis: DNA methylation potential of the related bacteriophages Z, SPR, SPbeta, Phi3T, and Rho11.
<3>J. Virol.
<4>46
<5>446-453
<6>1983
<7>The DNA methylation capacity and some other properties of the related temperate
Bacillus subtilis phages Z, SPR, SPbeta, Phi3T, and Rho11 are compared.  With
phage mutants affected in their methylation potential, we show that phage-coded
methyltransferase genes are interchangeable among the phages studied.  DNA/DNA
hybridization experiments indicate that phage methyltransferase genes are
structurally related, whereas no such relationship is observed to a bacterial
gene, specifying a methyltransferase with the same specificity.

<>

<1>Noyer-Weidner, M., Pawlek, B., Jentsch, S., Gunthert, U., Trautner, T.A.
<2>Restriction and modification in Bacillus subtilis: gene coding for a BsuR-specific modification methyltransferase in the temperate bacteriophage Phi3T.
<3>J. Virol.
<4>38
<5>1077-1080
<6>1981
<7>The resistance of Phi3T DNA to degradation by the restriction enzyme BsuR or
its isoschizomer HaeIII is due to obligatory modification of such DNA.
Biochemical and genetical experiments indicate that Phi3T codes for a
methyltransferase, which methylates Phi3T DNA itself or heterologous DNA at
target sites 5'-GG*CC.

<>

<1>Noyer-Weidner, M., Reiners-Schramm, L.
<2>Highly efficient positive selection of recombinant plasmids using a novel rglB-based Escherichia coli K-12 vector system.
<3>Gene
<4>66
<5>269-278
<6>1988
<7>We have developed pBR328-derived vectors which allow highly efficient positive
selection of recombinant plasmids.  The system is based on the rglB-coded
restriction activity of Escherichia coli K-12 directed against 5-methylcytosine
(5mC)-containing DNA.  The vectors code for cytosine-specific,
temperature-sensitive DNA methyltransferases (ts-Mtases), whose specificity
elicits RglB restriction.  5mC-free vector DNA - a prerequisite to allow
establishment of such plasmids in cells expressing the RglB nuclease activity -
can be prepared from cultures grown at 42C.  At 30C the vector plasmids are
vulnerable to RglB restriction due to the expression of suicidal Mtase
activity.  Cloning a DNA fragment into the ts-Mtase-coding gene disrupts the
lethal methylation and thus permits selection of such recombinant plasmids at
30C.  The standard vector used, pBN73, contains unique recognition sites for
nine restriction enzymes within the ts-Mtase-coding gene, which can be used
independently or in combination for the construction of recombinant plasmids
selectable by the rglB-coded activity.  Plasmid pBN74, which carries the
determinants for both the ts-Mtase and the RglB nuclease, contains seven unique
sites within the ts-Mtase-coding gene.  While selection of recombinant plasmids
derived from pBN73 obligatorily requires the employment of rglB+ strains,
selection of pBN74 derivatives can be performed independent of the E. coli host
genotype.  It remains to be elucidated whether positive selection of
pBN74-derived recombinant plasmids can also be achieved in hosts other than E.
coli.  Plasmids pBN73, pBN74 and the recombinants are structurally stable.
Generally applicable procedures, as developed during the establishment of this
vector system, are described; they allow the isolation of ts-Mtases and
facilitate the cloning of genes coding for nucleases directed against
5mC-containing DNA.

<>

<1>Noyer-Weidner, M., Trautner, T.A.
<2>Methylation of DNA in prokaryotes.
<3>DNA Methylation: Molecular Biology and Biological Significance, Birkhauser Verlag, Jost, J.P., Saluz, H.P., Basel
<4>0
<5>39-108
<6>1993
<7>A much wider variety of biological functions of postreplicative DNA methylation is observed in
prokaryotes than in eukaryotes. In eukaryotes DNA methylation is primarily a means of the
control of gene expression. Many chapters of this book are devoted to various aspects of this
function. In prokaryotes, DNA methylation affects such diverse phenomena as determination of
accessiblity of DNA to digestion by endonucleases, control of initiation of DNA replication,
and the definition of origins of packaging in the maturation of phage DNA, which will be dealt
with in this article. We shall also be concerned with enzymes which facilitate methylation,
the DNA methyltransferases. In the eukaryotes, as far as we know at this time, the various DNA
methyltransferases encountered represent a rather homogenous group, whereas in prokaryotes, we
find a very diverse set of DNA methyltransferases. Beyond their biological significance, DNA
methyltransferases represent a remarkable class of enzyme in their own right. Not only are
they paradigms for sequence specific DNA binding proteins, but they also show specificity in
their catalytic interaction with defined DNA sequences. Furthermore, their universal
distribution, the multitude of enzymes with different or identical specificities observed
among prokaryotes and the obligatory coexistence of isospecific restriction and methylating
enzymes in restriction/modification systems make DNA methyltransferases choice candidates for
evolutionary studies.

<>

<1>Noyer-Weidner, M., Walter, J., Terschuren, P.-A., Chai, S., Trautner, T.A.
<2>M.Phi3TII: a new monospecific DNA (cytosine-C5) methyltransferase with pronounced amino acid sequence similarity to a family of adenine-N6-DNA-methyltransferases.
<3>Nucleic Acids Res.
<4>22
<5>4066-4072
<6>1994
<7>The temperate B.subtilis phages Phi3T and Rho11s code, in addition to the multispecific DNA
(cytosine-C5) methyltransferases (C5-MTases) M.Phi3TI and M.Rho11sI, which were previously
characterized, for the identical monospecific C5-MTases M.Phi3TII and M.Rho11s.II. These
enzymes modify the C of TCGA sites, a novel target specificity among C5-MTases. The primary
sequence of M.Phi3TII (326 amino acids) shows all conserved motifs typical of the building
plan of C5-MTases. The degree of relatedness between M.Phi3TII and all other mono- or
multispecific C5-MTases ranges from 30-40% amino acid identity. Particularly M.Phi3TII does
not show pronounced similarity to M.Phi3TI indicating that both MTase genes were not generated
from one another but were acquired independently by the phage. The amino terminal part of the
M.Phi3TII (preceding the variable region 'V'), which predominantly constitutes the catalytic
domain of the enzyme, exhibits pronounced sequence similarity to the amino termini of a family
of A-N6-MTases, which -- like M. TaqI -- recognize the general sequence TNNA. This suggests
that recently described similarities in the general three dimensional organization of C5- and
A-N6-MTases imply divergent evolution of these enzymes originating from a common molecular
ancestor.

<>

<1>Noyer-Weidner, M., Walter, J., Terschuren, P.-A., Chai, S., Trautner, T.A.
<2>M.Phi3TII: a new monospecific DNA (cytosine-C5) methyltransferase with pronounced amino acid sequence similarity to a family of adenine-N6-DNA-methyltransferases.
<3>Nucleic Acids Res.
<4>22
<5>5517-5523
<6>1994
<7>The temperate B.subtilis phages Phi3T and Rho11S code, in addition to the multispecific DNA
(cytosine-C5) methyltransferases (C5-MTases) M.Phi3TI and M.Rho11SI, which were previously
characterized, for the identical monospecific C5-MTases M.Phi3TII and M.Rho11SII. These
enzymes modify the C of TCGA sites, a novel target specificity among C5-MTases. The primary
sequence of M.Phi3TII (326 amino acids) shows all conserved motifs typical of the building
plan of C5-MTases. The degree of relatedness between M.Phi3TII and all other mono- or
multispecific C5-MTases ranges from 30-40% amino acid identity. Particularly M.Phi3TII does
not show pronounced similarity to M.Phi3TI indicating that both Mtase genes were not generated
from one another but were acquired independently by the phage. The amino terminal part of the
M.Phi3TII (preceding the variable region 'V'), which predominantly constitutes the catalytic
domain of the enzyme, exhibits pronounced sequence similarity to the amino termini of a family
of A-N6-MTases, which -- like M. TaqI -- recognize the general sequence TNNA. This suggests
that recently described similarities in the general three dimensional organization of C5- and
A-N6-MTases imply divergent evolution of these enzymes originating from a common molecular
ancestor.

<>

<1>Ntougias, S., Lapidus, A., Han, J., Mavromatis, K., Pati, A., Chen, A., Klenk, H.P., Woyke, T., Fasseas, C., Kyrpides, N.C., Zervakis, G.I.
<2>High quality draft genome sequence of Olivibacter sitiensis type strain (AW-6(T)), a diphenol degrader with genes involved in the catechol pathway.
<3>Standards in Genomic Sciences
<4>9
<5>783-793
<6>2014
<7>Olivibacter sitiensis Ntougias et al. 2007 is a member of the family Sphingobacteriaceae,
phylum Bacteroidetes. Members of the genus Olivibacter are
phylogenetically diverse and of significant interest. They occur in diverse
habitats, such as rhizosphere and contaminated soils, viscous wastes, composts,
biofilter clean-up facilities on contaminated sites and cave environments, and
they are involved in the degradation of complex and toxic compounds. Here we
describe the features of O. sitiensis AW-6(T), together with the permanent-draft
genome sequence and annotation. The organism was sequenced under the Genomic
Encyclopedia for Bacteria and Archaea (GEBA) project at the DOE Joint Genome
Institute and is the first genome sequence of a species within the genus
Olivibacter. The genome is 5,053,571 bp long and is comprised of 110 scaffolds
with an average GC content of 44.61%. Of the 4,565 genes predicted, 4,501 were
protein-coding genes and 64 were RNA genes. Most protein-coding genes (68.52%)
were assigned to a putative function. The identification of 2-keto-4-pentenoate
hydratase/2-oxohepta-3-ene-1,7-dioic acid hydratase-coding genes indicates
involvement of this organism in the catechol catabolic pathway. In addition,
genes encoding for beta-1,4-xylanases and beta-1,4-xylosidases reveal the
xylanolytic action of O. sitiensis.

<>

<1>Nunoura, T., Takaki, Y., Kakuta, J., Nishi, S., Sugahara, J., Kazama, H., Chee, G.J., Hattori, M., Kanai, A., Atomi, H., Takai, K., Takami, H.
<2>Insights into the evolution of Archaea and eukaryotic protein modifier systems revealed by the genome of a novel archaeal group.
<3>Nucleic Acids Res.
<4>39
<5>3204-3223
<6>2011
<7>The domain Archaea has historically been divided into two phyla, the Crenarchaeota and
Euryarchaeota. Although regarded as members of the
Crenarchaeota based on small subunit rRNA phylogeny, environmental
genomics and efforts for cultivation have recently revealed two novel
phyla/divisions in the Archaea; the 'Thaumarchaeota' and 'Korarchaeota'.
Here, we show the genome sequence of Candidatus 'Caldiarchaeum
subterraneum' that represents an uncultivated crenarchaeotic group. A
composite genome was reconstructed from a metagenomic library previously
prepared from a microbial mat at a geothermal water stream of a
sub-surface gold mine. The genome was found to be clearly distinct from
those of the known phyla/divisions, Crenarchaeota (hyperthermophiles),
Euryarchaeota, Thaumarchaeota and Korarchaeota. The unique traits suggest
that this crenarchaeotic group can be considered as a novel archaeal
phylum/division. Moreover, C. subterraneum harbors an ubiquitin-like
protein modifier system consisting of Ub, E1, E2 and small Zn RING finger
family protein with structural motifs specific to eukaryotic system
proteins, a system clearly distinct from the prokaryote-type system
recently identified in Haloferax and Mycobacterium. The presence of such a
eukaryote-type system is unprecedented in prokaryotes, and indicates that
a prototype of the eukaryotic protein modifier system is present in the
Archaea.

<>

<1>Nunvar, J., Elhottova, D., Chronakova, A., Schneider, B., Licha, I.
<2>Draft Genome Sequence of Stenotrophomonas maltophilia Strain 5BA-I-2, a Soil Isolate and a Member of a Phylogenetically Basal Lineage.
<3>Genome Announcements
<4>2
<5>e00134-14
<6>2014
<7>Stenotrophomonas maltophilia is an omnipresent environmental bacterium emerging as an
opportunistic human pathogen and exhibiting multidrug resistance. Here, we
report the draft genome sequence of S. maltophilia strain 5BA-I-2, a soil isolate
and a member of a phylogenetically basal lineage.

<>

<1>Nur, I., Szyf, M., Razin, A., Glaser, G., Rottem, S., Razin, S.
<2>Procaryotic and Eucaryotic Traits of DNA Methylation in Spiroplasmas (Mycoplasmas).
<3>J. Bacteriol.
<4>164
<5>19-24
<6>1985
<7>Differences in the type of base methylated (cytosine or adenine) and in the
extent of methylation were detected by high-pressure liquid chromatography in
the DNAs of five spiroplasmas.  Nearest neighbor analysis and digestion by
restriction enzyme isoschizomers also revealed differences in methylation
sequence specificity.  Whereas in Spiroplasma floricola and Spiroplasma sp.
strain PPS-1 5-methylcytosine was found on the 5' side of each of the four
major bases, the cytosine in Spiroplasma apis DNA was methylated only when its
3' neighboring base was adenine or thymine.  In Spiroplasma sp. strain MQ-1
over 95% of the methylated cytosine was in C-G sequences.  Essentially all of
the C-G sequences in the MQ-1 DNA were methylated.  Partially purified extracts
of S. apis and Spiroplasma sp. strain MQ-1 were used to study substrate and
sequence specificity of the methylase activity.  Methylation by the MQ-1 enzyme
was exclusively at C-G sequences, resembling in this respect eucaryotic DNA
methylases.  However, the MQ-1 emthylase differed from eucaryotic methylases by
showing high activity on nonmethylated DNA duplexes, low activity with
hemimethylated DNA duplexes, and no activity on single-stranded DNA.

<>

<1>Nurminsky, D.I., Hartl, D.L.
<2>Design of compact multiple cloning sites.
<3>Biotechniques
<4>15
<5>209-213
<6>1993
<7>Cloning procedures in molecular biology are usually enhanced by the availability of desired
restriction sites in vectors engineering for cloning or expression purposes. In the absence of
such sites, cloning required blunt ends or the addition of linkers, which requires more
manipulation, both being less efficient. During the past decade, many vectors have been
developed that have a multiple cloning site (MCS) containing many restriction sites found only
once within each vector. An ideal MCS would be one that has many unique restriction sites that
would satisfy general cloning needs. The upper limit to the number of restriction sites that
can be placed in an MCS is usually determined by the absence of identical recognition
specificities in the rest of the vector and the functional length available for the MCS. Here
we present a simple method to design compact MCSs which are 50% shorter than presently
available MCSs as a result of the presence of restriction sites that overlap each other.

<>

<1>Nutter, R.L., Bullas, L.R., Siapco, B.J., Pearson, C.
<2>The role of methylation in host-induced modification of Salmonella bacteriophage P3.
<3>Bacteriol. Proc.
<4>71
<5>196
<6>1971
<7>The temperate Salmonella potsdam bacteriophage P3 lyses E. coli K with high
efficiency.  It is modified in this passage and subsequently restricted by S.
potsdam.  Studies with P-32 tagged phage showed that the E. coli passed P3
phage were adsorbing to S. potsdam, however, and infecting their DNA.  When a
methionine requiring mutant of S. potsdam was deprived of methionine and
infected with restricted P3 phage, the restriction was reduced to less than
half of its former value.  Experiments employing adenine-2-H3 revealed that the
unrestricted phage DNA from S. potsdam has about 1.2% as many MAP groups as
adenine whereas the restricted phage DNA from E. coli has about 0.6%.

<>

<1>Nutter, R.L., Bullas, L.R., Siapco, B.S., Pearson, C.A.
<2>Change in methylation of Salmonella bacteriophage P3 deoxyribonucleic acid with host-controlled modification by Escherichia coli.
<3>J. Virol.
<4>10
<5>560-562
<6>1972
<7>Modification of bacteriophage P3 by passage through Escherichia coli K was
correlated with a 54% decrease in the content of 6-methylaminopurine in the
phage deoxyribonucleic acid.

<>

<1>Nwankwo, D., Wilson, G.
<2>Cloning of two type II methylase genes that recognise asymmetric nucleotide sequences:  FokI and HgaI.
<3>Mol. Gen. Genet.
<4>209
<5>570-574
<6>1987
<7>The modification genes of Flavobacterium okeanokoites and Haemophilus
gallinarum have been cloned into the vector of pBR322 and expressed in
Escherichia coli cells.  FokI methylase gene is contained on a 3.80 kb piece of
F. okeanokoites DNA.  Plasmid constructs carrying this fragment of DNA are
resistant to digestion by FokI restriction endonuclease but are sensitive to
cleavage by HindIII, EcoRI and PstI.  Unmodified lambda DNA molecules, exposed
in vitro to cell extracts prepared from cells harbouring this plasmid, became
resistant to digestion by FokI.  The smallest HgaI methylase clone carries the
pBR322 plasmid containing a 3.50 kb piece of H. gallinarum DNA.  This plasmid
is resistant to digestion by HgaI.  Neither the FokI nor the HgaI restriction
endonuclease was detected in either clone.  This is the first report of cloning
modification genes whose protein products recognise asymmetric nucleotide
sequences.

<>

<1>Nwankwo, D.O.
<2>Cloning of a pair of genes encoding isoschizomeric restriction endonucleases from Bacillus species: the BspEI and BspMII restriction and modification systems.
<3>Gene
<4>157
<5>31-35
<6>1995
<7>The respective genes (R-M) encoding restriction and modification systems from two Bacillus
species which recognize the same nucleotide sequence, 5'TCCGGA, have been cloned and
expressed in Escherichia coli.  The BspEI R-M genes were cloned on a 3.6-kb HindIII fragment,
whereas the BspMII R-M genes were cloned on three contiguous HindIII fragments totalling 9.8
kb.  Upon thermal induction, E. coli carrying the bspEIR clones under the control of the phage
lambda PL promoter, express high levels of R.BspEI (106 units/g wet cell paste).  The
bspMIIR  gene, on the other hand, is only poorly expressed (about 4 x 10/3 units/g wet cell
paste) following induction.  Although the enzymes of both R-M systems recognize the same
sequence and  the restriction endonucleases (ENases) cleave DNA at the same position, the
modification specified by the methyltransferases (MTases) differ.  The internal cytosine is
the site for M.BspMII modification (TCmeCGGA), whereas the external cytosine is modified by
M.BspEI.

<>

<1>Nwankwo, D.O., Lynch, J.J., Moran, L.S., Fomenkov, A., Slatko, B.E.
<2>The XmnI restriction-modification system: cloning, expression, sequence organization and similarity between the R and M genes.
<3>Gene
<4>173
<5>121-127
<6>1996
<7>The xmn1RM genes from Xanthomonas manihotis 7AS1 have been cloned and expressed in Escherichia
coli.  The nucleotide (nt) sequences of both genes were determined.  The XmnI
methyltransferase (Mtase)-encoding gene is 1861 bp in length and codes for 620 amino acids
(aa) (68,660 Da).  The restriction endonuclease (Enase)-encoding gene is 969 bp long and
therefore codes for a 319-aa protein (35,275 Da).  The two genes are aligned tail to tail and
they overlap at their respective stop codons.  About 4 x 10^4 units/g wet cell paste of R.XmnI
was obtained following IPTG induction in a suitable E. coli host.  The xmnIR gene is expressed
from the T7 promoter.  M.XmnI probably modifies the first A in the sequence, GAA(N)4TTC.  The
xmnIR and M genes contain regions of conserved similarity and probably evolved from a common
ancestor.  M.XmnI is losely related to M.EcoRI.  The XmnI R-M system and the type-I R-M
systems probably derived from a common ancestor.

<>

<1>Nwankwo, D.O., Maunus, R.E., Xu, S.-Y.
<2>Cloning and expression of AatII restriction-modification system.
<3>Gene
<4>185
<5>105-109
<6>1997
<7>The genes encoding the AatII restriction endonuclease and methylase from Acetobacter aceti
have been cloned and expressed in Escherichia coli.  The nucleotide sequences of aatIIM and
aatIIR genes were determined.  The aatIIM and aatIIR genes are 996 bp and 1038 bp,
respectively, encoding the 331-aa methylase with a predicted molecular mass of 36.9 kDa, and
the 345-aa AatII restriction endonuclease with a predicted molecular mass of 38.9 kDa.  The
two genes overlap by 4 base pairs and are transcribed in the same orientation.  The aatIIRM
genes are located next to a putative gene for plasmid mobilization.  A stable overproducing
strain was constructed, in which the aatIIM gene was expressed from a pSC101-derived plasmid.
The aatIIR gene was inserted into a modified T7 expression vector that carries transcription
terminators upstream from the T7 promoter.  The recombinant AatII restriction endonuclease was
purified to near homogeneity by chromatography through DEAE Sepharose, Heparin Sepharose, and
phosphocellulose columns.

<>

<1>Nwankwo, D.O., Moran, L.S., Slatko, B.E., Waite-Rees, P.A., Dorner, L.F., Benner, J.S., Wilson, G.G.
<2>Cloning, analysis and expression of the HindIII R-M-encoding genes.
<3>Gene
<4>150
<5>75-80
<6>1994
<7>The genes encoding the HindIII restriction endonuclease (R.HindIII ENase) and
methyltransferase (M.HindIII MTase) from Haemophilus influenzae Rd were cloned and expressed
in Escherichia coli and their nucleotide (nt) sequences were determined. The genes are
transcribed in the same orientation, with the ENase-encoding gene (hindIIIR) preceding the
MTase-encoding gene (hindIIIM). The two genes overlap by several nt. The ENase is predicted to
be 300 amino acids (aa) in length (34950 Da); the MTase is predicted to be 309 aa (35550 Da).
The HindIII ENase and MTase activities increased approx. 20-fold when the genes were brought
under the control of an inducible lambda pL promoter. Highly purified HindIII ENase and MTase
proteins were prepared and their N-terminal aa sequences determined. In H. influenzae Rd, the
HindIII R-M genes are located between the holC and valS genes; they are not closely linked to
the HindII R-M genes.

<>

<1>Nwankwo, D.O., Wilson, G.G.
<2>Cloning and expression of the MspI restriction and modification genes.
<3>Gene
<4>64
<5>1-8
<6>1988
<7>The genes for the MspI restriction (R) and modification enzymes (recognition
sequence CCGG) have been cloned into Escherichia coli using the vector pBR322.
Clones carrying both genes have been isolated from libraries prepared with
EcoRI, HindIII and BamHI.  The smallest fragment that encodes both activities
is a 3.6-kb HindIII fragment.  Plasmids purified from the clones are fully
resistant to digestion by MspI, indicating that the modification gene is
functional in E. coli.  The clones remain sensitive to phage infection,
however, indicating that the endonuclease is dysfunctional.  When the R gene is
brought under the control of the inducible leftward promoter from phage lambda,
the level of endonuclease increases and the level of methylase decreases,
suggesting that the genes are transcribed in opposite directions.

<>

<1>Nwosu, V.U.
<2>Overexpression of the wild-type gene coding for Escherichia coli DNA adenine methylase (dam).
<3>Biochem. J.
<4>283
<5>745-750
<6>1992
<7>The gene coding for Escherichia coli dam methylase was isolated from a dam+ K12 strain by the
PCR method. The gene was subcloned into an overexpression vector under the control of the
strong lambdaPL promoter. The resultant construct produced the dam methylase at about 20% of
total cellular protein. Purification of the protein was achieved with two chromatography
columns and yielded 6 mg of pure methylase per gram cell paste. The methylase readily
methylates the synthetic dodecamer GACTGATCAGTC containing its recognition sequence. It also
methylates a synthetic dodecamer containing the EcoRV recognition sequence GATATC. However,
methyl transfer is to the second adenine in the EcoRV sequence.

<>

<1>Nwosu, V.U., Connolly, B.A., Halford, S.E., Garnett, J.
<2>The cloning, purification and characterization of the EcoRV modification methylase.
<3>Nucleic Acids Res.
<4>16
<5>3705-3720
<6>1988
<7>The gene for the EcoRV methylase has been cloned into a plasmid under control
of the strong lambda PL promoter and overexpressed in E. coli.  This plasmid,
pVIC1, gives reliable overexpression of the methylase at levels of about 20% of
total protein.  Maximum yields of soluble protein are achieved after about 6
hours of induction.  If the cells are harvested later than this much of the
enzyme is found in the pellet fraction following centrifugation.  A two column
purification scheme using phosphocellulose and Blue-Sepharose chromatography
has been developed.  This yielded pure methylase in amounts of 5mg per gram E.
coli cell paste.  The enzyme is monomeric and methylates the first
deoxyadenosine residue in its recognition sequence GATATC.

<>

<1>Nyarko, E., Tabata, M., Watanabe, K.
<2>Enhanced DNA cleavage by mercury(II) porphyrin at a low concentration of HaeIII restriction enzyme.
<3>Chem. Lett.
<4>0
<5>932-933
<6>2001
<7>Mercury(II) porphyrin enhanced DNA cleavage in the presence of 0.2 units per microL of HaeIII
at which concentration the restriction enzyme
could not cleave DNA in the absence of the porphyrin. This was ascribed
to the synergistic effect of the bound Hg2+ ions to DNA and the
intercalated free base porphyrin, released from the mercury(II)
porphyrin complex upon binding to DNA, which was confirmed by
UV-visible and CD spectroscopic measurements.

<>

<1>Nyengaard, N., Vogensen, F.K., Josephsen, J.
<2>LlaAI and LlaBI, two type-II restriction endonucleases from Lactococcus lactis subsp. cremoris W9 and W56 recognizing, respectively, 5'-/GATC-3' and 5'-C/TRYAG-3'.
<3>Gene
<4>136
<5>371-372
<6>1993
<7>Two type-II restriction endonucleases have been purified from Lactococus lactis subsp.
cremoris W9 and W56, the strains isolated from a mixed Cheddar starter. Their characterization
showed that LlaAI was an isoschizomer of MboI from Moraxella bovis with the cleaving sequence
5'-/GATC-3' being sensitive to methylation of the adenine residue; LlaBI was an isoschizomer
to SfcI from Streptococcus faecium with the cleaving sequence 5'C/TRYAG-3'. Both LlaAI and
LlaBI restriction-modification (R-M) systems are encoded by the plasmids, respectively, pFW094
and pJW563, protecting the harboring strain against phage attack.

<>

<1>Nyengaard, N., Vogensen, F.K., Josephsen, J.
<2>Restriction-modification systems in Lactococcus lactis.
<3>Gene
<4>157
<5>13-18
<6>1995
<7>Several restriction-modification (R-M) systems have been identified in Lactococcus lactis.
Most of the systems have been plasmid encoded and function as phage-resistance mechanisms. At
least five different type-II R-M systems, LlaAI, LlaBI, LlaCI, LlaDI and LlaEI, were
identified in isolates from a mixed Cheddar starter culture. LlaAI and LlaBI recognized the
DNA sequences 5'-/GATC-3' and 5'-C/TRYAG-3', respectively. The genes coding for the LlaAI
and LlaBI R-M systems have been cloned and sequenced. The LlaAI R-M system had two genes
coding for methyltransferases (MTases) and one gene coding for a restriction endonuclease
(ENase). The MTases showed high homology to the MTases from DpnII. The LlaBI R-M system had
one gene coding for a MTase and one gene coding for an ENase.

<>

<1>Nyengaard, N.R., Falkenberg-Klok, J., Josephsen, J.
<2>Cloning and analysis of the restriction-modification system LlaBI, a bacteriophage resistance system from Lactococcus lactis subsp. cremoris W56.
<3>Appl. Environ. Microbiol.
<4>62
<5>3494-3498
<6>1996
<7>The genes coding for the type II restriction-modification (R/M) system LlaBI, which recognized
the sequence 5'-C/TRYAG-3', have been cloned from a plasmid in Lactococcus lactis subsp.
cremoris W56 and sequenced.  The DNA sequence predicts an endonuclease of 299 amino acids (33
kDa) and a methylase of 580 amino acids (65 kDa).  A 4.0-kb HindIII fragment in pSA3 was able
to restrict bacteriophages, showing that the cloned R/M system can function as a phage defense
mechanism in L. lactis.

<>

<1>Nyyssonen, M., Tran, H.M., Karaoz, U., Weihe, C., Hadi, M.Z., Martiny, J.B., Martiny, A.C., Brodie, E.L.
<2>Coupled high-throughput functional screening and next generation sequencing for identification of plant polymer decomposing enzymes in metagenomic libraries.
<3>Front. Microbiol.
<4>4
<5>282
<6>2013
<7>Recent advances in sequencing technologies generate new predictions and
hypotheses about the functional roles of environmental microorganisms. Yet, until
we can test these predictions at a scale that matches our ability to generate
them, most of them will remain as hypotheses. Function-based mining of
metagenomic libraries can provide direct linkages between genes, metabolic traits
and microbial taxa and thus bridge this gap between sequence data generation and
functional predictions. Here we developed high-throughput screening assays for
function-based characterization of activities involved in plant polymer
decomposition from environmental metagenomic libraries. The multiplexed assays
use fluorogenic and chromogenic substrates, combine automated liquid handling and
use a genetically modified expression host to enable simultaneous screening of
12,160 clones for 14 activities in a total of 170,240 reactions. Using this
platform we identified 374 (0.26%) cellulose, hemicellulose, chitin, starch,
phosphate and protein hydrolyzing clones from fosmid libraries prepared from
decomposing leaf litter. Sequencing on the Illumina MiSeq platform, followed by
assembly and gene prediction of a subset of 95 fosmid clones, identified a broad
range of bacterial phyla, including Actinobacteria, Bacteroidetes, multiple
Proteobacteria sub-phyla in addition to some Fungi. Carbohydrate-active enzyme
genes from 20 different glycoside hydrolase (GH) families were detected. Using
tetranucleotide frequency (TNF) binning of fosmid sequences, multiple enzyme
activities from distinct fosmids were linked, demonstrating how
biochemically-confirmed functional traits in environmental metagenomes may be
attributed to groups of specific organisms. Overall, our results demonstrate how
functional screening of metagenomic libraries can be used to connect microbial
functionality to community composition and, as a result, complement large-scale
metagenomic sequencing efforts.

<>

<1>O'Connell-Motherway, M. et al.
<2>Functional genome analysis of Bifidobacterium breve UCC2003 reveals type IVb tight adherence (Tad) pili as an essential and conserved host-colonization factor.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>11217-11222
<6>2011
<7>Development of the human gut microbiota commences at birth, with bifidobacteria being among
the first colonizers of the sterile newborn gastrointestinal tract. To date, the genetic basis
of Bifidobacterium colonization and persistence remains poorly understood. Transcriptome
analysis of the Bifidobacterium breve UCC2003 2.42-Mb genome in a murine colonization model
revealed differential expression of a type IVb tight adherence (Tad) pilus-encoding gene
cluster designated "tad(2003)." Mutational analysis demonstrated that the tad(2003) gene
cluster is essential for efficient in vivo murine gut colonization, and immunogold
transmission electron microscopy confirmed the presence of Tad pili at the poles of B. breve
UCC2003 cells. Conservation of the Tad pilus-encoding locus among other B. breve strains and
among sequenced Bifidobacterium genomes supports the notion of a ubiquitous pili-mediated host
colonization and persistence mechanism for bifidobacteria.

<>

<1>O'Connell-Motherway, M., O'Driscoll, J., Fitzgerald, G.F., van Sinderen, D.
<2>Overcoming the restriction barrier to plasmid transformation and targeted mutagenesis in Bifidobacterium breve UCC2003.
<3>Micro. Biotech.
<4>2
<5>321-332
<6>2009
<7>In silico analysis of the Bifidobacterium breve UCC2003 genome predicted two distinct loci,
which encode three different restriction/modification systems, each comprising a modification
methylase and a restriction endonuclease.  Based on sequence homology and observed protection
against restriction we conclude that the first restriction endonuclease, designated BbrI, is
an isoschizomer of BbeI, the second, BbrII, is a neoschizomer of SalI, while the third,
BbrIII, is an isoschizomer of PstI.  Expression of each of the B. breve UCC2003
methylase-encoding genes in B. breve JCM 7017 established that BbrII and BbrIII are active and
restrict incoming DNA.  By exploiting knowledge on restriction/modification in B. breve
UCC2003 we successfully increased the transformation efficiency to a level that allows the
reliable generation of mutants by homologous recombination using a non-replicative plasmid.

<>

<1>O'Connell-Motherway, M., Watson, D., Bottacini, F., Clark, T.A., Roberts, R.J., Korlach, J., Garault, P., Chervaux, C., van-Hylckama-Vlieg, J.E., Smokvina, T., van-Sinderen, D.
<2>Identification of Restriction-Modification Systems of Bifidobacterium animalis subsp. lactis CNCM I-2494 by SMRT Sequencing and Associated Methylome Analysis.
<3>PLoS ONE
<4>9
<5>e94875
<6>2014
<7>Bifidobacterium animalis subsp. lactis CNCM I-2494 is a component of a commercialized
fermented dairy product for which beneficial effects on health has
been studied by clinical and preclinical trials. To date little is known about
the molecular mechanisms that could explain the beneficial effects that
bifidobacteria impart to the host. Restriction-modification (R-M) systems have
been identified as key obstacles in the genetic accessibility of bifidobacteria,
and circumventing these is a prerequisite to attaining a fundamental
understanding of bifidobacterial attributes, including the genes that are
responsible for health-promoting properties of this clinically and industrially
important group of bacteria. The complete genome sequence of B. animalis subsp.
lactis CNCM I-2494 is predicted to harbour the genetic determinants for two type
II R-M systems, designated BanLI and BanLII. In order to investigate the
functionality and specificity of these two putative R-M systems in B. animalis
subsp. lactis CNCM I-2494, we employed PacBio SMRT sequencing with associated
methylome analysis. In addition, the contribution of the identified R-M systems
to the genetic accessibility of this strain was assessed.

<>

<1>O'Connor, C.D., Humphreys, G.O.
<2>Expression of the EcoRI restriction-modification system and the construction of positive-selection cloning vectors.
<3>Gene
<4>20
<5>219-229
<6>1982
<7>The genes encoding the EcoRI restriction-modification (R/M) system have been
separately cloned onto compatible plasmids.  We have shown that the EcoRI
restriction gene is expressed in the total absence of methylase enzyme and
confirmed that a temperature-sensitive mutant is defective in EcoRI
modification activity at higher temperatures.  Insertion of transcriptional
terminators into the restriction gene had no detectable effect on EcoRI
modification activity.  This strongly suggests that a separate promoter exists
for the methylase gene.  Analysis of the published sequence shows that the
methylase gene promoter may overlap with the COOH-terminal region of the
endonuclease structural gene.  The temperature-sensitive EcoRI system has been
exploited in the construction of two plasmid cloning vectors, pLV57 and pLV59,
which can be used to select positively for transformants bearing recombinant
plasmids; cloning of a DNA fragment into pLV57 or pLV59 at the unique HindIII,
BglII, or PstI sites inactivates the EcoRI restriction gene and permits the
hybrid plasmid to survive at 37C.  The temperature-sensitive modification
activity of these vectors should also facilitate the introduction of EcoRI
linkers into DNA cloned in this way.

<>

<1>O'Connor, C.D., Metcalf, E., Wrighton, C.J., Harris, T.J.R., Saunders, J.R.
<2>RsrII - a novel restriction endonuclease with a heptanucleotide recognition site.
<3>Nucleic Acids Res.
<4>12
<5>6701-6708
<6>1984
<7>A sequence-specific endonuclease present in extracts of Rhodopseudomonas
sphaeroides 630 has been purified and characterized.  The enzyme, RsrII,
recognises and cleaves the palindromic heptanucleotide sequence:  5' -
CG^G(A/T)CCG - 3' By virtue of its unusual specificity, RsrII cuts most DNA
molecules very infrequently which should facilitate the physical mapping of
large genomes.

<>

<1>O'Connor, C.D., Timmis, K.N.
<2>Highly repressible expression system for cloning genes that specify potentially toxic proteins.
<3>J. Bacteriol.
<4>169
<5>4457-4462
<6>1987
<7>A highly repressible expression vector system that allows the cloning of potentially
deleterious genes has been constructed.  Undesired expression of a cloned gene was prevented
(I) at the level of initiation of transcription, by the presence of the strong but highly
repressible leftward promoter of bacteriophage lambda, lambda PL, and (ii) at the level of
transcript elongation or translation, through synthesis of antisense RNA complementary to the
mRNA of the cloned gene.  The system was tested by measuring the inhibition of expression of
traT, the gene for the TraT major outer membrane lipoprotein.  Direct detection and functional
assays indicated that an essentially complete inhibition of traT expression was obtained.  As
a further test of the system, the gene encoding the EcoRI restriction endonuclease was cloned
in the absence of the gene of the corresponding protective EcoRI modification methylase.
Transformants harboring this construct were only viable when both repression controls were
operational.

<>

<1>O'Connor, C.D., Walker, J.N.B., Saunders, J.R.
<2>RsrII:  a restriction endonuclease with a heptanucleotide recognition sequence.
<3>Methods Enzymol.
<4>155
<5>11-15
<6>1987
<7>The purple nonsulfur photosynthetic bacterium Rhodopseudomonas sphaeroides 630
produces two restriction endonucleases, RsrI and RsrII, that can be isolated
free of contaminating endonucleases and exonucleases.  Although RsrI is an
isoschizomer of the well-characterized Escherichia coli restriction enzyme
EcoRI, RsrII is the only enzyme described to date that recognizes and cleaves a
palindromic heptanucleotide sequence.  We describe here an improved procedure
for the purification of RsrII and discuss some uses of the enzymes.

<>

<1>O'Connor, T.J., Adepoju, Y., Boyd, D., Isberg, R.R.
<2>Minimization of the Legionella pneumophila genome reveals chromosomal regions involved in host range expansion.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>14733-14740
<6>2011
<7>Legionella pneumophila is a bacterial pathogen of amoebae and humans.
Intracellular growth requires a type IVB secretion system that
translocates at least 200 different proteins into host cells. To
distinguish between proteins necessary for growth in culture and those
specifically required for intracellular replication, a screen was
performed to identify genes necessary for optimal growth in nutrient-rich
medium. Mapping of these genes revealed that the L. pneumophila chromosome
has a modular architecture consisting of several large genomic islands
that are dispensable for growth in bacteriological culture. Strains
lacking six of these regions, and thus 18.5% of the genome, were viable
but required secondary point mutations for optimal growth. The
simultaneous deletion of five of these genomic loci had no adverse effect
on growth of the bacterium in nutrient-rich media. Remarkably, this
minimal genome strain, which lacked 31% of the known substrates of the
type IVB system, caused only marginal defects in intracellular growth
within mouse macrophages. In contrast, deletion of single regions reduced
growth within amoebae. The importance of individual islands, however,
differed among amoebal species. The host-specific requirements of these
genomic islands support a model in which the acquisition of foreign DNA
has broadened the L. pneumophila host range.

<>

<1>O'Cuiv, P., Klaassens, E.S., Smith, W.J., Mondot, S., Durkin, A.S., Harkins, D.M., Foster, L., McCorrison, J., Torralba, M., Nelson, K.E., Morrison, M.
<2>Draft Genome Sequence of Enterococcus faecalis PC1.1, a Candidate Probiotic Strain Isolated from Human Feces.
<3>Genome Announcements
<4>1
<5>e00160-12
<6>2013
<7>is commonly isolated from the gastrointestinal tract of healthy infants and adults, where it
contributes to host health and well-being. We describe here the
draft genome sequence of PC1.1, a candidate probiotic strain isolated from human
feces.

<>

<1>O'Cuiv, P., Klaassens, E.S., Smith, W.J., Mondot, S., Durkin, A.S., Harkins, D.M., Foster, L., McCorrison, J., Torralba, M., Nelson, K.E., Morrison, M.
<2>Draft Genome Sequence of Enterococcus faecium PC4.1, a Clade B Strain Isolated from Human Feces.
<3>Genome Announcements
<4>2
<5>e00022-14
<6>2014
<7>Enterococcus faecium is commonly isolated from the human gastrointestinal tract;  however,
important intraspecies variations exist with relevance for host health
and well-being. Here, we describe the draft genome sequence of E. faecium PC4.1,
a clade B strain isolated from human feces.

<>

<1>O'Donnell, M.M., Harris, H.M., O'Toole, P.W., Ross, R.P.
<2>The Genome of the Predominant Equine Lactobacillus Species, Lactobacillus equi, Is Reflective of Its Lifestyle Adaptations to an Herbivorous Host.
<3>Genome Announcements
<4>2
<5>e01155-13
<6>2014
<7>We report the draft genome sequence of Lactobacillus equi strain DPC6820, isolated from equine
feces. L. equi is a predominant Lactobacillus species in the
horse hindgut microbiota. An examination of the genome identified genes and
enzymes highlighting L. equi adaptations to the herbivorous gastrointestinal
tract of the horse, including fructan hydrolases. This genome sequence may help
us further understand the microbial ecology of the equine hindgut and the
influence lactobacilli have on it.

<>

<1>O'Driscoll, J., Fitzgerald, G.F., van Sinderen, D.
<2>A dichotomous epigenetic mechanism governs expression of the LlaJI restriction/modification system.
<3>Mol. Microbiol.
<4>57
<5>1532-1544
<6>2005
<7>The LlaJI restriction/modification (R/M) system is comprised of two 5mC MTase-encoding genes,
llaJIM1 and llaJIM2, and two genes required for
restriction activity, llaJIR1 and llaJIR2. Here, we report the
molecular mechanism by which this R/M system is transcriptionally
regulated. The recognition sequence for the LlaJI MTases was deduced to
be 5'GACGC'3 for M1.LlaJI and 5'GCGTC'3 for M2.LlaJI, thus together
constituting an asymmetric complementary recognition site. Two
recognition sequences for both LlaJI MTases are present within the
LlaJI promoter region, indicative of an epigenetic role. Following in
vivo analysis of expression of the LlaJI promoter, we established that
both LlaJI MTases were required for complete transcriptional
repression. A mutational analysis and DNA binding studies of this
promoter revealed that the methylation of two specific cytosines by
M2.LlaJI within this region was required to trigger the specific and
high affinity binding of M1.LlaJI, which serves to regulate expression
of the LlaJI operon. This regulatory system therefore represents the
amalgamation of an epigenetic stimulation coupled to the formation of a
MTase/repressor:promoter complex.

<>

<1>O'Driscoll, J., Glynn, F., Cahalane, O., O'Connell-Motherway, M., Fitzgerald, G.F., van Sinderen, D.
<2>Lactococcal plasmid pNP40 encodes a novel, temperature-sensitive restriction-modification system.
<3>Appl. Environ. Microbiol.
<4>70
<5>5546-5556
<6>2004
<7>A novel restriction-modification system, designated LlaJI, was identified on pNP40, a
naturally occurring 65-kb plasmid from Lactococcus lactis. The system comprises four adjacent
similarly oriented genes that are predicted to encode two m(5)C methylases and two restriction
endonucleases. The LlaJI system, when cloned into a low-copy-number vector, was shown to
confer resistance against representatives of the three most common lactococcal phage species.
This phage resistance phenotype was found to be strongly temperature dependent, being most
effective at 19 degrees C. A functional analysis confirmed that the predicted
methylase-encoding genes, llaJIM1 and llaJIM2, were both required to mediate complete
methylation, while the assumed restriction enzymes, specified by llaJIR1 and llaJIR2, were
both necessary for the complete restriction phenotype. A Northern blot analysis revealed that
the four LlaJI genes are part of a 6-kb operon and that the relative abundance of the
LlaJI-specific mRNA in the cells does not appear to contribute to the observed
temperature-sensitive profile. This was substantiated by use of a LlaJI promoter-lacZ fusion,
which further revealed that the LlaJI operon appears to be subject to transcriptional
regulation by an as yet unidentified element(s) encoded by pNP40.

<>

<1>O'Driscoll, J., Glynn, F., Fitzgerald, G.F., van Sinderen, D.
<2>Sequence Analysis of the Lactococcal Plasmid pNP40: a Mobile Replicon for Coping with Environmental Hazards.
<3>J. Bacteriol.
<4>188
<5>6629-6639
<6>2006
<7>The conjugative lactococcal plasmid pNP40, identified in Lactococcus lactis subsp.
diacetylactis DRC3, possesses a potent complement of bacteriophage resistance systems, which
has stimulated its application as a fitness-improving, food-grade genetic element for
industrial starter cultures. The complete sequence of this plasmid allowed the mapping of
previously known functions including replication, conjugation, bacteriocin resistance, heavy
metal tolerance, and bacteriophage resistance. In addition, functions for cold shock
adaptation and DNA damage repair were identified, further confirming pNP40's contribution to
environmental stress protection. A plasmid cointegration event appears to have been part of
the evolution of pNP40, resulting in a "stockpiling" of bacteriophage resistance systems.

<>

<1>O'Driscoll, J., Heiter, D.F., Wilson, G.G., Fitzgerald, G.F., Roberts, R., van Sinderen, D.
<2>A genetic dissection of the LlaJI restriction cassette reveals insights on a novel bacteriophage resistance system.
<3>BMC Microbiol.
<4>6
<5>40
<6>2006
<7>ABSTRACT: BACKGROUND: Restriction/modification systems provide the dual function of protecting
host DNA against restriction by methylation of
appropriate bases within their recognition sequences, and restriction of
foreign invading un-methylated DNA, such as promiscuous plasmids or
infecting bacteriphage. The plasmid-encoded LlaJI restriction/modification
system from Lactococcus lactis recognizes an asymmetric, complementary DNA
sequence, consisting of 5-GACGC-3 in one strand and 5-GCGTC-3 in the other
and provides a prodigious barrier to bacteriophage infection. LlaJI is
comprised of four similarly oriented genes, encoding two 5mC-MTases
(M1.LlaJI and M2.LlaJI) and two subunits responsible for restriction
activity (R1.LlaJI and R2.LlaJI). Here we employ a detailed genetic
analysis of the LlaJI restriction determinants in an attempt to
characterize mechanistic features of this unusual hetero-oligomeric
endonuclease. RESULTS: Detailed bioinformatic analysis confirmed the
presence of a conserved GTP binding and hydrolysis domain within the
C-terminal half of the R1.LlaJI amino acid sequence whilst the N-terminal
half appeared to be entirely unique. This domain architecture was
homologous with that of the B subunit of the GTP-dependent,
methyl-specific McrBC endonuclease from E.coli K-12. R1.LlaJI did not
appear to contain a catalytic centre, whereas this conserved motif;
PD...D/EXK, was clearly identified within the amino acid sequence for
R2.LlaJI. Both R1.LlaJI and R2.LlaJI were found to be absolutely required
for detectable LlaJI activity in vivo. The LlaJI restriction subunits were
purified and examined in vitro, which allowed the assignment of R1.LlaJI
as the sole specificity determining subunit, whilst R2.LlaJI is believed
to mediate DNA cleavage. CONCLUSIONS: The hetero-subunit structure of
LlaJI, wherein one subunit mediates DNA binding whilst the other subunit
is predicted to catalyze strand hydrolysis distinguishes LlaJI from
previously characterized restriction-modification systems. Furthermore,
this distinction is accentuated by the fact that whilst LlaJI behaves as a
conventional Type IIA system in vivo, in that it restricts un-methylated
DNA, it resembles the Type IV McrBC endonuclease, an enzyme specific for
methylated DNA. A number of similar restriction determinants were
identified in the database and it is likely LlaJI together with these
homologous systems, comprise a new subtype of the Type II class
incorporating features of Type II and Type IV systems.

<>

<1>O'Gara, M., Adams, G.M., Gong, W., Kobayashi, R., Blumenthal, R.M., Cheng, X.
<2>Expression, purification, mass spectrometry, crystallization and multiwavelength anomalous diffraction of selenomethionyl PvuII DNA methyltransferase (cytosine-N4-specific).
<3>Eur. J. Biochem.
<4>247
<5>1009-1018
<6>1997
<7>The type II DNA-methyltransferase (cytosine N4-specific) M.PvuII was overexpressed in
Escherichia coli, starting from the internal translation initiator at Met14.  Selenomethionine
was efficiently incorporated into this short form of M.PvuII by a strain prototrophic for
methionine.  Both native and selenomethionyl M.PvuII were purified to apparent homogeneity by
a two-column chromatography procedure.  The yield of purified protein was approximately 1.8
mg/g bacterial paste.  Mass spectrometry analysis of selenomethionyl M.PvuII revealed three
major forms that probably differ in the degree of selenomethionine incorporation and the
extent of selenomethionine oxidation.  Amino acid sequencing and mass spectrometry analysis of
selenomethionine-containing peptides suggests that Met30, Met51, and Met261 were only
partially replaced by selenomethionine.  Furthermore, amino acid 261 may be preferentially
oxidized in both native and selenomethionyl form.  Selenomethionyl and native M.PvuII were
crystallized separately as binary complexes of the methyl donor S-adenosyl-L-methionine in the
monoclinic space group P21.  Two complexes were present per asymmetric unit.  Six out of nine
selenium positions (per molecule), including the three that were found to be partially
substituted, were identified crystallographically.

<>

<1>O'Gara, M., Horton, J.R., Roberts, R.J., Cheng, X.
<2>Structures of HhaI methyltransferase complexed with substrates containing mismatches at the target base.
<3>Nat. Struct. Biol.
<4>5
<5>872-877
<6>1998
<7>Three structures have been determined for complexes between HhaI methyltransferase (M.HhaI)
and oligonucleotides containing a G:A, G:U or G:AP (AP = abasic or apurinic/apyrimidinic)
mismatch at the target base pair. The mismatched adenine, uracil and abasic site are all
flipped out of the DNA helix and located in the enzyme's active-site pocket, adopting the
same conformation as in the flipped-out normal substrate. These results, particularly the
flipped-out abasic deoxyribose sugar, provide insight into the mechanism of base flipping. If
the process involves the protein pushing the base out of the helix, then the push must take
place not on the base, but rather on the sugar-phosphate backbone. Thus rotation of the DNA
backbone is probably the key to base flipping.

<>

<1>O'Gara, M., Klimasauskas, S., Roberts, R.J., Cheng, X.
<2>Enzymatic C5-cytosine methylation of DNA: Mechanistic implications of new crystal structures for HhaI methyltransferase-DNA-AdoHcy complexes.
<3>J. Mol. Biol.
<4>261
<5>634-645
<6>1996
<7>The refined crystal structures of HhaI methyltransferase complexed with cognate unmethylated
or methylated DNA together with S-adenosyl-L-homocysteine, along with the previously-solved
binary and covalent ternary structures, offer a detailed picture of the active site at
individual stages throughout the reaction cycle.  This picture supports and extends a proposed
mechanism for C5-cytosine methylation that may be general for the whole family of C5-cytosine
methyltransferases.  The structures of the two new complexes have been refined to
crystallographic R-factors of 0.189 and 0.178, respectively, at 2.7 A resolution.  We observe
that both unmethylated 2'-deoxycytidine and 5-methyl-2'-deoxycytidine flip out of the DNA
helix and fit into the active site of the enzyme.  The catalytic sulfur atom of Cys81
interacts strongly with C6.  The C5 methyl group of the flipped 5-methyl-2'-deoxycytidine is
bent ~50o out of the plane of the cytosine ring and towards the sulfur atom of
S-adenosyl-L-homocysteine.  This unusual position is probably due to partial sp3 character at
C5 and C6 and to steric effects of the conserved amino acid residues Pro80 and Cys81.  Two
water molecules are held near the hydrophobic edge (C5 and C6) of the flipped cytosine by two
conserved amino acid residues (Gln82 and Asn304) and the phosphoryl oxygen atom of the
phosphate group 3' to the flipped nucleotide, and one of them may serve as the general base
for eliminating the proton from C5.  Protonation of the cytosine N3 during the methylation
reaction may involve Glu119, which itself might be protonated via a water-mediated interaction
between the terminal carboxyl group of Glu119 and the amino group of the methionine moiety of
S-adenosyl-L-methionine.  The cofactor thus plays two key roles in the reaction.

<>

<1>O'Gara, M., McCloy, K., Malone, T., Cheng, X.
<2>Structure-based sequence alignment of three AdoMet-dependent DNA methyltransferases.
<3>Gene
<4>157
<5>135-138
<6>1995
<7>M.HhaI, M.TaqI and COMT are DNA methyltransferases (MTases) which catalyze the transfer of a
methyl group from the cofactor AdoMet to C5 of cytosine, to N6 of adenine and to a hydroxyl
group of catechol, respectively.  The larger catalytic domains of the bilobal proteins, M.HhaI
and M.TaqI, and the entire single domain of COMT have an alpha/beta structure containing a
mixed central beta-sheet.  These domains have very similar folding.  By allowing appropriate
'insertions' or 'deletions' in the bakbones of the three structures, it was possible to
find more conserved motifs in M.TaqI and COMT.  The similarity in protein folding and the
equivalence of amino-acid sequences revealed by the structural alignment indicate that many
AdoMet-dependent MTases may share a common catalytic domain structure.

<>

<1>O'Gara, M., Roberts, R.J., Cheng, X.
<2>A structural basis for the preferential binding of hemimethylated DNA by HhaI DNA methyltransferase.
<3>J. Mol. Biol.
<4>263
<5>597-606
<6>1996
<7>The crystal structure of HhaI methyltransferase complexed with non-palindromic duplex DNA,
containing a hemimethylated recognition sequence, and with the cofactor analog
S-adenosyl-L-homocysteine (AdoHcy), has been determined.  The structure provides an
explanation for the stronger affinities of DNA methyltransferases for hemimethylated DNA than
for unmethylated or fully methylated DNA in the presence of AdoHcy.  The unmethylated target
2'-deoxycytidine flips out of the DNA helix and the CH group at position 5 makes van der
Waals' contacts with the sulfur atom of AdoHcy.  Selectivity/preference for hemimethylated
over fully methylated DNA may thus reflect interactions among the chemical substituent (H or
CH3) at the C5 position of the flipped cytosine, protein and the bound AdoHcy.  The
5-methyl-2'-deoxycytidine on the complementary strand remains in the DNA helix, with the
methyl group almost perpendicular to the carboxylate group of Glu239, which is part of the
sequence recognition loop.  Thus, selectivity/preference for hemimethylated over unmethylated
DNA appears to result largely from van der Waals' contacts between the planar Glu239
carboxylate and the methyl group of the 5-methyl-2'-deoxycytidine.  Furthermore, the positive
electrostatic potential originating from the bound AdoHcy extends to the DNA phosphate groups
flanking the flipped cytosine.  The increased binding to DNA by long-range electrostatic
interactions should also occur with the methyl donor S-adenosyl-L-methionine.

<>

<1>O'Gara, M., Zhang, X., Roberts, R.J., Cheng, X.
<2>Structure of a binary complex of HhaI methyltransferase with S-adenosyl-L-methionine formed in the presence of a short non-specific DNA oligonucleotide.
<3>J. Mol. Biol.
<4>287
<5>201-209
<6>1999
<7>We have determined a structure for a complex formed between HhaI methyltransferase and
S-adenosyl-L-methionine in the presence of a non-specific short oligonucleotide.  M.HhaI binds
to the non-specific short oligonucleotides in solution.  Although no DNA is incorporated in
the crystal, AdoMet binds in a primed orientation, identical with that observed in the ternary
complex of the enzyme, cognate DNA, and AdoMet or S-adenosyl-L-homocysteine.  This orientation
differs from the previously observed unprimed orientation in the M.HhaI-AdoMet binary complex,
where the S+-CH3 unit of AdoMet is protected by a favorable cation-pi interaction with Trp41.
The structure suggests that the presence of DNA can guide AdoMet into the primed orientation.
These results shed new light on the proposed ordered mechanism of binding and explains the
stable association between Ado-Met and M.HhaI.

<>

<1>O'Loughlin, T.J., Xu, Q., Kucera, R.B., Dorner, L.F., Sweeney, S., Schildkraut, I., Guo, H.-C.
<2>Crystallization and preliminary X-ray diffraction analysis of MspI restriction endonuclease in complex with its cognate DNA.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>56
<5>1652-1655
<6>2000
<7>The MspI restriction endonuclease is a type II restriction enzyme. Unlike all other
restriction enzymes with known structures, MspI recognizes the palindromic tetranucleotide
sequence 5'-C/CGG and cleaves it as indicated by the '/' to produce DNA products with 5'
two-base overhangs. Owing to the nature of its cleavage pattern, it is likely that MspI would
represent a new structural class of restriction endonucleases.  Crystals of the dimeric MspI
restriction enzyme bound to a duplex DNA molecule containing the specific recognition sequence
have been obtained by vapor-diffusion techniques in the presence of polyethylene glycol as
precipitant. The crystals belong to the monoclinic space group P2(1), with unit-cell
parameters a = 50.2, b = 131.6, c = 59.3 Angstroms, beta = 109.7 degrees. The crystals contain
one dimeric complex in the asymmetric unit.  A complete native data set has been collected to
a resolution of 2.05 Angstroms by cryo-crystallographic methods, with an R(merge) of 4.0%.

<>

<1>O'Neill, M., Chen, A., Murray, N.E.
<2>The restriction-modification genes of Escherichia coli K-12 may not be selfish: They do not resist loss and are readily replaced by alleles conferring different specificities.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>14596-14601
<6>1997
<7>Type II restriction and modification genes have been described as selfish because they have
been shown to impose selection for the maintenance of the plasmid that encodes them. In our
experiments, the type I R-M system EcoKI does not behave in the same way.  The genes
specifying EcoKI are, however, normally residents of the chromosome and therefore our analyses
were extended to monitor the deletion of chromosomal genes rather than loss of plasmid vector.
If EcoKI were to behave in the same way as the plasmid-encoded type II R-M systems, the loss
of the relevant chromosomal genes by mutation of recombination should lead to cell death
because the cell would become deficient in modification enzyme and the bacterial chromosome
would be vulnerable to the restriction endonuclease.  Our data contradict this prediction;
they reveal that functional type I R-M genes in the chromosome are readily replaced by mutant
alleles and by alleles encoding a type I R-M system of different specificity.  The acquisition
of allelic genes conferring a new sequence specificity, but not the loss of the resident
genes, is dependent on the product of an unlinked gene, one predicted to be relevant to
control of expression of the genes that encode EcoKI.  Our evidence suggests that not all R-M
systems are evolving as "selfish" units; rather, the diversity and distribution of the family
of type I enzymes we have investigated require an alternative selective pressure.

<>

<1>O'Neill, M., Dryden, D.T.F., Murray, N.E.
<2>Localization of a protein-DNA interface by random mutagenesis.
<3>EMBO J.
<4>17
<5>7118-7127
<6>1998
<7>The type I restriction and modification enzymes do not possess obvious DNA-binding motifs
within their target recognition domains of 150-180 amino acids.  To identify residues involved
in DNA recognition, changes were made in the amino-TRD of EcoKI by random mutagenesis.  Most
of the 101 substitutions affecting 79 residues had no effect on the phenotype.  Changes at
only seven positions caused the loss of restriction and modification activities.  The seven
residues identified by mutation are not randomly distributed throughout the primary sequence
of the TRD: five are within the interval between residues 80 and 110.  Sequence analyses have
led to the suggestion that the TRDs of type I restriction enzymes include a tertiary structure
similar to the TRD of the HhaI methyltransferase, and to a model for a DNA-protein interface
in EcoKI.  In this model, the residues within the interval identified by the five mutations
are close to the protein-DNA interface.  Three additional residues close to the DNA in the
model were changed; each substitution impaired both activities.  Proteins from twelve mutants
were purified: six from mutants with partial or wild-type activity and six from mutants
lacking activity.  There is a strong correlation between phenotype and DNA-binding affinity,
as determined by fluorescence anisotropy.

<>

<1>O'Neill, M., Powell, L.M., Murray, N.E.
<2>Target recognition by EcoKI: the recognition domain is robust and restriction-deficiency commonly results from the proteolytic control of enzyme activity.
<3>J. Mol. Biol.
<4>307
<5>951-963
<6>2001
<7>We report a genetic and biochemical analysis of a target recognition domain (TRD) of EcoKI, a
type I restriction and modification enzyme. The TRDs of type I R-M systems are within the
specificity subunit (HsdS) and HsdS confers sequence specificity to a complex endowed with
both restriction and modification activities. Random mutagenesis has revealed that most
substitutions within the amino TRD of EcoKI, a region comprising 157 amino acid residues, have
no detectable effect on the phenotype of the bacterium, even when the substitutions are non-
conservative. The structure of the TRD appears to be robust. All but one of the six
substitutions that confer a restriction-deficient, modification-deficient (r(-)m(-)) phenotype
were found to be in the interval between residues 80 and 110, a region predicted by sequence
comparisons to form part of the protein-DNA interface. Additional site-directed mutations
affecting this interval commonly impair both restriction and modification. However, we show
that an r(-) phenotype cannot be taken as evidence that the EcoKI complex lacks endonuclease
activity; in response to even a slightly impaired modification efficiency, the endonuclease
activity of EcoKI is destroyed by a process dependent upon the ClpXP protease. Enzymes from
mutants with an r(-)m(-) phenotype commonly retain some sequence-specific activity; methylase
activity can be detected on hemimethylated DNA substrates and residual endonuclease activity
is implied whenever the viability of the r(-)m(-) bacterium is dependent on ClpXP. Conversely,
the viability of ClpX(-) r(-)m(-) bacteria can be used as evidence for little, or no,
endonuclease activity. Of 14 mutants with an r(-)m(-) phenotype, only six are viable in the
absence of ClpXP. The significance of four of the six residues (G91, G105, F107 and G141) is
enhanced by the finding that even conservative substitutions for these residues impair
modification, thereby conferring an r(-)m(-) phenotype.

<>

<1>O'Sullivan, D., Coffey, A., Fitzgerald, G.F., Hill, C., Ross, R.P.
<2>Design of a phage-insensitive lactococcal dairy starter via sequential transfer of naturally occurring conjugative plasmids.
<3>Appl. Environ. Microbiol.
<4>64
<5>4618-4622
<6>1998
<7>The plasmid-free Lactococcus lactis subsp. cremoris MG1614 is highly phage sensitive and lacks
lactose fermenting ability (Lac) and primary casein degrading ability (Prt). Food grade gene
transfer systems were used to sequentially superimpose different phage defense systems on this
background, resulting in a gradual increase in resistance to bacteriophage in the derivatives.
pLP712, encoding Lac and Prt, was then transferred to one of these hosts, into which plasmids
encoding adsorption inhibition, restriction modification, and abortive infection had already
been introduced. This resulted in a phage-resistant strain which was successfully used as a
single-strain starter for cheddar cheese manufacture under industrial conditions.

<>

<1>O'Sullivan, D., Ross, R.P., Twomey, D.P., Fitzgerald, G.F., Hill, C., Coffey, A.
<2>Naturally occurring lactococcal plasmid pAH90 links bacteriophage resistance and mobility functions to a food-grade selectable markerpotential use to impart phage resistance to dairy fermentation starter culture.
<3>Appl. Environ. Microbiol.
<4>67
<5>929-937
<6>2001
<7>Phage resistance plasmid pAH90 (26,490 bp) of Lactococcus lactis subsp. lactis biovar.
diacetylactis DPC721 is a natural cointegrate plasmid
formed by homologous recombination between the type I
restriction-modification specificity determinants (hsdS) of plasmid
pAH33 (6,159 bp) and plasmid pAH82 (20,331 bp), giving rise to a
phage-sensitive mutant following phage challenge. The recombinant event
is favored by phage infection. The nucleotide sequence of pAH90 was
determined, identifying 24 open reading frames responsible for
restriction-modification, phage adsorption inhibition, plasmid
replication, cadmium resistance, cobalt transport and conjugative
mobilization phenotypes. The cadmium resistance property, encoded by
the cadA gene, facilitated the selection of pAH90 in other
phage-sensitive lactococci after electroporation. The fortuitous
association of multiple phage resistance systems, which acted at
different stages in the phage lytic cycle, with a food-grade selectable
marker on a mobilizable plasmid makes pAH90 an ideal candidate for use
in food-grade starter culture improvement in industrial dairy
fermentations.

<>

<1>O'Sullivan, D., Twomey, D.P., Coffey, A., Hill, C., Fitzgerald, G.F., Ross, R.P.
<2>Novel type I restriction specificities through domain shuffling of HsdS subunits in lactococcus lactis.
<3>Mol. Microbiol.
<4>36
<5>866-875
<6>2000
<7>This study identifies a natural system in Lactococcus lactis, in which a restriction
modification specificity subunit resident on a 6159 bp plasmid (pAH33) alters the specificity
of a functional R/M mechanism encoded by a 20.3 kb plasmid, pAH82. The new specificity was
identified after phenotypic and molecular analysis of a 26.5 kb co-integrate plasmid (pAH90),
which was detected after bacteriophage challenge of the parent strain. Analysis of the regions
involved in the co-integration revealed that two novel hybrid hsdS genes had been formed
during the co-integration event. The HsdS chimeras had interchanged the C- and N-terminal
variable domains of the parent subunits, generating two new restriction specificities.
Comparison of the parent hsdS genes with other type I specificity determinants revealed that
the region of the hsdS genes responsible for the co-integration event is highly conserved
among lactococcal type I hsdS determinants. Thus, as hsdS determinants are widespread in the
genus Lactococcus, new restriction specificities may evolve rapidly after homologous
recombination between these genes. This study demonstrates that, similar to previous
observations in Gram-negative bacteria, a Gram-positive bacterium can acquire novel
restriction specificities naturally through domain shuffling of resident HsdS subunits.

<>

<1>O'Sullivan, D.J., Klaenhammer, T.R.
<2>C.LlaI is a bifunctional regulatory protein of the LlaI restriction modification operon from Lactococcus lactis.
<3>Dev. Biol. Stand.
<4>85
<5>591-595
<6>1995
<7>Strains of Lactococcus lactis are used commercially as starter bacteria in dairy
fermentations.  Bacteriophage attack of these bacteria represents a major problem in the
industry.  Plasmid-borne phage resistance traits have been found naturally in some strains and
the mobilization of these plasmids into desired industrial starter strains has provided a
dynamic new source of cultures in the industry.  One of these plasmids, pTR2030, has been
studied extensively in this laboratory and shown to encode two phage-resistance determinants;
the AbiA abortive infection mechanism and the LlaI restriction modification system.  The LlaI
methylase is a bifunctional type IIS methylase with 39% identity to M.FokI and is encoded by
1.9 kb on pTR2030, ~5 kb upstream from the abiA gene.  The LlaI restriction component is
encoded by three genes, llaI.1, llaI.2, llaI.3, positioned downstream from llaIM. A
GTP-binding site on the deduced protein from llaI.2 suggests that LlaI restriction is probably
energy-dependent.  Data bank searches did not reveal any significant homologies with these
deduced proteins, except for the GTP-binding motif on LlaI.2 with a corresponding motif on the
Escherichia coli McrB protein, which is part of a GTP-dependent, multi-subunit restriction
enzyme.  From the molecular studies on the LlaI R/M system, it is suggested that the LlaI
restriction enzyme is a novel type with properties reminiscent of both type IIS and type I
enzymes.

<>

<1>O'Sullivan, D.J., Klaenhammer, T.R.
<2>Control of expression of LlaI restriction in Lactococcus lactis.
<3>Mol. Microbiol.
<4>27
<5>1009-1020
<6>1998
<7>The plasmid encoded LlaI R/M sytem from Lactococcus lactis ssp. lactis consists of a bidomain
methylase, with close evolutionary ties to type IIS methylases, and a tri-subunit restriction
complex.  Both the methylase and restriction subunits are encoded on a polycistronic 6.9 kb
operon.  In this study, the 5' end of the llaI 6.9 kb transcript was determined by primer
extension analysis to be 254 bp upstream from the first R/M gene on the operon, llaIM.
Deletion of this promoter region abolished LlaI restriction in L. lactis.  Analysis of the
intervening sequence revealed a 72-amino-acid open reading frame, designated llaIC, with a
conserved ribosome binding site and helix-turn-helix domain.  Overexpression of llaIC in
Escherichia coli with a T7 expression vector produced the predicted protein of 8.2 kDa.
Mutation and in trans complementation analysis indicated that C.LlaI positively enhanced LlaI
restriction activity in vivo.  Northern analysis and transcriptional fusions of the llaI
promoter to a lacZ reporter gene indicated that C.LlaI did not enhance transcription of the
llaI operon.  Databank searches with the deduced protein sequence for llaIC revealed
significant homologies to the E. coli Rop regulatory and mRNA stabilizer protein.
Investigation of the effect of C.LlaI on enhancement of LlaI restriction in L. lactis revealed
that growth at elevated temperatures (40oC) completely abolished any enhancement of
restriction activity.  These data provide molecular evidence for a mechanism on how the
expression of a restriction system in a prokaryote can be drastically reduced during elevated
growth temperatures, by a small regulatory protein.

<>

<1>O'Sullivan, D.J., Zagula, K., Klaenhammer, T.R.
<2>In vivo restriction by LlaI is encoded by three genes, arranged in an operon with llaIM, on the conjugative Lactococcus plasmid pTR2030.
<3>J. Bacteriol.
<4>177
<5>134-143
<6>1995
<7>The LlaI restriction and modification (R/M) system is encoded on pTR2030, a 46.2-kb
conjugative plasmid from Lactococcus lactis. The llaI methylase gene, sequenced previously,
encodes a functional type IIS methylase and is located approximately 5 kb upstream from the
abiA gene, encoding abortive phage resistance. In this study, the sequence of the region
between llaIM and abiA was determined and revealed four consecutive open reading frames
(ORFs). Northern (RNA) analysis showed that the four ORFs were part of a 7-kb operon with llaM
and the downstream abiA gene on a separate transcriptional unit. The deduced protein sequence
of ORF2 revealed a P-loop consensus motif for ATP/GTP-binding sites and a three-part consensus
motif for GTP-binding proteins. Data bank searches with the deduced protein sequences for all
four ORFs revealed no homology except for ORF2 with McrB, in three regions that coincided with
the GTP-binding motifs in both proteins. To phenotypically analyze the llaI operon, a 9.0-kb
fragment was cloned into a high-copy-number lactococcal shuttle vector, pTRKH2. The resulting
construct, pTRK370, exhibited a significantly higher level of in vivo restriction and
modification in L. lactis NCK203 than the low-copy-number parental plasmid, pTR2030. A
combination of deletion constructions and frameshift mutations indicated that the first three
ORFs were involved in LlaI restriction, and they were therefore designated llaI.1, llaI.2, and
llaI.3. Mutating llaI.1 completely abolished restriction, while disrupting llaI.2 or llaI.3
allowed an inefficient restriction of phage DNA to occur, manifested primarily by a variable
plaque phenotype. ORF4 had no discernible effect on in vivo restriction. A frameshift mutation
in llaIM proved lethal to L. lactis NCK203, implying that the restriction component was active
without the modification subunit. These results suggested that the LlaI R/M system is unlike
any other R/M system studied to date and has diverged from the type IIS class of restriction
enzymes by acquiring some characteristics reminiscent of type I enzymes.

<>

<1>O'Sullivan, O., O'Callaghan, J., Sangrador-Vegas, A., McAuliffe, O., Slattery, L., Kaleta, P., Callanan, M., Fitzgerald, G.F., Ross, R.P., Beresford, T.
<2>Comparative genomics of lactic acid bacteria reveals a niche-specific gene set.
<3>BMC Microbiol.
<4>9
<5>50
<6>2009
<7>Background: The recently sequenced genome of Lactobacillus helveticus DPC4571 [1] revealed a
dairy organism with significant homology (75% of
genes are homologous) to a probiotic bacteria Lb. acidophilus NCFM [2].
This led us to hypothesise that a group of genes could be determined
which could define an organism's niche.
Results: Taking 11 fully sequenced lactic acid bacteria (LAB) as
our target, (3 dairy LAB, 5 gut LAB and 3 multi-niche LAB), we
demonstrated that the presence or absence of certain genes involved in
sugar metabolism, the proteolytic system, and restriction modification
enzymes were pivotal in suggesting the niche of a strain. We identified
9 niche specific genes, of which 6 are dairy specific and 3 are gut
specific. The dairy specific genes identified in Lactobacillus
helveticus DPC4571 were lhv 1161 and lhv 1171, encoding components of
the proteolytic system, lhv 1031 lhv 1152, lhv 1978 and lhv 0028
encoding restriction endonuclease genes, while bile salt hydrolase
genes lba 0892 and lba 1078, and the sugar metabolism gene lba 1689
from Lb. acidophilus NCFM were identified as gut specific genes.
Conclusion: Comparative analysis revealed that if an organism had
homologs to the dairy specific geneset, it probably came from a dairy
environment, whilst if it had homologs to gut specific genes, it was
highly likely to be of intestinal origin.
We propose that this "barcode" of 9 genes will be a useful initial
guide to researchers in the LAB field to indicate an organism's ability
to occupy a specific niche.

<>

<1>O'Sullivan, T., van Sinderen, D., Fitzgerald, G.
<2>Structural and functional analysis of pCI65st, a 6.5 kb plasmid from Streptococcus thermophilus NDI-6.
<3>Microbiology
<4>145
<5>127-134
<6>1999
<7>The 6.5 kb cryptic plasmid pCI65st from Streptococcus thermophilus NDI-6, a strain isolated
from the Indian fermented milk dahi, was subcloned and sequenced. Five putative ORFs were
identified. ORF1 could encode a 315 aa polypeptide almost identical to the RepA protein of
previously sequenced S. thermophilus plasmids, indicating that pCI65st is one of the pC194
group of small gram-positive rolling-circle plasmids. ORFs 2 and 4 were virtually identical
and could specify proteins of approximately 150 aa with significant similarity to the small
heat-shock proteins described from a variety of gram-positive bacteria. ORF3 could encode a
415 aa protein similar to enolase, an enzyme involved in glycolysis and gluconeogenesis. ORF5
could encode a 412 aa protein which had high similarity to the HsdS (specificity) proteins of
type I restriction-modification systems. Variants of strain NDI-6 which lacked pCI65st were
readily isolated after subculture of the parent strain at 32 degrees C. The plasmid-bearing
parent culture was significantly more resistant to a temperature shift from 42 degrees C to 62
degrees C than its plasmid-free variant and expressed proteins which corresponded with the
predicted translation products from ORF2 and ORF4. In addition, plasmid-free mutants were
lysed in broth by bacteriophages to which the parent culture was resistant.

<>

<1>O'Toole, P., Fitzgerald, G.F.
<2>A product.
<3>International Patent Office
<4>WO 2007020617 A
<5>
<6>2007
<7>The present invention relates to the identification and isolation of genes in the
Lactobacillus salivarius genome involved in probiotic activity or function, and a novel
megaplasmid that may be exploited for strain alteration.

<>

<1>O'Toole, R.F., Johari, B.M., Mac, A.M., Rogers, T.R., Bower, J.E., Basu, I., Freeman, J.T.
<2>Draft Genome Sequence of the First Isolate of Extensively Drug-Resistant Mycobacterium tuberculosis in New Zealand.
<3>Genome Announcements
<4>2
<5>e00319-14
<6>2014
<7>Extensively drug-resistant (XDR) tuberculosis has now been described in >90 countries
worldwide. The first case of XDR tuberculosis (XDR-TB) in New Zealand
was recorded in 2010. We report the draft whole-genome sequence of the New
Zealand isolate, NZXDR1, and describe a number of single-nucleotide polymorphisms
that relate to drug resistance.

<>

<1>O-Cuiv, P., Klaassens, E.S., Durkin, A.S., Harkins, D.M., Foster, L., McCorrison, J., Torralba, M., Nelson, K.E., Morrison, M.
<2>Draft genome sequence of Bacteroides vulgatus PC510, a strain isolated from human feces.
<3>J. Bacteriol.
<4>193
<5>4025-4026
<6>2011
<7>Although Bacteroides vulgatus is one of the most prevalent microorganisms in the human
gastrointestinal tract little is known about the genetic
potential of this species. Here we describe the annotated draft genome
sequence of B. vulgatus PC510 isolated from human faeces.

<>

<1>Oakeley, E.J.
<2>DNA methylation analysis: a review of current methodologies.
<3>Pharmacol. Ther.
<4>84
<5>389-400
<6>1999
<7>The relationship between levels of DNA methylation and gene activity has been known for some
time. Many of the early procedures developed gave only
somewhat limited information about methylation patterns, for example, the total level of
5-methyl cytosine in the genome or the frequency of methylation of cytosines within
certain restriction sites. However, in the last few years, there has been an explosion of
interest in DNA methylation, and with it, many new and powerful techniques have
been developed to facilitate its study. In this paper, the key techniques currently available
are reviewed and the advantages, disadvantages, and potential artifacts of each are
discussed.

<>

<1>Oakeley, E.J., Schmitt, F., Jost, J.P.
<2>Quantification of 5-methylcytosine in DNA by the chloroacetaldehyde reaction.
<3>Biotechniques
<4>27
<5>744-746
<6>1999
<7>The study of changes in genome-wide levels of DNA methylation has become a key focus for
understanding the epigenetic regulation of gene expression. Many procedures exist to study DNA
methylation, falling into two categories: gene-specific and genome-wide. Genome-wide
methylation analysis is best performed by DNA hydrolysis followed by HPLC; however, it
requires access to an HPLC machine, which is not always available. Alternative procedures,
such as the radioactive labeling of CpG sites using SssI DNA methyltransferase, have been
developed to address this problem, but it can only monitor CpG methylation changes, and CpNpG
methylation is not detected. Here, we present a method for the analysis of DNA methylation in
any sequence context by fluorescent labeling. We present control analyses using synthetic
oligonucleotides of known methylation levels and a comparison of genomic DNA from two
transgenic tobacco lines known to differ in their methylation levels. The results indicate
that hygromycin-induced hypermethylation acts equally on all classes of methylatable cytosine,
perhaps indicating a common mechanism.

<>

<1>Oakeson, K.F., Gil, R., Clayton, A.L., Dunn, D.M., von Niederhausern, A.C., Hamil, C., Aoyagi, A., Duval, B., Baca, A., Silva, F.J., Vallier, A., Jackson, D.G., Latorre, A., Weiss, R.B., Heddi, A., Moya, A., Dale, C.
<2>Genome Degeneration and Adaptation in a Nascent Stage of Symbiosis.
<3>Genome Biol. Evol.
<4>6
<5>76-93
<6>2014
<7>Symbiotic associations between animals and microbes are ubiquitous in nature, with an
estimated15%of all insect species harboring
intracellular bacterial symbionts. Most bacterial symbionts sharemany genomic features
including small genomes, nucleotide composition
bias, high coding density, and a paucity ofmobileDNA, consistent with long-term host
association. In this study,wefocus on
the early stages of genome degeneration in a recently derived insect-bacterial mutualistic
intracellular association. We present the
completegenomesequenceandannotationof Sitophilusoryzae primary endosymbiont (SOPE).We
alsopresent the finishedgenome
sequence and annotation of strainHS, a close free-living relative of SOPEand other insect
symbionts of the Sodalis-allied clade,whose
gene inventory is expected to closely resemble the putative ancestor of this group.
Structural, functional, and evolutionary analyses
indicate that SOPE has undergone extensive adaptation toward an insect-associated lifestyle in
a very short timeperiod. The genome
of SOPE is large in sizewhen compared withmany ancient bacterial symbionts; however, almost
half of the protein-coding genes in
SOPE are pseudogenes. There is also evidence for relaxed selection on the remaining intact
protein-coding genes. Comparative
analyses of the whole-genome sequence of strain HS and SOPE highlight numerous genomic
rearrangements, duplications, and
deletions facilitated by a recent expansion of insertions sequence elements, some of which
appear to have catalyzed adaptive
changes. Functional metabolic predictions suggest that SOPE has lost the ability to synthesize
several essential amino acids and
vitamins. Analyses of the bacterial cell envelope and genes encoding secretion systems suggest
that these structures and elements
have become simplified in the transition to a mutualistic association.

<>

<1>Oakey, H.J., Cullen, B.R., Owens, L.
<2>The complete nucleotide sequence of the Vibrio harveyi bacteriophage VHML.
<3>J. Appl. Microbiol.
<4>93
<5>1089-1098
<6>2002
<7>Aims: To determine the complete nucleotide sequence of the bacteriophage VHML and establish a
hypothesis for the virulence conversion caused by
VHML infection of Vibrio harveyi. Methods and Results: The complete
nucleotide sequence of VHML was determined (43 193 bp) and used to
identify putative genes. The translated products of these genes were
compared with reported sequences to assign hypothetical functions. All
anticipated structural genes and putative genes for lysogeny were
identified. In addition, we found a complete N6-adenine methyltransferase
(Dam) gene that appeared to have an essential site for ADP-ribosylating
toxins at the C-terminal of the translated product. Conclusions: Virulence
conversion of V. harveyi by VHML may be associated with Dam
transcriptional regulation. The Dam gene may also encode for a toxin
component similar to ADP-ribosylating toxins. Significance and Impact of
Study: This manuscript lays the foundation for understanding the virulence
of toxin-producing V. harveyi. Further research into aspects discussed
here will lead to a greater comprehension regarding the invertebrate
disease vibriosis and its control in the farming of these animals.

<>

<1>Oana, H., Tsumoto, K., Yoshikawa, Y., Yoshikawa, K.
<2>Folding transition of large DNA completely inhibits the action of a restriction endonuclease as revealed by single-chain observation.
<3>FEBS Lett.
<4>530
<5>143-146
<6>2002
<7>The biochemical characteristics of lambda DNA chains in folded/unfolded states upon cleavage
by the restriction enzyme ApaLI were investigated in the presence of spermine. These
characteristics of DNA chains depending on their higher-order structure were studied at the
single-molecule level using fluorescence microscopy. With a low concentration of spermine,
lambda DNA takes a random coiled conformation and allows digestion by the enzyme, while under
a high concentration of spermine, lambda DNA takes a compact folded structure and inhibits
such attack. Together with comparative experiments on short oligomeric DNA, our results
suggest that the transition in the higher-order structure causes on/off-type switching of
sensitivity to the enzyme.

<>

<1>Obarska, A., Blundell, A., Feder, M., Vejsadova, S., Sisakova, E., Weiserova, M., Bujnicki, J.M., Firman, K.
<2>Structural model for the multisubunit Type IC restriction-modification DNA methyltransferase M.EcoR124I in complex with DNA.
<3>Nucleic Acids Res.
<4>34
<5>1992-2005
<6>2006
<7>Recent publication of crystal structures for the putative DNA-binding subunits (HsdS) of the
functionally uncharacterized Type I
restriction-modification (R-M) enzymes MjaXIP and MgeORF438 have provided
a convenient structural template for analysis of the more extensively
characterized members of this interesting family of multisubunit molecular
motors. Here, we present a structural model of the Type IC M.EcoR124I DNA
methyltransferase (MTase), comprising the HsdS subunit, two HsdM subunits,
the cofactor AdoMet and the substrate DNA molecule. The structure was
obtained by docking models of individual subunits generated by
fold-recognition and comparative modelling, followed by optimization of
inter-subunit contacts by energy minimization. The model of M.EcoR124I has
allowed identification of a number of functionally important residues that
appear to be involved in DNA-binding. In addition, we have mapped onto the
model the location of several new mutations of the hsdS gene of M.EcoR124I
that were produced by misincorporation mutagenesis within the central
conserved region of hsdS, we have mapped all previously identified
DNA-binding mutants of TRD2 and produced a detailed analysis of the
location of surface-modifiable lysines. The model structure, together with
location of the mutant residues, provides a better background on which to
study protein-protein and protein-DNA interactions in Type I R-M systems.

<>

<1>Obarska-Kosinska, A., Taylor, J.E., Callow, P., Orlowski, J., Bujnicki, J.M., Kneale, G.G.
<2>HsdR Subunit of the Type I Restriction-Modification Enzyme EcoR124I: Biophysical Characterisation and Structural Modelling.
<3>J. Mol. Biol.
<4>376
<5>438-452
<6>2008
<7>Type I restriction-modification (RM) systems are large, multifunctional enzymes composed of
three different subunits. HsdS and HsdM form a complex
in which HsdS recognizes the target DNA sequence, and HsdM carries out
methylation of adenosine residues. The HsdR subunit, when associated with
the HsdS-HsdM complex, translocates DNA in an ATP-dependent process and
cleaves unmethylated DNA at a distance of several thousand base-pairs from
the recognition site. The molecular mechanism by which these enzymes
translocate the DNA is not fully understood, in part because of the
absence of crystal structures. To date, crystal structures have been
determined for the individual HsdS and HsdM subunits and models have been
built for the HsdM-HsdS complex with the DNA. However, no structure is
available for the HsdR subunit. In this work, the gene coding for the HsdR
subunit of EcoR124I was re-sequenced, which showed that there was an error
in the published sequence. This changed the position of the stop codon and
altered the last 17 amino acid residues of the protein sequence. An
improved purification procedure was developed to enable HsdR to be
purified efficiently for biophysical and structural analysis. Analytical
ultracentrifugation shows that HsdR is monomeric in solution, and the
frictional ratio of 1.21 indicates that the subunit is globular and fairly
compact. Small angle neutron-scattering of the HsdR subunit indicates a
radius of gyration of 3.4 nm and a maximum dimension of 10 nm. We
constructed a model of the HsdR using protein fold-recognition and
homology modelling to model individual domains, and small-angle neutron
scattering data as restraints to combine them into a single molecule. The
model reveals an ellipsoidal shape of the enzymatic core comprising the
N-terminal and central domains, and suggests conformational heterogeneity
of the C-terminal region implicated in binding of HsdR to the HsdS-HsdM
complex.

<>

<1>Obayashi, A.
<2>Restriction enzyme production by culturing Synechoccus strain.
<3>Japanese Patent Office
<4>JP 6066980
<5>
<6>1985
<7>Restriction enzyme is an endonuclease which cuts high molecular weight DNA into specific
sizes.  It is useful for anlaysis of primary structure of DNA, isolation of gene, and genetic
recombination.

<>

<1>Obayashi, A., Hiraoka, N., Kita, K., Nakajima, H.
<2>New restriction enzyme and process for producing the same.
<3>US Patent Office
<4>US 4808531
<5>2168
<6>1989
<7>The invention provides restriction endonuclease MflI capable of recognizing the base sequence
as shown below on a double-stranded DNA molecule and cleaving the DNA chain at the
arrow-marked positions, but has no such action when A is methylated 5'-Pu/GATC Py-3'3'-Py
CTAG/Pu-5' (wherein A represents adenosine, G guanosine, T thymidine, C cytidine, Pu
adenosine or guanosine, and Py thymidine or cytidine). The restriction endonuclease is
produced by culturing Microbacterium flavum IAM 1642, FERM BP-938 in a culture medium and
recovering it from the culture.

<>

<1>Obayashi, A., Hiraoka, N., Kita, K., Nakajima, H.
<2>Production of restriction endonuclease by cultivating Halococcus strains especially from H. acetoinfaciens IAM 12094 for use in genetic manipulations, disease detection etc.
<3>US Patent Office
<4>US 4724209
<5>
<6>1988
<7>Production of a restriction endonuclease capable of recognising the nucleotide sequence ^GATC
on a DNA chain and specifically cleaving the double-stranded chain at the positions marked by
the arrows comprises: (a) growing a suitable Halococcus micro-organism; and (b) collecting the
enzyme from the culture medium. Preferably the endonuclease is obtained by cultivation of H.
acetoinfaciens IAM 12094 in a nutrient medium. The enzyme is formed mainly in the cells and is
recovered by centrifugation etc.

<>

<1>Obayashi, A., Hiraoka, N., Kita, K., Nakajima, H.
<2>Process for producing a restriction enzyme.
<3>US Patent Office
<4>US 4668631
<5>
<6>1987
<7>A restriction endonuclease having the ability to recognize the same base sequence and cleavage
sites as SacII and SstII can be produced from Gluconobacter and isolated in pure form because
no other restriction enzyme is formed.

<>

<1>Obayashi, A., Hiraoka, N., Kita, K., Nakajima, H.
<2>Process for producing a restriction enzyme.
<3>United Kingdom Patent Office
<4>GB 2164946
<5>
<6>1987
<7>A restriction endonuclease having the ability to recognize the same base sequence and cleavage
sites as SacII and SstII can be produced from Gluconobacter and isolated in pure form because
no other restriction enzyme is formed.

<>

<1>Obayashi, A., Hiraoka, N., Kita, K., Nakajima, H.
<2>Production of restriction endonuclease by cultivating Halococcus strains especially from H. acetoinfaciens IAM 12094 for use in genetic manipulations, disease detection etc.
<3>United Kingdom Patent Office
<4>GB 2166744
<5>
<6>1988
<7>Production of a restriction endonuclease capable of recognising the nucleotide sequence ^GATC
on a DNA chain and specifically cleaving the double-stranded chain at the positions marked by
the arrows comprises: (a) growing a suitable Halococcus micro-organism; and (b) collecting the
enzyme from the culture medium. Preferably the endonuclease is obtained by cultivation of H.
acetoinfaciens IAM 12094 in a nutrient medium. The enzyme is formed mainly in the cells and is
recovered by centrifugation etc.

<>

<1>Oberfelder, R., Longo, M., Flynn, E., Chatterjee, D.
<2>Methods for production of proteins.
<3>European Patent Office
<4>EP 1911841 A
<5>
<6>2008
<7>The current invention provides methods for producing a polypeptide as inclusion bodies in
bacterial host cells.  The present methods are carried out by forming a gene construct
comprising the genetic sequence encoding a polypeptide operatively linked to that of an
inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such
that host cells comprising the gene construct produce the polypeptide as inclusion bodies.
The methods of the present invention facilitate the rapid isolation and purification of
recombinant proteins.  In addition, the present methods may be useful for producing
polypeptides or proteins which are small and are typically difficult to express, as well as
those proteins that are toxic to host cells such as E. coli.  The present invention also
provides plasmids, vectors and host cells to be used in the present invention for production
of polypeptides and methods of production of polypeptides using these vectors and host cells.
The invention further provides methods for producing protein molecular weight ladders for use
in protein gel electrophoresis, as well as proteins and protein molecular weight ladders
produced by these methods.

<>

<1>Oberfelder, R., Longo, M., Flynn, E., Chatterjee, D.
<2>Methods for production of proteins.
<3>European Patent Office
<4>EP 2196536 A
<5>
<6>2010
<7>The current invention provides methods for producing a polypeptide as inclusion bodies in
bacterial host cells.  The present methods are carried out by forming a gene construct
comprising the genetic sequence encoding a polypeptide operatively linked to that of an
inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such
that host cells comprising the gene construct produce the polypeptide as inclusion bodies.
The methods of the present invention facilitate the rapid isolation and purification of
recombinant protein.  In addition, the present methods may be useful for producing
polypeptides or protein which are small and are typically difficult to express, as well as
those proteins that are toxic to host cells such as E. coli.  The present invention also
provides plasmids, vectors and host cells to be used in the present invention for production
of polypeptides and methods of production of polypeptides using these vectors as host cells.
The invention further provides methods for producing protein molecular weight ladders produced
by these methods.

<>

<1>Oberfelder, R., Longo, M., Flynn, E., Chatterjee, D.
<2>Methods for production of proteins.
<3>European Patent Office
<4>EP 2336313 A
<5>
<6>2011
<7>The current invention provides methods for producing a polypeptide as inclusion bodies in
bacterial host cells.  The present methods are carried out by forming a gene construct
comprising the genetic sequence encoding a polypeptide operatively linked to that of an
inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such
that host cells comprising the gene construct produce the polypeptide as inclusion bodies.
The methods of the present invention facilitate the rapid isolation and purification of
recombinant proteins.  In addition, the present methods may be useful for producing
polypeptides or proteins which are small and are typically difficult to express, as well as
those proteins that are toxic to host cells such as E. coli.  The present invention also
provides plasmids, vectors and host cells to be used in the present invention for production
of polypeptides using these vectors and host cells.  The invention further provides methods
for producing protein molecular weight ladders for use in protein gel electrophoresis, as well
as proteins and protein molecular weight ladders produced by these methods.

<>

<1>Oberto, J., Gaudin, M., Cossu, M., Gorlas, A., Slesarev, A., Marguet, E., Forterre, P.
<2>Genome Sequence of a Hyperthermophilic Archaeon, Thermococcus nautili 30-1, That  Produces Viral Vesicles.
<3>Genome Announcements
<4>2
<5>e00243-14
<6>2014
<7>Thermococcus nautili 30-1 (formerly Thermococcus nautilus), an anaerobic hyperthermophilic
marine archaeon, was isolated in 1999 from a deep-sea
hydrothermal vent during the Amistad campaign. Here, we present the complete
sequence of T. nautili, which is able to produce membrane vesicles containing
plasmid DNA. This property makes T. nautili a model organism to study horizontal
gene transfer.

<>

<1>Obregon, V., Garcia, P., Lopez, R., Garcia, J.L.
<2>VO1, a temperate bacteriophage of the type 19A multiresistant epidemic 8249 strain of Streptococcus pneumoniae: Analysis of variability of  lytic and putative C5 methyltransferase genes.
<3>Microb. Drug Resist.
<4>9
<5>7-15
<6>2003
<7>A temperate bacteriophage (VO1) has been isolated from the Streptococcus pneumoniae type 19F
multiresistant epidemic 8249 strain (South African
strain). Structural analysis of the specific integration site, protein
composition, restriction patterns, and molecular dissection of the lytic
system of this phage revealed high sequence similarity with MM1, a
temperate phage from the Spain23F-1 strain of pneumococcus, another
multiresistant epidemic clone. The different pneumococcal strains
sequenced so far exhibit an identical and single attB located in the same
site of the genome. Remarkably, the LytA amidase coded by VO1 showed clear
differences with that of the host bacterium in contrast with the situation
previously documented for bacterial- and phage-coded amidases of
pneumococcus. In addition, a new gene (orfmet) putatively coding for a C5
methyltransferase has been identified. A noticeable variability affecting
the presence (or absence) of this supernumerary gene(s) in the same region
of the genomes of three otherwise highly similar phages (i.e., VO1, MM1,
and HB-3) suggests frequent recombinational events leading to introduce
variability in this genome region. The peculiarities of genes like lytA
and orfmet in VO1 provide interesting insights on mechanisms of horizontal
transfer and lysogenic state co-evolution.

<>

<1>Ochiai, H., Inoue, Y., Takeya, M., Sasaki, A., Kaku, H.
<2>Genome sequence of Xanthomonas oryzae pv. oryzae suggests contribution of large numbers of effector genes and insertion sequences to its race diversity.
<3>Japan Agricultural Research Quarterly
<4>39
<5>275-287
<6>2005
<7>The plant-pathogenic prokaryote Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial blight,
one of the most important diseases of rice.  The bacterium is a model organism for the
analysis of plant-pathogen interaction, because more than 30 races differing in virulence and
25 resistance genes in rice have been reported to date.  We present here the complete genome
sequence of Xoo strain MAFF 311018.  The size of the genome was 4,940,217 bp, in a single
circular chromosome.  The genome structure of Xoo MAFF 311018 was characterized by large
numbers of effector (avr) genes of the avrBs3/pth family and insertion sequences (Iss).  RFLP
analysis of diverse strains using ISxo1 as a probe suggests that the prevalence of mobile
elements in this species, which can bring about genome inversions and rearrangement, may have
played a major role in generating the high degree of genetic diversity and race
differentiation characteristic of this pathogen.  The Xoo MAFF 311018 sequence was also highly
similar to those of X. axonopodis pv. citri and X. campestris pv. campestris with the
exception of the large number of effectors and IS elements, and numerous inversions and
rearrangements.

<>

<1>Ochiai, H., Shibata, H., Sawa, Y., Ashida, N.
<2>Restriction endonucleases from Phormidium lapideum, a strain of filamentous and thermophilic Cyanobacteria.
<3>Bull. Fac. Agr. Shimane Univ.
<4>23
<5>184-191
<6>1989
<7>A couple of restriction endonucleases, PlaI and PlaII, have been purified from
a filamentous and thermophilic cyanobacterium, Phormidium lapideum.  PlaI
proved to be an isoschizomer of HaeIII and cleaved the site GG^CC and was a
monomeric protein that had a molecular weight of about 40 kilodaltons.  PlaII
was an isoschizomer of Nsp(7524)V and recognized the site TTCGAA and was
estimated as a heterotetrameric protein (alpha2,beta2).  PlaII has an apparent
molecular mass of 176 kilodaltons and that of the alpha subunit was 63
kilodaltons, beta subunit 31 kilodaltons.  Characteristics of PlaI and PlaII
were investigated in comparison with that of the respective isoschizomers,
HaeIII and Nsp(7524)V.

<>

<1>Oda, K., Marmur, J.
<2>Purification and properties of deoxyribonucleic acid methylase from Bacillus subtilis.
<3>Biochemistry
<4>5
<5>761-773
<6>1966
<7>DNA methylase was purified about 100-fold from Bacillus subtilis strain 6633.
The novel features of this enzyme are as follows: (1) During growth of
bacteria, enzyme activity was detected principally in the early exponential
phase.  (2) The enzyme catalyzes the methylation of both native and
heat-denatured DNA.  (3) The product of the enzymatic methylation of DNA was
5-methylcytosine (5MC) only.  (4) The extent of the methylation of both native
and heat-denatured bacterial DNA is approximately proportional to its GC
(guanine plus cytosine) content.  The higher the per cent GC in DNA, the
greater the extent of methylation that takes place, even if the ratio of 5MC to
total cytosine residues in DNA is considered.  The possibility of the selective
methylation of one strand of DNA was also examined by using B. subtilis
bacteriophage 2C DNA, whose strands, after denaturation, can be fractionated
because of a bias in base composition.  The H and L strands of phage 2C DNA
were separated by MAK column chromatography or by centrifugation in alkaline
CsCl density gradients, and it was found that both strands were equally
methylated in vivo and in vitro.

<>

<1>Oda, M., Tanaka, S., Shiota, K.
<2>DNA methyltransferase 1 is essential for establishment of trophoblast stem cells in culture.
<3>Biol. Reprod.
<4>64
<5>175-176
<6>2001
<7>DNA methylation patterns in mammals are stage- and tissue-specific.  This suggests that
methylation patterns contribute to proper cell differentiation and/or to the maintenance of
cellular characteristics.  Several different DNA methyltransferases exist that both establish
and maintain proper methylation patterns.  Dnmt1 is largely a maintenance methyltransferase
that ensures an established methylation pattern is inherited by daughter cells.  Disruption of
Dnmt1 causes growth delay at gastrulation and homozygous mutants are embryonic lethal.  To
evaluate the role of Dnmt1 in placental development, we attempted to establish trophoblast
stem cells from homozygous Dnmt1 hypomorphic mutant embryos.  TS cells exclusively contribute
to the trophoblast lineage in vivo in chimeras.  At the blastocyst stage, TS cells appear in
the polar trophectoderm, which is adjacent to the inner cell mass.  TS cells can be maintained
in culture with media supplemented with FGF-4, and differentiate into several subtypes of
trophoblast cells.  To establish homozygous mutant TS cells, we used blastocysts from
heterozygous matings.  The 39 blastocysts from 6 female mice were incubated in FGF4-contained
medium with feeder cells.  Of 39 blastocysts, 28 generated stem cell colonies; 3 were
homozygous, 10 were wild type, 13 were heterozygous clones, and 2 could not be genotyped, but
were probably also homozygous.  All clones were passed two more times and most (70-80%) of
wild type and heterozygous clones survived.  However, no homozygous clone survived beyond the
second passage.  Some autonomous differentiation was observed in the homozygous TS cells.  Our
results indicate that Dnmt1-hypomorphic TS cells cannot be established in cell culture, as
normal and heterozygous TS cells can.  This is in contrast to Dnmt1-hypomorphic embryonic stem
cells, which can be maintained in culture.  These data suggest that the trophoblast lineage is
more dependent on Dnmt1 than the embryo proper.

<>

<1>Odom, O.W., Holloway, S.P., Deshpande, N.N., Lee, J., Herrin, D.L.
<2>Mobile self-splicing Group I introns from the psbA gene of Chlamydomonas reinhardtii: highly efficient homing of an exogenous intron containing its own promoter.
<3>Mol. Cell. Biol.
<4>21
<5>3472-3481
<6>2001
<7>Introns 2 and 4 of the psbA gene of Chlamydomonas reinhardtii chloroplasts (Cr.psbA2 and
Cr.psbA4, respectively) contain large free-standing open reading frames (ORFs). We used
transformation of an intronless-psbA strain (IL) to test whether these introns undergo homing.
Each intron, plus short exon sequences, was cloned into a chloroplast expression vector in
both orientations and then cotransformed into IL along with a spectinomycin resistance marker
(16S rrn). For Cr.psbA2, the sense construct gave nearly 100% cointegration of the intron
whereas the antisense construct gave 0%, consistent with homing. For Cr.psbA4, however, both
orientations produced highly efficient cointegration of the intron. Efficient cointegration of
Cr.psbA4 also occurred when the intron was introduced as a restriction fragment lacking any
known promoter. Deletion of most of the ORF, however, abolished cointegration of the intron,
consistent with homing. The Cr.psbA4 constructs also contained a
3-(3,4-dichlorophenyl)-1,1-dimethylurea resistance marker in exon 5, which was always present
when the intron integrated, thus demonstrating exon coconversion. Remarkably, primary
selection for this marker gave >100-fold more transformants (>10,000/?g of DNA) than did the
spectinomycin resistance marker. A trans homing assay was developed for Cr.psbA4; the
ORF-minus intron integrated when the ORF was cotransformed on a separate plasmid. This assay
was used to identify an intronic region between bp -88 and -194 (relative to the ORF) that
stimulated homing and contained a possible bacterial (-10, -35)-type promoter. Primer
extension analysis detected a transcript that could originate from this promoter. Thus, this
mobile, self-splicing intron also contains its own promoter for ORF expression. The
implications of these results for horizontal intron transfer and organelle transformation are
discussed.

<>

<1>Oehlerking, J., Kube, M., Felder, K.M., Matter, D., Wittenbrink, M.M., Schwarzenbach, S., Kramer, M.M., Hoelzle, K., Hoelzle, L.E.
<2>The complete genome sequence of the hemotrophic Mycoplasma suis_KI3806.
<3>J. Bacteriol.
<4>193
<5>2369-2370
<6>2011
<7>Mycoplasma suis, a member of the hemotrophic mycoplasma (HM) group, parasitize erythrocytes of
pigs. Increasing evidences suggest that M. suis is also a zoonotic agent. Highly pathogenic
strains of M. suis (e.g. M. suis_KI3806) have been demonstrated to invade erythrocytes. This
complete sequenced and manually annotated genome of M. suis_KI3806 is the first available from
this species and from all HM. The DNA was isolated from blood-samples of experimentally
infected pigs due to the lack of an in vitro cultivation system. The small circular chromosome
of 709270 bp with an unexpected-high number of hypothetical proteins and limited transport and
metabolic capacities could reflect the unique life-style of HM on the surface of erythrocytes.

<>

<1>Oelgeschlager, T., Geiger, R., Ruter, T., Alves, J., Fliess, A., Pingoud, A.
<2>Probing the function of individual amino acid residues in the DNA binding site of the EcoRI restriction endonuclease by analysing the toxicity of genetically engineered mutants.
<3>Gene
<4>89
<5>19-27
<6>1990
<7>We have developed an assay that allows analysis of the activity of EcoRI
restriction endonuclease (ENase) and its mutants in vivo.  This assay is based
on the fact that wild type (wt) EcoRI ENase is toxic for Escherichia coli cells
not expressing the EcoRI methyltransferase (MTase).  The viability factor
defined by the ratio of the viable counts of E. coli cultures having or not
having expressed the ecoRIR gene for a defined time is 10-6 for wt EcoRI ENase
and close to one for a totally inactive EcoRI ENase mutant.  While the EcoRI
MTase (M.EcoRI) provides substantial protection against the toxic effects of
the wt EcoRI ENase and several of the mutants, some mutants become more toxic
in the presence of M.EcoRI.  Twenty-four different DNA-binding-site mutants of
EcoRI ENase were characterized in their activity in vivo with this assay.  The
results obtained allow us to conclude that the structural integrity of the
region at and around aa 200 seems to be very critical for the enzymatic
function of EcoRI ENase: nonconservative replacements there lead to viability
factors of 1-10/2.  While our results indicate that the region around aa 144
and 145 is also involved in the EcoRI ENase-catalyzed reaction, it is also
evident that the effects of mutation there are not as large:  viability factors
of approx. 10-3 are obtained even for drastic replacements.  These results are
discussed in the light of the x-ray structure analysis of an EcoRI ENase-DNA
recognition complex.

<>

<1>Ogata, H., Audic, S., Renesto-Audiffren, P., Fournier, P.-E., Barbe, V., Samson, D., Roux, V., Cossart, P., Weissenbach, J., Claverie, J.-M., Raoult, D.
<2>Mechanisms of evolution in Rickettsia conorii and R. prowazekii.
<3>Science
<4>293
<5>2093-2098
<6>2001
<7>Rickettsia conorii is an obligate intracellular bacterium that causes Mediterranean spotted
fever in humans. We determined the 1,268,755-nucleotide complete genome sequence of R.
conorii, containing 1374 open reading frames. This genome exhibits 804 of the 834 genes of the
previously determined R. prowazekii genome plus 552 supplementary open reading frames and a
10-fold increase in the number of repetitive elements. Despite these differences, the two
genomes exhibit a nearly perfect colinearity that allowed the clear identification of
different stages of gene alterations with gene remnants and 37 genes split in 105 fragments,
of which 59 are transcribed. A 38-kilobase sequence inversion was dated shortly after the
divergence of the genus.

<>

<1>Ogata, H., La Scola, B., Audic, S., Renesto, P., Blanc, G., Robert, C., Fournier, P.E., Claverie, J.M., Raoult, D.
<2>Genome sequence of Rickettsia bellii illuminates the role of amoebae in gene exchanges between intracellular pathogens.
<3>PLoS Genet.
<4>2
<5>e76
<6>2006
<7>The recently sequenced Rickettsia felis genome revealed an unexpected plasmid carrying several
genes usually associated with DNA transfer,
suggesting that ancestral rickettsiae might have been endowed with a
conjugation apparatus. Here we present the genome sequence of Rickettsia
bellii, the earliest diverging species of known rickettsiae. The 1,552,076
base pair-long chromosome does not exhibit the colinearity observed
between other rickettsia genomes, and encodes a complete set of putative
conjugal DNA transfer genes most similar to homologues found in
Protochlamydia amoebophila UWE25, an obligate symbiont of amoebae. The
genome exhibits many other genes highly similar to homologues in
intracellular bacteria of amoebae. We sought and observed sex pili-like
cell surface appendages for R. bellii. We also found that R. bellii very
efficiently multiplies in the nucleus of eukaryotic cells and survives in
the phagocytic amoeba, Acanthamoeba polyphaga. These results suggest that
amoeba-like ancestral protozoa could have served as a genetic "melting
pot" where the ancestors of rickettsiae and other bacteria promiscuously
exchanged genes, eventually leading to their adaptation to the
intracellular lifestyle within eukaryotic cells.

<>

<1>Ogata, H., Renesto, P., Audic, S., Robert, C., Blanc, G., Fournier, P.E., Parinello, H., Claverie, J.M., Raoult, D.
<2>The genome sequence of Rickettsia felis identifies the first putative conjugative plasmid in an obligate intracellular parasite.
<3>PLoS Biology
<4>3
<5>e248
<6>2005
<7>We sequenced the genome of Rickettsia felis, a flea-associated obligate intracellular
alpha-proteobacterium causing spotted fever in humans.
Besides a circular chromosome of 1,485,148 bp, R. felis exhibits the first
putative conjugative plasmid identified among obligate intracellular
bacteria. This plasmid is found in a short (39,263 bp) and a long (62,829
bp) form. R. felis contrasts with previously sequenced Rickettsia in terms
of many other features, including a number of transposases, several
chromosomal toxin-antitoxin genes, many more spoT genes, and a very large
number of ankyrin- and tetratricopeptide-motif-containing genes.
Host-invasion-related genes for patatin and RickA were found. Several
phenotypes predicted from genome analysis were experimentally tested:
conjugative pili and mating were observed, as well as beta-lactamase
activity, actin-polymerization-driven mobility, and hemolytic properties.
Our study demonstrates that complete genome sequencing is the fastest
approach to reveal phenotypic characters of recently cultured obligate
intracellular bacteria.

<>

<1>Ogawa, S., Matsuo, K., Angata, K., Yanagisawa, K., Tanaka, Y.
<2>Group-I introns in the cytochrome c oxidase genes of Dictyostelium discoideum: two related ORFs in one loop of a group-I intron, a cox1/2 hybrid gene and an unusually large cox3 gene.
<3>Curr. Genet.
<4>31
<5>80-88
<6>1997
<7>The DNA sequences of cytochrome oxidase (subunits 1, 2 and 3) genes of the cellular slime mold
Dictyostelium discoideum mitochondria were determined.  The genes for subunits 1 and 2 have a
single continuous ORF (COX1/2) which contains four group-I introns.  The insertion sites of
the two group-I introns (DdOX1/2.2 and DdOX1/2.3) coincide with those of fungal and algal
group-I introns, as well as a liverwort group-I intron, in the cytochrome oxidase subunit 1.
Interestingly, intron DdOX1/2.2 has two free-standing ORFs in a loop (L8) which have similar
amino-acid sequences and are homologous to ai4 DNA endonuclease (I-Sce II) and bi4 RNA
maturase found in group-I introns of Saccharomyces cerevisiae mitochondrial DNA.  Two group-I
introns (DdOX1/2.3 and DdOX1/2.4) also have a free-standing ORF in loop 1 and loop 2,
respectively.  These results show that these group-I introns and the intronic ORFs have
evolved from the same ancestral origin, but that these ORFs have been propagated
independently.

<>

<1>Ogawa, S., Naito, K., Angata, K., Morio, T., Urushihara, H., Tanaka, Y.
<2>A site-specific DNA endonuclease specified by one of two ORFs encoded by a group I intron in Dictyostelium discoideum mitochondrial DNA.
<3>Gene
<4>191
<5>115-121
<6>1997
<7>The second intron (DdOX1/2.2) of Dictyostelium discoideum cytochrome oxidase subunit 1/2 fused
gene has two free-standing ORF genes (Dd ai2a and Dd ai2b) in a loop, which have similar amino
acid sequences and are homologous to aI4 DNA endonuclease (I-SceII) of Saccharomyces
cerevisiae.  To elucidate the functions of these ORFs, we cloned the ORFs into an expression
vector and introduced the composite vectors into E. coli.  The expression of Dd ai2a in E.
coli caused growth inhibition and degradation of the E. coli genomic DNA.  To determine
whether Dd ai2a protein is a homing type DNA endonuclease, the ability to cleave the homing
site of its intron in vivo was examined.  Dd ai2a cleaved only one strand of intronless DNA
sequence at the site which coincides with the I-SceII cleavage recognition site.  We suppose
that Dd ai2a functions actually as a homing type DNA endonuclease in D. discoideum
mitochondria by virtue of other factors.  To obtain further information about the relationship
between the existence of introns and the mating system, we carried out in vitro self-splicing
assay and polymerase chain reaction analysis using 13 strains of the cellular slime mold.

<>

<1>Ogawa, Y., Ooka, T., Shi, F., Ogura, Y., Nakayama, K., Hayashi, T., Shimoji, Y.
<2>The Genome of Erysipelothrix rhusiopathiae, the Causative Agent of Swine Erysipelas, Reveals New Insights into the Evolution of Firmicutes and the Organism's Intracellular Adaptations.
<3>J. Bacteriol.
<4>193
<5>2959-2971
<6>2011
<7>Erysipelothrix rhusiopathiae is a Gram-positive bacterium that represents
a new class, Erysipelotrichia, in the phylum Firmicutes. The organism is a
facultative intracellular pathogen that causes swine erysipelas, as well
as a variety of diseases in many animals. Here, we report the first
complete genome sequence analysis of a member of the class
Erysipelotrichia. The E. rhusiopathiae genome (1,787,941 bp) is one of the
smallest genomes in the phylum Firmicutes. Phylogenetic analyses based on
the 16S rRNA gene and 31 universal protein families suggest that E.
rhusiopathiae is phylogenetically close to Mollicutes, which comprises
Mycoplasma species. Genome analyses show that the overall features of the
E. rhusiopathiae genome are similar to those of other Gram-positive
bacteria; it possesses a complete set of peptidoglycan biosynthesis genes,
two-component regulatory systems, and various cell wall-associated
virulence factors, including a capsule and adhesins. However, it lacks
many orthologous genes for the biosynthesis of wall teichoic acids (WTA)
and lipoteichoic acids (LTA) and the dltABCD operon, which is responsible
for d-alanine incorporation into WTA and LTA, suggesting that the organism
has an atypical cell wall. In addition, like Mollicutes, its genome shows
a complete loss of fatty acid biosynthesis pathways and lacks the genes
for the biosynthesis of many amino acids, cofactors, and vitamins,
indicating reductive genome evolution. The genome encodes nine antioxidant
factors and nine phospholipases, which facilitate intracellular survival
in phagocytes. Thus, the E. rhusiopathiae genome represents evolutionary
traits of both Firmicutes and Mollicutes and provides new insights into
its evolutionary adaptations for intracellular survival.

<>

<1>Oger, P., Sokolova, T.G., Kozhevnikova, D.A., Chernyh, N.A., Bartlett, D.H., Bonch-Osmolovskaya, E.A., Lebedinsky, A.V.
<2>Complete Genome Sequence of the Hyperthermophilic Archaeon Thermococcus sp. Strain AM4, Capable of Organotrophic Growth and Growth at the Expense  of Hydrogenogenic or Sulfidogenic Oxidation of Carbon Monoxide.
<3>J. Bacteriol.
<4>193
<5>7019-7020
<6>2011
<7>Analysis of the complete genome of Thermococcus sp. strain AM4, which was the first
lithotrophic Thermococcales isolate described and the first
archaeal isolate to exhibit a capacity for hydrogenogenic carboxydotrophy,
reveals a proximity with Thermococcus gammatolerans, corresponding to
close but distinct species that differ significantly in their lithotrophic
capacities.

<>

<1>Ogg, C.D., Patel, B.K.
<2>Draft Genome Sequence of Caloramator australicus strain RC3T a thermoanaerobe from the Great Artesian Basin of Australia.
<3>J. Bacteriol.
<4>193
<5>2664-2665
<6>2011
<7>Caloramator australicus strain RC3(T) (JCM 15081(T) = KCTC 5601(T)) is the type strain of a
newly identified thermophilic species, which was isolated from red-coloured microbial mats
that thrive at 66  degrees C in the runoff channel of a Great Artesian Basin bore (New Lorne
bore, registered number 17263) in outback Queensland, Australia. The ability of C. australicus
strain to use metals as terminal electron acceptors has led to concerns that it could colonise
and enhance corrosion of the metal casing of Great Artesian Basin bore-well pipes, and this
could subsequently lead to bore failure and loss of water availability for the community which
is so reliant on it. The genome of C. australicus strain has been sequenced, and annotation of
the  approximately  2.65 Mb sequence indicates that the attributes are consistent with
physiological and phenotypic traits.

<>

<1>Ogino, H., Azuma, Y., Hosoyama, A., Nakazawa, H., Matsutani, M., Hasegawa, A., Otsuyama, K., Matsushita, K., Fujita, N., Shirai, M.
<2>Complete Genome Sequence of NBRC 3288, a Unique Cellulose-Nonproducing Strain of Gluconacetobacter xylinus Isolated from Vinegar.
<3>J. Bacteriol.
<4>193
<5>6997-6998
<6>2011
<7>Gluconacetobacter xylinus is involved in the industrial production of cellulose. We have
determined the genome sequence of G. xylinus NBRC 3288,
a cellulose-nonproducing strain. Comparative analysis of genomes of G.
xylinus NBRC 3288 with those of the cellulose-producing strains clarified
the genes important for cellulose production in Gluconacetobacter.

<>

<1>Ogunremi, D., Devenish, J., Amoako, K., Kelly, H., Dupras, A.A., Belanger, S., Wang, L.R.
<2>High resolution assembly and characterization of genomes of Canadian isolates of Salmonella Enteritidis.
<3>BMC Genomics
<4>15
<5>713
<6>2014
<7>BACKGROUND: There is a need to characterize genomes of the foodborne pathogen,
Salmonella enterica serovar Enteritidis (SE) and identify genetic information
that could be ultimately deployed for differentiating strains of the organism, a
need that is yet to be addressed mainly because of the high degree of clonality
of the organism. In an effort to achieve the first characterization of the
genomes of SE of Canadian origin, we carried out massively parallel sequencing of
the nucleotide sequence of 11 SE isolates obtained from poultry production
environments (n = 9), a clam and a chicken, assembled finished genomes and
investigated diversity of the SE genome. RESULTS: The median genome size was
4,678,683 bp. A total of 4,833 chromosomal genes defined the pan genome of our
field SE isolates consisting of 4,600 genes present in all the genomes, i.e.,
core genome, and 233 genes absent in at least one genome (accessory genome).
Genome diversity was demonstrable by the presence of 1,360 loci showing single
nucleotide polymorphism (SNP) in the core genome which was used to portray the
genetic distances by means of a phylogenetic tree for the SE isolates. The
accessory genome consisted mostly of previously identified SE prophage sequences
as well as two, apparently full- sized, novel prophages namely a 28 kb sequence
provisionally designated as SE-OLF-10058 (3) prophage and a 43 kb sequence
provisionally designated as SE-OLF-10012 prophage. CONCLUSIONS: The number of
SNPs identified in the relatively large core genome of SE is a reflection of
substantial diversity that could be exploited for strain differentiation as shown
by the development of an informative phylogenetic tree. Prophage sequences can
also be exploited for SE strain differentiation and lineage tracking. This work
has laid the ground work for further studies to develop a readily adoptable
laboratory test for the subtyping of SE.

<>

<1>Oguro, K., Tamura, K., Yamane, J., Shimizu, M., Yamamoto, T., Ikawa, T., Ohnishi, K., Oshima, S., Imajoh, M.
<2>Draft Genome Sequences of Two Genetic Variant Strains of Edwardsiella piscicida,  JF1305 and RSB1309, Isolated from Olive Flounder (Paralichythys olivaceus) and  Red Sea Bream (Pagrus major) Cultured in Japan, Respectively.
<3>Genome Announcements
<4>2
<5>e00546-14
<6>2014
<7>Edwardsiella piscicida is a new species discovered within the group of organisms
traditionally classified as Edwardsiella tarda. We present draft genome sequences
of two variant strains of E. piscicida, JF1305 and RSB1309. Differences in
protein-coding sequence between these isolates are associated with virulence,
disease, and defense, suggesting differences in pathogenicity.

<>

<1>Oguro, K., Yamane, J., Yamamoto, T., Ohnishi, K., Oshima, S., Imajoh, M.
<2>Draft Genome Sequence of Streptococcus parauberis Strain SK-417, Isolated from Diseased Sebastes ventricosus in Kagoshima, Japan.
<3>Genome Announcements
<4>2
<5>e00453-14
<6>2014
<7>Streptococcus parauberis strain SK-417 was isolated from the brain of a diseased  Sebastes
ventricosus, collected from an aquaculture farm in April 2013 in
Kagoshima Prefecture, Japan. The draft genome sequence, obtained with a 454 GS
Junior sequencing system, consists of 33 large contigs of >500 bp, totaling
1,958,836 bp, and has a G+C content of 35.4%.

<>

<1>Oh, C., Heo, S.J., De Zoysa, M., Affan, A., Jung, W.K., Park, H.S., Lee, Y., Lee, J., Yoon, K.T., Kang, D.H.
<2>Whole-Genome Sequence of the Xylanase-Producing Mesoflavibacter zeaxanthinifaciens Strain S86.
<3>J. Bacteriol.
<4>193
<5>5557
<6>2011
<7>We isolated Mesoflavibacter zeaxanthinifaciens S86 as xylanase-producing bacteria from
seawater sampled in Micronesia. Analysis of the M.
zeaxanthinifaciens genome revealed that it contains a single circular
chromosome of 3,704,661 bp with 3,249 putative open reading frames.

<>

<1>Oh, C., Kwon, Y.K., Heo, S.J., De Zoysa, M., Affan, A., Lee, Y., Lee, J., Choi, Y.U., Park, H.S., Jung, K.H., Lee, H.Y., Kang, D.H.
<2>Complete genome sequence of strain s85, a novel member of the family flavobacteriaceae.
<3>J. Bacteriol.
<4>193
<5>6107
<6>2011
<7>An agar-degrading marine bacterium identified as a novel member of the family
Flavobacteriaceae (strain S85) was isolated from seawater in
Micronesia. The sequenced strain S85 genome is composed of 3,384,629 bp in
a circular chromosome, which includes 2,883 complete open reading frames.

<>

<1>Oh, C., Zoysa, M.D., Kwon, Y.K., Heo, S.J., Affan, A., Jung, W.K., Park, H.S., Lee, J., Son, S.K., Yoon, K.T., Kang, D.H.
<2>Complete genome sequence of the agarase-producing marine bacterium strain s89, representing a novel species of the genus alteromonas.
<3>J. Bacteriol.
<4>193
<5>5538
<6>2011
<7>We report here the annotated genome sequence of the marine bacterium Alteromonas sp. S89 and
the identification of six genes coding for
agar-degrading enzymes. The sequenced Alteromonas sp. S89 genome is
composed of a 3,864,871-bp circular chromosome that includes 3,236
complete open reading frames.

<>

<1>Oh, H.M., Giovannoni, S.J., Ferriera, S., Johnson, J., Cho, J.C.
<2>The Complete Genome Sequence of Erythrobacter litoralis HTCC2594.
<3>J. Bacteriol.
<4>191
<5>2419-2420
<6>2009
<7>Erythrobacter litoralis has been known as a bacteriochlorophyll a-containing aerobic
anoxygenic phototrophic bacterium. Here we announce
the complete genome sequence of E. litoralis HTCC2594 that is devoid of
phototrophic potential. E. litoralis HTCC2594, isolated by
dilution-to-extinction culturing from seawater, could not carry out
aerobic anoxygenic phototrophy and lacked genes for bacteriochlorophyll a
biosynthesis and photosynthetic reaction center proteins.

<>

<1>Oh, H.M., Giovannoni, S.J., Lee, K., Ferriera, S., Johnson, J., Cho, J.C.
<2>Complete Genome Sequence of Robiginitalea biformata HTCC2501.
<3>J. Bacteriol.
<4>191
<5>7144-7145
<6>2009
<7>Robiginitalea biformata HTCC2501, isolated by dilution-to-extinction culturing from the
Sargasso Sea, has been known as an aerobic chemoheterotroph with carotenoid pigments and
dimorphic growth phases. Here we announce the complete genome sequence of R. biformata
HTCC2501 that contained genes for carotenoid biosynthesis and several macromolecule-degrading
enzymes.

<>

<1>Oh, H.M., Kang, I., Ferriera, S., Giovannoni, S.J., Cho, J.C.
<2>Genome Sequence of an Oligotrophic Marine Gammaproteobacterium HTCC2143 Isolated from Oregon Coast.
<3>J. Bacteriol.
<4>192
<5>4530-4531
<6>2010
<7>Strain HTCC2143 was isolated from Oregon Coast surface waters using dilution-to-extinction
culturing. Here we present the genome of strain
HTCC2143 from BD1-7 clade of the oligotrophic marine Gammaproteobacteria
group (OMG). The genome of HTCC2143 encodes genes for carotenoid
biosynthesis, proteorhodopsin, and genes that have potential
biotechnological significance: epoxide hydrolases, Baeyer-Villiger
monooxygenases, and polyketide synthases.

<>

<1>Oh, H.M., Kang, I., Lee, K., Jang, Y., Lim, S.I., Cho, J.C.
<2>Complete Genome Sequence of Strain IMCC9063 Belonging to SAR11 subgroup 3, Isolated from the Arctic Ocean.
<3>J. Bacteriol.
<4>193
<5>3379-3380
<6>2011
<7>Strain IMCC9063 is a novel isolate of the SAR11 clade and is distantly related to other
cultured representatives in this clade. The strain was
isolated off the coast of Svalbard, Norway by applying high-throughput
culturing methods based on dilution-to-extinction. Here we present the
finished genome sequence of strain IMCC9063.

<>

<1>Oh, H.M., Kang, I., Vergin, K.L., Kang, D., Rhee, K.H., Giovannoni, S.J., Cho, J.C.
<2>Complete Genome Sequence of Parvularcula bermudensis HTCC2503T, the type species of the order 'Parvularculales' in the Alphaproteobacteria.
<3>J. Bacteriol.
<4>193
<5>305-306
<6>2010
<7>The order 'Parvularculales' represents the seventh order in the class Alphaproteobacteria.
Parvularcula bermudensis, the type species of the order, was isolated from the Sargasso Sea
using dilution-to-extinction culturing. We present here the complete genome sequence of
Parvularcula bermudensis HTCC2503(T) that contains genes for carotenoid biosynthesis,
dimethylsulfoniopropionate demethylase, and transduction-like gene transfer agents.

<>

<1>Oh, H.M., Kang, I., Vergin, K.L., Lee, K., Giovannoni, S.J., Cho, J.C.
<2>Genome Sequence of Oceanicaulis sp. HTCC2633, Isolated from the Western Sargasso Sea.
<3>J. Bacteriol.
<4>193
<5>317-318
<6>2010
<7>The genus Oceanicaulis represents dimorphic rods that were originally isolated from a marine
dinoflagellate. Here we announce the genome sequence of Oceanicaulis sp. strain HTCC2633,
isolated by dilution-to-extinction culturing from the Sargasso Sea. The genome information of
strain HTCC2633 indicates a chemoorganotrophic way of life of this strain.

<>

<1>Oh, H.M., Kang, I., Yang, S.J., Jang, Y., Vergin, K.L., Giovannoni, S.J., Cho, J.C.
<2>Complete Genome Sequence of strain HTCC2170, a Novel Member of the Genus Maribacter in the Family Flavobacteriaceae.
<3>J. Bacteriol.
<4>193
<5>303-304
<6>2010
<7>Strain HTCC2170 was isolated from surface waters off the Oregon coast using
dilution-to-extinction culturing. Here we present the finished genome sequence of a marine
bacterium, Maribacter sp. HTCC2170. Maribacter sp. HTCC2170 is predicted to be a facultatively
aerobic chemoorganotroph that is capable of macromolecule degradation and anaerobic
respiration based on genomic sequence analysis.

<>

<1>Oh, H.M., Kwon, K.K., Kang, I., Kang, S.G., Lee, J.H., Kim, S.J., Cho, J.C.
<2>Complete genome sequence of 'Candidatus Puniceispirillum marinum' IMCC1322, a representative of the SAR116 clade in the Alphaproteobacteria.
<3>J. Bacteriol.
<4>192
<5>3240-3241
<6>2010
<7>The complete genome sequence of 'Candidatus Puniceispirillum marinum' IMCC1322, the first
cultured representative of the SAR116 clade in the
Alphaproteobacteria, is reported here. The genome contains genes for
proteorhodopsin, aerobic-type carbon monoxide dehydrogenase,
dimethylsulfoniopropionate demethylase, and C(1) compound metabolism. The
genome information proposes the SAR116 group to be metabolic generalists
in ocean nutrient cycling.

<>

<1>Oh, H.M., Lee, K., Jang, Y., Kang, I., Kim, H.J., Kang, T.W., Kim, S.Y., Cho, J.C.
<2>Genome Sequence of Strain IMCC9480, a Xanthorhodopsin-bearing Betaproteobacterium Isolated from the Arctic Ocean.
<3>J. Bacteriol.
<4>193
<5>3421
<6>2011
<7>Strain IMCC9480 is a novel member of the family Oxalobacteraceae of the Betaproteobacteria,
isolated from the Arctic Ocean by
dilution-to-extinction culturing. Here we present the draft genome
sequence of strain IMCC9480. The genome is predicted to contain genes for
xanthorhodopsin, retinoid biosynthesis, carbon monoxide dehydrogenase, and
C1 metabolism.

<>

<1>Oh, J.D., Kling-Backhed, H., Giannakis, M., Xu, J., Fulton, R.S., Fulton, L.A., Cordum, H.S., Wang, C., Elliott, G., Edwards, J., Mardis, E.R., Engstrand, L.G., Gordon, J.I.
<2>The complete genome sequence of a chronic atrophic gastritis Helicobacter pylori strain: Evolution during disease progression.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>9999-10004
<6>2006
<7>Helicobacter pylori produces acute superficial gastritis in nearly all of its human hosts.
However, a subset of individuals develops chronic atrophic gastritis (ChAG), a condition
characterized in part by diminished numbers of acid-producing parietal cells and increased
risk for development of gastric adenocarcinoma. Previously, we used a gnotobiotic transgenic
mouse model with an engineered ablation of parietal cells to show that loss of parietal cells
provides an opportunity for a H. pylori isolate from a patient with ChAG (HPAG1) to bind to,
enter, and persist within gastric stem cells. This finding raises the question of how ChAG
influences H. pylori genome evolution, physiology, and tumorigenesis. Here we describe the
1,596,366-bp HPAG1 genome. Custom HPAG1 Affymetrix GeneChips, representing 99.6% of its
predicted ORFs, were used for whole-genome genotyping of additional H. pylori ChAG isolates
obtained from Swedish patients enrolled in a case-control study of gastric cancer, as well as
ChAG- and cancer-associated isolates from an individual who progressed from ChAG to gastric
adenocarcinoma. The results reveal a shared gene signature among ChAG strains, as well as
genes that may have been lost or gained during progression to adenocarcinoma. Whole-genome
transcriptional profiling of HPAG1's response to acid during in vitro growth indicates that
genes encoding components of metal uptake and utilization pathways, outer membrane proteins,
and virulence factors are among those associated with H. pylori's adaptation to ChAG.

<>

<1>Oh, S., Zhang, R., Wu, Q.L., Liu, W.T.
<2>Draft Genome Sequence of a Novel SAR11 Clade Species Abundant in a Tibetan Lake.
<3>Genome Announcements
<4>2
<5>e01137-14
<6>2014
<7>SAR11 clade bacteria are abundant and play a key role in the nutrient cycles of marine and,
presumably, inland aquatic environments. We report here the draft
genome sequence of a novel species in the SAR11 cluster, reconstructed from a
metagenomic data set obtained from a Tibetan lake.

<>

<1>Ohji, S., Yamazoe, A., Hosoyama, A., Tsuchikane, K., Ezaki, T., Fujita, N.
<2>The Complete Genome Sequence of Pseudomonas putida NBRC 14164T Confirms High Intraspecies Variation.
<3>Genome Announcements
<4>2
<5>e00029-14
<6>2014
<7>Pseudomonas putida has attracted much interest for its environmental, industrial,
biotechnological, and clinical importance. Here, we report the complete genome
sequence of the type strain P. putida NBRC 14164. This genome sequence will
assist to further elucidate the molecular mechanisms of the characteristic traits
among strains belonging to the species P. putida.

<>

<1>Ohkuma, M., Yuki, M., Oshima, K., Suda, W., Oshida, Y., Kitamura, K., Iida, T., Hattori, M.
<2>Draft Genome Sequence of the Alkaliphilic and Xylanolytic Paenibacillus sp. Strain JCM 10914, Isolated from the Gut of a Soil-Feeding Termite.
<3>Genome Announcements
<4>2
<5>e01144-13
<6>2014
<7>Panibacillus sp. strain JCM 10914 is a xylanolytic alkaliphile isolated from the  gut of a
soil-feeding termite. Its draft genome sequence revealed various genes
for hydrolytic enzymes and will facilitate studies on adaptation to the highly
alkaline gut environment and its role in digesting soil organic matter in the
gut.

<>

<1>Ohnishi, N., Maruyama, F., Ogawa, H., Kachi, H., Yamada, S., Fujikura, D., Nakagawa, I., Hang'ombe, M.B., Thomas, Y., Mweene, A.S., Higashi, H.
<2>Genome Sequence of a Bacillus anthracis Outbreak Strain from Zambia, 2011.
<3>Genome Announcements
<4>2
<5>e00116-14
<6>2014
<7>In August 2011, an anthrax outbreak occurred among Hippopotamus amphibius hippopotamuses and
humans in Zambia. Here, we report the draft genome sequence of
the Bacillus anthracis outbreak strain CZC5, isolated from tissues of H.
amphibius hippopotamuses that had died in the outbreak area.

<>

<1>Ohnishi, Y., Ishikawa, J., Hara, H., Suzuki, H., Ikenoya, M., Ikeda, H., Yamashita, A., Hattori, M., Horinouchi, S.
<2>Genome sequence of the streptomycin-producing microorganism Streptomyces griseus IFO 13350.
<3>J. Bacteriol.
<4>190
<5>4050-4060
<6>2008
<7>We determined the complete genome sequence of Streptomyces griseus IFO 13350, a soil bacterium
producing an antituberculosis agent, streptomycin, which is the first aminoglycoside
antibiotic, discovered more than 60 years ago. The linear chromosome consists of 8,545,929
base pairs (bp), with an average G+C content of 72.2%, predicting 7,138 open reading frames,
six rRNA operons (16S-23S-5S), and 66 tRNA genes. It contains extremely long terminal inverted
repeats (TIRs) of 132,910 bp each. The telomere's nucleotide sequence and secondary
structure, consisting of several palindromes with a loop sequence of 5'-GGA-3', are
different from those of typical telomeres conserved among other Streptomyces species. In
accordance with the difference, the chromosome has pseudogenes for a conserved terminal
protein (Tpg) and a telomere-associated protein (Tap), and a novel pair of Tpg and Tap
proteins is instead encoded by the TIRs. Comparisons with the genomes of two related species,
Streptomyces coelicolor A3(2) and Streptomyces avermitilis, clarified not only the
characteristics of the S. griseus genome but also the existence of 24 Streptomyces-specific
proteins. The S. griseus genome contains 34 gene clusters or genes for the biosynthesis of
known or unknown secondary metabolites. Transcriptome analysis using a DNA microarray showed
that at least four of these clusters, in addition to the streptomycin biosynthesis gene
cluster, were activated directly or indirectly by AdpA, which is a central transcriptional
activator for secondary metabolism and morphogenesis in the A-factor (a gamma-butyrolactone
signaling molecule) regulatory cascade in S. griseus.

<>

<1>Ohno, S., Handa, N., Watanabe-Matsui, M., Takahashi, N., Kobayashi, I.
<2>Maintenance forced by a restriction-modification system can be modulated by a region in its modification enzyme not essential for methyltransferase activity.
<3>J. Bacteriol.
<4>190
<5>2039-2049
<6>2008
<7>Several type II restriction-modification gene complexes can force their maintenance on their
host bacteria by killing cells that have lost them
in a process called postsegregational killing or genetic addiction. It
is likely to proceed by dilution of the modification enzyme molecule
during rounds of cell division following the gene loss, which exposes
unmethylated recognition sites on the newly replicated chromosomes to
lethal attack by the remaining restriction enzyme molecules. This
process is in apparent contrast to the process of the classical types
of postsegregational killing systems, in which built-in metabolic
instability of the antitoxin allows release of the toxin for lethal
action after the gene loss. In the present study, we characterize a
mutant form of the EcoRII gene complex that shows stronger capacity in
such maintenance. This phenotype is conferred by an L80P amino acid
substitution (T239C nucleotide substitution) mutation in the
modification enzyme. This mutant enzyme showed decreased DNA
methyltransferase activity at a higher temperature in vivo and in vitro
than the nonmutated enzyme, although a deletion mutant lacking the
N-terminal 83 amino acids did not lose activity at either of the
temperatures tested. Under a condition of inhibited protein synthesis,
the activity of the L80P mutant was completely lost at a high
temperature. In parallel, the L80P mutant protein disappeared more
rapidly than the wild-type protein. These results demonstrate that the
capability of a restriction-modification system in forcing maintenance
on its host can be modulated by a region of its antitoxin, the
modification enzyme, as in the classical postsegregational killing
systems.

<>

<1>Ohsawa, K., Imai, Y., Ito, D., Kohsaka, S.
<2>Molecular cloning and characterization of annexin V-binding proteins with highly hydrophilic peptide structure.
<3>J. Neurochem.
<4>67
<5>89-97
<6>1996
<7>We previously reported that annexin V promoted the survival of cultured rat neocortical
neurons.  In an effort to elucidate the mechanism underlying this neurotrophic activity of
annexin V, we have attempted to identify the target or binding proteins of annexin V in
neuronal cells.  Herein, we screened an embryonic day 17 rat brain cDNA library by western
blot using glutathione S-transferase-annexin V fusion protein as a probe and then isolated
four clones showing binding to annexin V in a ca2+ - and phospholipid-dependent manner.
Although these cDNAs encoded different polypeptides, all four polypeptides shared the unique
feature of containing highly hydrophilic stretches with high Lys, Glu, and Ser contents.
Deduced amino acid sequences of two clones showed high homology with human X-linked Helicase2
(XH2) and DNA (cytosine-5) methyltransferase (DMTase) sequences, whereas the other two were
not related to any known peptide sequence.  These results suggest that XH2 and DMTase are
candidates for annexin V-binding proteins and thus may mediate the biological activity of
annexin V.

<>

<1>Ohshima, H., Matsuoka, S., Asai, K., Sadaie, Y.
<2>Molecular organization of intrinsic restriction and modification genes BsuM of Bacillus subtilis Marburg.
<3>J. Bacteriol.
<4>184
<5>381-389
<6>2002
<7>Transcriptional analysis and disruption of five open reading frames (ORFs), ydiO, ydiP, ydiR,
ydiS, and ydjA, in the prophage 3 region of the chromosome of Bacillus subtilis Marburg
revealed that they are component genes of the intrinsic BsuM restriction and modification
system of this organism. The classical mutant strain RM125, which lacks the restriction and
modification system of B. subtilis Marburg, lacks the prophage 3 region carrying these five
ORFs. These ORFs constitute two operons, the ydiO-ydiP operon and the ydiR-ydiS-ydjA operon,
both of which are expressed during the logarithmic phase of growth. The predicted gene
products YdiO and YdiP are the orthologues of cytosine DNA methyltransferases. The predicted
YdiS product is an orthologue of restriction nucleases, while the predicted YdiR and YdjA
products have no apparent paralogues and orthologues whose functions are known. Disruption of
the ydiR-ydiS-ydjA operon resulted in enhanced transformation by plasmid DNA carrying multiple
BsuM target sequences. Disruption of ydiO or ydiP function requires disruption of at least one
of the following genes on the chromosome: ydiR, ydiS, and ydjA. The degrees of methylation of
the BsuM target sequences on chromosomal DNAs were estimated indirectly by determining the
susceptibility to digestion with XhoI (an isoschizomer of BsuM) of DNAs extracted from the
disruptant strains. Six XhoI (BsuM) sites were examined. XhoI digested at the XhoI sites in
the DNAs from disruptants with disruptions in both operons, while XhoI did not digest at the
XhoI sites in the DNAs from the wild-type strain or from the disruptants with disruptions in
the ydiR-ydiS-ydjA operon. Therefore, the ydiO-ydiP operon and the ydiR-ydiS-ydjA operon are
considered operons that are responsible for BsuM modification and BsuM restriction,
respectively.

<>

<1>Ohta, K., Keszenman-Pereyra, D., Nicolas, A., Shibata, T.
<2>A novel meiosis-induced site-specific endonuclease activity in yeast mitochondria.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>174
<6>1993
<7>Sequence-specific endonucleases in eukaryotic cells have been shown to play a crucial role in
the initiation of genetic recombination. We have found a sequence-specific DNA endonuclease
activity that cuts DNAs of phage Phi105c and phage Phix174 in a cell-free extracts of a
synchronously-sporulating strain (SK1) of S. cerevisiae. The preparation from SK1 cells in
early meiotic phase had 40-fold higher activity than that in exponential growing phase.
Subcellular fractionation revealed that this activity is condensed in the crude mitochondrial
fraction. In addition, the activity to cut Phi105c DNA was not detected in the extract from
p-mutant (large deletion mutations that block all mitochondrial protein synthesis) of the SK1
strain. Furthermore, the ability to produce the endonuclease was able to be transferred into a
po strain by mitochondrial transfer through cytoduction. Comparison of nucleotide sequences
around the cutting sites in Phi 105c and Phix174 DNA suggests that the endonuclease recognizes
a -20 base pair sequence and generates ends with 4 base-5' overhang. The sequences around the
cutting sites are different from those by an other yeast mitochondrial endonucleases. As well
as other yeast mitochondrial sequence-specific endonucleases, the endonuclease in SK1 strain
is speculated to initiate gene conversion in the mitochondrial genome.

<>

<1>Ohta, K., Nicolas, A., Keszenman-Pereyra, D., Shibata, T.
<2>Endo.SK1: an inducible site-specific endonuclease from yeast mitochondria.
<3>Mol. Gen. Genet.
<4>250
<5>395-404
<6>1996
<7>Site-specific endonucleases have been found in various eukaryotic organelles such
as mitochondria, chloroplasts and nuclei.  These endonucleases initiate site-specific or
homologous
gene conversion in mitochondrial and nuclear DNA.  Here, we report a new site-specific
endonuclease activity, Endo.SK1, identified in mitochondria of strain SK1, a homothallic
diploid
strain of Saccharomyces cerevisiae.  Nucleotide sequences around the Endo.SK1-cleavage sites
are
different from those of known yeast site-specific endonucleases.  The Endo.SK1 activity is, at
least partly, specified by a gene in the Sk1-derived mitochondria.  A novel feature of the
Endo.SK1 activity is its inducibility: the endonuclease activity was induced by ca. 40-fold by
transfer of cells from a glucose medium into an acetate medium, and was then repressed.  This
transient induction was independent of the ploidy level of the cells, and coincided with
induction of
fumarase, a mitochondrial enzyme involved in the TCA cycle.  Co-induction and co-repression of
the mitochondrial site-specific endonuclease activity and a respiration-related enzyme
indicate that
the endonuclease activity is regulated in response to physiological conditions, and suggest a
possible role for the endonuclease in mitochondrial DNA metabolism.

<>

<1>Ohtani, N., Sato, M., Tomita, M., Itaya, M.
<2>Restriction on conjugational transfer of pLS20 in Bacillus subtilis 168.
<3>Biosci. Biotechnol. Biochem.
<4>72
<5>2472-2475
<6>2008
<7>Conjugational transfer of pLS20 in Bacillus subtilis Marburg 168 is restricted by the BsuM
restriction modification system. Restriction
efficiency was measured using pLS20 derivatives possessing various
numbers of XhoI sites, which are known to be recognized by BsuM. An
increase in XhoI sites clearly reduced the conjugational efficiency of
pLS20 as compared with that of pUB110 plasmid lacking XhoI.

<>

<1>Ohtani, N., Tomita, M., Itaya, M.
<2>The third plasmid pVV8 from Thermus thermophilus HB8: isolation, characterization, and sequence determination.
<3>Extremophiles
<4>16
<5>237-244
<6>2012
<7>The extremely thermophilic bacterium Thermus thermophilus is a model organism for structural
biology and systems biology, and the so-called "Structural and Functional Whole-Cell Project
for T. thermophilus HB8" is in progress. The released genomic sequence of the strain HB8 is
composed of chromosome, pTT27 megaplasmid, and pTT8 plasmid. In this paper, however, a third
plasmid was demonstrated and its sequence was determined. Although this plasmid pVV8 had been
reported before, limited information and an unfortunate dropout in the substrain, whose
genomic sequence was determined, would have prevented the plasmid from coming to public
attention. The intrinsic circular plasmid, which was estimated to be six to ten copies in a
cell, is 81151 bp and its G + C content is 68%. Among the identified 91 ORFs, a single gene
has been experimentally analyzed before and is known as xylose isomerase. The phnCDEGHIJKLMX
operon related to phosphonate metabolism, alkaline phosphatase, putative transcriptional
regulators, several sets of toxin-antitoxin system, and transposase-like ORFs are also encoded
on the pVV8 plasmid. Although association with cell aggregation was the one phenotypic
characteristic of the plasmid that had been reported, it was never confirmed. Comparison of T.
thermophilus HB8 strains suggests that the pVV8 is nonessential for growth.

<>

<1>Ohtsubo, Y., Fujita, N., Nagata, Y., Tsuda, M., Iwasaki, T., Hatta, T.
<2>Complete Genome Sequence of Ralstonia pickettii DTP0602, a 2,4,6-Trichlorophenol  Degrader.
<3>Genome Announcements
<4>1
<5>e00903-13
<6>2013
<7>Ralstonia pickettii strain DTP0602 utilizes 2,4,6-trichlorophenol as its sole carbon and
energy source. Here, we report the complete genome sequence of strain
DTP0602, which comprises three chromosomes and no plasmids. We also found that
the two had gene clusters responsible for the degradation of
2,4,6-trichlorophenol are located on the 2.9-Mb chromosome 2.

<>

<1>Ohtsubo, Y., Kishida, K., Sato, T., Tabata, M., Kawasumi, T., Ogura, Y., Hayashi, T., Tsuda, M., Nagata, Y.
<2>Complete Genome Sequence of Pseudomonas sp. Strain TKP, Isolated from a gamma-Hexachlorocyclohexane-Degrading Mixed Culture.
<3>Genome Announcements
<4>2
<5>e01241-13
<6>2014
<7>Pseudomonas sp. strain TKP does not degrade gamma-hexachlorocyclohexane (gamma-HCH), but it
persistently coexists with the gamma-HCH-degrading
Sphingobium sp. strain TKS in a mixed culture enriched by gamma-HCH. Here, we
report the complete genome sequence of strain TKP, which consists of one circular
chromosome with a size of 7 Mb.

<>

<1>Ohtsubo, Y., Maruyama, F., Mitsui, H., Nagata, Y., Tsuda, M.
<2>Complete Genome Sequence of Acidovorax sp. Strain KKS102, a Polychlorinated-Biphenyl Degrader.
<3>J. Bacteriol.
<4>194
<5>6970-6971
<6>2012
<7>We report the complete genome sequence of Acidovorax sp. strain KKS102, a
polychlorinated-biphenyl-degrading strain isolated from a soil sample in Tokyo.
The genome contains a single circular 5,196,935-bp chromosome and no plasmids.

<>

<1>Ohtsubo, Y., Sato, T., Kishida, K., Tabata, M., Ogura, Y., Hayashi, T., Tsuda, M., Nagata, Y.
<2>Complete Genome Sequence of Pseudomonas aeruginosa MTB-1, Isolated from a Microbial Community Enriched by the Technical Formulation of  Hexachlorocyclohexane.
<3>Genome Announcements
<4>2
<5>e01130-13
<6>2014
<7>Pseudomonas aeruginosa MTB-1 does not degrade gamma-hexachlorocyclohexane (gamma-HCH), but
this bacterium persistently coexists with a gamma-HCH-degrading
strain, Sphingomonas sp. MM-1, in a microbial community enriched by the technical
formulation of HCH. Here we report the complete MTB-1 genome sequence, with a
6.6-Mb circular chromosome.

<>

<1>Ohtsuka, E., Ishino, Y., Ibaraki, K., Ikehara, M.
<2>Recognition by restriction endonuclease EcoRI of deoxyoctanucleotides containing modified sugar moieties.
<3>Eur. J. Biochem.
<4>139
<5>447-450
<6>1984
<7>The deoxyribooctanucleotide d(G-G-A-A-T-T-C-C), containing the recognition sequence for EcoRI,
d(G-A-A-T-T-C), and analogs containing modified sugar moieties were tested for their activity
in cleavage with EcoRI.  These analogs, with replacement in the third position from the 5'
end, were synthesized using 9-b-D-arabinosyladenine (aA), 2'-deoxy-2'-fluoroadenosine (Afl)
and adenosine (rA).  Duplex formation by the three analogs was confirmed by measurements of
ultraviolet/temperature profiles.  It was found that EcoRI cleaved these duplexes less
efficiently than d(G-G-A-A-T-T-C-C).  The adenosine-containing analog d(G-G)-rA-d(A-T-T-C-C)
was cleaved much more slowly than d(G-G)-aA-d(A-T-T-C-C) and d(G-G-Afl-A-T-T-C-C).  The
corresponding ribooctamer G-G-A-A-U-U-C-C showed a higher melting temperature than the
deoxyoctamers but its duplex was not cleaved by this enzyme.  An analog with
2'-deoxy-2'-fluoroguanosine at the second position was cleaved by the endonuclease faster
than the natural deoxyoctamer.

<>

<1>Ohtsuka, E., Morisawa, H., Ikehara, M.
<2>Studies on deoxynucleic acids and related compounds.  IV. Syntheses of an octanucleotide containing a recognition site for restriction enzyme EcoRI and of an arabinosyladenine analog.
<3>Chem. Pharm. Bull. (Tokyo)
<4>30
<5>874-880
<6>1982
<7>An octanucleotide containing a recognition site for EcoRI and its
arabinosyladenine (araA) analog, dGGAATTCC and dGGaraAATTCC, were synthesized
by the phosphotriester approach with phosphoro-p-anisidate as the protecting
group for 3'-phosphodiesters.  araA was converted to
5'-dimethoxytrityl-N,2'-O-O-benzoyl 3'-p-chlorophenyl phosphate and condensed
with N-benzoyldeoxyadenosine 3'-p-chlorophenyl phosphoro-p-anisidate to yield
the protected araAdAp.  Other deoxynucleotide blocks (dAAp, dGGp) were prepared
similarly and condensed with a 5'-deblocked dTTCC block having the 3'-O-benzoyl
group after removal of the p-anisidate group with isoamyl nitrite.

<>

<1>Ohtsuka, K., Ohnishi, H., Nozaki, E., Pais, R.J., Tortoli, E., Yonetani, S., Matsushima, S., Tateishi, Y., Matsumoto, S., Watanabe, T.
<2>Whole-Genome Sequence of Mycobacterium kyorinense.
<3>Genome Announcements
<4>2
<5>e01062-14
<6>2014
<7>We report here the first draft genome sequence of Mycobacterium kyorinense, which was
described in 2009 and exhibits significant pathogenicity to humans.

<>

<1>Oishi, K., Aoi, S., Ehara, Y., Otsuka, Y., Shimamura, K., Higuchi, Y., Nomoto, M.
<2>Inhibition of restriction endonucleases by commercial polysaccharides.
<3>J. Ferment. Bioeng.
<4>69
<5>360-361
<6>1990
<7>Commercial polysaccharide preparations were investigated for their restriction
enzyme-inhibitory activities.  Dextran sulfate (S content 18.5%) and laminaran
from Eisenia arborea (0.88%) had marked inhibitory activity and haparin (13.1%)
and iota-, kappa-, and lambda-carrageenans (3.0, 3.8, and 4.3%) showed moderate
inhibition.  The effects of sulfation level and the structure of the
carbohydrate moiety on the inhibitory activity were discussed.

<>

<1>Ojala, T., Kuparinen, V., Koskinen, J.P., Alatalo, E., Holm, L., Auvinen, P., Edelman, S., Westerlund-Wikstrom, B., Korhonen, T.K., Paulin, L., Kankainen, M.
<2>Genome Sequence of Lactobacillus crispatus ST1.
<3>J. Bacteriol.
<4>192
<5>3547-3548
<6>2010
<7>Lactobacillus crispatus is a common member of the beneficial microbiota present in the
vertebrate gastrointestinal and human genitourinary tracts.
Here, we report the genome sequence of L. crispatus ST1, a chicken isolate
displaying strong adherence to vaginal epithelial cells.

<>

<1>Oka, M., Meacham, A.M., Hamazaki, T., Rodic, N., Chang, L.-J., Terada, N.
<2>De novo DNA methyltransferases Dnmt3a and Dnmt3b primarily mediate the cytotoxic effect of 5-aza-2'-deoxycytidine.
<3>Oncogene
<4>24
<5>3091-3099
<6>2005
<7>The deoxycytidine analog 5-aza-2'-deoxycitidine (5-aza-dC) is a potent chemotherapeutic agent
effective against selective types of cancer. The molecular mechanism by which 5-aza-dC induces
cancer cell death, however, is not fully understood. It has been accepted that the mechanism
of toxicity is due to the covalent binding between the DNA methyltransferase (Dnmt) and
5-aza-dC-substituted DNA. In order to define which member of the Dnmt family plays a dominant
role in the cytotoxicity, we examined the effect of 5-aza-dC on cell growth and apoptosis in
various Dnmt null mutant embryonic stem (ES) cells. Of interest, Dnmt3a-Dnmt3b double null ES
cells were highly resistant to 5-aza-dC when compared to wild type, Dnmt3a null, Dnmt3b null,
or Dnmt1 null ES cells. The cellular sensitivity to 5-aza-dC correlated well with the
expression status of Dnmt3 in both undifferentiated and differentiated ES cells. When
exogenous Dnmt3a or Dnmt3b was expressed in double null ES cells, the sensitivity to 5-aza-dC
was partially restored. These results suggest that the cytotoxic effect of 5-aza-dC may be
mediated primarily through Dnmt3a and Dnmt3b de novo DNA methyltransferases. Further, the
ability to form Dnmt-DNA adducts was similar in Dnmt1 and Dnmt3, and the expression level of
Dnmt3 was not higher than that of Dnmt1 in ES cells. Therefore, Dnmt3-DNA adducts may be more
effective for inducing apoptosis than Dnmt1-DNA adducts. These results imply a therapeutic
potential of 5-aza-dC to cancers expressing Dnmt3.

<>

<1>Okada, K., Na-Ubol, M., Natakuathung, W., Roobthaisong, A., Maruyama, F., Nakagawa, I., Chantaroj, S., Hamada, S.
<2>Comparative Genomic Characterization of a Thailand-Myanmar Isolate, MS6, of Vibrio cholerae O1 El Tor, Which Is Phylogenetically Related to a 'US Gulf Coast' Clone.
<3>PLoS ONE
<4>9
<5>E98120
<6>2014
<7>BACKGROUND: The cholera outbreaks in Thailand during 2007-2010 were exclusively
caused by the Vibrio cholerae O1 El Tor variant carrying the cholera toxin gene
of the classical biotype. We previously isolated a V. cholerae O1 El Tor strain
from a patient with diarrhea and designated it MS6. Multilocus sequence-typing
analysis revealed that MS6 is most closely related to the U. S. Gulf Coast clone
with the exception of two novel housekeeping genes. METHODOLOGY/PRINCIPAL
FINDINGS: The nucleotide sequence of the genome of MS6 was determined and
compared with those of 26 V. cholerae strains isolated from clinical and
environmental sources worldwide. We show here that the MS6 isolate is distantly
related to the ongoing seventh pandemic V. cholerae O1 El Tor strains. These
strains differ with respect to polymorphisms in housekeeping genes, seventh
pandemic group-specific markers, CTX phages, two genes encoding predicted
transmembrane proteins, the presence of metY (MS6_A0927) or hchA/luxR in a highly
conserved region of the V. cholerae O1 serogroup, and a superintegron (SI). We
found that V. cholerae species carry either hchA/luxR or metY and that the V.
cholerae O1 clade commonly possesses hchA/luxR, except for MS6 and U. S. Gulf
Coast strains. These findings illuminate the evolutionary relationships among V.
cholerae O1 strains. Moreover, the MS6 SI carries a quinolone-resistance gene
cassette, which was closely related with those present in plasmid-borne integrons
of other gram-negative bacteria. CONCLUSIONS/SIGNIFICANCE: Phylogenetic analysis
reveals that MS6 is most closely related to a U. S. Gulf Coast clone, indicating
their divergence before that of the El Tor biotype strains from a common V.
cholerae O1 ancestor. We propose that MS6 serves as an environmental aquatic
reservoir of V. cholerae O1.

<>

<1>Okada, K., Ogura, Y., Hayashi, T., Abe, A., Kuwae, A., Horiguchi, Y., Abe, H.
<2>Complete Genome Sequence of Bordetella bronchiseptica S798, an Isolate from a Pig with Atrophic Rhinitis.
<3>Genome Announcements
<4>2
<5>e00436-14
<6>2014
<7>Bordetella bronchiseptica colonizes the respiratory tracts of a wide variety of mammals and
causes a range of diseases, from lethal pneumonia to asymptomatic
chronic infection. We report the complete genome sequence of Bordetella
bronchiseptica strain S798, isolated from a pig with atrophic rhinitis in Japan.

<>

<1>Okada, M.
<2>Host-controlled restriction and modification of Salmonella typhimurium.
<3>Keio J. Med.
<4>18
<5>81-97
<6>1969
<7>Since the discovery of sexuality in bacteria, hybridization between Escherichia
coli and Salmonella strains has been intensively studied by a number of
workers.  Despite initial uniformal negative results, hybrids did occur at much
lower frequencies than those between E. coli strains.  In the mating of
Salmonella, Hfr strains of E. coli have been employed as donors.  The hybrids
were viable and more fertile as recipients than the parent Salmonella strains,
in the sense of the frequency of hybrid formation per donor cell.  It was not
known, however, whether this barrier in gene exchange between these species
might be poor mating capacity or poor integration capacity or both.  In the
present paper, I shall show that fertile mutants, which were selected for
abnormally increased recipient ability in the transmission of an R factor 222
from E. coli (R+), are simultaneously accompanied by acquisition of the
increased recipient ability for E. coli chromosome and some alteration of
host-controlled restriction and modification as to R factors and phage P22.

<>

<1>Okada, R., Matsumoto, M., Zhang, Y., Isaka, M., Tatsuno, I., Hasegawa, T.
<2>Emergence of type I restriction modification system-negative emm1 type Streptococcus pyogenes clinical isolates in Japan.
<3>APMIS
<4>122
<5>914-921
<6>2014
<7>Streptococcus pyogenes emm1 type is the dominant cause of streptococcal toxic shock syndrome
(STSS) in Japan and many other developed countries. Recently, the number of STSS patients in
Japan was reported to be increasing. Hence, we analyzed the S. pyogenes clinical isolates
detected in Japan after 2005. We found that the regions encoding the Spy1908-1910
two-component regulatory system and the adjacent type I restriction modification system were
deleted in some emm1 type isolates. The isolates with the deletion were detected only in the
emm1 strains that were isolated between 2010 and 2013, but not before 2010. Twenty-six of 46
(56.5%) emm1 type isolates were isolated in 2010-2013, and among these isolates, five of seven
(71.4%) emm1 type STSS isolates were shown to have that deletion. PFGE and PCR analysis for
the presence of several pyrogenic exotoxin-related genes suggested that the emm1 isolates with
and without the deletion shared the same genetic background. The emm1 isolates with the
deletion could incorporate exogenous plasmids by experimental electroporation transformation
far more efficiently. These results suggested that the novel emm1 isolates have occupied a
fairly large part of total emm1 isolates.

<>

<1>Okamoto, A., Lee, H., Yabutani, M., Yamada, K., Ohta, M.
<2>Draft Genome Sequence of a Legionella pneumophila Serogroup 4 Strain Causing Legionellosis.
<3>Genome Announcements
<4>2
<5>e00602-14
<6>2014
<7>Here, we report the draft genome sequence of the Legionella pneumophila Nagoya-1  strain,
serogroup 4, which was isolated from a clinical sample from a patient
with legionellosis. Several virulence-associated genes, including those encoding
the type IV (Dot/Icm) secretion system and effector proteins, were highly
conserved.

<>

<1>Okamoto, A., Tanabe, K., Saito, I.
<2>Site-specific discrimination of cytosine and 5-methylcytosine in duplex DNA by peptide nucleic acids.
<3>J. Am. Chem. Soc.
<4>124
<5>10262-10263
<6>2002
<7>For site-specific discrimination of cytosine (C) and 5-methylcytosine ((m)C) in duplex DNA, we
developed a new method using peptide nucleic acids (PNAs). The combination of a PNA-assisted
DNA displacement complex and a fluorescein-labeled probe oligomer allowed the detection of
(m)C at the defined sites in target DNA using a restriction enzyme. After treatment of the
complex with a restriction enzyme, strong fluorescence emission was observed for the complex
containing C at the target site, whereas the fluorescence intensity for the complex containing
(m)C was extremely weak.

<>

<1>Okano, M.
<2>DNA methylation and DNA methyltransferases in mammals.
<3>Saibo Kogaku
<4>20
<5>381-386
<6>2001
<7>
<>

<1>Okano, M.
<2>DNA methyltransferases Dnmt3a and Dnmt3b are essential for de novo methylation in mouse early development.
<3>Jikken Igaku
<4>18
<5>468-471
<6>2000
<7>
<>

<1>Okano, M., Bell, D.W., Haber, D.A., Li, E.
<2>DNA methyltransferases Dnmt3a and Dnmt3b are essential for de novo methylation and mammalian development.
<3>Cell
<4>99
<5>247-257
<6>1999
<7>The establishment of DNA methylation patterns requires de novo methylation that occurs
predominantly during early development and gametogenesis in mice.  Here we demonstrate that
two recently identified DNA methyltransferases, Dnmt3a and Dnmt3b, are essential for de novo
methylation and for mouse development.  Inactivation of both genes by gene targeting blocks de
novo methylation in ES cells and early embryos, but it has no effect on maintenance of
imprinted methylation patterns.  Dnmt3a and Dnmt3b also exhibit nonoverlapping functions in
development, with Dnmt3b specifically required for methylation of centromeric minor satellite
repeats.  Mutations of human DNMT3B are found in ICF syndrome, a developmental defect
characterized by hypomethylation of pericentromeric repeats.  Our results indicate that both
Dnmt3a and Dnmt3b function as de novo methyltransferases that play important roles in normal
development and disease.

<>

<1>Okano, M., Li, E.
<2>Genetic analyses of DNA methyltransferase genes in mouse model system.
<3>J. Nutr.
<4>132
<5>2462S-2465S
<6>2002
<7>DNA methylation regulates important biological processes and is involved in tumorigenesis and
several human diseases, such as Rett and immunodeficiency, centromeric instability and facial
anomalies (ICF). The major objective of our research is to investigate the roles of DNA
methylation in mammals through genetic analysis of DNA methyltransferase genes in mouse and
human. Previously, we found that Dnmt1 knockout embryonic stem (ES) cells are capable of
methylating retroviral DNA de novo. In search of enzymes responsible for de novo methylation,
we have cloned a novel family of mammalian DNA methyltransferase genes, Dnmt3a and Dnmt3b.
Although extensive sequence similarity was found between Dnmt3a and Dnmt3b, little homology
was observed between Dnmt1 and Dnmt3a/3b in the catalytic domain as well as in the N-terminal
domain. Additionally, biochemical analysis revealed that, unlike Dnmt1, neither Dnmt3a nor
Dnmt3b had a strong preference to hemimethylated DNA substrates. Genetic analysis demonstrated
that Dnmt3a and Dnmt3b were required for de novo methylation activities in ES cells and during
early embryogenesis and were essential for early development. Interestingly, phenotype
analyses of single homozygous mice for either Dnmt3a or Dnmt3b suggested that the functions of
Dnmt3a and Dnmt3b also were required at the late developmental stage and even at the adult
stage.

<>

<1>Okano, M., Takebayashi, S., Okumura, K., Li, E.
<2>Assignment of cytosine-5 DNA methyltransferases Dnmt3a and Dnmt3b to mouse chromosome bands 12A2-A3 and 2H1 by in situ hybridization.
<3>Cytogenet. Cell Genet.
<4>86
<5>333-334
<6>1999
<7>Methylation of cytosine at the C-5 position is a major form of DNA modification in vertebrates
and plays important roles in regulation of gene expression and development.  Previously, only
one mammalian cytosine-5 methyltransferase (now termed Dnmt1) was known and shown to be
required for maintaining global DNA methylation levels.  Recently, we cloned a family of novel
cytosine-5 methyltransferase genes, termed Dnmt3a and Dnmt3b, which do not share sequence
similarities to any known eukaryotic cytosine-5 methyltransferase genes.  Expression pattern
and biological analysis suggest that Dnmt3a and Dnmt3b are probably responsible for de novo
methylation, a key process by which DNA methylation patterns are established during
development.  In this study we have mapped chromosome locations of Dnmt3a and Dnmt3b to
12A2-A3 and 2H1, respectively, by FISH.

<>

<1>Okano, M., Xie, S., Li, E.
<2>Dnmt2 is not required for de novo and maintenance methylation of viral DNA in embryonic stem cells.
<3>Nucleic Acids Res.
<4>26
<5>2536-2540
<6>1998
<7>We have shown previously that de novo methylation activities persist in mouse embryonic stem
cells homozygous for a null mutation of Dnmt1 that encodes the major DNA cytosine
methyltransferase.  In this study, we have cloned a putative mammalian DNA methyltransferase
gene, termed Dnmt2, that is homologous to pmt1 of fission yeast.  Different from pmt1 in which
the catalytic Pro-Pro-Cys motif is 'mutated' to Pro-Ser-Cys, Dnmt2 contains all the
conserved methyltransferase motifs, thus likely encoding a functional cytosine
methyltransferase.  However, baculovirus-expressed Dnmt2 protein failed to methylate DNA in
vitro.  To investigate whether Dnmt2 functions as a DNA methyltransferase in vivo, we
inactivated the Dnmt2 gene by targeted deletion of the putative catalytic PPC motif in ES
cells.  We showed that endogenous virus was fully methylated in Dnmt2-deficient mutant ES
cells.  Furthermore, newly integrated retrovirus DNA was methylated de novo in infected mutant
ES cells as efficiently as in wild-type cells.  These results indicate that Dnmt2 is not
essential for global de novo or maintenance methylation of DNA in ES cells.

<>

<1>Okano, M., Xie, S., Li, E.
<2>Cloning and characterization of a family of novel mammalian DNA (cytosine-5) methyltransferases.
<3>Nat. Genet.
<4>19
<5>219-220
<6>1998
<7>De novo methylation of genomic DNA is a developmentally regulated process that is believed to
play a pivotal role in regulation of genomic imprinting and X-chromosome inactivation in
mammals.  Aberrant de novo methylation of growth regulatory genes has been associated with
tumorigenesis in humans.  We have shown previously that de novo methylation persists in
embryonic stem cells lacking Dnmt1, which encodes the constitutive DNA methyltransferase Dnmt1
(or MT1), indicating the existence of independently encoded de novo methyltransferases.

<>

<1>Okhapkina, S.S., Netesova, N.A., Golikova, L.N., Seregina, E.V., Sosnovtsev, S.V., Abdurashitov, M.A., Degtyarev, S.K.
<2>Comparison of the homologous SfeI and LlaBI restriction-modification  systems.
<3>Mol. Biol. (Mosk)
<4>36
<5>432-437
<6>2002
<7>A fragment containing the SfeI restriction-modification system (RMS)  operon was cloned from a
Streptococcus faecalis SE72 plasmid. Nucleotide sequence analysis revealed its high (99.2%)
homology to the operon for Lactococcus lactis subsp. cremoris W56 LlaBI RMS recognizing the
same site, 5'-CTRYAG-3'. A substantial difference was that SfeI RMS operon had an additional
198-bp fragment and a larger gene for the putative control protein. No homology was observed
between operon-flanking sequences of the two closely related species, suggesting horizontal
transfer of the operon.

<>

<1>Okinaka, R.T., Challacombe, J., Drees, K., Birdsell, D.N., Janke, N., Naumann, A., Seymour, M., Hornstra, H., Schupp, J., Sahl, J., Foster, J.T., Pearson, T., Turnbull, P., Keim, P.
<2>Genome Sequence of Bacillus anthracis STI, a Sterne-Like Georgian/Soviet Vaccine  Strain.
<3>Genome Announcements
<4>2
<5>e00853-14
<6>2014
<7>The Bacillus anthracis strain STI is a Soviet vaccine strain that lacks the pXO2  plasmid.
Previous data indicate that this isolate forms a new branch within the
B. anthracis sub-group originally identified as A. Br.008/009.

<>

<1>Okstad, O.A., Hegna, I., Lindback, T., Rishovd, A.-L., Kolsto, A.-B.
<2>Genome organization is not conserved between Bacillus cereus and Bacillus subtilis.
<3>Microbiology
<4>145
<5>621-631
<6>1999
<7>The opportunistic pathogen Bacillus cereus is the genetically stable member of a group of
closely related bacteria including the insect pathogen Bacillus thuringiensis and the
mammalian pathogen Bacillus anthracis. Physical maps of B. cereus and B. thuringiensis strains
show considerable variations in discrete parts of the chromosome, suggesting that certain
genome regions are more prone to rearrangements. B. cereus belongs to the same subgroup of
Bacillus species as Bacillus subtilis, by both phenotypic and rRNA sequence classification.
The analysis of 80 kb of genome sequence sampled from different regions of the B. cereus ATCC
10987 chromosome is reported.  Analysis of the sequence and comparison of the localization of
the putative genes with that of B. subtilis orthologues show the following: (1) gene
organization is not conserved between B. cereus and B. subtilis; (2) several putative genes
are more closely related to genes from other bacteria and archaea than to B. subtilis, or may
be absent in B. subtilis 168; (3) B. cereus contains a 155 bp repetitive sequence that is not
present in B. subtilis. By hybridization, this repeat is present in all B. cereus and B.
thuringiensis strains so far investigated.

<>

<1>Oktavcovca, B., Godany, A., Pristas, P., Stevcikova, B., Farkasovska, J.
<2>Isolation and characterization of the modification methylase M.SauLPI from Streptomyces aureofaciens B-96.
<3>Nucleic Acids Res.
<4>21
<5>4843
<6>1993
<7>In our previous work we reported the presence of a GCC/GGC recognizing restriction system in
tetracycline producing strains of Streptomyces aureofaciens. In this paper we describe the
characterization of the cognate modification DNA methyltransferase M.SauLPI from
S.aureofaciens strain B-96.

<>

<1>Okubo, T. et al.
<2>Complete Genome Sequence of Bradyrhizobium sp. S23321: Insights into Symbiosis Evolution in Soil Oligotrophs.
<3>Microbes Environ.
<4>27
<5>306-315
<6>2012
<7>Bradyrhizobium sp. S23321 is an oligotrophic bacterium isolated from paddy field soil.
Although S23321 is
phylogenetically close to Bradyrhizobium japonicum USDA110, a legume symbiont, it is unable to
induce root nodules
in siratro, a legume often used for testing Nod factor-dependent nodulation. The genome of
S23321 is a single circular
chromosome, 7,231,841 bp in length, with an average GC content of 64.3%. The genome contains
6,898 potential
protein-encoding genes, one set of rRNA genes, and 45 tRNA genes. Comparison of the genome
structure between
S23321 and USDA110 showed strong colinearity; however, the symbiosis islands present in
USDA110 were absent
in S23321, whose genome lacked a chaperonin gene cluster (groELS3) for symbiosis regulation
found in USDA110.
A comparison of sequences around the tRNA-Val gene strongly suggested that S23321 contains an
ancestral-type
genome that precedes the acquisition of a symbiosis island by horizontal gene transfer.
Although S23321 contains a
nif (nitrogen fixation) gene cluster, the organization, homology, and phylogeny of the genes
in this cluster were more
similar to those of photosynthetic bradyrhizobia ORS278 and BTAi1 than to those on the
symbiosis island of USDA110.
In addition, we found genes encoding a complete photosynthetic system, many ABC transporters
for amino acids and
oligopeptides, two types (polar and lateral) of flagella, multiple respiratory chains, and a
system for lignin monomer
catabolism in the S23321 genome. These features suggest that S23321 is able to adapt to a wide
range of environments,
probably including low-nutrient conditions, with multiple survival strategies in soil and
rhizosphere.

<>

<1>Okubo, T., Fukushima, S., Itakura, M., Oshima, K., Longtonglang, A., Teaumroong, N., Mitsui, H., Hattori, M., Hattori, R., Hattori, T., Minamisawa, K.
<2>Soil oligotrophic bacterium Agromonas oligotrophica (Bradyrhizobium oligotrophicum) is a nitrogen-fixing symbiont of Aeschynomene indica as suggested by genome analysis.
<3>Appl. Environ. Microbiol.
<4>79
<5>2542-2551
<6>2013
<7>Agromonas oligotrophica (Bradyrhizobium oligotrophicum) S58T is a nitrogen-fixing oligotrophic
bacterium isolated from paddy field soil that is able to grow in extra-low nutrient
environments.  Here, the complete genome sequence of S58 was determined.  The S58 genome was
found to comprise a circular chromosome of 8,264,165 bp with an average GC content of 65.1%
lacking nodABC genes and typical symbiosis island.  The genome showed a high level of
similarity to the genomes of Bradyrhizobium sp. ORS278 and Bradyrizobium sp. BTAil including
nitrogen fixation and photosynthesis gene clusters, which nodulate an aquatic legume plant,
Aeschynomene indica, in a Nod factor-independent manner.  Although non-symbiotic
(brady)rhizobia are significant components of rhizobial populations in soil, we found that
most genes important for nodule development (ndv) and symbiotic nitrogen fixation (nif and
fix) with A. indica were well conserved between the ORS278 and S58 genomes.  Therefore, we
performed inoculation experiments with five A. oligotrophica strians (S58, S42, S55, S72, and
S80).  Surpirsingly, all five strains of A. oligotrophica formed effective nitrogen-fixing
nodules on the roots and/or stems of A. indica with differentiated bacteroids.  Non-symbiotic
(brady)rhizobia are known to be significant components of rhizobial populations without
symbiosis isoland or symbiotic plasmids in soil, but the present results indicate that
soil-dwelling A. oligotrophica generally possesses the ability to establish symbiosis with A.
indica is a common trait of nodABC- and symbiosis island-lacking strains within the members of
photosynthetic Bradyrhizobium clade including A. oligotrophica.

<>

<1>Okuda, Y., Sasaki, D., Nogami, S., Kaneko, Y., Ohya, Y., Anraku, Y.
<2>Occurrence, horizontal transfer and degeneration of VDE intein family in Saccharomycete yeasts.
<3>Yeast
<4>20
<5>563-573
<6>2003
<7>VDE is a homing endonuclease gene originally discovered as an intervening element in VMA1s of
Saccharomyces cerevisiae. There have been two
independent subfamilies of VDE, one from S. cerevisiae strain X2180-1A and
the other from Saccharomyces sp. DH1-1A in the host VMA1 gene, and they
share the identity of 96.3%. In order to search the occurrence,
intra/interspecies transfer and molecular degeneration of VDE, complete
sequences of VMA1 in 10 strains of S. cerevisiae, eight species of
saccharomycete yeasts, Candida glabrata and Kluyveromyces lactis were
determined. We found that six of 10 S. cerevisiae strains contain VDEs
99.7-100% identical to that of the strain X2180-1A, one has no VDE,
whereas the other three harbour VDEs 100% identical to that of the strain
DH1-1A. S. carlsbergensis has two VMA1s, one being 99.8% identical to that
of the strain X2180-1A with VDE 100% identical to that of the strain
DH1-1A and the other containing the same VMA1 in S. pastorianus with no
VDE. This and other evidence indicates that intra/interspecies
transmissions of VDEs have occurred among saccharomycete yeasts.
Phylogenetic analyses of VMA1 and VDE suggest that the S. cerevisiae VDEs
had branched earlier than other VDEs from an ancestral VDE and had invaded
into the host loci as relatively late events. The two VDEs seemed to
degenerate in individual host loci, retaining their splicing capacity
intact. The degeneration of the endonuclease domains was distinct and, if
compared, its apparent rate was much faster than that of the
protein-splicing domains.

<>

<1>Okumura, K., Arai, R., Okura, M., Kirikae, T., Takamatsu, D., Osaki, M., Miyoshi-Akiyama, T.
<2>Complete Genome Sequence of Melissococcus plutonius ATCC 35311.
<3>J. Bacteriol.
<4>193
<5>4029-4030
<6>2011
<7>We report the first completely annotated genome sequence of Melissococcus plutonius ATCC
35311. M. plutonius is a one genus one species bacterium,
and the etiological agent of European foulbrood of the honey bee. The
genome sequence will provide new insights into the molecular mechanisms
underlying its pathogenicity.

<>

<1>Okura, M., Takamatsu, D., Maruyama, F., Nozawa, T., Nakagawa, I., Osaki, M., Sekizaki, T., Gottschalk, M., Kumagai, Y., Hamada, S.
<2>Genetic Analysis of Capsular Polysaccharide Synthesis Gene Clusters from All Serotypes of Streptococcus suis: Potential Mechanisms for the Generation of Capsular Variation.
<3>Appl. Environ. Microbiol.
<4>79
<5>2796-2806
<6>2013
<7>Streptococcus suis strains are classified into 35 serotypes on the basis of the
antigenicity of their capsular polysaccharides (CPs). CP synthesis genes are
known to be clustered on the chromosome (cps gene cluster). The entire cps gene
clusters of S. suis have so far been sequenced in 15 serotypes and found to be
located between orfZ and aroA. In this study, to provide comprehensive
information about S. suis CPs, we sequenced the entire cps gene clusters of the
remaining serotypes and analyzed the complete set of S. suis cps gene clusters.
Among the 35 cps gene clusters, 22 were located between orfZ and aroA, whereas
the other 13 were flanked by other gene(s) on the chromosomes, and the
chromosomal locus was classified into five patterns. By clustering analysis, the
predicted products of cps genes found in the 35 serotypes were assigned into 291
homology groups, and all serotypes possessed a serotype-specific gene, except for
serotypes 1, 2, 1/2 and 14. Because of the presence of genes encoding flippase
(wzx) and polymerase (wzy), CPs of all serotypes were thought to be synthesized
by the Wzx/Wzy pathway. Our data also implied the possibility of the transfer of
the entire or partial cps gene clusters among S. suis strains, as well as the
influence of spontaneous mutations in a single or a few genes on the antigenicity
of some serotypes. Accumulation of these gene transfers and small-scale mutations
may have generated the antigenic diversity of S. suis CPs.

<>

<1>Olano, C., Cano-Prieto, C., Losada, A.A., Bull, A.T., Goodfellow, M., Fiedler, H.P., Mendez, C., Salas, J.A.
<2>Draft Genome Sequence of Marine Actinomycete Streptomyces sp. Strain NTK 937, Producer of the Benzoxazole Antibiotic Caboxamycin.
<3>Genome Announcements
<4>2
<5>e00534-14
<6>2014
<7>Streptomyces sp. strain NTK 937 is the producer of the benzoxazole antibiotic caboxamycin,
which has been shown to exert inhibitory activity against
Gram-positive bacteria, cytotoxic activity against several human tumor cell
lines, and inhibition of the enzyme phosphodiesterase. In this genome
announcement, we present a draft genome sequence of Streptomyces sp. NTK 937 in
which we identified at least 35 putative secondary metabolite biosynthetic gene
clusters.

<>

<1>Old, R., Murray, K., Roizes, G.
<2>Recognition Sequence of Restriction Endonuclease III from Hemophilus influenzae.
<3>J. Mol. Biol.
<4>92
<5>331-339
<6>1975
<7>An endonuclease, R.HindIII, prepared from Hemophilus influenzae strain Rd,
degrades foreign DNA, but not homologous DNA.  Phage T7 DNA is also resistant
to the enzyme.  Fragments of phage lambda DNA produced by treatment with
R.HindIII have been labelled at their 5' termini and analysis of the
radioactive nucleotides in pancreatic DNAase digests of these fragments
revealed a single 5' terminal sequence.  From this and other data we conclude
that the enzyme recognizes and cleaves DNA at the following nucleotide
sequence, 3' -N-T-T-C-G-A-A-N- 5' 5' -N-A-A-G-C-T-T-N- 3' giving termini
bearing short cohesive ends.

<>

<1>Oleastro, M., Monteiro, L., Lehours, P., Megraud, F., Menard, A.
<2>Identification of Markers for Helicobacter pylori Strains Isolated from Children with Peptic Ulcer Disease by Suppressive Subtractive Hybridization.
<3>Infect. Immun.
<4>74
<5>4064-4074
<6>2006
<7>Peptic ulcer disease (PUD) occurs after a long-term Helicobacter pylori
infection. However, the disease can develop earlier, and rare cases have
been observed in children, suggesting that these H. pylori strains may be
more virulent. We used suppressive subtractive hybridization for
comparative genomics between H. pylori strains isolated from a 5-year-old
child with duodenal ulcer and from a sex- and age-matched child with
gastritis only. The prevalence of the 30 tester-specific subtracted
sequences was determined on a collection of H. pylori strains from
children (15 ulcers and 30 gastritis) and from adults (46 ulcers and 44
gastritis). Two of these sequences, jhp0562 (80.0% versus 33.3%, P =
0.008) and jhp0870 (80.0% versus 36.7%, P = 0.015), were highly associated
with PUD in children and a third sequence, jhp0828, was less associated
(40.0% versus 10.0%, P = 0.048). Among adult strains, none of the 30
sequences was associated with PUD. However, both jhp0562 and jhp0870 were
less prevalent in adenocarcinoma strains than in PUD strains from children
and adults, the difference being statistically significant for jhp0870. In
conclusion, two H. pylori genes were identified as being strongly
associated with PUD in children, and their putative roles as an outer
membrane protein for jhp0870 and in lipopolysaccharide biosynthesis for
jhp0562, suggest that they may be novel virulence factors of H. pylori.

<>

<1>Olhoft, P.M.
<2>Cloning and characterization of the 5-methylcytosine methyltransferase gene in maize plants and tissue cultures.
<3>Diss. Abstr.
<4>59
<5>4638
<6>1999
<7>The genomic sequence of maize containing the methyltransferase gene called Zmet1 was
successfully cloned and sequenced.  Seven clones were identified from a genomic library by
using a highly conserved region from an Arabidopsis EST homologous to the Met1
methyltransferase gene as a probe.  The assembled genomic sequence of four overlapping clones
covers both the 5' and 3' flanking regions of the maize methyltransferase gene totaling
7,955 nucleotides.  Sequence alignments with the Arabidopsis Met1 cDNA revealed that the open
reading frame of Zmet1 encodes a putative protein of 1,525 amino acids, which is interrupted
in the genomic sequence by ten introns.  Northern analysis confirmed this result by the
identification of a single 4.6 kb RNA transcript.  The structure of the Zmet1 methylase is
similar to the other eukaryotic maintenance methylases; a large N-terminal domain of 1,054
amino  acids linked to a smaller C-terminal domain of 471 amino acids by a lysine-rich
sequence.  Zmet1 is highly expressed in actively dividing cells, namely in seedling tissue and
rapidly dividing callus tissue.  Restriction analysis suggests that there are at least two
Zmet1 loci in the maize genome, one of which maps to the short arm of chromosome 7 in bin 2.
The percent of 5-methylcytosine in maize DNA was shown to be dependent on the type of tissue
and the stage in development.  Although the amount of repetitive DNA remained stable in the
overall G/C to A/T ratio, methylation levels were found to significantly decrease from
15-day-old embryos to one-week-old seedlings.  Remethylation occurs between the first and
second week of seedling growth.  These changes indicate that there are both de novo
methylation and demethylation activities in early development.  The methylation pattern at
four low-copy sequences remained unchanged throughout plant and callus development except for
a possible hypermethylation event in the third month of cell culture.  However, there was
significant demethylation in the repetitive sequences, rDNA and COS 12, throughout an eight
month period in tissue culture.  The identification of stages or tissues which are undergoing
demethylation and de novo methylation may be important in identifying undiscovered methylase
and demethylase enzymes, developmentally regulated genes, as well as interacting proteins that
may regulate methylase functions.

<>

<1>Oliveira, D.C., Wu, S.W., de Lencastre, H.
<2>Genetic organization of the downstream region of the mecA element in methicillin-resistant Staphylococcus aureus isolates carrying different polymorphisms of this region.
<3>Antimicrob. Agents Chemother.
<4>44
<5>1906-1910
<6>2000
<7>We describe here the genetic organization of the mec element downstream of
the mecA gene in 34 different methicillin-resistant Staphylococcus aureus
(MRSA) clinical isolates carrying 13 of the most frequent polymorphisms of
mecA and representing the major epidemic clones of MRSA. All polymorphisms
carried three common genetic elements: the hypervariable region, a copy of
IS431, and a unique 2-kb sequence (downstream constant segment, or dcs)
for which no homologous sequences are found in data banks. Polymorphisms
of the downstream region were shown to be caused by the presence of
linearized plasmids flanked by insertion sequences (pUB110, pT181, and
pI258) and the autonomous insertion sequence IS256.

<>

<1>Oliveira, L.C. et al.
<2>Genome Sequence of Lactococcus lactis subsp. lactis NCDO 2118, a GABA-Producing Strain.
<3>Genome Announcements
<4>2
<5>e00980-14
<6>2014
<7>Lactococcus lactis subsp. lactis NCDO 2118 is a nondairy lactic acid bacterium, a xylose
fermenter, and a gamma-aminobutyric acid (GABA) producer isolated from
frozen peas. Here, we report the complete genome sequence of L. lactis NCDO 2118,
a strain with probiotic potential activity.

<>

<1>Oliveira, P.H., Touchon, M., Rocha, E.P.
<2>The interplay of restriction-modification systems with mobile genetic elements and their prokaryotic hosts.
<3>Nucleic Acids Res.
<4>42
<5>10618-10631
<6>2014
<7>The roles of restriction-modification (R-M) systems in providing immunity against horizontal
gene transfer (HGT) and in stabilizing mobile genetic elements (MGEs)
have been much debated. However, few studies have precisely addressed the
distribution of these systems in light of HGT, its mechanisms and its vectors. We
analyzed the distribution of R-M systems in 2261 prokaryote genomes and found
their frequency to be strongly dependent on the presence of MGEs, CRISPR-Cas
systems, integrons and natural transformation. Yet R-M systems are rare in
plasmids, in prophages and nearly absent from other phages. Their abundance
depends on genome size for small genomes where it relates with HGT but saturates
at two occurrences per genome. Chromosomal R-M systems might evolve under cycles
of purifying and relaxed selection, where sequence conservation depends on the
biochemical activity and complexity of the system and total gene loss is
frequent. Surprisingly, analysis of 43 pan-genomes suggests that solitary R-M
genes rarely arise from the degradation of R-M systems. Solitary genes are
transferred by large MGEs, whereas complete systems are more frequently
transferred autonomously or in small MGEs. Our results suggest means of testing
the roles for R-M systems and their associations with MGEs.

<>

<1>Oliynyk, M., Samborskyy, M., Lester, J.B., Mironenko, T., Scott, N., Dickens, S., Haydock, S.F., Leadlay, P.F.
<2>Complete genome sequence of the erythromycinproducing bacterium Saccharopolyspora erythraea NRRL23338.
<3>Nat. Biotechnol.
<4>25
<5>447-453
<6>2007
<7>Saccharopolyspora erythraea is used for the industrial-scale production of the antibiotic
erythromycin A, derivatives of which play a vital role in medicine. The sequenced chromosome
of this soil bacterium comprises 8,212,805 base pairs, predicted to encode 7,264 genes. It is
circular, like those of the pathogenic actinomycetes Mycobacterium tuberculosis and
Corynebacterium diphtheriae, but unlike the linear chromosomes of the model actinomycete
Streptomyces coelicolor A3(2) and the closely related Streptomyces avermitilis. The S.
erythraea genome contains at least 25 gene clusters for production of known or predicted
secondary metabolites, at least 72 genes predicted to confer resistance to a range of common
antibiotic classes and many sets of duplicated genes to support its saprophytic lifestyle. The
availability of the genome sequence of S. erythraea will improve insight into its biology and
facilitate rational development of strains to generate high-titer producers of clinically
important antibiotics.

<>

<1>Oller, A.R., Vanden Broek, W., Conrad, M., Topal, M.D.
<2>Ability of DNA and spermidine to affect the activity of restriction endonucleases from several bacterial species.
<3>Biochemistry
<4>30
<5>2543-2549
<6>1991
<7>Previous work has described the novel ability to modulate in vitro the activity
of restriction endonuclease NaeI from Nocardia aerocoligenes by using cleavable
DNA and spermidine [Conrad & Topal (1989) Proc. Natl. Acad. Sci. 86,
9707-9711].  In this paper we report the results of a study of 49 type II
restriction enzymes from a variety of bacterial species.  On the basis of the
rates of cleavage observed, we found that in addition to expected cleavable
sites a number of enzymes had slow and resistant cognate recognition sites.
Resistant sites were identified for BspMI, NaeI, and NarI; slow sites were
identified for HpaII, NaeI, and SacII.  Cleavage of these sites was found to be
signficantly enhanced by the addition of cleavable DNA or spermidine.  We
demonstrate that for BspMI, as for NaeI, activator DNAs increased Vmax without
altering Km, whereas for HpaII, NarI, and SacII activator DNAs decreased Km
without changing Vmax.  Comparison among the Kms for NaeI cleavage of several
different substrates demonstrated that distant DNA sequences can affect DNA
recognition by the activated enzyme.  Our observations extend DNA activation of
the Nocardia NaeI endonuclease to restriction endonucleases from Nocardia
argentinensis (NarI), Bacillus species M (BspMI), Haemophilus parainfluenza
(HpaII), and Streptomyces achromogenes (SacII).  In addition, activation has
now been found to affect slow as well as resistant recognition sites.

<>

<1>Olmos, A., Henriquez-Piskulich, P., Sanchez, C., Rojas-Herrera, M., Moreno-Pino, M., Gomez, M., Rodriguez, Da.S.R., Maracaja-Coutinho, V., Aldea, P., Trombert, A.N.
<2>Draft Genome of Chilean Honeybee (Apis mellifera) Gut Strain Lactobacillus kunkeei MP2.
<3>Genome Announcements
<4>2
<5>e01013-14
<6>2014
<7>Here, we report the first draft genome sequence of Lactobacillus kunkeei strain MP2, isolated
from a Chilean honeybee gut. The sequenced genome has a total size
of 1.58 Mb distributed into 44 contigs and 1,356 protein-coding sequences.

<>

<1>Olsen, D.B., Kotzorek, G., Eckstein, F.
<2>Investigation of the inhibitory role of phosphorothioate internucleotidic linkages on the catalytic activity of the restriction endonuclease EcoRV.
<3>Biochemistry
<4>29
<5>9546-9551
<6>1990
<7>The inhibitory effect of phosphorothioate residues, located within one strand
of double-stranded DNA, on the hydrolytic activity of the restriction
endonuclease EcoRV was investigated.  Specific incorporation of a
phosphorothioate group at the site of cleavage yielded the sequence
5'-GATsATC-3'.  This modified sequence was cleaved at a relative rate of 0.1
compared to the unmodified substrate.  Substrates 5'-GATsAsTC-3' and
5'-GsATsATC-3', both containing one additional phosphorothioate substitution,
were linearized at a rate of 0.04 relative to unmodified DNA.  However, under
the same conditions, fully dAMPS-substituted DNA was found to be virtually
resistant to the hydrolytic activity of EcoRV.  Further experiments showed that
double-stranded DNA fragments generated by PCR containing phosphorothioate
groups within both strands are potent inhibitors of EcoRV catalysis.  The
inhibition was independent of whether the inhibitor fragment contained an EcoRV
recognition site.  We concluded that substitution of the phosphate group at the
site of cleavage by a phosphorothioate residue decreases the rate of
EcoRV-catalyzed hydrolysis most significantly.  Substitution of other phosphate
groups within the recognition sequence plays a limited role in enzyme
inhibition.  The presence of multiple dNMPS residues at regions of the DNA
removed from the EcoRV recognition site may decrease the amount of enzyme
available for catalysis by nonspecific binding to EcoRV.

<>

<1>Olsen, D.B., Kotzorek, G., Sayers, J.R., Eckstein, F.
<2>Inhibition of the restriction endonuclease BanII using modified DNA substrates.
<3>J. Biol. Chem.
<4>265
<5>14389-14394
<6>1990
<7>The restriction endonuclease BanII catalyzes the cleavage of double-stranded
DNA and recognizes the degenerate sequence 5'-GPuGCPyC-3'.  The polylinker of
M13mp18 contains one such sequence, 5'-GAGCTC-3'.  The three other possible
sites recognized by the enzyme were prepared by site-directed mutagenesis.  The
substitution of phosphate groups by phosphorothioate residues at some positions
within the various recognition sites had relatively little effect on the rate
of cleavage of the DNA.  However, when the DNA contained a phosphorothioate
group at the site of cleavage the rate of linearization of the DNA was
decreased by a factor of 9.  Interestingly, DNA which contained an additional
phosphorothioate internucleotidic linkage immediately 3'-outside the
recognition site could not be linearized by the enzyme.  The results indicate
that an important contact between enzyme and substrate is perturbed by the
presence of the sulfur atom at this position.

<>

<1>Olsen, D.B., Sayers, J.R., Kotzorek, G., Eckstein, F.
<2>Interaction of restriction endonucleases with phosphorothioate-containing DNA.
<3>Nucleosides and Nucleotides
<4>10
<5>665-667
<6>1991
<7>The requirements for inhibition of cleavage of phosporothioate-containing DNA
by the restriction enzymes BanII and EcoRV with respect to number and position
of these groups was determined.

<>

<1>Olszewski, J., Wasserman, B.P.
<2>Effect of glutaraldehyde on the activity of some DNA restriction endonucleases.
<3>Appl. Biochem. Biotechnol.
<4>13
<5>29-35
<6>1986
<7>The effect of the bifunctional crosslinking reagent glutaraldehyde on the
activity of the restriction enzymes BamHI, HindIII, EcoRI, and Tth111I was
investigated.  The four enzymes exhibited differential sensitivity to
inactivation.  Tth111I was the most sensitive, with activity losses occurring
at levels of 0.0025% and above.  HindIII was the most stable of the four and
remained fully active at concentrations as high as 0.075%.  Addition of BSA to
incubation mixtures generally had a stabilizing effect.  Implications of these
results for the design of glutaraldehyde-based methods for the immobilization
of restriction endonucleases are discussed.

<>

<1>Omura, S., Ikeda, H., Ishikawa, J., Horikawa, H., Shiba, T., Sakaki, Y., Hattori, M.
<2>Actinomycetes polynucleotides.
<3>European Patent Office
<4>EP 1852508 A
<5>
<6>2007
<7>Novel polynucleotides derived from microorganisms belonging to actinomycetes and fragments
thereof, polypeptides encoded by the polynucleotides and fragments thereof, polynucleotide
arrays comprising the polynucleotides and fragments thereof, recording media in which the
nucleotide sequences of the polynucleotide and fragments thereof have been recorded which are
readable in a computer, and use of them.

<>

<1>Ong, K.S., Aw, Y.K., Gan, H.M., Yule, C.M., Lee, S.M.
<2>Draft Genome Sequences of Two Antimicrobial-Producing Burkholderia sp. Strains, MSh1 and MSh2, Isolated from Malaysian Tropical Peat Swamp Forest Soil.
<3>Genome Announcements
<4>2
<5>e01032-14
<6>2014
<7>We report the draft genome sequences of two antimicrobial-producing isolates, Burkholderia sp.
strains MSh1 and MSh2, which were isolated from tropical peat
swamp forest soil. Putative genes related to different antimicrobial production
have been annotated in both genome sequences.

<>

<1>Ong, S.Y., Pratap, C.B., Wan, X., Hou, S., Abdul, R.A.Y., Saito, J.A., Nath, G., Alam, M.
<2>Complete Genome Sequence of Salmonella enterica subsp. enterica Serovar Typhi P-stx-12.
<3>J. Bacteriol.
<4>194
<5>2115-2116
<6>2012
<7>We report here the complete genome sequence of Salmonella enterica subsp. enterica serovar
Typhi P-stx-12, a clinical isolate obtained from a typhoid
carrier in India.

<>

<1>Ong, S.Y., Pratap, C.B., Wan, X., Hou, S., Rahman, A.Y., Saito, J.A., Nath, G., Alam, M.
<2>The Genomic Blueprint of Salmonella enterica subspecies enterica serovar Typhi P-stx-12.
<3>Standards in Genomic Sciences
<4>7
<5>483-496
<6>2013
<7>Salmonella enterica subspecies enterica serovar Typhi is a rod-shaped, Gram-negative,
facultatively anaerobic bacterium. It belongs to the family
Enterobacteriaceae in the class Gammaproteobacteria, and has the capability of
residing in the human gallbladder by forming a biofilm and hence causing the
person to become a typhoid carrier. Here we present the complete genome of
Salmonella enterica subspecies enterica serotype Typhi strain P-stx-12, which was
isolated from a chronic carrier in Varanasi, India. The complete genome comprises
a 4,768,352 bp chromosome with a total of 98 RNA genes, 4,691 protein-coding
genes and a 181,431 bp plasmid. Genome analysis revealed that the organism is
closely related to Salmonella enterica serovar Typhi strain Ty2 and Salmonella
enterica serovar Typhi strain CT18, although their genome structure is slightly
different.

<>

<1>Ono, A., Matsuo, Y., Matsuda, A., Ueda, T.
<2>Nucleosides and nucleotides. CXIX. Inhibition of DNA-cytosine methylase HhaI by a self-complementary oligonucleotide containing 5-fluorocytosine.
<3>Biol. Pharm. Bull.
<4>16
<5>529-533
<6>1993
<7>A self-complementary decadeoxyribonucleotide, 5'd(GAAGFGCTTC)3', containing 5-fluorocytosine
(F) in substitution for cytosine at the methylation site of DNA-cytosine methylase HhaI
(mHhaI) has been synthesized M.HhaI was inhibited by the pre-incubation of the enzyme with
d(GAAGFGCTTC).

<>

<1>Ono, A., Ohtani, Y., Sato, M., Ueda, T.
<2>Oligodeoxynucleotides containing 7-deazaadenine: synthesis and recognition by restriction.
<3>Nucleic Acids Symp. Ser.
<4>12
<5>67-70
<6>1983
<7>Deoxydecanucleotides containing a recognition sequence of BglII and Sau3AI, and their
7-deazaadenine analogs were synthesized by the phosphotriester method.  The decanucleotides
containing 7-deazaadenine in place of adenine were partially or strongly resistant to the
hydrolysis by these restriction endonucleases.

<>

<1>Ono, A., Sato, M., Ohtani, Y., Ueda, T.
<2>Synthesis of deoxyoligonucleotides containing 7-deazaadenine: recognition and cleavage by restriction endonuclease BglII and Sau3AI.
<3>Nucleic Acids Res.
<4>12
<5>8939-8949
<6>1984
<7>Deoxydecanucleotides having a recognition sequence of BglII and Sau3AI, and
their 7-deazaadenine analogs were synthesized.  The decanucleotides containing
7-deazaadenine in place of adenine were partially resistant to the hydrolysis
by Sau3AI and strongly resistant to that by BglII.  A new hypothesis on the
mode of recognition and cleavage of specific nucleotide sequences by BglII,
recognizing one strand and cleaving the other strand, is presented.

<>

<1>Ono, A., Ueda, T.
<2>Synthesis of decadeoxyribonucleotides containing N6-methyladenine, N4-methylcytosine, and 5-methylcytosine: recognition and cleavage by restriction endonucleases (nucleosides and nucleotides part 74).
<3>Nucleic Acids Res.
<4>15
<5>219-232
<6>1987
<7>The naturally-occurring modified bases, N6-methyladenine, N4-methylcytosine,
and 5-methylcytosine were chemically introduced in place of the adenine or
cytosine in the decadeoxyribonucleotides containing recognition sequences of
BglII, Sau3AI, MboI and MflI.  The modified oligomers bind to the enzymes but
the rates of cleavage by the enzymes are variable.

<>

<1>Ono, A., Ueda, T.
<2>Minor-groove-modified oligonucleotides: synthesis of decadeoxynucleotides containing hypoxanthine, N2-methylguanine and 3-deazaadenine, and their interactions with restriction endonucleases BglII, Sau3AI, and MboI.
<3>Nucleic Acids Res.
<4>15
<5>3059-3072
<6>1987
<7>Decadeoxynucleotides containing hypoxanthine, N2-methylguanine, 3-deazaadenine in the
recognition sequences of restriction endonucleases BglII, Sau3AI, and MboI were synthesized.
These decanucleotides modified in the base moieties facing into the minor groove were strongly
resistant to hydrolysis by BglII and partially resistant to that of Sau3AI and MboI.  The
decadeoxynucleotide containing 3-deazaadenine in place of adenine was bound to BglII strongly,
whereas the nucleotides containing hypoxanthine and N2-methylguanine were bound less tightly.

<>

<1>Ono, C., Oda, N.
<2>Oligonucleotides and methods for detection of N.gonorrhoeae using.
<3>Japanese Patent Office
<4>JP 2005278443 A
<5>
<6>2005
<7>
<>

<1>Onodera, N.T., Ryu, J., Durbic, T., Nislow, C., Archibald, J.M., Rohde, J.R.
<2>Genome Sequence of Shigella flexneri Serotype 5a Strain M90T Sm.
<3>J. Bacteriol.
<4>194
<5>3022
<6>2012
<7>Bacteria of the genus Shigella are a major cause of death worldwide (L. von Seidlein et al.,
PLoS Med. 3:e353, 2006). We sequenced the genome of Shigella flexneri strain M90T Sm (serotype
5a) and compared it to the published genome sequence of S. flexneri strain 8401 (serotype 5b).

<>

<1>Onyango, M., Wang, Y., Nickel, O., Zhao, C., Zhang, X., Hartke, A., Hemberger, J., Cemic, F.
<2>First Genome Sequence of Potential Mycotoxin-Degrading Bacterium Devosia nanyangense DDB001.
<3>Genome Announcements
<4>2
<5>e00922-14
<6>2014
<7>Devosia sp. nov. DDB001, isolated from mycotoxin-contaminated soil, is a potential
mycotoxin-degrading alphaproteobacterium. To our knowledge, this is the
first draft genome announcement of a Devosia species.

<>

<1>Ooi, S.K., Qiu, C., Bernstein, E., Li, K., Jia, D., Yang, Z., Erdjument-Bromage, H., Tempst, P., Lin, S.P., Allis, C.D., Cheng, X., Bestor, T.H.
<2>DNMT3L connects unmethylated lysine 4 of histone H3 to de novo methylation of DNA.
<3>Nature
<4>448
<5>714-717
<6>2007
<7>Mammals use DNA methylation for the heritable silencing of retrotransposons and imprinted
genes and for the inactivation of the X
chromosome in females. The establishment of patterns of DNA methylation
during gametogenesis depends in part on DNMT3L, an enzymatically inactive
regulatory factor that is related in sequence to the DNA
methyltransferases DNMT3A and DNMT3B. The main proteins that interact in
vivo with the product of an epitope-tagged allele of the endogenous Dnmt3L
gene were identified by mass spectrometry as DNMT3A2, DNMT3B and the four
core histones. Peptide interaction assays showed that DNMT3L specifically
interacts with the extreme amino terminus of histone H3; this interaction
was strongly inhibited by methylation at lysine 4 of histone H3 but was
insensitive to modifications at other positions. Crystallographic studies
of human DNMT3L showed that the protein has a carboxy-terminal
methyltransferase-like domain and an N-terminal cysteine-rich domain.
Cocrystallization of DNMT3L with the tail of histone H3 revealed that the
tail bound to the cysteine-rich domain of DNMT3L, and substitution of key
residues in the binding site eliminated the H3 tail-DNMT3L interaction.
These data indicate that DNMT3L recognizes histone H3 tails that are
unmethylated at lysine 4 and induces de novo DNA methylation by
recruitment or activation of DNMT3A2.

<>

<1>Oosterkamp, M.J. et al.
<2>Genome sequences of Alicycliphilus denitrificans strains BC and K601T.
<3>J. Bacteriol.
<4>193
<5>5028-5029
<6>2011
<7>Alicycliphilus denitrificans strain BC and A. denitrificans strain K601(T) degrade cyclic
hydrocarbons. These strains have been isolated from a
mixture of wastewater treatment plant material and benzene polluted soil
and from a wastewater treatment plant, respectively, suggesting their role
in bioremediation of soil and water. Although the strains are
phylogenetically closely related, there are some clear physiological
differences. The hydrocarbon cyclohexanol, for example, can be degraded by
strain K601(T), but not by strain BC. Furthermore, both strains can use
nitrate and oxygen as an electron acceptor, but only strain BC can use
chlorate as electron acceptor. To better understand nitrate and chlorate
reduction mechanisms coupled to the oxidation of cyclic compounds, the
genomes of A. denitrificans strain BC and K601(T) were sequenced. Here, we
report the complete genome sequences of A. denitrificans strain BC and
K601(T).

<>

<1>Ordinario, D.D., Burke, A.M., Long, P., Jocson, J.-M., Wang, H., Dickson, M.N., Gorodetsky, A.A.
<2>Sequence Specific Detection of Restriction Enzymes at DNA-Modified Carbon Nanotube Field Effect Transistors.
<3>Anal. Chem.
<4>86
<5>8628-8633
<6>2014
<7>Protein-DNA interactions play a central role in many cellular processes, and their
misregulation has been implicated in a number of human diseases. Thus, there is a pressing
need for the development of analytical strategies for interrogating the binding of proteins to
DNA. Herein, we report the electrical monitoring of a prototypical DNA-binding protein, the
PvuII restriction enzyme, at microfluidic-encapsulated, DNA-modified carbon nanotube field
effect transistors. Our integrated platform enables the sensitive, sequence specific detection
of PvuII at concentrations as low as 0.5 pM in a volume of 0.025 mu L (corresponding to
similar to 7500 proteins). These figures of merit compare favorably to state of the art values
reported for alternative fluorescent and electrical assays. The overall detection strategy
represents a step toward the massively parallel electrical monitoring, identification, and
quantification of protein DNA interactions at arrayed nanoscale devices.

<>

<1>Ordogh, L., Hunyadkurti, J., Voros, A., Horvath, B., Szucs, A., Urban, E., Kereszt, A., Kondorosi, E., Nagy, I.
<2>Complete Genome Sequence of Propionibacterium avidum Strain 44067, Isolated from  a Human Skin Abscess.
<3>Genome Announcements
<4>1
<5>e00337-13
<6>2013
<7>Propionibacterium avidum is an anaerobic Gram-positive bacterium that forms part  of the
normal human cutaneous microbiota, colonizing moist areas such as the
vestibule of the nose, axilla, and perineum. Here we present the complete genome
sequence of P. avidum strain 44067, which was isolated from a carbuncle of the
trunk.

<>

<1>Ordonez, O.F., Lanzarotti, E., Kurth, D., Gorriti, M.F., Revale, S., Cortez, N., Vazquez, M.P., Farias, M.E., Turjanski, A.G.
<2>Draft Genome Sequence of the Polyextremophilic Exiguobacterium sp. Strain S17, Isolated from Hyperarsenic Lakes in the Argentinian Puna.
<3>Genome Announcements
<4>1
<5>e00480-13
<6>2013
<7>Exiguobacterium sp. strain S17 is a moderately halotolerant, arsenic-resistant bacterium that
was isolated from Laguna Socompa stromatolites in the Argentinian
Puna. The draft genome sequence suggests potent enzyme candidates that are
essential for survival under multiple environmental extreme conditions, such as
high levels of UV radiation, elevated salinity, and the presence of critical
arsenic concentrations.

<>

<1>Orduna, P., Cevallos, M.A., de Leon, S.P., Arvizu, A., Hernandez-Gonzalez, I.L., Mendoza-Hernandez, G., Lopez-Vidal, Y.
<2>Genomic and proteomic analyses of Mycobacterium bovis BCG Mexico 1931 reveal a diverse immunogenic repertoire against tuberculosis infection.
<3>BMC Genomics
<4>12
<5>493
<6>2011
<7>BACKGROUND: Studies of Mycobacterium bovis BCG strains used in different
countries and vaccination programs show clear variations in the genomes
and immune protective properties of BCG strains. The aim of this study was
to characterise the genomic and immune proteomic profile of the BCG 1931
strain used in Mexico. RESULTS: BCG Mexico 1931 has a circular chromosome
of 4,350,386 bp with a G+C content and numbers of genes and pseudogenes
similar to those of BCG Tokyo and BCG Pasteur. BCG Mexico 1931 lacks
Region of Difference 1 (RD1), RD2 and N-RD18 and one copy of IS6110,
indicating that BCG Mexico 1931 belongs to DU2 group IV within the BCG
vaccine genealogy. In addition, this strain contains three new RDs, which
are 53 (RDMex01), 655 (RDMex02) and 2,847 bp (REDMex03) long, and 55
single-nucleotide polymorphisms representing non-synonymous mutations
compared to BCG Pasteur and BCG Tokyo. In a comparative proteomic
analysis, the BCG Mexico 1931, Danish, Phipps and Tokyo strains showed
812, 794, 791 and 701 protein spots, respectively. The same analysis
showed that BCG Mexico 1931 shares 62% of its protein spots with the BCG
Danish strain, 61% with the BCG Phipps strain and only 48% with the BCG
Tokyo strain. Thirty-nine reactive spots were detected in BCG Mexico 1931
using sera from subjects with active tuberculosis infections and positive
tuberculin skin tests. CONCLUSIONS: BCG Mexico 1931 has a smaller genome
than the BCG Pasteur and BCG Tokyo strains. Two specific deletions in BCG
Mexico 1931 are described (RDMex02 and RDMex03). The loss of RDMex02
(fadD23) is associated with enhanced macrophage binding and RDMex03
contains genes that may be involved in regulatory pathways. We also
describe new antigenic proteins for the first time.

<>

<1>Ordway, J.M., Bedell, J.A., Citek, R.W., Nunberg, A.N., Jeddeloh, J.A.
<2>MethylIMapper: a method for high-throughput, multilocus bisulfite sequence analysis and reporting.
<3>Biotechniques
<4>39
<5>464-468
<6>2005
<7>Understanding the phenotypic contribution of epigenetic components is making DNA methylation
pattern analysis more important in higher eukaryotic genomes as well as human disease.
Bisulfite sequencing protocols report DNA methylation occupancy information as a positive
assay output that allows methylation patterns to be elucidated from particular developmental
or disease states.  Reported here is a new method for bisulfite sequencing project management,
data analysis, and site-specific methylation test development that is designed for integration
in high-throughput genomic and bioinformatics analyses.

<>

<1>Orekhov, A.V., Rebentish, B.A., Debabov, V.G.
<2>A new site-specific endonuclease from Streptomyces -- SgrII.
<3>Dokl. Akad. Nauk.
<4>263
<5>217-220
<6>1982
<7>At the present time 21 restriction enzymes have been isolated from various
species of Streptomyces.  According to the genetic data of T.A. Voeikova, the
same restriction-modification systems tested with the acid of actinophage Pg81,
have been detected in the strains S. griseus, Kr. 20 and Rcg 2, a recombinant
obtained in a cross of S. coelicolor A3/2 and S. griseus Kr. 15.  In this work
we describe the isolation and characterization of a new type of restriction
enzyme, called SgrII.  These enzymes have been subsequently renamed Sgr20I and
Scg2I.

<>

<1>Orekhov, A.V., Strokina, I.V., Foors, A.R.
<2>Restriction of shuttle Escherichia coli - Streptomyces plasmids in Streptomyces Lividans 66.
<3>Genetika
<4>25
<5>614-625
<6>1989
<7>The shuttle Escherichia coli - Streptomyces plasmids were used to transform S. lividans 66.
Plasmid DNAs isolated from this strain transform it 10-1000-fold more efficiently than DNAs
from E. coli. A rare transformant cured of the most restricted plasmid is a more efficient
recipient of plasmid DNA from E. coli and has the property of R+/-M+ mutant. Restriction in S.
lividans 66 correlates with the appearance in DNA from E. coli of the sites susceptible to
Scg2I restriction endonuclease. The latter was isolated earlier from recombinant strain Rcg2,
a hybrid between S. griseus Kr. 15 and S. coelicolor A3(2). Scg2I possesses the recognition
sequence CC(T/A)GG, like EcoRII, MvaI and Eco dcm methylase. The DNA resistant to Scg2I
cleavage retained this ability after in vitro modification by EcoRII methylase. So, the
resistance of DNA to Scg2I cleavage is not connected with methylation at 4th and 5th position
of second cytosine in the recognition sequence. Neither restriction of plasmid DNA in S.
lividans 66 is dependent on dcm modification in E. coli, though its dependence on dam
modification is not excluded. It is assumed that the restriction in S. lividans 66 is
specified by endonuclease analogous to Scg2I.

<>

<1>Orlowski, J., Boniecki, M., Bujnicki, J.M.
<2>I-Ssp6803I: the first homing endonuclease from the PD-(D/E)XK superfamily exhibits an unusual mode of DNA recognition.
<3>Bioinformatics
<4>23
<5>527-530
<6>2007
<7>Motivation: Restriction endonucleases (REases) and homing endonucleases (HEases) are
biotechnologically important enzymes. Nearly all
structurally characterized REases belong to the PD-(D/E) XK superfamily
of nucleases, while most HEases belong to an unrelated LAGLIDADG
superfamily. These two protein folds are typically associated with very
different modes of protein-DNA recognition, consistent with the
different mechanisms of action required to achieve high specificity.
REases recognize short DNA sequences using multiple contacts per base
pair, while HEases recognize very long sites using a few contacts per
base pair, thereby allowing for partial degeneracy of the target
sequence. Thus far, neither REases with the LAGLIDADG fold, nor HEases
with the PD-(D/E) XK fold, have been found.Results: Using protein fold
recognition, we have identified the first member of the PD-(D/E) XK
superfamily among homing endonucleases, a cyanobacterial enzyme
I-Ssp6803I. We present a model of the I-Ssp6803I-DNA complex based on
the structure of Type II restriction endonuclease R. BglI and predict
the active site and residues involved in specific DNA sequence
recognition by I-Ssp6803I. Our finding reveals a new unexpected
evolutionary link between HEases and REases and suggests how PD-(D/E)
XK nucleases may develop a 'HEase-like' way of interacting with the
extended DNA sequence. This in turn may be exploited to study the
evolution of DNA sequence specificity and to engineer nucleases with
new substrate specificities.Contact: iamb@genesilico.pl.

<>

<1>Orlowski, J., Bujnicki, J.M.
<2>Structural and evolutionary classification of Type II restriction enzymes based on theoretical and experimental analyses.
<3>Nucleic Acids Res.
<4>36
<5>3552-3569
<6>2008
<7>For a very long time, Type II restriction enzymes (REases) have been a paradigm of ORFans:
proteins with no detectable similarity to each other and to any other protein in the database,
despite common cellular and biochemical function. Crystallographic analyses published until
January 2008 provided high-resolution structures for only 28 of 1637 Type II REase sequences
available in the Restriction Enzyme database (REBASE). Among these structures, all but two
possess catalytic domains with the common PD-(D/E)XK nuclease fold. Two structures are
unrelated to the others: R.BfiI exhibits the phospholipase D (PLD) fold, while R.PabI has a
new fold termed 'half-pipe'. Thus far, bioinformatic studies supported by site-directed
mutagenesis have extended the number of tentatively assigned REase folds to five (now
including also GIY-YIG and HNH folds identified earlier in homing endonucleases) and provided
structural predictions for dozens of REase sequences without experimentally solved structures.
Here, we present a comprehensive study of all Type II REase sequences available in REBASE
together with their homologs detectable in the nonredundant and environmental samples
databases at the NCBI. We present the summary and critical evaluation of structural
assignments and predictions reported earlier, new classification of all REase sequences into
families, domain architecture analysis and new predictions of three-dimensional folds. Among
289 experimentally characterized (not putative) Type II REases, whose apparently full-length
sequences are available in REBASE, we assign 199 (69%) to contain the PD-(D/E)XK domain. The
HNH domain is the second most common, with 24 (8%) members. When putative REases are taken
into account, the fraction of PD-(D/E)XK and HNH folds changes to 48% and 30%, respectively.
Fifty-six characterized (and 521 predicted) REases remain unassigned to any of the five REase
folds identified so far, and may exhibit new architectures. These enzymes are proposed as the
most interesting targets for structure determination by high-resolution experimental methods.
Our analysis provides the first comprehensive map of sequence-structure relationships among
Type II REases and will help to focus the efforts of structural and functional genomics of
this large and biotechnologically important class of enzymes.

<>

<1>Orlowski, J., Mebrhatu, M.T., Michiels, C.W., Bujnicki, J.M., Aertsen, A.
<2>Mutational analysis and a structural model of methyl-directed restriction enzyme Mrr.
<3>Biochem. Biophys. Res. Commun.
<4>377
<5>862-866
<6>2008
<7>The Mrr protein of Escherichia coli K12 is a cryptic Type IV restriction endonuclease whose
activity appears to be triggered by high
pressure stress. In this report We used high pressure to isolate and
analyze several Mrr mutants, and generated a new structural model of
the Mrr protein. The activity of a number of spontaneous and
Strategically Constructed Mrr mutants is discussed in the light of this
model, providing a first insight into the Structure-function
relationships of the Mrr enzyme.

<>

<1>Ormeno-Orrillo, E., Menna, P., Almeida, L.G., Ollero, F.J., Nicolas, M.F., Pains, R., E, S.N., A, S.B., J, S., Oliveira, C., L, M., Souza, R.C., Ribeiro, V., A, T., Megias, M., Hungria, M., Martinez-Romero, E.
<2>Genomic basis of broad host range and environmental adaptability of Rhizobium tropici CIAT 899 and Rhizobium sp. PRF 81 which are used in inoculants for common bean (Phaseolus vulgaris L.).
<3>BMC Genomics
<4>13
<5>735
<6>2012
<7>ABSTRACT: BACKGROUND: Rhizobium tropici CIAT 899 and Rhizobium sp. PRF 81 are
alpha-Proteobacteria that establish nitrogen-fixing symbioses with a range of
legume hosts. These strains are broadly used in commercial inoculants for
application to common bean (Phaseolus vulgaris) in South America and Africa. Both
strains display intrinsic resistance to several abiotic stressful conditions such
as low soil pH and high temperatures, which are common in tropical environments,
and to several antimicrobials, including pesticides. The genetic determinants of
these interesting characteristics remain largely unknown. RESULTS: Genome
sequencing revealed that CIAT 899 and PRF 81 share a highly-conserved symbiotic
plasmid (pSym) that is present also in Rhizobium leucaenae CFN 299, a rhizobium
displaying a similar host range. This pSym seems to have arisen by a
co-integration event between two replicons. Remarkably, three distinct nodA genes
were found in the pSym, a characteristic that may contribute to the broad host
range of these rhizobia. Genes for biosynthesis and modulation of plant-hormone
levels were also identified in the pSym. Analysis of genes involved in stress
response showed that CIAT 899 and PRF 81 are well equipped to cope with low pH,
high temperatures and also with oxidative and osmotic stresses. Interestingly,
the genomes of CIAT 899 and PRF 81 had large numbers of genes encoding
drug-efflux systems, which may explain their high resistance to antimicrobials.
Genome analysis also revealed a wide array of traits that may allow these strains
to be successful rhizosphere colonizers, including surface polysaccharides,
uptake transporters and catabolic enzymes for nutrients, diverse iron-acquisition
systems, cell wall-degrading enzymes, type I and IV pili, and novel T1SS and T5SS
secreted adhesins. CONCLUSIONS: Availability of the complete genome sequences of
CIAT 899 and PRF 81 may be exploited in further efforts to understand the
interaction of tropical rhizobia with common bean and other legume hosts.

<>

<1>Ormeno-Orrillo, E., Rogel, M.A., Chueire, L.M., Tiedje, J.M., Martinez-Romero, E., Hungria, M.
<2>Genome Sequences of Burkholderia sp. Strains CCGE1002 and H160, Isolated from Legume Nodules in Mexico and Brazil.
<3>J. Bacteriol.
<4>194
<5>6927
<6>2012
<7>The genome sequences of Burkholderia sp. strains CCGE1002 from Mexico and H160 from Brazil,
isolated from legume nodules, are reported. Their gene contents in
relation to plant-microbe interactions and xenobiotic degradation are discussed.

<>

<1>Oroguchi, T., Hashimoto, H., Shimizu, T., Sato, M., Ikeguchi, M.
<2>Intrinsic Dynamics of Restriction Endonuclease EcoO109I Studied by Molecular Dynamics Simulations and X-Ray Scattering Data Analysis.
<3>Biophys. J.
<4>96
<5>2808-2822
<6>2009
<7>EcoO109I is a type II restriction endonuclease that functions as a dimer in solution. Upon DNA
binding to the enzyme, the two subunits
rotate counterclockwise relative to each other, as the two catalytic
domains undergo structural changes to capture the cognate DNA. Using a
150-ns molecular dynamics simulation, we investigated the intrinsic
dynamics of the DNA-free enzyme in solution to elucidate the
relationship between enzyme dynamics and structural changes. The
simulation revealed that the enzyme is considerably flexible, and thus
exhibits large fluctuations in the radius of gyration. The small-angle
x-ray scattering profile calculated from the simulation, including
scattering from explicit hydration water, was in agreement with the
experimentally observed profile. Principal component analysis revealed
that the major dynamics were represented by the open-close and
counterclockwise motions: the former is required for the enzyme to
access DNA, whereas the latter corresponds to structural changes upon
DNA binding. Furthermore, the intrinsic dynamics in the catalytic
domains were consistent with motions capturing the cognate DNA. These
results indicate that the structure of EcoO109I is intrinsically
flexible in the direction of its functional movement, to facilitate
effective structural changes for sequence-specific DNA recognition and
processing.

<>

<1>Orru, L., Salvetti, E., Cattivelli, L., Lamontanara, A., Michelotti, V., Capozzi, V., Spano, G., Keller, D., Cash, H., Martina, A., Torriani, S., Felis, G.E.
<2>Draft Genome Sequence of Bacillus coagulans GBI-30, 6086, a Widely Used Spore-Forming Probiotic Strain.
<3>Genome Announcements
<4>2
<5>e01080-14
<6>2014
<7>Bacillus coagulans GBI-30, 6086 is a safe strain, already available on the market, and
characterized by certified beneficial effects. The draft genome
sequence presented here constitutes the first pillar toward the identification of
the molecular mechanisms responsible for its positive features and safety.

<>

<1>Ortet, P., Barakat, M., Lalaouna, D., Fochesato, S., Barbe, V., Vacherie, B., Santaella, C., Heulin, T., Achouak, W.
<2>Complete Genome Sequence of a beneficial plant root-associated bacterium Pseudomonas brassicacearum.
<3>J. Bacteriol.
<4>193
<5>3146
<6>2011
<7>To shed light on the genetic equipment of the beneficial plant-associated bacterium
Pseudomonas brassicacearum, we sequenced the whole genome of the
strain NFM421. Its genome consists of one chromosome equipped with a
repertoire of plant growth beneficial factors. In addition a complete T3SS
and two complete T6SS were identified. We report here the first genome
sequence of this species.

<>

<1>Osama, A., Gan, H.M., Teh, C.S., Yap, K.P., Thong, K.L.
<2>Genome Sequence and Comparative Genomics Analysis of a Vibrio cholerae O1 Strain  Isolated from a Cholera Patient in Malaysia.
<3>J. Bacteriol.
<4>194
<5>6933
<6>2012
<7>The genome sequence analysis of a clinical Vibrio cholerae VC35 strain from an outbreak case
in Malaysia indicates multiple genes involved in host adaptation
and a novel Na(+)-driven multidrug efflux pump-coding gene in the genome of
Vibrio cholerae with the highest similarity to VMA_001754 of Vibrio mimicus
VMA223.

<>

<1>Oshima, H., Asai, K., Sadaie, Y.
<2>Restriction and modification system genes of Bacillus subtilis.
<3>Genes Genet. Syst.
<4>75
<5>380
<6>2000
<7>We found a set of genes for DNA restriction and modification in the third prophage region of
the Bacillus subtilis chromosome, where only five orfs are found in the 12kb region.  Two orfs
code for proteins similar to DNA methylation enzymes and constitute an operon.  The other
three orfs make another operon and the middle orf codes for a protein similar to DNA
restriction enzyme.  They must be RM system genes which recognize XhoI site and are deleted in
a classical RM strain of Bacillus subtilis.  XhoI sites on the DNA extracted only from RM
knockout strain could be digested with XhoI, indicating they are involved in restriction and
modification of Bacillus subtilis chromosome.

<>

<1>Oshima, T. et al.
<2>A 718-kb DNA sequence of the Escherichia coli K-12 genome corresponding to the 12.7-28.0 min region on the linkage map (supplement).
<3>DNA Res.
<4>3
<5>211-223
<6>1996
<7>none

<>

<1>Oshima, T. et al.
<2>A 718-kb DNA sequence of the Escherichia coli K-12 genome corresponding to the 12.7-28.0 min Region on the linkage map.
<3>DNA Res.
<4>3
<5>137-155
<6>1996
<7>The 718,122 base pair sequence of the Escherichia coli K-12 genome corresponding to the region
from 12.7 to 28.0 minutes on the genetic map is described.  This region contains at least 681
potential open reading frames, of which 277 (41%) have been previously identified, 147 (22%)
are homologous to other known genes, 139 (20%) are identical or similar to the hypothetical
genes registered in databases, and the remaining 118 (17%) do not show a significant
similarity to any other gene.  In this region, we assigned a cluster of cit genes encoding
multienzyme citrate lyase, two clusters of fimbrial genes and a set of lysogenic phage genes
encoding integrase, excisionase and repressor in the e14 genetic element.  In addition, a new
valine tRNA gene, designated valZ, and a family of long directly repeated sequences, LDR-A, -B
and -C, were found.

<>

<1>Oshima, T., Wada, C., Kawagoe, Y., Ara, T., Maeda, M., Masuda, Y., Hiraga, S., Mori, H.
<2>Genome-wide analysis of deoxyadenosine methyltransferase-mediated control of gene expression in Escherichia coli.
<3>Mol. Microbiol.
<4>45
<5>673-695
<6>2002
<7>Deoxyadenosine methyltransferase (Dam) methylates the deoxyadenine residues in 5'-GATC-3'
sequences and is important in many cellular processes in Escherichia coli. We performed a
computational analysis of the entire E. coli genome and confirmed that GATC sequences are
distributed unevenly in regulatory regions, which suggests that Dam might regulate gene
transcription. To test this, a high-density DNA microarray of 4097 E. coli genes was
constructed and used to assess the gene expression profiles of the wild type and the
dam-16::kam mutant strain grown under four different conditions. We also used two-dimensional
electrophoretic analysis of the proteome to assess the protein profiles. The expression of a
large number of genes was affected by the dam deficiency. Genes involved in aerobic
respiration, stress and SOS responses, amino acid metabolism and nucleotide metabolism were
expressed at higher levels in the mutant cells, especially in aerobic conditions. In contrast,
transcription of genes participating in anaerobic respiration, flagella biosynthesis,
chemotaxis and motility was decreased in the dam mutant strain under both aerobic and low
aerobic conditions. Thus, Dam-controlled genes are involved in adjusting the metabolic and
respiratory pathways and bacterial motility to suit particular environmental conditions. The
promoters of most of these Dam-controlled genes were also found to contain GATC sequences that
overlap with recognition sites for two global regulators, fumarate nitrate reduction (Fnr) and
catabolite activator protein (CRP). We propose that Dam-mediated methylation plays an
important role in the global regulation of genes, particularly those with Fnr and CRP binding
sites.

<>

<1>Osipiuk, J., Walsh, M.A., Joachimiak, A.
<2>Crystal structure of MboIIA methyltransferase.
<3>Nucleic Acids Res.
<4>31
<5>5440-5448
<6>2003
<7>DNA methyltransferases (MTases) are sequence-specific enzymes which transfer a methyl group
from S-adenosyl-L-methionine (AdoMet) to the amino
group of either cytosine or adenine within a recognized DNA sequence.
Methylation of a base in a specific DNA sequence protects DNA from
nucleolytic cleavage by restriction enzymes recognizing the same DNA
sequence. We have determined at 1.74 A resolution the crystal structure of
a beta-class DNA MTase MboIIA (M.MboIIA) from the bacterium Moraxella
bovis, the smallest DNA MTase determined to date. M.MboIIA methylates the
3' adenine of the pentanucleotide sequence 5'-GAAGA-3'. The protein
crystallizes with two molecules in the asymmetric unit which we propose to
resemble the dimer when M.MboIIA is not bound to DNA. The overall
structure of the enzyme closely resembles that of M.RsrI. However, the
cofactor-binding pocket in M.MboIIA forms a closed structure which is in
contrast to the open-form structures of other known MTases.

<>

<1>Oskam, L., Hillenga, D.J., Venema, G., Bron, S.
<2>The integrated state of the rolling-circle plasmid pTB913 in the composite Bacillus plasmid pTB19.
<3>Mol. Gen. Genet.
<4>233
<5>462-468
<6>1992
<7>pTB19, a 27 kb plasmid originating from a thermophilic Bacillus species,
contains integrated copies of two rolling-circle type plasmids on a 10.6
kb DNA fragment. In the present study we analysed the part of pTB19 that
contains the rolling-circle plasmid pTB913 and the region in between the
two rolling-circle plasmids. We show that, in the integrated state, pTB913
was flanked by a 55 bp direct repeat that duplicated part of the
replication initiation gene repB. Since repB was interrupted, the
integrated pTB913 could not initiate rolling-circle replication.
Autonomously replicating pTB913 was produced from pTB19, probably through
recombination between the 55 bp direct repeats; this was a rare event.
Since the second integrated rolling-circle type plasmid also contained a
non-functional replication initiation gene, replication of pTB19 must be
controlled by the RepA determinant. Theta-type replication, controlled by
RepA is likely to account for the high stability of pTB19. In between the
two integrated rolling-circle plasmids was present an open reading frame
(447 codons) which could encode a protein of unknown function.

<>

<1>Ossandon, F.J., Cardenas, J.P., Corbett, M., Quatrini, R., Holmes, D.S., Watkin, E.
<2>Draft Genome Sequence of the Iron-Oxidizing, Acidophilic, and Halotolerant 'Thiobacillus prosperus' Type Strain DSM 5130.
<3>Genome Announcements
<4>2
<5>e01042-14
<6>2014
<7>'Thiobacillus prosperus' is a halotolerant mesophilic acidophile that gains energy through
iron and sulfur oxidation. Its physiology is poorly understood.
Here, we describe the principal genomic features of the type strain of T.
prosperus, DSM 5130. This is the first public genome sequence of an acidophilic
halotolerant bacterium.

<>

<1>Ostendorf, T., Cherepanov, P., deVries, J., Wachernagel, W.
<2>Characterization of a dam mutant of Serratia marcescens and nucleotide sequence of the dam region.
<3>J. Bacteriol.
<4>181
<5>3880-3885
<6>1999
<7>The DNA of Serratia marcescens has N6-adenine methylation in GATC sequences. Among
2-aminopurine-sensitive mutants isolated from S. marcescens Sr41, one was identified which
lacked GATC methylation. The mutant showed up to 30-fold increased spontaneous mutability and
enhanced mutability after treatment with 2-aminopurine, ethyl methanesulfonate, or UV light.
The gene (dam) coding for the adenine methyltransferase (Dam enzyme) of S. marcescens was
identified on a gene bank plasmid which alleviated the 2-aminopurine sensitivity and the
higher mutability of a dam-13::Tn9 mutant of Escherichia coli. Nucleotide sequencing revealed
that the deduced amino acid sequence of Dam (270 amino acids; molecular mass, 31.3 kDa) has
72% identity to the Dam enzyme of E. coli. The dam gene is located between flanking genes
which are similar to those found to the sides of the E. coli dam gene. The results of
complementation studies indicated that like Dam of E. coli and unlike Dam of Vibrio cholerae,
the Dam enzyme of S. marcescens plays an important role in mutation avoidance by allowing the
mismatch repair enzymes to discriminate between the parental and newly synthesized strands
during correction of replication errors.

<>

<1>Osterlund, M., Luthman, H., Nilsson, S.V., Magnusson, G.
<2>Ethidium-bromide-inhibited restriction endonucleases cleave one strand of circular DNA.
<3>Gene
<4>20
<5>121-125
<6>1982
<7>We analyzed the effect of ethidium bromide (EtBr) on the cleavage of closed
circular pBR322 DNA molecules by six restriction enzymes which make staggered
or flush cuts (EcoRI, HindIII, BglI, PstI, HincII, PvuII).  EtBr concentrations
and reaction temperatures were determined at which DNA molecules with
single-strand breaks were the major reaction product of digestion by all the
enzymes.  However, the amounts of intermediates which could be isolated
differed for various enzymes.  The results extend previous studies, showing
that sequential cleavage of the DNA strands probably is a general property of
restriction endonucleases.

<>

<1>Osterman, D.G., DePillis, G.D., Wu, J.C., Matsuda, A., Santi, D.V.
<2>5-Fluorocytosine in DNA is a mechanism-based inhibitor of HhaI methylase.
<3>Biochemistry
<4>27
<5>5204-5210
<6>1988
<7>5-Fluorodeoxycytidine (FdCyd) was incorporated into a synthetic DNA polymer
containing the GCGC recognition sequence of HhaI methylase to give a polymer
with about 80% FdCyd.  In the absence of AdoMet, poly(FdC-dG) bound
competitively with respect to poly(dG-dC) (Ki=3nM).  In the presence of AdoMet,
the analogue caused a time-dependent, first-order (k=0.05 min-1) inactivation
of the enzyme.  There is an ordered mechanism of binding in which enzyme first
binds to poly(FdC-dG), then binds to AdoMet, and subsequently forms stable,
inactive complexes.  The complexes did not dissociate over the course of 3 days
and were stable to heat (95C) in the prsence of 1% SDS.  Gel filtration of a
complex formed with HhaI methylase, poly(FdC-dG), and [methyl-3H]AdoMet gave a
peak of radioactivity eluting near the void volume.  Digestion of the DNA in
the complex resulted in a reduction of the molecular weight to the size of the
methylase, and the radioactivity in this peak was shown to be associated with
protein.  These data indicate that the complexes contain covalently bound HhaI
methylase, poly(FdC-dG), and methyl groups and that 5-fluorodeoxycytidine is a
mechanism-based inactivator of the methylase.  By analogy with other
pyrimidine-modifying enzymes and recent studies on the mechanism of HhaI
methylase (Wu & Santi, 1987), these results suggest that an enzyme nucleophile
attacks FdCyd residues at C-6, activating the 5-position for one-carbon
transfer.  Subsequent transfer of the methyl group of AdoMet to the activated
FdCyd forms a stable complex in which the enzyme is covalently bound to the
6-position of FdCyd in the polymer and a methyl group is attached to C-5.  The
effect of 5-fluorodeoxycytidine on the inhibition of DNA-cytosine
methyltransferases is thus due to irreversible, covalent inactivation.

<>

<1>Osterman, J., Marsh, J., Laine, P., Zeng, Z., Alatalo, E., Sullivan, J.T., Young, J.P., Thomas-Oates, J., Paulin, L., Lindstrom, K.
<2>Genome sequencing of two Neorhizobium galegae strains reveals a noeT gene responsible for the unusual acetylation of the nodulation factors.
<3>BMC Genomics
<4>15
<5>500
<6>2014
<7>BACKGROUND: The species Neorhizobium galegae comprises two symbiovars that induce
nodules on Galega plants. Strains of both symbiovars, orientalis and officinalis,
induce nodules on the same plant species, but fix nitrogen only in their own host
species. The mechanism behind this strict host specificity is not yet known. In
this study, genome sequences of representatives of the two symbiovars were
produced, providing new material for studying properties of N. galegae, with a
special interest in genomic differences that may play a role in host specificity.
RESULTS: The genome sequences confirmed that the two representative strains are
much alike at a whole-genome level. Analysis of orthologous genes showed that N.
galegae has a higher number of orthologs shared with Rhizobium than with
Agrobacterium. The symbiosis plasmid of strain HAMBI 1141 was shown to transfer
by conjugation under optimal conditions. In addition, both sequenced strains have
an acetyltransferase gene which was shown to modify the Nod factor on the residue
adjacent to the non-reducing-terminal residue. The working hypothesis that this
gene is of major importance in directing host specificity of N. galegae could
not, however, be confirmed. CONCLUSIONS: Strains of N. galegae have many genes
differentiating them from strains of Agrobacterium, Rhizobium and Sinorhizobium.
However, the mechanism behind their ecological difference is not evident.
Although the final determinant for the strict host specificity of N. galegae
remains to be identified, the gene responsible for the species-specific
acetylation of the Nod factors was identified in this study. We propose the name
noeT for this gene to reflect its role in symbiosis.

<>

<1>Ostroff, G.R., Pene, J.J.
<2>Molecular cloning with bifunctional plasmid vectors in Bacillus subtilis:  Isolation of a spontaneous mutant of Bacillus subtilis with enhanced transformability for Escherichia coli-propagated chimeric plasmid DNA.
<3>J. Bacteriol.
<4>156
<5>934-936
<6>1983
<7>Hybrid plasmid DNA cloned in Escherichia coli undergoes deletions when returned
to competent Bacillus subtilis, even in defined restriction and modification
mutants of strain 168.  We have isolated a mutant of B. subtilis MI112 which is
stably transformed at high frequency by chimeric plasmid DNA propagated in E.
coli.

<>

<1>Ostroff, G.R., Pene, J.J.
<2>Molecular cloning with bifunctional plasmid vectors in Bacillus subtilis.  II.  Transfer of sequences propagated in Escherichia coli to B. subtilis.
<3>Mol. Gen. Genet.
<4>193
<5>306-311
<6>1984
<7>Recombinant plasmid DNA cloned in E. coli via the bifunctional vector pDH5060
suffered deletions when returned to B. subtilis.  However, DNA preparations of
identical chimeras containing homologous or heterologous sequences stably
transformed B. subtilis at high efficiency when isolated from B. subtilis.  The
vector pDH5060, however, was not affected and could be stably shuttled between
E. coli and B. subtilis at high frequency.  These problems affected the
transfer of clone pools and individual chimeras, irrespective of the
restriction or recombination phenotype of B. subtilis recipients.  Deleted
chimeras lost at least one end of cloned inserts, and in most cases, flanking
plasmid sequences.  Single plasmid forms (intact or deleted) were isolated from
several hundred individual Cmr-transformants.  This suggests that events
leading to deletion of chimeric plasmid DNA occur during transformation by
restriction of unmodified insert sequences propagated in the intermediate host,
E. coli.  This conclusion is discussed with regard to the mechanism of plasmid
transformation in B. subtilis.

<>

<1>Osuna, J., Flores, H., Soberon, X.
<2>Combinatorial mutagenesis of three major groove-contacting residues of EcoRI: single and double amino acid replacements retaining methyltransferase-sensitive activities.
<3>Gene
<4>106
<5>7-12
<6>1991
<7>A library of mutant ecoRIR genes encoding EcoRI restriction endonuclease was
generated using trinucleotide blocks and a combination of recombinant DNA
procedures, including primer extension and the polymerase chain reaction.
Codons corresponding to three amino acids (E144, R145 and R200), previously
implicated in the specific, recognition of the DNA substrate, were
combinatorially mutated so as to generate a library that potentially contains
all 8000 possible single, double and triple aa replacements, in a balanced
distribution.  Inspection of the phenotypes of Escherichia coli colonies
bearing the mutant genes showed that several of them retained activities that
were deleterious to the cells but were still protected by the EcoRI
methyltransferase.  These included new enzyme variants, including
non-conservative single (Thr or Val for Glu144) and double (Val for Glu144 and
Thr for Arg145) replacements.

<>

<1>Oswald, T., Hornbostel, G., Rinas, U., Anspach, F.B.
<2>Purification of (His)6EcoRV [recombinant restriction endonuclease EcoRV fused to a (His)6 affinity domain] by metal-chelate affinity chromatography.
<3>Biotechnol. Appl. Biochem.
<4>25
<5>109-115
<6>1997
<7>The chromatographic purification of (His)6EcoRV, a fusion protein consisting of a
hexahistidine affinity domain and restriction endonuclease EcoRV produced from recombinant
Escherichia coli, led to high product concentrations (>-1 mg/ml) in the preparative mode.
Increasing the amount of applied crude cell homogenate caused competition with host-specific
proteins, leading to a decrease of recovery and purity of the fusion protein.  Reduction of
host-specific proteins was achieved by pre-adsorption on to a DEAE anion-exchange sorbent.
This, in combination with 0.5-1 M NaCl in the adsorption buffer, assured a purity >95% and a
total protein recovery of ~34% in the preparative mode.  Contamination of the product with
about 2 mol of Ni(II)/mol of (His)6EcoRV was found due to metal-ion transfer to the N-terminal
high affinity binding site at (His)6.  Tris(carboxymethyl)ethylenediamine (TED)-Sepharose was
employed as an Ni(II) adsorber.  One passage of Ni(II)-contaminated protein solutions through
the TED-Sepharose column resulted in a decrease in the Ni(II) content in the (His)6EcoRV
fractions below the detection limit (~0.02 mg/l) of the atomic-adsorption spectrophotometer.

<>

<1>Oswald, T., Rinas, U.
<2>Chloramphenicol resistance interferes with purification of histidine-tagged fusion proteins from recombinant Escherichia coli.
<3>Anal. Biochem.
<4>236
<5>357-358
<6>1996
<7>The production of oligohistidine-tagged fusion proteins has become a widespread technique in
order to facilitate recombinant protein purification by using immobilized metal ion affinity
chromatography (IMAC).  In many cases a one-step purification procedure is sufficient to
obtain the protein of interest as 95% pure.  However, several host-specific proteins have been
identified which show a high affinity to the IMAC materials and are copurified with the
oligohistidine-tagged fusion protien.  Here we describe impaired purification of (His)6-tagged
fusion proteins by IMAC from chloramphenicol-resistant Escherichia coli cells.

<>

<1>Oswald, T., Wende, W., Pingoud, A., Rinas, U.
<2>Comparison of N-terminal affinity fusion domains: effect on expression level and product heterogeneity of recombinant restriction endonuclease EcoRV.
<3>Appl. Microbiol. Biotechnol.
<4>42
<5>73-77
<6>1994
<7>The influence of different N-terminal affinity fusion domains on the product heterogeneity of
recombinant proteins expressed in Escherichia coli was investigated. N-Terminal extended forms
of the restriction endonuclease EcoRV with either glutathione-S-transferase [GST], histidine
hexapeptide [(His)6], or a combination of GST and (His)6 [GST-(His)6] were compared to native
EcoRV with respect to expression level, susceptibility to inclusion body formation and protein
fragmentation. Fingerprinting of product heterogeneity was done by using two-dimensional (2-D)
non-equilibrium pH-gradient electrophoresis with subsequent immunoblotting. Fusion proteins
containing GST were poorly expressed compared to native EcoRV. In addition, GST fusion
proteins were highly susceptible to in vivo aggregation and fragmentation and displayed more
heterogeneity on 2-D immunoblots. However, the sole presence of oligohistidine at the
N-terminus of EcoRV proved to be advantageous. Fragmentation of (His)6-EcoRV was not observed
and 2-D immunoblots did not show heterogeneous forms of the recombinant protein. In addition,
fusion of the histidine-hexapeptide to the N-terminus of native EcoRV increased the expression
level of the recombinant protein twofold compared to native EcoRV. Inclusion body formation of
the (His)6-EcoRV fusion protein was intensive when cells were grown at 37oC but not at 30oC.
The advantage of oligohistidine fusion to EcoRV was finally demonstrated by purifying soluble
(His)6-EcoRV in a single-step procedure from crude cell lysates using immobilized metal
chelate affinity chromatography.

<>

<1>Ota, T., Isogai, T., Nishikawa, T., Hayashi, K., Saito, K., Yamamoto, J., Ishii, S., Sugiyama, T., Wakamatsu, A., Nagai, K., Otsuki, T.
<2>Primers for synthesising full-length cDNA and their use.
<3>European Patent Office
<4>EP 1074617 A
<5>
<6>2001
<7>
<>

<1>Otani, J., Nankumo, T., Arita, K., Inamoto, S., Ariyoshi, M., Shirakawa, M.
<2>Structural basis for recognition of H3K4 methylation status by the DNA methyltransferase 3A ATRX-DNMT3-DNMT3L domain.
<3>EMBO Rep.
<4>10
<5>1235-1241
<6>2009
<7>DNMT3 proteins are de novo DNA methyltransferases that are responsible for the establishment
of DNA methylation patterns in mammalian genomes. Here,
we have determined the crystal structures of the ATRX-DNMT3-DNMT3L (ADD)
domain of DNMT3A in an unliganded form and in a complex with the
amino-terminal tail of histone H3. Combined with the results of
biochemical analysis, the complex structure indicates that DNMT3A
recognizes the unmethylated state of lysine 4 in histone H3. This finding
indicates that the recruitment of DNMT3A onto chromatin, and thereby de
novo DNA methylation, is mediated by recognition of the histone
modification state by its ADD domain. Furthermore, our biochemical and
nuclear magnetic resonance data show mutually exclusive binding of the ADD
domain of DNMT3A and the chromodomain of heterochromatin protein 1alpha to
the H3 tail. These results indicate that de novo DNA methylation by DNMT3A
requires the alteration of chromatin structure.

<>

<1>Ott, J., Eckstein, F.
<2>31P NMR spectral analysis of the dodecamer d(CGCGAATTCGCG).
<3>Biochemistry
<4>24
<5>2530-2535
<6>1985
<7>The resonances in the 31P NMR spectrum of the dodecamer d(CGCGAATTCGCG) have
been assigned by use of regiospecific labeling with oxygen-17.  At 19C the
resonsances of 9 of the 11 dinucleoside phosphates are resolved.  Most
noticeably, different chemical shifts are observed for the phosphates of d(GpC)
at positions 2 and 10 as well as for d(CpG) at positions 1,3,9, and 11,
indicating that the position in an oligonucleotide influences the chemical
shift.  For the central d(GAATTC) portion of this dodecamer, a close
relationship between the chemical shift of the phosphate groups and their
position in the sequence of the oligonucleotide exists, in that the more
central the phosphate residue is the more the signal appears at higher field.
This finding parallels that found for the octamer d(GGAATTCC) [Connolly, B.A. &
Eckstein, F. (1984) Biochemistry 23,5523-5527].  The signals of the phosphate
residues at positions 3 and 9, however, are found at lower field strength than
expected from their position in the sequence, indicating a break in
conformation at these two locations.  A discontinuity of structure is also
observed at these positions in the X-ray structure of this dodecamer
[Dickerson, R.F., & Drew, H.R. (1981) J. Mol. Biol. 149, 761-786] as shown by
the anomalous twist angles between the third and fourth as well as the ninth
and tenth base pairs.  The dependence of the chemical shift on temperature
indicates different mobilities for each of the 11 phosphate groups.  There
seems to be no fraying at the ends but conformational changes particularly at
the central A-T base pairs at the center of the molecule, consistent with the
data obtained by H NMR spectroscopy [Patel, D.J., Kozlowski, S.A., Marky, L.A.,
Broka, C., Rice, J.A., Itakura, K., & Breslauer, K.J. (1982) Biochemistry 21,
428-436].

<>

<1>Ou, H.Y., He, X., Shao, Y., Tai, C., Rajakumar, K., Deng, Z.
<2>dndDB: a database focused on phosphorothioation of the DNA backbone.
<3>PLoS ONE
<4>4
<5>e5132
<6>2009
<7>BACKGROUND: The Dnd DNA degradation phenotype was first observed during electrophoresis of
genomic DNA from Streptomyces lividans more than 20
years ago. It was subsequently shown to be governed by the five-gene dnd
cluster. Similar gene clusters have now been found to be widespread among
many other distantly related bacteria. Recently the dnd cluster was shown
to mediate the incorporation of sulphur into the DNA backbone via a
sequence-selective, stereo-specific phosphorothioate modification in
Escherichia coli B7A. Intriguingly, to date all identified dnd clusters
lie within mobile genetic elements, the vast majority in laterally
transferred genomic islands. METHODOLOGY: We organized available data from
experimental and bioinformatics analyses about the DNA phosphorothioation
phenomenon and associated documentation as a dndDB database. It contains
the following detailed information: (i) Dnd phenotype; (ii) dnd gene
clusters; (iii) genomic islands harbouring dnd genes; (iv) Dnd proteins
and conserved domains. As of 25 December 2008, dndDB contained data
corresponding to 24 bacterial species exhibiting the Dnd phenotype
reported in the scientific literature. In addition, via in silico
analysis, dndDB identified 26 syntenic dnd clusters from 25 species of
Eubacteria and Archaea, 25 dnd-bearing genomic islands and one dnd plasmid
containing 114 dnd genes. A further 397 other genes coding for proteins
with varying levels of similarity to Dnd proteins were also included in
dndDB. A broad range of similarity search, sequence alignment and
phylogenetic tools are readily accessible to allow for to individualized
directions of research focused on dnd genes. CONCLUSION: dndDB can
facilitate efficient investigation of a wide range of aspects relating to
dnd DNA modification and other island-encoded functions in host organisms.
dndDB version 1.0 is freely available at http://mml.sjtu.edu.cn/dndDB/.

<>

<1>Oude Essink, B.B., Berkhout, B.
<2>The restriction enzyme BanI is inhibited by dcm-methylation of the GGCGCm5C site.
<3>Nucleic Acids Res.
<4>22
<5>108
<6>1994
<7>We constructed a synthetic tRNAlys,3 gene that was cloned into pUC9. A BanI site was
introduced at the 3' end to allow run-off transcription by T7 RNA polymerase. Surprisingly,
we were unable to digest this BanI site, while three BanI sites in the pUC9 plasmid were
efficiently cleaved (Figure 1, lane 7). Similar results were obtained upon prolonged
incubation at either 37 degrees C or 50 degrees C which is the optimal temperature for BanI
(results not shown). Since BanI recognizes different sequences (G/GPyPuCC), this could be due
to site-preference. However, the BanI site in the tRNA gene was identical to one of the pUC9
sites (GGCGCC). Inspection of the downstream sequences indicated the presence of an
overlapping dcm-methylation site (GGCGCCAGG). Because the plasmid was grown in a dcm+ host
(DH5), this will result in C5-methylation of the final C of the BanI site (GGCGCm5C). Next, we
grew pUC-tRNAlys3 in a dcm-host (GM48), which resulted in complete digestion of the plasmid
(lane 3). These results indicate that the BanI enzyme is not active on GGCGCm5C sites.

<>

<1>Ovechkina, L.G., Zinoviev, V.V., Gorbunov, Y.A., Malygin, E.G.
<2>The oligomerization of phage T4 DNA-(adenine-N-6)-methyltransferase and its effect on the catalytic characteristics of the enzyme.
<3>Bioorg. Khim.
<4>26
<5>940-943
<6>2000
<7>The structural and catalytic properties of the phage T4 DNA-(adenine-N-6)-methyltransferase
(EC 2.1;1.72) were studied at
different enzyme-substrate concentration ratios by chemical
cross-linking of the protein subunits and by measuring the presteady
state kinetics of the reactions. Various structural states of the
methyltransferase were correlated with its catalytic activity, and it
was shown that the oligomeric forms of the enzyme are catalytically
active but are characterized by the reaction parameters different from
those of the monomer.

<>

<1>Overholt, W.A., Green, S.J., Marks, K.P., Venkatraman, R., Prakash, O., Kostka, J.E.
<2>Draft genome sequences for oil-degrading bacterial strains from beach sands impacted by the deepwater horizon oil spill.
<3>Genome Announcements
<4>1
<5>e01015-13
<6>2013
<7>We report the draft genome sequences of 10 proteobacterial strains isolated from  beach sands
contaminated with crude oil discharged from the Deepwater Horizon
spill, which were cultivated under aerobic and anaerobic conditions with crude
oil as the sole carbon source. All strains contain multiple putative genes
belonging to hydrocarbon degradation pathways.

<>

<1>Ozdemir, O.
<2>The construction of a mammalian transfection vector for expression of cytosine-5 specific DNA methyltransferase gene M.MspI in cultured cells.
<3>Turkish J. Biol.
<4>22
<5>161-170
<6>1998
<7>The expression vectors are designed for expression and purification of normal or recombinant
genes of interest.  There is a wide variety of stable and transferable selectable mammalian
expression vectors.  The vector pCDM8 and its derivatives, pcDNAI/Ampicillin are widely used
for cloning and analyzing of genes in higher eukaryotic cells.  The vectors pRC/CMV, pcDNA3
and pRC/RSV are designed for high-level expression of recombinant genes in mammalian cells.
In the present study, we have been able to transfect and express the monospecific bacterial
(Moraxella sp.) cytosine-5 DNA methyltransferase M.MspI gene in cultured cells within a newly
constructed mammalian transfection vector.  We have constructed a very efficient, stable or
transferable eukaryotic expression vector analogous to the well-known expression system in COS
cells.  A bacterial cytosine-5 MTase gene with a vertebrate nuclear targeting signal SV40 VP1
and marker enzyme glutathione-S-transferase genes were transferred into the eukaryotic shuttle
vector pcDNA3 using a PCR based method.  This novel plasmid which contains a strong
cytomegalovirus and T7 promoters and the SV40 origin of replication for autonomous replication
in mammalian cells was called pOZT4.  Human kidney epithelial 293 and CHO cells were
transfected with pOZT4 which was encoding the fusion gene and it was established that the
genomic DNA of both cells were methylated at the CCCG sites by the active enzyme.

<>

<1>Ozdemir, O., Hornby, D.
<2>In vivo DNA methylation of Escherichia coli DH5a and top10F' strains by bacterial cytosine-5 methyltransferase M.MspI.
<3>Turkish J. Biol.
<4>22
<5>143-151
<6>1998
<7>At the chromatin level, methylated CpG dinucleotides are R.MspI resistant compared with
nonmethylated counterparts.  The DNA of two E. coli strains was analyzed following
transformation with bacterial cytosine-5-methyltransferase gene M.MspI in the mammalian
transfection vector pcDNA3.  Expression of the M.MspI was tested by R.MspI digestion.  The
results suggest that the DNA of both strains was fully methylated at the CCGG sequences, by
the active enzyme under the control of T7 and cytomegalovirus promoters.  Methylated DNA
cannot be digested and it exhibits higher fragment sizes in 1% agarose gel in contrast to the
untransformed cell DNAs in vivo.

<>

<1>Ozer, E.A., Allen, J.P., Hauser, A.R.
<2>Draft Genome Sequence of the Pseudomonas aeruginosa Bloodstream Isolate PABL056.
<3>J. Bacteriol.
<4>194
<5>5999
<6>2012
<7>Pseudomonas aeruginosa is an important cause of disease in hospitalized and immunocompromised
patients. The genome of P. aeruginosa is among the largest of
bacteria pathogenic to humans. We present the draft genome sequence of P.
aeruginosa strain PABL056, a human bloodstream isolate with the largest genome
yet reported in P. aeruginosa.

<>

<1>Ozer, E.A., Fitzpatrick, M.A., Hauser, A.R.
<2>Draft Genome Sequence of Acinetobacter baumannii Strain ABBL099, a Multidrug-Resistant Clinical Outbreak Isolate with a Novel Multilocus Sequence  Type.
<3>Genome Announcements
<4>2
<5>e00738-14
<6>2014
<7>Acinetobacter baumannii is associated with hospital-acquired infections and can cause
persistent outbreaks. Here we report the draft genome sequence of ABBL099,
a multidrug-resistant clinical isolate of A. baumannii belonging to a novel
sequence type and representative of clonal isolates cultured from patients at one
institution over a 4-year time period.

<>

<1>Pabo, C.O.
<2>Specificity by design - The specificity of a homing endonuclease has been altered using computational modeling of the protein-DNA interface.
<3>Nat. Biotechnol.
<4>24
<5>954-955
<6>2006
<7>Endonucleases that cleave DNA with high specificity have been exploited in biotechnology for
gene cloning and for nuclease-induced recombination.  As methods are developed that can
retarget such proteins to recognize any endogenous DNA site of interest, they should become
powerful tools that facilitate new approaches in gene therapy and genome editing.  In this
context, a recent Nature paper by David Baker and colleagues marks an important milestone.
The work builds on Barry Stoddard's long-standing interest in the study of homing
endonucleases and on recent studies from the Baker laboratory in which a computational model
of the protein-DNA interface was tested and optimized.  Using this model, Baker and colleagues
have now designed a variant of the I-MsoI homing endonuclease that binds and cleaves DNA at a
new site.  Their experiment had a relatively modest goal -- only one base pair was changed in
each recognition half-site -- but it nonetheless represents a critical conceptual and
methodological advance for the field.

<>

<1>Pachebat, J.A., van Keulen, G., Whitten, M.M., Girdwood, S., Del Sol, R., Dyson, P.J., Facey, P.D.
<2>Draft Genome Sequence of Rhodococcus rhodnii Strain LMG5362, a Symbiont of Rhodnius prolixus (Hemiptera, Reduviidae, Triatominae), the Principle Vector of  Trypanosoma cruzi.
<3>Genome Announcements
<4>1
<5>e00329-13
<6>2013
<7>We report the 4,385,577-bp high-quality draft assembly of the bacterial symbiont  Rhodococcus
rhodnii strain LMG5362, isolated from the gut of Rhodnius prolixus
(Hemiptera, Reduviidae, Triatominae), the principle vector of the protozoan
Trypanosoma cruzi, the etiological agent of Chagas disease. This sequence might
provide useful information for subsequent studies of the symbiotic relationship
between Rd. prolixus and Rc. rhodnii, while also providing a starting point for
the development of biotechnological applications for the control of Rd. prolixus.

<>

<1>Pachkunov, D.M., Kramarov, B.M., Dobritsa, A.P., Matvienko, N.I.
<2>Site-specific endonuclease BmeI from Bacillus Megaterium 216.
<3>Bioorg. Khim.
<4>9
<5>127-129
<6>1983
<7>A site-specific endonuclease BmeI has been isolated from Bacillus megaterium
216 by gel filtration on ultragel AcA-44 with a subsequent chromatography on
heparin-sepharose 6B.  On the double-stranded DNA the endonuclease recognizes
the pentanucleotide sequence 5' - GG (A) CC - 3' 3' - CC (T) GG - 5' and
hydrolyzes it in the points shown by arrows.  At gel filtration the
endonuclease is eluted in the volume corresponding to a molecular mass of
60,000.

<>

<1>Pack, S.P., Doi, A., Choi, Y.S., Kodaki, T., Makino, K.
<2>Biomolecular response of oxanine in DNA strands to T4 polynucleotide kinase, T4 DNA ligase, and restriction enzymes.
<3>Biochem. Biophys. Res. Commun.
<4>391
<5>118-122
<6>2010
<7>Oxanine (Oxa) generated from guanine (Gua) by NO- or HNO2-induced nitrosative oxidation, has
been thought to cause mutagenic problems in
cellular systems. In this study, the response of Oxa to different
enzymatic functions was explored to understand how similarly it can
participate in biomolecular reactions compared to the natural base,
Gua. The phosphorylation efficiency of the T4 polynucleotide kinase was
highest when Oxa was located on the 5'-end of single stranded DNAs
compared to when other nucleobases were in this position. The order of
phosphorylation efficiency was as follows, Oxa > Gua > adenine (Ade)
similar to thymine (Thy) > cylosine (Cyt) Base-pairing of Oxa and Cyt
(Oxa Cyt) between the ligation fragment and template was found to
influence the ligation performance of the T4 DNA ligase to a lesser
degree compared to Gua:Cyt. In addition, EcoRl and BglII showed higher
cleavage activities on DNA substrates containing Oxa:Cyt than those
containing Gua:Cyt, while BamHl, HindIII and EcoRV showed lower
cleavage activity; however, this decrease in activity was relatively
small.

<>

<1>Padegimiene, E., Maneliene, Z., Petrusyte, M., Janulaitis, A.
<2>OliI, a unique restriction endonuclease that recognizes the discontinuous sequence 5'-CACNN/NNGTG-3'.
<3>Nucleic Acids Res.
<4>29
<5>e30
<6>2001
<7>A new type II restriction endonuclease designated OliI has been partially purified from the
halophilic bacterium Oceanospirillum linum 4-5D. OliI recognizes the interrupted
hexanucleotide palindrome 5'-CACNNNNGTG-3' and cleaves it in the center generating
blunt-ended DNA fragments.

<>

<1>Padhy, R.N., Hottat, F.G., Coene, M.M., Hoet, P.P.
<2>Restriction analysis and quantitative estimation of methylated bases of filamentous and unicellular cyanobacterial DNAs.
<3>J. Bacteriol.
<4>170
<5>1934-1939
<6>1988
<7>The DNAs of strains of three cyanobacterial genera (Anabaena, Plectonema, and Synechococcus)
were found to be partially or fully resistant to many restriction endonucleases. This could be
due to the absence of specific sequences or to modifications, rendering given sequences
resistant to cleavage. The latter explanation is substantiated by the content of
N6-methyladenine and 5-methylcytosine in these genomes, which is high in comparison with that
in other bacterial genomes. dcm- and dam-like methylases are present in the three strains
(based on the restriction patterns obtained with the appropriate isoschizomeric enzymes).
Their contribution to the overall content of methyladenine and methylcytosine in the genomes
was calculated. Partial methylation of GATC sequences was observed in Anabaena DNA. In
addition, the GATC methylation patterns might not have been random in the three cyanobacterial
DNA preparations, as revealed by the appearance of discrete fragments (possibly of plasmid
origin) withstanding cleavage by DpnI (which requires the presence of methyladenine in the
GATC sequence).

<>

<1>Padmanabhan, R., Dubourg, G., Lagier, J.C., Couderc, C., Michelle, C., Raoult, D., Fournier, P.E.
<2>Genome sequence and description of Corynebacterium ihumii sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1128-1143
<6>2014
<7>Corynebacterium ihumii strain GD7(T) sp. nov. is proposed as the type strain of a new species,
which belongs to the family Corynebacteriaceae of the class
Actinobacteria. This strain was isolated from the fecal flora of a 62 year-old
male patient, as a part of the culturomics study. Corynebacterium ihumii is a
Gram positive, facultativly anaerobic, nonsporulating bacillus. Here, we describe
the features of this organism, together with the high quality draft genome
sequence, annotation and the comparison with other member of the genus
Corynebacteria. C. ihumii genome is 2,232,265 bp long (one chromosome but no
plasmid) containing 2,125 protein-coding and 53 RNA genes, including 4 rRNA
genes. The whole-genome shotgun sequence of Corynebacterium ihumii strain GD7(T)
sp. nov has been deposited in EMBL under accession number GCA_000403725.

<>

<1>Padmanabhan, R., Dubourg, G., Nguyen, T.T., Couderc, C., Rossi-Tamisier, M., Caputo, A., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Collinsella massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1144-1158
<6>2014
<7>Collinsella massiliensis strain GD3(T) is the type strain of Collinsella massiliensis sp.
nov., a new species within the genus Collinsella. This strain,
whose genome is described here, was isolated from the fecal flora of a
53-year-old French Caucasoid woman who had been admitted to intensive care unit
for Guillain-Barre syndrome. Collinsella massiliensis is a Gram-positive,
obligate anaerobic, non motile and non sporulating bacillus. Here, we describe
the features of this organism, together with the complete genome sequence and
annotation. The genome is 2,319,586 bp long (1 chromosome, no plasmid), exhibits
a G+C content of 65.8% and contains 2,003 protein-coding and 54 RNA genes,
including 1 rRNA operon.

<>

<1>Padmanabhan, R., Lagier, J.C., Dangui, N.P., Michelle, C., Couderc, C., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Megasphaera massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>8
<5>525-538
<6>2013
<7>Megasphaera massiliensis strain NP3(T) sp. nov. is the type strain of Megasphaera massiliensis
sp. nov., a new species within the genus Megasphaera. This strain,
whose genome is described here, was isolated from the fecal flora of an
HIV-infected patient. M. massiliensis is a Gram-negative, obligate anaerobic
coccobacillus. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 2,661,757 bp long genome (1
chromosome but no plasmid) contains 2,577 protein-coding and 61 RNA genes,
including 5 rRNA genes.

<>

<1>Padmanabhan, R., Robert, C., Fenollar, F., Raoult, D., Fournier, P.E.
<2>Draft Genome Sequence of Necropsobacter rosorum Strain P709T.
<3>Genome Announcements
<4>2
<5>e00913-14
<6>2014
<7>Necropsobacter is a recently described genus that contains a single species, N. rosorum, and
belongs to the family Pasteurellaceae. Here, we present the draft
genome of N. rosorum strain P709(T), which is the first genome sequence from this
species.

<>

<1>Pagani, I. et al.
<2>Complete genome sequence of Desulfobulbus propionicus type strain (1pr3).
<3>Standards in Genomic Sciences
<4>4
<5>100-110
<6>2011
<7>Desulfobulbus propionicus Widdel 1981 is the type species of the genus Desulfobulbus, which
belongs to the family Desulfobulbaceae. The species is of
interest because of its great implication in the sulfur cycle in aquatic
sediments, its large substrate spectrum and a broad versatility in using various
fermentation pathways. The species was the first example of a pure culture known
to disproportionate elemental sulfur to sulfate and sulfide. This is the first
completed genome sequence of a member of the genus Desulfobulbus and the third
published genome sequence from a member of the family Desulfobulbaceae. The
3,851,869 bp long genome with its 3,351 protein-coding and 57 RNA genes is a part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Pagani, I. et al.
<2>Complete genome sequence of Marivirga tractuosa type strain (H-43).
<3>Standards in Genomic Sciences
<4>4
<5>154-162
<6>2011
<7>Marivirga tractuosa (Lewin 1969) Nedashkovskaya et al. 2010 is the type species of the genus
Marivirga, which belongs to the family Flammeovirgaceae. Members of
this genus are of interest because of their gliding motility. The species is of
interest because representative strains show resistance to several antibiotics,
including gentamicin, kanamycin, neomycin, polymixin and streptomycin. This is
the first complete genome sequence of a member of the family Flammeovirgaceae.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. The 4,511,574 bp long chromosome and the 4,916 bp
plasmid with their 3,808 protein-coding and 49 RNA genes are a part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Pagie, L., Hogeweg, P.
<2>Individual- and population-based diversity in restriction-modification systems.
<3>Bull. Math. Biol.
<4>62
<5>759-774
<6>2000
<7>Restriction-modification (RM) systems are cognate gene complexes that code for an endonuclease
and a methylase. They are often thought to have developed in bacteria as protection against
invading genetic material, e.g., phage DNA. The high diversity of RM systems, as observed in
nature, is often ascribed to the coevolution of RM systems (which 'invent' novel types) and
phages. However, the extent to which phages are insensitive to RM systems casts doubts on the
effectiveness of RM systems as protection against infection and thereby on the reason for the
diversity of RM systems. We present an eco-evolutionary model in order to study the evolution
of the diversity of RM systems. The model predicts that in general diversity of RM systems is
high. More importantly, the diversity of the RM systems is expressed either at the individual
level or at the population level. In the first case all individuals carry RM systems of all
sequence specificities, whereas in the second case they carry only one RM system or no RM
systems at all. Nevertheless, in the second case the same number of sequence specificities are
present in the population.

<>

<1>Pagnier, I., Boughalmi, M., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Genome Sequence of Legionella tunisiensis Strain LegMT, a New Legionella Species  Isolated from Hypersaline Lake Water.
<3>J. Bacteriol.
<4>194
<5>5978
<6>2012
<7>Legionella tunisiensis is a gammaproteobacterium from the class Legionellaceae, growing in
amoebae. We sequenced the genome from strain LegM(T). It is composed
of 3,508,121 bp and contains 4,747 protein-coding genes and 38 RNA genes,
including 3 rRNA genes.

<>

<1>Pagnier, I., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Non-contiguous finished genome sequence and description of Anaerococcus provenciensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1198-1210
<6>2014
<7>Anaerococcus provenciensis strain 9402080(T) sp. nov. is the type strain of A. provenciensis
sp. nov., a new species within the genus Anaerococcus. This strain
was isolated from a cervical abscess sample. A. provenciensis is a Gram-positive
anaerobic cocci. Here, we describe the features of this organism, together with
the complete genome sequence and annotation. The 2.26 Mbp long genome contains
2099 protein-coding and 57 RNA genes including 8 rRNA genes and exhibits a G+C
content of 33.48%.

<>

<1>Pagnier, I., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Non-contiguous finished genome sequence and description of Fenollaria massiliensis gen. nov., sp. nov., a new genus of anaerobic bacterium.
<3>Standards in Genomic Sciences
<4>9
<5>704-717
<6>2014
<7>Fenollaria massiliensis strain 9401234(T), is the type strain of Fenollaria massiliensis gen.
nov., sp. nov., a new species within a new genus Fenollaria.
This strain, whose genome is described here, was isolated from an osteoarticular
sample. F. massiliensis strain 9401234(T) is an obligate anaerobic Gram-negative
bacillus. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 1.71 Mbp long genome exhibits a G+C
content of 34.46% and contains 1,667 protein-coding and 30 RNA genes, including 3
rRNA genes.

<>

<1>Pagnier, I., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Genome Sequence of Legionella massiliensis, Isolated from a Cooling Tower Water Sample.
<3>Genome Announcements
<4>2
<5>e01068-14
<6>2014
<7>We present the draft genome sequence of Legionella massiliensis strain LegA(T), recovered from
a cooling tower water sample, using an amoebal coculture
procedure. The strain described here is composed of 4,387,007 bp, with a G+C
content of 41.19%, and its genome has 3,767 protein-coding genes and 60 predicted
RNA genes.

<>

<1>Pagnier, I., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Genome Sequence of Reyranella massiliensis, a Bacterium Associated with Amoebae.
<3>J. Bacteriol.
<4>194
<5>5698
<6>2012
<7>Reyranella massiliensis is an Alphaproteobacterium member of the class Rhodospirillaceae,
growing in amoebae. We sequenced the genome of type strain
521(T). It is composed of a 5,792,218-bp chromosome and encodes 5,675
protein-coding genes and 53 RNA genes, including 3 rRNA genes.

<>

<1>Pagnier, I., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Genome Sequence of Afipia birgiae, a Rare Bacterium Associated with Amoebae.
<3>J. Bacteriol.
<4>194
<5>7018
<6>2012
<7>Afipia birgiae is an alphaproteobacterium from the family Bradyrhizobiaceae, growing in
amoebae, and a potential human pathogen. We sequenced the genome of
type strain 34632(T). It is composed of 5,325,467 bp and contains 5,160
protein-coding genes and 53 RNA genes, including 3 rRNA genes.

<>

<1>Pagnier, I., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Non-contiguous finished genome sequence and description of Anaerococcus pacaensis sp. nov., a new species of anaerobic bacterium.
<3>Standards in Genomic Sciences
<4>8
<5>548-560
<6>2013
<7>Anaerococcus pacaensis strain 9403502(T), is the type strain of Anaerococcus pacaensis sp.
nov., a new species within a new genus Anaerococcus. This strain,
whose genome is described here, was isolated from a blood sample. A. pacaensis
strain 9403502(T) is an obligate anaerobic Gram-positive coccus. Here we describe
the features of this organism, together with the complete genome sequence and
annotation. The 2.36 Mbp long genome exhibits a G+C content of 35.05% and
contains 2,186 protein-coding and 72 RNA genes, including 3 rRNA genes.

<>

<1>Pagnier, I., Croce, O., Robert, C., Raoult, D., La Scola, B.
<2>Genome Sequence of Legionella anisa, Isolated from a Respiratory Sample, Using an Amoebal Coculture Procedure.
<3>Genome Announcements
<4>2
<5>e00031-14
<6>2014
<7>Legionella anisa is a gammaproteobacterium from the class Legionellaceae, which is responsible
for nosocomial pneumonia. We sequenced the genome from the L.
anisa strain Linanisette, which was recovered from a clinical sample using an
amoebal coculture procedure but not with standard culture methods.

<>

<1>Pai, S.-H., Chuang, J.-Z., Kou, M.-C., Hsu, T.-T.
<2>Purification of EcoRI endonuclease on heparin sepharose-4B column chromatography.
<3>Proc. Natl. Sci. Counc. Repub. China B
<4>8
<5>41-45
<6>1984
<7>Restriction endonucleases play a very important role in genetic engineering and
DNA mapping.  Among hundreds of restriction endonucleases, the EcoRI enzyme is
the most useful and widely investigated enzyme.  After sonication and
ultracentrifugation, crude extracts of E. coli RY 13 were purified by employing
the polyethyleneimine precipitate, ammonium sulfate precipitate and heparin
Sepharose-4B affinity column chromatography.  The EcoRI enzyme were purified at
about 42 fold and the specific activity was about 100,000 U/mg of protein.  The
whole purification procedure was finished within two days.  The recovery was
about 42%.  The enzyme was sufficiently concentrated for direct specific DNA
hydrolysis.

<>

<1>Paigen, K., Weinfeld, H.
<2>Cooperative infection by host-modified lambda phage.
<3>Virology
<4>19
<5>565-572
<6>1963
<7>Host-modified lambda which arises by phage growth in Escherichia coli strain C
plates with an efficiency of 10-3 to 10-4 on E. coli strain K when infection is
performed at a low multiplicity.  The occasional infection which results arises
from the presence in K populations of rare cells in which lambda.C is capable
of multiplying.  When several lambda.C phage particles infect a single K cell,
a form of cooperative infection, analogous to multiplicity reactivation,
occurs, such that at high multiplicities more than 10% of the infected cells
yield progeny.  The latent period and burst size are normal when lambda.C
infects strain K either singly or multiply.  In both cases all the progeny that
are formed in a single cycle of growth are of the unrestricted or lambda.K
type.  The occurrence of cooperative infection is explained by assuming that
the restriction present in host modification is applied independently to
genetic structures smaller than the entire phage chromosome.  Each susceptible
site has a small chance of escaping restriction, and undamaged sites in
separate phage chromosomes can complement each other to produce an infection.
The quantitative dependence of the number of successful infections upon the
multiplicity of infection suggests that approximately 10 independent phage
sites are involved, each with approximately a 15% change of surviving.

<>

<1>Pajunen, M.I., Elizondo, M.R., Skurnik, M., Kieleczawa, J., Molineux, I.J.
<2>Complete nucleotide sequence and likely recombinatorial origin of bacteriophage T3.
<3>J. Mol. Biol.
<4>319
<5>1115-1132
<6>2002
<7>We report the complete genome sequence (38,208 bp) of bacteriophage T3 and
provide a bioinformatic comparative analysis with other completely
sequenced members of the T7 group of phages. This comparison suggests that
T3 has evolved from a recombinant between a T7-like coliphage and a
yersiniophage. To assess this, recombination between T7 and the Yersinia
enterocolitica serotype O:3 phage phiYeO3-12 was accomplished in vivo;
coliphage progeny from this cross were selected that had many biological
properties of T3. This represents the first experimentally observed
recombination between lytic phages whose normal hosts are different
bacterial genera.

<>

<1>Pak, C.M., Kim, U.Y., Ra, S.R.
<2>Study on recognization and cleavage characteristics of restriction enzyme Bci528I.
<3>Choson Minjujuui Inmin Konghwaguk Kwahagwon Tongbo
<4>98
<5>50-51
<6>2009
<7>We find that restriction enzyme Bci528I isolated Bacillus circulans 528 recognizes
pallindromic hexanucleotide sequence.

<>

<1>Pal, K.K., Dey, R., Sherathia, D., Dalsania, T., Savsani, K., Patel, I., Thomas, M., Ghorai, S., Vanpariya, S., Rupapara, R., Acharya, N., Rawal, P., Joshi, P., Sukhadiya, B., Mandaliya, M., Saxena, A.K.
<2>Draft Genome Sequence of Salinibacillus aidingensis Strain MSP4, an Obligate Halophilic Bacterium Isolated from a Salt Crystallizer of the Rann of Kutch,  India.
<3>Genome Announcements
<4>1
<5>e00253-13
<6>2013
<7>We report the 7.42-Mbp draft whole genome sequence of Salinibacillus aidingensis  strain MSP4,
an obligate halophilic bacterium, isolated from a salt crystallizer
of the Rann of Kutch in India. Analysis of the genome of this organism will lead
to a better understanding of the genes and metabolic pathways involved in
imparting osmotolerance.

<>

<1>Pal, K.K., Dey, R., Sherathia, D., Sukhadiya, B., Dalsania, T., Patel, I., Savsani, K., Thomas, M., Vanpariya, S., Mandaliya, M., Rupapara, R., Rawal, P., Ghorai, S., Bhayani, S., Shah, A., Saxena, A.K.
<2>Draft Genome Sequence of an Obligate and Moderately Halophilic Bacterium, Thalassobacillus devorans Strain MSP14, the First Draft Genome of the Genus  Thalassobacillus.
<3>Genome Announcements
<4>1
<5>e01103-13
<6>2013
<7>We report the 3.93-Mbp first draft genome sequence of a species of the genus Thalassobacillus,
Thalassobacillus devorans strain MSP14, a moderate but obligate
halophile, isolated from a salt crystallizer of the Little Rann of Kutch, India.
Exploring the genome of this organism will facilitate understanding the
mechanism(s) of its obligate halophilism.

<>

<1>Pal, K.K., Dey, R., Sherathia, D., Vanpariya, S., Patel, I., Dalsania, T., Savsani, K., Sukhadiya, B., Mandaliya, M., Thomas, M., Ghorai, S., Rupapara, R., Rawal, P., Shah, A., Bhayani, S.
<2>Draft Genome Sequence of a Moderately Halophilic Bacillus megaterium Strain, MSP20.1, Isolated from a Saltern of the Little Rann of Kutch, India.
<3>Genome Announcements
<4>2
<5>e01134-13
<6>2014
<7>The 4.37-Mbp draft genome of a moderately halophilic Bacillus megaterium strain,  MSP20.1,
isolated from a saltern of the Little Rann of Kutch, India, is reported
here. To understand the mechanism(s) of moderate halophilism and to isolate the
gene(s) involved in osmotolerance and adaptation, the genome of MSP20.1 was
sequenced.

<>

<1>Pal, K.K., Dey, R., Thomas, M., Sherathia, D., Dalsania, T., Patel, I., Savsani, K., Ghorai, S., Vanpariya, S., Sukhadiya, B., Mandaliya, M., Rupapara, R., Rawal, P.
<2>Draft Genome Sequence of the Extremely Halophilic Bacillus sp. Strain SB49, Isolated from a Salt Crystallizer Pond of the Little Rann of Kutch, India.
<3>Genome Announcements
<4>1
<5>e00869-13
<6>2013
<7>Here we report the draft whole-genome sequence (3.72 Mbp) of Bacillus sp. strain  SB49, an
extremely halophilic bacterium isolated from a salt crystallizer pond of
the Little Rann of Kutch in India. Unraveling the genome of this organism will
facilitate understanding and isolation of the genes involved in imparting extreme
osmotolerance.

<>

<1>Pal, K.K., Dey, R., Thomas, M., Sherathia, D., Dalsania, T., Patel, I., Savsani, K., Ghorai, S., Vanpariya, S., Sukhadiya, B., Mandaliya, M., Rupapara, R., Rawal, P., Saxena, A.K.
<2>Draft Genome Sequence of Bacillus sp. Strain SB47, an Obligate Extreme Halophile  Isolated from a Salt Pan of the Little Rann of Kutch, India.
<3>Genome Announcements
<4>1
<5>e00816-13
<6>2013
<7>Here, we report the 4.46-Mbp draft genome sequence of Bacillus sp. strain SB47, an extreme
halophile isolated from a salt pan of the Little Rann of Kutch, India.
Exploring the genome of this organism will facilitate the understanding and
isolation of the gene(s) involved in its extreme osmotolerance.

<>

<1>Palaniappan, K. et al.
<2>Genome sequence of the moderately thermophilic sulfur-reducing bacterium Thermanaerovibrio velox type strain (Z-9701(T)) and emended description of the  genus Thermanaerovibrio.
<3>Standards in Genomic Sciences
<4>9
<5>57-70
<6>2013
<7>Thermanaerovibrio velox Zavarzina et al. 2000 is a member of the Synergistaceae,  a family in
the phylum Synergistetes that is already well-characterized at the
genome level. Members of this phylum were described as Gram-negative staining
anaerobic bacteria with a rod/vibrioid cell shape and possessing an atypical
outer cell envelope. They inhabit a large variety of anaerobic environments
including soil, oil wells, wastewater treatment plants and animal
gastrointestinal tracts. They are also found to be linked to sites of human
diseases such as cysts, abscesses, and areas of periodontal disease. The
moderately thermophilic and organotrophic T. velox shares most of its morphologic
and physiologic features with the closely related species, T. acidaminovorans. In
addition to Su883(T), the type strain of T. acidaminovorans, stain Z-9701(T) is
the second type strain in the genus Thermanaerovibrio to have its genome sequence
published. Here we describe the features of this organism, together with the
non-contiguous genome sequence and annotation. The 1,880,838 bp long chromosome
(non-contiguous finished sequence) with its 1,751 protein-coding and 59 RNA genes
is a part of the G enomic E ncyclopedia of Bacteria and Archaea project.

<>

<1>Palecek, E., Boublikova, P., Galazka, G., Klysik, J.
<2>Inhibition of restriction endonuclease cleavage due to site-specific chemical modification of the B-Z junction in supercoiled DNA.
<3>Gen. Physiol. Biophys.
<4>6
<5>327-341
<6>1987
<7>Structural distortions on the boundary between right-handed B and left-handed Z
DNA segments in plasmid pRW751 (a derivative of pBR322 containing (dC-dG)13 and
(dC-dG)16 segments) were studied by means of chemical probes.  Samples of
supercoiled DNA were treated with the respective chemical probe, linearized
with EcoRI and inhibition of BamHI (whose recognition sequence GGATCC lies on
the boundary between the (dC-dG)n segments and the pBR322 nucleotide sequence)
cleavage was tested.  Treatment with osmium tetroxide in the presence of
pyridine or 2,2'-bipyridine, respectively, resulted in a strong inhibition of
the BamHI cleavage at both restriction sites, provided the (dC-dG)n segments
were in the left-handed form.  In the presence of 2,2'-bipyridine submillimolar
concentrations of OsO4 (at 26C) were sufficient to induce the inhibition of
BamHI.  Chloroacetaldehyde was used as a probe reacting selectively with atoms
involved in the Watson-Crick hydrogen bonding.  Similarly as in the case of
osmium tetroxide treatment of pRW751 with this agent resulted in the inhibition
of BamHI cleavage.  It was concluded that the B-Z junction regions in pRW751
contain few solitary bases with disturbed hydrogen bonding or non-Watson-Crick
base pairs.

<>

<1>Palenik, B., Brahamsha, B., Larimer, F.W., Land, M., Hauser, L., Chain, P., Lamerdin, J., Regala, W., Allen, E.E., McCarren, J., Paulsen, I., Dufresne, A., Partensky, F., Webb, E.A., Waterbury, J.
<2>The genome of a motile marine Synechococcus.
<3>Nature
<4>424
<5>1037
<6>2003
<7>Marine unicellular cyanobacteria are responsible for an estimated 20-40% of chlorophyll
biomass and carbon fixation in the oceans. Here we have
sequenced and analysed the 2.4-megabase genome of Synechococcus sp. strain
WH8102, revealing some of the ways that these organisms have adapted to
their largely oligotrophic environment. WH8102 uses organic nitrogen and
phosphorus sources and more sodium-dependent transporters than a model
freshwater cyanobacterium. Furthermore, it seems to have adopted
strategies for conserving limited iron stores by using nickel and cobalt
in some enzymes, has reduced its regulatory machinery (consistent with the
fact that the open ocean constitutes a far more constant and buffered
environment than fresh water), and has evolved a unique type of swimming
motility. The genome of WH8102 seems to have been greatly influenced by
horizontal gene transfer, partially through phages. The genetic material
contributed by horizontal gene transfer includes genes involved in the
modification of the cell surface and in swimming motility. On the basis of
its genome, WH8102 is more of a generalist than two related marine
cyanobacteria.

<>

<1>Palenik, B., Ren, Q., Dupont, C.L., Myers, G.S., Heidelberg, J.F., Badger, J.H., Madupu, R., Nelson, W.C., Brinkac, L.M., Dodson, R.J., Durkin, A.S., Daugherty, S.C., Sullivan, S.A., Khouri, H., Mohamoud, Y., Halpin, R., Paulsen, I.T.
<2>Genome sequence of Synechococcus CC9311: Insights into adaptation to a coastal environment.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>13555-13559
<6>2006
<7>Coastal aquatic environments are typically more highly productive and dynamic than open ocean
ones. Despite these differences, cyanobacteria
from the genus Synechococcus are important primary producers in both types
of ecosystems. We have found that the genome of a coastal cyanobacterium,
Synechococcus sp. strain CC9311, has significant differences from an open
ocean strain, Synechococcus sp. strain WH8102, and these are consistent
with the differences between their respective environments. CC9311 has a
greater capacity to sense and respond to changes in its (coastal)
environment. It has a much larger capacity to transport, store, use, or
export metals, especially iron and copper. In contrast, phosphate
acquisition seems less important, consistent with the higher concentration
of phosphate in coastal environments. CC9311 is predicted to have
differences in its outer membrane lipopolysaccharide, and this may be
characteristic of the speciation of some cyanobacterial groups. In
addition, the types of potentially horizontally transferred genes are
markedly different between the coastal and open ocean genomes and suggest
a more prominent role for phages in horizontal gene transfer in
oligotrophic environments.

<>

<1>Palma, L., Munoz, D., Murillo, J., Caballero, P.
<2>Draft Genome Sequence of Bacillus thuringiensis Serovar Tolworthi Strain Na205-3, an Isolate Toxic for Helicoverpa armigera.
<3>Genome Announcements
<4>2
<5>e00187-14
<6>2014
<7>We report here the complete annotated 6,510,053-bp draft genome sequence of Bacillus
thuringiensis serovar tolworthi strain Na205-3, which is toxic for
Helicoverpa armigera. This strain potentially contains nine insecticidal toxin
genes homologous to cry1Aa12, cry1Ab1, cry1Ab8, cry1Ba1, cry1Af1, cry1Ia10,
vip1Bb1, vip2Ba2, and vip3Aa6.

<>

<1>Palmer, B.R., Marinus, M.G.
<2>The dam and dcm strains of Escherichia coli--a review.
<3>Gene
<4>143
<5>1-12
<6>1994
<7>The construction of a variety of strains deficient in the methylation of adenine and cytosine
residues in DNA by the methyltransferases (MTases) Dam and Dcm has allowed the study of the
role of these enzymes in the biology of Escherichia coli. Dam methylation has been shown to
play a role in coordinating DNA replication initiation. DNA mismatch repair and the regulation
of expression of some genes. The regulation of expression of dam has been found to be complex
and influenced by five promoters. A role for Dcm methylation in the cell remains elusive and
dcm- cells have no obvious phenotype, dam- and dcm- strains have a range of uses in molecular
biology and bacterial genetics, including preparation of DNA for restriction by some
restriction endonucleases, for transformation into other bacterial species, nucleotide
sequencing and site-directed mutagenesis. A variety of assays are available for rapid
detection of both the Dam and Dcm phenotypes. A number of restriction systems in E. coli have
been described which recognise foreign DNA methylation, but ignore Dam and Dcm methylation.
Here, we describe the most commonly used mutant alleles of dam and dcm and the characteristics
of a variety of the strains that carry these genes. A description of several plasmids that
carry dam gene constructs is also included.

<>

<1>Palmer, K.L., Carniol, K., Manson, J.M., Heiman, D., Shea, T., Young, S., Zeng, Q., Gevers, D., Feldgarden, M., Birren, B., Gilmore, M.S.
<2>High-quality draft genome sequences of 28 Enterococcus sp. isolates.
<3>J. Bacteriol.
<4>192
<5>2469-2470
<6>2010
<7>The enterococci are low-GC Gram-positive bacteria that have emerged as leading causes of
hospital-acquired infection. They are also commensals of
the gastrointestinal tract of healthy humans and most other animals with
gastrointestinal flora and are important for food fermentations. Here we
report the availability of draft genome sequences for 28 enterococcal
strains of diverse origin, including the species Enterococcus faecalis, E.
faecium, E. casseliflavus, and E. gallinarum.

<>

<1>Palmer, M.E., Lipsitch, M., Moxon, E.R., Bayliss, C.D.
<2>Broad Conditions Favor the Evolution of Phase-Variable Loci.
<3>MBio
<4>4
<5>e00430-12
<6>2013
<7>Simple sequence repeat (SSR) tracts produce stochastic on-off switching, or phase variation,
in the expression of a panoply of
surface molecules in many bacterial commensals and pathogens. A change
to the number of repeats in a tract may alter the phase of the
translational reading frame, which toggles the on-off state of the
switch. Here, we construct an in silico SSR locus with mutational
dynamics calibrated to those of the Haemophilus influenzae mod locus.
We simulate its evolution in a regimen of two alternating environments,
simultaneously varying the selection coefficient, s, and the epoch
length, T. Some recent work in a simpler (two-locus) model suggested
that stochastic switching in a regimen of two alternating environments
may be evolutionarily favored only if the selection coefficients in the
two environments are nearly equal ('symmetric') or selection is very
strong. This finding was puzzling, as it greatly restricted the
conditions under which stochastic switching might evolve. Instead, we
find agreement with other recent theoretical work, observing selective
utility for stochastic switching if the product sT is large enough for
the favored state to nearly fix in both environments. Symmetry is
required neither in s nor in sT. Because we simulate finite populations
and use a detailed model of the SSR locus, we are also able to examine
the impact of population size and of several SSR locus parameters. Our
results indicate that conditions favoring evolution and maintenance of
SSR loci in bacteria are quite broad.
IMPORTANCE Bacteria experience frequent changes of environment
during the infection cycle. One means to rapidly adapt is stochastic
switching: a bacterial lineage will stochastically produce a variety of
genotypes, so that some descendants will survive if the environment
changes. Stochastic switching mediated by simple sequence repeat (SSR)
loci is widespread among bacterial commensals and pathogens and
influences critical interactions with host surfaces or immune
effectors, thereby affecting host persistence, transmission, and
virulence. Here, we use the most detailed in silico model of an SSR
locus to date, with its phase variation calibrated to match the mod
locus of Haemophilus influenzae. The type III restriction-modification
system encoded by mod participates in the regulation of multiple other
genes; thus, SSR-mediated phase variation of mod has far-reaching
cis-regulatory effects. This coupling of phase-variable switching to
complex phenotypic effects has been described as the 'phasevarion' and
is central to understanding the infection cycle of bacterial commensals
and pathogens.

<>

<1>Pan, X.S., Chen, Z.F.
<2>A new Type II restriction endonuclease, BsaOI, from Bacillus stearothermophilus.
<3>Chinese Sci. Bull.
<4>36
<5>1231-1232
<6>1991
<7>A new Type II restriction endonuclease, BsaOI, has been isolated from the thermophile Bacillus
stearothermophilus O-122. This enzyme cleaves pBR322 DNA at 7 sites, pUC19 DNA at 5 sites and
PhiX174 DNA at one site.

<>

<1>Pan, Y., Kong, K.F., Tsang, J.S.
<2>Draft Genome Sequence of the Haloacid-Degrading Burkholderia caribensis Strain MBA4.
<3>Genome Announcements
<4>2
<5>e00047-14
<6>2014
<7>Burkholderia caribensis MBA4 was isolated from soil for its ability to utilize 2-haloacid. An
inducible haloacid operon, encoding a dehalogenase and a permease,
is mainly responsible for the biotransformation. Here, we report the draft genome
sequence of this strain.

<>

<1>Pan, Y., Yang, X., Duan, J., Lu, N., Leung, A.S., Tran, V., Hu, Y., Wu, N., Liu, D., Wang, Z., Yu, X., Chen, C., Zhang, Y., Wan, K., Liu, J., Zhu, B.
<2>Whole-genome sequences of four Mycobacterium bovis BCG vaccine strains.
<3>J. Bacteriol.
<4>193
<5>3152-3153
<6>2011
<7>Bacille Calmette-Guerin (BCG) is the only vaccine available against tuberculosis (TB).
Currently there are a number of BCG strains that are in
use, which exhibit biochemical and genetic differences. We report the
genome sequences of four BCG strains representing different lineages,
which will help to design more effective TB vaccines.

<>

<1>Pan, Y., Yang, X., Li, J., Zhang, R., Hu, Y., Zhou, Y., Wang, J., Zhu, B.
<2>The genome sequence of spinosyns-producing bacterium Saccharopolyspora spinosa NRRL 18395.
<3>J. Bacteriol.
<4>193
<5>3150-3151
<6>2011
<7>Saccharopolyspora spinosa is a Gram-positive bacterium that produces spinosad, a well-known
biodegradable insecticide used for agricultural
pests control and has an excellent environmental and mammalian
toxicological profile. Here, we present the first draft genome sequence of
the type strain Saccharopolyspora spinosa NRRL 18395, which consists of 22
scaffolds.

<>

<1>Pan, Y.J., Lin, T.L., Lin, Y.T., Su, P.A., Chen, C.T., Hsieh, P.F., Hsu, C.R., Chen, C.C., Hsieh, Y.C., Wang, J.T.
<2>Identification of capsular types in carbapenem-resistant Klebsiella pneumoniae strains by wzc sequencing and implications in capsule depolymerase treatment.
<3>Antimicrob. Agents Chemother.
<4>0
<5>0
<6>2014
<7>Klebsiella pneumoniae is an important human pathogen associated with a variety of
diseases and the prevalence of multiple drug resistant K. pneumoniae (MDRKP) was
rapidly increasing. Here we determined the capsular types of 85
carbapenem-resistant K. pneumoniae (CRKP) by wzc sequencing and investigated the
presence of carbapenemases and integrons among CRKP. Ten strains (12%) of the
CRKP was positive for carbapenemase (IMP: 6/85, KPC: 3/85, and VIM: 1/85).
Capsular type K64 accounted for 32 (38%) of CRKP, followed by K62 (13%), K24
(8%), KN2 (7%) and K28 (6%). Sequence types (STs) were determined by multilocus
sequence typing (MLST) and the results indicated that ST11 which accounted for
47% (40/85) of these CRKP was the major ST. We further isolated a K64 specific
capsule depolymerase (K64dep) which can enhance serum and neutrophil killing in
vitro and increase survival rate in K64 K. pneumoniae inoculated mice. The
toxicity study demonstrated that mice treated with K64dep showed normal
biochemical parameters and no significant histopathological changes of liver,
kidney and spleen, indicating enzyme treatment did not cause toxicity in mice.
Therefore, the findings of capsular type clustering among CRKP and an effective
treatment of capsule depolymerase for MDRKP infections are important for
capsule-based vaccine development and therapy.

<>

<1>Panayotatos, N.
<2>Practical consequences of restriction site symmetry.
<3>Gene
<4>31
<5>291-294
<6>1984
<7>Because of the palindromic character of most 6-bp restriction sites, filling-in
and ligation of the protruding ends create symmetric sequences which include
new 6-bp restriction sites.  The old site is, in most cases, lost.  After
cleavage at the new palindromic site and removal of the protruding ends, a new
center of symmetry is created which is often part of yet another 6-bp
restriction site.  A compilation of potential and available sites as presented
should prove useful in genetic engineering.

<>

<1>Pandiyan, A., Ray, M.K.
<2>Draft Genome Sequence of the Antarctic Psychrophilic Bacterium Pseudomonas syringae Strain Lz4W.
<3>Genome Announcements
<4>1
<5>e00377-13
<6>2013
<7>The psychrophilic bacterium Pseudomonas syringae strain Lz4W was isolated from soil samples
from Antarctica to decipher the mechanisms of low-temperature
adaptation. We report here the 4.982-Mb draft genome sequence of P. syringae
Lz4W. This sequence will provide insights into the genomic basis of the
psychrophilicity of this bacterium.

<>

<1>Pang, J., Dong, M., Li, N., Zhao, Y., Liu, B.
<2>Functional characterization of a rice de novo DNA methyltransferase, OsDRM2, expressed in Escherichia coli and yeast.
<3>Biochem. Biophys. Res. Commun.
<4>432
<5>157-162
<6>2013
<7>DNA methylation of cytosine nucleotides is an important epigenetic modification that occurs in
most eukaryotic organisms and is established and maintained by various DNA methyltransferases
together with their co-factors. There are two major categories of DNA methyltransferases: de
novo and maintenance. Here, we report the isolation and functional characterization of a de
novo methyltransferase, named OsDRM2, from rice (Oryza sativa L.). The full-length coding
region of OsDRM2 was cloned and transformed into Escherichia coil and Saccharomyces
cerevisiae. Both of these organisms expressed the OsDRM2 protein, which exhibited stochastic
de novo methylation activity in vitro at CG, CHG, and CHH di- and tri-nucleotide patterns. Two
lines of evidence demonstrated the de novo activity of OsDRM2: (1) a 5'-CCGG-3' containing
DNA fragment that had been pre-treated with OsDRM2 protein expressed in E. coli was protected
from digestion by the CG-methylation-sensitive isoschizomer HpaII; (2) methylation-sensitive
amplified polymorphism (MSAP) analysis of S. cerevisiae genomic DNA from transformants that
had been introduced with OsDRM2 revealed CG and CHG methylation levels of 3.92-9.12%, and
2.88-6.93%, respectively, whereas the mock control S. cerevisiae DNA did not exhibit cytosine
methylation. These results were further supported by bisulfite sequencing of the 18S rRNA and
EAF5 genes of the transformed S. cerevisiae, which exhibited different DNA methylation
patterns, which were observed in the genomic DNA. Our findings establish that OsDRM2 is an
active de novo DNA methyltransferase gene with conserved activity in both prokaryotic and
eukaryotic non-host species. (C) 2013 Elsevier Inc. All rights reserved.

<>

<1>Pang, S., Renvoise, A., Perret, C., Guinier, M., Chelghoum, N., Brossier, F., Capton, E., Jarlier, V., Sougakoff, W.
<2>Whole-Genome Sequence of Mycobacterium abscessus Clinical Strain V06705.
<3>Genome Announcements
<4>1
<5>e00690-13
<6>2013
<7>Infection caused by Mycobacterium abscessus strains is a growing cause of concern in both
community-acquired and health care-associated diseases, as these
organisms naturally display multiple drug resistances. We report an annotated
draft genome sequence of M. abscessus strain V06705 obtained from a patient in
France.

<>

<1>Panina, E.M., Mironov, A.A., Gelfand, M.S.
<2>Statistical analysis of complete bacterial genomes: Avoidance of palindromes and restriction-modification systems.
<3>Mol. Biol. (Mosk)
<4>34
<5>246-252
<6>2000
<7>Absence of 4, 5, and 6-letter palindromes is observed in genome sequences of a broad spectrum
of bacteria. Recognition sites of
restrictases of a particular species (or of a species closely related
to it) comprise a significant portion of such palindromes. A
significant role is played by the horizontal transfer of genes that
encode the restriction-modification systems (R-M systems). In organisms
that are practically isolated from the effects of such systems (for
example, in Mycoplasma), such phenomenon is not observed. Common trends
to preference and "avoidance" of nucleotides were studied in
representatives of 33 bacterial families on the basis available
bacterial genomes. The results of the study present additional data on
intra- and interrelationships in established taxonomic groups.

<>

<1>Panis, G., Lambert, C., Viollier, P.H.
<2>Complete genome sequence of Caulobacter crescentus bacteriophage phiCbK.
<3>J. Virol.
<4>86
<5>10234-10235
<6>2012
<7>CbK is a B3 morphotype bacteriophage of the Siphoviridae family that infects Caulobacter
crescentus, the preeminent model system for bacterial cell cycle studies. The last 4 decades
of research with CbK as a genetic and cytological tool to study the biology of the host
warrant an investigation of the phage genome composition. Herein, we report the complete
genome sequence of CbK and highlight unusual features that emerged from its annotation. The
complete genome analysis of the CbK phage provides
new insight into its characteristics and potential interactions with its Caulobacter
crescentus host, setting the stage for future functional studies with CbK.

<>

<1>Panne, D., Muller, S.A., Wirtz, S., Engel, A., Bickle, T.A.
<2>The McrBC restriction endonuclease assembles into a ring structure in the presence of G nucleotides.
<3>EMBO J.
<4>20
<5>3210-3217
<6>2001
<7>McrBC from Escherichia coli K-12 is a restriction enzyme that belongs to the family of AAA(+)
proteins and cuts DNA containing modified cytosines. Two proteins are expressed from the mcrB
gene: a full-length version, McrB(L), and a short version, McrB(S). McrB(L) binds specifically
to the methylated recognition site and is, therefore, the DNA-binding moiety of the McrBC
endonuclease. McrB(S) is devoid of DNA-binding activity. We observed that the quaternary
structure of the endonuclease depends on binding of the cofactors. In gel filtration
experiments, McrB(L) and McrB(S) form high molecular weight oligomers in the presence of
Mg(2+) and GTP, GDP or GTP-gamma-S. Oligomerization did not require the presence of DNA and
was independent of GTP hydrolysis. Electron micrographs of negatively stained McrB(L) and
McrB(S) revealed ring-shaped particles with a central channel. Mass analysis by scanning
transmission electron microscopy indicates that McrB(L) and McrB(S) form single heptameric
rings as well as tetradecamers. In the presence of McrC, a subunit that is essential for DNA
cleavage, the tetradecameric species was the major form of the endonuclease.

<>

<1>Panne, D., Raleigh, E.A., Bickle, T.A.
<2>McrBS, a modulator peptide for McrBC activity.
<3>EMBO J.
<4>17
<5>5477-5483
<6>1998
<7>McrBC is a methylation-dependent endonuclease from Escherichia coli K-12.  The enzyme
recognizes DNA with modified cytosine preceded by a purine.  McrBC restricts DNA that contains
at least two methylated recognition sites separated by 40-80 bp.  Two gene products, McrBL and
McrBS, are produced from the mcrB gene and one, McrC, from the mcrC gene.  DNA cleavage in
vitro requires McrBL, McrC, GTP and Mg2+.  We found that DNA cleavage was optimal at a ratio
of 3-5 McrBL per molecule of McrC, suggesting that formation of a multisubunit complex with
several molecules of McrBL is required for cleavage.  To understand the role of McrBS, we have
purified the protein and analyzed its role in vitro.  At the optimal ratio of 3-5 McrBL per
molecule of McrC, McrBS acted as an inhibitor of DNA cleavage.  Inhibition was due to
sequestration of McrC and required the presence of GTP, suggesting that the interaction is GTP
dependent.  If McrC was in excess, a condition resulting in suboptimal DNA cleavage, addition
of McrBS enhanced DNA cleavage, presumably due to sequestration of excess McrC.  We suggest
that the role of McrBS is to modulate McrBC activity by binding to McrC.

<>

<1>Panne, D., Raleigh, E.A., Bickle, T.A.
<2>The McrBC endonuclease translocates DNA in a reaction dependent on GTP hydrolysis.
<3>J. Mol. Biol.
<4>290
<5>49-60
<6>1999
<7>McrBC specifically recognizes and cleaves methylated DNA in a reaction dependent on GTP
hydrolysis. DNA cleavage requires at least two recognition sites that are optimally separated
by 40-80 bp, but can be spaced as far as 3 kb apart. The nature of the communication between
two recognition sites was analyzed on DNA substrates containing one or two recognition sites.
DNA cleavage of circular DNA required only one methylated recognition site, whereas the
linearized form of this substrate was not cleaved. However, the linearized substrate was
cleaved if a Lac repressor was bound adjacent to the recognition site. These results suggest a
model in which communication between two remote sites is accomplished by DNA translocation
rather than looping. A mutant protein with defective GTPase activity cleaved substrates with
closely spaced recognition sites, but not substrates where the sites were further apart. This
indicates that McrBC translocates DNA in a reaction dependent on GTP hydrolysis. We suggest
that DNA cleavage occurs by the encounter of two DNA-translocating McrBC complexes, or can be
triggered by non-specific physical obstacles like the Lac repressor bound on the enzyme's
path along DNA. Our results indicate that McrBC belongs to the general class of DNA "motor
proteins", which use the free energy associated with nucleoside 5'-triphosphate hydrolysis to
translocate along DNA.

<>

<1>Papa, C.M., Springer, N.M., Muszynski, M.G., Meeley, R., Kaeppler, S.M.
<2>Maize chromomethylase Zea methyltransferase2 is required for CpNpG methyl ation.
<3>Plant Cell
<4>13
<5>1919-1928
<6>2001
<7>A cytosine DNA methyltransferase containing a chromodomain, Zea methyltransferase2 (Zmet2),
was cloned from maize. The sequence of ZMET2 is similar to that of the Arabidopsis
chromomethylases CMT1 and CMT3, with C-terminal motifs characteristic of eukaryotic and
prokaryotic DNA methyltransferases. We used a reverse genetics approach to determine the
function of the Zmet2 gene. Plants homozygous for a Mutator transposable element insertion
into motif IX had a 13% reduction in methylated cytosines. DNA gel blot analysis of these
plants with methylation-sensitive restriction enzymes and bisulfite sequencing of a 180-bp
knob sequence showed reduced methylation only at CpNpG sites. No reductions in methylation
were observed at CpG or asymmetric sites in heterozygous or homozygous mutant plants. Our
research shows that chromomethylase Zmet2 is required for in vivo methylation of CpNpG
sequences.

<>

<1>Papadakos, G.A.
<2>The effect of active site mutations on the homodimeric behavior of the PvuII restriction endonuclease.
<3>Ph.D. Thesis, University of Missouri
<4>
<5>
<6>2008
<7>The PvuII restriction endonuclease, a homodimer of two 18 kDa subunits, belongs to the type II
family of restriction enzymes. As a part of the Proteus vulgaris RM system, it specifically
cleaves the 5'-CAG|CTG-3' sequence in the presence of Mg2+ ions. Located in the active site
of PvuII, Tyrosine 94 has previously been shown to be involved in the metal ion binding by the
enzyme. The profile of the Ca2+ dependence of the DNA binding to the Y94F variant is shown to
be clearly biphasic. The application of
a sequential binding model yielded two weak binding constants in the upper phase with a
coupling energy (Go coop) at -0.3, while two tight binding constants are shown for the lower
phase with -1.4 kcal/mole interaction energy. The similar metal binding pattern between the
Y94F and the WT PvuII for Mg2+, Ca2+, Tb3+ and Eu3+ in the absence of DNA is also shown. The
application of 1H-15N HSQC spectroscopy in the presence of Ca2+ and DNA and the chemical
denaturation of the Y94F variant confirm the conformational impact of Tyr94. It is concluded
that the removal of the aromatic iii
hydroxyl group of Tyr94 slightly repositions the metal ions in the active site of PvuII
affecting the intra and/or inter-subunit interactions among the metal binding sites. The
single chain (SC) PvuII bearing a covalent linker between the two subunits is utilized in the
exploration of the modes of cooperativity among the metal binding sites. The heterodimeric
WT|E68A-SC PvuII was prepared and studied in parallel to the WTSC homodimer. Global analysis
of DNA binding isotherms at different Ca2+ concentrations for the WT|E68A-SC variant returned
an intra-subunit Go coop at +1.6 and +1.0 kcal/mole in the absence and presence of DNA,
respectively. Combined with
similar analysis for the WT-SC variant, the corresponding values for the inter-subunit Go
coop are shown at -2.8 and -1.1 kcal/mole for the occupation of two sites simultaneously. The
sequential binding of metal ions in the absence and presence of DNA is overall unfavorable
with significant negative interaction being observed between the metal sites. It is shown that
the effect of Ca2+ ions on DNA binding is greater than the effect of the DNA on the affinity
for metal ions. The cleavage of plasmid DNA under
single turnover conditions reveals a similar dependence of the nicking and linearization rates
on the concentration of Mg2+ ions for the WT-SC and the WT|E68A-SC PvuII. The series of events
leading to the linear product (DNA association, nicking, release of the intermediate,
re-association and linearization) in the presence of metal ions in one PvuII subunit is not
significantly slower than the synchronized double strand cleavage in the
presence of metal ions in both PvuII subunits.

<>

<1>Papadimitriou, K., Ferreira, S., Papandreou, N.C., Mavrogonatou, E., Supply, P., Pot, B., Tsakalidou, E.
<2>Complete Genome Sequence of the Dairy Isolate Streptococcus macedonicus ACA-DC 198.
<3>J. Bacteriol.
<4>194
<5>1838-1839
<6>2012
<7>The species Streptococcus macedonicus is associated with the food environment, especially with
fermented dairy products. Here we present the complete 2.1-Mb
genome sequence of strain ACA-DC 198, which was isolated from naturally fermented
Greek kasseri cheese.

<>

<1>Papagiannitsis, C.C., Miriagou, V., Giakkoupi, P., Tzouvelekis, L.S., Vatopoulos, A.C.
<2>Characterization of pKP1433, a Novel KPC-2-Encoding Plasmid from Klebsiella pneumoniae ST340.
<3>Antimicrob. Agents Chemother.
<4>57
<5>3427-3429
<6>2013
<7>The nucleotide sequence of pKP1433 (55417 bp), a blaKPC-2-carrying plasmid from
Klebsiella pneumoniae sequence type 340 was determined. pKP1433 displayed
extensive sequence and structural similarities with the IncN plasmids possessing
the KPC-2-encoding Tn4401b isoform. However, replication, partitioning, and
stability of pKP1433 were determined by sequences related to diverse non-IncN
plasmids.

<>

<1>Papagiannitsis, C.C., Tzouvelekis, L.S., Kotsakis, S.D., Tzelepi, E., Miriagou, V.
<2>Sequence of pR3521, an IncB Plasmid from Escherichia coli Encoding ACC-4, SCO-1, and TEM-1 {beta}-Lactamases.
<3>Antimicrob. Agents Chemother.
<4>55
<5>376-381
<6>2011
<7>The sequence of pR3521, a self-transmissible plasmid from Escherichia
coli, was determined. pR3521 (110,416 bp) comprised a contiguous IncB
sequence (84,034 bp) sharing extensive similarities with IncI replicons
and an acquired region (26,382 bp) carrying sequences of diverse origin,
containing bla(ACC-4), bla(SCO-1), bla(TEM-1b) (two copies), strA, strB,
sul2, and aacC2.

<>

<1>Papapanagiotou, I., Streeter, S.D., Cary, P.D., Kneale, G.G.
<2>DNA structural deformations in the interaction of the controller protein C.AhdI with its operator sequence.
<3>Nucleic Acids Res.
<4>35
<5>2643-2650
<6>2007
<7>Controller proteins such as C.AhdI regulate the expression of bacterial
restriction-modification genes, and ensure that methylation of the host DNA precedes
restriction by delaying transcription of the endonuclease. The operator DNA sequence to which
C.AhdI binds consists of two adjacent binding sites, O(L) and O(R). Binding of C.AhdI to O(L)
and to O(L) + O(R) has been investigated by circular permutation DNA-bending assays and by
circular dichroism (CD) spectroscopy. CD indicates considerable distortion to the DNA when
bound by C.AhdI. Binding to one or two sites to form dimeric and tetrameric complexes
increases the CD signal at 278 nm by 40 and 80% respectively, showing identical local
distortion at both sites. In contrast, DNA-bending assays gave similar bend angles for both
dimeric and tetrameric complexes (47 and 38 degrees , respectively). The relative orientation
of C.AhdI dimers in the tetrameric complex and the structural role of the conserved Py-A-T
sequences found at the centre of C-protein-binding sites are discussed.

<>

<1>Papke, R.T., de la Haba, R.R., Infante-Dominguez, C., Perez, D., Sanchez-Porro, C., Lapierre, P., Ventosa, A.
<2>Draft Genome Sequence of the Moderately Halophilic Bacterium Marinobacter lipolyticus Strain SM19.
<3>Genome Announcements
<4>1
<5>e00379-13
<6>2013
<7>Marinobacter lipolyticus strain SM19, isolated from saline soil in Spain, is a moderately
halophilic bacterium belonging to the class Gammaproteobacteria. Here,
we report the draft genome sequence of this strain, which consists of a 4.0-Mb
chromosome and which is able to produce the halophilic enzyme lipase LipBL.

<>

<1>Pappas, K.M., Kouvelis, V.N., Saunders, E., Brettin, T.S., Bruce, D., Detter, C., Balakireva, M., Han, C., Savvakis, G., Kyrpides, N.C., Typas, M.A.
<2>Genome sequence of the ethanol-producing Zymomonas mobilis subsp. mobilis lectotype strain ATCC 10988.
<3>J. Bacteriol.
<4>193
<5>5051-5052
<6>2011
<7>Zymomonas mobilis ATCC 10988 is the type strain of the Z. mobilis subsp. mobilis taxon,
members of which are some of the most rigorous
ethanol-producing bacteria. Isolated from Agave cactus fermentations in
Mexico, ATCC 10988 is of the first Z. mobilis strains to be described and
studied. Its robustness in sucrose-substrate fermentations, physiological
characteristics, large number of plasmids and overall genomic plasticity
render this strain important to the study of the species. Here we report
the finishing and annotation of the ATCC 10988 chromosomal and plasmid
genome.

<>

<1>Paques, F.
<2>Engineering of I-CreI homing endonuclease variants having novel cleavage specificity and use for genetic engineering, genome therapy and antiviral therapy.
<3>International Patent Office
<4>WO 200749156 A
<5>
<6>2007
<7>A method for engineering I-CreI homing endonuclease variants able to cleave mutant I-CreI
sites having variation in positions +/-8 to +/-10.  A I-CreI homing endonuclease variant
obtainable by said method,a vector encoding said variant, a cell, an animal or a plant
modified by said vector.  Use of said I-CreI endonuclease variant and derived products for
genetic engineering genome therapy and antiviral therapy.

<>

<1>Paques, F.
<2>Engineering of Laglidadg homing endonuclease variants having mutations in two functional subdomains, and use in genetic engineering and gene therapy.
<3>International Patent Office
<4>WO 200749095 A
<5>
<6>2007
<7>A LAGLIDADG homing endonuclease variant, having mutants in two separate subdomains, each
binding a distinct part of a modified DNA target half-site, said LAGLIDADG homing endonuclease
variant being able to cleave a chimeric DNA target sequence comprising the nucleotides bound
by each subdomain.  Use of said herodimeric meganuclease and derived products for genetic
engineering, genome therapy and antiviral therapy.

<>

<1>Paques, F.
<2>Engineering I-CreI homing endonuclease variants with modified cleavage specificity and use for genetic engineering and therapy.
<3>International Patent Office
<4>WO 200760495 A
<5>
<6>2007
<7>A method for engineering I-CreI homing endonuclease variants able to cleave mutant I-CreI
sites having variation in positions +/-8 to +/-10.  A I-CreI homing endonuclease variant
obtainable by said method, a vector encoding said variant, a cell, an animal or a plant
modified by said vector.  Use of said I-CreI endonuclease variant and derived products for
genetic engineering, genome therapy and antiviral therapy.

<>

<1>Paques, F.
<2>LAGLIDADG homing endonuclease variants having mutations in two functional subdomains and their use for cleaving defined target sequences.
<3>International Patent Office
<4>WO 200757781 A
<5>
<6>2007
<7>A LAGLIDADG homing endonuclease variant, having mutations in two separate subdomains, each
binding a distinct part of a modified DNA target half-site, said LAGLIDADG homing endonuclease
variant being able to cleave a chimeric DNA target sequence comprising the nucleotides bound
by each subdomain.  Use of said herodimeric meganuclease and derived products for genetic
engineering, genome therapy and antiviral therapy.

<>

<1>Paques, F., Duchateau, P.
<2>Meganucleases and DNA double-strand break-induced recombination: Perspectives for gene therapy.
<3>Curr. Gene Ther.
<4>7
<5>49-66
<6>2007
<7>Meganucleases are sequence-specific endonucleases recognizing large (> 12 bp) sequence sites
and several laboratories have used these proteins
to induce highly efficient gene targeting in mammalian cells. The
recent development of artificial endonucleases with tailored
specificities has opened the door for a wide range of new applications,
including therapeutic ones: redesigned endonucleases cleaving chosen
sequences could be used to in gene therapy to correct mutated genes or
introduce transgenes in chosen loci. Such "targeted" approaches
markedly differ from current gene therapy strategies based on the
random insertion of a complementing virus-borne transgene. As a
consequence, they should bypass the odds of random insertion.
Artificial fusion proteins including Zinc-Finger binding domains have
provided important proofs of concept, however the toxicity of these
proteins is still an issue. Today custom-designed homing endonucleases,
the natural meganucleases, could represent an efficient alternative.
After a brief description of the origin of the technology, current
systems based on redesigned endonucleases will be presented, with a
special emphasis on the recent advances in homing endonuclease
engineering. Finally, we will discuss the main issues that will need to
be addressed in order to bring this promising technology to the
patient.

<>

<1>Paquin, B., Laforest, M.-J., Lang, B.F.
<2>Interspecific transfer of mitochondrial genes in fungi and creation of a homologous hybrid gene.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>11807-11810
<6>1994
<7>In eukaryotes, horizontal gene transfer is a rare event.  Here we show that the mitochondrial
genome of a lower fungus, Allomyces macrogynus, has an extra DNA segment not present in a
close relative, Allomyces arbusculus.  This insert consists of the C terminus of a foreign
gene encoding a subunit of the ATP synthetase complex (atp6) plus an open reading frame
encoding an endonuclease.  The inserted atp6 portion is fused in phase to the resident gene,
resulting in expression of a hybrid atp6 gene and the displacement of the original C-terminal
atp6 region.  We present evidence that this insertion may have been acquired by interspecific
transfer and we discuss the possible role of the endonuclease in this process.

<>

<1>Paquin, B., Lang, B.F.
<2>The mitochondrial DNA of Allomyces macrogynus: the complete genomic sequence from an ancestral fungus.
<3>J. Mol. Biol.
<4>255
<5>688-701
<6>1996
<7>We have determined the complete nucleotide sequence of the circular mitochondrial DNA (mtDNA)
of the chytridiomycete fungus, Allomyces macrogynus (57,473 bp; A + T content 60.5%).  The
identified genes that are typical for most fungal mitochondria include those for the large
(rnl) and small subunit (rns) ribosomal RNAs, a complete set of 25 tRNAs, three ATPase
subunits (atp6, atp8 and atp9), apocytochrome b (cob), three subunits of the cytochrome
oxidase complex (cox1, cox2 and cox3), and seven subunits of the NADH dehydrogenase complex
(nad1, nad2, nad3, nad4, nad4L, nad5 and nad6).  A total of 28 introns of both groups are
found, some of which contain open reading frames (ORFs) coding for potential endonucleases
(group I) or reverse-transcriptases (group II).  All  mitochondrial genes are transcribed from
the same DNA strand, as is the case in many other eufungi.  Particular features of the A.
macrogynus mtDNA include: (1) the first documented case of a fungal mitochondrial ribosomal
protein gene (rps3) that is clearly identified by similarity with bacterial homologues; (2)
four unique ORFs; (3) the presence of an insert in the atp6 gene that may have been acquired
by interspecific transfer; (4) more than 67 short, highly structured and conserved DNA
elements inserted in intergenic spacers, introns, and variable regions of the rnl and rns
genes: these elements are unusually G + C rich; (5) rRNA structures that resemble more closely
those of eubacteria than their counterparts in other fungal mitochondria.  The high degree of
conservation of the A. macrogynus mitochondrial rRNA secondary structures, the existence of a
mitochondrial rps3 gene (common to protist but unique in fungal mtDNAs), and phylogenetic
relationships inferred from highly conserved protein genes, demonstrate consistently the
ancestral character of this fungal mitochondrial genome.

<>

<1>Paquin, B., O'Kelly, C.J., Lang, B.F.
<2>Intron-encoded open reaading frame of the GIY-YIG subclass in a plastid gene.
<3>Curr. Genet.
<4>28
<5>97-99
<6>1995
<7>Group-I introns, containing open reading frames that code for homing endonucleases, are widely
distributed amongst eukaryotic organellar genomes.  However, endonucleases of the GIY-YIG
subclass have a restricted distribution in mitochondria and bacteriophages, and have never
been observed in plastids.  We have found the GIY-YIG motif in an intronic ORF within the
previously published psbA gene sequence from Chlamydomonas reinhardtii chloroplasts.  Based on
phylogenetic analysis and an evaluation of amino-acid substitutions, this ORF is not closely
related to any of the other GIY-YIG ORFs.  These results suggest that GIY-YIG ORFs have a
longer evolutionary history than previously assumed.

<>

<1>Parashar, V., Capalash, N., Sharma, P.
<2>Demonstration of REBASE-assisted restriction mapping to determine the recognition site of unknown restriction endonucleases.
<3>Biochem. Mol. Biol. Educ.
<4>35
<5>337-341
<6>2007
<7>An important step in the characterization of a new restriction enzyme involves determination
of its recognition site. Comparison of its DNA
substrate digestion fragment patterns with those obtained using enzymes
of known specificity indicates whether the enzyme recognizes a novel
sequence or is an isoschizomer of already existing prototype. REBASE
(Restriction Enzyme dataBASE: hftp://www.neb.com/rebase)-assisted
restriction mapping is described in this paper for a rare cutter [TspMI
(REBASE No. 7191)] and a frequent cutter [Bfll (REBASE No. 4910)] as a
practical exercise for undergraduate students to understand how to
determine recognition sequence of a REase.

<>

<1>Parashar, V., Capalash, N., Xu, S.Y., Sako, Y., Sharma, P.
<2>TspMI, a thermostable isoschizomer of XmaI (5' C/CCGGG 3'): characterization and single molecule imaging with DNA - involving vector-mediated gene transfer and expression in host cell with restriction endonuclease activity.
<3>Appl. Microbiol. Biotechnol.
<4>72
<5>917-923
<6>2006
<7>AUTHOR ABSTRACT - TspMI, a thermostable isoschizomer of XmaI from a Thermus sp., has been
characterized. The enzyme was purified to homogeneity using Cibacron-Blue 3GA agarose, Heparin
agarose, SP sephadex C50, and Mono-Q fast protein liquid chromatography and was found to be a
homodimer of 40 kDa. Restriction mapping and run-off sequencing of TspMI-cleaved DNA ends
depicted that it cleaved at 5'C/CCGGG3' to generate a four-base, 5'-CCGG overhang. The
enzyme was sensitive to methylation of second and third cytosines in its recognition sequence.
TspMI worked optimally at 60 degrees C with 6 mM Mg2+, no Na+/K+, and showed no star activity
in the presence of 25% glycerol. The enzyme could efficiently digest the DNA labeled with a
higher concentration of YOYO-I (one dye molecule to one nucleotide), making it a useful
candidate for real-time imaging experiments. Single molecule interaction between TspMI and
lambda DNA was studied using total internal reflection fluorescence microscopy. The enzyme
survived 30 polymerase chain reaction (PCR) cycles in the presence of 10% glycerol and 0.5 M
trehalose without any activity loss and, hence, is suitable for incorporation in
restriction-endonuclease-mediated selective-PCR for various applications.

<>

<1>Pardo, C.E., Carr, I.M., Hoffman, C.J., Darst, R.P., Markham, A.F., Bonthron, D.T., Kladde, M.P.
<2>MethylViewer: computational analysis and editing for bisulfite sequencing and methyltransferase accessibility protocol for individual  templates (MAPit) projects.
<3>Nucleic Acids Res.
<4>39
<5>e5
<6>2011
<7>Bisulfite sequencing is a widely-used technique for examining cytosine DNA methylation at
nucleotide resolution along single DNA strands.
Probing with cytosine DNA methyltransferases followed by bisulfite
sequencing (MAPit) is an effective technique for mapping protein-DNA
interactions. Here, MAPit methylation footprinting with M.CviPI, a GC
methyltransferase we previously cloned and characterized, was used to
probe hMLH1 chromatin in HCT116 and RKO colorectal cancer cells.
Because M. CviPI-probed samples contain both CG and GC methylation, we
developed a versatile, visually-intuitive program, called MethylViewer,
for evaluating the bisulfite sequencing results. Uniquely, MethylViewer
can simultaneously query cytosine methylation status in
bisulfite-converted sequences at as many as four different user-defined
motifs, e.g. CG, GC, etc., including motifs with degenerate bases. Data
can also be exported for statistical analysis and as
publication-quality images. Analysis of hMLH1 MAPit data with
MethylViewer showed that endogenous CG methylation and accessible GC
sites were both mapped on single molecules at high resolution.
Disruption of positioned nucleosomes on single molecules of the PHO5
promoter was detected in budding yeast using M.CviPII, increasing the
number of enzymes available for probing protein-DNA interactions.
MethylViewer provides an integrated solution for primer design and
rapid, accurate and detailed analysis of bisulfite sequencing or MAPit
datasets from virtually any biological or biochemical system.

<>

<1>Parini, C., Fortina, M.G.
<2>Site-specific restriction endonucleases in Bacillus licheniformis.
<3>FEMS Microbiol. Lett.
<4>132
<5>285-289
<6>1995
<7>We systematically studied site-specific restriction endonucleases in Bacillus licheniformis
strains and detected endonuclease activity in 25 of 217 strains tested.  Three different
activities were obtained.  One of these activities detected in 21 strains was the most
representative within the species and produced a banding pattern, after digestion of lambda
DNA, identical to that seen with ClaI.  Two other strains isolated from soil samples from
China and USA were found to produce a DNA-cleaving enzyme with the same recognition sequence
as BsaI.  One producer strain, isolated from a Peruvian soil sample, possessed a mixture of
two isoschizomers, ClaI and BsaI.  Finally, one strain produced an endonuclease activity, not
previously described in B. licheniformis, that showed the same recognition sites as Bsu36I.

<>

<1>Parizzi, L.P., Grassi, M.C., Llerena, L.A., Carazzolle, M.F., Queiroz, V.L., Lunardi, I., Zeidler, A.F., Teixeira, P.J., Mieczkowski, P., Rincones, J., Pereira, G.A.
<2>The genome sequence of Propionibacterium acidipropionici provides insights into its biotechnological and industrial potential.
<3>BMC Genomics
<4>13
<5>562
<6>2012
<7>ABSTRACT: BACKGROUND: Synthetic biology allows the development of new biochemical
pathways for the production of chemicals from renewable sources. One major
challenge is the identification of suitable microorganisms to hold these pathways
with sufficient robustness and high yield. In this work we analyzed the genome of
the propionic acid producer Actinobacteria Propionibacterium acidipropionici
(ATCC 4875). RESULTS: The assembled P. acidipropionici genome has 3,656,170 base
pairs (bp) with 68.8% G + C content and a low-copy plasmid of 6,868 bp. We
identified 3,336 protein coding genes, approximately 1000 more than P.
freudenreichii and P. acnes, with an increase in the number of genes putatively
involved in maintenance of genome integrity, as well as the presence of an
invertase and genes putatively involved in carbon catabolite repression. In
addition, we made an experimental confirmation of the ability of P.
acidipropionici to fix CO2, but no phosphoenolpyruvate carboxylase coding gene
was found in the genome. Instead, we identified the pyruvate carboxylase gene and
confirmed the presence of the corresponding enzyme in proteome analysis as a
potential candidate for this activity. Similarly, the phosphate acetyltransferase
and acetate kinase genes, which are considered responsible for acetate formation,
were not present in the genome. In P. acidipropionici, a similar function seems
to be performed by an ADP forming acetate-CoA ligase gene and its corresponding
enzyme was confirmed in the proteome analysis. CONCLUSIONS: Our data shows that
P. acidipropionici has several of the desired features that are required to
become a platform for the production of chemical commodities: multiple pathways
for efficient feedstock utilization, ability to fix CO2, robustness, and
efficient production of propionic acid, a potential precursor for valuable
3-carbon compounds.

<>

<1>Park, C., Shin, H.H., Kwon, E.Y., Choi, S.M., Kim, S.H., Park, S.H., Choi, J.H., Yoo, J.H., Lee, D.G., Shin, W.S.
<2>Two variants of staphylococcal cassette chromosome mec type IVA in community-associated meticillin-resistant Staphylococcus aureus strains in South Korea.
<3>J. Med. Microbiol.
<4>58
<5>1314-1321
<6>2009
<7>Meticillin-resistant Staphylococcus aureus (MRSA) strains harbouring staphylococcal cassette
chromosome mec (SCCmec) type IVA are known to be more prevalent in South Korea than in other
countries. Variations in the SCCmec IVA structure have been identified, including in sequence
type (ST)
1 and ST72 strains. This study compared and investigated the genetic characteristics of two
subtypes common in South Korea. Type IVA SCCmec of
ST1 strains was characterized by type IV features with the linearized pUB110 at the junkyard
(J) 3 region. However, that of ST72 strains carried a variant class B mec complex, ccrA2, with
an identity of approximately 96 % and the linearized pUB110 at the J3 region. In SCCmec of
ST72 strains, the organization of the class B variant and the J3 region may be more similar to
that of type IA than to other types, but the ccr type and other J regions seemed to be derived
from type IV. These genetic characteristics showed that type IVA appears to result from the
dynamic genetic exchange and recombination of SCC DNA.

<>

<1>Park, C.K., Joshi, H.K., Agrawal, A., Ghare, M.I., Little, E.J., Dunten, P.W., Bitinaite, J., Horton, N.C.
<2>Domain swapping in allosteric modulation of DNA specificity.
<3>PLoS Biology
<4>8
<5>e1000554
<6>2010
<7>SgrAI is a type IIF restriction endonuclease that cuts an unusually long recognition sequence
and exhibits allosteric self-modulation of cleavage
activity and sequence specificity. Previous studies have shown that DNA
bound dimers of SgrAI oligomerize into an activated form with higher DNA
cleavage rates, although previously determined crystal structures of SgrAI
bound to DNA show only the DNA bound dimer. A new crystal structure of the
type II restriction endonuclease SgrAI bound to DNA and Ca(2+) is now
presented, which shows the close association of two DNA bound SgrAI
dimers. This tetrameric form is unlike those of the homologous enzymes
Cfr10I and NgoMIV and is formed by the swapping of the amino-terminal 24
amino acid residues. Two mutations predicted to destabilize the swapped
form of SgrAI, P27W and P27G, have been made and shown to eliminate both
the oligomerization of the DNA bound SgrAI dimers as well as the
allosteric stimulation of DNA cleavage by SgrAI. A mechanism involving
domain swapping is proposed to explain the unusual allosteric properties
of SgrAI via association of the domain swapped tetramer of SgrAI bound to
DNA into higher order oligomers.

<>

<1>Park, D.H., Thapa, S.P., Choi, B.S., Kim, W.S., Hur, J.H., Cho, J.M., Lim, J.S., Choi, I.Y., Lim, C.K.
<2>Complete Genome Sequence of Japanese Erwinia Strain Ejp617, a Bacterial Shoot Blight Pathogen of Pear.
<3>J. Bacteriol.
<4>193
<5>586-587
<6>2010
<7>The Japanese Erwinia strain Ejp617 is a plant pathogen that causes bacterial shoot blight of
pear (BSBP) in Japan. Here, we report the
complete genome sequence of strain Ejp617 isolated from Nashi pears in
Japan to provide for further valuable insight among related Erwinia
species.

<>

<1>Park, D.S., Bae, K.S., Kim, H., Shin, K.S., Choi, S.H., Kim, D.S., Kim, B.W., Oh, H.W.
<2>Draft Genome Sequence of the Novel Enteric Bacterium Galloisinimonas intestini B14T KCTC 32180, Isolated from the Gut of a Galloisiana Species (Notoptera:  Grylloblattidae) Fossil Insect.
<3>J. Bacteriol.
<4>194
<5>6648
<6>2012
<7>We report the 3.74-Mb genome sequence of Galloisinimonas intestini B14(T), isolated from the
gut of one of the world's rarest insect species, Galloisiana
sp., collected at a Mosan cave, Moonkyung, Gyungsangbook-do, South Korea. Strain
B14(T) is a novel genus candidate of the family Enterobacteriaceae.

<>

<1>Park, G.S., Hong, S.J., Lee, C.H., Khan, A.R., Ullah, I., Jung, B.K., Choi, J., Kwak, Y., Back, C.G., Jung, H.Y., Shin, J.H.
<2>Draft Genome Sequence of Chryseobacterium sp. Strain P1-3, a Keratinolytic Bacterium Isolated from Poultry Waste.
<3>Genome Announcements
<4>2
<5>e01237-14
<6>2014
<7>Chryseobacterium sp. strain P1-3, harboring keratin degrading activity, has recently been
isolated from poultry waste. Here, we report the 4.6-Mbp draft
genome sequence of the keratinolytic bacterium with a G+C content of 37.0% and
4,087 protein-coding genes.

<>

<1>Park, G.S., Khan, A.R., Hong, S.J., Jang, E.K., Ullah, I., Jung, B.K., Choi, J., Yoo, N.K., Park, K.J., Shin, J.H.
<2>Draft Genome Sequence of Entomopathogenic Bacterium Photorhabdus temperata Strain M1021, Isolated from Nematodes.
<3>Genome Announcements
<4>1
<5>e00747-13
<6>2013
<7>Photorhabdus temperata strain M1021 is an entomopathogenic bacterium belonging to the family
Enterobacteriaceae and is symbiotically associated with nematodes. The
draft genome sequence of P. temperata strain M1021 consists of 5,598,253 bp with
a G+C content of 43.7%, and it has 6,120 protein-coding genes.

<>

<1>Park, H.J., Kim, D.
<2>Draft Genome Sequence of a Humic Substance-Degrading Paenibacillus sp. Isolated from the Subarctic Grasslands at Low Temperature.
<3>Genome Announcements
<4>1
<5>e00170-12
<6>2013
<7>The Paenibacillus sp. strain PAMC 26794 was isolated from the tundra grasslands in Alaska for
its high ability to degrade humic acids. We sequenced the PAMC
26794 genome to discover the degradative genes for natural humic substances and
we propose the degradation pathway(s) of an abundant bacterial group (genus
Paenibacillus) that inhabits cold environments.

<>

<1>Park, H.J., Shin, S.C., Kim, D.
<2>Draft Genome Sequence of Arctic Marine Bacterium Pseudoalteromonas issachenkonii  PAMC 22718.
<3>J. Bacteriol.
<4>194
<5>4140
<6>2012
<7>The psychrotolerant Pseudoalteromonas issachenkonii PAMC 22718 was isolated for its higher
chitinase and protease activities from cold seawater in the Kara Sea,
Arctic. Here, we present the draft genome sequence of PAMC 22718 to provide
further information for the ecological function of the genus Pseudoalteromonas in
a cold marine environment.

<>

<1>Park, H.J., Shin, S.C., Kim, D.
<2>Draft genome sequence of a subarctic humic substance-degrading pseudomonad.
<3>Genome Announcements
<4>1
<5>e00070-12
<6>2013
<7>The Pseudomonas sp. PAMC 26793 was isolated because of its high ability to degrade humic acids
from a subarctic grassland in Alaska. We sequenced the PAMC 26793 genome to discover the genes
for degradation of natural humic substances and to provide further information for the
degradation process of soil bacteria in a low-temperature environment.

<>

<1>Park, J., Zhang, Y., Buboltz, A.M., Zhang, X., Schuster, S.C., Ahuja, U., Liu, M., Miller, J.F., Sebaihia, M., Bentley, S.D., Parkhill, J., Harvill, E.T.
<2>Comparative genomics of the classical Bordetella subspecies: the evolution and exchange of virulence-associated diversity amongst closely related pathogens.
<3>BMC Genomics
<4>13
<5>545
<6>2012
<7>ABSTRACT: BACKGROUND: The classical Bordetella subspecies are phylogenetically
closely related, yet differ in some of the most interesting and important
characteristics of pathogens, such as host range, virulence and persistence. The
compelling picture from previous comparisons of the three sequenced genomes was
of genome degradation, with substantial loss of genome content (up to 24%)
associated with adaptation to humans. RESULTS: For a more comprehensive picture
of lineage evolution, we employed comparative genomic and phylogenomic analyses
using seven additional diverse, newly sequenced Bordetella isolates. Genome-wide
single nucleotide polymorphism (SNP) analysis supports a reevaluation of the
phylogenetic relationships between the classical Bordetella subspecies, and
suggests a closer link between ovine and human B. parapertussis lineages than has
been previously proposed. Comparative analyses of genome content revealed that
only 50% of the pan-genome is conserved in all strains, reflecting substantial
diversity of genome content in these closely related pathogens that may relate to
their different host ranges, virulence and persistence characteristics.
Strikingly, these analyses suggest possible horizontal gene transfer (HGT) events
in multiple loci encoding virulence factors, including O-antigen and pertussis
toxin (Ptx). Segments of the pertussis toxin locus (ptx) and its secretion system
locus (ptl) appear to have been acquired by the classical Bordetella subspecies
and are divergent in different lineages, suggesting functional divergence in the
classical Bordetellae. CONCLUSIONS: Together, these observations, especially in
key virulence factors, reveal that multiple mechanisms, such as point mutations,
gain or loss of genes, as well as HGTs, contribute to the substantial phenotypic
diversity of these versatile subspecies in various hosts.

<>

<1>Park, J.H., Cho, Y.J., Chun, J., Seok, Y.J., Lee, J.K., Kim, K.S., Lee, K.H., Park, S.J., Choi, S.H.
<2>Complete Genome Sequence of Vibrio vulnificus MO6-24/O.
<3>J. Bacteriol.
<4>193
<5>2062-2063
<6>2011
<7>Vibrio vulnificus is the causative agent of life-threatening septicemia and severe wound
infections. Here, we announce the complete annotated
genome sequence of V. vulnificus MO6-24/O, isolated from a patient with
septicemia. When it is compared with previously known V. vulnificus
genomes, the genome of this bacterium shows a unique genetic makeup,
including phagelike elements, carbohydrate metabolism-related genes, and
the superintegron.

<>

<1>Park, J.Y., Han, S.H., Lee, J.H., Han, Y.S., Lee, Y.S., Rong, X., McSpadden, G.B.B., Park, H.S., Kim, Y.C.
<2>Draft Genome Sequence of the Biocontrol Bacterium Pseudomonas putida B001, an Oligotrophic Bacterium That Induces Systemic Resistance to Plant  Diseases.
<3>J. Bacteriol.
<4>193
<5>6795-6796
<6>2011
<7>Pseudomonas putida B001 is a rhizobacterium that was isolated on the basis of its abilities to
grow under low-nutrient conditions and induce systemic
resistance against bacterial, fungal, and viral diseases of plants. Here
we report the draft genome sequence and automatic annotation of strain
B001. Comparison of this sequence to the sequenced genome of P. putida
KT2440 points to a subset of gene functions that may be related to the
defense-inducing functions of B001.

<>

<1>Park, J.Y., Kim, S., Kim, S.M., Cha, S.H., Lim, S.K., Kim, J.
<2>Complete Genome Sequence of Multidrug-Resistant Acinetobacter baumannii Strain 1656-2, Which Forms Sturdy Biofilm.
<3>J. Bacteriol.
<4>193
<5>6393-6394
<6>2011
<7>Acinetobacter baumannii is a Gram-negative bacterium causing nosocomial infections worldwide.
To gain quick insight into the molecular basis of
biofilm formation in A. baumannii, we determined the complete genome
sequence of A. baumannii strain 1656-2, which forms sturdy biofilm and is
resistant to multiple drugs.

<>

<1>Park, M.A., Kwon, M.G., Hwang, J.Y., Jung, S.H., Kim, D.W., Park, J.Y., Kim, J.S., Na, Y.J., Kim, M.Y., Kim, D.S., Chae, S.H., Seo, J.S.
<2>Genome Sequence of Streptococcus parauberis Strain KCTC11980, Isolated from Diseased Paralichthys olivaceus.
<3>Genome Announcements
<4>1
<5>e00780-13
<6>2013
<7>Streptococcus parauberis is a coccoid, nonmotile, alpha-hemolytic, Gram-positive  bacterium of
the Streptococcaceae family. Streptococcus parauberis strain
KCTC11980 was isolated from the kidney of a diseased olive flounder collected
from an aquaculture farm on Jeju Island in 2010. The 2.12-Mb genome sequence
consists of 44 large contigs in 16 scaffolds and contains 2,214 predicted
protein-coding genes, with a G+C content of 35.4%.

<>

<1>Park, S.-Y., Lee, H.-J., Song, J.-M., Sun, J., Hwang, H.-J., Nishi, K., Kim, J.-S.
<2>Structural characterization of a modification subunit of a putative type I restriction enzyme from Vibrio vulnificus YJ016.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>68
<5>1570-1577
<6>2012
<7>In multifunctional type I restriction enzymes, active methyl-transferases (MTases) are
constituted of methylation (HsdM) and
specificity (HsdS) subunits. In this study, the crystal structure of a
putative HsdM subunit from Vibrio vulnificus YJ016 (vvHsdM) was
elucidated at a resolution of 1.80 angstrom. A cofactor-binding site
for S-adenosyl-L-methionine (SAM, a methyl-group donor) is formed
within the C-terminal domain of an alpha/beta-fold, in which a number
of residues are conserved, including the GxGG and (N/D) PP(F/Y) motifs,
which are likely to interact with several functional moieties of the
SAM methyl-group donor. Comparison with the N6 DNA MTase of Thermus
aquaticus and other HsdM structures suggests that two aromatic rings
(Phe199 and Phe312) in the motifs that are conserved among the HsdMs
may sandwich both sides of the adenine ring of the recognition sequence
so that a conserved Asn residue (Asn309) can interact with the N6 atom
of the target adenine base (a methyl-group acceptor) and locate the
target adenine base close to the transferred SAM methyl group.

<>

<1>Park, S.H., Kim, H.U., Kim, T.Y., Park, J.S., Kim, S.S., Lee, S.Y.
<2>Metabolic engineering of Corynebacterium glutamicum for L-arginine production.
<3>Nat. Commun.
<4>5
<5>4618
<6>2014
<7>L-Arginine is an important amino acid for diverse industrial and health product
applications. Here we report the development of metabolically engineered
Corynebacterium glutamicum ATCC 21831 for the production of L-arginine. Random
mutagenesis is first performed to increase the tolerance of C. glutamicum to
L-arginine analogues, followed by systems metabolic engineering for further
strain improvement, involving removal of regulatory repressors of arginine
operon, optimization of NADPH level, disruption of L-glutamate exporter to
increase L-arginine precursor and flux optimization of rate-limiting L-arginine
biosynthetic reactions. Fed-batch fermentation of the final strain in 5 l and
large-scale 1,500 l bioreactors allows production of 92.5 and 81.2 g l(-1) of
L-arginine with the yields of 0.40 and 0.35 g L-arginine per gram carbon source
(glucose plus sucrose), respectively. The systems metabolic engineering strategy
described here will be useful for engineering Corynebacteria strains for the
industrial production of L-arginine and related products.

<>

<1>Park, S.J., Ghai, R., Martin-Cuadrado, A.B., Rodriguez-Valera, F., Jung, M.Y., Kim, J.G., Rhee, S.K.
<2>Draft Genome Sequence of the Sulfur-Oxidizing Bacterium 'Candidatus Sulfurovum sediminum' AR, Which Belongs to the Epsilonproteobacteria.
<3>J. Bacteriol.
<4>194
<5>4128-4129
<6>2012
<7>Sulfur-oxidizing bacteria are common microorganisms in a variety of sulfide-rich
environments. They play important roles in the global sulfur cycle on earth.
Here, we present a high-quality draft genome sequence of a sulfur-oxidizing
bacterium, 'Candidatus Sulfurovum sediminum' strain AR, which belongs to the
class Epsilonproteobacteria and dominated an enrichment culture from a marine
sediment collected off Svalbard, within the Arctic Circle. Its genome contains
genes for sulfur oxidation and carbon fixation. The size of the draft genome is
2.12 Mb, and the G+C content is 39.4%.

<>

<1>Park, S.J., Kim, J.G., Jung, M.Y., Kim, S.J., Cha, I.T., Ghai, R., Martin-Cuadrado, A.B., Rodriguez-Valera, F., Rhee, S.K.
<2>Draft Genome Sequence of an Ammonia-Oxidizing Archaeon, 'Candidatus Nitrosopumilus sediminis' AR2, from Svalbard in the Arctic Circle.
<3>J. Bacteriol.
<4>194
<5>6948-6949
<6>2012
<7>Ammonia-oxidizing archaea (AOA) typically predominate over ammonia-oxidizing bacteria in
marine sediments. We herein present the draft genome sequence of an
ammonia-oxidizing archaeon, 'Candidatus Nitrosopumilus sediminis' AR2, which was
enriched in culture from a marine sediment obtained off Svalbard, within the
Arctic Circle. The typical genes involved in archaeal ammonia oxidation and
carbon fixation necessary for chemolithoautotrophic growth were observed.
Interestingly, the AR2 genome sequence was revealed to possess, uniquely among
cultivated AOA from marine environments, a capability for urea utilization.

<>

<1>Park, S.J., Kim, J.G., Jung, M.Y., Kim, S.J., Cha, I.T., Kwon, K., Lee, J.H., Rhee, S.K.
<2>Draft Genome Sequence of an Ammonia-Oxidizing Archaeon, 'Candidatus Nitrosopumilus koreensis' AR1, from Marine Sediment.
<3>J. Bacteriol.
<4>194
<5>6940-6941
<6>2012
<7>Ammonia-oxidizing archaea (AOA) are ubiquitous in various marine environments and play
important roles in the global nitrogen and carbon cycles. We here present a
high-quality draft genome sequence of an ammonia-oxidizing archaeon, 'Candidatus
Nitrosopumilus koreensis' AR1, which was found to dominate an ammonia-oxidizing
enrichment culture in marine sediment off Svalbard, the Arctic Circle. Despite a
significant number of nonoverlapping genes (ca. 30%), similarities of this strain
to 'Candidatus Nitrosopumilus maritimus' were revealed by core genes for archaeal
ammonia oxidation and carbon fixation, G+C content, and extensive synteny
conservation.

<>

<1>Park, S.K., Roh, S.W., Whon, T.W., Bae, J.W.
<2>Genome Sequence of Brachybacterium squillarum M-6-3T, Isolated from Salt-Fermented Seafood.
<3>J. Bacteriol.
<4>193
<5>6416-6417
<6>2011
<7>Brachybacterium squillarum M-6-3(T) was isolated from salt-fermented seafood in Korea and
belongs to the Dermabacteraceae, a rather isolated
family within the actinobacterial suborder Micrococcineae. Here, we
present the draft genome sequence of the type strain Brachybacterium
squillarum M-6-3(T) (3,191,479 bp), a Gram-positive bacterium with high
(72.8%) G+C content.

<>

<1>Park, S.N., Cho, E., Kim, H.S., Kim, D.S., Jung, J., Baek, J.H., Kyong, L.Y., Jo, E., Chang, Y.H., Hwan, S.J., Choi, S.H., Kang, J., Choi, Y., Kong, S.W., Han, S.E., Park, H.S., Kim, H., Kook, J.K.
<2>Draft Genome Sequence of Fusobacterium nucleatum subsp. animalis ChDC F324, Isolated from a Human Subgingival Plaque in the Republic of Korea.
<3>Genome Announcements
<4>1
<5>e01042-13
<6>2013
<7>Five subspecies of Fusobacterium nucleatum have been classified: animalis, nucleatum,
polymorphum, vincentii, and fusiforme. F. nucleatum subsp. animalis
ChDC F324 (KCOM 1325) was isolated from a human subgingival plaque in the
Republic of Korea. Here, we report the draft genome sequence of the strain.

<>

<1>Park, S.N., Cho, E., Kim, H.S., Kim, D.S., Jung, J., Baek, J.H., Kyong, L.Y., Jo, E., Chang, Y.H., Hwan, S.J., Choi, S.H., Kang, J., Choi, Y., Park, H.S., Kim, H., Kook, J.K.
<2>Draft Genome Sequence of Fusobacterium nucleatum subsp. nucleatum ChDC F316, Isolated from a Human Peri-implantitis Lesion in the Republic of Korea.
<3>Genome Announcements
<4>1
<5>e01041-13
<6>2013
<7>Fusobacterium nucleatum is a Gram-negative anaerobe and is one of the causative agents of
periodontal diseases, including peri-implantitis. Fusobacterium
nucleatum subsp. nucleatum ChDC F316 (KCOM 1322) was isolated from a human
peri-implantitis lesion. Here, we report the draft genome sequence of this
strain.

<>

<1>Park, S.N., Cho, E., Kim, H.S., Kim, D.S., Jung, J., Baek, J.H., Kyong, L.Y., Jo, E., Chang, Y.H., Hwan, S.J., Kim, J., Choi, S.H., Kang, J., Choi, Y., Park, H.S., Kim, H., Kook, J.K.
<2>Draft Genome Sequence of Fusobacterium nucleatum subsp. vincentii ChDC F8, Isolated from a Human Subgingival Plaque in the Republic of Korea.
<3>Genome Announcements
<4>1
<5>e01040-13
<6>2013
<7>Fusobacterium nucleatum is a Gram-negative, nonmotile, obligately anaerobic rod bacterium
which might play an important role in the initiation and progression of
periodontal diseases. F. nucleatum subsp. vincentii ChDC F8 (KCOM 1231) was
isolated from a human gingivitis lesion. Here, we report the draft genome
sequence of the strain.

<>

<1>Park, S.N., Cho, E., Lim, Y.K., Kim, H.S., Kim, D.S., Jung, J., Baek, J.H., Jo, E., Chang, Y.H., Shin, J.H., Choi, S.H., Kang, J., Choi, Y., Park, H.S., Kim, H., Kook, J.K.
<2>Draft Genome Sequences of Fusobacterium nucleatum ChDC F145, ChDC F174, ChDC F206, and ChDC F300, Isolated from Human Subgingival Plaques in the Republic of  Korea.
<3>Genome Announcements
<4>2
<5>e01233-13
<6>2014
<7>Recently, five strains were isolated from human subgingival plaques and were proposed as a
novel subspecies of Fusobacterium nucleatum. Here, we report the
draft genome sequences of the strains, except one for which the draft sequence
was already introduced.

<>

<1>Park, S.N., Kong, S.W., Kim, H.S., Park, M.S., Lee, J.W., Cho, E., Lim, Y.K., Choi, M.H., Chang, Y.H., Shin, J.H., Park, H.S., Choi, S.H., Kook, J.K.
<2>Draft Genome Sequence of Fusobacterium nucleatum ChDC F128, Isolated from a Periodontitis Lesion.
<3>J. Bacteriol.
<4>194
<5>6322-6323
<6>2012
<7>Fusobacterium nucleatum is classified into five subspecies. F. nucleatum ChDC F128 was
isolated from a periodontitis lesion and proposed as a new subspecies
based on the comparison of the nucleotide sequences of the RNA polymerase beta
subunit and zinc protease genes. Here, we report the draft genome sequence of the
strain.

<>

<1>Park, S.N., Kong, S.W., Park, M.S., Lee, J.W., Cho, E., Lim, Y.K., Choi, M.H., Kim, H.S., Chang, Y.H., Shin, J.H., Park, H.S., Choi, S.H., Kook, J.K.
<2>Draft Genome Sequence of Fusobacterium nucleatum subsp. fusiforme ATCC 51190T.
<3>J. Bacteriol.
<4>194
<5>5445-5446
<6>2012
<7>Fusobacterium nucleatum, one of the major causative bacteria of periodontitis, is classified
into five subspecies (nucleatum, polymorphum, vincentii, animalis, and
fusiforme) on the basis of the several phenotypic characteristics and DNA
homology. This is the first report of the draft genome sequence of F. nucleatum
subsp. fusiforme ATCC 51190(T).

<>

<1>Park, T.H., Choi, B.S., Choi, A.Y., Choi, I.Y., Heu, S., Park, B.S.
<2>Genome Sequence of Pectobacterium carotovorum subsp. carotovorum Strain PCC21, a  Pathogen Causing Soft Rot in Chinese Cabbage.
<3>J. Bacteriol.
<4>194
<5>6345-6346
<6>2012
<7>Pectobacterium carotovorum is a plant-pathogenic enterobacterium responsible for  soft rot in
various commercially important plants. Here we report the complete
genome sequence and automatic annotation of strain PCC21.

<>

<1>Park, Y., Kim, G.-D., Choi, T.-J.
<2>Molecular cloning and characterization of the DNA adenine methyltransferase gene in Feldmannia sp virus.
<3>Virus Genes
<4>34
<5>177-183
<6>2007
<7>The genome of Feldmannia sp. virus (FsV), a marine brown alga virus, contains a putative DNA
adenine methyltransferase (dam) gene of 1,245
bp that encodes a polypeptide of 45.8 kDa. A BLAST search with the FsV
dam gene showed high amino acid identity to two putative
methyltransferase genes, ORF B29 of Feldmannia irregularis virus
(FirrV, 54%) and ORF129 of Ectocarpus siliculosus virus (EsV, 36%); and
a PSI BLAST search revealed similarity to the N-6-adenine
methyltransferases (MTases) of other species. Most conserved motifs of
beta-class MTases were observed in the FsV dam gene. However, neither
of the highly conserved sequences in motifs I (FxGxG) or IV
[(S/N/D)PP(Y/F/W)] perfectly matched those in the FsV dam gene. The
highly conserved DPPY consensus sequence in motif IV was NTPW in the
FsV dam gene, perfectly matching the sequences in ORF B29 of FirrV and
ORF129 of EsV. Therefore, the dam genes in brown algae viruses may
belong to a yet undiscovered group. The FsV Dam protein expressed from
the cloned FsV dam gene methylated E. coli chromosomal DNA. This is the
first report showing that a virus infecting marine filamentous brown
algae encodes a functional Dam protein.

<>

<1>Park, Y.J., Park, H.J., Jung, K.H.
<2>Feldmannia.
<3>Korean Patent Office
<4>KR 1020050121681 A
<5>
<6>2005
<7>
<>

<1>Park, Y.K., Kang, H., Yoo, H., Lee, S.H., Roh, H., Kim, H.J., Ryoo, S.
<2>Whole-Genome Sequence of Mycobacterium tuberculosis Korean Strain KIT87190.
<3>Genome Announcements
<4>2
<5>e01103-14
<6>2014
<7>Mycobacterium tuberculosis is a contagious agent that causes tuberculosis. A specific type
(called the K cluster) of M. tuberculosis with 10 copies of IS6110
in restriction fragment length polymorphism (RFLP) has been found in about 4% of
M. tuberculosis isolates in Korea. Here, we report the complete genome sequence
of M. tuberculosis Korean strain KIT87190 belonging to the K cluster.

<>

<1>Parker, B., Marinus, M.G.
<2>A simple and rapid method to obtain substitution mutations in Escherichia coli:  isolation of a dam deletion/insertion mutation.
<3>Gene
<4>73
<5>531-535
<6>1988
<7>We describe the isolation of a strain of Escherichia coli bearing a deletion/insertion (i.e.,
a substitution mutation) in the dam gene (dam-16). The mutagenesis protocol used should be
applicable to any cloned non-essential gene of E. coli. The substitution mutation confers
resistance to kanamycin and can easily be transferred to other strains by standard genetic
techniques. The amount of Dam methyltransferase (MTase) in dam-16 strains as determined either
in vitro or in vivo is below the level of detection. We conclude that the Dam MTase is not
required for viability of E. coli.

<>

<1>Parker, C.T., Gorski, L., Huynh, S.
<2>Complete Genome Sequence of Salmonella enterica subsp. enterica Serovar Thompson  Strain RM6836.
<3>Genome Announcements
<4>1
<5>e00900-13
<6>2013
<7>Salmonella enterica subsp. enterica serovar Thompson strain RM6836 was isolated from lettuce
in 2002. We report here the complete sequence and annotation of the
genome of S. Thompson RM6836. This is the first reported complete genome sequence
for S. Thompson and it will enhance our understanding of this serovar and provide
another point for comparative studies between Salmonella enterica strains.

<>

<1>Parker, D., Narechania, A., Sebra, R., Deikus, G., Larussa, S., Ryan, C., Smith, H., Prince, A., Mathema, B., Ratner, A.J., Kreiswirth, B., Planet, P.J.
<2>Genome Sequence of Bacterial Interference Strain Staphylococcus aureus 502A.
<3>Genome Announcements
<4>2
<5>e00284-14
<6>2014
<7>Staphylococcus aureus 502A was a strain used in bacterial interference programs
during the 1960s and early 1970s. Infants were deliberately colonized with 502A
with the goal of preventing colonization with more invasive strains. We present
the completed genome sequence of this organism.

<>

<1>Parker, M.M., Belisle, M., Belfort, M.
<2>Intron homing with limited exon homology. Illegitimate double-strand-break repair in intron acquisition by phage t4.
<3>Genetics
<4>153
<5>1513-1523
<6>1999
<7>The td intron of bacteriophage T4 encodes a DNA endonuclease that initiates intron homing to
cognate intronless alleles by a double-strand-break (DSB) repair process. A genetic assay was
developed to analyze the relationship between exon homology and homing efficiency. Because
models predict exonucleolytic processing of the cleaved recipient leading to homologous strand
invasion of the donor allele, the assay was performed in wild-type and exonuclease-deficient
(rnh or dexA) phage. Efficient homing was supported by exon lengths of 50 bp or greater,
whereas more limited exon lengths led to a precipitous decline in homing levels. However,
extensive homology in one exon still supported elevated homing levels when the other exon was
completely absent. Analysis of these "one-sided" events revealed recombination junctions at
ectopic sites of microhomology and implicated nucleolytic degradation in illegitimate DSB
repair in T4. Interestingly, homing efficiency with extremely limiting exon homology was
greatly elevated in phage deficient in the 3'-5' exonuclease, DexA, suggesting that the
length of 3' tails is a major determinant of the efficiency of DSB repair. Together, these
results suggest that illegitimate DSB repair may provide a means by which introns can invade
ectopic sites.

<>

<1>Parkhill, J. et al.
<2>The genome sequence of the food-borne pathogen Campylobacter jejuni reveals hypervariable sequences.
<3>Nature
<4>403
<5>665-668
<6>2000
<7>Campylobacter jejuni, from the delta-epsilon group of proteobacteria, is a microaerophilic,
Gram-negative, flagellate, spiral bacterium-properties it shares with the related gastric
pathogen Helicobacter pylori.  It is the leading cause of bacterial food-borne diarrhoeal
disease throughout the world.  In addition, infection with C. jejuni is the most frequent
antecedent to a form of neuromuscular paralysis known as Guillain-Barre syndrome.  Here we
report the genome sequence of C. jejuni NCTC11168.  C. jejuni has a circular chromosome of
1,641,481 base pairs (30.6% G+C) which is predicted to encode 1,654 proteins and 54 stable RNA
species.  The genome is unusual in that there are virtually no insertion sequences or
phage-associated sequences and very few repeat sequences.  One of the most striking findings
in the genome was the presence of hypervariable sequences.  These short homopolymeric runs of
nucleotides were commonly found in genes encoding the biosynthesis or modification of surface
structures, or in closely linked genes of unknown function.  The apparently high rate of
variation of these homopolymeric tracts may be important in the survival strategy of C.
jejuni.

<>

<1>Parkhill, J. et al.
<2>Complete DNA sequence of a serogroup A strain of Neisseria meningitidis Z2491.
<3>Nature
<4>404
<5>502-506
<6>2000
<7>Neisseria meningitidis causes bacterial meningitis and is therefore responsible for
considerable morbidity and mortality in both the developed and the developing world.
Meningococci are opportunistic pathogens that colonize the nasopharynges and oropharynges of
asymptomatic carriers. For reasons that are still mostly unknown, they occasionally gain
access to the blood, and subsequently to the cerebrospinal fluid, to cause septicaemia and
meningitis. N. meningitidis strains are divided into a number of serogroups on the basis of
the immunochemistry of their capsular polysaccharides; serogroup A strains are responsible for
major epidemics and pandemics of meningococcal disease, and therefore most of the morbidity
and mortality associated with this disease. Here we have determined the complete genome
sequence of a serogroup A strain of Neisseria meningitidis, Z2491. The sequence is 2,184,406
base pairs in length, with an overall G+C content of 51.8%, and contains 2,121 predicted
coding sequences. The most notable feature of the genome is the presence of many hundreds of
repetitive elements, ranging from short repeats, positioned either singly or in large multiple
arrays, to insertion sequences and gene duplications of one kilobase or more. Many of these
repeats appear to be involved in genome fluidity and antigenic variation in this important
human pathogen.

<>

<1>Parkhill, J. et al.
<2>Genome sequence of Yersinia pestis, the causative agent of plague.
<3>Nature
<4>413
<5>523-527
<6>2001
<7>The Gram-negative bacterium Yersinia pestis is the causative agent of the systemic invasive
infectious disease classically referred to as plague, and has been responsible for three human
pandemics: the Justinian plague (sixth to eighth centuries), the Black Death (fourteenth to
nineteenth centuries) and modern plague (nineteenth century to the present day). The recent
identification of strains resistant to multiple drugs and the potential use of Y. pestis as an
agent of biological warfare mean that plague still poses a threat to human health. Here we
report the complete genome sequence of Y. pestis strain CO92, consisting of a 4.65-megabase
(Mb) chromosome and three plasmids of 96.2 kilobases (kb), 70.3 kb and 9.6 kb. The genome is
unusually rich in insertion sequences and displays anomalies in GC base-composition bias,
indicating frequent intragenomic recombination. Many genes seem to have been acquired from
other bacteria and viruses (including adhesins, secretion systems and insecticidal toxins).
The genome contains around 150 pseudogenes, many of which are remnants of a redundant
enteropathogenic lifestyle. The evidence of ongoing genome fluidity, expansion and decay
suggests Y. pestis is a pathogen that has undergone large-scale genetic flux and provides a
unique insight into the ways in which new and highly virulent pathogens evolve.

<>

<1>Parkhill, J. et al.
<2>Complete genome sequence of a multiple drug resistant Salmonella enterica serovar Typhi CT18.
<3>Nature
<4>413
<5>848-852
<6>2001
<7>Salmonella enterica serovar Typhi (S. typhi) is the aetiological agent of typhoid fever, a
serious invasive bacterial disease of humans with an annual global burden of approximately 16
million cases, leading to 600,000 fatalities. Many S. enterica serovars actively invade the
mucosal surface of the intestine but are normally contained in healthy individuals by the
local immune defence mechanisms. However, S. typhi has evolved the ability to spread to the
deeper tissues of humans, including liver, spleen and bone marrow. Here we have sequenced the
4,809,037-base pair (bp) genome of a S. typhi (CT18) that is resistant to multiple drugs,
revealing the presence of hundreds of insertions and deletions compared with the Escherichia
coli genome, ranging in size from single genes to large islands. Notably, the genome sequence
identifies over two hundred pseudogenes, several corresponding to genes that are known to
contribute to virulence in Salmonella typhimurium. This genetic degradation may contribute to
the human-restricted host range for S. typhi. CT18 harbours a 218,150-bp
multiple-drug-resistance incH1 plasmid (pHCM1), and a 106,516-bp cryptic plasmid (pHCM2),
which shows recent common ancestry with a virulence plasmid of Yersinia pestis.

<>

<1>Parraga, A., Portugal, J.
<2>Detection of elsamicin-DNA binding specificity by restriction enzyme cleavage.
<3>FEBS Lett.
<4>300
<5>25-29
<6>1992
<7>The sequence specificity of elsamicin A, an anti-tumour antibotic, binding to
DNA was elucidated considering the inhibition of the rate of digestion of
linearised pBR322 DNA by AatII, ClaI, EcoRI, HindIII and NruI restriction
enzymes.  Elsamicin A inhibits the rate of digestion by NruI (recognition
sequence TCG/CGA) to a greater extent than it does for the other enzymes, thus
evidencing the sequence-selective binding of elsamicin to CGC regions in DNA.
Our results also show the important role of the neighbouring sequences in the
elsamicin A-DNA interactions and their effects on the cleavage by restriction
enzymes.

<>

<1>Parreira, V.R., Costa, M., Eikmeyer, F., Blom, J., Prescott, J.F.
<2>Sequence of Two Plasmids from Clostridium perfringens Chicken Necrotic Enteritis Isolates and Comparison with C. perfringens Conjugative Plasmids.
<3>PLoS ONE
<4>7
<5>E49753
<6>2012
<7>Twenty-six isolates of Clostridium perfringens of different MLST types from
chickens with necrotic enteritis (NE) (15 netB-positive) or from healthy chickens
(6 netB-positive, 5 netB-negative) were found to contain 1-4 large plasmids, with
most netB-positive isolates containing 3 large and variably sized plasmids which
were more numerous and larger than plasmids in netB-negative isolates. NetB and
cpb2 were found on different plasmids consistent with previous studies. The
pathogenicity locus NELoc1, which includes netB, was largely conserved in these
plasmids whereas NeLoc3, present in the cpb2 containing plasmids, was less well
conserved. A netB-positive and a cpb2-positive plasmid were likely to be
conjugative, and the plasmids were completely sequenced. Both plasmids possessed
the intact tcp conjugative region characteristic of C. perfringens conjugative
plasmids. Comparative genomic analysis of nine CpCPs, including the two plasmids
described here, showed extensive gene rearrangements including pathogenicity
locus and accessory gene insertions around rather than within the backbone
region. The pattern that emerges from this analysis is that the major
toxin-containing regions of the variety of virulence-associated CpCPs are
organized as complex pathogenicity loci. How these different but related CpCPs
can co-exist in the same host has been an unanswered question. Analysis of the
replication-partition region of these plasmids suggests that this region controls
plasmid incompatibility, and that CpCPs can be grouped into at least four
incompatibility groups.

<>

<1>Parro, V., Moreno-Paz, M., Gonzalez-Toril, E.
<2>Analysis of environmental transcriptomes by DNA microarrays.
<3>Environ. Microbiol.
<4>9
<5>453-464
<6>2007
<7>In this work we investigated the correlations between global gene expression patterns and
environmental parameters in natural ecosystems. We
studied the preferential gene expression of the iron oxidizer bacterium
Leptospirillum ferrooxidans to adapt its physiology to changes in the
physicochemical parameters in its natural medium. Transcriptome analysis
by DNA microarrays can proportionate an instant picture about the
preferential gene expression between two different environmental samples.
However, this type of analysis is very difficult and complex in natural
ecosystems, mainly because of the broad biodiversity and multiple
environmental parameters that may affect gene expression. The necessity of
high-quality RNA preparations as well as complicated data analysis are
also technological limitations. The low prokaryotic diversity of the
extremely acidic and iron-rich waters of the Tinto River (Spain)
ecosystem, where L. ferrooxidans is abundant, allows the opportunity to
achieve global gene expression studies and to associate gene function with
environmental parameters. We applied a total RNA amplification protocol
validated previously for the amplification of the environmental
transcriptome (meta-transcriptome). The meta-transcriptome of two sites
from the Tinto River mainly differing in the salt and oxygen contents were
amplified and analysed by a L. ferrooxidans DNA microarray. The results
showed a clear preferential induction of genes involved in certain
physicochemical parameters like: high salinity (ectAB, otsAB), low oxygen
concentration (cydAB), iron uptake (fecA-exbBD-tonB), oxidative stress
(carotenoid synthesis, oxyR, recG), potassium (kdpBAC) or phosphate
concentrations (pstSCAB), etc. We conclude that specific gene expression
patterns can be useful indicators for the physiological conditions in a
defined ecosystem. Also, the upregulation of certain genes and operons
reveals information about the environmental conditions (nutrient
limitations, stresses, etc.).

<>

<1>Parry, D., Moon, S.A., Liu, H.H., Heslop, P., Connolly, B.A.
<2>DNA Recognition by the EcoRV Restriction Endonuclease Probed using Base Analogues.
<3>J. Mol. Biol.
<4>331
<5>1005-1016
<6>2003
<7>The EcoRV restriction endonuclease recognises palindromic GATATC sequences and cuts between
the central T and dA bases in a reaction that has an
absolute requirement for a divalent metal ion, physiologically Mg(2+). Use
has been made of base analogues, which delete hydrogen bonds between the
protein and DNA (or hydrophobic interactions in the case of the 5-CH(3)
group of thymine), to evaluate the roles of the outer two base-pairs
(GATATC) in DNA recognition. Selectivity arises at both the binding steps
leading to the formation of the enzyme-DNA-metal ion ternary complex
(assayed by measuring the dissociation constant in the presence of the
non-reactive metal Ca(2+)) and the catalytic step (evaluated using
single-turnover hydrolysis in the presence of Mg(2+)), with each
protein-DNA contact contributing to recognition. With the A:T base-pair,
binding was reduced by the amount expected for the simple loss of a single
contact; much more severe effects were observed with the G:C base-pair,
suggesting additional conformational perturbation. Most of the modified
bases lowered the rate of hydrolysis; furthermore, the presence of an
analogue in one strand of the duplex diminished cutting at the second,
unmodified strand, indicative of communication between DNA binding and the
active site. The essential metal ion Mg(2+) plays a key role in mediating
interactions between the DNA binding site and active centre and in many
instances rescue of hydrolysis was seen with Mn(2+). It is suggested that
contacts between the GATATC site are required for tight binding and for
the correct assembly of metal ions and bound water at the catalytic site,
functions important in providing acid/base catalysis and transition state
stabilisation.

<>

<1>Parsa, Y.L., Azarbaijani, R., Sarikhan, S., Mousavi, H., Ramezani, M., Amoozegar, M.A., Shahzadeh, F.A., Salekdeh, G.H.
<2>Complete Genome Sequence of Oceanimonas sp. GK1, a Halotolerant Bacterium from Gavkhouni Wetland in Iran.
<3>J. Bacteriol.
<4>194
<5>2123-2124
<6>2012
<7>Oceanimonas sp. GK1 (IBRC-M 10197) is a marine halotolerant gammaproteobacterium  which was
characterized as producing large amounts of poly-beta-hydroxybutyrate.
Here we present the whole-genome sequence of Oceanimonas sp. GK1, which consists
of a single circular chromosome of 3,514,537 bp and two plasmids 8,462 and 4,245
bp in length.

<>

<1>Parschat, K., Hauer, B., Kappl, R., Kraft, R., Huttermann, J., Fetzner, S.
<2>Gene cluster of Arthrobacter ilicis Ru61a involved in the degradation of quinaldine to anthranilate: characterization and functional expression of the quinaldine 4-oxidase qoxLMS genes.
<3>J. Biol. Chem.
<4>278
<5>27483-27494
<6>2003
<7>A genetic analysis of the anthranilate pathway of quinaldine degradation was
performed. A 23-kb region of DNA from Arthrobacter ilicis Ru61a was cloned into
the cosmid pVK100. Although Escherichia coli clones containing the recombinant
cosmid did not transform quinaldine, cosmids harboring the 23-kb region, or a
10.8-kb stretch of this region, conferred to Pseudomonas putida KT2440 the
ability to cometabolically convert quinaldine to anthranilate. The 10.8-kb
fragment thus contains the genes coding for quinaldine 4-oxidase (Qox),
1H-4-oxoquinaldine 3-monooxygenase, 1H-3-hydroxy-4-oxoquinaldine 2,4-dioxygenase,
and N-acetylanthranilate amidase. The qoxLMS genes coding for the molybdopterin
cytosine dinucleotide-(MCD-), FeSI-, FeSII-, and FAD-containing Qox were inserted
into the expression vector pJB653, generating pKP1. Qox is the first
MCD-containing enzyme to be synthesized in a catalytically fully competent form
by a heterologous host, P. putida KT2440 pKP1; the catalytic properties and the
UV-visible and EPR spectra of Qox purified from P. putida KT2440 pKP1 were
essentially like those of wild-type Qox. This provides a starting point for the
construction of protein variants of Qox by site-directed mutagenesis. Downstream
of the qoxLMS genes, a putative gene whose deduced amino acid sequence showed 37%
similarity to the cofactor-inserting chaperone XdhC was located. Additional open
reading frames identified on the 23-kb segment may encode further enzymes (a
glutamyl tRNA synthetase, an esterase, two short-chain dehydrogenases/reductases,
an ATPase belonging to the AAA family, a 2-hydroxyhepta-2,4-diene-1,7-dioate
isomerase/5-oxopent-3-ene-1,2,5-tricarboxylate decarboxylase-like protein, and an
enzyme of the mandelate racemase group) and hypothetical proteins involved in
transcriptional regulation, and metabolite transport.

<>

<1>Parschat, K., Overhage, J., Strittmatter, A.W., Henne, A., Gottschalk, G., Fetzner, S.
<2>Complete Nucleotide Sequence of the 113-Kilobase Linear Catabolic Plasmid pAL1 of Arthrobacter nitroguajacolicus Ru61a and Transcriptional Analysis of Genes Involved in Quinaldine Degradation.
<3>J. Bacteriol.
<4>189
<5>3855-3867
<6>2007
<7>The nucleotide sequence of the linear catabolic plasmid pAL1 from the 2-methylquinoline
(quinaldine)-degrading strain Arthrobacter nitroguajacolicus Rü61a comprises 112,992 bp. A
total of 103 open reading frames (ORFs) were identified on pAL1, 49 of which had no
annotatable function. The ORFs were assigned to the following functional groups: (i)
catabolism of quinaldine and anthranilate, (ii) conjugation, and (iii) plasmid maintenance and
DNA replication and repair. The genes for conversion of quinaldine to anthranilate are
organized in two operons that include ORFs presumed to code for proteins involved in assembly
of the quinaldine-4-oxidase holoenzyme, namely, a MobA-like putative molybdopterin cytosine
dinucleotide synthase and an XdhC-like protein that could be required for insertion of the
molybdenum cofactor. Genes possibly coding for enzymes involved in anthranilate degradation
via 2-aminobenzoyl coenzyme A form another operon. These operons were expressed when cells
were grown on quinaldine or on aromatic compounds downstream in the catabolic pathway.
Single-stranded 3' overhangs of putative replication intermediates of pAL1 were predicted to
form elaborate secondary structures due to palindromic and superpalindromic terminal
sequences; however, the two telomeres appear to form different structures. Sequence analysis
of ORFs 101 to 103 suggested that pAL1 codes for one or two putative terminal proteins,
presumed to be covalently bound to the 5' termini, and a multidomain telomere-associated
protein (Tap) comprising 1,707 amino acids. Even if the putative proteins encoded by ORFs 101
to 103 share motifs with the Tap and terminal proteins involved in telomere patching of
Streptomyces linear replicons, their overall sequences and domain structures differ
significantly.

<>

<1>Partridge, S.R., Ginn, A.N., Paulsen, I.T., Iredell, J.R.
<2>pEl1573 carrying blaIMP-4 from Sydney, Australia, is closely related to other IncL/M plasmids.
<3>Antimicrob. Agents Chemother.
<4>56
<5>6029-6032
<6>2012
<7>Complete sequencing of pEl1573, a representative IncL/M plasmid carrying blaIMP-4 from Sydney,
Australia, revealed a ~60 kb backbone almost identical to those of IncL/M plasmids
pCTX-M3,from Poland, and pCTX-M360, from China, and less closely related to pNDM-HK, pOXA-48a
and pEL60, suggesting different lineages. The ~28 kb Tn2-derived multiresistance region in
pEl1573 is inserted in the same location as those in pCTX-M3 and pNDM-HK and shares some of
the same components, but has undergone rearrangements.

<>

<1>Partridge, S.R., Zong, Z., Iredell, J.R.
<2>Recombination in IS26 and Tn2 in the Evolution of Multiresistance Regions Carrying blaCTX-M-15 on Conjugative IncF Plasmids from Escherichia coli.
<3>Antimicrob. Agents Chemother.
<4>55
<5>4971-4978
<6>2011
<7>CTX-M-15 now appears to be the dominant extended-spectrum beta-lactamase
worldwide, and a number of different factors may contribute to this
success. These include associations between bla(CTX-M-15) and particular
plasmids (IncF) and/or strains, such as Escherichia coli ST131, as well as
the genetic contexts in which this gene is found. We previously identified
bla(CTX-M-15) as the dominant ESBL gene in the western Sydney area,
Australia, and found that it was carried mainly on IncF or IncI1 plasmids.
Here, we have mapped the multiresistance regions of the 11 conjugative
plasmids with one or more IncF replicons obtained from that survey and
conducted a limited comparison of plasmid backbones. Two plasmids with
only an IncFII replicon appear to be very similar to the published
plasmids pC15-1a and pEK516. The remaining nine plasmids, with multiple
IncF replicons, have multiresistance regions related to those of pC15-1a
and pEK516, but eight contain additional modules previously found in
resistance plasmids from different geographic locations that carry a
variety of different resistance genes. Differences between the
multiresistance regions are largely due to IS26-mediated deletions,
insertions, and/or rearrangements, which can explain the observed variable
associations between bla(CTX-M-15) and certain other resistance genes. We
found no evidence of independent movement of bla(CTX-M-15) or of a large
multiresistance region between different plasmid backbones. Instead,
homologous recombination between common components, such as IS26 and Tn2,
appeared to be more important in creating new multiresistance regions, and
this may be coupled with recombination in plasmid backbones to reassort
multiple IncF replicons as well as components of multiresistance regions.

<>

<1>Pascopella, L., Raupach, B., Ghori, N., Monack, D., Falkow, S., Small, P.L.C.
<2>Host restriction phenotypes of Salmonella typhi and Salmonella gallinarum.
<3>Infect. Immun.
<4>63
<5>4329-4335
<6>1995
<7>Salmonella typhi and Salmonella gallinarum phenotypes correlated with mouse host restriction
have been identified by using in vitro and in vivo systems. S. typhi is capable of entering
the murine intestinal epithelium via M cells, as is Salmonella typhimurium, which causes
systemic infection in the mouse.  But, unlike S. typhimurium, S. typhi does not destroy the
epithelium and is cleared from the Peyer's patches soon after M-cell entry, S. gallinarum
appears to be incapable of entering the murine Peyer's patch epithelium.  Our in vitro
evidence suggests that S. gallinarum is taken up in murine phagocytic cells by a mechanism
different from that of S. typhimurium.  S. typhimurium is taken up at a higher frequency and
is maintained at higher viable counts throughout a 24-h time course in a murine
macrophage-like cell line than are S. gallinarum and S. typhi.

<>

<1>Pashenkov, A.L., Nikolskaya, I.I., Nigmatullin, T.G., Debov, S.S.
<2>Search for DNA host specificity system in Salmonella typhi.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>4
<5>3-6
<6>1985
<7>Seventeen pure lines of S. typhi bacteriophages have been obtained from mother races O and Vi;
of these, three were used to study 152 S. typhi strains with a view to detecting their DNA
host specificity systems.  10 S. typhi strains having the DNA host specificity system have
been detected by the rough determination of the lytic spectrum and the cross titration of
phage Vi IX.

<>

<1>Passerini, D., Vuillemin, M., Laguerre, S., Amari, M., Loux, V., Gabriel, V., Robert, H., Morel, S., Monsan, P., Gabriel, B., Fontagne-Faucher, C., Remaud-Simeon, M., Moulis, C.
<2>Complete Genome Sequence of Leuconostoc citreum Strain NRRL B-742.
<3>Genome Announcements
<4>2
<5>e01179-14
<6>2014
<7>Leuconostoc citreum belongs to the group of lactic acid bacteria and plays an important role
in fermented foods of plant origin. Here, we report the complete
genome of the Leuconostoc citreum strain NRRL B-742, isolated in 1954 for its
capacity to produce dextran.

<>

<1>Passos-da-Silva, D., Devescovi, G., Paszkiewicz, K., Moretti, C., Buonaurio, R., Studholme, D.J., Venturi, V.
<2>Draft Genome Sequence of Erwinia toletana, a Bacterium Associated with Olive Knots Caused by Pseudomonas savastanoi pv. Savastanoi.
<3>Genome Announcements
<4>1
<5>e00205-13
<6>2013
<7>Erwinia toletana was first reported in 2004 as a bacterial species isolated from  olive knots
caused by the plant bacterium Pseudomonas savastanoi pv. savastanoi.
Recent studies have shown that the presence of this bacterium in the olive knot
environment increases the virulence of the disease, indicating possible
interspecies interactions with P. savastanoi pv. savastanoi. Here, we report the first draft
genome sequence of an E. toletana strain.

<>

<1>Pasternak, Z., Pietrokovski, S., Rotem, O., Gophna, U., Lurie-Weinberger, M.N., Jurkevitch, E.
<2>By their genes ye shall know them: genomic signatures of predatory bacteria.
<3>ISME J.
<4>7
<5>756-769
<6>2013
<7>Predatory bacteria are taxonomically disparate, exhibit diverse predatory
strategies and are widely distributed in varied environments. To date, their
predatory phenotypes cannot be discerned in genome sequence data thereby limiting
our understanding of bacterial predation, and of its impact in nature. Here, we
define the 'predatome,' that is, sets of protein families that reflect the
phenotypes of predatory bacteria. The proteomes of all sequenced 11 predatory
bacteria, including two de novo sequenced genomes, and 19 non-predatory bacteria
from across the phylogenetic and ecological landscapes were compared. Protein
families discriminating between the two groups were identified and quantified,
demonstrating that differences in the proteomes of predatory and non-predatory
bacteria are large and significant. This analysis allows predictions to be made,
as we show by confirming from genome data an over-looked bacterial predator. The
predatome exhibits deficiencies in riboflavin and amino acids biosynthesis,
suggesting that predators obtain them from their prey. In contrast, these genomes
are highly enriched in adhesins, proteases and particular metabolic proteins,
used for binding to, processing and consuming prey, respectively. Strikingly,
predators and non-predators differ in isoprenoid biosynthesis: predators use the
mevalonate pathway, whereas non-predators, like almost all bacteria, use the DOXP
pathway. By defining predatory signatures in bacterial genomes, the predatory
potential they encode can be uncovered, filling an essential gap for measuring
bacterial predation in nature. Moreover, we suggest that full-genome proteomic
comparisons are applicable to other ecological interactions between microbes, and
provide a convenient and rational tool for the functional classification of
bacteria.

<>

<1>Patel, B.K.
<2>Draft Genome Sequence of Fervidicella metallireducens Strain AeBT, an Iron-Reducing Thermoanaerobe from the Great Artesian Basin.
<3>Genome Announcements
<4>2
<5>e00345-14
<6>2014
<7>The genome sequence of Fervidicella metallireducens strain AeB(T), a curved, heterotrophic,
thermoanaerobic, and iron-reducing bacterium isolated from a gray
microbial mat colonizing the free-flowing waters of a Great Artesian Basin (GAB)
bore well located in outback Queensland, Australia, is reported here. The
analysis of the 2.9-Mb sequence indicates that the attributes of the genome are
consistent with its physiological and phenotypic traits.

<>

<1>Patel, D.J.
<2>A molecular handshake.
<3>Nature
<4>367
<5>688-690
<6>1994
<7>Review of the M.HhaI/DNA crystal structure paper.

<>

<1>Patel, H.K., Passos-da-Silva, D., Devescovi, G., Maraite, H., Paszkiewicz, K., Studholme, D.J., Venturi, V.
<2>Draft Genome Sequence of Pseudomonas fuscovaginae, a Broad-Host-Range Pathogen of Plants.
<3>J. Bacteriol.
<4>194
<5>2765-2766
<6>2012
<7>Pseudomonas fuscovaginae was first reported as a pathogen of rice causing sheath  rot in
plants grown at high altitudes. P. fuscovaginae is now considered a
broad-host-range plant pathogen causing disease in several economically important
plants. We report what is, to our knowledge, the first draft genome sequence of a
P. fuscovaginae strain.

<>

<1>Patel, J., Firman, K.
<2>The domain structure of the DNA specificity subunit of type I restriction endonucleases. I. Cloning, mutagenesis and over-production of the EcoR124 DNA methyltransferase.
<3>Proc. Eur. Meet. Genet. Transform., Intercept, Balla, E., Berencsi, G., Szentirmai, A., Andover, UK
<4>0
<5>179-187
<6>1993
<7>Type I restriction endonucleases consist of three subunits encoded by the genes hsdR, hsdM and
hsdS.  The hsdS gene product is responsible for DNA specificity and together with the hsdM
gene product is sufficient for modification (methylation) of the DNA recognition sequence;
hsdR is required for endonuclease activity.  The endonuclease requires ATP, SAM and Mg2+ as
cofactors while the DNA methyltransferase requires only SAM and Mg2+.  Evidence has been
presented to support the hypothesis that the HsdS protein has two domains responsible for DNA
recognition, separated by a spacer region.  The EcoR124 R-M system is of particular interest
in that an alternative DNA specificity (EcoR124/3) has been identified which differs from the
EcoR124 DNA specificity by the presence of an extra non-specific nucleotide:  EcoR124 -
GAANNNNNNRTCG (or GAAN6RTCG), EcoR124/3 - GAANNNNNNNRTCG (or GAAN7RTCG).  EcoR124 and its
variant form EcoR124/3 have been cloned producing the recombinant plasmids pCP1005 and pUNG31
respectively, their transcripts mapped, the genes sequenced and low levels of the endonuclease
have been purified.  These data show that they are members of a new sub-class of type I R-M
systems: type IC.  The only difference between EcoR124 and EcoR124/3 lies within their hsdS
genes.  The EcoR124/3 hsdS gene has an extra copy of a 12-bp repeat within the predicted
spacer region (repeated twice in EcoR124 and three times in EcoR124/3).

<>

<1>Patel, J., Taylor, I., Dutta, C.F., Kneale, G., Firman, K.
<2>High-level expression of the cloned genes encoding the subunits of and intact DNA methyltransferase, M.EcoR124.
<3>Gene
<4>112
<5>21-27
<6>1992
<7>We have cloned the genes coding for the two subunits (HsdM and HsdS) of the
type-I DNA methyltransferase (MTase), M.EcoR124, into the specially constructed
expression vector, pJ119.  These subunit have been synthesized together as an
intact MTase.  WE have also cloned the individual subunit-encoding genes under
the control of the T7 gene 10 promoter or the lacUV5 promoter.  High levels of
expression have been obtained in all cases.  While HsdM was found to be
soluble, HsdS was insoluble.  However, in the presence of the co-produced HsdM
subunit, HsdS was found in the soluble fraction as part of an active MTase.  We
have partially purified the cloned multi-subunit enzyme and shown that it is
capable of DNA methylation both in vivo and in vitro.

<>

<1>Patel, N., Graslund, A., Berglund, H., Nilsson, L., Rigler, R., McLaughlin, L.W.
<2>Interaction of a minor groove binder with a fluorescent DNA oligomer containing the EcoRI recognition sequence.
<3>Nucleosides and Nucleotides
<4>10
<5>547-548
<6>1991
<7>The minor groove binding drug netropsin quenches the 2-aminopurine (2AP)
fluorescence in the duplex d(CTGA(2AP)TTCAG)2.  Drug binding constants, K
approx.10/5 M-1 were established between 5-25C.  A preliminary evaluation of
the thermodynamic data indicated a predominantly entropy driven interaction.

<>

<1>Patel, P.A., Kothari, V.V., Kothari, C.R., Faldu, P.R., Domadia, K.K., Rawal, C.M., Bhimani, H.D., Parmar, N.R., Nathani, N.M., Koringa, P.G., Joshi, C.G., Kothari, R.K.
<2>Draft Genome Sequence of Petroleum Hydrocarbon-Degrading Pseudomonas aeruginosa Strain PK6, Isolated from the Saurashtra Region of Gujarat, India.
<3>Genome Announcements
<4>2
<5>e00002-14
<6>2014
<7>Pseudomonas aeruginosa strain PK6, a potential petroleum hydrocarbon-degrading soil bacterium,
was isolated from a site contaminated by a petroleum hydrocarbon
spill from an automobile service station in Junagadh, Gujarat, India. Here, we
provide the 6.04-Mb draft genome sequence of strain PK6, which has genes encoding
enzymes for potential and related metabolic pathways of the strain.

<>

<1>Patel, P.H., Suzuki, M., Adman, E., Shinkai, A., Loeb, L.A.
<2>Prokaryotic DNA polymerase I: evolution, structure, and "base flipping" mechanism for nucleotide selection.
<3>J. Mol. Biol.
<4>308
<5>823-837
<6>2001
<7>Accurate transmission of DNA material from one generation to the next is crucial for prolonged
cell survival. Following the discovery of DNA polymerse I in Escherichia coli, the DNA
polymerase I class of enzymes has served as the prototype for studies on structural and
biochemical mechanisms of DNA replication. Recently, a series of genomic, mutagenesis and
structural investigations have provided key insights into how Pol I class of enzymes function
and evolve. X-ray crystal structures of at least three Pol I class of enzymes have been solved
in the presence of DNA and dNTP, thus allowing a detailed description of a productive
replication complex. Rapid-quench stop-flow studies have helped define individual steps during
nucleotide incorporation and conformational changes that are rate limiting during catalysis.
Studies in our laboratory have generated large libraries of active mutant enzymes (8000)
containing a variety of substitutions within the active site, some of which exhibit altered
biochemical properties. Extensive genomic information of Pol I has recently become available,
as over 50 polA genes from different prokaryotic species have been sequenced. In light of
these advancements, we review here the structure-function relationships of Pol I, and we
highlight those interactions that are responsible for the high fidelity of DNA synthesis. We
present a mechanism for "flipping" of the complementary template base to enhance interactions
with the incoming nucleotide substrate during DNA synthesis.

<>

<1>Patel, Y., Nelson, M., McClelland, M.
<2>Methylation at overlapping dam (Gm6ATC) sites does not block cleavage by the NruI (TCGCGA) isoschizomer restriction endonucleases AmaI, SalDI and Sbo13I.
<3>Nucleic Acids Res.
<4>17
<5>3613
<6>1989
<7>AmaI, NruI, SalDI and Sbo13I endonucleases were tested for sensitivity to TCGCGm6A at an
overlapping dam site in Ad2 DNA (pos 7723). NruI cleavage was blocked by m6A modification, but
the other endonucleases were not.

<>

<1>Patel, Y., Van Cott, E., Wilson, G.G., McClelland, M.
<2>Cleavage at the twelve-base-pair sequence 5'-TCTAGATCTAGA-3' using M.XbaI (TCTAGm6A) methylation and DpnI (Gm6A/TC) cleavage .
<3>Nucleic Acids Res.
<4>18
<5>1603-1607
<6>1990
<7>The DNA methylase M.XbaI was isolated from an E. coli recombinant clone.  We
deduce that the enzyme methylates at the sequence 5'-TCTAGm6A-3'.  In
combination with the methylation-dependent restriction endonuclease, DpnI
(5'-Gm6A/TC-3'), DNA cleavage occurs at the sequence 5'-TCTAGA/TCTAGA-3'.  This
twelve-base-pair site should occur once every 16,000,000 base pairs in a random
sequence of DNA.  The exceptional rarity of the M.XbaI/DpnI sequence makes it
an ideal candidate for transpositional integration of a unique cleavage site
into bacterial genomes.  Retrotransposition into mammalian genomes is also an
attractive possibility.

<>

<1>Patell, V., Rajyashri, K., Rodrigue, M., Vernet, G.
<2>Construction of a comparative database and identification of virulence factors through comparison of polymorphic regions in clinical isolates of infectious organisms.
<3>International Patent Office
<4>EP 1789577 A, WO 2006008575 A
<5>
<6>2006
<7>The present invention is directed to novel nucleotide sequences to be used for diagnosis,
identification of the strain, typing of the strain and giving orientation to its potential
degree of virulence, infectivity and/or latency for all infectious diseases more particularly
tuberculosis.  The present invention also includes method for the identification and selection
of polymorphisms associated with the virulence' and/or infectivity in infectious diseases
more particularly in tuberculosis by a comparative genomic analysis of the sequences of
different clinical isolates/strains of infectious organisms.  The regions of polymorphisms,
can also act as potential drug targets and vaccine targets.  More particularly, the invention
also relates to identifying virulence factors of M. tuberculosis strains and other infectious
organisms to be included in a diagnostic DNA chip allowing identification of the strain,
typing of the strain and finally giving orientation to its potential degree of virulence.
Although the present invention has been illustrated with specific reference to the polymorphic
region in the Mycobacterium tuberculosis, the said invention is not to be understood and
constructed as being limited to Tuberculosis but is applicable to all infectious diseases.

<>

<1>Pathak, A., Green, S.J., Ogram, A., Chauhan, A.
<2>Draft Genome Sequence of Rhodococcus opacus Strain M213 Shows a Diverse Catabolic Potential.
<3>Genome Announcements
<4>1
<5>e00144-12
<6>2013
<7>Soil-borne Gram-positive bacteria from the genus Rhodococcus metabolize a range of aromatic
hydrocarbons and also produce a variety of value-added products, such
as triacylglycerols and steroids. We report the draft genome sequence of
Rhodococcus opacus strain M213 (9,193,504 bp with a G+C content of 66.99%),
providing a comprehensive understanding of the repertoire of metabolic genes of
this strain.

<>

<1>Pati, A. et al.
<2>Complete genome sequence of Bacteroides helcogenes type strain (P 36-108).
<3>Standards in Genomic Sciences
<4>4
<5>45-53
<6>2011
<7>Bacteroides helcogenes Benno et al. 1983 is of interest because of its isolated phylogenetic
location and, although it has been found in pig feces and is known
to be pathogenic for pigs, occurrence of this bacterium is rare and it does not
cause significant damage in intensive animal husbandry. The genome of B.
helcogenes P 36-108(T) is already the fifth completed and published type strain
genome from the genus Bacteroides in the family Bacteroidaceae. The 3,998,906 bp
long genome with its 3,353 protein-coding and 83 RNA genes consists of one
circular chromosome and is a part of the Genomic Encyclopedia of Bacteria and
Archaea project.

<>

<1>Pati, A. et al.
<2>Non-contiguous finished genome sequence of the opportunistic oral pathogen Prevotella multisaccharivorax type strain (PPPA20).
<3>Standards in Genomic Sciences
<4>5
<5>41-49
<6>2011
<7>Prevotella multisaccharivorax Sakamoto et al. 2005 is a species of the large genus Prevotella,
which belongs to the family Prevotellaceae. The species is of
medical interest because its members are able to cause diseases in the human oral
cavity such as periodontitis, root caries and others. Although 77 Prevotella
genomes have already been sequenced or are targeted for sequencing, this is only
the second completed genome sequence of a type strain of a species within the
genus Prevotella to be published. The 3,388,644 bp long genome is assembled in
three non-contiguous contigs, harbors 2,876 protein-coding and 75 RNA genes and
is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Pati, A. et al.
<2>Complete genome sequence of Oceanithermus profundus type strain (506).
<3>Standards in Genomic Sciences
<4>4
<5>210-220
<6>2011
<7>Oceanithermus profundus Miroshnichenko et al. 2003 is the type species of the genus
Oceanithermus, which belongs to the family Thermaceae. The genus currently
comprises two species whose members are thermophilic and are able to reduce
sulfur compounds and nitrite. The organism is adapted to the salinity of sea
water, is able to utilize a broad range of carbohydrates, some proteinaceous
substrates, organic acids and alcohols. This is the first completed genome
sequence of a member of the genus Oceanithermus and the fourth sequence from the
family Thermaceae. The 2,439,291 bp long genome with its 2,391 protein-coding and
54 RNA genes consists of one chromosome and a 135,351 bp long plasmid, and is a
part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Pati, A. et al.
<2>Complete genome sequence of Cellulophaga lytica type strain (LIM-21).
<3>Standards in Genomic Sciences
<4>4
<5>221-232
<6>2011
<7>Cellulophaga lytica (Lewin 1969) Johansen et al. 1999 is the type species of the  genus
Cellulophaga, which belongs to the family Flavobacteriaceae within the
phylum 'Bacteroidetes' and was isolated from marine beach mud in Limon, Costa
Rica. The species is of biotechnological interest because its members produce a
wide range of extracellular enzymes capable of degrading proteins and
polysaccharides. After the genome sequence of Cellulophaga algicola this is the
second completed genome sequence of a member of the genus Cellulophaga. The
3,765,936 bp long genome with its 3,303 protein-coding and 55 RNA genes consists
of one circular chromosome and is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Pati, A. et al.
<2>Complete genome sequence of Saccharomonospora viridis type strain (P101).
<3>Standards in Genomic Sciences
<4>1
<5>141-149
<6>2009
<7>Saccharomonospora viridis (Schuurmans et al. 1956) Nonomurea and Ohara 1971 is the type
species of the genus Saccharomonospora which belongs to the family
Pseudonocardiaceae. S. viridis is of interest because it is a Gram-negative
organism classified among the usually Gram-positive actinomycetes. Members of the
species are frequently found in hot compost and hay, and its spores can cause
farmer's lung disease, bagassosis, and humidifier fever. Strains of the species
S. viridis have been found to metabolize the xenobiotic pentachlorophenol (PCP).
The strain described in this study has been isolated from peat-bog in Ireland.
Here we describe the features of this organism, together with the complete genome
sequence, and annotation. This is the first complete genome sequence of the
family Pseudonocardiaceae, and the 4,308,349 bp long single replicon genome with
its 3906 protein-coding and 64 RNA genes is part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Pati, A. et al.
<2>Complete genome sequence of Arcobacter nitrofigilis type strain (CI).
<3>Standards in Genomic Sciences
<4>2
<5>300-308
<6>2010
<7>Arcobacter nitrofigilis (McClung et al. 1983) Vandamme et al. 1991 is the type species of the
genus Arcobacter in the family Campylobacteraceae within the
Epsilonproteobacteria. The species was first described in 1983 as Campylobacter
nitrofigilis [1] after its detection as a free-living, nitrogen-fixing
Campylobacter species associated with Spartina alterniflora Loisel roots [2]. It
is of phylogenetic interest because of its lifestyle as a symbiotic organism in a
marine environment in contrast to many other Arcobacter species which are
associated with warm-blooded animals and tend to be pathogenic. Here we describe
the features of this organism, together with the complete genome sequence, and
annotation. This is the first complete genome sequence of a type stain of the
genus Arcobacter. The 3,192,235 bp genome with its 3,154 protein-coding and 70
RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Pati, A. et al.
<2>Complete genome sequence of Sphaerobacter thermophilus type strain (S 6022).
<3>Standards in Genomic Sciences
<4>2
<5>49-56
<6>2010
<7>Sphaerobacter thermophilus Demharter et al. 1989 is the sole and type species of  the genus
Sphaerobacter, which is the type genus of the family
Sphaerobacteraceae, the order Sphaerobacterales and the subclass
Sphaerobacteridae. Phylogenetically, it belongs to the genomically little studied
class of the Thermomicrobia in the bacterial phylum Chloroflexi. Here, the genome
of strain S 6022(T) is described which is an obligate aerobe that was originally
isolated from an aerated laboratory-scale fermentor that was pulse fed with
municipal sewage sludge. We describe the features of this organism, together with
the complete genome and annotation. This is the first complete genome sequence of
the thermomicrobial subclass Sphaerobacteridae, and the second sequence from the
chloroflexal class Thermomicrobia. The 3,993,764 bp genome with its 3,525
protein-coding and 57 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Pati, A. et al.
<2>Complete genome sequence of Brachyspira murdochii type strain (56-150).
<3>Standards in Genomic Sciences
<4>2
<5>260-269
<6>2010
<7>Brachyspira murdochii Stanton et al. 1992 is a non-pathogenic, host-associated spirochete of
the family Brachyspiraceae. Initially isolated from the intestinal
content of a healthy swine, the 'group B spirochaetes' were first described as
Serpulina murdochii. Members of the family Brachyspiraceae are of great
phylogenetic interest because of the extremely isolated location of this family
within the phylum 'Spirochaetes'. Here we describe the features of this organism,
together with the complete genome sequence and annotation. This is the first
completed genome sequence of a type strain of a member of the family
Brachyspiraceae and only the second genome sequence from a member of the genus
Brachyspira. The 3,241,804 bp long genome with its 2,893 protein-coding and 40
RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Patrick, S., Blakely, G.W., Houston, S., Moore, J., Abratt, V.R., Bertalan, M., Cerdeno-Tarraga, A.M., Quail, M.A., Corton, N., Corton, C., Bignell, A., Barron, A., Clark, L., Bentley, S.D., Parkhill, J.
<2>Twenty-eight divergent polysaccharide loci specifying within- and amongst-strain capsule diversity in three strains of Bacteroides fragilis.
<3>Microbiology
<4>156
<5>3255-3269
<6>2010
<7>Comparison of the complete genome sequence of Bacteroides fragilis 638R, originally isolated
in the USA, was made with two previously
sequenced strains isolated in the UK (NCTC 9343) and Japan (YCH46). The
presence of 10 loci containing genes associated with polysaccharide
(PS) biosynthesis, each including a putative Wzx flippase and Wzy
polymerase, was confirmed in all three strains, despite a lack of
cross-reactivity between NCTC 9343 and 638R surface PS-specific
antibodies by immunolabelling and microscopy. Genomic comparisons
revealed an exceptional level of PS biosynthesis locus diversity. Of
the 10 divergent PS-associated loci apparent in each strain, none is
similar between NCTC 9343 and 638R. YCH46 shares one locus with NCTC
9343, confirmed by mAb labelling, and a second different locus with
638R, making a total of 28 divergent PS biosynthesis loci amongst the
three strains. The lack of expression of the phase-variable large
capsule (LC) in strain 638R, observed in NCTC 9343, is likely to be due
to a point mutation that generates a stop codon within a putative
initiating glycosyltransferase, necessary for the expression of the LC
in NCTC 9343. Other major sequence differences were observed to arise
from different numbers and variety of inserted extra-chromosomal
elements, in particular prophages. Extensive horizontal gene transfer
has occurred within these strains, despite the presence of a
significant number of divergent DNA restriction and modification
systems that act to prevent acquisition of foreign DNA. The level of
amongst-strain diversity in PS biosynthesis loci is unprecedented.

<>

<1>Pattabiraman, V., Bopp, C.A.
<2>Draft Whole-Genome Sequences of 10 Serogroup O6 Enterotoxigenic Escherichia coli  Strains.
<3>Genome Announcements
<4>2
<5>e01274-14
<6>2014
<7>Entertotoxigenic Escherichia coli (ETEC) is a major cause of global diarrhea, resulting in
approximately 200 million occurrences and 300,000 to 400,000 deaths
annually, primarily in children under the age of five. Here, we announce the
release of the draft genomes of 10 ETEC isolates belonging to serogroup O6.

<>

<1>Patterson, N.H., Pauling, C.
<2>Evidence for two restriction-modification systems in Halobacterium cutirubrum.
<3>J. Bacteriol.
<4>163
<5>783-784
<6>1985
<7>Data from plating experiments indicated that Halobacterium cutirubrum NRC34001
has at least two separate restriction-modifiction systems.  A spontaneous or
induced loss of one or both systems resulted in four restriction-modification
phenotypes.  There was a positive correlation between changes in gas
vacuolation phenotypes and either restriction-modifiction system.

<>

<1>Paul, D., Austin, F.W., Arick, T., Bridges, S.M., Burgess, S.C., Dandass, Y.S., Lawrence, M.L.
<2>Genome sequence of the solvent-producing bacterium Clostridium carboxidivorans strain P7T.
<3>J. Bacteriol.
<4>192
<5>5554-5555
<6>2010
<7>Clostridium carboxidivorans strain P7(T) is a strictly anaerobic acetogenic bacterium that
produces acetate, ethanol, butanol, and
butyrate. The C. carboxidivorans genome contains all the genes for the
carbonyl branch of the Wood-Ljungdahl pathway for CO(2) fixation, and it
encodes enzymes for conversion of acetyl coenzyme A into butanol and
butyrate.

<>

<1>Paul, D., Bridges, S., Burgess, S.C., Dandass, Y., Lawrence, M.L.
<2>Genome sequence of the chemolithoautotrophic bacterium Oligotropha carboxidovorans OM5T.
<3>J. Bacteriol.
<4>190
<5>5531-5532
<6>2008
<7>Oligotropha carboxidovorans OM5(T) (DSM 1227, ATCC 49405) is a
chemolithoautotrophic bacterium with the capability to utilize carbon
monoxide, carbon dioxide, and hydrogen. It is also capable of
heterotrophic growth under appropriate environmental conditions. Here we
report the annotated genome sequence of the circular chromosome of this
organism.

<>

<1>Paul, R., Jinkerson, R.E., Buss, K., Steel, J., Mohr, R., Hess, W.R., Chen, M., Fromme, P.
<2>Draft Genome Sequence of the Filamentous Cyanobacterium Leptolyngbya sp. Strain Heron Island J, Exhibiting Chromatic Acclimation.
<3>Genome Announcements
<4>2
<5>e01166-13
<6>2014
<7>Leptolyngbya sp. strain Heron Island is a cyanobacterium exhibiting chromatic acclimation.
However, this strain has strong interactions with other bacteria,
making it impossible to obtain axenic cultures for sequencing. A protocol
involving an analysis of tetranucleotide frequencies, G+C content, and BLAST
searches has been described for separating the cyanobacterial scaffolds from
those of its cooccurring bacteria.

<>

<1>Paul, W., Wehrkamp-Richter, S., Laffaire, J.-B.
<2>Modified restriction enzymes for stimulation of homologous recombination in plants.
<3>International Patent Office
<4>WO 2007135022 A
<5>
<6>2007
<7>The invention relates to modified restriction enzymes capable of being used for promoting
homologous recombination in organisms, in particular plants, making it possible to either
target gene integration or excise unwantd DNA sequences in the genome of said organisms.

<>

<1>Paulsen, I. et al.
<2>The Brucella suis genome reveals fundamental similarities between animal and plant pathogens and symbionts.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>13148-13153
<6>2002
<7>The 3.31-Mb genome sequence of the intracellular pathogen and potential bioterrorism agent,
Brucella suis, was determined. Comparison of B. suis with Brucella melitensis has defined a
finite set of differences that could be responsible for the differences in virulence and host
preference between these organisms, and indicates that phage have played a significant role in
their divergence. Analysis of the B. suis genome reveals transport and metabolic capabilities
akin to soil/plant-associated bacteria. Extensive gene synteny between B. suis chromosome 1
and the genome of the plant symbiont Mesorhizobium loti emphasizes the similarity between this
animal pathogen and plant pathogens and symbionts. A limited repertoire of genes homologous to
known bacterial virulence factors were identified.

<>

<1>Paulsen, I.T. et al.
<2>Role of mobile DNA in the evolution of vancomycin-resistant Enterococcus faecalis.
<3>Science
<4>299
<5>2071-2074
<6>2003
<7>The complete genome sequence of Enterococcus faecalis V583, a vancomycin-resistant clinical
isolate, revealed that more than a quarter
of the genome consists of probable mobile or foreign DNA. One of the
predicted mobile elements is a previously unknown vanB
vancomycin-resistance conjugative transposon. Three plasmids were
identified, including two pheromone-sensing conjugative plasmids, one
encoding a previously undescribed pheromone inhibitor. The apparent
propensity for the incorporation of mobile elements probably contributed
to the rapid acquisition and dissemination of drug resistance in the
enterococci.

<>

<1>Paulsen, I.T. et al.
<2>Complete genome sequence of the plant commensal Pseudomonas fluorescens Pf-5.
<3>Nat. Biotechnol.
<4>23
<5>873-878
<6>2005
<7>Pseudomonas fluorescens Pf-5 is a plant commensal bacterium that inhabits the rhizosphere and
produces secondary metabolites that suppress soilborne
plant pathogens. The complete sequence of the 7.1-Mb Pf-5 genome was
determined. We analyzed repeat sequences to identify genomic islands that,
together with other approaches, suggested P. fluorescens Pf-5's recent
lateral acquisitions include six secondary metabolite gene clusters, seven
phage regions and a mobile genomic island. We identified various features
that contribute to its commensal lifestyle on plants, including broad
catabolic and transport capabilities for utilizing plant-derived
compounds, the apparent ability to use a diversity of iron siderophores,
detoxification systems to protect from oxidative stress, and the lack of a
type III secretion system and toxins found in related pathogens. In
addition to six known secondary metabolites produced by P. fluorescens
Pf-5, three novel secondary metabolite biosynthesis gene clusters were
also identified that may contribute to the biocontrol properties of P.
fluorescens Pf-5.

<>

<1>Pavan, M.E., Pavan, E.E., Lopez, N.I., Levin, L., Pettinari, M.J.
<2>Genome Sequence of the Melanin-Producing Extremophile Aeromonas salmonicida subsp. pectinolytica Strain 34melT.
<3>Genome Announcements
<4>1
<5>e00675-13
<6>2013
<7>The genome of Aeromonas salmonicida subsp. pectinolytica strain 34mel(T), isolated from a
heavily polluted river, contains several genomic islands and
putative virulence genes. The identification of genes involved in resistance to
different kinds of stress sheds light on the mechanisms used by this strain to
thrive in an extreme environment.

<>

<1>Pavco, P.A., Steege, D.A.
<2>A DNA binding protein acts as a transcriptional roadblock.
<3>J. Cell Biochem. Suppl.
<4>12D
<5>145
<6>1988
<7>What results when an elongating RNA polymerase encounters DNA-bound protein is
of general interest.  To ask if a protein bound tightly to a specific sequence
blocks further elongation by E. coli RNA polymerase, transcription on templates
associated with a mutant of the EcoRI endonuclease has been studied in vitro.
This protein (E111G) binds to the wild type recognition sequence with high
affinity yet carries out no appreciable cleavage.  When a DNA template
containing one EcoRI site is transcribed in the presence of E111G and
rifampicin, two RNA products are seen:  a full-length runoff transcript and a
truncated RNA species whose 3' endpoint is immediately upstream of the EcoRI
recognition sequence.  Under conditions of complete binding by E111G, the
shorter transcript is the major RNA species appearing.  Blockage appears
long-lived and not dependent on the DNA sequence context upstream of the EcoRI
site.  During steady state transcription an additional RNA 3' endpoint, due to
a second RNA polymerase stalled behind the first, is seen.  From analysis of
the RNA 3' ends, the position of the 3' terminal ribonucleotide with respect to
the leading edge of the RNA polymerase ternary complex has been located.

<>

<1>Pavlopoulou, A., Kossida, S.
<2>Plant cytosine-5 DNA methyltransferases: Structure, function, and molecular evolution.
<3>Genomics
<4>90
<5>530-541
<6>2007
<7>A detailed analysis of the structure and function, along with evolutionary aspects, of the
main plant cytosine-5 DNA
methyltransferases (C5-MTases) is presented. The evolutionary
relationships between the already known and four candidate plant
C5-MTases identified in this work were investigated using the distance,
maximum-parsimony, and maximum-likelihood approaches. The topologies of
the trees were overall congruent: four monophyletic groups
corresponding to the four plant C5-MTase families were clearly
distinguished. In addition, sequence analyses of the plant C5-MTase
target recognition domain sequences were performed and phylogenetic
trees were reconstructed showing that there is good conservation among
but not within the plant C5-MTase families. Furthermore, a conserved
dipeptide that plays an important role in flipping the target base into
the catalytic site of the C5-MTases was identified in all plant
C5-MTases under study.

<>

<1>Pavlopoulou, A., Kossida, S.
<2>Phylogenetic analysis of the eukaryotic RNA (cytosine-5)-methyltransferases.
<3>Genomics
<4>93
<5>350-357
<6>2009
<7>RNA (cytosine-5)-methyltransferases (RCMTs) have been characterized both in prokaryotic and
eukaryotic organisms. The RCMT family, however, remains largely uncharacterized, as opposed to
the family of DNA (cytosine-5)-methyltransferases which has been studied in depth. In the
present study, an in silico identification of the putative 5-methylcytosine RNA-generating
enzymes in the eukaryotic genomes was performed. A comprehensive phylogenetic analysis of the
putative eukaryotic RCMT-related proteins has been performed in order to redefine subfamilies
within the RCMT family. Five distinct eukaryotic subfamilies were identified, including the
three already known (NOP2, NCL1 and YNL022c), one novel subfamily (RCMT9) and a fifth one
which hitherto was considered to exist exclusively in prokaryotes (Fmu). The potential
evolutionary relationships among the different eukaryotic RCMT subfamilies were also
investigated.  Furthermore, the results of this study add further support to a previous
hypothesis that RCMTs represent evolutionary intermediates of RNA
(uridine-5)-methyltransferases and DNA (cytosine-5)-methyltransferases.

<>

<1>Pavlovic, G., Burrus, V., Gintz, B., Decaris, B., Guedon, G.
<2>Evolution of genomic islands by deletion and tandem accretion by site-specific recombination: ICESt1-related elements from Streptococcus thermophilus.
<3>Microbiology
<4>150
<5>759-774
<6>2004
<7>The 34 734-bp integrative and potentially conjugative element (putative ICE)
ICESt1 has been previously found to be site-specifically integrated in the 3' end
of the fda locus of Streptococcus thermophilus CNRZ368. Four types of genomic
islands related to ICESt1 are integrated in the same position in seven other
strains of S. thermophilus. One of these elements, ICESt3, harbours conjugation
and recombination modules closely related to those of ICESt1 and excises by
site-specific recombination. Two other types of elements, CIME19258 and CIME302,
are flanked by site-specific attachment sites closely related to attL and attR of
ICESt1 and ICESt3, whereas Delta CIME308 only possesses a putative attR site;
none of these three elements carry complete conjugation and recombination
modules. ICESt1 contains a functional internal recombination site, attL', that is
almost identical to attL of CIME19258. The recombination between attL' and attR
of ICESt1 leads to the excision of the expected circular molecule (putative ICE);
a cis-mobilizable element (CIME) flanked by an attL site and an attB' site
remains integrated into the 3' end of fda. Furthermore, sequences that could be
truncated att sites were found within ICESt1, ICESt3 and CIME302. All together,
these data suggest that these genomic islands evolved by deletion and tandem
accretion of ICEs and CIMEs resulting from site-specific recombination. A model
for this evolution is proposed and its application to other genomic islands is
discussed.

<>

<1>Pavlovic, G., Burrus, V., Toulmay, A., Choulet, F., Decaris, B., Guedon, G.
<2>Characterization and evolution of a family of integrative and potentially conjugative or mobilizable elements from Streptococcus  thermophilus.
<3>Lait
<4>84
<5>7-14
<6>2004
<7>The integrative and conjugative elements (ICEs) excise by site-specific recombination.
self-transfer the resulting circular form by conjugation
and integrate into the genome of the recipient bacterium. The 34.7-kb
element from Streptococcus thermophilus CNRZ368, ICEStl, excises and
integrates by site-specific recombination. This element also possesses
a conjugation module distantly related to that of the conjugative
transposon Tn9l6 from Enterococcus faecalis. Therefore, ICEStl is
probably an ICE. Four types of elements related to ICEStl are
integrated into the same location as ICEStl in seven other strains of
S. thermophilus. One of these elements, ICESt3, is probably an ICE
whereas the three others (CIMEs) would have arisen from ICEs by
deletion of the conjugation and recombination modules. These elements
also encode functions that are not involved in element maintenance or
transfer, Such as restriction-modification systems. Each of these
elements has a chimerical Structure resulting from the acquisition of
modules from different origins. Sequence analyses indicate that these
elements are involved in horizontal transfers with various species of
dairy or pathogenic lactic acid bacteria. DeltaCIME308 has exchanged
restriction-modification and cadmium resistance modules with plasmids.
CIME19258 has acquired a cadmium resistance module by the integration
of an ICE related to Tn9l6 within the CIME. The site-specific
recombination between an internal anL-related site and attR of ICEStl
leads to the excision of a circular molecule which could be another
ICE, ICEt2, suggesting that ICEStl has arisen by accretion of a CIME
and ICEt2. CIME302, ICESt2 and ICESt3 would also have arisen by
site-specific accretion of CIMEs and ICEs and/or mobilization of CIMEs
by ICEs. An ICE would integrate by site-specific recombination in the
attR site of a CIME; then the CIME-ICE would excise by site-specific
recombination and transfer by conjugation.

<>

<1>Pawar, S.P., Dhotre, D.P., Shetty, S.A., Chowdhury, S.P., Chaudhari, B.L., Shouche, Y.S.
<2>Genome Sequence of Janibacter hoylei MTCC8307, Isolated from the Stratospheric Air.
<3>J. Bacteriol.
<4>194
<5>6629-6630
<6>2012
<7>Janibacter hoylei MTCC8307 was isolated from stratospheric air at an altitude of  41.4 km over
Hyderabad, India. Here, we present the draft genome of Janibacter
hoylei MTCC8307, which contains 3,139,099 bp with a G+C content of 72.8 mol%,
2,972 protein-coding genes, and 57 structural RNAs.

<>

<1>Pawlak, S.D., Radlinska, M., Chmiel, A.A., Bujnicki, J.M., Skowronek, K.J.
<2>Inference of relationships in the 'twilight zone' of homology using a combination of bioinformatics and site-directed mutagenesis: a case study   of restriction endonucleases Bsp6I and PvuII.
<3>Nucleic Acids Res.
<4>33
<5>661-671
<6>2005
<7>Thus far, identification of functionally important residues in Type II restriction
endonucleases (REases) has been difficult using conventional
methods. Even though known REase structures share a fold and marginally
recognizable active site, the overall sequence similarities are
statistically insignificant, unless compared among proteins that recognize
identical or very similar sequences. Bsp6I is a Type II REase, which
recognizes the palindromic DNA sequence 5'GCNGC and cleaves between the
cytosine and the unspecified nucleotide in both strands, generating a
double-strand break with 5'-protruding single nucleotides. There are no
solved structures of REases that recognize similar DNA targets or generate
cleavage products with similar characteristics. In straightforward
comparisons, the Bsp6I sequence shows no significant similarity to REases
with known structures. However, using a fold-recognition approach, we have
identified a remote relationship between Bsp6I and the structure of PvuII.
Starting from the sequence-structure alignment between Bsp6I and PvuII, we
constructed a homology model of Bsp6I and used it to predict functionally
significant regions in Bsp6I. The homology model was supported by
site-directed mutagenesis of residues predicted to be important for
dimerization, DNA binding and catalysis. Completing the picture of
sequence-structure-function relationships in protein superfamilies becomes
an essential task in the age of structural genomics and our study may
serve as a paradigm for future analyses of superfamilies comprising
strongly diverged members with little or no sequence similarity.

<>

<1>Pawlak, S.D., Skowronek, K., Radlinska, M., Bujnicki, J.M.
<2>Fold-recognition, homology modeling and mutagenesis of restriction enzyme Bsp6I.
<3>FEBS J.
<4>272
<5>95
<6>2005
<7>Identification of functionally important residues in type II restriction enzymes (REases) has
been difficult using conventional methods. Even though known REase structures share a common
fold and marginally recognizable active site, the overall sequence similarities are
statistically insignificant, unless compared among proteins that recognize identical or very
similar sequences. Bsp6I is a Type II REase, which recognizes the palindromic DNA sequence
5'GCNGC and cleaves between the cytosine and the unspecified nucleotide in both strands,
generating a double strand break with 5'-protruding single nucleotides. There are no solved
structures of REases that recognize similar DNA targets or generate cleavage products with
similar characteristics. The Bsp6I sequence shows no significant similarity to REases with
known structures. However, using a protein fold-recognition approach, we have identified a
remote relationship between Bsp6I and the structure of PvuII, which allowed us to construct a
homology model of Bsp6I and use it to predict functionally important regions and residues in
Bsp6I. The model of the Bsp6I structure was built using the "Frankenstein's monster" method
and tested by the characterization of the effects of single amino acid substitutions of
residues predicted to be directly involved in involved in cleavage, DNA-binding and
dimerization. The endonuclease activity of an extensive panel of mutants was tested in vivo
using the bacteriophage lambda-plating assay. All mutations in residues predicted as
catalytic, involved in DNA binding dimerization decreased the restriction level to less than
1% of the wild type (wt) activity. A subset of mutants exhibiting different levels of
reduction of the in vivo activity was recloned into an expression vector, overexpressed,
purified and tested in an in vitro cleavage assay. The results agreed with the in vivo
analyses, thus corroborating the model-based predictions. Our study represents an example of
how the computational protein fold-recognition followed by model-based identification and
experimental validation of functionally important residues can be used to reduce the "white
spaces" on the structural map of a protein superfamily by providing links between known
structures and the sequences of their remote homologs. Confident identification of a protein
fold, which is very difficult in the case of restriction enzymes, is important for the
selection of targets for high-resolution studies. Completing the picture of
sequence-structure-function relationships in protein superfamilies becomes an essential task
in the age of structural genomics and our study may serve as a paradigm for future analyses.

<>

<1>Peakman, L.J., Antognozzi, M., Bickle, T.A., Janscak, P., Szczelkun, M.D.
<2>S-Adenosyl Methionine Prevents Promiscuous DNA Cleavage by the EcoP1I type III Restriction Enzyme.
<3>J. Mol. Biol.
<4>333
<5>321-335
<6>2003
<7>DNA cleavage by the type III restriction endonuclease EcoP1I was analysed on circular and
catenane DNA in a variety of buffers with different salts.
In the presence of the cofactor S-adenosyl methionine (AdoMet), and
irrespective of buffer, only substrates with two EcoP1I sites in inverted
repeat were susceptible to cleavage. Maximal activity was achieved at a
Res2Mod2 to site ratio of approximately 1:1 yet resulted in cleavage at
only one of the two sites. In contrast, the outcome of reactions in the
absence of AdoMet was dependent upon the identity of the monovalent buffer
components, in particular the identity of the cation. With Na+, cleavage
was observed only on substrates with two sites in inverted repeat at
elevated enzyme to site ratios (>15:1). However, with K+ every substrate
tested was susceptible to cleavage above an enzyme to site ratio of
approximately 3:1, including a DNA molecule with two directly repeated
sites and even a DNA molecule with a single site. Above an enzyme to site
ratio of 2:1, substrates with two sites in inverted repeat were cleaved at
both cognate sites. The rates of cleavage suggested two separate events: a
fast primary reaction for the first cleavage of a pair of inverted sites;
and an order-of-magnitude slower secondary reaction for the second
cleavage of the pair or for the first cleavage of all other site
combinations. EcoP1I enzymes mutated in either the ATPase or nuclease
motifs did not produce the secondary cleavage reactions. Thus, AdoMet
appears to play a dual role in type III endonuclease reactions: Firstly,
as an allosteric activator, promoting DNA association; and secondly, as a
"specificity factor", ensuring that cleavage occurs only when two
endonucleases bind two recognition sites in a designated orientation.
However, given the right conditions, AdoMet is not strictly required for
DNA cleavage by a type III enzyme.

<>

<1>Peakman, L.J., Szczelkun, M.D.
<2>DNA communications by Type III restriction endonucleases - confirmation of 1D translocation over 3D looping.
<3>Nucleic Acids Res.
<4>32
<5>4166-4174
<6>2004
<7>DNA cleavage by Type III restriction enzymes is governed strictly by the relative arrangement
of recognition sites on a DNA substrate.  Xendonuclease activity is usually only triggered by
sequences in head-to-head orientation. Tens to thousands of base pairs can separate these
sites. Long distance communication over such distances could occur by either one-dimensional
(1D) DNA translocation or 3D DNA looping. To distinguish between these alternatives, we
analysed the activity of EcoPI and EcoP15I on DNA catenanes in which the recognition sites
were either on the same or separate rings. While substrates with a pair of sites located on
the same ring were cleaved efficiently, catenanes with sites on separate rings were not
cleaved. These results exclude a simple 3D DNA-looping activity. To characterize the
interactions further, EcoPI was incubated with plasmids carrying two recognition sites
interspersed with two 21res sites for site-specific recombination by Tn21 resolvase;
inhibition of recombination would indicate the formation of stable DNA loops. No inhibition
was observed, even under conditions where EcoPI translocation could also occur.

<>

<1>Peakman, L.J., Szczelkun, M.D.
<2>S-Adenosyl homocysteine and DNA ends stimulate promiscuous nuclease activities in the Type III restriction endonuclease EcoPI.
<3>Nucleic Acids Res.
<4>37
<5>3934-3945
<6>2009
<7>In the absence of the methyl donor S-adenosyl methionine and under certain permissive reaction
conditions, EcoPI shows non-specific endonuclease
activity. We show here that the cofactor analogue S-adenosyl homocysteine
promotes this promiscuous DNA cleavage. Additionally, an extensive
exonuclease-like processing of the DNA is also observed that can even
result in digestion of non-specific DNA in trans. We suggest a model for
how DNA communication events initiating from non-specific sites, and in
particular free DNA ends, could produce the observed cleavage patterns.

<>

<1>Pearce, S.L., Pandey, R., Dorrian, S.J., Russell, R.J., Oakeshott, J.G., Pandey, G.
<2>Genome sequence of the newly isolated chemolithoautotrophic Bradyrhizobiaceae strain SG-6C.
<3>J. Bacteriol.
<4>193
<5>5057
<6>2011
<7>Strain SG-6C (DSM 23264, CCM 7827) is a chemolithoautotrophic bacterium, of the family
Bradyrhizobiaceae. It can also grow heterotrophically under
appropriate environmental conditions. Here we report the annotated genome
sequence of this strain in a single 4.3-Mb circular scaffold.

<>

<1>Pearce, S.L., Pushiri, H., Oakeshott, J.G., Russell, R.J., Pandey, G.
<2>Draft Genome Sequence of Ralstonia sp. Strain GA3-3, Isolated from Australian Suburban Soil.
<3>Genome Announcements
<4>1
<5>e00414-13
<6>2013
<7>Ralstonia sp. strain GA3-3 is a hexachlorocyclohexane (HCH)-degrading bacterial strain
isolated from suburban soil in Canberra, Australia. The genome of strain
GA3-3 was sequenced to investigate its ability to degrade alpha-HCH. Here, we
report the annotated genome sequence of this strain.

<>

<1>Pearson, B.M., Gaskin, D.J., Segers, R.P., Wells, J.M., Nuijten, P.J., Mvan, V.A.H.
<2>The Complete Genome Sequence of Campylobacter jejuni Strain 81116 (NCTC11828).
<3>J. Bacteriol.
<4>189
<5>8402-8403
<6>2007
<7>Campylobacter jejuni is a major human enteric pathogen that displays genetic variability via
genomic reorganization and phase variation. This
variability can adversely affect the outcomes and reproducibility of
experiments. C. jejuni strain 81116 (NCTC11828) has been suggested to be a
genetically stable strain (G. Manning, B. Duim, T. Wassenaar, J. A.
Wagenaar, A. Ridley, and D. G. Newell, Appl. Environ. Microbiol.
67:1185-1189, 2001), is amenable to genetic manipulation, and is infective
for chickens. Here we report the finished annotated genome sequence of C.
jejuni strain 81116.

<>

<1>Pearson, B.M., Rokney, A., Crossman, L.C., Miller, W.G., Wain, J., van Vliet, A.H.
<2>Complete Genome Sequence of the Campylobacter coli Clinical Isolate 15-537360.
<3>Genome Announcements
<4>1
<5>e01056-13
<6>2013
<7>Campylobacter coli strain 15-537360 was originally isolated in 2001 from a 42-year-old patient
with gastroenteritis. Here, we report its complete genome
sequence, which comprises a 1.7-Mbp chromosome and a 29-kbp conjugative cryptic
plasmid. This is the first complete genome sequence of a clinical isolate of C.
coli.

<>

<1>Pearson, M.D., Noller, H.F.
<2>The Draft Genome of Planococcus donghaensis MPA1U2 Reveals Nonsporulation Pathways Controlled by a Conserved Spo0A Regulon.
<3>J. Bacteriol.
<4>193
<5>6106
<6>2011
<7>The Planococcaceae are extreme survivors, having been cultured from environments such as deep
sea sediments, marine solar salterns, glaciers,
permafrost, Antarctic deserts, and sea ice brine. The family contains both
sporulating and nonsporulating genera. Here we present the unclosed, draft
genome sequence of Planococcus donghaensis strain MPA1U2, a nonsporulating
psychrotrophic bacterium isolated from surface coastal water of the
Pacific Ocean.

<>

<1>Pearson, M.M. et al.
<2>Complete genome sequence of uropathogenic Proteus mirabilis, a master of both adherence and motility.
<3>J. Bacteriol.
<4>190
<5>4027-4037
<6>2008
<7>The gram-negative enteric bacterium Proteus mirabilis is a frequent cause of urinary tract
infections in individuals with long-term indwelling
catheters or with complicated urinary tracts (e.g., due to spinal cord
injury or anatomic abnormality). P. mirabilis bacteriuria may lead to
acute pyelonephritis, fever, and bacteremia. Most notoriously, this
pathogen uses urease to catalyze the formation of kidney and bladder
stones or to encrust or obstruct indwelling urinary catheters. Here we
report the complete genome sequence of P. mirabilis HI4320, a
representative strain cultured in our laboratory from the urine of a
nursing home patient with a long-term (> or =30 days) indwelling urinary
catheter. The genome is 4.063 Mb long and has a G+C content of 38.88%.
There is a single plasmid consisting of 36,289 nucleotides. Annotation of
the genome identified 3,685 coding sequences and seven rRNA loci. Analysis
of the sequence confirmed the presence of previously identified virulence
determinants, as well as a contiguous 54-kb flagellar regulon and 17 types
of fimbriae. Genes encoding a potential type III secretion system were
identified on a low-G+C-content genomic island containing 24 intact genes
that appear to encode all components necessary to assemble a type III
secretion system needle complex. In addition, the P. mirabilis HI4320
genome possesses four tandem copies of the zapE metalloprotease gene,
genes encoding six putative autotransporters, an extension of the atf
fimbrial operon to six genes, including an mrpJ homolog, and genes
encoding at least five iron uptake mechanisms, two potential type IV
secretion systems, and 16 two-component regulators.

<>

<1>Pech, M., Streeck, R.E., Zachau, H.G.
<2>Patchwork structure of a bovine satellite DNA.
<3>Cell
<4>18
<5>883-893
<6>1979
<7>According to a previous restriction nuclease analysis, bovine 1.706 satellite
DNA (density 1.706 g/cm3 in CsCl) is organized in an unusual structure of
superimposed long- and short-range repeats (Streeck and Zachau, 1978).  We have
now determined the nucleotide sequence of this satellite DNA in both cloned
fragments and fragments from the total satellite DNA.  Each long-range repeat
unit (about 2350 bp) is divided into four segments.  Each segment consists of
different variants of a basic 23 bp sequence which is itself composed of a
dodecanucleotide and a related undecanucleotide.  A total of 2400 nucleotides
have been sequenced.  Detailed analysis of the sequence divergence reveals that
both the overall extent of divergence and the frequency of base changes at
individual positions of the 23 bp repeats are characteristically different in
the various segments.  Preferentially methylated sites and a high incidence of
symmetry elements are found.  In two of the four segments, 22 of 23 bp of the
prototype sequence are included in six overlapping elements of dyad symmetry
and in a palindrome.  A scheme for the evolution of the satellite DNA from a
basic dodecanucleotide is proposed which is based on the different degrees of
divergence for the various repeats superimposed in this satellite DNA.

<>

<1>Pedersen, K., Bengtsson, A., Edlund, J., Rabe, L., Hazen, T., Chakraborty, R., Goodwin, L., Shapiro, N.
<2>Complete Genome Sequence of the Subsurface, Mesophilic Sulfate-Reducing Bacterium Desulfovibrio aespoeensis Aspo-2.
<3>Genome Announcements
<4>2
<5>e00509-14
<6>2014
<7>Desulfovibrio aespoeensis Aspo-2, DSM 10631(T), is a mesophilic, hydrogenotrophic
sulfate-reducing bacterium sampled from a 600-m-deep subsurface aquifer in hard
rock under the island of Aspo in southeastern Sweden. We report the genome
sequence of this bacterium, which is a 3,629,109-bp chromosome; plasmids were not
found.

<>

<1>Pedersen, T.B., Kot, W.P., Hansen, L.H., Sorensen, S.J., Broadbent, J.R., Vogensen, F.K., Ardo, Y.
<2>Genome Sequence of Leuconostoc mesenteroides subsp. cremoris Strain T26, Isolated from Mesophilic Undefined Cheese Starter.
<3>Genome Announcements
<4>2
<5>e00485-14
<6>2014
<7>Leuconostoc is the main group of heterofermentative bacteria found in mesophilic  dairy
starters. They grow in close symbiosis with the Lactococcus population and
are able to degrade citrate. Here we present a draft genome sequence of
Leuconostoc mesenteroides subsp. cremoris strain T26.

<>

<1>Pedersen, T.B., Kot, W.P., Hansen, L.H., Sorensen, S.J., Broadbent, J.R., Vogensen, F.K., Ardo, Y.
<2>Genome Sequences of Two Leuconostoc pseudomesenteroides Strains Isolated from Danish Dairy Starter Cultures.
<3>Genome Announcements
<4>2
<5>e00484-14
<6>2014
<7>The lactic acid bacterium Leuconostoc pseudomesenteroides can be found in mesophilic cheese
starters, where it produces aromatic compounds from, e.g.,
citrate. Here, we present the draft genome sequences of two L.
pseudomesenteroides strains isolated from traditional Danish cheese starters.

<>

<1>Pedulla, M.L. et al.
<2>
<3>Cell
<4>113
<5>171-182
<6>2003
<7>Bacteriophages are the most abundant organisms in the biosphere and play major roles in the
ecological balance of microbial life. The genomic
sequences of ten newly isolated mycobacteriophages suggest that the
bacteriophage population as a whole is amazingly diverse and may represent
the largest unexplored reservoir of sequence information in the biosphere.
Genomic comparison of these mycobacteriophages contributes to our
understanding of the mechanisms of viral evolution and provides compelling
evidence for the role of illegitimate recombination in horizontal genetic
exchange. The promiscuity of these recombination events results in the
inclusion of many unexpected genes including those implicated in
mycobacterial latency, the cellular and immune responses to mycobacterial
infections, and autoimmune diseases such as human lupus. While the role of
phages as vehicles of toxin genes is well established, these observations
suggest a much broader involvement of phages in bacterial virulence and
the host response to bacterial infections.

<>

<1>Peek, R.M. Jr., Thompson, S.A., Atherton, J.C., Blaser, M.J., Miller, G.G.
<2>Expression of ICEA, a novel ulcer-associated H. pylori gene, is induced by contact with gastric epithelial cells and is associated with enhanced mucosal IL-8.
<3>Gut
<4>39
<5>A71
<6>1996
<7>cagA+tox+ H. pylori strains are linked with peptic ulceration but most persons infected with
such isolates remain disease-free; thus, other unidentified virulence genes may be important
in pathogenesis.  For H. pylori, adherence to gastric epithelium may provide a stimulus for
induction of virulence gene expression.  iceA is a novel H. pylori gene that is selectively
up-regulated following contact with gastric epithelial cells.  The aims of this study were to
characterize iceA allellic diversity, correlate iceA genotypes with H. pylori virulence
determinants, peptic ulcer disease and in vivo IL-8 production, and examine expression of iceA
alleles following contact with gastric epithelial cells.

<>

<1>Pein, C.-D., Reuter, M., Cech, D., Kruger, D.H.
<2>Oligonucleotide duplexes containing CC(A/T)GG stimulate cleavage of refractory DNA by restriction endonuclease EcoRII.
<3>FEBS Lett.
<4>245
<5>141-144
<6>1989
<7>Some DNA species are resistant towards the restriction endonuclease EcoRII
despite the presence of unmodified recognition sites.  We show that 14
base-pair oligonucleotide duplexes containing the EcoRII recognition site
5'-CC(A/T)GG are cleaved by this enzyme and are able to stimulate EcoRII
cleavage of such resistant DNA molecules (e.g. DNA of bacterial virus T3).  A
direct correlation between the concentration of oligonucleotide duplex
molecules and the degree of EcoRII digestion of the primarily resistant DNA is
observed.  This indicates a stoichiometric rather than a catalytic mode of
enzyme activation.  An excess of DNA devoid of EcoRII sites (non-site DNA, e.g.
MvaI-digested T7 DNA) does not interfere with the activity of EcoRII.

<>

<1>Pein, C.-D., Reuter, M., Meisel, A., Cech, D., Kruger, D.H.
<2>Activation of restriction endonuclease EcoRII does not depend on the cleavage of stimulator DNA.
<3>Nucleic Acids Res.
<4>19
<5>5139-5142
<6>1991
<7>The restriction endonuclease EcoRII is unable to cleave DNA molecules when
recognition sites are very far apart.  The enzyme, however can be activated in
the presence of DNA molecules with a high frequency of EcoRII sites or by
oligonucleotides containing recognition sites:  Addition of the activator
molecules stimulates cleavage of the refractory substrate.  We now show that
endonucleolysis of the stimulator molecules is not a necessary prerequisite of
enzyme activation.  A total EcoRII digest of pBR322 DNA or oligonucleotide
duplexes with simulated EcoRII ends (containing the 5' phosphate group), as
well as oligonucleotide duplexes containing modified bases within the EcoRII
site, making them resistant to cleavage, are all capable of enzyme activation.
For activation EcoRII requires interaction with at least two recognition sites.
The two sites may be on the same DNA molecule, on different oligonucleotide
duplexes, or on one DNA molecule and one oligonucleotide duplex.  The
efficiency of functional intramolecular cooperation decreases with increasing
distance between the sites.  Intermolecular site interaction is inversely
related to the size of the stimulator oligonucleotide duplex.  The data are in
agreement with a model whereby EcoRII simultaneously interacts with two
recognition sites in the active complex, but cleavage of the site serving as an
allosteric activator is not necessary.

<>

<1>Pein, C.D., Cech, D., Gromova, E.S., Orezkaya, T.S., Shabarova, Z.A., Kubareva, E.A.
<2>Interaction of the MvaI restriction enzyme with synthetic DNA fragments.
<3>Nucleic Acids Symp. Ser.
<4>SS18
<5>225-228
<6>1987
<7>The cleavage of synthetic DNA duplexes by the restriction endonuclease MvaI has
been studied.  The main result of the cleavage experiments is that MvaI cleaves
unmodified duplexes in two single strand scissions in separate events and that
the two strands are cleaved at significantly different rates.  One strand nicks
within the recognition site do not affect the cleavage.  Furthermore, neither a
pyrophosphate internucleotide bond modification in one strand nor the absence
of one phosphate group at the central dA-residue of the recognition site
inhibits the cleavage of the second strand.

<>

<1>Pelaez, A.I., Ribas-Aparicio, R.M., Gomez, A., Rodicio, M.R.
<2>Establishment of a hybrid SalI-HgiDII type II restriction-modification system.
<3>Biol. Chem.
<4>379
<5>583-584
<6>1998
<7>In the SalI system, endonuclease activity can be only achieved in the presence of a functional
modification gene.  Thus, the DNA methyltransferase is involved in the control of restriction.
By fusion of the restriction gene of the SalI system to the modification gene of the
isospecific HgiDII system a hybrid type II restriction-modification system was created.
Although in the hybrid situation the level of endonuclease activity was significantly lower
than in the natural system, the HgiDII modification enzyme clearly supports SalI restriction.
The mechanism by which the two isospecific methyltransferases control restriction is currently
under study.

<>

<1>Pellenz, S., Dujon, B., Schafer, B.
<2>Target site cleavage by the homing endonuclease I-SpomI from fission yeast mitochondria.
<3>Yeast
<4>20
<5>S47
<6>2003
<7>Proteins encoded by mobile group I introns promote invasion into target DNAs.  The LAGLIDADG
homing endonuclease I-SpomI from the intron cox1Ilb of S. pombe mitochondria recognizes a
target DNA-sequence of 20bp.  As other representatives of this enzyme class I-SpomI generates
a 4nt 3' overhang.  Because of the length of the recognition sequence it can be employed for
the induction of specific double strand brakes in vitro and in vivo.  Since the recognition
site is almost palindromic, we changed it in such a way to give rise to a complete palindrome.
Since one of those variants was cut as well as the wild-type sequence, we assume that the
enzyme forms dimers and only one of the two LAGLIDADG-motifs is involved in the recognition of
the DNA-substrate.  An antibody against I-SpomI allows the determination of the active form in
mitochondrial extracts of the fission yeast.  Furthermore we introduced mutations into both
LADLIDADG-motifs: in motif P1 the aspartic residues were changed into alanine, in P2 the two
glutamic residues.  Inactivation of one of those motifs employed in the cutting mechanism was
supposed to result in a DNA single strand break actively of the mutant protein.  The
experiments were done in parallel with I-SceI from S. cerevisiae to have a direct comparison
between the different enzymes.  Enzymes with nicking activity can serve to study as well the
repair mechanisms of SSB and to give insights into the cutting mechanism of LAGLIDADG homing
endonucleases.

<>

<1>Pellenz, S., Harington, A., Dujon, B., Wolf, K., Schaefer, B.
<2>Characterization of the I-SpomI endonuclease from fission yeast: Insights into the evolution of a group I intron-encoded homing  endonuclease.
<3>J. Mol. Evol.
<4>55
<5>302-313
<6>2002
<7>The first group I intron in the cox1 gene (cox1I1b) of the mitochondrial genome of the fission
yeast Schizosaccharomyces pombe is
a mobile DNA element. The mobility is dependent on an endonuclease
protein that is encoded by an intronic open reading frame (ORF). The
intron-encoded endonuclease is a typical member of the LAGLIDADG
protein family of endonucleases with two consensus motifs. In addition
to this, analysis of several intron mutants revealed that this protein
is required for intron splicing. However, this protein is one of the
few group I intron-encoded proteins that functions in RNA splicing
simultaneously with its DNA endonuclease activity. We report here on
the biochemical characterization of the endonuclease activity of this
protein artificially expressed in Escherichia coli. Although the
intronic ORF is expressed as a fusion protein with the upstream exon in
vivo, the experiments showed that a truncated translation product
consisting of the C-terminal 304 codons of the cox1I1b ORF restricted
to loop 8 of the intron RNA secondary structure is sufficient for the
specific endonuclease activity in vitro. Based on the results, we
speculate on the evolution of site-specific homing endonucleases
encoded by group I introns in eukaryotes.

<>

<1>Pelletier, E., Kreimeyer, A., Bocs, S., Rouy, Z., Gyapay, G., Chouari, R., Riviere, D., Ganesan, A., Daegelen, P., Sghir, A., Cohen, G.N., Medigue, C., Weissenbach, J., Le Paslier, D.
<2>"Candidatus Cloacamonas acidaminovorans": genome sequence reconstruction provides a first glimpse of a new bacterial division.
<3>J. Bacteriol.
<4>190
<5>2572-2579
<6>2008
<7>Many microorganisms live in anaerobic environments. Most of these microorganisms have not yet
been cultivated. Here, we present, from a
metagenomic analysis of an anaerobic digester of a municipal wastewater
treatment plant, a reconstruction of the complete genome of a bacterium
belonging to the WWE1 candidate division. In silico proteome analysis
indicated that this bacterium might derive most of its carbon and energy
from the fermentation of amino acids, and hence, it was provisionally
classified as "Candidatus Cloacamonas acidaminovorans." "Candidatus
Cloacamonas acidaminovorans" is probably a syntrophic bacterium that is
present in many anaerobic digesters. This report highlights how
environmental sequence data might provide genomic and functional
information about a new bacterial clade whose members are involved in
anaerobic digestion.

<>

<1>Pelludat, C., Mirold, S., Hardt, W.D.
<2>The SopEPhi Phage Integrates into the ssrA Gene of Salmonella enterica Serovar Typhimurium A36 and Is Closely Related to the Fels-2 Prophage.
<3>J. Bacteriol.
<4>185
<5>5182-5191
<6>2003
<7>Salmonella spp. are enteropathogenic gram-negative bacteria that use a large array of
virulence factors to colonize the host, manipulate host
cells, and resist the host's defense mechanisms. Even closely related
Salmonella strains have different repertoires of virulence factors.
Bacteriophages contribute substantially to this diversity. There is
increasing evidence that the reassortment of virulence factor repertoires
by converting phages like the GIFSY phages and SopEPhi may represent an
important mechanism in the adaptation of Salmonella spp. to specific hosts
and to the emergence of new epidemic strains. Here, we have analyzed in
more detail SopEPhi, a P2-like phage from Salmonella enterica serovar
Typhimurium DT204 that encodes the virulence factor SopE. We have cloned
and characterized the attachment site (att) of SopEPhi and found that its
47-bp core sequence overlaps the 3' terminus of the ssrA gene of serovar
Typhimurium. Furthermore, we have demonstrated integration of SopEPhi into
the cloned attB site of serovar Typhimurium A36. Sequence analysis of the
plasmid-borne prophage revealed that SopEPhi is closely related to (60 to
100% identity over 80% of the genome) but clearly distinct from the Fels-2
prophage of serovar Typhimurium LT2 and from P2-like phages in the serovar
Typhi CT18 genome. Our results demonstrate that there is considerable
variation among the P2-like phages present in closely related Salmonella
spp.

<>

<1>Pembroke, J.T., Piterina, A.V.
<2>A novel ICE in the genome of Shewanella putrefaciens W3-18-1: comparison with the SXT/R391 ICE-like elements.
<3>FEMS Microbiol. Lett.
<4>264
<5>80-88
<6>2006
<7>A novel R391-like ICE (integrating conjugative element) has been detected in the  4.2 MB
genome of Shewanella putrefaciens W3-18-1 located on three different
contigs. Assembly of the ICE encoding contigs based on similarity with R391
revealed a mosaic element of plasmid, phage and transposon-like sequences typical
of SXT/R391 ICE-like elements. The element, which is 110 057 bp in length, was
highly similar to R391 sequences, with most related ORFs showing >96% amino acid
sequence identity. The element, designated ICESpuPO1, contained a number of
inserts determining resistance to copper and other heavy metals and a
broad-spectrum RND efflux pump similar to antibiotic efflux systems. The element
was integrated into the Shewanella prfC gene in a manner similar to related
ICE-like elements. The chromosomal element junctions contained a 17-bp
SXT/R391-like attL and attR site and an unannotated ORF between attL and the ICE
integrase encoding a putative recombinational directional factor necessary for
excision, with 100% amino acid identity to the R391 ORF4 product.

<>

<1>Pena, A., Busquets, A., Gomila, M., Bosch, R., Nogales, B., Garcia-Valdes, E., Lalucat, J., Bennasar, A.
<2>Draft Genome of Pseudomonas stutzeri Strain ZoBell (CCUG 16156), a Marine Isolate and Model Organism for Denitrification Studies.
<3>J. Bacteriol.
<4>194
<5>1277-1278
<6>2012
<7>Pseudomonas stutzeri strain ZoBell, formerly a strain of Pseudomonas perfectomarina (CCUG
16156 = ATCC 14405), is a model organism for
denitrification. It was isolated by ZoBell in 1944 from a marine sample, and here
we report the first genome draft of a strain assigned to genomovar 2 of the
species P. stutzeri.

<>

<1>Pena, A., Busquets, A., Gomila, M., Mayol, J., Bosch, R., Nogales, B., Garcia-Valdes, E., Bennasar, A., Lalucat, J.
<2>Draft Genome of Pseudomonas stutzeri Strain NF13, a Nitrogen Fixer Isolated from  the Galapagos Rift Hydrothermal Vent.
<3>Genome Announcements
<4>1
<5>e0011313
<6>2013
<7>Pseudomonas stutzeri strain NF13 was isolated from a water sample taken at a hydrothermal vent
in the Galapagos rift. It was selected for its ability to
metabolize sulfur compounds and to grow diazotrophically. Here, we report the
first draft genome of a member of genomovar 19 of the species.

<>

<1>Pena-Montenegro, T.D., Dussan, J.
<2>Genome sequence and description of the heavy metal tolerant bacterium Lysinibacillus sphaericus strain OT4b.31.
<3>Standards in Genomic Sciences
<4>9
<5>42-56
<6>2013
<7>Lysinibacillus sphaericus strain OT4b.31 is a native Colombian strain having no larvicidal
activity against Culex quinquefasciatus and is widely applied in the
bioremediation of heavy-metal polluted environments. Strain OT4b.31 was placed
between DNA homology groups III and IV. By gap-filling and alignment steps, we
propose a 4,096,672 bp chromosomal scaffold. The whole genome (consisting of
4,856,302 bp long, 94 contigs and 4,846 predicted protein-coding sequences)
revealed differences in comparison to the L. sphaericus C3-41 genome, such as
syntenial relationships, prophages and putative mosquitocidal toxins.
Sphaericolysin B354, the coleopteran toxin Sip1A and heavy metal resistance
clusters from nik, ars, czc, cop, chr, czr and cad operons were identified.
Lysinibacillus sphaericus OT4b.31 has applications not only in bioremediation
efforts, but also in the biological control of agricultural pests.

<>

<1>Peng, J., Yang, L., Yang, F., Yang, J., Yan, Y., Nie, H., Zhang, X., Xiong, Z., Jiang, Y., Cheng, F., Xu, X., Chen, S., Sun, L., Li, W., Shen, Y., Shao, Z., Liang, X., Xu, J., Jin, Q.
<2>Characterization of ST-4821 complex, a unique Neisseria meningitidis clone.
<3>Genomics
<4>91
<5>78-87
<6>2008
<7>Ten outbreaks of a new serogroup C meningococcal disease emerged during
2003-2005 in China. The multilocus sequence typing results indicated that
unique sequence type 4821 clone meningococci were responsible for these
outbreaks. Herein, we determined the entire genomic DNA sequence of
serogroup C isolate 053442, which belongs to ST-4821. Comparison of 053442
gene contents with other meningococcal genomes shows that they have
similar characteristics, including thousands of repetitive elements and
simple sequence repeats, numerous phase-variable genes, and similar
virulence-related factors. However, many strain-specific regions were
found in each genome. We also present the results of a genomic comparison
of 28 ST-4821 complex isolates that were isolated from different
serogroups using comparative genomic hybridization analysis. Genome
comparison between the newly emerged hyperinvasive isolates belonging to
different serogroups will further our understanding of their respective
pathogenetic mechanisms.

<>

<1>Pennell, R.I., Dang, V.
<2>Methods and compositions for altering seed phenotypes.
<3>International Patent Office
<4>EP 1687438 A, WO 2005038040 A
<5>
<6>2005
<7>Plants are disclosed that express a cytosine DNA methyltransferase and that can be used to
confer an altered seed phenotype, e.g., an increase in seed weight.  Also disclosed are plants
in which expression of an endogenous cytosine DNA methyltransferase is inhibited and that
exhibit an altered seed phenotype, e.g., an increase in seed weight.  Also disclosed are
nucleic acids and polypeptides suitable for conferring such phenotypes.

<>

<1>Penner, M., Morad, I., Snyder, L., Kaufmann, G.
<2>Phage T4-coded Stp: double-edged effector of coupled DNA and tRNA-restriction systems.
<3>J. Mol. Biol.
<4>249
<5>857-868
<6>1995
<7>The optional Escherichia coli prr locus encodes two physically associated restriction systems:
the type IC DNA restriction-modification enzyme EcoprrI and the tRNALys-specific anticodon
nuclease, specified by the PrrC polypeptide. Anticodon nuclease is kept latent as a result of
this interaction. The activation of anticodon nuclease, upon infection by phage T4, may cause
depletion of tRNALys and, consequently, abolition of T4 protein synthesis. However, this
effect is counteracted by the repair of tRNALys in consecutive reactions catalysed by the
phage enzymes polynucleotide kinase and RNA ligase. Stp, a short polypeptide encoded by phage
T4, has been implicated with activation of the anticodon nuclease. Here we confirm this notion
and also demonstrate a second function of Stp: inhibition of EcoprrI restriction. Both effects
depend, in general, on the same residues within the N-proximal 18 residue region of Stp. We
propose that Stp alters the conformation of EcoprrI and, consequently, of PrrC, allowing
activation of the latent anticodon nuclease. Presumably, Stp evolved to offset a DNA
restriction system of the host cell but was turned, eventually, against the phage as an
activator of the appended tRNA restriction enzyme.

<>

<1>Perea, J., Desdouets, C., Schapira, M., Jacq, C.
<2>I-SceIII: A novel group I intron-encoded endonuclease from the yeast mitochondria.
<3>Nucleic Acids Res.
<4>21
<5>358
<6>1993
<7>Re-engineered gene with "good" codons and expressed in E. coli. Cleavage site determined.

<>

<1>Perelman, E.V., Shtanchaeva, S.M., Bulk, V.F., Tarasov, A.P., Bakh, N.L., Semina, I.S.
<2>Use of different media for growing the producers of restricting enzyme XbaI.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>12
<5>48-50
<6>1984
<7>The possibility of using culture media prepared from local ingredients and
intended for growing the producers of restricting enzyme XbaI has been
demonstrated.  The yield of restricting enzyme XbaI per g of crude biomass,
obtained with the use of peptone-yeast medium prepared from ingredients
supplied by Difco Laboratories (USA), has proved to be 4 times greater than
that obtained with the use of peptone-yeast medium prepared from local
ingredients.  At the same time the use of casein-saline medium ensures the
yield of the enzyme, similar to that obtained with the use of peptone-yeast
medium prepared from ingredients supplied by Difco Laboratories, but with a
greater content of nonspecific nucleases.

<>

<1>Perevyazova, T.A., Rogulin, E.A., Zheleznaya, L.A., Matvienko, N.I.
<2>Cloning and sequencing of the gene of site-specific nickase N.BspD6I.
<3>Biokhimiia
<4>68
<5>1203-1207
<6>2003
<7>A fragment of chromosomal DNA from Bacillus species D6 containing the gene of nickase N.BspD6I
and the regions adjacent to its 5;- and 3;-ends was cloned and sequenced. The nucleotide
sequence of the nickase gene, except of one neutral change, is homologous to the nicking
endonuclease N.BstNBI gene sequenced by Higgens et al. (2001). After integration of a PCR-copy
of the nickase gene into an expression vector pET28b under the control of the phage T7
promoter, specific nicking activity was detected in the lysates of transformed E. coli cells.

<>

<1>Pericone, C.D., Bae, D., Shchepetov, M., McCool, T., Weiser, J.N.
<2>Short-sequence tandem and nontandem DNA repeats and endogenous hydrogen peroxide production contribute to genetic instability of Streptococcus   pneumoniae.
<3>J. Bacteriol.
<4>184
<5>4392-4399
<6>2002
<7>Loss-of-function mutations in the following seven pneumococcal genes were detected and
analyzed: pspA, spxB, xba, licD2, lytA, nanA, and atpC.
Factors associated with these mutations included (i) frameshifts caused by
reversible gain and loss of single bases within homopolymeric repeats as
short as 6 bases, (ii) deletions caused by recombinational events between
nontandem direct repeats as short as 8 bases, and (iii) substitutions of
guanine residues caused at an increased frequency by the high levels of
hydrogen peroxide (>2 mM) typically generated by this species under
aerobic growth conditions. The latter accounted for a frequency as high as
2.8 x 10(-6) for spontaneous mutation to resistance to optochin and was
10- to 200-fold lower in the absence of detectable levels of H2O2. Some of
these mutations appear to have been selected for in vivo during
pneumococcal infection, perhaps as a consequence of immune pressure or
oxidative stress.

<>

<1>Peris-Bondia, F., Muraille, E., Van Melderen, L.
<2>Complete Genome Sequence of the Escherichia coli PMV-1 Strain, a Model Extraintestinal Pathogenic E. coli Strain Used for Host-Pathogen Interaction  Studies.
<3>Genome Announcements
<4>1
<5>e00913-13
<6>2013
<7>Escherichia coli is a highly versatile species, causing diverse intestinal and extraintestinal
infections. Here, we present the complete genome sequence of
PMV-1, an O18:K1 extraintestinal pathogenic E. coli (ExPEC) strain that is used
as a model for peritonitis in mice and was useful for deciphering the innate
immune response triggered by ExPEC infections.

<>

<1>Perler, F.B.
<2>InBase, the New England Biolabs Intein Database.
<3>Nucleic Acids Res.
<4>27
<5>346-347
<6>1999
<7>Inteins are intervening sequences that splice as proteins, not RNA.  InBase, the New England
Biolabs Intein Database (http://www.neb.com/neb.inteins.html), is a comprehensive on-line
database that includes the Intein Registry, along with detailed information about each intein
and its host protein, tabulated comparisons and a comprehensive bibliography including papers
in press.

<>

<1>Perler, F.B.
<2>Hyperthermophilic inteins.
<3>Methods Enzymol.
<4>334
<5>270-280
<6>2001
<7>Inteins are intervening sequences that are posttranslationally excised from protein
precursors.  They are the protein equivalent of introns, which are intervening sequences that
splice from precursor RNAs.  The sequences flanking both sides of the intein are called
exteins.  During protein splicing, the intein is excised from a precursor protein and the
flanking exteins are joined by a peptide bond.  This ligation of exteins differentiates
protein splicing from other forms of proteolytic processing.  The self-catalytic protein
splicing reaction is mediated by the intein plus the first carboxy-extein amino acid, which
are capable of splicing in heterologous exteins.  However, each intein has its own "substrate"
specificity that dictates allowable proximal extein residues.  As of December 31, 1999, there
were 100 putative inteins listed in the Intein Registry, representing all three domains of
life (see InBase2 at http://www.neb.com/neb/intins.html); 74% of these inteins are found in
thermophilic organisms, mainly Archaea.  Thermophilic inteins were among the first inteins
discovered and played a key role in establishing protein splicing as a fundamental method of
protein biosynthesis.  The proof that inteins were spliced from precursor proteins rather than
from precursor RNAs and the mechanism of protein splicing were initially demonstrated using
archaeal inteins.  Since their discovery in 1990, inteins have been harnessed to perform
numerous protein engineering processes.

<>

<1>Perler, F.B.
<2>Inteins - A historical perspective.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Belfort, M., Berlin Heidelberg
<4>16
<5>193-210
<6>2005
<7>Protein splicing elements, termed inteins, were first identified in 1990.  Since then,
post-translational protein splicing has been demonstrated and the self-catalytic mechanism
deciphered.  The robust nature of these single turnover enzymes is evidenced by the expanding
list of naturally occurring variations in the protein splicing mechanism.  Protein splicing
must be efficient and neutral, and must not cause detrimental effects to the spliced extein;
otherwise, selective pressure would lead to intein loss.  Inteins are probably ancient
elements, but their original function can only be speculated upon, because invasion by homing
endonucleases mobilized them into new locations and converted them into selfish DNA.  To date,
there is no evidence of regulation of protein splicing in native systems.  The sporadic
distribution of inteins may relate more to the types of genes found in mobile elements capable
of spreading inteins, than to the function of those genes.  Inteins tend to be found in
conserved host protein motifs, which may be due to conservation of homing endonuclease
recognition sites, difficulty in removing inteins from essential regions or the ease of
accepting an insertion sequence in a conserved substrate or cofactor binding site designed to
interact with the environment.  The ability to cleave peptide bonds, to ligate protein
fragments and to generate carboxy-terminal alpha-thioesters have made inteins the fastest
growing tool for protein engineering and biotechnology.

<>

<1>Perler, F.B., Comb, D.G., Jack, W.E., Moran, L.S., Qiang, B., Kucera, R.B., Benner, J., Slatko, B.E., Nwankwo, D.O., Hempstead, S.K., Carlow, C.K.S., Jannasch, H.
<2>Intervening sequences in an Archaea DNA polymerase gene.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>5577-5581
<6>1992
<7>The DNA polymerase gene from the Archaea Thermococcus litoralis has been cloned and expressed
in Escherichia coli. It is split by two intervening sequences (IVSs) that form one continuous
open reading frame with three polymerase exons. To our knowledge, neither IVS is similar to
previously described introns. However, the deduced amino acid sequences of both IVSs are
similar to open reading frames present in mobile group I introns. The second IVS (IVS2)
encodes an endonuclease, I-TliI, that cleaves at the exon 2-exon 3 junction after IVS2 has
been deleted. IVS2 self-splices in E. coli to yield active polymerase, but processing is
abolished if the IVS2 reading frame is disrupted. Silent changes in the DNA sequence at the
exon 2-IVS2 junction that maintain the original protein sequence do not inhibit splicing.
These data suggest that protein rather than mRNA splicing may be responsible for production of
the mature polymerase.

<>

<1>Perler, F.B., Davis, E.O., Dean, G.E., Gimble, F.S., Jack, W.E., Neff, N., Noren, C.J., Thorner, J., Belfort, M.
<2>Protein splicing elements: inteins and exteins--a definition of terms and recommended nomenclature.
<3>Nucleic Acids Res.
<4>22
<5>1125-1127
<6>1994
<7>Several archaeal, eubacterial and eucaryotic genes have been identified with in-frame
insertions that are excised at the protein level, not at the RNA level. This process is termed
protein splicing. Initially, a single precursor polypeptide is synthesized. The intervening
protein sequences is then excised from within the precursor, and the flanking protein
sequences are joined. Thus, protein splicing results in the production of two proteins from a
single primary translation product, the internal protein and the protein formed by the joining
of the external sequences. The removal of the internal segment, concomitant with the formation
of a normal peptide bond joining the external polypeptide sequences, distinguishes protein
splicing from simple autoproteolysis. The rapid production of the mature products suggests
that protein splicing is very efficient. The protein precursor rarely accumulates, even when
the native gene is expressed in heterologous systems, both in vivo and in vitro. Evidence to
date suggests that protein splicing is autocatalytic.

<>

<1>Perler, F.B., Olsen, G.J., Adam, E.
<2>Compilation and analysis of intein sequences.
<3>Nucleic Acids Res.
<4>25
<5>1087-1093
<6>1997
<7>We have compiled a list of all the inteins (protein splicing elements) whose sequences have
been published or were available from on-line
sequence databases as of September 18, 1996. Analysis of the 36 available
intein sequences refines the previously described intein motifs and
reveals the presence of another intein motif, Block H. Furthermore,
analysis of the new inteins reshapes our view of the conserved splice
junction residues, since three inteins lack the intein penultimate His
seen in prior examples. Comparison of intein sequences suggests that, in
general, (i) inteins present in the same location within extein homologs
from different organisms are very closely related to each other in paired
sequence comparison or phylogenetic analysis and we suggest that they
should be considered intein alleles; (ii) multiple inteins present in the
same gene are no more similar to each other than to inteins present in
different genes; (iii) phylogenetic analysis indicates that inteins are so
divergent that trees with statistically significant branches cannot be
generated except for intein alleles.

<>

<1>Perlloni, A., Brown, E.W., LeClerc, J.E., Cebula, T.A.
<2>Phylogenetic evidence for horizontal gene transfer of type I restriction and modification (hsd) genes among highly homogenous Salmonella strains.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>105
<5>284
<6>2005
<7>Previous studies from our laboratory have documented the prevalence of horizontal gene
transfer (HGT) among strains of Salmonella enterica subspecies I. In comparing subspecies, a
recombination gradient was noted wherein the incidence of HGT is inversely correlated with the
genetic diversity separating individual strains. These findings suggested that there are
barriers, either genetic or ecological, that restrict exchange between disparate serovars of
Salmonella. The compatibility of restriction-modification (R-M) systems among strains has been
proposed as one explanation to account for the contrasting recombination rates. To explore
this possibility, 40 closely-related strains of the highly homogenous Typhimurium complex were
compared by cladistic analysis of the hsd genes, R, M, and S, which compose the type I R-M
system in Salmonella enterica subspecies I. The resultant trees revealed a prominent role for
HGT in the evolution of the hsd operon. Several Salmonella strains were found to be
evolutionarily discordant when hsd gene trees were compared to known markers of S. enterica
chromosome evolution (e.g., mdh). Additionally, several distinct clusters of mdh and mutS
alleles were coalesced into single hsd clades for the three type I R-M loci. Analyses of
congruence among hsd genes showed nearly unanimous discordance, the only exception being the
hsdM/S2 comparison (p = 1.0). This finding would suggest that the type I R-M operon is
anevolutionary mosaic, subject to numerous HGT events. This conclusion is further supported by
the identification of unique cassettes of sequence at two sites in hsdS. The hsdS2 segment
comprised two distinct sequences while the hsdS3 segment contained one of three unique
alleles. One of the hsdS3 alleles showed high homology to an E. coli hsd insert, suggesting a
recent cross-species transfer of this sequence. These data demonstrate that HGT has been a
common occurrence in hsd gene evolution and indicate an overall genetic compatibility among
closely-related salmonellae. This may explain in part why Salmonella known to share homologous
genomes and common niches are permitted to exchange DNA more freely.

<>

<1>Perna, N.T. et al.
<2>Genome sequence of enterohaemorrhagic Escherichia coli O157:H7.
<3>Nature
<4>409
<5>529-533
<6>2001
<7>The bacterium Escherichia coli O157:H7 is a worldwide threat to public health and has been
implicated in many outbreaks of haemorrhagic colitis, some of which included fatalities caused
by haemolytic uraemic syndrome.  Close to 75,000 cases of O157:H7 infection are now estimated
to occur annually in the United States.  The severity of disease, the lack of effective
treatment and the potential for large-scale outbreaks from contaminated food supplies have
propelled intensive research on the pathogenesis and detection of E. coli O157:H7.  Here we
have sequenced the genome of E. coli O157:H7 to identify candidate genes responsible for
pathogenesis, to develop better methods of strain detection and to advance our understanding
of the evolution of E. coli, through comparison with the genome of the non-pathogenic
laboratory strain E. coli K-12.  We find that lateral gene transfer is far more extensive than
previously anticipated.  In fact, 1,387 new genes encoded in strain-specific clusters of
diverse sizes were found in O157:H7.  These include candidate virulence factors, alternative
metabolic capacities, several prophages and other new functions - all of which could be
targets for surveillance.

<>

<1>Pernstich, C., Halford, S.E.
<2>Illuminating the reaction pathway of the FokI restriction endonuclease by fluorescence resonance energy transfer.
<3>Nucleic Acids Res.
<4>40
<5>1203-1213
<6>2012
<7>The FokI restriction endonuclease is a monomeric protein that recognizes an asymmetric
sequence and cleaves both DNA strands at fixed loci downstream of the
site. Its single active site is positioned initially near the recognition
sequence, distant from its downstream target 13 nucleotides away. Moreover, to
cut both strands, it has to recruit a second monomer to give an assembly with two
active sites. Here, the individual steps in the FokI reaction pathway were
examined by fluorescence resonance energy transfer (FRET). To monitor DNA binding
and domain motion, a fluorescence donor was attached to the DNA, either
downstream or upstream of the recognition site, and an acceptor placed on the
catalytic domain of the protein. A FokI variant incapable of dimerization was
also employed, to disentangle the signal due to domain motion from that due to
protein association. Dimerization was monitored separately by using two samples
of FokI labelled with donor and acceptor, respectively. The stopped-flow studies
revealed a complete reaction pathway for FokI, both the sequence of events and
the kinetics of each individual step.

<>

<1>Pero, J., Hannett, N.M., Talkington, C.
<2>Restriction cleavage map of SP01 DNA: general location of early, middle, and late genes.
<3>J. Virol.
<4>31
<5>156-171
<6>1979
<7>A detailed restriction endonuclease map for the genome of Bacillus subtilis phage SP01 is
presented. Sites of cleavage for the restriction
enzymes BglII, EcoRI, HaeIII, and SalI were determined. This physical map
showed that SP01 DNA was 140 kilobases in length and contained a repeated
sequence of 12.4 kilobases at its termini. Combined with previously
published information, we were also able to identify the general locations
of genes expressed at early, middle, or late times in the phage lytic
cycle. In particular, early genes were largely clustered in the terminal
repeats, whereas a major cluster of late genes was located in the
left-central portion of the genome.

<>

<1>Perona, J.J.
<2>Structural pathways of phosphoryl transfer in type II restriction endonucleases.
<3>ACS Abstracts
<4>223
<5>C-38-C-39
<6>2002
<7>Crystal structures of EcoRV endonuclease bound to DNA and divalent metal
ions have elucidated important aspects of the structural pathway of DNA
bending, the stereochemical mechanism of catalysis, and the coupling of
sequence selectivity to phosphoryl transfer. Three distinct divalent metal
ion binding sites have been located in the vicinity of the scissile DNA
phosphates, while a fourth site specific to manganese is found at the
interface of the enzyme with the DNA flanks. Mutational analysis of
active-site residues places important constraints on the reaction mechanism,
and suggests that moderate-range electrostatic effects play an important
role in facilitating rate enhancement. Measurements of the pH and
metal-dependence of the chemical step, by rapid-quench kinetics, lend
support to a detailed model of the reaction pathway derived from the crystal
structures. Comparisons of the EcoRV mechanism with those of other
restriction endonucleases offer further insight into the origins of
catalytic power.

<>

<1>Perona, J.J.
<2>Type II restriction endonucleases.
<3>Methods
<4>28
<5>353-364
<6>2002
<7>Type II restriction endonucleases have emerged as important paradigms for the study of
protein-nucleic acid interactions. This is due to their ability to catalyse phosphodiester
bond cleavage with very large rate enhancements while also maintaining exquisite sequence
selectivities. The principles and methods developed to analyze site-specific binding and
catalysis for restriction endonucleases can be applied to other enzymes which also operate on
nucleic acids. This paper reviews biochemical and structural approaches to characterization of
these enzymes, with particular attention to the multiple crucial roles of divalent metal ions,
the possibilities for use of alternative substrates in binding and catalytic experiments, the
strategies for exploring the detailed chemistry of phosphoryl transfer, and the use of X-ray
crystallography to provide descriptions of conformational pathways at specific, nonspecific,
and noncognate DNA sites.

<>

<1>Perona, J.J., Horton, N.C., Sam, M.D.
<2>Catalytic mechanism of EcoRV endonuclease derived from crystal structures and transient kinetics.
<3>Transactions ACA
<4>35
<5>9-17
<6>2000
<7>High-resolution cocrystal structures of wild-type and mutant forms of EcoRV endonuclease bound
to duplex DNA and divalent metal ions allow construction of a detailed model for the
pre-transition state configuration.  Three distinct metal-ion binding sites have been revealed
in different structural analyses.  In particular, a new site (site I) was recently found in a
ternary complex of the T93A mutant bound to cognate DNA and Ca2+ ions.  The same site is
occupied by Ca2+, Mg2+ or Mn2+ in three high-resolution structures of EcoRV bound to duplex
DNA containing 3'S-phosphorothiolate linkages (3'-PS) at the scissile phosphates.  Each of
these four structures traps a pre-transition state conformation in which the DNA is not
cleaved.  The new site I metal is ligated through an inner-sphere water molecule to the
phosphate group located 3' to the scissile phosphate.  A second inner-sphere water on this
metal is positioned approximately in-line for attack on the scissile phosphate.  These
structures corroborate the observation that the pro-SP phosphoryl oxygen on the adjacent
3'-phosphate cannot be modified without severe loss of catalytic efficiency.  Together with
previous cocrystal structures, these data allow construction of a detailed model for the
pre-transition state configuration in EcoRV.  This model features three divalent metal ions
per active site, and invokes assistance in the bond-making step by a conserved lysine, which
stabilizes the attacking hydroxide ion nucleophile.  The model is supported by pre-steady
state and single-turnover kinetic data, in which the metal and pH-dependencies of the
phosphoryl transfer step are directly evaluated.  The structural equivalence of key groups,
conserved in the active sites of many type II restriction endonucleases, suggests that
ligation of a catalytic divalent metal ion to the adjacent 3'-phosphate may occur in many
type II restriction enzymes.

<>

<1>Perona, J.J., Martin, A.M.
<2>Conformational transitions and structural deformability of EcoRV endonuclease revealed by crystallographic analysis.
<3>J. Mol. Biol.
<4>273
<5>207-225
<6>1997
<7>The structures of wild-type and mutant forms of the unliganded EcoRV endonuclease dimer have
been determined at 2.4 angstroms resolution in a new crystal lattice.  Comparison of these
structures with that of the free enzyme determined with different packing constraints shows
that the conformations of the domain interfaces are not conserved between crystal forms.  The
unliganded enzyme and the enzyme-DNA complex delineate two distinct quaternary states
separated by a 25 degree intersubunit rotation, but considerable conformational heterogeneity,
of the order of 10^6 domain rotations, exists within each of these states.  Comparison of the
free enzyme structure between the two crystal forms further reveals that the C-terminal 28
amino acid residues are disordered and undergo an extensive local folding transition upon DNA
binding.  Introduction of the mutation T93A at the DNA-binding cleft causes large-scale
effects on the protein conformation.  Structural changes in the mutated unliganded enzyme
propagate some 20 to 25 angstroms to the dimerization interface and lead to a rearrangement of
monomer subunits.  Comparative analysis of these structures, a new structure of the enzyme
cocrystallized with DNA and calcium ions, and previously determined cocrystal structures
suggests important roles for a number of amino acid residues in facilitating the intersubunit
motions and local folding transitions.  In particular, the T93A structure reveals a pathway
through the protein, by which DNA-binding may cause the domain movements required for proper
alignment of catalytic groups.  The key active-site residue Glu45 is located on a flexible
helix inside the pathway, and this provides a direct means by which essential catalytic
functions are coupled to the protein conformational change.  It appears that indirect
perturbation of the Glu45 conformation via an altered quaternary structure may be a
contributing factor to the decreased catalytic efficiency of T93A, and this mechanism may also
explain the diminished activities of other active site variants of EcoRV.

<>

<1>Perotto, S., Nepote-Fus, P., Saletta, L., Bandi, C., Young, J.P.W.
<2>A diverse population of introns in the nuclear ribosomal genes of ericoid mycorrhizal fungi includes elements with sequence similarity to endonuclease-coding genes.
<3>Mol. Biol. Evol.
<4>17
<5>44-59
<6>2000
<7>Ericoid mycorrhizal fungi form symbioses with the roots of members of the Ericales. Although
only two genera have been identified in culture,
the taxonomic diversity of ericoid symbionts is certainly wider.
Genetic variation among 40 ericoid fungal isolates was investigated in
this study. PCR amplification of the nuclear small-subunit ribosomal
DNA (SSU rDNA) and of the internal transcribed spacer (ITS), followed
by sequencing, led to the discovery of DNA insertions of various sizes
in the SSU rDNA of most isolates. They reached sizes of almost 1,800 bp
and occurred in up to five different insertion sites. Their positions
and sizes were generally correlated with morphological and ITS-RFLP
grouping of the isolates, although some insertions were found to be
optional among isolates of the same species, and insertions were not
always present in all SSU rDNA repeats within an isolate. Most
insertions were identified as typical group I introns, possessing the
conserved motifs characteristic of this group. However, other
insertions lack these motifs and form a distinct group that includes
other fungal ribosomal introns. Alignments with almost 70 additional
sequences from fungal nuclear SSU rDNA introns indicate that introns
inserted at the same site along the rDNA gene are generally homologous,
but they also suggest the possibility of some horizontal transfers. Two
of the ericoid fungal introns showed strong homology with a conserved
motif found in endonuclease genes from nuclear rDNA introns.

<>

<1>Perreten, V., Chanchaithong, P., Prapasarakul, N., Rossano, A., Blum, S.E., Elad, D., Schwendener, S.
<2>Novel Pseudo-Staphylococcal Cassette Chromosome mec Element ({Psi}SCCmec57395) in Methicillin-Resistant Staphylococcus pseudintermedius CC45.
<3>Antimicrob. Agents Chemother.
<4>57
<5>5509-5515
<6>2013
<7>Genetic characterization of methicillin-resistant Staphylococcus pseudintermedius
(MRSP) from Thailand and Israel revealed the presence of a predominant atypical
clonal lineage which was not typeable by SmaI-PFGE and SCCmec typing. All the
atypical isolates (n = 34) belonged to CC45 (30 ST45 and 2 ST179 isolates, 1 ST57
isolate, and 1 ST85 isolate). The isolates originated from healthy and diseased
dogs and cats, as well as from the environment of one clinic. Cfr9I-pulsed-field
gel electrophoresis (Cfr9I-PFGE) and dru typing permitted the further distinction
of CC45 isolates from the two different countries. Microarray analysis identified
genes that confer resistance to beta-lactams (mecA; blaZ), aminoglycosides
[aac(6')-Ie-aph(2')-Ia; aph(3')-III; ant(6)-Ia], macrolides and lincosamides
[erm(B)], tetracyclines [tet(M)], trimethoprim [dfr(G)], streptothricin (sat4),
and chloramphenicol (catpC221). Fluoroquinolone resistance was attributed to
specific amino acid substitutions, i.e., Ser84Leu in GyrA and Ser80Ile and
Asp84Asn in GrlA. A novel pseudo-staphylococcal cassette chromosome
(PsiSCCmec57395) element was identified in MRSP strain 57395 (sequence type ST45)
by whole-genome sequencing. The 12,282-bp PsiSCCmec57395 element contained a
class C1 mec gene complex but no ccr genes. In addition to the methicillin
resistance gene mecA, PsiSCCmec57395 also carried determinants of resistance to
heavy metals, such as arsenic, cadmium, and copper. Bsu36I restriction analysis
of the PsiSCCmec57395 element amplified by long-range PCR revealed the presence
of PsiSCCmec57395 in the 33 additional isolates of MRSP CC45. The PsiSCCmec57395
element represents a new class of SCCmec and has been identified in MRSP of CC45,
which is a predominant clonal lineage in Israel and Thailand.

<>

<1>Perrin, A., Buckle, M., Dujon, B.
<2>Asymmetrical recognition and activity of the I-SceI endonuclease on its site and on intron-exon junctions.
<3>EMBO J.
<4>12
<5>2939-2947
<6>1993
<7>Group I intron-encoded endonucleases represent a new class of double strand cutting
endonucleases whose function is to initiate the homing of introns by generating double strand
breaks in site-specific sequences. We have studied the mechanism of interaction of the I-SceI
endonuclease with different DNA substrates derived from its natural site in the intron-less
gene or from intron-exon junctions in the gene with an intron. We show that the enzyme
recognizes it asymmetrical site with high affinity binding to the sequence corresponding to
the downstream exon followed by binding to the upstream exon and catalysis of phosphodiester
bond hydrolysis. Asymmetrical nicking activity is observed as an intermediate of the cleavage
reaction. In the intron-containing gene, the enzyme recognizes the downstream intron-exon
junction without any cleavage activity. This binding raises the possibility of a specific
function of homing endonucleases in either gene expression or intron homing steps subsequent
to DNA cleavage.

<>

<1>Perrody, E., Cirinesi, A.M., Desplats, C., Keppel, F., Schwager, F., Tranier, S., Georgopoulos, C., Genevaux, P.
<2>A Bacteriophage-Encoded J-Domain Protein Interacts with the DnaK/Hsp70 Chaperone and Stabilizes the Heat-Shock Factor sigma(32) of Escherichia coli.
<3>PLoS Genet.
<4>8
<5>E1003037
<6>2012
<7>The universally conserved J-domain proteins (JDPs) are obligate cochaperone
partners of the Hsp70 (DnaK) chaperone. They stimulate Hsp70's ATPase activity,
facilitate substrate delivery, and confer specific cellular localization to
Hsp70. In this work, we have identified and characterized the first functional
JDP protein encoded by a bacteriophage. Specifically, we show that the ORFan gene
057w of the T4-related enterobacteriophage RB43 encodes a bona fide JDP protein,
named Rki, which specifically interacts with the Escherichia coli host
multifunctional DnaK chaperone. However, in sharp contrast with the three known
host JDP cochaperones of DnaK encoded by E. coli, Rki does not act as a generic
cochaperone in vivo or in vitro. Expression of Rki alone is highly toxic for
wild-type E. coli, but toxicity is abolished in the absence of endogenous DnaK or
when the conserved J-domain of Rki is mutated. Further in vivo analyses revealed
that Rki is expressed early after infection by RB43 and that deletion of the rki
gene significantly impairs RB43 proliferation. Furthermore, we show that
mutations in the host dnaK gene efficiently suppress the growth phenotype of the
RB43 rki deletion mutant, thus indicating that Rki specifically interferes with
DnaK cellular function. Finally, we show that the interaction of Rki with the
host DnaK chaperone rapidly results in the stabilization of the heat-shock factor
sigma(32), which is normally targeted for degradation by DnaK. The mechanism by
which the Rki-dependent stabilization of sigma(32) facilitates RB43 bacteriophage
proliferation is discussed.

<>

<1>Persson, T. et al.
<2>Genome Sequence of 'Candidatus Frankia datiscae' Dg1, the Uncultured Microsymbiont from Nitrogen-Fixing Root Nodules of the Dicot Datisca  glomerata.
<3>J. Bacteriol.
<4>193
<5>7017-7018
<6>2011
<7>Members of the noncultured clade of Frankia enter into root nodule symbioses with actinorhizal
species from the orders Cucurbitales and
Rosales. We report the genome sequence of a member of this clade
originally from Pakistan but obtained from root nodules of the American
plant Datisca glomerata without isolation in culture.

<>

<1>Pertry, I., Vaclavikova, K., Depuydt, S., Galuszka, P., Spichal, L., Temmerman, W., Stes, E., Schmulling, T., Kakimoto, T., Van Montagu, M.C., Strnad, M., Holsters, M., Tarkowski, P., Vereecke, D.
<2>Identification of Rhodococcus fascians cytokinins and their modus operandi to reshape the plant.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>929-934
<6>2009
<7>Decades ago, the importance of cytokinins (CKs) during Rhodococcus
fascians pathology had been acknowledged, and an isopentenyltransferase
gene had been characterized in the fas operon of the linear virulence
plasmid, but hitherto, no specific CK(s) could be associated with
virulence. We show that the CK receptors AHK3 and AHK4 of Arabidopsis
thaliana are essential for symptom development, and that the CK perception
machinery is induced upon infection, underlining its central role in the
symptomatology. Three classical CKs [isopentenyladenine, trans-zeatin, and
cis-zeatin (cZ)] and their 2-methylthio (2MeS)-derivatives were identified
by CK profiling of both the pathogenic R. fascians strain D188 and its
nonpathogenic derivative D188-5. However, the much higher CK levels in
strain D188 suggest that the linear plasmid is responsible for the
virulence-associated production. All R. fascians CKs were recognized by
AHK3 and AHK4, and, although they individually provoked typical CK
responses in several bioassays, the mixture of bacterial CKs exhibited
clear synergistic effects. The cis- and 2MeS-derivatives were poor
substrates of the apoplastic CK oxidase/dehydrogenase enzymes and the
latter were not cytotoxic at high concentrations. Consequently, the
accumulating 2MeScZ (and cZ) in infected Arabidopsis tissue contribute to
the continuous stimulation of tissue proliferation. Based on these
results, we postulate that the R. fascians pathology is based on the local
and persistent secretion of an array of CKs.

<>

<1>Pertsev, A.V., Denmukhametov, M.M., Anoshkin, A.G., Ariskina, E.V., Berezin, I.A., Solonin, A.S., Kuzmin, N.P.
<2>Restriction endonucleases from various bacterial strains displaying an ice-nucleating activity.
<3>Prikl. Biokhim. Mikrobiol.
<4>36
<5>13-16
<6>2000
<7>Six strains containing Type II site-specific endonucleases were selected from a collection of
45
ice-nucleating bacterial strains isolated from the
rhizosphere of plants growing in various geographical regions.
Endonucleases Pfl21I, Psp8I, and Psp23I were isolated and purified
from two Pseudomonas sp. strains and a Pseudomonas fluorescens strain.
Restriction endonucleases Pfl21I and Psp23I were shown to recognize and
cleave the DNA nucleotide sequence 5'-CTGCA^G-3'.
Endonuclease Psp8I recognized and cleaved the DNA nucleotide sequence
5'-G^GATCC-3'. These endonucleases were found to be true
isoschizomers of PstI and BamHI, respectively.

<>

<1>Pertsev, A.V., Kravets, A.N.
<2>New Klebsiella azeanae strain produces restriction endonuclease Kaz48I - useful for recognition and splitting of specific nucleotide sequence.
<3>Russian Patent Office
<4>RU 2044055
<5>
<6>1995
<7>
<>

<1>Pertsev, A.V., Kravets, A.N.
<2>New Escherichia coli strain produces restriction endonuclease Eco110k1 - new strain for large-scale production of restriction enzyme.
<3>Russian Patent Office
<4>RU 2044054
<5>
<6>1995
<7>
<>

<1>Pertsev, A.V., Kravets, A.N., Beletskaya, I.V., Zakharova, M.V., Solonin, A.S.
<2>An E. coli strain, producing Eco75KI restriction endonuclease - Escherichia coli culture for enzyme production for use in genetic engineering.
<3>Russian Patent Office
<4>RU 2053298 C
<5>
<6>1996
<7>
<>

<1>Pertzev, A.V., Kravetz, A.N., Mayorov, S.G., Zakharova, M.V., Solonin, A.S.
<2>Isolation of a strain overproducing endonuclease Eco29kI: Purification and characterization of the homogeneous enzyme.
<3>Biokhimiia
<4>62
<5>858-867
<6>1997
<7>The physical map of the plasmid pSACII1 carrying the genes of the restriction-modification
system Eco29kI (isoschizomer of SacII) was determined.  The cloning of the Eco29kI
endonuclease methylase genes into the plasmid vector pUC129 produced recombinant strain
Escherichia coli K802 [pECO29A15] with Eco29kI synthesis level about 100 times higher than in
the parent strain.  The restriction endonuclease was purified from Escherichia coli K802
[pECO29A15] cells to near homogeneity using column chromatography sequentially on
phosphocellulose, hydroxyapatite, and heparin-Sepharose and rechromatography on
phosphocellulose.  Biochemical characterization of the homogeneous R.Eco29kI is given.  The
enzyme has molecular mass 24.5 kD and is present in the solution as a monomer.

<>

<1>Pertzev, A.V., Ruban, N.M., Zakharova, M.V., Beletzkaja, I.V., Petrov, S.I., Kravetz, A.N., Solonin, A.S.
<2>Eco29kI, a novel plasmid encoded restriction endonuclease from Escherichia coli.
<3>Nucleic Acids Res.
<4>20
<5>1991
<6>1992
<7>Eco29kI is a type II restriction endonuclease from clinical strain Escherichia coli 29
isolated in Kiev. The genes for the Eco29kI restriction modification system were located on
one of its plasmids, namely pSACII1 about 4.0 kb in size as was determined by plasmid
transformation of E. coli K802 (Figure 1) according to (1) except that the phage Phi 80 vir
was used for selection of clones.

<>

<1>Pester, M. et al.
<2>Complete Genome Sequences of Desulfosporosinus orientis DSM765T, Desulfosporosinus youngiae DSM17734T, Desulfosporosinus meridiei DSM13257T, and  Desulfosporosinus acidiphilus DSM22704T.
<3>J. Bacteriol.
<4>194
<5>6300-6301
<6>2012
<7>Desulfosporosinus species are sulfate-reducing bacteria belonging to the Firmicutes. Their
genomes will give insights into the genetic repertoire and
evolution of sulfate reducers typically thriving in terrestrial environments and
able to degrade toluene (Desulfosporosinus youngiae), to reduce Fe(III)
(Desulfosporosinus meridiei, Desulfosporosinus orientis), and to grow under
acidic conditions (Desulfosporosinus acidiphilus).

<>

<1>Pestova, E.V., Morrison, D.A.
<2>Isolation and characterization of three Streptococcus pneumoniae transformation-specific loci by use of a lacZ reporter insertion vector.
<3>J. Bacteriol.
<4>180
<5>2701-2710
<6>1998
<7>Although more than a dozen new proteins are produced when Streptococcus pneumoniae cells
become competent for genetic transformation, only a few of the corresponding genes have been
identified to date.  To find genes responsible for the production of competence-specific
proteins, a random lacZ transcriptional fusion library was constructed in S. pneumoniae by
using the insertional lacZ reporter vector pEVP3.  Screening the library for clones with
competence-specific B-galactosidase production yielded three insertion mutants with induced
beta-Gal levels of about 4, 10, and 40 Miller units.  In all three clones, activation of the
lacZ reporter correlated with competence and depended on competence-stimulating peptide.
Chromosomal loci adjacent to the integrated vector were subcloned from the insertion mutants,
and their nucleotide sequences were determined.  Genes at two of the loci exhibited strong
similarity to parts of Bacillus subtilis com operons.  One locus contained open reading frames
homologous to the comEA and comEC genes in B. subtilis but lacked a comEB homolog.  A second
locus contained four ORFs with homology to the B. subtilis comG gene ORFs 1 to 4, but comG
gene ORFs 5 to 7 were replaced in S. pneumoniae with an ORF encoding a protein homologous to
transport ATP-binding proteins.  Genes at all three loci were confirmed to be required for
transformation by mutagenesis using pEVP3 for insertion duplications or an erm cassette for
gene disruptions.

<>

<1>Petering, H., Hammerschmidt, S., Frosch, M., van Putten, J.P.M., Ison, C.A., Robertson, B.D.
<2>Genes associated with the meningococcal capsule complex are also found in Neisseria gonorrhoeae.
<3>J. Bacteriol.
<4>178
<5>3342-3345
<6>1996
<7>A homolog of the meningococcal cps locus region E has been identified in Neisseria gonorrhoeae
immediately upstream of the gonococcal region D locus.  Region E has no detectable function in
capsule biosynthesis in Neisseria meningitidis or in lipopolysaccharide biosynthesis in either
organism.  The open reading frame is homologous to proteins of unknown function in Escherichia
coli and Haemophilus influenzae.  Further analysis of the N. meningitidis cps cluster has
identified a second copy of region D encoding three additional open reading frames, including
homologs of DNA methyltransferases.  The organization of the region D and E genes in N.
gonorrhoeae and N. meningitidis in relation to the cps genes provides some insight into the
evolutionary origin of encapsulation in N. meningitidis.

<>

<1>Peters, I.
<2>Changing the sequence specificity of the restriction endonuclease EcoRI.
<3>Ph.D. Thesis, Germany
<4>
<5>
<6>2003
<7>EcoRI is one of the best studied restriction endonucleases of type II.  It recognizes the
palindromic DNA sequence GAATTC with high specificity and cleaves it between G and A in the
presence of Mg2+ ions.  This specific recognition is due to a highly complex and redundant
network of hydrogen bonds, ionic and hydrophobic contacts.  In order to change the sequence
specificity of the enzyme concerning the GC base pair at the end of the recognition sequence
three amino acid residues (Met137, Arg200 and Arg203) were mutated.  Met137 was replaced by
Asn, Cys, Gln, Leu and Lys, Arg200 by Lys and Tyr and Arg203 by Lys.  Already the single
mutation M137Q results in an obvious change of specificity, because of its preference for
5-methyl cytosine instead of cytosine during cleavage experiments with both
oligodeoxynucleotide and plasmid substrates depending on the sequence context within the DNA.
All other mutants at amino acid Met137 revealed a strong reduction of cleavage activity.  Thus
only the single mutant M137Q was used as a starting point for further mutations within one
protein.  The four mutations M137Q, R200K, R200Y and R203K were combined with each other to
form any variation possible in order to generate multiple mutants.  Except for M137Q/R203K all
mutants showed heavy loss of cleavage activity.  The mutant M137Q/R203K revealed an increased
ability to cleave star sites.  In this case the stringent coupling of specific DNA recognition
and catalysis is relaxed and the cleavage of DNA also takes place even if there are more
interrupted DNA contacts than those to the GC base pair.  By combining the double mutation
M137Q/R203K with R200Y an enzyme was created, that despite very low activity was able to
cleave all four palindromic hexamer variations with a central AATT sequence at star
conditions, whereas the sequence CAATTG was obviously preferred.  Thus an alteration of
sequence specificity was obtained for the triple mutant M137Q/R200Y/R203K.

<>

<1>Peters, W.
<2>New applications of (S)-adenosyl-L-methionine analogues with protein methyltransferases and click chemistry for sequence specific protein labelling.
<3>Ph.D. Thesis, Germany
<4>
<5>
<6>2007
<7>
<>

<1>Peterson, K.R., Mount, D.W.
<2>Analysis of the genetic requirements for viability of Escherichia coli K-12 DNA adenine methylase (dam) mutants.
<3>J. Bacteriol.
<4>175
<5>7505-7508
<6>1993
<7>RecBCD protein, necessary for Escherichia coli dam mutant viability, is directly required for
DNA repair. Recombination genes recF+, recN+, recO+, and recQ+ are not essential for dam
mutant viability; they are required for recBC sbcBC dam mutant survival, mutH, mutL, or mutS
mutations do not suppress subinduction of SOS genes in dam mutants.

<>

<1>Peterson, K.R., Wertman, K.F., Mount, D.W., Marinus, M.G.
<2>Viability of Escherichia coli K-12 DNA adenine methylase (dam) mutants requires increased expression of specific genes in the SOS regulon.
<3>Mol. Gen. Genet.
<4>201
<5>14-19
<6>1985
<7>We have examined the level of expression of the SOS regulon in cells lacking DNA adenine
methylase activity (dam-). Mud (Ap,lac) fusions to several SOS operons (recA, lexA, uvrA,
uvrB,uvrD, sulA, dinD and dinF) were found to express higher levels of beta-galactosidase in
dam- strains than in isogenic dam+ strains. The attempted construction of dam- strains that
were also mutant in one of several SOS genes indicated that the viability of
methylase-deficient strains correlates with the inactivation of the SOS repressor (LexA
protein). Consistent with this, the wild-type functions of two LexA-repressed genes (recA and
ruv) appear to be required for dam- strain viability.

<>

<1>Peterson, R.C.
<2>Prediction of restriction endonuclease site distribution based on dinucleotide frequency.
<3>Fed. Proc.
<4>45
<5>1850
<6>1986
<7>Most analyses of the frequency of restriction endonuclease recognition sites
have assumed a random distribution of nucleotides.  Nearest neighbor analysis
and the sequence of nucleic acids indicate that the distribution of nucleotides
is nonrandom at least at the dinucleotide level.  Recent analyses have made use
of the nonrandom dinucleotide distribution observed in sequenced nucleic acids
to predict the frequency of restriction endonuclease sites.  The frequency of
sites in DNAs which have not been extensively sequenced can also be predicted
using the dinucleotide frequencies measured by nearest neighbor analysis.
Taking into account the dinucleotide bias, the site frequencies for a large
number of restriction endonucleases have been calculated for the DNAs from
several organisms.  Some restriction endonucleases which have the same base
composition in their recognition sites [e.g. EcoRV (GATATC) and HindIII
(AAGCTT)] have widely different site frequencies in the DNA from some
organisms.  The observed distribution of DNA fragments generated by selected
restriction endonucleases correlates with the predicted frequency of the
recognition site.  This type of analysis should be useful in the selection of
suitable restriction endonucleases for mapping genomic DNAs, for generating DNA
fragments for subcloning, and for screening for restriction fragment length
polymorphisms.

<>

<1>Peterson, R.C.
<2>Prediction of the frequencies of restriction endonuclease recognition sequences using di- and mononucleotide frequencies.
<3>Biotechniques
<4>6
<5>34-39
<6>1988
<7>The calculation of probabilities of nucleotide sequences from the freqeuencies
of dinucleotides is described.  The dinucleotide and mononucleotide frequencies
used can be obtained from nearest neighbor analysis or from databank sequences.
If dinucleotide and mononucleotide frequencies from nearest neighbor analysis
are used, probabilities for oligonucleotides can be calculated for genomes in
which there is little or no sequence data.  Within a given genome, a broad
range of probabilities for hexanucleotide palindromes with the same base
composition is predicted and shown.

<>

<1>Peterson, S.N., Reich, N.O.
<2>GATC flanking sequences regulate Dam activity: evidence for how Dam specificity may influence pap expression.
<3>J. Mol. Biol.
<4>355
<5>459-472
<6>2005
<7>Escherichia coli DNA adenine methyltransferase (Dam) plays essential roles in DNA replication,
mismatch repair and gene regulation. The differential methylation by Dam of the two GATC
sequences in the pap promoter regulates the expression of pili genes necessary for
uropathogenic E.coli cellular adhesion. Dam processively methylates GATC sites in various DNA
substrates, yet the two pap GATC sites are not processively methylated. We previously proposed
that the flanking sequences surrounding the two pap GATC sites contribute to the enzyme's
distributive methylation. We show here that replacement of the poorly methylated pap GATC
sites with sites predicted to be processively methylated indeed results in an increase in Dam
processivity. The increased processivity is due to a change in the methyltransfer kinetics and
not the binding efficiency of Dam. A competition experiment in which the flanking sequences of
only one pap GATC site were altered demonstrates that the GATC flanking sequences directly
regulate the enzyme's catalytic efficiency. The GATC flanking sequences in Dam-regulated
promoters in E.coli and other bacteria are similar to those in the pap promoter. Gene
regulation from some of these promoters involves mechanisms and proteins that are quite
different from those in the pap operon. Further, GATC sequences previously identified to
remain unmethylated within the E.coli genome, but whose function remains largely unassigned,
are flanked by sequences predicted to be poorly methylated. We conclude that the GATC flanking
sequences may be critical for expression of pap and other Dam-regulated genes by affecting the
activity of Dam at such sites and, thus, its processivity. A model is proposed, illustrating
how the sequences flanking the GATC sites in Dam-regulated promoters may contribute to this
epigenetic mechanism of gene expression, and how flanking sequences contribute to the diverse
biological roles of Dam.

<>

<1>Pethick, F.E. et al.
<2>Complete Genome Sequences of Corynebacterium pseudotuberculosis Strains 3/99-5 and 42/02-A, Isolated from Sheep in Scotland and Australia, Respectively.
<3>J. Bacteriol.
<4>194
<5>4736-4737
<6>2012
<7>Here, we report the whole-genome sequences of two ovine-pathogenic Corynebacterium
pseudotuberculosis isolates: strain 3/99-5, which represents the
first C. pseudotuberculosis genome originating from the United Kingdom, and
42/02-A, the second from Australia. These genome sequences will contribute to the
objective of determining the global pan-genome of this bacterium.

<>

<1>Pethick, F.E., Macfadyen, A.C., Tang, Z., Sangal, V., Liu, T.T., Chu, J., Kosec, G., Petkovic, H., Guo, M., Kirby, R., Hoskisson, P.A., Herron, P.R., Hunter, I.S.
<2>Draft Genome Sequence of the Oxytetracycline-Producing Bacterium Streptomyces rimosus ATCC 10970.
<3>Genome Announcements
<4>1
<5>e0006313
<6>2013
<7>We report the draft genome of Streptomyces rimosus (ATCC 10970), a soil isolate that produces
oxytetracycline, a commercially important and clinically useful
antibiotic.

<>

<1>Petranovic, M., Petranovic, D., Dohet, C., Brooks, P., Radman, M.
<2>Some restriction endonucleases tolerate single mismatches of the pyrimidine-purine type.
<3>Nucleic Acids Res.
<4>18
<5>2159-2162
<6>1990
<7>DNAs from phage mutants M13mp18 and M13mp18/MP-1 were used to construct two
closed circular heteroduplexes.  One of them carried the sequence 5'-CCTGGG-3'
3'-GGGCCC-5' with a T-G mismatch at the position 6248.  The other carried the
sequence  5'-CCCGGG-3' 3'-GGACCC-5' with a C-A mismatch at the same position.
Heteroduplexes were exposed to 7 restriction endonucleases having recognition
sites within the sequence  5'-CCCGGG-3'  3'-GGGCCC-5' and to 1 restriction
endonuclease having a recognition site within the sequence  5'-CCTGGG-3'
3'-GGACCC-5'.  All tested enzymes cleaved at least one mismatch-containing
sequence although with reduced efficiency.  SmaI and XmaI tolerated both
mismatch-containing sequences.  AvaI, HpaII, MspI, NciI and NspIII were able to
tolerate only the T-G containing sequence while BstNI was able to tolerate only
the C-A containing sequence.  It is inferred that the tolerance displayed by
SmaI and XmaI depends on the presence of either the original purines or the
original pyrimidines in mismatches of both the T-G and C-A type and that all
other tested enzymes require the presence of the original purines in mismatches
of both types.

<>

<1>Petrauskene, O., Kubareva, E.A., Gromova, E.S.
<2>A spectrophotometric method for studying the cleavage of DNA duplexes by restriction endonucleases.
<3>Mol. Biol. (Mosk)
<4>25
<5>1424-1426
<6>1991
<7>A spectrophotometric method for continuously monitoring the cleavage of DNA duplexes by type
II restriction endonucleases was proposed.  The time course of cleavage of a 14-membered DNA
duplex by MvaI endonuclease was obtained.  The spectrophotometric method is characterized by
rapidity and high precision in determining the kinetic parameters of the reaction.  It can be
recommended for testing preparations for the presence of restriction endonucleases, rapid
determination of the activity of any restriction endonucleases, highly precise quantitative
analysis of the restriction enzyme catalysed reactions.

<>

<1>Petrauskene, O.V., Babkina, O.V., Tashlitsky, V.N., Kazankov, G.M., Gromova, E.S.
<2>EcoRII endonuclease has two identical DNA-binding sites and cleaves one of two co-ordinated recognition sites in one catalytic event.
<3>FEBS Lett.
<4>425
<5>29-34
<6>1998
<7>EcoRII is a typical restriction enzyme that cleaves DNA using a two-site mechanism.  EcoRII
endonuclease is unable to cleave DNA which contains a small number of EcoRII recognition sites
but the enzyme activity can be stimulated in the presence of DNA with a high frequency of
EcoRII sites.  To investigate the mechanism of activation, the kinetics of stimulated EcoRII
cleavage has been studied.  A 14 bp substrate activated the cleavage of the 71 bp substrate,
containing one EcoRII recognition site (trans-activation) by a competitive mechanism: the
activator increased substrate binding but not catalysis.  The activation increased if the
substrate concentration decreased and if the activator had a lower affinity for the enzyme
than the substrate.  The introduction of the second recognition site into the 71 bp duplex
also enabled cleavage of this substrate (cis-activation).  Pyrophosphate bonds were
incorporated into one of two recognition sites to switch off the cleavage of the
phosphodiester bonds.  Analysis of cleavage products of these modified substrates showed that
EcoRII cuts one of two coordinated recognition sites in one catalytic event.

<>

<1>Petrauskene, O.V., Gromova, E.S., Romanova, E.A., Volkov, E.M., Oretskaya, T.S., Shabarova, Z.A.
<2>DNA duplexes containing methylated bases or non-nucleotide inserts in the recognition site are cleaved by restriction endonuclease R.EcoRII in presence of canonical substrate.
<3>Gene
<4>157
<5>173-176
<6>1995
<7>DNA duplexes containing the natural methylated bases N6-methyladenine (m6Ade),
N4-methylcytosine (m4Cyt) or C5-methylcytosine (m5Cyt) in one strand of the recognition
sequence are resistant to EcoRII restriction endonuclease (R.EcoRII).  Hydrolysis of these
modified duplexes was observed in the presence of the canonical substrate.  Incorporation of
m4Cyt or m5Cyt into both strands of the recognition sequence precludes such activation by a
canonical substrate.  R.EcoRII also fails to cleave substrate analogs in which one of the
nucleosides in the recognition site is replaced by the 1,2-dideoxyribose (D) or by
1,3-propanediol (Prd) (modeling DNA with an abasic site).  The hydrolysis of DNA duplexes with
non-nucleotide inserts is also activated in the presence of canonical substrate.  Thus, the
two-substrate mechanism of EcoRII-DNA interaction allows hydrolysis of apurinic/apyrimidinic
and hemimethylated DNA.

<>

<1>Petrauskene, O.V., Karpova, E.A., Gromova, E.S., Guschlbauer, W.
<2>Two subunits of EcoRII restriction endonuclease interact with two DNA recognition sites.
<3>Biochem. Biophys. Res. Commun.
<4>198
<5>885-890
<6>1994
<7>The cleavage of a 14 base pair DNA duplex containing one EcoRII recognition site by EcoRII
restriction endonuclease (R.EcoRII) was studied in single turnover experiments with varying
enzyme concentrations in the micromolar range. The reaction rate increased with enzyme
concentration until a ratio of one dimeric R.EcoRII enzyme to two double stranded
olionucleotide molecules. Excess of R.EcoRII lead to inhibition of cleavage. Maximum cleavage
was also found with pBR322 DNA containing six EcoRII recognition sites at a ratio of one
dimeric enzyme to two EcoRII recognition sites of the plasmid DNA. At higher ratios inhibition
was observed. These observations indicate that the active enzyme complex is formed when two
subunits of the enzyme interact with two R. EcoRII recognition sites.

<>

<1>Petrauskene, O.V., Krynetskaya, N.F., Tashlitsky, V.N., Belkov, V.M., Kubareva, E.A., Gromova, E.S., Guschlbauer, W., Shabarova, Z.A.
<2>Use of UV spectroscopy for the study of nucleic acid cleavage by E.coli RNase H and restriction endonucleases.
<3>Biochem. Mol. Biol. Int.
<4>37
<5>1127-1135
<6>1995
<7>A one-step spectrophotometric method for monitoring of nucleic acid cleavage by ribonuclease H
from E.coli and type II restriction endonucleases has been proposed.  It is based on recording
the increase in the UV absorbance at 260 nm during the course of enzymatic reaction.  Duplexes
stable under the reaction conditions were chosen as substrates for the enzymes being studied.
In order to obtain duplex dissociation following their cleavage by the enzyme appropriate
temperature conditions were selected.  The spectrophotometric method may be applied for rapid
testing of the nuclease activity in protein preparations as well as for precise quantitative
analysis of nucleic acid degradation by enzymes.  This method may be successfully employed in
kinetic studies of nucleic acid - protein interactions.

<>

<1>Petrauskene, O.V., Kubareva, E.A., Gromova, E.S., Pein, C.-D., Cech, D., Shabarova, Z.A.
<2>Synthetic DNA duplexes as a tool in studying the mechanism of EcoRII activation.
<3>Mol. Biol. (Mosk)
<4>27
<5>507-518
<6>1993
<7>The efficiency of EcoRII cleavage of synthetic DNA duplexes with one EcoRII recognition site
decreases with increasing substrate length. This enzyme virtually fails to cleave DNA duplexes
longer than 215 bp. However, EcoRII cleaves long DNA duplexes with one recognition site in the
presence of 11-14 bp substrates. The extent of hydrolysis activation depends on the length and
concentration of the added substrate. A model system is suggested for studying the molecular
and kinetic mechanism of EcoRII activation. This system includes a 30-bp substrate with one
EcoRII recognition site, and DNA duplexes as activating substrates that contain modified
heterocyclic bases and internucleotide phosphate groups. DNA duplexes with a modified EcoRII
recognition site may activate hydrolysis of the 30-bp substrate and of phage T3 DNA. Their
catalytic effect on the cleavage of extended duplexes depends on the type of modification and
its localization in the recognition site. Cooperative interaction of EcoRII with two
recognition sites in DNA has been shown to be essential for the functioning of the enzyme.

<>

<1>Petrauskene, O.V., Kubareva, E.A., Gromova, E.S., Shabarova, Z.A.
<2>Mechanism of the interaction of EcoRII restriction endonuclease with two recognition sites - probing of modified DNA duplexes as activators of the enzyme.
<3>Eur. J. Biochem.
<4>208
<5>617-622
<6>1992
<7>The efficiency of cleavage of DNA duplexes with single EcoRII recognition sites by the EcoRII
restriction endonuclease decreases with increasing substrate length. DNA duplexes of more than
215 bp are not effectively cleaved by this enzyme. Acceleration of the hydrolysis of long
single-site substrates by EcoRII is observed in the presence of 11-14 bp substrates. The
stimulation of hydrolysis depends on the length and concentration of the second substrate. To
study the mechanism of EcoRII endonuclease stimulation, DNA duplexes with base analogs and
modified internucleotide phosphate groups in the EcoRII site have been investigated as
activators. These modified duplexes are cleaved by EcoRII enzyme with different efficiences or
are not cleaved at all. It has been discovered that the resistance of some of them can be
overcome by incubation with a susceptible canonical substrate. The acceleration of cleavage of
long single-site substrates depends on the type of modification of the activator. The modified
DNA duplexes can activate EcoRII catalyzed hydrolysis if they can be cleaved by EcoRII
themselves or in the presence of the second canonical substrate. It has been demonstrated that
EcoRII endonuclease interacts in a cooperative way with two recognition sites in DNA. The
cleavage of one of the recognition sites depends on the cleavage of the other. We suggest that
the activator is not an allosteric effector but acts as a second substrate.

<>

<1>Petrauskene, O.V., Schmidt, S., Karyagina, A.S., Nikolskaya, I.I., Gromova, E.S., Cech, D.
<2>The interaction of DNA duplexes containing 2-aminopurine with restriction endonucleases EcoRII and SsoII.
<3>Nucleic Acids Res.
<4>23
<5>2192-2197
<6>1995
<7>Oligonucleotides containing 2-aminopurine (2-AP) in place of G or A in the recognition site of
EcoRII (CCT/AGG) or SsoII (CCNGG) restriction endonucleases have been synthesized in order to
investigate the specific interaction of DNA with these enzymes. Physicochemical properties (CD
spectra and melting behaviour) have shown that DNA duplexes containing 2-aminopurine exist
largely in a stable B-like form. 2-Aminopurine base paired with cytidine, however, essentially
influences the helix structure. The presence of a 2-AP.C mismatch strongly reduces the
stability of the duplexes in comparison with the natural double strand, indicated by a
biphasic melting behaviour. SsoII restriction endonuclease recognizes and cleaves the modified
substrate with a 2-AP.T mismatch in the centre of the recognition site, but it does not cleave
the duplexes containing 2-aminopurine in place of inner and outer G, or both. EcoRII
restriction endonuclease does not cleave duplexes containing 2-aminopurine at all. The
two-substrate mechanism of EcoRII-DNA interaction, however, allows hydrolysis of the duplex
containing 2-aminopurine in place of adenine in the presence of the canonical substrate.

<>

<1>Petrauskene, O.V., Tashlitsky, V.N., Brevnov, M.G., Bakman, Y., Gromova, E.S.
<2>Kinetic modeling of allosteric interaction of EcoRII restriction endonuclease with two DNA sites.
<3>Biokhimiia
<4>61
<5>1257-1269
<6>1996
<7>The effect of correlations between kinetic parameters of two inducible substrates on
allosteric activation of EcoRII endonuclease hydrolysis of one substrate was studied.  The
pairs of DNA duplexes were constructed that were the substrates of EcoRII restriction
endonuclease or their analogs and had different kinetic constants of interaction with the
enzyme; the effects of their concentrations on mutual hydrolysis induction were studied.  A
kinetic mechanism is suggested considering the allosteric effects of two DNA recognition sites
on a dimeric molecule of EcoRII.  Mathematic modelling was used to analyze the kinetic
mechanism and evaluate optimal characteristics of the inductor.  Thus, activation increases
when substrate concentration decreases, enzyme binding of two inductor or substrate molecules
decreases, binding of one substrate molecule increases versus binding of one inductor
molecule, and kcat of the enzyme-substrate complex including one substrate and one inductor
increases.

<>

<1>Petrauskene, O.V., Yakovleva, J.N., Alekseev, Y.I., Subach, F.V., Babkina, O.V., Gromova, E.S.
<2>DNA duplexes containing altered sugar residues as probes of EcoRII and MvaI endonuclease interactions with sugar-phosphate backbone.
<3>J. Biomol. Struct. Dyn.
<4>17
<5>857-870
<6>2000
<7>Oligonucleotides containing 1-(beta-D-2'-deoxy-threo-pentofuranosyl)cytosine (dCx) and/or
1-(beta-D-2'-deoxy-threo-pentofuranosyl)thymine (dTx) in place of dC and dT residues in the
EcoRII and MvaI recognition site CC(A/T)GG were synthesized in order to investigate specific
recognition of the DNA sugar-phosphate backbone by EcoRII and MvaI restriction endonucleases.
In 2'-deoxyxylosyl moieties of dCx and dTx, 3'-hydroxyl groups were inverted, which perturbs
the related individual phosphates. Introduction of a single 2'-deoxyxylosyl moiety into a dC
x dG pair resulted in a minor destabilization of double-stranded DNA structure. In the case of
a dA x dT pair the effect of a 2'-deoxyxylose incorporation was much more pronounced.
Multiple dCx modifications and their combination with dTx did not enhance the destabilization
effect. Hydrolysis of dCx-containing DNA duplexes by EcoRII endonuclease was blocked and
binding affinity was strongly dependent on the location of an altered sugar. A DNA duplex
containing a dTx residue was cleaved by the enzyme, but kcat/K(M) was slightly reduced. In
contrast, MvaI endonuclease efficiently cleaved both types of sugar-altered substrate analogs.
However it did not cleave conformationally perturbed scissile bonds, when the corresponding
unmodified bonds were perfectly hydrolyzed in the same DNA duplexes. Based on these data the
possible contributions of individual phosphates in the recognition site to substrate
recognition and catalysis by EcoRII were proposed. We observed strikingly non-equivalent
inputs for different phosphates with respect to their effect on EcoRII-DNA complex formation.

<>

<1>Petrauskiene, L.J., Klimasauskas, S.J., Butkus, V.V., Janulaitis, A.
<2>Synthesis of oligodeoxynucleotides containing N4-methylcytosine.
<3>Bioorg. Khim.
<4>12
<5>1597-1603
<6>1986
<7>With deoxyuridine as starting material, N4-methyldeoxycytidine and its fully protected
mononucleotide, suitable for oligonucleotide synthesis, have been prepared.  By means of the
phosphotriester approach, the fully protected mononucleotide was used for the synthesis of
seven dodecadeoxynucleotides containing either m4C or m5C in various positions of the CCCGGG
sequence, the recognition site of some restriction endonucleases.

<>

<1>Petronella, N., Kenwell, R., Pagotto, F., Pightling, A.W.
<2>Draft Genome Sequences of Two Clostridium botulinum Group II (Nonproteolytic) Type B Strains (DB-2 and KAPB-3).
<3>Genome Announcements
<4>2
<5>e01111-14
<6>2014
<7>Clostridium botulinum is important for food safety and studies of neurotoxins associated with
human botulism. We present the draft genome sequences of two
strains belonging to group II type B: one collected from Pacific Ocean sediments
(DB-2) and another obtained during a botulism outbreak (KAPB-3).

<>

<1>Petroni, E.A., Bocca, S.N., Ielpi, L.
<2>Sequence-specific DNA modification in Acetobacter xylinum.
<3>Cell. Mol. Biol. (Noisy-le-grand)
<4>42
<5>759-767
<6>1996
<7>Two cryptic plasmids have been discovered in Acetobacter xylinum B42 and in its derivative
PEA-1, a cellulose defective mutant.  These two plasmids were designated pAX1 and pAX2 (50 and
105 in size, respectively).  A restriction map was constructed for pAX1.  Attempts to cure
these plasmids were unsuccessful.  Enzyme restriction analysis showed that these plasmids
contain protected EcoRI and ApoI sites.  Using Southern blot and hybridization techniques, the
protection was extended to chromosomal DNA.  Enzyme restriction analysis of several plasmids,
from different origins and containing different incompatibility groups, isolated from strain
PEA-1 also showed EcoRI and ApoI protection.  The presence of modifications on specific
sequences was not found in A. xylinum 8747.  These results strongly suggest the presence of a
modification system in A. xylinum B42 that recognizes the tetranucleotide 5'-AATT.

<>

<1>Petronzio, T., Schildkraut, I.
<2>Altered specificity of restriction endonucleases HinfI.
<3>Nucleic Acids Res.
<4>18
<5>3666
<6>1990
<7>None

<>

<1>Petrosino, J.F., Xiang, Q., Karpathy, S.E., Jiang, H., Yerrapragada, S., Liu, Y., Gioia, J., Hemphill, L., Gonzalez, A., Raghavan, T.M., Uzman, A., Fox, G.E., Highlander, S., Reichard, M., Morton, R.J., Clinkenbeard, K.D., Weinstock, G.M.
<2>Chromosome Rearrangement and Diversification of Francisella tularensis Revealed by the Type B (OSU18) Genome Sequence.
<3>J. Bacteriol.
<4>188
<5>6977-6985
<6>2006
<7>The gamma-proteobacterium Francisella tularensis is one of the most
infectious human pathogens, and the highly virulent organism F. tularensis
subsp. tularensis (type A) and less virulent organism F. tularensis subsp.
holarctica (type B) are most commonly associated with significant disease
in humans and animals. Here we report the complete genome sequence and
annotation for a low-passage type B strain (OSU18) isolated from a dead
beaver found near Red Rock, Okla., in 1978. A comparison of the F.
tularensis subsp. holarctica sequence with that of F. tularensis subsp.
tularensis strain Schu4 (P. Larsson et al., Nat. Genet. 37:153-159, 2005)
highlighted genetic differences that may underlie different pathogenicity
phenotypes and the evolutionary relationship between type A and type B
strains. Despite extensive DNA sequence identity, the most significant
difference between type A and type B isolates is the striking amount of
genomic rearrangement that exists between the strains. All but two
rearrangements can be attributed to homologous recombination occurring
between two prominent insertion elements, ISFtu1 and ISFtu2. Numerous
pseudogenes have been found in the genomes and are likely contributors to
the difference in virulence between the strains. In contrast, no
rearrangements have been observed between the OSU18 genome and the genome
of the type B live vaccine strain (LVS), and only 448 polymorphisms have
been found within non-transposase-coding sequences whose homologs are
intact in OSU18. Nonconservative differences between the two strains
likely include the LVS attenuating mutation(s).

<>

<1>Petrosova, H., Zobanikova, M., Cejkova, D., Mikalova, L., Pospisilova, P., Strouhal, M., Chen, L., Qin, X., Muzny, D.M., Weinstock, G.M., Smajs, D.
<2>Whole Genome Sequence of Treponema pallidum ssp. pallidum, Strain Mexico A, Suggests Recombination between Yaws and Syphilis Strains.
<3>PLoS Neglected Trop. Dis.
<4>6
<5>E1832
<6>2012
<7>BACKGROUND: Treponema pallidum ssp. pallidum (TPA), the causative agent of
syphilis, and Treponema pallidum ssp. pertenue (TPE), the causative agent of
yaws, are closely related spirochetes causing diseases with distinct clinical
manifestations. The TPA Mexico A strain was isolated in 1953 from male, with
primary syphilis, living in Mexico. Attempts to cultivate TPA Mexico A strain
under in vitro conditions have revealed lower growth potential compared to other
tested TPA strains. METHODOLOGY/PRINCIPAL FINDINGS: The complete genome sequence
of the TPA Mexico A strain was determined using the Illumina sequencing
technique. The genome sequence assembly was verified using the whole genome
fingerprinting technique and the final sequence was annotated. The genome size of
the Mexico A strain was determined to be 1,140,038 bp with 1,035 predicted ORFs.
The Mexico A genome sequence was compared to the whole genome sequences of three
TPA (Nichols, SS14 and Chicago) and three TPE (CDC-2, Samoa D and Gauthier)
strains. No large rearrangements in the Mexico A genome were found and the
identified nucleotide changes occurred most frequently in genes encoding putative
virulence factors. Nevertheless, the genome of the Mexico A strain, revealed two
genes (TPAMA_0326 (tp92) and TPAMA_0488 (mcp2-1)) which combine TPA- and TPE-
specific nucleotide sequences. Both genes were found to be under positive
selection within TPA strains and also between TPA and TPE strains.
CONCLUSIONS/SIGNIFICANCE: The observed mosaic character of the TPAMA_0326 and
TPAMA_0488 loci is likely a result of inter-strain recombination between TPA and
TPE strains during simultaneous infection of a single host suggesting horizontal
gene transfer between treponemal subspecies.

<>

<1>Petrossian, T., Clarke, S.
<2>Bioinformatic identification of novel methyltransferases.
<3>Epigenomics
<4>1
<5>163-175
<6>2009
<7>Methylation of DNA, protein and even RNA species are integral processes in epigenesis. Enzymes
that catalyze these reactions using the donor S-adenosylmethionine fall into several
structurally distinct classes. The members in each class share sequence similarity that can be
used to identify additional methyltransferases. Here, we characterize these classes and in
silico approaches to infer protein function. Computational methods, such as hidden Markov
model profiling and the Multiple Motif Scanning program, can be used to analyze known
methyltransferases and relay information into the prediction of new ones. In some cases, the
substrate of methylation can be inferred from hidden Markov model sequence similarity
networks. Functional identification of these candidate species is much more difficult; we
discuss one biochemical approach.

<>

<1>Petrossian, T.C., Clarke, S.G.
<2>Computational methods to identify novel methyltransferases.
<3>BMC Bioinformatics
<4>10
<5>7
<6>2009
<7>1.2% of the yeast genes are estimated to encode enzymes that catalyze the transfer of a methyl
group from S-adenosylmethionine to protein, nucleic acid, lipid, and small molecule
substrates.  These enzymes function in biosynthesis, regulating metabolic pathways and
controlling gene expressiopn, including writing the histone code.  BLAST and MEM/MAST analysis
using the amino acid sequence of motifs have previously generated a list of putative Class I
methyltransferases.  Recently we have used a combination of a new search algorithm and
structural information to refine this analysis.  This study utilizes these updated methods of
identifying motifs and scanning the proteome to predict new members of the different families
of methyltransferases in different organisms.  These new members may function in novel
pathways or new modes of regulation.

<>

<1>Petrov, N.A., Gorbunov, Y.A., Malygin, E.G.
<2>Interaction of T4 phage DNA-[N6-adenine]-methyltransferase with substrates containing defective recognition sites.
<3>Mol. Biol. (Mosk)
<4>31
<5>973-977
<6>1997
<7>The effect of the structure of the recognition site GATC in a substrate duplex on its complex
form ation with DNA-[N6-adenine]-methyltransferase of T4 phage was studied.  The gel
retardation method was employed to reveal the complexes.  As compared with the native
duplexes, the majority of defective duplexes had the same or even better affinity to T4 MTase;
however, no correlation was found between the complex stability and effectiveness of the
duplexes as substrates for methylation.  Apparently, formation of a stale enzyme-DNA complex
does not require continuity of the two strands of the duplex and perfect base pairing in the
recognition site.  A half of the constituents of the recognition site suffices for stable
complexes with T4 MTase to form.  Deoxyguanosine residues in both strands of the modified GATC
are shown to be indispensable for complex formation.

<>

<1>Petrov, N.A., Gorbunov, Y.A., Naumochkin, A.N., Malygin, E.G.
<2>Substrate complexes of DNA-[N6-adenine]-methyltransferases of T-even phages registered by the gel retardation method.
<3>Mol. Biol. (Mosk)
<4>31
<5>966-972
<6>1997
<7>DNA-[N6-adenine]-methyltransferases of T4 and T2 phages (T4 and T2 Mtases) recognize, in
double-stranded DNA, the palindrome GATC and catalyze the transfer of the methyl group from
S-adenosyl-L-methionine (SAM) to position N6 of the adenine residue.  The gel retardation
method was used to study the relative effectiveness of complex formation of T4 and T2 MTases
with oligonucleotide substrates of varying length containing GATC in the middle of the duplex.
It is shown that T4 MTase forms stable complexes with 20-mer duplexes bearing a nonmodified or
hemimethylated GATC site.  The binding of the duplex to T4 MTase is enhanced in the presence
of SAM.  Parameters of the interaction of SAM with MTase not bound and bound with the 20-mer
duplex are determined.  T2 MTase, which has a higher catalytic activity than the T4 enzyme,
forms less stable complexes with oligonucleotides.  Thus, there is no direct relation between
the stability of the enzyme-substrate complexes and the catalytic activity.

<>

<1>Petrov, V.M., Ratnayaka, S., Nolan, J.M., Miller, E.S., Karam, J.D.
<2>Genomes of the T4-related bacteriophages as windows on microbial genome evolution.
<3>Virol. J.
<4>7
<5>292
<6>2010
<7>ABSTRACT: The T4-related bacteriophages are a group of bacterial viruses
that share morphological similarities and genetic homologies with the
well-studied Escherichia coli phage T4, but that diverge from T4 and each
other by a number of genetically determined characteristics including the
bacterial hosts they infect, the sizes of their linear double-stranded
(ds) DNA genomes and the predicted compositions of their proteomes. The
genomes of about 40 of these phages have been sequenced and annotated over
the last several years and are compared here in the context of the factors
that have determined their diversity and the diversity of other microbial
genomes in evolution. The genomes of the T4 relatives analyzed so far
range in size between ~160,000 and ~250,000 base pairs (bp) and are
mosaics of one another, consisting of clusters of homology between them
that are interspersed with segments that vary considerably in genetic
composition between the different phage lineages. Based on the known
biological and biochemical properties of phage T4 and the proteins encoded
by the T4 genome, the T4 relatives reviewed here are predicted to share a
genetic core, or "Core Genome" that determines the structural design of
their dsDNA chromosomes, their distinctive morphology and the process of
their assembly into infectious agents (phage morphogenesis). The Core
Genome appears to be the most ancient genetic component of this phage
group and constitutes a mere 12-15% of the total protein encoding
potential of the typical T4-related phage genome. The high degree of
genetic heterogeneity that exists outside of this shared core suggests
that horizontal DNA transfer involving many genetic sources has played a
major role in diversification of the T4-related phages and their spread to
a wide spectrum of bacterial species domains in evolution. We discuss some
of the factors and pathways that might have shaped the evolution of these
phages and point out several parallels between their diversity and the
diversity generally observed within all groups of interrelated dsDNA
microbial genomes in nature.

<>

<1>Petrovski, S., Seviour, R.J., Tillett, D.
<2>Prevention of Gordonia and Nocardia Stabilized Foam Formation by Using Bacteriophage GTE7.
<3>Appl. Environ. Microbiol.
<4>77
<5>7864-7867
<6>2011
<7>Most activated sludge treatment plants suffer from the presence of foams
on the surfaces of their aeration reactors. These are often stabilized by
hydrophobic mycolic acid-synthesizing actinobacterial species. A
polyvalent Siphoviridae phage, GTE7, which lysed several Gordonia and
Nocardia species, is described here. Its genome has a modular structure
similar to that described for Rhodococcus phage ReqiDocB7. In
laboratory-scale experiments, we showed that GTE7 prevents stabilization
of foams by these Gordonia and Nocardia species.

<>

<1>Petruska, J., Horn, D.
<2>Sequence-specific responses of restriction endonucleases to bromodeoxyuridine substitution in mammalian DNA.
<3>Nucleic Acids Res.
<4>11
<5>2495-2510
<6>1983
<7>Substitution of BrdU for dT in mammalian DNA alters the rates of DNA cleavage
by restriction endonucleases in a manner that can be related to the specificity
of cleavage.  A formula is proposed that describes inhibitory and stimulatory
contributions arising from the substitution of a Br atom for the CH3 group on
T.  The larger Br atom is postulated to sterically hinder the nuclease from
binding to adjacent groups in the DNA cleavage site, while allowing a tighter
binding to itself.  The inhibition caused by steric hindrance is predicted to
vary inversely with distance from the point of cleavage, whereas the
stimulation caused by tighter binding is predicted to be independent of
distance.  The resultant formula gives a good fit to the data obtained for
thirteen different restriction nucleases of known specificity.  The parameters
in the formula appear to be simple functions of ionic strength.  This formula
can be used to predict the effect of BrdU substitution on any endonuclease
whose specificity of cleavage is known.

<>

<1>Petrusyte, M., Bitinaite, J., Menkevicius, S., Klimasauskas, S., Butkus, V., Janulaitis, A.
<2>Restriction endonucleases of a new type.
<3>Gene
<4>74
<5>89-91
<6>1988
<7>Meeting Abstract

<>

<1>Petrusyte, M., Rudokas, K., Maneliene, Z., Kiuduliene, E., Butkus, V., Janulaitis, A.
<2>Bsp1407I, a restriction endonuclease from Bacillus stearothermophilus, which recognizes novel palindromic sequence 5'-T^GTACA-3'.
<3>Nucleic Acids Res.
<4>21
<5>2514
<6>1993
<7>A new type II restriction endonuclease, Bsp1407I, has been isolated from Bacillus
stearothermophilus RFL1407. Bsp1407I recognizes the palindromic hexanucleotide 5'- TGTACA-3'
generating 5'-protruding tetranucleotide. Bsp1407I cuts lambda DNA at five sites, SV40 at two
sites and does not cleave phiX174 and pBR322 DNA. We found that the computer calculated number
of fragments that would be generated at the sequences 5'-TGTACA, correlated with the observed
cleavage frequency of the above mentioned substrates. Double digestion with Bsp1407I and
Bsp1201 (ApaI), Pfl23II(SplI), Eco81I(SauI) and XhoI were used to map the Bsp1407I sites on
lambda DNA. The mapped positions matched those predicted by cleavage at the sequence
5'-TGTACA. The cleavage site of Bsp1407I was determined using a synthetic oligonucleotide
duplex which contains the Bsp1407I recognition sequence:
5'-GAGTGTACACTC-3'
3'-CTCACATGTGAG-5'.

<>

<1>Petrusyte, M.P., Bitinaite, J.B., Kersulyte, D.R., Menkevicius, S.J., Butkus, V.V., Janulaitis, A.
<2>New types of restriction endonucleases.
<3>Dokl. Akad. Nauk.
<4>295
<5>1250-1253
<6>1987
<7>The restriction enzymes GsuI and Eco57I are described.  The cleavage sites were
characterized as CTGGAG (16/14) for GsuI and CTGAAG (16/14) for Eco57I.  Both
enzymes are activated by SAM.  Complete fragmentation was not possible!
Endonuclease and methylase activities could not be separated for Eco57I.  For
GsuI no methylase activity was detected.

<>

<1>Petrusyte, M.P., Janulaitis, A.
<2>Isolation and some properties of the restriction endonuclease BcnI from Bacillus centrosporus.
<3>Eur. J. Biochem.
<4>121
<5>377-381
<6>1982
<7>A specific type-II restriction endonuclease BcnI from Bacillus centrosporus has
been purified to electrophoretic homogeneity in three chromatographic steps.
Around 15 micrograms of such a preparation can be isolated from 1g of the cell
paste.  The yield of the enzyme is higher than that of any type-II restriction
endonuclease so far reported.The molecular weight of the enzyme determined by
gel filtration and polyacrylamide gel electrophoresis in the presence of sodium
dodecyl sulphate equals 27500 and 28000 respectively.  The activity of the
restriction endonuclease is maximal at pH 9.2 and 40-45C.  The optimal
magnesium concentration was estimated to be 7.5mM.  The activity of BcnI may
also be observed in the presence of Co2+, Mn2+, Ni2+ and Zn2+ but it is
markedly less than in the presence of Mg2+.

<>

<1>Petrusyte, M.P., Janulaitis, A.
<2>Specific methylase from Bacillus centrosporus.
<3>Bioorg. Khim.
<4>7
<5>1885-1887
<6>1981
<7>A new site-specific methylase, BcnI, has been isolated from the Bacillus centrosporus strain.
The enzyme recognizes the sequence 5'CmC (C/G)GG in double-stranded DNA and methylating the
underlined cytosine residue.

<>

<1>Pettersson, B.M., Behra, P.R., Manduva, S., Das S, D.S., Bhattacharya, A., Kirsebom, L.A.
<2>Draft Genome Sequence of Saccharopolyspora rectivirgula.
<3>Genome Announcements
<4>2
<5>e01117-13
<6>2014
<7>We have sequenced the genome of Saccharopolyspora rectivirgula, the causative agent of
farmer's lung disease. The draft genome consists of 182 contigs totaling
3,977,051 bp, with a GC content of 68.9%.

<>

<1>Petty, N.K., Bulgin, R., Crepin, V.F., Cerdeno-Tarraga, A.M., Schroeder, G.N., Quail, M.A., Lennard, N., Corton, C., Barron, A., Clark, L., Toribio, A.L., Parkhill, J., Dougan, G., Frankel, G., Thomson, N.R.
<2>The Citrobacter rodentium genome sequence reveals convergent evolution with human pathogenic Escherichia coli.
<3>J. Bacteriol.
<4>192
<5>525-538
<6>2010
<7>Citrobacter rodentium (formally Citrobacter freundii biotype 4280) is a
highly infectious pathogen that causes colitis and transmissible colonic
hyperplasia in mice. In common with enteropathogenic and enterohemorrhagic
Escherichia coli (EPEC and EHEC, respectively), C. rodentium exploits a
type III secretion system (T3SS) to induce attaching and effacing (A/E)
lesions that are essential for virulence. Here, we report the fully
annotated genome sequence of the 5.3-Mb chromosome and four plasmids
harbored by C. rodentium strain ICC168. The genome sequence revealed key
information about the phylogeny of C. rodentium and identified 1,585 C.
rodentium-specific (without orthologues in EPEC or EHEC) coding sequences,
10 prophage-like regions, and 17 genomic islands, including the locus for
enterocyte effacement (LEE) region, which encodes a T3SS and effector
proteins. Among the 29 T3SS effectors found in C. rodentium are all 22 of
the core effectors of EPEC strain E2348/69. In addition, we identified a
novel C. rodentium effector, named EspS. C. rodentium harbors two type VI
secretion systems (T6SS) (CTS1 and CTS2), while EHEC contains only one
T6SS (EHS). Our analysis suggests that C. rodentium and EPEC/EHEC have
converged on a common host infection strategy through access to a common
pool of mobile DNA and that C. rodentium has lost gene functions
associated with a previous pathogenic niche.

<>

<1>Pfeifer, G.P., Kohlmaier, L., Tomassetti, A., Schleicher, R., Follmann, H., Pfohl-Leszkowicz, A., Dirheimer, G., Drahovsky, D.
<2>Polypeptide composition and an immunological analysis of DNA methyltransferases from different species.
<3>Arch. Biochem. Biophys.
<4>268
<5>388-392
<6>1989
<7>The cross-reactivity of the monoclonal anti-human placental DNA
methyltransferase antibody M2B10 with DNA methyltransferases isolated from
other species was investigated.  This antibody immunoprecipitates DNA
methyltransferases from mammalian cells, i.e., human placenta, mouse P815
cells, and rat liver cells.  No cross-reactivity is observed with DNA
methyltransferases from wheat germ and with bacterial DNA methyltransferases
HpaII and EcoRI.  The mammalian enzymes are characterized by polypeptides of
molecular mass 150-190 kDa.  Polypeptides smaller than 190 kDa are presumably
generated by proteolysis of the native 190-kDa DNA methyltransferase.  Trypsin
digestion of the 190-kDa polypeptide isolated from mouse cells results in
progressive appearance of DNA methyltransferase polypeptides of 150-190, 110,
100, and 52-60 kDa.

<>

<1>Pfeiffer, F., Schuster, S.C., Broicher, A., Falb, M., Palm, P., Rodewald, K., Ruepp, A., Soppa, J., Tittor, J., Oesterhelt, D.
<2>Evolution in the laboratory: The genome of Halobacterium salinarum strain R1 compared to that of strain NRC-1.
<3>Genomics
<4>91
<5>335-346
<6>2008
<7>We report the sequence of the Halobacterium salinarum strain R1 chromosome and its four
megaplasmids. Our set of protein-coding genes is supported by
extensive proteomic and sequence homology data. The structures of the
plasmids, which show three large-scale duplications (adding up to 100 kb),
were unequivocally confirmed by cosmid analysis. The chromosome of strain
R1 is completely colinear and virtually identical to that of strain NRC-1.
Correlation of the plasmid sequences revealed 210 kb of sequence that
occurs only in strain R1. The remaining 350 kb shows virtual sequence
identity in the two strains. Nevertheless, the number and overall
structure of the plasmids are largely incompatible. Also, 20% of the
protein sequences differ despite the near identity at the DNA sequence
level. Finally, we report genome-wide mobility data for insertion
sequences from which we conclude that strains R1 and NRC-1 originate from
the same natural isolate. This exemplifies evolution in the laboratory.

<>

<1>Pfleiderer, A., Mishra, A.K., Lagier, J.C., Robert, C., Caputo, A., Raoult, D., Fournier, P.E.
<2>Non-contiguous finished genome sequence and description of Alistipes ihumii sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1221-1235
<6>2014
<7>Alistipes ihumii strain AP11(T) sp. nov. is the type strain of A. ihumii sp. nov., a new
species within the genus Alistipes. This strain, whose genome is
described here, was isolated from the fecal flora of a 21-year-old French
Caucasian female, suffering from a severe restrictive form of anorexia nervosa
since the age of 12 years. A. ihumii is a Gram-negative anaerobic bacillus. Here
we describe the features of this organism, together with the complete genome
sequence and annotation. The 2,753,264 bp long genome (one chromosome but no
plasmid) contains 2,254 protein-coding and 47 RNA genes, including 3 rRNA genes.

<>

<1>Pfreundt, U., Stal, L.J., Voss, B., Hess, W.R.
<2>Dinitrogen fixation in a unicellular chlorophyll d-containing cyanobacterium.
<3>ISME J.
<4>6
<5>1367-1377
<6>2012
<7>Marine cyanobacteria of the genus Acaryochloris are the only known organisms that
use chlorophyll d as a photosynthetic pigment. However, based on chemical
sediment analyses, chlorophyll d has been recognized to be widespread in oceanic
and lacustrine environments. Therefore it is highly relevant to understand the
genetic basis for different physiologies and possible niche adaptation in this
genus. Here we show that unlike all other known isolates of Acaryochloris, the
strain HICR111A, isolated from waters around Heron Island, Great Barrier Reef,
possesses a unique genomic region containing all the genes for the structural and
enzymatically active proteins of nitrogen fixation and cofactor biosynthesis.
Their phylogenetic analysis suggests a close relation to nitrogen fixation genes
from certain other marine cyanobacteria. We show that nitrogen fixation in
Acaryochloris sp. HICR111A is regulated in a light-dark-dependent fashion. We
conclude that nitrogen fixation, one of the most complex physiological traits
known in bacteria, might be transferred among oceanic microbes by horizontal gene
transfer more often than anticipated so far. Our data show that the two powerful
processes of oxygenic photosynthesis and nitrogen fixation co-occur in one and
the same cell also in this branch of marine microbes and characterize
Acaryochloris as a physiologically versatile inhabitant of an ecological niche,
which is primarily driven by the absorption of far-red light.

<>

<1>Phale, P.S., Paliwal, V., Raju, S.C., Modak, A., Purohit, H.J.
<2>Genome Sequence of Naphthalene-Degrading Soil Bacterium Pseudomonas putida CSV86.
<3>Genome Announcements
<4>1
<5>e00234-12
<6>2013
<7>CSV86, a soil isolate, preferentially utilizes naphthalene over glucose as a source of carbon
and energy. We present the draft genome sequence, which is 6.4
Mb in size; analysis suggests the chromosomal localization of genes coding for
naphthalene utilization. The operons coding for glucose and other aromatic
compounds might also be annotated in another study.

<>

<1>Phalke, S., Nickel, O., Walluscheck, D., Hortig, F., Onorati, M.C., Reuter, G.
<2>Retrotransposon silencing and telomere integrity in somatic cells of Drosophila depends on the cytosine-5 methyltransferase DNMT2.
<3>Nat. Genet.
<4>41
<5>696-702
<6>2009
<7>Here we show that the cytosine-5 methyltransferase DNMT2 controls retrotransposon silencing in
Drosophila somatic cells. In Drosophila,
significant DNMT2-dependent DNA methylation occurs during early
embryogenesis. Suppression of white gene silencing by Mt2 (Dnmt2) null
mutations in variegated P[w(+)] element insertions identified functional
targets of DNMT2. The enzyme controls DNA methylation at retrotransposons
in early embryos and initiates histone H4K20 trimethylation catalyzed by
the SUV4-20 methyltransferase. In somatic cells, loss of DNMT2 eliminates
H4K20 trimethylation at retrotransposons and impairs maintenance of
retrotransposon silencing. In Dnmt2 and Suv4-20 null genotypes,
retrotransposons are strongly overexpressed in somatic but not germline
cells, where retrotransposon silencing depends on an RNAi mechanism. DNMT2
also controls integrity of chromosome 2R and 3R telomeres. In Dnmt2 null
strains, we found stable loss of the subtelomeric clusters of defective
Invader4 elements. Together, these results demonstrate a previously
unappreciated role of DNA methylation in retrotransposon silencing and
telomere integrity in Drosophila.

<>

<1>Pham, T.T., Jacobs-Sera, D., Pedulla, M.L., Hendrix, R.W., Hatfull, G.F.
<2>Comparative genomic analysis of mycobacteriophage Tweety: evolutionary insights and construction of compatible site-specific integration vectors for mycobacteria.
<3>Microbiology
<4>153
<5>2711-2723
<6>2007
<7>Mycobacteriophage Tweety is a newly isolated phage of Mycobacterium
smegmatis. It has a viral morphology with an isometric head and a long
flexible tail, and forms turbid plaques from which stable lysogens can
be
isolated. The Tweety genome is 58 692 bp in length, contains 109
protein-coding genes, and shows significant but interrupted nucleotide
sequence similarity with the previously described mycobacteriophages
Llij,
PMC and Che8. However, overall the genome possesses mosaic architecture,
with gene products being related to other mycobacteriophages such as
Che9d, Omega and Corndog. A gene encoding an integrase of the
tyrosine-recombinase family is located close to the centre of the
genome,
and a putative attP site has been identified within a short intergenic
region immediately upstream of int. This Tweety attP-int cassette was
used
to construct a new set of integration-proficient plasmid vectors that
efficiently transform both fast- and slow-growing mycobacteria through
plasmid integration at a chromosomal locus containing a tRNA(Lys) gene.
These vectors are maintained well in the absence of selection and are
completely compatible with integration vectors derived from
mycobacteriophage L5, enabling the simple construction of complex
recombinants with genes integrated simultaneously at different
chromosomal
positions.

<>

<1>Phelippeau, M., Robert, C., Croce, O., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium neoaurum Strain DSM 44074T.
<3>Genome Announcements
<4>2
<5>e00699-14
<6>2014
<7>We report the draft genome sequence of Mycobacterium neoaurum strain DSM 44074(T), a
nontuberculosis species responsible for opportunistic infections in
immunocompromised patients. The strain described here is composed of 5,536,033
bp, with a G+C content of 66.24%, and carries 5,274 protein-coding genes and 72
RNA genes.

<>

<1>Phiasivongsa, P., Redkar, S.G., Gamage, S., Brooke, D., Denny, W., Bearss, D.J., Vankayalapati, H., Xu, Y., Swierczek, K.
<2>Quinoline derivatives for modulating DNA methylation.
<3>US Patent Office
<4>US 20090285772
<5>
<6>2009
<7>Quinoline derivatives, particularly 4-anilinoquinoline derivatives, are provided.  Such
quinoline derivatives can be used for modulation of DNA methylation, such as effective
inhibition of methylation of cytosine at the C-5 position, for example via selective
inhibition of DNA methyltransferase DNMT1.  Methods for synthesizing numerous
4-anilinoquinoline derivatives and for modulating DNA methylation are provided.  Also provided
are methods for formualting and administering those compounds or compositions to treat
conditions such as cancer and hematological disorders.

<>

<1>Philippe, N., Legendre, M., Doutre, G., Coute, Y., Poirot, O., Lescot, M., Arslan, D., Seltzer, V., Bertaux, L., Bruley, C., Garin, J., Claverie, J.M., Abergel, C.
<2>Pandoraviruses: amoeba viruses with genomes up to 2.5 Mb reaching that of parasitic eukaryotes.
<3>Science
<4>341
<5>281-286
<6>2013
<7>Ten years ago, the discovery of Mimivirus, a virus infecting Acanthamoeba,
initiated a reappraisal of the upper limits of the viral world, both in terms of
particle size (>0.7 micrometers) and genome complexity (>1000 genes), dimensions
typical of parasitic bacteria. The diversity of these giant viruses (the
Megaviridae) was assessed by sampling a variety of aquatic environments and their
associated sediments worldwide. We report the isolation of two giant viruses, one
off the coast of central Chile, the other from a freshwater pond near Melbourne
(Australia), without morphological or genomic resemblance to any previously
defined virus families. Their micrometer-sized ovoid particles contain DNA
genomes of at least 2.5 and 1.9 megabases, respectively. These viruses are the
first members of the proposed "Pandoravirus" genus, a term reflecting their lack
of similarity with previously described microorganisms and the surprises expected
from their future study.

<>

<1>Phillips, K.E., Schipma, M.J., Satchell, K.J.
<2>Draft Genome Sequence of Israeli Outbreak-Associated Vibrio vulnificus Biotype 3  Clinical Isolate BAA87.
<3>Genome Announcements
<4>2
<5>e00032-14
<6>2014
<7>Vibrio vulnificus is a seafood-associated pathogen that causes severe wound and intestinal
infections. Biotype 3 of V. vulnificus emerged in 1996 as the cause of
an Israeli outbreak associated with the handling of infected tilapia. Here, we
describe the whole-genome sequence of the ATCC biotype 3 clinical isolate BAA87
(CDC9530-96).

<>

<1>Phillips, P.L.
<2>Cloning and characterization of a methyl-dependent restriction endonuclease and a cell cycle regulating DNA methyltransferase from Zymomonas mobilis subspecies mobilis CP4.
<3>Ph.D. Thesis, Univ. of Florida, Gainesville
<4>
<5>1-220
<6>2005
<7>A Zymomonas mobilis CP4 genomic library was screened using the E. coli indicator strains
AP1-200-9 and ER1992 to isolate clones of enzymes that cause DNA damage.  Sequence analysis of
positive clones identified two open reaading frames encoding DNA modification enzymes: (a) a
924 base pair open reading frame with sequence similarity to mrr, a methyl-dependant
restriction endonuclease, which was designated ZmCP4mrr, and (2) a 1149 bp open reading frame
with amino acid sequence similarity to ccrM, a cell cycle regulating DNA methyltransferase,
which was designated ZmCP4ccM.  Sequence analysis indicates that ZmCP4mrr is a solitary
methyl-dependent restriction endonuclease without a cognate DNA methyltransferase.
Transformation of Eschichia coli K12 strains with various DNA methyltransferase backgrounds
demonstrated that a plasmid borne ZmCP4mrr gene readily transforms E. coli strains that
experess dcm, hsdM, and EcoKccrM DNA methyltransferases, indicating that ZmCP4Mrr does not
recognize and restrict sites methylated by these DNA methyltransferases.  E. coli strains that
express the dam DNA methyltransferase could only be transformed if expression of plasmid borne
ZmCP4mrr was repressed.  Subsequent induction of ZmCP4mrr expression in these cells resulted
in inhibition of growth and cell death, indicating that ZmCP4Mrr specifically restricts Dam
N6-adenine methylated DNA (5'-GmATC-3').  Plasmid DNA originating from dam deficient E. coli
strains did not improve transformation efficiency, indicating that Z. mobilis CP4 has a
restriction system in addition to ZmCP4Mrr contributing to low frequency of gene transfer from
foreign DNA.  Sequence analysis indicates that AmCP4ccrM is a solitary DNA methyltransferase
with two possible in-frame translation initiation sites.  A ribosomal binding site containing
a sequence, 5'-AGGA-3', conserved in Z. mobilis promoters of highly expressed genes is
located adjacent to the first possible translation initiation site and not the second,
suggesting that ZmCP4ccrM is being expressed from the first translational initiation site. The
specificity for ZmCP4CcrM methylation was directly determined to be the N6-adenine of its
recognition site 5'-GANTC-3'.  Z. mobilis CP4 cells overexpressing ZmCP4ccrM exhibited a
subpopulation of filamentous cells, ranging from 10-90 uM in length, with multiple
chromosomes.  Overexpression of ZmCP4ccrM caused disruption of normal cell division and
chromosomal segregation, suggesting that ZmCP4CcrM is involved in cell cycle regulation.

<>

<1>Phillips, S.E.V.
<2>Induced flip.
<3>Nat. Struct. Biol.
<4>1
<5>76-77
<6>1994
<7>Hhal methyltransferase, caught in the act of methylating a cytosine on a DNA substrate,
reveals how the enzyme overcomes the problem of chemically modifying bases in the relatively
inaccessible environment of the DNA duplex.

<>

<1>Phung, L.T., Silver, S., Trimble, W.L., Gilbert, J.A.
<2>Draft Genome of Halomonas Species Strain GFAJ-1 (ATCC BAA-2256).
<3>J. Bacteriol.
<4>194
<5>1835-1836
<6>2012
<7>Halomonas strain GFAJ-1 was reported in Science magazine to be a remarkable microbe for which
there was 'arsenate in macromolecules that normally contain phosphate, most notably nucleic
acids.' The draft genome of the bacterium was determined (NCBI accession numbers AHBC01000001
through AHBC01000103). It appears to be a typical gamma proteobacterium.

<>

<1>Phung, L.T., Trimble, W.L., Meyer, F., Gilbert, J.A., Silver, S.
<2>Draft Genome Sequence of Alcaligenes faecalis subsp. faecalis NCIB 8687 (CCUG 2071).
<3>J. Bacteriol.
<4>194
<5>5153
<6>2012
<7>Alcaligenes faecalis subsp. faecalis NCIB 8687, the betaproteobacterium from which arsenite
oxidase had its structure solved and the first 'arsenate gene
island' identified, provided a draft genome of 3.9 Mb in 186 contigs (with the
largest 15 comprising 90% of the total) for this opportunistic pathogen species.

<>

<1>Picardeau, M. et al.
<2>Genome sequence of the saprophyte Leptospira biflexa provides insights into the evolution of Leptospira and the pathogenesis of leptospirosis.
<3>PLoS ONE
<4>3
<5>e1607
<6>2008
<7>Leptospira biflexa is a free-living saprophytic spirochete present in aquatic environments. We
determined the genome sequence of L. biflexa,
making it the first saprophytic Leptospira to be sequenced. The L. biflexa
genome has 3,590 protein-coding genes distributed across three circular
replicons: the major 3,604 chromosome, a smaller 278-kb replicon that also
carries essential genes, and a third 74-kb replicon. Comparative sequence
analysis provides evidence that L. biflexa is an excellent model for the
study of Leptospira evolution; we conclude that 2052 genes (61%) represent
a progenitor genome that existed before divergence of pathogenic and
saprophytic Leptospira species. Comparisons of the L. biflexa genome with
two pathogenic Leptospira species reveal several major findings. Nearly
one-third of the L. biflexa genes are absent in pathogenic Leptospira. We
suggest that once incorporated into the L. biflexa genome, laterally
transferred DNA undergoes minimal rearrangement due to physical
restrictions imposed by high gene density and limited presence of
transposable elements. In contrast, the genomes of pathogenic Leptospira
species undergo frequent rearrangements, often involving recombination
between insertion sequences. Identification of genes common to the two
pathogenic species, L. borgpetersenii and L. interrogans, but absent in L.
biflexa, is consistent with a role for these genes in pathogenesis.
Differences in environmental sensing capacities of L. biflexa, L.
borgpetersenii, and L. interrogans suggest a model which postulates that
loss of signal transduction functions in L. borgpetersenii has impaired
its survival outside a mammalian host, whereas L. interrogans has retained
environmental sensory functions that facilitate disease transmission
through water.

<>

<1>Picchi, S.C., Vilas-Boas, L.A., Ceresini, P.C., de Macedo-Lemos, E.G., Lemos, M.V.
<2>Strain variability in the DNA immigration control region (ICR) of Xylella fastidiosa.
<3>Res. Microbiol.
<4>157
<5>254-262
<6>2006
<7>The genome of the bacterium Xylella fastidiosa contains four ORFs (XF2721, XF2725, XF2739 and
XF0295) related to the restriction
modification type I system, ordinarily named R-M. This system belongs
to the DNA immigration control region (ICR). Each CIRF is related to
different operon structures, which are homologues among themselves and
with subunit Hsd R from the endonuclease coding genes. In addition,
these ORFs are highly homologous to genes in Pseudomonas aeruginosa,
Methylococcus capsulatus str. Bath, Legionella pneumophila,
Helicobacter pylori, Xanthomonas oryzae pv. Oryzae and Silicibacter
pomeroyi, as well as to genes from X. fastidiosa strains that infect
grapevine, almond and oleander plants. This study was carried out on
R-M ORFs from forty-three X. fastidiosa strains isolated from citrus,
coffee, grapevine, periwinkle, almond and plum trees, in order to
assess the genetic diversity of these loci through PCR-RFLP. PCR-RFLP
analysis of the four ORFs related to the R-M system from these strains
enabled the detection of haplotypes for these loci. When the haplotypes
were defined, wide genetic diversity and a large range of similar
strains originating from different hosts were observed. This analysis
also provided information indicating differences in population genetic
structures, which led to detection of different levels of gene transfer
among the groups of strains.

<>

<1>Piechaczyk, M., Jeanteur, P., Louarn, J.-M.
<2>An easy method for the selection of restriction- and modification-deficient mutants of Escherichia coli K-12.
<3>Gene
<4>11
<5>173-175
<6>1980
<7>An easy and rapid method for selecting restriction- and modification-
defective mutants of
Escherichia coli K-12 is described. This method employs selection of tetracycline-resistant
lysogens after
infection with lambda::Tn10 phage and results in a high yield of spontaneous rk-mk- and
rk-mk- mutants.

<>

<1>Piechula, S., Kim, S.C., Podhajska, A.J.
<2>PamI and PamII restriction endonucleases from Phormidium ambiguum.
<3>Nucleic Acids Res.
<4>20
<5>619
<6>1992
<7>PamI and PamII are type II restriction endonucleases from the Cyanobacterial
strain Phormidium ambiguum GOM (CCALA Hindak 1965/117).  PamI and PamII are
isoschizomers of MstI and AcyI respectively.  The enzymes were purified using
two chromatographic steps: 1) phosphocellulose, 2) DEAE-sephadex G-25.  The
enzymes were free of contaminating nuclease activity.  All digestions were
performed at 37C in a buffer containing 50 mM NaCl, 10 mM Tris-HCl (pH 7.5), 10
mM MgCl2.

<>

<1>Piechula, S., Kur, J., Bielawski, K., Podhajska, A.J.
<2>Isolation and identification of the restriction endonuclease PtaI from Phormidium tadzschicicum, an isoschizomer of BspMII.
<3>Nucleic Acids Res.
<4>20
<5>6738
<6>1992
<7>PtaI is a type II restriction endonuclease from the cyanobacterial strain Phormidium
tadzschicicum. PtaI recognizes the sequence TCCGGA and cleaves between T and C. It is an
isoschizomer of BspMII.

<>

<1>Piechula, S., Kur, J., Woszczyk, J., Podhajska, A.J.
<2>Purification and characterization of two restriction endonucleases isolated from Phormidium inundatum.
<3>Gene
<4>157
<5>315-316
<6>1995
<7>We have isolated two restriction endonucleases, PinBI and PinBII, from the cyanobacterial
strain Phormidium inundatum, and identified them as isoschizomers of AvaIII and BspMII,
respectively.

<>

<1>Piechula, S., Piosik, J., Bielawski, K., Podhajska, A.J.
<2>Isolation and characterization of the restriction endonuclease PpeI from Phormidium persicinum.
<3>Mol. Biotechnol.
<4>5
<5>97-99
<6>1996
<7>PpeI is a type II restriction endonuclease isolated from cyanobacterial strain
Phormidium persicinum.  The endonuclease PpeI, an isoschizomer of ApaI, recognizes the
hexanucleotide sequence (5'-GGGCC/C-3') and cleaves, after the second C, producing
four nucleotide 3'-cohesive ends.

<>

<1>Piechula, S., Skowron, P.M., Piatyszek, M., Podhajska, A.J.
<2>Isolation and identification of two new Synechococcus-derived restriction endonucleases, SleI and SspAI isoschizomers of EcoRII.
<3>Nucleic Acids Res.
<4>19
<5>2782
<6>1991
<7>Two new type-II restriction endonucleases, SleI and SspAI, have been isolated
from Synechococcus leopoliensis.  Strain 1402-1 was obtained from the Institut
Pasteur Culture Collection of Cyanobacterial Strains.  Synechococcus sp. AN6301
was obtained from Pflanzenphysiologisches Institut, Universitat Gottingen,
Nikolausberg, FRG.  Both strains were grown aerobically at 30C in BG medium
under fluorescent light.  The cells were harvested and stored in liquid
nitrogen.  Frozen cells were disrupted by sonication.  Purification of the
enzymes was carried out by the following steps: (I) DEAE-cellulose
chromatography (II) DNA-cellulose chromatography, (III) QAE-Sephadex
chromatography.  The digestion pattern of pBR322 DNA with BstNI was identical
to the patterns obtained with SleI and SspAI, indicating that SleI and SspAI
are isoschizomers of BstNI, and therefore, of its isoschizomer EcoRII.  It was
found that SleI and SspAI enzymes generate 5 nucleotide (nt) cohesive ends, as
assessed by digestion of lambda DNA and fill-in reaction with T7 DNA
polymerase, [alpha-35S]dATP and the other dNTPs.  EcoRII and BstNI recognize
the same sequence, but cut between different nt generating 5-nt and 1-nt
cohesive ends, respectively.  Generally following the approach described by
Brown et al., we have directly determined the sequence of the cleavage site of
SleI and SspAI in M13mp18 double stranded DNA.  The first restriction site
recognized by EcoRII is located 167 nt from the first nt of the 17-mer
sequencing primer, and thus can easily be sequenced.  We found that the SleI
and SspAI cut sites (represented by arrows) are shifted by two nt (Fig. 2)
within the recognition site with respect to the BstNI cuts (represented by
dots), and thus are identical to the EcoRII cuts.

<>

<1>Piechula, S., Waleron, K., Swiatek, W., Biedrzycka, I., Podhajska, A.J.
<2>Mesophilic cyanobacteria producing thermophilic restriction endonucleases.
<3>FEMS Microbiol. Lett.
<4>198
<5>135-140
<6>2001
<7>When searching for the site-specific endonucleases in several strains of Phormidium we made
the following observations. Among the 16 strains
that originated from 15 species of Phormidium. 12 produced one or more
restriction enzymes, of which two produced the highly thermophilic
restriction endonucleases PtaI and PpaAII with their optimum activity
at 65-80 C, which is far above the lethal temperature for the
host microorganism (40 C). These two temperature-resistant
enzymes are isoschizomers of known BspMII and TaqI endonucleases,
respectively. The presence of the thermophilic TaqI isoschizomer does
not seem to play any role in the mesophilic host microorganism, which
does not even contain an active cognate methyltransferase. Among the
remaining 10 strains, six produced isoschizomers of endonucleases which
we first described in cyanobacteria, namely: PfuAII (NdeI), PinBII and
PtaI(BspMII). PlaAII (RsaI), PpaAII PpeI (ApaI). Two enzymes, PauAII
(AhaIII) and PfaAII (NdeI), belong to a group of a very rarely
occurring isoschizomers. Out of 21 cyanobacterial endonucleases
investigated by us, Four were active in a wide range of temperatures
(from 15 to 60 C) which also extended the optimal growth
temperature of the hosts. We assume that our observation on the
presence of temperature-resistant restriction enzymes in mesophilic
hosts supports the idea of horizontal gene transfer. Restriction
modification systems may be an excellent tool for investigation of that
phenomenon.

<>

<1>Piekarowicz, A.
<2>HineI is an isoschizomer of HinfIII restriction endonuclease.
<3>J. Mol. Biol.
<4>157
<5>373-381
<6>1982
<7>HineI is a restriction enzyme isolated from Haemophilus influenzae strain Re.
Like other type III restriction endonucleases it requires ATP for cleavage and
S-adenosyl-methionine for methylation of DNA.  This enzyme recognises the same
sequence as HinfIII (Piekarowicz et al., 1981) and cleaves and methylates DNA
in a manner similar to all type III restriction enzymes.

<>

<1>Piekarowicz, A.
<2>The influence of methionine deprivation on restriction properties of Haemophilus influenzae Rd and Ra strains.
<3>Acta Microbiol. Pol. A
<4>6
<5>71-74
<6>1974
<7>The influence of methionine starvation on the restriction properties of H.
influenzae Rd and Ra has been examined.  It was shown that the methionine
deprivation of Rd and Ra cells does not change their capacity to restrict HPlcl
phage.  These results suggest that S-adenosylmethionine may not be required for
the action of H. influenzae Rd and Ra restriction endonucleases.

<>

<1>Piekarowicz, A.
<2>Preferential cleavage by restriction endonuclease HinfIII.
<3>Acta Biochim. Pol.
<4>31
<5>453-464
<6>1984
<7>The efficiency of endonucleolytic scission by restriction endonuclease HinfIII
varies markedly for different recognition sites.  The relative frequencies of
cleavage at these sites have been determined on the basis of analysis of
specific unit length linear molecules formed.  The efficiency of restriction
reaction depends also on the number of recognition sites in the DNA substrate.
Cleavage by HinfIII in the absence or presence of S-adenosylmethionine is
observed only when at least three recognition sites are present.  HinfIII also
shows preferential methylation of certain sites observable even for a substrate
with one recognition site.  The nucleotide sequences at sites cleaved or
methylated at high frequency have been compared.

<>

<1>Piekarowicz, A.
<2>Identification of a new restriction endonuclease R.NciII, from Neisseria cinerea.
<3>Acta Microbiol. Pol.
<4>43
<5>103-105
<6>1994
<7>Site-specific restriction endonuclease R.NciII has been purified from Neisseria cinerea strain
32615. The enzyme recognizes the sequence 5'GATC3' and its activity is inhibited by the
presence of methylated adenine residue within the recognition sequence.

<>

<1>Piekarowicz, A.
<2>DNA methyltransferases of Neisseria gonorrhoeae.
<3>Acta Microbiol. Pol.
<4>43
<5>269-277
<6>1994
<7>The DNA of both prokaryotic and eukaryotic organisms can undergo postreplicative modification.
The most widely known type of modification is the addition of the methyl groups to either
cytosine or adenine residues.  This process is carried out by the enzymes called DNA
methyltransferases or MTases.  Methylation by all types of MTases requires
S-adenosylmethionine (SAM).  The function of SAM is in most of the cases limited to serving as
a methyl group donor.  In the case of Escherichia coli Dam MTase, however, it also affects
binding of the enzyme to the target site.

<>

<1>Piekarowicz, A., Baj, J.
<2>Host specificity of DNA in Haemophilus influenzae:  The physiological and genetical bases of instability of restriction and modification of DNA in strain RD.
<3>Acta Microbiol. Pol. A
<4>8
<5>119-130
<6>1975
<7>Further investigations of the instability of restriction and modification
properties of H. influenzae Rd strain were carried out.  It has been shown that
the instable properties of hsd HindI system are maintained even after transfer
of this system to another H. influenzae strain.  The expression of hsd HindI
system is very sensitive to various physiological changes which do not
influence the other hsd systems present in the same Rd strain.  The instability
of hsd HindI system is postulated to be connected with some regulator gene(s).

<>

<1>Piekarowicz, A., Bickle, T.A., Shepherd, J.C.W., Ineichen, K.
<2>The DNA sequence recognised by the HinfIII restriction endonuclease.
<3>J. Mol. Biol.
<4>146
<5>167-172
<6>1981
<7>HinfIII is a type III restriction enzyme (Kaue & Piekarowicz, 1978) isolated
from Haemophilus influenzae Rf.  Like other type III restriction endonucleases,
the enzyme also catalyses the modification of susceptible DNA.  It requires ATP
for DNA cleavage and S-adenosyl methionine for DNA methylation.  We have
determined the DNA sequence recognised by HinfIII to be: 5'-C-G-A-A-T-3'
3'-G-C-T-T-A-5' In restriction, the enzyme cleaves the DNA about 25 base-pairs
to the right of this sequence.  In the modification reaction only one of the
strands is methylated, that containing the 5'-C-G-A-A-T-3' sequence.

<>

<1>Piekarowicz, A., Brzezinski, R.
<2>Cleavage and methylation of DNA by the restriction endonuclease HinfIII isolated from Haemophilus influenzae Rf.
<3>J. Mol. Biol.
<4>144
<5>415-429
<6>1980
<7>HinfIII is a restriction enzyme isolated from Haemophilus influenzae strain Rf. It requires
ATP for cleavage and S-adenosyl-methionine for methylation of DNA. This enzyme can be present
in two forms: one with AdoMet bound to it, and a second form free of this cofactor.  In the
presence of AdoMet and ATP the enzyme cleaves ColE1 DNA molecules once, to produce unit-length
linear molecules.  The HinfIII endonuclease cleaves at unique sites, though not every site on
every molecule is cut.  The five HinfIII cleavage sites were mapped relative to the EcoRI
restriction endonuclease cleavage site.  If AdoMet is omitted from the enzyme reaction
mixture, the second form of HinfIII enzyme cleaves ColE1 DNA into several fragments.  An
average of 6.2 +/- 1 methyl groups are transferred to ColE1 DNA from AdoMet.  The methyl
groups were mapped relative to the HaeIII restriction endonuclease fragments.  The position of
methylation sites correlates well with the cleavage sites. The restriction activity of the
HinfIII enzyme shows some dependence upon the structure of the substrate DNA.  The linear
molecule of ColE1 DNA is a poorer substrate than the supercoiled molecules.  Lambda DNA
fragments with molecular weights smaller than approximately 2,000,000 are not cleaved by
HinfIII enzyme, but since they can be methylated the enzyme is able to recognize the specific
sequences on these fragments.

<>

<1>Piekarowicz, A., Brzezinski, R., Kauc, L.
<2>Host specificity of DNA in Haemophilus influenzae: DNA restriction enzyme from H. Influenzae Rf232.
<3>Acta Microbiol. Pol.
<4>25
<5>307-312
<6>1976
<7>A restriction endonuclease has been partially purified from Haemophilus
influenzae Rf232 containing the genetically determined system of restriction
and modification of DNA.  The enzyme requires ATP for the degradation of
transfecting phage DNA.

<>

<1>Piekarowicz, A., Brzezinski, R., Kauc, L.
<2>Host Specificity of DNA in Haemophilus influenzae:  The in vivo Action of the Restriction Endonucleases on Phage and Bacterial DNA.
<3>Acta Microbiol. Pol. A
<4>7
<5>51-65
<6>1975
<7>In Haemophilus influenzae strains only the type 1 of the restriction
endonucleases have an in vivo effect on phage and bacterial transforming DNA.
The type 2 of restriction endonucleases which act very efficiently in vitro are
completely inactive in vivo.  The reasons for this inactivity is unknown.

<>

<1>Piekarowicz, A., Bujnicki, J.
<2>Cloning of the Dam methyltransferase gene from Haemophilus influenzae bacteriophage HP1.
<3>Acta Microbiol. Pol.
<4>48
<5>123-129
<6>1999
<7>The putative product of orf13 from the genome of Haemophilus influenzae HP1 bacteriophage
shows homology only to bacteriophage T1 Dam methyltransferase, and a weak similarity to the
conserved amino acids sequence motifs characteristic of m6A-methyltransferases.  Especially
interesting is the lack of characteristic motif I responsible for binding of
S-adenosylmethionine.  Despite this fact, a DNA sequence of HP1 bacteriophage of Haemophilus
influenzae encoding methyltransferase activity was cloned and expressed in Escherichia coli
using pMPMT4omega expression vector.  The cloned methyltransferase recognizes the sequence
5'-GATC-3' and methylates an adenine residue.  The enzyme methylates both double- and
single-stranded DNA substrates.

<>

<1>Piekarowicz, A., Glover, S.W.
<2>Host specificity of DNA in Haemophilus influenzae:  the two restriction and modification systems in strain Ra.
<3>Mol. Gen. Genet.
<4>116
<5>11-25
<6>1972
<7>Rough R strains of Haemophilus influenzae derived from the smooth (S) serotypes
Sa, Sb, Sd, Se and Sf each carry DNA restriction and modification systems.  The
DNA host specificity determined by Re and Rf may be the same but is different
from that for Ra, Rb and Rd all of which can be distinguished from one another.
Strain Ra carries two genetically distinct host specificity systems Al and A2
each of which is able to restrict Haemophilus phage HP1, and each of which
confers a specific modification on phage grown in strain Ra.  Among
restriction-deficient mutants isolated from strain Ra, seven of the eight
possible phenotypes for these two systems were obtained after either one or two
mutational steps.

<>

<1>Piekarowicz, A., Goguen, J.D.
<2>The DNA sequence recognized by the EcoDXXI restriction endonuclease.
<3>Eur. J. Biochem.
<4>154
<5>295-298
<6>1986
<7>EcoDXXI is a type-I restriction enzyme coded for by the plasmid pDXX1.  Like
other type-I restriction endonucleases, the enzyme catalyses the modification
of susceptible DNA.  We have determined the DNA sequence recognised by EcoDXXI
to be:5'-TCANNNNNNNATTC-3' 3'-AGTNNNNNNNTAAG-5'where N can be any nucleotide.
This sequence has an overall structure very similar to previously determined
type-I sequences.

<>

<1>Piekarowicz, A., Goguen, J.D., Skrzypek, E.
<2>The EcoDXXI restriction and modification system of Escherichia coli ET7.  Purification, subunit structure and properties of the restriction endonuclease.
<3>Eur. J. Biochem.
<4>152
<5>387-393
<6>1985
<7>The Escherichia coli plasmid pDXX1 codes for a new restriction-modification system. The
specific restriction endonuclease coded by this system has been purified by a procedure that
includes phosphocellulose and heparin-agarose chromatography. Sedimentation on glycerol
gradients showed one peak of activity with a value of about 12S. The highly purified enzyme
require ATP and Mg2+ for activity as well as S-adenosylmethionine, although some
S-adenosylmethionine molecules are probably bound to the enzyme. The enzyme does not cleave
lambda DNA at well-defined sites and has a strong non-modified DNA-dependent ATPase activity.
The enzyme has also methylase activity acting against non-modified DNA. te is repeated in
inverse orientation. The additional base pair in the non-specific spacer of the mutant
recognition sequence maintains the proper spacing between the two methylatable adenine groups.
Sequencing of both the wild type and mutant EcoDXXI hsdS genes showed that the Tn5 insertion
occurred at nucleotide 673 of the 1221 bp gene. This effectively deletes the entire
carboxyl-terminal DNA binding domain which recognizes the 3' half of the EcoDXXI binding
site. The truncated hsdS gene still encodes both the amino-terminal DNA binding domain and the
conserved repeated sequence that defines the length of the recognition site spacer region. We
propose that the EcoDXXI mutant methylase utilizes two truncated hsdS subunits to recognize
its binding site. The implications of this finding in terms of subunit interactions and the
malleability of the type I R-M systems will be discussed.

<>

<1>Piekarowicz, A., Golaszewska, M., Sunday, A.O., Siwinska, M., Stein, D.C.
<2>The HaeIV restriction modification system of Haemophilus aegyptius is encoded by a single polypeptide.
<3>J. Mol. Biol.
<4>293
<5>1055-1065
<6>1999
<7>The HaeIV restriction endonuclease (ENase) belongs to a distinct class of ENases,
characterized by its ability to cleave double-stranded DNA on both sides of its recognition
sequence, excising a short DNA fragment that includes the recognition sequence. The gene
encoding the HaeIV ENase was cloned from Haemophilus aegyptius into pUC19 using a previously
described system that does not need the knowledge that a particular ENase is produced by a
bacterial strain. DNA sequence analysis of the insert contained on this plasmid identified a
single open reading frame (ORF), with the predicted protein having an apparent molecular mass
of approximately 110 kDa. The protein encoded by this ORF was purified to homogeneity from
Escherichia coli strain ER1944 carrying the haeIVRM gene on a recombinant plasmid under the
control of the inducible ara promoter. The protein possessed both ENase and methyltransferase
(MTase) activities. Amino acid sequence analysis was able to identify several conserved motifs
found in DNA MTases, located in the middle of the protein. The enzyme recognizes the
interrupted palindromic sequence 5' GAPyNNNNNPuTC 3', cleaving double-stranded DNA on both
strands upstream and downstream of the recognition sequence, releasing an approximately 33 bp
fragment. The ENase possessed an absolute requirement only for Mg(+2). ATP had no influence on
ENase or MTase activities. The ENase made the first strand cleavage randomly on either side of
the recognition sequence, but the second cleavage occurred more slowly. The MTase activity
modified symmetrically located adenine residues on both strands within the recognition
sequence yielding N6-methyl adenine. Furthermore, the MTase was active as a dimer.

<>

<1>Piekarowicz, A., Kalinowska, J.
<2>Host specificity of DNA in Haemophilus influenzae: Similarity between host-specificity types of Haemophilus influenzae Re and Rf.
<3>J. Gen. Microbiol.
<4>81
<5>405-411
<6>1974
<7>Strain Re of Haemophilus influenzae carries two genetically distinct
host-specificity systems EI and E2 each of which is able to restrict
Haemophilus phage HPIcI and each of which confers a specific modification upon
phage grown in strain Re.  These two systems are apparently identical to the
host-specificity systems of H. influenzae Rf F1 and F2.

<>

<1>Piekarowicz, A., Kauc, L., Glover, S.W.
<2>Host specificity of DNA in Haemophilus influenzae: The restriction and modification systems in strains Rb and Rf.
<3>J. Gen. Microbiol.
<4>81
<5>391-403
<6>1974
<7>Haemophilus influenzae Rf possesses two distinct host specificity systems FI
and F2 each of which is able to restrict and modify Haemophilus phage HPICI,
while strain Rb posseses only one system, B.  Among restriction-deficient
mutants isolated from strain Rf, the r-m+ as well as r-m- phenotypes for these
two systems were obtained after either one or two mutational steps.  The FI
system was introduced into H. influenzae Rd by genetic transformation to show
that the DI and FI systems are not allelic.

<>

<1>Piekarowicz, A., Klyz, A., Kwiatek, A., Stein, D.C.
<2>Analysis of type I restriction modification systems in the Neisseriaceae: Genetic organization and properties of the gene products.
<3>Mol. Microbiol.
<4>41
<5>1199-1210
<6>2001
<7>The hsd locus (host specificity of DNA) was identified in the Neisseria gonorrhoeae genome.
The DNA fragment encoding this locus produced an active restriction and modification (R/M)
system when cloned into Escherichia coli. This R/M system was designated NgoAV. The cloned
genomic fragment (7800 bp) has the potential to encode seven open reading frames (ORFs).
Several of these ORFs had significant homology with other proteins found in the databases:
ORF1, the hsdM, a methylase subunit (HsdM); ORF2, a homologue of dinD; ORF3, a homologue of
hsdS; ORF4, a homologue of hsdS; and ORF5, an endonuclease subunit hsdR. The endonuclease and
methylase subunits possessed strongest protein sequence homology to the EcoR124II R/M system,
indicating that NgoAV belongs to the type IC R/M family. Deletion analysis showed that only
ORF3 imparted the sequence specificity of the RM.NgoAV system, which recognizes an interrupted
palindrome sequence (GCAN8TGC). The genetic structure of ORF3 (208 amino acids) is almost
identical to the structure of the 5' truncated hsdS genes of EcoDXXI or EcoR124II R/M systems
obtained by in vitro manipulation. Genomic sequence analysis allowed us to identify hsd loci
with a very high homology to RM.NgoAV in two strains of Neisseria meningitidis. However,
significant differences in the organization and structure of the hsdS genes in both these
systems suggests that, if functional, they would possess recognition sites that differ from
the gonococcus and from themselves.

<>

<1>Piekarowicz, A., Radlinska, M.
<2>Sensitivity of the restriction endonucleases HaeIII, BsrI, EaeI and CfrI to cytosine N4-methylation.
<3>Acta Microbiol. Pol.
<4>47
<5>405-407
<6>1998
<7>HaeIII, BsrI and NgoII are isochizomers that recognize the sequence GGCC while EaeI and CfrI
recognize the overlapping sequence YGGCCR. It has previously been shown that all these enzymes
are inhibited by cytosine C5-methylation within the recognition sequence. The methylation
sensitivities of these enzymes to cytosine N4-methylation have not been previously reported.
In this paper we present data demonstrating that all these enzymes, except NgoII, are
inhibited by cytosine N4-methylation of the second 5' cytosine residue within the recognition
sequence.

<>

<1>Piekarowicz, A., Radlinska, M., Wiernicka-Gnas, M.
<2>DNA methyltransferases of Neisseria gonorrhoeae.
<3>Bull. Acad. Pol. Sci. [Biol]
<4>44
<5>205-210
<6>1996
<7>An individual strain of Neisseria gonorrhoeae may produce up to 16 different DNA
methyltransferases.  We have used a novel cloning system that is able to detect MTase clones
in the absence of direct selection to identify different MTase clones.  The characterization
of six of these clones showed that MTase genes are linked to restriction endonuclease systems
but none of these six R-M systems are genetically linked on the chromosome.  The initial
characterization of four other clones indicates that none of the encoded MTase genes are
linked to the restriction endonuclease systems.  On the other hand, several of these MTases
show genetical linkage on the chromosome.  Four of these MTase clones have been characterized
by DNA sequence analysis, and the open reading frames encoding each of these MTases have been
identified.  These MTases belong either to the 5mC or N4mC group of MTases.  Some of the 5mC
MTases show the presence of typical MTase conserved motifs.  However, some other cloned 5mC
MTase show the lack of these motifs.

<>

<1>Piekarowicz, A., Skowronek, K.
<2>Identification of a new restriction endonuclease R.BcrAI from Bacillus cremoris.
<3>Acta Microbiol. Pol.
<4>44
<5>315-316
<6>1995
<7>Site specific restriction endonuclease R.BcrAI has been purified from Bacillus cremoris.  The
enzyme recognizes the sequence 5' CTCTTC 3'.

<>

<1>Piekarowicz, A., Stasiak, A., Stanczak, J.
<2>Specific restriction endonucleases from Haemophilus influenzae JC9.
<3>Acta Microbiol. Pol.
<4>29
<5>151-156
<6>1980
<7>Two types of restriction endonucleases have been isolated from Haemophilus
influenzae.  The presence of type III restriction enzymes is in vivo correlated
with the activity against Haemophilus phages HP1, S2 and N3 (Piekarowicz,
Brzezinski and Kauc, 1975, Kauc and Piekarowicz, 1978).

<>

<1>Piekarowicz, A., Stein, D.C.
<2>Purification and characterization of a new DNA methyltransferase from Neisseria gonorrhoeae.
<3>Gene
<4>157
<5>101-102
<6>1995
<7>A new DNA methyltransferase, M.NgoBVII, was isolated from Neisseria gonorrhoeae strain WR302.
M.NgoVII recognizes the sequence 5'-GCNGC-3'.
[ The enzyme called NgoBVII in this abstract has been renamed NgoBXII, Jan/1998. ]

<>

<1>Piekarowicz, A., Weglenska, A.
<2>Improvement of the strain for the rapid identification of genes encoding restriction and modification enzymes.
<3>Acta Microbiol. Pol.
<4>43
<5>229-231
<6>1994
<7>The E. coli AP1-200-9 strain used for rapid identification of genes encoding restriction and
modification enzymes carries a temperature sensitive lacZ gene fused to the damage-inducible
dinD locus. A derivative of this strain was constructed that has a wild-type form of this
locus which allows for a more efficient identification of recombinant plasmids encoding
restriction and modification enzymes.

<>

<1>Piekarowicz, A., Yuan, R., Stein, D.C.
<2>Identification of a new restriction endonuclease, R.NgoBI, from Neisseria gonorrhoeae.
<3>Nucleic Acids Res.
<4>16
<5>9868
<6>1988
<7>As a species, Neisseria gonorrhoeae produces five restriction endonucleases, and several other
DNA methyltransferases.  We have purified a methyltransferase that recognizes the sequence 5'
TCACC 3' and report here the purification from N. gonorrhoeae WR302 of a restriction
endonuclease, R.NgoBI, that also recognizes this sequence.  Purification scheme:  The
purification scheme employed was as previously described (2) except the (NH4)2SO4 precipitate
was dissolved in buffer A (20 mM KPO4, 1 mM EDTA, 10% glycerol, 10 mM 2-mercaptoethanol, pH
7.5) before being purified by chromatography through a 2x20 cm phosphocellulose column.  The
enzyme activity eluted at 0.15 M NaCl. Active fractions were further purified through an Accel
QMA column and active fractions eluted at 0.1 M NaCl.  The recognition sequence for R.NgoBI
was determined by digesting lambda DNA with it and comparing the banding pattern obtained with
computer generated patterns obtained with all known enzymes.  The data indicated that this
enzyme cleaved lambda DNA at the same sequence as HphI.  Figure 1 is a comparison of the
fragments obtained after digesting pUC8 with NgoBI and HphI.  The restriction enzyme was most
active in a buffer containing 25 mM Tris-HCl, 25 mM KCl, 10 mM MgCl2, 2 mM 2-mercaptoethanol,
pH 7.8.
[ The enzyme called NgoBI in this abstract has been renamed NgoBVIII, Jan/1998. ]

<>

<1>Piekarowicz, A., Yuan, R., Stein, D.C.
<2>Purification and characterization of DNA methyltransferases from Neisseria gonorrhoeae.
<3>Nucleic Acids Res.
<4>16
<5>5957-5972
<6>1988
<7>Three DNA methyltransferases, M.NgoAI, and M.NgoBI and M.NgoBII, free of any nuclease
activities were isolated from Neisseria gonorrhoeae strains WR220 and MUG116 respectively.
M.NgoAI recognizes the sequence 5' GGCC 3' and methylates the first 5' cytosine on both
strands.  M.NgoBI and M.NgoBII recognize 5' TCACC 3' and 5' GTANNNNNCTC 3' respectively.
M.NgoBII 5' GTANNNNNmCTC 3'.
[ The enzyme called NgoAI in this abstract has been renamed NgoGII, Jan/1998. ]
[ The enzyme called NgoBI in this abstract has been renamed NgoBVIII, Jan/1998. ]
[ The enzyme called NgoBII in this abstract has been renamed NgoBIX, Jan/1998. ]

<>

<1>Piekarowicz, A., Yuan, R., Stein, D.C.
<2>Construction of a temperature-sensitive mutation for the direct identification of plasmids encoding DNA methyltransferases.
<3>Gene
<4>74
<5>233-235
<6>1988
<7>Meeting Abstract

<>

<1>Piekarowicz, A., Yuan, R., Stein, D.C.
<2>Neisseria gonorrhoeae M.NgoAI DNA methyltransferase:  physical and catalytic properties of the homogeneous enzyme.
<3>Gene
<4>74
<5>93-97
<6>1988
<7>A DNA methyltransferase, M.NgoAI, was purified to homogeneity from Neisseria
gonorrhoeae strain WR220 by successive column chromatography.  Its Mr is 25000,
as determined by both gel filtration and denaturing polyacrylamide gel
electrophoresis.  Maximal enzymatic activity was obtained in 50 mM Tris.HCl (pH
7.4), 10 mM EDTA, with incubation at 37C.  An apparent Km value for
S-adenosylmethionine and 5'-GGCC sites was determined to be 1.25 microM and
89.6 nM, respectively.

<>

<1>Piekarowicz, A., Yuan, R., Stein, D.C.
<2>Cleavage of DNA by HaeII is inhibited by the presence of 5-methylcytosine at the second cytosine within the recognition sequence.
<3>Nucleic Acids Res.
<4>17
<5>10132
<6>1989
<7>None

<>

<1>Piekarowicz, A., Yuan, R., Stein, D.C.
<2>A new method for the rapid identification of genes encoding restriction and modification enzymes.
<3>Nucleic Acids Res.
<4>19
<5>1831-1835
<6>1991
<7>We have constructed derivatives of Escherichia coli that can be used for the
rapid identification of recombinant plasmids encoding DNA restriction enzymes
and methyltransferases.  The induction of the DNA-damage inducible SOS response
by the Mcr and Mrr systems, in the presence of methylated DNA, is used to
select plasmids encoding DNA methyltransferases.  The strains of E. coli that
we have constructed are temperature-sensitive for the Mcr and Mrr systems and
have been further modified to include a lacZ gene fused to the damage-inducible
dinD locus of E. coli.  The detection of recombinant plasmids encoding DNA
methyltransferases and restriction enzymes is a simple, one step procedure that
is based on the induction at the restrictive temperature of the lacZ gene.
Transformants encoding DNA methyltransferase genes are detected on LB agar
plates supplemented with X-gal as blue colonies.  Using this method, we have
cloned a variety of DNA methyltransferase genes from diverse species such as
Neisseria, Haemophilus, Treponema, Pseudomonas, Xanthomonas and
Saccharopolyspora.

<>

<1>Piekarowicz, A., Yuan, R., Stein, D.C.
<2>Isolation of temperature-sensitive McrA and McrB mutations and complementation analysis of the McrBC region of Escherichia coli K-12.
<3>J. Bacteriol.
<4>173
<5>150-155
<6>1991
<7>We isolated temperature-sensitive mcrA and mcrBC mutants of Escherichia coli.
At 42C, they were unable to restrict the T-even bacteriophages T6gt and Tegt or
plasmids encoding cloned DNA methylase genes whose specificities confer
sensitivity to the McrA and McrBC nucleases.  Complementation analysis of the
McrBC region (mcrB251) with the complete cloned McrBC system or a derivative
with mcrB alone indicated that the mutation shows an absolute defect for the
restriction of DNA containing hydroxymethylcytosine and a thermosensitive
defect for the restriction of DNA containing methylcytosine.  The properties of
the McrA temperature-sensitive mutants suggest that some of these mutations can
also influence the restriction of DNA containing hydroxymethylcytosine or
methylcytosine residues.

<>

<1>Pieper, U., Brinkmann, T., Kruger, T., Noyer-Weidner, M., Pingoud, A.
<2>Characterization of the interaction between the restriction endonuclease McrBC from E. coli and its cofactor GTP.
<3>J. Mol. Biol.
<4>272
<5>190-199
<6>1997
<7>McrBC, a GTP-dependent restriction enzyme from E. coli K-12, cleaves DNA containing methylated
cytosine residues 40 to 80 residues apart and 3'-adjacent to a purine residue
(pumCN40-80PumC).  The presence of the three consensus sequences characteristic for guanine
nucleotide binding proteins in one of the two subunits of McrBC suggests that this subunit is
responsible for GTP binding and hydrolysis.  We show here that (i) McrB binds GTP with an
affinity of 106 M^-1 and that GTP binding stabilizes McrB against thermal denaturation.  (ii)
McrB binds GDP about 50-fold and ATP at least three orders of magnitude more weakly than GTP.
(iii) McrB hydrolyzes GTP in the presence of Mg2+ with a steady-state rate of approximately
0.5 min^-1.  (iv) McrC stimulates GTP hydrolysis 30-fold, but substrate DNA has no detectable
effect on the GTPase activity of McrB, neither by itself nor in the presence of McrC.  (v)
Substitution of N339 and N376 with alanine allowed us to identify NTAD (339 to 342) rather
than NKKA (376 to 379) as the equivalent of the third consensus sequence motif characteristic
for guanine nucleotide binding proteins, NKXD.

<>

<1>Pieper, U., Groll, D.H., Wunsch, S., Gast, F.U., Speck, C., Mucke, N., Pingoud, A.
<2>The GTP-dependent restriction enzyme McrBC from Escherichia coli forms high-molecular mass complexes with DNA and produces a cleavage pattern with a characteristic 10-base pair repeat.
<3>Biochemistry
<4>41
<5>5245-5254
<6>2002
<7>The GTP-dependent restriction enzyme McrBC consists of two polypeptides: one (McrB) that is
responsible for GTP binding and hydrolysis as well as DNA binding and another (McrC) that is
responsible for DNA cleavage. It recognizes two methylated or hemimethylated RC sites (R(m)C)
at a distance of approximately 30 to more than 2000 base pairs and cleaves the DNA close to
one of the two R(m)C sites. This process is strictly coupled to GTP hydrolysis and involves
the formation of high-molecular mass complexes. We show here using footprinting techniques,
surface plasmon resonance, and scanning force microscopy experiments that in the absence of
McrC, McrB binds to a single R(m)C site. If a second R(m)C site is present on the DNA, it is
occupied independently by McrB. Whereas the DNA-binding domain of McrB forms 1:1 complexes
with each R(m)C site and shows a clear footprint on both R(m)C sites, full-length McrB forms
complexes with a stoichiometry of at least 4:1 at each R(m)C site, resulting in a slightly
more extended footprint. In the presence of McrC, McrB forms high-molecular mass complexes of
unknown stoichiometry, which are considerably larger than the complexes formed with McrB
alone. In these complexes and when GTP is present, the DNA is cleaved next to one of the R(m)C
sites at distances differing by one to five helical turns, suggesting that in the McrBC-DNA
complex only a few topologically well-defined phosphodiester bonds of the DNA are accessible
for the nucleolytic center of McrC.

<>

<1>Pieper, U., Pingoud, A.
<2>A mutational analysis of the PD...D/EXK motif suggests that McrC harbors the catalytic center for DNA cleavage by the GTP-dependent restriction enzyme McrBC from Escherichia coli.
<3>Biochemistry
<4>41
<5>5236-5244
<6>2002
<7>McrBC is a unique restriction enzyme which binds specifically to the bipartite recognition
sequence RmCNa~30-2000RmC and in the
presence of GTP translocates the DNA and cleaves both strands at
multiple positions within the two RmC "half-sites". It is known that
McrBC is composed of two subunits: McrB which binds and hydrolyzes GTP
and specifically interacts with DNA and McrC whose function is not
clear but which has been suspected to harbor the catalytic center for
DNA cleavage. A multiple-sequence alignment of the amino acid sequence
of Escherichia coli McrC and of six presumably homologous open reading
frames from various bacterial species shows that a sequence motif found
in many restriction enzymes, but also in other nucleases, the
PD....D/EXK motif, is conserved among these sequences. A mutational
analysis, in which the carboxylates (aspartic acid in McrC) of this
motif were substituted with alanine or asparagine and lysine was
substituted with alanine or arginine, strongly suggests that Asp244,
Asp257, and Lys259 represent the catalytic center of E. coli McrC.
Whereas the variants D244A (or -N), D257A (or -N), and K259A are
inactive in DNA cleavage (K259R has residual DNA cleavage activity),
they interact with McrB like wild-type McrC, as can be deduced from the
finding that they stimulate the McrB-catalyzed GTP hydrolysis to the
same extent as wild-type McrC. Thus, whereas McrC variants defective in
DNA cleavage can stimulate the GTPase activity of McrB, the DNase
activity of McrC is not supported by McrB variants defective in GTP
hydrolysis.

<>

<1>Pieper, U., Schweitzer, T., Groll, D.H., Gast, F.-U., Pingoud, A.
<2>The GTP-binding domain of McrB: More than just a variation on common theme?
<3>J. Mol. Biol.
<4>292
<5>547-556
<6>1999
<7>The methylation-dependent restriction endonuclease McrBC from Escherichia coli K12 cleaves DNA
containing two R(m)C dinucleotides separated by about 40 to 2000 base-pairs. McrBC is unique
in that cleavage is totally dependent on GTP hydrolysis. McrB is the GTP binding and
hydrolyzing subunit, whereas MrC stimulates its GTP hydrolysis. The C-terminal part of McrB
contains the sequences characteristic for GTP-binding proteins, consisting of the GxxxxGK(S/T)
motif (position 201-208), followed by the DxxG motif (position 300-303). The third motif
(NKxD) is present only in a non-canonical form (NTAD 333-336). Here we report a mutational
analysis of the putative GTP-binding domain of McrB. Amino acid substitutions were initially
performed in the three proposed GTP-binding motifs. Whereas substitutions in motif 1 (P203V)
and 2 (D300N) show the expected, albeit modest effects, mutation in the motif 3 is at variance
with the expectations. Unlike the corresponding EF-Tu and ras -p21 variants, the D336N
mutation in McrB does not change the nucleotide specificity from GTP to XTP, but results in a
lack of GTPase stimulation by McrC. The finding that McrB is not a typical G protein motivated
us to perform a search for similar sequences in DNA databases. Eight microbial sequences were
found, mainly from unfinished sequencing projects, with highly conserved sequence blocks
within a presumptive GTP-binding domain. From the five sequences showing the highest homology,
17 invariant charged or polar residues outside the classical three GTP-binding motifs were
identified and subsequently exchanged to alanine. Several mutations specifically affect GTP
affinity and/or GTPase activity. Our data allow us to conclude that McrB is not a typical
member of the superfamily of GTP-binding proteins, but defines a new subfamily within the
superfamily of GTP-binding proteins, together with similar prokaryotic proteins of as yet
unidentified function.

<>

<1>Pieper, U., Schweitzer, T., Groll, D.H., Pingoud, A.
<2>Defining the location and function of domains of McrB by deletion mutagenesis.
<3>Biol. Chem.
<4>380
<5>1225-1230
<6>1999
<7>The GTP-dependent restriction endonuclease McrBC of E. coli K12, which recognizes
cytosine-methylated DNA, consists of two protein subunits, McrB and McrC. We have investigated
the structural assignment and interdependence of the McrB subunit functions, namely (i)
specific DNA recognition and (ii) GTP binding and hydrolysis. Extending earlier work, we have
produced McrB variants comprising N- and C-terminal fragments. The variants McrB1-162 and
McrB1-170 are still capable of specific DNA binding. McrB169-465 shows GTP binding and
hydrolysis characteristics indistinguishable from full-length McrB as well as wild-type like
interaction with McrC. Thus, DNA and GTP binding are spatially separated on the McrB molecule,
and the respective domains function quite independently.

<>

<1>Pieretti, I., Royer, M., Barbe, V., Carrere, S., Koebnik, R., Cociancich, S., Couloux, A., Darrasse, A., Gouzy, J., Jacques, M.A., Lauber, E., Manceau, C., Mangenot, S., Poussier, S., Segurens, B., Szurek, B., Verdier, V., Arlat, M., Rott, P.
<2>The complete genome sequence of Xanthomonas albilineans provides new insights into the reductive genome evolution of the xylem-limited Xanthomonadaceae.
<3>BMC Genomics
<4>10
<5>616
<6>2009
<7>BACKGROUND: The Xanthomonadaceae family contains two xylem-limited plant
pathogenic bacterial species, Xanthomonas albilineans and Xylella
fastidiosa. X. fastidiosa was the first completely sequenced plant
pathogen. It is insect-vectored, has a reduced genome and does not possess
hrp genes which encode a Type III secretion system found in most plant
pathogenic bacteria. X. fastidiosa was excluded from the Xanthomonas group
based on phylogenetic analyses with rRNA sequences. RESULTS: The complete
genome of X. albilineans was sequenced and annotated. X. albilineans,
which is not known to be insect-vectored, also has a reduced genome and
does not possess hrp genes. Phylogenetic analysis using X. albilineans
genomic sequences showed that X. fastidiosa belongs to the Xanthomonas
group. Order of divergence of the Xanthomonadaceae revealed that X.
albilineans and X. fastidiosa experienced a convergent reductive genome
evolution during their descent from the progenitor of the Xanthomonas
genus. Reductive genome evolutions of the two xylem-limited
Xanthomonadaceae were compared in light of their genome characteristics
and those of obligate animal symbionts and pathogens. CONCLUSION: The two
xylem-limited Xanthomonadaceae, during their descent from a common
ancestral parent, experienced a convergent reductive genome evolution.
Adaptation to the nutrient-poor xylem elements and to the cloistered
environmental niche of xylem vessels probably favoured this convergent
evolution. However, genome characteristics of X. albilineans differ from
those of X. fastidiosa and obligate animal symbionts and pathogens,
indicating that a distinctive process was responsible for the reductive
genome evolution in this pathogen. The possible role in genome reduction
of the unique toxin albicidin, produced by X. albilineans, is discussed.

<>

<1>Piet, J.R., Huis, I.V.R.A., van Schaik, B.D., van Kampen, A.H., Baas, F., van de Beek, D., Pannekoek, Y., van der Ende, A.
<2>Genome Sequence of Neisseria meningitidis serogroup B strain H44/76.
<3>J. Bacteriol.
<4>193
<5>2371-2372
<6>2011
<7>Neisseria meningitidis is an obligate human pathogen. While it is a frequent commensal of the
upper respiratory tract, in some individuals the bacterium spreads to the bloodstream causing
meningitis and/or sepsis, serious conditions with high morbidity and mortality. Here we report
the availability of the genome sequence of the widely used serogroup B laboratory strain
H44/76.

<>

<1>Pietila, M.K., Laurinmaki, P., Russell, D.A., Ko, C.C., Jacobs-Sera, D., Hendrix, R.W., Bamford, D.H., Butcher, S.J.
<2>Structure of the archaeal head-tailed virus HSTV-1 completes the HK97 fold story.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>110
<5>10604-10609
<6>2013
<7>It has been proposed that viruses can be divided into a small number of
structure-based viral lineages. One of these lineages is exemplified by bacterial
virus Hong Kong 97 (HK97), which represents the head-tailed dsDNA bacteriophages.
Seemingly similar viruses also infect archaea. Here we demonstrate using genomic
analysis, electron cryomicroscopy, and image reconstruction that the major coat
protein fold of newly isolated archaeal Haloarcula sinaiiensis tailed virus 1 has
the canonical coat protein fold of HK97. Although it has been anticipated
previously, this is physical evidence that bacterial and archaeal head-tailed
viruses share a common architectural principle. The HK97-like fold has previously
been recognized also in herpesviruses, and this study expands the HK97-like
lineage to viruses from all three domains of life. This is only the second
established lineage to include archaeal, bacterial, and eukaryotic viruses. Thus,
our findings support the hypothesis that the last common universal ancestor of
cellular organisms was infected by a number of different viruses.

<>

<1>Pietrokovski, S.
<2>Conserved sequence features of inteins (protein introns) and their use in identifying new inteins and related proteins.
<3>Protein Sci.
<4>3
<5>2340-2350
<6>1994
<7>Inteins (protein introns) are internal portions of protein sequences that are
posttranslationally excised while the flanking regions are spliced together, making an
additional protein product.  Inteins have been found in a number of homologous genes in
yeast, mycobacteria, and extreme thermophile archaebacteria.  The inteins are probably
multifunctional, autocatalyzing their own splicing, and some were also shown to be DNA
endonucleases.  The splice junction regions and two regions similar to homing
endonucleases were thought to be the only common sequence features of inteins.  This
work analyzed all published intein sequences with recently developed methods for detecting
weak, conserved sequence features.  The methods complemented each other in the
identification and assessment of several patterns characterizing the intein sequences.  New
intein conserved features are discovered and the known ones are quantitatively described
and localized.  The general sequence description of all the known inteins is derived from
the motifs and their relative positions.  The intein sequence description is used to search
the
sequence databases for intein-like proteins.  A sequence region in a mycobacterial open
reading frame possessing all of the intein motifs and absent from sequences homologous to
both of its flanking sequences is identified as an intein.  A newly discovered putative intein
in red algae chloroplasts is found not to contain the endonuclease motifs present in all other
inteins.  The yeast HO endonuclease is found to have an overall intein-like structure and a
few viral polyprotein cleavage sites are found to be significantly similar to the inteins
amino-end splice junction motif.  The intein features described may serve for detection of
intein sequences.

<>

<1>Pietrokovski, S.
<2>A new intein in cyanobacteria and its significance for the spread of inteins.
<3>Trends Genet.
<4>12
<5>287-288
<6>1996
<7>Inteins are protein 'introns' encoded inside the polypeptide sequence of
other proteins.  The inteins splice out post-translationally by a proteolytic cleavage and
ligation process.  Inteins appear to autocatalyze their own excision and some are site-
specific endonucleases.  Inteins are mobile genetic elements and at least one can home, that
is, insert a copy of its DNA into its integration site in an intein-less allele.  Fifteen
inteins
have been found in various organisms, including mycobacteria, thermophilic
archaebacteria, yeast and chloroplast of red alga.  Inteins are not very similar to one
another, but homologous sites in archaebacterial DNA polymerases and in mycobacterial
gyrase-A proteins contain homologous inteins.  However, the mycobacterial RecA proteins
and DNA polymerases also contain different inteins in different integration sites.

<>

<1>Pietrokovski, S.
<2>Modular organization of inteins and C-terminal autocatalytic domains.
<3>Protein Sci.
<4>7
<5>64-71
<6>1998
<7>Analysis of the conserved sequence features of inteins (protein "introns") reveals that they
are composed of three distinct modular domains.  The N-terminal and C-terminal domains are
predicted to perform different parts of the autocatalytic protein splicing reaction.  An
optional endonuclease domain is shown to correspond to different types of homing endonucleases
in different inteins.  The N domain contains motifs predicted to catalyze the first steps of
protein splicing, leading to the cleavage of the intein N terminus from its protein host.
Intein N domain motifs are also found in C-terminal autocatalytic domains present in hedgehog
and other protein families.  Specific residues in the N domain of intein and CADs are proposed
to form a charge relay system involved in cleaving their N-termini.  The intein C domain is
apparently unique to inteins and contains motifs that catalyze the final protein splicing
steps: ligation of the intein flanks and cleavage of its C terminus to release the free intein
and spliced host protein.  All intein EN domains known thus far have dodecapeptide (DOD,
LAGLI-DADG) type homing endonuclease motifs.  This work identifies an EN domain with an HNH
homing-endonuclease motif and two new small inteins with no EN domains.  One of these small
inteins might be inactive or a "pseudo intein".  The results suggest a modular architecture
for inteins, clarify their origin and relationship to other protein families, and extend
recent experimental findings on the functional roles of intein N, C and EN motifs.

<>

<1>Pietrokovski, S.
<2>Intein spread and extinction in evolution.
<3>Trends Genet.
<4>17
<5>465-472
<6>2001
<7>Inteins are selfish DNA elements found within coding regions. They are translated with their
host protein, but then catalyze their own
excision and the formation of a peptide bond between their flanking
protein regions. Understanding what drives and selects inteins is
relevant for assessing whether they have unidentified biological
functions and whether they can invade and become established in new
genes and organisms. Inteins are suggested to have been present and
more common in the progenitors of eukaryotes and prokaryotes. In these
cells, inteins had some beneficial function or had evolved from an
unknown beneficial protein. Since then, this putative benefit has been
lost and inteins are gradually becoming extinct. The proteins in which
inteins are currently found are proposed to be proteins vital for the
survival of the organism, where intein removal is most difficult.

<>

<1>Pightling, A.W., Lin, M., Pagotto, F.
<2>Draft Genome Sequence of Listeria monocytogenes Strain LI0521 (syn. HPB7171), Isolated in 1983 during an Outbreak in Massachusetts Caused by Contaminated  Cheese.
<3>Genome Announcements
<4>2
<5>e00729-14
<6>2014
<7>Listeria monocytogenes, a pathogenic food-borne bacterium, is the causative agent of both
sporadic and outbreak cases of human listeriosis. Here, we present the
genome sequence of L. monocytogenes reference strain LI0521, isolated during an
outbreak involving contaminated cheese, which has been used as the model during
several proteomic studies.

<>

<1>Pightling, A.W., Pagotto, F.
<2>Draft Genome Sequence of Cronobacter sakazakii Clonal Complex 45 Strain HPB5174,  Isolated from a Powdered Infant Formula Facility in Ireland.
<3>Genome Announcements
<4>2
<5>e00778-14
<6>2014
<7>Cronobacter sakazakii is a food-borne pathogenic bacterium that may cause severe  illness in
neonates and the elderly. We present the genome sequence of a rare
strain (ST40, CC45), commonly found in multiple food processing facilities and in
powdered infant formula and only indicted in a single clinical case.

<>

<1>Pignot, M., Pljevaljcic, G., Weinhold, E.
<2>Efficient synthesis of S-adenosyl-L-homocysteine natural product analogues and their use to elucidate the structural determinant for cofactor binding of the DNA methyltransferase M.HhaI.
<3>Eur. J. Org. Chem.
<4>2000
<5>549-555
<6>2000
<7>5'-Acetylthio-5'-deoxy-2',3'-O-isopropylideneadenosine was directly prepared from
commercially available 2',3'-O-isopropylideneadenosine and thioacetic acid under Mitsunobu
conditions in almost quantitative yield.  In situ cleavage of the acetylthio function of
5'-Acetylthio-5'-deoxy-2',3'-O-isopropylideneadenosine followed by coupling with different
alkyl bromides proceeded with high yields.  Deprotection of the obtained 5'-thionucleosides
yielded the S-adenosyl-L-homocysteine analogues decarboxylated AdHcy, deaminated AdoHcy and
5'-[3-(cyano)propylthio]-5'-deoxyadenosine in good overall yields.  Direct deprotection of
the thionucleoside 5'-Acetylthio-5'-deoxy-2',3'-O-isopropylideneadenosine delivered
5'-thio-5'-deoxyadenosine in excellent yield.  In addition, binding constants of these
AdoHcy analogues and the DNA methyltransferase M.HhaI were determined in a fluorescence assay.

<>

<1>Pignot, M., Siethoff, C., Linscheid, M., Weinhold, E.
<2>Coupling of a nucleoside with DNA by a methyltransferase.
<3>Angew. Chem. Int. Ed. Engl.
<4>37
<5>2888-2891
<6>1998
<7>S-Adenosyl-L-methionine-dependent methyltransferases catalyze the transfer of the activated
methyl group from the cofactor S-adenosyl-L-methionine to sulfur, nitrogen, oxygen, and carbon
acceptors (Scheme 1) of small molecules, phospholipids, proteins, RNA and DNA with high
specificity.  The transfer of larger chemical entities in a Mtase-catalyzed reaction has not
been reported and thus represents an interesting challenge for bioorganic chemists.  In
principal, covalent linking of the activated methyl group with the gamma-C atom of 1 would
yield a three-membered thiiranium compound, which could lead to a coupling of the whole
cofactor to the target substrate.  Since thiiranium compounds are known to be unstable in
nucleophilic solvents, we concentrated on the more stable aziridine analogues, which can be
activated as alkylating reagents upon protonation of their ring nitrogen atom.
N-adenosylaziridine was synthesized by nucleophilic substitution of the tosylate group of
5'-deoxy-5'-tosyladenosine (tosyl=Ts=toluene-4-sulfonyl) with aziridine (Scheme 2).

<>

<1>Pikin, S.A.
<2>Physical aspects of the structure and function of helicases as rotary molecular motors.
<3>Crystallogr. Rep.
<4>54
<5>929-936
<6>2009
<7>Helicases were shown to have common physical properties with rotary molecular motors, such as
F (0) F (1)-ATP synthase and type I
restriction-modification (RM) enzymes. The necessary conditions for
action of molecular motors are chirality, the presence of the C (2) (or
lower) symmetry axis within rather large atomic groups, and
polarization properties. The estimates were made for the material
parameters of helicases, which translocate DNA due to moving chiral
kinks without DNA cleavage and are characterized by higher viscosity,
low mobility, and smaller chiral kinetic coefficients than type II RM
enzymes. This paper discusses the efficiency of helicases with opposite
polarities that drive DNA translocation in opposite directions.

<>

<1>Pikin, S.A.
<2>On the distinction of the mechanisms of DNA cleavage by restriction enzymes - The I-, II-, and III-type molecular motors.
<3>Crystallogr. Rep.
<4>53
<5>858-867
<6>2008
<7>A comparative physical description is given for the functioning of various restriction enzymes
and for their processes of DNA cleavage.
The previously proposed model system of kinetic equations is applied to
the I- and III-type enzymes, which use ATP molecules as an energy
source, while the II-type enzymes work thanks to catalytic reactions
with participation of an electric field. All the enzymes achieved
bending and twisting DNA, providing for either the linear motion of the
II-type enzyme along the DNA chain or the DNA translocation by the
I-and III-type enzymes due to moving chiral kinks. A comparative
estimation of the considered linear and angular velocities is
performed. The role of stalling forces for enzyme-DNA complexes, which
induce the observed cutting of the DNA either inside the enzyme (II) or
in some "weak" places outside enzymes I and III, which results in the
supercoiling of the DNA, is shown. The role of ionic screening for the
described processes is discussed.

<>

<1>Pikin, S.A.
<2>On the DNA cleavage by restriction enzymes - molecular motors with polarization properties.
<3>Mol. Cryst. Liq. Cryst.
<4>508
<5>403-413
<6>2009
<7>In the paper, on the general physical basis, the attempt was done to explain the operation of
restriction enzymes of different types.  The physical model lies in the DNA deformation in the
protein zone which is caused by catalytic processes taking place here.  It is shown that some
phenomena are similar to liquid-crystalline effects.  The DNA molecule either forms locally a
chiral kink moving together with a protein (the II type enzymes) or realizes the translocation
through protein (the I and III type enzymes).  The velocity of linear motion of the enzymes
was estimated on the basis of proper kinetic equations which include the action of a
longitudinal stalling force, the role of this force in DNA cleavage being different for
different enzymes.  The supercoiling of DNA during tis translocation is discussed.

<>

<1>Piknova, M., Filova, M., Javorsky, P., Pristas, P.
<2>Different restriction and modification phenotypes in ruminal lactate-utilizing bacteria.
<3>FEMS Microbiol. Lett.
<4>236
<5>91-95
<6>2004
<7>Analysis of restriction and modification activities in lactate-utilizing bacteria belonging to
the Megasphaera elsdenii and Mitsuokella multiacida species revealed the presence of
GATC-specific, MboI isospecific, restriction-modification (R-M) systems in all strains tested.
While restriction endonucleases isolated from M. elsdenii strains were found to be sensitive
to Dam methylation, enzymes from M. multiacida cleaved DNA irrespective of Dam methylation.
The comparison of type 11 R-M systems specificities in three closely related lactate-utilizing
ruminal bacterial species indicated complete lack of restriction and/or modification enzymes
previously characterized from Selenomonas ruminantium in tested M. elsdenii and M. multiacida
strains. R-M systems are believed to represent the main defense tool against phage infection.
Based on the results of our experiments it could be assumed that M. elsdenii and M. multiacida
use the different strategy for bacteriophage protection compared to S. ruminantium.

<>

<1>Piknova, M., Filova, M., Javorsky, P., Pristas, P.
<2>A Unique Pair of GATC Specific DNA Methyltransferases in Mitsuokella multiacida.
<3>Mol. Biol. Rep.
<4>32
<5>281-284
<6>2005
<7>Two GATC specific methylases together with Sau3AI isoschizomeric restriction endonuclease were
partially characterized in Mitsuokella
multiacida 46/5. This is the first report on the presence of solitary Dam
methyltransferase alongside GATC specific restriction-modification system
resulting in the unusual two-fold methylation of the GATC motifs.

<>

<1>Piknova, M., Javorsky, P., Pristas, P.
<2>Multiple restriction-modification systems are present in rumen treponemes.
<3>FEMS Microbiol. Lett.
<4>251
<5>99
<6>2005
<7>Type II restriction endonucleases were purified by heparin-sepharose followed by ion
chromatography from Treponema strains. The results
indicate that in addition to frequently cutting GATC-specific
restriction enzymes, the tested strains also possess rarely cutting
endonucleases. The purified restriction endonucleases represent four
different sequence specificities, comprising isoschizomers of Drdl,
AflII, Tth111II and NdeI. The data presented show that three rumen
Treponema strains possess altogether seven type II
restriction-modification systems. Thus, individual Treponema strains
may be considered an interesting source of multiple type II restriction
enzymes.

<>

<1>Piknova, M., Pristas, P., Javorsky, P.
<2>GATC-specific restriction-modification systems in ruminal bacteria.
<3>Folia Microbiol. (Praha)
<4>49
<5>191-193
<6>2004
<7>The GATC-specific restriction and modification activities were analyzed in 11 major bacterial
representatives of ruminal microflora.
Modification phenotype was observed in 13 out of 40 ruminal strains.
MboI isoschizomeric restriction endonucleases were detected in 10
bacterial strains tested; three strains lacked any detectable
corresponding endonuclease activity. The only examined strain of
Mitsuokella multiacida was found to possess a different type of
endonuclease activity. This is the first report on restriction activity
in ruminal treponemes M. multiacida and Megasphaera elsdenii.

<>

<1>Piknova, M., Pristas, P., Javorsky, P.
<2>Some evolutionary aspects of biology of the type II restriction-modification systems.
<3>Biol. Listy
<4>69
<5>15-28
<6>2004
<7>Restriction-modification (R-M) systems occur exclusively among prokaryotic organisms, mainly
bacteria, and they represent the main protection mechanism against bacteriophage infections.
R-M systems comprise two enzymatic activities: a restriction endonuclease activity that
specifically cleaves DNA; and a corresponding methyltransferase activity that specifically
methylates the DNA, thereby protecting the genome of the host bacterium from cleavage by a
partner's restriction enzyme.  There are three main groups of R-M systems called Types I, II
and III and recently a new Type IV has been added to accommodate a class of methyl-dependent
restriction enzymes.  R-M systems are widely distributed among bacteria and more than 3500
restriction enzymes and 600 methyltransferases have been identified to date.  The most
abundant are type II R-M systems, which form a very large family of enzymes of similar
function.  Considering the dissimilarities in amino acid sequences and, paradoxically,
structural resemblances of restriction enzymes, it is very hard to decide whether restriction
endonucleases are the outcome of divergent evolution from a very distant ancestor or whether
they evolved independently.  While generally accepted that R-M systems act as barrier against
genetic exchanges, there is increasing evidence for their behavior as mobile genetic elements.

<>

<1>Piknova, M., Pristas, P., Javorsky, P., Kasperowic, A., Michalowski, T.
<2>GATC-specific restriction and modification systems in treponemes.
<3>Lett. Appl. Microbiol.
<4>38
<5>311-314
<6>2004
<7>Aims: To investigate the presence of GATC-specific modification and restriction activities in
rumen isolates of Treponema sp.
Methods: The presence of N-6-methyladenine within GATC (Dam)
sequences was analysed using isoschizomeric restriction endonucleases
having different sensitivities to the methylation of the target
sequence. A fast screening method was used for testing of site-specific
endonuclease activities directly in crude cell extracts. Three out of
six rumen isolates of Treponema sp. showed restriction activities.
Restriction endonucleases were further purified by Heparin-Sepharose
chromatography. Using PCR and specific primers, no sequence homologous
to the T. pallidum dam gene was found.
Conclusions: Three rumen treponemal strains were documented to
possess MboI isoschizomeric restriction- modification systems.
Significance: This is the first report on restriction activity in
rumen treponemes.

<>

<1>Pinarbasi, E., Elliott, J., Hornby, D.P.
<2>Activation of a yeast pseudo DNA methyltransferase by deletion of a single amino acid.
<3>J. Mol. Biol.
<4>257
<5>804-813
<6>1996
<7>The biological methylation of cytosine bases in DNA is central to such diverse phenomena as
restriction and modification in bacteria, repeat induced point-mutation (RIPing) in fungi and
for programming gene expression patterns in vertebrates.  Structural studies on HhaI DNA
methyltransferase, together with the sequence comparisons of around 40 cytosine-specific DNA
methyltransferases, have recently provided a molecular framework for understanding the
mechanism of action of the related group of enzymes that catalyse this base modification.
There are, however, a number of organisms, including Saccharomyces cerevisiae,
Schizosaccharomyces pombe and Drosophila melanogaster, which have no detectable DNA
methylation.  Here we report that the product of the pmt1 gene recently identified in S.
pombe, which contains most of the primary structure elements of a typical cytosine-specific
DNA methyltransferase, is catalytically inert owing to the insertion of a Ser residue between
the Pro-Cys motif found at the active site of all such DNA methyltransferases.  Following
deletion of this Ser residue, catalytic activity is restored and, using a range of DNA binding
experiments, it is shown that the enzyme recognises and methylates the sequence CC(A/T)GG, the
same sequence that is modified by the product of the Escherichia coli dcm gene.  The pmt gene
of S. pombe therefore encodes a pseudo DNA methyltransferase, which we have called psiM.SpoI.

<>

<1>Pinarbasi, E., Kan, M.S., Duran, C., Ford, G.C., Hornby, D.P.
<2>Substitution of the conserved phenylalanine in the S-adenosyl-L-methionine binding site of M.MspI with tyrosine modifies the kinetic properties of the enzyme.
<3>Biol. Chem.
<4>379
<5>591-594
<6>1998
<7>Cytosine (C-5)-specific DNA methyltransferases share a set of ten conserved motifs distributed
evenly throughout the entire polypeptide chain.  The first conserved motif contains a Phe,
which is intimately associated with cofactor recognition.  In the pseudo-DNA methyltransferase
M.SpoI, encoded by the pmt1 gene in Schizosaccharomyces pombe, a Tyr replaces this Phe
residue.  We describe the properties of a mutant form of M.MspI, a typical cytosine
(C-5)-specific DNA methyltransferase, in which Tyr replaces the conserved Phe.  This mutant
shows differences in ternary complex formation and in the pattern of covalent complex
formation with an inhibitory, fluorinated DNA duplex which may be due to anomalous hydrogen
bonding between the mutant Tyr hydroxyl group and the catalytic loop of the enzyme or through
interference with cofactor binding.

<>

<1>Pinarbasi, E., Pinarbasi, H., Hornby, D.P.
<2>Subcloning and expression of a cDNA encoding a Schizosaccharomyces pombe putative C-5 DNA methyltransferase.
<3>Turkish J. Biol.
<4>20
<5>325-331
<6>1996
<7>The pmt1+ gene, which shows a striking similarity to bacterial C-5 DNA methyltransferases, has
recently been isolated from Schizosaccharomyces pombe.  In this study, we have subcloned pmt1+
cDNA into the bacterial expression vector pET14b, which contains a sequence coding six
histidine residues upstream of the coding site so that the recombinant protein possesses a
histidine tag (Hig-Tag) at its N-terminus.  The constructed vector, which we have called
pETSPO1, encoding the pmt1+ cDNA, was then introduced into E. coli BL21 (DE3)pLysS cells and
the expressed recombinant protein was purified to homogeneity by nickel-chelate-affinity
chromatography.  Approximately 5 mg of His-Tag-pmt1+ fusion protein was purified from one
liter of induced culture.

<>

<1>Pinarbasi, H., Pinarbasi, E., Hornby, D.
<2>Recombinant alpha and beta subunits of M.AquI constitute an active DNA methyltransferase.
<3>J. Biochem. Mol. Biol.
<4>35
<5>348-351
<6>2002
<7>AquI DNA methyltransferase, M.AquI, catalyses the transfer of a methyl group from
S-adenosyl-L-methionine to the C5 position of the outermost
deoxycytidine base in the DNA sequence 5'CYCGRG3'. M.AquI is encoded by
two overlapping ORFs (termed a and P) instead of the single ORF that is
customary for Class II methyltransferase genes. The structural
organization of the M.AquI protein sequence is quite similar to that of
other bacteria] C5-DNA methyltransferases. Ten conserved motifs are
also present in the correct order, but only on two polypeptides. We
separately subcloned the genes that encode the alpha and beta subunits
of M.AquI into expression vectors. The overexpressed His-fusion alpha
and beta subunits of the enzyme were purified to homogeneity in a
single step by Nickel-chelate affinity chromatography. The purified
recombinant proteins were assayed for biological activity by an in
vitro DNA tritium transfer assay. The alpha and beta subunits of M.AquI
alone have no DNA methyltransferase activity, but when both subunits
are included in the assay, an active enzyme that catalyses the transfer
of the methyl group from S-adenosyl-L-methionine to DNA is
reconstituted. We also showed that the beta subunit alone contains all
of the information that is required to generate recognition of specific
DNA duplexes in the absence of the alpha subunit.

<>

<1>Pinarbasi, H., Pinarbasi, E., Hornby, D.P.
<2>Substitution of the conserved cysteine with glycine (Cys82Gly) of Agmenellum quadruplicatum methylase AquI (M.AquI) is not cytotoxic to  E. coli.
<3>Turkish J. Biol.
<4>25
<5>177-184
<6>2001
<7>Cytosine 5 DNA methyltransferases share ten conserved motifs. Motif IV contains an absolutely
conserved proline-cysteine dipeptide. The
cysteine residue of this motif is involved in catalysis by forming a
covalent bond with the 6-position of cytosine prior to methyl group
transfer. AquI DNA methyltransferase recognising the sequence CCCGGG is
a heterodimer unlike other C5 DNA methyltransferases. We changed the
conserved cysteine (Cys82) of M.AquI to serine and glycine. The
presence of mutations was confirmed by automated DNA sequencing.
Mutants were tested by in vivo plasmid protection assay and also by
transformation into mcrA+BC+ strain of E. coli. Replacement of the
conserved cysteine with serine led to an apparent loss of the
methyltransferring ability of the enzyme. Interestingly, it was found
that substitution of cysteine with glycine is not cytotoxic to E. coli
in the case of M.AquI.

<>

<1>Pinarbasi, H., Pinarbasi, E., Hornby, D.P.
<2>The small subunit of M.AquI is responsible for sequence-specific DNA recognition and binding in the absence of the catalytic domain.
<3>J. Bacteriol.
<4>185
<5>1284-1288
<6>2003
<7>AquI DNA methyltransferase (M. AquI) catalyzes the transfer of a methyl group from
S-adenosyl-L-methionine to the C5 position of the outermost
deoxycytidine base in the DNA sequence 5'-CCCGGG-3'. M. AquI is a
heterodimer in which the polypeptide chain is separated at the junction
between the two equivalent structural domains in the related enzyme M.
HhaI. Recently, we reported the subcloning, overexpression, and
purification of the subunits (alpha and beta) of M. AquI separately. Here
we describe the DNA binding properties of M. AquI. The results presented
here indicate that the beta subunit alone contains all of the information
for sequence-specific DNA recognition and binding. The first step in the
sequence-specific recognition of DNA by M. AquI involves the formation of
binary complex with the target recognition domain in conjunction with
conserved sequence motifs IX and X, found in all known C5 DNA
methyltransferases, contained in the beta subunit. The alpha subunit
enhances the binding of the beta subunit to DNA specifically and
nonspecifically. It is likely that the addition of the alpha subunit to
the beta subunit stabilizes the conformation of the beta subunit and
thereby enhances its affinity for DNA indirectly. Addition of
S-adenosyl-L-methionine and its analogues S-adenosyl-L-homocysteine and
sinefungin enhances binding, but only in the presence of the alpha
subunit. These compounds did not have any effect on DNA binding by the
beta subunit alone. Using a 30-mer oligodeoxynucleotide substrate
containing 5-fluorodeoxycytidine (5-FdC), it was found that the beta
subunit alone did not form a covalent complex with its specific sequence
in the absence or presence of S-adenosyl-L-methionine. However, the
addition of the alpha subunit to the beta subunit led to the formation of
a covalent complex with specific DNA sequence containing 5-FdC.

<>

<1>Pinard, M.
<2>Multitiered regulation of 5-cytosine DNA methyltransferase expression.
<3>Diss. Abstr.
<4>60
<5>5938
<6>2000
<7>The methylation of DNA at the 5 position of the cytosine moiety within CpG dinucleotides
sequences forms a complex pattern which delimits non-expressed genes.  For many years,
especially through studies of the correlation between methylation and gene silencing,
researchers have tried to decipher this genomic subtext.  Probably, a better way to achieve an
understanding of DNA methylation is by first elucidating what regulates the apparatus
responsible for its establishment.  The DNA MeTase is the enzyme responsible for the catalysis
of the DNA methylation reaction.  It was therefore the objective of this thesis to define some
of the factors that regulate the expression of this enzyme.  Preceding work had described a
promoter for the gene encoding the DNA MeTase.  Recent data has prompted some to put in doubt
the validity of this promoter.  In the first part of this thesis I present a report which
describes a selection between two possible promoters for the DNA MeTase gene effected by
methylation in P19 cells.  This data not only revalidates the previous description of the
promoter, but adds a new level of possible control.  In the second part, I elucidated the
elements within the minimal region of the aforementioned promoter which are necessary for
activity.  The results illustrate that most of the sequence plays a role in the promoter
function, and pRb is found to be able to modulate the activity in a novel cooperation between
AP1 and Sp1 which is absolutely necessary for the promoter to fulfil its purpose.  The third
part of the thesis is based on an effort to investigate the generality of oncogenic control of
DNA MeTase expression.  The promoter was previously demonstrated to be responsive to the
Ras/AP1 pathway leading to oncogenesis.  Therefore, another oncogene, the SV40 large T
antigen, known to act through a separate pathway from Ras and AP1, was transfected into BALB/c
3T3 cells in order to see how the expression of the DNA MeTase would be affected during this
transformation.  It was found that T antigen, through its actions on pRb, can induce DNA
MeTase expression by stabilizing its mRNA.  These studies establish that the DNA MeTase is a
nodal point of control by oncogenes and tumor suppressors which may need this enzyme in order
to execute their effects on the cell.

<>

<1>Pinard, M., Szyf, M.
<2>Regulation of cytosine DNA methyltransferase expression by tumor suppressors.
<3>Mol. Biol. Cell
<4>7
<5>174a
<6>1996
<7>In mammalian genomes, 60-80% of CpG dinucleotide sequences are methylated on the cytosine 5'
position.  These methylated sequences are distributed in a tissue specific pattern which is
correlated with gene silencing.  Recent data suggest a critical role for DNA MeTase in
oncogenesis.  The DNA methyltransferase gene expression has previously been shown to respond
to the oncogenic Ras pathway via AP-1 sites found in its promoter.  The effective silencing of
certain tumor suppressors by DNA methylation has also been demonstrated.  We hypothesize that
DNA MeTase acts as a pivotal point in the cross-talk between oncogenes and tumor suppressors.
In order to elucidate the effect of tumor suppressors on DNA MeTase expression, the SV40 T
antigen was stably transfected into BalbC3T3 cells to inhibit pRb activity.  We demonstrate
that expression of T antigen includes expression of DNA MeTase RNA as determined by RNAase
protection, of DNA MeTase protein as shown by Western blot analysis and of DNA MeTase activity
as assessed by an enzymatic activity assay.  DNA MeTase induction is at least partly
transcriptional as shown by nuclear Run-off assays.  By Southern blot assays we have observed
differences in the levels of methylation in several CpG island genes although not all to the
same extent.  Therefore, we clearly show that T antigen affects the regulation of DNA MeTase
and, in consequence, affects the methylation of specific genes.  These results may be the
first to show a feedback regulation of DNA MeTase by tumor suppressors which may mean that the
DNA MeTase is a center point in growth regulation by both oncogenes and tumor suppressors.

<>

<1>Pingoud, A.
<2>Spermidine increases the accuracy of type II restriction endonucleases.  Suppression of cleavage at degenerate, non-symmetrical sites.
<3>Eur. J. Biochem.
<4>147
<5>105-109
<6>1985
<7>The non-specific cleavage of DNA by type II restriction endonucleases (BamHI,
BsuRI, EcoRI, EcoRV, HindIII, PstI and SalI) can be effectively suppressed by
spermidine in millimolar concentrations, regardless of whether the non-specific
cleavage is induced by high concentrations of enzyme under optimal buffer
conditions or by high pH, low ionic strength, organic solvents and Mn2+ ions.
The increased specificity of restriction endonucleases in the presence of
spermidine is due to an enhancement of the cleavage rate at the canonical site
and a slowing down of the cleavage rate at related sites.  It is argued that
spermidine is essential for the high accuracy of restriction endonucleases in
vivo.

<>

<1>Pingoud, A., Alves, J., Fliess, A., Geiger, R., Rueter, T., Wolfes, H.
<2>Site-directed mutagenesis of the EcoRI restriction endonuclease.
<3>Biol. Chem. Hoppe Seyler
<4>368
<5>1093
<6>1987
<7>EcoRI is the only restriction endonuclease for which detailed structural
information is available.  Based on this information we have begun to study the
involvement of individual amino acid residues in the recognition process and in
catalysis by site directed mutagenesis.  In an effort to identify the essential
Glu and His residues of EcoRI we have replaced Glu 96, Glu 111, Glu 144 by Gln
and His 14, His 31 and His 114 by Asn.  All these single point mutations
drastically lower the catalytic effectivity of EcoRI, mainly by affecting the
kcat-value.  The Glu 96 mutant, furthermore, shows reduced specificity towards
its DNA substrate.  At present these results can only in part be explained by
the 3D structure of the EcoRI recognition complex as deduced from the x-ray
analysis of an EcoRI oligodeoxynucleotide complex crystallized in the absence
of Mg2+.

<>

<1>Pingoud, A., Alves, J., Geiger, R.
<2>Restriction Enzymes.
<3>Methods Mol. Biol.
<4>16
<5>107-200
<6>1993
<7>A review focussing on the practical aspects of the use of these enzymes.

<>

<1>Pingoud, A., Fuxreiter, M., Pingoud, V., Wende, W.
<2>Type II restriction endonucleases: structure and mechanism.
<3>Cell. Mol. Life Sci.
<4>62
<5>685-707
<6>2005
<7>Type II restriction endonucleases are components of restriction modification systems that
protect bacteria and archaea against invading
foreign DNA. Most are homodimeric or tetrameric enzymes that cleave DNA
at defined sites of 4 - 8 bp in length and require Mg2+ ions for
catalysis. They differ in the details of the recognition process and
the mode of cleavage, indicators that these enzymes are more diverse
than originally thought. Still, most of them have a similar structural
core and seem to share a common mechanism of DNA cleavage, suggesting
that they evolved from a common ancestor. Only a few restriction
endonucleases discovered thus far do not belong to the PD...D/ExK
family of enzymes, but rather have active sites typical of other
endonuclease families. The present review deals with new developments
in the field of Type II restriction endonucleases. One of the more
interesting aspects is the increasing awareness of the diversity of
Type II restriction enzymes. Nevertheless, structural studies
summarized herein deal with the more common subtypes. A major emphasis
of this review will be on target site location and the mechanism of
catalysis, two problems currently being addressed in the literature.

<>

<1>Pingoud, A., Geiger, R., Alves, J., Oelgeschlager, T., Ruter, T.
<2>Functional sites of the EcoRI restriction endonuclease probed by site directed mutagenesis.
<3>Biol. Chem. Hoppe Seyler
<4>370
<5>943-944
<6>1989
<7>The EcoRI restriction endonuclease recognizes with high specificity the double
stranded DNA sequence G^AATTC/CTTAA^G and in the presence of Mg2+ cleaves the
DNA as indicated.  A 3 Angstrom X-ray structure analysis of an EcoRI-DNA
complex, crystallized in the absence of Mg2+, indicates that the amino acid
residues in positions 144 and 145 as well as 200 are involved in DNA binding.
Site directed mutagenesis experiments have shown that their involvement in the
recognition process, however, is more intricate than proposed by the authors of
the X-ray structure analysis.  To define more precisely the role of these and
adjacent amino acids we have produced single and double mutants of EcoRI with
amino acid replacements in position 144, 145, 147 as well as 199, 200, 203 and
analyzed their activity in vivo and in vitro after purification to homogeneity.
The results of the in vivo assay allows us to conclude that the structural
integrity of the region at and around position 200 is very critical for the
enzymatic function of EcoRI:  nonconservative mutations lead to a dramatic
decrease in activity.  In contrast, the region around position 144 and 145
seems to be of minor importance for the enzymatic activity:  non-conservative
mutations in this region have similar moderate effects as conservative
mutations.  The analysis of the purified EcoRI mutants confirms these results.
In addition, they demonstrate that some mutants which have a very low activity
are still specific for the EcoRI site (Arg200->Gly), while others show a
relaxed specificity (Arg200->Glu or Gln and Asn199->Asp).  The Asn199->Asp
mutant, furthermore, attacks preferentially the central phosphodiester bond in
its degenerate hexanucleotide recognition sequence.  Taken together our results
suggest that not only Glu144, Arg145 and Arg200 are involved in specificity
determining interactions but other amino acids as well.  The recognition
process, furthermore, does not only depend on hydrogen bonds but also on the
overall complementarity of charge dipoles, and presumably is highly redundant.
We have begun to locate the catalytic center and the Mg2+ binding site of
EcoRI.  Two candidate regions were analyzed:
1.  92GGIVEVKD - DYGEWR
2. 126LLVGKRGDQDLMAAG
which show a homology to a consensus sequence proposed to be involved in Mg2+
binding in a variety of polymerases.  Preliminary results demonstrate that the
second region is more critical than the first one for the catalytic action of
EcoRI.

<>

<1>Pingoud, A., Jeltsch, A.
<2>Recognition and cleavage of DNA by type-II restriction endonucleases.
<3>Eur. J. Biochem.
<4>246
<5>1-22
<6>1997
<7>Restriction endonucleases are enzymes which recognize short DNA sequences and cleave the DNA
in both strands.  Depending on the enzymological properties different types are distinguished.
Type II restriction endonucleases are homodimers which recognize short palindromic sequences
4-8 bp in length and, in the presence of Mg2+, cleave the DNA within or next to the
recognition site.  They are capable of non-specific binding to DNA and make use of linear
diffusion to locate their target site.  Binding and recognition of the specific site involves
contacts to the bases of the recognition sequence and the phosphodiester backbone over
approximately 10-12 bp.  In general, recognition is highly redundant which explains the
extreme specificity of these enzymes.  Specific binding is accompanied by conformational
changes over both the protein and the DNA.  This mutual induced fit leads to the activation of
the catalytic centers.  The precise mechanism of cleavage has not yet been established for any
restriction endonuclease.  Currently two models are discussed: the substrate-assisted
catalysis mechanism and the two-metal-ion mechanism.  Structural similarities identified
between EcoRI, EcoRV, BamHI, PvuII and Cfr10I suggest that many type II restriction
endonucleases are not only functionally but also evolutionarily related.  A review.

<>

<1>Pingoud, A., Jeltsch, A.
<2>Structure and function of type II restriction endonucleases.
<3>Nucleic Acids Res.
<4>29
<5>3705-3727
<6>2001
<7>More than 3000 type II restriction endonucleases have been discovered. They recognize short,
usually palindromic, sequences of 4-8 bp and, in the presence of Mg(2+), cleave the DNA within
or in close proximity to the recognition sequence. The orthodox type II enzymes are homodimers
which recognize palindromic sites. Depending on particular features subtypes are classified.
All structures of restriction enzymes show a common structural core comprising four
beta-strands and one alpha-helix. Furthermore, two families of enzymes can be distinguished
which are structurally very similar (EcoRI-like enzymes and EcoRV-like enzymes). Like other
DNA binding proteins, restriction enzymes are capable of non-specific DNA binding, which is
the prerequisite for efficient target site location by facilitated diffusion. Non-specific
binding usually does not involve interactions with the bases but only with the DNA backbone.
In contrast, specific binding is characterized by an intimate interplay between direct
(interaction with the bases) and indirect (interaction with the backbone) readout. Typically
approximately 15-20 hydrogen bonds are formed between a dimeric restriction enzyme and the
bases of the recognition sequence, in addition to numerous van der Waals contacts to the bases
and hydrogen bonds to the backbone, which may also be water mediated. The recognition process
triggers large conformational changes of the enzyme and the DNA, which lead to the activation
of the catalytic centers. In many restriction enzymes the catalytic centers, one in each
subunit, are represented by the PD...D/EXK motif, in which the two carboxylates are
responsible for Mg(2+) binding, the essential cofactor for the great majority of enzymes. The
precise mechanism of cleavage has not yet been established for any enzyme, the main
uncertainty concerns the number of Mg(2+) ions directly involved in cleavage. Cleavage in the
two strands usually occurs in a concerted fashion and leads to inversion of configuration at
the phosphorus. The products of the reaction are DNA fragments with a 3'-OH and a
5'-phosphate.

<>

<1>Pingoud, A., Jeltsch, A., Lanio, T., Schulze, C., Selent, U., Stahl, F., Wende, W., Wenz, C.
<2>Mechanism of DNA recognition and cleavage by restriction enzymes.
<3>Biol. Chem. Hoppe Seyler
<4>376
<5>S138
<6>1995
<7>The crystal structure analyses of specific DNA complexes of EcoRI, EcoRV and PvuII have
greatly aided in our understanding of how these enzymes recognize and cleave their DNA
substrate.  Many questions, however, remain, in particular, how is the target site located,
what is the relative contribution of direct and indirect readout for the recognition process,
how do the two subunits of the homodimeric restriction enzymes cooperate in recognition as
well as cleavage, and, what is the precise mechanism of phosphodiester bond hydrolysis.  Using
EcoRV as a model restriction enzyme we have tried to answer some of these questions.  It is
apparent from our studies that EcoRV makes use of facilitated diffusion along the DNA to
locate its target site and that Mg ions are required for specific binding which leads to a
pronounced DNA bending.  During recognition, contacts are formed to the bases and the
backbone: base contacts are in general more important than phosphate contacts, the latter,
however, are needed for binding, linear diffusion and to make sites with different flanking
sequences equally accessible for EcoRV,  Mutants of EcoRV have been constructed that exhibit
preferences for sites containing a modified base or backbone as well as for certain flanking
sequences.  We have shown that the two identical subnits of EcoRV communicate with each other
in DNA recognition, not, however, in catalysis per se.  Accordingly, artifical heterodimers
could be produced that exhibit a pronounced nicking activity.  While the catalytic amino acid
residues and the stereochemical course for the reaction have been identfied for EcoRV, it is
still not clear how the attacking hydroxide ion is generated and how many metal ions are
involved in the catalytic mechanism.  Based on the current experimental evidence, we believe
that a water molecule is activated by the phosphate residue 3' to the scissile phosphodiester
bond, that only one Mg ion is needed to polarize the P-O bond, to stabilize the transition
state and to supply a water molecule for protonation of the leaving group.  It is becoming
increasingly clear that restriction enzymes, which in general do not share significant
sequence homology, have structural and mechanistic features in common.  Indeed, using a
genotypic and phenotypic analysis it has been possible to demonstrate a common evolutionary
history for these enzymes.

<>

<1>Pingoud, A., Jeltsch, A., Maxwell, A., Sherratt, D.
<2>Enzymes that keep DNA under control.
<3>EMBO Rep.
<4>2
<5>271-276
<6>2001
<7>Life demands not only the faithful and controlled replication of DNA but, in addition, many
other enzymatic processes involving DNA, including topoisomerase action, recombination,
repair, restriction and modification.  These topics and their interrelationships were
discussed at the IUBMB symposium in Bangalore (India) on DNA enzymes: structures and
mechanisms (December 1-3, 2000), which was organized by V. Nagaraja and D.N. Rao (Bangalore,
India) and brought together about 200 scientists from all over the world.  Whereas most
presentations focused on detailed biochemical and genetic mechanisms, several reminded us that
enzymes and DNA are components of complex living organisms that live, die and evolve.

<>

<1>Pingoud, A., Pingoud, V., Wende, W.
<2>Homing endonucleases - multifaceted tools for genome engineering.
<3>BIOspektrum
<4>9
<5>592-595
<6>2003
<7>
<>

<1>Pingoud, A., Silva, G.H.
<2>Precision genome surgery.
<3>Nat. Biotechnol.
<4>25
<5>743-744
<6>2007
<7>Zinc finger nucleases hold great potential for gene therapy as they allow one to cleave DNA at
a chosen target sequence.  Unfortunately, however, they also cleave prolifically at off-target
sites, resulting in unacceptable toxicity.  Two papers in this issue, by Miller et al. and
Szczepek et al., show that off-target cleavage can be greatly reduced by rationally
redesigning the ZFN dimer interface to inhibit homodimerization.  The goal of gene therapy is
to repair genetic defects without othewise modifying the genome.  One of the most promising
strategies for gene correction is homologous recombination.  In this approach, the correct DNA
sequence is saupplied in trans and is integrated into the genome by an endogenous mechanism of
site-specific recombination.  Ideally, the deficient gene is replaced with a functional copy
in its natural context while leaving the rest of the genome untouched.  Homologous
recombination can also be used for gene inactivation, insertion or deletion.

<>

<1>Pingoud, A., Thielking, V., Selent, U., Kohler, E., Liedtke, M., Wolfes, H., Pieper, U., Geiger, R., Winkler, F.K.
<2>Identification of functional sites in the EcoRV restriction endonuclease by site-directed mutagenesis.
<3>J. Biomol. Struct. Dyn.
<4>8
<5>A165
<6>1991
<7>Guided by the X-ray structure analysis of a crystalline EcoRV d(GGGATATCCC)
complex (Winkler, unpublished) we have replaced various amino acid residues in
EcoRV by site-directed mutagenesis in order to identify those amino acids which
participate in the binding and cleavage of DNA.  Mutant enzymes were isolated
and characterized physico-chemically and biochemically.  Our results show that
three regions are of importance for DNA binding: region I around Ser183,
Asn185, Thr186 and Asn188, region II around Gln69 and Asn70 and region III
comprising the C-terminus.  The conservative substitution of these amino acids
leads to mutant enzymes of no or considerably reduced activity but unaltered
specificity, indicating that the process of recognition is tightly coupled to
catalysis and highly redundant.  Region I contains a sequence motif
(-SerGlyXXXAsnIIeXSer-) which is also found in other restriction enzymes
recognizing the sequence G--/--C or G-/-C, the dashes representing A or T.
Region III is homologous to a sequence found in SmaI.  Presumably the catalytic
center is formed by Asp74 and Asp90, which may serve to bind Mg2+ and activate
water for a nucleophilic attack, and Lys92 which may hold the scissile
phosphodiester bond in place.  The mutation of Asp74 to Gln leads to a nearly
inactive enzyme.  Other mutations are currently being characterized.  It is
noteworthy that in EcoRI a similar sequence motif (-ProAsp...Glu(Asp)XLys-) is
found in the vicinity of the scissile phosphodiester bond.

<>

<1>Pingoud, A., Urbanke, C., Alves, J., Ehbrecht, H.-J., Zabeau, M., Gualerzi, C.
<2>Effect of polyamines and basic proteins on cleavage DNA by restriction endonucleases.
<3>Biochemistry
<4>23
<5>5697-5703
<6>1984
<7>We have investigated the effect of the polyamines spermine, spermidine, and
putrescine and the prokaryotic histone-like proteins NS1 and NS2 on the
restriction endonuclease EcoRI catalyzed cleavage of plasmid and bacteriophage
DNAs.  At low concentrations of spermine and spermidine, the rate of DNA
cleavage by EcoRI is increased, while high concentrations of spermine as well
as of spermidine are inhibitory.  These phenomena are also observed with other
restriction endonucleases.  They are, therefore, probably due to the
interaction of the polyamines with the DNA.  Putrescine does not have such an
effect within the concentration range investigated.  Remarkably, low
concentrations of spermine and spermidine very efficiently suppress EcoRI
activity.  An inhibition of the EcoRI-catalyzed cleavage of DNA is also
observed with NS1 and NS2, an effect that can be mimicked with other basic
proteins that interact with DNA.  The results are discussed in terms of the
mechanism of restriction in vivo.

<>

<1>Pingoud, A., Wende, W.
<2>A sliding restriction enzyme pauses.
<3>Structure
<4>15
<5>391-393
<6>2007
<7>In this issue of Structure, Aggarwal and colleagues (Townson et al., 2007) present the crystal
structure of the restriction endonuclease BstYI in
complex with a near-cognate substrate. This structure most likely reflects
the conformation BstYI adopts as it scans DNA and pauses upon encountering
a site similar to its recognition sequence.

<>

<1>Pingoud, A., Wende, W.
<2>Generation of Novel Nucleases with Extended Specificity by Rational and Combinatorial Strategies.
<3>Chembiochem
<4>12
<5>1495-1500
<6>2011
<7>After the discovery of DNA as the molecule carrying the hereditary information of all living
organisms, DNA processing enzymes, in particular restriction endonucleases, led to the
development of recombinant DNA technology. Today, in the postgenomic era, a steadily growing
number of entire genome sequences is available.  The emerging genome-engineering technology
requires more sophisticated nucleases with very high specificity, that ideally target a unique
sequence in a complex genome.  The design and control of these rare-cutting endonucleases by
using rational and evolutionary strategies is the focus of this Minireview.

<>

<1>Pingoud, A., Wilson, G.G., Wende, W.
<2>Type II restriction endonucleases-a historical perspective and more.
<3>Nucleic Acids Res.
<4>42
<5>7489-7527
<6>2014
<7>This article continues the series of Surveys and Summaries on restriction endonucleases
(REases) begun this year in Nucleic Acids Research. Here we discuss 'Type II' REases, the
kind used for DNA analysis and cloning. We focus on their biochemistry: what they are, what
they do, and how they do it. Type II REases are produced by prokaryotes to combat
bacteriophages. With extreme accuracy, each recognizes a particular sequence in
double-stranded DNA and cleaves at a fixed position within or nearby. The discoveries of these
enzymes in the 1970s, and of the uses to which they could be put, have since impacted every
corner of the life sciences. They became the enabling tools of molecular biology, genetics and
biotechnology, and made analysis at the most fundamental levels routine. Hundreds of different
REases have been discovered and are available commercially. Their genes have been cloned,
sequenced and overexpressed. Most have been characterized to some extent, but few have been
studied in depth. Here, we describe the original discoveries in this field, and the properties
of the first Type II REases investigated. We discuss the mechanisms of sequence recognition
and catalysis, and the varied oligomeric modes in which Type II REases act. We describe the
surprising heterogeneity revealed by comparisons of their sequences and structures.

<>

<1>Pingoud, V., Conzelmann, C., Kinzebach, S., Sudina, A., Metelev, V., Kubareva, E., Bujnicki, J.M., Lurz, R., Luder, G., Xu, S.Y., Pingoud, A.
<2>PspGI, a type II restriction endonuclease from the extreme thermophile Pyrococcus sp.: structural and functional studies to investigate an evolutionary relationship with several mesophilic restriction enzymes.
<3>J. Mol. Biol.
<4>329
<5>913-929
<6>2003
<7>We present here the first detailed biochemical analysis of an archaeal restriction enzyme.
PspGI shows sequence similarity to SsoII, EcoRII,
NgoMIV and Cfr10I, which recognize related DNA sequences. We demonstrate
here that PspGI, like SsoII and unlike EcoRII or NgoMIV and Cfr10I,
interacts with and cleaves DNA as a homodimer and is not stimulated by
simultaneous binding to two recognition sites. PspGI and SsoII differ in
their basic biochemical properties, viz. stability against chemical
denaturation and proteolytic digestion, DNA binding and the pH, MgCl(2)
and salt-dependence of their DNA cleavage activity. In contrast, the
results of mutational analyses and cross-link experiments show that PspGI
and SsoII have a very similar DNA binding site and catalytic center as
NgoMIV and Cfr10I (whose crystal structures are known), and presumably
also as EcoRII, in spite of the fact that these enzymes, which all
recognize variants of the sequence -/CC-GG- (/ denotes the site of
cleavage), are representatives of different subgroups of type II
restriction endonucleases. A sequence comparison of all known restriction
endonuclease sequences, furthermore, suggests that several enzymes
recognizing other DNA sequences also share amino acid sequence
similarities with PspGI, SsoII and EcoRII in the region of the presumptive
active site. These results are discussed in an evolutionary context.

<>

<1>Pingoud, V., Geyer, H., Geyer, R., Kubareva, E., Bujnicki, J.M., Pingoud, A.
<2>Identification of base-specific contacts in protein-DNA complexes by photocrosslinking and mass spectrometry: A case study using the  restriction endonuclease SsoII.
<3>Mol. BioSyst.
<4>1
<5>135-141
<6>2005
<7>Specific protein-nucleic acid interactions are of paramount importance for the propagation,
maintenance and expression of genetic information.  Restriction endonucleases serve as model
systems to study the mechanisms of DNA recognition by proteins.  SsoII is a Type II
restriction endonuclease that recognizes the double stranded sequence / CCNGG and cleaves it
in the presence of Mg2+-ions, as indicated.  SsoII shows sequence similarity over a stretch of
~70 amino acid residues with several other restriction endonucleases that recognize a similar
sequence as SsoII (Cfr10I, EcoRII, NgoMIV, PspGI).  In the NgoMIV this stretch is involved in
DNA recognition and cleavage, as shown by the crystal structure analysis of an enzyme-product
complex.  To find out whether the presumptive DNA recognition region in SsoII is indeed in
contact with DNA we have photocrosslinked SsoII with an oligodeoxyribonucleotide in which the
first guanine of the recognition sequence was replaced by 5-iodouracil.  Following digestion
by trypsin, the peptide-oligodeoxyribonucleotide conjugate was purified by Fe3+-IMAC and then
incubated with hydrogen fluoride, which hydrolyzes the oligodeoxyribonucleotide to yield the
peptide-deoxyuridine conjugate.  The site of photocrosslinking was identified by MALDI-TOF-MS
and MALDI-TOF-MS/MS to be Trp189, adjacent to Arg188, which aligns with Arg194 in NgoMIV,
involved in recognition of the second guanine in the NgoMIV recognition sequence G/CCGGC.
This result confirms previously published conclusions drawn on the basis of a mutational
analysis of SsoII.  The methodology that was employed here can be used in principle to
identify the DNA binding site of any protein.

<>

<1>Pingoud, V., Grindl, W., Wende, W., Thole, H., Pingoud, A.
<2>Structural and functional analysis of the homing endonuclease PI-SceI by limited proteolytic cleavage and molecular cloning of partial digestion products.
<3>Biochemistry
<4>37
<5>8233-8243
<6>1998
<7>PI-SceI is a member of an unusual class of rare cutting homing endonucleases produced by an
autocatalytic protein splicing from a precursor.  To analyze the structural and functional
domain organization of the endonuclease PI-SceI and to examine whether the DNA binding
activity can be structurally separated from the catalytic activity, we performed limited
proteolytic digestion experiments with various proteases.  Two protease-resistant fragments
spanning the N- and C-terminal halves of the nuclease were identified using different
proteases which cleave the protein in the same region. Each fragment contains one of the two
conserved LAGLIDADG motifs.  The products of the limited proteolytic digests were shown to
remain associated and to exhibit specific DNA binding but to be inactive in DNA cleavage.
Different from what is observed with native PI-SceI, only one complex is formed as shown in an
electrophoretic mobility shift assay.  Expression clones for the N- and C-terminal protein
fragments obtained by tryptic digestion were constructed, and the proteins PI-SceI-N and
PI-SceI-C were purified.  Only PI-SceI-N exhibits DNA binding activity.  Bending experiments
with PI-SceI-N, a mixture of PI-SceI-N and PI-SceI-C, as well as the products of the limited
tryptic digest show that a DNA substrate with the full length recognition sequence is bent by
45 degrees.  This degree of bending is also observed with a DNA containing only the right side
of the recognition sequence, corresponding to one of the DNA cleavage products of PI-SceI.
Our results demonstrate that the N-terminal half of PI-SceI which lacks one of the two
LAGLDADG motifs is able to bind to DNA specifically and to induce one of the distortions
observed to occur in the process of DNA binding by PI-SceI.  These results are discussed in
light of the recently solve crystal structure of PI-SceI and used to refine a model for the
mechanism of DNA binding and cleavage by PI-SceI.

<>

<1>Pingoud, V., Kubareva, E., Stengel, G., Friedhoff, P., Bujnicki, J.M., Urbanke, C., Sudina, A., Pingoud, A.
<2>Evolutionary relationship between different subgroups of restriction endonucleases.
<3>J. Biol. Chem.
<4>277
<5>14306-14314
<6>2002
<7>The type II restriction endonuclease SsoII shows sequence similarity with 10 other restriction
endonucleases, among them the type IIE
restriction endonuclease EcoRII, which requires binding to an effector
site for efficient DNA cleavage, and the type IIF restriction
endonuclease NgoMIV, which is active as a homotetramer and cleaves DNA
with two recognition sites in a concerted reaction. We show here that
SsoII is an orthodox type II enzyme, which is active as a homodimer and
does not require activation by binding to an effector site.
Nevertheless, it shares with EcoRII and NgoMIV a very similar
DNA-binding site and catalytic center as shown here by a mutational
analysis, indicative of an evolutionary relationship between these
three enzymes. We suggest that a similar relationship exists between
other orthodox type II, type IIE, and type IIF restriction
endonucleases. This may explain why similarities may be more pronounced
between members of different subtypes of restriction enzymes than among
the members of a given subtype.

<>

<1>Pingoud, V., Sudina, A., Geyer, H., Bujnicki, J.M., Lurz, R., Luder, G., Morgan, R., Kubareva, E., Pingoud, A.
<2>Specificity changes in the evolution of Type II restriction endonucleases - A biochemical and bioinformatic analysis of restriction  enzymes that recognize unrelated sequences.
<3>J. Biol. Chem.
<4>280
<5>4289-4298
<6>2005
<7>How restriction enzymes with their different specificities and mode of cleavage evolved has
been a long standing question in evolutionary
biology. We have recently shown that several Type II restriction
endonucleases, namely SsoII (down arrow CCNGG), PspGI (^CCWGG), Eco-RII (^CCWGG), NgoMIV
(G^CCGGC), and Cfr10I (R^CCGGY), which recognize similar DNA sequences (as indicated, where ^
denotes cleavage position), share limited
sequence similarity over an interrupted stretch of approximately 70 amino
acid residues with MboI, a Type II restriction endonuclease from
Moraxella bovis (Pingoud, V., Conzelmann, C., Kinzebach, S., Sudina,
A., Metelev, V., Kubareva, E., Bujnicki, J.M., Lurz, R., Luder, G., Xu,
S. Y., and Pingoud, A. (2003) J. Mol. Biol 329, 913-929). Nevertheless,
MboI has a dissimilar DNA specificity (^GATC) compared with
these enzymes. In this study, we characterize MboI in detail to
determine whether it utilizes a mechanism of DNA recognition similar to
SsoII, PspGI, EcoRII, NgoMIV, and Cfr10I. Mutational analyses and
photocross-linking experiments demonstrate that MboI exploits the
stretch of approximately 70 amino acids for DNA recognition and cleavage.
It is therefore likely that MboI shares a common evolutionary origin
with SsoII, PspGI, EcoRII, NgoMIV, and Cfr10I. This is the first
example of a relatively close evolutionary link between Type
II restriction enzymes of widely different specificities.

<>

<1>Pingoud, V., Thole, H., Christ, F., Grindl, W., Wende, W., Pingoud, A.
<2>Photocross-linking of the homing endonuclease PI-SceI to its recognition sequence.
<3>J. Biol. Chem.
<4>274
<5>10235-10243
<6>1999
<7>PI-SceI is an intein-encoded protein that belongs to the LAGLIDADG family of homing
endonucleases.  According to the crystal structure and mutational studies, this endonuclease
consists of two domains, one responsible for protein splicing, the other for DNA cleavage, and
both presumably for DNA binding.  To define the DNA binding site of PI-SceI,
photocross-linking was used to identify amino acid residues in contact with DNA.  Sixty-three
double-stranded oligodeoxynucleotides comprising the minimal recognition sequence and
containing single 5-iodopyrimidine substitutions in almost all positions of the recognition
sequence were synthesized and irradiated in the presence of PI-SceI with a helium/cadmium
laser (325 nm).  The best cross-linking yield (approximately 30%) was obtained with an
oligodeoxynucleotide with a 5-iododeoxyuridine at position +9 in the bottom strand.  The
subsequent analysis showed that cross-linking had occurred with amino acid His-333, 6 amino
acids after the second LAGLIDADG motif.  With the H333A variant of PI-SceI or in the presence
of excess unmodified oligodeoxynucleotide, no cross-linking was observed, indicating the
specificity of the cross-linking reaction.  Chemical modification of His residues in PI-SceI
by diethylpyrocarbonate leads to a substantial reduction in the binding and cleavage activity
of PI-SceI.  This inactivation can be suppressed by substrate binding.  This result further
supports the finding that at least one His residue is in close contact to the DNA.  Based on
these and published results, conclusions are drawn regarding the DNA binding site of PI-SceI.

<>

<1>Pingoud, V., Wende, W., Friedhoff, P., Reuter, M., Alves, J., Jeltsch, A., Mones, L., Fuxreiter, M., Pingoud, A.
<2>On the Divalent Metal Ion Dependence of DNA Cleavage by Restriction Endonucleases of the EcoRI Family.
<3>J. Mol. Biol.
<4>393
<5>140-160
<6>2009
<7>Restriction endonucleases of the PD...D/EXK family need Mg(2+) for DNA cleavage. Whereas
Mg(2+) (or Mn(2+)) promotes catalysis, Ca(2+) (without
Mg(2+)) only supports DNA binding. The role of Mg(2+) in DNA cleavage by
restriction endonucleases has elicited many hypotheses, differing mainly
in the number of Mg(2+) involved in catalysis. To address this problem, we
measured the Mg(2+) and Mn(2+) concentration dependence of DNA cleavage by
BamHI, BglII, Cfr10I, EcoRI, EcoRII (catalytic domain), MboI, NgoMIV,
PspGI, and SsoII, which were reported in co-crystal structure analyses to
bind one (BglII and EcoRI) or two (BamHI and NgoMIV) Me(2+) per active
site. DNA cleavage experiments were carried out at various Mg(2+) and
Mn(2+) concentrations at constant ionic strength. All enzymes show a
qualitatively similar Mg(2+) and Mn(2+) concentration dependence. In
general, the Mg(2+) concentration optimum (between approximately 1 and 10
mM) is higher than the Mn(2+) concentration optimum (between approximately
0.1 and 1 mM). At still higher Mg(2+) or Mn(2+) concentrations, the
activities of all enzymes tested are reduced but can be reactivated by
Ca(2+). Based on these results, we propose that one Mg(2+) or Mn(2+) is
critical for restriction enzyme activation, and binding of a second Me(2+)
plays a role in modulating the activity. Steady-state kinetics carried out
with EcoRI and BamHI suggest that binding of a second Mg(2+) or Mn(2+)
mainly leads to an increase in K(m), such that the inhibitory effect of
excess Mg(2+) or Mn(2+) can be overcome by increasing the substrate
concentration. Our conclusions are supported by molecular dynamics
simulations and are consistent with the structural observations of both
one and two Me(2+) binding to these enzymes.

<>

<1>Pinto, S.B., Stainton, K., Harris, S., Kambris, Z., Sutton, E.R., Bonsall, M.B., Parkhill, J., Sinkins, S.P.
<2>Transcriptional Regulation of Culex pipiens Mosquitoes by Wolbachia Influences Cytoplasmic Incompatibility.
<3>PLoS Pathog.
<4>9
<5>E1003647
<6>2013
<7>Cytoplasmic incompatibility (CI) induced by the endosymbiont Wolbachia pipientis
causes complex patterns of crossing sterility between populations of the Culex
pipiens group of mosquitoes. The molecular basis of the phenotype is yet to be
defined. In order to investigate what host changes may underlie CI at the
molecular level, we examined the transcription of a homolog of the Drosophila
melanogaster gene grauzone that encodes a zinc finger protein and acts as a
regulator of female meiosis, in which mutations can cause sterility. Upregulation
was observed in Wolbachia-infected C. pipiens group individuals relative to
Wolbachia-cured lines and the level of upregulation differed between lines that
were reproductively incompatible. Knockdown analysis of this gene using RNAi
showed an effect on hatch rates in a Wolbachia infected Culex molestus line.
Furthermore, in later stages of development an effect on developmental
progression in CI embryos occurs in bidirectionally incompatible crosses. The
genome of a wPip Wolbachia strain variant from Culex molestus was sequenced and
compared with the genome of a wPip variant with which it was incompatible. Three
genes in inserted or deleted regions were newly identified in the C. molestus
wPip genome, one of which is a transcriptional regulator labelled wtrM. When this
gene was transfected into adult Culex mosquitoes, upregulation of the grauzone
homolog was observed. These data suggest that Wolbachia-mediated regulation of
host gene expression is a component of the mechanism of cytoplasmic
incompatibility.

<>

<1>Pintor-Toro, J.A.
<2>Adenine methylation in zein genes.
<3>Biochem. Biophys. Res. Commun.
<4>147
<5>1082-1087
<6>1987
<7>This paper reports the novel finding of adenine methylation in higher plants. Comparison of
restriction patterns of genomic maize DNA digested with enzymes MboI and Sau3AI enabled us to
detect the existence of adenine methylation in zein genes. Adenine methylation within or
around zein genes turned out to be similar in endosperm (where zeins are actively synthesized)
and in seedling tissue (where zein genes are not expressed). Furthermore, adenine methylation
patterns were found to be similar both in wild-type and opaque-2 mutant plants. These lines of
evidence suggest that adenine methylation is unrelated to the regulation of gene expression.

<>

<1>Pique, N., Aquilini, E., Alioto, T., Minana-Galbis, D., Tomas, J.M.
<2>Genome Sequence of Plesiomonas shigelloides Strain 302-73 (Serotype O1).
<3>Genome Announcements
<4>1
<5>e00404-13
<6>2013
<7>Plesiomonas shigelloides, the only species of the genus, is an emergent pathogenic bacterium
associated with human diarrheal and extraintestinal disease.
We present the whole-genome sequence analysis of the representative strain for
the O1 serotype (strain 302-73), providing a tool for studying bacterial
outbreaks, virulence factors, and accurate diagnostic methods.

<>

<1>Pirrotta, V.
<2>Two restriction endonucleases from Bacillus globigii.
<3>Nucleic Acids Res.
<4>3
<5>1747-1760
<6>1976
<7>The sites of action of the restriction enzyme BglII on lambda DNA are mapped. This enzyme
recognises the sequence 5'...AGATCT...3' and makes staggered cuts producing sticky ends. In
lambda DNA, the second A in this sequence is methylated about 50% of the time by a bacterial
methylase absent in E. coli dam-. In contrast to BglII, BglI makes many cuts in lambda DNA and
produces 5' terminals which are not substrates for polynucleotide kinase.

<>

<1>Pirrotta, V., Bickle, T.A.
<2>General purification schemes for restriction endonucleases.
<3>Methods Enzymol.
<4>65
<5>89-95
<6>1980
<7>The range of organisms used for the production of restriction enzymes and the
various properties of these enzymes is such that it is difficult to devise a
purification scheme of general application.  the combination of
polyethyleneimine precipitation and chromatography on heparin-agarose discussed
in this article has a sufficiently wide applicability and a number of
advantages to recommend it as the backbone of a general purification scheme or
as a first approach in the isolation of new restriction endonuclease
activities.  In individual cases, however, additions or modifications of this
procedure are necessary to optimize the results.  In this article we will first
present the basic procedure and then discuss alternatives or modifications
suitable to certain particular cases.

<>

<1>Pitluck, S. et al.
<2>Complete genome sequence of Calditerrivibrio nitroreducens type strain (Yu37-1).
<3>Standards in Genomic Sciences
<4>4
<5>54-62
<6>2011
<7>Calditerrivibrio nitroreducens Iino et al. 2008 is the type species of the genus
Calditerrivibrio. The species is of interest because of its important role in the
nitrate cycle as nitrate reducer and for its isolated phylogenetic position in
the Tree of Life. Here we describe the features of this organism, together with
the complete genome sequence and annotation. This is the third complete genome
sequence of a member of the family Deferribacteraceae. The 2,216,552 bp long
genome with its 2,128 protein-coding and 50 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Pitluck, S. et al.
<2>Complete genome sequence of Thermosediminibacter oceani type strain (JW/IW-1228P).
<3>Standards in Genomic Sciences
<4>3
<5>108-116
<6>2010
<7>Thermosediminibacter oceani (Lee et al. 2006) is the type species of the genus
Thermosediminibacter in the family Thermoanaerobacteraceae. The anaerobic,
barophilic, chemoorganotrophic thermophile is characterized by straight to curved
Gram-negative rods. The strain described in this study was isolated from a core
sample of deep sea sediments of the Peruvian high productivity upwelling system.
This is the first completed genome sequence of a member of the genus
Thermosediminibacter and the seventh genome sequence in the family
Thermoanaerobacteraceae. The 2,280,035 bp long genome with its 2,285
protein-coding and 63 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Pitluck, S. et al.
<2>Non-contiguous finished genome sequence of Aminomonas paucivorans type strain (GLU-3).
<3>Standards in Genomic Sciences
<4>3
<5>285-293
<6>2010
<7>Aminomonas paucivorans Baena et al. 1999 is the type species of the genus Aminomonas, which
belongs to the family Synergistaceae. The species is of
interest because it is an asaccharolytic chemoorganotrophic bacterium which
ferments quite a number of amino acids. This is the first finished genome
sequence (with one gap in a rDNA region) of a member of the genus Aminomonas and
the third sequence from the family Synergistaceae. The 2,630,120 bp long genome
with its 2,433 protein-coding and 61 RNA genes is a part of the
GenomicEncyclopedia ofBacteria andArchaea project.

<>

<1>Pitsikas, P., Cupples, C.
<2>Effects of increasing DCM expression on Vsr repair of site specific and related sequences.
<3>FASEB J.
<4>11
<5>A1369
<6>1997
<7>DCM, the only cytosine methylase in Escherichia coli, methylates the second cytosine of this
specific site: CCAGG.  However, overexpression of dcm leads to methylation of other related
sequences as well.  5-methylcytosine spontaneously deaminates to thymine.  Vsr repairs T/G
mismatches, and restores the cytosine both at CTAGG sites and at related sequences.  Our
experiments aim to demonstrate what sequences are methylated by Dcm and which ones are
repaired by Vsr, and show if there is overlap between the two.  We have regulated the
expression of Dcm by placing dcm under the PBAD promoter and by varying the concentration of
arabinose from 0% to 0.2%.  The results showed variation over 3 orders of magnitude in Dcm
expression.  Further work will be done to analyze Vsr repair.

<>

<1>Pitsikas, P., Cupples, C.G.
<2>Fate of Dcm cytosine methylase after treatment of Escherichia coli with 5-Azacytidine.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>99
<5>334
<6>1999
<7>5-azacytidine is a chemotherapeutic agent used in the treatment of leukemia.  When
incorporated into DNA as 5-azacytosine, it causes C-to-G transversion mutations.  In addition,
when it replaces cytosine in methylase recognition sites, transfer of the methyl group cannot
take place, and the enzyme remains covalently bound to the DNA.  The resulting inactivation of
the methyltransferase leads to DNA demethylation.  We are interested in determining how the
DNA-methylase complexes are removed, how the DNA is repaired, and what becomes of the enzyme.
As a first step, we have made an antibody to Dcm, the sole cytosine methylase in Escherichia
coli, and used it to quantify the levels of free protein present in the cells after treatment
with 5-azacytidine.  We find that levels of analog which are sufficient to cause an increase
in C-to-G mutations result in a large reduction in the amount of Dcm.  The levels of an
enzymatically inactive mutant are unaffected.  Transcription of the dcm gene is apparently not
autoregulated.  We are currently combining western analysis with Southern analysis to
determine the fate of the DNA/protein complexes.

<>

<1>Pittard, J.
<2>Effect of phage-controlled restriction on genetic links in bacterial crosses.
<3>J. Bacteriol.
<4>87
<5>1256-1257
<6>1964
<7>Arber and Dussoix (J. Mol. Biol. 5:18, 1962) demonstrated that
lambda-bacteriophage, when grown on Escherichia coli K-12, plates with an
efficiency of 1 on E. coli K-12 but with an efficiency of only 2 X 10-5 on E.
coli K-12 (P1) which is lysogenic for bacteriophage Pl.  They showed that the
low efficiency of plating is a result of the breakdown of the
lambda-deoxyribonucleic acid (DNA) injected into the Pl lysogenic host. The
present paper reports some studies of recombination between a P1 sensitive Hfr,
AB2229, and a P1 lysogenic recipient, AB2147, in which two effects of
restriction on the recovery of recombinants were observed.

<>

<1>Pittet, V., Abegunde, T., Marfleet, T., Haakensen, M., Morrow, K., Jayaprakash, T., Schroeder, K., Trost, B., Byrns, S., Bergsveinson, J., Kusalik, A., Ziola, B.
<2>Complete Genome Sequence of the Beer Spoilage Organism Pediococcus claussenii ATCC BAA-344T.
<3>J. Bacteriol.
<4>194
<5>1271-1272
<6>2012
<7>Pediococcus claussenii is a common brewery contaminant. We have sequenced the chromosome and
plasmids of the type strain P. claussenii ATCC BAA-344. A ropy
variant was chosen for sequencing to obtain genetic information related to growth
in beer, as well as exopolysaccharide and possibly biofilm formation by this
organism.

<>

<1>Pittet, V., Ewen, E., Bushell, B.R., Ziola, B.
<2>Genome Sequence of Lactobacillus rhamnosus ATCC 8530.
<3>J. Bacteriol.
<4>194
<5>726
<6>2012
<7>Lactobacillus rhamnosus is found in the human gastrointestinal tract and is important for
probiotics. We became interested in L. rhamnosus isolate
ATCC 8530 in relation to beer spoilage and hops resistance. We report here
the genome sequence of this isolate, along with a brief comparison to
other available L. rhamnosus genome sequences.

<>

<1>Pizza, M., Hickey, E.T., Peterson, J.T., Tettelin, H.T., Masignani, V., Galeotti, C., Mora, M., Ratti, G., Scarselli, M., Scarlato, V., Rappuoli, R., Fraser, C.M., Grandi, G., Venter, C.T.
<2>
<3>European Patent Office
<4>EP 1605061 A
<5>
<6>2005
<7>The invention provides methods of obtaining immunogenic proteins from genomic sequences
including Neisseria, including the amino acid sequences and the corresponding nucleotide
sequences, as well as the genomic sequence of Neisseria meningitidis B.  The proteins so
obtained are useful antigens for vaccines, immunogenic compositions, and/or diagnostics.

<>

<1>Planet, P.J., Rampersaud, R., Hymes, S.R., Whittier, S., Della-Latta, P.A., Narechania, A., Daugherty, S.C., Santana-Cruz, I., Desalle, R., Ravel, J., Ratner, A.J.
<2>Genome Sequence of the Human Abscess Isolate Streptococcus intermedius BA1.
<3>Genome Announcements
<4>1
<5>e00117-12
<6>2013
<7>Streptococcus intermedius is a human pathogen with a propensity for abscess formation. We
report a high-quality draft genome sequence of S. intermedius strain BA1, an isolate from a
human epidural abscess. This sequence provides insight into the biology of S. intermedius and
will aid investigations of pathogenicity.

<>

<1>Platonov, M.E., Blouin, Y., Evseeva, V.V., Afanas'ev, M.V., Pourcel, C., Balakhonov, S.V., Vergnaud, G., Anisimov, A.P.
<2>Draft Genome Sequences of Five Yersinia pseudotuberculosis ST19 Isolates and One  Isolate Variant.
<3>Genome Announcements
<4>1
<5>e00122-13
<6>2013
<7>We report the first draft genome sequences of five Yersinia pseudotuberculosis isolates of
sequence type (ST) 19 and of a variant from one of the five isolates.
The total length of assemblies ranged from 4,226,485 bp to 4,274,148 bp,
including between 3,808 and 3,843 predicted coding sequences.

<>

<1>Plesch, G., Blau, A., Daeschner, K.
<2>Method for identifying substances having a herbicide action.
<3>International Patent Office
<4>WO 2004022780 A
<5>
<6>2004
<7>The invention relates to a method for identifying compounds having a herbicide action.  The
invention also relates to nucleic acid constructs, to vectors containing said nucleic acid
constructs, to transgenic organisms, and to the use of the same.  Also disclosed are
substances which have been identified by means of the above-mentioned method.

<>

<1>Plessis, A., Perrin, A., Haber, J.E., Dujon, B.
<2>Site-specific recombination determined by I-SceI, a mitochondrial group I intron-encoded endonuclease expressed in the yeast nucleus.
<3>Genetics
<4>130
<5>451-460
<6>1992
<7>The Saccharomyces cerevisiae mitochondrial endonuclease I-SceI creates a double-strand break
as the initiating step in the gene conversional transfer of the omega+ intron to omega- DNA.
We have expressed a glactose-inducible synthetic I-SceI gene in the nucleus of yeast that also
carries the I-SceI recognition site on a plasmid substrate.  We find that the
galactose-induced I-SceI protein can be active in the nucleus and efficiently catalyze
recombination.  With a target plasmid containing direct repeats of the Escherichia coli lacZ
gene, one copy of which is interrupted by a 24-bp cutting site, galactose induction produces
both deletions and gene conversions.  Both the kinetics and the proportion of deletions and
gene conversions are very similar to analogous events initiated by a galactose-inducible HO
endonuclease gene.  We also find that, in a rad52 mutant strain, the repair of double-strand
breaks initiated by I-SceI and by HO are similarly affected: the formation of deletions is
reduced, but not eliminated.  Altogether, these results suggest either that the two
endonucleases act in the same way after double-strand break formation or that the two
endonucleases are not involved in subsequent steps.

<>

<1>Pljevaljcic, G., Pignot, M., Weinhold, E.
<2>Design of a new fluorescent cofactor for DNA methyltransferases and sequence-specific labeling of DNA.
<3>J. Am. Chem. Soc.
<4>125
<5>3486-3492
<6>2003
<7>Sequence-specific labeling of DNA is of immense interest for analytical and functional studies
of DNA. We present a novel approach for
sequence-specific labeling of DNA using a newly designed fluorescent
cofactor for the DNA methyltransferase from Thermus aquaticus (M.TaqI).
Naturally, M.TaqI catalyzes the nucleophilic attack of the exocyclic amino
group of adenine within the double-stranded 5'-TCGA-3' DNA sequence onto
the methyl group of the cofactor S-adenosyl-L-methionine (AdoMet) leading
to methyl group transfer. The design of a new fluorescent cofactor for
covalent labeling of DNA was based on three criteria: (1) Replacement of
the methionine side chain of the natural cofactor AdoMet by an aziridinyl
residue leads to M.TaqI-catalyzed nucleophilic ring opening and coupling
of the whole nucleoside to DNA. (2) The adenosyl moiety is the molecular
anchor for cofactor binding. (3) Attachment of a fluorophore via a
flexible linker to the 8-position of the adenosyl moiety does not block
cofactor binding. According to these criteria the new fluorescent cofactor
8-amino[1'-(N'-dansyl)-4'-aminobutyl]-5'-(1-aziridinyl)-5'-deoxyadenosi
ne (3) was synthesized. 3 binds about 4-fold better than the natural
cofactor AdoMet to M.TaqI and is coupled with a short duplex
oligodeoxynucleotide by M.TaqI. The identity of the expected modified
nucleoside was verified by electrospray ionization mass spectrometry after
enzymatic fragmentation of the product duplex. In addition, the new
cofactor 3 was used to sequence-specifically label plasmid DNA in a
M.TaqI-catalyzed reaction.

<>

<1>Pljevaljcic, G., Schmidt, F., Peschlow, A., Weinhold, E.
<2>Sequence-specific DNA labeling using methyltransferases.
<3>Methods Mol. Biol.
<4>283
<5>145-161
<6>2004
<7>Sequence-specific labeling of native deoxyribonucleic acid (DNA) still represents a
more-or-less unsolved problem. Difficulties mainly arise from
the necessity to combine two different functions: sequence-specific
recognition of DNA and covalent bond formation between the label and DNA.
DNA methyltransferases (MTases) naturally possess these two functions and
transfer a methyl group from the cofactor S-adenosyl-L-methionine (AdoMet)
to adenine or cytosine residues within specific DNA sequences, typically
ranging from two to eight base pairs. Unfortunately, the methyl group
itself is a very limited reporter group and it would be desirable to
transfer larger chemical entities with DNA MTases. Replacement of the
methionine side chain of the natural cofactor AdoMet by an aziridinyl
residue leads to the synthetic cofactor N-adenosylaziridine, which is
quantitatively, base- and sequence-specifically coupled with DNA in a DNA
MTase-catalyzed reaction. By attaching interesting reporter groups to a
suitable position of N-adenosylaziridine a large variety of new synthetic
cofactors are obtained for sequence-specific labeling of DNA. This method
is illustrated by coupling primary amino groups and biotin to short duplex
oligodeoxynucleotides or plasmid DNA using the DNA MTase M.TaqI.

<>

<1>Pljevaljcic, G., Schmidt, F., Scheidig, A.J., Lurz, R., Weinhold, E.
<2>Quantitative labeling of long plasmid DNA with nanometer precision.
<3>Chembiochem
<4>8
<5>1516-1519
<6>2007
<7>
<>

<1>Pljevaljcic, G., Schmidt, F., Weinhold, E.
<2>Sequence-specific methyltransferase-induced labeling of DNA (SMILing DNA).
<3>Chembiochem
<4>5
<5>265-269
<6>2004
<7>A new concept for sequence-specific labeling of DNA by using chemically modified cofactors for
DNA methyltransferases is presented. Replacement
of the amino acid side chain of the natural cofactor
S-adenosyl-L-methionine with an aziridine group leads to a cofactor
suitable for DNA methyltransferase-catalyzed sequence-specific coupling
with DNA. Sequence-specifically fluorescently labeled plasmid DNA was
obtained by using the DNA methyltransferase from Thermus aquaticus
(M.TaqI) as catalyst and attaching a fluorophore to the aziridine
cofactor. First results suggest that all classes of DNA
methyltransferases with different recognition sequences can be used. In
addition, this novel method for DNA labeling should be applicable to a
wide variety of reporter groups.

<>

<1>Plugge, C.M., Henstra, A.M., Worm, P., Swarts, D.C., Paulitsch-Fuchs, A.H., Scholten, J.C., Lykidis, A., Lapidus, A.L., Goltsman, E., Kim, E., McDonald, E., Rohlin, L., Crable, B.R., Gunsalus, R.P., Stams, A.J., McInerney, M.J.
<2>Complete genome sequence of Syntrophobacter fumaroxidans strain (MPOB(T)).
<3>Standards in Genomic Sciences
<4>7
<5>91-106
<6>2012
<7>strain MPOB is the best-studied species of the genus . The species is of interest because of
its anaerobic syntrophic lifestyle, its involvement in the conversion
of propionate to acetate, H and CO during the overall degradation of organic
matter, and its release of products that serve as substrates for other
microorganisms. The strain is able to ferment fumarate in pure culture to CO and
succinate, and is also able to grow as a sulfate reducer with propionate as an
electron donor. This is the first complete genome sequence of a member of the
genus and a member genus in the family . Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
4,990,251 bp long genome with its 4,098 protein-coding and 81 RNA genes is a part
of the Microbial Genome Program (MGP) and the Genomes to Life (GTL) Program
project.

<>

<1>Plumbridge, J.
<2>The role of dam methylation in controlling gene expression.
<3>Biochimie
<4>69
<5>439-443
<6>1987
<7>The E. coli gene dam encodes an adenine-specific methylase which converts all adenine residues
in the sequence GATC into 6-methyl adenine. dam type methylases seem to be ubiquitous in
Enterobacteriaceae and Haemophilus, since DNA from these strains cross hybridizes with the
cloned dam gene and they contain DNA resistant to MboI cleavage, which only cleaves
non-methylated DNA. However, dam methylation is absent from most species of eubacteria not
phylogenetically related to E. coli and presumably evolved late in prokaryotic evolution. At
least, one reason why people have become more aware of dam (and dcm) methylation in the last
decade is because certain restriction enzymes are sensitive to DNA methylation and hence
overlapping dam and restriction sites are not cleaved. A third phenotype for the dam mutations
has been described in the literature: that of controlling the level of expression of a
miscellaneous selection of genes with GATC sequences in their promoter regions. It is this
third phenotype which is the subject of this paper, but initially it is worth summarizing the
evidence for the dam methylation affecting other phenotypes of the E. coli bacteria.

<>

<1>Plunkett, G. III, Rose, D.J., Durfee, T.J., Blattner, F.R.
<2>Sequence of Shiga toxin 2 phage 933W from Escherichia coli O157:H7: Shiga toxin as a phage late-gene product.
<3>J. Bacteriol.
<4>181
<5>1767-1778
<6>1999
<7>Lysogenic bacteriophages are major vehicles for the transfer of genetic
information between bacteria, including pathogenicity and/or virulence
determinants. In the enteric pathogen Escherichia coli O157:H7, which
causes hemorrhagic colitis and hemolytic-uremic syndrome, Shiga toxins 1
and 2 (Stx1 and Stx2) are phage encoded. The sequence and analysis of the
Stx2 phage 933W is presented here. We find evidence that the toxin genes
are part of a late-phage transcript, suggesting that toxin production may
be coupled with, if not dependent upon, phage release during lytic growth.
Another phage gene, stk, encodes a product resembling eukaryotic
serine/threonine protein kinases. Based on its position in the sequence,
Stk may be produced by the prophage in the lysogenic state, and, like the
YpkA protein of Yersinia species, it may interfere with the signal
transduction pathway of the mammalian host. Three novel tRNA genes present
in the phage genome may serve to increase the availability of rare tRNA
species associated with efficient expression of pathogenicity
determinants: both the Shiga toxin and serine/threonine kinase genes
contain rare isoleucine and arginine codons. 933W also has homology to
lom, encoding a member of a family of outer membrane proteins associated
with virulence by conferring the ability to survive in macrophages, and
bor, implicated in serum resistance.

<>

<1>Poch, M.T., Somkuti, G.A.
<2>Detection of restriction endonuclease activity in Streptococcus thermophilus ST132.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>93
<5>330
<6>1993
<7>Genetic development of Streptococcus thermophilus (ST), an important industrial bacterium in
dairy fermentations, requires the understanding of enzymes that influence the survival and
replication of transforming DNAs. We have detected a new restriction endonuclease (R-ENase),
designated as Sth132I, in sonically disrupted cell extracts of strain ST132. Ion exchange
chromatography (DEAE-cellulose) of crude extracts with a continuous salt gradient resolved
Sth132I (50-100mM Kcl) and nonspecific exonucleases which coprecipitated by ammonium sulfate
fractionation. Sth132I had several recognition sites on PhiX174 (5.38 kbp), a single site each
on pVA736 (7.6 kbp) and pERB (2.2 kbp) but failed to digest pBR322 (4.36 kbp) and lambda DNA.
Single, double and triple digestions performed on the model pER8 (a native plasmid of ST108)
allowed tentative positioning of the Sth132I recognition sie at coordinate 0.22 kbp but the
enzyme was not identifiable as one of the known single-restriction R-NEses. Sth132I required
Mg++ and retained activity up to 55oC.

<>

<1>Poch, M.T., Somkuti, G.A.
<2>Rapid isolation of restriction endonucleases from Streptococci.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>385
<6>1994
<7>Streptococcus thermophilus can be genetically-engineered to favorably alter characteristics of
fermented dairy products.  However, endogeneous restriction-modification systems in these
bacteria can influence the survival and replication of transforming DNAs. A rapid, microscale
Heparin Sepharose CL-6B affinity gel procedure was developed for detection of restriction
endonucleases (RE-Nases) in S. thermophilus and other Streptococcus species.
RE-Nase-containing extracts free of DNA, RNA and nonspecific nuclease activity were produced
for forty or more strains daily. RE-Nase activity was detected in 10 of 44 strains of
Streptococci assayed.        	SagI, a new isoschizomer of HaeIII was purified using this
single-step method from S. agalactiae. This highly active isoschizomer (12,000 U/g of dry
cells) was free of nonspecific nuclease activity and was suitable for use in molecular biology
procedures. The isolation procedures was applicable for the rapid isolation of RE-Nases from
other lactic acid bacteria.

<>

<1>Poch, M.T., Somkuti, G.A.
<2>Isolation of SagI, a new HaeIII isoschizomer from Streptococcus agalactiae.
<3>Appl. Microbiol. Biotechnol.
<4>43
<5>282-284
<6>1995
<7>A new HaeIII isoschizomer from Streptococcus agalactiae was isolated by a single-step
purification method. The highly active restriction endonuclease, SagI, was free of nonspecific
nuclease activity and was suitable for use in molecular biology procedures. The rapid
isolation procedure may be applicable for the recovery of other restriction endonucleases from
bacteria.

<>

<1>Poch, M.T., Somkuti, G.A.
<2>Rapid screening of lactic acid bacteria for restriction endonuclease activity.
<3>Biotechnol. Tech.
<4>7
<5>781-784
<6>1993
<7>A rapid microscale heparin Sepharose CL-6B affinity gel procedure was developed for detecting
restriction endonuclease (RE-Nase) activity in a variety of lactic acid bacteria.
Re-Nase-containing extracts free of DNA, RNA and nonspecific nuclease activity can be produced
for forty or more strains daily and only 10-12 ml of each log phase culture as required for
screening. RE-Nase activity was tested in several streptococci and lactobacilli. With
appropriate modifications, this procedure should allow rapid detection of RE-Nase activity in
other bacterial species.

<>

<1>Poch, M.T., Somkuti, G.A., Solaiman, D.K.Y.
<2>Sth132I, a novel class-IIS restriction endonuclease of Streptococcus thermophilus ST132.
<3>Gene
<4>195
<5>201-206
<6>1997
<7>The Sth132I restriction endonuclease (R.Sth132I) was detected in Streptococcus thermophilus
ST132 and purified to near homogeneity by heparin Sepharose CL-6B affinity chromatography.
Fragments from Sth132I digestion of plasmid DNA were subcloned into pUC19 in Escherichia coli
DH5a and sequenced.  Sequence analysis of inserts and their ligation junction sites revealed
that Sth132I is a novel class-IIS restriction endonuclease, which recognizes the
non-palindromic sequence 5'-CCCG(N)4-3' 3'-GGGC(N)8-5'.

<>

<1>Podgorska, B., Kujawska, G., Skurzewski, M., Batsko, O., Kaczorowski, T.
<2>A rapid and simple method for detection of type II restriction endonucleases in cells of bacteria with high activity of nonspecific nucleases.
<3>Acta Biochim. Pol.
<4>59
<5>669-672
<6>2012
<7>In this work we describe a novel, rapid and simple microscale procedure for identification of
restriction endonuclease activity in bacteria
lysates, which contain high levels of non-specific DNA nucleases.

<>

<1>Podhajska, A.J., Kim, S.C., Szybalski, W.
<2>Conferring new specificities on restriction enzymes: cleavage at any predetermined site by combining adapter oligodeoxynucleotide and Class-IIS enzyme.
<3>Methods Enzymol.
<4>216
<5>303-309
<6>1992
<7>
<>

<1>Podhajska, A.J., Szybalski, W.
<2>Conversion of the FokI endonuclease to a universal restriction enzyme:  cleavage of phage M13mp7 DNA at predetermined sites.
<3>Gene
<4>40
<5>175-182
<6>1985
<7>Endonuclease FokI belongs to class IIS of restriction enzymes, for which the
DNA cut points lie outside the enzyme-recognition sites.  This permitted
conferring new cleavage specificities by combining FokI with tailored
oligodeoxynucleotide adapters.  Such adapters carry a single-stranded (ss)
target-recognition domain, complementary to the selected ss target DNA, and a
double-stranded (ds) enzyme-recognition site.  Neither enzyme nor adapter alone
has endonucleolytic activity toward phage M13mp7 ss DNA, whereas the
enzyme-adapter complex cleaves this ss target DNA at the particular sites fore
ordained by the sequence of the ss domain of the adapter.  Two kinds of
adapters (32 and 34 nucleotides long), with opposing orientations of the
asymmetric FokI recognition site, were constructed and shown to direct specific
cleavage under a variety of conditions.  In addition to FokI, other class IIS
enzymes, HphI, MboII and BbvI, which alone do not cleave ss DNA, are suitable
for construction of tailored enzyme-adapter complexes with predictable cleavage
specificities.  This report provides a preliminary experimental confirmation
for the proposal of Szybalski [Gene 40 (1985) 169-173] for the design of
adapter-enzyme complexes with novel and predictable specificities.
Theoretically, using this approach any sequence could be precisely cleaved at a
predetermined point.

<>

<1>Poehlein, A., Alghaithi, H.S., Chandran, L., Chibani, C.M., Davydova, E., Dhamotharan, K., Ge, W., Gutierrez-Gutierrez, D.A., Jagirdar, A., Khonsari, B., Nair, K.P., Daniel, R.
<2>First Insights into the Genome of the Amino Acid-Metabolizing Bacterium Clostridium litorale DSM 5388.
<3>Genome Announcements
<4>2
<5>e00754-14
<6>2014
<7>Clostridium litorale is a Gram-positive, rod-shaped, and spore-forming bacterium, which is
able to use amino acids such as glycine, sarcosine, proline, and betaine
as single carbon and energy sources via Stickland reactions. The genome consists
of a circular chromosome (3.41 Mb) and a circular plasmid (27 kb).

<>

<1>Poehlein, A., Andreesen, J.R., Daniel, R.
<2>Complete Genome Sequence of Amino Acid-Utilizing Eubacterium acidaminophilum al-2 (DSM 3953).
<3>Genome Announcements
<4>2
<5>e00573-14
<6>2014
<7>Eubacterium acidaminophilum is a strictly anaerobic, Gram-positive, rod-shaped
bacterium which belongs to cluster XI of the Clostridia. It ferments amino acids
by a Stickland reaction. The genome harbors a chromosome (2.25 Mb) and a
megaplasmid (0.8 Mb). It contains several gene clusters coding for
selenocysteine-containing, glycine-derived, and amino acid-degrading reductases.

<>

<1>Poehlein, A., Daniel, R., Simeonova, D.D.
<2>Draft Genome Sequence of Desulfotignum phosphitoxidans DSM 13687 Strain FiPS-3.
<3>Genome Announcements
<4>1
<5>e00227-13
<6>2013
<7>We report the 5.008-Mbp assembled draft genome sequence of Desulfotignum phosphitoxidans
strain FiPS-3 (DSM 13687), which gains metabolic energy from the
oxidation of phosphite to phosphate. Its genome provides insights into the
composition and architecture of the phosphite-utilizing and energy-transducing
systems required to live with phosphite as electron donor.

<>

<1>Poehlein, A., Deutzmann, J.S., Daniel, R., Simeonova, D.D.
<2>Draft Genome Sequence of the Methanotrophic Gammaproteobacterium Methyloglobulus  morosus DSM 22980 Strain KoM1.
<3>Genome Announcements
<4>1
<5>e01078-13
<6>2013
<7>Here, we report the draft genome sequence of the methanotrophic gammaproteobacterium
Methyloglobulus morosus DSM 22980 strain KoM1, which is
proposed to be the type species for the novel genus Methyloglobulus. The genome
(4.143 Mb) consists of a single circular chromosome and harbors genes for
2-aminoethylphosphonate (ciliatine) biosynthesis.

<>

<1>Poehlein, A., Freese, H.M., Daniel, R., Simeonova, D.D.
<2>Draft Genome Sequence of Serratia sp. Strain DD3, Isolated from the Guts of Daphnia magna.
<3>Genome Announcements
<4>2
<5>e00903-14
<6>2014
<7>We report the draft genome sequence of Serratia sp. strain DD3, a gammaproteobacterium from
the family Enterobacteriaceae. It was isolated from
homogenized guts of Daphnia magna. The genome size is 5,274 Mb.

<>

<1>Poehlein, A., Gottschalk, G., Daniel, R.
<2>First Insights into the Genome of the Gram-Negative, Endospore-Forming Organism Sporomusa ovata Strain H1 DSM 2662.
<3>Genome Announcements
<4>1
<5>e00734-13
<6>2013
<7>The genome of Sporomusa ovata strain H1 DSM 2662, an anaerobic, Gram-negative
endospore-forming bacterium, was sequenced. S. ovata uses N-methyl compounds,
primary alcohols, fatty acids, and H2 and CO2 as energy and carbon sources to
produce acetate. The genome harbors one chromosome, which encodes proteins
typical for sporulation.

<>

<1>Poehlein, A., Hartwich, K., Krabben, P., Ehrenreich, A., Liebl, W., Durre, P., Gottschalk, G., Daniel, R.
<2>Complete Genome Sequence of the Solvent Producer Clostridium saccharobutylicum NCP262 (DSM 13864).
<3>Genome Announcements
<4>1
<5>e00997-13
<6>2013
<7>Clostridium saccharobutylicum was employed for the production of acetone and butanol in South
Africa until the 1970s. The genome comprises a single replicon
(5,107,814 bp) harboring all the genes necessary for solvent production and the
degradation of various organic compounds, such as fructose, cellobiose, sucrose,
and mannose.

<>

<1>Poehlein, A., Krabben, P., Durre, P., Daniel, R.
<2>Complete Genome Sequence of the Solvent Producer Clostridium saccharoperbutylacetonicum Strain DSM 14923.
<3>Genome Announcements
<4>2
<5>e01056-14
<6>2014
<7>Clostridium saccharoperbutylacetonicum strain DSM 14923 is known as a butanol-producing
bacterium. Various organic compounds such as glucose, fructose,
sucrose, mannose, and cellobiose are fermented. The genome consists of one
chromosome and one circular megaplasmid. C. saccharoperbutylacetonicum was used
in industrial fermentation processes to produce the solvents acetone, butanol,
and ethanol.

<>

<1>Poehlein, A., Kusian, B., Friedrich, B., Daniel, R., Bowien, B.
<2>Complete genome sequence of the type strain Cupriavidus necator N-1.
<3>J. Bacteriol.
<4>193
<5>5017
<6>2011
<7>Here we announce the complete genome sequence of the copper-resistant bacterium Cupriavidus
necator N-1, the type strain of the genus
Cupriavidus. The genome consists of two chromosomes and two circular
plasmids. Based on genome comparison the chromosomes of C. necator N-1
share a high degree of similarity with the two chromosomal replicons of
the bioplastic-producing hydrogen bacterium Ralstonia eutropha H16. The
two strains differ in their plasmids and the presence of hydrogenase
genes, which are absent in strain N-1.

<>

<1>Poehlein, A., Zverlov, V.V., Daniel, R., Schwarz, W.H., Liebl, W.
<2>Complete Genome Sequence of Clostridium stercorarium subsp. stercorarium Strain DSM 8532, a Thermophilic Degrader of Plant Cell Wall Fibers.
<3>Genome Announcements
<4>1
<5>e0007313
<6>2013
<7>Clostridium stercorarium strain DSM 8532 is a thermophilic bacterium capable of efficiently
degrading polysaccharides in plant biomass and converting the
resulting sugars to ethanol and acetate. The complete genome sequence of 2.96 Mbp
reveals a multitude of genes for hydrolytic enzymes and enables further study of
the organism and its enzymes, and their exploitation for biotechnological
processes.

<>

<1>Pogge von Strandmann, R., Stadtler, R., Walter, T., Frey, B., Kaluza, K., Hengstenberg, W., Schmitz, G.
<2>BpuAI, a novel BbsI and BbvII isoschizomer from Bacillus pumilus recognizing 5'-GAAGAC-3'.
<3>Nucleic Acids Res.
<4>20
<5>4664
<6>1992
<7>We have isolated BpuAI, a novel class-IIs restriction endonuclease from Bacillus pumilus
recognizing the sequence 5'-GAAGAC-3', cutting at N2-3' and N6-5'. High amounts of
activity can be purified due to a fast protocol and the presence of high amounts of specific
activity in the crude extract.

<>

<1>Pogolotti, A.L., Ono, A., Subramaniam, R., Santi, D.V.
<2>On the mechanism of DNA-adenine methylase.
<3>J. Biol. Chem.
<4>263
<5>7461-7464
<6>1988
<7>Experiments were performed to determine whether EcoRI methylase catalyzes the
transfer of the methyl group of S-adenosylmethionine (a) directly to the N6 of
adenine in DNA or (b) initially to N1 to give N1-methyladenine followed by
isomerization of the N1-methylamino and 6-NH2 to give N6-methyladenine (Dimroth
rearrangement).  A facile synthesis of highly enriched [6-15N]deoxyadenosine
and a dodecamer substrate of EcoRI methylase with [6-15N]adenine in the
methylation site are reported.  In the product of EcoRI enzymatic methylation,
all of the isotope remains at the N6 position of the N6-methyladenine product.
It is concluded that, contrary to existing chemical precedent, the methylation
occurs by direct transfer from S-adenosylmethionine to the N6 of adenine in
DNA.

<>

<1>Pohl, F.M., Thomae, R., Karst, A.
<2>Temperature dependence of the activity of DNA-modifying enzymes: endonucleases and DNA ligase.
<3>Eur. J. Biochem.
<4>123
<5>141-152
<6>1982
<7>The activities of 17 endonucleases: the restriction endonucleases AvaI, BamHI,
EcoRI, HindIII, PstI and SalI, which cleave pBR322 DNA once: AluI, AvaII, CfoI,
HaeIII, HhaI, HinfI, HpaII and TaqI, which cut pBR322 DNA several times, and
three "unspecific" nucleases (S1 nuclease, staphylococcal nuclease and DNase I
from bovine pancreas) were determined between 0C and 65C.  The reaction was
followed by the disappearance of covalently closed circular pBR322 DNA, using
the alkaline ethidium fluorescence assay of Morgan et al. [Nucl. Acids Res.
(1979) 7, 547-594]; the activity of T4 DNA ligase was similarly measured by the
conversion of nicked circular DNA to closed circular DNA.  For each enzyme,
small aliquots of the same solution were incubated at different temperatures
simultaneously in a temperature gradient device, resulting in a high relative
precision.  The experimental results are summarized by the simplest possible
theoretical description, using linear or exponential kinetics and apparent
activation energies Ea for the enzymatic reaction, Ei for the enzyme
inactivation and Ti for the inactivation temperature.  To a good approximation
these three parameters suffice for describing the temperature dependence of the
activity of most of the enzymes.

<>

<1>Poland, B.W., Xu, M.Q., Quiocho, F.A.
<2>Structural Insights into the Protein Splicing Mechanism of PI-SceI.
<3>J. Biol. Chem.
<4>275
<5>16408-16413
<6>2000
<7>PI-SceI is a member of a class of proteins (inteins) that excise themselves from a precursor
protein and in the process ligate the flanking protein sequences (exteins). We report here the
2.1-A resolution crystal structure of a PI-SceI miniprecursor (VMA29) containing 10 N-terminal
extein residues and 4 C-terminal extein residues. Mutations at the N- and C-terminal splicing
junctions, blocking in vivo protein splicing, allowed the miniprecursor to be purified and
crystallized. The structure reveals both the N- and C-terminal scissile peptide bonds to be in
distorted trans conformations (tau approximately 100 degrees ). Modeling of the wild-type
PI-SceI based on the VMA29 structure indicates a large conformational change (movement of >9
A) must occur to allow transesterification to be completed. A zinc atom was discovered at the
C-terminal splicing junction. Residues Cys(455), His(453), and Glu(80) along with a water
molecule (Wat(53)) chelate the zinc atom. The crystal structure of VMA29 has captured the
intein in its pre-spliced state.

<>

<1>Polishchuk, L.V., Lukyanchuk, V.V., Matselyuch, B.P.
<2>Site-specific endonucleases of Streptomycetes.
<3>Actinomycetes
<4>10
<5>13-15
<6>2000
<7>Production of restriction enzymes is widespread among soil streptomycetes. More than 15% of
fresh soil isolates showed this ability. Of eight strains showing enzyme activity, seven
formed isoschizomers of AsuII. Enzymes with such specificity were not previously found amongst
streptomycetes.  Enzymes of restriction-modification (RM} systems are present in large amounts
in streptomycetes and some strains are strong producers of endonucleases (e.g., SacI, SacII,
SalGI). Streptomycetes also produce isoschizomers of EcoRI, PstI and others. Studies on these
enzymes are valuable for understanding regulation and functioning of RM systems, actual
production of the enzymes themselves and for pharmaceutical and other biotechnological
applications (Rodicio & Chater, 1988}. The aim of the present study was to investigate the
amount of restrictases among fresh soil isolates.

<>

<1>Polisky, B., Greene, P., Garfin, D.E., McCarthy, B.J., Goodman, H.M., Boyer, H.W.
<2>Specificity of substrate recognition by the EcoRI restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>72
<5>3310-3314
<6>1975
<7>The substrate specificity of the EcoRI restriction endonuclease can be varied
in vitro by changing the pH and the ionic environment of the reaction.
Phosphodiester bond cleavage occurs at a DNA hexanucleotide sequence
d(N-G-A-A-T-T-C-N) d(N-C-T-T-A-A-G-N)	when the ionic strength is high, 100mM
Tris-HCl, 50 mM NaCl, 5 mM MgCl2, and the pH is approximately 7.3.  Lowering
the ionic strength to 25 mM Tris-HCl, 2 mM MgCl2, and adjusting the pH to 8.5
reduces the recognition specificity of the EcoRI endonuclease to the
tetranucleotide sequence, d(N-A-A-T-T-N) d(N-T-T-A-A-N)	The enzymatic activity
responsible for this substrate recognition is referred to as EcoRI*.  Cleavage
of pVH51 plasmid DNA under EcoRI* conditions results in a number of partial
digest fragments, some of which disappear slowly over a prolonged digestion
period.  This suggests that different recognition sites are cleaved at
different rates.  Comparison of DNA fragment patterns of modified and
unmodified pVH51 DNA indicates that the canonical EcoRI sequence is the most
rapidly cleaved site under EcoRI* conditions.  DNA modified in vivo by the
EcoRI methylase is not cleaved by the EcoRI endonuclease under standard
conditions, but is cleaved under EcoRI* conditions at sites other than the
standard EcoRI substrate.

<>

<1>Polisson, C.
<2>MscI, a type II restriction endonuclease from Micrococcus species which recognizes 5' TGGCCA 3' .
<3>Nucleic Acids Res.
<4>17
<5>5858
<6>1989
<7>None

<>

<1>Polisson, C.
<2>Type II restriction endonuclease obtainable from Pseudomonas alcaligenes and a process for producing the same.
<3>US Patent Office
<4>US 5098839
<5>
<6>1992
<7>*
The present invention provides a novel Type II restriction endonuclease obtainable from
Pseudomonas alcaligenes. The endonuclease known as PacI, recognizes the following nucleotide
sequence and has a cleavage point indicated by the arrows:   5'TTAAT^TAA3'
                                                             3'AAT^TAATT5'
Also described is a process for obtaining PacI from P. alcaligenes.


<>

<1>Polisson, C.
<2>A novel type II restriction endonuclease obtainable from Pseudomonas alcaligenes and a process for producing the same.
<3>European Patent Office
<4>EP 0456356 B
<5>
<6>1994
<7>*
The present invention provides a novel Type II restriction endonuclease obtainable from
Pseudomonas alcaligenes. The endonuclease known as PacI, recognizes the following nucleotide
sequence and has a cleavage point indicated by the arrows:

    5'-TTAAT TAA-3'
    3'-ATT TAATT-5'

Also described is a process for obtaining PacI from P. alcaligenes.


<>

<1>Polisson, C., Barsomian, J.M.
<2>Type II restriction endonuclease obtainable from Kluyvera ascorbata and a process for producing the same.
<3>US Patent Office
<4>US 5147794
<5>
<6>1992
<7>*
The present invention provides a novel restriction endonuclease obtainable from the
bacterium Kluyvera ascorbata, hereinafter referred to as "KasI", which endonuclease:

(1) recognizes the base sequence in a double-stranded DNA molecule as shown below,
       5' G^GCGCC 3'
       3' CCGCG^G 5'
    (wherein C and G represent cytosine and guanine, respectively),

(2) cleaves said sequence in the phosphodiester bonds between G and G as indicated
    with the vertical arrows; and

(3) cleaves double-stranded pUC, M13mp18, and lambda DNA in one position, PhiX172,
    and T7 DNA in two positions, and Adeno2 DNA at 20 positions, while not cleaving
    SV40 DNA. The present invention further provides a recombinant DNA encoding the
    KasI restriction endonuclease and modification methylase obtainable from K.
    ascorbata and methods for the production of the recombinant DNA encoding those
    enzymes. Methods for producing the KasI restriction endonuclease and modification
    methylase in substantially pure form are also provided.


<>

<1>Polisson, C., Morgan, R.D.
<2>EarI, a restriction endonuclease from Enterobacter aerogenes which recognizes 5'CTCTTC3'.
<3>Nucleic Acids Res.
<4>16
<5>9872
<6>1988
<7>EarI, a TypeII restriction endonuclease, has been isolated from Enterobacter aerogenes
(NEB#450).  EarI, an isoschizomer of Ksp632I, recognizes the six base non-palindromic sequence
5'CTCTTC3', and cleaves one nucleotide 3' of the 3' cytosine on one strand, and four
nucleotides 5' of the 5' guanine on the opposite strand, to generate a three base 5'
overhang.  The single cleavage site of EarI on SV40 DNA was mapped to approximately position
4450 by analysis against BglI, EcoRI and TaqI cleaved SV40 DNA (figure 1, lanes B-E).  The
sequence 5'CTCTTC3' occurs at position 4437.  The number and sizes of the fragments
generated by digestion with EarI on eight DNA molecules (119 sites) match the computer
predicted number and sizes of the fragments that would be generated by cleavage at the
sequence 5'CTCTTC3'.  EarI has the following number of recognition sites on these commonly
used DNAs:  pUC19, 3; pBR322, 2; phiX174, 2; M13mp18, 2; SV40, 1; Adeno2, 29; T7, 46; and
Lambda, 34 (fig. 1, lanes F-l).  From these data we conclude that EarI recognizes the sequence
5'CTCTTC3'.  The crude extract contained approximately 8,000 units EarI per gram of cells.
The cleavage site of EarI was determined by cleavage of a primed synthesis reaction.  Using
M13mp18 DNA as template with an appropriate primer, the four standard dideoxy DNA sequencing
reactions were performed and a fifth reaction containing no dideoxy terminations was extended
through the EarI site.  The fifth reaction was terminated by heat treatment.  EarI was added
to the fifth reaction.  The cleaved product resulted in a single band (fig. II, lane -) which
comigrates with the first 3'nucleotide outside of the recognition sequence.  The addition of
Klenow subsequent to EarI digestion resulted in a single band, three nucleotides longer,
comigrating with the fourth 3' nucleotide outside of the recognition sequence (fig. II, lane
+).  These results indicate that EarI cleaves one nucleotide 3' of the recognition sequence
on the 5'CTCTTC3' strand, and four nucleotides 5' of the 5' G on the opposite strand
sequence 5'GAAGAG3', generating a three base 5' overhang.
5'CTCTTCN3'
3'GAGAAGNNNN5'.

<>

<1>Polisson, C., Morgan, R.D.
<2>AseI, a restriction endonuclease from Aquaspirillum serpens which recognizes 5'ATTAAT3'.
<3>Nucleic Acids Res.
<4>16
<5>10365
<6>1988
<7>AseI and AseII, type II restriction endonucleases, have been isolated from
Aquaspirillum serpens (NEB#448).  AseI, an isoschizomer of VspI, recognizes the
six base palindromic sequence 5'ATTAAT3', and cleaves 3' of the 5' T, to
generate a two base 5' overhang, 5' AT^TAAT3'.  AseII is an isoschizomer of
NciI (data not shown).

<>

<1>Polisson, C., Morgan, R.D.
<2>BsrI, a unique restriction endonuclease from Bacillus stearothermophilus which recognizes 5'ACTGG3'.
<3>Nucleic Acids Res.
<4>16
<5>5205
<6>1988
<7>A new type II restriction endonuclease, BsrI, has been isolated from Bacillus
stearothermophilus (NEB#447).  BsrI recognizes the five base non-palindromic
sequence 5' ACTGG 3'.  It cleaves one nucleotide outside of the recognition
sequence on one strand, and within the recognition sequence on the opposite
strand, to generate a two base 3' overhang.

<>

<1>Polisson, C., Morgan, R.D.
<2>AciI, a unique restriction endonuclease from Arthrobacter citreus which recognizes 5' CCGC3'.
<3>Nucleic Acids Res.
<4>18
<5>5911
<6>1990
<7>None

<>

<1>Polisson, C., Morgan, R.D.
<2>DrdI, a unique restriction endonuclease from Deinococcus radiodurans which recognizes 5'GACN6GTC3'.
<3>Nucleic Acids Res.
<4>17
<5>3316
<6>1989
<7>None

<>

<1>Polisson, C., Robinson, D.
<2>ApoI, a unique restriction endonuclease from Arthrobacter protophormiae which recognizes 5' RAATTY-3'.
<3>Nucleic Acids Res.
<4>20
<5>2888
<6>1992
<7>ApoI, a novel type II restriction endonuclease, has been isolated from Arthrobacter
protophormiae (NEB#723). ApoI recognizes the six base palindromic sequence 5' R|AATTY 3',
and cleaves after the first base pair, as indicated by the arrow, to create a four base 5'
extension.

<>

<1>Polisson, C., Robinson, D., Lunnen, K.
<2>Novel Type II restriction endonuclease ApoI, obtainable from Arthrobacter protophormiae and a process for producing the same.
<3>US Patent Office
<4>US 5200337
<5>
<6>1993
<7>*
The present invention provides a novel type II restriction endonuclease obtainable from
Arthrobacter protophormiae. The endonuclease known as ApoI, recognizes the following
nucleotide sequence and has a cleavage point indicated by the arrows:   5' Pu^AATTPy 3'
                                                                        3' PyTTAA^Pu 5'
Also described is a process for obtaining ApoI from Arthrobacter protophormiae.


<>

<1>Polisson, C., Robinson, D., Lunnen, K.
<2>A Type II restriction endonuclease, ApoI, obtainable from Arthrobacter protophormiae and a process for producing the same.
<3>European Patent Office
<4>EP 0539160 B
<5>
<6>1994
<7>The present invention relates to a new Type II restriction endonuclease, ApoI, obtainable from
Arthrobacter protophormiae and for a process for producing the same.

<>

<1>Pollak, A.J., Reich, N.O.
<2>Proximal Recognition Sites Facilitate Intrasite Hopping by DNA Adenine Methyltransferase MECHANISTIC EXPLORATION OF EPIGENETIC GENE REGULATION.
<3>J. Biol. Chem.
<4>287
<5>22873-22881
<6>2012
<7>The methylation of adenine in palindromic 5'-GATC-3' sites by Escherichia coli Dam supports
diverse roles, including the essential
regulation of virulence genes in several human pathogens. As a result
of a unique hopping mechanism, Dam methylates both strands of the same
site prior to fully dissociating from the DNA, a process referred to as
intrasite processivity. The application of a DpnI restriction
endonuclease-based assay allowed the direct interrogation of this
mechanism with a variety of DNA substrates. Intrasite processivity is
disrupted when the DNA flanking a single GATC site is longer than 400
bp on either side. Interestingly, the introduction of a second GATC
site within this flanking DNA reinstates intrasite methylation of both
sites. Our results show that intrasite methylation occurs only when
GATC sites are clustered, as is found in gene segments both known and
postulated to undergo in vivo epigenetic regulation by Dam methylation.
We propose a model for intrasite methylation in which Dam bound to
flanking DNA is an obligate intermediate. Our results provide insights
into how intrasite processivity, which appears to be context-dependent,
may contribute to the diverse biological roles that are carried out by
Dam.

<>

<1>Polosina, Y.Y., Mui, J., Pitsikas, P., Cupples, C.G.
<2>The Escherichia coli Mismatch Repair Protein MutL Recruits the Vsr and MutH Endonucleases in Response to DNA Damage.
<3>J. Bacteriol.
<4>191
<5>4041-4043
<6>2009
<7>The activities of the Vsr and MutH endonucleases of Escherichia coli are stimulated by MutL.
The interaction of MutL with each enzyme is enhanced
in vivo by 2-aminopurine treatment and by inactivation of the mutY gene.
We hypothesize that MutL recruits the endonucleases to sites of DNA
damage.

<>

<1>Poly, F., Read, T., Tribble, D.R., Baqar, S., Lorenzo, M., Guerry, P.
<2>Genome sequence of a clinical isolate of Campylobacter jejuni from Thailand.
<3>Infect. Immun.
<4>75
<5>3425-3433
<6>2007
<7>Campylobacter jejuni CG8486, which belongs to the HS4 complex, was isolated from a patient
with inflammatory diarrhea in Thailand.  This strain caused a diarrheal disease in ferrets
comparable to that caused by C. jejuni strain 81-176, but it was much less invasive for
epithelial cells in vitro than 81-176.  Complete genome sequencing of CG8486 revealed a
1.65-Mb genome that was very similar to the other two published genomes of clinical isolates
of C. jejuni, the genomes of 81-176 and NCTC 11168, with a limited number of CG8486-specific
genes mapping outside the hypervariable carbohydrate biosynthesis loci.  These data suggest
that the genes required for induction of inflammatory diarrhea are among the genes shared by
CG8486 and 81-176 but that either major changes in the carbohydrate loci and/or more subtle
changes in other genes may modulate virulence.

<>

<1>Poly, F., Read, T.D., Chen, Y.H., Monteiro, M.A., Serichantalergs, O., Pootong, P., Bodhidatta, L., Mason, C.J., Rockabrand, D., Baqar, S., Porter, C.K., Tribble, D., Darsley, M., Guerry, P.
<2>Characterization of two Campylobacter jejuni strains for use in volunteer experimental-infection studies.
<3>Infect. Immun.
<4>76
<5>5655-5667
<6>2008
<7>The development of vaccines against Campylobacter jejuni would be facilitated by
the ability to perform phase II challenge studies. However, molecular mimicry of
the lipooligosaccharide (LOS) of most C. jejuni strains with human gangliosides
presents safety concerns about the development of Guillain-Barre syndrome.
Clinical isolates of C. jejuni that appeared to lack genes for the synthesis of
ganglioside mimics were identified by DNA probe analyses. Two clinical isolates
from Southeast Asia (strains BH-01-0142 and CG8421) were determined to express
the LOS type containing N-acetyl quinovosamine. No ganglioside structures were
observed to be present in the LOSs of these strains, and pyrosequence analyses of
the genomes of both strains confirmed the absence of genes involved in
ganglioside mimicry. The capsule polysaccharide (CPS) of BH-01-0142 was
determined to be composed of galactose (Gal), 6-deoxy-ido-heptose, and, in
smaller amounts, D-glycero-D-ido-heptose, and the CPS of CG8421 was observed to
contain Gal, 6-deoxy-altro-heptose, N-acetyl-glucosamine, and minor amounts of
6-deoxy-3-O-Me-altro-heptose. Both CPSs were shown to carry
O-methyl-phosphoramidate. The two genomes contained strain-specific zones, some
of which could be traced to a plasmid origin, and both contained a large
chromosomal insertion related to the CJEI3 element of C. jejuni RM1221. The
genomes of both strains shared a high degree of similarity to each other and,
with the exception of the capsule locus of CG8421, to the type strain of the HS3
serotype, TGH9011.

<>

<1>Poly, F., Threadgill, D., Stintzi, A.
<2>Identification of Campylobacter jejuni ATCC 43431-specific genes by whole microbial genome comparisons.
<3>J. Bacteriol.
<4>186
<5>4781-4795
<6>2004
<7>This study describes a novel approach to identify unique genomic DNA sequences from the
unsequenced strain C. jejuni ATCC 43431 by comparison with the sequenced strain C. jejuni NCTC
11168. A shotgun DNA microarray was constructed by arraying 9,600 individual DNA fragments
from a C. jejuni ATCC 43431 genomic library onto a glass slide. DNA fragments unique to C.
jejuni ATCC 43431 were identified by competitive hybridization to the array with genomic DNA
of C. jejuni NCTC 11168. The plasmids containing unique DNA fragments were sequenced, allowing
the identification of up to 130 complete and incomplete genes. Potential biological roles were
assigned to 66% of the unique open reading frames. The mean G+C content of these unique genes
(26%) differs significantly from the G+C content of the entire C. jejuni genome (30.6%). This
suggests that they may have been acquired through horizontal gene transfer from an organism
with a G+C content lower than that of C. jejuni. Because the two C. jejuni strains differ by
Penner serotype, a large proportion of the unique ATCC 43431 genes encode proteins involved in
lipooligosaccharide and capsular biosynthesis, as expected. Several unique open reading frames
encode enzymes which may contribute to genetic variability, i.e., restriction-modification
systems and integrases. Interestingly, many of the unique C. jejuni ATCC 43431 genes show
identity with a possible pathogenicity island from Helicobacter hepaticus and components of a
potential type IV secretion system. In conclusion, this study provides a valuable resource to
further investigate Campylobacter diversity and pathogenesis.

<>

<1>Poly, F., Threadgill, D., Stintzi, A.
<2>Genomic diversity in Campylobacter jejuni: Identification of C-jejuni 81-176-specific genes.
<3>J. Clin. Microbiol.
<4>43
<5>2330-2338
<6>2005
<7>Since the publication of the complete genomic sequence of Campylobacter jejuni NCTC 11168 in
February 2000, evidence has been compiling that
suggests C. jejuni strains exhibit high genomic diversity. In order to
investigate this diversity, the unique genomic DNA sequences from a
nonsequenced Campylobacter strain, C. jejuni 81-176, were identified by
comparison with C. jejuni NCTC 11168 by using a shotgun DNA microarray
approach. Up to 63 kb of new chromosomal DNA sequences unique to this
pathogen were obtained. Eighty-six open reading frames were identified
by the presence of uninterrupted coding regions encoding a minimum of
40 amino acids. In addition, this study shows that the whole-plasmid
shotgun microarray approach is effective and provides a comprehensive
coverage of DNA regions that differ between two closely related
genomes. The two plasmids harbored by this Campylobacter strain, pTet
and pVir, were also sequenced, with coverages of 2.5- and 2.9-fold,
respectively, representing 72 and 92% of their complete nucleotide
sequences. The unique chromosomal genes encode proteins involved in
capsule and lipooligosaccharide biosynthesis, restriction and
modification systems, and respiratory metabolism. Several of these
unique genes are likely associated with C. jejuni 81-176 fitness and
virulence. Interestingly, the comparison of C. jejuni 81-176 unique
genes with those of C. jejuni ATCC 43431 revealed a single gene which
encodes a probable TraG-like protein. The product of this gene might be
associated with the mechanism of C. jejuni invasion into epithelial
cells. In conclusion, this study extends the repertoire of C. jejuni
genes and thus will permit the construction of a composite and more
comprehensive microarray of C. jejuni.

<>

<1>Polyachenko, V.M., Melnikova, V.A., Gruber, I.M., Kiuduliene, L.J., Raskin, B.M., Smirnova, G.A.
<2>Nutrient medium producing restriction endonuclease(s) from Haemophilus-type bacteria, containing fermented animal blood and acid casein hydrolysate, yeast extract, sodium chloride, etc.
<3>Soviet Patent Office
<4>SU 1479507
<5>
<6>1989
<7>Use of fermented animal blood hydrolysate (I) and acid casein hydrolysate (II) as the N
source, and edible yeast extract (III) as the source of vitamins in the nutrient medium used
for producing restriction endonucleases from the Haemophilus-type bacteria, improves its
properties.  The mixture contains (in wt. %): (I) 0.35-0.45, (II) 0.35-0.45, (III) 0.1-0.2 NAD
0.0002, haemin 0.001, NaCl 0.1-0.15 and balance distilled water.  Tests show that although
the yield of biomass is roughly the same as that using the known medium, the yield of ferment
is increased by more than 10 times.

<>

<1>Polyachenko, V.M., Melnikova, V.A., Gruber, I.M., Smirnova, G.A., Raskin, B.M.
<2>Development of a culture medium for bacteria of the genus Haemophilus, producing restriction endonucleases.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>3
<5>3-8
<6>1989
<7>A culture medium for the cultivation of hemophilic bacteria, containing acidic
casein hydrolysate, aminopeptide and fodder yeast extract, has been proposed.
The growth-stimulating properties of this medium have been studied on 5 strains
producing restrictases differing in their specificity.  In growing these
producer strains in a model AHKYM-2 fermenter with the supply of carbohydrate
substrates (glucose, sucrose, glycerin) the yield of biomass, considered to be
high for hemophilic bacteria (10-14 g wet weight from 1 liter of the medium),
has been achieved.  As shown on H. influenzae Rc B-2297 used as an example, an
increase in the yield of microbial biomass leads to a decrease in restrictase
specific activity.

<>

<1>Polyanovsky, O.L., Nosikov, V.V., Zhuze, A.L., Braga, E.A., Karlyshev, A.V.
<2>Regulation of restriction endonuclease activity with antibiotics.
<3>Adv. Enzyme Regul.
<4>17
<5>307-321
<6>1979
<7>Restriction-modification systems were made known in the early 1960's.  Active investigation,
however, has begun in the last years.  These systems serve to protect the bacterial cell from
invasion by foreign DNA.  Simultaneously, restriction enzymes are widely used as a tool for
specific DNA cleavage and obtaining of recombinant DNA molecules in vitro.

<>

<1>Pompejus, M., Kroeger, B., Schroder, H., Zelder, O., Haberhauer, G.
<2>Corynebacterium glutamicum genes encoding novel proteins.
<3>US Patent Office
<4>US 6962989 A
<5>
<6>2005
<7>Isolated nucleic acid molecules, designated MCP nucleic acid molecules, which encode novel MCP
proteins from Corynebacterium glutamicum are described.  The invention also provides antisense
nucleic acid molecules, recombinant expression vectors containing MCP nucleic acid molecules,
and host cells into which the expression vectors have been introduced.  The invention still
further provides isolated MCP proteins, mutated MCP proteins, fusion proteins, antigenic
peptides and methods for the improvement of production of a desired compound from C.
glutamicum based on genetic engineering of MCP genes in this organism.

<>

<1>Pompejus, M., Kroeger, B., Schroeder, H., Zelder, O., Haberhauer, G.
<2>Corynebacterium glutamicum genes encoding proteins involved in homeostasis and adaptation.
<3>International Patent Office
<4>WO 0100842 A
<5>
<6>2001
<7>Isolated nucleic acid molecules, designated HA nucleic acid molecules, which encode novel HA
proteins from Corynebacterium glutamicum are described.  The invention also provides antisense
nucleic acid molecules, recombinant expression vectors containing HA nucleic acid molecules,
and host cells into which the expression vectors have been introduced.  The invention still
further provides isolated HA proteins, mutated HA proteins, fusion proteins, antigenic
peptides and methods for the improvement of production of a desired compound from C.
glutamicum based on genetic engineering of HA genes in this organism.

<>

<1>Pompejus, M., Kroeger, B., Schroeder, H., Zelder, O., Haberhauer, G.
<2>Putative gene coding for Catechol 1,2-Dioxygenase from Corynebacterium glutamicum.
<3>European Patent Office
<4>EP 1683859 A
<5>
<6>2006
<7>Isolated nucleic acid molecules, designated HA nucleic acid molecules, which encode novel HA
proteins from Corynebacterium glutamicum are described.  The invention also provides antisense
nucleic acid molecules.  Recombinant expression vectors containing HA nucleic acid molecules,
and host cells into which the expression vectors have been introduced.  The invention still
further provides isolated HA proteins, mutated HA proteins, fusion proteins, antigenic
peptides and methods for the improvement of production of a desired compound from C.
glutamicum based on genetic engineering of HA genes in this organism.

<>

<1>Pompejus, M., Kroeger, B., Schroeder, H., Zelder, O., Haberhauer, G.
<2>Corynebacterium glutamicum genes encoding proteins involved in homeostasis and adaptation.
<3>Korean Patent Office
<4>KR 1020017016584 A
<5>
<6>2001
<7>
<>

<1>Pompejus, M., Kroeger, B., Schroeder, H., Zelder, O., Haberhauer, G.
<2>CORYNEBACTERIUM GLUTAMICUM GENES ENCODING PROTEINSINVOLVED IN HOMEOSTASIS AND ADAPTATION.
<3>Korean Patent Office
<4>KR 1020067022695 A
<5>
<6>2006
<7>
<>

<1>Ponger, L., Li, W.H.
<2>Evolutionary diversification of DNA methyltransferases in eukaryotic Genomes.
<3>Mol. Biol. Evol.
<4>22
<5>1119-1128
<6>2005
<7>In eukaryotes, C5-cytosine methylation is a common mechanism associated with a variety of
functions such as gene regulation or control of
genomic stability. Different subfamilies of eukaryotic
methyltransferases (MTases) have been identified, mainly in metazoa,
plants, and fungi. In this paper, we used hidden Markov models to
detect MTases in completed or almost completed eukaryotic genomes,
including different species of Protozoa. A phylogenetic analysis of
MTases enabled us to define six subfamilies of MTases, including two
new subfamilies. The dnmt1 subfamily that includes all the known MTases
with a maintenance activity seems to be absent in the Protozoa. The
dnmt2 subfamily seems to be the most widespread, being present even in
the nonmethylated Dictyostelium discoideum. We also found two dnmt2
members in the bacterial genus Geobacter, suggesting that horizontal
transfers of MTases occurred between eukaryotes and prokaryotes. Even
if the direction of transfer cannot be determined, this relationship
might be useful for understanding the function of this enigmatic
subfamily of MTases. Globally, our analysis reveals a great diversity
of MTases in eukaryotes, suggesting the existence of different
methylation systems. Our results also suggest acquisitions and losses
of different MTases in every eukaryotic lineage studied and that some
eukaryotes appear to be devoid of methylation.

<>

<1>Pope, W.H. et al.
<2>Expanding the diversity of mycobacteriophages: insights into genome architecture and evolution.
<3>PLoS ONE
<4>6
<5>E16329
<6>2011
<7>Mycobacteriophages are viruses that infect mycobacterial hosts such as
Mycobacterium smegmatis and Mycobacterium tuberculosis. All
mycobacteriophages characterized to date are dsDNA tailed phages, and have
either siphoviral or myoviral morphotypes. However, their genetic
diversity is considerable, and although sixty-two genomes have been
sequenced and comparatively analyzed, these likely represent only a small
portion of the diversity of the mycobacteriophage population at large.
Here we report the isolation, sequencing and comparative genomic analysis
of 18 new mycobacteriophages isolated from geographically distinct
locations within the United States. Although no clear correlation between
location and genome type can be discerned, these genomes expand our
knowledge of mycobacteriophage diversity and enhance our understanding of
the roles of mobile elements in viral evolution. Expansion of the number
of mycobacteriophages grouped within Cluster A provides insights into the
basis of immune specificity in these temperate phages, and we also
describe a novel example of apparent immunity theft. The isolation and
genomic analysis of bacteriophages by freshman college students provides
an example of an authentic research experience for novice scientists.

<>

<1>Popovski, B.
<2>Structure and mechanisms of action of restrictional endonucleases.
<3>Vet. Med. Nauki
<4>23
<5>13-17
<6>1986
<7>The structure and the mode of restrictional endonucleases are dealt with in
detail.  Described are some more important representatives of the three types
of endonucleases.  Stated are their role and place in present-day molecular
biology.

<>

<1>Popovski, B.
<2>Phenotypic characteristics of a new modificational and restriction system in Escherichia coli.
<3>Vet. Med. Nauki
<4>24
<5>19-24
<6>1987
<7>It has been demonstrated phenotypically that there exists a new
modification-and-restrictional system as synthesized by the recombinant
Escherichia coli tF strain.  A series of passages of the T3 and T7 phages and
their mutants in E. coli tF has made it possible to ascertain a specific
modification of the phage DNA, which was shown to be induced by the host
strain.  The high level of adsorption of these phages on the cell surface of E.
coli tF has ruled out the possibility of the existance of a nonclassical
modification and restriction of DNA.  In view of the further characterizing of
this modificational-and-restrictional system of E. coli tF it has been
comparatively studied with the already known modification-and-restrictional
systems isolated from various Escherichia coli strains.  Results have shown
that no identity exists between the tested systems and the one in E. coli tF.
It is stated that the new modificational-and-restrictional system of E. coli tF
belongs to none of the three known types of restrictional endonucleases.

<>

<1>Popovski, B.
<2>The location of genes coding the specific modification and restriction system in the recombinant E. coli strain tF.
<3>Vet. Med. Nauki
<4>24
<5>9-12
<6>1987
<7>The recombinant Escherichia coli tF strain has been shown to have its own specific
modification and restriction system.  In order to establish the location of genes, coding this
system a plasmid has been isolated from the investigated strain, which substantiates the
resistance to streptomycin.  The plasmid DNA has been transformed into a recipient strain, E.
coli O, which has no modification and restriction system of its own.  The newly obtained E.
coli O (ptF) transformants also have proved negative with regard to their testing for the
presence of a specific modification and restriction system.  The conclusion follows that the
genes, coding the modification and restriction system of the E. coli tF strain are not located
in the plasmid isolated from it.

<>

<1>Porcellato, D., Ostlie, H.M., Skeie, S.B.
<2>Draft Genome Sequence of Enterococcus hirae Strain INF E1 Isolated from Cultured  Milk.
<3>Genome Announcements
<4>2
<5>e00498-14
<6>2014
<7>Here, we present the draft genome of Enterococcus hirae INF E1, found as a contaminant in
cultured milk and studied for its ability to metabolize milk fat
globule membrane glycoconjugates.

<>

<1>Pore, S.D., Arora, P., Dhakephalkar, P.K.
<2>Draft Genome Sequence of Geobacillus sp. Strain FW23, Isolated from a Formation Water Sample.
<3>Genome Announcements
<4>2
<5>e00352-14
<6>2014
<7>The thermophilic Geobacillus sp. strain FW23 was isolated from the Mehsana oil wells in
Gujrat, India, during a screening for oil-degrading bacteria. Here, we
report the draft genome sequence of Geobacillus sp. FW23, which may help reveal
the genomic differences between this strain and the earlier reported species of
the genus Geobacillus.

<>

<1>Port, G.C., Paluscio, E., Caparon, M.G.
<2>Complete Genome Sequence of emm Type 14 Streptococcus pyogenes Strain HSC5.
<3>Genome Announcements
<4>1
<5>e00612-13
<6>2013
<7>Streptococcus pyogenes causes a greater diversity of human disease than any other bacterial
pathogen. Here, we present the complete genome sequence of the emm type
14 S. pyogenes strain HSC5. This strain is a robust producer of the cysteine
protease SpeB and is capable of producing infection in several different animal
models.

<>

<1>Porter, S.L., Wilkinson, D.A., Byles, E.D., Wadhams, G.H., Taylor, S., Saunders, N.J., Armitage, J.P.
<2>Genome Sequence of Rhodobacter sphaeroides strain WS8N.
<3>J. Bacteriol.
<4>193
<5>4027-4028
<6>2011
<7>R. sphaeroides is a metabolically diverse photosynthetic alpha-proteobacterium found
ubiquitously in soil and in fresh water
habitats. Here we present the annotated genome sequence of Rhodobacter
sphaeroides WS8N.

<>

<1>Portillo, F.G.D., Pucciarelli, M.G., Casadesus, J.
<2>DNA adenine methylase mutants of Salmonella typhimurium show defects in protein secretion, cell invasion, and M cell cytotoxicity.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>11578-11583
<6>1999
<7>Mutants of Salmonella typhimurium lacking DNA adenine methylase are attenuated for virulence
in BALB/c mice.  LD(50) values of a DNA adenine methylation (Dam)(-) mutant are at least
10(3)- to 10(4)-fold higher than those of the parental strain when administrated by oral or
intraperitoneal routes. Dam(-) mutants are unable to proliferate in target organs but persist
in low numbers in these locations. Efficient protection to challenge with the virulent
parental strain is observed in mice infected with a Dam(-) mutant. Use of the ileal loop assay
shows that Dam(-) mutants are less cytotoxic to M cells and fail to invade enterocytes. In the
tissue culture model, lack of DNA adenine methylation causes reduced ability to invade
nonphagocytic cells. In contrast, no effect is observed either in intracellular proliferation
within nonphagocytic cells or in survival within macrophages. The invasion defect of Dam(-)
mutants is correlated with a distinct pattern of secreted proteins, which is observed in both
PhoP(+) and PhoP(-) backgrounds. Altogether, our observations suggest a multifactorial role of
Dam methylation in Salmonella virulence.

<>

<1>Posey, K.L., Gimble, F.S.
<2>Insertion of a reversible redox switch into a rare-cutting DNA endonuclease.
<3>Biochemistry
<4>41
<5>2184-2190
<6>2002
<7>Target sites for homing endonucleases occur infrequently in complex genomes. As a consequence,
these enzymes can be used in mammalian systems to introduce double-strand breaks at
recognition sites inserted within defined loci to study DNA repair by homologous and
nonhomologous recombination. Using homing endonucleases for gene targeting in vivo would be
more feasible if temporal or spatial regulation of their enzymatic activity were possible.
Here, we show that the DNA cleavage activity of the yeast PI-SceI homing endonuclease can be
turned on and off using a redox switch. Two cysteine pairs (Cys-64/Cys-344 and Cys-67/Cys-365)
were separately inserted into flexible DNA binding loop(s) to create disulfide bonds that lock
the endonuclease into a nonproductive conformation. The cleavage activities of the reduced
Cys-64/Cys-344 and Cys-67/Cys-365 variants are similar or slightly lower than that of the
control protein, but the activities of the proteins in the oxidized state are decreased more
than 30-fold. Modulating the activity of the proteins is easily accomplished by adding or
removing the reducing agent. We show that defects in DNA binding account for the decreased DNA
cleavage activities of the proteins containing disulfide bonds. Interestingly, the
Cys-67/Cys-365 variant toggles between two different DNA binding conformations under reducing
and oxidizing conditions, which may permit the identification of structural differences
between the two states. These studies demonstrate that homing endonuclease activity can be
controlled using a molecular switch.

<>

<1>Posey, K.L., Koufopanou, V., Burt, A., Gimble, F.S.
<2>Evolution of divergent DNA recognition specificities in VDE homing endonucleases from two yeast species.
<3>Nucleic Acids Res.
<4>32
<5>3947-3956
<6>2004
<7>Homing endonuclease genes (HEGs) are mobile DNA elements that are thought to confer no benefit
to their host. They encode site-specific DNA endonucleases that perpetuate the element within
a species population by homing and disseminate it between species by horizontal transfer.
Several yeast species contain the VMA1 HEG that encodes the intein-associated VMA1-derived
endonuclease (VDE). The evolutionary state of VDEs from 12 species was assessed by assaying
their endonuclease activities. Only two enzymes are active, PI-ZbaI from Zygosaccharomyces
bailii and PI-ScaI from Saccharomyces cariocanus. PI-ZbaI cleaves the Z.bailii recognition
sequence significantly faster than the Saccharomyces cerevisiae site, which differs at six
nucleotide positions. A mutational analysis indicates that PI-ZbaI cleaves the S.cerevisiae
substrate poorly due to the absence of a contact that is analogous to one made in PI-SceI
between Gln-55 and nucleotides +9/+10. PI-ZbaI cleaves the Z.bailii substrate primarily due to
a single base-pair substitution (A/T+5  T/A+5). Structural modeling of the PI-ZbaI/DNA complex
suggests that Arg-331, which is absent in PI-SceI, contacts T/A+5, and the reduced activity
observed in a PI-ZbaI R331A mutant provides evidence for this interaction. These data
illustrate that homing endonucleases evolve altered specificity as they adapt to recognize
alternative target sites.

<>

<1>Posfai, G., Baldauf, F., Erdei, S., Posfai, J., Venetianer, P., Kiss, A.
<2>Structure of the gene coding for the sequence-specific DNA-methyltransferase of the B. subtilis phage SPR.
<3>Nucleic Acids Res.
<4>12
<5>9039-9049
<6>1984
<7>The nucleotide sequence of the gene coding for the 5'GGCC and 5'CCGG specific
DNA methyltransferase of the Bacillus subtilis phage SPR was determined by the
Maxam-Gilbert procedure.  Transcriptional and translational signals of the
sequence were assigned with the help of S1 mapping and translation in E. coli
minicells.  The gene codes for a 49 kd polypeptide.  The amino acid sequence of
the SPR methylase shows regions of homology with the sequence of the
5'-GGCC-specific BspRI modification methylase.

<>

<1>Posfai, G., Kim, S.C., Szilak, L., Kovacs, A., Venetianer, P.
<2>Complementation by detached parts of GGCC-specific DNA methyltransferases.
<3>Nucleic Acids Res.
<4>19
<5>4843-4847
<6>1991
<7>Individually inactive N- and C-terminal fragments of the m5C-methyltransferase
M.BspRI can complement each other resulting in specific, in vivo methylation of
the DNA.  This was shown by cloning the coding regions for N- and C-terminal
parts of the enzyme in compatible plasmids and co-transforming them into E.
coli cells.  The enzyme could be detached at several different sites, producing
either non-overlapping or partially overlapping fragments capable of
complementation.  Reconstitution of the active methyltransferase from inactive
fragments was demonstrated in vitro, as well.  Another GGCC-specific
methyltransferase, M.BsuRI, showed a similar complementation phenomenon.
Moreover, interspecies complmentation was observed between appropriate
fragments of the two closely related enzymes M.BspRI and M.BsuRI.  Fragments of
structurally and functionally more different methyltransferases were unable to
complement each other.

<>

<1>Posfai, G., Kiss, A., Erdei, S., Posfai, J., Venetianer, P.
<2>Structure of the Bacillus sphaericus R modification methylase gene.
<3>J. Mol. Biol.
<4>170
<5>597-610
<6>1983
<7>A 2500 base-pair segment of Bacillus sphaericus R DNA cloned in Escherichia
coli has previously been shown to carry the functional BspRI modification
methylase gene.  The approximate location of the gene on this DNA segment and
its direction of transcription were established by subcloning experiments.  The
nucleotide sequence of the relevant region was determined by the Maxam-Gilbert
procedure.  An open reading frame that can code for a 424 amino acid protein
was found.  The calculated molecular weight (48,264) of this protein is in fair
agreement with previous estimates (50,000 to 52,000).  The synthesis of this
protein was demonstrated in E. coli minicells.  The initiation point of
transcription by E. coli RNA polymerase was localized by in vitro transcription
experiments.  The open reading frame starts 29 base-pairs downstream from the
transcription initiation site and it is preceded by a sequence showing
extensive Shine-Dalgarno complementarity.  Subcloning experiments and
translation in minicells suggest that after removal of this translational
initiation site, a secondary start site 29 amino acids downstream can also
start translation in E. coli and this shorter protein retains the methylase
activity.  The overall base composition of the gene and the codon usage
indicate a strong preference for A.T base-pairs.

<>

<1>Posfai, G., Kiss, A., Venetianer, P.
<2>Overproduction of the Bacillus sphaericus R modification methylase in Escherichia coli and its purification to homogeneity.
<3>Gene
<4>50
<5>63-67
<6>1986
<7>A DNA fragment containing the information coding for the GGCC-specific Bacillus
sphaericus R modification methylase, BspR, was inserted into plasmid vector
pKK223-3 under the control of the strong and inducible tac promoter, and
transformed into Escherichia coli HB101.  Upon induction this strain
accumulated the methylase enzyme (while cell growth was inhibited) up to
several percent of total cellular protein.  Homogeneous methylase could be
prepared in three purification steps.

<>

<1>Posfai, G., Szybalski, W.
<2>Increasing the FokI cleavage specificity from 5 to 7 base pairs by two-step methylation.
<3>Nucleic Acids Res.
<4>16
<5>6245
<6>1988
<7>Non-cognate methylation of specific nucleotides of the recognition sequence
could inhibit the methylase but not the endonuclease of the same
restriction-modification system.  This permits an increase in the cleavage
specificity of the restriction enzyme by two-step methylation of the DNA, as
shown here for the three-component M.MspI-M.FokI-FokI combination, changing
FokI specificity from 5 to 7 bp.  The cleavage specificity of FokI is
5'-GGATG(N)9/13.  M.FokI methylates only one strand of the recognition site
resulting in GGmATG sequence.  MspI and FokI sites can overlap in the 7-bp
sequence CCGGATG or in the 8-bp sequence CCGGGATG.  Methylation of such sites
by M.MspI (methylation specificity:  mCCGG) produces non-cognate methylation of
the overlapping FokI sites, resulting in  (a)G GATG    CmCTAC or (b)GGATG
mCCTAC sequences, respectively.  We show here that in case (a), M.FokI is
inhibited and FokI is unaffected (Fig. 2, lanes 3 and 2, respectively), while
in case (b), M.FokI is unaffected and FokI is partially inhibited (Fig. 2,
lanes 6 and 5, respectively, where M.HpaII was used to produce the (b) type
non-cognate methylation of FokI sites on the same 7-bp sequences).  As a
result, when the DNA is premethylated by M.MspI, M.FokI cannot methylate those
7-bp CCGGATG sequences (which thus remain susceptible to FokI), whereas all
other FokI sites would be methylated and thus protected from FokI cleavage.
However, cleavage by FokI at the CCGGATG sites may not be complete (max 90%),
because M.FokI has some residual affinity to the sites premethylated by M.MspI,
especially when excess M.FokI is used (data not shown).  Nevertheless, in many
cases this does not affect the applicability of the system.

<>

<1>Posfai, G., Szybalski, W.
<2>A simple method for locating methylated bases in DNA, as applied to detect asymmetric methylation by M.FokIA.
<3>Gene
<4>69
<5>147-151
<6>1988
<7>Class-IIS restriction enzymes, which cut the DNA outside their recognition sequence, could be
used for locating the bases methylated by a DNA-modification methylase.  This is possible
because methylation of the class-IIS cut sites does not interfere with the cleavage.  The
method consists of (i) selection of a nucleotide sequence with appropriate overlap between the
methylase recognition site and the class-IIS enzyme cut site, (ii) methylation using
S-adenosylmethionine as [3H]methyl donor, (iii) cleavage of the methylated sequence with the
class-IIS enzyme, (iv) separation of the cleavage products and identification of the
3H-labelled fragment.  Using this simple and straightforward method, we have shown that
M.FokIA is an adenine methylase and methylates asymmetrically one strand of the FokI
recognition site, resulting in the
5'-GGmATG(N)9
CC TAC(N)13
sequence.  In addition, it was observed that another class-IIS restriction enzyme, SfaNI, is
completely inhibited by methylation of its recognition site,  5'-GCA TC/CGTmAG, by M.FokIA.

<>

<1>Posfai, G., Szybalski, W.
<2>A simple method for locating methylated bases in DNA using class-IIS restriction enzymes.
<3>Gene
<4>74
<5>179-181
<6>1988
<7>Meeting Abstract

<>

<1>Posfai, J., Bhagwat, A.S., Posfai, G., Roberts, R.J.
<2>Predictive motifs derived from cytosine methyltransferases.
<3>Nucleic Acids Res.
<4>17
<5>2421-2435
<6>1989
<7>Thirteen bacterial DNA methyltransferases that catalyze the formation of
5-methylcytosine within specific DNA sequences possess related structures.
Similar building blocks (motifs), containing invariant positions, can be found
in the same order in all thirteen sequences.  Five of these blocks are highly
conserved while a further five contain weaker similarities.  One block, which
has the most invariant residues, contains the proline-cysteine dipeptide of the
proposed catalytic site.  A region in the second half of each sequence is
unusually variable both in length and sequence composition.  Those
methyltransferases that exhibit significant homology in this region share
common specificity in DNA recognition.  The five highly conserved motifs can be
used to discriminate the known 5-methylcytosine forming methyltransferases from
all other methyltransferases of known sequence, and from all other identified
proteins in the PIR, GenBank and EMBL databases.  These five motifs occur in a
mammalian methyltransferase responsible for the formation of 5-methylcytosine
within CG dinucleotides.  By searching the unidentified open reading frames
present in the GenBank and EMBL databases, two potential 5-methylcytosine
forming methyltransferases have been found.

<>

<1>Posfai, J., Bhagwat, A.S., Roberts, R.J.
<2>Sequence motifs specific for cytosine methyltransferases.
<3>Gene
<4>74
<5>261-265
<6>1988
<7>Using a new alignment method, the sequences of 13 m5C methyltransferases
(MTases) have been examined.  Five extremely well-conserved blocks of sequence
have been detected and have been used as fixed points for the alignment of the
13 sequences.  Following this initial alignment, five further blocks of
similarity have been identified to give a total of ten recognizable blocks of
sequence homology that are all arranged in a common order.  The structures of
these MTases consist of a variable-length N-terminal arm followed by eight
well-conserved blocks each separated by small variable-length regions.  A large
variable-length segment of 90 to 270 amino acids (aa) then follows.  After this
are two blocks, and a variable-length C-terminal segment completes the
sequence.  Within the final alignment, 20 aa in the protein sequences, and 86
nucleotides in the nucleotide sequences are invariant.  The strongest
conservation is found in proximity to a suspected functional site that contains
the dipeptide proline-cysteine.  Consensus patterns can be defined for the five
best conserved blocks and, when used as search motifs, are able to clearly
distinguish between the m5C MTases and all other identified proteins in the PIR
database.  This suggests they may be of use in identifying putative MTases
among protein sequences of unknown function.

<>

<1>Posfai, J., Roberts, R.J.
<2>Predictive motifs of cytosine methylases.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>213
<6>1989
<7>Fourteen bacterial DNA methyltransferases that catalyze the formation of
5-methylcytosine within specific DNA sequences possess related structures.
Similar building blocks (motifs), containing invariant positions, can be found
in the same order in all fourteen sequences.  Five of these blocks are highly
conserved while a further five contain weaker similarities.  One block which
has the most invariant residues, contains the proline-cysteine dipeptide of the
proposed catalytic site.  A region in the second half of each sequence is
unusually variable both in length and sequence composition.  Those
methyltransferases that exhibit significant homology in this region share
common specificity in DNA recognition.  The five highly conserved motifs can be
used to discriminate the known 5-methylcytosine forming methyltransferases from
all other methyltransferases of known sequence and from all other identified
proteins in the PIR, GenBank and EMBL databases.  These five motifs occur in
the eukaryotic mammalian methyltransferase.  By searching the unidentified open
reading frames present in the GenBank and EMBL databases two potential cytosine
methyltransferases have been found that were not recognized previously.

<>

<1>Potter, A.A., Loutit, J.S.
<2>Exonuclease activity from Pseudomonas aeruginosa which is missing in phenotypically restrictionless mutants.
<3>J. Bacteriol.
<4>151
<5>1204-1209
<6>1982
<7>A phenotypically restrictionless strain of Pseudomonas aeruginosa was found to
lack a deoxyribonuclease specific for linear duplex DNA.  The purified enzyme
had an optimum pH of 8.5, required MgCl2 (10 mM) for maximum activity, and did
not require ATP.  Neither the degradation of heat-denatured DNA nor the
degradation of bacteriophage F116 DNA was detected.  The genome of
bacteriophage F116 was shown to possess single-stranded terminal regions, which
account for the resistance to degradation and for the ability of the phage to
transfect restriction-proficient strains.

<>

<1>Potter, B.V.L., Eckstein, F.
<2>Cleavage of phosphorothioate-substituted DNA by restriction endonucleases.
<3>J. Biol. Chem.
<4>259
<5>14243-14248
<6>1984
<7>M13 RF DNA was synthesized in vitro in the presence of various single
deoxynucleoside 5'-O-(1-thiotriphosphate) phosphorothioate analogues, and the
three other appropriate deoxynucleoside triphosphates using a M13
(+)-single-stranded template, Escherichia coli DNA polymerase I and T4 DNA
ligase.  The resulting DNAs contained various restriction endonuclease
recognition sequences which had been modified at their cleavage points in the
(-)-strand by phosphorothioate substitution.  The behavior of the restriction
enzymes AvaI, BamHI, EcoRI, HindIII, and SalI towards these substituted DNAs
was investigated.  EcoRI, BamHI, and HindIII were found to cleave appropriate
phosphorothioate-substituted DNA at a reduced rate compared to normal M13 RF
DNA, and by a two-step process in which all of the DNA is converted to an
isolable intermediate nicked molecule containing a specific discontinuity at
the respective recognition site presumably in the (+)-strand.  By contrast,
SalI cleaved substituted DNA effectively without the intermediacy of a nicked
form.  AvaI, however, is only capable of cleaving the unsubstituted (+)-strand
in appropriately modified DNA.

<>

<1>Potter, S.S., Bott, K.F., Newbold, J.E.
<2>Two-dimensional restriction analysis of the Bacillus subtilis genome: gene purification and ribosomal ribonucleic acid gene organization.
<3>J. Bacteriol.
<4>129
<5>492-500
<6>1977
<7>With two-dimensional restriction enzyme analysis we have been able to cleave
the Bacillus subtilis genome and resolve the resulting deoxyribonucleic acid
(DNA) segments into discrete bands on agarose gels.  A general procedure for
gene purification has been developed by coupling multidimensional restriction
analysis with a biological assay for gene detection.  The organization of
ribosomal ribonucleic acid (rRNA) genes was studied by hybridizing 16S and 23S
rRNA probes to the two-dimensional DNA banding patterns.

<>

<1>Pouillot, F., Fayolle, C., Carniel, E.
<2>A putative DNA adenine methyltransferase is involved in Yersinia pseudo tuberculosis pathogenicity.
<3>Microbiology
<4>153
<5>2426-2434
<6>2007
<7>Some adenine methyltransferases have been shown not only to protect specific DNA restriction
sites from cleavage by a restriction
endonuclease, but also to play a role in various bacterial processes
and sometimes in bacterial virulence. This study focused on a type I
restriction-modification system (designated yrml) of Y.
pseudotuberculosis. This system is composed of three adjacent genes
which could potentially encode an N-6-adenine DNA methylase (YamA), an
enzyme involved in site-specific recognition (YrsA) and a restriction
endonuclease (YreA). Screening of 85 isolates of Y. pestis and Y.
pseudotuberculosis indicated that the yrml system has been lost by Y.
pestis and that yamA (but not yrsA or yreA) is present in all Y
pseudotuberculosis strains tested, suggesting that it may be important
at some stages of the epidemiological cycle of this species. To further
investigate the role of yamA in Y pseudo tuberculosis survival,
multiplication or virulence, a Delta yamA mutant of Y
pseudotuberculosis IP32953 was constructed by allelic exchange with a
kanamycin cassette. The fact that Delta yamA mutants were obtained
indicated that this gene is not essential for Y pseudotuberculosis
viability. The IP32953 Delta yamA mutant strain grew as well as the
wild-type in a rich medium at both 28 degrees C and 37 degrees C. It
also grew normally in a chemically defined medium at 28 degrees C, but
exhibited a growth defect at 37 degrees C. In contrast to the Dam
adenine methyltransferase, a mutation in yamA did not impair the
functions of DNA repair or resistance to detergents. However, the Delta
yamA mutant exhibited a virulence defect in a mouse model of
intragastric infection. The in silico analysis indicated that the
chromosomal region carrying the Y pseudotuberculosis yrml locus has
been replaced in Y. pestis by a horizontally acquired region which
potentially encodes another methyltransferase. YamA might thus be
dispensable for Y. pestis growth and virulence because this species has
acquired another gene fulfilling the same functions.

<>

<1>Poulter, R.T.M., Goodwin, T.J.D., Butler, M.I.
<2>The nuclear-encoded inteins of fungi.
<3>Fungal Genet. Biol.
<4>44
<5>153-179
<6>2007
<7>An intein is a protein sequence embedded within a precursor protein that is excised during
protein maturation. Inteins were first found
encoded in the VMA gene of Saccharomyces cerevisiae. Subsequently, they
have been found in diverse organisms (eukaryotes, archaea, eubacteria
and viruses). The VMA intein has been found in various saccharomycete
yeasts but not in other fungi. Different inteins have now been found
widely in the fungi (ascomycetes, basidiomycetes, zygomycetes and
chytrids) and in diverse proteins. A protein distantly related to
inteins, but closely related to metazoan hedgehog proteins, has been
described from Glomeromycota. Many of the newly described inteins
contain homing endonucleases and some of these are apparently active.
The enlarged fungal intein data set permits insight into the evolution
of inteins, including the role of horizontal transfer in their
persistence. The diverse fungal inteins provide a resource for
biotechnology using their protein splicing or homing endonuclease
capabilities.

<>

<1>Povilionis, P.I., Lubys, A.A., Janulaitis, A.
<2>Cloning of the restriction-modification genes of Bacillus centrosporus in Escherichia coli.
<3>Genetika
<4>24
<5>210-215
<6>1988
<7>Using the pBR327 vector we have produced a genomic library of Bacillus
centrosporus.  The total plasmid DNA of the library was cleaved by the
restriction endonuclease BcnI and then transformed in Escherichia coli RR1.
Among the transformants obtained we identified two clones possessing
restriction and DNA modification profiles of BcnI and containing 13.3 kb and
9.05 kb plasmids, respectively.  Restriction mapping of both plasmids showed
that they contained two sites for HindIII, one site for Eco31I, and one site
for Eco47III located at equal distances.  Deletion mapping of the recombinant
plasmids confirmed that this was the region of the location of the BcnI
restriction-modification genes.  On the basis of the results obtained we
discuss the special features of cloning of the restriction-modification genes.

<>

<1>Povilionis, P.I., Lubys, A.A., Vaisvila, R.I., Kulakauskas, S.T., Janulaitis, A.
<2>Investigation of methyl-cytosine specific restriction in Escherichia coli K-12.
<3>Genetika
<4>25
<5>753-755
<6>1989
<7>Experiments on transformation of Escherichia coli K-12 cells by plasmids
carrying RM systems with different recognition sites containing
5-methylcytosine have shown that the gene mcrB dtermines the function of
restriction.  The data obtained made it possible to believe that E. coli
possesses no restriction system recognizing specifically cytosine methylated in
position 4.

<>

<1>Powell, I.B., Davidson, B.E.
<2>Resistance to in vitro restriction of DNA from lactic streptococcal bacteriophage c6A.
<3>Appl. Environ. Microbiol.
<4>51
<5>1358-1360
<6>1986
<7>DNA isolated from streptococcal bacteriophage c6A was cut only infrequently by
many restriction endonucleases.  Fragments of c6A DNA cloned in Escherichia
coli plasmids were similarly resistant to cleavage.  We conclude that the low
frequency of cleavage is due to an unusually low number of restriction enzyme
recognition sequences in c6A DNA.

<>

<1>Powell, L.M., Connolly, B.A., Dryden, D.T.F.
<2>The DNA binding characteristics of the trimeric EcoKI methyltransferase and its partially assembled dimeric form determined by fluorescence polarization and DNA footprint.
<3>J. Mol. Biol.
<4>283
<5>947-961
<6>1998
<7>The type I DNA restriction and modification systems of enteric bacteria display several
enzymatic activities due to their oligomeric structure.  Partially assembled forms of the
EcoKI enzyme from E. coli K12 can display specific DNA binding properties and modification
methyltransferase activity.  The heterodimer of one specificity (S) subunit and one
modification (M) subunit can only bind DNA whereas  the addition of a second modification
subunit to form M2S1 also confers methyltransferase activity.  We have examined the DNA
binding specificity of M1S1 and M2S1 using the change in fluorescence anisotropy which occurs
on binding of a DNA probe labelled with a hexachlorofluorescein fluorophore.  The dimer has
much weaker affinity for the EcoKI target sequence than the trimer and slightly less ability
to discriminate against other DNA sequences.  Binding of both proteins is strongly dependent
on salt concentration.  The fluorescence results compare favorably with those obtained with
the gel retardation method.  DNA footprinting using exonucleaseIII and DNaseI, and methylation
interference show no asymmetry, with both DNA strands being protected by the dimer and the
trimer.  This indicates that the dimer is a mixture of the two possible forms, M1S1 and S1M1.
The dimer has a footprint on the DNA substrate of the same length as the trimer implying that
the modification subunits are located on either side of the DNA helical axis rather than lying
along the helical axis.

<>

<1>Powell, L.M., Dryden, D.T.F., Murray, N.E.
<2>Sequence-specific DNA binding by EcoKI, a type IA DNA restriction enzyme.
<3>J. Mol. Biol.
<4>283
<5>963-976
<6>1998
<7>The type I DNA restriction and modification enzymes of prokaryotes are multimeric enzymes that
cleave unmethylated, foreign DNA in a complex process involving recognition of the methylation
status of a DNA target sequence, extensive translocation of DNA in both directions towards the
enzyme bound at the target sequence, ATP hydrolysis, which is believed to drive the
translocation possibly via a helicase mechanism, and eventual endonucleolytic cleavage of the
DNA.  We have examined the DNA binding affinity and exonuclease III footprint of the EcoKI
type IA restriction enzyme on oligonucleotide duplexes that either contain or lack the target
sequence.  The influence of the cofactors, S-adenosyl methionine and ATP, on binding to DNA of
different methylation states has been assessed.  EcoKI in the absence of ATP, with or without
S-adenosyl methionine, binds tightly even to DNA lacking the target site and the exonuclease
footprint is large, approximately 45 base-pairs.  The protection is weaker on DNA lacking the
target site.  Partially assembled EcoKI lacking one or both of the subunits essential for DNA
cleavage, is unable to bind tightly to DNA lacking the target site but can bind tightly to the
recognition site.  The addition of ATP to EcoKI, in the presence of AdoMet, allows tight
binding only to the target site and the footprint shrinks to 30 base pairs, almost identical
to that of the modification enzyme which makes up the core of EcoKI.  The same effect occurs
when S-adenosyl homocysteine or sinefungin are substituted for S-adenosyl methionine, and ADP
or ATPgammaS are substituted for ATP.  It is proposed that the DNA binding surface of EcoKI
comprises three regions: a "core" region which recognizes the target sequence and which is
present on the modification enzyme, and a region on each DNA cleavage subunit.  The cleavage
subunits make tight contacts to any DNA molecule in the absence of cofactors, but this contact
is weakened in the presence of cofactors to allow the protein conformational changes required
for DNA translocation when a target site is recognized by the core modification enzyme.  This
weakening of the interaction between the DNA cleavage subunits and the DNA could allow more
access of exonuclease III to DNA and account for the shorter footprint.

<>

<1>Powell, L.M., Dryden, D.T.F., Willcock, D.F., Pain, R.H., Murray, N.E.
<2>DNA recognition by the EcoK methyltransferase. The influence of DNA methylation and the cofactor S-adenosyl-L-methionine.
<3>J. Mol. Biol.
<4>234
<5>60-71
<6>1993
<7>The methyltransferase of the EcoK type I restriction/modification system is trimeric, M2S1,
where the S subunit determines the sequence specificity of the enzyme. The methyltransferase
has a strong preference for hemimethylated substrate DNA and therefore, we have investigated
the effect of the methylation state of DNA on binding by the enzyme, together with the effects
on binding of the cofactor S-adenosyl-L-methionine. Our results indicate that the
methyltransferase has two non-interacting S-adenosyl-L-methionine binding sites, each with a
dissociation constant of 3.60 (+-0.42)uM determined by equilibrium dialysis, or 2.21 (+-0.29)
uM determined by the displacement of a fluorescent probe. Ultraviolet light-induced
crosslinking showed that S-adenosyl-L-methionine binds strongly only to the modification (M)
subunits. Changes in the sedimentation velocity of the methyltransferases imply a protein
conformational change due to S-adenosyl-L-methionine binding. Gel retardation results show
that the binding of S-adenosyl-L-methionine to the methyltransferase enhances binding to both
specific and non-specific DNAs, but the enhancement is greater for the specific DNA.
Differences in binding affinities contribute to the recognition of the specific nucleotide
sequence AAC(N)6GTGC by the methyltransferase in preference to a non-specific sequence. In
contrast, although the complexes of unmodified and hemimethylated DNAs with the
methyltransferase have different mobilities in non-denaturing gels, there appears to be no
contribution of binding affinity to the distinction between these two substrates. Therefore,
the preference for the hemimethylated substrate must be due to a difference in catalysis.

<>

<1>Powell, L.M., Lejeune, E., Hussain, F.S., Cronshaw, A.D., Kelly, S.M., Price, N.C., Dryden, D.T.
<2>Assembly of EcoKI DNA methyltransferase requires the C-terminal region of the HsdM modification subunit.
<3>Biophys. Chem.
<4>103
<5>129-137
<6>2003
<7>The methyltransferase component of type I DNA restriction and modification systems comprises
three subunits, one DNA sequence specificity subunit and
two DNA modification subunits. Limited proteolysis of the EcoKI
methyltransferase shows that a 55-kDa N-terminal fragment of the 59-kDa
modification subunit is resistant to degradation. We have purified this
fragment and determined by mass spectrometry that proteolysis removes 43
or 44 amino acids from the C-terminus. The fragment fails to interact with
the other subunits even though it still possesses secondary and tertiary
structure and the ability to bind the S-adenosylmethionine cofactor. We
conclude that the C-terminal region of the modification subunit of EcoKI
is essential for the assembly of the EcoKI methyltransferase.

<>

<1>Powell, L.M., Murray, N.E.
<2>S-adenosyl methionine alters the DNA contacts of the EcoKI methyltransferase.
<3>Nucleic Acids Res.
<4>23
<5>967-974
<6>1995
<7>The EcoKI methyltransferase methylates two adenines on opposite strands of its bipartite DNA
recognition sequence AAC(N6)GTGC. The enzyme has a strong preference for hemimethylated DNA
substrates, but the methylation state of the DNA does not influence its binding affinity.
Methylation interference was used to compare the contacts made by the EcoKI methyltransferase
with unmodified, hemimethylated or fully modified DNAs. Contacts were seen at or near the N7
position of guanine, in the major groove, for all of the guanines in the EcoKI recognition
sequence, and at two guanines on the edge of the intervening spacer sequence. The presence of
the cofactor and methyl donor S-adenosyl methionine had a striking effect on the interference
pattern for unmodified DNA which could not be mimicked by the presence of the cofactor
analogue S-adenosyl homocysteine. In contrast, S-adenosyl methionine had no effect on the
interference patterns for either kind of hemimethylated DNA, or for fully modified DNA.
Differences between the interference patterns for the unmodified DNA provide evidence that
methylation of the target sequence influences the conformation of the protein-DNA interface,
and illustrate the importance of S-adenosyl methionine in the distinction between unmodified
and methylated DNA by the methyltransferase.

<>

<1>Powell, R.J., Bachvaroff, T.R., Hill, R.T.
<2>Draft Genome Sequence of the Alga-Aggregating Bacterium Bacillus sp. Strain RP1137.
<3>Genome Announcements
<4>2
<5>e00973-13
<6>2014
<7>Bacillus sp. strain RP1137 is a bacterium that is able to rapidly and efficiently aggregate
biofuel-producing microalgae. By 16S rRNA gene sequencing, it was found
to be related to the industrially important Bacillus megaterium. Here, we report
the draft genome sequence of Bacillus sp. strain RP1137.

<>

<1>Power, K.A., Yan, Q., Fox, E.M., Cooney, S., Fanning, S.
<2>Genome Sequence of Cronobacter sakazakii SP291, a Persistent Thermotolerant Isolate Derived from a Factory Producing Powdered Infant Formula.
<3>Genome Announcements
<4>1
<5>e0008213
<6>2013
<7>Cronobacter is an opportunistic pathogen associated with meningitis in neonates.  Based on
long-term surveillance of a powdered infant formula production facility,
a persistent and thermotolerant isolate, denoted Cronobacter sakazakii SP291, was
detected. Here we report the complete genome along with the sequences of three
plasmids identified in this organism.

<>

<1>Power, S.E., Harris, H.M., Bottacini, F., Ross, R.P., O'Toole, P.W., Fitzgerald, G.F.
<2>Draft Genome Sequence of Lactobacillus crispatus EM-LC1, an Isolate with Antimicrobial Activity Cultured from an Elderly Subject.
<3>Genome Announcements
<4>1
<5>e01070-13
<6>2013
<7>Here we report the 1.86-Mb draft genome sequence of Lactobacillus crispatus EM-LC1, a fecal
isolate with antimicrobial activity. This genome sequence is
expected to provide insights into the antimicrobial activity of L. crispatus and
improve our knowledge of its potential probiotic traits.

<>

<1>Powney, R., Smits, T.H., Sawbridge, T., Frey, B., Blom, J., Frey, J.E., Plummer, K.M., Beer, S.V., Luck, J., Duffy, B., Rodoni, B.
<2>Genome Sequence of an Erwinia amylovora Strain with Restricted Pathogenicity to Rubus Plants.
<3>J. Bacteriol.
<4>193
<5>785-786
<6>2010
<7>Here we present the genome of a strain of Erwinia amylovora, the fire blight pathogen, with
restricted pathogenicity to Rubus spp. Comparative
genomics of ATCC BAA-2158 with E. amylovora strains from non-Rubus hosts
identified significant genetic differences but supports the inclusion of
this strain within the species E. amylovora.

<>

<1>Pozidis, C., Vlatakis, G., Bouriotis, V.
<2>Sequence-specific DNA affinity chromatography: application of a group-specific adsorbent for the isolation of restriction endonucleases.
<3>J. Chromatogr.
<4>630
<5>151-157
<6>1993
<7>The use of sequence-specific DNA affinity adsorbents for the isolation of restriction
endonuclease EcoRI and SphI to near homogeneity has been reported. However, the high cost of
these adsorbents is a limiting factor for their wider application. This paper reports the
application of sequence-specific DNA affinity ligands containing recognition sequences for 34
restriction endonucleases as group specific ligands in the isolation of restriction
endonuclease. Crude samples of six restriction endonucleases, namely BshFI, BamHI, SmaI,
SacII, PvuII and SalI, were shown to bind to these adsorbents and could be eluted at different
KCl concentrations. High purification factors and recoveries were obtained. Restriction
endonuclease BshFI, an isoschizomer of HaeIII, from the microorganism Bacillus sphaericus was
purified to near homogeneity employing a two-step procedure which involves DNA-cellulose
chromatography and oligonucleotide-ligand affinity chromatography. The enzyme exists as a
monomer with an apparent relative molecular mass of 34000 as determined by both sodium dodecyl
sulphate-polyacrylamide gel electrophoresis and size exclusion chromatography.

<>

<1>Pozidis, C., Vlatakis, G., Bouriotis, V.
<2>Sequence-specific DNA affinity chromatography: Application of a group specific adsorbent for the isolation of restriction endonucleases.
<3>Prog. Biotechnol.
<4>9
<5>543-546
<6>1994
<7>Several rapid and effective methods have been described to obtain restriction endonucleases
suitable for commercial exploitation. However lengthy and laborious protocols have been
necessary to obtain homogeneous enzymes. The use of sequence-specific DNA affinity
chromatography to purify restriction endonucleases EcoRI and SphI to near homogeneity in a two
step procedure has been recently reported. However, the high cost of these adsorbents is a
limiting factor for their wider application. The application of a sequence-specific DNA
affinity ligand containing recognition sequences for 34 restriction endonucleases as
group-specific ligand for the isolation of restriction endonucleases is now reported. Crude
samples of six restriction endonucleases namely BshFI, BamHI, SmaI, SacII, PvuII and SalI were
shown to bind to this adsorbent and could be eluted at different Kcl concentrations with
purification factors obtained varying from 8 to greater than 300 fold and recoveries from
75-94%. Furthermore, restriction endonuclease BshFI, an isoschizomer of HaeIII from the
microorganism Bacillus sphaericus was purified to near homogeneity employing a two step
procedure which involves DNA cellulose chromatography and oligonucleotide ligand affinity
chromatography.

<>

<1>Prabhakara, S., Khedkar, S., Loganathan, R.M., Chandana, S., Gowda, M., Arakere, G., Seshasayee, A.S.
<2>Draft Genome Sequence of Staphylococcus aureus 118 (ST772), a Major Disease Clone from India.
<3>J. Bacteriol.
<4>194
<5>3727-3728
<6>2012
<7>We report the draft genome sequence of an ST772 Staphylococcus aureus disease isolate carrying
staphylococcal cassette chromosome mec (SCCmec) type V from a
pyomyositis patient. Our de novo short read assembly is approximately 2.8 Mb and
encodes a unique Panton-Valentine leukocidin (PVL) phage with structural genes
similar to those of varphi7247PVL and novel lysogenic genes at the N termini.

<>

<1>Pradel, N., Ji, B., Gimenez, G., Talla, E., Lenoble, P., Garel, M., Tamburini, C., Fourquet, P., Lebrun, R., Bertin, P., Denis, Y., Pophillat, M., Barbe, V., Ollivier, B., Dolla, A.
<2>The First Genomic and Proteomic Characterization of a Deep-Sea Sulfate Reducer: Insights into the Piezophilic Lifestyle of Desulfovibrio piezophilus.
<3>PLoS ONE
<4>8
<5>E55130
<6>2013
<7>Desulfovibrio piezophilus strain C1TLV30(T) is a piezophilic anaerobe that was
isolated from wood falls in the Mediterranean deep-sea. D. piezophilus represents
a unique model for studying the adaptation of sulfate-reducing bacteria to
hydrostatic pressure. Here, we report the 3.6 Mbp genome sequence of this
piezophilic bacterium. An analysis of the genome revealed the presence of seven
genomic islands as well as gene clusters that are most likely linked to life at a
high hydrostatic pressure. Comparative genomics and differential proteomics
identified the transport of solutes and amino acids as well as amino acid
metabolism as major cellular processes for the adaptation of this bacterium to
hydrostatic pressure. In addition, the proteome profiles showed that the
abundance of key enzymes that are involved in sulfate reduction was dependent on
hydrostatic pressure. A comparative analysis of orthologs from the
non-piezophilic marine bacterium D. salexigens and D. piezophilus identified
aspartic acid, glutamic acid, lysine, asparagine, serine and tyrosine as the
amino acids preferentially replaced by arginine, histidine, alanine and threonine
in the piezophilic strain. This work reveals the adaptation strategies developed
by a sulfate reducer to a deep-sea lifestyle.

<>

<1>Pradhan, S., Adams, R.L.P.
<2>DNA methylation in plants: involvement of two different methyltransferases.
<3>Biochem. Soc. Trans.
<4>22
<5>297S
<6>1994
<7>The DNA of higher eukaryotes is methylated at carbon 5 of some cytosine residues. In
vertebrates, 3 to 8% of cytosine residues are methylated, whereas in plants as many as 30% of
the total cytosines are methylated. The higher content of methylated cytosine in the DNA of
some plants could be partly attributed to their large genome with many repetitive sequences.
However, in the vertebrate genome 5-methylcytosine (5mC) is largely confined to CG
dinucleotides, whereas in higher plants both CG dinucleotides and CNG trinucleotides are
methylated. In non-vascular plants, methylation appears to occur only at CNG trinucleotides.
Methylation of DNA in plants, as in vertebrates, is implicated in the regulation of gene
expression; an effect that may be direct, through DNA:transcription factor interaction, or
indirect via an alteration in chromatin structure.

<>

<1>Pradhan, S., Adams, R.L.P.
<2>Distinct CG and CNG DNA methyltransferases in Pisum sativum.
<3>Plant J.
<4>7
<5>471-481
<6>1995
<7>DNA methyltransferase activity, present in low salt extracts of nuclei from young pea shoot
apices, has been fractionated into two different species by assaying with model substrates.
The CG methyltransferase (an unstable enzyme believed to be of 140 kDa) methylates cytosine
only in oligonucleotides with CG and CI dinucleotide targets while an enzyme of 110 kDa (the
CNG methyltransferase) methylates the cytosines in 5'-CAG-3' and 5'-CTG-3' target
sequences, especially when hemimethylated, but not in 5'-CCG-3' nor in 5'-CGG-3' target
sequences present in oligonucleotides.

<>

<1>Pradhan, S., Bacolla, A., Wells, R.D., Roberts, R.J.
<2>Recombinant human DNA (cytosine-5) methyltransferase.
<3>J. Biol. Chem.
<4>274
<5>33002-33010
<6>1999
<7>A method is described to express and purify human DNA (cytosine-5) methyltransferase (human
DNMT1) using a protein splicing (intein) fusion partner in a baculovirus expression vector.
The system produces ~1 mg of intact recombinant enzyme >95% pure per 1.5 x 10^9 insect cells.
The protein lacks any affinity tag and is identical to the native enzyme except for the two
C-terminal amino acids, proline and glycine, that were substituted for lysine and aspartic
acid for optimal cleavage from the intein affinity tag. Human DNMT1 was used for steady-state
kinetic analysis with poly(dI-dC).poly(dI-dC) and unmethylated and hemimethylated 36- and
75-mer oligonucleotides. The turnover number (k(cat)) was 131-237 h(-1) on
poly(dI-dC).poly(dI-dC), 1.2-2.3 h(-1) on unmethylated DNA, and 8.3-49 h(-1) on hemimethylated
DNA. The Michaelis constants for DNA (K(m)(CG)) and S-adenosyl-L-methionine (AdoMet)
(K(m)(AdoMet)) ranged from 0.33-1.32 and 2.6-7.2 muM, respectively, whereas the ratio of
k(cat)/K(m)(CG) ranged from 3.9 to 44 (237-336 for poly(dI-dC).poly(dI-dC)) x 10(6) M(-1)
h(-1). The preference of the enzyme for hemimethylated, over unmethylated, DNA was 7-21-fold.
The values of k(cat) on hemimethylated DNAs showed a 2-3-fold difference, depending upon which
strand was pre-methylated. Furthermore, human DNMT1 formed covalent complexes with substrates
containing 5-fluoro-CNG, indicating that substrate specificity extended beyond the canonical
CG dinucleotide. These results show that, in addition to maintenance methylation, human DNMT1
may also carry out de novo and non-CG methyltransferase activities in vivo.

<>

<1>Pradhan, S., Cummings, M., Roberts, R.J., Adams, R.L.P.
<2>Isolation, characterization and baculovirus-mediated expression of cDNA encoding cytosine DNA methyltransferase from Pisum sativum.
<3>Nucleic Acids Res.
<4>26
<5>1214-1222
<6>1998
<7>A series of overlapping clones complementy to the Arabidopsis cytosine-5 DNA methyltransferase
has been isolated from pea cDNA libraries.  The assembled nucleic acid sequence contains an
open reading frame of 4761 bp encoding a protein of 1554 amino acids.  Like other eukaryotic
C-5 MTases, the inferred protein has a presumed regulatory N-terminal region linked to a
catalytic C-terminal domain, which has eight of the ten conserved motifs found in prokaryotic
C-5 MTases.  The pea C-5 MTase has 65% identity at the nucleotide level and 61% identity at
the protein level, with the Arabidopsis C-5 MTase.  The catalytic domain of the pea enzyme
shares 78% identity with Arabidopsis and ~52% identity with murine and human C-5 MTases,
including the relative position of the proline-cysteine dipeptides of the catalytic center.
Using the conserved region of the cDNA as a probe, we have identified a transcript of 5 kb.
Southern blot analysis of pea genomic DNA with the above probe indicates the presence of a
single gene.  Using poly(A)+ RNA from different developmental stages and different tissues, we
have observed that expression is confined mostly to the rapidly dividing tissues of the plant.
Expression of this assembled cDNA in a baculovirus system gives a protein of ~174 kDa.  The
expressed protein can be cross-linked, in an AdoMet-dependent manner, to duplex
oligonucleotide substrates containing FdC in place of target cytosines in either CG or CAG/CTG
sequences.

<>

<1>Pradhan, S., Esteve, P.O.
<2>Allosteric activator domain of maintenance human DNA (cytosine-5) methyltransferase and its role in methylation spreading.
<3>Biochemistry
<4>42
<5>5321-5332
<6>2003
<7>The human maintenance DNA (cytosine-5) methyltransferase (hDNMT1) consists of a large
N-terminal regulatory domain fused to a catalytic C-terminal
domain by randomly repeated Gly-Lys dipeptides. Several N-terminal
deletion mutants of hDNMT1 were made, purified, and tested for substrate
specificity. Deletion mutants lacking 121, 501, 540, or 580 amino acids
from the N-terminus still functioned as DNA methyltransferases, methylated
CG sequences, and preferred hemimethylated to unmethylated DNA, as did the
full-length hDNMT1. Methylated DNA stimulated methylation spreading on
unmethylated CpG sequences for the full-length and the 121 amino acid
deletion hDNMT1 equally well but not for the mutants lacking 501, 540, or
580 amino acids, indicating the presence of an allosteric activation
determinant between amino acids 121 and 501. Peptides from the N- and
C-termini bound methylated DNA independently. Point mutation analysis
within the allosteric region revealed that amino acids 284-287 (KKHR) were
involved in methylated DNA-mediated allosteric activation. Allosteric
activation was reduced in the double point mutant enzymes D25 (K284A and
K285A) and D12 (H286A and R287A). Retinoblastoma gene product (Rb), a
negative regulator of DNA methylation, bound to the allosteric site of
hDNMT1 and inhibited methylation, suggesting Rb may regulate methylation
spreading.

<>

<1>Pradhan, S., Esteve, P.O.
<2>Mammalian DNA (cytosine-5) methyltransferases and their expression.
<3>Clin. Immunol.
<4>109
<5>6-16
<6>2003
<7>Two classes of functional DNA (cytosine-5) methyltransferases have been discovered in mammals
to date. One class methylates the unmodified DNA and
is designated as the de novo enzyme, whereas the other maintains the
methylation status of the daughter strand during DNA replication and thus
is referred to as a maintenance DNA methyltransferase. Each enzyme
catalyzes methyl group transfer from S-adenosyl-L-methionine to cytosine
bases in DNA. During methylation the enzyme flips its target base out of
the DNA duplex into a typically concave catalytic pocket. This flipped
cytosine base is then a substrate for the enzyme-catalyzed reaction. The
newly formed 5-methylcytosine confers epigenetic information on the
parental genome without altering nucleotide sequences. This epigenetic
information is inherited during DNA replication and cell division. In
mammals, DNA methylation participates in gene expression, protection of
the genome against selfish DNA, parental imprinting, mammalian X
chromosome inactivation, developmental regulation, T cell development, and
various diseases.

<>

<1>Pradhan, S., Houlston, C., Cummings, M., Adams, R.L.P.
<2>CG and CNG methyltransferases in plants.
<3>Gene
<4>157
<5>289-291
<6>1995
<7>We have purified two distinct DNA methyltransferases from pea shoot tips and analyzed their
sequence specificity using synthetic oligodeoxyribonucleotide substrates and chemical
sequencing methods.  One methylates only CG target sequences, whereas the other methylates
only CAG or CTG target sequences.  We have found no evidence for methylation of the 5'
cytosine in CCG target sequences either in vivo or in vitro.  Using amino-acid sequence data,
PCR-amplified fragments from conserved sequences and heterologous probes, we have isolated
several cDNA clones that react with mRNA molecules of different sizes.

<>

<1>Pradhan, S., Kim, G.D.
<2>The retinoblastoma gene product interacts with maintenance human DNA (cytosine-5) methyltransferase and modulates its activity.
<3>EMBO J.
<4>21
<5>779-788
<6>2002
<7>The mammalian DNA (cytosine-5) methyltransferase (Dnmt1) is involved in the maintenance of
methylation patterns in the genome during DNA replication and development.  The retinoblastoma
gene product, Rb, is a cell cycle regulator protein that represses transcription by recruiting
histone deacetylase (HDAC1).  In vivo, histone deacetylase associates with Dnmt1.  Here we
show that Rb itself associates with human Dnmt1 (hDnmt1) independently of its own
phosphorylation status.  Methyltransferase activity was co-purified with Rb.  The regulatory
domain of hDnmt1 binds strongly to the B and C pockets of Rb (amino acids 701-872) and
inhibits methyltransferase activity by disruption of the hDnmt1-DNA binary complex.  Weak
interaction of Rb pockets A and B with Dnmt1 was also observed.  Overexpression of Rb leads to
hypomethylation of the cellular DNA, suggesting that Rb may modulate Dnmt1 activity during DNA
replication in the cell cycle.

<>

<1>Pradhan, S., Roberts, R.J.
<2>Hybrid mouse-prokaryotic DNA (cytosine-5) methyltransferases retain the specificity of the parental C-terminal domain.
<3>EMBO J.
<4>19
<5>2103-2114
<6>2000
<7>The mouse (cytosine-5) DNA methyltransferase (Dnmt1) consists of a regulatory N-terminal and a
catalytic C-terminal domain, which are fused by a stretch of Gly-Lys dipeptide repeats. The
C-terminal region contains all of the conserved motifs found in other cytosine-5 DNA
methyltransferases including the relative position of the catalytic Pro-Cys dipeptide. In
prokaryotes, the methyltransferases are simpler and lack the regulatory N-terminal domain. We
constructed three hybrid methyltransferases, containing the intact N-terminus of the murine
Dnmt1 and most of the coding sequences from M.HhaI (GCGC), M.HpaII (CCGG) or M.SssI (CG).
These hybrids are biologically active when expressed in a baculovirus system and show the
specificity of the parental C-terminal domain. Expression of these recombinant constructs
leads to de novo methylation of both host and viral genomes in a sequence-specific manner.
Steady-state kinetic analyses were performed on the murine Dnmt1-HhaI hybrid using
poly(dG-dC).poly (dG-dC), unmethylated and hemimethylated oligonucleotides as substrates. The
enzyme has a slow catalytic turnover number of 4.38 h(-1) for poly(dG-dC). poly(dG-dC), and
exhibits 3-fold higher catalytic efficiency for hemimethylated substrates.

<>

<1>Pradhan, S., Talbot, D., Sha, M., Benner, J., Hornstra, L., Li, E., Jaenisch, R., Roberts, R.J.
<2>Baculovirus-mediated expression and characterization of the full-length murine DNA methyltransferase.
<3>Nucleic Acids Res.
<4>25
<5>4666-4673
<6>1997
<7>The original cDNA sequence reported for the murine DNA methyltransferase was not full length.
Recently, additional cDNA sequences have been reported that lie upstream of the original and
contain an extended open reading frame with three additional ATGs in frame with the coding
region.  Genomic DNA upstream of this ATG contains two more ATGs in frame and no obvious
splice site.  We have constructed, and expressed in baculovirus, MTase clones that begin at
each of these four ATGs and examined their properties.  Constructs beginning with any of the
first three ATGs as their initiator methionines give a predominant DNA MTase band of ~185 kDa
on SDS-PAGE corresponding to translational initiation at the third ATG.  The fourth ATG
construct gives a much smaller protein band of 173 kDa.  The 185 kDa protein was purified by
HPLC, characterized by mass spectrometry and has a measured molecular mass of 184+/- 0.5 kDa.
All of these MTases were functional in vitro and steady state kinetic analysis showed that the
recombinant proteins exhibit similar kinetic properties irrespective of their length.  The
homogeneous recombinant enzyme from the fourth ATG construct shows a 2.5-fold preference for a
hemi-methylated DNA substrate as compared to an unmethylated substrate, whereas the 185 kDa
protein is equally active on both substrates.  The kinetic properties of the recombinant
enzyme are similar to those reported for the native MTase derived from murine erythroleukemia
cells.  The new clones are capable of yielding large quantities of intact MTases for further
structural and functional studies.

<>

<1>Prajapati, J.B., Khedkar, C.D., Chitra, J., Suja, S., Mishra, V., Sreeja, V., Patel, R.K., Ahir, V.B., Bhatt, V.D., Sajnani, M.R., Jakhesara, S.J., Koringa, P.G., Joshi, C.G.
<2>Whole-Genome Shotgun Sequencing of an Indian-Origin Lactobacillus helveticus Strain, MTCC 5463, with Probiotic Potential.
<3>J. Bacteriol.
<4>193
<5>4282-4283
<6>2011
<7>Lactobacillus helveticus MTCC 5463 was isolated from a vaginal swab from a healthy adult
female. The strain exhibited potential probiotic properties,
with their beneficial role in the gastrointestinal tract and their ability
to reduce cholesterol and stimulate immunity. We sequenced the whole
genome and compared it with the published genome sequence of Lactobacillus
helveticus DPC4571.

<>

<1>Prajapati, J.B., Khedkar, C.D., Chitra, J., Suja, S., Mishra, V., Sreeja, V., Patel, R.K., Ahir, V.B., Bhatt, V.D., Sajnani, M.R., Jakhesara, S.J., Koringa, P.G., Joshi, C.G.
<2>Whole-Genome Shotgun Sequencing of Lactobacillus rhamnosus MTCC 5462, a Strain with Probiotic Potential.
<3>J. Bacteriol.
<4>194
<5>1264-1265
<6>2012
<7>Lactobacillus rhamnosus MTCC 5462 was isolated from infant gastrointestinal flora. The strain
exhibited an ability to reduce cholesterol and stimulate
immunity. The strain has exhibited positive results in alleviating
gastrointestinal discomfort and good potential as a probiotic. We sequenced the
whole genome of the strain and compared it to the published genome sequence of
Lactobacillus rhamnosus GG (ATCC 53103).

<>

<1>Prakash, A., Chung, S., Ryu, J.-I.
<2>Study of the expression of hsd genes of E. coli K-12 after conjugal transfer.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>165
<6>1991
<7>The enzyme for DNA restriction and modification in E. coli K-12 is encoded by
three contiguous genes: hsdRK (restriction), hsdMK (modification) and hsdSK
(specificity).  Two promoters are known pres upstream of hsdRK, and pmod
upstream of hsdMK and hsdSK.  F plasmids carrying the wild-type hsd genes can
be transferred to a non-K modified recipient (e.g., E. coli) without killing
the recipient strain.  Thus there must be control at either the
transcriptional, translational or post-translational level which allows the
modification of the recipient's chromosome, thereby preventing its restriction.
F plasmids carrying various hsd-lacZ operon fusions were constructed and
subsequent F transfer experiments showed simultaneous expression of both pres
and pmod promoters, and thus no control at the transcriptional level was
indicated.  Further we have isolated a mutant of E. coli C which is killed
(efficiency of transfer <10/4) upon conjugal transfer of the wild type hsd
genes, suggesting a genetic basis for this control phenomenon.  A plasmid
expressing the R subunit was then introduced into this mutant as well as into
wild type E. coli C.  Subsequent conjugal transfer of an F plasmid carrying the
hsdRK hsdM+K hsdS+K genes into this mutant resulted in complementation between
the incoming M and S subunits with the pre-existing R subunit, as evidenced by
the killing of the mutant recipient.  However, in the case of the wild type
recipient strain harboring the plasmid expressing the R subunit, modification
of the recipient DNA occurred, resulting in successful conjugation.  This
suggests that there is control of the incoming hsd gene expression in the
recipient at the post-translational level.

<>

<1>Prakash, A., Valinluck, B., Ryu, J.-I.
<2>Genomic hsd-Mu(lac) operon fusion mutants of Escherichia coli K-12.
<3>Gene
<4>99
<5>9-14
<6>1991
<7>Genomic (chromosomal) hsd-Mu(lac) operon fusions have been constructed in two
strains of Escherichia coli K-12 for the three hsd genes, hsdRK, hsdMK and
hsdSK, using MudX and lambda placMu53.  Expression of hsdK mutants ranged from
16 to 74 units (u) (with a mean of 52 u) for fusions to promoter Pres and
ranged from 26-75 u (also with a mean of 52 u) for fusions to promoter Pmod.
The expression of the two hsdK promoters was measured in different stages of
growth.  The Pres fusion mutant showed a lag in Beta-galactosidase (BetaGal)
production, as compared to the Pmod fusion mutant.  One rK-mK mutant (JR205)
showed more than ten times the BetaGal activity of other insertion mutants.
The activity of this mutant decreased by 20-fold upon the transfer of F101-102,
which includes the wild-type hsd region.  Positive gene-dosage effect was
observed using F' plasmids containing the hsd-lacZ region.

<>

<1>Prakash, T., Oshima, K., Morita, H., Fukuda, S., Imaoka, A., Kumar, N., Sharma, V.K., Kim, S.-W., Takahashi, M., Saitou, N., Taylor, T.D., Ohno, H., Umesaki, Y., Hattori, M.
<2>Complete Genome Sequences of Rat and Mouse Segmented Filamentous Bacteria, a Potent Inducer of Th17 Cell Differentiation.
<3>Cell Host Microbe
<4>10
<5>273-284
<6>2011
<7>Segmented filamentous bacteria (SFB) are noncultivable
commensals inhabiting the gut of various vertebrate
species and have been shown to induce Th17
cells in mice. We present the complete genome
sequences of both rat and mouse SFB isolated from
SFB-monocolonized hosts. The rat and mouse SFB
genomes each harbor a single circular chromosome
of 1.52 and 1.59 Mb encoding 1346 and 1420
protein-coding genes, respectively. The overall nucleotide
identity between the two genomes is 86%, and
the substitution rate was estimated to be similar to
that of the free-living E. coli. SFB genomes encode
typical genes for anaerobic fermentation and spore
and flagella formation, but lack most of the amino
acid biosynthesis enzymes, reminiscent ofpathogenic
Clostridia, exhibiting large dependency on the host.
However, SFB lack most of the clostridial virulencerelated
genes. Comparative analysis with clostridial
genomes suggested possible mechanisms for host
responses and specific adaptations in the intestine.

<>

<1>Prakash-Cheng, A., Chung, S.S., Ryu, J.-I.
<2>The expression and regulation of hsdK genes after conjugative transfer.
<3>Mol. Gen. Genet.
<4>241
<5>491-496
<6>1993
<7>The type I restriction and modification genes of Escherichia coli can be transferred to other
non-modified strains by conjugation without killing the recipient, implying that the
restriction function must be regulated. In this study, two isogenic F plasmids (r+k and r-K)
served as donors in quantitative conjugation experiments with various restriction-deficient
strains of E. coli and Salmonella typhimurium. Conjugation studies with hsd:lacZ operon
fusions in F' plasmids indicate that both the hsdK promoters, Pres and Pmod, express
simultaneously following conjugative transfer. Thus these genes do not appear to be regulated
at the transcriptional level. A spontaneous mutant of E. coli C was discovered that is
presumably killed upon conjugative transfer of the hsdK genes (defined as a Crc- phenotype).
The gene that is defective in the mutant was tentatively designated hsdC (control). Hfr gene
replacement studies led to the localization of the putative hsdC gene between 6 and 16 min on
the E. coli genetic map.

<>

<1>Prakash-Cheng, A., Ryu, J.
<2>Delayed expression of in vivo restriction activity following conjugal transfer of Escherichia coli hsdk (restriction-modification) genes.
<3>J. Bacteriol.
<4>175
<5>4905-4906
<6>1993
<7>Following conjugal transfer of the hsdk genes (hsdRK, hsdMk, and hsdSk) of Escherichia coli
K-12, restriction activity was first detected only after approximately 15 generations, whereas
modification activity was observed immediately. This sequential expression explains the
establishment of hsdk genes in a nonmodified host and suggests regulation of restriction
activity after conjugal transfer.

<>

<1>Prangishvili, D.A., Chinchaladze, D.Z., Chelidze, M.G.
<2>Thermostability of nucleic acid and restriction endonuclease template synthesis enzymes from extremely thermophilic archebacteria.
<3>Biotekhnologiya
<4>3
<5>440-446
<6>1987
<7>A study has been made of the thermostability of DNA-dependent RNA polymerases, DNA-dependent
DNA polymerases, and restriction endonucleases from extremely thermophilic archebacteria. The
optimum temperature for enzyme catalytic activity (T opt) lay in the optimum temperature
region for growth of the organisms involved: at 85C for RNA polymerases from Thermoproteus
tenax and Desulfurococcus mucosus, at 55-65C and 70-75C for three DNA polymerases and
restrictase SuaI from Sulfolobus acidocaldarius, respectively, and at 55-60C for DNA
polymerase and restrictase ThaI from Thermoplasma acidophilum. All the enzymes investigated
were highly stable at temperatures up to T opt.

<>

<1>Prangishvili, D.A., Vashakidze, R.P., Chelidze, M.G., Chinchaladze, D.Z., Tsalkalamanidze, N.V.
<2>Isolation and properties of a restriction endonuclease SuaI from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius.
<3>Biokhimiia
<4>52
<5>1043-1050
<6>1987
<7>A new restriction endonuclease SuaI was isolated from the thermoacidophilic
archaebacterium Sulfolobus acidocaldarius.  The enzyme is an isoschizomer of
BspRI; it recognizes the tetranucleotide GGCC and cleaves DNA in the center of
this sequence.  SuaI requires Mg2+, the optimal concentration being 6 mM.  KCl
at concentrations above 25 mM significantly inhibits the enzyme activity.  The
pH optimum lies within the range of 6-7 at 70C, the temperature optimum is at
70-75C.  the enzyme is highly stable at temperatures up to 80C.  DNA of S.
acidocaldarius is not cleaved by the enzyme.

<>

<1>Prangishvili, D.A., Vashakidze, R.P., Chelidze, M.G., Gabriadze, I.Y.
<2>A restriction endonuclease SuaI from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius.
<3>FEBS Lett.
<4>192
<5>57-60
<6>1985
<7>A type II restriction endonuclase (SuaI) has been isolated from the
thermoacidophilic archaebacterium Sulfolobus acidocaldarius.  The enzyme is an
isoschizomer of BspRI.  It does not cut S. acidocaldarius DNA, as the
recognition sequence GGCC in this DNA contains modified nucleotide(s).  The
enzyme is most active at 60-70C and is highly thermostable.

<>

<1>Prasad, B.J., Sabnis, K., Deobagkar, D.D., Deobagkar, D.N.
<2>Deinococcus radiodurans strain R1 contains N6-methyladenine in its genome.
<3>Biochem. Biophys. Res. Commun.
<4>335
<5>412-416
<6>2005
<7>Methylation of DNA is known to be involved in DNA repair mechanisms in bacteria. Deinococcus
radiodurans strain R1 on exposure to high
radiation undergoes significant DNA damage, which is repaired without
mutations. However, the presence of modified nucleotides has not been
reported in its genome. We report here the detection of
N6-methyladenine in the genome of D. radiodurans strain R1 using
immunochemical techniques. This N6-methyladenine is not a part of GATC
restriction-modification system. D. radiodurans cell extract also
exhibited a DNA adenine methyltransferase activity which was reduced in
the early post-irradiation recovery phase.

<>

<1>Prasannan, C.B.
<2>Modulation of restriction enzyme PvuII activity by metal ion cofactors.
<3>Ph.D. Thesis, University of Missouri
<4>
<5>
<6>2009
<7>Many nucleases require cofactors (usually Mg2+) for activity. Metal ions like Ca2+, Mn2+,
Tb3+, Eu3+ etc., can bind at the active site and have varying effects on the enzymatic
activity. In a crystal structure with DNA, two Ca2+ ions were bound to the active site of each
PvuII monomer, and the metals share the ligands. Ca2+ ions promote DNA binding and not
cleavage in many nucleases. We explored the role of Mg2+ and Ca2+ binding at the metal binding
sites of PvuII to elucidate the role of each metal site. Ca2+ binding at the catalytic metal
site would inhibit the cleavage activity,
whereas its binding at the regulatory site can have varied effects. In EcoRV enzyme, it was
seen that Ca2+ at the regulatory site increases the activity of the enzyme. We monitored the
cleavage kinetics of PvuII in presence of mixed metals. The cleavage kinetics data for Ca2+
and Mg2+ set were modeled using parameters from previous studies on PvuII. Our global analysis
on single turnover cleavage kinetics datasets show best fit to a model in which mixed metal
species are formed and active. The cleavage rate constants for the mixed metal species ranged
from 0.01-0.08 sec-1, which is similar to the rate when only one metal is bound. From earlier
work in our lab, Tb3+ was shown to have a tight (2 ìM) binding site and a weak binding site in
PvuII. The difference in affinity allows one site to be filled with Tb3+ and the other with
another metal. Indirect Tb3+ luminescence spectroscopy of the Tb3+ bound to enzyme in presence
of other metals indicates that Ca2+ and Mn2+ displace Tb3+ from the enzyme. This was observed
by the decrease in the luminescence intensity of E-Tb3+ complex with the addition of Ca2+/Mn2+
ions. Under similar conditions, the addition of Mg2+ ions to the E-Tb3+ complex results in an
increase in the signal observed. This indicates the formation of the mixed species
E-Tb3+-Mg2+. No enzymatic activity was detected for the enzyme with the addition of Mg2+ to
the E-Tb3+ complex, whereas with the addition of Mn2+ ions there was detectable activity.
The observed activity with Mn2+ ion was due to the displacement of Tb3+ ions from the active
site, forming the active EMn2+Mn2+ species. Although the E-Mg2+-Tb3+ species is catalytically
inactive, it does bind the DNA as confirmed by fluorescence anisotropy using nonhydrolyzable
phosphoramidate DNA.

<>

<1>Prere, M.F., Fayet, O.
<2>DNA methylase activities in a Neisseria gonorrhoeae extract.
<3>FEMS Microbiol. Lett.
<4>33
<5>37-41
<6>1986
<7>Resistance of Neisseria gonorrhoeae DNA to cleavage by various restriction endonucleases
suggests the existence of modification enzymes which protect the corresponding recognition
sequences. We indeed found methylase activities in N. gonorrhoeae extracts. These activities
lead to the methylation of adenine and cytosine residues in bacteriophage lambda DNA and DNA
from an Escherichia coli Dam- strain. They also result in partial protection of lambda DNA to
cleavage by the restriction endonucleases HaeII, HaeIII, BamHI and SacII.

<>

<1>Prere, M.F., Fayet, O.
<2>Susceptibility of Neisseria Gonorrhoeae DNA to Cleavage by Restriction Endonuclease KpnI.
<3>Ann. Inst. Pasteur Microbiol.
<4>136A
<5>329-338
<6>1985
<7>We compared the susceptibility of Escherichia coli and Neisseria gonorrhoeae
DNA to cleavage by KpnI and found that KpnI has a lower affinity for gonococcal
DNA.  Site-specific methylation is suggested as the cause of this altered
affinity.

<>

<1>Prere, M.F., Fayet, O.
<2>DNA modification in Neisseria gonorrhoeae: Resistance of DNA of 19 strains to cleavage by restriction enzymes.
<3>Ann. Inst. Pasteur Microbiol.
<4>136A
<5>323-328
<6>1985
<7>Site-specific modification is frequently encountered in the DNA of bacteria.  It is generally
due to the methylation of either a cytosine or an adenine in short sequences 4- to
6-base-pairs long.  One way to reveal the presence of site-specific methylation in a DNA
sample is to demonstrate its resistance to cleavage by a restriction endonuclease.  In
Neisseria gonorrhoeae, studies performed in a limited number of strains revealed that their
DNA is resistant to digestion by several restriction enzymes.  Those included HaeII, HaeIII,
SacII, BamHI, NarI and KpnI.  In order to determine whether this pattern of resistance is a
general property, or if there is variation from strain to strain, we have analyzed the DNA of
a collection of 19 gonococcal strains for resistance to various restriction endonucleases.  We
have also investigated the plasmid content of these strains to determine whether this
correlates with a given resistance pattern.

<>

<1>Press, M.
<2>Method for purification of enzymes HindII and HindIII for gene manipulation.
<3>Czech Patent Office
<4>CS 272934
<5>
<6>1991
<7>A method for purification of restriction endonucleases HindII and HindIII by using column
liquid chromatography is described.  Both activities are separated on phosphocellulose, blue
cellulose or heparin-cellulose, respectively.  Combination of these columns or combination
with DEAE-cellulose results in higher efficiency of separation.

<>

<1>Press, M., Hamsikova, E.
<2>Purification of restriction endonuclease BamHI.
<3>Czech Patent Office
<4>CS 267506
<5>
<6>1990
<7>
<>

<1>Price, C., Bickle, T.A.
<2>A possible role for DNA restriction in bacterial evolution.
<3>Microbiol. Sci.
<4>3
<5>296-299
<6>1986
<7>The phenomena of restriction and modification (R-M) were first described more
than 30 years ago and it is now more than 20 years since Arber & Dussoix
proposed, rightly, that R-M was due to the action of endonucleases and
methylases acting at the same DNA sequences.  Briefly, the endonuclease (or
restriction enzyme) recognizes a specific DNA sequence as a signal to cleave
the DNA unless the sequence has previously been methylated by a modification
methylase of the same sequence specificity.  The DNA of a cell carrying a R-M
system will normally be modified and is therefore not a substrate for the
restriction enzyme.  The only natural substrate for a restriction enzyme is
invasive foreign DNA containing unmethylated recognition sequences.
Restriction is independent of the mode of introduction of the DNA into the
cell, and phage infection, plasmid conjugation or transformation all lead to
restriction.

<>

<1>Price, C., Bickle, T.A.
<2>Evolution of DNA sequence specificity in type I restriction enzymes.
<3>Biochem. Soc. Trans.
<4>16
<5>942-943
<6>1988
<7>None

<>

<1>Price, C., Gubler, M., Braguglia, D., Meister, J., Tyndall, C., Piekarowicz, A., Bickle, T.A.
<2>Evolution of DNA sequence specificity in type 1C restriction and modification systems.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>151
<6>1993
<7>The type I restriction-modification (R-M) systems of the Enterobacteriaceae form three
genetically complementing families, one of which is the type 1C family. All type I R-M systems
recognize DNA sequences that are asymmetric and split into two by a spacer that can have any
sequence but which, for a given R-M system, has fixed length. The specific DNA-binding
components of both restriction and modification enzymes are the products of the hsdS genes,
which form complexes with the hsdM and hsdR gene products. Within a family of type I systems
the hsdS genes have a characteristic structure with two variable regions, each coding a
protein domain (called Target Recognition Domains, TRD) that recognizes one half of the
recognition sequence, and two (three in one case) conserved regions that are thought to code
regions of the protein that are important for protein-protein interactions. The type IA and IC
R-M systems are so far the only DNA binding protein that have been show to alter their binding
specificity by natural means. Type IA enzymes can reassort their TRDs by recombination within
a conserved region of the hsdS gene, leading to the production of enzymes that have new,
hybrid recognition sequences. Type IC systems (and, presumable, type IB systems, although it
has not yet been demonstrated) can also change their specificity in this way. In addition type
IC systems can change the length of the central conserved region of the hsdS gene by unequal
crossing over at a 12 bp long repeated sequence. This leads to enzymes that recognize the same
specific sequence but with different lengths of non-specific spacer. We have discovered yet a
third way in which type IC enzymes can change their specificity. We have recovered a mutant
that has a transposon inserted in the middle of the hsdS gene and which nevertheless produces
an active R-M system with altered sequence specificity. The structure of the mutant enzymes
and the sequence that it recognizes with be presented. The sequence is the only known type I
recognition sequence that is palindromic. Finally, we will present data on a type IC R-M
system of as yet unknown DNA specificity that appears to be on a transposable element. This
system is particulary interesting because it is closely integrated, both genetically and
physically, with an RNA restriction system that is specific for cells infected by T even
phages.

<>

<1>Price, C., Lingner, J., Bickle, T.A., Firman, K., Glover, S.W.
<2>Basis for changes in DNA recognition by the EcoR124 and EcoR124/3 Type I DNA restriction and modification enzymes.
<3>J. Mol. Biol.
<4>205
<5>115-125
<6>1989
<7>EcoR124I and EcoR124/3I are type I DNA restriction and modification systems.
The EcoR124/3I system arose from the EcoR124I system some 15 years ago and at
the electron microscopic DNA heteroduplex level the genes for both systems are
still apparently identical.  We have shown that the DNA sequences recognized by
the two systems are GAA(N6)RTCG for EcoR124I and GAA(N7)RTCG for EcoR124/3I.
The sequences thus differ only in the length of the non-specific spacer.  This
difference nevertheless places the two specific domains of the EcoR124/3I
recognition sequence 0.34 nm further apart and rotates them 36C with respect to
those of EcoR124I, which implies major structural differences in the proteins
recognizing these sequences.  We have now determined the nucleotide sequences
of the hsdS and hsdM genes of both systems and of the hsdR gene of EcoR124/3I.
The hsdS gene products provide DNA sequence specificity in both restriction and
modification, the hsdM gene products are necessary for modification and all
three hsd gene products are required for restriction.  The only difference that
we have detected between the two systems is that a 12 base-pair sequence
towards the middle of the hsdS gene is repeated twice in the EcoR124I gene and
three times in the EcoR124/3I gene.  We have deleted one of the repeats in the
EcoR124/3I gene and shown that this changes the specificity to that of
EcoR124I.  Thus, the extra four amino acids in the middle of the EcoR124/3I
hsdS gene product, which in an alpha-helical configuration would extend 0.6 nm,
are sufficient to explain the differences in sequence recognition.  We suggest
that this kind of specificity change should not be rare in Nature.

<>

<1>Price, C., Pripfl, T., Bickle, T.A.
<2>EcoR124 and EcoR124/3:  the first members of a new family of type I restriction and modification systems.
<3>Eur. J. Biochem.
<4>167
<5>111-115
<6>1987
<7>We have purified the EcoR124 and EcoR124/3 restriction enzymes and shown that
they are type I enzymes by several criteria:  subunit composition, DNA and
S-adenosylmethionine-dependent ATPase activity, and site-specific DNA methylase
activity.  By immunochemical criteria these enzymes are related to each other
but are unrelated to the two previously investigated families of type I
restriction enzymes.  They form therefore a new family which we call type IC.
The arrangement of the structural genes coding for these enzymes and their
transcriptional organisation have been determined.  These are different from
the common arrangement found for the other two families of type I enzymes.

<>

<1>Price, C., Shepherd, J.C.W., Bickle, T.A.
<2>DNA recognition by a new family of type I restriction enzymes:  a unique relationship between two different DNA specificities.
<3>EMBO J.
<4>6
<5>1493-1497
<6>1987
<7>The DNA sequences recognized by the allelic type I restriction enzymes EcoR124
and EcoR124/3 were determined.  EcoR124 recognizes 5'-GAA(N6)RTCG-3' and
EcoR124/3 recognizes 5'-GAA(N7)RTCG-3'.  These are typical of sequences
recognized by type I recognition enzymes in that they consist of two specific
domains separated by a non-specific spacer sequence.  For these two enzymes,
the specific sequences are identical but the length of the non-specific spacer
is different.  The specific domains of EcoR124/3 are thus 3.4 angstroms further
apart than those of EcoR124 and rotated with respect to each other through a
further 36 degrees.

<>

<1>Prichodko, E.A., Rechnukova, N.I., Degtyarev, S.K.
<2>Determination of substrate specificity of restriction endonuclease Tru9I.
<3>Sib. Biol. J.
<4>1
<5>57-59
<6>1991
<7>Tru9I, type II restriction endonuclease, has been isolated from Thermus ruber
9.  The recognition sequence and cleavage point of restriction endonuclease
Tru9I have been determined as 5'T^TAA.  Tru9I is an isoschizomer of MseI.

<>

<1>Prichodko, G.G., Degtyarev, S.K., Rechkunova, N.I., Sosnovsev, S.V., Tchigikov, V.E.
<2>General method for determining the sites of DNA cleavage by restriction endonucleases.
<3>Biotekhnologiya
<4>1
<5>12-16
<6>1990
<7>A method for determining substrate specificity of large block cleaving restrictases is
suggested. Plasmid vectors pUBS18 and pUBS19, that permit labelling at one end of a Bse21I
site for rapid DNA sequencing were constructed. Restriction endonuclease BimI recognizing the
sequence TT^CGAA was isolated from the bacterial strain Brevibacterium immotum.

<>

<1>Prichodko, G.G., Petrov, N.A., Chizhikov, V.E., Degtyarev, S.K.
<2>A modified method for determining the sites of DNA cleavage by restriction endonucleases.
<3>Biotekhnologiya
<4>4
<5>618-620
<6>1988
<7>The modification merely involves labelling the 3' end of restriction fragments
using Klenow so as to avoid problems of the extra phosphate group which arise
when kinase-labelled fragments are compared with chemically degraded fragments.

<>

<1>Prichodko, G.G., Petrov, N.A., Repin, V.E., Degtyarev, S.K.
<2>Establishing the substrate specificity of restriction endonuclease Bsu15I.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>14
<5>108-109
<6>1988
<7>The recognition sequence and cleavage point of restriction endonuclease Bsu15I has been
determined as 5'AT^CGAT.

<>

<1>Prichodko, G.G., Rechnukova, N.I., Repin, V.E., Degtyarev, S.K.
<2>Determination of substrate specificity of restriction endonuclease Acc65I.
<3>Sib. Biol. J.
<4>1
<5>59-60
<6>1991
<7>The recognition sequence and cleavage point of restriction endonuclease Acc65I
have been determined as 5'G^GTACC.

<>

<1>Pride, D.T., Blaser, M.J.
<2>Identification of horizontally acquired genetic elements in Helicobacter pylori and other prokaryotes using oligonucleotide difference analysis.
<3>Genome Lett.
<4>1
<5>2-15
<6>2002
<7>The Helicobacter pylori genome contains genetic elements believed to have been horizontally
acquired. We compared a Markov chain method dependent on genomic oligonucleotide frequencies
with
methods based upon G + C content to identify potential horizontally acquired gene clusters.
Compared with G +
C content analysis, oligonucleotide difference analysis (ODA) identified a unique set of
genes, with a few genes
shared by the two methods. However, each method identified a large number of cag island and
plasticity zone
genes, with 12-18% identified by both. ODA also was applicable to other prokaryotes,
identifying as foreign
prophages in Bacillus subtilis, insertion elements in Campylobacter jejuni, genes with
repetitive elements in
Mycobacterium tuberculosis, and the integron island in Vibrio cholerae. GATC, the cognate
sequence for the
hpyIII restriction-modification system in H. pylori, is substantially underrepresented in the
plasticity zone,
suggesting a role for that R-M system in limiting horizontal transfer events. That GATC is not
substantially
underrepresented in the cag island suggests that the two islands likely have different
origins. The information
provided by ODA is complementary to G + C content for identifying horizontal acquisitions in
prokaryotes.

<>

<1>Pridgeon, J.W., Zhang, D., Zhang, L.
<2>Complete Genome Sequence of the Highly Virulent Aeromonas hydrophila AL09-71 Isolated from Diseased Channel Catfish in West Alabama.
<3>Genome Announcements
<4>2
<5>e00450-14
<6>2014
<7>Aeromonas hydrophila AL09-71 was isolated from diseased channel catfish in west Alabama during
a 2009 disease outbreak. The full genome of A. hydrophila AL09-71
is 5,023,861 bp. The availability of this genome will allow comparative genomics
to identify genes involved in pathogenesis or immunogens for the purpose of
vaccine development.

<>

<1>Pridgeon, J.W., Zhang, D., Zhang, L.
<2>Complete Genome Sequence of a Moderately Virulent Aeromonas hydrophila Strain, pc104A, Isolated from Soil of a Catfish Pond in West Alabama.
<3>Genome Announcements
<4>2
<5>e00554-14
<6>2014
<7>Aeromonas hydrophila pc104A is a moderately virulent strain isolated from the soil of a
catfish pond in west Alabama in 2010. Its full genome is 5,023,829 bp.
The availability of this genome will allow comparative genomics to identify the
virulence genes that are important for pathogenesis or immunogens for the purpose
of vaccine development.

<>

<1>Pridgeon, J.W., Zhang, D., Zhang, L.
<2>Complete Genome Sequence of a Virulent Strain, Streptococcus iniae ISET0901, Isolated from Diseased Tilapia.
<3>Genome Announcements
<4>2
<5>e00553-14
<6>2014
<7>Streptococcus iniae ISET0901 is a virulent strain isolated in 2007 from diseased  tilapia. Its
full genome is 2,070,856 bp. The availability of this genome will
allow comparative genomics to identify virulence genes important for the
pathogenesis of streptococcosis caused by S. iniae, as well as possible
immunogens for vaccine development.

<>

<1>Pridgeon, J.W., Zhang, D., Zhang, L.
<2>Complete Genome Sequence of the Attenuated Novobiocin-Resistant Streptococcus iniae Vaccine Strain ISNO.
<3>Genome Announcements
<4>2
<5>e00510-14
<6>2014
<7>Streptococcus iniae ISNO is an attenuated novobiocin-resistant vaccine strain. Its full genome
is 2,070,182 bp in length. The availability of this genome will
allow comparative genomics to identify potential virulence genes important for
pathogenesis of S. iniae and potential mechanisms associated with novobiocin
resistance in this strain.

<>

<1>Pridmore, R.D., Berger, B., Desiere, F., Vilanova, D., Barretto, C., Pittet, A.-C., Zwahlen, M.-C., Rouvet, M., Altermann, E., Barrangou, R., Mollet, B., Mercenier, A., Klaenhammer, T., Arigoni, F., Schell, M.A.
<2>The genome sequence of the probiotic intestinal bacterium Lactobacillus johnsonii NCC 533.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>101
<5>2512-2517
<6>2004
<7>Lactobacillus johnsonii NCC 533 is a member of the acidophilus group of intestinal
lactobacilli that has been extensively studied for their "probiotic" activities that include,
pathogen inhibition, epithelial cell attachment, and immunomodulation. To gain insight into
its physiology and identify genes potentially involved in interactions with the host, we
sequenced and analyzed the 1.99-Mb genome of L. johnsonii NCC 533. Strikingly, the organism
completely lacked genes encoding biosynthetic pathways for amino acids, purine nucleotides,
and most cofactors. In apparent compensation, a remarkable number of uncommon and often
duplicated amino acid permeases, peptidases, and phosphotransferase-type transporters were
discovered, suggesting a strong dependency of NCC 533 on the host or other intestinal microbes
to provide simple monomeric nutrients. Genome analysis also predicted an abundance (>12) of
large and unusual cell-surface proteins, including fimbrial subunits, which may be involved in
adhesion to glycoproteins or other components of mucin, a characteristic expected to affect
persistence in the gastrointestinal tract (GIT). Three bile salt hydrolases and two bile acid
transporters, proteins apparently critical for GIT survival, were also detected. In silico
genome comparisons with the >95% complete genome sequence of the closely related Lactobacillus
gasseri revealed extensive synteny punctuated by clear-cut insertions or deletions of single
genes or operons. Many of these regions of difference appear to encode metabolic or structural
components that could affect the organisms competitiveness or interactions with the GIT
ecosystem.

<>

<1>Pridmore, R.D., Mollet, B., Arigoni, F., Hermanns, J.
<2>La1 - the genome of a lactobacillus strain.
<3>International Patent Office
<4>WO 03084989 A
<5>
<6>2003
<7>The present invention pertains to the use of the DNA sequence of a Lactobacillus johnsonii
strain, in particular to its genomic sequence for elucidating interactions of micro-organism
with hosts they colonize, and moreover for elucidating the basis of probiotic properties
exhibited by such strain.  In addition, the present invention also relates to methods of
detecting nucleic acids or polypeptides of Lactobacilli and related species, respectively.  A
data carrier is provided comprising nucleotide sequences and/or polypeptide sequences of La1.

<>

<1>Prieto, J., Epinat, J.C., Redondo, P., Ramos, E., Padro, D., Cedrone, F., Montoya, G., Paques, F., Blanco, F.J.
<2>Generation and analysis of mesophilic variants of the thermostable archaeal I-DmoI homing endonuclease.
<3>J. Biol. Chem.
<4>283
<5>4364-4374
<6>2008
<7>The hyperthermophilic archaeon Desulfurococcus mobilis I-DmoI protein belongs to the family of
proteins known as homing endonucleases (HEs).
HEs are highly specific DNA-cleaving enzymes that recognize long
stretches of DNA and are powerful tools for genome engineering. Because
of its monomeric nature, I-DmoI is an ideal scaffold for generating
mutant enzymes with novel DNA specificities, similarly reported for
homodimeric HEs, but providing single chain endonucleases instead of
dimers. However, this would require the use of a mesophilic variant
cleaving its substrate at temperatures of 37 C and below. We have
generated mesophilic mutants of I-DmoI, using a single round of
directed evolution that relies on a functional assay in yeast. The
effect of mutations identified in the novel proteins has been
investigated. These mutations are located distant to the DNA-binding
site and cause changes in the size and polarity of buried residues,
suggesting that they act by destabilizing the protein. Two of the novel
proteins have been produced and analyzed in vitro. Their overall
structures are similar to that of the parent protein, but they are
destabilized against thermal and chemical denaturation. The
temperature-dependent activity profiles for the mutants shifted toward
lower temperatures with respect to the wild-type activity profile.
However, the most destabilized mutant was not the most active at low
temperatures, suggesting that other effects, like local structural
distortions and/or changes in the protein dynamics, also influence
their activity. These mesophilic I-DmoI mutants form the basis for
generating new variants with tailored DNA specificities.

<>

<1>Prieto, J., Molina, R., Montoya, G.
<2>Molecular scissors for in situ cellular repair.
<3>Crit. Rev. Biochem. Mol. Biol.
<4>47
<5>207-221
<6>2012
<7>The engineering of protein-DNA interactions in different protein scaffolds may provide
'toolkits' to modify the genome. Homing
endonucleases are powerful tools for genome manipulation through
homologous recombination, as these enzymes possess a very low frequency
of DNA cleavage in eukaryotic genomes due to their high specificity.
Therefore, the combination of a precise 'cutter' with the presence of a
natural or modified homologous DNA donor provides a potentially useful
means to modify the genome. However, the basis of protein-DNA
recognition must be understood to generate tailored enzymes that target
the DNA at sites of interest. The engineering of homing endonucleases
and alternative scaffolds, such as zinc fingers or transcription
activator-like effector domains, has demonstrated the potential of
these approaches to create new specific instruments to target genes for
inactivation or repair. Customized homing endonucleases targeting
selected human genes can excise or correct regions of genes implicated
in monogenic diseases, thereby representing important tools for
intervention in eukaryotic genomes.

<>

<1>Prieto, J., Redondo, P., Padro, D., Arnould, S., Epinat, J.C., Paques, F., Blanco, F.J., Montoya, G.
<2>The C-terminal loop of the homing endonuclease I-CreI is essential for site recognition, DNA binding and cleavage1.
<3>Nucleic Acids Res.
<4>35
<5>3262-3271
<6>2007
<7>Meganucleases are sequence-specific endonucleases with large cleavage sites that can be used
to induce efficient homologous gene targeting in
cultured cells and plants. These enzymes open novel perspectives for
genome engineering in a wide range of fields, including gene therapy. A
new crystal structure of the I-CreI dimer without DNA has allowed the
comparison with the DNA-bound protein. The C-terminal loop displays a
different conformation, which suggests its implication in DNA binding. A
site-directed mutagenesis study in this region demonstrates that whereas
the C-terminal helix is negligible for DNA binding, the final C-terminal
loop is essential in DNA binding and cleavage. We have identified two
regions that comprise the Ser138-Lys139 and Lys142-Thr143 pairs whose
double mutation affect DNA binding in vitro and abolish cleavage in vivo.
However, the mutation of only one residue in these sites allows DNA
binding in vitro and cleavage in vivo. These findings demonstrate that the
C-terminal loop of I-CreI endonuclease plays a fundamental role in its
catalytic mechanism and suggest this novel site as a region to take into
account for engineering new endonucleases with tailored specificity.

<>

<1>Prinz, B., Lechner, M., Frey, B., Jarsch, M.
<2>Type II restriction endonuclease SwaI.
<3>US Patent Office
<4>US 5158878
<5>
<6>1992
<7>*
The novel type II restriction endonuclease SwaI has the following recognition sequence:

    5'-ATTT/AAAT-3'
    3'-TAAA/TTTA-5'

and preferably cleaves at the cleavage sites indicated by the line. It is preferably
obtainable from microorganisms of the genus Staphylococcus.


<>

<1>Prinz, B., Lechner, M., Frey, B., Jarsch, M.
<2>Type II Restriction endonuclease SwaI.
<3>European Patent Office
<4>EP 0456086 A, EP 0456086 B
<5>
<6>1995
<7>
 The new Type II restriction endonuclease SwaI possesses the following recognition
 sequence, cleavage favors at the marked cleavage site:
 

     5'-ATTT^AAAT-3'
     3'-TAAA^TTTA-5'
 	

 It is especially obtainable from the microorganism with the genus Staphylococcus.


<>

<1>Prior, J.L., Prior, R.J., Hitchen, P.G., Dell, A., Titball, R.W.
<2>Immunogenic Sequences.
<3>Japanese Patent Office
<4>JP 2005528108 A
<5>
<6>2005
<7>
<>

<1>Pristas, P., Fliegerova, K., Javorsky, P.
<2>Two restriction endonucleases in Selenomonas ruminantium subsp. lactilytica.
<3>Lett. Appl. Microbiol.
<4>27
<5>83-85
<6>1998
<7>Crude protein extract from a recently isolated ruminal bacterium identified as Selenomonas
ruminantium subsp. lactilytica specifically cleaved DNA.  This ability was due to the presence
of two site-specific restriction endonucleases.  SrlI, a NaeI isoschizomer, recognizes the
5'-GCCGGC-3' sequence.  SrlII, an NsiI isoschizomer, recognizes 5'-ATGCAT-3'.

<>

<1>Pristas, P., Godany, A., Sevcikova, B., Oktavcova, B., Farkasovska, J.
<2>Characterization of restriction endonuclease activities in tetracycline producing strains of Streptomyces aureofaciens.
<3>Nucleic Acids Res.
<4>20
<5>4364
<6>1992
<7>We have tested several strains of streptomyces aureofaciens from different sources for
restriction endonuclease. In all tested tetracycline producing strains (B-96, NMU, 16, R8/26)
restriction endonuclease was isolated and characterized (SauLPI, SauNI, SauSI, SauHPI
respectively). Previously described restriction endonuclease from strain S.aureofaciens R8/26I
(1) was renamed for SauHPI in this report. Endonucleases were purified from cell extracts by
phosphocellulose and heparin-sepharose chromatography according to (2) with minor
modifications. The restriction endonucleases were free of contaminating nucleases activity.
All isolated restriction endonucleases produced the same cleavage pattern on tested DNA
substrates. Computer-derived mapping data predict the sequence 5'-GCCGCC-3'. The comparison
of cleavage patterns of pBR322 by S.aureofaciens restrictases to NaeI confirmed the
recognition sequence (Figure 1).

<>

<1>Pristas, P., Javorsky, P.
<2>Restriction-modification systems in ruminal bacteria: occurrence and some evolutionary implications.
<3>Reprod. Nutr. Dev., Elsevier, Chesson, A., Stewart, C.S., Flint, J.H., 
<4>37
<5>74-75
<6>1997
<7>Type II restriction modification systems involve a DNA methyltransferase and an endonuclease
of the same recognition sequence specificity.  It is generally accepted that these systems act
primarily to protect bacteria from foreign DNA, particularly from infection by bacteriophages.
The study of the biology of restriction-modification systems has revealed some general
features and it has been shown that the composition of the bacterial chromosome and the
restriction-modification systems present within cells are evolutionarily linked.  Restriction
endonucleases have been found in bacteria from all taxonomic and ecological groups.  Ruminal
bacteria have been shown to be a promising source of these enzymes.  Up to now ten restriction
endonucleases have been isolated from bacteria of this ecological group.  Our studies on
variability of endonucleolytic activity in S. ruminantium have demonstrated a high frequency
and diversity of restriction endonucleases in this species, and at least ten different
specificities have been characterized.  In addition the observed frequency of restriction
endonucleases, which were present in more than one-third of strains tested, is higher than
observed in bacteria from other ecological groups.  A high frequency of restriction
endonucleases in S. ruminantium can also be inferred from the analysis of DNA.  Using the
method of Karlin et al., it was shown that average counts of perfect 4- and 6- base pairs
palindromes observed within S. ruminantium DNA are lower than in other bacteria, and even
lower than those observed among phage DNAs.  The observed low frequency of short palindromes
is therefore in good agreement with the high frequency of restriction endonucleases observed
in this genus.  Similarly other ruminal bacteria show lower counts of palindromes than would
be predicted from a random distribution.  We suppose that the consistently low frequencies of
4-bp and 6-bp palindromes observed within the DNA of ruminal bacteria is probably a result of
the variety and multiplicity of restriction systems found in bacteria from this ecological
group.  Possibly, there is a correlation between bacterial population density and the
frequency of restriction endonucleases.  Bacterial counts in the rumen are higher than in any
other environment.  Together with the high bacterial counts, there are also unusually high
concentrations of bacteriophages.  If the protection of cells from bacteriophage infection is
a primary role of restriction-modification systems, these systems should be more frequent in
the rumen than in environments with lower populations of bacteriophages.  In such a strongly
competitive ecosystem as the rumen of herbivorous animals the possession of restriction
activity can provide a selective advantage for survival of both the individual bacterial clone
and the species as a whole.

<>

<1>Pristas, P., Molnarova, V., Javorsky, P.
<2>Detection of N6-methyladenine in GATC sequences of Selenomonas ruminantium.
<3>J. Basic Microbiol.
<4>38
<5>283-287
<6>1998
<7>The presence of N6-methyladenine in GATC sequences in DNA of Selenomonas ruminantium was
investigated using sensitive methylation discriminating isoschizomeric restriction enzyme
analysis.  Methylated adenine was detected in 8 out of 18 tested strains belonging to the
subsp. lactilytica of S. ruminantium.  No corresponding restriction activity was detected in
three tested strains.  No GATC methylation was detected in 3 analyzed S. ruminantium subsp.
ruminantium strains.

<>

<1>Pristas, P., Molnarova, V., Javorsky, P.
<2>Restriction and modification systems of ruminal bacteria.
<3>Folia Microbiol. (Praha)
<4>46
<5>71-72
<6>2001
<7>A high frequency of type II restriction endonuclease activities was detected in Selenomonas
ruminantium but not in other rumen bacteria tested. Eight different restriction endonucleases
were characterized in 17 strains coming from genetically homogeneous local populations.
Chromosomal DNA isolated from S. ruminantium strains was found to be refractory to cleavage by
various restriction enzymes, implying the presence of methylase activities additional to those
required for protection against the cellular endonucleases. The presence of Dam methylation
was detected in S. ruminantium strains as well as in several other species belonging to the
Sporomusa sub-branch of low G + C Gram-positive bacteria (Megasphaera elsdenii, Mitsuokella
multiacidus).

<>

<1>Pristas, P., Molnarova, V., Javorsky, P.
<2>Diversity of restriction and modification phenotypes in local population of rumen bacterium Selenomonas ruminantium.
<3>Biologia (Bratisl)
<4>57
<5>777-782
<6>2002
<7>Type II restriction endonuclease activities detected in various strains of Selenomonas
ruminantium species, coming from genetically homogenous
local population, were characterised. The recognition sequence of
Srl55DI was determined to be 5'-G/AATTC-3', identical to that of EcoRI.
Srl5DI restriction endonuclease recognises 5'-CTGCA/G-3' sequence and
is true isoschizomer of PstI. The Srl56DI restriction endonuclease was
found to recognise and cleave 5-C/TRYAG-3' sequence and is true
isoschizomer of SfcI. All other restriction activities characterised
were duplicates of restriction endonucleases, which have previously
been detected in bacteria of S. ruminantium species. In total, 9
different restriction endonucleases were found in the tested bacteria.
Much higher variability of modification phenotypes was observed, when
totally 13 types of modification profiles were found in the studied S.
ruminantium strains. Chromosomal DNA isolated from S. ruminantium
strains was found to be refractive to cleavage by various restriction
enzymes, implying the presence of methylase activities additional to
those required for protection against the cellular endonucleases. While
extremely diverse in restriction and modification phenotypes the tested
strains showed very low genetic diversity, when all but one strain
produced identical profiles in ARDREA analysis. Based on close
similarity of the tested strains the lateral gene transfer is proposed
as a source of the observed diversity of restriction and modification
phenotypes in S. ruminantium.

<>

<1>Pristas, P., Piknova, M.
<2>Underrepresentation of short palindromes in Selenomonas ruminantium DNA: evidence for horizontal gene transfer of restriction and  modification systems?
<3>Can. J. Microbiol.
<4>51
<5>315-318
<6>2005
<7>Molecular analysis of isolates of the rumen bacterium Selenomonas ruminantium revealed a high
variety and frequency of site-specific
(restriction) endonucleases. While all known S. ruminantium restriction
and modification systems recognize hexanucleotide sequences only,
consistently low counts of both 6-bp and 4-bp palindromes were found in
DNA sequences of S. ruminantium. Statistical analysis indicated that
there is some correlation between the degree of underrepresentation of
tetranucleotide words and the number of known restriction endonucleases
for a given sequence. Control analysis showed the same correlation in
lambda DNA but not in human adenovirus DNA. Based on the data
presented, it could be proposed that there is a much higher historical
occurrence of restriction and modification systems in S. ruminantium
and (or) frequent horizontal gene transfer of restriction and
modification gene complexes.

<>

<1>Pristas, P., Vanat, I., Godany, A., Javorsky, P.
<2>Restriction endonucleases from Selenomonas ruminantium which recognize and cleave 5'-AT/TAAT-3'.
<3>Arch. Microbiol.
<4>161
<5>439-441
<6>1994
<7>Two natural isolates from fallow-deer rumen identified as Selenomonas ruminantium were found
to produce a restriction endonuclease which we called Sru4DI. This enzyme was isolated from
cell extracts by phosphocellulose chromatography. Analysis of the Sru4DI recognition site
showed that Sru4DI recognizes the hexanucleotide sequence 5'-AT/TAAT-3' generating 5'
dinucleotide protruding ends upon cleavage and thus is a true isoschizomer of VspI, a
restriction enzyme isolated from Vibrio sp.

<>

<1>Pristas, P., Vanat, I., Javorsky, P.
<2>Isolation and characterization of a new restriction endonuclease, Sru30DI, from Selenomonas ruminantium.
<3>Gene
<4>158
<5>139-140
<6>1995
<7>The restriction endonuclease (ENase) Sru30DI, an isoschizomer of StuI, which recognizes the
sequence 5'-AGG/CCT-3', was purified from a natural isolate of Selenomonas ruminantium. The
ENase was isolated from cell extracts using single-step purification by phosphocellulose
column chromatography. Activity of Sru30DI is inhibited by overlapping Dcm methylation. The
ENase is extremely stable at 37oC and is active over a wide range of pH, temperature and salt
concentrations.

<>

<1>Pristas, P., Vanat, I., Javorsky, P.
<2>Variability of endonucleolytic activity indicates high genetic diversity within the natural population of Selenomonas ruminantium.
<3>Folia Microbiol. (Praha)
<4>42
<5>121-124
<6>1997
<7>The population of bacteria of Selenomonas ruminantium species in the rumen of fallow-deer was
analyzed using endonucleolytic activity assay and plasmid profiles, indicating the presence of
the different specificity nucleases, have been observed.  Site-specific endonucleases were
detected in 17 out of 45 strains tested.  In other strains a various level of non-specific
activity was detected.  Plasmid DNAs ranging in size from 0.9 to more than 25 kbp were
detected in 60% of strains analyzed.  No or little correlation was observed between the
endonuclease activity and the plasmid content.  The presence of different specificity
endonucleases, as well as differences of plasmid profiles of isolates possessing identical
specific activity indicate that the population of S. ruminantium in the rumen of an individual
animal consists of at least 10 different clones.

<>

<1>Pritchard, L., Humphris, S., Baeyen, S., Maes, M., Van Vaerenbergh, J., Elphinstone, J., Saddler, G., Toth, I.
<2>Draft Genome Sequences of Four Dickeya dianthicola and Four Dickeya solani Strains.
<3>Genome Announcements
<4>1
<5>e00087-12
<6>2013
<7>Dickeya dianthicola and 'Dickeya solani' are currently the dominant bacterial pathogens of
potatoes in Europe. Here, we present the draft genome sequences of
four strains of each pathogen.

<>

<1>Pritchard, L., Humphris, S., Saddler, G.S., Elphinstone, J.G., Pirhonen, M., Toth, I.K.
<2>Draft Genome Sequences of 17 Isolates of the Plant Pathogenic Bacterium Dickeya.
<3>Genome Announcements
<4>1
<5>e00978-13
<6>2013
<7>Dickeya (formerly Erwinia chrysanthemi) species cause diseases on a wide range of crops and
ornamental plants worldwide. Here we present the draft sequences of 17
Dickeya isolates spanning four Dickeya species, including five isolates that are
currently unassigned to a species.

<>

<1>Privman, E.
<2>Bioinformatic Identification of Homing Endonucleases and Their Target Sites.
<3>Methods Mol. Biol.
<4>1123
<5>27-35
<6>2014
<7>Homing endonuclease genes (HEGs) are a large, phylogenetically diverse superfamily of enzymes
with high specificity for especially long target sites. The public genomic sequence databases
contain thousands of HEGs. This is a large and diverse arsenal of potential genome editing
tools. To make use of this natural resource, one needs to identify candidate HEGs. Due to
their special relationship with a host gene, it is also possible to predict their cognate
target sequences. Here I describe the HomeBase algorithm that was developed to this end. A
detailed description of the computational pipeline is provided with emphasis on technical and
methodological caveats of the approach.

<>

<1>Proenca, D.N., Espirito, S.C., Grass, G., Morais, P.V.
<2>Draft Genome Sequence of Serratia sp. Strain M24T3, Isolated from Pinewood Disease Nematode Bursaphelenchus xylophilus.
<3>J. Bacteriol.
<4>194
<5>3764
<6>2012
<7>Here we report the draft genome sequence of Serratia sp. strain M24T3, which is associated
with pinewood nematode Bursaphelenchus xylophilus, the causative agent
of pine wilt disease. Serratia sp. strain M24T3 has been identified as a
bionematocide for B. xylophilus in vitro, and multiple genes potentially involved
in virulence and nematotoxity were identified.

<>

<1>Proenca, D.N., Espirito, S.C., Grass, G., Morais, P.V.
<2>Draft Genome Sequence of Pseudomonas sp. Strain M47T1, Carried by Bursaphelenchus xylophilus Isolated from Pinus pinaster.
<3>J. Bacteriol.
<4>194
<5>4789-4790
<6>2012
<7>The draft genome sequence of Pseudomonas sp. strain M47T1, carried by the Bursaphelenchus
xylophilus pinewood nematode, the causative agent of pine wilt
disease, is presented. In Pseudomonas sp. strain M47T1, genes that make this a
plant growth-promoting bacterium, as well as genes potentially involved in
nematotoxicity, were identified.

<>

<1>Proffitt, J.H., Davie, J.R., Swinton, D., Hattman, S.
<2>5-methylcytosine is not detectable in Saccharomyces cerevisiae.
<3>Mol. Cell. Biol.
<4>4
<5>985-988
<6>1984
<7>We examined the DNA of Saccharomyces cerevisiae by both HpaII-MspI restriction enzyme
digestion and high-performance liquid chromatography analysis for the possible presence of
5-methylcytosine.  Both of these methods failed to detect cytosine methylation within this
yeast DNA; i.e. there is <1 5-methylcytosine per 3,100 to 6,000 cytosine residues.

<>

<1>Progulske-Fox, A., Hillman, J.D., Handfield, M.
<2>Identification of porphyromonas gingivalis virulence polynucleotides for diagnosis, treatment, and monitoring of periodontal diseases.
<3>International Patent Office
<4>WO 2005019249 A
<5>
<6>2005
<7>The invention provides compositions and methods for the detection of Porphyromonas gingivalis
and for the treatment and prevention of diseases and infections caused by P. gingivalis.

<>

<1>Progulske-Fox, A., Hillman, J.D., Handfield, M.
<2>Identification of Actinobacillus actinomycetemcomitans Antigens for Use in the Diagnosis, Treatment, and Monitoring of Periodontal Diseases.
<3>Japanese Patent Office
<4>JP 2005531288 A
<5>
<6>2005
<7>
<>

<1>Progulske-Fox, A., Hillman, J.D., Handfield, M.
<2>Identification of Actinobacillus actinomycetemcomitans Antigens for Use in the Diagnosis, Treatment, and Monitoring of Periodontal Diseases.
<3>Japanese Patent Office
<4>JP 2006519586 A
<5>
<6>2006
<7>
<>

<1>Protozanova, E., Demidov, V.V., Nielsen, P.E., Frank-Kamenetskii, M.D.
<2>Pseudocomplementary PNAs as selective modifiers of protein activity on duplex DNA: the case of type IIs restriction enzymes.
<3>Nucleic Acids Res.
<4>31
<5>3929-3935
<6>2003
<7>This study evaluates the potential of pseudocomplementary peptide nucleic acids (pcPNAs) for
sequence-specific modification of enzyme activity
towards double-stranded DNA (dsDNA). To this end, we analyze the ability
of pcPNA-dsDNA complexes to site-selectively interfere with the action of
four type IIs restriction enzymes. We have found that pcPNA-dsDNA
complexes exhibit a different degree of DNA protection against
cleaving/nicking activity of various isoschizomeric endonucleases under
investigation (PleI, MlyI and N.BstNBI) depending on their type and mutual
arrangement of PNA-binding and enzyme recognition/cleavage sites. We have
also found that the pcPNA targeting to closely located PleI or BbsI
recognition sites on dsDNA generates in some cases the nicking activity of
these DNA cutters. At the same time, MlyI endonuclease, a PleI
isoschizomer, does not exhibit any DNA nicking/cleavage activity, being
completely blocked by the nearby pcPNA binding. Our results have general
implications for effective pcPNA interference with the performance of
DNA-processing proteins, thus being important for prospective applications
of pcPNAs.

<>

<1>Protozanova, E., Demidov, V.V., Soldatenkov, V., Chasovskikh, S., Frank-Kamenetskii, M.D.
<2>Tailoring the activity of restriction endonuclease PleI by PNA-induced DNA looping.
<3>EMBO Rep.
<4>3
<5>956-961
<6>2002
<7>DNA looping is one of the key factors allowing proteins bound to different DNA sites to signal
one another via direct contacts. We demonstrate that DNA looping can be generated in an
arbitrary chosen site by sequence-directed targeting of double-stranded DNA with
pseudocomplementary peptide-nucleic acids (pcPNAs). We designed pcPNAs to mask the DNA from
cleavage by type IIs restriction enzyme PleI while not preventing the enzyme from binding to
its primary DNA recognition site. Direct interaction between two protein molecules (one bound
to the original recognition site and the other to a sequence-degenerated site) results in a
totally new activity of PleI: it produces a nick near the degenerate site. The PNA-induced
nicking efficiency varies with the distance between the two protein-binding sites in a phase
with the DNA helical periodicity. Our findings imply a general approach for the fine-tuning of
proteins bound to DNA sites well separated along the DNA chain.

<>

<1>Protsenko, A., Zakharova, M., Nagornykh, M., Solonin, A., Severinov, K.
<2>Transcription regulation of restriction-modification system Ecl18kI.
<3>Nucleic Acids Res.
<4>37
<5>5322-5330
<6>2009
<7>Restriction-modification (R-M) system Ecl18kI is representative of R-M systems whose
coordinated transcription is achieved through a separate
DNA-binding domain of the methyltransferase. M.Ecl18kI recognizes an
operator sequence located in the noncoding region that separates the
divergently transcribed R and M genes. Here we show that, contrary to
previous predictions, the two ecl18kI promoters are not divergent, but
actually face one another. The binding of M.Ecl18kI to its operator
prevents RNA polymerase (RNAP) binding to the M promoter by steric
exclusion, but has no direct effect on RNAP interaction with the R
promoter. The start point for R transcription is located outside of the
intergenic region, opposite the initiation codon of the M gene. Regulated
transcription of the potentially toxic ecl18kI R gene is accomplished (i)
at the stage of promoter complex formation, through direct competition
from complexes formed at the M promoter, and (ii) at the stage of promoter
clearance, since R promoter-bound RNAP escapes the promoter more slowly
than RNAP bound to the M promoter.

<>

<1>Proux, C., Van Sinderen, D., Suarez, J., Garcia, P., Ladero, V., Fitzgerald, G.F., Desiere, F., Brussow, H.
<2>The dilemma of phage taxonomy illustrated by comparative genomics of sfi21-like siphoviridae in lactic acid bacteria.
<3>J. Bacteriol.
<4>184
<5>6026-6036
<6>2002
<7>The complete genome sequences of two dairy phages, Streptococcus thermophilus phage 7201 and
Lactobacillus casei phage A2, are reported. Comparative genomics reveals that both phages are
members of the recently proposed Sfi21-like genus of Siphoviridae, a widely distributed phage
type in low-GC-content gram-positive bacteria. Graded relatedness, the hallmark of evolving
biological systems, was observed when different Sfi21-like phages were compared. Across the
structural module, the graded relatedness was represented by a high level of DNA sequence
similarity or protein sequence similarity, or a shared gene map in the absence of sequence
relatedness. This varying range of relatedness was found within Sfi21-like phages from a
single species as demonstrated by the different prophages harbored by Lactococcus lactis
strain IL1403. A systematic dot plot analysis with 11 complete L. lactis phage genome
sequences revealed a clear separation of all temperate phages from two classes of virulent
phages. The temperate lactococcal phages share DNA sequence homology in a patchwise fashion
over the nonstructural gene cluster. With respect to structural genes, four DNA homology
groups could be defined within temperate L. lactis phages. Closely related structural modules
for all four DNA homology groups were detected in phages from Streptococcus or Listeria,
suggesting that they represent distinct evolutionary lineages that have not uniquely evolved
in L. lactis. It seems reasonable to base phage taxonomy on data from comparative genomics.
However, the peculiar modular nature of phage evolution creates ambiguities in the definition
of phage taxa by comparative genomics. For example, depending on the module on which the
classification is based, temperate lactococcal phages can be classified as a single phage
species, as four distinct phage species, or as two if not three different phage genera. We
propose to base phage taxonomy on comparative genomics of a single structural gene module
(head or tail genes). This partially phylogeny-based taxonomical system still mirrors some
aspects of the current International Committee on Taxonomy in Virology classification system.
In this system the currently sequenced lactococcal phages would be grouped into five genera:
c2-, sk1, Sfi11-, r1t-, and Sfi21-like phages.

<>

<1>Prozorov, A.A., Belova, T.S., Surikov, N.N.
<2>Transformation and transduction of Bacillus subtilis strains with the BsuR restriction-modification system by means of modified and unmodified DNA of pUB110 plasmid.
<3>Mol. Gen. Genet.
<4>180
<5>135-138
<6>1980
<7>During transformation of B. subtilis cells with the BsuR restriction-modification system by
means of pUB110 plasmid, restriction and modification of the plasmid DNA occurs.  The effect
of restriction on the transformation frequency is relatively weak, bringing about a 20-fold
decrease only.  When using cells of a modifying recipient, the frequency of AR9 phage-mediated
transduction of unmodified plasmid DNA is also relatively little decreased.  The frequency of
transduction by chromosomal markers, under the same conditions, falls much lower.

<>

<1>Pryshliak, M., Hammerl, J.A., Reetz, J., Strauch, E., Hertwig, S.
<2>Vibrio vulnificus Phage PV94 Is Closely Related to Temperate Phages of V. cholerae and Other Vibrio Species.
<3>PLoS ONE
<4>9
<5>E94707
<6>2014
<7>BACKGROUND: Vibrio vulnificus is an important pathogen which can cause serious
infections in humans. Yet, there is limited knowledge on its virulence factors
and the question whether temperate phages might be involved in pathogenicity, as
is the case with V. cholerae. Thus far, only two phages (SSP002 and VvAW1)
infecting V. vulnificus have been genetically characterized. These phages were
isolated from the environment and are not related to Vibrio cholerae phages. The
lack of information on temperate V. vulnificus phages prompted us to isolate
those phages from lysogenic strains and to compare them with phages of other
Vibrio species. RESULTS: In this study the temperate phage PV94 was isolated from
a V. vulnificus biotype 1 strain by mitomycin C induction. PV94 is a myovirus
whose genome is a linear double-stranded DNA of 33,828 bp with 5'-protruding
ends. Sequence analysis of PV94 revealed a modular organization of the genome.
The left half of the genome comprising the immunity region and genes for the
integrase, terminase and replication proteins shows similarites to V. cholerae
kappa phages whereas the right half containing genes for structural proteins is
closely related to a prophage residing in V. furnissii NCTC 11218. CONCLUSION: We
present the first genomic sequence of a temperate phage isolated from a human V.
vulnificus isolate. The sequence analysis of the PV94 genome demonstrates the
wide distribution of closely related prophages in various Vibrio species.
Moreover, the mosaicism of the PV94 genome indicates a high degree of horizontal
genetic exchange within the genus Vibrio, by which V. vulnificus might acquire
virulence-associated genes from other species.

<>

<1>Ptashne, M.
<2>Replication and host modification of DNA transferred during bacterial mating.
<3>J. Mol. Biol.
<4>11
<5>829-838
<6>1965
<7>An experiment is presented which shows that during the course of bacterial
mating between Escherichia coli F-prime (lambda) male and an E. coli (lambda)
female, the transferred lambda prophages all replicate.  This strongly suggests
that all the transferred F-prime factors carrying the lambda prophage replicate
during the mating.  A separate experiment, employing host-induced modification,
shows that at least 20% of the strands injected into the female are synthesized
in the male just prior to injection.  It is argued that all the injected DNA
replicates in the male just prior to injection, and that most of the newly made
strands are not endowed with the host-induced modification properties normally
associated with DNA synthesized in the male.

<>

<1>Pu, A.T., Radany, E.H.
<2>Regulated expression of restriction endonuclease activity in mammalian cells.
<3>J. Cell. Biochem.
<4>S21
<5>327
<6>1995
<7>DNA double-strand breaks (dsbs) are the primary lethal lesion in cells exposed to ionizing
radiation (IR).  Detecting cellular responses specific to the dsbs formed by IR is hampered by
a broad spectrum of damage.  Transfer of restriction enzymes (REs) into mammalian cells
affords access to dsbs in the absence of other lesions; however, this approach is not
quantitatively radiomimetic due to the variable introduction of REs into cells in quantities
ranging from none to much greater than average.  While this effect might be eliminated by
regulated, homogeneous RE gene expression in mammalian cells, such a strategy has been limited
by leaky transcriptional control.  Braselman et al. reported stringent regulation by estrogen
of a chimeric transcription factor::estrogen receptor (ER) fusion protein activity in
mammalian cells in an effort to create a radiomimetic dsb model system tightly controlled by
estrogen.  PCR was used to modify a RE gene (PvuII) to allow efficient translation in
eucaryotic cells; expression of the modified gene in E. coli verified function by phage
restriction.  This gene was installed in a mammalian expression system expected to afford
estrogen-dependent regulation; the resulting construct has been transferred into rodent cells.
Recovery of robust G418-resistant stable transfectants at similar frequencies of RE
gene-containing and control constructs (in the absence of estrogen) indicates that any basal
PvuII expression in the former case occurs at a level tolerated by the cells.  Preliminary
results suggest cytotoxicity associated with estrogenic steroid exposure in the former, but
not the latter, cells.  The dependence of killing on estrogen exposure duration is currently
under investigation, as is the ability of putative RE-induced dsb to function as sublethal
damage with respect to subsequent IR exposure.  Direct measurement of dsb yields will be
performed using pulse field gels.  The PvuII gene has been engineered for regulated mammalian
cell expression.  This system apparently affords tight control of intracellular RE activity by
estrogen, providing a means to study quantitative DNA dsb effects in the absence of other
changes generated in X-irradiated cells.  Current results will be presented.

<>

<1>Pucciarelli, M.G., Prieto, A.I., Casadesus, J., Portillo, F.G.D.
<2>Envelope instability in DNA adenine methylase mutants of Salmonella  enterica.
<3>Microbiology
<4>148
<5>1171-1182
<6>2002
<7>Mutants of Salmonella enterica serovar Typhimurium lacking DNA
adenine (Dam) methylase show reduced secretion of invasion effectors
encoded in the Salmonella-pathogenicity island 1 (SPI-1). Concomitant with
this alteration, a high number and quantity of extracellular proteins are
detected in cultures of Dam(-) mutants. This study shows by subcellular
fractionation analysis that the presence of numerous extracellular proteins
in cultures of Dam(-) mutants is linked to an exacerbated release of
membrane particulate material. The membrane 'leaky' phenotype and the
impaired functionality of type III secretion systems were, however,
unrelated since exacerbated release of proteins to the medium was evident
in Dam(-) strains carrying mutations in either SPI-1 (invA, invJ) or
flagellar (flhD) genes. This result supports the view that Dam methylation
controls a plethora of cellular processes. Electron microscopy analysis
demonstrated that the accumulation of membrane particulate material occurs
preferentially as vesicles in stationary cultures of Dam(-) strains. In
addition, a reduction in the relative amount of peptidoglycan-associated
lipoprotein (PAL), TolB, OmpA and murein lipoprotein (Lpp) bound to
peptidoglycan was observed in actively growing Dam(-) mutants. The
existence of an envelope defect was further confirmed by the increased
sensitivity to deoxycholate exhibited by Dam(-) mutants, mostly during
exponential growth. Unexpectedly, lack of Dam methylation neither increased
envelope instability nor impaired the association of PAL-Tol-Lpp proteins
to the peptidoglycan in Escherichia coli. Accordingly, E. coli Dam(-)
mutants did not show sensitivity to deoxycholate. Altogether, these results
indicate that, besides its role in modulating the secretion of effectors by
the SPI-1-encoded type III apparatus, Dam methylation controls cell
envelope integrity in S. enterica.

<>

<1>Puchkova, L.I., Andreeva, I.S., Repin, V.E., Krivopalova, G.N.
<2>Bacterial Streptomyces species ST-12 produces a restriction endonuclease recognizing the sequence 5' CTGCAG 3'.
<3>Russian Patent Office
<4>RU 2135581 C
<5>
<6>1999
<7>
<>

<1>Puchkova, L.I., Andreeva, I.S., Saranina, I.V., Pechurkina, N.I., Afonina, V.S., Repin, V.E.
<2>Strain of Paenibacillus sp, producer of restriction endonuclease Psp 1009I.
<3>Russian Patent Office
<4>RU 2340663 C
<5>
<6>2008
<7>New strain Paenibacillus sp. has been obtained, which produces site specific restriction
endonuclease Psp 1009I, capable of recognising and splitting the sequence of nucleotides
5'-GCCNNNNNGGC-3'.  The strain may be used for the production of the highly active
endonuclease Psp 1009I.  New connections contain useful biological properties.

<>

<1>Puchkova, L.I., Andreeva, I.S., Ushakova, T.A., Radionenko, Y.V., Repin, V.E., Polushkina, A.F.
<2>Strain of Bacillus stearothermophilus 22 as producer of site-specific restriction endonuclease recognizing and cleaving nucleotide sequence  5'-CCNNNNNNNGG-3' and eliciting growth-stimulating activity with  respect to cultural plant seeds.
<3>Russian Patent Office
<4>RU 2238972 C
<5>
<6>2004
<7>
<>

<1>Puchkova, L.I., Kalmykova, G.V., Burtseva, L.I., Repin, V.E.
<2>Entomapathogenic bacteria Bacillus thuringiensis as producers of restriction endonucleases.
<3>Prikl. Biokhim. Mikrobiol.
<4>38
<5>140-144
<6>2002
<7>A total of 115 collection strains of Bacillus thuringiensis, belonging to various subspecies,
have been studied for the presence of DNA restriction-modification systems.  Restriction
endonucleases of 13 strains have been isolated and characterized.  No considerable
correlations between the taxonomic positions of the bacteria and the specificities of the
endonucleases isolated have been detected.  It is concluded that the enzymes with identical
specificities are present in both the crystalliferous and acrystalliferous strains of the same
subspecies.

<>

<1>Puchkova, L.I., Krivopalova, G.N., Andreeva, I.S., Selina, A.V., Serov, G.D., Rechkunova, N.I., Degtyarev, S.K.
<2>Streptomyces fradiae is the producer of the restriction endonuclease Sfr274I.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>1
<5>32-34
<6>1990
<7>Experiments were carried out to assay for restrictase producers in
microorganisms from the Streptomyces genus.  Streptomyces fradiae Ac 149 was
found to be a producer of the restriction endonuclease Sfr274I - an
isoschizomer of the restrictase XhoI (recognition site CTCGAG).

<>

<1>Puchkova, L.I., Mikhailova, V.K.
<2>Method of purification of restriction endonuclease HinfI including thermal treatment, chromatography on DEAE cellulose DE-52 using NaCl buffer followed by chromatography on phosphocellulose P-11 and heparin Sepharose.
<3>Russian Patent Office
<4>RU 2136749 C
<5>
<6>1999
<7>
<>

<1>Puchkova, L.I., Radionenko, Y.V., Ushakova, T.A., Repin, V.E.
<2>Sphingobacterium strain Mizutae-32 as producer of SpmI restriction endonuclease, recognizing and cleaving 5'-AT'CGAT-3' nucleotide sequence.
<3>Russian Patent Office
<4>RU 2266323 C
<5>
<6>2005
<7>Invention relates to Bacterium strain sphingobacterium Mizutae-32 producing restriction
endonuclease named as SpmI, which recognizes and cleaves 5'-AT'CGAT-3' nucleotide sequence.
Strain is isolated from melt water in process of searching endonuclease producing strain with
high activity at low temperatures.  Strain is capable of growth at 6oC, is stable for two week
storage at 20oC.  Strain-produced enzyme hydrolyzes phage-lambda DNA and T7 at 6-10oC.  Enzyme
yield is 300000 U/10g of cells.  Effective strain for restriction endonuclease production.

<>

<1>Puchkova, L.I., Serov, G.D.
<2>Method of producing restriction enzymes.
<3>Soviet Patent Office
<4>SU 1406159
<5>
<6>1988
<7>
<>

<1>Puchkova, L.I., Totmenina, O.D., Afonina, V.S., Polushkina, A.F., Repin, V.E.
<2>Arthrobacter strain Mn372 producing Asi3721 restriction endonuclease that recognizes and cleaves the 5'-ATGCA-3' nucleotide sequence.
<3>Russian Patent Office
<4>RU 2302459 C
<5>
<6>2007
<7>
<>

<1>Puchkova, L.I., Ushakova, T.A., Mikhailova, V.K., Serov, G.D., Krivopalova, G.N., Repin, V.E.
<2>Testing and isolation of high-purity restriction endonucleases.
<3>Prikl. Biokhim. Mikrobiol.
<4>38
<5>20-24
<6>2002
<7>A new method of testing restriction nucleases is proposed.  This method is based on
high-temperature treatment of crude cell extracts.  Disrupted cells were heated at 50-60 C,
centrifuged, and assayed for restrictases.  This method provides the opportunity for screening
new enzymes in microbial strains enriched with nonspecific restrictases.  High-temperature
treatment of cell extracts of certain producers reduces the number of steps of the procedure
used for isolating high-purity restrictases; the resulting preparations are capable of
maintaining high enzymatic activity during long-term storage.  It was shown that
high-temperature treatment can be applied not only to thermophilic but also to mesophilic
strains of microorganisms of different taxa.

<>

<1>Puchta, H., Dujon, B., Hohn, B.
<2>Homologous recombination in plant cells in enhanced by in vivo induction of double strand breaks into DNA by a site-specific endonuclease.
<3>Nucleic Acids Res.
<4>21
<5>5034-5040
<6>1993
<7>Induction of double strand breaks (DSBs) is coupled to meiotic and mitotic recombination in
yeast. We show that also in a higher eukaryote induction of DSBs is directly correlated with a
strong enhancement of recombination frequencies. We cotransfected Nicotiana plumbaginifolia
protopasts with a plasmid carrying a synthetic I-SceI gene, coding for a highly sequence
specific endonuclease, together with recombination substrates carrying an I-SceI-site adjacent
to their homologous sequences. We measured efficiencies of extrachromosomal recombination,
using a well established transient B-glucuronidase (GUS) assay. GUS enzyme activities were
strongly increased when a plasmid carrying the I-SceI gene in sense but not in antisense
orientation with respect to the promoter was included in the transfections. The in vivo
induced DSBs were detected in the recombination substrates by Southern blotting, demonstrating
that the yeast enzyme is functional in plant cells. At high ratios of transfected I-SceI-genes
to I-SceI-sites the majority of the I-SceI-sites in the recombination substrates are cleaved,
indicating that the induction of the DSBs is the rate limiting step in the described
recombination reaction. These results imply that in vivo induction of transient breaks at
specific sites in the plant genome could allow foreign DNA to be targeted to these sites via
homologous recombination.

<>

<1>Pues, H., Bleimling, N., Holz, B., Wolcke, J., Weinhold, E.
<2>Functional roles of the conserved aromatic amino acid residues at position 108 (Motif IV) and position 196 (Motif VIII) in base flipping and catalysis by the N6-adenine DNA methyltransferase from Thermus aquaticus.
<3>Biochemistry
<4>38
<5>1426-1434
<6>1999
<7>The DNA methyltransferase from Thermus aquaticus catalyzes the transfer of the activated
methyl group of S-adenosyl-L-methionine to the N6 position of adenine within the
double-stranded DNA sequence 5'-TCGA-3'.  To achieve catalysis M.TaqI flips the target
adenine out of the DNA helix.  On the basis of the three-dimensional structure of M.TaqI in
complex with the cofactor and its structural homology to the C5-cytosine DNA Mtase from
Haemophilus haemolyticus, Tyr108 and Phe196 were suggested to interact with the extrahelical
adenine.  The functional roles of these two aromatic amino acid residues in M.TaqI were
investigated by mutational analysis.  The obtained mutant Mtases were analyzed in an improved
kinetic assay, and their ability to flip the target base was studied in a fluorescence-based
assay using a duplex oligodeoxynucleotide containing the fluorescent base analogue
2-aminopurine at the target position.  While the mutant Mtases containing the aromatic amino
acid Trp at position 108 or 196 (Y108W and F196W) showed almost wild-type catalytic activity,
the mutant Mtases with the nonaromatic amino acid Ala (Y108A and F196A) had a strongly reduced
catalytic constant.  Y108A was still able to flip the target base, whereas F196A was strongly
impaired in base flipping.  These results indicate that Phe196 is important for stabilizing
the extrahelical target adenine and suggest that Tyr108 is involved in placing the
extrahelical target base in an optimal position for methyl group transfer.  Since both
aromatic amino acids belong to the conserved motifs IV and XIII found in N6-adenine and
N4-cytosine DNA Mtases as well as in N6-adenine RNA Mtases, a similar function of aromatic
amino acid residues within these motifs is expected for the different Mtases.

<>

<1>Pugatsch, T., Weber, H.
<2>A thermostable, sequence-specific restriction endonuclease from Bacillus stearothermophilus: BstPI.
<3>Nucleic Acids Res.
<4>7
<5>1429-1444
<6>1979
<7>A restriction endonuclease, BstPI, was purified from a strain of B.
stearothermophilus, and its cleavage specificity was determined.  The enzyme
cleaves at palindromic sites of the general structure: 5' -G^-G-T-N-A-C-C- 3'
3' -C-C-A-N-T-G^-G- 5' where N-N' can be any base pair.  It produces
phosphorylated 5'-termini which are single stranded over a length of 5
nucleotides.  Ends generated by cleavage with BstPI can be rejoined by DNA
ligase.

<>

<1>Puiu, D., Salzberg, S.L.
<2>Re-assembly of the genome of Francisella tularensis subsp. holarctica OSU18.
<3>PLoS ONE
<4>3
<5>E3427
<6>2008
<7>Francisella tularensis is a highly infectious human intracellular pathogen
that is the causative agent of tularemia. It occurs in several major
subtypes, including the live vaccine strain holarctica (type B). F.
tularensis is classified as category A biodefense agent in part because a
relatively small number of organisms can cause severe illness. Three
complete genomes of subspecies holarctica have been sequenced and
deposited in public archives, of which OSU18 was the first and the only
strain for which a scientific publication has appeared. We re-assembled
the OSU18 strain using both de novo and comparative assembly techniques,
and found that the published sequence has two large inversion
mis-assemblies. We generated a corrected assembly of the entire genome
along with detailed information on the placement of individual reads
within the assembly. This assembly will provide a more accurate basis for
future comparative studies of this pathogen.

<>

<1>Pukall, R. et al.
<2>Complete genome sequence of Deinococcus maricopensis type strain (LB-34).
<3>Standards in Genomic Sciences
<4>4
<5>163-172
<6>2011
<7>Deinococcus maricopensis (Rainey and da Costa 2005) is a member of the genus Deinococcus,
which is comprised of 44 validly named species and is located within
the deeply branching bacterial phylum Deinococcus-Thermus. Strain LB-34(T) was
isolated from a soil sample from the Sonoran Desert in Arizona. Various species
of the genus Deinococcus are characterized by extreme radiation resistance, with
D. maricopensis being resistant in excess of 10 kGy. Even though the genomes of
three Deinococcus species, D. radiodurans, D. geothermalis and D. deserti, have
already been published, no special physiological characteristic is currently
known that is unique to this group. It is therefore of special interest to
analyze the genomes of additional species of the genus Deinococcus to better
understand how these species adapted to gamma- or UV ionizing-radiation. The
3,498,530 bp long genome of D. maricopensis with its 3,301 protein-coding and 66
RNA genes consists of one circular chromosome and is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Pukall, R. et al.
<2>Complete genome sequence of Jonesia denitrificans type strain (Prevot 55134).
<3>Standards in Genomic Sciences
<4>1
<5>262-269
<6>2009
<7>Jonesia denitrificans (Prevot 1961) Rocourt et al. 1987 is the type species of the genus
Jonesia, and is of phylogenetic interest because of its isolated
location in the actinobacterial suborder Micrococcineae. J. denitrificans is
characterized by a typical coryneform morphology and is able to form irregular
nonsporulating rods showing branched and club-like forms. Coccoid cells occur in
older cultures. J. denitrificans is classified as a pathogenic organism for
animals (vertebrates). The type strain whose genome is described here was
originally isolated from cooked ox blood. Here we describe the features of this
organism, together with the complete genome sequence and annotation. This is the
first completed genome sequence of a member of the genus for which a complete
genome sequence is described. The 2,749,646 bp long genome with its 2558
protein-coding and 71 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Pukall, R. et al.
<2>Complete genome sequence of Slackia heliotrinireducens type strain (RHS 1).
<3>Standards in Genomic Sciences
<4>1
<5>234-241
<6>2009
<7>Slackia heliotrinireducens (Lanigan 1983) Wade et al. 1999 is of phylogenetic interest because
of its location in a genomically yet uncharted section of the
family Coriobacteriaceae, within the deep branching Actinobacteria. Strain RHS
1(T) was originally isolated from the ruminal flora of a sheep. It is a
proteolytic anaerobic coccus, able to reductively cleave pyrrolizidine alkaloids.
Here we describe the features of this organism, together with the complete genome
sequence, and annotation. This is the first complete genome sequence of the genus
Slackia, and the 3,165,038 bp long single replicon genome with its 2798
protein-coding and 60 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Pukall, R. et al.
<2>Complete genome sequence of Conexibacter woesei type strain (ID131577).
<3>Standards in Genomic Sciences
<4>2
<5>212-219
<6>2010
<7>The genus Conexibacter (Monciardini et al. 2003) represents the type genus of the family
Conexibacteraceae (Stackebrandt 2005, emend. Zhi et al. 2009) with
Conexibacter woesei as the type species of the genus. C. woesei is a
representative of a deep evolutionary line of descent within the class
Actinobacteria. Strain ID131577(T) was originally isolated from temperate forest
soil in Gerenzano (Italy). Cells are small, short rods that are motile by
peritrichous flagella. They may form aggregates after a longer period of growth
and, then as a typical characteristic, an undulate structure is formed by
self-aggregation of flagella with entangled bacterial cells. Here we describe the
features of the organism, together with the complete sequence and annotation. The
6,359,369 bp long genome of C. woesei contains 5,950 protein-coding and 48 RNA
genes and is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Pukall, R. et al.
<2>Complete genome sequence of Kribbella flavida type strain (IFO 14399).
<3>Standards in Genomic Sciences
<4>2
<5>186-193
<6>2010
<7>The genus Kribbella consists of 15 species, with Kribbella flavida (Park et al. 1999) as the
type species. The name Kribbella was formed from the acronym of the
Korea Research Institute of Bioscience and Biotechnology, KRIBB. Strains of the
various Kribbella species were originally isolated from soil, potato, alum slate
mine, patinas of catacombs or from horse racecourses. Here we describe the
features of K. flavida together with the complete genome sequence and annotation.
In addition to the 5.3 Mbp genome of Nocardioides sp. JS614, this is only the
second completed genome sequence of the family Nocardioidaceae. The 7,579,488 bp
long genome with its 7,086 protein-coding and 60 RNA genes and is part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Pukkila, P.J., Peterson, J., Herman, G., Modrich, P., Meselson, M.
<2>Effects of high levels of DNA adenine methylation on methyl-directed mismatch repair in Escherichia coli.
<3>Genetics
<4>104
<5>571-582
<6>1983
<7>Two methods were used in an attempt to increase the efficiency and strand
selectivity of methyl-directed mismatch repair of bacteriophage lambda
heteroduplexes in E. coli.  Previous studies of such repair used lambda DNA
that was only partially methylated as the source of methylated chains.  Also,
transfection was carried out in methylating strains.  Either of these factors
might have been responsible for the incompleteness of the strand selectivity
observed previously.  In the first approach to increasing strand selectivity,
heteroduplexes were transfected into a host deficient in methylation, but no
changes in repair frequencies were observed.  In the second approach,
heteroduplexes were prepared using DNA that had been highly methylated in vitro
with purified DNA adenine methylase as the source of methylated chains.  In
heteroduplexes having a repairable cI/+ mismatch, strand selectivity was indeed
enhanced.  In heteroduplexes with one chain highly methylated and the
complementary chain unmethylated, the frequency of repair on the unmethylated
chain increased to nearly 100%.  Heteroduplexes with both chains highly
methylated were not repaired at a detectable frequency.  Thus, chains highly
methylated by DNA adenine methylase were refractory to mismatch repair by this
system, regardless of the methylation of the complementary chain.  These
results support the hypothesis that methyl-directed mismatch repair acts to
correct errors of replication, thus lowering the mutation rate.

<>

<1>Pullan, S.T., Chandra, G., Bibb, M.J., Merrick, M.
<2>Genome-wide analysis of the role of GlnR in Streptomyces venezuelae provides new insights into global nitrogen regulation in actinomycetes.
<3>BMC Genomics
<4>12
<5>175
<6>2011
<7>ABSTRACT: BACKGROUND: GlnR is an atypical response regulator found in
actinomycetes that modulates the transcription of genes in response to
changes in nitrogen availability. We applied a global in vivo approach to
identify the GlnR regulon of Streptomyces venezuelae, which, unlike many
actinomycetes, grows in a diffuse manner that is suitable for
physiological studies. Conditions were defined that facilitated analysis
of GlnR-dependent induction of gene expression in response to rapid
nitrogen starvation. Microarray analysis identified global transcriptional
differences between glnR+ and glnR mutant strains under varying nitrogen
conditions. To differentiate between direct and indirect regulatory
effects of GlnR, chromatin immuno-precipitation (ChIP) using antibodies
specific to a FLAG-tagged GlnR protein, coupled with microarray analysis
(ChIP-chip), was used to identify GlnR binding sites throughout the S.
venezuelae genome. RESULTS: GlnR bound to its target sites in both
transcriptionally active and apparently inactive forms. Thirty-six GlnR
binding sites were identified by ChIP-chip analysis allowing derivation of
a consensus GlnR-binding site for S. venezuelae. GlnR-binding regions were
associated with genes involved in primary nitrogen metabolism, secondary
metabolism, the synthesis of catabolic enzymes and a number of
transport-related functions. CONCLUSIONS: The GlnR regulon of S.
venezuelae is extensive and impacts on many facets of the organism's
biology. GlnR can apparently bind to its target sites in both
transcriptionally active and inactive forms.

<>

<1>Pullinger, G.D., Bevir, T., Lax, A.J.
<2>The Pasteurella multocida toxin is encoded within a lysogenic bacteriophage.
<3>Mol. Microbiol.
<4>51
<5>255-269
<6>2004
<7>Toxigenic strains of Pasteurella multocida produce a 146 kDa toxin (PMT)
that acts as a potent mitogen. Sequence analysis of the structural gene
for PMT, toxA, previously suggested it was horizontally acquired, because
it had a low G + C content relative to the P. multocida genome. To address
this, the sequence of DNA flanking toxA was determined. The sequence
analysis showed the presence of homologues to bacteriophage tail protein
genes and a bacteriophage antirepressor, suggesting that the toxin gene
resides within a prophage. In addition to phage genes, the toxA flanking
DNA contained a homologue of a restriction/modification system that was
shown to be functional. The presence of a bacteriophage was demonstrated
in spent medium from toxigenic P. multocida isolates. Its production was
increased by mitomycin C addition, a treatment that is known to induce the
lytic cycle of many temperate bacteriophages. The genomes of
bacteriophages from three different toxigenic P. multocida strains had
similar but not identical restriction profiles, and were approximately
45-50 kb in length. The prophages from two of these had integrated at the
same site in the chromosome, in a tRNA gene. Southern blot analysis
confirmed that these bacteriophages contained the toxA gene.

<>

<1>Pullinger, G.D., Dziva, F., Charleston, B., Wallis, T.S., Stevens, M.P.
<2>Identification of Salmonella enterica serovar Dublin-specific sequences by subtractive hybridization and analysis of their role in intestinal colonization and systemic translocation in cattle.
<3>Infect. Immun.
<4>76
<5>5310-5321
<6>2008
<7>Salmonella enterica serovar Dublin is a host-restricted serovar associated
with typhoidal disease in cattle. In contrast, the fowl-associated serovar
S. enterica serovar Gallinarum is avirulent in calves, yet it invades
ileal mucosa and induces enteritis at levels comparable to those induced
by S. enterica serovar Dublin. Suppression subtractive hybridization was
employed to identify S. enterica serovar Dublin strain SD3246 genes absent
from S. enterica serovar Gallinarum strain SG9. Forty-one S. enterica
serovar Dublin fragments were cloned and sequenced. Among these, 24
mobile-element-associated genes were identified, and 12 clones exhibited
similarity with sequences of known or predicted function in other
serovars. Three S. enterica serovar Dublin-specific regions were
homologous to regions from the genome of Enterobacter sp. strain 638.
Sequencing of fragments adjacent to these three sequences revealed the
presence of a 21-kb genomic island, designated S. enterica serovar Dublin
island 1 (SDI-1). PCR analysis and Southern blotting showed that SDI-1 is
highly conserved within S. enterica serovar Dublin isolates but rarely
found in other serovars. To probe the role of genes identified by
subtractive hybridization in vivo, 24 signature-tagged S. enterica serovar
Dublin SD3246 mutants lacking loci not present in Salmonella serovar
Gallinarum SG9 were created and screened by oral challenge of cattle.
Though attenuation of tagged SG9 and SD3246 Salmonella pathogenicity
island-1 (SPI-1) and SPI-2 mutant strains was detected, no obvious defects
of these 24 mutants were detected. Subsequently, a DeltaSDI-1 mutant was
found to exhibit weak but significant attenuation compared with the parent
strain in coinfection of calves. SDI-1 mutation did not impair invasion,
intramacrophage survival, or virulence in mice, implying that SDI-1 does
not influence fitness per se and may act in a host-specific manner.

<>

<1>Puopolo, G., Sonego, P., Engelen, K., Pertot, I.
<2>Draft Genome Sequence of Lysobacter capsici AZ78, a Bacterium Antagonistic to Plant-Pathogenic Oomycetes.
<3>Genome Announcements
<4>2
<5>e00325-14
<6>2014
<7>Lysobacter capsici AZ78, isolated from tobacco rhizosphere, effectively controls  Phytophthora
infestans and Plasmopara viticola on tomato and grapevine plants, respectively. We report the
first draft genome sequence of the L. capsici species.

<>

<1>Pupo, E., Perez, E., Trujillo, L.E., Miranda, F., Gonzalez, E., Brito, J.
<2>Validation of radioactive methods in the quality control of DNA restriction enzymes.
<3>Biotecnol. Apl.
<4>13
<5>197-200
<6>1996
<7>In this paper two radioactive substrates obtained from lambda DNA digested with the
restriction enzyme HpaII were evaluated for the detection of 5' to 3', 3' to 5' single and
double stranded-DNA dependent exonuclease and phosphatase activities found in DNA restriction
and modifying enzyme preparations.  A cloning simulation assay was performed using the same
conditions established for the radioactive assay taking into account enzyme units and pmols of
DNA ends used as substrate.  As a result, it was found that for degradation of the radioactive
DNA substrate per enzyme unit below 0.5%, the false positives in the cloning simulation assay
became less than 5%.  Finally, the use of the radiolabeled [gamma 32P] ATP lambda HpaII DNA
substrate to detect 5' to 3' single stranded-DNA dependent exonuclease and phosphatase
contaminating activities is described at certain critical steps of the purification process of
the restriction enzyme KpnI.

<>

<1>Puranik, S., Talkal, R., Qureshi, A., Khardenavis, A., Kapley, A., Purohit, H.J.
<2>Genome Sequence of the Pigment-Producing Bacterium Pseudogulbenkiania ferrooxidans, Isolated from Loktak Lake.
<3>Genome Announcements
<4>1
<5>e01115-13
<6>2013
<7>The whole genome of a pigment-producing isolate from a lake in northern India,
Pseudogulbenkiania ferrooxidans strain EGD-HP2, has been sequenced to study the
spectrum of biosynthesis of secondary metabolites. The genome annotation data
revealed an operon for violacein, which showed homology with the reported operon
of a Chromobacterium sp., and also a quinone cofactor.

<>

<1>Purdy, D., O'Keeffe, T.A., Elmore, M., Herbert, M., McLeod, A., Bokori-Brown, M., Ostrowski, A., Minton, N.P.
<2>Conjugative transfer of clostridial shuttle vectors from Escherichia coli to Clostridium difficile through circumvention of the restriction  barrier.
<3>Mol. Microbiol.
<4>46
<5>439-452
<6>2002
<7>Progress towards understanding the molecular basis of virulence in Clostridium difficile has
been hindered by the lack of effective gene transfer systems. We have now, for the first time,
developed procedures that may be used to introduce autonomously replicating vectors into this
organism through their conjugative, oriT-based mobilization from Escherichia coli donors.
Successful transfer was achieved through the use of a plasmid replicon isolated from an
indigenous C. difficile plasmid, pCD6, and through the characterization and subsequent
circumvention of host restriction/modification (RM) systems. The characterized replicon is the
first C. difficile plasmid replicon to be sequenced and encodes a large replication protein
(RepA) and a repetitive region composed of a 35 bp iteron sequence repeated seven times.
Strain CD6 has two RM systems, CdiCD6I/M.CdiCD6I and CdiCD6II/M.CdiCD6II, with equivalent
specificities to Sau96I/M. Sau96I (5'-GGNmCC-3') and Mbol/M. Mbol (5'-GmATC-3')
respectively. A second strain (CD3) possesses a type IIs restriction enzyme, CdiI, which
cleaves the sequence 5'-CATCG-3' between the fourth and fifth nucleotide to give a
blunt-ended fragment. This is the first time that an enzyme with this specificity has been
reported. The sequential addition of this site to vectors showed that each site caused between
a five- and 16-fold reduction in transfer efficiency. The transfer efficiencies achieved with
both strains equated to between 1.0x10^-6 and 5.5x10^-5 transconjugants per donor.

<>

<1>Purdy, M.M., Holz-Schietinger, C., Reich, N.O.
<2>Identification of a second DNA binding site in human DNA methyltransferase 3A by substrate inhibition and domain deletion.
<3>Arch. Biochem. Biophys.
<4>498
<5>13-22
<6>2010
<7>The human DNA methyltransferase 3A (DNMT3A) is essential for establishing DNA methylation
patterns. Knowing the key factors involved
in the regulation of mammalian DNA methylation is critical to
furthering understanding of embryonic development and designing
therapeutic approaches targeting epigenetic mechanisms. We observe
substrate inhibition for the full length DNMT3A but not for its
isolated catalytic domain, demonstrating that DNMT3A has a second
binding site for DNA. Deletion of recognized domains of DNMT3A reveals
that the conserved PWWP domain is necessary for substrate inhibition
and forms at least part of the allosteric DNA binding site. The PWWP
domain is demonstrated here to bind DNA in a cooperative manner with mu
M affinity. No clear sequence preference was observed, similar to
previous observations with the isolated PWWP domain of Dnmt3b but with
one order of magnitude weaker affinity. Potential roles for a low
affinity, low specificity second DNA binding site are discussed.

<>

<1>Purdy, M.M., Reich, N.O.
<2>DNA binding by HhaI DNA methyltransferase domain interface mutants.
<3>FASEB J.
<4>20
<5>A901
<6>2006
<7>The bacterial HhaI DNA methyltransferase (M.HhaI) is a structurally and mechanistically
characterized member of the cytosine C-5 DNA
methyltransferase family. A Statistical Coupling Analysis, which uses
genetic covariation to estimate energetic coupling between protein
residues, was performed on this enzyme family. This analysis identified
a network of co-evolving residues centered on two regions - the
catalytic loop and domain interface, both of which were previously
implicated by crystallography in large scale conformational changes
upon DNA binding. Mutation of several domain interface residues in
M.HhaI resulted in 6 - 100 fold decreases in DNA affinity, despite their 7 - 20 angstrom
distances from the DNA. The effects of these
mutations suggest a role of this network in positioning or moving the
two domains and the importance of proper domain positioning in DNA
binding. The crystal structure of the I308A mutant was solved to 2.2
angstrom resolution. It is unclear if this mutation alters the
structure enough to explain the 35 fold loss in DNA affinity,
suggesting that this mutation may perturb the domain - domain motions
necessary for DNA binding.

<>

<1>Purdy, M.M., Reich, N.O.
<2>DNA binding and hemimethylation preference of HhaI DNA methyltransferase domain interface mutants.
<3>ACS Abstracts
<4>230
<5>U626
<6>2005
<7>Recent studies indicate potential interactions between mammalian DNA methyltransferase (DNMT)
enzymes and RNA. Transcriptional silencing by short or antisense RNA's directed to promoter
regions has been proposed to involve an RNA directed DNA methylation component, though this
remains controversial. Binding of duplex RNA to the DNMT3 isoforms has been demonstrated,
along with inhibition of DNMT1 by single stranded (ss) DNA. The present work shows that an ss
RNA is a more potent DNMT1 inhibitor than the corresponding ss DNA. ss RNA, being more
abundant than ss DNA, may represent an endogenous DNMT1 inhibitor. Preliminary results
indicate that an RNA/DNA hybrid duplex, suggested in some mechanisms for RNA directed DNA
methylation, is a poor DNMT1 substrate. Studies on interactions of the DNMT3 isoforms with ss
RNA and RNA/DNA duplexes are underway.

<>

<1>Purdy, M.M., Reich, N.O.
<2>Modulation of mammalian DNA methyltransferase activity by RNA.
<3>ACS Abstracts
<4>232
<5>244
<6>2006
<7>Recent studies indicate potential interactions between mammalian DNA methyltransferase enzymes
and RNA.  Transcriptional silencing by short or antisense RNA's directed to promoter regions
has been proposed to involve an RNA directed DNA methylation component, though this remains
controversial.  Binding of duplex RNA to the DNMT3 isoforms has been demonstrated, along with
inhibition of DNMT1 by single stranded DNA.  The present work shows that an ss RNA is a more
potent DNMT1 inhibitor than the corresponding ssDNA.  ssRNA, being more abundant than ssDNA,
may represent an endogenous DNMT1 inhibitor.  Preliminary results indicate that an RNA/DNA
hybrid duplex, suggested in some mechanisms for RNA directed DNA methylation, is a poor DNMT1
substrate.  Studies on interactions of the DNMT3 isoforms with ssRNA and RNA/DNA duplexes are
underway.

<>

<1>Purmal, A.A., Shabarova, Z.A., Gumport, R.I.
<2>A new affinity reagent for the site-specific, covalent attachment of DNA to active-site nucleophiles: application to the EcoRI and RsrI restriction and modification enzymes.
<3>Nucleic Acids Res.
<4>20
<5>3713-3719
<6>1992
<7>A modified oligodeoxyribonucleotide duplex containing an unnatural internucleotide
trisubstituted 3' to 5' pyrophosphate bond in one strand
[5'(oligo1)3'-P(OCH3)P-5'(oligo2)3'] reacts with nucleophiles in aqueous media by acting
as a phosphorylating affinity reagent. When interacted with a protein, a portion of the
oligonucleotide [-P-5'(oligo2)3'] becomes attached to an amino acid nucleophilic group
through a phosphate of the O-methyl-modified pyrophosphate linkage. We demonstate the affinity
labeling of nucleophilic groups at the active sites of the EcoRI and RsrI restriction and
modification enzymes with an oligodeoxyribonucleotide duplex containg a modified scissile bond
in the EcoRI recognitin site. With the EcoRI and RsrI endonuclease in molar excess
approximately 1% of the oligonucleotide becomes attached to the protein and with the companion
methyltransferases the yield approaches 40% for the EcoRI enzyme and 30% for the RsrI
methyltransferase. Crosslinking proceeds only upon formation of a sequence-specific enzyme-DNA
complex and generates a covalent bond between the 3'-phosphate of the modified pyrophosphate
in the substrate and a nucleophilic group at the active site of the enzymes. The reaction
results in the elimination of an oligodeoxyribonucleotide remnant that contains the
3'-O-methylphosphate [5'(oligo1)3'-P(OCH3)] derived from the modified phosphate of the
pyrophosphate linkage. Hydrolysis properties of the covalent protein-DNA adducts indicate that
phosphoamide (P-N) bonds are formed with the EcoRI endonuclease and methyltransferase.

<>

<1>Purmal, A.A., Vinogradova, M.N., Elov, A.A., Gromova, E.S., Drutsa, V.L.
<2>Interaction of EcoRII restriction and modification enzymes with DNA duplexes containing pyrophosphate bonds.
<3>Dokl. Akad. Nauk.
<4>276
<5>992-995
<6>1984
<7>
<>

<1>Purvis, I.J., Moseley, B.E.B.
<2>Isolation and characterisation of DraI, a type II restriction endonuclease recognising a sequence containing only A:T basepairs, and inhibition of its activity by uv irradiation of substrate DNA.
<3>Nucleic Acids Res.
<4>11
<5>5467-5474
<6>1983
<7>A type II restriction endonuclease, DraI, isolated from Deinococcus radiophilus
ATCC 27603 recognises the palindromic hexanucleotide sequence5'-T-T-T^A-A-A-3'
3'-A-A-A^T-T-T-5'and cleaves it, as indicated by the arrows, to produce
blunt-ended fragments.  The yield of enzyme is 100 to 1000 times that of the
only other known type II restriction endonuclease that recognises a sequence
composed solely of A:T basepairs, the isoschizomer AhaIII.  Ultraviolet
irradiation of the DNA substrate at relatively low doses inhibits the activity
of DraI by "protecting" the recognition sequence and this may be exploited to
give control of partial digestion of DNA by DraI.

<>

<1>Pushiri, H., Pearce, S.L., Oakeshott, J.G., Russell, R.J., Pandey, G.
<2>Draft Genome Sequence of Pandoraea sp. Strain SD6-2, Isolated from Lindane-Contaminated Australian Soil.
<3>Genome Announcements
<4>1
<5>e00415-13
<6>2013
<7>Pandoraea sp. strain SD6-2 is a delta-hexachlorocyclohexane-degrading bacterial strain
isolated from lindane-contaminated soil in Queensland, Australia. The
genome of SD6-2 was sequenced to investigate its ability to degrade
delta-hexachlorocyclohexane. Here we report the annotated genome sequence of this
strain.

<>

<1>Pustoshilova, N.M., Shingareva, N.V.
<2>Method for isolation of the restriction endonuclease HhaI.
<3>Soviet Patent Office
<4>SU 1613490 A
<5>
<6>1990
<7>The present invention is directed to a method for purification of the HhaI restriction
endonuclease. The cells of Haemophilus haemolyticus were disrupted by sonication. The lysate
was then extracted in a two-phase mixture of 7% polyethyleneglycol, 2% dextran, 38-45 mM NaCl,
pH 7.6. The upper phase was loaded onto a phosphocellulose PII column, and then a linear salt
gradient from 0.2 to 1.0 M NaCl in the buffer was applied. The HhaI yield was 4-6,000 units/g
cells with specific activity of HhaI of 25,000 units/ml. After incubation of 15-fold excess of
enzyme with DNA for 20 hours, no nuclease activity was detected.

<>

<1>Puvvada, M.S., Hartley, J.A., Jenkins, T.C., Thurston, D.E.
<2>A quantitative assay to measure the relative DNA-binding affinity of pyrrolo[2,1-c][1,4]benzodiazepine (PBD) antitumor antibiotics based on the inhibition of restriction endonuclease BamHI.
<3>Nucleic Acids Res.
<4>21
<5>3671-3675
<6>1993
<7>An assay has been developed (restriction endonuclease digestion assay - RED100) based on
inhibition of the restriction endonuclease BamHI that is capable of quantitive evaluation of
the relative DNA-binding affinity of pyrrolo[2,1-c][1,4]benzodiazepine (PBD) antitumor
antibiotics. This method provides comparable results to those obtained from thermal
denaturation and ethidium bromide displacement assays but is much more sensitive,
discriminating between molecules of similar structure such as DC-81, iso-DC-81 and
neothramycin. The results reveal a trend between relative DNA-binding affinity and in vitro
cytotoxicity for the PBDs in two tumour cell lines studied.

<>

<1>Puzio, P., Ascenzi, R., Mittendorf, V.
<2>Pathogen control genes and methods of use in plants.
<3>International Patent Office
<4>WO 2009027313
<5>
<6>2009
<7>The invention provides methods for conferring increased pathogen resistance to a plant.
Specifically, the invention relates to methods of producing transgenic plants with increased
nematode resistance, expression vectors comprising polynucleotides encoding polypeptides with
anti-nematode activity, and transgenic plants and seeds generated thereof.

<>

<1>Puzio, P., Blau, A., Plesch, G., Kamlage, B., Looser, R., Schmitz, O., Wendel, B.
<2>Process for the production of lutein.
<3>European Patent Office
<4>EP 2096177 A
<5>
<6>2009
<7>The present invention relates further to a  process for the control of the production of fine
chemical in a micro-organism, a plant cell, a plant, a plant tissue or in one or mroe parts
thereof.  The invention furthermroe relates to nucleic acid molecules, polypeptides, nucleic
acid constructs, vectors, antisense molecules, antibodies, host cells, plant tissue,
propagation material, harvested material, plants, microorganisms as well as agricultural
compositions and to their use.

<>

<1>Puzio, P., Blau, A., Plesch, G., Kamlage, B., Looser, R., Schmitz, O., Wendel, B.
<2>Process for the control of production of fine chemicals.
<3>European Patent Office
<4>EP 2199304 A
<5>
<6>2010
<7>The present invention relates further to a process for the control of the production of fine
chemical in a microorganism, a plant cell, a plant, a plant tissue or in one or more parts
thereof. The invention furthermore relates to nucleic acid molecules, polypeptides, nucleic
acid
constructs, vectors, antisense molecules, antibodies, host cells, plant tissue, propagation
material, harvested material, plants, microorganisms as well as agricultural compositions and
to their use.

<>

<1>Pyne, M.E., Moo-Young, M., Chung, D.A., Chou, C.P.
<2>Development of an electrotransformation protocol for genetic manipulation of Clostridium pasteurianum.
<3>Biotechnol. Biofuels.
<4>6
<5>50
<6>2013
<7>Background: Reducing the production cost of, and increasing revenues from, industrial biofuels
will greatly facilitate their proliferation
and co-integration with fossil fuels. The cost of feedstock is the
largest cost in most fermentation bioprocesses and therefore represents
an important target for cost reduction. Meanwhile, the biorefinery
concept advocates revenue growth through complete utilization of
by-products generated during biofuel production. Taken together, the
production of biofuels from low-cost crude glycerol, available in
oversupply as a by-product of bioethanol production, in the form of
thin stillage, and biodiesel production, embodies a remarkable
opportunity to advance affordable biofuel development. However, few
bacterial species possess the natural capacity to convert glycerol as a
sole source of carbon and energy into value-added bioproducts. Of
particular interest is the anaerobe Clostridium pasteurianum, the only
microorganism known to convert glycerol alone directly into butanol,
which currently holds immense promise as a high-energy biofuel and bulk
chemical. Unfortunately, genetic and metabolic engineering of C.
pasteurianum has been fundamentally impeded due to lack of an efficient
method for deoxyribonucleic acid (DNA) transfer.
Results: This work reports the development of an
electrotransformation protocol permitting high-level DNA transfer to C.
pasteurianum ATCC 6013 together with accompanying selection markers and
vector components. The CpaAI restriction-modification system was found
to be a major barrier to DNA delivery into C. pasteurianum which we
overcame by in vivo methylation of the recognition site (5'-CGCG-3')
using the M. FnuDII methyltransferase. With proper selection of the
replication origin and antibiotic-resistance marker, we initially
electroporated methylated DNA into C. pasteurianum at a low efficiency
of 2.4 x 10(1) transformants mu g(-1) DNA by utilizing conditions
common to other clostridial electroporations. Systematic investigation
of various parameters involved in the cell growth, washing and pulse
delivery, and outgrowth phases of the electrotransformation procedure
significantly elevated the electrotransformation efficiency, up to 7.5
x 10(4) transformants mu g(-1) DNA, an increase of approximately three
order of magnitude. Key factors affecting the electrotransformation
efficiency include cell-wall-weakening using glycine, ethanol-mediated
membrane solubilization, field strength of the electric pulse, and
sucrose osmoprotection.
Conclusions: C. pasteurianum ATCC 6013 can be electrotransformed at
a high efficiency using appropriately methylated plasmid DNA. The
electrotransformation method and tools reported here should promote
extensive genetic manipulation and metabolic engineering of this
biotechnologically important bacterium.

<>

<1>Pyne, M.E., Utturkar, S., Brown, S.D., Moo-Young, M., Chung, D.A., Chou, C.P.
<2>Improved Draft Genome Sequence of Clostridium pasteurianum Strain ATCC 6013 (DSM  525) Using a Hybrid Next-Generation Sequencing Approach.
<3>Genome Announcements
<4>2
<5>e00790-14
<6>2014
<7>We present an improved draft genome sequence for Clostridium pasteurianum strain  ATCC 6013
(DSM 525), the type strain of the species and an important
solventogenic bacterium with industrial potential. Availability of a
near-complete genome sequence will enable strain engineering of this promising
bacterium.

<>

<1>Qi, G.R., Wong, P., Cedergren, R.
<2>Restriction of single-stranded M13 DNA using synthetic oligonucleotides:  the structural requirement of restriction enzymes.
<3>Biochem. Cell Biol.
<4>65
<5>50-55
<6>1987
<7>A targeted ss (single stranded) DNA cleavage technique is reported which
involves the use of synthetic oligomers complementary to the ss M12 DNA
polylinker.  BamHI, SmaI, and KpnI restriction enzymes were tested with a
partial duplex DNA formed from ss M13 DNA and a nested series of fragments
derived from a synthetic 21-mer which were complementary to the polylinker
region.  These enzymes require up to two flanking nucleotides in addition to
the hexameric recognition site for efficient cleavage.  This technique could be
useful for effecting unique cleavages of DNA with enzymes which generally give
a large number of fragments and for strategies of ss DNA manipulation.

<>

<1>Qi, J., Guo, A., Cui, P., Chen, Y., Mustafa, R., Ba, X., Hu, C., Bai, Z., Chen, X., Shi, L., Chen, H.
<2>Comparative Geno-Plasticity Analysis of Mycoplasma bovis HB0801 (Chinese Isolate).
<3>PLoS ONE
<4>7
<5>E38239
<6>2012
<7>Mycoplasma bovis pneumonia in cattle has been epidemic in China since 2008. To
investigate M. bovis pathogenesis, we completed genome sequencing of strain
HB0801 isolated from a lesioned bovine lung from Hubei, China. The genomic
plasticity was determined by comparing HB0801 with M. bovis strain ATCC(R)
25523/PG45 from cow mastitis milk, Chinese strain Hubei-1 from lesioned lung
tissue, and 16 other Mycoplasmas species. Compared to PG45, the genome size of
HB0801 was reduced by 11.7 kb. Furthermore, a large chromosome inversion (580 kb)
was confirmed in all Chinese isolates including HB0801, HB1007, a strain from cow
mastitis milk, and Hubei-1. In addition, the variable surface lipoproteins (vsp)
gene cluster existed in HB0801, but contained less than half of the genes, and
had poor identity to that in PG45, but they had conserved structures. Further
inter-strain comparisons revealed other mechanisms of gene acquisition and loss
in HB0801 that primarily involved insertion sequence (IS) elements, integrative
conjugative element, restriction and modification systems, and some lipoproteins
and transmembrane proteins. Subsequently, PG45 and HB0801 virulence in cattle was
compared. Results indicated that both strains were pathogenic to cattle. The
scores of gross pathological assessment for the control group, and the PG45- and
HB0801-infected groups were 3, 13 and 9, respectively. Meanwhile the scores of
lung lesion for these three groups were 36, 70, and 69, respectively. In
addition, immunohistochemistry detection demonstrated that both strains were
similarly distributed in lungs and lymph nodes. Although PG45 showed slightly
higher virulence in calves than HB0801, there was no statistical difference
between the strains (P>0.05). Compared to Hubei-1, a total of 122 SNP loci were
disclosed in HB0801. In conclusion, although genomic plasticity was thought to be
an evolutionary advantage, it did not apparently affect virulence of M. bovis
strains in cattle.

<>

<1>Qi, M., Nelson, K.E., Daugherty, S.C., Nelson, W.C., Hance, I.R., Morrison, M., Forsberg, C.W.
<2>Novel molecular features of the fibrolytic intestinal bacterium Fibrobacter intestinalis not shared with Fibrobacter succinogenes as  determined by suppressive subtractive hybridization.
<3>J. Bacteriol.
<4>187
<5>3739-3751
<6>2005
<7>Suppressive subtractive hybridization was conducted to identify unique genes coding for plant
cell wall hydrolytic enzymes and other properties
of the gastrointestinal bacterium Fibrobacter intestinalis DR7 not shared
by Fibrobacter succinogenes S85. Subtractive clones from F. intestinalis
were sequenced and assembled to form 712 nonredundant contigs with an
average length of 525 bp. Of these, 55 sequences were unique to F.
intestinalis. The remaining contigs contained 764 genes with BLASTX
similarities to other proteins; of these, 80% had the highest similarities
to proteins in F. succinogenes, including 30 that coded for carbohydrate
active enzymes. The expression of 17 of these genes was verified by
Northern dot blot analysis. Of genes not exhibiting BLASTX similarity to
F. succinogenes, 30 encoded putative transposases, 6 encoded restriction
modification genes, and 45% had highest similarities to proteins in other
species of gastrointestinal bacteria, a finding suggestive of either
horizontal gene transfer to F. intestinalis or gene loss from F.
succinogenes. Analysis of contigs containing segments of two or more
adjacent genes revealed that only 35% exhibited BLASTX similarity and were
in the same orientation as those of F. succinogenes, indicating extensive
chromosomal rearrangement. The expression of eight transposases, and three
restriction-modification genes was confirmed by Northern dot blot
analysis. These data clearly document the maintenance of carbohydrate
active enzymes in F. intestinalis necessitated by the preponderance of
polysaccharide substrates available in the ruminal environment. It also
documents substantive changes in the genome from that of F. succinogenes,
which may be related to the introduction of the array of transposase and
restriction-modification genes.

<>

<1>Qi, M., Wang, D., Bradley, C.A., Zhao, Y.
<2>Genome Sequence Analyses of Pseudomonas savastanoi pv. glycinea and Subtractive Hybridization-Based Comparative Genomics with Nine Pseudomonads.
<3>PLoS ONE
<4>6
<5>E16451
<6>2011
<7>Bacterial blight, caused by Pseudomonas savastanoi pv. glycinea (Psg), is a
common disease of soybean. In an effort to compare a current field isolate with
one isolated in the early 1960s, the genomes of two Psg strains, race 4 and B076,
were sequenced using 454 pyrosequencing. The genomes of both Psg strains share
more than 4,900 highly conserved genes, indicating very low genetic diversity
between Psg genomes. Though conserved, genome rearrangements and recombination
events occur commonly within the two Psg genomes. When compared to each other,
437 and 163 specific genes were identified in B076 and race 4, respectively. Most
specific genes are plasmid-borne, indicating that acquisition and maintenance of
plasmids may represent a major mechanism to change the genetic composition of the
genome and even acquire new virulence factors. Type three secretion gene clusters
of Psg strains are near identical with that of P. savastanoi pv. phaseolicola
(Pph) strain 1448A and they shared 20 common effector genes. Furthermore, the
coronatine biosynthetic cluster is present on a large plasmid in strain B076, but
not in race 4. In silico subtractive hybridization-based comparative genomic
analyses with nine sequenced phytopathogenic pseudomonads identified dozens of
specific islands (SIs), and revealed that the genomes of Psg strains are more
similar to those belonging to the same genomospecies such as Pph 1448A than to
other phytopathogenic pseudomonads. The number of highly conserved genes (core
genome) among them decreased dramatically when more genomes were included in the
subtraction, suggesting the diversification of pseudomonads, and further
indicating the genome heterogeneity among pseudomonads. However, the number of
specific genes did not change significantly, suggesting these genes are indeed
specific in Psg genomes. These results reinforce the idea of a species complex of
P. syringae and support the reclassification of P. syringae into different
species.

<>

<1>Qian, L., Kussell, E.
<2>Evolutionary Dynamics of Restriction Site Avoidance.
<3>Phys. Rev. Lett.
<4>108
<5>158105
<6>2012
<7>Molecular noise in bacterial restriction-modification systems can cause rare events of host
DNA cleavage at restriction sites. Such
noise-induced selective pressure may result in evolved sequences
exhibiting restriction site avoidance. We identify a two-state regime
of evolutionary dynamics, in which populations either develop avoidance
or go extinct. Using perturbation theory, we show that equilibrium
sequence statistics exhibit power-law scaling in the ratio of
restriction strength to mutation rate. Noise levels comparable to
mutation rates can be sufficient to evolve detectable avoidance.

<>

<1>Qian, Y., Xi, Y., Cheng, B., Zhu, S.
<2>Genome-wide identification and expression profiling of DNA methyltransferase gene family in maize.
<3>Plant Cell
<4>33
<5>1661-1672
<6>2014
<7>In this study, we identified eight DNA MTase genes in maize and the diversity of expression
patterns of them was presented by EST mining, microarray and semi-quantitative expression
profile analyses.DNA methylation plays a pivotal role in promoting genomic stability through
diverse biological processes including regulation of gene expression during development and
chromatin organization. Although this important biological process is mainly regulated by
several conserved Cytosine-5 DNA methyltransferases encoded by a smaller multigene family in
plants, investigation of the plant C5-MTase-encoding gene family will serve to elucidate the
epigenetic mechanism diversity in plants. Recently, genome-wide identification and
evolutionary analyses of the C5-MTase-encoding gene family have been characterized in multiple
plant species including Arabidopsis, rice, carrot and wheat. However, little is known
regarding the C5-MTase-encoding genes in the entire maize genome. Here, genome-wide
identification and expression profile analyses of maize C5-MTase-encoding genes (ZmMETs) were
performed from the latest version of the maize (B73) genome. Phylogenetic analysis indicated
that the orthologs from the three species (maize, Arabidopsis and rice) were categorized into
four classes. Chromosomal location of these genes revealed that they are unevenly distributed
on 6 of all 10 chromosomes with three chromosomal/segmental duplication events, suggesting
that gene duplication played a key role in expansion of the maize C5-MTase-encoding gene
family. Furthermore, EST expression data mining, microarray data and semi-quantitative
expression profile analyses detected in the leaves by two different abiotic stress treatments
have demonstrated that these genes had temporal and spatial expression pattern and exhibited
different expression levels in stress treatments, suggesting that functional diversification
of ZmMET genes family. Overall, our study will serve to present signification insights to
explore the plant C5-MTase!
-encodin
g gene expression and function and also be beneficial for future experimental research to
further unravel the mechanisms of epigenetic regulation in plants.

<>

<1>Qiang, B.-Q., McClelland, M., Poddar, S., Spokauskas, A., Nelson, M.
<2>The apparent specificity of NotI (5'-GCGGCCGC-3') is enhanced by M.FnuDII or M.BepI methyltransferases (5'-mCGCG-3'): cutting bacterial chromosomes into a few large pieces .
<3>Gene
<4>88
<5>101-105
<6>1990
<7>The restriction endonuclease (ENase) NotI is blocked by methylation within its
recognition sequence at 5'GCGGCmCGC-3'.  This sensitivity to methylation can be
used to enhance the specificity of NotI in vivo and in vitro.  Modification by
M.FnuDII or M.BepI methyltransferases (MTase)(5'-mCGCG-3') will block NotI
(5'-GCGGCCGC-3') cleavage at overlapping MTase/ENase sites 5'-CGCGGCCGC-3'
(equivalent to 5'-GCGGCCGCG-3'), and increase the apparent cleavage specificity
of NotI about twofold.  This cross-protection procedure reduces the number of
NotI fragments in the genomes of Escherichia coli and Bacillus subtilis, as
resolved by pulsed field electrophoresis.  Application of this method to large
DNAs in vitro requires the preparation of highly purified DNA MTases.

<>

<1>Qiang, B.-Q., Schildkraut, I.
<2>Two unique restriction endonucleases from Neisseria lactamica.
<3>Nucleic Acids Res.
<4>14
<5>1991-1999
<6>1986
<7>Two new site-specific endonucleases, NlaIII and NlaIV, have been isolated from
Neisseria lactamica.  NlaIII recognizes the sequence, CATG, and cleaves 3' of
the sequence to produce a four base 3' extension.  NlaIV recognizes the
sequence, GGNNCC, and cleaves between the two N's to produce blunt ended
fragments.

<>

<1>Qiang, B.-Q., Schildkraut, I.
<2>A type II restriction endonuclease with an eight nucleotide specificity from Streptomyces fimbriatus.
<3>Nucleic Acids Res.
<4>12
<5>4507-4515
<6>1984
<7>A new site-specific endonuclease, SfiI, has been isolated from Streptomyces
fimbriatus.  This is the first report of a type II restriction endonuclease
whose recognition specificity requires eight nucleotides.  SfiI cleaves the
sequence, GGCCNNNN^NGGCC, symmetrically to produce a three base, 3' extension.

<>

<1>Qiang, B.-Q., Schildkraut, I.
<2>NotI and SfiI:  restriction endonucleases with octanucleotide recognition sequences.
<3>Methods Enzymol.
<4>155
<5>15-21
<6>1987
<7>While there are over 500 reported Type II restriction endonucleases, only two,
NotI and SfiI, require an octanucleotide recognition sequence.  These two
endonucleases cleave DNA less frequently than conventional tetranucleotide-,
pentanucleotide-, and hexanucleotide-recognizing restriction endonucleases.  On
average, NotI and SfiI will cleave DNA only once every 64,000 nucleotides.
With the advent of physical methods that can separate very large DNA molecules,
NotI and SfiI have become useful analytical reagents for molecular biologists.
NotI is isolated from the bacterium Nocardia otitidis-caviarum ATCC 14630, and
recognizes the DNA sequence     5'...GC^GGCCGC...3'     3'...CGCCGG^CG...5'
cleaving the phosphodiester bonds in both strands as indicated.  SfiI is
isolated from the bacterium Streptomyces fimbriatus ATCC 25051 and recognizes
the DNA sequence     5'...GGCCNNNN^NGGCC...3'     3'...CCGGN^NNNNCCGG...5'
cleaving the phosphodiester bonds in both strands as indicated.

<>

<1>Qiao, J., Chen, L., Li, Y., Wang, J., Zhang, W., Chen, S.
<2>Whole-Genome Sequence of Nocardiopsis alba Strain ATCC BAA-2165, Associated with  Honeybees.
<3>J. Bacteriol.
<4>194
<5>6358-6359
<6>2012
<7>The actinomycete Nocardiopsis alba was reportedly associated with honeybees in separate
occurrences. We report the complete genome of Nocardiopsis alba ATCC
BAA-2165 isolated from honeybee guts. It will provide insights into the
metabolism and genetic regulatory networks of this genus of bacteria that enable
them to live in a range of environments.

<>

<1>Qiao, J., Liu, Y., Liang, X., Hu, Y., Du, Y.
<2>Draft Genome Sequence of Root-Colonizing Bacterium Bacillus sp. Strain PTS-394.
<3>Genome Announcements
<4>2
<5>e00038-14
<6>2014
<7>Here, we report the high-quality draft genome sequence of Bacillus sp. strain PTS-394,
isolated from the rhizosphere of tomatoes grown on Putuo Mountain
(Xiamen, Fujian province, China), which exhibited excellent colonization ability
on plant roots. The 4.0-Mb genome uncovered the mechanism for its potential root
colonization ability and may provide novel insights into plant-bacterium
interactions.

<>

<1>Qin, N., Ding, W., Yao, J., Su, K., Wu, L., Li, L.
<2>Genome Sequence of Staphylococcus capitis QN1, Which Causes Infective Endocarditis.
<3>J. Bacteriol.
<4>194
<5>4469-4470
<6>2012
<7>Staphylococcus capitis is a subtype of coagulase-negative staphylococci (CoNS) which could
emerge as a significant pathogen causing infective endocarditis,
prosthetic valve endocarditis, and late-onset sepsis. We isolated S. capitis
strain QN1 from the skin swab sample of a female. Here we prepared a genome
sequence for this strain consisting of 30 contigs totaling 2,430,101 bases and a
GC content of 32.76%.

<>

<1>Qin, N., Zheng, B., Yang, F., Chen, Y., Guo, J., Hu, X., Li, L.
<2>Genome Sequence of Aerococcus viridans LL1.
<3>J. Bacteriol.
<4>194
<5>4143
<6>2012
<7>Aerococcus viridans is a catalase-negative Gram-positive bacterium and has been described as
an airborne organism widely distributed in the hospital environment
or in clinical specimens. We isolated A. viridans strain LL1 from indoor dust
samples collected by a patient. Here, we prepared a genome sequence for this
strain consisting of 31 contigs totaling 1,994,039 bases and a GC content of
39.42%.

<>

<1>Qin, Q.L., Li, Y., Zhang, Y.J., Zhou, Z.M., Zhang, W.X., Chen, X.L., Zhang, X.Y., Zhou, B.C., Wang, L., Zhang, Y.Z.
<2>Comparative genomics reveals a deep-sea sediment-adapted life style of Pseudoalteromonas sp. SM9913.
<3>ISME J.
<4>5
<5>274-284
<6>2011
<7>Deep-sea sediment is one of the most important microbial-driven ecosystems, yet it is not well
characterized. Genome sequence analyses of deep-sea sedimentary bacteria would shed light on
the understanding of this ecosystem. In this study, the complete genome of deep-sea
sedimentary bacterium Pseudoalteromonas sp. SM9913 (SM9913) is described and compared with
that of the closely related Antarctic surface sea-water ecotype Pseudoalteromonas haloplanktis
TAC125 (TAC125). SM9913 has fewer dioxygenase genes than TAC125, indicating a possible
sensitivity to reactive oxygen species. Accordingly, experimental results showed that SM9913
was less tolerant of H(2)O(2) than TAC125. SM9913 has gene clusters related to both polar and
lateral flagella biosynthesis. Lateral flagella, which are usually present in deep-sea
bacteria and absent in the related surface bacteria, are important for the survival of SM9913
in deep-sea environments. With these two flagellar systems, SM9913 can swim in sea water and
swarm on the sediment particle surface, favoring the acquisition of nutrients from particulate
organic matter and reflecting the particle-associated alternative lifestyle of SM9913 in the
deep sea. A total of 12 genomic islands were identified in the genome of SM9913 that may
confer specific features unique to SM9913 and absent from TAC125, such as drug and heavy metal
resistance. Many signal transduction genes and a glycogen production operon were also present
in the SM9913 genome, which may help SM9913 respond to food pulses and store carbon and energy
in a deep-sea environment.

<>

<1>Qin, Q.L., Xie, B.B., Shu, Y.L., Rong, J.C., Zhao, D.L., Zhang, X.Y., Chen, X.L., Zhou, B.C., Zhang, Y.Z.
<2>Genome Sequence of Proteorhodopsin-Containing Sea Ice Bacterium Glaciecola punicea ACAM 611T.
<3>J. Bacteriol.
<4>194
<5>3267
<6>2012
<7>Here, we report the draft genome sequence of Antarctic sea ice bacterium Glaciecola punicea
ACAM 611(T), the type species of the genus Glaciecola. A
blue-light-absorbing proteorhodopsin gene is present in the 3.08-Mb genome. This
genome sequence can facilitate the study of the physiological metabolisms and
ecological roles of sea ice bacteria.

<>

<1>Qin, Q.L., Zhang, X.Y., Wang, X.M., Liu, G.M., Chen, X.L., Xie, B.B., Dang, H.Y., Zhou, B.C., Yu, J., Zhang, Y.Z.
<2>The complete genome of Zunongwangia profunda SM-A87 reveals its adaptation to the deep-sea environment and ecological role in sedimentary organic nitrogen degradation.
<3>BMC Genomics
<4>11
<5>247
<6>2010
<7>BACKGROUND: Zunongwangia profunda SM-A87, which was isolated from deep-sea
sediment, is an aerobic, gram-negative bacterium that represents a new
genus of Flavobacteriaceae. This is the first sequenced genome of a
deep-sea bacterium from the phylum Bacteroidetes. RESULTS: The Z. profunda
SM-A87 genome has a single 5 128 187-bp circular chromosome with no
extrachromosomal elements and harbors 4 653 predicted protein-coding
genes. SM-A87 produces a large amount of capsular polysaccharides and
possesses two polysaccharide biosynthesis gene clusters. It has a total of
130 peptidases, 61 of which have signal peptides. In addition to
extracellular peptidases, SM-A87 also has various extracellular enzymes
for carbohydrate, lipid and DNA degradation. These extracellular enzymes
suggest that the bacterium is able to hydrolyze organic materials in the
sediment, especially carbohydrates and proteinaceous organic nitrogen.
There are two clustered regularly interspaced short palindromic repeats in
the genome, but their spacers do not match any sequences in the public
sequence databases. SM-A87 is a moderate halophile. Our protein
isoelectric point analysis indicates that extracellular proteins have
lower predicted isoelectric points than intracellular proteins. SM-A87
accumulates organic osmolytes in the cell, so its extracelluar proteins
are more halophilic than its intracellular proteins. CONCLUSION: Here, we
present the first complete genome of a deep-sea sedimentary bacterium from
the phylum Bacteroidetes. The genome analysis shows that SM-A87 has some
common features of deep-sea bacteria, as well as an important capacity to
hydrolyze sedimentary organic nitrogen.

<>

<1>Qin, S., Zhang, H., Li, F., Zhu, B., Zheng, H.
<2>Draft Genome Sequence of Marine Streptomyces sp. Strain W007, Which Produces Angucyclinone Antibiotics with a Benz[a]anthracene Skeleton.
<3>J. Bacteriol.
<4>194
<5>1628-1629
<6>2012
<7>A series of angucyclinone antibiotics have been isolated from marine Streptomyces sp. strain
W007 and identified. Here, a draft genome sequence of Streptomyces sp.
W007 is presented. The genome contains an intact biosynthetic gene cluster for
angucyclinone antibiotics, which provides insight into the combinatorial
biosynthesis of angucyclinone antibiotics produced by marine streptomycetes.

<>

<1>Qin, T., Cui, Y., Cen, Z., Liang, T., Ren, H., Yang, X., Zhao, X., Liu, Z., Xu, L., Li, D., Song, Y., Yang, R., Shao, Z., Song, Y.
<2>Draft Genome Sequences of Two Legionella dumoffii Strains, TEX-KL and NY-23.
<3>J. Bacteriol.
<4>194
<5>1251-1252
<6>2012
<7>Legionella (Fluoribacter) dumoffii is one of the agents causing Legionnaires' disease. Here,
we used Illumina second-generation sequencing technology to
decipher for the first time the whole-genome sequences of two strains of this
species, TEX-KL and NY-23. The assembly results for both strains consist of one
chromosome and two plasmids.

<>

<1>Qin, W., Yung, L.Y.L.
<2>Efficient manipulation of nanoparticle-bound DNA by restriction endonuclease.
<3>ACS Abstracts
<4>230
<5>U1077
<6>2005
<7>
<>

<1>Qin, W.H., Leonhardt, H., Spada, F.
<2>Usp7 and Uhrf1 Control Ubiquitination and Stability of the Maintenance DNA Methyltransferase Dnmt1.
<3>J. Cell. Biochem.
<4>112
<5>439-444
<6>2011
<7>In mammals Dnmt1 is the DNA methyltransferase chiefly responsible for maintaining genomic
methylation patterns through DNA replication
cycles, but how its maintenance activity is controlled is still not
well understood. Interestingly, Uhrf1, a crucial cofactor for
maintenance of DNA methylation by Dnmt1, is endowed with E3 ubiquitin
ligase activity. Here, we show that both Dnmt1 and Uhrf1 coprecipitate
with ubiquitin specific peptidase 7 (Usp7), a de-ubiquitinating enzyme.
Overexpression of Uhrf1 and Usp7 resulted in opposite changes in the
ubiquitination status and stability of Dnmt1. Our findings suggest
that, by balancing Dnmt1 ubiquitination, Usp7 and Uhrf1 fine tune Dnmt1
stability.

<>

<1>Qin, W.J., Yung, L.Y.L.
<2>Efficient manipulation of nanoparticle-bound DNA via restriction endonuclease.
<3>Biomacromolecules
<4>7
<5>3047-3051
<6>2006
<7>As a programmable biopolymer, DNA has shown great potential in the fabrication and
construction of nanometer-scale assemblies and devices.
In this report, we described a strategy for efficient manipulation of
gold nanoparticle-bound DNA using restriction endonuclease. The
digestion efficiency of this restriction enzyme was studied by varying
the surface coverage of stabilizer, the size of nanoparticles, as well
as the distance between the nanoparticle surface and the enzyme-cutting
site of particle-bound DNA. We found that the surface coverage of
stabilizer is crucial for achieving high digestion efficiency. In
addition, this stabilizer surface coverage can be tailored by varying
the ion strength of the system. Based on the results of polyacrylamide
gel electrophoresis and fluorescent study, a high digestion efficiency
of 90+% for particle-bound DNA was achieved for the first time. This
restriction enzyme manipulation can be considered as an additional
level of control of the particle-bound DNA and is expected to be
applied to manipulate more complicated nanostructures assembled by DNA.

<>

<1>Qin, X., Evans, J.D., Aronstein, K.A., Murray, K.D., Weinstock, G.M.
<2>Genome sequences of the honey bee pathogens Paenibacillus larvae and Ascosphaera apis.
<3>Insect Mol. Biol.
<4>15
<5>715-718
<6>2006
<7>Genome sequences offer a broad view of host-pathogen interactions at the systems
biology level. With the completion of the sequence of the honey bee, interest in
the relevant pathogens is heightened. Here we report the genome sequences of two
of the major pathogens of honey bees, the bacterium Paenibacillus larvae
(causative agent for American foulbrood disease) and the fungus Ascosphaera apis.
(causative agent for chalkbrood disease). Ongoing efforts to characterize the
genomes of these species can be used to understand and mitigate the effects of
two important pathogens, and will provide a contrast with pathogenic, benign and
freeliving relatives.

<>

<1>Qin, Z., Deng, Z., Zhou, Q., Chen, H.
<2>Overcoming restriction of Streptomyces hygroscopicus 10-22 by the modification of S. fradiae -- An attempt to develop a transformation system for S. hygroscopicus 10-22.
<3>I Chuan Hsueh Pao
<4>20
<5>180-184
<6>1993
<7>No transformant was obtained when pIJ702 (tsr, mel+) from S. lividans TK24 was used to
transform S. hygroscopicus 10-22. pIJ702 isolated from S. fradiae ATCC 10745, however, was
transformed into 10-22 at a frequency of 10/3-10/4 tranformants/ug DNA. Among the transformant
colonies, only 1/1000 of them were black in colour (mel+) while a great majority of them
remained white (mel-). The plasmid pIJ702 band was only visualized on agarose gels from the
black colonies but not from the white colonies. However, when pIJ702 isolated from both black
and white transformants were used to transform S. lividans TK24, the mel gene was expressed
normally in the recipients. The preparation was also successful in transforming S.
hygroscopicus 10-22, and again gave rise to 1/1000 of black colonies only. When the 10-22
(pIJ702) black colonies were plated on non-selective medium, among the majority of black
colonies grown, there were a few white colonies, which proved to be host mutants of 10-22.
These mutants were transformable by pIJ702 and homogeneous black colonies were obtained.

<>

<1>Qin, Z., Peng, K., Zhou, X., Lliang, R., Zhou, Q., Chen, H., Hopwood, D.A., Kieser, T., Deng, Z.
<2>Development of a gene cloning system for Streptomyces hygroscopicus subsp. yingchengensis, a producer of three useful antifungal compounds, by elimination of three barriers to DNA transfer.
<3>J. Bacteriol.
<4>176
<5>2090-2095
<6>1994
<7>Streptomyces hygroscopicus 10-22 could not be transformed with any of the commonly used
Streptomyces plasmid vectors and was resistant to plaque formation by the Streptomyces phages
C31 and R4. Repeated selection resulted in the isolation of derivatives of S. hygroscopicus
10-22 that could be transformed with pIJ101- and pJV1-derived cloning vectors and of
restriction-deficient derivatives that could accept DNA propagated in Streptomyces lividans
66. These new strains, which include three that still produce the original antibiotics, can be
used as hosts for gene cloning. Insertion of nonreplicating vectors by homologous
recombination and transposition of Tn4560 were demonstrated in S. hygroscopicus 10-22.

<>

<1>Qiu, C., Sawada, K., Zhang, X., Cheng, X.
<2>The PWWP domain of mammalian DNA methyltransferase Dnmt3b defines a new family of DNA-binding folds.
<3>Nat. Struct. Biol.
<4>9
<5>217-224
<6>2002
<7>The PWWP domain is a weakly conserved sequence motif found in >60 eukaryotic proteins,
including the mammalian DNA methyltransferases Dnmt3a and Dnmt3b. These proteins often contain
other chromatin-association domains. A 135-residue PWWP domain from mouse Dnmt3b (amino acids
223--357) has been structurally characterized at 1.8 A resolution. The N-terminal half of this
domain resembles a barrel-like five-stranded structure, whereas the C-terminal half contains a
five-helix bundle. The two halves are packed against each other to form a single structural
module that exhibits a prominent positive electrostatic potential. The PWWP domain alone binds
DNA in vitro, probably through its basic surface. We also show that recombinant Dnmt3b2
protein (a splice variant of Dnmt3b) and two N-terminal deletion mutants (Delta218 and
Delta369) have approximately equal methyl transfer activity on unmethylated and hemimethylated
CpG-containing oligonucleotides. The Delta218 protein, which includes the PWWP domain, binds
DNA more strongly than Delta369, which lacks the PWWP domain.

<>

<1>Qiu, D., Eisinger, V.M., Head, N.E., Pier, G.B., Yu, H.D.
<2>ClpXP proteases positively regulate alginate overexpression and mucoid conversion in Pseudomonas aeruginosa.
<3>Microbiology
<4>154
<5>2119-2130
<6>2008
<7>Overproduction of the exopolysaccharide alginate and conversion to a mucoid
phenotype in Pseudomonas aeruginosa are markers for the onset of chronic lung
infection in cystic fibrosis (CF). Alginate production is regulated by the
extracytoplasmic function (ECF) sigma factor AlgU/T and the cognate anti-sigma
factor MucA. Many clinical mucoid isolates carry loss-of-function mutations in
mucA. These mutations, including the most common mucA22 allele, cause C-terminal
truncations in MucA, indicating that an inability to regulate AlgU activity by
MucA is associated with conversion to the mucoid phenotype. Here we report that a
mutation in a stable mucoid strain derived from the parental strain PAO1,
designated PAO581, that does not contain the mucA22 allele, was due to a
single-base deletion in mucA (DeltaT180), generating another type of C-terminal
truncation. A global mariner transposon screen in PAO581 for non-mucoid isolates
led to the identification of three regulators of alginate production, clpP
(PA1801), clpX (PA1802), and a clpP paralogue (PA3326, designated clpP2). The
PAO581 null mutants of clpP, clpX and clpP2 showed decreased AlgU transcriptional
activity and an accumulation of haemagglutinin (HA)-tagged N-terminal MucA
protein with an apparent molecular mass of 15 kDa. The clpP and clpX mutants of a
CF mucoid isolate revert to the non-mucoid phenotype. The ClpXP and ClpP2
proteins appear to be part of a proteolytic network that degrades the cytoplasmic
portion of truncated MucA proteins to release the sequestered AlgU, which drives
alginate biosynthesis.

<>

<1>Qiu, D., Wei, H., Tu, Q., Yang, Y., Xie, M., Chen, J., Pinkerton, M.H., Liang, Y., He, Z., Zhou, J.
<2>Combined Genomics and Experimental Analyses of Respiratory Characteristics of Shewanella putrefaciens W3-18-1.
<3>Appl. Environ. Microbiol.
<4>79
<5>5250-5257
<6>2013
<7>It has previously been shown that the Shewanella putrefaciens W3-18-1 strain produces
remarkably high current in microbial fuel cells (MFCs)
and can form magnetite at 0 degrees C. To explore the underlying
mechanisms, we developed a genetic manipulation method by deleting the
restriction-modification system genes of the SGI1 (Salmonella genome
island 1)-like prophage and analyzed the key genes involved in
bacterial respiration. W3-18-1 has less respiratory flexibility than
the well-characterized S. oneidensis MR-1 strain, as it possesses fewer
cytochrome c genes and lacks the ability to oxidize sulfite or reduce
dimethyl sulfoxide (DMSO) and timethylamine oxide (TMAO). W3-18-1 lacks
the hydrogen-producing Fe-only hydrogenase, and the hydrogen-oxidizing
Ni-Fe hydrogenase genes were split into two separate clusters. Two
periplasmic nitrate reductases (NapDAGHB and NapDABC) were functionally
redundant in anaerobic growth of W3-18-1 with nitrate as the electron
acceptor, though napDABC was not regulated by Crp. Moreover, nitrate
respiration started earlier in W3-18-1 than in MR-1 (with NapDAGHB
only) under microoxic conditions. These results indicate that
Shewanella putrefaciens W3-18-1 is well adapted to habitats with higher
oxygen levels. Taken together, the results of this study provide
valuable insights into bacterial genome evolution.

<>

<1>Qiu, J., Ren, J.
<2>Database management system for recognition sequences and sequences assembly of restriction endonuclease and methylase.
<3>Zhongguo Yaoke Daxue Xuebao
<4>20
<5>185-187
<6>1989
<7>None

<>

<1>Qiu, X., Han, R., Yan, X., Liu, M., Cao, L., Yoshiga, T., Kondo, E.
<2>Identification and Characterization of a Novel Gene Involved in the trans-Specific Nematicidal Activity of Photorhabdus luminescens LN2.
<3>Appl. Environ. Microbiol.
<4>75
<5>4221-4223
<6>2009
<7>Photorhabdus luminescens subsp. akhurstii LN2 from Heterorhabditis indica
LN2 showed nematicidal activity against axenic Heterorhabditis
bacteriophora H06 infective juveniles (IJs). Transposon mutagenesis
identified an LN2 mutant that supports the growth of H06 nematodes. Tn5
disrupted the namA gene, encoding a novel 364-residue protein and
involving the nematicidal activity. The green fluorescent protein-labeled
namA mutant was unable to colonize the intestines of H06 IJs.

<>

<1>Qiu, X., Zhan, Z.B., Yan, X., Han, R.
<2>Draft Genome Sequence and Annotation of the Entomopathogenic Bacterium Photorhabdus luminescens LN2, Which Shows Nematicidal Activity against  Heterorhabditis bacteriophora H06 Nematodes.
<3>Genome Announcements
<4>2
<5>e01268-14
<6>2014
<7>We present here the 5.6-Mb genome sequence of Photorhabdus luminescens strain LN2, a
Gram-negative bacterium that is a symbiont of Heterorhabditis indica LN2
and shows nematicidal activity against Heterorhabditis bacteriophora H06
nematodes.

<>

<1>Qu, Y., Zhang, X., Yu, H., Tang, H., Shen, E., Zhou, H., Ma, Q., Cao, X., Zhou, J., Xu, P.
<2>Genome Sequence of Sphingomonas xenophaga QYY, an Anthraquinone-Degrading Strain.
<3>Genome Announcements
<4>1
<5>e00031-12
<6>2013
<7>Sphingomonas xenophaga QYY is an efficient anthraquinone-degrading strain. Here,  we present a
4.2-Mb assembly of the first genome sequence of S. xenophaga. We have annotated 36 coding
sequences (CDSs) encoding aromatic catabolism and 216 CDSs responsible for toxic resistance
and stress response, which may provide insights into the degradation of complex aromatics.

<>

<1>Quada, J.C., Izbicka, E., Rashidi, H.H.
<2>Dual flipping and novel motifs of DNA methyltransferase.
<3>Proc. Amer. Assoc. Cancer Res.
<4>41
<5>79
<6>2000
<7>Structural superimposition of three resolved structures of DNA cytosine methyltransferase
(DCMTase) M.HhaI revealed the dual flipping feature of the enzyme and the cofactor
S-adenosyl-L-methionine (SAM) or its demethylated form, S-adenosyl-L-homocysteine (SAH).  In
the absence of DNA, enzyme-bound SAM is solvent exposed.  In the presence of DNA, SAM flips
from the surface of the enzyme into the catalytic site and complements the flipped out
cytosine methylation target in DNA.  Analysis of amino acid residues interacting with the
cofactor identified 16 contact amino acid residues for unflipped SAM and 24 residues for
flipped SAM/SAH in M.HhaI.  These contact residues are also conserved in higher eukaryotic
DCMTases.  Human Dnmt1 and M.HhaI bound to SAH and DNA and M.HhaI bound to flipped SAM reveals
that SAH and flipped SAM are coordinated in the same binding pocket.  Analysis of the
cofactor-interacting residues in M.HhaI, human Dnmt1 and other known DCMTases enabled us to
define two novel flipped SAM/SAH binding sequence motifs expressed in the same order in all
DCMTases, providing a specific tool for analysis of related proteins.  High sequence homology
of the cofactor interacting residues implies their conserved function and further underlines
the structural and functional similarity between bacterial M.HhaI and human Dnmt1 DCMTases.
Our findings also support the alternative binding mechanism in which SAM binding precedes DNA
binding in M.HhaI.

<>

<1>Que, Q., Zhang, Y., Nelson, M., Ropp, S., Burbank, D.E., Van Etten, J.L.
<2>Chlorella virus SC-1A encodes at least five functional and one nonfunctional DNA methyltransferases.
<3>Gene
<4>190
<5>237-244
<6>1997
<7>Chlorella virus SC-1A encodes at least six DNA methyltransferases: four N6-methyldeoxyadenine
(m6A) Mtases, M.CviSI (TGCmA), M.CviSII (CmATG), M.CviSIII (TCGmA) and M.CviSIV (GmATC), one
5-methyldeoxycytosine (m5C) Mtase, M.CviSV (~RCmCG), and one nonfunctional m5C MTase,
M.CviSVI, which is homologous to the MTase M.CviJI [RGmC(T/C/G)] produced by another chlorella
virus IL-3A.  Genes encoding three of the SC-1A m6A MTases (M.CviSI, M.CviSII, and M.CviSIII)
and the nonfunctional m5C MTase were cloned and sequenced.  Neither M.CviSI nor M.CviSIII
genes hybridized to genes for their respective isomethylomers, M.CviRI and M.CviBIII, from
other chlorella viruses.  However, the M.CviSII gene hybridized strongly to its M.CviAII
isomethylomer gene from virus PBCV-1.  Like the prototype chlorella virus PBCV-1, the SC-1A
genome contains inverted terminal repeats, one of which is adjacent to the nonfunctional m5C
MTase.  The three cloned m6A MTase genes are distributed throughout the approx. 345 kb SC-1A
genome.

<>

<1>Quick, J., Constantinidou, C., Pallen, M.J., Oppenheim, B., Loman, N.J.
<2>Draft Genome Sequence of Elizabethkingia meningoseptica Isolated from a Traumatic Wound.
<3>Genome Announcements
<4>2
<5>e00355-14
<6>2014
<7>We report the draft genome assembly of Elizabethkingia meningoseptica strain 502. The sample
was isolated from the wound of a repatriated military serviceperson
who suffered major trauma from an improvised explosive device (IED), resulting in
wounds with extensive environmental contamination. E. meningoseptica was isolated
from wounds in both legs. The draft genome assembly has 21 contigs with a total
size of 3,960,744 bases. The genome contains genes encoding 26 putative
beta-lactamases.

<>

<1>Quint, A., Cedar, H.
<2>In vitro methylation of DNA with HpaII methylase.
<3>Nucleic Acids Res.
<4>9
<5>633-646
<6>1981
<7>The enzyme HpaII methylase extracted and partially purified from Haemophilus
parainfluenza catalyzes the methylation of the tetranucleotide sequence CCGG at
the internal cytosine.  The enzyme will methylate this sequence if both DNA
strands are unmethylated or if only one strand is unmethylated.  Conditions
have been developed for producing fully methylated DNA from various sources.
In vitro methylation of this site protects the DNA against digestion by the
restriction enzyme HpaII as well as the enzyme SmaI which recognizes the
hexanucleotide sequence CCCGGG.  These properties make this enzyme a valuable
tool for analyzing methylation in eukaryotic DNA where the sequence CCGG is
highly methylated.  The activity of this methylase on such DNA indicates the
degree of undermethylation of the CCGG sequence.  Several examples show that
this technique can be used to detect small changes in the methylation state of
eukaryotic DNA.

<>

<1>Quirk, S.M., Bell-Pedersen, D., Belfort, M.
<2>Intron mobility in the T-even phages:  high frequency inheritance of group I introns promoted by intron open reading frames.
<3>Cell
<4>56
<5>455-465
<6>1989
<7>Intron mobility in the T-even phages has been demonstrated. Efficient nonreciprocal conversion
of intron minus (In-) alleles to intron plus (In+) occurred for the td and sunY genes, but not
for nrdB. Conversion to In+ was absolutely dependent on expression of the respective intron
open reading frame (ORF). Introns were inserted at their cognate sites in an intronless phage
genome via an RNA-independent, DNA-based, duplicative recombination event that was stimulated
by exon homology. The fd intron ORF product directs the endonucleolytic cleavage of DNA,
targeting the site of intron integration. A 21 nucleotide deletion of the integration site
abolished high frequency intron inheritance. These experiments provide a novel example of gene
conversion in prokaryotes, while suggesting a molecular rationale for the inconsistent
distribution of introns within highly conserved exon contexts of the T-even phage genomes.

<>

<1>Qureshi, A., Itankar, Y., Ojha, R., Mandal, M., Khardenavis, A., Kapley, A., Purohit, H.J.
<2>Genome Sequence of Lactobacillus plantarum EGD-AQ4, Isolated from Fermented Product of Northeast India.
<3>Genome Announcements
<4>2
<5>e01122-13
<6>2014
<7>We present a draft genome sequence of Lactobacillus plantarum strain EGD-AQ4, isolated from
nonalcoholic fermented bamboo shoot products of Northeast India.
The size of the draft genome sequence is the largest among all the reported
genome sequences of Lactobacillus plantarum, thus enabling the exploration of new
gene clusters involved in various functional and probiotic attributes.

<>

<1>Ra, S.R., Ri, D.C.
<2>Isolation-purification of a new restriction endonuclease, CglI and its optimum reaction condition.
<3>Choson Minjujuui Inmin Konghwaguk Kwahagwon Tongbo
<4>0
<5>36-38
<6>2000
<7>A new restriction endonuclease CglI is isolated and purified from Corynebacterium glutamicum
165 by a ultrasonication, Biogel filtered phosphocellulose and DEAE-cellulose column
chromatography.  We establish some reasonable reaction condition for the purified restriction
endonuclease, CglI.

<>

<1>Raaijmakers, H., Toro, I., Birkenbihl, R., Kemper, B., Suck, D.
<2>Conformational flexibility in T4 endonuclease VII revealed by crystallography: implications for substrate binding and cleavage.
<3>J. Mol. Biol.
<4>308
<5>311-323
<6>2001
<7>The structure of the N62D mutant of the junction-resolving endonuclease VII (EndoVII) from
phage T4 has been refined at 1.3 A, and a second wild-type crystal form solved and refined at
2.8 A resolution. Comparison of the mutant with the wild-type protein structure in two
different crystal environments reveals considerable conformational flexibility at the dimer
level affecting the substrate-binding cleft, the dimerization interface and the orientation of
the C-terminal domains. The opening of the DNA-binding cleft, the orientation of the
C-terminal domains relative to the central dimerization domain as well as the relative
positioning of helices in the dimerization interface appear to be sensitive to the crystal
packing environment. The highly unexpected rearrangement within the extended hydrophobic
interface does change the contact surface area but keeps the number of hydrophobic contacts
about the same and will therefore not require significant energy input. The conformational
flexibility most likely is of functional significance for the broad substrate specificity of
EndoVII. Binding of sulphate ions in the mutant structure and their positions relative to the
active-site metal ions and residues known to be essential for catalysis allows us to propose a
possible catalytic mechanism. A comparison with the active-site geometries of other
magnesium-dependent nucleases, among them the homing endonuclease I-PpoI and Serratia
endonuclease, shows common features, suggesting related catalytic mechanisms. Copyright 2001
Academic Press.

<>

<1>Rabinkova, E.V., Torosyan, M.V., Fradkin, G.E.
<2>Restriction alleviation of phage lambda in Escherichia coli K-12 cells after gamma-irradiation.
<3>Radiobiologiia
<4>27
<5>563-564
<6>1987
<7>In gamma-irradiated cells of Escherichia coli K-12 restriction alleviation of an unmodified
phage lambda is only observed in AB1157 strain.  No restriction alleviation by gamma-rays is
registered in AB1157 mutants (recA and ssb-1).

<>

<1>Rabsch, W., Helm, R.A., Eisenstark, A.
<2>Diversity of phage types among archived cultures of the demerec collection of Salmonella enterica serovar Typhimurium strains.
<3>Appl. Environ. Microbiol.
<4>70
<5>664-669
<6>2004
<7>The existence of several thousand Salmonella enterica serovar Typhimurium LT2 and LT7 cultures
originally collected by M. Demerec and
sealed in agar stab vials for 33 to 46 years is a resource for
evolutionary and mutational studies. Cultures from 74 of these vials,
descendants of cells sealed and stored in nutrient agar stabs several
decades ago, were phage typed by the Callow and Felix, Lilleengen, and
Anderson systems. Among 53 LT2 archived strains, 16 had the same phage
type as the nonarchival sequenced LT2 strain. The other 37 archived
cultures differed in phage typing pattern from the sequenced strain.
These 37 strains were divided into 10 different phage types. Among the
19 LT7 strains, only one was similar to the parent by phage typing,
while 18 were different. These 18 strains fell into eight different
phage types. The typing systems were developed to track epidemics from
source to consumer, as well as geographic spread. The value of phage
typing is dependent upon the stability of the phage type of any given
strain throughout the course of the investigation. Thus, the variation
over time observed in these archived cultures is particularly
surprising. Possible mechanisms for such striking diversity may include
loss of prophages, prophage mosaics as a result of recombination
events, changes in phage receptor sites on the bacterial cell surface,
or mutations in restriction-modification systems.

<>

<1>Rabus, R., Kube, M., Heider, J., Beck, A., Heitmann, K., Widdel, F., Reinhardt, R.
<2>The genome sequence of an anaerobic aromatic-degrading denitrifying bacterium, strain EbN1.
<3>Arch. Microbiol.
<4>183
<5>27-36
<6>2005
<7>Recent research on microbial degradation of aromatic and other refractory compounds in anoxic
waters and soils has revealed that nitrate-reducing
bacteria belonging to the Betaproteobacteria contribute substantially to
this process. Here we present the first complete genome of a metabolically
versatile representative, strain EbN1, which metabolizes various aromatic
compounds, including hydrocarbons. A circular chromosome (4.3 Mb) and two
plasmids (0.21 and 0.22 Mb) encode 4603 predicted proteins. Ten anaerobic
and four aerobic aromatic degradation pathways were recognized, with the
encoding genes mostly forming clusters. The presence of paralogous gene
clusters (e.g., for anaerobic phenylacetate oxidation), high sequence
similarities to orthologs from other strains (e.g., for anaerobic phenol
metabolism) and frequent mobile genetic elements (e.g., more than 200
genes for transposases) suggest high genome plasticity and extensive
lateral gene transfer during metabolic evolution of strain EbN1. Metabolic
versatility is also reflected by the presence of multiple respiratory
complexes. A large number of regulators, including more than 30
two-component and several FNR-type regulators, indicate a finely tuned
regulatory network able to respond to the fluctuating availability of
organic substrates and electron acceptors in the environment. The absence
of genes required for nitrogen fixation and specific interaction with
plants separates strain EbN1 ecophysiologically from the closely related
nitrogen-fixing plant symbionts of the Azoarcus cluster. Supplementary
material on sequence and annotation are provided at the Web page
http://www.micro-genomes.mpg.de/ebn1/.

<>

<1>Rabus, R., Ruepp, A., Frickey, T., Rattei, T., Fartmann, B., Stark, M., Bauer, M., Zibat, A., Lombardot, T., Becker, I., Amann, J., Gellner, K., Teeling, H., Leuschner, W.D., Glockner, F.O., Lupas, A.N., Amann, R., Klenk, H.P.
<2>The genome of Desulfotalea psychrophila, a sulfate-reducing bacterium from permanently cold Arctic sediments.
<3>Environ. Microbiol.
<4>6
<5>887-902
<6>2004
<7>Summary Desulfotalea psychrophila is a marine sulfate-reducing delta-proteobacterium that is
able to grow at in situ temperatures below 0
degrees C. As abundant members of the microbial community in permanently
cold marine sediments, D. psychrophila-like bacteria contribute to the
global cycles of carbon and sulfur. Here, we describe the genome sequence
of D. psychrophila strain LSv54, which consists of a 3 523 383 bp circular
chromosome with 3118 predicted genes and two plasmids of 121 586 bp and 14
663 bp. Analysis of the genome gave insight into the metabolic properties
of the organism, e.g. the presence of TRAP-T systems as a major route for
the uptake of C(4)-dicarboxylates, the unexpected presence of genes from
the TCA cycle, a TAT secretion system, the lack of a beta-oxidation
complex and typical Desulfovibrio cytochromes, such as c(553), c(3) and
ncc. D. psychrophila encodes more than 30 two-component regulatory
systems, including a new Ntr subcluster of hybrid kinases, nine putative
cold shock proteins and nine potentially cold shock-inducible proteins. A
comparison of D. psychrophila's genome features with those of the only
other published genome from a sulfate reducer, the hyperthermophilic
archaeon Archaeoglobus fulgidus, revealed many striking differences, but
only a few shared features.

<>

<1>Rachinger, M., Volland, S., Meinhardt, F., Daniel, R., Liesegang, H.
<2>First Insights into the Completely Annotated Genome Sequence of Bacillus licheniformis Strain 9945A.
<3>Genome Announcements
<4>1
<5>e00525-13
<6>2013
<7>Strains of the species Bacillus licheniformis are widely used in biotechnology for the
production of enzymes and antibiotics (M. Schallmey, A. Singh, and O. P.
Ward, Can. J. Microbiol. 50:1-17, 2004). However, research and application of B.
licheniformis strains are adversely affected by poor genetic accessibility. Thus,
for a closer inspection of natural competence in B. licheniformis, the genome of
strain 9945A, of which derivatives are known to be naturally competent (C. B.
Thorne and H. B. Stull, J. Bacteriol. 91:1012-1020, 1966), was completely
sequenced and manually annotated.

<>

<1>Rachkevych, N., Sybirna, K., Boyko, S., Boretsky, Y., Sibirny, A.
<2>Improving the efficiency of plasmid transformation in Shewanella oneidensis MR-1 by removing ClaI restriction site.
<3>J. Microbiol. Methods
<4>99
<5>35-37
<6>2014
<7>Here we demonstrate that elimination of ClaI restriction site from the sequence of a plasmid
DNA increases the efficiency of transformation of Shewanella oneidensis MR-1 significantly. To
achieve reliable transformation of S. oneidensis MR-1 plasmids either lacking ClaI site or
isolated from primary transformants of S. oneidensis should be used. (C) 2014 Published by
Elsevier B.V.

<>

<1>Radlinska, M., Bujnicki, J.M.
<2>Cloning of enterohemorrhagic Escherichia coli Phage VT-2 Dam methyltransferase.
<3>Acta Microbiol. Pol.
<4>50
<5>161-167
<6>2001
<7>Enterobacterial GATC-specific DNA adenine methyltransferase plays an essential role in
regulation of DNA replication, methyl-directed mismatch repair, transposition and gene
expression.  In Salmonella typhimurium it has been shown to directly control virulence.  In
this paper we report cloning and expression of the dam gene from the Shiga toxin-producing
VT2-Sa prophage of enterohemorrhagic Escherichia coli O157.  Comparisons of the predicted
amino acid sequence indicates that Dam methyltransferases of E. coli phages VT2-Sa, 933W, T1
and Haemophilus influenzae phage HP1 make up a separate subgroup of adenine-N6
methyltransferases.  These proteins are similar to the gamma subfamily of
amino-methyltransferases in respect to the linear order of sequence motifs and the presence of
the hallmark "NPPY" tetrapeptide.  However, they apparently lack an autonomous
target-recognizing domain at the C-terminus of the catalytic domain and therefore we propose
to dub them as a "mini-gamma" subfamily.

<>

<1>Radlinska, M., Bujnicki, J.M.
<2>Site-directed mutagenesis defines the catalytic aspartate in the active site of the atypical DNA: m4C methyltransferase M.NgoMXV.
<3>Acta Microbiol. Pol.
<4>50
<5>97-105
<6>2001
<7>M.NgoMXV is one of the few atypical DNA:m4C methyltransferases that does not possess a serine
residue in its predicted active site.  We previously reported a homology model of M.NgoMXV and
argued that the aspartate side chain at a corresponding position, similarly to some
DNA:m6A-specific enzymes, is essential for the methyltransferase activity (Radlinska et al.
1999).  Here we reported the corrected amino acid sequence of M.NgoMXV and the analysis of
substitution of D68 with alanine or serine, which both render the enzyme totally inactive.

<>

<1>Radlinska, M., Bujnicki, J.M., Piekarowicz, A.
<2>Structural characterization of two tandemly arranged DNA methyltransferase genes from Neisseria gonorrhoeae MS11: N4-cytosine specific M.NgoMXV and nonfunctional 5-cytosine-type M.NgoMorf2P.
<3>Proteins
<4>37
<5>717-728
<6>1999
<7>Two adjacent genes encoding DNA methyltransferases (MTases) of Neisseria gonorrhoeae MS11, an
active N4-cytosine specific M. NgoMXV and an inactive 5-cytosine type M.NgoMorf2P, were cloned
into Escherichia coli and sequenced. We analyzed the deduced amino acid sequence of both gene
products and localized conserved regions characteristic for DNA MTases. Structure prediction,
threading-derived alignments, and comparison with the common fold for DNA MTases allowed for
construction of super-secondary and tertiary models for M.NgoMorf2P and M.NgoMXV,
respectively. These models helped in identification of amino acids and structural elements
essential for function of both enzymes. The implications of this putative structural model on
the catalytic mechanism of M.NgoMXV and its possible relation to the common ancestor of modern
DNA amino-MTases are also discussed.

<>

<1>Radlinska, M., Kondrzycka-Dada, A., Piekarowicz, A., Bujnicki, J.M.
<2>Identification of amino acids important for target recognition by the DNA:m(5)C methyltransferase M.NgoPII by alanine-scanning mutagenesis  of residues at the protein-DNA interface.
<3>Proteins
<4>58
<5>263-270
<6>2005
<7>DNA:m(5)C MTases comprise a catalytic domain with conserved residues of the active site and a
strongly diverged TRD with variable residues
involved in DNA recognition and binding. To date, crystal structures of
2 DNA:m(5)C MTases complexed with the substrate DNA have been obtained;
however, for none of these enzymes has the importance of the whole set
of DNA-binding residues been comprehensively studied. We built a
comparative model of M.NgoPII, a close homologue and isomethylomer of
M.HaeIII, and systematically analyzed the effect of alanine
substitutions for the complete set of amino acid residues from its TRD
predicted to be important for DNA binding and target recognition. Our
data demonstrate that only 1 Arg residue is indispensable for the MTase
activity in vivo and in vitro, and that mutations of only a few other
residues cause significant reduction of the activity in vitro, with
little effect on the activity in vivo. The identification of
dispensable protein-DNA contacts in the wild-type MTase will serve as a
platform for exhaustive combinatorial mutagenesis aimed at the design
of new contacts, and thus construction of enzyme variants that retain
the activity but exhibit potentially new substrate preferences.

<>

<1>Radlinska, M., Piekarowicz, A.
<2>Cloning and characterization of the gene encoding a new DNA methyltransferase from Neisseria gonorrhoeae.
<3>Biol. Chem.
<4>379
<5>1391-1395
<6>1998
<7>A HindIII fragment of N. gonorrhoeae MS11 DNA coding for DNA methyltransferase activity was
cloned and expressed in E. coli AP1-200-9 cells.  The sequence of 4681 bp was determined, and
its analysis revealed two open reading frames sharing some similarity with known DNA MTases.
ORF1 encodes an active N4mC MTase (M.NgoMV).  The enzyme modifies only one strand of double
stranded DNA and preferentially recognizes the sequence GCCHR although it is able to methylate
other sites.  The exact recognition sequence cannot be precisely defined due to a relaxed
specificity.  The second ORF shows high homology to 5mC Mtases, but we were unable to
demonstrate DNA methylating activity of its product either in vivo or in vitro.

<>

<1>Radlinska, M., Piekarowicz, A., Galimand, M., Bujnicki, J.M.
<2>Cloning and preliminary characterization of a GATC-specific beta(2)-class DNA:m(6)A methyltransferase encoded by transposon Tn1549  from Enterococcus spp.
<3>Pol. J. Microbiol.
<4>54
<5>249-252
<6>2005
<7>A recent study revealed a subfamily of N6-adenine (m(6)A) methyltransferases that comprises a
few functionally studied eukaryotic
members acting on mRNA and prokaryotic members acting on DNA as well as
numerous uncharacterized open reading frames. Here, we report cloning
and functional characterization of a prokaryotic member of this family
encoded by transposon Tn1549 from Enterococcus spp.

<>

<1>Radlinska, M., Skowronek, K.
<2>Novel procedure for the detection of 5-methylcytosine.
<3>Acta Microbiol. Pol.
<4>47
<5>327-334
<6>1998
<7>Bisulfite converts non-methylated cytosine in DNA to uracil leaving 5-methylcytosine
unaltered.  In this communication, we present a new approach omitting the conventional PCR
amplification step.  Bisulfite-converted methylated DNA is directly sequenced.  The
effectiveness of the new protocol is demonstrated by using it for the detection of
5-methylation of cytosine residues introduced by three different DNA methyltransferases
(M.HaeIII, M.HpaII and M.HhaI).  A simple experimental system useful to determine the sequence
specificity of DNA methyltransferases is also presented.

<>

<1>Radnedge, L., Pinney, R.J.
<2>Ultraviolet light induction of lambda from dcm host strains alleviates EcoRII restriction of phage.
<3>FEMS Microbiol. Lett.
<4>121-125
<5>121-126
<6>1991
<7>A new form of restriction alleviation is demonstrated for phage induced by ultraviolet light
from dcm strains of Escherichia coli K-12. EcoRII restriction of the induced phage is
alleviated, which is the first report of Type II restriction alleviation. Unlike previously
reported restriction alleviation, the increase in phage-plating efficiency is not dependent
upon irradiation of the plating host for its induction.

<>

<1>Raftis, E.J., Forde, B.M., Claesson, M.J., O'Toole, P.W.
<2>Unusual genome complexity in Lactobacillus salivarius JCM1046.
<3>BMC Genomics
<4>15
<5>771
<6>2014
<7>BACKGROUND: Lactobacillus salivarius strains are increasingly being exploited for
their probiotic properties in humans and animals. Dissemination of antibiotic
resistance genes among species with food or probiotic-association is undesirable
and is often mediated by plasmids or integrative and conjugative elements. L.
salivarius strains typically have multireplicon genomes including circular
megaplasmids that encode strain-specific traits for intestinal survival and
probiotic activity. Linear plasmids are less common in lactobacilli and show a
very limited distribution in L. salivarius. Here we present experimental evidence
that supports an unusually complex multireplicon genome structure in the porcine
isolate L. salivarius JCM1046. RESULTS: JCM1046 harbours a 1.83 Mb chromosome,
and four plasmids which constitute 20% of the genome. In addition to the known
219 kb repA-type megaplasmid pMP1046A, we identified and experimentally validated
the topology of three additional replicons, the circular pMP1046B (129 kb), a
linear plasmid pLMP1046 (101 kb) and pCTN1046 (33 kb) harbouring a conjugative
transposon. pMP1046B harbours both plasmid-associated replication genes and
paralogues of chromosomally encoded housekeeping and information-processing
related genes, thus qualifying it as a putative chromid. pLMP1046 shares limited
sequence homology or gene synteny with other L. salivarius plasmids, and its
putative replication-associated protein is homologous to the RepA/E proteins
found in the large circular megaplasmids of L. salivarius. Plasmid pCTN1046
harbours a single copy of an integrated conjugative transposon (Tn6224) which
appears to be functionally intact and includes the tetracycline resistance gene
tetM. CONCLUSION: Experimental validation of sequence assemblies and plasmid
topology resolved the complex genome architecture of L. salivarius JCM1046. A
high-coverage draft genome sequence would not have elucidated the genome
complexity in this strain. Given the expanding use of L. salivarius as a
probiotic, it is important to determine the genotypic and phenotypic organization
of L. salivarius strains. The identification of Tn6224-like elements in this
species has implications for strain selection for probiotic applications.

<>

<1>Raghavendra, N.K., Bheemanaik, S., Rao, D.N.
<2>Mechanistic insights into type III restriction enzymes.
<3>Front. Biosci.
<4>17
<5>1094-1107
<6>2012
<7>Type III restriction-modification (R-M) enzymes need to interact with two separate
unmethylated DNA sequences in indirectly repeated, head-to-head orientations for efficient
cleavage to occur at a defined location next to only one of the two sites. However, cleavage
of sites that are not in head-to-head orientation have been observed to occur under certain
reaction conditions in vitro. ATP hydrolysis is required for the long-distance communication
between the sites prior to cleavage. Type III R-M enzymes comprise two subunits, Res and Mod
that form a homodimeric Mod(2) and a heterotetrameric Res(2)Mod(2) complex. The Mod subunit in
M-2 or R2M2 complex recognizes and methylates DNA while the Res subunit in R2M2 complex is
responsible for ATP hydrolysis, DNA translocation and cleavage. A vast majority of biochemical
studies on Type III R-M enzymes have been undertaken using two closely related enzymes, EcoP1I
and EcoP15I. Divergent opinions about how the long-distance interaction between the
recognition sites exist and at least three mechanistic models based on 1D- diffusion and/or
3D-DNA looping have been proposed.

<>

<1>Raghavendra, N.K., Rao, D.N.
<2>Functional cooperation between exonucleases and endonucleases - basis for the evolution of restriction enzymes.
<3>Nucleic Acids Res.
<4>31
<5>1888-1896
<6>2003
<7>Many types of restriction enzymes cleave DNA away from their recognition site. Using the type
III restriction enzyme, EcoP15I, which cleaves DNA
25-27 bp away from its recognition site, we provide evidence to show that
an intact recognition site on the cleaved DNA sequesters the restriction
enzyme and decreases the effective concentration of the enzyme. EcoP15I
restriction enzyme is shown here to perform only a single round of DNA
cleavage. Significantly, we show that an exonuclease activity is essential
for EcoP15I restriction enzyme to perform multiple rounds of DNA cleavage.
This observation may hold true for all restriction enzymes cleaving DNA
sufficiently far away from their recognition site. Our results highlight
the importance of functional cooperation in the modulation of enzyme
activity. Based on results presented here and other data on
well-characterised restriction enzymes, a functional evolutionary
hierarchy of restriction enzymes is discussed.

<>

<1>Raghavendra, N.K., Rao, D.N.
<2>Unidirectional translocation from recognition site and a necessary interaction with DNA end for cleavage by Type III restriction enzyme.
<3>Nucleic Acids Res.
<4>32
<5>5703-5711
<6>2004
<7>Type III restriction enzymes have been demonstrated to require two unmethylated asymmetric
recognition sites oriented head-to-head to elicit
double-strand break 25-27 bp downstream of one of the two sites. The
proposed DNA cleavage mechanism involves ATP-dependent DNA translocation.
The sequence context of the recognition site was suggested to influence
the site of DNA cleavage by the enzyme. In this investigation, we
demonstrate that the cleavage site of the R.EcoP15I restriction enzyme
does not depend on the sequence context of the recognition site.
Strikingly, this study demonstrates that the enzyme can cleave linear DNA
having either recognition sites in the same orientation or a single
recognition site. Cleavage occurs predominantly at a site proximal to the
DNA end in the case of multiple site substrates. Such cleavage can be
abolished by the binding of Lac repressor downstream (3' side) but not
upstream (5' side) of the recognition site. Binding of HU protein has also
been observed to interfere with R.EcoP15I cleavage activity. In accordance
with a mechanism requiring two enzyme molecules cooperating to elicit
double-strand break on DNA, our results convincingly demonstrate that the
enzyme translocates on DNA in a 5' to 3' direction from its recognition
site and indicate a switch in the direction of enzyme motion at the DNA
ends. This study demonstrates a new facet in the mode of action of these
restriction enzymes.

<>

<1>Raghavendra, N.K., Rao, D.N.
<2>Exogenous AdoMet and its analogue sinefungin differentially influence DNA cleavage by R.EcoP15I-Usefulness in SAGE.
<3>Biochem. Biophys. Res. Commun.
<4>334
<5>803-811
<6>2005
<7>While it has been demonstrated that AdoMet is required for DNA cleavage by Type III
restriction enzymes, here we show that in the presence of
exogenous AdoMet, the head-to-head oriented recognition sites are cleaved
only on a supercoiled DNA. On a linear DNA, exogenous AdoMet strongly
drives methylation while inhibiting cleavage reaction. Strikingly, the AdoMet
analogue sinefungin results in cleavage at all recognition sites
irrespective of the topology of DNA. The cleavage reaction in the presence
of sinefungin is ATP dependent. The site of cleavage is comparable with
that in the presence of AdoMet. The use of EcoP15I restriction in presence
of sinefungin as an improved tool for serial analysis of gene expression
is discussed.

<>

<1>Rai, K.
<2>Roles of DNA and histone methyltransferases during zebrafish development.
<3>Ph.D. Thesis, Univ. of Utah
<4>
<5>1-160
<6>2006
<7>The research work presented in this dissertation describes the roles of the major DNA
methyltransferases and histone methyltransferases in zebrafish development.  DNA and histone
methylation are two key processes that regulate transcription of genes in mammals.  Both of
these processes are essential for proper development of an organism and their misregulation
leads to multiple diseases including cancer.  A better understanding of these two processes
will help us understand how an organism develops and will also help in designing more
effective drugs against cancer.  Chapter 1 is an introduction on DNA and histone methylation
and the enzymes which catalyze these processes.  This chapter summarizes the literature on the
normal various organisms including zebrafish, how abnormalities in this process lead to
cancer, the protein structure and function of the known DNA methyltransferases and findings
about relationship between histone methylation and DNA methylation.  Chapter 2 describes the
function of DNA methyltransferase-1 during zebrafish development.  The focus of the studies
described in this chapter is on the function of this enzyme during later differentiation
during tissue-specific development.  Also, the epistatic relationship between Dnmt1 and
Suv39h1, the major H3K9 methyltransferase, is described in this chapter.  Chapter 3 shows our
findings about role of Dnmt3, a member of other sub-class of DNA methyltransferase family,
during zebrafish development.  Here I have shown novel roles of this enzyme during development
of vertebrate brain.  In Chapter 4, I have described our novel findings regarding the role of
Dnmt2, a mysterious DNA/RNA methyltransferase, during zebrafish development.  Here, I have
also described the importance of its function in the cytoplasm.  Chapter 5 is an attempt to
compare the phenotypes obtained by knocking-down protein levels of different DNA
methyltransferases.  This chapter is the crux of our studies which shows striking similarities
and differences of in vivo functions of different classes of DNA methyltransferases.  In
Chapter 6, the main conclusions of my work are summarized.  I have also discussed the
implications of our findings and the future approaches to unravel the mysteries of
developmental functions of DNA methyltransferases.

<>

<1>Raiol, T., De-Souza, M.T., Oliveira, J.V., Silva, H.S., Orem, J.C., Cavalcante, D.A., Almeida, N.F., Telles, G.P., Setubal, J.C., Brigido, M.M., Torres, F.A., Stadler, P.S., Walter, M.E., Moraes, L.M.
<2>Draft Genome Sequence of FT9, a Novel Bacillus cereus Strain Isolated from a Brazilian Thermal Spring.
<3>Genome Announcements
<4>2
<5>e01027-14
<6>2014
<7>A Bacillus cereus strain, FT9, isolated from a hot spring in the midwest region of Brazil, had
its entire genome sequenced.

<>

<1>Raiol, T., Ribeiro, G.M., Maranhao, A.Q., Bocca, A.L., Silva-Pereira, I., Junqueira-Kipnis, A.P., Brigido, M.M., Kipnis, A.
<2>Complete Genome Sequence of Mycobacterium massiliense.
<3>J. Bacteriol.
<4>194
<5>5455
<6>2012
<7>Mycobacterium massiliense is a rapidly growing bacterium associated with opportunistic
infections. The genome of a representative isolate (strain GO 06)
recovered from wound samples from patients who underwent arthroscopic or
laparoscopic surgery was sequenced. To the best of our knowledge, this is the
first announcement of the complete genome sequence of an M. massiliense strain.

<>

<1>Raizis, A.M., Schmitt, F., Jost, J.-P.
<2>A bisulfite method of 5-methylcytosine mapping that minimizes template degradation.
<3>Anal. Biochem.
<4>226
<5>161-166
<6>1995
<7>The bisulfite method is a highly sensitive approach to 5-methylcytosine mapping that utilizes
the capability of the polymerase chain reaction to exponentially amplify DNA. We have observed
that the bisulfite reaction results in a significant level of template degradation due to DNA
depurination.  Furthermore, our data suggest that the DNA fragmentation which occurs limits
the sensitivity of the method.  We describe a simple solution to limit degradation of the DNA
template.

<>

<1>Raja, M.C., Dharmalingam, K.
<2>Heat shock-induced relaxation of restriction enzyme specificity in Escherichia coli.
<3>J. Genetics
<4>70
<5>91-101
<6>1991
<7>Methylated and hydroxymethylated cytosine containing DNA was restricted by
proteins encoded by the mcrBC (rglB) loci of E. coli.  In vivo, RglB proteins
recognize and cleave hmCT2 and hmCT4 DNAs at 30C and 42C but hmCT6 DNA was
unaffected at both temperatures.  However, cells carrying the rglB genes cloned
on pBR322 (pDSS17) did not restrict hmCT6 at 30C, but hmCT6 DNA was cleaved
efficiently at 42C.  Heat shock treatment for five minutes was enough to induce
this promiscuity in recognition specificity.  We call this activity RglB star.
A single copy of rglB located on the chromosome or cloned on a low copy vector
pMU575 failed to show RglB star activity.  De novo protein synthesis was not
required for the manifestation of RglB star activity.

<>

<1>Rajagopal, K.
<2>Draft Genome Sequence of Enterococcus raffinosus Strain CFTRI 2200, Isolated from Infant Fecal Material.
<3>Genome Announcements
<4>1
<5>e00932-13
<6>2013
<7>The complete genome sequence of Enterococcus raffinosus strain CFTRI 2200, isolated from the
fecal material of a 7-month-old infant, is reported. The
complete genome consists of 4.237 Mbp with a G+C content of 39.47% and 4,242
protein-coding genes, 54 tRNAs, and 46 rRNAs.

<>

<1>Rajanna, C., Revazishvili, T., Rashid, M.H., Chubinidze, S., Bakanidze, L., Tsanava, S., Imnadze, P., Bishop-Lilly, K.A., Sozhamannan, S., Gibbons, H.S., Morris, J.G., Sulakvelidze, A.
<2>Characterization of pPCP1 Plasmids in Yersinia pestis Strains Isolated from the Former Soviet Union.
<3>Int. J. Microbiol.
<4>2010
<5>760-819
<6>2010
<7>Complete sequences of 9.5-kb pPCP1 plasmids in three Yersinia pestis strains from the former
Soviet Union (FSU) were determined and compared with those of pPCP1 plasmids in three
well-characterized, non-FSU Y.
pestis strains (KIM, CO92, and 91001). Two of the FSU plasmids were from strains C2614 and
C2944, isolated from plague foci in Russia, and one plasmid was from strain C790 from
Kyrgyzstan. Sequence analyses identified four sequence types among the six plasmids. The pPCP1
plasmids in the FSU strains were most genetically related to the pPCP1 plasmid in the KIM
strain and least related to the pPCP1 plasmid in Y. pestis 91001. The FSU strains generally
had larger pPCP1 plasmid copy numbers compared to strain CO92. Expression of the plasmid's
pla gene was significantly (P </= .05) higher in strain C2944 than in strain CO92. Given
pla's role in Y. pestis virulence, this difference may have important implications for the
strain's virulence.

<>

<1>Rajski, S.R., Kumar, S., Roberts, R.J., Barton, J.K.
<2>Protein-modulated DNA electron transfer.
<3>J. Am. Chem. Soc.
<4>121
<5>5615-5616
<6>1999
<7>Long-range oxidative damage to the 5'-guanine of 5'-GG-3' sequences in DNA readily occurs
as a result of electron migration through the pi-stack on long-range electron transfer, since
upon binding, the protein induces and stabilizes a pi-gap using a novel DNA base-flipping
mechanism.  Long-range oxidation of 5'-GG-3' sites was first shown with a rhodium
intercalator.  The rhodium photochemistry bound to DNA yields base photooxidation upon
irradiation at low energy (365nm), whereas irradiation at high energy (313 nm) leads to direct
strand scission, marking the sites of intercalation.  Other DNA-bound photooxidants also
promote DNA damage at long range.

<>

<1>Raleigh, E., Dila, D., Sutherland, E., Kelleher, J., Moran, L., Slatko, B., Briggs, P.
<2>McrBC, a novel multisubunit GTP-dependent restriction endonuclease.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>152
<6>1993
<7>The McrBC system is one of four restriction systems used by E. coli K-12 to monitor the origin
of invading DNA and determine its fate. Like McrA and Mrr, the McrBC system is specific for
modified DNA. The system is encoded by two genes of low GC composition flanked by two similar
dyad symmetries, suggesting that the system may have been imported from elsewhere. Both genes
are required for restriction in vivo of a variety of modified targets, including those with
5-methylcytosine, 5-hydroxymethylcytosine and N4-methylcytosine. A few modified targets are
sensitive to restriction mediated by mcrB in the absence of mcrC. Three proteins are expressed
from the two genes. Only two of the three are required for in vitro activity. The in vitro
cleavage activity reflects the in vivo properties of the system in its requirement for a
modified substrate and in the spectrum of site-specific modifications that are sensitive to
cleavage. GTP is required for cleavage. Non-hydrolysable analogues of GTP inhibit the
reaction, as does ATP. Our current model is that cleavage requires the sequence
RmC(N40-80)RmC, with multiple cleavage positions on both strands distributed within the spacer
region. The roles played by the two proteins are under investigation genetically. Twelve mcrB
mutants with dominant phenotypes have been isolated. These fall into three phenotypic classes.
All are defective in McrC-dependent restriction; they differ in their ability in vivo to
inhibit restriction by wild type genes in trans and in their ability to carry out
McrC-independent restriction. Sequence analysis reveals that each class corresponds to a
particular portion of the protein, one of which is the GTP-binding site motif identified in
the polypeptide sequence of McrB.

<>

<1>Raleigh, E.A.
<2>Restriction and modification in vivo by Escherichia coli K12.
<3>Methods Enzymol.
<4>152
<5>130-141
<6>1987
<7>This chapter focuses on how restriction of newly introduced DNA by Escherichia coli can
interfere with cloning and subcloning work, and in particular on how the pattern of
methylation of the DNA influences this. A principal aim is to acquaint the reader with three
E. coli restriction systems that attack DNA only when it is appropriately methylated. The
second part of this chapter describes biological restriction and modification in general
terms. The third part discusses the particular restriction systems found in E. coli K12, first
briefly the familiar K and P1 restriction systems, and then in detail the methyl-specific
McrA, McrB and Mrr systems. Some common strains are discussed with special reference to their
restriction phenotypes. The fourth part briefly reviews the E. coli methylation functions, Dam
and Dcm, as they affect sensitivity to digestion of DNA in vitro.

<>

<1>Raleigh, E.A.
<2>Organization and function of the mcrBC genes of Escherichia coli K-12.
<3>Mol. Microbiol.
<4>6
<5>1079-1086
<6>1992
<7>Many natural DNA sequences are restricted in Escherichia coli K-12, not only by the classic
Type I restriction system EcoK, but also by one of three modification-specific restriction
systems found in K-12. The McrBC system is the best studied of these. We infer from the base
composition of the mcrBC genes that they were imported from an evolutionarily distant source.
The genes are located in a hypervariable cluster of restriction genes that may play a
significant role in generation of species identity in enteric bacteria. Restriction activity
requires the products of two genes for activity both in vivo and in vitro. The mcrB gene
elaborates two protein products, only one of which is required for activity in vitro, but both
of which contain a conserved amino acid sequence motif identified as a possible GTP-binding
site. The mcrC gene product contains a leucine heptad repeat that could play a role in
protein-protein interactions. McrBC activity in vivo and in vitro depends on the presence of
modified cytosine in a specific sequence context; three different modifications are
recognized. The in vitro activity of this novel multi-subunit restriction enzyme displays an
absolute requirement for GTP as a cofactor.

<>

<1>Raleigh, E.A., Benner, J., Bloom, F., Braymer, H.D., DeCruz, E., Dharmalingam, K., Heitman, J., Noyer-Weidner, M., Piekarowicz, A., Kretz, P.L., Short, J.M., Woodcock, D.
<2>Nomenclature relating to restriction of modified DNA in Escherichia coli.
<3>J. Bacteriol.
<4>173
<5>2707-2709
<6>1991
<7>At least three restriction systems that attack DNA containing naturally
modified bases have been found in common Escherichia coli K-12 strains.  These
systems are McrA, McrBC, and Mrr.  A brief summary of the genetic and phenotype
properties so far observed in laboratory strains is set forth, together with a
proposed nomenclature for describing these properties.

<>

<1>Raleigh, E.A., Brooks, J.E.
<2>Restriction modification systems: where they are and what they do.
<3>Bacterial Genomes, Chapman & Hall, De Bruijn, F.J., Lupski, J.R., Weinstock, G.M., New York
<4>
<5>78-92
<6>1998
<7>The review concentrates on restriction-modification in the context of bacterial genome
evolution and how the systems affect bacterial populations.  RM systems regulate the entry of
foreign DNA into cells.  A model of how the systems work is shown in Figure 8-1.  Foreign DNA
is restricted by a restriction endonuclease that recognizes a specific sequence and cleaves
the DNA unless the sequence is protected. Typically, a modification methyltransferase confers
protection, by methylating a particular base within the sequence recognized by the restriction
enzyme, thereby rendering it resistant to cleavage.  Alternatively, however, some restriction
enzymes recognize a sequence only when it is methylated.  In this case, methylation of a
suitable base confers sensitivity to restriction and protection arises from failure to
methylate the relevant sequence.  Both sorts of restriction can act to limit the transfer of
DNA into cells.  One key feature of RM is that the systems can be effective only if they are
variable and fluid within a bacterial population.  Modifications made to foreign DNA escaping
restriction are epigenetic, i.e., not heritable.

<>

<1>Raleigh, E.A., Hauck, E.S.
<2>Process for producing recombinant McrBC endonuclease and cleavage of methylated DNA.
<3>US Patent Office
<4>US 5405760
<5>
<6>1995
<7>The present invention relates to a recombinant McrBC endonuclease obtainable from Escherichia
coli, two components of which, McrBL and McrC, have been purified in active form. McrBC is
active in the presence of GTP and at a low pH. The McrBC is active in the presence of GTP and
at a low pH. The McrBC endonuclease is also substantially free of a third component, McrBs,
which is believed to inhibit or otherwise interfere with the activity of the enzyme. McrBC has
various desirable properties, including the ability to recognize a methylated DNA sequence and
also its ability to cleave such a sequence in the presence of GTP. Also provided is a process
for the production of recombinant McrBC endonuclease, a process for the determination of the
modification state of DNA, a process for the determination of an epigenetic alteration or
defect (including "imprinting"), as well as a process for identifying and isolating additional
enzymes that cleave modified DNA.

<>

<1>Raleigh, E.A., Murray, N.E., Revel, H., Blumenthal, R.M., Westaway, D., Reith, A.D., Rigby, P.W.J., Elhai, J., Hanahan, D.
<2>McrA and McrB restriction phenotypes of some E. coli strains and implications for gene cloning.
<3>Nucleic Acids Res.
<4>15
<5>1563-1575
<6>1988
<7>The McrA and McrB (modified cytosine restriction) systems of E. coli interfere
with incoming DNA containing methylcytosine.  DNA from many organisms,
including all mammalian and plant DNA, is expected to be sensitive, and this
could interfere with cloning experiments.  The McrA and B phenotypes of a few
strains have been reported previously.  The Mcr phenotypes of 94 strains,
primarily derived from E. coli K12, are tabulated here.  We briefly review some
evidence suggesting that McrB restriction of mouse-modified DNA does occur in
vivo and does in fact interfere with cloning of specific mouse sequences.

<>

<1>Raleigh, E.A., Sutherland, E., Dila, D., Briggs, P., Kelleher, J., Coe, L., Slatko, B., Moran, L.
<2>Molecular analysis of McrBC, a GTP-dependent restriction endonuclease from E. coli K-12.
<3>J. Cell Biochem. Suppl.
<4>16B
<5>21
<6>1992
<7>The McrBC system is one of three modification-dependent restriction systems used by E. coli
K-12 to monitor the origin of invading DNA and determine its fate. The system is encoded by
two genes of low GC composition flanked by two similar dyad symmetries, suggesting that the
system may have been imported from elsewhere. Both genes are required for restriction in vivo
of a variety of modified targets, including those with 5-methylcytosine,
5-hydroxymethylcytosine and N4-methylcytosine. Three proteins are expressed from the two
genes, two of which are required for in vitro activity. For each of these proteins (McrBL and
McrC), constructs that forced translation initiation at either of two potential start codons
yielded enzymatically active product. One of each was purified to >90% purity. The in vitro
cleavage activity reflected the in vivo properties of the system in its requirement for a
modified substrate and in the spectrum of site-specific modifications that were sensitive to
cleavage. GTP was required for cleavage. Non-hydrolysable analogues of GTP inhibited the
reaction, as did ATP. We are unaware of any other nuclease with an absolute requirement for a
guanosine nucleotide. The nature of the cleavage site was examined further by mapping sites on
natural substrates, delimiting the cleavage site by primer extension, and cleaving synthetic
oligonucleotide model substrates. Different sites are cleaved with differing efficiencies. Our
current model is that cleavage requires the sequence RmC(N40-70)RmC, with multiple cleavage
positions on both strands distributed within the spacer region.

<>

<1>Raleigh, E.A., Trimarchi, R., Revel, H.
<2>Genetic and physical mapping of the mcrA (rglA) and mcrB (rglB) loci of Escherichia coli K-12.
<3>Genetics
<4>122
<5>279-296
<6>1989
<7>We have genetically analyzed, cloned and physically mapped the modified
cytosine-specific restriction determinants mcrA (rglA) and mcrB (rglB) of
Escherichia coli K-12.  The independently discovered Rgl and Mcr restriction
systems are shown to be identical by three criteria:  1) mutants with the RglA-
or RglB- phenotypes display the corresponding McrA- or McrB- phenotypes, and
vice versa; 2) the gene(s) for RglA and McrA reside together at one locus,
while gene(s) for RglB and McrB are coincident at a different locus; and 3)
RglA+ and RglB+ recombinant clones complement for the corresponding
Mcr-deficient lesions.  The mcrA (rglA) gene(s) is on the excisable element
e14, just clockwise of purB at 25 min.  The mcrB (rglB) gene(s), at 99 min, is
in a cluster of restriction functions that includes hsd and mrr, determinants
of host-specific restriction (EcoK) and methyladenine-specific restriction
respectively.  Gene order is mcrB-hsdS-hsdM-hsdR-mrr-serB.  Possible models for
the acquisition of these restriction determinants by enteric bacteria are
discussed.

<>

<1>Raleigh, E.A., Vaisvila, R., Morgan, R.D.
<2>Restriction enzyme gene discovery method.
<3>International Patent Office
<4>WO 9964632
<5>
<6>1999
<7>The invention is directed to direct cloning of intact genes, with a high probability that the
orientation of expression is known in advance, and with a low probability of being associated
with extraneous possibly toxic genes.  The invention is particularly directed to obtaining
genes encoded in DNA cassettes comprised of repeat sequences flanking variable open reading
frames.  The invention encompasses obtaining such cassette-encoded genes using
oligonucleotides hybridizing to the repeated elements, cloning them and expressing them.
Expression may employ tightly regulated vectors and useful strains disclosed.  Methods for
identifying restriction endonuclease and DNA methyltransferase genes in the absence of prior
information about the sequences or biochemical specificities of these are also disclosed.

<>

<1>Raleigh, E.A., Vaisvila, R., Morgan, R.D.
<2>Method of finding restriction enzyme.
<3>Japanese Patent Office
<4>JP 2002517260 A
<5>
<6>2002
<7>
<>

<1>Raleigh, E.A., Wilson, G.
<2>Escherichia coli K-12 restricts DNA containing 5-methylcytosine.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>83
<5>9070-9074
<6>1986
<7>We have observed that plasmids containing certain cloned modification methylase genes of type
II restriction-modification systems cannot be transformed into many laboratory strains of
Escherichia coli K-12.  The investigation of this phenomenon, reported here, has revealed (i)
DNA containing 5-methylcytosine is biologically restricted by these strains, while DNA
containing 6-methyladenine is not; (ii) restriction is due to two genetically distinct systems
that differ in their sequence specificities, which we have named mcrA and mcrB (for modified
cytosine restriction).  Since 5-methylcytosine containing DNA is widespread in nature, the Mcr
systems probably have a broad biological role. Mcr restriction may seriously interfere with
molecular cloning of 5-methylcytosine-containing foreign DNAs.  The Mcr phenotypes of some
commonly used strains of E. coli K-12 are reported.

<>

<1>Ralph, D., Que, Q., Van Etten, J.L., McClelland, M.
<2>Leptospira genomes are modified at 5'-GTAC.
<3>J. Bacteriol.
<4>175
<5>3913-3915
<6>1993
<7>Genomic DNAs of 14 strains from seven species of the spirochete Leptospira were resistant to
cleavage by the restriction endonuclease RsaI (5'-GTAC). A modified base comigrating with m4C
was detected by chromatography. Genomic DNAs from other spirochetes, Borrelia group VS461, and
Serpulina strains were not resistant to RsaI digestion. Modification at 5'-GTAm4C may occur
in most or all strains of all species of Leptospira but not in all genera of spirochetes.
Genus-wide DNA modification has rarely been observed in bacteria.

<>

<1>Ralston, D.J., Baer, B.S.
<2>A new property of phage group II Staphylococcus aureus strains:  Host restriction of phage K14.
<3>J. Gen. Microbiol.
<4>36
<5>1-16
<6>1964
<7>Various strains of Staphylococcus aureus which type exclusively with phages of
lytic group II were found to modify phage K14 so that its ability to form
plaques on host K1N was lessened.  The restricted phage formed plaques with
high efficiency on all strains of lytic group II.  In general, it plated at
lower titres on strains of lytic groups I, II, IV, and on some strains of
miscellaneous typing characteristics; however, there were some variations among
separate cultures of the same strains.  For example, the restricted phage
plated at high titre on strains 52A/79a, 73, and 44A, but formed significantly
fewer numbers of plaques on strain 52A, 79b and on a second culture of 44A.
Strain K1N was found to dissociate into apt (K1H1) and non-apt (K1N2) forms.
The probability of plaque development by restricted phage on strain K1N was
dependent upon the nutritional state of the cocci and also upon the proportion
of apt and non-apt cells.  The restriction of phage K14 was eliminated during
its propagation on all strains other than lytic group II.  The unrestricted
progeny particles tended to assay at equal titre on all the indicator strains.
In all cases the genotype of the phage - susceptibility to
host-control-remained unchanged.  The observations add to existing data which
indicate that strains of phage group II form a genetically distinct group.  The
suggestion is made that this phenomenon might help in taxonomic classifiction
of strains of S. aureus.

<>

<1>Ralston, D.J., Kruger, A.P.
<2>Phage multiplication on two hosts, isolation and activity of variants of Staphylococcus phage P1.
<3>Proc. Soc. Exp. Biol. Med.
<4>80
<5>217-220
<6>1952
<7>Analysis of phage activity by titration on two hosts has revealed the presence
of variants hitherto undetected in stock phage P1.  One isolate, Phage 14,
exhibits great changes in titration ratio on passage through the two hosts K1
and WF 145.  Produced on K1 cells, the ratio of free phage assayed on two hosts
is 1.3, whereas made on 145 cells,the free phage titrates in a ratio 145/K1 of
ca 40.  This occurs regardless of the host employed in previous passage and is
reproduced in the very first burst from infected cells.  Attempts to isolate
different strains from this phage by usual plaque isolation technics were not
successful.  The high ratio obtained with free phage made on 145 cells could
not be explained on the basis of differences in adsorption onto or latent
periods in the two hosts.  The difference was traced to the production of a
large number of particles from host 145 which adsorbed on K1 but formed no
plaques.  No evidence was found for an inhibitor of K1 activity associated with
phage 14 production on 145 cells.  Heat inactivation destroyed phage activity
for K1 cells much more rapidly than for 145 cells.  There was no interaction of
heat killed and active phage on exposure to 59C. Evidence has been accumulated
which indicates that the phage P14 is altered on passage through host 145.  The
fact that phage particles surviving a heat treatment which destroyed all
activity for K1 cells produce on strain 145 a mixture of two phages (one active
on K1 and on - or both - active on strain 145) points to an unusual host effect
on virus reproduction.

<>

<1>Ramakrishna, J., Mathee, K., Plaut, A.G., Wright, A.
<2>Restriction enzyme systems of Helicobacter pylori.
<3>Gastroenterology
<4>108
<5>A200
<6>1995
<7>To date, the analysis of H. pylori using methods such as RFLP's, PCR and pulsed field gel
electrophoresis, has revealed considerable genetic diversity from strain to strain.  Due to
this diversity, there are as yet no methods for classifying this organism.  Also there are
considerable differences in the transformability of different clinical isolates, some being
easily transformable while others are not.  It has therefore been postulated that the organism
may have restriction-modification systems which differ among isolates.  Restriction enzymes
are produced by bacteria that cleave DNA at specific recognition sequences; these bacteria
therefore have a modification system to methylate their own DNA at the recognition sites to
protect it from cleavage.  No restriction enzymes have been described in H. pylori so far.
Since restriction-modification systems are highly conserved in a given strain of a pathogenic
organism, this can be a reliable method for classifying clinical isolates.  We cultured eleven
H. pylori isolates on Campylobacter agar Skirrow (Difco) plates supplemented with 10%
defibrinated sheep blood (Remel) in microaerobic chambers.  Cells were resuspended in a
suitable buffer and sonicated.  Protein extracts were prepared by precipitation with 60%
ammonium sulfate and subsequent dialysis, and these were used for restriction digests.
Preliminary digests were done using a standard DNA template, the plasmid pBR322, which has a
known restriction map and DNA sequence.  The patterns of these digests suggested the existence
of restriction enzymes.  The site specificity of the restriction enzymes was elucidated using
pBR322 that was linearized and labelled at one end with 32P.  Partial digests of the 32P
labelled DNA with Hp extracts were fractionated by electrophoresis on polyacrylamide and
agarose gels alongside appropriate radiolabelled markers.  By determining the sizes of the
fragments of DNA generated we were able to determine the restriction sites on pBR322.  The
various clinical isolates studied by us so far all have distinct restriction patterns.  Two
have so far been identified.  The enzyme present in Hp 32 has a specificity corresponding to
Sau96I while that in Hp 64 corresponds to DdeI.  Work is currently under way in our laboratory
to identify the restriction enzymes in other clinical isolates.  We anticipate that this will
yield a pattern that will form the bais for a classification system for H. pylori strains.

<>

<1>Ramalingam, R., Prasad, R., Shivapriya, R., Dharmalingam, K.
<2>Molecular cloning and sequencing of mcrA locus and identification of McrA protein in Escherichia coli.
<3>J. Biosci.
<4>17
<5>217-232
<6>1992
<7>The Mcr systems (previously known as Rgl systems) of Escherichia coli recognize and cleave
specific sequences carrying methylated or hydroxymethylated cytosines.  We have cloned the
mcrA gene and determined its nucleotide sequence.  An 831 base pair sequence encodes the McrA
protein.  Analysis of the sequence data reveals that there are additional ORFs internal to the
above.  A phage T7 expression system was used to determine the protein products encoded by the
cloned mcrA gene.  The results clearly show that a 31 kDa polypeptide is responsible for McrA
activity.  This is in agreement with the molecular weight deduced from sequence data.  McrA
protein was found to be localized in the outer membrane of Escherichia coli.  To our knowledge
this is the first restriction enzyme localized in the outer membrane of Escherichia coli.

<>

<1>Ramalingam, S., Kandavelou, K., Rajenderan, R., Chandrasegaran, S.
<2>Creating Designed Zinc-Finger Nucleases with Minimal Cytotoxicity.
<3>J. Mol. Biol.
<4>405
<5>630-641
<6>2011
<7>Zinc-finger nucleases (ZFNs) have emerged as powerful tools for delivering a targeted genomic
double-strand break (DSB) to either stimulate local
homologous recombination with investigator-provided donor DNA or induce
gene mutations at the site of cleavage in the absence of a donor by
nonhomologous end joining both in plant cells and in mammalian cells,
including human cells. ZFNs are formed by fusing zinc-finger proteins to
the nonspecific cleavage domain of the FokI restriction enzyme.
ZFN-mediated gene targeting yields high gene modification efficiencies
(>10%) in a variety of cells and cell types by delivering a recombinogenic
DSB to the targeted chromosomal locus, using two designed ZFNs. The
mechanism of DSB by ZFNs requires (1) two ZFN monomers to bind to their
adjacent cognate sites on DNA and (2) the FokI nuclease domains to
dimerize to form the active catalytic center for the induction of the DSB.
In the case of ZFNs fused to wild-type FokI cleavage domains, homodimers
may also form; this could limit the efficacy and safety of ZFNs by
inducing off-target cleavage. In this article, we report further
refinements to obligate heterodimer variants of the FokI cleavage domain
for the creation of custom ZFNs with minimal cellular toxicity. The
efficacy and efficiency of the reengineered obligate heterodimer variants
of the FokI cleavage domain were tested using the green fluorescent
protein gene targeting reporter system. The three-finger and four-finger
zinc-finger protein fusions to the REL_DKK pair among the newly generated
FokI nuclease domain variants appear to eliminate or greatly reduce the
toxicity of designer ZFNs to human cells.

<>

<1>Ramanathan, S.P., van Aelst, K., Sears, A., Peakman, L.J., Diffin, F.M., Szczelkun, M.D., Seidel, R.
<2>Type III restriction enzymes communicate in 1D without looping between their target sites.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>1748-1753
<6>2009
<7>To cleave DNA, Type III restriction enzymes must communicate the relative orientation of two
asymmetric recognition sites over hundreds of base
pairs. The basis of this long-distance communication, for which ATP
hydrolysis by their helicase domains is required, is poorly understood.
Several conflicting DNA-looping mechanisms have been proposed, driven
either by active DNA translocation or passive 3D diffusion. Using
single-molecule DNA stretching in combination with bulk-solution assays,
we provide evidence that looping is both highly unlikely and unnecessary,
and that communication is strictly confined to a 1D route. Integrating our
results with previous data, a simple communication scheme is concluded
based on 1D diffusion along DNA.

<>

<1>Ramanujam, R., Heaster, J., Huang, C., Jolly, J., Koelbl, J., Lively, C., Ogutu, E., Ting, E., Treml, S., Aldous, B., Hatley, R., Mathias, S., Franks, F., Burdick, B.
<2>Ambient temperature-stable molecular biology reagents.
<3>Biotechniques
<4>14
<5>470-474
<6>1993
<7>We have processed biological materials to generate several reagents that are ambient
temperature stable and ready to use. Stabilized biomolecules in a glassy matrix of
carbohydrate polymers offer water-soluble reagents for complex molecular biology applications.
This approach is particulary useful for reagent systems composed of enzymes, nucleotides and
other components dispensed in single-use aliquots. Reconstitution of the glassy matrix
delivers buffered enzymes and/or nucleotides for restriction, modification, sequencing and/or
amplification of nucleic acids. These ambient-temperature-stable reagents allow a high level
of reproducibility as they minimize the potential for pipetting errors. They also provide
advantages in shipping, storage and subsequent handling. Added convenience includes
elimination of setup time, cross contamination and refrigeration. Applications of
ambient-temperature-stable biological reagents for routine molecular biology methods are
presented.

<>

<1>Ramaraj, T., Matyi, S.A., Sundararajan, A., Lindquist, I.E., Devitt, N.P., Schilkey, F.D., Lamichhane-Khadka, R., Hoyt, P.R., Mudge, J., Gustafson, J.E.
<2>Draft Genome Sequences of Vancomycin-Susceptible Staphylococcus aureus Related to Heterogeneous Vancomycin-Intermediate S. aureus.
<3>Genome Announcements
<4>2
<5>e01033-14
<6>2014
<7>We report the draft genome sequences of three vancomycin-susceptible methicillin-resistant
Staphylococcus aureus strains. S. aureus strain MV8 is a
sequence type 8 (ST-8) staphylococcal cassette chromosome mec element type IV
(SCCmec IV) derivative, while the other two strains (S. aureus MM25 and MM61) are
ST-5 SCCmec II strains. MM61 is also closely related to the heterogeneous
vancomycin-intermediate S. aureus strain MM66.

<>

<1>Ramaraj, T., Sundararajan, A., Schilkey, F.D., Delvecchio, V.G., Donlon, M., Ziemer, C., Mudge, J.
<2>Improved Hybrid Genome Assemblies of Two Strains of Bacteroides xylanisolvens, SD_CC_1b and SD_CC_2a, Obtained Using Illumina and 454 Sequencing Technologies.
<3>Genome Announcements
<4>2
<5>e00237-14
<6>2014
<7>Bacteroides xlyanisolvens strains (SD_CC_1b, SD_CC_2a) isolated from human feces  were grown
on crystalline cellulose. Cellulolytic properties are not common in Bacteroides species. Here,
we report improved genome sequences of both of the B. xlyanisolvens strains.

<>

<1>Ramasamy, D., Dubourg, G., Robert, C., Caputo, A., Papazian, L., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Enorma timonensis sp.  nov.
<3>Standards in Genomic Sciences
<4>9
<5>970-986
<6>2014
<7>Enorma timonensis strain GD5(T) sp. nov., is the type strain of E. timonensis sp. nov., a new
member of the genus Enorma within the family Coriobacteriaceae. This
strain, whose genome is described here, was isolated from the fecal flora of a
53-year-old woman hospitalized for 3 months in an intensive care unit. E.
timonensis is an obligate anaerobic rod. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
2,365,123 bp long genome (1 chromosome but no plasmid) contains 2,060
protein-coding and 52 RNA genes, including 4 rRNA genes.

<>

<1>Ramasamy, D., Kokcha, S., Lagier, J.C., Nguyen, T.T., Raoult, D., Fournier, P.E.
<2>Genome sequence and description of Aeromicrobium massiliense sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>246-257
<6>2012
<7>Aeromicrobium massiliense strain JC14(T)sp. nov. is the type strain of Aeromicrobium
massiliense sp. nov., a new species within the genus Aeromicrobium.
This strain, whose genome is described here, was isolated from the fecal
microbiota of an asymptomatic patient. Aeromicrobium massiliense is an aerobic
rod-shaped gram-positive bacterium. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
3,322,119 bp long genome contains 3,296 protein-coding and 51 RNA genes.

<>

<1>Ramasamy, D., Lagier, J.C., Gorlas, A., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Bacillus massiliosenegalensis sp. nov.
<3>Standards in Genomic Sciences
<4>8
<5>264-278
<6>2013
<7>Bacillus massiliosenegalensis strain JC6(T) sp. nov. is the type strain of Bacillus
massiliosenegalensis sp. nov., a new species within the genus Bacillus.
This strain was isolated from the fecal flora of a healthy Senegalese patient. B.
massiliosenegalensis is an aerobic Gram-positive rod-shaped bacterium. Here we
describe the features of this organism, together with the complete genome
sequence and annotation. The 4,981,278-bp long genome comprises a 4,957,301-bp
chromosome and a 23,977-bp plasmid. The chromosome contains 4,925 protein-coding
and 72 RNA genes, including 4 rRNA genes. The plasmid contains 29 protein-coding
genes.

<>

<1>Ramasamy, D., Lagier, J.C., Nguyen, T.T., Raoult, D., Fournier, P.E.
<2>Non contiguous-finished genome sequence and description of Dielma fastidiosa gen. nov., sp. nov., a new member of the Family Erysipelotrichaceae.
<3>Standards in Genomic Sciences
<4>8
<5>336-351
<6>2013
<7>Dielma fastidiosa strain JC13(T) gen. nov., sp. nov. is the type strain of D. fastidiosa gen.
nov., sp. nov., the type species of a new genus within the family
Erysipelotrichaceae. This strain, whose draft genome is described here, was
isolated from the fecal flora of a healthy 16-year-old male Senegalese volunteer.
D. fastidiosa is a Gram-negative anaerobic rod. Here we describe the features of
this organism, together with the complete genome sequence and annotation. The
3,574,031 bp long genome comprises a 3,556,241-bp chromosome and a 17,790-bp
plasmid. The chromosome contains 3,441 protein-coding and 50 RNA genes, including
3 rRNA genes, whereas the plasmid contains 17 protein-coding genes.

<>

<1>Ramasamy, D., Lagier, J.C., Rossi-Tamisier, M., Pfleiderer, A., Michelle, C., Couderc, C., Raoult, D., Fournier, P.E.
<2>Genome sequence and description of Bacteroides timonensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1181-1197
<6>2014
<7>Bacteroides timonensis strain AP1(T) (= CSUR P194 = DSM 26083) is the type strain of B.
timonensis sp. nov. This strain, whose genome is described here, was
isolated from the fecal flora of a 21-year-old French Caucasoid female who
suffered from severe anorexia nervosa. Bacteroides timonensis is a Gram-negative,
obligate anaerobic bacillus. Here we describe the features of this organism,
together with the complete genome sequence and annotation. The 7,130,768 bp long
genome (1 chromosome, no plasmid) exhibits a G+C content of 43.3% and contains
5,786 protein-coding and 59 RNA genes, including 2 rRNA genes.

<>

<1>Ramasubban, G., Lakshmipathy, D., Vetrivel, U., Kulandai, L.T., Madhavan, H.N., Sridhar, R., Meenakshi, N.
<2>Whole-Genome Sequencing of Streptomycin-Resistant Mycobacterium tuberculosis Isolate VRFCWCF MRTB 180 Reveals Novel and Potential Mutations for Resistance.
<3>Genome Announcements
<4>2
<5>e00919-14
<6>2014
<7>We announce the draft genome sequence of a streptomycin monoresistant Mycobacterium
tuberculosis strain (VRFCWCF MRTB 180) isolated from sputum of a
clinically suspected tuberculosis patient.

<>

<1>Rambach, A.
<2>Purification and properties of the SstI endonuclease.
<3>Methods Enzymol.
<4>65
<5>170-173
<6>1980
<7>The SstI endonuclease is a restriction enzyme purified from a Streptomyces species which has
been named Streptomyces stanford and is available from the American Type Culture Collection as
ATCC No. 29415.  A crude lysate of these cells appears to cleave phage lambda DNA into two
large fragments and phage lambda plac5 DNA into three large fragments.  SstI endonuclease is
the major endonuclease which can be found in the lysate.  The enzyme produces cohesive ends
and seems to cut most DNAs tested at rare sites.

<>

<1>Ramchandani, S., Bhattacharya, S.K., Cervoni, N., Szyf, M.
<2>DNA methylation is a reversible biological signal.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>6107-6112
<6>1999
<7>The pattern of DNA methylation plays an important role in regulating different genome
functions.  To test the hypothesis that DNA methylation is a reversible biochemical process,
we purified a DNA demethylase from human cells that catalyzes the cleavage of a methyl residue
from 5-methyl cytosine and its release as methanol.  We show that similar to DNA
methyltransferase, DNA demethylase shows CpG dinucleotide specificity, can demethylate mdCpdG
sites in different sequence contexts, and demethylates both fully methylated and
hemimethylated DNA.  Thus, contrary to the commonly accepted model, DNA methylation is a
reversible signal, similar to other physiological biochemical modifications.

<>

<1>Ramchandani, S., Bigey, P., Szyf, M.
<2>Genomic structure of the human DNA methyltransferase gene.
<3>Biol. Chem.
<4>379
<5>535-540
<6>1998
<7>We determined the genomic  structure of the gene encoding human DNA methyltransferase.  Six
overlapping human genomic DNA clones which include all of the known cDNA sequence were
isolated.  Analysis of these clones demonstrates that the human DNA MTase gene consists of at
least 40 exons and 39 introns spanning a distance of 60 kilobases.  Elucidation of the
chromosomal organization of the human DNA MTase gene provides the template for future
structure-function analysis of the properties of mammalian DNA MTase.

<>

<1>Ramchandani, S., MacLeod, A.R., Pinard, M., von Hofe, E., Szyf, M.
<2>Inhibition of tumorigenesis by a cytosine-DNA, methyltransferase, antisense oligodeoxynucleotide.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>684-689
<6>1997
<7>This paper tests the hypothesis that cytosine DNA methyltransferase (DNA MeTase) is a
candidate target for anticancer therapy.  Several observations have suggested recently that
hyperactivation of DNA MeTase plays a critical role in initiation and progression of cancer
and that its up-regulation is a component of the Ras oncogenic signaling pathway.  We show
that a phosphorothioate-modified, antisense oligodeoxynucleotide directed against the DNA
MeTase mRNA reduces the level of DNA MeTase mRNA, inhibits DNA MeTase activity, and inhibits
anchorage independent growth of Y1 adrenocortical carcinoma cells ex vivo in a dose-dependent
manner.  Injection of DNA MeTase antisense oligodeoxynucleotides i.p. inhibits the growth of
Y1 tumors in syngeneic LAF1 mice, reduces the level of DNA MeTase, and induces demethylation
of the adrenocortical-specific gene C21 and its expression in tumors in vivo.  These results
support the hypothesis that an increase in DNA MeTase activity is critical for tumorigenesis
and is reversible by pharmacological inhibition of DNA MeTase.

<>

<1>Ramchandani, S., Szyf, M.
<2>A novel RNA element mediates the cell cycle dependent posttranscriptional regulation of DNA methyltransferase.
<3>Mol. Biol. Cell
<4>7
<5>302a
<6>1996
<7>DNA methyltransferase is the enzyme responsible for methylation of DNA at CpG dinucleotide
sequences.  Patterns of methylation have been shown to be an important control over gene
regulation.  Recent evidence has linked the disregulation of the DNA MeTase to oncogenesis.
The only mechanism of regulation of DNA MeTase gene expression thoroughly studied has been
transcriptional.  However, in untransformed cells the regulation of DNA MeTase has been
described to be posttranscriptional and dependent on the cell cycle.  The stability of DNA
MeTase mRNA is dependent on protein synthesis as the stability of the message is enhanced at
G0 upon the addition of cycloheximide.  Differential mRNA stabilities can be conferred by the
make-up of a transcript's 3' untranslated region (3'UTR).  Homology comparison between the
3' UTR's of the human, mouse, and chicken DNA MeTases have revealed two stretches of greater
than 90% homology between all three mRNAs and the AU content of these stretches is greater
than 85%.  To test the hypothesis that this sequence element regulates DNA MeTase mRNA it has
been inserted 3' to the rabbit-beta-globin gene and transfected into BALB/c 3T3 fibroblast
cells.  The chimeric globin-MeTase 3'UTR stability profile and its regulation with the cell
cycle recapitulates that of the endogenous DNA MeTase mRNA.  Our data suggests that a sequence
element in the DNA MeTase 3' UTR is a novel cell-cycle dependent regulator of mRNA stability.

<>

<1>Ramchandani, S.K., Rouleau, J., Szyf, M.
<2>Cloning of the human DNA methyltransferase gene.
<3>Am. J. Hum. Genet.
<4>55
<5>A137
<6>1994
<7>During the process of carcinogenesis it has been observed that DNA methylation is deregulated.
Increasing levels of DNA methyltransferase (DNA MeTase) mRNA has been shown to parallel the
progression of cells through neoplasia to tumour cells. At least two levels of regulation of
the mouse DNA MeTase have been shown; at the transcriptional level, via its promoter, and at
the post transcriptional level in a cell cycle dependent fashion. Previously in our lab, the
mouse 5' region was identified and was shown to be regulated by known oncogenic pathways. The
sequence of the complete DNA MeTase gene has not yet been reported. Identifying the promoter
of the human gene along with its complete genomic sequence is essential for a thorough study
of the mechanisms involved in the multi-tiered regulation of DNA MeTase and to understand
it's deregulation in cancer. Using a probe generated by PCR of the human DNA MeTase cDNA
(position +156 to +507), a human genomic library was screened and a clone of approximately 22
kilobases (kb) was isolated. It was found that this clone contains the complete coding
sequence of the DNA MeTase enzyme. Sequence analysis along with restriction enzyme digests
have allowed us to construct a partial map of the physical structure of the human DNA MeTase
gene. This partial structure has already revealed some interesting aspects related to the
genetic evolution of the human DNA MeTase. First, the proposed catalytic domain of the human
DNA MeTase is extremely homologous to all other cytosine DNA MeTases, even to those that are
found in bacteria, and this catalytic domain is conserved within one complete exon in the
human gene. This is very different from the structure of the 5' region of the gene, which is
fragmented into numerous little introns and exons. Within one of the small introns that have
been identified, a trinucleotide repeat of ATG occurs (9 times in a row), and this repeat is
upstream of the proposed start site of translation. Trinucleotide repeat expansion has been
shown to be a genetic hot spot of mutation, but even more interesting is the nature of the
repeat, ATG, which is the translation start codon and this repeat appears to be in frame with
the "normal" coding sequence. The implications being that possible alternative
methyltransferases may be translated under certain conditions such as cancer.

<>

<1>Ramesh, R., Gaitonde, S., Achari, G., Asolkar, T., Singh, N.P., Carrere, S., Genin, S., Peeters, N.
<2>Genome Sequencing of Ralstonia solanacearum Biovar 3, Phylotype I, Strains Rs-09-161 and Rs-10-244, Isolated from Eggplant and Chili in India.
<3>Genome Announcements
<4>2
<5>e00323-14
<6>2014
<7>Ralstonia solanacearum Indian strains Rs-09-161 and Rs-10-244 were isolated from  the coastal
region of Goa and from the Andaman Islands. We report the draft
genome sequences of these representative isolates infecting solanaceous
vegetables in India.

<>

<1>Ramirez, M.S., Adams, M.D., Bonomo, R.A., Centron, D., Tolmasky, M.E.
<2>Genomic Analysis of Acinetobacter baumannii A118 by Comparison of Optical Maps: Identification of Structures Related to its Susceptibility Phenotype.
<3>Antimicrob. Agents Chemother.
<4>55
<5>1520-1526
<6>2011
<7>Acinetobacter baumannii A118, a naturally competent clinical isolate, is unusually susceptible
to several antibiotics. Comparison of the optical map of strain A118 with in silico-generated
restriction maps of sequenced genomes and sequence analyses showed that the AbaR region,
commonly found inserted within the comM gene in other isolates, is missing in strain A118,
which could in part explain the susceptible phenotype exhibited by this isolate. These
comparative studies also showed differences in regions where genes coding for functions that
may be involved in drug resistance or susceptibility are located. Further sequencing
demonstrated that cat and blaADC, named blaADC-55, are present but that a tet(A) gene usually
found in other strains is not. In addition, carO and pbp2, which may play a role in
susceptibility to carbapenems, are present in strain A118. These findings support the idea
that A. baumannii strains possess multiple mechanisms that contribute to antibiotic
resistance, and the presence of some of them is not sufficient for a resistant phenotype. The
results shown here indicate that optical mapping is a useful tool for preliminary comparative
genomic analysis.

<>

<1>Ramirez, M.S., Xie, G., Johnson, S., Davenport, K., van Duin, D., Perez, F., Bonomo, R.A., Chain, P., Tolmasky, M.E.
<2>Genome Sequences of Two Carbapenemase-Resistant Klebsiella pneumoniae ST258 Isolates.
<3>Genome Announcements
<4>2
<5>e00558-14
<6>2014
<7>Klebsiella pneumoniae, an ESKAPE group (Enterococcus faecium, Staphylococcus aureus,
Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa,
and Enterobacter species) pathogen, has acquired multiple antibiotic resistance
genes and is becoming a serious public health threat. Here, we report the genome
sequences of two representative strains of K. pneumoniae from the emerging K.
pneumoniae carbapenemase (KPC) outbreak in northeast Ohio belonging to sequence
type 258 (ST258) (isolates Kb140 and Kb677, which were isolated from blood and
urine, respectively). Both isolates harbor a blaKPC gene, and strain Kb140
carries blaKPC-2, while Kb677 carries blaKPC-3.

<>

<1>Ramirez-Diaz, M.I., Diaz-Magana, A., Meza-Carmen, V., Johnstone, L., Cervantes, C., Rensing, C.
<2>Nucleotide sequence of Pseudomonas aeruginosa conjugative plasmid pUM505 containing virulence and heavy-metal resistance genes.
<3>Plasmid
<4>66
<5>7-18
<6>2011
<7>We determined the complete nucleotide sequence of conjugative plasmid
pUM505 isolated from a clinical strain of Pseudomonas aeruginosa. The
plasmid had a length of 123,322bp and contained 138 complete coding
regions, including 46% open reading frames encoding hypothetical proteins.
pUM505 can be considered a hybrid plasmid because it presents two
well-defined regions. The first region corresponded to a larger DNA
segment with homology to a pathogenicity island from virulent Pseudomonas
strains; this island in pUM505 was comprised of genes probably involved in
virulence and genes encoding proteins implicated in replication,
maintenance and plasmid transfer. Sequence analysis identified pil genes
encoding a type IV secretion system, establishing pUM505 as a member of
the family of IncI1 plasmids. Plasmid pUM505 also contained virB4/virD4
homologues, which are linked to virulence in other plasmids. The second
region, smaller in length, contains inorganic mercury and chromate
resistance gene clusters both flanked by putative mobile elements.
Although no genes for antibiotic resistance were identified, when pUM505
was transferred to a recipient strain of P. aeruginosa it conferred
resistance to the fluoroquinolone ciprofloxacin. pUM505 also conferred
resistance to the superoxide radical generator paraquat. pUM505 could
provide Pseudomonas strains with a wide variety of adaptive traits such as
virulence, heavy-metal and antibiotic resistance and oxidative stress
tolerance which can be selective factors for the distribution and
prevalence of this plasmid in diverse environments, including hospitals
and heavy metal contaminated soils.

<>

<1>Ramos, R.T. et al.
<2>Genome Sequence of the Corynebacterium pseudotuberculosis Cp316 Strain, Isolated  from the Abscess of a Californian Horse.
<3>J. Bacteriol.
<4>194
<5>6620-6621
<6>2012
<7>The bacterium Corynebacterium pseudotuberculosis is of major veterinary importance because it
affects livestock, particularly sheep, goats, and horses,
in several countries, including Australia, Brazil, the United States, and Canada,
resulting in significant economic losses. In the present study, we describe the
complete genome of the Corynebacterium pseudotuberculosis Cp316 strain, biovar
equi, isolated from the abscess of a North American horse.

<>

<1>Ramsahoye, B.H., Biniszkiewicz, D., Lyko, F., Clark, V., Bird, A.P., Jaenisch, R.
<2>Non-CpG methylation is prevalent in embryonic stem cells and may be mediated by DNA methyltransferase 3a.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>5237-5242
<6>2000
<7>Current evidence indicates that methylation of cytosine in mammalian DNA is restricted to both
strands of the symmetrical sequence CpG, although there have been sporadic reports that
sequences other than CpG may also be methylated. We have used a dual-labeling nearest neighbor
technique and bisulphite genomic sequencing methods to investigate the nearest neighbors of
5-methylcytosine residues in mammalian DNA. We find that embryonic stem cells, but not somatic
tissues, have significant cytosine-5 methylation at CpA and, to a lesser extent, at CpT. As
the expression of the de novo methyltransferase Dnmt3a correlates well with the presence of
non-CpG methylation, we asked whether Dnmt3a might be responsible for this modification.
Analysis of genomic methylation in transgenic Drosophila expressing Dnmt3a reveals that Dnmt3a
is predominantly a CpG methylase but also is able to induce methylation at CpA and at CpT.

<>

<1>Ramsahoye, B.H., Burnett, A.K., Taylor, C.
<2>Restriction endonuclease isoschizomers ItaI, BsoFI and Fsp4HI are characterized by differences in their sensitivities to CpG methylation.
<3>Nucleic Acids Res.
<4>25
<5>3196-3198
<6>1997
<7>BsoFI, ItaI and Fsp4HI are isoschizomers of Fnu4HI (5'-GC/NGC-3').  Both Fnu4HI and BsoFI
have previously been shown to be inhibited by cytosine-specific methylation within the
recognition sequence.  Fnu4HI is inhibited if either the internal cytosine at position 2 or
the external cytosine at position 5 of the restriction sequence is methylated, but the precise
nature of the methylation sensitivity of BsoFI is unclear from the literature.  The
methylation sensitivities of ItaI and Fsp4HI have not previously been reported.  By
methylating the plasmid pUC18 with M.SssI (a DNA cytosine-5'-methyltransferase with a
specificity for CpG), we have determined that ItaI is sensitive only to methylation of
internal CpG sites within the restriction sequence.  The methylation sensitivity of Fsp4HI is
identical to that of Fnu4HI, being inhibited by methylation of either internal CpG sites or
overlapping CpG sites.  BsoFI, like the other isoschizomers tested, is sensitive to a
combination of internal and overlapping CpG methylation.  BsoFI is also sensitive to
overlapping CpG methylation (in the absence of internal CpG methylation) if CpG overlap with
both sides of the recognition sequence.  Sites containing one overlapping CpG (in the absence
of internal CpG) are cut when methylated but show marked individual variation in their rates
of cleavage.  Considerable variation in the rate of cleavage by BsoFI is also observed at
sites containing only internal methylated CpG.  Some sites are cut slowly, whilst others fail
to cut even after prolonged incubation with excess of enzyme.

<>

<1>Ramsden, J.J., Dreier, J.
<2>Kinetics of the interaction between DNA and the type IC restriction enzyme EcoR124II.
<3>Biochemistry
<4>35
<5>3746-3753
<6>1996
<7>Optical waveguide mode spectroscopy was used to determine the binding constants characterizing
the interaction of EcoR124II, a type IC restriction modification enzyme from Salmonella
typhimurium, with DNA.  The DNA is immobilized on the surface of an optical waveguide, and the
enzyme is introduced in bulk solution flowing over the DNA under controlled hydrodynamic
conditions.  The binding kinetics of the protein to the DNA can be directly observed and the
number of bound protein molecules per base pair determined to a high accuracy.  Dissociation
of the protein was measured by switching flowing protein to protein-free buffer.  Binding to
two different kinds of DNA, with and without the specific sequence recognized by EcoR124II,
was investigated.  Protein binding and dissociation (M-RnonspecificM-S binding), quantified by
association and dissociation rate coefficients ka and kd, were the same for both types, but
the DNA carrying the recognition site showed an additional process, M-RirreversibleM-S
association (i.e. dissociation was not observed on the time scale of the experiments) of the
protein, quantified by a rate coefficient ks.  Some inferences regarding the mechanism of base
pair searching are made from the measured ka, kd and ks values.

<>

<1>Rand, K.N., Young, G.P., Ho, T., Molloy, P.L.
<2>Sensitive and selective amplification of methylated DNA sequences using helper-dependent chain reaction in combination with a methylation-dependent restriction enzymes.
<3>Nucleic Acids Res.
<4>41
<5>e15
<6>2013
<7>We have developed a novel technique for specific amplification of rare methylated DNA
fragments in a high background of unmethylated sequences
that avoids the need of bisulphite conversion. The
methylation-dependent restriction enzyme GlaI is used to selectively
cut methylated DNA. Then targeted fragments are tagged using specially
designed 'helper' oligonucleotides that are also used to maintain
selection in subsequent amplification cycles in a process called
'helper-dependent chain reaction'. The process uses disabled primers
called 'drivers' that can only prime on each cycle if the helpers
recognize specific sequences within the target amplicon. In this way,
selection for the sequence of interest is maintained throughout the
amplification, preventing amplification of unwanted sequences. Here we
show how the method can be applied to methylated Septin 9, a promising
biomarker for early diagnosis of colorectal cancer. The GlaI digestion
and subsequent amplification can all be done in a single tube. A
detection sensitivity of 0.1% methylated DNA in a background of
unmethylated DNA was achieved, which was similar to the
well-established Heavy Methyl method that requires bisulphite-treated
DNA.

<>

<1>Rao, B.S., Buckler-White, A.
<2>Direct visualization of site-specific and strand-specific DNA methylation patterns in automated DNA sequencing data.
<3>Nucleic Acids Res.
<4>26
<5>2505-2507
<6>1998
<7>We report here a simple method of directly visualizing in automated DNA sequencing
chromatograms DNA methylations of different types including cytosine
methylations in Hpa II and dcm sites as well as adenine methylations in dam
sites. This is made possible by the observation that the extent of incorporation
of fluorescently labeled dideoxynucleotides is influenced by the methylated bases
in template DNA. This simple approach involves routine automated DNA sequencing
without any prior treatment of DNA specific for detecting DNA methylation.

<>

<1>Rao, D.N., Dryden, D.T., Bheemanaik, S.
<2>Type III restriction-modification enzymes: a historical perspective.
<3>Nucleic Acids Res.
<4>42
<5>45-55
<6>2014
<7>Restriction endonucleases interact with DNA at specific sites leading to cleavage of DNA.
Bacterial DNA is protected from restriction endonuclease cleavage by modifying the DNA using a
DNA methyltransferase. Based on their molecular structure, sequence recognition, cleavage
position and cofactor requirements, restriction-modification (R-M) systems are classified into
four groups. Type III R-M enzymes need to interact with two separate unmethylated DNA
sequences in inversely repeated head-to-head orientations for efficient cleavage to occur at a
defined location (25-27 bp downstream of one of the recognition sites). Like the Type I R-M
enzymes, Type III R-M enzymes possess a sequence-specific ATPase activity for DNA cleavage.
ATP hydrolysis is required for the long-distance communication between the sites before
cleavage. Different models, based on 1D diffusion and/or 3D-DNA looping, exist to explain how
the long-distance interaction between the two recognition sites takes place. Type III R-M
systems are found in most sequenced bacteria. Genome sequencing of many pathogenic bacteria
also shows the presence of a number of phase-variable Type III R-M systems, which play a role
in virulence. A growing number of these enzymes are being subjected to biochemical and genetic
studies, which, when combined with ongoing structural analyses, promise to provide details for
mechanisms of DNA recognition and catalysis.

<>

<1>Rao, D.N., Eberle, H., Bickle, T.A.
<2>Characterization of mutations of the bacteriophage P1 mod gene encoding the recognition subunit of the EcoPI restriction and modification system.
<3>J. Bacteriol.
<4>171
<5>2347-2352
<6>1989
<7>This study characterized several mutations of the bacteriophage P1 mod gene.
This gene codes for the subunit of the EcoPI restriction enzyme that is
responsible for DNA sequence recognition and for modification methylation.  We
cloned the mutant mod genes into expression vectors and purified the mutant
proteins to near homogeneity.  Two of the mutant mod genes studied were the c2
clear-plaque mutants described by Scott (Virology 41:66-71, 1970).  These
mutant proteins can recognize EcoPI sites in DNA and direct restriction but are
unable to modify DNA.  Methylation assays as well as S-adenosylmethionine (SAM)
binding studies showed that the c2 mutants are methylation deficient because
they do not bind SAM, and we conclude that the mutations destroy the
SAM-binding site.  Both of the c2 mutations lie within a region of the EcoPI
mod gene that is not conserved when compared with the mod gene of the related
EcoPI5 system.  EcoPI5 and EcoPI recognize different DNA sequences, and we
believe that this region of the protein may code for the DNA-binding site of
the enzyme.  The other mutants characterized were made by site-directed
mutagenesis at codon 240.  Evidence is presented that one of them, Ser-240
->Pro, simultaneously lost the capacity to bind SAM and may also have changed
its DNA sequence specificity.

<>

<1>Rao, D.N., Page, M.G.P., Bickle, T.A.
<2>Cloning, over-expression and the catalytic properties of the EcoP15 modification methylase from Escherichia coli.
<3>J. Mol. Biol.
<4>209
<5>599-606
<6>1989
<7>The EcoP15 modification methylase gene from the p15B plasmid of Escherichia
coli 15T- has been cloned and expressed at high levels in a plasmid vector
system.  We have purified the enzyme to near homogeneity in large amounts and
have studied some of its enzymatic properties.  Initial rates of methyl
transfer are first order in methylase concentration and, with a pUC19 DNA as
substrate, the reaction proceeds by a random mechanism in which either DNA or
S-adenosylmethionine can bind to the free enzyme.  After methyltransfer to DNA,
the methylated DNA and S-adenosylhomocysteine appear to dissociate in random
order.  As expected in such a mechanism, S-adenosylhomocysteine is a
non-competitive inhibitor with respect to both S-adenosylmethionine and DNA.
DNA-dependent substrate inhibition by S-adenosylmethionine at concentrations
not much above its KM suggests that release of methylated DNA may be the
rate-limiting step.  This suggestion is strengthened by the fact that a mutant
of the closely related EcoP1 does not show such substrate inhibition.

<>

<1>Rao, D.N., Saha, S., Krishnamurthy, V.
<2>ATP-dependent restriction enzymes.
<3>Prog. Nucleic Acid Res. Mol. Biol.
<4>64
<5>1-63
<6>2000
<7>The phenomenon of restriction and modification (R-M) was first observed in the course of
studies on bacteriophages in the early 1950s. It was only in the 1960s that work of Arber and
colleagues provided a molecular explanation for the host specificity. DNA restriction and
modification enzymes are responsible for the host-specific barriers to interstrain and
interspecies transfer of genetic information that have been observed in a variety of bacterial
cell types. R-M systems comprise an endonuclease and a methyltransferase activity. They serve
to protect bacterial cells against bacteriophage infection, because incoming foreign DNA is
specifically cleaved by the restriction enzyme if it contains the recognition sequence of the
endonuclease. The DNA is protected from cleavage by a specific methylation within the
recognition sequence, which is introduced by the methyltransferase. Classic R-M systems are
now divided into three types on the basis of enzyme complexity, cofactor requirements, and
position of DNA cleavage, although new systems are being discovered that do not fit readily
into this classification. This review concentrates on multisubunit, multifunctional
ATP-dependent restriction enzymes. A growing number of these enzymes are being subjected to
biochemical and genetic studies that, when combined with ongoing structural analyses, promise
to provide detailed models for mechanisms of DNA recognition and catalysis. It is now clear
that DNA cleavage by these enzymes involves highly unusual modes of interaction between the
enzymes and their substrates. These unique features of mechanism pose exciting questions and
in addition have led to the suggestion that these enzymes may have biological functions beyond
that of restriction and modification. The purpose of this review is to describe the exciting
developments in our understanding of how the ATP-dependent restriction enzymes recognize
specific DNA sequences and cleave or modify DNA.

<>

<1>Rao, M., Suvarna, K., Srinivasan, M.C., Vasanti, D.
<2>Development of Chainia as a host for xlanase gene cloning: evidence for occurrence of a restriction-modification system.
<3>Biotechnol. Lett.
<4>18
<5>327-332
<6>1996
<7>Conditions for genetic transformation of the xylanase-negative (X-) strain of
Chainia with pIJ 702 were optimized.  The growth of Chainia at 30oC for 36-40h and addition of
gelatin (1%) to the medium resulted in the maximum yield of protoplasts and regeneration
efficiency.  Poor transformation efficiency of Chainia (X-) protoplasts by native pIJ 702
versus
improved efficiency (16 transformants/ug of plasmid DNA) by prior heating of protoplasts at
42oC
for 10 min suggests the occurrence of a restriction system in Chainia.  Increased
transformation
efficiency by passage of the plasmid through Chainia together with the altered methylation
status of
the transformant plasmid presents evidence for the existence of an operative modification
system in
Chainia.  Development of thiostrepton resistance and formation of melamin pigment in Chainia
(X-
) by transformation with pIJ 702 reveal that genes from Streptomyces can be functionally
expressed by Chainia (X-).

<>

<1>Rao, Q., Wang, S., Su, Y.L., Bing, X.L., Liu, S.S., Wang, X.W.
<2>Draft Genome Sequence of 'Candidatus Hamiltonella defensa,' an Endosymbiont of the Whitefly Bemisia tabaci.
<3>J. Bacteriol.
<4>194
<5>3558
<6>2012
<7>'Candidatus Hamiltonella defensa' is a facultative endosymbiont of the whitefly Bemisia
tabaci. Herein, we report the first draft genome sequence of 'Candidatus
Hamiltonella defensa' from the invasive Mediterranean cryptic species of the B.
tabaci complex. The 1.84-Mbp genome sequence comprises 404 contigs and contains
1,806 predicted protein-coding genes.

<>

<1>Rao, Q., Wang, S., Zhu, D.T., Wang, X.W., Liu, S.S.
<2>Draft Genome Sequence of Rickettsia sp. Strain MEAM1, Isolated from the Whitefly  Bemisia tabaci.
<3>J. Bacteriol.
<4>194
<5>4741-4742
<6>2012
<7>We report the draft genome sequence of the Rickettsia sp. strain MEAM1, which is  a
facultative symbiont from an invasive species of the whitefly Bemisia tabaci.
The total length of the assembled genome is approximately 1.24 Mb, with 335
scaffolds and 1,247 coding sequences predicted within the genome.

<>

<1>Rao, S.B., Gupta, V.K., Kumar, M., Hegde, N.R., Splitter, G.A., Reddanna, P., Radhakrishnan, G.K.
<2>Draft Genome Sequence of the Field Isolate Brucella melitensis Strain Bm IND1 from India.
<3>Genome Announcements
<4>2
<5>e00497-14
<6>2014
<7>Brucella spp. are facultative intracellular bacterial pathogens causing the zoonotic disease
brucellosis. Here, we report the draft genome sequence of the
Brucella melitensis strain from India designated Bm IND1, isolated from stomach
contents of an aborted goat fetus.

<>

<1>Rapa, R.A., Islam, A., Monahan, L.G., Mutreja, A., Thomson, N., Charles, I.G., Stokes, H.W., Labbate, M.
<2>A genomic island integrated into recA of Vibrio cholerae contains a divergent recA and provides multi-pathway protection from DNA damage.
<3>Environ. Microbiol.
<4>0
<5>0
<6>2014
<7>Lateral gene transfer (LGT) has been crucial in the evolution of the cholera
pathogen, Vibrio cholerae. The two major virulence factors are present on two
different mobile genetic elements, a bacteriophage containing the cholera toxin
genes and a genomic island (GI) containing the intestinal adhesin genes.
Non-toxigenic V. cholerae in the aquatic environment are a major source of novel
DNA that allows the pathogen to morph via LGT. In this study, we report a novel
GI from a non-toxigenic V. cholerae strain containing multiple genes involved in
DNA repair including the recombination repair gene recA that is 23% divergent
from the indigenous recA and genes involved in the translesion synthesis pathway.
This is the first report of a GI containing the critical gene recA and the first
report of a GI that targets insertion into a specific site within recA. We show
that possession of the island in Escherichia coli is protective against DNA
damage induced by UV-irradiation and DNA targeting antibiotics. This study
highlights the importance of genetic elements such as GIs in the evolution of V.
cholerae and emphasizes the importance of environmental strains as a source of
novel DNA that can influence the pathogenicity of toxigenic strains.

<>

<1>Rappert, S., Song, L., Sabra, W., Wang, W., Zeng, A.P.
<2>Draft Genome Sequence of Type Strain Clostridium pasteurianum DSM 525 (ATCC 6013), a Promising Producer of Chemicals and Fuels.
<3>Genome Announcements
<4>1
<5>e00232-12
<6>2013
<7>, an anaerobic bacterium able to utilize atmospheric free nitrogen for biosynthesis, has
recently been proven to be a promising producer of chemicals
and fuels, such as 1,3-propanediol and -butanol. Here, we report the high-quality
draft genome sequence of DSM 525, a type strain of .

<>

<1>Rappuoli, R., Fraser, C., Pizza, M., Scarselli, M., Serruto, D., Tetelin, H.
<2>Polypeptides from Neisseria meningitidis.
<3>International Patent Office
<4>WO 2008001224 A
<5>
<6>2008
<7>Various specific meningococcal proteins are disclosed.  The invention provides related
polypeptides, nucleic acids, antibodies and methods.  These can all be used in medicine for
treating or preventing disease and/or infection caused by meningococcus, such as bacterial
meningitis.

<>

<1>Raschke, E.
<2>Comprehensive restriction enzyme lists to update to any DNA sequence computer program.
<3>Genet. Anal. Tech. Appl.
<4>10
<5>49-60
<6>1993
<7>Restriction enzyme lists are presented for the practical working geneticist to update any DNA
computer program. These lists combine formerly scattered information and contain all presently
known restriction enzymes with a unique recognition sequence, a cut site, or methylation
(in)sensitivity. The lists are in the shortest possible form to also be functional with small
DNA computer programs, and will produce clear restriction maps without any redundancy or loss
of information. The lists discern between commercial and noncommercial enzymes, and prototype
enzymes and different isoschizomers are cross-referenced. Differences in general methylation
sensitivities and (in)sensitivities against Dam and Dcm methylases of Escherichia coli are
indicated. Commercial methylases and intron-encoded endonucleases are included. An address
list is presented to contact commercial suppliers. The lists are constantly updated and
available in electronic form as pure US ASCI filed, and in formats for the DNA computer
programs DNA-Strider for Apple Macintosh, and DNAsis for IBM personal computers or compatibles
via e-mail from the internet address: NETSERV@EMBLHEIDELBERG.DE by sending only the message
HELP RELIBRARY.

<>

<1>Rashtchian, A., Chatterjee, D., Guan, N., Schuster, D.
<2>Compositions and methods for positive selection cloning.
<3>International Patent Office
<4>WO 9838205
<5>
<6>1998
<7>Nucleic acid molecules are disclosed which comprise a toxic gene operably linked to a
regulatory DNA sequence, wherein the regulatory DNA sequence contains one or more cloning
sites allowing insertional inactivation of the toxic gene.  Also disclosed are kits for
cloning and selecting nucleic acid molecules in a transformed host cell using the methods and
compositions of the invention.  In the practice of the invention, when a genetic insert is
inserted into a cloning site within the nucleic acid molecule comprising the toxic gene, the
toxic gene is rendered substantially non-toxic to the host cell, allowing for the isolation of
viable transformants which comprise the gene to be cloned and selected.  The invention also
provides stable compositions for cloning and selecting a nucleic acid molecule, methods for
producing recombinant polypeptides using these nucleic acid molecules and compositions, and
recombinant polypeptides produced according to these methods.

<>

<1>Rasko, D.A. et al.
<2>Origins of the E. coli strain causing an outbreak of hemolytic-uremic syndrome in Germany.
<3>N. Engl. J. Med.
<4>365
<5>709-717
<6>2011
<7>BACKGROUND: A large outbreak of diarrhea and the hemolytic-uremic syndrome
caused by an unusual serotype of Shiga-toxin-producing Escherichia coli
(O104:H4) began in Germany in May 2011. As of July 22, a large number of
cases of diarrhea caused by Shiga-toxin-producing E. coli have been
reported--3167 without the hemolytic-uremic syndrome (16 deaths) and 908
with the hemolytic-uremic syndrome (34 deaths)--indicating that this
strain is notably more virulent than most of the Shiga-toxin-producing E.
coli strains. Preliminary genetic characterization of the outbreak strain
suggested that, unlike most of these strains, it should be classified
within the enteroaggregative pathotype of E. coli. METHODS: We used
third-generation, single-molecule, real-time DNA sequencing to determine
the complete genome sequence of the German outbreak strain, as well as the
genome sequences of seven diarrhea-associated enteroaggregative E. coli
serotype O104:H4 strains from Africa and four enteroaggregative E. coli
reference strains belonging to other serotypes. Genomewide comparisons
were performed with the use of these enteroaggregative E. coli genomes, as
well as those of 40 previously sequenced E. coli isolates. RESULTS: The
enteroaggregative E. coli O104:H4 strains are closely related and form a
distinct clade among E. coli and enteroaggregative E. coli strains.
However, the genome of the German outbreak strain can be distinguished
from those of other O104:H4 strains because it contains a prophage
encoding Shiga toxin 2 and a distinct set of additional virulence and
antibiotic-resistance factors. CONCLUSIONS: Our findings suggest that
horizontal genetic exchange allowed for the emergence of the highly
virulent Shiga-toxin-producing enteroaggregative E. coli O104:H4 strain
that caused the German outbreak. More broadly, these findings highlight
the way in which the plasticity of bacterial genomes facilitates the
emergence of new pathogens.

<>

<1>Rasko, D.A., Ravel, J., Okstad, O.A., Helgason, E., Cer, R.Z., Jiang, L., Shores, K.A., Fouts, D.E., Tourasse, N.J., Angiuoli, S.V., Kolonay, J., Nelson, W.C., Kolsto, A.-B., Fraser, C.M., Read, T.D.
<2>The genome sequence of Bacillus cereus ATCC 10987 reveals metabolic adaptations and a large plasmid related to Bacillus anthracis pXO1.
<3>Nucleic Acids Res.
<4>32
<5>977-988
<6>2004
<7>We sequenced the complete genome of Bacillus cereus ATCC 10987, a non-lethal dairy isolate in
the same genetic subgroup as Bacillus anthracis. Comparison of the chromosomes demonstrated
that B.cereus ATCC 10987 was more similar to B.anthracis Ames than B.cereus ATCC 14579, while
containing a number of unique metabolic capabilities such as urease and xylose utilization and
lacking the ability to utilize nitrate and nitrite. Additionally, genetic mechanisms for
variation of capsule carbohydrate and flagella surface structures were identified. Bacillus
cereus ATCC 10987 contains a single large plasmid (pBc10987, of  208 kb, that is similar in
gene content and organization to B.anthracis pXO1 but is lacking the pathogenicity-associated
island containing the anthrax lethal and edema toxin complex genes. The chromosomal similarity
of B.cereus ATCC 10987 to B.anthracis Ames, as well as the fact that it contains a large
pXO1-like plasmid, may make it a possible model for studying B.anthracis plasmid biology and
regulatory cross-talk.

<>

<1>Rasko, T., Der, A., Klement, E., Slaska-Kiss, K., Posfai, E., Medzihradszky, K.F., Marshak, D.R., Roberts, R.J., Kiss, A.
<2>BspRI restriction endonuclease: cloning, expression in Escherichia coli and sequential cleavage mechanism.
<3>Nucleic Acids Res.
<4>38
<5>7155-7166
<6>2010
<7>The GGCC-specific restriction endonuclease BspRI is one of the few Type IIP restriction
endonucleases, which were suggested to be a monomer. Amino
acid sequence information obtained by Edman sequencing and mass
spectrometry analysis was used to clone the gene encoding BspRI. The
bspRIR gene is located adjacently to the gene of the cognate modification
methyltransferase and encodes a 304 aa protein. Expression of the bspRIR
gene in Escherichia coli was dependent on the replacement of the native
TTG initiation codon with an ATG codon, explaining previous failures in
cloning the gene using functional selection. A plasmid containing a single
BspRI recognition site was used to analyze kinetically nicking and
second-strand cleavage under steady-state conditions. Cleavage of the
supercoiled plasmid went through a relaxed intermediate indicating
sequential hydrolysis of the two strands. Results of the kinetic analysis
of the first- and second-strand cleavage are consistent with cutting the
double-stranded substrate site in two independent binding events. A
database search identified eight putative restriction-modification systems
in which the predicted endonucleases as well as the methyltransferases
share high sequence similarity with the corresponding protein of the BspRI
system. BspRI and the related putative restriction endonucleases belong to
the PD-(D/E)XK nuclease superfamily.

<>

<1>Rasko, T., Finta, C., Kiss, A.
<2>DNA bending induced by DNA (cytosine-5) methyltransferases.
<3>Nucleic Acids Res.
<4>28
<5>3083-3091
<6>2000
<7>DNA bending induced by six DNA (cytosine-5) methyltransferases was studied using circular
permutation gel mobility shift assay. The following bend angles were obtained: M.BspRI
(GG(m5)CC), 46-50 degrees; M.HaeIII (GG(m5)CC), 40-43 degrees; M.SinI (GGW(m5)CC), 34-37
degrees; M.Sau96I (GGN(m5)CC), 52-57 degrees; M.HpaII (C(m5)CGG), 30 degrees; and M.HhaI
(G(m5)CGC), 13 degrees. M.HaeIII was also tested with fragments carrying a methylated binding
site, and it was found to induce a 32 degrees bend. A phase-sensitive gel mobility shift
assay, using a set of DNA fragments with a sequence-directed bend and a single
methyltransferase binding site, indicated that M.HaeIII and M.BspRI bend DNA toward the
minor groove. The DNA curvature induced by M.HaeIII contrasts with the lack of DNA bend
observed for a covalent M.HaeIII-DNA complex in an earlier X-ray study. Our results and data
from other laboratories show a correlation between the bending properties and the recognition
specificities of (cytosine-5) methyltransferases: enzymes recognizing a cytosine 3' to the
target cytosine tend to induce greater bends than enzymes with guanine in this position. We
suggest that the observed differences indicate different mechanisms employed by (cytosine-5)
methyltransferases to stabilize the helix after the target base has flipped out.

<>

<1>Rasmussen, L.J., Lobner-Olesen, A., Marinus, M.G.
<2>Growth-rate-dependent transcription initiation from the dam P2 promoter.
<3>Gene
<4>157
<5>213-215
<6>1995
<7>Transcription of the dam gene in Escherichia coli is dependent on growth rate.  Using
single-copy promoter::lacZYA fusions we found that of the five promoter regions which affect
dam expression, only the P2 promoter shows growth-rate dependence.  The determinants for
growth-rate control must lie in the region -52 to +27 relative to the transcription start
point.

<>

<1>Rasmussen, L.J., Marinus, M.G., Lcbner-Olesen, A.
<2>Regulation of expression of the dam methyltransferase gene (dam) from Escherichi coli.
<3>J. Cell Biochem. Suppl.
<4>17E
<5>303
<6>1993
<7>The level of Dam protein (DNA adenine methyltransferase) in E. coli is critical for the
efficient action of Dam-directed mismatch repair and synchronous initiation of chromosome
replication from oriC. A drastic increase or decrease from the normal level of 130 molecules
per cell causes hypermutability and asynchronous initiation. The Dam methyltransferase is
encoded by the dam gene located at 74 min on the genetic map. The dam gene is part of a
transcriptional unit which includes five promoters and at least four genes: aroK (shikimic
acids kinase I), aroB (3-dehydroquinate synthase), urf74.3 (an unidentified open reading
frame) and dam (Dam methyltransferase). As a first step to elucidate the mechanism of
regulation we have found that the expression of the dam gene is growth rate regulated and
coordinated with cell growth. In order to determine the region responsible for this regulation
we have separated and characterized the promoters individually.

<>

<1>Rasmussen, L.J., Marinus, M.G., Lobner-Olesen, A.
<2>Novel growth rate control of dam gene expression in Escherichia coli.
<3>Mol. Microbiol.
<4>12
<5>631-638
<6>1994
<7>Transcription of the dam gene in Escherichia coli is growth rate regulated by a mechanism
distinct from that used for ribosomal RNA gene promoters.
Single-copy operon fusions to lacZ indicated that the major promoter, P2,
is responsible for most or all of the growth rate dependence. Promoter P2
is a typical sigma 70 promoter with 18 bp spacing between the -10 and -35
hexamers. Primer extension analysis was used to show that there was no
inhibition of transcription from promoter P2 in cells induced for the
stringent response. Beta-galactosidase specific activity from a
single-copy dam::lacZ fusion was unaffected by either excess rrnB RNA or
the level of Fis protein. Thus growth rate control of dam gene expression
differs from that of the rRNA and tRNA genes by its lack of response to
stringent control, ribosomal feedback and enhanced transcription by Fis
protein. We devised a procedure for selection of mutant cells in which dam
gene expression was unregulated. One such mutant (cde-4), obtained by
miniTn10 insertion, showed the same level of beta-galactosidase activity
at all growth rates tested. In contrast, growth rate-dependent expression
of the rrnB gene was unaffected by cde-4 confirming the different modes of
regulation. The cde-4::miniTn10 insertion is located close to kilobase 670
on the physical map in or near the lipB gene.

<>

<1>Rasmussen, L.J., Samson, L., Marinus, M.G.
<2>Dam-directed DNA mismatch repair.
<3>DNA Damage and Repair, Humana Press, Inc., Nickloff, J.A. and Hoekstra, M.F., Totowa, NJ
<4>1
<5>205-228
<6>1998
<7>Base mismatches in duplex DNA are repaired by a variety of systems in Escherichia coli,
including: Very Short Patch repair; MutY-dependent repair; and DNA adenine
methyltransferase-directed DNA mismatch repair.  The first two systems have a restricted
substrate specificity (T.G and A.G mismatches, respectively), whereas the third system can
repair 11 out of the 12 possible base mispairs with only the C.C mismatch being refractory.
There is no known repair pathway for C.C mismatches in duplex DNA.  The DDMR pathway also acts
on insertions/deletions ("loops") of up to 4 bases.  In this chapter are reviewed the VSP
repair and DDMR systems with emphasis on the latter.  The MutY-pathway is discussed in Chapter
6.

<>

<1>Rastorguev, S.M., Zavilgelskii, G.B., Suzuki, K., Sakka, K.
<2>Alleviation of type I restriction in Escherichia coli K12 in the presence of the arsR gene from pKW301 of Acidiphilium multivorum AIU 301.
<3>Mol. Biol. (Mosk)
<4>35
<5>79-82
<6>2001
<7>A study was made of the antirestriction activity of Acidiphilium multivorum AIU 301 ArsR, a
repressor of the ars operon which confers resistance to arsenite and arsenate and is on
pKW301. In Escherichia coli, arsR cloned under the control of Plac in a multi-copy vector
alleviated restriction of nonmodified lambda DNA by a factor of 120, six times more
efficiently than its analogs of conjugal plasmids R64 (incI1) and R773 (incFI). Amino acid
sequence analysis showed that the three ArsR proteins have a homologous region of 38 residues,
including the antirestriction motif, in their N domains, whereas the motif is in the C domain
in the Ard proteins. The other regions are nonhomologous, and pKW301 ArsR is 33 residues
shorter than R64 and R773 ArsRs. The total charge is -4 in pKW301 ArsR and +2 in R64 and R733
ArsRs. A total negative charge was assumed to contribute to the antirestriction activity.

<>

<1>Rastorguev, S.M., Zavilgelsky, G.B.
<2>Role of "antirestriction" motif in functional activity of antirestriction protein ArdA pKM101 (IncN).
<3>Genetika
<4>39
<5>286-292
<6>2003
<7>A number of mutant forms of the antirestriction protein ArdA encoded by the ardA gene located
in a transmissive IncN plasmid pKM101 have been
constructed. Proteins belonging to the Ard family are specific inhibitors
of type I restriction--modification enzymes. Single mutational
substitutions of negatively charged amino acid residues located in the
"antirestriction motif" with hydrophobic alanine, E134A, E137A, D144A, or
a double substitution E134A, E137A do not affect the antirestriction
activity (Ard) of ArdA but almost completely abolish the antimodification
activity (Amd). Mutational substitutions F107D and A110D in the assumed
interface ArdA, which determines contact between monomers in the active
dimer (Ard)2, cause an approximately 100-fold decrease in the
antirestriction protein activity. It is hypothesized that the ArdA protein
forms two complexes with the type I restriction--modification enzyme
(R2M2S): (1) with a specific region in the S subunit involved in contact
with the sK site in DNA; and (2) with a nonspecific region in the R
subunit involved in DNA translocation and degradation by restriction
endonucleases. The association of ArdA with the specific region inhibits
restriction endonuclease and methyltransferase activities simultaneously,
whereas the association of ArdA with a nonspecific region inhibits only
restriction endonuclease activity of the R2M2S enzyme.

<>

<1>Rastorguev, S.M., Zavilgelsky, G.B., Tchurikov, N.A.
<2>IncI1 plasmid R64 encodes the ArsR protein that alleviates type I restriction.
<3>FEBS Lett.
<4>426
<5>21-23
<6>1998
<7>The host-controlled EcoK restriction of unmodified phage lambda was five-fold alleviated in
the wild-type Escherichia coli strain K12 carrying the R64 plasmid of the incompatibility
group I1. The relevant gene was mapped between the origin of vegetative replication (rep,
oriV) and the tet(r) gene about 60 kbp downstream from the origin of transfer, oriT. We cloned
this gene inside the 613 bp long EcoRI-PstI fragment and sequenced it. Only one 351 bp long
open reading frame (ORF) starting at 124 bp from the beginning of the insert was found in the
sequence. Computer search in the current databases revealed that the putative protein is
identical to the ArsR protein specified by the IncFI plasmid R773. ArsR is a repressor of the
arsenical resistance (ars) operon, arsRDABC. There are no arsABC genes in the R64 plasmid
since plasmid R64- (or pSR8)-mediated resistance of E. coli K12 cells to the arsenicals
arsenate and arsenite was not detected. The gene arsR and the antirestriction genes ard (ardA
and ardB) are non-homologous. However, comparison of the deduced amino acid sequence of ArsR
with the ArdA and ArdB sequences revealed only one small region of similarity, a 9 amino acid
motif found in different antirestriction proteins that is hypothesized to be an interaction
site for antirestriction proteins with restriction endonucleases.

<>

<1>Rathert, P., Rasko, T., Roth, M., Slaska-Kiss, K., Pingoud, A., Kiss, A., Jeltsch, A.
<2>Reversible inactivation of the CG specific Sssl DNA (cytosine-C5)-methyltransferase with a photocleavable protecting group.
<3>Chembiochem
<4>8
<5>202-207
<6>2007
<7>Caging of proteins by conjugation with a photocleavable group is a powerful approach for
reversibly blocking enzymatic activity. Here we
describe the covalent modification of the bacterial Sssl DNA
methyltransferase (M.Sssl) with the cysteine-specific reagent
4,5-dimethoxy-2-nitrobenzyibromide (DMNBB). M.Sssl contains two
cysteine residues; replacement of the active-site Cys141 with Ser
resulted in an approximately 700-fold loss of enzymatic activity; this
indicates an important role for this residue in catalysis. However,
replacement of Cys368 with Ala did not affect methyltransferase
activity. Treatment of the Cys368Ala mutant enzyme with DMNBB led to an
almost complete loss of activity. Irradiation of the inactivated enzyme
with near-ultraviolet light (320400 nm) restored 60% of the catalytic
activity. This indicates that caging by DMNBB can be used for the
reversible inactivation of M.Sssl.

<>

<1>Raveh, D.
<2>DNA segments encoding a domain of HO-endonuclease.
<3>International Patent Office
<4>WO 9836079
<5>
<6>1998
<7>The invention provides an isolated DNA segment encoding a domain of HO-endonuclease, said
segment being selected from the group consisting of an isolated DNA segment encoding the
N-terminus of HO-endonuclease which contains the sequence-specific catalytic nuclease activity
of said HO-endonuclease, said DNA fragment comprising at least 755 nt and encoding for at
least 251 amino acids, and said endonuclease being characterized by the presence of at least
one copy of the dodecapeptide motif LAGLI-DADG and an isolated DNA segment encoding the
C-terminus of HO-endonuclease which contains the DNA binding/recognition activity of said
HO-endonuclease, said C-terminus comprising 360 nt and encoding for 120 amino acids.

<>

<1>Raven, N.D.H., Kelly, C.D., Carter, N.D., Eastlake, P., Brown, C., Williams, R.A.D.
<2>A new restriction endonuclease, TspEI, from the genus Thermus that generates cohesive termini compatible with those of EcoRI.
<3>Gene
<4>131
<5>83-86
<6>1993
<7>The detection, isolation and properties of the restriction endonuclease TspEI are described.
The canonical recognition sequence (AATT) is the same as the 4-bp core of the 6-bp sequence
(GAATTC) of EcoRI. Hydrolysis occurs 5' to the palindromic tetramer so that TspEI produces
the same cohesive termini as EcoRI. TspEI therefore has an obvious application in producing
partial digests of DNA for ligation to EcoRI-digested cloning vectors.

<>

<1>Raven, N.D.H., Williams, R.A.D., Smith, K.E., Kelly, C.D., Carter, N.D.
<2>Tsp45I, a new thermostable site-specific endonuclease that cleaves the recognition sequence 5'-GTSAC-3'.
<3>Nucleic Acids Res.
<4>21
<5>4397
<6>1993
<7>A site-specific nuclease from Thermus strain 45 was purified by ammonium sulphate
fractionation and then by chromatography on Phospho-Ultrogel, DEAE-Fractogel 650 M, and
Sephadex G200 columns. The Mr by gel filtration was 80 kD, and a single band on SDS-PAGE was
estimated as 38 kD. On the basis of the fragment pattern of digests of pBR322 and phiX174 RF
DNA< and by reference to reported fragment sizes for these substrates, it was concluded that
the recognition site was GTSAC. This was confirmed by the identity of Tsp45I patterns with
those predicted by computer simulations of substrate hydrolysis at this site (kindly provided
by R.Roberts), and by the agreement between observed and expected fragment sized for digests
of lambda DNA, M13mp18RF DNA, pHC624 and pUC18. The optimum activity was observed at 65oC in
10 mM Tris chloride buffer, pH7.3 at 25oC containing no NaCl, l mM MgCl2, 2 mM dithiothreitol
and 100 mg/l albumin. The enzyme has a low requirement for inorganic ions, indeed activity was
detected even when magnesium ions were omitted from the assay buffer. The cleavage points were
determined by the primed synthesis method from the M13 -40 sequencing primer on a recognition
site provided by two complementary synthetic oligonucleotides 5'CCGTCGACGTGACGGATCCCC and 5'
GGGGATCCGTCACGTCGACGG that were annealed together and digested with SalI and BamHI. The
central fragment that contained the recognition site was ligated into M13mp8 digested with the
same enzymes and DNA was extracted and purified. The hydrolysis sites of Tsp45I (Figure 1) lie
outside the recognition site, 5' to the G residues in each strand: 5'^GTGAC-3', 3'-CACTG^5'.

<>

<1>Ravetch, J.V., Horiuchi, K., Zinder, N.D.
<2>Nucleotide sequence of the recognition site for the restriction-modification enzyme of Escherichia coli B.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>75
<5>2266-2270
<6>1978
<7>The nucleotide sequence of the recognition site for the restriction-modifiction
enzyme of Escherichia coli B (SB site) has been determined.  The recognition
site is a 15-nucleotide sequence consisting of the trimer 5'TGA3', followed by
an 8-nucleotide domain of variable sequence, which in turn is followed by the
tetramer 5'TGCT3'.  The sequence has no 2-fold rotational symmetry.  Single
base changes in the constant nucleotide domains result in the loss of
sensitivity to both restriction and modification.  Our data are also consistent
with modification occurring by methylation of two adenine residues per SB site:
one on the adenine of the trimer 5'TGA3' and the other on the complementary
strand on the adenine complementary to the first thymine of the tetramer
5'TGCT3'.  All nine independently isolated spontaneous mutants at the SB1 site
of bacteriophage f1 are caused by a C-to-T transversion.  Mutations at the SB2
site are caused by various single base changes.

<>

<1>Ravi, R.S., Sozhamannan, S., Dharmalingam, K.
<2>Transposon mutagenesis and genetic mapping of the rglA and rglB loci of Escherichia coli.
<3>Mol. Gen. Genet.
<4>198
<5>390-392
<6>1985
<7>The rglA and rglB genes code for two different proteins which cleave the
hydroxymethylated cytosine residues of T-even phages.  We isolated Tn10 and Tn5
insertion mutants of the above genes and of the genes in and around the rglA
and rglB loci.  These insertions were used to cocnstruct a detailed genetic
map.  Our results show that the rglA gene maps at 25.24 min and the rglB gene
at 98.39 min on the standard Escherichia coli K12 genetic map.

<>

<1>Ravin, N.V., Mardanov, A.V., Beletsky, A.V., Kublanov, I.V., Kolganova, T.V., Lebedinsky, A.V., Chernyh, N.A., Bonch-Osmolovskaya, E.A., Skryabin, K.G.
<2>Complete genome sequence of the anaerobic, protein-degrading hyperthermophilic crenarchaeon Desulfurococcus kamchatkensis.
<3>J. Bacteriol.
<4>191
<5>2371-2379
<6>2009
<7>Desulfurococcus kamchatkensis is an anaerobic organotrophic hyperthermophilic crenarchaeon
isolated from a terrestrial hot spring. Its
genome consists of a single circular chromosome of 1365223 bp with no
extrachromosomal elements. A total of 1474 protein-coding genes were
annotated of which 205 are exclusive for D. kamchatkensis. The search for
a replication origin site revealed a single region coinciding with a
global extreme of the nucleotide composition disparity curve and
containing a set of crenarchaeal-type origin recognition boxes. Unlike in
most archaea, two genes encoding homologs of eukaryotic initiator proteins
Orc1/Cdc6 are located distantly from this site. A number of mobile
elements are present in the genome, including seven transposons
representing IS607 and IS200/IS605 families, and multiple copies of
miniature inverted repeat transposable elements. Two large clusters of
regularly interspaced repeats are present; none of the spacer sequences
matches with known archaeal extrachromosomal elements except that one
spacer matches the sequence of a resident gene of D. kamchatkensis. Many
of the predicted metabolic enzymes are associated with the fermentation of
peptides and sugars, including more than thirty peptidases with diverse
specificities, a number of polysaccharide degradation enzymes, and many
transporters. Consistently, the genome encodes both enzymes of the
modified Embden-Meyerhof pathway of glucose oxidation and a set of enzymes
needed for gluconeogenesis. The genome structure and content reflect the
organism's nutritionally diverse, competitive natural environment
periodically invaded by viruses and other mobile elements.

<>

<1>Ravin, V., Raisanen, L., Alatossava, T.
<2>A conserved C-terminal region in Gp71 of the small isometric-head phage LL-H and ORF474 of the prolate-head phage JCL1032 is implicated in  specificity of adsorption of phage to its host, Lactobacillus  delbrueckii.
<3>J. Bacteriol.
<4>184
<5>2455-2459
<6>2002
<7>Thirty-five phage-resistant mutants of Lactobacillus delbrueckii subsp. lactis ATCC 15808 were
selected. Thirty-three of these mutants were
assigned to the Bes group, while the remaining two were grouped under
the Ads designation. Bes group mutants adsorbed phage LL-H but did not
allow efficient phage development. Preliminary evidence suggests that
these strains exhibit a mutation that changes the DNA specificity of a
restriction-modification system. The Ads group mutants did not adsorb
the small isometric-head phage LL-H. The results suggest that there are
at least three different types of phage receptors in L. delbrueckii:
two that are specific for small isometric-head phages and one that is
specific for prolate-head phage JCL1032. Five LL-H host-range mutants
which could overcome the adsorption block (a-type mutants) were
selected and investigated by sequencing the genes g71 and g17, which
encode minor and major tail proteins, respectively. Each of the a-type
mutants carried a nucleotide change at the 3' end of gene g71. No
mutations were observed in gene g17. Comparison of the gene product of
g71 of phage LL-H with its homolog in JCL1032 (ORF474) showed that
these proteins had very similar C-terminal regions. No similarities
were found at the N-terminal part of the proteins. We conclude that the
C-terminal portion of the protein encoded by g71 of phage LL-H and its
homolog in phage JCL1032 determines the adsorption specificities of
these phages on L. delbrueckii.

<>

<1>Ravin, V., Ravin, N., Casjens, S., Ford, M.E., Hatfull, G.F., Herdrix, R.W.
<2>Genomic sequence and analysis of the atypical temperate bacteriophage N15.
<3>J. Mol. Biol.
<4>299
<5>53-73
<6>2000
<7>N15 is a temperate bacteriophage that forms stable lysogens in Escherichia coli. While its
virion is morphologically very similar to
phage lambda and its close relatives, it is unusual in that the
prophage form replicates autonomously as a linear DNA molecule with
closed hairpin telomeres. Here, we describe the genomic architecture of
N15, and its global pattern of gene expression, which reveal that N15
contains several plasmid-derived genes that are expressed in N15
lysogens. The tel site, at which processing occurs to form the prophage
ends is close to the center of the genome in a similar location to that
occupied by the attachment site, attP, in lambda and its relatives and
defines the boundary between the left and right arms. The left arm
contains a long cluster of structural genes that are closely related to
those of the lambda-like phages, but also includes homologs of umuD',
which encodes a DNA polymerase accessory protein, and the plasmid
partition genes, sopA and sopB. The right arm likewise contains a
mixture of apparently phage- and plasmid-derived genes including genes
encoding plasmid replication functions, a phage repressor, a
transcription antitermination system, as well as phage host cell lysis
genes and two putative DNA methylases. The unique structure of the N15
genome suggests that the large global population of bacteriophages may
exhibit a much greater diversity of genomic architectures than was
previously recognized.

<>

<1>Rawal, C.M., Raval, V.H., Bhimani, H.D., Bhensdadia, D.V., Kothari, C.R., Patel, A.B., Bhatt, V.D., Parmar, N.R., Sajnani, M.R., Koringa, P.G., Joshi, C.G., Kothari, R.K., Singh, S.P.
<2>Whole-Genome Shotgun Sequencing of the Extremophile Alkalibacillus haloalkaliphilus C-5, of Indian Origin.
<3>J. Bacteriol.
<4>194
<5>4775
<6>2012
<7>Alkalibacillus haloalkaliphilus C-5 is a haloalkaliphilic bacterium that was isolated from a
soil sample from the salty Sambhar Lake, Rajasthan, India. The
organism is capable of alkaline protease production under conditions of pH 10 and
10% (wt/vol) salt. We sequenced and have reported the whole genome of
Alkalibacillus haloalkaliphilus C-5, of Indian origin, for the first time.

<>

<1>Rawat, S.R., Mannisto, M.K., Starovoytov, V., Goodwin, L., Nolan, M., Hauser, L., Land, M., Davenport, K.W., Woyke, T., Haggblom, M.M.
<2>Complete genome sequence of Granulicella tundricola type strain MP5ACTX9(T), an Acidobacteria from tundra soil.
<3>Standards in Genomic Sciences
<4>9
<5>449-461
<6>2014
<7>Granulicella tundricola strain MP5ACTX9(T) is a novel species of the genus Granulicella in
subdivision 1 Acidobacteria. G. tundricola is a predominant
member of soil bacterial communities, active at low temperatures and nutrient
limiting conditions in Arctic alpine tundra. The organism is a cold-adapted
acidophile and a versatile heterotroph that hydrolyzes a suite of sugars and
complex polysaccharides. Genome analysis revealed metabolic versatility with
genes involved in metabolism and transport of carbohydrates, including gene
modules encoding for the carbohydrate-active enzyme (CAZy) families for the
breakdown, utilization and biosynthesis of diverse structural and storage
polysaccharides such as plant based carbon polymers. The genome of G. tundricola
strain MP5ACTX9(T) consists of 4,309,151 bp of a circular chromosome and five
mega plasmids with a total genome content of 5,503,984 bp. The genome comprises
4,705 protein-coding genes and 52 RNA genes.

<>

<1>Rawat, S.R., Mannisto, M.K., Starovoytov, V., Goodwin, L., Nolan, M., Hauser, L., Land, M., Davenport, K.W., Woyke, T., Haggblom, M.M.
<2>Complete genome sequence of Terriglobus saanensis type strain SP1PR4(T), an Acidobacteria from tundra soil.
<3>Standards in Genomic Sciences
<4>7
<5>59-69
<6>2012
<7>SP1PR4 is a novel species of the genus . is of ecological interest because it is  a
representative of the phylum , which are dominant members of bacterial soil
microbiota in Arctic ecosystems. is a cold-adapted acidophile and a versatile
heterotroph utilizing a suite of simple sugars and complex polysaccharides. The
genome contained an abundance of genes assigned to metabolism and transport of
carbohydrates including gene modules encoding for carbohydrate-active enzyme
(CAZyme) family involved in breakdown, utilization and biosynthesis of diverse
structural and storage polysaccharides. SP1PR4 represents the first member of
genus with a completed genome sequence, consisting of a single replicon of
5,095,226 base pairs (bp), 54 RNA genes and 4,279 protein-coding genes. We infer
that the physiology and metabolic potential of is adapted to allow for resilience
to the nutrient-deficient conditions and fluctuating temperatures of Arctic
tundra soils.

<>

<1>Rawat, S.R., Mannisto, M.K., Starovoytov, V., Goodwin, L., Nolan, M., Hauser, L.J., Land, M., Davenport, K.W., Woyke, T., Haggblom, M.M.
<2>Complete genome sequence of Granulicella mallensis type strain MP5ACTX8(T), an acidobacterium from tundra soil.
<3>Standards in Genomic Sciences
<4>9
<5>71-82
<6>2013
<7>Granulicella mallensis MP5ACTX8(T) is a novel species of the genus Granulicella in subdivision
1of Acidobacteria. G. mallensis is of ecological interest being a
member of the dominant soil bacterial community active at low temperatures and
nutrient limiting conditions in Arctic alpine tundra. G. mallensis is a
cold-adapted acidophile and a versatile heterotroph that hydrolyzes a suite of
sugars and complex polysaccharides. Genome analysis revealed metabolic
versatility with genes involved in metabolism and transport of carbohydrates.
These include gene modules encoding the carbohydrate-active enzyme (CAZyme)
family involved in breakdown, utilization and biosynthesis of diverse structural
and storage polysaccharides including plant based carbon polymers. The genome of
Granulicella mallensis MP5ACTX8(T) consists of a single replicon of 6,237,577
base pairs (bp) with 4,907 protein-coding genes and 53 RNA genes.

<>

<1>Raychaudhuri, S., Saha, A., Ghoshal, T., Thakur, A.R.
<2>Draft Genome Sequence of Ammonia-Producing Aeromonas sp. MDS8 Strain MCC2167 from Sludge of a Dairy Effluent Treatment Plant.
<3>Genome Announcements
<4>1
<5>e00710-13
<6>2013
<7>The draft genome sequence of an amylase-, protease-, lipase-, oxidase-, and catalase-producing
Gram-negative bacillus (Aeromonas sp. MDS8 strain MCC2167)
with the ability to produce ammonia during 16 h of growth at 37 degrees C,
isolated from dairy sludge, with a size of 4,841,753 bp and a G+C content of
63.1%, is reported here.

<>

<1>Razin, A.
<2>Methylated bases in the single-stranded DNA phages.
<3>The Single-Stranded DNA Phages, Cold Spring Harbor Laboratory Press, Denhardt, D.T., Dressler, D., D.S. Ray., New York
<4>0
<5>165-175
<6>1978
<7>The widespread occurrence of methylated bases in the DNA of living organisms and the
species-specific pattern of DNA methylation (Hall 1971) strongly suggest that these methyl
groups play an important biological role.  At the present time, however, few biological
processes can be correlated with methylated bases in DNA.  Most of the methylated bases in the
bacterial chromosome are not related to the modificatiton activity in these cells, since
bacteria that are devoid of a restriction-modification (R-M) system still methylate their own
DNA.  The function of these methyl groups, as well as of those found in eukaryotic cell DNA,
is obscure.  The single-stranded DNA bacteriophages, which possess a few methyl groups per
genome, are ideal tools for studying the role of DNA methylation.  Thus, the filamentous
bacteriophages f1, fd, and M13 have been investigated with respect to the function of the
methylated bases in the R-M phenomena (Arber 1968).  The isometric phage PhiX174, which is not
subject to R-M and is propagated in Escherichia coli C, a strain devoid of any known R-M
system, was chosen to study the role of methylated bases not related to R-M.  A single methyl
group has been found in the PhiX genome and it has been suggested that this group is essential
in the final stages of phage maturation.

<>

<1>Razin, A.
<2>DNA methylases.
<3>Genet. Eng. (N Y), Plenum Press, Setlow, J.K., New York
<4>11
<5>1-11
<6>1989
<7>A review.

<>

<1>Razin, A., Friedman, J.
<2>DNA methylation and its possible biological roles.
<3>Prog. Nucleic Acid Res. Mol. Biol.
<4>25
<5>33-52
<6>1981
<7>Bases modified by methylation have been known to occur at a low frequency in DNA for more than
three decades.  This modification of DNA is carried out by specific methyltransferases (DNA
methylases) that transfer the chemically active methyl group from S-adenosylmethionine
(AdoMet) to either carbon 5 of cytosine residues or the exocyclic amino group attached to
carbon 6 of adenine residues of the DNA chain.
I. Introduction.
II.  Methylases and their specificity.
A. Substrate specificity.
B. Sequence specificity.
III. Distribution of methylated bases along the chromosome.
A. Distribution with respect to DNA sequences.
B. Distribution with respect to chromosomal proteins.
C. Distribution with respect to chromosome ultrastructure.
D. Methylated and unmethylated domains.
IV. The mode of methylation in vivo.
A. Semiconservative methylation.
B. DeNovo methylation.
C. "Origins" of methylation.
V. Possible functions of methylated bases in DNA.
A. Cell differentiation and gene activity.
B. Restriction and modification.
C. Interplay between DNA replication and methylation.
D. Mutation, recombination, and repair.
VI. Conclusions and prospects. References.

<>

<1>Razin, A., Goren, D., Friedman, J.
<2>Studies on the biological role of DNA methylation:  inhibition of methylation and maturation of the bacteriophage PhiX174 by nicotinamide.
<3>Nucleic Acids Res.
<4>2
<5>1967-1974
<6>1975
<7>Nicotinamide was found to be a potent inhibitor of DNA methylation in vivo without
interferring with protein or DNA synthesis.  The inhibition of DNA methylation in a
phage-infected cell resulted in a parallel decrease in the production of viable virus
particles.  In vitro experiments revealed that nicotinamide inhibits DNA methylase activity in
a competitive fashion with respect to S-adenosylmethionine and non-competitively with respect
to DNA. These results were interpreted to mean that DNA methylation is an essential step in
the process of maturation of the bacteriophage PhiChi174.

<>

<1>Razin, A., Razin, S.
<2>Methylated bases in mycoplasmal DNA.
<3>Nucleic Acids Res.
<4>8
<5>1383-1390
<6>1980
<7>The DNAs of four Mycoplasma and one Acholeplasma species were found to contain methylated
bases.  All of the five species contained 6-methyladenine (m6Ade), the methylated base
characteristic of prokaryotic DNA.  The extent of methylation of adenine residues in the
mycoplasmal DNA ranged from 0.2% in Mycoplasma capricolum to about 2% in Mycoplasma arginini
and Mycoplasma hyorhinis with intermediate methylation values for Mycoplasma orale and
Acholeplasma laidlawii DNAs.  About 5.8% of the cytosine residues in M. hyorhinis DNA were
methylated also.  Analysis of cell culture DNA for the presence of m6Ade as a means for
detection of contamination by mycoplasmas, and the phylogenetic implications of the finding of
methylated bases in mycoplasmal DNAs are discussed.

<>

<1>Razin, A., Riggs, D.
<2>DNA methylation and gene function.
<3>Science
<4>210
<5>604-610
<6>1980
<7>In most higher organisms, DNA is modified after synthesis by the enzymatic conversion of many
cytosine residues to 5-methylcytosine.  For several years, control of gene activity by DNA
methylation has been recognized as a logically attractive possibility, but experimental
support has proved elusive.  However, there is now reason to believe, from recent studies,
that DNA methylation is a key element in the hierarchy of control mechanisms that govern
vertebrate gene function and differentiation.

<>

<1>Razin, A., Rottem, S., Renbaum, P.F.
<2>Method for producing the Spiroplasma SP. DNA methylase.
<3>European Patent Office
<4>EP 0412676 B
<5>
<6>1995
<7>The present invention is directed to a method for cloning and producing the Spiroplasma sp.
strain MQ1 DNA methylase by (1) introducing the Spiroplasma methylase gene into a host whereby
the methylase gene is expressed; (2) fermenting the host which contains the vector encoding
and expressing the Spiroplasma methylase and (3) purifying the Spiroplasma methylase from the
fermented host which contains the vector encoding and expressing the Spiroplasma DNA methylase
activity.

<>

<1>Razin, A., Rottem, S., Renbaum, P.F.
<2>Method for producing the Spiroplasma species.
<3>Japanese Patent Office
<4>JP 2538806
<5>
<6>1996
<7>
<>

<1>Razin, A., Rottem, S., Renbaum, P.F.
<2>DNA encoding Spiroplasma sp. DNA methylase.
<3>US Patent Office
<4>US 5296371
<5>
<6>1994
<7>The present invention is directed to a method for cloning and producing the Spiroplasma sp.
strain MQ1 DNA methylase by (1) introducing the Spiroplasma methylase gene into a host whereby
the methylase gene is expressed; (2) fermenting the host which contains the vector encoding
and expressing the Spiroplasma methylase and (3) purifying the Spiroplasma methylase from the
fermented host which contains the vector encoding and expressing the Spiroplasma DNA methylase
activity.

<>

<1>Razin, A., Sedat, J.
<2>Analysis of 5-methylcytosine in DNA.
<3>Anal. Biochem.
<4>77
<5>370-377
<6>1977
<7>A method for analyzing 5-methylcytosine in DNA by gas chromatography is described.  The method
is based on degradation of the DNA to its free bases by treatment with trifluoroacetic acid
and gas chromatography of the trimethylsilyl derivatives of the free bases.  Chromatography of
microgram amounts of derivatized material is conducted at isothermal conditions using a 3%
SE-30 or 2% OV-225 column.  The peak areas corresponding to cytosine and 5-methylcytosine are
used to calculate the 5-methylcytosine/cytosine molar ratio in DNA.  The lower limit for
detection of 5-methylcytosine in DNA by this method is a 5-methylcytosine/cytosine molar ratio
of 0.001.

<>

<1>Razin, A., Urieli, S., Pollack, Y., Gruenbaum, Y., Glaser, G.
<2>Studies on the biological role of DNA methylation; IV.  Mode of methylation of DNA in E. coli cells.
<3>Nucleic Acids Res.
<4>8
<5>1783-1792
<6>1980
<7>Two pairs of restriction enzyme isoschizomers were used to study in vivo
methylation of E. coli and extrachromosomal DNA.  By use of the restriction
enzymes MboI (which cleaves only the unmethylated GATC sequence) and its
isoschizomer Sau3A (indifferent to a methylated adenine at this sequence), we
found that all the GATC sites in E. coli and in extrachromosomal DNAs are
symmetrically methylated on both strands.  The calculated number of GATC sites
in E. coli DNA can account for all its m6Ade residues.  Foreign DNA, like mouse
mtDNA, which is not methylated at GATC sites became fully methylated at these
sequences when introduced by transfection into E. coli cells.  This experiment
provides the first evidence for the operation of a de novo methylation
mechanism for E. coli methylases not involved in restriction modification.
When the two restriction enzyme isoschizomers, EcoRII and ApyI, were used to
analyze the methylation pattern of CCA/TGG sequences in E. coli C and PhiX174
DNA, it was found that all these sites are methylated.  The number of CCA/TGG
sites in E. coli C DNA does not account for all m5Cyt residues.

<>

<1>Read, T.D. et al.
<2>Genome sequences of Chlamydia trachomatis MoPn and Chlamydia pneumoniae AR39.
<3>Nucleic Acids Res.
<4>28
<5>1397-1406
<6>2000
<7>The genome sequences of Chlamydia trachomatis mouse pneumonitis (MoPn) strain Nigg (1 069 412
nt) and Chlamydia pneumoniae strain AR39 (1 229 853 nt) were determined using a random shotgun
strategy. The MoPn genome exhibited a general conservation of gene order and content with the
previously sequenced C. trachomatis serovar D. Differences between C. trachomatis strains were
focused on an ~50 kb 'plasticity zone' near the termination origins. In this region MoPn
contained three copies of a novel gene encoding a >3000 amino acid toxin homologous to a
predicted toxin from Escherichia coli 0157:H7 but had apparently lost the tryptophan
biosyntheis genes found in serovar D in this region. The C. pneumoniae AR39 chromosome was
>99.9% identical to the previously sequenced C. pneumoniae CWL029 genome, however, comparative
analysis identified an invertible DNA segment upstream of the uridine kinase gene which was in
different orientations in the two genomes. AR39 also contained a novel 4524 nt circular
single-stranded (ss)DNA bacteriophage, the first time a virus has been reported infecting C.
pneumoniae. Although the chlamydial genomes were highly conserved, there were intriguing
differences in key nucleotide salvage pathways: C. pneumoniae has a uridine kinase gene for
dUTP production, MoPn has a uracil phosphororibosyl transferase, while C. trachomatis serovar
D contains neither gene. Chromosomal comparison revealed that there had been multiple large
inversion events since the species divergence of C. trachomatis and C. pneumoniae, apparently
oriented around the axis of the origin of replication and the termination region. The striking
synteny of the Chlamydia genomes and prevalence of tandemly duplicated genes are evidence of
minimal chromosome rearrangement and foreign gene uptake, presumably owing to the ecological
isolation of the obligate intracellular parasites. In the absence of genetic analysis,
comparative genomics will continue to provide insight into the virulence mechanisms of these
important human pathogens.

<>

<1>Read, T.D. et al.
<2>The genome sequence of Bacillus anthracis Ames and comparison to closely related bacteria.
<3>Nature
<4>423
<5>81-86
<6>2003
<7>Bacillus anthracis is an endospore-forming bacterium that causes inhalational anthrax. Key
virulence genes are found on plasmids
(extra-chromosomal, circular, double-stranded DNA molecules) pXO1 (ref. 2)
and pXO2 (ref. 3). To identify additional genes that might contribute to
virulence, we analysed the complete sequence of the chromosome of B.
anthracis Ames (about 5.23 megabases). We found several chromosomally
encoded proteins that may contribute to pathogenicity--including
haemolysins, phospholipases and iron acquisition functions--and identified
numerous surface proteins that might be important targets for vaccines and
drugs. Almost all these putative chromosomal virulence and surface
proteins have homologues in Bacillus cereus, highlighting the similarity
of B. anthracis to near-neighbours that are not associated with anthrax.
By performing a comparative genome hybridization of 19 B. cereus and
Bacillus thuringiensis strains against a B. anthracis DNA microarray, we
confirmed the general similarity of chromosomal genes among this group of
close relatives. However, we found that the gene sequences of pXO1 and
pXO2 were more variable between strains, suggesting plasmid mobility in
the group. The complete sequence of B. anthracis is a step towards a
better understanding of anthrax pathogenesis.

<>

<1>Read, T.D. et al.
<2>Genome sequence of Chlamydophila caviae (Chlamydia psittaci GPIC): examining the role of niche-specific genes in the evolution of the   Chlamydiaceae.
<3>Nucleic Acids Res.
<4>31
<5>2134-2147
<6>2003
<7>The genome of Chlamydophila caviae (formerly Chlamydia psittaci, GPIC isolate) (1 173 390 nt
with a plasmid of 7966 nt) was determined,
representing the fourth species with a complete genome sequence from the
Chlamydiaceae family of obligate intracellular bacterial pathogens. Of
1009 annotated genes, 798 were conserved in all three other completed
Chlamydiaceae genomes. The C.caviae genome contains 68 genes that lack
orthologs in any other completed chlamydial genomes, including tryptophan
and thiamine biosynthesis determinants and a ribose-phosphate
pyrophosphokinase, the product of the prsA gene. Notable amongst these was
a novel member of the virulence-associated invasin/intimin family (IIF) of
Gram-negative bacteria. Intriguingly, two authentic frameshift mutations
in the ORF indicate that this gene is not functional. Many of the unique
genes are found in the replication termination region (RTR or plasticity
zone), an area of frequent symmetrical inversion events around the
replication terminus shown to be a hotspot for genome variation in
previous genome sequencing studies. In C.caviae, the RTR includes several
loci of particular interest including a large toxin gene and evidence of
ancestral insertion(s) of a bacteriophage. This toxin gene, not present in
Chlamydia pneumoniae, is a member of the YopT effector family of type
III-secreted cysteine proteases. One gene cluster (guaBA-add) in the RTR
is much more similar to orthologs in Chlamydia muridarum than those in the
phylogenetically closest species C.pneumoniae, suggesting the possibility
of horizontal transfer of genes between the rodent-associated Chlamydiae.
With most genes observed in the other chlamydial genomes represented,
C.caviae provides a good model for the Chlamydiaceae and a point of
comparison against the human atherosclerosis-associated C.pneumoniae. This
crucial addition to the set of completed Chlamydiaceae genome sequences is
enabling dissection of the roles played by niche-specific genes in these
important bacterial pathogens.

<>

<1>Read, T.D., Salzberg, S.L., Pop, M., Shumway, M., Umayam, L., Jiang, L., Holtzapple, E., Busch, J.D., Smith, K.L., Schupp, J.M., Solomon, D., Keim, P., Fraser, C.M.
<2>Comparative genome sequencing for discovery of novel polymorphisms in Bacillus anthracis.
<3>Science
<4>296
<5>2028-2033
<6>2002
<7>Comparison of the whole-genome sequence of Bacillus anthracis isolated from a victim of a
recent bioterrorist anthrax attack with a reference
reveals 60 new markers that include single nucleotide polymorphisms
(SNPs), inserted or deleted sequences, and tandem repeats. Genome
comparison detected four high-quality SNPs between the two sequenced B.
anthracis chromosomes and seven differences among different preparations
of the reference genome. These markers have been tested on a collection of
anthrax isolates and were found to divide these samples into distinct
families. These results demonstrate that genome-based analysis of
microbial pathogens will provide a powerful new tool for investigation of
infectious disease outbreaks.

<>

<1>Read, T.D., Thomas, A.T., Wilkins, B.M.
<2>Evasion of type I and type II DNA restriction systems by IncI1 plasmid Collb-P9 during transfer by bacterial conjugation.
<3>Mol. Microbiol.
<4>6
<5>1933-1941
<6>1992
<7>Transmission of unmodified plasmid CoIIb-P9 by bacterial conjugation is markedly resistant to
restriction compared with transfer by transformation. One process allowing evasion of type I
and II restriction systems involves conjugative transfer of multiple copies of the plasmid. A
more specialized evasion mechanism requires the Ard (alleviation of restriction of DNA) system
encoded by CoIIb. The ard gene is transferred early in conjugation and specifically alleviates
DNA restriction by all known families of type I enzyme, including EcoK.  CoIIb has no effect
on EcoK modification but this activity is impaired by multicopy recombinant plasmids
supporting overexpression of ard. Genetic evidence shows that Ard protects CoIIb from EcoK
restriction following conjugative transfer and that this protection requires expression of the
gene on the immigrant plasmid. It is proposed that carriage of ard facilitates transfer of
CoIIb between its natural enterobacterial hosts and that the route of DNA entry is important
to the restriction-evasion mechanism.

<>

<1>Reale, A., Lindsay, H., Saluz, H.P., Pradhan, S., Adams, R.L.P., Jost, J.-P., Strom, R.
<2>DNA binding and methyl transfer catalysed by mouse DNA methyltransferase.
<3>Biochem. J.
<4>312
<5>855-861
<6>1995
<7>By using a purified fraction of mouse DNA methyltransferase we have shown, by gel-retardation
analysis, that the enzyme forms a low-affinity complex preferentially with hemimethylated DNA;
the complexes formed with unmethylated or with fully methylated DNA are of even lower
affinity, and only very weak interaction occurs with DNA lacking CG dinucleotides.
Interaction is inhibited by N-ethylmaleimide.  Methyl transfer from S-adenosyl-methionine is
associated with the release of the fully methylated product from the complex.  Complexes
formed with the intact enzyme are extremely large, but limited trypsin treatment allows a
major complex to enter the gel.  DNA binding is not inhibited by this limited proteolysis of
the native enzyme.

<>

<1>Reaston, J., Duyvesteyn, M.G.C., de Waard, A.
<2>Nostoc PCC7524, a cyanobacterium which contains five sequence-specific deoxyribonucleases.
<3>Gene
<4>20
<5>103-110
<6>1982
<7>Five nucleotide sequence-specific deoxyribonucleases present in cell-free
extracts of the filamentous cyanobacterium Nostoc PCC7524 have been purified
and characterized.  One of these enzymes, designated Nsp(7524)I cleaves at a
new kind of nucleotide sequence, i.e. 5'-PuCATG^Py-3'.  The other four
restriction enzymes in this organism, designated Nsp(7524)II, Nsp(7524)III,
Nsp(7524)IV and Nsp(7524)V, are isoschizomers of enzymes which have been
previously described.  The cleavage site of Nsp(7524)II which is an
isoschizomer of SduI was determined.

<>

<1>Rebentish, B.A., Bolotin, A.P., Grachova, I.M., Ren, L.S., Uyan, J.M.
<2>A new site-specific endodeoxyribonuclease FmuI from the strain Flavobacterium multivorum.
<3>Biotekhnologiya
<4>3
<5>15-16
<6>1994
<7>A new site-specific endodeoxyribonuclease (restrictase) has been isolated from the strain
Flavobacterium multivorum and some of its characteristics were examined. It was named FmuI and
shown to determine the following nucleotide sequence: 5'-GGNC/C-3'. The new restrictase
turned out to be a false isoschizomer (or an alternative prototype) of the enzyme AsuI.

<>

<1>Rechkunova, N.I., Lokhov, S.G., Gorbunov, Y.A., Zinovev, V.V., Buryanov, Y.I., Malygin, E.G.
<2>Stability of the secondary structure of the oligonucleotide substrates.  The effect of Ecodam DNA-methylase.
<3>Biopol. Kletka
<4>5
<5>43-49
<6>1989
<7>Oligonucleotide complexes containing various defects in the Ecodam methylase recognition site
have been investigated for their stability. A partial duplex structure for the single-stranded
20-base long oligonucleotide containing a self-complementary hexanucleotide sequence is
observed only below 5C. Other complexes melt within the narrow temperature range of 22-31C in
30 mM of potassium-phosphate buffer, pH 7.8. The presence of the Ecodam methylase results in
an increase of the melting point of the complex at least by 5C.

<>

<1>Rechkunova, N.I., Ovetchkina, L.G., Jashina, L.N., Vtorushina, I.A., Gorbunov, J.A., Zinoviev, V.V., Malygin, E.G.
<2>Effects of oligonucleotide structure on the kinetic values of the interaction with BamHI endonuclease.
<3>Mol. Biol. (Mosk)
<4>22
<5>217-223
<6>1988
<7>Kinetic values of the BamHI endonuclease interaction with synthetic oligonucleotides,
containing some defects, have been determined.  These defects were: the absence of the one
internucleotide phosphate in the GGATCC sequence; substitution of a phosphate linkage by a
methylphosphonate one; 5'-protruding end of the double-stranded oligonucleotide substrate.
Some modifications resulted in the increase of the initial rates of cleavage due to higher
Vmax values for these substrates.  Several structural defects in the oligonucleotide
substrates have been shown to intensify the formation of productive complexes with the enzyme,
which can be explained by the significant role of the polynucleotide chain kinds in the
recognition process.  Studies on oligonucleotides with different defects made it possible to
reveal the phosphate groups essential for the interaction with BamHI endonuclease.

<>

<1>Rechkunova, N.I., Zinovev, V.V., Malygin, E.G., Gorbunov, Y.A., Popov, S.G., Nesterenko, V.F., Buryanov, Y.I.
<2>Dimerization of Ecodam-methylase induced by the oligonucleotide substrate.
<3>Biopol. Kletka
<4>3
<5>152-154
<6>1987
<7>The Ecodam-methylase has been investigated for its interaction with the 20-base
oligonucleotide duplex containing the enzyme recognition site. An increase of the enzyme
molecular weight was detected by the gel-filtration and sucrose density gradient
centrifugation methods. The maximal value of the complex molecular weight was observed when
concentrations of the enzyme and the substrate were equal. The results obtained prove the
formation of the dimeric enzyme form in the presence of the substrate.

<>

<1>Reckmann, B., Krauss, G.
<2>The cleavage of single-stranded DNA by the isoschizomeric restriction endonuclease HhaI and CfoI.
<3>Biochim. Biophys. Acta
<4>908
<5>90-96
<6>1987
<7>The cleavage of single-stranded (ss) M13mp8(+) DNA by the isoschizomeric restriction
endonucleases HhaI and CfoI has been investigated.  The two enzymes differ considerably in
their ability to cleave ssDNA.  HhaI cleaves ssDNA about two orders of magnitude faster than
does CfoI, although both enzymes show the same activity when assayed on double-stranded DNA.
From the cleavage of oligonucleotides and of M13mp8(+) DNA fragments it is concluded that
cleavage of ssDNA occurs via transiently formed double-stranded hairpin structures.  A rough
correlation exists between the stability of the secondary structures and the cleavage
efficiency.

<>

<1>Reckmann, B., Rieke, E.
<2>Determination of nucleic acids - hybridization with single-stranded nucleic acid probes containing 5-aza-cytidine or 2-deoxy-5-aza-cytidine and detection with DNA-methylase.
<3>European Patent Office
<4>EP 0286958
<5>
<6>1987
<7>In a new procedure for the determination of nucleic acids in a specimen by conversion of the
nucleic acid sequences to be determined into single strands and reaction of the single strands
with a complementary polynucleotide probe, cytidine in the polynucleotide probe is replaced by
5-aza-2'-deoxy-cytidine (I) or 5-azacytidine (II) and the probe is detected by binding of
optionally labelled DNA-methylase and detection of the methylase by means of specific,
optionally labelled anti-methylase antibodies by means of the label. New agents for the
determination of nucleic acids contain (1) a polynucleotide probe containing
5-aza-2'-deoxycytidine or 5-azacytidine in place of cytidine, (2) an optimally labelled
methylase, and optionally (3) labelled anti-methylase antibodies.

<>

<1>Recktenwald, J., Schmidt, H.
<2>The nucleotide sequence of Shiga toxin (Stx) 2e-encoding phage phiP27 is not related to other Stx phage genomes, but the modular genetic structure is Conserved.
<3>Infect. Immun.
<4>70
<5>1896-1908
<6>2002
<7>In this study we determined the complete nucleotide sequence of Shiga
toxin 2e-encoding bacteriophage phi P27, isolated from the Shiga
toxin-producing Escherichia coli patient isolate 2771/97. phi P27 is
integrated as a prophage in the chromosomal yecE gene. This integration
generates identity segments of attL and attR sites with lengths of 11
nucleotides. The integrated prophage genome has a size of 42,575 bp. We
identified 58 open reading frames (ORFs), each with a length of >150
nucleotides. The deduced proteins of 44 ORFs showed significant homologies
to other proteins present in sequence databases, whereas 14 putative
proteins did not. For 29 proteins, we could deduce a putative function.
Most of these are related to the basic phage propagation cycle. The phi
P27 genome represents a mosaic composed of genetic elements which are
obviously derived from related and unrelated phages. We identified five
short linker sequences of 22 to 151 bp in the phi P27 sequence which have
also been detected in a couple of other lambdoid phages. These linkers are
located between functional modules in the phage genome and are thought to
play a role in genetic recombination. Although the overall DNA sequence of
phi P27 is not highly related to other known phages, the data obtained
demonstrate a typical lambdoid genome structure.

<>

<1>Redaschi, N., Bickle, T.A.
<2>Posttranscriptional regulation of EcoP1I and EcoP15I restriction activity.
<3>J. Mol. Biol.
<4>257
<5>790-803
<6>1996
<7>Efficient establishment of a DNA restriction-modification (R-M) system in a non-modified cell
requires a tight control of the potentially lethal activity of the restriction enzyme.  The
type III R-M systems EcoP1I and EcoP15I can be transferred to non-modified Escherichia coli
cells by transfection, conjugation or transformation and become established without
difficulty.  Modification activity is expressed immediately after the R-M genes enter the
cell, whereas the expression of restriction activity is delayed until complete protection of
the cellular DNA is achievedby methylation.  We have shown by Western blot analysis that the
expression of the modification polypeptide subunit positively regulates the amount of
restriction subunit present in the cell.  The finding that ribosomal alterations affected the
expression of restriction activity pointed to additional control at the translational level.
The analysis of EcoP1I expression in E. coli strains mutated in either of the ribosomal
proteins S12 (rpsL) or S4 (rpsD) suggests that the level of in vivo restriction activity can
be modulated both by a decrease in the efficiency of translation and by varying ribosomal
accuracy conditions.  In addition, we have preliminary evidence from in vivo gene fusion
studies that the res gene may code for more than one gene product.

<>

<1>Redaschi, N., Bickle, T.A.
<2>DNA restriction and modification systems.
<3>Escherichia coli and Salmonella: Cellular and Molecular Biology, ASM Press, F.C. Neidhardt, Washington, DC
<4>1
<5>773-781
<6>1996
<7>The phenomenon of restriction and modification was first observed in the course of studies on
bacterial viruses in the early 1950s.  Several authors reported that certain strains of
bacteria inhibited ("restricted") the growth of bacterial viruses previously propagated on a
different strain.  The molecular explanation for this effect was discovered in the early
1960s: the restriction of viral growth was due to endonucleolytic cleavage of the viral DNA by
site-specific endonucleases.  Some of these restriction endonucleases were found to possess
very useful properties, and their subsequent exploitation in the 1970s was the key to the
development of genetic engineering technology.

<>

<1>Reddy, G.S., Ara, S., Singh, A., Kumar, P.A., Shivaji, S.
<2>Draft Genome Sequence of Psychrobacter aquaticus Strain CMS 56T, Isolated from a  Cyanobacterial Mat Sample Collected from Water Bodies in the McMurdo Dry Valley  Region of Antarctica.
<3>Genome Announcements
<4>1
<5>e00918-13
<6>2013
<7>We report the 3.2-Mb draft genome sequence of Psychrobacter aquaticus strain CMS  56(T),
isolated from a cyanobacterial mat sample collected from a water body in
the McMurdo Dry Valley region of Antarctica.

<>

<1>Reddy, G.S., Sreenivas, A., Shivaji, S.
<2>Draft Genome Sequence of Cryobacterium roopkundensis Strain RuGl7, Isolated from  a Soil Sample in the Vicinity of Roopkund Lake, Himalayas, India.
<3>Genome Announcements
<4>2
<5>e01206-14
<6>2014
<7>We report the 4.36-Mb genome of Cryobacterium roopkundensis strain RuGl7, isolated from a soil
sample collected in the periphery of Roopkund Lake, Himalayas, India. The draft genome
consists of 4,356,863 bp, 4,048 protein-coding sequences, and 50 RNAs, with 65.3% G+C DNA
content.

<>

<1>Reddy, Y.V., Rao, D.N.
<2>Binding of EcoP15I DNA methyltransferase to DNA reveals a large structural distortion within the recognition sequence.
<3>J. Mol. Biol.
<4>298
<5>597-610
<6>2000
<7>EcoP15I DNA methyltransferase, a member of the type III restriction-modification system, binds
to the sequence 5'-CAGCAG-3' transferring a methyl group from S-adenosyl-l-methionine to the
second adenine base. We have investigated protein-DNA interactions in the methylase-DNA
complex by three methods. Determination of equilibrium dissociation constants indicated that
the enzyme had higher affinity for DNA containing mismatches at the target base within the
recognition sequence. Potassium permanganate footprinting studies revealed that there was a
hyper-reactive permanganate cleavage site coincident with adenine that is the target base for
methylation. More importantly, to detect DNA conformational alterations within the enzyme-DNA
complexes, we have used a fluorescence-based assay. When EcoP15I DNA methyltransferase bound
to DNA containing 2-aminopurine substitutions within the cognate sequence, an eight to tenfold
fluorescent enhancement resulting from enzymatic flipping of the target adenine base was
observed. Furthermore, fluorescence spectroscopy analysis showed that the changes attributable
to structural distortion were specific for only the bases within the recognition sequence.
More importantly, we observed that both the adenine bases in the recognition site appear to be
structurally distorted to the same extent. While the target adenine base is probably flipped
out of the DNA duplex, our results also suggest that fluorescent enhancements could be derived
from protein-DNA interactions other than base flipping. Taken together, our results support
the proposed base flipping mechanism for adenine methyltransferases.

<>

<1>Reddy, Y.V.R., Rao, D.N.
<2>Probing the role of cysteine residues in the EcoP15I DNA methyltransferase.
<3>J. Biol. Chem.
<4>273
<5>23866-23876
<6>1998
<7>Chemical modification using thiol-directed agents and site-directed mutagenesis has been used
to investigate the role of cysteine residues of EcoP15I DNA methyltransferase.  Irreversible
inhibition of enzymatic activity was provoked by chemical modification of the enzyme by
N-ethylmaleimide and iodoacetamide.  5,5'-Dithiobis(2-nitrobenzoic acid) titration of the
enzyme under nondenaturing and denaturing conditions confirmed the presence of six cysteine
residues without any disulfides in the protein.  Aware that relatively bulky reagents
inactivate the methyltransferase by directly occluding the substrate-binding site or by
locking the methyltransferase in an inactive conformation, we used site-directed mutagenesis
to sequentially replace each of the six cysteines in the protein at positions 30, 213, 344,
434, 553, and 577.  All the resultant mutation methylases except for the C344S and C344A
enzymes retained significant activity as assessed by in vivo and in vitro assays.  The effects
of the substitutions on the function of EcoP15I DNA methyltransferase were investigated by
substrate binding assays, activity measurements, and steady-state kinetic analysis of
catalysis.  Our results clearly indicate that the cysteines at positions other than 344 are
not essential for activity.  In contrast, the C344A enzyme showed a marked loss of enzymatic
activity.  More importantly, whereas the inactive C344A mutant enzyme bound
S-adenosyl-L-methionine, it failed to bind to DNA.  Furthermore, in double and triple mutants
where two or three cysteine residues were replaced by serine, all such mutants in which the
cysteine at position 344 was changed, were inactive.  Taken together, these results
convincingly demonstrate that the Cys-344 is necessary for enzyme activity and indicate an
essential role for it in DNA binding.

<>

<1>Redenbach, M., Ikeda, K., Yamasaki, M., Kinashi, H.
<2>Cloning and physical mapping of the EcoRI fragments of the giant linear plasmid SCP1.
<3>J. Bacteriol.
<4>180
<5>2796-2799
<6>1998
<7>A cosmid library was constructed for the 350-kb giant linear plasmid SCP1
and aligned on a successive linear map. Only a 0.8-kb gap has remained
uncloned in the terminal inverted repeats close to both ends. Partial
digestion of the aligned cosmids with EcoRI and hybridization with the
flanking fragments of the vector enabled physical mapping of all of the
EcoRI fragments. On this map, the methylenomycin biosynthetic gene
cluster, the insertion sequence IS466, and the sapCDE genes coding for
spore-associated proteins were localized.

<>

<1>Redenbach, M., Kieser, H.M., Denapaite, D., Eichner, A., Cullum, J., Kinashi, H., Hopwood, D.A.
<2>A set of ordered cosmids and a detailed genetic and physical map for the 8 Mb Streptomyces coelicolor A3(2) chromosome.
<3>Mol. Microbiol.
<4>21
<5>77-96
<6>1996
<7>A Supercos-1 library carrying chromosomal DNA of a plasmid-free derivative of Streptomyces
coelicolor A3(2) was organized into an ordered encyclopaedia of overlapping clones by
hybridization. The minimum set of overlapping clones representing the entire chromosome (with
three short gaps) consists of 319 cosmids. The average insert size is 37.5 kb and the set of
clones therefore divides the chromosome into 637 alternating unique and overlapping segments
which have an average length of approx. 12.5 kb. More than 170 genes, gene clusters and other
genetic markers were mapped to their specific segment by hybridization to the encyclopaedia.
Genes could be cloned by direct transformation and complementation of S. coelicolor mutants
with cosmids isolated from Escherichia coli, selecting for insertion into the chromosome by
homologous recombination. As in other streptomycetes, the ends of the chromosome have long
terminal inverted repeats.

<>

<1>Redondo, P., Merino, N., Villate, M., Blanco, F.J., Montoya, G., Molina, R.
<2>Crystallization and preliminary X-ray diffraction analysis of the homing endonuclease I-CvuI from Chlorella vulgaris in complex with its target DNA.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>70
<5>256-259
<6>2014
<7>Homing endonucleases are highly specific DNA-cleaving enzymes that recognize long stretches of
DNA. The engineering of these enzymes provides novel instruments for genome modification in a
wide range of fields, including gene targeting, by inducing specific double-strand breaks.
I-CvuI is a homing endonuclease from the green alga Chlorella vulgaris. This enzyme was
purified after overexpression in Escherichia coli. Crystallization experiments of I-CvuI in
complex with its DNA target in the presence of Mg2+ yielded crystals suitable for X-ray
diffraction analysis. The crystals belonged to the orthorhombic space group P2(1)2(1)2(1),
with unit-cell parameters a = 62.83, b = 83.56, c = 94.40 angstrom. The self-rotation function
and the Matthews coefficient suggested the presence of one protein-DNA complex per asymmetric
unit. The crystals diffracted to a resolution limit of 1.9 angstrom using synchrotron
radiation.

<>

<1>Redondo, P., Prieto, J., Munoz, I.G., Alibes, A., Stricher, F., Serrano, L., Cabaniols, J.P., Daboussi, F., Arnould, S., Perez, C., Duchateau, P., Paques, F., Blanco, F.J., Montoya, G.
<2>Molecular basis of xeroderma pigmentosum group C DNA recognition by engineered meganucleases.
<3>Nature
<4>456
<5>107-111
<6>2008
<7>Xeroderma pigmentosum is a monogenic disease characterized by hypersensitivity to ultraviolet
light. The cells of xeroderma pigmentosum
patients are defective in nucleotide excision repair, limiting their
capacity to eliminate ultraviolet-induced DNA damage, and resulting in a
strong predisposition to develop skin cancers. The use of rare cutting DNA
endonucleases-such as homing endonucleases, also known as
meganucleases-constitutes one possible strategy for repairing DNA lesions.
Homing endonucleases have emerged as highly specific molecular scalpels
that recognize and cleave DNA sites, promoting efficient homologous gene
targeting through double-strand-break-induced homologous recombination.
Here we describe two engineered heterodimeric derivatives of the homing
endonuclease I-CreI, produced by a semi-rational approach. These two
molecules-Amel3-Amel4 and Ini3-Ini4-cleave DNA from the human XPC gene
(xeroderma pigmentosum group C), in vitro and in vivo. Crystal structures
of the I-CreI variants complexed with intact and cleaved XPC target DNA
suggest that the mechanism of DNA recognition and cleavage by the
engineered homing endonucleases is similar to that of the wild-type
I-CreI. Furthermore, these derivatives induced high levels of specific
gene targeting in mammalian cells while displaying no obvious
genotoxicity. Thus, homing endonucleases can be designed to recognize and
cleave the DNA sequences of specific genes, opening up new possibilities
for genome engineering and gene therapy in xeroderma pigmentosum patients
whose illness can be treated ex vivo.

<>

<1>Redondo, P., Prieto, J., Ramos, E., Blanco, F.J., Montoya, G.
<2>Crystallization and preliminary x-ray diffraction analysis on the homing endonuclease I-Dmo-I in complex with its target DNA.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>63
<5>1017-1020
<6>2007
<7>Homing endonucleases are highly specific DNA-cleaving enzymes that recognize long stretches of
base pairs. The availability of these
enzymes has opened novel perspectives for genome engineering in a wide
range of fields, including gene therapy, by taking advantage of the
homologous gene-targeting enhancement induced by a double-strand break.
I-Dmo-I is a well characterized homing endonuclease from the archaeon
Desulfurococcus mobilis. The enzyme was cloned and overexpressed in
Escherichia coli. Crystallization experiments of I-Dmo-I in complex
with its DNA target in the presence of Ca2+ and Mg2+ yielded crystals
that were suitable for X-ray diffraction analysis. The crystals
belonged to the monoclinic space group P2(1), with unit-cell parameters
a = 106.75, b = 70.18, c = 106.85 angstrom, alpha = gamma = 90, beta =
119.93 degrees. The self-rotation function and the Matthews coefficient
suggested the presence of three protein-DNA complexes per asymmetric
unit. The crystals diffracted to a resolution limit of 2.6 angstrom
using synchrotron radiation at the Swiss Light Source (SLS) and the
European Synchrotron Radiation Facility (ESRF).

<>

<1>Redondo-Nieto, M. et al.
<2>Genome Sequence of the Biocontrol Strain Pseudomonas fluorescens F113.
<3>J. Bacteriol.
<4>194
<5>1273-1274
<6>2012
<7>Pseudomonas fluorescens F113 is a plant growth-promoting rhizobacterium (PGPR) that has
biocontrol activity against fungal plant pathogens and is a model for
rhizosphere colonization. Here, we present its complete genome sequence, which
shows that besides a core genome very similar to those of other strains sequenced
within this species, F113 possesses a wide array of genes encoding specialized
functions for thriving in the rhizosphere and interacting with eukaryotic
organisms.

<>

<1>Redzuan, R.A., Abu, B.N., Rozano, L., Badrun, R., Mat, A.N., Mohd, R.M.F.
<2>Draft Genome Sequence of Erwinia mallotivora BT-MARDI, Causative Agent of Papaya  Dieback Disease.
<3>Genome Announcements
<4>2
<5>e00375-14
<6>2014
<7>Erwinia mallotivora was isolated from papaya trees infected with dieback disease, which were
planted at the Malaysian Agricultural Research and Development
Institute (MARDI), Malaysia. Here, we report a draft genome sequence of E.
mallotivora BT-MARDI, which offers an important source of information for
understanding pathogen and host interaction during papaya dieback development.

<>

<1>Reeben, M., Prydz, H.
<2>An improved method for detection of 5-methylcytosine by PCR-based genomic sequencing.
<3>Biotechniques
<4>16
<5>416-417
<6>1994
<7>
<>

<1>Reed, S.G., Lodes, M.J., Mohamath, R., Secrist, H.
<2>Compounds for therapy and diagnosis of lung cancer and methods for their use.
<3>Japanese Patent Office
<4>JP 2002540790 A
<5>
<6>2002
<7>
<>

<1>Reed, S.G., Skeiky, Y.A.W., Dillon, D.C.
<2>Compositions and methods for the prevention and treatment of M. tuberculosis infection.
<3>US Patent Office
<4>US 6350456 B
<5>
<6>2002
<7>Compositions and methods for treatment and vaccination against tuberculosis are disclosed.  In
one aspect the compositions provided include at least two polypeptides that contain an
immunogenic portion of a M. tuberculosis antigen or at least two DNA molecules encoding such
polypeptides.  In a second aspect, the compositions provided include a fusion protein
comprising at least two polypeptides that contain an immunogenic portion of a M. tuberculosis
antigen.  Such compositions may be formulated into vaccines and/or pharmaceutical compositions
for immunization against M. tuberculosis infection, or may be used for the diagnosis of
tuberculosis.

<>

<1>Reed, S.G., Skeiky, Y.A.W., Dillon, D.C., Campos-Neto, A., Houghton, R., Vedvick, T.S., Twardzik, D.R., Lodes, M.J., Hendrickson, R.C.
<2>Compounds and methods for immunotherapy and diagnosis of tuberculosis.
<3>US Patent Office
<4>US 6962710 A
<5>
<6>2005
<7>Compounds and methods for inducing protective immunity against tuberculosis are disclosed.
The compounds provided include polypeptides that contain at least one immunogenic portion of
one or more M. tuberculosis proteins and DNA molecules encoding such polypeptides.  Such
compounds may be formulated into vaccines and/or pharmaceutical compositions for immunization
against M. tuberculosis infection, or may be used for the diagnosis of tuberculosis.

<>

<1>Reed, S.G., Skeiky, Y.A.W., Dillon, D.C., Campos-Neto, A., Houghton, R., Vedvick, T.S., Twardzik, D.R., Lodes, M.J., Hendrickson, R.C.
<2>Compounds and methods for diagnosis of tuberculosis.
<3>US Patent Office
<4>US 6949246 A
<5>
<6>2005
<7>Compounds and methods for diagnosing tuberculosis are disclosed.  The compounds provided
include polypeptides that contain at least one antigenic portion of one or more M.
tuberculosis proteins, and DNA sequences encoding such polypeptides.  Diagnostic kits
containing such polypeptides or DNA sequences and a suitable detection reagent may be used for
the detection of M. tuberculosis infection in patients and biological samples.  Antibodies
directed against such polypeptides are also provided.

<>

<1>Reed, S.G., Skeiky, Y.A.W., Dillon, D.C., Campos-Neto, A., Houghton, R., Vedvick, T.S., Twardzik, D.R., Lodes, M.J., Hendrickson, R.C.
<2>Compounds and methods for diagnosis of tuberculosis.
<3>US Patent Office
<4>US 7122196 A
<5>
<6>2006
<7>Compounds and methods for diagnosing tuberculosis are disclosed.  The compounds provided
include polypeptides that contain at least one antigenic portion of one or more M.
tuberculosis proteins, and DNA sequences encoding such polypeptides.  Diagnostic kits
containing such polypeptides or DNA sequences and a suitable detection reagent may be used for
the detection of M. tuberculosis infection in patients and biological samples.  Antibodies
directed against such polypeptides are also provided.

<>

<1>Reed, S.G., Skeiky, Y.A.W., Dillon, D.C., Neto, A.C., Houghton, R., Vedvick, T.S., Twardzik, D.R., Lodes, M.J.
<2>Compounds and methods for diagnosis of Tuberculosis.
<3>Japanese Patent Office
<4>JP 2001500383
<5>
<6>2001
<7>
<>

<1>Rees, P.A., Nwankwo, D.O., Wilson, G.G., Benner, J.S.
<2>Cloning, purification and characterization of the HincII and HindII methyltransferases from Haemophilus influenzae.
<3>Gene
<4>74
<5>37
<6>1988
<7>Meeting Abstract

<>

<1>Reeve, J.N., Amann, E., Tailor, R., Gunthert, U., Scholz, K., Trautner, T.A.
<2>Unusual behaviour of SPO1 DNA with respect to restriction and modification enzymes recognizing the sequence 5'-G-G-C-C.
<3>Mol. Gen. Genet.
<4>178
<5>229-231
<6>1980
<7>SPO1 DNA contains only 5 cleavage sites for restriction enzymes which recognize
and cleave the sequence 5'-G-G-C-C (HaeIII or BsuR).  Fragments of SPO1 DNA
cloned in E. coli to substitute 5'-hydroxymethyluracil (HMU) by thymine (T)
remain resistant to HaeIII indicating that this unexpectedly small number of
cleavages by HaeIII is not correlated with the presence of HMU in the normal
phage DNA.  It was previously shown that SPO1 is neither subject to B. subtilis
R restriction (Trautner et al., 1974) nor modification in vivo (Gunthert et
al., 1975).  We now show that SPO1 DNA can however be restricted and modified
in vitro.

<>

<1>Reeve, W. et al.
<2>Complete genome sequence of Mesorhizobium australicum type strain (WSM2073(T)).
<3>Standards in Genomic Sciences
<4>9
<5>410-419
<6>2013
<7>Mesorhizobium australicum strain WSM2073(T) was isolated from root nodules on the pasture
legume Biserrula pelecinus growing in Australia in 2000. This aerobic,
motile, gram negative, non-spore-forming rod is poorly effective in N2 fixation
on B. pelecinus and has gained the ability to nodulate B. pelecinus following in
situ lateral transfer of a symbiosis island from the original inoculant strain
for this legume, Mesorhizobium ciceri bv. biserrulae WSM1271. We describe that
the genome size of M. australicum strain WSM2073(T) is 6,200,534 bp encoding
6,013 protein-coding genes and 67 RNA-only encoding genes. This genome does not
contain any plasmids but has a 455.7 kb genomic island from Mesorhizobium ciceri
bv. biserrulae WSM1271 that has been integrated into a phenylalanine-tRNA gene.

<>

<1>Reeve, W. et al.
<2>Genome sequence of the Lebeckia ambigua-nodulating 'Burkholderia sprentiae' strain WSM5005(T.).
<3>Standards in Genomic Sciences
<4>9
<5>385-394
<6>2013
<7>'Burkholderia sprentiae' strain WSM5005(T) is an aerobic, motile, Gram-negative,
non-spore-forming rod that was isolated in Australia from an effective N2-fixing
root nodule of Lebeckia ambigua collected in Klawer, Western Cape of South
Africa, in October 2007. Here we describe the features of 'Burkholderia
sprentiae' strain WSM5005(T), together with the genome sequence and its
annotation. The 7,761,063 bp high-quality-draft genome is arranged in 8 scaffolds
of 236 contigs, contains 7,147 protein-coding genes and 76 RNA-only encoding
genes, and is one of 20 rhizobial genomes sequenced as part of the DOE Joint
Genome Institute 2010 Community Sequencing Program.

<>

<1>Reeve, W. et al.
<2>Complete genome sequence of Mesorhizobium opportunistum type strain WSM2075(T.).
<3>Standards in Genomic Sciences
<4>9
<5>294-303
<6>2013
<7>Mesorhizobium opportunistum strain WSM2075(T) was isolated in Western Australia in 2000 from
root nodules of the pasture legume Biserrula pelecinus that had been
inoculated with M. ciceri bv. biserrulae WSM1271. WSM2075(T) is an aerobic,
motile, Gram negative, non-spore-forming rod that has gained the ability to
nodulate B. pelecinus but is completely ineffective in N2 fixation with this
host. This report reveals that the genome of M. opportunistum strain WSM2075(T)
contains a chromosome of size 6,884,444 bp, encoding 6,685 protein-coding genes
and 62 RNA-only encoding genes. The genome contains no plasmids, but does harbor
a 455.7 kb genomic island from Mesorhizobium ciceri bv. biserrulae WSM1271 that
has been integrated into a phenylalanine-tRNA gene.

<>

<1>Reeve, W. et al.
<2>Genome sequence of the Trifolium rueppellianum -nodulating Rhizobium leguminosarum bv. trifolii strain WSM2012.
<3>Standards in Genomic Sciences
<4>9
<5>283-293
<6>2013
<7>Rhizobium leguminosarum bv. trifolii WSM2012 (syn. MAR1468) is an aerobic, motile,
Gram-negative, non-spore-forming rod that was isolated from an
ineffective root nodule recovered from the roots of the annual clover Trifolium
rueppellianum Fresen growing in Ethiopia. WSM2012 has a narrow, specialized host
range for N2-fixation. Here we describe the features of R. leguminosarum bv.
trifolii strain WSM2012, together with genome sequence information and
annotation. The 7,180,565 bp high-quality-draft genome is arranged into 6
scaffolds of 68 contigs, contains 7,080 protein-coding genes and 86 RNA-only
encoding genes, and is one of 20 rhizobial genomes sequenced as part of the DOE
Joint Genome Institute 2010 Community Sequencing Program.

<>

<1>Reeve, W. et al.
<2>Genome sequence of the lupin-nodulating Bradyrhizobium sp. strain WSM1417.
<3>Standards in Genomic Sciences
<4>9
<5>273-282
<6>2013
<7>Bradyrhizobium sp. strain WSM1417 is an aerobic, motile, Gram-negative, non-spore-forming rod
that was isolated from an effective nitrogen (N2) fixing
root nodule of Lupinus sp. collected in Papudo, Chile, in 1995. However, this
microsymbiont is a poorly effective N2 fixer with the legume host Lupinus
angustifolius L.; a lupin species of considerable economic importance in both
Chile and Australia. The symbiosis formed with L. angustifolius produces less
than half of the dry matter achieved by the symbioses with commercial inoculant
strains such as Bradyrhizobium sp. strain WSM471. Therefore, WSM1417 is an
important candidate strain with which to investigate the genetics of effective N2
fixation in the lupin-bradyrhizobia symbioses. Here we describe the features of
Bradyrhizobium sp. strain WSM1417, together with genome sequence information and
annotation. The 8,048,963 bp high-quality-draft genome is arranged in a single
scaffold of 2 contigs, contains 7,695 protein-coding genes and 77 RNA-only
encoding genes, and is one of 20 rhizobial genomes sequenced as part of the DOE
Joint Genome Institute 2010 Community Sequencing Program.

<>

<1>Reeve, W. et al.
<2>Genome sequence of the South American clover-nodulating Rhizobium leguminosarum bv. trifolii strain WSM597.
<3>Standards in Genomic Sciences
<4>9
<5>264-272
<6>2013
<7>Rhizobium leguminosarum bv. trifolii strain WSM597 is an aerobic, motile, Gram-negative,
non-spore-forming rod isolated from a root nodule of the annual
clover Trifolium pallidum L. growing at Glencoe Research Station near Tacuarembo,
Uruguay. This strain is generally ineffective for nitrogen (N2) fixation with
clovers of Mediterranean, North American and African origin, but is effective on
the South American perennial clover T. polymorphum Poir. Here we describe the
features of R. leguminosarum bv. trifolii strain WSM597, together with genome
sequence information and annotation. The 7,634,384 bp high-quality-draft genome
is arranged in 2 scaffolds of 53 contigs, contains 7,394 protein-coding genes and
87 RNA-only encoding genes, and is one of 20 rhizobial genomes sequenced as part
of the DOE Joint Genome Institute 2010 Community Sequencing Program.

<>

<1>Reeve, W. et al.
<2>Genome sequence of the Ornithopus/Lupinus-nodulating Bradyrhizobium sp. strain WSM471.
<3>Standards in Genomic Sciences
<4>9
<5>254-263
<6>2013
<7>Bradyrhizobium sp. strain WSM471 is an aerobic, motile, Gram-negative, non-spore-forming rod
that was isolated from an effective nitrogen- (N2) fixing
root nodule formed on the annual legume Ornithopus pinnatus (Miller) Druce
growing at Oyster Harbour, Albany district, Western Australia in 1982. This
strain is in commercial production as an inoculant for Lupinus and Ornithopus.
Here we describe the features of Bradyrhizobium sp. strain WSM471, together with
genome sequence information and annotation. The 7,784,016 bp high-quality-draft
genome is arranged in 1 scaffold of 2 contigs, contains 7,372 protein-coding
genes and 58 RNA-only encoding genes, and is one of 20 rhizobial genomes
sequenced as part of the DOE Joint Genome Institute 2010 Community Sequencing
Program.

<>

<1>Reeve, W. et al.
<2>Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain TA1.
<3>Standards in Genomic Sciences
<4>9
<5>243-253
<6>2013
<7>Rhizobium leguminosarum bv. trifolii strain TA1 is an aerobic, motile, Gram-negative,
non-spore-forming rod that is an effective nitrogen fixing
microsymbiont on the perennial clovers originating from Europe and the
Mediterranean basin. TA1 however is ineffective with many annual and perennial
clovers originating from Africa and America. Here we describe the features of R.
leguminosarum bv. trifolii strain TA1, together with genome sequence information
and annotation. The 8,618,824 bp high-quality-draft genome is arranged in a 6
scaffold of 32 contigs, contains 8,493 protein-coding genes and 83 RNA-only
encoding genes, and is one of 20 rhizobial genomes sequenced as part of the DOE
Joint Genome Institute 2010 Community Sequencing Program.

<>

<1>Reeve, W. et al.
<2>Genome sequence of the Listia angolensis microsymbiont Microvirga lotononidis strain WSM3557(T.).
<3>Standards in Genomic Sciences
<4>9
<5>540-550
<6>2014
<7>Microvirga lotononidis is a recently described species of root-nodule bacteria that is an
effective nitrogen- (N2) fixing microsymbiont of the symbiotically
specific African legume Listia angolensis (Welw. ex Bak.) B.-E. van Wyk & Boatwr.
M. lotononidis possesses several properties that are unusual in root-nodule
bacteria, including pigmentation and the ability to grow at temperatures of up to
45 degrees C. Strain WSM3557(T) is an aerobic, motile, Gram-negative,
non-spore-forming rod isolated from a L. angolensis root nodule collected in
Chipata, Zambia in 1963. This is the first report of a complete genome sequence
for the genus Microvirga. Here we describe the features of Microvirga lotononidis
strain WSM3557(T), together with genome sequence information and annotation. The
7,082,538 high-quality-draft genome is arranged in 18 scaffolds of 104 contigs,
contains 6,956 protein-coding genes and 84 RNA-only encoding genes, and is one of
20 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010
Community Sequencing Program.

<>

<1>Reeve, W. et al.
<2>Complete genome sequence of Rhizobium leguminosarum bv trifolii strain WSM2304, an effective microsymbiont of the South American clover Trifolium polymorphum.
<3>Standards in Genomic Sciences
<4>2
<5>66-76
<6>2010
<7>Rhizobium leguminosarum bv trifolii is the effective nitrogen fixing microsymbiont of a
diverse range of annual and perennial Trifolium (clover)
species. Strain WSM2304 is an aerobic, motile, non-spore forming, Gram-negative
rod, isolated from Trifolium polymorphum in Uruguay in 1998. This microsymbiont
predominated in the perennial grasslands of Glencoe Research Station, in Uruguay,
to competitively nodulate its host, and fix atmospheric nitrogen. Here we
describe the basic features of WSM2304, together with the complete genome
sequence, and annotation. This is the first completed genome sequence for a
nitrogen fixing microsymbiont of a clover species from the American center of
origin. We reveal that its genome size is 6,872,702 bp encoding 6,643
protein-coding genes and 62 RNA only encoding genes. This multipartite genome was
found to contain 5 distinct replicons; a chromosome of size 4,537,948 bp and four
circular plasmids of size 1,266,105 bp, 501,946 bp, 308,747 bp and 257,956 bp.

<>

<1>Reeve, W. et al.
<2>Complete genome sequence of the Medicago microsymbiont Ensifer (Sinorhizobium) medicae strain WSM419.
<3>Standards in Genomic Sciences
<4>2
<5>77-86
<6>2010
<7>Ensifer (Sinorhizobium) medicae is an effective nitrogen fixing microsymbiont of  a diverse
range of annual Medicago (medic) species. Strain WSM419 is an aerobic,
motile, non-spore forming, Gram-negative rod isolated from a M. murex root nodule
collected in Sardinia, Italy in 1981. WSM419 was manufactured commercially in
Australia as an inoculant for annual medics during 1985 to 1993 due to its
nitrogen fixation, saprophytic competence and acid tolerance properties. Here we
describe the basic features of this organism, together with the complete genome
sequence, and annotation. This is the first report of a complete genome sequence
for a microsymbiont of the group of annual medic species adapted to acid soils.
We reveal that its genome size is 6,817,576 bp encoding 6,518 protein-coding
genes and 81 RNA only encoding genes. The genome contains a chromosome of size
3,781,904 bp and 3 plasmids of size 1,570,951 bp, 1,245,408 bp and 219,313 bp.
The smallest plasmid is a feature unique to this medic microsymbiont.

<>

<1>Reeve, W. et al.
<2>Complete genome sequence of Rhizobium leguminosarum bv. trifolii strain WSM1325,  an effective microsymbiont of annual Mediterranean clovers.
<3>Standards in Genomic Sciences
<4>2
<5>347-356
<6>2010
<7>Rhizobium leguminosarum bv trifolii is a soil-inhabiting bacterium that has the capacity to be
an effective nitrogen fixing microsymbiont of a diverse range of
annual Trifolium (clover) species. Strain WSM1325 is an aerobic, motile,
non-spore forming, Gram-negative rod isolated from root nodules collected in 1993
from the Greek Island of Serifos. WSM1325 is produced commercially in Australia
as an inoculant for a broad range of annual clovers of Mediterranean origin due
to its superior attributes of saprophytic competence, nitrogen fixation and
acid-tolerance. Here we describe the basic features of this organism, together
with the complete genome sequence, and annotation. This is the first completed
genome sequence for a microsymbiont of annual clovers. We reveal that its genome
size is 7,418,122 bp encoding 7,232 protein-coding genes and 61 RNA-only encoding
genes. This multipartite genome contains 6 distinct replicons; a chromosome of
size 4,767,043 bp and 5 plasmids of size 828,924 bp, 660,973 bp, 516,088 bp,
350,312 bp and 294,782 bp.

<>

<1>Reeve, W., Ballard, R., Drew, E., Tian, R., Brau, L., Goodwin, L., Huntemann, M., Han, J., Tatiparthi, R., Chen, A., Mavrommatis, K., Markowitz, V., Palaniappan, K., Ivanova, N., Pati, A., Woyke, T., Kyrpides, N.
<2>Genome sequence of the Medicago-nodulating Ensifer meliloti commercial inoculant  strain RRI128.
<3>Standards in Genomic Sciences
<4>9
<5>602-613
<6>2014
<7>Ensifer meliloti strain RRI128 is an aerobic, motile, Gram-negative, non-spore-forming rod.
RRI128 was isolated from a nodule recovered from the roots
of barrel medic (Medicago truncatula) grown in the greenhouse and inoculated with
soil collected from Victoria, Australia. The strain is used in commercial
inoculants in Australia. RRI128 nodulates and forms an effective symbiosis with a
diverse range of lucerne cultivars (Medicago sativa) and several species of
annual medic (M. truncatula, Medicago littoralis and Medicago tornata), but forms
an ineffective symbiosis with Medicago polymorpha. Here we describe the features
of E. meliloti strain RRI128, together with genome sequence information and
annotation. The 6,900,273 bp draft genome is arranged into 156 scaffolds of 157
contigs, contains 6,683 protein-coding genes and 87 RNA-only encoding genes, and
is one of 100 rhizobial genomes sequenced as part of the DOE Joint Genome
Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria
(GEBA-RNB) project.

<>

<1>Reeve, W., Ballard, R., Howieson, J., Drew, E., Tian, R., Brau, L., Munk, C., Davenport, K., Chain, P., Goodwin, L., Pagani, I., Huntemann, M., Mavrommatis, K., Pati, A., Markowitz, V., Ivanova, N., Woyke, T., Kyrpides, N.
<2>Genome sequence of Ensifer medicae strain WSM1115; an acid-tolerant Medicago-nodulating microsymbiont from Samothraki, Greece.
<3>Standards in Genomic Sciences
<4>9
<5>514-526
<6>2014
<7>Ensifer medicae strain WSM1115 forms effective nitrogen fixing symbioses with a range of
annual Medicago species and is used in commercial inoculants in
Australia. WSM1115 is an aerobic, motile, Gram-negative, non-spore-forming rod.
It was isolated from a nodule recovered from the root of burr medic (Medicago
polymorpha) collected on the Greek Island of Samothraki. WSM1115 has a broad host
range for nodulation and N2 fixation capacity within the genus Medicago, although
this does not extend to all medic species. WSM1115 is considered saprophytically
competent in moderately acid soils (pH(CaCl2) 5.0), but it has failed to persist
at field sites where soil salinity exceeded 10 ECe (dS/m). Here we describe the
features of E. medicae strain WSM1115, together with genome sequence information
and its annotation. The 6,861,065 bp high-quality-draft genome is arranged into 7
scaffolds of 28 contigs, contains 6,789 protein-coding genes and 83 RNA-only
encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE
Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root
Nodule Bacteria (GEBA-RNB) project.

<>

<1>Reeve, W., Drew, E., Ballard, R., Melino, V., Tian, R., De Meyer, S., Brau, L., Ninawi, M., Daligault, H., Davenport, K., Erkkila, T., Goodwin, L., Gu, W., Munk, C., Teshima, H., Xu, Y., Chain, P., Kyrpides, N.
<2>Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain SRDI943.
<3>Standards in Genomic Sciences
<4>9
<5>232-242
<6>2013
<7>Rhizobium leguminosarum bv. trifolii SRDI943 (strain syn. V2-2) is an aerobic, motile,
Gram-negative, non-spore-forming rod that was isolated from a root nodule
of Trifolium michelianum Savi cv. Paradana that had been grown in soil collected
from a mixed pasture in Victoria, Australia. This isolate was found to have a
broad clover host range but was sub-optimal for nitrogen fixation with T.
subterraneum (fixing 20-54% of reference inoculant strain WSM1325) and was found
to be totally ineffective with the clover species T. polymorphum and T. pratense.
Here we describe the features of R. leguminosarum bv. trifolii strain SRDI943,
together with genome sequence information and annotation. The 7,412,387 bp
high-quality-draft genome is arranged into 5 scaffolds of 5 contigs, contains
7,317 protein-coding genes and 89 RNA-only encoding genes, and is one of 100
rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010
Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB)
project.

<>

<1>Reeve, W., Drew, E., Ballard, R., Melino, V., Tian, R., De Meyer, S., Brau, L., Ninawi, M., Teshima, H., Goodwin, L., Chain, P., Liolios, K., Pati, A., Mavromatis, K., Ivanova, N., Markowitz, V., Woyke, T., Kyrpides, N.
<2>Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain SRDI565.
<3>Standards in Genomic Sciences
<4>9
<5>220-231
<6>2013
<7>Rhizobium leguminosarum bv. trifolii SRDI565 (syn. N8-J) is an aerobic, motile, Gram-negative,
non-spore-forming rod. SRDI565 was isolated from a nodule
recovered from the roots of the annual clover Trifolium subterraneum subsp.
subterraneum grown in the greenhouse and inoculated with soil collected from New
South Wales, Australia. SRDI565 has a broad host range for nodulation within the
clover genus, however N2-fixation is sub-optimal with some Trifolium species and
ineffective with others. Here we describe the features of R. leguminosarum bv.
trifolii strain SRDI565, together with genome sequence information and
annotation. The 6,905,599 bp high-quality-draft genome is arranged into 7
scaffolds of 7 contigs, contains 6,750 protein-coding genes and 86 RNA-only
encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE
Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root
Nodule Bacteria (GEBA-RNB) project.

<>

<1>Reeve, W., Parker, M., Tian, R., Goodwin, L., Teshima, H., Tapia, R., Han, C., Han, J., Liolios, K., Huntemann, M., Pati, A., Woyke, T., Mavromatis, K., Markowitz, V., Ivanova, N., Kyrpides, N.
<2>Genome sequence of Microvirga lupini strain LUT6(T), a novel Lupinus alphaproteobacterial microsymbiont from Texas.
<3>Standards in Genomic Sciences
<4>9
<5>1159-1167
<6>2014
<7>Microvirga lupini LUT6(T) is an aerobic, non-motile, Gram-negative, non-spore-forming rod that
can exist as a soil saprophyte or as a legume
microsymbiont of Lupinus texensis. LUT6(T) was isolated in 2006 from a nodule
recovered from the roots of the annual L. texensis growing in Travis Co., Texas.
LUT6(T) forms a highly specific nitrogen-fixing symbiosis with endemic L.
texensis and no other Lupinus species can form an effective nitrogen-fixing
symbiosis with this isolate. Here we describe the features of M. lupini LUT6(T),
together with genome sequence information and its annotation. The 9,633,614 bp
improved high quality draft genome is arranged into 160 scaffolds of 1,366
contigs containing 10,864 protein-coding genes and 87 RNA-only encoding genes,
and is one of 20 rhizobial genomes sequenced as part of a DOE Joint Genome
Institute 2010 Community Sequencing Project.

<>

<1>Reeve, W., Tian, R., Brau, L., Goodwin, L., Munk, C., Detter, C., Tapia, R., Han, C., Liolios, K., Huntemann, M., Pati, A., Woyke, T., Mavrommatis, K., Markowitz, V., Ivanova, N., Kyrpides, N., Willems, A.
<2>Genome sequence of Ensifer arboris strain LMG 14919(T); a microsymbiont of the legume Prosopis chilensis growing in Kosti, Sudan.
<3>Standards in Genomic Sciences
<4>9
<5>473-483
<6>2014
<7>Ensifer arboris LMG 14919(T) is an aerobic, motile, Gram-negative, non-spore-forming rod that
can exist as a soil saprophyte or as a legume
microsymbiont of several species of legume trees. LMG 14919(T) was isolated in
1987 from a nodule recovered from the roots of the tree Prosopis chilensis
growing in Kosti, Sudan. LMG 14919(T) is highly effective at fixing nitrogen with
P. chilensis (Chilean mesquite) and Acacia senegal (gum Arabic tree or gum
acacia). LMG 14919(T) does not nodulate the tree Leucena leucocephala, nor the
herbaceous species Macroptilium atropurpureum, Trifolium pratense, Medicago
sativa, Lotus corniculatus and Galega orientalis. Here we describe the features
of E. arboris LMG 14919(T), together with genome sequence information and its
annotation. The 6,850,303 bp high-quality-draft genome is arranged into 7
scaffolds of 12 contigs containing 6,461 protein-coding genes and 84 RNA-only
encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE
Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root
Nodule Bacteria (GEBA-RNB) project.

<>

<1>Reeves, P.R., Liu, B., Zhou, Z., Li, D., Guo, D., Ren, Y., Clabots, C., Lan, R., Johnson, J.R., Wang, L.
<2>Rates of Mutation and Host Transmission for an Escherichia coli Clone over 3 Years.
<3>PLoS ONE
<4>6
<5>E26907
<6>2011
<7>Although over 50 complete Escherichia coli/Shigella genome sequences are
available, it is only for closely related strains, for example the O55:H7
and O157:H7 clones of E. coli, that we can assign differences to
individual evolutionary events along specific lineages. Here we sequence
the genomes of 14 isolates of a uropathogenic E. coli clone that persisted
for 3 years within a household, including a dog, causing a urinary tract
infection (UTI) in the dog after 2 years. The 20 mutations observed fit a
single tree that allows us to estimate the mutation rate to be about 1.1
per genome per year, with minimal evidence for adaptive change, including
in relation to the UTI episode. The host data also imply at least 6 host
transfer events over the 3 years, with 2 lineages present over much of
that period. To our knowledge, these are the first direct measurements for
a clone in a well-defined host community that includes rates of mutation
and host transmission. There is a concentration of non-synonymous
mutations associated with 2 transfers to the dog, suggesting some
selection pressure from the change of host. However, there are no changes
to which we can attribute the UTI event in the dog, which suggests that
this occurrence after 2 years of the clone being in the household may have
been due to chance, or some unknown change in the host or environment. The
ability of a UTI strain to persist for 2 years and also to transfer
readily within a household has implications for epidemiology, diagnosis,
and clinical intervention.

<>

<1>Rego, R.O.M., Bestor, A., Rosa, P.A.
<2>Defining the plasmid-encoded restriction-modification systems of the Lyme disease spirochete Borrelia burgdorferi.
<3>J. Bacteriol.
<4>193
<5>1161-1171
<6>2010
<7>The restriction-modification (R-M) systems of many bacteria present a barrier to the stable
introduction of foreign DNA. The Lyme disease spirochete Borrelia burgdorferi has two
plasmid-encoded putative R-M genes, bbe02 and bbq67, whose presence limits transformation by
shuttle vector DNA from E. coli. We show that both the bbe02 and bbq67 loci in recipient B.
burgdorferi limit transformation with shuttle vector DNA from E. coli, irrespective of its
dam, dcm, or hsd methylation status. However, plasmid DNA purified from B. burgdorferi
transformed naive B. burgdorferi much more efficiently than plasmid DNA from E. coli,
particularly when the bbe02 and bbq67 genotypes of the B. burgdorferi DNA source matched that
of the recipient. We detected adenine methylation of plasmid DNA prepared from B. burgdorferi
that carried bbe02 and bbq67. These results indicate that the bbe02 and bbq67 loci of B.
burgdorferi encode distinct R-M enzymes that methylate endogenous DNA and cleave foreign DNA
lacking the same sequence-specific modification. Our findings have basic implications for
horizontal gene transfer among B. burgdorferi strains with distinct plasmid contents. Further
characterization and identification of the nucleotide sequences recognized by BBE02 and BBQ67
will facilitate efficient genetic manipulation of this pathogenic spirochete.

<>

<1>Rehman, M.A., Ziebell, K., Nash, J.H., Kropinski, A.M., Ross, A., Al-Lami, M., Boerlin, P., Chui, L., Devenish, J., Bekal, S., Graham, M., Amoako, K.K., Johnson, R.P.
<2>Complete Genome Sequences of 16 Canadian Strains of Salmonella enterica subsp. enterica Serovar Enteritidis.
<3>Genome Announcements
<4>2
<5>e00330-14
<6>2014
<7>Salmonella enterica subsp. enterica serovar Enteritidis is an important zoonotic food-borne
pathogen causing serious human illnesses frequently linked to poultry products. Here, we
report fully assembled genome sequences of 16 S. Enteritidis strains with common pulsed-field
gel electrophoresis (PFGE) and phage types (8, 13, 13a, and 14b) that predominate in North
America.

<>

<1>Rehman, M.A., Ziebell, K., Nash, J.H., Kropinski, A.M., Zong, Z., Nafziger, E., Boerlin, P., Chui, L., Devenish, J., Bekal, S., Graham, M., Johnson, R.P.
<2>High-Quality Draft Whole-Genome Sequences of 162 Salmonella enterica subsp. enterica Serovar Enteritidis Strains Isolated from Diverse Sources in Canada.
<3>Genome Announcements
<4>2
<5>e00348-14
<6>2014
<7>We report the high-quality draft genome sequences of 162 strains of Salmonella enterica subsp.
enterica serovar Enteritidis representing diverse phage types and
pulsed-field gel electrophoresis (PFGE) profiles. The analysis of these genomes
will enable the identification of markers that are useful for differentiating
strains of this highly clonal serovar and will provide insights into the
evolution, virulence, and epidemiology of the strains.

<>

<1>Rehvathy, V., Tan, M.H., Gunaletchumy, S.P., Teh, X., Wang, S., Baybayan, P., Singh, S., Ashby, M., Kaakoush, N.O., Mitchell, H.M., Croft, L.J., Goh, K.L., Loke, M.F., Vadivelu, J.
<2>Multiple Genome Sequences of Helicobacter pylori Strains of Diverse Disease and Antibiotic Resistance Backgrounds from Malaysia.
<3>Genome Announcements
<4>1
<5>e00687-13
<6>2013
<7>Helicobacter pylori causes human gastroduodenal diseases, including chronic gastritis and
peptic ulcer disease. It is also a major microbial risk factor for
the development of gastric adenocarcinoma and mucosa-associated lymphoid tissue
(MALT) lymphoma. Twenty-one strains with different ethnicity, disease, and
antimicrobial susceptibility backgrounds were sequenced by use of Illumina HiSeq
and PacBio RS platforms.

<>

<1>Reich, N.O., Danzitz, M.J.
<2>Non-additivity of sequence-specific enzyme-DNA interactions in the EcoRI DNA methyltransferase.
<3>Nucleic Acids Res.
<4>19
<5>6587-6594
<6>1991
<7>We describe a novel strategy to characterize protein-DNA interactions involving
monomeric enzymes such as DNA methyltransferases (Mtases).  This strategy is
applied to our investigation of the EcoRI DNA Mtase, which binds its double
stranded recognition site 5'-G-AATTC-3' and methylates the central adenosine of
each strand using S-adenosyl-L-methionine as the methyl donor.  We show that
prior methylation of adenosine in either strand does not perturb catalysis.  In
contrast, substrates substituted with deoxyinosine at either guanosine position
(T-BM15 and T15-BM) show the minor groove residing N2 amino group of both
guanosines contribute to DNA recognition since specificity constants for the
modified substrates are reduced 13 and 39 fold.  Similar analysis of a
substrate containing deoxyinosine at both positions (T15-BM15) clearly shows
that some communication occurs between the sites.  To determine the extent to
which structural changes in the DNA alone contribute to this lack of
additivity, we performed DNA melting analysis of the singly and doubly
substituted substrates, and also found nonadditivity.  Although our functional
and structural analyses suggest that deoxyinosine incorporation causes long
range conformational effects, the similarity of Km(AdoMet) for all substrates
suggests that no large-scale structural changes occur in the Mtase-DNA-AdoMet
complex.  Our results support the following conclusions: 1) The non-additivity
shown in this system contrasts with the widespread demonstration of additivity
involving repressors (Lehming et al., 1990; Takeda et al., 1989; Ebright et
al., 1987), suggesting that sequence discrimination by enzymes may involve more
complex mechanisms.  Further, this non-additivity precludes quantitative
assignment of individual interactions and we suggest that future analyses of
this and related enzyme systems with base analogs include detailed information
about the long range structural consequences of individual substitutions.  2)
Although T15-BM and T-BM15 are shown to be radically different by thermodynamic
analysis, the similar specificity constants with the Mtase suggest that the
underlying structural differences (e.g., altered helical parameters of the DNA)
are not critical for sequence-recognition.  3) The significance of minor groove
Mtase-DNA interactions to specificity is confirmed.

<>

<1>Reich, N.O., Danzitz, M.J.
<2>High resolution mapping of the DNA-protein interface in the EcoRI DNA methylase system.
<3>ACS Abstracts
<4>194
<5>63
<6>1987
<7>This monomeric, S-Adenosyl-Methionine (SAM) dependent enzyme recognizes the
palindromic double stranded DNA sequence 5' GAATTC 3' and methylates the
exocyclic amino of the second adenine.  As part of our effort to understand the
origins of sequence specific DNA binding we are characterizing the interactions
between the methylase and its DNA substrate.  The experimental methods include
the use of small, well defined oligonucleotides in conjunction with DNase I and
hydroxy radical (PNAS (1986) 83 5469) mapping.  Results with altered flanking
DNA sequences, hemimethylated recognition sequences, and modulation of binding
via SAM (and SAM analogs) will be presented.

<>

<1>Reich, N.O., Danzitz, M.J.
<2>EcoRI DNA methyltransferase-DNA interactions.
<3>Biochemistry
<4>31
<5>1937-1945
<6>1992
<7>We present a novel strategy with synthetic hemimethylated DNA substrates
containing uracil for thymine and inosine for guanosine replacements and EcoRI
DNA methyltransferase to characterize the importance of major groove
hydrophobic groups to the sequence-specific modification of DNA.  The bacterial
MTase uses S-adenosyl-L-methionine to methylate the double-stranded DNA site
5'GAATTC3' at the N6 position of the central adenosine of each strand.  Uracil
substitution in either strand at the outer thymine (5'GAATUC3') causes 2.2- and
1.7-fold improvements in specificity (kcat/KmDNA).  The fact that the
specificity constant for the substrate containing uracil in both strands is
identifical to the value expected for noninteracting substitutions suggests
that no significant methyltransferase-DNA interactions are altered beyond the
site of etiher substitution.  Similar analysis of the internal thymine
(5'GAAUTC3') also shows these methyl groups to make a negative contribution to
specificity, although the observed non-additivity with the doubly modified
substrate clearly shows methyltransferase-DNA interactions beyond the site of
substitution to be affected in this case.  To further probe the effect of
analogue incorporation on methyltransferase-DNA interactions beyond the site of
substitution, the relatively silent and additive uracil changes (5'GAATUC3')
were combined with inosine for guanosine substitutions (e.g., 5'IAATTC3') known
to have significant negative effects on specificity.  In contrast to the
additivity observed with the outer thymines, these studies show significant
changes in methyltransferase-DNA interactions caused by the removal of the
thymine methyls.  Our results implicate a complex and flexible
methyltransferase-DNA interface in which subtle structural changes in the
substrate are transmitted over the entire canonical site.  Thermal stability
analyses and determination of Delta H and Delta S for the double- to
single-stranded transition for single and doubly substituted substrates show no
additivity.  This suggests that structural changes in the DNA alone may occur
beyond the site of substitution.  Interestingly, substrates with widely varying
enthalpy and entropy terms show similar specificity with the Mtase, suggesting
the Mtase is insensitive to the underlying conformational differences.

<>

<1>Reich, N.O., Danzitz, M.J., Osti, F.
<2>Mechanisms of substrate discrimination in the EcoRI DNA methylase.
<3>FASEB J.
<4>4
<5>A1793
<6>1990
<7>We have investigated how this prokaryotic Type II DNA methylase selectively
modifies the second adenine in the double stranded canonical site, 5' GAATTC
3'.  Non-selfcomplementary 14 basepair DNA substrates have been submitted to
detailed functional analysis by comparisons of true steady-state kinetic
parameters.  Modifications within the recognition hexanucleotide include  i)
methylation of one strand, ii) removal of single sites of (potential)
methylase-DNA interaction (e.g. minor groove H-bond donor, major groove
thymidine-methyl) iii) substitution of basepairs within the recognition site.
Prior methylation of one strand does not effect kinetic parameters (Kcat, Km or
Kcat/Km).  A significant contribution toward specificity (kcat/Km)DNA derives
from minor groove interactions; this is largely a result of increases in KmDNA.
In contrast, discrimination against substrates related by single basepair
changes occur through decreases in kcat of up to one million fold.  No effects
on KmAdoMet were detected for any DNA substrate.  The structural integrity of
the modified DNA substrates was determined with melting temperature and second
site analyses.

<>

<1>Reich, N.O., DiMichele, L.
<2>Functional analysis of cysteines in the EcoRI DNA methylase.
<3>ACS Abstracts
<4>194
<5>64
<6>1987
<7>This monomeric, S-adenosyl-methionine (SAM) dependent enzyme recognizes the
palindromic double stranded DNA sequence 5' GAATTC 3' and methylates the
exocyclic amino of the second adenine.  As part of our effort to understand the
origins of sequence specific DNA binding we are characterizing the functional
significance of the enzyme's seven cysteines (326 total amino acids).  Cysteine
labeling experiments with N-Ethyl-Maleimide (NEM) lead to inactivation of
enzyme activity.  Substrate and cofactor (SAM) protection are being used to
implicate specific cysteines.  Proteolytic digestion of 3H NEM labeled enzyme
followed by amino acid sequencing affords the assignment of specific
cysteine(s).

<>

<1>Reich, N.O., Everett, E.A.
<2>Identification of peptides involved in S-Adenosylmethionine binding in the EcoRI DNA methylase.
<3>J. Biol. Chem.
<4>265
<5>8929-8934
<6>1990
<7>The Mr 38,050 monomeric EcoRI DNA methylase is part of a bacterial
restriction-modification system.  The methylase transfers the methyl group from
S-adenosylmethionine (AdoMet) to the second adenine in the double-stranded DNA
sequence 5'-GAATTC-3'.  We have used the radiolabeled photoaffinity analog
8-azido-S-adenosylmethionine (8-N3-AdoMet) to identify peptides at the AdoMet
binding site in the binary methylase-cofactor analog complex.  The dissociation
constants in the absence of DNA for the analog and AdoMet are 12.9 and 4.8
microM, respectively.  The apparent kcat and Km values, obtained with the
double-stranded DNA substrate 5'-CGCGAATTCGCG-3', are 5.0 s-1 and 0.710 microM
(8-N3-AdoMet) and 4.3 s-1 and 0.335 microM (AdoMet).  Photolabeling by
8-N3-AdoMet occurs upon irradiation with ultraviolet light and is inhibited by
AdoMet.  Digestion of the adducted methylase with subtilisin generated several
radiolabeled peptides.  Peptide sequencing from independent photolabeling
experiments revealed two radiolabeled peptides containing amino acids 206-212
and 213-221.  Instability of the adducted peptides precluded assignment of
modified amino acids.

<>

<1>Reich, N.O., Flynn, J.
<2>Modulators of DNA cytosine-5 methyltransferase and methods for use thereof.
<3>US Patent Office
<4>US 07138384
<5>
<6>2006
<7>A synthetic oligonucleotide comprising a C-5 methylcytosine and which recognizes and binds an
allosteric site on DNA methyltransferase
thereby inhibiting DNA methyltransferase activity is disclosed. Also
disclosed is a composition comprising a synthetic oligonucleotide of
the invention. The composition is useful for inhibiting DNA
methyltransferase activity, thereby inhibiting the methylation of DNA.
The composition can be a pharmaceutical composition useful for treating
disorders associated with methylation defects, such as cancer and
certain developmental disorders. Also disclosed is a method of
inhibiting methylation of DNA. The method involves contacting a DCMTase
with a synthetic oligonucleotide of the invention in the presence of
the DNA, thereby resulting in an enzyme/synthetic oligonucleotide
complex. The presence of the complex prevents catalysis, thereby
inhibiting DNA methyltransferase activity. Also disclosed is a method
of treating a disorder of cell proliferation or development by
administering to a subject a synthetic oligonucleotide of the
invention. The inhibition of DNA methyltransferase prevents the
methylation of DNA thereby treating the disorder of cell proliferation
or development.

<>

<1>Reich, N.O., Maegley, K.A., Shoemaker, D.D., Everett, E.
<2>Structural and functional analysis of EcoRI DNA methyltransferase by proteolysis.
<3>Biochemistry
<4>30
<5>2940-2946
<6>1991
<7>Native EcoRI DNA methyltransferase (Mtase, Mr 38050) is proteolyzed by trypsin
to generate an intermediate 36-kDa fragment (p36) followed by the formation of
two polypeptides of Mr23000 and 13000 (p23 and p13, respectively).  Protein
sequence analysis of the tryptic fragments indicates that p36 results from
removal of the first 14 or 16 amino acids, p23 spans residues 15-216, and p13
spans residues 217-325.  The relative resistance to further degradation of p23
and p13 suggests stable domain structures.  This is further supported by the
generation of similar fragments with SV8 endoprotease which has entirely
different peptide specificities.  Our results suggest that Mtase is a
two-domain protein connected by a highly flexible interdomain hinge.  The
putative hinge region encompasses previously identified peptides implicated in
AdoMet binding [Reich, N.O. & Everett, E. (1990) J. Biol. Chem. 265, 8929-8934]
and catalysis [Everett et al. (1990) J. Biol. Chem. 265, 17713-17719].
Protection studies with DNA, S-adenosylmethionine (AdoMet),
S-adenosylhomocysteine (AdoHcy), and sinefungin (AdoMet analogue) show that the
Mtase undergoes significant conformational changes upon ligand binding.
Trypsinolysis of the AdoMet-bound form of the Mtase generates different
fragments, and the AdoMet-bound form is over 800 times more stable than unbound
Mtase.  The sequence-specific ternary complex (Mtase-DNA-sinefungin) is 2000
times more resistant to degradation by trypsin; cleavage eventually generates
26- and 12-kDa fragments which span residues 104-325 and 1-103, respectively
(p26 and p12).  The first 14 or 16 amino acids of the Mtase are not essential
since p36 retains activity.  Activity analysis of the p26 and p12 mixture also
indicates retention of activity.  Therefore, either p26 is catalytically active
or the two fragments remain associated to create a functional enzyme.  The
former rationale is supported by the fact that for EcoRI Mtase, all peptide
regions implicated in DNA binding, AdoMet binding, and catalysis residue in the
p26 fragment.

<>

<1>Reich, N.O., Mashhoon, N.
<2>Inhibition of EcoRI DNA methylase with cofactor analogs.
<3>J. Biol. Chem.
<4>265
<5>8966-8970
<6>1990
<7>Four analogs of the natural cofactor S-adenosylmethionine (AdoMet) were tested
for their ability to bind and inhibit the prokaryotic enzyme, EcoRI adenine DNA
methylase.  The EcoRI methylase transfers the methyl group from AdoMet to the
second adenine in the double-stranded DNA sequence 5' GAATTC 3'.  Dissociation
constants (KD) of the binary methylase-analog complexes obtained in the absence
of DNA with S-adenosylhomocysteine (AdoHcy), sinefungin, N-methyl-AdoMet, and
N-ethylAdoMet are 225, 43, >1000, and >2000 microM, respectively.  In the
presence of a DNA substrate, all four analogs show simple competitive
inhibition with respect to AdoMet.  The product of the enzymic reaction,
AdoHcy, is a poor inhibitor of the enzyme (KI(AdoHcy)=9 microM; KM(AdoMet)=0.60
microM).  Two synthetic analogs, N-methyl-AdoMet and N-ethyl-AdoMet, were also
shown to be poor inhibitors with KI values of 50 and >1000 microM,
respectively.  In contrast, the naturally occurring analog sinefungin was shown
to be a highly potent inhibitor (KI=10 microM).  Gel retardation assays confirm
that the methylase-DNA-sinefungin complex is sequence-specific.  The ternary
complex is the first sequence-specific complex detected for any DNA methylase.
Potential applications to structural studies of methylase-DNA interations are
discussed.

<>

<1>Reich, N.O., Mashhoon, N.
<2>Kinetic mechanism of the EcoRI DNA methyltransferase.
<3>Biochemistry
<4>30
<5>2933-2939
<6>1991
<7>We present kinetic analysis of the EcoRI DNA N6-adenosine methyltransferase
(Mtase).  The enzyme catalyzes the S-adenosylmethionine (AdoMet)-dependent
methylation of a short, synthetic 14 base pair DNA substrate and plasmid pBR322
DNA substrate with kcat/Km values of 0.51 x 10/8 and 4.1 x 10/8 S-1-M-1,
respectively.  The Mtase is thus one of the most efficient biocatalysts known.
Our data are consistent with an ordered bi-bi steady-state mechanism in which
AdoMet binds first, followed by DNA addition.  One of the reaction products,
S-adenosylhomocysteine (AdoHcy), is an uncompetitive inhibitor with respect to
DNA and a competititive inhibitor with respect to AdoMet.  Thus, initial DNA
binding followed by AdoHcy binding leads to formation of a ternary dead-end
complex (Mtase-DNA-AdoHcy).  We suggest that the product inhibition patterns
and apparent order of substrate binding can be reconciled by a mechanism in
which the Mtase binds AdoMet and noncanonical DNA randomly but that recognition
of the canonical site requires AdoMet to be bound.  Pre-steady-state and
isotope partitition analyses starting with the binary Mtase-AdoMet complex
confirm its catalytic competence.  Moreover, the methyl transfer step is at
least 10 times faster than catalytic turnover.

<>

<1>Reich, N.O., Mashhoon, N.
<2>Presteady state kinetics of an S-adenosylmethionine-dependent enzyme.
<3>J. Biol. Chem.
<4>268
<5>9191-9193
<6>1993
<7>We present the first presteady state kinetic analysis of an S-adenosylmethionine-dependent
enzyme. The target enzyme is the bacterial EcoRI DNA methyltransferase, which transfers the
methyl group to the second adenine in the DNA sequence GAATTC. The rate constant for
conversion of the central complex (enzyme-DNA-S-adenosylmethionine) to products
(enzyme-methylated DNA-S-adenosylhomocysteine) (41 +/- 7 s-1) is over 300-fold faster than
kcat, consistent with our demonstration that steps after methyltransfer are rate-limiting
(Reich, N.O., and Mashhoon, N. (1991) Biochemistry 30, 2933-2939). Methyltransfer at the N6
amino moiety of adenine on each strand requires a single binding orientation.

<>

<1>Reich, N.O., Mashhoon, N., Everett, E., Danzitz, M.
<2>Structure-function analysis of enzyme-DNA interactions in the EcoRI DNA methylase.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>213
<6>1989
<7>Our goal is to understand the molecular basis of DNA sequence recognition and
the modulation of this process by the cofactor S-adenosylmethionine, [AdoMet]
in this prokaryotic system.  The kinetic mechanism and the rate determining
step[s] have been elucidated: AdoMet binds first, followed by the double
stranded DNA substrate [5'GAATTC3'].  Rapid transfer of the methyl group to the
DNA in the ternary methylase-AdoMet-DNA complex is followed by rate limiting
step[s].  One portion of the methylase involved in AdoMet binding has been
identified:  it is a flexible peptide connecting two stable domains.  This
flexible hinge region and the domains were characterized using photo affinity
analogs, in vitro proteolysis, and peptide sequencing.  Hydroxy radical
footprinting and synthetic oligonucleotides have been used to characterize the
methylase-DNA topology.  Moreover, DNA substrates with modified bases [uracil,
inosine, deaza-adenine, etc.] have been submitted to comparative specificity
analysis [kcat/KM].  Data from both analyses will be presented; the
contribution to overall specificity deriving from individual methylase-DNA
interactions has been elucidated.

<>

<1>Reich, N.O., Olsen, C., Osti, F., Murphy, J.
<2>In vitro specificity of EcoRI DNA methyltransferase.
<3>J. Biol. Chem.
<4>267
<5>15802-15807
<6>1992
<7>The sequence selectivity of enzyme-DNA interactions was analyzed by comparing discrimination
between synthetic oligonucleotides containing the canonical site GAATTC and altered DNA
sequences with the EcoRI DNA methyltransferase. The specificities (kcat/Km DNA) are decreased
from 5- to 23,000-fold relative to the unmodified site. For several substrates the decrease in
kcat makes a disproportionate contribution to the specificity difference, suggesting that
discrimination is mediated by the placement of critical catalytic residues rather than binding
interaction. This is supported by our observation that specificity changes are generally not
followed by changes in the stability of the methyltransferase-DNA complexes. Also, base pair
substitution near the site of methylation results in greater decreases in complex stability,
suggesting that recognition and catalytic mechanisms overlap.

<>

<1>Reich, N.O., Olsen, C., Osti, F., Murphy, J., Smith, D., Crowder, S.
<2>In vitro and in vivo specificity analysis of the EcoRI DNA methyltransferase.
<3>FASEB J.
<4>6
<5>A217
<6>1992
<7>The EcoRI DNA methyltransferase modifies the second adenine in the site GAATTC
using the cofactor S-adenosylmethionine.  We determined the in vitro
sequence-selectivity with a family of related hemimethylated 14mers
X=methyladenosine):
5' GGCCGGAATTCGCGG 3'
3' CCGGCCTTXAGCGCC 5'
Comparisons of true kcat, KDNA, KmAdomet and kcat/KmDNA show specificity
decreases from 5(GAATCC) to 23,000 (GGATTC) fold (top strand shown).  For
several substrates the decrease in kcat makes a disproportionate contribution
toward specificity, suggesting that discrimination is mediated by the placement
of critical catalytic residues.  Further evidence for this conclusion is
provided by our observation that the stability of the methyltransferase-DNA
complexes (determined by gel shift analysis) do not generally follow
specificity changes.  In constrast, no methylation of noncanonical sites was
detectable in vivo.  We used three assays to show that TAATTC, CAATTC, GTATTC,
GGATTC and GAGTTC are not methylated in vivo under conditions where the
canonical site is protected.  A possible reconciliation of these results with
our in vitro data is that noncanonical methylation does occur in vivo but is
actively repaired.  The possible involvement of the recently identified mrr
(methylated adenine recognition and repair) locus in this capacity is being
investigated.

<>

<1>Reich, N.O., Olsen, C., Osti, F., Murphy, J., Smith, D., Crowder, S.
<2>In vitro and in vivo specificity analysis of the EcoRI DNA methyltransferase.
<3>Biochemistry
<4>31
<5>2208
<6>1992
<7>The EcoRI DNA methyltransferase modifies the second adenine in the site GAATTC
using the cofactor S-adenosylmethionine.  We determined the in vitro sequence
selectivity with a family of related hemimethylated 14mers (X =
methyladenosine):
5'GGCGGAATTCGCGG3'
3'CCGCCTTXAGCGCC5'
Comparisons of true kcat, KmDNA, KmAdoMet, and kcat/Km DNA show specificity
decreases from 5-(GAATCC) to 23000-(GGATTC) fold (top strand shown).  For
several substrates the decrease in kcat makes a disproportionate contribution
toward specificity, suggesting that discrimination is mediated by the placement
of critical catalytic residues.  Further evidence for this conclusion is
provided by our observation that the stability of the methyltransferase-DNA
complexes (determined by gel shift analysis) do not generally follow
specificity changes.  In contrast, no methylation of noncanonical sites was
detectable in vivo.  We used three assays to show that TAATTC, CAATTC, GTATTC,
GGATTC, and GAGTTC are not methylated in vivo under conditions where the
canonical site is protected.  A possible reconciliation of these results with
our in vitro data is that noncanonical methylation does occur in vivo but is
actively repaired.  The possible involvement of the recently identified mrr
(methylated adenine recognition and repair locus in this capacity is being
investigated.

<>

<1>Reich, N.O., Olsen, C., Osti, F., Murphy, J., Smith, D., Crowder, S.
<2>In vitro and in vivo specificity analysis of the EcoRI DNA methyltransferase.
<3>ACS Abstracts
<4>203
<5>113-BIOL
<6>1992
<7>The EcoRI DNA methyltransferase modifies the second adenine in the site GAATTC using the
cofactor S-adenosylmethionine. We determined the in vitro sequence-selectivity with a family
of related hemimethylated 14mers (X=methyladenosine):
5' GGCGGAATTCGCGG 3'
3' CCGCCTTXAGCGCC 5'
Comparisons of true kcat, KmDNA, KmAdoMet and Kcat/KmDNA show specificity decreases from 5
(GAATCC) to 23,000 (GGATTC) fold (top strand shown). For several substrates the decrease in
kcat makes a disproportionate contribution toward specificity, suggesting that discrimination
is mediated by the placement of critical catalytic residues. Further evidence for this
conclusion is provided by our observation that the stability of the methyltransferase-DNA
complexes (determined by gel shift analysis) do not generally follow specificity changes. In
contrast, no methylation of noncanonical sites was detectable in vivo. We used three assays to
show that TAATTC, CAATTC, GTATTC, GGATTC and GAGTTC are not methylated in vivo under
conditions where the canonical site is protected. A possible reconciliation of these results
with our in vitro data is that noncanonical methylation does occur in vivo but is actively
repaired. The possible involvement of the recently identified mrr (methylated adenine
recognition and repair) locus in this capacity is being investigated.

<>

<1>Reich, N.O., Olsen, C., Osti, F., Murphy, J., Smith, D., Crowder, S.
<2>In vitro and in vivo specificity analysis of the EcoRI DNA methyltransferase.
<3>Biophys. J.
<4>61
<5>A217
<6>1992
<7>The EcoRI DNA methyltransferase modifies the second adenine in the site GAATTC
using the cofactor S-adenosylmethionine.  We determined the in vitro
sequence-selectivity with a family of related hemimethylated 14mers
(X-methyladenosine): top 5GGCGGAATTCGCGG3/5CCGCGAXTTCCGCC3' bottom.
Comparisons of the true kcat, KDNA, KmAdoMet and kcat/KmDNA show specificity
decreases from 5(GAATCC) to 23,000 (GGATTC) fold (top strand shown).  For
several substrates the decrease in kcat makes a disproportionate contribution
toward specificity, suggesting that discrimination is mediated by the placement
of critical catalytic residues.  Further evidence for this conclusion is
provided by our observation that the stability of the methyltransferase-DNA
complexes (determined by gel shift analysis) do not generally follow
specificity changes.  In contrast, no methylation of noncanonical sites was
detectable in vivo.  We used three assays to show that TAATTC, CAATTC, GTATTC,
GGATTC and GAGTTC are not methylated in vivo under conditions where the
canonical site is protected.  A possible reconciliation of these results with
our in vitro data is that noncanonical methylation does occur in vivo but is
actively repaired.  The possible involvement of the recently identified mrr
(methylated adenine recognition and repair) locus in this capacity is being
investigated.

<>

<1>Reich, N.O., Svedruzic, Z., Flynn, J., Aubol, B.
<2>The enzymology of epigenetics: Mechanism and inhibition of mammalian DNA cytosine methyltransferase.
<3>Proc. Amer. Assoc. Cancer Res.
<4>41
<5>346-347
<6>2000
<7>The major DNA cytosine methyltransferase in mammals, Dnmt1, is potently inhibited by a novel
allosteric inhibitor.  The inhibitor decreases DNA methylation in mammalian cells.  Other
compounds that decrease cellular DNA methylation levels have been shown to reverse
tumorigenesis in animals.  Thus, our inhibitor is being tested in tumor cell lines as a new
anticancer therapeutic.  The expression of newly integrated viral DNA is often stopped by DNA
methylation.  "DNA methylation spreading" results from the "activation" of the enzyme by
proximal 5-methylcytosine groups adjacent to the target CpG that undergoes methylation.  This
occurs primarily in single-stranded DNA, the 5-methylcytosine can be anywhere from 3 to 27
nucleotides removed from the CpG, and the effect is due to an increased affinity of the enzyme
for its substrate.  Our results have implications for the potential methylation-dependent
silencing of genes during gene therapy.  Dnmt1 prefers hemimethylated substrates; this occurs
during the initial attack at the C6 position of the target cytosine, rather than the
subsequent methyl transfer step.  Dnmt1 catalyzes the exchange of the C5 hydrogen of cytosine,
but only in the presence of the natural cofactor AdoMet, or certain cofactor analogs.  Similar
results were reported for the related deamination reaction catalyzed by bacterial enzymes,
suggesting that exchange and demamination occur through similar reaction intermediates.  Our
results suggest that Dnmt1 may not be mutagenic under the conditions previously demonstrated
for the bacterial enzyme.

<>

<1>Reich, S.
<2>Reaction mechanism of type III restriction endonuclease EcoP151 and possible application in molecular diagnostics.
<3>Ph.D. Thesis, Germany
<4>
<5>
<6>2003
<7>EcoP15I is a multifunctional, hetero-oligomeric Type III restriction enzyme.  Type III
restriction enzymes are of general interest in medicine and functional genome analysis because
they cut DNA 25 bp downstream of their recognition site.  EcoP15I recognises the DNA sequence
5'-CAGCAG and needs two inverse oriented recognition sites for effective DNA cleavage.
According to the present translocation collision model DNA cleavage was proposed to result
from ATP dependent DNA translocation, which is expected to induce DNA loop formation, and
collision of two enzyme-DNA complexes.  Experiments show that EcoP15 moves rather in a
three-dimensional than in a 'sliding' process in search for its recognition site.
Huntington's disease is a progressive neurodegenerative disorder with autosomal-dominant
inheritance.  The disease is caused by a CAG trinucleotide repeat expansion located in the
first exon of the HD gene.  To diagnose the illness the exact determination of the number of
CAG repeats is necessary.  This study shows that the number of CAG repeats in the HD gene can
be determined by restriction of the DNA with the endonuclease EcoP15I and subsequent
high-resolution analysis of the restriciton fragment pattern using the ALPexpress DNA Analysis
System.  Here, for the first time DNA translocation by the Type III restriction enzyme EcoP15I
is demonstrated.  The postulated EcoP15-DNA loops are visualised using scanning force
microscopy.  This confirms the translocation-collision model for DNA cleavage by EcoP15.
Similarities and differences between the DNA cleavage processes of the Type III restriciton
enzyme EcoP15I and other restriction enzymes are discussed.

<>

<1>Reich, S., Gossl, D., Reuter, M., Rabe, J.P., Kruger, D.H.
<2>Scanning force microscopy of DNA translocation by the type III restriction enzyme EcoP15I.
<3>J. Mol. Biol.
<4>341
<5>337-343
<6>2004
<7>Type III restriction enzymes are multifunctional heterooligomeric enzymes that cleave DNA at a
fixed position downstream of a non-symmetric recognition site. For effective DNA cleavage
these restriction enzymes need the presence of two unmethylated, inversely oriented
recognition sites in the DNA molecule. DNA cleavage was proposed to result from ATP-dependent
DNA translocation, which is expected to induce DNA loop formation, and collision of two
enzyme-DNA complexes. We used scanning force microscopy to visualise the protein interaction
with linear DNA molecules containing two EcoP15I recognition sites in inverse orientation. In
the presence of the cofactors ATP and Mg2+, EcoP15I molecules were shown to bind specifically
to the recognition sites and to form DNA loop structures. One of the origins of the
protein-clipped DNA loops was shown to be located at an EcoP15I recognition site, the other
origin had an unspecific position in between the two EcoP15I recognition sites. The data
demonstrate for the first time DNA translocation by the Type III restriction enzyme EcoP15I
using scanning force microscopy. Moreover, our study revealed differences in the
DNA-translocation processes mediated by Type I and Type III restriction enzymes.

<>

<1>Reich, S., Mucke, M., Moncke-Buchner, E., Reuter, M., Kruger, D.H.
<2>DNA cleavage by the restriction endonuclease EcoP15I.
<3>Biochem. Soc. Trans.
<4>28
<5>A331
<6>2000
<7>The type III restriction endonuclease EcoP15I recognizes the asymmetric DNA sequence
5'-CAGCAG and cleaves it 25-27 bp downstream.  Two unmethylated, inversely oriented
recognition sites in head-to-head configuration (5'-CAGCAG...NNN...CTGCTG) are preferred
substrates in DNA cleavage.  An intrinsic ATPase activity is the potential driving force of
DNA translocation bringing two EcoP15I-DNA complexes together.  In this work we investigated
EcoP15I cleavage efficiency in dependence on distance of two inversely oriented recognition
sites.  We showed that EcoP15I proceeds to cleave DNA efficiently even in the case of two
adjacent head-to-head oriented recognition sites, but cleavage rate rapidly diminished with
increasing distance of two tail-to-tail oriented recognition sites.  EcoP15I cleavage was
abolished in the presence of non-hydrolysable ATP analogs instead of ATP, even on substrates
with recognition sites in close vicinity.  This confirmed a role of ATP hydrolysis for the
phosphodiester bond cleavage.  Furthermore, DNaseI footprint experiments were performed to get
insight into the spatial requirements of the enzyme on substrate DNA with one recognition site
and revealed a 36 base-footprint rather symmetrical in both strands.  It became evident that
the enzyme did not cover the region around the cleavage site.  Presence of ATP caused a change
in the footprint pattern.  Analyzing a DNA fragment with two head-to-head oriented recognition
sites, an additional region between the two cooperative cleavage sites was protected against
DNaseI digestion.  Our experimental data allow a refinement of the tracking-collision model
discussed for type III enzymes and give suggestions on the spatial organization of the
collision complex.

<>

<1>Reid, S.L., Parry, D., Liu, H.-H., Connolly, B.A.
<2>Binding and recognition of GATATC target sequences by the EcoRV restriction endonuclease: A study using fluorescent oligonucleotides and fluorescence polarization.
<3>Biochemistry
<4>40
<5>2484-2494
<6>2001
<7>Oligonucleotides labeled with hexachlorofluorescein (hex) have enabled the interaction of the
restriction endonuclease EcoRV with DNA to be
evaluated using fluorescence anisotropy. The sensitivity of hex allowed
measurements at oligonucleotide concentrations as low as 1 nM, enabling
KD values in the low nanomolar range to be measured. Both direct
titration, i.e., addition of increasing amounts of the endonuclease to
hex-labeled oligonucleotides, and displacement titration, i.e.,
addition of unlabeled oligonucleotide to preformed
hex-oligonucleotide/EcoRV endonuclease complexes, have been used for KD
determination. Displacement titration is the method of choice;
artifacts due to any direct interaction of the enzyme with the dye are
eliminated, and higher fluorescent-labeled oligonucleotide
concentrations may be used, improving signal-to-noise ratio. Using this
approach (with three different oligonucleotides) we found that the
EcoRV restriction endonuclease showed a preference of between 1.5 and
6.5 for its GATATC target sequence at pH 7.5 and 100 mM NaCl, when the
divalent cation Ca2+ is absent. As expected, both the presence of Ca2+
and a decrease in pH value stimulated the binding of specific sequences
but had much less effect on nonspecific ones.

<>

<1>Reik, W., Allen, N.D.
<2>Imprinting with and without methylation.
<3>Curr. Biol.
<4>4
<5>145-147
<6>1994
<7>Methyltransferase-deficient mice reveal that DNA methylation is required for the somatic-cell
maintenance of parental imprinting, which alters the expression of a gene according to the
parent from which it was inherited.

<>

<1>Reik, W., Kelsey, G., Walter, J.
<2>Dissecting de novo methylation.
<3>Nat. Genet.
<4>23
<5>380-382
<6>1999
<7>The methylation of DNA is fundamental to mammalian development.  This is vividly illustrated
by the fact that mouse embryos deficient in Dnmt1, a DNA methyltransferase that methylates
cytosine groups, die as a result of genome-wide demethylation.  Dnmt1 seems to be the main
enzyme responsible for maintaining methylation after each round of DNA replication.  But
studies of Dnmt1-deficient embryonic stem cells have revealed that other enzymes must exist to
methylate the genome de novo after the wave of global demethylation that occurs during early
embryonic development.  Okano et al. recently reported that murine Dnmt3a and Dnmt3b are the
long-sought mammalian de novo methyltransferases.  Their findings, as well as those of Xu et
al. and Hansen et al., also show that mutations of DNMT3B cause a disorder associated with
immunodeficiency, centromere instability and facial anomalies (ICF syndrome).  This disorder
is the only known human condition with constitutive abnormalities in DNA methylation.

<>

<1>Reik, W., Walter, J.
<2>Imprinting mechanisms in mammals.
<3>Current Opinion in Genetics
<4>8
<5>154-164
<6>1998
<7>Imprinting is a genetic mechanism that determines expression or repression of genes according
to their parental origin.  Some imprinted genes occur in clusters in the genome.  Recent work
using transgenic mice shows that multiple cis-acting sequences are needed for correct
imprinting.  Mutation analysis in a normal chromosomal context reveals the importance of
imprinting centers for regional establishment or maintenance of imprinting in a cluster.
Elements that contribute to the function of imprinting centers and regional propagation of the
imprints are CpG-rich differentially methylated regions (that during development retain
germline imposed methylation or demethylation), direct repeat clusters, and unusual RNA's
(antisense, non-translated etc.).  The interaction of these cis elements with transacting
factors such as methylase and chromatin factors establishes a hierarchical control system with
local and regional effects.

<>

<1>Reimundo, P., Rivas, A.J., Osorio, C.R., Mendez, J., Perez-Pascual, D., Navais, R., Gomez, E., Sotelo, M., Lemos, M.L., Guijarro, J.A.
<2>Application of suppressive subtractive hybridization to the identification of genetic differences between two Lactococcus garvieae strains showing distinct differences in virulence for rainbow trout and mouse.
<3>Microbiology
<4>157
<5>2106-2119
<6>2011
<7>Lactococcus garvieae is the causative microbial agent of lactococcosis, an important and
damaging fish disease in aquaculture. This bacterium has also been isolated from vegetables,
milk, cheese, meat and sausages, from cow and buffalo as a mastitis agent, and even from
humans, as an opportunistic infectious agent. In this work pathogenicity experiments were
performed in rainbow trout and mouse models with strains isolated from human (L. garvieae HF)
and rainbow trout (L. garvieae UNIUDO74; henceforth referred to as 074). The mean LD(50) value
in rainbow trout obtained for strain 074 was 2.1x10(2)+/-84 per fish. High doses of the
bacteria caused specific signs of disease as well as histological alterations in mice. In
contrast, strain HF did not prove to be pathogenic either for rainbow trout or for mice. Based
on these virulence differences, two suppressive subtractive hybridizations were carried out to
identify unique genetic sequences present in L. garvieae HF (SSHI) and L. garvieae 074
(SSHII). Differential dot-blot screening of the subtracted libraries allowed the
identification of 26 and 13 putative ORFs specific for L. garvieae HF and L. garvieae 074,
respectively. Additionally, a PCR-based screening of 12 of the 26 HF-specific putative ORFs
and the 13 074-specific ones was conducted to identify their presence/absence in 25 L.
garvieae strains isolated from different origins and geographical areas. This study
demonstrates the existence of genetic heterogeneity within L. garvieae isolates and provides a
more complete picture of the genetic background of this bacterium.

<>

<1>Rein, T., DePamphilis, M.L., Zorbas, H.
<2>Identifying 5-methylcytosine and related modifications in DNA genomes.
<3>Nucleic Acids Res.
<4>26
<5>2255-2264
<6>1998
<7>Intense interest in the biological roles of DNA methylation, particularly in eukaryotes, has
produced at least eight different methods for identifying 5-methylcytosine and related
modifications in DNA genomes.  However, the utility of each method depends not only on its
simplicity but on its specificity, resolution, sensitivity and potential artifacts.  Since
these parameters affect the interpretation of data, they should be considered in any
application.  Therefore, we have outlined the principles and applications of each method,
quantitatively evaluated their specificity, resolution and sensitivity, identified potential
artifacts and suggested solutions, and discussed a paradox in the distribution of m5C in
mammalian genomes that illustrates how methodological limitations can affect interpretation of
data.  Hopefully, the information and analysis provided here will guide new investigators
entering this exciting field.

<>

<1>Reinecke, S.N., Morgan, R.D.
<2>BfaI, a new MaeI isoschizomer from Bacteroides fragilis, recognizes the sequence 5' C^TAG 3'.
<3>Nucleic Acids Res.
<4>19
<5>1152
<6>1991
<7>A new type II restriction endonuclease, BfaI, has been isolated from
Bacteroides fragilis (NEB #668).  BfaI recognizes the four base sequence 5'
CTAG 3' and cleaves between the C and the T residues to produce a 2 base 5'
extension; C/TAG.

<>

<1>Reinhard, B.M., Sheikholeslami, S., Mastroianni, A., Alivisatos, A.P., Liphardt, J.
<2>Use of plasmon coupling to reveal the dynamics of DNA bending and cleavage by single EcoRV restriction enzymes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>2667-2672
<6>2007
<7>Pairs of Au nanoparticles have recently been proposed as "plasmon rulers" based on the
dependence of their light scattering on the interparticle distance. Preliminary work has
suggested that plasmon rulers can be used to measure and monitor dynamic distance changes over
the 1- to 100-nm length scale in biology. Here, we substantiate that plasmon rulers can be
used to measure dynamical biophysical processes by applying the ruler to a system that has
been investigated extensively by using ensemble kinetic measurements: the cleavage of DNA by
the restriction enzyme EcoRV. Temporal resolutions of up to 240 Hz were obtained, and the
end-to-end extension of up to 1,000 individual dsDNA enzyme substrates could be simultaneously
monitored for hours. The kinetic parameters extracted from our single-molecule cleavage
trajectories agree well with values obtained in bulk through other methods and confirm well
known features of the cleavage process, such as DNA bending before cleavage. Previously
unreported dynamical information is revealed as well, for instance, the degree of softening of
the DNA just before cleavage. The unlimited lifetime, high temporal resolution, and high
signal/noise ratio make the plasmon ruler a unique tool for studying macromolecular assemblies
and conformational changes at the single-molecule level.

<>

<1>Reinisch, K.M.
<2>The crystal structure of HaeIII methylase covalently complexed to DNA: An extrahelical cytosine and rearranged base pairing.
<3>Diss. Abstr.
<4>56
<5>3746B
<6>1996
<7>Many organisms expand the information content of their genome through enzymatic methylation of
cytosine residues.  The structure of M.HaeIII, a bacterial DNA (cytosine-5) methyltransferase
(DCMtase), bound covalently to DNA, is presented here.  The structure has been solved by X-ray
diffraction using the multiple isomorphous replacement method.  It has been refined to 2.8 A
with a crystallographic Rfree-value of 32.6% (R-value of 22.6%) and root mean square
deviations from ideal values of 0.012 Angstroms and 2.3 degrees for bond lengths and angles,
respectively.  In the M.HaeIII-DNA complex, the substrate cytosine is extruded from the DNA
helix and inserted into the active site of the enzyme, as was observed for another DCMtase,
M.HhaI.  The DNA is bound in a cleft between the two domains of the protein and is distorted
from the characteristic B-form conformation at its recognition sequence.  A comparison of
structures shows that the M.HaeIII complex differs from that of M.HhaI in that the remaining
bases in the M.HaeIII recognition sequence undergo an extensive rearrangement in their
pairing.  In this process, the bases are unstacked, and a gap 8 Angstroms long opens in the
DNA.

<>

<1>Reinisch, K.M., Chen, L., Verdine, G.L., Lipscomb, W.N.
<2>Crystallization and preliminary crystallographic anlaysis of a DNA (cytosine-5)-methyltransferase from Haemophilus aegyptius bound covalently to DNA.
<3>J. Mol. Biol.
<4>238
<5>626-629
<6>1994
<7>A DNA (cytosine)-5-methyltransferase from Haemophilus aegyptius (M.HaeIII), which catalyzes
methyl transfer from S-adenosyl-L-methionine to DNA, has been crystallized as a covalent
complex with a suicide oliogonucleotide substrate. Crystals of the co-complex were grown by
vapor diffusion with hanging droplets, using polyethylene glycol 3500 as the precipitant. The
crystals belong to the orthorhombic space group P212121; the unit cell parameters are a=57.6A,
b=108.0A, c=155.8A with two protein-DNA complexes in the asymmetric unit. Complete sets of
native and derivative data have been collected to 2.7A using a laboratory source.

<>

<1>Reinisch, K.M., Chen, L., Verdine, G.L., Lipscomb, W.N.
<2>The crystal structure of HaeIII methyltransferase covalently complexed to DNA: an extrahelical cytosine and rearranged base pairing.
<3>Cell
<4>82
<5>143-153
<6>1995
<7>Many organisms expand the information content of their genome through enzymatic methylation of
cytosine residues.  Here we report the 2.8 A crystal structure of a bacterial DNA
(cytosine-5)-methyltransferase (DCMtase), M. HaeIII, bound covalently to DNA.  In this
complex, the substrate cytosine is extruded from the DNA helix and inserted into the active
site of the enzyme, as has been observed for another DCMtase, M.HhaI.  The DNA is bound in a
cleft between the two domains of the protein and is distorted from the characteristic B-form
conformation at its recognition sequence.  A comparison of structures shows a variation in the
mode of DNA recognition: M.HaeIII differs from M.HhaI in that the remaining bases in its
recognition sequence undergo an extensive rearrangement in their pairing.  In this process,
the bases are unstacked, and a gap 8 A long opens in the DNA.

<>

<1>Reisenauer, A., Kahng, L.S., McCollum, S., Shapiro, L.
<2>Bacterial DNA Methylation: a Cell Cycle Regulator?
<3>J. Bacteriol.
<4>181
<5>5135-5139
<6>1999
<7>A MiniReview of the role of CcrM (M.CcrMI) and Dam (M.EcoDam) methylases in regulating various
cellular processes such as DNA replication, cell-cycle and others.

<>

<1>Reisenauer, A., Shapiro, L.
<2>DNA methylation affects the cell cycle transcription of the CtrA global regulator in Caulobacter.
<3>EMBO J.
<4>21
<5>4969-4977
<6>2002
<7>The Caulobacter chromosome changes progressively from the fully methylated to the
hemimethylated state during DNA replication. These changes in DNA methylation could signal
differential binding of regulatory proteins to activate or repress transcription. The gene
encoding CtrA, a key cell cycle regulatory protein, is transcribed from two promoters. The P1
promoter fires early in S phase and contains a GAnTC sequence that is recognized by the CcrM
DNA methyltransferase. Using analysis of CcrM mutant strains, transcriptional reporters
integrated at different sites on the chromosome, and a ctrA P1 mutant, we demonstrate that
transcription of the P1 promoter is repressed by DNA methylation. Moreover moving the native
ctrA gene to a position near the chromosomal terminus, which delays the conversion of the ctrA
promoter from the fully to the hemimethylated state until late in the cell cycle, inhibited
ctrA P1 transcription, and altered the time of accumulation of the CtrA protein and the size
distribution of swarmer cells. Together, these results show that CcrM-catalyzed methylation
adds another layer of control to the regulation of ctrA expression.

<>

<1>Reiser, J., Yuan, R.
<2>Purification and properties of the P15 specific restriction endonuclease from Escherichia coli.
<3>J. Biol. Chem.
<4>252
<5>451-456
<6>1977
<7>The specific restriction endonuclease of the Escherichia coli plasmid, P15, has
been purified to apparent homogeneity by a procedure that includes
DNA-cellulose chromatography as well as a new endonuclease assay.
Sedimentation on glycerol gradients showed two peaks of activity with values of
11.3 S and 15.7 S.  The highly purified enzyme requires ATP and Mg2+ for
activity and is stimulated by S-adenosylmethionine.  A methylase activity is
observed in the course of the endonucleolytic reaction which protects some of
the DNA sites from cleavage.

<>

<1>Reiser, J., Yuan, R.
<2>A new cleavage assay for restriction endonucleases.
<3>Experientia
<4>31
<5>745
<6>1975
<7>Four different assays have recently been used for restriction enzymes.  Both
sucrose gradient and gel electrophoresis assays do not give quantitative values
whereas transfection is very time consuming.  The standard filter-binding assay
is based on the formation of an enzyme-DNA complex which can then be trapped on
nitrocellulose filters.  However, the restriction endonucleases from E. coli
(P1) and E. coli (15) do not show such binding, and a new cleavage assay that
was quick and quantitative had to be developed.  The basis for the assay is the
observation of Saucier and Wang (Biochem. 12, 2755, 1973) that circular lambda
DNA binds to nitrocellulose filters under specified conditions, whereas linear
lambda DNA will pass through.  For the cleavage assay linear lambda DNA is
circularized to form hydrogen-bonded circles (Hershey circles).  The circles
are then exposed to the enzyme in a complete reaction mixture; the reaction is
stopped by SDS and filtered rapidly through nitrocellulose filters without any
subsequent washing.  The filters are dried and counted in a liquid
scintillation counter.  The assay is specific (as checked with modified DNA and
dependence on cofactors), linear and quantitative.  It has been used
successfully in the purification of the restriction endonucleases from E. coli
(P1) and E. coli (15).

<>

<1>Reith, M., Munholland, J.
<2>Complete nucleotide sequence of the Porphyra purpurea chloroplast genome.
<3>Plant Mol. Biol. Rep.
<4>13
<5>333-335
<6>1995
<7>The complete nucleotide sequence of the chloroplast genome of the red alga Porphyra purpurea
has been determined (accession number = U38804).  The circular genome is 191,028 bp in length
and encodes approximately 250 genes.

<>

<1>Reither, S., Li, F., Gowher, H., Jeltsch, A.
<2>Catalytic mechanism of DNA-(cytosine-C5)-methyltransferases revisited: covalent intermediate formation is not essential for methyl group transfer by the murine Dnmt3a enzyme.
<3>J. Mol. Biol.
<4>329
<5>675-684
<6>2003
<7>Co-transfections of reporter plasmids and plasmids encoding the catalytic domain of the murine
Dnmt3a DNA methyltransferase lead to inhibition of
reporter gene expression. As Dnmt3a mutants with C-->A and E-->A exchanges
in the conserved PCQ and ENV motifs in the catalytic center of the enzyme
also cause repression, we checked for their catalytic activity in vitro.
Surprisingly, the activity of the cysteine variant and of the
corresponding full-length Dnmt3a variant is only two to sixfold reduced
with respect to wild-type Dnmt3a. In contrast, enzyme variants carrying
E-->A, E-->D or E-->Q exchanges of the ENV glutamate are catalytically
almost inactive, demonstrating that this residue has a central function in
catalysis. Since the glutamic acid residue contacts the flipped base, its
main function could be to hold the target base at a position that supports
methyl group transfer. Whereas wild-type Dnmt3a and the ENV variants form
covalent complexes with 5-fluorocytidine modified DNA, the PCN variant
does not. Therefore, covalent complex formation is not essential in the
reaction mechanism of Dnmt3a. We propose that correct positioning of the
flipped base and the cofactor and binding to the transition state of
methyl group transfer are the most important roles of the Dnmt3a enzyme in
the catalytic cycle of methyl group transfer.

<>

<1>Remenant, B., Babujee, L., Lajus, A., Medigue, C., Prior, P., Allen, C.
<2>Sequencing of K60, Type Strain of the Major Plant Pathogen Ralstonia solanacearum.
<3>J. Bacteriol.
<4>194
<5>2742-2743
<6>2012
<7>Ralstonia solanacearum is a widespread and destructive plant pathogen. We present the genome
of the type strain, K60 (phylotype IIA, sequevar 7). Sequevar 7
strains cause ongoing tomato bacterial wilt outbreaks in the southeastern United
States. K60 generally resembles R. solanacearum CFBP2957, a Caribbean tomato
isolate, but has almost 360 unique genes.

<>

<1>Remenant, B., Coupat-Goutaland, B., Guidot, A., Cellier, G., Wicker, E., Allen, C., Fegan, M., Pruvost, O., Elbaz, M., Calteau, A., Salvignol, G., Mornico, D., Mangenot, S., Barbe, V., Medigue, C., Prior, P.
<2>Genomes of three tomato pathogens within the Ralstonia solanacearum species complex reveal significant evolutionary divergence.
<3>BMC Genomics
<4>11
<5>379
<6>2010
<7>ABSTRACT: BACKGROUND: The Ralstonia solanacearum species complex includes
thousands of strains pathogenic to an unusually wide range of plant
species. These globally dispersed and heterogeneous strains cause
bacterial wilt diseases, which have major socio-economic impacts.
Pathogenicity is an ancestral trait in R. solanacearum and strains with
high genetic variation can be subdivided into four phylotypes, correlating
to isolates from Asia (phylotype I), the Americas (phylotype IIA and IIB),
Africa (phylotype III) and Indonesia (phylotype IV). Comparison of genome
sequences strains representative of this phylogenetic diversity can help
determine which traits allow this bacterium to be such a pathogen of so
many different plant species and how the bacteria survive in many
different habitats. RESULTS: The genomes of three tomato bacterial wilt
pathogens, CFBP2957 (Phy. IIA), CMR15 (phy. III) and PSI07 (phy. IV) were
sequenced and manually annotated. These genomes were compared with those
of three previously sequenced R. solanacearum strains: GMI1000 (tomato,
phy. I), IPO1609 (potato, phy. IIB), and Molk2 (banana, phy. IIB). The
major genomic features (size, G+C content, number of genes) were conserved
across all of the six sequenced strains. Despite relatively high genetic
distances (calculated from average nucleotide identity) and many genomic
rearrangements, more than 60% of the genes of the megaplasmid and 70% of
those on the chromosome are syntenic. The three new genomic sequences
revealed the presence of several previously unknown traits, probably
acquired by horizontal transfers, within the genomes of R. solanacearum,
including a type IV secretion system, a rhi-type anti-mitotic toxin and
two small plasmids. Genes involved in virulence appear to be evolving at a
faster rate than the genome as a whole. CONCLUSIONS: Comparative analysis
of genome sequences and gene content confirmed the differentiation of R.
solanacearum species complex strains into four phylotypes. Genetic
distances between strains, in conjunction with CGH analysis of a larger
set of strains, revealed differences great enough to consider
reclassification of the R. solanacearum species complex into three
species. The data are still too fragmentary to link genomic classification
and phenotypes, but these new genome sequences identify a pan-genome more
representative of the diversity in the R. solanancearum species complex.

<>

<1>Remenant, B., de Cambiaire, J.C., Cellier, G., Jacobs, J.M., Mangenot, S., Barbe, V., Lajus, A., Vallenet, D., Medigue, C., Fegan, M., Allen, C., Prior, P.
<2>Phylotype IV strains of Ralstonia solanacearum, R. syzygii and the blood disease bacterium form a single genomic species despite their divergent life-styles.
<3>PLoS ONE
<4>6
<5>e24356
<6>2011
<7>
<>

<1>Remus-Emsermann, M.N., Kim, E.B., Marco, M.L., Tecon, R., Leveau, J.H.
<2>Draft Genome Sequence of the Phyllosphere Model Bacterium Pantoea agglomerans 299R.
<3>Genome Announcements
<4>1
<5>e00036-13
<6>2013
<7>Bacteria belonging to the genus are common colonizers of plant leaf surfaces. Here, we present
the draft genome sequence of 299R, a phyllosphere isolate that
has become a model strain for studying the ecology of plant leaf-associated
bacterial commensals.

<>

<1>Ren, G., Li, D., Zhang, Q., Ye, X.
<2>Soluble NotI gene-expressing engineered Escherichia coli strain, its construction method and application.
<3>Chinese Patent Office
<4>CN 101948795 A
<5>
<6>2011
<7>
<>

<1>Ren, L.S., Oreshkin, E.N., Grachyov, Y.P.
<2>Biosynthesis of restriction endonuclease SalGI and optimal conditions of its purification.
<3>Prikl. Biokhim. Mikrobiol.
<4>22
<5>736-741
<6>1986
<7>The effect of the growing phase of Streptomyces albus G. and the components of
the culture medium on the biosynthesis of restriction endonuclease SalGI was
studied.  The conditions of DNA sedimentation with streptomycin sulfate and
polyethylenimine and separation of proteins with ammonium sulfate were
investigated as well.  A maximal quantity of the enzyme was observed in the
cells in 18-20 h of cultivation in flasks on a shaker.  The optimal
concentrations of the medium components (g/l) were found by mathematical
methods of the experiment planning:  glucose - 19,0; peptone - 5.2;
K2HPO4.3H2O-5.0.  The optimal concentrations for DNA sedimentation with
streptomycin sulfate and polyethylenimine were found to be 1.8-2.0% and
0.2-0.3%, respectively.  The optimal concentration of ammonium sulfate for
protein separation is 70% of saturation.

<>

<1>Ren, S.-X. et al.
<2>Unique physiological and pathogenic features of Leptospira interrogans revealed by whole-genome sequencing.
<3>Nature
<4>422
<5>888
<6>2003
<7>Leptospirosis is a widely spread disease of global concern. Infection causes flu-like episodes
with frequent severe renal and hepatic damage,
such as haemorrhage and jaundice. In more severe cases, massive pulmonary
haemorrhages, including fatal sudden haemoptysis, can occur. Here we
report the complete genomic sequence of a representative virulent serovar
type strain (Lai) of Leptospira interrogans serogroup Icterohaemorrhagiae
consisting of a 4.33-megabase large chromosome and a 359-kilobase small
chromosome, with a total of 4,768 predicted genes. In terms of the genetic
determinants of physiological characteristics, the facultatively parasitic
L. interrogans differs extensively from two other strictly parasitic
pathogenic spirochaetes, Treponema pallidum and Borrelia burgdorferi,
although similarities exist in the genes that govern their unique
morphological features. A comprehensive analysis of the L. interrogans
genes for chemotaxis/motility and lipopolysaccharide synthesis provides a
basis for in-depth studies of virulence and pathogenesis. The discovery of
a series of genes possibly related to adhesion, invasion and the
haematological changes that characterize leptospirosis has provided clues
about how an environmental organism might evolve into an important human
pathogen.

<>

<1>Ren, W., Liu, G., Yin, J., Chen, S., Li, T., Kong, X., Peng, Y., Yin, Y., Hardwidge, P.R.
<2>Draft Genome Sequence of Enterotoxigenic Escherichia coli Strain W25K.
<3>Genome Announcements
<4>2
<5>e00593-14
<6>2014
<7>Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrheal disease in  humans and
newly weaned pigs. Here, we report the draft genome sequence of ETEC
strain W25K, which causes diarrhea in piglets.

<>

<1>Ren, Y., Ren, Y., Zhou, Z., Guo, X., Li, Y., Feng, L., Wang, L.
<2>Complete genome sequence of Enterobacter cloacae subsp. cloacae type strain ATCC 13047.
<3>J. Bacteriol.
<4>192
<5>2463-2464
<6>2010
<7>Enterobacter cloacae is an important nosocomial pathogen. Here, we report the completion of
the genome sequence of E. cloacae ATCC 13047, the type
strain of E. cloacae subsp. cloacae. Multiple sets of virulence
determinant and heavy-metal resistance genes have been found in the
genome. To the best of our knowledge, this is the first complete genome
sequence of the E. cloacae species.

<>

<1>Renbaum, P., Abrahamove, D., Fainsod, A., Wilson, G.G., Rottem, S., Razin, A.
<2>Cloning, characterization, and expression in Escherichia coli of the gene coding for the CpG DNA methylase from Spiroplasma sp. strain MQ1(M.SssI) .
<3>Nucleic Acids Res.
<4>18
<5>1145-1152
<6>1990
<7>We describe here the cloning, characterization and expression in E. coli of the
gene coding for a DNA methylase from Spiroplasma sp. strain MQ1 (M.SssI).  This
enzyme methylates completely and exclusively CpG sequences.  The Spiroplasma
gene was transcribed in E. coli using its own promoter.  Translation of the
entire message required the use of an opal suppressor, suggesting that UGA
triplets code for tryptophan in Spiroplasma.  Sequence analysis of the gene
revealed several UGA triplets, in a 1158 bp long open reading frame.  The
deduced amino acid sequence revealed in M.SssI all common domains
characteristic of bacterial cytosine DNA methylases.  The putative sequence
recognition domain of M.SssI showed no obvious similarities with that of the
mouse DNA methylase, in spite of their common sequence specificity.  The cloned
enzyme methylated exclusively CpG sequences both in vivo and in vitro.  In
contrast to the mammalian enzyme which is primarily a maintenance methylase,
M.SssI displayed de novo methylase activity, characteristic of prokaryotic
cytosine DNA methylases.

<>

<1>Renbaum, P., Razin, A.
<2>Mode of action of the Spiroplasma CpG methylase M.SssI.
<3>FEBS Lett.
<4>313
<5>243-247
<6>1992
<7>The cytosine DNA methylase from the wall-less prokaryote, Spiroplasma strain MQ1 (M.SssI)
methylates completely and exclusively CpG-containing sequences, thus showing sequence
specificity which is similar to that of mammalian DNA methylases. M.SssI is shown here to
methylate duplex DNA processively as judged by kinetic analysis of methylated intermediates.
The cytosine DNA methylases. M.HpaII and M.HhaI from other prokaryotic organisms, appear to
methylate in a non-processive manner or with a very low degree of processivity. The
Spiroplasma enzyme interacts with duplex DNA irrespective of the presence of CpG sequences in
the substrate DNA. The enzyme proceeds along a CpG-containing DNA substrate molecule
methylating one strand of DNA at a time.

<>

<1>Renbaum, P., Razin, A.
<2>Interaction of M.SssI and M.HhaI with single-base mismatched oligodeoxynucleotide duplexes.
<3>Gene
<4>157
<5>177-179
<6>1995
<7>As part of an attempt to elucidate the mode of interaction of the CpG methyltransferase M.SssI
with its substrate, we have prepared a series of double-stranded oligodeoxyribonucleotides
containing one mismatch in the CpG recognition site.  The mismatched duplexes were used to
analyze the binding capabilities and enzymatic activity of M.SssI and M.HhaI (recognizes
GCGC).  We demonstrate here that M.SssI binds specifically to substrates containing either a
C/A or G/T mismatch in the recognition sequence, i.e., 5'-GCGC/CACG-5' or 5'-GCGC/CGTG-5',
respectively.  The enzyme also shows significant enzymatic activity with these mismatched
substrates.  These results suggest that site recognition and methylation of M.SssI take place
on the same strand.  M.HhaI bound and methylated the C/A mismatch very efficiently, but
recognition of the G/T mismatch was scarcely detectable.

<>

<1>Renbaum, P., Razin, A.
<2>Footprint analysis of M.SssI and M.HhaI methyltransferases reveals extensive interactions with the substrate DNA backbone.
<3>J. Mol. Biol.
<4>248
<5>19-26
<6>1995
<7>The interactions of the CpG methyltransferases M.SssI and M.HhaI (GCGC) with substrate DNA
were investigated using three different footprinting techniques. The two structurally related
enzymes displayed similar specific and non-specific contacts with DNA while bound to their
target sequences. DNase I footprinting implicated a region of 18 to 21 base-pairs with which
these enzymes interact. Dimethylsulfate protection experiments mostly revealed specific base
interactions; each enzyme was shown to interact predominantly with bases at its recognition
site in the major groove. However, hydroxyl radical footprints demonstrated extensive
interactions with the sugar-phosphate backbone on both strands of the DNA substrate. Both
enzymes protected a 16 nucleotide region, in a staggered fashion, covering 9 to 10 nucleotides
on each strand. The protected regions, extending for almost a full turn of DNA on each strand,
were offset by 6 to 7 nucleotides in the 5' direction, placing both regions on the same face
of the double helix, bracketing the major groove. The results suggest that these
methyltransferases straddle the major groove from the backbone, but protrude into the groove
only to specifically interact with their recognition sites. The sequence-independent
interactions observed on the sugar-phosphate backbone may explain the ability of the enzymes
to recognize a small sequence, as well as their processive mode of action.

<>

<1>Rendulic, S., Jagtap, P., Rosinus, A., Eppinger, M., Baar, C., Lanz, C., Keller, H., Lambert, C., Evans, K.J., Goesmann, A., Meyer, F., Sockett, R.E., Schuster, S.C.
<2>A predator unmasked: life cycle of Bdellovibrio bacteriovorus from a genomic perspective.
<3>Science
<4>303
<5>689-692
<6>2004
<7>Predatory bacteria remain molecularly enigmatic, despite their presence in
many microbial communities. Here we report the complete genome of
Bdellovibrio bacteriovorus HD100, a predatory Gram-negative bacterium that
invades and consumes other Gram-negative bacteria. Its surprisingly large
genome shows no evidence of recent gene transfer from its prey. A plethora
of paralogous gene families coding for enzymes, such as hydrolases and
transporters, are used throughout the life cycle of B. bacteriovorus for
prey entry, prey killing, and the uptake of complex molecules.

<>

<1>Rensing, S.A. et al.
<2>The Physcomitrella genome reveals evolutionary insights into the conquest of land by plants.
<3>Science
<4>319
<5>64-69
<6>2008
<7>We report the draft genome sequence of the model moss Physcomitrella patens and compare its
features with those of flowering plants, from which it is separated
by more than 400 million years, and unicellular aquatic algae. This comparison
reveals genomic changes concomitant with the evolutionary movement to land,
including a general increase in gene family complexity; loss of genes associated
with aquatic environments (e.g., flagellar arms); acquisition of genes for
tolerating terrestrial stresses (e.g., variation in temperature and water
availability); and the development of the auxin and abscisic acid signaling
pathways for coordinating multicellular growth and dehydration response. The
Physcomitrella genome provides a resource for phylogenetic inferences about gene
function and for experimental analysis of plant processes through this plant's
unique facility for reverse genetics.

<>

<1>Renvoise, A., Pang, S., Bernard, C., Brossier, F., Veziris, N., Capton, E., Jarlier, V., Sougakoff, W.
<2>First Whole-Genome Sequence of a Clinical Isolate of Multidrug-Resistant Mycobacterium bovis BCG.
<3>Genome Announcements
<4>2
<5>e00611-14
<6>2014
<7>The attenuated BCG strain of Mycobacterium bovis is widely used as a vaccine against
tuberculosis. However, in rare cases, it can be pathogenic to humans.
Here, we report the first draft of a whole-genome sequence of a
multidrug-resistant clinical isolate of M. bovis BCG.

<>

<1>Repik, A.V.
<2>Structural and functional organization of the genes of the EcoRII restriction-modification system.
<3>Ph.D. Thesis, Russian Academy of Sciences
<4>
<5>
<6>1994
<7>
<>

<1>Repin, V.E.
<2>New bacterium Bacillus coagulans, a producer of site-specific thermally stable endonuclease - thermostable restriction endonuclease production.
<3>Russian Patent Office
<4>RU 2040540
<5>
<6>1995
<7>
<>

<1>Repin, V.E.
<2>A strain of Bacillus stearothermophilus - a producer of restriction endonuclease able to recognize and split the 5'-CC(A/T)GG-3' nucleotide sequence.
<3>Russian Patent Office
<4>RU 2038380
<5>
<6>1995
<7>
<>

<1>Repin, V.E.
<2>A strain of Bacillus species able to recognize and cleave the nucleotide sequence 5'-(A/G)CCGG(T/C)-3'. Site-specific thermostable restriction endonuclease Bse118I production.
<3>Russian Patent Office
<4>RU 2037522
<5>
<6>1995
<7>
<>

<1>Repin, V.E., Andreeva, I.S., Kileva, E.V., Shevchenko, A.V., Abdurashitov, M.A., Repin, M.V., Degtyarev, S.K.
<2>MspR91, a new isoschizomer of ScrFI restriction endonuclease from Micrococcus species.
<3>Prikl. Biokhim. Mikrobiol.
<4>33
<5>284-286
<6>1997
<7>MspR91 restriction endonuclease was isolated from Micrococcus species identified by screening
soil bacteria for site-specific endonucleases.  The enzyme recognizes and cleaves the
palindromic sequence 5'-CC/NGG-3'.

<>

<1>Repin, V.E., Babkin, I.V., Tereshchenko, T.A.
<2>Bse118I, an isoschizomer of the Cfr10I restriction endonuclease from Bacillus coagulans.
<3>Bioorg. Khim.
<4>19
<5>406-409
<6>1993
<7>The recognition sequence and cleavage point for Bse118I restriction endonuclease have been
determined as (5') R^CCGGY.

<>

<1>Repin, V.E., Burtseva, L.I., Burlak, V.A., Trusova, S.I.
<2>Search of restriction endonuclease producers with required specificity.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>11
<5>20-22
<6>1991
<7>Six site-specific restriction endonucleases were isolated from Bacillus
thuringiensis strains chosen from 58 strains sought purposefully for the
production of restriction enzymes.  All six strains produce isoschizomers of
Sau3AI.

<>

<1>Repin, V.E., Chizhikov, V.E., Tereshchenko, T.A., Lebedev, L.R.
<2>Bco116I, An isoschizomer of Ksp632I from Bacillus coagulans.
<3>Bioorg. Khim.
<4>4
<5>410-413
<6>1993
<7>The recognition sequence and cleavage point of restriction endonuclease Bco116I have been
determined as (5') CTCTTC (1/4).

<>

<1>Repin, V.E., Degtyarev, S.K.
<2>Comparison of express-methods used for the detection of restriction endonucleases in microorganisms.
<3>Prikl. Biokhim. Mikrobiol.
<4>28
<5>152-155
<6>1992
<7>The sensitivity of several express-methods used for the detection of bacterial endonucleases
was compared. The most sensitive method is that employing Triton X-100.

<>

<1>Repin, V.E., Degtyarev, S.K.
<2>New non-pathogenic strain of Acinetobacter calcoaceticus VKPN V-6337 - useful as a producer of restriction endonuclease Acc65I.
<3>Russian Patent Office
<4>RU 2034922
<5>
<6>1995
<7>
<>

<1>Repin, V.E., Degtyarev, S.K., Petrov, N.A., Prihodko, E.A., Bendukidze, K.A., Fodor, I.I.
<2>Bacillus subtilis strain V-3667 - is used as a producer of the restriction endonuclease Bsu15I giving an increased yield.
<3>Soviet Patent Office
<4>SU 1449583 A
<5>
<6>1989
<7>The use of the strain of Bacillus subtilus deposited under the number V-3667 as producer of
restriction endonuclease Bsu15I, increases the process efficiency. The restrictase Bsu15I is
an isoschisomer of the restrictase ClaI. Bsu15I splits the nucleotide sequence 5'-ATCGAT-3'
in double-stranded DNA. The biomass is grown with aeration in bouillon based on sprat
hydrolysate until the growth ceases, and the cells are disintegrated with ultrasound.
Centrifuging and purification by chromatography completes the process.

<>

<1>Repin, V.E., Degtyarev, S.K., Rechkunova, N.I.
<2>A strain of bacterium Bacillus pumilis, a producer of restriction endonucleases BR119-1 - restriction endonuclease, e.g. Bpu19-1 preparation and purification.
<3>Russian Patent Office
<4>RU 2046142 C
<5>
<6>1995
<7>
<>

<1>Repin, V.E., Degtyarev, S.K., Rechkunova, N.I., Krivopalova, G.N., Vorodeva, T.I., Fish, A.M.
<2>Restriction endonuclease Vha464I production by Vibrio harveyi - application in cloning, mapping, etc.
<3>Russian Patent Office
<4>SU 1806192 A
<5>
<6>1993
<7>
<>

<1>Repin, V.E., Lebedev, L.R., Andreeva, I.S., Puchkova, L.I., Zernov, Y.P., Serov, G.D., Tereshchenko, T.A., Aphinogenova, G.N., Pustoshilova, N.M.
<2>The producers of restriction endonucleases from natural microbe isolates and the development on this basis of enzyme production technologies.
<3>Biotekhnologiya
<4>0
<5>18-27
<6>1998
<7>The screening of natural bacterial strains isolated from various ecological niches in order to
discover site specific nucleases has been carried out using different modifications of the
express method for detection of restriction endonucleases in bacterial colonies.  More than 60
easy to grow strains belonging to different bacterial taxa and promising for practical use
were obtained.  Some approaches to increase the strain efficiency as well as the purification
schemes to produce highly pure preparations of restriction endonuclease were suggested.

<>

<1>Repin, V.E., Lebedev, L.R., Puchkova, L., Serov, G.D., Tereschenko, T., Chizikov, V.E., Andreeva, I.
<2>New restriction endonucleases from thermophilic soil bacteria.
<3>Gene
<4>157
<5>321-322
<6>1995
<7>Using the most effective rapid method for the detection of restriction endonucleases (ENases)
in microorganisms, 32 thermophilic producers have been isolated.  All strains belong to the
genus Bacillus.  Thermostable isoschizomers of ENases, such as AvaI, BbvI, BbvII, BclI, BsaBI,
BsiYI, BsrI, BstEII, BstNI, Cfr10I, ClaI, FspI, HaeIII, HpaII, Ksp632I and SfeI, were
isolated.

<>

<1>Repin, V.E., Malup, T.K., Nikolaenkova, A.A., Rechkunova, N.I., Sviridov, S.M., Degtyarev, S.K.
<2>Culture medium for obtaining biomass of Streptomyces enriched in restriction endonuclease SfiI.
<3>Soviet Patent Office
<4>SU 1701745 A
<5>
<6>1991
<7>The present invention describes a method of fermentation of Streptomyces fimbriatus to produce
higher yields of the SfiI restriction endonuclease. Colonies of Streptomyces fimbriatus were
taken from cells plated on solid media and used to innoculate liquid media containing 3.5-3.7%
of hydrolyzed pilchard. The yield of cells was 18-20g/L culture; the yield of SfiI was 1,000
units/g cells.

<>

<1>Repin, V.E., Polshina, S.V., Bozhko, N.A., Degtyarev, S.K.
<2>The cultivation of Bacillus species 21, producer of restriction endonuclease Bse21I.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>15
<5>108-111
<6>1989
<7>The effect of the components of the culture medium, temperature of cultivation
and the growing phase of Bacillus species 21 on the yield of the restrictase
activity of this microorganism was studied.  A maximal quantity of the enzyme
per 1 g cells was observed at the beginning of the stationary phase of growth.
Using mathematical methods for planning experiments allowed us to raise the
Bse21I yield more than 8-fold.

<>

<1>Repin, V.E., Prichodko, E.A., Degtyarev, S.K.
<2>Search of soil microorganisms for production of restriction endonucleases.
<3>Izv. Sib. Otd. Akad. Nauk SSSR
<4>14
<5>110-113
<6>1988
<7>A search of soil microorganisms for the production of restriction endonucleases
was carried out.  Three strains belonging to species of Bacillus subtilis,
Bacillus megaterium and Aeromonas punctata were found and identified.  The
strains' restrictases were shown to be isoschizomers of ClaI and Sau3AI.  The
optimal conditions for cultivation of Bacillus subtilis 15 to increase enzyme
outcome were established.

<>

<1>Repin, V.E., Puchkova, L.I., Ananko, G.G., Reshetnikov, O.V., Kurilovich, S.A.
<2>HpyNI is the first restriction endonuclease isolated from Helicobacter pylori.
<3>Gut
<4>45
<5>A6
<6>1999
<7>Restriction-modification systems serve as a biological tool to ensure relative stability of
genetic material of the microbe, degrading alien DNA penetrated into bacterial cell.  The aim
of the study was to isolate and characterize restriction endonucleases of H. pylori.  H.
pylori strain was isolated from the gastric mucosa of duodenal ulcer patient.  The strain had
typical morphological and biochemical properties of H. pylori and was highly resistant to
metronidazole.  To study site-specificity viral and plasmid DNA's were used: T7, lambda, and
pBR322.  MsrR9I and Bst38I restrictases were utilized as standard.  The given strain of H.
pylori showed nuclease activity, and endonuclease obtained was called HpyNI according to the
generally accepted nomenclature.  The enzyme restricted standard substrate DNAs with the site
specific to base sequence 5'-CCNGG-3'.  Additional hydrolysis with Bst38I could not yield
novel fragments.  However, hydrolysis with HpyNI and MsrR9I showed complete homogeneity.  Thus
HpyNI proved to be an isoschizomer of ScrFI, in which this recognition site was first
described. Similar site specific endonucleases are widely represented in various microbial
taxons.  Horizontal transfer occurs between genomes of various bacteria, and restriction may
play a part in this process.  H. pylori is evolutionarily close to Escherichia coli and the
latter showed the greatest number of ScrFI isoschizomers, thus one could not exclude the
exchange of genes between these two bacteria.

<>

<1>Repin, V.E., Puchkova, L.I., Rodicheva, E.K., Vydryakova, G.A.
<2>Luminescent bacteria as producers of specific restriction endonucleases.
<3>Microbiology
<4>64
<5>751-755
<6>1995
<7>Luminescent bacteria isolated from the waters of the Black Sea and the Indian and Pacific
Oceans were tested for the presence of restriction endonucleases.  Restriction sites were
determined for 12 of the 19 restriction enzyme producers revealed.  The enzymes were
identified as isoschizomers of AflII, BanI, HaeIII, PstI, Sau3AI, and Sau96I.  Possible
participation of the restriction and modification enzymes in the interaction of bacterial
symbionts with the animal macrosymbiont is discussed.

<>

<1>Repin, V.E., Puchkova, L.I., Rodicheva, E.K., Vydryakova, G.A.
<2>Restriction endonucleases from symbiotic luminous bacteria.
<3>Prikl. Biokhim. Mikrobiol.
<4>33
<5>152-155
<6>1997
<7>Marine luminous bacteria from the Black Sea and the Indian and Pacific Oceans were assayed for
restriction endonucleases.  Nineteen producers of restriction endonucleases were found; the
recognition sites were determined for 13 identified restriction endonucleases.  The identified
enzymes were isoschizomers of AflII, BanI, HaeIII, NarI, PstI, Sau3AI, or Sau96I restriction
endonucleases.  The role of restriction-modification enzymes in the symbiotic relationships
between bacteria and their hosts was suggested.

<>

<1>Repin, V.E., Puchkova, L.I., Rodicheva, E.K., Vydryakova, G.A.
<2>Strain of bacterium Photobacterium phosphoreum, a producer of restriction endonuclease recognizing and splitting nucleotide sequence 5'-gg(t/c)(ag)cc-3'.
<3>Russian Patent Office
<4>RU 2110573 C
<5>
<6>1998
<7>
<>

<1>Repin, V.E., Puchkova, L.I., Ushakova, T.A., Mikhailova, V.K., Repina, M.V., Litosh, V.G.
<2>Strain of bacterium Deleya aquamarina, a producer of restriction endonuclease recognizing and cleaving nucleotide sequence 5'-GTGCAC-3'.
<3>Russian Patent Office
<4>RU 2105811 C
<5>
<6>1998
<7>
<>

<1>Repin, V.E., Rechkunova, N.I., Degtyarev, S.K., Hachaturyan, A.A., Afrikyan, E.K.
<2>Aerobic spore-forming bacteria as producers of restriction endonucleases.
<3>Biol. J. Armenia
<4>42
<5>969-972
<6>1989
<7>From 58 strains of aerobic sporeforming bacteria studied, including
their thermophilic forms, 5 strains have restrictase activity.  All
restrictases
produced by these strains are isoschizomers of known restrictases
characteristic
of bacilli.  For the forst time specific restrictase producers have been
indicated, which reveal the order of hexanucleotides.

<>

<1>Repin, V.E., Rechkunova, N.I., Tarasova, O.D., Degtyarev, S.K.
<2>Novel Moraxella species strain VKPM B-3678 - used in manufacturing site-specific endonuclease capable of recognising and splitting nucleotide(s).
<3>Soviet Patent Office
<4>SU 1514774 A
<5>
<6>1989
<7>Novel bacterial strain VKPM B-3678 of Moraxella species is used as producer of site-specific
endonuclease capable of recognising and splitting the sequence of nucleotides
5'-C(A/C)GC(G/T)G-3' is easily grown on the usual media and disintegrates under the action
of ultrasonics. The restriction endonuclease MspAI has site specificity identical to that of
NspBII and can replace it in all genetic engineering applications. A 1g portion of the biomass
yields 5000 units of the ferment with specific activity of 50000 units/ml.

<>

<1>Repin, V.E., Serov, G.D., Puchkova, L.I., Tereshenko, T.A., Lebedev, L.R., Chigikov, V.E.
<2>New restriction endonucleases from thermophilic bacteria of the genus Bacillus.
<3>Bioorg. Khim.
<4>19
<5>583-585
<6>1993
<7>Restriction endonucleases have been isolated from 26 thermophilic strains of the Bacillus
genus, their recognition sequences were determined, and for 15 of them cleavage sites
identified. The enzymes proved to be isoschizomers of known endonucleases BstNI, EarI, HaeIII,
HpaII, Cfr10I, BsiYI, BclI, BbvII, BbvI, BstEII, BsaBI, BsrI, FspI, ClaI, SfeI.

<>

<1>Repin, V.E., Serov, G.D., Shcheglova, I.K.
<2>New strain of Micrococcus varians VKPM B-5942 - useful for restriction endonuclease Mva161 production.
<3>Russian Patent Office
<4>RU 2018533
<5>
<6>1994
<7>
<>

<1>Repin, V.E., Shelkunov, S.N.
<2>Occurrence and function of restriction endonucleases (A Review).
<3>Usp. Sovrem. Biol.
<4>110
<5>34-47
<6>1990
<7>Modern nomenclature and classification of restriction endonucleases and related
topics are discussed.  Particular emphasis is placed on the distribution of
restriction enzymes among various taxons of microorganisms.  The possible
functional role of restriction enzymes in vivo and their evolution within
bacteria is discussed.  Basic methods for screening for type II enzymes are
described with a rationale for the use of rapid methods for analyzing large
numbers of bacterial strains.  Research perspectives in this branch of
molecular biology are presented.

<>

<1>Repin, V.E., Shilyayeva, O.H.
<2>Screening microorganisms isolated from air for restriction endonuclease producers.
<3>Microbiological investigations in Western Siberia, Nauka Siberian Division, Klevenskaya, I.L., Novosibirsk
<4>0
<5>14-17
<6>1989
<7>None

<>

<1>Repin, V.E., Tereshchenko, T.A., Lebedev, L.R.
<2>Strain of bacterium Bacillus coagulans - a producer of site-specific endonuclease which recognizes and splits the nucleotide sequence 5'-CTCTTC-3'.
<3>Russian Patent Office
<4>RU 2054040
<5>
<6>1996
<7>
<>

<1>Repin, V.E., Tereshchenko, T.A., Lebedev, L.R.
<2>A strain of the bacterium Bacillus coagulans, a producer of restriction endonuclease, recognizing the nucleotide sequence 5'-GATNNNNATC - Bco631I production.
<3>Russian Patent Office
<4>RU 2057806 C
<5>
<6>1996
<7>
<>

<1>Repin, V.E., Tereshchenko, T.A., Lebedev, L.R.
<2>A strain of Bacillus schlegelii manufacturing a restriction endonuclease recognizing and cleaving the nucleotide sequence 5'-CCNNNNNNNGG-3'.
<3>Russian Patent Office
<4>RU 2073717 C
<5>
<6>1997
<7>
<>

<1>Resch, G., Kulik, E.M., Dietrich, F.S., Meyer, E.
<2>Complete genomic nucleotide sequence of the temperate bacteriophage Aaphi23 of Actinobacillus actinomycetemcomitans.
<3>J. Bacteriol.
<4>186
<5>5523-5528
<6>2004
<7>The entire double-stranded DNA genome of the Actinobacillus actinomycetemcomitans
bacteriophage AaPhi23 was sequenced. Linear DNA
contained in the phage particles is circularly permuted and terminally
redundant. Therefore, the physical map of the phage genome is circular.
Its size is 43,033 bp with an overall molar G+C content of 42.5 mol%.
Sixty-six potential open reading frames (ORFs) were identified,
including an ORF resulting from a translational frameshift. A putative
function could be assigned to 23 of them. Twenty-three other ORFs share
homologies only with hypothetical proteins present in several bacteria
or bacteriophages, and 20 ORFs seem to be specific for phage AaPhi23.
The organization of the phage genome and several genetic functions
share extensive similarities to that of the lambdoid phages. However,
AaPhi23 encodes a DNA adenine methylase, and the DNA packaging strategy
is more closely related to the P22 system. The attachment sites of
AaPhi23 (attP) and several A. actinomycetemcomitans hosts (attB) are 49
bp long.

<>

<1>Reuter, M., Kruger, D.H., Scholz, D., Rosenthal, H.A.
<2>Protection of foreign DNA against host-controlled restriction in bacterial cells.  II. Protection of pSF2124 plasmid by the gene function of bacteriophages T3 and T7.
<3>Z. Allg. Mikrobiol.
<4>20
<5>345-354
<6>1980
<7>When restriction-active Escherichia coli cells (r+p1m+p1) are transformed
with the pSF2124 plasmid, a common vector in experimental gene transfer, the efficiency
of transformation (e.o.t.) is lowered by 2 orders of magnitude compared with restriction-
negative (r-p1m-p1 or r-p1m+p1) recipient cells due to restriction of the pSF2124 DNA by
endoR.EcoP1.  Preinfection of r+p1m+p1 recipient cells with pSF2124 attains the same
high value as that of r-m- cells.  The specific role of the ocr+ gene function was
demonstrated by the use of ocr- mutants (T3/R7, T7/D111): Preinfection with such phage
mutants does not increase the e.o.t. of r+p1m+p1 cells.  An unspecific e.o.t. alteration of
restriction-negative (r-m-) recipient cells by ocr+ or ocr- phages was excluded.  The ocr+
gene function can be exploited to protect pSF2124 against DNA restriction.  The recipient
cells survive the process of phage preinfection and transformation and stably replicate
themselves as well as the plasmid DNA.

<>

<1>Reuter, M., Kupper, D., Meisel, A., Schroeder, C., Kruger, D.H.
<2>Cooperative binding properties of restriction endonuclease EcoRII with DNA recognition sites.
<3>J. Biol. Chem.
<4>273
<5>8294-8300
<6>1998
<7>EcoRII is a member of the expanding group of type IIe restriction endonucleases that share the
distinguishing feature of requiring cooperativity between two recognition sites in their
substrate DNA.  To determine the stoichiometry of the active NDA-enzyme complex and the mode
of cooperative interaction, we have investigated the dependence of EcoRII cleavage on the
concentration of EcoRII dimers.  Maximal restriction was observed at dimer/site ratios of 0.25
and 0.5.  The molecular weight of the DNA-enzyme complex eluted from a gel filtration column
also corresponds to a dimeric enzyme structure bound to two substrate sites.  We conclude that
one EcoRII dimer is sufficient to interact cooperatively with two DNA recognition sites.  A
Lac repressor "barrier" bound between two normally reactive EcoRII sites did not inhibit
restriction endonuclease activity, indicating that cooperativity between EcoRII sites is
achieved by bending or looping of the intervening DNA stretch.  Comparative cleavage of linear
substrates with differently spaced interacting sites revealed an inverse correlation between
cleavage rate and site distance.  At the optimal distance of one helical turn, EcoRII cleavage
is independent of the orientation of the recognition sequence in the DNA double strand.

<>

<1>Reuter, M., Kupper, D., Pein, C.D., Petrusyte, M., Siksnys, V., Frey, B., Kruger, D.H.
<2>Use of specific oligonucleotide duplexes to stimulate cleavage of refractory DNA sites by restriction endonucleases.
<3>Anal. Biochem.
<4>209
<5>232-237
<6>1993
<7>There are numerous restriction endonuclease (ENases) which are known never to achieve total
cleavage of certain unmethylated target DNAs. In addition to EcoRII we found seven ENasese
(AtuBI, Cfr9I, Eco57I, Ksp6321, NaeI, NarI, and SauBMKI) that were stimulated by
oligodeoxyribonucleotide (oligo) duplexes containing enzyme-specific recognition sequences to
cut the target DNAs much more efficiently and in most cases even to completion. These enzymes
are class-II and class-IIS ENases isolated from different bacterial species and possess a
varying number of specific sites in the refractory DNA substrates. For DNA analysis and
large-scale preparation of certain restriction fragments where complete digestions are
essential we recommend taking into account the fact that various ENases can be activiated by
specific oligo duplexes to drive restriction digestions to completion.

<>

<1>Reuter, M., Muecke, M., Krueger, D.H.
<2>Structure and function of Type IIE restriction endonucleases or: From a plasmid that restricts phage replication to a new molecular DNA recognition mechanism.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>261-295
<6>2004
<7>Restriction endonucleases and DNA methyltransferases are encoded by chromosomal, plasmid, or
viral genes.  In eubacteria as well as in archaea, they often form biologically active DNA
restriction-modification systems.  The first R-M systems, discovered by reversible growth
reduction of bacterial viruses, were found to be encoded by chromosomal genes in Escherichia
coli K-12, Escherichia coli B, and Salmonella typhimurium.  Besides the prophage P1 and the
related plasmid p15 also naturally occurring drug resistance plasmids (R factors) of the fi-
(fertility inhibition-minus) type have been shown to restrict and modify infecting
bacteriophage or trans-conjugated plasmid DNAs in vivo.  The first R-factor controlled R-M
systems, called EcoRI and EcoRII today, have been defined by those phenotypical properties.
Endonucleolysis and cytosine methylation, respectively, have been proposed as the molecular
mechanisms of EcoRII-specific restriction and modification.  EcoRII was among the very first
R-M enzymes for which the DNA substrate site was identified.  The REase EcoRII recognizes the
sequence 5'-CC(A/T)GG which exhibits a twofold rotational symmetry with a dyad axis
corresponding to the central A-T pair.  EcoRII cleaves the phosphodiester bond at the 5' end
of the first cytosine of the unmethylated sequence.

<>

<1>Reuter, M., Pein, C.-D., Butkus, V., Kruger, D.H.
<2>An improved method for the detection of Dcm methylation in DNA molecules.
<3>Gene
<4>95
<5>161-162
<6>1990
<7>The intrinsic insensitivity of EcoRII recognition sites in RF DNAs of phage M13 and vector
M13mp18 towards this restriction endonuclease can be overcome by adding site-specific
oligodeoxyribonucleotide duplexes to the restriction sample. Since Dcm- DNA but not Dcm+
-methylated DNA becomes susceptible under these conditions, this procedure constitutes an
improvement of the Dcm methylation assay.

<>

<1>Reuter, M., Schneider-Mergener, J., Kupper, D., Meisel, A., Mackeldanz, P., Kruger, D.H., Schroeder, C.
<2>Regions of endonuclease EcoRII involved in DNA target recognition identified by membrane-bound peptide repertoires.
<3>J. Biol. Chem.
<4>274
<5>5213-5221
<6>1999
<7>Target sequence-specific DNA binding regions of the restriction endonuclease EcoRII were
identified by screening a membrane-bound EcoRII-derived peptide scan with an EcoRII
recognition site (CCWGG) oligonucleotide duplex.  Dodecapeptides overlapping by nine amino
acids and representing the complete protein were prepared by spot synthesis.  Two separate DNA
binding regions, amino acids 88-102 and amino acids 256-273, which share the consensus motif
KXRXXK, emerged.  Screening 570 single substitution analogues obtained by exchanging every
residue of both binding sites for all other amino acids demonstrated that replacing basic
residues in the consensus motifs significantly reduced DNA binding.  EcoRII mutant enzymes
generated by substituting alanine or glutamic acid for the consensus lysine residues in DNA
binding site I expressed attentuated DNA binding, whereas corresponding substitutions in DNA
binding site II caused impaired cleavage, but enzyme secondary structure was unaffected.
Furthermore, Glu96, which is part of a potential catalytic motif and also locates to DNA
binding site I, was demonstrated to be critical for DNA cleavage and binding.  Homology
studies of DNA binding site II revealed strong local homology to SsoII (recognition sequence,
CCNGG) and patterns of sequence conservation, suggesting the existence of functionally related
DNA binding sites in diverse restriction endonucleases with recognition sequences containing
terminal C:G or G:C pairs.

<>

<1>Reuter, V.M., Bogdarina, I.G., Kruger, D.H.
<2>Evidence for a specific inhibitor protein of DNA methyltransferases.
<3>Biol. Rundsch.
<4>24
<5>323-325
<6>1986
<7>The ocr protein encoded by the bacterial virus T7 is the first known inhibitor protein which
blocks DNA methyltransferases specifically. It inhibits the methyl transfer to DNA conferred
by the methylase M.EcoK, however, it is not active against the methylases M.dam and M.EcoRII.

<>

<1>Reva, O.M., Lukyanchuk, V.V., Polishchuk, L.V.
<2>Bsu5044I - Isoschisomer of endonuclease restriction AsuI.
<3>Mikrobiol. Zh.
<4>64
<5>57-59
<6>2002
<7>Endonuclease of restriction of type II has been found in Bacillus subtilis B-5044 during
testing of endophytic strains of cotton plants bacilli.  The restrictase Bsu5044I hydrolysed
DNA of M13mp18 phage in 4 sites; pUC19 DNA in 5 sites; pBR322 in 15 sites.  There were a lot
of sites restriction in DNAs of phages T7 and lambda.  A comparison of electrophoretical
divisions of Bsu5044I and Cfr13I fragments of phages and plasmid DNAs showed their identity.
Thus, the found restrictase Bsu5044I is an isoschisomer of AsuI.

<>

<1>Revel, H.R.
<2>DNA Modification:  Glucosylation.
<3>Bacteriophage T4, Amer. Soc. Microbiol., Mathews, C.K., Kutter, E.M., Mosig, G., Berget, P., Washington, DC
<4>0
<5>156-165
<6>1983
<7>None

<>

<1>Revel, H.R.
<2>Restriction of nonglucosylated T-even bacteriophage: properties of permissive mutants of Escherichia coli B and K12.
<3>Virology
<4>31
<5>688-701
<6>1967
<7>Mutants of Escherichia coli B and K12 which are permissive for either nonglucosylated
T6 alone or for all the nonglucosylated T-even bacteriophages have been isolated
after mutagenesis with nitrosoguanidine. The fully permissive mutants of B, derived
in a single step, require vitamin Bl, whereas those of K12, obtained in two steps, have
no novel nutritional requirements. Extracts of related permissive and reskicting
bacteria show no differences in the levels or specificities of their endonuclease I and
exonuclease III activities. Strain KlZ-1100, an endonuclease-deficient strain, restricts
all nonglucosylated T-even bacteriophages. Spheroplasts of restrict.ing bacteria
restrict ?r particles of the nonglucosylated T-even phages.

<>

<1>Revel, H.R., Georgopoulos, C.P.
<2>Restriction of nonglucosylated T-even bacteriophages by prophage P1.
<3>Virology
<4>39
<5>1-17
<6>1969
<7>Glucosyl transferase (gt) mutants of all the T-even bacteriophages, with less
than 3% of the normal complement of glucose on their DNA, fall into two groups
with respect to restriction by prophage P1.  Mutants of the first group, called
rp1, are restricted by P1 prophage irrespective of the nature of the hosts on
which the phage has grown; mutants of the second group, called rcp1, are
restricted by P1 only if they have grown on a permissive host lacking
UDPG-PPase.  Mutant prophages P1 r-m+ and P1 r-m-, which do not restrict
unmodified lambda phage, also fail to restrict gt rp1 or rcp1 phage.  The gt
mutants grown on P1 r-m+ lysogens are not modified.  Single-step mutants of gt
phage, called up1, which are insensitive to P1 restriction have been isolated
from T1gt rp1 and T4gt rp1, but not from T6gt rp1.  The up1 mutation does not
arise from loss or gain of a diffusible function and probably represents an
alteration in a site on the DNA recognized by the P1 restricting enzyme.  The
rp1 site maps outside the gt genes.  Both gt rp1 and gt up1 phages are
restricted by the resistance transfer factor N1.

<>

<1>Revel, H.R., Hattman, S.M.
<2>Mutants of T2gt with altered DNA methylase activity:  relation to restriction by prophage P1.
<3>Virology
<4>45
<5>484-495
<6>1971
<7>Glucosyl transferase (gt) mutants of T-even phages of a class called rp1 are restricted by
prophage P1. Unrestricted mutants up1, derived from T2gt rp1, have hypermethylated DNA. From
up1 one obtains in turn uRp1 mutants, which lack methyl groups on their DNA. Phage up1 is
dominant to either rp1 or uRp1 phage in mixed infections. The uRp1 mutations map close to and
on either side of the up1 mutation suggesting that both classes of mutation affect a single
gene. Analysis of DNA methylase activity in crude extracts of Sh infected with T2 or its
various gt derivatives reveals that the phage methylase induced by up1 phage differs in
specificity from the methylase of wild-type phage. The uRp1 phage fail to induce any methylase
activity.

<>

<1>Revel, H.R., Luria, S.E.
<2>DNA-glucosylation in T-even phage:  Genetic determination and role in phage-host interaction.
<3>Annu. Rev. Genet.
<4>4
<5>177-192
<6>1970
<7>Host-controlled modification phenomena.  The term "host-controlled
modification" covers a number of phenomena, first discovered in work on
bacteriophage:  a phage may become unable to infect successfully a given host
strain because of having previously grown in a different host.  Analysis of
this phenomenon in its most common form, especially with Escherichia coli and
phages lambda and fd, has revealed that these "restrictions" in host range are
due to changes in the DNA molecules that made them susceptible to a limited
endonucleolytic attack upon entering the "restricting" bacterial host.  The
attack occurs at specific "sites," presumably specific nucleotide sequences, in
the DNA molecules, and is accompanied in vivo by partial exonucleolytic
solubilization of the DNA fragments.  The term "modification" refers to the
fact that DNA produced in a given host strain is not restricted when it enters
cells of that same strain.  Modification consists of a specific methylation of
some base(s), adenine or cytosine, which renders the sites insensitive to the
specific restriction.  Because restriction can affect sites in the bacterial
DNA, it is not surprising that ability to restrict and ability to modify a
given site are usually associated in a given strain.  In fact, the restriction
and modification activities in a given E. coli strain are determined by a group
of three adjacent genes, r,m, and s, which together constitute the hs (host
specificity) system.  The r and m genes specify the restricting and modifying
functions.  The s gene determines the specificity of the phenomenon, presumably
by specifying a component common to a restricting and a modifying enzyme, which
recognizes the target sequence in the DNA: the m enzyme methylates the target
sequence if it is inside the cell; the r enzyme breaks it if it enters
unmethylated from the outside.  Both hs modification and restriction require
S-adenosyl methionine (SAM), as a methyl donor in modification  and also as a
cofactor in restriction.  Methylation of DNA generates 6-methyl-aminopurine
(6-MAP) from adenine residues and 5-methylcytosine (MC) from cytosine.  Mutants
r- do not restrict; mutants s- neither restrict nor modify.  Mutations m- are
found only together with r- mutations; it has been suggested that in an r+m-
bacterium there would be a suicidal attack on the bacterial DNA by the r+
function.  Besides its own hs system, which may be present or absent in a given
strain, a bacterium may exhibit other restriction systems due to episomes or
plasmids such as prophages or "resistance" factors.

<>

<1>Revez, J., Schott, T., Rossi, M., Hanninen, M.L.
<2>Complete Genome Sequence of a Variant of Campylobacter jejuni NCTC 11168.
<3>J. Bacteriol.
<4>194
<5>6298-6299
<6>2012
<7>Campylobacter jejuni NCTC 11168 is widely used in research, but at least two variants have
been reported. The available genome was sequenced from a variant
which later showed a different phenotype and gene expression profile. Here we
present the complete genome sequence of a second variant of C. jejuni NCTC 11168.

<>

<1>Rexer, B.U., Jarsch, M., Sagmeister, C., Gluck, B., Berger, G., Kessler, C.
<2>AsnI:  a novel class II restriction endonuclease from Arthrobacter sp., strain N-CM, recognizing 5'-AT/TAAT-3'.
<3>FEBS Lett.
<4>235
<5>241-246
<6>1988
<7>A new class II restriction endonuclease, AsnI, with a novel sequence
specificity was isolated from the Gram-positive eubacterium Arthrobacter
species, strain N-CM.  AsnI recognizes the unambiguously defined palindromic
hexanucleotide  5'-AT^TAAT-3' 3'-TAAT^TA-5' consisting of A- and T-residues.
The novel enzyme in the presence of Mg2+ cleaves specifically both strands as
indicated by the arrows.  The staggered cuts generate 5'-protruding ends with
single-stranded 5'-TA-3' dinucleotide extensions.  The novel enzyme may be a
useful tool for cloning experiments by complementation of the few enzymes such
as PstI and PvuI cutting only once in the Ampr-gene of plasmids pBR322 and
pBR328.

<>

<1>Rey, M.W. et al.
<2>Complete genome sequence of the industrial bacterium Bacillus licheniformis and comparisons with closely related Bacillus species.
<3>Genome Biology
<4>5
<5>R77
<6>2004
<7>Bacillus licheniformis is a Gram-positive, spore-forming soil
bacterium that is used in the biotechnology industry to manufacture
enzymes, antibiotics, biochemicals and consumer products. This species is
closely related to the well studied model organism Bacillus subtilis, and
produces an assortment of extracellular enzymes that may contribute to
nutrient cycling in nature.  We determined the complete nucleotide sequence
of the B. licheniformis ATCC 14580 genome which comprises a circular
chromosome of 4,222,336 base-pairs (bp) containing 4,208 predicted
protein-coding genes with an average size of 873 bp, seven rRNA operons,
and 72 tRNA genes. The B. licheniformis chromosome contains large regions
that are colinear with the genomes of B. subtilis and Bacillus halodurans,
and approximately 80% of the predicted B. licheniformis coding sequences
have B. subtilis orthologs.  Despite the unmistakable organizational
similarities between the B. licheniformis and B. subtilis genomes, there
are notable differences in the numbers and locations of prophages,
transposable elements and a number of extracellular enzymes and secondary
metabolic pathway operons that distinguish these species. Differences
include a region of more than 80 kilobases (kb) that comprises a cluster of
polyketide synthase genes and a second operon of 38 kb encoding plipastatin
synthase enzymes that are absent in the B. licheniformis genome. The
availability of a completed genome sequence for B. licheniformis should
facilitate the design and construction of improved industrial strains and
allow for comparative genomics and evolutionary studies within this group
of Bacillaceae.

<>

<1>Reysenbach, A.-L., Flores, G.E.
<2>Electron microscopy encounters with unusual thermophiles helps direct genomic analysis of Aciduliprofundum boonei.
<3>Geobiology
<4>6
<5>331-336
<6>2008
<7>Terry Beveridge's enthusiasm about the ingenuity of microorganisms has stimulated many new
avenues of microbial research. One example where Terry's observations helped direct the
scientific process was in the analysis of the draft genome of the thermoacidophilic archaeum,
Aciduliprofundum boonei. This deep-sea vent heterotroph ferments peptides as its primary
metabolic pathway, using numerous enzymes encoding for proteolytic or peptidolytic activities.
An almost complete modified Embden-Meyerhof-Parnas pathway operates in the gluconeogenic
direction. Terry was particularly intrigued by the S-layer and flagellum of A. boonei.
Although only putative genes for the S-layer protein could be identified, several genes
encoding for glycosyl transferases were located in the draft genome that could glycosylate the
S-layer proteins and protect the proteins from the acidic environment. Furthermore, A. boonei
possesses a unique organization to its flagellum genes and may represent a third
organizational type within the Archaea.

<>

<1>Reysenbach, A.L., Hamamura, N., Podar, M., Griffiths, E., Ferreira, S., Hochstein, R., Heidelberg, J., Johnson, J., Mead, D., Pohorille, A., Sarmiento, M., Schweighofer, K., Seshadri, R., Voytek, M.A.
<2>Complete and draft genome sequences of six members of the Aquificales.
<3>J. Bacteriol.
<4>191
<5>1992-1993
<6>2009
<7>The Aquificales are widespread in marine and terrestrial hydrothermal
environments. Here, we report the complete and draft genome sequences of
six new members of the Aquificales: two marine species, Persephonella
marina strain EX-H1 and Hydrogenivirga strain 128-5-R1 (from the East
Pacific Rise, 9 degrees 50.3'N, 104 degrees 17.5'W, and the Eastern Lau
Spreading Center, 176 degrees 11.5'W, 20 degrees 45.8'S, respectively),
and four terrestrial isolates, Sulfurihydrogenibium azorense strain
Az-Fu1, Sulfurihydrogenibium yellowstonense strain SS-5, and
Sulfurihydrogenibium strain Y03AOP1 (from Furnas, Azores, Portugal, and
Calcite Springs and Obsidian Pool in Yellowstone National Park, United
States, respectively), and the only thermoacidophilic isolate,
Hydrogenobaculum strain Y04AAS1 (from a stream adjacent to Obsidian Pool).
Significant differences among the different species exist that include
nitrogen metabolism, hydrogen utilization, chemotaxis, and signal
transduction, providing insights into their ecological niche adaptations.

<>

<1>Rhee, I., Bachman, K.E., Park, B.H., Jair, K.-W., Yen, R.-W.C., Schuebel, K.E., Cui, H., Feinberg, A.P., Lengauer, C., Kinzler, K.W., Baylin, S.B., Vogelstein, B.
<2>DNMT1 and DNMT3b cooperate to silence genes in human cancer cells.
<3>Nature
<4>416
<5>552-556
<6>2002
<7>Inactivation of tumour suppressor genes is central to the development of all common forms of
human cancer. This inactivation often results from epigenetic silencing associated with
hypermethylation rather than intragenic mutations. In human cells, the mechanisms underlying
locus-specific or global methylation patterns remain unclear. The prototypic DNA
methyltransferase, Dnmt1, accounts for most methylation in mouse cells, but human cancer cells
lacking DNMT1 retain significant genomic methylation and associated gene silencing. We
disrupted the human DNMT3b gene in a colorectal cancer cell line. This deletion reduced global
DNA methylation by less than 3%. Surprisingly, however, genetic disruption of both DNMT1 and
DNMT3b nearly eliminated methyltransferase activity, and reduced genomic DNA methylation by
greater than 95%. These marked changes resulted in demethylation of repeated sequences, loss
of insulin-like growth factor II (IGF2) imprinting, abrogation of silencing of the tumour
suppressor gene p16INK4a, and growth suppression. Here we demonstrate that two enzymes
cooperatively maintain DNA methylation and gene silencing in human cancer cells, and provide
compelling evidence that such methylation is essential for optimal neoplastic proliferation.

<>

<1>Rhee, I., Jair, K.W., Yen, R.W., Lengauer, C., Herman, J.G., Kinzler, K.W., Vogelstein, B., Baylin, S.B., Schuebel, K.E.
<2>CpG methylation is maintained in human cancer cells lacking DNMT1.
<3>Nature
<4>404
<5>1003-1007
<6>2000
<7>Hypermethylation is associated with the silencing of tumour susceptibility genes in several
forms of cancer; however, the mechanisms responsible for this aberrant methylation are poorly
understood. The prototypic DNA methyltransferase, DNMT1, has been widely assumed to be
responsible for most of the methylation of the human genome, including the abnormal
methylation found in cancers. To test this hypothesis, we disrupted the DNMT1 gene through
homologous recombination in human colorectal carcinoma cells. Here we show that cells lacking
DNMT1 exhibited markedly decreased cellular DNA methyltransferase activity, but there was only
a 20% decrease in overall genomic methylation. Although juxtacentromeric satellites became
significantly demethylated, most of the loci that we analysed, including the tumour suppressor
gene p16INK4a, remained fully methylated and silenced. These results indicate that DNMT1 has
an unsuspected degree of regional specificity in human cells and that methylating activities
other than DNMT1 can maintain the methylation of most of the genome.

<>

<1>Rhee, J.I., Bode, J., Diaz-Ricci, J.C., Poock, D., Weigel, B., Kretzmer, G., Schugerl, K.
<2>Influence of the medium composition and plasmid combination on the growth of recombinant Escherichia coli JM109 and on the production of the fusion protein EcoRI::SPA.
<3>J. Biotechnol.
<4>55
<5>69-83
<6>1997
<7>Plasmid-free and plasmid-harboring E. coli JM109 strains were investigated in shaken flasks,
stirred tanks in batch and continuous operation.  The shaken flask cultivations were performed
in M9 minimal medium and in media with various protein supplements.  The host hardly grows on
M9 minimal medium as opposed to the plasmid-harboring cells, which grow well on this medium.
All of the investigated cells propagate well on protein-containing media.  The influence of
the combinations of repressor plasmid pRK248cI, the protection plasmid EcoR4 and the
production plasmid pMTC48 were determined on the initial specific growth rate of the E. coli
JM109 without gene expression, on the yield coefficient of cell growth, acetate concentration
and acetate yield coefficient in the yeast extract-containing (HM) medium.  The influence of
various media on the induction of the gene expression were evaluated.  In cultivation media
with protein supplement, the growth rate and yield coefficient increased.  The variation of
the volumetric and specific beta-lactamase activities with the cultivation time were
determined in a stirred tank reactor in HM medium.  With increasing dilution rate the process
performance decreased.  Simple relationships exist between the substrate uptake rate and the
specific growth rate of the continuous cultivated cells in M9 and HM media.  The influence of
the dilution rate on the cell mass concentration, colony forming units, acetate formation,
yield coefficients of growth and acetate formation, substrate uptake rate, CO2 production
rate, ammonium formation rate and beta-lactamase activity in M9 and HM media were determined
as well.  Carbon balances of the batch and continuous cultivations indicated high carbon
recoveries.  On account of the higher growth rate of plasmid-harboring cells than that of the
plasmid-free cells, the behavior of the investigated plasmid-free and plasmid-harboring E.
coli JM109 cells deviates from the published properties of other plasmid-free and
plasmid-harboring E. coli cells.

<>

<1>Rhee, J.I., Schuegerl, K.
<2>Mathematical simulation of the growth of a three plasmid harboring Escherichia coli JM109 strain and the production of the fusion protein EcoRI::SPA with a four-compartment model.
<3>Bioprocess Eng.
<4>19
<5>261-267
<6>1998
<7>In the previous papers the process variables of plasmid-free, one-, two- and three-plasmid
harboring E. coli JM109 cells were investigated in batch and continuous cultivation as a
function of the medium composition, plasmid content, dilution rate and cultivation
(generation) time.  In the present paper the growth of the recombinant E. coli JM109 [pEcoR4,
pRK248cI, pMTC48] and the production of the fusion protein EcoRI::SPA are simulated by using a
four-compartment model, consisting of the active cell components (ribosomes, mRNA, tRNA, and
others) (A), the structure forming materials and chromosomal DNA (Z), the plasmid-DNA (G) and
the recombinant enzyme protein (E).  At the first time, all of the three plasmids: the
production plasmid (Gp), the repressor plasmid (Gr) and the protection plasmid (Gs) are taken
into account in the plasmid DNA-compartment of the model.  The calculated and measured courses
of the cell mass, the concentrations of glucose and acetate, and the products as well as the
particular plasmids agree well.

<>

<1>Rhee, M.S., Moritz, B.E., Xie, G., Glavina-del-Rio, T., Dalin, E., Tice, H., Bruce, D., Goodwin, L., Chertkov, O., Brettin, T., Han, C., Detter, C., Pitluck, S., Land, M.L., Patel, M., Ou, M., Harbrucker, R., Ingram, L.O., Shanmugam, K.T.
<2>Complete genome sequence of a thermotolerant sporogenic lactic acid bacterium, Bacillus coagulans strain 36D1.
<3>Standards in Genomic Sciences
<4>5
<5>331-340
<6>2011
<7>Bacillus coagulans is a ubiquitous soil bacterium that grows at 50-55 °C and pH 5.0 and
fer-ments various sugars that constitute plant biomass to L (+)-lactic acid. The ability of
this spo-rogenic lactic acid bacterium to grow at 50-55 °C and pH 5.0 makes this organism an
attrac-tive microbial biocatalyst for production of optically pure lactic acid at industrial
scale not only from glucose derived from cellulose but also from xylose, a major constituent
of hemi-cellulose. This bacterium is also considered as a potential probiotic. Complete genome
se-quence of a representative strain, B. coagulans strain 36D1, is presented and discussed.

<>

<1>Ri, S., Ra, S.R.
<2>Determination of cleavage sites of restriction endonuclease FmuI to plasmid pUB110 DNA.
<3>Choson Minjujuui Inmin Konghwaguk Kwahagwon Tongbo
<4>0
<5>50-53
<6>2002
<7>We cleave the plasmid pUB110 DNA with FmuI and some restriction endonucleases and map the
restriction sites according to the cleavage fragments.  The restriction endonuclease, FmuI
acts on 6 cleavage sites in the plasmid pUB110 DNA.

<>

<1>Riazuddin, S., Athar, A., Sohail, A., Ahmed, Z.
<2>Molecular mechanism of host controlled restriction and modification in Haemophilus influenzae 1.  Isolation of a restriction positive mutant.
<3>Pak. J. Zool.
<4>19
<5>393-405
<6>1987
<7>Wild type Haemophilus influenzae strain Rd has been mutagenized by acridine
orange treatment to produce a mutant MB69 which has cell division time,
transforming ability and radiosensitivity similar to that of the parent, but
does not allow normal growth of Haemophilus phage.  The mutant is sensitive to
adsorption of phage HP1c1 at similar levels as the parent strain.  However,
after entry into the host, phage HP1c1 DNA is specifically degraded into
discrete fragments of molecular sizes about one fifth of the parent phage DNA
estimated at 4.5 x 10/6 daltons.  Evidence is presented to support the
suggestion that the isolated mutant is restriction positive.

<>

<1>Riazuddin, S., Sohail, A., Maqbool, T., Khan, E., Mushtaq, R.
<2>Presence of new TypeII specificity restriction enzymes in local bacteria.
<3>Pak. J. Sci. Ind. Res.
<4>30
<5>819-824
<6>1987
<7>Eight bacterial strains isolated from local environments have been screened for
the presence of new Type II restriction enzymes by analyzing lambda DNA
fragments resulting from protein DNA interactions.  Partially purified protein
extracts of two of these strains, namely Pseudomonas aeruginosa A and
Citrobacter freundii A4 contain endonuclease activities which have been highly
purified by a combination of gel filtration, ion exchange and affinity
chromatography.  The identity of the new enzymes, designated as PaeAI and
CfrA4I, have been confirmed by analyses of incised lambda, PhiX174RFI, pBR322,
adeno-2 and M13 mp 19 RFI DNA substrates as truly Type II restriction enzymes.
The isolated enzymes, by analogy with known enzymes, seem to recognize CCGCGG
and CTGCAG base sequences respectively.

<>

<1>Ribeiro, F.J., Przybylski, D., Yin, S., Sharpe, T., Gnerre, S., Abouelleil, A., Berlin, A.M., Montmayeur, A., Shea, T.P., Walker, B.J., Young, S.K., Russ, C., Nusbaum, C., Maccallum, I., Jaffe, D.B.
<2>Finished bacterial genomes from shotgun sequence data.
<3>Genome Res.
<4>22
<5>2270-2277
<6>2012
<7>Exceptionally accurate genome reference sequences have proven to be of great
value to microbial researchers. Thus, to date, about 1800 bacterial genome
assemblies have been "finished" at great expense with the aid of manual
laboratory and computational processes that typically iterate over a period of
months or even years. By applying a new laboratory design and new assembly
algorithm to 16 samples, we demonstrate that assemblies exceeding finished
quality can be obtained from whole-genome shotgun data and automated computation.
Cost and time requirements are thus dramatically reduced.

<>

<1>Riccelli, P.V., Vallone, P.M., Kashin, I., Faldasz, B.D., Lane, M.J., Benight, A.S.
<2>Thermodynamic, spectroscopic, and equilibrium binding studies of DNA sequence context effects in six 22-base pair deoxyoligonucleotides.
<3>Biochemistry
<4>38
<5>11197-11208
<6>1999
<7>Effects of different end sequences on stability, circular dichroism spectra (CD), and enzyme
binding properties were investigated for six 22-base pair, non-self-complementary duplex DNA
oligomers. The center sequences of these deoxyoligonucleotides have 8-14 base pairs in common
and are flanked on both sides by sequences differing in context and A-T content.
Temperature-induced melting transitions monitored by differential scanning calorimetry (DSC)
and ultraviolet absorbance were measured for the six duplexes in buffered 115 mM Na(+)
solutions. Values of the melting transition enthalpy, DeltaH(cal), and entropy, DeltaS(cal),
were obtained directly from DSC experiments. Melting transition parameters, DeltaH(vH) and
DeltaS(vH), were also estimated from van't Hoff analysis of optical melting curves collected
as a function of DNA concentration, assuming a two-state melting transition. Melting free
energies (20 degrees C) of the six DNAs evaluated from DSC experiments ranged from -18.7 to
-32.7 kcal/mol. van't Hoff estimates of the free energies ranged from -18.5 to -48.0
kcal/mol. With either method, the trends in free energy as a function of sequence were
identical. Equilibrium binding by BamHI restriction endonuclease to the 22-base pair DNAs was
also investigated. The central eight base pairs of all six molecules, 5'-A-GGATCC-A-3',
contained a BamHI recognition sequence bounded by A-T base pairs. Magnesium free binding
assays were performed by titering BamHI against a constant concentration of each of the
deoxyoligonucleotide substrates and analyzing reaction products by gel retardation. Binding
isotherms of the total amount of bound DNA versus protein concentration were constructed which
provided semiquantitative estimates of the equilibrium dissociation constants for dissociation
of BamHI from the six DNA oligomers. Dissociation constants ranged from 0.5 x 10(-)(9) to 12.0
x 10(-)(9) M with corresponding binding free energies of -12.5 to -10.6 (+/-0. 1) kcal/mol. An
inverse relationship is found when binding and stability are compared.

<>

<1>Riccelli, P.V., Vallone, P.M., Lane, M.J., Benight, A.S.
<2>Investigations of DNA context effects: influences of flanking sequence stability on site specific binding of BamHI restriction enzyme to duplex DNA oligomers.
<3>J. Biophys.
<4>72
<5>A95
<6>1997
<7>Binding of BamHI restriction enzyme was investigated for short DNA duplex oligomer substrates
containing the cognate site 5'-GGATCC-3' flanked on both sides by sequences of different A-T
or G-C composition.  Binding reactions were conducted in buffer containing 10 mM CaCl2 and
analyzed by gel-shift assays.  While cleavage activity of the enzyme was eliminated under
these conditions, site specific binding was retained.  For each DNA substrate, binding
isotherms were constructed and equilibrium binding constants evaluated.  Binding constants
greater than 10^9 M-1 were observed and found to vary at least 10-fold as a function of
flanking sequence.  Significantly higher binding of BamHI was observed for duplex substrates
containing A-T flanking sequences.  Optical melting curves of the DNAs were also measured in
the binding buffer.  From these results, the thermodynamic stabilities of the DNA substrates
were evaluated.  Comparisons of the results of the binding assays with those of the melting
analysis revealed an inverse correlation between flanking sequence stability and binding
affinity of BamHI suggesting stability of flanking DNA context may comprise a significant
component of DNA recognition by site-specific binding agents.

<>

<1>Ricci, G., Ferrario, C., Borgo, F., Eraclio, G., Fortina, M.G.
<2>Genome Sequences of Two Lactococcus garvieae Strains Isolated from Meat.
<3>Genome Announcements
<4>1
<5>e00018-12
<6>2013
<7>Lactococcus garvieae is an important fish pathogen and an emerging opportunistic  human
pathogen, as well as a component of natural microbiota in dairy and meat products. We present
the first report of genome sequences of L. garvieae I113 and Tac2 strains isolated from a meat
source.

<>

<1>Ricci, G., Ferrario, C., Borgo, F., Rollando, A., Fortina, M.G.
<2>Genome Sequences of Lactococcus garvieae TB25, Isolated from Italian Cheese, and  Lactococcus garvieae LG9, Isolated from Italian Rainbow Trout.
<3>J. Bacteriol.
<4>194
<5>1249-1250
<6>2012
<7>Lactococcus garvieae is a fish pathogen and an emerging zoonotic opportunistic pathogen as
well as a component of natural microbiota in dairy products. Here, we
present the first report of a genome sequence of L. garvieae TB25, isolated from
a dairy source, and that of L. garvieae LG9, isolated from rainbow trout.

<>

<1>Rice, M.R., Blumenthal, R.M.
<2>Recognition of native DNA methylation by the PvuII restriction endonuclease.
<3>Nucleic Acids Res.
<4>28
<5>3143-3150
<6>2000
<7>Recognizing the methylation status of specific DNA sequences is central to the function of
many systems in eukaryotes and prokaryotes. Restriction-modification systems have to
distinguish between 'self' and 'non-self' DNA and depend on the inability of restriction
endonucleases to cleave their DNA substrates when the DNA is appropriately methylated. These
endonucleases thus provide a model system for studying the recognition of DNA methylation by
proteins. We have characterized the interaction of R.PvuII with DNA containing the
physiologically relevant N4-methylcytosine modification. R.PvuII binds (N4m)C-modified DNA
and cleaves it very slowly. Methylated strands in hemimethylated duplexes were cleaved at a
higher rate than in fully methylated duplexes, in parallel with a higher binding affinity for
hemimethylated DNA. The co-crystal structures of R.PvuII-DNA, together with a mutagenesis
study, have implicated specific amino acids in recognition of the methylatable base; one of
these is His84. We report that replacing His84 with Ala reduced the rate of cleavage of
unmodified DNA but, in contrast, slightly increased the cleavage of (N4m)C-modified DNA.

<>

<1>Rice, M.R., Calvin-Koons, M.D., Blumenthal, R.M.
<2>The PvuII (CAGCTG) endonuclease binds to and cleaves CAG5mCTG sites.
<3>FASEB J.
<4>9
<5>A1400
<6>1995
<7>The PvuII restriction modification system recognizes the sequence CAGCTG. Cleavage by the
restriction endonuclease (R.PvuII) occurs between the central bases, leaving blunt ends. The
PvuII methylase modifies the internal cytosine residue at the N4 position, but the sequence
CAG5mCTG has also been reported to be protected from cleavage by R.PvuII. Our evidence
suggests that cleavage of this sequence does in fact occur. This conclusion is based on the
following criteria. First, a cell expressing M.AluI, which generates AG5mCT, cannot be stably
transformed with R.PvuII, indicating that the protection provided by M.AluI is incomplete.
Second, a synthetic oligonucleotide with 5-methylcytosine substituted from the internal
cytosine is bound by R.PvuII in gel mobility shift assays. Third, the same methylated
oligonucleotide is slowly cleaved by R.PvuII (but not at all by R.AluI). These results are
consistent with the recently-determined structure of R.PvuII-DNA cocrystals.

<>

<1>Rice, M.R., Koons, M.D., Blumenthal, R.M.
<2>Substrate recognition by the PvuII endonuclease: binding and cleavage of CAG5mCTG sites.
<3>Nucleic Acids Res.
<4>27
<5>1032-1038
<6>1999
<7>The PvuII restriction endonuclease (R.PvuII) cleaves CAG/CTG sequences as indicated, leaving
blunt ends.  Its cognate methyltransferase generates N4-methylcytosine, yielding CAGN4mCTG,
though the mechanism by which this prevents cleavage by R.PvuII is unknown.  The heterologous
5-methylcytosine methylation CAG5mCTG has also been reported to prevent cleavage by R.PvuII
and this has been used in some cloning methods.  Since this heterologous methylation occurs at
the native methylated base, it can provide insights into the detection of DNA methylation by
R.PvuII.  We found that the cloned gene for R.PvuII could not stably transform cells protected
only by M.AluI (AG5mCT) and then determined that R.PvuII cleaves CAG5mCTG in vitro, even when
both strands are methylated.  DNase I footprint analysis and competition experiments reveal
that R.PvuII binds to CAG5mCTG specifically, though with reduced affinity relative to the
unmethylated sequence.  These results provide biochemical support for the published structures
of R.PvuII complexed with DNA containing CAGCTG and CAG5-iodoCTG and support a model for how
methylation interferes with DNA cleavage by this enzyme.

<>

<1>Richards, D.F., Linnett, P.E., Oultram, J.D., Young, M.
<2>Restriction endonucleases in Clostridium pasteurianum ATCC 6013 and Clostridium thermohydrosulfuricum DSM568.
<3>J. Gen. Microbiol.
<4>134
<5>3151-3157
<6>1988
<7>A small collection of Clostridia were surveyed for type II restriction endonucleases. Enzymes
were detected in two organisms. Clostridium pasteurianum ATCC 6013 contains an isoschizomer of
ThaI (FnuDII) [5'-CGCG-3'] and preliminary evidence suggests that cleavage generates
blunt-ended fragments. Clostridium thermohydrosulfuricum DSM568 contains an isoschizomer of
MboI (Sau3A) [5'-GATC-3'], that is inactive on dam methylated substrates. The DNA of this
latter organism shows dam methylation.

<>

<1>Richards, G.P., Bono, J.L., Watson, M.A., Needleman, D.S.
<2>Complete Genome Sequence for the Shellfish Pathogen Vibrio coralliilyticus RE98 Isolated from a Shellfish Hatchery.
<3>Genome Announcements
<4>2
<5>e01253-14
<6>2014
<7>Vibrio coralliilyticus is a pathogen of corals and larval shellfish. Publications on strain
RE98 list it as a Vibrio tubiashii; however, whole genome sequencing
confirms RE98 as V. coralliilyticus containing a total of 6,037,824 bp consisting
of two chromosomes (3,420,228 and 1,917,482 bp) and two megaplasmids (380,714 and
319,400 bp).

<>

<1>Richards, G.P., Needleman, D.S., Watson, M.A., Bono, J.L.
<2>Complete Genome Sequence of the Larval Shellfish Pathogen Vibrio tubiashii Type Strain ATCC 19109.
<3>Genome Announcements
<4>2
<5>e01252-14
<6>2014
<7>Vibrio tubiashii is a larval shellfish pathogen. Here, we report the first closed genome
sequence for this species (ATCC type strain 19109), which consists of two
chromosomes (3,294,490 and 1,766,582 bp), two megaplasmids (251,408 and 122,808
bp), and two plasmids (57,076 and 47,973 bp).

<>

<1>Richards, V.P., Lang, P., Pavinski, B., P, D., Lefebure, T., Schukken, Y.H., Zadoks, R.N., Stanhope, M.J.
<2>Comparative genomics and the role of lateral gene transfer in the evolution of bovine adapted Streptococcus agalactiae.
<3>Infect. Genet. Evol.
<4>11
<5>1263-1275
<6>2011
<7>In addition to causing severe invasive infections in humans, Streptococcus
agalactiae, or group B Streptococcus (GBS), is also a major cause of bovine
mastitis. Here we provide the first genome sequence for S. agalactiae isolated
from a cow diagnosed with clinical mastitis (strain FSL S3-026). Comparison to
eight S. agalactiae genomes obtained from human disease isolates revealed 183
genes specific to the bovine strain. Subsequent polymerase chain reaction (PCR)
screening for the presence/absence of a subset of these loci in additional bovine
and human strains revealed strong differentiation between the two groups (Fisher
exact test: p<0.0001). The majority of the bovine strain-specific genes (
approximately 85%) clustered tightly into eight genomic islands, suggesting these
genes were acquired through lateral gene transfer (LGT). This bovine GBS also
contained an unusually high proportion of insertion sequences (4.3% of the total
genome), suggesting frequent genomic rearrangement. Comparison to other
mastitis-causing species of bacteria provided strong evidence for two cases of
interspecies LGT within the shared bovine environment: bovine S. agalactiae with
Streptococcus uberis (nisin U operon) and Streptococcus dysgalactiae subsp.
dysgalactiae (lactose operon). We also found evidence for LGT, involving the
salivaricin operon, between the bovine S. agalactiae strain and either
Streptococcus pyogenes or Streptococcus salivarius. Our findings provide insight
into mechanisms facilitating environmental adaptation and acquisition of
potential virulence factors, while highlighting both the key role LGT has played
in the recent evolution of the bovine S. agalactiae strain, and the importance of
LGT among pathogens within a shared environment.

<>

<1>Richardson, B., Yung, R.
<2>Role of DNA methylation in the regulation of cell function.
<3>J. Lab. Clin. Med.
<4>134
<5>333-340
<6>1999
<7>The methylation of DNA helps stabilize chromatin in an inactive configuration and inhibits
gene transcription. This mechanism of gene regulation is involved in essential genetic events
including differentiation, genomic imprinting, and X chromosome inactivation. The alteration
of methylation patterns can result in abnormal gene expression, with significant pathologic
effects including carcinogenesis, autoimmunity, and some of the changes in gene expression
associated with aging. The mechanisms establishing, maintaining, and modifying methylation
patterns in normal and pathologic states are only now becoming understood, as are the
mechanisms relating DNA methylation to gene expression and chromosome inactivation. Further
characterization of these mechanisms holds promise for delaying or preventing the changes in
methylation patterns that contribute to cancer, autoimmunity, and aging.

<>

<1>Richardson, C., Elliott, B., Jasin, M.
<2>Chromosomal double strand breaks induced in mammalian cells by expression of I-SceI endonuclease.
<3>Methods Mol. Biol., Humana Press, Inc., Henderson, D.S., Totowa
<4>113
<5>453-463
<6>1999
<7>Until recently, investigators interested in analyzing the repair of chromosomal double-strand
breaks in mammalian cells have been limited by the inability to introduce defined DSBs within
the genome.  Traditional methods of introducing breaks have included irradiation or the
introduction of restriction enzymes; however, both of these methods cause multiple lesions at
different chromosomal loci.  Many of these types of studies have relied on cytogenetics for
the detection of gross genomic changes owing to misrepair at these damaged sites.

<>

<1>Richardson, E.J., Limaye, B., Inamdar, H., Datta, A., Manjari, K.S., Pullinger, G.D., Thomson, N.R., Joshi, R.R., Watson, M., Stevens, M.P.
<2>Genome sequences of Salmonella enterica serovar Typhimurium, Choleraesuis, Dublin and Gallinarum strains of highly defined virulence in  food-producing animals.
<3>J. Bacteriol.
<4>193
<5>3162-3163
<6>2011
<7>Salmonella enterica is an animal and zoonotic pathogen of worldwide importance and may be
classified into serovars differing in virulence and
host range. We sequenced and annotated the genomes of serovar Typhimurium,
Choleraesuis, Dublin and Gallinarum strains of defined virulence in each
of three food-producing animal hosts. This provides valuable measures of
intra-serovar diversity and opportunities to formally link genotypes to
phenotypes in target animals.

<>

<1>Richardson, F.C., Richardson, K.K.
<2>Alterations in DNA-restriction enzyme interactions by O4-alkyldeoxythymidines.
<3>Mol. Carcinog.
<4>4
<5>162-168
<6>1991
<7>O4-Alkyldeoxythymidines have been extensively studied for their ability to cause mutations and
to induce cancer.  Since these adducts can change DNA conformation, they may also have a more
immediate effect of altering DNA-protein interactions.  To address this issue, the effects of
these adducts on restriction enzyme activity were examined.  Oligodeoxyribonucleosides
containing O4-ethyldeoxythymidine (O4-EtdT) or O4-methyldeoxythymidine (O4-MedT) at a unique
site within the sequence 5'-GAATGGATCCTAATGAGATC-3' were constructed by automated DNA
synthesis.  This sequence contains the recognition site for various restriction enzymes.
These oligomers were annealed to various complementary strands and digested with restriction
enzymes:  BamHI or BstI (GGATCC); Sau3A, NdeII, or MboI (GATC); DpnI (GmATC); and BstYI, MflI,
or XhoII (PuGATCPy).  Analysis of the digests demonstrated that the presence of either O4-EtdT
or O4-MedT abolished the ability of XhoII, MboI, MflI, or NdeII to cut at the restriction
site.  DpnI failed to cut any of the oligomers.  BamHI, Sau3A, BstI, and BstYI exhibited
alterations in cutting specificity depending upon the oligomers used.  These results
demonstrated that O4-alkyldeoxythymidine adducts alter DNA-restriction enzyme interactions in
a protein- and sequence-dependent manner.  Because of the importance of natural methylation in
genetic regulation it is possible that aberrant methylation in the form of DNA adducts could
also alter protein-DNA interactions in cells exposed to DNA-modifying agents.

<>

<1>Richardson, F.C., Richardson, K.K., Kroin, J.S., Hertel, L.W.
<2>Synthesis and restriction enzyme analysis of oligodeoxyribonucleotides containing the anti-cancer drug 2', 2'-difluoro-2'-deoxycytidine.
<3>Nucleic Acids Res.
<4>20
<5>1763-1768
<6>1992
<7>The anti-cancer drug 2',2'-difluoro-2'-deoxycytidine (dFdC) is internally incorporated into
DNA in vitro. To determine the effects of this incorporation on DNA structure and function,
the beta-cyanoethyl phosphoramidite of dFdC was synthesized and oligodeoxyribonucleotides
containing dFdC were made using automated solid phase DNA synthesis techniques. Extension of
the coupling time was required to achieve high coupling efficiency, suggesting a significant
reduction in the rate of phosphotriester formation. Insertion of dFdC 5' into the recognition
sequence of restriction enzymes HpaII and KpnI reduced the rate of cutting by 4% and 14% over
60 minutes. This reduction is similar to the effects seen with arabinofuranosylcytidine
(ara-C) but small compared to the reductions caused by base analogues and phosphothioates.
Insertion of dFdC into the BamHI recognition sequence, but not 5' to the cut site, did not
alter the rate of cutting/recognition. The presence of a single dFdC reduced the Tm's of
oligomers by 2-4C, depending on sequence and location. These results demonstrate that, once
incorporated into DNA, dFdC does not greatly alter recognition between DNA and restriction
enzymes; however, it does significantly alter duplex stability.

<>

<1>Richter, M., Kube, M., Bazylinski, D.A., Lombardot, T., Glockner, F.O., Reinhardt, R., Schuler, D.
<2>Comparative genome analysis of four magnetotactic bacteria reveals a complex set of group-specific genes implicated in magnetosome biomineralization and function.
<3>J. Bacteriol.
<4>189
<5>4899-4910
<6>2007
<7>Magnetotactic bacteria (MTB) are a heterogeneous group of aquatic
prokaryotes with a unique intracellular organelle, the magnetosome, which
orients the cell along magnetic field lines. Magnetotaxis is a complex
phenotype, which depends on the coordinate synthesis of magnetosomes and
the ability to swim and orient along the direction caused by the
interaction with the Earth's magnetic field. Although a number of putative
magnetotaxis genes were recently identified within a conserved genomic
magnetosome island (MAI) of several MTB, their functions have remained
mostly unknown, and it was speculated that additional genes located
outside the MAI might be involved in magnetosome formation and
magnetotaxis. In order to identify genes specifically associated with the
magnetotactic phenotype, we conducted comparisons between four sequenced
magnetotactic Alphaproteobacteria including the nearly complete genome of
Magnetospirillum gryphiswaldense strain MSR-1, the complete genome of
Magnetospirillum magneticum strain AMB-1, the complete genome of the
magnetic coccus MC-1, and the comparative-ready preliminary genome
assembly of Magnetospirillum magnetotacticum strain MS-1 against an
in-house database comprising 426 complete bacterial and archaeal genome
sequences. A magnetobacterial core genome of about 891 genes was found
shared by all four MTB. In addition to a set of approximately 152
genus-specific genes shared by the three Magnetospirillum strains, we
identified 28 genes as group specific, i.e., which occur in all four
analyzed MTB but exhibit no (MTB-specific genes) or only remote
(MTB-related genes) similarity to any genes from nonmagnetotactic
organisms and which besides various novel genes include nearly all mam and
mms genes previously shown to control magnetosome formation. The
MTB-specific and MTB-related genes to a large extent display synteny,
partially encode previously unrecognized magnetosome membrane proteins,
and are either located within (18 genes) or outside (10 genes) the MAI of
M. gryphiswaldense. These genes, which represent less than 1% of the 4,268
open reading frames of the MSR-1 genome, as yet are mostly of unknown
functions but are likely to be specifically involved in magnetotaxis and,
thus, represent prime targets for future experimental analysis.

<>

<1>Riddihough, G.
<2>No limits on restriction.
<3>Nature
<4>370
<5>78
<6>1994
<7>The structure of the endonuclease R.PvuII in the absence and presence of its recognition site
indicates how the interaction with DNA might proceed.

<>

<1>Riedel, T. et al.
<2>Genome sequence of the orange-pigmented seawater bacterium Owenweeksia hongkongensis type strain (UST20020801(T)).
<3>Standards in Genomic Sciences
<4>7
<5>120-130
<6>2012
<7>Lau . 2005 is the sole member of the monospecific genus in the family a poorly characterized
family at the genome level thus far. This family comprises seven
genera within the class . Family members are known to be psychrotolerant,
rod-shaped and orange pigmented (beta-carotene), typical for . For growth,
seawater and complex organic nutrients are necessary. The genome of UST20020801
is only the second genome of a member of the family whose sequence has been
deciphered. Here we describe the features of this organism, together with the
complete genome sequence and annotation. The 4,000,057 bp long chromosome with
its 3,518 protein-coding and 45 RNA genes is a part of the project.

<>

<1>Riedel, T. et al.
<2>Genome sequence of the Antarctic rhodopsins-containing flavobacterium Gillisia limnaea type strain (R-8282(T)).
<3>Standards in Genomic Sciences
<4>7
<5>107-119
<6>2012
<7>Van Trappen et al. 2004 is the type species of the genus , which is a member of the well
characterized family . The genome of R-8282 is the first sequenced
genome (permanent draft) from a type strain of the genus . Here we describe the
features of this organism, together with the permanent-draft genome sequence and
annotation. The 3,966,857 bp long chromosome (two scaffolds) with its 3,569
protein-coding and 51 RNA genes is a part of the of and project.

<>

<1>Riedel, T. et al.
<2>Genome sequence of the Leisingera aquimarina type strain (DSM 24565(T)), a member of the marine Roseobacter clade rich in extrachromosomal elements.
<3>Standards in Genomic Sciences
<4>8
<5>389-402
<6>2013
<7>Leisingera aquimarina Vandecandelaere et al. 2008 is a member of the genomically  well
characterized Roseobacter clade within the family Rhodobacteraceae.
Representatives of the marine Roseobacter clade are metabolically versatile and
involved in carbon fixation and biogeochemical processes. They form a
physiologically heterogeneous group, found predominantly in coastal or polar
waters, especially in symbiosis with algae, in microbial mats, in sediments or
associated with invertebrates. Here we describe the features of L. aquimarina DSM
24565(T) together with the permanent-draft genome sequence and annotation. The
5,344,253 bp long genome consists of one chromosome and an unusually high number
of seven extrachromosomal elements and contains 5,129 protein-coding and 89 RNA
genes. It was sequenced as part of the DOE Joint Genome Institute Community
Sequencing Program 2010 and of the activities of the Transregional Collaborative
Research Centre 51 funded by the German Research Foundation (DFG).

<>

<1>Riedel, T., Fiebig, A., Goker, M., Klenk, H.P.
<2>Complete genome sequence of the bacteriochlorophyll a-containing Roseibacterium elongatum type strain (DSM 19469(T)), a representative of the Roseobacter group  isolated from Australian coast sand.
<3>Standards in Genomic Sciences
<4>9
<5>840-854
<6>2014
<7>Roseibacterium elongatum Suzuki et al. 2006 is a pink-pigmented and bacteriochlorophyll
a-producing representative of the Roseobacter group within
the alphaproteobacterial family Rhodobacteraceae. Representatives of the marine
'Roseobacter group' were found to be abundant in the ocean and play an important
role in global and biogeochemical processes. In the present study we describe the
features of R. elongatum strain OCh 323(T) together with its genome sequence and
annotation. The 3,555,102 bp long genome consists of one circular chromosome with
no extrachromosomal elements and is one of the smallest known Roseobacter
genomes. It contains 3,540 protein-coding genes and 59 RNA genes. Genome analysis
revealed the presence of a photosynthetic gene cluster, which putatively enables
a photoheterotrophic lifestyle. Gene sequences associated with quorum sensing,
motility, surface attachment, and thiosulfate and carbon monoxide oxidation could
be detected. The genome was sequenced as part of the activities of the
Transregional Collaborative Research Centre 51 (TRR51) funded by the German
Research Foundation (DFG).

<>

<1>Riedel, T., Fiebig, A., Han, J., Huntemann, M., Spring, S., Petersen, J., Ivanova, N.N., Markowitz, V., Woyke, T., Goker, M., Kyrpides, N.C., Klenk, H.P.
<2>Genome sequence of the Wenxinia marina type strain (DSM 24838(T)), a representative of the Roseobacter group isolated from oilfield sediments.
<3>Standards in Genomic Sciences
<4>9
<5>855-865
<6>2014
<7>Wenxinia marina Ying et al. 2007 is the type species of the genus Wenxinia, a representative
of the Roseobacter group within the alphaproteobacterial family
Rhodobacteraceae, isolated from oilfield sediments of the South China Sea. This
family was shown to harbor the most abundant bacteria especially from coastal and
polar waters, but was also found in microbial mats, sediments and attached to
different kind of surfaces. Here we describe the features of W. marina strain
HY34(T) together with the genome sequence and annotation of strain DSM 24838(T)
and novel aspects of its phenotype. The 4,181,754 bp containing genome sequence
encodes 4,047 protein-coding genes and 59 RNA genes. The genome of W. marina DSM
24838(T) was sequenced as part of the activities of the Genomic Encyclopedia of
Type Strains, Phase I: the one thousand microbial genomes (KMG) project funded by
the DoE and the Transregional Collaborative Research Centre 51 (TRR51) funded by
the German Research Foundation (DFG).

<>

<1>Riedel, T., Fiebig, A., Petersen, J., Gronow, S., Kyrpides, N.C., Goker, M., Klenk, H.P.
<2>Genome sequence of the Litoreibacter arenae type strain (DSM 19593(T)), a member  of the Roseobacter clade isolated from sea sand.
<3>Standards in Genomic Sciences
<4>9
<5>117-127
<6>2013
<7>Litoreibacter arenae Kim et al. 2012 is a member of the genomically well-characterized
Rhodobacteraceae clade within the Roseobacter clade.
Representatives of this clade are known to be metabolically versatile and
involved in marine carbon-producing and biogeochemical processes. They form a
physiologically heterogeneous group of Alphaproteobacteria and were mostly found
in coastal or polar waters, especially in symbiosis with algae, in microbial
mats, in sediments or together with invertebrates and vertebrates. Here we
describe the features of L. arenae DSM 19593(T), including novel aspects of its
phenotype, together with the draft genome sequence and annotation. The 3,690,113
bp long genome consists of 17 scaffolds with 3,601 protein-coding and 56 RNA
genes. This genome was sequenced as part of the activities of the Transregional
Collaborative Research Centre 51 funded by the German Research Foundation (DFG).

<>

<1>Riedel, T., Spring, S., Fiebig, A., Petersen, J., Goker, M., Klenk, H.P.
<2>Genome sequence of the pink to light reddish-pigmented Rubellimicrobium mesophilum type strain (DSM 19309(T)), a representative of the Roseobacter group   isolated from soil, and emended description of the species.
<3>Standards in Genomic Sciences
<4>9
<5>902-913
<6>2014
<7>Rubellimicrobium mesophilum Dastager et al. 2008 is a mesophilic and light reddish-pigmented
representative of the Roseobacter group within the
alphaproteobacterial family Rhodobacteraceae. Representatives of the Roseobacter
group play an important role in the marine biogeochemical cycles and were found
in a broad variety of marine environments associated with algal blooms, different
kinds of sediments, and surfaces of invertebrates and vertebrates. Roseobacters
were shown to be widely distributed, especially within the total bacterial
community found in coastal waters, as well as in mixed water layers of the open
ocean. Here we describe the features of R. mesophilum strain MSL-20(T) together
with its genome sequence and annotation generated from a culture of DSM 19309(T).
The 4,927,676 bp genome sequence consists of one chromosome and probably one
extrachromosomal element. It contains 5,082 protein-coding genes and 56 RNA
genes. As previously reported, the G+C content is significantly different from
the actual genome sequence-based G+C content and as the type strain tests
positively for oxidase, the species description is emended accordingly. The
genome was sequenced as part of the activities of the Transregional Collaborative
Research Centre 51 (TRR51) funded by the German Research Foundation (DFG).

<>

<1>Riedel, T., Spring, S., Fiebig, A., Petersen, J., Kyrpides, N.C., Goker, M., Klenk, H.P.
<2>Genome sequence of the exopolysaccharide-producing Salipiger mucosus type strain  (DSM 16094(T)), a moderately halophilic member of the Roseobacter clade.
<3>Standards in Genomic Sciences
<4>9
<5>1333-1345
<6>2014
<7>Salipiger mucosus Martinez-Canovas et al. 2004 is the type species of the genus Salipiger, a
moderately halophilic and exopolysaccharide-producing representative
of the Roseobacter lineage within the alphaproteobacterial family
Rhodobacteraceae. Members of this family were shown to be the most abundant
bacteria especially in coastal and polar waters, but were also found in microbial
mats and sediments. Here we describe the features of the S. mucosus strain DSM
16094(T) together with its genome sequence and annotation. The 5,689,389-bp
genome sequence consists of one chromosome and several extrachromosomal elements.
It contains 5,650 protein-coding genes and 95 RNA genes. The genome of S. mucosus
DSM 16094(T) was sequenced as part of the activities of the Transregional
Collaborative Research Center 51 (TRR51) funded by the German Research Foundation
(DFG).

<>

<1>Rieger, A., Nassal, M.
<2>Restriction endonuclease AlwNI is blocked by overlapping Dcm methylation.
<3>Nucleic Acids Res.
<4>21
<5>4148
<6>1993
<7>Many restriction enzymes are unable to cleave DNA if their recognition sequence contains
methylated residues. The two sequence-specific methylases present in most laboratory strains
of Escherichia coli are the Dam methylase, which methylates the N6 position of A within the
sequence GATC, and the Dcm methylase, which methylates the C5 position of the internal C
residue with the sequence CC(A/T)GG. During the construction of derivatives of plasmid
pCHG-3122, which contains three recognition sites for the restriction endonuclease AlwNI, we
observed that one of the sites was only poorly cleaved when the plasmid DNA was isolated from
E. coli strain DH5-alpha. Inspection of the sequences flanking the three AlwNI recognition
sites showed that one of them located inside the LacZ fragment (ant position 2145) overlapped
with a recognition site for Dcm Methylase (see Table 1).

<>

<1>Rifat, D., Wright, N.T., Varney, K.M., Weber, D.J., Black, L.W.
<2>Restriction Endonuclease Inhibitor IPI* of Bacteriophage T4: A Novel Structure for a Dedicated Target.
<3>J. Mol. Biol.
<4>375
<5>720-734
<6>2008
<7>Phage T4 protects its DNA from the two-gene-encoded gmrS/gmrD (glucose-modified
hydroxymethylcytosine restriction endonuclease) CT of
pathogenic Escherichia coli, CT596, by injecting several hundred copies of
the 76-amino-acid-residue nuclease inhibitor, IPI*, into the infected
host. Here, the three-dimensional solution structure of mature IPI* is
reported as determined by nuclear magnetic resonance techniques using 1290
experimental nuclear Overhauser effect and dipolar coupling constraints (
approximately 17 constraints per residue). Close examination of this
oblate-shaped protein structure reveals a novel fold consisting of two
small beta-sheets (beta1: B1 and B2; beta2: B3-B5) flanked at the N- and
C-termini by alpha-helices (H1 and H2). Such a fold is very compact in
shape and allows ejection of IPI* through the narrow 30-A portal and tail
tube apertures of the virion without unfolding. Structural and dynamic
measurements identify an exposed hydrophobic knob that is a putative
gmrS/gmrD-binding site. A single gene from the uropathogenic E. coli
UT189, which codes for a gmrS/gmrD-like UT fusion enzyme (with
approximately 90% identity to the heterodimeric CT enzyme), has evolved
IPI* inhibitor immunity. Analysis of the gmrS/gmrD restriction
endonuclease enzyme family and its IPI* family phage antagonists reveals
an evolutionary pathway that has elaborated a surprisingly diverse and
specifically fitted set of coevolving attack and defense structures.

<>

<1>Riggs, A.D.
<2>X chromosome inactivation, differentiation, and DNA methylation revisited, with a tribute to Susumu Ohno.
<3>Cytogenet. Genome Res.
<4>99
<5>17-24
<6>2002
<7>X chromosome inactivation and DNA methylation are reviewed, with emphasis on the contributions
of Susumu Ohno and the predictions made
in my 1975 paper (Riggs, 1975) in which I proposed the "maintenance
methylase" model for somatic inheritance of methylation patterns and
suggested that DNA methylation would be involved in mammalian X
chromosome inactivation and development. The maintenance methylase
model is discussed and updated to consider methylation patterns in cell
populations that have occasional, stochastic methylation changes by de
novo methylation or demethylation, either active or passive. The "way
station" model for the spread of X inactivation by LINE- 1 elements is
also considered, and some recent results from my laboratory are briefly
reviewed.

<>

<1>Riipinen, K.A., Forsman, P., Alatossava, T.
<2>The genomes and comparative genomics of Lactobacillus delbrueckii phages.
<3>Arch. Virol.
<4>156
<5>1217-1233
<6>2011
<7>Lactobacillus delbrueckii phages are a great source of genetic diversity.
Here, the genome sequences of Lb. delbrueckii phages LL-Ku, c5 and JCL1032
were analyzed in detail, and the genetic diversity of Lb. delbrueckii
phages belonging to different taxonomic groups was explored. The lytic
isometric group b phages LL-Ku (31,080 bp) and c5 (31,841 bp) showed a
minimum nucleotide sequence identity of 90% over about three-fourths of
their genomes. The genomic locations of their lysis modules were unique,
and the genomes featured several putative overlapping transcription units
of genes. LL-Ku and c5 virions displayed peptidoglycan hydrolytic activity
associated with a ~36-kDa protein similar in size to the endolysin.
Unexpectedly, the 49,433-bp genome of the prolate phage JCL1032
(temperate, group c) revealed a conserved gene order within its structural
genes. Lb. delbrueckii phages representing groups a (a phage LL-H), b and
c possessed only limited protein sequence homology. Genomic comparison of
LL-Ku and c5 suggested that diversification of Lb. delbrueckii phages is
mainly due to insertions, deletions and recombination. For the first time,
the complete genome sequences of group b and c Lb. delbrueckii phages are
reported.

<>

<1>Rimbara, E., Matsui, M., Mori, S., Suzuki, S., Suzuki, M., Kim, H., Sekizuka, T., Kuroda, M., Shibayama, K.
<2>Draft Genome Sequence of Helicobacter fennelliae Strain MRY12-0050, Isolated from a Bacteremia Patient.
<3>Genome Announcements
<4>1
<5>e00512-13
<6>2013
<7>Helicobacter fennelliae, a human enterohepatic pathogen, causes bacteremia and colitis. We
isolated H. fennelliae strain MRY12-0050 from a female patient; this
strain was isolated from 2 other patients from the same hospital during the same
period, suggesting human-to-human transmission. This is the first report of an H.
fennelliae genome sequence.

<>

<1>Rimseliene, R., Janulaitis, A.
<2>Saturation mutagenesis of Thr862, the amino acid essential for substrate specificity of Eco57I restriction endonuclease.
<3>Biologija
<4>1
<5>11-14
<6>2005
<7>Type IIG restriction endonuclease Eco57I cleaves DNA 16/14 nucleotides away from the
asymmetric recognition sequence 5'-CTGAAG.  The enzyme also possesses methyltransferase
activity that modifies the second A base within the 5'-CTGAAG strand of the target duplex
(underlined).  In previous studies, Eco57I mutants with altered substrate specificity
5'-CTGRAG were isolated.  These mutant enzymes have Asn or Ser instead of Thr in the 862th
position of the protein.  In order to evaluate the impact of T862 on the substrate
specificity, it was changed to the other 17 amino acids.  The in vivo cleavage activity and
substrate specificity of the resulting mutant enzymes was examined (i) by testing lethality of
the mutants to the host cells in the absence or presence of Eco57I (specificity 5'-CTGAAG) and
GsuI (specificity 5'-CTGGAG) methyltransferases, and (ii) by testing the ability of the
mutants to induce SOS DNA repair response in the absence or presence of protecting
methyltransferases.  The results indicate that mutants T862G, T862C and, probably, T862A and
T862D could display altered substrate specificity.  The recognition sequence of T862F, H, K,
L, Q, M and Y mutants was the same as that of the wild type enzyme.  The remaining
substitutions rendered the enzyme catalytically inactive.

<>

<1>Rimseliene, R., Janulaitis, A.
<2>Mutational analysis of two putative catalytic motifs of the Type IV restriction endonuclease Eco57I.
<3>J. Biol. Chem.
<4>276
<5>10492-10497
<6>2001
<7>The role of two sequence motifs (SM) as putative cleavage catalytic centers (77)PD(X13)EAK (SM
I) and (811)PD(X20)DQK (SM II) of type IV restriction endonuclease Eco57I was studied by
site-directed mutational analysis. Substitutions within SM I; D78N, D78A, D78K, and E92Q
reduced cleavage activity of Eco57I to a level undetectable both in vivo and in vitro.
Residual endonucleolytic activity of the E92Q mutant was detected only when the Mg(2+) in the
standard reaction mixture was replaced with Mn(2+). The mutants D78N and E92Q retained the
ability to interact with DNA specifically. The mutants also retained DNA methylation activity
of Eco57I. The properties of the SM I mutants indicate that Asp(78) and Glu(92) residues are
essential for cleavage activity of the Eco57I, suggesting that the sequence motif
(77)PD(X13)EAK represents the cleavage active site of this endonuclease. Eco57I mutants
containing single amino acid substitutions within SM II (D812A, D833N, D833A) revealed only a
small or moderate decrease of cleavage activity as compared with wild-type Eco57I, indicating
that the SM II motif does not represent the catalytic center of Eco57I. The results, taken
together, allow us to conclude that the Eco57I restriction endonuclease has one catalytic
center for cleavage of DNA.

<>

<1>Rimseliene, R., Maneliene, Z., Lubys, A., Janulaitis, A.
<2>Engineering of restriction endonucleases: using methylation activity of the bifunctional endonuclease Eco571 to select the mutant with a novel sequence specificity.
<3>J. Mol. Biol.
<4>327
<5>383-391
<6>2003
<7>Type II restriction endonucleases (REs) are widely used tools in molecular biology,
biotechnology and diagnostics. Efforts to generate new
specificities by structure-guided design and random mutagenesis have been
unsuccessful so far. We have developed a new procedure called the
methylation activity-based selection (MABS) for generating REs with a new
specificity. MABS uses a unique property of bifunctional type II REs to
methylate DNA targets they recognize. The procedure includes three steps:
(1) conversion of a bifunctional RE into a monofunctional DNA-modifying
enzyme by cleavage center disruption; (2) mutagenesis and selection of
mutants with altered DNA modification specificity based on their ability
to protect predetermined DNA targets; (3) reconstitution of the cleavage
center's wild-type structure. The efficiency of the MABS technique was
demonstrated by altering the sequence specificity of the bifunctional RE
Eco57I from 5'-CTGAAG to 5'-CTGRAG, and thus generating the mutant
restriction endonuclease (and DNA methyltransferase) of a specificity not
known before. This study provides evidence that MABS is a promising
technique for generation of REs with new specificities.

<>

<1>Rimseliene, R., Timinskas, A.
<2>Site-directed mutagenesis of type IV restriction endonuclease Eco57I.
<3>Biologija
<4>1
<5>31-33
<6>1997
<7>Amino acid sequence analysis of type IV restriction endonuclease Eco57I revealed two sequence
motifs 77PDX13EXK and 811PDX20 DXK as putative catalytic sites.  We have used a site-directed
mutational analysis of these regions in order, to determine their role in catalysis.
Catalytic properties of the mutants were studied in vivo and in vitro.  The replacement of D78
and E92, respectively, by mutations N78, A78, K78, Q92, resulted in the complete loss of
cleavage activity.  The D833N mutant cleaves DNA with reduced rate.  The activity of the D812A
mutant seems to be the same as that of the wild type R. Eco57I.  The results suggest that
amino acids D87 and E92 are essential for cleavage activity of the Eco57I restriction
endonuclease, and the sequence motif 77PDX13EXK is a catalytic center of the enzyme.

<>

<1>Rimseliene, R., Vaisvila, R.
<2>Cloning the PaeI restriction-modification system in Escherichia coli.
<3>Biologija
<4>2
<5>77-78
<6>1998
<7>PaeI, type II restriction-modification system from bacterium Pseudomonas aeruginosa,
recognizes DNA sequence 5'-CGATCG-3'.  The PaeI methyltransferase (Mtase)-encoding gene,
paeIM, was cloned into Escherichia coli using biochemical Mtase selection method.  According
to the results of Southern-blot analysis, chromosomal map of Pseudomonas aeruginosa was
generated localizing the paeIM gene and flanking regions.  The paeIR gene was cloned as a 2.1
kb Eco47III DNA fragment into pBR322 vector.  In order to increase the PaeI restriction
endonuclease expression and yield in E. coli, the paeIR gene was subcloned into multicopy
plasmid pIC-19H.  80000 units of R.PaeI per gram of wet-weight cells were produced, approx. 20
times more than were produced by P. aeruginosa.

<>

<1>Rimseliene, R., Vaisvila, R., Janulaitis, A.
<2>The eco72IC gene specifies a trans-acting factor which influences expression of both DNA methyltransferase and endonuclease from the Eco72I restriction-modification system.
<3>Gene
<4>157
<5>217-219
<6>1995
<7>Eco72I from Escherichia coli RFL72 is a type-II restriction-modification (R-M) system
recognizing and cleaving the sequence 5'-CAC/GTG-3'.  The R-M genes are transcribed
divergently and between the two genes is a small open reading frame codirectional to the R
gene.  This small ORF acts both to stimulate ENase expression and to depress DNA
methyltransferase synthesis.  The activity of beta-Gal produced from the eco72IM::lacZ
translational fusion increased ten-fold, and eco72IR::lacZ translational fusion beta-Gal
activity decreased 130-fold when eco72IC was inactivated by a frameshift mutation.  Analysis
of nucleotide sequences of R-M systems, containing C genes, revealed a 5'-ACCTTATAGTC-3'
consensus sequence upstream from the regulatory genes in all six analysed R-M systems.  This
sequence, named the C-box, may play the role of an operator sequence.

<>

<1>Rina, M., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BsiSI.
<3>Nucleic Acids Res.
<4>18
<5>1654
<6>1990
<7>None

<>

<1>Rina, M., Bouriotis, V.
<2>Cloning, purification and characterization of the BseCI DNA methyltransferase from Bacillus stearothermophilus.
<3>Gene
<4>133
<5>91-94
<6>1993
<7>The gene (bseCIM) encoding the BseCI DNA methyltransferase (MTase; M.BseCI) from a Bacillus
stearothermophilus species was cloned and expressed in Escherichia coli using plasmid vector
pBR322. Selection of transformants carrying bseCIM was based on resistance of the modified
plasmid to cleavage by BseCI. The MTase was purified to homogeneity and further characterized.
Its size as determined by sodium dodecyl sulfate-polyacrylamide-gel electrophoresis and size
exclusion chromatography was 68 kDa, suggesting that the MTase exists as a monomer. When phage
lambda DNA was used as a substrate, the optimum temperature for MTase activity was determined
to be 50-55 degrees C and optimum pH approx 7.4. M.BseCI is inhibited by concentrations of
NaCl and KCl greater than 50 mM, and it does not require Mg2+ for activity. Finally, M.BseCI
methylates the 3' adenine residue in the sequence, 5'ATCGAT 3', similarly to its
isoschizomer M.ClaI.

<>

<1>Rina, M., Caufrier, F., Markaki, M., Mavromatis, K., Kokkinidis, M., Bouriotis, V.
<2>Cloning and characterization of the gene encoding PspPI methyltransferase from the Antarctic psychrotroph Psychrobacter sp. strain TA137.
<3>Gene
<4>197
<5>353-360
<6>1997
<7>The gene (pspPIM) encoding the PspPI DNA methyltransferase (MTase) associated with the PspPI
restriction-modification system (5'-GGNCC-3') of Psychrobacter species TA137 has been cloned
and expressed in E. coli, and its nucleotide sequence has been determined.  The coding region
was 1248 nt in length and capable of specifying a 46,826-Da protein of 416 amino acids.  The
predicted sequence of the MTase protein displays ten sequence motifs characteristic of all
prokaryotic m5C-MTases and shows the highest similarity to other MTases that methylate the
GGNCC sequence, namely M.Eco47II and M.Sau96I.  All three MTases methylate the internal
cytosine within their recognition sequence.  Sequence similarities between M.PspPI and its
isospecific M.Eco47II and M.Sau96I as well as with four other m5C-MTases that methylate the
related GGWCC sequence, namely M.SinI, M.HgiCII, M.HgiBI, M.HgiEI have been also found within
the variable region of these proteins.  On the basis of structural information from M.HhaI and
M.HaeIII, several M.PspPI residues that are expected to interact with DNA can be predicted.
Furthermore, an organization of the variable region of m5C-MTases into two segments exhibiting
a pattern of conserved residues and a considerable degree of structural homologies is
described.

<>

<1>Rina, M., Clark, D., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BsiKI.
<3>Nucleic Acids Res.
<4>18
<5>1655
<6>1990
<7>None

<>

<1>Rina, M., Dialektakis, D., Clark, D., Pagomenou, M., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BseCl.
<3>Nucleic Acids Res.
<4>20
<5>1807
<6>1992
<7>BseCI, an isoschizomer of ClaI has been purified from Bacillus species. BseCI recognizes the
sequence 5' ...ATCGAT ...3' and cleaves between T and C. The enzyme was purified using the
following chromatographic steps: 1. Phosphocellulose, 2. Heparin-Sepharose, 3. DEAE-cellulose.

<>

<1>Rina, M., Karagouni, A., Pagomenou, M., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease SgrBI.
<3>Nucleic Acids Res.
<4>19
<5>6342
<6>1991
<7>SgrBI, an isoschizomer of SacII has been purified from Streptomyces griseus.
SgrBI recognises the palindromic sequence 5'...CCGCGG...3' generating
3'-protruding GC-dinucleotides.

<>

<1>Rina, M., Karagouni, A., Pagomenou, M., Tsigos, I., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease SseAI.
<3>Nucleic Acids Res.
<4>19
<5>6341
<6>1991
<7>SseAI, an isoschizomer of NarI, has been purified from Streptomyces species.
SseAI recognises the palindromic sequence 5'...GGCGCC...3' and cleaves between
the second G and C.

<>

<1>Rina, M., Markaki, M., Bouriotis, V.
<2>Sequence of the cloned bseCIM gene: M.BseCI reveals high homology to M.BanIII.
<3>Gene
<4>150
<5>71-73
<6>1994
<7>The bseCIM gene, encoding M.BseCI methyltransferase (MTase) from a Bacillus stearothermophilus
strain, has been previously cloned and expressed in Escherichia coli. The nucleotide (nt)
sequence of a 2357-bp BspMII-EcoRI fragment encoding bseCIM has now been determined. The
sequence predicts a MTase of 579 amino acids (aa), 66.7 kDa. Comparison of the deduced aa
sequence of M.BseCI with sequences of various MTases revealed a significant homology to
m6A-MTases, especially to its isoschizomer M.BanIII from Bacillus aneurinolyticus.

<>

<1>Rina, M., Stratidakis, I., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BssAI.
<3>Nucleic Acids Res.
<4>18
<5>6161
<6>1990
<7>None

<>

<1>Rina, M., Tsigos, I., Karagouni, A., Pagomenou, M., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BseBI.
<3>Nucleic Acids Res.
<4>19
<5>4776
<6>1991
<7>BseBI, an isoschizomer of BstNI has been purified from Bacillus
stearothermophilus species.  BseBI recognises the sequence 5'...CCWGG...3' (W=A
or T) and cleaves between C and W.

<>

<1>Rina, M., Tzanodaskalaki, M., Karagouni, A., Pagomenou, M., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease SseBI.
<3>Nucleic Acids Res.
<4>20
<5>1808
<6>1992
<7>SseBI, an isoschizomer of StuI has been purified from Streptomyces species. SseBI recognizes
the sequence 5' ... AGGCCT ...3' and cleaves between G and C. The enzyme was purified using
the following chromatographic steps: 1. Blue Sepharose F3GA, 2. Heparin-Sepharose.

<>

<1>Rina, M., Tzanodaskalaki, M., Karagouni, A., Pagomenou, M., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease MspCI.
<3>Nucleic Acids Res.
<4>20
<5>1806
<6>1992
<7>MspCI, an isoschizomer of AflII, has been purified from Micrococcus species. MspCI recognizes
the sequence 5' ...CTTAAG ...3' and cleaves between C and T. The enzyme was purified using
the following chromatographic steps: 1. Blue Sepharose F3GA; 2. Heparin-Sepharose; 3. DEAE
Sepharose.

<>

<1>Rine, J., Barnes, G.
<2>Entry of a procaryotic endonuclease into the nucleus of Saccharomyces cerevisiae.
<3>Yeast Cell Biology, Alan R. Liss, Inc., Hicks, J., NY
<4>0
<5>395-413
<6>1986
<7>Recent experiment have documented the existence of amino acid sequences, acting
as nuclear signal sequences, that result in the accumulation of proteins in the
nucleus.  In this paper, the issue of whether or not a protein must possess a
nuclear signal in order to enter the nucleus is examined.  For these
experiments, a plasmid capable of expressing EcoRI endonuclease in the yeast
Saccharomyces cerevisiae has been constructed and transformed into several
yeast strains.  Two results demonstrate that this bacterial protein can enter
the yeast nucleus:  First, yeast cells expressing the endonuclease gene die
with kinetics that are proportional to the capacity of the strain to repair
double stranded breaks in nuclear DNA.  Secondly, the nuclear DNA contains
extensive double stranded breaks at EcoRI sites and only at EcoRI sites.
Therefore, there is no apparent requirement for a protein to contain a complex
nuclear localization signal in order to enter the nucleus.  Additional
experiments demonstrate that in-frame fusion of an open reading frame to the 5'
end of the endonuclease structural gene results in synthesis of a hybrid
protein that retains endonucleolytic activity.  Mutants of the endonuclease are
described that allow the activity of the enzyme to be modulated independently
of its synthesis.

<>

<1>Ringquist, S., Smith, C.L.
<2>The Escherichia coli chromosome contains specific, unmethylated dam and dcm sites.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>4539-4543
<6>1992
<7>The Escherichia coli chromosome encodes two methylases, dam and dcm, which recognize the
sequences GATC and CC(A/T)GG, respectively. Specific dam and dcm sites on the E. coli
chromosome were found to be unmethylated in vivo by using pulsed-field gel electrophoresis
experiments scanning megabase regions of DNA. Some sites were totally unmethylated. The dam
sites display variable methylation depending on the local sequence, and, in general, their
methylation shows complex modulation by growth conditions and growth rate, suggesting multiple
protection mechanisms. Sites resistant to complete dam or dcm methylation appear to be
distributed throughout the chromosome. These unusual sites may identify regions of the
chromosome with interesting biological functions.

<>

<1>Rinkel, L.J., van der Marel, G.A., van Boom, J.H., Altona, C.
<2>Influence of N6-methylation of residue A(5) on the conformational behaviour of d(C-C-G-A-A-T-T-C-G-G) in solution studied by 1H-NMR spectroscopy 1. The duplex form.
<3>Eur. J. Biochem.
<4>163
<5>275-286
<6>1987
<7>One- and two-dimensional NMR studies at 300 MHz and 500 MHz were carried out on
the two oligonucleotides d(C-C-G-A-A-T-T-C-G-G) and d(C-C-G-A-m6A-T-T-C-G-G) in
aqueous solution.  NMR spectra was observed at 10 mM sample concentration over
the temperature range 273-368 K.  Assignments are given of the base, H1', H2',
H2", H3' and of some H4' resonances, based upon a combination of
two-dimensional correlation spectra (COSY) and two-dimensional nuclear
Overhauser effect spectra (NOESY); imino-proton resonances were assigned with
the aid of a two-dimensional NOE experiment.  Chemical shift vs temperature
profiles were constructed in order to gain insight into the influence of
N6-methylation of residue A(5) on the temperature-dependent conformational
behaviour of the decamer and to determine thermodynamic parameters for the
duplex-to-coil tranasition.  The NOESY spectra, the imino-proton spectra and
the shift profiles of the two compounds, under conditions where each forms a
B-DNA-type duplex, are very similar.  This is taken to indicate that the
influence of N6-methylation of residue A(5) on the local structure of the
duplex must be small.  However, the temperature dependence of the
(non-)exchangeable proton resonances of the two compounds reveals that
methylation slows down the duplex- single-strand exchange.  Furthermore, a
thermodynamic analysis of the two compounds indicates that N6-methylation
slightly decreases the stability of the duplex relative to the monomeric forms
(Tm is reduced from 332 K down to 325 K at 10 mM sample concentration).
Proton-proton couplings were obtained by means of one-dimensional and
two-dimensional NMR experiments and were used in a conformational analysis of
the sugar ring of each residue of the two compounds in the duplex form.  The
analysis indicated that all sugar rings display conformational flexibility in
the intact duplex:  population S-type sugar conformation ranges from 70% to
100%. A more refined analysis of the sugar rings of the parent compound
revealed a sequence-dependent variation of the sugar geometry.  This variation
does not follow well the trend predicted by the Calladine/Dickerson Sigma3-sum
rule [Dickerson, R.E.(1983).  J. Mol. Biol. 166, 419-441; Calladine, C.R.
(1982) J. Mol. Biol. 161, 343-352}; moreover the actual variations appear to be
smaller in solution than those expected on the basis of known X-ray structures.

<>

<1>Risser, R., Hopkins, N., Davis, R.W., Delius, H., Mulder, C.
<2>Action of Escherichia coli P1 restriction endonuclease on Simian Virus 40 DNA.
<3>J. Mol. Biol.
<4>89
<5>517-544
<6>1974
<7>The P1 restriction endonuclease prepared from a P1 lysogen of Escherichia coli makes one
double-strand break in simian virus (SV40) DNA.  In the presence of cofactors
S-adenosylmethionine and ATP the enzyme cleaves 70% of the closed circular SV40 DNA molecules
once to produce unit-length linear molecules and renders the remaining 30% resistant to
further cleavage.  No molecules were found by electron microscopy or by gel electrophoresis
that were cleaved more than once.  It would appear that the double-strand break is made by two
nearly simultaneous single-strand breaks, since no circular DNA molecules containing one
single-strand break were found as intermediates during the cleavage reaction.  The EcoP1
endonuclease-cleaved linear SV40 DNA molecules are not cleaved at a unique site, as shown by
the generation of about 65% circular molecules after denaturation and renaturation.  These
EcoP1 endonuclease-cleaved, renatured circular molecules are resistant to further cleavage by
EcoP1 endonuclease.  The EcoP1 endonuclease cleavage sites on SV40 DNA were mapped relative to
the partial denaturation map and to the EcoRI and HpaII restriction endonuclease cleavage
sites.  These maps suggest there are a minimum of four unique but widely space cleavage sites
at 0.09, 0.19, 0.52, and 0.66 SV40 units relative to the EcoRI site.  The frequency of
cleavage at any particular site differs from that at another site.  If S-adenosylmethionine is
omitted from the enzyme reaction mix, SV40 DNA is cleaved into several fragments.  An average
of 4.6 +/- 1 methyl groups are transferred to SV40 DNA from S-adenosylmethionine during the
course of a normal reaction containing the cofactors.  Under conditions which optimize this
methylation, 7 +/- 1 methyl groups can be transferred to DNA.  This methylation protects most
of the molecules from further cleavage.  he methyl groups were mapped relative to the
Hemophilus influenzae restriction endonuclease fragments.  The A fragment receives three to
four methyl groups and the B and G fragments each receive one to two methyl groups.  These
fragments correspond to those in which cleavage sites are located.

<>

<1>Ritchie, L., Podger, D.M., Hall, R.M.
<2>A mutation in the DNA adenine methylase gene (dam) of Salmonella typhimurium decreases susceptibility to 9-aminoacridine-induced frameshift mutagenesis.
<3>Mutat. Res.
<4>194
<5>131-141
<6>1988
<7>A mutant of Salmonella typhimurium with a reduced response to mutation
induction by 9-aminoacridine (9AA) has been isolated.  The mutation (dam-2) is
located in the DNA adenine methylase gene.  The dam-2 mutant strain exhibits a
level of sensitivity to 2-aminopurine (2AP) intermediate between that of the
dam+ and the DNA adenine methylation-deficit dam-1 strain, and 2AP sensitivity
was reversed by introduction of a mutH mutation or of the plasmid pMQ148 (which
carries a functional Escherichia coli dam+ gene).  However, the dam-2 strain is
not grossly defective in DNA adenine methylase activity.  Whole cell DNA
appears full methylated at -GATC- sites.  The levels of 9AA required to induce
equivalent levels of frameshift mutagenesis in the dam-2 strain were
approximately 2-fold higher than for the dam+ strain.  Introduction of pMQ148
dam+ reduced the level of 9AA required for induction of frameshift mutations
4-fold in the dam-2 strain and 2-fold in the dam+ strain.  The dam-2 mutation
had no effect on the levels of ICR191 required for induction of frameshift
mutations, but introduction of pMQ148 reduced the ICR191-induced mutagenesis
2-fold.  The dam+/pMQ148, dam-2/pMQ148 and dam-1/pMQ148 strains showed
identical dose-response curves for both 9AA and ICR191.  These results are
consistent with a slightly reduced (dam-2) or increased (pMQ148) rate of
methylation at the replication fork.  The 2AP sensitivity of the dam-2 strain
cannot be simply explained.  Furthermore, addition of methionine to the assay
medium reverses the 2AP sensitivity of the dam-2 strain, but has no effect on
9AA mutagenesis.

<>

<1>Ritchie, L.J., Hall, R.M., Podger, D.M.
<2>Mutant of Salmonella typhimurium LT2 deficient in DNA adenine methylation.
<3>J. Bacteriol.
<4>167
<5>420-422
<6>1986
<7>A mutant of Salmonella typhimurium LT2 deficient in methylation of the adenine residues in the
sequence 5'-GATC-3' was isolated.  The mutation (dam-1) was linked to the cysG locus, and
the properties of the mutant were similar to those of Escherichia coli dam mutants.  Reversion
of the hisC3076 frameshift marker by 9-aminoacridine was substantially enhanced by the dam-1
mutation, implying a direct role for adenine methylation in the prevention of frameshift
mutation induction.

<>

<1>Ritchot, N., Roy, P.H.
<2>DNA methylation in Neisseria gonorrhoeae and other Neisseriae.
<3>Gene
<4>86
<5>103-106
<6>1990
<7>It has been reported in the literature that Neisseria gonorrhoeae DNA is modified by the
methyltransferases (MTases) M.NgoI, M.NgoII, and M.NgoIII, as well as three other cytosine
MTases and one adenine MTase, even if the corresponding restriction endonucleases are not
present.  We envisioned the possibility of cloning one of the N. gonorrhoeae MTase-encoding
genes for use as a species-specific DNA probe.  We therefore undertook a survey of methylation
patterns of several clinical isolates of N. gonorrhoeae and N. meningitidis as well as ATCC
strains of other Neisseriae.  We found, from digestion patterns with isoschizomers, one N.
gonorrhoeae strain that lacked M.NgoII and two that lacked M.NgoIII.  All N. meningitidis
strains (save one) were resistant to digestion with NlaIV thus possessing an MTase like NgoV,
and one was resistant to SstII, thus having an NgoIII-like MTase.  None were resistant to
isoschizomers of NgoI, NgoIII and NgoIV.  Some other Neisseriae had an MTase with NlaIV (NgoV)
specificity, but none had NgoI, NgoIV, NgoII or NgoIII specificity, except for the
Branhamella-like N. caviae-ovis group and N. lactamica where these specificities were present
in at least one strain of this group.  Therefore, among the Neisseriae other than N. caviae
only M.NgoI is N. gonorrhoeae-specific.
[ The enzyme called NgoI in this abstract has been renamed NgoWI, Jan/1998. ]
[ The enzyme called NgoII in this abstract has been renamed NgoCII, Jan/1998. ]
[ The enzyme called NgoIII in this abstract has been renamed NgoKIII, Jan/1998. ]

<>

<1>Rival, A., Jaligot, E., Beule, T., Finnegan, E.J.
<2>Isolation and expression analysis of genes encoding MET, CMT, and DRM methyltransferases in oil palm (Elaeis guineensis Jacq.) in relation to the mantled somaclonal variation.
<3>J. Exp. Bot.
<4>59
<5>3271-3281
<6>2008
<7>In oil palm (Elaeis guineensis Jacq.), 5% of somatic embryo-derived regenerants show homeotic
changes during floral development, involving
an apparent feminization of male parts in flowers of both sexes, called
the 'mantled' phenotype. This variant phenotype is associated with a
reduction in the level of global DNA methylation. To explore possible
relationships between DNA methylation level and accumulation of
DNA-(cytosine-5) methyltransferase (DNMT) transcripts, the full-length
coding sequences corresponding to three different DNMT families in oil
palm, namely the MET, CMT, and DRM classes, have been isolated and
characterized. The corresponding genes were designated as EgMET1,
EgCMT1, and EgDRM1, and encode predicted polypeptides of 1543, 925, and
591 amino acid residues, respectively. Expression of oil palm DNMTs was
compared between normal and variant calli and in florescence tissues
using quantitative reverse-transcription PCR. A consistent increase in
transcript levels of EgMET1 and EgCMT1 was found in variant
fast-growing calli relative to nodular-compact calli. Nodular-compact
calli give rise to about 5% of abnormal regenerants whereas
fast-growing calli generate 95% of 'mantled' palms in their clonal
offspring and were previously demonstrated as having markedly
hypomethylated DNA. In immature abnormal in florescences only EgMET1
transcript levels were increased, while no changes in relative
abundance of the EgCMT1 or EgDRM1 transcripts were observed.

<>

<1>Rivera, D., Revale, S., Molina, R., Gualpa, J., Puente, M., Maroniche, G., Paris, G., Baker, D., Clavijo, B., McLay, K., Spaepen, S., Perticari, A., Vazquez, M., Wisniewski-Dye, F., Watkins, C., Martinez-Abarca, F., Vanderleyden, J., Cassan, F.
<2>Complete Genome Sequence of the Model Rhizosphere Strain Azospirillum brasilense  Az39, Successfully Applied in Agriculture.
<3>Genome Announcements
<4>2
<5>e00683-14
<6>2014
<7>We present the complete genome sequence of Azospirillum brasilense Az39, isolated from wheat
roots in the central region of Argentina and used as inoculant in
extensive and intensive agriculture during the last four decades. The genome
consists of 7.39 Mb, distributed in six replicons: one chromosome, three
chromids, and two plasmids.

<>

<1>Riveros-Mckay, F., Campos, I., Giles-Gomez, M., Bolivar, F., Escalante, A.
<2>Draft Genome Sequence of Leuconostoc mesenteroides P45 Isolated from Pulque, a Traditional Mexican Alcoholic Fermented Beverage.
<3>Genome Announcements
<4>2
<5>e01130-14
<6>2014
<7>Leuconostoc mesenteroides P45 was isolated from the traditional Mexican pulque beverage. We
report its draft genome sequence, assembled in 6 contigs consisting
of 1,874,188 bp and no plasmids. Genome annotation predicted a total of 1,800
genes, 1,687 coding sequences, 52 pseudogenes, 9 rRNAs, 51 tRNAs, 1 noncoding
RNA, and 44 frameshifted genes.

<>

<1>Roach, P.L., Wood, R.J., Maynard-Smith, M.D.
<2>Partially-methylated break light assay for DNA methyltransferase activity.
<3>International Patent Office
<4>WO 2008107711 A
<5>
<6>2008
<7>An assay for DNA methyltransferase activity wherein methylation at a single nucleotide
position in a partially methylated methylation site is coupled to preferential restriction
endonuclease cleavage of the substrate and detection of such cleavage, e.g. by means of a
fluorophore-quencher pair in a break light type assay.  Such an assay is favoured for
identification of inhibitors of DNA methyltransferases as it can be run in continuous format
and enable ready determination of Ki for any tested inhibitor.

<>

<1>Robbins, J.B., Smith, D., Belfort, M.
<2>Redox-Responsive Zinc Finger Fidelity Switch in Homing Endonuclease and  Intron Promiscuity in Oxidative Stress.
<3>Curr. Biol.
<4>21
<5>243-248
<6>2011
<7>It is well understood how mobile introns home to allelic sites, but how
they are stimulated to transpose to ectopic locations on an
evolutionary timescale is unclear [1]. Here we show that a group I
intron can move to degenerate sites under oxidizing conditions. The
phage T4 td intron endonuclease, I-Tevl, is responsible for this
infidelity. We demonstrate that I-Tevl, which promotes mobility and is
subject to autorepression [2] and translational control [3], is also
regulated posttranslationally by a redox mechanism. Redox regulation is
exercised by a zinc finger (ZF) in a linker that connects the catalytic
domain of I-Tevl to the DNA binding domain. Four cysteines coordinate
Zn2+ in the ZF, which ensures that I-Tevl cleaves its DNA substrate at
a fixed distance, 23-25 nucleotides upstream of the intron insertion
site [4]. We show that the fidelity of I-Tevl cleavage is controlled by
redox-responsive Zn2+ cycling. When the ZF is mutated, or after
exposure of the wild-type I-Tevl to H2O2, intron homing to degenerate
sites is increased, likely because of indiscriminate DNA cleavage.
These results suggest a mechanism for rapid intron dispersal, joining
recent descriptions of the activation of biomolecular processes by
oxidative stress through cysteine chemistry [5, 6].

<>

<1>Robbins, J.B., Stapleton, M., Stanger, M.J., Smith, D., Dansereau, J.T., Derbyshire, V., Belfort, M.
<2>Homing endonuclease I-TevIII: dimerization as a means to a double-strand break.
<3>Nucleic Acids Res.
<4>35
<5>1589-1600
<6>2007
<7>Homing endonucleases are unusual enzymes, capable of recognizing lengthy DNA sequences and
cleaving site-specifically within genomes. Many homing
endonucleases are encoded within group I introns, and such enzymes promote
the mobility reactions of these introns. Phage T4 has three group I
introns, within the td, nrdB and nrdD genes. The td and nrdD introns are
mobile, whereas the nrdB intron is not. Phage RB3 is a close relative of
T4 and has a lengthier nrdB intron. Here, we describe I-TevIII, the H-N-H
endonuclease encoded by the RB3 nrdB intron. In contrast to previous
reports, we demonstrate that this intron is mobile, and that this mobility
is dependent on I-TevIII, which generates 2-nt 3' extensions. The enzyme
has a distinct catalytic domain, which contains the H-N-H motif, and
DNA-binding domain, which contains two zinc fingers required for
interaction with the DNA substrate. Most importantly, I-TevIII, unlike the
H-N-H endonucleases described so far, makes a double-strand break on the
DNA homing site by acting as a dimer. Through deletion analysis, the
dimerization interface was mapped to the DNA-binding domain. The unusual
propensity of I-TevIII to dimerize to achieve cleavage of both DNA strands
underscores the versatility of the H-N-H enzyme family.

<>

<1>Roberts, G.A., Chen, K., Bower, E.K.M., Madrzak, J., Woods, A., Barker, A.M., Cooper, L.P., White, J.H., Blakely, G.W., Manfield, I., Dryden, D.T.F.
<2>Mutations of the domain forming the dimeric interface of the ArdA protein affect dimerization and antimodification activity but not antirestriction activity.
<3>FEBS J.
<4>280
<5>4903-4914
<6>2013
<7>ArdA antirestriction proteins are encoded by genes present in many conjugative plasmids and
transposons within bacterial genomes. Antirestriction is the ability to prevent cleavage of
foreign incoming DNA by restriction-modification (RM) systems. Antimodification, the ability
to inhibit modification by the RM system, can also be observed with some antirestriction
proteins. As these mobile genetic elements can transfer antibiotic resistance genes, the ArdA
proteins assist their spread. The consequenc of antirestriction is therefore the enhanced
dissemination of mobile genetic elements. ArdA proteins cause antirestriction by mimicking the
DNA structure bound by TypeI RM enzymes. The crystal structure of ArdA showed it to be a
dimeric protein with a highly elongated curved cylindrical shape [McMahon SA etal. (2009)
Nucleic Acids Res 37, 4887-4897]. Each monomer has three domains covered with negatively
charged side chains and a very small interface with the other monomer. We investigated the
role of the domain forming the dimer interface for ArdA activity via site-directed
mutagenesis. The antirestriction activity of ArdA was maintained when up to seven mutations
per monomer were made or the interface was disrupted such that the protein could only exist as
a monomer. The antimodification activity of ArdA was lost upon mutation of this domain. The
ability of the monomeric form of ArdA to function in antirestriction suggests, first, that it
can bind independently to the restriction subunit or the modificat ion subunits of the RM
enzyme, and second, that the many ArdA homologues with long amino acid extensions, present in
sequence databases, may be active in antirestriction.Structured digital abstract ArdA and ArdA
bind by molecular sieving (1, 2) ArdA and ArdA bind by cosedimentation in solution (1, 2)

<>

<1>Roberts, G.A., Chen, K., Cooper, L.P., White, J.H., Blakely, G.W., Dryden, D.T.
<2>Removal of a frameshift between the hsdM and hsdS genes of the EcoKI Type IA DNA  restriction and modification system produces a new type of system and links the  different families of Type I systems.
<3>Nucleic Acids Res.
<4>40
<5>10916-10924
<6>2012
<7>The EcoKI DNA methyltransferase is a trimeric protein comprised of two modification subunits
(M) and one sequence specificity subunit (S). This enzyme
forms the core of the EcoKI restriction/modification (RM) enzyme. The 3' end of
the gene encoding the M subunit overlaps by 1 nt the start of the gene for the S
subunit. Translation from the two different open reading frames is
translationally coupled. Mutagenesis to remove the frameshift and fuse the two
subunits together produces a functional RM enzyme in vivo with the same
properties as the natural EcoKI system. The fusion protein can be purified and
forms an active restriction enzyme upon addition of restriction subunits and of
additional M subunit. The Type I RM systems are grouped into families, IA to IE,
defined by complementation, hybridization and sequence similarity. The fusion
protein forms an evolutionary intermediate form lying between the Type IA family
of RM enzymes and the Type IB family of RM enzymes which have the frameshift
located at a different part of the gene sequence.

<>

<1>Roberts, G.A., Cooper, L.P., White, J.H., Su, T.J., Zipprich, J.T., Geary, P., Kennedy, C., Dryden, D.T.
<2>An investigation of the structural requirements for ATP hydrolysis and DNA cleavage by the EcoKI Type I DNA restriction and modification enzyme.
<3>Nucleic Acids Res.
<4>39
<5>7667-7676
<6>2011
<7>Type I DNA restriction/modification systems are oligomeric enzymes capable of switching
between a methyltransferase function on hemimethylated host
DNA and an endonuclease function on unmethylated foreign DNA. They have
long been believed to not turnover as endonucleases with the enzyme
becoming inactive after cleavage. Cleavage is preceded and followed by
extensive ATP hydrolysis and DNA translocation. A role for dissociation of
subunits to allow their reuse has been proposed for the EcoR124I enzyme.
The EcoKI enzyme is a stable assembly in the absence of DNA, so recycling
was thought impossible. Here, we demonstrate that EcoKI becomes unstable
on long unmethylated DNA; reuse of the methyltransferase subunits is
possible so that restriction proceeds until the restriction subunits have
been depleted. We observed that RecBCD exonuclease halts restriction and
does not assist recycling. We examined the DNA structure required to
initiate ATP hydrolysis by EcoKI and find that a 21-bp duplex with
single-stranded extensions of 12 bases on either side of the target
sequence is sufficient to support hydrolysis. Lastly, we discuss whether
turnover is an evolutionary requirement for restriction, show that the ATP
hydrolysis is not deleterious to the host cell and discuss how foreign DNA
occasionally becomes fully methylated by these systems.

<>

<1>Roberts, G.A., Houston, P.J., White, J.H., Chen, K., Stephanou, A.S., Cooper, L.P., Dryden, D.T., Lindsay, J.A.
<2>Impact of target site distribution for Type I restriction enzymes on the evolution of methicillin-resistant Staphylococcus aureus (MRSA) populations.
<3>Nucleic Acids Res.
<4>41
<5>7472-7484
<6>2013
<7>A limited number of Methicillin-resistant Staphylococcus aureus (MRSA) clones are responsible
for MRSA infections worldwide, and those of different lineages carry
unique Type I restriction-modification (RM) variants. We have identified the
specific DNA sequence targets for the dominant MRSA lineages CC1, CC5, CC8 and
ST239. We experimentally demonstrate that this RM system is sufficient to block
horizontal gene transfer between clinically important MRSA, confirming the
bioinformatic evidence that each lineage is evolving independently. Target sites
are distributed randomly in S. aureus genomes, except in a set of large
conjugative plasmids encoding resistance genes that show evidence of spreading
between two successful MRSA lineages. This analysis of the identification and
distribution of target sites explains evolutionary patterns in a pathogenic
bacterium. We show that a lack of specific target sites enables plasmids to evade
the Type I RM system thereby contributing to the evolution of increasingly
resistant community and hospital MRSA.

<>

<1>Roberts, G.A., Stephanou, A.S., Kanwar, N., Dawson, A., Cooper, L.P., Chen, K., Nutley, M., Cooper, A., Blakely, G.W., Dryden, D.T.
<2>Exploring the DNA mimicry of the Ocr protein of phage T7.
<3>Nucleic Acids Res.
<4>40
<5>8129-8143
<6>2012
<7>DNA mimic proteins have evolved to control DNA-binding proteins by competing with the target
DNA for binding to the protein. The Ocr protein of bacteriophage T7 is
the most studied DNA mimic and functions to block the DNA-binding groove of Type
I DNA restriction/modification enzymes. This binding prevents the enzyme from
cleaving invading phage DNA. Each 116 amino acid monomer of the Ocr dimer has an
unusual amino acid composition with 34 negatively charged side chains but only 6
positively charged side chains. Extensive mutagenesis of the charges of Ocr
revealed a regression of Ocr activity from wild-type activity to partial activity
then to variants inactive in antirestriction but deleterious for cell viability
and lastly to totally inactive variants with no deleterious effect on cell
viability. Throughout the mutagenesis the Ocr mutant proteins retained their
folding. Our results show that the extreme bias in charged amino acids is not
necessary for antirestriction activity but that less charged variants can affect
cell viability by leading to restriction proficient but modification deficient
cell phenotypes.

<>

<1>Roberts, M.D., Martin, N.L., Kropinski, A.M.
<2>The genome and proteome of coliphage T1.
<3>Virology
<4>318
<5>245-266
<6>2004
<7>The genome of enterobacterial phage T1 has been sequenced, revealing that
its 50.7-kb terminally redundant, circularly permuted sequence contains
48,836 bp of nonredundant nucleotides. Seventy-seven open reading frames
(ORFs) were identified, with a high percentage of small genes located at
the termini of the genomes displaying no homology to existing phage or
prophage proteins. Of the genes showing homologs (47%), we identified
those involved in host DNA degradation (three endonucleases) and T1
replication (DNA helicase, primase, and single-stranded DNA-binding
proteins) and recombination (RecE and Erf homologs). While the tail genes
showed homology to those from temperate coliphage N15, the capsid
biosynthetic genes were unique. Phage proteins were resolved by 2D gel
electrophoresis, and mass spectrometry was used to identify several of the
spots including the major head, portal, and tail proteins, thus verifying
the annotation.

<>

<1>Roberts, R.J.
<2>An amazing distortion in DNA induced by a methyltransferase (Nobel lecture).
<3>Angew. Chem. Int. Ed. Engl.
<4>33
<5>1222-1228
<6>1994
<7>
<>

<1>Roberts, R.J.
<2>Restriction enzymes.
<3>Molecular Genetic Analysis of Populations: A Practical Approach, IRL Press, Hoelzel, A.R., Oxford
<4>
<5>379-397
<6>1998
<7>A summary of the properties of the commercially available Type II restriction enzymes,
including digestion conditions. The information in this list is taken from Roberts, R.J. and
Macelis, D. (1996) 24, 223-235 plus updates from REBASE, the restriction enzyme database (URL
- http://www.neb.com/rebase).

<>

<1>Roberts, R.J.
<2>Analysis of Restriction Modification systems from whole genome sequences.
<3>Abstracts AAAS Ann. Mtg.
<4>166
<5>A37
<6>2000
<7>Until recently all of the 3200 restriction enzymes known to man had been found by obtaining
bacteria from culture collections or environ-mental samples and assaying them biochemically
and genetically.  During the last 15 years, many of these Restriction-Modification (RM)
systems have been cloned and sequenced and it is now possible to use quite so-phisticated
search algorithms to screen new DNA sequences for the presence of DNA methyltransferase genes.
Experience among known systems has shown that restriction enzyme genes always lie close to
their cognate methyltransferase genes.  Analysis of the bacterial and archaeal genome
sequences shows that methyltransferase genes are more common than one would have expected on
the basis of previous biochemical screening.  Frequently, they clearly form part of an RM
system, because the adjacent open reading frames show similarity to known restriction enzyme
genes.  Very often, though, the adjacent open reading frames have no homologs in GenBank and
become candidates either for restriction enzymes with novel specificities or for new examples
of previously uncloned specificities. We are developing methods to allow these candidate genes
quickly to be tested biochemically. Initial results are promising and it seems clear that
screening DNA sequence databases and websites will be a very productive method to find
restriction enzymes with new specificities.

<>

<1>Roberts, R.J.
<2>How restriction enzymes became the workhorses of molecular biology.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>5905-5908
<6>2005
<7>Restriction enzymes have proved to be invaluable for the physical mapping of DNA. They offer
unparalleled opportunities for diagnosing DNA sequence content and are used in fields as
disparate as criminal forensics and basic research. In fact, without restriction enzymes, the
biotechnology industry would certainly not have flourished as it has. The first experiments
demonstrating the utility of restriction enzymes were carried out by Danna and Nathans and
reported in 1971. This pioneering study set the stage for the modern practice of molecular
biology in which restriction enzymes are ubiquitous tools, although they are often taken for
granted.

<>

<1>Roberts, R.J.
<2>Inferring function from shotgun sequencing data.
<3>International Patent Office
<4>WO 2005121946 A
<5>
<6>2005
<7>Methods are described for detecting genes that encode toxic proteins using maps derived from
shotgun libraries by determining the presence of gaps in clone start sites on either side of
open reading frames.  The method is exemplified by identifying a previously unknown
restriction endonuclease gene.

<>

<1>Roberts, R.J.
<2>Inferring function from shotgun sequencing data.
<3>US Patent Office
<4>US 20060014179 A
<5>
<6>2006
<7>Methods are described for detecting genes that encode toxic proteins using maps derived from
shotgun libraries by determining the presence of gaps in clone start sites on either side of
open reading frames.  The method is exemplified by identifying a previously unknown
restriction endonuclease gene.

<>

<1>Roberts, R.J.
<2>An amazing distortion in DNA induced by a methyltransferase.
<3>Biosci. Rep.
<4>14
<5>103-117
<6>1994
<7>republication of the Nobel lecture

<>

<1>Roberts, R.J.
<2>Restriction endonucleases, DNA sequencing and computers.
<3>In Physics and Contemporary Needs, Plenum Press, Khan, A.M., Riazuddin, S., Qadir, A., Qazi, M.N., New York
<4>6
<5>305-316
<6>1984
<7>Among the 250 TypeII restriction endonucleases now characterized, there are
more than 70 different specificities and yet there is no indication that the
range of specificities is exhausted.  Indeed, there is good reason to believe
that hundreds, if not thousands, of different specificities would be found if a
diligent search were carried out.  One reason for this speculation is
illustrated in Table 1, which shows the range of sequence patterns with which
different Type II restriction endonucleases interact.  Among the simple
symmetric hexanucleotide sequences designated here as Class A, almost half of
the possible sequence patterns are already represented by well-characterized
enzymes.  There is no reason to believe that a similar number of enzymes will
not be found for the other patterns in Classes B through F.  Similarly, it
seems likely that enzymes recognizing degenerate patterns, like HgiAI and AccI,
are not the sole representatives of the class.  Within the last year alone,
five new classes (C,D,F,N., and O) were added to this list.

<>

<1>Roberts, R.J.
<2>Restriction endonucleases.
<3>Nucleases, Cold Spring Harbor Laboratory Press, Linn, S.M., Roberts, R.J., New York
<4>0
<5>311-340
<6>1982
<7>None

<>

<1>Roberts, R.J.
<2>Restriction endonucleases:  a new role in vivo?
<3>Nature
<4>271
<5>502
<6>1978
<7>Few enzymes have been exploited as thoroughly as the bacterial restriction
enzymes.  In recent years they have been instrumental in dramatic advances in
DNA sequence analysis, genetic engineering, and studies of gene structure.

<>

<1>Roberts, R.J.
<2>The Nobel Prizewinners 1978: Medicine.
<3>Nature
<4>275
<5>689-690
<6>1978
<7>The restriction endonucleases, which have become so familiar to the molecular
biologist, have finally come of age with the award of this year's Nobel Prize
in Physiology and Medicine to Dr. Werner Arber of the University of Basel and
to Drs. Daniel Nathans and Hamilton O. Smith of Johns Hopkins University.  They
each played a critical but separate role in drawing attention to these
bacterial enzymes which dominate so much present research.

<>

<1>Roberts, R.J.
<2>Restriction enzymes.
<3>Nucleic Acid Hybridisation: A Practical Approach, IRL Press, Hames, B.D., Higgins, S.J., Oxford, Washington DC
<4>0
<5>203-210
<6>1985
<7>Restriction enzymes are endodeoxyribonucleases that recognise short, specific
sequences within DNA molecules and then catalyse double-strand cleavage of the
DNA.  Three distinct classes of restriction enzymes are known:  Type I, Type
II, Type III.  In this Appendix, restriction enzymes and their isoschizomers
are listed alphabetically by prototype.  Their availability from three major
commercial sources is indicated, as are the buffer conditions recommended by
the manufacturer.  It should be noted that for most restriction enzymes, their
activity varies little over a wide range of ionic strength and pH, and the
values listed in general have not rigorously been shown to be optimal.  The
information in this list is taken from Roberts, R.J. Nucleic Acids Res. (1984)
12, r167-r204, and the New England Biolabs catalogue (1984 edition).

<>

<1>Roberts, R.J.
<2>Restriction enzymes.
<3>Molecular Genetic Analysis of Populations: A Practical Approach, Oxford University Press, Hoelzel, A.R., NY
<4>0
<5>281-296
<6>1992
<7>Restriction enzymes are endodeoxyribonucleases that recognize short, specific
sequences within DNA molecules and then catalyse double-strand cleavage of the
DNA.  Three distinct classes of restriction enzymes are known: (a) Type I
enzymes (b) Type II enzymes (c) Type III enzymes.  In this Appendix restriction
enzymes and their isoschizomers are listed alphabetically by prototype.  Their
availability from three major commercial sources is indicated, as are the
buffer conditions recommended by the manufacturer.  It should be noted that for
most restriction enzymes, their activity varies little over a wide range of
ionic strength and pH, and the values listed in general have not rigorously
been shown to be optimal.  The information in this list is taken from Roberts,
R.J., Nucleic Acids Res. (1984), 12, r167=t204, and the New England Biolabs
catalogue (1991 edition).

<>

<1>Roberts, R.J.
<2>Directory of restriction endonucleases.
<3>Methods Enzymol.
<4>65
<5>1-15
<6>1980
<7>This article is intended to serve as a directory to the restriction endonucleases which have
not been characterized.  All endonucleases which cleave DNA at a specific sequence have been
considered to be restriction enzymes, although in most cases there is no direct genetic
evidence for the presence of a host-controlled restriction-modification system.  Certain
strains are omitted from the table to save space.  Thus the many different Staphylococcus
aureus isolates which contain an isoschizomer of Sau3A are not listed individually.  Similarly
the many strains of gliding bacteria (orders: Myxobacterales and Cytophagales) which showed
evidence of specific endonucleases during a large-scale screening are still rather poorly
characterized.  Within the table the source of each microorganism is given either as an
individual or a National Culture Collection.  The enzymes are named in accordance with the
proposal of Smith and Nathans.  When two enzymes recognize the same sequence (i.e., are
isoschizomers), the prototype (i.e., the first example isolated) is indicated in parentheses
in column 3 of the table.  The recognition sequences (column 4 of the table) are abbreviated
so that only one strand, reading 5'-3', is indicated and the point of cleavage, when known,
is indicated by an arrow.  When two bases appear in parentheses, either one may appear at that
position within the recognition sequence.  Where known, the base modified by the corresponding
methylase is indicated by an asterisk.  A* is N6-methyladenosine; C* is 5-methylcytosine.  The
frequency of cleavage (columns five to eight) is experimentally determined for bacteriophage
lambda and adenovirus-2 DNAs, but represents the computer-derived values from the published
sequences of SV40 and PhiX174 DNAs.  When more than one reference appears (column 9 of the
table), the first contains the purification procedure for the restriction enzyme, the second
concerns its recognition sequence, the third contains the purification procedure for the
methylase, and the fourth describes its recognition sequence.  In some cases two references
appear in one of these categories when two independent groups have reached similar
conclusions.

<>

<1>Roberts, R.J.
<2>Directory of restriction endonucleases.
<3>Methods Enzymol.
<4>68
<5>27-41
<6>1979
<7>Table I is intended to serve as a directory to the restriction endonucleases
that have now been characterized.  In forming the list, all endonucleases that
cleave DNA at a specific sequence have been considered restriction enzymes,
although in most cases there is no direct genetic evidence for the presence of
a host-controlled restriction-modification system.  Certain strains have been
omitted from this list to save space.  Thus the many different Staphylococcus
aureus isolates containing an isoschizomer of Sau3A are not listed
individually.  Similarly the numerous strains of gliding bacteria (orders
Myxobacterales and Cytophagales) that showed evidence of specific endonucleases
during a large-scale screening are still rather poorly characterized.  Within
Table I the source of each microorganism is given either as an individual or a
national culture collection.  The enzymes are named in accordance with the
proposal of Smith and Nathans.

<>

<1>Roberts, R.J.
<2>Restriction endonucleases.
<3>Microbiology-1982, American Society for Microbiology, Schlessinger, D., Washington
<4>0
<5>5-9
<6>1978
<7>None

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Nucleic Acids Res.
<4>10
<5>r117-r144
<6>1982
<7>Since the last compilation of restriction endonucleases, 97 new entries have been added,
including 17 new specificities. Most notable among the new specificities is AhaIII (TTTAAA),
which is the first restriction enzyme to recognize only A/T base pairs. Other valuable new
specificities are ApaI (GGGCCC), AflII (CTTAAG), CfrI (PYGGCCPu), EcoRV (GATATC), FokI
(GGATG), HgiJII (GPuGCPyC), MluI (ACGCGT), NdeI (CATATG), NaeI (GCCGGC), NarI (GGCGCC), NcoI
(CCATGG), NruI (TCGCGA), NspBII (GCC/GGC), NspCI (PuCATGPy), ScrFI (CCNGG) and XmnI
(GAA[N]4TTC). Two entries have been removed, RruI and RruII, because the strain producing them
has been lost. Fortunately an isoschizomer of RruI has been found. This is ScaI (AGTACT).
Among the 355 enzymes listed, there are a minimum of 85 different specificities.

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Nucleic Acids Res.
<4>8
<5>r63-r80
<6>1980
<7>Since the last compilation of restriction endonucleases, more than 30 Type II
restriction endonucleases have been discovered, including some valuable new
specificities.  These include AcyI (GPuCGPyC), DdeI (CTNAG), Fnu4HI (GCNGC),
RsaI (GTAC), SphI (GCAGTC), and XmaIII (CGGCCG).  In addition, a number of new
isoschizomers have been discovered and further information about the
recognition sequences of some old entries is now available.  AvaX is renamed
AvaIII.

<>

<1>Roberts, R.J.
<2>Restriction enzymes and their isoschizomers.
<3>Nucleic Acids Res.
<4>18
<5>2331-2365
<6>1990
<7>A review

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Nucleic Acids Res.
<4>12
<5>r167-r204
<6>1984
<7>Since the last compilation of restriction endonucleases, 80 new entries have
been added, including 12 new specificities.  These are Cfr101 (PuCCGGPy),
Eco47III (AGCGCT), EcoA (GAG(N)7GTCA), MaeI (CTAG), MaeII (ACGT), MaeIII
(GTNAC), NlaIII (CATG), NlaIV (GGNNCC), NotI (GCGGCCGC), SnaBI (TACGTA), ScaI
(AGTACT) and SfiI (GGCCNNNNNGGCC).  NotI and SfiI are the first example of Type
II enzymes that recognize octanucleotide sequences.  In addition to these two
new sequence patterns, one additional new sequence pattern is recognized by
NlaIV.  The first example of unusual methylation is provided in the BcnI system
where the methylase protects by the formation of N4-methylcytosine.  Among the
475 enzymes listed, there are a minimum of 103 different specificities.  New
entries, together with new information about recognition sequences, are
indicated (@).  In forming this list, all endonucleases cleaving DNA at a
specific sequence have been considered to be restriction enzymes, although in
most cases there is no direct genetic evidence for the presence of a
restriction-modification system.  These endonucleases are named in accordance
with the proposal of Smith and Nathans.  Within the table, the source of each
microorganism is given either as an individual or a National Culture
Collection.  If further information is required, it can be found either in the
first reference which, in each case, refer to the purification procedure for
the restriction enzyme, or from the individuals who have provided their
unpublished results.  Where more than one reference appears, the second
concerns the recognition sequence for the restriction enzyme, the third
describes the purification procedure for the methylase and the fourth describes
the recognition sequence of the methylase.  In some cases, several references
appear in one of these categories when independent groups have reached similar
conclusions.

<>

<1>Roberts, R.J.
<2>Restriction endonucleases, DNA sequencing, and computers.
<3>Developmental Biology Using Purified Genes, Academic Press, Inc., Brown, D.D., 
<4>0
<5>621-634
<6>1981
<7>Among the 250 Type II restriction endonucleases now characterized, there are
more than 70 different specificities and yet there is no indication that the
range of specificities is exhausted.  Indeed, there is good reason to believe
that hundreds, if not thousands, of different specificities would be found if a
diligent search were carried out.

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Nucleic Acids Res.
<4>9
<5>r75-r96
<6>1981
<7>Since the last compilation of Type II restriction endonucleases, more than 45 new enzymes have
been discovered. Among the valuable new specificities are GdiI and its isoschizomer StuI
(AGGCCT), GdiII (PyGGCCG), HgiEII (ACC[N]6GGT), RruI (AGTACT), Tth111I and its isoschizomers
TtrI and TteI (GACNNNGTC), and Tth111II (CAAPuCA). The new enzyme NciI (CC[G/C]GG) turns out
to be an isoschizomer of CauII whose recognition has recently been determined. The recognition
sequences of SnaI (GTATAC) and SauI (CDTNAGG) have also been newly determined. Among the 258
enzymes listed, there are at least 69 different specificities. New entries, together with new
information about recognition sequences, are indicated. .

<>

<1>Roberts, R.J.
<2>The role of restriction endonucleases in genetic engineering.
<3>In: Recombinant Molecules: Impact on Science Society, Raven Press, Beers, R.F.Jr., Barrett, E.G., New York
<4>0
<5>21-32
<6>1977
<7>The class II restriction endonucleases have played a key role in the
development of recombinant DNA technology although, of the many enzymes now
available, only EcoRI and HindIII have been used extensively.  This chapter
describes some of the newly discovered restriction endonucleases which seem to
provide alternative possibilities for genetic engineering and suggests schemes
whereby the specificity of the nucleases can be exploited in the creation of
new recombinant genomes.

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Gene
<4>8
<5>329-343
<6>1980
<7>Since the last compilation of restriction endonucleases, more than 30 type II
restriction endonucleases have been discovered, including some with valuable
new specificities.  These include AcyI (GPuCGPyC), AsuII (TTCGAA), DdeI
(CTNAG), Fnu4HI (GCNGC), RsaI (GTAC), SphI (GCATGC) and XmaIII (CGGCCG).  In
addition, a number of new isoschizomers have been discovered and further
information about the recognition sequences of some old entries is now
available.  AvaX is renamed AvaIII.  In forming this list, all endonucleases
cleaving DNA at a specific sequence have been considered to be restriction
enzymes although, in most cases, there is no direct genetic evidence for the
presence of a restriction modification system.

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequence.
<3>Gene Amplif. Anal., Elsevier, Chirikjian, J.G., 
<4>5
<5>1-49
<6>1987
<7>Since the last published compilation of restriction endonucleases 130 new
enzymes have been discovered, including many new specificities.  Especially
noteworthy are the enzymes NotI (GCGGGCCGC) and SfiI (GGCCNNNNNGGCC) which are
the first Type II enzymes to recognize octanucleotide sequences.  They have the
useful property of cutting DNA sufficiently infrequently so that their sites
provide useful landmarks for mapping bacterial genomes and eukaryotic
chromosomes.  Among the 645 enzymes listed there are now a minimum of 137
different specificities.  In forming this list all endonucleases cleaving DNA
at a specific sequence have been considered to be restriction enzymes, although
in most cases there is no direct genetic evidence for the presenece of a
restriction-modification system.  These endonucleases are named in accordance
with the proposal of Smith and Nathans.  Within the table the source of each
microorganism is given either as an individual or a national culture
collection.  If further information is required it can be found either in the
first reference which in each case refers to the purification procedure for the
restriction enzyme, or from the individuals who have provided their unpublished
results.  Where more than one reference appears, the second concerns the
recognition sequence for the restriction enzyme, the third describes the
purification procedure for the methylase and the fourth describes the
recognition sequence of the methylase.  In some cases, several references
appear in one of these categories when independent groups have reached similar
conclusions.

<>

<1>Roberts, R.J.
<2>Restriction enzymes and their isoschizomers.
<3>Nucleic Acids Res.
<4>17
<5>r347-r387
<6>1989
<7>Since the last compilation of restriction enzymes (1), 156 new entries have
been added including 12 new specificities.  With the growing size of this
database and the recognition that the most widespread use of the information is
as a database for computer programs predicting restriction enzyme cleavage
patterns, the new format has been continued.  This format is intended to
contain the minimal amount of information required by a computer program.  It
should be noted that only enzymes for which the recognition sequence is known
are included.  This new list is shown in the first Table, while an alphabetical
listing of all Type II enzymes is presented in the second Table.  A copy of the
restriction enzyme data base in its previous format (2), including enzymes of
unknown recognition sequence, will be available upon request.  It should also
be noted that an alternative compilation of these enzymes has recently been
produced (3).

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Nucleic Acids Res.
<4>11
<5>r135-r167
<6>1983
<7>Since the last compilation of restriction endonucleases, 43 new entries have
been added, including 6 new specificities.  These are AatII (GACGTC), AflIII
(ACPuPyGT), BinI (GGATC), EcopDXI (ATCA(N)^ATTC), NspBII (C(A/C)GC(T/G)G and
SduI (G(G/A/T)GC(C/A/T)C).  EcopDXI is the first example of a Type I enzyme
that recognizes an octanucleotide sequence.  Both NspBII and SduI recognize
sequence patterns that have not been described before.

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Nucleic Acids Res.
<4>13
<5>r165-r200
<6>1985
<7>Since the last compilation of restriction endonucleases forty-nine new entries
have been added, including nine new specificities, these are DraII (PuGGNCCPy),
DraIII (CACNNNGTG), EcoD (TTA(N)7GTCPy), EspI (GCTNAGC), NheI (GCTAGC), RsrII
(CGG(A/T)CCG), StyI (CC(A/T)(A/T)GG), SspI (AATATT) and SpeI (ACTAGT).  In
addition, the enzyme Asp718 is an interesting isoschizomer of KpnI.  It cleaves
the recognition sequence to leave a 5' terminal extension instead of the 3'
terminal extension left by KpnI.

<>

<1>Roberts, R.J.
<2>Restriction enzymes and their isoschizomers.
<3>Nucleic Acids Res.
<4>16
<5>r271-r313
<6>1988
<7>Since the last compilation of restriction enzymes, 156 new entries have been
added including 12 new specificities.  With the growing size of this database
and the recognition that the most widespread use of the information is as a
database for computer programs predicting restriction enzyme cleavage patterns,
the new format has been continued.  This format is intended to contain the
minimal amount of information required by a computer program.  It should be
noted that only enzymes for which the recognition sequence is known are
included.  This new list is shown in the first Table, while an alphabetical
listing of all Type II enzymes is presented in the second Table.  A copy of the
restriction enzyme data base in its previous format, including enzymes of
unknown recognition sequence, will be available upon request.  It should also
be noted that an alternative compilation of these enzymes has recently been
produced.  The database shown in these Tables is available online through the
BIONET computer resource.  A version corresponding to the printed text is
located in the file <ROBERTS>RESTRICT.NAR several alternative versions are
available and are documented in <ROBERTS>RESTRICT.DOC  In forming this list,
all endonucleases cleaving DNA at a specific sequence have been considered to
be restriction enzymes, although in most cases there is no direct genetic
evidence for the presence of a restriction-modification system.  The
endonucleases are named in accordance with the proposal of Smith and Nathans.
Several enzymes appear in this list with revised names.  These revisions were
made to avoid confusion with existing enzymes or to increase the uniformity of
the names.  In each case the name changes were made with the approval of the
appropriate authors.

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>In Handbook of Biochemistry and Molecular Biology, CRC Press, Fasman, G.D., Ohio
<4>0
<5>532-534
<6>1976
<7>None

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>DNA Insertion Elements, Plasmids, and Episomes., Cold Spring Harbor Laboratory Press, Bukhari, A.I., Shapiro, J.A., Adhya, S.L., New York
<4>0
<5>757-768
<6>1977
<7>None

<>

<1>Roberts, R.J.
<2>Restriction endonucleases.
<3>CRC Crit. Rev. Biochem.
<4>4
<5>123-164
<6>1976
<7>This review provides a comprehensive account of the current status of the biology and
biochemistry of restriction endonucleases. Both Class I and Class II restriction endonucleases
will be considered. However, emphasis will be placed on the Class II group, which recognizes
and cleaves a specific duplex DNA sequence. Their occurrence, purification, and
characterization is discussed in detail. The characterization includes physical mapping
information and determination of recognition sequences. In addition to detailed discussions of
the biochemical properties of the enzymes, considerable attention is paid to the uses of these
enzymes as tools for research in molecular biology. These uses include physical mapping of
genomes and their transcripts, genetic analysis (marker rescue, etc.), DNA sequence analysis,
analysis of complex genomes, and genetic engineering. Specific examples of each use are
outlined. Practical aspects of both the isolation and use of the restriction endonucleases
form the major theme of this review.

<>

<1>Roberts, R.J.
<2>Restriction and modification enzymes and their recognition sequences.
<3>Gene
<4>4
<5>183-193
<6>1978
<7>During the last few years many bacterial strains have been examined for the
presence of Type II restriction endonucleases and a large number of these
enzymes have now been characterized.  Much of the information available has
never been formally published.  While this reflects the lengthy time which can
elapse between discovery and publication, increasingly it results from the fact
that a newly discovered endonuclease is an isoschizomer of a more familiar one.
Thtus, unless the new source offers some advantage, there is a natural trend
to avoid formal publication.  To some extent, review articles fill this gap;
however, they quickly become outdated.  The present compilation is an attempt
to extend current awareness of the enzymes now available.

<>

<1>Roberts, R.J.
<2>Restriction enzymes and their isoschizomers.
<3>Nucleic Acids Res.
<4>15
<5>r189-r217
<6>1987
<7>Since the last compilation of restriction enzymes, 251 new entries have been
added including 21 new specificities.  With the growing size of this database
and the recognition that the most widespread use of the information is as a
database for computer programs predicting restriction enzyme cleavage patterns,
a new format has been adopted.  This new format is intended to contain the
minimal amount of information required by a computer program.  It should be
noted that only enzymes for which the recognition sequence is known are
included.

<>

<1>Roberts, R.J.
<2>Hans Krebs Lecture Bacterial methylomes.
<3>FEBS J.
<4>280
<5>0
<6>2013
<7>Bacterial DNA methyltransferases are best known as orphan enzymes such as the Dam methylase of
E. coli or as components of restriction-modification systems.  Until recently, rigorously
determining the specificity of MTases has been a tedious process.  When they were components
of Type II restriction systems it has been assumed that the MTases would have the same
specificity as the cognate restriction enzyme.  For Type I and Type III RM systems specificity
determination was rarely attempted.  With the advent of SMRT sequencing from Pacific
Biosciences this situation has changed dramatically.  Now it has become very simple to
determine MTase recognition sequences both for individual MTases cloned in plasmids and also
for whole bacterial genomes.  This offers new insights into the functioning of bacteria and
has led to the discovery of several novel MTases with unexpected properties.

<>

<1>Roberts, R.J. et al.
<2>A nomenclature for restriction enzymes, DNA methyltransferases, homing endonucleases and their genes.
<3>Nucleic Acids Res.
<4>31
<5>1805-1812
<6>2003
<7>A nomenclature is described for restriction endonucleases, DNA methyltransferases, homing
endonucleases and related genes and gene
products. It provides explicit categories for the many different Type II
enzymes now identified and provides a system for naming the putative genes
found by sequence analysis of microbial genomes.

<>

<1>Roberts, R.J. et al.
<2>A nomenclature for restriction enzymes, DNA methyltransferases, homing endonucleases, and their genes.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>1-18
<6>2004
<7>There are three main groups of restriction endonucleases (REases) called Types I, II, and III.
Since 1973, REases and DNA methyltransferases (MTases) have been named based on an original
suggestion by Smith and Nathans.  They proposed that the enzyme names should begin with a
three-letter acronym in which the first letter was the first letter of the genus from which
the enzyme was isolated and the next two letters were the first two letters of the species
name.  Extra letters or numbers could be added to indicate individual strains or serotypes.
Thus, the enzyme HindII was one of four enzymes isolated from Haemophilus influenzae serotype
d.  The first three letters of the name were italicized.  Later, a formal proposition for
naming the genes encoding REases and MTases was adopted.  When there were only a handful of
enzymes known, these schemes were very useful, but as more enzymes have been found, often from
different genera and species with names whose three-letter acronyms would be identical,
considerable laxity in naming conventions has appeared.  In addition, we now know that each
major type of enzyme can contain subtypes.  This especially applies to the Type II enzymes, of
which more than 3500 have been characterized.  In this paper we revisit the naming conventions
and outline an updated scheme that incorporates current knowledge about the complexities of
these enzymes.  We describe a set of naming conventions for REases and their associated
MTases.  Since the homing endonucleases have been named in an analogous fashion, we proposed
that similar guidelines be applied to that group of enzymes.  Finally, it is important to
realize that the aim of this document is to provide a nomenclature for these enzymes, not to
provide a rigorous classification.

<>

<1>Roberts, R.J., Breitmeyer, J.B., Tabachnik, N.F., Myers, P.A.
<2>A second specific endonuclease from Haemophilus aegyptius.
<3>J. Mol. Biol.
<4>91
<5>121-123
<6>1975
<7>A second restriction-like endonuclease has been partially purified from
Haemophilus aegyptius.  This enzyme cleaves bacteriophage lambda DNA and
adenovirus 2 DNA at many sites, but cleaves simian virus 40 DNA at only one
site.

<>

<1>Roberts, R.J., Byrd, D.R., Morgan, R.D., Patti, J., Noren, C.J.
<2>Method for screening restriction endonucleases.
<3>International Patent Office
<4>WO 9911821
<5>
<6>1999
<7>A method is provided for identifying a restriction endonuclease, which includes the steps of
(a) screening a target DNA sequence for the presence of known methylase sequence motifs, (b)
identifying any open reading frames which lie close to the methylase sequence motifs screened
in step (a), and (c) assaying the protein products of these open reading frames for
restriction endonuclease activity.  Methods for identifying isoschizomers of known restriction
endonucleases, which isoschizomers possess a desired physical property, such as
thermostability, are also provided by the present invention, as are several novel restriction
endonucleases isolated from M. jannaschii, MjaIII and MjaIV.  Additionally, a gene was
identified that encoded a previously observed endonuclease activity, designated MjaII.  Also
provided by the present invention are vectors suitable for cloning a DNA sequence encoding a
cytotoxic protein, via independent transcription promoters which may be selectively controlled
by several conditions.  A method for producing these cytotoxic proteins using such vectors is
also provided, as are stable clones of PacI and NlaIII.

<>

<1>Roberts, R.J., Byrd, D.R., Morgan, R.D., Patti, J., Noren, C.J.
<2>Method for screening restriction endonucleases.
<3>US Patent Office
<4>US 6383770
<5>
<6>2002
<7>A method is provided for identifying a restriction endonuclease, which includes the steps of
(a) screening a target DNA sequence for the
presence of known methylase sequence motifs, (b) identifying any open
reading frames which lie close to the methylase sequence motifs
screened in step (a), and (c) assaying the protein products of these
open reading frames for restriction endonuclease activity. Methods for
identifying isoschizomers of known restriction endonucleases, which
isoschizomers possess a desired physical property, such as
thermostability, are also provided by the present invention, as are
several novel restriction endonucleases isolated from M. jannaschii,
MjaIII and MjaIV. Additionally, a gene was identified that encoded a
previously observed endonuclease activity, designated MjaII. Also
provided by the present invention are vectors suitable for cloning a
DNA sequence encoding a cytotoxic protein, via independent
transcription promotors which may be selectively controlled by several
conditions. A method for producing these cytotoxic proteins using such
vectors is also provided, as are stable clones of PacI and NlaIII.

<>

<1>Roberts, R.J., Byrd, D.R., Morgan, R.D., Patti, J., Noren, C.J.
<2>Method for screening restriction endonucleases from Methanococcus jannaschii.
<3>US Patent Office
<4>US 20030119027 A
<5>25
<6>2003
<7>A method is provided for identifying a restriction endonuclease, which includes the steps of
(a) screening a target DNA sequence for the presence of known methylase sequence motifs, (b)
identifying any open reading frames which lie close to the methylase sequence motifs screened
in step (a), and (c) assaying the protein products of these open reading frames for
restriction endonuclease activity.  Methods for identifying isoschizomers of known restriction
endonucleases, which isoschizomers possess a desired physical property, such as
themostability, are also provided by the present invention, as are several novel restriction
endonucleases isolated from M. jannaschii, MjaIII and MjaIV.  Additionally, a gene was
identified that encoded a previously observed endonuclease activity, designated MjaII.  Also
provided by the present invention are vectors suitable for cloning a DNA sequence encoding a
cytotoxic protein, via independent transcription promoters which may be selectively controlled
by several conditions.  A method for producing these cytotoxic proteins using such vectors is
also provided, as are stable clones of PacI and NlaIII.

<>

<1>Roberts, R.J., Byrd, D.R., Morgan, R.D., Patti, J., Noren, C.J.
<2>Method for screening restriction endonucleases based on database homology searching of cognate DNA methylase sequences.
<3>US Patent Office
<4>US 6689573 B
<5>
<6>2004
<7>A method is provided for identifying a restriction endonuclease that includes: screening a
target DNA sequence for the presence of known methylase sequence motifs, identifying any open
reading frames which lie close to the screened methylase sequence motif and assaying the
protein products of the open reading frames for restriction endonuclease activity.

<>

<1>Roberts, R.J., Byrd, D.R., Morgan, R.D., Patti, J., Noren, C.J.
<2>Method for screening restriction enzymes.
<3>US Patent Office
<4>US 6905837 B
<5>
<6>2005
<7>A method is provided for identifying a restriction endonuclease, which includes the steps of
(a) screening a target DNA sequence for the presence of known methylase sequence motifs, (b)
identifying any open reading frames which lie close to the methylase sequence motifs screened
in step (a), and (c) assaying the protein products of these open reading frames for
restriction endonuclease activity.  Methods for identifying isoschizomers of known restriction
endonucleases, which isoschizomers possess a desired physical property, such as
thermostability, are also provided by the present invention, as are several novel restriction
endonucleases isolated from M. jannaschii, MjaIII and MjaIV.  Additionally, a gene was
identified that encoded a previously observed endonuclease activity, designated MjaII.  Also
provided by the present invention are vectors suitable for cloning a DNA sequence encoding a
cytotoxic protein, via independent transcription promoters which may be selectively controlled
by several conditions.  A method for producing these cytotoxic proteins using such vectors is
also provided, as are stable clones of PacI and NlaIII.

<>

<1>Roberts, R.J., Cheng, X.
<2>Base flipping.
<3>Annu. Rev. Biochem.
<4>67
<5>181-198
<6>1998
<7>Base flipping is the phenomenon whereby a base in normal B-DNA is swung completely out of the
helix into an extrahelical position.  It was discovered in 1994 when the first co-crystal
structure was reported for a cytosine-5 DNA methyltransferase binding to DNA.  Since then it
has been shown to occur in many systems where enzymes need access to a DNA base to perform
chemistry on it.  Many DNA glycosylases that remove abnormal bases from DNA use this
mechanism.  This review describes systems known to use base flipping as well as many systems
where it is likely to occur but has not yet been rigorously demonstrated.  The mechanism and
evolution of base flipping are also discussed.  A review.

<>

<1>Roberts, R.J., Halford, S.E.
<2>Type II restriction enzymes.
<3>Nucleases, Cold Spring Harbor Laboratory Press, Linn, S.M., Lloyd, R.S., Roberts, R.J., Cold Spring Harbor
<4>0
<5>35-88
<6>1993
<7>
    I. Introduction and history
   II. Recognition sequences and cleavage properties
            A. Type IIs enzymes
            B. Degenerate recognition sequences
            C. Unusual type II enzymes
            D. Determination of cleavage sites
            E. Effects of methylation
            F. Single-stranded DNA cleavage
  III. Genes and their organization
            A. Cloning
            B. Genetic location
            C. Sequences
   IV. DNA Binding
            A. Enzymes that bind specifically to their recognition sites
            B. Enzymes that fail to bind specifically to their recognition sites
            C. Transfer to recognition sites
    V. DNA Cleavage
            A. Plasmid substrates
            B. Oligonucleotide substrates
            C. Specificity
   VI. Crystallography
            A. Protein structures
            B. DNA structures
            C. DNA-protein interfaces
  VII. Phosphodiester hydrolysis
 VIII. DNA recognition functions
            A. Altered enzymes
            B. Altered substrates
            C. Coupling recognition to catalysis
   IX. Evolution
    X. Conclusions and future prospects


<>

<1>Roberts, R.J., Macelis, D.
<2>Restriction enzymes and their isoschizomers.
<3>Nucleic Acids Res.
<4>20
<5>2167-2180
<6>1992
<7>The restriction enzyme database, REBASE, contains information about restriction enzymes and
their associated methylases. Since the last description of the contents of REBASE, 204 new
entries have been added including 5 new Type II enzymes and 4 new Type I enzymes. A complete
list of these new enzymes can be found in Table I. A total of 2103 restriction enzymes are now
known and include 17 different Type I specificities, 179 different Type II specificities and 4
different Type III specificities. Table II contains a listing of all prototype restriction
enzymes (Types I, II and III), together with their commercially available isoschizomers and
neoschizomers that cleave at a position different from their prototype.

<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE-restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>21
<5>3125-3137
<6>1993
<7>The restriction enzyme database, REBASE, is a collection of information about restriction
enzymes and methylases. Since the last description of the contents of REBASE (1), 265 new
entries have been added including 8 new Type II enzymes: AclI, AA^CGTT; Bce83I, CTTGAG
(16/14); BscGI, CCCGT; BseRI, GAGGAG (10/8); Bsp1407I, T^GTACA; BspLU11I, A^CATGT; BsrDI,
GCAATG (2/0) and SexAI A^CCWGGT. A complete list of these new enzymes can be found in Table I.
A total of 2393 restriction enzymes is now known including 17 different Type I specificities,
188 different Type II specificities and 4 different Type II specificities. Table II contains a
listing of all prototype restriction enzymes (Types I, II and III), together with their
commercially available isoschizomers and neoschizomers that cleave at a position different
from their prototype.

<>

<1>Roberts, R.J., Macelis, D.
<2>The restriction enzymes.
<3>Nucleases, Cold Spring Harbor Laboratory Press, Linn, S.M., Lloyd, R.S., Roberts, R.J., Cold Spring Harbor
<4>0
<5>439-444
<6>1993
<7>
<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE - restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>22
<5>3628-3639
<6>1994
<7>REBASE is a comprehensive database of information about restriction enzymes and their
associated methylases, including their recognition and cleavage sites and their commercial
availability. Information from REBASE is available via monthly electronic mailings as well as
via WAIS and anonymous ftp. Specialized files are available that can be used directly by many
software packages.

<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE--restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>24
<5>223-235
<6>1996
<7>REBASE is a comprehensive database of information about restriction enzymes and their
associated methylases, including their recognition and cleavage sites and their commercial
availability.  Information from REBASE is available via monthly electronic mailings as well as
via WAIS, anonymouse ftp and through the World Wide Web (htp://www.neb.com/rebase).
Specialized files are available that can be used directly by many software packages.

<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE-restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>25
<5>248-262
<6>1997
<7>REBASE is a comprehensive database of information about restriction enzymes and their
associated methylases, including their recognition and cleavage sites and their commercial
availability.  Information from REBASE is available via monthly electronic mailings as well as
via anonymous ftp, WAIS/gopher and through the World Wide Web (http://www.neb.com/rebase).
Specialized files are available that can be used directly by many software packages.

<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE - restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>26
<5>338-350
<6>1998
<7>REBASE is a comprehensive database of information about restriction enzymes and their
associated methylases, including their recognition and cleavage sites and their commercial
availability. Also included is a listing of homing endonucleases. Information from REBASE is
available via monthly electronic mailings as well as via anonymous ftp and through the World
Wide Web. The REBASE web site, http://www.neb.com/rebase , is where we maintain a web page for
every enzyme, reference and supplier. Additionally, there is a search facility, help and NEWS
pages, and a complete description of our various services. Specialized files are available
that can be used directly by many software packages.

<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE-restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>27
<5>312-313
<6>1999
<7>REBASE is a comprehensive database of information about restriction enzymes and their
associated methylases, including their recognition and cleavage sites and their commercial
availability.  Also included is a listing of homing endonucleases.  Information from REBASE is
distributed via monthly electronic mailings as well as through anonymous ftp and the World
Wide Web.  The REBASE web site (http://www.neb.com/rebase) contains a web page for every
enzyme, reference and supplier.  Additionally, there is a search facility, help and NEWS
pages, and a complete description of our various services.  Specialized files are available
that can be used directly by many software packages.

<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE - restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>28
<5>306-307
<6>2000
<7>REBASE is a comprehensive database of information about restriction enzymes and related
proteins. It contains published and unpublished references, recognition and cleavage sites,
isoschizomers, commercial availability, methylation sensitivity, crystal and sequence data.
DNA methyltransferases, homing endonucleases, nicking enzymes, specificity subunits and
control proteins are also included. Most recently, putative DNA methyltransferases and
restriction enzymes, as predicted from analysis of genomic sequences, are also listed. The
data is distributed via Email, ftp (ftp.neb.com), and the Web (http://rebase.neb.com).

<>

<1>Roberts, R.J., Macelis, D.
<2>REBASE-restriction enzymes and methylases.
<3>Nucleic Acids Res.
<4>29
<5>268-269
<6>2001
<7>REBASE contains comprehensive information about restriction enzymes, DNA methylases and
related proteins such as nicking enzymes, specificity subunits and control proteins.
It contains published and unpublished references, recognition and cleavage sites,
isoschizomers, commercial availability, methylation sensitivity, crystal data and
sequence data. Homing endonucleases are also included. Most recently, extensive
information about the methylation sensitivity of restriction enzymes has been added
and a new feature contains complete analyses of the putative restriction systems in
the sequenced bacterial and archaeal genomes. The data is distributed via email,
ftp (ftp.neb.com) and the Web (http://rebase.neb.com).

<>

<1>Roberts, R.J., Macelis, D.
<2>Restriction enzymes and their isoschizomers.
<3>Nucleic Acids Res.
<4>19
<5>2077-2109
<6>1991
<7>A review of all known restriction enzymes and a description of the REBASE
database.

<>

<1>Roberts, R.J., Meyertons, J.L., Lechevalier, M.P.
<2>Restriction endonuclease FseI.
<3>International Patent Office
<4>WO 9101371
<5>
<6>1991
<7>A novel Type II restriction endonuclease isolated from Frankia species DDB13250120 is
provided. The enzyme, designated FseI, recognizes the octanucleotide sequence, GGCCGG^CC, and
cleaves the DNA at the sites indicated by the arrows.

<>

<1>Roberts, R.J., Meyertons, J.L., Lechevalier, M.P.
<2>Restriction endonuclease FseI.
<3>US Patent Office
<4>US 5061628
<5>
<6>1991
<7>A novel type II restriction endonuclease isolated from Frankia species DDB13250120 is
provided.  The enzyme, designated FseI, recognizes the octanucleotide sequence 5'
GGCCGG/CC3' 3'CC/GGCCGG5' and cleaves adenovirus 2 DNA at the sites indicated by the
arrows.

<>

<1>Roberts, R.J., Myers, P.A., Morrison, A., Murray, K.
<2>A specific endonuclease from Arthrobacter luteus.
<3>J. Mol. Biol.
<4>102
<5>157-165
<6>1976
<7>A new restriction-like endonuclease, AluI, has been partially purified from Arthrobacter
luteus. This enzyme cleaves bacteriophage lambda DNA, adenovirus-2 DNA and simian virus 40 DNA
at may sites including all sites cleaved by the endonuclease HindIII from Haemophilus
influenzae serotype d. Radioactive oligonucleotides in pancreatic DNAase digests of
(5'-32P)-labelled fragments of phage lambda DNA relased by the action of AluI had the 5'
terminal sequence pC-T-N-. The enzyme recognizes the tetranucleotide sequence
3'-T-C-^-G-A-5' 5'-A-G-^C-T-3' and cleaves it at the position marked by the arrows.

<>

<1>Roberts, R.J., Myers, P.A., Morrison, A., Murray, K.
<2>A specific endonuclease from Haemophilus haemolyticus.
<3>J. Mol. Biol.
<4>103
<5>199-208
<6>1976
<7>A restriction-like endonuclease, HhaI, has been partially purified from
Haemophilus haemolyticus.  This enzyme cleaves bacteriophage lambda DNA and
adenovirus-2 DNA at many sites, and cleaves simian virus 40 DNA at only two
sites.  It recognizes the sequence 5'-G-C-G-^C-3' 3'-C-^G-C-G-5' and cuts at
the sites indicated by the arrows.

<>

<1>Roberts, R.J., Stickel, S.
<2>Cleavage of RNA by restriction endonucleases.
<3>International Patent Office
<4>WO 2005040342 A
<5>
<6>2005
<7>Methods of and uses for cleaving RNA/DNA duplexes with restriction endonucleases are provided
as well as methods for determining whether a restriction endonuclease is capable of such
cleavage.

<>

<1>Roberts, R.J., Stickel, S.
<2>Cleavage of RNA by restriction endonucleases.
<3>US Patent Office
<4>US 20050053990
<5>
<6>2005
<7>NOVELTY - Cleaving a RNA/DNA duplex comprises combining a restriction endonuclease,
isoschizomer or its modification
with a RNA/DNA duplex in a mixture and cleaving the RNA/DNA duplex. The
restriction endonuclease, isoschizomer or its modification is capable
of cleaving the RNA/DNA duplex to form RNA/DNA duplex fragments of
specific sizes with defined ends. DETAILED DESCRIPTION - INDEPENDENT
CLAIMS are also included for: (1) determining whether a restriction
endonuclease is capable of cleaving a RNA within a RNA/DNA duplex; (2)
detecting a pathogenic RNA virus; (3) treating a subject infected with
a RNA-containing virus to reduce viral load; (4) obtaining a
double-stranded RNA (dsRNA) fragment having a defined length and
terminal sequence; (5) gene silencing; (6) mapping a long RNA molecule;
(7) detecting alternative spliced forms of messenger RNAs (mRNAs); (8)
generating RNA primers for DNA polymerase or reverse transcriptase; and
(9) RNA sequence shuffling for expressing a novel protein.
BIOTECHNOLOGY - Preferred Method: Cleaving a RNA/DNA duplex comprises
combining a restriction endonuclease, isoschizomer or its modification
with an RNA/DNA duplex in a mixture and cleaving the RNA/DNA duplex. The
restriction endonuclease, isoschizomer or its modification is capable
of cleaving the RNA/DNA duplex to form RNA/DNA duplex fragments of
specific sizes with defined ends. The restriction endonuclease is a
modified restriction endonuclease that selectively cleaves RNA in the
RNA/DNA duplexes without substantial cleavage of double-stranded DNA
(dsDNA). The mixture further comprises metal ions other than magnesium
for inhibiting DNA duplex cleavage by the restriction endonuclease. The
restriction endonuclease recognizes a specific sequence on the RNA/DNA
duplex, so that the size of the RNA/DNA duplex is at least 2
nucleotides longer than the recognition sequence. The method further
comprises denaturing the duplex to form single-stranded RNA (ssRNA)
fragments of defined size and ends. The restriction endonuclease is
AvaII, Cac8I, BtsI, SfaNI or Sau3AI. Determining whether a restriction
endonuclease is capable of cleaving a RNA within a RNA/DNA duplex
comprises: (1) obtaining a labeled RNA/DNA oligonucleotide duplex; (2)
cleaving the RNA/DNA duplex with a restriction endonuclease; and (3)
analyzing the products of the reaction by size separation to determine
whether the restriction endonuclease is capable of cleaving the RNA in
the duplex in the absence of ribonuclease activity. The restriction
endonuclease has a known DNA cleavage specificity under standard
reaction conditions. Detecting a pathogenic RNA virus comprises: (1)
hybridizing viral RNA in a biological sample with a ssDNA fragment; (2)
cleaving the RNA/DNA duplex with one or more restriction endonucleases,
isoschizomers or their modifications having known recognition and
cleavage specificities; (3) denaturing the RNA/DNA duplex to produce
RNA having characteristic fragment sizes in a RNA profile; and (4)
detecting the pathogenic RNA virus from the RNA profile. Treating a
subject infected with a RNA-containing virus to reduce viral load
comprises administering one or more restriction endonucleases,
isoschizomers or their modifications in a pharmaceutical formulation.
The viral pathogen is HIV. Obtaining a double-stranded RNA (dsRNA)
fragment having a defined length and terminal sequence comprises: (1)
cleaving a RNA/DNA duplex with a restriction endonuclease, isoschizomer
or its modification having known cleavage specificity; (2) denaturing
the cleaved RNA/DNA duplex so that the RNA hybridizes to itself or a
second RNA to form a RNA/DNA duplex; and (3) obtaining the dsRNA
fragment having a defined length and terminal sequence. The method of
gene silencing comprises: (1) cleaving a RNA/DNA duplex with one or
more restriction endonucleases, isoschizomers or their modifications;
(2) denaturing the cleaved RNA/DNA duplex to provide a ssRNA; (3)
permitting the ssRNA to reanneal into a hairpin or RNA duplex

<>

<1>Roberts, R.J., Vincze, T., Posfai, J., Macelis, D.
<2>REBASE: restriction enzymes and methyltransferases.
<3>Nucleic Acids Res.
<4>31
<5>418-420
<6>2003
<7>REBASE contains comprehensive information about restriction enzymes, DNA methyltransferases
and related proteins such as nicking enzymes,
specificity subunits and control proteins. It contains published and
unpublished references, recognition and cleavage sites, isoschizomers,
commercial availability, crystal and sequence data. Homing endonucleases
are also included. REBASE contains the most complete and up-to-date
information about the methylation sensitivity of restriction
endonucleases. In addition, there is extensive information about the known
and putative restriction-modification (R-M) systems in more than 100
sequenced bacterial and archaeal genomes. The data is available on the web
(http://rebase.neb.com/rebase/rebase.html), through ftp (ftp.neb.com) and
as monthly updates via email.

<>

<1>Roberts, R.J., Vincze, T., Posfai, J., Macelis, D.
<2>REBASE - Restriction enzymes and DNA methyltransferases.
<3>Nucleic Acids Res.
<4>33
<5>D230-D232
<6>2005
<7>REBASE is a comprehensive database of information about restriction enzymes, DNA
methyltransferases and related proteins involved in restriction-modification.  It contains
both published and unpublished work with information about recognition and cleavage sites,
isoschizomers, commercial availability, crystal and sequence data.  Experimentally
characterized homing endonucleases are also included.  Additionally, REBASE contains complete
and up-to-date information about the methylation sensitivity of restriction endonucleases.  An
extensive analysis is included of the restriction-modification systems that are predicted to
be present in the sequenced bacterial and archaeal genomes from GenBank.  The contents of
REBASE are available by browsing from the web (http://rebase.neb.com/rebase/rebase.html) and
through selected compilations by ftp (ftp.neb.com) and as monthly updates that can be
requested via email.

<>

<1>Roberts, R.J., Vincze, T., Posfai, J., Macelis, D.
<2>REBASE--enzymes and genes for DNA restriction and modification.
<3>Nucleic Acids Res.
<4>35
<5>D269-D270
<6>2007
<7>REBASE is a comprehensive database of information about restriction enzymes, DNA
methyltransferases and related proteins involved in the biological process of
restriction-modification. It contains fully referenced information about recognition and
cleavage sites, isoschizomers, neoschizomers, commercial availability, methylation
sensitivity, crystal and sequence data. Experimentally characterized homing endonucleases are
also included. All newly sequenced genomes are analyzed for the presence of putative
restriction systems and these data are included within the REBASE. The contents or REBASE may
be browsed from the web (http://rebase.neb.com/rebase/rebase.ftp.html) and selected
compilations can be downloaded by ftp (ftp.neb.com). Additionally, monthly updates can be
requested via email.

<>

<1>Roberts, R.J., Vincze, T., Posfai, J., Macelis, D.
<2>REBASE--a database for DNA restriction and modification: enzymes, genes and genomes.
<3>Nucleic Acids Res.
<4>38
<5>D234-D236
<6>2010
<7>REBASE is a comprehensive database of information about restriction enzymes, DNA
methyltransferases and related proteins involved in the
biological process of restriction-modification (R-M). It contains fully
referenced information about recognition and cleavage sites,
isoschizomers, neoschizomers, commercial availability, methylation
sensitivity, crystal and sequence data. Experimentally characterized
homing endonucleases are also included. The fastest growing segment of
REBASE contains the putative R-M systems found in the sequence databases.
Comprehensive descriptions of the R-M content of all fully sequenced
genomes are available including summary schematics. The contents of REBASE
may be browsed from the web (http://rebase.neb.com) and selected
compilations can be downloaded by ftp (ftp.neb.com). Additionally, monthly
updates can be requested via email.

<>

<1>Roberts, R.J., Wilson, G.A., Young, F.E.
<2>Recognition sequence of specific endonuclease BamHI from Bacillus amyloliquefaciens H.
<3>Nature
<4>265
<5>82-84
<6>1977
<7>Many specific endonucleases (restriction endonucleases) have been isolated and
recognition sequences have been determined for a number of them.  The isolation
of a new specific endonuclease, BamHI, from Bacillus amyloliquefaciens H has
recently been described.  We have determined the recognition sequence of BamHI
and find that it cleaves the two-fold rotationallly symmetric sequence
5'-G-^G-A-T-C-C-3' 3'-C-C-T-A-G-^G-5' at the positions indicated by the arrows
generating fragments with cohesive termini.

<>

<1>Robertson, A.K., Geiman, T.M., Sankpal, U.T., Hager, G.L., Robertson, K.D.
<2>Effects of chromatin structure on the enzymatic and DNA binding functions of DNA methyltransferases DNMT1 and Dnmt3a in vitro.
<3>Biochem. Biophys. Res. Commun.
<4>322
<5>110-118
<6>2004
<7>DNA methylation is an epigenetic modification of the genome critical for numerous processes,
including transcriptional repression and
maintenance of chromatin structure. Recent studies have revealed
connections between DNA methylation and other epigenetic modifications
such as ATP-dependent chromatin remodeling. It remains unclear,
however, exactly how chromatin and epigenetic chromatin modifications
affect the biological properties of the DNA methyltransferases (DNMT1,
DNMT3A, and DNMT3B). Using a highly purified system and the 5S rDNA
gene as free DNA or assembled into a mononucleosome, we have compared
the effects of chromatin structure on DNMT1 and Dnmt3a. The catalytic
efficiency for both enzymes decreased on the mononucleosome, similar
to8-fold for DNMT1 and 17-fold for Dnmt3a. DNMT1 and Dnmt3a bound to
DNA and mononucleosomal substrates in gel shift experiments with
approximately equal affinity and in a cooperative manner. We also show
that DNMT1 interacts with hSNF2H chromatin remodeling enzyme and that
DNMT1 binds mononucleosomes with higher affinity in the presence of
hSNF2H. These findings raise interesting implications about the
interactions of mammalian DNA methyltransferases with chromatin and
provide the first evidence that a chromatin remodeling enzyme can alter
the biological properties of a DNMT.

<>

<1>Robertson, G.T., Reisenauer, A., Wright, R., Jensen, R.B., Jensen, A., Shapiro, L., Roop, R.M.
<2>The Brucella abortus CcrM DNA methyltransferase is essential for viability, and its overexpression attenuates intracellular replication in murine macrophages.
<3>J. Bacteriol.
<4>182
<5>3482-3489
<6>2000
<7>The CcrM DNA methyltransferase of the alpha-proteobacteria catalyzes the methylation of the
adenine in the sequence GAnTC.  Like Dam in the enterobacteria, CcrM plays a regulatory role
in Caulobacter crescentus and Rhizobium meliloti.  CcrM is essential for viability in both of
these organisms, and we show here that it is also essential in Brucella abortus.  Further,
increased copy number of the ccrM gene results in striking changes in B. abortus morphology,
DNA replication, and growth in murine macrophages.  We generated strains that carry ccrM
either on a low-copy-number plasmid (strain GR131) or on a moderate-copy-number plasmid
(strain GR132).  Strain GR131 has wild-type morphology and chromosome number, as assessed by
flow cytometry.  In contrast, strain GR132 has abnormal branched morphology, suggesting
aberrant cell division, and increased chromosome number.  Although these strains exhibit
different morphologies and DNA content, the replication of both strains in macrophages is
attenuated.  These data imply that the reduction in survival in host cells is not due solely
to a cell division defect but is due to additional functions of CcrM.  Because CcrM is
essential in B. abortus and increased ccrM copy number attentuates survival in host cells, we
propose that CcrM is an appropriate target for new antibiotics.

<>

<1>Robertson, K.
<2>DNA methyltransferase function and regulation.
<3>J. Nutr.
<4>133
<5>3845S
<6>2003
<7>DNA methylation is an epigenetic modification of the genome catalyzed by a group of 3 DNA
methyltransferases: DNMT1, 3A, and 3B.  Methyl groups are not randomly distributed in
mammalian cells but rather are compartmentalized in repetitive DNA, heterochromatic regions,
and parasitic elements.  Other regions of the genome, such as CpG island promoters, are almost
always unmethylated.  Given the minimal sequence requirements of the DNMTs (CpG), it is likely
that they are directed to sequences that are to be methylated by interactions with other
proteins, particularly chromatin-associated factors.  This compartmentalization is essential
because genetic knockouts of the DNMTs lead to embryonic lethality, and reversal of the normal
DNA methylation patterns is a hallmark of the most transformed cells.

<>

<1>Robertson, K.D.
<2>DNA methylation, methyltransferases, and cancer.
<3>Oncogene
<4>20
<5>3139-3155
<6>2001
<7>The field of epigenetics has recently moved to the forefront of studies relating to diverse
processes such as transcriptional regulation, chromatin structure, genome integrity, and
tumorigenesis. Recent work has revealed how DNA methylation and chromatin structure are linked
at the molecular level and how methylation anomalies play a direct causal role in
tumorigenesis and genetic disease. Much new information has also come to light regarding the
cellular methylation machinery, known as the DNA methyltransferases, in terms of their roles
in mammalian development and the types of proteins they are known to interact with. This
information has forced a new view for the role of DNA methyltransferases. Rather than enzymes
that act in isolation to copy methylation patterns after replication, the types of
interactions discovered thus far indicate that DNA methyltransferases may be components of
larger complexes actively involved in transcriptional control and chromatin structure
modulation. These new findings will likely enhance our understanding of the myriad roles of
DNA methylation in disease as well as point the way to novel therapies to prevent or repair
these defects.

<>

<1>Robertson, K.D., Ait-Si-Ali, S., Yokochi, T., Wade, P.A., Jones, P.L., Wolffe, A.P.
<2>DNMT1 forms a complex with Rb, E2F1 and HDAC1 and represses transcription from E2F-responsive promoters.
<3>Nat. Genet.
<4>25
<5>338-342
<6>2000
<7>Methylation of CpG islands is associated with transcriptional silencing and the formation of
nuclease-resistant chromatin structures enriched in hypoacetylated histones.
Methyl-CpG-binding proteins, such as MeCP2, provide a link between methylated DNA and
hypoacetylated histones by recruiting histone deacetylase, but the mechanisms establishing the
methylation patterns themselves are unknown. Whether DNA methylation is always causal for the
assembly of repressive chromatin or whether features of transcriptionally silent chromatin
might target methyltransferase remains unresolved. Mammalian DNA methyltransferases show
little sequence specificity in vitro, yet methylation can be targeted in vivo within
chromosomes to repetitive elements, centromeres and imprinted loci. This targeting is
frequently disrupted in tumour cells, resulting in the improper silencing of tumour-suppressor
genes associated with CpG islands. Here we show that the predominant mammalian DNA
methyltransferase, DNMT1, co-purifies with the retinoblastoma (Rb) tumour suppressor gene
product, E2F1, and HDAC1 and that DNMT1 cooperates with Rb to repress transcription from
promoters containing E2F-binding sites. These results establish a link between DNA
methylation, histone deacetylase and sequence-specific DNA binding activity, as well as a
growth-regulatory pathway that is disrupted in nearly all cancer cells.

<>

<1>Robertson, K.D., Jones, P.A.
<2>DNA methylation: past, present and future directions.
<3>Carcinogenesis
<4>21
<5>461-467
<6>2000
<7>DNA methylation, or the covalent addition of a methyl group to cytosine within the context of
the CpG dinucleotide, has profound effects on the mammalian genome. These effects include
transcriptional repression via inhibition of transcription factor binding or the recruitment
of
methyl-binding proteins and their associated chromatin remodeling factors, X chromosome
activation, imprinting and the suppression of parasitic DNA sequences. DNA methylation is also
essential for proper embryonic development; however, its presence can add an additional burden
to the genome. Normal methylation patterns are frequently disrupted in tumor cells with global
hypomethylation accompanying region-specific hypermethylation. When these hypermethylation
events occur within the promoter of a tumor suppressor gene they will silence the gene and
provide the cell with a growth advantage in a manner akin to deletions or mutations. Recent
work indicating that DNA methylation is an important player in both DNA repair and genome
stability as well as the discovery of a new family of DNA methyltransferases makes now a very
exciting period for the methylation field. This review will highlight the major findings in
the methylation field over the past 20 years then summarize the most important and interesting
future directions the field is likely to take in the next millennium.

<>

<1>Robertson, K.D., Keyomarsi, K., Gonzales, F.A., Velicescu, M., Jones, P.A.
<2>Differential mRNA expression of the human DNA methyltransferases (DNMTs) 1, 3a and 3b during the G0/G1 to S phase transition in normal and tumor cells.
<3>Nucleic Acids Res.
<4>28
<5>2108-2113
<6>2000
<7>DNA methylation is essential for mammalian development, X-chromosome inactivation, and
imprinting yet aberrant methylation patterns are one of the most common features of
transformed cells. One of the proposed causes for these defects in the methylation machinery
is overexpression of one or more of the three known catalytically active DNA
methyltransferases (DNMTs) 1, 3a and 3b, yet there are clearly examples in which
overexpression is minimal or non-existent but global methylation anomalies persist. An
alternative mechanism which could give rise to global methylation errors is the improper
expression of one or more of the DNMTs during the cell cycle. To begin to study the latter
possibility we examined the expression of the mRNAs for DNMT1, 3a and 3b during the cell cycle
of normal and transformed cells. We found that DNMT1 and 3b levels were significantly
downregulated in G0/G1 while DNMT3a mRNA levels were less sensitive to cell cycle
alterations and were maintained at a slightly higher level in tumor lines compared to normal
cell strains. Enzymatic activity assays revealed a similar decrease in the overall methylation
capacity of the cells during G0/G1 arrest and again revealed that a tumor cell line
maintained a higher methylation capacity during arrest than a normal cell strain. These
results reveal a new level of control exerted over the cellular DNA methylation machinery, the
loss of which provides an alternative mechanism for the genesis of the aberrant methylation
patterns observed in tumor cells.

<>

<1>Robertson, K.D., Uzvolgyi, E., Liang, G., Talmadge, C., Sumegi, J., Gonzales, F.A., Jones, P.A.
<2>The human DNA methyltransferases 1, 3a and 3b: coordinate mRNA expression in normal tissues and overexpression in tumors.
<3>Nucleic Acids Res.
<4>27
<5>2291-2298
<6>1999
<7>DNA methylation in mammals is required for embryonic development, X chromosome inactivation
and imprinting.  Previous studies have shown that methylation patterns become abnormal in
malignant cells and may contribute to tumorigenesis by improper de novo methylation and
silencing of the promoters for growth-regulatory genes.  RNA and protein levels of the DNA
methyltransferase DNMT1 have been shown to be elevated in tumors, however murine stem cells
lacking Dnmt1 are still able to de novo methylate viral DNA.  The recent cloning of a new
family of DNA methyltransferases (Dmnt3a and Dmnt3b) in mouse which methylate hemimethylated
and unmethylated templates with equal efficiencies make them candidates for the long sought de
novo methyltransferases.  We have investigated the expression of human DNMT1, 3a and 3b and
found widespread, coordinate expression of all three transcripts in most normal tissues.
Chromosomal mapping placed DNMT3a on chromosome 2p23 and DNMT3b on chromosome 20q11.2.
Significant overexpression of DNMT3b was seen in tumors while DNMT1 and DNMT3a were only
modestly overexpressed and with lower frequency.  Lastly, several novel alternatively spliced
forms of DNMT3b, which may have altered enzymatic activity, were found to be expressed in a
tissue-specific manner.

<>

<1>Robertson, K.D., Wolffe, A.P.
<2>DNA methylation in health and disease.
<3>Genetics
<4>1
<5>11-19
<6>2000
<7>DNA methylation has recently moved to centre stage in the aetiology of human
neurodevelopmental syndromes such as the fragile X, ICF and Rett syndromes.  These diseases
result from the misregulation of genes that occurs with the loss of appropriate epigenetic
controls during neuronal development.  Recent advances have connected DNA methylation to
chromatin-remodelling enzymes, and understanding this link will be central to the design of
new therapeutic tools.

<>

<1>Robinson, A., Gorringe, A.R., Hudson, M.J., Bracegirdle, P., West, D.M., Oliver, K.J., Kroll, J.S., Langford, P.R.
<2>Pathogenic and commensal vaccine antigens.
<3>International Patent Office
<4>WO 02077648 A
<5>
<6>2002
<7>The invention provides methods of screening commensal and pathogenic bacteria for previously
unidentified vaccine antigens, based upon identifying polypeptide antigens that bind to sera
raised against commensal bacterial proteins.  Also provided are vaccine compositions and
methods of preparing vaccine compositions comprising the antigens identified by the screening
methods.  Antigens and uses thereof are also described.

<>

<1>Robinson, C.R., Sligar, S.G.
<2>Molecular recognition mediated by bound water. A mechanism for star activity of the restriction endonuclease EcoRI.
<3>J. Mol. Biol.
<4>234
<5>302-306
<6>1993
<7>Many restriction endonucleases such as EcoRI lose some specificity for their recognition
sequence under certain buffer conditions. The cause of this disruption of accurate protein-DNA
recognition has never been explained. By cleaving DNA with EcoRI in the presence of several
osmolytes, we show that the extent of this EcoRI "star activity" depends strongly upon osmotic
pressure. The loss of specificity accompanying decreased water activity implies a role for one
or more water molecules in recognition of specific sequences of DNA. Water mediation may
constitute a general motif for sequence-specific DNA recognition by restriction enzymes and
other DNA-binding proteins.

<>

<1>Robinson, C.R., Sligar, S.G.
<2>Hydrostatic pressure reverses osmotic pressure effects on the specificity of EcoRI-DNA interactions.
<3>Biochemistry
<4>33
<5>3787-3793
<6>1994
<7>To characterize the role of water in protein-DNA interactions, we have studied the specificity
of the EcoRI restriction endonuclease as a function of osmotic and hydrostatic pressure. The
extent of cleavage by the enzyme at noncanonical ("star") sites is shown to depend uniquely
upon the osmotic pressure in the reaction as controlled by the addition of a wide variety of
neutral solutes. Alteration of cleavage specificity ("EcoRI* activity") is not uniformly
correlated with any other colligative solvent property such as dielectric constant, viscosity,
or water concentration. The application of hydrostatic pressure reverses the effects of
osmotic pressure, restoring the natural selectivity of the enzyme for its canonical site
GAATTC. This combination of observations provides compelling evidence that the site-specific
recognition of canonical site DNA by EcoRI is mediated by discretely bound water molecules and
that the release of these waters induces a fundamental change in the specificity of the
interaction, leading to cleavage at alternative sites. This comprehensive analysis of solvent
effects facilitates the unambiguous identification of structurally and functionally specific
waters involved in macromolecular recognition events.

<>

<1>Robinson, C.R., Sligar, S.G.
<2>Mediation of molecular recognition in protein-DNA complexes by bound water: a mechanism for "star activity" of restriction endonucleases.
<3>Biophys. J.
<4>66
<5>A34
<6>1994
<7>For many restriction endonucleases such as EcoRI, accurate protein-DNA recognition is
disrupted by changes in buffer composition, leading to an unexplained loss of specificity
termed "star activity". We have found that the extent of cleavage by EcoRI at non-canonical
sites is strongly correlated with the osmotic pressure in the reaction. This relationship is
unique to osmotic pressure, and is independent of other physical or chemical properties of the
osmolyte. An analogous correlation between osmotic pressure and star activity is observed for
other restriction enzymes including PvuII and BamHI. Specificity for cleavage at canonical
sites is restored by the application of hydrostatic pressure to counteract the effects of
osmotic pressure. Elevated osmotic pressures induce a fundamental change in the selectivity of
EcoRI -- at 100 atm osmotic pressure the rate of cleavage at the canonical site actually
decreases, whereas the rate of cleavage at "star" sites increases. The alteration of
specificity accompanying release of water clearly implicates one or more water molecules in
mediating accurate recognition of specific sequences of DNA by the enzyme. The change in
selectivity is manifested in both the association and catalytic steps of the reaction. Under
standard conditions, water may participate as a general mediator for sequence specific
recognition of DNA by restriction enzymes and other DNA-binding proteins.

<>

<1>Robinson, C.R., Sligar, S.G.
<2>Mediation of molecular recognition in protein-DNA complexes by bound water: a mechanism for "star activity" of restriction endonucleases.
<3>J. Cell Biochem. Suppl.
<4>18C
<5>138
<6>1994
<7>For many restriction endonucleases such as EcoRI, accurate protein-DNA recognition is
disrupted by changes in buffer composition, leading to an unexplained loss of specificity
termed "star activity". We have found that the extent of cleavage by EcoRI at non-canonical
sites is strongly correlated with the osmotic pressure in the reaction. This relationship is
unique to osmotic pressure, and is independent of other physical or chemical properties of the
osmolyte. An analogous correlation between osmotic pressure and star activity is observed for
other restriction enzymes including PvuII and BamHI. For EcoRI, specificity for cleavage at
the canonical site is restored by the application of hydrostatic pressure to counteract the
effects of osmotic pressure. Elevated osmotic pressures induce a fundamental change in the
selectivity of EcoRI -- at 100 atm osmotic pressure the rate of cleavage at the canonical sie
actually decreases, whereas the rate of cleavage at "star" sites increases. The alteration of
specificity accompanying release of water clearly implicates one or more water molecules in
mediating accurate recognition of specific sequences of DNA by the enzyme. The change in
selectivity is manifested in both the association and catalytic steps of the reaction. Under
standard conditions, water may participate as a general mediator for sequence specific
recognition of DNA by restriction enzymes and other DNA-binding proteins.

<>

<1>Robinson, C.R., Sligar, S.G.
<2>Heterogeneity in molecular recognition by restriction endonucleases: Osmotic and hydrostatic pressure effects on BamHI, PvuII, and EcoRV specificity.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>92
<5>3444-3448
<6>1995
<7>The cleavage specificity of the PvuII and BamHI restriction endonucleases is found to be
dramatically reduced at elevated osmotic pressure. Relaxation in specificity of these
otherwise highly accurate and specific enzymes, previously termed "star activity", is uniquely
correlated with osmotic pressure between 0 and 100 atmospheres. No other colligative solvent
property exhibits a uniform correlation with star activity for all of the compounds tested.
Application of hydrostatic pressure counteracts the effects of osmotic pressure and restores
the natural selectivity of the enzymes for their canonical recognition sequences. These
results indicate that water solvation plays an important role in the site-specific recognition
of DNA by many restriction enzymes. Osmotic pressure did not induce an analogous effect on the
specificity of the EcoRV endonuclease, implying that selective hydration effects do not
participate in DNA recognition in this system. Hydrostatic pressure was found to have little
effect on the star activity induced by changes in ionic strength, pH, or divalent cation,
suggesting that distinct mechanisms may exist for these observed alterations in specificity.
Recent evidence has indicated that BamHI and EcoRI share similar structural motifs, while
PvuII and EcoRV belong to a different structural family. Evidently, the use of hydration water
to assist in site-specific recognition is a motif neither limited to nor defined by structural
families.

<>

<1>Robinson, C.R., Sligar, S.G.
<2>Changes in solvation during DNA binding and cleavage are critical to altered specificity of the EcoRI endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>2186-2191
<6>1998
<7>Restriction endonucleases such as EcoRI bind and cleave DNA with great specificity and
represent a paradigm for protein-DNA interactions and molecular recognition.  Using osmotic
pressure to induce water release, we demonstrate the participation of bound waters in the
sequence discrimination of substrate DNA by EcoRI.  Changes in solvation can play a critical
role in directing sequence-specific DNA binding by EcoRI and are also crucial in assisting
site discrimination during catalysis.  By measuring the volume change for complex formation,
we show that at the cognate sequence (GAATTC) EcoRI binding releases about 70 fewer water
molecules than binding at an alternate DNA sequence (TAATTC), which differs by a single base
pair.  EcoRI complexation with nonspecific DNA releases substantially less water than either
of these specific complexes.  In cognate substrates (GAATTC) kcat decreases as osmotic
pressure is increased, indicating that binding of about 30 water molecules accompanies the
cleavage reaction.  For the alternate substrate (TAATTC), release of about 40 water molecules
accompanies the reaction, indicated by a dramatic acceleration of the rate when osmotic
pressure is raised.  These large differences in solvation effects demonstrate that water
molecules can be key players in the molecular recognition process during both association and
catalytic phases of the EcoRI reaction, acting to change the specificity of the enzyme.  For
both the protein-DNA complex and the transition state, there may be substantial conformational
differences between cognate and alternate sites, accompanied by significant alterations in
hydration and solvent accessibility.

<>

<1>Robinson, D., Walsh, P.R., Bonventre, J.A.
<2>Restriction endonucleases.
<3>Molecular biology problem solver., Wiley-Liss, Inc., Gerstein, A.S., New York
<4>
<5>225-266
<6>2001
<7>Molecular biologists routinely use restriction enzymes as key reagents for a variety of
applications including genomic mapping, restriction fragment length polymorphism (RFLP)
analysis, DNA sequencing, and a host of recombinant DNA methodologies.  Few would argue that
these enzymes are not indispensable tools for the variety of techniques used in the
manipulation of DNA, but like many common tools that are easy to use, they are not always
applied as efficiently and effectively as possible.  This chapter focuses on the biochemical
attributes and requirements of restriction enzymes and delivers strategies to optimize their
use in simple and complex reactions.

<>

<1>Robinson, V.L., Oyston, P.C., Titball, R.W.
<2>No title supplied.
<3>International Patent Office
<4>WO 2005120561 A
<5>
<6>2005
<7>A recombinant microorganism which is a recombinant Yersinia species which has been mutated so
that DNA adenine methylase activity is downregulated, for use as a prophylactic or therapeutic
vaccine against infection by a Yersinia species.  In particular, the microorganism is an
isogenic mutant, in which the dam gene is at least partially replaced, with for example a
selection marker.  These can provide cross protection against a range of Yersinia species such
as Y. pestis and Y. pseudotuberculosis.

<>

<1>Robinson, V.L., Oyston, P.C., Titball, R.W.
<2>A dam mutant of Yersinia pestis is attenuated and induces protection against plague.
<3>FEMS Microbiol. Lett.
<4>252
<5>251-256
<6>2005
<7>We have constructed a dam mutant of Yersinia pestis GB. In BALB/c mice inoculated
subcutaneously, the median lethal dose of the mutant was at least 2000-fold
higher than the wild type. Mice inoculated with sub-lethal doses of the mutant
were protected against a subsequent challenge with virulent Y. pestis. The effect
of dam inactivation on gene expression was examined using a DNA microarray, which
revealed increased expression of a number of genes associated with the SOS
response. These results confirm the key role of Dam in the regulation of
virulence, and its potential role as a target for the generation of attenuated
strains of pathogenic bacteria.

<>

<1>Rocap, G. et al.
<2>Genome divergence in two Prochlorococcus ecotypes reflects oceanic niche differentiation.
<3>Nature
<4>424
<5>1042-1047
<6>2003
<7>The marine unicellular cyanobacterium Prochlorococcus is the smallest-known oxygen-evolving
autotroph. It numerically dominates the
phytoplankton in the tropical and subtropical oceans, and is responsible
for a significant fraction of global photosynthesis. Here we compare the
genomes of two Prochlorococcus strains that span the largest evolutionary
distance within the Prochlorococcus lineage and that have different
minimum, maximum and optimal light intensities for growth. The
high-light-adapted ecotype has the smallest genome (1,657,990 base pairs,
1,716 genes) of any known oxygenic phototroph, whereas the genome of its
low-light-adapted counterpart is significantly larger, at 2,410,873 base
pairs (2,275 genes). The comparative architectures of these two strains
reveal dynamic genomes that are constantly changing in response to myriad
selection pressures. Although the two strains have 1,350 genes in common,
a significant number are not shared, and these have been differentially
retained from the common ancestor, or acquired through duplication or
lateral transfer. Some of these genes have obvious roles in determining
the relative fitness of the ecotypes in response to key environmental
variables, and hence in regulating their distribution and abundance in the
oceans.

<>

<1>Rocha, E.P., Danchin, A., Viari, A.
<2>Evolutionary role of restriction/modification systems as revealed by comparative genome analysis.
<3>Genome Res.
<4>11
<5>946-958
<6>2001
<7>Type II restriction modification systems (RMSs) have been regarded either as defense tools or
as molecular parasites of bacteria. We extensively analyzed their evolutionary role from the
study of their impact in the complete genomes of 26 bacteria and 35 phages in terms of
palindrome avoidance. This analysis reveals that palindrome avoidance is not universally
spread among bacterial species and that it does not correlate with taxonomic proximity.
Palindrome avoidance is also not universal among bacteriophage, even when their hosts code for
RMSs, and depends strongly on the genetic material of the phage. Interestingly, palindrome
avoidance is intimately correlated with the infective behavior of the phage. We observe that
the degree of palindrome and restriction site avoidance is significantly and consistently less
important in phages than in their bacterial hosts. This result brings to the fore a larger
selective load for palindrome and restriction site avoidance on the bacterial hosts than on
their infecting phages. It is then consistent with a view where type II RMSs are considered as
parasites possibly at the verge of mutualism. As a consequence, RMSs constitute a nontrivial
third player in the host-parasite relationship between bacteria and phages.

<>

<1>Rocha, E.P.C., Blanchard, A.
<2>Genomic repeats, genome plasticity and the dynamics of Mycoplasma evolution.
<3>Nucleic Acids Res.
<4>30
<5>2031-2042
<6>2002
<7>Mycoplasmas evolved by a drastic reduction in genome size, but their genomes contain numerous
repeated sequences with important roles in their evolution. We have established a
bioinformatic strategy to detect the major recombination hot-spots in the genomes of
Mycoplasma pneumoniae, Mycoplasma genitalium, Ureaplasma urealyticum and Mycoplasma pulmonis.
This allowed the identification of large numbers of potentially variable regions, as well as a
comparison of the relative recombination potentials of different genomic regions. Different
trends are perceptible among mycoplasmas, probably due to different functional and structural
constraints. The largest potential for illegitimate recombination in M. pulmonis is found at
the vsa locus and its comparison in two different strains reveals numerous changes since
divergence. On the other hand, the main M. pneumoniae and M. genitalium adhesins rely on large
distant repeats and, hence, homologous recombination for variation. However, the relation
between the existence of repeats and antigenic variation is not necessarily straightforward,
since repeats of P1 adhesin were found to be anti-correlated with epitopes recognized by
patient antibodies. These different strategies have important consequences for the structures
of genomes, since large distant repeats correlate well with the major chromosomal
rearrangements. Probably to avoid such events, mycoplasmas strongly avoid inverse repeats, in
comparison to co-oriented repeats.

<>

<1>Rochaix, J.D., Rahire, M., Michel, F.
<2>The chloroplast ribosomal intron of Chlamydomonas reinhardii codes for a polypeptide related to mitochondrial maturases.
<3>Nucleic Acids Res.
<4>13
<5>975-984
<6>1985
<7>The sequences of the 888bp chloroplast ribosomal intron and of the flanking 23S rRNA gene
regions of Chlamydomonas reinhardii have been established. The intron can be folded with a
secondary structure which is typical of group I introns of fungal mitochondrial genes. It
contains a 489bp open reading frame encoding a potential polypeptide that is related to
mitochondrial maturases.

<>

<1>Rochepeau, P., Selinger, L.B., Hynes, M.F.
<2>Transposon-like structure of a new plasmid-encoded restriction-modification system in Rhizobium leguminosarum VF39SM.
<3>Mol. Gen. Genet.
<4>256
<5>387-396
<6>1997
<7>Total DNA isolated from Rhizobium leguminosarum VF39SM cells is resistant to cleavage by the
restriction endonuclease PstI.  Plasmid curing and transfer studies localized this phenotype
to pRleVF39b, the second smallest of six plasmids found in this bacterium.  In vitro selection
for vector modification was employed to isolate a presumptive methylase gene (M.Rle39BI) from
a plasmid gene library.  Total and plasmid DNAs isolated from E. coli containing M.RleBI were
resistant to digestion by PstI.  Sequence data suggested that a putative restriction
endonuclease (R.Rle39BI) was also encoded on the same fragment.  The two genes were flanked by
identical copies of a putative insertion sequence, which was also present in several copies
elsewhere in the VF39SM genome.  The presence of this element in other strains examined
suggested that this element is indeed an insertion sequence.  The differences in G/C content
between the DNA coding for the R/M system and that of the IS element suggest that this DNA
region may have been acquired by horizontal transfer.

<>

<1>Rockah-Shmuel, L., Tawfik, D.S.
<2>Evolutionary transitions to new DNA methyltransferases through target site expansion and shrinkage.
<3>Nucleic Acids Res.
<4>40
<5>11627-11637
<6>2012
<7>DNA-binding and modifying proteins show high specificity but also exhibit a certain level of
promiscuity. Such latent promiscuous activities comprise the
starting points for new protein functions, but this hypothesis presents a
paradox: a new activity can only evolve if it already exists. How then, do novel
activities evolve? DNA methyltransferases, for example, are highly divergent in
their target sites, but how transitions toward novel sites occur remains unknown.
We performed laboratory evolution of the DNA methyltransferase M.HaeIII. We found
that new target sites emerged primarily through expansion of the original site,
GGCC, and the subsequent shrinkage of evolved expanded sites. Variants evolved
for sites that are promiscuously methylated by M.HaeIII [GG((A)/(T))CC and
GGCGCC] carried mutations in 'gate-keeper' residues. They could thereby methylate
novel target sites such as GCGC and GGATCC that were neither selected for nor
present in M.HaeIII. These 'generalist' intermediates were further evolved to
obtain variants with novel target specificities. Our results demonstrate the ease
by which new DNA-binding and modifying specificities evolve and the mechanism by
which they occur at both the protein and DNA levels.

<>

<1>Roddy, E.S., Price, M., Ewing, A.G.
<2>Continuous monitoring of a restriction enzyme digest of DNA on a microchip with automated capillary sample introduction.
<3>Anal. Chem.
<4>75
<5>3704-3711
<6>2003
<7>Continuous analysis of a DNA restriction enzyme digest on a microfabricated device is
demonstrated with minimal intervention and
enhanced time resolution. A 62-base-pair fragment of dsDNA containing a
KpnI site was used to demonstrate this process. A capillary was used to
transfer sample from a single reaction mix to a microfabricated chip with
parallel separation lanes. The 6-carboxyfluorescein-labeled DNA fragments
were detected with a CCD camera as they separated in the lanes, which were
filled with linear polyacrylamide. The products of the restriction enzyme
digest were monitored for up to 60 min at an average sampling rate of 1
injection/46 s, with consecutive injections as short as 1 injection/14 s.
The digest was injected directly into the chip, eliminating the need for
any sample-handling steps after addition of the enzyme to the reaction
mix. The effects of temperature and restriction enzyme concentration were
briefly examined, as well. This work shows the potential of this method to
provide valuable information about the process of restriction enzyme
cleavage.

<>

<1>Rodicio, M.R., Alvarez, M.A., Chater, K.F.
<2>Isolation and genetic structure of IS112, an insertion sequence responsible for the inactivation of the SalI restriction-modification system of Streptomyces albus G.
<3>Mol. Gen. Genet.
<4>225
<5>142-147
<6>1991
<7>IS112 is a transposable element identified in Streptomyces albus G by its
frequent mutagenic insertion into the genes for the SalI
restriction-modification system.  IS112 is present in several copies in the
genome of S. albus G.  Homologous sequences were detected in other Streptomyces
strains.  Sequence analysis revealed that IS112 has a length of 883 bp with a
GC content of 67.4%.  The copy that was isolated contained imperfect inverted
repeats (16/20 match) at its ends and was flanked by a 2 bp duplication at the
target site, which was located within the gene (salIR) for the SalI
endonuclease.  A long open reading frame (ORF) encoding a putative polypeptide
of 256-253 amino acids spans almost the entire sequence.  Significant homology
was detected between this polypeptide and that corresponding to ORFB of IS493,
an insertion sequence recently isolated from Streptomyces lividans 66.

<>

<1>Rodicio, M.R., Chater, K.F.
<2>Cloning and expression of the SalI restriction-modification genes of Streptomyces albus G.
<3>Mol. Gen. Genet.
<4>213
<5>346-353
<6>1988
<7>The Streptomyces albus G genes (salR and salM) for the class II restriction
enzyme SalI (SalGI) and its cognate modification enzyme were cloned in
Streptomyces lividans 66.  Selection was initially for the salR gene.  From a
library of S. albus G DNA in the high copy number plasmid pIJ486 several clones
of S. lividans were obtained that were resistant to phage PhiC31 unmodified at
the many SalI sites in its DNA, but were sensitive to modified phages last
propagated on a restriction-deficient, modification- proficient mutant of S.
albus G.  SalI activity was detected in cell-free extracts of the clones,
though only at levels comparable with that in S. albus G.  Five different
recombinant plasmids were isolated, with inserts of 5.6, 5.7, 8.9, 10 and 18.9
kb that contained a common region of 4.5 kb.  These plasmids could not be
digested by SalI, although the vector has four recognition sites for this
enzyme, indicating that the salM gene was also cloned and expressed.
Subcloning experiments in S. lividans indicated the approximate location of
salR and salM, and in Escherichia coli led to detectable expression of salM but
not of salR.  A variety of previously isolated S. albus G mutants affected in
aspects of SalI-specific restriction and modification were complemented by the
cloned DNA; they included a mutant temperature-sensitive for growth apparently
because of a mutation in salM.  Southern blotting showed that DNA homologous to
the cloned sal genes was present in Xanthomonas and Rhodococcus strains, but
not detectably in Herpetosiphon strains, all of which produce SalI
isoschizomers.

<>

<1>Rodicio, M.R., Chater, K.F.
<2>The SalI (SalGI) restriction-modification system of Streptomyces albus G.
<3>Gene
<4>74
<5>39-42
<6>1988
<7>The salIR and salM genes of Streptomyces albus G specify the SalGI (SalI)
restriction enzyme and its cognate methyltransferase, respectively.  These
enzymes are responsible for restriction and modification of bacteriophages.
Some phages carry genes that interfere with SalI-specific modification.  The
sal genes have been cloned in a Streptomyces host-vector system.  Use of the
cloned DNA as a hybridization probe reveals that sal mutants frequently arise
from transposition of a DNA segment of approx. 1 kb into the sal genes.  Some,
but not all, other bacteria that produce SalGI isoschizomers contain nucleotide
sequences that hybridize with sal DNA.

<>

<1>Rodicio, M.R., Quinton-Jager, T., Moran, L.S., Slatko, B.E., Wilson, G.G.
<2>Organization and sequence of the SalI restriction-modification system.
<3>Gene
<4>151
<5>167-172
<6>1994
<7>The organization and nucleotide (nt) sequences were determined for the genes encoding the SalI
restriction and modification (R-M) system (recognition sequence 5'-GTCGAC-3') from
Streptomyces albus G. The system comprises two genes, salIR, coding for the restriction
endonuclease (ENase, R.SalI; probably 315 amino acids (aa), a predicted Mr of 35,305; product,
GTCGAC) and salIM, coding for the methyltransferase (MTase, M.SalI; probably 587 aa, a
predicted Mr of 64,943; product, GRCGm6AC). The genes are adjacent, they have the same
orientation, and they occur in the order salIR then SalIM. R.SalI contains a putative
magnesium-binding motif similar to those at the active sites of R.EcoRI and R.EcoRV, but
otherwise it bears little aa sequence similarity to other ENases. M.SalI is a member of the
m6A-gamma class of MTases. In aa sequence it resembles M.AccI, another m5A-gamma-MTase whose
recognition sequence includes the SalI recognition sequence as a subset.

<>

<1>Rodriguez, A., Lleonart, R., Martinez, A.S., Ryes, G., Gonzalez, E., Brito, J.E.
<2>Cloning and expression of genes coding for Haemophilus influenzae Rf restriction and modification enzymes.  Purification of the recombinant endonuclease.
<3>Biotecnol. Apl.
<4>12
<5>156-159
<6>1995
<7>The genes coding for HinfI restriction and modification enzymes were cloned from the
Haemophilus influenzae Rf strain using pUC18 plasmid as vector.  To select the genes from the
library, the classical modification phenotype selection was used.  When introduced into
Escherichia coli HB101 strain, the recombinant plasmid pERHinf4 was able to direct the
expression of the restriction phenotype.  The crude extracts from the transformed E. coli
contained roughly twice the amount of restrictase expressed by the natural source.  A
purification procedure for the restrictase is described which when applied to the recombinant
enzyme, renders a preparation free of contaminant exonucleases and phosphatases.  The overall
recovery of the purification procedure was about 33% of the total activity.

<>

<1>Rodriguez, R.J.
<2>Polyphosphate present in DNA preparations from Filamentous fungal species of Colletotrichum inhibits restriction endonucleases and other enzymes.
<3>Anal. Biochem.
<4>209
<5>291-297
<6>1993
<7>During the development of a procedure for the isolation of total genomic DNA from filamentous
fungi (Rodriguez, R.J., and Yoder, O.C., Exp. Myco. 15, 232-242, 1991) a cell fraction was
isolated which inhibited the digestion of DNA by restriction enzymes. After elimination of
DNA, RNA, proteins, and lipids, the active compund was purified by gel filtration to yield a
single fraction capable of complete inhibition of restriction enzyme activity. The inhibitor
did not absorb uv light above 220 nm, and was resistant to alkali and acid at 25 degrees C and
to temperatures as high as 100 degrees C. More extensive analyses demonstrated that the
inhibitor was also capable of inhibiting T4 DNA ligase and TaqI DNA polymerase, but not DNase
or RNase. Chemical analyses indicated that the inhibitor was devoid of carbohydrates, protein,
lipids and nucleic acids but rich in phosphorus. A combination of nuclear magnetic resonance,
metachromatic shift of toluidine blue, and gel filtration indicated that the inhibitor was a
polyphosphate (polyP) containing approximately 60 phosphate molecules. The mechanism of
inhibition appeared to involve complexing of polyP to the enzymatic proteins. All species of
Colletotrichum analyzed produced polyP equivalent in chain length and concentration. A
modification to the original DNA extraction procedure is described which eliminates polyP and
reduces the time necessary to obtain DNA of sufficient purity for restriction enzyme digestion
and TaqI polymerase amplification.

<>

<1>Rodriguez-Osorio, N., Wang, H.F., Rupinski, J., Bridges, S.M., Memili, E.
<2>Comparative functional genomics of mammalian DNA methyltransferases.
<3>Reprod. Biomed. Online
<4>20
<5>243-255
<6>2010
<7>DNA methylation involves biochemical modification of DNA by addition of methyl groups onto CpG
dinucleotides, and this epigenetic mechanism
regulates gene expression in disease and development. Mammalian DNA
methyltransferases, DNMT (DNMT1, DNMT3A and DNMT3B), together with the
accessory protein DNMT3L establish specific DNA methylation patterns in
the genome during gametogenesis, embryogenesis and somatic tissue
development. The present study addresses the structural and functional
conservation of the DNMT in humans, mice and cattle and the patterns of
mRNA abundance of the different enzymes during embryogenesis to improve
understanding of epigenetic regulation in early development. The
findings showed a high degree of structural and functional conservation
among the human, mouse, and bovine DNMT. The results also showed
similar patterns of transcript abundance for all of the proteins at
different stages of early embryo development. Remarkably, all of the
DNMT with an important role in DNA methylation (DNMT1, DNMT3A, DNMT3B,
and DNMT3L) show a greater degree of structural similarity between
human and bovine than that between human and mouse. These results have
important implications for the selection of an appropriate model for
study of DNA methylation during early development in humans.

<>

<1>Roer, L., Aarestrup, F.M., Hasman, H.
<2>EcoKI Type I Restriction-Modification System in Escherichia coli Affect, but is Not an Absolute Barrier for, Conjugation.
<3>J. Bacteriol.
<4>197
<5>337-342
<6>2014
<7>The rapid evolution of bacteria is crucial to their survival, and is among other  things
caused by exchange, transfer and uptake of DNA. Conjugation is one of the
main mechanisms by which bacteria share their DNA, and is thought to be
controlled by varied bacterial immune systems. Contradictory results, based on
phenotypical studies, have been presented on Restriction-Modification systems as
a barrier for conjugation and other means of uptake of exogenous DNA. In this
study, we show that inactivation of the R.EcoKI restriction enzyme in strain
Escherichia coli K-12 strain MG1655 increases the conjugational transfer of
plasmid pOLA52, which carriers two EcoKI recognition sites. Interestingly, the
results were not absolute, and uptake of un-methylated pOLA52 was still observed
in the wild-type strain (with intact hsdR gene), but with a reduction of 85%
compared to the mutant recipient having a disrupted hsdR gene. This leads to the
conclusion that EcoKI Restriction-Modification affects the uptake of DNA by
conjugation, but is not a major barrier for plasmid transfer.

<>

<1>Rogulin, E.A., Perevyazova, T.A., Zheleznaya, L.A., Matvienko, N.I.
<2>Plasmid pRARE as a vector for cloning to construct a superproducer of the site-specific nickase N.BspD6I.
<3>Biokhimiia
<4>69
<5>1123-1127
<6>2004
<7>The gene of methylase M.SccL1I that protects DNA against hydrolysis with the nickase N.BspD6I
was inserted into plasmid pRARE carrying
genes of tRNA, which are rare in E. coli. The insertion of the gene
sscML1I into pRARE was reasoned by incompatibility of pRARE and the
plasmid carrying the gene sscML1I, because both plasmids contained the
same ori-site. Upon transformation of E. coli TOP10F' cells with both
the recombinant plasmid pRARE/MSsc and the expression vector pET28b
containing the nickase gene bspD6IN under the phage T7 promoter, a
strain of E. coli was obtained which produced 7(.)10(5) units of the
nickase N.BspD6I per 1 g wet biomass, and this yield was two orders of
magnitude higher than the yield of the enzyme from the strain free of
pRARE/MSsc.

<>

<1>Roh, H., Ko, H.J., Kim, D., Choi, D.G., Park, S., Kim, S., Chang, I.S., Choi, I.G.
<2>Complete Genome Sequencing of A Carbon Monooxide Utilizing Acetogen, Eubacterium limosum KIST612.
<3>J. Bacteriol.
<4>193
<5>307-308
<6>2010
<7>Eubacterium limosum KIST612 is an anaerobic acetogenic bacterium that uses CO as a sole
carbon/energy source and produces acetate, butyrate and
ethanol. To evaluate its potential as a syngas microbial catalyst, we have
sequenced the complete 4.3 Mb genome of E. limosum KIST612.

<>

<1>Roh, H., Uguru, G.C., Ko, H.J., Kim, S., Kim, B.Y., Goodfellow, M., Bull, A.T., Kim, K.H., Bibb, M.J., Choi, I.G., Stach, J.E.
<2>Genome Sequence of the Abyssomicin- and Proximicin-Producing Marine Actinomycete Verrucosispora maris AB-18-032.
<3>J. Bacteriol.
<4>193
<5>3391-3392
<6>2011
<7>Verrucosispora maris AB-18-032 is a marine actinomycete that produces atrop-abyssomicin C and
proximicin A, both of which have novel structures
and modes of action. In order to understand the biosynthesis of these
compounds, to identify further biosynthetic potential and to facilitate
rational improvement of secondary metabolite titers, we have sequenced the
complete 6.7 Mb genome of Verrucosispora maris AB-18-032.

<>

<1>Roh, H., Yun, E.J., Lee, S., Ko, H.J., Kim, S., Kim, B.Y., Song, H., Lim, K.I., Kim, K.H., Choi, I.G.
<2>Genome Sequence of Vibrio sp. Strain EJY3, an Agarolytic Marine Bacterium Metabolizing 3,6-Anhydro-L-Galactose as a Sole Carbon Source.
<3>J. Bacteriol.
<4>194
<5>2773-2774
<6>2012
<7>The metabolic fate of 3,6-anhydro-l-galactose (l-AHG) is unknown in the global marine carbon
cycle. Vibrio sp. strain EJY3 is an agarolytic marine bacterium
that can utilize l-AHG as a sole carbon source. To elucidate the metabolic
pathways of l-AHG, we have sequenced the complete genome of Vibrio sp. strain
EJY3.

<>

<1>Roh, S.W., Nam, Y.D., Nam, S.H., Choi, S.H., Park, H.S., Bae, J.W.
<2>Complete Genome Sequence of Halalkalicoccus jeotgali B3T, an Extremely Halophilic Archaeon.
<3>J. Bacteriol.
<4>192
<5>4528-4529
<6>2010
<7>Halalkalicoccus jeotgali B3(T), isolated from salt-fermented seafood from Korea, is an
extremely halophilic archaeon belonging to the family
Halobacteriaceae. Here, we present the complete genome sequence of the
type strain Hac. jeotgali B3(T) (3,698,650 bp, with a G+C content of
62.5%), which consists of one chromosome and six plasmids. This is the
first complete genome sequence of the Halalkalicoccus species.

<>

<1>Rohmer, L., Brittnacher, M., Svensson, K., Buckley, D., Haugen, E., Zhou, Y., Chang, J., Levy, R., Hayden, H., Forsman, M., Olson, M., Johansson, A., Kaul, R., Miller, S.I.
<2>Potential source of Francisella tularensis live vaccine strain attenuation determined by genome comparison.
<3>Infect. Immun.
<4>74
<5>6895-6906
<6>2006
<7>Francisella tularensis is a bacterial pathogen that causes the zoonotic
disease tularemia and is important to biodefense. Currently, the only
vaccine known to confer protection against tularemia is a specific live
vaccine strain (designated LVS) derived from a virulent isolate of
Francisella tularensis subsp. holarctica. The origin and source of
attenuation of this strain are not known. To assist with the design of a
defined live vaccine strain, we sought to determine the genetic basis of
the attenuation of LVS. This analysis relied primarily on the comparison
between the genome of LVS and Francisella tularensis holarctica strain
FSC200, which differ by only 0.08% of their nucleotide sequences. Under
the assumption that the attenuation was due to a loss of function(s), only
coding regions were examined in this comparison. To complement this
analysis, the coding regions of two slightly more distantly related
Francisella tularensis strains were also compared against the LVS coding
regions. Thirty-five genes show unique sequence variations predicted to
alter the protein sequence in LVS compared to the other Francisella
tularensis strains. Due to these polymorphisms, the functions of 15 of
these genes are very likely lost or impaired. Seven of these genes were
demonstrated to be under stronger selective constraints, suggesting that
they are the most probable to be the source of LVS attenuation and useful
for a newly defined vaccine.

<>

<1>Roizes, G., Charlieu, J.-P., Marcais, B., Bellis, M.
<2>Use of the DNA fragments generated by a restriction enzyme (Bcg I) for the construction of overlapping clone libraries.
<3>DNA Seq.
<4>2
<5>65-67
<6>1991
<7>The human genome contains at intervals the sequence recognized by the
restriction endonuclease BcgI.  We propose to take advantage of the presence of
this sequence for the construction of overlapping clone libraries.  The
proposal holds for any other genome in which there are similar projects of
physical mapping and sequencing.

<>

<1>Roizes, G., Nardeux, P.C., Monier, R.
<2>A new specific endonuclease from Anabaena variabilis.
<3>FEBS Lett.
<4>104
<5>39-44
<6>1979
<7>A large number of site-specific endonucleases have been isolated from Anabaena
variabilis and we now describe a third from this organism, AvaIII, which cuts
SV40 DNA three times.  From the positions of these breaks on the physical map
of SV40, the DNA sequence recognised by the enzyme can be deduced.  It has not
yet been possible to separate this enzyme from AvaI.

<>

<1>Roizes, G., Pages, M., Lecou, C., Patillon, M., Kovoor, A.
<2>A new site-specific endonuclease showing phenotypical crypticity in a tumorigenic strain of Agrobacterium tumefaciens.
<3>Gene
<4>6
<5>43-50
<6>1979
<7>AtuBVI, an endonuclease showing new site-specificity, has been isolated from
the tumorigenic strain IIBV7 of Agrobacterium tumefaciens, and is undetectable
in the non-tumorigenic sister strain IIBNV6.  AtuBVI degrades IIBV7 DNA in
vitro and should, therefore, be regarded as being phenotypically cryptic in the
bacterial cell; it also shows anomalous behavior under certain incubation
conditions.  These properties point to a possible role for this enzyme in the
insertion of exogenous Ti-plasmid DNA into plant tissues during tumorigenesis.

<>

<1>Roizes, G., Patillon, M., Kovoor, A.
<2>A restriction endonuclease from Agrobacterium tumefaciens.
<3>FEBS Lett.
<4>82
<5>69-70
<6>1977
<7>The tumorous growth of plant tissues known as Crowngall is induced by certain
strains of Agrobacterium tumefaciens (Smith and Townsend, Conn.) and there is
considerable evidence that foreign DNA is integrated in the genome of the
transformed host cell.  A specific endonuclease of the bacterium could then
conceivably play a role in the insertion process.  In this letter we describe
the isolation of a sequence-specific endonuclease from a tumorigenic strain,
B6, of Agrobacterium tumefaciens.  The recognition site as well as the
modification specificity of this enzyme, named AtuI, are shown to be identical
to those of a known restriction enzyme, EcoRI.

<>

<1>Rojas, J.D., Starcevic, A., Baranasic, D., Ferreira-Torres, M.A., Contreras, C.A., Garrido, L.M., Araujo, W.L., de Souza, R.F., Zucko, J., Hranueli, D., Long, P.F., Cullum, J., Padilla, G.
<2>Genome Sequence of Streptomyces olindensis DAUFPE 5622, Producer of the Antitumoral Anthracycline Cosmomycin D.
<3>Genome Announcements
<4>2
<5>e00541-14
<6>2014
<7>Streptomyces olindensis DAUFPE 5622, which was isolated from a Brazilian soil sample, produces
the antitumor anthracycline cosmomycin D. The genome sequence is
9.4 Mb in length, with a G+C content of 71%. Thirty-four putative secondary
metabolite biosynthetic gene clusters were identified, including the cosmomycin D
cluster.

<>

<1>Rojas, T.C., Maluta, R.P., Parizzi, L.P., Koenigkan, L.V., Yang, J., Yu, J., Pereira, G.A., Dias-da-Silveira, W.
<2>Genome Sequences of Avian Pathogenic Escherichia coli Strains Isolated from Brazilian Commercial Poultry.
<3>Genome Announcements
<4>1
<5>e0011013
<6>2013
<7>Avian pathogenic Escherichia coli (APEC) infections are responsible for significant losses in
the poultry industry worldwide. The disease might present
as different local infections or as septicemia. Here, we present the draft genome
sequences of three Brazilian APEC strains isolated from different kinds of
infections. The availability of these APEC genome sequences is important for
gaining a thorough understanding of the genomic features of E. coli, particularly
those of this pathotype.

<>

<1>Rojas, T.C., Parizzi, L.P., Tiba, M.R., Chen, L., Pereira, G.A., Sangal, V., Yang, J., Yu, J., Dias-da-Silveira, W.
<2>Draft Genome of a Brazilian Avian-Pathogenic Escherichia coli Strain and In Silico Characterization of Virulence-Related Genes.
<3>J. Bacteriol.
<4>194
<5>3023
<6>2012
<7>Avian-pathogenic Escherichia coli (APEC) strains cause extraintestinal diseases in avian
species. Here, we present the draft genome of an APEC strain (SCI-07) from Brazil that was
isolated from skin lesions (gelatinous edema) on the head and periorbital tissues of a laying
hen with swollen head syndrome.

<>

<1>Rolain, J.M., Vayssier-Taussat, M., Gimenez, G., Robert, C., Fournier, P.E., Raoult, D.
<2>Genome Sequence of Bartonella birtlesii, a Bacterium Isolated from Small Rodents  of the Genus Apodemus.
<3>J. Bacteriol.
<4>194
<5>4779
<6>2012
<7>Bartonella birtlesii is a facultative intracellular bacterium isolated from the blood of small
mammals of the genus Apodemus. The present study reports the draft
genome of Bartonella birtlesii strain IBS 135(T) (CIP 106691(T)).

<>

<1>Romanenko, E.B., Pushkareva, M.Y., Alessenko, A.V., Vanyushin, B.F.
<2>Effects of Sphingomyelin and its enzymatic hydrolysis products on heterologous methylation of calf thymus DNA by cytosine-DNA methyltransferase EcoRII.
<3>Biokhimiia
<4>56
<5>295-300
<6>1991
<7>It was found that sphingomyelin and its enzymatic hydrolysis products, choline
and sphingosine, influence the degree of DNA methylation in the reaction of
heterologous methylation by methylase EcoRII in vitro.  Sphingomyelin was found
to be able to activate (by 20%), sphingosine and choline inhibit methylation.
Phosphatidylcholine had no effect on DNA methylation in an in vitro system.
The role of lipids in the regulation of gene expression during enzymatic
modification (methylation) of DNA is discussed.

<>

<1>Romanenkov, A.S., Kisil, O.V., Zatsepin, T.S., Yamskova, O.V., Karyagina, A.S., Metelev, V.G., Oretskaya, T.S., Kubareva, E.A.
<2>DNA-methyltransferase SsoII as a bifunctional protein: Features of the interaction with the promoter region of SsoII restriction-modification genes.
<3>Biokhimiia
<4>71
<5>1341-1349
<6>2006
<7>DNA duplexes bearing an aldehyde group at the 2'-position of the sugar moiety were used for
affinity modification of (cytosine-5)-DNA
methyltransferase SsoII. It is shown that lysine residues of M.SsoII
N-terminal region are located in proximity to DNA sugar-phosphate
backbone of a regulatory sequence of promoter region of SsoII
restriction-modification enzyme coding genes. The ability of the two
M.SsoII subunits to interact with DNA regulatory sequence has been
demonstrated by affinity modification using DNA duplexes with two
2'-aldehyde groups. Changes in nucleotide sequence of one half of the
regulatory region prevented cross-linking of the second M.SsoII
subunit. The results on sequential affinity modification of M.SsoII by
two types of modified DNA ligands (i.e. by 2'-aldehyde-containing and
phosphoryldisulfide-containing) have demonstrated the possibility of
covalent attachment of the protein to two different DNA recognition
sites: regulatory sequence and methylation site.

<>

<1>Romano, A., Trip, H., Campbell-Sills, H., Bouchez, O., Sherman, D., Lolkema, J.S., Lucas, P.M.
<2>Genome Sequence of Lactobacillus saerimneri 30a (Formerly Lactobacillus sp. Strain 30a), a Reference Lactic Acid Bacterium Strain Producing Biogenic Amines.
<3>Genome Announcements
<4>1
<5>e00097-12
<6>2013
<7>Lactobacillus sp. strain 30a (Lactobacillus saerimneri) produces the biogenic amines
histamine, putrescine, and cadaverine by decarboxylating their amino acid
precursors. We report its draft genome sequence (1,634,278 bases, 42.6% G+C
content) and the principal findings from its annotation, which might shed light
onto the enzymatic machineries that are involved in its production of biogenic
amines.

<>

<1>Romanovskaya, V.A., Alexeyev, M.F., Gunkovskaya, N.V., Stolyar, S.M., Shatohina, E.S., Malashenko, Y.R.
<2>Screening for restriction endonucleases in methane-oxidizing bacteria.
<3>Mikrobiol. Zh.
<4>54
<5>32-39
<6>1992
<7>51 methane-oxidizing bacterial strains such as Methylomonas methanica, M.rubra, Methylococcus
capsulatus, M. thermophilus, M. luteus, M. ucrainicus, M. whittenburyi, Methylosinus
trichosporium, M. sporium, Methylocystis parvus isolated from various ecological niches and
geographical regions of the Ukraine and also the strains received from R. Whittenbury and Y.
Heyer were screened for restriction endonucleases. Type II restriction endonucleases were
detected in IMV B-3112 (=12 b), IMV B-3027 (=26), IMV B-3019 (=9 c), IMV B-3017 (=17 c). IMV
B-3226 (=26 v). IMV B-3033 (=Y), IMV B-3100 (=100) and IMV B-3494 (=IE494). The results
obtained were indicative of a relatively high frequency of occurrence of restriction enzymes
in methane-oxidizing bacteria. There were KpnI (Asp718I) restriction endonuclease
isoschizomers in crude extracts of IMV B-3112, B-3017, B-3019, B-3027 isolated from
fresh-water silt as well as in IMV B-3226 strain isolated from waste-water silt. Although
these isolates had been previously considered as atypical strains of M. ucrainicus, more
detailed study of their properties allowed placing them with Methylovarius luteus
(=Methylococcus luteus). IMV B-3494 strain was identified as Methylococcus capsulatus. Strain
IMV B-3033 had earlier been allocated to Methylovarius whittenburyi (=Methylococcus
whittenburyi). Specificity of restriction endonucleases of this strain was not tested.
Therefore, for the first time restriction endonucleases were detected in methane-oxidizing
bacteria. 8 strains (3 species) among 51 strains (13 species) were found to produce
restriction endonucleases displaying three different types of specificity at least. Producers
of restriction endonucleases having KpnI (Asp718I) specificity were isolated from different
water and silt samples of the Dnieper flood-land more than 20 years ago.

<>

<1>Romero, D.A., Klaenhammer, T.R.
<2>Abortive phage infection and restriction/modification activities directed by pTR2030 determinants are enhanced by recombination with conjugal elements in lactococci.
<3>J. Gen. Microbiol.
<4>136
<5>1817-1824
<6>1990
<7>The recombinant plasmid pTK6 is composed of a 13.6 kb fragment from pTR2030
encoding phage resistance determinants for restriction/modification (R+/M+) and
abortive phage infection (Hsp+) cloned into shuttle vector pSA3 (erythromycin
resistance, Emr).  Conjugal matings were performed to mobilize pTK6-encoded
markers from Lactococcus lactis subsp. lactis MMS362 and MG1363.  Emr
transconjugants were recovered at 10-6 per input donor and harboured pTK6 or
recombinant plasmids not found in either parental strain.  The recombinant
plasmids (pTRK78 and pTRK79) encoded Emr, Hsp+ and R+/M+, and transferred at
high frequency in second-round matings.  Mobilization of pTK6 from the
otherwise plasmid-free donor, L. lactis MG1363, confirmed the presence of a
conjugal element in this strain.  Phage resistance in transconjugants
containing pTRK78 and pTRK79 was markedly enhanced over a pTK6-directed Hsp+
and R+/M+.  In L. lactis LM2345 transconjugants, a reduction in plaque size was
accompanied by a significant decrease in the efficiency of plaquing for phages
c2 (10-2 to 10-6) and p2(<10-9).  L. lactis NCK203 transconjugants containing
pTRK78 or pTRK79 exhibited an additional 100-1000-fold reduction in the
plaquing efficiency of Phi48 (10-4 to 10-5) over pTK6 imposed restriction
(10-2).  Increased resistance to phage was a consequence of the physical
interaction of pTR2030-derived sequences on pTK6 with a conjugal element
resident in the donor strains.

<>

<1>Romine, M.F., Stillwell, L.C., Wong, K.-K., Thurston, S.J., Sisk, E.C., Sensen, C., Gaasterland, T., Fredrickson, J.K., Saffer, J.D.
<2>Complete sequence of a 184-kilobase catabolic plasmid from Sphingomonas aromaticivorans F199.
<3>J. Bacteriol.
<4>181
<5>1585-1602
<6>1999
<7>The complete 184,457-bp sequence of the aromatic catabolic plasmid, pNL1,
from Sphingomonas aromaticivorans F199 has been determined.  A total of 186 open
reading frames are predicted to encode proteins, of which 79 are likely directly
associated with catabolism or transport of aromatic compounds.  Genes that encode
enzymes associated with the degradation of biphenyl, naphthalene, m-xylene, and p-
cresol are predicted to be distributed among 15 gene clusters.  The unusual coclustering
of genes associated with different pathways appears to have evolved in response to
similarities in biochemical mechanisms required for the degradation of intermediates in
different pathways.  A putative efflux pump and several hypothetical membrane-
associated proteins were identified and predicted to be involved in the transport of
aromatic compounds and/or intermediates in catabolism across the cell wall.  Several
genes associated with integration and recombination, including two group II intron-
associated maturases, were identified in the replication region, suggesting that pNL1 is
able to undergo integration and excision events with the chromosome and/or other
portions of the plasmid.  Conjugative transfer of pNL1 to another Sphingomonas sp. was
demonstrated, and genes associated with this function were found in two large clusters.
Approximately one-third of the ORFs (59 of them) have no obvious homology to known
genes.

<>

<1>Rong, X., Baysal, G.F., Meulia, T., McSpadden, G.B.B.
<2>Draft Genome Sequences of the Pseudomonas fluorescens Biocontrol Strains Wayne1R and Wood1R.
<3>J. Bacteriol.
<4>194
<5>724-725
<6>2012
<7>Pseudomonas fluorescens strains Wayne1R and Wood1R have proven capacities to improve plant
health. Here we report the draft genome sequences and
automatic annotations of both strains. Genome comparisons reveal
similarities with P. fluorescens strain Pf-5, reveal the novelty of
Wood1R, and indicate some genes that may be related to biocontrol.

<>

<1>Rong, X., Gurel, F.B., Meulia, T., McSpadden, G.B.B.
<2>Draft Genome Sequences of the Biocontrol Bacterium Mitsuaria sp. Strain H24L5A.
<3>J. Bacteriol.
<4>194
<5>734-735
<6>2012
<7>Mitsuaria sp. strain H24L5A is a plant-associated bacterium with proven capacities to suppress
plant pathogens. Here, we report the draft genome
sequences and automatic annotation of H24L5A. Comparative genomic analysis
indicates H24L5A's similarity to the Leptothrix and Methylibium species,
as well as several genes potentially contributing to its biocontrol
activities.

<>

<1>Ronka, J., Hjorleifsdottir, S., Tenkanen, T., Pitkanen, K., Mattila, P., Kristjansson, J.K.
<2>RmaI, a type II restriction endonuclease from Rhodothermus marinus which recognizes 5' CTAG 3'.
<3>Nucleic Acids Res.
<4>19
<5>2789
<6>1991
<7>RmaI, an isoschizomer of MaeI has been isolated from Rhodothermus marinus. RmaI recognizes the
palindromic sequence 5'-CTAG-3'.  Like its isoschizomer RmaI cleaves the sequence, C/TAG,
generating 5'-protruding TA-dinucleotides. The recognition sequence of RmaI was determined
using double digestions on pBR322- and phiX174 DNAs (figure 1, lanes 2-6 and 8-12).  The
cleavage patterns obtained were compared with computer-derived mapping data.  The data
predicts the sequence 5'-CTAG-3'.  The sequence was also tested by digesting Lambda- and
M13mp18 DNAs with RmaI (figure 1, lanes 13 and 14).  The fragments obtained matched the
computer predicted fragments that would be produced when cleaving at 5'-CTAG-3'.  RmaI has
the following number of recognition sites on these commonly used DNAs: pUC19, pBR322, phiX174,
SV40, M13mp18, T7, Lambda and Adeno2.  The cleavage site of RmaI was determined by cleavage of
primed synthesis reaction.  M13mp19 DNA containing the recognition site of RmaI was used as a
template.  Using an M13 sequencing primer the DNA was sequenced according to Sanger et al.  In
addition to the four standard reactions a fifth reaction was performed which was extended
through the RmaI recognition site. The reaction was terminated by heat treatment and the
product was cleaved with RmaI.  The cleaved product was divided in two.  The addition of
Klenow to one part resulted in a band migrating with the A-band (figure 2, lane +).  The
cleaved product (figure 2, lane -) migrated with the C-band.  Thus, the cleavage site for RmaI
is:
5'-C/TA-G-3'
3'-G-AT/C-5'.

<>

<1>Rosamond, J., Endlich, B., Linn, S.
<2>Electron microscopic studies of the mechanism of action of the restriction endonuclease of Escherichia coli B.
<3>J. Mol. Biol.
<4>129
<5>619-635
<6>1979
<7>Reaction intermediates and products formed by the restriction endonuclease of Escherichia coli
B with fd replicative form DNA substrates containing recognition sites in known positions and
orientations have been characterized by electron microscopy.  After exposure of these
substrates to enzyme, loops of duplex DNA were frequently observed, usually at or near the
termini.  Analysis of the size and structure of the loops observed with various DNA substrates
suggests that the enzyme binds initially to the recognition site then remains bound to the DNA
in the region of this site while tracking towards a site of cleavage.  Tracking appears to
occur only on the 5' side of the asymmetric recognition sequence, 5' . . .
T-G-A-(N)8-T-G-C-T . . . 3'; however, the location of the cleavage sites appears to be
random, at least within certain limits of distance from the recognition site.  Enzyme-DNA
complexes remain intact even after the double-strand cleavage is completed, and this complex
acts as a potent ATPase with no obvious function.  This latter reaction might represent an
artifactual uncoupling of ATP hydrolysis from the tracking of the enzyme along the DNA;
alternatively, it might indicate an in vivo function for the enzyme of which we are unaware.

<>

<1>Rosati, O., Srivastava, T.K., Katti, S.B., Alves, J.
<2>Importance of phosphate contacts for sequence recognition by EcoRI restriction enzyme.
<3>Biochem. Biophys. Res. Commun.
<4>295
<5>198-205
<6>2002
<7>We have studied the importance of charge and hydrogen-bonding potential of the phosphodiester
backbone for binding and cleavage by EcoRI restriction endonuclease.  We used 12-mer
oligodeoxynucleotide substrates with single substitutions of phosphates by chiral
methylphosphonates at each position of the recognition sequence -pGpApApTpTpCp-.  Binding was
moderately reduced between 4- and 400-fold more or less equally for the RP and SP-analogues
mainly caused by missing charge interaction.  The range of cleavage effects was much wider.
Four substrates were not cleaved at all.  At both flanking positions and in the purine half of
the sequence up to the central position, cleavage was more impaired than binding and
differences between RP and SP diastereomers were more pronounced.  These effects are easily
interpreted by direct phosphate contacts seen in the crystal structure.  For the effects of
substitutions in the pyrimidine half of the recognition sequence, more indirect effects have
to be discussed.

<>

<1>Rose, T.M., Schultz, E.R., Henikoff, J.G., Pietrokovski, S., McCallum, C.M., Henikoff, S.
<2>Consensus-degenerate hybrid oligonucleotide primers for amplification of distantly related sequences.
<3>Nucleic Acids Res.
<4>26
<5>1628-1635
<6>1998
<7>We describe a new primer design strategy for PCR amplification of unknown targets that are
related to multiply-aligned protein sequences.  Each primer consists of a short 3' degenerate
core region and a longer 5' consensus clamp region. Only 3-4 highly conserved amino acid
residues are necessary for design of the core, which is stabilized by the clamp during
annealing to template molecules.  During later rounds of amplification, the non-degenerate
clamp permits stable annealing to product molecules.  We demonstrate the practical utility of
this hybrid primer method by detection of diverse reverse transcriptase-like genes in a human
genome, and by detection of C5 DNA methyltransferase homologs in various plant DNAs.  In each
case, amplified products were sufficiently pure to be cloned without gel fractionation.  This
Consensus-Degenerate Hybrid Oligonucleotide Primer strategy has been implemented as a computer
program that is accessible over the World Wide Web (http://blocks.fhcrc.org/codehop.html) and
is directly linked from the BlockMaker multiple sequence alignment site for hybrid primer
prediction beginning with a set of related protein sequences.

<>

<1>Rosen, L.E., Morrison, H.A., Masri, S., Brown, M.J., Springstubb, B., Sussman, D., Stoddard, B.L., Seligman, L.M.
<2>Homing endonuclease I-CreI derivatives with novel DNA target specificities - mutant homing enzyme via mutagenesis for DNA cleavage and gene therapy.
<3>Nucleic Acids Res.
<4>34
<5>4791-4800
<6>2006
<7>Homing endonucleases are highly specific enzymes, capable of recognizing and cleaving unique
DNA sequences in complex genomes. Since such DNA
cleavage events can result in targeted allele-inactivation and/or
allele-replacement in vivo, the ability to engineer homing endonucleases
matched to specific DNA sequences of interest would enable powerful and
precise genome manipulations. We have taken a step-wise genetic approach
in analyzing individual homing endonuclease I-CreI protein/DNA contacts,
and describe here novel interactions at four distinct target site
positions. Crystal structures of two mutant endonucleases reveal the
molecular interactions responsible for their altered DNA target
specificities. We also combine novel contacts to create an endonuclease
with the predicted target specificity. These studies provide important
insights into engineering homing endonucleases with novel target
specificities, as well as into the evolution of DNA recognition by this
fascinating family of proteins.

<>

<1>Rosenberg, A.H., Simon, M.N., Studier, F.W., Roberts, R.J.
<2>Survey and mapping of restriction endonuclease cleavage sites in bacteriophage T7 DNA.
<3>J. Mol. Biol.
<4>135
<5>907-915
<6>1979
<7>A survey of restriction endonucleases having different cleavage specificities
has identified 10 that do not cut wild-type bacteriophage T7 DNA, 11 that cut
at six or fewer sites, four that cut at 18 to 45 sites, and 12 that cut at more
than 50 sites.  All the cleavage sites for the 13 enzymes that cut a 26 or
fewer sites have been mapped.  Cleavage sites for each of the 10 enzymes that
do not cut T7 DNA would be expected to occur an average of 9 to 10 times in a
random nucleotide sequence the length of T7 DNA.  A possible explanation for
the lack of any cleavage sites for these enzymes might be that T7 encounters
enzymes having these specificities in natural hosts, and that the sites have
been eliminated from T7 DNA by natural selection.  Five restriction
endonucleases were found to cut within the terminal repetition of T7 DNA; one
of these, KpnI, cuts at only three additional sites in the T7 DNA molecule.
The length of the terminal repetition was estimated by two independent means to
be approximately 155 to 160 base-pairs.

<>

<1>Rosenberg, J., Wang, B., Frederick, C., Reich, N., Greene, P., Grable, J., McClarin, J.
<2>Development of a Protein Design Strategy for EcoRI Endonuclease.
<3>Protein Eng., A.R. Liss Inc., Oxender, D., Fox, C.F., 
<4>0
<5>237-250
<6>1987
<7>The 3 angstrom structure of a co-crystalline recognition complex between EcoRI
endonuclease and the cognate oligonucleotide TCGCGAATTCGCG has been solved.
Each subunit of the endonuclease is organized into a domain with a/b
architecture.  The b-sheet consists of anti-parallel and parallel motifs.  The
amino acid residues which interact directly with the DNA to perform the
hydrolysis are located primarily in the anti-parallel motif, while those which
form sequence specific hydrogen bonds with the bases are found in the parallel
motif.  The hydrolytic active site is located in a cleft which is not fully
assembled in this structure (which was obtained in the absence of magnesium).
The DNA conformation departs significantly from those which have been observed
in crystals containing pure DNA; suggesting that the altered conformation has
been stabilized by the enzyme.  The conformations seen only upon protein
binding are termed neo-conformations to distinguish them from those which are
intrinsically stable in the absence of protein.  The determinants of sequence
specificity include "modular" interactions based on the crossover
alpha-helices, i.e., those which connect the b-strands of the parallel segment
of the principal B-sheet.  They are pointing into the major groove of the DNA
and amino acid side chains at the amino ends of these helices form bidentate
hydrogen bonding interactions with the bases.  The inner recognition module
consists of two symmetry-related alpha-helices which recognize the inner
tetranucleotide (AATT), while the two symmetry-equivalent outer recognition
modules are single alpha-helices which recognize the GC base pairs.

<>

<1>Rosenberg, J.M.
<2>Structure and function of restriction endonucleases.
<3>Curr. Opin. Struct. Biol.
<4>1
<5>104-113
<6>1991
<7>The past year has seen significant advances in our understanding of the
structure and function of restriction endonucleases.  The highlights include a
revised chain tracing for EcoRI endonuclease from Escherichia coli, structures
soon to be reported for E. coli EcoRV endonuclease and significant advances in
the biochemistry and molecular genetics of both enzymes.

<>

<1>Rosenberg, J.M., Boyer, H.W., Greene, P.
<2>The structure and function of the EcoRI restriction endonuclease.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>131-164
<6>1981
<7>*
  I. DNA mapping
 II. X-ray diffraction analysis of EcoRI endonuclease crystals
III. DNA binding properties of EcoRI methylase and endonuclease
 IV. Degeneracies in restriction recognition sequences


<>

<1>Rosenberg, J.M., Dickerson, R.E., Greene, P.J., Boyer, H.W.
<2>Preliminary X-ray diffraction analysis of crystalline EcoRI endonuclease.
<3>J. Mol. Biol.
<4>122
<5>241-245
<6>1978
<7>EcoRI endonuclease crystallizes in space group C2 with unit cell parameters a =
209 angstroms, b = 129 angstroms, c = 50 angstroms and B = 98.4o.  Four 29,000
molecular weight subunits per asymmetric unit would give a reasonable Vm value
of 2.87 cubic angstroms/dalton.  EcoRI endonuclease is the first protein which
recognizes a specific sequence of bases in DNA to be crystallized in a form
suitable for high resolution structure analysis.

<>

<1>Rosenberg, J.M., Greene, P.
<2>EcoRI* specificity and hydrogen bonding.
<3>DNA
<4>1
<5>117-124
<6>1982
<7>Under standard conditions, EcoRI endonuclease uniquely recognizes the inverted
repeat GAATTC.  However, this specificity breaks down under non-standard
conditions into what has been termed EcoRI* specificity, wherein many other
sequences are recognized.  We show here that the hydrolysis rates at all known
EcoRI* sites can be summarized by the hierarchies: G > > A > T > > C at the
first position, A > > [G,C] > > T at the second and third position, and the
corresponding complements at the last three positions.  This is consistent with
a recognition model which assumes that there are two specific hydrogen bonds
per base pair under standard conditions.  One or more of these are randomly
replaced by water under EcoRI* conditions and the position of a sequence within
the appropriate bonds are common recognition features that can be identified by
examining the DNA.  The recognition points thereby identified for EcoRI all
fall within the major groove of the DNA.

<>

<1>Rosenberg, J.M., McClarin, J., Grable, J., Frederick, C., Samudzi, C., Jen-Jacobson, L.
<2>Structure of DNA-EcoRI endonuclease complex at 3 angstroms resolution.
<3>J. Cell Biochem. Suppl.
<4>9B
<5>101
<6>1985
<7>The 3 angstrom structure of a co-crystalline recognition complex between EcoRI
endonuclease and the oligonucleotide TCGCGAATTCGCG will be reported.  The DNA
and the protein share a common (crystallographic) two-fold axis of rotational
symmetry, as expected from the intrinsic symmetry of the recognition site
(GAATTC).  The DNA conformation within the complex is different from that found
in the absence of protein suggesting that the set of conformational states
which are accessable to protein-free DNA is expanded by the binding of sequence
specific proteins.  These include the torsional neo-1 kink which widens the
major groove by unwinding the DNA by approximately 25o thereby facilitating
contact between the edges of the purine bases and amino acid side chains.  A
second (neo-2) kink is located three base-pairs away which also facilitates
access of protein to DNA and/or has a role in the hydrolytic mechanism of this
enzyme.  A five stranded a/b structure forms the foundation of each
endonuclease subunit with the strands of beta-sheet and the alpha-helices
oriented approximately perpendicular to the average DNA helix axis.
Polypeptide loops at the carboxy edge of the beta-sheet form a cleft which
contains the segment of DNA backbone spanning the scissile bond.  (DNA
hydrolysis was inhibited via omission of Mg+2).  The cleft is complementary to
one strand of double helical DNA and its shape determined by the intrinsic
twist of b-sheet.  This novel feature suggests that DNA and protein are
intrinsically complementary at fundamental level.  Sequence specificity is
determined by "modular" interactions.  One large symmetric module recognizes
the inner tetranucleotide (AATT while two additional symmetry-equivalent
modules recognizes the outer base pairs.  The inner module consists of two
symmetry equivalent alpha-helices which project from the a/b units.  Lysine and
glutamic acid side chains at the ends of the alpha-helices hydrogen bond to the
adenine residues thereby determining the specificity for the inner
tetranucleotide.  The outer module is formed by a separate segment of the
polypeptide chain containing an arginine side chain which hydrogen bonds to
guanine.

<>

<1>Rosenberg, J.M., McClarin, J.A., Frederick, C.A., Grable, J., Boyer, H.W., Greene, P.J.
<2>Structure and function of the EcoRI restriction endonuclease.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>119-145
<6>1987
<7>The highly specific recognition of the double-stranded sequence d(GAATTC) by
EcoRI endonuclease offers compelling advantages as a system for investigating
sequence specific recognition of DNA.  It is a small protein (31,065 daltons,
276 amino acids) of known sequence which forms highly stable catalytically
active dimers in solution.  The enzyme hydrolyzes the phosphodiester bond
between the guanylic and adenylic acid residues resulting in a 5'-phosphate.
The reaction proceeds with inversion of configuration at the reactive
phosphorus, implying that there is an odd number of chemical events during the
hydrolysis.  The simplest interpretation of this observation is that the enzyme
does not form a covalent intermediate with the DNA.  Although EcoRI
endonuclease requires Mg2+ for phosphodiester bond hydrolysis, it binds
specifically to its cognate hexanucleotide in the absence of Mg2+ with a
dissociation constant on the order of 10-11 M-1.  The enzyme also binds DNA in
a nonspecific manner ie., at sites other than GAATTC; this does not result in
hydrolysis of the DNA.  It has been postulated that the nonspecific complex
enhances the rate of formation of formation of the specific complex by
facilitated diffusion along the DNA.  Both the EcoRI endonuclease and the EcoRI
methylase recognize the same hexanucleotide; however, the latter methylates the
central adenine residues of both strands at the exocyclic N-6 amino group.
When either one or both groups is methtylated the endonuclease no longer
cleaves the DNA.  Thus, EcoRI endonuclease not only discriminates between its
hexanucleotide and all other hexanucleotides, it also discriminates between
different methtylation states of the same hexanucleotide.  Cocrystals of DNA
and protein that diffract to high resolution are required for a full
understanding of sequence specificity in the EcoRI system.  We have obtained
cocrystals and determined their structure.  Here we report the structure of the
recognition complex including interactions involved in sequence specificity.

<>

<1>Rosenberg, J.M., McClarin, J.A., Frederick, C.A., Wang, B.-C., Boyer, H.W., Grable, J., Greene, P.
<2>The structure and function of EcoRI endonuclease.
<3>Biological Organization: Macromolecular Interactions at High Resolution., Academic Press, Burnett, R.M., Vogle, H.J., 
<4>0
<5>11-43
<6>1987
<7>The 3 angstrom structure of a co-crystalline recognition complex between EcoRI
endonuclease and the cognate oligonucleotide TCGCGAATTCGCG has been solved by
the ISIR method using a platinum isomorphous derivative.  Each subunit of the
endonuclease is organized into an a/b domain based on a five stranded
beta-sheet and an extension, called the "arm" which wraps around the DNA.  The
primary beta-sheet consists of anti-parallel and parallel motifs which contain
the sites of DNA strand scission and sequence specific recognition,
respectively.  The hydrolytic active site is located in a cleft which binds the
DNA backbone in the vicinity of the scissile bond.  The DNA conformation
departs significantly from those which have been observed in crystals
containing pure DNA; suggesting that the altered conformation has been
stabilized by the enzyme.  The conformations seen only upon protein binding are
termed neo-conformations to distinguish them from those which are intrinsically
stable in the absence of protein.  Sequence specificity is determined by
"modular" interactions based on the crossover alpha-helices, i.e., those which
connect the beta-strands of the parallel segment of the principal beta-sheet.
They are pointing into the major groove of the DNA and amino acid side chains
at the amino ends of these helices form bidentate interactions with the bases.
The inner recognition module consists of two symmetry-related alpha-helices
which recognize the inner tetranucleotide (AATT), while the two
symmetry-equivalent outer recognition modules are single alpha-helices wihch
recognize the GC base pairs.

<>

<1>Rosenberg, J.M., McClarin, J.A., Frederick, C.A., Wang, B.-C., Boyer, H.W., Greene, P.
<2>The 3 angstrom structure of a DNA-EcoRI endonuclease recognition complex.
<3>Chem. Scr.
<4>26B
<5>147-157
<6>1986
<7>The 3 angstrom structure of a co-crystalline recognition complex between EcoRI
endonuclease and the cognate oligonucleotide TCGCGAATTCGCG has been solved by
the ISIR method using a platinum isomorphous derivative.  Each subunit of the
endonuclease is organized into an a/b domain based on a five stranded
beta-sheet and an extension, called the "arm", which wraps around the DNA.  The
primary beta-sheet consists of anti-parallel and parallel sub-domains which
contain the sites of DNA strand scission and sequence specific recognition,
respectively.  The hydrolytic active site is located in a cleft which binds the
DNA backbone in the vicinity of the scissile bond.  The DNA conformation
departs significantly from those which have been observed in crystals
containing pure DNA; suggesting that the altered conformation has been
stabilized by the enzyme.  The conformations seen only upon protein binding are
termed neo-conformations to distinguish them from those which are intrinsically
stable in the absence of protein.  Sequence specificity is determined by
"modular" interactions based on the crossover alpha-helices, i.e., those which
connect the beta-strands of the parallel segment of the principal beta-sheet.
They are pointing into the major groove of the DNA and amino acid side chains
at the amino ends of these helices form bidentate interactions with the bases.
The inner recognition module consists of two symmetry-related alpha-helices
which recognize the inner tetranucleotide (AATT), while the two
symmetry-equivalent outer recognition modules are single alpha-helices which
recognize the GC base pairs.

<>

<1>Rosenberg, J.M., McClarin, J.A., Frederick, C.A., Wang, B.-C., Grable, J., Boyer, H.W., Greene, P.
<2>Structure and recognition mechanism of EcoRI endonuclease.
<3>Trends Biochem. Sci.
<4>12
<5>395-398
<6>1987
<7>The structure of a complex between EcoRI endonuclease and a cognate
oligonucleotide shows that sequence specificity is mediated by 12 protein-DNA
hydrogen bonds.  These interactions discriminate the EcoRI recognition site
from all other sequences because any base substitution would rupture at least
one of these hydrogen bonds.

<>

<1>Rosenberg, J.M., McClarin, J.A., Frederick, C.A., Wang, B.-C., Grable, J., Boyer, H.W., Greene, P.
<2>Structure of the DNA-EcoRI endonuclease recognition complex.
<3>Structure, Dynamics and Function of Biomolecules, Springer-Verlag, Ehrenberg, A., New York
<4>0
<5>255-259
<6>1987
<7>The 3 angstrom structure of the co-crystalline recognition complex between
EcoRI endonuclease and the cognate oligonucleotide TCGCGAATTCGCG has been
solved by the ISIR method using a platinum isomorphous derivative.  Refinement
is in progress.  The endonuclease-DNA recognition complex consists of a
distorted double helix and a protein dimer composed of identical subunits
related by a two-fold axis of rotational symmetry.  The distortions of the DNA
are induced by the binding of the protein.  They are concentrated into separate
features which are localized disruptions of the double helical symmetry.  These
disruptions appear to have structural consequences which propagate over long
distances through the DNA via twisting and perhaps bending effects.  They are
therefore referred to as neo-kinks.  The Type-I neo-kink spans the central
two-fold symmetry axis of the complex and it introduces a net unwinding of 25
degrees into the DNA.  This increases the separation of the DNA backbones
across the major groove thereby facilitating access by the protein to the base
edges, which are at the floor of the groove.  The Type-I neo-kink also realigns
adjacent adenine residues within the central AATT tetranucleotide so as to
create the detailed geometry necessary for amino acid side chains to bridge
across these purines.

<>

<1>Rosenberg, S.M.
<2>EcoK restriction during in vitro packaging of coliphage lambda DNA.
<3>Gene
<4>39
<5>313-315
<6>1985
<7>The K restriction system of Escherichia coli works in vitro [Meselson and Yuan,
Nature 217 (1968) 1110-1114].  E. coli C lacks the K restriction system.  I
show that in vitro packaging in standard E. coli K-12-derived systems effects a
loss of plaque-former output from K-unmodified lambda DNA relative to
K-modified lambda DNA when compared with packaging in the E. coli C-derived
system of Rosenberg et al. [Gene 38 (1985) 165-175].  I conclude that the EcoK
restriction system is active in standard in vitro packaging systems.  EcoK
restriction during in vitro packaging could specifically depress recovery of
some lambdaand cosmid clones of eukaryotic DNA or any other DNA not modified
for EcoK restriction.

<>

<1>Rosenstein, R., Nerz, C., Biswas, L., Resch, A., Raddatz, G., Schuster, S.C., Gotz, F.
<2>Genome analysis of the meat starter culture bacterium Staphylococcus carnosus TM300.
<3>Appl. Environ. Microbiol.
<4>75
<5>811-822
<6>2009
<7>The Staphylococcus carnosus genome has the highest GC content of all
sequenced staphylococcal genomes, with 34.6%, and therefore represents a
species that is set apart from S. aureus, S. epidermidis, S.
saprophyticus, and S. haemolyticus. With only 2.56 Mbp, the genome belongs
to a family of smaller staphylococcal genomes, and the ori and ter regions
are asymmetrically arranged with the replichores I (1.05 Mbp) and II (1.5
Mbp). The events leading up to this asymmetry probably occurred not that
long ago in evolution, as there was not enough time to approach the
natural tendency of a physical balance. Unlike the genomes of pathogenic
species, the TM300 genome does not contain mobile elements such as
plasmids, insertion sequences, transposons, or STAR elements; also, the
number of repeat sequences is markedly decreased, suggesting a
comparatively high stability of the genome. While most S. aureus genomes
contain several prophages and genomic islands, the TM300 genome contains
only one prophage, PhiTM300, and one genomic island, nuSCA1, which is
characterized by a mosaic structure mainly composed of species-specific
genes. Most of the metabolic core pathways are present in the genome. Some
open reading frames are truncated, which reflects the nutrient-rich
environment of the meat starter culture, making some functions
dispensable. The genome is well equipped with all functions necessary for
the starter culture, such as nitrate/nitrite reduction, various sugar
degradation pathways, two catalases, and nine osmoprotection systems. The
genome lacks most of the toxins typical of S. aureus as well as genes
involved in biofilm formation, underscoring the nonpathogenic status.

<>

<1>Rosenthal, A.Z., Matson, E.G., Eldar, A., Leadbetter, J.R.
<2>RNA-seq reveals cooperative metabolic interactions between two termite-gut spirochete species in co-culture.
<3>ISME J.
<4>5
<5>1133-1142
<6>2011
<7>The hindguts of wood-feeding termites typically contain hundreds of
microbial species. Together with their insect host, these gut microbes
degrade lignocellulose into usable catabolites. Although past research
revealed many facets of the stepwise flow of metabolites in this scheme,
not much is known about the breadth of interactions occurring between
termite-gut microbes. Most of these microbes are thought to depend on, and
to have co-speciated with, their host and each other for millions of
years. In this study, we explored the interactions of two spirochetes
previously isolated from the very same termite species. As hydrogen (H(2))
is the central free intermediate in termite-gut lignocellulose digestion,
we focused on interactions between two closely related termite-gut
spirochetes possessing complementary H(2) physiologies: one produces H(2),
while the other consumes it. In vitro, these two Treponema species
markedly enhanced each other's growth. RNA sequencing resolved the
transcriptomes of these two closely related organisms, revealing that
co-cultivation causes comprehensive changes in global gene expression. The
expression of well over a 100 genes in each species was changed >twofold,
with over a dozen changed >10-fold. Several changes implicating
synergistic cross-feeding of known metabolites were validated in vitro.
Additionally, certain activities beneficial to the host were
preferentially expressed during consortial growth. However, the majority
of changes in gene expression are not yet understandable, but indicate a
broad, comprehensive and mutualistic interaction between these closely
related, co-resident gut symbionts. The results suggest that staggeringly
intricate networks of metabolic and gene interactions drive lignocellulose
degradation and co-evolution of termite gut microbiota.

<>

<1>Rosewarne, C.P., Cheung, J.L., Smith, W.J., Evans, P.N., Tomkins, N.W., Denman, S.E., O, C.P., Morrison, M.
<2>Draft Genome Sequence of Treponema sp. Strain JC4, a Novel Spirochete Isolated from the Bovine Rumen.
<3>J. Bacteriol.
<4>194
<5>4130
<6>2012
<7>Morphologically and biochemically diverse members of the Treponema genus are present in the
gastrointestinal tract of ruminants, yet very little is understood
about their functional importance to this microbiome. Here we describe the
annotated draft genome sequence of Treponema sp. strain JC4, a novel spirochete
isolated from a bovine rumen sample.

<>

<1>Rosewarne, C.P., Greenfield, P., Li, D., Tran-Dinh, N., Bradbury, M.I., Midgley, D.J., Hendry, P.
<2>Draft Genome Sequence of Clostridium sp. Maddingley, Isolated from Coal-Seam Gas  Formation Water.
<3>Genome Announcements
<4>1
<5>e00081-12
<6>2013
<7>Clostridium sp. Maddingley was isolated as an axenic culture from a brown coal-seam formation
water sample collected from Victoria, Australia. It lacks the solventogenesis genes found in
closely related clostridial strains. Metabolic reconstructions suggest that volatile fatty
acids are the main fermentation end products.

<>

<1>Rosewarne, C.P., Greenfield, P., Li, D., Tran-Dinh, N., Midgley, D.J., Hendry, P.
<2>Draft Genome Sequence of Methanobacterium sp. Maddingley, Reconstructed from Metagenomic Sequencing of a Methanogenic Microbial Consortium Enriched from  Coal-Seam Gas Formation Water.
<3>Genome Announcements
<4>1
<5>e00082-12
<6>2013
<7>The draft genome of Methanobacterium sp. Maddingley was reconstructed from metagenomic
sequencing of a methanogenic microbial consortium enriched from coal-seam gas formation water.
It is a hydrogenotrophic methanogen predicted to grow using hydrogen and carbon dioxide.

<>

<1>Rosinski-Chupin, I., Sauvage, E., Mairey, B., Mangenot, S., Ma, L., Da Cunha, V., Rusniok, C., Bouchier, C., Barbe, V., Glaser, P.
<2>Reductive evolution in Streptococcus agalactiae and the emergence of a host adapted lineage.
<3>BMC Genomics
<4>14
<5>252
<6>2013
<7>BACKGROUND: During host specialization, inactivation of genes whose function is
no more required is favored by changes in selective constraints and evolutionary
bottlenecks. The Gram positive bacteria Streptococcus agalactiae (also called
GBS), responsible for septicemia and meningitis in neonates also emerged during
the seventies as a cause of severe epidemics in fish farms. To decipher the
genetic basis for the emergence of these highly virulent GBS strains and of their
adaptation to fish, we have analyzed the genomic sequence of seven strains
isolated from fish and other poikilotherms. RESULTS: Comparative analysis shows
that the two groups of GBS strains responsible for fish epidemic diseases are
only distantly related. While strains belonging to the clonal complex 7 cannot be
distinguished from their human CC7 counterparts according to their gene content,
strains belonging to the ST260-261 types probably diverged a long time ago. In
this lineage, specialization to the fish host was correlated with a massive gene
inactivation and broad changes in gene expression. We took advantage of the low
level of sequence divergence between GBS strains and of the emergence of
sublineages to reconstruct the different steps involved in this process.
Non-homologous recombination was found to have played a major role in the genome
erosion. CONCLUSIONS: Our results show that the early phase of genome reduction
during host specialization mostly involves accumulation of small and likely
reversible indels, followed by a second evolutionary step marked by a higher
frequency of large deletions.

<>

<1>Roske, K., Calcutt, M.J., Wise, K.S.
<2>The Mycoplasma fermentans prophage phiMFV1: genome organization, mobility and variable expression of an encoded surface protein.
<3>Mol. Microbiol.
<4>52
<5>1703-1720
<6>2004
<7>The approximately 16 kb genome of the Mycoplasma fermentans phiMFV1 prophage is described, and
its mobility, replication and effect on the
mycoplasma surface phenotype are demonstrated. In various M. fermentans
strains, phiMFV1 was either absent or integrated at diverse (and sometimes
multiple) chromosomal sites, each marked by a conserved TTTTTA target
sequence that is duplicated upon integration. Precise excision,
replication of an extrachromosomal form and loss of phiMFV1 from the
mycoplasmal genome were documented in a series of clonal derivatives of M.
fermentans propagated in culture. Of 18 open reading frames (ORFs) encoded
by phiMFV1, most can be ascribed functions related to phage biology,
whereas one encodes a unique coiled-coil membrane surface protein, Mem,
that was confirmed to be expressed in propagating populations of M.
fermentans. With the exception of Mem and other minor ORFs, the striking
similarity between the deduced proteomes of phiMFV1 and the recently
described phiMAV1 of arthritogenic strains of Mycoplasma arthritidis,
along with the prominent gene synteny between these elements, provides the
taxonomic basis for a new family of prophage. Their coding features are
consistent with long-term residence in mycoplasma genomes and the
divergence of species within a phylogenetic clade of mycoplasmas. The
unique Mem protein expressed from phiMFV1 and the unique hypothetical
surface lipoproteins encoded by phiMAV1 and phiMFV1 also suggest that
prophage-associated genes may provide specific, selectable phenotypic
traits during co-evolution of mycoplasma species with their respective
mammalian hosts. Retention of these labile prophage elements in organisms
with such drastically reduced genome sizes implies a significant role in
adaptation and survival.

<>

<1>Rosner, J.L.
<2>Modification-deficient mutants of bacteriophage Pl.  I. Restriction by Pl cryptic lysogens.
<3>Virology
<4>52
<5>213-222
<6>1973
<7>The Pl restriction-modification system is responsible for the inefficient
plating of c2 and c3 mutants of bacteriophage Pl on Pl cryptic lysogens.  Pl
cryptic is a defective prophage which does not express Pl immunity but which
does express Pl modification and restriction.  The rare c2 or c3 phage which do
grow on the Pl cryptic lysogens [abbreviated (Plcry)], lose their ability to do
so after growth on a nonlysogenic host.  Temperature-sensitive c2 mutants grown
on a nonlysogenic host at the permissive temperature plate efficiently on
(Plcry).  If grown at a nonpermissive temperature, however, the c2 ts mutants
plate inefficiently on (Plcry).  Plr-m+, a nonrestricting P1 phage, plates
efficiently on (Plcry), but Plr-m-, which neither restricts nor modifies,
plates inefficiently on (Plcry).  These results are explained as follows:  Pl
DNA is itself a substrate for the Pl directed modification-restriction system.
Normally, during lytic growth, Pl DNA is modified.  However, Plr-m-, c2, and c3
mutants are modification-defective.  Thus, when their unmodified DNA enters
(Plcry), it is subject to Pl restriction.  Direct evidence for this hypothesis
was obtained from experiments in which the abilitiy of Pl phage to modify
lambda was studied.  Plr-m-, c2 and c3 mutants cannot modify lambda whereas Pl
wild type and Plr-m+ are able to do so.  Furthermore, c2 and c3 mutants can
complement each other to express Pl modification.  Plr-m- is not complemented
by either c2 or c3 mutants.  It is concluded that c2 and c3 are two cistrons
required for P1 modification.  Plr-m- may be missing, or unable to transcribe,
the c2 and c3 genes.

<>

<1>Ross, B.C.
<2>P. gingivalis polynucleotides and uses thereof.
<3>US Patent Office
<4>US 6444799 A
<5>
<6>2002
<7>The present invention relates to isolated Porphorymonas gingivalis polynucleotides.  The
polynucleotides comprises a continguous sequence of at least 20 nucleotides which is identical
to a contiguous sequence of at least 20 nucleotides within a sequence selected from the group
consisting of SEQ. ID. NO. 1 to SEQ. ID. NO. 1120 and sequences complementary thereto.

<>

<1>Ross, D.W.
<2>Restriction enzymes.
<3>Arch. Pathol. Lab. Med.
<4>114
<5>906
<6>1990
<7>None

<>

<1>Ross, T.K., Achberger, E.C., Braymer, H.D.
<2>Characterization of the Escherichia coli modified cytosine restriction (mcrB) gene.
<3>Gene
<4>61
<5>277-289
<6>1987
<7>The McrB restriction system of Escherichia coli K-12 is responsible for the
inactivation of 5-methylcytosine-containing DNA.  The mcrB mutation of E. coli
strain K802 was complemented by hybrid plasmid pUC9-14 which consists of a
5.5-kb BglII-EcoRI fragment from the E. coli K-12 chromosome cloned in pUC9
(Ross and Braymer, 1987).  The limits of the mcrB gene within the 5.5-kb insert
were defined by deletion portions of the fragment and assaying for McrB
restriction of M. AluI-methylated DNA.   A 51-kDa polypeptide was identified as
the mcrB gene product based on an analysis of maxicell-labeled polypeptides
from pUC9-14 and deletion derivatives of this plasmid.  Deletion analyses and
transcription initiation assays enabled us to determine the direction of
transcription and translation of mcrB.  Transcription initiates approx. 710 bp
beyond the end of the hsdS gene, and proceeds in the same direction as the
transcription of the hsdR, hsdM, and hsdS genes, which is clockwise on the
conventional E. coli map.

<>

<1>Ross, T.K., Achberger, E.C., Braymer, H.D.
<2>Nucleotide sequence of the McrB region of Escherichia coli K-12 and evidence for two independent translational initiation sites at the mcrB locus.
<3>J. Bacteriol.
<4>171
<5>1974-1981
<6>1989
<7>The McrB restriction system of Escherichia coli K-12 is responsible for the
biological inactivation of foreign DNA that contains 5-methylcytosine residues.
Within the McrB region of the chromosome is the mcrB gene, which encodes a
protein of 51 kilodaltons (kDa), and the mcrC gene, the product of which is 39
kDa.  The nucleotide sequence of a 2695-base-pair segment encompassing the McrB
region was determined.  The deduced amino acid sequence was used to identify
two open reading frames specifying peptides of 455 and 348 amino acids,
corresponding to the products of the mcrB and mcrC genes, respectively.  A
single-nucleotide overlap was found to exist between the termination codon of
the mcrB gene and the proposed initiation codon of the mcrC gene.  The presence
of an additional peptide of 33 kDa in strains containing various recombinant
plasmids with portions of the McrB region has been reported by Ross et al.  The
analysis of frameshift and deletion mutants of one such hybrid plasmid,
pRAB-13, provided evidence for a second translational initiation site within
the McrB open reading frame.  The proposed start codon for translation of the
33-kDa peptide lies 481 nucleotides downstream from the initation codon for the
51-kDa mcrB gene product.  The 33-kDa peptide may play a regulatory role in the
McrB restriction of DNA containing 5-methylcytosine.

<>

<1>Ross, T.K., Achberger, E.C., Braymer, H.D.
<2>Identification of a second polypeptide required for McrB restriction of 5-methylcytosine-containing DNA in Escherichia coli K12.
<3>Mol. Gen. Genet.
<4>216
<5>402-407
<6>1989
<7>The McrB restriction system in Escherichia coli K12 causes sequence-specific
recognition and inactivation of DNA containing 5-methylcytosine residues.  We
have previously located the mcrB gene near hsdS at 99 min on the E. coli
chromosome and demonstrated that it encodes a 51 kDa polypeptide required for
restriction of M.AluI methylated (A-G-5mC-T) DNA.  We show here, by analysis of
maxicell protein synthesis of various cloned fragments from the mcrB region,
that a second protein of approximately 39 kDa is also required for
McrB-directed restriction.  The new gene, designated mcrC, is adjacent to mcrB
and located distally to hsdS.  The McrB phenotype has been correlated
previously with restriction of 5-hydroxy-methylcytosine (HMC)-containing T-even
phage DNA that lacks the normal glucose modification of HMC, formally
designated RglB (for restriction of glucoseless phage).  This report reveals a
difference between the previously correlated McrB and RglB restriction systems:
while both require the mcrB gene product only the McrB system requires the
newly identified mcrC-encoded 39-kDa polypeptide.

<>

<1>Ross, T.K., Braymer, H.D.
<2>Localization of a genetic region involved in McrB restriction by Escherichia coli K-12.
<3>J. Bacteriol.
<4>169
<5>1757-1759
<6>1987
<7>A 5,500-base-pairs BglII-EcoRI fragment proximal to the hsd genes of
Escherichia coli K-12 has been cloned in the plasmid vector pUC9.  The
resultant hybrid plasmid was shown to complement the mcrR mutation of E. coli
K802.  The presence of the hybrid plasmid in strain K802 caused an 18.3-fold
drop in transformation efficiency with AluI-methylated pACYC184 relative to
unmethylated pACYC184.  These results indicate that the cloned DNA is involved
in the McrB system of restriction of 5-methylcytosine DNA.

<>

<1>Rossi, S., Buera, M.P., Moreno, S., Chirife, J.
<2>Stabilization of the restriction enzyme EcoRI dried with Trehalose and other selected glass-forming solutes.
<3>Biotechnol. Prog.
<4>13
<5>609-616
<6>1997
<7>The stabilization of the restriction enzyme EcoRI by its incorporation into aqueous
glass-forming carbohydrate or polymer solutions, followed by vacuum-drying to low moisture,
has been studied.  Glass-forming solutes included trehalose, sucrose, lactose, maltose,
raffinose, maltodextrin, DE 10, and poly(vinylpyrrolidone) (molecular weight 40,000, PVP).
Among the solutes examined, trehalose and sucrose protected the enzyme most effectively during
storage at 37 and 45 C.  The restriction enzyme dried with trehalose or sucrose maintained its
activity without detectable loss for at least 20 days at 37 C and 12 days at 45 C.  In
contrast, the activity of the enzyme dried with maltodextrin or PVP was reduced during vacuum
desiccation and also it decreased remarkably during storage at the same temperatures.  Stored
(37/45 C) vacuum-dried trehalose and sucrose systems were either a dense paste or a very
viscous syrup, and this indicated that they were not glassy.  Moreover, no relationship was
found between the glass transition temperatures (Tg) of the pure added solute and enzyme
protection during storage, since, e.g., sucrose which has significantly lower Tg values
protected the enzyme much better than either maltose, lactose, maltodextrin, or PVP.  The
trisaccharide raffinose offered good protection of enzyme activity, and its role as a novel
excipient matrix for labile enzyme stabilization deserves further investigation.  The
stability of enzyme EcoRI was rapidly lost when the vacuum-dried trehalose and sucrose systems
were humidified to 58% relative humidity and stored at 45 C, and this was attributed to
disaccharide crystallization.

<>

<1>Rossino, R., Gosalvez, J., Mezzanotte, R.
<2>The effects of enzyme inactivation and incubation buffer on digestion in situ with restriction endonucleases.
<3>Biotech. Histochem.
<4>73
<5>325-328
<6>1998
<7>Previous studies have shown that components of the incubation reaction other than the
restriction endonucleases in an in situ restriction enzyme digest of chromosomes may induce
G-like banding patterns.  To determine whether factors other than DNA base composition play a
role in determining restriction enzyme induced bands, we investigated the effect of reaction
buffers alone or in the presence of heat inactivated enzymes.  Our results show that enzymes
such as AluI, RsaI and MspI become inactivated during 3-24 hr incubations at 37 C and that
reaction buffers alone failed to produce G-like bands when inactive endonucleases were
included.

<>

<1>Rosubatchi, V., Harettosukai, A.
<2>Sequences for detection and identification of methicillin-resistant Staphylococcus aureus.
<3>Japanese Patent Office
<4>JP 2004534537 A
<5>
<6>2004
<7>
<>

<1>Roszczyk, E., Goodgal, S.
<2>Methylase Activities from Haemophilus influenzae that protect Haemophilus parainfluenzae Transforming Deoxyribonucleic Acid from Inactivation by Haemophilus influenzae Endonuclease R.
<3>J. Bacteriol.
<4>123
<5>287-293
<6>1975
<7>Specific methylases that have the properties of deoxyribonucleic acid (DNA)
modification enzymes have been isolated from Haemophilus influenzae strain Rd.
Two activities (methylase IIa and methylase III) were found to protect
transforming DNA of H. parainfluenzae from the action of H. influenzae
restriction enzymes.  To determine the specificity of the protection, a
procedure based on biological activity was developed for the separation and
purification of the restriction endonucleases from H. influenzae strain Rd.
Two endonuclease R activities presumably corresponding to HindII and HindIII
(P.H. Roy and H.O. Smith, 1973; H.O. Smith and K.W. Wilcox, 1970) were
characterized by differences in their chromatographic properties, ability to
attack T7 DNA, and inactivation of the transforming activity of different
markers of H. parainfluenzae DNA.  One endonuclease R enzyme (HindII) attacked
T7 DNA and was found to inactivate the dalacin resistance marker (<0.01%
activity remaining) with only a slight effect on the streptomycin resistance
marker (83% activity remaining).  Methylase IIa treatment protected 40% of the
dalacin resistance marker of H. parainfluenzae DNA from inactivation by HindII.
The other restriction activity (HindIII) was inert towards T7 DNA and
inactivated the streptomycin resistance marker of H. parainfluenzae DNA (<0.01%
activity remaining) without any effect on the dalacin resistance marker.  The
methylation of H. parainfluenzae DNA accomplished by methylase III protected
60% of the transforming activity of the streptomycin resistance marker of H.
parainfluenzae DNA from the action of HindIII.

<>

<1>Roth, M., Helm-Kruse, S., Friedrich, T., Jeltsch, A.
<2>Functional roles of conserved amino acid residues in DNA methyltransferases investigated by site-directed mutagenesis of the EcoRV adenine-M6-methyltransferase.
<3>J. Biol. Chem.
<4>273
<5>17333-17342
<6>1998
<7>All DNA methyltransferases have similar catalytic domains containing nine blocks of conserved
amino acid residues.  We have investigated by site-directed mutagenesis the function of 17
conserved residues in the EcoRV alpha-adenine-N6-DNA methyltransferase.  The structure of this
class of MTases has been predicted recently.  The variants were characterized with respect to
their catalytic activities and their abilities to bind to DNA and the S-adenosylmethionine
cofactor.  Amino acids located in motifs X, I, and II are shown to be involved in AdoMet
binding (Lys16, Glu37, Phe39, and Asp58).  Some of the mutants defective in AdoMet binding are
also impaired in DNA binding, suggesting allosteric interactions between the AdoMet and DNA
binding site.  Asp78 (motif III), which was supposed to form a hydrogen bond to the AdoMet on
the basis of the structure predictions, turned out not to be important for AdoMet binding,
suggesting that motif III has not been identified correctly.  R128A and N130A, having
mutations in the putative DNA binding domain, are unable to bind to DNA.  Residues located in
motifs IV, V, VI, and VIII are involved in catalysis (Asp193, Tyr196, Asp211, Ser229, Trp231,
and Tyr258), some of them presumably in binding the flipped target base, because mutations at
these residues fail to significantly interfere with DNA and AdoMet binding but strongly reduce
catalysis. Our results are in substantial agreement with the structure prediction for EcoRV
alpha-adenine-N6-methyltransferase and x-ray structures of other MTases.

<>

<1>Roth, M., Jeltsch, A.
<2>Biotin-Avidin microplate assay for the quantitative analysis of enzymatic methylation of DNA by DNA methyltransferases.
<3>Biol. Chem.
<4>381
<5>269-272
<6>2000
<7>An assay is described to measure methylation of biotinylated oligonucleotide substrates by DNA
methyltransferases using [methyl-3H]-AdoMet.  After the methylation reaction the
oligonucleotides are immobilized on an avidin-coated microplate.  The incorporation of [3H]
into the DNA is quenched by addition of unlabeled AdoMet to the binding buffer.  Unreacted
AdoMet and enzyme are removed by washing.  To release the radioactivity incorporated into the
DNA, the wells are incubated with a non-specific endonuclease and the radioactivity determined
by liquid scintillation counting.  As an example, we have studied methylation of DNA by the
EcoRV DNA methyltransferase.  The reaction progress curves measured with this assay are linear
with respect to time.  Methylation rates linearly increase with enzyme concentration.  The
rates are comparable to results obtained with the same enzyme using a different assay.  The
biotin-avidin assay is inexpensive, convenient, quantitative, fast and well suited to process
many samples in parallel.  The accuracy of the assay is high, allowing to reproduce results
within +-10%. The assay is very sensitive as demonstrated by the detection of incorporation of
0.8 fmol methyl groups into the DNA. Under the experimental conditions, this corresponds to
methylation of only 0.03% of all target sites of the substrate.  Using this assay, the DNA
methylation activity of some M.EcoRV variants could be detected that was not visible by other
in vitro methylation assays.

<>

<1>Roth, M., Jeltsch, A.
<2>Changing the target base specificity of the EcoRV DNA methyltransferase by rational de novo protein-design.
<3>Nucleic Acids Res.
<4>29
<5>3137-3144
<6>2001
<7>The EcoRV DNA-(adenine-N(6))-methyltransferase (M.EcoRV) specifically modifies the first
adenine residue within GATATC sequences. During catalysis, the enzyme flips its target base
out of the DNA helix and binds it into a target base-binding pocket, which is formed in part
by Lys16 and Tyr196. A cytosine residue is accepted by wild-type M.EcoRV as a substrate at a
31-fold reduced efficiency with respect to the k(cat)/K(M) values if it is located in a CT
mismatch substrate (GCTATC/GATATC). Cytosine residues positioned in a CG base pair
(GCTATC/GATAGC) are modified at much more reduced rates, because flipping out the target base
is much more difficult in this case. We intended to change the target base specificity of
M.EcoRV from adenine-N(6) to cytosine-N(4). To this end we generated, purified and
characterized 15 variants of the enzyme, containing single, double and triple amino acid
exchanges following different design approaches. One concept was to reduce the size of the
target base-binding pocket by site-directed mutagenesis. The K16R variant showed an altered
specificity, with a 22-fold preference for cytosine as the target base in a mismatch
substrate. This corresponds to a 680-fold change in specificity, which was accompanied by only
a small loss in catalytic activity with the cytosine substrate. The K16R/Y196W variant no
longer methylated adenine residues at all and its activity towards cytosine was reduced only
17-fold. Therefore, we have changed the target base specificity of M.EcoRV from adenine to
cytosine by rational protein design. Because there are no natural paragons for the variants
described here, a change of the target base specificity of a DNA interacting enzyme was
possible by rational de novo design of its active site.

<>

<1>Rotman, E., Kouzminova, E., Plunkett, G. III, Kuzminov, A.
<2>Genome of Enterobacteriophage Lula/phi80 and Insights into Its Ability To Spread in the Laboratory Environment.
<3>J. Bacteriol.
<4>194
<5>6802-6817
<6>2012
<7>The novel temperate bacteriophage Lula, contaminating laboratory Escherichia coli
strains, turned out to be the well-known lambdoid phage phi80. Our previous
studies revealed that two characteristics of Lula/phi80 facilitate its spread in
the laboratory environment: cryptic lysogen productivity and stealthy
infectivity. To understand the genetics/genomics behind these traits, we
sequenced and annotated the Lula/phi80 genome, encountering an E. coli-toxic gene
revealed as a gap in the sequencing contig and analyzing a few genes in more
detail. Lula/phi80's genome layout copies that of lambda, yet homology with other
lambdoid phages is mostly limited to the capsid genes. Lula/phi80's DNA is
resistant to cutting with several restriction enzymes, suggesting DNA
modification, but deletion of the phage's damL gene, coding for DNA adenine
methylase, did not make DNA cuttable. The damL mutation of Lula/phi80 also did
not change the phage titer in lysogen cultures, whereas the host dam mutation did
increase it almost 100-fold. Since the high phage titer in cultures of Lula/phi80
lysogens is apparently in response to endogenous DNA damage, we deleted the only
Lula/phi80 SOS-controlled gene, dinL. We found that dinL mutant lysogens release
fewer phage in response to endogenous DNA damage but are unchanged in their
response to external DNA damage. The toxic gene of Lula/phi80, gamL, encodes an
inhibitor of the host ATP-dependent exonucleases, RecBCD and SbcCD. Its own
antidote, agt, apparently encoding a modifier protein, was found nearby.
Interestingly, Lula/phi80 lysogens are recD and sbcCD phenocopies, so GamL and
Agt are part of lysogenic conversion.

<>

<1>Rouet, P., Smih, F., Jasin, M.
<2>Introduction of double-strand breaks into the genome of mouse cells by expression of a rare-cutting endonuclease.
<3>Mol. Cell. Biol.
<4>14
<5>8096-8106
<6>1994
<7>To maintain genomic integrity, double-strand breaks (DSBs) in chromosomal DNA must be
repaired. In mammalian systems, the analysis of the repair of chromosomal DSBs has been
limited by the inability to introduce well-defined DSBs in genomic DNA. In this study, we
created specific DSBs in mouse chromosomes for the first time, using an expression system for
a rare-cutting endonuclease, I-SceI. A genetic assay has been devised to monitor the repair of
DSBs, whereby cleavage sites for I-SceI have been integrated into the mouse genome in two
tandem neomycin phosphotransferase genes. We find that cleavage of the I-SceI sites is very
efficient, with at least 12% of stably transfected cells having at least one cleavage event
and, of these, more than 70% have undergone cleavage at both I-SceI sites. Cleavage of both
sites in a fraction of clones deletes 3.8 kb of intervening chromosomal sequences. We find
that the DSBs are repaired by both homologous and nonhomologous mechanisms. Nonhomologous
repair events frequently result in small deletions after rejoining of the two DNA ends. Some
of these appear to occur by simple blunt-ended ligation, whereas several others may occur
through annealing of short regions of terminal homology. The DSBs are apparently
recombinogenic, stimulating gene targeting of a homologous fragment by more than 2 orders of
magnitude. Whereas gene-targeted clones are nearly undetectable without endonuclease
expression, they represent approximately 10% of cells transfected with the I-SceI expression
vector. Gene targeted clones are of two major types, those that occur by two-sided homologous
recombination with the homologous fragment and those that occur by one-sided homologous
recombination. Our results are expected to impact a number of areas in the study of mammalian
genome dynamics, including the analysis of the repair of DSBs and homologous recombination
and, potentially, molecular genetic analyses of mammalian genomes.

<>

<1>Rouet, P., Smith, F., Jasin, M.
<2>Expression of a site-specific endonuclease stimulates homologous recombination in mammalian cells.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>6064-6068
<6>1994
<7>Double-strand breaks introduced into DNA in vivo have been shown to enhance homologous
recombination in a variety of chromosomal and extrachromosomal loci in Saccharomyces
cerevisiae. To introduce double-strand breaks in DNA at defined locations in mammalian cells,
we have constructed a mammalian expression vector for a modified form of I-SceI, a yeast
mitochondrial intron-encoded endonuclease with an 18-bp recognition sequence. Expression of
the modified I-SceI endonuclease in COS1 cells results in cleavage of model recombination
substrates and enhanced extrachromosomal recombination, as assayed by chloramphenicol
acetyltransferase activity and Southern blot analysis. Constitutive expression of the
endonuclease in mouse 3T3 cells is not lethal, possibly due to either the lack of I-SceI sites
in the genome or sufficient repair of them. Expression of an endonuclease with such a long
recognition sequence will provide a powerful approach to studying a number of molecular
processes in mammalian cells, including homologous recombination.

<>

<1>Rouleau, J., MacLeod, A.R., Szyf, M.
<2>Regulation of the DNA methyltransferase by the Ras-AP-1 signaling pathway.
<3>J. Biol. Chem.
<4>270
<5>1595-1601
<6>1995
<7>Using deletion analysis and site-specific mutagenesis to map the 5' regulatory region of the
DNA methyltransferase (MeTase) gene, we show that a 106-bp sequence (at -1744 to -1650)
bearing three AP-1 sites is responsible for induction of DNA MeTase promoter activity. Using
transient cotransfection chloramphenicol acetyltransferase assays in P19 cells, we show that
the DNA MeTase promoter is induced by c-Jun or Ha-Ras but not by a dominant negative mutant of
Jun, 99. The activation of the DNA MeTase promoter by Jun is inhibited in a ligand dependent
manner by the glucocorticoid receptor. Stable expression of Ha-Ras in P19 cells results in
induction of transcription of the DNA MeTase mRNA as determined by nuclear run-on assays and
the steady state levels of DNA MeTase mRNA as determined by an RNase protection assay. These
experiments establish a potential molecular link between nodal cellular signaling pathways and
the control of expression of the DNA MeTase gene. This provides us with a possible molecular
explanation for the hyperactivation of DNA MeTase in many cancer cells and suggests that DNA
MeTase is one possible downstream effector of Ras.

<>

<1>Rouleau, J., Szyf, M.
<2>Regulation of the mouse DNA methyltransferase by signal transduction pathways.
<3>Mol. Biol. Cell
<4>3
<5>A25
<6>1992
<7>A hallmark of DNA methylation in vertebrates is the fact that only a fraction of the CpG
sequences is methylated (60-80%) and that nonmethylated cytosines are distributed in a
nonrandom fashion, generating a pattern of methylation that is gene and tissue specific. DNA
methylation is catalyzed by the DNA methyltransferase enzyme (DNA MeTase). We have previously
hypothesized that regulated changes in the level of DNA MeTase gene expression might be an
important mechanism through which DNA methylation patterns are generated(Szyf et al., J. Biol.
Chem. 266, 10027-10030, 1991). If this is true the DNA MeTase gene should be responsive to
cellular signal transduction pathways that are involved in cellular differentiation. Sequence
analysis of the promoter region revealed several potential binding sites for transcript factor
complexes (AP-1, GRE and E-boxes) that might be involved in the regulation of the DNA MeTase
gene expression by different signal transduction pathways (Rouleau et al., J. Biol. Chem.,
267, 7368-7377. 1992). To test this hypothesis we cotransfected P19 cells with a chimeric
construct containing 2.3 kb sequences from the 5' region of the DNA MeTase fused to
CAT(pMetCAT+) with fos and jun. DNA MeTase gene promoter activity was induced 60-fold by fos
and jun but not by a mutant of jun lacking the DNA binding domain. This induction was
inhibited by deletion of the 7 AP-1 sites in the 5' region of the DNA MeTase. Gel retardation
assays demonstated the formation of an AP-1 complex with an AP-1 binding site in the promoter
region. Induction of DNA MeTase transcripton by fos and jun was inhibited by the human
glucocorticoid receptor while expression of the receptor per se had no effect on DNA MeTase
activity. The DNA MeTase is responsive also to the myogenesis regulator MyoD which expression
results in a 20 fold induction of the DNA MeTase gene. This work suggests for the first time a
molecular link between extracellular signals and possible changes in the covalent structure of
the genome.

<>

<1>Rouleau, J., Tanigawa, G., Szyf, M.
<2>The mouse DNA methyltransferase 5'-region .
<3>J. Biol. Chem.
<4>267
<5>7368-7377
<6>1992
<7>We have cloned and characterized 5'-flanking sequences of the DNA methyltransferase (MeTase)
gene. DNA MeTase gene transcription is initiated at a few discrete sites: 343 and 90 base
pairs upstream of the translation initiation site as determined by RNase protection and primer
extension assays. The promoter sequences that regulate expression of DNA MeTase, as defined by
chloramphenicol acetyltransferase assays, reside between position - 171 and the transcription
start site. The promoter of DNA MeTase does not contain TATAA or CAAT boxes and is unusual
because it does not contain the CG-rich elements characteristic of TATAA-less housekeeping
genes. The 5'-flanking region of DNA MeTase contains AP-1, AP-2 and glucocorticoid response
elements, suggesting possible regulation by cellular transduction pathways. The base
composition of the DNA MeTase promoter is markedly different from that of other housekeeping
genes. Whereas most housekeeping genes are characterized by CG-rich areas in their
5'-flanking regions, the TG dinucleotide is over-represented in DNA MeTase 5'-flanking
sequences, including a perfect tandem repeat of T/G between positions -685 and -650. DNA
methylation patterns play an important role in the developmental regulation of gene expression
in vertebrates. DNA MeTase activity is probably regulated to maintain this pattern of
methylation. We suggest that the DNA MeTase promoter represents a new class of housekeeping
gene promoters that was designed to ensure high fidelity regulation of gene expression.

<>

<1>Rouli, L., Robert, C., Raoult, D., Yagupsky, P.
<2>Kingella kingae KK247, an Atypical Pulsed-Field Gel Electrophoresis Clone A Strain.
<3>Genome Announcements
<4>2
<5>e01228-14
<6>2014
<7>Kingella kingae strain KK247 was isolated from an adult Israeli patient with endocarditis. It
belongs to pulsed-field gel electrophoresis clone A, has a
2,113,021-bp genome, a 15,507-bp plasmid that carries genes encoding
beta-lactamases, and possesses 45 transposases, compared to the 5 detected in
other K. kingae strains.

<>

<1>Rouli, L., Rolain, J.M., El Filali, A., Robert, C., Raoult, D.
<2>Genome Sequence of Coxiella burnetii 109, a Doxycycline-Resistant Clinical Isolate.
<3>J. Bacteriol.
<4>194
<5>6939
<6>2012
<7>Coxiella burnetii 109, with a 2.03-Mb genome, is a doxycycline-resistant human isolate that
was isolated from the cardiac valve of a German male patient with Q
fever endocarditis who died during the course of the treatment due to the
bacterium's resistance to doxycycline. This new genome can be useful for future
comparative genomic or Q fever studies.

<>

<1>Roulland-Dussoix, D., Boyer, H.W.
<2>The Escherichia coli B restriction endonuclease.
<3>Biochim. Biophys. Acta
<4>195
<5>219-229
<6>1969
<7>1.  The restriction endonuclease of Escherichia coli B has been purified
1000-fold from crude extracts. 2.  It has been found to be similar to the K12
and PI restriction endonucleases, i.e., it requires ATP,
S-adenosyl-L-methionine and Mg2+ and unmodified DNA for enzymatic activity and
has an estimated large molecular weight (approx. 300,000 daltons). 3.  It
introduces a limited number of double strand scissions in unmodified lambda vir
DNA, Escherichia coli chromosomal DNA and probably one double strand scission
per fd replicative form DNA.  The double strand scission in the unmodified fd
replicative form DNA occurs by a two-step mechanism. 4.  Replicative form DNA
generated from an fd mutant which is only restricted by Escherichia coli B by a
factor of 1.10-2 (versus 1.10-4 for wild type fd) is also a substrate for the B
restriction endonuclease.  Cosedimentation of the endonuclease-treated wild
type and mutant replicative form DNA results in qualitatively identical
patterns of DNA distribution.

<>

<1>Roulland-Dussoix, D., Boyer, H.W.
<2>B Restriction endonuclease of Escherichia coli.
<3>Bacteriol. Proc.
<4>69
<5>46
<6>1969
<7>We have purified the B restriction endonuclease by a factor of 1,500 from a
crude extract of E. coli.  The enzyme preparation is free of detectable
exonuclease and nonspecific endonuclease activities under conditions optimum
for the restriction endonuclease activity.  The B restriction endonuclease
activity has an absolute requirement for SAM, ATP, Mg++ and unmodified duplex
DNA (i.e. DNA originating in a strain without B modification).  The enzyme
produces a limited number of double strand scissions in unmodified lambda++,
lambda vir,  fd RF, colE1 and E. coli DNA.  The double strand scission is made
by two proximal single strand scissions.  The molecular weight of the enzyme
has been estimated at about 140,000 daltlons.  With the exception of substrate
specificity, the B restriction endonuclease appears to be similar to the K
restriction endonuclease.  We have found that the B restriction endonuclease
produces a linear molecule from the fd RF with an estimated molecular weight of
2.8 x 10/6 daltons and sediments a little slower than the nicked RF molecules
(ratio =  1.14).  We conclude that there is one double strand scission per RF
molecule and have used this as a substrate to study the kinetics of the
reaction.  RF, prepared from a mutant fd which is restricted in vivo 100-fold
less than WT fd, is attacked at a rate less than one-half that of the WT RF.

<>

<1>Roulland-Dussoix, D., Yoshimori, R., Greene, P., Betlach, M., Goodman, H.M., Boyer, H.W.
<2>R Factor-Controlled Restriction and Modification of Deoxyribonucleic Acid.
<3>Microbiology-1974, American Society for Microbiology, Schlesinger, D., Washington, D.C.
<4>0
<5>187-198
<6>1975
<7>The restriction and modification of deoxyribonucleic acid (DNA) appear to be
quite prevalent in bacteria although by no means ubiquitous.  In Escherichia
coli strains alone, there appear to be over six different restriction and
modification systems in terms of enzymatic specificity.  In E. coli strains,
the enzymes responsible for the restriction and modification of DNA are
genetically controlled by chromosomal, plasmid, or viral genes.  At least in
some cases, it is clear that the restriction endonuclease and modification
methylase of a given "host specificity" interact with the same substrate, a
specific sequence of nucleotide base pairs.  This sequence defines the host
specificity of a bacterial cell which possesses a set of restriction and
modification enzymes. Naturally occurring bacterial strains or strains
constructed in the laboratory can be demonstrated to have several sets of
restriction and modification enzymes.

<>

<1>Rountree, M.R., Bachman, K.E., Baylin, S.B.
<2>DNMT1 binds HDAC2 and a new co-repressor, DMAP1, to form a complex at replication foci.
<3>Nat. Genet.
<4>25
<5>269-277
<6>2000
<7>DNA methylation can contribute to transcriptional silencing through several transcriptionally
repressive complexes, which include methyl-CpG binding domain proteins (MBDs) and histone
deacetylases (HDACs).  We show here that the chief enzyme that maintains mammalian DNA
methylation, DNMT1, can also establish a repressive transcription complex. The non-catalytic
amino terminus of DNMT1 binds to HDAC2 and a new protein, DMAP1 (for DNMT1 associated
protein), and can mediate transcriptional repression. DMAP1 has intrinsic transcription
repressive activity, and binds to the transcriptional co-repressor TSG101. DMAP1 is targeted
to replication foci through interaction with the far N terminus of DNMT1 throughout S phase,
whereas HDAC2 joins DNMT1 and DMAP1 only during late S phase, providing a platform for how
histones may become deacetylated in heterochromatin following replication. Thus, DNMT1 not
only maintains DNA methylation, but also may directly target, in a heritable manner,
transcriptionally repressive chromatin to the genome during DNA replication.

<>

<1>Rountree, P.M.
<2>Variations in a related series of staphylococcal bacteriophages.
<3>J. Gen. Microbiol.
<4>15
<5>266-279
<6>1956
<7>Stocks of staphylococcal phage 47C (serological group A) contained some group B
phage.  This was found to have originated in a lysogenic staphylococcus
previously used to propagate phage 47C and which had subsequently lost its
lysogenicity.  The lysogenic phage was thus perpetuated in the phage stocks as
a lytic phage.  The characters of the two phages are described.  When they
lysogenized five different strains of staphylococci changes in typing pattern
were produced.  There was evidence which indicated that, in the prophage state,
the two phages occupy different sites and that there is no cross-immunity
between them.  The propagation of the phages in different hosts resulted in
changes in their host range.  A virulent mutant of the B prophage was induced
by the application of a variety of phages to the strain carrying it.

<>

<1>Roustan-Espinosa, I., Guerrero, D., Flores, E., Huete-Perez, J.
<2>Restriction enzymes in native bacteria of Nicaragua.
<3>Revista Univ. Centroamer., Revista de la Universidad Centroamericana, , 
<4>52
<5>10-18
<6>2000
<7>Advances in genetic engineering and molecular biology have led to the utilization of bacteria
in the biotechnology industry.  In this study, restriction enzymes present in bacteria
collected from aqueous medium in Nicaragua have been identified and classified.  Restriction
activity was found in 25% of the total of bacteria analyzed.  The process of purification of
bacterial protein extracts with Sau96I and PvuII activities is discussed.  This work is a
result of an effort to implement modern biotechnological methods of research in Nicaragua.

<>

<1>Roux, V., El Karkouri, K., Lagier, J.C., Robert, C., Raoult, D.
<2>Non-contiguous finished genome sequence and description of Kurthia massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>7
<5>221-232
<6>2012
<7>Kurthia massiliensis strain JC30(T) sp. nov. is the type strain of K. massiliensis sp. nov., a
new species within the genus Kurthia. This strain, whose
genome is described here, was isolated from the fecal flora of a healthy patient.
K. massiliensis is a Gram-positive aerobic rod. Here we describe the features of
this organism, together with the complete genome sequence and annotation. The
3,199,090 bp long genome contains 3,240 protein-coding genes and 86 RNA genes,
including between 3 and 4 rRNA genes.

<>

<1>Roux, V., Lagier, J.C., Gorlas, A., Robert, C., Raoult, D.
<2>Non-contiguous finished genome sequence and description of Kurthia senegalensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>1321-1332
<6>2014
<7>Kurthia senegalensis strain JC8E(T) sp. nov. is the type strain of K. senegalensis sp. nov., a
new species within the genus Kurthia. This strain, whose
genome is described here, was isolated from the fecal flora of a healthy patient.
K. senegalensis is an aerobic rod. Here we describe the features of this
organism, together with the complete genome sequence and annotation. The
2,975,103 bp long genome contains 2,889 protein-coding genes and 83 RNA genes,
including between 4 and 6 rRNA genes.

<>

<1>Roux, V., Million, M., Robert, C., Magne, A., Raoult, D.
<2>Non-contiguous finished genome sequence and description of Oceanobacillus massiliensis sp. nov.
<3>Standards in Genomic Sciences
<4>9
<5>370-384
<6>2013
<7>Oceanobacillus massiliensis strain N'Diop(T) sp. nov. is the type strain of O. massiliensis
sp. nov., a new species within the genus Oceanobacillus. This
strain, whose genome is described here, was isolated from the fecal flora of a
healthy patient. O. massiliensis is an aerobic rod. Here we describe the features
of this organism, together with the complete genome sequence and annotation. The
3,532,675 bp long genome contains 3,519 protein-coding genes and 72 RNA genes,
including between 6 and 8 rRNA operons.

<>

<1>Roux, V., Robert, C., Gimenez, G., Gharbi, R., Raoult, D.
<2>Draft Genome Sequence of Actinomyces massiliensis Strain 4401292T.
<3>J. Bacteriol.
<4>194
<5>5121
<6>2012
<7>A draft genome sequence of Actinomyces massiliensis, an anaerobic bacterium isolated from a
patient's blood culture, is described here. CRISPR-associated
proteins, insertion sequences, and toxin-antitoxin loci were found on the genome.

<>

<1>Roux, V., Robert, C., Gimenez, G., Raoult, D.
<2>Draft Genome Sequence of Brevibacterium massiliense Strain 541308T.
<3>J. Bacteriol.
<4>194
<5>5151-5152
<6>2012
<7>A draft genome sequence of Brevibacterium massiliense, an aerobic bacterium isolated from a
human ankle discharge, is described here. CRISPR-associated
proteins were found to be encoded in the genome, and analysis of transport
proteins was performed.

<>

<1>Roux, V., Robert, C., Gimenez, G., Raoult, D.
<2>Draft Genome Sequence of Staphylococcus massiliensis Strain 5402776T.
<3>J. Bacteriol.
<4>194
<5>6984-6985
<6>2012
<7>A draft genome sequence of Staphylococcus massiliensis, Gram-positive cocci isolated from a
human brain abscess sample, is described here. One clustered
regularly interspaced short palindromic repeat, three transposases, six putative
transposases, and one potential provirus were characterized.

<>

<1>Roux, V., Robert, C., Raoult, D.
<2>Non-contiguous finished genome sequence of Phocaeicola abscessus type strain 7401987(T.).
<3>Standards in Genomic Sciences
<4>9
<5>351-358
<6>2013
<7>Phocaeicola abscessus strain 7401987(T) is the sole member of the genus Phocaeicola. This
bacterium is Gram-negative, non-spore-forming, coccoid to
rod-shaped and motile by lophotrichous flagella. It was isolated from a human
brain abscess sample. In this work, we describe a set of features of this
organism, together with the complete genome sequence and annotation. The
2,530,616 bp long genome contains 2,090 protein-coding genes and 54 RNA genes,
including 4 rRNA operons.

<>

<1>Roux, V., Robert, C., Raoult, D.
<2>Non-contiguous finished genome sequence of Prevotella timonensis type strain 4401737(T.).
<3>Standards in Genomic Sciences
<4>9
<5>1346-1353
<6>2014
<7>Prevotella timonensis strain 4401737(T) is a member of the genus Prevotella, which contains
anaerobic Gram-negative bacteria. It was isolated from a human
breast abscess. In this work, we describe a set of features of this organism,
together with the complete genome sequence and annotation. The 3,169,464 bp long
genome contains 2,746 protein-coding genes and 56 RNA genes, including 3 or 4
rRNA operons.

<>

<1>Roux, V., Robert, C., Raoult, D.
<2>Non-contiguous finished genome sequence of Corynebacterium timonense type strain  5401744(T.).
<3>Standards in Genomic Sciences
<4>9
<5>948-955
<6>2014
<7>Corynebacterium timonense strain 5401744(T) is a member of the genus Corynebacterium which
contains Gram-positive bacteria with a high G+C content. It
was isolated from the blood of a patient with endocarditis. In this work, we
describe a set of features of this organism, together with the complete genome
sequence and annotation. The 2,553,575 bp long genome contains 2,401
protein-coding genes and 55 RNA genes, including between 5 and 6 rRNA operons.

<>

<1>Rowland, G.C., Lim, P.P., Glass, R.E.
<2>'Stop-codon-specific' restriction endonucleases: their use in mapping and gene manipulation.
<3>Gene
<4>116
<5>21-26
<6>1992
<7>Certain restriction endonucleases recognize target sequences that contain the stop triplet TAG
and are commonly either 4 or 6 bp in length. Interestingly, these restriction targets do not
occur at the frequency expected on the basis of base composition and size. For example, the
tetranucleotide MaeI recognition sequence (CTAG) occurs considerably less commonly (5-8 fold)
in the genome of Escherichia coli (and many other eubacteria) than expected from
mononucleotide frequencies. This surprising rarity is particularly evident in protein-encoding
genes and is largely dictated by codon usage. Thus, amber (TAG) nonsense mutations frequently
give rise to novel MaeI (CTAG) sites which are unique within a translated region. Such
amber/MaeI sites, whether arising spontaneously or created in vitro by site-directed
mutagensis, act as a useful physical marker for the presence of the nonsense mutation and are
a convenient startpoint for a range of diverse procedures. These features provide a useful
supplement to protein engineering methods which use nonsense suppression to mediate amino acid
replacements.

<>

<1>Roy, A.S., Baruah, R., Gogoi, D., Borah, M., Singh, A.K., Deka, B.H.P.
<2>Draft Genome Sequence of Pseudomonas aeruginosa Strain N002, Isolated from Crude  Oil-Contaminated Soil from Geleky, Assam, India.
<3>Genome Announcements
<4>1
<5>e00104-12
<6>2013
<7>Here, we report the draft genome sequence of crude oil-degrading Pseudomonas aeruginosa strain
N002, isolated from a crude oil-polluted soil sample from
Geleky, Assam, India. Multiple genes potentially involved in crude oil
degradation were identified.

<>

<1>Roy, K.B., Vrushank, D.
<2>Bam HI cleaves the self complementary dodecamer d-CGCGGAGCCGCG, before the two G's and possibly binds in the DNA major groove.
<3>Biochem. Mol. Biol. Int.
<4>36
<5>759-770
<6>1995
<7>Oligodeoxynucleotides with GT or GA mispairs within the Bam HI recognition sequence (GGATCC),
have been prepared. Binding and cleavage of the native
vis a vis the mismatch substrates by Bam HI are analysed. UV melting
curves and CD spectra of the oligomers suggest a double stranded B-DNA
conformation. The enzyme Bam HI binds with varying affinities to the
oligomers except the one with the GT wobble base pair. Bam HI cleaves the
cognate sequence, GGATCC, between the two Guanines but cleaves GGAGCC
before the guanines. The unusual cleavage is due to a local distortion in
the DNA structure. Kinetic analysis of the cleavage reactions using the
35S labeled hexadecamers, d-ATGGCGGATCCGCCAT and d-ATGGCGGAGCCGCCAT, as
substrates gives Km values 11.08 nM and 1.16 nM with corresponding Kcat of
11.04 and 0.62 min-1 respectively. The results are consistent with the
binding of Bam HI in the major groove.

<>

<1>Roy, K.B., Vrushank, D., Jayaram, B.
<2>Use of isotope-dilution phenomenon to advantage in the determination of kinetic constants Km and Kcat for BamHI restriction endonuclease: an empirical and interative approach.
<3>Anal. Biochem.
<4>220
<5>160-164
<6>1994
<7>An assay using a very small amount of 35S-labeled deoxyoligonucleotide as a substrate for the
determination of Km and Kcat for the restriction enzyme BamHI is described. Two synthetic
deoxyoligonucleotides, ATGGCGGATCCGC and ATGGCGGAGCCGC, containing the cognate and a mismatch
BamHI sequence, respectively, were labeled by an end-filling reaction using the Klenow
fragment of DNA polymerase and [35S]dATP to generate the labeled self-complementary
substrates. The dependence of BamHI hydrolysis on substrate concentration was investigated
using mixtures of a fixed amount of radiolabeled substrate and varying amounts of cold-labeled
substrate over a wide range. The apparent competitive inhibition observed due to the
phenomenon of carrier dilution was analytically corrected by an empirical as well as an
iterative approach to give Km values comparable to those reported in the literature. We have
found that the values obtained using the empirical formula are very close to the precise
values obtained through iteration. Our procedure has used isotopic dilution to advantage to
make the assay less expensive and can be applied effectively to any enzyme-substrate reaction
in which the substrate and the product have radioactive labels. The method would be especially
useful for a rapid analysis and comparison of kinetic constants of various mutant enzymes or
substrates.

<>

<1>Roy, P.H., Smith, H.O.
<2>DNA methylases of Hemophilus influenzae Rd.  II. Partial recognition site base sequences.
<3>J. Mol. Biol.
<4>81
<5>445-459
<6>1973
<7>A small percentage of the adenine bases in Hemophilus influenzae strain Rd DNA
are methylated in the 6-amino position.  The methyl groups are introduced
specifically by at least four different DNA methylases (I, II, III and IV).  A
method is described for determining the 3' and 5' nearest-neighbor bases to
methylated adenine so as to reveal the specificity of each methylase.
Tritium-labeled methyl groups are introduced into the DNA.  The DNA is then
digested to dinucleotides using the Bacillus subtilis phage SP3 DNase, followed
by removal of the terminal 5'-phosphoryl group with phosphatase to produce
dinucleoside monophosphates.  These are analyzed by Aminex A25 (Bio-Rad)
chromatography.  Dinucleoside monophosphate species containing the 3' neighbor
or the 5' neighbor are resolved so that a trinucleotide is determined that
contains the centrally placed methylated adenine.  H. influenzae Rd DNA
contains seven dinucleoside monophosphate species, about 80% representing GpmA
and mApT in approximately equal amount.  DNA methylases I, II, III and IV
introduce methyl groups into sequences containing the trinucleotides CpmApC,
PupmApC, NpmApA and GpmApT, respectively.  The sequence methylated by DNA
methylase II is consistent with the recognition site determined by Kelly &
Smith (1970) for the H. influenzae restriction enzyme, endonuclease R.

<>

<1>Roy, P.H., Smith, H.O.
<2>DNA methylases of Hemophilus influenzae Rd I.  Purification and properties.
<3>J. Mol. Biol.
<4>81
<5>427-444
<6>1973
<7>Hemophilus influenzae strain Rd DNA contains small amounts of 5-methylcytosine
(0.012%) and significantly greater amounts of N-6-methyladenine (0.34%).  Four
DNA adenine methylases have been identified and purified from crude extracts of
H. influenzae Rd by means of phosphocellulose chromatography.  Each of the four
enzymes requires S-adenosyl-L-methionine as a methyl group donor and each
differs in its ability to methylate various DNAs in vitro.  DNA methylase I is
related to the genetically described modification-restriction system in H.
influenzae Rd, and is presumably the modification enzyme for that system.  DNA
methylase II introduces approximately 130 methyl groups into a phage T7 DNA
molecule and protects T7 DNA from the H. influenzae Rd restriction enzyme,
endonuclease R, described by Smith & Wilcox (1970).  These findings indicate
that DNA methylase II is the modification enzyme corresponding to endonuclease
R.  A third modification-restriction system, which does not affect T7 DNA, has
been detected in H. influenzae Rd.  DNA methylase III is apparently the
modification enzyme for this system.  The biological function of DNA methylase
IV remains unknown.

<>

<1>Roy, P.H., Tetu, S.G., Larouche, A., Elbourne, L., Tremblay, S., Ren, Q., Dodson, R., Harkins, D., Shay, R., Watkins, K., Mahamoud, Y., Paulsen, I.T.
<2>Complete genome sequence of the multiresistant taxonomic outlier Pseudomonas aeruginosa PA7.
<3>PLoS ONE
<4>5
<5>e8842
<6>2010
<7>Pseudomonas aeruginosa PA7 is a non-respiratory human isolate from Argentina that is
multiresistant to antibiotics. We first sequenced gyrA,
gyrB, parC, parE, ampC, ampR, and several housekeeping genes and found
that PA7 is a taxonomic outlier. We report here the complete sequence of
the 6,588,339 bp genome, which has only about 95% overall identity to
other strains. PA7 has multiple novel genomic islands and a total of 51
occupied regions of genomic plasticity. These islands include antibiotic
resistance genes, parts of transposons, prophages, and a pKLC102-related
island. Several PA7 genes not present in PAO1 or PA14 are putative
orthologues of other Pseudomonas spp. and Ralstonia spp. genes. PA7
appears to be closely related to the known taxonomic outlier DSM1128
(ATCC9027). PA7 lacks several virulence factors, notably the entire TTSS
region corresponding to PA1690-PA1725 of PAO1. It has neither exoS nor
exoU and lacks toxA, exoT, and exoY. PA7 is serotype O12 and pyoverdin
type II. Preliminary proteomic studies indicate numerous differences with
PAO1, some of which are probably a consequence of a frameshift mutation in
the mvfR quorum sensing regulatory gene.

<>

<1>Roycroft, E., Mac, A.M., O'Toole, R.F., Fitzgibbon, M., Rogers, T.R.
<2>Draft Genome Sequence of the First Isolate of Extensively Drug-Resistant Mycobacterium tuberculosis in Ireland.
<3>Genome Announcements
<4>2
<5>e01002-14
<6>2014
<7>Extensive drug resistance is an emerging threat to the control of tuberculosis (TB) worldwide,
even in countries with low TB incidence. We report the draft
whole-genome sequence of the first reported extensively drug-resistant TB
(XDR-TB) strain isolated in Ireland (a low-incidence setting) and describe a
number of single-nucleotide variations that correlate with its XDR phenotype.

<>

<1>Rozanov, A.S., Bryanskaya, A.V., Kotenko, A.V., Malup, T.K., Peltek, S.E.
<2>Draft Genome Sequence of Anoxybacillus flavithermus Strain 25, Isolated from the  Garga Hot Spring in the Barguzin Valley, Baikal Region, Russian Federation.
<3>Genome Announcements
<4>2
<5>e01258-14
<6>2014
<7>Anoxybacillus flavithermus strain 25 was isolated from a sediment sample from the Garga hot
spring in the Barguzin Valley, Baikal Region, Russian Federation (54
degrees 19'3.72'N, 110 degrees 59'38.4'E). The sequenced and annotated genome
is 2,838,680 bp and encodes 3,009 genes.

<>

<1>Rozanov, A.S., Logacheva, M.D., Peltek, S.E.
<2>Draft Genome Sequences of Geobacillus stearothermophilus Strains 22 and 53, Isolated from the Garga Hot Spring in the Barguzin River Valley of the Russian  Federation.
<3>Genome Announcements
<4>2
<5>e01205-14
<6>2014
<7>Geobacillus stearothermophilus strains 22 and 53 were isolated from sediment samples isolated
from the Garga hot spring (72 degrees C) located in the valley
of the river Barguzin (the Baikal region, Russian Federation) (54 degrees
19'3.72'N, 110 degrees 59'38.4'E).

<>

<1>Ruan, H., Lunnen, K.D., Pelletier, J.J., Xu, S.-Y.
<2>Overexpression of BsoBI restriction endonuclease in E. coli, purification of the recombinant BsoBI, and identification of catalytic residues of BsoBI by random mutagenesis.
<3>Gene
<4>188
<5>35-39
<6>1997
<7>BsoBI is a type II restriction enzyme found in Bacillus stearothermophilus JN209 that
recognizes the symmetric sequence 5'-CYCGRG-3' (Y=C or T; R=A or G) and cleaves between the
first and second base to generate a four-base 5' extension.  The cloning and sequencing of
BsoBI restriction-modification system has been described by Ruan et al.  Here we report the
overexpression of the BsoBI restriction endonuclease gene in E. coli by insertion of the
endonuclease gene into an expression vector pRRS.  The recombinant BsoBI was purified to
homogeneity and its N-terminus sequence was determined.  It has the same N-terminal aa
sequence as the native enzyme.  The constitutive expression of BsoBI from pRRS is lethal to E.
coli in the absence of the cognate methylase.  The BsoBIR gene was mutagenized with either
hydroxylamine or by error-prone polymerase chain reaction in vitro and transferred into E.
coli via plasmid vectors in the absence of the cognate methylase.  Surviving transformants
were selected that carry BsoBI variants which lost endonuclease activity.  DNA sequencing of
the mutant alleles revealed that G123, D124, D212, D246, E252 and H253 are important residues
for enzymatic activity.  An electrophoretic mobility shift assay was used to identify
binding-proficient and cleavage-deficient variants.  Seven variants I95M&D124Y, G123R, D212N,
K207R&D212V, D246N, D246G and E252K can still bind DNA despite the loss of cleavage activity.
Thus, residues D124, D212, D246 and E252 may be located near or within the catalytic center,
and are likely involved in metal ion binding.

<>

<1>Ruan, H., Lunnen, K.D., Scott, M.E., Moran, L.S., Slatko, B.E., Pelletier, J.J., Hess, E.J., Benner, J., Wilson, G.G., Xu, S.-Y.
<2>Cloning and sequence comparison of AvaI and BsoBI restriction-modification systems.
<3>Mol. Gen. Genet.
<4>252
<5>695-699
<6>1996
<7>AvaI and BsoBI restriction endonucleases are isoschizomers which recognize the symmetric
sequence 5'CYCGRG3' and cleave between the first C and second Y to generate a four-base 5'
extension.  The AvaI restriction endonuclease gene (avaIR) and methylase gene (avaIM) were
cloned into Escherichia coli by the methylase selection method.  The BsoBI restriction
endonuclease gene (bsoBIM) were cloned by the "endo-blue" method (SOS induction assay), and
the remainder of bsoBIM was cloned by inverse PCR.  The nucleotide sequences of the two
restriction-modification (RM) systems were determined.  Comparisons of the predicted amino
acid sequences indicated that AvaI and BsoBI endonucleases share 55% identity, whereas the two
methylases share 41% identity.  Although the two systems show similarity in protein sequence,
their gene organization differs.  The avaIM gene precedes avaIR in the AvaI RM system, while
the bsoBIR gene is located upstream of bsoBIM in the BsoBI RM system.  Both AvaI and BsoBI
methylases contain  motifs conserved among the N4 cytosine methylases.

<>

<1>Ruan, L., Xu, X.
<2>Sequence analysis and characterizations of two novel plasmids isolated from Thermus sp. 4C.
<3>Plasmid
<4>58
<5>84-87
<6>2007
<7>Two novel plasmids, named pS4C and pL4C, were isolated from the thermophilic bacterium Thermus
sp. 4C. The pS4C with a length of 5015bp
and 58.25% of G+C content, contains 9 putative open reading frames (ORFs).
The larger plasmid, pL4C, consisting of 21,248bp, has a G+C content of
68.60% and 34 putative ORFs. Both plasmids encode their own replication
protein. The ORF 22 of pL4C and the ORF 4 of pS4C encode proteins with
high sequence similarities to integrase (97%) and transposase (97%),
respectively, which are both involved in DNA rearrangement and exchange.
Furthermore, sequence analysis of pL4C also showed that several
plasmid-encoded genes may be involved in DNA modification and repair, such
as DNA G:T-mismatch repair endonuclease and micrococcal nuclease-like
protein. These proteins may be involved in raising the repair efficiency
and other minor editing needs. Interestingly, the elimination of plasmids
significantly lowered the growth temperature of Thermus sp. 4C. Few
reports dealing with the DNA repair enzymes on the plasmid from Thermus
strains were published so far.

<>

<1>Ruan, Z., Wang, Y., Song, J., Zhai, Y., Zhang, C., Chen, C., Li, Y., Zhao, B., Zhao, B.
<2>Draft Genome Sequence of Kurthia huakuii LAM0618T, an Organic-Pollutant-Degrading Strain Isolated from Biogas Slurry.
<3>Genome Announcements
<4>2
<5>e01158-13
<6>2014
<7>Kurthia huakuii LAM0618(T) is a facultative anaerobic pollutant-degrading bacterium isolated
from biogas slurry. An analysis of the draft genome sequence
of LAM0618(T) reveals a genome size of 3,585,165 bp, with a mean G+C content of
39.1%. The genome contains 3,560 coding sequences and 112 tRNA and 33 rRNA genes.

<>

<1>Ruben, G., Spielman, P., Tu, C.-P.D., Jay, E., Siegel, B., Wu, R.
<2>Relaxed circular SV40 DNA as cleavage intermediate of two restriction endonucleases.
<3>Nucleic Acids Res.
<4>4
<5>1803-1813
<6>1977
<7>We have determined the mode of cleavage of superhelical SV40 DNA (Form I) by
restriction endonucleases EcoRI and HpaII at 37C.  By analysis with agarose gel
electrophoresis and direct examination with dark field electron microscopy, we
found that a large amount of the single-nicked circular DNA (FormII) was
produced before the linear SV40 DNA (Form III) appeared.  Thus, both
restriction enzymes cleave only one strand of the superhelical DNA first.  The
second cleavage on the complementary strand occurred after a lag period.  The
first order rate constant for the second cleavage by EcoRI endonuclease was
determined and a kinetic reaction scheme for both enzymes is proposed.

<>

<1>Rubenfield, M.J., Nolling, J., Deloughery, C., Bush, D.
<2>Nucleic acid and amino acid sequences relating to pseudomonas aeruginosa for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6551795 A
<5>
<6>2003
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Pseudomonas aeruginosa that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Rubin, R.A., Modrich, P.
<2>Substrate dependence of the mechanism of EcoRI endonuclease.
<3>Nucleic Acids Res.
<4>5
<5>2991-2997
<6>1978
<7>The mechanism of EcoRI endonuclease is substrate dependent.  At 37C,
dissociation of the enzyme-Form II DNA intermediates of ColE1 ENA and
bacteriophage G4 RFI DNA is negligible.  Therefore, both DNA strands within the
EcoRI sequence are cleaved during a single binding event.  However, double
strand cleavage of SV40 DNA occurs without dissociation of the enzyme in only
75% of the catalytic events.  This mechanistic difference presumably reflects
sequence differences about the EcoRI sites of these DNA's.

<>

<1>Rubin, R.A., Modrich, P.
<2>EcoRI methylase physical and catalytic properties of the homogeneous enzyme.
<3>J. Biol. Chem.
<4>252
<5>7265-7272
<6>1977
<7>Escherichia coli RI methylase has been isolated by a procedure which is suitable for large
scale use and which yields enzyme with a specific activity 10-fold higher than previous
methods.  The purified methylase is homogeneous as judged by polyacrylamide gel
electrophoresis, isoelectric focusing, and analytical sedimentation.  The methylase is a basic
protein composed of a single polypeptide chain of molecular weight 39,000, the stable form in
solution being a 3.0 S monomer.  No aggregation has been observed at concentrations up to 0.3
mg/ml in the temperature range of 4-30C, and the presence of S-adenosyl-L-methionine is
without effect.  Catalytic studies have demonstrated that the enzyme functions as a monomer.
Initial rates of methyl transfer are first order in methylase concentration, and the enzyme
obeys Michaelis-Menten kinetics with respect to both substrates.  At 37C, the Km for the EcoRI
site of ColE1 DNA is 1.3 nM, that for S-adenosyl-L-methionine is 0.26 microm, and the turnover
number is three methyl transfers per min.  The mechanism of methyl transfer to unmodified DNA
is also consistent with the functional form of the enzyme being a monomer.  The enzyme
transfers methyl groups to the EcoRI sequence one at a time and dissociates from the DNA prior
to any subsequent catalytic events.  Furthermore, the kinetic parameters for addition of a
second methyl group to a site which is already methylated on one strand are not more favorable
than those for addition of the first.  These properties of the methylase are in marked
contrast to those of the endonuclease (Modrich, P., and Zabel, D. (1976) J. Biol. Chem. 251,
5866-5874).  Thus, we suggest that the two proteins interact with their common recognition
sequence in different ways.

<>

<1>Rubin, R.A., Modrich, P.
<2>Purification and properties of EcoRI endonuclease.
<3>Methods Enzymol.
<4>65
<5>96-104
<6>1980
<7>The Escherichia coli RI (EcoRI) DNA restriction and modification enzymes
recognize a common twofold symmetrical hexanucleotide sequence in duplex DNA.
d(pG^pApA*pTpTpCp) d(pCpTpTpA*pAp^Gp) EcoRI restriction endonuclease cleaves
the DNA duplex within this sequence (see arrows in above sequence), while the
modification enzyme methylates the two adenine residues adjacent to the axis of
symmetry (asterisks) to yield 6-methylaminopurine.  The presence of one
6-methylaminopurine residue within this sequence is sufficient to block single-
or double-stranded cleavage by the endonuclease.  EcoRI endonuclease has been
extensively used as a reagent for the preparation of recombinant molecules.  In
addition, the EcoRI enzymes are biochemically simple and hence provide an ideal
system for study of sequence-specific DNA-protein interaction.  We describe
here a convenient method for isolation of large quantities of the endonuclease,
a rapid assay procedure for quantitation of specific endonucleolytic activity,
as well as physical and catalytic properties of the homogeneous protein.

<>

<1>Rubin, R.A., Modrich, P., Vanaman, T.C.
<2>Partial NH2- and C00H-Terminal Sequence Analyses of EcoRI DNA Restriction and Modification Enzymes.
<3>J. Biol. Chem.
<4>256
<5>2140-2142
<6>1981
<7>NH2- and C00H-terminal amino acid sequences of the EcoRI restriction and modification enzymes
have been determined.  The results allow localization of the coding regions within the DNA
segment which controls activity of both enzymes.  Processing of the endonuclease is limited to
removal of NH2-terminal formylmethionine whereas, in the case of the methylase, formylMet-Ala
is removed.

<>

<1>Ruby, E.G., Urbanowski, M., Campbell, J., Dunn, A., Faini, M., Gunsalus, R., Lostroh, P., Lupp, C., McCann, J., Millikan, D., Schaefer, A., Stabb, E., Stevens, A., Visick, K., Whistler, C., Greenberg, E.P.
<2>Complete genome sequence of Vibrio fischeri: A symbiotic bacterium with pathogenic congeners.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>3004-3009
<6>2005
<7>Vibrio fischeri belongs to the Vibrionaceae, a large family of marine gamma-proteobacteria
that includes several dozen species known to engage in a diversity of beneficial or pathogenic
interactions with animal tissue. Among the small number of pathogenic Vibrio species that
cause human diseases are Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus, the
only members of the Vibrionaceae that have had their genome sequences reported. Nonpathogenic
members of the genus Vibrio, including a number of beneficial symbionts, make up the majority
of the Vibrionaceae, but none of these species has been similarly examined. Here we report the
genome sequence of V. fischeri ES114, which enters into a mutualistic symbiosis in the light
organ of the bobtail squid, Euprymna scolopes. Analysis of this sequence has revealed
surprising parallels with V. cholerae and other pathogens.

<>

<1>Ruckert, C., Albersmeier, A., Al-Dilaimi, A., Bednarz, H., Niehaus, K., Szczepanowski, R., Kalinowski, J.
<2>Genome sequence of the squalene-degrading bacterium Corynebacterium terpenotabidum type strain Y-11(T) (= DSM 44721(T)).
<3>Standards in Genomic Sciences
<4>9
<5>505-513
<6>2014
<7>Corynebacterium terpenotabidum Takeuchi et. al 1999 is a member of the genus Corynebacterium,
which contains Gram-positive and non-spore forming bacteria with
a high G+C content. C. terpenotabidum was isolated from soil based on its ability
to degrade squalene and belongs to the aerobic and non-hemolytic Corynebacteria.
It displays tolerance to salts (up to 8%) and is related to Corynebacterium
variabile involved in cheese ripening. As this is a type strain of
Corynebacterium, this project describing the 2.75 Mbp long chromosome with its
2,369 protein-coding and 72 RNA genes will aid the G enomic E ncyclopedia of
Bacteria and Archaea project.

<>

<1>Ruckert, C., Albersmeier, A., Al-Dilaimi, A., Niehaus, K., Szczepanowski, R., Kalinowski, J.
<2>Genome sequence of the halotolerant bacterium Corynebacterium halotolerans type strain YIM 70093(T) (= DSM 44683(T)).
<3>Standards in Genomic Sciences
<4>7
<5>284-293
<6>2012
<7>Corynebacterium halotolerans Chen et al. 2004 is a member of the genus
Corynebacterium which contains Gram-positive bacteria with a high G+C content. C.
halotolerans, isolated from a saline soil, belongs to the non-lipophilic,
non-pathogenic corynebacteria. It displays a high tolerance to salts (up to 25%)
and is related to the pathogenic corynebacteria C. freneyi and C. xerosis. As
this is a type strain in a subgroup of Corynebacterium without complete genome
sequences, this project describing the 3.14 Mbp long chromosome and the 86.2 kbp
plasmid pCha1 with their 2,865 protein-coding and 65 RNA genes will aid the
Genomic Encyclopedia ofBacteria andArchaea project.

<>

<1>Ruckert, C., Kalinowski, J., Heide, L., Apel, A.K.
<2>Draft Genome Sequence of Streptomyces roseochromogenes subsp. oscitans DS 12.976, Producer of the Aminocoumarin Antibiotic Clorobiocin.
<3>Genome Announcements
<4>2
<5>e01147-13
<6>2014
<7>Streptomyces roseochromogenes subsp. oscitans DS 12.976 is the producer of the
gyrase-inhibiting aminocoumarin antibiotic clorobiocin. Here, we present a draft
genome sequence of this strain, in which we identified the clorobiocin gene
cluster as well as an unusually high number (43) of further putative secondary
metabolite clusters.

<>

<1>Ruckert, C., Licht, K., Kalinowski, J., Espirito, S.C., Antwerpen, M., Hanczaruk, M., Reischl, U., Holzmann, T., Gessner, A., Tiemann, C., Grass, G.
<2>Draft Genome Sequence of Bacillus anthracis UR-1, Isolated from a German Heroin User.
<3>J. Bacteriol.
<4>194
<5>5997-5998
<6>2012
<7>We report the draft genome sequence of Bacillus anthracis UR-1, isolated from a fatal case of
injectional anthrax in a German heroin user. Analysis of the genome
sequence of strain UR-1 may aid in describing phylogenetic relationships between
virulent heroin-associated isolates of B. anthracis isolated in the United
Kingdom, Germany, and other European countries.

<>

<1>Rueckert, C., Blom, J., Chen, X.H., Reva, O., Borriss, R.
<2>Genome sequence of B. amyloliquefaciens type strain DSM7T reveals differences to plant-associated B. amyloliquefaciens FZB42.
<3>J. Biotechnol.
<4>155
<5>78-85
<6>2011
<7>The complete genome sequence of Bacillus amyloliquefaciens type strain DSM7T is presented. A
comparative
analysis between the genome sequences of the plant associated strain FZB42 (Chen et al., 2007)
with the genome of B. amyloliquefaciens DSM7T revealed obvious differences in the variable
part of the
genomes, whilst the core genomes were found to be very similar. The strains FZB42 and DSM7T
have in
common 3345 genes (CDS) in their core genomes; whilst 547 and 344 CDS were found to be unique
in
DSM7T and FZB42, respectively. The core genome shared by both strains exhibited 97.89%
identity on
amino acid level. The number of genes representing the core genome of the strains FZB42,
DSM7T, and
Bacillus subtilis DSM10T was calculated as being 3098 and their identity was 92.25%. The
3,980,199 bp
genome of DSM7T contains numerous genomic islands (GI) detected by different methods. Many of
them
were located in vicinity of tRNA, glnA, and glmS gene copies. In contrast to FZB42, but
similar to B. subtilis
DSM10T, the GI were enriched in prophage sequences and often harbored transposases, integrases
and recombinases. Compared to FZB42, B. amyloliquefaciens DSM7T possessed a reduced potential
to
non-ribosomally synthesize secondary metabolites with antibacterial and/or antifungal action.
B. amyloliquefaciens
DSM7T did not produce the polyketides difficidin and macrolactin and was impaired in its
ability to produce lipopeptides other than surfactin. Differences established within the
variable part of
the genomes, justify our proposal to discriminate the plant-associated ecotype represented by
FZB42
from the group of type strain related B. amyloliquefaciens soil bacteria.

<>

<1>Ruepp, A., Graml, W., Santos-Martinez, M.-L., Koretke, K.K., Volker, C., Mewes, H.W., Frishman, D., Stocker, S., Lupas, A.N., Baumeister, W.
<2>The genome sequence of the thermoacidophilic scavenger Thermoplasma acidophilum.
<3>Nature
<4>407
<5>508-513
<6>2000
<7>Thermoplasma acidophilum is a thermoacidophilic archaeon that thrives at 59 C and pH2, which
was isolated from self-heating coal refuse piles and solfatara fields.  Species of the genus
Thermoplasma do not possess a rigid cell wall, but are only delimited by a plasma membrane.
Many macromolecular assemblies from Thermoplasma, primarily proteases and chaperones, have
been pivotal in elucidating the structure and function of their more complex eukaryotic
homologues.  Our interest in protein folding and degradation led us to seek a more complete
representation of the proteins involved in these pathways by determining the genome sequence
of the organism.  Here we have sequenced the 1,564,905-base-pair genome in just 7,855
sequencing reactions by using a new strategy.  The 1,509 open reading frames identify
Thermoplasma as a typical euryarchaeon with a substantial complement of bacteria-related
genes; however, evidence indicates that there has been much lateral gene transfer between
Thermoplasma and Sulfolobus solfataricus, a phylogenetically distance crenarchaeon inhabiting
the same environment.  At least 252 open reading frames, including a complete protein
degradation pathway and various transport proteins, resemble Sulfolobus proteins most closely.

<>

<1>Ruffing, A.M., Castro-Melchor, M., Hu, W.S., Chen, R.R.
<2>Genome Sequence of the Curdlan-Producing Agrobacterium sp. Strain ATCC 31749.
<3>J. Bacteriol.
<4>193
<5>4294-4295
<6>2011
<7>Agrobacterium sp. ATCC 31749 is an industrial strain for the commercial production of curdlan,
an important exopolysaccharide with food and
medical applications. Here we report the genome sequence of the
curdlan-producing strain ATCC 31749. Genome sequencing is the first step
toward the understanding of regulation of curdlan biosynthesis.

<>

<1>Ruiz, S.M., Letourneau, S., Cupples, C.G.
<2>Isolation and characterization of an Escherichia coli strain with a high frequency of C-to-T mutations at 5-methylcytosines.
<3>J. Bacteriol.
<4>175
<5>4985-4989
<6>1993
<7>We used a genetic selection system to isolate a strain of Escherichia coli with a high
frequency of C-to-T transition mutations at the second C of the sequence CCAGG. Cytosines in
other sequences do not mutate to thymine at a high frequency in this strain, and the
frequencies of other base substitution mutations are not increased to the same extent. The
gene responsible for the mutator phenotype has been mapped to 43 min on the E. coli
chromosome. Several lines of evidence indicate that this gene is distinct from the very short
patch repair gene vsr.

<>

<1>Ruiz-Herrera, J., Ruiz-Medrano, R., Dominguez, A.
<2>Selective inhibition of cytosine-DNA methylases by polyamines.
<3>FEBS Lett.
<4>357
<5>192-196
<6>1995
<7>We have advanced the hypothesis that polyamines affect DNA methylation and thus promote the
expression of developmentally controlled genes. We demonstrate that the activity of
cytosine-DNA methyltransferases HpaII, HhaI, HaeIII and SssI is inhibited by physiological
concentrations of polyamines. On the other hand, activity of the adenine-DNA methyltransferase
EcoRI, and restriction enzymes HpaII, HhaI, HaeIII and EcoRI, is insensitive to polyamine
concentrations up to 40 mM. Our results indicate that the effect of polyamines on cytosine-DNA
methyltransferases is rather selective and suggest a possible mode of action in vivo.

<>

<1>Rump, L.V., Strain, E.A., Cao, G., Allard, M.W., Fischer, M., Brown, E.W., Gonzalez-Escalona, N.
<2>Draft Genome Sequences of Six Escherichia coli Isolates from the Stepwise Model of Emergence of Escherichia coli O157:H7.
<3>J. Bacteriol.
<4>193
<5>2058-2059
<6>2011
<7>Enterohemorrhagic Escherichia coli (EHEC) of serotype O157:H7 has been implicated in
food-borne illnesses worldwide. An evolutionary model was
proposed in which the highly pathogenic EHEC O157:H7 serotype arose from
its ancestor, enteropathogenic E. coli (EPEC) O55:H7 (sorbitol fermenting
[SOR(+)] and beta-glucuronidase positive [GUD(+)]), through sequential
gain of virulence, phenotypic traits, and serotype change. Here we report
six draft genomes of strains belonging to this evolutionary model: two
EPEC O55:H7 (SOR(+) GUD(+)) strains, two nonmotile EHEC O157:H(-) strains
(SOR(+) GUD(+)) containing plasmid pSFO157, one EHEC O157:H7 (SOR(-)
GUD(+)) strain, and one O157:H7 strain containing plasmid pSFO157 (SOR(+)
GUD(+)).

<>

<1>Ruoff, B., Johansen, S., Vogt, V.M.
<2>Characterization of the self-splicing products of a mobile intron from the nuclear rDNA of Physarum polycephalum.
<3>Nucleic Acids Res.
<4>20
<5>5899-5906
<6>1992
<7>We have characterized the splicing products formed in vitro from RNA derived from the mobile
group I intron in the nuclear rDNA of Physarum polycephalum, Pp LSU 3. This intron is a close
relative of the well known Tetrahymena intron Tt LSU 1, being inserted at exactly the same
position in the rDNA and sharing about 90% sequence identity with Tt LSU 1 in the conserved
elements characteristic of the catalytic core of all group I introns. However, Pp LSU 3
differs from Tt LSU 1 in that it encodes a site-specific endonuclease, which mediates the
homing of the intron to unoccupied target sites. The endonuclease, I-Ppo, would appear to be a
unique example of a protein encoded by an RNA polymerase I transcript. To gain clues to the
splicing products formed in vivo, and to the nature of the messenger RNA for I-Ppo, we
subjected Pp LSU 3 RNA to standard self-splicing conditions in vitro, and then analyzed the
products by size, by northern blotting, and by primer extension. The results show two novel
features. First, in addition to the expected 5' splice site, there is an alternative 5'
splice site in the upstream exon, just preceding the first codon of the I-Ppo open reading
frame. Second, at the position corresponding to the major circularization site in Tt LSU 1
there is an internal processing site, leading to the efficient separation of two halves of the
excised intron, the 5' half encoding I-Ppo and 3' half containing the ribozyme.
Surprisingly, this cleavage appears not to be due to circularization followed by the
hydrolytic opening of the circle, but rather to G addition. The formation of these products in
vitro suggests how the messenger RNA for the I-Ppo endonuclease may be generated in vivo.

<>

<1>Rusch, D.B. et al.
<2>The Sorcerer II Global Ocean Sampling Expedition: Northwest Atlantic through Eastern Tropical Pacific.
<3>PLoS Biology
<4>5
<5>e77
<6>2007
<7>The world's oceans contain a complex mixture of micro-organisms that are for the most part,
uncharacterized both genetically and biochemically. We report here a metagenomic study of the
marine planktonic microbiota in which surface (mostly marine) water samples were analyzed as
part of the Sorcerer II Global Ocean Sampling expedition. These samples, collected across a
several-thousand km transect from the North Atlantic through the Panama Canal and ending in
the South Pacific yielded an extensive dataset consisting of 7.7 million sequencing reads (6.3
billion bp). Though a few major microbial clades dominate the planktonic marine niche, the
dataset contains great diversity with 85% of the assembled sequence and 57% of the unassembled
data being unique at a 98% sequence identity cutoff. Using the metadata associated with each
sample and sequencing library, we developed new comparative genomic and assembly methods. One
comparative genomic method, termed "fragment recruitment," addressed questions of genome
structure, evolution, and taxonomic or phylogenetic diversity, as well as the biochemical
diversity of genes and gene families. A second method, termed "extreme assembly," made
possible the assembly and reconstruction of large segments of abundant but clearly nonclonal
organisms. Within all abundant populations analyzed, we found extensive intra-ribotype
diversity in several forms: (1) extensive sequence variation within orthologous regions
throughout a given genome; despite coverage of individual ribotypes approaching 500-fold, most
individual sequencing reads are unique; (2) numerous changes in gene content some with direct
adaptive implications; and (3) hypervariable genomic islands that are too variable to
assemble. The intra-ribotype diversity is organized into genetically isolated populations that
have overlapping but independent distributions, implying distinct environmental preference. We
present novel methods for measuring the genomic similarity between metagenomic samples and
show how they may be grouped into several community types. Specific functional adaptations can
be identified both within individual ribotypes and across the entire community, including
proteorhodopsin spectral tuning and the presence or absence of the phosphate-binding gene
PstS.

<>

<1>Rusch, D.B., Lombardo, M.J., Yee-Greenbaum, J., Novotny, M., Brinkac, L.M., Lasken, R.S., Dupont, C.L.
<2>Draft Genome Sequence of a Single Cell of SAR86 Clade Subgroup IIIa.
<3>Genome Announcements
<4>1
<5>e00030-12
<6>2013
<7>SAR86 denotes a 16S clade of gammaproteobacteria that are ubiquitous in ocean surface waters.
So far, SAR86 is resistant to cultivation; thus, little is known about the genome contents or
physiology of this clade. Recently, four partial genome sequences for SAR86 subclades I and II
were published. Here, we present the draft genome sequence of a single cell from SAR86
subgroup IIIa isolated from coastal waters in San Diego, CA.

<>

<1>Ruscher, K., Reuter, M., Kupper, D., Trendelenburg, G., Dirnagl, U., Meisel, A.
<2>A fluorescence based non-radioactive electrophoretic mobility shift assay.
<3>J. Biotechnol.
<4>78
<5>163-170
<6>2000
<7>Electrophoretic mobility shift assay (EMSA) or gel shift assay is one of the most powerful
methods for studying protein-DNA interactions. Typically,
32P-labeled DNA probes containing the sequence bound by the protein of interest are used in
EMSA (rEMSA). Although rEMSA is sensitive and practicable, it relies on the
handling of hazardous radioisotopes, and does not easily allow quantification. We developed a
non-radioactive procedure using fluorescence (Cyano dye Cy5) labeled
oligodeoxynucleotide duplexes as specific probes (fEMSA) and an automatic DNA sequencer for
analysis. Testing different DNA-binding proteins (restriction endonuclease
EcoRII, transcription factor NFkappaB and it's subunit p50) the results in fEMSA and rEMSA
are similar in regard to quality, reproducibility, and sensitivity. fEMSA allows
a semiquantitative screening of large amounts of samples for specific DNA binding activities
and is, therefore, a high throughput technology for semiquantitative analysis of
DNA-protein interaction.

<>

<1>Rushizky, G.W.
<2>Purification of the sequence-specific endonuclease PalI.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>239-242
<6>1981
<7>*
  I. Growth of cells
 II. Purification of PalI
III. Physical and chemical properties of the enzyme
 IV. Purification of other enzymes


<>

<1>Rusling, D.A., Laurens, N., Pernstich, C., Wuite, G.J., Halford, S.E.
<2>DNA looping by FokI: the impact of synapse geometry on loop topology at varied site orientations.
<3>Nucleic Acids Res.
<4>40
<5>4977-4987
<6>2012
<7>Most restriction endonucleases, including FokI, interact with two copies of their recognition
sequence before cutting DNA. On DNA with two sites they act in cis
looping out the intervening DNA. While many restriction enzymes operate
symmetrically at palindromic sites, FokI acts asymmetrically at a non-palindromic
site. The directionality of its sequence means that two FokI sites can be bridged
in either parallel or anti-parallel alignments. Here we show by biochemical and
single-molecule biophysical methods that FokI aligns two recognition sites on
separate DNA molecules in parallel and that the parallel arrangement holds for
sites in the same DNA regardless of whether they are in inverted or repeated
orientations. The parallel arrangement dictates the topology of the loop trapped
between sites in cis: the loop from inverted sites has a simple 180 degrees bend,
while that with repeated sites has a convoluted 360 degrees turn. The ability of
FokI to act at asymmetric sites thus enabled us to identify the synapse geometry
for sites in trans and in cis, which in turn revealed the relationship between
synapse geometry and loop topology.

<>

<1>Rusmintratip, V., Riggs, A.D., Sowers, L.C.
<2>Examination of the DNA substrate selectivity of DNA cytosine methyltransferases using mass tagging.
<3>Nucleic Acids Res.
<4>28
<5>3594-3599
<6>2000
<7>The biological significance of cytosine methylation is as yet incompletely understood, but
substantial and growing evidence strongly suggests that perturbation of methylation patterns,
resulting from the infidelity of DNA cytosine methyltransferase, is an important component of
the development of human cancer. We have developed a novel in vitro assay that allows us to
quantitatively determine the DNA substrate preferences of cytosine methylases. This approach,
which we call mass tagging, involves the labeling of target cytosine residues in synthetic DNA
duplexes with stable isotopes, such as (15)N. Methylation is then measured by the formation of
5-methylcytosine (5mC) by gas chromatography/mass spectrometry. The DNA substrate selectivity
is determined from the mass spectrum of the product 5mC. With the non-symmetrical duplex DNA
substrate examined in this study we find that the bacterial methyltransferase HpaII (duplex
DNA recognition sequence CCGG) methylates the one methylatable cytosine of each strand
similarly. Introduction of an A-C mispair at the methylation site shifts methylation
exclusively to the mispaired cytosine residue. In direct competition assays with HpaII
methylase we observe that the mispaired substrate is methylated more extensively than the
fully complementary, normal substrate, although both have one HpaII methylation site. Through
the use of this approach we will be able to learn more about the mechanisms by which
methylation patterns can become altered.

<>

<1>Rusniok, C., Couve, E., Da Cunha, V., El Gana, R., Zidane, N., Bouchier, C., Poyart, C., Leclercq, R., Trieu-Cuot, P., Glaser, P.
<2>Genome Sequence of Streptococcus gallolyticus: Insights into Its Adaptation to the Bovine Rumen and Its Ability To Cause Endocarditis.
<3>J. Bacteriol.
<4>192
<5>2266-2276
<6>2010
<7>Streptococcus gallolyticus (formerly known as Streptococcus bovis biotype I) is an increasing
cause of endocarditis among streptococci and
frequently associated with colon cancer. S. gallolyticus is part of the
rumen flora but also a cause of disease in ruminants as well as in birds.
Here we report the complete nucleotide sequence of strain UCN34,
responsible for endocarditis in a patient also suffering from colon
cancer. Analysis of the 2,239 proteins encoded by its 2,350-kb-long genome
revealed unique features among streptococci, probably related to its
adaptation to the rumen environment and its capacity to cause
endocarditis. S. gallolyticus has the capacity to use a broad range of
carbohydrates of plant origin, in particular to degrade polysaccharides
derived from the plant cell wall. Its genome encodes a large repertoire of
transporters and catalytic activities, like tannase, phenolic compounds
decarboxylase, and bile salt hydrolase, that should contribute to the
detoxification of the gut environment. Furthermore, S. gallolyticus
synthesizes all 20 amino acids and more vitamins than any other sequenced
Streptococcus species. Many of the genes encoding these specific functions
were likely acquired by lateral gene transfer from other bacterial species
present in the rumen. The surface properties of strain UCN34 may also
contribute to its virulence. A polysaccharide capsule might be implicated
in resistance to innate immunity defenses, and glucan mucopolysaccharides,
three types of pili, and collagen binding proteins may play a role in
adhesion to tissues in the course of endocarditis.

<>

<1>Russell, D.A., Bowman, C.A., Hatfull, G.F.
<2>Genome Sequence of Salmonella enterica subsp. enterica Strain Durban.
<3>Genome Announcements
<4>2
<5>e00399-14
<6>2014
<7>We report the genome sequence of Salmonella enterica subsp. enterica strain Durban, isolated
from a patient with salmonellosis and typhoid fever. The strain
is closely related to S. enterica subsp. enterica strain P125109 but differs in
loss of the SE20 prophage and acquisition of a prophage similar to ELPhiS.

<>

<1>Russell, D.F., Betts, D.H.
<2>Alternative splicing and expression analysis of bovine DNA methyltransferase 1.
<3>Dev. Dyn.
<4>237
<5>1051-1059
<6>2008
<7>Methylation of specific CG residues in the mammalian genome results in tissue-specific
patterns of gene expression, which are critical for
cell differentiation. Embryos that fail to establish and maintain
proper DNA methylation patterns show severe developmental abnormalities
as is the case of DNA methyltransferase 1 (Dnmt1) -deficient embryos.
Dnmt1 is the main maintenance methyltransferase in the mouse and its
expression is regulated by a splicing mechanism that dictates the
expression of stage-specific isoforms. Little is known about Dnmt1
expression in the cow and isoforms of Dnmt1 are yet unknown in this
species. Here we demonstrate that the previously described bovine Dnmt1
transcript is ubiquitously expressed in embryos and fetal tissue. In
addition, we report the identification of a splice variant of the
bovine Dnmt1, which shows a ubiquitous expression pattern. This new
transcript was detected using 5'RACE and genomic mapping and its
expression pattern was shown to be consistent with a tissue-specific
mode of regulation. Furthermore, our analysis shows that the expression
of an oocyte-specific isoform of Dnmt1 is unlikely to occur in cattle.
The newly reported isoform of Dnmt1 was demonstrated to be, similarly
to Dnmt1a, polyadenylated and if translated possess the functional
domains necessary for maintenance and de novo methyltransferase
activity.

<>

<1>Russell, D.W., Hirata, R.K.
<2>The detection of extremely rare DNA modifications.
<3>J. Biol. Chem.
<4>264
<5>10787-10794
<6>1989
<7>DNA methylation in Escherichia coli plays a role in many key cellular processes, including DNA
replication, repair, restriction, and transcription. However, several mutant bacterial strains
exist which are deficient in DNA methylase activities. Thus, it has been suggested that
methylation produced by the dam (DNA adenine methylase) gene is required for the viability of
E. coli and that dam- strains still produce low levels of methylation. Current experimental
methods are not sensitive enough to detect a few potentially essential methylated sites per
genome. Here we describe a method for the detection of N6-methyladenine at specific sites with
a sensitivity of one site in more than 10 megabases. We show that methylation produced by both
the dam and hsd (EcoK) genes is not required for the growth of E. coli and identify the site
of EcoK modification. Minor adaptations of the technique should enable the identification of
other rare DNA modifications.

<>

<1>Russell, D.W., Zinder, N.D.
<2>Hemimethylation prevents DNA replication in E. coli.
<3>Cell
<4>50
<5>1071-1079
<6>1987
<7>The DNA adenine methylase of E. coli methylates adenines at GATC sequences. Strains deficient
in this methylase are transformed poorly by methylated plasmids that depend on either the
pBR322 or the chromosomal origins for replication. We show here that hemimethylated plasmids
also transform dam- bacteria poorly but that unmethylated plasmids transform them at high
frequencies. Hemimethylated daughter molecules accumulate after the transformation of dam-
strains by fully methylated plasmids, suggesting that hemimethylation prevents DNA
replication. We also show that plasmids purified from dam+ bacteria are hemimethylated at
certain sites. These results can explain why newly formed daughter molecules are not
substrates for an immediate reinitiation of DNA replication in wild-type E. coli.

<>

<1>Russo, T.A., Gill, S.R.
<2>Draft Genome Sequence of the Hypervirulent Klebsiella pneumoniae Strain hvKP1, Isolated in Buffalo, New York.
<3>Genome Announcements
<4>1
<5>e0006513
<6>2013
<7>Hypervirulent variants of Klebsiella pneumoniae have been primarily reported in the Asian
Pacific Rim, but they are spreading across the globe. We report the
sequence of K. pneumoniae strain hvKP1, which caused liver-splenic abscesses in
an otherwise healthy 24-year-old from Buffalo, NY, which will assist in
determining why these variants are more pathogenic than 'classic' K. pneumoniae
strains.

<>

<1>Rusu, V., Dorobat-Baron, O., Cosman, M., Lazaroae, D.
<2>Restriction and modification of typing phages by an R factor in S. typhi.
<3>Zentralbl. Bakteriol. [Orig. A]
<4>234
<5>491-501
<6>1976
<7>We have investigated the qualities of one R factor 552 discovered on a strain of S. typhi
resistant to A, C, S, T, non-typable, isolated from stool cultures; from the same patient,
before starting the treatment we isolated, from his blood sample, the strain S. typhi 221,
sensitive to A, C, T, degraded phage-type Vi A.  Factor R 552 fi- when infecting strains of S.
typhi Vi A and of A degraded 221- leads to the conversion of the respective phage-types into
non-typable ones, as a result of the restricting and modifying effect on phage ViA and on the
derivatives resulting from it.  Derivative R 552-1 as a resistance marker to ampicillin has a
restrictive effect on the phage of S. panama A 47 too.  Not taking into account possible
causes such as spontaneous mutation, lysogeny, and adsorption of phages, we reach the
conclusion that R factor 552, through its restrictive effect, is the only cause responsible
for the existence in the same patient of two strains of S. typhi different from the point of
view of phage-type antibiotype.

<>

<1>Rutebuka, O.B.
<2>Molecular studies of restriction genes in enteric bacteria.
<3>Diss. Abstr.
<4>57
<5>4210
<6>1997
<7>Bacterial cells contain various restriction-modification systems to protect themselves from
incoming foreign DNA.  Most bacteria have at least one such system.  In this study, the
restriction genes and the location of the genes on the chromosome were examined in three
enteric bacteria, Salmonella typhimurium LT2, Klebsiella pneumoniae GM 236, and Klebsiella
pneumoniae M5a1.  The 98 minute region of the Salmonella typhimurium LT2 chromosome comprises
at least two R-M systems.  From genetic data this region was previously thought to be only one
minute in length but was found to be larger than two minutes using a physical mapping
technique, pulsed field gel electrophoresis.  The KpnAI and KpnBI R-M systems were recognized
in K. pneumoniae strain M5a1 and GM236, respectively.  A macro-restriction map of Klebsiella
pneumoniae GM236 was constructed in this study using PFGE and Southern hybridization.  The
genome was digested with three rare cutting restriction enzymes (BlnI, I-CeuI, and XbaI).  The
estimated size of the GM236 genome was 4,582 (+/-80) kb, in accordance with the range of other
enteric bacteria.  The partial digest of I-CeuI allowed a tentative ordering of the eight
fragments on a circular map.  The map was remarkably similar to the map of Escherichia coli
and S. typhimurium with the exception of two fragments.  I-CeuI cuts the GM236 chromosome in
eight fragments whereas it cuts E. coli and Salmonella genomes in seven.  This suggests a
possible duplication of the ribosomal cluster in the GM236 strain.  The constructed map,
although tentative, has permitted the mapping of hsdRKpnBI, a restriction gene located on a
322 kb BlnI fragment and also on a 40 kb XbaI fragment.  In this study, hsdRKpnAI, another
restriction gene recognized in K. pneumoniae M5a1, was subcloned and sequenced.  The DNA
sequence of 4.5 kb was determined and only one open reading frame of 3,305 base pairs was
considered as the coding region for the HsdRKpnAI polypeptide.  The nucleotide sequence of the
hsdRKpnAI gene showed no significant similarity to any other sequences in the GenBank
database.  However, the deduced amino acid sequence showed a moderate degree of homology (26%)
with EcoR124II (former EcoR124/3I), a type IC restriction enzyme and KpnBI.  After alignment
of the three proteins, seven helicase motifs, typical of type I and III restriction enzymes,
were found.  This similarity between the three proteins and other current evidence show that
KpnAI and KpnBI are probably the first type I restriction endonucleases recognized in K.
pneumoniae.

<>

<1>Rutebuka, O.B., Lee, N.S., Ryu, J.
<2>DNA sequence analysis of the KpnAI restriction gene of Klebsiella pneumoniae M5a1.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>95
<5>522
<6>1995
<7>We have recently cloned the restriction genes of the KpnAI and KpnBI restriction-modification
systems of K. pneumoniae strains M5aI and GM236 and have reported the nucleotide and deduced
amino acid sequence of the KpnBI gene.  We have now sequenced and analyzed a 4.5 kb
Sau3AI/BamHI restriction fragment of the KpnAI clone, pNLA2, which expresses restriction
activity in r-KpnAI m+KpnAI mutants.  From the DNA sequence, we deduce an open reading frame
(ORF) of 3.3 kb (nucleotides 1156 to 4458) that appears to encode an HsdR polypeptide.
However, the ORF does not show any nucleotide sequence similarity with other genes that are
available from the PC/Gene GenBank.  In spite of the lack of nucleotide sequence similarity,
the deduced amino acid sequence shows a high degree of homology with EcoR124/3, a type IC
restriction enzyme, and the KpnBI polypeptide.  The KpnAI restriction endonuclease, like other
type I and type III endonuclease, also contains sequence motifs characteristic of
superfamily-II helicases which may be involved in DNA unwinding at the cleavage site.  Our
data suggests that the KpnAI system is most likely to be a type I restriction-modification
system.

<>

<1>Rutkauskas, D., Petkelyte, M., Naujalis, P., Sasnauskas, G., Tamulaitis, G., Zaremba, M., Siksnys, V.
<2>Restriction Enzyme Ecl18kI-Induced DNA Looping Dynamics by Single-Molecule FRET.
<3>J. Phys. Chem. B
<4>118
<5>8575-8582
<6>2014
<7>Many type II restriction endonucleases require binding of two copies of a recognition site for
efficient DNA cleavage. Simultaneous interaction of the enzyme with two DNA sites results in
DNA loop formation. It was demonstrated with the tethered particle motion technique that such
looping is a dynamic process where a DNA loop is repeatedly formed and disrupted. Here we use
a better and in the context of protein-induced DNA looping virtually unexploited strategy of
single-molecule Forster resonance energy transfer of surface immobilized biomolecules to
quantitatively study the dynamics of Ecl18kI endonuclease-induced DNA looping and determine
the rate constants of loop formation and disruption. We show that two DNA-bound Ecl18kI dimers
efficiently form a bridging tetramer looping out intervening DNA with a rate that is only a
few orders of magnitude lower than the diffusion limited rate. On the other hand, the
existence of Ecl18kI tetramer is only transient, and the loop is rapidly disrupted within
about 1 s.

<>

<1>Rutkowska, S.M., Skowron, P.M., Bielawski, K., Podhajska, A.J.
<2>SacNI, an isoschizomer of BanII isolated from Streptomyces achromogenes recognizes the 5'-GRGCY/C sequence.
<3>Gene
<4>157
<5>319-320
<6>1995
<7>SacNI, an isoschizomer of the restriction endonuclease, BanII, has been isolated from
Streptomyces achromogenes N-J-H.  SacNI recognizes the palindromic sequence, 5'-GRGCY/C, and
cleaves within the recognition sequence, generating a 3' protruding RGCY end (where R=A or G,
and Y=C or G).

<>

<1>Rutkowska, S.M., Skowron, P.M., Podhajska, A.J.
<2>Purification and characterization of the restriction endonuclease AvcI from Actinomyces cristalomycini.
<3>Gene
<4>157
<5>317-318
<6>1995
<7>The restriction endonuclease AvcI, an isoschizomer of Sau96I was purified from Actinomyces
cristalomycini.  AvcI recognizes a 5-bp palindromic sequence, 5'-G/GNCC and cleaves it after
the first G residue producing a 3-nucleotide 5'-overhang.

<>

<1>Rutter, J.M.
<2>Recent Advances in Microbial Genetics.
<3>Br. Vet. J.
<4>131
<5>284-289
<6>1975
<7>Evolution depends on genetic diversity, and the genetic information contained
within double-stranded deoxyribonucleic acid (DNA) of bacterial cells can be
altered by mutation or by recombination.  Mutations include changes in genes
that occur naturally or are brought about experimentally by agents such as
chemicals or by ionizing radiation.  In contrast, recombination results from an
interaction between one micro-organism with another distinct and separate
micro-organism.  Recombination provides a rapid method for producing changes in
genotype and is generally confined to a sexual process in higher animals.  The
prokaryotic unicellular bacterium differs from eukaryotic animal and plant
cells in that it possesses a single, circular, closed chromosome.  A variety of
different recombination mechanisms have been discovered in bacteria; these
include the entry of naked DNA into bacterial cells (a process called
transformation), the transfer of genetic information incorporated in
bacteriophages (termed transduction), and a sexual process involving transfer
of genetic material between bacterial cells in close apposition (called
conjugation).  Studies of these mechanisms have led to a deeper understanding
of the molecular basis of gene action and are well described (see Hayes, 1968).
Recent advances in our knowledge of nucleic acids have led to the creation in
the laboratory of new combinations of genetic material.  This has been achieved
by splicing together DNA from entirely different sources to form hybrid
molecules that are less likely to occur during evolutionary processes.  This
review is confined to a discussion of these techniques which have caused so
much public disquiet.

<>

<1>Ryan, K.A., Lo, Y.C.
<2>Characterization of a CACAG pentanucleotide repeat in Pasteurella haemolytica and its possible role in modulation of a novel type III restriction-modification system.
<3>Nucleic Acids Res.
<4>27
<5>1505-1511
<6>1999
<7>In a previous study, a recombinant plasmid that contains a CACAG pentanucleotide repeat was
isolated from a Pasteurella haemolytica A1 library.  Southern hybridization analysis using a
(CACAG)5 probe indicated the presence of two loci that contain the pentanucleotide repeats on
the genome of P. haemolytica A1. Additional hybridization analyses against genomic DNA from
related microorganisms indicated that the repeats are only present in P. haemolytica and
Pasteurella trehalosi T3.  The various serotypes of P. haemolytica were found to have either
one or two of the CACAG repeat-containing loci.  Examination of the locus designated Rpt2 by
PCR and sequence analysis indicated that the number of CACAG repeats could change upon serial
subculture which most likely occurs as a result of DNA slipped-strand mispairing.  A plasmid
carrying the Rpt2 locus was isolated and characterized.  Sequence analysis indicated that the
CACAG repeats are contained within the 5'-end of a gene that showed homology to mod genes of
type III restriction-modification systems.  A second open reading frame downstream was
identified which showed homology to res genes of type III restriction-modification systems.
Both the modification and restriction proteins could be expressed and polypeptides of the
expected sizes were detected by SDS-PAGE.  Restriction activity could also be detected in
crude cytoplasmic extracts of Escherichia coli strains carrying the mod and res genes on
recombinant plasmids.

<>

<1>Ryazanova, A., Migur, A., Norkin, M., Timofeyeva, N., Fedorova, O., Kubareva, E.
<2>DNA methyltransferase SsoII: a balance between DNA methylation and transcription repression.
<3>FEBS J.
<4>280
<5>127
<6>2013
<7>Restriction-modification systems serve as primitive immune systems protecting bacterial cells
from phage infection.  R-M system SsoII from Shigella sonnei consists of a DNA
methyltransferase and a restriction endonuclease.  M.SsoII methylates C5 atom of the second
cytosine in the sequence 5'-CCNGG-3' (N=A, C, G or T) in dsDNA while R.SsoII cleaves this
site in case it remains unmethylated.  Since phage DNA is not methylated, R.SsoII hydrolyzes
it and protects the host cell.  In case the endonuclease activity exceeds the
methyltransferase activity, the host cell DNA can be cleaved as well.  Therefore, M. SsoII and
R.SsoII expression should be strictly coordinated.  This coordination is provided by M.SsoII
itself: it represses transcription of its own gene and stimulates transcription of the R.SsoII
gene via binding to the regulatory site which is located in the promoter region of the R-M
system SsoII.  Surpringly, M.SsoII forms a much more stable complex with the regulatory site
than with the methylation site.  Moreover, M.SsoII binding with the former one prevents its
binding with the latter one.  Does M.SsoII still methylate DNA?  We show that (i) M.SsoII
association rate with the methylation site is higher than that for the regulatory site; (ii)
amount of the methylation sites in a bacterial cell is on average 10,000 times higher than
amount of the regulatory sites.  Both these factors direct M.SsoII to the DNA methylation.
M.SsoII has an extremely high affinity to the regulatory site (Kd < nM, i.e. effective binding
even when there are only two regulatory sites per cell).  We conclude that the plasmid-encoded
R-M system SsoII is naturally adjusted for acting at low concentrations.  Therefore, this
system is not a great burden for a cell and has spread among different species and genders of
Enterobacteriaceae.

<>

<1>Ryazanova, A.Y., Kubareva, E.A., Grman, I., Lavrova, N.V., Ryazanova, E.M., Oretskaya, T.S., Hianik, T.
<2>The study of the interaction of (cytosine-5)-DNA methyltransferase SsoII with DNA by acoustic method.
<3>Analyst
<4>136
<5>1227-1233
<6>2011
<7>The interaction of (cytosine-5)-DNA methyltransferase SsoII (M. SsoII) with double-stranded
DNA was studied by means of thickness shear mode
acoustic method (TSM) and gel electrophoresis. M. SsoII recognizes in
double-stranded DNA the methylation site 5'-CCNGG-3' (N = A, C, G, T)
and methylates the inner cytosine residue. M. SsoII also acts as a
transcription factor via binding to the regulatory site
5'-AGGACAAATTGTCCT-3' in the promoter region of SsoII
restriction-modification system. We designed three 60-mer biotinylated
DNA duplexes: with the methylation site (60met), with the regulatory
site (60reg), and without a specific binding site (60oct). A strong
binding of M. SsoII with each one of the studied DNA immobilized on the
TSM transducer has been shown. The equilibrium dissociation constants,
K-D, of the M. SsoII-DNA complexes decreased in the order 60oct > 60reg
> 60met, suggesting a higher stability of M. SsoII-60met complex in
comparison with the others. The association rate constant, k(a), was
also higher for 60met, while similar values were obtained for 60reg and
60oct. The difference in the kinetic parameters for 60met and 60reg
suggested a possible way of coordination between the two M. SsoII
functions in a cell.

<>

<1>Ryazanova, A.Y., Molochkov, N.V., Abrosimova, L.A., Alexeevsky, A.V., Karyagina, A.S., Protsenko, A.S., Friedhoff, P., Oretskaya, T.S., Kubareva, E.A.
<2>Secondary structure of SsoII-like (Cytosine-5)-DNA methyltransferases N-terminal region determined by Circular dichroism spectroscopy.
<3>Mol. Biol. (Mosk)
<4>44
<5>911-921
<6>2010
<7>(Cytosine-5)-DNA methyltransferase SsoII (M.SsoII) has a long N-terminal region (1-71
residues) preceding the sequence with conservative motifs, which are characteristic for all
DNA methyltrans-ferases of such kind. The presence of this region provides M.SsoII capability
to act as a transcription regulator in SsoII restriction-modification system. To perform its
regulatory function, M.SsoII binds specifically to a 15-mer inverted repeat in the promoter
region of SsoII restriction-modification system genes. In the present work, properties of the
protein Delta(72-379)M.Ecl18kI are studied, which is a deletion mutant of the SsoII-like
DNA-methyltransferase M.Ecl18kI and is homologous to M.SsoII N-terminal region.
Delta(72-379)M.Ecl18kI capability to bind specifically a DNA duplex containing the regulatory
site is demonstrated. However, such a binding takes place only in the presence of high protein
excess relative to DNA, which could indicate an altered structure in the deletion mutant in
comparison with the full-length M.SsoII. Circular dichroism spectroscopy demonstrated that
Delta(72-379)M.Ecl18kI has a strongly pronounced secondary structure and contains 32%
alpha-helices and 20% beta-strands. Amino acid sequences alignment of M.SsoII N-terminal
region and transcription factors of known spatial structure is made. An assumption is made how
alpha-helices and beta-strands are arranged in M.SsoII N-terminal region.

<>

<1>Ryazanova, A.Y., Winkler, I., Friedhoff, P., Viryasov, M.B., Oretskaya, T.S., Kubareval, E.A.
<2>Crosslinking of (cytosine-5)-DNA methyltransferase SsoII and its complexes with specific DNA duplexes provides an insight into their structures.
<3>Nucleosides Nucleotides Nucleic Acids
<4>30
<5>632-650
<6>2011
<7>(Cytosine-5)-DNA methyltransferase SsoII (M.SsoII) functions as a methyltransferase and also
as a transcription factor Chemical and photochemical crosslinking was used for exploring the
structure of M.SsoII-DNA complexes and M.SsoII in the absence of DNA. Photocrosslinking with
4-(N-maleimido)benzophenone demonstrated that in the M.SsoII complex with DNA containing the
regulatory site, the M.SsoII region responsible for methylation was bound to DNA flanking the
regulatory site, which contained no methylation sequence. This required high flexibility of
the linker connecting the M.SsoII N-terminal domain and the M.SsoII region responsible for
methylation. The flexibility was demonstrated by crosslinking with bis-maleimidoethane and
1,11-bis-maleimidotetraethyleneglycol.

<>

<1>Rye, P.T.
<2>Biochemical characterization of the Escherichia coli very short patch repair pathway and its coordination with methyltransferase repair of  O6-methylguanine.
<3>Ph.D. Thesis, MIT, USA
<4>
<5>
<6>2006
<7>The E. coli Very Short Patch Repair (VSPR) system corrects T:G mismatches that arise through
Dcm-mediated methylation and subsequent deamination of the underlined cytosine residue in the
palindromic sequence 5'-CCWGG-3' (W is an adenine or thymine). Vsr initiates VSPR by
producing a single stranded nick on the 5' side of the mismatched T. The MutS and MutL
mismatch recognition proteins stimulate this activity, as cells lacking either of these
proteins display diminished VSPR. Genetic studies also indicate that Pol I is responsible for
removing and replacing a short tract of nucleotides downstream of the incision site and that
DNA Ligase seals the nick to complete the repair event. However, until now, biochemical
investigation of the repair steps downstream of Vsr incision have been lacking.  Herein, we
describe two novel in vitro assays used to probe the biochemical events of VSPR. The first was
used to verify the reconstitution of VSPR using purified E. coli Vsr, Pol I, and DNA Ligase
enzymes, while the second was used to measure the distribution of VSPR patch sizes in whole
cell extracts. By monitoring the loss of radiosignal from a series of substrates that
contained the label at prescribed distances downstream of the T:G mismatch, we were able to
determine that VSPR patches are distributed around 2 to 4 deoxynucleotides in length.
Interestingly, under certain reaction conditions, the addition of DNA Ligase improved the
efficiency of repair initiation by Vsr, suggesting that VSPR may be optimal in the context of
a multi-protein complex.  Lastly, we investigated the effect of VSPR proteins on
methyltransferase (MTase) repair of O6-methylguanine (6mG). MTase repair of O6mG opposite T
results in a G:T mismatch that must be further processed to yield the native G:C base pairing.
The G:T mismatch is therefore an intersection of the two pathways and led us to hypothesize
that MTase and VSPR proteins might interact. Indeed, cells lacking the functions of MutS,
MutL, or Vsr proteins displayed decreased MTase repair in vivo, revealing a previously unknown
interaction. The cooperation between proteins of these two repair systems may shed light on
the biological significance of the VSPR system.

<>

<1>Ryu, C.J., Lee, C.H., Yoo, O.J.
<2>A new restriction endonuclease, Bsp1894I, from Bacillus sphaericus 1894.
<3>Korean Biochem. J.
<4>22
<5>444-447
<6>1989
<7>A new restriction endonuclease, Bsp1894I, has been isolated from Bacillus
sphaericus 1894 (KCTC 1188), and its catalytic properties have been studied.
This enzyme recognizes the DNA sequence 5'-G^GNCC-3' and cleaves at the site as
indicated by the arrow.  Bsp1894I is an isoschizomer of AsuI, Sau96I, and
Cfr13I.  It shows maximum activity at a pH range between 6 and 7 in the
presence of 10 mM MgCl2.  The optimum reaction temperature for Bsp1894I is 42C.
Unlike its isoschizomers, Bsp1894I does not require NaCl for optimum activity.

<>

<1>Ryu, J.
<2>The restriction enzyme KpnAI of Klebsiella pneumoniae is a new member of the type ID family.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>97
<5>285
<6>1997
<7>We have described the KpnAI restriction and modification system in Klebsiella pneumoniae M5a1.
The restriction gene (hsdR, 3.3 kb) was cloned into pBR322 and the DNA was sequenced.  In this
project, the corresponding modification genes were identified in the 5' flanking region of
the hsdR gene and cloned into pUC19.  The DNA sequence reveals two open reading frames that
are designated hsdM (1.6kb) and hsdS (1.3kb).  These two genes form a single transcriptional
unit and their coding sequences overlap by five nucleotides.  Thus the KpnAI R-M system
consists of three genes in the order of hsdM, hsdS and hsdR and organized into two
transcriptional units.  These features are typical to the type I restriction enzymes. Homology
studies reveal that the predicted R, M and S peptides share 97%, 98% and 58% homology,
respectively, with the corresponding peptides of the StySBLI restriction enzyme, a prototype
of the type ID family recently identified in Salmonella blegdam.  These high homologies
indicate that the KpnAI restriction enzyme is another member of the type ID family. The KpnAI
enzyme is the first type I restriction enzyme identified in Klebsiella species.

<>

<1>Ryu, J.
<2>Establishment of the KpnAI R-M system of Klebsiella pneumoniae requires prior host modification.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>0
<5>288
<6>1998
<7>Type I restriction-modification systems have been classified into 4 families: IA, IB, IC and
newly identified ID.  They consist of 3 consecutive genes, hsdR, hsdM and hsdS which encode
for subunits R(restriction), M(modification) and S(specificity), respectively.  When entire
R-M genes were transferred to bacteria, without corresponding modification, through either
conjugation, transduction or transformation, the expression of restriction enzymes are somehow
regulated that the R-M genes can be established without degrading recipient DNA.  The basis of
the mechanism of "establishment" still remains to be elucidated.  We have recently cloned R
and MS genes separately for Klebsiella pneumoniae KpnAI R-M system.  In the present project,
the "establishment" of the KpnAI system was studied by using plasmid transformation as a gene
transfer system.  All the three (R, M and S) genes were combined and cloned into pUC18.  When
the entire system was transferred into recipient (E. coli DH5a), no Amp-resistant
transformants were obtained.  However, when the recipient cells were modified with a
modification proficient plasmid, many transformants were obtained.  This result is in sharp
contrast with other known type I enzymes where the entire system is transferred without
killing the recipient.  Thus, the KpnAI R-M enzymes seems to be the first example of the type
I R-M system which requires prior methylation of the chromosome before their establishments.

<>

<1>Ryu, J., Rowsell, E.
<2>Quick identification of Type I restriction enzyme isoschizomers using newly developed pTypeI and reference plasmids.
<3>Nucleic Acids Res.
<4>36
<5>e81
<6>2008
<7>Although DNA-recognition sequences are among the most important characteristics of restriction
enzymes and their corresponding methylases,
determination of the recognition sequence of a Type-I restriction enzyme
is a complicated procedure. To facilitate this process we have previously
developed plasmid R-M tests and the computer program RM search. To
specifically identify Type-I isoschizomers, we engineered a pUC19
derivative plasmid, pTypeI, which contains all of the 27 Type-I
recognition sequences in a 248-bp DNA fragment. Furthermore, a series of
27 plasmids (designated 'reference plasmids'), each containing a unique
Type-I recognition sequence, were also constructed using pMECA, a
derivative of pUC vectors. In this study, we tried those vectors on 108
clinical E. coli strains and found that 48 strains produced isoschizomers
of Type I enzymes. A detailed study of 26 strains using these 'reference
plasmids' revealed that they produce seven different isoschizomers of the
prototypes: EcoAI, EcoBI, EcoKI, Eco377I, Eco646I, Eco777I and Eco826I.
One strain EC1344 produces two Type I enzymes (EcoKI and Eco377I).

<>

<1>Ryu, J.-I., Rajadas, P.T., Bullas, L.R.
<2>Complementation and hybridization evidence for additional families of Type I DNA restriction and modification genes in Salmonella serotypes.
<3>J. Bacteriol.
<4>170
<5>5785-5788
<6>1988
<7>Of eight Salmonella, serB-linked hsd genes for the restriction and modification
of DNA transferred to Escherichia coli/Salmonella hybrids, only two-those with
SM and ST (S. muenchen and S. thompson, respectively) specificities - may have
weakly complemented rSB- and non complemented rK-.  An A-specific DNA probe
failed to hybridize to HindIII-restricted fragments of each of the hybrids, but
an SB (S. typhimurium)-specific probe hybridized to DNA from the hybrid with ST
specificity.  These results indicate that additional families of the type I hsd
genes may exist.

<>

<1>Saavedra, C., Gonzalez, E., Vasquez, C.
<2>Studies on the heterologous expression of BstVI restriction endonuclease in Escherichia coli.
<3>Biochem. Mol. Biol. Int.
<4>44
<5>391-397
<6>1998
<7>Bacterial restriction and modification systems must be regulated to avoid self-restriction.
It is generally accepted that cognate DNA methyltransferases normally protect both, the
host's chromosome and extrachromosomal elements from the activity of their endonuclease
counterparts.  When the bstVIRm genes from Bacillus stearothermophilusV were subcloned into
Escherichia coli, several clones exhibiting a r+m- phenotype were found.  The present work was
undertaken to analyze the possibility that mechanisms other than DNA methylation could account
for the viability of these cells.  No evidence was found for an inhibitory agent or
endonuclease compartmentation.  In vivo experiments showed that lambda phage multiplication
was poorly restricted by the heterologous enzyme.  The restricting activity against the
incoming phage increased however when phage adsortion was performed at higher temperatures.
Analogous experiments in which a DNA-repair deficient strain was used as a host for the
thermophilic R-M system suggested, to some extent, the participation of the repair machinery
in the viability of r-m- clones.

<>

<1>Saavedra, C., Vasquez, C., Encinas, M.V.
<2>Structural studies of the BstVI restriction-modification proteins by fluorescence spectroscopy.
<3>Eur. J. Biochem.
<4>263
<5>65-70
<6>1999
<7>Structural studies of the proteins of the BstVI restriction-modification system of Bacillus
stearothermophilus V were carried out using intrinsic fluorescence techniques. The exposure
and environments of their tryptophanyl residues were determined using collisional quenchers.
Quenching of BstVI endonuclease by iodide suggested a heterogeneous class of tryptophan
residues, while the results obtained with M.BstVI methylase were consistent with a rather
exposed tryptophan population. A comparison of the quenching efficiencies at 20 degrees C and
55 or 60 degrees C showed that their structures are more flexible and open at the temperature
at which they exhibit maximal activity. The endonuclease reached its active conformation only
after 1 h of incubation at 60 degrees C. Fluorescence changes were observed upon Mn2+ and Mg2+
binding, with Kd values in the range 3-5 microM.  The binding of S-adenosyl-L-methionine to
the methylase produced conformational changes, which were consistent with binding to a single
site of Kd 550 and 680 microM at 20 degrees C and 55 degrees C, respectively. Quenching
experiments with iodide showed that the presence of S-adenosyl-L-methionine leads to different
conformational states at 20 degrees C and 55 degrees C.  These results were interpreted in
terms of differences in the structural characteristics of these restriction-modification
proteins as well as in terms of differences in the conformational states that these enzymes
exhibit at 20 degrees C and at the temperature at which they are most active.

<>

<1>Sabat, A.J., Kock, R., Akkerboom, V., Hendrix, R., Skov, R.L., Becker, K., Friedrich, A.W.
<2>Novel Organization of the Arginine Catabolic Mobile Element and Staphylococcal Cassette Chromosome mec Composite Island and Its Horizontal Transfer between Distinct Staphylococcus aureus Genotypes.
<3>Antimicrob. Agents Chemother.
<4>57
<5>5774-5777
<6>2013
<7>In this study, 425 methicillin-resistant Staphylococcus aureus (MRSA) isolates
recovered in the Dutch-German Euregio were investigated for the presence of the
arginine catabolic mobile element (ACME). Sequence analysis by whole-genome
sequencing revealed an entirely new organization of the ACME-staphylococcal
cassette chromosome mec composite island (SCCmec-CI), with truncated ACME type II
located downstream of SCCmec. An identical nucleotide sequence of ACME-SCCmec-CI
was found in two distinct MRSA lineages (t064-ST8 and t002-ST5), which has not
been reported previously in S. aureus.

<>

<1>Sabehi, G., Beja, O., Suzuki, M.T., Preston, C.M., DeLong, E.F.
<2>Different SAR86 subgroups harbour divergent proteorhodopsins.
<3>Environ. Microbiol.
<4>6
<5>903-910
<6>2004
<7>Proteorhodopsins (PRs), bacterial photoactive proton pumps, were originally detected in the
uncultured marine gamma-proteobacterial SAR86
group. PRs are now known to occur in both the gamma and alpha marine
proteobacterial lineages. Recent environmental shotgun sequence analysis
in the Sargasso Sea has added yet more diversity, and a potentially
broader taxonomic distribution, to the PR family. Much remains to be
learned, however, about within-taxon PR variability and the broader
organismal distribution of different PR types. We report here genomic
analyses of large genome fragments from different subgroups of the SAR86
lineage, recovered from naturally occurring bacterioplankton populations
in coastal Red Sea and open ocean Pacific waters. Sequence comparisons
were performed on large bacterial artificial chromosomes (BACs) bearing
both rRNA and PR genes, derived from different SAR86 subgroups. Our
analyses indicated the presence of different PR sequence types within the
same SAR86 rRNA subgroup. The data suggested that the distribution of
particular PR types does not necessarily parallel the phylogenetic
relationship inferred from highly conserved genes such as rRNA. Further
analyses of the genomic regions flanking PR also revealed a potential
pathway for the biosynthesis of retinal, the PR chromophore that is
required to generate the functionally active photoprotein. Finally,
comparison of our results with recently reported Sargasso Sea
environmental shotgun sequence assemblies demonstrated the utility of BAC
clones for interpreting environmental shotgun sequence data, much of which
is represented in short contigs that have an overall low depth of
coverage.

<>

<1>Sabelnikov, A.G., Greenberg, B., Lacks, S.A.
<2>An extended -10 promoter alone directs transcription of the DpnII operon of Streptococcus pneumoniae.
<3>J. Mol. Biol.
<4>250
<5>144-155
<6>1995
<7>The genetic cassette encoding the DpnII restriction-modification system of Streptococcus
pneumoniae gave transcription products of approximately 2.7 and 1.8 kilobases. The larger,
mRNA1, covered both of the methylase genes, dpnM and dpnA, and the endonuclease gene dpnB; the
smaller, mRNA2, covered only the dpnA and dpnB genes. Transcription of mRNA1 was shown to
begin at the translation start site for dpnM, thereby producing an mRNA without any apparent
ribosome-binding site for translation of the DpnM methylase. The promoter for mRNA1 was shown
by base substitution and deletion analysis to consist of an extended -10 site, TaTGgTATAAT,
with no required -35 site. A possible promoter further upstream with close matches to a -35
site and a nonextended -10 site was not used. A survey of 36 proven and putative promoters
used by S. pneumoniae revealed that 61% of them contained the full -10 extension, although,
other than the dpmM promoter, they matched at a -35 site, as well. It appears that, unlike
those found in Escherchia coli, S. pneumoniae promoters frequently require an extended -10
site, and such a site can function naturally without a -35 site.

<>

<1>Sabirova, J.S., Xavier, B.B., Hernalsteens, J.P., De Greve, H., Ieven, M., Goossens, H., Malhotra-Kumar, S.
<2>Complete Genome Sequences of Two Prolific Biofilm-Forming Staphylococcus aureus Isolates Belonging to USA300 and EMRSA-15 Clonal Lineages.
<3>Genome Announcements
<4>2
<5>e00610-14
<6>2014
<7>Methicillin-resistant Staphylococcus aureus (MRSA) causes serious infections that are even
more difficult to treat when associated with a biofilm phenotype that
facilitates evasion of the host immune system and antibiotics. As a first step
toward understanding the mechanisms underlying biofilm formation, we sequenced
the genomes of two prolific biofilm-forming strains belonging to the two most
important globally disseminated clonal lineages, USA300 and EMRSA-15.

<>

<1>Sack, G.H. Jr., Nathans, D.
<2>Studies of SV40 DNA. IV.  Cleavage of SV40 DNA by restriction endonuclease from Hemophilus parainfluenzae.
<3>Virology
<4>51
<5>517-520
<6>1973
<7>None

<>

<1>Sadri, R., Hornsby, P.J.
<2>Rapid analysis of DNA methylation using new restriction enzyme sites created by bisulfite modification.
<3>Nucleic Acids Res.
<4>24
<5>5058-5059
<6>1996
<7>Bisulfite converts non-methylated cytosine in DNA to uracil leaving 5-methylcytosine
unaltered.  Here, predicted changes in restriction enzyme sites following reaction of genomic
DNA with bisulfite and amplification of the product by the polymerase chain reaction (PCR)
were used to assess the methylation of CpG sites.  This procedure differs from conventional
DNA methylation analysis by methylation-sensitive restriction enzymes because it does not rely
on an absence of cleavage to detect methylated sites, the two strands of DNA produce different
restriction enzyme sites and may be differentially analyzed, and closely related sequences may
be separately analyzed by using specific PCR primers.

<>

<1>Sadykov, M., Asami, Y., Niki, H., Handa, N., Itaya, M., Tanokura, M., Kobayashi, I.
<2>Multiplication of a restriction-modification gene complex.
<3>Mol. Microbiol.
<4>48
<5>417-427
<6>2003
<7>Previous works have suggested that some gene complexes encoding a restriction (R) enzyme and a
cognate modification (M) enzyme may behave as
selfish mobile genetic elements. RM gene complexes, which destroy
'non-self' elements marked by the absence of proper methylation, are often
associated with mobile genetic elements and are involved in various genome
rearrangements. Here, we found amplification of a restriction-modification
gene complex. BamHI gene complex inserted into the Bacillus chromosome
showed resistance to replacement by a homologous stretch of DNA. Some
cells became transformed with the donor without losing BamHI. In most of
these transformants, multiple copies of BamHI and the donor allele were
arranged as tandem repeats. When a clone carrying one copy of each allele
was propagated, extensive amplification of BamHI and the donor unit was
observed in a manner dependent on restriction enzyme gene. This suggests
that restriction cutting of the genome participates in the amplification.
Visualization by fluorescent in situ hybridization revealed that the
amplification occurred in single cells in a burst-like fashion that is
reminiscent of induction of provirus replication. The multiplication
ability in a bacterium with natural capacity for DNA release, uptake and
transformation will be discussed in relation to spreading of RM gene
-complexes.

<>

<1>Sadykov, M., Handa, N., Asami, Y., Tanokura, M., Itaya, M., Kobayashi, I.
<2>Selfish maintenance and amplification of BamHI restriction modification gene complex on Bacillus subtilis chromosome.
<3>Mutat. Res.
<4>483
<5>S159
<6>2001
<7>It has been shown that some type II RM (restriction modification) gene complexes on a plasmid
resist replacement by an incompatible plasmid through post-segregational host killing.  Here
we present experimental evidence for selfish maintenance of BamHI RM gene complex on Bacillus
subtilis chromosome.  pBR322 derivatives carrying BamHI restriction +/- modification gene
complex and neomycin resistance gene were inserted into B. subtilis chromosome.  We then tried
to replace them by a homologous stretch of DNA that contains spectinomycin resistance gene by
natural transformation.  The efficiency of apparent replacement was several fold less, and the
resulting transformant colonies were smaller in the r+ cells than in the r- cells.  Moreover,
some of these clones were able to grow on media containing both spectinomycin and neomycin.
By PCR, we found that some of these clones retained the recipient RM gene complex and neomycin
phosphotransferase gene as well as the donor spectinomycin resistance gene.  Southern analysis
of chromosomal DNA provided supporting evidence.  Moreover, by Southern analyses and DNA
sequencing we found genome rearrangements, i.e. "amplicon" structures with different level of
amplification.  Possible mechanisms of gene amplification will be discussed.

<>

<1>Saenz, H.L., Engel, P., Stoeckli, M.C., Lanz, C., Raddatz, G., Vayssier-Taussat, M., Birtles, R., Schuster, S.C., Dehio, C.
<2>Genomic analysis of Bartonella identifies type IV secretion systems as host adaptability factors.
<3>Nat. Genet.
<4>39
<5>1469-1476
<6>2007
<7>The bacterial genus Bartonella comprises 21 pathogens causing characteristic intraerythrocytic
infections. Bartonella bacilliformis is a
severe pathogen representing an ancestral lineage, whereas the other
species are benign pathogens that evolved by radial speciation. Here, we
have used comparative and functional genomics to infer pathogenicity genes
specific to the radiating lineage, and we suggest that these genes may
have facilitated adaptation to the host environment. We determined the
complete genome sequence of Bartonella tribocorum by shotgun sequencing
and functionally identified 97 pathogenicity genes by signature-tagged
mutagenesis. Eighty-one pathogenicity genes belong to the core genome
(1,097 genes) of the radiating lineage inferred from genome comparison of
B. tribocorum, Bartonella henselae and Bartonella quintana. Sixty-six
pathogenicity genes are present in B. bacilliformis, and one has been lost
by deletion. The 14 pathogenicity genes specific for the radiating lineage
encode two laterally acquired type IV secretion systems, suggesting that
these systems have a role in host adaptability.

<>

<1>Sagarkar, S., Bhardwaj, P., Yadav, T.C., Qureshi, A., Khardenavis, A., Purohit, H.J., Kapley, A.
<2>Draft genome sequence of atrazine-utilizing bacteria isolated from Indian agricultural soil.
<3>Genome Announcements
<4>2
<5>e01149-13
<6>2014
<7>We report the draft genome sequences of two tropical bacterial isolates capable of degrading
the herbicide atrazine. Alcaligenes sp. strain EGD-AK7 and
Arthrobacter sp. strain AK-YN10 were isolated from Indian agricultural soil in
which sugarcane is grown, with a reported history of atrazine use. EGD-AK7 has
the atzABCDEF genes and AK-YN10 has the trzN and atzBC genes for atrazine
degradation.

<>

<1>Sagawa, H., Ohshima, A., Kato, I.
<2>Sse8387I, a useful eight base cutter for mammalian genome analysis (influence of methylation on the activity of Sse8387I).
<3>Nucleic Acids Res.
<4>23
<5>2367-2370
<6>1995
<7>To develop restriction enzymes that are useful for genome analysis, we previously performed
screening and isolated Sse8387I from Streptomyces sp. strain 8387. Sse8387I is a restriction
enzyme that recognizes 5'-CCTGCA/GG-3' and cleaves DNA at the site shown by the diagonal.
The present study evaluated the effects of methylation that is important when Sse8387I is used
for genome analysis. Sse8387I lost cleavage activity after methylation of adenine or
methylation of cytosine at any site in the recognition sequence. However, the recognition
sequence of Sse8387I contains no CG sequence, which is the mammalian methylation sequence. In
addition, we evaluated the effects of methylation of CG at sites other than the recognition
sequence. The cleavage activity of Sse8387I was maintained even when CG sequences were present
immediately before or after, or near the recognition sequence, and cytosine was methylated.
These results suggest that CG methylation does not affect the cleavage activity of Sse8387I.
Therefore, Sse8387I seems to be very useful for mammalian genome analysis.

<>

<1>Sagawa, H., Takagi, M., Nomura, Y., Inagaki, K., Tano, T., Kishimato, N., Kotani, H., Nakajima, K.
<2>Isolation and identification of restriction endonuclease Aor51HI from Acidiphilium organovorum 51H.
<3>Nucleic Acids Res.
<4>20
<5>365
<6>1992
<7>Aor5HI, a type II restriction endonuclease, has been isolated from Acidiphilium
organovorum 51H, an isoschizomer of Eco47III, recognizes the palindromic six
base sequence 5'-AGCGCT-3', and cleaves the center of recognition sequence
giving the blunted end.

<>

<1>Sagawa, H., Uemori, T., Mukai, H., Yamamoto, J., Tomono, J., Kobayashi, E., Enoki, T., Asada, K., Kato, I.
<2>A stabilization method and a preservation method for a reagent for nucleic acid amplification or detection reaction.
<3>International Patent Office
<4>WO 02101042 A
<5>
<6>2002
<7>A method of stabilizing a reaction reagent for highly sensitively and specifically amplifying
a target nucleic acid in a sample with the use of a chimeric oligonucleotide primer and a
method of storing the same over a long time; and a method of highly sensitively detecting a
pathogenic microorganism and a virus.

<>

<1>Sagawa, H., Uemori, T., Mukai, H., Yamamoto, J., Tomono, J., Kobayashi, E., Enoki, T., Asada, K., Kato, I.
<2>A stabilization method and a preservation method for a reagent fornucleic acid amplification or detection reaction.
<3>Korean Patent Office
<4>KR 1020037015577 A
<5>
<6>2003
<7>
<>

<1>Sagawa, H., Ueno, H., Oshima, A., Kato, I.
<2>Plasmid vector for expression of genes such as AccIII restriction enzyme gene - plasmid pACE601, plasmid pACE611, plasmid pACE701 and plasmid pACE702 for inducible recombinant restriction endonuclease production in Escherichia coli.
<3>International Patent Office
<4>WO 9734006
<5>
<6>1997
<7>A plasmid vector characterized by having a promoter sequence that can be recognized by an RNA
polymerase which is not inherent in a host and that controls the expression of target genes
and a replication origin that increases the number of copies under the induction by foreign
factors; methods of expression and isolation of target genes by using the vector; a
polypeptide having the activity of an AccIII restriction enzyme; and a DNA encoding the
polypeptide.  The invention provides for the first time a plasmid vector which can introduce a
foreign target gene encoding proteins which are fatal or noxious to hosts into said hosts, a
method of efficiently expressing the proteins by using the vector, and also a method of
permitting a restriction enzyme gene constituting a restriction-modification system to be
isolated even in the absence of a modification enzyme gene, which has been difficult according
to prior art.

<>

<1>Sagawa, H., Ueno, H., Oshima, A., Kato, I.
<2>Plasmid.
<3>US Patent Office
<4>US 6165749
<5>none
<6>2000
<7>A plasmid vector characterized by comprising a promoter sequence that can be recognized by an
RNA polymerase which is not inherent in a host
and that controls the expression of desired genes and a replication
origin that increases the number of copies under the induction by
exogenous factors; methods for expression and isolation of target genes
by using the vector; a polypeptide having the activity of an AccIII
restriction endonuclease; and a DNA encoding the polypeptide. The
invention provides for the first time a plasmid vector which can
introduce an exogenous desired gene encoding proteins which are lethal
or harmful to hosts into the hosts, a method for efficiently expressing
the proteins by using the vector, and also a method for permitting a
restriction endonuclease gene constituting a restriction-modification
system to be isolated even in the absence of a modification enzyme
gene, which has been difficult in the prior arts.

<>

<1>Sagawa, H., Ueno, H., Oshima, A., Kato, I.
<2>Restriction enzyme and gene thereof.
<3>Japanese Patent Office
<4>JP 2000325094
<5>
<6>2000
<7>The cloned AccIII RM system.

<>

<1>Sagawa, H., Ueno, H., Oshima, A., Kato, I.
<2>Gene isolation method.
<3>US Patent Office
<4>US 6770481 A
<5>
<6>2004
<7>A plasmid vector characterized by comprising a promoter sequence that can be recognized by an
RNA polymerase which is not inherent in a host and that controls the expression of desired
genes and a replication origin that increases the number of copies under the induction by
exogenous factors; methods for expression and isolation of target genes by using the vector; a
polypeptide having the activity of an AccIII restriction endonuclease; and a DNA encoding the
polypeptide.  The invention provides for the first time a plasmid vector which can introduce
an exogenous desired gene encoding proteins which are lethal or harmful to hosts into the
hosts, a method for efficiently expressing the proteins by using the vector, and also a method
for permitting a restriction endonuclease-gene constituting a restriction-modification system
to be isolated even in the absence of a modification enzyme gene, which has been difficult in
the prior arts.

<>

<1>Sager, R., Kitchin, R.
<2>Selective Silencing of eukaryotic DNA.
<3>Science
<4>189
<5>426-433
<6>1975
<7>A molecular basis is proposed for programmed inactivation or loss of eukaryotic
DNA.

<>

<1>Sager, R., Lane, D.
<2>Molecular basis of maternal inheritance.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>69
<5>2410-2413
<6>1972
<7>The mechanism of preferential transmission (i.e., maternal inheritance) of
cytoplasmic genes was investigated with chloroplast DNA of Chlamydomonas as a
model system.  The behavior of nuclear and chloroplast DNAs were compared in
the sexual cycle; DNAs from male and female parents were distinguished by
labeling with 14N- or 15NH4Cl and then by making the crosses: 14N (female) X
15N (male) and the reciprocal.  Chloroplast DNAs from the two parents followed
different paths in the zygote, but nuclear DNAs showed no differences.
Chloroplast DNA from the female parent persists in the zygote, but undergoes a
density shift of 0.003-0.005 g/cm3 to a lighter buoyant density, whereas that
from the male disappears soon after zygote formation.  The possibility is
discussed that a modification-restriction system may be involved.

<>

<1>Sagitov, V.R., Aleksandrov, A.A.
<2>Selectivity of DNA methylase BspRI.
<3>Dokl. Akad. Nauk.
<4>298
<5>1266-1268
<6>1988
<7>None

<>

<1>Saguez, C., Lecellier, G., Koll, F.
<2>Intronic GIY-YIUG endonuclease gene in the mitochondrial genome of Podospora curvicolla: evidence for mobility.
<3>Nucleic Acids Res.
<4>28
<5>1299-1306
<6>2000
<7>Endonuclease genes encoded in invasive introns are themselves supposed to be mobile elements
which, during evolution, have colonized pre-existing introns converting them into invasive
elements.  This hypothesis is supported by numerous data concerning the LAGLI-DADG subclass of
intronic endonucleases. Less is known about the GIY-YIG ORFs which constitute another family
of endonucleases. In this paper we describe the presence of one optional GIY-YIG ORF in the
second intron of the mitochondrial cytochrome b gene in the fungus Podospora curvicolla. We
show that this GIY-YIG ORF is efficiently transferred from an ORF-containing intron to an
ORF-less allele. We also show that the products of both the GIY-YIG ORF and the non-canonical
LAGLI-DADG-GIY-YIG ORF, which is generated by its integration, have endonuclease activities
which recognize and cut the insertion site of the optional sequence. This constitutes the
first direct evidence for potential mobility of an intronic GIY-YIG endonuclease. We discuss
the role that such a mobile sequence could have played during evolution.

<>

<1>Saha, S., Ahmad, I., Reddy, Y.V.R., Krishnamurthy, V., Rao, D.N.
<2>Functional analysis of conserved motifs in type III restriction-modification enzymes.
<3>Biol. Chem.
<4>379
<5>511-517
<6>1998
<7>EcoP1I and EcoP15I are members of type III restriction-modification enzymes.  EcoPI and
EcoP15I DNA methyltransferases transfer a methyl group from S-adenosyl-L-methionine to the N6
position of the second adenine residues in their recognition sequences, 5'-AGACC-3' and
5'-CAGCAG-3' respectively.  We have altered various residues in two highly conserved
sequences, FxGxG (motif I) and DPPY (motif IV) in these proteins by site-directed mutagenesis.
Using a mixture of in vivo and in vitro assays, our results on the mutational analysis of
these methyltransferases demonstrate the universal role of motif I in AdoMet binding and a
role for motif IV in catalysis.  All six cysteine residues in EcoP15I DNA methyltransferase
have been substituted with serine and the role of cysteine residues in EcoP15I DNA
methyltransferase catalyzed reaction assessed.  The Res subunits of type III restriction
enzymes share a distant sequence similarity with and contain the motifs characteristic of the
DEAD box proteins.  We have carried out site-directed mutagenesis of the conserved residues in
two of the helicase motifs of the EcoP1I restriction enzyme in order to investigate the role
of motifs in DNA cleavage by this enzyme.  Our findings indicate that certain conserved
residues in these motifs are involved in ATP hydrolysis while the other residues are involved
in coupling restriction of DNA to ATP hydrolysis.  Taken collectively, these results form the
basis for a detailed structure-function analysis of EcoP1I and EcoP15I restriction enzymes.

<>

<1>Saha, S., Rao, D.N.
<2>ATP hydrolysis is required for DNA cleavage by EcoPI restriction enzyme.
<3>J. Mol. Biol.
<4>247
<5>559-567
<6>1995
<7>The type III restriction endonuclease EcoPI, coded by bacteriophage P1, cleaves unmodified DNA
in the presence of ATP and magnesium ions. We show that purified EcoPI restriction enzyme
fails to cleave DNA in the presence of non-hydrolyzable ATP analogs. More importantly, this
study demonstrates that EcoPI restriction enzyme has an inherent ATPase activity, and ATP
hydrolysis is necessary for DNA cleavage. Furthermore, we show that the progress curve of the
reaction with EcoPI restriction enzyme exhibits a lag which is dependent on the enzyme
concentration. Kinetic analysis of the progress curves of the reaction suggest slow
transitions that can occur during the reaction, characteristic of hysteretic enzymes. The role
of ATP in the cleavage mechanism of type III restriction enzymes is discussed.

<>

<1>Saha, S., Rao, D.N.
<2>Mutations in the Res subunit of the EcoPI restriction enzyme that affect ATP-dependent reactions.
<3>J. Mol. Biol.
<4>269
<5>342-354
<6>1997
<7>The Res subunits of the type III restriction-modification enzymes share a statistically
significant amino acid sequence similarity with several RNA and DNA helicases of the so-called
DEAD family.  It was postulated that in type III restriction enzymes a DNA helicase activity
may be required for local unwinding at the cleavage site.  The members of this family share
seven conserved motifs, all of which are found in the Res subunit of the type III restriction
enzymes.  To determine the contribution, if any, of these motifs in DNA cleavage by EcoPI, a
type III restriction enzyme, we have made changes in motifs I and II.  While mutations in
motif I (GTGKT) clearly affected ATP hydrolysis and resulted in loss of DNA cleavage activity,
mutation in motif II (DEPH) significantly decreased ATP hydrolysis but had no effect on DNA
cleavage.  The double mutant R.EcoPIK90R-H229K showed no significant ATPase or DNA restriction
activity though ATP binding was not affected.  These results imply that there are at least two
ATPase reaction centers in EcoPI restriction enzyme.  Motif I appears to be involved in
coupling DNA restriction to ATP hydrolysis.  Our results indicate that EcoPI restriction
enzyme does not have a strand separation activity.  We suggest that these motifs play a role
in the ATP-dependent translocation that has been proposed to occur in the type III restriction
enzymes.

<>

<1>Sahl, J.W., Stone, J.K., Gelhaus, H.C., Warren, R.L., Cruttwell, C.J., Funnell, S.G., Keim, P., Tuanyok, A.
<2>Genome Sequence of Burkholderia pseudomallei NCTC 13392.
<3>Genome Announcements
<4>1
<5>e00183-13
<6>2013
<7>Here, we describe the draft genome sequence of Burkholderia pseudomallei NCTC 13392. This
isolate has been distributed as K96243, but distinct genomic
differences have been identified. The genomic sequence of this isolate will
provide the genomic context for previously conducted functional studies.

<>

<1>Saikawa, Y., Kubota, T., Maeda, S., Otani, Y., Kumai, K., Kitajima, M.
<2>Inhibition of DNA methyltransferase by antisense oligodeoxynucleotide modifies cell characteristics in gastric cancer cell lines.
<3>Oncol. Rep.
<4>12
<5>527-531
<6>2004
<7>DNA (cytosine-5-)-methyltransferase 1 (DNMT1) plays an important role in the maintenance of
DNA methylation patterns via complicated networks
including signaling pathways and transcriptional factors, relating to
cell differentiation or carcinogenesis. In the present study, we
designed an antisense oligodeoxynucleotide of DNMT1 (AS/MT: 5'-CGGTAC
GCGCCGGCATCT-3') and demonstrated successful inhibition of DNMT1
expression by AS/MT at the protein level, using gastric cancer cell
lines in vitro. E-cadherin protein expression was increased, and both
cyclin D1 and PCNA were decreased by AS/MT treatment. AS/MT also
induced suppression of cell growth as determined by BrDU uptake
incorporation, in a dose-dependent manner, suggesting specificity of
AS/MT. Simultaneously, morphological alterations were observed in both
TMK-1 and MKN-45 cells after 24 h incubation with 2 muM of AS/MT. The
cells changed shape from their original forms to dispersed,
fibroblast-like cells with neurite-like processes, accompanied by an
increased adhesive potential of the cells. An in vivo model of
peritoneal dissemination using the nude mouse system showed an
increased malignant potential of AS/MT treated TMK-1 cells as
demonstrated by a greater number of peritoneal tumor nodules in the
AS/MT as compared to the NS/MT treated group, 34.8+/-4.3 vs. 22.4+/-3.0
nodules, respectively (p=0.0039). The total wet tumor weight in the
AS/MT group (350+/-47.4 g) was significantly greater than that in the
NS/MT group (248+/-41.5 g) (p=0.0065). In conclusion, the inhibition of
DNA methylation by DNMT1 by an antisense oligodeoxynucleotide
influences cell morphology and adhesion, as well as cell growth in
gastric cancer cells in vitro. Moreover, these alterations in the
characteristics of cancer cells resulted in an increased ability to
attach onto the peritoneum in the nude mouse system in vivo, suggesting
that strict clinical guidelines will be necessary to utilize such a DNA
methylation inhibitor, since it does not always mean a therapeutic
antitumor strategy.

<>

<1>Sain, B., Murray, N.E.
<2>The hsd (host specificity) genes of E. coli K12.
<3>Mol. Gen. Genet.
<4>180
<5>35-46
<6>1980
<7>The hsd genes of E. coli K12 have been cloned in phage lambda by a combination
of in vitro and in vivo techniques.  Three genes, whose products are required
for K-specific restriction and modification, have been identified by
complementation tests as hsdR, M, and S.  The order of these closely linked
genes was established as R, M, S by analysis of the DNA of genetically
characterised deletion derivatives of lambda hsd phages.  The three genes are
transcribed in the same direction but not necessarily as a single operon.
Genetic evidence identifies two promoters, one from which transcription of hsdM
and S is initiated and a second for the hsdR gene.  The hsdR gene codes for a
polypeptide of molecular weight ~130,000; hsdM for one of 62-65,000 and the
hsdS gene was associated with two polypeptides of approximately 50000.
Circumstantial evidence suggest that one of these two polypeptides may be a
degradation, or processed, derivative of the other.  The hsdS polypeptide of E.
coli B has a slightly higher mobility in an SDS-polyacrylamide gel than does
that of E. coli K12.  A probe comprising most of the hsdR gene and all of the
hsdM and S genes of E. coli K12 shares extensive homology with the DNA of E.
coli B but none with that of E. coli C.

<>

<1>Sait, M., Aitchison, K., Wheelhouse, N., Wilson, K., Lainson, F.A., Longbottom, D., Smith, D.G.
<2>Genome Sequence of Lawsonia intracellularis Strain N343, Isolated from a Sow with Hemorrhagic Proliferative Enteropathy.
<3>Genome Announcements
<4>1
<5>e00027-13
<6>2013
<7>is the etiological agent of proliferative enteropathy (PE), causing mild or acute hemorrhagic
diarrhea in infected animals. Here we report the genome sequence of
strain N343, isolated from a sow that died of hemorrhagic PE. N343 contains 24
single nucleotide polymorphisms and 90 indels compared to the reference strain
PHE/MN1-00.

<>

<1>Saito, H., Shibata, T., Ando, T.
<2>Mapping of genes determining nonpermissiveness and host-specific restriction to bacteriophages in Bacillus subtilis Marburg.
<3>Mol. Gen. Genet.
<4>170
<5>117-122
<6>1979
<7>Bacillus subtilis Marburg is nonpermissive for the multiplication of
bacteriophages SP10 and uNR2.  A permissive mutant was derived from the Marburg
strain, and the genetic determinants of nonpermissiveness were analyzed by PBS1
transduction.  The simultaneous presence of two genes as mutant alleles, nonA
and nonB, was necessary for permissiveness.  The gene nonA is linked very
closely to rfm (cotransfer: 90%); nonB is located between dal and purB
(cotransfer of nonB and purB6: 48%).  The genetic determinant of host-specific
restriction intrinsic to the Marburg strain (hsrM) was found to be identical or
very closely linked to nonB.  The segregation of nonB and hsrM has never been
observed in the course of transduction analysis.  The mutation, hsrM1,
diminishes the restriction activity, but not the host-controlled modifiction.

<>

<1>Sakaguchi, Y., Hayashi, T., Kurokawa, K., Nakayama, K., Oshima, K., Fujinaga, Y., Ohnishi, M., Ohtsubo, E., Hattori, M., Oguma, K.
<2>The genome sequence of Clostridium botulinum type C neurotoxin-converting phage and the molecular mechanisms of unstable lysogeny.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>17472-17477
<6>2005
<7>Botulinum neurotoxins (BoNTXs) produced by Clostridium botulinum are among
the most poisonous substances known. Of the seven types of BoNTXs, genes
for type C1 and D toxins (BoNTX/C1 and D) are carried by bacteriophages.
The gene for exoenzyme C3 also resides on these phages. Here, we present
the complete genome sequence of c-st, a representative of
BoNTX/C1-converting phages. The genome is a linear double-stranded DNA of
185,682 bp with 404-bp terminal direct repeats, the largest known
temperate phage genome. We identified 198 potential protein-coding
regions, including the genes for production of BoNTX/C1 and exoenzyme C3.
Very exceptionally, as a viable bacteriophage, a number of insertion
sequences were found on the c-st genome. By analyzing the molecular
structure of the c-st genome in lysogens, we also found that it exists as
a circular plasmid prophage. These features account for the unstable
lysogeny of BoNTX phages, which has historically been called
"pseudolysogeny." The PCR scanning analysis of other BoNTX/C1 and D phages
based on the c-st sequence further revealed that BoNTX phages comprise a
divergent phage family, probably generated by exchanging genomic segments
among BoNTX phages and their relatives.

<>

<1>Sakaguchi, Y., Hosomi, K., Uchiyama, J., Ogura, Y., Umeda, K., Sakaguchi, M., Kohda, T., Mukamoto, M., Misawa, N., Matsuzaki, S., Hayashi, T., Kozaki, S.
<2>Draft Genome Sequence of Clostridium botulinum Type B Strain Osaka05, Isolated from an Infant Patient with Botulism in Japan.
<3>Genome Announcements
<4>2
<5>e01010-13
<6>2014
<7>Clostridium botulinum strain Osaka05, which has been isolated from an infant patient with
botulism in Japan, is the first strain producing botulinum
neurotoxin subtype B6. Here, we report the draft genome sequence of C. botulinum
Osaka05.

<>

<1>Sakamoto, M., Oshima, K., Suda, W., Kitamura, K., Iida, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequences of Three Strains of Bacteroides pyogenes Isolated from a Cat and Swine.
<3>Genome Announcements
<4>2
<5>e01242-13
<6>2014
<7>Here, we report the draft genome sequences of Bacteroides pyogenes JCM 6294(T), JCM 6292, and
JCM 10003, which were isolated from a cat and swine and were
recently classified into a single species, B. pyogenes. Comparative analyses of
these genomes revealed the diversification of B. pyogenes strains isolated from
different animals.

<>

<1>Sakamoto, M., Tanaka, N., Shiwa, Y., Yoshikawa, H., Ohkuma, M.
<2>Draft Genome Sequences of Porphyromonas crevioricanis JCM 15906T and Porphyromonas cansulci JCM 13913T Isolated from a Canine Oral Cavity.
<3>Genome Announcements
<4>1
<5>e00483-13
<6>2013
<7>Here, we report the draft genome sequences of Porphyromonas crevioricanis JCM 15906(T) and
Porphyromonas cansulci JCM 13913(T), which were isolated from a
canine oral cavity and were recently united under the single species P.
crevioricanis. These two genome sequences are very similar, and yet a high degree
of genome rearrangements is observed.

<>

<1>Sakurai, T., Kosaka, T.
<2>Restriction enzyme manufacture by culturing Bifidobacterium and an enzyme recognising a specific arrangement in double-stranded DNA.
<3>US Patent Office
<4>US 4542099
<5>
<6>1983
<7>The method comprises culturing a bacterial strain of Bifidobacterium bifidum which produces a
restriction enzyme which can recognise the base arrangement GRCGYC in double-stranded DNA. The
restriction enzyme is then collected from the bacteria. Preferably the restriction enzyme
(abbr. BbiII) produced by B. bifidum is the isoschizomer of AcyI. B. bifidum can be easily
cultured in large quantities. AcyI has been extracted from algal bodies and has been difficult
to prepare industrially. The obtained enzyme is much stabler than AcyI. B. bifidum
YIT-4005(FERM-P3372) and YIT-4007(FERM-P5871) are used. The culture is preferably at 25-45
(36-38) degrees C at pH 5-7 (6-7). After the culture, the bacterial body is preferably treated
with lysozyme and smashed with supersonic waves. Noncellular extract is obtained by
centrifuging the crushed product and refined by applying known methods.

<>

<1>Sakurai, T., Kosaka, T.
<2>Restriction enzyme manufacture by culturing Bifidobacterium and an enzyme recognising a specific arrangement in double-stranded DNA.
<3>Japanese Patent Office
<4>JP 5863387
<5>
<6>1983
<7>The method comprises culturing a bacterial strain of Bifidobacterium bifidum which produces a
restriction enzyme which can recognise the base arrangement GRCGYC in double-stranded DNA. The
restriction enzyme is then collected from the bacteria. Preferably the restriction enzyme
(abbr. BbiII) produced by B. bifidum is the isoschizomer of AcyI. B. bifidum can be easily
cultured in large quantities. AcyI has been extracted from algal bodies and has been difficult
to prepare industrially. The obtained enzyme is much stabler than AcyI. B. bifidum
YIT-4005(FERM-P3372) and YIT-4007(FERM-P5871) are used. The culture is preferably at 25-45
(36-38) degrees C at pH 5-7 (6-7). After the culture, the bacterial body is preferably treated
with lysozyme and smashed with supersonic waves. Noncellular extract is obtained by
centrifuging the crushed product and refined by applying known methods.

<>

<1>Salaj-Smic, E., Donjerkovic, D., Trgovcevic, Z.
<2>In vivo modulation of the EcoK restriction endonuclease.
<3>Periodicum Biologorum
<4>96
<5>357-358
<6>1994
<7>Recently, we have found a novel type of restriction alleviation (RA) which occurs in strains
carrying recB alone or recB in combination with additional mutations (recBC sbcBC, recBC sbcBC
recJ, recBC sbcBC recF, recBC sucBC recA).  It occurs in the absence of UV irraditation and
depends on the presence of the commonly used plasmids (such as pACYC184).  It is, however,
independent of the host and phage recombination systems.  The mechanism of this type of RA is
unknown.  Therefore, it was interesting to see whether UV-induced RA and plasmid-mediated RA
have a common mechanism.  For this purpose, we measured the efficiency of plating of
unmodified lambda on three sets of UV-irradiated bacteria: 1. E. coli JC7623 recB21 recC22
sbcB15 sbcC201 carrying the plasmid pACYC184 and, as a control, plasmid-free derivative; 2. E.
coli N2281 rec B21 recC22 sbcB15 sbcC201 recA13 carrying pACYC184 and its plasmid-free
derivative.  3. E. coli AB1157 (wild type) carrying pACYC184 and its plasmid-free derivative.

<>

<1>Salaj-Smic, E., Marsic, N., Trgovcevic, Z., Lloyd, R.G.
<2>Modulation of EcoKI restriction in vivo: Role of the lambda gam protein and plasmid metabolism.
<3>J. Bacteriol.
<4>179
<5>1852-1856
<6>1997
<7>Two novel types of alleviation of DNA restriction by the EcoKI restriction endonuclease are
described.  The first type depends on the presence of the gam gene product (Gam protein) of
bacteriophage lambda.  The efficiency of plating of unmodified phage lambda is greatly
increased when the restricting Escherichia coli K-12 host carries a gam+ plasmid.  The effect
is particularly striking in wild-type strains and, to a lesser extent, in the presence of sbcC
and recA mutations.  In all cases, Gam-dependent alleviation of restriction requires active
recBCD genes of the host and recombination (red) genes of the infecting phage.  The enhanced
capacity of Gam-expressing cells to repair DNA strand breaks might account for this
phenomenon.  The second type is caused by the presence of a plasmid in a restricting host
lacking RecBCD enzyme.  Commonly used plasmids such as the cloning vector pACYC184  can
produce such an effect in strains carrying recB single mutations or in recBC sbcBC strains.
Plasmid-mediated restriction alleviation in recBC sbcBC strains is independent of the host
RecF, RecJ, and RecA proteins and phage recombination functions.  The presence of plasmids can
also relieve restriction in recD strains.  This effect depends, however, on the RecA function
in the host.  The molecular mechanism of the plasmid-mediated restriction alleviation remains
unclear.

<>

<1>Salama, N., Guillemin, K., McDaniel, T.K., Sherlock, G., Tompkins, L., Falkow, S.
<2>A whole-genome microarray reveals genetic diversity among Helicobacter pylori strains.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>14668-14673
<6>2000
<7>Helicobacter pylori colonizes the stomach of half of the world's population, causing a wide
spectrum of disease ranging from
asymptomatic gastritis to ulcers to gastric cancer. Although the basis
for these diverse clinical outcomes is not understood, more severe
disease is associated with strains harboring a pathogenicity island. To
characterize the genetic diversity of more and less virulent strains,
we examined the genomic content of 15 H. pylori clinical isolates by
using a whole genome H. pylori DNA microarray. We found that a full 22%
of H. pylori genes are dispensable in one or more strains, thus
defining a minimal functional core of 1281 H. pylori genes. While the
core genes encode most metabolic and cellular processes, the
strain-specific genes include genes unique to H. pylori, restriction
modification genes, transposases, and genes encoding cell surface
proteins, which may aid the bacteria under specific circumstances
during their long-term infection of genetically diverse hosts. We
observed distinct patterns of the strain-specific gene distribution
along the chromosome, which may result from different mechanisms of
gene acquisition and loss. Among the strain-specific genes, we have
found a class of candidate virulence genes identified by their
coinheritance with the pathogenicity island.

<>

<1>Salama, N.R., Gonzalez-Valencia, G., Deatherage, B., Aviles-Jimenez, F., Atherton, J.C., Graham, D.Y., Torres, J.
<2>Genetic analysis of Helicobacter pylori strain populations colonizing the stomach at different times postinfection.
<3>J. Bacteriol.
<4>189
<5>3834-3845
<6>2007
<7>Genetic diversity of the human gastric pathogen Helicobacter pylori in an
individual host has been observed; whether this diversity represents
diversification of a founding strain or a mixed infection with distinct
strain populations is not clear. To examine this issue, we analyzed
multiple single-colony isolates from two to four separate stomach biopsies
of eight adult and four pediatric patients from a high-incidence Mexican
population. Eleven of the 12 patients contained isolates with identical
random amplified polymorphic DNA, amplified fragment length polymorphism,
and vacA allele molecular footprints, whereas a single adult patient had
two distinct profiles. Comparative genomic hybridization using
whole-genome microarrays (array CGH) revealed variation in 24 to 67 genes
in isolates from patients with similar molecular footprints. The one
patient with distinct profiles contained two strain populations differing
at 113 gene loci, including the cag pathogenicity island virulence genes.
The two strain populations in this single host had different spatial
distributions in the stomach and exhibited very limited genetic exchange.
The total genetic divergence and pairwise genetic divergence between
isolates from adults and isolates from children were not statistically
different. We also analyzed isolates obtained 15 and 90 days after
experimental infection of humans and found no evidence of genetic
divergence, indicating that transmission to a new host does not induce
rapid genetic changes in the bacterial population in the human stomach.
Our data suggest that humans are infected with a population of closely
related strains that vary at a small number of gene loci, that this
population of strains may already be present when an infection is
acquired, and that even during superinfection genetic exchange among
distinct strains is rare.

<>

<1>Salanoubat, M. et al.
<2>Genome sequence of the plant pathogen Ralstonia solanacearum.
<3>Nature
<4>415
<5>497-502
<6>2002
<7>Ralstonia solanacearum is a devastating, soil-borne plant pathogen with a global distribution
and an unusually wide host range. It is a model system for the dissection of molecular
determinants governing pathogenicity. We present here the complete genome sequence and its
analysis of strain GMI1000. The 5.8-megabase (Mb) genome is organized into two replicons: a
3.7-Mb chromosome and a 2.1-Mb megaplasmid. Both replicons have a mosaic structure providing
evidence for the acquisition of genes through horizontal gene transfer. Regions containing
genetically mobile elements associated with the percentage of G+C bias may have an important
function in genome evolution. The genome encodes many proteins potentially associated with a
role in pathogenicity. In particular, many putative attachment factors were identified. The
complete repertoire of type III secreted effector proteins can be studied. Over 40 candidates
were identified. Comparison with other genomes suggests that bacterial plant pathogens and
animal pathogens harbour distinct arrays of specialized type III-dependent effectors.

<>

<1>Salauen, L., Linz, B., Suerbaum, S., Saunders, N.J.
<2>The diversity within an expanded and redefined repertoire of phase-variable genes in Helicobacter pylori.
<3>Microbiology
<4>150
<5>817-830
<6>2004
<7>Phase variation is a common mechanism used by pathogenic bacteria to generate intra-strain
diversity that is important in niche adaptation and is strongly associated with virulence
determinants.  Previous analyses of the complete sequences of the Helicobacter pylori strains
26695 and J99 have identified 36 putative phase-variable genes among the two genomes through
their association with homopolymeric tracts and dinucleotide repeats.  Here a comparative
analysis of the two genomes is reported and an updated and expanded list of 46 candidate
phase-variable genes in H. pylori is described.  These have been systematically investigated
by PCR and sequencing for the presence of the genes, and the presence and variability in
length of the repeats in strains 26695 and J99 and in a collection of unrelated H. pylori
strains representative of the main global subdivisions recently suggested.  This provides
supportive evidence for the phase variability of 30 of the 46 candidates.  Other differences
in this subset of genes were observed (i) in the repeats, which can be present or absent among
the strains, or stabilized in different strains and (ii) in the gene-complements of the
strains.  Differences between genes were not consistently correlated with the geographic
population distribution of the strains.  This study extends and provides new evidence for
variation of this type in H. pylori, and of the high degree of diversity of the repertoire of
genes which display phase-variable switching within individual strains.

<>

<1>Saldanha, R., Chen, B., Wank, H., Matsuura, M., Edwards, J., Lambowitz, A.M.
<2>RNA and protein catalysis in group II intron splicing and mobility reactions using purified components.
<3>Biochemistry
<4>38
<5>9069-9083
<6>1999
<7>Group II introns encode proteins with reverse transcriptase activity. These proteins also
promote RNA splicing (maturase activity) and then, with the excised intron, form a
site-specific DNA endonuclease that promotes intron mobility by reverse splicing into DNA
followed by target DNA-primed reverse transcription. Here, we used an Escherichia coli
expression system for the Lactococcus lactis group II intron Ll.LtrB to show that the
intron-encoded protein (LtrA) alone is sufficient for maturase activity, and that RNP
particles containing only the LtrA protein and excised intron RNA have site-specific DNA
endonuclease and target DNA-primed reverse transcriptase activity. Detailed analysis of the
splicing reaction indicates that LtrA is an intron-specific splicing factor that binds to
unspliced precursor RNA with a K(d) of </=0.12 pM at 30 degrees C. This binding occurs in a
rapid bimolecular reaction, which is followed by a slower step, presumably an RNA
conformational change, required for splicing to occur. Our results constitute the first
biochemical analysis of protein-dependent splicing of a group II intron and demonstrate that a
single intron-encoded protein can interact with the intron RNA to carry out a coordinated
series of reactions leading to splicing and mobility.

<>

<1>Sales, C.M., Mahendra, S., Grostern, A., Parales, R.E., Goodwin, L., Woyke, T., Nolan, M., Lapidus, A., Chertkov, O., Ovchinnikova, G., Szcyrba, A., Alvarez-Cohen, L.
<2>Genome sequence of 1,4-dioxane degrading Pseudonocardia dioxanivorans strain CB1190.
<3>J. Bacteriol.
<4>193
<5>4549-4550
<6>2011
<7>Pseudonocardia dioxanivorans CB1190 is the first bacterium reported to be capable of growth on
the environmental contaminant 1,4-dioxane, and the
first member of the genus Pseudonocardia for which there is an annotated
genome sequence. Preliminary analysis of the genome (chromosome and three
plasmids) indicates that strain CB1190 possesses several multicomponent
monooxygenases that could be involved in the aerobic degradation of
1,4-dioxane and other environmental contaminants.

<>

<1>Salka, I., Srivastava, A., Allgaier, M., Grossart, H.P.
<2>The Draft Genome Sequence of Sphingomonas sp. Strain FukuSWIS1, Obtained from Acidic Lake Grosse Fuchskuhle, Indicates Photoheterotrophy and a Potential for  Humic Matter Degradation.
<3>Genome Announcements
<4>2
<5>e01183-14
<6>2014
<7>Sphingomonas spp. are Alphaproteobacteria considered to be versatile bacteria that can utilize
a variety of natural substrates available in terrestrial and
aquatic systems. Sphingomonas sp. strain FukuSWIS1 was isolated from the
eutrophic and acidic freshwater Lake Grosse Fuchskuhle in northeastern Germany.
The strain has a genome size of 3.89 Mb, possesses a set of photosynthetic genes,
and expresses photopigment BChl a under oxic conditions. Thus, this strain
belongs to the aerobic anoxygenic phototrophic (AAP) bacteria, which are most
likely involved in humic matter degradation as indicated by the presence of
organic compound mineralizing genes.

<>

<1>Salwan, R., Swarnkar, M.K., Singh, A.K., Kasana, R.C.
<2>First draft genome sequence of a member of the genus planomicrobium, isolated from the chandra river, India.
<3>Genome Announcements
<4>2
<5>e01259-13
<6>2014
<7>We report the first draft genome sequence of a member of the genus Planomicrobium, isolated
from a soil sample from the Chandra River, located in
the cold deserts of Himachal Pradesh, India. The draft genome assembly for
Planomicrobium glaciei strain CHR43 has a size of 3,900,800 bp with a G+C content
of 46.97%.

<>

<1>Salzberg, S.L. et al.
<2>Genome sequence and rapid evolution of the rice pathogen Xanthomonas oryzae pv. oryzae PXO99A.
<3>BMC Genomics
<4>9
<5>204
<6>2008
<7>BACKGROUND: Xanthomonas oryzae pv. oryzae causes bacterial blight of rice (Oryza sativa L.), a
major disease that constrains production of this
staple crop in many parts of the world. We report here on the complete
genome sequence of strain PXO99A and its comparison to two previously
sequenced strains, KACC10331 and MAFF311018, which are highly similar to
one another. RESULTS: The PXO99A genome is a single circular chromosome of
5,240,075 bp, considerably longer than the genomes of the other strains
(4,941,439 bp and 4,940,217 bp, respectively), and it contains 5083
protein-coding genes, including 87 not found in KACC10331 or MAFF311018.
PXO99A contains a greater number of virulence-associated transcription
activator-like effector genes and has at least ten major chromosomal
rearrangements relative to KACC10331 and MAFF311018. PXO99A contains
numerous copies of diverse insertion sequence elements, members of which
are associated with 7 out of 10 of the major rearrangements. A
rapidly-evolving CRISPR (clustered regularly interspersed short
palindromic repeats) region contains evidence of dozens of phage
infections unique to the PXO99A lineage. PXO99A also contains a unique,
near-perfect tandem repeat of 212 kilobases close to the replication
terminus. CONCLUSION: Our results provide striking evidence of genome
plasticity and rapid evolution within Xanthomonas oryzae pv. oryzae. The
comparisons point to sources of genomic variation and candidates for
strain-specific adaptations of this pathogen that help to explain the
extraordinary diversity of Xanthomonas oryzae pv. oryzae genotypes and
races that have been isolated from around the world.

<>

<1>Sam, M.D., Horton, N.C., Nissan, T.A., Perona, J.J.
<2>Catalytic efficiency and sequence selectivity of a restriction endonuclease modulated by a distal manganese ion binding site.
<3>J. Mol. Biol.
<4>306
<5>851-861
<6>2001
<7>Crystal structures of EcoRV endonuclease bound in a ternary complex with cognate duplex DNA
and manganese ions have previously revealed an Mn2+-binding site located between the enzyme
and the DNA outside of the dyad-symmetric GATATC recognition sequence. In each of the two
enzyme subunits, this metal ion bridges between a distal phosphate group of the DNA and the
imidazole ring of His71. The new metal-binding site is specific to Mn2+ and is not occupied in
ternary cocrystal structures with either Mg2+ or Ca2+. Characterization of the H71A and H71Q
mutants of EcoRV now demonstrates that these distal Mn2+ sites significantly modulate activity
toward both cognate and non-cognate DNA substrates. Single-turnover and steady-state kinetic
analyses show that removal of the distal site in the mutant enzymes increases Mn2+-dependent
cleavage rates of specific substrates by tenfold. Conversely, the enhancement of non-cognate
cleavage at GTTATC sequences by Mn2+ is significantly attenuated in the mutants. As a
consequence, under Mn2+ conditions Eco RV-H71A and EcoRV-H71Q are 100 to 700-fold more
specific than the wild-type enzyme for cognate DNA relative to the GTTATC non-cognate site.
These data reveal a strong dependence of DNA cleavage efficiency upon metal ion-mediated
interactions located some 20 ? distant from the scissile phosphodiester linkages.  They also
show that discrimination of cognate versus non-cognate DNA sequences by EcoRV depends in part
on contacts with the sugar-phosphate backbone outside of the target site.

<>

<1>Sam, M.D., Perona, J.J.
<2>Mn2+-dependent catalysis by restriction enzymes: Pre-steady-state analysis of EcoRV endonuclease reveals burst kinetics and the origins of reduced activity.
<3>J. Am. Chem. Soc.
<4>121
<5>1444-1447
<6>1999
<7>The origins of divalent metal-dependent catalytic properties in phosphoryl transfer by EcoRV
endonuclease have been investigated by transient kinetic methods.  Pre-steady-state
measurements on short oligodeoxynucleotide substrates reveal a burst of product formation for
both Mg2+- and Mn2+-catalyzed DNA cleavage reactions, indicating that for each metal ion the
product release step is partially or completely rate-limiting.  However, the steepness of the
burst is far greater for Mn2+ reactions, and analysis of the steady-state portions of the
reaction profiles shows that the overall rate is 6-fold slower in the presence of this
cofactor.  The strongly rate-limiting product release step in Mn2+ reactions may arise from
the higher intrinsic affinity of this metal ion for phosphates.  Single-turnover experiments
carried out with enzyme in molar excess over DNA were also used to isolate the chemical step
of the reaction.  In contrast to the slower steady-state rates, both these measurements and
the pre-steady-state reaction bursts show that the bond-breaking and bond-making steps are
significantly better catalyzed by Mn2+.  This supports models for catalysis deduced from X-ray
crystal structures of the enzyme-substrate DNA complex, in which a divalent metal ion is
directly ligated to the pro-Sp oxygen of the scissile group.

<>

<1>Sam, M.D., Perona, J.J.
<2>Catalytic roles of divalent metal ions in phosphoryl transfer by EcoRV endonuclease.
<3>Biochemistry
<4>38
<5>6576-6586
<6>1999
<7>The rate constant for the phosphoryl transfer step in site-specific DNA cleavage by EcoRV
endonuclease has been determined as a function of pH and identity of the required divalent
metal ion cofactor, for both wild-type and T93A mutant enzymes. These measurements show
bell-shaped pH-rate curves for each enzyme in the presence of Mg2+ as a cofactor, indicating
general base catalysis for the nucleophilic attack of hydroxide ion on the scissile phosphate,
and general acid catalysis for protonation of the leaving 3'-O anion. The kinetic data
support a model for phosphoryl transfer based on wild-type and T93A cocrystal structures, in
which the ionizations of two distinct metal-ligated waters respectively generate the attacking
hydroxide ion and the proton for donation to the leaving group. The model concurs with recent
observations of two metal ions bound in the active sites of the type II restriction
endonucleases BamHI and BglI, suggesting the possibility of a similar catalytic mechanism
functioning in many or all members of this enzyme family.

<>

<1>Samko, O.T., Kalugin, A.A., Eldarov, M.A., Karpychev, I.V., Anikeitcheva, N.V., Khoroshutina, E.B., Skryabin, K.G., Librik, G.I., Sokolov, N.N.
<2>Isolation, purification and characterization of new site-specific endonucleases BciBI and BciBII produced by Bacillus circulans.
<3>Bioorg. Khim.
<4>20
<5>1327-1333
<6>1994
<7>New site-specific endonucleases BciBI and BciBII have been detected in Bacillus circulans. The
enzymes were purified by fractionation of cell-free extract with polyethylene imine and
ammonium sulphate (40-80% of saturation) followed by chromatography on DEAE-sepharose,
blue-sepharose and phosphocellulose. The endonucleases BciBI and BciBII were separated only at
the final step of the purification -- by chromatography on the phosphocellulose column. The
yields of BciBI and BciBII were 600 and 10,000 U/g of cells. It was found that restriction
endonucleases BciBI and BciBII are isoschizomers of ClaI and BstNI, respectively.

<>

<1>Sampaio, J.L., Ribeiro, V.B., Campos, J.C., Rozales, F.P., Magagnin, C.M., Falci, D.R., da Silva, R.C., Dalarosa, M.G., Luz, D.I., Vieira, F.J., Antochevis, L.C., Barth, A.L., Zavascki, A.P.
<2>Detection of OXA-370, an OXA-48-Related Class D beta-Lactamase, in Enterobacter hormaechei from Brazil.
<3>Antimicrob. Agents Chemother.
<4>58
<5>3566-3567
<6>2014
<7>The class D -lactamase OXA-48 (1) and its variants have
emerged as important determinants of carbapenem resistance
in Enterobacteriaceae, representing a public health concern in
some countries (2-4). The objective of this study was to report a
new OXA-48 variant.

<>

<1>Sampath, J., Vijayakumar, M.N.
<2>A DNA cytosine methyltransferase-like gene in Streptococcal conjugative transposon, Tn5252.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>95
<5>511
<6>1995
<7>Tn5252, a 47 kb element, belongs to a distinct class of conjugative transposons mediating
multiple antibiotic resistance in Streptococci.  To determine the physical structure of the
element, fragments of transposon DNA were cloned into Escherichia coli.  However, a 3.27 kb
EcoRI segment of DNA carrying the right junction region could not be cloned on several of the
commonly used high copy E. coli plasmid vectors, even though portions of this DNA segment
could be independently cloned.  Using these the DNA sequence of this region was obtained and
was found to carry an open reading frame (ORF6).  Genbank analysis of the predicted amino acid
sequence of ORF6 detected extensive homology to a number of prokaryotic cytosine
methyltransferases.  To investigate the possible role of the cytosine methyltransferase-like
gene in the conjugative transposition of Tn5252, insertion mutagenesis of this locus was
carried out using the E. coli plasmid, pVA891.  Blot hybridization experiments confirmed the
intended insertion mutation.  The newly created strain would be used as a donor in
filter-mating experiments to determine the transferability of the element carrying the
mutation.  Also we have cloned the 3.27 kb EcoRI fragment carrying the ORF6 in a Streptococcal
plasmid, pLS1, which would be used in complementation experiments.

<>

<1>Sampath, J., Vijayakumar, M.N.
<2>Identification of a DNA cytosine methyltransferase gene in conjugative transposon Tn5252.
<3>Plasmid
<4>39
<5>63-76
<6>1998
<7>The nucleotide sequence of the 3.5-kb right junction fragment of the streptococcal conjugative
transposon Tn5252 was obtained.  The DNA fragment was found to carry four putative genes one
of which displayed a high degree of similarity to prokaryotic 5C-cytosine methyltransferases
carrying multiple sequence specificities.  No cognate endonuclease gene was detected in the
sequenced DNA.  Purified methylase polypeptide synthesized in a T7 promoter-controlled
overexpression system was found to lack methylase activity while the cell extracts of host
cells containing the recombinant plasmid carrying the methylase gene were active.  In vivo
mutations in the methylase gene did not seem to affect the transferability of the element.

<>

<1>Samrakandi, M.M., Cirillo, S.L.G., Ridenour, D.A., Bermudez, L.E., Cirillo, J.D.
<2>Genetic and phenotypic differences between Legionella pneumophila strains.
<3>J. Clin. Microbiol.
<4>40
<5>1352-1362
<6>2002
<7>Legionnaires' disease is a potentially lethal pneumonia that is primarily due to infection by
the species Legionella pneumophila, although more than 40 other species are known. Certain L.
pneumophila subgroups, particularly serogroup 1, are associated with the majority of the
epidemics. The genetic bases for these differences in virulence have not been determined.
Three strains, AA100, JR32, and Lp01, have been used in many molecular pathogenesis studies of
L. pneumophila. We found genetic differences between these strains by PCR and Southern
analyses that may be related to their ability to cause disease. We also examined the
distribution of these genetic loci in clinical and environmental isolates of Legionella and
found a correlation between the presence of two of these loci, rtxA and lvh, and the ability
to cause disease in humans. Examination of the interactions of these strains with host cells
suggested that they differ in important phenotypic characteristics including adherence, entry,
and intracellular replication. Furthermore, in the mouse model of infection they display
differing levels of replication in lungs. These studies emphasize the importance of further
investigation into the genetic makeup of these strains, which is likely to lead to the
identification of additional factors involved in Legionella pathogenesis.

<>

<1>Samson, J.E., Belanger, M., Moineau, S.
<2>Effect of the abortive infection mechanism and type III toxin/antitoxin system AbiQ on the lytic cycle of Lactococcus lactis phages.
<3>J. Bacteriol.
<4>195
<5>3947-3956
<6>2013
<7>To survive in phage-containing environments, bacteria have evolved an array of
anti-phage systems. Similarly, phages have overcome these hurdles through various
means. Here, we investigated how phages are able to circumvent the Lactococcus
lactis AbiQ system, a type III toxin-antitoxin with antiviral activities.
Lactococcal phage-escaping mutants were obtained in the laboratory and their
genome sequenced. Three unrelated genes of unknown function were mutated in
derivatives of three distinct lactococcal siphophages: orf38 of phage P008, m1 of
phage bIL170, and e19 of phage c2. One-step growth curve experiments revealed
that the phage mutations had a fitness cost while transcriptional analyses showed
that AbiQ modified the early-expressed phage mRNAs profile. The L. lactis AbiQ
system was also transferred into E. coli MG1655 and tested against several
coliphages. While AbiQ was efficient against phages T4 (Myoviridae) and T5
(Siphoviridae), escaping mutants of only phage 2 (Myoviridae) could be isolated.
Genome sequencing revealed a mutation in gene orf210, a putative DNA polymerase.
Taken altogether, different phages genes or genes products are targeted or
involved in AbiQ phenotype. Moreover, this antiviral system is active against
various phages families infecting Gram-positive and Gram-negative bacteria. A
model for the mode of action of AbiQ is proposed.

<>

<1>Samson, J.E., Magadan, A.H., Sabri, M., Moineau, S.
<2>Revenge of the phages: defeating bacterial defences.
<3>Nat. Rev. Microbiol.
<4>11
<5>675-687
<6>2013
<7>Bacteria and their viral predators (bacteriophages) are locked in a constant battle. In order
to proliferate in phage-rich environments, bacteria have an impressive arsenal of defence
mechanisms, and in response, phages have evolved counter-strategies to evade these antiviral
systems. In this Review, we describe the various tactics that are used by phages to overcome
bacterial resistance mechanisms, including adsorption inhibition, restriction-modification,
CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated
proteins) systems and abortive infection. Furthermore, we consider how these observations have
enhanced our knowledge of phage biology, evolution and phage-host interactions.

<>

<1>Samson, J.E., Moineau, S.
<2>Characterization of Lactococcus lactis phage 949 and comparison with other lactococcal phages.
<3>Appl. Environ. Microbiol.
<4>76
<5>6843-6852
<6>2010
<7>The virulent Lactococcus lactis phage 949 was isolated in 1975 from cheese
whey in New Zealand. This phage is a member of the Siphoviridae family and
of a rare lactococcal phage group that bears its name (949 group). It has
an icosahedral capsid (79-nm diameter) and a very long noncontractile tail
(length, 500 nm; width, 12 nm). It infected 7 of 59 tested L. lactis
strains, a somewhat expanded host range for a rare lactococcal phage. The
abortive phage infection defense mechanisms AbiQ and AbiT strongly
inhibited the multiplication of phage 949, but AbiK and AbiV did not. Its
double-stranded DNA (dsDNA) genome of 114,768 bp is, to date, the largest
among lactococcal phages. Its GC content was calculated at 32.7%, which is
the lowest reported for a lactococcal phage. Its 154 open reading frames
(ORFs) share limited identity with database sequences. In addition,
terminal redundancy was observed as well as the presence of six tRNAs, one
group I intron, and putative recombinases. SDS-PAGE coupled with mass
spectrometry identified 13 structural proteins. The genomes of the members
of the 10 currently known L. lactis phage groups were used to construct a
proteomic tree. Each L. lactis phage group separated into distinct genetic
clusters, validating the current classification scheme. Of note, members
of the polythetic P335 groups were clearly separated into subgroups.

<>

<1>Samuel, B.S., Hansen, E.E., Manchester, J.K., Coutinho, P.M., Henrissat, B., Fulton, R., Latreille, P., Kim, K., Wilson, R.K., Gordon, J.I.
<2>Genomic and metabolic adaptations of Methanobrevibacter smithii to the human gut.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>10643-10648
<6>2007
<7>The human gut is home to trillions of microbes, thousands of bacterial phylotypes, as well as
hydrogen-consuming methanogenic archaea. Studies in
gnotobiotic mice indicate that Methanobrevibacter smithii, the dominant
archaeon in the human gut ecosystem, affects the specificity and
efficiency of bacterial digestion of dietary polysaccharides, thereby
influencing host calorie harvest and adiposity. Metagenomic studies of the
gut microbial communities of genetically obese mice and their lean
littermates have shown that the former contain an enhanced representation
of genes involved in polysaccharide degradation, possess more archaea, and
exhibit a greater capacity to promote adiposity when transplanted into
germ-free recipients. These findings have led to the hypothesis that M.
smithii may be a therapeutic target for reducing energy harvest in obese
humans. To explore this possibility, we have sequenced its 1,853,160-bp
genome and compared it to other human gut-associated M. smithii strains
and other Archaea. We have also examined M. smithii's transcriptome and
metabolome in gnotobiotic mice that do or do not harbor Bacteroides
thetaiotaomicron, a prominent saccharolytic bacterial member of our gut
microbiota. Our results indicate that M. smithii is well equipped to
persist in the distal intestine through (i) production of surface glycans
resembling those found in the gut mucosa, (ii) regulated expression of
adhesin-like proteins, (iii) consumption of a variety of fermentation
products produced by saccharolytic bacteria, and (iv) effective
competition for nitrogenous nutrient pools. These findings provide a
framework for designing strategies to change the representation and/or
properties of M. smithii in the human gut microbiota.

<>

<1>Samuelson, J., Xu, S.-Y.
<2>Method for cloning and expression of Pseudomonas putida PpuMI restriction endonuclease and PpuMI methylase in Escherichia coli.
<3>US Patent Office
<4>US 20030215907 A
<5>
<6>2003
<7>The present invention relates to recombinant DNA encoding the PpuMI restriction endonuclease
as well as PpuMI methylase, expression of PpuMI restriction endonuclease and PpuMI methylase
in E. coli cells containing the recombinant DNA.

<>

<1>Samuelson, J., Xu, S.-y.
<2>Method for cloning and expression of PpuMI restriction endonuclease and PpuMI methylase in E. coli.
<3>US Patent Office
<4>US 6794172 B
<5>
<6>2004
<7>The present invention relates to recombinant DNA encoding the PpuMI restriction endonuclease
as well as PpuMI methylase, expression of PpuMI restriction endonuclease and PpuMI methylase
in E. coli cells containing the recombinant DNA.

<>

<1>Samuelson, J., Xu, S.-Y., O'Loane, D.
<2>Method for cloning and expression of Acinetobacter calcoaceticus AcuI restriction endonuclease and AcuI methylase in E. coli.
<3>US Patent Office
<4>US 20040209257 A
<5>27
<6>2004
<7>The present invention relates to recombinant DNA encoding the AcuI restriction endonuclease as
well as AcuI methylase, and expression of AcuI restriction endonuclease and AcuI methylase in
E. coli cells containing the recombinant DNA.

<>

<1>Samuelson, J., Xu, S.-Y., O'Loane, D.
<2>Method for cloning and expression of AcuI restriction endonuclease and AcuI methylase in E. coli.
<3>US Patent Office
<4>US 7011966 A
<5>
<6>2006
<7>The present invention relates to recombinant DNA encoding the AcuI restriction endonuclease as
well as AcuI methylase, and expression of AcuI restriction endonuclease and AcuI methylase in
E. coli cells containing the recombinant DNA.

<>

<1>Samuelson, J.C., Luo, J.
<2>Methods and compositions for targeting proteins or interest to the host cell envelope.
<3>International Patent Office
<4>WO 2009026089 A
<5>
<6>2009
<7>Methods and compositions are provided for producing membrane proteins or toxic proteins from
recombinant DNA introduced into a prokaryotic host cell by targeting the expressed proteins to
the envelope of the host cell.  The methods and compositions utilize a protein vehicle fused
to a  protein of interest.  The fusion protein may contain one or more protease cleavage sites
to separate the protein of interest from the protein vehicle either in vivo or in vitro.  The
protein vehicle is characterized by a membrane-targeting peptide and a trans-membrane segment
separated by a cytoplasmic amino acid sequence that includes a cytoplasmic affinity-binding
domain.

<>

<1>Samuelson, J.C., Morgan, R.D., Benner, J.S., Claus, T.E., Packard, S.L., Xu, S.Y.
<2>Engineering a rare-cutting restriction enzyme: genetic screening and selection of NotI variants.
<3>Nucleic Acids Res.
<4>34
<5>796-805
<6>2006
<7>Restriction endonucleases (REases) with 8-base specificity are rare specimens in nature. NotI
from Nocardia otitidis-caviarum (recognition
sequence 5'-GCGGCCGC-3') has been cloned, thus allowing for mutagenesis
and screening for enzymes with altered 8-base recognition and cleavage
activity. Variants possessing altered specificity have been isolated by
the application of two genetic methods. In step 1, variant E156K was
isolated by its ability to induce DNA-damage in an indicator strain
expressing M.EagI (to protect 5'-NCGGCCGN-3' sites). In step 2, the E156K
allele was mutagenized with the objective of increasing enzyme activity
towards the alternative substrate site: 5'-GCTGCCGC-3'. In this procedure,
clones of interest were selected by their ability to eliminate a
conditionally toxic substrate vector and induce the SOS response. Thus,
specific DNA cleavage was linked to cell survival. The secondary
substitutions M91V, F157C and V348M were each found to have a positive
effect on specific activity when paired with E156K. For example, variant
M91V/E156K cleaves 5'-GCTGCCGC-3' with a specific activity of 8.2 x 10(4)
U/mg, a 32-fold increase over variant E156K. A comprehensive analysis
indicates that the cleavage specificity of M91V/E156K is relaxed to a
small set of 8 bp substrates while retaining activity towards the NotI
sequence.

<>

<1>Samuelson, J.C., Xu, S.-Y.
<2>Directed evolution of restriction endonuclease BstYI to achieve increased substrate specificity.
<3>J. Mol. Biol.
<4>319
<5>673-683
<6>2002
<7>Restriction endonucleases have proven to be especially resistant to engineering altered
substrate specificity, in part, due to the requirement of a cognate DNA methyltransferase for
cellular DNA protection. The thermophilic restriction endonuclease BstYI recognizes and
cleaves all hexanucleotide sequences described by 5'-RGATCY-3' (where R=A or G and Y=C or
T). The recognition of a degenerate sequence is a relatively common feature of the more than
3000 characterized restriction endonucleases. However, very little is known concerning
substrate recognition by such an enzyme. Our objective was to investigate the substrate
specificity of BstYI by attempting to increase the specificity to recognition of only AGATCT.
By a novel genetic selection/screening process, two BstYI variants were isolated with a
preference for AGATCT cleavage. A fundamental element of the selection process is modification
of the Escherichia coli host genomic DNA by the BglII N4-cytosine methyltransferase to protect
AGATCT sites. The amino acid substitutions resulting in a partial change of specificity were
identified and combined into one superior variant designated NN1. BstYI variant NN1 displays a
12-fold preference for cleavage of AGATCT over AGATCC or GGATCT. Moreover, cleavage of the
GGATCC sequence is no longer detected. This study provides further evidence that laboratory
evolution strategies offer a powerful alternative to structure-guided protein design.

<>

<1>Samuelson, J.C., Xu, S.-Y.
<2>Alteration of restriction endonuclease specificity by genetic selection.
<3>International Patent Office
<4>WO 200360152
<5>51
<6>2003
<7>Methods and compositions are provided for altering the DNA recognition and cleavage
characteristics of an endonuclease without prior knowledge of the endonuclease's
three-dimensional structure and/or amino acid residues responsible for activity and/or
specificity.  Methods include subjecting a mutagenized endonucloease gene library to a genetic
selection in prokaryotic cells which tolerate the expression of mutated endonuclease and where
the endonuclease is active and determining the altered recognition-site specificity for the
endonuclease.

<>

<1>Samuelson, J.C., Xu, S.-y.
<2>Alteration of restriction endonuclease specificity by genetic selection.
<3>US Patent Office
<4>US 7052897 B
<5>
<6>2006
<7>Methods and compositions are provided for altering the DNA recognition and cleavage
characteristics of an endonuclease without prior knowledge of the endonuclease's
three-dimensional structure and/or amino acid residues responsible for activity and/or
specificity.  Methods include subjecting a mutagenized endonuclease gene library to a genetic
selection in prokaryotic cells which tolerate the expression of mutated endonuclease and where
the endonuclease is active and determining the altered recognition-site specificity for the
endonuclease.

<>

<1>Samuelson, J.C., Zhu, Z., Xu, S.-y.
<2>The isolation of strand-specific nicking endonucleases from a randomized SapI expression library.
<3>Nucleic Acids Res.
<4>32
<5>3661-3671
<6>2004
<7>The Type IIS restriction endonuclease SapI recognizes the DNA sequence 5'-GCTCTTC-3' (top
strand by convention) and cleaves downstream (N1/N4) indicating top- and bottom-strand
spacing, respectively. The asymmetric nature of DNA recognition presented the possibility that
one, if not two, nicking variants might be created from SapI. To explore this possibility, two
parallel selection procedures were designed to isolate either top-strand nicking or
bottom-strand nicking variants from a randomly mutated SapI expression library. These
procedures take advantage of a SapI substrate site designed into the expression plasmid, which
allows for in vitro selection of plasmid clones possessing a site-specific and strand-specific
nick. A procedure designed to isolate bottom-strand nicking enzymes yielded Nb.SapI-1
containing a critical R420I substitution near the end of the protein. The top-strand procedure
yielded several SapI variants with a distinct preference for top-strand cleavage. Mutations
present within the selected clones were segregated to confirm a top-strand nicking phenotype
for single variants Q240R, E250K, G271R or K273R. The nature of the amino acid substitutions
found in the selected variants provides evidence that SapI may possess two active sites per
monomer. This work presents a framework for establishing the mechanism of SapI DNA cleavage.

<>

<1>San Martin-Uriz, P., Gomez, M.J., Arcas, A., Bargiela, R., Amils, R.
<2>Draft Genome Sequence of the Electricigen Acidiphilium sp. Strain PM (DSM 24941).
<3>J. Bacteriol.
<4>193
<5>5585-5586
<6>2011
<7>Acidiphilium sp. strain PM (DSM 24941) was isolated from Rio Tinto's acidic, heavy metal-rich
waters. Voltammetry experiments revealed that
this strain is capable of electricity production even under aerobic
conditions. Here we report the draft genome sequence of Acidiphilium sp.
PM and a preliminary genome analysis that reveals a versatile respiratory
metabolism.

<>

<1>Sanchez, D.O., Zandomeni, R.O., Cravero, S., Verdun, R.E., Pierrou, E., Faccio, P., Diaz, G., Lanzavecchia, S., Aguero, F., Frasch, A.C.C., Andersson, S.G.E., Rosetti, O.L., Grau, O., Ugalde, R.A.
<2>Gene discovery through genomic sequencing of Brucella abortus.
<3>Infect. Immun.
<4>69
<5>865-868
<6>2001
<7>Brucella abortus is the etiological agent of brucellosis, a disease that
affects bovines and human. We generated DNA random sequences from the
genome of B. abortus strain 2308 in order to characterize molecular
targets that might be useful for developing immunological or
chemotherapeutic strategies against this pathogen. The partial sequencing
of 1,899 clones allowed the identification of 1,199 genomic sequence
surveys (GSSs) with high homology (BLAST expect value < 10(-5)) to
sequences deposited in the GenBank databases. Among them, 925 represent
putative novel genes for the Brucella genus. Out of 925 nonredundant GSSs,
470 were classified in 15 categories based on cellular function. Seven
hundred GSSs showed no significant database matches and remain available
for further studies in order to identify their function. A high number of
GSSs with homology to Agrobacterium tumefaciens and Rhizobium meliloti
proteins were observed, thus confirming their close phylogenetic
relationship. Among them, several GSSs showed high similarity with genes
related to nodule nitrogen fixation, synthesis of nod factors, nodulation
protein symbiotic plasmid, and nodule bacteroid differentiation. We have
also identified several B. abortus homologs of virulence and pathogenesis
genes from other pathogens, including a homolog to both the Shda gene from
Salmonella enterica serovar Typhimurium and the AidA-1 gene from
Escherichia coli. Other GSSs displayed significant homologies to genes
encoding components of the type III and type IV secretion machineries,
suggesting that Brucella might also have an active type III secretion
machinery.

<>

<1>Sanchez, J., Barbes, C., Hernandez, A., de los Reyes-Gavilan, C.G., Hardisson, C.
<2>Restriction-modification systems in Streptomyces antibioticus.
<3>Can. J. Microbiol.
<4>31
<5>942-946
<6>1985
<7>Several restriction systems were detected in different strains of Streptomyces
antibioticus by using actinophages as biological indicators. Adsorption of
phages to the bacteria, together with the study of the efficiency of plating
gave an initial indication of restriction in three strains. The alternation of
efficiency of plating values obtained from restricting and nonrestricting
hosts, detected among the different strains tested. Two specific endonucleases
with a possible role in restriction were detected in strains ATCC 11891 and ETH
7451, respectively.

<>

<1>Sanchez-Fernandez, R., Biesgen, C., Leps, M., Brown, J.A.
<2>Recombination cassettes and methods for sequence excision in plants.
<3>International Patent Office
<4>WO 2006032426 A
<5>
<6>2006
<7>The invention relates to improved recombination systems and methods for eliminating maker
sequences from the genome of plants.  Particularly the invention is based on use of an
expression cassette comprising the parsley ubiquitin promoter, and operably linked thereto a
nucleic acid sequence coding for a sequence specific DNA-endonuclease.

<>

<1>Sanchez-Porro, C., de la Haba, R.R., Cruz-Hernandez, N., Gonzalez, J.M., Reyes-Guirao, C., Navarro-Sampedro, L., Carballo, M., Ventosa, A.
<2>Draft Genome of the Marine Gammaproteobacterium Halomonas titanicae.
<3>Genome Announcements
<4>1
<5>e0008313
<6>2013
<7>Halomonas titanicae strain BH1 is a heterotrophic, aerobic marine bacterium which was isolated
from rusticles of the RMS Titanic wreck. Here we report the draft
genome sequence of this halophilic gammaproteobacterium.

<>

<1>Sanchez-Vizuete, P., Tanaka, K., Bridier, A., Shirae, Y., Yoshida, K., Bouchez, T., Aymerich, S., Briandet, R., Le Coq, D.
<2>Genome Sequences of Two Nondomesticated Bacillus subtilis Strains Able To Form Thick Biofilms on Submerged Surfaces.
<3>Genome Announcements
<4>2
<5>e00946-14
<6>2014
<7>Genomes of two nondomesticated strains of Bacillus subtilis subspecies subtilis,  NDmed and
NDfood, have been sequenced. Both strains form very thick and spatially
complex biofilms on submerged surfaces. Moreover, biofilms of the NDmed isolate
were shown to be highly resistant to antimicrobials action.

<>

<1>Sandal, I., Seleem, M.N., Elswaifi, S.F., Sriranganathan, N., Inzana, T.J.
<2>Construction of a high-efficiency shuttle vector for Histophilus somni.
<3>J. Microbiol. Methods
<4>74
<5>106-109
<6>2008
<7>The genetic manipulation of Histophilus somni is limited due to its high-fidelity
restriction-modification system. The broad host-range
shuttle plasmid pLS88 is capable of transforming some strains of H.
somni, but is an inefficient vector. We have constructed an improved
version of pLS88, pNS3K, that transforms H. somni strain 2336 100-fold
more efficiently than its predecessor. The transformation efficiency
was further increased when pNS3K was isolated from H. somni and
retransformed into the same strain. As proof of principle, the
lipooligosaccharide biosynthesis gene lob-2A was cloned into pNS3K and
expressed in H. somni strain 129Pt, which lacks this gene. Thus, pNS3K
is a useful shuttle vector for H. somni and a potential vector for
genetic manipulation of this bacterium.

<>

<1>Sandegren, L., Linkevicius, M., Lytsy, B., Melhus, A., Andersson, D.I.
<2>Transfer of an Escherichia coli ST131 multiresistance cassette has created a Klebsiella pneumoniae-specific plasmid associated with a major nosocomial outbreak.
<3>J. Antimicrob. Chemother.
<4>67
<5>74-83
<6>2012
<7>ObjectivesTo characterize the complete sequence, horizontal spread and
stability of the CTX-M-15-encoding multiresistance plasmid of a Klebsiella
pneumoniae strain involved in a large nosocomial outbreak.MethodsThe 220
kbp plasmid pUUH239.2 was completely sequenced using 454 technology. The
conjugational host range, conjugation frequencies, plasmid stability and
fitness cost of plasmid carriage were studied in vitro. Conjugational
spread during the outbreak was assessed retrospectively by multiplex PCR
screening, restriction fragment length polymorphism and
PFGE.ResultsPlasmid pUUH239.2 encodes resistance to beta-lactams
(bla(CTX-M-15), bla(TEM-1) and bla(OXA-1)), aminoglycosides
[aac-(6')-1b-cr and aadA2], tetracyclines [tet(A) and tetR], trimethoprim
(dhfrXII), sulphonamides (sul1), quaternary ammonium compounds
(qacEDelta1), macrolides [mph(A)-mxr-mphR(A)] and heavy metal ions
(silver, copper and arsenic). The plasmid consists of a backbone, highly
similar to the K. pneumoniae plasmid pKPN3, and a 41 kbp resistance
region, highly similar to the resistance regions of plasmids pEK499 and
pC15-1a previously isolated from Escherichia coli strains belonging to the
outbreak lineage ST131 (where ST stands for sequence type). The pUUH239.2
plasmid is stable in K. pneumoniae but unstable in E. coli and confers a
fitness cost when introduced into a naive host cell. Transfer of pUUH239.2
from the outbreak K. pneumoniae clone to the E. coli of the patients'
intestinal floras has occurred on multiple occasions during the
outbreak.ConclusionsThe plasmid pUUH239.2 is a composite of the pKPN3 K.
pneumoniae plasmid backbone and the bla(CTX-M-15)-encoding multiresistance
cassette associated with the internationally recognized outbreak strain E.
coli ST131. The resulting plasmid differs in stability between K.
pneumoniae and E. coli, and this has probably limited the spread of this
plasmid during the outbreak.

<>

<1>Sandegren, L., Nord, D., Sjoberg, B.M.
<2>SegH and Hef: two novel homing endonucleases whose genes replace the mobC and mobE genes in several T4-related phages.
<3>Nucleic Acids Res.
<4>33
<5>6203-6213
<6>2005
<7>T4 contains two groups of genes with similarity to homing endonucleases, the seg-genes ((s)
over bar imilarity to (e) over bar
ndonucleases encoded by (g) over bar roup I introns) containing GIY-YIG
motifs and the mob-genes (similarity to mobile endonucleases)
containing H-N-H motifs. The four seg-genes characterized to date
encode homing endonucleases with cleavage sites close to their
respective gene loci while none of the mob-genes have been shown to
cleave DNA. Of 18 phages screened, only T4 was found to have mobC while
mobE genes were found in five additional phages. Interestingly, three
phages encoded a seg-like gene (hereby called segH) with a GIY-YIG
motif in place of mobC. An additional phage has an unrelated gene
called hef ((h) over bar oming (e) over bar ndonuclease-like (f) over
bar unction) in place of the mobE gene. The gene products of both novel
genes displayed homing endonuclease activity with cleavage site
specificity close to their respective genes. In contrast to intron
encoded homing endonucleases, both SegH and Hef can cleave their own
DNA as well as DNA from phages without the genes. Both segH and mobE
(and most likely hef) can home between phages in mixed infections. We
discuss why it might be a selective advantage for phage freestanding
homing endonucleases to cleave both HEG-containing and HEG-less
genomes.

<>

<1>Sandegren, L., Sjoberg, B.M.
<2>Distribution, sequence homology, and homing of group I introns among T-even-like bacteriophages: evidence for recent transfer of old introns.
<3>J. Biol. Chem.
<4>279
<5>22218-22227
<6>2004
<7>Self-splicing group I introns are being found in an increasing number of bacteriophages. Most
introns contain an open reading frame coding for a homing endonuclease that confers mobility
to both the intron and the homing endonuclease gene (HEG). The frequent occurrence of
intron/HEG has raised questions whether group I introns are spread via horizontal transfer
between phage populations. We have determined complete sequences for the known group I introns
among T-even-like bacteriophages together with sequences of the intron-containing genes td,
nrdB, and nrdD from phages with and without introns. A previously uncharacterized phage
isolate, U5, is shown to contain all three introns, the only phage besides T4 found with a
"full set" of these introns. Sequence analysis of td and nrdB genes from intron-containing and
intronless phages provides evidence that recent horizontal transmission of introns has
occurred among the phages. The fact that several of the HEGs have suffered deletions rendering
them non-functional implies that the homing endonucleases are of no selective advantage to the
phage and are rapidly degenerating and probably dependent upon frequent horizontal
transmissions for maintenance within the phage populations. Several of the introns can home to
closely related intronless phages during mixed infections. However, the efficiency of homing
varies and is dependent on homology in regions flanking the intron insertion site. The
occurrence of optional genes flanking the respective intron-containing gene can strongly
affect the efficiency of homing. These findings give further insight into the mechanisms of
propagation and evolution of group I introns among the T-even-like bacteriophages.

<>

<1>Sanders, K.L., Catto, L.E., Bellamy, S.R., Halford, S.E.
<2>Targeting individual subunits of the FokI restriction endonuclease to specific DNA strands.
<3>Nucleic Acids Res.
<4>37
<5>2105-2115
<6>2009
<7>Many restriction endonucleases are dimers that act symmetrically at palindromic DNA sequences,
with each active site cutting one strand. In
contrast, FokI acts asymmetrically at a non-palindromic sequence, cutting
'top' and 'bottom' strands 9 and 13 nucleotides downstream of the site.
FokI is a monomeric protein with one active site and a single monomer
covers the entire recognition sequence. To cut both strands, the monomer
at the site recruits a second monomer from solution, but it is not yet
known which DNA strand is cut by the monomer bound to the site and which
by the recruited monomer. In this work, mutants of FokI were used to show
that the monomer bound to the site made the distal cut in the bottom
strand, whilst the recruited monomer made in parallel the proximal cut in
the top strand. Procedures were also established to direct FokI activity,
either preferentially to the bottom strand or exclusively to the top
strand. The latter extends the range of enzymes for nicking specified
strands at specific sequences, and may facilitate further applications of
FokI in gene targeting.

<>

<1>Sanders, M.E.
<2>Phage resistance in lactic acid bacteria.
<3>Biochimie
<4>70
<5>411-422
<6>1988
<7>The interactions between lactic acid bacteria and their phages are commercially
significant.  Current research has focused on the elucidation of the mechanisms
and genetics of phage resistance.  Phage resistance genes have been linked to
plasmid DNA for Streptococcus lactis and Streptococcus cremoris, and
preliminary studies suggest the operation of mechanisms such as the prevention
of phage adsorption, restriction/modification, and abortive infection.  Some
phage resistance plasmids can be conjugally transferred, providing a means of
dissemination among phage-sensitive strains for the construction of
phage-resistant starter cultures.

<>

<1>Sanders, M.E., Klaenhammer, T.R.
<2>Evidence for plasmid linkage of restriction and modification in Streptococcus cremoris KH.
<3>Appl. Environ. Microbiol.
<4>42
<5>944-950
<6>1981
<7>Restriction and modification have been demonstrated in Streptococcus cremoris
KH cells when infected by Streptococcus lactis C2 phage (designated c2) at an
efficiency of plating of 2 X 10-7.  The growth of c2 phage through KH cells
produces modified progeny phage capable of unrestricted growth on KH cells.
The ability of single-colony isolates of S. cremoris KH cultures to restrict
and modify c2 phage was found to be variable.  From 2 to 6.5% of colonies
isolated were partially deficient in restrictive capacity, permitting a greater
plaquing ability by c2 phage of 1.8 to 2.9 log cycles.  No completely
restrictionless mutants were isolated from 1,000 colonies examined.  Mutants
were shown to be deficient in both restriction and modification capabilities of
the same specificity.  The frequent occurrence of a genotypic change that
resulted in the loss of both restriction and modification capacities indicated
the involvement of plasmid deoxyribonucleic acid in genetically determining
this specific restriction and modification system.  S. cremoris KH was found to
harbor 11 plasmid molecules, with molecular weights (X10-6) estimated to be 50,
41, 24, 18, 10, 7.4, 3.3, 3.0, 2.8, 2.5, and 1.5.  Of the 27 mutants examined,
25 were missing the 10-megadalton plasmid.  This consistent plasmid difference
among the majority of mutants isolated supports the involvement of this plasmid
in restriction and modification.  Plasmid linkage of restriction and
modification systems provides a genetic mechanism for the rapid development of
phage-sensitive starter cultures due to the inherent instability of
extrachromosomal elements.

<>

<1>Sanders, M.E., Klaenhammer, T.R.
<2>Phage resistance in a phage-insensitive strain of Streptococcus lactis:  temperature-dependent phage development and host-controlled phage replication.
<3>Appl. Environ. Microbiol.
<4>47
<5>979-985
<6>1984
<7>Streptococcus lactis ME2 is a dairy starter strain that is insensitive to a
variety of phage, including Phi18.  The efficiency of plating of Phi18 on ME2
and N1 could be increased from <1 X 10-9 to 5.0 X 10-2 and from 7.6 X 10-7 to
2.1 X 10-2, respectively, when the host strains were subcultured at 40C before
plating the phage and the phage assay plates were incubated at 40C.
Host-dependent replication was demonstrated in N1 at 30C and in N1 and ME2 at
40C, suggesting the operation of a temperature-sensitive restriction and
modification system in MER2 and N1.  The increased sensitivity of ME2 and N1 to
Phi18 at 40C was also demonstrated by lysis of broth cultures and increased
plaque size.  ME2 grown at 40C showed an increased ability to adsorb Phi18,
indicating a second target for temperature-dependent phage sensitivity in ME2.
Challenge of N1 with a Phi18 preparation that had been previously modified for
growth on N1 indicated that at 40C phage development was characterized by a
shorter latent period and larger burst size than at 30C.  The evidence
presented suggests that the high degree of phage insensitivity expressed by ME2
consists of a variety of temperature-sensitive mechanisms, including (i) the
prevention of phage adsorption, (ii) host-controlled restriction of phage, and
(iii) suppression of phage development.  At 30C these factors appear to act
cooperatively to prevent the successful emergence of lytic phage active against
S. lactis ME2.

<>

<1>Sanders, M.E., Klaenhammer, T.R.
<2>Restriction and modification in group N streptococci: Effect of heat on development of modified lytic bacteriophage.
<3>Appl. Environ. Microbiol.
<4>40
<5>500-506
<6>1980
<7>The appearance of lytic bacteriophage against newly introduced starter strains used during
commercial cheese manufacture occurs rapidly, and their origin is not well understood.  In
this study, members of the group N streptococci were examined for the presence of
bacteriophage restriction and modification systems.  Two streptococcal phages from
Streptococcus cremoris TR and Streptococcus lactis C2 (phage designations tr and c2) showed
restricted lytic development on S. cremoris 799 and KH, respectively.  Efficiency of plaquing
was 1.9 x 10^-7 for tr plaqued on 799 and 2.1 x 10^-7 for c2 plaqued on KH.  After passage
through the restrictive hosts, these phages demonstrated high  lytic ability for formerly
restrictive hosts.  Stress of the restrictive host strains at temperatures of 40 to 50oC
resulted in a significant increase in the efficiency of plaquing of restricted bacteriophages.
Elevated temperatures are encountered during commercial cheese manufacture.  The results
suggested that the temporary loss of host restriction activity with the resulting modification
of nonspecific bacteriophage may contribute directly to the appearance of lytic phage against
new starter strains.

<>

<1>Sanders, M.E., Klaenhammer, T.R.
<2>Characterization of phage-sensitive mutants from a phage-insensitive strain of Streptococcus lactis: Evidence for a plasmid determinant that prevents phage adsorption.
<3>Appl. Environ. Microbiol.
<4>46
<5>1125-1133
<6>1983
<7>A phage-insensitive strain of Streptococcus lactis, designated ME2, was used as a prototype
strain for the study of mechanisms and genetics of phage resistance in the lactic
streptococci.  Mutants sensitive to a Streptococcus cremoris phage, Phi18, were isolated at a
level of 17% from cultures of ME2 after sequential transfer at 30oC.  Phage-sensitive mutants
of ME2 were not fully permissive to Phi18.  The efficiency of plating of Phi18 on the mutants
was 5 x 10^-7 as compared with <10^-0 for Phi18 on ME2.  Further characterization of the
mutants showed that they efficiently adsorbed Phi18 at levels of >99.8%, whereas ME2 adsorbed
only 20 to 40% of Phi18.  These results suggest that increased phage susceptibility of the
mutants may result from the loss of a mechanism that inhibits phage adsorption.  Moreover, the
high frequency of spontaneous mutation in ME2 indicates the involvement of an unstable genetic
determinant in this phage defense mechanism.  ME2 was shown to possess 13 plasmids ranging in
size from 1.6 to 34 megadaltons.  Of 40 mutants examined that had increased efficiencies of
plating, all were missing a 30-megadalton plasmid, pME0030.  These data suggest that pME0030
codes for a function that prevents phage adsorption.  Further phenotypic characterization of
the phage-sensitive mutants showed that some mutants were deficient in the ability to ferment
lactose and hydrolyze milk proteins.  However, the Lac+ and Prt+ phenotype segregated
independently of the phage-sensitivity phenotype.  One phage-sensitive adsorption mutant,
designated N1, was tested for susceptibility to 14 different phages.  N1 showed increased
capacity to adsorb 4 and to replicate 2 of these 14 phages, thereby indicating a phage
resistance mechanism in ME2 that generalizes to phage interactions other than the specific
o18-ME2 phage-host interaction.  These data provide evidence for a unique plasmid-linked phage
defense mechanism in phage-insensitive strains of lactic streptococci.

<>

<1>Sanders, M.E., Shultz, J.W.
<2>Cloning of phage resistance genes from Lactococcus lactis ssp. cremoris KH.
<3>J. Dairy Sci.
<4>73
<5>2044-2053
<6>1990
<7>A 17.5-kb plasmid conferring restriction and modification-type phage resistance
in Lactococcus lactis ssp. cremoris KH was transformed into Lactococcus lactis
LM0230 and cloned onto an origin of replication-deficient vector directly into
L. lactis.  Expression of phage resistance was seen in L. lactis but not in E.
coli.  A restriction map was generated of the cloned plasmid, and the region
encoding phage resistance was localized by insertional inactivation and
deletion analysis.

<>

<1>Sands, T.W., Petras, M.L., Van Wijngaarden, J.
<2>A computer program to assist in the choice of restriction endonucleases for use in DNA analysis.
<3>Int. J. Biomed. Comput.
<4>26
<5>39-52
<6>1990
<7>Type II restriction endonucleases cleave double stranded DNA molecules at sites characterized
by one or more sets of nucleotide pairs sequences. These digestions are essential in such
procedures as DNA cloning, DNA sequencing and restriction fragment length polymorphism (RFLP)
analyses. A large number of enzymes with different sequence specificities are available. To
date, most choices of restriction endonucleases have been made by trial and error. A computer
program, REDI, has been developed that predicts the ability of a particular restriction enzyme
to detect mutations. Characteristics of both the restriction endonuclease used and the DNA
being cut are incorporated as variables in the program. The program was tested using mouse
mitochondrial DNA (mtDNA) and bacteriophage lambda DNA because these have been sequenced and
are well characterized. REDI was strongly correlated (rs = +0.862, n = 11, P less than 0.001)
with mouse mtDNA RFLP detected by Ferris et al. [1] (Genetics, 105 (1983) 681-721). Even
though predictions may be altered by a non-random association of nucleotides, which varies
among DNA molecules, the predictions increase the probability of selecting the most efficient
enzymes for use in the analysis of a particular DNA molecule.

<>

<1>Sangal, V., Tucker, N.P., Burkovski, A., Hoskisson, P.A.
<2>The Draft Genome Sequence of Corynebacterium diphtheriae bv. mitis NCTC 3529 Reveals Significant Diversity between the Primary Disease-Causing Biovars.
<3>J. Bacteriol.
<4>194
<5>3269
<6>2012
<7>We report the draft genome of the human pathogen Corynebacterium diphtheriae bv.  mitis NCTC
3529. This is the first C. diphtheriae bv. mitis strain to be
sequenced and reveals significant differences from the other primary biovar, C.
diphtheriae bv. gravis.

<>

<1>Sangal, V., Tucker, N.P., Burkovski, A., Hoskisson, P.A.
<2>Draft Genome Sequence of Corynebacterium diphtheriae Biovar Intermedius NCTC 5011.
<3>J. Bacteriol.
<4>194
<5>4738
<6>2012
<7>We report an annotated draft genome of the human pathogen Corynebacterium diphtheriae bv.
intermedius NCTC 5011. This strain is the first C. diphtheriae
bv. intermedius strain to be sequenced, and our results provide a useful
comparison to the other primary disease-causing biovars, C. diphtheriae bv.
gravis and C. diphtheriae bv. mitis. The sequence has been deposited at
DDBJ/EMBL/GenBank with the accession number AJVH01000000.

<>

<1>Sanjar, F. et al.
<2>Genome Sequence of Escherichia coli O157:H7 Strain 2886-75, Associated with the First Reported Case of Human Infection in the United States.
<3>Genome Announcements
<4>2
<5>e01120-13
<6>2014
<7>First identified in 1982 as a human pathogen, enterohemorrhagic Escherichia coli  of the
O157:H7 serotype is a major cause of food-borne acquired human infections.
Here, we report the genome sequence of the first known strain of this serotype
isolated in the United States.

<>

<1>Sankpal, U.T., Rao, D.N.
<2>Mutational analysis of conserved residues in HhaI DNA methyltransferase.
<3>Nucleic Acids Res.
<4>30
<5>2628-2638
<6>2002
<7>HhaI DNA methyltransferase belongs to the C5-cytosine methyltransferase family, which is
characterized by the presence of a set of highly conserved amino acids and motifs present in
an invariant order. HhaI DNA methyltransferase has been subjected to a lot of biochemical and
crystallographic studies. A number of issues, especially the role of the conserved amino acids
in the methyltransferase activity, have not been addressed. Using sequence comparison and
structural data, a structure-guided mutagenesis approach was undertaken, to assess the role of
conserved amino acids in catalysis. Site-directed mutagenesis was performed on amino acids
involved in cofactor S-adenosyl-L-methionine (AdoMet) binding (Phe18, Trp41, Asp60 and
Leu100). Characterization of these mutants, by in vitro /in vivo restriction assays and
DNA/AdoMet binding studies, indicated that most of the residues present in the AdoMet-binding
pocket were not absolutely essential. This study implies plasticity in the recognition of
cofactor by HhaI DNA methyltransferase.

<>

<1>Sankpal, U.T., Rao, D.N.
<2>Structure, function, and mechanism of HhaI DNA methyltransferases.
<3>Crit. Rev. Biochem. Mol. Biol.
<4>37
<5>167-197
<6>2002
<7>A vast amount of literature has accumulated on the characterization of DNA methyltransferases.
The HhaI DNA methyltransferase, a C5-cytosine methyltransferase, has been the subject of
investigation for the last 2 decades. Biochemical and kinetic characterization have led to an
understanding of the catalytic and kinetic mechanism of the methyltransfer reaction. The HhaI
methyltransferase has also been subjected to extensive structural analysis, with the
availability of 12 structures with or without a cofactor and a variety of DNA substrates. The
mechanism of base flipping, first described for the HhaI methyltransferase, is conserved among
all DNA methyltransferases and is also found to occur in numerous DNA repair enzymes. Studies
with other methyltransferase reveal a significant structural and functional similarity among
different types of methyltransferases. This review aims to summarize the available information
on the HhaI DNA methyltransferase.

<>

<1>Sano, H., Koizumi, N., Yamaguchi, Y., Wada, N.
<2>Preparation of tobacco DNA methyltransferase and use of the enzyme for treatment of plant diseases.
<3>Japanese Patent Office
<4>JP 2004222581 A
<5>29
<6>2004
<7>
<>

<1>Sano, H., Sager, R.
<2>Deoxyribonucleic acid methyltransferase from the eukaryote, Chlamydomonas reinhardi.
<3>Eur. J. Biochem.
<4>105
<5>471-480
<6>1980
<7>DNA methyltransferase was purified 310-fold from a green alga. Chlamydomonas reinhardi
vegetative cells. The native enzyme of molecular weight 55000-58000 catalyzed the transfer of
methyl groups from S-adenosylmethionine to the 5 position of cytosine in DNA. Native DNA
accepted methyl groups 10-fold more than did denatured DNA. The sequence specificity analysis
of methylated deoxycytidine in vitro revealed that the enzyme introduces methyl groups
preferentially into sequences containing 5'd(T-mC-R)3'. Kinetic anlaysis of the reaction
indicated that the enzyme obeys a random sequential mechanism. The extent of saturation with
methyl groups depends upon the species from which the DNA was obtained. Kinetic analysis of
the reaction catalyzes by RNA polymerase II has indicated that DNA methylation decreases the
rate of initiation of RNA synthesis, but does not affect the rate of RNA chain elongation.

<>

<1>Sansom, C.E., Biro, J.C., Benyo, B., Benyo, Z.
<2>Codes in the codons: codon-amino acid complementarity revealed in the structures of restriction enzyme - DNA complexes.
<3>FEBS J.
<4>272
<5>100
<6>2005
<7>In the early years after the elucidation of the DNA structure, there was controversy about
whether there was any chemical rationale underlying the genetic code. In 1967, Carl Woese
proposed that there was stereochemical affinity between amino acids and the base sequences
that code for them. The alternative hypothesis proposed by Crick among others, that the exact
genetic code was largely accidental, became largely accepted by the molecular biology
community. We have now constructed a "periodic table" linking the chemical properties of the
amino acids and the sequences of their associated codons. The amino acid table showed
significant periodicity and indicated the importance of the central base in determining the
chemical properties of amino acids. This adds support to Woese' original hypothesis. If this
stereochemical and structural affinity were true, we would expect interactions between amino
acids and their associated codons to be favoured in DNA-protein complexes. We originally
tested this hypothesis using known structures of restriction enzyme - DNA complexes. We found
that, not only were cleavage-site like base sequences found disproportionately often in the
DNA sequences of restriction enzymes, but that, in the complex structures, the amino acids
coded by those site-like sequences were found close to the restriction sites themselves. The
average distance between the closest atoms in the codon and amino acid was significantly
lowest when the amino acid involved was positively charged. We now update this work to include
restriction enzyme - DNA complexes that have entered the PDB since December 2003.

<>

<1>Santagati, M., Iannelli, F., Cascone, C., Campanile, F., Oggioni, M.R., Stefani, S., Pozzi, G.
<2>The novel conjugative transposon tn1207.3 carries the macrolide efflux gene mef(A) in Streptococcus pyogenes.
<3>Microb. Drug Resist.
<4>9
<5>243-247
<6>2003
<7>The macrolide efflux gene mef(A) of the Streptococcus pyogenes clinical strain 2812A was found
to be carried by a 52-kb chromosomal genetic
element that could be transferred by conjugation to the chromosome of
other streptococcal species. The characteristics of this genetic element
are typical of conjugative transposons and was named Tn1207.3. The size of
Tn1207.3 was established by pulsed-field gel electrophoresis (PFGE), and
DNA sequencing analysis showed that the 7,244 bp at the left end of
Tn1207.3 were identical to those of the pneumococcal Tn1207.1 element.
Tn1207.3-like genetic elements were found to be inserted at a single
specific chromosomal site in 12 different clinical isolates S. pyogenes
exhibiting the M phenotype of resistance to macrolides and carrying the
mef(A) gene. Tn1207.3 was transferred from S. pyogenes 2812A to
Streptococcus pneumoniae, and sequence analysis carried out on six
independent transconjugants showed that insertion of Tn1207.3 in the
pneumococcal genome always occurred at a single specific site as in
Tn1207.1. Using MF2, a representative S. pneumoniae transconjugant, as a
donor, Tn1207.3 was transferred again by conjugation to S. pyogenes and
Streptococcus gordonii. The previously described nonconjugative element
Tn1207.1 of S. pneumoniae appears to be a defective element, part of a
longer conjugative transposon that carries mef(A) and is found in clinical
isolates of S. pyogenes.

<>

<1>Santhoshkumar, P., Sharma, K.K.
<2>Analysis of alpha-crystallin chaperone function using restriction enzymes and citrate synthase.
<3>Mol. Vis.
<4>7
<5>172-177
<6>2001
<7>PURPOSE: To compare the abilities of [alpha]A-crystallin, [alpha]B-crystallin, and
mini-[alpha]A-crystallin (a synthetic peptide
chaperone representing the functional unit of [alpha]A-crystallin) to
protect against heat-induced inactivation of citrate synthase (CS) and
restriction enzymes, SmaI and NdeI. METHODS: Restriction enzymes, SmaI and
NdeI were heated at different temperatures in the presence of various
amounts of molecular chaperones and tested for their ability to cleave
plasmid DNA. The aggregation of CS was measured at 43 degrees C while the
loss in activity was monitored at 37 degrees C in the presence of various
crystallins. RESULTS: Restriction enzyme activities were protected by the
crystallin subunits up to 37 degrees C for SmaI and 43 degrees C for NdeI.
However, the mini-[alpha]A-crystallin was unable to protect endonuclease
activity. The crystallin subunits and the peptide chaperone were able to
suppress thermal aggregation of CS at 43 degrees C, but failed to
stabilize its activity at 37 degrees C. CONCLUSIONS: The ability of
[alpha]-crystallin subunits to stabilize denaturing proteins varies from
enzyme to enzyme as evidenced by the inactivation of CS and protection of
SmaI and NdeI activity in the presence of [alpha]-crystallin subunits.
Additionally, our results show that there could be more than one site in
[alpha]A-crystallin responsible for its chaperone-like action. By addition
of crystallin subunits to restriction enzymes prior to or during storage,
transport, or assay would maintain or improve their activity thereby
decreasing their cost.

<>

<1>Santi, D.V., Garrett, C.E., Barr, P.J.
<2>On the mechanism of inhibition of DNA-cytosine methyltransferases by cytosine analogs.
<3>Cell
<4>33
<5>9-10
<6>1983
<7>In recent years 5-methylcytosine residues in DNA have been implicated to have an important
role in the control of eucaryotic gene expression (see Razin and Riggs, Science 210, 604-610,
1980). Consequently, there has been much interest in cytosine analogs such as 5-azacytosine
(5-azaC) and 5-fluorocytosine (5-fluoro-C), which, when incorporated into DNA, inhibit
methylation and profoundly affect gene expression and differentiation (see Taylor and Jones,
JBM 162, 679-692, 1982). The degree of hypomethylation far exceeds the levels of such analogs
in DNA and is not simply a result of their inability to serve as methyl acceptors. It is now
clear that DNAs containing low levels of these analogs are potent inhibitors of DNA-cytosine
methyltansferase (DCMT), but the mechanism of inhibition is unresolved. In this report, we
review pertinent literature and formulate a proposal for the molecular mechanism by which DCMT
is inhibited bly DNA containing 5-aza-C or 5-fluoro-C. A similar mechanism explains how small
amounts of 5-aza-C or 5-fluoro-C in tRNA cause specific loss of tRNA-cytosine
methyltransferase activity (Lu et al., Biochem. Pharmacol. 28, 489-495, 1979; Lu and
Randerath, Cancer Res. 40, 2701-2705, 1980).

<>

<1>Santi, D.V., Norment, A., Garrett, C.E.
<2>Covalent bond formation between a DNA-cytosine methyltransferase and DNA containing 5-azacytosine.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>81
<5>6993-6997
<6>1984
<7>DNA containing 5-azacytosine (azaC) has previously been shown to be a
potent inhibitor of DNA-cytosine methyltransferases.  In this report, we describe
experiments which demonstrate that azaC-DNA forms a covalent complex with HpaII
methylase, a bacterial enzyme that methylates the internal C of C-C-G-G sequences.  The
complex does not undergo detectable dissociation over at least 3 days and is stable to
denaturation with NaDodSO4.  After extensive digestion of the complex with DNase and
phosphodiesterase, gel filtration gave the methylase bound to approximately one equivalent
of azaC; the digested complex had an apparent molecular weight similar to that of the native
enzyme.  Although prior treatment of azaC-DNA with HpaII endonuclease had only a slight
effect on binding of the methylase, treatment with MspI endonuclease, which also cleaves
at C-C-G-G sequences, resulted in a significant reduction in binding; this indicates that
azaC residues in the recognition sequence of HpaII are an important component in the
covalent interaction of the methylase.  However, since there was residual binding it is
possible that azaC residues elsewhere in DNA also covalently bind to the methylase.  These
results provide an explanation of why azaC-DNA is such a potent inhibitor of cytosine
methyltransferases and how the incorporation of such low levels of azaC into DNA can
result in dramatic decreases in the methylation of cytosine.  Finally, consideration of the
probable catalytic mechanism of cytosine methylases and the chemical properties of azaC
suggests that the inhibition is, at least in part, an active-site directed process and permits
a
proposal for the structure of the covalent complex.

<>

<1>Santini, S., Jeudi, S., Bartoli, J., Poirot, O., Lescot-David, M., Barbe, V., Wommack, E.K., Abergel, C., Brussaard, C., Claverie, J.-M.
<2>The genome of Phaeocystis globosa virus PgV-16T highlights the common ancestry of the largest known DNA viruses infecting eukaryotes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>110
<5>10800-10805
<6>2013
<7>Large dsDNA viruses are involved in the population control of many
globally distributed species of eukaryotic phytoplankton and have
a prominent role in bloomtermination. The genus Phaeocystis (Haptophyta,
Prymnesiophyceae) includes several high-biomass-forming
phytoplankton species, such as Phaeocystis globosa, the blooms of
which occur mostly in the coastal zone of the North Atlantic and the
North Sea. Here,we report the 459,984-bp-long genomesequence of
P. globosa virus strain PgV-16T, encoding 434 proteins and eight
tRNAs and, thus, the largest fully sequenced genome to date among
viruses infecting algae. Surprisingly, PgV-16T exhibits no phylogenetic
affinity with other viruses infecting microalgae (e.g., phycodnaviruses),
including those infecting Emiliania huxleyi, another
ubiquitous bloom-forming haptophyte. Rather, PgV-16T belongs to
anemerging clade (theMegaviridae) clustering the viruses endowed
with the largest known genomes, including Megavirus, Mimivirus
(both infecting acanthamoeba), and a virus infecting the marine
microflagellate grazer Cafeteria roenbergensis. Seventy-five percent
of the best matches of PgV-16T-predicted proteins correspond to
two viruses [Organic Lake phycodnavirus (OLPV)1 and OLPV2] from
a hypersaline lake in Antarctica (Organic Lake), the hosts of which
are unknown. As for OLPVs and other Megaviridae, the PgV-16T
sequence data revealed the presence of a virophage-like genome.
However, no virophage particle was detected in infected P. globosa
cultures. The presence of many genes found only in Megaviridae in
its genome and the presence of an associated virophage strongly
suggest that PgV-16T shares a common ancestry with the largest
known dsDNA viruses, the host range ofwhich already encompasses
the earliest diverging branches of domain Eukarya.

<>

<1>Santopolo, L., Marchi, E., Decorosi, F., Galardini, M., Brilli, M., Giovannetti, L., Viti, C.
<2>Draft Genome Sequence of Chromate-Resistant and Biofilm-Producing Strain Pseudomonas alcaliphila 34.
<3>Genome Announcements
<4>1
<5>e00125-12
<6>2013
<7>We report the draft genome sequence of 34, a Cr(VI)-hyperresistant and biofilm-producing
bacterium that might be used for the bioremediation of
chromate-polluted soils. The genome sequence might be helpful in exploring the
mechanisms involved in chromium resistance and biofilm formation.

<>

<1>Santos, A.P., Guimaraes, A.M.S., do Nascimento, N.C., SanMiguel, P.J., Martin, S.W., Messick, J.B.
<2>Genome of Mycoplasma haemofelis, unraveling its strategies for survival and persistence.
<3>Vet. Res.
<4>42
<5>102
<6>2011
<7>Mycoplasma haemofelis is a mycoplasmal pathogen (hemoplasma) that attaches to the host's
erythrocytes. Distributed worldwide, it has a
significant impact on the health of cats causing acute disease and,
despite treatment, establishing chronic infection. It might also have a
role as a zoonotic agent, especially in immunocompromised patients.
Whole genome sequencing and analyses of M. haemofelis strain Ohio2 was
undertaken as a step toward understanding its survival and persistence.
Metabolic pathways are reduced, relying on the host to supply many of
the nutrients and metabolites needed for survival. M. haemofelis must
import glucose for ATP generation and ribose derivates for RNA/DNA
synthesis. Hypoxanthine, adenine, guanine, uracil and CMP are scavenged
from the environment to support purine and pyrimidine synthesis. In
addition, nicotinamide, amino acids and any vitamins needed for growth,
must be acquired from its environment. The core proteome of M.
haemofelis contains an abundance of paralogous gene families,
corresponding to 70.6% of all the CDSs. This 'paralog pool' is a rich
source of different antigenic epitopes that can be varied to elude the
host's immune system and establish chronic infection. M. haemofelis
also appears to be capable of phase variation, which is particularly
relevant to the cyclic bacteremia and persistence, characteristics of
the infection in the cat. The data generated herein should be of great
use for understanding the mechanisms of M. haemofelis infection.
Further, it will provide new insights into its pathogenicity and clues
needed to formulate media to support the in vitro cultivation of M.
haemofelis.

<>

<1>Santos, F., Yarza, P., Parro, V., Briones, C., Anton, J.
<2>The metavirome of a hypersaline environment.
<3>Environ. Microbiol.
<4>12
<5>2965-2976
<6>2010
<7>Hypersaline environments harbour the highest number of virus-like
particles reported for planktonic systems. However, very little is known
about the genomic diversity of these virus assemblages since most of the
knowledge on halophages is based on the analysis of a few isolates
infecting strains of hyperhalophilic Archaea that may not be
representatives of the natural microbiota. Here, we report the
characterization, through a metagenomic approach, of the viral assemblage
inhabiting a crystallizer pond (CR30) from a multi-pond solar saltern in
Santa Pola (SE Spain). A total of 1.35 Mbp were cloned that yielded a
total of 620 kb sequenced viral DNA. The metavirome was highly diverse and
different from virus communities of marine and other aquatic environments
although it showed some similarities with metaviromes from high-salt ponds
in solar salterns in San Diego (SW USA), indicating some common traits
between high-salt viromes. A high degree of diversity was found in the
halophages as revealed by the presence of 2479 polymorphic nucleotides.
Dinucleotide frequency analysis of the CR30 metavirome showed a good
correlation with GC content and enabled the establishment of different
groups, and even the assignment of their putative hosts: the archaeon
Haloquadratum walsbyi and the bacterium Salinibacter ruber.

<>

<1>Santos-Garcia, D., Farnier, P.A., Beitia, F., Zchori-Fein, E., Vavre, F., Mouton, L., Moya, A., Latorre, A., Silva, F.J.
<2>Complete Genome Sequence of 'Candidatus Portiera aleyrodidarum' BT-QVLC, an Obligate Symbiont That Supplies Amino Acids and Carotenoids to Bemisia tabaci.
<3>J. Bacteriol.
<4>194
<5>6654-6655
<6>2012
<7>The genome of 'Candidatus Portiera aleyrodidarum,' the primary endosymbiont of the whitefly
Bemisia tabaci (Mediterranean species), is reported. It presents a
reduced genome (357 kb) encoding the capability to synthetize, or participate in
the synthesis of, several amino acids and carotenoids, being the first insect
endosymbiont capable of supplying carotenoids.

<>

<1>Santourlidis, S., Schulz, W.A.
<2>The cytotoxic effect of DNA methyltransferase I over-expression is mediated by the N-terminal.
<3>Biochem. Soc. Trans.
<4>28
<5>A182
<6>2000
<7>DNA methyltransferase 1, the essential enzyme maintaining DNA methylation patterns in
mammalian cells, contains a 500 amino acid catalytic domain and an N-terminal 1200 amino acid
complex regulatory domain determining specificity and intracellular localization.
Overexpression of the enzyme in somatic cells is very difficult, even more of the
oocyte-specific form lacking the first 120 amino acids.  To elucidate the underlying causes,
the mouse and human cDNAs encoding this form were transfected into mouse F9 cells and various
human cell lines.  Few cell clones were obtained which contained rearranged cDNAs after two
weeks, but the number of transfected cells was not notably diminished up to 3 days.
Surprisingly, deletion of the catalytic domain did not obliterate cytotoxicity; rather a 150
amino acid subdomain containing the PCNA-binding site and an NLS was found to be sufficient.
Mutation of the PCNA-binding sequence abolished cytotoxicity.  Thus, DNMT1 overexpression
causes a slow-acting cytotoxic effect via binding to PCNA, possibly by interfering with DNA
replication.  DNMT1 may be a multifunctional enzyme important beyond DNA methylation.

<>

<1>Sapienza, P., Kurpiewski, M., Jen-Jacobson, L.
<2>Thermodynamic and kinetic basis of promiscuity in mutant EcoRI endonucleases.
<3>Biophys. J.
<4>84
<5>367a
<6>2003
<7>We have used fluorescence-quenching measurements to characterize the partitioning of a variety
of indolyl-labeled phospho- and sphingolipids
between gel or liquid-ordered and liquid-disordered lipid domains in
several types of lipid bilayers where such domains coexist. In both
cholesterol-free and cholesterol-containing lipid mixtures, sphingolipids
with diverse polar headgroups (ranging from sphingomyelin and
monoglycosylceramides to ganglioside GM1) show a net preference for
partitioning into ordered domains, which varies modestly in magnitude with
varying headgroup structure. The affinities of different sphingolipids for
ordered lipid domains do not vary in a consistent manner with the size or
other simple structural properties of the polar headgroup, such that for
example ganglioside GM1 partitions between ordered and disordered lipid
domains in a manner very similar to sphingomyelin. Ceramide exhibits a
dramatically higher affinity for ordered lipid domains in both
cholesterol-free and cholesterol-containing bilayers than do other
sphingolipids. Our findings suggest that sphingolipids with a variety of
headgroup structures will be enriched by substantial factors in
liquid-ordered versus liquid-disordered regions of membranes, in a manner
that is only modestly dependent on the nature of the polar headgroup.
Ceramide is predicted to show a very strong enrichment in such domains,
supporting previous suggestions that ceramide-mediated signaling may be
compartmentalized to liquid-ordered (raft and raft-related) domains in the
plasma membrane.

<>

<1>Sapienza, P.J., Dela-Torre, C.A., McCoy, W.H., Jana, S.V., Jen-Jacobson, L.
<2>Thermodynamic and kinetic basis for the relaxed DNA sequence specificity of "promiscuous" mutant EcoRI endonucleases.
<3>J. Mol. Biol.
<4>348
<5>307-324
<6>2005
<7>Promiscuous mutant EcoRI endonucleases produce lethal to sublethal effects because they cleave
Escherichia coli DNA despite the presence of the EcoRI methylase. Three promiscuous mutant
forms, Ala138Thr, Glu192Lys and His114Tyr, have been characterized with respect to their
binding affinities and first-order cleavage rate constants towards the three classes of DNA
sites: specific, miscognate (EcoRI*) and non-specific. We have made the unanticipated and
counterintuitive observations that the mutant restriction endonucleases that exhibit relaxed
specificity in vivo nevertheless bind more tightly than the wild-type enzyme to the specific
recognition sequence in vitro, and show even greater preference for binding to the cognate
GAATTC site over miscognate sites. Binding preference for EcoRI* over non-specific DNA is also
improved. The first-order cleavage rate constants of the mutant enzymes are normal for the
cognate site GAATTC, but are greater than those of the wild-type enzyme at EcoRI* sites. Thus,
the mutant enzymes use two mechanisms to partially bypass the multiple fail-safe mechanisms
that protect against cleavage of genomic DNA in cells carrying the wild-type EcoRI
restriction-modification system: (a) binding to EcoRI* sites is more probable than for
wild-type enzyme because non-specific DNA is less effective as a competitive inhibitor; (b)
the combination of increased affinity and elevated cleavage rate constants at EcoRI* sites
makes double-strand cleavage of these sites a more probable outcome than it is for the
wild-type enzyme. Semi-quantitative estimates of rates of EcoRI* site cleavage in vivo,
predicted using the binding and cleavage constants measured in vitro, are in accord with the
observed lethal phenotypes associated with the three mutations.

<>

<1>Sapienza, P.J., Rosenberg, J.M., Jen-Jacobson, L.
<2>Structural and Thermodynamic Basis for Enhanced DNA Binding by a Promiscuous Mutant EcoRI Endonuclease.
<3>Structure
<4>15
<5>1368-1382
<6>2007
<7>Promiscuous mutant EcoRI endonucleases bind to the canonical site GAATTC more tightly than
does the wild-type endonuclease, yet cleave variant
(EcoRI( *)) sites more rapidly than does wild-type. The crystal structure
of the A138T promiscuous mutant homodimer in complex with a GAATTC site is
nearly identical to that of the wild-type complex, except that the Thr138
side chains make packing interactions with bases in the 5'-flanking
regions outside the recognition hexanucleotide while excluding two bound
water molecules seen in the wild-type complex. Molecular dynamics
simulations confirm exclusion of these waters. The structure and
simulations suggest possible reasons why binding of the A138T protein to
the GAATTC site has DeltaS degrees more favorable and DeltaH degrees less
favorable than for wild-type endonuclease binding. The interactions of
Thr138 with flanking bases may permit A138T, unlike wild-type enzyme, to
form complexes with EcoRI( *) sites that structurally resemble the
specific wild-type complex with GAATTC.

<>

<1>Sapranauskas, R.
<2>Investigation of type IIS restriction endonuclease BfiI domain organization by using a new random gene dissection approach.
<3>Ph.D. Thesis, Vilnius University
<4>
<5>1-42
<6>2008
<7>Conclusions
1. The genes of BfiI restriction-modification system were cloned. The system
comprises two cytosine-N4-methyltransferases and a restriction endonuclease.
2. A new method for investigation of protein domain organization, Random
Gene Dissection, was proposed. The interdomain region of the chosen model protein, FokI REase,
was determined using the new method, and it was in perfect agreement with the linker region
predicted from the tertiary structure of FokI.
3. Interdomain region of BfiI REase was determined by Random Gene
Dissection method.
4. Indirect data (induction of SOS response, toxicity) indicate that the
N-terminal domain of BfiI acts as a nuclease.
5. The DNA binding assay using purified C-terminal domain of BfiI indicates
that it is responsible for target recognition.
6. The complementary fragments of FokI and BfiI were isolated using Random
Gene Dissection. Therefore the Random Gene Dissection method can be
used not only for identification of interdomain regions, but also for isolation of
complementing fragments of proteins.

<>

<1>Sapranauskas, R., Lubys, A.
<2>Random gene dissection: a tool for the investigation of protein structural organization.
<3>Biotechniques
<4>39
<5>395-402
<6>2005
<7>To investigate the domain structure of proteins and the function of individual domains,
proteins are usually subjected to limited proteolysis, followed by isolation of protein
fragments and determination of their functions.  We have developed an approach we call random
gene dissection for the identification of functional protein domains and their interdomain
regions as well as their in vivo complementing fragments.  The approach was tested on a
two-domain protein, the type IIS restriction endonuclease BfiI.  The collection of BfiI
insertional mutants was screened for those that are endonucleolytically active and thus induce
the SOS DNA repair response.  Sixteen isolated mutants of the wild-type specificity contained
insertions that were dispersed in a relatively large region of the target recognition domain.
They split the gene into two complementing parts that separately were unable to induce the SOS
DNA repair response.  In contrast, all 19 mutants of relaxed specificity contained the
cassette inserted into a very narrow interdomain region that connects BfiI domains responsible
for DNA recognition and for cleavage.  As expected, only the N-terminal fragment of BfiI was
required to induce SOS response.  Our results demonstrate that RGD can be used as a general
method to identify complementing fragments and functional domains in enzymes.

<>

<1>Sapranauskas, R., Sasnauskas, G., Lagunavicius, A., Vilkaitis, G., Lubys, A., Siksnys, V.
<2>Novel subtype of type IIs restriction enzymes. BfiI endonuclease exhibits similarities to the EDTA-resistant nuclease Nuc of Salmonella typhimurium.
<3>J. Biol. Chem.
<4>275
<5>30878-30885
<6>2000
<7>The type IIs restriction enzyme BfiI recognizes the non-palindromic nucleotide sequence
5'-ACTGGG-3' and cleaves complementary DNA strands 5/4 nucleotides downstream of the
recognition sequence. The genes coding for the BfiI restriction-modification (R-M) system were
cloned/sequenced and biochemical characterization of the BfiI restriction enzyme was
performed. The BfiI R-M system contained three proteins: two N4-methylcytosine
methyltransferases and a restriction enzyme. Sequencing of bisulfite-treated methylated DNA
indicated that each methyltransferase modifies cytosines on opposite strands of the
recognition sequence.  The N-terminal part of the BfiI restriction enzyme amino acid sequence
revealed intriguing similarities to an EDTA-resistant nuclease of Salmonella typhimurium.
Biochemical analyses demonstrated that BfiI, like the nuclease of S. typhimurium, cleaves DNA
in the absence of Mg(2+) ions and hydrolyzes an artificial substrate bis(p-nitrophenyl)
phosphate. However, unlike the nonspecific S. typhimurium nuclease, BfiI restriction enzyme
cleaves DNA specifically. We propose that the DNA-binding specificity of BfiI stems from the
C-terminal part of the protein. The catalytic N-terminal subdomain of BfiI radically differs
from that of type II restriction enzymes and is presumably similar to the EDTA-resistant
nonspecific nuclease of S. typhimurium; therefore, BfiI did not require metal ions for
catalysis. We suggest that BfiI represents a novel subclass of type IIs restriction enzymes
that differs from the archetypal FokI endonuclease by the fold of its cleavage domain, the
domain location, and reaction mechanism.

<>

<1>Saranathan, R., Tomar, A., Sudhakar, P., Arunkumar, K.P., Prashanth, K.
<2>Draft Genome Sequence of a Multidrug-Resistant Acinetobacter baumannii PKAB07 Clinical Strain from India Belonging to Sequence Type 195.
<3>Genome Announcements
<4>2
<5>e00184-14
<6>2014
<7>Acinetobacter baumannii has emerged as one of the most common nosocomial pathogens and is
considered to be a significant threat to public health
worldwide. Here, we present the draft genome sequence of a multidrug-resistant
clinical strain of A. baumannii PKAB07 isolated from a wound infection in India
during 2011 to 2012.

<>

<1>Saravanan, M., Bujnicki, J.M., Cymerman, I.A., Rao, D.N., Nagaraja, V.
<2>Type II restriction endonuclease R.KpnI is a member of the HNH nuclease superfamily.
<3>Nucleic Acids Res.
<4>32
<5>6129-6135
<6>2004
<7>The restriction endonuclease (REase) R.KpnI is an orthodox Type IIP enzyme, which binds to DNA
in the absence of metal ions and cleaves the DNA sequence 5'-GGTAC^C-3' in the presence of
Mg2+ as shown generating 3' four base overhangs. Bioinformatics analysis reveals that R.KpnI
contains a beta-beta-alpha-Me-finger fold, which is characteristic of many HNH-superfamily
endonucleases, including homing endonuclease I-HmuI, structure-specific T4 endonuclease VII,
colicin E9, sequence non-specific Serratia nuclease and sequence-specific homing endonuclease
I-PpoI. According to our homology model of R.KpnI, D148, H149 and Q175 correspond to the
critical D, H and N or H residues of the HNH nucleases. Substitutions of these three conserved
residues lead to the loss of the DNA cleavage activity by R.KpnI, confirming their importance.
The mutant Q175E fails to bind DNA at the standard conditions, although the DNA binding and
cleavage can be rescued at pH 6.0, indicating a role for Q175 in DNA binding and cleavage. Our
study provides the first experimental evidence for a Type IIP REase that does not belong to
the PD...D/EXK superfamily of nucleases, instead is a member of the HNH superfamily.

<>

<1>Saravanan, M., Elango, K., Chandrashekaran, S., Anand, N., Nagaraja, V.
<2>A new type II restriction endonuclease, OfoI from nonheterocystous cynobacterium Oscillatoria foreaui.
<3>Curr. Sci.
<4>85
<5>188-190
<6>2003
<7>Although restriction enzymes are widely distributed in nature, many bacterial genera are yet
to be explored for the presence of this important class of enzymes.  We have purified and
characterized a new type II restriction endonuclease, OfoI from a nonheterocyst cyanobacterium
Oscillatoria foreaui.  The recognition sequence has been determined by primer extension
analysis.  The purified enzyme OfoI recognizes and cleaves the palindromic hexanucleotide
5'-C/YCGRG-3', generating 5'-protruding ends.

<>

<1>Saravanan, M., Vasu, K., Ghosh, S., Nagaraja, V.
<2>Dual role for Zn2+ in maintaining structural integrity and inducing DNA sequence specificity in a promiscuous endonuclease.
<3>J. Biol. Chem.
<4>282
<5>32320-32326
<6>2007
<7>We describe two uncommon roles for Zn2+ in enzyme KpnI restriction endonuclease (REase). Among
all of the REases studied, KpnI REase is unique in its DNA binding and cleavage
characteristics. The enzyme is a poor discriminator of DNA sequences, cleaving DNA in a
promiscuous manner in the presence of Mg2+. Unlike most Type II REases, the active site of the
enzyme comprises an HNH motif, which can accommodate Mg2+, Mn2+, or Ca2+. Among these metal
ions, Mg2+ and Mn2+ induce promiscuous cleavage by the enzyme, whereas Ca2+-bound enzyme
exhibits site-specific cleavage. Examination of the sequence of the protein revealed the
presence of a zinc finger CCCH motif rarely found in proteins of prokaryotic origin. The zinc
binding motif tightly coordinates zinc to provide a rigid structural framework for the enzyme
needed for its function. In addition to this structural scaffold, another atom of zinc binds
to the active site to induce high fidelity cleavage and suppress the Mg2+- and Mn2+-mediated
promiscuous behavior of the enzyme. This is the first demonstration of distinct structural and
catalytic roles for zinc in an enzyme, suggesting the distinct origin of KpnI REase.

<>

<1>Saravanan, M., Vasu, K., Kanakaraj, R., Rao, D.N., Nagaraja, V.
<2>R.KpnI, an HNH superfamily REase, exhibits differential discrimination at non-canonical sequences in the presence of Ca2+ and Mg2+.
<3>Nucleic Acids Res.
<4>35
<5>2777-2786
<6>2007
<7>KpnI REase recognizes palindromic sequence, GGTAC downward arrowC, and forms complex in the
absence of divalent metal ions, but requires the ions for DNA cleavage. Unlike most other
REases, R.KpnI shows promiscuous DNA cleavage in the presence of Mg(2+). Surprisingly, Ca(2+)
suppresses the Mg(2+)-mediated promiscuous activity and induces high fidelity cleavage. To
further analyze these unique features of the enzyme, we have carried out DNA binding and
kinetic analysis. The metal ions which exhibit disparate pattern of DNA cleavage have no role
in DNA recognition. The enzyme binds to both canonical and non-canonical DNA with comparable
affinity irrespective of the metal ions used. Further, Ca(2+)-imparted exquisite specificity
of the enzyme is at the level of DNA cleavage and not at the binding step. With the canonical
oligonucleotides, the cleavage rate of the enzyme was comparable for both Mg(2+)- and
Mn(2+)-mediated reactions and was about three times slower with Ca(2+). The enzyme
discriminates non-canonical sequences poorly from the canonical sequence in Mg(2+)-mediated
reactions unlike any other Type II REases, accounting for the promiscuous behavior. R.KpnI,
thus displays properties akin to that of typical Type II REases and also endonucleases with
degenerate specificity in its DNA recognition and cleavage properties.

<>

<1>Saravanan, M., Vasu, K., Nagaraja, V.
<2>Evolution of sequence specificity in a restriction endonuclease by a point mutation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>10344-10347
<6>2008
<7>Restriction endonucleases (REases) protect bacteria from invading foreign DNAs and are endowed
with exquisite sequence specificity. REases have
originated from the ancestral proteins and evolved new sequence
specificities by genetic recombination, gene duplication, replication
slippage, and transpositional events. They are also speculated to have
evolved from nonspecific endonucleases, attaining a high degree of
sequence specificity through point mutations. We describe here an example
of generation of exquisitely site-specific REase from a highly-promiscuous
one by a single point mutation.

<>

<1>Sargueil, B., Delahodde, A., Hatat, D., Tian, G.L., Lazowska, J., Jacq, C.
<2>A new specific DNA endonuclease activity in yeast mitochondria.
<3>Mol. Gen. Genet.
<4>225
<5>340-341
<6>1991
<7>Two group I intron-encoded proteins from the yeast mitochondrial genome have already been
shown to have a specific DNA endonuclease activity. This activity mediates intron insertion by
cleaving the DNA sequence corresponding to the splice junction of an intronless strain. We
have discovered in mitochondrial extracts from the yeast strain 777-3A a new DNA endonuclease
activity which cleaves the fused exon A3-exon A4 junction sequence of the COXI gene.

<>

<1>Sargueil, B., Hatat, D., Delahodde, A., Jacq, C.
<2>In vivo and in vitro analyses of an intron-encoded DNA endonuclease from yeast mitochondria.  Recognition site by site-directed mutagenesis.
<3>Nucleic Acids Res.
<4>18
<5>5659-5665
<6>1990
<7>The pal 4 nuclease (termed I-Sce II) is encoded in the group I aI 4 intron of the COX I gene
of Saccharomyces cerevisiae. It introduces a specific double-strand break at the junction of
the two exons A4-A5 and thus mediates the insertion of the intron into an intronless strain.
To define the sequence recognized by pal 4 we introduced 35 single mutations in its target
sequence and examined their cleavage properties either in vivo in E. coli (when different
forms of the paI 4 proteins were artificially produced) or in vitro with mitochondrial
extracts of a mutant yeast strain blocked in the splicing of the aI 4 intron. We also detected
the pal 4 DNA endonuclease activity in extracts of the wild type strain. The results suggest
that 6 to 9 noncontiguous bases in the 17 base-pair region examined are necessary for pal 4
nuclease to bind and cleave its recognition site. We observed that the pal 4 nuclease
specificity can be significantly different with the different forms of the protein thus
explaining why only some forms are highly toxic in E. coli. This study shows that pal 4
recognition site is a complex phenomenon and this might have evolutionary implications on the
transfer properties of the intron.

<>

<1>Saridaki, A., Sapountzis, P., Harris, H.L., Batista, P.D., Biliske, J.A., Pavlikaki, H., Oehler, S., Savakis, C., Braig, H.R., Bourtzis, K.
<2>Wolbachia Prophage DNA Adenine Methyltransferase Genes in Different Drosophila-Wolbachia Associations.
<3>PLoS ONE
<4>6
<5>e19708
<6>2011
<7>Wolbachia is an obligatory intracellular bacterium which often manipulates the reproduction of
its insect and isopod hosts. In
contrast, Wolbachia is an essential symbiont in filarial nematodes.
Lately, Wolbachia has been implicated in genomic imprinting of host DNA
through cytosine methylation. The importance of DNA methylation in cell
fate and biology calls for in depth studing of putative
methylation-related genes. We present a molecular and phylogenetic
analysis of a putative DNA adenine methyltransferase encoded by a
prophage in the Wolbachia genome. Two slightly different copies of the
gene, met1 and met2, exhibit a different distribution over various
Wolbachia strains. The met2 gene is present in the majority of strains,
in wAu, however, it contains a frameshift caused by a 2 bp deletion.
Phylogenetic analysis of the met2 DNA sequences suggests a long
association of the gene with the Wolbachia host strains. In addition,
our analysis provides evidence for previously unnoticed multiple
infections, the detection of which is critical for the molecular
elucidation of modification and/or rescue mechanism of cytoplasmic
incompatibility.

<>

<1>Sarikhan, S., Azarbaijani, R., Yeganeh, L.P., Fazeli, A.S., Amoozegar, M.A., Salekdeh, G.H.
<2>Draft Genome Sequence of Nesterenkonia sp. Strain F, Isolated From Aran-Bidgol Salt Lake in Iran.
<3>J. Bacteriol.
<4>193
<5>5580
<6>2011
<7>The draft genome of the aerobic, Gram-positive, halophilic chemoorganotroph Nesterenkonia sp.
strain F consists of a 2,812,133-bp
chromosome. This study is the first to report the shotgun-sequenced draft
genome of a member of the genus Nesterenkonia.

<>

<1>Sarma, M.H., Dhingra, M.M., Gupta, G., Sarma, R.H.
<2>Conformational Microheterogeniety in a DNA Double Helix:  Structure of Restriction Endonuclease BamHI Recognition Site.
<3>Biochem. Biophys. Res. Commun.
<4>131
<5>269-276
<6>1985
<7>Structural studies using 500 MHz 1H NMR spectroscopy on BamHI recognition site
d(GGATCC)2 in solution at 19C is reported.  The resonances from the sugar ring
and base protons have been assigned from the 2D-COSY and NOESY spectra.
Analyses of the NOESY cross-peaks between the base protons H8/H6 and sugar
protons H2'/H2", H3' reveal that the nucleotide units G2, A3 and C6 adopt
(C3'-endo, x = 200o-220o) conformation while G1, T4 and C5 exhibit (C2'-endo,
x= 240o-260o) conformation.  NMR data clearly suggest that the two strands of
d(GGATCC)2 are conformationally equivalent and there is a structural two-fold
between the two A-T pairs.  The above information and the NOESY data are used
to generate a structural model of d(GGATCC)2.  The important features are: (i)
G1-G2 stack, the site of cleavage, shows an alternation in sugar pucker i.e.
C2'-endo, C3'-endo as in a B-A junction, (ii) G2-A3 stack adopts a mini A-DNA,
both the sugars being C3'-endo, (iii) A3-T4 stack, the site of two-fold,
displays an A-B junction with alternation in sugar pucker as C3'-endo,
C2'-endo, (iv) T4-C5 stack adopts a mini B-DNA both the sugars being C2'-endo
and (v) C5-C6 stack exhibits a B-A junction with C2'-endo, C3'-endo sugar
puckers.  Thus, our studies demonstrate that conformational microheterogeniety
with a structural two fold, is present in the BamHI recognition site.

<>

<1>Sarrade-Loucheur, A., Xu, S.Y., Chan, S.H.
<2>The Role of the Methyltransferase Domain of Bifunctional Restriction Enzyme RM.BpuSI in Cleavage Activity.
<3>PLoS ONE
<4>8
<5>e80967
<6>2013
<7>Restriction enzyme (REase) RM.BpuSI can be described as a Type IIS/C/G REase for its cleavage
site outside of the recognition sequence (Type IIS), bifunctional polypeptide possessing both
methyltransferase (MTase) and endonuclease activities (Type IIC) and endonuclease activity
stimulated by S-adenosyl-L-methionine (SAM) (Type IIG). The stimulatory effect of SAM on
cleavage activity presents a major paradox: a co-factor of the MTase activity that renders the
substrate unsusceptible to cleavage enhan es the cleavage activity. Here we show that the RM.
BpuSI MTase activity modifies both cleavage substrate and product only when they are
unmethylated. The MTase activity is, however, much lower than that of M1. BpuSI and is thought
not to be the major MTase for host DNA protection. SAM and sinefungin (SIN) increase the V-max
of the RM. BpuSI cleavage activity with a proportional change in Km, suggesting the presence
of an energetically more favorable pathway is taken. We further showed that RM. BpuSI under
goes substantial conformational changes in the presence of Ca2+, SIN, cleavage substrate
and/or product. Distinct conformers are inferred as the pre-cleavage/cleavage state (in the
presence of Ca2+, substrate or both) and MTase state (in the presence of SIN and substrate,
SIN and product or product alone). Interestingly, RM. BpuSI adopts a unique conformation when
only SIN is present. This SIN-bound state is inferred as a branch point for cleavage and MTase
activity and an intermediate to an energetically  favorable pathway for cleavage, probably
through increasing the binding affinity of the substrate to the enzyme under cleavage
conditions. Mutation of a SAM-binding residue resulted in altered conformational changes in
the presence of substrate or Ca2+ and eliminated cleavage activity. The present study
underscores the role of the MTase domain as facilitator of efficient cleavage activity for RM.
BpuSI.

<>

<1>Sarver, J., Stone, K., Townsend, J., Sapienza, P., Jen-Jacobson, L., Saxena, S.
<2>In the Arms of EcoRI - probing the Binding Specificity of the Restriction Endonuclease Using Electron Spin Resonance.
<3>Biophys. J.
<4>100
<5>144a-145a
<6>2011
<7>Pulsed electron spin resonance (ESR) was used to probe the binding specificity of EcoRI, a
restriction endonuclease that binds to and cleaves a six base pair sequence of DNA. EcoRI
binds to the specific sequence GAATTC with an affinity that is 50,000-90,000-fold greater than
that of a miscognate site that differs by only one base pair. Low binding affinity is also
exhibited at non-specific binding sites which differ from the specific sequence by two or more
base pairs.  Distance measurements were performed on several spin labeled EcoRI mutants when
bound to specific, miscognate, and non-specific sequences of DNA using Double
Electron-Electron Resonance. These distances demonstrated that on average the arms of EcoRI,
thought to play a major role in binding specificity, are similarly positioned. Additionally,
noncognate (miscognate and non-specific) complexes demonstrated broader distance distributions
indicating that the flexibility
of the arms is greater in these complexes. Room temperature continuous
wave (CW) experiments were also performed on the EcoRI mutant complexes at both X-band and
W-band to probe the arm region dynamics. Higher sensitivity
to the fast motional dynamics of the spin label at W-band resolved differences in two of the
EcoRI complexes that were not apparent in the X-band CW spectra. Molecular dynamics (MD)
simulations were performed on the
spin-label-modified specific EcoRI-DNA crystal structure to model the average
nitroxide orientation. Disparity in average distance as well as distribution indicates a need
for further sampling of the spin label in silico. This work is supported by NSF.

<>

<1>Sasaki, A., Oka, M., Shigenori, M.
<2>Composition where restriction enzyme SfiI is stabilised - containing magnesium chloride in glycerol solution.
<3>Japanese Patent Office
<4>JP 4023987
<5>
<6>1992
<7>The composition comprises a glycerol soln. contg. MgCl2 and the restriction enzyme SfiI.
Preferably the composition contains the restriction enzyme by 2-200U/microlitre and MgCl2 by
1-20mM, pref 5-20mM. It contains a buffer pref the tris hydrochloride buffer soln, in order to
adjust to pH7-8 and opt sodium chloride, potassium chloride, ethylenediamine tetraacetic acid
or dithiothreitol.

USE/ADVANTAGE - The restriction enzyme SfiI generated by Streptomyces fimbriatus ATCC 15051
is stabilised in the composition.


<>

<1>Sasaki, H., Ishikawa, H., Asano, R., Ueshiba, H., Matsumoto, T., Boot, R., Kawamoto, E.
<2>Draft Genome Sequence of the Rodent Opportunistic Pathogen Pasteurella pneumotropica ATCC 35149T.
<3>Genome Announcements
<4>2
<5>e00771-14
<6>2014
<7>Pasteurella pneumotropica is an opportunistic pathogen in rodents that is commonly isolated
from upper respiratory tracts in laboratory rodents. Here, we
report the draft genome sequence of the P. pneumotropica type strain ATCC 35149,
which was first isolated and characterized as biotype Jawetz.

<>

<1>Sasaki, J., Murakami, M., Yamada, Y.
<2>Purification, properties and recognition sequence of site-specific restriction endonuclease from Gluconobacter cerinus IFO 3285.
<3>Agric. Biol. Chem.
<4>49
<5>3017-3022
<6>1985
<7>A type II restriction endonuclease designated as GceCLI, was purified from
cells of Gluconobacter cerinus IFO 3285.  The purified enzyme was found to be
homogeneous on polyacrylamide gel disc electrophoresis.  The enzyme worked best
at 37C and pH 7.5 and required 7mm MgCl2 and 100 mm NaCl.  The purified enzyme
was stable when preincubated over a pH range of 7.5 to 9.5 for 12 hr at 4C and
a temperature range of 37 to 40C for 5 min at pH 7.5.  The enzyme was shown to
cleave lambda, PhiX174 RF, SV40, pBR322, M13 mp7 RF and Ad2 DNAs at 4, 1,0,0,0
and 25 or more sites, respectively, and to recognize the DNA sequence of
5'-C-C-G-G-3' and to cut between C and G on the right side of the sequence,
being an isoschizomer of SacII of Streptomyces achromogenes ATCC 12767.

<>

<1>Sasaki, J., Yamada, Y.
<2>Purification, properties and recognition sequence of site-specific restriction endonuclease from Acetobacter liquefaciens AJ 2881.
<3>Agric. Biol. Chem.
<4>48
<5>3027-3034
<6>1984
<7>A type II restriction endonuclease, designated as AliAJI, was purified from
cells of Acetobacter liquefaciens AJ 2881 by combined column chromatography on
heparin-Sepharose CL-6B, DEAE-Sepharose CL-6B and blue Sepharose CL-6B.  The
purified enzyme was homogeneous on polyacrylamide gel disc electrophoresis, and
the enzyme preparation was free from other nuclease activities, as judged by
constancy of lambda DNA-digest electrophoretic patterns after prolonged
incubation fro 24 hr.  The enzyme was optimally active at 37C at pH 7.5,
required neither sodium chloride nor ammonium sulfate, both of which rather
inhibited enzyme activity at high concentration (100 and 75 mM, respectively),
and cleaved lambda, PhiX174 RF, SV40, pBR322, M13 mp7RF and Ad2 DNAs at 18, 1,
2, 1, 1 and 25 more sites, respectively.  The recognition sequence of the
enzyme on DNA molecules was determined to be 5'-C-T-G-C-A-G-3', and the enzyme
was found to cut between A and G in the sequence, being an isoschizomer of the
endonuclease of Providencia stuartii 164 (PstI).

<>

<1>Sasaki, Y., Ishikawa, J., Yamashita, A., Oshima, K., Kenri, T., Furuya, K., Yoshino, C., Horino, A., Shiba, T., Sasaki, T., Hattori, M.
<2>The complete genomic sequence of Mycoplasma penetrans, an intracellular bacterial pathogen in humans.
<3>Nucleic Acids Res.
<4>30
<5>5293-5300
<6>2002
<7>The complete genomic sequence of an intracellular bacterial pathogen, Mycoplasma penetrans
HF-2 strain, was determined. The HF-2 genome consists of a 1 358 633 bp single circular
chromosome containing 1038 predicted coding sequences (CDSs), one set of rRNA genes and 30
tRNA genes. Among the 1038 CDSs, 264 predicted proteins are common to the Mycoplasmataceae
sequenced thus far and 463 are M.penetrans specific. The genome contains the two-component
system but lacks the essential cellular gene, uridine kinase. The relatively large genome of
M.penetrans HF-2 among mycoplasma species may be accounted for by both its rich core proteome
and the presence of a number of paralog families corresponding to 25.4% of all CDSs. The
largest paralog family is the p35 family, which encodes surface lipoproteins including the
major antigen, P35. A total of 44 genes for p35 and p35 homologs were identified and 30 of
them form one large cluster in the chromosome. The genetic tree of p35 paralogs suggests the
occurrence of dynamic chromosomal rearrangement in paralog formation during evolution. Thus,
M.penetrans HF-2 may have acquired diverse repertoires of antigenic variation-related genes to
allow its persistent infection in humans.

<>

<1>Sasnauskas, G., Connolly, B.A., Halford, S.E., Siksnys, V.
<2>Site-specific DNA transesterification catalyzed by a restriction enzyme.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>104
<5>2115-2120
<6>2007
<7>Most restriction endonucleases use Mg2+ to hydrolyze phosphodiester bonds at specific DNA
sites. We show here that Bfil, a
metal-independent restriction enzyme from the phospholipase D
superfamily, catalyzes both DNA hydrolysis and transesterification
reactions at its recognition site. In the presence of alcohols such as
ethanol or glycerol, it attaches the alcohol covalently to the 5'
terminus of the cleaved DNA. Under certain conditions, the terminal
3'-OH of one DNA strand can attack the target phosphodiester bond in
the other strand to create a DNA hairpin. Transesterification reactions
on DNA with phosphorothioate linkages at the target bond proceed with
retention of stereoconfiguration at the phosphorus, indicating,
uniquely for a restriction enzyme, a two-step mechanism. We propose
that Bfil first makes a covalent enzyme-DNA intermediate, and then it
resolves it by a nucleophilic attack of water or an alcohol, to yield
hydrolysis or transesterification products, respectively.

<>

<1>Sasnauskas, G., Connolly, B.A., Halford, S.E., Siksnys, V.
<2>Template-directed addition of nucleosides to DNA by the BfiI restriction enzyme.
<3>Nucleic Acids Res.
<4>36
<5>3969-3977
<6>2008
<7>Restriction endonucleases catalyse DNA cleavage at specific sites. The BfiI endonuclease cuts
DNA to give staggered ends with 1-nt 3'-extensions.
We show here that BfiI can also fill in the staggered ends: while cleaving
DNA, it can add a 2'-deoxynucleoside to the reaction product to yield
directly a blunt-ended DNA. We propose that nucleoside incorporation
proceeds through a two-step reaction, in which BfiI first cleaves the DNA
to make a covalent enzyme-DNA intermediate and then resolves it by a
nucleophilic attack of the 3'-hydroxyl group of the incoming nucleoside,
to yield a transesterification product. We demonstrate that base pairing
of the incoming nucleoside with the protruding DNA end serves as a
template for the incorporation and governs the yield of the elongated
product. The efficiency of the template-directed process has been
exploited by using BfiI for the site-specific modification of DNA
5'-termini with an amino group using a 5'-amino-5'-deoxythymidine.

<>

<1>Sasnauskas, G., Halford, S.E., Siksnys, V.
<2>How the BfiI restriction enzyme uses one active site to cut two DNA strands.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>6410-6415
<6>2003
<7>Unlike other restriction enzymes, BfiI functions without metal ions. It recognizes an
asymmetric DNA sequence, 5'-ACTGGG-3', and cuts top and
bottom strands at fixed positions downstream of this sequence. Many
restriction enzymes are dimers of identical subunits, with one active site
for each DNA strand. Others, like FokI, dimerize transiently during
catalysis. BfiI is also a dimer but it has only one active site, at the
dimer interface. We show here that BfiI remains a dimer as it makes
double-strand breaks in DNA and that its single active site acts
sequentially, first on the bottom and then the top strand. Hence, after
cutting the bottom strand, a rearrangement of either the protein and/or
the DNA in the BfiI-DNA complex must switch the active site to the top
strand. Low pH values selectively block top-strand cleavage, converting
BfiI into a nicking enzyme that cleaves only the bottom strand. The switch
to the top strand may depend on the ionization of the cleaved 5' phosphate
in the bottom strand. BfiI thus uses a mechanism for making double-strand
breaks that is novel among restriction enzymes.

<>

<1>Sasnauskas, G., Jeltsch, A., Pingoud, A., Siksnys, V.
<2>Plasmid DNA cleavage by MunI restriction enzyme: Single-turnover and steady-state kinetic analysis.
<3>Biochemistry
<4>38
<5>4028-4036
<6>1999
<7>Mutational analysis has previously indicated that D83 and E98 residues are essential for DNA
cleavage activity and presumably chelate a Mg2+ ion at the active site of MunI restriction
enzyme. In the absence of metal ions, protonation of an ionizable residue with a pKa > 7.0,
most likely one of the active site carboxylates, controls the DNA binding specificity of MunI.
Thus, competition between H+ and Mg2+ binding at the active site of MunI presumably plays an
important role in catalysis/binding. In the present study we have identified elementary steps
and intermediates in the reaction pathway of plasmid DNA cleavage by MunI and elucidated the
effect of pH and Mg2+ ions on the individual steps of the DNA cleavage reaction. The kinetic
analysis indicated that the multiple-turnover rate of plasmid cleavage by MunI is limited by
product release     throughout the pH range 6.0-9.3. Quenched-flow experiments revealed that
open circle DNA is an obligatory intermediate in the reaction pathway. Under optimal reaction
conditions, open circle DNA remains bound to the MunI; however it is released into the
solution at low [MgCl2]. Rate constants for the phosphodiester bond hydrolysis of the first
(k1) and second (k2) strand of plasmid DNA at pH 7.0 and 10 mM MgCl2 more than 100-fold exceed
the kcat value which is limited by product dissociation. The analysis of the pH and [Mg2+]
dependences of k1 and k2 revealed that both H+ and Mg2+ ions compete for the binding to the
same residue at the active site of MunI. Thus, the decreased rate of phosphodiester hydrolysis
by MunI at pH < 7.0 may be due to the reduction of affinity for the Mg2+ binding at the active
site. Kinetic analysis of DNA cleavage by MunI yielded estimates for the
association-dissociation rate constants of enzyme-substrate complex and demonstrated the
decreased stability of the MunI-DNA complex at pH values above 8.0.

<>

<1>Sasnauskas, G., Kostiuk, G., Tamulaitis, G., Siksnys, V.
<2>Target site cleavage by the monomeric restriction enzyme BcnI requires translocation to a random DNA sequence and a switch in enzyme orientation.
<3>Nucleic Acids Res.
<4>39
<5>8844-8856
<6>2011
<7>Endonucleases that generate double-strand breaks in DNA often possess two identical subunits
related by rotational symmetry, arranged so that the
active sites from each subunit act on opposite DNA strands. In contrast to
many endonucleases, Type IIP restriction enzyme BcnI, which recognizes the
pseudopalindromic sequence 5'-CCSGG-3' (where S stands for C or G) and
cuts both DNA strands after the second C, is a monomer and possesses a
single catalytic center. We show here that to generate a double-strand
break BcnI nicks one DNA strand, switches its orientation on DNA to match
the polarity of the second strand and then cuts the phosphodiester bond on
the second DNA strand. Surprisingly, we find that an enzyme flip required
for the second DNA strand cleavage occurs without an excursion into bulk
solution, as the same BcnI molecule acts processively on both DNA strands.
We provide evidence that after cleavage of the first DNA strand, BcnI
remains associated with the nicked intermediate and relocates to the
opposite strand by a short range diffusive hopping on DNA.

<>

<1>Sasnauskas, G., Zakrys, L., Zaremba, M., Cosstick, R., Gaynor, J.W., Halford, S.E., Siksnys, V.
<2>A novel mechanism for the scission of double-stranded DNA: BfiI cuts both 3'-5' and 5'-3' strands by rotating a single active site.
<3>Nucleic Acids Res.
<4>38
<5>2399-2410
<6>2010
<7>Metal-dependent nucleases that generate double-strand breaks in DNA often possess two
symmetrically-equivalent subunits, arranged so that the active
sites from each subunit act on opposite DNA strands. Restriction
endonuclease BfiI belongs to the phospholipase D (PLD) superfamily and
does not require metal ions for DNA cleavage. It exists as a dimer but has
at its subunit interface a single active site that acts sequentially on
both DNA strands. The active site contains two identical histidines
related by 2-fold symmetry, one from each subunit. This symmetrical
arrangement raises two questions: first, what is the role and the
contribution to catalysis of each His residue; secondly, how does a
nuclease with a single active site cut two DNA strands of opposite
polarities to generate a double-strand break. In this study, the roles of
active-site histidines in catalysis were dissected by analysing
heterodimeric variants of BfiI lacking the histidine in one subunit. These
variants revealed a novel mechanism for the scission of double-stranded
DNA, one that requires a single active site to not only switch between
strands but also to switch its orientation on the DNA.

<>

<1>Sass, P., Berscheid, A., Jansen, A., Oedenkoven, M., Szekat, C., Strittmatter, A., Gottschalk, G., Bierbaum, G.
<2>Genome Sequence of Staphylococcus aureus VC40, a Vancomycin- and Daptomycin-Resistant Strain, To Study the Genetics of Development of Resistance  to Currently Applied Last-Resort Antibiotics.
<3>J. Bacteriol.
<4>194
<5>2107-2108
<6>2012
<7>The increasing emergence of multidrug-resistant Staphylococcus aureus is a problem of global
importance. Here, we report the genome of S. aureus VC40, which
is resistant to the last-resort antibiotics vancomycin and daptomycin. Its genome
sequence will allow insights into the mechanisms that convey full resistance to
these compounds.

<>

<1>Sassera, D., Gaiarsa, S., Scaltriti, E., Morganti, M., Bandi, C., Casadei, G., Pongolini, S.
<2>Draft Genome Sequence of Salmonella enterica subsp. enterica Serovar Manhattan Strain 111113, from an Outbreak of Human Infections in Northern Italy.
<3>Genome Announcements
<4>1
<5>e00632-13
<6>2013
<7>We announce the draft genome sequence of Salmonella enterica subsp. enterica serovar Manhattan
strain 111113, isolated from a patient during an outbreak in
northern Italy. The genome, which was obtained with Illumina MiSeq technology, is
composed of 21 contigs for a total of 4,684,342 bp, with a G+C content of 52.17%.

<>

<1>Sassera, D., Leardini, I., Villa, L., Comandatore, F., Carta, C., Almeida, A., do Ceu, S.M., Gaiarsa, S., Marone, P., Pozio, E., Caccio, S.M.
<2>Draft Genome Sequence of Stenotrophomonas maltophilia Strain EPM1, Found in Association with a Culture of the Human Parasite Giardia duodenalis.
<3>Genome Announcements
<4>1
<5>e00182-13
<6>2013
<7>We report the draft genome sequence of the Stenotrophomonas maltophilia strain EPM1, found in
association with a culture of Giardia duodenalis. The draft genome
sequence of S. maltophilia strain EPM1, obtained with Roche 454 GS-FLX Titanium
technology, is composed of 19 contigs totaling 4,785,869 bp, with a G+C content
of 66.37%.

<>

<1>Sassera, D., Lo, N., Epis, S., D'Auria, G., Montagna, M., Comandatore, F., Horner, D., Pereto, J., Luciano, A.M., Franciosi, F., Ferri, E., Crotti, E., Bazzocchi, C., Daffonchio, D., Sacchi, L., Moya, A., Latorre, A., Bandi, C.
<2>Phylogenomic evidence for the presence of a flagellum and cbb3 oxidase in the free-living mitochondrial ancestor.
<3>Mol. Biol. Evol.
<4>28
<5>3285-3296
<6>2011
<7>The initiation of the intracellular symbiosis that would give rise to
mitochondria and eukaryotes was a major event in the history of life on
earth. Hypotheses to explain eukaryogenesis fall into two broad and
competing categories: those proposing that the host was a phagocytotic
proto-eukaryote that preyed upon the free-living mitochondrial ancestor
(hereafter FMA) and those proposing that the host was an archaebacterium
that engaged in syntrophy with the FMA. Of key importance to these
hypotheses are whether the FMA was motile or non-motile, and the
atmospheric conditions under which the FMA thrived. Reconstructions of the
FMA based on genome content of Rickettsiales representatives - generally
considered to be the closest living relatives of mitochondria - indicate
that it was non-motile and aerobic. We have sequenced the genome of
Candidatus Midichloria mitochondrii, a novel and phylogenetically
divergent member of the Rickettsiales. We found that it possesses unique
gene sets found in no other Rickettsiales, including 26 genes associated
with flagellar assembly, and a cbb(3)-type cytochrome oxidase.
Phylogenomic analyses show that these genes were inherited in a vertical
fashion from an ancestral alpha-proteobacterium, and indicate that the FMA
possessed a flagellum, and could undergo oxidative phosphorylation under
both aerobic and microoxic conditions. These results indicate that the FMA
played a more active and potentially parasitic role in eukaryogenesis than
currently appreciated, and provide an explanation for how the symbiosis
could have evolved under low levels of oxygen.

<>

<1>Sassi, M., Croce, O., Robert, C., Raoult, D., Drancourt, M.
<2>Draft Genome Sequence of Mycobacterium triplex DSM 44626.
<3>Genome Announcements
<4>2
<5>e00499-14
<6>2014
<7>We announce the draft genome sequence of Mycobacterium triplex strain DSM 44626,  a
nontuberculosis species responsible for opportunistic infections. The genome
described here is composed of 6,382,840 bp, with a G+C content of 66.57%, and
contains 5,988 protein-coding genes and 81 RNA genes.

<>

<1>Sassi, M., Felden, B., Augagneur, Y.
<2>Draft Genome Sequence of Staphylococcus aureus subsp. aureus Strain HG003, an NCTC8325 Derivative.
<3>Genome Announcements
<4>2
<5>e00855-14
<6>2014
<7>We report the draft genome sequence of a Staphylococcus aureus NCTC8325 derivative, strain
HG003. HG003 contains functional global regulators rsbU and
tcaR and is therefore considered as a reference for studies of regulation and
virulence. The genome is composed of 2,797,898 bp and will be essential for
subsequent RNAseq analysis.

<>

<1>Sassi, M., Robert, C., Raoult, D., Drancourt, M.
<2>Non-contiguous genome sequence of Mycobacterium simiae strain DSM 44165(T.).
<3>Standards in Genomic Sciences
<4>8
<5>306-317
<6>2013
<7>Mycobacterium simiae is a non-tuberculosis mycobacterium causing pulmonary infections in both
immunocompetent and imunocompromized patients. We announce the
draft genome sequence of M. simiae DSM 44165(T). The 5,782,968-bp long genome
with 65.15% GC content (one chromosome, no plasmid) contains 5,727 open reading
frames (33% with unknown function and 11 ORFs sizing more than 5000 -bp), three
rRNA operons, 52 tRNA, one 66-bp tmRNA matching with tmRNA tags from
Mycobacterium avium, Mycobacterium tuberculosis, Mycobacterium bovis,
Mycobacterium microti, Mycobacterium marinum, and Mycobacterium africanum and 389
DNA repetitive sequences. Comparing ORFs and size distribution between M. simiae
and five other Mycobacterium species M. simiae clustered with M. abscessus and M.
smegmatis. A 40-kb prophage was predicted in addition to two prophage-like
elements, 7-kb and 18-kb in size, but no mycobacteriophage was seen after the
observation of 10(6) M. simiae cells. Fifteen putative CRISPRs were found. Three
genes were predicted to encode resistance to aminoglycosides, betalactams and
macrolide-lincosamide-streptogramin B. A total of 163 CAZYmes were annotated. M.
simiae contains ESX-1 to ESX-5 genes encoding for a type-VII secretion system.
Availability of the genome sequence may help depict the unique properties of this
environmental, opportunistic pathogen.

<>

<1>Sassi, M., Robert, C., Raoult, D., Drancourt, M.
<2>Noncontiguous Genome Sequence of Mycobacterium septicum Strain DSM 44393T.
<3>Genome Announcements
<4>1
<5>e00574-13
<6>2013
<7>The rapidly growing Mycobacterium septicum rarely causes pulmonary infections. We report here
the draft genome sequence of M. septicum strain DSM 44393(T),
isolated from catheter-related bacteremia and initially identified as a member of
Mycobacterium fortuitum.

<>

<1>Sato, H., Suzuki, T., Yamada, Y.
<2>Purification of restriction endonuclease from Acetobacter aceti IFO 3281 (AatII) and its properties.
<3>Agric. Biol. Chem.
<4>54
<5>3319-3325
<6>1990
<7>The restriction endonuclease AatII was purified from cell-free extracts of
Acetobacter aceti IFO 3281 by streptomycin treatment, ammonium sulfate
fractionation, combined column chromatographies on DEAE-Toyopearl 650S,
heparin-Sepharose CL-6B and DEAE-Sepharose CL-6B and FPLC on Mono Q and on
Superose 12 (gel filtration).  The purified enzyme was homogeneous on
SDS-polyacrylamide gel disk electrophoresis.  The relative molecular mass of
the purified enzyme was 190,000 daltons by gel filtration.  The
SDS-polyacrylamide gel disk electrophoresis gave the relative molecular mass of
47,500 daltons.  These data indicated that the purified, native enzyme is a
tetramer (190,000 daltons) composed of four 47,500-dalton subunits.  The
isoelectric point of the enzyme was 6.0.  The purified enzyme was intensely
activated by manganese ion (50-fold increase or more when compared with
magnesium ion).  The enzyme worked best at 37C and pH 8.5 in a reaction mixture
(50 microliters) containing 1.0 micrograms lambda DNA, 10 mM Tris-HCI, 7 mM
2-mercaptoethanol, 7 mM MnCl2 and 50 mM NaCl.  The enzyme recognizes the same
palindromic hexanucleotide sequence 5'-GACGTC-3', cuts between T and C and
produces a 3'-tetranucleotide extension in the presence of MnCl2, as it does in
the presence of MgCl2.

<>

<1>Sato, H., Yamada, Y.
<2>The restriction endonuclease AatI from Acetobacter aceti IFO 3281, an isoschizomer of StuI, has a dimeric structure.
<3>J. Gen. Appl. Microbiol.
<4>36
<5>273-277
<6>1990
<7>The restriction endonuclease AatI, an isoschizomer of the StuI endonuclease,
was first reported in Acetobacter aceti IFO 3281 by Sugisaki et al.  However, a
detailed enzymatic study has not been done as yet regarding the AatI
endonuclease.  During the course of our studies on acetic acid bacteria, we
purified the AatI endonuclease to a homogeneous state and found that the enzyme
has a dimeric structure.  This paper describes the purification and properties
of the AatI endonuclease.

<>

<1>Sato, S., Hutchison, C.A. III, Harris, J.I.
<2>A thermostable sequence-specific endonuclease from Thermus aquaticus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>74
<5>542-546
<6>1977
<7>A sequence-specific endonuclease, TaqI, of novel specificity has been partially purified from
an extreme thermophile, Thermus aquaticus.  The enzyme cleaves bacteriophage lambda DNA at
many (>30) sites and bacteriophage PhiX174 RF DNA at 10 sites.  The enzyme is active at
temperatures up to 79C.  The cleavage sites on PhiX174 RF DNA have been mapped.  The sequence
recognized and cleaved by TaqI has been shown to be the symmetrical tetranucleotide:
5' T-^C-G-A 3'
3' A-G-C-^T 5'.

<>

<1>Sato, S., Nakazawa, K., Shinomiya, T.
<2>A DNA methylase from Thermus thermophilus HB8.
<3>J. Biochem. (Tokyo)
<4>88
<5>737-747
<6>1980
<7>A DNA methylase was purified in a homogeneous state from an extremely
thermophilic bacterium, Thermus thermophilus HB8, by chromatography on,
successively, phosphocellulose, CM-cellulose, and haparin-Sepharose.  The
molecular weight of the enzyme was determined to be about 44,000 by gel
filtration on a Sephadex G-100 column and 41,000 by SDS-polyacrylamide gel
electrophoresis, and these findings suggest a single polypeptide enzyme.  The
enzyme develops maximum activity around pH 7.4 and at 70C.  Enzymatic activity
is completely inhibited by 0.2M NaCl or 2 mM HgCl2.  The enzyme transfers
methyl groups from S-adenosyl-L-methionine to a double stranded DNA.  The sole
product of the reaction was identified as N-6-methyl adenine after hydrolysis
of the DNA with formic acid.  The enzyme kinetics obey the Michaelis-Menten
equation and Km values for S-adenosylmethionine and lambda phage DNA were
determined to be 0.8 microM and 10 microgram/ml, respectively.  The enzyme does
not transfer methyl groups to TthHB8I endonuclease digested DNA as well as the
host (T. thermophilus HB8) DNA.  The number of methyl groups of the fully
methylated PhiX174 RF DNA was about twice as many as TthHB8I endonuclease sites
on the DNA.  The distribution of the methyl groups of PhiX174 RF DNA among the
HaeIII  fragments was the same as that of TthHB8I endonuclease sites,
suggesting that this DNA methylase is the other component of the
modification-restriction system including TthHB8I endonuclease.  The enzyme
probably recognizes the sequence, 5'-TCGA-3', in a double stranded DNA and
probably methylates adenine in the above sequence.

<>

<1>Sato, S., Shinomiya, T.
<2>An isoschizomer of TaqI from Thermus thermophilus HB8.
<3>J. Biochem. (Tokyo)
<4>84
<5>1319-1321
<6>1978
<7>A site-specific endonuclease has been isolated from Thermus thermophilus HB8
and named TthHB81.  It recognizes the same sequences as TaqI from Thermus
aquaticus TY-1 does.  The amount of Tth HB81 in the cells was comparable to
that of TaqI.  T. thermophilus HB8 has an advantage over T. aquaticus YT-1 for
preparation of a TaqI-like enzyme since it is easier to obtain T. thermophilus
HB8 cells in quantity.

<>

<1>Sato, S., Umemura, M., Koike, H., Habe, H.
<2>Draft Genome Sequence of Gluconobacter frateurii NBRC 103465, a Glyceric Acid-Producing Strain.
<3>Genome Announcements
<4>1
<5>e00369-13
<6>2013
<7>Gluconobacter frateurii strain NBRC 103465 can efficiently produce glyceric acid  (GA) from
raw glycerol feedstock derived from biodiesel fuel production
processes. Here, we report the 3.4-Mb draft genome sequence of G. frateurii NBRC
103465. The draft genome sequence can be applied to examine the enzymes and
electron transport system involved in GA production.

<>

<1>Sato, T., Shimizu, T., Watarai, M., Kobayashi, M., Kano, S., Hamabata, T., Takeda, Y., Yamasaki, S.
<2>Genome analysis of a novel Shiga toxin 1 (Stx1)-converting phage which is closely related to Stx2-converting phages but not to other Stx1-converting phages.
<3>J. Bacteriol.
<4>185
<5>3966-3971
<6>2003
<7>Two Stx-converting phages, designated Stx1 phi and Stx2 phi-II, were isolated from an
Escherichia coli O157:H7 strain, Morioka V526, and their
entire nucleotide sequences were determined. The genomes of both phages
were similar except for the stx gene-flanking regions. Comparing these
phages to other known Stx-converting phages, we concluded that Stx1 phi is
a novel Stx1-converting phage closely related to Stx2-converting phages so
far reported.

<>

<1>Satou, K., Shiroma, A., Teruya, K., Shimoji, M., Nakano, K., Juan, A., Tamotsu, H., Terabayashi, Y., Aoyama, M., Teruya, M., Suzuki, R., Matsuda, M., Sekine, A., Kinjo, N., Kinjo, F., Yamaoka, Y., Hirano, T.
<2>Complete Genome Sequences of Eight Helicobacter pylori Strains with Different Virulence Factor Genotypes and Methylation Profiles, Isolated from Patients with Diverse Gastrointestinal Diseases on Okinawa Island, Japan.
<3>Genome Announcements
<4>2
<5>e00286-14
<6>2014
<7>We report the complete genome sequences of eight Helicobacter pylori strains isolated from
patients with gastrointestinal diseases in Okinawa, Japan.
Whole-genome sequencing and DNA methylation detection were performed using the PacBio
platform. De novo assembly determined a single, complete contig for each strain. Furthermore,
methylation analysis identified virulence factor genotype-dependent motifs.

<>

<1>Saunders, E. et al.
<2>Complete genome sequence of Haloterrigena turkmenica type strain (4k).
<3>Standards in Genomic Sciences
<4>2
<5>107-116
<6>2010
<7>Haloterrigena turkmenica (Zvyagintseva and Tarasov 1987) Ventosa et al. 1999, comb. nov. is
the type species of the genus Haloterrigena in the euryarchaeal
family Halobacteriaceae. It is of phylogenetic interest because of the yet
unclear position of the genera Haloterrigena and Natrinema within the
Halobacteriaceae, which created some taxonomic problems historically. H.
turkmenica, was isolated from sulfate saline soil in Turkmenistan, is a
relatively fast growing, chemoorganotrophic, carotenoid-containing, extreme
halophile, requiring at least 2 M NaCl for growth. Here we describe the features
of this organism, together with the complete genome sequence, and annotation.
This is the first complete genome sequence of the genus Haloterrigena, but the
eighth genome sequence from a member of the family Halobacteriaceae. The
5,440,782 bp genome (including six plasmids) with its 5,287 protein-coding and 63
RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Saunders, E. et al.
<2>Complete genome sequence of Eggerthella lenta type strain (IPP VPI 0255).
<3>Standards in Genomic Sciences
<4>1
<5>174-182
<6>2009
<7>Eggerthella lenta (Eggerth 1935) Wade et al. 1999, emended Wurdemann et al. 2009  is the type
species of the genus Eggerthella, which belongs to the
actinobacterial family Coriobacteriaceae. E. lenta is a Gram-positive,
non-motile, non-sporulating pathogenic bacterium that can cause severe
bacteremia. The strain described in this study has been isolated from a rectal
tumor in 1935. Here we describe the features of this organism, together with the
complete genome sequence, and annotation. This is the first complete genome
sequence of the genus Eggerthella, and the 3,632,260 bp long single replicon
genome with its 3123 protein-coding and 58 RNA genes is part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Saunders, N., Snyder, L.
<2>The minimal mobile element.
<3>Microbiology
<4>148
<5>3756-3760
<6>2002
<7>Horizontal transfer of genes is an integral component of bacterial evolution and is
particularly associated with processes related to environmental adaptation and virulence.  The
association of many host adaptive and virulence genes with mobile genetic elements such as
transposases and bacteriophages rejects this.  Presumably the association of the gene
conferring some competitive advantage for the recipient strain facilitates the dissemination
of the mobile element.  The presence of larger elements such as pathogenicity islands or
islands of horizontal transfer is also characteristic of some bacterial species.  However,
analysis of complete bacterial genomes suggests a previously unrecognized mechanism of
mobilization that utilizes natural transformation and homologous recombination, and that is
independent of transposases and other mobilization mechanisms.

<>

<1>Saunders, N.J., Jeffries, A.C., Peden, P.F., Hood, D.W., Tettelin, H., Rappuoli, R., Moxon, E.R.
<2>Repeat-associated phase variable genes in the complete genome sequence of Neisseria meningitidis strain MC58.
<3>Mol. Microbiol.
<4>37
<5>207-215
<6>2000
<7>Phase variation, mediated through variation in the length of simple sequence repeats, is
recognized as an important mechanism for
controlling the expression of factors involved in bacterial virulence.
Phase variation is associated with most of the currently recognized
virulence determinants of Neisseria meningitidis. Based upon the
complete genome sequence of the N. meningitidis serogroup B strain
MC58, we have identified tracts of potentially unstable simple sequence
repeats and their potential functional significance determined on the
basis of sequence context. Of the 65 potentially phase variable genes
identified, only 13 were previously recognized. Comparison with the
sequences from the other two pathogenic Neisseria sequencing projects
shows differences in the length of the repeats in 36 of the 65 genes
identified, including 25 of those not previously known to be phase
variable. Six genes that did not have differences in the length of the
repeat instead had polymorphisms such that the gene would not be
expected to be phase variable in at least one of the other strains. A
further 12 candidates did not have homologues in either of the other
two genome sequences. The large proportion of these genes that are
associated with frameshifts and with differences in repeat length
between the neisserial genome sequences is further corroborative
evidence that they are phase variable. The number of potentially phase
variable genes is substantially greater than for any other species
studied to date, and would allow N. meningitidis to generate a very
large repertoire of phenotypes through expression of these genes in
different combinations. Novel phase variable candidates identified in
the strain MC58 genome sequence include a spectrum of genes encoding
glycosyltransferases, toxin related products, and metabolic activities
as well as several restriction/modification and bacteriocin-related
genes and a number of open reading frames (ORFs) for which the function
is currently unknown. This suggests that the potential role of phase
variation in mediating bacterium-host interactions is much greater than
has been appreciated to date. Analysis of the distribution of
homopolymeric tract lengths indicates that this species has
sequence-specific mutational biases that favour the instability of
sequences associated with phase variation.

<>

<1>Saunders, N.J., Peden, J.F., Hood, D.W., Moxon, E.R.
<2>Simple sequence repeats in the Helicobacter pylori genome.
<3>Mol. Microbiol.
<4>27
<5>1091-1098
<6>1998
<7>We describe an integrated system for the analysis of DNA sequence motifs within complete
bacterial genome sequences.  This system is based around ACeDB, a genome database with an
integrated graphical user interface; we identify and display motifs in the context of genetic,
sequence and bibliographic data.  Tomb et al. (1997) previously reported the identification of
contingency genes in Helicobacter pylori through their association with homopolymeric tracts
and dinucleotide repeats.  With this as a starting point, we validated the system by a search
for this type of repeat and used the contextual information to assess the likelihood that they
mediate phase variation in the associated open reading frames.  We found all of the repeats
previously described, and identified 27 putative phase-variable genes (including 17 previously
described).  These could be divided into three groups: lipopolysaccharide biosynthesis,
cell-surface-associated proteins and DNA restriction/modification systems.  Five of the
putative genes did not have obvious homologues in any of the public domain sequence databases.
The reading frame of some ORFs was disrupted by the presence of the repeats, including the
alpha(1-2) fucosyltransferase gene, necessary for the synthesis of the Lewis Y epitope.  An
additional benefit of this approach is that the results of each search can be analyzed further
and compared with those from other genomes.  This revealed that H. pylori has an unusually
high frequency of homopurine:homopyrimidine repeats suggesting mechanistic biases that favor
their presence and instability.

<>

<1>Saves, I., Eleaume, H., Dietrich, J., Masson, J.-M.
<2>The Thy Pol-2 intein of Thermococcus hydrothermalis is an isoschizomer of PI-TliI and PI-TfuII endonucleases.
<3>Nucleic Acids Res.
<4>28
<5>4391-4396
<6>2000
<7>Thy Pol-2 intein, from Thermococcus hydrothermalis, belongs to the same allelic family as
TliPol-2 (PI-TliI), Tfu Pol-2 (PI-TfuII) and TspTY Pol-3 mini-intein, all inserted at the
pol-c site of archaeal DNA polymerase genes. This new intein was cloned, expressed in
Escherichia coli and purified. The intein is a specific endonuclease (PI-ThyI) which cleaves
the inteinless sequence of the ThyDNA pol gene.  Moreover, PI-TliI, PI-TfuII and PI-ThyI are
very similar endonucleases which cleave DNA in the same optimal conditions at 70 degrees C
yielding similar 3'-hydroxyl overhangs of 4 bp and the reaction is subject to product
inhibition. The three enzymes are able to cleave the three DNA sequences spanning the pol-c
site and a 24 bp consensus cleavage site was defined for the three isoschizomers. However, the
exact size of the minimal cleavage site depends both on the substrate sequence and the
endonuclease. The inability of the isoschizomers to cleave the inteinless DNA polymerase gene
from Pyrococcus spp. KOD is due to point substitutions on the 5' side of the pol-c site,
suggesting that the absence of inteins of this allelic family in DNA polymerase genes from
Pyrococcus spp. can be linked to small differences in the target site sequence.

<>

<1>Saves, I., Morlot, C., Thion, L., Rolland, J.-L., Dietrich, J., Masson, J.-M.
<2>Investigating the endonuclease activity of four Pyrococcus abyssi inteins.
<3>Nucleic Acids Res.
<4>30
<5>4158-4165
<6>2002
<7>Among the 14 inteins of the Pyrococcus abyssi genome, 10 harbour the LAGLIDADG motifs of
dodecapeptide endonucleases.  Four of these were cloned, expressed in Escherichia coli and
purified to assay their potential endonuclease activity.  PabRIR1-2 and PabRIR1-3 are specific
endonucleases, named PI-PabI and PI-PabII, respectively, cleaving the sequence spanning their
homing site.  This is consistent with their size and with the relative positions and sequences
of their endonuclease motifs.  However, PI-PabI is 10-fold more active than PI-PabII and a
discrepancy of the DNA recognition and cleavage mechanisms was observed between the two
inteins.  In particular, analysis of the DNA cleavage reactions by MALDI-TOF highlighted that
while the cleavage of DNA by PI-PabI consists of two steps corresponding to the cleavage of
each DNA strand, PI-PabII processes the two DNA strands simultaneously.  Furthermore, the two
inteins interact differently with DNA.  In addition, we did not detect any endonuclease
activity for PabLon and PabRIR1-1.  deletions in the intein sequences and mutations in the
putative endonuclease motifs probably abolish this activity.  Hence, inteins from the same
archaebacteria, even if contained in the same host protein, did not evolve uniformly and are
presumably at different stages of the invasion cycle.

<>

<1>Saves, I., Ozanne, V., Dietrich, J., Masson, J.M.
<2>Inteins of Thermococcus fumicolans DNA Polymerase Are Endonucleases with Distinct Enzymatic Behaviors.
<3>J. Biol. Chem.
<4>275
<5>2335-2341
<6>2000
<7>The DNA polymerase gene of Thermococcus fumicolans harbors two intein genes. Both inteins have
been produced in Escherichia coli and purified either as naturally spliced products from the
expression of the complete DNA polymerase gene or directly from the cloned inteins genes. Both
recombinant inteins exhibit endonuclease activity, with an optimal temperature of 70 degrees
C. The Tfu pol-1 intein, which belongs to the Psp KOD pol-1 allelic family, recognizes and
cleaves a minimal sequence of 16 base pairs (bp) on supercoiled DNA with either Mn(2+) or
Mg(2+) as cofactor. It cleaves linear DNA only with Mn(2+) and requires a 19-bp minimal
recognition sequence. The Tfu pol-2 intein, which belongs to the Tli pol-2 allelic family, is
a highly active homing endonuclease using Mg(2+) as cofactor. Its minimal recognition and
cleavage site is 21 bp long either on linear or circular DNA substrates. Its endonuclease
activity is strongly inhibited by the 3' digestion product, which remains bound to the enzyme
after the cleavage reaction. According to current nomenclature, these endonucleases were named
PI-TfuI and PI-TfuII. These two inteins thus exhibit different requirements for metal cofactor
and substrate topology as well as different mechanism of action.

<>

<1>Saves, I., Westrelin, F., Daffe, M., Masson, J.-M.
<2>Identification of the first eubacterial endonuclease coded by an intein allele in the pps1 gene of mycobacteria.
<3>Nucleic Acids Res.
<4>29
<5>4310-4318
<6>2001
<7>A survey of a vast range of mycobacterial strains led us to discover a new Pps1 intein allele
in Mycobacterium gastri which differs from those of Mycobacterium tuberculosis and
Mycobacterium leprae in both its sequence and insertion site. While little is known about
Pps1, except that it belongs to the YC24 family of ABC transporters, we show that, unlike the
other inteins described so far from Eubacteria, the MgaPps1 intein possesses a specific
endonuclease activity. The intein is the first eubacterial intein to be characterised as an
endonuclease. Like other intein endonucleases, its minimal sequence for recognition and
cleavage is quite large, with 22 bp spanning the Pps1-c site. The fact that an active
endonuclease is found among the mycobacterial inteins supports the concept of a cyclical model
of invasion by horizontal transfer of these genes, followed by degeneration and loss until a
new invasion event, thus explaining their long-term persistence in closely related eubacterial
species.

<>

<1>Savin, C., Frangeul, L., Ma, L., Bouchier, C., Moszer, I., Carniel, E.
<2>Draft Genome Sequence of a Clinical Strain of Yersinia enterocolitica (IP10393) of Bioserotype 4/O:3 from France.
<3>Genome Announcements
<4>1
<5>e00150-12
<6>2013
<7>We sequenced the genome of a clinical isolate of (IP10393) from France. This strain belongs to
bioserotype 4/O:3, which is the most common pathogenic subgroup
worldwide. The draft genome has a size of 4,463,212 bp and a G+C content of
47.0%, and it is predicted to contain 4,181 coding sequences.

<>

<1>Saw, J.H., Yuryev, A., Kanbe, M., Hou, S., Young, A.G., Aizawa, S., Alam, M.
<2>Complete genome sequencing and analysis of Saprospira grandis str. Lewin, a predatory marine bacterium.
<3>Standards in Genomic Sciences
<4>6
<5>84-93
<6>2012
<7>Saprospira grandis is a coastal marine bacterium that can capture and prey upon other marine
bacteria using a mechanism known as 'ixotrophy'. Here, we present
the complete genome sequence of Saprospira grandis str. Lewin isolated from La
Jolla beach in San Diego, California. The complete genome sequence comprises a
chromosome of 4.35 Mbp and a plasmid of 54.9 Kbp. Genome analysis revealed
incomplete pathways for the biosynthesis of nine essential amino acids but
presence of a large number of peptidases. The genome encodes multiple copies of
sensor globin-coupled rsbR genes thought to be essential for stress response and
the presence of such sensor globins in Bacteroidetes is unprecedented. A total of
429 spacer sequences within the three CRISPR repeat regions were identified in
the genome and this number is the largest among all the Bacteroidetes sequenced
to date.

<>

<1>Saw, J.H.W., Schatz, M., Brown, M.V., Kunkel, D.D., Foster, J.S., Shick, H., Christensen, S., Hou, S., Wan, X., Donachie, S.P.
<2>Cultivation and Complete Genome Sequencing of Gloeobacter kilaueensis sp. nov., from a Lava Cave in Kilauea Caldera, Hawai'i.
<3>PLoS ONE
<4>8
<5>e76376
<6>2013
<7>The ancestor of Gloeobacter violaceus PCC 7421T is believed to have diverged from that of all
known cyanobacteria before
the evolution of thylakoid membranes and plant plastids. The long and largely independent
evolutionary history of G.
violaceus presents an organism retaining ancestral features of early oxygenic photoautotrophs,
and in whom cyanobacteria
evolution can be investigated. No other Gloeobacter species has been described since the genus
was established in 1974
(Rippka et al., Arch Microbiol 100:435). Gloeobacter affiliated ribosomal gene sequences have
been reported in
environmental DNA libraries, but only the type strain's genome has been sequenced. However,
we report here the
cultivation of a new Gloeobacter species, G. kilaueensis JS1T, from an epilithic biofilm in a
lava cave in Ký'lauea Caldera,
Hawai'i. The strain's genome was sequenced from an enriched culture resembling a
low-complexity metagenomic sample,
using 9 kb paired-end 454 pyrosequences and 400 bp paired-end Illumina reads. The JS1T and G.
violaceus PCC 7421T
genomes have little gene synteny despite sharing 2842 orthologous genes; comparing the genomes
shows they do not
belong to the same species. Our results support establishing a new species to accommodate
JS1T, for which we propose the
name Gloeobacter kilaueensis sp. nov. Strain JS1T has been deposited in the American Type
Culture Collection (BAA-2537),
the Scottish Marine Institute's Culture Collection of Algae and Protozoa (CCAP 1431/1), and
the Belgian Coordinated
Collections of Microorganisms (ULC0316). The G. kilaueensis holotype has been deposited in the
Algal Collection of the US
National Herbarium (US# 217948). The JS1T genome sequence has been deposited in GenBank under
accession number
CP003587. The G+C content of the genome is 60.54 mol%. The complete genome sequence of G.
kilaueensis JS1T may
further understanding of cyanobacteria evolution, and the shift from anoxygenic to oxygenic
photosynthesis.

<>

<1>Sawaragi, H., Yokoyama, S., Nomura, Y., Kimizuka, F., Nakajima, K.
<2>Stabilized restriction endonuclease composition - used in genetic engineering.
<3>Japanese Patent Office
<4>JP 6343468
<5>
<6>1994
<7>
<>

<1>Sawaya, M.R., Zhu, Z., Mersha, F., Chan, S.-h., Dabur, R., Xu, S.-y., Balendiran, G.K.
<2>Crystal structure of the restriction-modification system control element C.BclI and mapping of its binding site.
<3>Structure
<4>13
<5>1837-1847
<6>2005
<7>Protection from DNA invasion is afforded by restriction-modification systems in many bacteria.
The efficiency of protection depends crucially on the relative expression levels of
restriction versus methyltransferase genes. This regulation is provided by a controller
protein, named C protein. Studies of the BclI system in E. coli suggest that C.BclI functions
as a negative regulator for M.BclI expression, implying that it plays a role in defense
against foreign DNA during virus infection. C.BclI binds (Kd = 14.3 nM) to a 2-fold symmetric
C box DNA sequence that overlaps with the putative END cut site unknown.

<>

<1>Saxena, A., Kumari, R., Mukherjee, U., Singh, P., Lal, R.
<2>Draft Genome Sequence of the Rifamycin Producer Amycolatopsis rifamycinica DSM 46095.
<3>Genome Announcements
<4>2
<5>e00662-14
<6>2014
<7>Amycolatopsis rifamycinica DSM 46095 is an actinobacterium that produces rifamycin SV, an
antibiotic used against Mycobacterium tuberculosis. Here, we
present the draft genome of DSM 46095, which harbors a novel rifamycin polyketide
biosynthetic gene cluster (rif PKS) that differed by 10% in nucleotide sequence
from the already reported rif PKS cluster of Amycolatopsis mediterranei S699.

<>

<1>Saxena, A., Nayyar, N., Sangwan, N., Kumari, R., Khurana, J.P., Lal, R.
<2>Genome Sequence of Novosphingobium lindaniclasticum LE124T, Isolated from a Hexachlorocyclohexane Dumpsite.
<3>Genome Announcements
<4>1
<5>e00715-13
<6>2013
<7>Novosphingobium lindaniclasticum LE124(T) is a hexachlorocyclohexane (HCH)-degrading bacterium
isolated from a high-dosage-point HCH dumpsite (450 mg
HCH/g soil) located in Lucknow, India (27 degrees 00'N and 81 degrees 09'E).
Here, we present the annotated draft genome sequence of strain LE124(T), which
has an estimated size of 4.86 Mb and is comprised of 4,566 coding sequences.

<>

<1>Saxena, D., Caufield, P.W., Li, Y., Brown, S., Song, J., Norman, R.
<2>Genetic Classification of Severe Early Childhood Caries Using Subtracted DNA Fragments from Streptococcus mutans.
<3>J. Clin. Microbiol.
<4>46
<5>2868-2873
<6>2008
<7>Streptococcus mutans is one of several members of the oral indigenous
biota linked with severe early childhood caries (S-ECC). Because most
humans harbor S. mutans, but not all manifest disease, it has been
proposed that the strains of S. mutans associated with S-ECC are
genetically distinct from those found in caries-free (CF) children. The
objective of this study was to identify common DNA fragments from S.
mutans present in S-ECC but not in CF children. Using suppressive
subtractive hybridization, we found a number of DNA fragments (biomarkers)
present in 88 to 95% of the S-ECC S. mutans strains but not in CF S.
mutans strains. We then applied machine learning techniques including
support vector machines and neural networks to identify the biomarkers
with the most predictive power for disease status, achieving a 92%
accurate classification of the strains as either S-ECC or CF associated.
The presence of these gene fragments in 90 to 100% of the 26 S-ECC
isolates tested suggested their possible functional role in the
pathogenesis of S. mutans associated with dental caries.

<>

<1>Sayers, J.R., Olsen, D.B., Eckstein, F.
<2>Inhibition of restriction endonuclease hydrolysis by phosphorothioate-containing DNA.
<3>Nucleic Acids Res.
<4>17
<5>9495
<6>1989
<7>None

<>

<1>Sayers, J.R., Schmidt, W., Wendeler, A., Eckstein, F.
<2>Strand specific cleavage of phosphorothioate-containing DNA by reaction with restriction endonucleases in the presence of ethidium bromide.
<3>Nucleic Acids Res.
<4>16
<5>803-813
<6>1988
<7>A method for achieving strand specific nicking of DNA has been developed.
Phosphorothioate groups were incorporated enzymatically into the (-) strand of
M13 RF IV DNA.  When such DNA is reacted with restriction endonucleases in the
presence of ethidium bromide, nicked DNA (RF II) is produced.  All of the
restriction enzymes tested linearised phosphorothioate-containing DNA in the
absence of this dye.  The strand specificity of the reaction was investigated
by employing the ethidium bromide mediated nicking reaction in the
phosphorothioate-based oligonucleotide-directed mutagenesis method.  The
mutational efficiences obtained were in the region of 64-89%, indicating that
these restriction enzymes hydrolyse the phosphodiester bond at the cleavage
site of the unsubstituted (+) strand.

<>

<1>Scalley-Kim, M., McConnell-Smith, A., Stoddard, B.L.
<2>Coevolution of a Homing Endonuclease and Its Host Target Sequence.
<3>J. Mol. Biol.
<4>372
<5>1305-1319
<6>2007
<7>We have determined the specificity profile of the homing endonuclease I-AniI and compared it
to the conservation of its host gene. Homing
endonucleases are encoded within intervening sequences such as group I
introns. They initiate the transfer of such elements by cleaving cognate
alleles lacking the intron, leading to their transfer via homologous
recombination. Each structural homing endonuclease family has arrived at
an appropriate balance of specificity and fidelity that avoids toxicity
while maximizing target recognition and invasiveness. I-AniI recognizes a
strongly conserved target sequence in a host gene encoding apocytochrome B
and has fine-tuned its specificity to correlate with wobble versus
nonwobble positions across that sequence and to the amount of degeneracy
inherent in individual codons. The physiological target site in the host
gene is not the optimal substrate for recognition and cleavage: at least
one target variant identified during a screen is bound more tightly and
cleaved more rapidly. This is a result of the periodic cycle of intron
homing, which at any time can present nonoptimal combinations of
endonuclease specificity and insertion site sequences in a biological
host.

<>

<1>Scarpelis, G., Moissidou, A., Rina, M., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BshGI.
<3>Nucleic Acids Res.
<4>17
<5>8883
<6>1989
<7>None

<>

<1>Scarselli, M., Masignani, V., Mora, M.
<2>STAPHYLOCOCCUS AUREUS PROTEINS AND NUCLEIC ACIDS.
<3>Japanese Patent Office
<4>JP 2005502326 A
<5>
<6>2005
<7>
<>

<1>Scavetta, R.D., Thomas, C.B., Walsh, M.A., Szegedi, S., Joachimiak, A., Gumport, R.I., Churchill, M.E.A.
<2>Structure of RsrI methyltransferase, a member of the N6-adenine beta class of DNA methyltransferases.
<3>Nucleic Acids Res.
<4>28
<5>3950-3961
<6>2000
<7>DNA methylation is important in cellular, developmental and disease processes, as well as in
bacterial restriction-modification systems. Methylation of DNA at the amino groups of cytosine
and adenine is a common mode of protection against restriction endonucleases afforded by the
bacterial methyltransferases.  The first structure of an N6-adenine methyltransferase
belonging to the beta class of bacterial methyltransferases is described here. The structure
of M.RsrI from Rhodobacter sphaeroides, which methylates the second adenine of the GAATTC
sequence, was determined to 1.75 A resolution using X-ray crystallography. Like other
methyltransferases, the enzyme contains the methylase fold and has well-defined substrate
binding pockets. The catalytic core most closely resembles the PvuII methyltransferase, a
cytosine amino methyltransferase of the same beta group. The larger nucleotide binding pocket
observed in M.RsrI is expected because it methylates adenine. However, the most striking
difference between the RsrI methyltransferase and the other bacterial enzymes is the structure
of the putative DNA target recognition domain, which is formed in part by two helices on an
extended arm of the protein on the face of the enzyme opposite the active site. This
observation suggests that a dramatic conformational change or oligomerization may take place
during DNA binding and methylation.

<>

<1>Schaefer, B., Wilde, B., Massardo, D.R., Manna, F., Del Giudice, L., Wolf, K.
<2>A mitochondrial group-I intron in fission yeast encodes a maturase and is mobile in crosses.
<3>Curr. Genet.
<4>25
<5>336-341
<6>1994
<7>The open reading frame in the first intron of the mitochondrial gene encoding subunit I of
cytochrome c oxidase encodes a maturase and stimulates homologous recombination in Escherichia
coli.  In this paper, we demonstrate that this intron is mobile in crosses, indicating that it
also encodes an endonuclease.  This is the first report on an intron which possesses mobility
and acts as a maturase.

<>

<1>Schaeffer, C.J., Huang, B., Tsai, M.-D.
<2>A new class IIS zinc finger restriction enzyme with specificity for SP1 binding sites.
<3>FASEB J.
<4>10
<5>A1241
<6>1996
<7>A new restriction endonuclease (Sp1ase) was constructed by fusing the DNA-cleavage domain of
the restriction endonuclease FokI in frame with the zinc-finger DNA-binding domain of the
transcription factor Sp1.  This construct was shown to selectively digest plasmid DNA carrying
consensus Sp1 sites.  Splase was also shown to selectively digest plasmid DNA carrying the
HIV-1 LTR.  Sp1ase is a genetically engineered restriction enzyme conveying specificity for
Sp1 binding sites.  The site-specific phosphodiesterase activity of Sp1ase has been
characterized and shown to have more specific cleavage of one strand of DNA than the other.
Sp1ase recognizes a ten base-pair DNA sequence and hydrolyzes phosphodiester bonds upstream of
the binding sequence.  The binding specificity of Sp1ase makes this a rare cutter
restriction enzyme which could be valuable in creating large DNA fragments for genome
sequencing projects.  This result presents the opportunity to creat other restriction enzymes
by altering the binding specificity of the zinc finger recognition helix.

<>

<1>Schaefler, S.
<2>Staphylococcus epidermidis BV:  Antibiotic resistance patterns, physiological characteristics, and bacteriophage susceptibility.
<3>Appl. Microbiol.
<4>22
<5>693-699
<6>1971
<7>Staphylococcus epidermidis BV is a group of mannitol-fermenting
coagulase-negative staphylococci characterized by multiple antibiotic
resistance, very similar biochemical characteristics, and phage susceptibility.
Clinical isolates belonging to this group are resistant to most antibiotics
tested, including oxacillin, lincomycin, and novobiocin.  The only antibiotic
to which all tested strains are sensitive is vancomycin.  Common biochemical
traits of the tested S. epidermidis BV strains include fermentation of
trehalose and ribose, phospho-b-glucosidase activity, growth on synthetic
medium with amino acids as carbon source, and lack of deoxyribonuclease,
phosphatase, lipase, and gelatinase activity.  Some of these characteristics
appear more frequently in mannitol-positive control strains than in
mannitol-negative strains.  S. epidermidis  BV strains carry lysogenic phages
with a host range restricted to this group.  These phages allow the
differentiation of individual strains.

<>

<1>Schafer, A., Schwarzer, A., Kalinowski, J., Puhler, A.
<2>Cloning and characterization of a DNA region encoding a stress-sensitive restriction system from Corynebacterium glutamicum ATCC 13032 and analysis of its role in intergeneric conjugation with Escherichia coli.
<3>J. Bacteriol.
<4>176
<5>7309-7319
<6>1994
<7>RP4-mediated transfer of mobilizable plasmids in intergeneric conjugation of Escherichia coli
donors with Corynebacterium glutamicum ATCC 13032 is severely affected by a restriction system
in the recipient that can be inactivated by a variety of exogenous stress factors. In this
study a rapid test procedure based on intergeneric conjugal plasmid transfer that permitted
the distinction between restriction-negative and restriction-positive C. glutamicum clones was
developed. By using this procedure, clones of the restriction-deficient mutant strain C.
glutamicum RM3 harboring a plasmid library of the wild-type chromosome were checked for their
restriction properties. A complemented clone with a restriction-positive phenotype was
isolated and found to contain a plasmid with a 7-kb insertion originating from the wild-type
chromosome. This plasmid, termed pRES806, is able to complement the restriction-deficient
phenotype of different C. glutamicum mutants. Sequence analysis revealed the presence of two
open reading frames (orf1 and orf2) on the complementing DNA fragment. The region comprising
orf1 and orf2 displayed a strikingly low G+C content and was present exclusively in C.
glutamicum strains. Gene disruption experiments with the wild type proved that orf1 is
essential for complementation, but inactivation of orf2 also resulted in a small but
significant increase in fertility. These results were confirmed by infection assays with the
bacteriophage CL31 from Corynebacterium lilium ATCC 15990.

<>

<1>Schafer, A., Tauch, A., Droste, N., Puhler, A., Kalinowski, J.
<2>The Corynebacterium glutamicum cglIM gene encoding a 5-cytosine methyltransferase enzyme confers a specific DNA methylation pattern in an McrBC-deficient Escherichia coli strain.
<3>Gene
<4>203
<5>95-101
<6>1997
<7>The cglIM gene of the coryneform soil bacterium Corynebacterium glutamicum ATCC 13032 has been
cloned and characterized.  The coding region comprises 1092 nucleotides and specifies a
protein of 363 amino acid residues with a deduced Mr of 40,700.  The amino acid sequence
showed striking similarities to methyltransferase enzymes generating 5-methylcytosine
residues, especially to M.NgoVII from Neisseria gonorrhoeae recognizing the sequence GCSGC.
The cglIM gene is organized in an unusual operon which contains, in addition, two genes
encoding stress-sensitive restriction enzymes.  Using PCR techniques the entire gene including
the promoter region was amplified from the wild-type chromosome and cloned in Escherichia
coli.  Expression of the cglIM gene in E. coli under the control of its own promoter conferred
the C. glutamicum-specific methylation pattern to co-resident shuttle plasmids and led to a
260-fold increase in the transformation rate of C. glutamicum.  In addition, the methylation
pattern produced by this methyltransferase enzyme is responsible for the sensitivity of DNA
from C. glutamicum to the modified cytosine restriction system of E. coli.

<>

<1>Schafer, B., Kaulich, K., Wolf, K.
<2>Mosaic structure of the cox2 gene in the petite negative yeast Schizosaccharomyces pombe: a group II intron is inserted at the same location as the otherwise unrelated group II introns in the mitochondria of higher plants.
<3>Gene
<4>214
<5>101-112
<6>1998
<7>In contrast to homologous genes in other fungal mitochondrial genomes, the gene encoding
subunit 2 of cytochrome oxidase (cox2) in several Schizosaccharomyces pombe strains contains a
large group II intron. Its 2436 nucleotides can be folded into a typical group II intron
secondary structure, possessing all the expected sequence motifs for subgroup IIA1 (Michel et
al., 1989).  This intron is remarkable for the following reasons: (i) Five nucleotide changes
were observed compared with the continuous form of the cox2 gene in the reference strain 50 at
the 3'-exon sequence, but not in the 5'-exon. (ii) One of these changes occurred at the
splice point leading to a serine instead of a threonine residue in the deduced cox2
polypeptide. In all cases, the alterations resulted in the replacement of more frequently used
codons by rare ones. (iii) Although the intron is able to undergo splicing, the sequence
motifs thought to be necessary for interaction between the 5'-exon and the intron during the
splicing process (the EBS1/IBS1 as well as the EBS2/IBS2 pairings) are unusual. (iv) The
intron is inserted at the same location in the cox2 gene as the otherwise unrelated intron
from higher plants.

<>

<1>Schaffert, L., Albersmeier, A., Bednarz, H., Niehaus, K., Kalinowski, J., Ruckert, C.
<2>Genome sequence of the marine bacterium Corynebacterium maris type strain Coryn-1(T) (= DSM 45190(T)).
<3>Standards in Genomic Sciences
<4>8
<5>516-524
<6>2013
<7>Corynebacterium maris Coryn-1(T) Ben-Dov et al. 2009 is a member of the genus Corynebacterium
which contains Gram-positive, non-spore forming bacteria with a
high G+C content. C. maris was isolated from the mucus of the Scleractinian coral
Fungia granulosa and belongs to the aerobic and non-haemolytic corynebacteria. It
displays tolerance to salts (up to 10%) and is related to the soil bacterium
Corynebacterium halotolerans. As this is a type strain in a subgroup of
Corynebacterium without complete genome sequences, this project, describing the
2.78 Mbp long chromosome and the 45.97 kbp plasmid pCmaris1, with their 2,584
protein-coding and 67 RNA genes, will aid the G enomic E ncyclopedia of Bacteria
and Archaea project.

<>

<1>Schapira, M., Desdouets, C., Jacq, C., Perea, J.
<2>I-SceIII an intron-encoded DNA endonuclease from yeast mitochondria. Asymmetrical DNA binding properties and cleavage reaction.
<3>Nucleic Acids Res.
<4>21
<5>3683-3689
<6>1993
<7>We have previously discovered the new intron-encoded endonuclease I-SceIII by expressing, in
E.coli, the ORF contained in the third intron of the yeast mitochondrial COX I gene. In this
work, we analyzed the in vitro properties of partially purified I-SceIII and found that it is
a very specific DNA endonuclease, tolerating relatively few base changes in its 20 base pair
long target site. I-SceIII should be a useful molecular tool to analyze the structure of large
genomes. Interestingly, I-SceIII is the first P1-P2 DNA endonuclease for which DNA binding
properties could be analyzed by band-shift experiments. Clearly, the cleavage products
corresponding to the upstream A3 exon and to the downstream A4 exon could compete with the
substrate A3-A4 in forming a DNA-protein complex. However, the A3 exon competes more
efficiently than the downstream A4 product. The cleavage of the two DNA strands is also
asymmetric; the top strand (non-transcribed strand) is cleaved faster than the bottom strand,
a property found under various experimental conditions. These findings sugest that this
intron-encoded DNA endonuclease may have a role in the RNA splicing process of the intron.

<>

<1>Scharnagl, M., Richter, S., Hagemann, M.
<2>The cyanobacterium Synechocystis sp. strain PCC 6803 expresses a DNA methyltransferase specific for the recognition sequence of the restriction endonuclease PvuI.
<3>J. Bacteriol.
<4>180
<5>4116-4122
<6>1998
<7>By use of restriction endonucleases, the DNA of the cyanobacterium Synechocystis sp. strain
PCC 6803 was analyzed for DNA-specific methylation.  Three different recognition sites of
methyltransferases, a dam-like site including N6-methyladenosine and two other sites with
methylcytosine, were identified, whereas no activities of restriction endonucleases could be
detected in this strain.  Slr0214, a Synechocystis gene encoding a putative methyltransferase
that shows significant similarities to C5-methylcytosine-synthesizing enzymes, was amplified
by PCR and cloned for further characterization.  Mutations in slr0214 were generated by the
insertion of an aphII gene cassette.  Analyses of chromosomal DNAs of such mutants
demonstrated that the methylation pattern was changed.  The recognition sequence of the
methyltransferase was identified as 5'-CGATCG-3', corresponding to the recognition sequence
of PvuI.  The specific methyltransferase activity was significantly reduced in protein
extracts obtained from mutant cells.  Mutation of slr0214 also led to changed growth
characteristics of the cells compared to wild-type cells.  These alterations led to the
conclusion that the methyltransferase Slr0214 might play a regulatory role in Synechocystis.
The Slr0214 protein was also overexpressed in Escherichia coli, and the purified protein
demonstrated methyltransferase activity and specificity for PvuI recognition sequences in
vitro.  We propose the designation SynMI (Synechocystis methyltransferase I) for the
slr0214-encoded enzyme.

<>

<1>Scheidt, G., Graessmann, A., Weber, H.
<2>Cloning of the methyltransferase gene from Arabidopsis thaliana.
<3>Biol. Chem. Hoppe Seyler
<4>372
<5>742-743
<6>1991
<7>DNA methylation is a widespread modification in eucaryotic cells and thought to be envolved in
gene regulation.  Recently we showed that in vitro methylation of transgenes was inherited in
tobacco plants and led to gene inactivation.  In plants DNA methylation differs from that in
animal cells in respect to a much higher amount of 5-methylcytosine content of the DNA and to
an altered sequence specificity of the methyltransferase (Mtase).  Cytosines in CpG and CpNpG
sequences were methylated in plant DNA whereas only 5mCpG can be found in animal DNA.
Furthermore it was shown that wheat Mtase had a preference for endogenous double stranded DNA
as substrate and a lower molecular mass distinguishes it from the mammalian enzyme.  Little is
known about the structure and regulation of eucaryotic Mtases, especially from plants and only
from mouse cells the corresponding cDNA had been cloned and sequenced.  To characterize the
Mtase gene from Arabidopsis a genomic library from nuclear DNA was established in EMBL3 and
screened with heterologous probes encoding the cDNA of mouse Mtase (gift from T. Bestor).
Several clones which hybridized under low stringent conditions were isolated and part of the
DNA sequence was determined.  We found significant homology to the mouse Mtase cDNA on
nucleotide and on amino acid level.  Furthermore this region is part of a conserved domain of
cytosine Mtases not only from eucaryotic but also from bacterial and phage origin.  Using the
cloned fragments as probes for hybridization experiments to each other and to total
Arabidopsis DNA we identified at least 3 different sequences which are related but not totally
homologous.  However they all hybridize to the mouse Mtase cDNA.  We conclude therefore that
Arabidopsis Mtase is encoded by a gene family.

<>

<1>Scheidt, G., Weber, H., Graessmann, M., Graessmann, A.
<2>Are there two DNA methyltransferase gene families in plant cells?  A new potential methyltransferase gene isolated from an Arabidopsis thaliana genomic library.
<3>Nucleic Acids Res.
<4>22
<5>953-958
<6>1994
<7>Using the 1kb 3' terminal DNA fragment of the mouse methyltransferase cDNA as a probe and low
stringent hybridisation conditions, a new potential methyltransferase (MTase) gene family was
isolated from an Arabidopsis thaliana genomic DNA library. One clone (MTase-11), which gave
the strongest signal at the Northern blot, was entirely sequenced (11483 bp) and further
characterised. Under consideration of the likely open reading frames and our preliminary cDNA
experiments we propose that the clone 11 gene encodes for an ~90 kD protein. As deduced from
the DNA sequence this protein contains all conserved sequence motifs specific for the 5m
cytosine MTases. MTase-11 gene expression was demonstrable in callus and during germination
but not in one month old plants or in leaves.

<>

<1>Scheinbart, L.S., Johnson, M.A., Gross, L.A., Edelstein, S.R., Richardson, B.C.
<2>Procainamide inhibits DNA methyltransferase in a human T cell line.
<3>J. Rheumatol.
<4>18
<5>530-534
<6>1991
<7>Procainamide, a widely used antiarrythmic, causes DNA hypomethylation in the
human T cell line Jurkat, but the mechanism is unknown.  We report that
procainamide inhibits the DNA methyltransferase catalyzed transfer of methyl
groups from S-adenosylmethionine to DNA, but has no effect on other known
regulators of DNA methylation.  Our results suggest that procainamide could
inhibit cellular DNA methylation by inhibiting DNA methyltransferase activity.

<>

<1>Schell, J.
<2>The mechanism of restriction of bacteriophage lambda in Escherichia coli strains: Demonstration of an in vivo requirement for S-adenosylmethionine.
<3>Virology
<4>39
<5>66-73
<6>1969
<7>Restriction of phage lambda by certain Escherichia coli strains involves the degradation of
the phage DNA in strains with a host-controlled modification system different from the one
carried by the restricted phage.  By infecting the restricting host strain with phage T3,
prior to infection with phage lambda, it was possible to inactivate the capacity of host cells
to restrict phage lambda.  We have shown that this inactivation is specifically due to the
production, in these T3-infected host cells, of an enzyme which cleaves S-adenosylmethionine.
Since SAM is necessary for methylation reactions, it follows from these observations that
methylation of the restricted lambda DNA might be required to make it susceptible to the
host-controlled restriction system.  Several E. coli strains have a host-controlled
restriction system, e.g., E. coli K12 and E. coli B, but also prophage P1 controls such a
system.  Our results show that both the E. coli K12 and B restriction systems require SAM for
activity.  The P1 restriction system, however, does not appear to have such a requirement.

<>

<1>Schell, J., Glover, S.W.
<2>The effect of heat on host-controlled restriction of phage lambda in Escherichia coli K(P1).
<3>J. Gen. Microbiol.
<4>45
<5>61-72
<6>1966
<7>The growth of phage lambda.C (i.e. phage lambda grown in Escherichia coli C) in
E. coli K lysogenized by phage P1 is normally restricted so that the efficiency
of plating of phage lambda.C on K(P1) compared to C is about 10-7.  When K(P1)
bacteria are heated before infection with phage lambda.C this restriction may
be decreased as much as a million-fold.  The time of exposure to elevated
temperature (49C and above) required to achieve this increase in e.o.p. of
phage lambda.C decreased with increasing temperature up to temperatures which
began to inhibit the capacity of bacteria to grow phage.  Heated K(P1) bacteria
recovered their ability to restrict phage lambda.C following the resumption of
growth at 37C.  Part of this recovery can be inhibited by chloramphenicol.  A
more dramatic recovery of restriction is observed when heated K(P1) bacteria
are resuspended in hypertonic media.  Experiments are described which indicate
that phage lambda.C is restricted at an early step after adsorption and that,
if phage escapes this restriction, it can grow in heated bacteria subsequently
converted into restricting hosts by resuspension in hypertonic media.

<>

<1>Schell, J., Glover, S.W.
<2>The effect of various physiological conditions on host-controlled restriction in Escherichia coli K (P1).
<3>Genet. Res.
<4>7
<5>273-276
<6>1966
<7>Growth of K(P1) bacteria under conditions which lead to a reduction in the level of nucleases
also leads to a reduction in their ability to restrict the growth of lambda.C.  Experiments
designed to estimate the time after adsorption at which restriction takes place indicate that
phage DNA is probably restricted by a nuclease while passing through the periplasm.

<>

<1>Schell, J., Glover, S.W.
<2>On the localization of a factor responsible for host-controlled restriction in Escherichia coli K(P1).
<3>Genet. Res.
<4>7
<5>277-279
<6>1966
<7>Recent experiments indicate that an essential step in the restriction of phage lambda.C by E.
coli K(P1) bacteria may involve an enzyme located on the surface of the cells, and a
surface-localized nuclease has been implicated in the restriction of non-glucosylated T4 DNA
by E. coli B.  Neu & Heppel have shown that several enzymes, alkaline phosphatase, latent
RNase, 5'-nucleotidase, acid phosphatase, cyclic phosphodiesterase and an RNA inhibited DNase
can be partly or completely released by E. coli cells during spheroplast formation.  These
authors also reported that surface-localized enzymes can also be released by EDTA treatment
followed by washing at 4 C.  The results reported in this paper show that the restriction of
phage lambda.C by K(P1) cells is markedly reduced when the cells are treated with EDTA and
washed several times with cold distilled water.

<>

<1>Schell, J., Glover, S.W., Stacey, K.A., Broda, P.M.A., Symonds, N.
<2>The restriction of phage T3 by certain strains of Escherichia coli.
<3>Genet. Res.
<4>4
<5>483-484
<6>1963
<7>The efficiency of plating (e.o.p.) of phage T3 is equal on the E. coli strains
B and C, and also on most F- strains of K12 (afterwards called K).  However, in
strains of K which carry the F episome its e.o.p. is greatly reduced.  On most
F+ strains of K its e.o.p. is about 10-5.  The phages which do multiply in the
F+ bacteria are not modified; they still have an e.o.p. of 10-5 when replated
on the same host.  The actual fraction of cells which accept T3 phages is
approximately 10-2, and these cells yield a small burst of unmodified T3.  It
seems to be this combination of restriction coupled with a small burst which
defines the level of the e.o.p. and produces the typical small plaques which
are observed on F+ strains.  As a result of screening a large number of male
and female strains of K two striking exceptions were found to the observation
stated above.  The F+ strain W1485 was found to plate efficiently, while some
F- derivatives of 58-161 plate T3 with a low e.o.p.  In an effort to elucidate
this situation a series of conjugation experiments was performed to establish
the relationship between F and T3 restriction.  The results of these
experiments are presented in Table 1.

<>

<1>Schell, M.A., Karmirantzou, M., Snel, B., Vilanova, D., Berger, B., Pessi, G., Zwahlen, M.C., Desiere, F., Bork, P., Delley, M., Prodmore, R.D., Arigoni, F.
<2>The genome sequence of Bifidobacterium longum reflects its adaptation to the human gastrointestinal tract.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>14422-14427
<6>2002
<7>Bifidobacteria are Gram-positive prokaryotes that naturally colonize the human
gastrointestinal tract (GIT) and vagina. Although not numerically dominant in the complex
intestinal microflora, they are considered as key commensals that promote a healthy GIT. We
determined the 2.26-Mb genome sequence of an infant-derived strain of Bifidobacterium longum,
and identified 1,730 possible coding sequences organized in a 60%-GC circular chromosome.
Bioinformatic analysis revealed several physiological traits that could partially explain the
successful adaptation of this bacteria to the colon. An unexpectedly large number of the
predicted proteins appeared to be specialized for catabolism of a variety of oligosaccharides,
some possibly released by rare or novel glycosyl hydrolases acting on "nondigestible" plant
polymers or host-derived glycoproteins and glycoconjugates. This ability to scavenge from a
large variety of nutrients likely contributes to the competitiveness and persistence of
bifidobacteria in the colon. Many genes for oligosaccharide metabolism were found in
self-regulated modules that appear to have arisen in part from gene duplication or horizontal
acquisition. Complete pathways for all amino acids, nucleotides, and some key vitamins were
identified; however, routes for Asp and Cys were atypical. More importantly, genome analysis
provided insights into the reciprocal interactions of bifidobacteria with their hosts. We
identified polypeptides that showed homology to most major proteins needed for production of
glycoprotein-binding fimbriae, structures that could possibly be important for adhesion and
persistence in the GIT. We also found a eukaryotic-type serine protease inhibitor (serpin)
possibly involved in the reported immunomodulatory activity of bifidobacteria.

<>

<1>Schenk, J.A., Heymann, S., Micheel, B.
<2>The accessibility of thiophosphorylated groups in DNA fragments to the enzymatic activity of ligases and restriction endonuclease BbsI.
<3>Biochem. Mol. Biol. Int.
<4>36
<5>1037-1043
<6>1995
<7>The aim of this paper was to test the possibility to ligate and hydrolyse DNA
sequences containing thiomodified ends and bonds.  T4 DNA ligase was shown to ligate DNA
fragments regardless of whether it contains phosphorylated or thiophosphorylated 5'-ends.  But
the cleavage of an internally thiomodified phosphodiester bond was found to be totally
inhibited
when using the nonpalindromic restrictase BbsI.  The special properties of this restriction
endonuclease should allow the development of an oriented cloning strategy when combined with
T4 ligase and a thiophosphorylation of DNA fragments.

<>

<1>Schermelleh, L., Haemmer, A., Spada, F., Rosing, N., Meilinger, D., Rothbauer, U., Cristina, C.M., Leonhardt, H.
<2>Dynamics of Dnmt1 interaction with the replication machinery and its role in postreplicative maintenance of DNA methylation.
<3>Nucleic Acids Res.
<4>35
<5>4301-4312
<6>2007
<7>Postreplicative maintenance of genomic methylation patterns was proposed to depend largely on
the binding of DNA methyltransferase 1 (Dnmt1) to PCNA, a core component of the replication
machinery. We investigated how
the slow and discontinuous DNA methylation could be mechanistically linked
with fast and processive DNA replication. Using photobleaching and
quantitative live cell imaging we show that Dnmt1 binding to PCNA is
highly dynamic. Activity measurements of a PCNA-binding-deficient mutant
with an enzyme-trapping assay in living cells showed that this interaction
accounts for a 2-fold increase in methylation efficiency. Expression of
this mutant in mouse dnmt1(-/-) embryonic stem (ES) cells restored CpG
island methylation. Thus association of Dnmt1 with the replication
machinery enhances methylation efficiency, but is not strictly required
for maintaining global methylation. The transient nature of this
interaction accommodates the different kinetics of DNA replication and
methylation while contributing to faithful propagation of epigenetic
information.

<>

<1>Schermelleh, L., Spada, F., Easwaran, H.P., Zolghadr, K., Margot, J.B., Cardoso, M.C., Leonhardt, H.
<2>Trapped in action: direct visualization of DNA methyltransferase activity in living cells.
<3>Nat. Methods
<4>2
<5>751-756
<6>2005
<7>DNA methyltransferases have a central role in the complex regulatory network of epigenetic
modifications controlling gene expression in
mammalian cells. To study the regulation of DNA methylation in living
cells, we developed a trapping assay using transiently expressed
fluorescent DNA methyltransferase 1 (Dnmt1) fusions and mechanism-based
inhibitors 5-azacytidine (5-aza-C) or 5-aza-2'-deoxycytidine (5-aza-dC).
These nucleotide analogs are incorporated into the newly synthesized DNA
at nuclear replication sites and cause irreversible immobilization, that
is, trapping of Dnmt1 fusions at these sites. We measured trapping by
either fluorescence bleaching assays or photoactivation of
photoactivatable green fluorescent protein fused to Dnmt1 (paGFP-Dnmt1) in
mouse and human cells; mutations affecting the catalytic center of Dnmt1
prevented trapping. This trapping assay monitors kinetic properties and
activity-dependent immobilization of DNA methyltransferases in their
native environment, and makes it possible to directly compare mutations
and inhibitors that affect regulation and catalytic activity of DNA
methyltransferases in single living cells.

<>

<1>Scherr, M., Reed, M., Huang, C.F., Riggs, A.D., Rossi, J.J.
<2>Oligonucleotide scanning of native mRNAs in extracts predicts intracellular ribozyme efficiency: ribozyme-mediated reduction of the murine DNA methyltransferase.
<3>Mol. Ther.
<4>2
<5>26-38
<6>2000
<7>Modulation of gene expression by catalytic RNA requires accessible ribozyme cleavage sites in
the target mRNA, and accessibility is determined by the secondary and tertiary structure of
the target RNA, as affected by its interactions with cellular proteins. As we previously
reported, an oligonucleotide-scanning approach using antisense oligonucleotides can be used to
determine RNA accessibility in cell extracts. To test whether this method can be used to
improve selection of ribozyme target sites, we designed ribozymes corresponding to the sites
identified by oligonucleotide scanning and have evaluated their catalytic activities, first in
cell extracts and then in transduced cell lines. As a target we used the mRNA of murine DNA
(cytosine-5)-methyltransferase 1 (MTase). For intracellular studies, the ribozyme genes were
inserted downstream of a Pol III tRNAVAL promoter, which in turn was cloned in the U3 region
of a retroviral vector. We find that the efficiency of the ribozymes both in cell extracts and
in vivo corresponds with the relative effectiveness predicted by the oligonucleotide-scanning
assay. The best ribozyme causes a 70-80% reduction in the MTase mRNA levels in NIH 3T3 cells
that are stably transduced with the retroviral constructs. This reduction in mRNA levels is
accompanied by a small decrease in the methylation of repetitive intercisternal A particle DNA
elements. Ribozyme expression also increased several-fold the reactivation frequency of a
methylation-silenced green fluorescent protein (GFP) transgene. Both the reduction in
methylation and reactivation of GFP were roughly equivalent to the effects obtained by
treating NIH 3T3 cells with 2.5 mM 5-azacytidine, which gives an effect of about 10% of
maximum. These results confirm the validity of the cell extract approach for ribozyme site
selection and provide a potentially useful ribozyme for future study of DNA methyltransferase
function.

<>

<1>Scherzer, E., Auer, B., Schweiger, M.
<2>Identification, purification and characterization of Escherichia coli virus T1 DNA methyltransferase.
<3>J. Biol. Chem.
<4>262
<5>15225-15231
<6>1987
<7>An Escherichia coli virus T1-induced DNA methyltransferase was identified by
activity gel analysis in homogenates of infected E. coli
DNA-adenine-methylation-deficient strains.  Although the Mr of this protein
(31,000) is in the same range as that of the E. coli DNA adenine
methyltransferase, the two proteins are not closely related; the E. coli dam
gene does not hybridize with T1 DNA. Selective conditions for measurement of
the T1 activity were developed, and the enzyme was purified to functional
homogeneity, as shown by activity analysis in polyacrylamide gels.
Requirements for optimal activity of the viral enzyme were determined to be pH
6.9, ionic strengths below 0.1 M KCl, and a temperature between 40 and 43C.
The Km for S-adenosyl-L-methionine is 4.9 microM.  The purified T1 DNA
methyltransferase is capable of methylating adenine in 5'-GATC-3' sites in
vitro.

<>

<1>Scheuring-Vanamee, E., Viadiu, H., Lukacs, C.M., Aggarwal, A.K.
<2>Two of a kind: BamHI and BglII.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>215-236
<6>2004
<7>Among the more than 3500 Type II restriction endonucleases identified to date, fourteen have
been structurally characterized so far, including EcoRI, EcoRV, BamHI, PvuII, FokI, Cfr10I,
BglI, BglII, BsoBI, NaeI, NgoMIV, Bse634I (an isoschizomer of Cfr10I), MunI, and HincII.  All
except Cfr10I and Bse634I have been found to be bound to their cognate DNA sites.  Despite the
lack of sequence homology, all REases consist of a central beta-sheet that is flanked by
alpha-helices on both sides.  Interestingly, a similar alpha/beta core is also present in
other DNA-acting enzymes such as lambda-exonuclease, MutH, Vsr endonuclease, and TnsA from the
Tn7 transposase.  In the common core only three beta-strands are absolutely conserved, two of
these strands contain the amino acid residues directly involved in catalysis.  The similarity
at the tertiary structure level is strongest between endonucleases that share a similar
cleavatge pattern, such as between BamHI and EcoRI which cleave DNA to leave four-base 5'
overhangs, or between EcoRV and PvuII which cleave DNA to produce blunt ends.  An exception is
BglI that has a similar fold as EcoRV and PvuII but cleaves DNA to leave 3' overhangs.
Overall, the similarity reflects constraints in positioning of the active sites: 17-19 A apart
to produce four-base 5' overhangs and ~2A apart to produce blunt ends.  The active sites
occur at one end of the central beta-sheet and contain at least three superimposable residues
that are critical for catalysis.  Two of these residues are acidic, while the third residue is
usually a lysine, except in BamHI, which has a glutamate and in BglII which has a glutamine.

<>

<1>Schierling, B., Dannemann, N., Gabsalilow, L., Wende, W., Cathomen, T., Pingoud, A.
<2>A novel zinc-finger nuclease platform with a sequence-specific cleavage module.
<3>Nucleic Acids Res.
<4>40
<5>2623-2638
<6>2012
<7>Zinc-finger nucleases (ZFNs) typically consist of three to four zinc fingers (ZFs) and the
non-specific DNA-cleavage domain of the restriction endonuclease FokI. In this configuration,
the ZFs constitute the binding module and the FokI domain the cleavage module. Whereas new
binding modules, e.g. TALE sequences, have been considered as alternatives to ZFs, no efforts
have been undertaken so far to replace the catalytic domain of FokI as the cleavage module in
ZFNs. Here, we have fused a three ZF array to the restriction endonuclease PvuII to generate
an alternative ZFN. While PvuII adds an extra element of specificity when combined with ZFs,
ZF-PvuII constructs must be designed such that only PvuII sites with adjacent ZF-binding sites
are cleaved. To achieve this, we introduced amino acid substitutions into PvuII that alter
K(m) and k(cat) and increase fidelity. The optimized ZF-PvuII fusion constructs cleave DNA at
addressed sites with a >1000-fold preference over unaddressed PvuII sites in vitro as well as
in cellula. In contrast to the 'analogous' ZF-FokI nucleases, neither excess of enzyme over
substrate nor prolonged incubation times induced unaddressed cleavage in vitro. These results
present the ZF-PvuII platform as a valid alternative to conventional ZFNs.

<>

<1>Schierling, B., Noel, A.J., Thi, L.H., Wende, W., Pingoud, A.
<2>Development of a 'photoswitchable' restriction endonuclease.
<3>FEBS J.
<4>276
<5>291
<6>2009
<7>The engineering of proteins in order to obtain light-responsive characteristics is in focus of
photochemical research, because
of the possibility to regulate protein function quickly and
reversibly, which is difficult to achieve by other means. As a
proof of principle for promising applications in future, we
tried to generate a restriction endonuclease that can be turned
on and off by irradiation with light, using the single chain (sc)
variant of the restriction enzyme PvuII. 'Photoswitchable' proteins
have been pro duced using azobenzene crosslinkers that
can be isomerized between the trans- and cis-state and vice
versa by illumination at specific wavelengths or by thermal
relaxation from the cis- to the more stable trans-state. Up to
now, a 7-fold higher activity could be obtained with a variant
of scPvuII modified by two intramolecular crosslinks with 4,4¢-
azobenzene-dimaleimide in the cis configuration than in the
trans configuration. Switching was shown to be fully reversible.
The higher activity of the scPvuII variant with the azobenzene
group in the cis state was shown to be due to an increase
mainly in Vmax.

<>

<1>Schierling, B., Noel, A.J., Wende, W., Hien, L.T., Volkov, E., Kubareva, E., Oretskaya, T., Kokkinidis, M., Rompp, A., Spengler, B., Pingoud, A.
<2>Controlling the enzymatic activity of a restriction enzyme by light.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>1361-1366
<6>2010
<7>For many applications it would be desirable to be able to control the activity of proteins by
using an external signal. In the present study, we have explored the possibility of modulating
the activity of a restriction
enzyme with light. By cross-linking two suitably located cysteine residues
with a bifunctional azobenzene derivative, which can adopt a cis- or
trans-configuration when illuminated by UV or blue light, respectively,
enzymatic activity can be controlled in a reversible manner. To determine
which residues when cross-linked show the largest 'photoswitch effect,'
i.e., difference in activity when illuminated with UV vs. blue light, > 30
variants of a single-chain version of the restriction endonuclease PvuII
were produced, modified with azobenzene, and tested for DNA cleavage
activity. In general, introducing single cross-links in the enzyme leads
to only small effects, whereas with multiple cross-links and additional
mutations larger effects are observed. Some of the modified variants,
which carry the cross-links close to the catalytic center, can be
modulated in their DNA cleavage activity by a factor of up to 16 by
illumination with UV (azobenzene in cis) and blue light (azobenzene in
trans), respectively. The change in activity is achieved in seconds, is
fully reversible, and, in the case analyzed, is due to a change in V(max)
rather than K(m).

<>

<1>Schierling, B., Wende, W., Pingoud, A.
<2>Redesigning the single-chain variant of the restriction endonuclease PvuII by circular permutation.
<3>FEBS Lett.
<4>586
<5>1736-1741
<6>2012
<7>The restriction endonuclease PvuII has been introduced as a sequence-specific cleavage module
in highly-specific nucleases for gene
targeting. Here, a structural reorganization of the single-chain
variant of PvuII (scPvuII) was performed by circular permutation as a
proof-of-concept in order to find out whether the relocated, new
termini next to structural elements important for DNA recognition and
catalysis could be used for the fusion with other regulatory protein
domains. Three circularly permuted variants of scPvuII were obtained
that all maintain the specific endonucleolytic activity of scPvuII.

<>

<1>Schiestl, R.H., Petes, T.D.
<2>Integration of DNA fragments by illegitimate recombination in Saccharomyces cerevisiae.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>88
<5>7585-7589
<6>1991
<7>DNA fragments (generated by BamHI treatment) with no homology to the yeast genome were
transformed into Saccharomyces cerevisiae. When the fragments were transformed in the presence
of the BamHI enzyme, they integrated into genomic BamHI sites. When the fragments were
transformed in the absence of the enzyme, they integrated into genomic G-A-T-C sites. Since
the G-A-T-C sequence is present at the ends of BamHI fragments, this result indicates that
four base pairs of homology are sufficient for some types of mitotic recombination.

<>

<1>Schildkraut, I.
<2>Screening for and characterizing restriction endonucleases.
<3>Genet. Eng. (N Y), Plenum Press, Setlow J.K., Hollaender, A., New York
<4>6
<5>117-140
<6>1984
<7>The ability to cleave DNA at specific sequences is the fundamental technology
which was responsible for the rapid development of genetic engineering.  The
type II restriction endonucleases are the enzymes which enable scientists to
cleave DNA at specific sequences.  In 1974, Richard Roberts distributed a list
of 30 restriction endonucleases.  Eighteen of these recognized unique
sequences; the remainder were isoschizomers, enzymes from different bacterial
sources that recognize the same sequence.  Presently the list contains 398
restriction endonucleases, 91 of which are unique.  The increased number of
restriction endonucleases available now has allowed greater flexibility and
versatility in experimental strategies and design.  Continuing the search for
and identifying new restriction endonucleases will further reduce the
limitations and permit more precision in genetic engineering.  This chapter
describes a method for screening bacterial cells for the presence of type II
restriction endonucleases, and procedures for characterizing these restriction
endonucleases with respect to recognition sequence and site of cleavage.  The
recognition sequence is defined here as the nucleotide sequence which is
required for cleavage.  The site of cleavage is defined as the position of the
cleavage with relation to the recognition sequence.  For example, the
recognition sequence for EcoRI is GAATTC.  The site of cleavage is between the
G and A residues which produces a four base 5' extension (GAATTC).

<>

<1>Schildkraut, I., Banner, C.D.B., Rhodes, C.S., Parekh, S.
<2>The cleavage site for the restriction endonuclease EcoRV is 5'-GAT^ATC-3'.
<3>Gene
<4>27
<5>327-329
<6>1984
<7>The cleavage site for the restriction endonuclease EcoRV has been found to be
5'-GAT^ATC-3', rather than 5'-GATAT^C-3' as reported earlier by Kholmina et al.
(Dokl. Akad. Nauk. SSSR 253 (1980) 495-497).

<>

<1>Schildkraut, I., Lynch, J., Morgan, R.
<2>The cleavage site for the restriction endonucleases BanI and HgiCI is 5' ...G^GPyPuCC ...3'.
<3>Nucleic Acids Res.
<4>15
<5>5492
<6>1987
<7>We have determined the cleavage site for the restriction endonucleases BanI and
HgiC 1 to be identical.  Both cleaving the sequence 5'... G^GPyPuCC ...3'
leaving a 4 base 5' extension.  This is in contrast to Kroger et al, who
reported the cleavage for HgiCI as 5' ...^GGPyPuCC...3' leaving a 6 base 5'
extension.

<>

<1>Schink, A.K., Kadlec, K., Schwarz, S.
<2>Detection of qnr genes among Escherichia coli isolates of animal origin and complete sequence of the conjugative qnrB19-carrying plasmid pQNR2078.
<3>J. Antimicrob. Chemother.
<4>67
<5>1099-1102
<6>2012
<7>OBJECTIVES: The aims of this study were to identify qnr genes among
quinolone-resistant Escherichia coli isolates from defined disease conditions of
companion and farm animals obtained in the BfT-GermVet study, and to gain insight
into their localization and the organization of the qnr gene regions. METHODS:
The qnr genes were detected by PCR and confirmed by sequencing. qnr-positive
isolates were checked for mutations in DNA gyrase and topoisomerase IV genes by
PCR and sequencing of the quinolone resistance-determining regions. Multilocus
sequence typing (MLST) was performed for the qnr-positive E. coli isolates.
Plasmids harbouring qnr genes were transferred by conjugation into E. coli
recipients, subjected to PCR-based replicon typing and plasmid-MLST, and one
qnrB19-carrying plasmid was sequenced completely. RESULTS: Only 2 of 417 E. coli
isolates investigated carried qnr genes. Both isolates originated from horses and
showed MLST type ST86. They harboured conjugative qnrB19-carrying plasmids, which
proved to be indistinguishable by restriction analysis, belonged to
incompatibility group IncN, showed plasmid-MLST type ST8 and did not carry other
resistance genes. The qnrB19 gene was flanked by copies of the insertion sequence
IS26. One of these plasmids, pQNR2078, was sequenced completely and had a size of
42 379 bp. Except for the resistance gene region, plasmid pQNR2078 closely
resembled the bla(CTX-M-65)-carrying plasmid pKC396 from E. coli. CONCLUSIONS:
qnr genes were rarely detected among E. coli from animals in the BfT-GermVet
study. The qnrB19 gene was detected on conjugative plasmids, with IS26 being
likely involved in the mobility of qnrB19.

<>

<1>Schinzel, R., Burger, K.J.
<2>A site-specific endonuclease activity in Halobacterium halobium.
<3>FEMS Microbiol. Lett.
<4>37
<5>325-329
<6>1986
<7>In Halobacterium halobium and some related strains, a site-specific
endonuclease activity was found.  This activity requires 3 M NaCl and 5-10 mM
Mg2+ ions for function.  The 3 cleavage sites in plasmid pBR322 were mapped,
but no homology between these sites was found.  H. halobium DNA is resistant to
cleavage, which may be due to a modification of the DNA.  The behaviour of the
endonuclease can be explained by the presence of a Type I or Type III-like
restriction-modification system.

<>

<1>Schirawski, J., van Sinderen, D., Fitzgerald, G.
<2>Bacteriophage resistance in Streptococcus thermophilus.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>476
<6>2001
<7>Thermophilic lactic acid bacteria are frequently used as starter cultures for the industrial
preparation of yoghurt or cheese.
Bacteriophage attack of these starter cultures has been described as
the main reason for fermentation failures. Natural phage resistance
mechanisms of bacteria include restriction/modification (R/M) systems,
enzyme complexes, that modify host DNA by methylation and degrade
unmodified incoming DNA. Type 1 R/M systems are encoded by three genes,
named hsdR, hsdM and hsdS. A complex of all three proteins, R, M and S,
is needed for degradation of unmodified DNA, whereas DNA modification
only requires the M and S subunits. The hsdM and hsdS genes are located
on the same operon, whereas the hsdR gene is usually transcribed from
its own promoter. A type I R/M system was identified on the genome of
S. thermophilus 4134, a phage resistant industrial strain used for the
production of yoghurt. The organization of the three genes is unusual,
in that they appear to be located on the same operon. The hsdR and hsdM
genes show high sequence identity to the corresponding genes of plasmid
located type 1 R/M systems from Lactococcos lactis or S. thermophilus,
suggesting a gene transfer event from a mobile genetic element into the
genome of S. thermophilus 4134.

<>

<1>Schirrmacher, E., Beck, C., Brueckner, B., Schmitges, F., Siedlecki, P., Bartenstein, P., Lyko, F., Schirrmacher, R.
<2>Synthesis and in vitro evaluation of biotinylated RG108: a high affinity compound for studying binding interactions with human DNA  methyltransferases.
<3>Bioconjugate Chem.
<4>17
<5>261-266
<6>2006
<7>Small-molecule inhibitors of DNA methyltransferases such as RG108 represent promising
candidates for cancer drug development. We report the
synthesis and in vitro analysis of a biotinylated RG108 conjugate,
2-(1,3-dioxo-1,3-dihydro-isoindol-2-yl)-3-(5-[3-[5-(2-oxo-hexahydro-thieno
[3,4-d]imidazol-4-yl)pentanoylamino]propoxy]-1H-indol-3-yl)propionic acid
(bio-RG108), for the evaluation of interactions with DNA methyltransferase
enzymes. The structural design of the chemically modified inhibitor was
aided by molecular modeling, which suggested the possibility for extensive
chemical modifications at the 5-position of the tryptophan moiety in
RG108. The inhibitory activity of the corresponding derivative was
confirmed in a cell-free biochemical assay, where bio-RG108 showed an
undiminished inhibition of DNA methyltransferase activity (IC50 = 40 nM).
Bio-RG108 therefore represents a suitable bioconjugate for the elucidation
of inhibitory mechanisms and for the affinity purification of
RG108-associated proteins.

<>

<1>Schirwitz, K., Lundin, A., Skoglund, A., Krabbe, M., Engstrand, L., Enroth, C.
<2>Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of HP1352, a putative DNA methyltransferase  in Helicobacter pylori.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>59
<5>719-720
<6>2003
<7>The putative methyltransferase gene HP1352 from Helicobacter pylori strain 26695 was
heterologously expressed in Escherichia coli. The 359-amino-acid
gene product was purified and crystallized. The crystals belong to space
group I2(1)2(1)2(1) and show diffraction to at least 2.5 A resolution. The
unit-cell parameters are a = 69.6, b = 86.6, c = 140.0 A. A greater than
90% complete native data set has been collected and structure
determination using the molecular-replacement method is ongoing.

<>

<1>Schlagman, S., Hattman, S.
<2>Mutants of the N-3 R-Factor conditionally defective in hspII modification and deoxyribonucleic acid-cytosine methylase activity.
<3>J. Bacteriol.
<4>120
<5>234-239
<6>1974
<7>The N-3 resistance (R) factor specifies a deoxyribonucleic acid (DNA)-cytosine
methylase and a DNA restriction-modification (hspII) system.  We have isolated
three independent mutants that are conditionally defective in their ability to
modify bacteriophage lambda and to methylate DNA-cytosine residues.  The ratio
of 5-methylcytosine to N6-methyladenine in bacterial DNA and in the DNA of
phages lambda and fd was determined after labeling with [methyl-3H]methionine
at various growth temperatures.  Although the ability of the wild-type N-3
factor to modify phage lambda and to methylate DNA-cytosine residues was
unaffected with increasing temperature, two of the mutants exhibited a parallel
loss in modification and cytosine methylation ability.  The ability of the
third mutant to carry out these functions was dependent on the presence or
absence of an amber suppressor mutation in the host genome.  These results
offer further support for the notion that hspII modification is mediated by a
DNA-cytosine methylase.  Evidence is also presented that the modification
methylase is responsible for the in vivo methylation of phage fd DNA (which is
not subject to hspII restriction in vivo).

<>

<1>Schlagman, S., Hattman, S., May, M.S., Berger, L.
<2>In vivo methylation by Escherichia coli K-12 mec+ Deoxyribonucleic Acid-cytosine methylase protects against in vitro cleavage by the RII restriction endonuclease (R.EcoRII).
<3>J. Bacteriol.
<4>126
<5>990-996
<6>1976
<7>We have analyzed the susceptibility of the deoxyribonucleic acid (DNA) of phage
fd replicative form (RF) and of Escherichia coli to in vitro cleavage by
purified RII restriction endonuclease (R.EcoRII).  The results are summarized
as follows:  (i) fd.mec- RFI, isolated from infected E. coli K-12 mec- bacteria
(a mutant strain lacking DNA-cytosine methylase activity), is cleaved into at
least two fragments, whereas fd.mec+ RFI, isolated from the parental mec+
strain, is not cleaved.  (ii) E. coli mec- DNA is extensively degraded, whereas
E. coli mec+ DNA is resistant to cleavage.  We conclude that the E. coli mec+
DNA-cytosine methylase acts as an RII modification enzyme.

<>

<1>Schlagman, S.L., Hattman, S.
<2>The bacteriophage T2 and T4 DNA-[N6-adenine] methyltransferase (Dam) sequence specificities are not identical.
<3>Nucleic Acids Res.
<4>17
<5>9101-9112
<6>1989
<7>Bacteriophages T2 and T4 encode DNA-[N6-adenine] methyltransferases (Dam) which differ from
each other by only three amino acids. The canonical recognition sequence for these enzymes in
both cytosine and 5-hydroxymethylcytosine-containing DNA is GATC; at a lower efficiency they
also recognize some non-canonical sites in sequences derived from GAY (where Y is cytosine or
thymine). We found that T4 Dam fails to methylate certain GATA and GATT sequences which are
methylated by T2 Dam. This indicates that T2 Dam and T4 Dam do not have identical sequence
specificities. We analyzed DNA sequence data files obtained from GenBank, containing bout 30%
of the T4 genome, to estimate the overall frequency of occurrence of GATC, as well as
non-canonical sites derived from GAY. The observed N6methyladenine (m6A) content of T4 DNA,
methylated exclusively at GATC (by Escherichia coli Dam), was found to be in good agreeement
with this estimate. Although GATC is fully methylated in virion DNA, only a small percentage
of the non-canonical sequences are methylated.

<>

<1>Schlagman, S.L., Hattman, S.
<2>Molecular cloning of a functional dam+ gene coding for phage T4 DNA adenine methylase.
<3>Gene
<4>22
<5>139-156
<6>1983
<7>Phages T2 and T4 induce synthesis of a DNA-adenine methylase which is coded for
by a phage gene, dam+. These enzymes methylate adenine residues in specific
sequences which include G-A-T-C, the methylation site of the host Escherichia
coli dam+ methylase.  Methylation of G-A-T-C to G-m6A-T-C protects the site
against cleavage by the MboI restriction nuclease.  We have taken advantage of
this property to enrich and screen for transformants which contain a cloned,
functional T4 dam+ gene.  These recombinant molecules consist of a 1.85-kb
HindIII fragment inserted into the plasmid pBR322; both orientations of the
fragment express the methylase gene, suggesting that transcription is from a T4
promoter.  We have tested the 1.85-kb insert for sensitivity to a variety of
restriction nucleases and have found single sites for EcoRI, BalI, XbaI, and at
least two sites for BstNI (EcoRII).  The relative positions of these
restriction sites have also been determined.  Physical mapping was carried out
by Southern blot hybridization with 32P-labeled (nick-translated clone) probe.
These experiments showed that the insert corresponds to a HindIII fragment
located on the physical map of T4 between positions 16.2 and 18.1 kb from the
T4rIIA-rIIB junction.  E. coli dam- possesses several phenotypic differences
from the wild-type dam+ parent, including an increased sensitivity to
2-aminopurine (2-AP).  We found that T4 dam+ clones could relieve dam- cells of
their increased sensitivity to 2-AP.

<>

<1>Schlagman, S.L., Hattman, S., Marinus, M.G.
<2>Direct role of the Escherichia coli Dam DNA methyltransferase in methylation-directed mismatch repair.
<3>J. Bacteriol.
<4>165
<5>896-900
<6>1986
<7>The T4 dam+ gene has been cloned (S. L. Schlagman and S. Hattman, Gene 22:139-156, 1983) and
transferred into an Escherichia coli dam-host. In this host, the T4 Dam DNA methyltransferase
methylates mainly, if not exclusively, the sequence 5'-GATC-3'; this sequence specificity is
the same as that of the E. coli Dam enzyme. Expression of the cloned T4 dam+ gene suppresses
almost all the phenotypic traits associated with E. coli dam mutants, with the exception of
hypermutability. In wild-type hosts, 20- to 500-fold overproduction of the E. coli Dam
methylase by plasmids containing the cloned E. coli dam+ gene results in a hypermutability
phenotype (G.E. Herman and P. Modrich, J. Bacteriol. 145:644-646, 1981; M.G. Marinus, A.
Poteete, and J.A. Arraj, Gene 28:123-125, 1984). In contrast, the same high level of T4 Dam
methylase activity, produced by plasmids containing the cloned T4 dam+ gene, does not result
in hypermutability. To account for these results we propose that the E. coli Dam methylase may
be directly involved in the process of methylation-instructed mismatch repair and that the T4
Dam methylase is unable to substitute for the E. coli enzyme.

<>

<1>Schlagman, S.L., Miner, Z., Feher, Z., Hattman, S.
<2>The DNA [adenine-N6]methyltransferase (Dam) of bacteriophage T4.
<3>Gene
<4>73
<5>517-530
<6>1988
<7>A functional bacteriophage T4 dam+ gene, which specifies a DNA [adenine-N6]
methyltransferase (Dam), was cloned on a 1.8-kb HindIII fragment [Schlagman and
Hattman, Gene 22 (1983) 139-156].  Sequence analysis [McDonald and Mosig, EMBO
J. 3(1984) 2863-2871] revealed two overlapping in-phase open reading frames
(ORFs).  The 5' proximal ORF initiates translation at an AUG and encodes a
30-kDA polypeptide, whereas the downstream ORF initiates translation at a GUG
and encodes a 26-kDa polypeptide.  Analysis of BAL 31 deletions in our original
dam+ clone has verified that at least one of these overlapping ORFs, in fact,
encodes T4 Dam.  To investigate where T4 Dam translation is initiated, we have
constructed plasmids in which a tac or lambda pL promoter is placed 5' to
either the longer ORF or just the shorter ORF.  Only clones which contain a
promoter in front of the longer ORF produce active T4 Dam.  This indicates that
the 26-kDa polypeptide alone cannot be T4 Dam.  Additional experiments suggest
that only the 30-kDa polypeptide is required for enzyme activity and that the
shorter ORF is not translated in plasmid-carrying cells.  We also present
evidence that T4 Dam is capable of methylating 5'-GATC-3', GATm5C, and GAThmC
sequences; non-canonical sites (e.g., GACC) are also methylated, but much less
efficiently.

<>

<1>Schleheck, D., Weiss, M., Pitluck, S., Bruce, D., Land, M.L., Han, S., Saunders, E., Tapia, R., Detter, C., Brettin, T., Han, J., Woyke, T., Goodwin, L., Pennacchio, L., Nolan, M., Cook, A.M., Kjelleberg, S., Thomas, T.
<2>Complete genome sequence of Parvibaculum lavamentivorans type strain (DS-1(T)).
<3>Standards in Genomic Sciences
<4>5
<5>298-310
<6>2011
<7>Parvibaculum lavamentivorans DS-1(T) is the type species of the novel genus Parvibaculum in
the novel family Rhodobiaceae (formerly Phyllobacteriaceae) of
the order Rhizobiales of Alphaproteobacteria. Strain DS-1(T) is a non-pigmented,
aerobic, heterotrophic bacterium and represents the first tier member of
environmentally important bacterial communities that catalyze the complete
degradation of synthetic laundry surfactants. Here we describe the features of
this organism, together with the complete genome sequence and annotation. The
3,914,745 bp long genome with its predicted 3,654 protein coding genes is the
first completed genome sequence of the genus Parvibaculum, and the first genome
sequence of a representative of the family Rhodobiaceae.

<>

<1>Schleif, R.
<2>Assaying of organisms for the presence of restriction endonucleases.
<3>Methods Enzymol.
<4>65
<5>19-23
<6>1980
<7>The utility of sequence-specific endonucleases recommends their use in a wide variety of
applications.  Typically a potential user is faced with the problem of determining which of
the large number of enzymes already known will cleave in acceptable locations.  Thus an easy,
uniform, and rapid scheme for partial purification of these enzymes would greatly assist the
initial screening.  A purification step fulfilling these requirements could also be of value
as a first step when pure enzyme is required or of use in searching for new varieties of
endonuclease.  Dextran-polyethylene glycol phase partition meets the requirements of speed and
convenience.  Here it is shown that adjustment of the salt concentration during the phase
partition step allows separation of most of the interfering nuclease activities in the crude
extracts of many strains containing site-specific nucleases.

<>

<1>Schleper, C.
<2>Metagenomic analysis of ammonia oxidizing archaea affiliated with the soil group.
<3>Front. Microbiol.
<4>3
<5>208
<6>2012
<7>Ammonia-oxidising archaea (AOA) have recently been recognized as a significant component of
many microbial communities and represent one of the most abundant prokaryotic groups in the
biosphere. However, only few AOA have been successfully cultivated so far and information on
the physiology and genomic content remains scarce. We have performed a metagenomic analysis to
extend the knowledge of the AOA affiliated with group I.1b that is widespread in terrestrial
habitats and of which no genome sequences has been described yet. A fosmid library was
generated from samples of a radioactive thermal cave (46°C) in the Austrian Central Alps in
which AOA had been found as a major part of the microbial community. Out of sixteen fosmids
that possessed either an amoA or 16S rRNA gene affiliating with AOA, five were fully
sequenced, four of which grouped with the soil/I.1b (Nitrososphaera-) lineage and one with
marine/I.1a (Nitrosopumilus-) lineage. Phylogenetic analyses of amoBC and an associated
conserved gene were congruent with earlier analyses based on amoA and 16S rRNA genes and
supported the separation of the soil and marine group. Several putative genes that did not
have homologues in  currently available marine thaumarchaeota genomes indicated that AOA of
the soil group contain specific genes that are distinct from their marine relatives. Potential
cis regulatory elements around conserved promoter motifs found upstream of the amo genes in
sequenced (meta-) genomes differed in marine and soil group AOA. On one fosmid, a group of
genes including amoA and amoB were flanked by identical transposable insertion sequences,
indicating that amoAB could potentially be co-mobilized in the form of a composite transposon.
This might be one of the mechanisms that caused the greater variation in gene order compared
to genomes in the marine counterparts. Our findings highlight the genetic diversity within the
two major and widespread lineages of thaumarchaeota.

<>

<1>Schleper, C., DeLong, E.F., Preston, C.M., Feldman, R.A., Wu, K.-Y., Swanson, R.V.
<2>Genomic analysis reveals chromosomal variation in natural populations of the uncultured psychrophilic archaeon Cenarchaeum symbiosum.
<3>J. Bacteriol.
<4>180
<5>5003-5009
<6>1998
<7>Molecular phylogenetic surveys have recently revealed an ecologically widespread crenarchaeal
group that inhabits cold and temperate terrestrial and marine environments.  To date these
organisms have resisted isolation in pure culture, and so their phenotypic and genotypic
characteristics remain largely unknown.  To characterize these archaea, and to extend
methodological approaches for characterizing uncultivated microorganisms, we initiated genomic
analyses of the nonthermophilic crenarchaeote Cenarchaeum symbiosum found living in
association with a marine sponge, Axinella mexicana.  Complex DNA libraries derived from the
host-symbiont population yielded several large clones containing the ribosomal operon from C.
symbiosum.  Unexpectedly, cloning and sequence analysis revealed the presence of two closely
related variants that were consistently found in the majority of host individuals analyzed.
Homologous regions from the two variants were sequenced and compared in detail.  The variants
exhibit >99.2% sequence identity in both small- and large-subunit rRNA genes and they contain
homologous protein-encoding genes in identical order and orientation over a 28-kbp overlapping
region.  Our study not only indicates the potential for characterizing uncultivated
prokaryotes by genome sequencing but also identifies the primary complication inherent in the
approach: the widespread genomic microheterogeneity in naturally occurring prokaryotic
populations.

<>

<1>Schlesinger, D.J., Shoemaker, N.B., Salyers, A.A.
<2>Possible Origins of CTnBST, a Conjugative Transposon Found Recently in a Human Colonic Bacteroides Strain.
<3>Appl. Environ. Microbiol.
<4>73
<5>4226-4233
<6>2007
<7>A previous survey of Bacteroides isolates suggested that the ermB gene
entered Bacteroides spp. recently. Previously, ermB had been found almost
exclusively in gram-positive bacteria. In one Bacteroides strain, ermB was
located on 100-kb conjugative transposon (CTn) CTnBST. To assess the
possible origin of this CTn, we obtained the full DNA sequence of CTnBST
and used this information to investigate its possible origins. Over
one-half of CTnBST had high sequence identity to a putative CTn found in
the genome of Bacteroides fragilis YCH46. This included the ends of the
CTn and genes involved in integration, transfer, and excision. However,
the region around the ermB gene contained genes that appeared to originate
from gram-positive organisms. In particular, a 7-kb segment containing the
ermB gene was 100% identical to an ermB region found in the genome of the
gram-positive bacterium Arcanobacterium pyogenes. A screen of Bacteroides
isolates whose DNA cross-hybridized with a CTnBST probe revealed that
several isolates did not carry the 7-kb region, implying that the
acquisition of this region may be more recent than the acquisition of the
entire CTnBST element by Bacteroides spp. We have also identified other
Bacteroides isolates that carry a slightly modified 7-kb region but have
no other traces of CTnBST. Thus, it is possible that this 7-kb region
could itself be part of a mobile element that has inserted in a
Bacteroides CTn. Our results show that CTnBST is a hybrid element which
has acquired a portion of its coding region from gram-positive bacteria
but which may originally have come from Bacteroides spp. or some related
species.

<>

<1>Schlickenrieder, M.
<2>Structure, localization/regulation and interaction-partners of the Dam-Methyltransferase of Escherichia coli.
<3>Ph.D. Thesis, Germany
<4>
<5>
<6>2006
<7>
<>

<1>Schluckebier, G., Kozak, M., Bleimling, N., Weinhold, E., Saenger, W.
<2>Differential binding of S-adenosylmethionine, S-adenosylhomocysteine and sinefungin to the adenine-specific DNA methyltransferase M.TaqI.
<3>J. Mol. Biol.
<4>265
<5>56-67
<6>1997
<7>The crystal structures of the binary complexes of the DNA methyltransferase M.TaqI with the
inhibitor Sinefungin and the reaction product S-adenosyl-L-homocysteine were determined, both
at 2.6 A resolution.  Structural comparison of these binary complexes with the complex formed
by M.TaqI and the cofactor S-adenosyl-L-methionine suggests that the key element for molecular
recognition of these ligands is the binding of their adenosine part in a pocket, and
discrimination between cofactor, reaction product and inhibitor is mediated by different
conformations of these molecules; the methionine part of S-adenosyl-L-methionine is located in
the binding cleft, whereas the amino acid moieties of Sinefungin and S-adenosyl-l-homocysteine
are in a different orientation and interact with the active site amino acid residues
105NPPY108.  Dissociation constants for the complexes of M.TaqI with the three ligands were
determined spectrofluorometrically.  Sinefungin binds more strongly than
S-adenosyl-L-homocysteine or S-adenosyl-L-methionine, with KD=0.34 microM, 2.4 microM and 2.0
microM, respectively.

<>

<1>Schluckebier, G., Labahn, J., Granzin, J., Saenger, W.
<2>Structure and function of the adenine specific DNA methyltransferase M.TaqI.
<3>J. Biomol. Struct. Dyn.
<4>12
<5>A206
<6>1995
<7>Methylation of DNA bases is frequently employed by most living organisms to enhance the
informational content of DNA.  DNA methyltransferases (MTases) transfer a methyl group from
the cofactor S-adenosyl-methionine (AdoMet) to either adenine N6, cytosine N4 (N-MTases) or
cytosine C5 (C-MTases) in a specific DNA sequence.  AdoMet is thereby converted to
S-adenosyl-homocysteine (AdoHcy), and a proton is released.  In procaryotes, DNA MTases are
usually part of restriction/modification systems (R/M system), which consist typically of an
endonuclease (ENase) and a DNA MTase with the same recognition sequence.  The ENase cleaves
unmethylated double stranded DNA, e.g. of invading viruses.  The cognate MTase methylates the
host's DNA making it insusceptable against cleavage.  In eucaryotes DNA methylation plays a
role in such different phenomena as cell differentiation, genomic imprinting and
carcinogenesis.  We determined the first crystal structure of an N-MTase from the R/M system I
of Thermus aquaticus (M.TaqI) in complex with its cofactor AdoMet at a resolution of 2.4
Angstroms.  It shows alpha/beta-folding of the enzyme into two domains of roughly equal size.
Both domains are oriented towards each other such that they form a cleft which is studded
with positively charged amino acid residues and is sufficiently wide to accommodate B-DNA.
The N-terminal domain binds AdoMet and contains all amino acid residues which are conserved
throughout all DNA MTases.  As with probably all AdoMet dependent MTases the cofactor is bound
at an alpha-beta-alpha-fold which resembles the Rossmann-fold of dinucleotide binding
proteins.  The C-terminal domain functions in DNA binding and molecular recognition of the DNA
substrate.  Additionally, crystals of the enzyme complexed with the reaction product AdoHcy
and the competitive inhibitor Sinefungin were grown.  The X-Ray structures show the same
overall folding of the polypeptide chain as with the M.TaqI-AdoMet complex.  Small, but
distinctive differences in the binding of the small molecules provide insight into the
molecular recognition of the cofactor.

<>

<1>Schluckebier, G., Labahn, J., Granzin, J., Saenger, W.
<2>M.TaqI: Possible catalysis via cation-pi interactions in N-specific DNA methyltransferases.
<3>Biol. Chem.
<4>379
<5>389-400
<6>1998
<7>The adenine-specific DNA methyltransferase M.TaqI transfers a methyl group from
S-adenosylmethionine to N6 of the adenine residue in the DNA sequence 5'-TCGA-3'.  In the
crystal structure of M.TaqI in complex with S-adenosylmethionine the enzyme is folded into two
domains: An N-terminal catalytic domain, whose fold is conserved among S-adenosylmethionine
dependent methyltransferases, and a DNA recognition domain which possesses a unique fold.  In
the active site, two aromatic residues, Tyr 108 and Phe 196, are postulated to bind the
flipped-out target DNA adenine which becomes methylated.  By lowering the energy of the
positively charged transition state via cationic-pi interactions, these two residues probably
hold a key role in catalysis.

<>

<1>Schluckebier, G., Labahn, J., Granzin, J., Schildkraut, I., Saenger, W.
<2>A model for DNA binding and enzyme action derived from crystallographic studies of the TaqI N6-adenine-methyltransferase.
<3>Gene
<4>157
<5>131-134
<6>1995
<7>The crystal structures of the DNA-N6-adenine-methyltransferase M.TaqI, in complexes with the
cofactor S-adenosyl-L-methionine (AdoMet) and the competitive inhibitor sinefungin (Sf) show
identical folding of the polypeptide chains into two domains.  The N-terminal domain carries
the cofactor-binding site, the C-terminal domain is thought to be implicated in
sequence-specific DNA binding.  Model building of the M.TaqI-DNA complex suggests that the
adenine to be methylated swings out of the double helix as found previously in the
cytosine-C5-MTase HhaI DNA co-crystal structure.  A torsion of the methionine moiety of the
cofactor is required to bring the methyl group within reach of the swung-out base and allow
methyl group transfer.

<>

<1>Schluckebier, G., O'Gara, M., Saenger, W., Cheng, X.
<2>Universal catalytic domain structure of AdoMet-dependent methyltransferases.
<3>J. Mol. Biol.
<4>247
<5>16-20
<6>1995
<7>The DNA methyltransferases, M.HhaI and M.TaqI, and catechol O-methyl-transferase (COMT)
catalyze the transfer of a methyl group from the cofactor S-adenosyl-L-methionine (AdoMet) to
carbon-5 of cytosine, to nitrogen-6 of adenine, and to a hydroxyl group of catechol,
respectively. The catalytic domains of the bilobal proteins, M.HhaI and M.TaqI, and the entire
single domain of COMT have similar folding with an alpha/beta structure containing a mixed
central beta-sheet. The functional residues are located in equivalent regions at the carboxyl
ends of the parallel beta-strands. The cofactor binding sites are almost identical and the
essential catalytic amino acids coincide. The comparable protein folding and the existence of
equivalent amino acids in similar secondary and tertiary positions indicate that many (if not
all) AdoMet-dependent methyltransferases have a common catalytic domain structure. This
permits tertiary structure prediction of other DNA, RNA, protein, and small-molecule
AdoMet-dependent methyltransferases from their amino acid sequences.

<>

<1>Schluter, A., Heuer, H., Szczepanowski, R., Poler, S.M., Schneiker, S., Puhler, A., Top, E.M.
<2>Plasmid pB8 is closely related to the prototype IncP-1beta plasmid R751 but transfers poorly to Escherichia coli and carries a new transposon encoding a small multidrug resistance efflux protein.
<3>Plasmid
<4>54
<5>135-148
<6>2005
<7>The IncP-1beta plasmid pB8, which confers resistance to amoxicillin,
spectinomycin, streptomycin, and sulfonamides, was previously isolated
from a sewage treatment plant. It was found to possess abnormal
conjugative transfer properties, i.e., transfer to Escherichia coli by
conjugation or electroporation could not be detected. We showed in this
study that plasmid pB8 is transferable to E. coli by conjugation, but only
at low frequencies and under specific experimental conditions, a
phenomenon that is very unusual for IncP-1 plasmids. Determination of the
complete 57,198bp pB8 nucleotide sequence revealed that the backbone of
the plasmid consists of a complete set of IncP-1beta-specific genes for
replication initiation, conjugative plasmid transfer, stable inheritance,
and plasmid control with an organisation identical to that of the
prototype IncP-1beta plasmid R751. All of the minor differences in the pB8
backbone sequence compared to that of R751 were also found in other
IncP-1beta plasmids known to transfer to and replicate in E. coli.
Plasmids pB8 and R751 can be distinguished with respect to their accessory
genetic elements. First, the pB8 region downstream of the replication
initiation gene trfA contains two transposable elements one of which is
similar to Tn5501. The latter transposon encodes a putative
post-segregational-killing system and the small multidrug resistance (SMR)
protein QacF, mediating quaternary ammonium compound resistance. The
accessory genes in this region are not responsible for the poor plasmid
transfer to E. coli since a pB8 deletion derivative devoid of all genes in
that region showed the same conjugative transfer properties as pB8. A
Tn5090/Tn402 derivative carrying a class 1 integron is located between the
conjugative transfer modules. The Tn5090/Tn402 integration-sites are
exactly identical on pB8 and R751 but in contrast to R751 the pB8 element
carries the resistance gene cassettes oxa-2 for amoxicillin resistance and
aadA4 for streptomycin/spectinomycin resistance, the integron-specific
conserved segment consisting of the genes qacEDelta1, sul1, and orf5, and
a truncated tni transposition module (tniAB). Although future work will
have to determine the molecular basis for the poor transfer of pB8 to E.
coli, our findings demonstrate that the host-range of typical IncP-1
plasmids may be less broad than expected.

<>

<1>Schmeisser, C. et al.
<2>Rhizobium sp. NGR234 possesses a remarkable number of secretion systems.
<3>Appl. Environ. Microbiol.
<4>75
<5>4035-4045
<6>2009
<7>Rhizobium sp. NGR234 is a unique alpha-proteobacterium (Order -
Rhizobiales) that forms nitrogen-fixing nodules with more legumes than any
other micro-symbiont. Here we report that the 3.93 Mbp chromosome
(cNGR234) encodes most functions required for cellular growth. Few
essential functions are encoded on the 2.43 Mbp mega-plasmid (pNGR234b)
and none are present on the second 0.54 Mbp symbiotic plasmid (pNGR234a).
Amongst many striking features, the 6.9 Mbp genome encodes more different
secretion systems than any other known rhizobia and probably most known
bacteria. Altogether 132 genes and proteins are linked to secretory
processes. Secretion systems identified include general and export
pathways (GSP

<>

<1>Schmid, E.N., von Recklinghausen, G., Ansorg, R.
<2>Bacteriophages in Helicobacter (Campylobacter) pylori.
<3>J. Med. Microbiol.
<4>32
<5>101-104
<6>1990
<7>Bacteriophages in different stages of maturation were found in thin sections of a clinical
isolate of Helicobacter (Campylobacter) pylori. Mature phage heads measured 70 x 60 nm and the
tail at least 120 nm. Lysogeny was maintained during subculture on blood agar for more than 3
months after isolation from a gastric biopsy.

<>

<1>Schmid, J., Koenig, S., Pick, A., Steffler, F., Yoshida, S., Miyamoto, K., Sieber, V.
<2>Draft Genome Sequence of Kozakia baliensis SR-745, the First Sequenced Kozakia Strain from the Family Acetobacteraceae.
<3>Genome Announcements
<4>2
<5>e00594-14
<6>2014
<7>Kozakia baliensis belongs to the family Acetobacteraceae and was described for the first time
in 2002. These acetic acid bacteria are able to produce acetic
acid from various carbon sources and 2- and 5-keto-d-gluconate from glucose. The
novel K. baliensis strain SR-745 was isolated from a pineapple fruit bought in a
German supermarket. The strain produces large amounts of organic acids when grown
on glucose-containing medium and accepts also glycerol, fructose, mannitol, and
sucrose as a C source. When grown under light and high-oxygen conditions in
submerged culture, the production of a pink pigment is observed after 72 h.

<>

<1>Schmid, K., Thomm, M., Laminet, A., Laue, F.G., Kessler, C., Stetter, K.O., Schmitt, R.
<2>Three new restriction endonucleases MaeI, MaeII and MaeIII from Methanococcus aeolicus.
<3>Nucleic Acids Res.
<4>12
<5>2619-2628
<6>1984
<7>Three type II restriction endonucleases, MaeI, MaeII and MaeIII, with novel
site specificities have been isolated and purified form the archaebacterium
Methanococcus aeolicus PL-15/H.  The recognition sequences of these enzymes are
C^T A G   (MaeI)  A^C G T   (MaeII) ^G T N A C  (MaeIII)  with the sites of
cleavage as indicated by the arrows.  The sequences were confirmed by
restriction and computer analyses on sequenced DNA's of plasmid pBR322,
bacteriophage lambda and PhiX174 and virus SV40.

<>

<1>Schmidt, A., Reinert, H., Venner, H., Bieber, J.
<2>Studies on the substrate specificity of the DNA methylase activity from Escherichia coli K-12.
<3>Z. Allg. Mikrobiol.
<4>19
<5>489-495
<6>1979
<7>A partially purified extract of DNA methylases from E. coli K-12 containing DNA-adenine as
well as DNA-cytosine methylase activities has been examined with respect to different DNA
species as substrates.  The results show that the natural content of 6-MAP in the applied DNA
represses the DNA-adenine methylase activity.  On the other hand, 5-MC, already present in the
substrate does not influence the activity of the DNA-cytosine methylase.  DNA from Micrococcus
radiodurans, which is completely free of methylated bases served as a comparison.  Since
netropsin preferentially binds to AT-rich regions of DNA, the influence of this oligopeptide
antibiotic on the methylation of DNA was investigated.  As expected the antibiotic
predominantly inhibits adenine methylation of DNA.  The degree of inhibition depends on the
molar ratio of netropsin to DNA phosphate.

<>

<1>Schmidt, F.H.-G., Hueben, M., Gider, B., Renault, F., Teulade-Fichou, M.-P., Weinhold, E.
<2>Sequence-specific methyltransferase-induced labelling (SMILing) of plasmid DNA for studying cell transfection.
<3>Bioorg. Med. Chem.
<4>16
<5>40-48
<6>2008
<7>Plasmid DNA (pUC19 and pBR322) was sequence-specifically, covalently labelled with Cy3
fluorophores using a newly synthesised
N-adenosylaziridine cofactor and the DNA methyltransferase M.TaqI. The
fluorescently labelled plasmids were used for transfection of mammalian
cells and their intracellular distribution was visualised by
epifluorescence and confocal fluorescence microscopy. Although these
prokaryotic plasmids do not contain nuclear import sequences,
translocation into the nuclei was observed.

<>

<1>Schmidt, S., Cech, D., Fritz, H.-J.
<2>Enzyme - substrate investigation of dcm methylase.
<3>Biol. Chem. Hoppe Seyler
<4>372
<5>748
<6>1991
<7>To investigate the enzyme-substrate complex of the dcm methylase we synthesized
5-fluoro-2'-deoxycytidine as a substrate analogue. The synthesis was successful by selective
amination of 5-fluoro-2'-deoxyuridine. 5-fluoro-2'-deoxycytidine has not been incorporated
into oligonucleotides chemically. The direct usage of 5-fluoro-2'-deoxycytidine for
oligonucleotide synthesis is made difficult due to the increased acid and alkaline sensitivity
of this monomer. As an alternative we investigated the incorporation of
4-methylmercapto-5-fluoro-2'-deoxyuridine and the subsequently amination of the 4' position.
In principal it is possible to incorporate the 4' methylmercapto derivative into
oligonucleotides using the phosporamidite method. However, during the oxidation step we
observed a partial conversion into 5-fluoro-2'deoxyuridine. Under the assumption that only
the 5-fluoro-2'-deoxycytidine-containing oligonucleotide will be recognized as a substrate we
used the oligonucleotide mixture for our first experiments.

<>

<1>Schmutte, C., Jones, P.A.
<2>Involvement of DNA methylation in human carcinogenesis.
<3>Biol. Chem.
<4>379
<5>377-388
<6>1998
<7>It is now generally accepted that the presence of 5-methylcytosine in human DNA has both a
genetic and an epigenetic effect on cellular development, differentiation and transformation.
First, 5mC is more unstable than its unmethylated counterpart cytosine.  Hydrolytic
deamination of 5mC leads to a G/T mismatch and subsequently, if unrepaired, to a C-T
transition mutation.  Sites of DNA methylation are mutational hotspots in many human tumors.
Second, DNA methylation of promoter regions is often correlated with the down regulation of
the corresponding gene.  Both of these effects have fundamental consequences for basic
functions of the cell like cellular differentiation, the development of cancer and possibly
other diseases, and on the evolutionary process.  Recent hypotheses also propose a role for
methylation in the process of aging.  In this review we will describe recent findings and
hypotheses about the function of 5mC in DNA with the focus on its involvement in human
carcinogenesis.

<>

<1>Schnegg, B., Hofschneider, P.H.
<2>Mutant of PhiX174 accessible to host-controlled modification.
<3>J. Virol.
<4>3
<5>541-542
<6>1969
<7>Host-controlled modification has been shown to occur with several bacteriophage
strains carrying their genetic information on a single-stranded
deoxyribonucleic acid (DNA) molecule.  All these bacteriophage strains (fd, fl,
M13, F12) are related inasmuch as their particles are rod shaped, contain
single-stranded DNA molecules of comparable size, and infect only male
bacteria.  Other single-stranded DNA phages unrelated to this group, such as
PhiX174 and S13, are unable to infect cells of Escherichia coli K-12 and B, so
that their sensitivity to K- and B- host specificity cannot be investigated
directly.  However, using spheroplasts of E. coli and infecting them with
PhiX174 DNA molecules, Benzinger showed that PhiX174 DNA is not restricted by
B- or Pl host specificity.  Lack of restriction of PhiX174 and the related
phage S13 by P1-host specificity has also been shown by Eskridge, Weinfeld, and
Paigen by using the host pair E. coli C and C (P1).  In the experiments
presented here, hybrids were selected from crosses with E. coli C and B (rB+
mB+), and C and K-12 (rK+ mK+), sensitive to infection by PhiX174 and carrying
the genes responsible for B- and K-host specificity, respectively.  With these
hybrids, E. coli BC and KC, respectively, it is shown that wild-type PhiX174 is
not susceptible to modification and restriction controlled by the genes
responsible for lambda DNA modification and restriction.  The findings
concerning B-host specificity thus confirm the results of Benzinger.  However,
a PhiX174 mutant was isolated which is accessible to restriction and
modification in the hybrid E. coli BC.

<>

<1>Schneider-Scherzer, E., Auer, B., de Groot, E.J., Schweiger, M.
<2>Primary structure of a DNA (N6-Adenine)-methyltransferase from Escherichia coli virus T1.
<3>J. Biol. Chem.
<4>265
<5>6086-6091
<6>1990
<7>Escherichia coli virus T1 encodes a DNA (N6-adenine)-methyltransferase (M.T1)
with the same sequence specificity as the E. coli DNA
(N6-adenine)-methyltransferase (M.Eco dam).  This enzyme was purified to
homogeneity and a partial amino acid sequence determined.  Oligonucleotides
were constructed and used not only as probes to map the gene on the T1 genome,
but also as primers in sequencing reactions to establish the nucleotide
sequence of the M.T1 locus by primer extension.  These data represent the first
analysis of the genomic organization of bacterial virus T1 on a molecular
level.  Significant homology to E. coli consensus transcription and
translation-initiation signals suggest that the gene for M.T1 is most probably
under control of its own promoter.  It may be transcribed as a polycistronic
mRNA, together with a downstream open reading frame which codes for a
polypeptide containing 83 amino acids (HP 83).  Both the deduced primary and
the secondary structure of the M.T1 were compared to those of other known DNA
methyltransferases, especially those recognizing the sequence, GATC; there is
little similarity of the T1 enzyme to the other members of this family.

<>

<1>Schneider-Scherzer, E., Auer, B., Schweiger, M.
<2>Identification, purification, characterization and sequencing of DNA-methyltransferase from E. coli virus T1.
<3>Biol. Chem. Hoppe Seyler
<4>369
<5>910
<6>1988
<7>Immediately after infection E. coli virus T1 induces a DNA-methyltransferase,
which methylates the GATC-sites at adenine in vivo and in vitro.  The activity
is associated with a protein of MW 31,000 and can be visualized by activity
analysis in polyacrylamide gels.  Assay conditions were developed which permit
selective measurement of the T1 activity in the presence of E. coli
DNA-methyltransferases.

<>

<1>Schneiker, S. et al.
<2>Complete genome sequence of the myxobacterium Sorangium cellulosum.
<3>Nat. Biotechnol.
<4>25
<5>1281-1289
<6>2007
<7>The genus Sorangium synthesizes approximately half of the secondary metabolites isolated from
myxobacteria, including the anti-cancer
metabolite epothilone. We report the complete genome sequence of the model
Sorangium strain S. cellulosum So ce56, which produces several natural
products and has morphological and physiological properties typical of the
genus. The circular genome, comprising 13,033,779 base pairs, is the
largest bacterial genome sequenced to date. No global synteny with the
genome of Myxococcus xanthus is apparent, revealing an unanticipated level
of divergence between these myxobacteria. A large percentage of the genome
is devoted to regulation, particularly post-translational phosphorylation,
which probably supports the strain's complex, social lifestyle. This
regulatory network includes the highest number of eukaryotic protein
kinase-like kinases discovered in any organism. Seventeen secondary
metabolite loci are encoded in the genome, as well as many enzymes with
potential utility in industry.

<>

<1>Schneiker-Bekel, S., Wibberg, D., Bekel, T., Blom, J., Linke, B., Neuweger, H., Stiens, M., Vorholter, F.J., Weidner, S., Goesmann, A., Puhler, A., Schluter, A.
<2>The complete genome sequence of the dominant Sinorhizobium meliloti field isolate SM11 extends the S. meliloti pan-genome.
<3>J. Biotechnol.
<4>155
<5>20-33
<6>2011
<7>Isolates of the symbiotic nitrogen-fixing species Sinorhizobium meliloti usually contain a
chromosome and two large megaplasmids encoding functions that are absolutely required for the
specific interaction of the microsymbiont with corresponding host plants leading to an
effective symbiosis. The complete genome sequence, including the megaplasmids pSmeSM11c
(related to pSymA) and pSmeSM11d (related to pSymB), was established for the dominant,
indigenous S. meliloti strain SM11 that had been isolated during a long-term field release
experiment with genetically modified S. meliloti strains. The chromosome, the largest replicon
of S. meliloti SM11, is 3,908,022bp in size and codes for 3785 predicted protein coding
sequences. The size of megaplasmid pSmeSM11c is 1,633,319bp and it contains 1760 predicted
protein coding sequences whereas megaplasmid pSmeSM11d is 1,632,395bp in size and comprises
1548 predicted coding sequences. The gene content of the SM11 chromosome is quite similar to
that of the reference strain S. meliloti Rm1021. Comparison of pSmeSM11c to pSymA of the
reference strain revealed that many gene regions of these replicons are variable, supporting
the assessment that pSymA is a major hot-spot for intra-specific differentiation. Plasmids
pSymA and pSmeSM11c both encode unique genes. Large gene regions of pSmeSM11c are closely
related to corresponding parts of Sinorhizobium medicae WSM419 plasmids. Moreover, pSmeSM11c
encodes further novel gene regions, e.g. additional plasmid survival genes (partition,
mobilisation and conjugative transfer genes), acdS encoding 1-aminocyclopropane-1-carboxylate
deaminase involved in modulation of the phytohormone ethylene level and genes having predicted
functions in degradative capabilities, stress response, amino acid metabolism and associated
pathways. In contrast to Rm1021 pSymA and pSmeSM11c, megaplasmid pSymB of strain Rm1021 and
pSmeSM11d are highly conserved showing extensive synteny with only few rearrangements. Most
remarkably, pSmeSM11b contains a new gene cluster predicted to be involved in polysaccharide
biosynthesis. Compilation of the S. meliloti SM11 genome sequence contributes to an extension
of the S. meliloti pan-genome.

<>

<1>Schnetz, K., Rak, B.
<2>Cleavage by EcoO109I and DraII is inhibited by overlapping dcm methylation.
<3>Nucleic Acids Res.
<4>16
<5>1623
<6>1988
<7>We have recently determined the nucleotide sequence of the bgl operon of E. coli. Sequence
analysis revealed the presence of two recognition sites for restriction endonuclease EcoO109I
and its isoschizomer DraII. Both enzymes recognize and cut the sequence RG'GNCCY. However,
digestion of plasmid pFDX733 containing the entire operon with either enzyme resulted in only
linearization (Fig. 1, lanes 1 and 3). We noted that the resistant site overlaps with a dcm
methylation site (CmCT/AGG). The sequence at this site is GGGGCCTGG. When we isolated plasmid
pFDX733 from a Dcm- host, the DNA was efficiently cut with both enzymes at both positions
(Fig. 1, lanes 2 and 4). We conclude that enzymes EcoO109I and DraII are sensitive to
overlapping dcm methylation at their recognition sites.

<>

<1>Schoen, C., Weber-Lehmann, J., Blom, J., Joseph, B., Goesmann, A., Strittmatter, A., Frosch, M.
<2>Whole-Genome Sequence of the Transformable Neisseria meningitidis Serogroup A Strain WUE2594.
<3>J. Bacteriol.
<4>193
<5>2064-2065
<6>2011
<7>Serogroup A meningococci are a leading cause of bacterial meningitis in children and young
adults worldwide. However, the genetic basis of
serogroup A strains' virulence and their epidemiological properties remain
poorly understood. Therefore, we sequenced the complete genome of the
transformable Neisseria meningitidis serogroup A strain WUE2594.

<>

<1>Schoenfeld, T.
<2>Bca77I: A restriction enzyme from Bacillus caldolyticus cleaving the novel hexanucleotide (A/T)^CCGG(A/T) .
<3>Biophys. J.
<4>61
<5>A77
<6>1992
<7>I describe a restriction enzyme, Bca77I, derived from the intermediate
thermophile, Bacillus caldolyticus (Promega strain 77).  The cognate sequence
for this restriction enzyme is the degenerate hexanucleotide, (A/T)CCGG(A/T)
and the cleavage site is 5' of the external cytosine.  The methylation
sensitivity of the endonuclease was examined.  Methylation of the external
cytosine (5mC) completely blocks restriction by Bca77I; methylation of the
internal cytosine (5mC) only decreases marginally the rate of cleavage.  It is
expected that Bca77I will be a valuable tool for the molecular biologist.

<>

<1>Schoenfeld, T.
<2>A Practical Guide to DNA Methylation.
<3>Promega Notes
<4>42
<5>22-27
<6>1993
<7>Most problems attributable to DNA methylation fall into one of two categories: inability to
cut DNA with restriction enzymes; inability to obtain transformants. This article contains
background information that explains why these problems ocur in cloning, followed by
troubleshooting tips that will help you determine if methylation is the problem and if so, how
to solve it. A final Section describes common eukaryotic methylation patterns and how to use
this information to your advantage. DNA methylation refers to the covalent modification of DNA
by transfer of a methyl group from S-adenosylmethionine to one of a few possible sites on
cyosine or adenine. The molecular biologist will primarily encounter m5C methylation of
cytosine and m6N methylation of adenine (Figure 1). The guidelines in this article, however,
also apply to other types of methylation such as m4C methylation of cytosine and m4C
hydroxymethylation of cytosine (1).

<>

<1>Schoenfeld, T.
<2>Bca77I: A restriction enzyme from Bacillus caldolyticus cleaving the novel hexanucleotide (A/T)^CCGG(A/T) .
<3>FASEB J.
<4>6
<5>A77
<6>1992
<7>I describe a restriction enzyme, Bca77I, derived from the intermediate
thermophile, Bacillus caldolyticus (Promega strain 77).  The cognate sequence
for this restriction enzyme is the degenerate hexanucleotide, (A/T)CCGG(A/T)
and the cleavage site is 5' of the external cytosine.  The methylation
sensitivity of the endonuclease was examined.  Methylation of the external
cytosine (m5C) completely blocks restriction by Bca77I; methylation of the
internal cytosine (5mC) only decreases marginally the rate of cleavage.  It is
expected that Bca77I will be a valuable tool for the molecular biologist.

<>

<1>Schoenfeld, T., Fiandt, M., Schink, M.
<2>Bst71I: an isoschizomer of the type-IIS restriction enzyme, BbvI, recognizing the GCAGC(8/12) site.
<3>Gene
<4>111
<5>141-142
<6>1992
<7>A new type-IIS restriction enzyme, Bst71I, with the specificity 5'-GCAGC(N)8
CGTCG(N)12
was isolated from Bacillus stearothermophilus (Promega No. 71).  This enzyme
is an isoschizomer of BbvI with somewhat improved characteristics for use by
molecular biologists.

<>

<1>Schoenfeld, T., King, K.B., Schink, M.
<2>Bca771: a new type-II restriction endonuclease from Bacillus caldolyticus cutting at the (A/T)^CCGG(A/T) site.
<3>Gene
<4>117
<5>99-101
<6>1992
<7>We describe a new restriction enzyme recognizing a degenerate hexanucleotide sequence and
cleaving the 5'-W^CCGGW site (W = A or T). This enzyme cuts with high efficiency all four
permutations of this sequence; ACCGGA (TCCGGT on the opposite stand), ACCGGT and TCCGGA.
Methylation of the external cytosine completely blocks restriction while methylation of the
internal cytosine only decreases the rate of restriction activity.

<>

<1>Schoenfeld, T., Mead, D.A., Fiandt, M.
<2>Purification and characterization of an isoschizomer of AsuII from Clostridium sporogenes.
<3>Nucleic Acids Res.
<4>17
<5>4417
<6>1989
<7>A new type II restriction enzyme, Csp45I, was isolated from Clostridium
sporogenes and characterized.  Digestion of a standard substrate and sequencing
confirmed that both the recognition sequence and the cut site, TT/CGAA, were
the same as those of AsuII and BstBI.

<>

<1>Schoenfeld, T.W., Murugapiran, S., Dodsworth, J.A., Floyd, S., Lodes, M., Mead, D.A., Hedlund, B.P.
<2>Lateral gene transfer of Family A DNA polymerases between thermophilic viruses, Aquificae, and Apicomplexa.
<3>Mol. Biol. Evol.
<4>30
<5>1653-1664
<6>2013
<7>Bioinformatics and functional screens identified a group of Family A-type DNA
Polymerase (polA) genes encoded by viruses inhabiting circumneutral and alkaline
hot springs in Yellowstone National Park and the US Great Basin. The proteins
encoded by these viral polA genes (PolAs) shared no significant sequence
similarity with any known viral proteins but were remarkably similar to PolAs
encoded by two of three families of the bacterial phylum Aquificae and by several
apicoplast-targeted PolA-like proteins found in the eukaryotic phylum
Apicomplexa, which includes the obligate parasites Plasmodium, Babesia, and
Toxoplasma. The viral gene products share signature elements previously
associated only with Aquificae and Apicomplexa PolA-like proteins and were
similar to proteins encoded by prophage elements of a variety of otherwise
unrelated Bacteria, each of which additionally encoded a prototypical bacterial
PolA. Unique among known viral DNA polymerases, the viral PolA proteins of this
study share with the Apicomplexa proteins large amino-terminal domains with
putative helicase/primase elements but low primary sequence similarity. The
genomic context and distribution, phylogeny, and biochemistry of these PolA
proteins suggest that thermophilic viruses transferred polA genes to the
Apicomplexa, likely through secondary endosymbiosis of a virus-infected
proto-apicoplast, and to the common ancestor of two of three Aquificae families,
where they displaced the orthologous cellular polA gene. On the basis of
biochemical activity, gene structure, and sequence similarity, we speculate that
the xenologous viral-type polA genes may have functions associated with
diversity-generating recombination in both Bacteria and Apicomplexa.

<>

<1>Schoettler, S., Christ, F., Pingoud, V., Wende, W., Pingoud, A.
<2>Identification of the catalytic centres of the homing endonuclease PI-SceI by site-directed mutagenesis.
<3>Biochem. Soc. Trans.
<4>28
<5>A332
<6>2000
<7>The homing endonuclease PI-SceI from Saccharomyces cerevisiae is composed of two domains:
Domain I is responsible for protein splicing and specific DNA binding and domain II harbours
the endonuclease function which mediates the mobility of the gene of PI-SceI by introducing a
double strand break into an allele that lacks it.  Cellular repair leads to the integration of
the sequence into this allele.  It was unknown whether the DNA cleavage is due to one
catalytic centre cleaving both DNA strands or two active sites each cleaving one strand.  To
solve this problem we exchanged amino acids in the vicinity of D218 and D326 of the
characteristic LAGLIDADG motifs which were already shown to be essential.  The amino acids
chosen for mutagenesis were selected based on a structural comparison with the homodimeric
enzyme I-CreI.  Results of experiments with substrates with a nick in the cleavage position
are taken as evidence for the existence of two catalytic sites: The variant D229N cleaves a
substrate with a nick in the top strand whereas a substrate with a nick in the bottom strand
is hardly cleaved.  T341N displays an opposite behavior, i.e. prefers to cleave a DNA
substrate nicked in the bottom strand.  Taking together all our results, including
characterization of PI-SceI with respect to DNA binding and cleavage, leads to this model of
DNA strand cleavage: PI-SceI contains two catalytic centres.  In catalytic centre I D218 and
D229 coordinate the metal ion and induce the cleavage of the top strand.  This is the rate
limiting step of cleavage.  The cleavage of the bottom strand is done by active site II
containing D326 as metal ion binding site.  Intriguingly, the mutants D229N and T341N which
are almost inactive in cleaving double stranded DNA can do so when crosslinked to the
substrate.  This could be interpreted to mean that crosslinking fixes the enzyme-substrate
complex in a conformation resembling the transition state, similarly as observed with EcoRV.

<>

<1>Schoettler, S., Wende, W., Pingoud, V., Pingoud, A.
<2>Identification of Asp218 and Asp326 as the principal Mg2+ binding ligands of the homing endonuclease PI-SceI.
<3>Biochemistry
<4>39
<5>15895-15900
<6>2000
<7>The monomeric homing endonuclease PI-SceI harbors two catalytic centers which cooperate in the
cleavage of the two strands of its extended
recognition sequence. Structural and biochemical data suggest that
catalytic center I contains Asp218, Asp229, and Lys403, while catalytic
center II contains Asp326, Thr341, and Lys301. The analogy with I-CreI,
for which the cocrystal structure with the DNA substrate has been
determined, suggests that Asp218 and Asp229 in catalytic center I and
Asp326 and Thr341 in catalytic center II serve as ligands for Mg2+, the
essential divalent metal ion cofactor which can be replaced by Mn2+ in
vitro. We have carried out a mutational analysis of these presumptive
Mg2+ ligands. The variants carrying an alanine or asparagine
substitution bind DNA, but (with the exception of the D229N variant)
are inactive in DNA cleavage in the presence of Mg2+, demonstrating
that these residues are important for cleavage. Our finding that the
PI-SceI variants carrying single cysteine substitutions at these
positions are inactive in the presence of the oxophilic Mg2+ but active
in the presence of the thiophilic Mn2+ suggests that the amino acid
residues at these positions are involved in cofactor binding. From the
fact that in the presence of Mn2+ the D218C and D326C variants are even
more active than the wild-type enzyme, it is concluded that Asp218 and
Asp326 are the principal Mg2+ ligands of PI-SceI. On the basis of these
findings and the available structural information, a model for the
composition of the two Mg2+ binding sites of PI-SceI is proposed.

<>

<1>Scholl, D., Merril, C.
<2>The genome of bacteriophage K1F, a T7-like phage that has acquired the ability to replicate on K1 strains of Escherichia coli.
<3>J. Bacteriol.
<4>187
<5>8499-8503
<6>2005
<7>Bacteriophage K1F specifically infects Escherichia coli strains that
produce the K1 polysaccharide capsule. Like several other K1
capsule-specific phages, K1F encodes an endo-neuraminidase (endosialidase)
that is part of the tail structure which allows the phage to recognize and
degrade the polysaccharide capsule. The complete nucleotide sequence of
the K1F genome reveals that it is closely related to bacteriophage T7 in
both genome organization and sequence similarity. The most striking
difference between the two phages is that K1F encodes the endosialidase in
the analogous position to the T7 tail fiber gene. This is in contrast with
bacteriophage K1-5, another K1-specific phage, which encodes a very
similar endosialidase which is part of a tail gene \"module\" at the end of
the phage genome. It appears that diverse phages have acquired
endosialidase genes by horizontal gene transfer and that these genes or
gene products have adapted to different genome and virion architectures.

<>

<1>Scholl, D.R., Patterson, R.B. Jr., Jollick, J.D.
<2>Modification of EcoRI restriction sites by Cauloacter vibrioides.
<3>Gene
<4>17
<5>163-166
<6>1982
<7>A comparison of EcoRI digestion profiles of plasmid RP1 isolated from
Caulobacter vibrioides WS48 and Escherichia coli CSH29 demonstrated that EcoRI
sites were modified by WS48.

<>

<1>Scholtissek, S., Pingoud, A., Maass, G., Zabeau, M.
<2>Polypeptide sequences involved in the cleavage of DNA by the restriction endonuclease EcoRI.
<3>J. Biol. Chem.
<4>261
<5>2228-2234
<6>1986
<7>We have prepared a variety of fragments of the restriction endonuclease EcoRI
by partial or total CNBr or acid cleavage of the protein.  These fragments were
isolated by preparatvie polyacrylamide gel electrophoresis in the presence of
sodium dodecyl sulfate.  They were analyzed in a qualitative manner for
phosphodiesterase activity.  Antibodies against these fragments were elicited
in rats and tested for binding to native EcoRI in an enzyme-linked immunoassay.
We conclude from these experiments that the DNA binding site of EcoRI is
located in the COOH-terminal half of the molecule, close to and probably
comprising amino acid residues 137 to 157.  This conclusion is reinforced by
the observation that this sequence shows homology to the sequences of the
recognition helix of other gene-regulatory proteins.

<>

<1>Schon, H., Thimm, O., Ritte, G., Blasing, O., Bruynseels, K., Hatzfeld, Y., Frankard, V., Sanz-Molinero, A.I., Reuzeau, C., Vandenabeele, S.
<2>Plants with increased yield (nue).
<3>International Patent Office
<4>WO 2010046221 A
<5>
<6>2010
<7>A method for producing a plant with increased yield as compared to a corresponding wild type
plant whereby the method comprises at least the following step: increasing or generating in a
plant or a part thereof one or more activities selected from the group conisting of 17.6 kDa
class I heat shock protein, 26.5 kDa class I small heat shock protein, 26S protease subunit,
2-Cys peroxiredoxin, 3-dehydroquinate synthase, 5-keto-D-gluconate-5-reductase, asparagine
synthetase A, aspartate 1-decarboxylase precursor, ATP-dependent RNA helicase, BO567-protein,
B1088-protein, B1289-protein, B2940-protein, calnexin homologue, CDS 5399-protein, chromatin
structure-remodeling complex protein, D-amino acid dehydrogenase, D-arabinono-1,4-lactone
oxidase, Delta 1-pyrroline-5-carboxylate reductase, glycine cleavage complex lipoylprotein,
ketodeoxygluconokinase, lipoyl synthase, low-molecular-weight heat-shock protein, Microsomal
cytochrome b reductase, mitochondrial ribosomal protein, mitotic check point protein,
monodehydroascorbate reductase, paraquat-inducible protein B, phosphatase, Phosphoglucosamine
mutase, protein disaggregation chaperone, protein kinase, pyruvate decarboxylase, recA family
protein, rhodanese-related sulfurtransferase, ribonuclease P protein component, ribosome
modulation factor, sensory histodine kinase, serine hydroxymethyltransferase, SLL1280-protein,
SLL1797-protein, small membrane lipoprotein, Small nucleolar ribonucleoprotein complex
subunit, Sulfatase, transcription initiation factor subunit, tretraspanin, tRNA ligase,
xyloglucan galactosyltransferase, YKL130C-protein, YLR443W-protein, YML096W-protein, and zinc
finger family protein-activity.

<>

<1>Schoner, B., Kelly, S., Smith, H.O.
<2>The nucleotide sequence of the HhaII restriction and modification genes from Haemophilus haemolyticus.
<3>Gene
<4>24
<5>227-236
<6>1983
<7>We have determined the nucleotide squence of a cloned 1710-bp segment of Haemophilus
haemolyticus DNA which contains the HhaII restriction (r) and modification (m) genes. The m
gene is 513 bp in length and the r gene is 681 bp in length. Both are in the same reading
frame, being separated by a 21-bp region. A ribosome-binding site is identified in front of
each gene, but no Haemophilus promoter is apparent on the cloned fragment. Transcription
originates from a plasmid promoter and proceeds in the direction m to r.

<>

<1>Schork, S., Schluter, A., Blom, J., Schneiker-Bekel, S., Puhler, A., Goesmann, A., Frosch, M., Schoen, C.
<2>Genome Sequence of a Neisseria meningitidis Capsule Null Locus Strain from the Clonal Complex of Sequence Type 198.
<3>J. Bacteriol.
<4>194
<5>5144-5145
<6>2012
<7>Neisseria meningitidis is a commensal and accidental pathogen exclusively of humans. Although
the production of polysaccharide capsules is considered to be
essential for meningococcal virulence, there have been reports of constitutively
unencapsulated strains causing invasive meningococcal disease (IMD). Here we
report the genome sequence of a capsule null locus (cnl) strain of sequence type
198 (ST-198), which is found in half of the reported cases of IMD caused by cnl
meningococcal strains.

<>

<1>Schott, T., Rossi, M., Hanninen, M.L.
<2>Genome sequence of Helicobacter bizzozeronii strain CIII-1, an isolate from human gastric mucosa.
<3>J. Bacteriol.
<4>193
<5>4565-4566
<6>2011
<7>The canine-adapted Helicobacter bizzozeronii is the only non-pylori Helicobacter species
isolated from human gastric biopsies. Here we present
the genome sequence of the strain CIII-1 isolated from a 45 year-old
female patient with severe gastric symptoms. This is the first genome
sequence of non-pylori gastric Helicobacter isolated from human gastritis.

<>

<1>Schottler, S., Wenz, C., Lanio, T., Jeltsch, A., Pingoud, A.
<2>Protein engineering of the restriction endonuclease EcoRV: structure-guided design of enzyme variants that recognize the base pairs flanking the recognition site.
<3>Eur. J. Biochem.
<4>258
<5>184-191
<6>1998
<7>We generated variants of the restriction endonuclease EcoRV that discriminate between
recognition sites with different flanking sequences.  This was achieved by designing new
contacts to the bases in the major groove of the DNA preceding and following the EcoRV
recognition site.  We selected Ala181 as the starting point for the extension of the site
specificity of EcoRV because, according to the structure of the specific EcoRV DNA complex,
this residue is involved in a water mediated contact with the bases flanking the recognition
sequence on the 5' side.  A substitution of this alanine residue by other amino acid residues
changes the protein-DNA interface in this region and potentially creates new contacts, such
that EcoRV variants could have an extended specificity, i.e. a greater selectivity for EcoRV
sites within a particular sequence context.  EcoRV variants with naturally occurring amino
acid residues at position 181 were produced and their selectivity analyzed with
oligodeoxynucleotide and plasmid substrates that differ only in the base pairs immediately
flanking the EcoRV site.  Some variants, having amino acid residues with long or bulky side
chains at position 181 showed altered preferences for the base pairs flanking the recognition
sequence with oligodeoxynucleotide substrates without losing their catalytic efficiency.  One
variant, A181K, is able to discriminate between purine and pyrimidine bases on the 5' side of
the recognition sequence, probably by means of a new hydrogen bond to the N7 of the purine
base.  Another variant, A181E, strongly prefers a thymine base on the 5' side of the
recognition sequence, presumably due to a hydrogen bond formed between the protonated glutamic
acid residue and the 04 of thymine.

<>

<1>Schouler, C., Clier, F., Luisa, A., Lerayer, L., Ehrlich, S.D., Chopin, M.-C.
<2>A type IC restriction-modification system in Lactococcus lactis.
<3>J. Bacteriol.
<4>180
<5>407-411
<6>1998
<7>Three genes coding for the endonuclease, methylase, and specificity subunits of a type I
restriction-modification (RM) system in the Lactococcus lactis plasmid pIL2614 have been
characterized.  Plasmid location, sequence homologies, and inactivation studies indicated that
this R-M system is most probably of type IC.

<>

<1>Schouler, C., Gautier, M., Ehrlich, S.D., Chopin, M.-C.
<2>Combinational variation of restriction modification specificities in Lactococcus lactis.
<3>Mol. Microbiol.
<4>28
<5>169-178
<6>1998
<7>Three genes coding for a type I R-M system related to the class C enzymes have been identified
on the chromosome of Lactococcus lactis strain IL1403.  In addition, plasmids were found that
encode only the HsdS subunit that directs R-M specificity.  The presence of these plasmids in
IL1403 conferred a new R-M phenotype on the host, indicating that the plasmid-encoded HsdS is
able to interact with the chromosomally encoded HsdR and hsdM subunits.  Such combinational
variation of type I R-M systems may facilitate the evolution of their specificity and thus
reinforce bacterial resistance against invasive foreign unmethylated DNA.

<>

<1>Schreier, H.J., Schott, E.J.
<2>Draft Genome Sequence of the Shellfish Bacterial Pathogen Vibrio sp. Strain B183.
<3>Genome Announcements
<4>2
<5>e00914-14
<6>2014
<7>We report the draft genome sequence of Vibrio sp. strain B183, a Gram-negative marine
bacterium isolated from shellfish that causes mortality in larval
mariculture. The availability of this genome sequence will facilitate the study
of its virulence mechanisms and add to our knowledge of Vibrio sp. diversity and
evolution.

<>

<1>Schreier, H.J., Schott, E.J.
<2>Draft Genome Sequence of the Oyster Larval Probiotic Bacterium Vibrio sp. Strain  OY15.
<3>Genome Announcements
<4>2
<5>e01006-14
<6>2014
<7>We report the draft genome sequence of Vibrio sp. strain OY15, a Gram-negative marine
bacterium isolated from an oyster (Crassostrea virginica) digestive tract
and shown to possess probiotic activity. The availability of this genome sequence
will facilitate the study of the mechanisms of probiotic activity as well as
virulence capacity.

<>

<1>Schroder, J., Glaub, A., Schneider, J., Trost, E., Tauch, A.
<2>Draft Genome Sequence of Corynebacterium bovis DSM 20582, Which Causes Clinical Mastitis in Dairy Cows.
<3>J. Bacteriol.
<4>194
<5>4437
<6>2012
<7>Bovine mastitis represents the most economically important disease in dairy cows  and can be
caused by Corynebacterium bovis, a commensal in the bovine udder. The
draft genome sequence provides insights into the adaptation of this bacterium to
the bovine habitat and its lipolytic capabilities to utilize components of cow's
milk.

<>

<1>Schroeder, C., Jurkschat, H., Meisel, A., Reich, J.G., Kruger, D.
<2>Unusual occurrence of EcoP1 and EcoP15 recognition sites and counterselection of type II methylation and restriction sequences in bacteriophage T7 DNA.
<3>Gene
<4>45
<5>77-86
<6>1986
<7>Selected and counterselected oligodeoxynucleotide sequences were identified in the total
sequence of bacteriophage T7 DNA using a statistical criterion derived for a probability model
of the Markov chain type. All extremely rare tetra- and pentadeoxynucleotides are (or contain)
recognition sequences for the Escherichia coli DNA methylases dam or dcm. Most of the 37
hexadeoxynucleotides absent from T7 DNA are recognition sequences for type II
modification/restriction enzymes of E. coli or related species.  In contrast
to most restriction sites counterselected during evolution, the EcoP1 site GGTCT
occurs 126 times in the T7 genome, and phage T7 replication is severly repressed in
P1-lysogenic host cells. We demonstrate that the frequency of the EcoP1 site is determined by
that of the overlapping recognition sites for T7 primase, an essential phage enzyme. The
recognition site of a type III enzyme, EcoP15, is also not counterselected. In T7 DNA all 36
EcoP15 sites are arranged in such a manner that the sequence CAGCAG is confined to the H
strand, the complementary sequence CTGCTG to the L strand. This "strand bias" is highly
significant and, therefore, very probably selected. A functional relational between this
strand bias and the refractive behaviour of phage T7 to EcoP15 restriction is suspected.

<>

<1>Schroeder, C., Reuter, M., Kruger, D.H.
<2>DNA methylation of T3 virus ocr+ and ocr- strains in Escherichia coli cells harbouring the EcoK DNA host specificity system.
<3>Biomed. Biochim. Acta
<4>43
<5>K1-K5
<6>1984
<7>The influence of the T3 gene functions ocr+ and sam+ on the extent of phage DNA
methylation in Escherichia coli K12 cells was studied by determining the
proportion of 6-methylaminopurine to adenine in the purified DNA of T3
wild-type, sam- and ocr-sam phage strains.  We demonstrate that the DNA of T3
ocr-sam- mutants carries 12 methyl groups as a result of the action of the
host-specificity methylase EcoK.  In contrast to this the DNA of ocr+ strains
is not EcoK-specifically methylated.

<>

<1>Schroeder, D.C., Park, Y., Yoon, H.M., Lee, Y.S., Kang, S.W., Meints, R.H., Ivey, R.G., Choi, T.J.
<2>Genomic analysis of the smallest giant virus--Feldmannia sp. virus 158.
<3>Virology
<4>384
<5>223-232
<6>2009
<7>Genomic analysis of Feldmannia sp. virus 158, the second phaeovirus to be sequenced in hits
entirety, provides further evidence that large double-stranded DNA viruses share similar
evolutionary pressures as cellular organisms.  Reductive evolution is clearly evident within
the phaeoviruses which occurred via several routes: the loss of genes from an ancestral virus
core genome most likely through genetic drift; and as a result of relatively large
recombination events that caused wholesale loss of genes.  The entire genome is 154,641 bp in
lenth and has 150 predicted coding sequences of which 87% have amino acid sequence
similarities to other algal virus coding sequences within the family Phycodnavirdae.
Significant similarities were found, for thirty eight coding sequences (25%), to genes in gene
databanks that are known to be involved in processes that include DNA replication, DNA
methylation, signal transduction, viral integration and transposition, and protein-protein
interactions.  Unsurprisingly, the greatest similarity was observed between the two known
viruses that infect Feldmannia, indicating the taxonomic linkage of these two viruses with
their hosts.  Moreover, comparative analysis of phycodnaviral genomic sequences revealed the
smallest set of core genes (10 out of a possible 31) required to make a functional
nucleocytoplasmic large dsDNA virus.

<>

<1>Schroeder, J.W., Simmons, L.A.
<2>Complete Genome Sequence of Bacillus subtilis Strain PY79.
<3>Genome Announcements
<4>1
<5>e01085-13
<6>2013
<7>Bacillus subtilis is a Gram-positive soil-dwelling and endospore-forming bacterium in the
phylum Firmicutes. B. subtilis strain PY79 is a prototrophic
laboratory strain that has been highly used for studying a wide variety of
cellular pathways. Here, we announce the complete whole-genome sequence of B.
subtilis PY79.

<>

<1>Schroeder, S.G.
<2>Structure-function studies of lima bean trypsin inhibitor and EcoRII methyltransferase.
<3>Diss. Abstr.
<4>61
<5>3052-B
<6>2000
<7>Lima bean trypsin inhibitor studies.  The crystal structure of a stable trypsin inhibitor from
lima bean was determined to 2.5 Angstrom resolution.  The space group is cubic, I213 with
a=110.67 A.  Native Lima Bean Trypsin Inhibitor (LBTI) crystals diffract x-rays to 1.65
Angstrom resolution and yield data that are 93.99% complete.  LBTI has a unique property in
that it is thermally stable to the extent that it can be boiled for ten minutes without
destroying its activity.  This protein also shows a high degree of homology with protease
inhibitors of the Bowman-Birk class: it contains 79 amino acid residues, including 14
cysteines.  Thus, the structure was determined by first building a homology model of LBTI
using adzuki bean trypsin inhibitor and of the molecular replacement method using the
homology-modeled LBTI as a search model.  In this study, the three-dimensional structure of
LBTI is presented and discussed.  Methyltransferase studies.  DNA methylation is believed to
be an important mechanism for DNA recognition, transcription regulation and DNA replication in
bacteria, plants and animals.  EcoRII methyltransferase (M.EcoRII) is a cytosine-C5 DNA
methylating enzyme.  A model of its three-dimensional structure is proposed on the basis of
homology modeling.  Crystal structures of two members of the same family of enzymes, HaeIII
and HhaI methyltransferases (M.HaeIII and M.HhaI respectively), were used as template
molecules.  Molecular dynamics calculations were used to ensure sampling of conformationally
stable structures.  The final model has good geometry.  The DNA and cofactor binding residues
are in expected positions to form proper interactions.  M.EcoRIII is 147 amino acids longer
than the template molecules, and hence the model contains several loops that are significantly
longer than those in M.HaeIII and M.HhaI.  The model provides a framework for interpretation
and designing site-directed mutants that have a potential to improve crystallization
experiments of this enzyme, and other similar enzymes.

<>

<1>Schroeder, S.G., Samudzi, C.T.
<2>Structural studies of EcoRII methylase: exploring similarities among methylases.
<3>Protein Eng.
<4>10
<5>1385-1393
<6>1997
<7>EcoRII methyltransferase is a cytosine-C5 DNA methylating enzyme.  A model of its
three-dimensional structure is proposed on the basis of homology modeling.  Crystal structures
of two members of the same family of enzymes, HaeIII and HhaI methyltransferases (M.HaeIII and
M.HhaI respectively), were used as template molecules.  Molecular dynamics was used to ensure
sampling of conformationally stable structures.  The final model has good geometry.  The DNA
and cofactor binding residues are in expected positions and form proper interactions.
M.EcoRII is 147 amino acids longer than the template molecules, and hence the model contains
several loops that are significantly longer than those in M.HaeIII and M.HhaI.  The model
provides a framework for interpretation and designing site-directed mutants that have a
potential to improve crystallization experiments of this enzyme, and possibly other similar
enzymes.

<>

<1>Schubert, H.L., Blumenthal, R.M., Cheng, X.
<2>Many paths to methyltransfer: a chronicle of convergence.
<3>Trends Biochem. Sci.
<4>28
<5>329-335
<6>2003
<7>S-adenosyl-L-methionine (AdoMet) dependent methyltransferases (MTases) are involved in
biosynthesis, signal transduction, protein repair, chromatin
regulation and gene silencing. Five different structural folds (I-V) have
been described that bind AdoMet and catalyze methyltransfer to diverse
substrates, although the great majority of known MTases have the Class I
fold. Even within a particular MTase class the amino-acid sequence
similarity can be as low as 10%. Thus, the structural and catalytic
requirements for methyltransfer from AdoMet appear to be remarkably
flexible.

<>

<1>Schubert, H.L., Blumenthal, R.M., Cheng, X.
<2>Protein Methyltransferases: Their Distribution Among the Five Structural Classes of AdoMet-Dependent Methyltransferases.
<3>Enzymes
<4>24
<5>3-28
<6>2006
<7>S-adenosyl-L-methionine (AdoMet) dependent methyltransferases (MTases) are involved in
biosynthesis, signal transduction, protein repair,
chromatin regulation, and gene silencing. Five different structural
folds (designated I through V) have been described that bind AdoMet and
catalyze methyltransfer to diverse substrates, although the great
majority of known MTases have the Class I fold. Even within a
particular MTase class the amino-acid sequence similarity can be as low
as 10%. Thus, the structural and catalytic requirements for
methyltransfer from AdoMet appear to be remarkably flexible. MTases
that act on protein substrates have been found to date among three of
the five structural classes (I, the classical fold; III, the corrin
MTase fold; and V, the SET fold).'There are many paths to the top of
the mountain, but the view is always the same.'-Chinese proverb The
Columbia World of Quotations, New York, Columbia University Press, 1996

<>

<1>Schubert, H.L., Phillips, J.D., Hill, C.P.
<2>Structures along the catalytic pathway of PrmC/HemK, an N(5)-glutamine AdoMet-dependent methyltransferase.
<3>Biochemistry
<4>42
<5>5592-5599
<6>2003
<7>Posttranslational methylation of release factors on the glutamine residue of a conserved GGQ
motif is required for efficient termination of protein
synthesis. This methylation is performed by an N(5)-glutamine
methyltransferase called PrmC/HemK, whose crystal structure we report here
at 2.2 A resolution. The electron density at the active site appears to
contain a mixture of the substrates, S-adenosyl-L-methionine (AdoMet) and
glutamine, and the products, S-adenosyl-L-homocysteine (AdoHcy) and
N(5)-methylglutamine. The C-terminal domain of PrmC adopts the canonical
AdoMet-dependent methyltransferase fold and shares structural similarity
with the nucleotide N-methyltransferases in the active site, including use
of a conserved (D/N)PPY motif to select and position the glutamine
substrate. Residues of the PrmC (197)NPPY(200) motif form hydrogen bonds
that position the planar Gln side chain such that the lone-pair electrons
on the nitrogen nucleophile are oriented toward the methyl group of
AdoMet. In the product complex, the methyl group remains pointing toward
the sulfur, consistent with either an sp(3)-hybridized, positively charged
Gln nitrogen, or a neutral sp(2)-hybridized nitrogen in a strained
conformation. Due to steric overlap within the active site, proton loss
and formation of the neutral planar methylamide product are likely to
occur during or after product release. These structures, therefore,
represent intermediates along the catalytic pathway of PrmC and show how
the (D/N)PPY motif can be used to select a wide variety substrates.

<>

<1>Schuch, R., Pelzek, A.J., Fazzini, M.M., Nelson, D.C., Fischetti, V.A.
<2>Complete Genome Sequence of Bacillus cereus Sensu Lato Bacteriophage Bcp1.
<3>Genome Announcements
<4>2
<5>e00334-14
<6>2014
<7>Bacillus cereus sensu lato organisms are an ecologically diverse group that includes etiologic
agents of food poisoning, periodontal disease, and anthrax.
The recently identified Bcp1 bacteriophage infects B. cereus sensu lato and is
being developed as a therapeutic decontamination agent and diagnostic
countermeasure. We announce the complete genome sequence of Bcp1.

<>

<1>Schugerl, K.
<2>Comparison of different reactor designs and performances.
<3>Proc. Ninth Int. Biotech. Symp. Expo., American Chemical Society, Ladisch, M.R., Bose, A., Washington, DC
<4>0
<5>232-235
<6>1992
<7>*
In the chemical industry, the production costs are mainly influenced by chemical and other
running expenditures. The same holds true for the manufacturing of biotechnological bulk
products. In contrast to the high-added-value products, the costs of product separation and
purification are low compared to the product formation costs.
	
In the case of production with aerobic microorganisms, the key factors are the costs for
chemicals (mainly substrate) and energy (including aeration and cooling). The reactor costs
are relatively unimportant. Therefore, for the selection of suitable reactors, not only their
volumetric peformance (e.g., volumetric productivity), but their specific performance with
respect to the key parameters (chemicals, energy) are decisive.

According to industrial practice, the medium composition has a much larger effect on the
process performance than on the reactor type, upon which, however, the optimal medium
composition depends, at least for filamentous molds.

Comparison of reactors and their performances can be carried out on different levels, i.e.,
with regard to
   - the oxygen transfer rate and efficiency,
   - the cell mass productivity and efficiency,
   - the metabolite or enzyme productivity.

The efficiency can be related to the substrate or energy consumption.


<>

<1>Schuldes, J., Rodriguez, O.M., Schmeisser, C., Krishnan, H.B., Daniel, R., Streit, W.R.
<2>Complete Genome Sequence of the Broad-Host-Range Strain Sinorhizobium fredii USDA257.
<3>J. Bacteriol.
<4>194
<5>4483
<6>2012
<7>Here we announce the complete genome sequence of the symbiotic and nitrogen-fixing bacterium
Sinorhizobium fredii USDA257. The genome shares a high
degree of sequence similarity with the closely related broad-host-range strains
S. fredii NGR234 and HH103. Most strikingly, the USDA257 genome encodes a wealth
of secretory systems.

<>

<1>Schultz, P.G.
<2>The interplay between chemistry and biology in the design of enzymatic catalysts.
<3>Science
<4>240
<5>426-433
<6>1988
<7>Chemists and biologist are focusing considerable effort on the development of
efficient, highly selective catalysts for the synthesis or modification of
complex molecules.  Two approaches are described here, the generation of
catalytic antibodies and hybrid enzymes, which exploit the binding and
catalytic machinery of nature in catalyst design.  Characterization of these
systems is providing additional insight into the mechanisms of molecular
recognition and catalysis which may, in turn, lead to the design of tailor-made
catalysts for applications in chemistry, biology and medicine.

<>

<1>Schultz, P.G., Dervan, P.B.
<2>Sequence-specific double-strand cleavage of DNA by penta-N-methylpyrrolecarboxamide-EDTA.Fe(II).
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>80
<5>6834-6837
<6>1983
<7>In the presence of O2 and 5 mM dithiothreitol,
penta-N-methylpyrrolecarboxamide-EDTA.Fe(II) [P5E.Fe(II)] at 0.5 microM cleaves
pBR322 plasmid DNA (50 microM in base pairs) on opposite strands to afford
discrete DNA fragments as analyzed by agarose gel electrophoresis.
High-resolution denaturing gel electrophoresis of a 32P-end-labeled
517-base-pair restriction fragment containing a major cleavage site reveals
that P5E.Fe(II) cleaves 3-5 base pairs contiguous to a 6-base-pair sequence,
5'-T-T-T-T-T-A-3' (4,323-4,328 base pairs).  The major binding orientation of
the pentapeptide occurs with the amino terminus at the adenine side of this
sequence.  In the presence of 5 mM dithiothreitol, 0.01 microM P5E.Fe(II)
converts form I pBR322 DNA at 022 microM plasmid (1.0 mM in base pairs) to 40%
form II, indicating the cleavage reaction is catalytic, turning over a minimum
of nine times.  This synthetic molecule achieves double-strand cleavage of DNA
(pH 7.9, 25C) at the 6-base-pair recognition level and may provide an approach
to the design of "artificial restriction enzymes".

<>

<1>Schulze, C., Jeltsch, A., Franke, I., Urbanke, C., Pingoud, A.
<2>Crosslinking the EcoRV restriction endonuclease across the DNA-binding site reveals transient intermediates and conformational changes of the enzyme during DNA binding and catalytic turnover.
<3>EMBO J.
<4>17
<5>6757-6766
<6>1998
<7>EcoRV completely encircles bound DNA with two loops, forming the entry and exit gate for the
DNA substrate.  These loops were crosslinked generating CL-EcoRV which binds and releases
linear DNA only slowly, because threading linear DNA into and out of the DNA-binding
'tunnel' of CL-EcoRV is not very effective.  If the crosslinking reaction is carried out
with a circular bound DNA, CL-EcoRV is hyperactive towards the trapped substrate which is
cleaved very quickly but not very accurately.  CL-EcoRV also binds to, but does not cleave,
circular DNA when added from the outside, because it cannot enter the active site.  Based on
these results a two-step binding model is proposed for EcoRV: initial DNMA binding occurs at
the outer side of the loops before the gate opens and then the DNA is transferred to the
catalytic center.

<>

<1>Schumann, J., Walter, J., Willert, J., Wild, C., Koch, D., Trautner, T.A.
<2>M.BssHII, a multispecific cytosine-C5-DNA-methyltransferase with unusual target recognizing properties.
<3>J. Mol. Biol.
<4>257
<5>949-959
<6>1996
<7>A new multispecific cytosine-C5-DNA-methyltransferase (C5-MTase), M.BssHII,
was identified in Bacillus stearothermophilus H3.  The M.BssHII gene was cloned and sequenced.
The amino acid sequence deduced shows the characteristic building plan of a C5-MTase.  By
sequencing bisulfite-treated DNA methylated by M.BssHII and by restriction enzyme analysis, we
defined the following methylation targets of M.BssHII: ACGCGT/CCGCGG (Mlu/SacII),
PuGCGCPy (HaeII), PuCCGGPy (Cfr10I) and GCGCGC (BssHII).  The relative location of the
specificity determinants in the C5-MTase was derived from the analysis of M.BssHII derivatives
carrying deletions within the variable region V and chimeric C5-MTases constructed between
M.BssHII and the related monospecific enzyme M.Phi3TII.  Four of the M.BssHII specificities
(MluI, SacII, Cfr10I and BssHII) could be associated with amino acid segments within the
variable region V.  The determinant for HaeII activity had to be assigned to sequences
defining
the enzyme core, the first example of a C5-MTase in which a sequence-specific methylation
potential is mediated by structures outside of the variable region.  Another intriguing result
came
from the analysis of one particular chimera made between M.BssHII and M.Phi3TII.  This
construct showed a relaxation of the methylation capacity, both with respect to the target
recognized and the targeting of methylation within this sequence.

<>

<1>Schumann, J., Willert, J., Wild, C., Waler, J., Trautner, T.A.
<2>M.BssHII: a new multispecific C5-DNA-methyltransferase.
<3>Gene
<4>157
<5>103-104
<6>1995
<7>M.BssHII is a new multispecific C5-DNA-methyltransferase recognizing five different targets.
As the enzyme has been isolated from a thermophilic Bacillus, the protein should show enhanced
intrinsic thermostability and therefore be a promising candidate for crystallizing a
multispecific MTase.

<>

<1>Schuster, A.M., Burbank, D.E., Meister, B., Skrdla, M.P., Meints, R.H., Hattman, S., Swinton, D., Van Etten, J.L.
<2>Characterization of viruses infecting a eukaryotic chlorella-like green alga.
<3>Virology
<4>150
<5>170-177
<6>1986
<7>Nineteen plaque-forming viruses of the unicellular, eukaryotic Chlorella-like
green alga, strain NC64A, were isolated from the various geographic regions in
the United States and characterized.  Like the previously described virus,
PBCV-1, all of the new viruses were large polyhedrons, sensitive to chloroform,
and contained large dsDNA genomes of ca.  300 kbp.  All of the viral DNAs
contained 5-methyldeoxycytidine which varied from 0.1 to 47% of the
deoxycytidine.  In addition, 10 of the viral DNAs contained
N6-methyldeoxyadenosine which varied from 8.1 to 37% of the deoxyadenosine.
These viruses, along with 11 previously described viruses which replicate in
the same Chlorella host, were grouped into 11 classes based on at least one of
the following properties:  plaque size, reaction with PBCV-1 antiserum, or the
nature and abundance of methylated bases in their genomic DNA.

<>

<1>Schutzer, S.E., Fraser-Liggett, C.M., Casjens, S.R., Qiu, W.G., Dunn, J.J., Mongodin, E.F., Luft, B.J.
<2>Whole-Genome Sequences of Thirteen Isolates of Borrelia burgdorferi.
<3>J. Bacteriol.
<4>193
<5>1018-1020
<6>2011
<7>Borrelia burgdorferi is a causative agent of Lyme disease in North America and Eurasia. The
first complete genome sequence of B. burgdorferi strain 31, available for more than a decade,
has assisted research on the
pathogenesis of Lyme disease. Because a single genome sequence is not
sufficient to understand the relationship between genotypic and geographic
variation and disease phenotype, we determined the whole-genome sequences
of 13 additional B. burgdorferi isolates that span the range of natural
variation. These sequences should allow improved understanding of
pathogenesis and provide a foundation for novel detection, diagnosis, and
prevention strategies.

<>

<1>Schutzer, S.E., Fraser-Liggett, C.M., Qiu, W.G., Kraiczy, P., Mongodin, E.F., Dunn, J.J., Luft, B.J., Casjens, S.R.
<2>Whole-Genome Sequences of Borrelia bissettii, Borrelia valaisiana, and Borrelia spielmanii.
<3>J. Bacteriol.
<4>194
<5>545-546
<6>2012
<7>It has been known for decades that human Lyme disease is caused by the three spirochete
species Borrelia burgdorferi, Borrelia afzelii, and
Borrelia garinii. Recently, Borrelia valaisiana, Borrelia spielmanii, and
Borrelia bissettii have been associated with Lyme disease. We report the
complete genome sequences of B. valaisiana VS116, B. spielmanii A14S, and
B. bissettii DN127.

<>

<1>Schwabe, G., Posseckert, G., Klingmuller, W.
<2>Restriction endonucleases in Azospirillum.
<3>Gene
<4>39
<5>113-116
<6>1985
<7>Azospirillum brasilense, A. amazonense, and A. lipoferum strains were screened
for restriction endonucleases using phage lambda DNA.  The extract of A.
brasilense 29711 cleaved lambda DNA into specific fragments.  It was concluded
that this strain possesses a class II restriction endonuclease which was named
AbrI.  AbrI has a single recognition site on lambda DNA at position of approx.
33500 bp. AbrI was characterized as an isoschizomer of XhoI, which cuts lambda
DNA at 33498 bp and cleaves double-stranded DNA at the sequence 5'-C^TCGAG-3'.
From other Azospirilla strains only A. amazonense QRZ42 extracts (AamI
activity) cleaved DNA into specific fragments under certain conditions.

<>

<1>Schwartz, E., Henne, A., Cramm, R., Eitinger, T., Friedrich, B., Gottschalk, G.
<2>Complete nucleotide sequence of pHG1: a Ralstonia eutropha H16 megaplasmid encoding key enzymes of H(2)-based ithoautotrophy and anaerobiosis.
<3>J. Mol. Biol.
<4>332
<5>369-383
<6>2003
<7>The self-transmissible megaplasmid pHG1 carries essential genetic information for the
facultatively lithoautotrophic and facultatively
anaerobic lifestyles of its host, the Gram-negative soil bacterium
Ralstonia eutropha H16. We have determined the complete nucleotide
sequence of pHG1. This megaplasmid is 452,156 bp in size and carries 429
potential genes. Groups of functionally related genes form loose clusters
flanked by mobile elements. The largest functional group consists of
lithoautotrophy-related genes. These include a set of 41 genes for the
biosynthesis of the three previously identified hydrogenases and of a
fourth, novel hydrogenase. Another large cluster carries the genetic
information for denitrification. In addition to a dissimilatory nitrate
reductase, both specific and global regulators were identified. Also
located in the denitrification region is a set of genes for cytochrome c
biosynthesis. Determinants for several enzymes involved in the
mineralization of aromatic compounds were found. The genes for conjugative
plasmid transfer predict that R.eutropha forms two types of pili. One of
them is related to the type IV pili of pathogenic enterobacteria. pHG1
also carries an extensive "junkyard" region encompassing 17 remnants of
mobile elements and 22 partial or intact genes for phage-type integrase.
Among the mobile elements is a novel member of the IS5 family, in which
the transposase gene is interrupted by a group II intron.

<>

<1>Schwarz, F.W., Toth, J., van Aelst, K., Cui, G., Clausing, S., Szczelkun, M.D., Seidel, R.
<2>The helicase-like domains of type III restriction enzymes trigger long-range diffusion along DNA.
<3>Science
<4>340
<5>353-356
<6>2013
<7>Helicases are ubiquitous adenosine triphosphatases (ATPases) with widespread roles in genome
metabolism.  Here, we report a previously undescribed functionality for ATPases with
helicase-like domains; namely, that ATP hydrolysis can trigger ATP-independent long-range
protein diffusion on DNA in one dimension (1D).  Specifically, using single-molecule
fluorescence microscopy we show that the Type III restriction enzyme EcoP15I uses its ATPase
to switch into a distinct structural state that diffuses on DNA over long distances and long
times.  The switching occurs only upon binding to the target site and requires hydrolysis of
~30 ATPs.  We define the mechanism for these enzymes and show how ATPase activity is involved
in DNA target site verification and 1D signaling, roles that are common in DNA metabolism: for
example, in nucleotide excision and mismatch repair.

<>

<1>Schwarz, F.W., Toth, J., van Aelst, K., Cui, G., Szczelkun, M.D., Seidel, R.
<2>Type III Restriction Enzymes Use 1D Diffusion to Communicate the Relative Orientation of their Distant Target Sites.
<3>Biophys. J.
<4>100
<5>191
<6>2011
<7>Type III restriction enzymes sense that relative orientation of their distant target sites and
cleave DNA only if at least two of them are situated in an inverted repeat.  The communication
process is strictly dependent on ATP hydrolysis catalyzed by their superfamily 2 helicase
domains.  Given the similarity to Type I restriction enzymes, which couple ATP hydrolysis to
directed motion on DNA, unidirectional loop translocation that may partially be accommpanied
by 3D diffusive looping has  been the suggested communication mechanism for Type III enzymes.
Based on magnetic tweezers single-molecule cleavage experiments and ATPase measurements we
suggest an alternative inter-site communication mechanism using 1D diffusion along the DNA
contour.  In order to verify this hypothesis we directly visualize the motion of quantum-dot
labeled Type III restriction enzymes along DNA.  For this we use a setup that combines
magnetic tweezers with total internal reflection fluorescence microscopy.  The enzymes undergo
a fast diffusive motion along DNA capable of scanning kbp distances per second.  We also find
that the affinity of the enzymes to non-specific and specific DNA is regulated by the presence
of ATP suggesting that ATP hydrolysis acts as a trigger for diffusion.  Thus, Type III
restricotn enzymes are the first DNa-mopdifying enzymes which communicate target site
orientations over long distances via 1D diffusion.

<>

<1>Schwarz, F.W., van Aelst, K., Toth, J., Seidel, R., Szczelkun, M.D.
<2>DNA cleavage site selection by Type III restriction enzymes provides evidence for head-on protein collisions following 1D bidirectional motion.
<3>Nucleic Acids Res.
<4>39
<5>8042-8051
<6>2011
<7>DNA cleavage by the Type III Restriction-Modification enzymes requires communication in 1D
between two distant indirectly-repeated recognitions
sites, yet results in non-specific dsDNA cleavage close to only one of the
two sites. To test a recently proposed ATP-triggered DNA sliding model, we
addressed why one site is selected over another during cleavage. We
examined the relative cleavage of a pair of identical sites on DNA
substrates with different distances to a free or protein blocked end, and
on a DNA substrate using different relative concentrations of protein.
Under these conditions a bias can be induced in the cleavage of one site
over the other. Monte-Carlo simulations based on the sliding model
reproduce the experimentally observed behaviour. This suggests that
cleavage site selection simply reflects the dynamics of the preceding
stochastic enzyme events that are consistent with bidirectional motion in
1D and DNA cleavage following head-on protein collision.

<>

<1>Schweikart, K.M., Ribeiro, E., Larcom, L.L.
<2>Analysis of the inhibition of restriction endonuclease cleavage by UV damage.
<3>Photochem. Photobiol.
<4>45
<5>76s
<6>1987
<7>It has been shown that thymine dimers in the recognition sequence of a
restriction endonuclease inhibit cleavage of the substrate.  Quantitative gel
electrophoresis analysis allows evaluation of the extent to which cleavage is
inhibited at different sites in the same DNA.  Since the recognition sequence
is the same at all sites, any differences in sensitivity to cleavage must
result from differences in the base sequences near the recognition sites.  We
have observed different amounts of cleavage inhibition for the EcoRI sites in
lambda DNA.  The amount of inhibition is correlated with the probability of
dimer formation within 10 bases on either side of the cleavage site.
Endonucleases BamHI, EcoRI and HaeII were found to be insensitive to damage by
254 nm UV.  DNA complexed with EcoRI was protected from UV damage by the bound
protein.

<>

<1>Schwibbert, K., Marin-Sanguino, A., Bagyan, I., Heidrich, G., Lentzen, G., Seitz, H., Rampp, M., Schuster, S.C., Klenk, H.-P., Pfeiffer, F., Oesterhelt, D., Kunte, H.J.
<2>A blueprint of ectoine metabolism from the genome of the Industrial producer Halomonas elongata DSM 2581(T).
<3>Environ. Microbiol.
<4>13
<5>1973-1994
<6>2010
<7>The halophilic g-proteobacterium Halomonas elongata DSM 2581T thrives at high salinity by
synthesizing and accumulating the compatible solute ectoine.  Ectoine levels are highly
regulated according to external
salt levels but the overall picture of its metabolism and control is not well understood.
Apart from its critical role in cell adaptation to halophilic environments, ectoine can be
used as a stabilizer for enzymes and as a cell protectant in skin and health care applications
and is thus produced annually on a scale of tons in an industrial process using H. elongata as
producer strain. This paper presents the complete genome sequence of H. elongata (4 061 296
bp) and includes experiments and analysis identifying and characterizing the entire ectoine
metabolism, including a newly discovered pathway for ectoine degradation and its cyclic
connection to ectoine synthesis.  The degradation of ectoine (doe) proceeds via hydrolysis of
ectoine (DoeA) to Na-acetyl-L-2,4-diaminobutyric acid, followed by deacetylation to
diaminobutyric acid (DoeB). In H. elongata, diaminobutyric acid can either flow off to
aspartate or re-enter the ectoine synthesis pathway, forming a cycle of ectoine synthesis and
degradation. Genome comparison revealed that the ectoine degradation pathway exists
predominantly in non-halophilic bacteria unable to synthesize ectoine. Based on the resulting
genetic and biochemical data, a metabolic flux model of ectoine metabolism was derived that
can be used to understand the way H. elongata survives under varying salt stresses and that
provides a basis for a model-driven improvement of industrial ectoine production.

<>

<1>Schwinghamer, E.A.
<2>Host-controlled modification of Rhizobium bacteriophage.
<3>Aust. J. Biol. Sci.
<4>18
<5>333-343
<6>1964
<7>Host-controlled phenotypic variation of host specificity was observed with two
rhizobiophage strains, OL1 and OL5, following growth on six strains of
Rhizobium leguminosarum and R. trifolii.  The six hosts could be assigned to
four groups, each group representing a different pattern of host specificity.
Initial adaptation of OL5 to hosts L2, L7, and L25 appeared to involve
mutation, although replication in cells of these hosts generally involved
additional phenotypic restriction.  Restricted and unrestricted forms of a
phage did not differ significantly in their ability to adsorb to several hosts.
One-step analysis of the L25-specific form of OL5 grown in L25 cells indicated
a low average burst size of approximately one unrestricted plaque-forming unit
in a small proportion of cells which were able to produce infective centres on
L4.  One-cycle analysis of L4-specific OL5 modified by growth in L25 confirmed
the phenotypic nature of phage variation in this phage-host system, and
indicated that specificity for L4 was not replicated in L25.

<>

<1>Schwudke, D., Ergin, A., Michael, K., Volkmar, S., Appel, B., Knabner, D., Konietzny, A., Strauch, E.
<2>Broad-host-range Yersinia phage PY100: genome sequence, proteome analysis of virions, and DNA packaging strategy.
<3>J. Bacteriol.
<4>190
<5>332-342
<6>2008
<7>PY100 is a lytic bacteriophage with a broad host range within the genus
Yersinia. The phage forms plaques on strains of the three human pathogenic
species Yersinia enterocolitica, Y. pseudotuberculosis, and Y. pestis at
37 degrees C. PY100 was isolated from farm manure and intended to be used
in phage therapy trials. PY100 has an icosahedral capsid containing
double-stranded DNA and a contractile tail. The genome consists of 50,291
bp and is predicted to contain 93 open reading frames (ORFs). PY100 gene
products were found to be homologous to the capsid proteins and proteins
involved in DNA metabolism of the enterobacterial phage T1; PY100 tail
proteins possess homologies to putative tail proteins of phage AaPhi23 of
Actinobacillus actinomycetemcomitans. In a proteome analysis of virion
particles, 15 proteins of the head and tail structures were identified by
mass spectrometry. The putative gene product of ORF2 of PY100 shows
significant homology to the gene 3 product (small terminase subunit) of
Salmonella phage P22 that is involved in packaging of the concatemeric
phage DNA. The packaging mechanism of PY100 was analyzed by hybridization
and sequence analysis of DNA isolated from virion particles. Newly
replicated PY100 DNA is cut initially at a pac recognition site, which is
located in the coding region of ORF2.

<>

<1>Schyns, G., Serra, C.R., Lapointe, T., Pereira-Leal, J.B., Potot, S., Fickers, P., Perkins, J.B., Wyss, M., Henriques, A.O.
<2>Genome of a Gut Strain of Bacillus subtilis.
<3>Genome Announcements
<4>1
<5>e00184-12
<6>2013
<7>Bacillus subtilis is a Gram-positive, rod-shaped, spore-forming bacterium. We present the
genome sequence of an undomesticated strain, BSP1, isolated from
poultry. The sequence of the BSP1 genome supports the view that B. subtilis has a
biphasic lifestyle, cycling between the soil and the animal gastrointestinal
tract, and it provides molecular-level insight into the adaptation of B. subtilis
to life under laboratory conditions.

<>

<1>Sclair, M., Edgell, M.H., Hutchison, C.A.
<2>Mapping of new Escherichia coli K and 15 restriction sites on specific fragments of bacteriophage PhiX174 DNA.
<3>J. Virol.
<4>11
<5>378-385
<6>1973
<7>We have isolated several new PhiX174 mutants which contain sites sensitive to
restriction by Escherichia coli.  One contains an E. coli 15 restriction site
and three are double mutants containing an E. coli K site as well as the E.
coli 15 site.  The replicative form (RF) DNA of one of the mutants containing a
K site has been shown to be restricted in spheroplasts of a K-12 strain.  The
infectivity of this RF, but not wild-type RF, has also been shown to be
inactivated by an E. coli K extract and by purified K restriction enzyme in
vitro.  The product of the RF treated with purified K restriction enzyme in
vitro is a full length linear molecule.  The mutant sites have also been
localized to specific regions of the PhiX174 genome by a fragment mapping
technique, making use of specific fragments of PhiX174 RF DNA obtained by
digestion with a specific endonuclease.

<>

<1>Scott, A., Song, J., Ewing, R., Wang, Z.
<2>Regulation of protein stability of DNA methyltransferase 1 by post-translational modifications.
<3>Acta Biochim. Biophys. Sin.
<4>46
<5>199-203
<6>2014
<7>DNA methylation is an important epigenetic mechanism that ensures correct gene expression and
maintains genetic stability. DNA methyltransferase 1 (DNMT1) is the primary enzyme that
maintains DNA methylation during replication. Dysregulation of DNMT1 is implicated in a
variety of diseases. DNMT1 protein stability is regulated via various post-translational
modifications, such as acetylation and ubiquitination, but also through protein-protein
interactions. These mechanisms ensure DNMT1 is properly activated during the correct time of
the cell cycle and at correct genomic loci, as well as in response to appropriate
extracellular cues. Further understanding of these regulatory mechanisms may help to design
novel therapeutic approaches for human diseases.

<>

<1>Scott, K.M. et al.
<2>The Genome of Deep-Sea Vent Chemolithoautotroph Thiomicrospira crunogena XCL-2.
<3>PLoS Biology
<4>4
<5>e383
<6>2006
<7>Presented here is the complete genome sequence of Thiomicrospira crunogena XCL-2,
representative of ubiquitous chemolithoautotrophic sulfur-oxidizing
bacteria isolated from deep-sea hydrothermal vents. This
gammaproteobacterium has a single chromosome (2,427,734 base pairs), and
its genome illustrates many of the adaptations that have enabled it to
thrive at vents globally. It has 14 methyl-accepting chemotaxis protein
genes, including four that may assist in positioning it in the redoxcline.
A relative abundance of coding sequences (CDSs) encoding regulatory
proteins likely control the expression of genes encoding carboxysomes,
multiple dissolved inorganic nitrogen and phosphate transporters, as well
as a phosphonate operon, which provide this species with a variety of
options for acquiring these substrates from the environment. Thiom.
crunogena XCL-2 is unusual among obligate sulfur-oxidizing bacteria in
relying on the Sox system for the oxidation of reduced sulfur compounds.
The genome has characteristics consistent with an obligately
chemolithoautotrophic lifestyle, including few transporters predicted to
have organic allocrits, and Calvin-Benson-Bassham cycle CDSs scattered
throughout the genome.

<>

<1>Sczesnak, A., Segata, N., Qin, X., Gevers, D., Petrosino, J.F., Huttenhower, C., Littman, D.R., Ivanov, I.I.
<2>The genome of th17 cell-inducing segmented filamentous bacteria reveals extensive auxotrophy and adaptations to the intestinal environment.
<3>Cell Host Microbe
<4>10
<5>260-272
<6>2011
<7>Perturbations of the composition of the symbiotic intestinal microbiota can have
profound consequences for host metabolism and immunity. In mice, segmented
filamentous bacteria (SFB) direct the accumulation of potentially proinflammatory
Th17 cells in the intestinal lamina propria. We present the genome sequence of
SFB isolated from monocolonized mice, which classifies SFB phylogenetically as a
unique member of Clostridiales with a highly reduced genome. Annotation analysis
demonstrates that SFB depend on their environment for amino acids and essential
nutrients and may utilize host and dietary glycans for carbon, nitrogen, and
energy. Comparative analyses reveal that SFB are functionally related to members
of the genus Clostridium and several pathogenic or commensal "minimal" genera,
including Finegoldia, Mycoplasma, Borrelia, and Phytoplasma. However, SFB are
functionally distinct from all 1200 examined genomes, indicating a gene
complement representing biology relatively unique to their role as a gut
commensal closely tied to host metabolism and immunity.

<>

<1>Sears, A., Peakman, L.J., Wilson, G.G., Szczelkun, M.D.
<2>Characterization of the Type III restriction endonuclease PstII from Providencia stuartii.
<3>Nucleic Acids Res.
<4>33
<5>4775-4787
<6>2005
<7>A new Type III restriction endonuclease designated PstII has been purified from Providencia
stuartii. PstII recognizes the hexanucleotide sequence 5'-CTGATG(N)(25-26/27-28)-3'.
Endonuclease activity requires a substrate with two copies of the recognition site in
head-to-head repeat and is dependent on a low level of ATP hydrolysis ( approximately 40
ATP/site/min). Cleavage occurs at just one of the two sites and results in a staggered cut
25-26 nt downstream of the top strand sequence to generate a two base 5'-protruding end.
Methylation of the site occurs on one strand only at the first adenine of 5'-CATCAG-3'.
Therefore, PstII has characteristic Type III restriction enzyme activity as exemplified by
EcoPI or EcoP15I. Moreover, sequence asymmetry of the PstII recognition site in the T7 genome
acts as an historical imprint of Type III restriction activity in vivo. In contrast to other
Type I and III enzymes, PstII has a more relaxed nucleotide specificity and can cut DNA with
GTP and CTP (but not UTP). We also demonstrate that PstII and EcoP15I cannot interact and
cleave a DNA substrate suggesting that Type III enzymes must make specific protein-protein
contacts to activate endonuclease activity.

<>

<1>Sears, A., Szczelkun, M.D.
<2>Subunit assembly modulates the activities of the Type III restriction-modification enzyme PstII in vitro.
<3>Nucleic Acids Res.
<4>33
<5>4788-4796
<6>2005
<7>We demonstrate that, like other Type III restriction endonuclease, PstII does not turnover
such that a DNA substrate is only fully cleaved at a
Res2Mod2-to-site ratio of approximately 1:1. However, unlike other Type
III enzymes, the cleavage rate profiles varied with protein concentration:
using 5 nM DNA and 25 nM PstII, approximately half of the DNA was cut at a
fast rate while the remainder was cut 24 times more slowly; in comparison,
with 100 nM PstII cleavage occurs at a single fast rate. The inclusion of
the methyl donor S-adenosyl methionine does not alter the rates with 100
nM PstII but with 25 nM PstII the reaction stopped after completion of the
initial fast cleavage phase owing to methylation. Concentration-dependent
rates were also observed in methylation assays: at 100 nM PstII, a single
slow rate was measured while at lower PstII concentrations both fast and
slow rates were measured. We propose a model in which the intact Res2Mod2
complex favoured at high PstII concentrations is a fast endonuclease/slow
methyltransferase while the various subassemblies which coexist at lower
concentrations are fast methyltransferases. A potential role for
disassembly in control of restriction activity in vivo is discussed.

<>

<1>Sears, L.E., Zhou, B., Aliotta, J.M., Morgan, R.D., Kong, H.
<2>BaeI, another unusual BcgI-like restriction endonuclease.
<3>Nucleic Acids Res.
<4>24
<5>3590-3592
<6>1996
<7>BcgI and BcgI-like restriction endonucleases have a very distinct characteristic which causes
them to differ from the other classified restriction enzymes; they all cleave double-stranded
DNA specifically on both sides of the recognition sites.  Here we report a new BcgI-like
restriction endonuclease, BaeI, isolated from Bacillus sphaericus.  Like BcgI, BaeI also
cleaves double-stranded DNA on both strands upstream and downstream of its recognition
sequence (10/15)ACNNNNGTAYC(12/7).  There are two dominant polypeptides in the final
preparation of BaeI with molecular masses of ~80 and 55 kDa.  Both are slightly larger than
the two BcgI subunits.  BaeI requires both Mg2+ and AdoMet to cleave DNA.  Accompanying
bilateral cleavage activity, the heteromeric BaeI also has an N6-adenine methyltransferase
activity which modifies the symmetrically located adenines within its recognition sequence.

<>

<1>Sebaihia, M. et al.
<2>Comparison of the genome sequence of the poultry pathogen Bordetella avium with those of B. bronchiseptica, B. pertussis, and B. parapertussis reveals extensive diversity in surface structures associated with host interaction.
<3>J. Bacteriol.
<4>188
<5>6002-6015
<6>2006
<7>Bordetella avium is a pathogen of poultry and is phylogenetically distinct from Bordetella
bronchiseptica, Bordetella pertussis, and Bordetella parapertussis, which are other species in
the Bordetella genus that infect mammals. In order to understand the evolutionary relatedness
of Bordetella species and further the understanding of pathogenesis, we obtained the complete
genome sequence of B. avium strain 197N, a pathogenic strain that has been extensively
studied. With 3,732,255 base pairs of DNA and 3,417 predicted coding sequences, it has the
smallest genome and gene complement of the sequenced bordetellae. In this study, the presence
or absence of previously reported virulence factors from B. avium was confirmed, and the
genetic bases for growth characteristics were elucidated. Over 1,100 genes present in B. avium
but not in B. bronchiseptica were identified, and most were predicted to encode surface or
secreted proteins that are likely to define an organism adapted to the avian rather than the
mammalian respiratory tracts. These include genes coding for the synthesis of a polysaccharide
capsule, hemagglutinins, a type I secretion system adjacent to two very large genes for
secreted proteins, and unique genes for both lipopolysaccharide and fimbrial biogenesis. Three
apparently complete prophages are also present. The BvgAS virulence regulatory system appears
to have polymorphisms at a poly(C) tract that is involved in phase variation in other
bordetellae. A number of putative iron-regulated outer membrane proteins were predicted from
the sequence, and this regulation was confirmed experimentally for five of these.

<>

<1>Sebaihia, M. et al.
<2>The multidrug-resistant human pathogen Clostridium difficile has a highly mobile, mosaic genome.
<3>Nat. Genet.
<4>38
<5>779-786
<6>2006
<7>We determined the complete genome sequence of Clostridium difficile strain 630, a virulent and
multidrug-resistant strain. Our analysis indicates
that a large proportion (11%) of the genome consists of mobile genetic
elements, mainly in the form of conjugative transposons. These mobile
elements are putatively responsible for the acquisition by C. difficile of
an extensive array of genes involved in antimicrobial resistance,
virulence, host interaction and the production of surface structures. The
metabolic capabilities encoded in the genome show multiple adaptations for
survival and growth within the gut environment. The extreme genome
variability was confirmed by whole-genome microarray analysis; it may
reflect the organism's niche in the gut and should provide information on
the evolution of virulence in this organism.

<>

<1>Sebaihia, M. et al.
<2>Genome sequence of a proteolytic (Group I) Clostridium botulinum strain Hall A and comparative analysis of the clostridial genomes.
<3>Genome Res.
<4>17
<5>1082-1092
<6>2007
<7>Clostridium botulinum is a heterogeneous Gram-positive species that comprises four genetically
and physiologically distinct groups of bacteria that share the
ability to produce botulinum neurotoxin, the most poisonous toxin known to man,
and the causative agent of botulism, a severe disease of humans and animals. We
report here the complete genome sequence of a representative of Group I
(proteolytic) C. botulinum (strain Hall A, ATCC 3502). The genome consists of a
chromosome (3,886,916 bp) and a plasmid (16,344 bp), which carry 3650 and 19
predicted genes, respectively. Consistent with the proteolytic phenotype of this
strain, the genome harbors a large number of genes encoding secreted proteases
and enzymes involved in uptake and metabolism of amino acids. The genome also
reveals a hitherto unknown ability of C. botulinum to degrade chitin. There is a
significant lack of recently acquired DNA, indicating a stable genomic content,
in strong contrast to the fluid genome of Clostridium difficile, which can form
longer-term relationships with its host. Overall, the genome indicates that C.
botulinum is adapted to a saprophytic lifestyle both in soil and aquatic
environments. This pathogen relies on its toxin to rapidly kill a wide range of
prey species, and to gain access to nutrient sources, it releases a large number
of extracellular enzymes to soften and destroy rotting or decayed tissues.

<>

<1>Sebaihia, M., Bocsanczy, A.M., Biehl, B.S., Quail, M.A., Perna, N.T., Glasner, J.D., Declerck, G.A., Cartinhour, S., Schneider, D.J., Bentley, S.D., Parkhill, J., Beer, S.V.
<2>Complete Genome Sequence Of The Plant Pathogen Erwinia amylovora Strain ATCC 49946.
<3>J. Bacteriol.
<4>192
<5>2020-2021
<6>2010
<7>Erwinia amylovora causes the economically important disease fire blight that affects rosaceous
plants, especially pear and apple. Here we report the complete genome sequence and annotation
of strain ATCC 49946. The analysis of the sequence and its comparison with sequenced genomes
of closely related enterobacteria revealed signs of pathoadaptation to rosaceous hosts.

<>

<1>Sebastiani, H., Jager, H.
<2>Bacteriophages of Streptococcus-salivarius ssp thermophilus: Characterization of hereditary relationships and determination of virus-host interaction.
<3>Milchwissenschaft
<4>48
<5>25-29
<6>1992
<7>Problems due to bacteriophage infection are widespread in all dairy fermentations. Since the
pioneering work of Whitehead and Cox, numerous studies in the past 50 years have led to a
higher degree in the control of starter activity. However, most of the research in this area
focussed on bacteriophages of mesophilic streptococci, which are now characterized quite well
down to the genomic level. On the other hand very little is known about the phages of
thermophilic streptococci. Up to now these viruses were mainly characterized on the basis of
ultrastructure and their host range. Problems like the frequency of temperate phages and the
risk of lysogenic strains in fermentation, as well as the spontaneous change of some host
strains in their sensitivity to bacteriophages or phage-defense mechanisms of Streptococcus
salivarius ssp. thermophilus are as poorly understood as the genome of the bacteriophage, but
demand a solution in order to isolate or construct stable phage-resistant starter cultures for
the dairy industry.

<>

<1>Sechler, J.M.
<2>Use of restriction endonucleases against viruses, including HIV.
<3>US Patent Office
<4>US 5523232
<5>
<6>1996
<7>Restriction endonucleases are administered to a patient topically or
internally to prevent or ameliorate viral infection.  The endonucleases are also useful to
treat
products, such as blood and blood derived products, to elminate viral transmission when
the products are administered to a patient.  In addition, the endonucleases may be used in
the manner of a disinfectant to inactivate viral contamination on objects.  The endonucleases
are used in their native form, modified to provide decreased immunoreactivity or increased
ability to target a site of infection or both, and alone or in combination with a suitable
excipient.  In addition to methods of treatment and disinfection, the invention provides
related products, such as pharmaceutical compositions.

<>

<1>Sechler, J.M.G., Clouse, K.A., Strebel, K., Weih, K.A., Rosenberg, A.S.
<2>Effect of restriction endonucleases on HIV infection.
<3>J. Cell Biochem. Suppl.
<4>17D
<5>76
<6>1993
<7>HIV RNA undergoes obligatory reverse transcription to a dsDNA form in the cytoplasm of
infected cells. This step is critical for the establishment of infection. However, no
treatment has yet been devised to target the dsDNA. If the dsDNA form of the virus could be
destroyed before translocating to the nucleus and integrating into the host genome, then
perhaps amplifiation of the virus within the organism might be prevented and disease
progression halted. Bacterial cells possess DNA site-specific restriction-modification systems
that provide protection from viral infections. The type II restriction endonucleases recognize
a particular sequence in DNA and cleave the DNA in the vicinity of that sequence. We
investigated whether the type II restriction endonucleases could modify the course of HIV
infection in human PBL. Studies performed to date indicate that enzymes known to cleave the
dsDNA form of the virus inhibit the development of RT activity in cultures of infected human
PBL whereas a control enzyme which lacks a restriction site on HIV dsDNA fails to inhibit
virus replication. The replication kinetics were delayed and the absolute RT level achieved in
the treated cultures was reduced. Additionally, morphologic changes associated with infection,
such as syncytia formation, were significantly delayed. Of critical importance, the
restriction endonucleases do not appear to damage host cell DNA as shown by their failure to
inhibit proliferation of activated human PBL in vitro. Thus, type II restriction endonucleases
may prove useful in the study and treatment of HIV infection.

<>

<1>Seeber, S., Kessler, C., Gotz, F.
<2>Cloning, expression and characterization of the Sau3AI restriction and modification genes in Staphylococcus carnosus TM300.
<3>Gene
<4>94
<5>37-43
<6>1990
<7>The genes encoding the restriction enzyme (ENase) and modification enzyme
(MTase) of Staphylococcus aureus 3A (recognition sequence 5'-GATC-3') have been
cloned in Staphylococcus carnosus TM300 using the vector pCA44.  Clones
carrying both genes were isolated from DNA libraries prepared with MboI +
BamHI.  The DNA region encoding M.Sau3AI was subcloned on a 3.66-kb EcoRV
fragment in vector pT181mcs.  Plasmids purified from the clones were resistant
to digestion with Sau3AI, indicating that the sau3AIM gene was expressed and
the product was functional in S. carnosus.  Cell lysates of clones with both
activities encoded on plasmid pSEM7, cut DNA with the same pattern as Sau3AI,
showing that the sau3AIR gene was also expressed and the ENase was functional
in S. carnosus.  Sequence analysis shows that both genes are transcribed in the
same direction and encode polypeptides with calculated Mrs of 56477 for
R.Sau3AI and 47300 for M.Sau3AI.  Efforts to clone one or both genes in
Escherichia coli have so far failed.

<>

<1>Seed, K.D., Bodi, K.L., Kropinski, A.M., Ackermann, H.W., Calderwood, S.B., Qadri, F., Camilli, A.
<2>Evidence of a Dominant Lineage of Vibrio cholerae-Specific Lytic Bacteriophages Shed by Cholera Patients over a 10-Year Period in Dhaka, Bangladesh.
<3>MBio
<4>2
<5>e00334-10
<6>2011
<7>ABSTRACT Lytic bacteriophages are hypothesized to contribute to the seasonality and duration
of cholera epidemics in Bangladesh. However, the bacteriophages contributing to this
phenomenon have yet to be characterized at a molecular genetic level. In this study, we
isolated and sequenced the genomes of 15 bacteriophages from stool samples from cholera
patients spanning a 10-year surveillance period in Dhaka, Bangladesh. Our results indicate
that a single novel bacteriophage type, designated ICP1 (for the International Centre for
Diarrhoeal Disease Research, Bangladesh cholera phage 1) is present in all stool samples from
cholera patients, while two other bacteriophage types, one novel (ICP2) and one T7-like
(ICP3), are transient. ICP1 is a member of the Myoviridae family and has a 126-kilobase genome
comprising 230 open reading frames. Comparative sequence analysis of ICP1 and related isolates
from this time period indicates a high level of genetic conservation. The ubiquitous presence
of ICP1 in cholera patients and the finding that the O1 antigen of lipopolysaccharide (LPS)
serves as the ICP1 receptor suggest that ICP1 is extremely well adapted to predation of
human-pathogenic V. cholerae O1. IMPORTANCE The severe diarrheal disease cholera is caused by
the bacterium Vibrio cholerae, which can be transmitted to humans from the aquatic
environment. Factors that affect V. cholerae in the environment can impact the occurrence of
cholera outbreaks; one of these factors is thought to be the presence of bacterial viruses, or
bacteriophages. Bacteriophages that prey on V. cholerae in the environment, and potentially in
humans, have not been extensively genetically characterized. Here, we isolated and sequenced
the genomes of bacteriophages from cholera patient stool samples collected over a 10-year
period in Dhaka, Bangladesh, a region that suffers from regular cholera outbreaks. We describe
a unique bacteriophage present in all samples, infer its evolution by sequencing multiple
isolates from different patients over time, and identify the host receptor that shows that the
bacteriophage specifically predates the serogroup of V. cholerae responsible for the majority
of disease occurrences.

<>

<1>Seed, K.D., Dennis, J.J.
<2>Isolation and characterization of bacteriophages of the Burkholderia cepacia complex.
<3>FEMS Microbiol. Lett.
<4>251
<5>273-280
<6>2005
<7>The Burkholderia cepacia complex consists of nine phenotypically similar
but genotypically distinct beta-proteobacteria that are metabolically
diverse and highly antibiotic resistant. Because of this exceptional
intrinsic antibiotic resistance, infections with B. cepacia complex
members are difficult to treat clinically and new alternative therapies
are required. One strategy that holds some promise is the use of naturally
occurring antibacterial bacteriophages that could potentially bind to and
lyse B. cepacia complex cells in vivo. Towards that end, we used
enrichment techniques to isolate lytic and lysogenic bacteriophages
specific to the B. cepacia complex. The newly isolated bacteriophages were
characterized by host range analysis, electron microscopy, genome
restriction analysis, and partial DNA sequencing. These isolates include a
bacteriophage with one of the broadest host ranges yet identified for any
bacteriophage specific to the B. cepacia complex, and the first
description of bacteriophages capable of lysing B. ambifaria.

<>

<1>Seedorf, H., Fricke, W.F., Veith, B., Bruggemann, H., Liesegang, H., Strittmatter, A., Miethke, M., Buckel, W., Hinderberger, J., Li, F., Hagemeier, C., Thauer, R.K., Gottschalk, G.
<2>The genome of Clostridium kluyveri, a strict anaerobe with unique metabolic features.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>2128-2133
<6>2008
<7>Clostridium kluyveri is unique among the clostridia; it grows anaerobically on ethanol and
acetate as sole energy sources. Fermentation products are butyrate, caproate, and H(2). We
report here the genome sequence of C. kluyveri, which revealed new insights into the metabolic
capabilities of this well studied organism. A membrane-bound energy-converting NADH:ferredoxin
oxidoreductase (RnfCDGEAB) and a cytoplasmic butyryl-CoA dehydrogenase complex (Bcd/EtfAB)
coupling the reduction of crotonyl-CoA to butyryl-CoA with the reduction of ferredoxin
represent a new energy-conserving module in anaerobes. The genes for NAD-dependent ethanol
dehydrogenase and NAD(P)-dependent acetaldehyde dehydrogenase are located next to genes for
microcompartment proteins, suggesting that the two enzymes, which are isolated together in a
macromolecular complex, form a carboxysome-like structure. Unique for a strict anaerobe, C.
kluyveri harbors three sets of genes predicted to encode for polyketide/nonribosomal peptide
synthetase hybrides and one set for a nonribosomal peptide synthetase. The latter is predicted
to catalyze the synthesis of a new siderophore, which is formed under iron-deficient growth
conditions.

<>

<1>Seegers, J.F.M.L., van Sinderen, D., Fitzgerald, G.F.
<2>Molecular characterization of the lactococcal plasmid pCIS3: natural stacking of specificity subunits of a type I restriction/modification system in a single lactococcal strain.
<3>Microbiology
<4>146
<5>435-443
<6>2000
<7>A 6.1 kb plasmid from the Lactococcus lactis subsp. cremoris strain UC509.9. named pCIS3, was
found to mediate a restriction/modification
(RIM) phenotype, Nucleotide sequence analysis of pCIS3 revealed the
presence of an hsdS gene, typical of type I R/M systems, The presence
of this plasmid resulted in a 10(4)-fold reduction in the efficiency of
plating (e.o.p.) of unmodified phage. In addition to the hsdS gene of
pCIS3, two more hsd5 genes were identified in strain UC509.9, one
located on the chromosome downstream of a gene highly homologous to
hsdM genes and a third on the smallest (4 kb) plasmid, named pCIS1. The
replication region of pCIS3 was highly similar to that of a large
family of lactococcal theta replicons, In addition, pCIS3 was found to
encode a member of the CorA family of magnesium transporters.

<>

<1>Seela, F., Driller, H.
<2>Palindromic oligonucleotides containing 7-deaza-2'-deoxyguanosine: solid-phase synthesis of d[(p)GG*AATTCC] octamers and recognition by the endodeoxyribonuclease EcoRI.
<3>Nucleic Acids Res.
<4>14
<5>2319-2332
<6>1986
<7>Octadeoxynucleotides with the sequence d[(p)GG*AATTCC] have been prepared by
solid-phase synthesis employing regular and base-modified phosphoramidites.
These oligomers which contain an isosterically altred recognition sequence of
the endodeoxyribonuclease EcoRI form duplexes under appropriate salt
conditions.  Since G* can represent 7-deaza-2'-deoxyguanosine the oligomers
were used as probes to study their cleavage by the endodeoxyribonuclease EcoRI.
The enzymatic hydrolysis of the modified octamer was strongly decreased
compared to the regular DNA-fragment.  This shows that quanine N-7 located at
the cleavage site is important for the recognition process by the enzyme.  The
residual enzymatic activity is discussed on the basis of reduced specificity
towards the recognition fragment.  The fact that this cleavage occurs already
under regular conditions indicates that the process described here bases on an
intrinsic property of the oligomer and is different from the star activity.

<>

<1>Seela, F., Kehne, A.
<2>Palindromic  octa-  and  dodecanucleotides containing 2'-deoxytubercidin: synthesis,  hairpin formation, and recognition by the endodeoxyribonuclease EcoRI.
<3>Biochemistry
<4>26
<5>2232-2238
<6>1987
<7>Octa- and dodecanucleotides containing 2'-deoxytubericidin within the
endodeoxyribonuclease   EcoRI  recognition  fragment  d(GAATTC)  have  been
prepared  by  solid-phase synthesis. Whereas octamers as well as dodecamers
with  a  "random"  flanking region formed duplexes in aqueous solution, the
dodecamer  d(CGCGAATTCGCG)  and  isosterically  modified  oligomers thereof
showed  a  strong tendency of hairpin formation. Due to this, cleavage with the
endodeoxyribonuclease EcoRI was strongly decreased.  In contrast,
d(GTAGAATTCTAC) was easily cleaved by the enzyme. Single replacement of one of
the  dA  residues by 2'-deoxytubercidin within the recognition sequence
decreased   the   cleavage   velocity  but  retained  specificity.  Twofold
modification  prevents cleavage of the oligomer. This implies that both N-7
purine  nitrogens  are  proton acceptor sites for the endodeoxyribonuclease
EcoRI.

<>

<1>Seela, F., Roling, A.
<2>7-Deazapurine containing DNA: efficiency of c7GdTP, c7AdTP and c7IdTP incorporation during PCR-amplification and protection from endodeoxyribonuclease hydrolysis.
<3>Nucleic Acids Res.
<4>20
<5>55-61
<6>1992
<7>The enzymatic synthesis of 7-deazapurine nucleoside containing DNA (501 bp) is performed by
PCR-amplification (Taq polymerase) using a pUC18 plasmid DNA as template and the triphosphates
of 7-deaza-2'-deoxyguanosine (c7Gd), -adenosine (c7Ad) and -inosine (c7Id). c7GdTP can fully
replace dGTP resulting in a completely modified DNA-fragment of defined size and sequence. The
other two 7-deazapurine triphosphates (c7AdTP) and (c7IdTP) require the presence of the parent
purine 2-deoxyribonucleotides. In purine/7-deazapurine nucleotide mixtures Taq polymerase
prefers purine over 7-deazapurine nucleotides but accepts c7GdTP much better than c7AdTP or
c7IdTP. As incorporation of 7-deazapurine nucleotides represents a modification of the major
groove of DNA it can be used to probe DNA/protein interaction. Regioselective phosphodiester
hydrolysis of the modified DNA-fragments was studied with 28 endodeoxyribonucleases. c7Gd is
able to protect the DNA from the phosphodiester hydrolysis in more than 20 cases, only a few
enzymes (MaeIII, RsaI, HindIII, PvuII or TaqI) do still hydrolyze the modified DNA. c7Ad
protects DNA less efficiently, as this DNA could only be modified in part. The absence of N-7
as potential binding position or a geometric distortion of the recognition duplex caused by
the 7-deazapurine base can account for protection of hydrolysis.

<>

<1>Seeman, N.C., Rosenberg, J.M., Rich, A.
<2>Sequence-specific recognition of double helical nucleic acids by proteins.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>73
<5>804-808
<6>1976
<7>The base pairs in double helical nucleic acids have been compared to see how
they can be recognized by proteins.  We conclude that a single hydrogen bond is
inadequate for uniquely identifying any particular base pair, as this leads to
numerous degeneracies.  However, using two hydrogen bonds, fidelity of base
pair recognition may be achieved.  We propose specific amino-acid side chain
interactions involving two hydrogen bonds as a component of the recognition
system for base pairs.  In the major groove we suggest that asparagine or
glutamine binds to adenine of the base pair, or arginine binds to guanine.  In
the minor groove, we suggest an interaction between asparagine or glutamine
with guanine of the base pair.  We also discuss the role that ions and other
amino-acid side chains may play in recognition interactions.

<>

<1>Segata, N., Ballarini, A., Jousson, O.
<2>Genome Sequence of Pseudomonas aeruginosa PA45, a Highly Virulent Strain Isolated from a Patient with Bloodstream Infection.
<3>Genome Announcements
<4>1
<5>e00289-13
<6>2013
<7>Pseudomonas aeruginosa is a ubiquitous opportunistic pathogen causing a broad range of
infections in humans. We provide the draft genome sequence of the
recently identified and highly virulent P. aeruginosa PA45 strain. Its 6.6-Mb
genome contains 6,822 genes, including an unparalleled number of virulence genes,
which might explain its aggressive phenotype.

<>

<1>Seib, K.L., Peak, I.R., Jennings, M.P.
<2>Phase variable restriction-modification systems in Moraxella catarrhalis.
<3>FEMS Immunol. Med. Microbiol.
<4>32
<5>159-165
<6>2002
<7>A repetitive DNA motif was used as a marker to identify novel genes in the mucosal pathogen
Moraxella catarrhalis. There is a high prevalence of such repetitive motifs in virulence genes
that display phase variable expression. Two repeat containing loci were identified using a
digoxigenin-labelled 5'-(CAAC)(6)-3' oligonucleotide probe. The repeats are located in the
methylase components of two distinct type III restriction-modification (R-M) systems. We
suggest that the phase variable nature of these R-M systems indicates that they have an
important role in the biology of M. catarrhalis.

<>

<1>Seidel, R., Bloom, J.G., Dekker, C., Szczelkun, M.D.
<2>Motor step size and ATP coupling efficiency of the dsDNA translocase EcoR124I.
<3>EMBO J.
<4>27
<5>1388-1398
<6>2008
<7>The Type I restriction-modification enzyme EcoR124I is an archetypical helicase-based dsDNA
translocase that moves unidirectionally along the
30-50 strand of intact duplex DNA. Using a combination of ensemble and
single-molecule measurements, we provide estimates of two
physicochemical constants that are fundamental to a full description of
motor protein activity-the ATP coupling efficiency (the number of ATP
consumed per base pair) and the step size (the number of base pairs
transported per motor step). Our data indicate that EcoR124I makes
small steps along the DNA of 1 bp in length with 1 ATP consumed per
step, but with some uncoupling of the ATPase and translocase cycles
occurring so that the average number of ATP consumed per base pair
slightly exceeds unity. Our observations form a framework for
understanding energy coupling in a great many other motors that
translocate along dsDNA rather than ssDNA.

<>

<1>Seidel, R., Szczelkun, M.D.
<2>Switching roles for a helicase.
<3>Cell Cycle
<4>12
<5>3125-3126
<6>2013
<7>Helicases are widespread enzymes that share a core RecA fold and characteristic amino acid
motifs which together form an ATP binding/hydrolysis site.  For the classical helicases (i.e.,
those which confirm to the family name), ATP-binding provides energy to drive the separation
of polynucleotide duplexes into single strands.  There are also many enzymes which on the
basis of structure and motifs appear to be helicases, but which fulfill alternative functions
without the necessity for strand separation.  These include nucleo-protein remodeling,
long-range motion along dsDNA and molecular signaling.  The activities and mechanisms of these
enzymes, often termed "helicase-like" or "pseudo-helicases", are not well understood.  Recent
studies that try to elucidate the full repertoire of helicase activity provide more and more
surprises for their numerous roles in genome metabolism.

<>

<1>Seidel, R., van der Scheer, C., van Noort, J., Firman, K., Dekker, C.
<2>Translocation of type I restriction endonuclease EcoR124I along DNA measured with magnetic tweezers.
<3>Biophys. J.
<4>86
<5>316a
<6>2004
<7>Type I restriction enzymes belong to the machinery of bacteria that protects them against
invasion by viruses. After sequence specific
binding to viral DNA they translocate up to several thousands of bp and
induce DNA cleavage remote from the recognition site. Using magnetic
tweezers, we measured the translocation process of this enzyme. A
magnetic particle is used to stretch DNA in an applied magnetic field.
By tracking the displacement of the particle, translocation activity
can be recorded in real time. We found that the enzyme is moving with a
constant speed as fast as 550+30 bp/s. By varying the applied force
the translocation velocity, the maximum translocated distance as well
as the time between translocation events, could be measured as a
function of the load exerted to the tethered DNA molecule.

<>

<1>Seidel, R., van Noort, J., van der Scheer, C., Bloom, J.G.P., Dekker, N.H., Dutta, C.F., Blundell, A., Robinson, T., Firman, K., Dekker, C.
<2>Real-time observation of DNA translocation by the type I restriction modification enzyme EcoR124I.
<3>Nat. Struct. Mol. Biol.
<4>11
<5>838-843
<6>2004
<7>Type I restriction enzymes bind sequence-specifically to unmodified DNA and subsequently pull
the adjacent DNA toward themselves. Cleavage then
occurs remotely from the recognition site. The mechanism by which these
members of the superfamily 2 (SF2) of helicases translocate DNA is
largely unknown. We report the first single-molecule study of DNA
translocation by the type I restriction enzyme EcoR124I.
Mechanochemical parameters such as the translocation rate and
processivity, and their dependence on force and ATP concentration, are
presented. We show that the two motor subunits of EcoR124I work
independently. By using torsionally constrained DNA molecules, we found
that the enzyme tracks along the helical pitch of the DNA molecule.
This assay may be directly applicable to investigating the tracking of
other DNA-translocating motors along their DNA templates.

<>

<1>Seipke, R.F., Crossman, L., Drou, N., Heavens, D., Bibb, M.J., Caccamo, M., Hutchings, M.I.
<2>Draft Genome Sequence of Streptomyces Strain S4, a Symbiont of the Leaf-Cutting Ant Acromyrmex octospinosus.
<3>J. Bacteriol.
<4>193
<5>4270-4271
<6>2011
<7>Streptomyces spp. are common symbionts of the leaf-cutting ant species Acromyrmex
octospinosus, which feeds on basidiomycete fungus leaf matter
and harvests the lipid- and carbohydrate-rich gongylidia as a food source.
A. octospinosus and other ant genera use antifungal compounds produced by
Streptomyces spp. and other actinomycetes in order to help defend their
fungal gardens from parasitic fungi. Herein, we report the draft genome
sequence of Streptomyces strain S4, an antifungal-producing symbiont of A.
octospinosus.

<>

<1>Seito, K.K.
<2>Novel restriction enzyme CbiI - which recognises the specified base configuration in DNA, obtained by culturing Clostridium bifermentans B-4.
<3>Japanese Patent Office
<4>JP 2215379 A
<5>
<6>1990
<7>Novel restriction enzyme CbiI, has the following properties: recognising the base
configuration TT^CGAA in double-stranded DNA and cutting as indicated; optimum pH 5.5 to 7.0;
stable pH range 4.0 to 7.5; optimum temperature 30 to 40 degrees C; stable temperature 55
degrees C (100% enzymatic activity can be kept after 5 mins. heating at this temperature);
stable salt concn. range 150mM (no sodium chloride nor salt requirement); mol.wt. 49000 (gel
filtration and SDS polyacrylamide gel electrophoresis). The enzyme is obtained by cultivation
of Clostridium bifermentans B-4 (FERM 10518).

<>

<1>Sekiguchi, H., Komiya, K., Kiga, D., Yamamura, M.
<2>A design and feasibility study of reactions comprising DNA molecular machine that walks autonomously by using a restriction enzyme.
<3>Nat. Comput.
<4>7
<5>303-315
<6>2008
<7>In this paper, we propose an autonomous molecular walking machine using DNA. This molecular
machine follows a track of DNA equipped with many single-strand DNA stators arranged in a
certain pattern. The molecular machine achieves autonomous walk by using a restriction enzyme
as source of power. With a proposed machine we can control its moving direction and we can
easily extend walking patterns in two or three dimensions. Combination of multiple legs and
ssDNA stators can control the walking pattern. We designed and performed a series of
feasibility study with computer simulation
and molecular biology experiments.

<>

<1>Sekine, M., Tanikawa, S., Omata, S., Saito, M., Fujisawa, T., Tsukatani, N., Tajima, T., Sekigawa, T., Kosugi, H., Matsuo, Y., Nishiko, R., Imamura, K., Ito, M., Narita, H., Tago, S., Fujita, N., Harayama, S.
<2>Sequence analysis of three plasmids harboured in Rhodococcus erythropolis strain PR4.
<3>Environ. Microbiol.
<4>8
<5>334-346
<6>2006
<7>Rhodococcus erythropolis strain PR4 has been isolated as an alkane-degrading bacterium. The
strain harbours one linear plasmid, pREL1
(271 577 bp) and two circular plasmids, pREC1 (104 014 bp) and pREC2 (3637
bp), all with some sequence similarities to other Rhodococcus plasmids.
For pREL1, pREC1 and pREC2, 298, 102 and 3 open reading frames,
respectively, were predicted. Linear plasmid pREL1 has several regions
homologous to plasmid pBD2 found in R. erythropolis BD2. Sequence analysis
of pREL1 and pBD2 identified common metal-resistance genes on both, but
pREL1 also encodes alkane-degradation genes not found on pBD2, with enzyme
constituents some of which are quite different from those of other
organisms. The alkane hydroxylase consisted of a cytochrome P450
monooxygenase, a 2Fe-2S ferredoxin, and a ferredoxin reductase. The
ferredoxin reductase amino acid sequence resembles the AlkT (rubredoxin
reductase) sequence. A zinc-containing alcohol dehydrogenase further
oxydizes alkanols, alkane oxidation products catalysed by alkane
hydroxylase. Of the circular plasmids, the pREC1 sequence is partially
similar to the sequence of pREAT701, the virulence plasmid found in
Rhodococcus equi. pREC1 has no pREAT701 virulence genes and encodes genes
for beta-oxidation of fatty acids. Thus, joint actions of enzymes encoded
by pREL1 and pREC1 may enable efficient mineralization of alkanes.

<>

<1>Sekizaki, T., Osaki, M., Takamatsu, D., Shimoji, Y.
<2>Distribution of the SsuDAT1I restriction-modification system among different serotypes of Streptococcus suis.
<3>J. Bacteriol.
<4>183
<5>5436-5440
<6>2001
<7>The SsuDAT1I restriction-modification (R-M) system, which contains two methyltransferases and
two restriction endonucleases with recognition sequence 5'-GATC-3', was first found in a
field isolate of Streptococcus suis serotype 2. Isoschizomers of the R-M system were found in
the same locus between purH and purD in a field isolate of serotype 1/2 and the reference
strains of serotypes 3, 7, 23, and 26 among 29 strains of different serotypes examined in this
study. The R-M gene sequences in serotypes 1/2, 3, 7, and 23 were very similar to those of
SsuDAT1I, whereas those in serotype 26 were less similar. These results indicate intraspecies
recombination among them and genetic divergence through their evolution.

<>

<1>Sekizaki, T., Otani, Y., Osaki, M., Takamatsu, D., Shimoji, Y.
<2>Evidence for horizontal transfer of SsuDAT1I restriction-modification genes to the Streptococcus suis genome.
<3>J. Bacteriol.
<4>183
<5>500-511
<6>2001
<7>Different strains of Streptococcus suis serotypes 1 and 2 isolated from pigs either contained
a restriction-modification (R-M) system or lacked it. The R-M system was an isoschizomer of
Streptococcus pneumoniae DpnII, which recognizes the nucleotide sequence 5'-GATC-3'. The
nucleotide sequencing of the genes encoding the R-M system in S. suis DAT1, designated
SsuDAT1I, showed that the SsuDAT1I gene region contained two methyltransferase genes,
designated ssuMA and ssuMB, as does the DpnII system. The deduced amino acid sequences of
M.SsuMA and M.SsuMB showed 70 and 90% identity to M.DpnII and M.DpnA, respectively. However,
the SsuDAT1I system contained two isoschizomeric restriction endonuclease genes, designated
ssuRA and ssuRB. The deduced amino acid sequence of R.SsuRA was 49% identical to that of
R.DpnII, and R.SsuRB was 72% identical to R.LlaDCHI of Lactococcus lactis subsp. cremoris
DCH-4. The four SsuDAT1I genes overlapped and were bounded by purine biosynthetic gene
clusters in the following gene order: purF-purM-purN-purH-ssuMA-ssuMB-ssuRA-ssuRB-purD-purE.
The G+C content of the SsuDAT1I gene region (34.1%) was lower than that of the pur region
(48.9%), suggesting horizontal transfer of the SsuDAT1I system. No transposable element or
long-repeat sequence was found in the flanking regions. The SsuDAT1I genes were functional by
themselves, as they were individually expressed in Escherichia coli. Comparison of the
sequences between strains with and without the R-M system showed that only the region from 53
bp upstream of ssuMA to 5 bp downstream of ssuRB was inserted in the intergenic sequence
between purH and purD and that the insertion target site was not the recognition site of
SsuDAT1I. No notable substitutions or insertions could be found, and the structures were
conserved among all the strains. These results suggest that the SsuDAT1I system could have
been integrated into the S. suis chromosome by an illegitimate recombination mechanism.

<>

<1>Sekizaki, T., Takamatsu, D., Osaki, M., Shimoji, Y.
<2>Different foreign genes incidentally integrated into the same locus of the Streptococcus suis genome.
<3>J. Bacteriol.
<4>187
<5>872-883
<6>2005
<7>Some strains of Streptococcus suis possess a type II restriction-modification (RM) system,
whose genes are thought to be
inserted into the genome between purH and purD from a foreign source by
illegitimate recombination. In this study, we characterized the purHD
locus of the S. suis genomes of 28 serotype reference strains by DNA
sequencing. Four strains contained the RM genes in the locus, as
described before, whereas 11 strains possessed other genetic regions of
seven classes. The genetic regions contained a single gene or multiple
genes that were either unknown or similar to hypothetical genes of
other bacteria. The mutually exclusive localization of the genetic
regions with the atypical G+C contents indicated that these regions
were also acquired from foreign sources. No transposable element or
long-repeat sequence was found in the neighboring regions. An alignment
of the nucleotide sequences, including the RM gene regions, suggested
that the foreign regions were integrated by illegitimate recombination
via short stretches of nucleotide identity. By using a thermosensitive
suicide plasmid, the RM genes were experimentally introduced into an S.
suis strain that did not contain any foreign genes in that locus.
Integration of the plasmid into the S. suis genome did not occur in the
purHD locus but occurred at various chromosomal loci, where there were
2 to 10 bp of nucleotide identity between the chromosome and the
plasmid. These results suggest that various foreign genes described
here were incidentally integrated into the same locus of the S. suis
genome.

<>

<1>Sekizuka, T., Kai, M., Nakanaga, K., Nakata, N., Kazumi, Y., Maeda, S., Makino, M., Hoshino, Y., Kuroda, M.
<2>Complete genome sequence and comparative genomic analysis of Mycobacterium massiliense JCM 15300 in the Mycobacterium abscessus group reveal a conserved genomic island MmGI-1 related to putative lipid metabolism.
<3>PLoS ONE
<4>9
<5>e114848
<6>2014
<7>Mycobacterium abscessus group subsp., such as M. massiliense, M. abscessus sensu stricto and
M. bolletii, are an environmental organism found in soil, water and other ecological niches,
and have been isolated from respiratory tract infection, skin and soft tissue infection,
postoperative infection of cosmetic surgery. To determine the unique genetic feature of M.
massiliense, we sequenced the complete genome of M. massiliense type strain JCM 15300
(corresponding to CCUG 48898). Comparative genomic analysis was performed among Mycobacterium
spp. and among M. abscessus group subspp., showing that additional ß-oxidation-related genes
and, notably, the mammalian cell entry (mce) operon were located on a genomic island, M.
massiliense Genomic Island 1 (MmGI-1), in M. massiliense. In addition, putative anaerobic
respiration system-related genes and additional mycolic acid cyclopropane synthetase-related
genes were found uniquely in M. massiliense. Japanese isolates of M. massiliense also
frequently possess the MmGI-1 (14/44, approximately 32%) and three unique conserved regions
(26/44; approximately 60%, 34/44; approximately 77% and 40/44; approximately 91%), as well as
isolates of other countries (Malaysia, France, United Kingdom and United States). The
well-conserved genomic island MmGI-1 may play an important role in high growth potential with
additional lipid metabolism, extra factors for survival in the environment or synthesis of
complex membrane-associated lipids. ORFs on MmGI-1 showed similarities to ORFs of
phylogenetically distant M. avium complex (MAC), suggesting that horizontal gene transfer or
genetic recombination events might have occurred within MmGI-1 among M. massiliense and MAC.

<>

<1>Sekizuka, T., Lee, K., Kuroda, M., Kusumoto, M., Iwata, T., Uchida, I., Tanaka, K., Tamamura, Y., Akiba, M.
<2>Whole-Genome Sequence of CMY-2 beta-Lactamase-Producing Salmonella enterica Serovar Typhimurium Strain L-3553.
<3>Genome Announcements
<4>2
<5>e00711-14
<6>2014
<7>Salmonella enterica serovar Typhimurium pulsed-field gel electrophoresis cluster  VII has been
isolated from cattle populations in Japan since the mid-2000s. Some
cluster VII isolates exhibited extended-spectrum cephalosporin resistance defined
by the blaCMY-2 gene located in a chromosomal genomic island, GI-VII-6. We
determined the whole-genome sequence of strain L-3553 as the reference strain.

<>

<1>Sekizuka, T., Matsui, M., Yamane, K., Takeuchi, F., Ohnishi, M., Hishinuma, A., Arakawa, Y., Kuroda, M.
<2>Complete sequencing of the blaNDM-1-positive IncA/C plasmid from Escherichia coli ST38 isolate suggests a possible origin from plant pathogens.
<3>PLoS ONE
<4>6
<5>e25334
<6>2011
<7>The complete sequence of the plasmid pNDM-1_Dok01 carrying New Delhi metallo-b-lactamase
(NDM-1) was determined
by whole genome shotgun sequencing using Escherichia coli strain NDM-1_Dok01 (multilocus
sequence typing type: ST38)
and the transconjugant E. coli DH10B. The plasmid is an IncA/C incompatibility type composed
of 225 predicted coding
sequences in 195.5 kb and partially shares a sequence with blaCMY-2-positive IncA/C plasmids
such as E. coli AR060302
pAR060302 (166.5 kb) and Salmonella enterica serovar Newport pSN254 (176.4 kb). The blaNDM-1
gene in pNDM-1_Dok01 is
terminally flanked by two IS903 elements that are distinct from those of the other
characterized NDM-1 plasmids,
suggesting that the blaNDM-1 gene has been broadly transposed, together with various mobile
elements, as a cassette gene.
The chaperonin groES and groEL genes were identified in the blaNDM-1-related composite
transposon, and phylogenetic
analysis and guanine-cytosine content (GC) percentage showed similarities to the homologs of
plant pathogens such as
Pseudoxanthomonas and Xanthomonas spp., implying that plant pathogens are the potential source
of the blaNDM-1 gene.
The complete sequence of pNDM-1_Dok01 suggests that the blaNDM-1 gene was acquired by a novel
composite transposon
on an extensively disseminated IncA/C plasmid and transferred to the E. coli ST38 isolate.

<>

<1>Sekizuka, T., Yamamoto, A., Komiya, T., Kenri, T., Takeuchi, F., Shibayama, K., Takahashi, M., Kuroda, M., Iwaki, M.
<2>Corynebacterium ulcerans 0102 carries the gene encoding diphtheria toxin on a prophage different from the C. diphtheriae NCTC 13129 prophage.
<3>BMC Microbiol.
<4>12
<5>72
<6>2012
<7>ABSTRACT: BACKGROUND: Corynebacterium ulcerans can cause a diphtheria-like
illness, especially when the bacterium is lysogenized with a tox gene-carrying
bacteriophage that produces diphtheria toxin. Acquisition of toxigenicity upon
phage lysogenization is a common feature of C. ulcerans and C. diphtheriae.
However, because of a lack of C. ulcerans genome information, a detailed
comparison of prophages has not been possible between these two clinically
important and closely related bacterial species. RESULTS: We determined the whole
genome sequence of the toxigenic C. ulcerans 0102 isolated in Japan.The genomic
sequence showed a striking similarity with that of Corynebacterium
pseudotuberculosis and, to a lesser extent, with that of C. diphtheriae. The 0102
genome contained three distinct prophages. One of these, PhiCULC0102-I, was a
tox-positive prophage containing genes in the same structural order as for
tox-positive C. diphtheriae prophages. However, the primary structures of the
individual genes involved in the phage machinery showed little homology between
the two counterparts. CONCLUSION: Taken together, these results suggest that the
tox-positive prophage in this strain of C. ulcerans has a distinct origin from
that of C. diphtheriae NCTC 13129.

<>

<1>Sektas, M., Kaczorowski, T., Podhajska, A.J.
<2>Purification and properties of the MboII, a class-IIS restriction endonuclease.
<3>Nucleic Acids Res.
<4>20
<5>433-438
<6>1992
<7>After five purification steps a homogeneous preparation of endonuclease MboII was obtained,
and several properties of the enzyme were determined. MboII is a monomer, with Mr under native
and denaturing conditions being 47 - 49,000 Da. Endonuclease MboII is a basic protein (pl 8.3)
which remains active when Mg2+ is replaced by Mn2+, Co2+, Ca2+, or Fe2+. MboII exhibits a star
activity in the presence of some of the following reagents or ions: DMSO, glycerol, ethanol
(and Co2+ or Mn2+ at pH 6). MboII does not bend DNA and is heat sensitive, losing activity
after 15 min at 50C.

<>

<1>Sektas, M., Kaczorowski, T., Podhajska, A.J.
<2>Interaction of the MboII restriction endonuclease with DNA.
<3>Gene
<4>157
<5>181-185
<6>1995
<7>The binding of the MboII restriction endonuclease (R.MboII; ENase) to DNA containing its
recognition site was investigated using a mobility shift assay.  R.MboII forms specific,
stable and immunodetectable complexes with its canonical target sequence.  The association
constant (a) of R.MboII was calculated to be 2.89 x 10/9M, and is about 10/4-fold higher than
the Ka value for non-specific binding.  Based on results obtained after sedimentation of the
R.MboII-DNA complex in a glycerol gradient and measurement of the retardation of the complexes
in polyacrylamide gels, we conclude that specific binding to the canonical sequence involves
a monomer of R.MboII.  DNase I footprinting has shown that the enzyme covers 16 nucleotides of
DNA on the 5'-GAAGA-3' strand.

<>

<1>Sela, D.A., Chapman, J., Adeuya, A., Kim, J.H., Chen, F., Whitehead, T.R., Lapidus, A., Rokhsar, D.S., Lebrilla, C.B., German, J.B., Price, N.P., Richardson, P.M., Mills, D.A.
<2>The genome sequence of Bifidobacterium longum subsp. infantis reveals adaptations for milk utilization within the infant microbiome.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>18964-18969
<6>2008
<7>Following birth, the breast-fed infant gastrointestinal tract is rapidly colonized by a
microbial consortium often dominated by bifidobacteria.
Accordingly, the complete genome sequence of Bifidobacterium longum subsp.
infantis ATCC15697 reflects a competitive nutrient-utilization strategy
targeting milk-borne molecules which lack a nutritive value to the
neonate. Several chromosomal loci reflect potential adaptation to the
infant host including a 43 kbp cluster encoding catabolic genes,
extracellular solute binding proteins and permeases predicted to be active
on milk oligosaccharides. An examination of in vivo metabolism has
detected the hallmarks of milk oligosaccharide utilization via the central
fermentative pathway using metabolomic and proteomic approaches. Finally,
conservation of gene clusters in multiple isolates corroborates the
genomic mechanism underlying milk utilization for this infant-associated
phylotype.

<>

<1>Selent, U., Pingoud, A.
<2>Altering the substrate specificity of the restriction endonuclease EcoRV.
<3>Biol. Chem. Hoppe Seyler
<4>376
<5>S152
<6>1995
<7>The type II restriction endonuclease EcoRV recognizes and cleaves the DNA sequence GAT/ATC. By
simultaneous exchange of several amino acids of the DNA recognition loop we want to generate
mutant enzymes that no longer cleave the cognate site but related ones, preferably those for
which at present no restriction enzyme is described or even commercially available. In our
approach, a large pool of EcoRV-mutants is generated by random mutagenesis and after an in
vivo selection, mutants with the desired specificity are selected.  At present, the work
concentrates on establishing a suitable screening system: a catalytically inactive EcoRV
mutant is used as a repressor for a toxic gene product, i.e. the restriction endonuclease
EcoRI.  Cells expressing EcoRI that are not protected by the corresponding methyltransferase
have a probability for survival that is 1:10^6.  An exchange of the operator sequence upstream
of the toxic gene, i.e. from GATATC to TATATA, should allow for a positive screening for
EcoRV-mutants that bind to the new sequence and thereby repress the expression of the toxic
gene.  In a subsequent step, the catalytic activity of the EcoRV mutant has to be restored by
site-directed mutagenesis.  Alternatively, we try to identify EcoRV mutants with altered
specificity directly.  For this purpose, an E. coli strain is used that induces the lac operon
upon DNA damage.  After UV irradiation or expression of a restriction enzyme in the absence of
the corresponding methylase, these cells grow to blue colonies on X-Gal agar plates. If the
cells are protected against cleavage at GATATC sites by the EcoRV methylase, the colonies are
white.  In contrast, EcoRV mutants with an altered specificity can cleave DNA in vivo even in
the presence of the methylase, and cells expressing such a variant will give blue colonies on
X-Gal agar.

<>

<1>Selent, U., Ruter, T., Kohler, E., Liedtke, M., Thielking, V., Alves, J., Oelgeschlager, T., Wolfes, H., Peters, F., Pingoud, A.
<2>A site-directed mutagenesis study to identify amino acid residues involved in the catalytic function of the restriction endonuclease EcoRV.
<3>Biochemistry
<4>31
<5>4808-4815
<6>1992
<7>We have used site-directed mutagenesis of the EcoRV restriction endonuclease to change amino
acid side chains that have been shown crystallographically to be in close proximity to the
scissile phosphodiester bond of the DNA substrate. DNA cleavage assays of the resulting mutant
proteins indicate that the larges effects on nucleolytic activity result from substitution of
Asp74, Asp90, and Lys92. We suggest on the basis of structural information, mutagenesis data,
and analogies with other nucleases that Asp74 and Asp90 might be involved in Mg2+ binding
and/or catalysis and that Lys92 probably stabilizes the pentacovalent phosphorus in the
transition state. These amino acids are part of a sequence motif, Pro-Asp...Asp/Glu-X-Lys,
which is also present in EcoRI. In both enzymes, it is located in a structurally similar
context near the scissile phosphodiester bond. A preliminary mutational analysis with EcoRI
indicates that this sequence motif is of similar functional importance for EcoRI and EcoRV. On
the basis of these results, a proposal is made for the mechanism of DNA cleavage by EcoRV and
EcoRI.

<>

<1>Seligman, L.M., Chisholm, K.M., Chevalier, B.S., Chadsey, M.S., Edwards, S.T., Savage, J.H., Veillet, A.L.
<2>Mutations altering the cleavage specificity of a homing endonuclease.
<3>Nucleic Acids Res.
<4>30
<5>3870-3879
<6>2002
<7>The homing endonuclease I-CreI recognizes and cleaves a particular 22 bp DNA sequence. The
crystal structure of I-CreI bound to homing site DNA has previously been determined, leading
to a number of predictions about specific protein-DNA contacts. We test these predictions by
analyzing a set of endonuclease mutants and a complementary set of homing site mutants. We
find evidence that all structurally predicted I-CreI/DNA contacts contribute to DNA
recognition and show that these contacts differ greatly in terms of their relative importance.
We also describe the isolation of a collection of altered specificity I-CreI derivatives. The
in vitro DNA-binding and cleavage properties of two such endonucleases demonstrate that our
genetic approach is effective in identifying homing endonucleases that recognize and cleave
novel target sequences.

<>

<1>Selker, E.U., Cambareri, E.B., Garrett, P.W., Haack, K.R., Jensen, B.C., Shabtach, E.
<2>DNA methylation and control of genome organization in Neurospora crassa.
<3>Gene
<4>74
<5>109-111
<6>1988
<7>Why are 5S rRNA genes dispersed in the genome of Neurospora, instead of tandemly arranged as
in most organisms? A likely answer to this question came from studying an exceptional pair of
adjacent Neurospora 5S genes, or pseudogenes, designated zeta and eta. The zeta-eta region
consists of a diverged direct tandem duplication of a 0.8-kb segment including a 5S rRNA gene.
Sequence comparisons of the zeta and eta 5S rRNA regions with each other and with other 5S
regions suggested that the approx. 15% divergence between the zeta and eta 'duplicate'
segments resulted exclusively from CG to TA mutations. Unlike other 5S rRNA regions examined,
the zeta-eta region is heavily methylated. Thus it seemed likely that these transition
mutations arose by deamination of mC residues. In the course of exploring the basis for the
extraordinarily heavy methylation in the zeta-eta region, we discovered a novel genetic
process that accounts for the methylation and provides a probable explanation for why 5S rRNA
genes are generally dispersed in Neurospora.

<>

<1>Selker, E.U., Freitag, M., Kothe, G.O., Margolin, B.S., Rountree, M.R., Allis, C.D., Tamaru, H.
<2>Induction and maintenance of nonsymmetrical DNA methylation in Neurospora.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>16485-16490
<6>2002
<7>One can imagine a variety of mechanisms that should result in self-perpetuating biological
states. It is generally assumed that cytosine methylation is propagated in eukaryotes by
enzymes that specifically methylate hemimethylated symmetrical sites (e.g., (5')CpG/GpC(5')
or (5')CpNpG/GpNpC(5')). Although there is wide support for this model, we and others have
found examples of methylation that must be propagated by a different mechanism. Most
methylated regions of the Neurospora genome that have been examined are products of
repeat-induced point mutation, a premeiotic genome defense system that litters duplicated
sequences with C.G to T.A mutations and typically leaves them methylated at remaining
cytosines. In general, such relics of repeat-induced point mutation are capable of triggering
methylation de novo. Nevertheless, some reflect a mechanism that can propagate heterogeneous
methylation at nonsymmetrical sites. We propose that de novo and maintenance methylation are
manifestations of a single mechanism in Neurospora, catalyzed by the DIM-2 DNA
methyltransferase. The action of DIM-2 is controlled by the DIM-5 histone H3 Lys-9
methyltransferase, which in turn is influenced by other modifications of histone H3. DNA
methylation indirectly recruits histone deacetylases, providing the framework of a
self-reinforcing system that could result in propagation of DNA methylation and the associated
silenced chromatin state.

<>

<1>Selker, E.U., Fritz, D.Y., Singer, M.J.
<2>Dense nonsymmetrical DNA methylation resulting from repeat-induced point mutation in Neurospora.
<3>Science
<4>262
<5>1724-1728
<6>1993
<7>Cytosine methylation has been implicated in epigenetic control of gene expression in animals,
plants and fungi. It has been assumed that all methylation in eukaryotes is at symmetrical
sequences such as CpG/GpC, because this can explain perpetuation of methylation states. Here
the bisulfite genomic sequencing method was used to examine methylation in DNA from a
Neurospora gene exposed to repeat-induced point mutation. 5-Methylcytosine was not limited to
symmetrical sites and individual molecules showed different patterns and amounts of
modification. The methylation extended beyond the mutated region and even beyond the edge of
the duplicated segment.

<>

<1>Selker, E.U., Tountas, N.A., Cross, S.H., Margolin, B.S., Murphy, J.G., Bird, A.P., Freitag, M.
<2>The methylated component of the Neurospora crassa genome.
<3>Nature
<4>422
<5>893-897
<6>2003
<7>Cytosine methylation is common, but not ubiquitous, in eukaryotes. Mammals and the fungus
Neurospora crassa have about 2-3% of cytosines methylated.
In mammals, methylation is almost exclusively in the under-represented CpG
dinucleotides, and most CpGs are methylated whereas in Neurospora,
methylation is not preferentially in CpG dinucleotides and the bulk of the
genome is unmethylated. DNA methylation is essential in mammals but is
dispensable in Neurospora, making this simple eukaryote a favoured
organism in which to study methylation. Recent studies indicate that DNA
methylation in Neurospora depends on one DNA methyltransferase, DIM-2
(ref. 6), directed by a histone H3 methyltransferase, DIM-5 (ref. 7), but
little is known about its cellular and evolutionary functions. As only
four methylated sequences have been reported previously in N. crassa, we
used methyl-binding-domain agarose chromatography to isolate the
methylated component of the genome. DNA sequence analysis shows that the
methylated component of the genome consists almost exclusively of relics
of transposons that were subject to repeat-induced point mutation--a
genome defence system that mutates duplicated sequences.

<>

<1>Sellem, C.H., Belcour, L.
<2>Intron open reading frames as mobile elements and evolution of a group I intron.
<3>Mol. Biol. Evol.
<4>14
<5>518-526
<6>1997
<7>Group I introns are proposed to have become mobile following the acquisition of open reading
frames (ORFs) that encode highly specific DNA endonucleases. This proposal implies that intron
ORFs could behave as autonomously mobile entities. This was supported by abundant
circumstantial evidence but no experiment of ORF transfer from an ORF-containing intron to its
ORF-less counterpart has been described. In this paper we present such experiments, which
demonstrate the efficient mobility of the mitochondrial nad1-i4-orf1 between two Podospora
strains. The homing of this mobile ORF was accompanied by a bidirectional co-conversion that
did not systematically involve the whole intron sequence. Orf1 acquisition would be the most
recent step in the evolution of the nad1-i4 intron, which has resulted in many strains of
Podospora having an intron with two ORFs (biorfic) and four splicing pathways. We show that
two of the splicing events that operate in this biorfic intron, as evidenced by PCR
experiments, are generated by a 5'-alternative splice site, which is most probably a remnant
of the monoorfic ancestral form of the intron. We propose a sequential evolution model that is
consistent with the four organizations of the corresponding nad1 locus that we found among
various species of the Pyrenomycete family; these organizations consist of no intron, an
intron alone, a monoorfic intron, and a biorfic intron.

<>

<1>Sellmann, E., Knoblich, I.-M., Kaluza, K., Frey, B., Schmitz, G.G., Westermann, P.
<2>Bsp423I, a novel isoschizomer of BbvI from Bacillus recognizing 5'-GCAGC-3'.
<3>Nucleic Acids Res.
<4>20
<5>2377
<6>1992
<7>We have isolated Bsp423I, a novel class-II restriction endonuclease from Bacillus species
recognizing the palindromic sequence 5'-GCAGC-3' generating 5'-protruding tetranucleotides
within the sequence complementary to 5'-GCAGC(N)8-3'. With respect to its isoschizomer BbvI
it can be isolated in higher purity and stability. From the mapping and sequencing data the
specificity of Bsp423I is concluded as: 5'-GCAGC(N)8/-3' 3'-CGTCG(N)12/-5'.

<>

<1>Selvakumar, K.S., Shanmugasundaram, S.
<2>Studies on DNA modification in Oscillatoria sp. MKU 178.
<3>Curr. Sci.
<4>64
<5>49-50
<6>1993
<7>Cyanobacterial DNA are generally believed to be resistant to restriction by endonucleases due
to extensive modification of the DNA. We report here the absence of a dam-mediated
modification in a cyanobacterium, Oscillatoria sp. MKU 178.

<>

<1>Selvaraj, S., Kono, H., Sarai, A.
<2>Specificity of Protein-DNA Recognition Revealed by Structure-based Potentials: Symmetric/Asymmetric and Cognate/Non-cognate Binding.
<3>J. Mol. Biol.
<4>322
<5>907
<6>2002
<7>Asymmetric binding of protein homodimers to DNA, which has been observed in a number of
protein-DNA complexes, leads to subtle structural differences between the two subunits. Such
structural differences are frequently observed when the subunits form cognate and non-cognate
protein-DNA complexes, respectively. Analysis of these structural effects on binding
specificity should provide insight into the mechanism of protein-DNA recognition. We
previously derived empirical potential functions for specific nucleotide base-amino acid
interactions from statistical analyses of the structures of many protein-DNA complexes and
used a combinatorial threading procedure to evaluate the fitness of the DNA sequences
involved. We then introduced Z-scores to measure the specificity with which proteins bind to
DNA within complexes, as compared to random DNA sequences. Here, we examined in detail the
structural effects of asymmetric and cognate/non-cognate binding on specificity. Marked
differences in the specificity of DNA binding were observed for the two subunits of lambda
repressor, the glucocorticoid receptor, and for transcription factors containing a Zn(2)Cys(6)
binuclear cluster domain, which are known to bind asymmetrically to DNA. Moreover, the
differences in the specificity with which BamHI and EcoRV endonucleases bind to their cognate
and non-cognate DNA sequences were clearly detected using this approach; indeed, analysis of
EcoRV binding enabled us to show the cooperative effect of sequence and structure on binding
specificity. The present results demonstrate the utility of this approach when examining the
structure-specificity relationship in protein-DNA recognition, as subtle structural
differences in symmetric/asymmetric and cognate/non-cognate binding were clearly shown to
cause marked differences in specificity. This method can also be used as a tool for checking
new structures of protein-DNA complexes for their specificity.

<>

<1>Semenova, E., Minakhin, L., Bogdanova, E., Nagornykh, M., Vasilov, A., Heyduk, T., Solonin, A., Zakharova, M., Severinov, K.
<2>Transcription regulation of the EcoRV restriction-modification system.
<3>Nucleic Acids Res.
<4>33
<5>6942-6951
<6>2005
<7>When a plasmid containing restriction-modification (R-M) genes enters a naive host, unmodified
host DNA can be destroyed by restriction endonuclease. Therefore, expression of R-M genes must
be regulated to ensure that enough methyltransferase is produced and that host DNA is
methylated before the endonuclease synthesis begins. In several R-M systems, specialized
Control (C) proteins coordinate expression of the R and the M genes. C proteins bind to DNA
sequences called C-boxes and activate expression of their cognate R genes and inhibit the M
gene expression, however the mechanisms remain undefined. Here, we studied the regulation of
gene expression in the C protein-dependent EcoRV system. We map the divergent EcoRV M and R
gene promoters and we define the site of C protein-binding that is sufficient for activation
of the EcoRV R transcription.

<>

<1>Semmler, T., Eichhorn, I., Bethe, A., Bauerfeind, R., Pickard, D., Kingsley, R.A., Dougan, G., Wieler, L.H.
<2>Genome Sequence of Porcine Escherichia coli Strain IMT8073, an Atypical Enteropathogenic E. coli Strain Isolated from a Piglet with Diarrhea.
<3>Genome Announcements
<4>1
<5>e00573-13
<6>2013
<7>Escherichia coli is a highly diverse bacterial species, with atypical enteropathogenic E. coli
(aEPEC) causing intestinal disease in both human and
animal hosts. Here, we report the first complete genome sequence of an aEPEC
strain of sequence type ST794 and serotype Ont:H7, isolated from a diseased
piglet.

<>

<1>Sen, A. et al.
<2>Draft Genome Sequence of Frankia sp. Strain QA3, a Nitrogen-Fixing Actinobacterium Isolated from the Root Nodule of Alnus nitida.
<3>Genome Announcements
<4>1
<5>e0010313
<6>2013
<7>Members of the actinomycete genus Frankia form a nitrogen-fixing symbiosis with 8 different
families of actinorhizal plants. We report a high-quality draft genome
sequence for Frankia sp. strain QA3, a nitrogen-fixing actinobacterium isolated
from root nodules of Alnus nitida.

<>

<1>Sen, D., Brown, C.J., Top, E.M., Sullivan, J.
<2>Inferring the Evolutionary History of IncP-1 Plasmids Despite Incongruence among Backbone Gene Trees.
<3>Mol. Biol. Evol.
<4>30
<5>154-166
<6>2012
<7>Plasmids of the incompatibility group IncP-1 can transfer and replicate in many
genera of the Proteobacteria. They are composed of backbone genes that encode a
variety of essential functions as well as accessory genes that have implications
for human health and environmental remediation. While it is well understood that
the accessory genes are transferred horizontally between plasmids, recent studies
have also provided examples of recombination in the backbone genes of IncP-1
plasmids. As a consequence, phylogeny estimation based on backbone genes is
expected to produce conflicting gene tree topologies. The main goal of this study
was therefore to infer the evolutionary history of IncP-1 plasmids in the
presence of both vertical and horizontal gene transfer. This was achieved by
quantifying the incongruence among gene trees and attributing it to known causes
such as, a) phylogenetic uncertainty, b) coalescent stochasticity, and c)
horizontal inheritance. Topologies of gene trees exhibited more incongruence than
could be attributed to phylogenetic uncertainty alone. Species-tree estimation
using a Bayesian framework that takes coalescent stochasticity into account was
well supported, but it differed slightly from the maximum likelihood tree
estimated by concatenation of backbone genes. After removal of the gene that
demonstrated a signal of intergroup recombination, the concatenated tree was
congruent with the species-tree estimate, which itself was robust to
inclusion/exclusion of the recombinant gene. Thus, in spite of horizontal gene
exchange both within and among IncP-1 subgroups, the backbone genome of these
IncP-1 plasmids retains a detectable vertical evolutionary history.

<>

<1>Sen, D., Van der Auwera, G.A., Rogers, L.M., Thomas, C.M., Brown, C.J., Top, E.M.
<2>Broad-Host-Range Plasmids from Agricultural Soils Have IncP-1 Backbones with Diverse Accessory Genes.
<3>Appl. Environ. Microbiol.
<4>77
<5>7975-7983
<6>2011
<7>Broad-host-range plasmids are known to spread genes between distinct phylogenetic
groups of bacteria. These genes often code for resistances to antibiotics and
heavy metals or degradation of pollutants. Although some broad-host-range
plasmids have been extensively studied, their evolutionary history and genetic
diversity remain largely unknown. The goal of this study was to analyze and
compare the genomes of 12 broad-host-range plasmids that were previously isolated
from Norwegian soils by exogenous plasmid isolation and that encode mercury
resistance. Complete nucleotide sequencing followed by phylogenetic analyses
based on the relaxase gene traI showed that all the plasmids belong to one of two
subgroups (beta and epsilon) of the well-studied incompatibility group IncP-1. A
diverse array of accessory genes was found to be involved in resistance to
antimicrobials (streptomycin, spectinomycin, and sulfonamides), degradation of
herbicides (2,4-dichlorophenoxyacetic acid and 2,4-dichlorophenoxypropionic
acid), and a putative new catabolic pathway. Intramolecular transposition of
insertion sequences followed by deletion was found to contribute to the diversity
of some of these plasmids. The previous observation that the insertion sites of a
Tn501-related element are identical in four IncP-1beta plasmids (pJP4, pB10,
R906, and R772) was further extended to three more IncP-1beta plasmids (pAKD15,
pAKD18, and pAKD29). We proposed a hypothesis for the evolution of these
Tn501-bearing IncP-1beta plasmids that predicts recent diversification followed
by worldwide spread. Our study increases the available collection of complete
IncP-1 plasmid genome sequences by 50% and will aid future studies to enhance our
understanding of the evolution and function of this important plasmid family.

<>

<1>Sen, S., Nilsson, L.
<2>Free energy calculations and molecular dynamics simulations of wild-type and variants of the DNA-EcoRI complex.
<3>Biophys. J.
<4>77
<5>1801-1810
<6>1999
<7>Molecular dynamics simulations and free energy calculations of the wild-type EcoRI-DNA complex
and several variants have been performed in aqueous solvent. In general, the theoretical
estimations of the free energy differences (DeltaDeltaA) qualitatively agree well with the
corresponding experimental data. The modifications which were experimentally found unfavorable
compared to the wild-type complex were also found to be so in theoretical estimates. The
mutant where the amino group of the base Ade(6) was replaced by a hydrogen atom eliminating
one H-bond between the DNA and the protein, was experimentally found to be more stable than
the wild-type complex. It was speculated that the modification also caused a structural
relaxation in the DNA making DeltaDeltaA favorable. Our theoretical estimate yields a positive
DeltaDeltaA in this case, but the difference is small, and no significant local structural
relaxation was observed. The major H-bonds between the DNA and the protein in the wild-type
complex are found to be maintained in the different mutants although the specific and
non-specific interaction energies between the interacting the DNA bases and the protein
residues are different in different mutants. The interaction pattern of the other nearby
nucleotides are significantly influenced by each modification. Thus, the alteration of the
non-specific interactions may also play an indirect role in determining the specificity of the
complex. The interaction of the Gua(4) of the DNA with the protein is found to be most
sensitive to any alteration in the recognition site. Because Gua(4) is the nucleotide closest
to the scissile bond, this extra sensitivity seems to play an important role in altering the
functional activity of the complex.

<>

<1>Sen, S., Nilsson, L.
<2>Structure, interaction, dynamics and solvent effects on the DNA-EcoRI complex in aqueous solution from molecular dynamics simulation.
<3>Biophys. J.
<4>77
<5>1782-1800
<6>1999
<7>A 0.7-ns molecular dynamics simulation of the DNA-EcoRI complex in a 7.0-A solvent shell
indicated a stable behavior of the system. No significant evaporation or smearing of the
solvent's outer boundary occurred. The structure and the intermolecular interactions were
found to be well maintained during the simulation. The interaction pattern in the simulation
was found to be very similar to that in the crystal structure. Most of the specific
interactions between the DNA and the protein were found to be enhanced in the simulation
compared to that in the crystal structure as a result of improved interaction geometry. The
nonspecific interactions were found to be stronger than the specific ones. The specific
interactions between the N7 atoms of Gua(4) or Ade(5) or Ade(6) and the protein were found to
be present over almost the entire time of the simulation, whereas hydrogen bonds involving the
amino groups of the Ade(5) and Ade(6) with the protein were found to be relatively weaker,
with lower probability and shorter lifetime. The time evolution of the root mean square
deviations of the DNA and the protein were highly correlated even at the later part of the
simulation, showing the tight binding between them. Several long-lived water bridges were
found between the DNA backbone atoms and the protein and also between the two protein
monomers, which increased the overall stability of the complex. The two protein monomers were
found to interact strongly with each other. The energy of the DNA kink deformation was
estimated as approximately 31 kcal/mol.

<>

<1>Senejani, A.G., Gogarten, J.P.
<2>Structural stability and endonuclease activity of a PI-SceI GFP-fusion protein.
<3>Int. J. Biol. Sci.
<4>3
<5>205-211
<6>2007
<7>Homing endonucleases are site-specific and rare cutting endonucleases often encoded by intron
or intein containing genes. They lead to the
rapid spread of the genetic element that hosts them by a process termed
'homing'; and ultimately the allele containing the element will be
fixed in the population.
PI-SceI, an endonuclease encoded as a protein insert or intein
within the yeast V-ATPase catalytic subunit encoding gene (vma1), is
among the best characterized homing endonucleases. The structures of
the Sce VMA1 intein and of the intein bound to its target site are
known. Extensive biochemical studies performed on the PI-SceI enzyme
provide information useful to recognize critical amino acids involved
in self-splicing and endonuclease functions of the protein. Here we
describe an insertion of the Green Fluorescence Protein (GFP) into a
loop which is located between the endonuclease and splicing domains of
the Sce VMA1 intein. The GFP is functional and the additional GFP
domain does not prevent intein excision and endonuclease activity.
However, the endonuclease activity of the newly engineered protein was
different from the wild-type protein in that it required the presence
of Mn2+ and not Mg2+ metal cations for activity.

<>

<1>Senesac, J.H., Allen, J.R.
<2>Oligonucleotide activation of the type IIe restriction enzyme NaeI for digestion of refractory sites.
<3>Biotechniques
<4>19
<5>990-993
<6>1995
<7>Certain restriction endonucleases previously shown to exhibit DNA site preferences have a
two-site DNA cleavage mechanism.  These type IIe restriction endonucleases include NaeI, NarI,
EcoRII, HpaII and SacII.  Because of this two-site mechanism, it is often difficult or
impossible to achieve complete digestion of DNA subsrtrate.  Inasmuch as these enzymes are
commonly used in molecular biology, a method for enzyme activation to provide complete DNA
digestion is useful.  We have commercialized such a method for NaeI using a double-stranded
oligonucleotide containing a modified NaeI recognition sequence.  Cleavage of resistant sites
requires the presence of a DNA sequence that is more cleavable to bind the activator site.
The regions flanking the recognition site on our NaeI oligonucleotide cause it to serve as
this more cleavable sequence.  This activates the enzyme to cleave the resistant sequence in
the catalytic site, while the oligonucleotide modification does not allow the activator to be
depleted during the reaction.  Turbo NaeI provides for rapid digestion of sites previously
found difficult or impossible to completely cleave and does not interfere with subsequent
molecular biology techniques that might be performed downstream on the substrate DNA, such as
ligation, end-labeling or nick translation.

<>

<1>Senesac, J.H., Romanin, J.K.
<2>Application of oligonucleotide activation to restriction endonuclease NarI.
<3>Biotechniques
<4>22
<5>1166-1168
<6>1997
<7>Restriction endonuclease NarI cleaves DNA using a two-site mechanism, placing it in the type
IIe class of restriction endonucleases.  Although these enzymes have very useful recognition
sequences, the two-site mechanism limits the practical application.  Site preferences often
cause incomplete substrate digestion.  Oligonucleotide activation of NarI eliminates
incomplete digestions, making it possible to use NarI restriction sites in many common
molecular biology techniques.  A modified oligonucleotide was chosen for optimal activation of
restriction endonuclease NarI.  This oligonucleotide was demonstrated to allow complete
digestion in many commonly used substrates.

<>

<1>Sentausa, E., Abdad, M.Y., Robert, C., Stenos, J., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Rickettsia gravesii, Isolated from Western Australian Ticks.
<3>Genome Announcements
<4>1
<5>e00975-13
<6>2013
<7>Rickettsia gravesii is a new Rickettsia species closely related to the human pathogen
Rickettsia massiliae. Here, we describe the genome sequence of R.
gravesii strain BWI-1, isolated from Amblyomma triguttatum triguttatum ticks
collected from humans on Barrow Island, Western Australia.

<>

<1>Sentausa, E., El Karkouri, K., Michelle, C., Caputo, A., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Rickettsia tamurae, a Recently Detected Human Pathogen in Japan.
<3>Genome Announcements
<4>2
<5>e00838-14
<6>2014
<7>Rickettsia tamurae is a member of the spotted fever group rickettsiae, which was  reported in
2011 to cause human infections in Japan. We report the draft genome
sequence of R. tamurae strain AT-1(T), isolated from Amblyomma testudinarium
ticks.

<>

<1>Sentausa, E., El Karkouri, K., Michelle, C., Raoult, D., Fournier, P.E.
<2>Draft Genome Sequence of Rickettsia aeschlimannii, Associated with Hyalomma marginatum Ticks.
<3>Genome Announcements
<4>2
<5>e00666-14
<6>2014
<7>Rickettsia aeschlimannii is a tick-associated human pathogen. We report here the  draft genome
of R. aeschlimannii strain MC16, isolated from Hyalomma marginatum
marginatum ticks collected in Morocco.

<>

<1>Sentausa, E., El Karkouri, K., Nguyen, T.T., Caputo, A., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Rickettsia hoogstraalii, a Geographically Widely Distributed Tick-Associated Bacterium.
<3>Genome Announcements
<4>2
<5>e01171-14
<6>2014
<7>Rickettsia hoogstraalii is a tick-associated member of the spotted fever group rickettsiae
that is geographically widely distributed. We report here the draft
genome of R. hoogstraalii strain Croatica(T) (=DSM 22243 = UTMB 00003), which was
isolated from Haemaphysalis sulcata ticks collected in Croatia.

<>

<1>Sentausa, E., El Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Genome Sequence of 'Rickettsia sibirica subsp. mongolitimonae'.
<3>J. Bacteriol.
<4>194
<5>2389-2390
<6>2012
<7>'Rickettsia sibirica subsp. mongolitimonae' is the agent of lymphangitis-associated
rickettsiosis, an emerging human disease that has been
diagnosed in Europe and Africa. The present study reports the draft genome of
Rickettsia sibirica subsp. mongolitimonae strain HA-91.

<>

<1>Sentausa, E., El Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Sequence and Annotation of Rickettsia sibirica sibirica Genome.
<3>J. Bacteriol.
<4>194
<5>2377
<6>2012
<7>Rickettsia sibirica sibirica is the causative agent of Siberian or North Asian tick typhus, a
tick-borne rickettsiosis known to exist in Siberia and eastern
China. Here we present the draft genome of Rickettsia sibirica sibirica strain
BJ-90 isolated from Dermacentor sinicus ticks collected in Beijing, China.

<>

<1>Sentausa, E., El Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Rickettsia conorii subsp. indica, the Agent of Indian Tick Typhus.
<3>J. Bacteriol.
<4>194
<5>3288-3289
<6>2012
<7>Rickettsia conorii subsp. indica is the agent of Indian tick typhus. The present  study
reports the draft genome of Rickettsia conorii subsp. indica strain ITTR
(ATCC VR-597).

<>

<1>Sentausa, E., El Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Rickettsia conorii subsp. caspia, the Agent of Astrakhan Fever.
<3>J. Bacteriol.
<4>194
<5>4763-4764
<6>2012
<7>Rickettsia conorii subsp. caspia is the agent of Astrakhan fever, a spotted fever group
rickettsiosis endemic to Astrakhan, Russia. The present study reports the
draft genome of Rickettsia conorii subsp. caspia strain A-167.

<>

<1>Sentausa, E., El Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Genome Sequence of Rickettsia conorii subsp. israelensis, the Agent of Israeli Spotted Fever.
<3>J. Bacteriol.
<4>194
<5>5130-5131
<6>2012
<7>Rickettsia conorii subsp. israelensis is the agent of Israeli spotted fever. The  present
study reports the draft genome of Rickettsia conorii subsp. israelensis
strain ISTT CDC1, isolated from a Rhipicephalus sanguineus tick collected in
Israel.

<>

<1>Sentis, C., Santos, J., Fernandez-Piqueras, J.
<2>In situ methylation of insect chromosomes with methylase HpaII.
<3>Chromosoma
<4>98
<5>105-108
<6>1989
<7>A method of in situ DNA methylation with the prokaryotic methylase HpaII has
been developed on fixed mitotic and meiotic chromosomes of the insect species
Baetica ustulata.  Incorporation of methyl groups into the chromosomal DNA is
revealed by autoradiography using a laelled substrate and by its ability to
prevent endonuclease digestion.  The method allows direct visualization of
clusters of methylatable CCGG sites.  The distribution of these clusters in the
chromosome complement of Baetica shows two separate domains of heterochromatic
DNA which differ in their methylation patterns.  Each is distributed at
equivalent locations in both homologus and nonhomologus chromosomes.  The
existence of two compartments, one methylated and the other unmethylated, in
the heterochromatic DNA could be interpreted as a remnant of the ancestral
echinoderm-like pattern of methylation.

<>

<1>Seo, J., Park, H., Kang, H., Chong, H.
<2>GENOME SEQUENCE OF ZYMONAS MOBILIS ZM4 AND NOVEL GENE INVOLVED INPRODUCTION OF ETHANOL.
<3>Korean Patent Office
<4>KR 1020040099447 A
<5>
<6>2004
<7>
<>

<1>Seo, J.H., Cho, Y.D.
<2>Studies on the role of cysteinyl residue in EcoRI methylase.
<3>Korean Biochem. J.
<4>24
<5>193-199
<6>1991
<7>The chemical modification of cysteinyl residue of EcoRI methylase by NEM and
DACM resulted in the loss of the enzyme activity following pseudo first order
kinetics.  Second order rate constants (K) for the inactivation of EcoRI
methylase by NEM and DACM were 685 M-1.min-1 and  99960 M-1.min-1,
respectively.  The reaction orders with respect to NEM and DACM were determined
from plot of log (100/t1/2) against log (NEM) and log (DACM).  When the data
are plotted as indicated above, slopes of 0.95 and 1.25 are obtained,
suggesting that inactivation is the result of the reaction of one cysteinyl
residue per active site of EcoRI methylase.  The inactivation of EcoRI
methylase by DTNB was reversed upon addition of excess 2-mercaptoethanol.  When
the enzyme solution was preincubated with the substrates, the inactivation of
the enzyme by NEM and DACM was protected.  Furtheremore, loss of SAM binding
capacity of EcoRI methylase was detected by nitrocellulose filter binding
assay.  Based on the data, we suggest that cysteinyl residue of EcoRI methylase
plays a very important role as a SAM binding residue.

<>

<1>Seo, J.H., Cho, Y.D.
<2>Studies on the role of arginyl residue in EcoRI methylase.
<3>Korean Biochem. J.
<4>25
<5>54-59
<6>1992
<7>Treatment of EcoRI methylase with phenylglyoxal resulted in time and concentration dependent
enzyme inactivation. The reaction followed pseudo first-order kinetics until 90 to 95% of the
enzyme had been inactivated, and prolonged incubation with phenylglyoxal resulted in complete
inactivation. Second order rate constant (K) for the inactivation of EcoRI methylase by
phenylglyoxal was 666 M-1 min-1. The slope determined from the plot of log (100/t1/2) against
log (phenylglyoxal) indicated that the reaction of 1 molecule of phenylglyoxal reagent in
necessary for inactivation of each active site in EcoRI methylase. Preincubation with pUC19 or
S-adenosylmethionine as substrates decreased significantly the rate of inactivation by
phenylglyoxal. By nitrocellulose binding assay, phenylglyoxal was observed to reduce DNA
binding capacity of the enzyme more rapidly than S-adenosylmethionine. Furthermore, DNA
binding capacity of the enzyme was preserved when the enzyme was preincubated with pUC19 DNA.
The cumulative results suggest that arginyl residue plays a very important role as a DNA
binding residue in EcoRI methylase.

<>

<1>Seo, Y.S., Lim, J.Y., Choi, B.S., Kim, H., Goo, E., Lee, B., Lim, J.S., Choi, I.Y., Moon, J.S., Kim, J., Hwang, I.
<2>Complete Genome Sequence of Burkholderia gladioli BSR3.
<3>J. Bacteriol.
<4>193
<5>3149
<6>2011
<7>We report the complete genome sequence of Burkholderia gladioli BSR3 isolated from a diseased
rice sheath in Korea.

<>

<1>Seoane, A., Sangari, F.J., Lobo, J.M.
<2>Complete nucleotide sequence of the fosfomycin resistance transposon Tn2921.
<3>Int. J. Antimicrob. Agents
<4>35
<5>413-414
<6>2010
<7>Fosfomycin is a cell wall-active antibiotic introduced into clinical practice around 1970.
Despite its broad spectrum of activity and good pharmacological properties, its use has been
somewhat hampered by the high number of spontaneous resistant mutants isolated following
antibiotic challenge in many bacterial pathogens, most of them carrying chromosomal mutations
impairing drug transport.

<>

<1>Seraphin, B., Faye, G., Hatat, D., Jacq, C.
<2>The yeast mitochondrial intron aI5a: associated endonuclease activity and in vivo mobility.
<3>Gene
<4>113
<5>1-8
<6>1992
<7>By analyzing crosses between yeast strains carrying different combinations of mitochondrial
(mt) introns, we have shown that the aI5a intron is mobile in vivo. Furthermore, we have
observed that the mobility of intron aI5a is affected by both the nuclear and mt genotypes. We
have also detected a restriction endonuclease (ENase) activity that cleaves intronless mt
genomes close to the aI5a intron insertion site and thus might be involved in intron mobility.
Furthermore, similar to other ENases encoded by mobile mt introns of yeast, the ENase
generates a cut with a four-base 3'-OH overhang. Thus, intron aI5a represents a
characteristic member of the family of mobile group-I introns.

<>

<1>Seraphin, B., Simon, M., Faye, G.
<2>A mitochondrial reading frame which may code for a maturase-like protein in Saccharomyces cerevisiae.
<3>Nucleic Acids Res.
<4>13
<5>3005-3014
<6>1985
<7>In S. cerevisiae, the large oxi3/oli2 mitochondrial transcript contains
the products of the oxi3, aap1 and oli2 genes and an unassigned reading
frame, RF3. In the work presented here, we have completed the nucleotide
sequence of RF3. We have shown that RF3 is composed of four fairly large
ORFs which overlap within GC rich sequences. Furthermore, a shift of +1
base was found between each pair of consecutive reading frames. We discuss
how these frameshifts could be removed to produce a 500 aminoacid long
protein containing the two well conserved P1 and P2 oligopeptide sequences
featuring several mitochondrial intron reading frames, suggesting,
thereby, a RNA-maturase-like activity for the putative RF3 protein. In
addition, we suggest that the insertion of GC clusters in a gene could
provide a novel way of regulating its expression.

<>

<1>Serepa, M.H., Gray, V.M.
<2>Draft Whole-Genome Sequence of Serratia marcescens Strain MCB, Associated with Oscheius sp. MCB (Nematoda: Rhabditidae) Isolated from South Africa.
<3>Genome Announcements
<4>2
<5>e00911-14
<6>2014
<7>Here we report on the draft genome sequence of Serratia marcescens strain MCB associated with
Oscheius sp. MCB (Nematoda: Rhabditidae) isolated from South
African soil. S. marcescens strain MCB has 5,304,212-bp genome size with 4,877
genes and a G+C content of 59.1%.

<>

<1>Serfiotis-Mitsa, D., Herbert, A.P., Roberts, G.A., Soares, D.C., White, J.H., Blakely, G.W., Uhrin, D., Dryden, D.T.
<2>The structure of the KlcA and ArdB proteins reveals a novel fold and antirestriction activity against Type I DNA restriction systems in vivo  but not in vitro.
<3>Nucleic Acids Res.
<4>38
<5>1723-1737
<6>2010
<7>Plasmids, conjugative transposons and phage frequently encode anti-restriction proteins to
enhance their chances of entering a new
bacterial host that is highly likely to contain a Type I DNA restriction
and modification (RM) system. The RM system usually destroys the invading
DNA. Some of the anti-restriction proteins are DNA mimics and bind to the
RM enzyme to prevent it binding to DNA. In this article, we characterize
ArdB anti-restriction proteins and their close homologues, the KlcA
proteins from a range of mobile genetic elements; including an ArdB
encoded on a pathogenicity island from uropathogenic Escherichia coli and
a KlcA from an IncP-1b plasmid, pBP136 isolated from Bordetella pertussis.
We show that all the ArdB and KlcA act as anti-restriction proteins and
inhibit the four main families of Type I RM systems in vivo, but fail to
block the restriction endonuclease activity of the archetypal Type I RM
enzyme, EcoKI, in vitro indicating that the action of ArdB is indirect and
very different from that of the DNA mimics. We also present the structure
determined by NMR spectroscopy of the pBP136 KlcA protein. The structure
shows a novel protein fold and it is clearly not a DNA structural mimic.

<>

<1>Serfiotis-Mitsa, D., Roberts, G.A., Cooper, L.P., White, J.H., Nutley, M., Cooper, A., Blakely, G.W., Dryden, D.T.
<2>The Orf18 Gene Product from Conjugative Transposon Tn916 Is an ArdA Antirestriction Protein that Inhibits Type I DNA Restriction-Modification  Systems.
<3>J. Mol. Biol.
<4>383
<5>970-981
<6>2008
<7>Gene orf18, which is situated within the intercellular transposition region of the conjugative
transposon Tn916 from the bacterial pathogen Enterococcus faecalis, encodes a putative ArdA
(alleviation of restriction of DNA A) protein. Conjugative transposons are generally resistant
to DNA restriction upon transfer to a new host. ArdA from Tn916 may be responsible for the
apparent immunity of the transposon to DNA restriction and modification (R/M) systems and for
ensuring that the transposon has a broad host range. The orf18 gene was engineered for
overexpression in Escherichia coli, and the recombinant ArdA protein was purified to
homogeneity. The protein appears to exist as a dimer at nanomolar concentrations but can form
larger assemblies at micromolar concentrations. R/M assays revealed that ArdA can efficiently
inhibit R/M by all four major classes of Type I R/M enzymes both in vivo and in vitro. These
R/M systems are present in over 50% of sequenced prokaryotic genomes. Our results suggest that
ArdA can overcome the restriction barrier following conjugation and so helps increase the
spread of antibiotic resistance genes by horizontal gene transfer.

<>

<1>Sergeev, V.N., Chalov, S.E., Drutsa, V.L., Gromova, E.S.
<2>A study of the Asp110-Glu112 region of EcoRII restriction endonuclease by site-directed mutagenesis.
<3>Biochemistry
<4>65
<5>1006-1010
<6>2000
<7>Site-directed mutagenesis of the ecoRII gene has been used to search for the active site of
the EcoRII restriction endonuclease. Plasmids
with point mutations in ecoRII gene resulting in substitutions of amino
acid residues in the Asp110-Glu112 region of the EcoRII endonuclease
(Asp110 --> Lys, Asn, Thr, Val, or Ile; Pro111 --> Arg, His, Ala, or
Leu; Glu112 --> Lys, Gin, or Asp) have been constructed. When expressed
in E. coli, ail these plasmids displayed EcoRII endonuclease activity,
We also constructed a plasmid containing a mutant ecoRII gene with
deletion of the sequence coding the Gln109-Pro111 region of the
protein. This mutant protein had no EcoRII endonuclease activity. The
data suggest that Asp110, Pro111, and Glu112 residues do not
participate in the formation of the EcoRII active site. However, this
region seems to be relevant for the formation of the tertiary structure
of the EcoRII endonuclease.

<>

<1>Serov, G.D., Repin, V.E., Puchkova, L.E., Kolesnikov, V.A.
<2>The strain of Bacillus sphaericus 45 is a producer of the restriction endonuclease Bsh45I.
<3>Russian Patent Office
<4>RU 2007453 C
<5>
<6>1994
<7>Isolation and characterization of Bsh45I

<>

<1>Serva, S., Klimasauskas, S., Weinhold, E.
<2>Fluorescence studies of the DNA base flipping induced by a cytosine-5 methyltransferase.
<3>Biologija
<4>1
<5>9-12
<6>1997
<7>A novel fluorescence-based method for detecting and studying DNA base flipping in enzyme-DNA
complexes is described.  The target cytosine for the DNA methyltransferase HhaI (GCGC) was
replaced by a fluorescent base, 2-aminopurine.  Constistent with the extrahelical trapping of
the target base, a 90-fold increase in the fluorescence intensity was observed upon binding of
M. HhaI to a 37-mer duplex substrate.  Similar substitutions of bases adjacent to the target
cytosine result in a relatively small change.  Stopped-flow fluorescence experiments under
non-catalytic conditions allowed to monitor the course of base-flipping directly.  Kinetic
analysis shows that the site-specific binding of the enzyme is a diffusion-controlled process,
while the subsequent flipping motion is achieved in 1 millisecond, or faster.  Other classes
of enzymes suspected to employ the base flipping in their mechanisms can be investigated with
this method.

<>

<1>Serva, S., Velyvis, R., Lazareviciute, L., Klimasauskas, S.
<2>High-level expression of MvaI DNA methyltransferase.
<3>Biologija
<4>0
<5>48-50
<6>1996
<7>Construction of a high level expression system for the DNA-(cytosine-N4)-methyltransferase
M.MvaI is described.  The entire M.MvaI coding region was introduced as a 1511bp Alw44I-BpiI
fragment into the high expression vector pPR594E9.  The resultant plasmid (pPR-MvaIM7) was
expressed in the E. coli strain GM119 by induction with IPTG.  In the induced cells,
catalytically active M.MvaI was produced at a level of about 10% of their total protein.  The
enzyme was purified to apparent homogeneity by a four-column chromatographic procedure in a
yield of about 0.2 mg of pure enzyme per 1g of cell paste.  The molecular weight and the amino
terminal sequence of the protein agrees with that predicted from the DNA sequence.

<>

<1>Serva, S., Weinhold, E., Klimasauskas, S.
<2>Stopped-flow fluorescence studies of DNA base flipping by HhaI methyltransferase.
<3>Biochem. Soc. Trans.
<4>2000
<5>A468
<6>2000
<7>Rotation of a nucleotide out of the DNA helix (base flipping) has been first observed for the
HhaI methyltransferase followed by numerous other DNA modification and repair enzymes.  M.HhaI
catalyzes transfer of a methyl group from cofactor S-adenosyl-L-methionine onto the C5
position of the first cytosine in the target sequence GCGC.  In this study, stopped-flow and
rapid-quench techniques were employed in combination with fluorescence detection for kinetic
characterization of individual steps on the reaction pathway of M.HhaI.  Selective labeling of
the DNA substrate was achieved by synthetic incorporation of 2-aminopurine at the target
position which showed a dramatic increase in fluorescence signal upon transition of the base
from the stacked to an extrahelical position.  Association, dissociation and single-turnover
experimental setups permitted the direct determination of microscopic rate constants for DNA
binding, flipping of the target base, methyl group transfer and release of methylated DNA.  We
demonstrate here that the target sites on short DNA duplexes are predominantly located via a
diffusion-collision pathway and the subsequent base flipping is nearly instantaneous (<1 ms)
for the G-2AP mismatch.  We show for the first time that the chemical methyl transfer step
(0.15 s-1) is faster than the steady-state turnover (0.02 s-1) and directly confirm that decay
of the ternary product complex (methylated DNA-M.HhaI-AdoHcy) is rate-limiting in the
catalytic cycle.  Global fitting and simulation analysis in conjunction with structural data
suggest detailed catalytic mechanisms for DNA base flipping and catalysis by DNA cytosine-5
methyltransferases.

<>

<1>Serva, S., Weinhold, E., Roberts, R.J., Klimasauskas, S.
<2>Chemical display of thymine residues flipped out by DNA methyltransferases.
<3>Nucleic Acids Res.
<4>26
<5>3473-3479
<6>1998
<7>The DNA cytosine-C5 methyltransferase M.HhaI flips its target base out of the DNA helix during
interaction with the substrate sequence GCGC.  Binary and ternary complexes between M.HhaI and
hemimethylated DNA duplexes were used to examine the suitability of four chemical methods to
detect flipped-out bases in protein-DNA complexes.  These methods probe the structural
peculiarities of pyrimidine bases in DNA.  We find that in cases when the target cytosine is
replaced with thymine (GTGC), KMnO4 proved an efficient probe for positive display of
flipped-out thymines.  The generality of this procedure was further verified by examining a
DNA adenine-N6 methyltransferase, M.TaqI, in which case an enhanced reactivity of thymine
replacing the target adenine (TCGT) in the recognition sequence TCGA was also observed.  Our
results support the proposed base-flipping mechanism for adenine methyltransferases, and offer
a convenient laboratory tool for detection of flipped-out thymines in protein-DNA complexes.

<>

<1>Servin-Garciduenas, L.E., Garrett, R.A., Amils, R., Martinez-Romero, E.
<2>Genome Sequence of the Acidophilic Bacterium Acidocella sp. Strain MX-AZ02.
<3>Genome Announcements
<4>1
<5>e00041-12
<6>2013
<7>Here, we report the draft genome sequence of Acidocella sp. strain MX-AZ02, an acidophilic and
heterotrophic alphaproteobacterium isolated from a geothermal lake in western Mexico.

<>

<1>Servin-Garciduenas, L.E., Martinez-Romero, E.
<2>Draft Genome Sequence of the Sulfolobales Archaeon AZ1, Obtained through Metagenomic Analysis of a Mexican Hot Spring.
<3>Genome Announcements
<4>2
<5>e00164-14
<6>2014
<7>The Sulfolobales archaea have been found inhabiting acidic hot springs all over the world.
Here, we report the 1.798-Mbp draft genome sequence of the
thermoacidophilic Sulfolobales archaeon AZ1, reconstructed from the metagenome of
a Mexican hot spring. Sequence-based comparisons revealed that the Sulfolobales
archaeon AZ1 represents a novel candidate genus.

<>

<1>Servin-Garciduenas, L.E., Peng, X., Garrett, R.A., Martinez-Romero, E.
<2>Genome sequence of a novel archaeal fusellovirus assembled from the metagenome of a mexican hot spring.
<3>Genome Announcements
<4>1
<5>e00164-13
<6>2013
<7>The consensus genome sequence of a new member of the family Fuselloviridae designated as SMF1
(Sulfolobales Mexican fusellovirus 1) is presented. The
complete circular genome was recovered from a metagenomic study of a Mexican hot
spring. SMF1 exhibits an exceptional coding strand bias and a reduced set of
fuselloviral core genes.

<>

<1>Servin-Garciduenas, L.E., Rogel, M.A., Ormeno-Orrillo, E., Delgado-Salinas, A., Martinez-Romero, J., Sanchez, F., Martinez-Romero, E.
<2>Genome Sequence of Rhizobium sp. Strain CCGE510, a Symbiont Isolated from Nodules of the Endangered Wild Bean Phaseolus albescens.
<3>J. Bacteriol.
<4>194
<5>6310-6311
<6>2012
<7>We present the genome sequence of Rhizobium sp. strain CCGE510, a nitrogen fixing bacterium
taxonomically affiliated with the R. leguminosarum-R. etli group,
isolated from wild Phaseolus albescens nodules grown in native pine forests in
western Mexico. P. albescens is an endangered bean species phylogenetically
related to P. vulgaris. In spite of the close host relatedness, Rhizobium sp.
CCGE510 does not establish an efficient symbiosis with P. vulgaris. This is the
first genome of a Rhizobium symbiont from a Phaseolus species other than P.
vulgaris, and it will provide valuable new insights about symbiont-host
specificity.

<>

<1>Servin-Garciduenas, L.E., Sanchez-Quinto, A., Martinez-Romero, E.
<2>Draft Genome Sequence of Commensalibacter papalotli MX01, a Symbiont Identified from the Guts of Overwintering Monarch Butterflies.
<3>Genome Announcements
<4>2
<5>e00128-14
<6>2014
<7>We report the draft genome sequence of Commensalibacter papalotli strain MX01, isolated from
the intestines of an overwintering monarch butterfly. The
2,332,652-bp AT-biased genome of C. papalotli MX01 is the smallest genome for a
member of the Acetobacteraceae family and provides the first evidence of plasmids
in Commensalibacter.

<>

<1>Servos, S., Silva, C., Dougan, G., Charles, I.G.
<2>The commercially available restriction enzyme BspHI is blocked by overlapping methylation.
<3>Nucleic Acids Res.
<4>19
<5>183
<6>1991
<7>This report shows that BspHI is sensitive to dam methylation

<>

<1>Seshadri, R. et al.
<2>Genome sequence of the PCE-Dechlorinating bacterium Dehalococcoides ethenogenes.
<3>Science
<4>307
<5>105-108
<6>2005
<7>Dehalococcoides ethenogenes is the only bacterium known to reductively dechlorinate the
groundwater pollutants, tetrachloroethene (PCE) and trichloroethene, to ethene. Its
1,469,720-base pair chromosome contains large dynamic duplicated regions and integrated
elements. Genes encoding 17 putative reductive dehalogenases, nearly all of which were
adjacent to genes for transcription regulators, and five hydrogenase complexes were
identified. These findings, plus a limited repertoire of other metabolic modes, indicate that
D. ethenogenes is highly evolved to utilize halogenated organic compounds and H2.
Diversification of reductive dehalogenase functions appears to have been mediated by recent
genetic exchange and amplification. Genome analysis provides insights into the organism's
complex nutrient requirements and suggests that an ancestor was a nitrogen-fixing autotroph.

<>

<1>Seshadri, R., Joseph, S.W., Chopra, A.K., Sha, J., Shaw, J., Graf, J., Haft, D., Wu, M., Ren, Q., Rosovitz, M.J., Madupu, R., Tallon, L., Kim, M., Jin, S., Vuong, H., Stine, O.C., Ali, A., Horneman, A.J., Heidelberg, J.F.
<2>Genome Sequence of Aeromonas hydrophila ATCC 7966T: Jack of All Trades.
<3>J. Bacteriol.
<4>188
<5>8272-8282
<6>2006
<7>The complete genome of Aeromonas hydrophila ATCC 7966(T) was sequenced. Aeromonas, a
ubiquitous waterborne bacterium, has been placed by the
Environmental Protection Agency on the Contaminant Candidate List because
of its potential to cause human disease. The 4.7-Mb genome of this
emerging pathogen shows a physiologically adroit organism with broad
metabolic capabilities and considerable virulence potential. A large array
of virulence genes, including some identified in clinical isolates of
Aeromonas spp. or Vibrio spp., may confer upon this organism the ability
to infect a wide range of hosts. However, two recognized virulence
markers, a type III secretion system and a lateral flagellum, that are
reported in other A. hydrophila strains are not identified in the
sequenced isolate, ATCC 7966(T). Given the ubiquity and free-living
lifestyle of this organism, there is relatively little evidence of
fluidity in terms of mobile elements in the genome of this particular
strain. Notable aspects of the metabolic repertoire of A. hydrophila
include dissimilatory sulfate reduction and resistance mechanisms (such as
thiopurine reductase, arsenate reductase, and phosphonate degradation
enzymes) against toxic compounds encountered in polluted waters. These
enzymes may have bioremediative as well as industrial potential. Thus, the
A. hydrophila genome sequence provides valuable insights into its ability
to flourish in both aquatic and host environments.

<>

<1>Seshasayee, A.S.
<2>An assessment of the role of DNA adenine methyltransferase on gene expression regulation in E coli.
<3>PLoS ONE
<4>2
<5>e273
<6>2007
<7>N6-Adenine methylation is an important epigenetic signal, which regulates various processes,
such as DNA replication and repair and transcription. In gamma-proteobacteria, Dam is a
stand-alone enzyme that methylates GATC sites, which are non-randomly distributed in the
genome. Some of these overlap with transcription factor binding sites. This work describes a
global computational analysis of a published Dam knockout microarray alongside other publicly
available data to throw insights into the extent to which Dam regulates transcription by
interfering with protein binding. The results indicate that DNA methylation by DAM may not
globally affect gene transcription by physically blocking access of transcription factors to
binding sites. Down-regulation of Dam during stationary phase correlates with the activity of
TFs whose binding sites are enriched for GATC sites.

<>

<1>Seshasayee, A.S., Singh, P., Krishna, S.
<2>Context-dependent conservation of DNA methyltransferases in bacteria.
<3>Nucleic Acids Res.
<4>40
<5>7066-7073
<6>2012
<7>DNA methytransferases (MTs) in bacteria are best understood in the context of
restriction-modification (R-M) systems, which act as bacterial immune systems
against incoming DNA including phages, but have also been described as selfish
elements. But several orphan MTs, which are not associated with any restriction
enzyme, have also been characterized and may protect against parasitism by R-M
systems. The occurrence of MTs in these two contexts, namely as part of R-M
systems or as orphans, is poorly understood. Here we report the results of a
comparative genomic survey of DNA MTs across approximately 1000 bacterial
genomes. We show that orphan MTs overwhelm R-M systems in their occurrence. In
general, R-M MTs are poorly conserved, whereas orphans are nearly as conserved
within a genus as any average gene. However, oligonucleotide usage and
conservation patterns across genera suggest that both forms of MTs might have
been horizontally acquired. We suggest that many orphan MTs might be
'degradation' products of R-M systems, based on the properties of orphan MTs
encoded adjacent to highly diverged REs. In addition, several fully degraded R-M
systems exist in which both the MT and the RE are highly divergent from their
corresponding reference R-M pair. Despite their sporadic occurrence, conserved
R-M systems are present in strength in two highly transformable genera, in which
they may contribute to selection against integration of foreign DNA.

<>

<1>Sethmann, S., Ceglowski, P., Willert, J., Iwanicka-Nowicka, R., Trautner, T.A., Walter, J.
<2>M.PhiBssHII, a novel cytosine-C5-DNA-methyltransferase with target-recognizing domains at separated locations of the enzyme.
<3>EMBO J.
<4>18
<5>3502-3508
<6>1999
<7>In all cytosine-C5-DNA-methyltransferases (MTases) from prokaryotes and eukaryotes, remarkably
conserved amino acid sequence elements responsible for general enzymatic functions are
arranged in the same canonical order. In addition, one variable region, which includes the
target-recognizing domain(s) (TRDs) characteristic for each enzyme, has been localized in one
region between the same blocks of these conserved elements. This conservation in the order of
conserved and variable sequences suggests stringent structural constraints in the primary
structure to obtain the correct folding of the enzymes. Here we report the characterization of
a new type of a multispecific MTase, M.PhiBssHII, which is expressed as two isoforms. Isoform
I is an entirely novel type of MTase which has, in addition to the TRDs at the conventional
location, one TRD located at a non-canonical position at its N-terminus. Isoform II is
represented by the same MTase, but without the N-terminal TRD. The N-terminal TRD provides
HaeII methylation specificity to isoform I. The TRD is fully functional when engineered into
either the conventional variable region of M.PhiBssHII or the related monospecific M.Phi3TII
MTase. The implications of this structural plasticity with respect to the evolution of MTases
are discussed.

<>

<1>Sethuraman, J., Majer, A., Friedrich, N.C., Edgell, D.R., Hausner, G.
<2>Genes within genes: multiple LAGLIDADG homing endonucleases target the ribosomal protein S3 gene encoded within an rnl group I intron of  Ophiostoma and related taxa.
<3>Mol. Biol. Evol.
<4>26
<5>2299-2315
<6>2009
<7>In some ascomycete fungi, ribosomal protein S3 (Rps3) is encoded within a group I intron
(mL2449) that is inserted in the U11 region of the
mitochondrial large subunit rDNA (rnl) gene. Previous characterization of
the mL2449 intron in strains of Ophiostoma novo-ulmi subspecies americana
(Dutch Elm Disease) revealed a complex genes-within-genes arrangement
whereby a LAGLIDADG homing endonuclease gene (HEG) is inserted into the
RPS3 gene near the 3' terminus, creating a hybrid Rps3-LAGLIDADG fusion
protein. Here, we examined 119 additional strains of Ophiostoma and
related taxa representing 85 different species by a polymerase chain
reaction- based survey and detected both short (approximately 1.6 kb) and
long (>2.2 kb) versions of the mL2449 intron in 88 and 31 strains,
respectively. Among the long versions encountered, 21 were sequenced,
revealing the presence of either intact or degenerated HEG-coding regions
inserted within the RPS3 gene. Surprisingly, we identified two new HEG
insertion sites in RPS3; one near the original C-terminal insertion site
and one near the N-terminus of RPS3. In all instances, the HEGs are fused
in-frame with the RPS3-coding sequences to create fusion proteins.
However, comparative sequence analysis showed that upon insertion, the
HEGs displaced a portion of the RPS3-coding region. Remarkably, the
displaced RPS3-coding segments are duplicated and fused in-frame to the 3'
end of RPS3, restoring a full-length RPS3 gene. We cloned and expressed
the LAGLIDADG portion of two Rps3-HEG fusions, and showed that I-OnuI and
I-LtrI generate 4 nucleotide (nt), 3' overhangs, and cleave at or 1 nt
upstream of the HEG insertion site, respectively. Collectively, our data
indicate that RPS3 genes are a refuge for distinct types of LAGLIDADG HEGs
that are defined by the presence of duplicated segments of the host gene
that restore the RPS3 gene, thus minimizing the impact of the HEG
insertion on Rps3 function.

<>

<1>Setubal, J.C. et al.
<2>Genome sequence of Azotobacter vinelandii, an obligate aerobe specialized to support diverse anaerobic metabolic processes.
<3>J. Bacteriol.
<4>191
<5>4534-4545
<6>2009
<7>Azotobacter vinelandii is a soil bacterium related to the Pseudomonas genus that fixes
nitrogen under aerobic conditions while simultaneously
protecting nitrogenase from oxygen damage. In response to carbon
availability, this organism undergoes a simple differentiation process to
form cysts that are resistant to drought and other physical and chemical
agents. Here we report the complete genome sequence of A. vinelandii DJ,
which has a single circular genome of 5,365,318 bp. In order to reconcile
an obligate aerobic lifestyle with exquisitely oxygen-sensitive processes,
A. vinelandii is specialized in terms of its complement of respiratory
proteins. It is able to produce alginate, a polymer that further protects
the organism from excess exogenous oxygen, and it has multiple
duplications of alginate modification genes, which may alter alginate
composition in response to oxygen availability. The genome analysis
identified the chromosomal locations of the genes coding for the three
known oxygen-sensitive nitrogenases, as well as genes coding for other
oxygen-sensitive enzymes, such as carbon monoxide dehydrogenase and
formate dehydrogenase. These findings offer new prospects for the wider
application of A. vinelandii as a host for the production and
characterization of oxygen-sensitive proteins.

<>

<1>Seurinck, J., Van de Voorde, A., Van Montagu, M.
<2>A new restriction endonuclease from Acetobacter pasteurianus.
<3>Nucleic Acids Res.
<4>11
<5>4409-4415
<6>1983
<7>A restriction endonuclease, ApaI, has been partially purified from Acetobacter
pasteurianus.  This enzyme cleaves bacteriophage lambda DNA and Simian virus 40
DNA at one site, adenovirus-2 DNA at more than nine sites, but it does not
cleave PhiX174 DNA nor plasmid pBR322 DNA.  This enzyme recognizes the
sequence5' GGGCC^C 3' 3' C^CGGG 5'and cuts at the sites indicated by the
arrows.

<>

<1>Shabarova, Z.A., Sheflyan, G.Y., Kuznetsova, S.A., Kubareva, E.A., Sysoev, O.N., Ivanovskaya, M.G., Gromova, E.S.
<2>Affinity modification of restriction-endonuclease EcoRII by DNA duplex containing a monosubstituted pyrophosphate internucleotide bond.
<3>Bioorg. Khim.
<4>20
<5>413-419
<6>1994
<7>An oligonucleotide duplex with an active monosubstituted pyrophosphate bond within the
recognition site of the EcoRII restriction endonuclease was cross-linked to this enzyme with a
yield of 10-15%. The cross-linking specificity was proved by the absence of the cross-linking
to a DNA duplex with the same modification but without the EcoRII recognition site as well as
by unmodified EcoRII substrate's inhibition of the cross-linking.

<>

<1>Shah, G.R., Karunakaran, R., Kumar, G.N.
<2>In vivo restriction endonuclease activity of the Anabaena PCC 7120 XisA protein in Escherichia coli.
<3>Res. Microbiol.
<4>158
<5>679-684
<6>2007
<7>Anabaena PCC 7120 genome contains three elements, which get excised out during late stages of
heterocyst differentiation by a site-specific
recombination process. The XisA protein, which excises the nifD
element, shows sequence homology with the integrase family of tyrosine
recombinase. The 11 by target site of XisA CGGAGTAATCC contains a 3 bp
inverted repeat. Here, we report restriction endonuclease activity of
XisA by specific loss of plasmids containing single or double target
sites. The pMX25 plasmid containing two tat-get sites demonstrated
endonuclease activity proportional to excision frequency. Different
plasmid substrates containing one base pair mutation in the inverted
repeat of the target site were monitored for endonuclease activity.
Mutation of A4C retained endonuclease activity, while other
modifications lost endonuclease activity. The presence of an additional
copy of the target site enhanced endonuclease activity. These results
suggest that the XisA protein could be an IIE type of restriction
endonuclease in addition to being a recombinase. (C) 2007 Elsevier
Masson SAS. All rights reserved.

<>

<1>Shah, N.R., Moksa, M., Novikov, A., Perry, M.B., Hirst, M., Caroff, M., Fernandez, R.C.
<2>Draft Genome Sequences of Bordetella hinzii and Bordetella trematum.
<3>Genome Announcements
<4>1
<5>e00838-13
<6>2013
<7>Bordetella hinzii colonizes the respiratory tracts of poultry but can also infect
immunocompromised humans. Bordetella trematum, however, only infects humans,
causing ear and wound infections. Here, we present the first draft genome
sequences of strains B. hinzii ATCC 51730 and B. trematum CCUG 13902.

<>

<1>Shahinas, D., Tamber, G.S., Arya, G., Wong, A., Lau, R., Jamieson, F., Ma, J.H., Alexander, D.C., Low, D.E., Pillai, D.R.
<2>Whole-Genome Sequence of Streptococcus pseudopneumoniae Isolate IS7493.
<3>J. Bacteriol.
<4>193
<5>6102-6103
<6>2011
<7>Streptococcus pseudopneumoniae is a member of the viridans group streptococci (VGS) whose
pathogenic significance is unclear. We announce
the complete genome sequence of S. pseudopneumoniae IS7493. The genome
sequence will assist in the characterization of this new organism and
facilitate the development of accurate diagnostic assays to distinguish it
from Streptococcus pneumoniae and Streptococcus mitis.

<>

<1>Shajani, Z., Varani, G.
<2>C-13 relaxation studies of the DNA target sequence for HhaI methyltransferase reveal unique motional properties.
<3>Biochemistry
<4>47
<5>7617-7625
<6>2008
<7>The goal of this work was to examine if sequence-dependent conformational flexibility in DNA
plays a role in base extrusion, a
common conformational change induced by many DNA-modifying enzymes. We
studied the dynamics of the double-stranded DNA target of the HhaI
methyltransferase by recording an extensive set of C-13 NMR relaxation
parameters. We observe that the cytidine furanose rings experience fast
(picosecond to nanosecond) motions that are not present in other
nucleotides; the methylation site experiences particularly high
mobility. We also observe that the bases of guanosine and cytidine
residues within the HhaI recognition sequence GCGC experience motions
on a much slower (1-100 mu s) time scale. We compare these observations
with previous solution and solid-state NMR studies of the EcoRI
nuclease target sequence, and solid-state NMR studies of a similar HhaI
target construct. While an increased mobility of cytidine furanose
rings compared to those of other nucleotides is observed for both
sequences, the slower motions are only observed in the HhaI target DNA.
We propose that this inherent flexibility lowers the energetic barriers
that must occur when the DNA binds to the HhaI methyltransferase and
for extrusion of the cytidine prior to its methylation.

<>

<1>Shambat, S., Nadig, S., Prabhakara, S., Bes, M., Etienne, J., Arakere, G.
<2>Clonal complexes and virulence factors of Staphylococcus aureus from several cities in India.
<3>BMC Microbiol.
<4>12
<5>64
<6>2012
<7>BACKGROUND: Diseases from Staphylococcus aureus are a major problem in Indian
hospitals and recent studies point to infiltration of community associated
methicillin resistant S. aureus (CA-MRSA) into hospitals. Although CA-MRSA are
genetically different from nosocomial MRSA, the distinction between the two
groups is blurring as CA-MRSA are showing multidrug resistance and are endemic in
many hospitals. Our survey of samples collected from Indian hospitals between
2004 and 2006 had shown mainly hospital associated methicillin resistant
Staphylococcus aureus (HA-MRSA) carrying staphylococcal cassette chromosome mec
(SCCmec) type III and IIIA. But S. aureus isolates collected from 2007 onwards
from community and hospital settings in India have shown SCCmec type IV and V
cassettes while several variations of type IV SCCmec cassettes from IVa to IVj
have been found in other parts of the world. In the present study, we have
collected nasal swabs from rural and urban healthy carriers and pus, blood etc
from in patients from hospitals to study the distribution of SCCmec elements and
sequence types (STs) in the community and hospital environment. We performed
molecular characterization of all the isolates to determine their lineage and
microarray of select isolates from each sequence type to analyze their toxins,
virulence and immune-evasion factors. RESULTS: Molecular analyses of 68 S. aureus
isolates from in and around Bengaluru and three other Indian cities have been
carried out. The chosen isolates fall into fifteen STs with all major clonal
complexes (CC) present along with some minor ones. The dominant MRSA clones are
ST22 and ST772 among healthy carriers and patients. We are reporting three novel
clones, two methicillin sensitive S. aureus (MSSA) isolates belonging to ST291
(related to ST398 which is live stock associated), and two MRSA clones, ST1208
(CC8), and ST672 as emerging clones in this study for the first time. Sixty nine
percent of isolates carry Panton- Valentine Leucocidin genes (PVL) along with
many other toxins. There is more diversity of STs among methicillin sensitive S.
aureus than resistant ones. Microarray analysis of isolates belonging to
different STs gives an insight into major toxins, virulence factors, adhesion and
immune evasion factors present among the isolates in various parts of India.
CONCLUSIONS: S. aureus isolates reported in this study belong to a highly diverse
group of STs and CC and we are reporting several new STs which have not been
reported earlier along with factors influencing virulence and host pathogen
interactions.

<>

<1>Shan, D., Li, X., Gu, Z., Wei, G., Gao, Z., Shao, Z.
<2>Draft Genome Sequence of the Agar-Degrading Bacterium Catenovulum sp. Strain DS-2, Isolated from Intestines of Haliotis diversicolor.
<3>Genome Announcements
<4>2
<5>e00144-14
<6>2014
<7>Catenovulum sp. strain DS-2, isolated from intestines of Haliotis diversicolor, is able to
degrade agar and produce agaro-oligosaccharides. Here, we report the
draft genome sequence of Catenovulum sp. strain DS-2.

<>

<1>Shan, D., Ying, J., Li, X., Gao, Z., Wei, G., Shao, Z.
<2>Draft Genome Sequence of the Carrageenan-Degrading Bacterium Cellulophaga sp. Strain KL-A, Isolated from Decaying Marine Algae.
<3>Genome Announcements
<4>2
<5>e00145-14
<6>2014
<7>Cellulophaga sp. strain KL-A, isolated from decaying marine algae, is able to degrade
iota-carrageenan. Here, we report the draft genome sequence of
Cellulophaga sp. strain KL-A.

<>

<1>Shan, W., Yang, X., Ma, W., Yang, Y., Guo, X., Guo, J., Zheng, H., Li, G., Xie, B.
<2>Draft Genome Sequence of Ralstonia solanacearum Race 4 Biovar 4 Strain SD54.
<3>Genome Announcements
<4>1
<5>e00890-13
<6>2013
<7>Ralstonia solanacearum is an important etiological agent that can cause serious bacterial wilt
in a very wide range of potential host plants, including ginger.
Here, we report the complete genome sequence of R. solanacearum SD54, a race 4
biovar 4 (R4B4) strain from a diseased ginger plant in China.

<>

<1>Shane, J.L., Bongio, N.J., Favia, G., Lampe, D.J.
<2>Draft Genome Sequence of Asaia sp. Strain SF2.1, an Important Member of the Microbiome of Anopheles Mosquitoes.
<3>Genome Announcements
<4>2
<5>e01202-13
<6>2014
<7>Asaia spp. are abundant members of the microbiota of Anopheles mosquitoes, the principle
vectors of malaria. Here, we report the draft genome sequence of Asaia
sp. strain SF2.1. This strain is under development as a platform to deliver
antimalarial peptides and proteins to adult female Anopheles mosquitoes.

<>

<1>Shang, Y., Zhang, N., Zhu, P., Luo, Y., Huang, K., Tian, W., Xu, W.
<2>Restriction enzyme cutting site distribution regularity for DNA looping technology.
<3>Gene
<4>534
<5>222-228
<6>2014
<7>The restriction enzyme cutting site distribution regularity and looping conditions were
studied systematically. We obtained the restriction enzyme
cutting site distributions of 13 commonly used restriction enzymes in 5 model
organism genomes through two novel self-compiled software programs. All of the
average distances between two adjacent restriction sites fell sharply with
increasing statistic intervals, and most fragments were 0-499bp. A shorter DNA
fragment resulted in a lower looping rate, which was also directly proportional
to the DNA concentration. When the length was more than 500bp, the concentration
did not affect the looping rate. Therefore, the best known fragment length was
longer than 500bp, and did not contain the restriction enzyme cutting sites which
would be used for digestion. In order to make the looping efficiencies reach
nearly 100%, 4-5 single cohesive end systems were recommended to digest the
genome separately.

<>

<1>Shankar, M., Ponraj, P., Ilakiam, D., Rajendhran, J., Gunasekaran, P.
<2>Genome Sequence of the Plant Growth-Promoting Bacterium Enterobacter cloacae GS1.
<3>J. Bacteriol.
<4>194
<5>4479
<6>2012
<7>Here, we present the genome sequence of Enterobacter cloacae GS1. This strain proficiently
colonizes rice roots and promotes plant growth by improving plant
nutrition. Analyses of the E. cloacae GS1 genome will throw light on the genetic
factors involved in root colonization, growth promotion, and ecological success
of this rhizobacterium.

<>

<1>Shankar, S., Tyagi, A.K.
<2>MhaAI, a novel isoschizomer of PstI from Mycobacterium habana recognizing 5'-CTGCA/G-3'.
<3>Nucleic Acids Res.
<4>20
<5>2891
<6>1992
<7>We have isolated a novel class II restriction endonuclease, MhaAI, from Mycobacterium habana
strain 206 which recognizes the sequence 5'-CTGCA/G-3' and generates 3' protruding
fragments. Unlike its isoschizomer PstI, MhaAI did not exhibit star activity even at glycerol
concentrations as high as 35% and at units/ug DNA ratios greater than 300. MhaAI was fully
active in low salt (10 mM Tris pH 8.0, 10 mM MgCl2) in comparison to PstI, which requires NaCl
at a concentration of 50-100 mM for optimal activity. MhaAI was also found equally active in
10 to 200 mM concentration range of KCl as well as MgCl2. The enzyme, however, was inhibited
by NaCl concentrations greater than 50 mM.

<>

<1>Shankar, S., Tyagi, A.K.
<2>MfoAI, a novel isoschizomer of HaeIII from Mycobacterium fortuitum recognizing 5'-GG/CC-3' .
<3>Nucleic Acids Res.
<4>20
<5>2890
<6>1992
<7>We have isolated a novel class II restriction endonuclease, MfoAI, from Mycobacterium
fortuitum TMC 1529 which recognizes the sequence 5'-GG/CC-3' generating blunt ended
fragments. MfoAI was found to have a very high affinity for phosphocellulose and eluted late
at a concentration of 1.8 M KCl. This permitted very rapid purification of the enzyme
completely free from non-specific nucleases. In contrast to the situation in Haemophilus
aegyptius which has HaeIII and HaeII, MfoAI was found to be the only activity associated with
M. fortuitum TMC 1529, suggesting this organism to be a better source for this enzyme. In
addition, the activity of MfoAI was independent of salt concentration and purified MfoAI was
equally active in 10-200 mM concentrations of KCl, MgCl2 and NaCl and in the wide pH range of
7.4-11.0.

<>

<1>Shankar, S., Tyagi, A.K.
<2>Purification and characterization of restriction endonuclease MgoI from Mycobacterium gordonae.
<3>Gene
<4>131
<5>153-154
<6>1993
<7>A restriction endonuclease, MgoI an isoschizomer of Sau3AI, was purified from Mycobacterium
gordonae TRC1318. As compared to Sau3AI, the yield of MgoI was seven-eight fold higher, the
enzyme was four times more stable at 37C, and in addition, had optimal activity over a much
broader range of salt concentrations.

<>

<1>Shankar, S., Tyagi, A.K.
<2>MchAI and MchAII, two class-II restriction endonucleases from Mycobacterium chelonei.
<3>Gene
<4>132
<5>119-123
<6>1993
<7>We have purified and characterized two class-II restriction endonucleases from the saprophyte
Mycobacterium chelonei AMB 82. MchAI was an isoschizomer of NotI recognizing and cleaving at
5'-GC/GGCCGC. MchAI did not require Triton X-100 for activity and was fully active at
concentrations over 10-200 mM KCl and MgCl2. MchAII was an isoschizomer of HaeIII recognizing
and cleaving at 5'-GG/CC. MchAII was active in 10 mM and was inactive at KCl concentrations
lower than 250 mM.

<>

<1>Shankarappa, B., Vijayananda, K., Ehrlich, G.
<2>Silmut: A Computer Program for the Identification of Regions Suitable for Silent Mutagenesis to Introduce Restriction Enzyme Recognition Sequences.
<3>Biotechniques
<4>12
<5>882-884
<6>1992
<7>We describe a set of IBM-compatible computer programs designed to selectively identify the
potential sites for silent mutagenesis within a target DNA sequence. This program is based on
a novel strategy of identifying amino acid motifs compatible with each restriction site
(BioTechniques 12:382-384m 1991). The programs can be used to identify the suitability for the
introduction of any 6-base nucleic acid sequences, such as restriction enzyme sites in
cassette mutagenesis strategies. The Table program generates a table of multiple amino acid
motifs for each restriction enzyme, obtained by translating each unique recognition sequence
in all three reading frames. The Silmut program which utilizes the features of Table, will
further identify the presence of a match betwen any amino acid motif of each restriction
enzyme and the input target sequence. Minor manipulations of the database files will enable
the individual researcher to identify the potential for introduction of any 6-base sequences
by silent mutagenesis.

<>

<1>Shao, C., Wang, C., Zang, J.
<2>Structural basis for the substrate selectivity of PvuRts1I, a 5-hydroxymethylcytosine DNA restriction endonuclease.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>70
<5>2477-2486
<6>2014
<7>5-Hydroxymethylation is a curious modification of cytosine that was discovered some decades
ago, but its functional role in eukaryotes still awaits elucidation. 5-Hydroxymethylcytosine
is an epigenetic marker that is crucial for multiple biological processes. The profile is
altered under certain disease conditions such as cancer, Huntington's disease and
Alzheimer's disease. Using the DNA-modification-dependent restriction endonuclease AbaSI
coupled with sequencing (Aba-seq), the hydroxymethylome can be deciphered at the resolution of
individual bases. The method is based on the enzymatic properties of AbaSI, a member of the
PvuRts1I family of endonucleases. PvuRts1I is a modification-dependent endonuclease with high
selectivity for 5-hydroxymethylcytosine over 5-methylcytosine and cytosine. In this study, the
crystal structure of PvuRts1I was determined in order to understand and improve the substrate
selectivity. A nuclease domain and an SRA-like domain are located at the N- and C-termini,
respectively. Through comparison with other SRA-domain structures, the SRA-like domain was
proposed to be the 5-hmC recognition module. Several mutants of PvuRts1I with enzymatic
activity restricted to 5-hydroxymethylcytosine only were generated based on the structural
analysis, and these enzyme variants are appropriate for separating the hydroxymethylome from
the wider methylome.

<>

<1>Shao, Y., Xu, M.-Q., Paulus, H.
<2>Protein splicing: Evidence for an N-O acyl rearrangement as the initial step in the splicing process.
<3>Biochemistry
<4>35
<5>3810-3815
<6>1996
<7>Protein splicing involves the self-catalyzed formation of a branched
intermediate, which then resolves into the excised intervening sequence and the spliced
protein.  A possible mechanism for branched intemediate formation is an N-O
rearrangement of the peptide bond involving the amino group of the conserved
serine/cysteine residue at the upstream splice junction to yield a linear peptide ester
intermediate.  This possibility was examined using an in vitro splicing system involving the
intervening sequence from the DNA polymerase of the extremely thermophilic archeon,
Pyrococcus sp. GB-D.  Because thioesters react much more rapidly with nitrogen
nucleophiles at neutral pH than do oxygen esters, protein-splicing precursors in which the
serine residue of interest was replaced by cysteine were constructed and purified.  In the
presence of 0.25M hydroxylamine or 0.1M ethylene diamine at pH 6 or higher, these
constructs underwent rapid cleavage at the upstream splice junction, consistent with the
aminolysis of a thioester.  The site of hydroxylaminolysis was identified by analysis of the
C-terminus of the polypeptide cleavage products.  Comparison of the C-terminal peptide
hydroxamate with the synthetic peptide hydroxamates with respect to chromatographic
mobility, colorimetric assay, amino acid composition, and high-resolution mass
spectrometry showed that the hydroxylamine-sensitive site in the splicing precursor was the
peptide bond adjacent to the serine residue at the upstream splice junction.  These results
provide evidence that the peptide bond at the upstream splice junction can undergo a self-
catalyzed N-O or N-S acyl rearrangement to yield a linear polypeptide ester intermediate
and suggest that this kind of rearrangement constitutes the first step in protein splicing.

<>

<1>Shao, Y., Xu, M.-Q., Paulus, H.
<2>Protein splicing: characterization of the aminosuccinimide residue at the carboxyl terminus of the excised intervening sequence.
<3>Biochemistry
<4>34
<5>10844-10850
<6>1995
<7>Protein splicing is a self-catalyzed, posttranslational process which
converts a precursor polypeptide into two new proteins by the excision of an internal
polypeptide segment and the ligation of the flanking polypeptides.  Evidence has been
presented that protein splicing involves a branched intermediate, which is resolved into the
two protein products by the cyclization of an asparagine residue to aminosuccinimide.  This
report describes the chemical synthesis of a peptide with a C-terminal aminosuccinimide
residue, corresponding to the putative C-terminus of the excised intervening sequence
(intein) derived from the thermostable DNA polymerase of Pyrococcus species GB-D.  The
synthetic aminosuccinimide peptide was compared with the C-terminal cyanogen bromide
peptide of the excised intein and found to be indistinguishable in terms of its
chromatographic properties, high-resolution mass spectrum, and colorimetric assay
involving reaction with hydroxylamine.  This establishes definitively that protein splicing is
accompanied by the cyclization of asparagine to yield an aminosuccinimide residue at the C-
terminus of the excised intein and that this unusual residue is therefore a natural
constituent
of spliced proteins.  The effects of pH and temperature on the stability of the synthetic
aminosuccinimide peptide are described.  The stability of the C-terminal aminosuccinimide
decreased with increasing pH, similar to the internal aminosuccinimide residues that occur
in many proteins as intermediates in protein deamidation, but the C-terminal
aminosuccinimide was 5-10 times more stable than internal aminosuccinimides, with a half-
life of about 80 h at 25oC and pH 7.4, accounting for its relative ease of isolation.

<>

<1>Shapiro, L., Benkovic, S.J., Wright, R., Stephens, C., Kahng, L.S., Berdis, A.
<2>DNA adenine methyltransferases and uses thereof.
<3>International Patent Office
<4>WO 9812206
<5>
<6>1998
<7>The present invention relates to the isolation and sequencing of a novel class of
methyltransferase genes, including the methyltransferase gene from Rhizobium meliloti,
Agrobacterium tumefaciens, Brucella abortus, and Helicobacter pylori.  The invention further
comprises efficient methods of assaying methyltransferase activity.

<>

<1>Shapovalova, N.I., Ivanov, L.Y., Matvienko, N.I.
<2>Site-specific endonuclease BstM6I from the natural isolate Bacillus stearothermophilus M6.
<3>Bioorg. Khim.
<4>18
<5>1186-1189
<6>1992
<7>A site-specific endonuclease activity was found in extracts of Bacillus stearothermophilus M6
isolated from molasses. The functionally pure enzyme designated as BstM6I was obtained by
consecutive chromatographies on blue sepharose, hydroxyapatite, and heparin-sepharose. The
endonuclease recognizes the nucleotide sequence CC^WGG in double-stranded DNA and cleaves it
as indicated by the arrow to give one-nucleotide 5'-protruding ends. Consequently, the
site-specific endonuclease BstM6I is an isoschizomer of BstNI.

<>

<1>Shapovalova, N.I., Zheleznaja, L.A., Matvienko, N.I.
<2>BspKT6I, a new site-specific endonuclease which cleaves the GATC site producing two nucleotide 5'-protruding ends.
<3>Nucleic Acids Res.
<4>21
<5>5794
<6>1993
<7>A new class II restriction endonuclease BspKT6I was isolated from soil thermophilic bacteria
Bacillus species KT6. The enzyme was purified by chromatography on blue-agarose,
heparin-Sepharose and hydroxylapatite. Digestion of T7 DNA has revealed that the enzyme is an
isoschizomer of Sau3AI. Cleavage points were determined by the primed-synthesis reaction using
M13mp18 DNA. The cleavage product resulted in a band that comigrated with G (Fig 1). Upon
addition of the DNA-polymerase, a band appeared two nucleotides higher. These results indicate
that the enzyme recognizes the palindromic sequence G^ATC and cleaves it as indicated creating
two nucleotide 5'-extensions. It is the first isomer of Sau3AI producing such ends. The DNA
fragments produced by this enzyme can be ligated with PvuI-produced DNA fragments. BspKT6I is
sensitive to dam-methylation. It does not cleave pBR322 isolated from the E. coli dam+ strain
and results in almost complete digestion of pBR322 isolated from the dam-strain (Fig. 2). The
optimal reaction buffer is 10 mM Tris-HCl (ph 7.5), 10 mM MgCl2.

<>

<1>Shapovalova, N.I., Zheleznaya, L.A., Matvienko, N.I.
<2>New site-specific endonuclease and methylase from the thermophilic strain Bacillus species KT6.
<3>Biokhimiia
<4>59
<5>1730-1738
<6>1994
<7>The site-specific endonuclease R.BspKT6I and the cognate site-specific methylase M.BspKT6I
were isolated to functionally pure state from the thermophilic strain Bacillus species KT6 by
Sephadex G-100 gel filtration followed by chromatography on heparin-Sepharose and
hydroxyapatite. Endonuclease BspKT6I is not an isoschizomer, but an isomer of Sau3AI and MboI.
It recognizes the GAT/C sequence and cleaves it as shown by arrow, and, in distinction to
Sau3AI and MboI, produces a protruding 3' dinucleotide. The cleavage of the site is inhibited
by dam methylation. The sticky ends resulting from BspKT6I cleavage are identical and
complementary to those formed after PvuI cleavage. The methylase isolated from B. species KT6
protects the DNA from subsequent cleavage by BspKT6I. The methylated base is adenine.

<>

<1>Shapovalova, N.I., Zheleznaya, L.A., Matvienko, N.N., Matvienko, N.I.
<2>A new site-specific endonuclease BspKT5I.
<3>Biokhimiia
<4>59
<5>485-493
<6>1994
<7>A new site-specific endonuclease (BspKT5I) has been isolated from the thermophilic soil
bacterium Bacillus KT5 by chromatography on Blue agarose, hydroxyapatite, and
heparin-Sepharose; it is free from interfering impurities. On double-stranded DNA, BspKT5I
recognizes the sequence 5'-CTGAAG16N^/3'-GACTTC14N^ and cleaves the DNA at the recognition
site indicated by the arrows to form 3'-protruding dinucleotide termini. The isolated
endonuclease is an isoschizomer of Eco57I. However, unlike Eco57I, it is not stimulated by
S-adenosylmethionine (SAM) and thus belongs to subclass IIS and not to class IV, as Eco57I.
Furthermore, endonuclease BspKT5I, in contrast to Eco57I, has no methylase activity.

<>

<1>Sharath, A.N., Weinhold, E., Bhagwat, A.S.
<2>Reviving a dead enzyme: cytosine deaminations promoted by an inactive DNA methyltransferase and an S-adenosylmethionine analogue.
<3>Biochemistry
<4>39
<5>14611-14616
<6>2000
<7>The enzymes that transfer a methyl group to C5 of cytosine within specific sequences (C5
Mtases) deaminate the target cytosine to uracil if the methyl donor S-adenosylmethionine (SAM)
is omitted from the reaction.  Recently, it was shown that cytosine deamination caused by C5
Mtases M.HpaII, M.SssI and M.MspI is enhanced in the presence of several analogues of SAM, and
a mechanism for this analogue-promoted deamination was proposed. According to this mechanism,
the analogues protonate C5 of the target cytosine, creating a dihydrocytosine intermediate
that is susceptible to deamination. We show here that one of these analogues,
5'-aminoadenosine (AA), enhances cytosine deamination by the Mtase M. EcoRII, but it does so
without enhancing protonation of C5. Further, we show that uracil is an intermediate in the
mutational pathway and propose an alternate mechanism for the analogue-promoted deamination.
The new mechanism involves a facilitated water attack at C4 but does not require attack at C6
by the enzyme. The latter feature of the mechanism was tested by using M.EcoRII mutants
defective in the nucleophilic attack at C6 in the deamination assay. We find that although
these proteins are defective in methyl transfer and cytosine deamination, they cause cytosine
deaminations in the presence of AA in the reaction. Our results point to a possible connection
between the catalytic mechanism of C5 Mtases and of enzymes that transfer methyl groups to
N(4) of cytosine. Further, they provide an unusual example where a coenzyme activates an
otherwise "dead" enzyme to perform catalysis by a new reaction pathway.

<>

<1>Sharkova, E.V., Nikolskaya, I.I., Shomodi, P., Feldesh, I., Debov, S.S.
<2>Isolation and fractionation of DNA-methylases from Mycobacterium butyricum.
<3>Vopr. Med. Khim.
<4>30
<5>72-76
<6>1984
<7>Properties of total preparation of methylases from M. butyricum strain were
studied using various methods of column chromatography.  Distinct heterogeneity
of the methylase preparation was demonstrated by gel filtration, anion exchange
chromatography on diethyl aminoethyl cellulose and affinity chromatography on
Sepharose blue.  Several methylases, dissimilar both in physico-chemical
properties and in their specificity to nitrogenous bases, were found in M.
butyricum cells.  In the strain studied methylases of adenine and cytosine were
found, which were responsible for biosynthesis of 6-methyladenine and
5-methylcytosine in the acceptor DNA at the ratio 9:1.

<>

<1>Sharkova, E.V., Nikolskaya, I.I., Shomodi, P., Feldesh, I., Debov, S.S.
<2>Effect of various conditions on stability of DNA-methylases from cells of Mycobacterium smegmatis butyricum.
<3>Vopr. Med. Khim.
<4>32
<5>125-129
<6>1986
<7>DNA-methylases from M. sm. butyricum, produced by means of isoelectrofocusing, were
characterized by slow spontaneous variations in the activity with amplitude of 75-80% under
conditions of storage in glycerol at -10 C.  Effect of various conditions on stabilization of
activity of adenine methylases M.Mbu4,2 and M.Mbu7,2 was studied.  Alterations in the
enzymatic activity were not found in presence of substances applied usually to stabilize the
enzymes, blood serum albumin and cations of two valent metals as the values of the alterations
occurred in the ranges of the enzyme activity variations.

<>

<1>Sharma, A., Hira, P., Shakarad, M., Lal, R.
<2>Draft Genome Sequence of Cellulosimicrobium sp. Strain MM, Isolated from Arsenic-Rich Microbial Mats of a Himalayan Hot Spring.
<3>Genome Announcements
<4>2
<5>e01020-14
<6>2014
<7>Microbial mats situated at the Manikaran hot springs (>95 degrees C) are characterized by
their high arsenic content (140 ppb), qualifying as a stressed
niche. Here, we report the annotated draft genome (3.85 Mb) of Cellulosimicrobium
sp. strain MM, isolated from these microbial mats, consisting of 3,718 coding
sequences, with an average % G+C of 74.4%.

<>

<1>Sharma, G., Khatri, I., Subramanian, S.
<2>Draft Genome Sequence of Kocuria palustris PEL.
<3>Genome Announcements
<4>2
<5>e01261-13
<6>2014
<7>We report the 2.9-Mb draft genome sequence of an actinobacterium, Kocuria palustris PEL, of
the family Micrococcaceae.

<>

<1>Sharma, M., Ellis, R.L., Hinton, D.M.
<2>Identification of a family of bacteriophage T4 genes encoding proteins similar to those present in group I introns of fungi and phage.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>6658-6662
<6>1992
<7>The bacteriophage T4 segA gene lies in a genetically unmapped region between the gene betagt
(beta-glucosyltransferase) and uvsX (recombination protein) and encodes a protein of 221 amino
acids. We have found that the first 100 amino acids of the SegA protein are highly similar to
the N termini of four other predicted T4 proteins, also of unknown function. Together these
five proteins, SegA-E (similar to endonucleases of group I introns), contain regions of
similarity to the endonuclease I-TevI, which is encoded by the mobile group I intron of the T4
td gene, and to putative endonucleases of group I introns present in the mitochondria of
Neurospora crassa, Podospora anserina, and Saccharomyces douglasii. Intron-encoded
endonucleases are required for the movement (homing) of the intron DNA into an intronless
gene, cutting at or near the site of intron insertion. Our in vitro assays indicate that SegA,
like I-TevI, is a Mg2+-dependent DNA endonuclease that has preferred sites for cutting. Unlike
the I-TevI gene, however, there is no evidence that segA (or the other seg genes) resides
within introns. Thus, it is possible that segA encodes an endonuclease that is involved in the
movement of the endonuclease-encoding DNA rather than in the homing of an intron.

<>

<1>Sharma, M., Hinton, D.M.
<2>Purification and characterization of the SegA protein of bacteriophage T4, an endonuclease related to proteins encoded by group I introns.
<3>J. Bacteriol.
<4>176
<5>6439-6448
<6>1994
<7>Although not encoded by an intron, the bacteriophage T4 SegA protein shares common amino acid
motifs with a family of proteins found within mobile group I introns present in fungi and
phage.  Each of these intron-encoded proteins is thought to initiate the homing of its own
intron by cleaving the intronless DNA at or near the site of insertion.  Previously, we have
found that SegA also cleaves DNA.  In this report, we have purfied the SegA protein and
characterized this endonuclease activity extensively.  SegA protein cleaved circular and
linear plasmids, DNA containing unmodified cytosine, and wild-type T4 DNA containing
hydromethylated, glucosylated cytosines.  In all cases, certain sites on the DNA were highly
preferred for cleavage, but with increasing protein concentration or time of incubation,
cleavage occurred at many sites.  SegA cleaving activity was stimulated by the presence of ATP
or ATPgammaS.  Sequence analysis of three highly preferred cleavage sites did not reveal a
simple consensus sequence, suggesting that even among highly preferred sites, SegA tolerates
many different sequences.  A T4 segA amber mutant that we constructed had no phenotype, and
PCR analyses indicated that several T-even-related phages lack the segA gene.  Taken together,
our results show that SegA is an endonuclease with a hierarchy of site specificity, and these
results are consistent with the insertion of SegA DNA into the T4 genome some time after the
divergence of the closely related T-even phages.

<>

<1>Sharma, P., D'Souza, D.R., Bhandari, D., Parashar, V., Capalash, N.
<2>Demonstration of the principles of restriction endonuclease cleavage reactions using thermostable BflI from Anoxybacillus flavithermus.
<3>Biochem. Mol. Biol. Educ.
<4>31
<5>392-396
<6>2003
<7>Restriction enzymes are basic tools in recombinant DNA technology. To shape the molecular
biology experiments, the students must know how to
work with these molecular scissors. Here, we describe an integrated set
of experiments, introduced in the "Advances in Molecular Biology and
Biotechnology" postgraduate course, which covers the important aspects
about restriction endonucleases: preparation of cell-lysate, one-column
partial-purification, assay, effect of temperature, buffers, Mg2+ ions
and glycerol on activity, and quality control of two-column partially
purified enzyme. Examples of study questions, which help students
understand the theoretical basis of the experiments, have been given.

<>

<1>Sharma, P., Diene, S.M., Gimenez, G., Robert, C., Rolain, J.M.
<2>Genome Sequence of Microbacterium yannicii, a Bacterium Isolated from a Cystic Fibrosis Patient.
<3>J. Bacteriol.
<4>194
<5>4785
<6>2012
<7>Microbacterium yannicii is a Gram-positive, aerobic, yellow-pigmented, rod-shaped, nonmotile,
oxidase-negative, and catalase-positive bacterium isolated
on Columbia colistin-nalidixic acid (CNA) agar with 5% sheep blood from the
sputum of a cystic fibrosis patient. The present study reports the draft genome
of a Microbacterium yannicii strain.

<>

<1>Sharma, P.K., Fu, J., Cicek, N., Sparling, R., Levin, D.B.
<2>Kinetics of medium-chain-length polyhydroxyalkanoate production by a novel isolate of Pseudomonas putida LS46.
<3>Can. J. Microbiol.
<4>58
<5>982-989
<6>2012
<7>Six bacteria that synthesize medium-chain-length polyhydroxyalkanoates (mcl-PHAs)
were isolated from sewage sludge and hog barn wash and identified as strains of
Pseudomonas and Comamonas by 16S rDNA gene sequencing. One isolate, Pseudomonas
putida LS46, showed good PHA production (22% of cell dry mass) in glucose medium,
and it was selected for further studies. While it is closely related to other P.
putida strains (F1, KT2440, BIRD-1, GB-1, S16, and W619), P. putida LS46 was
genetically distinct from these other strains on the basis of nucleotide sequence
analysis of the cpn60 gene hypervariable region. PHA production was detected as
early as 12 h in both nitrogen-limited and nitrogen-excess conditions. The
increase in PHA production after 48 h was higher in nitrogen-limited cultures
than in nitrogen-excess cultures. Pseudomonas putida LS46 produced mcl-PHAs when
cultured with glucose, glycerol, or C(6)-C(14) saturated fatty acids as carbon
sources, and mcl-PHAs accounted for 56% of the cell dry mass when cells were
batch cultured in medium containing 20 mmol/L octanoate. Although
3-hydroxydecanoate was the major mcl-PHA monomer (58.1-68.8 mol%) in P. putida
LS46 cultured in glucose medium, 3-hydroxyoctanoate was the major monomer
produced in octanoate medium (88 mol%).

<>

<1>Sharma, P.K., Fu, J., Zhang, X., Fristensky, B.W., Davenport, K., Chain, P.S., Sparling, R., Levin, D.B.
<2>Draft Genome Sequence of Medium-Chain-Length Polyhydroxyalkanoate-Producing Pseudomonas putida Strain LS46.
<3>Genome Announcements
<4>1
<5>e00151-13
<6>2013
<7>We describe the draft genome sequence of Pseudomonas putida strain LS46, a novel  isolate that
synthesizes medium-chain-length polyhydroxyalkanoates. The draft
genome of P. putida LS46 consists of approximately 5.86 million bp, with a G+C
content of 61.69%. A total of 5,316 annotated genes and 5,219 coding sequences
(CDS) were identified.

<>

<1>Sharma, R., Singh, R.K.M., Malik, G., Deveshwar, P., Tyagi, A.K., Kapoor, S., Kapoor, M.
<2>Rice cytosine DNA methyltransferases - gene expression profiling during reproductive development and abiotic stress.
<3>FEBS J.
<4>276
<5>6301-6311
<6>2009
<7>DNA methylation affects important developmental processes in both plants and animals. The
process of methylation of cytosines at C-5 is
catalysed by DNA methyltransferases (MTases), which are highly
conserved, both structurally and functionally, in eukaryotes. In this
study, we identified and characterized cytosine DNA MTase genes that
are activated with the onset of reproductive development in rice. The
rice genome (Oryza sativa L. subsp. japonica) encodes a total of 10
genes that contain the highly conserved MTase catalytic domain. These
genes have been categorized into subfamilies on the basis of
phylogenetic relationships. A microarray-based gene expression profile
of all 10 MTases during 22 stages/tissues that included 14 stages of
reproductive development and five vegetative tissues together with
three stresses, cold, salt and dehydration stress, revealed specific
windows of MTase activity during panicle and seed development. The
expression of six methylases was specifically/preferentially
upregulated with the initiation of floral organs. Significantly, one of
the MTases was also activated in young seedlings in response to cold
and salt stress. The molecular studies presented here suggest a greater
role for these proteins and the epigenetic process in affecting genome
activity during reproductive development and stress than was previously
anticipated.

<>

<1>Sharma, V., Midha, S., Ranjan, M., Pinnaka, A.K., Patil, P.B.
<2>Genome Sequence of Xanthomonas axonopodis pv. punicae Strain LMG 859.
<3>J. Bacteriol.
<4>194
<5>2395
<6>2012
<7>We report the 4.94-Mb genome sequence of Xanthomonas axonopodis pv. punicae strain LMG 859,
the causal agent of bacterial leaf blight disease in pomegranate.
The draft genome will aid in comparative genomics, epidemiological studies, and
quarantine of this devastating phytopathogen.

<>

<1>Sharma, V., Singh, P.K., Midha, S., Ranjan, M., Korpole, S., Patil, P.B.
<2>Genome Sequence of Brevibacillus laterosporus Strain GI-9.
<3>J. Bacteriol.
<4>194
<5>1279
<6>2012
<7>We report the 5.18-Mb genome sequence of Brevibacillus laterosporus strain GI-9,  isolated
from a subsurface soil sample during a screen for novel strains
producing antimicrobial compounds. The draft genome of this strain will aid in
biotechnological exploitation and comparative genomics of Brevibacillus
laterosporus strains.

<>

<1>Sharma, V., Youngblood, B., Reich, N.
<2>Residues distal from the active site that alter enzyme function in M.HhaI DNA cytosine methyltransferase.
<3>J. Biomol. Struct. Dyn.
<4>22
<5>533-543
<6>2005
<7>Ten M.HhaI residues were replaced with alanine to probe the importance of distal protein
elements to substrate/cofactor binding, methyl
transfer, and product release. The substitutions, ranging from 6-20 A
from the active site were evaluated by thermodynamic analysis, AdoMet
DNA pre-steady and steady-state kinetics, to obtain K-d(AdoMet),
K-d(DNA), k(cat)/K-m(DNA), k(cat), and k(methyltransfer) values. For
the wild-type M.Hhal, product release steps dominate catalytic turnover
while the 4-fold faster internal microscopic constant kmethyltransfer
presents an upper limit. The methyl transfer reaction has
Delta H and Delta S values of 10.3
kcal/mol and 29.4 cal/(mol K), respectively, consistent with a
compressed transition state similar to that observed in the gas phase.
Although the ten mutants remained largely unperturbed in methyl
transfer, long-range effects influencing substrate/cofactor binding and
product release were observed. Positive enhancements were seen in
Asp(73)Ala, which showed a 25-fold improvement in AdoMet affinity and
in Val(282)Ala, which showed a 4-fold improvement in catalytic
turnover. Based on an analysis of the positional probability within the
C-5-cytosine DNA methyltransferase family we propose that certain
conserved distal residues may be important in mediating long-range
effects.

<>

<1>Sharp, P.A., Sugden, B., Sambrook, J.
<2>Detection of two restriction endonuclease activities in Haemophilus parainfluenzae using analytical agarose-ethidium bromide electrophoresis.
<3>Biochemistry
<4>12
<5>3055-3063
<6>1973
<7>A rapid assay for restriction enzymes has been developed using electrophoresis
of DNA through 1.4% agarose gels in the presence of 0.5 microgram/ml of
ethidium bromide.  The method eliminates lengthy staining and destaining
procedures and resolves species of DNA which are less than 7 Mdaltons.  As
little as 0.05 microgram of DNA can easily be detected by direct examination of
the gels in ultraviolet light.  Using this technique, we have identified two
different restricting activities in extracts of Haemophilus parainfluenzae.
The two activities have different chromatographic properties on
phosphocellulose and Bio-Gel A-0.5m, and they attack SV40 DNA at different
sites.  One activity (HpaII) cleaves SV40 DNA at a single position situated
0.38 fractional genome length from the insertion point of SV40 sequences into
the adenovirus SV40 hybrid Ad2++ND1.  The other activity (HpaI) cleaves SV40
DNA at three sites which appear to coincide with 3 of the 11 cleavage points
attacked by a restriction system isolated from H. influenzae strain Rd.

<>

<1>Sharp, P.M.
<2>Molecular evolution of bacteriophages: Evidence of selection against the recognition sites of host restriction enzymes.
<3>Mol. Biol. Evol.
<4>3
<5>75-83
<6>1986
<7>Restriction enzymes produced by bacteria serve as a defense against invading
bacteriophages, and so phages without other protection would be expected to
undergo selection to eliminate recognition sites for these enzymes from their
genomes.  The observed frequencies of all restriction sites in the genomes of
all completely sequenced DNA phages (T7, lambda, PhiX174, G4, M13, fl, fd, and
IKe) have been compared to expected frequencies derived from trinucleotide
frequencies.  Attention was focused on 6-base palindromes since they comprise
the typical recognition sites for type II restriction enzymes.  All of these
coliphages, with the exception of lambda and G4, exhibit significant avoidance
of the particular sequences that are enterobacterial restriction sites.  As
expected, the sequenced fraction of the genome of Phi29, a Bacillus subtilis
phage, lacks Bacillus restriction sites.  By contrast, the RNA phage MS2,
several viruses that infect eukaryotes (EBV, adenovirus, papilloma, and SV40),
and three mitochondrial genomes (human, mouse and cow) were found not to lack
restriction sites.  Because the particular palindromes avoided correspond
closely with the recognition sites for host enzymes and because other viruses
and small genomes do not show this avoidance, it is concluded that the effect
indeed results from natural selection.

<>

<1>Sharp, P.M., Kelleher, J.E., Daniel, A.S., Cowan, G.M., Murray, N.E.
<2>Roles of selection and recombination in the evolution of type I restriction-modification systems in enterobacteria.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>9836-9840
<6>1992
<7>Restriction-modification systems can protect bacteria against viral infection. Sequences of
the hsdM gene, encoding one of the three subunits of type I restriction-modification systems,
have been determined for four strains of enterobacteria. Comparison with the known sequences
of EcoKI and EcoR124I indicated that all are homologous, though they fall into three families
(exemplified by EcoKI, EcoAI, and EcoR124I), the first two of which are apparently allelic.
The extent of amino acid sequence identity between EcoKI and EcoAI is so low that the genes
encoding them might be better termed pseudoalleles; this almost cerainly reflects genetic
exchange among highly divergent species. Within the EcoKI family the ratio of intra- to
interspecific divergence is very high. The extent of divergence between the genes from
Escherichia coli K-12 and Salmonella typhimurium LT2 is similar to that for other genes with
the same level of codon usage bias. In contrast, intraspecific divergence (between E. coli
strains B and K-12) is extremely high and may reflect the action of frequency-dependent
selection mediated by bacteriophages. There is also evidence of lateral transfer of a short
sequence between E. coli and S. typhimurium.

<>

<1>Sharrocks, A.D., Hornby, D.P.
<2>Transcriptional analysis of the restriction and modification genes of bacteriophage P1.
<3>Mol. Microbiol.
<4>5
<5>685-694
<6>1991
<7>Bacteriophage P1 res and mod genes encode the restriction and modification
polypeptides of the Type III restriction enzyme EcoPI.  Northern blot analysis
using res- and mod-specific probes revealed the presence of two separate
transcripts in strains harbouring the EcoPI restriction and modification genes.
Furthermore, by constructing a series of fusions with a promoterless lacZ
gene, we show that both the res and mod genes are transcribed from separate
promoters.  A more detailed investigation of the mod promoter region revealed
two promoters located some 70 and 140 bp upstream from the translational start
codon.  In addition, another pair of promoters and a further separate promoter
are located more than 500 bp upstream from this start codon.  Two short open
reading frames are located between these distal and proximal promoter clusters.
Transcription of the res gene is initiated from within the mod open reading
frame from two adjacent promoters.  In addition a functional promoter is
located on the antisense strand close to the res promoter region.  The
relationship between the transcription units of the res and mod genes is
discussed.

<>

<1>Shaw, P.C., Clark, D.R., Kam, K.M., Mok, Y.K.
<2>Isolation and characterization of thermophilic restriction endonucleases.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>199
<6>1991
<7>Restriction endonucleases BsiBI, BsiEI, BsiGI, BsiUI and BsiYI were isolated from thermophilic
Bacillus spp. BsiBI, BsiEI and BsiUI were purified by using a DEAE sephacel column and a
heparin sepharose column, while BsiYI was followed by a FPLC mono Q column. Unpurified BsiGI
was used. Recognition sites of these enzymes were found by means of single and double
digestions on different DNA of known sequences. The cleavage sites were determined according
to FEMS Microbiol. Lett. 66:153-156 (1990). BsiGI and BsiUI were found to recognize TCCGGA and
CCWGG respectively. The recognition and cleavage sites of the other three enzymes are: BsiBI:
GATNN^NNATC; BsiEI: CGRY^CG and BsiYI: CCNNNNN^NNATC. These enzymes worked well at 55C. BsiBI
and BsiUI were sensitive to dam and dcm methylation respectively. The optimal ionic strength
for BsiEI and BsiYI was medium salt buffer, while that for BsiBI, BsiGI and BsiUI was high
salt buffer. During the characterization of BsiYI, we found that a G nucleotide is missed at
position 1227 of pACYC177 recorded in GenBank. This error also occurs in pACYC184 and p15A.

<>

<1>Shaw, P.C., Mok, Y.K.
<2>XcmI as a universal restriction enzyme for single-stranded DNA.
<3>Gene
<4>133
<5>85-89
<6>1993
<7>Single-stranded DNA can be cleaved into defined fragments at any predetermined site by
interaction with a specially designed oligodeoxyribonucleotide (oligo) adaptor and the
class-IIN restriction endonuclease, XcmI. The oligo adaptor has the structure
(CCANNNNNNNNNTGG
 GGTNNNNNNNNNACC). Upon hybridization to the target DNA through the central 9-nucleotide
region and with the addition of XcmI, the template DNA is specifically cleaved to near
completion. Hairpin structures on the template close to the hybridization site reduce the
efficacy of cleavage.

<>

<1>Shaw, Y.J., Chen, C.S.
<2>Structure-activity relationship study of (-)-epicatechin analogues as DNA methyltransferase inhibitors.
<3>ACS Abstracts
<4>229
<5>U186-U187
<6>2005
<7>Hypermethylation of DNA cPG islands is an important epigenetic mechanism for aberrant gene
silencing in cancer.  Although nucleoside analogue inhibitors of DNA methyltransferases can
reverse abnormal DNA hypermethylation in cancer cells, their clinical utility is limited due
to side effects.  The development of nonnucleoside DNMTs that lack systemic toxicity is a
promising approach to targeting epigenetic mechanisms for cancer therapy.  (-)-Epicatechin
analogues extracted from green tea inhibit DNMT.  Among them, EGCG and ECG exhibit inhibitory
activity with IC50 values of 20-30 uM.  Our objective is to synthesize (-)-epicatechin
scaffold, a series of derivaties have been synthesized and screened for activity in an in
vitro recombinant DNMT assay system.  Lead compounds with potent inhibitory activity will
subsequently be evaluated in cancer cell-based models of viability, apoptosis and DNA
methylation status.

<>

<1>She, Q. et al.
<2>The complete genome of the crenarchaeon Sulfolobus solfataricus P2.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>7835-7840
<6>2001
<7>The genome of the crenarchaeon Sulfolobus solfataricus P2 contains 2,992,245 bp on a single
chromosome and encodes 2,977 proteins and many RNAs. One-third of the encoded proteins have no
detectable homologs in other sequenced genomes. Moreover, 40% appear to be archaeal-specific,
and only 12% and 2.3% are shared exclusively with bacteria and eukarya, respectively. The
genome shows a high level of plasticity with 200 diverse insertion sequence elements, many
putative nonautonomous mobile elements, and evidence of integrase-mediated insertion events.
There are also long clusters of regularly spaced tandem repeats. Different transfer systems
are used for the uptake of inorganic and organic solutes, and a wealth of intracellular and
extracellular proteases, sugar, and sulfur metabolizing enzymes are encoded, as well as
enzymes of the central metabolic pathways and motility proteins. The major metabolic electron
carrier is not NADH as in bacteria and eukarya but probably ferredoxin. The essential
components required for DNA replication, DNA repair and recombination, the cell cycle,
transcriptional initiation and translation, but not DNA folding, show a strong eukaryal
character with many archaeal-specific features. The results illustrate major differences
between crenarchaea and euryarchaea, especially for their DNA replication mechanism and cell
cycle processes and their translational apparatus.

<>

<1>Shearman, C., Godon, J.-J., Gasson, M.
<2>Splicing of a group II intron in a functional transfer gene of Lactococcus lactis.
<3>Mol. Microbiol.
<4>21
<5>45-53
<6>1996
<7>A chromosomally located sex factor that controls conjugation in Lactococcus lactis 712 has
been cloned and sequenced, leading to the discovery of an open reading frame with homology to
the maturases of group II self-splicing introns.  Reverse transcriptase polymerase chain
reaction amplification was used to demonstrate that the intron was spliced out of mRNA in
vivo, and sequence analysis revealed the site of splicing.  The intron was inserted within a
sex-factor gene which encodes a protein with homology to proteins involved in rolling-circle
DNA replication.  Gene-disruption experiments were used to demonstrate that this mobA gene was
essential for sex-factor transfer and this suggests that intron splicing is a necessary part
of the conjugation process.  The sequence of the intron was modelled to produce a secondary
structure that exhibited several features characteristic of the IIA subgroup.  Here we report
the characterization of a new group II intron in the Gram-positive bacterium L. lactis and
demonstrate for the first time in bacteria both splicing in vivo and an active role for the
gene carrying the intron.

<>

<1>Sheflyan, G.Y., Kubareva, E.A., Gromova, E.S., Shabarova, Z.A.
<2>Hydrolysis of 5-fluorodeoxycytidine-containing DNA duplexes using restriction endonucleases.
<3>Biokhimiia
<4>58
<5>1806-1811
<6>1993
<7>Cleavage by restriction endonucleases MvaI and EcoRII of DNA duplexes, in which the internal
deoxycytidine in one of the strands of the recognition site is substituted for
5-fluorodeoxycytidine, has been studied. It has been found that the modified strands of these
duplexes are cleaved by endonuclease MvaI with a greater efficiency than the intact ones.
Endonuclease EcoRII does not practically cleave such substrate analogs. The UV absorbance
spectra of 5-fluorodeoxycytidine and the 14-member oligodeoxyribonucleotide with modification
in the middle of the strand have been measured. A hypothesis has been put forward about the
structural features of 5-fluorodeoxycytidine-containing DNA duplexes.

<>

<1>Sheflyan, G.Y., Kubareva, E.A., Gromova, E.S., Shabarova, Z.A.
<2>Conformational transition of restriction endonuclease MvaI-substrate complex under the influence of Mg2+ probed by DNA-protein linking studies.
<3>Bioconjugate Chem.
<4>9
<5>703-707
<6>1998
<7>The method of protein affinity modification by DNA analogues was used to study the
characteristic features of restriction endonuclease MvaI interaction with DNA.
Oligonucleotide duplexes containing a monosubstituted pyrophosphate internucleotide bond were
used for cross-linking to the enzyme.  The conditions of the reaction of MvaI endonuclease
with these reagents were investigated.  On the basis of data obtained, the model of successive
inclusion of two Mg2+ ions into an MvaI endonuclease-substrate complex was proposed and
confirmed by the kinetic scheme of the process.

<>

<1>Sheflyan, G.Y., Kubareva, E.A., Kuznetsova, S.A., Karyagina, A.S., Nikolskaya, I.I., Gromova, E.S., Shabarova, Z.A.
<2>Cross-linking of SsoII restriction endonuclease to cognate and non-cognate DNAs.
<3>FEBS Lett.
<4>390
<5>307-310
<6>1996
<7>Specific and non-specific interactions of SsoII restriction endonuclease (R.SsoII) were probed
by the method of covalent attachment to modified DNA containing an active monosubstituted
pyrophosphate internucleotide bond instead of a phosphodiester one.  R.SsoII with six
N-terminal His residues was shown to be cross-linked to duplexes with this type of
modification, either containing or not the recognition sequence.  Competition experiments with
covalent attachment of R.SsoII to activated DNAs demonstrated the similar affinity of the
enzyme to cognate and noncognate DNAs in the absence of cofactor, Mg2+ ions.

<>

<1>Sheflyan, G.Y., Kubareva, E.A., Volkov, E.M., Oretskaya, T.S., Gromova, E.S., Shabarova, Z.A.
<2>Chemical cross-linking of MvaI and EcoRII enzymes to DNA duplexes containing monosubstituted pyrophosphate internucleotide bond.
<3>Gene
<4>157
<5>187-190
<6>1995
<7>DNA duplexes containing a monosubstituted pyrophosphate internucleotide group, instead of a
phosphodiester bond, were used as cross-linking reagents for the affinity modification of the
restriction endonucleases EcoRII and MvaI (R.EcoRII and R.MvaI).  An active group was
introduced into the enzyme's recognition site or between the recognition site and flanking
sequence.  The substrate properties of such DNA duplexes were determined.  Cross-linking
specificity was demonstrated by competition experiments with unmodified substrate, as well as
by the absence of cross-linking to an active duplex lacking a recognition site.  It was shown
that the nucleophilicity of the buffer solution and the presence of the enzyme cofactor Mg2+
dramatically affected the cross-linking yield.

<>

<1>Sheflyan, G.Y., Tashlitskii, V.N., Kubareva, E.A.
<2>Interaction of restriction endonuclease MvaI with synthetic DNA duplexes.
<3>Vestn. Mosk. Univ.
<4>34
<5>516-520
<6>1993
<7>
<>

<1>Sheh, A., Piazuelo, M.B., Wilson, K.T., Correa, P., Fox, J.G.
<2>Draft Genome Sequences of Helicobacter pylori Strains Isolated from Regions of Low and High Gastric Cancer Risk in Colombia.
<3>Genome Announcements
<4>1
<5>e00736-13
<6>2013
<7>The draft genome sequences of six Colombian Helicobacter pylori strains are presented. These
strains were isolated from patients from regions of high and low
gastric cancer risk in Colombia and were characterized by multilocus sequence
typing. The data provide insights into differences between H. pylori strains of
different phylogeographic origins.

<>

<1>Sheh, A., Shen, Z., Fox, J.G.
<2>Draft Genome Sequences of Eight Enterohepatic Helicobacter Species Isolated from  Both Laboratory and Wild Rodents.
<3>Genome Announcements
<4>2
<5>e01218-14
<6>2014
<7>The draft genome sequences of eight enterohepatic Helicobacter species, H. muridarum, H.
trogontum, H. typhlonius, and five unnamed helicobacters, are
presented here. Using laboratory mice pervasively infected with helicobacters, we
characterized the presence of known virulence factors.

<>

<1>Sheikhnejad, G., Brank, A., Christman, J.K., Goddard, A., Alvarez, E., Ford, H. Jr., Marquez, V.E., Marasco, C.J., Sufrin, J.R., O'Gara, M., Cheng, X.
<2>Mechanism of inhibition of DNA (cytosine C5)-methyltransferases by oligodeoxyribonucleotides containing 5,6-dihydro-5-azacytosine.
<3>J. Mol. Biol.
<4>285
<5>2021-2034
<6>1999
<7>A key step in the predicted mechanism of enzymatic transfer of methyl groups from
S-adenosyl-L-methionine to cytosine residues in DNA is the transient formation of a
dihydrocytosine intermediate covalently linked to cysteine in the active site of a DNA
(cytosine C5)-methyltransferase.  Crystallographic analysis of complexes formed by HhaI
methyltransferase, AdoMet and a target oligodeoxyribonucleotide containing 5-fluorocytosine
confirmed the existence of this dihydrocytosine intermediate.  Based on the premise that
5,6-dihydro-5-azacytosine, a cytosine analog with an sp3-hybridized carbon (CH2) at position 6
and an NH group at position 5, could mimic the non-aromatic character of the cytosine ring in
this transition state, we synthesized a series of synthetic substrates for DNA C5-MTase
containing DZCyt.  Substitution of DZCyt for target cytosines in C-G dinucleotides of
single-stranded or double-stranded oligodeoxyribonucleotide substrates led to complete
inhibition of methylation by murine DNA C5-MTase.  Substitution of DZCyt for the target
cytosine in G-C-G-C sites in double-stranded oligodeoxyribonucleotides had a similar effect on
methylation by M.HhaI.  Oligodeoxyribonucleotides containing DZCyt formed a tight but
reversible complex with M.HhaI, and were consistently more potent as inhibitors of DNA
methylation than oligodeoxyribonucleotides identical in sequence containing 5-fluorocytosine.
Crystallographic analysis of a ternary complex involving M.HhaI, S-adenosyl-L-homocysteine and
a double-stranded 13-mer oligodeoxyribonucleotide containing DZCyt at the target position
showed that the analog is flipped out of the DNA helix in the same manner as cytosine,
5-methylcytosine, and 5-fluorocytosine.  However, no formation of a covalent bond was detected
between the sulfur atom of the catalytic site nucleophile, cysteine 81, and the pyrimidine C6
carbon.  These results indicate that DZCyt can occupy the active site of M.HhaI as a
transition state mimic and, because of the high degree of affinity of its interaction with the
enzyme, it can act as a potent inhibitor of methylation.

<>

<1>Sheikhnejad, G., Marasco, C., Sufrin, J., Christman, J.K.
<2>Comparison of substrate specificity of mammalian and bacterial CpG DNA methyltransferases (MTases).
<3>Biomed. Pharmacother.
<4>46
<5>24
<6>1992
<7>Methylation of C residues at specific sites in DNA of higher eukaryotes appears to play a role
in regulating gene expression during development and diferentiation. Silencing genes by
methylation can in part explain genomic imprinting and alterations in patterns of methylation
during the early stages of carcinogenesis may account for some changes in gene expression in
tumor cells. Although the distribution of 5mC in mammalian DNA is tissue- and
species-specific, little is known about MTases role in establishing specific patterns of
methylation. Mammalian DNA (mDNA) MTase is most active in methylating C residues in
hemi-methylated CpG sites and prefers substrates with high GC content and multiple CpGs approx
15 residues apart.  To more fully characterize the specificity of mDNA MTase, we analyzed
its activity with a variety of defined DNA substrates and compared it with a bacterial MTase,
SssI, which is reported to act on hemi-methylated CpG sites with equal efficency. Using
defined unmethylated single (ss)- and double stranded (ds)-DNAs as substrates, we find that:1)
the initial rate of methylation of both ss-and ds-DNA-by SssI and mDNA MTase is influenced by
sequence, often in opposite ways; 2) depending on sequence, SssI may be more active on ds-
than ss-DNA. Activity of mDNA MTase with ss-DNA is highly variable and dependent on sequence
but is always low with unmethylated ds-DNA. Using 5mC-substituted substrates, we find that
hemi-methylation either has no effect or inhibits activity of SssI but always activates mDNA
MTase. mDNA MTase is most active with hemi-methylated ds-DNA ans ss-DNA with 5mC near the
5'-end. These results demonstrate that although both MTases catalyze the same reaction,
methylation of C residues in CpG dinucleotides, they have quite different specificities. This
should be considered when these enzymes are used to quantitate hypomethylation at CpG sites in
DNA resulting from exposure of mammalian cells or tissues to conditions or agents that affect
DNA methylation. Finally, our findings suggest that 5mC residues in specific sequences in
ss-DNA may serve to activate de novo methylation in mammalian cells allowing alteration of
existing methylation patterns.

<>

<1>Shell, S.S., Prestwich, E.G., Baek, S.H., Shah, R.R., Sassetti, C.M., Dedon, P.C., Fortune, S.M.
<2>DNA methylation impacts gene expression and ensures hypoxic survival of Mycobacterium tuberculosis.
<3>PLoS Pathog.
<4>9
<5>e1003419
<6>2013
<7>DNA methylation regulates gene expression in many organisms. In eukaryotes, DNA methylation is
associated with gene repression, while it exerts both activating
and repressive effects in the Proteobacteria through largely locus-specific
mechanisms. Here, we identify a critical DNA methyltransferase in M.
tuberculosis, which we term MamA. MamA creates N(6)-methyladenine in a six base
pair recognition sequence present in approximately 2,000 copies on each strand of
the genome. Loss of MamA reduces the expression of a number of genes. Each has a
MamA site located at a conserved position relative to the sigma factor -10
binding site and transcriptional start site, suggesting that MamA modulates their
expression through a shared, not locus-specific, mechanism. While strains lacking
MamA grow normally in vitro, they are attenuated in hypoxic conditions,
suggesting that methylation promotes survival in discrete host microenvironments.
Interestingly, we demonstrate strikingly different patterns of DNA
methyltransferase activity in different lineages of M. tuberculosis, which have
been associated with preferences for distinct host environments and different
disease courses in humans. Thus, MamA is the major functional adenine
methyltransferase in M. tuberculosis strains of the Euro-American lineage while
strains of the Beijing lineage harbor a point mutation that largely inactivates
MamA but possess a second functional DNA methyltransferase. Our results indicate
that MamA influences gene expression in M. tuberculosis and plays an important
but strain-specific role in fitness during hypoxia.

<>

<1>Sheluho, D., Khariwala, S., Yebra, M.J., Bhagwat, A.S.
<2>Ability of cytosine methyltransferases to bind substrates containing mismatches is not sufficient for causing C to T mutations.
<3>FASEB J.
<4>10
<5>A965
<6>1996
<7>Hydrolytic deamination of cytosine to uracil and of 5-methylcytosine to thymine create
mismatches, which - if unrepaired - can cause C to T mutations.  It was found that cytosine
methyltransferases (C5 Mtases) M.HhaI and M.HpaII bind tightly to these mismatches suggesting
that the C5 Mtases may prevent repair of U:G and T:G mismatches by competing with mismatch
correction systems.  To test this, we studied the binding of WT M.EcoRII and its mutant in
which active site cysteine was replaced with serine (C186S) to oligonucleotide duplexes
containing U:G or T:G mismatches.  We found that C186S mutant binds more tightly to duplexes
containing mismatches than the wild type enzyme.  The tightness of binding to the mismatched
duplexes was generally comparable to that with the cognate sequence.  In contrast, when the
ability of the C186S mutant to promote C to T mutations was tested using a genetic reversion
system, we found that the magnitude of reversion caused by the WT M.EcoRII was about 50 fold
higher than that caused by the C186S mutant.  This shows that the ability of C5 Mtases to bind
U:G or T:G mismatch containing duplexes is not sufficient to promote C to T mutations.

<>

<1>Sheluho, D., Yebra, M.J., Shariwala, S.S., Bhagwat, A.S.
<2>Lack of correlation between binding of EcoRII methyltransferase to DNA duplexes containing mismatches and the promotion of C to T mutations.
<3>Mol. Gen. Genet.
<4>255
<5>54-59
<6>1997
<7>The cytosine methyltransferases (MTases) M.HhaI and M.HpaII bind substrates in which the
target cytosine is replaced by uracil or thymine, i.e. DNA containing a U:G or a T:G mismatch.
We have extended this observation to the EcoRII MTase (M.EcoRII) and determined the apparent
Kd for binding.  Using a genetic assay we have also tested the possibility that MTase binding
to U:G mismatches may interfere with repair of the mismatches and promote C:G to T:A
mutations.  We have compared two mutants of M.EcoRII that are defective for catalysis by the
wild-type enzyme for their ability to bind DNA containing U:G or T:G mismatches and for their
ability to promote C to T mutations.  We find that although all three proteins are able to
bind DNAs with mismatches, only the wild-type enzyme promotes C:G to T:A mutations in vivo.
Therefore, the ability of M.EcoRII to bind U:G mismatched duplexes is not sufficient for their
mutagenic action in cells.

<>

<1>Shemer, R., Birger, Y., Riggs, A.D., Razin, A.
<2>Structure of the imprinted mouse Snrpn gene and establishment of its parental-specific methylation pattern.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>94
<5>10267-10272
<6>1997
<7>The mouse Snrpn gene encodes the Smn protein, which is involved in RNA splicing.  The gene
maps to a region in the central part of chromosome 7 that is syntenic to the
Prader-Willi/Angelman syndromes region on human chromosome 15q11-q13.  The mouse gene, like
its human counterpart, is imprinted and paternally expressed, primarily in brain and heart.
We provide here a detailed description of the structural features and differential methylation
pattern of the gene.  We have identified a maternally methylated region at the 5' end (DMR1),
which correlates inversely with the Snrpn paternal expression.  We also describe a region at
the 3' end of the gene (DMR1), which correlates inversely with the Snrpn paternal expression.
We also describe a region at the 3' end of the gene (DMR2) that is preferentially methylated
on the paternal allele.  Analysis of Snrpn mRNA levels in a methylase-deficient mouse embryo
revealed that maternal methylation of DMR1 may play a role in silencing the maternal allele.
Yet both regions, DMR1 and DMR2, inherit the parental-specific methylation profile from the
gametes.  This methylation pattern is erased in 12.5 days postcoitum primordial germ cells and
reestablished during gametogenesis.  DMR1 is remethylated during oogenesis, whereas DMR2 is
remethylated during spermatogenesis.  Once established, these methylation patterns are
transmitted to the embryo and maintained, protected from methylation changes during
embryogenesis and cell differentiation.  Transfections of DMR1 and DMR2 into embryonic stem
cells and injection into pronuclei of fertilized eggs reveal that embryonic cells lack the
capacity to establish anew the differential methylation pattern of Snrpn.  That all PWS
patients lack DMR1, together with the overall high resemblance of the mouse gene to the human
SNRPN, offers an excellent experimental tool to study the regional control of this imprinted
chromosomal domain.

<>

<1>Shemer, R., Razin, A.
<2>Establishment of imprinted methylation patterns during development.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, Russo, V.E.A., Martienssen, R.A., Riggs, A.D., Cold Spring Harbor
<4>0
<5>215-229
<6>1996
<7>Genomic imprinting is a classic example of epigenetic control of gene expression in mammals.
It represents a process that marks the parental origin of certain genes, resulting in their
allele-specific expression.  Imprinting, which is implicated in the inequality of the maternal
and paternal genomes in mammals, results in the developmental failure of isoparental embryos
to develop properly.  An imbalance of specific parental chromosomes in the embryo or aberrant
expression of the imprinted genes results in a number of genetic disorders in man and may also
contribute to tumor development.  Several model explanations have been proposed for the
evolutionary acquisition of genomic imprinting as a developmental control mechanism in
mammals.  One of these models suggests that imprinting might have evolved in mammals because
of the conflicting "interests" of maternal and paternal genes within the embryo.  This model
proposes that paternally expressed genes promote embryonic growth, whereas maternal genes act
to restrain the use of maternal resources.  A suitable example that corroborates this argument
is the paternally expressed Igf2 gene and its counterpart, maternally expressed receptor
(Igf2r).  The different contribution to the embryonic phenotype of paternal and maternal genes
had in fact been demonstrated in parthenogenetic and androgenetic embryos.  Although the
development of extraembryonic tissues in parthenogenetic embryos harboring two copies of the
maternal genome is impaired, the development of the embryo proper is normal.  In contrast, the
embryo proper in androgenones carrying two copies of the paternal genome does not develop
properly whereas extraembryonic tissues develop normally.

<>

<1>Shemesh, M., Gurevich, M., Stram, Y., Benvenisti, L., Shore, L.
<2>High efficiency methods and compositions for integrating exogenous DNA into genomic DNA of sperm.
<3>International Patent Office
<4>WO 9942569
<5>
<6>1999
<7>The present invention relates to high efficiency methods and compositions for stably
integrating exogenous double-stranded DNA into the chromosomal DNA of sperm cells.  The
exogenous DNA to be integrated into the sperm is converted to a linear double stranded DNA
possessing single-stranded cohesive ends, by contacting said exogenous DNA with a type II
restriction enzyme that upon scission, generates such ends.  The DNA to be cut can be a
circular DNA such as in a plasmid, such that it possesses at least one recognition and cutting
site outside of the genes or regulatory regions critical to the desired, post-integration
function of the DNA, and no recognition and cutting sites within the critical regions.  The
exogenous DNA is then introduced into sperm cells, as is some of the type II restriction
enzyme used above to generate the single-stranded cohesive ends on the exogenous DNA.  The DNA
and the corresponding restriction enzyme can be introduced into the sperm cells, together or
sequentially, by electroporation or lipofection or by any other technique that preserves
enough of said sperm's mobility and fertilization functions.  The sperm containing the
integrated exogenous DNA can then be used to fertilize eggs by artificial insemination, in
vitro fertilization, or other fertilization methods known in the art, thereby producing
transgenic animals wherein all or a plurality of cells now contain the exogenous DNA stably
integrated into their respective genomes.  The present invention also relates to transgenic
animals or their descendants produced by methods or compositions herein disclosed.  The
present invention further relates to cells or cell lines derived from transgenic animals or
their descendants produced by methods or compositions herein disclosed.  The present invention
further relates to proteins or other gene products encoded for by the exogenously introduced
DNA isolated from said transgenic animals or their descendants.  The present invention further
relates to using the method of the present invention to produce transgenic animals resistant
to diseases, and to the disease-resistant animals thereby produced.

<>

<1>Shemesh, M., Pasvolsky, R., Sela, N., Green, S.J., Zakin, V.
<2>Draft Genome Sequence of Alicyclobacillus acidoterrestris Strain ATCC 49025.
<3>Genome Announcements
<4>1
<5>e00638-13
<6>2013
<7>Alicyclobacillus acidoterrestris is a spore-forming Gram-positive, thermo-acidophilic,
nonpathogenic bacterium which contaminates commercial
pasteurized fruit juices. The draft genome sequence for A. acidoterrestris strain
ATCC 49025 is reported here, providing genetic data relevant to the successful
adaptation and survival of this strain in its ecological niche.

<>

<1>Shen, B.W., Heiter, D.F., Chan, S.H., Wang, H., Xu, S.Y., Morgan, R.D., Wilson, G.G., Stoddard, B.L.
<2>Unusual target site disruption by the rare-cutting HNH restriction endonuclease PacI.
<3>Structure
<4>18
<5>734-743
<6>2010
<7>The crystal structure of the rare-cutting HNH restriction endonuclease PacI in complex with
its eightbase-pair target recognition sequence 50-TTAATT AA-30 has been determined to 1.9 A°
resolution. The enzyme forms an extended homodimer, with each subunit containing two
zinc-bound motifs surrounding a bba-metal catalytic site. The latter is unusual in that a
tyrosine residue likely initiates strand cleavage. PacI dramatically distorts its target
sequence from Watson-Crick duplex DNA base pairing, with every base separated from its
original partner. Two bases on each strand are unpaired, four are engaged in noncanonical A:A
and T:T base pairs, and the remaining two bases are matched with new Watson-Crick partners.
This represents a highly unusual DNA binding mechanism for a restriction endonuclease, and
implies that initial recognition of the target site might involve significantly different
contacts from those visualized in the DNA-bound cocrystal structures.

<>

<1>Shen, B.W., Landthaler, M., Shub, D.A., Stoddard, B.L.
<2>DNA binding and cleavage by the HNH homing endonuclease I-HmuI.
<3>J. Mol. Biol.
<4>342
<5>43-56
<6>2004
<7>The structure of I-HmuI, which represents the last family of homing endonucleases without a
defining crystallographic structure, has been
determined in complex with its DNA target. A series of diverse protein
structural domains and motifs, contacting sequential stretches of
nucleotide bases, are distributed along the DNA target. I-HmuI contains
an N-terminal domain with a DNA-binding surface found in the I-PpoI
homing endonuclease and an associated HNH/N active site found in the
bacterial colicins, and a C-terminal DNA-binding domain previously
observed in the I-TevI homing endonuclease. The combination and
exchange of these features between protein families indicates that the
genetic mobility associated with homing endonucleases extends to the
level of independent structural domains. I-HmuI provides an unambiguous
structural connection between the His-Cys box endonucleases and the
bacterial colicins, supporting the hypothesis that these enzymes
diverged from a common ancestral nuclease.

<>

<1>Shen, B.W., Xu, D., Chan, S.H., Zheng, Y., Zhu, Z., Xu, S.Y., Stoddard, B.L.
<2>Characterization and crystal structure of the type IIG restriction endonuclease RM.BpuSI.
<3>Nucleic Acids Res.
<4>39
<5>8223-8236
<6>2011
<7>A type IIG restriction endonuclease, RM.BpuSI from Bacillus pumilus, has been characterized
and its X-ray crystal structure determined at 2.35A
resolution. The enzyme is comprised of an array of 5-folded domains that
couple the enzyme's N-terminal endonuclease domain to its C-terminal
target recognition and methylation activities. The REase domain contains a
PD-x(15)-ExK motif, is closely superimposable against the FokI
endonuclease domain, and coordinates a single metal ion. A helical bundle
domain connects the endonuclease and methyltransferase (MTase) domains.
The MTase domain is similar to the N6-adenine MTase M.TaqI, while the
target recognition domain (TRD or specificity domain) resembles a
truncated S subunit of Type I R-M system. A final structural domain, that
may form additional DNA contacts, interrupts the TRD. DNA binding and
cleavage must involve large movements of the endonuclease and TRD domains,
that are probably tightly coordinated and coupled to target site
methylation status.

<>

<1>Shen, J., Rideout, W.M. III, Jones, P.A.
<2>High frequency mutagenesis by a DNA methyltransferase.
<3>Cell
<4>71
<5>1073-1080
<6>1992
<7>HpaII methylase (M. HpaII), an example of a DNA(cytosine-5)-methyltransferase, was found to
induce directly a high frequency of C to U transition mutations in double-stranded DNA. A
mutant pSV2-neo plasmid, constructed with an inactivating T to C transition mutation creating
a CCGG site, was incubated with M.HpaII in the absence of S-adenosylmethionine (SAM). This
caused an approximately 104-fold increase in the rate of reversion when the mutant neo plasmid
was transformed into bacteria lacking uracil-DNA glycosylase. The mutation frequency was very
sensitive to SAM concentration and was reduced to background when the concentration of the
methyl donor exceeded 300 nM. The data support current models for the formation of a covalent
complex between the methyltransferase and cytosine. They also suggest that the occurrence of
muational hot spots at CpG sites may not always be due to spontaneous deamination of
5-methylcytosine, but might also be initiated by enzymatic deamination of cytosine and proceed
through a C to U to T pathway.

<>

<1>Shen, J., Rideout, W.M. III, Jones, P.A.
<2>The rate of hydrolytic deamination of 5-methylctyosine in double-stranded DNA.
<3>Nucleic Acids Res.
<4>22
<5>972-976
<6>1994
<7>The modified base, 5-methylcytosine, constitutes approximately 1% of human DNA, but sites
containing 5-methylcytosine account for at least 30% of all germline and somatic point
mutations. A genetic assay with a sensitivity of 1 in 10/7, based on reversion to neomycin
resistance of a mutant pSV2-neo plasmid, was utilized to determine and compare the deamination
rates of 5-methylcytosine and cytosine in double-stranded DNA for the first time. The rate
constants for spontaneous hydrolytic deamination of 5-methylcytosine and cytosine in
double-stranded DNA at 37 C were 5.8x10/-13 s/-1 and 2.6 x 10/-13 s/-1, respectively. These
rates are more than sufficient to explain the observed frequency of mutation at sites
containing 5-methylcytosine and emphasize the importance of hydrolytic deamination as a major
source of human mutations.

<>

<1>Shen, J.-C., Zingg, J.-M., Yang, A.S., Schmutte, C., Jones, P.A.
<2>A mutant HpaII methyltransferase functions as a mutator enzyme.
<3>Nucleic Acids Res.
<4>23
<5>4275-4282
<6>1995
<7>DNA (cytosine-5)-methyltransferases can cause deamination of cytosine when the cofactor
S-adenosylmethionine (AdoMet) is limiting and thus function as sequence-specific C-U mutator
enzymes.  Here we explored whether mutations causing inactivation of the cofactor binding
activity of the HpaII methyltransferase, thus mimicking conditions of limiting AdoMet
concentration, could convert a DNA methyltransferase to a C-U mutator enzyme.  We created two
mutator enzymes from the HpaII methyltransferase (F38S and G40D) which both showed enhanced
cytosine deamination activities in vitro and in vivo.  Interestingly, the G:U mispairs
generated by these enzymes were not repaired completely in bacteria equipped with uracil-DNA
glycosylase-initiated repair machinery, giving rise to a potent mutator phenotype.  This is
the first report showing the creation of mutator enzymes from a DNA methyltransferase and the
demonstration of their mutagenicity in living cells.

<>

<1>Shen, K. et al.
<2>Extensive genomic plasticity in Pseudomonas aeruginosa revealed by identification and distribution studies of novel genes among clinical isolates.
<3>Infect. Immun.
<4>74
<5>5272-5283
<6>2006
<7>The distributed genome hypothesis (DGH) states that each strain within a
bacterial species receives a unique distribution of genes from a
population-based supragenome that is many times larger than the genome of
any given strain. The observations that natural infecting populations are
often polyclonal and that most chronic bacterial pathogens have highly
developed mechanisms for horizontal gene transfer suggested the DGH and
provided the means and the mechanisms to explain how chronic infections
persist in the face of a mammalian host's adaptive defense mechanisms.
Having previously established the validity of the DGH for obligate
pathogens, we wished to evaluate its applicability to an opportunistic
bacterial pathogen. This was accomplished by construction and analysis of
a highly redundant pooled genomic library containing approximately 216,000
functional clones that was constructed from 12 low-passage clinical
isolates of Pseudomonas aeruginosa, 6 otorrheic isolates and 6 from other
body sites. Sequence analysis of 3,214 randomly picked clones (mean insert
size, approximately 1.4 kb) from this library demonstrated that 348
(10.8%) of the clones were unique with respect to all genomic sequences of
the P. aeruginosa prototype strain, PAO1. Hypothetical translations of the
open reading frames within these unique sequences demonstrated protein
homologies to a number of bacterial virulence factors and other proteins
not previously identified in P. aeruginosa. PCR and reverse
transcription-PCR-based assays were performed to analyze the distribution
and expression patterns of a 70-open reading frame subset of these
sequences among 11 of the clinical strains. These sequences were unevenly
distributed among the clinical isolates, with nearly half (34/70) of the
novel sequences being present in only one or two of the individual
strains. Expression profiling revealed that a vast majority of these
sequences are expressed, strongly suggesting they encode functional
proteins.

<>

<1>Shen, L., Gao, G., Zhang, Y., Zhang, H., Ye, Z., Huang, S., Huang, J., Kang, J.
<2>A single amino acid substitution confers enhanced methylation activity of mammalian Dnmt3b on chromatin DNA.
<3>Nucleic Acids Res.
<4>38
<5>6054-6064
<6>2010
<7>Dnmt3a and Dnmt3b are paralogous enzymes responsible for de novo DNA methylation but with
distinguished biological functions. In mice,
disruption of Dnmt3b but not Dnmt3a causes global DNA hypomethylation,
especially in repetitive sequences, which comprise the large majority of
methylated DNA in the genome. By measuring DNA methylation activity of
Dnmt3a and Dnmt3b homologues from five species, we found that mammalian
Dnmt3b possessed significantly higher methylation activity on chromatin
DNA than Dnmt3a and non-mammalian Dnmt3b. Sequence comparison and
mutagenesis experiments identified a single amino acid substitution
(I662N) in mammalian Dnmt3b as being crucial for its high chromatin DNA
methylation activity. Further mechanistic studies demonstrated this
substitution markedly enhanced the binding of Dnmt3b to nucleosomes and
hence increased the chromatin DNA methylation activity. Moreover, this
substitution was crucial for Dnmt3b to efficiently methylate repetitive
sequences, which increased dramatically in mammalian genomes. Consistent
with our observation that Dnmt3b evolved more rapidly than Dnmt3a during
the emergence of mammals, these results demonstrated that the I662N
substitution in mammalian Dnmt3b conferred enhanced chromatin DNA
methylation activity and contributed to functional adaptation in the
epigenetic system.

<>

<1>Shen, P., Jiang, Y., Zhou, Z., Zhang, J., Yu, Y., Li, L.
<2>Complete nucleotide sequence of pKP96, a 67 850 bp multiresistance plasmid encoding qnrA1, aac(6')-Ib-cr and blaCTX-M-24 from Klebsiella pneumoniae.
<3>J. Antimicrob. Chemother.
<4>62
<5>1252-1256
<6>2008
<7>OBJECTIVES: The multiresistance plasmid pKP96 from Klebsiella pneumoniae
was sequenced completely and analysed concerning its genetic environment
and distributing of antimicrobial resistance genes. METHODS: The complete
sequence of the plasmid was determined using a whole-genome shotgun
approach. MICs of 13 antimicrobial agents were determined using Etests. A
conjugation experiment was performed in liquid medium. RESULTS: pKP96 is a
circularly closed 67 850 bp multiresistance plasmid with an IncN
incompatibility group. Seventy putative genes were identified according to
the annotation of the finished sequence. The backbone region of the
plasmid, comprising the conjugal transfer and plasmid replication regions,
showed 91% identity to the IncN plasmid R46. Several mobile elements were
found to be inserted into pKP96 together with antimicrobial resistance
genes, including qnrA1, aac(6')-Ib-cr and bla(CTX-M-24). CONCLUSIONS:
Plasmid pKP96 is a chimera that has acquired its multiple antimicrobial
resistance determinants horizontally from different sources. It may have
evolved from an ancestor plasmid similar to R46 through the stepwise
events of integration or recombination.

<>

<1>Shen, S., Li, Q., Yan, P., Zhou, B., Ye, S., Lu, Y., Wang, D.
<2>Restriction endonucleases from three strains of Haemophilus influenzae.
<3>Sci. Sin.
<4>23
<5>1435-1442
<6>1980
<7>The restriction endonucleases, Hin P1I, Hin S1I and Hin S2I are isolated from
three strains of Haemophilus influenzae respectively.  By polymin P treatment,
ammonium sulphage precipitation and column chromatography on phosphocellulose
and on heparin-Sepharose, Hin P1I is partially purified.  No contaminating
deoxyribonuclease activities have been detected in this purified enzyme
preparation.  The face that the digestion patterns of Hin P1I and HhaI on phage
lambda, plasmids ColE1 and pBR 322 DNAs are identical indicates that they are
isoschizomers but their splitting sites are different.  The banding patterns of
Hin S1I and Hin S2I are also the same as that of HhaI.

<>

<1>Shen, X., Chen, M., Hu, H., Wang, W., Peng, H., Xu, P., Zhang, X.
<2>Genome Sequence of Pseudomonas chlororaphis GP72, a Root-Colonizing Biocontrol Strain.
<3>J. Bacteriol.
<4>194
<5>1269-1270
<6>2012
<7>Pseudomonas chlororaphis GP72 is a root-colonizing biocontrol strain isolated from a green
pepper rhizosphere. It can produce several secondary metabolites to
suppress phytopathogens. Here we present a 6.6-Mb assembly of its genome, which
is the first genome sequence of the P. chlororaphis group and may provide
insights into its antifungal activities.

<>

<1>Shen, X., Wang, Q., Xia, L., Zhu, X., Zhang, Z., Liang, Y., Cai, H., Zhang, E., Wei, J., Chen, C., Song, Z., Zhang, H., Yu, D., Hai, R.
<2>Complete Genome Sequences of Yersinia pestis from Natural Foci in China.
<3>J. Bacteriol.
<4>192
<5>3551-3552
<6>2010
<7>Yersinia pestis, the causative agent of plague, is a deadly bacterium that affects humans.
Strain D106004 was isolated from a new plague focus in
Yulong County, China, in 2006. To gain insights into the epidemic origin,
we have sequenced the genomes of D106004 and strains Z176003 and D182038,
isolated from neighboring regions.

<>

<1>Shen, Z., Sheh, A., Young, S.K., Abouelliel, A., Ward, D.V., Earl, A.M., Fox, J.G.
<2>Draft genome sequences of six enterohepatic helicobacter species isolated from humans and one from rhesus macaques.
<3>Genome Announcements
<4>2
<5>e00857-14
<6>2014
<7>Draft genome sequences of seven enterohepatic Helicobacter species, H. bilis, H.  canadensis,
H. canis, H. cinaedi, H. winghamensis, H. pullorum, and H. macacae,
are presented. These isolates were obtained from clinical patients and a nonhuman
primate. Due to potential zoonotic risks, we characterized antibiotic resistance
markers and Helicobacter virulence factors.

<>

<1>Shendure, J., Porreca, G.J., Reppas, N.B., Lin, X., McCutcheon, J.P., Rosenbaum, A.M., Wang, M.D., Zhang, K., Mitra, R.D., Church, G.M.
<2>Accurate multiplex polony sequencing of an evolved bacterial genome.
<3>Science
<4>309
<5>1728-1732
<6>2005
<7>We describe a DNA sequencing technology in which a commonly available, inexpensive
epifluorescence microscope is converted to rapid
nonelectrophoretic DNA sequencing automation. We apply this technology to
resequence an evolved strain of Escherichia coli at less than one error
per million consensus bases. A cell-free, mate-paired library provided
single DNA molecules that were amplified in parallel to 1-micrometer beads
by emulsion polymerase chain reaction. Millions of beads were immobilized
in a polyacrylamide gel and subjected to automated cycles of sequencing by
ligation and four-color imaging. Cost per base was roughly one-ninth as
much as that of conventional sequencing. Our protocols were implemented
with off-the-shelf instrumentation and reagents.

<>

<1>Sheng, B., Ni, J., Gao, C., Ma, C., Xu, P.
<2>Draft Genome Sequence of the Gluconobacter oxydans Strain DSM 2003, an Important  Biocatalyst for Industrial Use.
<3>Genome Announcements
<4>2
<5>e00417-14
<6>2014
<7>Gluconobacter oxydans strain DSM 2003 can efficiently produce some industrially important
building blocks, such as (R)-lactic acid and (R)-2-hydroxybutyric acid.
Here, we present a 2.94-Mb assembly of its genome sequence, which might provide
further insights into the molecular mechanism of its biocatalysis in order to
further improve its biotechnological applications.

<>

<1>Shenoy, S., Daigle, K., Ehrlich, K.C., Gehrke, C.W., Ehrlich, M.
<2>Hydrolysis by restriction endonucleases at their DNA recognition sequences substituted with mismatched base pairs.
<3>Nucleic Acids Res.
<4>14
<5>4407-4420
<6>1986
<7>Restriction endonucleases were tested for their ability to catalyze the
cleavage of mismatch-containing recognition sites in DNA.  These mismatched
base pairs were T.G, U.G, or A.C in covalently closed, circular heteroduplexes
prepared by in vitro extension of chemically synthesized oligonucleotide
primers annealed to a bacteriophage M13-derived viral DNA.  None of the
restriction enzymes was able to completely cleave the mismatch-containing
recognition sites under standard conditions.  However, three of them, SmaI,
SalI, and SstI, catalyzed partial digestion leading to an accumulation of DNA
singly nicked at the mismatched recognition site.  The ability of SmaI and SstI
to partially cleave at a mismatch was shown to depend on the nature and
position of the mismatch within the corresponding recognition site.  In
contrast, little or no digestion was obtained with AccI, HincII, HindIII, and
KpnI at mismatch-containing sites.  Therefore, in some cases a transition-type
substitution in only one strand of a recognition site inhibits restriction
endonuclease-catalyzed digestion at that site although in others partial
digestion occurs.

<>

<1>Shepard, S.M., Danzeisen, J.L., Isaacson, R.E., Seemann, T., Achtman, M., Johnson, T.J.
<2>Genome sequences and phylogenetic analysis of K88- and F18-positive porcine enterotoxigenic Escherichia coli.
<3>J. Bacteriol.
<4>194
<5>395-405
<6>2012
<7>Porcine enterotoxigenic Escherichia coli (ETEC) continues to result in major morbidity and
mortality in the swine industry via postweaning diarrhea. The key
virulence factors of ETEC strains, their serotypes, and their fimbrial components
have been well studied. However, most studies to date have focused on
plasmid-encoded traits related to colonization and toxin production, and the
chromosomal backgrounds of these strains have been largely understudied. Here, we
generated the genomic sequences of K88-positive and F18-positive porcine ETEC
strains and examined the phylogenetic distribution of clinical porcine ETEC
strains and their plasmid-associated genetic content. The genomes of porcine ETEC
strains UMNK88 and UMNF18 were both found to contain remarkable plasmid
complements containing known virulence factors, potential novel virulence
factors, and antimicrobial resistance-associated elements. The chromosomes of
these strains also possessed several unique genomic islands containing
hypothetical genes with similarity to classical virulence factors, although
phage-associated genomic islands dominated the accessory genomes of these
strains. Phylogenetic analysis of 78 clinical isolates associated with neonatal
and porcine diarrhea revealed that a limited subset of porcine ETEC lineages
exist that generally contain common toxin and fimbrial profiles, with many of the
isolates belonging to the ST10, ST23, and ST169 multilocus sequencing types.
These lineages were generally distinct from existing human ETEC database
isolates. Overall, most porcine ETEC strains appear to have emerged from a
limited subset of E. coli lineages that either have an increased propensity to
carry plasmid-encoded virulence factors or have the appropriate ETEC core genome
required for virulence.

<>

<1>Sheppard, A.E., Poehlein, A., Rosenstiel, P., Liesegang, H., Schulenburg, H.
<2>Complete Genome Sequence of Bacillus thuringiensis Strain 407 Cry-.
<3>Genome Announcements
<4>1
<5>e00158-12
<6>2013
<7>Bacillus thuringiensis is an insect pathogen that has been used widely as a biopesticide.
Here, we report the genome sequence of strain 407 Cry-, which is
used to study the genetic determinants of pathogenicity. The genome consists of a
5.5-Mb chromosome and nine plasmids, including a novel 502-kb megaplasmid.

<>

<1>Sherburne, C.K., Lawley, T.D., Gilmour, M.W., Blattner, F.R., Burland, V., Grotbeck, E., Rose, D.J., Taylor, D.E.
<2>The complete DNA sequence and analysis of R27, a large IncHI plasmid from Salmonella typhi that is temperature sensitive for transfer.
<3>Nucleic Acids Res.
<4>28
<5>2177-2186
<6>2000
<7>Salmonella typhi, the causative agent of typhoid fever, annually infects 16 million people and
kills 600,000 world wide. Plasmid-encoded multiple drug resistance in S. typhi is always
encoded by plasmids of incompatibility group H (IncH). The complete DNA sequence of the large
temperature-sensitive conjugative plasmid R27, the prototype for the IncHI1 family of
plasmids, has been compiled and analyzed. This 180 kb plasmid contains 210 open reading frames
(ORFs), of which 14 have been previously identified and 56 exhibit similarity to other plasmid
and prokaryotic ORFs. A number of insertion elements were found, including the full Tn 10
transposon, which carries tetracycline resistance genes. Two transfer regions, Tra1 and Tra2,
are present, which are separated by a minimum of 64 kb. Homologs of the DNA-binding proteins
TlpA and H-NS that act as temperature-regulated repressors in other systems have been located
in R27. Sequence analysis of transfer and replication regions supports a mosaic-like structure
for R27. The genes responsible for conjugation and plasmid maintenance have been identified
and mechanisms responsible for thermosensitive transfer are discussed.

<>

<1>Sheth, A., Ravikumar, M., Hosur, R.V., Govil, G.
<2>Two dimensional NMR studies on the solution structure of d-CTCGAGCTCGAG.
<3>Biochem. Biophys. Res. Commun.
<4>144
<5>26-34
<6>1987
<7>Two dimensional (2D) FT-NMR investigations have been carried out on the
self-complementary dodecanucleotide d-CTCGAGCTCGAG, which has cleavage sites
for the restriction enzyme XhoI (between C and T).  The central TCG portion is
also known to show a preference for DNAase activity.  Complete resonance
assignments have been obtained for the non-exchangeable sugar and base protons
of the oligonucleotide.  Information regarding sugar geometries, glycosidic
torsion angles and other structural parameters has been obtained from the
relative intensities of the cross peaks in the COSY and NOESY spectra.  The
results indicate that deoxyribose rings of C1 and C7 adopt a conformation
different from the remaining sugars in the double helical oligonucleotide.  The
Central TCG portion also exhibits variations in the backbone structure.  The
base stacking in the double helix shows interesting sequence dependent effects
suggesting that the sequence effects are not localised to nearest neighbours
but extended over longer stretches.

<>

<1>Shetty, S.A., Marathe, N.P., Munot, H., Antony, C.P., Dhotre, D.P., Murrell, J.C., Shouche, Y.S.
<2>Draft Genome Sequence of Methylophaga lonarensis MPLT, a Haloalkaliphilic (Non-Methane-Utilizing) Methylotroph.
<3>Genome Announcements
<4>1
<5>e00202-13
<6>2013
<7>Methylophaga lonarensis strain MPL(T) is a haloalkaliphilic methylotroph isolated from Lonar
Lake, a saline and alkaline lake in Maharashtra, India. Strain MPL(T)
utilizes methanol as its sole carbon and energy source. Here, we present the
draft genome sequence of M. lonarensis MPL(T) (VKM B-2684(T) = MCC 1002(T)).

<>

<1>Shevchuk, T., Kretzner, L., Munson, K., Axume, J., Clark, J., Dyachenko, O.V., Caudill, M., Buryanov, Y., Smith, S.S.
<2>Transgene-induced CCWGG methylation does not alter CG methylation patterning in human kidney cells.
<3>Nucleic Acids Res.
<4>33
<5>6124-6136
<6>2005
<7>Several reports suggest that C(m)CWGG methylation tends not to co-exist with (m)CG methylation
in human cells. We have asked whether or not
methylation at CCWGG sites can influence CG methylation. DNA from cells
expressing an M.EcoRII-GFP fusion was actively methylated at CCWGG sites.
CG methylation as measured by R.HpaII/R.MspI ratios was unchanged in cells
expressing the transgene. Cloned representatives of C(m)CWGG methylated
DNA often contained, or were adjacent to an ALU repeat, suggesting that
M.EcoRII-GFP actively methylated gene-rich R-band DNA. The transgenic
methyltransferase applied C(m)CWGG methylation to a representative human
promoter that was heavily methylated at CG dinucleotides (the SERPINB5
promoter) and to a representative promoter that was essentially
unmethylated at CG dinucleotides (the APC promoter). In each case, the CG
methylation pattern remained in its original state, unchanged by the
presence of neighboring C(m)CWGG sites. Q-PCR measurements showed that RNA
expression from the APC gene was not significantly altered by the presence
of C(m)CWGG in its promoter. Kinetic studies suggested that an adjacent
C(m)CWGG methylation site influences neither the maintenance nor the de
novo methylation activities of purified human Dnmt1. We conclude that
C(m)CWGG methylation does not exert a significant effect on CG methylation
in human kidney cells.

<>

<1>Shevchuk, T.V., Buryanov, Y.I.
<2>DNA methyltransferase-based assay for the cytosine methylation level in the DNA sequence CCWGG.
<3>Bioorg. Khim.
<4>25
<5>630-633
<6>1999
<7>An assay for the cytosine methylation level in the eukaryotic DNA CCWGG sequence is proposed.
The method is based on the ability of DNA methylase BstNI to methylate DNA containing in a
CCWGG site a nonmodified or 5-methylated cytosine to yield N4-methyl- or
N4,5-dimethylcytosine, respectively.

<>

<1>Shevchuk, T.V., Zakharchenko, N.S., Dyachenko, O.V., Buryanov, Y.I.
<2>The effect of the gene encoding EcoRII DNA methyltransferase on the phenotype of transgenic tumor cell lines of Nicotiana tabacum and the  methylation level of CpNpG sequences in their genome.
<3>Russ. J. Plant Physiol.
<4>48
<5>478-482
<6>2001
<7>Several tumor cell lines were obtained by transforming Nicotiana tabacum plants with the
recombinant Ti plasmid comprising the gene
encoding EcoRII DNA methyltransferase (M.EcoRII) and subjected to
analysis. The transformed lines differed in their morphology, growth
dependence on hormones, and nopaline-synthesizing capacity. Southern
blot-hybridization showed that the M.EcoRII gene was present in the
cells of all transformed lines. However, genome analysis using
polymerase chain reaction with the oligonucleotide primers recognizing
5'-ends of the M.EcoRII gene did not exhibit the full-length copies of
the gene. Lower methylation of CpNpG sequences characteristic of all
transformed cells could result from the disturbance of one of several
plant DNA methyltransferase genes following its homologous
recombination with the M.EcoRII gene.

<>

<1>Shevtsov, A., Tarlykov, P., Zholdybayeva, E., Momynkulov, D., Sarsenova, A., Moldagulova, N., Momynaliev, K.
<2>Draft Genome Sequence of Rhodococcus erythropolis DN1, a Crude Oil Biodegrader.
<3>Genome Announcements
<4>1
<5>e00846-13
<6>2013
<7>We report the 6,548-Mb genome sequence of Rhodococcus erythropolis strain DN1, isolated from
the oil-contaminated soil in the Karagandy region of Kazakhstan.
The draft genome sequence of strain DN1 might provide new insights into the
genetic mechanisms of crude oil biodegradation.

<>

<1>Shevtsov, A., Tarlykov, P., Zholdybayeva, E., Shevtsova, E., Momynkulov, D., Sytnik, I., Karibaev, T., Chsherbakov, A., Momynaliev, K.
<2>Draft Genome Sequence of the Live Vaccine Strain Brucella abortus 82.
<3>Genome Announcements
<4>1
<5>e01101-13
<6>2013
<7>Vaccination is a crucial part of the brucellosis eradication programs worldwide.  A live
vaccine strain of Brucella abortus 82 has been successfully used for the
vaccination of cattle against brucellosis in the former Soviet republics for the
last 39 years. Here, we report the genome sequence of Brucella abortus 82.

<>

<1>Shi, C.Y., Jia, K.T., Yang, B., Huang, J.
<2>Complete genome sequence of a Megalocytivirus (family Iridoviridae) associated with turbot mortality in China.
<3>Virol. J.
<4>7
<5>159
<6>2010
<7>ABSTRACT: BACKGROUND: Turbot reddish body iridovirus (TRBIV) causes
serious systemic diseases with high mortality in the cultured turbot,
Scophthalmus maximus. We here sequenced and analyzed the complete genome
of TRBIV, which was identified in Shandong province, China. RESULTS: The
genome of TRBIV is a linear double-stranded DNA of 110,104 base pairs,
comprising 55% G + C. Total 115 open reading frames were identified,
encoding polypeptides ranging from 40 to 1168 amino acids. Amino acid
sequences analysis revealed that 39 of the 115 potential gene products of
TRBIV show significant homology to other iridovirus proteins. Phylogenetic
analysis of conserved genes indicated that TRBIV is closely related to
infectious spleen and kidney necrosis virus (ISKNV), rock bream iridovirus
(RBIV), orange-spotted grouper iridovirus (OSGIV), and large yellow
croaker iridovirus (LYCIV). The results indicated that TRBIV belongs to
the genus Megalocytivirus (family Iridoviridae). CONCLUSIONS: The
determination of the genome of TRBIV will provide useful information for
comparative study of Megalocytivirus and developing strategies to control
outbreaks of TRBIV-induced disease.

<>

<1>Shi, L., Suetake, I., Kawakami, T., Aimoto, S., Tajima, S.
<2>Xenopus Eggs Express an Identical DNA Methyltransferase, Dnmt1, to Somatic Cells.
<3>J. Biochem. (Tokyo)
<4>130
<5>359-366
<6>2001
<7>In mouse, an oocyte-specific short isoform of DNA methyltransferase-1 (Dnmt1) lacking amino
terminal 118 amino acid residues exists and plays a crucial role in maintaining the
methylation state of imprinted genes during early embryogenesis [Howell et al. (2001) Cell
104, 829-838]. To address the question of whether or not Xenopus oocyte expresses such a short
isoform, we raised monoclonal antibodies against the amino-terminal portion of Xenopus Dnmt1.
Two of the isolated monoclonal antibodies, 3C6 and 4A8, were determined to recognize (1-32)
and (115-126) of Xenopus Dnmt1, respectively. The amounts of Dnmt1 in Xenopus eggs were
determined to be similar, 10.0 2.5, 8.0 0.8, and 8.2 0.2 ng per egg with monoclonal antibodies
3C6 and 4A8, and polyclonal antibodies, respectively. This indicated that Dnmt1 in Xenopus
mature eggs had an identical amino-terminal sequence to the amino acid sequence deduced from
the cDNA. Together with the fact that Dnmt1 in A6 cells immuno-reacted with all the monoclonal
antibodies isolated and with the polyclonal antibodies, we concluded that Dnmt1 expressed in
Xenopus mature eggs possesses an identical amino-terminal sequence to that in somatic cells.
Immuno-purified Xenopus Dnmt1 in mature eggs showed similar specific activity to that in
proliferating A6 cells and that of mouse recombinant Dnmt1.

<>

<1>Shi, X., Yu, M., Yan, S., Dong, S., Zhang, X.H.
<2>Genome Sequence of the Thermostable-Agarase-Producing Marine Bacterium Catenovulum agarivorans YM01T, Which Reveals the Presence of a Series of  Agarase-Encoding Genes.
<3>J. Bacteriol.
<4>194
<5>5484
<6>2012
<7>Marine bacterium Catenovulum agarivorans YM01(T) can produce highly thermostable  agarases.
The draft genome of YM01(T) is about 5.36 Mb and harbors approximately
4,913 genes, including 15 agarase (2 alpha-agarase and 13 beta-agarase)-encoding
genes, which will provide references to functional characterization of various
agarases from marine bacteria.

<>

<1>Shi, X., Zhu, Y., Li, Y., Jiang, M., Lin, Y., Qiu, Y., Chen, Q., Yuan, Y., Ni, P., Hu, Q., Huang, S.
<2>Genome Sequence of Proteus mirabilis Clinical Isolate C05028.
<3>Genome Announcements
<4>2
<5>e00167-14
<6>2014
<7>Genomic DNA of Proteus mirabilis C05028 was sequenced by an Illumina HiSeq platform and was
assembled to 39 scaffolds with a total length of 3.8 Mb. Next,
open reading frames (ORFs) were identified and were annotated by the KEGG, COG,
and NR databases. Finally, we found special virulence factors only existing in P.
mirabilis C05028.

<>

<1>Shia, M.A., Wong, G.G.
<2>Analyzing nucleic acids (NA) by direct labeling of methylated sites on NAs using agents that bind to methylated nucleotides, and by enzymatic modification of the NAs resulting in labeled nucleotides - methylated DNA labelling useful for genomics.
<3>International Patent Office
<4>WO 2002101353
<5>
<6>2002
<7>NOVELTY - Analyzing nucleic acids (NA) by direct labeling of methylated sites on NAs (using
agents that bind to methylated
nucleotides), and by enzymatic modification of NAs resulting in labeled
nucleotides. Direct labeling methods use methylated NA binding proteins
(MBPs) or methylation-specific antibodies or antibody fragments. Enzymatic
modification methods use labeled methylation cofactors e.g., S-adenosyl
methionine (or cofactor derivatives). DETAILED DESCRIPTION - Analyzing NA
molecule involves (M1-M5): (a) exposing a NA molecule to a
sequence-specific methylase and an S-adenosyl methionine (SAM) labeled
derivative, allowing the sequence-specific methylase to label NA molecule
with the SAM labeled derivative, and determining a labeling pattern in the
NA molecule using a linear polymer analysis system, where the labeling
pattern is indicative of a methylation pattern of the NA molecule; (b)
exposing NA to a labeled sequence-specific methylase and a SAM derivative,
allowing the labeled sequence-specific methylase to bind to NA with the
SAM derivative and label NA, and determining a labeling pattern in the NA
molecule using a linear polymer analysis system, where the labeling
pattern is indicative of a methylation pattern of the NA molecule; (c)
exposing NA molecule to MBP, and determining the pattern of binding of MBP
to NA molecule using a linear polymer analysis system, where the pattern
of binding of MBP is indicative of methylation pattern of NA molecule; (d)
exposing NA molecule to a methylation-specific antibody or its fragment,
determining the pattern of binding of the methylation-specific antibody or
its fragment to NA molecule using a linear polymer analysis system, where
the pattern of binding of the methylation-specific antibody or its
fragment is indicative of methylation pattern of NA; or (e) exposing NA to
a sequence-specific methylase and a labeled SAM, allowing the SAM to label
the NA with labeled SAM, and determining a labeling pattern in NA using a
linear polymer analysis system, where the labeling pattern is indicative
of a methylation pattern of the NA.INDEPENDENT CLAIMS are also included
for: (1) a system (I) for optically analyzing NA molecule comprising an
optical source for emitting optical radiation of a known wavelength, an
interaction station for receiving the optical radiation in an optical path
and for receiving the NA molecule that is exposed to the optical radiation
to produce detectable signals, dichroic reflectors in the optical path for
creating at least two separate wavelength bands of the detectable signals,
optical detectors constructed to detect radiation including the signals
resulting from interaction of NA molecule with the optical radiation, and
a processor constructed and arranged to analyze the NA molecule based on
the detected radiation including the signals, where the NA molecule is
labeled according to its methylation status; and (2) a method (M6) for
analyzing NA molecules.

<>

<1>Shiba, T., Hattori, M., Sakaki, Y., Horikawa, H., Kikuchi, H., Ishikawa, J., Ikeda, H., Omura, S.
<2>Novel polynucleotides.
<3>Japanese Patent Office
<4>JP 2003284572 A
<5>
<6>2003
<7>
<>

<1>Shibata, T., Ando, T.
<2>The restriction endonucleases in Bacillus amyloliquefaciens N strain.  Substrate specificities.
<3>Biochim. Biophys. Acta
<4>442
<5>184-196
<6>1976
<7>Two species of restriction endonuclease were isolated by gel filtration and
DEAE-cellulose chromatography from a cell-free extract of Bacillus
amyloliquefaciens (B. subtilis) N strain; a lower molecular weight endonuclease
(endonuclease R.BamNI) and a higher molecular-weight one (endonuclease
R.BamNx).  Both of them required only Mg2+ for their activities.  Endonuclease
R.BamNx introduced a larger number of site-specific scissions in Escheria coli
phage lambda DNA than endonuclease R.BamNI did.  Endonuclease R.BamNx cleaved
Bacillus phage Phi105C DNA at the specific sites which are classified into two
groups: one type of sites is modified by B. amyloliquefaciens H strain in vivo
while the other is not affected.  It was also active on DNAs of E. coli phage
T7, lambda dvl, Simian virus 40 (SV40) and colicinogenic factor ColEI and was
inactive on DNAs of Bacillus phages Phi29 and M2.  Endonuclease R.BamNi cleaved
DNA in the same nucleotide sequences as endonuclease R.BamHI isolated from H
strain by Wilson and Young.  This endonuclease was active on DNAs of phage
lambda, lambda dvl and SV40, and was inactive of phages Phi105C, Phi29, M2 and
T7, and ColEI DNA.

<>

<1>Shibata, T., Ando, T.
<2>In vitro Modification and Restriction of Phage Phi105C DNA with Bacillus subtilis N Cell-free Extract.
<3>Mol. Gen. Genet.
<4>138
<5>269-279
<6>1975
<7>The enzymes involved in host-controlled modification and restriction by
Bacillus subtilis strain N were detected in cell-free extracts.  In the
presence of Mg2+, the N-specific endonuclease cleaved unmodified DNA but did
not attack Phi105C.N DNA carrying N-specific modification.  The restriction
endonuclease required neither SAM nor ATP for its activity.  The N-specific
modification enzyme was active only in the presence of SAM, indicating that
modification in this system is a methylation of DNA.

<>

<1>Shibata, T., Ando, T.
<2>Host controlled modification and restriction in Bacillus subtilis.
<3>Mol. Gen. Genet.
<4>131
<5>275-280
<6>1974
<7>The host controlled modification and restriction was found in Bacillus subtilis
Marburg 168, Bacillus subtilis (B. amyloliquefaciens) N and H, by use of a
clear plaque mutant of temperate phage Phi105 (Phi105C).  This phenomenon of
"modification and restriction" was similar to that found in Escherichia coli,
Haemophilus influenzae and other micro-organisms.  Phi105 carrying Marburg 168
specific modification (Phi105C.168) plated on B. subtilis N and H with an
efficiency of 10-5 and 10-2, respectively.  Phi105C carrying the modification
endowed by B. subtilis N (Phi105C.N) had an efficiency of plating on B.
subtilis 168 of 4 x 10-2 and was not restricted by B. subtilis H.  Phi105C
carrying H specific modification (Phi105C.H) plated on B. subtilis 168 and N
with an efficiency of 10-1-10-2 and 10-5, respectively.  Modification type of
Phi105C was determined by the last host strain and was not genetic behavior of
the phage.  Efficiencies of plating of Bacillus phages Phi29 and SPP1 were not
affected by the modification and restriction described in the ent paper.

<>

<1>Shibata, T., Hayase, E., Ando, T.
<2>Simultaneous preparation of a DNA ligase and restriction endonucleases from Bacillus amyloliquefaciens N.
<3>Agric. Biol. Chem.
<4>42
<5>1613-1615
<6>1978
<7>None

<>

<1>Shibata, T., Ikawa, S., Ando, T.
<2>Genetic study of restriction endonucleases in Bacillus subtilis.
<3>Microbiology-1982, American Society for Microbiology, Schlessinger, D., Washington
<4>0
<5>71-74
<6>1982
<7>During this decade, research has revealed that various microorganisms have a
type of intracellular endodeoxyribonuclease that recognizes specific nucleotide
sequences in double-stranded DNA and introduces double-stranded scissions at or
near those sequences.  These endonucleases are called site-specific
endonucleases or type II restriction endonucleases, although only a few of them
have been proven to be involved in restriction and modification of genetically
foreign entities invading into a cell.  Therefore, this type of endonuclease
might have other biological roles in living cells, such as recombination.  In
Bacillus subtilis, restriction and modification of phages were not known until
1974, when Trautner et al. and we found, independently, that strains of B.
subtilis, including the 168 strain, exhibited restriction and modification of
phages SPP1 or Phi105C.  We then found that 13 of 40 B. subtilis strains
(including B. amyloliquefaciens) and 11 strains of other Bacillus species have
site-specific endonucleases.  As an initial step in elucidating the biological
roles of site-specific endonucleases, we analyzed the genes for such
endonucleases.

<>

<1>Shibata, T., Ikawa, S., Kim, C., Ando, T.
<2>Site-specific Deoxyribonucleases in Bacillus subtilis and other Bacillus strains.
<3>J. Bacteriol.
<4>128
<5>473-476
<6>1976
<7>We systematically studied site-specific deoxyribonucleases in Bacillus strains
and detected deoxyribonuclease activities in 20 of 62 strains tested.

<>

<1>Shibata, T., Ikawa, S., Komatsu, Y., Ando, T., Saito, H.
<2>Introduction of host-controlled modification and restriction systems of Bacillus subtilis IAM1247 into Bacillus subtilis 168.
<3>J. Bacteriol.
<4>139
<5>308-310
<6>1979
<7>Bacillus subtilis IAM1247 had two modification and restriction systems
(Bsu12471 and Bsu1247II), the former producing an isoschizomer of PstI
endonuclease.  A transformant clone was isolated which had Bsu 168, BsuR, and
Bsu12471 systems coexisting within a genome.

<>

<1>Shibata, T., Morishima, N.
<2>Genetic recombination and site-specific endonucleases in mitochondria.
<3>Tanpakushitsu Kakusan Koso
<4>39
<5>589-600
<6>1994
<7>
<>

<1>Shibata, T., Morishima, N., Nakagawa, K.
<2>A mitochondrial multi-site specific endonuclease, endo SceI of Saccharomyces cerevisiae.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>154
<6>1993
<7>There are two classes of sequence specific endonucleases active on double-stranded DNA in
eukaryotic cells. Endonucleases in one class exhibit very strict site-specificity which allows
to cut DNA at any one (or a few) sites among the whole genome. These endonucleases have been
shown to initiate site-specific genetic recombination in vivo including "intron homing".
Endonucleases in the other class introduce a number of double-stranded breaks in various
double-stranded DNA like bacterial restriction endonucleases. Until now, three endonucleases
were biochemically or genetically identified to belong to the latter class; i.e. Endo.SceI and
Endo.SceII of Saccharomyces cerevisiae, and Endo.SuvI from Saccharomyces uvarum (S.
carlsbergensis). The active form of Endo.SceII is a heterodimer of 75kDa- and 50kDa- subunits
and localized in mitochondria. Endo.SceI and Endo.SuvI contain the 50kDa subunit, and each of
them is a product of the allele of a mitochondrial gene (ENS2). The difference in sequence
specificity between Endo.SceI and Endo.SuvI is caused by the substitution of two amino acids.
The cutting sites of Endo.SceI have complex features and the cutting creates staggered ends
with 4 bases protruding at the 3' ends. These are common featues of all sequence-specific
endonucleases from yeast. Another common feature is the presence of two clusters of partially
conserved 12 amino acid sequences. We looked at events around a genetic marker (Oli') which
was located at ca 200 bps from a cutting site for Endo.SceI in a mitochondrial gene, oli2. The
site was shown to be partially cleaved in vivo in mitotic cells having active Endo.SceI. We
found mating-dependent introduction of a double-stranded break at the cutting site in the oli2
gene in a haploid lacking Endo.ScII upon the mating with a partner having active Endo.SceI. At
the same time, we observed polarized gene conversion at oli2 locus. The disparity in
conversion depends on the presence of active Endo.SceI in a parent and the difference in the
sensitivity of the cutting sites to Endo.SceI between parental strains. Endo.SceI cuts
mitochondrial DNA at more than 30 sites in vitro and we detected in vivo cutting at the
cutting sites for Enco.SceI other than that in oli2 in haploid or diploid cells having active
Endo.SceI. These results suggest that the recombination induced by Endo.SceI is probably
homologous rather than site-specific recombination. The subunit structure and the mechanism to
protect genomic DNA from complete cleavage by the endonuclease are different between Endo.SceI
and endonucleases of the other class. The 75kDa-subunit of Endo.SceI is HSP70, which is
involved in the import of protein into mitochondria. This suggests a regulatory role for the
larger subunit of Endo.SceI in vivo. Although Endo.SceI is dispensable, we found another
site-specific endonuclease from S. cerevisiae lacking a gene for Endo.SceI (Ohta K., Nicolas,
A., Keszenman-Pereyra, D. and T.S.). Thus, mitochondria have multiple species of site-specific
endonucleases and the endonucleases or the recombination induced by them plays a basic role in
this organelle.

<>

<1>Shibata, T., Nakagawa, K.-I., Morishima, N.
<2>Multi-site-specific endonucleases and the initiation of homologous genetic recombination in yeast.
<3>Adv. Biophys.
<4>31
<5>77-91
<6>1995
<7>The notion that homologous recombination is a regulated biological process is not a familiar
one.  In yeasts, homologous recombination and most site-specific ones are initiated by
site-specific double-stranded breaks that are introduced within cis-acting elements for the
recombination.  On the other hand, yeasts have a group of site-specific endonucleases
(multi-site-specific endonucleases) that have a number of cleavage sites on each DNA.  One of
them, Endo.SceI of S. cerevisiae, was shown to introduce double-stranded breaks at a number of
well-defined sites on the mitochondrial DNA in vivo.  An Endo.SceI-induced double-stranded
break was demonstrated to induce homologous recombination in mitochondria.  Like the case of
homologous recombination of nuclear chromosomes, the double-stranded break induces gene
conversion of both genetic markers flanking and in the proximity of the cleavage site, and the
cleaved DNA acts as a recipient of genetic information from the uncleaved partner DNA.  The 70
kDa-heat-shock protein (HSP70)-subunit of Endo.SceI and a general role of the HSP70 in the
regulation of protein-folding suggest the regulation of nucleolytic activity of Endo.SceI and
a general role of the HSP70 in the regulation of protein-folding suggest the regulation of
nucleolytic activity of Endo.SceI.

<>

<1>Shibata, T., Watabe, H., Kaneko, T., Iino, T., Ando, T.
<2>On the nucleotide sequence recognized by a eukaryotic site-specific endonuclease, Endo.SceI from yeast.
<3>J. Biol. Chem.
<4>259
<5>10499-10506
<6>1984
<7>Endo.SceI which is isolated from cells of Saccharomyces cerevisiae is a
eukaryotic site-specific endonuclease active on double-stranded DNA.  At each
cleavage site, Endo.SceI cuts only a defined phosphodiester bond in each strand
of the double helix.  We compared nucleotide sequences around five cleavage
sites for Endo.SceI using a computer.  We could not find any common specific
sequence consisting of five base pairs or more among them.  However, we found a
26-base pair consensus sequence which included 15 conserved nucleotides,
allowing any of the five sequences to include a few nucleotides deviated from
the consensus sequence.  The consensus sequence is
5'-CAn*PYnnAnnCYYGTTnnnPnYnnYA-3', where P, Y, n, and * denote purine,
pyrimidine, any nucleotide, and the center of the cleavage site, respectively.
The numbers of sites at which the consensus sequence appears in pBR322 DNA,
PhiX174 replicative form DNA, fd replicative form DNA, or SV40 DNA are close to
those of the cleavage sites for Endo.SceI.  We found that a 33-base pair
fragment was efficiently cut at the defined phosphodiester bonds by Endo.SceI.
This 33-base pair fragment included 25 base pairs out of the 26-base pair
consensus sequence.  The fragments in which a part of the consensus sequence
was missing were not cut by Endo.SceI.  These observations suggest that the
consensus sequence described above is the major characteristic around the
cleavage sites recognized by Endo.SceI and that the mode of recognition of
cleavage sites by Endo.SceI is different from that by restriction
endonucleases.  We found homology between the consensus sequence for Endo.SceI
and the sequences around the cleavage sites for two other site-specific
endonucleases of S. cerevisiae: Endo.SceII and YZ-Endo which is involved in
mating type switching.

<>

<1>Shibata, T.F., Maeda, T., Nikoh, N., Yamaguchi, K., Oshima, K., Hattori, M., Nishiyama, T., Hasebe, M., Fukatsu, T., Kikuchi, Y., Shigenobu, S.
<2>Complete Genome Sequence of Burkholderia sp. Strain RPE64, Bacterial Symbiont of  the Bean Bug Riptortus pedestris.
<3>Genome Announcements
<4>1
<5>e00441-13
<6>2013
<7>We isolated Burkholderia symbiont strain RPE64 from the bean bug Riptortus pedestris. Analysis
of the complete 6.96-Mb genome, which consists of three
chromosomes and two plasmids, will facilitate further understanding of
insect-microbe symbiosis and the development of pest-control technologies.

<>

<1>Shieh, F.K., Reich, N.O.
<2>AdoMet-dependent Methyl-transfer: Glu(119) Is Essential for DNA C5-Cytosine Methyltransferase M.HhaI.
<3>J. Mol. Biol.
<4>373
<5>1157-1168
<6>2007
<7>The role of Glu119 in S-adenosyl-l-methionine-dependent DNA methyltransferase M.HhaI-catalyzed
DNA methylation was studied. Glu119
belongs to the highly conserved Glu/Asn/Val motif found in all DNA
C5-cytosine methyltransferases, and its importance for M.HhaI function
remains untested. We show that formation of the covalent intermediate
between Cys81 and the target cytosine requires Glu119, since conversion to
Ala, Asp or Gln lowers the rate of methyl transfer 10(2)-10(6) fold.
Further, unlike the wild-type M.HhaI, these mutants are not trapped by the
substrate in which the target cytosine is replaced with the
mechanism-based inhibitor 5-fluorocytosine. The DNA binding affinity for
the Glu119Asp mutant is decreased 10(3)-fold. Thus, the ability of the
enzyme to stabilize the extrahelical cytosine is coupled directly to tight
DNA binding. The structures of the ternary protein/DNA/AdoHcy complexes
for both the Glu119Ala and Glu119Gln mutants (2.70 A and 2.75 A,
respectively) show that the flipped base is positioned nearly identically
with that observed in the wild-type M.HhaI complex. A single water
molecule in the Glu119Ala structure between Ala119 and the extrahelical
cytosine N3 is lacking in the Glu119Gln and wild-type M.HhaI structures,
and most likely accounts for this mutant's partial activity. Glu119 has
essential roles in activating the target cytosine for nucleophilic attack
and contributes to tight DNA binding.

<>

<1>Shieh, F.K., Youngblood, B., Reich, N.O.
<2>The Role of Arg165 Towards Base Flipping, Base Stabilization and Catalysis in M.HhaI.
<3>J. Mol. Biol.
<4>362
<5>516-527
<6>2006
<7>Arg165 forms part of a previously identified base flipping motif in the bacterial DNA cytosine
methyltransferase, M.HhaI. Replacement of Arg165
with Ala has no detectable effect on either DNA or AdoMet affinity, yet
causes the base flipping and restacking transitions to be decreased
approximately 16 and 190-fold respectively, thus confirming the importance
of this motif. However, these kinetic changes cannot account for the
mutant's observed 10(5)-fold decreased catalytic rate. The mutant
enzyme/cognate DNA cocrystal structure (2.79 A resolution) shows the
target cytosine to be positioned approximately 30 degrees into the major
groove, which is consistent with a major groove pathway for nucleotide
flipping. The pyrimidine-sugar chi angle is rotated to approximately +171
degrees , from a range of -95 degrees to -120 degrees in B DNA, and -77
degrees in the WT M.HhaI complex. Thus, Arg165 is important for
maintaining the cytosine positioned for nucleophilic attack by Cys81. The
cytosine sugar pucker is in the C2'-endo-C3'-exo (South conformation), in
contrast to the previously reported C3'-endo (North conformation)
described for the original 2.70 A resolution cocrystal structure of the WT
M.HhaI/DNA complex. We determined a high resolution structure of the WT
M.HhaI/DNA complex (1.96 A) to better determine the sugar pucker. This new
structure is similar to the original, lower resolution WT M.HhaI complex,
but shows that the sugar pucker is O4'-endo (East conformation),
intermediate between the South and North conformers. In summary, Arg165
plays significant roles in base flipping, cytosine positioning, and
catalysis. Furthermore, the previously proposed M.HhaI-mediated changes in
sugar pucker may not be an important contributor to the base flipping
mechanism. These results provide insights into the base flipping and
catalytic mechanisms for bacterial and eukaryotic DNA methyltransferases.

<>

<1>Shields, S.L., Burbank, D.E., Grabherr, R., Van Etten, J.L.
<2>Cloning and sequencing the cytosine methyltransferase gene M. CviJI from Chlorella virus IL-3A.
<3>Virology
<4>176
<5>16-24
<6>1990
<7>The Chlorella virus IL-3A gene encoding the DNA methyltransferase M.CviJI,
which methylates the internal cytosine in (G/A)GC(T/C/G) sequences, was cloned
and expressed in Escherichia coli.  The region containing the M.CviJI gene was
sequenced and a single open reading frame of 1101 bp was identified that could
code for a polypeptide of 367 amino acids with a predicted molecular weight of
41,864.  M.CviJI contained regions of amino acids which were similar to
bacterial cytosine methyltransferases.  Eighteen other Chlorella viruses, of 36
tested, contained DNA sequences which hybridized to the M.CviJI gene; DNA from
some, but not all, of these 18 viruses also contained 5-methylcytosine in
(G/A)GC(T/C/G) sequences.

<>

<1>Shields, S.L., Burbank, D.E., Van Etten, J.L.
<2>Molecular cloning and characterization of the gene encoding the cytosine methyltransferase.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>214
<6>1989
<7>The gene encoding the DNA methyltransferase, M.CviJI from Chlorella virus IL-3A
was cloned in pUC19 and expressed in E. coli RR1.  Recombinant plasmid
pIL-3A.22.8 containing a 3.6 kbp KpnI/Sau3A restriction fragment encoding
M.CviJI methylates the internal cytosine in PuGCPy sequences or a subset of
PuGCPy sequences in vivo.  Methylation of these sequences by M.CviJI prevents
digestion of pIL-3A.22.8 by restriction endonucleases sensitive to cytosine
methylation in PuGCPy recognition sequences.  Transposon Tn5 mutagenesis
localized the M.CviJI functional domain on pIL-3A.22.8.  Restriction fragments
from the HpaI restriction site, 185 bp from the termini of the terminal repeat
of Tn5, to the SalI and KpnI restriction site of pIL-3A.22.8 polylinker were
isolated from individual Tn5 mutants.  These restriction fragments containing
the functional domain and flanking sequences were subcloned in single stranded
sequencing vectors M13mp18 and M13mp19 for nucleotide sequencing.  The amino
acid sequence deduced from M.CviJI nucleotide sequence was compared to the
amino acid sequence of 5-methylcytosine methyltransferases from prokaryotes to
determine potentially shared protein domains.  The M.CviJI gene was not
essential for IL-3A replication since a M.CviJI deletion mutant also replicated
in Chlorella.

<>

<1>Shields-Menard, S.A., Brown, S.D., Klingeman, D.M., Indest, K., Hancock, D., Wewalwela, J.J., French, W.T., Donaldson, J.R.
<2>Draft Genome Sequence of Rhodococcus rhodochrous Strain ATCC 21198.
<3>Genome Announcements
<4>2
<5>e00054-14
<6>2014
<7>Rhodococcus rhodochrous is a Gram-positive red-pigmented bacterium commonly found in the soil.
The draft genome sequence for R.Â rhodochrous strain ATCC 21198 is presented here to provide
genetic data for a better understanding of its lipid-accumulating capabilities.

<>

<1>Shier, V.K.
<2>Characterization and inhibition of an essential adenine DNA methyltransferase from Caulobacter crescentus.
<3>Diss. Abstr.
<4>62
<5>3185-3186
<6>2002
<7>Genetic evidence has established that DNA methylation provides an obligatory signal for the
proper progression through the Caulobacter crescentus cell cycle.  The enzyme responsible for
catalyzing this reaction is a cell-cycle regulated adenine DNA methyltransferase (CcrM).  CcrM
activity is tightly restricted to predivisional cells by a delicate coordination between
transcription and proteolytic elimination by a Lon-mediated pathway.  Further underscoring the
importance of this enzyme is the observation that CcrM is essential for C. crescentus
viability and that it is only the second known DNA methyltransferase that lacks a cognate
restriction endonuclease.  To further our understanding of the regulatory role played by CcrM,
we sought to investigate the biophysical and biochemical properties of this enzyme.
Equilibrium ultracentrifugation, velocity ultracentrifugation, and chemical crosslinking
reveal that CcrM is dimeric at physiological concentrations.  However, surface plasmon
resonance experiments evince that CcrM binds as a monomer to DNA containing the CcrM canonical
methylation sequence, GANTC.  Collectively, these findings suggest that CcrM dimerization
serves to stabilize CcrM from premature in vivo proteolysis.  The recently sequenced C.
crescentus genome unveiled the presence of 4,496 CcrM methylation sites.  Following
semiconservative DNA replication, CcrM must methylate a total of 8,992 sites on the two
resulting copies of the genome within the strict temporal constraints of the predivisional
cells.  Using defined, synthetic substrates that contain multiple methylation sites, CcrM
processively methylates DNA over a minimum distance of 82 base pairs.  Moreover, CcrM is
capable of methylating GANTC clusters with no apparent effect on methylation efficiency.
Despite slow kinetic rate constants, these experiments demonstrate that CcrM is kinetically
competent to support full genomic methylation within the enforced activity interval.
Homologues of CcrM are found throughout the alpha-subdivision of proteobacteria.  Many of the
members of this group are pathogens.  The observation that CcrM activity is essential for
cellular viability in all of these microorganisms, coupled with the absence of adenine methyl
modification in eukaryotic species, has implicated CcrM as a target for antimicrobial drug
design.  Efforts to identify and characterize potential inhibitors are discussed with an
emphasis on a panel of diphenyl borinic esters.

<>

<1>Shier, V.K., Hancey, C.J., Benkovic, S.J.
<2>Identification of the active oligomeric state of an essential adenine DNA methyltransferase from Caulobacter crescentus.
<3>J. Biol. Chem.
<4>276
<5>14744-14751
<6>2001
<7>Caulobacter crescentus contains one of the two known prokaryotic DNA methyltransferases that
lacks a cognate endonuclease. This endogenous cell cycle regulated adenine DNA
methyltransferase (CcrM) is essential for C. crescentus cellular viability. DNA methylation
catalyzed by CcrM provides an obligatory signal for the proper progression through the cell
cycle. To further our understanding of the regulatory role played by CcrM, we sought to
investigate its biophysical properties. In this paper we employed equilibrium
ultracentrifugation, velocity ultracentrifugation, and chemical cross-linking to show that
CcrM is dimeric at physiological concentrations. However, surface plasmon resonance
experiments in the presence of S-adenosyl-homocysteine evince that CcrM binds as a monomer to
a defined hemi-methylated DNA substrate containing the canonical methylation sequence, GANTC.
Initial velocity experiments demonstrate that dimerization of CcrM does not affect DNA
methylation. Collectively, these findings suggest that CcrM is active as a monomer and
provides a possible in vivo role for dimerization as a means to stabilize CcrM from premature
catabolism.

<>

<1>Shigdel, U.K.
<2>Developing DNA probes to trap DNA cytosine-5 methyltransferases involved in promoter hypermethylation in cancer.
<3>Ph.D. Thesis, Univ. of Chicago
<4>
<5>
<6>2009
<7>A major limitation to making significant advances in diagnosing and treating
cancer is that we do not have a thorough understanding of the mechanisms leading to
abnormal DNA methylation in cancer cells. Although there is a plethora of information
on genes that are hypermethylated in cancer cells, we rarely know which DNA cytosine-5
methyltransferase (DNMTs) and what complex is involved in methylating particular
promoters in cancer cells. I developed two types of DNA probes, a diazirine and a
disulfide based, which can be incorporated at specific positions of oligonucleotides that
have potential to trap DNMTs and their cofactors at a particular promoter. A diazirine
photophore was introduced into either the major or minor groove of DNA via a
convertible nucleoside methodology. The resulting DNA probes efficiently cross-linked
with two different proteins studied as examples, the E. coli DNA adenine
methyltransferase (EcoDam) and the human 06-alkylguanine-DNA alkyltransferase
(hAGT). Efficient cross-linking of diazirine can be utilized to trap proteins from cell
extracts. Taking advantage of DNMTs' invariant active site Cys and their base flipping
property, a new disulfide-based DNA probe, l'-methylenedisulfide deoxyribose, which
can efficiently cross-link Haemophilus haemolyticus methylase (M. Hhal) and
Spiroplasma sp. Methylase (M. SssI) was also developed. Compared to commercially
available 5-FdC, the new probe cross-links DNMTs quickly. Using a disulfide tether on
the N4 position of cytosine, catalytic domain of DNMT3A (DNMT3 AC) has been crosslinked
to DNA in high efficiency. Such a high cross-linking yield of DNMT3AC holds
great promise in identifying DNMTs and their cofactors that act on particular promoters,
as well as in structurally characterizing the DNMT3 AC-DNA complex.

<>

<1>Shigdel, U.K., He, C.
<2>A New 1'-Methylenedisulfide Deoxyribose that Forms an Efficient Cross-Link to DNA Cytosine-5 Methyltransferase (DNMT).
<3>J. Am. Chem. Soc.
<4>130
<5>17634-17635
<6>2008
<7>Although only four bases, adenine, guanine, cytosine, and thymine,
encode all genetic information in DNA, there is a heritable "fifth" base,
5-methylcytosine, which can induce epigenetic changes that alter
chromatin structures. Methylation at the 5-position of cytosine in a
CpG dinucleotide is catalyzed by a conserved group of proteins called
DNA cytosine-5 methyltransferases (DNMTs) by using S-adenosylmethionine
(SAM) as the cofactor (Figure 1A). This modification has
a profound effect on gene expression. Many cancer cells are characterized
by abnormal DNA methylation. Repetitive DNA sequences and
some genes are hypomethylated and transcriptionally active, whereas
many genes are hypermethylated and transcriptionally inactive.1
Inhibition of human DNMTs has been shown to be an effective strategy
to treat various cancers.

<>

<1>Shigemoto, N., Kuwahara, R., Kayama, S., Shimizu, W., Onodera, M., Yokozaki, M., Hisatsune, J., Kato, F., Ohge, H., Sugai, M.
<2>Emergence in Japan of an imipenem-susceptible, meropenem-resistant Klebsiella pneumoniae carrying blaIMP-6.
<3>Diagn. Microbiol. Infect. Dis.
<4>72
<5>109-112
<6>2012
<7>We identified 5 Klebsiella pneumoniae isolates showing high resistance to
beta-lactams except imipenem and designated them ISMRK (imipenem-susceptible but
meropenem-resistant Klebsiella). They carried the bla(IMP-6) and bla(CTX-M-2) on
a self-transmissible plasmid. ISMRK may be falsely categorized as susceptible to
carbapenems if imipenem is used to screen carbapenem resistance.

<>

<1>Shigenobu, S., Watanabe, H., Hattori, M., Sakaki, Y., Ishikawa, H.
<2>Genome sequence of the endocellular bacterial symbiont of aphids Buchnera sp. APS.
<3>Nature
<4>407
<5>81-86
<6>2000
<7>Almost all aphid species (Homoptera, Insecta) have 60-80 huge cells called bacteriocytes,
within which are round-shaped bacteria that are designated Buchnera. These bacteria are
maternally transmitted to eggs and embryos through host generations, and the mutualism between
the host and the bacteria is so obligate that neither can reproduce independently. Buchnera is
a close relative of Escherichia coli, but it contains more than 100 genomic copies per cell,
and its genome size is only a seventh of that of E. coli. Here we report the complete genome
sequence of Buchnera sp. strain APS, which is composed of one 640,681-base-pair chromosome and
two small plasmids. There are genes for the biosyntheses of amino acids essential for the
hosts in the genome, but those for non-essential amino acids are missing, indicating
complementarity and syntrophy between the host and the symbiont. In addition, Buchnera lacks
genes for the biosynthesis of cell-surface components, including lipopolysaccharides and
phospholipids, regulator genes and genes involved in defense of the cell. These results
indicate that Buchnera is completely symbiotic and viable only in its limited niche, the
bacteriocyte.

<>

<1>Shikara, M.
<2>Identification of a restriction endonuclease (SacC1) from Saccharomyces cerevisiae.
<3>J. Yeast Fungal Res.
<4>1
<5>127-135
<6>2010
<7>palindromic sequence 5'CTCGAC3' cleaving both DNA strands upstream and downstream of its
recognition sequence and makes a staggered cut at the distance of five bases from the
recognition sequence on the upper strand and at the seventh base on the complementary strand.
It shares similar
characteristics with Sac I from Streptomyces achromogenes as well as Sst1 from Streptomyces
Stanford and Psp124B1 from Pseudomonas species. It has been purified by ammonium sulphate
precipitation, dialysis, and gel filtration using phosphocellulose, DEAE-cellulose and
Sephadex G-100
with an optimal pH range (7.5-8.5), active at 37°C and dependent on Mg+2 or Mn2+ which
increases its activity by 4- and 2-folds, respectively, while other cations decrease its
activity to some extents.  Cleavage on both sides of the recognition sequence is
characteristic of Type IIB systems but all IIB
enzymes studied so far have been found to recognize discontinuous sites and a distinctive
subunit/domain organization that is not present in the SacC1 enzyme. There are similarities
between SacC1 and other homing endonucleases belonging to the LAGLIDADG family such as a
requirement for Mg2+ (or Mn2+) for cleavage to take place, optimal activity at alkaline pH and
stimulation of the reaction by moderate concentrations of the monovalent cation.

<>

<1>Shikara, M.
<2>A specific inhibitory protein to a restriction enzyme from Saccharomyces cerevisiae.
<3>J. Yeast Fungal Res.
<4>1
<5>174-182
<6>2010
<7>A specific protein inhibitor for the restriction enzyme (SacC1) has been purified from
Saccharomyces
cerevisiae approximately 21,000 fold and its inhibitory properties have been characterized.
The
isoelectric points (pI) of SacCI and its inhibitor are 9.0 and 5.22, respectively. The
molecular weight of
SacC1, the inhibitor and SacC1-inhibitor complex were estimated by gel filtration on a
Sephadex G-100
column to be 64,000, 32,000 and 85,000, respectively. The inhibitor protein inhibits SacC1
catalytic
activities efficiently, but has no effect on other restriction enzymes tested. Inhibition does
not occur
unless SacC1 enzyme is exposed to the inhibitor protein prior to the reaction of the enzyme
with DNA.
The inhibitory activity is independent of temperature. The inhibition increased linearly with
the addition
of inhibitor to various amounts of SacC1, up to 85% inhibition. The slope of inhibition was
constant
irrespective of the initial amount of SacC1 and Ki value of 3.45 x 10-12 was obtained. The
inhibitor
interacts strongly with SacC1 and this interaction could increase the stability of the
complex, possibly
manifesting itself as SacC1 decreases in the dissociation rate due to the electrostatic
attraction between
the two groups or the stability may increase by potentially stronger electrostatic
interaction. The
conformational specificity between SacC1 and its inhibitor seems to be essential for their
interaction.
The extremely strong affinity of the inhibitor to SacC1 is remarkable and stronger than the
affinity of
several restriction enzymes.

<>

<1>Shilov, I., Tashlitsky, V., Khodoun, M., Vasilev, S., Alekseev, Y., Kuzubov, A., Kubareva, E., Karyagina, A.
<2>DNA-methyltransferase SsoII interaction with own promoter region binding site.
<3>Nucleic Acids Res.
<4>26
<5>2659-2664
<6>1998
<7>The investigation of SsoII DNA-methyltransferase interaction with the intergenic region of
SsoII restriction-modification system was carried out.  Seven guanine residues protected by
M.SsoII from methylation with dimethylsulfate and thus probably involved in enzyme-DNA
recognition were identified.  Six of them are located symmetrically within the 15 bp inverted
repeat inside the SsoII promoter region.  The crosslinking of SsoII methyltransferase with DNA
duplexes containing 5-bromo-2'-deoxyuridine instead of thymidine was performed.  The
crosslinked products were obtained in all cases, thus proving that tested thymines were in
proximity with enzyme.  The ability to produce the crosslinked products in one case was
2-5-fold higher than in other ones.  This allowed us to imply that the thymine residue in this
position of the inverted repeat could be in contact with M.SsoII.  Based on the experimental
data, two symmetrical 4 bp clusters (GGAC), which could be involved in the interaction with
M.SsoII in the DNA-protein complex, were identified.  The model of M.SsoII interaction with
its own promoter was proposed.

<>

<1>Shimado, H., Ishino, Y., Katou, I.
<2>BalI restriction;modification-based enzyme gene.
<3>Japanese Patent Office
<4>JP 1996205870 A
<5>
<6>1996
<7>
<>

<1>Shimado, H., Ishino, Y., Katou, I.
<2>BalI restriction/modification-based enzyme gene.
<3>Japanese Patent Office
<4>JP 8205870
<5>
<6>1996
<7>Purpose: To obtain the subject gene useful to mass-produce the BalI restriction enzyme useful
as a genetically engineered reagent.  Constitution: The objective BalI restiction enzyme gene
and BalI modification enzyme gene are obtainable from Brevibacterium albidum ATCC15831, having
e.g. amino acid sequences of formula I and formula II, respectively.  The BalI restriction
enzyme can be obtained in large quantities from a cultured product of transformant Escherichia
coli ER1648/pBSR1/pSRB8 (FERM P-14625) produced by transfecting a vector pSRB8 formed by
modifying the initiation codon of DRF2 into ATG and binding the BalI restriction enzyme gene
on a downstream site of lac promoter and a second vector pBSR1 capable of manifesting BalI
modification enzyme into E. coli ER1648 strain.

<>

<1>Shimizu, T., Ohtani, K., Hirakawa, H., Ohshima, K., Yamashita, A., Shiba, T., Ogasawara, N., Hattori, M., Kuhara, S., Hayashi, H.
<2>Complete genome sequence of Clostridium perfringens, an anaerobic flesh-eater.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>996-1001
<6>2002
<7>Clostridium perfringens is a Gram-positive anaerobic spore-forming bacterium that causes
life-threatening gas gangrene and mild
enterotoxaemia in humans, although it colonizes as normal intestinal flora of humans and
animals. The organism is known to produce a
variety of toxins and enzymes that are responsible for the severe myonecrotic lesions. Here we
report the complete 3,031,430-bp sequence
of C. perfringens strain 13 that comprises 2,660 protein coding regions and 10 rRNA genes,
showing pronounced low overall G + C
content (28.6%). The genome contains typical anaerobic fermentation enzymes leading to gas
production but no enzymes for the
tricarboxylic acid cycle or respiratory chain. Various saccharolytic enzymes were found, but
many enzymes for amino acid biosynthesis were
lacking in the genome. Twenty genes were newly identified as putative virulence factors of C.
perfringens, and we found a total of five
hyaluronidase genes that will also contribute to virulence. The genome analysis also proved an
efficient method for finding four members of
the two-component VirR/VirS regulon that coordinately regulates the pathogenicity of C.
perfringens. Clearly, C. perfringens obtains
various essential materials from the host by producing several degradative enzymes and toxins,
resulting in massive destruction of the host
tissues.

<>

<1>Shimizu-Kadota, M., Kato, H., Shiwa, Y., Oshima, K., Machii, M., Araya-Kojima, T., Zendo, T., Hattori, M., Sonomoto, K., Yoshikawa, H.
<2>Genomic Features of Lactococcus lactis IO-1, a Lactic Acid Bacterium That Utilizes Xylose and Produces High Levels of L-Lactic Acid.
<3>Biosci. Biotechnol. Biochem.
<4>77
<5>1804-1808
<6>2013
<7>Lactococcus lactis IO-1 (JCM7638) produces L-lactic acid predominantly when grown at high
xylose concentrations, and its utilization is highly desired in the green plastics industry.
Therefore it is worthwhile studying its genomic traits. In this study, we focused on (i) genes
of possible horizontal transfer derivation (prophages, the nisin-sucrose transposon, and
several restriction-modification systems), and (ii) genes for the synthetic pathways of amino
acids and vitamins in the IO-1 genome. In v ew of the results of this analysis, we consider
their meanings in strain IO-1.

<>

<1>Shimizu-Kadota, M., Shibahara-Sone, H.
<2>M.HhaI-methylated DNA is resistant to cleavage by R.BbeI.
<3>Agric. Biol. Chem.
<4>53
<5>2841-2842
<6>1989
<7>Shows that BbeI can cleave GGCG5mCC, GGCGC5mC, but not GG5mCGCC.

<>

<1>Shimoda, N., Yamakoshi, K., Miyake, A., Takeda, H.
<2>Identification of a gene required for de novo DNA methylation of the zebrafish no tail gene.
<3>Dev. Dyn.
<4>233
<5>1509-1516
<6>2005
<7>The zebrafish no tail gene (ntl) is indispensable for tail and notochord development. We have
shown previously that ntl is de novo methylated
during early embryogenesis. To find the gene that de novo methylates ntl
and understand the meaning of this methylation, we cloned seven genes that
encode the conserved catalytic domain of methyltransferases. We found that
injection of antisense morpholino oligonucleotides against one of them,
termed dnmt7, into eggs significantly reduced the level of ntl
methylation, although no apparent phenotype was induced by the injection.
Inhibition of Dnmt7 activity did not change the level of genome-wide
methylation nor did it affect de novo methylation of injected plasmid DNA,
indicating that Dnmt7 specifically methylates ntl in the genome.

<>

<1>Shimotsu, H., Takahashi, H., Saito, H.
<2>Site-specific endonucleases in Streptomyces strains.
<3>Agric. Biol. Chem.
<4>44
<5>1665-1666
<6>1980
<7>A large number of class II restriction endonucleases have now been isolated
from various microorganisms.  Since their discovery they have become invaluable
tools in researches of physical mapping DNA sequencing and gene cloning.  Only
four Streptomyces strains have been reported to produce restriction-like
endonucleases; Streptomyces achromogenes (SacI, SacII, and SacIII), S. albus G
(SalI and SalII), and S. lavendulae (SlaI).  The first two strains produce
exactly identical enzymes.  In this communication, we report the results of
survey of restriction-like endonucleases among Streptomyces strains.

<>

<1>Shimotsu, H., Takahashi, H., Saito, H.
<2>A new site-specific endonuclease StuI from Streptomyces tubercidicus.
<3>Gene
<4>11
<5>219-225
<6>1980
<7>A new sequence-specific endonuclease, StuI, produced by Streptomyces
tubercidicus KCC S-0054, was identified and partially purified.  StuI
recognizes the hexanucleotide "palindromic" sequence:
5'-AGG^CCT-3'
3'-TCC^GGA-5'  and cleaves it at the middle, producing blunt ends.

<>

<1>Shin, H., Lee, J.H., Ahn, C.S., Ryu, S., Cho, B.C.
<2>Complete genome sequence of marine bacterium Pseudoalteromonas phenolica bacteriophage TW1.
<3>Arch. Virol.
<4>159
<5>159-162
<6>2014
<7>For molecular study of marine bacteria Pseudoalteromonas phenolica using
bacteriophage, a novel bacteriophage, TW1, belonging to the family Siphoviridae,
was isolated, and its genome was completely sequenced and analyzed. The phage TW1
genome consists of 39,940-bp-length double-stranded DNA with a GC content of
40.19 %, and it was predicted to have 62 open reading frames (ORFs), which were
classified into functional groups, including phage structure, packaging, DNA
metabolism, regulation, and additional function. The phage life style prediction
using PHACTS showed that it may be a temperate phage. However, genes related to
lysogeny and host lysis were not detected in the phage TW1 genome, indicating
that annotation information about P. phenolica phages in the genome databases may
not be sufficient for the functional prediction of their encoded proteins. This
is the first report of a P. phenolica-infecting phage, and this phage genome
study will provide useful information for further molecular research on P.
phenolica and its phage, as well as their interactions.

<>

<1>Shin, H., Lee, J.H., Choi, Y., Ryu, S.
<2>Complete Genome Sequence of the Opportunistic Food-Borne Pathogen Cronobacter sakazakii ES15.
<3>J. Bacteriol.
<4>194
<5>4438-4439
<6>2012
<7>Cronobacter sakazakii is an emerging pathogen associated with several outbreaks of food-borne
illness in premature infants. To characterize its physiology and
pathogenicity at the molecular level, C. sakazakii ES15 was isolated and its
genome was completely sequenced and analyzed. Here, the results are announced and
major findings from its annotation data are reported.

<>

<1>Shin, H., Lee, J.H., Kim, Y., Ryu, S.
<2>Complete Genome Sequence of Cronobacter sakazakii Bacteriophage CR3.
<3>J. Virol.
<4>86
<5>6367-6368
<6>2012
<7>Due to the high risk of Cronobacter sakazakii infection in infants fed powdered
milk formula and the emergence of antibiotic-resistant strains, an alternative
biocontrol agent using bacteriophage is needed to control this pathogen. To
further the development of such an agent, the C. sakazakii-targeting
bacteriophage CR3 was isolated and its genome was completely sequenced. Here, we
announce the genomic analysis results of the largest C. sakazakii phage known to
date and report the major findings from the genome annotation.

<>

<1>Shin, H., Lee, J.H., Lim, J.A., Kim, H., Ryu, S.
<2>Complete Genome Sequence of Salmonella enterica Serovar Typhimurium Bacteriophage SPN1S.
<3>J. Virol.
<4>86
<5>1284-1285
<6>2012
<7>To understand the interaction between the host of pathogenic Salmonella enterica
serovar Typhimurium and its bacteriophage, we isolated the bacteriophage SPN1S.
It is a lysogenic phage in the Podoviridae family and uses the O-antigen of
lipopolysaccharides (LPS) as a host receptor. Comparative genomic analysis of
phage SPN1S and the S. enterica serovar Anatum-specific phage epsilon15 revealed
different host specificities, probably due to the low homology of host
specificity-related genes. Here we report the complete circular genome sequence
of S. Typhimurium-specific bacteriophage SPN1S and show the results of our
analysis.

<>

<1>Shin, J., Soo-Ko, K.
<2>Single origin of three plasmids bearing blaCTX-M-15 from different Klebsiella pneumoniae clones.
<3>J. Antimicrob. Chemother.
<4>69
<5>969-972
<6>2013
<7>OBJECTIVES: To determine and compare the complete nucleotide sequences of plasmids carrying
blaCTX-M-15 from three different Klebsiella pneumoniae clones.
METHODS: IncFII-type plasmids pKP02022, pKP09085 and pKP007 were extracted from three K.
pneumoniae strains. These strains belong to sequence types (STs) ST15,
ST48 and ST23, respectively, and were isolated in Korea. Plasmids were sequenced using the 454
Genome Sequencer FLX system. RESULTS: The three plasmids, pKP02022
(203577 bp), pKP09085 (213019 bp) and pKP007 (246 176 bp), all exhibited a very similar
structure, with a pKPN3-like backbone and a resistance region including blaOXA-1,
aac(6')-Ib-cr and cat genes as well as blaCTX-M-15. They were also very similar to pUUH239.2,
previously isolated in Sweden. Iron (III) uptake-related genes were found in pKP007 from the
ST23 strain, which has been reported to be associated with liver abscesses. The resistance
region contained several insertion sequences, such as IS26, which may play an important role
in structural rearrangements of plasmids. CONCLUSIONS: The very similar structure of the three
plasmids, extracted from different clones, suggests that the spread of CTX-M-producing K.
pneumoniae isolates might result from the horizontal transfer of plasmids and subsequent
integration and recombination.

<>

<1>Shin, N.R., Whon, T.W., Roh, S.W., Kim, M.S., Jung, M.J., Lee, J., Bae, J.W.
<2>Genome Sequence of Corynebacterium nuruki S6-4T, Isolated from Alcohol Fermentation Starter.
<3>J. Bacteriol.
<4>193
<5>4257
<6>2011
<7>Corynebacterium nuruki S6-4(T), isolated from Korean alcohol fermentation starter, is a
strictly aerobic, nonmotile, Gram-positive, and rod-shaped
bacterium belonging to the genus Corynebacterium and the actinomycete
group. We report here the draft genome sequence of C. nuruki strain
S6-4(T) (3,106,595 bp, with a G+C content of 69.5%).

<>

<1>Shin, S.C., Ahn, D.H., Lee, J.K., Kim, S.J., Hong, S.G., Kim, E.H., Park, H.
<2>Genome Sequence of Sphingomonas sp. Strain PAMC 26605, Isolated from Arctic Lichen (Ochrolechia sp.).
<3>J. Bacteriol.
<4>194
<5>1607
<6>2012
<7>The endosymbiotic bacterium Sphingomonas sp. strain PAMC 26605 was isolated from  Arctic
lichens (Ochrolechia sp.) on the Svalbard Islands. Here we report the
draft genome sequence of this strain, which could provide further insights into
the symbiotic mechanism of lichens in extreme environments.

<>

<1>Shin, S.C., Kim, S.J., Ahn, D.H., Lee, J.K., Lee, H., Lee, J., Hong, S.G., Lee, Y.M., Park, H.
<2>Genome Sequence of a Salinibacterium sp. Isolated from Antarctic Soil.
<3>J. Bacteriol.
<4>194
<5>2404
<6>2012
<7>The draft genome of Salinibacterium sp. PAMC 21357, isolated from permafrost soil of
Antarctica, was determined. Here we present a 3.1-Mb draft genome sequence of
Salinibacterium sp. that could provide further insight into the genetic
determination of its cold-adaptive properties.

<>

<1>Shin, S.C., Kim, S.J., Ahn-do, H., Lee, J.K., Park, H.
<2>Draft Genome Sequence of Sphingomonas echinoides ATCC 14820.
<3>J. Bacteriol.
<4>194
<5>1843
<6>2012
<7>Sphingomonas is a Gram-negative, yellow-pigmented, chemoheterotrophic, strictly aerobic
bacterium. The bacterium is known to be metabolically versatile and can
utilize a wide range of natural compounds as well as some types of environmental
contaminants, such as creosote, polychlorinated biphenyls, etc. Here, we report
the draft genome sequence of Sphingomonas echinoides ATCC 14820, which will
provide additional information to enhance our understanding of metabolic
versatility of Sphingomonas.

<>

<1>Shin, S.C., Kim, S.J., Hong, S.G., Ahn-do, H., Lee, Y.M., Lee, H., Lee, J., Park, H.
<2>Genome Sequence of Pseudomonas sp. Strain PAMC 25886, Isolated from Alpine Glacial Cryoconite.
<3>J. Bacteriol.
<4>194
<5>1844
<6>2012
<7>Pseudomonas spp. have shown characteristics of efficiently metabolizing environmental
pollutants and also producing exopolysaccharides known as biofilms.
Here we present the draft genome sequence of Pseudomonas sp. strain PAMC 25886,
which was isolated from glacier cryoconite in the Alps mountain permafrost region
and which may provide further insight into biodegradative and/or
biofilm-producing mechanisms in a cold environment.

<>

<1>Shin, S.H., Kim, S., Kim, J.Y., Lee, S., Um, Y., Oh, M.K., Kim, Y.R., Lee, J., Yang, K.S.
<2>Complete Genome Sequence of Enterobacter aerogenes KCTC 2190.
<3>J. Bacteriol.
<4>194
<5>2373-2374
<6>2012
<7>This is the first complete genome sequence of the Enterobacter aerogenes species. Here we
present the genome sequence of E. aerogenes KCTC 2190, which contains
5,280,350 bp with a G + C content of 54.8 mol%, 4,912 protein-coding genes, and
109 structural RNAs.

<>

<1>Shin, S.H., Kim, S., Kim, J.Y., Lee, S., Um, Y., Oh, M.K., Kim, Y.R., Lee, J., Yang, K.S.
<2>Complete Genome Sequence of the 2,3-Butanediol-Producing Klebsiella pneumoniae Strain KCTC 2242.
<3>J. Bacteriol.
<4>194
<5>2736-2737
<6>2012
<7>Here we report the full genome sequence of Klebsiella pneumoniae KCTC 2242,consisting of a
5.26-Mb chromosome (57.6% GC%; 5,035 genes [4,923 encoding
known proteins, 112 RNA genes]) and a 202-kb plasmid (50.2% GC%; 229 genes [229
encoding known proteins]).

<>

<1>Shin, S.H., Kim, S., Kim, J.Y., Lee, S., Um, Y., Oh, M.K., Kim, Y.R., Lee, J., Yang, K.S.
<2>Complete Genome Sequence of Klebsiella oxytoca KCTC 1686, Used in Production of 2,3-Butanediol.
<3>J. Bacteriol.
<4>194
<5>2371-2372
<6>2012
<7>Here we report the full genome sequence of Klebsiella oxytoca KCTC 1686, which is used in
production of 2,3-butanediol. The KCTC 1686 strain contains 5,974,109 bp
with G+C content of 56.05 mol% and contains 5,488 protein-coding genes and 110
structural RNAs.

<>

<1>Shin, S.H., Kim, S., Kim, J.Y., Song, H.Y., Cho, S.J., Kim, D.R., Lee, K.I., Lim, H.K., Park, N.J., Hwang, I.T., Yang, K.S.
<2>Genome Sequence of Paenibacillus terrae HPL-003, a Xylanase-Producing Bacterium Isolated from Soil Found in Forest Residue.
<3>J. Bacteriol.
<4>194
<5>1266
<6>2012
<7>This article reports on the full genome sequence of Paenibacillus terrae HPL-003, which is a
Gram-positive, endospore-forming, xylanase-producing bacterium
isolated from soil found in forest residue on Gara Mountain. The strain HPL-003
contains 6,083,395 bp with a G+C content of 46.77 mol%, 2,633 protein-coding
genes, and 117 structural RNAs.

<>

<1>Shin, S.H., Um, Y., Beak, J.H., Kim, S., Lee, S., Oh, M.K., Kim, Y.R., Lee, J., Yang, K.S.
<2>Complete Genome Sequence of Raoultella ornithinolytica Strain B6, a 2,3-Butanediol-Producing Bacterium Isolated from Oil-Contaminated Soil.
<3>Genome Announcements
<4>1
<5>e00395-13
<6>2013
<7>Here we report the full genome sequence of Raoultella ornithinolytica strain B6,  a
Gram-negative aerobic bacillus belonging to the family Enterobacteriaceae. This
2,3-butanediol-producing bacterium was isolated from oil-contaminated soil on
Backwoon Mountain in South Korea. Strain B6 contains 5,398,151 bp with 4,909
protein-coding genes, 104 structural RNAs, and 55.88% G+C content.

<>

<1>Shinomiya, T., Kobayashi, M., Sato, S.
<2>A second site specific endonuclease from Thermus thermophilus 111, Tth111II.
<3>Nucleic Acids Res.
<4>8
<5>3275-3285
<6>1980
<7>A second site specific endonuclease with novel specificity has been purified
from Thermus thermophilus strain 111 and named Tth111II.  the enzyme is active
at temperature up to 80C and requires Mg2+ for endonuclease activity.  Tth111II
cleaves PhiX174 RFDNA into 11 fragments and lambda DNA into more than 25
fragments.  From the 5' -terminal sequences of Tth111II fragments of PhiX174
RFDNA determined by the two dimensional homochromatography and the survey on
nucleotide sequence PhiX174 RFDNA, it was concluded that Tth111II recognizes
the DNA sequence 5'CAAPuCA(N)11 ^ 3' 3'GTTPyGT(N)9 ^ 5' and cleaves the sites
as indicated by the arrows.

<>

<1>Shinomiya, T., Kobayashi, M., Sato, S.
<2>A second site specific endonuclease from Thermus thermophilus 111, Tth111II.
<3>Nucleic Acids Symp. Ser.
<4>SS8
<5>S181-S184
<6>1980
<7>A second site specific endonuclease with a novel specificity has been isolated
from Thermus thermophilus strain 111 and named Tth111II.  The enzyme is active
at temperature up to 80C and requires Mg2+ or Mn2+ for activity.  Tth111II
cleaves PhiX174 RF into 11 fragments.  From the analysis of 5' terminal
sequences of the PhiX 174 RF DNA fragments produced by Tth111II action, it was
concluded that Tth111II recognized the DNA sequence 5'CAAPuCA(N)11^3'
3'GTTPyGT(N)9^5' and cleaved the sites as indicated by arrows.

<>

<1>Shinomiya, T., Kobayashi, M., Sato, S.
<2>Restriction endonuclease preparation from Thermus thermophilus culture useful in cleaving double helix DNA sequence.
<3>Japanese Patent Office
<4>JP 57012994
<5>
<6>1982
<7>A restriction endonuclease (II) having the following properties is new: (I) cleaves a double
stranded helical DNA at the position shown below. CAAPuCA(N)9NN^ GTTPyGT(N)9^ The two chains
shown should be complementary, (2) optimal pH 7-8 (3) stable pH-range 5-9. (II) is expected to
be useful as a reagent for investigation about genetic technology and primary structure of
DNA. (I) is prepared from the ground mycelium of Thermus thermophilus III. The microorganism
is incubated in a culture medium containing such carbon source as glucose, galactose, maltose,
etc. NaCl, vitamins, etc. The obtained culture medium is ground in beta-mercaptoethanol and
then solids in the medium are separated by centrifugation followed by collection of (I) by
diethylamino ethyl cellulose chromatography, affinity-chromatograph, etc. ph, etc.

<>

<1>Shinomiya, T., Kobayashi, M., Sato, S., Uchida, T.
<2>A new aspect of a restriction endonuclease Tth111I.  It has a degenerated specificity (Tth111I*).
<3>J. Biochem. (Tokyo)
<4>92
<5>1823-1832
<6>1982
<7>We previously reported that Thermus thermophilus 111 contained two restriction
enzymes, Tth111I and Tth111II.  The former does not cleave PhiX174 RFDNA and
the latter does.  We have now found another endonuclease activity able to
cleave PhiX174 RFDNA in the cell extract of T. thermophilus 111.

<>

<1>Shinomiya, T., Sato, S.
<2>A site specific endonuclease from Thermus thermophilus 111, Tth111I.
<3>Nucleic Acids Res.
<4>8
<5>43-56
<6>1980
<7>A site specific endonuclease with novel specificity has been isolated from
Thermus thermophilus strain 111 and named Tth111I.  Tth111I cleaves lambda DNA
into three fragments of 23.5, 25.7 and 50.8% of the total length, and ColE1 DNA
into two fragments of nearly equal length.  The sequences around Tth111I
cleavage sites of ColE1 and lambda DNA were determined by the Maxam and Gilbert
method and the two dimensional mapping method.  The results suggest that
Tth111I recognizes the DNA sequence 5'-TGACN^NNGTC-3' 3'-ACTGNN^NCAG-5' site as
indicated by the arrows.  Assuming that the first T.A pair in the sequence can
be replaced for any base pair, the Tth111I recognition sequence has the
symmetry with the two-fold axis as most type II restriction endonucleases do.

<>

<1>Shintani, M., Hosoyama, A., Ohji, S., Tsuchikane, K., Takarada, H., Yamazoe, A., Fujita, N., Nojiri, H.
<2>Complete Genome Sequence of the Carbazole Degrader Pseudomonas resinovorans Strain CA10 (NBRC 106553).
<3>Genome Announcements
<4>1
<5>e00488-13
<6>2013
<7>Pseudomonas resinovorans strain CA10 can grow on carbazole as its sole carbon and nitrogen
source. Here, we report the complete nucleotide sequence of the CA10
genome (a 6,285,863-bp chromosome and a 198,965-bp plasmid). CA10 carries a
larger number of genes that are potentially responsible for aromatic hydrocarbon
metabolism than do other previously sequenced Pseudomonas spp.

<>

<1>Shintani, M., Ohtsubo, Y., Fukuda, K., Hosoyama, A., Ohji, S., Yamazoe, A., Fujita, N., Nagata, Y., Tsuda, M., Hatta, T., Kimbara, K.
<2>Complete Genome Sequence of the Thermophilic Polychlorinated Biphenyl Degrader Geobacillus sp. Strain JF8 (NBRC 109937).
<3>Genome Announcements
<4>2
<5>e01213-13
<6>2014
<7>Geobacillus sp. strain JF8 (NBRC 109937) utilizes biphenyl and naphthalene as sole carbon
sources and degrades polychlorinated biphenyl (PCB) at 60 degrees C.
Here, we report the complete nucleotide sequence of the JF8 genome (a
3,446,630-bp chromosome and a 39,678-bp plasmid). JF8 has the smallest genome
among the known PCB degraders.

<>

<1>Shirai, M., Hirakawa, H., Kimoto, M., Tabuchi, M., Kishi, F., Ouchi, K., Shiba, T., Ishii, K., Hattori, M., Kuhara, S., Nakazawa, T.
<2>Comparison of whole genome sequences of Chlamydia pneumoniae J138 from Japan and CWL029 from USA.
<3>Nucleic Acids Res.
<4>28
<5>2311-2314
<6>2000
<7>Chlamydia pneumoniae is a widespread pathogen of humans causing pneumonia and bronchitis.
There are many reports of an association between C. pneumoniae infection and atherosclerosis.
We determined the whole genome sequence of C. pneumoniae strain J138 isolated in Japan in 1994
and compared it with the sequence of strain CWL029 isolated in the USA before 1987.  The J138
circular chromosome consists of 1,226,565 nt (40.7% G&C) with 1072 likely protein-coding genes
that is 3665 nt shorter than the CWL029 genome.  Plasmids, phage- or transposon-like sequences
were not identified.  The overall genomic organization, gene order and predicted proteomes of
the two strains are very similar, suggesting a high level of structural and functional
conservation between the two unrelated isolates.  The most conspicuous differences in the J138
genome relative to the CWL029 genome are the absence of five DNA segments, ranging in size
from 89 to 1649 nt, and the presence of three DNA segments, ranging from 27 to 84 nt.  The
complex organization of these 'different zones' may be attributable to a unique system of
recombination.

<>

<1>Shiraishi, H., Tabuse, Y.
<2>The AplI Restriction-Modification System in an Edible Cyanobacterium, Arthrospira (Spirulina) platensis NIES-39, Recognizes the Nucleotide Sequence 5'-CTGCAG-3'.
<3>Biosci. Biotechnol. Biochem.
<4>77
<5>782-788
<6>2013
<7>The degradation of foreign DNAs by restriction enzymes in an edible cyanobacterium,
Arthrospira platensis, is a potential barrier for gene-transfer experiments in this
economically valuable organism. We overproduced in Escherichia coli the proteins involved in a
putative restriction-modification system of A. platensis NIES-39. The protein produced from
the putative type II restriction enzyme gene NIES39_K04640 exhibited an endonuclease activity
that cleaved DNA within the sequence 5'-CTGCAG-3' between the A at the fifth position and
the G at the sixth position. We designated this enzyme AplI. The protein from the adjacent
gene NIES39_K04650, which encodes a putative DNA (cytosine-5-)-methyltransferase, rendered DNA
molecules resistant to AplI by modifying the C at the fourth position (but not the C at the
first position) in the recognition sequence. This modification enzyme, M.AplI, should be
useful for converting DNA molecules into AplI-resistant forms for use in gene-transfer
experiments. A summary of restriction enzymes in various Arthrospira strains is also presented
in this paper.

<>

<1>Shirane, K., Toh, H., Kobayashi, H., Miura, F., Chiba, H., Ito, T., Kono, T., Sasaki, H.
<2>Mouse Oocyte Methylomes at Base Resolution Reveal Genome-Wide Accumulation of Non-CpG Methylation and Role of DNA Methyltransferases.
<3>PLoS Genet.
<4>9
<5>e1003439
<6>2013
<7>DNA methylation is an epigenetic modification that plays a crucial role in normal mammalian
development, retrotransposon silencing, and
cellular reprogramming. Although methylation mainly occurs on the
cytosine in a CG site, non-CG methylation is prevalent in pluripotent
stem cells, brain, and oocytes. We previously identified non-CG
methylation in several CG-rich regions in mouse germinal vesicle
oocytes (GVOs), but the overall distribution of non-CG methylation and
the enzymes responsible for this modification are unknown. Using
amplification-free whole-genome bisulfite sequencing, which can be used
with minute amounts of DNA, we constructed the base-resolution
methylome maps of GVOs, non-growing oocytes (NGOs), and mutant GVOs
lacking the DNA methyltransferase Dnmt1, Dnmt3a, Dnmt3b, or Dnmt3L. We
found that nearly two-thirds of all methylcytosines occur in a non-CG
context in GVOs. The distribution of non-CG methylation closely
resembled that of CG methylation throughout the genome and showed clear
enrichment in gene bodies. Compared to NGOs, GVOs were over four times
more methylated at non-CG sites, indicating that non-CG methylation
accumulates during oocyte growth. Lack of Dnmt3a or Dnmt3L resulted in
a global reduction in both CG and non-CG methylation, showing that
non-CG methylation depends on the Dnmt3a-Dnmt3L complex. Dnmt3b was
dispensable. Of note, lack of Dnmt1 resulted in a slight decrease in CG
methylation, suggesting that this maintenance enzyme plays a role in
non-dividing oocytes. Dnmt1 may act on CG sites that remain
hemimethylated in the de novo methylation process. Our results provide
a basis for understanding the mechanisms and significance of non-CG
methylation in mammalian oocytes.

<>

<1>Shiryaev, S.A., Zheleznaya, L.A., Matvienko, N.I.
<2>Three site-specific endonucleases from Thermophilic strain Bacillus species LA are isoschizomers of HhaI, AsuII, and HindIII.
<3>Biokhimiia
<4>62
<5>280-290
<6>1997
<7>Screening of thermophilic bacterial strains revealed a strain containing three site-specific
endonucleases: BspLAI, BspLAII, and BspLAIII.  These endonucleases were purified to functional
purity by sequential chromatography.  Recognition sites, DNA cleavage sites, and some
properties of the endonucleases were determined.  BspLAI recognizes the sequence 5'-GCG/C-3'
on the DNA molecule and is an isoschizomer of endonuclease HhaI.  BspLAII recognizes the
sequence 5'-TT/CGAA-3' and is an isoschizomer of AsuII.  BspLAIII recognizes site
5'-A/AGCTT-3' and is an isoschizomer of endonuclease HindIII.  All the three enzymes exhibit
maximal activity at 55oC.  The optimal buffer is MRB, pH 7.4.  They retain activity on storage
for 3 weeks at room temperature and thus are highly stable.

<>

<1>Shiryaev, S.A., Zheleznyakova, E.N., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>Two new site-specific endonucleases from Staphylococcus species strain D5.
<3>Biokhimiia
<4>65
<5>553-561
<6>2000
<7>Staphylococcus species strain D5 containing two site-specific endonucleases, SspD5I and
SspD5II, was found during screening of a bacterial strain collection from soil. These
endonucleases
were purified to functional homogeneity by sequential chromatography. Site-specific
endonuclease SspD5I recognizes the sequence 5'-GGTGA(8N/8N)^3' on DNA. Unlike HphI, it
cleaves DNA at a distance of 8 nucleotides from the recognition sequence on both chains
producing blunt-end DNA fragments, while endonuclease HphI cleaves DNA forming mononucleotide
3'-OH protruding ends. Thus, endonuclease SspD5I is a new type II site-specific endonuclease
and a neoschizomer of endonuclease HphI. The advantage of this new endonuclease is that the
blunt-end DNA products of this enzyme can be inserted without additional treatment into vector
DNAs cleaved with endonucleases yielding DNA blunt-ends. Endonuclease SspD5II recognizes the
site
5'-ATGCA^T-3' and thus is an isoschizomer of endonuclease NsiI. The molecular
mass of SspD5I is about 35 kD and that of SspD5II is 40 kD. The enzymes exhibit maximal
activity at 37 C. The optimal buffer for the reaction is HRB (10 mM Tris-HCl, pH 7.5,
10 mM MgCl2, 100 mM NaCl, and 1 mM dithiothreitol).

<>

<1>Shishido, K., Berg, P.
<2>Restriction endonuclease from Haemophilus gallinarum (HgaI) cleaves polyoma DNA at four locations.
<3>J. Virol.
<4>18
<5>793-798
<6>1976
<7>A restriction endonuclease obtained from Haemophilus gallinarum (HgaI) cleaves
polyoma DNA at four specific sites.  Using the EcoRI, HindIII, and HpaII
endonuclease restriction sites as reference, the four HgaI cleavage sites were
mapped at 0.02, 0.14, 0.27, and 0.48 fractional lengths, clockwise, from the
single EcoRI cleavage site.

<>

<1>Shivaji, S., Ara, S., Bandi, S., Singh, A., Kumar, P.A.
<2>Draft Genome Sequence of Arthrobacter gangotriensis Strain Lz1yT, Isolated from a Penguin Rookery Soil Sample Collected in Antarctica, near the Indian Station  Dakshin Gangotri.
<3>Genome Announcements
<4>1
<5>e00347-13
<6>2013
<7>We report here the 4.3-Mb genome of Arthrobacter gangotriensis strain Lz1y(T), isolated from a
penguin rookery soil sample collected in Antarctica, near the
Indian station Dakshin Gangotri.

<>

<1>Shivaji, S., Ara, S., Begum, Z., Ruth, M., Singh, A., Kumar, P.A.
<2>Draft Genome Sequence of Bhargavaea cecembensis Strain DSE10T, Isolated from a Deep-Sea Sediment Sample Collected at a Depth of 5,904 m from the  Chagos-Laccadive Ridge System in the Indian Ocean.
<3>Genome Announcements
<4>1
<5>e00346-13
<6>2013
<7>Here, we report the 3.2-Mbp draft genome sequence of Bhargavaea cecembensis strain DSE10(T),
isolated from a sediment sample collected from the
Chagos-Laccadive ridge system in the Indian Ocean at a depth of 5,904 m.

<>

<1>Shivaji, S., Ara, S., Begum, Z., Srinivas, T.N., Singh, A., Kumar, P.A.
<2>Draft Genome Sequence of Cesiribacter andamanensis Strain AMV16T, Isolated from a Soil Sample from a Mud Volcano in the Andaman Islands, India.
<3>Genome Announcements
<4>1
<5>e00240-13
<6>2013
<7>Here we report the 4.75-Mb genome of Cesiribacter andamanensis strain AMV16(T), isolated from
a soil sample from a mud volcano in the Andaman Islands, India.

<>

<1>Shivaji, S., Ara, S., Prasad, S., Manasa, B.P., Begum, Z., Singh, A., Kumar, P.A.
<2>Draft Genome Sequence of Arcticibacter svalbardensis Strain MN12-7T, a Member of  the Family Sphingobacteriaceae Isolated from an Arctic Soil Sample.
<3>Genome Announcements
<4>1
<5>e00484-13
<6>2013
<7>The 4.69-Mb genome sequence of Arcticibacter svalbardensis strain MN12-7(T), isolated from an
Arctic soil sample, is reported.

<>

<1>Shivaji, S., Ara, S., Singh, A., Kumar, P.A.
<2>Draft Genome Sequence of Cyclobacterium qasimii Strain M12-11BT, Isolated from Arctic Marine Sediment.
<3>Genome Announcements
<4>1
<5>e00642-13
<6>2013
<7>A 6.29-Mb genome sequence of Cyclobacterium qasimii strain M12-11B(T), isolated from an Arctic
marine sediment sample, is reported.

<>

<1>Shivaji, S., Ara, S., Singh, A., Pinnaka, A.K.
<2>Draft Genome Sequence of Cecembia lonarensis Strain LW9T, Isolated from Lonar Lake, a Haloalkaline Lake in India.
<3>J. Bacteriol.
<4>194
<5>6631
<6>2012
<7>The draft genome sequence (4.84 Mb) of Cecembia lonarensis strain LW9(T), isolated from a
water sample (4.5-m depth) from Lonar Lake, a meteorite-created
haloalkaline lake in India, is reported. The enzymes produced by these
microorganisms need to be stable under alkaline conditions prevailing in its
habitat. Such enzymes would be of immense importance for enzymatic processes
operating at high pH.

<>

<1>Shivaji, S., Ara, S., Singh, S.K., Bandi, S., Singh, A., Pinnaka, A.K.
<2>Draft Genome Sequence of Bacillus isronensis Strain B3W22, Isolated from the Upper Atmosphere.
<3>J. Bacteriol.
<4>194
<5>6624-6625
<6>2012
<7>We report the 4.0-Mb genome sequence of Bacillus isronensis strain B3W22 isolated from air
collected at an altitude ranging from 27 to 30 km above the city of
Hyderabad, in India. This genome sequence will contribute to the objective of
determining the microbial diversity of the upper atmosphere.

<>

<1>Shivapriya, R., Prasad, R., Narayanan, I.L., Krishnaswamy, S., Dharmalingam, K.
<2>Expression of the mcrA gene of Escherichia coli is regulated post-transcriptionally, possibly by sequestration of the Shine-Dalgarno region.
<3>Gene
<4>157
<5>201-207
<6>1995
<7>The polypeptides encoded by the mcrA gene were analysed using a T7 expression system.  Cloned
fragments of 1.6 and 1.0 kb displayed an McrA+/RglA+ phenotype and directed synthesis of a
31-kDa polypeptide.  A derivative of these clones altered at an internal HindIII site
displayed an McrA+/RglA- phenotype and directed production of a 23-kDa polypeptide.  A
construct carrying the 1.0-kb mcrA insert yielded a single 1.3-kb transcript.  The mcrA
transcript was found to start from C, G, T and G, namely the fourth, fifth, sixth and seventh
nucleotides (nt), respectively, downstream from the last nt of the putative -10 region.  Two
mcrA transcriptional/translational fusions were made in the pT7-7 expression vector and the
protein encoded by these constructs were analysed.  Regulation of mcrA expression was studied
by quantitative Northern anlaysis of RNA from various mcrA clones.  Together with a computer
analysis of the translation initiation region in these mRNAs, the results suggest that the
expression of mcrA may be regulated at the translational level.

<>

<1>Shkoporov, A.N., Efimov, B.A., Khokhlova, E.V., Chaplin, A.V., Kafarskaya, L.I., Durkin, A.S., McCorrison, J., Torralba, M., Gillis, M., Sutton, G., Weibel, D.B., Nelson, K.E., Smeianov, V.V.
<2>Draft Genome Sequences of Two Pairs of Human Intestinal Bifidobacterium longum subsp. longum Strains, 44B and 1-6B and 35B and 2-2B, Consecutively Isolated from  Two Children after a 5-Year Time Period.
<3>Genome Announcements
<4>1
<5>e00234-13
<6>2013
<7>We report the genome sequences of four isolates of a human gut symbiont, Bifidobacterium
longum. Strains 44B and 35B were isolated from two 1-year-old
infants, while 1-6B and 2-2B were isolated from the same children 5 years later.
The sequences permit investigations of factors enabling long-term colonization of
bifidobacteria.

<>

<1>Shlyapnikov, S.V., Lunin, V.V., Blagova, E.V., Abaturov, L.V., Perbandt, M., Betzel, C., Mikhailov, A.M.
<2>A comparative structure-function analysis and molecular mechanism of action of endonucleases from Serratia marcescens and Physarum  polycephalum.
<3>Bioorg. Khim.
<4>28
<5>23-31
<6>2002
<7>Structural and functional characteristics were compared for wild-type nuclease from Serratia
marcescens, which belongs to the family of DNA/RNA nonspecific endonucleases, its mutational
forms, and the nuclease I-PpoI from Physarum polycephalum, which is a representative of the
Cys-His box-containing subgroup of the superfamily of extremely specific intron-encoded homing
DNases. Despite the lack of sequence homology and the overall different topology of the
Serratia marcescens and I-PpoI nucleases, their active sites have a remarkable structural
similarity. Both of them have a unique magnesium atom in the active site, which is a part of
the coordinately bonded water-magnesium complex involved in their catalytic acts. In the
enzyme-substrate complexes, the Mg2+ ion is chelated by an Asp residue, coordinates two oxygen
atoms of DNA, and stabilizes the transition state of the phosphate anion and 3'-OH group of
the leaving nucleotide. A new mechanism of the phosphodiester bond cleavage, which is common
for the Serratia marcescens and I-PpoI nucleases and differs from the known functioning
mechanism of the restriction and homing endonucleases, was proposed. It presumes a His residue
as a general base for the activation of a non-cluster water molecule at the nucleophilic in
line displacement of the 3'-leaving group. A strained metalloenzyme-substrate complex is
formed during hydrolysis and relaxes to the initial state after the reaction.

<>

<1>Shock, L.S., Thakkar, P.V., Peterson, E.J., Moran, R.G., Taylor, S.M.
<2>DNA methyltransferase 1, cytosine methylation, and cytosine hydroxymethylation in mammalian mitochondria.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>3630-3635
<6>2011
<7>Mitochondrial DNA (mtDNA) has been reported to contain 5-methylcytosine (5mC) at CpG
dinucleotides, as in the nuclear genome, but neither the
mechanism generating mtDNA methylation nor its functional significance
is known. We now report the presence of 5-hydroxymethylcytosine (5hmC)
as well as 5mC in mammalian mtDNA, suggesting that previous studies
underestimated the level of cytosine modification in this genome. DNA
methyltransferase 1 (DNMT1) translocates to the mitochondria, driven by
a mitochondrial targeting sequence located immediately upstream of the
commonly accepted translational start site. This targeting sequence is
conserved across mammals, and the encoded peptide directs a
heterologous protein to the mitochondria. DNMT1 is the only member of
the three known catalytically active DNA methyltransferases targeted to
the mitochondrion. Mitochondrial DNMT1 (mtDNMT1) binds to mtDNA,
proving the presence of mtDNMT1 in the mitochondrial matrix. mtDNMT1
expression is up-regulated by NRF1 and PGC1 alpha, transcription
factors that activate expression of nuclear-encoded mitochondrial genes
in response to hypoxia, and by loss of p53, a tumor suppressor known to
regulate mitochondrial metabolism. Altered mtDNMT1 expression
asymmetrically affects expression of transcripts from the heavy and
light strands of mtDNA. Hence, mtDNMT1 appears to be responsible for
mtDNA cytosine methylation, from which 5hmC is presumed to be derived,
and its expression is controlled by factors that regulate mitochondrial
function.

<>

<1>Shome, R., Krithiga, N., Muttannagouda, R.B., Veeregowda, B.M., Swati, S., Shome, B.R., Vishnu, U., Sankarasubramanian, J., Sridhar, J., Gunasekaran, P., Rahman, H., Rajendhran, J.
<2>Draft Genome Sequence of Brucella melitensis Strain ADMAS-G1, Isolated from Placental Fluids of an Aborted Goat.
<3>Genome Announcements
<4>1
<5>e00809-13
<6>2013
<7>Here, we report the draft genome sequence and annotation of the Brucella melitensis strain
designated ADMAS-G1, isolated from placental fluids of an
aborted goat. The length of the genome is 3,284,982 bp, with a 57.3% GC content.
A total of 3,325 protein-coding genes and 63 RNA genes were predicted.

<>

<1>Shome, R., Krithiga, N., Padmashree, B.S., Sankarasubramanian, J., Vishnu, U.S., Sridhar, J., Gunasekaran, P., Rajendhran, J., Rahman, H.
<2>Draft Genome Sequence of Brucella abortus S99: Designated Antigenic Smooth Reference Strain Used in Diagnostic Tests in India.
<3>Genome Announcements
<4>2
<5>e00824-14
<6>2014
<7>Brucella abortus strain S99 is widely used for the preparation of colored, plain, recombinant
and smooth lipopolysaccharide antigens for the preparation of
Brucella diagnostic kits. The genome of this strain was sequenced and the length
of the genome was 3,253,175 bp, with 57.2% G+C content. A total of 3,365 protein
coding genes and 53 RNA genes were predicted.

<>

<1>Shore, A., Rossney, A.S., Keane, C.T., Enright, M.C., Coleman, D.C.
<2>Seven Novel Variants of the Staphylococcal Chromosomal Cassette mec in Methicillin-Resistant Staphylococcus aureus Isolates from Ireland.
<3>Antimicrob. Agents Chemother.
<4>49
<5>2070-2083
<6>2005
<7>Methicillin-resistant Staphylococcus aureus (MRSA) isolates recovered in
Irish hospitals between 1971 and 2002 were characterized using multilocus
sequence typing (MLST) (n = 130) and SCCmec typing (n = 172). Where
atypical SCCmec typing results were obtained, PCR amplification of entire
SCCmec elements, analysis of amplimer mobility, and nucleotide sequencing
were undertaken. MLST revealed that 129/130 isolates had the same
genotypes as internationally spread MRSA clones, including ST239, ST247,
ST250, ST5, ST22, ST36, and ST8. A novel genotype, ST496, was identified
in one isolate. Half of the isolates (86/172) had SCCmec type I, IA, II,
III, or IV. The remaining 86 isolates harbored novel SCCmec variants in
three distinct genetic backgrounds: (i) 74/86 had genotype ST8 and either
one of five novel SCCmec II (IIA, IIB, IIC, IID, and IIE) or one of two
novel SCCmec IV (IVE and IVF) variants; (ii) 3/86 had genotype ST239 and a
novel SCCmec III variant; (iii) 9/86 had a novel SCCmec I variant
associated with ST250. SCCmec IVE and IVF were similar to SCCmec IVc and
IVb, respectively, but differed in the region downstream of mecA. The five
SCCmec II variants were similar to SCCmec IVb in the region upstream of
the ccr complex but otherwise were similar to SCCmec II, except for the
following regions: SCCmec IIA and IID had a novel mec complex, A.4 (Delta
mecI-IS1182-Delta mecI-mecR1-mecA-IS431mec); SCCmec IIC and IIE had a
novel mec complex, A.3 (IS1182-Delta mecI-mecR1-mecA-IS431mec); SCCmec IID
and IIE lacked pUB110; SCCmec IIC and IIE lacked a region of DNA between
Tn554 and the mec complex; and SCCmec IIB lacked Tn554. This study has
demonstrated a hitherto-undescribed degree of diversity within SCCmec.

<>

<1>Shore, A.C., Rossney, A.S., O'Connell, B., Herra, C.M., Sullivan, D.J., Humphreys, H., Coleman, D.C.
<2>Detection of staphylococcal cassette chromosome mec-associated DNA segments in multiresistant methicillin-susceptible Staphylococcus aureus  (MSSA) and identification of Staphylococcus epidermidis ccrAB4 in both  methicillin-resistant S. aureus and MSSA.
<3>Antimicrob. Agents Chemother.
<4>52
<5>4407-4419
<6>2008
<7>Methicillin-susceptible Staphylococcus aureus (MSSA) can arise from methicillin-resistant S.
aureus (MRSA) following partial or complete
excision of staphylococcal cassette chromosome mec (SCCmec). This study
investigated whether multiresistant MSSA isolates from Irish hospitals,
where MRSA has been endemic for decades, harbor SCCmec DNA. Twenty-five
multiresistant MSSA isolates recovered between 2002 and 2006 were tested
for SCCmec DNA by PCR and were genotyped by multilocus sequence typing and
spa typing. All isolates lacked mecA. Three isolates (12%) harbored SCCmec
DNA; two of these (genotype ST8/t190) harbored a 26-kb SCCmec IID
(II.3.1.2) remnant that lacked part of mecI and all of mecR1, mecA, and
IS431; the third isolate (ST8/t3209) harbored the SCCmec region from dcs
to orfX. All three isolates were detected as MRSA using the BD GeneOhm and
Cepheid's Xpert MRSA real-time PCR assays. Six isolates (ST8/t190, n = 4;
ST5/t088, n = 2), including both isolates with the SCCmec IID remnant,
harbored ccrAB4 with 100% identity to ccrAB4 from the Staphylococcus
epidermidis composite island SCC-CI. This ccrAB4 gene was also identified
in 23 MRSA isolates representative of ST8/t190-MRSA with variant SCCmec II
subtypes IIA to IIE, which predominated previously in Irish hospitals.
ccrAB4 was located 5,549 bp upstream of the left SCCmec junction in both
the MRSA and MSSA isolates with SCCmec elements and remnants and 5,549 bp
upstream of orfX in the four MSSA isolates with ccrAB4 only on an SCC-CI
homologous region. This is the first description of a large SCCmec remnant
with ccr and partial mec genes in MSSA and of the S. epidermidis SCC-CI
and ccrAB4 genes in S. aureus.

<>

<1>Shorning, B.Y., Vanyushin, B.F.
<2>Putative DNA-(amino)methyltransferases in eucaryotes.
<3>Biokhimiia
<4>66
<5>753-762
<6>2001
<7>By computer analysis of the known data bases, we have established that the open reading frames
(ORF) coding for proteins that possess high degree of homology with procaryotic
DNA-(amino)methyltransferases are present in the genomes of Leishmania major, Saccharomyces
cerevisiae, Schizosaccharomyces pombe, Arabidopsis thaliana, Drosophila melanogaster,
Caenorhabditis elegans, and Homo sapiens. Conservative motifs typical for bacterial
DNA-(amino)methyltransferases are detected in the amino acid sequences of these putative
proteins. The ORF of all putative eucaryotic DNA-(amino)methyltransferases found are encoded
in nuclear DNA. In mitochondrial genomes including a few fully sequenced higher plant mtDNA,
nucleotide sequences significantly homologous to genes of procaryotic
DNA-(amino)methyltransferases are not found. Thus, ORF homologous to bacterial adenine
DNA-methyltransferases are present in nuclei of protozoa, yeasts, insects, nematodes,
vertebrates, higher plants, and other eucaryotes. A special search for corresponding proteins
and, in particular, adenine DNA-methyltransferases in these organisms and a study of their
functions are quite promising.

<>

<1>Shouche, Y.S., Ramesh, N., Brahmachari, S.K.
<2>Probing of unusual DNA structures in topologically constrained form V DNA: use of restriction enzymes as structural probes.
<3>Nucleic Acids Res.
<4>18
<5>267-275
<6>1990
<7>The ability of DNA sequences to adopt unusual structures under superhelical
torsional stress has been studied.  Sequences that are forced to adopt an
unusual conformation in topologically constrained pBR322 form V DNA (Lk=0) were
mapped using restriction enzymes as probes.  Restriction enzymes such as BamHI,
PstI, AvaI and HindIII could not cleave their recognition sequences.  The
removal of topological constraint relieved this inhibition.  The influence of
neighbouring sequences on the ability of a given sequence to adopt an unusual
DNA structure, presumbly a left handed Z conformation, was studied through
single hit analysis.  Using multiple cut restriction enzymes such as NarI and
FspI, it could be shown that under identical topological strain, the extent of
structural alteration is greatly influenced by the neighbouring sequences.  In
light of the variety of sequences and locations that could be mapped to adopt
non-B conformation in pBR322 form V DNA, restriction enzymes appear as
potential structural probes for natural DNA sequences.

<>

<1>Shrestha, P.M., Kube, M., Reinhardt, R., Liesack, W.
<2>Transcriptional activity of paddy soil bacterial communities.
<3>Environ. Microbiol.
<4>11
<5>960-970
<6>2008
<7>Summary Bulk mRNA was used to explore the transcriptional activity of bacterial communities in
oxic versus anoxic paddy soil. Two microbial cDNA
libraries were constructed from composite samples using semi-randomly
primed RT-PCR. cDNAs averaged 500-600 bp in length and were treated as
expressed sequence tags (ESTs). Clustering analysis of 805 random cDNAs
resulted in 179 and 155 different ESTs for the oxic and anoxic zones
respectively. Using an E-value threshold of e(-10), a total of 218
different ESTs could be assigned by blastx, while 116 ESTs were predicted
novel. Both the proportion and significance of the EST assignments
increased with cDNA length. Taxonomic assignment was more powerful in
discriminating between the aerobic and anaerobic bacterial communities
than functional inference, as most ESTs in both oxygen zones were putative
indicators of similar housekeeping functions, in particular ABC-type
transporters. A few ESTs were putative indicators for community function
in a biogeochemical context, such as beta-oxidation of long-chain fatty
acids specifically in the oxic zone. Expressed sequence tags assigned to
Alpha- and Betaproteobacteria were predominantly found in the oxic zone,
while those affiliated with Deltaproteobacteria were more frequently
detected in the anoxic zone. At the genus level, multiple assignments to
Bradyrhizobium and Geobacter were unique to the oxic and anoxic zones
respectively. The phylum-level affiliations of 93 16S rRNA sequences
corresponded well with two taxonomically distinct EST patterns. Expressed
sequence tags affiliated with Acidobacteria and Chloroflexi were
frequently detected in both oxygen zones. In summary, the soil
metatranscriptome is accessible for global analysis and such studies have
great potential in elucidating the taxonomic and functional status of soil
bacterial communities, but study significance depends on the number and
length of cDNAs being randomly analysed.

<>

<1>Shrestha, R., Park, D.H., Cho, J.M., Cho, S., Wilson, C., Hwang, I., Hur, J.H., Lim, C.K.
<2>Genetic organization of the hrp genes cluster in Erwinia pyrifoliae and characterization of HR active domains in HrpNEp protein by mutational analysis.
<3>Mol. Cells
<4>25
<5>30-42
<6>2008
<7>The disease-specific (dsp) region and the hypersensitive response and
pathogenicity (hrp) genes, including the hrpW, hrpNEp, and hrpC operons
have previously been sequenced in Erwinia pyrifoliae WT3 [Shrestha et al.
(2005a)]. In this study, the remaining hrp genes, including the hrpC,
hrpA, hrpS, hrpXY, hrpL and hrpJ operons, were determined. The hrp genes
cluster (ca. 38 kb) was comprised of eight transcriptional units and
contained nine hrc (hrp conserved) genes. The genetic organization of the
hrp/hrc genes and their orientation for the transcriptions were also
similar to and collinear with those of E. amylovora, showing > or = 80%
homologies. However, ORFU1 and ORFU2 of unknown functions, present between
the hrpA and hrpS operons of E. amylovora, were absent in E. pyrifoliae.
To determine the HR active domains, several proteins were prepared from
truncated fragments of the N-terminal and the C-terminal regions of
HrpN(Ep) protein of E. pyrifoliae. The proteins prepared from the
N-terminal region elicited HR, but not from those of the C-terminal region
indicating that HR active domains are located in only N-terminal region of
the HrpN(Ep) protein. Two synthetic oligopeptides produced HR on tobacco
confirming presence of two HR active domains in the HrpN(Ep). The HR
positive N-terminal fragment (HN delta C187) was further narrowed down by
deleting C-terminal amino acids and internal amino acids to investigate
whether amino acid insertion region have role in faster and stronger HR
activity in HrpN(Ep) than HrpN(Ea). The HrpN(Ep) mutant proteins HN delta
C187 (D1AIR), HN delta C187 (D2AIR) and HN delta C187 (DM41) retained
similar HR activation to that of wild-type HrpN(Ep). However, the HrpN(Ep)
mutant protein HN delta C187 (D3AIR) lacking third amino acid insertion
region (102 to 113 aa) reduced HR when compared to that of wild-type
HrpN(Ep). Reduction in HR elicitation could not be observed when single
amino acids at different positions were substituted at third amino acids
insertion region. But, substitution of amino acids at L103R, L106K and
L110R showed reduction in HR activity on tobacco suggesting their
importance in activation of HR faster in the HrpN(Ep) although it requires
further detailed analysis.

<>

<1>Shtratnikova, V.Y., Bragin, E.Y., Dovbnya, D.V., Pekov, Y.A., Schelkunov, M.I., Strizhov, N., Ivashina, T.V., Ashapkin, V.V., Donova, M.V.
<2>Complete Genome Sequence of Sterol-Transforming Mycobacterium neoaurum Strain VKM Ac-1815D.
<3>Genome Announcements
<4>2
<5>e01177-13
<6>2014
<7>Mycobacterium neoaurum strain VKM Ac-1815D produces 4-androstene-3,17-dione as a  major
compound from phytosterols. Here, we report the complete genome sequence of
the strain. The genome consists of a single circular 5,438,190-bp chromosome,
with a G+C content of 66.88%, containing 5,318 putative open reading frames
(ORFs), 46 tRNAs, and 6 rRNAs. Arrays of cholesterol metabolism genes are
randomly clustered throughout the chromosome.

<>

<1>Shu, H.W., Liu, T.T., Chan, H.I., Liu, Y.M., Wu, K.M., Shu, H.Y., Tsai, S.F., Hsiao, K.J., Hu, W.S., Ng, W.V.
<2>Genome Sequence of the Repetitive-Sequence-Rich Mycoplasma fermentans Strain M64.
<3>J. Bacteriol.
<4>193
<5>4302-4303
<6>2011
<7>Mycoplasma fermentans is a microorganism commonly found in the genitourinary and respiratory
tracts of healthy individuals and AIDS
patients. The complete genome of the repetitive-sequence-rich M.
fermentans strain M64 is reported here. Comparative genomics analysis
revealed dramatic differences in genome size between this strain and the
recently completely sequenced JER strain.

<>

<1>Shub, D.A., Goodrich-Blair, H.
<2>Protein introns: A new home for endonucleases.
<3>Cell
<4>71
<5>183-186
<6>1992
<7>An underlying principle of the classical era of molecular biology--that RNA is an exact copy
of information in one of the DNA strands--was shattered forever with the discovery of RNA
splicing. Although at first introns could be considered an aberration of eukaryotes and their
viruses, self-splicing RNAs were soon discovered in mitochondrial, chloroplastic, nuclear,
bacteriophage, and bacterial genes. Additional modes of gene organization and expression
continue to emerge with no end in sight. Over the past several years we have witnessed the
discovery of programmed ribosomal frameshifting and hopping, trans-splicing, RNA editing, and
cotranslational suppression of nonsense codons. The latest of these surprises is protein
splicing: the excision of an internal segment of a polypeptide and religation of the flanking
regions to create a functional protein. The evidence for protein splicing rests on data from
three genes: VMA1/TFP1, which encodes a subunit of yeast vacuolar ATPase, the gene encoding
DNA polymerase of the archaen thermophile Thermococcus litoralis (with two "introns"), and the
recA gene of Mycobacterium tuberculosis. Although the number of examples is small, there is
one from each of the three major phylogenetic divisions, so the phenomenon may be very widely
distributed. Phylogenetic distance notwithstanding, these phenomena are remarkably similar:
findings in each system have foreshadowed similar results in the others. Therefore, although
specific citations are given, a summary of data from all three systems will be presented.

<>

<1>Shub, D.A., Goodrich-Blair, H.
<2>Amino acid sequence motif of group I intron endonucleases is conserved in open reading frames of group II introns.
<3>Trends Biochem. Sci.
<4>19
<5>402-404
<6>1994
<7>Both group I and group II intron RNAs are capable of self splicing, with cleavage-ligation
proceeding by concerted transesterification reactions. Although they tend to be localized to
the same cellular compartments (mitochondria and chloroplasts), these two categories of
self-splicing RNAs bear no features that would indicate a common evolutionary origin. Recent
attempts to ascribe similarities to the intermolecular structures formed in nuclear
spliceosomes and the intramolecular structures in group I and group II introns underscore the
current interest in determining the evolutionary origins and interactions of these
self-splicing intron classes.

<>

<1>Shub, D.A., Gott, J.M., Xu, M.Q., Lang, B.F., Michel, F., Tomashewski, J., Pedersen-Lane, J., Belfort, M.
<2>Structural conservation among three homologous introns of bacteriophage T4 and the group I introns of eukaryotes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>85
<5>1151-1155
<6>1988
<7>Three group I introns of bacteriophage T4 have been compared with respect to their sequence
and structural properties. The introns include the td intervening sequence, as well as the two
newly described introns in the nrdB and sunY genes of T4. The T4 introns are very closely
related, containing phylogenetically conserved sequence elements that allow them to be folded
into a core structure that is characteristic of eukaryotic group IA introns. Similarities
extend outward to the exon sequences surrounding the three introns. All three introns contain
open reading frames (ORFs). Although the intron ORFs are not homologous and occur at different
positions, all three ORFs are looped-out of the structure models, with only the 3' ends of
each of the ORFs extending into the secondary structure. This arrangement invites interesting
speculations on the regulation of splicing by translation. The high degree of similarity
between the T4 introns and the eukaryotic group I introns must reflect a common ancestry,
resulting either from vertical acquisition of a primordial RNA element or from horizontal
transfer.

<>

<1>Shub, D.A., Xu, M.-Q., Gott, J.M., Zeeh, A., Wilson, L.D.
<2>A family of autocatalytic group I introns in bacteriophage T4.
<3>Cold Spring Harb. Symp. Quant. Biol.
<4>52
<5>193-200
<6>1987
<7>The first intron to be discovered in a prokaryotic mRNA was found in the td gene of
bacteriophage T4.  The presence of an intron in this gene, which encodes thymidylate synthase,
was detected by sequence comparisons at the DNA and protein levels.  Thus, a stop codon was
encountered in the DNA sequence before the end of the protein-coding sequence.  Subsequent
experimentation revealed that the intron in the td gene was excised autocatalytically from
precursor RNA.  The splicing mechanism resembled that used by group I introns of eukaryotes: a
series of transesterification reactions triggered by nucleophilic attack by guanosine (or GTP)
at the 5' splice site.  The primary intron excision product was linear, containing a noncoded
G at the 5' end.  Although the discovery of an intron in phage T4 was entirely unexpected, it
encouraged us (in collaboration with M. Belfort) to look for additional group I introns in the
T4 genome.  Further examples would permit sequence and structural comparisons that might lend
insight into their evolutionary origin.  Additionally, we hoped that their locations within
the T4 genome would infer a possible regulatory function in prokaryotic gene expression.  We
took advantage of the fact that, since G is added to the 5' end of the intron, autocatalytic
group I introns could be specifically labeled in vitro for use as probes for DNA blotting
experiments.  If group I introns were in more than just the td gene, multiple RNA species
would be labeled when total RNA is extracted from T4-infected cells and incubated with
[a-32P]GTP in vitro.  When used as a probe for a Southern blot of T4 DNA, this RNA should
hybridize to several DNA bands.

<>

<1>Shukla, H., Kobayashi, Y., Arenstorf, H., Yasukochi, Y., Weissman, S.M.
<2>Purification of BsuE methyltransferase and its application in genome mapping.
<3>Nucleic Acids Res.
<4>19
<5>4233-4239
<6>1991
<7>We have used a combination of BsuE methyltransferase (M.BsuEII) and NotI
restriction enzyme to cut genomic DNA at a subset of NotI sites.  The
usefulness of this system is shown in a re-examination of the restriction map
of the human MHC.  Combinations of methylases and restriction enzymes can be
used to generate cuts at different frequencies in genomic DNA, such that they
generate ends complementary to NotI ends, and can be used in conjunction with
NotI linking clones in chromosome jumping experiments.  These enzyme
combinations have the potential to produce cutting sites in genomic DNA spaced
at intervals favorable for extensive mapping, fragment enrichment, and cloning
efforts.

<>

<1>Shulman, M.J.
<2>Model for wandering restriction enzymes.
<3>Nature
<4>252
<5>76-78
<6>1974
<7>None

<>

<1>Shutiashunaida, U., Dewasthaly, S., Hannah, M., Meinke, A., Nagy, E.
<2>S. pneumoniae antigens.
<3>Japanese Patent Office
<4>JP 2007525157 A
<5>
<6>2007
<7>
<>

<1>Sibley, M.H., Raleigh, E.A.
<2>Cassette-like variation of restriction enzyme genes in Escherichia coli C and relatives.
<3>Nucleic Acids Res.
<4>32
<5>522-534
<6>2004
<7>A surprising result of comparative bacterial genomics has been the large amount of DNA found
to be present in one strain but not in another of the same species. We examine in detail one
location where gene content varies extensively, the restriction cluster in Escherichia coli.
This region is designated the Immigration Control Region (ICR) for the density and variability
of restriction functions found there. To better define the boundaries of this variable locus,
we determined the sequence of the region from a restrictionless strain, E.coli C. Here we
compare the 13.7 kb E.coli C sequence spanning the site of the ICR with corresponding
sequences from five E.coli strains and Salmonella typhimurium LT2. To discuss this variation,
we adopt the term 'framework' to refer to genes that are stable components of genomes within
related lineages, while 'migratory' genes are transient inhabitants of the genome.
Strikingly, seven different migratory DNA segments, encoding different sets of genes and gene
fragments, alternatively occupy a single well-defined location in the seven strains examined.
The flanking framework genes, yjiS and yjiA, display approximately normal patterns of
conservation. The patterns observed are consistent with the action of a site-specific
recombinase. Since no nearby gene codes for a likely recombinase of known families, such a
recombinase must be of a new family or unlinked.

<>

<1>Siddaramappa, S. et al.
<2>Complete genome sequence of Dehalogenimonas lykanthroporepellens type strain (BL-DC-9(T)) and comparison to 'Dehalococcoides' strains.
<3>Standards in Genomic Sciences
<4>6
<5>251-264
<6>2012
<7>Dehalogenimonas lykanthroporepellens is the type species of the genus Dehalogenimonas, which
belongs to a deeply branching lineage within the phylum
Chloroflexi. This strictly anaerobic, mesophilic, non spore-forming,
Gram-negative staining bacterium was first isolated from chlorinated solvent
contaminated groundwater at a Superfund site located near Baton Rouge, Louisiana,
USA. D. lykanthroporepellens was of interest for genome sequencing for two
reasons: (a) an unusual ability to couple growth with reductive dechlorination of
environmentally important polychlorinated aliphatic alkanes and (b) a
phylogenetic position that is distant from previously sequenced bacteria. The
1,686,510 bp circular chromosome of strain BL-DC-9(T) contains 1,720 predicted
protein coding genes, 47 tRNA genes, a single large subunit rRNA (23S-5S) locus,
and a single, orphan, small subunit rRNA (16S) locus.

<>

<1>Siddavattam, D., Karegoudar, T.B., Mudde, S.K., Kumar, N., Baddam, R., Avasthi, T.S., Ahmed, N.
<2>Genome of a Novel Isolate of Paracoccus denitrificans Capable of Degrading N,N-Dimethylformamide.
<3>J. Bacteriol.
<4>193
<5>5598-5599
<6>2011
<7>The bacterial genus Paracoccus is comprised of metabolically versatile organisms having
diverse degradative capabilities and potential industrial
and environmental applications for bioremediation in particular. We report
a de novo-assembled sequence and annotation of the genome of a novel
isolate of Paracoccus denitrificans originally sourced from coal mine
tailings in India. The isolate was capable of utilizing
N,N-dimethylformamide (DMF) as a source of carbon and nitrogen and
therefore holds potential for bioremediation and mineralization of
industrial pollutants. The genome sequence and biological circuitry
revealed thereupon will be invaluable in understanding the metabolic
capabilities, functioning, and evolution of this important bacterial
organism.

<>

<1>Siddique, A., Jurkowska, R.Z., Jurkowski, T.P., Jeltsch, A.
<2>Auto-methylation of the mouse DNA-(cytosine C5)-methyltransferase Dnmt3a at its active site cysteine residue.
<3>FEBS J.
<4>278
<5>2055-2063
<6>2011
<7>The Dnmt3a DNA methyltransferase is responsible for establishing DNA methylation patterns
during mammalian development. We show here that
the mouse Dnmt3a DNA methyltransferase is able to transfer the methyl
group from S-adenosyl-L-methionine (AdoMet) to a cysteine residue in
its catalytic center. This reaction is irreversible and relatively
slow. The yield of auto-methylation is increased by addition of Dnmt3L,
which functions as a stimulator of Dnmt3a and enhances its AdoMet
binding. Auto-methylation was observed in binary Dnmt3a AdoMet
complexes. In the presence of CpG containing dsDNA, which is the
natural substrate for Dnmt3a, the transfer of the methyl group from
AdoMet to the flipped target base was preferred and auto-methylation
was not detected. Therefore, this reaction might constitute a
regulatory mechanism which could inactivate unused DNA
methyltransferases in the cell, or it could simply be an aberrant side
reaction caused by the high methyl group transfer potential of AdoMet.

<>

<1>Siddiqui, H., Yoder-Himes, D.R., Mizgalska, D., Nguyen, K.A., Potempa, J., Olsen, I.
<2>Genome Sequence of Porphyromonas gingivalis Strain HG66 (DSM 28984).
<3>Genome Announcements
<4>2
<5>e00947-14
<6>2014
<7>Porphyromonas gingivalis is considered a major etiologic agent in adult periodontitis.
Gingipains are among its most important virulence factors, but
their release is unique in strain HG66. We present the genome sequence of HG66
with a single contig of 2,441,680 bp and a G+C content of 48.1%.

<>

<1>Siddiqui, M.A., Rashid, N., Ayyampalayam, S., Whitman, W.B.
<2>Draft Genome Sequence of Geobacillus thermopakistaniensis Strain MAS1.
<3>Genome Announcements
<4>2
<5>e00559-14
<6>2014
<7>Geobacillus thermopakistaniensis strain MAS1 was isolated from a hot spring located in the
Northern Areas of Pakistan. The draft genome sequence was 3.5 Mb
and identified a number of genes of potential industrial importance, including
genes encoding glycoside hydrolases, pullulanase, amylopullulanase, glycosidase,
and alcohol dehydrogenases.

<>

<1>Sidorova, N., Muradymov, S., Rau, D.C.
<2>Specific versus Nonspecific DNA Binding of the Restriction Endonuclease EcoRV Measured by Self-Cleavage Assay.
<3>J. Biomol. Struct. Dyn.
<4>26
<5>896-897
<6>2009
<7>
<>

<1>Sidorova, N., Rau, D.C.
<2>The role of water in the EcoRI-DNA binding.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>319-337
<6>2004
<7>In many respects, Type II restriction endonucleases are prototypical DNA-binding proteins.  In
order to avoid catastrophic consequences for the cell, however, these enzymes must be far more
stringent in recognition of their target sequences and subsequent DNA cleavage than other
specific sequence recognition proteins that regulate gene activity.  In contrast to E. coli
Lac and lambda Cro repressors, for example, that show gradually decreasing binding energies as
the recognition sequence is changed, many restriction nucleases are exquisitely specific.
EcoRI will bind to its recognition sequence, GAATTC, with an association equilibrium constant
Ka,sp ~10^11 M-1 and to a completely nonspecific sequence with Ka,nonsp ~10^7 M-1.  A change
of even a single base pair is sufficient to decrease the binding constant at least by 10^3,
bringing it within a factor ~10 or less of nonspecific binding.

<>

<1>Sidorova, N.Y., Muradymov, S., Rau, D.C.
<2>Differences in hydration coupled to specific and nonspecific competitive binding and to specific DNA binding of the restriction endonuclease BamHI.
<3>J. Biol. Chem.
<4>281
<5>35656-35666
<6>2006
<7>Using the osmotic stress technique together with a self-cleavage assay we measure directly
differences in sequestered water between specific
and nonspecific DNA-BamHI complexes as well as the numbers of water
molecules released coupled to specific complex formation. The
difference between specific and nonspecific binding free energy of the
BamHI scales linearly with solute osmolal concentration for seven
neutral solutes used to set water activity. The observed osmotic
dependence indicates that the nonspecific DNA-BamHI complex sequesters
some 120-150 more water molecules than the specific complex. The weak
sensitivity of the difference in number of waters to the solute
identity suggests that these waters are sterically inaccessible to
solutes. This result is in close agreement with differences in the
structures determined by x-ray crystallography. We demonstrate
additionally that when the same solutes that were used in competition
experiments are used to probe changes accompanying the binding of free
BamHI to its specific DNA sequence, the measured number of water
molecules released in the binding process is strikingly
solute-dependent ( with up to 10-fold difference between solutes). This
result is expected for reactions resulting in a large change in a
surface exposed.

<>

<1>Sidorova, N.Y., Muradymov, S., Rau, D.C.
<2>A novel self-cleavage assay for measuring DNA binding of restriction endonucleases.
<3>FEBS J.
<4>272
<5>565
<6>2005
<7>DNA-protein interactions can be analyzed by methods based on physical separation of complexes
from free DNA fragments (gel mobility shift and filter binding assays) or by techniques
probing DNA-protein equilibrium directly in a solution (e.g. fluorescence and calorimetry).
These later techniques require larger amounts of materials and do not provide enough
sensitivity for measuring equilibrium association constants in the range 10^9-10^12 M-1. Both
gel mobility shift and filter binding assays that can measure these large binding constants,
however, have been criticized for the danger of disrupting equilibrium by physically
separating DNA-protein complexes from free species. We have developed a novel self-cleavage or
self-footprinting assay that uses the nuclease activity of restriction endonucleases to
measure sensitively their specific binding to DNA. At sufficiently high concentrations of
neutral osmolytes cleavage reaction can be triggered at only those DNA fragments with bound
enzyme. Under these conditions the fraction of specific DNA fragment cleaved reliably reflects
the fraction of DNA initially bound to the enzyme. The self-cleavage assay allows measurement
of binding equilibrium and kinetics directly in solution avoiding the intrinsic problems of
gel mobility shift and filter binding assays while providing the same sensitivity level. We
compare here the self-cleavage and gel mobility shift assays applied to the dissociation
kinetics and binding equilibrium of two restriction endonucleases, EcoRI and BamHI. The
technique can be used, in principle, for any protein possessing DNA cleavage activity if its
interactions with DNA are sensitive to osmotic stress.

<>

<1>Sidorova, N.Y., Muradymov, S., Rau, D.C.
<2>Solution parameters modulating DNA binding specificity of the restriction endonuclease EcoRV.
<3>FEBS J.
<4>278
<5>2713-2727
<6>2011
<7>The DNA binding stringency of restriction endonucleases is crucial for their proper function.
The X-ray structures of the specific and
non-cognate complexes of the restriction nuclease EcoRV are
considerably different suggesting significant differences in the
hydration and binding free energies. Nonetheless, the majority of
studies performed at pH 7.5, optimal for enzymatic activity, have found
a < 10-fold difference between EcoRV binding constants to the specific
and nonspecific sequences in the absence of divalent ions. We used a
recently developed self-cleavage assay to measure EcoRV-DNA competitive
binding and to evaluate the influence of water activity, pH and salt
concentration on the binding stringency of the enzyme in the absence of
divalent ions. We find the enzyme can readily distinguish specific and
nonspecific sequences. The relative specific-nonspecific binding
constant increases strongly with increasing neutral solute
concentration and with decreasing pH. The difference in number of
associated waters between specific and nonspecific DNA-EcoRV complexes
is consistent with the differences in the crystal structures. Despite
the large pH dependence of the sequence specificity, the osmotic
pressure dependence indicates little change in structure with pH. The
large osmotic pressure dependence means that measurement of protein-DNA
specificity in dilute solution cannot be directly applied to binding in
the crowded environment of the cell. In addition to divalent ions,
water activity and pH are key parameters that strongly modulate binding
specificity of EcoRV.

<>

<1>Sidorova, N.Y., Muradymov, S.R., Royal, R.M., Rau, D.C.
<2>Unusual DNA-Binding Kinetics of the Restriction Endonuclease EcoRV.
<3>Biophys. J.
<4>100
<5>72
<6>2011
<7>We have applied a self-cleavage assay, developed previously by us, to measure
EcoRV-DNA binding in solution. Self-cleavage assay monitors only enzymatically
competent complexes of the endonuclease. This technique does not have
the limitations of more commonly used gel mobility shift assay while providing
the same level of sensitivity.Wefound that the EcoRV has quite unusual kinetics
of specific complex formation in the absence of divalent ions that was not reported
previously. A significant fraction of the total enzyme, ~ 45%, forms enzymatically
competent complexes unusually slowly, especially at pH 7.6. This
novel result can be explained by a very slow transition between two conformations
of the free enzyme in solution. The equilibrium distribution of the slowly
and quickly associating protein structures and their exchange kinetics may depend
on many parameters including pH, salt, osmolytes, and divalent cations.
The observation of at least two kinetics components in association indicates
that EcoRV is an allosteric protein with at least two conformations. Allosterism
is now recognized as important concept for DNA-protein complexes, offering an
additional level of control over binding and activity. We are continuing our investigation
into the EcoRV structures responsible for the different kinetic classes
of association.

<>

<1>Sidorova, N.Y., Rau, D.C.
<2>Differences in water release for the binding of EcoRI to specific and nonspecific DNA sequences.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>12272-12277
<6>1996
<7>The free energy difference between complexes of the restriction nuclease EcoRI with
nonspecific DNA and with the enzyme's recognition sequence is linearly dependent on the water
chemical potential of the solution, set using several very different solutes, ranging from
glycine and glycerol to triethylene glycol and sucrose.  This osmotic dependence indicates
that the nonspecific complex sequesters some 110 waters more than the specific complex with
the recognition sequence.  The insensitivity of the difference in number of waters released to
the solute identity further indicates that this water is sequestered in a space that is
sterically inaccessible to solutes, most likely at the protein-DNA interface of the
nonspecific complex.  Calculations based on the structure of the specific complex suggest that
the apposing DNA and protein surfaces in the nonspecific complex retain approximately a full
hydration layer of water.

<>

<1>Sidorova, N.Y., Rau, D.C.
<2>Removing water from an EcoRI-noncognate DNA complex with osmotic stress.
<3>J. Biomol. Struct. Dyn.
<4>17
<5>19-31
<6>1999
<7>We recently showed that a nonspecific complex of the restriction nuclease EcoRI with poly
(dI-dC) sequesters significantly more water at the protein-DNA interface than the complex with
the specific recognition sequence. The nonspecific complex seems to retain almost a full
hydration layer at the interface. We now find that at low osmotic pressures a complex of the
restriction nuclease EcoRI with a DNA sequence that differs by only one base pair from the
recognition site (a 'star' sequence) sequesters about 70 waters more than the specific one,
a value virtually indistinguishable from nonspecific DNA. Unlike complexes with oligo (dI-dC)
or with a sequence that differs by two base pairs from the recognition sequence, however, much
of the water in the 'star' sequence complex is removed at high osmotic pressures. The energy
of removing this water can be calculated simply from the osmotic pressure work done on the
complex. The ability to measure not only the changes in water sequestered by DNA-protein
complexes for different sequences, but also the work necessary to remove this water is a
potentially powerful new tool for coupling inferred structural changes and thermodynamics.

<>

<1>Sidorova, N.Y., Rau, D.C.
<2>The dissociation rate of the EcoRI-DNA-specific complex is linked to water activity.
<3>Biopolymers
<4>53
<5>363-368
<6>2000
<7>In many respects, the dissociation rate constant of a DNA-protein or protein-protein complex
is as important a physical parameter as the equilibrium constant.  The regulation of most
cellular activities and developmental control are dynamic rather than static processes.  With
many techniques, the successful physiochemical characterization of a complex depends
ciritically on the lifetime of the complex during isolation or measurement.  With present
technologies, the very powerful, single molecule methods used for mapping the kinetic barriers
of complex dissociation reactions require lifetimes on the order of minutes.  We report here
that the dissociation rate of the specific complex between the restriction nuclease EcoRI and
its recognition DNA sequence is strongly dependent on water activity (in addition to its known
dependence on salt activity).  This observation means that the dissociation rate of complexes
in the crowded conditions found within cells cannot be straightforwardly predicted from dilute
solution measurements, even though salt, temperature, and pH conditions are fixed to those
found in vivo.  In addition, these results suggest a practical method to extend the lifetime
of "weak" complexes sufficiently to perform biophysical and biochemical characterizations.

<>

<1>Sidorova, N.Y., Rau, D.C.
<2>Linkage of EcoRI dissociation from its specific DNA recognition site to water activity, salt concentration, and pH: separating their roles in specific and non-specific binding.
<3>J. Mol. Biol.
<4>310
<5>801-816
<6>2001
<7>We have measured the dependencies of both the dissociation rate of specifically bound EcoRI
endonuclease and the ratio of non-specific and specific association constants on water
activity, salt concentration, and pH in order to distinguish the contributions of these
solution components to specific and non-specific binding. For proteins such as EcoRI that
locate their specific recognition site efficiently by diffusing along non-specific DNA, the
specific site dissociation rate can be separated into two steps: an equilibrium between
non-specific and specific binding of the enzyme to DNA, and the dissociation of
non-specifically bound protein. We demonstrated previously that the osmotic dependence of the
dissociation rate is dominated by the equilibrium between specific and non-specific binding
that is independent of the osmolyte nature. The remaining osmotic sensitivity linked to the
dissociation of non-specifically bound protein depends significantly on the particular
osmolyte used, indicating a change in solute-accessible surface area. In contrast, the
dissociation of non-specifically bound enzyme accounts for almost all the pH and
salt-dependencies. We observed virtually no pH-dependence of the equilibrium between specific
and non-specific binding measured by the competition assay. The observed weak salt-sensitivity
of the ratio of specific and non-specific association constants is consistent with an osmotic,
rather than electrostatic, action. The seeming lack of a dependence on viscosity suggests the
rate-limiting step in dissociation of non-specifically bound protein is a discrete
conformational change rather than a general diffusion of the protein away from the DNA.

<>

<1>Sidorova, N.Y., Rau, D.C.
<2>Self-footprint of the restriction endonuclease EcoRI.
<3>Biophys. J.
<4>78
<5>299A
<6>2000
<7>The dissociation rate of the EcoRI-DNA specific complex is strongly linked to water activity.
In the presence of osmolytes (betaine, sucrose, methyl glucoside, t-butanol, etc.) stability
of the specific EcoRI-DNA complex increases dramatically.  Osmotic stress technique offers a
practical way of manipulating dissociation rates controllably and to measure complex
properties on an experimentally convenient time scale.  It has been shown recently that
osmotic stress also slows down cleavage reaction of the EcoRI at its specific sequence.  We
believe this effect is at least partly due to the slowing of the EcoRI dissociation from the
product of the cleavage reaction.  Using osmotic stress technique we have developed a method
for using the nuclease activity of EcoRI to measure sensitively its specific binding.  We show
using gel shift assay that only those sites on DNA initially occupied by protein immediately
prior to adding Mg2+ are apparently cleaved.  The level of specific DNA fragment cleavage
remains constant for as long as 40 min.  This self-footprint assay allows measurement of EcoRI
binding equilibrium and kinetics avoiding intrinsic gel-shift and filter-binding assay
problems.  There is the possibility of using such a technique for any protein possessing DNA
cleavage activity given that its interactions with DNA are sensitive to osmotic stress.

<>

<1>Siebers, B., Zaparty, M., Raddatz, G., Tjaden, B., Albers, S.V., Bell, S.D., Blombach, F., Kletzin, A., Kyrpides, N., Lanz, C., Plagens, A., Rampp, M., Rosinus, A., von Jan, M., Makarova, K.S., Klenk, H.P., Schuster, S.C., Hensel, R.
<2>The Complete Genome Sequence of Thermoproteus tenax: A Physiologically Versatile Member of the Crenarchaeota.
<3>PLoS ONE
<4>6
<5>E24222
<6>2011
<7>Here, we report on the complete genome sequence of the hyperthermophilic
Crenarchaeum Thermoproteus tenax (strain Kra1, DSM 2078(T)) a type strain
of the crenarchaeotal order Thermoproteales. Its circular 1.84-megabase
genome harbors no extrachromosomal elements and 2,051 open reading frames
are identified, covering 90.6% of the complete sequence, which represents
a high coding density. Derived from the gene content, T. tenax is a
representative member of the Crenarchaeota. The organism is strictly
anaerobic and sulfur-dependent with optimal growth at 86 degrees C and pH
5.6. One particular feature is the great metabolic versatility, which is
not accompanied by a distinct increase of genome size or information
density as compared to other Crenarchaeota. T. tenax is able to grow
chemolithoautotrophically (CO/H) as well as chemoorganoheterotrophically
in presence of various organic substrates. All pathways for synthesizing
the 20 proteinogenic amino acids are present. In addition, two presumably
complete gene sets for NADH:quinone oxidoreductase (complex I) were
identified in the genome and there is evidence that either NADH or reduced
ferredoxin might serve as electron donor. Beside the typical archaeal
AA-ATP synthase, a membrane-bound pyrophosphatase is found, which might
contribute to energy conservation. Surprisingly, all genes required for
dissimilatory sulfate reduction are present, which is confirmed by growth
experiments. Mentionable is furthermore, the presence of two proteins
(ParA family ATPase, actin-like protein) that might be involved in cell
division in Thermoproteales, where the ESCRT system is absent, and of
genes involved in genetic competence (DprA, ComF) that is so far unique
within Archaea.

<>

<1>Siedlecki, P., Boy, R.G., Comagic, S., Schirrmacher, R., Wiessler, M., Zielenkiewicz, P., Suhai, S., Lyko, F.
<2>Establishment and functional validation of a structural homology model for human DNA methyltransferase 1.
<3>Biochem. Biophys. Res. Commun.
<4>306
<5>558-563
<6>2003
<7>Changes in DNA methylation patterns play an important role in tumorigenesis. The DNA
methyltransferase 1 (DNMT1) protein represents a
major DNA methyltransferase activity in human cells and is therefore a
prominent target for experimental cancer therapies. However, there are
only few available inhibitors and their high toxicity and low specificity
have so far precluded their broad use in chemotherapy. Based on the strong
conservation of catalytic DNA methyltransferase domains we have used a
homology modeling approach to determine the three-dimensional structure of
the DNMT1 catalytic domain. Our results suggest an overall structural
conservation with other DNA methyltransferases but also indicate local
conformational differences. To prove the validity of our model we used it
as a template to design a novel derivative of the known DNA
methyltransferase inhibitor 5-azacytidine. The resulting compound
(N4-fluoroacetyl-5-azacytidine) functioned as an efficient inhibitor of
DNA methylation in human tumor cell lines and also provides novel
opportunities for pharmacological applications.

<>

<1>Siedlecki, P., Boy, R.G., Musch, T., Brueckner, B., Suhai, S., Lyko, F., Zielenkiewicz, P.
<2>Discovery of two novel, small-molecule inhibitors of DNA methylation.
<3>J. Med. Chem.
<4>49
<5>678-683
<6>2006
<7>DNA methyltransferases are promising targets for cancer therapy. In many cancer cells
promoters of tumor suppressor genes are hypermethylated,
which results in gene inactivation. It has been shown that DNA
methyltransferase inhibitors can suppress tumor growth and have
significant therapeutic value. However, the established inhibitors are
limited in their application due to their substantial cytotoxicity. To
discover novel compounds for the inhibition of human DNA
methyltransferases, we have screened a set of small molecules available
from the NCI database. Using a 3-dimensional model of the human DNA
methyltransferase 1 and a modified docking and scoring procedure, we have
identified a small list of molecules with high affinities for the active
site of the enzyme. The two highest scoring structures were found to
inhibit DNA methyltransferase activity in vitro and in vivo. The newly
discovered inhibitors validate our screening procedure and also provide a
useful basis for further rational drug development.

<>

<1>Siegl, T., Petzke, L., Welle, E., Luzhetskyy, A.
<2>I-SceI endonuclease: a new tool for DNA repair studies and genetic manipulations in streptomycetes.
<3>Appl. Microbiol. Biotechnol.
<4>87
<5>1525-1532
<6>2010
<7>Actinomycetes are Gram-positive bacteria with a complex life cycle. They
produce many pharmaceutically relevant secondary metabolites, including
antibiotics and anticancer drugs. However, there is a limited number of
biotechnological applications available as opposed to genetic model
organisms like Bacillus subtilis or Escherichia coli. We report here a
system for the functional expression of a synthetic gene encoding the
I-SceI homing endonuclease in several streptomycetes. Using the synthetic
sce(a) gene, we were able to create controlled genomic DNA double-strand
breaks. A mutagenesis system, based on the homing endonuclease I-SceI, has
been developed to construct targeted, non-polar, unmarked gene mutations
in Streptomyces sp. Tu6071. In addition, we have shown that homologous
recombination is a major pathway in streptomycetes to repair an
I-SceI-generated DNA double-strand break. This novel I-SceI-based tool
will be useful in fundamental studies on the repair mechanism of DNA
double-strand breaks and for a variety of biotechnological applications.

<>

<1>Sievert, S.M., Scott, K.M., Klotz, M.G., Chain, P.S., Hauser, L.J., Hemp, J., Hugler, M., Land, M., Lapidus, A., Larimer, F.W., Lucas, S., Malfatti, S.A., Meyer, F., Paulsen, I.T., Ren, Q., Simon, J., USF, G.C.
<2>Genome of the epsilonproteobacterial chemolithoautotroph Sulfurimonas denitrificans.
<3>Appl. Environ. Microbiol.
<4>74
<5>1145-1156
<6>2008
<7>Sulfur-oxidizing epsilonproteobacteria are common in a variety of
sulfidogenic environments. These autotrophic and mixotrophic
sulfur-oxidizing bacteria are believed to contribute substantially to the
oxidative portion of the global sulfur cycle. In order to better
understand the ecology and roles of sulfur-oxidizing
epsilonproteobacteria, in particular those of the widespread genus
Sulfurimonas, in biogeochemical cycles, the genome of Sulfurimonas
denitrificans DSM1251 was sequenced. This genome has many features,
including a larger size (2.2 Mbp), that suggest a greater degree of
metabolic versatility or responsiveness to the environment than seen for
most of the other sequenced epsilonproteobacteria. A branched electron
transport chain is apparent, with genes encoding complexes for the
oxidation of hydrogen, reduced sulfur compounds, and formate and the
reduction of nitrate and oxygen. Genes are present for a complete,
autotrophic reductive citric acid cycle. Many genes are present that could
facilitate growth in the spatially and temporally heterogeneous sediment
habitat from where Sulfurimonas denitrificans was originally isolated.
Many resistance-nodulation-development family transporter genes (10 total)
are present; of these, several are predicted to encode heavy metal efflux
transporters. An elaborate arsenal of sensory and regulatory
protein-encoding genes is in place, as are genes necessary to prevent and
respond to oxidative stress.

<>

<1>Sievert, U.
<2>Determination of the specific recognition sequence of the restriction enzyme Rlu-1I from Rhizobium lupini.
<3>Ph.D. Thesis, Erlangen University, W. Germany
<4>
<5>1-109
<6>1982
<7>This is the enzyme RluI.

<>

<1>Siezen, R.J., Bayjanov, J., Renckens, B., Wels, M., van Hijum, S.A., Molenaar, D., van Hylckama, V.J.E.
<2>Complete genome sequence of Lactococcus lactis subsp. lactis KF147, a plant-associated lactic acid bacterium.
<3>J. Bacteriol.
<4>192
<5>2649-2650
<6>2010
<7>Lactococcus lactis is a lactic acid bacterium used in the production of many fermented dairy
products. We report the complete genome sequence of
L. lactis subsp. lactis KF147, a nondairy strain isolated from mung bean
sprouts. The circular chromosome of 2,598,144 bp, the largest among the
sequenced lactococcal strains, encodes many properties related to
adaptation to the plant environment.

<>

<1>Siezen, R.J., Francke, C., Renckens, B., Boekhorst, J., Wels, M., Kleerebezem, M., van Hijum, S.A.F.T.
<2>Complete resequencing and reannotation of the Lactobacillus plantarum WCFS1 genome.
<3>J. Bacteriol.
<4>194
<5>195-196
<6>2011
<7>There is growing interest in the beneficial effects of Lactobacillus plantarum on human
health. The genome of L. plantarum WCFS1, first sequenced in 2001, was resequenced using
Solexa technology. We identified 116 nucleotide corrections and improved function prediction
for nearly 1,200 proteins, with a focus on metabolic functions and cell surface-associated
proteins.

<>

<1>Siezen, R.J., Renckens, B., van Swam, I., Peters, S., van Kranenburg, R., Kleerebezem, M., de Vos, W.M.
<2>Complete Sequences of Four Plasmids of Lactococcus lactis subsp. cremoris SK11 Reveal Extensive Adaptation to the Dairy Environment.
<3>Appl. Environ. Microbiol.
<4>71
<5>8371-8382
<6>2005
<7>Lactococcus lactis strains are known to carry plasmids encoding industrially important traits.
L. lactis subsp. cremoris SK11 is widely used by the dairy industry in cheese making. Its
complete plasmid complement was sequenced and found to contain the plasmids pSK11A (10,372
bp), pSK11B (13,332 bp), pSK11L (47,165 bp), and pSK11P (75,814 bp). Six highly homologous
repB-containing replicons were found, all belonging to the family of lactococcal theta-type
replicons. Twenty-three complete insertion sequence elements segment the plasmids into
numerous modules, many of which can be identified as functional units or containing
functionally related genes. Plasmid-encoded functions previously known to reside on L. lactis
SK11 plasmids were now mapped in detail, e.g., lactose utilization (lacR-lacABCDFEGX), the
proteolytic system (prtM-prtP, pepO, pepF), and the oligopeptide permease system (oppDFBCA).
Newly identified plasmid-encoded functions could facilitate the uptake of various cations,
while the pabA and pabB genes could be essential for folate biosynthesis. A competitive
advantage could be obtained by using the putative flavin adenine dinucleotide-dependent
d-lactate dehydrogenase and oxalate:formate antiporter for enhanced ATP synthesis, while the
activity of the predicted alpha-acetolactate decarboxylase may contribute to the formation of
an additional electron sink. Various stress response proteins are plasmid encoded, which could
enhance strain robustness. A substantial number of these "adaptation" genes have not been
described before on L. lactis plasmids. Moreover, several genes were identified for the first
time in L. lactis, possibly reflecting horizontal gene transfer.

<>

<1>Siezen, R.J., Starrenburg, M.J.C., Boekhorst, J., Renckens, B., Molenaar, D., van Hylckama-Vlieg, J.E.T.
<2>Genome-scale genotype-phenotype matching of two Lactococcus lactis isolates from plants identifies mechanisms of adaptation to the plant niche.
<3>Appl. Environ. Microbiol.
<4>74
<5>424-436
<6>2008
<7>Lactococcus lactis is a primary constituent of many starter cultures used for the
manufacturing of fermented dairy products, but the species
also occurs in various nondairy niches such as (fermented) plant
material. Three genome sequences of L. lactis dairy strains (IL-1403,
SK11, and MG1363) are publicly available. An extensive molecular and
phenotypic diversity analysis was now performed on two L. lactis plant
isolates. Diagnostic sequencing of their genomes resulted in over 2.5
Mb of sequence for each strain. A high synteny was found with the
genome of L. lactis IL-1403, which was used as a template for contig
mapping and locating deletions and insertions in the plant L. lactis
genomes. Numerous genes were identified that do not have homologs in
the published genome sequences of dairy L. lactis strains. Adaptation
to growth on substrates derived from plant cell walls is evident from
the presence of gene sets for the degradation of complex plant polymers
such as xylan, arabinan, glucans, and fructans but also for the uptake
and conversion of typical plant cell wall degradation products such as
alpha-galactosides, P-glucosides, arabinose, xylose, galacturonate,
glucuronate, and gluconate. Further niche-specific differences are
found in genes for defense (nisin biosynthesis), stress response
(nonribosomal peptide synthesis and various transporters), and
exopolysaccharide biosynthesis, as well as the expected differences in
various mobile elements such as prophages, plasmids,
restrictionmodification systems, and insertion sequence elements. Many
of these genes were identified for the first time in Lactococcus
lactis. In most cases good correspondence was found with the phenotypic
characteristics of these two strains.

<>

<1>Sikora, A.E., Smith, J.R., Campbell, S.A., Firman, K.
<2>AFM protein-protein interactions within the EcoR124I molecular motor.
<3>Soft Matter
<4>8
<5>6358-6363
<6>2012
<7>Dynamic Force Spectroscopy (DFS), an Atomic Force Microscopy (AFM) technique, has been used to
investigate the interaction between the
HsdR subunit and the core methylase (MTase) of the Type I
Restriction-Modification (R-M) enzyme EcoR124I. Such systems are of
interest in bionanotechnology owing to their ability to translocate
DNA, thus acting as molecular motors. Forces between a glutathione
S-transferase (GST)-HsdR(PrrI) motor subunit attached to an AFM tip
using a polyethylene gycol linker and the core MTase on poly-L-lysine
pre-treated mica were measured at different loading rates. In the
absence of an applied force, the position of energy barrier x(diss),
bond dissociation rate k(diss)(0) and lifetime of the bond tau(0) were
calculated to be 1.35 +/- 0.17 nm, 0.16 s(-1) and 6.3 s, respectively.
The k(diss)(0) value was a little lower than that obtained from
magnetic tweezers (0.4 s(-1)), suggesting that the thermodynamic
equilibrium may be affected by the presence of DNA. This work
demonstrates that kinetic data concerning protein-protein interactions
between subunits within Type I R-M enzymes are accessible via AFM. Such
information is important for structure elucidation and the development
of nanodevices.

<>

<1>Sikorski, J. et al.
<2>Complete genome sequence of Mahella australiensis type strain (50-1 BON).
<3>Standards in Genomic Sciences
<4>4
<5>331-341
<6>2011
<7>Mahella australiensis Bonilla Salinas et al. 2004 is the type species of the genus Mahella,
which belongs to the family Thermoanaerobacteraceae. The species
is of interest because it differs from other known anaerobic spore-forming
bacteria in its G+C content, and in certain phenotypic traits, such as carbon
source utilization and relationship to temperature. Moreover, it has been
discussed that this species might be an indigenous member of petroleum and oil
reservoirs. This is the first completed genome sequence of a member of the genus
Mahella and the ninth completed type strain genome sequence from the family
Thermoanaerobacteraceae. The 3,135,972 bp long genome with its 2,974
protein-coding and 59 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Sikorski, J. et al.
<2>Complete genome sequence of Ilyobacter polytropus type strain (CuHbu1).
<3>Standards in Genomic Sciences
<4>3
<5>304-314
<6>2010
<7>Ilyobacter polytropus Stieb and Schink 1984 is the type species of the genus Ilyobacter, which
belongs to the fusobacterial family Fusobacteriaceae. The
species is of interest because its members are able to ferment quite a number of
sugars and organic acids. I. polytropus has a broad versatility in using various
fermentation pathways. Also, its members do not degrade poly-beta-hydroxybutyrate
but only the monomeric 3-hydroxybutyrate. This is the first completed genome
sequence of a member of the genus Ilyobacter and the second sequence from the
family Fusobacteriaceae. The 3,132,314 bp long genome with its 2,934
protein-coding and 108 RNA genes consists of two chromosomes (2 and 1 Mbp long)
and one plasmid, and is a part of the Genomic Encyclopedia of Bacteria and
Archaea project.

<>

<1>Sikorski, J. et al.
<2>Complete genome sequence of Sulfurospirillum deleyianum type strain (5175).
<3>Standards in Genomic Sciences
<4>2
<5>149-157
<6>2010
<7>Sulfurospirillum deleyianum Schumacher et al. 1993 is the type species of the genus
Sulfurospirillum. S. deleyianum is a model organism for studying sulfur
reduction and dissimilatory nitrate reduction as an energy source for growth.
Also, it is a prominent model organism for studying the structural and functional
characteristics of cytochrome c nitrite reductase. Here, we describe the features
of this organism, together with the complete genome sequence and annotation. This
is the first completed genome sequence of the genus Sulfurospirillum. The
2,306,351 bp long genome with its 2,291 protein-coding and 52 RNA genes is part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Sikorski, J. et al.
<2>Complete genome sequence of Sulfurimonas autotrophica type strain (OK10).
<3>Standards in Genomic Sciences
<4>3
<5>194-202
<6>2010
<7>Sulfurimonas autotrophica Inagaki et al. 2003 is the type species of the genus Sulfurimonas.
This genus is of interest because of its significant contribution
to the global sulfur cycle as it oxidizes sulfur compounds to sulfate and by its
apparent habitation of deep-sea hydrothermal and marine sulfidic environments as
potential ecological niche. Here we describe the features of this organism,
together with the complete genome sequence and annotation. This is the second
complete genome sequence of the genus Sulfurimonas and the 15(th) genome in the
family Helicobacteraceae. The 2,153,198 bp long genome with its 2,165
protein-coding and 55 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Sikorski, J. et al.
<2>Complete genome sequence of Acetohalobium arabaticum type strain (Z-7288).
<3>Standards in Genomic Sciences
<4>3
<5>57-65
<6>2010
<7>Acetohalobium arabaticum Zhilina and Zavarzin 1990 is of special interest because of its
physiology and its participation in the anaerobic C(1)-trophic chain in
hypersaline environments. This is the first completed genome sequence of the
family Halobacteroidaceae and only the second genome sequence in the order
Halanaerobiales. The 2,469,596 bp long genome with its 2,353 protein-coding and
90 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Sikorski, J. et al.
<2>Complete genome sequence of Segniliparus rotundus type strain (CDC 1076).
<3>Standards in Genomic Sciences
<4>2
<5>203-211
<6>2010
<7>Segniliparus rotundus Butler 2005 is the type species of the genus Segniliparus,  which is
currently the only genus in the corynebacterial family Segniliparaceae.
This family is of large interest because of a novel late-emerging genus-specific
mycolate pattern. The type strain has been isolated from human sputum and is
probably an opportunistic pathogen. Here we describe the features of this
organism, together with the complete genome sequence and annotation. This is the
first completed genome sequence of the family Segniliparaceae. The 3,157,527 bp
long genome with its 3,081 protein-coding and 52 RNA genes is part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Sikorski, J. et al.
<2>Complete genome sequence of Meiothermus silvanus type strain (VI-R2).
<3>Standards in Genomic Sciences
<4>3
<5>37-46
<6>2010
<7>Meiothermus silvanus (Tenreiro et al. 1995) Nobre et al. 1996 belongs to a thermophilic genus
whose members share relatively low degrees of 16S rRNA gene
sequence similarity. Meiothermus constitutes an evolutionary lineage separate
from members of the genus Thermus, from which they can generally be distinguished
by their slightly lower temperature optima. M. silvanus is of special interest as
it causes colored biofilms in the paper making industry and may thus be of
economic importance as a biofouler. This is the second completed genome sequence
of a member of the genus Meiothermus and only the third genome sequence to be
published from a member of the family Thermaceae. The 3,721,669 bp long genome
with its 3,667 protein-coding and 55 RNA genes is a part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Siksnys, V., Grazulis, S., Huber, R.
<2>Structure and function of the tetrameric restriction enzymes.
<3>Nucleic Acids Mol. Biol.
<4>14
<5>237-259
<6>2004
<7>Type II restriction endonucleases recognize specific DNA sequences, typically 4-8 bp in
length, and cleave phosphodiester bonds in the presence of Mg2+, within or close to their
recognition sites.  Around 3500 species, from variety of bacteria with nearly 240 differing
specificities, have now been characterized.  Most of the sequences recognized by Type II
restriction endonucleases are palindromic, i.e. possess a twofold rotational axis of symmetry.
On the basis of this observation, Kelly and Smith proposed the first model for the interaction
of these restriction enzymes with DNA.  According to their model, recognition of the
palindromic DNA sequence is achieved by two identical protein subunits related by a twofold
axis of symmetry.  Each subunit faces the same nucleotide sequence on the opposite DNA strand
and contains one active site.  Symmetrical nicking of opposite DNA strands by both monomers
within a homodimer generates the double-strand break.  Hence, the symmetry of the recognition
sequence implies the oligomeric state of the restriction enzyme.

<>

<1>Siksnys, V., Pleckaityte, M.
<2>Role of the reactive cysteine residue in restriction endonuclese Cfr9I.
<3>Biochim. Biophys. Acta
<4>1160
<5>199-205
<6>1992
<7>Chemical modification studies were performed to elucidate the role of Cys-residues in the
catalysis/binding of restriction endonuclease Cfr9I. Incubation of restriction endonuclease
Cfr9I with N-ethylmaleimide (NEM), iodoacetate, 5-5'-dithiobis (2-nitrobenzoic acids) at pH
7.5 led to a complete loss of the catalytic activity. However, no enzyme inactivation was
detectable after modification of the enzyme with iodoacetamide and methyl
methanethiosulfonate. Complete protection of the enzyme against inactivation by NEM was
observed in the presence of substrate implying that Cys-residues may be lcoated at or in the
vicinity of the active site of enzyme. Direct substrate-binding studies of native and modified
restriction endonuclease Cfr9I using a gel-mobility shift assay indicated that the
modification of the enzyme by NEM was hindered by substrate binding. A single Cys-residue was
modified during the titration of the enzyme with DTNB with concomitant loss of the catalytic
activity. The pH-dependence of inactivation of Cfr9I by NEM revealed the modification of the
residue with a pKa value of 8.9 +/- 0.2. The dependence of the reaction rate of substrate
hydrolysis by Cfr9I versus pH revealed two essential residues with pKa values of 6.3 +/- 0.15
and 8.7 +/- 0.15, respectively. The evidence presented suggests that the restriction
endonuclease Cfr9I contains a reactive sulfhydryl residue which is non-essential for
catalysis, but is located at or near the substrate binding site.

<>

<1>Siksnys, V., Pleckaityte, M.
<2>Catalytic and binding properties of restriction endonuclease Cfr9I.
<3>Eur. J. Biochem.
<4>217
<5>411-419
<6>1993
<7>The Cfr9I restriction endonuclease recognizes and cleaves the duplex DNA sequence C^CCGG. The
binding of restriction endonuclease Cfr9I to DNA was examined in the absence of Mg2+ using
gel-mobility-shift and nitrocellulose-filter-binding assays. It was shown that restriction
endonuclease Cfr9I bound DNA fragments either containing or lacking the canonical recognition
sequence with equal affinity. These results suggest that the specificity of restriction
endonuclease Cfr9I is expressed during the catalytic step. The cleavage of supercoiled pUC18
DNA by restriction endonuclease Cfr9I showed that at low concentrations of MgCl2, only with
open-circular DNA, nicks appeared in one strand at the recognition sequence, while the
cleavage of the second strand was very slow. At these conditions the supercoiled DNA was
converted to open-circular and linear forms simultaneously rather than consecutively. It was
shown that open-circular DNA was a poor substrate for restriction endonuclease Cfr9I. These
results suggested that both Mg2+ and intact recognition sequence are required to drive the
enzyme into the correct conformation to ensure DNA cleavage.

<>

<1>Siksnys, V., Skirgaila, R., Sasnauskas, G., Urbanke, C., Cherny, D., Grazulis, S., Huber, R.
<2>The Cfr10I restriction enzyme is functional as a tetramer.
<3>J. Mol. Biol.
<4>291
<5>1105-1118
<6>1999
<7>It is thought that most of the type II restriction endonucleases interact with DNA as
homodimers. Cfr10I is a typical type II restriction enzyme that recognises the 5'-Pu^CCGGPy
sequence and cleaves it as indicated by the arrow.  Gel-filtration and analytical
ultracentrifugation data presented here indicate that Cfr10I is a homotetramer in isolation.
Only the SfiI restriction enzyme that recognises the long interrupted recognition sequence
5'-GGCCNNNNNGGCC has been previously reported to operate as a tetramer however, its structure
is unknown. Analysis of Cfr10I crystals revealed that a single molecule in the asymmetric unit
is repeated by D2 symmetry to form a tetramer. To determine whether the packing of the Cfr10I
in the crystal reflects the quaternary structure of the protein in solution, the tryptophan
W220 residue located at the putative dimer-dimer interface was mutated to alanine, and the
structural and functional consequences of the substitution were analysed. Equilibrium
sedimentation experiments revealed that, in contrast to the wild-type Cfr10I, the W220A mutant
exists in solution predominantly as a dimer. In addition, the tetramer seems to be a
catalytically important form of Cfr10I, since the DNA cleavage activity of the W220A mutant is
< 0.1% of that of the wild-type enzyme. Further, analysis of plasmid DNA cleavage suggests
that the Cfr10I tetramer is able to interact with two copies of the recognition sequence,
located on the same DNA molecule. Indeed, electron microscopy studies demonstrated that two
distant recognition sites are brought together through the DNA looping induced by the
simultaneous binding of the Cfr10I tetramer to both sites. These data are consistent with the
tetramer being a functionally important form of Cfr10I.

<>

<1>Siksnys, V., Tamulaitis, G., Zaremba, M., Szczepanowski, R., Bochtler, M.
<2>How do restriction enzymes acquire specificities for different target sites?
<3>FEBS J.
<4>275
<5>10
<6>2008
<7>Restriction enzymes recognize 4-8 bp DNA sequences and cut at their target site with exquisite
specificity.  Many orthodox restriction endonucleases interacting with symmetric recognition
sites achieve sequence specificity making direct nucleobase-amino acid contacts with their
target sites.  Ecl18kI and EcoRII-C/PspGI restriction enzymes recognize interrupted
pentanucleotide sites CCNGG and CCWGG, respectively.  Structural studies demonstrate that
Ecl18kI/EcoRII-C/PspGI and the NgoMIV restriction enzyme specific for the non-interrupted
GCCGGC site share striking structural similarities and interact identically with the
symmetrical CC:GG half-sites.  It turned out that Ecl18kI specific for the CCNGG and
EcoRII-C/PspGI specific for the CCWGG site flip central nucleotides from DNA double helix to
interact with the conserved CC:GG parts of their target sequences and achieve cleavage.
However, Ecl18kI and EcoRII-C/PspGI accept different base pairs at the center, raising a
question whether the base pair strength/stability or a direct readout of the flipped out bases
determine the differences in specificity.  Using a combination of the X-ray structural
analysis and biochemical methods we show that EcoRII-C/PspGI achieve the specificity for the
CCWGG sequence by a double check mechanism by probing both the stability of the central base
pair and identity of the flipped out base in the pocket.

<>

<1>Siksnys, V., Timinskas, A., Klimasauskas, S., Butkus, V., Janulaitis, A.
<2>Sequence similarity among type-II restriction endonucleases, related by their recognized 6-bp target and tetranucleotide-overhang cleavage.
<3>Gene
<4>157
<5>311-314
<6>1995
<7>The type-II restriction endonucleases (ENases) EcoRI (recognition sequence G/AATTC), RsrI
(G/AATTC), XcyI (C/CCGGG), Cfr9I (C/CCGGG) and MunI (C/AATTG), all cleave hexanucleotide
palindromic sequences, leaving tetranucleotide 5'-overhangs.  Two regions of similarity that
appear in the same order and relative position were identified among the amino-acid sequences
of ENases.  These regions map to the structural elements of EcoRI involved in the building of
the catalytic site and in interactions with the central nucleotides of the recognized
sequence.  We propose that these ENases might all share a similar structural organization of
the active site and structural motifs involved in interactions with specific DNA recognition
sequences.

<>

<1>Siksnys, V., Zareckaja, N., Vaisvila, R., Timinskas, A., Stakenas, P., Butkus, V., Janulaitis, A.
<2>CAATTG-specific restriction-modification munI genes from Mycoplasma: sequence similarities between R.MunI and R.EcoRI.
<3>Gene
<4>142
<5>1-8
<6>1994
<7>The genes coding for the MunI restriction-modification (R-M) system, which recognize the
sequence 5'-CAATTG, have been cloned and expressed in Escherichia coli, and their nucleotide
sequences have been determined. The restriction endonuclease (ENase: R.MunI) is encoded by an
open reading frame (ORF) of 606 bp, and a 699-bp ORF codes for the methyltransferase (MTase).
The two genes are transcribed divergently from a 355-bp region. The gene encoding the ENase is
preceded by a short co-linear ORF of 222 bp. The deduced amino acid (aa) sequence of this
short ORF (SORF) closely resembles the sequences of a family of regulatory proteins that are
associated with other type-II R-M systems. Comparative analysis of the deduced aa sequence of
R.MunI revealed several regions of similarity to the EcoRI and RsrI ENases that recognize the
GAATTC sequence. The similar mode of interaction of MunI, EcoRI and RsrI with the
tetranucleotide AATT, common to the recognition sequences of these ENases, was suggested.

<>

<1>Silanskas, A., Foss, M., Wende, W., Urbanke, C., Lagunavicius, A., Pingoud, A., Siksnys, V.
<2>Photocaged Variants of the MunI and PvuII Restriction Enzymes.
<3>Biochemistry
<4>50
<5>2800-2807
<6>2011
<7>Regulation of proteins by light is a new and promising strategy for the external control of
biological processes. In this study, we demonstrate
the ability to regulate the catalytic activity of the MunI and PvuII
restriction endonucleases with light. We used two different approaches
to attach a photoremovable caging compound, 2-nitrobenzyl bromide
(NBB), to functionally important regions of the two enzymes. First, we
covalently attached a caging molecule at the dimer interface of Muni to
generate an inactive monomer. Second, we attached NBB at the DNA
binding site of the single-chain variant of PvuII (scPvuII) to prevent
binding and cleavage of the DNA substrate. Upon removal of the caging
group by UV irradiation, nearly 50% of the catalytic activity of Muni
and 80% of the catalytic activity of PvuII could be restored.

<>

<1>Silanskas, A., Zaremba, M., Sasnauskas, G., Siksnys, V.
<2>Catalytic Activity Control of Restriction Endonuclease-Triplex Forming Oligonucleotide Conjugates.
<3>Bioconjugate Chem.
<4>23
<5>203-211
<6>2012
<7>
<>

<1>Silber, K.R., Polisson, C., Rees, P.A., Benner, J.S.
<2>Cloning, purification and characterization of the M.NdeI methyltransferase from Neisseria denitrificans.
<3>Gene
<4>74
<5>43-44
<6>1988
<7>Meeting Abstract

<>

<1>Silberstein, Z., Shalit, M., Cohen, A.
<2>Heteroduplex strand-specificity in restriction-stimulated recombination by the RecE pathway of Escherichia coli.
<3>Genetics
<4>133
<5>439-448
<6>1993
<7>The RecE recombination pathway is active in Escherichia coli recB recC sbcA mutants. To
isolate and characterize products and intermediates of RecE-mediated, break-induced,
intramolecular recombination, we infected recB recC sbcA mutants, expressing EcoRI
endonuclease, with chimeric gamma phages that allow EcoRI-mediated release of cloned linear
recombination substrates. Substrates with direct terminal repeats recombined to yield a
circular product with one copy of the repeated sequence. Some recombinations were
heteroallelic for the recombining markers. Markers distant to the break were recovered in the
circular product at a higher frequency than markers close to the break. To examine the
heteroduplex structures that may have yielded the heteroallelic recombinants, non-replicative
substrates were employed. Some of the nonreplicative recombination products contained
heteroduplexes, with a strong bias for paired strands ending 3' at the break. This strand
bias in heteroduplex formation is consistent with recombination models that postulate
homologous pairing of protruding 3' single-stranded ends.

<>

<1>Silva, A. et al.
<2>Complete Genome Sequence of Corynebacterium pseudotuberculosis I-19, strain isolated from Israel Bovine mastitis.
<3>J. Bacteriol.
<4>193
<5>323-324
<6>2010
<7>This work portrays a report on complete and annotated genome sequence of Corynebacterium
pseudotuberculosis I-19 isolated from an Israel dairy cow
with sever clinical mastitis. To present the whole-genome sequence, de
novo assembly approach using 33 million short (25 bp) mate paired SOLiD
reads only was applied. More so, the automatic, functional and manual
annotation was attained with the use of several algorithms in a multi-step
process.

<>

<1>Silva, A.S., Barauna, R.A., de Sa, P.C., das Gracas, D.A., Carneiro, A.R., Thouvenin, M., Azevedo, V., Badell, E., Guiso, N., da Silva, A.L., Ramos, R.T.
<2>Draft Genome Sequence of Corynebacterium ulcerans FRC58, Isolated from the Bronchitic Aspiration of a Patient in France.
<3>Genome Announcements
<4>2
<5>e01132-13
<6>2014
<7>Corynebacterium ulcerans is a bacterial species with high importance because it causes
infections in animals and, rarely, in humans. Its virulence mechanisms
remain unclear. The current study describes the draft genome of C. ulcerans
FRC58, which was isolated from the bronchitic aspiration of a patient in France.

<>

<1>Silva, C.A., Blondel, C.J., Quezada, C.P., Porwollik, S., Andrews-Polymenis, H.L., Toro, C.S., Zaldivar, M., Contreras, I., McClelland, M., Santiviago, C.A.
<2>Infection of Mice by Salmonella enterica Serovar Enteritidis Involves Additional Genes That Are Absent in the Genome of Serovar Typhimurium.
<3>Infect. Immun.
<4>80
<5>839-849
<6>2012
<7>Salmonella enterica serovar Enteritidis causes a systemic, typhoid-like infection in newly
hatched poultry and mice. In the present study, a
library of 54,000 transposon mutants of S. Enteritidis phage type 4
(PT4) strain P125109 was screened for mutants deficient in the in vivo
colonization of the BALB/c mouse model using a microarray-based
negative-selection screening. Mutants in genes known to contribute to
systemic infection (e.g., Salmonella pathogenicity island 2 [SPI-2],
aro, rfa, rfb, phoP, and phoQ) and enteric infection (e.g., SPI-1 and
SPI-5) in this and other Salmonella serovars displayed colonization
defects in our assay. In addition, a strong attenuation was observed
for mutants in genes and genomic islands that are not present in S.
Typhimurium or in most other Salmonella serovars. These genes include a
type I restriction/modification system (SEN4290 to SEN4292), the peg
fimbrial operon (SEN2144A to SEN2145B), a putative pathogenicity island
(SEN1970 to SEN1999), and a type VI secretion system remnant SEN1001,
encoding a hypothetical protein containing a lysin motif (LysM) domain
associated with peptidoglycan binding. Proliferation defects for
mutants in these individual genes and in exemplar genes for each of
these clusters were confirmed in competitive infections with wild-type
S. Enteritidis. A Delta SEN1001 mutant was defective for survival
within RAW264.7 murine macrophages in vitro. Complementation assays
directly linked the SEN1001 gene to phenotypes observed in vivo and in
vitro. The genes identified here may perform novel virulence functions
not characterized in previous Salmonella models.

<>

<1>Silva, G.H., Belfort, M.
<2>Analysis of the LAGLIDADG interface of the monomeric homing endonuclease I-DmoI.
<3>Nucleic Acids Res.
<4>32
<5>3156-3168
<6>2004
<7>The general structural fold of the LAGLIDADG endonuclease family consists of two similar
alpha/beta domains (alpha beta beta alpha beta beta alpha) that assemble either as homodimers
or monomers with the domains related by pseudo-two-fold symmetry. At the center of this
symmetry is the closely packed LAGLIDADG two-helix bundle that forms the main inter- or
intra-molecular contact region between the domains of single- or double-motif proteins,
respectively. In this work, we further examine the role of the LAGLIDADG residues involved in
the helix?helix interaction. The interchangeability of the LAGLIDADG helix interaction was
explored by grafting interfacial residues from the homodimeric I-CreI into the corresponding
positions in the monomeric I-DmoI. The resulting LAGLIDADG exchange mutant is partially
active, preferring to nick dsDNA rather than making the customary double-strand break. A
series of partial revertants within the mutated LAGLIDADG region are shown to restore cleavage
activity to varying degrees resulting in one I-DmoI mutant that is more active than wild-type
I-DmoI. The phenotype of some of these mutants was reconciled on the basis of similarity to
the GxxxG helix interaction found in transmembrane proteins. Additionally, a split variant of
I-DmoI was created, demonstrating that the LAGLIDADG helices of I-DmoI are capable of forming
and maintaining the protein?protein interface in trans to create an active heterodimer.

<>

<1>Silva, G.H., Belfort, M., Wende, W., Pingoud, A.
<2>From monomeric to homodimeric endonucleases and back: Engineering novel specificity of LAGLIDADG enzymes.
<3>J. Mol. Biol.
<4>361
<5>744-754
<6>2006
<7>Monomeric homing endonucleases of the LAGLIDADG family recognize DNA in a bipartite manner,
reflecting the underlying structural assembly of two protein domains (A and B) related by
pseudo 2-fold symmetry. This architecture allows for changes in DNA specificity via the
distinct combination of these half-site domains. The key to engineering such hybrid proteins
lies in the LAGLIDADG two-helix bundle that forms both the domain interface and the
endonuclease active site. In this study, we utilize domain A of the monomeric I-DmoI to
demonstrate the feasibility of generating functional homodimeric endonucleases that recognize
palindromic DNA sequences derived from the original, non-palindromic target. Wild-type I-DmoI
domain A is capable of forming a homodimer (H-DmoA) that binds tightly to, but does not cleave
efficiently, its anticipated DNA target. Partial restoration of DNA cleavage ability was
obtained by re-engineering the LAGLIDADG dimerization interface (H-DmoC). Upon fusing two
copies of H-DmoC via a short peptide linker, a novel, site-specific DNA endonuclease was
created (H-DmoC2). Like I-DmoI, H-DmoC2 is thermostable and cleaves the new target DNA to
generate the predicted 4 nt 3'-OH overhangs but, unlike I-DmoI, H-DmoC2 retains stringent
cleavage specificity when substituting Mn(2+) for Mg(2+) as co-factor. This novel endonuclease
allows speculation regarding specificity of monomeric LAGLIDADG proteins, while it supports
the evolutionary genesis of these proteins by a gene duplication event.

<>

<1>Silva, G.H., Dalgaard, J.Z., Belfort, M., Van Roey, P.
<2>Crystal structure of the thermostable archaeal intron-encoded endonuclease I-DmoI.
<3>J. Mol. Biol.
<4>286
<5>1123-1136
<6>1999
<7>I-DmoI is a 22 kDa endonuclease encoded by an intron in the 23 S rRNA gene of the
hyperthermophilic archaeon Desulfurococcus mobilis.  The structure of I-DmoI has been
determined to 2.2 A resolution using multi-wave-length anomalous diffraction techniques.
I-DmoI, a protein of the LAGLIDADG motif family, represents the first structure of a
freestanding endonuclease with two LALIDADG motifs, and the first of a thermostable homing
endonuclease.  I-DmoI consists of two similar alpha/beta domains (alpha beta beta alpha beta
beta alpha) related by pseudo 2-fold symmetry.  The LAGLIDADG motifs are located at the
carboxy-terminal end of the first alpha-helix of each domain.  These helices form a two-helix
bundle at the interface between the domains and are perpendicular to a saddle-shaped DNA
binding surface, formed by two four-stranded antiparallel beta-sheets. Despite substantially
different sequences, the overall fold of I-DmoI is similar to that of two other LAGLIDADG
proteins for which the structures are known, I-CreI and the endonuclease domain of PI-SceI.
The three structures differ most in the loops connecting the beta-strands, relating to the
respective DNA target site sizes and geometries.  In addition, the absence of conserved
residues surrounding the active site, other than those within the LAGLIDADG motif, is of
mechanistic importance.  Finally, the carboxy-terminal domain of I-DmoI is smaller and has a
more irregular fold than the amino-terminal domain, which is more similar to I-CreI, a
symmetric homodimeric endonuclease.  This is reversed compared to PI-SceI, where the
amino-terminal domain is more similar to carboxy-terminal domain of I-DmoI and to I-CreI, with
interesting evolutionary implications.

<>

<1>Simala-Grant, J.L., Lam, E., Keelan, M., Taylor, D.E.
<2>Characterization of the DNA adenine 5 '-GATC-3 ' methylase HpyIIIM from Helicobacter pylori.
<3>Curr. Microbiol.
<4>49
<5>47-54
<6>2004
<7>The effect of inactivation of the 5'-GATC-3' methylase HpyIIIM in Helicobacter pylori (H.
pylori) on mismatch repair, adherence, and in
vitro fitness was examined. Chromosomal DNA from 90 H. pylori strains
was isolated, and restriction enzyme digestion indicated all strains
examined possess HpyIIIM. Wild-type H. pylori and a strain with an
inactive HpyIIIM were found to have rifampicin mutation frequencies of
2.93 x 10(-7) and 1.05 x 10(-7) (P > 0.05), respectively, indicating
that HpyIIIM does not appear to be important in mismatch repair.
Adherence of H. pylori in an in vitro model cell system was also
unaffected by inactivation of HpyIIIM. Inactivation of HpyIIIM did not
result in a decrease in fitness, as determined by liquid in vitro
competition experiments.

<>

<1>Simbochova, G., Timko, J., Zelinkova, E., Zelinka, J.
<2>Properties and recognition sequence of site-specific endonuclease from Streptomyces aureofaciens.
<3>Biologia (Bratisl)
<4>41
<5>357-365
<6>1986
<7>A type II restriction endonuclease Sau3239I was purified from Streptomyces
aureofaciens 3239.  The recognition sequence for the enzyme was determined 5' -
C ^ T C G A G - 3' 3' - G A G C T ^ C - 5' 	and cleaved at the position
indicated by the arrows, producing a tetranucleotide 5'-terminal extension.
Therefore Sau 3239I endonuclease is an isoschizomer of XhoI endonuclease.	The
contribution of the type II restriction enzymes to recombinant DNA technology
has stimulated extensive search for these enzymes which have been isolated from
a great variety of bacteria, including the actinomycetes {Streptomyces
aureofaciens IKA 18/4} is SauI {Timko et al., 1981}.  Another specific
restriction endonuclease was found in the chlortetracycline {CTC} producing
prototrophic strain of Streptomyces aureofaciens CCM 3239 {Gasperik et al.,
1983}.  We report here this endonuclease designated as Sau 3239I which was
isolated from this strain.  The isolation procedure and determination of the
recognition sequence are described.

<>

<1>Simbochova, G., Timko, J., Zelinkova, E., Zelinka, J.
<2>Some physical and chemical properties of site-specific endonuclease Sau3239I from Streptomyces aureofaciens.
<3>Biologia (Bratisl)
<4>42
<5>1129-1136
<6>1987
<7>Sau3239I was isolated and purified from Streptomyces aureofaciens CCM 3239. The
substrate specificity and the cleavage site of this enzyme is 5' - C^TCGAG - 3'
and thus it is an isoschizomer of XhoI (Simbochova et al., 1986).  Some of its
physical properties and substrate specificity are described.

<>

<1>Simcox, T.G., Fabian, L., Kretz, K., Hedden, V., Simcox, M.E.C.
<2>SanDI, a new type-II restriction endonuclease that recognizes 5'-GG/GWCCC-3'.
<3>Gene
<4>155
<5>129-130
<6>1995
<7>A new restriction endonuclease (ENase), SanDI, has been isolated from an unidentified species
of Streptomyces. SanDI recognizes the 7-bp interrupted palindrome 5'-GG/GWCCC-3' (W=A or T)
and cleaves double-stranded DNA after the second G in the sequence, producing 3-nt long 5'
protruding ends. SanDI is a rare-cutting ENase and should therefore be useful for megabase
mapping and vector constructions.

<>

<1>Simcox, T.G., Marsh, S.J., Gross, E.A., Lernhardt, W., Davis, S., Simcox, M.E.C.
<2>SrfI, a new type-II restriction endonuclease that recognizes the octanucleotide sequence,.
<3>Gene
<4>109
<5>121-123
<6>1991
<7>A new restriction endonuclease, SrfI, has been isolated from an unidentified
species of Streptomyces.  SrfI recognizes the 8-bp palindrome, 5'-GCCCGGGC and
cleaves double-stranded DNA after the third C in the sequence, producing blunt
ends.  SrfI is a rare-cutting enzyme and should therefore be useful for
megabase mapping.

<>

<1>Simcox, T.G., Simcox, M.E.
<2>Octanucleotide restriction endonuclease, SrfI, and method for producing the same.
<3>US Patent Office
<4>US 5300432
<5>
<6>1994
<7>*
This invention provides restriction endonuclease, SrfI, capable of recognizing the eight
nucleotide base palindromic sequence shown below on a double-stranded DNA molecule and
cleaving the DNA chain at the asterisk-marked position resulting in blunt ends

      5' GCCC*GGGC 3'
      3' CGGG*CCCG 5'

(wherein C and G respectively represent cytosine and guanine). The restriction endonuclease is
produced by culturing Streptomyces rf in a culture medium and recovering it from the culture.


<>

<1>Simcox, T.G., Simcox, M.E.
<2>Octanucleotide restriction endonuclease, SrfI, and method for producing same.
<3>US Patent Office
<4>US 6054307
<5>
<6>2000
<7>A new purified restriction endonuclease SrfI (RE) (I) produced by a host cell (from
Streptomyces sp.) transformed with an expression vector
is claimed. (I) containing a polynucleotide which encodes RE, operably
linked with a promoter has specificity for and a cleavage site in
double stranded (ds) sequence 5'GCCC/GGGC3', in which / represents the
Srfl cleavage site in the ds nucleotide sequence. The enzyme is used in
molecular biology techniques for DNA identification and in mammalian
genome analysis. The enzyme is also used for manipulating the DNA
molecule and analyzing the resulting construction. For example, eight
Streptomyces bacterial strains were screened for endonuclease
activities. The endonuclease assays were performed with FPLC. It was
noted that five possible palindromic recognition sequences were
identified. Of these possible sequences, three eight bp recognition
sequences were found along with one ten bp and one twelve bp
palindrome.

<>

<1>Simmon, V.F., Lederberg, S.
<2>Degradation of bacteriophage lambda deoxyribonucleic acid after restriction by Escherichia coli K-12.
<3>J. Bacteriol.
<4>112
<5>161-169
<6>1972
<7>Wild-type bacteria which restrict the deoxyribonucleic acid (DNA) of infecting
phage when the phage do not carry the proper host modification rapidly degrade
that restricted DNA to acid-soluble products.  The purified restriction enzyme
acts as an endonuclease in vitro to cleave restrictable DNA and does not
further degrade the DNA fragment products.  We have examined mutants of
Escherichia coli K-12 which lack various nucleases in order to determine which
nucleases are involved in the rapid acid solubilization in vivo of unmodified
lambda DNA following restriction.  Bacteria which are wild type, recA-, or
polA1- degrade about 50% of the unmodified phage DNA within 10 min of
infection, with little subsequent degradation.  Mutants which are recB- or
recC- degrade unmodified DNA very slowly, solubilizing about 15% of the DNA by
10 min after infection.  Two classes of phenotypic revertants of recB-/C-
mutants were also tested.  Bacteria which are sbcA- restrict poorly and do not
degrade much of the restricted DNA.  Bacteria which are sbcB- restrict
normally.  This mutation does not appear to affect degradation of restricted
phage DNA in recB-/C- mutants, but such degradation is decreased in recB+/C+
bacteria.  The presence of a functional lambda exonuclease gene is not required
for degradation after restriction.

<>

<1>Simoliunas, E., Kaliniene, L., Truncaite, L., Klausa, V., Zajanckauskaite, A., Meskys, R.
<2>Genome of Klebsiella sp.-Infecting Bacteriophage vB_KleM_RaK2.
<3>J. Virol.
<4>86
<5>5406
<6>2012
<7>Despite the fact that multidrug-resistant Klebsiella sp. strains emerge rapidly
(Xu J, et al., Adv. Mater. Res. 268-270:1954-1956, 2011) and bacteriophages have
been reported to be useful in controlling these bacteria (Kumari S, Harjai K,
Chhibber S, J. Med. Microbiol. 60:205-210, 2011), the complete genome sequences
of only five Klebsiella phages (four siphoviruses and one myovirus) can be found
in databases. In this paper, we report on the complete genome sequence of
Klebsiella sp.-infecting bacteriophage vB_KleM_RaK2. With a genome size of
345,809 bp, this is the second largest myovirus and the largest Klebsiella phage
sequenced to date. This phage differs substantially from other myoviruses since
411 out of 534 vB_KleM_RaK2 open reading frames have no known functions and lack
any reliable database matches. Comparative analysis of the genome sequence of
vB_KleM_RaK2 suggests that this phage forms a distinct phylogenetic branch within
the family Myoviridae of tailed bacteriophages.

<>

<1>Simon, D., Grunert, F., Acken, U.V., Doring, H.P., Kroger, H.
<2>DNA-methylase from regenerating rat liver: purification and characterisation.
<3>Nucleic Acids Res.
<4>5
<5>2153-2167
<6>1978
<7>DNA methylase has been purified 660-fold from nuclei from regenerating rat liver. The enzyme
is able to methylate single stranded (ss) and double stranded (ds) DNA, the only reaction
product being 5-methylcytosine. Previously unmethylated double stranded DNA from prokaryotes
(M. luteus) as well as from eukaryotes (Ascaris suis) can serve as substrates. The synthetic
copolymers (dG-dC)n. (dC-dG)n and (dG,dC)n are also methylated. While SV40 DNA is almost not
methylated, PM2 DNA is a good substrate even in the supercoiled form. The enzyme methylates 1
in 17 bases in heterologous M. luteus DNA, but only 1 in 590 in homologous rat liver DNA. The
high methylation level of M. luteus DNA, an analysis of the methylated pyrimidine isostichs
and a preliminary dinucleotide analysis suggest that all the CpGs in a DNA can be methylated.

<>

<1>Simoncsits, A., Tjornhammar, M.L., Rasko, T., Kiss, A., Pongor, S.
<2>Covalent joining of the subunits of a homodimeric type II restriction endonuclease: single-chain PvuII endonuclease.
<3>J. Mol. Biol.
<4>309
<5>89-97
<6>2001
<7>The PvuII restriction endonuclease has been converted from its natural homodimeric form into a
single polypeptide chain by tandemly linking
the two subunits through a short peptide Linker. The arrangement of the
single-chain PvuII (sc PvuII) is (2-157)-GlySerGlyGly-(2-157), where
(2-157) represents the amino acid residues of the enzyme subunit and
Gly-SerGlyGly is the peptide linker. By introducing the corresponding
tandem gene into Escherichia coli, PvuII endonuclease activity could
be detected in functional in vivo assays. The sc enzyme was expressed
at high level as a soluble protein. The purified enzyme was shown to
have the molecular mass expected for the designed sc protein. Based on
the DNA cleavage patterns obtained with different substrates, the
cleavage specificity of the sc PvuII is indistinguishable from that of
the wild-type (wt) enzyme. The sc enzyme binds specifically to the
cognate DNA site under non-catalytic conditions, in the presence of
Ca2+, with the expected 1:1 stoichiometry. Under standard catalytic
conditions, the sc enzyme cleaves simultaneously the two DNA strands in
a concerted manner. Steady-state kinetic parameters of DNA cleavage by
the sc and wt PvuII showed that the sc enzyme is a potent, but somewhat
less efficient catalyst; the k(cat)/K-M values are 1.11 x 10^9 and
3.50 x 10^9 min^-1 M^-1 for the sc and wt enzyme, respectively. The
activity decrease is due to the lower turnover number and to the lower
substrate affinity. The sc arrangement provides a facile route to
obtain asymmetrically modified heterodimeric enzymes.

<>

<1>Simons, M., Diffin, F.M., Szczelkun, M.D.
<2>ClpXP protease targets long-lived DNA translocation states of a helicase-like motor to cause restriction alleviation.
<3>Nucleic Acids Res.
<4>42
<5>12082-12091
<6>2014
<7>We investigated how Escherichia coli ClpXP targets the helicase-nuclease (HsdR) subunit of the
bacterial Type I restriction-modification enzyme EcoKI during restriction alleviation (RA). RA
is a temporary reduction in endonuclease activity that occurs when Type I enzymes bind
unmodified recognition sites on the host genome. These conditions arise upon acquisition of a
new system by a naive host, upon generation of new sites by genome rearrangement/mutation or
during homologous recombination between hemimethylated DNA. Using recombinant DNA and proteins
in vitro, we demonstrate that ClpXP targets EcoKI HsdR during dsDNA translocation on circular
DNA but not on linear DNA. Protein roadblocks did not activate HsdR proteolysis. We suggest
that DNA translocation lifetime, which is elevated on circular DNA relative to linear DNA, is
important to RA. To identify the ClpX degradation tag (degron) in HsdR, we used bioinformatics
and biochemical assays to design N- and C-terminal mutations that were analysed in vitro and
in vivo. None of the mutants produced a phenotype consistent with loss of the degron,
suggesting an as-yet-unidentified recognition pathway. We note that an EcoKI nuclease mutant
still produces cell death in a clpx- strain, consistent with DNA damage induced by unregulated
motor activity.

<>

<1>Simons, M., Szczelkun, M.D.
<2>Recycling of protein subunits during DNA translocation and cleavage by Type I restriction-modification enzymes.
<3>Nucleic Acids Res.
<4>39
<5>7656-7666
<6>2011
<7>The Type I restriction-modification enzymes comprise three protein subunits; HsdS and HsdM
that form a methyltransferase (MTase) and HsdR
that associates with the MTase and catalyses Adenosine-5'-triphosphate
(ATP)-dependent DNA translocation and cleavage. Here, we examine whether
the MTase and HsdR components can 'turnover' in vitro, i.e. whether they
can catalyse translocation and cleavage events on one DNA molecule,
dissociate and then re-bind a second DNA molecule. Translocation
termination by both EcoKI and EcoR124I leads to HsdR dissociation from
linear DNA but not from circular DNA. Following DNA cleavage, the HsdR
subunits appear unable to dissociate even though the DNA is linear,
suggesting a tight interaction with the cleaved product. The MTases of
EcoKI and EcoAI can dissociate from DNA following either translocation or
cleavage and can initiate reactions on new DNA molecules as long as free
HsdR molecules are available. In contrast, the MTase of EcoR124I does not
turnover and additional cleavage of circular DNA is not observed by
inclusion of RecBCD, a helicase-nuclease that degrades the linear DNA
product resulting from Type I cleavage. Roles for Type I restriction
endonuclease subunit dynamics in restriction alleviation in the cell are
discussed.

<>

<1>Simpson, A.J. et al.
<2>The genome sequence fo the plant pathogen Xylella fastidiosa.
<3>Nature
<4>406
<5>151-157
<6>2000
<7>Xylella fastidiosa is a fastidious, xylem-limited bacterium that causes a range of
economically important plant diseases.  Here we report the complete genome sequence of X.
fastidiosa clone 9a5c, which causes citrus variegated chlorosis-a serious disease of orange
trees.  The genome comprises a 52.7% GC-rich 2,679,305-base-pair circular chromosome and two
plasmids of 51,158 bp and 1,285 bp.  We can assign putative functions to 47% of the 2,904
predicted coding regions.  Efficient metabolic functions are predicted, with sugars as the
principal energy and carbon source, supporting existence in the nutrient-poor xylem sap.  The
mechanisms associated with pathogenicity and virulence involve toxins, antibiotics and ion
sequestration systems, as well as bacterium-bacterium and bacterium-host interactions mediated
by a range of proteins.  Orthologues of some of these proteins have only been identified in
animal and human pathogens; their presence in X. fastidiosa indicates that the molecular basis
for bacterial pathogenicity is both conserved and independent of host.  At least 83 genes are
bacteriophage-derived and include virulence-associated genes from other bacteria, providing
direct evidence of phage-mediated horizontal gene transfer.

<>

<1>Sims, D. et al.
<2>Complete genome sequence of Kytococcus sedentarius type strain (541).
<3>Standards in Genomic Sciences
<4>1
<5>12-20
<6>2009
<7>Kytococcus sedentarius (ZoBell and Upham 1944) Stackebrandt et al. 1995 is the type strain of
the species, and is of phylogenetic interest because of its
location in the Dermacoccaceae, a poorly studied family within the
actinobacterial suborder Micrococcineae. Kytococcus sedentarius is known for the
production of oligoketide antibiotics as well as for its role as an opportunistic
pathogen causing valve endocarditis, hemorrhagic pneumonia, and pitted
keratolysis. It is strictly aerobic and can only grow when several amino acids
are provided in the medium. The strain described in this report is a free-living,
nonmotile, Gram-positive bacterium, originally isolated from a marine
environment. Here we describe the features of this organism, together with the
complete genome sequence, and annotation. This is the first complete genome
sequence of a member of the family Dermacoccaceae and the 2,785,024 bp long
single replicon genome with its 2639 protein-coding and 64 RNA genes is part of
the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Simuth, J., Trnovsky, J., Jelokova, J.
<2>Inhibition of bacterial DNA-dependent RNA polymerases and restriction endonuclease by UV-absorbing components from Propolis.
<3>Pharmazie
<4>41
<5>131-132
<6>1986
<7>Several UV-absorbing substances inhibiting the DNA-dependent RNA polymerases of
Escherichia coli and Streptomyces aureofaciens, as well as the restriction
endonuclease EcoRI have been isolated from the water-soluble extract of
Propolis by two-dimensional paper chromatography.  The inhibition of bacterial
RNA-polymerases by the components of Propolis was probably due to the loss of
their ability to bind to DNA.  The general characteristic of the UV-absorbing
component of Propolis with the most pronounced inhibitory effect upon
transcription in vitro is described.

<>

<1>Sineva, E.V., Zakharova, G.G., Tarutina, Z.E., Kravetz, A.N., Solonin, A.S.
<2>Class II restriction-modification systems in Enterobacter cloacae.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>10-13
<6>1993
<7>Various clinical strains of Enterobacter cloacae have been examined for the presence of
site-specific endonuclease activities. Type II restriction endonucleases have been isolated
from 6 strains. Recognition sequences for all of these enzymes have been determined and the
cleavage sites were identified for two of them. The enzymes proved to be isoschizomers of
EcoRII and PstI. Restriction endonucleases Ecl2zI and Ecl37kI recognize the nucleotide
sequences CTGCA/G and are true isoschizomers of PstI. The genes for Ecl54kI and Ecl57kI
restriction modification systems (isoschizomers of EcoRII) were found to be located on the
IncN group of plasmids, whereas the genes for Ecl2zI and Ecl699kI seem to be located on the
chromosomes of the host cells.

<>

<1>Sing, W.D., Klaenhammer, T.R.
<2>Characterization of a recombinant plasmid, pTRK12, encoding restriction/modification and proteolytic activities in lactic streptococci.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>88
<5>147
<6>1988
<7>Restriction and modification (R+/M+) activities are widely distributed in
lactic streptococci and often associated with plasmid DNA.  A
lactose-fermenting (Lac+) transconjugant (designated Streptococcus lactis
NCK40) was isolated from matings between S. cremoris TDM1 and a plasmid-free
recipient.  NCK40 contained a plasmid approximating 100 kb (pTRK11) that
correlated with Lac+ and conjugal transfer ability (Tra+) and a 13 kb plasmid
(pTRK10) which was linked to proteolytic activity (Prt+).  S. lactis NCK40
exhibited phage restriction and modification; the efficiency of plaquing (EOP)
for phage c2 was 10-3.  Two types of Lac- derivatives of NCK40, both cured of
pTRK11, were isolated.  One exhibited R-/M- Prt+ and contained pTRK10.  The
second type was R+/M+ Prt+ and contained a new 30 kb plasmid (pTRK12).  pTRK10
and pTRK11 were not detected in the R+/M+ Prt+ variant.  Restriction analyses
demonstrated that pTRK12 (R+/M+ Prt+) contained all detectable regions of
pTRK10 (Prt+).  Hybridization experiments using a 32P-pTRK11 probe identified
regions of pTRK12 that originated from the 100 kb plasmid, pTRK11.  These data
demonstrated that formation of a new 30 kb plasmid encoding R+/M+ and Prt+ was
the result of recombination events between pTRK10 and pTRK11 occurring upon
destabilization of Lac+.

<>

<1>Sing, W.D., Klaenhammer, T.R.
<2>Characterization of restriction-modification plasmids from Lactococcus lactis ssp. cremoris and their effects when combined with pTR2030.
<3>J. Dairy Sci.
<4>74
<5>1133-1144
<6>1991
<7>Three different restriction-modification plasmids (pTRK12, pTRK30, pTRK317)
were isolated from an industrial starter strain, Lactococcus lactis ssp.
cremoris TDM1.  A lactose-fermenting transconjugant, Lactococcus lactis ssp.
lactis NCK40, was isolated from matings between L. lactis ssp. cremoris TDM1
and a plasmid-free recipient.  The NCK40 transmissible plasmid (pTRK11)
encoding for restriction modification and a 13.5-kb plasmid (pTRK10) encoding
proteolytic activity.  Following isolation of lactose-negative derivatives from
NCK40, a 30.5-kb plasmid, pTRK12, was identified that encoded proteolytic and
restriction-modification of the identical specificity as pTRK11.  Restriction
analyses and hybridization experiments indicated that pTRK12 contained
sequences from pTRK11 and all of pTRK10.  Cotransformation of total plasmid DNA
from L. lactis ssp. cremoris TDM1 with vector pVS2 identified two other
restriction-modification plasmids, pTRK30 (28.0 kb) and pTRK317 (15.5 kb).
Efficiencies of plaquing from phage c2 on restriction-modification
transconjugants and transformants was 10/2 to 10/4.  The specificity of
restriction-modification activities conferred by each of the three plasmids was
different.  When the abortive infection plasmid pTR2030 was combined with
pTRK30, both phage inhibition phenotypes were expressed.  However, when pTR2030
was combined with pTRK12, the abortive infection phenotype was not fully
expressed.  Significant cell death occurred when abortive infection cells
containing only pTR2030 were infected with phage.  Combining the
restriction-modification system of pTRK30 with pTR2030 significantly improved
cell survival following phage infection.  Operation of restriction-modification
systems in conjunction with the abortive defense mechanism maximized cell
survival.  The data suggest that cell death is minimized when the lytic cycle
is halted by restriction before abortive infection responses induce phage
abortion and kill the cell.

<>

<1>Sing, W.D., Klaenhammer, T.R.
<2>Conjugal transfer of bacteriophage resistance determinants on pTR2030 into Streptococcus cremoris strains.
<3>Appl. Environ. Microbiol.
<4>51
<5>1264-1271
<6>1986
<7>Agar surface conjugal matings were used to introduce heat-sensitive phage
resistance (hsp+) determinants carried on the conjugal plasmid pTR2030 into
Streptococcus cremoris KH, HP, 924, and TDM1.  Lactose-fermenting (Lac+)
transconjugants were selected from matings of Lac- variants of S. cremoris KH,
HP, 924, and TDM1 with Streptococcus lactis ME2 or a high-frequency donor, S.
lactis T-EK1 (pTR1040, Lac+; pTR2030, Hsp+).  For all of the S. cremoris
strains examined, select Lac+ transconjugants were completely resistant to
plaquing by their homologous lytic phages.  In all cases the plaquing
efficiencies were less than 10-9.  Acquisition of a 30-megadalton plasmid
(pTR2030) in the S. cremoris phage-resistant transconjugants was demonstrated
by direct plasmid analysis, by hybridization with 32P-labeled probes, or by
conugal transfer of pTR2030 out of the phage-resistant transconjugants into a
plasmid-cured recipient, S. lactis LM2302.  Acid production, coagulation
ability, and proteolytic activity of phage-resistant transconjugants in milk
were comparable to those of their phage-sensitive parents.  Further, S.
cremoris phage-resistant transconjugants were not attacked by phage in starter
culture activity tests, which included a 40C incubation period.  The results
demonstrated that phage resistance determinants on pTR2030 could be conjugally
transferred to a variety of S. cremoris strains and confer resistance to phage
under conditions encountered during cheese manufacture.  Phage-resistant
transconjugants of S. cremoris M43 and HP were also constructed without the use
of antibiotic markers to select conjugal recipients from mating mixtures.

<>

<1>Sing, W.D., Klaenhammer, T.R.
<2>Characteristics of phage abortion conferred in lactococci by the conjugal plasmid pTR2030.
<3>J. Gen. Microbiol.
<4>136
<5>1807-1815
<6>1990
<7>The effect of pTR2030 on phage DNA injection, transfection, release of progeny phage, and cell
death was evaluated for a number of lactococcal phages. Infection by prolate phage c2 and
small isometric phage p2 of derivatives of Lactococcus lactis LM2301 with or without pTR2030,
and infection by small isometric phage phi31 of derivatives of L. lactis NCK202 with or
without pTR2030 was studied. Phage DNA injection was not affected by pTR2030 when examined
using blender-resistant-complex assays with 32P-labelled DNA or by observaton of phage
labelled with the fluorescent dye 4',6-diamidino-2-phenylindole(DAPI). Successful
transfection of hosts bearing pTR2030 indicated that the plasmid did not retard passage of
naked phage DNA across the membrane. Infective-centre assays were used to determine whether
progeny were released from phage-infected pTR2030 hosts that do not support plaque formation
by small isometric phages. In all cases, pTR2030 reduced the number of infected hosts which
generated viable phage. When progeny were released, the phage burst size was reduced. The data
confirmed that pTR2030 interferes with development of prolate and small isometric phages in a
similar manner via a classical abortive infection mechanism.

<>

<1>Sing, W.D., Klaenhammer, T.R.
<2>A strategy for rotation of different bacteriophage defenses in a lactococcal single-strain started culture system.
<3>Appl. Environ. Microbiol.
<4>59
<5>365-372
<6>1993
<7>A new strategy for starter culture rotations was developed for a series of phage-resistant
clones genetically derived from a single strain of Lactococcus lactis subsp. lactis.
Phage-resistant derivatives carrying different defense systems were constructed via
conjugatiion with various plasmids encoding abortive infection (Abi/Hsp) and/or restriction
and modification (R/M) systems of different specificity. The plasmids included pTR2030 (Hsp+
R+/M+),pTN20 (Abi+ R+/M+), pTRK11 (R+/M+), and pTRK68 (R+/M+). Selected phage-resistant
transconjugants or transformants were evaluated in different rotation sequences through cycles
of the Heap-Lawrence starter culture activity test in milk contaminated with phage and when
from the previous cycle. When used in consecutive sequences, derivative strains carrying the
R/M systems encoded by pTN20, pTRK11, and pTRK68 retarded phage development when the initial
levels of phage contamination were below 102 PFU/ml but not when levels were increased to 103
PFU/ml. Use of a derivative bearing pTR2030 (Hsp+ R+/M+) at the beginning of the rotation
prevented phage development, even when the inital levels of phage contamination were high (106
PFU/ml). Alternating the type and specificity of R/M and Abi defenses through the rotation
prevented phage proliferation and in some cases eliminated contaminting phages. A model
rotation sequence for the phage defense rotation strategy was developed and performed
sucessfully over nine cycles of the Heap-Lawrence starter culture activity test in the
presence of high-titer commercial phage composites. This phage defense rotation strategy is
designed to protect a highly specilized Lactococcus strain from phage attack during continuous
and extended use in the dairy industry.

<>

<1>Sing, W.D., Klaenhammer, T.R.
<2>Plasmid-induced abortive infection in Lactococci: a review.
<3>J. Dairy Sci.
<4>73
<5>2239-2251
<6>1990
<7>The longevity of mesophilic lactococci in dairy fermentations depends to a large extent on
whether or not the strains carry effective phage-resistance mechanisms.  Among the different
systems that exist in lactococci, abortive infection is highly significant because it is the
cell's strongest defense against the phages that most often disrupt cheese making.
Phage-resistant strains that carry plasmids encoding abortive defenses have already been
constructed using genetic strategies.  These strains have been used successfully in commercial
cheese making since 1986.  Still, our knowledge of the molecular mechanisms underlying
abortive infection and the means through which it retards phage development is limited.  This
review addresses abortive infection in lactococci relative to similar phenomena in other
bacteria.  Further understanding of the abortive infection process should accelerate genetic
efforts to strengthen this phage defense as well as facilitate efforts to combine it with
other mechanisms in the construction of specialized strains that are insensitive to attack by
phage.

<>

<1>Singh, A., Jangir, P.K., Kumari, C., Sharma, R.
<2>Genome Sequence of Nitratireductor aquibiodomus Strain RA22.
<3>J. Bacteriol.
<4>194
<5>6307
<6>2012
<7>The genus Nitratireductor represents nitrate-reducing bacteria from the family
Phyllobacteriaceae. Here we report the draft genome sequence of Nitratireductor
aquibiodomus strain RA22, which contains 4,592,790 bp, with a G+C content of
61.30%, and has 4,241 protein coding genes.

<>

<1>Singh, A., Kumar, J.P., Sharma, R., Singh, A., Kumar, P.A., Shivaji, S.
<2>Draft Genome Sequence of Indibacter alkaliphilus Strain LW1T, Isolated from Lonar Lake, a Haloalkaline Lake in the Buldana District of Maharashtra, India.
<3>Genome Announcements
<4>1
<5>e00513-13
<6>2013
<7>We report the 5.0-Mb genome sequence of Indibacter alkaliphilus strain LW1(T), isolated from a
haloalkaline crater lake in the Buldana district, Maharashtra,
India.

<>

<1>Singh, A., Sreenivas, A., Sathyanarayana, R.G., Pinnaka, A.K., Shivaji, S.
<2>Draft Genome Sequence of Lutibaculum baratangense Strain AMV1T, Isolated from a Mud Volcano in Andamans, India.
<3>Genome Announcements
<4>2
<5>e00735-14
<6>2014
<7>The 4.3-Mb genome of Lutibaculum baratangense strain AMV1(T), isolated from a soil sample
collected from a mud volcano in Andamans, India, is reported. The
draft genome of strain Lutibaculum baratangense AMV1(T) consists of 4,300,776 bp
with a G+C content of 66.93 mol% and 4,198 predicted coding regions, including 56
RNAs.

<>

<1>Singh, A., Zubko, E., Meyer, P.
<2>Cooperative activity of DNA methyltransferases for maintenance of symmetrical and non-symmetrical cytosine methylation in Arabidopsis thaliana.
<3>Plant J.
<4>56
<5>814-823
<6>2008
<7>Maintenance of cytosine methylation in plants is controlled by three DNA methyltransferases.
MET1 maintains CG methylation, and DRM1/2 and
CMT3 act redundantly to enforce non-CG methylation. RPS, a repetitive
hypermethylated DNA fragment from Petunia hybrida, attracts DNA
methylation when transferred into Petunia or other species. In
Arabidopsis thaliana, which does not contain any RPS homologues, RPS
transgenes are efficiently methylated in all sequence contexts. To test
which DNA methylation pathways regulate RPS methylation, we examined
maintenance of RPS methylation in various mutant backgrounds.
Surprisingly, CG methylation was lost in a drm1/2/cmt3 mutant, and
non-CG methylation was almost completely eliminated in a met1 mutant.
An unusual cooperative activity of all three DNA methyltransferases is
therefore required for maintenance of both CG and non-CG methylation in
RPS. Other unusual features of RPS methylation are the independence of
its non-CG methylation from the RNA-directed DNA methylation (RdDM)
pathway and the exceptional maintenance of methylation at a CC(m)TGG
site in some epigenetic mutants. This is indicative of activity of a
methylation system in plants that may have evolved from the DCM
methylation system that controls CC(m)WGG methylation in bacteria. Our
data suggest that strict separation of CG and non-CG methylation
pathways does not apply to all target regions, and that caution is
required in generalizing methylation data obtained for individual
genomic regions.

<>

<1>Singh, A.K., Chaudhary, P., Macwan, A.S., Diwedi, U.N., Kumar, A.
<2>Selective loss of lin genes from hexachlorocyclohexane-degrading Pseudomonas aeruginosa ITRC-5 under different growth conditions.
<3>Appl. Microbiol. Biotechnol.
<4>76
<5>895-901
<6>2007
<7>The chlorinated insecticide ã-hexachlorocyclohexane
(ã-HCH) is sequentially metabolized by the
products of linA, linB, linC, linD, linE, and linF genes to
â-ketoadipate, which is subsequently mineralized. Two or
more copies of these genes are present in the bacterium
Pseudomonas aeruginosa ITRC-5 that was isolated earlier
by selective enrichment on technical-HCH. At least one
copy of linA, linB, linC, linD, and possibly linE is lost from
ITRC-5 upon its growth on ã-HCH. All the lin genes,
however, are lost when the bacterium was grown in Luria-
Bertani (LB) medium. The loss of lin genes is accompanied
with the loss/rearrangement of insertion sequence IS6100
genes. Concomitant to the loss of lin genes, the degradation
of HCH-isomers by "ã-HCH grown cells" is slower, when
compared with "technical-HCH grown cells", and is
completely lost by "LB-grown cells". The selective loss
of lin genes during different growth conditions has not been
reported before and is expected to help in understanding the
dynamism of degradative genes.

<>

<1>Singh, A.K., Sangwan, N., Sharma, A., Gupta, V., Khurana, J.P., Lal, R.
<2>Draft Genome Sequence of Sphingobium quisquiliarum Strain P25T, a Novel Hexachlorocyclohexane (HCH)-Degrading Bacterium Isolated from an HCH Dumpsite.
<3>Genome Announcements
<4>1
<5>e00717-13
<6>2013
<7>Here, we report the draft genome sequence (4.2 Mb) of Sphingobium quisquiliarum strain P25(T),
a natural lin (genes involved in degradation of hexachlorocyclohexane [HCH] isomers) variant
genotype, isolated from a heavily contaminated (450 mg HCH/g of soil) HCH dumpsite.

<>

<1>Singh, I., Beuck, C., Bhattacharya, A., Hecker, W., Parmar, V.S., Weinhold, E., Seitz, O.
<2>Abasic site stabilization by aromatic DNA base surrogates: High-affinity binding to a base-flipping DNA-methyltransferase.
<3>Pure Appl. Chem.
<4>76
<5>1563-1570
<6>2004
<7>DNA-methyltransferases catalyze the sequence-specific transfer of the methyl group of
S-adenosylmethionine to target bases in genomic DNA.
For gaining access to their target embedded within a double-helical
structure, DNA-methyltransferases (DNA-MTases) rotate the target base
out of the DNA helix. This base-flipping leads to the formation of an
apparent abasic site. MTases such as cytosine-specific M.HhaI and
M.HaeIII and also the repair enzyme uracil DNA glycosylase (UDG)
insert amino acid side chains into the opened space and/or rearrange
base-pairing. The adenine-specific DNA MTase M.TaqI binds without
amino acid insertion. This binding mode allows for a substitution of
the orphaned thymine with larger DNA base surrogates without steric
interference by inserted amino acid side chains. DNA containing
pyrenyl, naphthyl, acenaphthyl, and biphenyl residues was tested in
M.TaqI binding studies. The synthesis of DNA building blocks required
the formation of a C-glycosidic bond, which was established by using
protected 1-chloro-2-deoxyribose as glycosyl donor and organocuprates
as glycosyl acceptors. It is shown that all of the base surrogates
enhanced the binding affinity to M.TaqI. Incorporation of pyrene
increased the binding affinity by a factor of 400. Interestingly, there
is a correlation between the observed order of dissociation constants
and the ability of a base surrogate to stabilize abasic sites in model
duplexes.

<>

<1>Singh, J., Klar, A.J.S.
<2>Active genes in budding yeast display enhanced in vivo accessibility to foreign DNA methylases: a novel in vivo probe for chromatin structure of yeast.
<3>Genes Dev.
<4>6
<5>186-196
<6>1992
<7>Unlike higher eukaryotes, where an inverse correlation has been generally
observed between gene expression and methylation of CpG sites, the budding
yeast Saccharomyces cerevisiae lacks DNA methylation.  Gene regulatory
mechanisms can function independently of DNA methylation in yeast, and yeast
strains expressing foreign DNA methylases that modify adenine and CpG residues
have been found to be viable.  We have used such strains to determine whether
the transcriptional status of genes can influence the level of their DNA
methylation in vivo.  Several genes were tested, for example, GAL1, -7, and
-10, PHO5, HMRa and HML alpha, and STE2 and STE3.  Surprisingly, we found that
all the genes displayed several fold more methylation in the expressed state as
compared to the repressed state.  This procedure serves as a novel in vivo
probe for the chromatin structure of yeast and potentially for higher
eukaryotes.

<>

<1>Singh, M.P., Lown, J.W.
<2>Inhibitory effects of a GC-sequence- and minor groove-binding Hoechst 33258 analogue on DNA cleavage by selected restriction endonucleases.
<3>J. Biomol. Struct. Dyn.
<4>12
<5>A220
<6>1995
<7>In continuance of our research efforts in the area of employing the important features of
ligand-DNA molecular recognition in expanding the design and development of minor groove
binding class of compounds, we recently reported a bis(pyridoimidazole) analogue of Hoechst
33258 and demonstrated its preference for selective binding to a GC-rich sequence.  Such
compounds offer an approach to the experimental manipulation of sequence-specific protein-DNA
interactions.  A convenient way to test this hypothesis is to look for the effects on the
DNA-cleavage activity of restriction enzymes with recognition sites that are comparable in
size and nature to the ligand binding sites.  We have observed distinct inhibitory effects of
the bis(pyridoimidazole) compound on the cleavage of EcoRI-linearized pBR322 DNA in individual
agarose gel electrophoresis assays by three selected endonucleases, MscI (at 5'-TGGCCA),
NruI (at 5'TCGCGA), and FspI (at 5'-TGCGCA).  The extent to which this minor groove binding
ligand provides protection of the GC-rich sites towards restriction cleavage can be ranked, in
qualitative terms, as TGGCCA >> TCGCGA > TGCGCA.  It is important to recognize that the
restriction enzymes are widely believed to interact through the major groove of their cognate
DNA sequences and the observed effects presumably arise from changes in the helix conformation
brought about by the minor groove binding ligand.

<>

<1>Singh, N., Duenas-Gonzalez, A., Lyko, F., Medina-Franco, J.L.
<2>Molecular Modeling and Molecular Dynamics Studies of Hydralazine with Human DNA Methyltransferase 1.
<3>ChemMedChem
<4>4
<5>792-799
<6>2009
<7>DNA methyltransferases (DNMTs) are a family of enzymes that methylate DNA at the C5 position
of cytosine residues and their inhibition is a
promising strategy for the treatment of various developmental and
proliferative diseases, particularly cancers. In the present study, a
binding model for hydralazine, with a validated homology model of human
DNMT, was developed by the use of automated molecular docking and
molecular dynamics simulations. The docking protocol was validated by
predicting the binding mode of 2'-deoxycytidine, 5-azacytidine, and
5-aza-2'-deoxycytidine. The inhibitory activity of hydralazine toward
DNMT may be rationalized at the molecular level by similar interactions
within the binding pocket (e.g., by a similar pharmacophore) as
established by substrate-like deoxycytidine analogues. These
interactions involve a complex network of hydrogen bonds with arginine
and glutamic acid residues that also play a major role in the mechanism
of DNA methylation. Despite the different scaffolds of other
non-nucleoside DNMT inhibitors such as procaine and procainamide, the
current modeling work reveals that these drugs exhibit similar
interactions within the DNMT1 binding site. These findings are valuable
in guiding the rational design and virtual screening of novel DNMT
inhibitors.

<>

<1>Singh, N.K., Kumar, S., Raghava, G.P., Mayilraj, S.
<2>Draft Genome Sequence of Acinetobacter baumannii Strain MSP4-16.
<3>Genome Announcements
<4>1
<5>e0013713
<6>2013
<7>We report the 4.0-Mb draft genome sequence of Acinetobacter baumannii strain MSP4-16, isolated
from a mangrove soil sample from Parangipettai (11 degrees
30'N, 79 degrees 47'E), Tamil Nadu, India. The draft genome sequence of strain
MSP4-16 consists of 3,944,542 bp, with a G+C content of 39%, 5,387 protein coding
genes, and 69 RNAs.

<>

<1>Singh, N.N., Lambowitz, A.M.
<2>Interaction of a group II intron ribonucleoprotein endonuclease with its DNA target site investigated by DNA footprinting and modification interference.
<3>J. Mol. Biol.
<4>309
<5>361-386
<6>2001
<7>Group II intron mobility occurs by a target DNA-primed reverse transcription mechanism in
which the intron RNA reverse splices directly into one strand of a double-stranded DNA target
site, while the intron-encoded protein cleaves the opposite strand and uses it as a primer to
reverse transcribe the inserted intron RNA. The group II intron endonuclease, which mediates
this process, is an RNP particle that contains the intron-encoded protein and the excised
intron RNA and uses both cooperatively to recognize DNA target sequences. Here, we analyzed
the interaction of the Lactococcus lactis Ll.LtrB group II intron endonuclease with its DNA
target site by DNA footprinting and modification-interference approaches. In agreement with
previous mutagenesis experiments showing a relatively large target site, DNase I protection
extends from position -25 to +19 from the intron-insertion site on the top strand and from -28
to +16 on the bottom strand. Our results suggest that the protein first recognizes a small
number of specific bases in the distal 5'-exon region of the DNA target site via major-groove
interactions. These base interactions together with additional phosphodiester-backbone
interactions along one face of the helix promote DNA unwinding, enabling the intron RNA to
base-pair to DNA top-strand positions -12 to +3 for reverse splicing. Notably, DNA unwinding
extends to at least position +6, somewhat beyond the region that base-pairs with the intron
RNA, but is not dependent on interaction of the conserved endonuclease domain with the 3'
exon. Bottom-strand cleavage occurs after reverse splicing and requires recognition of a small
number of additional bases in the 3' exon, the most critical being T+5 in the now
single-stranded downstream region of the target site. Our results provide the first detailed
view of the interaction of a group II intron endonuclease with its DNA target site. Copyright
2001 Academic Press.

<>

<1>Singh, P., Kumari, R., Mukherjee, U., Saxena, A., Sood, U., Lal, R.
<2>Draft Genome Sequence of Rifamycin Derivatives Producing Amycolatopsis mediterranei Strain DSM 46096/S955.
<3>Genome Announcements
<4>2
<5>e00837-14
<6>2014
<7>Amycolatopsis mediterranei DSM 46096 produces antibiotics of the rifamycin family,
27-demethoxy-27-hydroxyrifamycin B,
25-desacetyl-27-demethoxy-27-hydroxyrifamycin, and
27-demethoxy-27-hydroxyrifamycin SV, which are effective against Gram-negative
bacteria. Here, we present the draft genome of A. mediterranei 46096 (approx.
10.2 Mbp) having 104 contigs with a GC content of 71.3% and 9,382 coding
sequences.

<>

<1>Singh, P., Springman, A.C., Davies, H.D., Manning, S.D.
<2>Whole-Genome Shotgun Sequencing of a Colonizing Multilocus Sequence Type 17 Streptococcus agalactiae Strain.
<3>J. Bacteriol.
<4>194
<5>6005
<6>2012
<7>This report highlights the whole-genome shotgun draft sequence for a Streptococcus agalactiae
strain representing multilocus sequence type (ST) 17,
isolated from a colonized woman at 8 weeks postpartum. This sequence represents
an important addition to the published genomes and will promote comparative
genomic studies of S. agalactiae recovered from diverse sources.

<>

<1>Singh, P., Tripathi, P., Muniyappa, K.
<2>Mutational analysis of active-site residues in the Mycobacterium leprae RecA intein, a LAGLIDADG homing endonuclease: Asp(122) and Asp(193) are  crucial to the double-stranded DNA cleavage activity whereas Asp(218) is not.
<3>Protein Sci.
<4>19
<5>111-123
<6>2010
<7>Mycobacterium leprae recA harbors an in-frame insertion sequence that encodes an intein homing
endonuclease (PI-MleI). Most inteins (intein
endonucleases) possess two conserved LAGLIDADG (DOD) motifs at their
active center. A common feature of LAGLIDADG-type homing endonucleases
is that they recognize and cleave the same or very similar DNA
sequences. However, PI-MleI is distinctive from other members of the
family of LAGLIDADG-type HEases for its modular structure with
functionally separable domains for DNA-binding and cleavage, each with
distinct sequence preferences. Sequence alignment analyses of PI-MleI
revealed three putative LAGLIDADG motifs; however, there is conflicting
bioinformatics data in regard to their identity and specific location
within the intein polypeptide. To resolve this conflict and to
determine the active-site residues essential for DNA target site
recognition and double-stranded DNA cleavage, we performed
site-directed mutagenesis of presumptive catalytic residues in the
LAGLIDADG motifs. Analysis of target DNA recognition and kinetic
parameters of the wild-type PI-MleI and its variants disclosed that the
two amino acid residues, Asp(122) (in Block C) and Asp(193) (in
functional Block E), are crucial to the double-stranded DNA
endonuclease activity, whereas Asp(218) (in pseudo-Block E) is not.
However, despite the reduced catalytic activity, the PI-MleI variants,
like the wild-type PI-MleI, generated a footprint of the same length
around the insertion site. The D122T variant showed significantly
reduced catalytic activity, and D122A and D193A mutations although
failed to affect their DNA-binding affinities, but abolished the
double-stranded DNA cleavage activity. On the other hand, D122C variant
showed approximately twofold higher double-stranded DNA cleavage
activity, compared with the wild-type PI-MleI. These results provide
compelling evidence that Asp(122) and Asp(193) in DOD motif I and II,
respectively, are bona fide active-site residues essential for DNA
cleavage activity. The implications of these results are discussed in
this report.

<>

<1>Singh, P., Tripathi, P., Silva, G.H., Pingoud, A., Muniyappa, K.
<2>Characterization of Mycobacterium leprae RecA Intein, a LAGLIDADG Homing Endonuclease, Reveals a Unique Mode of DNA Binding, Helical Distortion, and Cleavage Compared with a Canonical LAGLIDADG Homing Endonuclease.
<3>J. Biol. Chem.
<4>284
<5>25912-25928
<6>2009
<7>Mycobacterium leprae, which has undergone reductive evolution leaving behind a minimal set of
essential genes, has retained intervening
sequences in four of its genes implicating a vital role for them in the
survival of the leprosy bacillus. A single in-frame intervening
sequence has been found embedded within its recA gene. Comparison of
the M. leprae recA intervening sequence with the known intervening
sequences indicated that it has the consensus amino acid sequence
necessary for being a LAGLIDADG-type homing endonuclease. In light of
massive gene decay and function loss in the leprosy bacillus, we sought
to investigate whether its recA intervening sequence encodes a
catalytically active homing endonuclease. Here we show that the
purified M. leprae RecA intein (PI-MleI) binds to cognate DNA and
displays endonuclease activity in the presence of alternative divalent
cations, Mg2+ or Mn2+. A combination of approaches, including four
complementary footprinting assays such as DNase I,
copper-phenanthroline, methylation protection, and KMnO4, enhancement
of 2-aminopurine fluorescence, and mapping of the cleavage site
revealed that PI-MleI binds to cognate DNA flanking its insertion site,
induces helical distortion at the cleavage site, and generates two
staggered double strand breaks. Taken together, these results implicate
that PI-MleI possesses a modular structure with separate domains for
DNA target recognition and cleavage, each with distinct sequence
preferences. From a biological standpoint, it is tempting to speculate
that our findings have implications for understanding the evolution of
the LAGLIDADG family of homing endonucleases.

<>

<1>Singh, R.N., Saldanha, R.J., D'Souza, L.M., Lambowitz, A.M.
<2>Binding of a group II intron-encoded reverse transcriptase/maturase to its high affinity intron RNA binding site involves sequence-specific recognition and autoregulates translation.
<3>J. Mol. Biol.
<4>318
<5>287-303
<6>2002
<7>Mobile group II introns encode reverse transcriptases that bind specifically to the intron
RNAs to promote both intron mobility and RNA splicing (maturase activity). Previous studies
with the Lactococcus lactis Ll.LtrB intron suggested a model in which the intron-encoded
protein (LtrA) binds first to a primary high-affinity binding site in intron subdomain DIVa,
an idiosyncratic structure at the beginning of the LtrA coding sequence, and then makes
additional contacts with conserved regions of the intron to fold the RNA into the
catalytically active structure. Here, we analyzed the DIVa binding site by iterative in vitro
selection and in vitro mutagenesis. Our results show that LtrA binds to a small region at the
distal end of DIVa that contains the ribosome-binding site and initiation codon of the LtrA
open reading frame. The critical elements are in a small stem-loop structure emanating from a
purine-rich internal loop, with both sequence and structure playing a role in LtrA
recognition. The ribosome-binding site falls squarely within the LtrA-binding region and is
sequestered directly by the binding of LtrA or by stabilization of the small stem-loop or
both. Finally, by using LacZ fusions in Escherichia coli, we show that the binding of LtrA to
DIVa down-regulates translation. This mode of regulation limits accumulation of the
potentially deleterious intron-encoded protein and may facilitate splicing by halting ribosome
entry into the intron. The recognition of the DIVa loop-stem-loop structure accounts, in part,
for the intron specificity of group II intron maturases and has parallels in
template-recognition mechanisms used by other reverse transcriptases.

<>

<1>Singh, S.V., Kumar, N., Singh, S.N., Bhattacharya, T., Sohal, J.S., Singh, P.K., Singh, A.V., Singh, B., Chaubey, K.K., Gupta, S., Sharma, N., Kumar, S., Raghava, G.P.
<2>Genome Sequence of the 'Indian Bison Type' Biotype of Mycobacterium avium subsp.  paratuberculosis Strain S5.
<3>Genome Announcements
<4>1
<5>e00005-13
<6>2013
<7>We report the 4.79-Mb genome sequence of the 'Indian Bison Type' biotype of subsp. strain
S5, isolated from a terminally sick Jamunapari goat at the CIRG
(Central Institute for Research on Goats) farm in India. This draft genome will
help in studying novelties of this biotype, which is widely distributed in
animals and human beings in India.

<>

<1>Singh, T.R., Pardasani, K.R.
<2>In silico Analysis of Evolutionary Patterns in Restriction Endonucleases.
<3>In Silico Biology
<4>9
<5>45-53
<6>2009
<7>Restriction endonucleases represent one of the best studied examples of DNA binding proteins.
Type II restriction endonucleases recognize short
sequences of foreign DNA and cleave the target on both strands with
remarkable sequence specificity. Type II restriction endonucleases are
part of restriction modification systems. Restriction modification
systems occur ubiquitously among bacteria and archaea. Restriction
endonucleases are indispensable tools in molecular biology and
biotechnology. They are important model system for specific
protein-nucleic acid interactions and also serve as good example for
investigating structural, functional and evolutionary relationships
among various biomolecules. The interaction between restriction
endonucleases and their recognition sequences plays a crucial role in
biochemical activities like catalytic site/metal binding, DNA repair
and recombination etc. We study various patterns in restriction
endonucleases type II and analyzed their structural, functional and
evolutionary role. Our studies support X-ray crystallographic studies,
arguing for divergence and molecular evolution. Conservation patterns
of the nuclease superfamily have also been analyzed by estimating
site-specific evolutionary rates for the analyzed structures related to
respective chains in this study.

<>

<1>Sinha, D., Bakhshi, M.R., Vora, R.K., Kirby, E.P., Budzynski, A.Z.
<2>Engineering DNA and protein chimeras utilizing coding sequences of restriction sites.
<3>Anal. Biochem.
<4>238
<5>205-208
<6>1996
<7>Various engineering strategies have been employed so far to generate chimeric proteins for the
purpose of structure-function studies.  None of these methods, except splicing by overlap
extension (SOE)3, however, can be considered as providing a general methodology for generating
chimeras.  SOE is a powerful technique considering its simplicity and versatility.  One
limitation of the SOE procedure, however, is the difficulty in amplification when the chimera
is large.  In the present report, we describe a procedure for generating chimeras by altering
the codon sequences of one amino acid pair of the seven around the junction using the PCR
technique.  The altered nucleotide sequence of the amino acid pair creates a restriction site
that is utilized for formation of the desired chimera.  This procedure, like the SOE method,
eliminates the need for single-stranded template and viral vector intermediates, and thus does
not require a cloning and screening step.  However, in engineering relatively long chimeras,
the described procedure is more advantageous because this method amplifies smaller fragments.
The present technique is simple and 100% efficient.

<>

<1>Sinha, D., Shamayeva, K., Ramasubramani, V., Reha, D., Bialevich, V., Khabiri, M., Guzanova, A., Milbar, N., Weiserova, M., Csefalvay, E., Carey, J., Ettrich, R.
<2>Interdomain communication in the endonuclease/motor subunit of type I restriction-modification enzyme EcoR124I.
<3>J. Mol. Model.
<4>20
<5>2334
<6>2014
<7>Restriction-modification systems protect bacteria from foreign DNA. Type I
restriction-modification enzymes are multifunctional heteromeric complexes with DNA-cleavage
and ATP-dependent DNA translocation activities located on endonuclease/motor subunit HsdR. The
recent structure of the first intact motor subunit of the type I restriction enzyme from
plasmid EcoR124I suggested a mechanism by which stalled translocation triggers DNA cleavage
via a lysine residue on the endonuclease domain that contacts ATP bound between the two
helicase domains. In the present work, molecular dynamics simulations are used to explore this
proposal. Molecular dynamics simulations suggest that the Lys-ATP contact alternates with a
contact with a nearby loop housing the conserved QxxxY motif that had been implicated in DNA
cleavage. This model is tested here using in vivo and in vitro experiments. The results
indicate how local interactions are transduced to domain motions within the endonuclease/motor
subunit.

<>

<1>Siozios, S., Cestaro, A., Kaur, R., Pertot, I., Rota-Stabelli, O., Anfora, G.
<2>Draft Genome Sequence of the Wolbachia Endosymbiont of Drosophila suzukii.
<3>Genome Announcements
<4>1
<5>e00032-13
<6>2013
<7>is one of the most successful and abundant symbiotic bacteria in nature, infecting more than
40% of the terrestrial arthropod species. Here we report the
draft genome sequence of a novel strain named 'Suzi' that was retrieved from the
genome sequencing of its host, the invasive pest .

<>

<1>Siqueira, F.M., Thompson, C.E., Virginio, V.G., Gonchoroski, T., Reolon, L., Almeida, L.G., da Fonseca, M.M., de Souza, R., Prosdocimi, F., Schrank, I.S., Ferreira, H.B., de Vasconcelos, A.T., Zaha, A.
<2>New insights on the biology of swine respiratory tract mycoplasmas from a comparative genome analysis.
<3>BMC Genomics
<4>14
<5>175
<6>2013
<7>BACKGROUND: Mycoplasma hyopneumoniae, Mycoplasma flocculare and Mycoplasma
hyorhinis live in swine respiratory tracts. M. flocculare, a commensal bacterium,
is genetically closely related to M. hyopneumoniae, the causative agent of
enzootic porcine pneumonia. M. hyorhinis is also pathogenic, causing
polyserositis and arthritis. In this work, we present the genome sequences of M.
flocculare and M. hyopneumoniae strain 7422, and we compare these genomes with
the genomes of other M. hyoponeumoniae strain and to the a M. hyorhinis genome.
These analyses were performed to identify possible characteristics that may help
to explain the different behaviors of these species in swine respiratory tracts.
RESULTS: The overall genome organization of three species was analyzed, revealing
that the ORF clusters (OCs) differ considerably and that inversions and
rearrangements are common. Although M. flocculare and M. hyopneumoniae display a
high degree of similarity with respect to the gene content, only some genomic
regions display considerable synteny. Genes encoding proteins that may be
involved in host-cell adhesion in M. hyopneumoniae and M. flocculare display
differences in genomic structure and organization. Some genes encoding adhesins
of the P97 family are absent in M. flocculare and some contain sequence
differences or lack of domains that are considered to be important for adhesion
to host cells. The phylogenetic relationship of the three species was confirmed
by a phylogenomic approach. The set of genes involved in metabolism, especially
in the uptake of precursors for nucleic acids synthesis and nucleotide
metabolism, display some differences in copy number and the presence/absence in
the three species. CONCLUSIONS: The comparative analyses of three mycoplasma
species that inhabit the swine respiratory tract facilitated the identification
of some characteristics that may be related to their different behaviors. M.
hyopneumoniae and M. flocculare display many differences that may help to explain
why one species is pathogenic and the other is considered to be commensal.
However, it was not possible to identify specific virulence determinant factors
that could explain the differences in the pathogenicity of the analyzed species.
The M. hyorhinis genome contains differences in some components involved in
metabolism and evasion of the host's immune system that may contribute to its
growth aggressiveness. Several horizontal gene transfer events were identified.
The phylogenomic analysis places M. hyopneumoniae, M. flocculare and M. hyorhinis
in the hyopneumoniae clade.

<>

<1>Sirota-Madi, A., Olender, T., Helman, Y., Brainis, I., Finkelshtein, A., Roth, D., Hagai, E., Leshkowitz, D., Brodsky, L., Galatenko, V., Nikolaev, V., Gutnick, D.L., Lancet, D., Ben-Jacob, E.
<2>Genome Sequence of the Pattern-Forming Social Bacterium Paenibacillus dendritiformis C454 Chiral Morphotype.
<3>J. Bacteriol.
<4>194
<5>2127-2128
<6>2012
<7>Paenibacillus dendritiformis is a Gram-positive, soil-dwelling, spore-forming social
microorganism. An intriguing collective faculty of this strain is
manifested by its ability to switch between different morphotypes, such as the
branching (T) and the chiral (C) morphotypes. Here we report the 6.3-Mb draft
genome sequence of the P. dendritiformis C454 chiral morphotype.

<>

<1>Sirota-Madi, A., Olender, T., Helman, Y., Ingham, C., Brainis, I., Roth, D., Hagi, E., Brodsky, L., Leshkowitz, D., Galatenko, V., Nikolaev, V., Mugasimangalam, R.C., Bransburg-Zabary, S., Gutnick, D.L., Lancet, D., Ben-Jacob, E.
<2>Genome sequence of the pattern forming Paenibacillus vortex bacterium reveals potential for thriving in complex environments.
<3>BMC Genomics
<4>11
<5>710
<6>2010
<7>BACKGROUND: The pattern-forming bacterium Paenibacillus vortex is notable for its
advanced social behavior, which is reflected in development of colonies with
highly intricate architectures. Prior to this study, only two other Paenibacillus
species (Paenibacillus sp. JDR-2 and Paenibacillus larvae) have been sequenced.
However, no genomic data is available on the Paenibacillus species with
pattern-forming and complex social motility. Here we report the de novo genome
sequence of this Gram-positive, soil-dwelling, sporulating bacterium. RESULTS:
The complete P. vortex genome was sequenced by a hybrid approach using 454 Life
Sciences and Illumina, achieving a total of 289x coverage, with 99.8% sequence
identity between the two methods. The sequencing results were validated using a
custom designed Agilent microarray expression chip which represented the coding
and the non-coding regions. Analysis of the P. vortex genome revealed 6,437 open
reading frames (ORFs) and 73 non-coding RNA genes. Comparative genomic analysis
with 500 complete bacterial genomes revealed exceptionally high number of
two-component system (TCS) genes, transcription factors (TFs), transport and
defense related genes. Additionally, we have identified genes involved in the
production of antimicrobial compounds and extracellular degrading enzymes.
CONCLUSIONS: These findings suggest that P. vortex has advanced faculties to
perceive and react to a wide range of signaling molecules and environmental
conditions, which could be associated with its ability to reconfigure and
replicate complex colony architectures. Additionally, P. vortex is likely to
serve as a rich source of genes important for agricultural, medical and
industrial applications and it has the potential to advance the study of social
microbiology within Gram-positive bacteria.

<>

<1>Sisakova, E., Seidel, R., Szczelkun, M., Weiserova, M.
<2>Study of DNA translocation by EcoR124I type I RM enzyme.
<3>FEBS J.
<4>274
<5>305
<6>2007
<7>EcoR124I, Type I restriction-modification enzyme, is multifunctional, hetero-oligomeric enzyme
complex, able of ATP hydrolysis coupled to DNA translocation, the 1-D motion along the DNA
lattice. These activities are conferred by superfamily 2 (SF2) helicase motifs in the HsdR
subunit, however, the helicase motifs are not involved in DNA unwinding.
For studying DNA translocation activity of this enzyme, we prevented in vitro experiments to
be hampered by cleavage of DNA substrates by making substitutions of the conserved amino acid
residues in endonuclease motif D-X13-14-E-X-K of the HsdR subunit, responsible for restriction
activity of the enzyme(the substitutions made: D151A, E165A, E165D, E165H and K167A). Mutant
HsdR subunits were purified and endonucleases reconstituted from the wt methylase and
individual mutant HsdR subunits were analysed in vitro. DNA translocation activity and ATPase
activity of these mutants were analysed using a combination of bulk stopped-flow experiments
and single-molecule magnetic tweezers technique. The cleavage mutants of the EcoR124I enzyme
showed initiation and translocation rates similar to wild-type. However, ATPase activity of
the mutants was lower than of the wt enzyme. These results open new questions about HsdR
structure and its conformation in the subunit assembly. Only the K167A mutant had DNA
translocation and ATPase activity comparable to the wild type, which could be used as a useful
model for further investigation of the mechanism of DNA translocation of SF2 helicases.

<>

<1>Sisakova, E., Stanley, L.K., Weiserova, M., Szczelkun, M.D.
<2>A RecB-family nuclease motif in the Type I restriction endonuclease EcoR124I.
<3>Nucleic Acids Res.
<4>36
<5>3939-3949
<6>2008
<7>The Type I restriction-modification enzyme EcoR124I is an ATP-dependent endonuclease that uses
dsDNA translocation to locate and cleave distant
non-specific DNA sites. Bioinformatic analysis of the HsdR subunits of
EcoR124I and related Type I enzymes showed that in addition to the
principal PD-(E/D)xK Motifs, I, II and III, a QxxxY motif is also present
that is characteristic of RecB-family nucleases. The QxxxY motif resides
immediately C-terminal to Motif III within a region of predicted
alpha-helix. Using mutagenesis, we examined the role of the Q and Y
residues in DNA binding, translocation and cleavage. Roles for the QxxxY
motif in coordinating the catalytic residues or in stabilizing the
nuclease domain on the DNA are discussed.

<>

<1>Sisakova, E., van Aelst, K., Diffin, F.M., Szczelkun, M.D.
<2>The Type ISP Restriction-Modification enzymes LlaBIII and LlaGI use a translocation-collision mechanism to cleave non-specific DNA distant from their  recognition sites.
<3>Nucleic Acids Res.
<4>41
<5>1071-1080
<6>2013
<7>The Type ISP Restriction-Modification (RM) enzyme LlaBIII is encoded on plasmid pJW566 and can
protect Lactococcus lactis strains against bacteriophage
infections in milk fermentations. It is a single polypeptide RM enzyme comprising
Mrr endonuclease, DNA helicase, adenine methyltransferase and target-recognition
domains. LlaBIII shares >95% amino acid sequence homology across its first three
protein domains with the Type ISP enzyme LlaGI. Here, we determine the
recognition sequence of LlaBIII (5'-TnAGCC-3', where the adenine complementary to
the underlined base is methylated), and characterize its enzyme activities.
LlaBIII shares key enzymatic features with LlaGI; namely, adenosine
triphosphate-dependent DNA translocation ( approximately 309 bp/s at 25 degrees
C) and a requirement for DNA cleavage of two recognition sites in an inverted
head-to-head repeat. However, LlaBIII requires K(+) ions to prevent non-specific
DNA cleavage, conditions which affect the translocation and cleavage properties
of LlaGI. By identifying the locations of the non-specific dsDNA breaks
introduced by LlaGI or LlaBIII under different buffer conditions, we validate
that the Type ISP RM enzymes use a common translocation-collision mechanism to
trigger endonuclease activity. In their favoured in vitro buffer, both LlaGI and
LlaBIII produce a normal distribution of random cleavage loci centred midway
between the sites. In contrast, LlaGI in K(+) ions produces a far more
distributive cleavage profile.

<>

<1>Sisakova, E., Weiserova, M.
<2>Mutagenic analysis of the HsdR motor subunit of type IC restriction modification enzyme EcoR124I.
<3>FEBS J.
<4>272
<5>336
<6>2005
<7>Enzyme EcoR124I belongs to the IC family of restriction modification enzymes, intelligent
molecular motors. It is able to detect the methylation status of its DNA target sequence and
respond with alternative activities, methylation or translocation of DNA. While bound to its
target site, it translocates DNA towards itself simultaneously in both directions (500bp/sec).
It uses the free energy associated with ATP hydrolysis to translocate DNA so that DNA cleavage
occurs remote from the asymmetric recognition site. The enzyme EcoR124I is a multifunctional,
multi-subunit enzyme, composed of three different subunits, which are encoded by the genes
hsdR, hsdM and hsdS. Products of all three genes are required for DNA cleavage, producing the
endonuclease. HsdR subunit is a multifunctional motor protein, which has been shown to possess
ATPase, helicase and restriction activity. To provide a fully functional molecular motor,
which can never cleave DNA, the amino acid motif X, the active site of the endonuclease domain
of the HsdR subunit, was subjected to site-directed mutagenesis. The complementation analysis
proved that the substitutions D151A, E165A, E165D, E165H, K167A in the HsdR subunit fully
removed the restriction activity in vivo of the EcoR124I enzyme. The mutant subunits were
separately overproduced, purified and mixed with purified methylase to reconstitute the
EcoR124I endonuclease in vitro. As a substrate for DNA cleavage in vitro we used the plasmid
pCFD30 containing a single site for EcoR124I. The test of restriction activity showed that
reconstituted endonucleases were not able to cleave covalently closed plasmid DNA to linear
DNA in contrast to the wild-type enzyme.

<>

<1>Sisakova, E., Weiserova, M., Dekker, C., Seidel, R., Szczelkun, M.D.
<2>The Interrelationship of Helicase and Nuclease Domains during DNA Translocation by the Molecular Motor EcoR124I.
<3>J. Mol. Biol.
<4>384
<5>1273-1286
<6>2008
<7>The type I restriction-modification enzyme EcoR124I comprises three subunits with the
stoichiometry HsdR(2)/HsdM(2)/HsdS(1). The HsdR subunits
are archetypical examples of the fusion between nuclease and helicase
domains into a single polypeptide, a linkage that is found in a great many
other DNA processing enzymes. To explore the interrelationship between
these physically linked domains, we examined the DNA translocation
properties of EcoR124I complexes in which the HsdR subunits had been
mutated in the RecB-like nuclease motif II or III. We found that nuclease
mutations can have multiple effects on DNA translocation despite being
distinct from the helicase domain. In addition to reductions in DNA
cleavage activity, we also observed decreased translocation and ATPase
rates, different enzyme populations with different characteristic
translocation rates, a tendency to stall during initiation and altered
HsdR turnover dynamics. The significance of these observations to our
understanding of domain interactions in molecular machines is discussed.

<>

<1>Sistla, S., Krishnamurthy, V., Rao, D.N.
<2>Single-stranded DNA binding and methylation by EcoP1I DNA methyltransferase.
<3>Biochem. Biophys. Res. Commun.
<4>314
<5>159-165
<6>2004
<7>EcoP1I methyltransferase (M.EcoP1I) belongs to the type III restriction-modification system
encoded by prophage PI that infects
Escherichia coli. Binding of M.EcoP1I to double-stranded DNA and
single-stranded DNA has been characterized. Binding to both single- and
double-stranded DNA could be competed out by unlabeled single-stranded
DNA. Metal ions did not influence DNA binding. Interestingly, M.EcoP1I
was able to methylate single-stranded DNA. Kinetic parameters were
determined for single and double-stranded DNA methylation. This feature
of the enzyme probably functions in protecting the phage genome from
restriction by type III restriction enzymes and thus could be
considered as an anti-restriction system. This study describing in
vitro methylation of single-stranded DNA by the type III
methyltransferase EcoP1I allows understanding of the mechanism of
action of these enzymes and also their role in the biology of
single-stranded phages.

<>

<1>Sistla, S., Rao, D.N.
<2>S-adenosyl-L-methionine-dependent restriction enzyme.
<3>Crit. Rev. Biochem. Mol. Biol.
<4>39
<5>1-19
<6>2004
<7>Restriction-modification (R-M) enzymes are classified into type I, II, III, and IV, based on
their recognition sequence, subunit composition,
cleavage position, and cofactor requirements. While the role of
S-Adenosyl-L-methionine (AdoMet) as the methyl group donor in the
methylation reaction is undisputed, its requirement in DNA cleavage
reaction has been subject to intense study. AdoMet is a prerequisite
for the DNA cleavage by most type I enzymes known so far, with the
exception of R.EcoR124I. A number of new type 11 restriction enzymes
belonging to the type IIB and IIG family were found to show AdoMet
dependence for their cleavage reaction. The type III enzymes have been
found to require AdoMet for their restriction function. AdoMet
functions as an allosteric effector of the DNA cleavage reaction and
has been shown to bring about conformational changes in the protein
upon binding.

<>

<1>Sitaraman, R.
<2>Helicobacter pylori DNA methyltransferases and the epigenetic field effect in cancerization.
<3>Front. Microbiol.
<4>5
<5>4
<6>2014
<7>Helicobacter pylori, a Gram-negative, microaerophilic bacterium, has co-existed with human
beings as a prominent member of their gastric microbiota for approximately 105 years.  It
infects approximately half the world's population, and most infected individuals are
asymptomatic, but histologically exhibit superficial gastritis.  Only a minority of infected
individuals develop gastric or duodenal ulcers that necessitate treatment.  Prolonged
inflammation caused by chronic (often lifelong) infection predisposes a small fraction of
infected individuals to develop gastric adenocarcinoma or lymphoma of the mucosa-associated
lymphoid tissue (MALT lymphoma).  Unfortunately, the prognosis for cases of gastric cancer is
very poor, with 5-year survival rates being lower than 15%.

<>

<1>Sitaraman, R., Dybvig, K.
<2>The hsd loci of Mycoplasma pulmonis: organization, rearrangements and expression of genes.
<3>Mol. Microbiol.
<4>26
<5>109-120
<6>1997
<7>Two paralogous, site-specific invertible loci, designated hsd1 and hsd2, have been identified
in the Mycoplasma pulmonis genome.  They encode putative type I restriction and modification
systems with maximum sequence homology to the type IC family, which includes EcoR124II and
EcoDXXI.  Each locus encodes an endonuclease subunit, a methylase subunit and two DNA
specificity subunits.  The gene organization at each locus is such that hsdR and hsdM are
flanked by two hsdS genes.  Within each locus, one of the hsdS genes, hsdR and hsdM, is
encoded in tandem by the same DNA strand, while the second hsdS gene is encoded by the
complementary strand but without overlap with the other three hsd genes.  The hsdR and hsdM
sequences of one locus are almost identical to their counterparts in the other.  The four hsdS
genes (two per locus) are highly homologous at their 5' ends and also share sequence
similarities in the 3' ends of their corresponding coding regions.  Owing to the disposition
of and sequence similarities among the hsdS genes, they form inverted repeats at each locus.
Analysis by polymerase chain reaction has shown that both loci behave as site-specific DNA
invertible elements with multiple inversion sites, termed 'vipareetus', occurring within the
hsdS genes.  The inversions lead to a reassortment of hsdS sequences, generating an array of
recombinant genes that probably encode S subunits possessing alternative DNA-binding
specificities.  Sequence information obtained from the analysis of hsd2 transcripts by 5'
RACE indicates that inversion induces the transcription of alternative hsdS genes by the
relocation of coding sequences downstream of a promoter and ribosome-binding sites situated at
one end of each locus.

<>

<1>Sitaraman, R., Dybvig, K.
<2>Functional analysis of a unique site-specific recombinase from Mycoplasma pulmonis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>388
<6>2001
<7>Mycoplasma pulmonis has been shown to undergo high-frequency phase variation at different loci
within its genome - the two paralogous hsd
(host specificity determinant) loci that encode type I restriction and
modification (R-M) systems, and the vsa (variable surface antigen)
locus that encodes surface lipoproteins. Analyses of the hsd and vsa
gene loci have shown that they are organized as site-specific
invertible elements. Recently, the genome of M. pulmonis strain UAB
CTIP has been completely sequenced. Based on sequence homology, an open
reading frame near the vsa locus was predicted to encode a
site-specific recombinase. This is the only known site-specific
recombinase identified in Mollicutes that is not part of a viral and/or
transposable element. To test the functionality and sequence
specificity of the predicted recombinase, two assay systems have been
designed in an E. coli background using information about the regions
of the hsd and vsa genes that are actually involved in site-specific
recombination. Two copies of the relevant hsd (or vsa) regions were
cloned in opposite orientation into plasmid vectors. E. coli harboring
these "reporter" plasmids were then transformed with a compatible
plasmid containing the putative recombinase gene. The clones so
obtained were screened by PCR for inversion at the hsd- or vsa-derived
inverted repeats. The resulting amplicons corresponding to DNA
inversions were also sequenced, further verifying the occurrence of the
predicted inversions. This work proves that the putative recombinase is
functional and catalyzes site-specific inversions of the hsd- and
vsa-derived sequences that are known to undergo inversions in M.
pulmonis. The use of a single enzyme to catalyze DNA inversions at
different genetic loci is consistent with the observed genetic economy
of mycoplasmas. In view of the sequence differences between the sites
of inversion at the vsa and hsd loci, experiments to determine the
minimal sequence requirements for site-specific recombination are in
progress.

<>

<1>Sitaraman, R., Dybvig, K.
<2>Restriction-modification systems and chromosomal rearrangements in mycoplasmas.
<3>Mol. Biol. Pathog. Mycoplasmas
<4>0
<5>371-390
<6>2002
<7>The restriction-modification (R-M) systems are so named because of their ability to degrade
foreign DNA (restrict viral infection) and to methylate (modify) unmethylated and
hemimethylated DNA.  They were discovered as genetic elements that determined the
susceptibility of E. coli and S. typhimurium to phage infection in a strain-dependent manner,
with bacteriophage plaquing efficiency being reduced upon infection of a heterologous host
strain.  At the time of writing, 3154 R-M enzymes, many with overlapping DNA recognition
specificities, have been catalogued in the restriction enzyme database (REBASE).  R-M systems
are ubiquitous and arose early in prokaryotic evolution.

<>

<1>Sitaraman, R., Leppla, S.H.
<2>Methylation-dependent DNA restriction in Bacillus anthracis.
<3>Gene
<4>494
<5>44-50
<6>2012
<7>Bacillus anthracis, the causative agent of anthrax, is poorly transformed with DNA that is
methylated on adenine or cytosine. Here we characterize
three genetic loci encoding type IV methylation-dependent restriction
enzymes that target DNA containing C5-methylcytosine (m5C). Strains in
which these genes were inactivated, either singly or collectively, showed
increased transformation by methylated DNA. Additionally, a triple mutant
with an ~30-kb genomic deletion could be transformed by DNA obtained from
Dam(+)Dcm(+)E. coli, although at a low frequency of ~10(-3)
transformants/10(6)cfu. This strain of B. anthracis can potentially serve
as a preferred host for shuttle vectors that express recombinant proteins,
including proteins to be used in vaccines. The gene(s) responsible for the
restriction of m6A-containing DNA in B. anthracis remain unidentified, and
we suggest that poor transformation by such DNA could in part be a
consequence of the inefficient replication of hemimethylated DNA in B.
anthracis.

<>

<1>Sitbon, E., Pietrokovski, S.
<2>New types of conserved sequence domains in DNA-binding regions of homing endonucleases.
<3>Trends Biochem. Sci.
<4>28
<5>473-477
<6>2003
<7>We have identified four new types of short conserved sequence domains in homing endonucleases
and related proteins. These domains are modular,
appearing in various combinations. One domain includes a motif known by
structure as a novel sequence-specific DNA-binding helix. Sequence
similarity shows two other domains to be new types of helix-turn-helix
DNA-binding domains. We term the new domains nuclease-associated modular
DNA-binding domains (NUMODs).

<>

<1>Siwek, W., Czapinska, H., Bochtler, M., Bujnicki, J.M., Skowronek, K.
<2>Crystal structure and mechanism of action of the N6-methyladenine-dependent type  IIM restriction endonuclease R.DpnI.
<3>Nucleic Acids Res.
<4>40
<5>7563-7572
<6>2012
<7>DNA methylation-dependent restriction enzymes have many applications in genetic engineering
and in the analysis of the epigenetic state of eukaryotic genomes.
Nevertheless, high-resolution structures have not yet been reported, and
therefore mechanisms of DNA methylation-dependent cleavage are not understood.
Here, we present a biochemical analysis and high-resolution DNA co-crystal
structure of the N(6)-methyladenine (m6A)-dependent restriction enzyme R.DpnI.
Our data show that R.DpnI consists of an N-terminal catalytic PD-(D/E)XK domain
and a C-terminal winged helix (wH) domain. Surprisingly, both domains bind DNA in
a sequence- and methylation-sensitive manner. The crystal contains R.DpnI with
fully methylated target DNA bound to the wH domain, but distant from the
catalytic domain. Independent readout of DNA sequence and methylation by the two
domains might contribute to R.DpnI specificity or could help the monomeric enzyme
to cut the second strand after introducing a nick.

<>

<1>Sizova, I.A., Glibin, E.N., Ginzburg, O.F., Tereshin, I.M.
<2>Inhibition of the restriction of DNA by endonucleases in the presence of Actinomycin D, Distamycin A, and their analogs.
<3>Bioorg. Khim.
<4>8
<5>470-477
<6>1982
<7>The cleavage of phage lambda cI857s7 by the restriction endonucleases EcoRI and
SmaI in the presence of actinomycin D, distamycin A, and the distamycin analog
distamine, and also of polyfunctional ligands -- distaccins -- has been
studied.  Distamine suppresses the cleavage of phage lambda DNA by endonuclease
EcoRI in the same way as distamycin A.  Actinomycin D selectively blocks the
action of endonucleases SmaI and EcoRI; in the latter case the cleavage of only
one recognition site is suppressed.  In the inhibition of the action of
endonuclease EcoRI simultaneously by two ligands, actinomycin D and distamycin
A or actinomycin D and distamine, "additivity" of their action on the
recognition site present at the point of contact of the EcoRI fragments E and F
was detected.  When DNA was cleaved by endonuclease SmaI in the presence of
these ligands no "additivity" was revealed.  In this case, a rise in the degree
of cleavage of the DNA at the points of contact of the SmaI fragments A and B,
and D and C, as compared with the action of actinomycin D alone, was observed.
Distaccins are nonspecific inhibitors of the cleavage of DNA under the
influence of the enzymes EcoRI and SmaI; distaccin-O protects the sections of
the recognition of endonuclease SmaI and does not block the action of
endonuclease EcoRI.

<>

<1>Sjodin, A., Ohrman, C., Backman, S., Larkeryd, A., Granberg, M., Lundmark, E., Karlsson, E., Nilsson, E., Vallesi, A., Tellgren-Roth, C., Stenberg, P., Thelaus, J.
<2>Complete Genome Sequence of Francisella endociliophora Strain FSC1006, Isolated from a Laboratory Culture of the Marine Ciliate Euplotes raikovi.
<3>Genome Announcements
<4>2
<5>e01227-14
<6>2014
<7>A strain of Francisella endociliophora was isolated from a laboratory culture of  the marine
ciliate Euplotes raikovi. Here, we report the complete genome sequence
of the bacterial strain FSC1006 (Francisella Strain Collection, Swedish Defence
Research Agency, Umea, Sweden).

<>

<1>Sjolund-Karlsson, M., Reimer, A., Folster, J.P., Walker, M., Dahourou, G., Batra, D.G., Martin, I., Joyce, K., Parsons, M.B., Boncy, J., Whichard, J.M., Gilmour, M.W.
<2>Drug-Resistance Mechanisms in Vibrio cholerae O1 Outbreak Strain, Haiti, 2010.
<3>Emerg. Infect. Dis.
<4>17
<5>2151-2154
<6>2011
<7>To increase understanding of drug-resistant Vibrio cholerae, we studied selected
molecular mechanisms of antimicrobial drug resistance in the 2010 Haiti V.
cholerae outbreak strain. Most resistance resulted from acquired genes located on
an integrating conjugative element showing high homology to an integrating
conjugative element identified in a V. cholerae isolate from India.

<>

<1>Sjostrom, J.E., Lofdahl, S., Philipson, L.
<2>Biological characteristics of a Type I restriction-modification system in Staphylococcus aureus.
<3>J. Bacteriol.
<4>133
<5>1144-1149
<6>1978
<7>Two restriction-modification systems, S1 and S2, are present in Staphylococcus aureus RN450
(S. Iordanescu and M. Surdeanu, J. Gen. Microbiol., 96:277-281,1976).  System S2 affects phage
multiplication after both infection and transfection.  Unmodified plasmid and chromosomal DNAs
are also not expressed following transduction and transformation into a restrictive host.
Restricted phages are, however, capable of conferring phage-mediated competence, although the
state of competence does not affect the restriction-modification system. The restricting
activity of system S2 is inactivated by heat treatment of the cells.  An enzymatic activity
that restricts unmodified phage DNA in the presence of ATP, Mg2+, and S-adenosylmethionine was
recovered from cell-free extracts of a strain RN450 derivative.

<>

<1>Skarin, H., Hafstrom, T., Westerberg, J., Segerman, B.
<2>Clostridium botulinum group III: a group with dual identity shaped by plasmids, phages and mobile elements.
<3>BMC Genomics
<4>12
<5>185
<6>2011
<7>ABSTRACT: BACKGROUND: Clostridium botulinum strains can be divided into
four physiological groups that are sufficiently diverged to be considered
as separate species. We here presentHere we present the first complete
genome of a C. botulinum strain from physiological group III, causing
animal botulism. We also compare the sequence to three new draft genomes
from the same physiological group. RESULTS: The 2.77 Mb chromosome was
highly conserved between the isolates and also closely related to that of
C. novyi. However, the sequence was very different from the human C.
botulinum group genomes. Replication-directed translocations were rare and
conservation of synteny was high. The largest difference between C.
botulinum group III isolates occurred within their surprisingly large
plasmidomes and in the pattern of mobile elements insertions. Five
plasmids, constituting 13.5% of the total genetic material, were present
in the completed genome. Interestingly, the set of plasmids differed
compared to other isolates. The largest plasmid, the botulinum- neurotoxin
carrying prophage, was conserved to at a level similar to that of the
chromosome while the medium-sized plasmids seemed to be undergoing faster
genetic drift. These plasmids also contained more mobile elements than
other replicons. Several toxins and resistance genes were identified, many
of which were located on the plasmids. CONCLUSIONS: The completion of the
genome of C. botulinum group III has revealed it to be a genome withof
dual identity. It belongs to the pathogenic species C. botulinum, but as a
genotypic species it should also include C. novyi and C. haemolyticum. The
genotypic species share a conserved chromosomal core that can be
transformed into various pathogenic variants by modulation of the highly
plastic plasmidome.

<>

<1>Skavronskaya, A.G., Aleshkin, G.I., Demkin, V.V.
<2>Specificity of modification-restriction of bacteriophages P1 and lambda in strains of Escherichia coli K-12 controlled by the R124 plasmid.
<3>Genetika
<4>15
<5>1719-1723
<6>1979
<7>The specificities of restriction of bacteriophages P1 and lambda controlled by
R plasmids in Escherichia coli have been investigated.  The isogenic strains
harbouring the plasmids pAS26 coding for restriction endonuclease R.EcoRI, R245
coding for restriction endonuclease R.EcoRII and R124 have been investigated in
the present work.  Modification-restriction controlled by R124 has been found
to differ in specificity from those controlled by R245 and pAS26.  Frequencies
of restriction of bacteriophages P1vir and lambdavir specified by R124 plasmid
differ from the frequencies in the strains harbouring pAS26 and R245 plasmids
as well.  The difference is due to the specificity of restriction-modification
controlled by R124.  The data obtained are consistent with the determination of
R124 specified restriction-modification activity as a novel one designated
R.EcoRIII.

<>

<1>Skeiky, Y.A.W., Persing, D.H., Mitcham, J.L., Wang, S.S., Bhatia, A., L'Maisonneuve, J.-F., Zhang, Y., Jen, S., Carter, D.
<2>Compositions and methods for the therapy and diagnosis of acne vulgaris.
<3>International Patent Office
<4>WO 0181581 A
<5>
<6>2001
<7>Compositions and methods for the therapy and diagnosis of acne vulgaris and other related
conditions are disclosed.  Compositions may comprise one or more Propionibacterium acnes
proteins, immunogenic portions thereof, or polynucleotides that encode such portions.
Alternatively, a therapeutic composition may comprise an antibody that binds a
Propionibacterium acnes protein, antigen presenting cell that expresses a Propionibacterium
acnes protein, or a T cell that is specific for cells expressing such a protein.  Such
compositions may be used, for example, for the prevention and/or treatment of acne.

<>

<1>Skennerton, C.T., Angly, F.E., Breitbart, M., Bragg, L., He, S., McMahon, K.D., Hugenholtz, P., Tyson, G.W.
<2>Phage Encoded H-NS: A Potential Achilles Heel in the Bacterial Defence System.
<3>PLoS ONE
<4>6
<5>e20095
<6>2011
<7>The relationship between phage and their microbial hosts is difficult to elucidate in complex
natural ecosystems. Engineered systems performing
enhanced biological phosphorus removal (EBPR), offer stable, lower
complexity communities for studying phage-host interactions. Here,
metagenomic data from an EBPR reactor dominated by Candidatus
Accumulibacter phosphatis (CAP), led to the recovery of three complete and
six partial phage genomes. Heat-stable nucleoid structuring (H-NS)
protein, a global transcriptional repressor in bacteria, was identified in
one of the complete phage genomes (EPV1), and was most similar to a
homolog in CAP. We infer that EPV1 is a CAP-specific phage and has the
potential to repress up to 6% of host genes based on the presence of
putative H-NS binding sites in the CAP genome. These genes include CRISPR
associated proteins and a Type III restriction-modification system, which
are key host defense mechanisms against phage infection. Further, EPV1 was
the only member of the phage community found in an EBPR microbial
metagenome collected seven months prior. We propose that EPV1 laterally
acquired H-NS from CAP providing it with a means to reduce bacterial
defenses, a selective advantage over other phage in the EBPR system. Phage
encoded H-NS could constitute a previously unrecognized weapon in the
phage-host arms race.

<>

<1>Skirgaila, R., Grazulis, S., Bozic, D., Huber, R., Siksnys, V.
<2>Structure-based redesign of the catalytic/metal binding site of Cfr10I restriction endonuclease reveals importance of spatial rather than sequence conservation of active centre residues.
<3>J. Mol. Biol.
<4>279
<5>473-481
<6>1998
<7>According to the crystal structure of Cfr10I restriction endonuclease the acidic residues
D134, E71 and E204 are clustered together and presumably chelate metal ion(s) at the active
site.  Indeed, investigation of the DNA cleavage properties of substitutional mutants of
Cfr10I D134A, E71Q, E71A and E204Q reveals that D134, E71 and E204 residues are essential for
cleavage activity, supporting their active site function.  Structural comparison indicates
that the D134 residue of Cfr10I spatially overlaps with aspartate residues D91 and D74, from
the invariant active site motifs 90PD(X19)EAK and 73 PD(X15)DIK of EcoRI and EcoRV,
respectively.  However, structural studies in conjunction with mutational analyses suggest
that the sequence motif 133PD(X55)K(X13)E corresponds to the active site of Cfr10I, but
differs from the canonical active site motifs of EcoRI and EcoRV.  According to the crystal
structure of Cfr10I the serine S188 residue from the 188SVK sequence motif is a spatial
equivalent of the acidic residue from the (E/D)XK-part of the active site motif, which is
conserved between EcoRI and EcoRV.  Site-directed mutagenesis experiments of Cfr10I, however,
revealed that S188 was not so important for catalysis while the E204 residue located 2.8A away
indeed was essential for cleavage, suggesting that the glutamate E204 rather than the S188
residue contributes to the metal binding site in Cfr10I.  In addition, model-building studies
suggest that mutual interchange of the E204 and S188 residues should lead only to minor
positional differences of the carboxylate residues of glutamate side-chains.  The double
mutant S188E/E204S was therefore prepared by site-directed mutagenesis where the active site
motif 133 PD(X55)K(X13)E of Cfr10I was changed to a canonical motif 133PD(X53)EVK, which is
similar to that of EcoRI and EcoRV.  Interestingly, the double mutant S188E/E204S of Cfr10I
with redesigned active site structure, exhibited 10% of Wt cleavage activity in a lambda DNA
cleavage assay.  Thus, structure guided redesign of the catalytic/metal binding site of
Cfr10I, provides novel experimental evidence to suggest that spatial rather than sequence
conservation plays the dominant role in the formation of restriction enzyme active sites.

<>

<1>Skirgaila, R., Siksnys, V.
<2>Ca2+-ions stimulate DNA binding specificity of Cfr10I restriction enzyme.
<3>Biol. Chem.
<4>379
<5>595-598
<6>1998
<7>The Cfr10I restriction enzyme recognizes the degenerate hexanucleotide sequence
5'-Pu/CCGGPy-3' and cleaves it as indicated.  DNA binding studies of Cfr10I endonuclease
were performed using gel mobility shift assay.  Analysis of Cfr10I binding to DAN revealed
that in the absence of metal ions Cfr10I binds to DNA containing or lacking the recognition
sequence with similar low affinity.  Addition of Ca2+ to the binding specificity of Cfr10I.

<>

<1>Sklar, R., Altman, D., Sager, R.
<2>An endonuclease from Chlamydomonas reinhardtii that cleaves the sequence TATA.
<3>J. Biol. Chem.
<4>261
<5>6806-6810
<6>1986
<7>An endodeoxyribonuclease, designated CreI, was purified 16,000-fold from
zygotes of the eukaryote Chlamydomonas reinhardtii.  CreI preferentially
attacks the sequence TATA producing double stsrand breaks with
3'-phosphomonoester and 5'-hydroxyl termini.  The endonuclease has an Mr =
27,000 and requires Ca2+ at pH 7.5 for optimal activity.

<>

<1>Skoglund, A., Bjorkholm, B., Nilsson, C., Andersson, A.F., Jernberg, C., Schirwitz, K., Enroth, C., Krabbe, M., Engstrand, L.
<2>Functional analysis of the M.HpyAIV DNA methyltransferase of Helicobacter pylori.
<3>J. Bacteriol.
<4>189
<5>8914-8921
<6>2007
<7>A large number of genes encoding restriction-modification (R-M) systems are found in the
genome of the human pathogen Helicobacter pylori. R-M
genes comprise approximately 10% of the strain-specific genes, but the
relevance of having such an abundance of these genes is not clear. The
type II methyltransferase (MTase) M.HpyAIV, which recognizes GANTC sites,
was present in 60% of the H. pylori strains analyzed, whereof 69% were
resistant to restriction enzyme digestion, which indicated the presence of
an active MTase. H. pylori strains with an inactive M.HpyAIV phenotype
contained deletions in regions of homopolymers within the gene, which
resulted in premature translational stops, suggesting that M.HpyAIV may be
subjected to phase variation by a slipped-strand mechanism. An M.HpyAIV
gene mutant was constructed by insertional mutagenesis, and this mutant
showed the same viability and ability to induce interleukin-8 in
epithelial cells as the wild type in vitro but had, as expected, lost the
ability to protect its self-DNA from digestion by a cognate restriction
enzyme. The M.HpyAIV from H. pylori strain 26695 was overexpressed in
Escherichia coli, and the protein was purified and was able to bind to DNA
and protect GANTC sites from digestion in vitro. A bioinformatic analysis
of the number of GANTC sites located in predicted regulatory regions of H.
pylori strains 26695 and J99 resulted in a number of candidate genes.
katA, a selected candidate gene, was further analyzed by quantitative
real-time reverse transcription-PCR and shown to be significantly
down-regulated in the M.HpyAIV gene mutant compared to the wild-type
strain. This demonstrates the influence of M.HpyAIV methylation in gene
expression.

<>

<1>Skoglund, A., Bjorkholm, B., Nilsson, C., Krabbe, M., Engstrand, L.
<2>Are hsdS-subunits in restriction-modification systems involved in Helicobacter pylori virulence?
<3>Helicobacter
<4>8
<5>492
<6>2003
<7>Few regulatory genes are present in the H. pylori genome, but there is an abundance of genes
homologous to restriction-modification systems, that may play a role in gene regulation.  Each
strain has its own unique set of R-M genes.  In a recent study, the presence of hsdS genes
(specificity subunits of type I R-M systems) in clinical isolates was associated with
induction of a more robust host response in gnotobiotic transgenic mice.  We aim to study hsdS
genes and their involvement in virulence.  By PCR-amplification, two hsdS genes were detected
in 100% of the isolates, however, the amplified products had variable sizes.  We have
sequenced one hsdS locus in nine clinical isolates.  The sequenced locus reveal large
variations between the strains, but there were also conserved regions.  One of the conserved
domain is known as the target recognition domain.  We have constructed insertion mutants in
type I hsdS and hsdM (modification subunit of type I systems) genes: jhp0726, HP0790, HP0462
and HP0463.  Preliminary results indicate that one of the mutant strains (HP0462::km) has
reduced growth compared with the wild-type strain.  Strains deficient in hsdS and hsdM genes
do not show altered ability to induce IL-8 production in AGS-cells.  The effects of
inactivation of the hsdS genes on global gene expression are now being investigated using a
whole-genome microarray.  To investigate the involvement of hsdS genes in virulence, the
host-response of mutant compared with wild-type strains will be studied in germfree mice.

<>

<1>Skoglund, A., Bjorkholm, B., Nilsson, C., Krabbe, M., Engstrand, L.
<2>Are hsdS-subunits in restriction-modification systems involved in Helicobacter pylori virulence?
<3>Int. J. Med. Microbiol.
<4>293
<5>125
<6>2003
<7>Few regulatory genes are present in the H. pylori genome, but there is an abundance of genes
homologous to restriction-modification systems, that may play a role in gene regulation.  Each
strain has its own unique set of R-M genes.  In a recent study, the presence of hsdS genes
(specifically subunits of type I R-M systems) in clinical isolates was associated with
induction of a more robust host response in gnotobiotic transgenic mice.  We aim to study hsdS
genes and their involvement in virulence.  By PCR-amplification, two hsdS genes were detected
in 100% of the isolates, however the amplified products had variable sizes.  We have sequenced
one hsdS locus in 9 clinical isolates.  The sequenced locus reveals large variations between
the strains, but there were also conserved regions.  One of the conserved domains is known as
the target recognition domain.  We have constructed insertion mutants in type I hsdS and hsdM
(modification subunit of type I systems) genes: jhp0726, HP0790, HP0462 and HP0463.
Preliminary results indicate that one of the mutant strains (HP0462::km) has reduced growth
compared to the wild-type strain.  Strains deficient in hsdS and hsdM genes do not show
altered ability to induce IL-8 production in AGS-cells.  The effects of inactivation of the
hsdS genes on global gene expression are now being investigated using a whole-genome
microarray.  To investigate the involvement of hsdS genes in virulence, the host-response of
mutant compared to wild-type strains will be studied in germfree mice.

<>

<1>Skoglund, A., Engstrand, L., Krabbe, M.E.
<2>DNA methyltransferases in Helicobacter pylori.
<3>Int. J. Med. Microbiol.
<4>291S
<5>94-95
<6>2001
<7>DNA methyltransferases are enzymes that modify specific sequences in DNA.  In bacteria, this
modification is important as protection of "self" DNA in restriction-modification systems and
in cellular processes such as DNA repair, control of cell division and regulation of virulence
genes.  We are interested in the role of multiple methyltransferases in H. pylori.  To
determine the distribution of methyltransferase genes in DNA prepared from clinical isolates
of H. pylori, oligonucleotide pairs directed towards conserved regions of 18 MT genes were
used in PCR.  The PCR reaction for four genes, HPO260, HPO263, HPO478 and HPO910, were present
in all the DNA isolates tested, whereas other MT genes were only present in a subset of the
strains.  We are now proceeding to inactivate these and four additional MT genes in H. pylori
strain 26695 by insertion of a Kanamycin cassette (aphA) from Campylobacter coli into the MT
reading frame to determine the phenotypical effect of loss of the MT activity.  The first of
our inactivation constructs, HP0483 (not conserved in clinical isolates) was interrupted
without any obvious effect on the growth of the mutant as compared with wild-type bacteria.
This indicates that the restriction enzyme encoded downstream of the MT gene was not expressed
under the conditions used, since that likely would have resulted in degradation of the
chromosomal DNA in the mutant.  We will continue to analyze this and other MT mutants for
their phenotype.  We are also interested in gene regulation and we will investigate whether
these MT genes are regulated by environmental stimuli.

<>

<1>Skoglund, C.M., Smith, H.O., Chandrasegaran, S.
<2>Construction of an efficient overproducer clone of HinfI restriction endonuclease using the polymerase chain reaction.
<3>Gene
<4>88
<5>1-5
<6>1990
<7>We describe the use of the polymerase chain reaction (PCR) technique to alter
transcriptional and translational signals surrounding a gene so as to achieve
overexpression in Escherichia coli.  By changing the ribosome-binding site
sequence preceding the hinfIR gene to match the consensus E. coli signal and by
adding a transcription terminator sequence immediately following the gene, the
yield of HinfI was increased about tenfold over that obtained from the natural
Haemophilus influenzae signals.  The addition of the positive retroregulator
stem-loop sequence derived from the crystal protein-encoding gene of Bacillus
thuringiensis downstream from the hinfIR gene further increased yields by
twofold to a level of 13% of the total cellular protein.

<>

<1>Skogman, G.S., Bjork, G.R.
<2>Effects of the phage P1 restriction system on Coliphage PhiW: Degradation and complex formation of phage PhiW DNA.
<3>J. Gen. Virol.
<4>31
<5>9-20
<6>1976
<7>Growth of phages PhiW and T7 was restricted in Escherichia coli lysogenic for phage P1.  Only
a fraction of the infected cells gave burst of phages.  Cells permitting phage growth gave
normal burst size.  Host strains carrying P1 mutants with defective endonuclease gave no
restriction of phages T7 and Phi3, the latter a host-range mutant of PhiW.  Degradation but
not modification of parental phage DNA could be demonstrated.  Although no DNA, RNA or protein
was synthesized in PhiW infected P1 lysogenic cells, the parental phage DNA was found in
increasingly larger complexes during the course of infection.  At early times after infection,
parental phage DNA was found to sediment about twice as fast as mature phage DNA.  At later
times during the infection the parental phage DNA was recovered as a very rapidly sedimenting
material.  Such material was also found in alkaline sucrose gradient centrifugation after
treatment of the cell extract with sodium dodecyl sulphate, pronase digestion and phenol
extractions.

<>

<1>Skov, K.A., Adomat, H., Konway, D.C., Farrell, N.P.
<2>Assessment of DNA binding of platinum-radio-sensitizer complexes by inhibition of restriction enzymes.
<3>Chem. Biol. Interact.
<4>62
<5>117-129
<6>1987
<7>A simple and rapid method has been used to compare the binding of platinum
complexes to DNA, in a relatively qualitative manner.  A compound bound at or
near the restriction site inhibits enzymatic cleavage of DNA; inhibition of
BamHI and EcoRI activity by complexes was assessed in this study using
linearized pSV2-gpt plasmid.  Our particular interest was in DNA binding by
complexes of platinum (Pt) with known organic radiosensitizers (RS), to
determine whether the Pt was able to target the RS to the DNA. Although the
Pt-RS complexes investiaged themselves have moderate radiosensitizing ability
(like the inorganic complexes, cis- or trans-diamminedichloroplatinum(II), c-
or t-DDP) none of the Pt-RS inhibit to the same extent as c- or t-DDP.
However, there appears to be some correlation between enhanced
radiosensitization by Pt-RS over Pt(RS), with the degree of Pt binding (as
assessed by our assay).  Our results using isolated DNA suggest that not all
complexes bind well (e.g. Pt with two RS ligands), but that in certain cases
(e.g. Pt with only one RS), it is possible to target the drug to the DNA.  An
ammine or amine ligand may be required in order to target a radiosensitizer to
DNA using platinum.

<>

<1>Skowron, P., Kaczorowski, T., Tucholski, J., Podhajska, A.J.
<2>Atypical DNA-binding properties of class-IIS restriction endonucleases:evidence for recognition of the cognate sequence by a FokI monomer.
<3>Gene
<4>125
<5>1-10
<6>1993
<7>The DNA-binding properties of the FokI restriction endonuclease were studied using the
gel-mobility-shift assay. Specific recognition of the cognate sequence and cleavage of DNA are
distinguishable functions and can be separated. FokI binds to its recognition site
predominantly as a monomer. At high concentrations, FokI exhibits a cooperative recognition
sequence-dependent aggregation. In 20 mM KCl/10 mM Tris-HCl buffer, the binding constant of
FokI to its cognate site is 6.0-7.9 x 10/8 per mol and is lower than the values for most
gene-regulatory proteins. FokI binding is 600-1500 times weaker to non-cognate double-stranded
DNA than to the GGATG site, and 30,000 times weaker to single-stranded DNA or tRNA. The method
of Bading [Nucleic Acids Res. 16 (1988) 5241-5248], used for determining the stoichometry of
protein bound to DNA by gel-mobility-shift assay, is extended.

<>

<1>Skowron, P.M., Majewski, J., Zylicz-Stachula, A., Rutkowska, S.M., Jaworowska, I., Harasimowicz-Slowinska, R.I.
<2>A new Thermus sp. class-IIS enzyme sub-family: isolation of a "twin" endonuclease TspDTI with a novel specificity 5'-ATGAA(N11/9)-3', related to TspGWI, TaqII and Tth111II.
<3>Nucleic Acids Res.
<4>31
<5>e74
<6>2003
<7>The TspDTI restriction endonuclease, which shows a novel recognition specificity
5'-ATGAA(N(11/9))-3', was isolated from Thermus sp. DT. TspDTI
appears to be a 'twin' of restriction endonuclease TspGWI from Thermus sp.
GW, as we have previously reported. TspGWI was isolated from the same
location as TspDTI, it recognizes a related sequence 5'-ACGGA(N(11/9))-3'
and has conserved cleavage positions. Both enzymes resemble two other
class-IIS endonucleases from Thermus sp.: TaqII and Tth111II. N-terminal
amino acid sequences of TspGWI tryptic peptides exhibit 88.9-100%
similarity to the TaqII sequence. All four enzymes were purified to
homogeneity; their polypeptide sizes (114.5-122 kDa) make them the largest
class-IIS restriction endonucleases known to date. The existence of a
Thermus sp. sub-family of class-IIS restriction endonucleases of a common
origin is herein proposed.

<>

<1>Skowron, P.M., Swaminathan, N., McMaster, K., George, D., Van Etten, J.L., Mead, D.A.
<2>Cloning and applications of the two/three-base restriction endonuclease R.CviJI from IL-3A virus-infected Chlorella.
<3>Gene
<4>157
<5>37-41
<6>1995
<7>The gene (cviJIR) encoding the two/three-base R.CviJI eukaryotic restriction endonuclease
(ENase) from IL-3A virus-infected Chlorella was cloned into Escherichia coli.  A high
frequency of DNA cleavage by R.CviJI required overexpression of the gene encoding the M.CviJI
methyltransferase prior to cloning the gene for the ENase.  Both genes were sequenced and
their organization was determined to be in head-to-tail order.  The open reading frame coding
for R.CviJI can potentially translate a 41.4-kDa protein; however, in the E. coli host, a
truncated version of the enzyme is produced (32.5 kDa).  The recombinant ENase does not
exhibit ATP-induced 'star' activity (R.CviJI cleaves at RGCY, while R.CviJI* also cleaves at
RGCR and YGCY, but not at YGCR), as is characteristic for native R.CviJI.  The very high
frequency of DNA cleavage by R.CviJI* was exploited in the development of a quasi-random
shotgun library method.  R.CviJI*-generated oligodeoxyribonucleotides were applied to improve
certain molecular biology applications, i.e., DNA labeling, detection, high-resolution
restriction mapping, amplification and epitope mapping.

<>

<1>Skowron, P.M., Vitkute, J., Ramanauskaite, D., Mitkaite, G., Jezewska-Frackowiak, J., Zebrowska, J., Zylicz-Stachula, A., Lubys, A.
<2>Three-stage biochemical selection: cloning of prototype class IIS/IIC/IIG restriction endonuclease-methyltransferase TsoI from the thermophile Thermus scotoductus.
<3>BMC Mol. Biol.
<4>14
<5>0
<6>2013
<7>Background: In continuing our research into the new family of bifunctional restriction
endonucleases (REases), we describe the
cloning of the tsoIRM gene. Currently, the family includes six
thermostable enzymes: TaqII, Tth111II, TthHB27I, TspGWI, TspDTI, TsoI,
isolated from various Thermus sp. and two thermolabile enzymes: RpaI
and CchII, isolated from mesophilic bacteria Rhodopseudomonas palustris
and Chlorobium chlorochromatii, respectively. The enzymes have several
properties in common. They are large proteins (molecular size app. 120
kDa), coded by fused genes, with the REase and methyltransferase
(MTase) in a single polypeptide, where both activities are affected by
S-adenosylmethionine (SAM). They recognize similar asymmetric cognate
sites and cleave at a distance of 11/9 nt from the recognition site.
Thus far, we have cloned and characterised TaqII, Tth111II, TthHB27I,
TspGWI and TspDTI.
Results: TsoI REase, which originate from thermophilic Thermus
scotoductus RFL4 (T. scotoductus), was cloned in Escherichia coli (E.
coli) using two rounds of biochemical selection of the T. scotoductus
genomic library for the TsoI methylation phenotype. DNA sequencing of
restriction-resistant clones revealed the common open reading frame
(ORF) of 3348 bp, coding for a large polypeptide of 1116 aminoacid (aa)
residues, which exhibited a high level of similarity to Tth111II (50%
identity, 60% similarity). The ORF was PCR-amplified, subcloned into a
pET21 derivative under the control of a T7 promoter and was subjected
to the third round of biochemical selection in order to isolate
error-free clones. Induction experiments resulted in synthesis of an
app. 125 kDa protein, exhibiting TsoI-specific DNA cleavage. Also, the
wild-type (wt) protein was purified and reaction optima were
determined.
Conclusions: Previously we identified and cloned the Thermus family
RM genes using a specially developed method based on partial
proteolysis of thermostable REases. In the case of TsoI the classic
biochemical selection method was successful, probably because of the
substantially lower optimal reaction temperature of TsoI (app. 10-15
degrees C). That allowed for sufficient MTase activity in vivo in
recombinant E. coli. Interestingly, TsoI originates from bacteria with
a high optimum growth temperature of 67 degrees C, which indicates that
not all bacterial enzymes match an organism's thermophilic nature, and
yet remain functional cell components. Besides basic research advances,
the cloning and characterisation of the new prototype REase from the
Thermus sp. family enzymes is also of practical importance in gene
manipulation technology, as it extends the range of available DNA
cleavage specificities.

<>

<1>Skowronek, K., Boniecki, M.J., Kluge, B., Bujnicki, J.M.
<2>Rational engineering of sequence specificity in R.MwoI restriction endonuclease.
<3>Nucleic Acids Res.
<4>40
<5>8579-8592
<6>2012
<7>R.MwoI is a Type II restriction endonucleases enzyme (REase), which specifically  recognizes a
palindromic interrupted DNA sequence 5'-GCNNNNNNNGC-3' (where N
indicates any nucleotide), and hydrolyzes the phosphodiester bond in the DNA
between the 7th and 8th base in both strands. R.MwoI exhibits remote sequence
similarity to R.BglI, a REase with known structure, which recognizes an
interrupted palindromic target 5'-GCCNNNNNGGC-3'. A homology model of R.MwoI in
complex with DNA was constructed and used to predict functionally important amino
acid residues that were subsequently targeted by mutagenesis. The model, together
with the supporting experimental data, revealed regions important for recognition
of the common bases in DNA sequences recognized by R.BglI and R.MwoI. Based on
the bioinformatics analysis, we designed substitutions of the S310 residue in
R.MwoI to arginine or glutamic acid, which led to enzyme variants with altered
sequence selectivity compared with the wild-type enzyme. The S310R variant of
R.MwoI preferred the 5'-GCCNNNNNGGC-3' sequence as a target, similarly to R.BglI,
whereas the S310E variant preferentially cleaved a subset of the MwoI sites,
depending on the identity of the 3rd and 9th nucleotide residues. Our results
represent a case study of a REase sequence specificity alteration by a single
amino acid substitution, based on a theoretical model in the absence of a crystal
structure.

<>

<1>Skowronek, K.J., Kosinski, J., Bujnicki, J.M.
<2>Theoretical model of restriction endonuclease HpaI in complex with DNA, predicted by fold recognition and validated by site-directed mutagenesis.
<3>Proteins
<4>63
<5>1059-1068
<6>2006
<7>Type II restriction enzymes are commercially important deoxyribonucleases and very attractive
targets for protein engineering
of new specificities. At the same time they are a very challenging test
bed for protein structure prediction methods. Typically, enzymes that
recognize different sequences show little or no amino acid sequence
similarity to each other and to other proteins. Based on
crystallographic analyses that revealed the same PD-(D/E)XK fold for
more than a dozen case studies, they were nevertheless considered to be
related until the combination of bioinformatics and mutational analyses
has demonstrated that some of these proteins belong to other, unrelated
folds PLD, HNH, and GIY-YIG. As a part of a large-scale project aiming
at identification of a three-dimensional fold for all type II REases
with known sequences (currently similar to 1000 proteins), we carried
out preliminary structure prediction and selected candidates for
experimental validation. Here, we present the analysis of Hpal REase,
an ORFan with no detectable homologs, for which we detected a
structural template by protein fold recognition, constructed a model
using the FRankenstein monster approach and identified a number of
residues important for the DNA binding and catalysis. These predictions
were confirmed by site-directed mutagenesis and in vitro analysis of
the mutant proteins. The experimentally validated model of Hpal will
serve as a low-resolution structural platform for evolutionary
considerations in the subgroup of blunt-cutting REases with different
specificities. The research protocol developed in the course of this
work represents a streamlined version of the previously used techniques
and can be used in a high-throughput fashion to build and validate
models for other enzymes, especially ORFans that exhibit no sequence
similarity to any other protein in the database.

<>

<1>Skrypina, N.A., Kramarov, V.M., Lyannaya, A.M., Smolyaninov, V.V., Goncharova, G.I.
<2>Restriction endonucleases from Bifidobacteria.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>15-16
<6>1988
<7>The site specific restriction endonucleases were found in four strains among
the twelve strains of anaerobic bacteria of the genus Bifidobacterium.  Two of
the restriction endonucleases studied, BadI from B. adolescentis LVA1 and BbfI
from B. bifidum LVA3, are isoschizomers of XhoI and recognize the nucleotide
sequence CTCGAG.  The restriction endonucleases Bbf7411I from B. bifidum 7411
and Bla7920I from B. lactentis 7920 recognize and hydrolyze the nucleotide
sequence TCCGGA having a specificity analogous to that of the restriction
endonuclease CauB3I.  Like CauB3I, these restriction endonucleases are unable
to hydrolyze DNA if the adenine residues in the recognition site are
methylated.

<>

<1>Skrzypek, E., Piekarowicz, A.
<2>The EcoDXXI restriction and modification system: cloning the genes and homology to Type I restriction and modification systems.
<3>Plasmid
<4>21
<5>195-204
<6>1989
<7>The Escherichia coli plasmid pDXX1 codes for a type I restriction and
modification system, EcoDXXI.  A 15.5-kb BamHI fragment from pDXX1 has been
cloned and contains the hsdR, hsdM, and hsdS genes that encode the EcoDXX1
system.  The EcoDXX1 hsd genes can complement the gene products of the EcoR124
and EcoR124/3 hsd systems, but not those of EcoK and EcoB.  Hybridization
experiments using EcoDXX1 hsd genes as a probe demonstrate homology between
EcoDXX1 and EcoK or EcoB systems.

<>

<1>Slack, A., Cervoni, N., Pinard, M., Szyf, M.
<2>Feedback regulation of DNA methyltransferase gene expression by methylation.
<3>Eur. J. Biochem.
<4>264
<5>191-199
<6>1999
<7>This paper tests the hypothesis that expression of the DNA methyltransferase, dnmt1, gene is
regulated by a methylation-sensitive DNA element. Methylation of DNA is an attractive system
for feedback regulation of DNA methyltransferase as the final product of the reaction,
methylated DNA, can regulate gene expression in cis. We show that an AP-1-dependent regulatory
element of dnmt1 is heavily methylated in most somatic tissues and in the mouse embryonal cell
line, P19, and completely unmethylated in a mouse adrenal carcinoma cell line, Y1. dnmt1 is
highly over expressed in Y1 relative to P19 cell lines. Global inhibition of DNA methylation
in P19 cells by 5-azadeoxycytidine results in demethylation of the AP-1 regulatory region and
induction of dnmt1 expression in P19cells, but not Y1 cells.  We propose that this regulatory
region of dnmt1 acts as a sensor of the DNA methylation capacity of the cell. These results
provide an explanation for the documented coexistence of global hypomethylation and high
levels of DNA methyltransferase activity in many cancer cells and for the carcinogenic effect
of hypomethylating diets.

<>

<1>Sladek, T.L., Maniloff, J.
<2>Endonuclease from Acholeplasma laidlawii strain JA1 associated with in vivo restriction of DNA containing 5-methylcytosine.
<3>Isr. J. Med. Sci.
<4>23
<5>423-426
<6>1987
<7>Transfection of Acholeplasma laidlawii strain JA1 with viral DNAs that have
been sequence-specifically methylated in vitro has led us to previously
postulate that JA1 cells contain an enzyme which cleaves only DNA containing
5-methylcytosine, regardless of the nucleotide sequence containing that base.
In this paper we show that an endonuclease activity is present in extracts from
JA1 cells but not in extracts from a restriction deficient variant of JA1.  The
partially purified enzyme cleaves to only DNA containing 5-methylcytosine, but
also DNA containing no methylated bases.  We discuss why this endonuclease
activity may not have the same in vitro specificity as would be expected from
in vivo experiments.

<>

<1>Sladek, T.L., Nowak, J.A., Maniloff, J.
<2>Mycoplasma restriction: identification of a new type of restriction specificity for DNA containing 5-methylcytosine.
<3>J. Bacteriol.
<4>165
<5>219-225
<6>1986
<7>Mycoplasma bacteriophage L51 single-stranded DNA and L2 double-stranded DNA are
host cell modified and restricted when they transfect Acholeplasma laidlawii
JA1 and K2 cells.  The L51 genome has a single restriction endonuclease MboI
site (recognition sequence GATC), which contains 5-methylcytosine when the DNA
is isolated from L51 phage grown in K2 cells but is unmethylated when the DNA
is from phage grown in JA1 cells.  This GATC sequence is nonessential, since an
L51 mutant in which the MboI site was deleted was still viable.  DNA from this
deletion mutant phage was not restricted during transfection of either strain
K2 or containing the sequence GAT 5-methylcytosine.  We conclude that K2 cells
have a restriction system specific for DNA containing the sequence GATC and
protect their DNA by methylating cytosine in this sequence.  In contrast, JA1
cells (which contain no methylated DNA bases) have a newly discovered type of
restriction-modification system.  From results of studies of the restriction of
specifically methylated DNAs, we conclude that JA1 cells restrict DNA
containing 5-methylcytosine, regardless of the nucleotide sequence containing
5-methylcytosine.  This is the first report of a DNA restriction activity
specific for a single (methylated) base.  Modification in this system is the
absence of cytosine methylating activity.  A restriction-deficient variant of
strain JA1, which retains the JA1 modification phenotype, was isolated,
indicating that JA1 cells have a gene product with restriction specificity for
DNA containing 5-methylcytosine.

<>

<1>Slaska-Kiss, K., Timar, E., Kiss, A.
<2>Complementation between inactive fragments of SssI DNA methyltransferase.
<3>BMC Mol. Biol.
<4>13
<5>17
<6>2012
<7>Background: Silencing mammalian genes by targeted DNA (cytosine-5) methylation of selected CG
sites in the genome would be a powerful technique to analyze epigenomic information and to
study the roles of DNA methylation in physiological and pathological states. A promising
approach of targeted DNA methylation is based on the ability of split fragments of a monomeric
DNA methyltransferase (C5-MTase) to associate and form active enzyme. A few C5-MTases of
different specificities have been shown to possess the ability of fragment complementation,
but a demonstration of this phenomenon for a C5-MTase, which has CG specificity and thus can
be targeted to methylate any CG site, has been lacking. The purpose of this study was to test
whether the CG-specific prokaryotic C5-MTase M.SssI shows the phenomenon of fragment
complementation.
Results: We show that truncated inactive N-terminal fragments of M.SssI can assemble with
truncated inactive C-terminal fragments to form active enzyme in vivo when produced in the
same E. coli cell. Overlapping and non-overlapping fragments as well as fragments containing
short appended foreign sequences had complementation capacity. In optimal combinations
C-terminal fragments started between conserved motif VIII and the predicted target recognizing
domain of M.SssI. DNA methyltransferase activity in crude extracts of cells with the best
complementing fragment pairs was ~ 4 per cent of the activity of cells producing the full
length enzyme.  Fusions of two N-terminal and two C-terminal fragments to 21.6 kDa zinc finger
domains only slightly reduced complementation ability of the fragments.
Conclusions: The CG-specific DNA methyltransferase M.SssI shows the phenomenon of fragment
complementation in vivo in E. coli. Fusion of the split fragments to six unit zinc finger
domains does not substantially interfere with the formation of active enzyme. These
observations and the large number of complementing fragment combinations representing a wide
range of MTase activity offer the possibility to develop M.SssI into a programmable DNA
ethyltransferase of high specificity.

<>

<1>Slater, S.C. et al.
<2>Genome Sequences of Three Agrobacterium Biovars Help Elucidate the Evolution of Multi-Chromosome Genomes in Bacteria.
<3>J. Bacteriol.
<4>8
<5>2501-2511
<6>2009
<7>The family Rhizobiaceae contains plant-associated bacteria with critical roles in ecology and
agriculture. Within this family, many Rhizobium and
Sinorhizobium strains are nitrogen-fixing plant mutualists, while many
strains designated as Agrobacterium are plant pathogens. These contrasting
lifestyles are primarily dependent on the transmissible plasmids each
strain harbors. Members of Rhizobiaceae also have diverse genome
architectures that include single chromosomes, multiple chromosomes, and
plasmids of various sizes. Agrobacterium strains have been divided into
three Biovars, based on physiological and biochemical properties. The
genome of a Biovar I strain, A. tumefaciens C58, has been previously
sequenced. In this study the genomes of the Biovar II strain A.
radiobacter K84, a commercially available biological control strain that
inhibits certain pathogenic agrobacteria, and the Biovar III strain A.
vitis S4, a narrow host range strain that infects grapes and invokes a
hypersensitive response on non-host plants, were fully sequenced and
annotated. Comparison with other sequenced members of the
alpha-proteobacteria provides new data on evolution of multi-partite
bacterial genomes. Primary chromosomes show extensive conservation of both
gene content and order. In contrast, secondary chromosomes share smaller
percentages of genes, and conserved gene order is restricted to short
blocks. We propose that secondary chromosomes originated from an ancestral
plasmid to which genes have been transferred from a progenitor primary
chromosome. Similar patterns are observed in select beta- and
gamma-proteobacteria species. Together these results define the evolution
of chromosome architecture and gene content among the Rhizobiaceae and
support a generalized mechanism for second chromosome formation among
bacteria.

<>

<1>Slatko, B.E., Benner, J.S., Jager-Quinton, T., Moran, L.S., Simcox, T.G., VanCott, E.M., Wilson, G.G.
<2>Cloning, sequencing and expression of the TaqI restriction-modification system.
<3>Nucleic Acids Res.
<4>15
<5>9781-9796
<6>1987
<7>The TaqI modification and restriction genes (recognition sequence TCGA) have
been cloned in E. coli and their DNA sequences have been determined.  Both
proteins were characterized and the N-terminal sequence of the endonuclease was
determined.  The genes have the same transcriptional orientation with the
methylase gene 5' to the endonuclease gene.  The methylase gene is 1089 bp in
length (363 amino acids, 40,575 daltons); the endonuclease gene is 702 bp in
length (234 amino acis, 27,523 daltons); they are separated by 132 bp.  Both
methylase and endonuclease activity can be detected in cell extracts.  The
clones fully modify the vector and chromosomal DNA but they fail to restrict
infecting phage.  Clones carrying only the restriction gene are viable even in
the absence of modification.  The restriction gene contains 7 TaqI sites; the
modification gene contains none.  This asymmetric distribution of sites could
be important in the regulation of the expression of the endonuclease gene.

<>

<1>Slatko, B.E., Croft, R., Moran, L.S., Wilson, G.G.
<2>Cloning and analysis of the HaeIII and HaeII methyltransferase genes.
<3>Gene
<4>74
<5>45-50
<6>1988
<7>The HaeIII methyltransferase (MTase) gene from Haemophilus aegyptius
(recognition sequence: 5'-GGCC-3') was cloned into Escherichia coli in the
plasmid vector pBR322.  The gene was isolated on a single EcoRI fragment and on
a single HindIII fragment.  Clones carrying additional adjacent fragments were
found to code also for the HaeII restriction endonuclease and HaeII
modification MTase (recognition sequence:  5'-PuGCGCPy-3').  The sequence of
the HaeIII modification gene was determined.  The inferred amino acid sequence
of the protein was found to share extensive similarity with other sequenced
m5C-MTases.  The central non-conserved region of the M.HaeIII MTase, thought to
form the nucleotide sequence-specificity domain, is almost identical to that of
the M.BsuRI, M.BspRI and M.NgoPII MTases, which also recognize the sequence
5'-GGCC-3'.

<>

<1>Slavin, J.W.J., Ivanisevic, A.
<2>Dual restriction enzyme digest of cationic-gold-coated DNA scaffolds.
<3>Int. J. Nanomedicine
<4>2
<5>821-825
<6>2007
<7>DNA strands coated with AuNPs were cleaved by restriction enzymes while in solution or on a
surface. Enzymatic activity was verified by gel
electrophoresis prior to surface analysis. Cleavage results suggest
that enzymes can recognize the AuNP-coated strands while on the
surfaces, though specificity in digestion has not yet been verified.
Development allows for advances in site specific localization of
components using biological media.

<>

<1>Slesarev, A.I., Mezhevaya, K.V., Makarova, K.S., Polushin, N.N., Shcherbinina, O.V., Shakhova, V.V., Belova, G.I., Aravind, L., Natale, D.A., Rogozin, I.B., Tatusov, R.L., Wolf, Y.I., Stetter, K.O., Malykh, A.G., Koonin, E.V., Kozyavkin, S.A.
<2>The complete genome of the hyperthermophile Methanopyrus kandleri AV19 and monophyly of Archaeal methanogens.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>4644-4649
<6>2002
<7>We have determined the complete 1,694,969-nt sequence of the GC-rich genome of Methanopyrus
kandleri by using a whole direct genome sequencing approach. This approach is based on
unlinking of genomic DNA with the ThermoFidelase version of M. kandleri topoisomerase V and
cycle sequencing directed by 2'-modified oligonucleotides (Fimers).
Sequencing redundancy (3.3x) was sufficient to assemble the genome with less than one error
per 40 kb. Using a combination of sequence database searches and coding potential prediction,
1,692 protein-coding genes and 39 genes for
structural RNAs were identified. M. kandleri proteins show an unusually high content of
negatively charged amino acids, which might be an adaptation to the high intracellular
salinity. Previous phylogenetic analysis of 16S RNA suggested that M. kandleri belonged to a
very deep branch, close to the root of the archaeal tree. However, genome comparisons indicate
that, in both trees constructed using concatenated alignments of ribosomal proteins and trees
based on gene content, M.
kandleri consistently groups with other archaeal methanogens. M. kandleri shares the set of
genes implicated in
methanogenesis and, in part, its operon organization with Methanococcus jannaschii and
Methanothermobacter
thermoautotrophicum. These findings indicate that archaeal methanogens are monophyletic. A
distinctive feature of M.
kandleri is the paucity of proteins involved in signaling and regulation of gene expression.
Also, M. kandleri appears to
have fewer genes acquired via lateral transfer than other archaea. These features might
reflect the extreme habitat of this
organism. Full author list: Slesarev,A.I., Mezhevaya,K.V., Makarova,K.S., Polushin,N.N.,
Shcherbinina,O.V., Shakhova,V.V., Belova,G.I., Aravind,L., Natale,D.A., Rogozin,I.B.,
Tatusov,R.L., Wolf,Y.I., Stetter,K.O., Malykh,A.G., Koonin,E.V., Kozyavkin,S.A.

<>

<1>Sletvold, H., Johnsen, P.J., Hamre, I., Simonsen, G.S., Sundsfjord, A., Nielsen, K.M.
<2>Complete sequence of Enterococcus faecium pVEF3 and the detection of an omega-epsilon-zeta toxin-antitoxin module and an ABC transporter.
<3>Plasmid
<4>60
<5>75-85
<6>2008
<7>Glycopeptide resistant Enterococcus faecium (GREF) persists on Norwegian
poultry farms despite the ban on the growth promoter avoparcin. The
biological basis for long-term persistence of avoparcin resistance is not
fully understood. This study presents the complete DNA sequence of the E.
faecium R-plasmid pVEF3 and functional studies of some plasmid-encoded
traits (a toxin-antitoxin (TA) system and an ABC transporter) that may be
of importance for plasmid persistence. The pVEF3 (63.1 kbp), isolated from
an E. faecium strain of poultry origin sampled in Norway in 1999, has 71
coding sequences including the vanA avoparcin/vancomycin resistance
encoding gene cluster. pVEF3 encodes the TA system omega-epsilon-zeta, and
plasmid stability tests and transcription analysis show that
omega-epsilon-zeta is functional in Enterococcus faecalis OGIX, although
with decreasing effect over time. The predicted ABC transporter was not
found to confer reduced susceptibility to any of the 28 substances tested.
The TA system identified in the pVEF-type plasmids may contribute to vanA
plasmid persistence on Norwegian poultry farms. However, size and
compositional heterogeneity among E. faecium vanA plasmids suggest that
additional plasmid maintenance systems in combination with host specific
factors and frequent horizontal gene transfer and rearrangement causes the
observed plasmid composition and distribution patterns.

<>

<1>Sloan, D.B., Moran, N.A.
<2>Genome reduction and co-evolution between the primary and secondary bacterial symbionts of psyllids.
<3>Mol. Biol. Evol.
<4>29
<5>3781-3792
<6>2012
<7>Genome reduction in obligately intracellular bacteria is one of the most
well-established patterns in the field of molecular evolution. In the extreme,
many sap-feeding insects harbor nutritional symbionts with genomes that are so
reduced that it is not clear how they perform basic cellular functions. For
example, the primary symbiont of psyllids (Carsonella) maintains one of the
smallest and most AT-rich bacterial genomes ever identified and has surprisingly
lost many genes that are thought to be essential for its role in provisioning its
host with amino acids. However, our understanding of this extreme case of genome
reduction is limited, as genomic data for Carsonella are available from only a
single host species, and little is known about the functional role of "secondary"
bacterial symbionts in psyllids. To address these limitations, we analyzed
complete Carsonella genomes from pairs of congeneric hosts in three divergent
genera within the Psyllidae (Ctenarytaina, Heteropsylla, and Pachypsylla) as well
as complete secondary symbiont genomes from two of these host species
(Ctenarytaina eucalypti and Heteropsylla cubana). Although the Carsonella genomes
are generally conserved in size, structure, and GC content and exhibit
genome-wide signatures of purifying selection, we found that gene loss has
remained active since the divergence of the host species and had a particularly
large impact on the amino acid biosynthesis pathways that define the symbiotic
role of Carsonella. In some cases, the presence of additional bacterial symbionts
may compensate for gene loss in Carsonella, as functional gene content indicates
a high degree of metabolic complementarity between co-occurring symbionts. The
genomes of the secondary symbionts also show signatures of long-term evolution as
vertically transmitted, intracellular bacteria, including more extensive genome
reduction than typically observed in facultative symbionts. Therefore, a history
of co-evolution with secondary bacterial symbionts can partially explain the
ongoing genome reduction in Carsonella. However, the absence of these secondary
symbionts in other host lineages indicates that the relationships are dynamic and
that other mechanisms, such as changes in host diet or functional coordination
with the host genome, must also be at play.

<>

<1>Slocum, H., Boyer, H.W.
<2>Host specificity of Salmonella typhimurium deoxyribonucleic acid restriction and modification.
<3>J. Bacteriol.
<4>113
<5>724-726
<6>1973
<7>The restriction and modification genes of Salmonella typhimurium which lie near
the thr locus were transferred to a restrictionless mutant of Escherichia coli.
These genes were found to be allelic to the E. coli K, B, and A restriction
and modification genes.  E. coli recombinants with the restriction and
modfication host specificity of S. typhimurium restricted phage lambda that had
been modified by each of the seven known host specificities of E. coli at an
efficiency of plating levels of abobut 10-2.  Phage lambda modified with the S.
typhimurium host specificity was restricted by six of the seven E. coli host
specificiteis but not by the RII (fi- R-factor controlled) host specificity.
It is proposed that the restriction and modification enzymes of this S.
typhimurium host specificity have two substrates, one of which is a substrate
for the RII host specificity enzymes.

<>

<1>Slocum, H.C.
<2>In vitro restriction of DNA.  Demonstration that the DNA of mutant phage are resistant to attack by restriction endonuclease.
<3>Tex. Rep. Biol. Med.
<4>29
<5>405
<6>1971
<7>Various bacterial systems (Escherichia coli, Hemophilus, and others)
demonstrate a mechanism for rejecting non-homologous DNA, i.e., DNA which
originates in a cell of a different strain.  This host-controlled restriction,
more specifically degradation, is brought about by a 2-step enzymatic mechanism
involving an endonuclease which introduces a limited number of double-strand
scissions at 2 specific sites (a site being a sequence of nucleotide base
pairs) on a DNA molecule.  The endonuclease does not recognize or introduce
double-strand scissions at these sites if there has been a prior methylation of
1 or 2 bases in the site (known as host-controlled modification of DNA).  The
modification methylase and restriction endonuclease recognize the same
substrate by virtue of the 2 enzymes having a common subunit which recognizes
the sequence of base pairs.  Small circular phage DNA have 2 sites recognized
by the endonuclease and methylase per molecule.  The biologic effect of the
restriction endonuclease is to abort the lytic cycle of the phage.  That is,
only 1 in 10,000 phage infected are successful.  Phage mutants can be obtained
that have successful lytic cycles of 1 in 100.  I recombined 2 independent
mutants to obtain a phage totally resistant to the endonuclease, i.e., a
probability of 1.0 that the lytic cycle takes place.  In vitro, the single
mutants are degraded to intact linear molecules, whereas the recombinant is
completely resistant to degradation, meaning no single strand breaks are made
at either mutant site.  The results can be interpreted on the basis of the
substrate being a sequence of base pairs with 2-fold dimensional symmetry.

<>

<1>Smajs, D., Zobanikova, M., Strouhal, M., Cejkova, D., Dugan-Rocha, S., Pospisilova, P., Norris, S.J., Albert, T., Qin, X., Hallsworth-Pepin, K., Buhay, C., Muzny, D.M., Chen, L., Gibbs, R.A., Weinstock, G.M.
<2>Complete Genome Sequence of Treponema paraluiscuniculi, Strain Cuniculi A: The Loss of Infectivity to Humans Is Associated with Genome Decay.
<3>PLoS ONE
<4>6
<5>e20415
<6>2011
<7>Treponema paraluiscuniculi is the causative agent of rabbit venereal spirochetosis. It is not
infectious to humans, although its
genome structure is very closely related to other pathogenic Treponema species including
Treponema pallidum subspecies pallidum, the etiological agent of syphilis. In this study, the
genome sequence of Treponema paraluiscuniculi, strain Cuniculi A, was determined by a
combination of several high-throughput sequencing strategies. Whereas the overall size
(1,133,390 bp), arrangement, and gene content of the Cuniculi A genome closely resembled those
of the T. pallidum genome, the T. paraluiscuniculi genome contained a markedly higher number
of pseudogenes and gene fragments (51). In addition to pseudogenes, 33 divergent genes were
also found in the T. paraluiscuniculi genome. A set of 32 (out of 84) affected genes encoded
proteins of known or predicted function in the Nichols genome. These proteins included
virulence factors, gene regulators and components of DNA repair and recombination. The
majority (52 or 61.9%) of the Cuniculi A
pseudogenes and divergent genes were of unknown function. Our results indicate that T.
paraluiscuniculi has evolved from a T. pallidum-like ancestor and adapted to a specialized
host-associated niche (rabbits) during loss of infectivity to humans. The genes that are
inactivated or altered in T. paraluiscuniculi are candidates for virulence factors important
in the infectivity and pathogenesis of T. pallidum subspecies.

<>

<1>Smet, A., Van Nieuwerburgh, F., Ledesma, J., Flahou, B., Deforce, D., Ducatelle, R., Haesebrouck, F.
<2>Genome Sequence of Helicobacter heilmannii Sensu Stricto ASB1 Isolated from the Gastric Mucosa of a Kitten with Severe Gastritis.
<3>Genome Announcements
<4>1
<5>e00033-12
<6>2013
<7>Here we report the genome sequence of Helicobacter heilmannii sensu stricto ASB1  isolated
from the gastric mucosa of a kitten with severe gastritis. Helicobacter heilmannii sensu
stricto has also been associated with gastric disease in humans. Availability of this genome
sequence will contribute to the identification of genes involved in the pathogen's virulence
and carcinogenic properties.

<>

<1>Smet, A., Van Nieuwerburgh, F., Vandekerckhove, T.T., Martel, A., Deforce, D., Butaye, P., Haesebrouck, F.
<2>Complete nucleotide sequence of CTX-M-15-plasmids from clinical Escherichia coli isolates: insertional events of transposons and insertion sequences.
<3>PLoS ONE
<4>5
<5>E11202
<6>2010
<7>BACKGROUND: CTX-M-producing Escherichia coli strains are regarded as major
global pathogens. METHODOLOGY/PRINCIPAL FINDINGS: The nucleotide sequence
of three plasmids (pEC_B24: 73801-bp; pEC_L8: 118525-bp and pEC_L46:
144871-bp) from Escherichia coli isolates obtained from patients with
urinary tract infections and one plasmid (pEC_Bactec: 92970-bp) from an
Escherichia coli strain isolated from the joint of a horse with arthritis
were determined. Plasmid pEC_Bactec belongs to the IncI1 group and carries
two resistance genes: bla(TEM-1) and bla(CTX-M-15). It shares more than
90% homology with a previously published bla(CTX-M)-plasmid from E. coli
of human origin. Plasmid pEC_B24 belongs to the IncFII group whereas
plasmids pEC_L8 and pEC_L46 represent a fusion of two replicons of type
FII and FIA. On the pEC_B24 backbone, two resistance genes, bla(TEM-1) and
bla(CTX-M-15), were found. Six resistance genes, bla(TEM-1),
bla(CTX-M-15), bla(OXA-1), aac6'-lb-cr, tetA and catB4, were detected on
the pEC_L8 backbone. The same antimicrobial drug resistance genes, with
the exception of tetA, were also identified on the pEC_L46 backbone.
Genome analysis of all 4 plasmids studied provides evidence of a seemingly
frequent transposition event of the bla(CTX-M-15)-ISEcp1 element. This
element seems to have a preferred insertion site at the tnpA gene of a
bla(TEM)-carrying Tn3-like transposon, the latter itself being inserted by
a transposition event. The IS26-composite transposon, which contains the
bla(OXA-1), aac6'-lb-cr and catB4 genes, was inserted into plasmids pEC_L8
and pEC_L46 by homologous recombination rather than a transposition event.
Results obtained for pEC_L46 indicated that IS26 also plays an important
role in structural rearrangements of the plasmid backbone and seems to
facilitate the mobilisation of fragments from other plasmids. CONCLUSIONS:
Collectively, these data suggests that IS26 together with ISEcp1 could
play a critical role in the evolution of diverse multiresistant plasmids
found in clinical Enterobacteriaceae.

<>

<1>Smirnova, N.I., Cherkasov, A.V., Krasnov, Y.M., Agafonov, D.A., Kutyrev, V.V.
<2>Draft Whole-Genome Sequence of a New Variant of Vibrio cholerae O1 El Tor Strain  Isolated from a Cholera Patient in Russia.
<3>Genome Announcements
<4>2
<5>e00432-14
<6>2014
<7>Draft whole-genome sequencing of the Vibrio cholerae capital O, Cyrillic1 El Tor  clinical
strain L3226, isolated in Moscow in 2010, was carried out. Various
mutations in the virulence-associated mobile elements were determined in its
genome that differentiated this strain from the reference V. cholerae capital O,
Cyrillic1 El Tor strain N16961.

<>

<1>Smith, A.E., Ford, K.G.
<2>Specific targeting of cytosine methylation to DNA sequences in vivo.
<3>Nucleic Acids Res.
<4>35
<5>740-754
<6>2007
<7>Development of methods that will allow exogenous imposition of inheritable gene-specific
methylation patterns has potential application in both
therapeutics and in basic research. An ongoing approach is the use of
targeted DNA methyltransferases, which consist of a fusion between
gene-targeted zinc-finger proteins and prokaryotic DNA cytosine
methyltransferases. These enzymes however have so far demonstrated
significant and unacceptable levels of non-targeted methylation. We now
report the development of second-generation targeted methyltransferase
enzymes comprising enhanced zinc-finger arrays coupled to
methyltransferase mutants that are functionally dominated by their
zinc-finger component. Both in vitro plasmid methylation studies and a
novel bacterial assay reveal a high degree of target-specific methylation
by these enzymes. Furthermore, we demonstrate for the first time transient
expression of targeted cytosine methyltransferase in mammalian cells
resulting in the specific methylation of a chromosomal locus. Importantly,
the resultant methylation pattern is inherited through successive cell
divisions.

<>

<1>Smith, B.A., Dougherty, K.M., Baltrus, D.A.
<2>Complete Genome Sequence of the Highly Transformable Pseudomonas stutzeri Strain  28a24.
<3>Genome Announcements
<4>2
<5>e00543-14
<6>2014
<7>Here, we report the complete genome sequence for an isolate of Pseudomonas stutzeri that is
highly competent for natural transformation. This sequence
enables insights into the genetic basis of natural transformation rate variations
and provides an additional data point for genomic comparisons across a ubiquitous
and highly diverse bacterial species.

<>

<1>Smith, C.J., Parker, A., Rogers, M.B.
<2>Plasmid transformation of Bacteroides spp. by electroporation.
<3>Plasmid
<4>24
<5>100-109
<6>1990
<7>Transformation of Bacterioides spp. with a variety of plasmid DNAs was accomplished using
electroporation. The standard transformation assay system used to deduce the optimal
electroporation parameters employed a 50-to 100-fold concentrated cell suspension of
mid-logarithmic phase Bacterioides fragilis strain 638 and the 5.4-kb clindamycin resistance
(Ccr) vector pBI191. A variety of electroporation buffers were used successfully in
transformation experiments but of these 1 mM MgCl2 in 10% glycerol was superior. The
incorporation of MgCl2 was essential for optimum viability prior to electroporation and for
optimum transformation. Transformants were routinely obtained using 5-ms pulses over a range
of field strengths from 5 to 12.5 kV/cm, with a maximum of >1000000/ug DNA at 12.5 kV/cm. The
number of transformants increased linearly with respect to DNA concentration over the range
0.01-2 ug tested. Recovery of transformants required an expression period of up to 2.5 h
following exposure to the electric field. This period, however, was dependent on the
antibiotic resistance marker used for selection of transformants, with a significantly shorter
incubation required when chloramphenicol rather than clindamycin was used in the selective
medium. The effect of the DNA source of transformation was tested using the shuttle vector
pFD288. Plasmid DNA isolated from Bacteriodes uniformis, Bacteroides ovatus, or Bacteroides
thetaiotaomicron transformed B. fragilis 638 at frequencies 7.5 to 12.5 fold less than those
observed for controls with homologous DNA. Further reductions were seen with Escherichia coli
purified pFD288, which transformed at 1000-fold lower frequencies. Finally, using homologous
pFD288 or pBI191 isolated from strain 638, several strains of B. fragilis, B. uniformis, and
B. ovatus were transformed successfully without modification of the standard assay system. Two
strains each of B. thetaiotaomicron and Baceriodes ruminicola were not transformed using the
methods described here.

<>

<1>Smith, D., Alm, R.A., Doig, P.C., Kabok, Z., Castriotta, L.M.
<2>Nucleic acid and amino acid sequences relating to Helicobacter pylori and vaccine compositions thereof.
<3>Japanese Patent Office
<4>JP 2001510992 A
<5>
<6>2001
<7>
<>

<1>Smith, D.D., Kirzinger, M.W., Stavrinides, J.
<2>Draft Genome Sequence of the Antibiotic-Producing Cystic Fibrosis Isolate Pantoea agglomerans Tx10.
<3>Genome Announcements
<4>1
<5>e00904-13
<6>2013
<7>Pantoea agglomerans is an enteric bacterium that is capable of causing both plant and human
disease. Here, we report the genome sequence of a cystic fibrosis
isolate, P. agglomerans Tx10, which produces an antibiotic that is effective
against Staphylococcus aureus.

<>

<1>Smith, D.D., Kirzinger, M.W., Stavrinides, J.
<2>Draft Genome Sequence of the Antibiotic-Producing Epiphytic Isolate Pantoea ananatis BRT175.
<3>Genome Announcements
<4>1
<5>e00902-13
<6>2013
<7>Pantoea is a member of the Enterobacteriaceae, whose members have been shown to produce novel
antibiotics. Here, we report the 4.8-Mb genome sequence of Pantoea
ananatis strain BRT175, an epiphytic isolate from strawberries that produces an
antibiotic that is effective against the fire blight pathogen, Erwinia amylovora.

<>

<1>Smith, D.E., Gemmen, G.J., Millin, R.
<2>DNA looping and cleavage by restriction enzymes studied by manipulation of single DNA molecules with optical tweezers.
<3>Proc. SPIE-Int. Soc. Opt. Eng., SPIE-The International Society for Optical Engineering, Dholakia, K., Spalding, G.C., 
<4>6326
<5>1-13
<6>2006
<7>Looping and cleavage of single DNA molecules by the two-site restriction endonuclease Sau3AI
were measured with optical tweezers.  A DNA template containing many recognition sites was
used, permitting loop sizes from ~10 to 10,000 basepairs.  At high enzyme concentration
cleavage events were detected within 5 seconds and nearly all molecules were cleaved within 5
minutes.  Activity decreased ~10-fold as the DNA tension was increased from 0.03 to 0.7 pN.
Substituting Ca2+ for Mg2+ blocked cleavage, permitting measurement of stable loops.  At low
tension, the initial rates of cleavage and looping were similar (~0.025 s-1 at 0.1 pN),
suggesting that looping is rate limiting.  Short loops formed more rapidly than long loops.
The optimum size decreased from ~250 to 45 bp and the average number of loops (in 1 minute)
from 4.2 to 0.75 as tension was increased from 0.03 to 0.7 pN.  No looping was detected at 5
pN.  These findings are in qualitative agreement with recent theoretical predictions
considering only DNA mechanics, but we observed weaker suppression with tension and smaller
loop sizes.  Our results suggest that the span and elasticity of the protein complex and
protein-induced DNA bending and wrapping play an important role.

<>

<1>Smith, D.I., Blattner, F.R., Davies, J.
<2>The isolation and partial characterization of a new restriction endonuclease from Providencia stuartii.
<3>Nucleic Acids Res.
<4>3
<5>343-353
<6>1976
<7>We describe the isolation of a new class 2 restriction endonuclease from
Providencia stuartii 164.  Using the procedure of osmotic shock treatment, we
have partially purified this enzyme (PstI) and have begun preliminary work to
characterize its specificity and requirements.  PstI requires Mg++ as the only
cofactor and produces more than 18 cleavages in wild type lambda.  We have
determined the location of 7 of these cleavages by the use of deletion and
insertion mutants of lambda.

<>

<1>Smith, D.I., Golembieski, W., Gilbert, J.D., Kizyma, L., Miller, O.J.
<2>Overabundance of rare-cutting restriction endonuclease sites in the human genome.
<3>Nucleic Acids Res.
<4>15
<5>1173-1184
<6>1987
<7>A human chromosome 3-specific cosmid library was constructed from a somatic
cell hybrid containing human chromosome 3 as its only human component.  This
library was screened to identify 230 human recombinants which contained an
average insert size of 37 kilobases.  DNA prepared from 54 of these cosmids,
representing 2000 kilobases of human DNA, was then tested for restriction
endonuclease sites for EcoRI, HindIII, KpnI, XhoI, and DraI, as well as those
of the rare-cutting restriction endonucleases NotI, SfiI, NruI, MluI, SacII,
and BssHII.  Sites for the latter enzymes were much more abundant than would be
expected from theoretical calculations, reflecting non-random clustering of
these sites.  This has important implications for the use of these enzymes in
the construction of physical maps of chromosomes.  Some individual cosmids
contained large numbers of rare sites, offering an alternative means of
physically mapping chromosomes based upon identifying clusters of rare
restriction sites.  These clusters appear to be spaced an average of 1000 kb
apart.

<>

<1>Smith, D.R.
<2>Restriction endonuclease digestion of DNA..
<3>Methods Mol. Biol.
<4>58
<5>11-15
<6>1996
<7>The ability to cleave DNA at specific sites is one of the cornerstones of today's methods of
DNA manipulation.  Restriction endonucleases are bacterial enzymes that cleave duplex DNA at
specific target sequences with the production of defined fragments.  These enzymes can be
purchased from the many manufacturers of biotechnology products.  The nomenclature of enzymes
is based on a simple system, proposed by Smith and Nathans.  The name of the enzyme (such as
BamHI, EcoRI, and so on) tells us about the origin of the enzyme, but does not give us any
information about the specificity of cleavage.  This has to be determined for each individual
enzyme.  The recognition site for most of the commonly used enzymes is a short palindromic
sequence, usually either 4, 5, or 6 bp in length, such as AGCT (for AluI), GAATTC (for EcoRI),
and so on.  Each enzyme cuts the palindrome at a particular site, and two different enzymes
may have the same recognition sequence, but cleave the DNA at different points within that
sequence.  The cleavage sites fall into three different categories, either flush (or blunt) in
which the recognition site is cut in the middle, or with either 5'- or 3'-overhangs, in
which case unpaired bases will be produced on both ends of the fragment.  For a comprehensive
review of restriction endonucleases, see Fuchs and Blakesley.

<>

<1>Smith, D.R. et al.
<2>Complete genome sequence of Methanobacterium thermoautotrophicum delta H: functional analysis and comparative genomics.
<3>J. Bacteriol.
<4>179
<5>7135-7155
<6>1997
<7>The complete 1,751,377-bp sequence of the genome of the thermophilic archaeon Methanobacterium
thermoautotrophicum deltaH has been determined by a whole-genome shotgun sequencing approach.
A total of 1,855 open reading frames have been identified that appear to encode polypeptides,
844 (46%) of which have been assigned putative functions based on their similarities to
database sequences with assigned functions.  A total of 514 (28%) of the ORF-encoded
polypeptides are related to sequences with unknown functions, and 496 (27%) have little or no
homology to sequences in public databases.  Comparisons with Eucarya-, Bacteria-, and
Archaea-specific databases reveal that 1,013 of the putative gene products (54%) are most
similar to polypeptide sequences described previously for other organisms in the domain
Archaea.  Comparisons with the Methanococcus jannaschii genome data underlie the extensive
divergence that has occurred between these two methanogens; only 352 (19%) of M.
thermoautotrophicum ORFs encode sequences that are >50% identical to M. jannaschii
polypeptides, and there is little conservation in the relative locations of orthologous genes.
When the M. thermoautotrophicum ORFs are compared to sequences from only the eucaryal and
bacterial domains, 786 (42%) are more similar to bacterial sequences and 241 (13%) are more
similar to eucaryal sequences.  The bacterial domain-like gene products include the majority
of those predicted to be involved in cofactor and small molecule biosyntheses, intermediary
metabolism, transport, nitrogen fixation, regulatory functions, and interactions with the
environment.  Most proteins predicted to be involved in DNA metabolism, transcription, and
translation are more similar to eucaryal sequences.  Gene structure and organization have
features that are typical of the Bacteria, including genes that encode polypeptides closely
related to eucaryal proteins.  There are 24 polypeptides that could form two-component sensor
kinase-response regulator systems and homologs of the bacterial Hsp70-response proteins DnaK
and DnaJ, which are notably absent in M. jannaschii.  DNA replication initiation and
chromosome packaging in M. thermoautotrophicum are predictd to have eucaryal features, based
on the presence of two Cdc6 homologs and three histones; however, the presence of an ftsZ gene
indicates a bacterial type of cell division initiation.  The DNA polymerases include an
X-family repair type and an unusual archaeal B type formed by two separate polypeptides.  The
DNA-dependent RNA polymerase subunits A', A", B', B" and H are encoded in a typical archaeal
RNAP operon, although a second A' subunit-encoding gene is present at a remote location.
There are two rRNA operons, and 39 tRNA genes are dispersed around the genome, although most
of these occur in clusters.  Three of the tRNA genes have introns, including the tRNAPro (GGG)
gene, which contains a second intron at an unprecedented location.  There is no
selenocysteinyl-tRNA gene nor evidence for classically organized IS elements, prophages, or
plasmids.  The genome contains one intein and two extended repeats (3.6 and 8.6 kb) that are
members of a family with 18 representatives in the M. jannaschii genome.

<>

<1>Smith, D.R., Mao, J.-i.
<2>Nucleic acid and amino acid sequences relating to Mycobacterium tuberculosis and leprae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 6583266 A
<5>
<6>2003
<7>Embodiments of the present invention feature nucleic acid and proteins derived from
Mycobacterium tuberculosis and leprae.  The proteins and nucleic acid of the present invention
have applications in diagnostics and therapeutics.

<>

<1>Smith, D.W., Crowder, S.W., Reich, N.O.
<2>In vivo specificity of EcoRI DNA methyltransferase.
<3>Nucleic Acids Res.
<4>20
<5>6091-6096
<6>1992
<7>The EcoRI adenine DNA methyltransferase forms part of a bacterial restriction/modification
system; the methyltransferase modifies the second adenine within the canonical site GAATTC,
thereby preventing the EcoRI endonuclease from cleaving this site. We show that five
noncanonical EcoRI sites (TAATTC, CAATTC, GTATTC, GGATTC and GAGTTC) are not methylated in
vivo under conditons when the canonical site is methylated. Only when the methyltransferase is
overxpressed is partial in vivo methylation of the five sites detected. Our results suggest
that the methyltransferase does not protect host DNA against potential endonuclease-mediated
cleavage at noncanonical sites. Our related in vitro analysis of the methyltransferase reveals
a low level of sequence-discrimination. We propose that the high in vivo specificity may be
due to the active removal of methylated sequences by DNA repair enzymes (J. Bacteriology
(1987), 169 3243-3250).

<>

<1>Smith, H., Akiyama, T., Foreman, C., Franklin, M., Woyke, T., Teshima, H., Davenport, K., Daligault, H., Erkkila, T., Goodwin, L., Gu, W., Xu, Y., Chain, P.
<2>Draft Genome Sequence and Description of Janthinobacterium sp. Strain CG3, a Psychrotolerant Antarctic Supraglacial Stream Bacterium.
<3>Genome Announcements
<4>1
<5>e00960-13
<6>2013
<7>Here we present the draft genome sequence of Janthinobacterium sp. strain CG3, a
psychrotolerant non-violacein-producing bacterium that was isolated from the
Cotton Glacier supraglacial stream. The genome sequence of this organism will
provide insight as to the mechanisms necessary for bacteria to survive in
UV-stressed icy environments.

<>

<1>Smith, H.J., Foreman, C.M., Ramaraj, T.
<2>Draft Genome Sequence of a Metabolically Diverse Antarctic Supraglacial Stream Organism, Polaromonas sp. Strain CG9_12, Determined Using Pacific Biosciences  Single-Molecule Real-Time Sequencing Technology.
<3>Genome Announcements
<4>2
<5>e01242-14
<6>2014
<7>Polaromonas species are found in a diversity of environments and are particularly common in
icy ecosystems. Polaromonas sp. strain CG9_12 is an aerobic,
Gram-negative, catalase-positive, white-pigmented bacterium of the Proteobacteria
phylum. Here, we present the draft genome sequence of Polaromonas sp. strain
CG9_12, isolated from an Antarctic supraglacial stream.

<>

<1>Smith, H.O.
<2>Restriction endonucleases as enzymatic tools for DNA analysis and genetic manipulation.
<3>PAABS Revista
<4>5
<5>313-319
<6>1976
<7>Living cells contain a variety of nucleases capable of cleaving the
phosphodiester bonds of nucleic acid chains.  These are generally of two kinds:
exonucleases that act at chain termini to release nucleotides or
oligonucleotides, and endonucleases that cleave at internal bonds.  Many
endonucleases act randomly, cleaving each phosphodiester bond with about equal
probability regardless of the local base sequence.  Several years ago a new
class of DNA nucleases that cleave both strands of double-stranded DNA at sites
of specific base sequence, four to six bases in length, was discovered in
bacteria.  These endonucleases are called restriction enzymes because they are
components of bacterial restriction-modification (R-M) systems that determine a
host-specific DNA immunity mechanism.  Each R-M system consists of a
restriction endonuclease and a modification methylase of similar site
specificity.  The modification enzyme methylates sites in the host chromosome
to protect against cleavage by their restriction endonuclease.  However,
improperly methylated foreign DNA entering the cell, for example, by virus
infection, will be cleaved and subsequently degraded to nucleotides by cellular
exonucleases.  Since their discovery, restriction endonucleases have been
widely exploited as new tools for DNA analysis because of their ability to
cleave DNA molecules at only a few sites to produce specific DNA fragments.
Restriction fragments from a given DNA molecule may be ordered into a physical
map that can serve as a framework for location of genetic functions and for
nucleotide sequence analysis.  The possibilities of using restriction
endonucleases for genetic manipulation have aroused great interest.  Novel
kinds of recombinant DNA molecules can be constructed by rearranging DNA
fragments within a chromosome or by joining fragments from one species to those
of another to form hybrid chromosomes.  Fragments bearing specific genes may be
joined to self-replicating viral or plasmid chromosomes and isolated by
molecular cloning in suitable host bacteria.  The recombinant plasmid or virus
DNAs often exist as multiple copies intracellularly, thus providing large
quantities of the inserted DNA fragments for sequence analysis and studies of
gene organization and function.  In many cases, products of inserted genes,
such as specific enzymes, are obtained in high yield.  In a brief 3 to 4 years,
a whole new recombinant DNA technology has arisen that has many important
potential commercial as well as research applications. This review will briefly
cover properties of restriction endonucleases and their major applications.
Comprehensive reviews have been written by Arber and Nathans and Smith.

<>

<1>Smith, H.O.
<2>Nucleotide sequence specificity of restriction endonucleases.
<3>Science
<4>205
<5>455-462
<6>1979
<7>In the past 7 to 8 years we have witnessed the development of a new DNA
technology that has fundamentally altered our approach to modern genetics.  The
basic ingredients of this new technology are the cleavage site-specific
restriction enzymes: a special class of bacterial endonucleases that can
recognize specific nucleotide sequenceas in duplex DNA and produce
double-stranded cleavages.  A collection of these enzymes, each with its own
particular sequence specificity, can be used to cleave DNA molecules into
unique sets of fragments for DNA sequencing, chromosome analysis, gene
isolation, and construction of recombinant DNA.  The latter, together with the
concept of molecular cloning, has given birth to the new field of genetic
engineering, and from this many new and exciting medical and research
applications are expected.  My own role in these developments occurred
primarily in the period of 1968 to 1970 when my colleagues and I made the
chance discovery of the first of the cleavage site-specific restriction
enzymes.  I now briefly present this work in historical context because it
leads naturally into the main part of my lecture, describing our present
knowledge of restriction and modification enzymes.  Although many applications
have been reviewed, I should like to describe in some detail the use of these
enzymes as model systems for studying sequence-specific interactions of protein
and DNA, which is one of the major research interests in my laboratory.

<>

<1>Smith, H.O.
<2>Restriction endonuclease from Hemophilus influenzae Rd.
<3>Methods Mol. Biol.
<4>7
<5>71-85
<6>1974
<7>A growing number of molecular biologists are turning to restriction enzymes as
tools for analysis of DNA molecules and chromosomal DNA.  Restriction enzymes
are site-specific endonucleases that can be isolated from various strains of
bacteria.  They are capable of recognizing particular base sequences within
native DNA molecules and producing double-strand cleavage.  Thus viral genomes
or other DNA molecules may be cleaved in a highly specific and reproducible
manner into a number of double-stranded fragments.  These are useful for
localization of genetic functions, DNA base sequencing, and a number of other
purposes.  This article describes the purification of restriction enzyme from
H. influenzae Rd.  The procedure is similar to that reported by Smith and
Wilcox but incorporates modifications which facilitate assay and large scale
purification.

<>

<1>Smith, H.O., Annau, T.M., Chandrasegaran, S.
<2>Finding sequence motifs in groups of functionally related proteins.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>87
<5>826-830
<6>1990
<7>We have developed a method for rapidly finding patterns of conserved amino acid
residues (motifs) in groups of functionally related proteins.  All 3-amino acid
patterns in a group of proteins of the type aa1 d1 aa2 d2 aa3, where d1 and d2
are distances that can be varied in a range up to 24 residues, are accumulated
into an array.  Segments of the proteins are aligned on each other by a scoring
method that obtains an average relatedness value for all the amino acids in
each column of the aligned sequence block based on the Dayhoff relatedness odds
matrix.  The automated method successfully finds and displays nearly all of the
sequence motifs that have been previously reported to occur in 33 reverse
transcriptases, 18 DNA integrases, and 30 DNA methyltransferases.

<>

<1>Smith, H.O., Birnstiel, M.L.
<2>A simple method for DNA restriction site mapping.
<3>Nucleic Acids Res.
<4>9
<5>2387-2399
<6>1976
<7>When a DNA molecule, enzymatically labelled with 32P at one end, is partially
digested with a restriction enzyme labelled DNA fragments are obtained which
form an overlapping series of molecules, all with a common labelled terminus.
A restriction map can then be constructed from an analysis of the size
distribution of these molecules.  This technique has been used for the
restriction site mapping of cloned histone DNA (h22) where as many as 35
cleavage sites may be accurately determined in a single experiment.

<>

<1>Smith, H.O., Kelly, S.V.
<2>Methylases of the type II restriction-modification systems.
<3>DNA Methylation. Biochemistry and Biological Significance., Springer-Verlag, Razin, A., Cedar, H., Riggs, A.D., New York
<4>0
<5>39-71
<6>1984
<7>Most bacteria contain a small fraction (0.5-2%) of methylated cytosine or
adenine bases in their chromosomes.  Some of this methylation apparently plays
a role in directing mismatch repair systems to the correct strands in newly
replicated DNA.  However, in many bacterial strains, a substantial fraction can
be identified with host-specific restriction-modification (RM) systems.  These
ubiquitous systems play an important biological role in protecting bacteria
against viral infections.  Each system has two functional components: 1) a
restriction endonuclease capable of recognizing sequence-specific sites in DNA
and producing double-stranded cleavage; and 2) a modification enzyme
recognizing the same DNA sites as the restriction enzyme and protecting them by
modification.  So far, all modifications found are either 5-methylcytosine or
6-methyladenine.  The modification enzymes are methyltransferases, and AdoMet
appears to be the exclusively methyl group donor.  (For reviews, see Arber,
1974; Modrich, 1979; Smith, 1979; Modrich and Roberts, 1982).

<>

<1>Smith, H.O., Kelly, T.J., Roy, P.H.
<2>Enzymatic methods for sequence analysis applied to DNA restriction and methylation sites.
<3>Methods Enzymol.
<4>29
<5>282-294
<6>1974
<7>Restriction enzymes are site-specific endonucleases produced by various
bacteria, bacterial plasmids, and viruses.  In those cases where they produce
double-stranded cleavage of DNA within their recognition sites, sequence
analysis of the site is equivalent to determination of the 5'- and 3'-terminal
bases of the restricted DNA.  We have developed special enzymatic methods for
limited analysis of this type.  In addition, every host that produces a
restriction endonuclease also produces a companion DNA modification enzyme,
usually a methylase, with identical or similar site recognition.  This enzyme
serves to modify and protect the restriction sites of the host chromosome.  In
the second section of this chapter we describe a procedure for partial analysis
of methylation sites.

<>

<1>Smith, H.O., Marley, G.M.
<2>Purification and properties of HindII and HindIII endonucleases from Haemophilus influenzae Rd.
<3>Methods Enzymol.
<4>65
<5>104-108
<6>1980
<7>Haemophilus influenzae Rd is the source for two restriction endonucleases,
HindII and HindIII, that may be separated and purified using conventional ion
exchange chromatography media.  The basic steps in the procedure are removal of
nucleic acids from the cell extract, separation of the two activities by
DEAE-cellulose chromatography, and chromatography of each activity on
phosphocellulose as a combined purification and concentration step.

<>

<1>Smith, H.O., Nathans, D.
<2>A suggested nomenclature for bacterial host modification and restriction systems and their enzymes.
<3>J. Mol. Biol.
<4>81
<5>419-423
<6>1973
<7>In the proposed nomenclature restriction-modification systems are named
according to host organism and strain.  Different R-M+ systems in a single host
are designated by Roman numerals.  Restriction nucleases and modification
methylases are given the general names endonuclase R and methylase M, followed
by their R-M system name.

<>

<1>Smith, H.O., Wilcox, K.W.
<2>A restriction enzyme from Hemophilus influenzae. I. Purification and general properties.
<3>J. Mol. Biol.
<4>51
<5>379-391
<6>1970
<7>Extracts of Hemophilus influenzae strain Rd contain an endonuclease activity
which produces a rapid decrease in the specific viscosity of a variety of
foreign native DNA's; the specific viscosity of H. influenzae DNA is not
altered under the same conditions.  This "restriction" endonuclease activity
has been purified approximately 200-fold.  The purified enzyme contains no
detectable exo- or endonucleolytic activity against H. influenzae DNA.
However, with native phage T7 DNA as substrate, it produces about 40
double-strand 5'-phosphoryl, 3'-hydroxyl cleavages.  The limit product has an
average length of about 1000 nucleotide pairs and contains no single-strand
breaks.  The enzyme is inactive on denatured DNA and it requires no special
co-factors other than magnesium ions.

<>

<1>Smith, H.R., Humphreys, G.O., Willshaw, G.A., Anderson, E.S.
<2>Characterisation of plasmids coding for the restriction endonuclease EcoRI.
<3>Mol. Gen. Genet.
<4>143
<5>319-325
<6>1976
<7>The properties of two plasmids coding for the CcoRI restriction and modification enzymes are
described. Both plasmids are non
auto-transferring (NTP) but can be mobilised by transfer factors. Strains
carrying NTP13 produce colicin E1 and the EcoRI enzymes. This plasmid has
a molecular weight of 6 X 10(6) daltons and is present as approximately 12
copies per chromosome. The second plasmid, NTP14, was detected after
mobilisation of the EcoRI plasmid with the R factor RI-19. NTP14 codes for
ampicillin resistance, synthesis of the EcoRI enzymes and colicin E1. The
molecular weight of NTP14 is 10.7 X 10(6) daltons and there are about 14
copies per chromosome. DNA-DNA reassociation experiments were performed to
determine the interrelationships of NTP13, NTP14, ColE1 and the R factor
R1-19. NTP13 and NTP14 continue to replicate when cellular protein
synthesis is inhibited by the addition of chloramphenicol.

<>

<1>Smith, H.S.
<2>The restriction of T2 bacteriophage by Escherichia coli, strain W.
<3>Diss. Abstr.
<4>29
<5>9292B
<6>1968
<7>This thesis presents studies on the restriction of T2 bacteriophage infection
by Escherichia coli strain W, a bacterial strain lysogenic for a prophage which
is related to bacteriophage P2.

<>

<1>Smith, J., Berg, J.M., Chandrasegaran, S.
<2>A detailed study of the substrate specificity of a chimeric restriction enzyme.
<3>Nucleic Acids Res.
<4>27
<5>674-681
<6>1999
<7>Recently, the crystal structure of the designed zinc finger protein, deltaQNK, bound to a
preferred DNA sequence was reported.  We have converted deltaQNK into a novel site-specific
endonuclease by linking it to the FokI cleavage domain (FN).  The substrate specificity and
DNA cleavage properties of the resulting chimeric restriction enzyme (deltaQNK-FN) were
investigated, and the binding affinities of deltaQNK and deltaQNK-FN for various DNA
substrates were determined.  Substrates that are bound by deltaQNK with high affinity are the
same as those that are cleaved efficiently by deltaQNK-FN.  The binding of deltaQNK-FN to each
substrate was ~2-fold weaker than that for deltaQNK.  Thus, the fusion of the FokI cleavage
domain to the zinc finger motif does not change the DNA sequence specificity of the zinc
finger protein and does not change its binding affinity significantly.

<>

<1>Smith, J., Bibikova, M., Whitby, F.G., Reddy, A.R., Chandrasegaran, S., Carroll, D.
<2>Requirements for double-strand cleavage by chimeric restriction enzymes with zinc finger DNA-recognition domains.
<3>Nucleic Acids Res.
<4>28
<5>3361-3369
<6>2000
<7>This study concerns chimeric restriction enzymes that are hybrids between a zinc finger
DNA-binding domain and the non-specific DNA-cleavage domain from the natural restriction
enzyme FokI. Because of the flexibility of DNA recognition by zinc fingers, these enzymes are
potential tools for cleaving DNA at arbitrarily selected sequences. Efficient double-strand
cleavage by the chimeric nucleases requires two binding sites in close proximity. When cuts
were mapped on the DNA strands, it was found that they occur in pairs separated by
approximately 4 bp with a 5' overhang, as for native FokI. Furthermore, amino acid changes
in the dimer interface of the cleavage domain abolished activity. These results reflect a
requirement for dimerization of the cleavage domain. The dependence of cleavage efficiency on
the distance between two inverted binding sites was determined and both upper and lower limits
were defined. Two different zinc finger combinations binding to non-identical sites also
supported specific cleavage. Molecular modeling was employed to gain insight into the precise
location of the cut sites. These results define requirements for effective targets of chimeric
nucleases and will guide the design of novel specificities for directed DNA cleavage in vitro
and in vivo.

<>

<1>Smith, J., Grizot, S., Arnould, S., Duclert, A., Epinat, J.C., Prieto, P.C., Redondo, P., Blanco, F.J., Bravo, J., Montoya, G., Paques, F., Duchateau, P.
<2>A combinatorial approach to create artificial homing endonucleases cleaving chosen sequences.
<3>Nucleic Acids Res.
<4>34
<5>e149
<6>2006
<7>Meganucleases, or homing endonucleases (HEs) are sequence-specific endonucleases with large
(>14 bp) cleavage sites that can be used to
induce efficient homologous gene targeting in cultured cells and plants.
These findings have opened novel perspectives for genome engineering in a
wide range of fields, including gene therapy. However, the number of
identified HEs does not match the diversity of genomic sequences, and the
probability of finding a homing site in a chosen gene is extremely low.
Therefore, the design of artificial endonucleases with chosen
specificities is under intense investigation. In this report, we describe
the first artificial HEs whose specificity has been entirely redesigned to
cleave a naturally occurring sequence. First, hundreds of novel
endonucleases with locally altered substrate specificity were derived from
I-CreI, a Chlamydomonas reinhardti protein belonging to the LAGLIDADG
family of HEs. Second, distinct DNA-binding subdomains were identified
within the protein. Third, we used these findings to assemble four sets of
mutations into heterodimeric endonucleases cleaving a model target or a
sequence from the human RAG1 gene. These results demonstrate that the
plasticity of LAGLIDADG endonucleases allows extensive engineering, and
provide a general method to create novel endonucleases with tailored
specificities.

<>

<1>Smith, J.D., Arber, W., Kuhnlein, U.
<2>Host specificity of DNA by Escherichia coli. XIV.  The role of nucleotide methylation in in vivo B-specific modification.
<3>J. Mol. Biol.
<4>63
<5>1-8
<6>1972
<7>N-6-methyladenine is the only methylated base detected in bacteriophage fd DNA.
Its frequency is (1) host dependent: fd grown on a strain providing B-specific
modification to DNA carries twice as many methyl moieties in its DNA as phage
grown on a non-modifying host; and (2) dependent on the number of sites with
affinity for B-specific restriction: the DNA of a B-restriction insensitive
double mutant of fd is only half as much methylated after growth on B as its
wild type parent.  These results permit one to correlate methylation of
specifically located adenines with B-specific modification.  Quantitative
measurements suggest that one N-6-methyladenine is carried per B-host
specificity site on the modified, single stranded fd DNA.  Wild-type fd has two
such sites.  The biological function assigned to methylation brought about by
B-specific modification is the protection of the DNA from its destruction by
restriction endonuclese R.B.

<>

<1>Smith, J.L., Goldberg, J.M., Grossman, A.D.
<2>Complete Genome Sequences of Bacillus subtilis subsp. subtilis Laboratory Strains JH642 (AG174) and AG1839.
<3>Genome Announcements
<4>2
<5>e00663-14
<6>2014
<7>The Gram-positive bacterium Bacillus subtilis is widely used for studies of cellular and
molecular processes. We announce the complete genomic sequences of
strain AG174, our stock of the commonly used strain JH642, and strain AG1839, a
derivative that contains a mutation in the replication initiation gene dnaB and a
linked Tn917.

<>

<1>Smith, L.A., Chirikjian, J.G.
<2>Purification and characterization of the sequence-specific endonuclease BamHI.
<3>J. Biol. Chem.
<4>254
<5>1003-1006
<6>1979
<7>The specific endonuclease BamHI from Bacillus amyloliquefaciens (RUB 500) has
been purified to apparent homogeneity.  Two active forms of the enzyme
corresponding to the dimeric and tetrameric forms have been isolated.  On
sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the enzyme
dissociated into Mr = 22,000 _+ 500 subunits.  BamHI has a broad pH optimum on
the alkaline side and requires Mg2+ which can be partially replaced by Mn2+.
The enzyme catalysis appears to be governed by a two-step mechanism.

<>

<1>Smith, M.A., Mernagh, D.R., Kneale, G.G.
<2>Expression and characterization of the N-terminal fragment of the hsdS subunit of M.EcoR124I.
<3>Biol. Chem.
<4>379
<5>505-509
<6>1998
<7>The type IC modification methyltransferase M.EcoR124I is a trimeric enzyme of 162 kDa
consisting of two copies of the modification subunit, HsdM, and a single DNA specificity
subunit, HsdS.  Studies to date have been largely restricted to the HsdM subunit or the intact
methyltransferase, since the HsdS subunit is insoluble when expressed independently of HsdM.
Using PCR, we have cloned and expressed 13 fragments of the gene for the hsdS subunit,
including the sequences encoding each of the variable and conserved domains and various
combinations of these.  Only two of these fragments were found to be soluble, an 8.6 kDa
fragment (S11) comprising the central conserved domain and a 25 kDa N-terminal fragment (S3)
containing the N-terminal variable domain and the central conserved domain.  Analysis of the
larger of these fragments by gel retardation shows that the protein binds DNA in the presence
of HsdM at a subunit stoichiometry of 1:1.  Gel filtration and CD spectroscopy indicate that
the protein is monomeric and predominantly alpha-helical.

<>

<1>Smith, M.A., Read, C.M., Kneale, G.G.
<2>Domain structure and subunit interactions in the type I DNA methyltransferase M.EcoR124I.
<3>J. Mol. Biol.
<4>314
<5>41-50
<6>2001
<7>The type IC DNA methyltransferase M.EcoR124I is a trimeric enzyme of 162 kDa consisting of two
modification subunits, HsdM, and a single specificity subunit, HsdS. Studies have been largely
restricted to the HsdM subunit or to the intact methyltransferase since the HsdS subunit is
insoluble when over-expressed independently of HsdM. Two soluble fragments of the HsdS subunit
have been cloned, expressed and purified; a 25 kDa N-terminal fragment (S3) comprising the
N-terminal target recognition domain together with the central conserved domain, and an 8.6
kDa fragment (S11) comprising the central conserved domain alone.  Analytical
ultracentrifugation shows that the S3 subunit exists principally as a dimer of 50 kDa. Gel
retardation and competition assays show that both S3 and S11 are able to bind to HsdM, each
with a subunit stoichiometry of 1:1. The tetrameric complex (S3/HsdM)(2) is required for
effective DNA binding. Cooperative binding is observed and at low enzyme concentration, the
multisubunit complex dissociates, leading to a loss of DNA binding activity. The (S3/HsdM)(2)
complex is able to bind to both the EcoR124I DNA recognition sequence GAAN(6)RTCG and a
symmetrical DNA sequence GAAN(7)TTC, but has a 30-fold higher affinity binding for the latter
DNA sequence. Exonuclease III footprinting of the (S3/HsdM)(2) -DNA complex indicates that 29
nucleotides are protected on each strand, corresponding to a region 8 bp on both the 3' and
5' sides of the recognition sequence bound by the (S3/HsdM)(2) complex.

<>

<1>Smith, M.D., Longo, M., Gerard, G.F., Chatterjee, D.K.
<2>Cloning and characterization of genes for the PvuI restriction and modification system.
<3>Nucleic Acids Res.
<4>20
<5>5743-5747
<6>1992
<7>The genes encoding the endonuclease and the methylase of the PvuI restriction and modification
system were cloned in E.coli and characterized. The genes were adjacent in tandem orientation
spanning a distance of 2200 bases. The PvuI endonuclease was a single polypeptide with a
calculated molecular weight of 27,950 daltons. The endonuclease was easily detectable when the
gene was expressed from its endogenous promotor and present on a low copy plasmid, but
expression was considerably enhanced when the endonuclease gene was placed under the control
of a strong promotor on a high copy plasmid. The methylase did not completely protect plasmid
DNA from R.PvuI digeston unitl the methylase gene was placed under lac promotor control in a
multicopy plasmid. In the absence of the M.PvuI methylase, expression of the R.PvuI
endonuclease from the lac promotor on a multicopy plasmid was not lethal to wild type E.coli,
but was lethal in a temperature-sensitive ligase mutant at the non-permissive temperature.
Moreover, induction of the R.PvuI endonuclease under lambda pL promotor control resulted in
complete digestion of the E. coli chromosome by R.PvuI.

<>

<1>Smith, M.D., Schmidt, B.J., Longo, M.C., Chatterjee, D.K.
<2>Cloning and expression of AluI restriction endonuclease.
<3>US Patent Office
<4>US 5334526
<5>
<6>1994
<7>The present invention is directed to recombinant hosts which contain and express the AluI
Type-II restriction endonuclease gene. The present invention is also directed to vectors or
DNA molecules which contain the gene, and to methods of producing the AluI enzyme. One source
of the enzyme is Arthrobacter luteus, although other microorganisms may be used to isolate
restriction endonuclease isoschizomers of the invention.

<>

<1>Smith, M.J., Jeddeloh, J.A.
<2>DNA methylation in lysogens of pathogenic Burkholderia spp. requires prophage induction and is restricted to excised phage DNA.
<3>J. Bacteriol.
<4>187
<5>1196-1200
<6>2005
<7>Burkholderia mallei-specific phage  E125 encodes DNA methyltransferases in both the lysogenic
and replication modules within its genome. Characterization of DNA methylation in recombinant
systems, specifically in  E125 lysogenic strains of B. mallei and Burkholderia thailandensis,
revealed that, upon induction, cytosine methylation was targeted specifically to the phage
episome but not the phage provirus or the host chromosome.

<>

<1>Smith, R.M., Diffin, F.M., Savery, N.J., Josephsen, J., Szczelkun, M.D.
<2>DNA cleavage and methylation specificity of the single polypeptide restriction-modification enzyme LlaGI.
<3>Nucleic Acids Res.
<4>37
<5>7206-7218
<6>2009
<7>LlaGI is a single polypeptide restriction-modification enzyme encoded on the
naturally-occurring plasmid pEW104 isolated from Lactococcus lactis
ssp. cremoris W10. Bioinformatics analysis suggests that the enzyme
contains domains characteristic of an mrr endonuclease, a superfamily 2
DNA helicase and a gamma-family adenine methyltransferase. LlaGI was
expressed and purified from a recombinant clone and its properties
characterised. An asymmetric recognition sequence was identified,
5'-CTnGAyG-3' (where n is A, G, C or T and y is C or T). Methylation of
the recognition site occurred on only one strand (the non-degenerate dA
residue of 5'-CrTCnAG-3' being methylated at the N6 position). Double
strand DNA breaks at distant, random sites were only observed when two
head-to-head oriented, unmethylated copies of the site were present;
single sites or pairs in tail-to-tail or head-to-tail repeat only
supported a DNA nicking activity. dsDNA nuclease activity was dependent
upon the presence of ATP or dATP. Our results are consistent with a
directional long-range communication mechanism that is necessitated by the
partial site methylation. In the accompanying manuscript [Smith et al.
(2009) The single polypeptide restriction-modification enzyme LlaGI is a
self-contained molecular motor that translocates DNA loops], we
demonstrate that this communication is via 1-dimensional DNA loop
translocation. On the basis of this data and that in the third
accompanying manuscript [Smith et al. (2009) An Mrr-family nuclease motif
in the single polypeptide restriction-modification enzyme LlaGI], we
propose that LlaGI is the prototype of a new sub-classification of
Restriction-Modification enzymes, named Type I SP (for Single
Polypeptide).

<>

<1>Smith, R.M., Jacklin, A.J., Marshall, J.J., Sobott, F., Halford, S.E.
<2>Organization of the BcgI restriction-modification protein for the transfer of one methyl group to DNA.
<3>Nucleic Acids Res.
<4>41
<5>405-417
<6>2012
<7>The Type IIB restriction-modification protein BcgI contains A and B subunits in a 2:1 ratio: A
has the active sites for both endonuclease and methyltransferase
functions while B recognizes the DNA. Like almost all Type IIB systems, BcgI
needs two unmethylated sites for nuclease activity; it cuts both sites upstream
and downstream of the recognition sequence, hydrolyzing eight phosphodiester
bonds in a single synaptic complex. This complex may incorporate four A(2)B
protomers to give the eight catalytic centres (one per A subunit) needed to cut
all eight bonds. The BcgI recognition sequence contains one adenine in each
strand that can be N(6)-methylated. Although most DNA methyltransferases operate
at both unmethylated and hemi-methylated sites, BcgI methyltransferase is only
effective at hemi-methylated sites, where the nuclease component is inactive.
Unlike the nuclease, the methyltransferase acts at solitary sites, functioning
catalytically rather than stoichiometrically. Though it transfers one methyl
group at a time, presumably through a single A subunit, BcgI methyltransferase
can be activated by adding extra A subunits, either individually or as part of
A(2)B protomers, which indicates that it requires an assembly containing at least
two A(2)B units.

<>

<1>Smith, R.M., Josephsen, J., Szczelkun, M.D.
<2>An Mrr-family nuclease motif in the single polypeptide restriction-modification enzyme LlaGI.
<3>Nucleic Acids Res.
<4>37
<5>7231-7238
<6>2009
<7>Bioinformatic analysis of the putative nuclease domain of the single polypeptide
restriction-modification enzyme LlaGI reveals amino acid
motifs characteristic of the Escherichia coli methylated DNA-specific Mrr
endonuclease. Using mutagenesis, we examined the role of the conserved
residues in both DNA translocation and cleavage. Mutations in those
residues predicted to play a role in DNA hydrolysis produced enzymes that
could translocate on DNA but were either unable to cleave the
polynucleotide track or had reduced nuclease activity. Cleavage by LlaGI
is not targeted to methylated DNA, suggesting that the conserved motifs in
the Mrr domain are a conventional sub-family of the PD-(D/E)XK superfamily
of DNA nucleases.

<>

<1>Smith, R.M., Josephsen, J., Szczelkun, M.D.
<2>The single polypeptide restriction-modification enzyme LlaGI is a self-contained molecular motor that translocates DNA loops.
<3>Nucleic Acids Res.
<4>37
<5>7219-7230
<6>2009
<7>To cleave DNA, the single polypeptide restriction-modification enzyme LlaGI must communicate
between a pair of indirectly repeated recognition
sites. We demonstrate that this communication occurs by a 1-dimensional
route, namely unidirectional dsDNA loop translocation rightward of the
specific recognition sequence 5'-CTnGAyG-3' as written (where n is either
A, G, C or T and y is either C or T). Motion across thousands of base
pairs is catalysed by the helicase domain and requires the hydrolysis of
1.5-2 ATP per base pair. DNA loop extrusion is accompanied by changes in
DNA twist consistent with the motor following the helical pitch of the
polynucleotide track. LlaGI is therefore an example of a polypeptide that
is a completely self-contained, multi-functional molecular machine.

<>

<1>Smith, R.M., Marshall, J.J., Jacklin, A.J., Retter, S.E., Halford, S.E., Sobott, F.
<2>Organization of the BcgI restriction-modification protein for the cleavage of eight phosphodiester bonds in DNA.
<3>Nucleic Acids Res.
<4>41
<5>391-404
<6>2012
<7>Type IIB restriction-modification systems, such as BcgI, feature a single protein with both
endonuclease and methyltransferase activities. Type IIB nucleases
require two recognition sites and cut both strands on both sides of their
unmodified sites. BcgI cuts all eight target phosphodiester bonds before
dissociation. The BcgI protein contains A and B polypeptides in a 2:1 ratio: A
has one catalytic centre for each activity; B recognizes the DNA. We show here
that BcgI is organized as A(2)B protomers, with B at its centre, but that these
protomers self-associate to assemblies containing several A(2)B units. Moreover,
like the well known FokI nuclease, BcgI bound to its site has to recruit
additional protomers before it can cut DNA. DNA-bound BcgI can alternatively be
activated by excess A subunits, much like the activation of FokI by its catalytic
domain. Eight A subunits, each with one centre for nuclease activity, are
presumably needed to cut the eight bonds cleaved by BcgI. Its nuclease reaction
may thus involve two A(2)B units, each bound to a recognition site, with two more
A(2)B units bridging the complexes by protein-protein interactions between the
nuclease domains.

<>

<1>Smith, R.M., Pernstich, C., Halford, S.E.
<2>TstI, a Type II restriction-modification protein with DNA recognition, cleavage and methylation functions in a single polypeptide.
<3>Nucleic Acids Res.
<4>42
<5>5809-5822
<6>2014
<7>Type II restriction-modification systems cleave and methylate DNA at specific sequences.
However, the Type IIB systems look more like Type I than conventional Type II schemes as they
employ the same protein for both restriction and modification and for DNA recognition. Several
Type IIB proteins, including the archetype BcgI, are assemblies of two polypeptides: one with
endonuclease and methyltransferase roles, another for DNA recognition. Conversely, some IIB
proteins express all three functions from separate segments of a single polypeptide. This
study analysed one such single-chain protein, TstI. Comparison with BcgI showed that the one-
and the two-polypeptide systems differ markedly. Unlike the heterologous assembly of BcgI,
TstI forms a homotetramer. The tetramer bridges two recognition sites before eventually
cutting the DNA in both strands on both sides of the sites, but at each site the first
double-strand break is made long before the second. In contrast, BcgI cuts all eight target
bonds at two sites in a single step. TstI also differs from BcgI in either methylating or
cleaving unmodified sites at similar rates. The site may thus be modified before TstI can make
the second double-strand break. TstI MTase acts best at hemi-methylated sites.

<>

<1>Smith, S.S.
<2>Biological implications of the mechanism of action of human DNA (cytosine-5)methyltransferase.
<3>Prog. Nucleic Acid Res. Mol. Biol.
<4>49
<5>65-111
<6>1994
<7>I. Mechanism of action of the human DNA (cytosine-5)methyltransferase
A.  Sequence of catalytic events
B.  sp2-sp3 energetics and stereochemistry at C-6 and C-5 of cytosine
C.  Conformational change in the enzyme--DNA complex
D.  Potential for proton-mediated hydrolytic deamination
II.  Selectivity of human DNA methyltransferases
A.  De Novo methylation
B.  Methyl-directed methylation
C.  Structurally induced methylation
D.  The three-nucleotide recognition motif
E.  Enzyme--DNA interaction at the asymmetric DNA-binding site
III.  Biological implications of the mechanism
A.  Specificity of human DNA methylation
B.  Pattern formation as the key to the function of vertebrate DNA methylation
C.  Key elements of pattern formation are demonstrated by the phenomenon of concerted
modification
D.  Enzymology of pattern formation mechanisms
E.  Enzymology of disturbances in patterning produced by DNA damage
F.  Deamination at C-G dinucleotides
IV.  Conclusions
References

<>

<1>Smith, S.S.
<2>Construction of nucleoprotein based assemblies comprising addressable components for nanoscale assembly and nanoprocessors.
<3>International Patent Office
<4>WO 9605325
<5>
<6>1996
<7>A nucleoprotein based nanoprocessor is described.  The nanoprocessor
includes one or more chimeric fusion proteins linked to a DNA scaffold.  Both components
of the fusion protein are enzymes.

<>

<1>Smith, S.S.
<2>Construction of nucleoprotein based assemblies comprising addressable components for nanoscale assembly and nanoprocessors.
<3>International Patent Office
<4>WO 9605214
<5>
<6>1996
<7>A nucleoprotein based nanoprocessor is described.  The nanoprocessor
includes one or more chimeric fusion proteins linked to a DNA scaffold.  Both components
of the fusion protein are enzymes.

<>

<1>Smith, S.S.
<2>Stalling of DNA methyltransferase in chromosome stability and chromosome remodelling (Review).
<3>Int. J. Mol. Med.
<4>1
<5>147-156
<6>1998
<7>As a consequence of their mechanism of action, DNA (cytosine-5) methyltransferases from both
prokaryotes and eukaryotes necessarily recognize mispaired bases in unusual DNA structures as
catalytic transition-state analogs.  A review of the available data suggests that the enzymes
are designed to stall at these sites because they are unable to release substrates or products
that are fixed in a conformation resembling the transition state.  The enzymes can operate by
a two-step process in which they first methylate extrahelical cytosines satisfying their
recognition requirements and subsequently stall at the site of methylation.  On RNA and
DNA-RNA hybrids they may operate by a similar one-step process in which they stall at
transition-state analogs without methylating cytosine moieties.  These natural capacities
suggest that the enzymes may physically participate in stable nucleoprotein assemblies formed
as components of normal chromatin structure or as intermediates in the repair of unusual
structures.  The methyltransferases, themselves, may physically participate in chromosome
remodelling as part of a mechanism of inactivation or imprinting by stabilizing RNA-DNA
hybrids or RNA-RNA secondary structure involving cis-acting untranslated RNAs like the product
of the Xist gene.  Methyltransferase may physically participate in the repair of certain
unusual structures by serving as a nucleation point.  The affinity for secondary structure in
nucleic acids may account for the spreading of DNA methylation patterns.  Titration of host
methyltransferase by RNA-DNA hybrids and RNA secondary structure formed during retroviral
replication in certain tumorigenic retroviruses, like MMTV, may account for global
hypomethylation observed in retrovirally transformed cells.  In a similar fashion, titration
of methyltransferase by secondary structures associated with chromosome instability may
account for global hypomethylation observed in association with local hypermethylation in
tumorigenesis.

<>

<1>Smith, S.S.
<2>Gilbert's conjecture: The search for DNA (cytosine-5) demethylases and the emergence of new functions for eukaryotic DNA (cytosine-5) methyltransferases.
<3>J. Mol. Biol.
<4>302
<5>1-7
<6>2000
<7>In 1985 Walter Gilbert challenged members of the DNA methylation community assembled at a
National Institutes of Health meeting organized by Giulio Cantoni and Ahron Razin with the
following words: "The most exciting aspect about the methyl groups on DNA is the thought that
they might provide a locally inherited change in a DNA structure. However, for that to be
interesting, those changes have to be different in different cells. Furthermore, the
alterations in methylation have to be freely imposable and have to be maintained. It is not
yet clear that all these properties are true. So I don't think one will find that methylation
ever is one of the primary, top-level controls on gene expression." In essence, Gilbert's
conjecture, that DNA methylation is not one of the top-level controls on gene expression,
assumes that evidence in favor of both of its testable propositions will not be obtained.
Evidence for the first proposition, that alterations in methylation status associated with
gene-expression states have to be maintained, was already available in 1985 and has been
strengthened by a number of very recent experiments. However, the extensive effort to obtain
evidence for the second proposition, that alterations in methylation status be freely
imposable, has not been successful in its original intent. The effort has, on the other hand,
resulted in the emergence of new functions for 5-methylcytosine and the cytosine
methyltransferases in eukaryotic DNA repair, recombination and chromosome stability.

<>

<1>Smith, S.S.
<2>Conformation space and the emergence of new functions for eukaryotic DNA (cytosine-5) methyltransferases.
<3>Int. J. Mol. Med.
<4>6
<5>S11
<6>2000
<7>The existence of a link correlating CpG methylation with gene silencing as seen in mammalian
gene imprinting, X-chromosome inactivation, and the repression of viral genomes and L1
elements now seems quite plausible.  The evidence suggests that methylation in certain regions
of a gene can provide one of the signals necessary for the regional assembly of
heterochromatin.  On the other hand, work with histone deacetylase inhibitors suggests that
methylation is not one of the primary controls on transcription because reactivation does not
require demethylation.  DNA (cytosine-5)methyltransferases, the enzymes that apply methylation
patterns are known to show a strong preference for DNA sequences from regions with high
conformation space (i.e. sequences capable of adopting multiple secondary structures).  Such
sequences are present at sites that are often prone to clastic mutation (e.g. triplet repeat
sequences that expand in fragile X-linked mental retardation).  Moreover, lesions in
methyltransferase genes have been shown to promote clastic mutation.  These findings suggest
that methyltransferases play an additional role in the recognition and repair of clastogenic
damage in organisms that must incorporate sequences with high conformation spaces into their
genomes.  Such organisms may use stalled methyltransferase proteins as a cue in the repair of
inactive chromatin domains containing high conformation-space sequences.  The preference for
high conformation-space sequences exhibited by these enzymes extends to RNA.  RNA:DNA hybrids
can bind to methyltransferase but are not methylated on either strand.  Short RNA sequences
from the Xist gene and the 451nt human telomerase RNA (hTR) adopt secondary structures that
bind to the human placental enzyme and inhibit its action.  Given the widely observed
elevation of hTR levels in human cancers, and the mechanistic similarities between dyskerin, a
putative pseudouracil synthase, and the DNA methyltransferase, the data raise the possibility
that a DNA methyltransferase-hTR complex may form during tumorigenesis, protecting hTR from
degradation and promoting chromosome instability by blocking the normal action of DNA
methyltransferase.  These findings suggest that the elevation of hTR levels in prostate cancer
cells that we have observed in expressed prostatic secretion may be partially responsible for
the disruptions in methylation patterning and chromosomal instability that have been noted in
these tumor specimens.

<>

<1>Smith, S.S.
<2>Inhibition of human DNA (cytosine-5) methyltransferase by the 451nt RNA component of human telomerase.
<3>Mol. Biol. Cell
<4>11
<5>438a-439a
<6>2000
<7>A link between telomerase expression and chromosome stability has been recognized for some
time, and links between chromosome instability and cytosine methylation have recently been
firmly established.  Most research indicates that the local hypermethylation and global
hypomethylation associated with tumorigenesis are markers of chromosome instability, which in
turn is a hallmark of tumor progression.  The enzymes that apply methylation patterns to DNA
[DNA (cytosine-5) methyltransferases], are known to show a strong preference for DNA sequences
capable of adopting multiple secondary structures.  Sequences of this type are present at
sites that are often prone to clastic mutation (e.g. triplet repeat sequences that expand and
become methylated in fragile X-linked mental retardation).  These findings, coupled with
reports that methyltransferase mutations induce clastic mutation, suggest that
methyltransferases may play a role in the recognition and repair of clastogenic damage in
organisms that must incorporate sequences prone to secondary structure formation into their
genomes.  The preference for sequences that adopt secondary structure exhibited by these
enzymes, extends to RNA.  RNA:DNA hybrids can bind to methyltransferase but are not methylated
on either strand.  The 451 nt human temomerase RNA (hTR) adopts secondary structures that bind
to the human placental enzyme and inhibit its action with an inhibition constant in the nM
range.  Given the widely observed elevation of h TR levels in human cancers, and the
mechanistic similarities between dyskerin, (a putative pseudouracil synthase involved in hTR
processing), and the DNA methyltransferase, these data raise the possibility that DNA
methyltransferase may form a complex with hTR-RNA during tumorigenesis, protecting hTRRNA from
degradation and promoting chromosome instability by blocking the normal action of DNA
methyltransferase.

<>

<1>Smith, S.S., Baker, D.J.
<2>Stalling of human methyltransferase at single-strand conformers from the Huntington's locus.
<3>Biochem. Biophys. Res. Commun.
<4>234
<5>73-78
<6>1997
<7>We describe evidence for a sequence of events in which the human DNA (cytosine-5)
methyltransferase first methylates spontaneous single-stranded conformers and then stalls at
the methylated site to produce a complex with the conformationally unusual DNA.  This property
of the enzyme is a result of its ability to respond to a general loss of symmetry at its CG
recognition site.  The data suggest that DNA methyltransferase, itself, may physically
participate in biological processes that distinguish between DNA that is in the normal
Watson-Crick paired conformation and DNA that is conformationally unusual (e.g. a hairpin loop
or misassembled replication intermediate).  The in vitro methylation of spontaneous SSCs from
the Huntington's locus illustrates the phenomenon.

<>

<1>Smith, S.S., Crocitto, L.
<2>DNA methylation in eukaryotic chromosome stability revisited: DNA methyltransferase in the management of DNA conformation space.
<3>Mol. Carcinog.
<4>26
<5>1-9
<6>1999
<7>A previous working hypothesis on the role of DNA methylation in eukaryotic stability presented
enzymological data suggesting the DNA methylation has evolved as a biological response to the
formation of unusual DNA structures.  That evidence suggested that human DNA methyltransferase
is uniquely suited to participate in a repair system that functions to suppress unusual DNA
structures.  The evolution of this repair system was suggested to be a prerequisite for the
incorporation of dynamic sequences into the genome, because such sequences are particularly
prone to the formation of unusual DNA structures that promote clastic mutagenesis.  In the
intervening period, considerable evidence has accumulated in support of this proposal.  Here,
we extend the previous working hypothesis in light of the current experimental data to give a
more detailed picture of the role of DNA methylation in eukaryotic chromosome stability.

<>

<1>Smith, S.S., Hardy, T.A., Baker, D.J.
<2>Human DNA (cytosine-5)methyltransferase selectively methylates duplex DNA containing mispairs.
<3>Nucleic Acids Res.
<4>15
<5>6899-6916
<6>1987
<7>The presence of the C-C mispair in a defined duplex oligodeoxynucleotide enhanced its capacity
to serve as a substrate for highly purified human DNA methyltransferase. Analysis of tritiated
reaction products showed that the C-C mispair acted as a "methylation acceptor" in that it was
itself rapidly methylated. The m5C-G base pair also enhanced the capacity of the
oligodeoxynucleotide to serve as a substrate for the enzyme. However, this complementary base
pair was found to act as a "methylation director". That is, the presence of the m5C in one
strand induced the enzyme to rapidly methylate at the cytosine residue on the opposite strand
in an adjacent C-G base pair.

<>

<1>Smith, S.S., Kan, J.L.C., Baker, D.J., Kaplan, B.E., Dembek, P.
<2>Recognition of unusual DNA structures by human DNA (cytosine-5) methyltransferase.
<3>J. Mol. Biol.
<4>217
<5>39-51
<6>1991
<7>The symmetry of the responses of the human DNA (cytosine-5) methyltransferase
to alternative placements of 5-methylcytosine in model oligodeoxynucleotide
duplexes containing unusual structures has been examined.  The results of these
experiments more clearly define the DNA recognition specificity of the enzyme.
A simple three-nucleotide recognition motif within the CG dinucleotide pair can
be identified in each enzymatically methylated duplex.  The data can be
summarized by numbering the four nucleotides in the dinucleotide pair thus: 1
4 / 2   3  With reference to this numbering scheme, position 1 can be occupied
by cytosine or 5-methylcytosine; position 2 can be occupied by guanosine or
inosine; position 3, the site of enzymatic methylation, can be occupied only by
cytosine; and position 4 can be occupied by guanosine, inosine,
O6-methylguanosine, cytosine, adenosine, an abasic site, or the 3' hydroxyl
group at the end of a gapped molecule.  Replacing the guanosine normally found
at position 4 with any of the moieties introduces unusual (non-Watson-Crick)
pairing at position 3 and generally enhances methylation of the cytosine at
that site.  The exceptional facility of the enzyme in actively methylating
unusual DNA structures suggests that the evolution of the DNA
methyltransferase, and perhaps DNA methylation itself may be linked to the
biological occurrence of unusual DNA structures.

<>

<1>Smith, S.S., Kaplan, B.E.
<2>Mechanism based inhibitors of DNA methyltransferase.
<3>International Patent Office
<4>WO 9206985
<5>
<6>1992
<7>Method for producing double stranded oligonucleotide compounds by providing a self-priming DNA
fragment and extending said double stranded DNA with a DNA polymerase and with a combination
of dNTP's and 5-fdCTP, and compounds directed to same.

<>

<1>Smith, S.S., Kaplan, B.E.
<2>Mechanism based inhibitors of DNA methyltransferase.
<3>US Patent Office
<4>US 5503975
<5>
<6>1996
<7>Novel synthetic oligomers of unique three-dimensional structure comprising homologous and
heterologous blocks, some of which may self-associate are described.  Also disclosed is a
synthetic oligonucleotide that includes 5-fluoro-cytosine residues at positions corresponding
to methylation sites of DNA-cytosine methyltransferase and a method for preparing such a
molecule.

<>

<1>Smith, S.S., Kaplan, B.E., Sowers, L.C., Newman, E.M.
<2>Mechanism of human methyl-directed DNA methyltransferase and the fidelity of cytosine methylation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>4744-4748
<6>1992
<7>The properties of the methyl-directed DNA (cytosine-5)-methyltransferase (EC2.1.1.37) suggest
that it is the enzyme that maintains patterns of methylation in the human genome. Proposals
for the enzyme's mechanism of action suggest that 5-methyldeoxycytidine is produced from
deoxycytidine via a dihydrocytosine intermediate. We have used an oligodeoxynucleotide
containing 5-fluorodeoxycytidine as a suicide substrate to capture the enzyme and the
dihydrocytosine intermediate. Gel retardation experiments demonstrate the formation of the
expected covalent complex between duplex DNA containing 5-fluorodeoxycytidine and the human
enzyme. Formation of the complex was dependent upon the presence of the methyl donor
S-adenosylmethionine, suggesting that it comprises an enzyme-linked 5-substituted
dihydrocytosine moiety in DNA. Dihydrocytosine derivatives are extremely labile toward
hydrolytic deamination in aqueous solution. Because C-to-T transition mutations are especially
prevalent at CG sites in human DNA, we have used high-performance liquid chromatography to
search for thymidine that might be generated by hydrolysis during the methyl transfer
reaction. Despite the potential for deamination inherent in the formation of the intermediate,
the methyltransferase did not produce detectable amounts of thymidine. The data suggest that
the ability of the human methyltransferase to preserve genetic information when copying a
methylation pattern (i.e., its fidelity) is comparable to the ability of a mammalian DNA
polymerase to preserve genetic information when copying a DNA sequence. Thus the high
frequency of C-to-T transitions at CG sites in human DNA does not appear to be due to the
normal enzymatic maintenance of methylation patterns.

<>

<1>Smith, S.S., Kho, M.R., Baker, D.J.
<2>DNA methyltransferases in the recognition and repair of hairpin loops lacking precise Watson-Crick homology.
<3>Anticancer Res.
<4>15
<5>1671
<6>1995
<7>Sequences from codon-12 of the c-Ha-ras gene, the trinucleotide repeat region of the FMR-1
gene and the trinucleotide repeat region of the HuntingtonM-Us disease gene spontaneously form
hairpin loops under physiological conditions even though these loops lack precise Watson-Crick
homology.  The formation of loops of this type during replication may provide a new driving
force for genetic damage during carcinogenesis and the expression of genetic diseases.  The
exceptional capacity of the human methyltransferase to recognize these loops is a consequence
of its mechanism of action which requires that it unstack the cytosine ring from the DNA helix
as it activates the ring for methylation by nucleophilic attack at C6.  The rate of
methylation is enhanced by the capacity of mispairs (e.g. C.C., C.C+ or A+.C mispairs) in the
stems of such loops to serve as transition state analogs for the catalysis.  These findings
suggest that the mechanisms by which methyl groups and chromosomal abnormalities accumulate in
the c-Ha-ras region of chromosome 11 during carcinogenesis and at the FMR-1 locus during
repeat expansion involve structurally-induced de novo methylation at sites undergoing local
conformational change.  Models for the participation of methyltransferases in the recognition
and repair of looped structures suggest that they might serve to mark looped structures for
repair.  Cycles of loop formation and repair are expected to produce spreading of de novo
methylation in the vicinity of affected genes.  Asymmetrically methylated duplexes produced by
repair of the looped structures would be rapidly converted to symmetrically methylated
duplexes through the methyl-directed activity of the human methyltransferase.

<>

<1>Smith, S.S., Laayoun, A., Baker, D.J., Kho, M.R.
<2>Recognition of telomere-like structures from the human C-Ha-Ras gene and the trinucleotide repeat of the FMR-1 gene of fragile X by human DNA methyltransferase.
<3>FASEB J.
<4>9
<5>A1324
<6>1995
<7>Telomere-like hairprins from the c-Ha-ras gene and from the FMR-1 gene are exceptional
substrates for human DNA(cytosine-5)methyltransferases. This is a consequence of the mechanism
of action of these enzymes which requires that a group at the active site carry out
nucleophilic attack of an unstacked-cytosine ring in DNA. Thus the rate of methylation is
enhanced by the capacity of mispairs like C.C or C.C+ found on telomere-like loops formed in
C-rich DNA strands to serve as transition state analogs for the catalysis. These findings
suggest that the mechanisms by which methyl groups and chromosomal abnormalities accumulate at
the FMR-1 locus during repeat expansion in fragile X syndrome and in the c-Ha-ras region of
chromosome 11 during carcinogenesis may involve structurally-induced de novo methylation at
sites undergoing local conformational change. Possible roles for cytosine methylation in loop
repair and trinucleotide repeat expansion will be discussed.

<>

<1>Smith, S.S., Lingeman, R.G., Kaplan, B.E.
<2>Recognition of foldback DNA by the human DNA (Cytosine-5-)-methyltransferase.
<3>Biochemistry
<4>31
<5>850-854
<6>1992
<7>In order to specifiy the recognition requirements of the human DNA
(cytosine-5-)-methyltransferase, two isomeric 48mers were synthesized so as to
link a long block of DNA with a shorter complementary block of DNA through a
tether consisting of five thymidine residues.  These isomeric foldback
molecules, differing only in the location of the 5-methyldeoxycytosine, were
shown to be unimolecular, to contain a region of duplex DNA, and to contain a
region of single-stranded DNA.  When used as substrates for the DNA
methyltransferase, only one of the isomers was methylated.  A comparison of the
structures of the two isomers allows us to begin to define the potential sites
of interaction between the enzyme and the three nucleotides forming a
structural motif consisting of 5-methyldeoxycytosine, its base-paired
deoxyguanosine, and a deoxycytosine 5' to the paired deoxyguanosine.

<>

<1>Smith, T.
<2>Restriction enzyme searches using the word processing software:  LOCOSCRIPT.
<3>Biochemical Education
<4>16
<5>224-226
<6>1988
<7>One of the most important routine applications of computers in biochemistry and
molecular biology is the determination of restriction enzyme sites.  In our
laboratory, we routinely carry out restriction enzyme searches on the
University's Vax System, using the Staden Molecular Biology software.
Dedicated molecular biology software, however, can be expensive, and is
frequently difficult to obtain for the computer system which happens to be
available, while for those involved in introducing a genetic
engineering/biotechnology component into the life science courses offered by
colleges of further education and schools, the purchase of software may prove a
major hurdle.  Even when the systems and software are available, competition
from colleagues for terminals can mean constant frustration and delay to the
biochemist wanting to do a simple restriction enzyme search which is well
within the capacity of the cheapest PCW.  We now describe a rapid and reliable
method of performing restriction enzyme searches and other molecular biology
functions using the word processing software LOCOSCRIPT included with the
Amstrad PCWQ 8256.  At a total cost for the complete system - computer,
monitor, dot matrix printer and LOCOSCRIPT - of 299 Pounds plus VAT, this
brings restriction enzyme searches and similar functions within the range of
all educational institutes and research labs, and even of the individual
scientist or teacher, many of whom already possess and use the system as a word
processor.

<>

<1>Smith, T.J.
<2>Aurintricarboxylic acid inhibition of restriction endonuclease is not recognition site-specific.
<3>FASEB J.
<4>2
<5>A589
<6>1988
<7>Aurintricarboxylic acid (ATA) has been previously shown to inhibit nucleases,
including restriction endonucleases.  The discovery of restriction
endonucleases which do not require guanine or cytosine bases within the
recognition sequence provides an opportunity to determine whether or not the
presence of these bases is required for inhibition by ATA.  In addition to the
endonuclease DraI, which does not contain guanine-cytosine (GC) base pairs
within the recognition sequence, two other restriction enzymes were included as
controls.  These endonucleases were HaeIII, which has a recognition site
without adenine-thymine (AT) base pairs, and HindIII, which contains both AT
and GC base pairs within the recognition sequence.  ATA was included in
restriction digests with the above endonucleases.  Following termination of the
reactions, DNA fragments were separated by agarose gel electrophoresis.  The
results indicate that inhibition by ATA does not require a specific base pair
within the recognition sequence.

<>

<1>Smith, T.J., Hill, K.K., Foley, B.T., Detter, J.C., Munk, A.C., Bruce, D.C., Doggett, N.A., Smith, L.A., Marks, J.D., Xie, G., Brettin, T.S.
<2>Analysis of the Neurotoxin Complex Genes in Clostridium botulinum A1-A4 and B1 Strains: BoNT/A3, /Ba4 and /B1 Clusters Are Located within  Plasmids.
<3>PLoS ONE
<4>2
<5>e1271
<6>2007
<7>BACKGROUND: Clostridium botulinum and related clostridial species express extremely potent
neurotoxins known as botulinum neurotoxins (BoNTs) that
cause long-lasting, potentially fatal intoxications in humans and other
mammals. The amino acid variation within the BoNT is used to categorize
the species into seven immunologically distinct BoNT serotypes (A-G) which
are further divided into subtypes. The BoNTs are located within two
generally conserved gene arrangements known as botulinum progenitor
complexes which encode toxin-associated proteins involved in toxin
stability and expression. METHODOLOGY/PRINCIPAL FINDINGS: Because serotype
A and B strains are responsible for the vast majority of human botulism
cases worldwide, the location, arrangement and sequences of genes from
eight different toxin complexes representing four different BoNT/A
subtypes (BoNT/A1-Ba4) and one BoNT/B1 strain were examined. The bivalent
Ba4 strain contained both the BoNT/A4 and BoNT/bvB toxin clusters. The
arrangements of the BoNT/A3 and BoNT/A4 subtypes differed from the BoNT/A1
strains and were similar to those of BoNT/A2. However, unlike the BoNT/A2
subtype, the toxin complex genes of BoNT/A3 and BoNT/A4 were found within
large plasmids and not within the chromosome. In the Ba4 strain, both BoNT
toxin clusters (A4 and bivalent B) were located within the same 270 kb
plasmid, separated by 97 kb. Complete genomic sequencing of the BoNT/B1
strain also revealed that its toxin complex genes were located within a
149 kb plasmid and the BoNT/A3 complex is within a 267 kb plasmid.
CONCLUSIONS/SIGNIFICANCE: Despite their size differences and the BoNT
genes they contain, the three plasmids containing these toxin cluster
genes share significant sequence identity. The presence of partial
insertion sequence (IS) elements, evidence of recombination/gene
duplication events, and the discovery of the BoNT/A3, BoNT/Ba4 and BoNT/B1
toxin complex genes within plasmids illustrate the different mechanisms by
which these genes move among diverse genetic backgrounds of C. botulinum.

<>

<1>Smits, T.H., Guerrero-Prieto, V.M., Hernandez-Escarcega, G., Blom, J., Goesmann, A., Rezzonico, F., Duffy, B., Stockwell, V.O.
<2>Whole-Genome Sequencing of Erwinia amylovora Strains from Mexico Detects Single Nucleotide Polymorphisms in rpsL Conferring Streptomycin Resistance and in the  avrRpt2 Effector Altering Host Interactions.
<3>Genome Announcements
<4>2
<5>e01229-13
<6>2014
<7>We report draft genome sequences of three Mexican Erwinia amylovora strains. A novel plasmid,
pEA78, was identified. Comparative genomics revealed an rpsL
chromosomal mutation conferring high-level streptomycin resistance in two
strains. In the effector gene avrRpt2, a single nucleotide polymorphism was
detected that overcomes fire blight disease resistance in Malus x robusta 5.

<>

<1>Smits, T.H., Rezzonico, F., Kamber, T., Goesmann, A., Ishimaru, C.A., Stockwell, V.O., Frey, J.E., Duffy, B.
<2>Genome Sequence of the Biocontrol Agent Pantoea vagans Strain C9-1.
<3>J. Bacteriol.
<4>192
<5>6486-6487
<6>2010
<7>Pantoea vagans is a Gram-negative enterobacterial plant epiphyte of a broad range of plants.
Here we report the 4.89-Mb genome sequence of P.
vagans strain C9-1 (formerly Pantoea agglomerans), which is commercially
registered for biological control of fire blight, a disease of pear and
apple trees caused by Erwinia amylovora.

<>

<1>Smoot, J.C., Barbian, K.D., Van Gompel, J.J., Smoot, L.M., Chaussee, M.S., Sylva, G.L., Sturdevant, D.E., Ricklefs, S.M., Porcella, S.F., Parkins, L.D., Beres, S.B., Campbell, D.S., Smith, T.M., Zhang, Q., Kapur, V., Daly, J.A., Veasy, L.G.
<2>Genome sequence and comparative microarray analysis of serotype M18 group A Streptococcus strains associated with acute rheumatic fever outbreaks.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>4668-4673
<6>2002
<7>Acute rheumatic fever (ARF), a sequelae of group A Streptococcus (GAS) infection, is the most
common cause of preventable childhood heart disease worldwide. The molecular basis of ARF and
the subsequent rheumatic heart disease are poorly understood. Serotype M18 GAS strains have
been associated for decades with ARF outbreaks in the U.S. As a first step toward gaining new
insight into ARF pathogenesis, we sequenced the genome of strain MGAS8232, a serotype M18
organism isolated from a patient with ARF. The genome is a circular chromosome of 1,895,017
bp, and it shares 1.7 Mb of closely related genetic material with strain SF370 (a sequenced
serotype M1 strain). Strain MGAS8232 has 178 ORFs absent in SF370. Phages, phage-like
elements, and insertion sequences are the major sources of variation between the genomes. The
genomes of strain MGAS8232 and SF370 encode many of the same proven or putative virulence
factors. Importantly, strain MGAS8232 has genes encoding many additional secreted proteins
involved in human-GAS interactions, including streptococcal pyrogenic exotoxin A (scarlet
fever toxin) and two uncharacterized pyrogenic exotoxin homologues, all phage-associated. DNA
microarray analysis of 36 serotype M18 strains from diverse localities showed that most
regions of variation were phages or phage-like elements. Two epidemics of ARF occurring 12
years apart in Salt Lake City, UT, were caused by serotype M18 strains that were genetically
identical, or nearly so. Our analysis provides a critical foundation for accelerated research
into ARF pathogenesis and a molecular framework to study the plasticity of GAS genomes.

<>

<1>Sneider, T.W.
<2>The 5'-cytosine in CCGG is methylated in two eukaryotic DNAs and MspI is sensitive to methylation at this site.
<3>Nucleic Acids Res.
<4>8
<5>3829-3840
<6>1980
<7>Novikoff rat hepatoma and bovine liver DNAs were digested with MspI or HpaII. Restriction
fragments were end-labeled using [a-32P]-dCTP and the Klenow fragment of E. coli DNA
polymerase I and then digested to 2'-deoxyribonucleoside-3'-monophosphates using micrococcal
nuclease and spleen phosphodiesterase. Mononucleotides were separated by two-dimensional thin
layer chromatography, localized by radioautography, and the [32P]-label quantitated by
scintillation spectrometry. This method, based on known specificities of MspI and HpaII, shows
that CCGG, CMGG, and MCGG (M refers to 5-methylcytosine) occur at frequencies of 89.6%, 1.4%,
and 9.0%, respectively, in the rat DNA and at 41.6%, 48.3%, and 10.0%, respectively, in the
ovine DNA. [32P] recovery in 3'-5-MedCMP from end-labeled MspI digests was negligible
compared to recovery from HpaII digests. Hence, MspI is sensitive to methylation at the 5'
cytosine in the sequence CCGG.

<>

<1>Snitkin, E.S., Zelazny, A.M., Thomas, P.J., Stock, F., Henderson, D.K., Palmore, T.N., Segre, J.A.
<2>Tracking a hospital outbreak of carbapenem-resistant Klebsiella pneumoniae with whole-genome sequencing.
<3>Sci. Transl. Med.
<4>4
<5>148RA116
<6>2012
<7>The Gram-negative bacteria Klebsiella pneumoniae is a major cause of nosocomial
infections, primarily among immunocompromised patients. The emergence of strains
resistant to carbapenems has left few treatment options, making infection
containment critical. In 2011, the U.S. National Institutes of Health Clinical
Center experienced an outbreak of carbapenem-resistant K. pneumoniae that
affected 18 patients, 11 of whom died. Whole-genome sequencing was performed on
K. pneumoniae isolates to gain insight into why the outbreak progressed despite
early implementation of infection control procedures. Integrated genomic and
epidemiological analysis traced the outbreak to three independent transmissions
from a single patient who was discharged 3 weeks before the next case became
clinically apparent. Additional genomic comparisons provided evidence for
unexpected transmission routes, with subsequent mining of epidemiological data
pointing to possible explanations for these transmissions. Our analysis
demonstrates that integration of genomic and epidemiological data can yield
actionable insights and facilitate the control of nosocomial transmission.

<>

<1>Snounou, G., Malcolm, A.D.B.
<2>Supercoiling and the mechanism of restriction endonucleases.
<3>Eur. J. Biochem.
<4>138
<5>275-280
<6>1984
<7>1. We have used topoisomerase I in the presence of netropsin and ethidium
bromide to generate DNA molecules of varying superhelical density.  2.
Digestion by endonuclease EcoRI is sensitive to supercoiling, being maximal for
the relaxed form.  Endonucleases AvaI and BamHI, by contrast, are relatively
unaffected.  3. The results are interpreted in terms of the base composition of
the DNA in the vicinity of these sites. dA + dT-rich regions are more
susceptible to deformation than are dG + dC-rich ones.  4. Analysis of the
rates of disappearance of linear molecules confirms a two-step mechanism for
EcoRI cleavage but suggests that BamHI and AvaI cleave both strands
simultaneously.

<>

<1>Snow, A.M., Sheardy, R.D.
<2>Locating cobalt-binding sites on DNA using restriction endonucleases.
<3>Methods Enzymol.
<4>340
<5>519-528
<6>2001
<7>The interactions of simple metal complexes with DNA have been widely studied.  The interaction
specificities of these molecules range from simple outside binding, such as observed with
[Co(NH3)6]3+, to covalent binding as observed with Pt(NH3)2Cl2, to reactions involving DNA
oxidative cleavage through Fenton or related chemistries.  One important feature of such
interactions, especially those leading to covalent binding, is the sequence specificity of the
reaction.  It has been well established that platination of DNA by Pt(NH3)2Cl2 is highly
sequence specific with a preferential reaction at -GG- sites, although reactions involving
-GA- and -GC- sites have also been noted.  Reaction at an isolated -GG- or -GA- site results
in an intrastrand cross-link composed of a pur(N7)-Pt-pur(N7) linkage, whereas the reaction at
a -GC- site results in an interstrand cross-link with the same type of linkage.  The sequence
specificity of these and related reactions is due to the accessibility to N7 of purines,
located in the major groove of the DNA, and to the nucleophilicity of N7, particularly that of
guanine bases.

<>

<1>Soares-Castro, P., Marques, D., Demyanchuk, S., Faustino, A., Santos, P.M.
<2>Draft Genome Sequences of Two Pseudomonas aeruginosa Clinical Isolates with Different Antibiotic Susceptibilities.
<3>J. Bacteriol.
<4>193
<5>5573
<6>2011
<7>Pseudomonas aeruginosa is a primary cause of opportunistic infections. We have sequenced and
annotated the genomes of two P. aeruginosa clinical
isolates evidencing different antibiotic susceptibilities. Registered
differences in the composition of their accessory genomes may provide
clues on P. aeruginosa strategies to thrive in different environments like
infection loci.

<>

<1>Soares-Castro, P., Santos, P.M.
<2>Towards the Description of the Genome Catalogue of Pseudomonas sp. Strain M1.
<3>Genome Announcements
<4>1
<5>e00146-12
<6>2013
<7>Pseudomonas sp. strain M1 is a soil isolate with remarkable biotechnological potential. The
genome of Pseudomonas sp. M1 was sequenced using both 454 and
Illumina technologies. A customized genome assembly pipeline was used to
reconstruct its genome sequence to a single scaffold.

<>

<1>Sobell, H.M.
<2>Symmetry in protein-nucleic acid interaction and its genetic implications.
<3>Adv. Genet.
<4>17
<5>411-490
<6>1973
<7>Symmetry principles are known to play a fundamental role in biological
organization, governing the assembly of macromolecular subunits into viruses,
membranes, oligomeric globular and fibrous proteins, and cellular organelles
(Crick and Watson, 1956; Caspar and Klug, 1962; Monod et al., 1965; for an
excellent review, see Engstrom and Strandberg, 1968).  Thus, for example, small
spherical viruses utilize icosahedral symmetry in their construction, identical
protein subunits being used to form large protein shells in which each subunit
has, as nearly as possible, the same local environment (Caspar and Klug, 1962;
Finch and Klug, 1966; Klug and Finch, 1968; Finch et al., 1970; for a review,
see Klug et al., 1966).

<>

<1>Sobotta, T., Pingoud, A., Jeltsch, A.
<2>Mapping of the functional regions of the EcoRV methyltransferase by random mutagenesis and screening for inactive mutants.
<3>Biol. Chem. Hoppe Seyler
<4>376
<5>S155
<6>1995
<7>To examine the structure and the mechanism of the EcoRV DNA methyltransferase several mutants
were produced by random mutagenesis and selection for inactive mutants.  Until now, 11
catalytically inactive single mutants were found: P8L; V20A; F115S; S121P; F139L; F172S;
C192R; D193G; E212G; W231R and F249V.  Two of the mutant proteins (F139L and F249V) were not
expressed in the cell, although the promotor sequence was intact, suggesting that these
proteins are misfolded and rapidly degraded.  For further characterization the other mutant
enzymes were purified by affinity-chromatography.  It turned out, that all proteins except
wild type and the W231R mutant were insoluble suggestive of structural alterations caused by
the mutations.  Whereas the purified wild type enzyme has a specific activity of 1.3 x 10^6
U/mg, W231R is catalytically inactive in vitro.  Currently, the DNA- and
S-adenosylmethionine-binding properties of the mutants are investigated.

<>

<1>Sobral, B.W.S., McClelland, M.
<2>Methyltransferases as tools to alter the specificity of restriction endonucleases.
<3>Methods Mol. Biol.
<4>12
<5>159-172
<6>1992
<7>Pulsed-field gel electrophoresis (PFGE) has allowed the resolution of very large DNA fragments
from any organism. To apply PFGE to practical problems such as genetic mapping and map-based
gene cloning, it is necessary to specifically generate large DNA fragments that can then be
separated by PFGE. Ideally, restriction enzymes would exist that could generate DNA fragments
of desired sizes. Other factors being equal, and supposing that DNA sequences were random,
then enzymes with larger target sequences should produce larger DNA fragments. In practice, no
restriction enzymes are known to have larger than 8-bp-long target sites and, of course, DNA
sequences are not random. These realities severely limit the observed sizes of DNA fragments
produced by restriction enzymes acting on genomic DNA particularly in the case of eukaryotic
genomes, which are large and complex. This chapter describes enzymatic strategies to generate
large DNA fragments and statistical tools that can aid researchers in choosing the restriction
enzymes that are most likely to generate large fragments in the genome in question, if a
sequence data base can be investigated.

<>

<1>Sogutcu, E., Emrence, Z., Arikan, M., Cakiris, A., Abaci, N., Oner, E.T., Ustek, D., Arga, K.Y.
<2>Draft Genome Sequence of Halomonas smyrnensis AAD6T.
<3>J. Bacteriol.
<4>194
<5>5690-5691
<6>2012
<7>Halomonas smyrnensis AAD6(T) is a Gram-negative, aerobic, exopolysaccharide-producing, and
moderately halophilic bacterium that produces
levan, a fructose homopolymer with many potential uses in various industries. We
report the draft genome sequence of H. smyrnensis AAD6(T), which will accelerate
research on the rational design and optimization of microbial levan production.

<>

<1>Sohail, A., Ives, C.L., Brooks, J.E.
<2>Purification and characterization of C.BamHI, a regulator of the BamHI restriction-modification system.
<3>Gene
<4>157
<5>227-228
<6>1995
<7>The bamHIC gene, controlling the BamHI restriction-modification (R-M) system can functionally
be replaced by providing pvuIIC or smaIC in trans.  C.BamHI, the protein product encoded by
bamHIC, has been purified and shown to bind a 345-bp DNA fragment within the BamHI R-M system.

<>

<1>Sohail, A., Lieb, M., Dar, M., Bhagwat, A.S.
<2>A gene required for very short patch repair in Escherichia coli is adjacent to the DNA cytosine methylase gene.
<3>J. Bacteriol.
<4>172
<5>4214-4221
<6>1990
<7>Deamination of 5-methylcytosine in DNA results in T/G mismatches. If unrepaired, these
mismatches can lead to C-to-T transition mutations. The very short patch (VSP) repair process
in Escherichia coli counteracts the mutagenic process by repairing the mismatches in favor of
the G-containing strand. Previously we have shown that a plasmid containing an 11-kilobase
fragment from the E. coli chromosome can complement a chromosomal mutation defective in both
cytosine methylation and VSP repair. We have now mapped the regions essential for the two
phenotypes. In the process, we have constructed plasmids that complement the chromosomal
mutation for methylation, but not for repair, and vice versa. The genes responsible for these
phenotypes have been identified by DNA sequence analysis. The gene essential for cytosine
methylation, dcm, is predicted to code for a 473-amino-acid protein and is not required for
VSP repair. It is similar to other DNA cytosine methylases and shares extensive sequence
similarity with its isoschizomer, EcoRII methylase. The segment of DNA essential for VSP
repair contains a gene that should code for a 156-amino-acid protein. This gene, named vsr, is
not essential for DNA methylation. Remarkably, the 5' end of this gene appears to overlap the
3' end of dcm. The two genes appear to be transcribed from a common promoter but are in
different translational registers. This gene arrangement may assure that Vsr is produced along
with Dcm and may minimize the mutagenic effects of cytosine methylation.

<>

<1>Sohail, A., Mushtaq, R., Khan, E., Maqbool, T., Riazuddin, S.
<2>Discovery of two new Type II restriction enzymes.
<3>Pak. J. Zool.
<4>19
<5>371-391
<6>1987
<7>Eight bacterial strains isolated from local environments have been screened for
the presence of new restriction enzymes judging from the analysis of 1.4%
agarose gel patterns of different DNA substrates.  Partially purified protein
extracts of two of these strains, namely Pseudomonas ovalis and Enterobacter
agglomerans exhibit the presence of Type II restriction endonucleases.  The new
enzymes have been highly purified by a combination of gel filtration, ion
exchange and affinity chromatography and are designated as PovI and EagI,
respectively.  The identity of the new enzymes have been further confirmed by
DNA double digest analysis with a variety of substrate DNAs which yielded no
extra fragment.  This enzyme has been renamed EagMI to avoid confusion with the
previous EagI.

<>

<1>Sohn, K.H., Jones, J.D., Studholme, D.J.
<2>Draft Genome Sequence of Pseudomonas syringae Pathovar Syringae Strain FF5, Causal Agent of Stem Tip Dieback Disease on Ornamental Pear.
<3>J. Bacteriol.
<4>194
<5>3733-3734
<6>2012
<7>Pseudomonas syringae FF5 causes stem tip dieback disease on ornamental pear (Pyrus
calleryana). Its genome encodes a complete type III secretion system
(T3SS) and HopAC1, HopM1, AvrE1, HopI1, HopAA1, HopJ1, HopAH2, HopAH1, HopAG1,
and HopAZ1. Lacking detectable homologues of other T3SS effectors, it may encode
novel, undiscovered effectors.

<>

<1>Sokolov, N.N.
<2>Current approaches to the search of new restriction endonucleases.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>3-7
<6>1989
<7>The main methodological approaches to the search for new restriction
endonucleases are reviewed.  These methods include obtaining acellular extracts
by ultrasonic disintegration of microbial cells, osmotic shock effects, the
effects of organic solvents, mechanical disruption of bacterial cells, biphasic
division after Albertson and others.  The resolving power of any method
discussed depends mainly on the level of restriction endonuclease activity, the
presence of nonspecific endonucleases in the biomass, the presence of
exonucleases and the taxonomy of the microorganisms used.

<>

<1>Sokolov, N.N.
<2>Search, isolation and study of restrictases.
<3>Vestn. Akad. Med. Nauk SSSR
<4>2
<5>47-51
<6>1995
<7>New Class II restrictases were searched in over 800 microorganisms by using a highly sensitive
toluene micro technique developed in the laboratory. This enabled site-specific endonuclease
activity to be revealed in 72 strains. Thirty-two new restriction endonucleases were
identified, which were highly purified and contained no impurities of nonspecific nucleases,
phosphatases. Many of them (LpII, PaeI, PaeBI, ApiI, CsiAI, BavAI, BavAIII, BbvAI, etc.) are
of interest for use in molecular genetic studies. Corynebacterium species cells were used to
isolate a new super coarse hissing restrictase CsiBI that recognizes the 8-nucleotide site
GCGGCCGC (the isoschizomer NotI) and a fine tool for obtaining enlarged fragments of pro- and
eukaryotic genome.

<>

<1>Sokolov, N.N.
<2>Restriction endonucleases: Isolation methods and schemes.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>1-6
<6>1990
<7>The great successes achieved in the last 10-15 years in the field of recombinant DNA
technology have been largely due to the practical use of site-specific endonucleases (class II
restriction endonucleases; EC 3.1.23.X).  We must agree with the opinion of F. Yang, who
believes that one of the major discoveries that determined the course of development of
genetic engineering was the discovery of restriction enzymes by S. Luria and M. Human.  The
enormous interest in restriction endonucleases as research tools is due, on one hand, to their
unique properites - their ability to form sticky ends on DNA fragments, the variety of
recognizable nucleotide sequences, and the positions of the points of cleavage of the
recognition sites.  On the other hand, restriction enodnucleases, which possess extremely high
specificity with respect to the DNA nucleotide sequences that they recognize and hydrolyze,
represent a splendid model for the study of the important general biological problems of
nucleic-protein recognition.  Considering the great interest in restriction endonucleases
among biochemists, molecular biologists, and specialists in allied fields, it is advisable to
examine the basic methodological approaches and the most widespread schemes of isolation of
these enzymes, to assess the difficulties and problems standing in the way of the production
of purified restriction endonuclease preparations.

<>

<1>Sokolov, N.N., Anikeitcheva, N.V., Eldarov, M.A., Fitsner, A.B., Karpichev, I.V., Kalugin, A.A., Samko, O.T., Choroshoutina, E.B., Lidrik, G.I.
<2>New strain of Bacillus coagulans - useful as producer of new restriction endonuclease BcoAI.
<3>Soviet Patent Office
<4>SU 1761804
<5>
<6>1992
<7>
<>

<1>Sokolov, N.N., Anikeitcheva, N.V., Eldarov, M.A., Fitsner, A.B., Karpytsev, I.V., Kalugin, A.A., Samko, O.T., Khoroshutina, E.B.
<2>A strain of the bacterium Bacillus brevis, a producer of restriction endonuclease BbvBI-enzyme production, purification and characterization.
<3>Soviet Patent Office
<4>SU 1832129
<5>
<6>1993
<7>
<>

<1>Sokolov, N.N., Anikeitcheva, N.V., Fitsner, A.B., Samko, O.T., Choroshoutina, E.B., Kalugin, A.A., Lidrik, G.I.
<2>New Bacillus alvei strain - useful for restriction endonuclease BavBII production.
<3>Soviet Patent Office
<4>SU 1761803
<5>
<6>1992
<7>
<>

<1>Sokolov, N.N., Anikeitcheva, N.V., Fitsner, A.B., Samko, O.T., Khoroshutina, E.B., Kalugin, A.A.
<2>Search of strains producing new restriction endonucleases.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>9
<5>86-89
<6>1988
<7>The search for restriction endonucleases in 154 strains belonging to 104
species of 32 genera of microorganisms has been carried out by the method of
rapid toluene assay.  In 10 strains the activity of endonucleases specifically
fragmenting the DNA of phage lambda in the presence of Mg2+ ions has been
detected.  Restriction enzymes PaeI and PaeII found in two Pseudomonas
aeruginosa strains have been identified as the true isoschizomers of
restriction endonucleases SphI and SmaI respectively.  The results of the
screening of restriction enzyme-producing strains indicate that the production
of restriction enzymes is widespread among microorganisms of the genus
Bacillus.

<>

<1>Sokolov, N.N., Eldarov, M.A., Anikeitcheva, N.V., Karpychev, I.V., Samko, O.T., Fitzner, A.B., Kalugin, A.A., Choroshoutina, E.B., Skryabin, K.G.
<2>Site-specific endonuclease BbvBI from Bacillus brevis.
<3>Bioorg. Khim.
<4>18
<5>47-51
<6>1992
<7>A new restriction endonuclease BbvBI free from contaminating non-specific nucleases and
phosphatases was isolated from the Bacillus brevis cells. The enzyme was purified by
fractionating the sonicated cell-free extract in a two-phase PEG/dextran system and subsequent
chromatographies on DEAE-sepharose, blue sepharose and heparin sepharose. The endonuclease
BbvBI displayed the maximal activity at 45 degrees C, pH between 8.0 and 8.5, MgCl2
concentration in the range of 5-10 mM and at the low ionic strength. It is shown that the
enzyme cleaves the sequence G^GYPC'C, with the preferential cleavage of GGTACC and GGACC site
as compared with GGTGCC and GGCGCC. Thus, the restriction endonuclease BbvBI is a true
isoschizomer of nuclease BanI.

<>

<1>Sokolov, N.N., Eldarov, M.A., Anikeitcheva, N.V., Karpychev, I.V., Samko, O.T., Fitzner, A.B., Kalugin, A.A., Choroshoutina, E.B., Skryabin, K.G.
<2>BcoAI, a new site-specific endonuclease from Bacillus coagulans.
<3>Bioorg. Khim.
<4>17
<5>1188-1192
<6>1991
<7>A new site-specific endonuclease was detected in toluene lysates of Bacillus
coagulans AUCM B-732 and designated as BcoAI.  The enzyme was purified by
fractionation of the cell-free extract in the two-phase PEG/dextran system
followed by chromatography on DEAE-sepharose and phosphocellulose and shown to
be free of nonspecific nucleases and phosphatases.  BcoAI has three cleavage
sites on lambda DNA, but does not cleave SV40, pBR322 and pUC19 DNA.  BcoAI
recognizes the sequence 5'CAC^GTG3' on double-stranded DNA and cleaves it as
indicated by the arrow to yield blunt-ended DNA fragments.  Thus, BcoAI is a
true isoschizomer of PmaCI from Pseudomonas maltophila C.

<>

<1>Sokolov, N.N., Eldarov, M.A., Rina, M., Korolev, S.V., Markaki, M., Kalugin, A.A., Omelyanuk, N.M., Skryabin, K.G., Bouriotis, V.
<2>Isolation and characterization of CspBI, a novel NotI isoschizomer from Corynebacterium species B recognizing 5'-GC/GGCCGC-3'.
<3>Biochem. Mol. Biol. Int.
<4>44
<5>433-441
<6>1998
<7>Sixty-seven bacterial strains were surveyed for the presence of type II restriction
endonucleases, especially concerning super-rare-cutting enzymes.  Fourteen strains were found
to contain specific enzymes.  One of them CspBI from Corynebacterium species B was purified
and characterized as an isoschizomer of NotI, which recognizes the palindromic octanucleotide
sequence 5'-GC/GGCCGC-3' and cleaves at the position shown by the arrow.  A comparison
between the cleavage patterns on different DNA's, obtained with partially purified
endonucleases from other detected producers including some strains of Corynebacterium,
Cellulomonas, and Rhizobium has shown that these enzymes do not belong to super-rare-cutting
restriction enodnucleases.

<>

<1>Sokolov, N.N., Fitsner, A.B., Anikeitcheva, N.V., Choroshoutina, Y.B., Samko, O.T., Kolosha, V.O., Fodor, I., Votrin, I.I.
<2>A site-specific endonuclease from Pseudomonas aeruginosa.
<3>Mol. Biol. Rep.
<4>10
<5>159-161
<6>1985
<7>PaeI, a new restriction endonuclease from Pseudomonas aeruginosa clinical
strain was isolated and chracterized.  It recognizes and cleaves the sequence
5'-GCATG^C-3' generating DNA fragments with 3'-tetranucleotide sticky ends.
DNAs of pBR322, SV40 and bacteriophage lambda have one, two and six PaeI
recognition sites, respectively.Seventy-two strains of Pseudomonas,
Clostridium, Escherichia coli, Shigella, Proteus and Saccharomyces were
screened for the presence of site-specific endonucleases.  Here we describe the
PaeI restriction enzyme found in Pseudomonas aeruginosa; other data will be
published elsewhere.Earlier Hinkle and Miller isolated from P. aeruginosa a
PaeR7 restriction endonuclease recognizing and cleaving a sequence
5'-C^TCGAG-3'.  Sequence analysis of DNAs cleaved by PaeI shows that the enzyme
is the isoschizomer of SphI.

<>

<1>Sokolov, N.N., Fitsner, A.B., Anikeitcheva, N.V., Khoroshutina, E.B., Samko, O.T.
<2>Effect of thio-specific reagent on Pseudomonas aeruginosa PaeI and PaeII restriction endonuclease activity.
<3>Biull. Eksp. Biol. Med.
<4>102
<5>695-697
<6>1986
<7>Because of their unique properties class II restriction endonucleases are
widely used in research in molecular biology and genetic engineering.  By now
more than 400 restriction endonucleases have been described, their
physicochemical and catalytic properties and the structure of many of these
enzymes have been reasonably well studied and techniques have been developed
for seeking them in microorganisms and isolating them.  However, there is
hardly any information in the literature on sulfhydryl groups of restriction
endonucleases and their role in interaction with the DNA substrate.  The only
exception is an investigation of the sensitivity of 11 restriction
endonucleases to the action of alkylating and mercury compounds.

<>

<1>Sokolov, N.N., Fitsner, A.B., Anikeitcheva, N.V., Samko, O.T., Khoroshutina, E.B., Kolosha, V.O., Fodor, I.I.
<2>Pseudomonas aeruginosa restrictases:  Physico-chemical properties, substrate specificity, prospects of application.
<3>Biotekhnologiya
<4>3
<5>578-585
<6>1987
<7>The optimal conditions of the reaction catalyzed by restrictases PaeI and PaeII
(the optimum of pH, temperature, effects of divalent cations) were studied.
The activity of restrictase PaeII is dependent on the K+ ions.  The molecular
mass of restrictases PaeI and PaeII is approximately 150 and 130 kD,
respectively.  On the base of data on the substrate specificity and the
structure of the recognition site, it was determined, that restrictases PaeI
and PaeII are isoschizomers of restrictases SphI and SmaI, correspondingly.

<>

<1>Sokolov, N.N., Fitsner, A.B., Anikeycheva, N.V., Kovalenko, N.A.
<2>Role of bivalent cations in endonucleolysis of DNA catalyzed by restrictases.
<3>Vopr. Med. Khim.
<4>6
<5>66-69
<6>1989
<7>Electrophoretic analysis of products obtained after hydrolysis of phage lambda
DNA by means of restrictase PaeI was carried out after preincubation of DNA or
the enzyme with Mg2+ as well as after preincubation of DNA simultaneously with
the enzyme and the subsequent addition of the required components into the
experimental samples.  The analysis showed that Mg2+ ions were apparently not
required for the enzyme-substrate complex formation and caused destabilization
of the complex.  Restrictase PaeI was active when Mg2+ was substituted by Mn2+,
Ca2+, Zn2+, Co2+ but not by Cd2+, Cu2+ or Ni2+.  Experiments with
o-phenanthroline showed that Zn2+ cations are important for the catalytic
activity of restrictase PaeI.  Possible functions of Zn2+ in protein-nucleic
acids recognition are discussed.

<>

<1>Sokolov, N.N., Fitsner, A.B., Boikov, Y.N., Choroshoutina, E.B., Kalugin, A.A.
<2>Production of restriction endonuclease PaeI - using Pseudomonas aeruginosa strain, by disintegration with mixture of lysozyme and sodium EDTA, fraction chromatography etc.
<3>Soviet Patent Office
<4>SU 1592333 A
<5>
<6>1990
<7>Restriction endonuclease PaeI is produced more efficiently by using Pseudomonas aeruginosa
GISK-123 strain as the producer. The cell biomass is disintegrated with a mixture of lysozyme
and Na EDTA taken in amounts of 75-150 microgms./ml. and 0.8-1.0 mM, respectively and the
lysate fractionated consecutively with 0.5-0.9% solution of polyethyleneimine and 0.1-15%
solution of polyethylene glycol 6000. Chromatography and dialysis complete the process.

<>

<1>Sokolov, N.N., Fitsner, A.B., Khoroshutina, E.B., Kheislere, M.Y.
<2>Determination of restriction endonuclease activity in Toluene lysates of bacterial cells.
<3>Biull. Eksp. Biol. Med.
<4>97
<5>163-165
<6>1984
<7>By now more than 350 restriction endonucleases have been described in the
literature and they are widely used in molecular biology and in bioengineering
research.  Selection of strains producing restriction endonucleases with a view
to finding new and unique restriction enzymes is continuously in progress.  For
this reason there is an urgent need for quick and reliable ways of determining
restriction endonuclease activity in bacterial cells.  The method generally
used to estimate restriction endonuclease activity in bacterial cells involves
disintegration of the cells with ultrasound followed by high-speed
centrifugation in order to obtain a cell-free extract.  An essential
shortcoming of this method is that the extracts contain activity of nonspecific
endo- and exonucleases, in the presence of whose action it is not always
possible to reveal specific activity of restriction endonucleases.  For
instance, according to data in the literature, activity of only three of the 16
restriction endonucleases studied can be found in unpurified extracts.

<>

<1>Sokolov, N.N., Fitsner, A.B., Samko, O.T., Khoroshutina, E.B., Kalugin, A.A.
<2>Effect of monovalent cations on activity of site-specific endonuclease PaeII.
<3>Vopr. Med. Khim.
<4>36
<5>65-67
<6>1990
<7>The activity of restrictase PaeII, contrary to known Type II restriction
enzymes (except of true isoschizomer SmaI), depended absolutely on monovalent
cations.  This pattern is atypical for Type II restrictases.  At the same time,
restrictase PaeII was able to hydrolyze DNA as a substrate in the absence of
exogenous Mg2+, if the incubation mixture contained cations K+, Rb+, Cs+ and
NH4+ but not Na+ or Li+.  Mg2+ was found to activate the enzyme in the presence
of monovalent cations.  Based on the protective effect of K+ against
inactivation of restrictase PaeII by means of thiol-affecting reagents and high
temperature as well as on stabilization of the enzyme by KCl during storage,
monovalent cations appear to participate in formation of protein molecular
structure, which is optimal for catalytic effect and resistant to inactivation.

<>

<1>Sokolov, N.N., Fitzner, A.B., Eldarov, M.A., Anikeicheva, N.B., Kalugin, A.A., Samko, O.T., Khoroshoutina, E.B., Fodor, I.
<2>BavAI, a restriction endonuclease from Bacillus alvei.
<3>Nucleic Acids Res.
<4>20
<5>2897
<6>1992
<7>A restriction endonuclease BavAI, free of contaminating nuclease activity was isolated from
Bacillus alvei strain 675 using column chromatography on DEAE cellulose and Blue Sepharose
CL-6B. Optimal conditions for BavAI digestion were determined at 30C in a buffer containing 10
mM MgCl2, 30 mM KCl at pH 7.5-8.3. The yield of the enzyme from 10 grams of wet cells was
22,000 units. The enzyme cleaves lambda phage DNA at 15 sites, Ad-2 DNA at 24 sites and
plasmid pBR322 DNA at a unique site. The single cleavage site of BavAI on pBR322 DNA was
mapped by standard double digestion analysis to approximately position 2000. From these data
we deduced that the restriction enzyme cuts at PvuII sites. This was confirmed by comparing
patterns of lambda DNA digests obtained with BavAI, PvuII and BavAI + PvuII simultaneously.

<>

<1>Sokolov, N.N., Fodor, I.I., Fitsner, A.B., Khoroshutina, E.B., Anikeitcheva, N.V., Samko, O.T., Kolosha, V.O.
<2>A new strain of Pseudomonas aeruginosa-18, a producer of a new restriction endonuclease PaeII - stable for at least 6 months and useful in microbiology and genetic engineering.
<3>Soviet Patent Office
<4>SU 1314669
<5>
<6>1994
<7>
<>

<1>Sokolov, N.N., Fodor, I.I., Fitsner, A.B., Khoroshutina, E.B., Antikejcheva, N.V., Kolosha, V.O., Votrin, I.I.
<2>Manufacture of PaeI cleaving at the site GCATGC from Pseudomonas aeruginosa BU278.
<3>Soviet Patent Office
<4>SU 1249935
<5>
<6>1994
<7>
<>

<1>Sokolov, N.N., Kheislere, M.Y., Alexandrova, S.S., Zildere, A.M., Ansberga, S.E.
<2>A systematic method to isolate restriction endonucleases.
<3>Izv. Akad. Nauk Latv. SSR
<4>12
<5>54-62
<6>1984
<7>None

<>

<1>Sokolov, N.N., Kolosha, V.O., Fitsner, A.B., Anikeitcheva, N.V., Khoroshutina, E.B., Samko, O.T., Fodor, I., Votrin, I.I.
<2>New specific endonucleases PaeI and PaeII from Pseudomonas aeruginosa.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>24-26
<6>1986
<7>Specific endonuclease activities have been found in two Pseudomonas aeruginosa
strains.  Isolation and purification of enzymes and determining their specific
activities have permitted one to find out that PaeI is an isoshizomer of SphI
and digests the sequence 5'-GCATG^C-3'.  Another isolated enzyme PaeII is an
isoschizomer of SmaI and cleaves DNA in a fragment 5'-CCC^GGG-3'.  The use of
PaeI and PaeII enzymes in genetical engineering and their advantages are
discussed.

<>

<1>Sokolov, N.N., Korolev, S.V., Rina, M., Eldarov, M.A., Gervaziev, Y.V., Skryabin, K.G., Bouriotis, V.
<2>New site-specific endonucleases from Brevibacterium species.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>9
<5>35-38
<6>1998
<7>New site-specific endonucleases BecAI and BecAII have been detected in Brevibacterium species
A.  Endonuclease BecAII free from contaminating nonspecific endonucleases, exonucleases, and
phosphatases was isolated by column chromatography on phosphocellulose, heparin sepharose, and
DNA cellulose.  It recognizes and cleaves the 5'-GG/CC-3' sequence and is a true
isoschizomer of HaeIII restriction enzyme.  The other restriction endonuclease, BecAI, cleaves
Ad2 DNA at least by 2 sites but not the DNA of phage lambda, T7, SV40, PhiX174, and plasmids
pBR322 and pUC19.  The substrate specificity of BecAI indicates its appurtenance to the super
rare restriction endonucleases.

<>

<1>Sokolov, N.N., Maneliene, Z.P., Butkus, V.V., Fitzner, A.B., Khoroshutina, E.B., Kalugin, A.A., Janulaitis, A.
<2>Site-specific endonucleases LplI and AagI.
<3>Bioorg. Khim.
<4>16
<5>1040-1044
<6>1990
<7>New site-specific endonucleases LplI and AagI have been isolated from the
Lactobacillus plantarum and Achromobacter agile cells, respectively.  The
enzymes' purification stages included treatment of cell-free extracts with
polyethylenimine, fractionation in a two-phase system by Albertsson's method,
chromatography on blue Sepharose and DEAE-cellulose.  The results of cleavage
of a 5'-32P-labelled oligodeoxynucleotide duplex by restriction endonucleases
LplI and AagI indicate that these enzymes recognize and cut the sequence
AT^CGAT, being therefore true isoschizomers of the ClaI restriction
endonuclease from Caryophanon latum.  The L. plantarum strain has 400 fold more
endonuclease production as compared with the ClaI producer and is preferred for
preparative isolation of LplI.

<>

<1>Sokolov, N.N., Samko, O.T., Anikeitcheva, N.V., Kalugin, A.A., Khoroshutina, E.B., Plutalov, O.V., Birikh, K.R., Berlin, Y.A.
<2>New site-specific endonucleases from Bacillus strains.
<3>Bioorg. Khim.
<4>20
<5>1334-1341
<6>1994
<7>In a search for new restriction endonucleases type II, among forty bacterial strains of the
Bacillus genus two strains producing site-specific endonucleases have been found.
Endonucleases BbvAIII and BspFI, isolated from B. brevis BLM B-677 and B. species F, are shown
to be true isoschizomers of BspMII (Kpn2I) and Sau3AI, respectively.

<>

<1>Sokolov, N.N., Votrin, I.I.
<2>Restriction endonucleases as an instrument in comparative biochemical studies.
<3>Zh. Evol. Biokhim. Fiziol.
<4>15
<5>8-21
<6>1979
<7>
<>

<1>Sokolov, N.N., Votrin, I.I., Fitsner, A.B., Anikeitcheva, N.V.
<2>Isolation and certain properties of restriction endonuclease from Bacillus amyloliquefaciens.
<3>Biokhimiia
<4>43
<5>865-871
<6>1978
<7>A partially purified preparation of the restriction endonuclease BamHI was
isolated from cells of Bacillus amyloliquefaciens.  The proposed method of
purification of restriction enzyme BamHI is a modification of the method
described by Wilson and Young.  Decomposition of the cells with ultrasound,
treatment with streptomycin sulfate, fractionation with ammonium sulfate,
chromatography on DEAE-cellulose and hydroxyapatite, and rechromatography on
DEAE-cellulose were used for the isolation and purification of the enzyme.  The
restriction enzyme BamHI splits the linear double-stranded DNA molecule of
phage lambda into six fragments.  The enzyme is stable to storage in ice in
sodium or potassium phosphate buffer with beta-mercaptoethanol (10 mM) for
1.5-2 months.

<>

<1>Sokolowska, M., Czapinska, H., Bochtler, M.
<2>Crystal structure of the {beta}{beta}{alpha}-Me type II restriction endonuclease Hpy99I with target DNA.
<3>Nucleic Acids Res.
<4>37
<5>3799-3810
<6>2009
<7>The betabetaalpha-Me restriction endonuclease (REase) Hpy99I recognizes the CGWCG target
sequence and cleaves it with unusual stagger (five
nucleotide 5'-recessed ends). Here we present the crystal structure of the
specific complex of the dimeric enzyme with DNA. The Hpy99I protomer
consists of an antiparallel beta-barrel and two beta4alpha2 repeats. Each
repeat coordinates a structural zinc ion with four cysteine thiolates in
two CXXC motifs. The betabetaalpha-Me region of the second beta4alpha2
repeat holds the catalytic metal ion (or its sodium surrogate) via Asp148
and Asn165 and activates a water molecule with the general base His149. In
the specific complex, Hpy99I forms a ring-like structure around the DNA
that contacts DNA bases on the major and minor groove sides via the first
and second beta4alpha2 repeats, respectively. Hpy99I interacts with the
central base pair of the recognition sequence only on the minor groove
side, where A:T resembles T:A and G:C is similar to C:G. The Hpy99I-DNA
co-crystal structure provides the first detailed illustration of the
betabetaalpha-Me site in REases and complements structural information on
the use of this active site motif in other groups of endonucleases such as
homing endonucleases (e.g. I-PpoI) and Holliday junction resolvases (e.g.
T4 endonuclease VII).

<>

<1>Sokolowska, M., Czapinska, H., Bochtler, M.
<2>Hpy188I-DNA pre- and post-cleavage complexes--snapshots of the GIY-YIG nuclease mediated catalysis.
<3>Nucleic Acids Res.
<4>39
<5>1554-1564
<6>2011
<7>The GIY-YIG nuclease domain is present in all kingdoms of life and has diverse functions. It
is found in the eukaryotic flap endonuclease and
Holliday junction resolvase Slx1-Slx4, the prokaryotic nucleotide excision
repair proteins UvrC and Cho, and in proteins of 'selfish' genetic
elements. Here we present the structures of the ternary pre- and
post-cleavage complexes of the type II GIY-YIG restriction endonuclease
Hpy188I with DNA and a surrogate or catalytic metal ion, respectively. Our
structures suggest that GIY-YIG nucleases catalyze DNA hydrolysis by a
single substitution reaction. They are consistent with a previous proposal
that a tyrosine residue (which we expect to occur in its phenolate form)
acts as a general base for the attacking water molecule. In contrast to
the earlier proposal, our data identify the general base with the GIY and
not the YIG tyrosine. A conserved glutamate residue (Glu149 provided in
trans in Hpy188I) anchors a single metal cation in the active site. This
metal ion contacts the phosphate proS oxygen atom and the leaving group
3'-oxygen atom, presumably to facilitate its departure. Taken together,
our data reveal striking analogy in the absence of homology between
GIY-YIG and betabetaalpha-Me nucleases.

<>

<1>Sokolowska, M., Kaus-Drobek, M., Czapinska, H., Tamulaitis, G., Szczepanowski, R.H., Urbanke, C., Siksnys, V., Bochtler, M.
<2>Monomeric Restriction Endonuclease BcnI in the Apo Form and in an Asymmetric Complex with Target DNA.
<3>J. Mol. Biol.
<4>369
<5>722-734
<6>2007
<7>Restriction endonuclease BcnI cleaves duplex DNA containing the sequence CC/SGG (S stands for
C or G, / designates a cleavage position) to generate
staggered products with single nucleotide 5'-overhangs. Here, we show that
BcnI functions as a monomer that interacts with its target DNA in 1:1
molar ratio and report crystal structures of BcnI in the absence and in
the presence of DNA. In the complex with DNA, BcnI makes specific contacts
with all five bases of the target sequence and not just with a half-site,
as the protomer of a typical dimeric restriction endonuclease. Our data
are inconsistent with BcnI dimerization and suggest that the enzyme
introduces double-strand breaks by sequentially nicking individual DNA
strands, although this remains to be confirmed by kinetic experiments.
BcnI is remotely similar to the DNA repair protein MutH and shares
approximately 20% sequence identity with the restriction endonuclease
MvaI, which is specific for the related sequence CC/WGG (W stands for A or
T). As expected, BcnI is structurally similar to MvaI and recognizes
conserved bases in the target sequence similarly but not identically. BcnI
has a unique machinery for the recognition of the central base-pair.

<>

<1>Sokolowska, M., Kaus-Drobeka, M., Czapinska, H., Tamulaitis, G., Siksnys, V., Bochtler, M.
<2>Restriction endonucleases that resemble a component of the bacterial DNA repair machinery.
<3>Cell. Mol. Life Sci.
<4>64
<5>2351-2357
<6>2007
<7>It has long been known that most Type II restriction endonucleases share a conserved core fold
and similar active-sites. The same core folding motif is also present in the MutH protein, a
component of the bacterial DNA mismatch repair machinery. In contrast to most Type II
restriction endonucleases, which assemble into functional dimers and catalyze double-strand
breaks, MutH is a monomer and nicks hemimethylated DNA. Recent biochemical and
crystallographic studies demonstrate that the restriction enzymes BcnI and MvaI share many
additional features with MutH-like proteins, but not with most other restriction
endonucleases. The structurally similar monomers all recognize approximately symmetric target
sequences asymmetrically. Differential sensitivities to slight substrate asymmetries, which
could be altered by protein engineering, determine whether the enzymes catalyze only
single-strand nicks or double-strand breaks.

<>

<1>Solaiman, D.K.Y., Somkuti, G.A.
<2>Isolation and characterization of a type II restriction endonuclease from Streptococcus thermophilus.
<3>FEMS Microbiol. Lett.
<4>67
<5>261-266
<6>1990
<7>A type II restriction endonuclease Sth134I was isolated from Streptococcus
thermophilus strain 134.  The enzyme is an isoschizomer of HpaII. The
restriction endonuclease is most active at Mg(II) >5 mM; in the pH range of
7.5-8; temperature of 50-55C; and (KCl) or (NaCl) below 100 mM.  Double
digestion and ligation experiments experiments showed that Sth134I apparently
recognized and cleaves DNA at the sequence CCGG to produce two-base,
5'-protruding ends.

<>

<1>Solaiman, D.K.Y., Somkuti, G.A.
<2>A type II restriction endonuclease of Streptococcus thermophilus ST117.
<3>FEMS Microbiol. Lett.
<4>80
<5>75-80
<6>1991
<7>Streptococcus thermophilus strain 117 produces a type II restriction
endonuclease designated as Sth117I.  This enzyme was isolated from cell
extracts by membrane filtration and ammonium sulfate fractionation.  Anion
exchange chromatography on DE52 yielded an enzyme preparation free of
nonspecific nucleases.  The optimal reaction conditions for Sth117I are: (1)
[MgCl2] > 5 mM; (2) pH range of 6.5-7; (3) incubation temperature between 37
and 50C; and (4) [NaCl] or [KCl] < 50 mM.  The results of single- and
double-digestion experiments indicated that the Sth117I was an isoschizomer of
BstNI and EcoRII with the recognition sequence of 5'-CCWGG-3'.  The cleavage
site of Sth117I was identified as 5'-CC^WGG-3' by 5'-end analysis.  This was
supported by the results of ligation experiments with Sth117I-restricted DNAs
and the BstNI- or EcoRII-generated fragments.

<>

<1>Solem, A., Chatterjee, P., Caprara, M.G.
<2>A novel mechanism for protein-assisted group I intron splicing.
<3>RNA
<4>8
<5>412-425
<6>2002
<7>Previously it was shown that the Aspergillus nidulans (A.n.) mitochondrial COB intron
maturase, I-AniI, facilitates splicing of the
COB intron in vitro. In this study, we apply kinetic analysis of
binding and splicing along with RNA deletion analysis to gain insight
into the mechanism of I-AniI facilitated splicing. Our results are
consistent with I-AniI and A.n. COB pre-RNA forming a specific but
labile encounter complex that is resolved into the native,
splicing-competent complex. Significantly, kinetic analysis of splicing
shows that the resolution step is rate limiting for splicing. RNA
deletion studies show that I-Anil requires most of the A.n. COB intron
for binding suggesting that the integrity of the I-AniI-binding site
depends on overall RNA tertiary structure. These results, taken
together with the observation that A.n. COB intron lacks significant
stable tertiary structure in the absence of protein, support a model in
which I-AniI preassociates with an unfolded COB intron via a "labile"
interaction that facilitates correct folding of the intron catalytic
core, perhaps by resolving misfolded RNAs or narrowing the number of
conformations sampled by the intron during its search for native
structure. The active intron conformation is then "locked in" by
specific binding of I-AniI to its intron interaction site.

<>

<1>Soler-Bistue, A.J., Birshan, D., Tomaras, A.P., Dandekar, M., Tran, T., Newmark, J., Bui, D., Gupta, N., Hernandez, K., Sarno, R., Zorreguieta, A., Actis, L.A., Tolmasky, M.E.
<2>Klebsiella pneumoniae multiresistance plasmid pMET1: similarity with the Yersinia pestis plasmid pCRY and integrative conjugative elements.
<3>PLoS ONE
<4>3
<5>E1800
<6>2008
<7>BACKGROUND: Dissemination of antimicrobial resistance genes has become an important public
health and biodefense threat. Plasmids are important contributors to the rapid acquisition of
antibiotic resistance by pathogenic bacteria. PRINCIPAL FINDINGS: The nucleotide sequence of
the Klebsiella pneumoniae multiresistance plasmid pMET1 comprises 41,723 bp and includes
Tn1331.2, a transposon that carries the bla(TEM-1) gene and a perfect duplication of a 3-kbp
region including the aac(6')-Ib, aadA1, and bla(OXA-9) genes. The replication region of pMET1
has been identified. Replication is independent of DNA polymerase I, and the replication
region is highly related to that of the cryptic Yersinia pestis 91001 plasmid pCRY. The
potential partition region has the general organization known as the parFG locus. The
self-transmissible pMET1 plasmid includes a type IV secretion system consisting of proteins
that make up the mating pair formation complex (Mpf) and the DNA transfer (Dtr) system. The
Mpf is highly related to those in the plasmid pCRY, the mobilizable high-pathogenicity island
from E. coli ECOR31 (HPI(ECOR31)), which has been proposed to be an integrative conjugative
element (ICE) progenitor of high-pathogenicity islands in other Enterobacteriaceae including
Yersinia species, and ICE(Kp1), an ICE found in a K. pneumoniae strain causing primary liver
abscess. The Dtr MobB and MobC proteins are highly related to those of pCRY, but the
endonuclease is related to that of plasmid pK245 and has no significant homology with the
protein of similar function in pCRY. The region upstream of mobB includes the putative oriT
and shares 90% identity with the same region in the HPI(ECOR31). CONCLUSIONS: The comparative
analyses of pMET1 with pCRY, HPI(ECOR31), and ICE(Kp1 )show a very active rate of genetic
exchanges between Enterobacteriaceae including Yersinia species, which represents a high
public health and biodefense threat due to transfer of multiple resistance genes to pathogenic
Yersinia strains.

<>

<1>Sollner, S., Berkner, S., Lipps, G.
<2>Characterisation of the novel restriction endonuclease SuiI from Sulfolobus islandicus.
<3>Extremophiles
<4>10
<5>629-634
<6>2006
<7>A restriction endonuclease activity from Sulfolobus islandicus REN2H1 was purified by
phosphocellulose and cation exchange chromatography.
The enzyme cuts DNA at the recognition site GCwGC as could be shown by
restriction analysis of plasmids and short synthetic duplex DNA. The
cleavage occurs after the first guanosine base and is inhibited by
5-methylcytosine methylation. The restriction activity is
salt-sensitive and has an optimal activity around 70 degrees C.

<>

<1>Solodukhina, L.I., Korotayev, A.I., Solonin, A.S., Kuzmin, N.P.
<2>Cloning of genes for restriction-modification system Sfl2aI from Shigella flexneri 2a.
<3>Genetika
<4>26
<5>1126-1128
<6>1990
<7>The Sfl2aI system of restriction-modification (RM) was revealed in the cells of
Shigella flexneri encoded by pKMR114 plasmid belonging to the IncN
incompatibility group.  The genes for the Sf12aI RM system were cloned.  The
system was ascribed to the enzymes of the EcoRII specificity, as shown by in
vivo and in vitro experiments.  Restriction analysis of these genes' region and
antigenic properties of the Sfl2aI endonuclease pointed to significant
differences between this and the EcoRII RM system.

<>

<1>Solodukhina, L.I., Manuvakhova, M.S., Korotayev, A.I.
<2>Type II restriction-modification systems from Shigella strains.
<3>Genetika
<4>25
<5>1571-1577
<6>1989
<7>Two restriction-modification systems specified by two plasmids have been
discovered in clinical species of Shigella.  The plasmids are designated
pKMR114 and pKMR115.  Both are of 60800 bp and belong to the IncN
incompatibility group.  The EcoRI, EcoRV, HindIII restriction patterns of both
plasmid DNAs are identical.  As shown by the efficiency of plating of
bacteriophage lambdavir on the strains harbouring plasmids encoding EcoRI,
EcoRII, EcoRIII, EcoRIV, EcoRV systems and plasmids studied, the new plasmids
control synthesis of enzymes with the specificity of EcoRII.  The main
distinctive feature of pKMR114 is the ability to decrease the efficiency of
plating of bacteriophage T4 having glycosylated DNA.

<>

<1>Solovyeva, N.Y., Rautenstein, Y.I.
<2>On the phenomenon of restriction and modification of temperate phages for the lysogenic Streptomyces hygroscopicus culture.
<3>Mikrobiologiia
<4>47
<5>956-959
<6>1978
<7>Temperate phages were isolated from the lysogenic culture of Streptomyces hygroscopicus 0485
in the indicator cultures of S. hygroscopicus 0477 and S. levoris 1331.  The phages were found
to be identical in the morphology of particles and serological properties.  The phenomenon of
cross limitation, by the culture of S. hygroscopicus 0477, of the phage growing on the culture
of S. levoris 1331, and vice versa, was established.  At the same time, the phages were shown
to be modified by the host cell.

<>

<1>Solovyeva, N.Y., Rautenstein, Y.I.
<2>On the restriction and modification of actinophages by the cultures of Streptomyces hygroscopicus and Streptomyces levoris.
<3>Mikrobiologiia
<4>49
<5>512-515
<6>1980
<7>The capability for restriction and modification was studied in the cultures of Streptomyces
hygroscopicus 0477 and Streptomyces levoris 1331, 2340, 2144 toward the active against them
temperate phages and three polyphages.  All these cultures were found to be capable of the
restriction and modification of the temperate phage.  Certain differences in restriction and
modification were established between S. levoris 1331 and the two other cultures of this
species.  The culture 1331 could modify the temperate phage only with respect to itself rather
than the two other cultures.  The phage growing on culture 1331 was restricted not only by
culture 0477, but also by the strains of S. levoris 2340 and 2144.  At the same time, the
phage growing on the strains 2340 and 2144 gave the identical effectiveness of inoculation on
any of these cultures, as well as on the culture 1331, and was restricted to the same degree
by the culture 0477.  One of the examined polyphages 14/3 was not restricted by any of the
tested cultures. Two other polyphages SH4 and p4 were restricted only by the culture 2144.
However, the modification by this culture was not observed.  Among the studied cultures of S.
levoris, the culture 2144 was most capable of the restriction.

<>

<1>Solow, B.T., Somkuti, G.A.
<2>Molecular properties of Streptococcus thermophilus plasmid pER35 encoding a restriction modification system.
<3>Curr. Microbiol.
<4>42
<5>122-128
<6>2001
<7>Bacteriophage attack on lactic fermentation bacteria (LFB) is costly to the dairy industry
because it results in product loss. One mechanism
used by LFB to protect themselves from bacteriophage attack is
restriction of foreign DNA. Three plasmids, pER16, pER35, and pER36,
from three different strains of the thermotolerant dairy fermentation
bacterium Streptococcus thermophilus were sequenced. One of these
plasmids, pER35, isolated from S. thermophilus ST135, encoded a type IC
restriction-modification (R-M) system very similar to those encoded on
plasmids pIL2614 in Lactococcus lactis subsp. lactis and pND861 in
Lactococcus lactis biovar diacetylactis. The high degree of identity
between the R-M systems encoded on pER35, pIL2614, and pND861 indicated
the potential for horizontal transfer of these genes between different
species of lactic fermentation bacteria. Similar to the functional R-M
system encoded on pIL2614 that protects the mesophilic L. lactis subsp.
lactis against phage attack, the R-M system on pER35 most likely
functions in the same role in S. thermophilus ST135. The plasmid pER16
was found to encode the specificity subunit of the R-M system, but not
the R or M subunits. In addition, all three plasmids encoded proteins
that are present on other S. thermophilus plasmids, including a protein
for rolling-circle replication (RepA) and a low-molecular-weight stress
protein (Hsp). The presence of a complete R-M system encoded on a
plasmid in S. thermophilus, a species that often lacks plasmids, is
novel and may be beneficial for protecting S. thermophilus from
bacteriophage attack under dairy fermentation conditions.

<>

<1>Som, S., Bhagwat, A.S., Friedman, S.
<2>Nucleotide sequence and expression of the gene encoding the EcoRII modification enzyme.
<3>Nucleic Acids Res.
<4>15
<5>313-332
<6>1987
<7>The gene coding for the EcoRII modification enzyme has been cloned and the
nucleotide sequence of 1933 base pairs containing the gene has been determined.
The gene codes for a protein of 477 amino acids.  Two transcriptional start
sites have been mapped by S1 mapping.  One deletion that removes 34 N-terminal
amino acids was found to have partial enzyme activity.  Comparison of the
EcoRII methylase sequence with other cytosine methylases revealed several
domains of partial homology among all cytosine methylases.  Cloning the gene in
multicopy pUC vectors increased the expression by 6-18 fold.  A 40 fold
overproduction of the EcoRII methylase was obtained by cloning the gene in the
expression vector carrying the lambda PL promoter.

<>

<1>Som, S., Friedman, S.
<2>Photolabeling of the EcoRII DNA methylase with S-adenosyl-L-methionine.
<3>FASEB J.
<4>2
<5>A567
<6>1988
<7>Ultraviolet irradiation of EcoRII methylase enzyme in the presence of its
substrate, S-adenosyl-L-methionine (SAM), results in the formation of a stable
enzyme-substrate adduct.  Although the extent of photolabeling is low (1-2%),
it is specific for the enzyme.  Heat inactivated enzyme or proteins for which
SAM is not a substrate undergo negligible adduct formation upon uv irradiation.
Formation of the enzyme-substrate adduct can be demonstrated by
SDS-polyacrylamide gel electrophoresis after irradiation of the enzyme in the
presence of either [3H]-methyl or [35S] labeled SAM.  At a concentration of 30
micrograms the SAM analogues S-adenosyl-L-homocysteine (Ki=0.83 micrograms) and
sinefungin (Ki=4.3 micrograms) are effective inhibitors of photolabeling
whereas S-adenosyl-D-homocysteine (Ki=46 micrograms) is a poor inhibitor.
These experiments indicate that SAM becomes covalently bound at the catalytic
SAM binding site.  Studies in which the SAM-photolabeled methylase has been
chemically cleaved at selective sites indicate that binding occurs at a
localized region of the protein.

<>

<1>Som, S., Friedman, S.
<2>Identification of a highly conserved domain in the EcoRII DNA methylase that can be photolabeled with S-adenosylmethionine.
<3>FASEB J.
<4>4
<5>A1839
<6>1990
<7>DNA methyltransferases can be specifically labeled with either [3H]-methyl or
[35S] S-adenosylmethionine (AdoMet) upon uv irradiation (Som and Friedman, J.
Biol. Chem., 1990, in press).  The labeling is believed to occur at the AdoMet
binding site.  With the purpose of localizing the AdoMet binding site of one
such enzyme, the EcoRII methylase (EcoRIIM), we cleaved [3H]-AdoMet labeled
EcoRIIM by chemical and enzyme-catalyzed reactions and isolated the
radiolabeled peptides by SDS-PAGE and by HPLC.  Amino-terminal sequencing of
all such fragments localized a common region where 75-80% of labeling occurred.
This region includes a highly conserved core sequence present in all the
cytosine methylases.  One such fragment was further digested with chymotrypsin
and the amino acid analysis of the resulting radioactive peptide was consistent
with the sequence AGFP(C)QPFSL.  However, the cysteine residue could not be
identified as carboxymethylcysteine.  This PC bond was found to be completely
protected from a cleavage reaction that specifically cleaves the peptide bond
N-terminal to a cysteine residue.  These results suggest that the cysteine
residue is modified by the labeling reaction and is probably located at or
close to the AdoMet binding site.

<>

<1>Som, S., Friedman, S.
<2>Identification of a highly conserved domain in the EcoRII methyltransferase which can be photolabeled with S-adenosyl-L-[methyl-3H] methionine.
<3>J. Biol. Chem.
<4>266
<5>2937-2945
<6>1991
<7>DNA methyltransferases can be photolabeled with S-adenosyl-L-methionine
(AdoMet).  Specific incorporation of radioactivity has been demonstrated after
photolabeling with either [methyl-3H]AdoMet or [35S]AdoMet (Som, S., and
Friedman, S. (1990) J. Biol. Chem. 265, 4278-4283).  The labeling is believed
to occur at the AdoMet binding site.  With the purpose of localizing the site
responsible for [methyl-3H]AdoMet photolabeling, we cleaved the labeled EcoRII
methyltransferase by chemical and enzymatic reactions and isolated the
radiolabeled peptides by sodium dodecylsulfate-polyacrylamide gel
electrophoresis and high pressure liquid chromatography.  The labeled peptides
were identified by amino-terminal sequencing.  A common region was localized
which accounted for 65-70% of the total label.  This region includes a highly
conserved core sequence present in all DNA (cytosine 5)-methyltransferases.
One such fragment was digested further with chymotrypsin, and amino acid
analysis of the resulting 3H-labeled peptide was consistent with the sequence
Ala-Gly-Phe-Pro-(Cys)-Gln-Pro-Phe-Ser-Leu.  However, the cysteine residue was
not recovered as carboxymethylcysteine.  The Pro-Cys bond was found to be
protected from cleavage at cysteine residues after cyanylation.  These results
suggest that the cysteine residue is modified by the labeling reaction.  The
chymotryptic fragment was hydrolyzed enzymatically to single amino acids, and
the labeled amino acid was identified as S-methylcytsteine by thin layer
chromatography.  These results indicate that the cysteine residue is located at
or close to the AdoMet binding site of EcoRII methyltransferase.

<>

<1>Som, S., Friedman, S.
<2>Direct photolabeling of the EcoRII methyltransferase with S-Adenosyl-L-methionine.
<3>J. Biol. Chem.
<4>265
<5>4278-4283
<6>1990
<7>Ultraviolet irradiation of EcoRII methyltransferase in the presence of its substrate,
S-adenosyl-L-methionine (AdoMet), results in the formation of a stable enzyme-substrate
adduct. This adduct can be demonstrated by sodium dodecyl sulfate-polyacrylamide gel
electrophoresis after irradiation of the enzyme in the presence of either [methyl-3H]AdoMet or
[35S]AdoMet. The extent of photolabeling is low. Under optimal conditions 4.5 pmol of
[3H]AdoMet is incorporated into 100 pmol of enzyme. Use of the 8-azido derivative of AdoMet as
the photolabeling substrate increases the incorporation by approximately 2-fold. However, this
adduct, unlike the one formed with AdoMet, is not stable when treated with thiol reagents or
precipitated with trichloracetic acid. A catalytically active conformation of the enzyme is
needed for AdoMet photolabeling. Heat-inactivated enzyme or proteins for which AdoMet is not a
substrate or cofactor do not undergo adduct formation. Two other methyltransferases, MspI and
dam methylases are also shown to form adducts with AdoMet upon UV irradiation. The binding
constant of the EcoRII methyltransferase for AdoMet determined with the photolabeling reaction
is 11 microM, which is similar to the binding constant of 9 microM previously reported
(Freidman, S. (1986) Nucleic Acids Res. 14, 4543-4556). The AdoMet analogs
S-adenosyl-L-homocysteine (Ki=0.83 microM) and sinefungin (Ki=4.3 microM) are effective
inhibitors of photolabeling, whereas S-adenosyl-D-homocysteine (Ki=46 microM) is a poor
inhibitor. These experiments indicate that AdoMet becomes covalently bound at the
AdoMet-binding site on the enzyme molecule. The EcoRII methyltransferase-AdoMet adduct is very
stable and could be used to identify the AdoMet-binding site on DNA methyltransferases.

<>

<1>Som, S., Friedman, S.
<2>Autogenous regulation of the EcoRII methylase gene at the transcriptional level: effect of 5-azacytidine.
<3>EMBO J.
<4>12
<5>4297-4303
<6>1993
<7>mRNA of the EcoRII methylase (M.EcoRII), a type II modification enzyme, was induced when
Escherichia coli carrying a cloned M.EcoRII gene was exposed to the bacteriocidal drug
5-azacytidine. Induction occurred only when transcription was initiated from its own promoter.
When the 5' promoter sequences were deleted or replaced with the lac promoter sequences, no
induction occurred. The induction was independent of the template DNA level, but the presence
of an intact M.EcoRII protein was a requirement. The drug is incorporated into DNA which then
inhibits M.EcoRII by binding tightly to the enzyme. A deletion within the M.EcoRII coding
region caused a marked increase in the basal level of mRNA transcribed from the M.EcoRII
promoter, but no induction occurred upon 5-azacytidine treatment. The level could be reduced
to normal by M.EcoRII in trans. In vitro, the enzyme bound to the sequence upstream of the
transcription start sites and inhibited the initiation of transcription. These experiments
indicate that expression of the M.EcoRII gene was autogenously regulated at the
transcriptional level. Similar regulation is also noted in another DNA (cytosine-5) methylase,
M.MspI.

<>

<1>Som, S., Friedman, S.
<2>Inhibition of transcription in vitro by binding of DNA(cytosine-5)-methylases to DNA templates containing cytosine analogs.
<3>J. Biol. Chem.
<4>269
<5>25986-25991
<6>1994
<7>DNA(cytosine-5)-methylases form tight complexes at their methylation sites when the target
cytosine residue is substituted by analogs such as 5-azacytosine or 5-fluorocytosine. To test
whether such complexes can block RNA transcription in vitro, template DNA-containing
methylation sites were prepared, in which cytosine residues in either the (+)- or (-)-strand
were substituted by the analogs. Such templates, irrespective of the strand in which
substitution was made, could effectively block the elongation of RNA at specific sites when
complexed with EcoRII methylase. The protein-DNA complex probably prevents the unwinding of
the template strands or might directly present itself as a steric block to the advancing RNA
polymerase. RNA synthesis was also inhibited at specific sites due to complex formation
between azacytosine-containing DNA and two other methylases, HhaI and HpaII. The 3' ends of
the interrupted transcripts were mapped and were found to lie within 13-14, 13, and 23
nucleotides of the binding sites on the (-)-strand for HhaI, HpaII, and EcoRII methylases,
respectively. Exonuclease III footprinting revealed that the boundaries of the complexed
methylases, HhaI, HpaII, and EcoRII, on the (-)-strand were within 2-3, 1-2, and 9-10
nucleotides, respectively, of the last nucleotide copied by the RNA polymerase.

<>

<1>Som, S., Friedman, S.
<2>Regulation of EcoRII methyltransferase: effect of mutations on gene expression and in vitro binding to the promoter region.
<3>Nucleic Acids Res.
<4>22
<5>5347-5353
<6>1994
<7>EcoRII methyltransferase (M.EcoRII) which methylates the second C in the sequence CCWGG
(W=A/T) is autogenously regulated by binding to the 5' regulatory region of its gene. DNase I
footprinting experiments demonstrated that purified M.EcoRII protected a 47-49 bp region of
DNA immediately upstream of the ecoRIIM coding region. We have studied this interaction with
mutants of the enzyme, in vitro by DNA binding and in vivo by investigating the repression in
trans of expression of beta-galactosidase from an ecoRIIM-lacZ operon fusion. Two
catalytically active mutants failed to repress expression of the fusion whereas catalytically
inactive mutants had repressor activity. However, with one of the catalytically inactive
mutants, C186S, in which the catalytic Cys was replaced with Ser, and which bound unmethylated
CCWGG sequences, repression could only be demonstrated when those sequences in cellular DNA
were methylated by supplying a cloned dcm gene in trans. In vitro binding of the DNA fragment
containing the ecoRIIIM regulatory region was detected only with the mutants that showed
repressor activity, including C186S. Results indicate that down-regulation of the gene in vivo
and binding to the promoter in vitro are not dependent on the catalytic properties of
M.EcoRII. Mobility shift experiments with C186S also revealed that it could bind either the
promoter or unmethylated CCWGG sites, but not both. We conclude that the concentration of
unmethylated CCWGG sites controls expression from the ecoRIIM promoter.

<>

<1>Som, S., Friedman, S.
<2>Characterization of the intergenic region which regulates the MspI restriction-modification system.
<3>J. Bacteriol.
<4>179
<5>964-967
<6>1997
<7>The 110-bp intergenic region between mspIM and mspIR, the genes encoding the MspI modification
(M.MspI) and restriction (R.MspI) enzymes, respectively, was fused, in both orientations, with
lacZ.  Expression of a single-copy mspIM-lacZ fusion is more than 400-fold stronger than
expression of an mspIR-lacZ fusion.  M.MspI in trans represses expression of the mspIM-lacZ
fusion by binding to the DNA but does not affect expression of the mspIRlacZ fusion.
Transcription start sites of the genes were identified, and a set of nonoverlapping promoters
was assigned.  DNase I footprinting showed that M.MspI binds to a site within the intergenic
region that includes only the mspIM regulatory elements.

<>

<1>Som, S., Yang, L.F., Friedman, S.
<2>Insertion and deletion mutants in the presumed target recognition domain of the EcoRII DNA methylase.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>A436
<6>1991
<7>In order to understand the function of various core domains of
DNA(cytosine-5)-methyltransferases (Mtases) and to locate the target
recognition domain (TRD), we prepared insertion and deletion mutants of the
EcoRII methylase.  Insertion of 4 amino acids between residues 107-108, 117-118
and 186-187 resulted in total loss of catalytic activity.  These insertion are
either within or near the motifs conserved in all Mtases.  Ten other mutants
with insertions placed in variable regions of the methylase retained activity.
In an attempt to prepare mutants with random internal deletions within the
variable region (residues 295-407) believed to contain the TRD, we randomly
fused N- and C-terminal segments deleted for part or all of the variable
region.  Two catalytically active mutants have a deletion of 5 to 7 amino
acids.  Forty-five active isolates were found to have duplications ranging from
a few to more than 100 amino acids.  The duplications occurred within the
entire 400 bp region.  These results indicate that unlike the N-terminal
variable region which can be deleted with retention of enzyme activity, this
internal region, although variable in length in different Mtases, cannot accept
large deletions with retention of activity.

<>

<1>Sommer, R., Schaller, H.
<2>Nucleotide sequence of the recognition site of the B-specific restriction modification system in E. coli.
<3>Mol. Gen. Genet.
<4>168
<5>331-335
<6>1979
<7>Two sB mutations in the genome of bacteriophage fd were located by sequence analysis in the fd
sequence at positions 971 and 6341.  Base changes at or close to these positions in phage M13
and in phage f1 am 124 also correlate with a loss of sensitivity to B restriction.  From the
sequence homology between the sequences at the two sB sites the recognition signal for the E.
coli B restriction/modification enzyme is predicted to be:  5' TGA---8N---TGCT 3' 3'
ACT---8N---ACGA 5'.

<>

<1>Song, C.X., Yu, M., Dai, Q., He, C.
<2>Detection of 5-hydroxymethylcytosine in a combined glycosylation restriction analysis (CGRA) using restriction enzyme Taq(alpha)I.
<3>Bioorg. Med. Chem. Lett.
<4>21
<5>5075-5077
<6>2011
<7>5-Hydroxymethylcytosine (5-hmC) is a newly discovered DNA base in mammalian cells that is
believed to be another important epigenetic
modification. Here we report the use of a methylation-insensitive
restriction enzyme Taq(alpha)I coupled with selective chemical labeling
of 5-hmC in a combined glycosylation restriction analysis (CGRA) to
detect 5-hmC in TCGA sequences. This method, differentiates fully
versus hemi-hydroxymethylated cytosine in the CpG dinucleotide, adds a
new tool to facilitate biological studies of 5-hmC.

<>

<1>Song, G.T., Chen, C.E., Ren, J.S., Qu, X.G.
<2>A Simple, Universal Colorimetric Assay for Endonuclease/Methyltransferase Activity and Inhibition Based on an  Enzyme-Responsive Nanoparticle System.
<3>ACS NANO
<4>3
<5>1183-1189
<6>2009
<7>An enzyme responsive nanoparticle system that uses a DNA-gold nanoparticle (AuNP) assembly as
the substrate has been developed for
the simple, sensitive, and universal monitoring of restriction
endonucleases in real time. This new assay takes advantage of the
palindromic recognition sequence of the restriction nucleases and the
unique optical properties of AuNPs and is simpler than the procedure
previously described by by Xu et al. (Angew. Chem. Int. Ed. Engl. 2007,
46, 3468-3470). Because it involves only one type of ssDNA modified
AuNPs, this assay can be directed toward most of the endonucleases by
simply changing the recognition sequence found within the linker DNA.
In addition, the endonuclease activity could be quantitatively analyzed
by the value of the reciprocal of hydrolysis half time (t(1/2)(-1).
Furthermore, our new design could also be applied to the assay of
methyltransferase activity since the methylation of DNA inhibits its
cleavage by the corresponding restriction endonuclease, and thus, this
new methodology can be easily adapted to high-throughput screening of
methyltransferase inhibitors.

<>

<1>Song, H.S., Lai, F.M., Roche, C.E., Brown, J.A.
<2>Method of excising a nucleic acid sequence from a plant genome.
<3>International Patent Office
<4>WO 2008145731 A
<5>
<6>2008
<7>The present invention relates to a method for excising a nucleic acid sequence from the genome
of a plant or a plant cell.  This method is based on the steps of transforming a plant cell
with a construct encoding a DNA double strand break inducing enzyme (DSBI), generating a
transgenic plant line, performing a transient assay to analyze the functionality of the
transgenic enzyme, crossing the plant line with a line containing a nucleic acid sequence to
be excised and performing an immature embryo conversion or a tissue culture regeneration
through callus formation.  The method can also be reversed, which means that a plant cell is
tranformed with a construct encoding a nucleic acid sequence to be excised, and the crossing
is performed with a plant line containing a DSBI.  As an alternative to the crossing step, a
re-transformation of a transgenic plant line with a second construct can also be performed.
The invention is also directed to a plant obtained by this method, or progeny, propagation
material, part, tissue, cell or cell culture, derived from such a plant.  Finally, the
invention relates to the use of a plant or progeny, propagation material, part, tissue, cell
or cell culture, derived from this method, as aliment, fodder or seeds or for the production
of pharmaceuticals or chemicals.

<>

<1>Song, J., Rechkoblit, O., Bestor, T.H., Patel, D.J.
<2>Structure of DNMT1-DNA complex reveals a role for autoinhibition in maintenance DNA methylation.
<3>Science
<4>331
<5>1036-1040
<6>2011
<7>Maintenance of genomic methylation patterns is mediated primarily by DNA methyltransferase-1
(DNMT1). We have solved structures of mouse and human
DNMT1 composed of CXXC, tandem bromo-adjacent homology (BAH1/2), and
methyltransferase domains bound to DNA-containing unmethylated CpG sites.
The CXXC specifically binds to unmethylated CpG dinucleotide and positions
the CXXC-BAH1 linker between the DNA and the active site of DNMT1,
preventing de novo methylation. In addition, a loop projecting from BAH2
interacts with the target recognition domain (TRD) of the
methyltransferase, stabilizing the TRD in a retracted position and
preventing it from inserting into the DNA major groove. Our studies
identify an autoinhibitory mechanism, in which unmethylated CpG
dinucleotides are occluded from the active site to ensure that only
hemimethylated CpG dinucleotides undergo methylation.

<>

<1>Song, J., Teplova, M., Ishibe-Murakami, S., Patel, D.J.
<2>Structure-based mechanistic insights into DNMT1-mediated maintenance DNA methylation.
<3>Science
<4>335
<5>709-712
<6>2012
<7>DNMT1, the major maintenance DNA methyltransferase in animals, helps to regulate  gene
expression, genome imprinting, and X-chromosome inactivation. We report on
the crystal structure of a productive covalent mouse DNMT1(731-1602)-DNA complex
containing a central hemimethylated CpG site. The methyl group of methylcytosine
is positioned within a shallow hydrophobic concave surface, whereas the cytosine
on the target strand is looped out and covalently anchored within the catalytic
pocket. The DNA is distorted at the hemimethylated CpG step, with side chains
from catalytic and recognition loops inserting through both grooves to fill an
intercalation-type cavity associated with a dual base flip-out on partner
strands. Structural and biochemical data establish how a combination of active
and autoinhibitory mechanisms ensures the high fidelity of DNMT1-mediated
maintenance DNA methylation.

<>

<1>Song, J.Y., Jeong, H., Yu, D.S., Fischbach, M.A., Park, H.S., Kim, J.J., Seo, J.S., Jensen, S.E., Oh, T.K., Lee, K.J., Kim, J.F.
<2>Draft Genome Sequence of Streptomyces clavuligerus NRRL 3585, a Producer of Diverse Secondary Metabolites.
<3>J. Bacteriol.
<4>192
<5>6317-6318
<6>2010
<7>Streptomyces clavuligerus is an important industrial strain that produces a number of
antibiotics, including clavulanic acid and cephamycin C. A
high-quality draft genome sequence of the S. clavuligerus NRRL 3585 strain
was produced by employing a hybrid approach that involved Sanger
sequencing, Roche/454 pyrosequencing, optical mapping, and partial
finishing. Its genome, comprising four linear replicons, one chromosome,
and four plasmids, carries numerous sets of genes involved in the
biosynthesis of secondary metabolites, including a variety of antibiotics.

<>

<1>Song, J.Y., Kim, H.A., Kim, J.S., Kim, S.Y., Jeong, H., Kang, S.G., Kim, B.K., Kwon, S.K., Lee, C.H., Yu, D.S., Kim, B.S., Kim, S.H., Kwon, S.Y., Kim, J.F.
<2>Genome Sequence of the Plant Growth-Promoting Rhizobacterium Bacillus sp. Strain  JS.
<3>J. Bacteriol.
<4>194
<5>3760-3761
<6>2012
<7>Volatile and nonvolatile compounds emitted from the plant growth-promoting rhizobacterium
Bacillus sp. strain JS enhance the growth of tobacco and lettuce.
Here, we report the high-quality genome sequence of this bacterium. Its 4.1-Mb
genome reveals a number of genes whose products are possibly involved in
promotion of plant growth or antibiosis.

<>

<1>Song, J.Y., Kwak, M.J., Lee, K.Y., Kong, H.G., Kim, B.K., Kwon, S.K., Lee, S.W., Kim, J.F.
<2>Draft Genome Sequence of the Antifungal-Producing Plant-Benefiting Bacterium Burkholderia pyrrocinia CH-67.
<3>J. Bacteriol.
<4>194
<5>6649-6650
<6>2012
<7>Burkholderia pyrrocinia CH-67 was isolated from forest soil as a biocontrol agent to be
utilized in agriculture. Here, we report the 8.05-Mb draft genome sequence
of this bacterium. Its genome contains genes involved in biosynthesis of
secondary metabolites and plant growth promotion, which may contribute to
probiotic effects on plants.

<>

<1>Song, J.Y., Yoo, R.H., Jang, S.Y., Seong, W.K., Kim, S.Y., Jeong, H., Kang, S.G., Kim, B.K., Kwon, S.K., Lee, C.H., Yu, D.S., Park, M.S., Cho, S.H., Kim, J.F.
<2>Genome Sequence of Enterohemorrhagic Escherichia coli NCCP15658.
<3>J. Bacteriol.
<4>194
<5>3749-3750
<6>2012
<7>Enterohemorrhagic Escherichia coli causes severe food-borne disease in the guts of humans and
animals. Here, we report the high-quality draft genome sequence of
E. coli NCCP15658 isolated from a patient in the Republic of Korea. Its genome
size was determined to be 5.46 Mb, and its genomic features, including genes
encoding virulence factors, were analyzed.

<>

<1>Song, L., Ren, L., Li, X., Yu, D., Yu, Y., Wang, X., Liu, G.
<2>Complete Genome Sequence of Marinobacter sp. BSs20148.
<3>Genome Announcements
<4>1
<5>e00236-13
<6>2013
<7>Marinobacter sp. BSs20148 was isolated from marine sediment collected from the Arctic Ocean at
a water depth of 3,800 m. Here we report the complete genome
sequence of Marinobacter sp. BSs20148. This genomic information will facilitate
the study of the physiological metabolism, ecological roles, and evolution of the
Marinobacter species.

<>

<1>Song, P., Xu, X., Jiang, L., Zhang, R., Wang, J., Xu, Q., Li, S.
<2>Genome Sequence of Bacillus subtilis SPZ1, an Evolved Strain for Higher Uptake Rate of Tributyrin.
<3>Genome Announcements
<4>1
<5>e00511-13
<6>2013
<7>The lipase-producing strain Bacillus subtilis SPZ1 is isolated from the medium by tributyrin
as the sole carbon source. Here, we present a 4.13-Mb assembly of its
genome sequence, which may provide various kinds of useful information related to
Bacillus spp., such as mechanisms and control of the substrate uptake and protein
secretion pathways.

<>

<1>Song, S., Yuan, X., Liu, S., Zhang, N., Wang, Y., Ke, Y., Xu, J., Huang, L., Chen, Z., Li, Y.
<2>Genome Sequence of Stenotrophomonas maltophilia S028, an Isolate Harboring the AmpR-L2 Resistance Module.
<3>J. Bacteriol.
<4>194
<5>6696
<6>2012
<7>Multidrug-resistant Stenotrophomonas maltophilia has emerged as an important cause of
nosocomial infections, which is attributable mainly to the production of
diverse beta-lactamases by S. maltophilia. The L2 beta-lactamase mediated by the
AmpR-L2 module is the most represented lactamase. Here, we announce the genome
sequence of S028, an isolate harboring the AmpR-L2 module.

<>

<1>Song, T., Kang, B.S., Kim, Y.M.
<2>XspI, a new Type II restriction endonuclease from a Xanthomonas species.
<3>Mol. Cells
<4>8
<5>370-373
<6>1998
<7>A new Type II restriction endonuclease, XspI, was purified 11-fold in seven steps to
homogeneity from Xanthomonas sp. strain YK1 grown aerobically in Luria broth.  The final
specific activity of the purified enzyme was 3890 micrograms of lambda DNA digested per h per
mg of protein.  The molecular weight of the native enzyme was determined to be 54,000.  Sodium
dodecyl sulfate-gel electrophoresis revealed two identical subunits of molecular weight
29,000.  The purified enzyme was most active at 37 C in the presence of 100 mM KCl and 5 mM
MgCl2 under pH range from 7.5 to 8.0.  The enzyme was stable at least for 1 h at 37 C, but was
inactivated after 20 min at 65 C.  XspI recognized the tetranucleotide sequence, 5'-CTAG-3',
and cleaved it between C and T, like its isoschizomers MaeI and BfaI.  The ability of the
source organism to grow aerobically, together with the heat inactivation of the enzyme,
confers practical advantages upon XspI over its known isoschizomers.

<>

<1>Song, Y., Jeong, Y., Shin, H.S., Cho, B.K.
<2>Draft Genome Sequence of Clostridium scatologenes ATCC 25775, a Chemolithoautotrophic Acetogenic Bacterium Producing 3-Methylindole and  4-Methylphenol.
<3>Genome Announcements
<4>2
<5>e00459-14
<6>2014
<7>Clostridium scatologenes ATCC 25775 is a strictly anaerobic and chemolithoautotrophic
acetogenic bacterium that converts syngas into multi-carbon
compounds such as acetate, indole, 3-methylindole, and 4-methylphenol. Here we
report the draft genome sequence of C. scatologenes ATCC 25775 (7.3 Mbp) to
elucidate its metabolic pathway for syngas fermentation.

<>

<1>Song, Y.-H., Rueter, T., Geiger, R.
<2>DNA cleavage by AatI and StuI is sensitive to Escherichia coli dcm methylation.
<3>Nucleic Acids Res.
<4>16
<5>2718
<6>1988
<7>While attempting to subclone the StuI - EcoNI fragment of pRIF 309+ we found that the AGGCCT
site was not cleaved by StuI or its isoschizomer AatI. Upon examination of the DNA sequence it
became evident that this site overlapped an E. coli dcm methylation site CC(T/)AGG, ie.
AGGCCTGG, which when methylated could be resistant to cleavage. To test this proposition we
prepared pRIF 309+ in the dcm- E. coli strain GM 2929 kindly supplied by Dr. B. Bachmann. As
can be seen in Fig. 1 pRIF 309+ prepared from a dcm- host is cleaved by StuI. An identical
result was obtained with AatI.

<>

<1>Sonoki, S., Higuchi, T., Hisamatsu, S.
<2>The method for presenting the biases of nucleic acid-constitutive.
<3>Japanese Patent Office
<4>JP 2010033344 A
<5>
<6>2010
<7>
<>

<1>Soper, B.W.
<2>Host restriction-modification as related to gene transfer in the nitrogen-fixing cyanobacterium, cyanothece sp.
<3>Diss. Abstr.
<4>56
<5>3585B
<6>1996
<7>Cyanothece sp. strain BH68K (ATCC 51142) is a marine unicellular cyanobacterium capable of
oxygenic photosynthesis and aerobic nitrogen fixation in a single cell without undergoing
morphological differentiation.  In order to develop a gene transfer system for this organism,
shuttle vectors are required that are capable of replication in E. coli and Cyanothece sp.
Therefore, the plasmids of several clonal isolates have been characterized.  Each Cyanothece
isolate contains three or four plasmids ranging in size from 4.8 kb to 40 kb.  A small 4.8 kb
plasmid (pSE480), from the clonal isolate Cyanothece sp. strain BH68F, has been subcloned and
restriction mapped.  Ten restriction sites have been mapped, five of which are unique and are
suitable for further subcloning.  Shuttle vectors derived from pSE480 were used in attempt to
transfer genes by natural competency and electroporation.  Lack of transformation prompted the
analysis of Cyanothece sp. strain BH68K for restriction barriers.  Cell wash supernatants
revealed a cell wall-associated nuclease that exhibited non-site-specific degradation of
covalently-closed circular and linear double-stranded DNA molecules, including Cyanothece sp.
strain BH68K cell extracts contain three type II restriction endonucleases, designated
Csp68KI, Csp68KII, and Csp68KIII.  Csp68KI is an isoschizomer of AvaII and recognizes the
nucleotide sequence 5'-G/GWCC-3' (W=A or T).  Cleavage occurred between the guanosine
nucleotides producing 3-bp 5' overhang ends.  Csp68KII is an isoschizomer of AsuII and
restricts DNA at the recognition sequence 5'-TT/CGAA-3'.  Cleavage occurred between thymine
and cytosine producing 2-bp 5' overhang ends.  The third restriction endonuclease, Csp68KIII,
is an isoschizomer of AvaIII and restricts DNA at the recognition sequence 5'-ATGCA/T-3'.
Cleavage occurred between the 3' adenosine and thymine nucleotides producing 4-bp 3'
overhang ends.  The methylase genes M.Csp68KI, M.Csp68KIV and M.Csp68KV have been cloned from
plasmid libraries of Cyanothece sp. strain BH68K.  The last two methylases inhibit restriction
by the four-base, site-specific enzymes MspI and HaeIII, respectively.  Cloned methylase genes
from host restriction-modification systems can be used to construct helper plasmids that
methylate shuttle vectors prior to gene transfer.

<>

<1>Soper, B.W.
<2>Host restriction-modification as related to gene transfer in the nitrogen-fixing cyanobacterium, Cyanothece sp.
<3>Ph.D. Thesis, State University of New York, Binghamton, USA
<4>
<5>1-132
<6>1995
<7>Cyanothece sp. strain BH68K (ATCC 51142) is a marine unicellular
cyanobacterium capable of oxygenic photosynthesis and aerobic nitrogen fixation in a
single cell without undergoing morphological differentiation.  In order to develop a gene
transfer system for this organism, shuttle vectors are required that are capable of
replication
in E. coli and Cyanothece sp.  Therefore, the plasmids of several clonal isolates have been
characterized.  Each Cyanothece isolate contains three or four plasmids ranging in size from
4.8 kb to 40 kb.  A small 4.8 kb plasmid (pSE480), from the clonal isolate Cyanothece sp.
strain BH68F, has been subcloned and restriction mapped.  Ten restriction sites have been
mapped, five of which are unique and are suitable for further subcloning.  Shuttle vectors
derived from pSE480 were used in an attempt to transfer genes by natural competency and
electroporation.  Lack of transformation prompted the analysis of Cyanothece sp. strain
BH68K for restriction barriers.  Cell wash supernatants revealed a cell wall-associated
nuclease that exhibited non-site-specific degradation of covalently- closed circular and
linear double-stranded DNA molecules, including Cyanothece sp. strain BH68K
chromosomal DNA.  The nuclease also degraded Cyanothece sp. total RNA and phage
M13mp18 ssDNA.  Cyanothece sp. strain BH68K cell extracts contain three type II
restriction endonucleases, designated Csp68KI, Csp68KII, and Csp68KIII.  Csp68KI is
an isoschizomer of AvaII and recognizes the nucleotide sequence 5'-G/GWCC-3' (W=A or
T).  Cleavage occurred between the guanosine nucleotides producing 3-bp 5' overhang
ends. Csp68KII is an isoschizomer of AsuII and restricts DNA at the recognition sequence
5'-TT/CGAA-3'.  Cleavage occurred between thymine and cytosine producing 2-bp 5'
overhang ends.  The third restriction endonuclease, Csp68KIII, is an isoschizomer of
AvaIII and restricts DNA at the recognition sequence 5'- ATGCA/T-3'.  Cleavage occurred
between the 3' adenosine and thymine nucleotides producing 4- bp 3' overhang ends.  The
methylase genes M.Csp68KI, M.Csp68KIV and M.Csp68KV have been cloned from
plasmid libraries of Cyanothece sp. strain BH68K.  The last two methylases inhibit
restriction by the four-base, site-specific enzymes MspI and HaeIII, respectively. Cloned
methylase genes from host restriction-modification systems can be used to construct helper
plasmids that methylate shuttle vectors prior to gene transfer.

<>

<1>Soper, B.W., Hollister, W.R., Reddy, K.J.
<2>Restriction-modification systems in the marine, aerobic, nitrogen-fixing unicellular cyanobacterium Cyanothece sp. ATCC 51142.
<3>J. Mar. Biotechnol.
<4>6
<5>183-185
<6>1998
<7>Cyanothece sp. ATCC 51142 possesses six host restriction-modification systems (HRMS), and
these HRMS were designated as Csp68KI to Csp68KVI.  So far, four restriction enzymes have been
characterized biochemically, and the other two were deduced based on the cloning of
corresponding DNA methyltransferase genes M. Csp68KIV and M.Csp68KV from the Cyanothece
genome.  Csp68KI, Csp68KII, Csp68KIII, Csp68KIV, Csp68KV, and Csp68KVI are isoschizomers of
AvaII, AsuII, AvaIII, MspI, HaeIII, and FnuDII, respectively.  The cleavage specificities for
Csp68KI, Csp68KII, Csp68KIII, and Csp68KVI were characterized.  The Cyanothece restriction
enzymes showed different temperature and salt requirements for their optimal activity.  For
example, the restriction enzyme Csp68KII functions optimally at 50 C and Csp68KIII requires
higher salt for its activity.

<>

<1>Soper, B.W., Hollister, W.R., Reddy, K.J.
<2>Characterization of additional host restriction-modification systems in the unicellular cyanobacterium Cyanothece sp.
<3>Biochem. Biophys. Res. Commun.
<4>223
<5>24-30
<6>1996
<7>In order to develop a gene transfer system for the unicellular diazotrophic cyanobacterium
Cyanothece sp. strain BH68K, this organism has been further investigated for the presence of
additional host restriction-modification enzymes other than Csp68KI, previously reported for
Cyanothece sp.  Analysis of cell extracts by phosphocellulose and Mono Q fast protein liquid
chromatography (FPLC) has led to the identification of three new restriction endonucleases.
These enzymes have been designated Csp68KII, Csp68KIII, and Csp68KVI.  Csp68KII is an
isoschizomer of AsuII and restricts DNA at the recognition sequence 5'-TT/CGAA-3'.  Cleavage
occurred between thymine and cytosine producing 2 bp 5' overhang ends.  The third restriction
enodnuclease, Csp68KII, is an isoschizomer of AvaIII and restricts DNA at the recognition
sequence 5'-ATGCA/T-3'.  Cleavage occurred between the 3' adenosine and thymine nucleotides
producing 4 bp 3' overhang ends.  The fourth enzyme identified, Csp68KVI, recognizes CGCG and
cleaves this sequence between the internal guanine and cytosine nucleotides producing blunt
ends.

<>

<1>Soper, B.W., Reddy, K.J.
<2>Identification of a nuclease and host restriction-modification in the unicellular, aerobic nitrogen-fixing cyanobacterium Cyanothece sp.
<3>J. Bacteriol.
<4>176
<5>5565-5570
<6>1994
<7>In the process of developing a gene transfer system for the marine, unicellular,
nitrogen-fixing cyanobacterium Cyanothece sp. strain BH68K, two major restriction barriers
have been identified. A cell wall-associated nuclease exhibited non-site-specific degradation
of covalently closed circular and linear double-stranded DNA molecules, including Cyanothece
sp. strain BH68K chromosomal DNA. The nuclease is easily released from intact cells by using
water or buffer containing Triton X-100. Nuclease activity was undetectable in cell extracts
prepared from water-washed cells. Comparison of the restriction endonuclease susceptibility of
Cyanothece sp. strain BH68K DNA to that of Anabaena sp. strain PCC 7120 revealed that these
organisms have a nearly identical pattern of restriction and therefore may contain similar
systems for DNA methylation. Restriction by DpnI, MboI, and Sau3AI indicated the presence of
adenine methylation. Cyanothece sp. strain BH68KI was easily detected in cell extracts without
extensive purification. Csp68KI is an isoschizomer of AvaII and recognizes the nucleotide
sequence 5'-GG(A/T)CC-3'. Cleavage occurs between the guanosine nucleotides producing 3-bp
5' overhang ends.

<>

<1>Soriano, N., Vincent, P., Moullec, S., Meygret, A., Lagente, V., Kayal, S., Faili, A.
<2>Closed Genome Sequence of Noninvasive Streptococcus pyogenes M/emm3 Strain STAB902.
<3>Genome Announcements
<4>2
<5>e00792-14
<6>2014
<7>We report a closed genome sequence of group A Streptococcus genotype emm3 (GAS M/emm3) strain
STAB902, isolated from a superficial pyodermatitis. The genome is
composed of 1,892,124 bp, 6 integrated prophages, and has 1,858 identified coding
sequences (CDSs). It has been fitted with the two available invasive GAS M/emm3
strains.

<>

<1>Soriano, N., Vincent, P., Piau, C., Moullec, S., Gautier, P., Lagente, V., Faili, A., Kayal, S.
<2>Complete Genome Sequence of Streptococcus pyogenes M/emm44 Strain STAB901, Isolated in a Clonal Outbreak in French Brittany.
<3>Genome Announcements
<4>2
<5>e01174-14
<6>2014
<7>We report the complete genome sequence of an invasive isolate of Streptococcus pyogenes
M/emm44, belonging to a clonal outbreak that occurred in French
Brittany. The genome is composed of 1,795,608 bp, with a GC content of 38.5%, has
1,358 identified coding sequences (CDSs), and harbors a novel Tn916-like
transposon (Tn6253).

<>

<1>Sorokin, D.Y., Lucker, S., Vejmelkova, D., Kostrikina, N.A., Kleerebezem, R., Rijpstra, W.I., Damste, J.S., Le Paslier, D., Muyzer, G., Wagner, M., van Loosdrecht, M.C., Daims, H.
<2>Nitrification expanded: discovery, physiology and genomics of a nitrite-oxidizing bacterium from the phylum Chloroflexi.
<3>ISME J.
<4>6
<5>2245-2256
<6>2012
<7>Nitrite-oxidizing bacteria (NOB) catalyze the second step of nitrification, a
major process of the biogeochemical nitrogen cycle, but the recognized diversity
of this guild is surprisingly low and only two bacterial phyla contain known NOB.
Here, we report on the discovery of a chemolithoautotrophic nitrite oxidizer that
belongs to the widespread phylum Chloroflexi not previously known to contain any
nitrifying organism. This organism, named Nitrolancetus hollandicus, was isolated
from a nitrifying reactor. Its tolerance to a broad temperature range (25-63
degrees C) and low affinity for nitrite (K(s)=1 mM), a complex layered cell
envelope that stains Gram positive, and uncommon membrane lipids composed of
1,2-diols distinguish N. hollandicus from all other known nitrite oxidizers. N.
hollandicus grows on nitrite and CO(2), and is able to use formate as a source of
energy and carbon. Genome sequencing and analysis of N. hollandicus revealed the
presence of all genes required for CO(2) fixation by the Calvin cycle and a
nitrite oxidoreductase (NXR) similar to the NXR forms of the proteobacterial
nitrite oxidizers, Nitrobacter and Nitrococcus. Comparative genomic analysis of
the nxr loci unexpectedly indicated functionally important lateral gene transfer
events between Nitrolancetus and other NOB carrying a cytoplasmic NXR, suggesting
that horizontal transfer of the NXR module was a major driver for the spread of
the capability to gain energy from nitrite oxidation during bacterial evolution.
The surprising discovery of N. hollandicus significantly extends the known
diversity of nitrifying organisms and likely will have implications for future
research on nitrification in natural and engineered ecosystems.

<>

<1>Sorokin, V., Severinov, K., Gelfand, M.S.
<2>Large-Scale Identification and Analysis of C-Proteins<BOOK> Computational Biology of Transcription Factor Binding.
<3>Methods Mol. Biol.
<4>674
<5>269-282
<6>2010
<7>The restriction-modification system is a toxin antitoxin mechanism of bacterial cells to
resist phage attacks. High efficiency comes at a
price of high maintenance costs: (1) a host cell dies whenever it loses
restriction-modification genes and (2) whenever a plasmid with
restriction-modification genes enters a naive cell, modification enzyme
(methylase) has to be expressed prior to the synthesis of the
restriction enzyme (restrictase) or the cell dies. These phenomena
imply a sophisticated regulatory mechanism. During the evolution
several such mechanisms were developed, of which one relies on a
special C(control)protein, a short autoregulatory protein containing an
HTH-domain. Given the extreme diversity among restriction-modification
systems, one could expect that C-proteins had evolved into several
groups that might differ in autoregulatory binding sites architecture.
However, only a few C-proteins (and the corresponding binding sites)
were known before this study. Bioinformatics studies applied to
C-proteins and their binding sites were limited to groups of well-known
C-proteins and lacked systematic analysis. In this work, the authors
use bioinformatics techniques to discover 201 C-protein genes with
predicted autoregulatory binding sites. The systematic analysis of the
predicted sites allowed for the discovery of 10 structural classes of
binding sites.

<>

<1>Sorokin, V., Severinov, K., Gelfand, M.S.
<2>Systematic prediction of control proteins and their DNA binding sites.
<3>Nucleic Acids Res.
<4>37
<5>441-451
<6>2009
<7>We present here the results of a systematic bioinformatics analysis of control (C) proteins, a
class of DNA-binding regulators that control
time-delayed transcription of their own genes as well as restriction
endonuclease genes in many type II restriction-modification systems.
More than 290 C protein homologs were identified and DNA-binding sites
for 70 of new and previously known C proteins were predicted by a
combination of phylogenetic footprinting and motif searches in DNA
upstream of C protein genes. Additional analysis revealed that a large
proportion of C protein genes are translated from leaderless RNA, which
may contribute to time-delayed nature of genetic switches operated by
these proteins. Analysis of genetic contexts of newly identified C
protein genes revealed that they are not exclusively associated with
restriction-modification genes; numerous instances of associations with
genes originating from mobile genetic elements were observed. These
instances might be vestiges of ancient horizontal transfers and
indicate that during evolution ancestral restriction-modification
system genes were the sites of mobile elements insertions.

<>

<1>Sorokulova, I.B., Kramarov, V.M., Reznik, S.R., Smirnov, V.V.
<2>Restriction endonucleases from the genus Bacillus.
<3>Mikrobiol. Zh.
<4>52
<5>8-11
<6>1990
<7>Production regularities of site-specific endonucleases by aerobic spore-forming bacteria,
isolated from different ecological sources, have been studied. It is shown that more than 1/3
of all the cultures studied produce site-specific endonucleases. A dependence of the frequency
of occurrence of bacterial producers on the ecological niche of their separation has been
noticed. The data on production of species-specific restrictases have been obtained which can
serve as additional characteristics for the differentiation of close species of Bacilli.

<>

<1>Sorsa, L.J., Dufke, S., Schubert, S.
<2>Identification of novel virulence-associated loci in uropathogenic Escherichia coli by suppression subtractive hybridization.
<3>FEMS Microbiol. Lett.
<4>230
<5>203-208
<6>2004
<7>To identify novel virulence-associated genes in uropathogenic Escherichia coli (UPEC) strains,
a suppression subtractive hybridization strategy was applied to genomic DNA of four clinical
UPEC isolates from patients suffering from cystitis or pyelonephritis. The genomic DNA of four
isolates (tester strains) was subtracted from the DNA of two different driver strains, the
well characterized UPEC strain CFT073 and the non-pathogenic E. coli K-12 strain MG1655. We
determined the sequence of 172 tester strain-specific DNA fragments, 86 of which revealed only
low or no homology to nucleotide sequences of public databases. We further determined the
virulence association of the 86 novel DNA fragments using each DNA fragment as a probe in
Southern hybridizations of a reference strain collection consisting of 60 extraintestinal
pathogenic E. coli isolates, and 40 non-virulent E. coli strains from stool samples. From
this, 19 novel DNA fragments were demonstrated to be significantly associated with virulent
strains and thus may represent new virulence traits. Our results support the idea of a
considerable genetic variability among UPEC strains and suggest that novel genomic
determinants might contribute to virulence of UPEC.

<>

<1>Sosio, M., Gallo, G., Pozzi, R., Serina, S., Monciardini, P., Bera, A., Stegmann, E., Weber, T.
<2>Draft Genome Sequence of the Microbispora sp. Strain ATCC-PTA-5024, Producing the Lantibiotic NAI-107.
<3>Genome Announcements
<4>2
<5>e01198-13
<6>2014
<7>We report the draft genome sequence of Microbispora sp. strain ATCC-PTA-5024, a soil isolate
that produces NAI-107, a new lantibiotic with the potential to treat
life-threatening infections caused by multidrug-resistant Gram-positive
pathogens. The draft genome of strain Microbispora sp. ATCC-PTA-5024 consists of
8,543,819 bp, with a 71.2% G+C content and 7,860 protein-coding genes.

<>

<1>Soslau, G., Parker, J., Nelson, J.W.
<2>Methylation and restriction endonuclease cleavage of linear Z-DNA in the presence of hexamminecobalt (III) ions.
<3>Nucleic Acids Res.
<4>14
<5>7237-7252
<6>1986
<7>These studies employed the synthetic linear DNA, poly dGdC, in the B and cobalt
hexammine chloride (Co)-induced Z form to determine the effect of conformation
on protein-DNA interactions.  The rate of the reaction of the restriction
endonucleases, HhaI and CfoI, are reduced with Z DNA as compared to B DNA.  The
ability of both restriction endonucleases to react with an aggregate form of Z
DNA (Z* DNA) is found to depend upon how the Z* DNA is formed.  When Z* DNA is
induced by low concentrations of Co (50 microM), the endonucleases remain
active.  In the presence of 100 microM Co, which causes increased aggregation,
the endonucleases are inactive.  The HhaI DNA methyltransferase reacts at equal
rates with the B, Z and low cobalt Z* form.  these results are significantly
different than those observed with Z form dGdC tracts inserted into circular
DNA molecules.

<>

<1>Soslau, G., Pirollo, K.
<2>Selective inhibition of restriction endonuclease cleavage by DNA intercalators.
<3>Biochem. Biophys. Res. Commun.
<4>115
<5>484-491
<6>1983
<7>The preferred dye binding sites and the microenvironment of known nucleotide sequences within
mitochondrial and plasmid pBR322 DNA was probed in a gross fashion with restriction
endonucleases.  The intercalating dyes, ethidium bromide and propidium iodide, do not inhibit
a given restriction endonuclease equally at all of the restriction sites within a DNA
molecule.  The selective inhibition may be explained, in part, by the potential B to Z
conformation transition of DNA flanking the restriction site and by preferred dye binding
sites.  Propidium iodide was found to be a more potent inhibitor than ethidium bromide and the
inhibition is independent of the type of cut made by the enzyme.

<>

<1>Sosnovtsev, S.V., Dedkov, V.S., Rechkunova, N.I., Zernov, I.P., Degtyarev, S.K.
<2>Determination of substrate specificity of restriction endonuclease Mlu113I.
<3>Sib. Biol. J.
<4>0
<5>66-67
<6>1991
<7>The recognition sequence and cleavage point of restriction endonuclease Mlu113I have been
determined as 5'GG/CGCC.

<>

<1>Soule, M., Kuhn, E., Loge, F., Gay, J., Call, D.R.
<2>Using DNA microarrays to identify library-independent markers for bacterial source tracking.
<3>Appl. Environ. Microbiol.
<4>72
<5>1843-1851
<6>2006
<7>Bacterial source tracking is used to apportion fecal pollution among
putative sources. Within this context, library-independent markers are
genetic or phenotypic traits that can be used to identify the host origin
without a need for library-dependent classification functions. The
objective of this project was to use mixed-genome Enterococcus microarrays
to identify library-independent markers. Separate shotgun libraries were
prepared for five host groups (cow, dog, elk/deer, human, and waterfowl),
using genomic DNAs (gDNAs) from ca. 50 Enterococcus isolates for each
library. Microarrays were constructed (864 probes per library), and 385
comparative genomic hybridizations were used to identify putative markers.
PCR assays were used to screen 95 markers against gDNAs from isolates from
known sources collected throughout the United States. This validation
process narrowed the selection to 15 markers, with 7 having no recognized
homologues and the remaining markers being related to genes involved in
metabolic pathways and DNA replication. In most cases, each marker was
exclusive to one of four Enterococcus species (Enterococcus casseliflavus,
E. faecalis, E. hirae, or E. mundtii). Eight markers were highly specific
to either cattle, humans, or elk/deer, while the remaining seven markers
were positive for various combinations of hosts other than humans. Based
on microarray hybridization data, the prevalence of host-specific markers
ranged from 2% to 45% of isolates collected from their respective hosts. A
20-fold difference in prevalence could present challenges for the
interpretation of library-independent markers.

<>

<1>Soundararajan, M., Chang, Z., Morgan, R.D., Heslop, P., Connolly, B.A.
<2>DNA Binding and Recognition by the IIs Restriction Endonuclease MboII.
<3>J. Biol. Chem.
<4>277
<5>887-895
<6>2002
<7>The type IIs restriction endonuclease MboII recognizes nonsymmetrical GAAGA sites, cutting 8
(top strand) and 7 (bottom strand) bases to the right. Gel retardation showed that MboII bound
specifically to GAAGA sequences, producing two distinct complexes each containing one MboII
and one DNA molecule. Interference analysis indicated that the initial species formed, named
complex 1, comprised an interaction between the enzyme and the GAAGA target. Complex 2
involved interaction of the protein with both the GAAGA and the cutting sites. Only in the
presence of divalent metal ions such as Ca(2+) is the conversion of complex 1 to 2 rapid.
Additionally, a very retarded complex was seen with Ca(2+), possibly a (MboII)(2)-(DNA)(2)
complex. Plasmids containing a single GAAGA site were hydrolyzed slowly by MboII. Plasmids
containing two sites were cut far more rapidly, suggesting that the enzyme requires two
recognition sites in the same DNA molecule for efficient hydrolysis. MboII appears to have a
mechanism similar to the best characterized type IIs enzyme, FokI. Both enzymes initially bind
DNA as monomers, followed by dimerization to give an (enzyme)(2)-(DNA)(2) complex.
Dimerization is efficient only when the two target sites are located in the same DNA molecule
and requires divalent metal ions.

<>

<1>Sowers, K.R.
<2>Restriction-modification systems of methanogenic Archaea.
<3>Archaea: Methanogens: A laboratory manual., Cold Spring Harbor Laboratory Press, Sowers, K.R., Schreier, H.J., Plainview, New York
<4>0
<5>505-506
<6>1995
<7>Several type II restriction endonucleases have been isolated from methanogenic Archaea.  MaeI
is the predominant restriction endonuclease isolated from Methanococcus aeolicus; MaeII and
MaeIII appear in lower concentrations.  MaeI and MaeII recognize unique palindromic
tetranucleotide sequences, and MaeIII recognizes a unique palindromic pentanucleotide
sequence.  All three endonucleases produce 5' protruding ends.  MvnI from Methanococcus
vanielii recognizes a tetranucleotide sequence and is an isoschizomer of the
blunt-end-generating endonucleases ThaI and FnuDII.  All four methanogen endonucleases are
available commercially.  Four endonuclease-methyltransferase systems have been identified.
MwoI endonuclease (R.MwoI) and methyltransferase (M.MwoI) isolated from Methanobacterium
wolfei are thermophilic (optimum activity 65oC) and recognize a unique sequence.  R.MwoI
yields three nucleotide 3'-terminal extensions; M.MwoI is hypothesized to methylate C
residues.  Both genes have been cloned and expressed at low levels in Escherichia coli.
Plasmid-encoded R.MthI from Methanobacterium thermoautotrophicum is an isoschizomer of
blunt-end-generating NgoPII.  Deduced amino acid sequence of R.MthTI is highly similar to that
of R.NgoII, and M.MthTI has high sequence similarity to M.NgoPII and M.HaeIII.
Plasmid-encoded MthFI and MthZI from M. thermoautotrophicum are isoschizomers of each other
and of MaeI.  The restriction endonuclease and methyltransferase recognize a palindromic
tetranucleotide.  The deduced amino acid sequence of M.MthZI shows significant similarity to
that of m4C-MTases.  The occurrence of three restriction-modification systems that recognize
CTAG within the methanogenic Archaea and reports that the CTAG sequence in DNA from species of
halophilic Archaea is refractory to digestion by CTAG-specific XbaI, SpeI, and NheI suggest
that CTAG-specific restriction-modification systems are more characteristic in the Archaea
than in the Bacteria.  Continued characterization of restriction-modification systems in the
methanogenic Archaea will be important for determining the compatibility of strains and DNA in
the development of effective transgenic systems among archaeal species.

<>

<1>Sozhamannan, S., Dharmalingam, K.
<2>Molecular cloning and identification of protein products of rglB locus of Escherichia coli.
<3>Gene
<4>74
<5>51-52
<6>1988
<7>Meeting Abstract

<>

<1>Spada, F., Rothbauer, U., Zolghadr, K., Schermelleh, L., Leonhardt, H.
<2>Regulation of DNA methyltransferase 1.
<3>Adv. Enzyme Regul.
<4>46
<5>224-234
<6>2006
<7>Although essentially all cells of a mammalian organism contain the same genetic information
they may differ dramatically in form and function.  This cellular differentiation is
established during development by cascades of transcription factors driving the expression of
cell-type specific sets of genes.  These cell type specific gene expression patterns are
established, maintained and changed in conjunction with epigenetic mechanisms including DNA
methylation, histone modification and chromatin structure.  The most tangible epigenetic mark
is DNA methylation, a postreplicative modification, which in vertebrates is mainly catalyzed
by DNA methyltransferases (Dnmts: EC 2.1.1.37) 1, 3a and 3b at palindromic CpG sites.  The DNA
methylation pattern changes along with cellular differentiation and is generally inversely
correlated with transcriptional activity.  Over the past decade, several functional links
between DNA methylation, histone modification, chromatin structure and transcriptional
regulation have been discovered.  In mammalian cells Dnmt1 is the major and ubiquitously
expressed Dnmt.  By direct and indirect interactions Dnmt1 plays a central role in the
epigenetic networks controlling gene expression in development and disease.  We will outline
results and open questions concerning the regulation of Dnmt1 and dicuss novel approaches to
study Dnmts in living cells.

<>

<1>Spang, A. et al.
<2>The genome of the ammonia-oxidizing Candidatus Nitrososphaera gargensis: Insights into metabolic versatility and environmental adaptations.
<3>Environ. Microbiol.
<4>14
<5>3122-3145
<6>2012
<7>The cohort of the ammonia-oxidizing archaea
(AOA) of the phylum Thaumarchaeota is a diverse,
widespread and functionally important group of
microorganisms in many ecosystems. However, our
understanding of their biology is still very rudimentary
in part because all available genome sequences
of this phylum are from members of the Nitrosopumilus
cluster. Here we report on the complete
genome sequence of Candidatus Nitrososphaera
gargensis obtained from an enrichment culture, representing
a different evolutionary lineage of AOA frequently found in high numbers in many terrestrial
environments. With its 2.83 Mb the genome is much
larger than that of other AOA. The presence of
a high number of (active) IS elements/transposases,
genomic islands, gene duplications and a complete
CRISPR/Cas defence system testifies to its dynamic
evolution consistent with low degree of synteny
with other thaumarchaeal genomes. As expected,
the repertoire of conserved enzymes proposed to be
required for archaeal ammonia oxidation is encoded
by N. gargensis, but it can also use urea and possibly
cyanate as alternative ammonia sources. Furthermore,
its carbon metabolism is more flexible at the
central pyruvate switch point, encompasses the
ability to take up small organic compounds and
might even include an oxidative pentose phosphate
pathway. Furthermore, we show that thaumarchaeota
produce cofactor F420 as well as polyhydroxyalkanoates.
Lateral gene transfer from bacteria and
euryarchaeota has contributed to the metabolic
versatility of N. gargensis. This organisms is well
adapted to its niche in a heavy metal-containing
thermal spring by encoding a multitude of heavy
metal resistance genes, chaperones and mannosylglycerate
as compatible solute and has the genetic
ability to respond to environmental changes by signal
transduction via a large number of two-component
systems, by chemotaxis and flagella-mediated
motility and possibly even by gas vacuole formation.
These findings extend our understanding of thaumarchaeal
evolution and physiology and offer many testable
hypotheses for future experimental research on
these nitrifiers.

<>

<1>Sparling, R., Bhatti, A.R.
<2>Restriction endonuclease activities of Neisseria meningitidis.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>83
<5>214
<6>1983
<7>Restriction endonuclease activities present in different strains of Neisseria
meningitidis were studied.  Enzymes from cell free sonicates were partially
purified by Blue-2 cross linked agarose (Sigma) column chromatography.
Restriction enzyme activity was eluted by 0.5 M NaCl in Tris-HCl buffer pH 7.6.
Excess NaCl was dialyzed out against Tris-HCl buffer pH 7.6.  Restriction
enzyme activity from different strains was assayed by 0.8% agarose slab gel
electrophoresis.  Enzyme preparations from three different strains (DRES-W34,
M1011 and DRES-30) gave three distinct restriction endonuclease (NmeI, NmeIII
and NmeIV) activities.  Using a linear gradient of NaCl (from 0.0 to 0.5 M),
two restriction endonuclease activities NmeI and NmeII present in DRES-W34 were
separated into two distinct peaks.  These two enzyme activities were eluted at
0.15 M NaCl and 0.25 NaCl respectively.  NmeI and NmeII gave 18 and 28 cleavage
fragments when incubated with lambda DNA.  NmeII did not cleave SV-40 and
PhiX174 DNA.  Restriction endonuclease NmeI was further characterized.  It was
inhibited by 0.4 M NaCl but did require MgCl2 for its enzyme activity.  MgCl2
at concentration above 0.02 M inhibited its enzyme activity completely.  NmeI
activity was abolished completely after 30 min of incubation at 65C.  NmeI, II
and III were quite stable when stored at -70C but NmeIV lost its activity.

<>

<1>Sparling, R., Bhatti, A.R.
<2>NmeI, a restriction endonuclease from Neisseria meningitidis.
<3>Microbios
<4>41
<5>73-79
<6>1984
<7>A restriction endonuclease, NmeI, present in Neisseria meningitidis was
partially purified by passing through a blue 2-cross linked agarose column, no
contaminating nucleases remained detectable.  This enzyme cleaved phage lambda,
adenovirus type 2 and PhiX174 DNA but did not cleave SV40 DNA.  It had an
absolute requirement for Mg2+ for its activity and was inhibited by high
concentrations of NaCl or MgCl2.  NmeI activity was completely abolished after
1 h of incubation at 65C.  S-adenosyl-L-methionine and ATP had no effect on its
activity suggesting that NmeI is a type II restriction endonuclease enzyme.  It
is the first report of a restriction enzyme present in N. meningitidis.

<>

<1>Spataro, N., Farfan, M., Albarral, V., Sanglas, A., Loren, J.G., Fuste, M.C., Bosch, E.
<2>Draft Genome Sequence of Aeromonas molluscorum Strain 848TT, Isolated from Bivalve Molluscs.
<3>Genome Announcements
<4>1
<5>e00382-13
<6>2013
<7>We report here the draft genome sequence of Aeromonas molluscorum 848T, the type  strain of
this Aeromonas species, which was isolated from wedge shells (Donax
trunculus) obtained from a retail market in Barcelona, Spain, in 1997.

<>

<1>Spear, M.A.
<2>Efficient DNA subcloning through selective restriction endonuclease digestion.
<3>Biotechniques
<4>28
<5>660-662
<6>2000
<7>Described here is a selective restriction endonuclease digestion method that eliminates the
electrophoresis step that is usually used during the subcloning of new DNA sequences into
typical E. coli-based plasmids. The method increases yield while decreasing laboratory
resource and time utilization. By using donor and acceptor sequences that contain unique
restriction sites found only outside of the intended recombination sequences, the initial
digestion products can be directly combined without electrophoresis if the ligation step is
followed by a selective digestion using the unique restriction enzymes before transformation.
This system is based on the several order of magnitude decrease in transformation efficiency
of linearized compared to circular plasmids. As an example, this method was used to obtain
recombinants between a 3.6 kb acceptor plasmid and 3.0 kb insert following one ligation
reaction after the failure of nine standard reactions using similar amounts of input DNA. It
is particularly applicable to situations in which low subcloning efficiencies are expected.
The technique can be extended to a large percentage of planned recombinations by using
nonidentical compatible cohesive or blunt-ended fragments, or site-directed mutagenesis.

<>

<1>Spence, R.J., Pavasovic, A., Smith, J.J., Prentis, P.J.
<2>Draft Genome Sequence of Enterococcus faecalis Strain PF3, Isolated from Adelie Penguin Feces from Antarctica.
<3>Genome Announcements
<4>2
<5>e01209-13
<6>2014
<7>Enterococcus faecalis is one of the leading causes of nosocomial infections and is a common
commensal organism in humans and other animals. In this study, we
report a draft genome sequence for the E. faecalis strain PF3, isolated from
Adelie penguin feces collected from Warriner Island, Antarctica.

<>

<1>Spencer, D.H., Kas, A., Smith, E.E., Raymond, C.K., Sims, E.H., Hastings, M., Burns, J.L., Kaul, R., Olsen, M.V.
<2>Whole-genome sequence variation among multiple isolates of Pseudomonas aeruginosa.
<3>J. Bacteriol.
<4>185
<5>1316-1325
<6>2003
<7>Whole-genome shotgun sequencing was used to study the sequence variation
of three Pseudomonas aeruginosa isolates, two from clonal infections of
cystic fibrosis patients and one from an aquatic environment, relative to
the genomic sequence of reference strain PAO1. The majority of the PAO1
genome is represented in these strains; however, at least three prominent
islands of PAO1-specific sequence are apparent. Conversely, approximately
10% of the sequencing reads derived from each isolate fail to align with
the PAO1 backbone. While average sequence variation among all strains is
roughly 0.5%, regions of pronounced differences were evident in
whole-genome scans of nucleotide diversity. We analyzed two such divergent
loci, the pyoverdine and O-antigen biosynthesis regions, by complete
resequencing. A thorough analysis of isolates collected over time from one
of the cystic fibrosis patients revealed independent mutations resulting
in the loss of O-antigen synthesis alternating with a mucoid phenotype.
Overall, we conclude that most of the PAO1 genome represents a core P.
aeruginosa backbone sequence while the strains addressed in this study
possess additional genetic material that accounts for at least 10% of
their genomes. Approximately half of these additional sequences are novel.

<>

<1>Spiegel, P.C., Chevalier, B., Sussman, D., Turmel, M., Lemieux, C., Stoddard, B.L.
<2>The Structure of I-CeuI Homing Endonuclease: Evolving Asymmetric DNA Recognition from a Symmetric Protein Scaffold.
<3>Structure
<4>14
<5>869-880
<6>2006
<7>Homing endonucleases are highly specific catalysts of DNA strand breaks, leading to the
transfer of mobile intervening sequences containing the
endonuclease ORF. We have determined the structure and DNA recognition
behavior of I-CeuI, a homodimeric LAGLIDADG endonuclease from
Chlamydomonas eugametos. This symmetric endonuclease displays unique
structural elaborations on its core enzyme fold, and it preferentially
cleaves a highly asymmetric target site. This latter property represents
an early step, prior to gene fusion, in the generation of asymmetric DNA
binding platforms from homodimeric ancestors. The divergence of the
sequence, structure, and target recognition behavior of homing
endonucleases, as illustrated by this study, leads to the invasion of
novel genomic sites by mobile introns during evolution.

<>

<1>Spiess, E., Tomassetti, A., Hernaiz-Driever, P., Pfeifer, G.P.
<2>Structure of mouse DNA (cytosine-5-)-methyltransferase.
<3>Eur. J. Biochem.
<4>177
<5>29-34
<6>1988
<7>DNA (cytosine-5-)-methyltransferase was purified as a single polypeptide (190
kDa by SDS-PAGE) from mouse P815 mastocytoma cells.  This enzyme transfers
methyl groups to unmethylated as well as to hemimethylated DNA sites with a
strong preference for the hemimethylated substrate.  A structural analysis of
the isolated enzyme by electron microscopical techniques was undertaken.  On
the basis of the results obtained, we propose a model for the enzyme structure.
This model describes the enzyme as a hemielliptical globular structure with
dimensions of 5.4-6.7 nm for the height h and 10.3-10.8 nm for the diameter d,
respectively; this globular structure bears a small appendix at the flat side.
A molecular mass of 235-250 kDa is calculated from the measured dimensions.
Limited trypsin digestion of the enzyme led to a 160-kDa fragment which
preserved the gross morphology of the original material.  The possible
structure function relationships are discussed.

<>

<1>Spitzer, E.D.
<2>Restriction enzymes.  To the Editor.
<3>Arch. Pathol. Lab. Med.
<4>114
<5>1190
<6>1990
<7>In his article on restriction enzymes in the Archives, Dr. Ross omits the important effects of
methylation on the activity of restriction enzymes.  The ability of a restriction endonuclease
to cleave and inactivate invading bacteriophage DNA without destroying the bacterial
chromosome depends on the presence within the cell of a corresponding modification methylase
that recognizes the same DNA sequence and methylates either an adenine or a cytosine, thus
rendering the sequence resistant to cleavage.  It is the methylation of restriction sites
present on the chromosome, not the absence of such sites (as suggested in the article), that
enables restriction-modification systems to distinguish "self" from "non- self" DNA;
bacteriophage DNA that has been modified by the EcoRI methylase is also resistant to the
action of the EcoRI endonuclease.

<>

<1>Spoerel, N., Herrlich, P.
<2>Colivirus-T3-coded S-adenosylmethionine hydrolase.
<3>Eur. J. Biochem.
<4>95
<5>227-233
<6>1979
<7>Bacteriophage T3 induces an enzyme activity which hydrolyzes
S-adenosylmethionine.  This S-adenosylmethionine hydrolase is interesting, not
only because of its unique activity, but also because the protein has to
overcome host restriction.  S-adenosylmethionine hydrolase was purified to
homogeneity using affinity chromatography on S-adenosylhomocysteine-Sepharose.
The enzyme occurs in two forms, A and B.  Form A consists of the viral peptide
chain only; its native and subunit molecular weight is 17,000.  Form B
contains, in addition, a host subunit with a molecular weight of 49,000.  The
host subunit does not modifiy S-adenosylmethionine cleavage in vitro and no
apparent relationship to the host-restriction system could be detected.

<>

<1>Spoerel, N., Herrlich, P., Bickle, T.A.
<2>A novel bacteriophage defence mechanism: the anti-restriction protein.
<3>Nature
<4>278
<5>30-34
<6>1979
<7>Bacteriophage T3 and T7 protect their DNA from restriction by producing, as the
earliest detectable phage functions, anti-restriction proteins.  Although the
two phage proteins differ in their chromatographic and antigenic properties,
they act by the same mechanism: the anti-restriction proteins inhibit E. coli
K12 restriction endonuclease by direct interaction.

<>

<1>Sprague, L.D., Neubauer, H.
<2>Genome Sequence of Yersinia similis Y228T, a Member of the Yersinia pseudotuberculosis Complex.
<3>Genome Announcements
<4>2
<5>e00216-14
<6>2014
<7>We report here on the genome sequence of Yersinia similis 228(T) isolated in Germany. The
genome has a size of 4.9 Mb and a G+C content of 47% and is
predicted to contain 4,135 coding sequences. Annotation of the 60,687-bp
extrachromosomal element predicted 67 coding sequences and a G+C content of
47.8%.

<>

<1>Spring, S. et al.
<2>Complete genome sequence of the sulfate-reducing firmicute Desulfotomaculum ruminis type strain (DL(T)).
<3>Standards in Genomic Sciences
<4>7
<5>304-319
<6>2012
<7>Desulfotomaculum ruminis Campbell and Postgate 1965 is a member of the large genus
Desulfotomaculum which contains 30 species and is contained in the family
Peptococcaceae. This species is of interest because it represents one of the few
sulfate-reducing bacteria that have been isolated from the rumen. Here we
describe the features of D. ruminis together with the complete genome sequence
and annotation. The 3,969,014 bp long chromosome with a total of 3,901
protein-coding and 85 RNA genes is the second completed genome sequence of a type
strain of the genus Desulfotomaculum to be published, and was sequenced as part
of the DOE Joint Genome Institute Community Sequencing Program 2009.

<>

<1>Spring, S. et al.
<2>Complete genome sequence of Desulfotomaculum acetoxidans type strain (5575).
<3>Standards in Genomic Sciences
<4>1
<5>242-253
<6>2009
<7>Desulfotomaculum acetoxidans Widdel and Pfennig 1977 was one of the first sulfate-reducing
bacteria known to grow with acetate as sole energy and carbon
source. It is able to oxidize substrates completely to carbon dioxide with
sulfate as the electron acceptor, which is reduced to hydrogen sulfide. All
available data about this species are based on strain 5575(T), isolated from
piggery waste in Germany. Here we describe the features of this organism,
together with the complete genome sequence and annotation. This is the first
completed genome sequence of a Desulfotomaculum species with validly published
name. The 4,545,624 bp long single replicon genome with its 4370 protein-coding
and 100 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea
project.

<>

<1>Spring, S. et al.
<2>Complete genome sequence of Desulfohalobium retbaense type strain (HR(100)).
<3>Standards in Genomic Sciences
<4>2
<5>38-48
<6>2010
<7>Desulfohalobium retbaense (Ollivier et al. 1991) is the type species of the polyphyletic genus
Desulfohalobium, which comprises, at the time of writing, two
species and represents the family Desulfohalobiaceae within the
Deltaproteobacteria. D. retbaense is a moderately halophilic sulfate-reducing
bacterium, which can utilize H(2) and a limited range of organic substrates,
which are incompletely oxidized to acetate and CO(2), for growth. The type strain
HR(100) (T) was isolated from sediments of the hypersaline Retba Lake in Senegal.
Here we describe the features of this organism, together with the complete genome
sequence and annotation. This is the first completed genome sequence of a member
of the family Desulfohalobiaceae. The 2,909,567 bp genome (one chromosome and a
45,263 bp plasmid) with its 2,552 protein-coding and 57 RNA genes is a part of
the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Spring, S. et al.
<2>Complete genome sequence of Thermosphaera aggregans type strain (M11TL).
<3>Standards in Genomic Sciences
<4>2
<5>245-259
<6>2010
<7>Thermosphaera aggregans Huber et al. 1998 is the type species of the genus Thermosphaera,
which comprises at the time of writing only one species. This
species represents archaea with a hyperthermophilic, heterotrophic, strictly
anaerobic and fermentative phenotype. The type strain M11TL(T) was isolated from
a water-sediment sample of a hot terrestrial spring (Obsidian Pool, Yellowstone
National Park, Wyoming). Here we describe the features of this organism, together
with the complete genome sequence and annotation. The 1,316,595 bp long single
replicon genome with its 1,410 protein-coding and 47 RNA genes is a part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Spring, S., Fiebig, A., Riedel, T., Goker, M., Klenk, H.P.
<2>Genome Sequence of Gammaproteobacterial Pseudohaliea rubra Type Strain DSM 19751, Isolated from Coastal Seawater of the Mediterranean Sea.
<3>Genome Announcements
<4>2
<5>e01208-14
<6>2014
<7>Pseudohaliea rubra strain DSM 19751T is an aerobic marine gammaproteobacterium that was
isolated from surface coastal seawater of the Mediterranean Sea. Here,
we present its genome sequence and annotation. Genome analysis revealed the
presence of genes involved in the synthesis of bacteriochlorophyll-a and the
reserve compound glycogen.

<>

<1>Spuesens, E.B., van de Kreeke, N., Estevao, S., Hoogenboezem, T., Sluijter, M., Hartwig, N.G., van Rossum, A.M., Vink, C.
<2>Variation in a surface-exposed region of the Mycoplasma pneumoniae P40 protein as a consequence of homologous DNA recombination between RepMP5  elements.
<3>Microbiology
<4>157
<5>473-483
<6>2011
<7>Mycoplasma pneumoniae is a human pathogen that causes a range of respiratory tract infections.
The first step in infection is adherence of
the bacteria to the respiratory epithelium. This step is mediated by a
specialized organelle, which contains several proteins (cytadhesins) that
have an important function in adherence. Two of these cytadhesins, P40 and
P90, represent the proteolytic products from a single

<>

<1>Spyridaki, A., Matzen, C., Lanio, T., Jeltsch, A., Simoncsits, A., Athanasiadis, A., Scheuring-Vanamee, E., Kokkinidis, M., Pingoud, A.
<2>Structural and Biochemical Characterization of a New Mg2+ Binding Site Near Tyr94 in the Restriction Endonuclease PvuII.
<3>J. Mol. Biol.
<4>331
<5>395-406
<6>2003
<7>We have determined the crystal structure of the PvuII endonuclease in the presence of Mg(2+).
According to the structural data, divalent metal ion
binding in the PvuII subunits is highly asymmetric. The PvuII-Mg(2+)
complex has two distinct metal ion binding sites, one in each monomer. One
site is formed by the catalytic residues Asp58 and Glu68, and has
extensive similarities to a catalytically important site found in all
structurally examined restriction endonucleases. The other binding site is
located in the other monomer, in the immediate vicinity of the hydroxyl
group of Tyr94; it has no analogy to metal ion binding sites found so far
in restriction endonucleases. To assign the number of metal ions involved
and to better understand the role of Mg(2+) binding to Tyr94 for the
function of PvuII, we have exchanged Tyr94 by Phe and characterized the
metal ion dependence of DNA cleavage of wild-type PvuII and the Y94F
variant. Wild-type PvuII cleaves both strands of the DNA in a concerted
reaction. Mg(2+) binding, as measured by the Mg(2+) dependence of DNA
cleavage, occurs with a Hill coefficient of 4, meaning that at least two
metal ions are bound to each subunit in a cooperative fashion upon
formation of the active complex. Quenched-flow experiments show that DNA
cleavage occurs about tenfold faster if Mg(2+) is pre-incubated with
enzyme or DNA than if preformed enzyme-DNA complexes are mixed with
Mg(2+). These results show that Mg(2+) cannot easily enter the active
center of the preformed enzyme-DNA complex, but that for fast cleavage the
metal ions must already be bound to the apoenzyme and carried with the
enzyme into the enzyme-DNA complex. The Y94F variant, in contrast to
wild-type PvuII, does not cleave DNA in a concerted manner and metal ion
binding occurs with a Hill coefficient of 1. These results indicate that
removal of the Mg(2+) binding site at Tyr94 completely disrupts the
cooperativity in DNA cleavage. Moreover, in quenched-flow experiments Y94F
cleaves DNA about ten times more slowly than wild-type PvuII, regardless
of the order of mixing. From these results we conclude that wild-type
PvuII cleaves DNA in a fast and concerted reaction, because the Mg(2+)
required for catalysis are already bound at the enzyme, one of them at
Tyr94. We suggest that this Mg(2+) is shifted to the active center during
binding of a specific DNA substrate. These results, for the first time,
shed light on the pathway by which metal ions as essential cofactors enter
the catalytic center of restriction endonucleases.

<>

<1>Sreenivas, A., Sathyanarayana, R.G., Shivaji, S.
<2>Draft Genome Sequence of a Psychrophilic Bacterium, Sphingomonas antarcticum, Isolated from the Soils of Schirmacher Oasis, Antarctica.
<3>Genome Announcements
<4>2
<5>e00696-14
<6>2014
<7>We report the 4.5-Mbp genome sequences of Sphingobacterium antarcticum 4BY, a psychrophilic
bacterium isolated from the soils of Schirmacher Oasis, Antarctica.
The draft genome of S. antarcticum strain 4BY consists of 4,566,318 bp with 40.4%
G+C content, 4,234 protein coding genes, and 52 RNAs.

<>

<1>Srikhanta, Y.N., Dowideit, S.J., Edwards, J.L., Falsetta, M.L., Wu, H.J., Harrison, O.B., Fox, K.L., Seib, K.L., Maguire, T.L., Wang, A.H.J., Maiden, M.C., Grimmond, S.M., Apicella, M.A., Jennings, M.P.
<2>Phasevarions Mediate Random Switching of Gene Expression in Pathogenic Neisseria.
<3>PLoS Pathog.
<4>5
<5>e1000400
<6>2009
<7>Many host-adapted bacterial pathogens contain DNA methyltransferases (mod genes) that are
subject to phase-variable expression
(high-frequency reversible ON/OFF switching of gene expression). In
Haemophilus influenzae, the random switching of the modA gene controls
expression of a phase-variable regulon of genes (a phasevarion), via
differential methylation of the genome in the modA ON and OFF states.
Phase-variable mod genes are also present in Neisseria meningitidis and
Neisseria gonorrhoeae, suggesting that phasevarions may occur in these
important human pathogens. Phylogenetic studies on phase-variable mod
genes associated with type III restriction modification (R-M) systems
revealed that these organisms have two distinct mod genes-modA and
modB. There are also distinct alleles of modA (abundant: modA11, 12,
13; minor: modA4, 15, 18) and modB (modB1, 2). These alleles differ
only in their DNA recognition domain. ModA11 was only found in N.
meningitidis and modA13 only in N. gonorrhoeae. The recognition site
for the modA13 methyltransferase in N. gonorrhoeae strain FA1090 was
identified as 5'-AGAAA-3'. Mutant strains lacking the modA11, 12 or 13
genes were made in N. meningitidis and N. gonorrhoeae and their
phenotype analyzed in comparison to a corresponding mod ON wild-type
strain. Microarray analysis revealed that in all three modA alleles
multiple genes were either upregulated or downregulated, some of which
were virulence-associated. For example, in N. meningitidis MC58
(modA11), differentially expressed genes included those encoding the
candidate vaccine antigens lactoferrin binding proteins A and B.
Functional studies using N. gonorrhoeae FA1090 and the clinical isolate
O1G1370 confirmed that modA13 ON and OFF strains have distinct
phenotypes in antimicrobial resistance, in a primary human cervical
epithelial cell model of infection, and in biofilm formation. This
study, in conjunction with our previous work in H. influenzae,
indicates that phasevarions may be a common strategy used by
host-adapted bacterial pathogens to randomly switch between
differentiated cell types.

<>

<1>Srikhanta, Y.N., Fox, K.L., Jennings, M.P.
<2>The phasevarion: phase variation of type III DNA methyltransferases controls coordinated switching in multiple genes.
<3>Nat. Rev. Microbiol.
<4>8
<5>196-206
<6>2010
<7>In several host-adapted pathogens, phase variation has been found to occur in genes that
encode methyltransferases associated with type III restriction-modification systems. It was
recently shown that in the human pathogens Haemophilus influenzae, Neisseria gonorrhoeae and
Neisseria meningitidis phase variation of a type III DNA methyltransferase, encoded by members
of the mod gene family, regulates the expression of multiple genes. This novel genetic system
has been termed the 'phasevarion' (phase-variable regulon). The wide distribution of
phase-variable mod family genes indicates that this may be a common strategy used by
host-adapted bacterial pathogens to randomly switch between distinct cell types.

<>

<1>Srikhanta, Y.N., Gorrell, R.J., Steen, J.A., Gawthorne, J.A., Kwok, T., Grimmond, S.M., Robins-Browne, R.M., Jennings, M.P.
<2>Phasevarion Mediated Epigenetic Gene Regulation in Helicobacter pylori.
<3>PLoS ONE
<4>6
<5>e27569
<6>2011
<7>Many host-adapted bacterial pathogens contain DNA methyltransferases (mod genes) that are
subject to phase-variable expression
(high-frequency reversible ON/OFF switching of gene expression). In
Haemophilus influenzae and pathogenic Neisseria, the random switching
of the modA gene, associated with a phase-variable type III restriction
modification (R-M) system, controls expression of a phase-variable
regulon of genes (a 'phasevarion'), via differential methylation of
the genome in the modA ON and OFF states. Phase-variable type III R-M
systems are also found in Helicobacter pylori, suggesting that
phasevarions may also exist in this key human pathogen. Phylogenetic
studies on the phase-variable type III modH gene revealed that there
are 17 distinct alleles in H. pylori, which differ only in their DNA
recognition domain. One of the most commonly found alleles was modH5
(16% of isolates). Microarray analysis comparing the wild-type P12modH5
ON strain to a P12 Delta modH5 mutant revealed that six genes were
either up-or down-regulated, and some were virulence-associated. These
included flaA, which encodes a flagella protein important in motility
and hopG, an outer membrane protein essential for colonization and
associated with gastric cancer. This study provides the first evidence
of this epigenetic mechanism of gene expression in H. pylori.
Characterisation of H. pylori modH phasevarions to define stable
immunological targets will be essential for vaccine development and may
also contribute to understanding H. pylori pathogenesis.

<>

<1>Srikhanta, Y.N., Maguire, T.L., Stacey, K.J., Grimmond, S.M., Jennings, M.P.
<2>The phasevarion: a genetic system controlling coordinated, random switching of expression of multiple genes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>5547-5551
<6>2005
<7>Several host-adapted bacterial pathogens contain methyltransferases associated with type III
restriction-modification (R-M) systems that are
subject to reversible, high-frequency on/off switching of expression
(phase variation). To investigate the role of phase-variable expression of
R-M systems, we made a mutant strain lacking the methyltransferase (mod)
associated with a type III R-M system of Haemophilus influenzae and
analyzed its phenotype. By microarray analysis, we identified a number of
genes that were either up- or down-regulated in the mod mutant strain.
This system reports the coordinated random switching of a set of genes in
a bacterial pathogen and may represent a widely used mechanism.

<>

<1>Srilunchang, M., Homchampa, P., Proungvitaya, T., Wongratanacheewin, S.
<2>Construction of a marker-free DNA adenine methylase (Dam) mutant of Burkholderia pseudomallei.
<3>Tissue Antigens
<4>66
<5>550
<6>2005
<7>Burkholderia pseudomallei is the causative agent of melioidosis, a severe disease of humans
and animals.  Presently no vaccine exists to protect against meliodosis.  It has recently been
shown that DNA adenine methylase mutants of Salmonella enterica serovar Typhimurium are
markedly attenuated but highly effective as live vaccines against Salmonella infection of mice
and chickens.  To determine if this observation could be extended to B. pseudomallei, a
marker-free Dam mutant of B. pseudomallei was constructed and characterized.  The deletion has
been fully defined at the molecular level with no foreign DNA or drug resistance gene remain
in the B. pseudomallei mutant strain.  The dam mutant of B. pseudomallei was constructed by
using in vitro directed mutagenesis.  The first step, the B. pseudomallei dam gene was
PCR-amplified and the PCR product was digested with SmaI/SalI and ligated into pUC18.  The
ligated products were introduced into E.coli DH5a by electroporation and one clone with the
predicted insert size was selected and designated pPHE157.  The cloned gene was sequenced and
found 100% identity with that of the published sequence.  The next step, the 175 bp StyI/EagI
fragment located within the dam gene in pPHE157 was deleted to yield pPHE158.  The SmaI/SalI
fragment containing the Delta-dam was cloned into pEG19Tc replacement vector containing the
counterselectable sacB marker and transferred from E. coli PHE122-1 to B. pseudomallei
wild-type strain by conjugation method.  One desired dam mutants of B. pseudomallei designated
PHB 136 was positively identified on the basis of sucrose tolerance on sucrose containing
media.  The phenotype of this dam mutant strain was confirmed by its sensitivity to the
2-aminopurine and by restriction analysis using MboI endonuclease, which recognize and cleaves
DNA at non-methylated GATC sequences.  The genotype of the dam mutant was further confirmed by
PCR and Southern blot analysis.  Studies aimed to determine the attenuation level and vaccine
efficacy in a mouse model of this dam mutant are underway.

<>

<1>Srinivasan, V., Metcalf, B.J., Knipe, K.M., Ouattara, M., McGee, L., Shewmaker, P.L., Glennen, A., Nichols, M., Harris, C., Brimmage, M., Ostrowsky, B., Park, C.J., Schrag, S.J., Frace, M.A., Sammons, S.A., Beall, B.
<2>vanG element insertions within a conserved chromosomal site conferring vancomycin resistance to Streptococcus agalactiae and Streptococcus anginosus.
<3>MBio
<4>5
<5>E01386-E01314
<6>2014
<7>Three vancomycin-resistant streptococcal strains carrying vanG elements (two
invasive Streptococcus agalactiae isolates [GBS-NY and GBS-NM, both serotype II
and multilocus sequence type 22] and one Streptococcus anginosus [Sa]) were
examined. The 45,585-bp elements found within Sa and GBS-NY were nearly identical
(together designated vanG-1) and shared near-identity over an ~15-kb overlap with
a previously described vanG element from Enterococcus faecalis. Unexpectedly,
vanG-1 shared much less homology with the 49,321-bp vanG-2 element from GBS-NM,
with widely different levels (50% to 99%) of sequence identity shared among 44
related open reading frames. Immediately adjacent to both vanG-1 and vanG-2 were
44,670-bp and 44,680-bp integrative conjugative element (ICE)-like sequences,
designated ICE-r, that were nearly identical in the two group B streptococcal
(GBS) strains. The dual vanG and ICE-r elements from both GBS strains were
inserted at the same position, between bases 1328 and 1329, within the identical
RNA methyltransferase (rumA) genes. A GenBank search revealed that although most
GBS strains contained insertions within this specific site, only sequence type 22
(ST22) GBS strains contained highly related ICE-r derivatives. The vanG-1 element
in Sa was also inserted within this position corresponding to its rumA homolog
adjacent to an ICE-r derivative. vanG-1 insertions were previously reported
within the same relative position in the E. faecalis rumA homolog. An ICE-r
sequence perfectly conserved with respect to its counterpart in GBS-NY was
apparent within the same site of the rumA homolog of a Streptococcus dysgalactiae
subsp. equisimilis strain. Additionally, homologous vanG-like elements within the
conserved rumA target site were evident in Roseburia intestinalis. Importance:
These three streptococcal strains represent the first known vancomycin-resistant
strains of their species. The collective observations made from these strains
reveal a specific hot spot for insertional elements that is conserved between
streptococci and different Gram-positive species. The two GBS strains potentially
represent a GBS lineage that is predisposed to insertion of vanG elements.

<>

<1>Srivastav, R., Singh, A., Jangir, P.K., Kumari, C., Muduli, S., Sharma, R.
<2>Genome Sequence of Staphylococcus massiliensis Strain S46, Isolated from the Surface of Healthy Human Skin.
<3>Genome Announcements
<4>1
<5>e00553-13
<6>2013
<7>Staphylococcus massiliensis strain S46 was isolated from the surface of healthy human skin.
Here, we report the draft genome sequence of S. massiliensis S46
(2,447,110 bp, with a G+C content of 36.3%).

<>

<1>Srivastava, R., Gopinathan, K.P., Ramakrishnan, T.
<2>Deoxyribonucleic acid methylation in mycobacteria.
<3>J. Bacteriol.
<4>148
<5>716-719
<6>1981
<7>Deoxyribonucleic acid modification in six strains of mycobacteria was investigated. The
presence of 5-methylcytosine in the virulent strain
Mycobacterium tuberculosis H37Rv and its absence in the avirulent strain M.
tuberculosis H37Ra and other saprophytic, fast-growing mycobacteria appear to be
the salient features. However, deoxyribonucleic acid from M. smegmatis SN2
lysogenized with the temperature phage I3 showed the presence of
5-methylcytosine. All of the strains had N6-methyladenine.

<>

<1>Srivastava, S., Moraes, C.T.
<2>Manipulating mitochondrial DNA heteroplasmy by a mitochondrially targeted restriction endonuclease.
<3>Hum. Mol. Genet.
<4>10
<5>3093-3099
<6>2001
<7>Mutations in the mitochondrial DNA (mtDNA) can cause a variety of human diseases. In most
cases, such mutations are heteroplasmic (i.e. mutated
and wild-type mtDNA coexist) and a small percentage of wild-type
sequences can have a strong protective effect against a metabolic
defect. Because a genetic approach to correct mtDNA mutations is not
currently available, the ability to modulate heteroplasmy would have a
major impact in the phenotype of many patients with mitochondrial
disorders. We show here that a restriction endonuclease targeted to
mitochondria has this ability. A mitochondrially targeted PstI degraded
mtDNA harboring PstI sites, in some cases leading to a complete loss of
mitochondrial genomes. Recombination between DNA ends released by PstI
was not observed. When expressed in a heteroplasmic rodent cell line,
containing one mtDNA haplotype with two sites for PstI and another
haplotype having none, the mitochondrial PstI caused a significant
shift in heteroplasmy, with an accumulation of the mtDNA haplotype
lacking PstI sites. These experiments provide proof of the principle
that restriction endonucleases are feasible tools for genetic therapy
of a sub-group of mitochondrial disorders. Although this approach is
limited by the presence of mutation-specific restriction sites,
patients with neuropathy, ataxia and retinitis pigmentosa (NARP) could
benefit from it, as the T8399G mutation creates a unique restriction
site that is not present in wild-type human mitochondrial DNA.

<>

<1>Srivastava, S., Moraes, C.T.
<2>Expression of a mitochondrially targeted restriction endonuclease in skeletal muscle causes a mitochondrial myopathy.
<3>Mitochondrion
<4>3
<5>158-159
<6>2003
<7>
<>

<1>Stabler, R.A., Negus, D., Pain, A., Taylor, P.W.
<2>Draft Genome Sequences of Pseudomonas fluorescens BS2 and Pusillimonas noertemannii BS8, Soil Bacteria That Cooperate To Degrade the  Poly-gamma-d-Glutamic Acid Anthrax Capsule.
<3>Genome Announcements
<4>1
<5>e00057-12
<6>2013
<7>A mixed culture of Pseudomonas fluorescens BS2 and Pusillimonas noertemannii BS8  degraded
poly-gamma-d-glutamic acid; when the 2 strains were cultured separately, no hydrolytic
activity was apparent. Here we report the draft genome sequences of both soil isolates.

<>

<1>Stacey, K.A.
<2>Intracellular modification of nucleic acids.
<3>Br. Med. Bull.
<4>21
<5>211-216
<6>1965
<7>None

<>

<1>Stackebrandt, E. et al.
<2>Genome sequence of the free-living aerobic spirochete Turneriella parva type strain (H(T)), and emendation of the species Turneriella parva.
<3>Standards in Genomic Sciences
<4>8
<5>228-238
<6>2013
<7>Turneriella parva Levett et al. 2005 is the only species of the genus Turneriella which was
established as a result of the reclassification of Leptospira parva
Hovind-Hougen et al. 1982. Together with Leptonema and Leptospira, Turneriella
constitutes the family Leptospiraceae, within the order Spirochaetales. Here we
describe the features of this free-living aerobic spirochete together with the
complete genome sequence and annotation. This is the first complete genome
sequence of a member of the genus Turneriella and the 13(th) member of the family
Leptospiraceae for which a complete or draft genome sequence is now available.
The 4,409,302 bp long genome with its 4,169 protein-coding and 45 RNA genes is
part of the G enomic E ncyclopedia of Bacteria and Archaea project.

<>

<1>Stackebrandt, E. et al.
<2>High-quality-draft genome sequence of the yellow-pigmented flavobacterium Joostella marina type strain (En5(T)).
<3>Standards in Genomic Sciences
<4>8
<5>37-46
<6>2013
<7>At present, Joostella marina Quan et al. 2008 is the sole species with a validly  published
name in the genus Joostella, family Flavobacteriacae, phylum
Bacteriodetes. It is a yellow-pigmented, aerobic, marine organism about which
little has been reported other than the chemotaxonomic features required for
initial taxonomic description. The genome of J. marina strain En5(T) complements
a list of 16 Flavobacteriaceae strains for which complete genomes and draft
genomes are currently available. Here we describe the features of this bacterium,
together with the complete genome sequence, and annotation. This is the first
member of the genus Joostella for which a complete genome sequence becomes
available. The 4,508,243 bp long single replicon genome with its 3,944
protein-coding and 60 RNA genes is part of the G enomic E ncyclopedia of Bacteria
and Archaea project.

<>

<1>Stackebrandt, E. et al.
<2>Complete genome sequence of Coriobacterium glomerans type strain (PW2(T)) from the midgut of Pyrrhocoris apterus L. (red soldier bug).
<3>Standards in Genomic Sciences
<4>8
<5>15-25
<6>2013
<7>Coriobacterium glomerans Haas and Konig 1988, is the only species of the genus Coriobacterium,
family Coriobacteriaceae, order Coriobacteriales, phylum
Actinobacteria. The bacterium thrives as an endosymbiont of pyrrhocorid bugs,
i.e. the red fire bug Pyrrhocoris apterus L. The rationale for sequencing the
genome of strain PW2(T) is its endosymbiotic life style which is rare among
members of Actinobacteria. Here we describe the features of this symbiont,
together with the complete genome sequence and its annotation. This is the first
complete genome sequence of a member of the genus Coriobacterium and the sixth
member of the order Coriobacteriales for which complete genome sequences are now
available. The 2,115,681 bp long single replicon genome with its 1,804
protein-coding and 54 RNA genes is part of the G enomic E ncyclopedia of Bacteria
and Archaea project.

<>

<1>Stadtler, P., von Strandmann, R.P., Walter, T., Frey, B., Auer, H., Hengstenberg, W., Schmitz, G.
<2>Purification and characterisation of the restriction endonuclease ItaI from Ilyobacter tartaricus recognizing 5'-GC^NGC-3'.
<3>Gene
<4>137
<5>347-348
<6>1993
<7>ItaI, an isoschizomer of the subclass IIW [Kessler and Manta, Gene 92 (1990) 1-248]
restriction endonuclease (ENase), Fnu4HI [Leung et al., Nucleic Acids Res. 6 (1979) 17-25],
has been isolated from Ilyobacter tartaricus. The ENase has the five-base palindromic
recognition sequence, 5'GC^NGC-3'. It cleaves behind the second nucleotide and produces a
one--nt 5' overhang.

<>

<1>Stahl, B., Barrangou, R.
<2>Complete Genome Sequences of Probiotic Strains Bifidobacterium animalis subsp. lactis B420 and Bi-07.
<3>J. Bacteriol.
<4>194
<5>4131-4132
<6>2012
<7>We present the complete genomes of Bifidobacterium animalis subsp. lactis B420 and Bi-07.
Comparative genomic analysis with the type strain DSMZ10140 revealed
40 to 55 single nucleotide polymorphisms (SNPs) and an indel in a clustered
regularly interspaced short palindromic repeat (CRISPR) locus. These genetic
differences provide a molecular basis for strain typing within the two main
phylogenetic groups of this monomorphic species.

<>

<1>Stahl, B., Barrangou, R.
<2>Complete Genome Sequence of Probiotic Strain Lactobacillus acidophilus La-14.
<3>Genome Announcements
<4>1
<5>e00376-13
<6>2013
<7>We present the 1,991,830-bp complete genome sequence of Lactobacillus acidophilus strain La-14
(SD-5212). Comparative genomic analysis revealed 99.98% similarity
overall to the L. acidophilus NCFM genome. Globally, 111 single nucleotide
polymorphisms (SNPs) (95 SNPs, 16 indels) were observed throughout the genome.
Also, a 416-bp deletion in the LA14_1146 sugar ABC transporter was identified.

<>

<1>Stahl, F., Wende, W., Jeltsch, A., Pingoud, A.
<2>Introduction of asymmetry in the naturally symmetric restriction endonuclease EcoRV to investigate intersubunit communication in the homodimeric protein.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>6175-6180
<6>1996
<7>Type II restriction endonucleases are dimers of two identical subunits that together form one
binding site for the double-stranded DNA substrate.  Cleavage within the palindromic
recognition site occurs in the two strands of the duplex in a concerted manner, due to the
action of two catalytic centers, one per subunit.  To investigate how the two identical
subunits of the restriction endonuclease EcoRV cooperate in binding and cleaving their
substrate, heterodimeric versions of EcoRV with different amino acid substitutions in the two
subunits were constructed.  For this purpose, the ecorV gene was fused to the coding region
for the glutathione-binding domain of the glutathione S-transferase and a His6-tag,
respectively.  Upon cotransformation of Escherichia coli cells with both gene fusions stable
homo- and heterodimers of the EcoRV variants are produced, which can be separated and purified
to homogeneity by affinity chromatography over Ni-nitrilotriacetic acid and glutathione
columns.  A steady-state kinetic analysis shows that the activity of a heterodimeric variant
with one inactive catalytic center is decreased by 2-fold, demonstrating that the two
catalytic centers operate independently from each other.  In contrast, heterodimeric variants
with a defect in one DNA-binding site have a 30- to 50-fold lower activity, indicating that
the two subunits of EcoRV cooperate in the recognition of the palindromic DNA sequence.  By
combining a subunit with an inactive catalytic center with a subunit with a defect in the
DNA-binding site, EcoRV heterodimers were produced that only nick DNA specifically within the
EcoRV recognition sequence.

<>

<1>Stahl, F., Wende, W., Jeltsch, A., Pingoud, A.
<2>The mechanism of DNA cleavage by the type II restriction enzyme EcoRV: Asp36 is not directly involved in DNA cleavage but serves to couple indirect readout to catalysis.
<3>Biol. Chem.
<4>379
<5>467-473
<6>1998
<7>Three different mechanisms have been proposed to describe DNA cleavage by the type II
restriction endonuclease EcoRV, which differ in the number and function of metal ions directly
involved in catalysis and the different roles assigned to amino acid residues in the active
sites and a phosphate group of the substrate.  There are only four acidic amino acid residues
close to the scissile bond: the essential Asp74 and Asp90, the non-essential Glu45, and Asp36.
We show here that Asp36 can be exchanged for alanine, with only minor effects on the cleavage
rate of the nearby phosphodiester bond, excluding that Asp36 could be directly involved in
catalysis.  Hence, the two versions of the two-metal-ion mechanism are not compatible with the
experimental data, because too few ligands for two metal ions are present near the active site
of EcoRV.  Our result, thus, supports the one-metal-ion mechanism for EcoRV.  We suggest that
Asp36 has an allosteric effect by which specific contacts between one strand of the DNA and
one subunit of the enzyme trigger the activation of one catalytic center.  Given the similar
structures of the active sites of EcoRV, EcoRI, BamHI, PvuII and FokI, as well as the
occurrence of a characteristic catalytic motif in several other restriction enzymes, we
conclude that these enzymes most likely share a similar mechanism of DNA cleavage, whose
characteristic feature is the involvement of only one Mg2+ ion in catalysis.

<>

<1>Stahl, F., Wende, W., Wenz, C., Jeltsch, A., Pingoud, A.
<2>Intra- vs intersubunit communication in the homodimeric restriction enzyme EcoRV: Thr37 and Lys38 involved in indirect readout are only important for the catalytic activity of their own subunit.
<3>Biochemistry
<4>37
<5>5682-5688
<6>1998
<7>EcoRV is a dimer of two identical subunits which together form one binding site for the
double-stranded DNA substrate.  Concerted cleavage of both strands of the duplex requires
intersubunit communication to synchronize the two catalytic centers of EcoRV.  Here we address
the question of how contacts to the DNA backbone trigger conformational changes which lead to
the activation of both catalytic centers.  The structure of the specific EcoRV-DNA complex
shows that a region including amino acids Thr37 and Lys38 is involved in interactions with the
DNA backbone and is a candidate for intersubunit communication.  Homodimeric EcoRV T37A and
K38A variants have a 1000-fold reduced catalytic activity.  To examine whether Thr37 and Lys38
of one subunit affect the catalytic center in the same subunit and/or in the other subunit, we
have produced heterodimeric variants containing a Thr37-Ala or Lys38-Ala substitution in one
subunit combined with a wild type subunit or with a subunit which contains an amino acid
substitution (Asp90-Ala) in the active site (D90A/T37A and D90A/K38A).  Cleavage experiments
with supercoiled pAT153 show that wt/T37A and wt/K38A preferentially nick the DNA.  A
steady-state kinetic analysis of the cleavage of an oligodeoxynucleotide substrate shows that
the activity of wt/T37A and wt/K38A is half of that of wild type EcoRV, whereas D90A/T37A and
D90A/K38A are almost inactive.  These results demonstrate that Thr37 and Lys38 affect
primarily the catalytic center in their own subunit and that both subunits of EcoRV can be
activated independently of each other.  We suggest that Thr37 and Lys38 control the catalytic
activity of the active site in their own subunit by positioning a-helix B.

<>

<1>Stakenas, P.S., Zaretskaya, N.M., Maneliene, Z.P., Mauricas, M.M., Butkus, V.V., Yanulaitis, A.A.
<2>MunI mycoplasmic restriction-modification system and its possible role in pathogenesis.
<3>Mol. Biol. (Mosk)
<4>26
<5>546-557
<6>1992
<7>A restriction-modification system named R-M MunI, isolated from Friend's murine
erythroleukemia cells, was purified and characterized. This site-specific endonuclease
recognizes and cleaves the 5'-C'AATTG nucleotide sequence and is an isoschizomer of the MfeI
restrictase from Mycoplasma fermentans. Site-specific methylation modifies the second adenine
residue in the same sequence (5'-CAm6ATTG). It was shown that this enzyme system is from a
Mycoplasma that contaminates the cell line. The Mycoplasma's DNA hybridizes with
species-specific probes for M. fermentans and Mycoplasma arginini. The possible role of
mycoplasmic restriction-modification enzymes in the pathogenesis of aquired immune deficiency
syndrome is discussed.

<>

<1>Stamm, L.V., Greene, S.R., Barnes, N.Y., Bergen, H.L., Hardham, J.M.
<2>Identification and characterization of a Treponema pallidum subsp. pallidum gene encoding a DNA adenine methyltransferase.
<3>FEMS Microbiol. Lett.
<4>155
<5>115-119
<6>1997
<7>The nucleotide sequence of a DNA adenine methyltransferase gene from Treponema pallidum has
been determined.  Southern blot analysis of T. pallidum chromosomal DNA indicated that this
gene is present as a single copy.  The dam gene encodes a 303 amino acid protein whose deduced
sequence has significant homology with DNA (N6-adenine) methyltransferases.  T. pallidum Dam
can be assigned to group alpha DNA amino methyltransferases based on the order of nine
conserved motifs that are present in the protein.  Digests of T. pallidum chromosomal DNA
performed with isoschizomer restriction endonucleases (Sau3AI, DpnI, and MboI) confirmed the
presence of methylated adenine residues in GATC sequences (Dam+ phenotype).

<>

<1>Stampeggioni, E., Palitti, F., Carotti, D.
<2>A convenient assay for DNA methyltransferase.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>226
<6>1989
<7>Assaying DNA methylase by measuring transfer of tritium from the 5 position of
DNA cytosine to water rather than labelled methyl transfer from S-adenosyl
methionine to DNA cuts post-incubation procedures from hours to minutes.  The
methodology and validating data obtained in our laboratory will be presented.

<>

<1>Stamps, B.W., Corsetti, F.A., Spear, J.R., Stevenson, B.S.
<2>Draft genome of a novel chlorobi member assembled by tetranucleotide binning of a hot spring metagenome.
<3>Genome Announcements
<4>2
<5>e00897-14
<6>2014
<7>The genome of a member of the phylum Chlorobi was assembled from a shotgun metagenomic
sequence of a hot spring in Mammoth Lakes, CA. This organism appears
to be a novel, aerobic, photosynthetic Chlorobi member, expanding the knowledge
of this underrepresented phylum.

<>

<1>Stamps, B.W., Losey, N.A., Lawson, P.A., Stevenson, B.S.
<2>Genome Sequence of Thermoanaerobaculum aquaticum MP-01T, the First Cultivated Member of Acidobacteria Subdivision 23, Isolated from a Hot Spring.
<3>Genome Announcements
<4>2
<5>e00570-14
<6>2014
<7>Thermoanaerobaculum aquaticum MP-01(T) is currently the only cultivated and described member
of Acidobacteria subdivision 23. Here, we report the genome
sequence for this novel microorganism that was isolated from a hot spring.

<>

<1>Stanchev, B.S., Grokhovskii, S.L., Khorlin, A.A., Gottikh, B.P., Zhuze, A.L., Skamrov, A.V., Bibilashvili, R.S.
<2>Netropsin, distamycin A, bis-netropsins as selective inhibitors of the effect of restrictases and DNaseI.
<3>Mol. Biol. (Mosk)
<4>20
<5>1614-1624
<6>1986
<7>The influence was investigated of netropsin, a number of bis-netropsins, and
distamycin A on hydrolysis of DNA fragments by restrictases.  In parallel to
inhibition of hydrolysis, binding sites were determined for ligands on the same
DNA fragments according to shielding from partial hydrolysis by DNaseI
(footprinting).  Combined analysis of ligand binding sites with their capacity
for inhibition of hydrolysis led to the following conclusions.  1) Inhibition
of the effect of restrictases by these ligands is associated with formation of
a complex of ligand with the recognition site on DNA.  Experimental results
obtained permit definition of the zone required for overlapping of the site of
restrictase recognition and the binding site of ligand at +/- nucleotides from
the axis of symmetry of the site of restrictase recognition.  The overlapping
of the site occupied by one molecule of ligand with the restrictase recognition
site is not an adequate condition for complete inhibition of hydrolysis.  2)
Based on known specificity of netropsin for a nucleotide sequence, the
restrictase recognition sites can be predicted at which inhibition of
hydrolysis of DNA by restrictases will be observed.  3)  Netropsin and
bis-netropsins display differing specificities and can be used for selective
inhibition of restrictases at different recognition sites.

<>

<1>Stancheva, I., Hensey, C., Meehan, R.R.
<2>Loss of the maintenance methyltransferase, xDnmt1, induces apoptosis in Xenopus embryos.
<3>EMBO J.
<4>20
<5>1963-1973
<6>2001
<7>DNA methylation is necessary for normal embryogenesis in animals. Here we show that loss of
the maintenance methyltransferase, xDnmt1p, triggers an apoptotic response during Xenopus
development, which accounts for the loss of specific cell populations in hypomethylated
embryos. Hypomethylation-induced apoptosis is accompanied by a stabilization in xp53 protein
levels after the mid-blastula transition. Ectopic expression of HPV-E6, which promotes xp53
degradation, prevents cell death, implying that the apoptotic signal is mediated by xp53. In
addition, inhibition of caspase activation by overexpression of Bcl-2 results in the
development of cellular masses that resemble embryonic blastomas. Embryonic tissue explant
experiments suggest that hypomethylation alters the developmental potential of early embryo
cells and that apoptosis is triggered by differentiation. Our results imply that loss of DNA
methylation in differentiated somatic cells provides a signal via p53 that activates cell
death pathways.

<>

<1>Stancheva, I., Koller, T., Sogo, J.M.
<2>Asymmetry of Dam remethylation on the leading and lagging arms of plasmid replicative intermediates.
<3>EMBO J.
<4>18
<5>6542-6551
<6>1999
<7>In Escherichia coli, adenine methylation at the sequence GATC allows coupling of cellular
processes to chromosome replication and the cell cycle. The transient presence of
hemimethylated DNA after replication facilitates post-replicative mismatch repair, induces
transcription of some genes and allows transposition of mobile elements. We were interested in
estimating the half-life of hemimethylated DNA behind the replication fork in plasmid
molecules and in determining whether Dam methyltransferase restores N6 adenine methylation
simultaneously on both replicative arms. We show that remethylation takes place asynchronously
on the leading and lagging daughter strands shortly after replication. On the leading arm the
fully methylated adenine is restored ~2000 bp (corresponding to 2 s) behind the replication
fork, while remethylation takes twice as long (at 3500-4000 bp or ~3.5-4 s) on the lagging
replicative arm. This observation suggests that Dam remethylation of the lagging arm requires
ligated Okazaki fragments.

<>

<1>Stancheva, I., Meehan, R.R.
<2>Transient depletion of xDnmt1 leads to premature gene activation in Xenopus embryos.
<3>Genes Dev.
<4>14
<5>313-327
<6>2000
<7>In Xenopus laevis zygotic transcription begins at the midblastula
transition (MBT). Prior to this the genome is organized into chromatin
that facilitates rapid cycles of DNA replication but not transcription.
Here we demonstrate that DNA methylation contributes to the overall
transcriptional silencing before MBT. Transient depletion of the maternal
DNA methyltransferase (xDnmt1) by anti sense RNA during cleavage stages is
associated with a decrease in the genomic 5-methyl-cytosine content and
leads to the activation of zygotic transcription approximately two cell
cycles earlier than normal. Hypomethylation allows the early expression of
mesodermal marker genes such as Xbra, Cerberus, and Otx2, which are
subsequently down-regulated during gastrulation of the xDnmt1-depleted
embryos. The temporal switch in gene expression may account for the
appearance of body plan defects that we observe. Loss of xDnmt1 can be
rescued by the coinjection of mouse or human Dnmt1 protein. These results
demonstrate that DNA methylation has a role in the regulation of
immediately early genes in Xenopus at MBT.

<>

<1>Stanford, N.P., Halford, S.E., Baldwin, G.S.
<2>DNA cleavage by the EcoRV restriction endonuclease: pH dependence and proton transfers in catalysis.
<3>J. Mol. Biol.
<4>288
<5>105-116
<6>1999
<7>To characterize the pH dependence of phosphodiester hydrolysis by the EcoRV endonuclease in
the presence of Mn2+, single turnover reactions on a 12 bp DNA substrate were examined by
stopped-flow and quench-flow methods between pH 6.0 and 8.5.  At each pH value, the apparent
rate constants for phosphodiester hydrolysis increased hyperbolically with the concentration
of MnCl2, thus allowing values to be determined for the intrinsic rate constant at saturation
with Mn2+ and the equilibrium dissociation constant for Mn2+.  The equilibrium constants
showed no systematic variation across the pH range tested, while the rate constants increased
steeply with increasing pH up to an asymptote above pH 7.5.  At low pH conditions, the
gradient of a plot of log (rate constant) against pH approached a value of 2.  DNA cleavage by
EcoRV thus requires the de-protonation of two acidic groups.  To determine whether aspartate
36 is one of the groups, mutants of EcoRV were made with other amino acid residues at position
36.  Glutamate caused a partial loss of activity, while all other replacements gave near-zero
activities.  In contrast to wild-type EcoRV, the mutant with glutamate required the
de-protonation of only one acidic group for DNA cleavage.  A mechanism for EcoRV is proposed
in which the water molecule that hydrolyses the phosphodiester bond is de-protonated by two
Bronsted bases, probably the ionized forms of aspartate 36 and glutamate 45.

<>

<1>Stanford, N.P., Szczelkun, M.D., Marko, J.F., Halford, S.E.
<2>One- and three-dimensional pathways for proteins to reach specific DNA sites.
<3>EMBO J.
<4>19
<5>6546-6557
<6>2000
<7>Proteins that interact with specific DNA sites bind to DNA at random and then translocate to
the target site. This may occur by one-dimensional diffusion along the DNA, or through
three-dimensional space via multiple dissociation/re-associations. To distinguish these
routes, reactions of the EcoRV endonuclease were studied on substrates with two EcoRV sites
separated by varied distances. The fraction of encounters between the DNA and the protein that
resulted in the processive cleavage of both sites decreased as the length of intervening DNA
was increased, but not in the manner demanded for one-dimensional diffusion. The variation in
processivity with inter-site spacing shows instead that protein moves from one site to another
through three-dimensional space, by successive dissociation/re-associations, though each
re-association to a new site is followed by a search of the DNA immediately adjacent to that
site.  Although DNA-binding proteins are usually thought to find their target sites by
one-dimensional pathways, three-dimensional routes may be more common than previously
anticipated.

<>

<1>Stankevicius, K., Lubys, A., Timinskas, A., Janulaitis, A.
<2>Bpu10I restriction endonuclease is composed of two nonidentical subunits.
<3>Biologija
<4>0
<5>51-53
<6>1996
<7>The genes of the Bpu10I restriction-modification (RM) system have been cloned and sequenced.
Nucleotide sequence analysis revealed four major open reading frames oriented tandemly.
Comparison of amino acid sequences, subcloning and deletion mapping experiments have been
applied to investigate all ORFs mentioned. ORFI and ORFII correspond to two
m5C-methyltransferases.  Four conserved aa blocks including the EXK motif, which is known to
be involved in the active center formation of restriction endonucleases, were detected by
comparing aa sequences of ORFIII with ORFIV.  It was demonstrated that the dsDNA could be
hydrolysed specifically only in the presence of both ORFIII and ORFIV intact, indicating that
Bpu10I Enase is composed from two heterosubunits.

<>

<1>Stankevicius, K., Lubys, A., Timinskas, A., Vaitkevicius, D., Janulaitis, A.
<2>Cloning and analysis of the four genes coding for Bpu10I restriction-modification enzymes.
<3>Nucleic Acids Res.
<4>26
<5>1084-1091
<6>1998
<7>The Bpu10I R-M system from Bacillus pumilus 10, which recognizes the asymmetric 5'-CCTNAGC
sequence, has been cloned, sequenced and expressed in Escherichia coli.  The system comprises
four adjacent, similarly oriented genes encoding two m5C MTases and two subunits of Bpu10I
ENase (34.5 and 34 kDa).  Both Bpu10IR genes either in cis or trans are needed for the
manifestation of R.Bpu10I activity.  Subunits of R.Bpu10I, purified to apparent homogeneity,
are both required for cleavage activity. This heterosubunit structure distinguishes the Bpu10I
restriction endonuclease from all other type II restriction enzymes described previously.  The
subunits reveal 25% amino acid identity.  Significant similarity was also identified between a
43 amino acid region of R.DdeI and one of the regions of higher identity shared between the
Bpu10I subunits, a region that could possibly include the catalytic/Mg2+ binding center.  The
similarity between Bpu10I and DdeI Mtases is not limited to the conserved motifs typical for
m5C MTases.  It extends into the variable region that lies between CMs VIII and IX.
Duplication of a progenitor gene, encoding an enzyme recognizing a symmetric nucleotide
sequence, followed by concerted divergent evolution, may provide a possible scenario leading
to the emergence of the Bpu10I ENase, which recognizes an overall asymmetric sequence and
cleaves within it symmetrically.

<>

<1>Stankevicius, K., Povilionis, P., Lubys, A., Menkevicius, S., Janulaitis, A.
<2>Cloning and characterization of the unusual restriction-modification system comprising two restriction endonucleases and one methyltransferase.
<3>Gene
<4>157
<5>49-53
<6>1995
<7>An Escherichia coli RFL47 DNA fragment containing the Eco47IR and Eco47II
restriction-modification (R-M) system has been cloned and sequenced.  A clone carrying this
system has been selected by its ability to restrict phage lambda in vivo.  The sequence of
5360 bp was determined, and its analysis revealed three major open reading frames (ORF)
corresponding to two restriction endonucleases (ENases) and one DNA methyltransferase (MTase):
R.Eco47II (239 amino acids (aa)), R.Eco47I (230 aa) and M.Eco47II (417 aa).  The M.Eco47II
aa  sequence possesses all conserved domains typical for m5C MTases and its variable region
has a  high homology with M.Sau96I and M.SinI.  The ORF harboring a predicted helix-turn-helix
motif  upstream from the eco47IR gene has been found.  No sequence resembling the eco47IM
gene has been detected in the complete fragment sequenced, although disrupted ORF, possibly
corresponding to the transposase-encoding gene, has been found in the intergenic area between
eco47IIM and eco47IR.  No homology was found between the ENases; however, both revealed
homology with their isoschizomers, R.SinI and R.Sau96I.

<>

<1>Stankevicius, K., Timinskas, A.
<2>Expression autoregulation of the Eco47II methyltransferase gene.
<3>Biologija
<4>0
<5>54-56
<6>1996
<7>Amino acid sequence analysis revealed a strong helix-turn-helix motif in the N-terminus of the
M.Eco47II, with a standard deviation of 7.1.  The autoregulatory mechanism of the expression
of the eco47IIM gene has been proved by using the lacZ fusion technique.  We have demonstrated
that the HTH motif of M.Eco47II is necessary for the repression of the M.Eco47II promoter but
not for the methylation of the DNA target.  Mutation in the catalytic center of M.Eco47II
strongly reduces the methylation function, but has no effect on repression.

<>

<1>Stanley, E., Walsh, L., van der Zwet, A., Fitzgerald, G.F., van Sinderen, D.
<2>Identification of four loci isolated from two Streptococcus thermophilus phage genomes responsible for mediating bacteriophage resistance.
<3>FEMS Microbiol. Lett.
<4>182
<5>271-277
<6>2000
<7>Sequence data derived from the Streptococcus thermophilus phages phiO1205 and phi7201
indicated that each of these phages contains a distinct DNA
region dedicated to replication. Southern blotting experiments showed that
phages infecting S. thermophilus may be divided into at least two groups,
each containing the presumptive replication functions of either

<>

<1>Stanley, L.K., Seidel, R., van der Scheer, C., Dekker, N.H., Szczelkun, M.D., Dekker, C.
<2>When a helicase is not a helicase: dsDNA tracking by the motor protein EcoR124l.
<3>EMBO J.
<4>25
<5>2230-2239
<6>2006
<7>Using a combination of single molecule and bulk solution measurements, we have examined the
DNA translocation activity of a helicase, the Type
I restriction modification enzyme EcoR124I. We find that EcoR124I can
translocate past covalent interstrand crosslinks, inconsistent with an
obligatory unwinding mechanism. Instead, translocation of the intact
dsDNA occurs principally via contacts to the sugar-phosphate backbone
and bases of the 30 - 50 strand; contacts to the 50 - 30 strand are not
essential for motion but do play a key role in stabilising the motor on
the DNA. A model for dsDNA translocation is presented that could be
applicable to a wide range of other enzyme complexes that are also
labelled as helicases but which do not have actual unwinding activity.

<>

<1>Stanley, L.K., Szczelkun, M.D.
<2>Direct and random routing of a molecular motor protein at a DNA junction.
<3>Nucleic Acids Res.
<4>34
<5>4387-4394
<6>2006
<7>With the aim of investigating how motor proteins negotiate DNA nanostructures, we produced
test circuits based on recombination
intermediates in which 1D translocation across a Holliday junction (HJ)
could be assessed by subsequent triplex displacement signals on each
DNA arm. Using the EcoR124I restriction-modification enzyme, a 3'-5'
double-strand DNA (dsDNA) translocase, we could show that the motor
will tend to follow its translocated strand across a junction.
Nonetheless, as the frequency of junction bypass events increases, the
motor will occasionally jump tracks.

<>

<1>Starkenburg, S.R., Larimer, F.W., Stein, L.Y., Klotz, M.G., Chain, P.S.G., Sayavedra-Soto, L.A., Poret-Peterson, A.T., Gentry, M.E., Arp, D.J., Ward, B., Bottomley, P.J.
<2>Complete genome sequence of Nitrobacter hamburgensis X14 and comparative genomic analysis of species within the genus Nitrobactei.
<3>Appl. Environ. Microbiol.
<4>74
<5>2852-2863
<6>2008
<7>The alphaproteobacterium Nitrobacter hamburgensis X14 is a gram-negative facultative
chemolithoautotroph that conserves energy
from the oxidation of nitrite to nitrate. Sequencing and analysis of
the Nitrobacter hamburgensis X14 genome revealed four replicons
comprised of one chromosome (4.4 Mbp) and three plasmids (294, 188, and
121 kbp). Over 20% of the genome is composed of pseudogenes and
paralogs. Whole-genome comparisons were conducted between N.
hamburgensis and the finished and draft genome sequences of Nitrobacter
winogradskyi and Nitrobacter sp. strain Nb-311A, respectively. Most of
the plasmid-borne genes were unique to N. hamburgensis and encode a
variety of functions (central metabolism, energy conservation,
conjugation, and heavy metal resistance), yet similar to 21 kb of a
similar to 28-kb "autotrophic" island on the largest plasmid was
conserved in the chromosomes of Nitrobacter winogradskyi Nb-255 and
Nitrobacter sp. strain Nb-311A. The N. hamburgensis chromosome also
harbors many unique genes, including those for heme-copper oxidases,
cytochrome b(561), and putative pathways for the catabolism of
aromatic, organic, and one-carbon compounds, which help verify and
extend its mixotrophic potential. A Nitrobacter "subcore" genome was
also constructed by removing homologs found in strains of the closest
evolutionary relatives, Bradyrhizobium japonicum and Rhodapseudomonas
palustris. Among the Nitrobacter subcore inventory (116 genes), copies
of genes or gene clusters for nitrite oxidoreductase (NXR), cytochromes
associated with a dissimilatory nitrite reductase (NirK), PII-like
regulators, and polysaccharide formation were identified. Many of the
subcore genes have diverged significantly from, or have origins
outside, the alphaproteobacterial lineage and may indicate some of the
unique genetic requirements for nitrite oxidation in Nitrobacter.

<>

<1>Starkenburg, S.R., Reitenga, K.G., Freitas, T., Johnson, S., Chain, P.S., Garcia-Pichel, F., Kuske, C.R.
<2>Genome of the Cyanobacterium Microcoleus vaginatus FGP-2, a Photosynthetic Ecosystem Engineer of Arid Land Soil Biocrusts Worldwide.
<3>J. Bacteriol.
<4>193
<5>4569-4570
<6>2011
<7>The filamentous cyanobacterium, Microcoleus vaginatus, is found in arid land soils worldwide.
The genome of M. vaginatus strain FGP-2 allows exploration of genes involved in
photosynthesis, desiccation tolerance, alkane production, and other features contributing to
this organism's ability function as a major component of biological soil crusts in arid
lands.

<>

<1>Starns, D., Oshima, K., Suda, W., Iino, T., Yuki, M., Inoue, J., Kitamura, K., Iida, T., Darby, A., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequence of Cytophaga fermentans JCM 21142T, a Facultative Anaerobe  Isolated from Marine Mud.
<3>Genome Announcements
<4>2
<5>e00206-14
<6>2014
<7>Cytophaga fermentans strain JCM 21142(T) is a marine-dwelling facultative anaerobe. The draft
genome sequence of this strain revealed its diverse
chemoorganotrophic potential, which makes it capable of metabolizing various
polysaccharide substrates. The genome data will facilitate further studies on its
taxonomic reclassification, its metabolism, and the mechanisms pertaining to
bacterial gliding.

<>

<1>Stecker, C., Johann, A., Herzberg, C., Averhoff, B., Gottschalk, G.
<2>Complete nucleotide sequence and genetic organization of the 210-Kilobase linear plasmid of Rhodococcus erythropolis BD2.
<3>J. Bacteriol.
<4>185
<5>5269-5274
<6>2003
<7>The complete nucleotide sequence of the linear plasmid pBD2 from Rhodococcus erythropolis BD2
comprises 210,205 bp. Sequence analyses of
pBD2 revealed 212 putative open reading frames (ORFs), 97 of which had an
annotatable function. These ORFs could be assigned to six functional
groups: plasmid replication and maintenance, transport and
metalloresistance, catabolism, transposition, regulation, and protein
modification. Many of the transposon-related sequences were found to flank
the isopropylbenzene pathway genes. This finding together with the
significant sequence similarities of the ipb genes to genes of the linear
plasmid-encoded biphenyl pathway in other rhodococci suggests that the ipb
genes were acquired via transposition events and subsequently distributed
among the rhodococci via horizontal transfer.

<>

<1>Steczkiewicz, K., Muszewska, A., Knizewski, L., Rychlewski, L., Ginalski, K.
<2>Sequence, structure and functional diversity of PD-(D/E)XK phosphodiesterase superfamily.
<3>Nucleic Acids Res.
<4>40
<5>7016-7045
<6>2012
<7>Proteins belonging to PD-(D/E)XK phosphodiesterases constitute a functionally diverse
superfamily with representatives involved in replication, restriction, DNA repair and
tRNA-intron splicing. Their malfunction in humans triggers severe diseases, such as Fanconi
anemia and Xeroderma pigmentosum. To date there have been several attempts to identify and
classify new PD-(D/E)KK phosphodiesterases using remote homology detection methods. Such
efforts are complicated, because the superfamily exhibits extreme sequence and structural
divergence. Using advanced homology detection methods supported with superfamily-wide domain
architecture and horizontal gene transfer analyses, we provide a comprehensive
reclassification of proteins containing a PD-(D/E)XK domain. The PD-(D/E)XK phosphodiesterases
span over 21 900 proteins, which can be classified into 121 groups of various families. Eleven
of them, including DUF4420, DUF3883, DUF4263, COG5482, COG1395, Tsp45I, HaeII, Eco47II, ScaI,
HpaII and Replic_Relax, are newly assigned to the PD-(D/E)XK superfamily. Some groups of
PD-(D/E)XK proteins are present in all domains of life, whereas others occur within small
numbers of organisms. We observed multiple horizontal gene transfers even between human
pathogenic bacteria or from Prokaryota to Eukaryota. Uncommon domain arrangements greatly
elaborate the PD-(D/E)XK world. These include domain architectures suggesting regulatory roles
in Eukaryotes, like stress sensing and cell-cycle regulation. Our results may inspire further
experimental studies aimed at identification of exact biological functions, specific
substrates and molecular mechanisms of reactions performed by these highly diverse proteins.

<>

<1>Steele, C.L., Donaldson, J.R., Paul, D., Banes, M.M., Arick, T., Bridges, S.M., Lawrence, M.L.
<2>Genome sequence of lineage III Listeria monocytogenes strain HCC23.
<3>J. Bacteriol.
<4>193
<5>3679-3680
<6>2011
<7>More than 98% of reported human listeriosis cases are caused by Listeria monocytogenes
serotypes within lineages I and II. Serotypes within lineage
III (4a and 4c) are commonly isolated from environmental and food
specimens. We report the first complete genome sequence of a lineage III
isolate, HCC23, which will be used for comparative analysis.

<>

<1>Steenson, L.R., Klaenhammer, T.R.
<2>Plasmid heterogeneity in Streptococcus cremoris M12R:  effects on proteolytic activity and host-dependent phage replication.
<3>J. Dairy Sci.
<4>69
<5>2227-2236
<6>1986
<7>Examination of single colony isolates from a culture of Streptococcus cremoris
M12R revealed a high degree of variability in plasmid deoxyribonucleic acid
composition.  Fifty percent of the M12R population displayed proteolytic
activity and harbored a 13-Mdalton plasmid (pLR2013).  This plasmid was not
present in proteinase-deficient variants isolated from the culture, which
provided correlative evidence for linkage of proteinase activity to pLR2013.
Four percent of the M12R population demonstrated resistance to phage m12r.M12.
This resistance was identified by restriction and modification activities
against m12r.M12 phage, which was dependent on the presence of a 20-Md plasmid,
pLR1020.  Loss of restriction and modification activities was observed upon
curing of pLR1020.  In conjugal mating studies with Streptococcus lactis ME2,
transfer frequency of lactose-fermenting ability to a restriction and
modification-deficient variant of M12R was 100-fold higher than to a variant
exhibiting restriction and modification activities.  The data provided evidence
for restriction and modification activities in select S. cremoris M12R variants
that are linked to pLR1020 and restrict both the plaquing ability of phage and
efficiency of plasmid transfer by conjugation.

<>

<1>Steenson, L.R., Klaenhammer, T.R.
<2>Streptococcus cremoris M12R transconjugants carrying the conjugal plasmid pTR2030 are insensitive to attack by lytic bacteriophages.
<3>Appl. Environ. Microbiol.
<4>50
<5>851-858
<6>1985
<7>Conjugal transfer of lactose-fermenting ability (Lac+), nisin resistance (Nisr), and phage
resistance (Hsp+) was demonstrated in mating between Streptococcus lactis ME2 (donor) and
Streptococcus cremoris M43a (recipient), a derivative of M12R. Transconjugants were detected
by transfer of Lac+ and were found to exhibit Nisr and harbor a 40-megadalton plasmid
(pTR1040). Fifty-six percent of Lac+ transconjugants were resistant to the S. cremoris M12R
lytic phage. Efficiency of plaquing for phage m12r.M12 on a phage-resistant transconjugant,
T2r-M43a, was <4.3 x 10 -10. Five additional phages which were virulent for S. cremoris M12R
and isolated from industrial sources failed to plaque on S. cremoris T2r-M43a. Mating
experiments with T2r-M43a revealed that phage resistance was accompanied by high-frequency
conjugation ability (Tra+) and the appearance of both pTR1040 and pTR2030 encoding Lac+ Nisr
and Tra+ Hsp+, respectively, in transconjugants of S. lactis mLM2302. Phage-sensitive Lac+
transconjugants of S. cremoris M43a. Unlike the S. lactis LM2302 transconjugant carrying
pTR2030, resistance of T2r-M43a to phage was not affective at high temperatures (35 to 40 deg.
C) or destabilized in repeated transfers through a starter culture activity test. These
results demonstrated that phage resistance conferred by pTR2030 in the S. cremoris
transconjugant was effective against industrially significant phages under fermentation
conditions normally encountered during cheese manufacture.

<>

<1>Steenstrup, T.D., Norby, P.L.
<2>Host cell protein knock-out cells for production of therapeutic proteins.
<3>International Patent Office
<4>WO 2007006808 A
<5>
<6>2007
<7>The present invention relates to methods and means for making Vitamin K-dependent protein
compositions which are devoid or substantially devoid of protein contaminants.  In particular,
methods and means useful for the reduction or elimination of protein contaminants also being
Vitamin K-dependent proteins are described.

<>

<1>Stefan, C., Xia, Y., Van Etten, J.L.
<2>Molecular cloning and characterization of the gene encoding the adenine methyltransferase M.CviRI from Chlorella virus XZ-6E.
<3>Nucleic Acids Res.
<4>19
<5>307-311
<6>1991
<7>The gene encoding the DNA methyltransferase M.CviRI from Chlorella virus XZ-6E
was cloned and expressed in Escherichia coli.  M.CviRI methylates adenine in
TGCA sequences.  DNA containing the M.CviRI gene was sequenced and a single
open reading frame of 1137 bp was identified which could code for a polypeptide
of 379 amino acids with a predicted molecular weight of 42,814.  Comparison of
the M.CviRI predicted amino acid sequence with another Chlorella virus and 14
bacterial adenine methyltransferases revealed extensive similarity to the other
Chlorella virus enzyme.

<>

<1>Stefanishina, T.V., Bogdarina, I.G., Buryanov, Y.I.
<2>Search for the modification-restriction systems.
<3>Biopol. Kletka
<4>3
<5>128-131
<6>1987
<7>The DNA modification-restriction systems have been studied for their presence
in 19 Agrobacteria strains, including the virulent ones with Ti-plasmids of
various classes, the avirulent octopine and nopaline strain derivatives
obtained by curing the Agrobacteria virulent strains from Ti-plasmids as well
as natural isolates of avirulent Agrobacteria.  It is shown that the presence
of the DNA modification-restriction system with EcoRII specificity is a
characteristic feature of the Agrobacteria octopine strains and of their cured
avirulent derivatives.  The Agrobacteria nopaline strains, their cured
avirulent derivatives, and most of the Agrobacteria natural avirulent strains
contain the DNA modification-restriction systems different from EcoRII.  The
chromosomal nature of these characters is shown.

<>

<1>Stegger, M., Driebe, E.M., Roe, C., Lemmer, D., Bowers, J.R., Engelthaler, D.M., Keim, P., Andersen, P.S.
<2>Genome Sequence of Staphylococcus aureus Strain CA-347, a USA600 Methicillin-Resistant Isolate.
<3>Genome Announcements
<4>1
<5>e00517-13
<6>2013
<7>The Staphylococcus aureus clonal lineage CC45 is a predominant colonizer of healthy
individuals in northern Europe and constitutes a highly basal cluster of
the S. aureus population. Here, we report the complete genome sequence of S.
aureus strain CA-347 (NRS648), a representative of the methicillin-resistant
USA600 clone predominantly found in the United States.

<>

<1>Stegger, M., Lindsay, J.A., Moodley, A., Skov, R., Broens, E.M., Guardabassi, L.
<2>Rapid PCR Detection of Staphylococcus aureus Clonal Complex 398 by Targeting the Restriction-Modification System Carrying sau1-hsdS1.
<3>J. Clin. Microbiol.
<4>49
<5>732-734
<6>2011
<7>A PCR targeting sau1-hsdS1 was developed for rapid detection of Staphylococcus aureus clonal
complex 398 (CC398). High sensitivity
(100%) and specificity (100%) were shown by evaluating the test on a
large strain collection (n = 1,307). We recommend this test for
accurate, rapid, and inexpensive diagnosis of methicillin-resistant S.
aureus (MRSA) CC398 in hospitals and on farms.

<>

<1>Stegger, M., Price, L.B., Larsen, A.R., Gillece, J.D., Waters, A.E., Skov, R., Andersen, P.S.
<2>Genome Sequence of Staphylococcus aureus Strain 11819-97, an ST80-IV European Community-Acquired Methicillin-Resistant Isolate.
<3>J. Bacteriol.
<4>194
<5>1625-1626
<6>2012
<7>The European methicillin-resistant Staphylococcus aureus (MRSA) clone ST80-IV has historically
dominated community-associated infections in major parts of Europe
and is a lineage strongly linked to skin and soft tissue infections. Here, we
report the genome sequence of an ST80-IV representative, 11819-97, isolated from
a skin infection in Denmark in 1997.

<>

<1>Steigerwald, S.D., Pfeifer, G.P., Riggs, A.D.
<2>Ligation-mediated PCR improves the sensitivity of methylation analysis by restriction enzymes and detection of specific DNA strand breaks.
<3>Nucleic Acids Res.
<4>18
<5>1435-1439
<6>1990
<7>DNA methylation at specific sites is most frequently studied by use of
methylation-sensitive restriction endonucleases and Southern blotting.  We
report here that the sensitivity of this method can be increased
several-hundred-fold by applying a ligation-mediated polymerase chain reaction
(LM-PCR) procedure followign enzyme treatment.  DNA is cleaved simultaneously
with two restriction enzymes, one sensitive and one insensitive to methylation.
After cleavage, a gene-specific oligonucleotide primer is used for primer
extension, followed by linker ligation and then conventional PCR.  Using this
technique, we demonstrate that DNA from 100 cells (about 0.6 ng) can be
prepared and qualitatively analyzed for methylation at sites in an X-linked CpG
island, and 50 ng of DNA can be analyzed quantitatively.  A site 23 bp
downstream of the major transcription start site of human phosphoglycerate
kinase-1 (PGK-1) is 52 +/- 7 percent methylated in DNA from female blood and
greater than 98 percent unmethylated in DNA from male blood or HeLa cells.
This method detects quantitatively specific breaks in either double stranded or
single stranded DNA.  Thus new assays for enzymes and DNA structure can be
devised.

<>

<1>Stein, D.C., Chien, R., Seifert, S.H.
<2>Construction of a Neisseria gonorrhoeae MS11 derivative deficient in NgoMI restriction and modification.
<3>J. Bacteriol.
<4>174
<5>4899-4906
<6>1992
<7>We have cloned from Neisseria gonorrhoeae MS11 the gene encoding a methylase that modifies the
sequence GCCGGC. The corresponding restriction enzyme was also encoded by this clone. Sequence
analysis demonstrated that the methylase shares sequence similarities with other cytosine
methylases, but the sequence organization of M.NgoMI is different from that seen for other
cytosine methylases. A deletion was introduced into the chromosome of N. gonorrhoeae MS11 to
produce strain MUG701, a strain that is inactivated in both the methylase and the restriction
genes. Although this strain no longer methylated its DNA at the NgoMI recognition sequence,
cells were viable and had no other significant phenotypic changes. Transformation data
indicated that MS11 does not produce enough restriction activity to block plasmid
transformation in the gonococcus, even though restriction activity could be demonstrated in E.
coli containing the cloned gene.
[ The enzyme called NgoMI in this abstract has been renamed NgoMIV, Jan/1998. ]

<>

<1>Stein, D.C., Gregoire, S., Piekarowicz, A.
<2>Restriction of plasmid DNA during transformation but not conjugation in Neisseria gonorrhoeae.
<3>Infect. Immun.
<4>56
<5>112-116
<6>1988
<7>Neisseria gonorrhoeae strains WR302 and PGH3-2 were characterized with respect to their
restriction-modification phenotype.  WR302 DNA was cleaved by HaeIII, indicating the lack of
methylation at the GGCC sequence.  PGH3-2 produced NgoSI (an isoschizomer of NgoII).  WR302
produced a restriction enzyme with a recognition sequence different from that of NgoI, NgoII,
or NgoIII.  Plasmid pFT180 isolated from WR302 was unable to transform PGH3-2, whereas plasmid
pFT180 isolated from PGH3-2 was able to transform PGH3-2 at a very high frequency.  When
plasmid pFT180 isolated from WR302 isolated from WR302 was methylated in vitro with methylase
M.HaeIII, this plasmid was able to transform PGH3-2.  NgoSI was able to restrict WR302 DNA in
vitro, whereas it was incapable of restricting PGH3-2 DNA in vitro.  When the
self-transmissible R factor pFT6 was mobilized from WR302 to PGH3-2 by conjugation, a
1-order-of-magnitude difference in transfer frequencies was observed, as compared with an
isogenic cross.  The data indicate that host-mediated restriction can prevent the gonococcus
from acquiring DNA via transformation but not via conjugation.
[ The enzyme called NgoSI in this abstract has been renamed NgoSII, Jan/1998. ]

<>

<1>Stein, D.C., Gunn, J.S., Piekarowicz, A.
<2>Sequence similarities between the genes encoding the S.NgoI and HaeII restriction/modification systems.
<3>Biol. Chem.
<4>379
<5>575-578
<6>1998
<7>The DNA sequence encoding the S.NgoI restriction/modification system was identified from a
gene bank made from Neisseria gonorrhoaea strain WR302 by identifying recombinant plasmids
that induced the reporter system in a methylase detection strain AP1-200-9 and were resistant
to digestion with NgoI.  The DNA sequence was determined from one of these (pUCP30).  M.NgoI
is a protein of 315aa with a predicted MW of 35,296 Da and R.NgoI is a protein of 350aa with a
predicted MW of 40,650 Da.  The termination codon of M.NgoI overlapped the start codon of
R.NgoI.  The same strategy was used to clone the R/M system encoding HaeII from Haemophilus
aegyptius strain ATCC 11116.  The DNA sequence from one clone representing this class (pAP704)
was determined.  HaeII methylase is a protein of 318aa with a predicted MW of 35,669 Da and
R.HaeII contains 352aa with a predicted MW of 40,800 Da.  Aa alignments between the two
methylases indicated that they were 74.3% identical and 79% similar. DNA sequence alignments
revealed 68% identity.  An aa alignment between the two restriction enzymes indicated that
they were 60% identical and 68% similar.  DNA sequence alignments revealed 61% identity.  The
DNA sequences flanking these two systems were identified and used to determine the genomic
organization of the two systems.  The S.NgoI genes were found between two genes, one with high
homology to GTP binding proteins of unknown function and one with homology to genes involved
in tRNA synthetase synthesis.  The HaeII R/M genes were located between two genes, mucF and
mucE.  The DNA sequence of the HaeII R/M system was compared to the genomic DNA sequence of H.
influenzae Rd. Although the DNA sequences flanking the HaeII system were >99% identical to
contiguous DNA fragments found in the genome of H. influenzae Rd, no homology was seen with
the DNA sequences encoding the HaeII R/M system, indicating that it is not found in this
strain.  Given the vast difference in the GC content of S.NgoI and HaeII, their apparent
insertion into polycistronic operons, and their difference in codon usage when compared to the
species from which they were isolated, the data suggest that these R/M systems originated in
an organism other than Neisseria or Haemophilus.

<>

<1>Stein, D.C., Gunn, J.S., Radlinska, M., Piekarowicz, A.
<2>Restriction and modification systems of Neisseria gonorrhoeae.
<3>Gene
<4>157
<5>19-22
<6>1995
<7>An individual strain of Neisseria gonorrhoeae may produce up to 16 different DNA
methyltransferases (MTases).  We have used a novel cloning system that is able to detect MTase
clones in the absence of direct selection to identify 14 different MTase clones.  Initial
characterization of these clones indicates that at least seven of these MTases are linked to
restriction endonuclease (ENase) systems.  Six of these systems have been characterized by DNA
sequence analysis, and the open reading frames encoding each of these systems have been
identified.  The recognition sequences for the cloned systems have the following
specificities: S.NgoI, RGCGCY; S.NgoII, GGCC; S.NgoIV, GCCGCC; S.NgoV, GGNNCC; S.NgoVII,
GCSGC; S.NgoVIIIA, GGTGA; and S.NgoVIIIC, TCACC.  Of those systems that have been cloned,
NgoI-NgoVII are typical type II R-M systems, with each encoding a DNA MTase that methylates
cytosine in position 5.  NgoVIII is a type IIS system, containing an ENase and two different
MTases.  One of these is a cytosine MTase (NgoVIIIC) and the other is an adenine MTase
(NgoVIIIA).  Although most of our clones encodes both the ENase and the MTase, none of the six
R-M systems are genetically linked on the chromosome.

<>

<1>Stein, D.C., Piekarowicz, A.S., Yuan, R.T.
<2>Mutant strain of E. coli for detection of methyltransferase clones.
<3>US Patent Office
<4>US 5491060
<5>
<6>1996
<7>E. coli bacterial strains encoding a restriction gene that degrades methylated DNA, and
prevents cloning of genes expressing the methyltransferase responsible for methylation, are
mutated by a chemical or physical mutagen, so as to make the restriction enzyme temperature
sensitive.  Mutant cells are rendered competent, and plasmids expected or known to contain
genes encoding methyltransferase enzymes are introduced.  The transformants grow at the
permissive temperature, where the restriction enzyme system is inactivated due to the mutated
gene. Successful clones, expressing a methyltransferase, can be quickly identified by those
which grow at the permissive temperature, but not at the non-permissive temperature.  The
valuable methyltransferases, as well as restriction enzymes associated therewith, can
accordingly be recovered in large quantity.

<>

<1>Stein, L.Y. et al.
<2>Genome sequence of the methanotrophic Alphaproteobacterium, Methylocystis sp. Rockwell (ATCC 49242).
<3>J. Bacteriol.
<4>193
<5>2668-2669
<6>2011
<7>Methylocystis sp. strain Rockwell (ATCC 49242) is an aerobic methane-oxidizing
Alphaproteobacterium isolated from an aquifer in southern California. Unlike most
methanotrophs in the Methylocystaceae family, this strain has a single pmo operon encoding
particulate methane monooxygenase but no evidence of the genes encoding soluble methane
monooxygenase. This is the first reported genome sequence of a member of the Methylocystis
species of the Methylocystaceae family in the order Rhizobiales.

<>

<1>Stein, L.Y. et al.
<2>Genome Sequence of the Obligate Methanotroph Methylosinus trichosporium Strain OB3b.
<3>J. Bacteriol.
<4>192
<5>6497-6498
<6>2010
<7>Methylosinus trichosporium OB3b (for 'oddball' strain 3b) is an obligate aerobic
methane-oxidizing alphaproteobacterium that was originally
isolated in 1970 by Roger Whittenbury and colleagues. This strain has
since been used extensively to elucidate the structure and function of
several key enzymes of methane oxidation, including both particulate and
soluble methane monooxygenase (sMMO) and the extracellular copper chelator
methanobactin. In particular, the catalytic properties of soluble methane
monooxygenase from M. trichosporium OB3b have been well characterized in
context with biodegradation of recalcitrant hydrocarbons, such as
trichloroethylene. The sequence of the M. trichosporium OB3b genome is the
first reported from a member of the Methylocystaceae family in the order
Rhizobiales.

<>

<1>Steinweg, C., Kuenne, C.T., Billion, A., Mraheil, M.A., Domann, E., Ghai, R., Barbuddhe, S.B., Karst, U., Goesmann, A., Puhler, A., Weisshaar, B., Wehland, J., Lampidis, R., Kreft, J., Goebel, W., Chakraborty, T., Hain, T.
<2>The complete genome sequence of L. seeligeri, a non-pathogenic member of the genus Listeria.
<3>J. Bacteriol.
<4>192
<5>1473-1474
<6>2010
<7>We report the complete and annotated genome sequence of the non-pathogenic L. seeligeri
SLCC3954 serovar 1/2b type strain harboring the smallest completely sequenced genome of the
genus Listeria.

<>

<1>Steitz, T.A.
<2>Structural studies of protein-nucleic acid interaction:  the source of sequence-specific binding.
<3>Q. Rev. Biophys.
<4>23
<5>205-280
<6>1990
<7>1. Introduction and overview
2. Principles of sequence-specific nucleic-acid recognition
2.1 The problem that is set: what is being recognized?
2.2 Role of the major groove in DNA recognition
2.3 Role of nucleic acid bendability
2.4 Role of water molecules in sequence recognition
2.5 Role of the minor groove in DNA and RNA recognition
3. DNA-binding structure motifs
3.1 Helix-turn-helix
3.2 Zinc fingers
3.3 Helices of the dinucleotide fold
3.4 other motifs
4. Similarities and differences in RNA and DNA recognition
5. Sequence-specific DNA-binding proteins
5.1 Repressors and activators
5.1.1 Lac operon regulation
(a) E. coli catabolite gene activator protein (CAP)
(b) E. coli lac repressor protein
5.1.2 Structural studies of the bacterial phage repressors
(a) 434 Repressor fragment complexed with DNA
(b) Lambda cro
(c) Lambda cI repressor fragment
5.1.3 E. coli Trp repressor
5.1.4 E. coli Met repressor
5.1.5 Zinc-containing DNA-binding domains (zinc fingers) TFIIIA
5.2 Restriction endonuclease - E. coli EcoRI
5.3 Site-specific recombination-Gamma Delta Resolvase
6. Sequence-independent DNA-binding proteins
6.1 Klenow fragment of E. coli DNA Polymerase I
6.2 Bovine pancreatic DNase I
6.3 E. coli Hu protein
7. Sequence-specific RNA-binding proteins
7.1 E. coli glutaminyl-tRNA synthetase complexed with tRNA
8. Conclusions and prospects
9. Acknowledgements
10. References

<>

<1>Stelling, S.C., Techtmann, S.M., Utturkar, S.M., Alshibli, N.K., Brown, S.D., Hazen, T.C.
<2>Draft Genome Sequence of Thalassotalea sp. Strain ND16A Isolated from Eastern Mediterranean Sea Water Collected from a Depth of 1,055 Meters.
<3>Genome Announcements
<4>2
<5>e01231-14
<6>2014
<7>Thalassotalea sp. strain ND16A belongs to the family Colwelliaceae and was isolated from
eastern Mediterranean Sea water at a depth of 1,055 m. Members of
Colwelliaceae are ubiquitous marine heterotrophs. Here, we report the draft
genome sequence of Thalassotalea sp. strain ND16A, a member of the newly
described genus Thalassotalea.

<>

<1>Stenger, D.C., Lee, M.W., Rogers, E.E., Chen, J.
<2>Plasmids of Xylella fastidiosa mulberry-infecting strains share extensive sequence identity and gene complement with pVEIS01 from the earthworm symbiont Verminephrobacter eiseniae.
<3>Physiol. Mol. Plant Pathol.
<4>74
<5>238-245
<6>2010
<7>A ~25 kbp plasmid was present in each of four Californian strains of Xylella fastidiosa from
mulberry affected with leaf scorch disease.  Fragments of each plasmid were cloned into
Escherichia coli, sequenced, and assembled into circular contigs of 25,105 bp (pXF-RIV11 and
pXF-RIV16) or 24,372 bp (pXF-RIV19 and pXF-RIV25).  The four plasmids shared >99.8% sequence
identity, excluding a 732 bp insertion common to pXF-RIV11 and pXF-RIV16.  BLAST searches
identified seven regions (totaling 19,252 bp) sharing greater than or equal to 75% nucleotide
sequence identity with pVEI201, a 31 kbp plasmid from the earthworm symbiont Verminephrobacter
eiseniae.  Using pXF-RIV11 as a query in BLASTX searches, putative functions of
plasmid-encoded open reading frames were identified.  Fourteen ORFs were associated with DNA
transfer (Type IV secretion), four with plasmid stability (plasmid toxin/anti-toxin
addiction), one with protein export (Type II secretion), one with plasmid DNA replication
initiation (trfA), and the remaining ORFs associated with other or unknown functions.  The
putative origin of DNA replication (oriV) was located adjacent to the trfA ORF and was similar
in structure to that of plasmids belonging to the incP-1 incompatibility group.  E. coli
plasmids bearing fragments of pXF-RIV11 and the nptII gene as a selectable marker were tested
for replication in X. fastidiosa strain Temecula1.  Only fragments bearing oriV and trfA were
competent for replication in X. fastidiosa.  Collectively, these results indicate that
mulberry strains of X. fastidiosa harbor plasmids encoding genes associated with DNA transfer
and plasmid stability not previously identified on the chromosome of sequenced X. fastidiosa
strains and that ancestors of distantly related bacterial species occupying different niches
appear to have exchanged genetic material.

<>

<1>Stepanov, V.G., Vaishampayan, P., Venkateswaran, K., Fox, G.E.
<2>Draft Genome Sequence of Deinococcus phoenicis, a Novel Strain Isolated during the Phoenix Lander Spacecraft Assembly.
<3>Genome Announcements
<4>2
<5>e00301-14
<6>2014
<7>Deinococcus phoenicis strain 1P10ME(T) is a radiation- and desiccation-resistant  bacterium
isolated from a cleanroom facility where the Phoenix Lander spacecraft was assembled. In order
to facilitate investigations of the nature of the extreme resistance of D. phoenicis to
bactericidal factors, a draft genome sequence of D. phoenicis was determined.

<>

<1>Stephan, R., Grim, C.J., Gopinath, G.R., Mammel, M.K., Sathyamoorthy, V., Trach, L.H., Chase, H.R., Fanning, S., Tall, B.D.
<2>Genome Sequence of Enterobacter turicensis Strain 610/05 (LMG 23731), Isolated from Fruit Powder.
<3>Genome Announcements
<4>1
<5>e00996-13
<6>2013
<7>We report the draft genome sequence of Enterobacter turicensis strain 610/05 (LMG 23731),
isolated from fruit powder. The draft genome has a size of 4,182,790 bp
and a G+C% content of 58.0.

<>

<1>Stephan, R., Lehner, A., Tischler, P., Rattei, T.
<2>Complete Genome Sequence of Cronobacter turicensis LMG 23827, a foodborne pathogen causing deaths in neonates.
<3>J. Bacteriol.
<4>193
<5>309-310
<6>2010
<7>Here we report the complete and annotated genome sequence of Cronobacter turicensis, an
opportunistic foodborne pathogen, which is known as rare but important causes of
live-threatening neonatal infections. Among all proteins of C. turicensis, 223 have been
annotated as virulence and disease-related proteins.

<>

<1>Stephanou, A.S., Roberts, G.A., Cooper, L.P., Clarke, D.J., Thomson, A.R., MacKay, C.L., Nutley, M., Cooper, A., Dryden, D.T.
<2>Dissection of the DNA mimicry of the bacteriophage T7 Ocr protein using chemical modification.
<3>J. Mol. Biol.
<4>391
<5>565-576
<6>2009
<7>The homodimeric Ocr (overcome classical restriction) protein of bacteriophage T7 is a
molecular mimic of double-stranded DNA and a highly
effective competitive inhibitor of the bacterial type I
restriction/modification system. The surface of Ocr is replete with acidic
residues that mimic the phosphate backbone of DNA. In addition, Ocr also
mimics the overall dimensions of a bent 24-bp DNA molecule. In this study,
we attempted to delineate these two mechanisms of DNA mimicry by
chemically modifying the negative charges on the Ocr surface. Our analysis
reveals that removal of about 46% of the carboxylate groups per Ocr
monomer results in an approximately 50-fold reduction in binding affinity
for a methyltransferase from a model type I restriction/modification
system. The reduced affinity between Ocr with this degree of modification
and the methyltransferase is comparable with the affinity of DNA for the
methyltransferase. Additional modification to remove approximately 86% of
the carboxylate groups further reduces its binding affinity, although the
modified Ocr still binds to the methyltransferase via a mechanism
attributable to the shape mimicry of a bent DNA molecule. Our results show
that the electrostatic mimicry of Ocr increases the binding affinity for
its target enzyme by up to approximately 800-fold.

<>

<1>Stephanou, A.S., Roberts, G.A., Tock, M.R., Pritchard, E.H., Turkington, R., Nutley, M., Cooper, A., Dryden, D.T.F.
<2>A mutational analysis of DNA mimicry by ocr, the gene 0.3 antirestriction protein of bacteriophage T7.
<3>Biochem. Biophys. Res. Commun.
<4>378
<5>129-132
<6>2009
<7>The ocr protein of bacteriophage T7 is a structural and electrostatic mimic of approximately
24 base pairs of double-stranded B-form DNA. As
such, it inhibits all Type I restriction and modification (R/M) enzymes
by blocking their DNA binding grooves and inactivates them. This allows
the infection of the bacterial cell by T7 to proceed unhindered by the
action of the R/M defence system. We have mutated aspartate and
glutamate residues on the surface of ocr to investigate their
contribution to the tight binding between the EcoKI Type I R/M enzyme
and ocr. Contrary to expectations, all of the single and double site
mutations of ocr constructed were active as anti-R/M proteins in vivo
and in vitro indicating that the mimicry of DNA by ocr is very
resistant to change.

<>

<1>Stephens, C., Reisenauer, A., Wright, R., Shapiro, L.
<2>A cell cycle-regulated bacterial DNA methyltransferase is essential for viability.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>1210-1214
<6>1996
<7>The CcrM adenine DNA methyltransferase, which specifically modifies GANTC sequences, is
necessary for viability in Caulobacter crescentus.  To our knowledge, this is the first
example of an essential prokaryotic DNA methyltransferase that is not part of a DNA
restriction/modification system.  Homologs of CcrM are widespread in the a subdivision of the
Proteobacteria, suggesting that methylation at GANTC sites may have important functions in
other members of this diverse group as well.  Temporal control of DNA methylation state has an
important role in Caulobacter development, and we show that this organism utilizes an unusual
mechanism for control of remethylation of newly replicated DNA.  CcrM is synthesized de novo
late in the cell cycle, coincident with full methylation of the chromosome, and is then
subjected to proteolysis prior to cell division.

<>

<1>Stephens, C., Shapiro, L.
<2>A common regulatory system controls transcription of flagellar genes and an essential DNA methyltransferase in Caulobacter.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>203
<6>1994
<7>Our lab has recently isolated an essential Caulobacter gene (ccrM) encoding a DNA
methyltransferase which modifies DNA in a cell-cycle dependent manner.  ccrM transcription,
and CcrM methylation activity, occur only in the predivisional cell as chromosomal replication
is nearing completion.  Proper temporal control of CcrM activity is crucial, as constitutive
expression of ccrM results in defects in cell morphology and timing of chromosomal
replication.  We are thus interested in understanding the control of ccrM transcription.  The
functional ccrM promoter region was defined by deletion analysis, and the start site of
transcription was determined.  Immediately upstream of the start site is a sequence with
striking similarity to the consensus promoter for Caulobacter Class II flagellar genes, which
are also activated in the predivisional cell.  The RNA polymerase species recognizing these
promoters is not yet known.  Mutations in bases conserved between the ccrM and Class II
promoters greatly reduce ccrM transcription.  In addition, ccrM and Class II promoters are
rapidly repressed when DNA synthesis is inhibited.  It is proposed, therefore, that ccrM is
transcribed by the same factors used for Class II flagellar genes.  Deletion analysis
indicated that 20 bp downstream of the transcription start site was also required for ccrM
promoter activity.  This region contains a 10 bp inverted repeat containing two CcrM
methylation sites.  Roles for this sequence, and perhaps its methylation state, in promoter
activation are being examined.

<>

<1>Stephens, C., Shapiro, L.
<2>An essential DNA methyltransferase in Caulobacter crescentus.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>95
<5>330
<6>1995
<7>In the dimorphic bacterium Caulobacter crescentus, methylation of GAnTC sites in newly
replicated DNA only occurs late in the cell cycle, shortly before cell division.  The
Caulobacter ccM gene, which encodes the DNA methyltransferase (M.CcrII) responsible for
methylation of GAnTC sites, is transcribed only in the predivisional cell.  We have found that
ccrM expression depends on a promoter homologous to that used by Caulobacter Class II
flagellar genes, demonstrating that the system controlling the flagellar genetic hierarchy
also regulates other genes in the predivisional cell.  Antibodies generated to M.CcrII were
used to show that this protein is highly unstable, and M.CcrII levels during the cell cycle
are tightly linked to transcription of ccrM.  Temporal regulation of methylation by M.CcrII is
necessary for normal development, as strains which express ccrM throughout the cell cycle,
and which therefore have continuously fully methylated DNA, exhibit abnormalities in control
of DNA replication and cell division.  We have used gene replacement experiments to
demonstrate that the ccrM gene is essential for growth and viability.  ccrM is to our
knowledge the first essential prokaryotic DNA methyltransferase.  To further examine the
function of M.CcrII in Caulobacter growth and development, we have constructed strains in
which ccrM expression can be controlled exogeneously.  We are examining the pysiological
consequences of blocking methylation completely, and of inducing methylation at inappropriate
times in the cell cycle.

<>

<1>Stephens, C.M., Zweiger, G., Shapiro, L.
<2>Coordinate cell cycle control of a Caulobacter DNA methyltransferase and the flagellar genetic hierarchy.
<3>J. Bacteriol.
<4>177
<5>1662-1669
<6>1995
<7>The expression of the Caulobacter ccrM gene and the activity of its product, the M.CcrII DNA
methyltransferase, are limited to a discrete portion of the cell cycle. Temporal control of
DNA methylation has been shown to be critical for normal development in the dimorphic
Caulobacter life cycle. To understand the mechanism by which ccrM expression is regulated
during the cell cycle, we have identified and characterized the ccrM promoter region. We have
found that it belongs to an unusual promoter family used by several Caulobacter class II
flagellar genes. The expression of these class II genes initiates assembly of the flagellum
just prior to activation of the ccrM promoter in the predivisional cell. Mutational analysis
of two M.CcrII methylation sites located 3' to the ccrM promoter suggests that methylation
might influence the temporally controlled inactivation of ccrM transcription. An additional
parallel between the ccrM and class II flagellar promoters is that their transcription
responds to a cell cycle DNA replication checkpoint. We propose that a common regulatory
system coordinates the expression of functionally diverse genes during the Caulobacter cell
cycle.

<>

<1>Stephens, M.A.
<2>Partial purification and cleavage specificity of a site-specific endonuclease, SciNI, isolated from Spiroplasma citri.
<3>J. Bacteriol.
<4>149
<5>508-514
<6>1982
<7>A site-specific endonuclease, SciNI, has been partially purified from the plant
pathogen Spiroplasma citri.  The enzyme recognizes the sequence 5'-G-C-G-C-3'
and cleaves between the first G and C.  3'-C-G-C-G-5' SciNI is an isoschizomer
of HhaI, but generates DNA fragments with 5' rather than 3' single-stranded
protrusions.

<>

<1>Stephens, R.S., Kalman, S., Lammel, C.J., Fan, J., Marathe, R., Aravind, L., Mitchell, W.P., Olinger, L., Tatusov, R.L., Zhao, Q., Koonin, E.V., Davis, R.W.
<2>Genome Sequence of an Obligate Intracellular Pathogen of Humans: Chlamydia trachomatis.
<3>Science
<4>282
<5>754-759
<6>1998
<7>Analysis of the 1,042,519-base pair Chlamydia trachomatis genome revealed unexpected features
related to the complex biology of chlamydiae. Although chlamydiae lack many biosynthetic
capabilities, they retain functions for performing key steps and interconversions of
metabolites obtained from their mammalian host cells. Numerous potential virulence-associated
proteins also were characterized. Several eukaryotic chromatin-associated domain proteins were
identified, suggesting a eukaryotic-like mechanism for chlamydial nucleoid condensation and
decondensation. The phylogenetic mosaic of chlamydial genes, including a large number of genes
with phylogenetic origins from eukaryotes, implies a complex evolution for adaptation to
obligate intracellular parasitism.  [ Comment in: Science 1998 Oct 23;282(5389):638-9 ]

<>

<1>Stephenson, F.H., Ballard, B.T., Boyer, H.W., Rosenberg, J.M., Greene, P.J.
<2>Comparison of the nucleotide and amino acid sequences of the RsrI and EcoRI restriction endonucleases.
<3>Gene
<4>85
<5>1-13
<6>1989
<7>The RsrI endonuclease, a type-II restriction endonuclease (ENase) found in
Rhodobacter sphaeroides, is an isoschizomer of the EcoRI ENase.  A clone
containing an 11-kb BamHI fragment was isolated from an R. sphaeroides genomic
DNA library by hybridization with synthetic oligodeoxyribonucleotide probes
based on the N-terminal amino acid (aa) sequence of RsrI.  Extracts of E. coli
containing a subclone of the 11-kb fragment display RsrI activity.  Nucleotide
sequence analysis reveals an 831-bp open reading frame encoding a polypeptide
of 277 aa.  A 50% identity exists within a 266-aa overlap between the deduced
aa sequences of RsrI and EcoRI.  Regions of 75-100% aa sequence identity
correspond to key structural and functional regions of EcoRI.  The type-II
ENases have many common properties, and a common origin might have been
expected.  Nevertheless, this is the first demonstration of aa sequence
similarity between ENases produced by different organisms.

<>

<1>Stephenson, F.H., Greene, P.J.
<2>Nucleotide sequence of the gene encoding the RsrI methyltransferase.
<3>Nucleic Acids Res.
<4>17
<5>10503
<6>1989
<7>None

<>

<1>Steponaviciene, D., Maneliene, Z., Petrusyte, M., Janulaitis, A.
<2>BseSI, a restriction endonuclease from Bacillus stearothermophilus Jo 10-553, which recognizes the novel hexanucleotide sequence 5'-G(G/T)GC(A/C)/C-3'.
<3>Nucleic Acids Res.
<4>27
<5>2644-2645
<6>1999
<7>A new restriction endonuclease BseSI has been isolated from Bacillus stearothermophilus
Jo10-553. BseSI recognizes a degenerate hexanucleotide sequence 5'-G(G/T)GC(A/C)^C-3' and
cleaves DNA to produce 3'-protruding tetranucleotide ends.

<>

<1>Stern, A., Sorek, R.
<2>The phage-host arms race: Shaping the evolution of microbes.
<3>Bioessays
<4>33
<5>43-51
<6>2011
<7>Bacteria, the most abundant organisms on the planet, are outnumbered by a factor of 10 to 1 by
phages that infect them. Faced with the rapid evolution and turnover of phage particles,
bacteria have evolved various mechanisms to evade phage infection and killing, leading to an
evolutionary arms race. The extensive co-evolution of both phage and host has resulted in
considerable diversity on the part of both bacterial and phage defensive and offensive
strategies. Here, we discuss the unique and common features of phage resistance mechanisms and
their role in global biodiversity. The commonalities between defense mechanisms suggest
avenues for the discovery of novel forms of these mechanisms based on their evolutionary
traits.

<>

<1>Sternberg, N.
<2>Evidence that adenine methylation influences DNA-protein interactions in Escherichia coli.
<3>J. Bacteriol.
<4>164
<5>490-493
<6>1985
<7>In this review, most of the information presented will be derived from studies
with Escherichia coli K-12 (E. coli) and its related bacteriophages, simply
because more is known about methylation in these organisms than in any other.
The two methylated bases that have been detected in E. coli are 6-methyladenine
(6-meAde) and 5-methylcytosine.  Since little is known about the biological
function of 6-methylcytosine, I will deal exclusively here with the studies on
6-meAde.

<>

<1>Sternberg, N., Coulby, J.
<2>Cleavage of the bacteriophage P1 packaging site (pac) is regulated by adenine methylation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>87
<5>8070-8074
<6>1990
<7>The packaging of bacteriophage P1 DNA is initiated when the phage packaging
site (pac) is recognized and cleaved and continues until the phage head is
full.  We have previously shown that pac is a 162-base-pair segment of P1 DNA
that contains seven DNA adenine methyltransferase methylation sites (5'-GATC).
We show here that cleavage of pac is methylation sensitive.  Both in vivo and
in vitro experiments indicate that methylated pac is cleavable, whereas
unmethylated pac is not.  Moreover, DNA isolated from P1 phage and containing
an uncut pac site was a poor substrate for in vitro cleavage until it was
methylated by the Escherichia coli DNA adenine methyltransferase.  Comparison
of that uncut pac DNA with other viral DNA fragments by digestion with
methylation-sensitive restriction enzymes indicated that the uncut pac DNA was
preferentially undermethylated.  In contrast, virion DNA containing a cut pac
site was not undermethylated.  We believe these results indicate that pac
cleavage is regulated by adenine methylation during the phage lytic cycle.

<>

<1>Sternglanz, H., Bugg, C.E.
<2>Conformation of N6-methyladenine, a base involved in DNA modification:restriction processes.
<3>Science
<4>182
<5>833-834
<6>1973
<7>Crystal structures of N6,N9-dimethyladenine and N6-methyladenine hydrochloride
were determined from three-dimensional x-ray diffraction data.  The bases
assume a conformation in which the N(6)-methyl group blocks one of the
hydrogen-bonding sites normally used by adenine to form Watson-Crick pairs with
thymine in double-helical DNA.  When in this conformation, N6-methyladenine
residues might alter the secondary structure of DNA, thereby preventing the
scission of modified DNA's by restriction enzymes.

<>

<1>Stetter, K.O., Schmitt, R.
<2>New type II restriction endonuclease MaeII from Methanococcus aeolicus, recognising the sequence 5-ACGT-3.
<3>German Patent Office
<4>DE 3401619 A
<5>
<6>1985
<7>Restriction endonuclease, designated MaeII, which recognises the sequence A^CGT and cuts it as
indicated, is new. MaeII is isolated from Methanococcus aeolicus DSM 2835. The cells are
grown anaerobically on H2/CO2 or formate, with optimum growth at 45 degrees C. in presence of
4% NaCl. The cells are lysed and the extract treated with polyethylenimine (PEI) to a
concentration of 0.65%. Insolubles were removed and the supernatant treated with (NH4)2SO4 to
90% saturation. The resulting precipitate can be further refined by molecular sieve
fractionation; chromatography on weakly basic and acidic ion-exchangers, then affinity
chromatography, preferably on immobilised heparin.

<>

<1>Stetter, K.O., Schmitt, R.
<2>Type II restriction endonuclease MaeI, a process for obtaining it and the use thereof.
<3>US Patent Office
<4>US 4693978
<5>1468
<6>1987
<7>The restriction endonuclease, designated MaeI, which recognises the sequence
(I) and cuts it along the line indicated, is new.  MaeI is isolated from
Methanococcus aeolicus DSM 2835.  The cells are grown anaerobically on H2/CO2
or formate, with optimum growth at 45C. in presence of 4% NaCl.  The cells are
lysed and the extract treated with polyethyleneimine (PEI) to a concentration
of 0.65%.  Insolubles are removed and the supernatant treated with (NH4)2S04 to
60% satn.  The resulting ppte. can be futher refined by molecular sieve
fractionation; chromatography on weakly basic and acidic ion exchangers, then
affinity chromatography, pref. on immobilised heparin.  USE - MaeI is used to
cut DNA at the sequence 5'-C-T-A-G-3', e.g. for DNA analysis.  No enzyme which
recognised this sequence has previously been available.

<>

<1>Stetter, K.O., Schmitt, R.
<2>New typeII restriction endonuclease MaeII from Methanococcus aeolicus.
<3>US Patent Office
<4>US 4693979
<5>1468
<6>1987
<7>Restriction endonuclease, designated MaeII, which recognises the sequence (I)
and cuts it along the line indicated, is new.  MaeII is isolated from
Methanococcus aeolicus DSM2835.  The cells are grown anaerobically on H2/C02 or
formate, with optimum growth at 45C in presence of 4% NaCl.  The cells are
lysed and the extract treated with polyethylenimine (PEI) to a concentration of
0.65%.  Insolubles were removed and the supernatant treated with (NH4) 2S04 to
90% satn.  The resulting ppte. can be further refined by molecular sieve
fractionation; chromatography on weakly basic and acidic ion-exchangers, then
affinity chromatography, pref. on immobilised heparin.  USE - MaeII is used to
cut DNA at the sequence 5'-A-C-G-T-3', e.g. for DNA analysis.  No enzyme which
recognised this sequence has previously been available.

<>

<1>Stetter, K.O., Schmitt, R.
<2>New type II restriction endonuclease MaeI from Methanococcus aeolicus, recognising the sequence 5-CTAG-3.
<3>German Patent Office
<4>DE 3401617 A
<5>
<6>1985
<7>The restriction endonuclease, designated MaeI, which recognises the sequence C^TAG and cuts it
as indicated, is new. MaeI is isolated from Methanococcus aeolicus DSM 2835. The cells are
grown anaerobically on H2/CO2 or formate, with optimum growth at 45 degrees C. in presence of
4% NaCl. The cells are lysed and the extract treated with polyethyleneimine (PEI) to a
concentration of 0.65%. Insolubles are removed and the supernatant treated with (NH4)2SO4 to
60% saturation. The resulting pecipitate can be further refined by molecular sieve
fractionation; chromatography on weakly basic and acidic ionexchangers, then affinity
chromatography, preferably on immobilised heparin.

<>

<1>Stetter, K.O., Schmitt, R., Laue, F.
<2>New type restriction endonuclease MaeIII from Methanococcus aeolicus, recognising the sequence 5-GTNAC-3.
<3>US Patent Office
<4>US 4693980
<5>1468-1469
<6>1987
<7>The restriction endonuclease, designated MaeIII, which recognises the sequence
(I) and cuts it along the line indicated, is new.  MaeIII is isolated from
Methanococcus aeolicus DSM2835.  The cells are grown anaerobically on H2/CO2 or
formate, with optimum growth at 45C in presence of 4% NaCl.  The cells are
lysed and the extract treated with polyethyleneimine (PEI) to a concentration
of 0.65%.  Insolubles were removed and a fraction recovered from the
supernatant at 60-95% (NH4)2S04 satn..  The resulting ppte. can be further
refined by molecular sieve fractionation; chromatography on weakly basic and
acidic ion-exchangers, then affinity chromatography, pref. on immobilised
heparin.

<>

<1>Stetter, K.O., Schmitt, R., Laue, F.
<2>New type restriction endonuclease MaeIII from Methanococcus aeolicus, recognising the sequence 5-GTNAC-3.
<3>German Patent Office
<4>DE 3401620 A
<5>
<6>1985
<7>The restriction endonuclease, designated MaeIII, which recognises the sequence ^GTNAC and cuts
it as indicated, is new. MaeIII is isolated from Methanococcus aeolicus DSM 2835. The cells
are grown anaerobically on H2/CO2 or formate, with optimum growth at 45 degrees C. in the
presence of 4% NaCl. The cells are lysed and the extract treated with polyethyleneimine (PEI)
to a concentration of 0.65%. Insolubles were removed and a fraction recovered from the
supernatant at 60-95% (NH4)2SO4 saturation. The resulting precipitate can be further refined
by molecular sieve fractionation; chromatography on weakly basic and acidic ion-exchangers,
then affinity chromatography, preferably on immobilised heparin.

<>

<1>Steuer, S., Pingoud, V., Pingoud, A., Wende, W.
<2>Chimeras of the homing endonuclease PI-Scel and the homologous Candida tropicalis Intein: A study to explore the possibility of exchanging  DNA-binding modules to obtain highly specific endonucleases with  altered specificity.
<3>Chembiochem
<4>5
<5>206-213
<6>2004
<7>Homing endonucleases are extremely specific endodeoxyribonucleases. In vivo, these enzymes
confer mobility on their genes by inducing a very
specific double-strand cut in cognate alleles that lack the cooling
sequence for the homing endonuclease; the cellular repair of the
double-strand break with the endonuclease-containing allele as a
template leads to integration of the endonuclease gene, completing the
homing process. As a result of their extreme sequence specificity
homing endonucleases are promising tools for genome engineering, For
this, purpose, it is desirable to design enzymes with defined new
specificities. To analyse which DNA-binding elements are potential
candidates for use in the design of enzymes with modified or even new
specificity, we produced several chimeric proteins derived from the
Saccharomyces cerevisiae VMA1 intein (PI-SceI) and the related Candida
tropicalis VMA1 intein. Although the mature Candida intein is devoid of
endonucleolytic activity the exchange of two DNA-binding modules of
PI-Scel with the homologous elements from the Candida intein results in
an active endonuclease. The low sequence homology in these modules
indicates that different protein - DNA contacts are responsible for the
recognition of related DNA sequences. This flexibility in DNA
recognition should, in principle, allow endonucleases to be produced
with new specificities useful for genome engineering.

<>

<1>Stevens, D.C., Young, J., Carmichael, R., Tan, J., Taylor, R.E.
<2>Draft Genome Sequence of Gephyronic Acid Producer Cystobacter violaceus Strain Cb vi76.
<3>Genome Announcements
<4>2
<5>e01299-14
<6>2014
<7>A draft genome sequence of Cystobacter violaceus strain Cb vi76, which produces the eukaryotic
protein synthesis inhibitor gephyronic acid, has been obtained.
The genome contains numerous predicted secondary metabolite clusters, including
the gephyronic acid biosynthetic pathway. This genome will contribute to the
investigation of secondary metabolism in other Cystobacter strains.

<>

<1>Stevens, M., Cheng, J.B., Li, D., Xie, M., Hong, C., Maire, C.L., Ligon, K.L., Hirst, M., Marra, M.A., Costello, J.F., Wang, T.
<2>Estimating absolute methylation levels at single-CpG resolution from methylation enrichment and restriction enzyme sequencing methods.
<3>Genome Res.
<4>23
<5>1541-1553
<6>2013
<7>Recent advancements in sequencing-based DNA methylation profiling methods provide an
unprecedented opportunity to map complete DNA
methylomes. These include whole-genome bisulfite sequencing (WGBS,
MethyiC-seq, or BS-seq), reduced-representation bisulfite sequencing
(RRBS), and enrichment-based methods such as MeDIP-seq, MBD-seq, and
MRE-seq. These methods yield largely comparable results but differ
significantly in extent of genomic CpG coverage, resolution,
quantitative accuracy, and cost, at least while using current
algorithms to interrogate the data. None of these existing methods
provides single-CpG resolution, comprehensive genome-wide coverage, and
cost feasibility for a typical laboratory. We introduce methylCRF, a
novel conditional random fields based algorithm that integrates
methylated DNA immunoprecipitation (MeDIP-seq) and
methylation-sensitive restriction enzyme (MRE-seq) sequencing data to
predict DNA methylation levels at single-CpG resolution. Our method is
a combined computational and experimental strategy to produce DNA
methylomes of all 28 million CpGs in the human genome for a fraction
(<10%) of the cost of whole-genome bisulfite sequencing methods.
methylCRF was benchmarked for accuracy against Infinium arrays, RRBS,
WGBS sequencing, and locus-specific bisulfite sequencing performed on
the same human embryonic stem cell line. methylCRF transformation of
MeDIP-seq/MRE-seq was equivalent to a biological replicate of WGBS in
quantification, coverage, and resolution. We used conventional
bisulfite conversion, PCR, cloning, and sequencing to validate loci
where our predictions do not agree with whole-genome bisulfite data,
and in 11 out of 12 cases, methylCRF predictions of methylation level
agree better with validated results than does whole-genome bisulfite
sequencing. Therefore, methylCRF transformation of MeDIP-seq/MRE-seq
data provides an accurate, inexpensive, and widely accessible strategy
to create full DNA methylomes.

<>

<1>Steward, G.F., Preston, C.M.
<2>Analysis of a viral metagenomic library from 200 m depth in Monterey Bay, California constructed by direct shotgun cloning.
<3>Virol. J.
<4>8
<5>287
<6>2011
<7>ABSTRACT: BACKGROUND: Viruses have a profound influence on both the
ecology and evolution of marine plankton, but the genetic diversity of
viral assemblages, particularly those in deeper ocean waters, remains
poorly described. Here we report on the construction and analysis of a
viral metagenome prepared from below the euphotic zone in a temperate,
eutrophic bay of coastal California. METHODS: We purified viruses from
approximately one cubic meter of seawater collected from 200m depth in
Monterey Bay, CA. DNA was extracted from the virus fraction, sheared, and
cloned with no prior amplification into a plasmid vector and propagated in
E. coli to produce the MBv200m library. Random clones were sequenced by
the Sanger method. Sequences were assembled then compared to sequences in
GenBank and to other viral metagenomic libraries using BLAST analyses.
RESULTS: Only 26% of the 881 sequences remaining after assembly had
significant (E </= 0.001) BLAST hits to sequences in the GenBank nr
database, with most being matches to bacteria (15%) and viruses (8%). When
BLAST analysis included environmental sequences, 74% of sequences in the
MBv200m library had a significant match. Most of these hits (70%) were to
microbial metagenome sequences and only 0.7% were to sequences from viral
metagenomes. Of the 121 sequences with a significant hit to a known virus,
94% matched bacteriophages (Families Podo-, Sipho-, and Myoviridae) and 6%
matched viruses of eukaryotes in the Family Phycodnaviridae (5 sequences)
or the Mimivirus (2 sequences). The largest percentages of hits to viral
genes of known function were to those involved in DNA modification (25%)
or structural genes (17%). Based on reciprocal BLAST analyses, the MBv200m
library appeared to be most similar to viral metagenomes from two other
bays and least similar to a viral metagenome from the Arctic Ocean.
CONCLUSIONS: Direct cloning of DNA from diverse marine viruses was
feasible and resulted in a distribution of virus types and functional
genes at depth that differed in detail, but were broadly similar to those
found in surface marine waters. Targeted viral analyses are useful for
identifying those components of the greater marine metagenome that
circulate in the subcellular size fraction.

<>

<1>Steward, N., Kusano, T., Sano, H.
<2>Expression of ZmMET1, a gene encoding a DNA methyltransferase from maize, is associated not only with DNA replication in actively proliferating cells, but also with altered DNA methylation status in cold-stressed quiescent cells.
<3>Nucleic Acids Res.
<4>28
<5>3250-3259
<6>2000
<7>A cDNA fragment encoding part of a DNA methyltransferase was isolated from maize. The putative
amino acid sequence identically matched that deduced from a genomic sequence in the database
(accession no. AF063403), and the corresponding gene was designated as ZmMET1. Bacterially
expressed ZmMET1 actively methylated DNA in vitro. Transcripts of ZmMET1 could be shown to
exclusively accumulate in actively proliferating cells of the meristems of mesocotyls and root
apices, suggesting ZmMET1 expression to be associated with DNA replication. This was confirmed
by simultaneous decrease of transcripts of ZmMET1 and histone H3, a marker for DNA
replication, in seedlings exposed to wounding, desiccation and salinity, all of which suppress
cell division. Cold stress also depressed both transcripts in root tissues. In contrast,
however, accumulation of ZmMET1 transcripts in shoot mesocotyls was not affected by cold
stress, whereas those for H3 sharply decreased. Such a differential accumulation of ZmMET1
transcripts was consistent with ZmMET1 protein levels as revealed by western blotting.
Expression of ZmMET1 is thus coexistent, but not completely dependent on DNA replication.
Southern hybridization analysis with a methylation-sensitive restriction enzyme revealed that
cold treatment induced demethylation of DNA in the Ac/Ds transposon region, but not in other
genes, and that such demethylation primarily occurred in roots. These results suggested that
the methylation level was decreased selectively by cold treatment, and that ZmMET1 may, at
least partly, prevent such demethylation.

<>

<1>Stewart, F., Dila, D., Raleigh, E.
<2>The mechanism of action of the McrBC restriction enzyme of E. coli K-12.
<3>J. Cell Biochem. Suppl.
<4>19A
<5>118
<6>1995
<7>As more restriction systems are characterised, their properties are being found to be more
diverse and the traditional three classes of restriction enzymes (Types I, II and III) are
becoming inadequate for classification. McrBC is one enzyme which fails to fit neatly into any
of the three classes. Like enzymes of classes I and III it is a multi-subunit enzyme
(consisting of 2 proteins, McrB and McrC) which requires two "half-sites" on the DNA for
cleavage. However, unlike the enzymes so far described in these classes, McrBC recognises only
methylated DNA, requiring at least one methylated C in each half-site, which is of the form
5'-RmC-3'. Unusually, the half-sites can be symmetric or asymmetric as the DNA will be
cleaved irrespective of which strand(s) the methylated Cs are on. Qualitatively, then, McrBC
can recognise the half-sites in an orientation-independent manner. However, using synthetic
oligos with the methylated bases in various configurations, we have shown that the efficiency
of cleavage varies with the configuration of the methylated Cs.
Spacing requirements for the half-sites were further investigated using a series of plasmids
which contain only two McrBC half-sites, flanking a polylinker into which were inserted DNA
fragments of various sizes, we have also found that cleavage efficiency depends on the spacing
between the half-sites. Maximal cleavage occurs with a spacing of approximately 40-80bp, but
cleavage can occur, less efficiently, with spacing of up to and including 1.2kb but not 3kb.
The DNA is cleaved neither at a single position close to the recognition site like Type III
enzymes, nor at a site very distant from the recognition site as with Type I enzymes, but
rather at multiple positions close to only one half-site in each molecule, with no apparent
preference for one half-site over the other.
As McrBC shows no sequence similarity to other restriction enzymes (this is the case for
many restriction enzymes) a clue as to its evolution may be obtained by elucidation of its
mechanism of action. It is again similar to enzymes of Types I and III in requiring a
nucleotide for cleavage but differs in its requirement for GTP rather than ATP. By gel
retardation assays using synthetic oligos containing two appropriately-methylated and
appropriately-spaced McrBC half-sites, we have shown that GTP is required for the initial
binding of McrBC to DNA but it remains to be determined whether GTP is further required to
allow communication between half-sites by McrBC as is true for ATP in the case of Type I and
Type III enzymes.

<>

<1>Stewart, F.J., Panne, D., Bickle, T.A., Raleigh, E.A.
<2>Methyl-specific DNA binding by McrBC, a modification-dependent restriction enzyme.
<3>J. Mol. Biol.
<4>298
<5>611-622
<6>2000
<7>McrBC, a GTP-requiring, modification-dependent endonuclease of Escherichia coli K-12,
specifically recognizes DNA sites of the form 5' R(m)C 3'. DNA cleavage normally requires
translocation-mediated coordination between two such recognition elements at distinct sites.
We have investigated assembly of the cleavage-competent complex with gel-shift and DNase I
footprint analysis. In the gel-shift system, McrB(L) binding resulted in a fast-migrating
specific shifted band, in a manner requiring both GTP and Mg(2+). The binding was specific for
methylated DNA and responded to local sequence changes in the same way that cleavage does.
Single-stranded DNA competed for McrB(L)-binding in a modification and sequence-specific
fashion. A supershifted species was formed in the presence of McrC and GTPgammaS. DNase I
footprint analysis showed modest cooperativity in binding to two sites, and a two-site
substrate displayed protection in non-specific spacer DNA in addition to the recognition
elements. The addition of McrC did not affect the footprint obtained. We propose that McrC
effects a conformational change in the complex rather than a reorganization of the DNA:protein
interface.

<>

<1>Stewart, F.J., Raleigh, E.A.
<2>Dependence of McrBC cleavage on distance between recognition elements.
<3>Biol. Chem.
<4>379
<5>611-616
<6>1998
<7>DNA cleavage by the modification-dependent restriction enzyme McrBC requires the presence of
two suitably modified recognition elements appropriately spaced in the substrate.  To
characterize the spacing requirement in more detail, we have constructed a plasmid with a
single McrBC cleavage site, in which the distance between recognition elements could be
systematically varied while preserving the local sequence surrounding the recognition
elements.  Optimal separation between elements was 55-103 base pairs, with detectable cleavage
observed at spacing of 32 bp to 2 kb; no cleavage was seen with spacing of 22 bp or less or
with 3 kb between elements.  Changing the spacing by 4 basepairs within the optimal range has
little effect on the efficiency of cleavage, suggesting that the recognition elements need not
lie on the same face of the DNA helix.

<>

<1>Stiens, M., Becker, A., Bekel, T., Godde, V., Goesmann, A., Niehaus, K., Schneiker-Bekel, S., Selbitschka, W., Weidner, S., Schluter, A., Puhler, A.
<2>Comparative genomic hybridisation and ultrafast pyrosequencing revealed remarkable differences between the Sinorhizobium meliloti genomes of the model strain Rm1021 and the field isolate SM11.
<3>J. Biotechnol.
<4>136
<5>31-37
<6>2008
<7>Genomic variation between the Sinorhizobium meliloti model strain Rm1021 and the field isolate
SM11 was assessed by using the genome-wide
S. meliloti Rm1021 Sm6k-oligonucleotide microarray in a comparative
genomic hybridisation experiment. Several gene clusters present in the
Rm1021 genome are missing in the SM11 genome. In detail, three missing
gene clusters were identified for the chromosome, five for megaplasmid
pSymA and two for megaplasmid pSymB. To confirm these hybridisation
results, the draft genome sequence of the S. meliloti field isolate
SM11 was established by 454-pyrosequencing. Three sequencing runs on
the ultrafast Genome Sequencer 20 System yielded 112.5 million bases.
These could be assembled into 905 larger contigs resulting in a nearly
15-fold coverage of the 7.1 Mb SM11 genome. The missing gene regions
identified by comparative genomic hybridisation could be confirmed by
the results of the 454-sequencing project. An in-depth analysis of
these gene regions resulted in the following findings: (i) a complete
type I restriction/modification system encoded by a composite
transposon is absent in the chromosome of strain SM11. (ii) Most of the
Rm1021 denitrification genes and the complete siderophore biosynthesis
operon were found to be missing on SM11 megaplasmid pSymA. (iii) S.
meliloti SM11 megaplasmid pSymB lacks a complete cell surface
carbohydrate synthesis gene cluster. (iv) Several genes that are absent
in the SM11 genome could be assigned to insertion sequences and
transposons.

<>

<1>Stiens, M., Schneiker, S., Puhler, A., Schluter, A.
<2>Sequence analysis of the 181-kb accessory plasmid pSmeSM11b, isolated from a dominant Sinorhizobium meliloti strain identified during a long-term field release experiment.
<3>FEMS Microbiol. Lett.
<4>271
<5>297-309
<6>2007
<7>The 181 251 bp accessory plasmid pSmeSM11b of Sinorhizobium meliloti
strain SM11, belonging to a dominant indigenous S. meliloti subpopulation
identified during a long-term field release experiment, was sequenced.
This plasmid has 166 coding sequences (CDSs), 42% of which encode proteins
with homology to proteins of known function. Plasmid pSmeSM11b is a member
of the repABC replicon family and contains a large gene region coding for
a conjugation system similar to that of other self-transmissible plasmids
in Rhizobium and Agrobacterium. Another pSmeSM11b gene region, possibly
involved in sugar metabolism and polysaccharide catabolism, resembled a
region of S. meliloti 1021 megaplasmid pSymB and in the genome of
Sinorhizobium medicae WSM419. Another module of plasmid pSmeSM11b encodes
proteins similar to those of the nitrogen-fixing actinomycete Frankia
CcI3, and which are likely to be involved in the synthesis of a secondary
metabolite. Several ORFs of pSmeSM11b were predicted to play a role in
nonribosomal peptide synthesis. Plasmid pSmeSM11b has many mobile genetic
elements, which contribute to the mosaic composition of the plasmid.

<>

<1>Stier, I., Kiss, A.
<2>The Type II restriction endonuclease MvaI has dual specificity.
<3>Nucleic Acids Res.
<4>38
<5>8231-8238
<6>2010
<7>The MvaI restriction endonuclease cuts 5'-CC downward arrowAGG-3'/5'-CC upward arrowTGG-3'
sites as indicated by the arrows. N4-methylation of the
inner cytosines (C(m4)CAGG/C(m4)CTGG) protects the site against MvaI
cleavage. Here, we show that MvaI nicks the G-strand of the related
sequence (CCGGG/CCCGG, BcnI site) if the inner cytosines are
C5-methylated: C(m5)C downward arrowGGG/CC(m5)CGG. At M.SssI-methylated
SmaI sites, where two oppositely oriented methylated BcnI sites partially
overlap, double-nicking leads to double-strand cleavage (CC(m5)C downward
arrowGGG/CC(m5)C upward arrowGGG) generating fragments with blunt ends.
The double-strand cleavage rate and the stringency of substrate site
recognition is lower at the methylation-dependent site than at the
canonical target site. MvaI is the first restriction endonuclease shown to
possess, besides the 'normal' activity on its unmethylated recognition
site, also a methylation-directed activity on a different sequence.

<>

<1>Stier, I., Kiss, A.
<2>Cytosine-to-Uracil Deamination by SssI DNA Methyltransferase.
<3>PLoS ONE
<4>8
<5>e79003
<6>2013
<7>The prokaryotic DNA(cytosine-5)methyltransferase M. SssI shares the specificity of eukaryotic
DNA methyltransferases (CG) and is an important model and experimental tool in the study of
eukaryotic DNA methylation. Previously, M. SssI was shown to be able to catalyze deamination
of the target cytosine to uracil if the methyl donor S-adenosyl-methionine (SAM) was missing
from the reaction. To test whether this side-activity of the enzyme can be used to distinguish
between unmethylated and C5-methylate cytosines in CG dinucleotides, we re-investigated, using
a sensitive genetic reversion assay, the cytosine deaminase activity of M. SssI. Confirming
previous results we showed that M. SssI can deaminate cytosine to uracil in a slow reaction in
the absence of SAM and that the rate of this reaction can be increased by the SAM analogue
5'-amino-5'-deoxyadenosine. We could not detect M. SssI-catalyzed deamination of
C5-methylcytosine (C-m5). We found conditions where the rate of M. SssI mediated C-to-U deamin
ation was at least 100-fold higher than the rate of C-m5-to-T conversion. Although this
difference in reactivities suggests that the enzyme could be used to identify C5-methylated
cytosines in the epigenetically important CG dinucleotides, the rate of M. SssI mediated
cytosine deamination is too low to become an enzymatic alternative to the bisulfite reaction.
Amino acid replacements in the presumed SAM binding pocket of M. SssI (F17S and G19D) resulted
in greatly reduced methyltransferase activity. The G1 9D variant showed cytosine deaminase
activity in E. coli, at physiological SAM concentrations. Interestingly, the C-to-U deaminase
activity was also detectable in an E. coli ung(+) host proficient in uracil excision repair.

<>

<1>Stierschneider, U., Noiges, B., Gabain, A.V., Cipps, T., Bakshi, S., Meinke, A., Zierer, D., Lundberg, U., Nagy, E., Satke, C., Pikalo, J., Hanner, M.
<2>Klebsiella antigens.
<3>Japanese Patent Office
<4>JP 2010532657 A
<5>
<6>2010
<7>
<>

<1>Stinear, T.P. et al.
<2>Insights from the complete genome sequence of Mycobacterium marinum on the evolution of Mycobacterium tuberculosis.
<3>Genome Res.
<4>18
<5>729-741
<6>2008
<7>Mycobacterium marinum, a ubiquitous pathogen of fish and amphibia, is a near relative of
Mycobacterium tuberculosis, the etiologic agent of
tuberculosis in humans. The genome of the M strain of M. marinum comprises
a 6,636,827-bp circular chromosome with 5424 CDS, 10 prophages, and a
23-kb mercury-resistance plasmid. Prominent features are the very large
number of genes (57) encoding polyketide synthases (PKSs) and nonribosomal
peptide synthases (NRPSs) and the most extensive repertoire yet reported
of the mycobacteria-restricted PE and PPE proteins, and related-ESX
secretion systems. Some of the NRPS genes comprise a novel family and seem
to have been acquired horizontally. M. marinum is used widely as a model
organism to study M. tuberculosis pathogenesis, and genome comparisons
confirmed the close genetic relationship between these two species, as
they share 3000 orthologs with an average amino acid identity of 85%.
Comparisons with the more distantly related Mycobacterium avium subspecies
paratuberculosis and Mycobacterium smegmatis reveal how an ancestral
generalist mycobacterium evolved into M. tuberculosis and M. marinum. M.
tuberculosis has undergone genome downsizing and extensive lateral gene
transfer to become a specialized pathogen of humans and other primates
without retaining an environmental niche. M. marinum has maintained a
large genome so as to retain the capacity for environmental survival while
becoming a broad host range pathogen that produces disease strikingly
similar to M. tuberculosis. The work described herein provides a
foundation for using M. marinum to better understand the determinants of
pathogenesis of tuberculosis.

<>

<1>Stobberingh, E.E., Meijers, J.A., Van Kats-Renaud, J.H.
<2>The sensitivity of phage DNA and plasmid DNA for a restriction enzyme from Staphylococcus aureus.
<3>Antonie Van Leeuwenhoek
<4>45
<5>19-23
<6>1979
<7>In Staphylococcus aureus transduction of different tetracycline and chloramphenicol plasmids
with a group I III modification was possible to group I and III strains.  Group II strains,
containing a restriction endonuclease, had a restriction both for the phage and the plasmids:
two restriction-deficient group II strains were good acceptors for these plasmids.

<>

<1>Stobberingh, E.E., Schiphof, R., Sussenbach, J.S.
<2>Occurrence of a Class II Restriction Endonuclease in Staphylococcus aureus.
<3>J. Bacteriol.
<4>131
<5>645-649
<6>1977
<7>The occurrence of class II restriction endonucleases (enzymes that both
recognize and cleave a specific nucleotide sequence in deoxyribonucleic acid
(DNA) in Staphyloccus aureus has been investigated by analysis of crude
extracts obtained from different propagating strains of the International Phage
Typing System.  Of the four main groups of strains in the International System,
only extracts of group II strains were found to contain class II restriction
endonucleases.  The identical cleavage patterns obtained by incubation of
different DNAs with cell extracts of group II strains suggest that these
enzymes all recognize and cleave the same nucleotide sequence.  This
recognition site has been determined to be 5'-G-A-T-C-3' 3'-C-T-A-G-5' for the
prototype of these enzymes, Sau3AI (J. Sussenbach et al., Nucl. Acids Res. 3:
3192-3202, 1976).  Evidence is presented that the classification of group II
strains is based on restriction modification and is correlated with the
presence of a class II restriction endonuclease that recognizes and cleaves the
above sequence.

<>

<1>Stobberingh, E.E., Winkler, K.C.
<2>Restriction-deficient mutants of Staphylococcus aureus.
<3>J. Gen. Microbiol.
<4>99
<5>359-367
<6>1977
<7>A series of restriction-deficient mutants was isolated from non-lysogenic strains of
Staphylococcus aureus belonging to phage groups I and II.  Some mutants were sensitive to all
phages tested.  With one possible exception, all the mutants were unaffected in their
modification systems.  The breakdown of DNA of phages, restricted in the parental strains, was
reduced in both the mutants that were tested.  The restriction in propagating strain 3A could
be transduced to its restriction-deficient mutant.  The transduction efficiency increased
after ultraviolet irradiation of the transducing phage suggesting that the gene for
restriction is present on the bacterial chromosome.

<>

<1>Stoddard, B., Belfort, M.
<2>Social networking between mobile introns and their host genes.
<3>Mol. Microbiol.
<4>78
<5>1-4
<6>2010
<7>P>Homing endonucleases have long been known as the orchestrators of intron mobility. However,
the extent of their influence on the intron
and its genetic and cellular environment is still being elucidated. The
accompanying paper emphasizes the importance of temporal control of
endonuclease expression on splicing, expression of the host gene and
cellular metabolism, while it raises questions to guide future inquiry.

<>

<1>Stoddard, B.L.
<2>Homing endonuclease structure and function.
<3>Q. Rev. Biophys.
<4>38
<5>49-95
<6>2005
<7>Homing endonucleases are encoded by open reading frames that are embedded within group I,
group II and archaeal introns, as well as inteins (intervening sequences that are spliced and
excised post-translationally). These enzymes initiate transfer of those elements (and
themselves) by generating strand breaks in cognate alleles that lack the intervening sequence,
as well as in additional ectopic sites that broaden the range of intron and intein mobility.
Homing endonucleases can be divided into several unique families that are remarkable in
several respects: they display extremely high DNA-binding specificities which arise from long
DNA target sites (14-40 bp), they are tolerant of a variety of sequence variations in these
sites, and they display disparate DNA cleavage mechanisms. A significant number of homing
endonucleases also act as maturases (highly specific cofactors for the RNA splicing reactions
of their cognate introns). Of the known homing group I endonuclease families, two (HNH and
His-Cys box enzymes) appear to be diverged from a common ancestral nuclease. While crystal
structures of several representatives of the LAGLIDADG endonuclease family have been
determined, only structures of single members of the HNH (I-HmuI), His-Cys box (I-PpoI) and
GIY-YIG (I-TevI) families have been elucidated. These studies provide an important source of
information for structure-function relationships in those families, and are the centerpiece of
this review. Finally, homing endonucleases are significant targets for redesign and selection
experiments, in hopes of generating novel DNA binding and cutting reagents for a variety of
genomic applications.

<>

<1>Stoddard, B.L.
<2>Homing endonucleases: from microbial genetic invaders to reagents for targeted DNA modification.
<3>Structure
<4>19
<5>7-15
<6>2011
<7>Homing endonucleases are microbial DNA-cleaving enzymes that mobilize their own reading frames
by generating double strand breaks at specific genomic invasion sites. These proteins display
an economy of size, and yet recognize long DNA sequences (typically 20 to 30 base pairs). They
exhibit a wide range of fidelity at individual nucleotide positions in a manner that is
strongly influenced by host constraints on the coding sequence of the targeted gene. The
activity of these proteins leads to site-specific recombination events that can result in the
insertion, deletion, mutation, or correction of DNA sequences. Over the past fifteen years,
the crystal structures of representatives from several homing endonuclease families have been
solved, and methods have been described to create variants of these enzymes that cleave novel
DNA targets. Engineered homing endonucleases proteins are now being used to generate targeted
genomic modifications for a variety of biotech and medical applications.

<>

<1>Stoddard, B.L., Jurica, M., Heath, P., Flick, K.
<2>The structure, function, and convergent evolution of intron-encoded homing endonucleases.
<3>Biochem. Soc. Trans.
<4>27
<5>A39
<6>1999
<7>The homing endonucleases are a diverse family of proteins encoded by open reading frames in
genetically mobile, self-splicing introns.  Similar endonucleases have also been identified as
optional, independently folded domains in self-splicing protein introns, termed 'inteins'.
These comparatively small enzymes share the ability to recognize and cleave long DNA sites of
20 to 40 bp, and promote the lateral transfer of their host intron or intein to these sites by
a targeted transposition.  These proteins also display flexibility of site-recognition, and
are capable of tolerating changes at any position in the target DNA site.  Our laboratory has
determined the structure of representative members of two families of homing endonucleases,
both unbound and complexed to their DNA targets: I-CreI (a LAGLIDADG endonuclease) and I-PpoI
(a his-cys box endonuclease).  The structures both demonstrate an impressive ability of these
proteins to adopt an economical, elongated fold and to form a DNA complex with sub-saturating
atomic contacts across the full length of the homing site.  The co-crystal structures indicate
that the enzymes probably follow two very different structural mechanisms for phosphodiester
hydrolysis.

<>

<1>Stoddard, B.L., Lambert, A.R., Takeuchi, R., Scharenberg, A.M., Baxter, S.K.
<2>Generation, expression and structure of engineered homing endonuclease I-OnuI and its homologues and uses for genome engineering.
<3>International Patent Office
<4>WO 2011156430 A
<5>
<6>2011
<7>The present disclosure provides compositions and methods for producing and expressing variant
or engineered I-Onu1 endonucleases, variant or engineered I-Onu1 homologues, and hybrids of
two I-Onu1 homologue domains that have a target site altered from the wild-type.  A method for
selecting a variant or engineered I-Onu1 endonuclease, I-Onu1 endonuclease homologue, and a
hyurid of two I-Onu1 or I-Onu1 homologue domains that have a target site altered from the
wild-type and directed to a site within a gene of interest is also provided.  In addition, the
present disclosure provides the crystal structure of the I-Onu1 and I-Ltr1 endonucleases; the
specificity profiles for both endonuclease for DNA binding and cleavage; the identity of amino
acid residue positions in the I-Onu1 and I-Ltr1 protein scaffold that determine DNA
recognition specificity; methods for determining amino acid substitutions at those positions
that alter DNA cleavage specificity; methods for the complete redesign of the DNA cleavage
specificity of I-Onu1 and its homologues for recognition and cleavage of a human gene of
interest; and the relationship of the amino acid sequence, structure and specificity of I-Onu1
to a collection of identifiable I-Onu1 endonuclease homologues.

<>

<1>Stoddard, B.L., Monnat, R.J., Baker, D., Chevalier, B., Kortemme, T., Chadsey, M.
<2>Methods and compositions concerning designed highly-specific nucleic acid binding proteins.
<3>International Patent Office
<4>WO 200431346
<5>
<6>2004
<7>NOVELTY - Creating a modified nuclease with nucleic acid sequence-specificity activity
comprises identifying potential contact points between the DNA binding domain and the specific
nucleic acid sequence.
DETAILED DESCRIPTION - The method comprises: (a) preparing a computational model of a complex
between an unmodified nuclease and the specific nucleic acid sequence, where the nuclease
comprises a catalytic domain and a DNA binding domain; (b) identifying
potential contact points between the DNA binding domain and the
specific nucleic acid sequence; and identifying an amino acid change
that creates or enhances a contact point to improve an interface
between the DNA binding domain and the nucleic acid sequence, and
further provides a design for the modified nuclease. INDEPENDENT CLAIMS
are included for the following: (1) a modified nuclease that has
altered sequence-specificity made by the method cited above; (2) a
method of designing a modified chimeric nucleic acid binding
polypeptide with sequence-specific activity; (3) a modified nuclease
with nucleic acid sequence specific activity;(4) a modified chimeric
nuclease with site-specific activity at a combined target nucleotide
sequence comprising a first DNA binding domain and a catalytic domain
from a first homing endonuclease, and a second binding domain from a
second homing endonuclease; (5) a modified chimeric homing endonuclease
comprising the DNA binding and catalytic domains of Dmo-I and the DNA
binding domain of Cre-I; and (6) a method of screening to identify a
modified DNA binding polypeptide with altered DNA sequence-specific
activity. BIOTECHNOLOGY - Preferred Method: In creating a modified
nuclease with nucleic acid sequence-specific activity, the nucleic acid
is double-stranded DNA. The specific nucleic acid sequence is at least
8 residues in length. The method further comprises preparing the
modified nuclease. The nuclease is a homing endonuclease, which is a
member of the LAGLIDADG family. The LAGLIDADG endonuclease is l-Dmo-l,
Cre-I, I-CeuI, I-SceI, I-SceE, I-SceV, I-SceVI, or I-LldI. The amino
acid change results in reduced steric hindrance or improper bonding
between the DNA binding domain and the nucleic acid sequence. The amino
acid change creates a contact point between the DNA binding domain and
the nucleic acid sequence. The amino acid changes are determined
computationally. The method comprises obtaining a crystal structure of
the unmodified nuclease. The crystal structure further comprises a
specific nucleic acid sequence of the unmodified nuclease. Obtaining a
crystal structure of the unmodified nuclease comprises generating the
crystal structure. The method further comprises detecting nucleic acid
sequence-specific activity of the modified nuclease by a binding assay.
Computationally determining amino acid changes comprises calculating
the free energy of amino acid residues at each contact point or of
amino acid residues along the interface, or evaluating a library of
amino acid side chain rotamer conformations in different
backbone-dependent rotameric states for reduced free energy. The
library excludes cysteine. The different backbone-dependent rotameric
states comprise retainers interacting with surrounding groups;
retainers interacting with a fixed portion of the molecule including
the backbone and all side chains not subject to substitution; and
pairwise rotamer to rotamer energies. The method further comprises
calculating the free energy of the designed modified nuclease. The
over-all minimum free energy is increased by providing at least one
polypeptide backbone models with different relative orientations of
LAGLIDADG sequences in the interface between the DNA binding domain and
the nucleic acid sequence; performing sequence design assays on the
polypeptide backbone; and obtaining different amino acid combinations
at each contact point. The number of amino acid combinations is reduced
by eliminating sequences that affect the activity of nearby contact
point res

<>

<1>Stoddard, B.L., Scharenberg, A.M., Monnat, R.J. Jr.
<2>Advances in engineering homing endonucleases for gene targeting: ten years after structures.
<3>Prog. Gene Ther., , Bertolotti, R., Ozawa, K., 
<4>3
<5>135-167
<6>2008
<7>Homing endonucleases are highly site-specific endonucleases that induce homologous
recombination or gene conversion in vivo by cleaving long (typically >18bp) DNA target sites.
Homing endonucleases are under development as tools for targeted genetic engineering
applications, ranging from therapeutic gene correction to metabolic and population
engineering.  The first structures of homing endonucleases were reported 10 years ago.  Since
that time, representative structures from each of the known families of homing endonucleases
have been determined, and the corresponding details of their mechanisms of DNA recognition and
cleavage have been elucidated.  Using this information, the LAGLIDADG homing endonuclease
family has been identified as the most tractable for further modification by structure-based
selection and/or engineering approaches.  Most recently, successful redesign of the I-CreI
endonuclease has led to the development of reagents that recognize and act on genes associated
with monogenic diseases, including the human RAG1 and XPC genes.  These studies demonstrate
the feasibility of using engineered homing endonucleases to promote efficient and target
site-specific modification of chromosomal loci.  Current studies are rapidly improving the
throughput and efficiency of homing endonuclease design and selection, and aim to optimize the
specificity and activity of the resulting endonucleases for targeted genomic applications in
medicine and biotechnology.

<>

<1>Stokes, H.W., Nesbo, C.L., Holley, M., Bahl, M.I., Gillings, M.R., Boucher, Y.
<2>Class 1 Integrons Potentially Predating the Association with Tn402-Like Transposition Genes Are Present in a Sediment Microbial Community.
<3>J. Bacteriol.
<4>188
<5>5722-5730
<6>2006
<7>Integrons are genetic elements that contribute to lateral gene transfer in bacteria as a
consequence of possessing a site-specific recombination system. This system facilitates the
spread of genes when they are part of mobile cassettes. Most integrons are contained within
chromosomes and are confined to specific bacterial lineages. However, this is not the case for
class 1 integrons, which were the first to be identified and are one of the single biggest
contributors to multidrug-resistant nosocomial infections, carrying resistance to many
antibiotics in diverse pathogens on a global scale. The rapid spread of class 1 integrons in
the last 60 years is partly a result of their association with a specific suite of
transposition functions, which has facilitated their recruitment by plasmids and other
transposons. The widespread use of antibiotics has acted as a positive selection pressure for
bacteria, especially pathogens, which harbor class 1 integrons and their associated antibiotic
resistance genes. Here, we have isolated bacteria from soil and sediment in the absence of
antibiotic selection. Class 1 integrons were recovered from four different bacterial species
not known to be human pathogens or commensals. All four integrons lacked the transposition
genes previously considered to be a characteristic of this class. At least two of these
integrons were located on a chromosome, and none of them possessed antibiotic resistance
genes. We conclude that novel class 1 integrons are present in a sediment environment in
various bacteria of the beta-proteobacterial class. These data suggest that the dispersal of
this class may have begun before the "antibiotic era."

<>

<1>Stokke, R., Hocking, W.P., Steinsbu, B.O., Steen, I.H.
<2>Complete Genome Sequence of the Thermophilic and Facultatively Chemolithoautotrophic Sulfate Reducer Archaeoglobus sulfaticallidus Strain  PM70-1T.
<3>Genome Announcements
<4>1
<5>e00406-13
<6>2013
<7>Dissimilatory sulfate-reducing archaea of the genus Archaeoglobus display divergent
preferences in the use of energy sources and electron acceptors. Here
we present the complete genome sequence of the thermophilic Archaeoglobus
sulfaticallidus strain PM70-1(T), which distinctly couples chemolithoautotrophic
growth on H2/CO2 to sulfate reduction in addition to heterotrophic growth.

<>

<1>Stolt, P., Grampp, B., Zillig, W.
<2>Genes for DNA cytosine methyltransferases and structural proteins, expressed during lytic growth by the phage Phi-H of the archaebacterium Halobacterium salinarium.
<3>Biol. Chem. Hoppe Seyler
<4>375
<5>747-757
<6>1994
<7>Lytic genes and transcription from the Halobacterium salinarium phage phi-H were studied.
Genes for three structural proteins were located to the left arm of the linear phage genome.
The right arm was shown to encode three DNA cytosine methyltransferases, the first such
sequences reported from an archaebacterium. One cytosine methyltransferase is of the
N(4)-methyltransferase type. The other two open reading frames (ORFs) seem to be parts of the
same gene, which has been split by a recombination event. This gene product is of the
C5-methyltransferase type. The methyltransferase genes are the first phi-H genes detected
showing high homology to eubacterial proteins. Five of the six described gene products have a
higher proportion of acidic over basic amino acid residues, a common characteristic of
halobacterial proteins. Lytic phi-H transcription was shown to produce three RNA species, two
shorter species encoding the methyltransferase genes and one large species transcribed from
both the right and the left phage arm and subsequently being processed upstream of the region
encoding the structural proteins.

<>

<1>Stolyar, S., Liu, Z., Thiel, V., Tomsho, L.P., Pinel, N., Nelson, W.C., Lindemann, S.R., Romine, M.F., Haruta, S., Schuster, S.C., Bryant, D.A., Fredrickson, J.K.
<2>Genome Sequence of the Thermophilic Cyanobacterium Thermosynechococcus sp. Strain NK55a.
<3>Genome Announcements
<4>2
<5>e01060-13
<6>2014
<7>The genome of the unicellular cyanobacterium Thermosynechococcus sp. strain NK55a, isolated
from the Nakabusa hot spring, Nagano Prefecture, Japan, comprises
a single, circular, 2.5-Mb chromosome. The genome is predicted to contain 2,358
protein-encoding genes, including genes for all typical cyanobacterial
photosynthetic and metabolic functions. No genes encoding hydrogenases or
nitrogenase were identified.

<>

<1>Stone, J.K., Johnson, S.L., Bruce, D.C., Detter, J.C., Mayo, M., Currie, B.J., Gelhaus, H.C., Keim, P., Tuanyok, A.
<2>Complete Genome Sequence of the Encephalomyelitic Burkholderia pseudomallei Strain MSHR305.
<3>Genome Announcements
<4>1
<5>e00656-13
<6>2013
<7>We describe the complete genome sequence of Burkholderia pseudomallei MSHR305, a  clinical
isolate taken from a fatal encephalomyelitis case, a rare form of
melioidosis. This sequence will be used for comparisons to identify the genes
that are involved in neurological cases.

<>

<1>Stover, C.K. et al.
<2>Complete genome sequence of Pseudomonas aeruginosa PA01, an opportunistic pathogen.
<3>Nature
<4>406
<5>959-964
<6>2000
<7>Pseudomonas aeruginosa is a ubiquitous environmental bacterium that is one of the top three
causes of opportunistic human infections. A major factor in its prominence as a pathogen is
its intrinsic resistance to antibiotics and disinfectants. Here we report the complete
sequence of P. aeruginosa strain PAO1. At 6.3 million base pairs, this is the largest
bacterial genome sequenced, and the sequence provides insights into the basis of the
versatility and intrinsic drug resistance of P. aeruginosa. Consistent with its larger genome
size and environmental adaptability, P. aeruginosa contains the highest proportion of
regulatory genes observed for a bacterial genome and a large number of genes involved in the
catabolism, transport and efflux of organic compounds as well as four potential chemotaxis
systems. We propose that the size and complexity of the P. aeruginosa genome reflect an
evolutionary adaptation permitting it to thrive in diverse environments and resist the effects
of a variety of antimicrobial substances.

<>

<1>Stover, T., Kohler, E., Fagin, U., Wende, W., Wolfes, H., Pingoud, A.
<2>Determination of the DNA bend angle induced by the restriction endonuclease EcoRV in the presence of Mg2++.
<3>J. Biol. Chem.
<4>268
<5>8645-8650
<6>1993
<7>We have used the method of Zinkel and Crothers (1990, Biopolymers 29: 29-38) to determine the
degree of bending induced by the binding of the restriction endonuclease EcoRV to its
recognition sequence (-GATATC-). A set of four calibration DNA fragments was constructed that
contained zero, two, four, or six phased A-tracts in their centers and an EcoRV site at the
5'-end to account for the electrophoretic influence of the bound protein. The mobilities of
these calibration molecules complexed with EcoRV were compared to that of a test DNA
containing a central EcoRV site also complexed with EcoRV. The EcoRV-induced bend angle was
found to be 44 +/- 4. These experiments were performed with a catalytically inactive EcoRV
mutant that still binds DNA specifically in the presence of Mg2+. In the absence of Mg2+,
which is necessary for specific binding, there is no difference in the mobilities of the
fragments with a peripheral or a central EcoRV site complexed with EcoRV, indicating that
nonspecific binding on average does not lead to measurable DNA bending.

<>

<1>Strabala, T.J., Macdonald, L., Liu, V., Smit, A.-M.
<2>Draft genome sequence of Novosphingobium nitrogenifigens Y88T.
<3>J. Bacteriol.
<4>194
<5>201
<6>2011
<7>Novosphingobium nitrogenifigens was originally isolated from pulp and paper mill wastewater, a
low-nitrogen, high-carbon environment.  N. nitrogenifigens is the first known nitrogen-fixing,
polyhydroxyalkanoate-accumulating sphingomonad, and we report the annotated draft genome
sequence of the type strain Y88T here.

<>

<1>Strachan, C.R., Singh, R., VanInsberghe, D., Ievdokymenko, K., Budwill, K., Mohn, W.W., Eltis, L.D., Hallam, S.J.
<2>Metagenomic scaffolds enable combinatorial lignin transformation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>111
<5>10143-10148
<6>2014
<7>Engineering the microbial transformation of lignocellulosic biomass is essential
to developing modern biorefining processes that alleviate reliance on
petroleum-derived energy and chemicals. Many current bioprocess streams depend on
the genetic tractability of Escherichia coli with a primary emphasis on
engineering cellulose/hemicellulose catabolism, small molecule production, and
resistance to product inhibition. Conversely, bioprocess streams for lignin
transformation remain embryonic, with relatively few environmental strains or
enzymes implicated. Here we develop a biosensor responsive to monoaromatic lignin
transformation products compatible with functional screening in E. coli. We use
this biosensor to retrieve metagenomic scaffolds sourced from coal bed bacterial
communities conferring an array of lignin transformation phenotypes that
synergize in combination. Transposon mutagenesis and comparative sequence
analysis of active clones identified genes encoding six functional classes
mediating lignin transformation phenotypes that appear to be rearrayed in nature
via horizontal gene transfer. Lignin transformation activity was then
demonstrated for one of the predicted gene products encoding a multicopper
oxidase to validate the screen. These results illuminate cellular and
community-wide networks acting on aromatic polymers and expand the toolkit for
engineering recombinant lignin transformation based on ecological design
principles.

<>

<1>Strathdee, G., Brown, R.
<2>Epigenetic cancer therapies: DNA methyltransferase inhibitors.
<3>Expert Opin. Invest. Drugs
<4>11
<5>747-754
<6>2002
<7>Human cancers frequently show altered patterns of DNA methylation, particularly at CpG
islands.  These CpG islands are sequences of DNA rich in CpG dinucleotides and are often found
close to gene promoters.  Methylation within islands has been shown to be associated with
transcriptional repression of the linked gene.  Genes involved in all facets of tumour
development and progression can become methylated and epigenetically silenced.  Re-expression
of such silenced genes can lead to suppression of tumour growth or sensitisation to anticancer
therapies.  Agents that can reverse DNA methylation include nucleoside and non-nucleoside
inhibitors of DNA methyltransferase.  Such agents are now undergoing preclinical evaluation
and clinical trials in cancer patients.

<>

<1>Strauch, E., Goelz, G., Knabner, D., Konietzny, A., Lanka, E., Appel, B.
<2>A cryptic plasmid of Yersinia enterocolitica encodes a conjugative transfer system related to the regions of CloDF13 Mob and IncX Pil.
<3>Microbiology
<4>149
<5>2829-2845
<6>2003
<7>Yersinia enterocolitica 29930 (biotype 1A; O : 7,8), the producing strain of the
phage-tail-like bacteriocin enterocoliticin, possesses a
plasmid-encoded conjugative type IV transfer system. The genes of the
conjugative system were found by screening of a cosmid library constructed
from total DNA of strain 29930. The cosmid Cos100 consists of the vector
SuperCos1 and an insert DNA of 40 303 bp derived from a cryptic plasmid of
strain 29930. The conjugative transfer system consists of genes encoding a
DNA transfer and replication system (Dtr) with close relationship to the
mob region of the mobilizable plasmid CloDF13 and a gene cluster encoding
a mating pair formation system (Mpf) closely related to the Mpf system of
the IncX plasmid R6K. However, a gene encoding a homologue of TaxB, the
coupling protein of the IncX system, is missing. The whole transfer region
has a size of approximately 17 kb. The recombinant plasmid Cos100 was
shown to be transferable between Escherichia coli and Yersinia with
transfer frequencies up to 0.1 transconjugants per donor. Mutations
generated by inserting a tetracycline cassette into putative tri genes
yielded a transfer-deficient phenotype. Conjugative transfer of the
cryptic plasmid could not be demonstrated in the original host Y.
enterocolitica 29930. However, a kanamycin-resistance-conferring
derivative of the plasmid was successfully introduced into E. coli K-12 by
transformation and was shown to be self-transmissible. Furthermore,
Southern blot hybridization and PCR experiments were carried out to
elucidate the distribution of the conjugative transfer system in YERSINIA:
In total, six Y. enterocolitica biotype 1A strains harbouring closely
related systems on endogenous plasmids were identified.

<>

<1>Strauch, E., Hammerl, J.A., Konietzny, A., Schneiker-Bekel, S., Arnold, W., Goesmann, A., Puhler, A., Beutin, L.
<2>Bacteriophage 2851 is a prototype phage for dissemination of the Shiga toxin variant gene 2c in Escherichia coli O157:H7.
<3>Infect. Immun.
<4>76
<5>5466-5477
<6>2008
<7>The production of Shiga toxin (Stx) (verocytotoxin) is a major virulence
factor of Escherichia coli O157:H7 strains (Shiga toxin-producing E. coli
[STEC] O157). Two types of Shiga toxins, designated Stx1 and Stx2, are
produced in STEC O157. Variants of the Stx2 type (Stx2, Stx2c) are
associated with high virulences of these strains for humans. A
bacteriophage designated 2851 from a human STEC O157 encoding the Stx2c
variant was described previously. Nucleotide sequence analysis of the
phage 2851 genome revealed 75 predicted coding sequences and indicated a
mosaic structure typical for lambdoid phages. Analyses of free phages and
K-12 phage 2851 lysogens revealed that upon excision from the bacterial
chromosome, the loss of a phage-encoded IS629 element leads to fusion of
phage antA and antB genes, with the generation of a recombined antAB gene
encoding a strong antirepressor. In wild-type E. coli O157 as well as in
K-12 strains, phage 2851 was found to be integrated in the sbcB locus.
Additionally, phage 2851 carries an open reading frame which encodes an
OspB-like type III effector similar to that found in Shigella spp.
Investigation of 39 stx(2c) E. coli O157 strains revealed that all except
1 were positive for most phage 2851-specific genes and possessed a
prophage with the same border sequences integrated into the sbcB locus.
Phage 2851-specific sequences were absent from most stx(2c)-negative E.
coli O157 strains, and we suggest that phage 2851-like phages contributed
significantly to the dissemination of the Stx2c variant toxin within this
group of E. coli.

<>

<1>Strauch, M.A.
<2>Delineation of AbrB-binding sites on the Bacillus subtilis spoOH, kinB, ftsAZ, and pbpE promoters and use of a derived homology to identify a previously unsuspected binding site in the bsuB1 methylase promoter.
<3>J. Bacteriol.
<4>177
<5>6999-7002
<6>1995
<7>DNase I footprinting experiments showed that AbrB binds to the regulatory regions of the
spoOH, kinB, ftsAZ, and pbpE genes.  A conserved motif was found in these and other
AbrB-binding sites.  A search for Bacillus subtilis DNA sequences containing this motif led to
the prediction that AbrB would bind to the promoter controlling the bsuB1 methylase gene.
DNase I footprinting experiments confirmed this prediction.

<>

<1>Strausberg, R.L. et al.
<2>Generation and initial analysis of more than 15,000 full-length human and mouse cDNA sequences.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>16899-16903
<6>2002
<7>The National Institutes of Health Mammalian Gene Collection (MGC) Program is a
multiinstitutional effort to identify and sequence a cDNA clone
containing a complete ORF for each human and mouse gene. ESTs were
generated from libraries enriched for full-length cDNAs and analyzed to
identify candidate full-ORF clones, which then were sequenced to high
accuracy. The MGC has currently sequenced and verified the full ORF for a
nonredundant set of >9,000 human and >6,000 mouse genes. Candidate
full-ORF clones for an additional 7,800 human and 3,500 mouse genes also
have been identified. All MGC sequences and clones are available without
restriction through public databases and clone distribution networks (see
http:mgc.nci.nih.gov).

<>

<1>Streeck, R.E.
<2>Single-strand and double-strand cleavage at half-modified and fully modified recognition sites for the restriction nucleases Sau3A and TaqI.
<3>Gene
<4>12
<5>267-275
<6>1980
<7>The influence of cytosine methylation on the cleavage of DNA by the restriction
nucleases Sau3A and TaqI has been investigated.  Bovine satellite DNA fragments
containing a GATCGA sequence, i.e. a Sau3A site overlapping with a TaqI site
have been used in this study.  The methylation of these fragments has been
determined by sequence analysis.  It has been found that a TaqI site (TCGA)
methylated at cytosine in both DNA strands is still sensitive to double-strand
cleavage.  A Sau3A site (GATC), however, is rendered resistant to double-strand
cleavage by methylation of a single cytosine.  Fragments containing the
"half-modified" Sau3A site are nicked in the unmethylated DNA strand.  It has
been shown by sequence analysis of nicked DNA that the single-strand break
occurs at the same position which is cleaved in unmodified DNA.

<>

<1>Streeter, S.D., McGeehan, J.E., Kneale, G.G.
<2>Overexpression, purification and preliminary X-ray diffraction analysis of the controller protein C.Csp231I from Citrobacter sp RFL231.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>F65
<5>898-901
<6>2009
<7>Restriction-modification controller proteins play an essential role in regulating the temporal
expression of restriction-modification genes.
The controller protein C.Csp231I represents a new class of controller
proteins. The gene was sublconed to allow overexpression in Escherichia
coli. The protein was purified to homogeneity and crystallized using
the hanging-drop vapour-diffusion method. The crystals diffracted to
2.0 angstrom resolution and belonged to space group P2(1). An
electrophoretic mobility-shift assay provided evidence of strong
binding of C.Csp231I to a sequence located upstream of the csp231IC
start codon.

<>

<1>Streeter, S.D., Papapanagiotou, I., McGeehan, J.E., Kneale, G.G.
<2>DNA footprinting and biophysical characterization of the controller protein C.AhdI suggests the basis of a genetic switch.
<3>Nucleic Acids Res.
<4>32
<5>6445-6453
<6>2004
<7>We have cloned and expressed the ahdIC gene of the AhdI restriction-modification system and
have purified the resulting controller (C) protein to homogeneity. The protein sequence shows
a HTH motif typical of that found in many transcriptional regulators. C.AhdI is found to form
a homodimer of 16.7 kDa; sedimentation equilibrium experiments show that the dimer dissociates
into monomers at low concentration, with a dissociation constant of 2.5 microM. DNase I and
Exo III footprinting were used to determine the C.AhdI DNA-binding site, which is found
approximately 30 bp upstream of the ahdIC operon. The intact homodimer binds cooperatively to
a 35 bp fragment of DNA containing the C-protein binding site with a dissociation constant of
5-6 nM, as judged both by gel retardation analysis and by surface plasmon resonance, although
in practice the affinity for DNA is dominated by protein dimerization as DNA binding by the
monomer is negligible. The location of the C-operator upstream of both ahdIC and ahdIR
suggests that C.AhdI may act as a positive regulator of the expression of both genes, and
could act as a molecular switch that is critically dependent on the K(d) for the monomer-dimer
equilibrium. Moreover, the structure and location of the C.AhdI binding site with respect to
the putative -35 box preceding the C-gene suggests a possible mechanism for autoregulation of
C.AhdI expression.

<>

<1>Streiff, M.B., Iida, S., Bickle, T.A.
<2>Expression and proteolytic processing of the darA antirestriction gene product of bacteriophage P1.
<3>Virology
<4>157
<5>167-171
<6>1987
<7>The darA gene coding for one of the two bacteriophage P1 antirestriction functions is
expressed late after infection or induction.  The protein is made as a high-molecular-weight
soluble precursor.  This is proteolytically cleaved to the mature form, which is a structural
component of the phage head.  Defective mutants of the phage have been found in which the
synthesis of gpdarA is normal but processing does not take place.  These mutations all map to
the same region of the P1 genome and we propose that they lie in the structural gene for the
processing protease.

<>

<1>Striebel, H.-M., Kessler, C.
<2>Novel specific endonuclease activity recognizing a 10-bp sequence.
<3>Gene
<4>172
<5>47-48
<6>1996
<7>We report here the generation of a novel restriction endonuclease (Enase) activity
with the 10-bp recognition sequence,
5'-GmATnnnGmA/-TCnnnA-TC-3'
3'-C-TAnnnC-T/mAGnnnTmAG-5'.
This specificity could be achieved by first methylating a substrate DNA with M.MamI in vivo,
followed by in vitro R.DpnI restriction.

<>

<1>Striebel, H.-M., Schmitz, G.G., Kaluza, K., Jarsch, M., Kessler, C.
<2>MamI, a novel class-II restriction endonuclease from Microbacterium ammoniaphilum recognizing 5'-GATNN^NNATC-3'.
<3>Gene
<4>91
<5>95-100
<6>1990
<7>A new site-specific class-II restriction endonuclease, MamI, has been discovered in the
nonsporulating Gram-positive Microbacterium ammoniaphilum. MamI recognition sequence and
cleavage positions were deduced using experimental and computer-assisted mapping and
sequencing approaches. MamI cleavage specificity corresponds to: 5'-GATNN^NNATC-3'
3'-CTANN^NNTAG-5'. The novel 43-kD enzyme recognizes a palindromic hexanucleotide
interrupted by four ambiguous nucleotides. MamI cleavage positions are both located in the
center of the recognition sequence resulting in blunt-ended fragments after cleavage in the
presence of Mg2+ ions. MamI is inhibited by N6-methyladenine residues. In case of overlapping
sequences of MamI and Escherichia coli-coded DNA modification methyltransferase M.EcodamI
(5'-GmATC-3'), cleavage of DNA isolated from E. coli wild-type cells will be inhibited. By
applying incubation conditions forcing star activity, relaxation of MamI sequence specificity
is observed (MamI*).

<>

<1>Striebel, H.-M., Seeber, S., Jarsch, M., Kessler, C.
<2>Cloning and characterization of the MamI restriction-modification system from Microbacterium ammoniaphilum in Escherichia coli.
<3>Gene
<4>172
<5>41-46
<6>1996
<7>The genes encoding a class-IIN restriction-modification (R-M) system (MamI,
sequence specificity 5'-GATnn/nnATC-3' / 3'-CTAnn/nnTAG-5' from Microbacterium
ammoniaphilum have been cloned in Escherichia coli.  The vector used for cloning was plasmid
pUC18 modified by the inclusion of three MamI recognition sites.  Recombinant clones
containing
the mamIM gene in its genomic context became fully methylated in vivo and remained completely
resistant against digestion with the R.MamI restriction enodnuclease (Enase).  Determination
of the
nucleotide (nt) sequence revealed three open reading frames with lengths of 1089 bp (ORF1),
276
bp (ORFc) and 927 bp (ORF2).  On the basis of expression and deletion experiments, the 1089-bp
ORF1 was assigned to mamIM encoding the M.MamI DNA methyltransferase (Mtase).  By amino
acid sequencing of the N terminus of R.MamI and comparison of the deduced nt sequence with
ORF2, the 927-bp ORF2 was idenified as the mamIR gene encoding R.MamI.  The 276-bp Orfc,
located between mamIR and mamIM, is part of the DNA sequence downstream from mamIM
shown to be necessary for controlled mamIM expression.

<>

<1>Strikhanov, S.N., Aleshkin, G.I., Skavronskaya, A.G.
<2>Plasmid recombination stimulated by restriction endonuclease EcoRI in vivo: specificity of recombinant plasmid formation in RecA cells of Escherichia coli.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>5
<5>13-19
<6>1985
<7>The restriction endonuclease EcoRI dependent recombination of compatible plasmids has been
studied in RecA cells of Escherichia coli.  Plasmid RP4 and the isogenic ColE1 type plasmids
pSA14 or pSA25, differing in restriction-modification RM EcoRI genes, have been used to study
this type of recombination.  EcoRI dependent recombination of plasmids is demonstrated in RecA
cells and, thus, is independent of general system of homologous recombination.  The classes of
recombinant plasmids isolated from RecA cells differ from the classes isolated from wild type
cells.  Levels of tetracyclin resistance conferred by plasmid RP4 are shown to be dependent on
the alleles of recA+ gene, being extremely low in RecA cells.  This property is demonstrated
to be useful for obtaining the multicopy recombinant plasmids resulting from EcoRI dependent
recombination in RecA cells of Escherichia coli.

<>

<1>Stringer, S.C., Carter, A.T., Webb, M.D., Wachnicka, E., Crossman, L.C., Sebaihia, M., Peck, M.W.
<2>Genomic and physiological variability within Group II (non-proteolytic) Clostridium botulinum.
<3>BMC Genomics
<4>14
<5>333
<6>2013
<7>BACKGROUND: Clostridium botulinum is a group of four physiologically and
phylogenetically distinct bacteria that produce botulinum neurotoxin. While
studies have characterised variability between strains of Group I (proteolytic)
C. botulinum, the genetic and physiological variability and relationships between
strains within Group II (non-proteolytic) C. botulinum are not well understood.
In this study the genome of Group II strain C. botulinum Eklund 17B (NRP) was
sequenced and used to construct a whole genome DNA microarray. This was used in a
comparative genomic indexing study to compare the relatedness of 43 strains of
Group II C. botulinum (14 type B, 24 type E and 5 type F). These results were
compared with characteristics determined from physiological tests. RESULTS: Whole
genome indexing showed that strains of Group II C. botulinum isolated from a wide
variety of environments over more than 75 years clustered together indicating the
genetic background of Group II C. botulinum is stable. Further analysis showed
that strains forming type B or type F toxin are closely related with only toxin
cluster genes targets being unique to either type. Strains producing type E toxin
formed a separate subset. Carbohydrate fermentation tests supported the
observation that type B and F strains form a separate subset to type E strains.
All the type F strains and most of type B strains produced acid from amylopectin,
amylose and glycogen whereas type E strains did not. However, these two subsets
did not differ strongly in minimum growth temperature or maximum NaCl
concentration for growth. No relationship was found between tellurite resistance
and toxin type despite all the tested type B and type F strains carrying tehB,
while the sequence was absent or diverged in all type E strains. CONCLUSIONS:
Although Group II C. botulinum form a tight genetic group, genomic and
physiological analysis indicates there are two distinct subsets within this
group. All type B strains and type F strains are in one subset and all type E
strains in the other.

<>

<1>Strittmatter, A.W. et al.
<2>Genome sequence of Desulfobacterium autotrophicum HRM2, a marine sulfate reducer oxidizing organic carbon completely to carbon dioxide.
<3>Environ. Microbiol.
<4>11
<5>1038-1055
<6>2009
<7>Summary Sulfate-reducing bacteria (SRB) belonging to the metabolically
versatile Desulfobacteriaceae are abundant in marine sediments and
contribute to the global carbon cycle by complete oxidation of organic
compounds. Desulfobacterium autotrophicum HRM2 is the first member of this
ecophysiologically important group with a now available genome sequence.
With 5.6 megabasepairs (Mbp) the genome of Db. autotrophicum HRM2 is about
2 Mbp larger than the sequenced genomes of other sulfate reducers (SRB). A
high number of genome plasticity elements (> 100 transposon-related
genes), several regions of GC discontinuity and a high number of
repetitive elements (132 paralogous genes Mbp(-1)) point to a different
genome evolution when comparing with Desulfovibrio spp. The metabolic
versatility of Db. autotrophicum HRM2 is reflected in the presence of
genes for the degradation of a variety of organic compounds including
long-chain fatty acids and for the Wood-Ljungdahl pathway, which enables
the organism to completely oxidize acetyl-CoA to CO(2) but also to grow
chemolithoautotrophically. The presence of more than 250 proteins of the
sensory/regulatory protein families should enable Db. autotrophicum HRM2
to efficiently adapt to changing environmental conditions. Genes encoding
periplasmic or cytoplasmic hydrogenases and formate dehydrogenases have
been detected as well as genes for the transmembrane TpII-c(3), Hme and
Rnf complexes. Genes for subunits A, B, C and D as well as for the
proposed novel subunits L and F of the heterodisulfide reductases are
present. This enzyme is involved in energy conservation in methanoarchaea
and it is speculated that it exhibits a similar function in the process of
dissimilatory sulfate reduction in Db. autotrophicum HRM2.

<>

<1>Strnad, H., Lapidus, A., Paces, J., Ulbrich, P., Vlcek, C., Paces, V., Haselkorn, R.
<2>Complete Genome Sequence of the Photosynthetic Purple Nonsulfur Bacterium Rhodobacter capsulatus SB 1003.
<3>J. Bacteriol.
<4>192
<5>3545-3546
<6>2010
<7>Rhodobacter capsulatus SB 1003 belongs to the group of purple nonsulfur bacteria. Its genome
consists of a 3.7-Mb chromosome and a 133-kb plasmid.
The genome encodes genes for photosynthesis, nitrogen fixation,
utilization of xenobiotic organic substrates, and synthesis of
polyhydroxyalkanoates. These features made it a favorite research tool for
studying these processes. Here we report its complete genome sequence.

<>

<1>Strnad, H., Patek, M., Fousek, J., Szokol, J., Ulbrich, P., Nesvera, J., Paces, V., Vlcek, C.
<2>Genome Sequence of Rhodococcus erythropolis Strain CCM2595, a Phenol Derivative-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e00208-14
<6>2014
<7>We announce the completion of the genome sequence of a phenol derivative-degrading bacterium,
Rhodococcus erythropolis strain CCM2595. This
bacterium is interesting in the context of bioremediation for its capability to
degrade phenol, catechol, resorcinol, hydroxybenzoate, hydroquinone,
p-chlorophenol, p-nitrophenol, pyrimidines, and sterols.

<>

<1>Strnad, H., Ridl, J., Paces, J., Kolar, M., Vlcek, C., Paces, V.
<2>Complete Genome Sequence of the Haloaromatic Acids-degrading Bacterium Achromobacter xylosoxidans A8.
<3>J. Bacteriol.
<4>193
<5>791-792
<6>2010
<7>Achromobacter xylosoxidans strain A8 was isolated from soil contaminated with polychlorinated
biphenyls. It can use 2-chlorobenzoate and
2,5-dichlorobenzoate as sole sources of carbon and energy. This property
makes it a good starting microorganism for further development towards a
bioremediation tool. The genome of A. xylosoxidans consists of a 7-Mb
chromosome and two large plasmids (98 kb, 248 kb). Besides genes for
utilization of xenobiotic organic substrates it encodes genes associated
with pathogenesis, toxin production and resistance. Here we report its
complete genome sequence.

<>

<1>Strobel, S.A., Doucette-Stamm, L.A., Riba, L., Housman, D.E., Dervan, P.B.
<2>Site-specific cleavage of human chromosome 4 mediated by triple-helix formation.
<3>Science
<4>254
<5>1639-1642
<6>1991
<7>Direct physical isolation of specific DNA segments from the human genome is a
necessary goal in human genetics.  For testing whether triple-helix mediated
enzymatic cleavage can liberate a specific segment of a human chromosome, the
tip of human chromosome 4, which contains the entire candidate region for the
Huntington's disease gene, was chosen as a target.  A 16-base pyrimidine
oligodeoxyribonucleotide was able to locate a 16-base pair purine target site
within more than 10 gigabase pairs of genomic DNA and mediate the exact
enzymatic cleavage at that site in more than 80 percent yield.  The recognition
motif is sufficiently generalizable that most cosmids should contain a sequence
targetable by triple-helix formation.  This method may facilitate the
orchestrated dissection of human chromosomes from normal and affected
individuals into megabase sized fragments and facilitate the isolation of
candidate gene loci.

<>

<1>Strobel, T., Al-Dilaimi, A., Blom, J., Gessner, A., Kalinowski, J., Luzhetska, M., Puhler, A., Szczepanowski, R., Bechthold, A., Ruckert, C.
<2>Complete genome sequence of Saccharothrix espanaensis DSM 44229T and comparison to the other completely sequenced Pseudonocardiaceae.
<3>BMC Genomics
<4>13
<5>465
<6>2012
<7>ABSTRACT: BACKGROUND: The genus Saccharothrix is a representative of the family
Pseudonocardiaceae, known to include producer strains of a wide variety of potent
antibiotics. Saccharothrix espanaensis produces both saccharomicins A and B of
the promising new class of heptadecaglycoside antibiotics, active against both
bacteria and yeast. RESULTS: To better assess its capabilities, the complete
genome sequence of S. espanaensis was established. With a size of 9,360,653 bp,
coding for 8,501 genes, it stands alongside other Pseudonocardiaceae with large
genomes. Besides a predicted core genome of 810 genes shared in the family, S.
espanaensis has a large number of accessory genes: 2,967 singletons when compared
to the family, of which 1,292 have no clear orthologs in the RefSeq database. The
genome analysis revealed the presence of 26 biosynthetic gene clusters
potentially encoding secondary metabolites. Among them, the cluster coding for
the saccharomicins could be identified. CONCLUSION: S. espanaensis is the first
completely sequenced species of the genus Saccharothrix. The genome discloses the
cluster responsible for the biosynthesis of the saccharomicins, the largest
oligosaccharide antibiotic currently identified. Moreover, the genome revealed 25
additional putative secondary metabolite gene clusters further suggesting the
strain's potential for natural product synthesis.

<>

<1>Strobl, J.S., Thompson, E.B.
<2>Methylation of either cytosine in the recognition sequence CGCG inhibits ThaI cleavage of DNA.
<3>Nucleic Acids Res.
<4>12
<5>8073-8083
<6>1984
<7>ThaI (CGCG) sites which overlap HhaI (GCGC) sites in PhiX174 and pBR322 DNA
were methylated in vitro with HhaI methylase and S-adenosylmethionine to yield
CGmCG, mCGmCG (5-methylcytosine, mC).  Methylation of either cytosine in the
ThaI recognition sequence rendered the DNA resistant to ThaI cleavage.  Rat
pituitary cell genomic DNA was digested with ThaI or 2 other known
methylation-sensitive enzymes, AvaI or XhoI.  After electrophoresis and
ethidium bromide staining of the DNA, all 3 enzymes showed the infrequent DNA
cleavage characteristic of methylation-sensitive enzymes.  Comparison of
pituitary growth hormone (GH) genes bearing strain-specific degrees of
methylation showed the less methylated gene to be more frequently cut by either
AvaI or ThaI.  ThaI resistant sites in GH genes were cleaved by ThaI after
exposing cells to 5-azacytidine, an inhibitor of DNA methylation.  We conclude
that ThaI is a useful restriction enzyme for the analysis of mC at CGCG
sequences in eukaryotic DNA.

<>

<1>Strous, M. et al.
<2>Deciphering the evolution and metabolism of an anammox bacterium from a community genome.
<3>Nature
<4>440
<5>790-794
<6>2006
<7>Anaerobic ammonium oxidation (anammox) has become a main focus in
oceanography and wastewater treatment.  It is also the nitrogen cycle's
major remaining biochemical enigma.  Among its features, the occurrence of
hydrazine as a free intermediate of catabolism, the biosynthesis of
ladderane lipids and the role of cytoplasm differentiation are unique in
biology.  Here we use environmental genomics - the reconstruction of
genomic data directly from the environment - to assemble the genome of the
uncultured anammox bacterium Kuenenia stuttgartiensis from a complex
bioreactor community.  The genome data illuminate the evolutionary history
of the Planctomycetes and allow us to expose the genetic blueprint of the
organism's special properties.  Most significantly, we identified candidate
genes responsible for ladderane biosynthesis and biological hydrazine
metabolism, and discovered unexpected metabolic versatility.

<>

<1>Strzelecka, T., Dorner, L., Schildkraut, I., Aggarawal, A.
<2>Structural studies of the BamHI restriction enzyme.
<3>Biophys. J.
<4>57
<5>68
<6>1990
<7>Type II restriction enzymes are ideal for studying protein-DNA interactions
because of their high sequence specificity and striking variety.  Large,
well-ordered BamHI crystals, diffracting to 2.3 angstrom, were obtained from
PEG solutions.  These crystals occur in two forms:  monoclinic (space group C2,
unit cell constants: a=76.24 angstrom, b=46.0 angstrom, c=69.4 angstrom and
beta=110.5 degrees) and orthorhombic (space group C222/1, unit cell constants:
a=46.7 angstrom, b=76.6 angstrom and c=143.6 angstrom).  In both crystal forms
there is one protein monomer per asymmetric unit.  Currently, we are searching
for heavy atom derivatives of the enzyme.  We are also attempting
cocrystallization of the enzyme with a 12-bp DNA fragment containing the BamHI
recognition site (5'-GGATCC-3').  We have recently obtained large, plate-like
crystals, which we are exploring for the presence of DNA by X-ray and
biochemical methods.

<>

<1>Strzelecka, T., Newman, M., Dorner, L.F., Knott, R., Schildkraut, I., Aggarwal, A.K.
<2>Crystallization and preliminary X-ray analysis of restriction endonuclease BamHI-DNA complex.
<3>J. Mol. Biol.
<4>239
<5>430-432
<6>1994
<7>Restriction endonuclease BamHI from Bacillus amyloliquefaciens has been co-crystallized with a
12 bp DNA fragment that encompasses its recognition site. The co-crystals diffract to at least
1.95 A resolution and belong to space group P212121. The unit cell parameters are a=108.8 A,
b=81.9 A, c=68.8 A, consistent with one complex in the crystallographic asymmetric unit. The
direction of the DNA appears to be along the b axis. In order to achieve end to end stacking
of DNA, the complex must lie on the screw axis along b. A self-rotation function has
determined the directions of the non-crystallographic 2-fold axes.

<>

<1>Stucken, K., John, U., Cembella, A., Murillo, A.A., Soto-Liebe, K., Fuentes-Valdes, J.J., Friedel, M., Plominsky, A.M., Vasquez, M., Glockner, G.
<2>The smallest known genomes of multicellular and toxic cyanobacteria: comparison, minimal gene sets for linked traits and the evolutionary implications.
<3>PLoS ONE
<4>5
<5>E9235
<6>2010
<7>Cyanobacterial morphology is diverse, ranging from unicellular spheres or
rods to multicellular structures such as colonies and filaments.
Multicellular species represent an evolutionary strategy to differentiate
and compartmentalize certain metabolic functions for reproduction and
nitrogen (N(2)) fixation into specialized cell types (e.g. akinetes,
heterocysts and diazocytes). Only a few filamentous, differentiated
cyanobacterial species, with genome sizes over 5 Mb, have been sequenced.
We sequenced the genomes of two strains of closely related filamentous
cyanobacterial species to yield further insights into the molecular basis
of the traits of N(2) fixation, filament formation and cell
differentiation. Cylindrospermopsis raciborskii CS-505 is a
cylindrospermopsin-producing strain from Australia, whereas Raphidiopsis
brookii D9 from Brazil synthesizes neurotoxins associated with paralytic
shellfish poisoning (PSP). Despite their different morphology, toxin
composition and disjunct geographical distribution, these strains form a
monophyletic group. With genome sizes of approximately 3.9 (CS-505) and
3.2 (D9) Mb, these are the smallest genomes described for free-living
filamentous cyanobacteria. We observed remarkable gene order conservation
(synteny) between these genomes despite the difference in repetitive
element content, which accounts for most of the genome size difference
between them. We show here that the strains share a specific set of 2539
genes with >90% average nucleotide identity. The fact that the CS-505 and
D9 genomes are small and streamlined compared to those of other
filamentous cyanobacterial species and the lack of the ability for
heterocyst formation in strain D9 allowed us to define a core set of genes
responsible for each trait in filamentous species. We presume that in
strain D9 the ability to form proper heterocysts was secondarily lost
together with N(2) fixation capacity. Further comparisons to all available
cyanobacterial genomes covering almost the entire evolutionary branch
revealed a common minimal gene set for each of these cyanobacterial
traits.

<>

<1>Stucken, K., Koch, R., Dagan, T.
<2>Cyanobacterial defense mechanisms against foreign DNA transfer and their impact on genetic engineering.
<3>Biol. Res.
<4>46
<5>373-382
<6>2013
<7>Cyanobacteria display a large diversity of cellular forms ranging from unicellular to complex
multicellular filaments or aggregates. Species in the group present a wide range of metabolic
characteristics including the fixation of atmospheric nitrogen, resistance to extreme
environments, production of hydrogen, secondary metabolites and exopolysaccharides. These
characteristics led to the growing interest in cyanobacteria across the fields of ecology,
evolution, cell biology and biotechnology. The nu ber of available cyanobacterial genome
sequences has increased considerably in recent years, with more than 140 fully sequenced
genomes to date. Genetic engineering of cyanobacteria is widely applied to the model
unicellular strains Synechocystis sp. PCC 6803 and Synechococcus elongatus PCC 7942. However
the establishment of transformation protocols in many other cyanobacterial strains is
challenging. One obstacle to the development of these novel model organisms is that many
species have doubling times of 48 h or more, much longer than the bacterial models E. coli or
B. subtilis. Furthermore, cyanobacterial defense mechanisms against foreign DNA pose a
physical and biochemical barrier to DNA insertion in most strains. Here we review the various
barriers to DNA uptake in the context of lateral gene transfer among microbes and the various
mechanisms for DNA acquisition within the prokaryotic domain. Understanding the cyanobacterial
defense mechanisms is expected to assist in the development and establishment of novel
transformation protocols that are specifically suitable for this group.

<>

<1>Stuckle, E.E., Emmrich, C., Grob, U., Nielsen, P.J.
<2>Statistical analysis of nucleotide sequences.
<3>Nucleic Acids Res.
<4>18
<5>6641-6647
<6>1990
<7>In order to scan nucleic acid databases for potentially relevant but as yet
unknown signals, we have developed an improved statistical model for pattern
analysis of nucleic acid sequences by modifying previous methods based on
Markov chains.  We demonstrate the importance of selecting the appropriate
parameters in order for the method to function at all.  The model allows the
simultaneous analysis of several short sequences with unequal base frequencies
and Markov order k+0 as is usually the case in databases.  As a test of these
modifications, we show that in E. coli sequences there is a bias against
palindromic hexamers which correspond to known restriction enzyme recognition
sites.

<>

<1>Studier, F.W.
<2>Gene 0.3 of Bacteriophage T7 Acts to Overcome the DNA Restriction System of the Host.
<3>J. Mol. Biol.
<4>94
<5>283-295
<6>1975
<7>Wild-type bacteriophage T7 is not subject to restriction by the Escherichia
coli B and K restriction systems, but T7 mutants that are susceptible to such
restriction have been isolated.  These mutants are all defective in gene 0.3,
the first T7 gene to be expressed after infection.  The gene 0.3 protein
apparently acts to prevent modification as well as restriction, suggesting that
it may interact with a component of the host restriction-modification system
that is required for both processes.  Mutants in which gene 0.3 is completely
deleted are only partially modified by growth on hosts with an active
restriction-modification system, presumably because the conditions of T7
infection overload the modifying capacity of the cells.  This is in contrast to
phages such as lambda that are completely modified during growth.  Since gene
0.3 is not essential for growth in non-restricting hosts, it has been possible
to isolate deletions which extend to the left of gene 0.3 into the region where
E. coli RNA polymerase initiates the synthesis of T7 early RNA.  Two of the
three strong initiators from which E. coli RNA polymerase transcribes the early
region can be deleted.  In the course of searching for T7 mutants that are
unable to overcome restriction, it was discovered that mutants defective in
gene 2 are able to plate on E. coli C with essentially normal efficiency, and
most gene 7 mutants are able to plate on both C and K strains.  It has not been
determined why genes 2 and 7 seem to be needed for growth in some E. coli
strains but not in others.

<>

<1>Studier, F.W., Bandyopadhyay, P.K.
<2>Model for how type I restriction enzymes select cleavage sites in DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>85
<5>4677-4681
<6>1988
<7>Under appropriate conditions, digestion of phage T7 DNA by the type I
restriction enzyme EcoK produces an orderly progression of discrete DNA
fragments.  All details of the fragmentation pattern can be explained on the
basis of the known properties of type I enzymes, together with two further
assumptions:  (i) in the ATP-stimulated translocation reaction, the enzyme
bound at the recognition sequence translocates DNA toward itself from both
direction simultaneously; and (ii) when translocation causes neighboring
enzymes to meet, they cut the DNA between them.  The kinetics of digestion at
37C indicates that the rate of translocation of DNA from each side of a bound
enzyme is about 200 base pairs per second, and the cuts are completed within
15-25 sec of the time neighboring enzymes meet.  The resulting DNA fragments
each contain a single recognition site with an enzyme (or subunit) remaining
bound to it.  At high enzyme concentrations, such fragments can be further
degraded, apparently by cooperation between the specifically bound and excess
enzymes.  This model is consistent with a substantial body of previous work on
the nuclease activity of EcoB and EcoK, and it explains in a simple way how
cleavage sites are selected.

<>

<1>Studier, F.W., Movva, N.R.
<2>SAMase gene of bacteriophage T3 is responsible for overcoming host restriction.
<3>J. Virol.
<4>19
<5>136-145
<6>1976
<7>Deletion and point mutants of T3 have been isolated and used to show that the
early region of T3 DNA is organized in the same way as that of T7 DNA.
Homologous early RNAs and proteins of the two phages have been identified by
electrophoresis on polyacrylamide gels in the presence of sodium dodecyl
sulfate.  Both phages have five early mRNA's, numbered 0.3, 0.7, 1,1.1, and 1.3
from left to right, although no T3 protein that corresponds to the 1.1 protein
of T7 has yet been identified.  In general, corresponding early RNAs and
proteins of the two phages migrate differently on gels, indicating that they
differ in molecular weight and/or conformation.  In both T7 and T3, gene 0.3 is
responsible for overcoming the DNA restriction system of the host, gene 0.7
specifies a protein kinase, gene 1 specifies a phage-specific RNA polymerase,
and gene 1.3 specifies a polynucleotide ligase.  The 0.3 protein of T3 is
responsible for the S-adenosylmethionine cleaving activity (SAMase) induced
after T3 (but not T7) infection.  However, cleaving of S-adenosylmethionine
does not appear to be the primary mechanism by which T3 overcomes host
restriction, since at least one mutant of T3 has lost the SAMase activity
without losing the ability to overcome host restriction.

<>

<1>Sturino, J.M., Rajendran, M., Altermann, E.
<2>Draft Genome Sequence of Lactobacillus animalis 381-IL-28.
<3>Genome Announcements
<4>2
<5>e00478-14
<6>2014
<7>Lactobacillus animalis 381-IL-28 is an integral component of a multistrain commercial culture
with food biopreservative and pathogen biocontrol
functionality. A draft sequence of the L. animalis 381-IL-28 genome is described
in this paper.

<>

<1>Sturino, J.M., Rajendran, M., Altermann, E.
<2>Draft Genome Sequence of the Pediocin-Encoding Biopreservative and Biocontrol Strain Pediococcus acidilactici D3.
<3>Genome Announcements
<4>1
<5>e00208-13
<6>2013
<7>We describe a draft genome sequence for Pediococcus acidilactici strain D3, a component of
multistrain commercial cultures with biopreservative and biocontrol
properties in food-based applications. Strain D3 encodes at least one
antimicrobial peptide, pediocin AMPd3. The AMPd3-encoding operon exhibits high
sequence similarity to the archetype pediocin, PA-1, encoded by P. acidilactici
PAC 1.0.

<>

<1>Sturm, G., Buchta, K., Kurz, T., Rensing, S.A., Gescher, J.
<2>Draft Genome Sequence of Leucobacter chromiiresistens, an Extremely Chromium-Tolerant Strain.
<3>J. Bacteriol.
<4>194
<5>540-541
<6>2012
<7>Here we present the draft genome of Leucobacter chromiiresistens. This is the first genome
sequence of an organism belonging to the genus
Leucobacter. L. chromiiresistens was sequenced due to its capability to
tolerate up to 300 mM Cr(VI) in the medium, which is so far a unique
feature for microorganisms.

<>

<1>Sturm, R.A., Yaciuk, P.
<2>DNA cleavage by restriction endonucleases PflMI is inhibited in recognition sites modified by dcm methylation.
<3>Nucleic Acids Res.
<4>17
<5>3615
<6>1989
<7>None

<>

<1>Sturrock, S.S., Dryden, D.T., Atanasiu, C., Dornan, J., Bruce, S., Cronshaw, A., Taylor, P., Walkinshaw, M.D.
<2>Crystallization and preliminary X-ray analysis of ocr, the product of gene 0.3 of bacteriophage T7.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>57
<5>1652-1654
<6>2001
<7>Ocr, the product of gene 0.3 of bacteriophage T7, prevents the action of restriction
endonucleases of the host bacteria. The amino-acid sequence of
ocr has less than 20% similarity to any protein of known three-dimensional
structure. Ocr has been crystallized in a number of different crystal
forms and X-ray data for the seleno-L-methionine-substituted form has been
collected to a resolution of 1.8 A. The presence of caesium was found to
be required for good crystal growth. Anomalous X-ray data was used to
identify possible positions for Se and Cs atoms in the unit cell.

<>

<1>Sturrock, S.S., Dryden, D.T.F.
<2>A prediction of the amino acids and structures involved in DNA recognition by type I DNA restriction and modification enzymes.
<3>Nucleic Acids Res.
<4>25
<5>3408-3414
<6>1997
<7>The S subunits of type I DNA restriction/modification enzymes are responsible for recognizing
the DNA target sequence for the enzyme.  They contain two domains of approximately 150 amino
acids, each of which is responsible for recognizing one half of the bipartite asymmetric
target.  In the absence of any known tertiary structure for type I enzymes or recognizable DNA
recognition motifs in the highly variable amino acid sequences of the S subunits, it has
previously not been possible to predict which amino acids are responsible for sequence
recognition.  Using a combination of sequence alignment and secondary structure prediction
methods to analyze the sequences of S subunits, we predict that all of the 51 known target
recognition domains have the same tertiary structure.  Furthermore, this structure is similar
to the structure of the TRD of the C5-cytosine methyltransferase, HhaI, which recognizes its
DNA target via interactions with two short polypeptide loops and a beta strand.  Our results
predict the location of these sequence recognition structures within the TRDs of all type I S
subunits.

<>

<1>Stuy, J.H.
<2>Restriction Enzymes Do Not Play a Significant Role in Haemophilus Homospecific or Heterospecific Transformation.
<3>J. Bacteriol.
<4>128
<5>212-220
<6>1976
<7>Competent Haemophilus influenzae Rd recipients, either as phage HP1 restricting
(r+) or nonrestricting (r-) nonlysogens or defective lysogens, were exposed to
deoxyribonucleic acids from various wild-type phage HP1 lysogenic H. influenzae
serotype strains (non-encapsulated derivatives of serotypes alpha, beta, c, d,
and e), to DNA from lysogenic Haemophilus parahaemolyticus, and to DNA from
modified and nonmodified phage HP1.  Transformation of antibiotic resistance
markers and of prophage markers in homospecific crosses was observed to be
unaffected by the recipient restriction phenotype, whereas the transfection
response was much reduced in r+ recipients.  Heterospecific transformation of
prophage markers was reduced by only 80 to 90%, whereas antibiotic resistance
marker transformation was 1,000 to 10,000 times lower.  Heterospecific
transfection was at least 100 times lower than homospecific transfection in
both r+ and r- recipients.  The general conclusion is that neither class I nor
class II restriction enzymes affect significantly the transformation efficiency
in homospecific and heterospecific crosses.  The efficiency of heterospecific
transformation may depend mainly on the deoxyribonucleic acid homology in the
genetic marker region.

<>

<1>Stynen, A.P. et al.
<2>Complete Genome Sequence of Type Strain Campylobacter fetus subsp. venerealis NCTC 10354T.
<3>J. Bacteriol.
<4>193
<5>5871-5872
<6>2011
<7>Campylobacter fetus subsp. venerealis is the etiologic agent of bovine genital
campylobacteriosis, a sexually transmitted disease of cattle that
is of worldwide importance. The complete sequencing and annotation of the
genome of the type strain C. fetus subsp. venerealis NCTC 10354(T) are
reported.

<>

<1>Styriak, I., Pristas, P., Javorsky, P.
<2>Lack of surface receptors not restriction-modification system determines F4 phage resistance in Streptococcus bovis II/1.
<3>Folia Microbiol. (Praha)
<4>43
<5>35-38
<6>1998
<7>The resistance of Streptococcus bovis strain II/1, the producer of SbvI restriction
endonuclease, to F4 phage infection was demonstrated by the double-agar-layer method.  Despite
the presence of restriction endonuclease SbvI which can cleave F4 phage DNA to numerous
fragments in vitro, the evidence that adsorption inhibition is the most important defense
mechanism in phage resistance of S. bovis II/1 strain was obtained by adhesion experiments in
vivo.  Electron microscopy of phage-host mixtures showed many phage particles on the bacterial
surface of phage-sensitive S. bovis 47/3 control strain in comparison with no phage particles
seen on S. bovis II/1 (phage-resistant) strain surface.

<>

<1>Su, C.C., Deng, W.L., Jan, F.J., Chang, C.J., Huang, H., Chen, J.
<2>Draft Genome Sequence of Xylella fastidiosa Pear Leaf Scorch Strain in Taiwan.
<3>Genome Announcements
<4>2
<5>e00166-14
<6>2014
<7>The draft genome sequence of Xylella fastidiosa pear leaf scorch strain PLS229, isolated from
the pear cultivar Hengshan (Pyrus pyrifolia) in Taiwan, is reported
here. The bacterium has a genome size of 2,733,013 bp, with a G+C content of
53.1%. The PLS229 genome was annotated and has 3,259 open reading frames and 50
RNA genes.

<>

<1>Su, F., Hua, D., Zhang, Z., Wang, X., Tang, H., Tao, F., Tai, C., Wu, Q., Wu, G., Xu, P.
<2>Genome Sequence of Bacillus pumilus S-1, an Efficient Isoeugenol-Utilizing Producer for Natural Vanillin.
<3>J. Bacteriol.
<4>193
<5>6400-6401
<6>2011
<7>Bacillus pumilus S-1 is an efficient isoeugenol-utilizing producer of natural vanillin. The
genome of B. pumilus S-1 contains the epoxide
hydrolase and six candidate monooxygenases that make it possible to
explore the mechanism involved in conversion of isoenguenol to vanillin in
the B. pumilus strain.

<>

<1>Su, F., Tao, F., Tang, H., Xu, P.
<2>Genome Sequence of the Thermophile Bacillus coagulans Hammer, the Type Strain of  the Species.
<3>J. Bacteriol.
<4>194
<5>6294-6295
<6>2012
<7>Here we announce a 3.0-Mb assembly of the Bacillus coagulans Hammer strain, which is the type
strain of the species within the genus Bacillus. Genomic analyses
based on the sequence may provide insights into the phylogeny of the species and
help to elucidate characteristics of the poorly studied strains of Bacillus
coagulans.

<>

<1>Su, F., Xu, K., Zhao, B., Tai, C., Tao, F., Tang, H., Xu, P.
<2>Genome Sequence of the Thermophilic Strain Bacillus coagulans XZL4, an Efficient Pentose-Utilizing Producer of Chemicals.
<3>J. Bacteriol.
<4>193
<5>6398-6399
<6>2011
<7>Bacillus coagulans XZL4 is an efficient pentose-utilizing producer of important platform
compounds, such as l-lactic acid, 2,3-butanediol, and
acetoin. Here we present a 2.8-Mb assembly of its genome. Simple and
efficient carbohydrate metabolism systems, especially the
transketolase/transaldolase pathway, make it possible to convert pentose
sugars to products at high levels.

<>

<1>Su, F., Yu, B., Sun, J., Ou, H.Y., Zhao, B., Wang, L., Qin, J., Tang, H., Tao, F., Jarek, M., Scharfe, M., Ma, C., Ma, Y., Xu, P.
<2>Genome Sequence of the Thermophilic Strain Bacillus coagulans 2-6, an Efficient Producer of High-Optical-Purity L-Lactic Acid.
<3>J. Bacteriol.
<4>193
<5>4563-4564
<6>2011
<7>Bacillus coagulans 2-6 is an efficient producer of lactic acid. The genome of B. coagulans 2-6
owns the smallest genome size among the members of genus Bacillus known to date. The
frameshift mutation at the start of D-lactate dehydrogenase might be responsible for the
production of high optical purity L-lactic-acid.

<>

<1>Su, H., Zhang, T., Bao, M., Jiang, Y., Wang, Y., Tan, T.
<2>Genome Sequence of a Promising Hydrogen-Producing Facultative Anaerobic Bacterium, Brevundimonas naejangsanensis Strain B1.
<3>Genome Announcements
<4>2
<5>e00542-14
<6>2014
<7>Brevundimonas naejangsanensis strain B1 is a newly isolated, facultative anaerobic bacterium
capable of producing hydrogen with high efficiency. Here, we
present a 2.94-Mb assembly of the genome sequence of strain B1, which may provide
further insights into the molecular mechanism of hydrogen production from
bioresource.

<>

<1>Su, J., Ye, J., Zhu, Y.
<2>Draft Genome Sequence of a Novel Bacterial Strain, LSJC7, Belonging to the Family Enterobacteriaceae with Dual Resistance to Arsenic and Tetracycline.
<3>J. Bacteriol.
<4>194
<5>7005-7006
<6>2012
<7>Strain LSJC7, with dual resistance to arsenic and tetracycline, was isolated from an antimony
tailing in China. Its 16S rRNA gene sequence has the highest
similarity to that of Enterobacter cloacae subsp. dissolvens LMG 2683(T)
(97.02%). Here we present the approximately 4.6-Mbp draft genome sequence of
strain LSJC7.

<>

<1>Su, L., Wang, T., Zhou, L., Wu, C., Guo, Y., Chang, D., Liu, Y., Jiang, X., Yin, S., Liu, C.
<2>Genome Sequence of Bacillus cereus Strain LCT-BC235, Carried by the Shenzhou VIII Spacecraft.
<3>Genome Announcements
<4>2
<5>e00665-13
<6>2014
<7>In order to explore the effect of space environments on Bacillus cereus, we determined the
draft genome sequence of a B. cereus strain, LCT-BC235, which was
isolated after space flight.

<>

<1>Su, L., Zhou, T., Zhou, L., Fang, X., Li, T., Wang, J., Guo, Y., Chang, D., Wang, Y., Li, D., Liu, C.
<2>Draft Genome Sequence of Bacillus cereus Strain LCT-BC244.
<3>J. Bacteriol.
<4>194
<5>3549
<6>2012
<7>Bacillus cereus is a prevalent, soil-dwelling, Gram-positive bacterium. Some strains are
harmful to humans and cause food-borne illness, while other strains
can be beneficial as probiotics for animals. To gain insight into the bacterial
genetic determinants, we report the genome sequence of a strain, LCT-BC244, which
was isolated from CGMCC 1.230.

<>

<1>Su, P., Im, H., Hsieh, H., Kanga, S., Dunn, N.W.
<2>LlaFI, a type III restriction and modification system in Lactococcus lactis.
<3>Appl. Environ. Microbiol.
<4>65
<5>686-693
<6>1999
<7>We describe a type III restriction and modification system, LlaFI, in Lactococcus lactis.
LlaFI is encoded by a 12-kb native plasmid, pND801, harbored in L. lactis LL42-1.  Sequencing
revealed two adjacent open reading frames.  One ORF encodes a 680-amino-acid polypeptide, and
this ORF is followed by a second ORF which encodes an 873-amino-acid polypeptide.  The two
ORFs appear to be organized in an operon.  A homology search revealed that the two ORFs
exhibited significant similarity to type III restriction and modification subunits.  The
complete amino acid sequence of the Mod subunit of LlaFI was aligned with the amino acid
sequences of four previously described type III methyltransferases.  Both the N-terminal
regions and the C-terminal regions of the Mod proteins are conserved, while the central
regions are more variable.  An S-adenosyl methionine binding motif (present in all adenine
methyltransferases) was found in the N-terminal region of the Mod protein.  The seven
conserved helicase motifs found in the previously described type III R/M systems were found at
the same relative positions in the LlaFI Res sequence, LlaFI has cofactor requirements for
activity that are characteristic of the previously described type III enzymes.  ATP and Mg2+
are required for endonucleolytic activity; however, the activity is not strictly dependent on
the presence of Ado-Met but is stimulated by it.  To our knowledge, this is the first type III
R/M system that has been characterized not just in lactic acid bacteria but also in
gram-positive bacteria.

<>

<1>Su, P., Ng, A., Kennelly, V., Costello, M., Harvey, M., Dunn, N.
<2>Additive effects of two different plasmid-linked restriction and modification systems in Lactococcus.
<3>Biotechnol. Lett.
<4>20
<5>515-518
<6>1998
<7>Two plasmids, pND801 and pND802, encoding different restriction and modification systems were
isolated from Lactococcus lactis ssp. lactis LL42-1 and Lactococcus lactis ssp. cremoris
LC14-1, respectively.  PND802 contained one SphI restriction enzyme site and the whole plasmid
was cloned into the SphI site of the streptococcal/E. coli shuttle vector pSA3 generating the
plasmid pND803.  PND803 was stably maintained in L. lactis MG1363 harboring pND801.  The
combination of the two R/M systems within L. lactis MG1363 resulted in an additive resistance
towards both isometric phage and prolate phage.

<>

<1>Su, S.Y., Shao, J.P., Monasu, R.E.
<2>Process for cloning and producing restriction endonuclease SapI in Escherichia coli.
<3>Japanese Patent Office
<4>JP 1998057082 A
<5>
<6>1998
<7>The present invention relates to recombinant DNA which encodes the NheI restriction
endonuclease as well as NheI methyltransferase, expression of NheI restriction endonuclease
from E. coli cells containing the recombinant DNA.  An internal NdeI site in the NheIR gene
was eliminated by a silent mutation.  A new NdeI site was engineered at the start codon of
NheIR gene.  An NdeI-BamHI fragment containing NheIR gene was cloned into a T7 expression
vector pAII17 and expressed in a premodified host ER2566 [pACYC-NheIM, pAII17-NheIR2].  The
recombinant clone produces approximately 10 million units of NheI per gram of wet cells.

<>

<1>Su, T.-J., Connolly, B.A., Darlington, C., Mallin, R., Dryden, D.T.F.
<2>Unusual 2-aminopurine fluorescence from a complex of DNA and the EcoKI methyltransferase.
<3>Nucleic Acids Res.
<4>32
<5>2223-2230
<6>2004
<7>The methyltransferase, M.EcoKI, recognizes the DNA sequence 5'-AACNNNNNNGTGC-3' and
methylates adenine at the underlined positions. DNA methylation has been shown by
crystallography to occur via a base flipping mechanism and is believed to be a general
mechanism for all methyltransferases. If no structure is available, the fluorescence of
2-aminopurine is often used as a signal for base flipping as it shows enhanced fluorescence
when its environment is perturbed. We find that 2-aminopurine gives enhanced fluorescence
emission not only when it is placed at the M.EcoKI methylation sites but also at a location
adjacent to the target adenine. Thus it appears that 2-aminopurine fluorescence intensity is
not a clear indicator of base flipping but is a more general measure of DNA distortion. Upon
addition of the cofactor S-adenosyl-methionine to the M.EcoKI:DNA complex, the 2-aminopurine
fluorescence changes to that of a new species showing excitation at 345 nm and emission at 450
nm. This change requires a fully active enzyme, the correct cofactor and the 2-aminopurine
located at the methylation site. However, the new fluorescent species is not a covalently
modified form of 2-aminopurine and we suggest that it represents a hitherto undetected
physicochemical form of 2-aminopurine.

<>

<1>Su, T.-J., Tock, M.R., Egelhaaf, S.U., Poon, W.C.K., Dryden, D.T.F.
<2>DNA bending by M.EcoKI methyltransferase is coupled to nucleotide flipping.
<3>Nucleic Acids Res.
<4>33
<5>3235-3244
<6>2005
<7>The maintenance methyltransferase M.EcoKI recognizes the bipartite DNA sequence
5'-AACNNNNNNGTGC-3', where N is any nucleotide. M.EcoKI preferentially methylates a sequence
already containing a methylated adenine at or complementary to the underlined bases in the
sequence. We find that the introduction of a single-stranded gap in the middle of the
non-specific spacer, of up to 4 nt in length, does not reduce the binding affinity of M.EcoKI
despite the removal of non-sequence-specific contacts between the protein and the DNA
phosphate backbone. Surprisingly, binding affinity is enhanced in a manner predicted by simple
polymer models of DNA flexibility. However, the activity of the enzyme declines to zero once
the single-stranded region reaches 4 nt in length. This indicates that the recognition of
methylation of the DNA is communicated between the two methylation targets not only through
the protein structure but also through the DNA structure. Furthermore, methylation recognition
requires base flipping in which the bases targeted for methylation are swung out of the DNA
helix into the enzyme. By using 2-aminopurine fluorescence as the base flipping probe we find
that, although flipping occurs for the intact duplex, no flipping is observed upon
introduction of a gap. Our data and polymer model indicate that M.EcoKI bends the non-specific
spacer and that the energy stored in a double-stranded bend is utilized to force or flip out
the bases. This energy is not stored in gapped duplexes. In this way, M.EcoKI can determine
the methylation status of two adenine bases separated by a considerable distance in
double-stranded DNA and select the required enzymatic response.

<>

<1>Su, Y.C., Horhold, F., Singh, B., Riesbeck, K.
<2>Complete Genome Sequence of Encapsulated Haemophilus influenzae Type f KR494, an  Invasive Isolate That Caused Necrotizing Myositis.
<3>Genome Announcements
<4>1
<5>e00470-13
<6>2013
<7>Haemophilus influenzae serotype f (Hif) is an etiologic agent of bacterial invasive disease.
Here, we report the first annotated genome sequence of the Hif
strain KR494, which was isolated from a patient suffering from sepsis and
necrotizing myositis. The genome sequence will increase the understanding of Hif
pathogenesis.

<>

<1>Suarez, N.E., Saavedra, L., Slozilova, I., Bonacina, J., Demnerova, K., Sesma, F.
<2>Draft Genome Sequence of Enterococcus faecium Strain CRL 1879, Isolated from a Northwestern Argentinian Artisanal Cheese.
<3>Genome Announcements
<4>1
<5>e00514-13
<6>2013
<7>We report the draft genome sequence of the bacteriocin producer Enterococcus faecium strain
CRL 1879, isolated from a northwestern Argentinian artisanal
cheese. The draft genome sequence is composed of 73 contigs for 2,886,747 bp,
with 3,140 protein-coding genes. Six biosynthetic clusters for bacteriocin class
II production were found. Typical virulence determinants, which have relevance in
food safety, were not present.

<>

<1>Suarez, V., Zago, M., Giraffa, G., Reinheimer, J., Quiberoni, A.
<2>Evidence for the presence of restriction/modification systems in Lactobacillus delbrueckii.
<3>J. Dairy Res.
<4>76
<5>433-440
<6>2009
<7>The bacteriophages Cb1/204 and Cb1/342 were obtained by induction from the commercial strain
Lactobacillus delbrueckii subsp. lactis Cb1, and
propagated on Lactobacillus delbrueckii subsp. lactis 204 (Lb.l 204)
and Lactobacillus delbrueckii subsp. bulgaricus 342 (Lb.b 342),
respectively. By cross sensitivity, it was possible to detect a delay
in the lysis of Lb.l 204 with Cb1/342 phage, while the adsorption rate
was high (99-5%). Modified and unmodified phages were isolated using
phage Cb1/342 and strain Lb.l 204. The EOP (Efficiency of Plaquing)
values for the four phages (Cb1/204, Cb1/342, Cb1/342 modified and
Cb1/342 unmodified) suggested that an R/M system modified the original
temperate phage, and the Bg/II-DNA restriction patterns of these phages
might point out the presence of a Type II R/M system. Also, the
existence of a Type I R/M system was demonstrated by PCR and nucleotide
sequence, being the percentages of alignment homology with Type I R/M
systems reported previously higher than 95%. In this study it was
possible to demonstrate that the native phage resistant mechanisms and
the occurrence of prophages in commercial host strains, contribute
strongly to diversify the phage population in a factory environment.

<>

<1>Suarez, V.B., Maciel, N., Guglielmotti, D., Zago, M., Giraffa, G., Reinheimer, J.
<2>Phage-resistance linked to cell heterogeneity in the commercial strain Lactobacillus delbrueckii subsp lactis Ab1.
<3>Int. J. Food Microbiol.
<4>128
<5>401-405
<6>2008
<7>The aim of this work was to study the relationship between the cell morphological
heterogeneity and the phage-resistance in the commercial
strain Lactobacillus delbrueckii subsp. lactis Ab1. Two morphological
variants (named C and T) were isolated from this strain.
Phage-resistant derivatives were isolated from them and the percentage
of occurrence of confirmed phage-resistant cells was 0.001% of the
total cellular population. Within these phage-resistant cell
derivatives there were T (3 out of 4 total isolates) and C (1 out of 4
total isolates) variants. The study of some technological properties
(e.g. proteolytic and acidifying activities) demonstrated that most of
phage-resistant derivatives were not as good as the parental strain.
However. for one derivative (a T variant), the technological properties
Were better than those of the parental strain. On the other hand, it
was possible to determinate that the system of phage-resistance in the
T variants was interference in adsorption step, with adsorption rates <
15%. For the C variant derivative it was possible to demonstrate the
presence of a restriction/modification system and, moreover, to
determinate that this system could be Type 1R/M.

<>

<1>Suarez-Suarez, L.Y., Brunet-Galmes, I., Pina-Villalonga, J.M., Christie-Oleza, J.A., Pena, A., Bennasar, A., Armengaud, J., Nogales, B., Bosch, R.
<2>Draft Genome Sequence of Citreicella aestuarii Strain 357, a Member of the Roseobacter Clade Isolated without Xenobiotic Pressure from a Petroleum-Polluted    Beach.
<3>J. Bacteriol.
<4>194
<5>5464-5465
<6>2012
<7>Citreicella aestuarii 357 is a member of the Roseobacter clade that was isolated  without
xenobiotic pressure from an oil-polluted sand sample from the Galician
coast (Spain). Its genome sequence suggests an organoheterotrophic metabolism,
including a wide catabolic potential for aromatic hydrocarbons.

<>

<1>Subach, F., Kirsanova, O., Liquier, J., Gromova, E.S.
<2>Resolution of the EcoRII restriction endonuclease-DNA complex structure in solution using fluorescence spectroscopy.
<3>Biophys. Chem.
<4>138
<5>107-114
<6>2008
<7>The X-ray structure for the type HE EcoRII restriction endonuclease has been resolved (X.E.
Zhou, Y. Wang, M. Reuter, M. Mucke, D.H. Kruger,
EJ. Meehan and L. Chen. Crystal structure of type HE restriction
enclonuclease EcoRII reveals an autoinhibition mechanism by a novel
effector-binding fold. J. Mol. Biol. 335 (2004) 307319.1, but the
structure of the R.EcoRII-DNA complex is still unknown. The aim of this
article was to examine the structure of the pre-reactive R.EcoRII-DNA
complex in solution by fluorescence spectroscopy. The structure for the
R.EcoRII-DNA complex was resolved by determining the fluorescence
resonance energy transfer (FRET) between two fluorescent dyes,
covalently attached near the EcoRII recognition sites, that were
located at opposite ends of a lengthy two-site DNA molecule. Analysis
of the FRET data from the two-site DNA revealed a likely model for the
arrangement of the two EcoRII recognition sites relative to each other
in the R.EcoRII-DNA complex in the presence of Ca2+ ions. According to
this model, the R.EcoRII binds the two-site DNA and forms a DNA loop in
which the EcoRII recognition sites are 20 +/- 10 A distant to each
other and situated at an angle of 70 +/- 10 degrees.

<>

<1>Subach, F.V., Liquier, J., Gromova, E.S.
<2>Investigation of restriction endonuclease EcoRII complex with DNA in solution by FTIR spectroscopy.
<3>Russ. J. Gen. Chem.
<4>78
<5>1103-1109
<6>2008
<7>The X-ray structure of type IIE EcoRII restriction endonuclease has been solved but the
structure of the R.EcoRII-DNA complex is still unknown. We report here on the structure of the
pre-reactive R.EcoRII-DNA-Ca2+ complex in solution examined by FTIR spectroscopy. The
secondary structure of R.EcoRII as well as the structure of the target DNA in the
R.EcoRII-DNA-Ca2+ complex was characterized. It was shown that the R.EcoRII-DNA-Ca2+ complex
formation is accompanied by changes in the spectrum of both DNA bases and DNA sugar-phosphate
backbone that suggest contacts of the enzyme with different groups of atoms in DNA. The change
of the R.EcoRII secondary structure in the R.EcoRII-DNA-Ca2+ complex is also observed.

<>

<1>Subach, F.V., Muller, S., Tashlitsky, V.N., Petrauskene, O.V., Gromova, E.S.
<2>The preparation of DNA duplexes containing internucleotide phosphorothioate groups in various positions of the recognition site for the EcoRII restriction endonuclease.
<3>Bioorg. Khim.
<4>29
<5>623-631
<6>2003
<7>Twenty-four 12-mer DNA duplexes, each containing a chiral phosphorothioate group successively
replacing one of the internucleotide phosphate groups
either in the EcoRII recognition site (5'CCA/TGG) or near to it, were
obtained for studying the interaction of the restriction endonuclease
EcoRII with internucleotide DNA phosphates. Twelve of the 12-mer
oligonucleotides were synthesized as Rp and Sp diastereomeric mixtures.
Six of them were separated by reversed-phase HPLC using various buffers.
Homogeneous diastereomers of the other oligonucleotides were obtained by
enzymatic ligation of the Rp and Sp diastereomers of 5- to 7-mer
oligonucleotides preliminarily separated by HPLC with the corresponding
short oligonucleotides on a complementary DNA template. The English
version of the paper: Russian Journal of Bioorganic Chemistry, 2003, vol.
29, no. 6; see also http://www.maik.ru.

<>

<1>Subach, O.M., Baskunov, V.B., Darii, M.V., Maltseva, D.V., Alexandrov, D.A., Kirsanova, O.V., Kolbanovskiy, A., Kolbanovskiy, M., Johnson, F., Bonala, R., Geacintov, N.E., Gromova, E.S.
<2>Impact of benzo[a] pyrene-2 '-deoxyguanosine lesions on methylation of DNA by SssI and HhaI DNA methyltransferases.
<3>Biochemistry
<4>45
<5>6142-6159
<6>2006
<7>DNA damage caused by the binding of the tumorigen 7R, 8S-diol 9S, 10R-epoxide (B[a]PDE), a
metabolite of bezo[a] pyrene, to guanine in
CpG dinucleotide sequences could affect DNA methylation and, thus,
represent a potential epigenetic mechanism of chemical carcinogenesis.
In this work, we investigated the impact of stereoisomeric (+)- and
(-)- trans-anti-B[a]P-N-2-dG adducts (B+ and B-) on DNA methylation by
prokaryotic DNA methyltransferases M. SssI and M. HhaI. These two
methyltransferases recognize (C) under bar pG and G (C) under bar GC
sequences, respectively, and transfer a methyl group to the C5 atom of
cytosine (C). A series of 18-mer unmethylated or hemimethylated
oligodeoxynucleotide duplexes containing trans-anti-B[a]P-N-2-dG
adducts was generated. The B+ or B- residues were introduced either 5 '
or 3 ' adjacent or opposite to the target 2 '-deoxycytidines. The B[a]
PDE lesions practically produced no effect on M. SssI binding to DNA
but reduced M. HhaI binding by 1-2 orders of magnitude. In most cases,
the benzo[ a] pyrenyl residues decreased the methylation efficiency of
hemimethylated and unmethylated DNA by M. SssI and M. HhaI. An absence
of the methylation of hemimethylated duplexes was observed when either
the (+)- or the (-)-trans-anti-B[a]P-N-2-dG adduct was positioned 5 '
to the target dC. The effects observed may be related to the minor
groove conformation of the bulky benzo[a]pyrenyl residue and to a
perturbation of the normal contacts of the methyltransferase catalytic
loop with the B[a]PDE-modified DNA. Our results indicate that a
trans-anti-B[a] P-N-2-dG lesion flanking a target dC in the CpG
dinucleotide sequence on its 5 '- side has a greater adverse impact on
methylation than the same lesion when it is 3 ' adjacent or opposite to
the target dC.

<>

<1>Subach, O.M., Khoroshaev, A.V., Gerasimov, D.N., Baskunov, V.B., Shchyolkina, A.K., Gromova, E.S.
<2>2-Pyrimidinone as a probe for studying the EcoRII DNA methyltransferase-substrate interaction.
<3>Eur. J. Biochem.
<4>271
<5>2391-2399
<6>2004
<7>EcoRII DNA methyltransferase (M.EcoRII) recognizes the 5'...CC*T/AGG...3' DNA sequence and
catalyzes the transfer of the
methyl group from S-adenosyl-L-methionine to the C5 position of the
inner cytosine residue (C*). Here, we study the mechanism of inhibition
of M.EcoRII by DNA containing 2-pyrimidinone, a cytosine analogue
lacking an NH2 group at the C4 position of the pyrimidine ring. Also,
DNA containing 2-pyrimidinone was used for probing contacts of M.EcoRII
with functional groups of pyrimidine bases of the recognition sequence.
2-Pyrimidinone was incorporated into the 5'...CCT/AGG...3' sequence
replacing the target and nontarget cytosine and central thymine
residues. Study of the DNA stability using thermal denaturation of
2-pyrimidinone containing duplexes pointed to the influence of the
bases adjacent to 2-pyrimidinone and to a greater destabilizing
influence of 2-pyrimidinone substitution for thymine than that for
cytosine. Binding of M.EcoRII to 2-pyrimidinone containing DNA and
methylation of these DNA demonstrate that the amino group of the outer
cytosine in the EcoRII recognition sequence is not involved in the
DNA-M.EcoRII interaction. It is probable that there are contacts
between the functional groups of the central thymine exposed in the
major groove and M.EcoRII. 2-Pyrimidinone replacing the target cytosine
in the EcoRII recognition sequence forms covalent adducts with
M.EcoRII. In the absence of the cofactor S-adenosyl-L-methionine,
proton transfer to the C5 position of 2-pyrimidinone occurs and in the
presence of S-adenosyl-L-methionine, methyl transfer to the C5 position
of 2-pyrimidinone occurs.

<>

<1>Subach, O.M., Maltseva, D.V., Shastry, A., Kolbanovskiy, A., Klimasauskas, S., Geacintov, N.E., Gromova, E.S.
<2>The stereochemistry of benzo(a)pyrene-2'-deoxyguanosine adducts affects DNA methylation by SssI and HhaI DNA methyltransferases.
<3>FEBS J.
<4>274
<5>2121-2134
<6>2007
<7>The biologically most significant genotoxic metabolite of the environmental pollutant
benzo[a]pyrene (B[a]P), (+)-7R,8S-diol 9S,10R-epoxide, reacts chemically with guanine in DNA,
resulting in the predominent formation of (+)-trans-B[a]P-N2-dG and, to a lesser extent,
(+)-cis-B[a]P-N2-dG adducts.  Here, we compare the effects of the adduct stereochemistry and
conformation on the methylation of cytosine catalyzed by two purified prokaryotic DNA
methyltransferases, SssI and HhaI, with the lesions positioned within or adjacent to their CG
and GCGC recognition sites, respectively.  The fluorescence properties of the pyrenyl residues
of the (+)-cis-B[a]aP-N2-dG and (+)-trans-B[a]P-N2-dG adducts in complexes with MTases are
enhanced, but to different extents, indicating that aromatic B[a]P residues are positioned in
different microenvironments in the DNA-protein complexes.  We have previously shown that the
(+)-trans-isomeric adduct inhibits both the binding and methylating efficiencies (kcat) of
both MTases.  Here we show that the stereoisomeric (+)-cis-B[a]P-N2-dG lesion has only a
minimal effect on the binding of these MTases and on kcat.  The minor-groove (+)-trans adduct
interferes with the formation of the normal DNA minor-groove contacts with the catalytic loop
of the MTases.  However, the intercalated base-displaced (+)-cis adduct does not interfere
with the minor-groove DNA-catalytic loop contacts, allowing near-normal binding of the MTases
and undiminished kcat values.

<>

<1>Subramaniam, R., Wang, Y., Mathews, C.K., Santi, D.V.
<2>On the inhibition of deoxycytidylate hydroxymethylase by 5-fluoro-2'-deoxycytidine 5'-monophosphate .
<3>Arch. Biochem. Biophys.
<4>275
<5>11-15
<6>1989
<7>Studies were performed to determine whether 5-fluoro-2'-deoxycytidine 5'-monophosphate
(FdCMP) is an inhibitor of deoxycytidylate hydroxymethylase and whether it could form an
isolable covalent complex with the enzyme and the cofactor, 5,10-methylenetetrahydrofolate.
The results showed that although FdCMP is a competitive inhibitor of dCMP hydroxymethylase, it
does not cause time-dependent inhibition of the enzyme in the presence of cofactor. Further,
although uv difference spectral evidence was found for a FdCMP-cofactor-enzyme complex, the
complex was not sufficiently stable to isolate on nitrocellulose filters. We conclude that
FdCMP is not a mechanism-based inhibitor of dCMP hydroxymethylase.

<>

<1>Suchkov, S.V., Lopatina, N.G., Arutyunyan, E.E., Nikolskaya, I.I., Debov, S.S.
<2>Study of the conditions of activation and stabilization of Shigella sonnei 47 DNA methylases in the process of fractionation, purification, and during storage.
<3>Biokhimiia
<4>51
<5>1369-1376
<6>1986
<7>A comparative investigation was made of the factors of activation and
conditions of stabilization of partially purified and individual fractions of
DNA methylases of the strain Shigella sonnei 47.  The influence of the
temperature system, glycerin, albumin, protease inhibitors, divalent ions, as
well as the conditions of storage at the value of the isoelectric point of the
protein on the activity of the enzymes of methylation was studied.  It was
shown that the effects of activating factors and levels of stabilization differ
appreciably, depending on the degree of purification and the composition of the
enzyme preparations  It was established that glycerin is ineffective as a
stabilizing agent.  It was shown that of all the divalent cations studied the
only activator universal for the methylases of Shigella sonnei 47 is Ca2+ ions.
A pronounced stabilizing effect of albumin on the activity of these DNA
methylases was demonstrated.  With the exception of one enzymatic fraction, the
protease inhibitors have virtually no effect on the level of methylating
activity of the preparations.  The phenomenon of spontaneous fluctuations of
the methylating activity of Shigella sonnei 47 enzyme preparations during
storage is described.

<>

<1>Suchkov, S.V., Nikolskaya, I.I., Debov, S.S.
<2>Isoelectrofocusing of DNA-methylases from Shigella sonnei 47.
<3>Vopr. Med. Khim.
<4>29
<5>117-122
<6>1983
<7>Fractionation of bacterial methylases was studied by means of the isoelectrofocusing
technique.  The method exhibited high accuracy, efficiency and reproducibility in studies of
methylases from Shigella sonnei 47.  High column capacity and distinct focusing effect
increased the experimental advantages of work with various amounts of protein as compared with
ion exchange chromatography.  By means of the isoelectrofocusing technique three methylases
were detected in the strain S. sonnei 47, which were dissimilar in their isoelectric points at
pH 6.5, 8.4 and 9.8.  These findings indicate the heterogeneous pattern of enzyme methylation
in the strain studied.  Complete removing of endogenous DNA was the main precondition for
correct fractionation of methylases by means of the isoelectrofocusing technique.  A procedure
is described for purification of S. sonnei 47 methylases from traces of endogenous DNA by
affinity chromatography on heparin-Sepharose.

<>

<1>Suchkov, S.V., Nikolskaya, I.I., Debov, S.S.
<2>Fractionation and purification of DNA-methylases of Shigella sonnei 47 cells.
<3>Biokhimiia
<4>50
<5>1797-1804
<6>1985
<7>Possible applications of various column chromatography techniques and isoelectrofocusing for
the study of DNA-methylases of Shigella sonnei 47 cells were analyzed.  A simple, rapid and
convenient procedure based on the use of cation-exchange chromatography was developed for
obtaining a highly active total preparation of methylases.  Affinity chromatography on
heparin-Sepharose was shown to be a promising approach for separating methylases according to
their specificity towards nitrogenous bases.  Isoelectrofocusing was used to identify in
Shigella sonnei 47 cells six individual methylating enzymes differing in their pI values.
Under the stipulation that Shigella sonnei 47 DNA-methylases show a tendency to aggregate in
the course of fractionation, column chromatography is of little or no use in isolating and
purifying individual methylating enzymes of the given strain.  The advantages of the
isoelectrofocusing technique and its utility in the study of different molecular forms of
site-specific enzymes are discussed.

<>

<1>Suck, D.
<2>Flip out and modify.
<3>Curr. Biol.
<4>4
<5>252-255
<6>1994
<7>The crystal structure of a complex between a methyltransferase and DNA shows that, remarkably,
the target cytosine base is swung out of the double helix and located next to the enzyme's
S-adenosyl-L-homocysteine cofactor.

<>

<1>Sudina, A., Volkov, E., Oretskaya, T., Naryshkin, N., Ivanovskaya, M., Kubareva, E.
<2>Detection of glycosylase, endonuclease and methyltransferase enzyme activities using immobilized oligonucleotides.
<3>Life
<4>56
<5>139-143
<6>2004
<7>A novel rapid assay for detection of DNA glycosylase, restriction endonuclease, and DNA
methyltransferase enzyme activities is presented.
The assay is based on enzyme-dependent label release (in the case of
glycosylase and endonuclease), or non-release (in the case of
methyltransferase) into solution from end-labeled DNA immobilized on a
solid support (CPG or Tenta Gel S-NH2). The assay has been validated
for monitoring activity of repair enzyme uracil-DNA glycosylase,
restriction endonucleases SsoII, MvaI and EcoRII and (cytosine-5)-DNA
methyltransferase SsoII. Two types of labels have been tested and found
compatible with the assay: radioactive (32P) and fluorescent
(rhodamine B and fluorescein). The enzyme activity is estimated as a
ratio of the label released into solution to the total amount of the
label. Use of fluorescent labeling facilitates detection while use of
solid phase-immobilized substrates facilitates product separation,
improved assay sensitivity, and increases throughput of assay. Proposed
technique provides an estimate of enzyme activity but not its specific
activity. Thus, the assay will most valuable in the applications where
rapid estimation of enzyme activity is necessary.

<>

<1>Sudina, A.E., Zatsepin, T.S., Pingoud, V., Pingoud, A., Oretskaya, T.S., Kubareva, E.A.
<2>Affinity modification of the restriction endonuclease SsoII by 2'-aldehyde-containing double stranded DNAs.
<3>Biochemistry
<4>70
<5>1137-1144
<6>2005
<7>Properties of 2'-aldehyde-containing double stranded DNAs (dsDNAs) have been studied for the
first time as substrate analogs of the restriction endonuclease SsoII. These reactive
oligonucleotides were successfully cross-linked to the restriction endonuclease SsoII by
reductive amination, and conditions for DNA-protein conjugate trypsinolysis followed by the
oligonucleotide-peptide conjugate purification were optimized. Use of MALDI-TOF mass
spectrometry revealed that covalent linkage forms between the sugar moiety of the central
pyrimidine nucleoside of the SsoII recognition site and Lys173 of the enzyme. The latter is
probably involved in initial steps of enzyme-substrate recognition during dsDNA readout.

<>

<1>Suen, G. et al.
<2>Complete Genome of the Cellulolytic Ruminal Bacterium Ruminococcus albus 7.
<3>J. Bacteriol.
<4>193
<5>5574-5575
<6>2011
<7>Ruminococcus albus 7 is a highly cellulolytic ruminal bacterium that is a member of the phylum
Firmicutes. Here, we describe the complete genome of
this microbe. This genome will be useful for rumen microbiology and
cellulosome biology and in biofuel production, as one of its major
fermentation products is ethanol.

<>

<1>Suerbaum, S. et al.
<2>The complete genome sequence of the carcinogenic bacterium Helicobacter hepaticus.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>7901-7906
<6>2003
<7>Helicobacter hepaticus causes chronic hepatitis and liver cancer in mice. It is the prototype
enterohepatic Helicobacter species and a close
relative of Helicobacter pylori, also a recognized carcinogen. Here we
report the complete genome sequence of H. hepaticus ATCC51449. H.
hepaticus has a circular chromosome of 1,799,146 base pairs, predicted to
encode 1,875 proteins. A total of 938, 953, and 821 proteins have
orthologs in H. pylori, Campylobacter jejuni, and both pathogens,
respectively. H. hepaticus lacks orthologs of most known H. pylori
virulence factors, including adhesins, the VacA cytotoxin, and almost all
cag pathogenicity island proteins, but has orthologs of the C. jejuni
adhesin PEB1 and the cytolethal distending toxin (CDT). The genome
contains a 71-kb genomic island (HHGI1) and several genomic islets whose
G+C content differs from the rest of the genome. HHGI1 encodes three basic
components of a type IV secretion system and other virulence protein
homologs, suggesting a role of HHGI1 in pathogenicity. The genomic
variability of H. hepaticus was assessed by comparing the genomes of 12 H.
hepaticus strains with the sequenced genome by microarray hybridization.
Although five strains, including all those known to have caused liver
disease, were indistinguishable from ATCC51449, other strains lacked
between 85 and 229 genes, including large parts of HHGI1, demonstrating
extensive variation of genome content within the species.

<>

<1>Suetake, I., Hayata, D., Tajima, S.
<2>The amino-terminus of mouse DNA methyltransferase 1 forms an independent domain and binds to DNA with the sequence involving PCNA binding motif.
<3>J. Biochem. (Tokyo)
<4>140
<5>763-776
<6>2006
<7>DNA methylation patterns in genome are maintained during replication by a DNA
methyltransferase Dnmt1. Mouse Dnmt1 is a 180 kDa protein
comprising the N-terminal regulatory domain, which covers 2/3 of the
molecule, and the rest C-terminal catalytic domain. In the present
study, we demonstrated that the limited digestion of full-length Dnmt1
with different proteases produced a common N-terminal fragment, which
migrated along with Dnmt1 (1-248) in SDS-polyacrylamide gel
electrophoresis. Digestion of the N-terminal domains larger than Dnmt1
(1-248) with chymotrypsin again produced the fragment identical to the
size of Dnmt1 (1-248). These results indicate that the N-terminal
domain of 1-248 forms an independent domain. This N-terminal domain
showed DNA binding activity, and the responsible sequence was narrowed
to the 79 amino acid residues involving the proliferating cell nuclear
antigen (PCNA) binding motif. The DNA binding activity did not
distinguish between DNA methylated and non-methylated states, but
preferred to bind to the minor groove of AT-rich sequence. The DNA
binding activity of the N-terminal domain competed with the PCNA
binding. We propose that DNA binding activity of the N-terminal domain
contributes to the localization of Dnmt1 to AT-rich sequence such as
Line 1, satellite, and the promoter of tissue-specific silent genes.

<>

<1>Suetake, I., Mishima, Y., Kimura, H., Lee, Y.-H., Goto, Y., Takeshima, H., Ikegami, T., Tajima, S.
<2>Characterization of DNA-binding activity in the N-terminal domain of the DNA methyltransferase Dnmt3a.
<3>Biochem. J.
<4>437
<5>141-148
<6>2011
<7>
<>

<1>Suetake, I., Miyazaki, J., Murakami, C., Takeshima, H., Tajima, S.
<2>Distinct enzymatic properties of recombinant mouse DNA methyltransferases Dnmt3a and Dnmt3b.
<3>J. Biochem. (Tokyo)
<4>133
<5>737-744
<6>2003
<7>Recombinant mouse Dnmt3a and Dnmt3b were expressed in sf9 cells and purified to near
homogeneity. The purified Dnmt3a and Dnmt3b gave specific
activities of 1.8 +/- 0.3 and 1.3 +/- 0.1 mol/h/mol enzyme towards
poly(dGdC)-poly(dGdC), respectively, which were the highest among those
reported. Dnmt3a or Dnmt3b showed similar K(m) values towards
poly(dIdC)-poly(dIdC) and poly(dGdC)-poly(dGdC). The K(m) values for
S-adenosyl-L-methionine were not affected by the methyl-group acceptors,
poly(dI-dC)-poly(dIdC) and poly(dG-dC)-poly(dGdC). The results indicate
that the enzymes are de novo-type DNA methyltransferases. Dnmt3a and
Dnmt3b activities were inhibited by Mn(2+) and Ni(2+) and showed broad pH
optima around neutral pH. Both enzymes were susceptible to sodium ions,
which inhibited their activity at around physiological ionic strength.
However, Dnmt3a was fully active at physiological potassium concentration,
but Dnmt3b was not. Using designed oligonucleotides for the analysis of
cytosine methylation, we demonstrated that, in addition to CpG, Dnmt3a
methylated CpA but not CpT and CpC, and that Dnmt3b methylated CpA and CpT
but scarcely CpC. The relative activity of Dnmt3b towards nonCpG sequences
was higher than that of Dnmt3a. These differences in enzymatic properties
of Dnmt3a and Dnmt3b may contribute to the distinct functions of these
enzymes in vivo.

<>

<1>Suetake, I., Shi, L.H., Watanabe, D., Nakamura, M., Tajima, S.
<2>Proliferation stage-dependent expression of DNA methyltransferase (Dnmt1) in mouse small intestine.
<3>Cell Struct. Funct.
<4>26
<5>79-86
<6>2001
<7>In cultured cells, the maintenance-type DNA methyltransferase (Dnmt1) is highly expressed
during the proliferation stage. In the present
study, we detected significant expression of Dnmt1 protein in the
nuclear fraction of mouse small intestine. From its mobility in SDS
polyacrylamide gel electrophoresis and the specific antibodies against
the somatic cell-type Dnmt1, Dnmt1 was determined as a somatic cell
type. Immunofluorescence study revealed that the Dnmt1 was highly
expressed in the proliferating stem cells in crypts, and was localized
in the nuclei. The present results indicate that the expression of
Dnmt1 in vivo is also under the control of cell proliferation as in
cultured cells.

<>

<1>Suetake, I., Shinozaki, F., Miyagawa, J., Takeshima, H., Tajima, S.
<2>DNMT3L stimulates the DNA methylation activity of Dnmt3a and Dnmt3b through a direct interaction.
<3>J. Biol. Chem.
<4>279
<5>27816-27823
<6>2004
<7>In mammals, the resetting of DNA methylation patterns in early embryos and germ cells is
crucial for development. Two DNA methyltransferases, Dnmt3a and Dnmt3b, are responsible for
the creation of DNA methylation patterns. Dnmt3L, a member of the Dnmt3 family, has been
reported to be necessary for maternal methylation imprinting, possibly by interacting with
Dnmt3a and/or Dnmt3b (Hata et al., Development 129, 1983-1993, 2002). In the present study,
the effect of DNMT3L, a human homologue of Dnmt3L, on the DNA methylation activity of mouse
Dnmt3a and Dnmt3b was examined in vitro. DNMT3L enhanced the DNA methylation activity of
Dnmt3a and Dnmt3b about 1.5-3 fold in a dose-dependent manner, but not that of Dnmt1. Although
the extents of stimulation were different, a stimulatory effect on the DNA methylation
activity was observed for all the substrate DNA sequences examined, such as those of the
maternally methylated SNRPN and Lit-1 imprinting genes, the paternally methylated H19
imprinting gene, the CpG island of the myoD gene, the 5S ribosomal RNA gene, an artificial 28
bp DNA, poly(dGdC)-poly(dGdC), and poly(dIdC)-poly(dIdC). DNMT3L could not bind to DNA but
could to Dnmt3a and Dnmt3b, indicating that the stimulatory effect of DNMT3L on the DNA
methylation activity may not be due to the guiding of Dnmt3a and Dnmt3b to the targeting DNA
sequence, but may comprise a direct effect on their catalytic activity. The carboxyl-terminal
half of DNMT3L was found to be responsible for the enhancement of the enzyme activity.

<>

<1>Sufrin, J.R., Christman, J.K., Marasco, C.J. Jr., Sheikhnejad, G.
<2>Substrate for detection of mammalian 5-C-DNA methyltransferase.
<3>US Patent Office
<4>US 5652105
<5>
<6>1997
<7>A substrate selective for detection of mammalian 5-C-DNA methyltransferase in the presence of
bacterial 5-C-DNA methyltransferase, said substrate comprising oligomeric DNA which contains
at least one 5-methylcytosine residue, and at least one cytosine or 5-fluorocytosine residue,
each of which are followed in linkage to a guanine residue.  The invention also includes a
method for measuring the presence of mammalian 5-C-DNA methyltransferase which comprises
contacting a sample containing 5-C-DNA methyltransferase which comprises contacting a sample
containing 5-C-DNA methyltransferase with the substrate.

<>

<1>Sugden, B., DeTroy, B., Roberts, R.J., Sambrook, J.
<2>Agarose slab-gel electrophoresis equipment.
<3>Anal. Biochem.
<4>68
<5>36-46
<6>1975
<7>Simple slab-gel molds which utilize the electrophoresis apparatus described by
F.W. Studier (J. Mol. Biol. 79, 237 (1973)) have been designed for pouring and
running agarose slab-gels.  Analytical gels in which many samples are run
simultaneously facilitate the assay of many enzymes which lead to physical
changes in DNA, whereas the preparative gels allow the separation of large
quantities (1-20 mg) of DNA fragments.

<>

<1>Sugisaki, H.
<2>Recognition sequence of a restriction endonuclease from Haemophilus gallinarum.
<3>Gene
<4>3
<5>17-28
<6>1978
<7>From a comparison of the sequences in and around the cleavage sites of
restriction endonuclease HgaI isolated from Haemophilus gallinarum, the
recognition sequence and cleavage site of this enzyme was deduced as below:
(5')-----N^pN-N-N-N-N-N-N-N-N-N-G-C-G-T-C-N-----(3')
(3')-----N-N-N-N-N-Np^N-N-N-N-N-C-G-C-A-G-N-----(5') This enzyme recognizes a
specific but asymmetric penta-nucleotide sequence, GCGTC, and introduces
staggered cleavages at appointed positions away from the recognition sequence,
generating protruding 5'-ends of five nucleotides.  The sequences surrounding
the cleavage sites bear no obvious relation to one another.

<>

<1>Sugisaki, H.
<2>Nucleotide sequence of the gene of HgaI restriction endonuclease.
<3>Bull. Inst. Chem. Res. Kyoto Univ.
<4>71
<5>338-342
<6>1993
<7>
<>

<1>Sugisaki, H., Kanazawa, S.
<2>New restriction endonucleases from Flavobacterium okeanokoites (FokI) and Micrococcus luteus (MluI).
<3>Gene
<4>16
<5>73-78
<6>1981
<7>Two new restriction endonucleases have been isolated from Flavobacterium
okeanokoites IFO12536 and Micrococcus luteus IFO12992 and named FokI and MluI,
respectively.  Based on analysis of the sequences around the restriction sites,
the recognition sequences and cleavage sites of these endonucleases were
deduced as below: FokI:	(5')--N-G-G-A-T-G-N-N-N-N-N-N-N-N-N^pN-N-N-N--N--(3')
	(3')--N-C-C-T-A-C-N-N-N-N-N-N-N-N-N--N-N-N-Np^N--(5') and
MluI:	(5')--N-A^pC-G-C-G--T-N--(3') 	(3')--N-T  G-C-G-Cp^A-N--(5') 	MluI
introduces double-strand cleavages at unique sequences that are completely
two-fold rotationally symmetric like most type II restriction endonucleases.
FokI belongs to a class of restriction endonucleases that recognize specific
but asymmetric nucleotide sequences and introduce staggered cleavages at
appointed positions away from the recognition sequences.

<>

<1>Sugisaki, H., Kita, K., Takanami, M.
<2>The FokI restriction-modification system.  II. Presence of two domains in FokI methylase responsible for modification of different DNA strands.
<3>J. Biol. Chem.
<4>264
<5>5757-5761
<6>1989
<7>Based on the previous findings that the FokI methylase (MFokI) consists of 647
amino acid residues and contains two copies of the segment specific for adenine
methylase, Asp-Pro-Pro-Tyr, at amino acid positions 218-221 and 548-551, the
role of these copies in the methylation reaction was investigated by
introduction of a mutation into each segment.  The MFokI gene was inserted into
M13 vectors, and the Asp residues in the two segments were converted to Gly and
Ala by oligonucleotide-directed mutagenesis.  The wild-type and mutant genes
were recloned into an expression vector, from which gene products were
purified.  A short DNA fragment carrying the FokI recognition site was treated
with each of these enzymes, and after separation of the two strands by duplex
formation with M13 viral DNAs carrying the respective strands, the presence or
absence of modification was judged from susceptibility to FokI endonuclease.
The results of analysis showed that different strands were modified in an
asymmetric way by the introduction of mutations into one of the two segments,
and that the segments at the N-terminal and C-terminal moieties participated in
modification of the strands carrying 5'-GGATG-3' and 3'-CCTAC-5', respectively.
We concluded that MFokI contained two functional domains each of which was
responsible for modification of different strands in the target DNA.

<>

<1>Sugisaki, H., Maekawa, Y., Kanazawa, S., Takanami, M.
<2>Screening of type II restriction endonucleases.  I.  Isolation and characterization of enzymes from fifteen bacterial strains.
<3>Bull. Inst. Chem. Res. Kyoto Univ.
<4>60
<5>328-335
<6>1982
<7>One hundred and forty-seven bacterial strains were surveyed for the presence of
type II restriction endonucleases, and eighteen species of enzymes were
successfully isolated from fifteen strains in which enzyme activities were
identified.  Based on analysis of the restriction patterns generated from viral
and plasmid DNAs and of the sequences around the cleavage-sites, four of
enzymes named AatII, BanII, FokII and MluI were found to have new
specificities.  The remaining fourteen enzymes, named AatI, AtuIAMI, BanIII,
BprI, EcoICRI, GglI, GinI, MauI, PaiI, PanI, PflI, PpuI, and SpaI, were
isoschizomers of known enzymes.

<>

<1>Sugisaki, H., Maekawa, Y., Kanazawa, S., Takanami, M.
<2>New restriction endonucleases from Acetobacter aceti and Bacillus aneurinolyticus.
<3>Nucleic Acids Res.
<4>10
<5>5747-5752
<6>1982
<7>*
Two restriction endonucleases with new sequence specificities have been
isolated from Acetobacter aceti IFO 3281 and Bacillus aneurinolyticus IAM 1077
and named AatII and BanII, respectively.  Based on analysis of the sequences
around the restriction sites, the recognition sequences and cleavage sites of
these endonucleases were deduced as below:

 AatII:  (5') ---N-G- A-C-G-T^pC-N---  (3')
         (3') ---N-Cp^T-G-C-A- G-N---  (5')

 BanII:  (5') --N-G -Pu-G-C-Py^pC-N--  (3')
         (3') --N-Cp^Py-C-G-Pu- G-N--  (5')


<>

<1>Sugisaki, H., Takanami, M.
<2>DNA Sequence restricted by restriction endonuclease AP from Haemophilus aphirophilus.
<3>Nature New Biol.
<4>246
<5>138-140
<6>1973
<7>Many bacterial strains contain strain-specific restriction endonucleases which
degrade foreign DNA at a limited number of sites.  The site-specific action of
these enzymes is thought to be a consequence of their ability to recognise
specific nucleotide sequences.  Such sequences restricted by endonuclease R
(endoR) from Haemophilus influenzae Rd and RI endonuclease (endo RI) from
Escherichia coli carrying R factor have been determined.  In our laboratory,
several species of similar enzymes have been isolated from different
Haemophilus strains.  Different enzymes seem to have different cleavage site
specificities, since each enzyme produces different sizes of fragments from
bacteriophage DNA.  We analysed the terminal nucleotide sequences of fragments
produced from fd RF-I (doubly closed replicative form) DNA and T3 DNA by
cleavage with one of the Haemophilus enzymes, endonuclease AP (endo AP)
isolated from H. aphirophilus, and found that this enzyme cleaved DNA at a
sequence of four nucleotide pairs with a two-fold rotational symmetry.

<>

<1>Sugisaki, H., Yamamoto, K., Takanami, M.
<2>The HgaI restriction-modification system contains two cytosine methylase genes responsible for modification of different DNA strands.
<3>J. Biol. Chem.
<4>266
<5>13952-13957
<6>1991
<7>A DNA fragment of about 3.4 kilobase pairs that expressed the HgaI modification activity was
cloned from the chromosomal DNA of Haemophilus gallinarum, and its nucleotide sequence was
determined. Two open reading frames (ORF) which could code for structurally similar proteins
were identified in the upstream and middle regions and a truncated ORF in the downstream
region in the same orientation. When the respective ORF's were separately cloned, the clones
carrying the upstream and middle ORF's both expressed the modification activity, indicating
that the two genes are involved in modification of the HgaI restriction modification system.
In order to determine the sites of modification precisely, the respective genes were recloned
into an expression vector, from which gene products were purified. A short DNA fragment
carrying the HgaI recognition site was treated with each of these enzymes, and after
separation of the two strands by duplex formation with M13 viral DNAs carrying the respective
strands, the presence or absence of modification was judged from susceptibility to HgaI
endonuclease. The results of analysis showed that different strands were modified in an
asymmetric way by each gene product. Analysis of the species and positions of modified bases
by the Maxam-Gilbert method further demonstrated that the gene products from the upstream and
middle ORF's participated in methylation of the inter cytosine residues of the strands
carrying 3'-CTGCG-5' and 5'-GACGC-3', respectively. We concluded that the HgaI
modification system consisted of two cytosine methylase genes responsible for modification of
different strands in the target DNA.

<>

<1>Suhaili, Z., Lean, S.S., Yahya, A., Mohd, D.M.N., Ali, A.M., Yeo, C.C.
<2>Draft Genome Sequence of Methicillin-Resistant Staphylococcus aureus KT/Y21, a Sequence Type 772 (ST772) Strain Isolated from a Pediatric Blood Sample in Terengganu, Malaysia.
<3>Genome Announcements
<4>2
<5>e00271-14
<6>2014
<7>Here, we report the draft genome sequence of a methicillin-resistant Staphylococcus aureus
(MRSA) strain, KT/Y21, isolated from a blood sample of a pediatric patient. This strain
belongs to sequence type 772 (ST772), harbors the staphylococcal cassette chromosome mec
element (SCCmec) type V, and is positive for the Panton-Valentine leukocidin (PVL) pathogenic
determinant.

<>

<1>Sui, Z., Zhou, W., Yao, K., Liu, L., Zhang, G., Yang, Y., Feng, J.
<2>Complete Genome Sequence of Streptococcus pneumoniae Strain A026, a Clinical Multidrug-Resistant Isolate Carrying Tn2010.
<3>Genome Announcements
<4>1
<5>e01034-13
<6>2013
<7>Streptococcus pneumoniae is a primary cause of bacterial infection in humans. Here, we present
the complete genome sequence of S. pneumoniae strain A026, which
is a multidrug-resistant strain isolated from cerebrospinal fluid.

<>

<1>Sukackaite, R., Grazulis, S., Bochtler, M., Siksnys, V.
<2>The recognition domain of the BpuJI restriction endonuclease in complex with cognate DNA at 1.3-A resolution.
<3>J. Mol. Biol.
<4>378
<5>1084-1093
<6>2008
<7>Type IIS restriction endonucleases recognize asymmetric DNA sequences and cleave both DNA
strands at fixed positions downstream of the recognition
site. The restriction endonuclease BpuJI recognizes the asymmetric
sequence 5'-CCCGT; however, it cuts at multiple sites in the vicinity of
the target sequence. BpuJI consists of two physically separate domains,
with catalytic and dimerization functions in the C-terminal domain and DNA
recognition functions in the N-terminal domain. Here we report the crystal
structure of the BpuJI recognition domain bound to cognate DNA at 1.3-A
resolution. This region folds into two winged-helix subdomains, D1 and D2,
interspaced by the DL subdomain. The D1 and D2 subdomains of BpuJI share
structural similarity with the similar subdomains of the FokI DNA-binding
domain; however, their orientations in protein-DNA complexes are
different. Recognition of the 5'-CCCGT target sequence is achieved by
BpuJI through the major groove contacts of amino acid residues located on
both the helix-turn-helix motifs and the N-terminal arm. The role of these
interactions in DNA recognition is also corroborated by mutational
analysis.

<>

<1>Sukackaite, R., Grazulis, S., Tamulaitis, G., Siksnys, V.
<2>The recognition domain of the methyl-specific endonuclease McrBC flips out 5-methylcytosine.
<3>Nucleic Acids Res.
<4>40
<5>7552-7562
<6>2012
<7>DNA cytosine methylation is a widespread epigenetic mark.  Biological effects of DNA
methylation are mediated by the proteins that preferentially bind to 5-methylcytosine (5mC) in
different sequence contexts.  Until now two different structural mechanisms have been
established for 5mC recognition in eukaryotes; however, it is still unknown how discrimination
of the 5mC modification is achieved in prokaryotes.  Here we report the crystal structure of
the N-terminal DNA-binding domain (McrB-N) of the methyl-specific endonuclease McrBC from
Escherichia coli.  The McrB-N protein shows a novel DNA-binding fold adapted for
5-mC-recognition.  In the McrB-N structure in complex with methylated DNA, the 5mC base is
flipped out from the DNA duplex and positioned within a binding pocket.  Base flipping
elegantly explains why McrBC system restricts only T4-even phages impaired in glycosylation
[Luria, S.E. and Human, M.L. (1952) A nonhereditary, host-induced variation of bacterial
viruses.  J. Bacteriol., 64, 557-569]: flipped out 5-hydroxymethylcytosine is accommodated in
the binding pocket but there is no room for the glycosylated base.  The mechanism for 5mC
recognition employed by McrB-N is highly reminiscent of that for eukaryotic SRA domains,
despite the differences in their protein folds.

<>

<1>Sukackaite, R., Lagunavicius, A., Stankevicius, K., Urbanke, C., Venclovas, C., Siksnys, V.
<2>Restriction endonuclease BpuJI specific for the 5'-CCCGT sequence is related to the archaeal Holliday junction resolvase family.
<3>Nucleic Acids Res.
<4>35
<5>2377-2389
<6>2007
<7>Type IIS restriction endonucleases (REases) recognize asymmetric DNA sequences and cleave both
DNA strands at fixed positions downstream of the
recognition site. REase BpuJI recognizes the asymmetric sequence 5'-CCCGT,
however it cuts at multiple sites in the vicinity of the target sequence.
We show that BpuJI is a dimer, which has two DNA binding surfaces and
displays optimal catalytic activity when bound to two recognition sites.
BpuJI is cleaved by chymotrypsin into an N-terminal domain (NTD), which
lacks catalytic activity but binds specifically to the recognition
sequence as a monomer, and a C-terminal domain (CTD), which forms a dimer
with non-specific nuclease activity. Fold recognition approach reveals
that the CTD of BpuJI is structurally related to archaeal Holliday
junction resolvases (AHJR). We demonstrate that the isolated catalytic CTD
of BpuJI possesses end-directed nuclease activity and preferentially cuts
3 nt from the 3'-terminus of blunt-ended DNA. The nuclease activity of the
CTD is repressed in the apo-enzyme and becomes activated upon specific DNA
binding by the NTDs. This leads to a complicated pattern of specific DNA
cleavage in the vicinity of the target site. Bioinformatics analysis
identifies the AHJR-like domain in the putative Type III enzymes and
functionally uncharacterized proteins.

<>

<1>Sullivan, J.T., Trzebiatowski, J.R., Cruickshank, R.W., Gouzy, J., Brown, S.D., Elliot, R.M., Fleetwood, D.J., McCallum, N.G., Rossbach, U., Stuart, G.S., Weaver, J.E., Webby, R.J., de Bruijn, F.J., Ronson, C.W.
<2>Comparative sequence analysis of the symbiosis island of Mesorhizobium loti strain R7A.
<3>J. Bacteriol.
<4>184
<5>3086-3095
<6>2002
<7>The Mesorhizobium loti strain R7A symbiosis island is a 502-kb
chromosomally integrated element which transfers to nonsymbiotic
mesorhizobia in the environment, converting them to Lotus symbionts. It
integrates into a phenylalanine tRNA gene in a process mediated by a
P4-type integrase encoded at the left end of the element. We have
determined the nucleotide sequence of the island and compared its deduced
genetic complement with that reported for the 611-kb putative symbiosis
island of M. loti strain MAFF303099. The two islands share 248 kb of DNA,
with multiple deletions and insertions of up to 168 kb interrupting highly
conserved colinear DNA regions in the two strains. The shared DNA regions
contain all the genes likely to be required for Nod factor synthesis,
nitrogen fixation, and island transfer. Transfer genes include a trb
operon and a cluster of potential tra genes which are also present on the
strain MAFF303099 plasmid pMLb. The island lacks plasmid replication
genes, suggesting that it is a site-specific conjugative transposon. The
R7A island encodes a type IV secretion system with strong similarity to
the vir pilus from Agrobacterium tumefaciens that is deleted from
MAFF303099, which in turn encodes a type III secretion system not found on
the R7A island. The 414 genes on the R7A island also include putative
regulatory genes, transport genes, and an array of metabolic genes. Most
of the unique hypothetical genes on the R7A island are strain-specific and
clustered, suggesting that they may represent other acquired genetic
elements rather than symbiotically relevant DNA.

<>

<1>Sullivan, K.M., Macdonald, H.J., Saunders, J.R.
<2>Characterization of DNA restriction and modification activities in Neisseria species.
<3>FEMS Microbiol. Lett.
<4>44
<5>389-393
<6>1987
<7>Type II restriction endonuclease activities detected in various Neisseria
species were characterized for sequence specificity and precise site of
cleavage.  NsiCI isolated from N. sicca C351 cleaves the sequence 5'-GAT^ATC-3'
(EcoRV isoschizomer); NmeCI from N. meningitidis C114 and NphI from N.
pharyngis C245 cleave 5'-N^GATCN-3' (MboI isoschizomers); NgoPII and NgoPIII
from N. gonorrhoeae P9-2 cleave at 5'-GG^CC-3' (HaeIII isoschizomer) and
5'-CC^GCGG-3' (SacII isoschizomer), respectively.  Chromosomal DNA isolated
from these strains and two other N. meningitidis strains (which lacked
detectable endonuclease activities), was found to be refractive to cleavage by
various restriction enzymes, implying the presence of methylase activities
additional to those required for protection against the cellular endonucleases.
[Note .. the printed abstract was wrong and is corrected here]

<>

<1>Sullivan, K.M., Saunders, J.R.
<2>Sequence analysis of the NgoPII methyltransferase gene from Neisseria gonorrhoeae P9:  homologies with other enzymes recognizing the sequence 5'-GGCC-3'.
<3>Nucleic Acids Res.
<4>16
<5>4369-4387
<6>1988
<7>Recombinant plasmids harbouring the functional M.NgoPII methyltransferase (specificity
5'-GGCC-3') were isolated from amplified gene libraries of gonococcal chromosomal DNA cloned
in pBR322 and in Escherichia coli RR1.  The M.NgoPII gene was localized by sub-cloning and the
nucleotide sequence of a cloned 1.6 kb segment of Neisseria gonorrhoeae DNA harbouring the
methylase gene was determined.  This data, coupled with sub-cloning experiments and in vitro
transcription-translation studies, indicates a theoretical size of 38.5 kd for the methylase
protein.  The predicted amino acid sequence of the methylase contains significant regions of
homology with the projected sequences of other cytosine-modifying methylases, upon which the
activity of these enzymes is likely to depend.

<>

<1>Sullivan, K.M., Saunders, J.R.
<2>Determination of the endonuclease and methylase content of Neisseria gonorrhoeae strain P9 and the cloning therefrom of two functional methylase genes.
<3>Gonococci and Meningococci, Kluwer Academic Publishers, Poolman, J.T., Dordrecht, The Netherlands
<4>0
<5>329-334
<6>1988
<7>Neisseria gonorrhoeae strain P9 possesses five DNA cytosine methyltransferases designated
M.NgoPI (M.HaeII isoschizomer), M.NgoPII (M.HaeIII), M.NgoPIII (M.SacII), M.NgoPIV (M.NaeI)
and M.NgoPV.  Two corresponding endonuclease activities, NgoPII (GGCC) and NgoPIII (CCGCGG)
were also detected.  Recombinant plasmids harbouring functional M.NgoPI (PuGCGCPy) and
M.NgoPII methyltransferases were isolated by restriction of an amplified gene library with the
appropriate endonuclease.  After transformation of E. coli RR1, plasmid DNA from individual
transformants was analysed for protection against HaeII or HaeIII respectively to obtain
clones carrying the methylase genes.  It was noted that certain E. coli strains, notably DH1
could not be transformed by plasmids containing the functional M.NgoPI or M.NgoPII genes.

<>

<1>Sullivan, K.M., Saunders, J.R.
<2>Nucleotide sequence and genetic organization of the NgoPII restriction-modification system of Neisseria gonorrhoeae.
<3>Mol. Gen. Genet.
<4>216
<5>380-387
<6>1989
<7>The NgoPII restriction endonuclease, which recognizes the sequences 5'-GG^CC-3', differs
from its isoschizomer HaeIII in being sensitive to methylation at the external cytosine
residue.  The entire nucleotide sequence of a cloned 3.3 kb segment of Neisseria gonorrhoeae
strain P9 chromosomal DNA which harbours the NgoPII restriction-modification system has been
determined. This data, coupled with sub-cloning experiments, indicates that the restriction
endonuclease (R.NgoPII) and modification (M.NgoPII) genes are transcribed from separate
promoters but are arranged in tandem, with the R.NgoPII gene being located on the 5' side of
the M.NgoPII gene.  Unlike all previously reported restriction systems the 3' end of the
endonuclease open reading frame overlaps the 5' end of the methylase open reading frame by 8
codons.  This overlap may have implications for the regulation of the NgoPII
restriction-modification system.

<>

<1>Sullivan, M.B. et al.
<2>Genomic analysis of oceanic cyanobacterial myoviruses compared with T4-like myoviruses from diverse hosts and environments.
<3>Environ. Microbiol.
<4>12
<5>3035-3056
<6>2010
<7>T4-like myoviruses are ubiquitous, and their genes are among the most
abundant documented in ocean systems. Here we compare 26 T4-like genomes,
including 10 from non-cyanobacterial myoviruses, and 16 from marine
cyanobacterial myoviruses (cyanophages) isolated on diverse
Prochlorococcus or Synechococcus hosts. A core genome of 38 virion
construction and DNA replication genes was observed in all 26 genomes,
with 32 and 25 additional genes shared among the non-cyanophage and
cyanophage subsets, respectively. These hierarchical cores are highly
syntenic across the genomes, and sampled to saturation. The 25 cyanophage
core genes include six previously described genes with putative functions
(psbA, mazG, phoH, hsp20, hli03, cobS), a hypothetical protein with a
potential phytanoyl-CoA dioxygenase domain, two virion structural genes,
and 16 hypothetical genes. Beyond previously described cyanophage-encoded
photosynthesis and phosphate stress genes, we observed core genes that may
play a role in nitrogen metabolism during infection through modulation of
2-oxoglutarate. Patterns among non-core genes that may drive niche
diversification revealed that phosphorus-related gene content reflects
source waters rather than host strain used for isolation, and that carbon
metabolism genes appear associated with putative mobile elements. As well,
phages isolated on Synechococcus had higher genome-wide %G+C and often
contained different gene subsets (e.g. petE, zwf, gnd, prnA, cpeT) than
those isolated on Prochlorococcus. However, no clear diagnostic genes
emerged to distinguish these phage groups, suggesting blurred boundaries
possibly due to cross-infection. Finally, genome-wide comparisons of both
diverse and closely related, co-isolated genomes provide a locus-to-locus
variability metric that will prove valuable for interpreting metagenomic
data sets.

<>

<1>Sullivan, M.B., Coleman, M.L., Weigele, P., Rohwer, F., Chisholm, S.W.
<2>Three prochlorococcus cyanophage genomes: signature features and ecological interpretations.
<3>PLoS Biology
<4>3
<5>E144
<6>2005
<7>The oceanic cyanobacteria Prochlorococcus are globally important,
ecologically diverse primary producers. It is thought that their viruses
(phages) mediate population sizes and affect the evolutionary trajectories
of their hosts. Here we present an analysis of genomes from three
Prochlorococcus phages: a podovirus and two myoviruses. The morphology,
overall genome features, and gene content of these phages suggest that
they are quite similar to T7-like (P-SSP7) and T4-like (P-SSM2 and P-SSM4)
phages. Using the existing phage taxonomic framework as a guideline, we
examined genome sequences to establish "core" genes for each phage group.
We found the podovirus contained 15 of 26 core T7-like genes and the two
myoviruses contained 43 and 42 of 75 core T4-like genes. In addition to
these core genes, each genome contains a significant number of
"cyanobacterial" genes, i.e., genes with significant best BLAST hits to
genes found in cyanobacteria. Some of these, we speculate, represent
"signature" cyanophage genes. For example, all three phage genomes contain
photosynthetic genes (psbA, hliP) that are thought to help maintain host
photosynthetic activity during infection, as well as an aldolase family
gene (talC) that could facilitate alternative routes of carbon metabolism
during infection. The podovirus genome also contains an integrase gene
(int) and other features that suggest it is capable of integrating into
its host. If indeed it is, this would be unprecedented among cultured
T7-like phages or marine cyanophages and would have significant
evolutionary and ecological implications for phage and host. Further, both
myoviruses contain phosphate-inducible genes (phoH and pstS) that are
likely to be important for phage and host responses to phosphate stress, a
commonly limiting nutrient in marine systems. Thus, these marine
cyanophages appear to be variations of two well-known phages-T7 and T4-but
contain genes that, if functional, reflect adaptations for infection of
photosynthetic hosts in low-nutrient oceanic environments.

<>

<1>Sullivan, N.L., Septer, A.N., Fields, A.T., Wenren, L.M., Gibbs, K.A.
<2>The Complete Genome Sequence of Proteus mirabilis Strain BB2000 Reveals Differences from the P. mirabilis Reference Strain.
<3>Genome Announcements
<4>1
<5>e00024-13
<6>2013
<7>We announce the complete genome sequence for Proteus mirabilis strain BB2000, a model system
for self recognition. This opportunistic pathogen contains a single,
circular chromosome (3,846,754 bp). Comparisons between this genome and that of
strain HI4320 reveal genetic variations corresponding to previously unknown
physiological and self-recognition differences.

<>

<1>Sumaoka, J., Komiyama, M.
<2>Super restriction enzymes for future biotechnology.
<3>Kino Zairyo
<4>15
<5>19-25
<6>1995
<7>
<>

<1>Sumby, P., Porcella, S.F., Madrigal, A.G., Barbian, K.D., Virtaneva, K., Ricklefs, S.M., Sturdevant, D.E., Graham, M.R., Vuopio-Varkila, J., Hoe, N.P., Musser, J.M.
<2>Evolutionary origin and emergence of a highly successful clone of serotype M1 group A Streptococcus involved multiple horizontal gene transfer   events.
<3>J. Infect. Dis.
<4>192
<5>771-782
<6>2005
<7>To better understand the molecular events involved in the origin of new pathogenic bacteria,
we studied the evolution of a highly virulent clone
of serotype M1 group A Streptococcus (GAS). Genomic, DNA-DNA microarray,
and single-nucleotide polymorphism analyses indicated that this clone
evolved through a series of horizontal gene transfer events that involved
(1) the acquisition of prophages encoding streptococcal pyrogenic exotoxin
A and extracellular DNases and (2) the reciprocal recombination of a 36-kb
chromosomal region encoding the extracellular toxins NAD(+)-glycohydrolase
(NADase) and streptolysin O (SLO). These gene transfer events were
associated with significantly increased production of SLO and NADase.
Virtual identity in the 36-kb region present in contemporary serotype M1
and M12 isolates suggests that a serotype M12 strain served as the donor
of this region. Multiple horizontal gene transfer events were a crucial
factor in the evolutionary origin and emergence of a very abundant
contemporary clone of serotype M1 GAS.

<>

<1>Sumby, P., Smith, M.C.
<2>Genetics of the phage growth limitation (Pgl) system of Streptomyces coelicolor A3(2).
<3>Mol. Microbiol.
<4>44
<5>489-500
<6>2002
<7>The phage growth limitation (Pgl) system, encoded by Streptomyces coelicolor A3(2), confers
protection against the temperate bacteriophage
phiC31 and its homoimmune relatives. The Pgl phenotype is characterized by
the ability of Pgl+ hosts to support a phage burst on initial infection
but subsequent cycles are severely attenuated. Previously, two adjacent
genes pglY and pglZ were shown to be required for Pgl. It had been shown
by Southern blotting that Streptomyces lividans, a close relative of S.
coelicolor and naturally Pgl-, does not contain homologues of pglYZ and
that introduction of pglYZ into S. lividans is not sufficient to confer a
Pgl+ phenotype. Moreover, the mechanism of the Pgl+<--> Pgl- phase
variation associated with this phenotype is also not understood. Here we
describe two novel genes, pglW and pglX, that were shown to be part of
this system by complementation of Pgl- mutants and by insertional
mutagenesis. pglW encodes a 169 kDa protein that includes putative motifs
for both serine/threonine protein kinase activity and DNA binding. pglX
encodes a 136 kDa protein with putative adenine-specific DNA
methyltransferase activity. pglW and pglX have overlapping stop-start
codons suggesting transcriptional and translational coupling. S1 mapping
of transcripts initiating up-stream of pglW indicated that, like pglYZ,
pglWX is expressed in uninfected cultures. A homologue of pglX with 76%
amino acid identity was identified in S. coelicolor, and insertional
mutagenesis indicated that this gene was not required for the Pgl+
phenotype. Southern blots indicated that S. lividans does not contain
homologues of pglW or pglX. A plasmid encoding pglWXYZ was able to confer
the Pgl+ phenotype to S. lividans implying that these four genes
constitute the whole system.

<>

<1>Sumegi, J., Breedveld, D., Hosenlopp, P., Chambon, P.
<2>A rapid procedure for purification of EcoRI endonuclease.
<3>Biochem. Biophys. Res. Commun.
<4>1
<5>78-85
<6>1977
<7>A convenient and rapid procedure has been developed to purify restriction
endonuclease EcoRI.  The method involves sonication of cells at low ionic
strength, precipitation of the endonuclease with Polymin P (a
polyethyleneimine), elution of the enzyme from the Polymin P precipitate,
ammonium sulfate precipitation and chromatography on phosphocellulose.  The
purified restriction endonuclease is free of exonuclease and other
endonucleases.

<>

<1>Summer, E.J., Gonzalez, C.F., Bomer, M., Carlile, T., Embry, A., Kucherka, A.M., Lee, J., Mebane, L., Morrison, W.C., Mark, L., King, M.D., LiPuma, J.J., Vidaver, A.K., Young, R.
<2>Divergence and mosaicism among virulent soil phages of the Burkholderia cepacia complex.
<3>J. Bacteriol.
<4>188
<5>255-268
<6>2006
<7>We have determined the genomic sequences of four virulent myophages, Bcep1, Bcep43, BcepB1A,
and Bcep781, whose hosts are soil isolates of the
Burkholderia cepacia complex. Despite temporal and spatial separations
between initial isolations, three of the phages (Bcep1, Bcep43, and
Bcep781, designated the Bcep781 group) exhibit 87% to 99% sequence
identity to one another and most coding region differences are due to
synonymous nucleotide substitutions, a hallmark of neutral genetic drift.
Phage BcepB1A has a very different genome organization but is clearly a
mosaic with respect to many of the genes of the Bcep781 group, as is a
defective prophage element in Photorhabdus luminescens. Functions were
assigned to 27 out of 71 predicted genes of Bcep1 despite extreme sequence
divergence. Using a lambda repressor fusion technique, 10 Bcep781-encoded
proteins were identified for their ability to support homotypic
interactions. While head and tail morphogenesis genes have retained
canonical gene order despite extreme sequence divergence, genes involved
in DNA metabolism and host lysis are not organized as in other phages.
This unusual genome arrangement may contribute to the ability of the
Bcep781-like phages to maintain a unified genomic type. However, the
Bcep781 group phages can also engage in lateral gene transfer events with
otherwise unrelated phages, a process that contributes to the
broader-scale genomic mosaicism prevalent among the tailed phages.

<>

<1>Summers, M.F., Powell, C., Egan, W., Byrd, R.A., Wilson, W.D., Zon, G.
<2>Alkyl phosphotriester modified oligodeoxyribonucleotides.  VI. NMR and UV spectroscopic studies of ethyl phosphotriester (Et) modified Rp-Rp and Sp-Sp duplexes, {d[GGAA(Et)TTCC]}2.
<3>Nucleic Acids Res.
<4>14
<5>7421-7436
<6>1986
<7>1H NMR chemical shift assignments for the title compounds were made for all but
a few H5' and H5" signals using two-dimensional nuclear Overhauser effect
(2D-NOE) data, which was also used for the first time to assign absolute
configuration at phosphorus.  The chemical shifts were, in general, similar to
those reported [Broido, M.S., et al. (1985) Eur. J. Biochem. 150, 117-128] for
the B-like conformation of the unmodified, parent duplex, {d(GGAATTCC)]2.
Differences in chemical shifts for corresponding protons were mostly localized
to the AA(Et)TT region, and showed some stereochemical dependence.  Unambiguous
assignment of the phosphotriester 31P signals was achieved in a novel way using
selective insensitive nucleus enhancement by polarization transfer (selective
INEPT) NMR.  The Rp-Rp duplex melted ca. 11C lower than either the Sp-Sp or
parent duplexes, as evidenced by Tm and variable temperature 1H/31P NMR
measurements.  The 2D-NOE data for the Rp-Rp duplex suggested possible steric
interactions between the ethyl group and the H3' of the flanking A residue.  At
low ionic strength, the Sp-Sp and parent duplexes had similar stability but at
high ionic strength the Sp-Sp duplex was less stable.

<>

<1>Sun, D.K., Yoo, O.J.
<2>Purification and characterization of a restriction endonuclease ZanI from Zymomonas anaerobia.
<3>Korean Biochem. J.
<4>21
<5>419-422
<6>1988
<7>We described the purification and characterization of a sequence restriction endonuclease,
ZanI, found from Zymomonas anaerobia (NCI B8227). The purified enzyme is essentially
homogeneous and the subunit molecular weight is 30,000+/- 1,000 as judged by 10%
polyacrylamide gel electrophoresis containing 0.1% SDS. The recognition sequence and the
cleavage site (indicated by the arrow) were determined to be 5'-CC^(AT) GG-3', the same
sequence as recognized by BstNI and EcoRII. ZanI endonuclease is able to cleave dcm-methylated
DNA and is maximally active at 37C.

<>

<1>Sun, H. et al.
<2>Complete genome sequence of Nocardiopsis dassonvillei type strain (IMRU 509).
<3>Standards in Genomic Sciences
<4>3
<5>325-336
<6>2010
<7>Nocardiopsis dassonvillei (Brocq-Rousseau 1904) Meyer 1976 is the type species of the genus
Nocardiopsis, which in turn is the type genus of the family
Nocardiopsaceae. This species is of interest because of its ecological
versatility. Members of N. dassonvillei have been isolated from a large variety
of natural habitats such as soil and marine sediments, from different plant and
animal materials as well as from human patients. Moreover, representatives of the
genus Nocardiopsis participate actively in biopolymer degradation. This is the
first complete genome sequence in the family Nocardiopsaceae. Here we describe
the features of this organism, together with the complete genome sequence and
annotation. The 6,543,312 bp long genome consist of a 5.77 Mbp chromosome and a
0.78 Mbp plasmid and with its 5,570 protein-coding and 77 RNA genes is a part of
the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Sun, H. et al.
<2>Complete genome sequence of Desulfarculus baarsii type strain (2st14).
<3>Standards in Genomic Sciences
<4>3
<5>276-284
<6>2010
<7>Desulfarculus baarsii (Widdel 1981) Kuever et al. 2006 is the type and only species of the
genus Desulfarculus, which represents the family Desulfarculaceae
and the order Desulfarculales. This species is a mesophilic sulfate-reducing
bacterium with the capability to oxidize acetate and fatty acids of up to 18
carbon atoms completely to CO(2). The acetyl-CoA/CODH (Wood-Ljungdahl) pathway is
used by this species for the complete oxidation of carbon sources and autotrophic
growth on formate. The type strain 2st14(T) was isolated from a ditch sediment
collected near the University of Konstanz, Germany. This is the first completed
genome sequence of a member of the order Desulfarculales. The 3,655,731 bp long
single replicon genome with its 3,303 protein-coding and 52 RNA genes is a part
of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Sun, J., He, Z., Nechushtai, R., Chitnis, P.R.
<2>Molecular cloning of the PsaA and PsaB genes for the core proteins of photosystem I from the thermophilic cyanobacterium Mastigocladus Iaminosus (Accession No. AF038558).
<3>Plant Physiol.
<4>116
<5>1192
<6>1998
<7>
<>

<1>Sun, J., Weinstein, H.
<2>The restriction enzyme BamHI is poised for sliding along DNA in a model of the nonspecific complex.
<3>Biophys. J.
<4>82
<5>463a
<6>2002
<7>The mechanism by which DNA-binding proteins find their specific binding sites is unclear.  To
understand how the protein reaches its cognate site within a long piece of DNA, the crystal
structures of the nonspecific and the specific BamHI-DNA complexes were used as templates to
study the electrostatic interactions and possible sliding pathways.  A model is proposed for
the initial nonspecific binding of BamHI to a long stretch of B-form DNA, in which BamHI and
DNA adopt an orientation different from that in the crystal structures.  This arrangement of a
nonspecific complex is supported by the salt-dependence of the interaction, which is in good
agreement with experimental results if calculated in this model, but not in the crystal
structure.  The electrostatic interaction calculated for different orientations of BamHI
relative to DNA appears to be sufficient to steer the protein to this favorable configuration
parallel to the DNA axis.  The results are independent of DNA sequence, so that BamHI could
bind equally to any site along DNA thus enabling it to slide.  The calculations suggest that
the DNA helical structure could affect the pathway of sliding: the preference for sliding
along the DNA helix pitch stems from the ~2.5kcal/mol energy barrier that opposes sliding
along one face of DNA.

<>

<1>Sun, K., Jiao, X.D., Zhang, M., Sun, L.
<2>DNA adenine methylase is involved in the pathogenesis of Edwardsiella tarda.
<3>Vet. Microbiol.
<4>141
<5>149-154
<6>2010
<7>Edwardsiella tarda is a serious aquaculture pathogen that can infect many Cultured fish
species. The aim of this study was to investigate
the potential importance of DNA adenine methylase (Dam) in E. tarda
pathogenesis. The E. tardo dam gene (dam(Et)) was cloned from a
pathogenic strain, TXD1, isolated from diseased fish. Dam(Et) shares
high (70.2%) sequence identity with the Dam proteins of Yersinia
enterocolitica and several other bacterial species. Recombinant Dam(Et)
is able to complement a dam-deficient Escherichia coli strain and
methylate the genomic DNA. Attenuation of dam(Et) expression by
antisense RNA interference had no apparent effect on the growth of
TXD1, but caused significant attenuation of overall bacterial virulence
and altered several stress responses including spontaneous mutation,
recovering from UV radiation and H2O2 exposure, binding to host mucus,
and dissemination in host blood and liver. In addition, attenuation of
dam(Et) expression increased luxS expression and AI-2 activities in E.
tarda. These results indicate that Dam(Et) is a virulence determinant
and plays a role in the pathogenesis of TXD1, and that temporal
expression of dam(Et), is essential for optimal bacterial infection.

<>

<1>Sun, L.
<2>Kinetic studies on cleavage of adenovirus type 5 DNA by restriction endonuclease EcoRI.
<3>Xibei Daxue Xuebao
<4>19
<5>71-76
<6>1989
<7>The two EcoRI restriction sites on Adenovirus Type 5 DNA have been affirmed and relative
lengths of EcoRI digest products of Ad5 DNA are 77%, 7%, 16% for the fragment A, C, B
respectively. The kinetic of cleavage of Ad5 DNA by EcoRI was studied using quantitative
evaluation of the ethidium bromide fluorescence of DNA fragments separated on agarose gel. The
overall rate constants of cleavage at different EcoRI site have been determined. The rate
constants for two cleavage sites depend on the enzyme concentration and reaction temperature.
Thermodynamic studies have shed light on the importance of the nucleotide sequences around the
restriction sites, which modulate the activation barriers for the cleavage process itself.

<>

<1>Sun, L., Wang, Y., Yu, C., Zhao, Y., Gan, Y.
<2>Genome Sequence of Clostridium tunisiense TJ, Isolated from Drain Sediment from a Pesticide Factory.
<3>J. Bacteriol.
<4>194
<5>6950-6951
<6>2012
<7>Clostridium tunisiense is a Gram-positive, obligate anaerobe that was first isolated in an
anaerobic evironment under eutrophication. Here we report the
first genome sequence of the Clostridium tunisiense TJ isolated from drain
sediment of a pesticide factory in Tianjin, China. The genome is of great
importance for both basic and application research.

<>

<1>Sun, L., Yin, J.
<2>Initial studies on chemical modification of several amino acid residues of the restriction endonuclease XhoI.
<3>Xibei Daxue Xuebao
<4>20
<5>65-71
<6>1990
<7>The restriction endonuclease XhoI can be inactivated by some specific protein
inhibitors.  XhoI is sensitive not only to sulfhydryl reagents but also to
reagents that modify lysine and arginine residues.  Inactivation of the enzyme
obeyed pseudo-first-order kinetics and resulted in complete elimination of
catalysis.  Lambda DNA as substrate is strongly protected against inactivation
by the reagents.  These data suggest that XhoI has essential lysine and/or
arginine residues and SH groups which are crucial to the enzymatic activity.

<>

<1>Sun, L.-K., Hong, R.
<2>The influence of DNA conformation and base composition flanking recognition sites on the cleavage rate of restriction.
<3>Acta Biochim. Biophys. Sin.
<4>21
<5>456-460
<6>1989
<7>The relations between the nucleotide sequences adjacent to the recognition sites and the
cleavage rate of DNA with restriction endonucleases were investigated.  Our data indicate that
the recognition sites flanked by the A-T rich sequences are cleaved faster than those flanked
by G-C rich sequences regardless of the DNA conformation.

<>

<1>Sun, L.-K., Ruan, H.
<2>Kinetics and thermodynamics of specific and non-specific interactions between BglI restriction endonuclease and pBR322-DNA.
<3>Chinese Biochem. J.
<4>6
<5>334-338
<6>1990
<7>Cleavage of pBR322-DNA by BglI was studied at different temperatures and various incubation
times. Kinetic constants and thermodynamic parameters of the reaction were evaluated.  The
similarity between linear and circular DNA in specific and non-specific interaction with
the restriction endonuclease has been demonstrated by their kinetic constants and
thermodynamic parameters. The specific binding is about 2 orders of magnitude stronger than
non-specific binding. Ks is more dependent on temperature then KN. Complex formation is
weakened with increasing temperature. The values of enthalpy and entropy show that both
specific and non-specific binding need no activation energy and the "disorder" of the molecule
is decreased with binding. The key element that limits the cleavage rate is Kc.  From
the experimental results, it is evident that the recognition site adjacent to the A-T
rich sequences has a low activation energy for cleavage and can be cleaved fast. This is
probably due to the fact that the sequences adjacent to the recognition site may regulate the
activation threshold of the cleavage.

<>

<1>Sun, L.-K., Wang, X., Hong, R.
<2>Calculation of cleavage rates of restriction endonuclease on circular DNA.
<3>Acta Biophys. Sinica
<4>4
<5>64-66
<6>1988
<7>In this paper, we have deduced a set of equations to calculate the cleavage rates of
restriction endonucleases on circular DNA by mathematical and physiochemical methods. They are
important to study the kinetics and modification of restriction endonucleases.

<>

<1>Sun, Q., Lan, R., Wang, Y., Wang, J., Wang, Y., Li, P., Du, P., Xu, J.
<2>Isolation and genomic characterization of SfI, a serotype-converting bacteriophage of Shigella flexneri.
<3>BMC Microbiol.
<4>13
<5>39
<6>2013
<7>ABSTRACT: BACKGROUND: All Shigella flexneri serotypes except serotype 6 share a
common O-antigen tetrasaccharide backbone and nearly all variation between
serotypes are due to glucosyl and/or O-acetyl modifications of the common O units
mediated by glycosyltransferases encoded by serotype-converting bacteriophages.
Several S. flexneri serotype-converting phages including SfV, SfX, Sf6 and SfII
have been isolated and characterized. However, S. flexneri serotype-converting
phage SfI which encodes a type I modification of serotype 1 (1a, 1b, 1c and 1d)
had not yet been characterized. RESULTS: The SfI phage was induced and purified
from a S. flexneri serotype 1a clinical strain 019. Electron microscopy showed
that the SfI phage has a hexagonal head and a long contractile tail,
characteristic of the members of Myoviridae family. SfI can convert serotype Y to
serotype 1a and serotype X to serotype 1d, but cannot convert 10 other S.
flexneri serotypes (1a, 1b, 2a, 2b, 3a, 3b, 4a, 4b, 5a, Xv) tested, suggesting
that SfI has a narrow host range. Similar to other S. flexneri
serotype-converting phages, SfI integrates into the tRNA-thrW gene adjacent to
proA of the host chromosome when lysogenized. The complete sequence of the SfI
genome was 38,389 bp, encoding 66 open reading frames and two tRNA genes. Phage
SfI shares significant homology with S. flexneri phage SfV, Escherichia coli
prophage e14 and lambda, and is classified into the lambdoid phage family. SfI
was found to use a cos mechanism for DNA packaging similar to that of phage SfV.
CONCLUSIONS: SfI contains features of lambdoid phages and is closely related to
S. flexneri phage SfV, E. coli prophage e14 and lambda. The characterization of
SfI enhances our understanding of serotype conversion of S. flexneri.

<>

<1>Sun, S., Yang, X., Yuan, Y., Dai, X., Yan, Y., Cao, H., Luo, T., Guo, R., Wang, X., Song, Y., Yang, R., Zhang, Y., Cui, Y.
<2>Draft Genome Sequence of Yersinia pestis Strain 2501, an Isolate from the Great Gerbil Plague Focus in Xinjiang, China.
<3>J. Bacteriol.
<4>194
<5>5447-5448
<6>2012
<7>We deciphered the genome of Yersinia pestis strain 2501, isolated from the Junggar Basin, a
newly discovered great gerbil plague focus in Xinjiang, China.
The total length of assembly was 4,597,322 bp, and 4,265 coding sequences were
predicted within the genome. It is the first Y. pestis genome from this plague
focus.

<>

<1>Sun, Y., Song, Y., Song, H., Liu, J., Wang, P., Qiu, S., Chen, S., Zhu, L., Ji, X., Wang, Z., Liu, N., Xia, L., Chen, W., Feng, S.
<2>Complete Genome Sequence of an Acinetobacter Strain Harboring the NDM-1 Gene.
<3>Genome Announcements
<4>1
<5>e00023-12
<6>2013
<7>The NDM-1 gene is a significant public health concern. Acinetobacter is one of the most
prevalent opportunistic pathogens causing recent nosocomial infections
with NDM-1, and drug-resistant strains pose serious threats to public health
worldwide. Herein, we present the genomic sequence of Acinetobacter calcoaceticus
subsp. anitratus XM1570, a multidrug-resistant isolate that carries the blaNDM-1
gene.

<>

<1>Sun, Z., Chen, X., Wang, J., Gao, P., Zhou, Z., Ren, Y., Sun, T., Wang, L., Meng, H., Chen, W., Zhang, H.
<2>Complete genome sequence of probiotic Bifidobacterium animalis subsp. lactis strain V9.
<3>J. Bacteriol.
<4>192
<5>4080-4081
<6>2010
<7>Bifidobacterium animalis subsp. lactis strain V9 is a Chinese commercial bifidobacteria with
several probiotic functions. It was isolated from a healthy Mongolian child in China. We
present here the complete genome sequence of V9 and compared it to 3 other published genomes
of B. animalis subsp. lactis strains. The result indicates the lack of polymorphism among
strains of this subspecies of different continent origins.

<>

<1>Sun, Z., Chen, X., Wang, J., Zhao, W., Shao, Y., Guo, Z., Zhang, X., Zhou, Z., Sun, T., Wang, L., Meng, H., Zhang, H., Chen, W.
<2>Complete genome sequence of Lactobacillus delbrueckii subsp. bulgaricus strain ND02.
<3>J. Bacteriol.
<4>193
<5>3426-3427
<6>2011
<7>Lactobacillus delbrueckii subsp. bulgaricus strain ND02 is a Chinese commercial dairy starter
used for the manufacture of yoghurt. It was
isolated from the naturally fermented yak milk in Qinghai, China. Here, we
report the main genome features of ND02 and several differences with two
other published genomes of Lactobacillus delbrueckii subsp. bulgaricus
strains.

<>

<1>Sun, Z., Chen, X., Wang, J., Zhao, W., Shao, Y., Wu, L., Zhou, Z., Sun, T., Wang, L., Meng, H., Zhang, H., Chen, W.
<2>Complete genome sequence of Streptococcus thermophilus strain ND03.
<3>J. Bacteriol.
<4>193
<5>793-794
<6>2010
<7>Streptococcus thermophilus strain ND03 is a Chinese commercial dairy starter used for the
manufacture of yoghurt. It was isolated from the naturally fermented yak milk in Qinghai,
China. We present here the complete genome sequence of ND03 and compared it to 3 other
published genomes of Streptococcus thermophilus strains.

<>

<1>Sun, Z.Y., Terragni, J., Borgaro, J.G., Liu, Y.W., Yu, L., Guan, S.X., Wang, H., Sun, D.P., Cheng, X.D., Zhu, Z.Y., Pradhan, S., Zheng, Y.
<2>High-Resolution Enzymatic Mapping of Genomic 5-Hydroxymethylcytosine in Mouse Embryonic Stem Cells.
<3>Cell Rep.
<4>3
<5>567-576
<6>2013
<7>We describe the use of a unique DNA-modification-dependent restriction endonuclease AbaSI
coupled with sequencing (Aba-seq) to map
high-resolution hydroxymethylome of mouse E14 embryonic stem cells. The
specificity of AbaSI enables sensitive detection of
5-hydroxymethylcytosine (5hmC) at low-occupancy regions. Bioinformatic
analysis suggests 5hmCs in genic regions closely follow the 5mC
distribution. 5hmC is generally depleted in CpG islands and only
enriched in a small set of repetitive elements. A regularly spaced and
oscillating 5hmC pattern was observed at the binding sites of CTCF.
5hmC is enriched at the poised enhancers with the monomethylated
histone H3 lysine 4 (H3K4me1) marks, but not at the active enhancers
with the acetylated histone H3 lysine 27 (H3K27Ac) marks. Non-CG
hydroxymethylation appears to be prevalent in the mitochondrial genome.
We propose that some amounts of transiently stable 5hmCs may indicate a
poised epigenetic state or demethylation intermediate, whereas others
may suggest a locally accessible chromosomal environment for the TET
enzymatic apparatus.

<>

<1>Sung, J.S., Chun, J., Choi, S., Park, W.
<2>Genome Sequence of the Halotolerant Staphylococcus sp. Strain OJ82, Isolated from Korean Traditional Salt-Fermented Seafood.
<3>J. Bacteriol.
<4>194
<5>6353-6354
<6>2012
<7>Staphylococcus sp. strain OJ82 was isolated from a Korean traditional fermented squid seafood,
ojingeo-jeotgal. Staphylococcus sp. OJ82 could grow and show
extracellular protease and beta-galactosidase activities in the presence of
extremely high saline (20%). Here, we report the genome sequence of
Staphylococcus sp. OJ82.

<>

<1>Surani, M.A.
<2>Imprinting and the initiation of gene silencing in the germ line.
<3>Cell
<4>93
<5>309-312
<6>1998
<7>Inheritance of genes in active or repressed states requires appropriate chromosomal
modifications and DNA methylation.  Imprinting of a determined state into the chromatin
structure therefore constitutes a memory of all developmental decisions taken within
individual cells.  Genomic or gametic imprinting is however a unique manifestation of such
epigenetic inheritance in which expression of certain genes is governed by their parental
origin, from generation to generation.  Some of these genes show expression after paternal
inheritance while others are expressed only when inherited from the maternal germ line.  The
most striking feature of imprinted genes therefore is that the active and inactive parental
alleles coexist within individual cells.  Over 20 imprinted genes have so far been identified.
Parental genomes are functionally nonequivalent during development because of the differential
expression of imprinted genes. A review.

<>

<1>Surby, M.A.
<2>Facilitated diffusion of the EcoRI DNA methyltransferase under both catalytic and noncatalytic conditions and the potential of Z DNA disrupt protein translocation on DNA.
<3>Diss. Abstr.
<4>57
<5>5631B-5632B
<6>1997
<7>Previous experiments in our lab have suggested that the EcoRI DNA methyltransferase might
utilize one-dimensional facilitated diffusion to locate its specific binding site on a DNA
substrate.  In the current study, initial experiments to characterize this phenomenon with
regard to the methyltransferase focus on the contribution of nonspecific DNA to enzyme
efficiency (kcat/Km).  This parameter increases 4-fold as DNA length increases from 14 to 429
basepairs and increases 2-fold as the distance from the site to the nearest end is increased
from 29 to 378 basepairs.  No changes in kcat/Km result from further increases in either case.
A facilitated diffusion mechanism is proposed in which the methyltransferase scans an average
of <400 base pairs prior to dissociation from a DNA molecule.  The methyltransferase was found
to methylate two sites on a single DNA molecule in a distributive rather than a processive
manner, suggesting that the enzyme dissociates from the DNA prior to release of the reaction
product S-adenosylhomocysteine.  A direct competition experiment with the EcoRI endonuclease
shows the methyltransferase to be slightly more efficient at specific site location and
catalysis.  A rationale for the role of facilitated diffusion in this type II
restriction-modification system is proposed.  In addition, the contribution of nonspecific DNA
to binding parameters (d, koff, and kon) was determined for the methyltransferase under
noncatalytic conditions.  An increase in DNA size from 14 to 775 base pairs causes a 20-fold
decrease in Kd, while koff remains constant over the same range.  The calculated kon increases
with longer substrates, consistent with a facilitated diffusion mechanism.  However, the
combined results deviate from the model developed to describe facilitated diffusion.  Our
results were successfully simulated using numerical integration of a kinetic scheme invoking
protein dissociation via the ends of DNA.  Consistent with this scheme, the methyltransferase
dissociates more slowly from a circularized DNA molecule than from the identical linearized
form.  The final set of experiments is an attempt to determine the effect of Z DNA on the
ability of proteins to translocate on DNA.  Z DNA formation in a linear substrate is confirmed
with Z DNA-specific antibody binding and preliminary results with the EcoRI endonuclease
suggest that Z DNA decreases the rate of DNA cleavage by 2- to 3-fold.  However, subsequent
experiments with more highly purified endonuclease do not support this conclusion.  Z DNA
cannot be detected in the linear substrates with chemical modification techniques, so the
assay was modified to use the methyltransferase and a supercoiled DNA substrate.  Although
there is an observable effect on kcat and Km in the supercoiled substrate, control experiments
make an assignment of this effect to the presence of Z DNA unclear.

<>

<1>Surby, M.A., Reich, N.O.
<2>The role of nonspecific DNA in the site location kinetics of the EcoRI DNA methyltransferase.
<3>FASEB J.
<4>6
<5>A356
<6>1992
<7>The EcoRI DNA methyltransferase is an extremely efficient enzyme, with a
kcat/Km of 4.1 X 10/8 s-1M-1 for plasmid DNA.  Its kcat/Km decreases to 0.51 X
10/8 s-1M-1 when a 14 basepair synthetic oligonucleotide is used as a
substrate.  A possible explanation for these observations is that the methylase
utilizes facilitated diffusion along nonspecific DNA as a mechanism for site
location.  This was first tested using a processivity assay developed for the
EcoRI endonuclease (J. Biol. Chem., Oct. 25, 1985, 260 (24) pp 13130-13137).
In contrast to our results with the endonuclease, no processive methylation was
observed with this assay.  The role of facilitated diffusion in initial site
location was then investigated by kinetic analysis of restriction fragments
ranging in size from 4363 to 108 basepairs, all of which contained a single
EcoRI site.  There were no differences observed in the rate of methyl group
incorporation for the various substrates.  This leads us to conclude that the
specificity enhancement observed for the plasmid DNA is due either to
nonspecific sequences <108 basepairs away from the canonical site of the DNA
substrate or to some other phenomenon (e.g. flanking sequence differences).

<>

<1>Surby, M.A., Reich, N.O.
<2>The role of nonspecific DNA in the site-location kinetics of the EcoRI DNA methyltransferase.
<3>Biochemistry
<4>31
<5>2208
<6>1992
<7>The EcoRI DNA methyltransferase is an extremely efficient enzyme, with a
kcat/Km of 4.1 x 10/8 S-1M-1 for plasmid DNA.  Its kcat/Km decreases to 0.51 x
10.8 S-1 M-1 when a 14-base-pair synthetic oligonucleotide is used as a
substrate.  Canonical site flanking sequence differences between the plasmid
and the 14-bp oligonucleotide were shown to have no effect on kcat/Km.  A
possible explanation for these observations is that the methylase utilizes
facilitated diffusion along nonspecific DNA as a mechanism for site location.
This was first tested using a processivity assay developed for the EcoRI
endonuclease [(1985) J. Biol. Chem. 260 (24), 13130-13137].  In contrast to our
results with the endonuclease, no processive methylation was observed with this
assay.  The role of facilitated diffusion in initial site location was then
investigated by kinetic analysis of restriction fragments ranging in size from
4363 to 108 bp, all of which contained a single EcoRI site.  There were no
differences observed in the rate of methyl group incorporation for various
substrates.  This leads us to conclude that the specificity enhancement
observed for the plasmid DNA is due to nonspecific sequences <108 bp away from
the canonical site of the DNA substrate.

<>

<1>Surby, M.A., Reich, N.O.
<2>The role of nonspecific DNA in the site-location kinetics of the EcoRI DNA methyltransferase.
<3>ACS Abstracts
<4>203
<5>110-BIOL
<6>1992
<7>The EcoRI DNA methyltransferase is an extremely efficient enzyme, with a kcat/Km of 4.1 x 10/8
S-1M-1 for plasmid DNA. Its kcat/Km decreases to 0.51 x 10/8 S-1M-1 when a 14 basepair
synthetic oligonucleotide is used as a substrate. Cannonical site flanking sequence
differences between the plasmid and the 14 basepair oligonucleotide were shown to have no
effect on kcat/Km. A possible explanation for these observations is that the methylase
utilizes facilitated diffusion along nonspecific DNA as a mechanism for site location. This
was first tested using a processivity assay developed for the EcoRI endonuclease (J. Biol.
Chem., Oct. 25,1985, 260 (24), pp 13130-13137). In contrast to our results with the
endonuclease, no processive methylation was observed with this assay. The role of facilitated
diffusion in initial site location was then investigated by kinetic analysis of restriction
fragments ranging in size from 4363 to 108 basepairs, all of which contained a single EcoRI
site. There were no differences observed in the rate of methyl group incorporation for the
various substrates. This leads us to conclude that the specificity enhancement observed for
the plasmid DNA is due to nonspecific sequences <108 basepairs aways from the canonical site
of the DNA substrate.

<>

<1>Surby, M.A., Reich, N.O.
<2>Facilitated diffusion of the EcoRI DNA methyltransferase is described by a novel mechanism.
<3>Biochemistry
<4>35
<5>2209-2217
<6>1996
<7>The contribution of nonspecific DNA to binding parameters (Kd, koff, and kon) was determined
for the EcoRI DNA methyltransferase under noncatalytic conditions.  An increase in DNA size
from 14 to 775 base pairs causes a 20-fold decrease in Kd, while koff remains constant over
the same range.  The calculated kon increases with longer substrates, consistent with a
facilitated diffusion mechanism.  However, the combined results deviate from the model
developed to describe facilitated diffusion.  Our results were successfully simulated using
numerical integration of a kinetic scheme invoking protein dissociation via the ends of DNA.
Consistent with this scheme, the methyltransferase dissociates more slowly from a circularized
DNA molecule than from the identical linearized form.  The simulation strategy correctly
models our data with the methyltransferase and should be generally useful for routine modeling
of facilitated diffusion involving protein-DNA systems.

<>

<1>Surby, M.A., Reich, N.O.
<2>Contribution of facilitated diffusion and processive catalysis to enzyme efficiency: Implications for the EcoRI restriction-modification system.
<3>Biochemistry
<4>35
<5>2201-2208
<6>1996
<7>The contribution of nonspecific DNA to enzyme efficiency (kcat/Km) is described for a
sequence-specific DNA-modifying enzyme.  Our investigation focuses on the EcoRI DNA
methyltransferase which transfers a methyl group from the cofactor S-adenosylmethionine to the
second adenine in the double-stranded DNA sequence GAATTC.  kcat/Km increases 4-fold as DNA
length increases from 14 to 429 base pairs and increases 2-fold as the distance from the site
to the nearest end is increased from 29 to 378 base pairs.  No changes in kcat/Km result from
further increases in either case.  A facilitated diffusion mechanism is proposed in which the
methyltransferase scans an average of <400 base pairs prior to dissociation from a DNA
molecule.  The methyltransferase was found to methylate two sites on a single DNA molecule in
a distributive rather than a processive manner, suggesting that the enzyme dissociates from
the DNA prior to release of the reaction product S-adenosylhomocysteine.  A direct competition
experiment with the EcoRI endonuclease shows the methyltransferase to be slightly more
efficient at specific site location and catalysis.  A rationale for the role of facilitated
diffusion in this type II restriction-modification system is proposed.

<>

<1>Suri, B., Bickle, T.A.
<2>EcoA:  The first member of a new family of Type I restriction modification systems - Gene organization and enzymatic activities.
<3>J. Mol. Biol.
<4>186
<5>77-85
<6>1985
<7>The characterization of the EcoA restriction-modification enzymes from
Escherichia coli 15T- is described.  The reactions catalysed by these enzymes
are very similar to those catalysed by the classical type I restriction and
modification enzymes, a family of genetically related proteins.  The detailed
mechanisms, particularly for DNA modification, differ.  The genetic and
transcriptional organizations are also very similar to those of the classical
systems, despite the fact that EcoA is not allelic to the others.  We
demonstrate that the expression of the EcoA genes is controlled following
conjugative transfer to other strains in such a way that no lethality is
observed, probably because the reciptient chromosome is completely modified
before restriction activity is expressed.

<>

<1>Suri, B., Nagaraja, V., Bickle, T.A.
<2>Bacterial DNA modification.
<3>Curr. Top. Microbiol. Immunol.
<4>108
<5>1-9
<6>1984
<7>As first proposed by Arber (1965), DNA restriction/modification systems (R/M
systems) are mediated by endonucleases and DNA methylases that recognize the
same DNA sequences.  The endonuclease recognizes its specific sequence as a
signal to cleave the DNA unless the sequence has been previously methylated by
the modification enzyme.  Chromosomal DNA from cells harboring the R/M system
is normally methylated, and is thus not a substrate for the restriction enzyme.
Foreign DNA lacking the specific methylation pattern and introduced into the
cell by phase infection, conjugation, or transformation is the only known
natural substrate for restriction.  R/M systems can therefore be considered
primitive prokaryotic analogues of the eukaryotic immune system.  A vast body
of literature on the genetics and biochemistry of R/M systems has accumulated
since they were first investigated 20 years ago (Arber and Dussoix 1962), and
it is now clear that R/M systems can be conveniently classified into three
types (Boyer 1971; Kauc and Piekarowicz 1978; Nathans and Smith 1975).  The
most complicated of these are the type-I systems, which are mediated by
complex, multifunctional enzymes and which were the first proteins shown to
recognize specific DNA sequences.  The restriction enzymes EcoK and EcoB from
the Escherichia coli strains K12 and B are the two prototypes, and are still
the only ones to have been studied in detail.

<>

<1>Suri, B., Shepherd, J.C.W., Bickle, T.A.
<2>The EcoA restriction and modification system of Escherichia coli 15T-: enzyme structure and DNA recognition sequence.
<3>EMBO J.
<4>3
<5>575-579
<6>1984
<7>The EcoA restriction enzyme from Escherichia coli 15T- has been isolated.  It
proves to be an unusual enzyme, clearly related functionally to the classical
type I restriction enzymes.  The basic enzyme is a two subunit modification
methylase.  Another protein species can be purified which by itself has not
enzymatic activities but which converts the modification methylase to an ATP
and S-adenosylmethionine-dependent restriction endonuclease.  The DNA
recognition sequence of EcoA has an overall structure that is very similar to
previously determined type I sequences.  It is:
5'-GAGNNNNNNNGTCA-3'3'-CTCNNNNNNNCAGT-5'where N can be any nucleotide.
Modification methylates the adenosyl residue in the specific trinucleotide and
the adenosyl residue in the lower strand of the specific tetranucleotide.

<>

<1>Susanti, D. et al.
<2>Complete Genome Sequence of Desulfurococcus fermentans, a Hyperthermophilic Cellulolytic Crenarchaeon Isolated from a Freshwater Hot Spring in Kamchatka,  Russia.
<3>J. Bacteriol.
<4>194
<5>5703-5704
<6>2012
<7>Desulfurococcus fermentans is the first known cellulolytic archaeon. This hyperthermophilic
and strictly anaerobic crenarchaeon produces hydrogen from
fermentation of various carbohydrates and peptides without inhibition by
accumulating hydrogen. The complete genome sequence reported here suggested that
D. fermentans employs membrane-bound hydrogenases and novel glycohydrolases for
hydrogen production from cellulose.

<>

<1>Sussenbach, J.S., Monfoort, C.H., Schiphof, R., Stobberingh, E.E.
<2>A restriction endonuclease from Staphylococcus aureus.
<3>Nucleic Acids Res.
<4>3
<5>3193-3202
<6>1976
<7>A specific endonuclease, Sau3AI, has been partially purified from
Staphylococcus aureus strain 3A by DEAE-cellulose chromatography.  The enzyme
cleaves adenovirus type 5 DNA many times, SV40 DNA eight times but does not
cleave double-stranded PhiX174 DNA.  It recognizes the sequence 5' ^-G-A-T-C-
3' 3'  -C-T-A-G-^ 5'  and cleaves as indicated by the arrows.  Evidence is
presented that this enzyme plays a role in the biological
restriction-modification system of Staphylococcus aureus strain 3A.

<>

<1>Sussenbach, J.S., Steenbergh, P.H., Rost, J.A., van Leeuwen, W.J., van Embden, J.D.A.
<2>A second site-specific restriction endonuclease from Staphylococcus aureus.
<3>Nucleic Acids Res.
<4>5
<5>1153-1163
<6>1978
<7>A site-specific restriction endonuclease has been isolated from Staphylococcus
aureus PS 96.  This enzyme, Sau96I, recognizes the DNA sequence
5'--G-^G-N-C-C--3' 3'--C-C-N-G-^G--5' and cleaves as indicated by the arrows.
The enzyme cleaves adenovirus type 5 and lambda DNA many times, SV40 DNA 10
times and PhiX174 RF DNA 2 times.  Evidence is presented that the enzyme is
involved in biological restriction-modification.

<>

<1>Sussman, D., Chadsey, M., Fauce, S., Engel, A., Bruett, A., Monnat, R., Stoddard, B.L., Seligman, L.M.
<2>Isolation and characterization of new homing endonuclease specificities at individual target site positions.
<3>J. Mol. Biol.
<4>342
<5>31-41
<6>2004
<7>Homing endonucleases are highly specific DNA endonucleases, encoded within mobile introns or
inteins, that induce targeted recombination,
double-strand repair and gene conversion of their cognate target sites.
Due to their biological function and high level of target specificity,
these enzymes are under intense investigation as tools for gene
targeting. These studies require that naturally occurring enzymes be
redesigned to recognize novel target sites. Here, we report studies in
which the homodimeric LAGLIDADG homing endonuclease I-CreI is altered
at individual side-chains corresponding to contact points to distinct
base-pairs in its target site. The resulting enzyme constructs drive
specific elimination of selected DNA targets in vivo and display
shifted specificities of DNA binding and cleavage in vitro. Crystal
structures of two of these constructs demonstrate that substitution of
individual side-chain/DNA contact patterns can occur with almost no
structural deformation or rearrangement of the surrounding complex,
facilitating an isolated, modular redesign strategy for homing
endonuclease activity and specificity.

<>

<1>Sutcliffe, B., Midgley, D.J., Rosewarne, C.P., Greenfield, P., Li, D.
<2>Draft Genome Sequence of Thermotoga maritima A7A Reconstructed from Metagenomic Sequencing Analysis of a Hydrocarbon Reservoir in the Bass Strait, Australia.
<3>Genome Announcements
<4>1
<5>e00688-13
<6>2013
<7>The draft genome sequence of Thermotoga maritima A7A was obtained from a metagenomic assembly
obtained from a high-temperature hydrocarbon reservoir in
the Gippsland Basin, Australia. The organism is predicted to be a motile anaerobe
with an array of catabolic enzymes for the degradation of numerous carbohydrates.

<>

<1>Sutcliffe, J.G., Church, G.M.
<2>The cleavage site of the restriction endonuclease AvaII.
<3>Nucleic Acids Res.
<4>5
<5>2313-2319
<6>1978
<7>We have determined that the type II restriction enzyme AvaII, isolated from
Anabaena variabilis, recognizes and cuts the sequence 5' - G^GTCC - 3' 3' -
CCAG^G - 5' The eight AvaII sites of pBR322 have been mapped, as well as a
unique site for AvaI.

<>

<1>Sutherland, E., Coe, L., Raleigh, E.A.
<2>McrBC:  a multisubunit GTP-dependent restriction endonuclease.
<3>J. Mol. Biol.
<4>225
<5>327-358
<6>1992
<7>McrBC-mediated restriction of modified DNA has been studied extensively by genetic methods,
but little is known of its molecular action. We have used overproducing plasmid constructs to
facilitate purification of the McrB L and McrC proteins, and report preliminary
characterization of the activity of the complex. Both proteins are required for cleavage of
appropriately modified DNA in vitro, in a reaction absolutely dependent on GTP. ATP inhibits
the reaction. The sequence and modification requirements for cleavage of the substrate reflect
those seen in vivo. The position of cleavage was examined at the nucleotide level, revealing
that cleavage occurs at multiple positions in a small region. Based upon these observations,
and upon cleavage of model oligonucleotide substrates, it is proposed that the recognition
site for this enzyme consists of the motif RmC(N40-80) RmC, with cleavage occurring at
multiple positions on both strands between the modified C residues. In subunit composition,
cofactor requirement, and relation between cleavage and recognition site, McrBC does not fit
into any of the classes (types I to IV) of restriction enzyme so far described.

<>

<1>Suzuki, H., Lefebure, T., Hubisz, M.J., Pavinski, B., P, L., P, S., A, S., M, J.
<2>Comparative Genomic Analysis of the Streptococcus dysgalactiae Species Group: Gene Content, Molecular Adaptation, and Promoter Evolution.
<3>Genome Biol. Evol.
<4>3
<5>168-185
<6>2011
<7>Comparative genomics of closely related bacterial species with different
pathogenesis and host preference can provide a means of identifying the specifics
of adaptive differences. Streptococcus dysgalactiae (SD) is comprised of two
subspecies: S. dysgalactiae subsp. equisimilis is both a human commensal organism
and a human pathogen, and S. dysgalactiae subsp. dysgalactiae is strictly an
animal pathogen. Here, we present complete genome sequences for both taxa, with
analyses involving other species of Streptococcus but focusing on adaptation in
the SD species group. We found little evidence for enrichment in biochemical
categories of genes carried by each SD strain, however, differences in the
virulence gene repertoire were apparent. Some of the differences could be
ascribed to prophage and integrative conjugative elements. We identified
approximately 9% of the nonrecombinant core genome to be under positive
selection, some of which involved known virulence factors in other bacteria.
Analyses of proteomes by pooling data across genes, by biochemical category,
clade, or branch, provided evidence for increased rates of evolution in several
gene categories, as well as external branches of the tree. Promoters were
primarily evolving under purifying selection but with certain categories of genes
evolving faster. Many of these fast-evolving categories were the same as those
associated with rapid evolution in proteins. Overall, these results suggest that
adaptation to changing environments and new hosts in the SD species group has
involved the acquisition of key virulence genes along with selection of
orthologous protein-coding loci and operon promoters.

<>

<1>Suzuki, H., Takahashi, S., Osada, H., Yoshida, K.
<2>Improvement of transformation efficiency by strategic circumvention of restriction barriers in Streptomyces griseus.
<3>J. Microbiol. Biotechnol.
<4>21
<5>675-678
<6>2011
<7>DNA methylation in Streptomyces griseus IFO 13350 was analyzed by high-performance liquid
chromatographic analysis and bisulfite-based analysis to
reveal two methylation sites, 5'-GC5mCGGC-3' and 5'-GAG5mCTC-3'. The methylation
was reconstituted in Escherichia coli by simultaneous expression of S. griseus
SGR4675 and S. achromogenes M.SacI. The E. coli cells produced plasmids that
mimicked the methylation profile of S. griseus DNA, which was readily introduced
into S. griseus. The results of this study raise the possibility of a promising
approach to establish efficient transformation in several streptomycetes.

<>

<1>Suzuki, H., Yoshida, K.
<2>Genetic Transformation of Geobacillus kaustophilus HTA426 by Conjugative Transfer of Host-Mimicking Plasmids.
<3>J. Microbiol. Biotechnol.
<4>22
<5>1279-1287
<6>2012
<7>We established an efficient transformation method for thermophile Geobacillus kaustophilus
HTA426 using conjugative transfer from
Escherichia coli of host-mimicking plasmids that imitate DNA
methylation of strain HTA426 to circumvent its DNA restriction
barriers. Two conjugative plasmids, pSTE33T and pUCG18T, capable of
shuttling between E. coli and Geobacillus spp., were constructed. The
plasmids were first introduced into E. coli BR408, which expressed one
inherent DNA methylase gene (dam) and two heterologous methylase genes
from strain HTA426 (GK1380-GK1381 and GK0343-GK0344). The plasmids were
then directly transferred from E. coli cells to strain HTA426 by
conjugative transfer using pUB307 or pRK2013 as a helper plasmid.
pUCG18T was introduced very efficiently (transfer efficiency,
10(-5)-10(-3) recipient(-1)). pSTE33T showed lower efficiency
(10(-7)-10(-6) recipient(-1)) but had a high copy number and high
segregational stability. Methylase genes in the donor substantially
affected the transfer efficiency, demonstrating that the host-mimicking
strategy contributes to efficient transformation. The transformation
method, along with the two distinguishing plasmids, increases the
potential of G kaustophilus HTA426 as a thermophilic host to be used in
various applications and as a model for biological studies of this
genus. Our results also demonstrate that conjugative transfer is a
promising approach for introducing exogenous DNA into thermophiles.

<>

<1>Suzuki, K., Nagao, K., Tokunaga, J., Hirosawa, M., Tsubone, H., Uyeda, M.
<2>DMI-1, A new DNA methyltransferase inhibitor produced by Streptomyces sp. strain No. 560.
<3>J. Enzym. Inhib.
<4>9
<5>243-252
<6>1995
<7>A new inhibitor of DNA methyltransferase named DMI-1 has been discovered in
the culture filtrate of Streptomyces sp. strain No. 560.  DMI-1 was purified by extraction
with
ethyl acetate followed by Diaion HP-20SS and silica gel column chromatography.  The structure
of
DMI-1 was determined to be 8-methylpentadecanoic acid (C16H32O2).  DMI-1 is a novel inhibitor
of methyltransferase isolated from microorganisms and is structurally different from
sinefungin
and A9145C which are structural analogs of S-adenosylmethionine (methyl donor).  DMI-1 was a
strong inhibitor of N6-methyladenine-DNA methyltransferase (M.EcoRI, EC2.1.1.72) in a
noncompetitive manner and its inhibition depended on the pH and temperature in the assay
media.

<>

<1>Suzuki, K., Nagao, K., Tokunaga, J., Katayama, N., Uyeda, M.
<2>Inhibition of DNA methyltransferase by microbial inhibitors and fatty acids.
<3>J. Enzym. Inhib.
<4>10
<5>271-280
<6>1996
<7>Streptomyces sp. strain No. 560 produces four kinds of DNA methyltransferase inhibitors in the
culture filtrate.  One of them, DMI-4 was distinguished from DMI-1, -2 and -3 previously
reported with respect to certain properties.  DMI-4 is considered to be a triglyceride
consisting of the fatty acids anteisopentadecanoic acid (C15:0), isopalmitic acid (C16:0) and
isostearic acid (C18:0) from the results of gas chromatography analysis.  Since DMI-4 contains
three molecules of fatty acid, and the previously reported DMI-1, 8-methylpentadecanoic acid,
is analogous to a fatty acid, the inhibitory activity of various fatty acids and their methyl
esters has been examined against EcoRI DNA methyltransferase (M.EcoRI).  Oleic acid (C18:1)
was found to be a potent inhibitor of M.EcoRI.  The inhibitory activity of oleic acid was
shown to be pH- and temperature-dependent and inhibited M.EcoRI in a noncompetitive manner
with respect to DNA or S-adenosylmethionine (SAM).  The number of carbon atoms and double
bonds in the fatty acid molecule affected the inhibitory activity, but their methyl esters
were not inhibitors.  Our results suggest that the length of the carbon chain, the number of
double bonds and the presence of a carboxyl group and branched methyl group in the fatty acid
molecule may play an important role in the inhibition of DNA methyltransferase.

<>

<1>Suzuki, M., Matsui, M., Suzuki, S., Rimbara, E., Asai, S., Miyachi, H., Takata, T., Hiraki, Y., Kawano, F., Shibayama, K.
<2>Genome Sequences of Multidrug-Resistant Acinetobacter baumannii Strains from Nosocomial Outbreaks in Japan.
<3>Genome Announcements
<4>1
<5>e00476-13
<6>2013
<7>Acinetobacter baumannii has emerged worldwide as an important nosocomial pathogen in medical
institutions. Here, we present the draft genome sequences of A.
baumannii strains MRY09-0642, MRY10-0558, and MRY12-0277 that were isolated from
nosocomial outbreaks in Japan between 2008 and 2012 and that are resistant to
antimicrobial agents, including carbapenems, fluoroquinolones, and
aminoglycosides.

<>

<1>Suzuki, M., Suzuki, S., Matsui, M., Hiraki, Y., Kawano, F., Shibayama, K.
<2>Genome Sequence of a Strain of the Human Pathogenic Bacterium Pseudomonas alcaligenes That Caused Bloodstream Infection.
<3>Genome Announcements
<4>1
<5>e00919-13
<6>2013
<7>Pseudomonas alcaligenes, a Gram-negative aerobic bacterium, is a rare opportunistic human
pathogen. Here, we report the whole-genome sequence of P.
alcaligenes strain MRY13-0052, which was isolated from a bloodstream infection in
a medical institution in Japan and is resistant to antimicrobial agents,
including broad-spectrum cephalosporins and monobactams.

<>

<1>Suzuki, N., Okayama, S., Nonaka, H., Tsuge, Y., Inui, M., Yukawa, H.
<2>Large-scale engineering of the Corynebacterium glutamicum genome.
<3>Appl. Environ. Microbiol.
<4>71
<5>3369-3372
<6>2005
<7>The engineering of Corynebacterium glutamicum is important for enhanced production of
biochemicals. To construct an improved C. glutamicum genome,
we developed a precise genome excision method based on the Cre/loxP
recombination system and successfully deleted 11 distinct genomic regions
identified by comparative analysis of C. glutamicum genomes. Despite the
loss of several predicted open reading frames, the mutant cells exhibited
normal growth under standard laboratory conditions. With a total of 250 kb
(7.5% of the genome), the 11 genomic regions were loaded with cryptic
prophages, transposons, and genes of unknown function which were
dispensable for cell growth, indicating recent horizontal acquisitions to
the genome. This provides an interesting background for functional genomic
studies and can be used in the improvement of cell traits.

<>

<1>Suzuki, S., Aono, T., Lee, K.B., Suzuki, T., Liu, C.T., Miwa, H., Wakao, S., Iki, T., Oyaizu, H.
<2>Rhizobial factors required for stem nodule maturation and maintenance in Sesbania rostrata-Azorhizobium caulinodans ORS571 symbiosis.
<3>Appl. Environ. Microbiol.
<4>73
<5>6650-6659
<6>2007
<7>The molecular and physiological mechanisms behind the maturation and maintenance of
N(2)-fixing nodules during development of symbiosis between
rhizobia and legumes still remain unclear, although the early events of
symbiosis are relatively well understood. Azorhizobium caulinodans ORS571
is a microsymbiont of the tropical legume Sesbania rostrata, forming
N(2)-fixing nodules not only on the roots but also on the stems. In this
study, 10,080 transposon-inserted mutants of A. caulinodans ORS571 were
individually inoculated onto the stems of S. rostrata, and those mutants
that induced ineffective stem nodules, as displayed by halted development
at various stages, were selected. From repeated observations on stem
nodulation, 108 Tn5 mutants were selected and categorized into seven
nodulation types based on size and N(2) fixation activity. Tn5 insertions
of some mutants were found in the well-known nodulation, nitrogen
fixation, and symbiosis-related genes, such as nod, nif, and fix,
respectively, lipopolysaccharide synthesis-related genes, C(4)
metabolism-related genes, and so on. However, other genes have not been
reported to have roles in legume-rhizobium symbiosis. The list of newly
identified symbiosis-related genes will present clues to aid in
understanding the maturation and maintenance mechanisms of nodules.

<>

<1>Suzuki, S., Kuenen, J.G., Schipper, K., van der Velde, S., Ishii, S., Wu, A., Sorokin, D.Y., Tenney, A., Meng, X.Y., Morrill, P.L., Kamagata, Y., Muyzer, G., Nealson, K.H.
<2>Physiological and genomic features of highly-alkaliphilic hydrogen-utilizing Betaproteobacteria from a continental serpentinizing site.
<3>Nat. Commun.
<4>5
<5>3900
<6>2014
<7>Serpentinization, or the aqueous alteration of ultramafic rocks, results in challenging
environments for life in continental sites due to the combination of extremely high pH, low
salinity and lack of obvious electron acceptors and carbon sources.  Nevertheless, certain
Betaproteobacteria have been frequently observed in such environments.  Here we describe
physiological and genomic features of three related Betaprobacterial strains isolated from
highly alkaline (pH 11.6) serpentinizing springs at The Cedars, California.  All three strains
are obligate alkaliphiles with an optimum for growth at pH 11 and are capable of autotrophic
growth with hydrogen, calcium carbonate and oxygen.  The three strains exhibit differences,
however, regarding the utilization of organic carbon and electron acceptors.  Their global
distribution and physiological, genomic and transcriptomic characteristics indicate that the
strains are adapted to the alkaline and calcium-rich environments represented by the
terrestrial serpentinizing ecosystems.  We propose placing these strains in a new genus
'Serpentinomonas'.

<>

<1>Suzuki, T., Sato, Y., Yamada, Y., Akagawa-Matsushita, M., Yamasato, K.
<2>A restriction endonuclease (DpaI) from Deleya pacifica IAM 14115, a marine bacterium, an isoschizomer of ScaI.
<3>Agric. Biol. Chem.
<4>55
<5>2647-2649
<6>1991
<7>The marine bacteria have been considered to be important, but unused, uncommon
and unknown resources in the fields of applied and industrial microbiology.  In
previous papers, we reported a new restriction endonuclease, designated AgeI,
from Agrobacterium gelatinovorum Ahrens 1968 IAM 12617, a marine bacterium.
During the course of our screenings, we have found another marine bacterium to
produce a restriction endonuclease (DpaI, isoschizomer of ScaI) in a large
amount within the cells and the enzyme to have unique physiochemical
properties.

<>

<1>Suzuki, T., Sugimoto, E., Tahara, Y., Yamada, Y.
<2>Cloning and nucleotide sequence of the AgeI methylase gene from Agrobacterium gelatinovorum IAM 12617, a marine bacterium.
<3>Biosci. Biotechnol. Biochem.
<4>60
<5>444-447
<6>1996
<7>The AgeI restriction-modification system from a marine bacterium, Agrobacterium
gelatinovorum IAM 12617, recognizes the nucleotide sequence ACCGGT.  The gene coding for
the AgeI methylase (M.AgeI) was cloned into Escherichia coli DH5 alphaMCR, and the
nucleotides of the gene were sequenced.  The M.AgeI gene coded for a protein of 429 amino acid
residues (molecular mass, 47,358 daltons).  The deduced amino acid sequence of M.AgeI was
compared with those of other methylases and showed that there are high degrees of similarity
in
some cytosine-5-methylases.

<>

<1>Suzuki, T., Sugimoto, E., Tahara, Y., Yamada, Y.
<2>Cloning and nucleotide sequence of ApaLI restriction-modification system from Acetobacter pasteurianus IFO 13753.
<3>Biosci. Biotechnol. Biochem.
<4>60
<5>1401-1405
<6>1996
<7>The ApaLI restriction-modification system from Acetobacter pasteurianus IFO 13753 recognizes
the nucleotide sequence GTGCAC.  The gene coding for the ApaLI methylase (M.ApaLI) was cloned
into Escherichia coli DH5aMCR, and the nucleotide sequence of the gene was analyzed.  The
M.ApaLI gene coded for a protein of 429 amino acid residues (molecular mass, 46,554 daltons).
The ApaLI restriction endonuclease (R.ApaLI) gene was analyzed by inverse polymerase chain
reaction.  The R.ApaLI gene coded for a protein of 375 amino acid residues (molecular mass,
42,143 daltons).  The two genes had the same orientation separated by two base pairs.  The
deduced amino acid sequence of M.ApaLI shows significant similarities to the family of
cytosine-5 methylases.  However, the deduced amino acid sequence of R.ApaLI did not have as
much relatedness in the nucleotide sequence, when compared with those of the other restriction
endonucleases already reported.

<>

<1>Suzuki, T., Yasui, K.
<2>Plasmid Artificial Modification: A Novel Method for Efficient DNA Transfer into Bacteria.
<3>Methods Mol. Biol.
<4>765
<5>309-326
<6>2011
<7>Bacterial transformation is an essential component of many molecular biological techniques,
but bacterial restriction-modification (R-M)
systems can preclude the efficient introduction of shuttle vector
plasmids into target bacterial cells. Whole-genome DNA sequences have
recently been published for a variety of bacteria. Using homology and
motif analyses, putative R-M genes can be identified from genome
sequences. Introducing DNA methyltransferase genes into Escherichia
coli cells causes subsequently transformed plasmids to be modified by
these enzymes.We propose a new method, designated Plasmid Artificial
Modification (PAM). A PAM plasmid encoding the modification enzymes
expressed by the target bacterial host is transformed into E. coli (PAM
host). Propagation of a shuttle vector from the PAM host to the target
bacterium ensures that the plasmid will be modified such that it is
protected from restriction endonuclease digestion in the target
bacterium. The result will be a higher transformation efficiency.Here,
we describe the use of PAM and electroporation to transform
Bifidobacterium adolescentis ATCC15703. By introducing two genes
encoding modification enzymes, we improved transformation efficiency
10(5)-fold.

<>

<1>Suzuki, Y., Gilmore, J.L., Yoshimura, S.H., Henderson, R.M., Lyubchenko, Y.L., Takeyasu, K.
<2>Visual Analysis of Concerted Cleavage by Type IIF Restriction Enzyme SfiI in Subsecond Time Region.
<3>Biophys. J.
<4>101
<5>2992-2998
<6>2011
<7>Many DNA regulatory factors require communication between distantly separated DNA sites for
their activity.  The type IIF restriction enzyme SfiI is often used as a model system of site
communication.  Here, we used fast-scanning atomic force microscopy to monitor the DNA
cleavage process with SfiI and the changes in the single SfiI-DNA complex in the presence of
either Mg2+ or Ca2+ at a scan rate of 1-2 fps.  The increased time resolution allowed us to
visualize the concerted cleavage of the protein at two cognate sites.  The four termini
generated by the cleavage were released in a multistep manner.  The high temporal resolution
enabled us to visualize the translocation of a DNA strand on a looped complex and
intersegmental transfer of the SfiI protein in which swapping of the site is performed without
protein dissociation.  On the basis of our results, we propose that the SfiI tetramer can
remain bound to one of the sites even after cleavage, allowing the other site on the DNA
molecule to fill the empty DNA-binding cleft by combining a one-dimensional diffusion-mediated
sliding and a segment transfer mechanism.

<>

<1>Svab, D., Horvath, B., Szucs, A., Maroti, G., Toth, I.
<2>Draft Genome Sequence of an Escherichia coli O157:H43 Strain Isolated from Cattle.
<3>Genome Announcements
<4>1
<5>e00263-13
<6>2013
<7>Here we report the draft genome sequence of an Escherichia coli O157:H43 strain,  designated
T22, with an atypical virulence gene profile and isolated from healthy
cattle. T22 produces cytolethal distending toxin V (CDT-V) and belongs to
phylogenetic group B1 and sequence type 155 (ST155).

<>

<1>Svadbina, I.V., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>Location of the bases modified by M.BcoKIA and M.BcoKIB methylases in the sequence 5'-CTCTTC-3'/5'-GAAGAG-3'.
<3>Biochemistry
<4>70
<5>1363-1366
<6>2005
<7>The strain Bacillus coagulans K contains two DNA-methyltransferases, M.BcoKIA and M.BcoKIB,
which recognize the sequence 5'-CTCTTC-3'/5'-GAAGAG-3' and possess N4-methylcytosine and
N6-methyladenine specificities, respectively.  A special construct containing the recognition
site of BcoKI and sits of four IIS restriction endonucleases (IIS restriction endonuclease
cassette) was designed to locate the nucleotides modified by the methylases.  The modified
bases were determined as: 5'm4CTCTTC-3'/5'-GAAGAm6G-3'.

<>

<1>Svadbina, I.V., Zelinskaya, N.V., Kovalevskaya, N.P., Zheleznaya, L.A., Matvienko, N.I.
<2>Isolation and characterization of site-specific DNA- methyltransferases from Bacillus coagulans K.
<3>Biokhimiia
<4>69
<5>372-379
<6>2004
<7>Two site-specific DNA methyltransferases, M.BcoKIA and M.BcoKIB, were isolated from the
thermophilic strain Bacillus coagulans K. Each of the methylases protects the recognition site
5'-CTCTTC-3'/5'-GAAGAG-3' from cleavage with the cognate restriction endonuclease BcoKI.
It is shown that M.BcoKIB is an N6-adenine specific methylase and M.BcoKIA is an N4-cytosine
specific methylase. According to bisulfite mapping, M.BcoKIA methylates the first cytosine in
the sequence 5'-CTCTTC-3'.

<>

<1>Svadbina, I.V., Zheleznyakova, E.N., Zheleznaya, L.A., Matvienko, N.I.
<2>Bacillus species LU4 is an effective producer of thermostable site-specific endonuclease BspLU4I, an isoschizomer of AvaI.
<3>Biokhimiia
<4>68
<5>429-435
<6>2003
<7>The site-specific endonuclease BspLU4I was discovered in the thermophilic Bacillus species LU4
strain and purified to functionally pure state by
chromatography on blue agarose, hydroxyapatite HTP, and heparin-Sepharose
columns. Analysis of cleavage patterns of different DNAs with known
nucleotide sequences demonstrated that the enzyme recognizes the CPyCGPuG
site on the DNA. Cleavage points in the sequence were determined with the
elongated primer method. It was shown that the endonuclease is an
isoschizomer of AvaI. The final yield of the enzyme is 2.25.10(6) units
per g wet biomass.

<>

<1>Svedruzic, A.M., Reich, N.O.
<2>DNA cytosine C-5 methyltransferase Dnmt1: Catalysis-dependent release of allosteric inhibition.
<3>Biochemistry
<4>44
<5>9472-9485
<6>2005
<7>We followed the cytosine C-5 exchange reaction with Dnmt1 to characterize its preference for
different DNA substrates, its
allosteric regulation, and to provide a basis for comparison with the
bacterial enzymes. We determined that the methyl transfer is
rate-limiting, and steps up to and including the cysteine-cytosine
covalent intermediate are in rapid equilibrium. Changes in these rapid
equilibrium steps account for many of the previously described features
of Dnmt1 catalysis and specificity including faster reactions with
premethylated DNA versus unmethylated DNA, faster reactions with DNA in
which guanine is replaced with inosine [poly(dC-dG) vs poly(dI-dC)],
and 10-100-fold slower catalytic rates with Dnmt1 relative to the
bacterial enzyme M.HhaI. Dnmt1 interactions with the guanine within the
CpG recognition site can prevent the premature release of the target
base and solvent access to the active site that could lead to mutagenic
deamination. Our results suggest that the beta-elimination step
following methyl transfer is not mediated by free solvent. Dnmt1 shows
a kinetic lag in product formation and allosteric inhibition with
unmethylated DNA that is not observed with premethylated DNA. Thus, we
suggest the enzyme undergoes a slow relief from allosteric inhibition
upon initiation of catalysis on unmethylated DNA. Notably, this relief
from allosteric inhibition is not caused by self-activation through the
initial methylation reaction, as the same effect is observed during the
cytosine C5 exchange reaction in the absence of AdoMet. We describe
limitations in the Michaelis-Menten kinetic analysis of Dnmt1 and
suggest alternative approaches.

<>

<1>Svedruzic, Z.M.
<2>Mammalian cytosine DNA methyltransferase Dnmt1: Enzymatic mechanism, novel mechanism-based inhibitors, and RNA-directed DNA methylation.
<3>Curr. Med. Chem.
<4>15
<5>92-106
<6>2008
<7>This is a review of the enzymatic mechanism of DNA methyltransferase Dnmt1 and analysis of its
implications on regulation of DNA methylation
in mammalian cells and design of novel mechanism-based inhibitors. The
methylation reaction by Dnmt1 has different phases that depend on DNA
substrate and allosteric regulation. Consequently, depending on the
phase, the differences in catalytic rates between unmethylated and
pre-methylated DNA can vary between 30-40 fold, 3-6 fold or only 1
fold. The allosteric site and the active site can bind different
molecules. Allosteric activity depends on DNA sequence, methylation
pattern and DNA structure (single stranded vs. double stranded). Dnmt1
binds poly(ADP-ribose) and some RNA molecules. The results on kinetic
preferences, allosteric activity and binding preference of Dnmt1 are
combined together in one comprehensive model mechanism that can address
regulation of DNA methylation in cells; namely, inhibition of DNA
methylation by poly(ADP-ribose), RNA-directed DNA methylation by
methylated and unmethylated non-coding RNA molecules, and transient
interactions between Dnmt1 and genomic DNA. Analysis of reaction
intermediates showed that equilibrium between base-flipping and
base-restacking events can be the key mechanism in control of enzymatic
activity. The two events have equal but opposite effect on accumulation
of early reaction intermediates and methylation rates. The accumulation
of early reaction intermediates can be exploited to improve the current
inhibitors of Dnmt1 and achieve inhibition without toxic modifications
in genomic DNA.
[1,2-dihydropyrimidin-2-one]-5-methylene-(methylsulfonium)-adenosyl is
described as the lead compound.

<>

<1>Svedruzic, Z.M., Reich, N.O.
<2>Enzymatic properties of mouse cytosine DNA methyltransferase DNMT1.
<3>ACS Abstracts
<4>223
<5>C75
<6>2002
<7>DNA methylation is a gene control and chromatin organization mechanism that has been
associated with oncogene activation, mutagenic cytosine deamination, cell cycle changes,
retrovirus silencing, cell differentiation, and aging. This work evaluates the main catalytic
features of Dnmt1, the principal methyltransferase in eukaryotic cells.  Pre-steady state and
steady state kinetics were used together with solvent kinetic isotope effect and pH studies to
analyze catalytic preference for pre-methylated DNA, enzyme processivity on its DNA substrate,
allosteric regulation and likelihood of mutagenic deamination. The preference for
pre-methylated DNA is due to a faster target base attack, faster methyl transfer and faster
acid base catalysis that is likely to come from a specific conformational change in the active
site. Unlike its bacterial counterparts, Dnmt1 can recognize DNA as pre-methylated even when
5mC is one out of fifteen cytosines, however the enzyme is not self activated early in the
methylation reaction on unmethylated DNA. A novel analysis of processivity rate constants
showed that the processivity of Dnmt1 depends on the substrate and can be regulated through
the allosteric site. The extent of allosteric regulation depends on catalytic activity, DNA
sequence and the methylation status. DNA binding at the allosteric site inhibits catalytic
action of Dnmt1 and initiates DNA release from its active site. Interaction between Dnmt1 and
proliferating cell nuclear antigen (PCNA) facilitates the on rate for the substrate DNA, and
has no effect on enzyme processivity. Analogues of S-Adenosylmethionine and solvent kinetic
isotope effect studies showed that enzyme mediated mutagenesis and cytosine deamination are
due to enzymes inability to protect its reactive intermediates. Mutagenesis is likely in
catalytic action by mammalian and bacterial cytosine methyltransferases, however mammalian
enzyme is 200 times less likely to be mutagenic in the absence of the cofactor. Presented
studies give enzymatic insights to biology of DNA methylation and show novel methods to study
C5 pyrimidine methyltransferases.

<>

<1>Svedruzic, Z.M., Reich, N.O.
<2>Mechanism of allosteric regulation of Dnmt1's processivity.
<3>Biochemistry
<4>44
<5>14977-14988
<6>2005
<7>We have analyzed the relationship between the allosteric regulation and processive catalysis
of DNA methyltransferase 1 (Dnmt1). Processivity is
described quantitatively in terms of turnover rate, DNA dissociation rate,
and processivity probability. Our results provide further evidence that
the active site and the allosteric sites on Dnmt1 can bind DNA
independently. Dnmt1's processive catalysis on unmethylated DNA is
partially inhibited when the allosteric site binds unmethylated DNA and
fully inhibited when the allosteric site binds a single-stranded
oligonucleotide inhibitor. The partial inhibition by unmethylated DNA is
caused by a decrease in the turnover rate and an increase in the substrate
DNA dissociation rate. Processive catalysis with premethylated DNA is not
affected if the allosteric site is exposed to premethylated DNA but is
fully inhibited if the allosteric site binds unmethylated DNA or
poly(dA-dT). In sum, the occupancy of the allosteric site modulates the
enzyme's commitment to catalysis, which reflects the nature of the
substrate and the DNA bound at the allosteric site. Our in vitro results
are consistent with the possibility that the processive action of Dnmt1
may be regulated in vivo by specific regulatory nucleic acids such as DNA,
RNA, or poly(ADP-ribose).

<>

<1>Svenning, M.M. et al.
<2>Genome Sequence of the Arctic Methanotroph Methylobacter tundripaludum SV96.
<3>J. Bacteriol.
<4>193
<5>6418-6419
<6>2011
<7>Methylobacter tundripaludum SV96(T) (ATCC BAA-1195) is a psychrotolerant aerobic
methane-oxidizing gammaproteobacterium (Methylococcales,
Methylococcaceae) living in High Arctic wetland soil. The strain was
isolated from soil harvested in July 1996 close to the settlement
Ny-Alesund, Svalbard, Norway (78 degrees 56'N, 11 degrees 53'E), and
described as a novel species in 2006. The genome includes pmo and pxm
operons encoding copper membrane monooxygenases (Cu-MMOs), genes required
for nitrogen fixation, and the nirS gene implicated in dissimilatory
nitrite reduction to NO but no identifiable inventory for further
processing of nitrogen oxides. These genome data provide the basis to
investigate M. tundripaludum SV96, identified as a major player in the
biogeochemistry of Arctic environments.

<>

<1>Swaminathan, C.P., Sankpal, U.T., Rao, D.N., Surolia, A.
<2>Water-assisted dual mode cofactor recognition by HhaI DNA methyltransferase.
<3>J. Biol. Chem.
<4>277
<5>4042-4049
<6>2002
<7>Energetically competent binary recognition of the cofactor S-adenosyl-L-methionine (AdoMet)
and the product S-adenosyl-L-homocysteine (AdoHcy) by the DNA (cytosine C-5) methyltransferase
(M.HhaI) is demonstrated herein. Titration calorimetry reveals a dual mode, involving a
primary dominant exothermic reaction followed by a weaker endothermic one, for the recognition
of AdoMet and AdoHcy by M.HhaI. Conservation of the bimodal recognition in W41I and W41Y
mutants of M.HhaI excludes the cation-Pi interaction between the methylsulfonium group of
AdoMet and the Pi face of the Trp(41) indole ring from a role in its origin. Small magnitude
of temperature-independent heat capacity changes upon AdoMet or AdoHcy binding by M.HhaI
preclude appreciable conformational alterations in the reacting species. Coupled
osmotic-calorimetric analyses of AdoMet and AdoHcy binding by M.HhaI indicate that a net
uptake of nearly eight and 10 water molecules, respectively, assists their primary
recognition. A change in water activity at constant temperature and pH is sufficient to
engender and conserve enthalpy-entropy compensation, consistent with a true osmotic effect.
The results implicate solvent reorganization in providing the major contribution to the origin
of this isoequilibrium phenomenon in AdoMet and AdoHcy recognition by M.HhaI. The observations
provide unequivocal evidence for the binding of AdoMet as well as AdoHcy to M.HhaI in solution
state. Isotope partitioning analysis and preincubation studies favor a random mechanism for
M.HhaI-catalyzed reaction. Taken together, the results clearly resolve the issue of cofactor
recognition by free M.HhaI, specifically in the absence of DNA, leading to the formation of an
energetically and catalytically competent binary complex.

<>

<1>Swaminathan, N., George, D., McMaster, K., Szablewski, J., Van Etten, J.L., Mead, D.A.
<2>Restriction generated oligonucleotides utilizing the two base recognition endonuclease CviJI.
<3>Nucleic Acids Res.
<4>22
<5>1470-1475
<6>1994
<7>The conversion of an anonymous DNA sample into numerous oligonucleotides is enzymatically
feasible using an unusual restriction endonuclease, CviJI. Depending on reaction conditions,
CviJI is capable of digesting DNA at a two or three base recognition sequence. CviJI normally
cleaves RGCY sites between the G and C to leave blunt ends. Under 'relaxed' conditions CviJI
cleaves RGCY, and RGCR/YGCY, but not YGCR sites. In theory, CviJI* restriction of pUC19 (2686
bp) should produce 157 fragments, 75% of which are smaller than 20 bp. Instead, 96% of the
CviJI* fragments were 18-56 bp long and none of the fragments generates sequence specific
oligonucleotides homologous for the cognate template. The enzymatic conversion of anonymous
DNA into sequence specific oligomers has implications for several conventional and novel
molecular biology procedures.

<>

<1>Swaminathan, N., Mead, D.A., McMaster, K., George, D., Van Etten, J.L., Skowron, P.M.
<2>Molecular cloning of the three base restriction endonuclease R.CviJI from eukaryotic Chlorella virus IL-3A.
<3>Nucleic Acids Res.
<4>24
<5>2463-2469
<6>1996
<7>R.CviJI is unique among site-specific restriction endonucleases in that its activity can be
modulated to recognize either a two or three base sequence.  Normally R.CviJI cleaves RGCY
sites between the G and C to leave blunt ends.  In the presence of ATP, R.CviJI cleaves RGCN
and YGCY sites, but not YGCR sites.  The gene encoding R.CviJI was cloned from the eukaryotic
Chlorella virus IL-3A and expressed in Escherichia coli.  The primary E. coli cviJIR gene
product is a 278 amino acid protein initiated from a GTG codon, rather than the expected 358
amino acid protein, displays the normal restriction activity but not the R.CviJI* activity of
the native enzyme.  Nine restriction and modification proteins which recognize a central GC or
CG sequence share short regions of identity with R.CviJI amino acids 144-235, suggesting that
this region is the recognition and/or catalytic domain.

<>

<1>Swanson, R.V., Feldman, R.A., Schleper, C.
<2>Nucleic acids and proteins from Cenarchaeum symbiosum.
<3>Japanese Patent Office
<4>JP 2002531060 A
<5>
<6>2002
<7>
<>

<1>Swanson, R.V., Feldman, R.A., Schleper, C.
<2>Nucleic acids and proteins from Cenarchaeum symbiosum.
<3>US Patent Office
<4>US 6632937 B
<5>
<6>2003
<7>The present application relates to nucleic acids and polypeptides from Cenarchaeum symbiosum.
Methods of making the polypeptides and antibodies against the polypeptides are also described.

<>

<1>Swarbreck, D., Wilks, C., Lamesch, P., Berardini, T.Z., Garcia-Hernandez, M., Foerster, H., Li, D., Meyer, T., Muller, R., Ploetz, L., Radenbaugh, A., Singh, S., Swing, V., Tissier, C., Zhang, P., Huala, E.
<2>The Arabidopsis Information Resource (TAIR): gene structure and function annotation.
<3>Nucleic Acids Res.
<4>36
<5>D1009-D1014
<6>2008
<7>The Arabidopsis Information Resource (TAIR, http://arabidopsis.org) is the model organism
database for the fully sequenced and intensively studied
model plant Arabidopsis thaliana. Data in TAIR is derived in large part
from manual curation of the Arabidopsis research literature and direct
submissions from the research community. New developments at TAIR include
the addition of the GBrowse genome viewer to the TAIR site, a redesigned
home page, navigation structure and portal pages to make the site more
intuitive and easier to use, the launch of several TAIR web services and a
new genome annotation release (TAIR7) in April 2007. A combination of
manual and computational methods were used to generate this release, which
contains 27,029 protein-coding genes, 3889 pseudogenes or transposable
elements and 1123 ncRNAs (32,041 genes in all, 37,019 gene models). A
total of 681 new genes and 1002 new splice variants were added. Overall,
10,098 loci (one-third of all loci from the previous TAIR6 release) were
updated for the TAIR7 release.

<>

<1>Swarnkar, M.K., Salwan, R., Kasana, R.C., Singh, A.K.
<2>Draft Genome Sequence of Psychrotrophic Acinetobacter sp. Strain MN12 (MTCC 10786), Which Produces a Low-Temperature-Active and Alkaline-Stable Peptidase.
<3>Genome Announcements
<4>2
<5>e01167-14
<6>2014
<7>We report here the draft genome sequence of Acinetobacter sp. strain MN12 (MTCC 10786), which
is a psychrotrophic bacterium that produces an extracellular
low-temperature-active and alkaline-stable peptidase. The draft genome assembly
of Acinetobacter sp. MN12 has a size of 4.31 Mbp, with a G+C content of 40.75%.

<>

<1>Sweetnam, K.R., Reich, N.O.
<2>Construction of a mechanism-based inhibitor for EcoRI DNA methyltransferase.
<3>Biochemistry
<4>31
<5>2206
<6>1992
<7>Our results using N-ethylmaleimide modification suggest that cysteine 223 is
critical for the EcoRI DNA methyltransferase catalyzed methylation of the
second adenine in the GAATTC recognition sequence.  A plausible catalytic
mechanism involving this cysteine is shown below: A key feature is the
formation of a methylase-DNA covalent intermediate.  We are testing this
mechanism by incorporating a 6-chloropurine riboside into the DNA substrate.
This substituted analogue is proposed to act as a mechanism-based or active
site directed inactivator due to the excellent leaving group potential of the
6-chloro moiety.  We are submitting this analogue to inactivation analysis and
isolating the peptide sequence attached to the DNA to determine the reactive
residue in the catalytic site of EcoRI DNA methyltransferase.

<>

<1>Sweetnam, K.R., Reich, N.O.
<2>Construction of a mechanism-based inhibitor for EcoRI DNA methyltransferase.
<3>ACS Abstracts
<4>203
<5>102-BIOL
<6>1992
<7>Our results using N-ethyl maleimide modification suggest that cysteine 223 is critical for the
EcoRI DNA methyltransferase catalyzed methylation of the second adenine in the GAATTC
recognition sequence. A plausible catalytic mechanism involving this cysteine is shown below:
a key feature is the formation of a methylase-DNA covalent intermediate. We are testing this
mechanism by incorporating a 6-chloropurine riboside into the DNA substrate. This substituted
analog is proposed to act as a mechanism based or active site directed inactivator due to the
excellent leaving group potential of the 6-chloro moiety. We are submitting this analog to
inactivation analysis and isolating the peptide sequence attached to the DNA to determine the
reactive residue in the catalytic site of EcoRI DNA methyltransferase.

<>

<1>Swingley, W.D. et al.
<2>Niche adaptation and genome expansion in the chlorophyll d-producing cyanobacterium Acaryochloris marina.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>2005-2010
<6>2008
<7>Acaryochloris marina is a unique cyanobacterium that is able to produce chlorophyll d as its
primary photosynthetic pigment and thus efficiently
use far-red light for photosynthesis. Acaryochloris species have been
isolated from marine environments in association with other oxygenic
phototrophs, which may have driven the niche-filling introduction of
chlorophyll d. To investigate these unique adaptations, we have sequenced
the complete genome of A. marina. The DNA content of A. marina is composed
of 8.3 million base pairs, which is among the largest bacterial genomes
sequenced thus far. This large array of genomic data is distributed into
nine single-copy plasmids that code for >25% of the putative ORFs. Heavy
duplication of genes related to DNA repair and recombination (primarily
recA) and transposable elements could account for genetic mobility and
genome expansion. We discuss points of interest for the biosynthesis of
the unusual pigments chlorophyll d and alpha-carotene and genes
responsible for previously studied phycobilin aggregates. Our analysis
also reveals that A. marina carries a unique complement of genes for these
phycobiliproteins in relation to those coding for antenna proteins related
to those in Prochlorococcus species. The global replacement of major
photosynthetic pigments appears to have incurred only minimal
specializations in reaction center proteins to accommodate these alternate
pigments. These features clearly show that the genus Acaryochloris is a
fitting candidate for understanding genome expansion, gene acquisition,
ecological adaptation, and photosystem modification in the cyanobacteria.

<>

<1>Swingley, W.D., Sadekar, S., Mastrian, S.D., Matthies, H.J., Hao, J., Ramos, H., Acharya, C.R., Conrad, A.L., Taylor, H.L., Dejesa, L.C., Shah, M.K., O'huallachain, M.E., Lince, M.T., Blankenship, R.E., Beatty, J.T., Touchman, J.W.
<2>The complete genome sequence of Roseobacter denitrificans reveals a mixotrophic rather than photosynthetic metabolism.
<3>J. Bacteriol.
<4>189
<5>683-690
<6>2007
<7>Purple aerobic anoxygenic phototrophs (AAPs) are the only organisms known to capture light
energy to enhance growth only in the presence of oxygen
but do not produce oxygen. The highly adaptive AAPs compose more than 10%
of the microbial community in some euphotic upper ocean waters and are
potentially major contributors to the fixation of the greenhouse gas CO2.
We present the complete genomic sequence and feature analysis of the AAP
Roseobacter denitrificans, which reveal clues to its physiology. The
genome lacks genes that code for known photosynthetic carbon fixation
pathways, and most notably missing are genes for the Calvin cycle enzymes
ribulose bisphosphate carboxylase (RuBisCO) and phosphoribulokinase.
Phylogenetic evidence implies that this absence could be due to a gene
loss from a RuBisCO-containing alpha-proteobacterial ancestor. We describe
the potential importance of mixotrophic rather than autotrophic CO2
fixation pathways in these organisms and suggest that these pathways
function to fix CO2 for the formation of cellular components but do not
permit autotrophic growth. While some genes that code for the
redox-dependent regulation of photosynthetic machinery are present, many
light sensors and transcriptional regulatory motifs found in purple
photosynthetic bacteria are absent.

<>

<1>Swinton, D., Hattman, S., Benzinger, R., Buchanan-Wollaston, V., Beringer, J.
<2>Replacement of the deoxycytidine residues in Rhizobium bacteriophage RL38JI DNA.
<3>FEBS Lett.
<4>184
<5>294-298
<6>1985
<7>Rhizobium phage RL38JI DNA is resistant to cleavage by a variety of restriction
endonucleases, and is only partially sensitive to digestion by pancreatic DNase
I or by micrococcal nuclease.  We have found that a mixture of DNase I, P1
nuclease, and bacterial alkaline phosphatase will quantitatively digest RL38JI
DNA to deoxyribonucleosides.  HPLC analysis revealed that dCyd is nearly
totally absent among these digestion products, while dGuo, dAdo, and Thd are
readily detected.  Three additional peaks are always present; their retention
properties correspond to no known modified deoxyribonucleosides.  Thus it
appears that dCyd is replaced in phage RL38JI DNA by as many as 3 different
modified residues.

<>

<1>Swinton, D., Hattman, S., Crain, P.F., Cheng, C.-S., Smith, D.L., McCloskey, J.A.
<2>Purification and characterization of the unusual deoxynucleoside, alpha-N-(9-beta-D-2'-deoxyribofuranosylpurin-t-yl) glycinamide, specified by the phage Mu modification function.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>80
<5>7400-7404
<6>1983
<7>Bacteriophage Mu encodes a protein that modifies ~15% of DNA adenine residues
to a new and unusual form.  Modified DNA was enzymatically digested to
deoxynucleosides, and the products were fractionated by HPLC.  A modified
adenine nucleoside, designated dA'/x, was purified and its molecular structure
was established by mass spectrometry.  We show that dA'/x is
alpha-N-(9-beta-D-2'-deoxyribofuranosylpurin-t-yl)-glycinamide.  The dA'/x
obtained from DNA was indistinguishable from the synthetic product with respect
to its chromatographic behavior (HPLC and gas chromatography) and mass
spectrum.  Acid hydrolysis degrades dA'/x to produce N6-carboxymethyladenine;
this compound corresponds to the base Ax observed in earlier studies.

<>

<1>Swithers, K.S. et al.
<2>Genome Sequence of Thermotoga sp. Strain RQ2, a Hyperthermophilic Bacterium Isolated from a Geothermally Heated Region of the Seafloor near  Ribeira Quente, the Azores.
<3>J. Bacteriol.
<4>193
<5>5869-5870
<6>2011
<7>Thermotoga sp. strain RQ2 is probably a strain of Thermotoga maritima. Its complete genome
sequence allows for an examination of the extent and
consequences of gene flow within Thermotoga species and strains.
Thermotoga sp. RQ2 differs from T. maritima in its genes involved in
myo-inositol metabolism. Its genome also encodes an apparent fructose
phosphotransferase system (PTS) sugar transporter. This operon is also
found in Thermotoga naphthophila strain RKU-10 but no other Thermotogales.
These are the first reported PTS transporters in the Thermotogales.

<>

<1>Swithers, K.S., Dipippo, J.L., Bruce, D.C., Detter, C., Tapia, R., Han, S., Goodwin, L.A., Han, J., Woyke, T., Pitluck, S., Pennacchio, L., Nolan, M., Mikhailova, N., Land, M.L., Nesbo, C.L., Gogarten, J.P., Noll, K.M.
<2>Genome Sequence of Kosmotoga olearia Strain TBF 19.5.1, a Thermophilic Bacterium with a Wide Growth Temperature Range, Isolated from the Troll B  Oil Platform in the North Sea.
<3>J. Bacteriol.
<4>193
<5>5566-5567
<6>2011
<7>Kosmotoga olearia strain TBF 19.5.1 is a member of the Thermotogales that grows best at 65
degrees C and very well even at 37 degrees C. Information
about this organism is important for understanding the evolution of
mesophiles from thermophiles. Its genome sequence reveals extensive gene
gains and a large content of mobile genetic elements. It also contains
putative hydrogenase genes that have no homologs in the other member of
the Thermotogales.

<>

<1>Switt, A.I., A, I., Orsi, R.H., den Bakker, H.C., Vongkamjan, K., Altier, C., Wiedmann, M.
<2>Genomic characterization provides new insight into Salmonella phage diversity.
<3>BMC Genomics
<4>14
<5>481
<6>2013
<7>Background
Salmonella is a widely distributed foodborne pathogen that causes tens of millions of
salmonellosis cases globally every year. While the genomic diversity of Salmonella is
increasingly well studied, our knowledge of Salmonella phage genomic diversity is still rather
limited, despite the contributions of both lysogenic and lytic phages to Salmonella
virulence, diversity and ecology (e.g., through horizontal gene transfer and Salmonella
lysis). To gain a better understanding of phage diversity in a specific ecological niche, we
sequenced 22 Salmonella phages isolated from a number of dairy farms from New York State
(United States) and analyzed them using a comparative genomics approach.
Results
Classification of the 22 phages according to the presence/absence of orthologous genes allowed
for classification into 8 well supported clusters. In addition to two phage clusters that
represent novel virulent Salmonella phages, we also identified four phage clusters that each
contained previously characterized phages from multiple continents. Our analyses also
identified two clusters of phages that carry putative virulence (e.g., adhesins) and
antimicrobial resistance (tellurite and bicyclomycin) genes as well as virulent and temperate
transducing phages. Insights into phage evolution from our analyses include (i) identification
of DNA metabolism genes that may facilitate nucleotide synthesis in phages with a G+C %
distinct from Salmonella, and (ii) evidence of Salmonella phage tailspike and fiber diversity
due to both single nucleotide polymorphisms and major re-arrangements, which may affect the
host specificity of Salmonella phages.
Conclusions Genomics-based characterization of 22 Salmonella phages isolated from dairy farms
allowed for identification of a number of novel Salmonella phages. While the comparative
genomics analyses of these phages provide a number of new insights in the evolution and
diversity of Salmonella phages, they only represent a first glimpse into the diversity of
Salmonella phages
that is likely to be discovered when phages from different environments are characterized.

<>

<1>Syddall, R., Stachow, C.
<2>A site-specific endonuclease from Caulobacter crescentus CB-13: restriction endonuclease CcrI.
<3>Biochim. Biophys. Acta
<4>825
<5>236-243
<6>1985
<7>A site-specific restriction endonuclease (CcrI) has been identified from
Caulobacter crescentus CB-13.  This enzyme has been purified to homogeneity and
the cleavage patterns with various DNAs and sequence data show that CcrI
recognizes the same sequence as the XhoI restriction endonuclease
(5'-C^T-C-G-A-G-3').  CcrI has an absolute requirement for magnesium ions with
an optimum concentration of 4 mM.  The enzyme is optimally active at pH 8.0 and
is stable up to 70C.  CcrI has a molecular weight of 65300 and exists as a
monomer in its native state.  Most of the physical characteristics observed for
CcrI were similar to those observed for XhoI.  Kinetic studies on CcrI and XhoI
suggest that the enzymes with lambda DNA in the same manner, however with
PhiX-174 RF DNA, CcrI has a greater affinity for the supercoiled molecule than
XhoI.

<>

<1>Sykilinda, N.N., Bondar, A.A., Gorshkova, A.S., Kurochkina, L.P., Kulikov, E.E., Shneider, M.M., Kadykov, V.A., Solovjeva, N.V., Kabilov, M.R., Mesyanzhinov, V.V., Vlassov, V.V., Drukker, V.V., Miroshnikov, K.A.
<2>Complete Genome Sequence of the Novel Giant Pseudomonas Phage PaBG.
<3>Genome Announcements
<4>2
<5>e00929-13
<6>2014
<7>The novel giant Pseudomonas aeruginosa bacteriophage PaBG was isolated from a water sample of
the ultrafreshwater Lake Baikal. We report the complete genome
sequence of this Myoviridae bacteriophage, comprising 258,139 bp of
double-stranded DNA containing 308 predicted open reading frames.

<>

<1>Szatmari, G., Hua, N.M., Vzdornov, D., Daigle, F., Smoragiewicz, W., Mamet-Bratley, M.D., Karska-Wysocki, B.
<2>In vitro expression of the restriction endonucleases LlaMI and ScrFI isolated from Lactococcus lactis M19 and UC503.
<3>J. Biotechnol.
<4>121
<5>144-153
<6>2006
<7>A new restriction endonuclease LlaMI has been characterized in Lactococcus lactis subsp.
cremoris M19. LlaMI recognizes the sequence 5'-CCNGG-3' and cuts after the second cytosine.
This restriction endonuclease is related to commercially available ScrFI but not identical to
it. Comparative analysis of the predicted amino acid sequences of LlaMI and ScrFI indicates
five non-conservative amino acid changes between these two restriction enzymes. These two
enzymes were expressed in vitro as histidine-tagged fusion proteins. LlaMI was shown to be
more sensitive to high salt concentration than ScrFI. Southern blotting and hybridization
analysis indicate that the gene for LlaMI R/M system is chromosomally encoded.

<>

<1>Szczelkun, M.D.
<2>How do proteins move along DNA?  Lessons from type-I and type-III restriction endonucleases.
<3>Essays Biochem., Portland Press, Banting, G., Higgins, S.J., London
<4>35
<5>131-143
<6>2000
<7>A fundamental requirement of nearly every genetic event, including DNA replication,
transcription, cleavage and repair, is the protein-mediated communication between distance DNA
sites.  Four general mechanisms for this 'action at a distance' were described by Adzuma and
Mizuuchi and are illustrated in Figure 1.  The first three models, DNA sliding, DNA hopping
and DNA looping, are driven passively by random thermal fluctuations of the protein or DNA,
with significant consequences for the efficiency of the reactions.  In DNA sliding, a protein
bound to non-specific DNA diffuses either leftwards or rightwards without dissociating,
altering the DNA-binding register by 1 bp.  By confining diffusion to one dimension, the
probability of finding a second site or protein is increased.  However, the number of physical
steps needed to move between two DNA loci by linear diffusion has a square power dependence on
the distance between those sites; for instance, travelling 1000 bp takes 1 x 10^6 steps, which
in turn requires a linear diffusion rate at least 1 x 10^6 times faster than the DNA
dissociation rate.  Generally, DNA-binding proteins associate transiently with non-specific
sequences and DNA sliding would only be feasible over quite short distances, less than 100 bp.
In DNA hopping, a protein physically releases DNA, diffuses in three dimensions and randomly
rebinds the same DNA molecule at an alternative site.  The DNA acts as a binding 'reservoir'
- given enough time, one protein could sample every available binding locus.
Three-dimensional diffusion on a random coil has a square-root power dependence on the
distance between target sites, but because sampling is non-linear, sites may be overlooked and
the protein could even diffuse away from the DNA altogether.  In DNA looping, a protein
remains bound to a specific DNA sequence whilst segmental diffusion of the polynucleotide
brings distant DNA sites (and any bound proteins) into close proximity.  In general, the
tethering of two proteins on to the same DNA molecule increases the probability of
protein-protein interactions.  However, the probability of juxtaposition decreases as the
distance between the sites increases, an effect that is particularly marked on linear DNA.
DNA-looping events are less likely over distances greater than ~1000 bp without the
cooperation of accessory factors, such as DNA-bending proteins.

<>

<1>Szczelkun, M.D.
<2>Kinetic models of translocation, head-on collision, and DNA cleavage by type I restriction endonucleases.
<3>Biochemistry
<4>41
<5>2067-2074
<6>2002
<7>Digestion of linear DNA by type I restriction endonucleases is generally activated following
the head-on collision of two
translocating enzymes. However, the resulting distributions of cleavage
loci along the DNA vary with different enzymes; in some cases, cleavage
is located in a discrete region midway between a pair of recognition
sites while in other cases cleavage is broadly distributed and occurs
at nearly every intervening locus. Statistical models for DNA
translocation, collision, and cleavage are described that can account
for these observations and that are generally applicable to other
DNA-based motor proteins. If translocation is processive (stepping
forward is significantly more likely than DNA dissociation), then the
linear distribution of an ensemble of proteins can be described simply
using a Poisson relationship. The pattern of cleavage sites resulting
from collision between two processive type I enzymes over a distance d
can then be described by a binomial distribution with a standard
deviation 0.5.d(1/2). Alternatively, if translocation is nonprocessive
(stepping forward or dissociating become equally likely events), the
linear distribution is described by a continuum of populated states and
is thus extended. Comparisons of model data to the kinetics of DNA
translocation and cleavage discount the nonprocessive model. Instead,
the observed differences between enzymes are due to asynchronous events
that occur upon collision. Therefore, type I restriction enzymes can be
described as having processive DNA translocation but, in some cases,
nonprocessive DNA cleavage.

<>

<1>Szczelkun, M.D.
<2>Translocation, switching and gating: potential roles for ATP in long-range communication on DNA by Type III restriction endonucleases.
<3>Biochem. Soc. Trans.
<4>39
<5>589-594
<6>2011
<7>To cleave DNA, the Type III RM (restriction-modification) enzymes must communicate the
relative orientation of two recognition sequences, which may be separated by many thousands of
base pairs. This long-range interaction requires ATP hydrolysis by a helicase domain, and both
active (DNA translocation) and passive (DNA sliding) modes of motion along DNA have been
proposed. Potential roles for ATP binding and hydrolysis by the helicase domains are
discussed, with a focus on bipartite ATPases that act as molecular switches.

<>

<1>Szczelkun, M.D.
<2>Roles for Helicases as ATP-Dependent Molecular Switches.
<3>Adv. Exp. Med. Biol.
<4>767
<5>225-244
<6>2013
<7>On the basis of the familial name, a 'helicase' might be expected to have an enzymatic
activity that unwinds duplex polynucleotides to form single strands. A more encompassing
taxonomy that captures alternative enzymatic roles has defined helicases as a sub-class of
molecular motors that move directionally and processively along nucleic acids, the so-called
'translocases'. However, even this definition may be limiting in capturing the full scope of
helicase mechanism and activity. Discussed here is another, alternative view of helicases-as
machines which couple NTP-binding and hydrolysis to changes in protein conformation to resolve
stable nucleoprotein assembly states. This 'molecular switch' role differs from the
classical view of helicases as molecular motors in that only a single catalytic NTPase cycle
may be involved. This is illustrated using results obtained with the DEAD-box family of RNA
helicases and with a model bacterial system, the ATP-dependent Type III
restriction-modification enzymes. Further examples are discussed and illustrate the
wide-ranging examples of molecular switches in genome metabolism.

<>

<1>Szczelkun, M.D., Connolly, B.A.
<2>Sequence-specific binding of DNA by the EcoRV restriction and modification enzymes with nucleic acid and cofactor analogues.
<3>Biochemistry
<4>34
<5>10724-10733
<6>1995
<7>The DNA-binding properties of the EcoRV restriction endonuclease and modification
methyltransferase with their recognition sequence (GATATC) were analyzed using the
electrophoretic band-shift assay.  It has previously been observed that the endonuclease does
not bind specifically to GATATC sequences in the absence of the essential cofactor Mg2+.  To
investigate any possible roles for Mg2+ in promoting specific DNA binding, a set of
hydrolysis-resistant oligonucleotide substrates were synthesized that contained either
phosphate (phosphorothioate, 3'-S-phosphorothiolate), sugar (4'-thiothymidine), or base
(7-deaza-2'-deoxyadenosine) modifications.  However, it was found that none of these were
specifically bound by the endonuclease in either the absence or the presence of Mg2+.  In
contrast, the methylase bound to GATATC sequences much more strongly than to nonspecific
sites, and it was possible to observe the formation of enzyme-DNA complexes by gel
retardation.  Binding to GATATC sequences was increased by the addition of sinefungin, a
nonreactive analogue of the essential cofactor S-adenosyl-L-methionine (AdoMet).  Presumably
this also occurs with AdoMet although methylation and turnover prevented its direct
observation.  In the presence of sinefungin the strongest binding was observed with
hemimethylated EcoRV sequences (Kd=11-13 nM), and unmethylated DNA was bound less well
(Kd=46nM).  Specific, albeit weaker binding was also seen with the dimethylated product
(Kd=143 nM).  A difference in electrophoretic mobility was observed between enzyme-substrate
and enzyme-product complexes suggestive of structural differences between them.  The Kapp
value found for sinefungin, with the hemimethylated EcoRV sequence, was 10.9 mM.

<>

<1>Szczelkun, M.D., Dillingham, M.S., Janscak, P., Firman, K., Halford, S.E.
<2>Repercussions of DNA tracking by the type IC restriction endonuclease EcoR124I on linear, circular and catenated substrates.
<3>EMBO J.
<4>15
<5>6335-6347
<6>1996
<7>Type I restriction endonucleases such as EcoR124I cleave DNA at undefined loci, distant from
their recognition sequences, by a mechanism that involves the enzyme tracking along the DNA
between recognition and cleavage sites.  This mechanism was examined on plasmids that carried
recognition sites for EcoR124I and recombination sites for resolvase, the latter to create DNA
catenanes.  Supercoiled substrates with either one or two restriction sites were linearized by
EcoR124I at similar rates, although the two-site molecule underwent further cleavage more
readily than the one-site DNA.  The catenane from the plasmid with one EcoR124I site, carrying
the site on the smaller of the two rings, was cleaved by EcoR124I exclusively in the small
ring, and this underwent multiple cleavage akin to the two-site plasmid.  Linear substrates
derived from the plasmids were cleaved by EcoR124I at very slow rates.  The communication
between recognition and cleavage sites therefore cannot stem from random looping.  Instead, it
must follow the DNA contour between the sites.  On a circular DNA, the translocation of
non-specific DNA past the specifically bound protein should increase negative supercoiling in
one domain and decrease it in the other.  The ensuing topological barrier may be the trigger
for DNA cleavage.

<>

<1>Szczelkun, M.D., Friedhoff, P., Seidel, R.
<2>Maintaining a sense of direction during long-range communication on DNA.
<3>Biochem. Soc. Trans.
<4>38
<5>404-409
<6>2010
<7>Many biological processes rely on the interaction of proteins with multiple DNA sites
separated by thousands of base pairs. These
long-range communication events can be driven by both the thermal
motions of proteins and DNA, and directional protein motions that are
rectified by ATP hydrolysis. The present review describes conflicting
experiments that have sought to explain how the ATP-dependent Type Ill
restriction-modification enzymes can cut DNA with two sites in an
inverted repeat, but not DNA with two sites in direct repeat. We
suggest that an ATPase activity may not automatically indicate a DNA
translocase, but can alternatively indicate a molecular switch that
triggers communication by thermally driven DNA sliding. The generality
of this mechanism to other ATP-dependent communication processes such
as mismatch repair is also discussed.

<>

<1>Szczelkun, M.D., Halford, S.E.
<2>Recombination by resolvase to analyse DNA communications by the SfiI restriction endonuclease.
<3>EMBO J.
<4>15
<5>1460-1469
<6>1996
<7>The SfiI endonuclease differs from other type II restriction enzymes by cleaving DNA
concertedly at two copies of its recognition site, its optimal activity being with two sites
on the same DNA molecule.  The nature of this communication event between distant DNA sites
was analysed on plasmids with recognition sites for SfiI interspersed with recombination sites
for resolvase.  These were converted by resolvase to catenanes carrying one SfiI site on each
ring.  The catenanes were cleaved by SfiI almost as readily as a single ring with two sites,
in contrast to the slow reactions on DNA rings with one SfiI site.  Interactions between SfiI
sites on the same DNA therefore cannot follow the DNA contour and, instead, must stem from
their physical proximity.  In buffer lacking Mg2+, where SfiI is inactive while resolvase is
active, the addition of SfiI to a plasmid with target sites for both proteins blocked
recombination by resolvase, due to the restriction enzyme bridging its sites and thus
isolating the sites for resolvase into separate loops.  The extent of DNA looping by SfiI
matched its extent of DNA cleavage in the presence of Mg2+.

<>

<1>Szczelkun, M.D., Halford, S.E.
<2>Restriction Endonuclease.
<3>Encyclopaedia of Genetics, Academic Press, Brenner, S., Miller, J., New York
<4>0
<5>1693-1696
<6>2001
<7>The term 'restriction enzyme' was first coined more than 30 years ago, following the classic
observation that phage grown on one bacterial species would grow poorly on another strain of
the same species; the phage were 'restricted' in their host range.  The restriction activity
was dependent on an endonuclease which cleaves double-stranded DNA upon binding a specific
sequence of nucleotides (the recognition site).  However, transfer of a methyl group from the
donor S-adenosylmethionine to a particular base on one or both strands of the recognition site
prevents cleavage.  This modification is catalyzed by a methyltransferase activity.  The host
DNA can therefore be protected from self-digestion, even during semiconservative replication
when one strand is temporally unmodified.  Conversely, any invasive DNA is unlikely to carry
the correct pattern of methylated bases, and will be a target for cleavage.  The presence in
parallel of endonuclease and methyltransferase activities produces a bacterial equivalent of
an immune system, capable of distinguishing self from foreign DNA.  Probably as a consequence
of this custodial role, restriction endonucleases are widespread in nature, appearing in every
bacterial generus examined.

<>

<1>Szczelkun, M.D., Janscak, P., Firman, K., Halford, S.E.
<2>Selection of non-specific DNA cleavage sites by the type IC restriction endonuclease EcoR124I.
<3>J. Mol. Biol.
<4>271
<5>112-123
<6>1997
<7>The type IC restriction endonuclease EcoR124I binds specifically to its recognition sequence
but subsequently translocates non-specific DNA past the complex in an ATP-dependent mechanism.
The enzyme thus has the potential to cleave DNA at loci distant from the recognition site.  We
have scrutinized the link between translocation and cleavage on linear and circular DNA
substrates.  On linear DNA carrying two recognition sites, the majority of cleavages at loci
distant from the recognition site occurred between the two sites, regardless of the inter-site
distance or relative orientations.  On circular DNA carrying one site, distant cleavages
occurred throughout the DNA but an equivalent linear molecule underwent considerably fewer
cleavages at distant loci.  These results agree with published models for DNA tracking.
However, on every molecule investigated, discrete cleavage sites were also observed within
+/-250bp of the recognition sites.  The localized cleavages were not confined to particular
DNA sequences and were independent of DNA topology.  We propose a model to account for both
distant and localized cleavage events.  The conformation of the DNA loop extruded during
tracking may result in two DNA segments being held in proximity to the restriction moiety on
the protein, one close to the EcoR124I site and another distant from the site: cleavage may
occur in either segment.  Alternatively, the cutting of DNA close to recogniton sites may be
the result of multiple nicks being generated in the expanding loop before any extensive
translocation.

<>

<1>Szczelkun, M.D., Jones, H., Connolly, B.A.
<2>Probing the protein-DNA interface of the EcoRV modification methyltransferase bound to its recognition sequence, GATATC.
<3>Biochemistry
<4>34
<5>10734-10743
<6>1995
<7>The DNA contacts produced between the EcoRV modification methyltransferase and its recognition
sequence, GATATC, have been determined.  The enzyme's general location in a
methylase/DNA/sinefungin ternary complex was evaluated by protection from exonuclease III
digestion.  Important phosphate contacts were resolved using N-ethyl-N-nitrosourea ethylation
interference footprinting.  Methylation protection and interference using dimethyl sulfate
were employed to assess significant contacts to purinic bases.  The protein-DNA interface was
further probed using oligodeoxynucleotides containing base analogues within the GATATC
sequence.  Most of the experiments were carried out using hemimethylated sequences, i.e.,
having 6-methyladenosine at the methylation site in one of the strands.  The momomeric
methylase was found to bind to the DNA in two different orientations for the methylation of
each strand.  The enzyme approaches the DNA, predominantly from one "side", and makes most of
its contacts in the major groove.  In either of the two binding events contacts are made to
the four phosphates NpNpNpGpA and the three bases GAT (where GAT represents the 5' half of
the GATATC site) on both DNA strands.  The phosphates and bases in the 3' ATC half are much
less important.  Although the contacts made to the equivalent locations on each strand are
similar, they display a slight but consistent change dependent on which strand contains the
6-methyldeoxyadenosine.  This strand variation shows completely reciprocal behavior, switching
around exactly, depending entirely on the methylated deoxyadenosine location.  It is this
that provides evidence for the two binding modes.  The results obtained are discussed in terms
of possible models for the protein-DNA interface.

<>

<1>Szczepanek, T., Gora, M., Monteilhet, C., Wysocka, M., Lazowska, J., Golik, P.
<2>In vivo analysis of the relationships between the splicing and homing activities of a group I intron-encoded I-ScaI/bi2-maturase of  Saccharomyces capensis produced in the yeast cytoplasm.
<3>FEMS Yeast Res.
<4>6
<5>823-835
<6>2006
<7>The I-ScaI/bi2-maturase of Saccharomyces capensis acts as a specific homing endonuclease
promoting intron homing, and as a maturase
promoting intron splicing. Using the universal code equivalent of the
mitochondrial gene encoding the I-ScaI/bi2-maturase, a number of
truncated forms of the synthetic gene were constructed, shortened on
either side, as were several mutated alleles of the protein. The
shortest translation product that fully retains both activities in vivo
corresponds to 228 codons of the C-terminal region of the bi2
intron-encoded protein, whereas proteins resulting from more extensive
deletions either at the N-terminus or at the C-terminus (up to 73 and
four residues, respectively) were able to complement wholly the lack of
endogenous maturase, but all lost the endonuclease activity. Similarly,
all introduced mutations completely abolished the I-ScaI activity while
some mutant proteins retained substantial splicing function.
Immunodetection experiments demonstrated that different cytoplasmically
translated forms of the I-ScaI/bi2-maturase protein were imported into
mitochondria and correctly processed. They appeared to be tightly
associated with mitochondrial membranes. Homology modelling of the
I-ScaI/bi2-maturase protein allowed us to relate both enzymatic
activities to elements of enzyme structure.

<>

<1>Szczepanek, T., Jamoussi, K., Lazowska, J.
<2>Critical base substitutions that affect the splicing and/or homing activities of the group I intron bi2 of yeast mitochondria.
<3>Mol. Gen. Genet.
<4>264
<5>137-144
<6>2000
<7>The second intron (bi2) of the cyt b gene from related Saccharomyces species has an
extraordinarily conserved sequence and can have
different functions in wild-type cells. The protein encoded by the S.
cerevisiae intron functions as a maturase to promote intron splicing,
while the homologous S. capensis intron encodes a bifunctional protein
that acts both as a maturase and as a homing endonuclease (I-ScaI)
promoting intron mobility. The protein encoded by intron bi2 belongs to
a large gene family characterized by the presence of two conserved
LAGLIDADG motifs (P1 and P2). In this study, we analysed a set of
splicing-deficient mutants of the S. cerevisiae intron bi2 that carry
non-directed mutations affecting the maturase activity, and a set of
directed missense mutations introduced into the bifunctional protein
encoded by the S. capensis intron. Analysis of these mutations has
allowed identification of the residues in the conserved P1 and P2
motifs which are crucial for splicing and homing activities. Moreover,
several mutations which are located in the C-terminal part of the
protein have been found to affect both functions.

<>

<1>Szczepanek, T., Lazowska, J.
<2>Replacement of two non-adjacent amino acids in the S. cerevisiae bi2 intron-encoded RNA maturase is sufficient to gain a homing-endonuclease activity.
<3>EMBO J.
<4>15
<5>3758-3767
<6>1996
<7>Two homologous group I introns, the second intron of the cyt b gene, from related
Saccharomyces species differ in their mobility.  The S. capensis intron is mobile and encodes
the I-ScaI endonuclease promoting intron homing, whilst the homologous S. cerevisiae intron is
not mobile, but functions as an RNA maturase promoting splicing. These two intron-encoded
proteins differ by only four amino acid substitutions.  Taking advantage of the remarkable
similarity of the two intron open reading frames and using biolistic transformation of
mitochondria, we show that the replacement of only two non- adjacent residues in the S.
cerevisiae maturase carboxy-terminal sequence is sufficient to induce a homing-endonuclease
activity without losing the splicing function.  Also, we demonstrate that these two activities
reside in the S. capensis bi2-encoded protein which functions in both splicing and intron
mobility in the wild-type cells.  These results provide new insight into our understanding of
the activity and the evolution of group I intron- encoded proteins.

<>

<1>Szczepanek, T., Macadre, C., Meunier, B., Lazowska, J.
<2>Two homologous introns from related Saccharomyces species differ in their mobility.
<3>Gene
<4>139
<5>1-7
<6>1994
<7>We have studied gene conversion initiated by the ai3 intron of the Saccharomyces cerevisiae
mitochondrial COX1 gene and its homologous intron (S.cap.ai1) from Saccharomyces capensis.
The approach used involved the measurement of intron transmission amongst the progeny of
crosses between constructed recipient and donor strains.  We found that the S. cerevisiae ai3
intron is extremely active as a donor in gene conversion, whereas its homologous S. capensis
intron is not.  We have established the sequence of S.cap.ai1 and compared its open reading
frame with that of I-SceIII encoded by the homologous S. cerevisiae intron.  The two
protein-coding intron sequences are almost identical, except that the S. capensis ORF contains
an in-frame stop codon.  This finding provides a strong indication that the 3' part of the S.
cerevisiae intron ORF encoding I-SceIII (which should not be translated in the S. capensis
intron) must be critical for function of mtDNA endonucleases to mediate intron mobility.

<>

<1>Szczepanowski, R., Eikmeyer, F., Harfmann, J., Blom, J., Rogers, L.M., Brown, C.J., Top, E.M., Schluter, A.
<2>Sequencing and comparative analysis of IncP-1alpha antibiotic resistance plasmids reveal a highly conserved backbone and differences within accessory regions.
<3>J. Biotechnol.
<4>155
<5>95-103
<6>2011
<7>In spite of the importance of IncP-1 plasmids in horizontal gene transfer
among bacteria, in particular antibiotic resistance spread, so far only
three plasmids from the subgroup IncP-1alpha have been completely
sequenced. In this study we doubled this number. The three IncP-1alpha
plasmids pB5, pB11 and pSP21 were isolated from bacteria of two different
sewage treatment plants and sequenced by a combination of next-generation
and capillary sequencing technologies. A comparative analysis including
the previously analysed IncP-1alpha plasmids RK2, pTB11 and pBS228
revealed a highly conserved plasmid backbone (at least 99.9% DNA sequence
identity) comprising 54 core genes loaded with different accessory
elements. The accessory elements of the plasmid pB5 constitute a class 1
integron interrupting the parC gene and an IS6100 copy inserted into the
integron. In addition, the tetracycline resistance genes tetAR and the
ISTB11-like element are located between the klc operon and the trfA-ssb
operon. Plasmid pB11 is loaded with a Tn5053-like mercury resistance
transposon between the parCBA and parDE operons and contains tetAR that
are identical to those identified in plasmid pB5 and the insertion
sequence ISSP21. Plasmid pSP21 harbours an ISPa7 element in a Tn402
transposon including a class 1 integron between the partitioning genes
parCBA and parDE. The IS-element ISSP21 (99.89% DNA sequence identity to
ISSP21 from pB11), inserted downstream of the tetR gene and a copy of
ISTB11 (identical to ISTB11 on pTB11) inserted between the genes pncA and
pinR. On all three plasmids the accessory genes are almost always located
between the backbone modules confirming the importance of the backbone
functions for plasmid maintenance. The striking backbone conservation
among the six completely sequenced IncP-1alpha plasmids is in contrast to
the much higher diversity within the IncP-1beta subgroup.

<>

<1>Szczepanowski, R.H., Carpenter, M.A., Czapinska, H., Zaremba, M., Tamulaitis, G., Siksnys, V., Bhagwat, A.S., Bochtler, M.
<2>Central base pair flipping and discrimination by PspGI.
<3>Nucleic Acids Res.
<4>36
<5>6109-6117
<6>2008
<7>PspGI is a representative of a group of restriction endonucleases that recognize a pentameric
sequence related to CCNGG. Unlike the previously
investigated Ecl18kI, which does not have any specificity for the central
base pair, PspGI prefers A/T over G/C in its target site. Here, we present
a structure of PspGI with target DNA at 1.7 A resolution. In this
structure, the bases at the center of the recognition sequence are
extruded from the DNA and flipped into pockets of PspGI. The flipped
thymine is in the usual anti conformation, but the flipped adenine takes
the normally unfavorable syn conformation. The results of this and the
accompanying manuscript attribute the preference for A/T pairs over G/C
pairs in the flipping position to the intrinsically lower penalty for
flipping A/T pairs and to selection of the PspGI pockets against guanine
and cytosine. Our data show that flipping can contribute to the
discrimination between normal bases. This adds a new role to base flipping
in addition to its well-known function in base modification and DNA damage
repair.

<>

<1>Szczepek, M., Brondani, V., Buchel, J., Serrano, L., Segal, D.J., Cathomen, T.
<2>Structure-based redesign of the dimerization interface reduces the toxicity of zinc-finger nucleases.
<3>Nat. Biotechnol.
<4>25
<5>786-793
<6>2007
<7>Artificial endonucleases consisting of a FokI cleavage domain tethered to engineered
zinc-finger DNA-binding proteins have proven useful for
stimulating homologous recombination in a variety of cell types. Because
the catalytic domain of zinc-finger nucleases (ZFNs) must dimerize to
become active, two subunits are typically assembled as heterodimers at the
cleavage site. The use of ZFNs is often associated with significant
cytotoxicity, presumably due to cleavage at off-target sites. Here we
describe a structure-based approach to reducing off-target cleavage. Using
in silico protein modeling and energy calculations, we increased the
specificity of target site cleavage by preventing homodimerization and
lowering the dimerization energy. Cell-based recombination assays
confirmed that the modified ZFNs were as active as the original ZFNs but
elicit significantly less genotoxicity. The improved safety profile may
facilitate therapeutic application of the ZFN technology.

<>

<1>Szczepek, M., Mackeldanz, P., Moncke-Buchner, E., Alves, J., Kruger, D.H., Reuter, M.
<2>Molecular analysis of restriction endonuclease EcoRII from Escherichia coli reveals precise regulation of its enzymatic activity by autoinhibition.
<3>Mol. Microbiol.
<4>72
<5>1011-1021
<6>2009
<7>Bacterial restriction endonuclease EcoRII requires two recognition sites to cleave DNA.
Proteolysis of EcoRII revealed the existence of
two stable domains, EcoRII-N and EcoRII-C. Reduction of the enzyme to
its C-terminal domain, EcoRII-C, unleashed the enzyme activity; this
truncated form no longer needed two recognition sites and cleaved DNA
much more efficiently than EcoRII wild-type. The crystal structure of
EcoRII showed that probably the N-terminal domain sterically occludes
the catalytic site, thus apparently controlling the cleavage activity.
Based on these data, EcoRII was the first restriction endonuclease for
which an autoinhibition mechanism as regulatory strategy was proposed.
In this study, we probed this assumption and searched for the
inhibitory element that mediates autoinhibition. Here we show that
repression of EcoRII-C is achieved by addition of the inhibitory domain
EcoRII-N or by single soluble peptides thereof in trans. Moreover, we
perturbed contacts between the N- and the C-terminal domain of EcoRII
by site-directed mutagenesis and proved that beta-strand B1 and
alpha-helix H2 are essential for autoinhibition; deletion of either
secondary structural element completely relieved EcoRII autoinhibition.
This potent regulation principle that keeps EcoRII enzyme activity
controlled might protect bacteria against suicidal restriction of rare
unmodified recognition sites in the cellular genome.

<>

<1>Szczepinska, T., Kutner, J., Kopczynski, M., Pawlowski, K., Dziembowski, A., Kudlicki, A., Ginalski, K., Rowicka, M.
<2>Probabilistic approach to predicting substrate specificity of methyltransferases.
<3>PLOS Comp. Biol.
<4>10
<5>e1003514
<6>2014
<7>We present a general probabilistic framework for predicting the substrate specificity of
enzymes. We designed this approach to be easily applicable to
different organisms and enzymes. Therefore, our predictive models do not rely on
species-specific properties and use mostly sequence-derived data. Maximum
Likelihood optimization is used to fine-tune model parameters and the Akaike
Information Criterion is employed to overcome the issue of correlated variables.
As a proof-of-principle, we apply our approach to predicting general substrate
specificity of yeast methyltransferases (MTases). As input, we use several
physico-chemical and biological properties of MTases: structural fold,
isoelectric point, expression pattern and cellular localization. Our method
accurately predicts whether a yeast MTase methylates a protein, RNA or another
molecule. Among our experimentally tested predictions, 89% were confirmed,
including the surprising prediction that YOR021C is the first known MTase with a
SPOUT fold that methylates a substrate other than RNA (protein). Our approach not
only allows for highly accurate prediction of functional specificity of MTases,
but also provides insight into general rules governing MTase substrate
specificity.

<>

<1>Szeberenyi, J.
<2>Two restriction endonucleases.
<3>Biochem. Mol. Biol. Educ.
<4>34
<5>228-229
<6>2006
<7>Terms to be familiar with before you start to solve the test: restriction endonucleases;
palindromic sequences; blunt ends; cohesive
("sticky") ends; in vitro DNA synthesis; radioactive nucleotide
precursors; template-primer complexes; DNA ligation.|

<>

<1>Szegedi, S.S., Gumport, R.I.
<2>DNA binding properties in vivo and target recognition domain sequence alignment analyses of wild-type and mutant RsrI [N6-adenine] DNA methyltransferases.
<3>Nucleic Acids Res.
<4>28
<5>3972-3981
<6>2000
<7>A genetic selection method, the P22 challenge-phage assay, was used to characterize DNA
binding in vivo by the prokaryotic beta class [N6-adenine] DNA methyltransferase M.RsrI.
M.RsrI mutants with altered binding affinities in vivo were isolated. Unlike the wild-type
enzyme, a catalytically compromised mutant, M.RsrI (L72P), demonstrated site-specific DNA
binding in vivo. The L72P mutation is located near the highly conserved catalytic motif IV,
DPPY (residues 65-68). A double mutant, M.RsrI (L72P/D173A), showed less binding in vivo than
did M.RsrI (L72P). Thus, introduction of the D173A mutation deleteriously affected DNA
binding. D173 is located in the putative target recognition domain (TRD) of the enzyme.
Sequence alignment analyses of several beta class MTases revealed a TRD sequence element that
contains the D173 residue. Phylogenetic analysis suggested that divergence in the amino acid
sequences of these methyltransferases correlated with differences in their DNA target
recognition sequences. Furthermore, MTases of other classes (alpha and gamma) having the same
DNA recognition sequence as the beta class MTases share related regions of amino acid
sequences in their TRDs.

<>

<1>Szegedi, S.S., Ma, Z., Gumport, R.I.
<2>A novel scheme for the purification of wildtype and mutant RsrI DNA methyltransferases (MTases).
<3>FASEB J.
<4>12
<5>A1437
<6>1998
<7>RsrI DNA MTase (M.RsrI) is an S-adenosylmethionine-dependent enzyme that recognizes the duplex
DNA sequence 5-GAATTC-3 and methylates the central adenine at the N6 position.  M.RsrI is
isocatalytic but not homologous with M.EcoRI.  These two proteins show only 15.6% amino acid
identity.  Our laboratory has purified M.RsrI and is generating M.RsrI mutants as part of an
effort to understand how the two dissimilar proteins perform the same catalytic function.  A
simpler method for purifying the wild-type M.RsrI has been developed, reducing the
purification steps from six to two.  The isolation of wild-type M.RsrI to >90% purity from
8.0g of E. coli cells paste yields approximately 7.5 mg of protein.  This amount represents a
>4-fold increase in the yield of protein as compared to our previous purification method.  A
mutant, M.RsrI (L72P), is currently being purified in a similar manner.  This mutant was
generated by random PCR mutagenesis and isolated using the in vivo DNA binding assay, the
bacteriophage P22 challenge-phage assay.  Wild-type M.RsrI is toxic and shows no DNA binding
in the assay.  M.RsrI (L72P), on the other hand, is non-toxic and gives reproducible binding.
Catalytic assays of >70% pure M.RsrI (L72P) have shown that it retains weak catalytic
activity.  This mutation lies close to the highly conserved catalytic motif "DPPY", and may
affect catalysis and/or AdoMet binding.  We will generate more M.RsrI for attempts at
crystallization, and will purify M.RsrI (L72P) and other mutants recently generated by
site-directed PCR mutagenesis.

<>

<1>Szegedi, S.S., Reich, N.O., Gumport, R.I.
<2>Characterization of the DNA binding and kinetic properties of wild-type Rsr[N6-adenine] methyltransferase (M.RsrI).
<3>FASEB J.
<4>13
<5>A1367
<6>1999
<7>M.RsrI recognizes the DNA sequence GAATTC and catalyzes the transfer of a methyl group from
S-adenosylmethionine to the exocyclic amino group of the central adenine to form GamATTC and
S-adenosylhomocysteine.  Our research focuses on the characterization for the DNA binding,
catalytic and structural properties of M.RsrI.  First the wild-type protein was purified to
>95% homogeneity using a simplified procedure involving two chromatographic steps.  Gel-shift
assays of the protein were then performed on 1 or 5 nM canonical unmethylated, hemimethylated,
and dimethylated, or control (CTTAAG) 25-mer DNA duplexes in the presence of sinefungin, a
potent inhibitory analogue of AdoMet.  The following KD values obtained: hemimethylated, 3-7
nM; unmethylated, 30nM; and dimethylated and control, > 2500 nM.  Some pre-steady state
kinetic parameters of M.RsrI have been determined.  Upon the addition of enzyme pre-incubated
with radiolabeled AdoMet, M.RsrI shows a "burst" of methylated product formation equivalent to
the enzyme concentration in the presence of excess hemimethylated 14-mer DNA and radiolabeled
AdoMet.  This is followed by a slower step that represents the catalytic turnover (kcat).  The
dissociation constants for AdoMet (8.4 microM), AdoHcy (3.5 microM), and sinefungin (4.6
microM) have been determined for M.RsrI using an intrinsic tryptophan fluorescence quenching
assay.

<>

<1>Szegedi, S.S., Reich, N.O., Gumport, R.I.
<2>Substrate binding in vitro and kinetics of RsrI [N6-adenine] DNA methyltransferase.
<3>Nucleic Acids Res.
<4>28
<5>3962-3971
<6>2000
<7>RsrI [N6-adenine] DNA methyltransferase (M.RsrI), which recognizes GAATTC and is a member of a
restriction-modification system in Rhodobacter sphaeroides, was purified to >95% homogeneity
using a simplified procedure involving two ion exchange chromatographic steps. Electrophoretic
gel retardation assays with purified M.RsrI were performed on unmethylated, hemimethylated,
dimethylated or non-specific target DNA duplexes (25 bp) in the presence of sinefungin, a
potent inhibitory analog of AdoMet. M.RsrI binding was affected by the methylation status of
the DNA substrate and was enhanced by the presence of the cofactor analog. M.RsrI bound DNA
substrates in the presence of sinefungin with decreasing affinities: hemimethylated >
unmethylated > dimethylated >> non-specific DNA. Gel retardation studies with DNA substrates
containing an abasic site substituted for the target adenine DNA provided evidence consistent
with M.RsrI extruding the target base from the duplex. Consistent with such base flipping, an
approximately 1.7-fold fluorescence intensity increase was observed upon stoichiometric
addition of M.RsrI to hemimethylated DNA containing the fluorescent analog 2-aminopurine in
place of the target adenine. Pre-steady-state kinetic and isotope-partitioning experiments
revealed that the enzyme displays burst kinetics, confirmed the catalytic competence of the
M.RsrI-AdoMet complex and eliminated the possibility of an ordered mechanism where DNA is
required to bind first. The equilibrium dissociation constants for AdoMet, AdoHcy and
sinefungin were determined using an intrinsic tryptophan fluorescence-quenching assay.

<>

<1>Szekeres, M.
<2>Phage-induced development of a site-specific endonuclease in Anacystis nidulans, a cyanobacterium.
<3>Virology
<4>111
<5>1-10
<6>1981
<7>Anacystis nidulans infected by cyanophage AS-1 produces a site-specific endonuclease cleaving
double-stranded DNA, as judged from the gel electrophoretic analysis of the reaction products
and the determination of the terminal nucleotide at the 5'-end of the fragments.  The enzyme
was purified to near electrophoretic homogeneity.  It has a molecular weight of 40,000 +/-
4,000.  Only Mg2+ ions are required for enzyme activity.  Limit digestion was not obtained
even after extensive digestion of substrate DNA with large amounts of the purified enzyme.
This suggests that the endonuclease splits the substrate at more than one nucleotide sequence
with a different efficiency for each sequence.  The following observations suggest that the
endonuclease takes part in the breakdown of host DNA in the infected cell: purified host DNA
is degraded by, while cyanophage AS-1 DNA is protected against, the enzyme, and the appearance
of the endonuclease during the lytic cycle coincides with the onset of intensive degradation
of host DNA.

<>

<1>Szekeres, M., Matveyev, A.V.
<2>Cleavage and sequence recognition of 2,6-diaminopurine-containing DNA by site-specific endonucleases.
<3>FEBS Lett.
<4>222
<5>89-94
<6>1987
<7>The susceptibility of 2,6-diaminopurine (DAP)-containing bacteriophage DNA to
several restriction and other endonucleases was examined.  With the only
exception of TaqI, these enzymes did not accept the modified base as a
substitute for adenine.  The phage DNA was extensively fragmented by the
restriction endonucleases which recognized only G and C-containing sites.
5'-terminal analysis of the MspI and SmaI fragments revealed that d(DAP-T)
basepairs can be mistaken by some enzymes for d(G-C) pairs.

<>

<1>Szekeres, M., Szmidt, A.E., Torok, I.
<2>Evidence for a restriction/modification-like system in Anacystis nidulans infected by cyanophage AS-1.
<3>Eur. J. Biochem.
<4>131
<5>137-141
<6>1983
<7>Anacystis nidulans infected by AS-1 cyanophage contains an endonuclease (AS-1
endonuclease) which splits host DNA but not AS-1 phage DNA [Szekeres, M. (1981)
Virology, 111, 1-10].  AS-1 phage DNA proved to be resistant not only to AS-1
endonuclease but also to a number of restriction endonucleases the recognition
sites of which contain a central dG-dC dinucleotide.  Since an unmodified
5'dG-dC dinucleotide was shown to be present at the sites at which DNA is
cleaved by AS-1 endonuclease, the results suggest that the sites attacked
preferentially by the AS-1 endonuclease are specifically protected on the AS-1
DNA molecule.  The modification of AS-1 DNA was shown to occur specifically in
infected Anacystis because AS-1 DNA fragments which are normally resistant to
AS-1 endonuclease became susceptible to this enzyme if inserted into pBR322
plasmid and cloned in Escherichia coli.  AS-1 DNA was shown to contain about 5%
of a modified nucleotide which was not 5-methyldeoxycytidylic acid.  Results
presented and our earlier data suggest that in Anacystis infected by AS-1
phage, a restriction/modification-like system operates which is able to
elimminate 'unwanted' (host) DNA selectively.

<>

<1>Szeto, M.D., Boissel, S.J.S., Baker, D., Thyme, S.B.
<2>Mining Endonuclease Cleavage Determinants in Genomic Sequence Data.
<3>J. Biol. Chem.
<4>286
<5>32617-32627
<6>2011
<7>Homing endonucleases have great potential as tools for targeted gene therapy and gene
correction, but identifying variants of these enzymes
capable of cleaving specific DNA targets of interest is necessary
before the widespread use of such technologies is possible. We
identified homologues of the LAGLIDADG homing endonuclease I-AniI and
their putative target insertion sites by BLAST searches followed by
examination of the sequences of the flanking genomic regions. Amino
acid substitutions in these homologues that were located close to the
target site DNA, and thus potentially conferring differences in target
specificity, were grafted onto the I-AniI scaffold. Many of these
grafts exhibited novel and unexpected specificities. These findings
show that the information present in genomic data can be exploited for
endonuclease specificity redesign.

<>

<1>Szilak, L., Der, A., Deak, F., Venetianer, P.
<2>Kinetic characterization of the EcaI methyltransferase.
<3>Eur. J. Biochem.
<4>218
<5>727-733
<6>1993
<7>A kinetic analysis of the EcaI adenine-N6-specific methyltransferase (MTase) is presented. The
enzyme catalyzes the transfer of a methyl group from S-adenosyl-L-methionine (AdoMet) to the
adenine of the GGTNACC sequence with a random rapid-equilibrium mechanism. Experiments with a
synthetic, 14-bp DNA substrate suggest that recognition of the specific site of DNA occurs
after the binding of AdoMet. Proton concentration does not affect the dissociation constant of
AdoMet while Vm and the dissociation constant of DNA show a maximum around pH 8. Increasing
the amount of S-adenosyl-L-homocysteine decreases the inhibitory effect of methylated DNA
which proves the active role of AdoMet in site recognition. Experiments with hemimethylated
DNA show that the methylase binds the double-stranded DNA asymmetrically.

<>

<1>Szilak, L., Finta, C., Patthy, A., Venetianer, P., Kiss, A.
<2>Self-methylation of BspRI DNA-methyltransferase.
<3>Nucleic Acids Res.
<4>22
<5>2876-2881
<6>1994
<7>The DNA (cytosine-5)-methyltransferase (m5C-MTase) M.BspRI is able to accept the methyl group
from the methyl donor S-adenosyl-L-methionine (AdoMet) in the absence of DNA. Transfer of the
methyl group to the enzyme is a slow reaction relative to DNA methylation. Self-methylation is
dependent on the native conformation of the enzyme and is inhibited by
S-adenosyl-L-homocysteine, DNA and sulfhydryl reagents. Amino acid sequencing of proteolytic
peptides obtained from M.BspRI, which had been methylated with [methyl-3H]AdoMet, and thin
layer chromatography of the modified amino acid identified two cysteines, Cys156 and Cys181
that bind the methyl group in the form of S-methylcysteine. One of the acceptor residues,
Cys156 is the highly conserved cysteine which plays the role of the catalytic nucleophile of
m5C-MTases.

<>

<1>Szilak, L., Finta, C., Patthy, A., Venetianer, P., Kiss, A.
<2>Self-methylation of the M.BspRI methyltransferase.
<3>Gene
<4>157
<5>105
<6>1995
<7>In the absence of DNA substrate, the DNA methyltransferase (MTase) M.BspRI can methylate
itself using the methyl donor S-adenosyl-L-methionine (AdoMet).  The methyl group is
transferred to two Cys residues of the MTase.

<>

<1>Szilak, L., Venetianer, P., Kiss, A.
<2>Purification and biochemical characterization of the EcaI DNA methyltransferase.
<3>Eur. J. Biochem.
<4>209
<5>391-397
<6>1992
<7>The EcaI GGTNACC-specific DNA-adenine modification methyltransferase has been purufied to
apparent homogeneity. The active form of the DNA methyltransferase is a single polypeptide.
The enzyme has a pH optimum at pH 8.0 and a temperature optimum at 25oC. EcaI DNA
methyltransferase transfers one methyl group to the adenine of the recognition site in a
single binding event. The Km was 170 nM for DNA and 1.8 microM for the methyl donor
S-adenosylmethionine. Methylated DNA is a competitive inhibitor with respect to DNA (Ki-3.5
nM). The other product of the DNA-methylation reaction, S-adenosylhomocysteine was found to be
a competitive inhibitor with respect to S-adenosylmethionine (Ki= 2.7 uM). The
S-adenosylmethionine analog sinefungin was shown to be a very strong inhibitor (Ki = 3.5 nM)
of the DNA methyltransferase reaction.

<>

<1>Szilak, L., Venetianer, P., Kiss, A.
<2>Cloning and nucleotide sequence of the genes coding for the Sau96I restriction and modification enzymes.
<3>Nucleic Acids Res.
<4>18
<5>4659-4664
<6>1990
<7>The genes coding for the GGNCC specific Sau96I restriction and modification
enzymes were cloned and expressed in E. coli.  The DNA sequence predicts a 430
amino acid protein (Mr: 49,252) for the methyltransferase and a 261 amino acid
protein (Mr: 30,486) for the endonuclease.  No protein sequence similarity was
detected between the Sau96I methyltransferase and endonuclease.  The
methyltransferase contains the sequence elements characteristic for m5C
methyltransferases.  In addition to this, M.Sau96I shows similarity, also in
the variable region with one m5C-methyltransferase (M.SinI) which has closely
related recognition specificity (GGA/TCC).  M.Sau96I methylates the internal
cytosine within the GGNCC recognition sequence.  The Sau96I endonuclease
appears to act as a monomer.

<>

<1>Sznyter, L.A., Brooks, J.E.
<2>The characterization and cloning of the NotI restriction-modification system.
<3>Heredity
<4>61
<5>308
<6>1988
<7>NotI, a Type II restriction-modification system from the actinomycete Nocardia
otitidis-caviarum, recognizes the sequence GCGGCCGC.  We are currently purifying and
characterizing the proteins of this system.  The endonuclease, which cleaves at GC^GGCCGC, has
been sized by gel filtration and estimated to be approximately 85,000 daltons.  The NotI
methylase protein has been partially purified and we are in the process of determining its
size as well as the site and type (N-4 or C-5) of modification it provides.  The N. otitidis
genomic DNA has been purified and analyzed by HPLC analysis for base composition and for the
presence of modified bases.  Two approaches are being used to clone the NotI system.  First
genomic libraries constructed in E. coli are being selected for methylase expression.  Second,
genomic libraries will also be made and selected in Streptomyces lividans.  In addition to
selection for methylase expression, we also will be selecting the libraries with phage.  This
approach has been successfully used in cloning the SalI restriction-modification system.

<>

<1>Sznyter, L.A., Brooks, J.E.
<2>The characterization and cloning of the EagI restriction-modification system.
<3>Gene
<4>74
<5>53
<6>1988
<7>Meeting Abstract

<>

<1>Sznyter, L.A., Marcus, A.M., Brooks, J.E.
<2>The cloning and characterization of the EagI restriction-modification system.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>89
<5>180
<6>1989
<7>EagI is a Type II restriction-modification system from the gram-negative
enteric bacterium Enterobacter agglomerans.  It recognizes the sequence CGGCCG
with the endonuclease cleaving at C^GGCCG.  The genomic DNA from this organism
has been purified and analyzed by HPLC analysis for base composition and for
the presence of modified bases.  The EagI system has been cloned on a 2.9 kb
PstI fragment in pLSN3 (a pBR322 derivative containing three NotI sites) by
selection with the NotI endonuclease.  The 2.9kb PstI fragment was then cloned
into pUC19 in both orientations.  On this higher copy plasmid, an increase of
endonuclease activity was found in the crude extracts.  The level of increase
is dependent on orientation.  These clones are currently being characterized.
We have explored the Mcr phenotype of the most active EagI containing clone and
find that it is highly restricted by mcrB.  The boundaries of the genes have
been assigned by deletion subcloning and Tn5 mutagenesis.  The protein products
of the EagI system are also being characterized.  The endonuclease has been
purified and sized by gel filtration.  EagI is estimated to be approximately
78,000 daltons.  The EagI methylase protein has been purified and the size as
well as site and type (N-4 or C-5) of modification it provides is being
determined.

<>

<1>Sznyter, L.A., Slatko, B., Moran, L., O'Donnell, K.H., Brooks, J.E.
<2>Nucleotide sequence of the DdeI restriction-modification system and characterization of the methylase protein.
<3>Nucleic Acids Res.
<4>15
<5>8249-8266
<6>1987
<7>The DdeI restriction-modification system was previously cloned and has been
maintained in E. coli on two separate and compatible plasmids.  The nucleotide
sequence of the endonuclease and methylase genes has now been determined; it
predicts proteins of 240 amino acids, Mr=27,808, and 415 amino acids,
Mr=47,081, respectively.  Inspection of the DNA sequence shows that the 3' end
of the methylase gene had been deleted during cloning.  The clone containing
the complete methylase gene was made and compared to that containing the
truncated gene; only clones containing the truncated form support the
endonuclease gene in E. coli.  Bal-31 deletion studies show that methylase
expression in the Dde clones is also dependent upon orientation of the gene
with respect to pBR322.  The truncated and complete forms of the methylase
protein were purified and compared; the truncated form appears to be more
stable and active in vitro.  Finally, comparison of the deduced amino acid
sequence of M.DdeI with that of other known cytosine methylases shows
significant regions of homology.

<>

<1>Sznyter, L.A., Vaccaro, C.M., Jager-Quinton, T., Wilson, G., Brooks, J.E.
<2>Cloning and characterization of the SphI restriction-modification system from S. phaeochromogenes.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>0
<5>361
<6>1989
<7>SphI, a Type II restriction-modification system from the actinomycete Streptomyces
phaeochromogenes, recognizes the sequence GCATGC.  The endonuclease cleaves at GCATG/C leaving
a 3' four base overhang.  A 5.4kb insert carrying the methylase gene has been cloned into the
PstI site of pBR322 and expressed in E. coli at a low level as evidenced by incomplete
modification of chromosomal DNA and lack of modification of phage DNA in vivo.  The clone has
no endonuclease activity.  The plasmid has been extensively mapped; Tn-5 insertion mutagenesis
is being used to locate the methylase gene.  The fragment is also being positioned by Southern
hybridization onto the S. phaeochromogenes chromosome.  Attempts will be made to clone and
express the complete system in Streptomyces.

<>

<1>Szomolanyi, I., Kiss, A., Venetianer, P.
<2>Cloning the modification methylase gene of Bacillus sphaericus R in Escherichia coli.
<3>Gene
<4>10
<5>219-225
<6>1980
<7>The gene coding for the sequence-specific modification methylase methM - BspI
of Bacillus sphaericus R has been cloned in Escherichia coli by means of
plasmid pBR322.  The selection was based on the expression of the cloned gene
which rendered the recombinant plasmid resistant to BspI restriction
endonuclease cleavage.  The gene is carried by a 9 kb BamHI fragment and by a
smaller 2.5 kb EcoRI fragment derived from the BamHI fragment.  The
Bsp-specific methylase level was found to be higher in the recombinant clones
than in the parental strain.  The methylase gene is probably located on the
Bacillus sphaericus chromosome, and not on a plasmid known to be carried by
this strain.  The recombinant clones do not exhibit any BspI restriction
endonuclease activity.

<>

<1>Szwagierczak, A., Brachmann, A., Schmidt, C.S., Bultmann, S., Leonhardt, H., Spada, F.
<2>Characterization of PvuRts1I endonuclease as a tool to investigate genomic 5-hydroxymethylcytosine.
<3>Nucleic Acids Res.
<4>39
<5>5149-5156
<6>2011
<7>In mammalian genomes a sixth base, 5-hydroxymethylcytosine ((hm)C), is generated by enzymatic
oxidation of 5-methylcytosine ((m)C). This
discovery has raised fundamental questions about the functional relevance
of (hm)C in mammalian genomes. Due to their very similar chemical
structure, discrimination of the rare (hm)C against the far more abundant
(m)C is technically challenging and to date no methods for direct
sequencing of (hm)C have been reported. Here, we report on a purified
recombinant endonuclease, PvuRts1I, which selectively cleaves
(hm)C-containing sequences. We determined the consensus cleavage site of
PvuRts1I as (hm)CN(11-12)/N(9-10)G and show first data on its potential to
interrogate (hm)C patterns in mammalian genomes.

<>

<1>Szybalski, W.
<2>Universal restriction endonucleases: designing novel cleavage specificities by combining adapter oligodeoxynucleotide and enzyme moieties.
<3>Gene
<4>40
<5>169-173
<6>1985
<7>Class IIS restriction endonucleases cleave double-stranded (ds) DNA at precise
distances from their recognition sequences.  A method is proposed which
utilizes this separation between the recognition site and the cut site to allow
a class IIS enzyme, e.g., FokI, to cleave practically any predetermined
sequence by combining the enzyme with a properly designed oligodeoxynucleotide
adapter.  Such an adapter is constructed from the constant recognition site
domain (a hairpin containing the ds sequence, e.g., G/C G/C A/T T/A G/C for
FokI) and a variable, single-stranded (ss) domain complementary to the ss
sequence to be cleaved (at 9 and 13 nucleotides on the paired strands from the
recognition sequence in the exampled of FokI).  The ss sequence designated to
be cleaved could be provided by ss phage DNA (e.g., M13), gapped ds plasmids,
or supercoiled ds plasmids that were alkali denatured and rapidly neutralized.
Combination of all three components, namely the class IIS enzyme, the ssDNA
target sequence, and the complementing adapter, would result in target DNA
cleavage at the specific predetermined site.  The target ss DNA could be
converted to the precisely cleaved ds DNA by DNA polymerase, utilizing the
adapter oligodeoxynucleotide as primer.  This novel procedure represents the
first example of changing enzyme specificity by synthetic design.  A
practically unlimited assortment of new restriction specificities could be
produced.  The method should have many specific and general applications when
its numerous ramification are exploited.

<>

<1>Szybalski, W.
<2>Modifying specificities of restriction enzymes.
<3>Biotechnology: Bridging Research and Applications, Kluwer, Kamely, D., Chakrabarty, A.M., Kornguth, S.E., Boston
<4>0
<5>371-376
<6>1991
<7>For the last five years, our laboratory has been studying various aspects of
controlled cleavage of DNA, both for (1) the 50 bp-10 kb range of
single-stranded (ss) and double-stranded (ds) DNA fragments, plasmids or
phages, and (2) for very large genomes of bacteria, yeast and higher
eukaryotes.  We are using commerical restriction enzymes (ENases) either (i) of
the class-IIS variety (ENase-IIS) (Szybalski, 1985), where the recognition site
on the DNA for the ENase is separated by several base pairs from the cleavage
site, together with an adapter oligodeoxynucleotide (oligo), or (ii) various
combinations of a particular ENase, its cognate methyltransferase (MTase) and
DNA-binding protein whose binding site contains the corresponding ENase/MTase
recognition site.  The ENase-IIS enzymes were used either for precise cutting
(Podhajska and Szybalski, 1985) or for precise trimming of DNA (Hasan et al,
1986), whereas the MTase/ENase/DNA-binding protein combination was used for
very rare cuts in large genomes (Koob et al., 1968; Grimes et al., 1990; Koob
and Szybalski, 1990).

<>

<1>Szybalski, W.
<2>Universal restriction endonuclease.
<3>US Patent Office
<4>US 4935357
<5>
<6>1990
<7>A universal restriction endonuclease for cleaving a target DNA at a predetermined site. The
universal restriction endonuclease utilizes a tailored oligodeoxynucleotide adapter in
conjunction with a restriction enzyme which cuts DNA at a known distance from its recognition
site. The oligodeoxynucleotide adapter mimics the relationship which exists in such enzymes
between its recognition site and cut site.

<>

<1>Szybalski, W.
<2>Universal restriction endonuclease.
<3>European Patent Office
<4>EP 234781
<5>
<6>1986
<7>A universal restriction endonuclease for cleaving a target DNA at a predetermined site. The
universal restriction endonuclease utilizes a tailored oligodeoxynucleotide adapter in
conjunction with a restriction enzyme which cuts DNA at a known distance from its recognition
site. The oligodeoxynucleotide adapter mimics the relationship which exists in such enzymes
between its recognition site and cut site.

<>

<1>Szybalski, W., Blumenthal, R.M., Brooks, J.E., Hattman, S., Raleigh, E.A.
<2>Nomenclature for bacterial genes coding for class-II restriction endonucleases and modification methyltransferases.
<3>Gene
<4>74
<5>279-280
<6>1988
<7>A proposed nomenclature for restriction enzyme and methylase genes.

<>

<1>Szybalski, W., Kim, S.C., Hasan, N., Podhajska, A.J.
<2>Class-IIS restriction enzymes - a review.
<3>Gene
<4>100
<5>13-26
<6>1991
<7>Class-IIS restriction enzymes (ENases-IIS) interact with two discrete sites on
double-stranded DNA: the recognition site, which is 4-7 bp long, and the
cleavage site, usually 1-20 bp away from the recognition site.  The recognition
sequences of ENases-IIS are totally (or partially) asymmetric and all of the
characterized ENases-IIS are monomeric.  A total of 35 ENases-IIS are described
(80, if all isoschizomers are taken into consideration) together with ten
related ENases (class IIT), and 15 cognate methyltransferases (MTases-IIS).
The physical, chemical, and molecular properties of the ENases-IIS and
MTases-IIS are reviewed and many unique applications of this class of enzymes
are described, including: precise trimming of DNA; retrieval of cloned
fragments; gene assembly; use as a universal restriction enzyme; cleavage of
single-stranded DNA; detection of point mutations; tandem amplification;
printing-amplification reaction; and localization of methylated bases.

<>

<1>Szybalski, W., Skalka, A.
<2>Nobel prizes and restriction enzymes.
<3>Gene
<4>4
<5>181-182
<6>1978
<7>We have two reasons to rejoice in the wise choices made by the Nobel Committee in 1978.  The
discovery of the restriction enzymes and their application in genetic mapping, a main topic of
our journal, were selected for this award, and one of our Editors, Dr. Hamilton O. Smith, was
honored as one of the three recipients.

<>

<1>Szyf, M.
<2>DNA methylation properties: consequences for pharmacology.
<3>Trends Pharmacol. Sci.
<4>15
<5>233-238
<6>1994
<7>DNA methylation plays an important role in controlling the profile of gene expression of
mammalian cells. The hypothesis presented in this article is that DNA methylation patterns are
determined by an interplay between the level of DNA methyltransferase and demethylase
activities and site-specific signals. The expression of the DNA methyltransferase gene is
regulated with the proliferative state of the cell and it is upregulated by cellular oncogenic
pathways, resulting in hypermethylation and repression of tumour-suppressing loci. DNA
methyltransferase inhibitors would inhibit the excessive activity of DNA methyltransferase in
cancer cells and induce the original cellular programme of tumour suppression. They can also
be used to turn on alternative programmes of gene expression. Specific DNA methyltransferase
antagonists might provide us with therapeutic agents directed at a nodal point of regulation
of genetic information.

<>

<1>Szyf, M.
<2>Inhibition of DNA methyltransferase.
<3>International Patent Office
<4>WO 9515373
<5>
<6>1995
<7>The present invention relates to the interplay between the level of DNA methyltransferase and
demethylase activities, to the role of the interplay between these levels on the
proliferative, differentiated, tumorigenic and homeostatic state of the cell, and to the DNA
methyltransferase and demethylase as therapeutic targets.  The invention further relates to a
reduction of the level of DNA methylation through inhibitors and antagonist in order to
inhibit the excessive activity or hypermethylation of DNA MeTase in cancer cells to induce the
original cellular tumor suppressing program, to turn on alternative gene expression programs,
to provide therapeutics directed at a nodal point of regulation of genetic information, and
to modulate the general level of methylase and demethylase enzymatic activity of a cell to
permit specific changes in the methylation pattern of a cell.

<>

<1>Szyf, M.
<2>Targeting DNA methyltransferase in cancer.
<3>Cancer Metastasis Rev.
<4>17
<5>219-231
<6>1998
<7>DNA methyltransferase is an enzyme responsible for generating and maintaining DNA methylation
patterns.  DNA methylation patterns control different genome functions, thus they are an
important component of the epigenetic information.  It has been recently postulated that DNA
methyltransferase plays an important role in oncogenesis and that it is a candidate target for
anticancer therapy.  This commentary discusses the possible mechanisms through which DNA
methyltransferase participates in oncogenesis and the rationale for targeting it in cancer.

<>

<1>Szyf, M.
<2>The role of DNA methyltransferase 1 in growth control.
<3>Front. Biosci.
<4>6
<5>D599-D609
<6>2001
<7>Vertebrate DNA contains in addition to the four bases comprising the genetic information a
modified base, 5-methylcytosine that plays an important role in the epigenome. The methylated
bases form a pattern of methylation that is cell specific and is faithfully inherited during
cell division. The enzyme DNA methyltransferase 1 DNMT1 is responsible for copying the DNA
methylation pattern but other de novo methyltransferase as well as demethylases might also be
involved. Multiple mechanisms are in place to ensure the coordinate inheritance of the DNA
methylation pattern with DNA replication. There is a bilateral relationship between the cell
cycle and DNMT1. The expression of DNMT1 is tightly regulated with the cell cycle while the
expression of DNMT1 can affect the cell cycle. DNMT1 protein might regulate cell cycle events
by mechanisms that are independent of its DNA methylation activity through its multiple
protein-protein interactions. The unique position of DNMT1 in the cell cycle is consistent
with the hypothesis that it plays an important role in cancer.

<>

<1>Szyf, M.
<2>The DNA methylation machinery as a therapeutic target.
<3>Curr. Drug Targets
<4>1
<5>101-118
<6>2000
<7>Pharmacology and therapeutics have traditionally focused on altering the activity of specific
proteins that play an important physiological role either to counteract disease processes or
to achieve changes in physiology that are of benefit to the patient.  The explosion in our
understanding of gene expression programs opens up new horizons for pharmacological
intervention.  Key processes reversibly controlling genetic programs are attractive drug
targets.  DNA methylation is a fundamental feature of genomes and the control of their
function and is therefore a candidate for pharmacological manipulation that might have
important therapeutic advantage.  This review argues that DNA methylation plays an important
role in the control of genomic processes, suggests how the DNA methylation machinery is
involved in cancer, identifies the enzymatic processes that are a target for drug
intervention, proposes potential therapeutic utilities for agents that manipulate the DNA
methylation machinery and discusses novel drugs that target the DNA methylation machinery.

<>

<1>Szyf, M.
<2>DNA methylation and cancer therapy.
<3>Drug Resistance Updates
<4>6
<5>341-353
<6>2003
<7>Vertebrate DNA is modified by methyl moieties at the 5'-position of cytosine rings residing
in the di-nucleotide sequence CpG. Approximately
80% of CpG dinucleotide sequences are methylated. The pattern of
distribution of methylated CGs is cell-type specific and correlates with
gene expression programming and chromatin structure. Three kinds of
seemingly contradictory aberrations in DNA methylation are observed in
cancer, global hypomethylation, and regional hypermethylation and
deregulated level of expression of DNA methyltransferases. It was
previously proposed that the DNA methylation machinery is a candidate
target for anticancer therapy. Inhibition of hypermethylation was the
first therapeutic target. However, recent data suggests that inhibition of
DNA methylation might have untoward effects such as induction of genes
involved in metastasis. This review discusses the relative role of the
three levels of alteration in the DNA methylation in cancer, proposes a
unified hypothesis on the relative roles of increased DNA
methyltransferase as well as the coexistence of hypo -and hyper-
methylation in cancer and its possible implications on anticancer therapy.

<>

<1>Szyf, M.
<2>Protein sequences of human and mouse DNA demethylase and uses in screening demethylase inhibitors for tumor therapy.
<3>US Patent Office
<4>US 20050003378 A
<5>23
<6>2005
<7>The present invention relates to a DNA demethylase (dMTase) inhibitor which comprises
S-adenosylmethionine, a metabolite of S-adenosylmethionine or a pharmaceutically acceptable
salt thereof.

<>

<1>Szyf, M., Avraham-Haetzni, K., Reifman, A., Shlomai, J., Kaplan, F., Oppenheim, A., Razin, A.
<2>DNA methylation pattern is determined by the intracellular level of the methylase.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>81
<5>3278-3282
<6>1984
<7>Extrachromosomal plasmid DNA is transiently undermethylated in Escherichia coli during
amplification in the presence of chloramphenicol. In addition, undermethylation of phage
lambda DNA was observed after thermal induction of a lambda cI857 lysogen while the integrated
lambda phage DNA was found to be fully methylated. these methylation pattern changes occur
under conditions (extensive replication) in which the intracellular methylase level becomes
limiting. In an E. coli strain that harbors a plasmid that carries the dam methylase gene and
therefore overproduces dam methylase, there is no undermethylation of dam sites in either of
the extrachromosomal DNAs. The sites that are methylated by the mec methylase in both plasmid
and lambda phage DNAs were undermethylated in the dam overproducer as well. These results
indicate that the intracellular level of the E. coli methylase determines the DNA methylation
pattern.

<>

<1>Szyf, M., Bhattacharya, S., Ramchandani, S.
<2>DNA demethylase, therapeutic and diagnostic uses thereof.
<3>International Patent Office
<4>WO 9923583, WO 9924583
<5>
<6>1999
<7>The present invention relates to a DNA demethylase enzyme having about 40 KDa, and wherein
said DNA demethylase enzyme is overexpressed in cancer cells and not in normal cells.  The
present invention also relates to the therapeutic and diagnostic uses of the DNA demethylase,

<>

<1>Szyf, M., Bigey, P.
<2>Specific inhibitors of DNA methyltransferase enzyme.
<3>International Patent Office
<4>WO 9744346
<5>
<6>1997
<7>The invention provides novel mechanism-based inhibitors of DNA methyltransferase enzyme, and
diagnostic and therapeutic uses for the same.  The novel inhibitors according to the invention
form stable, noncovalent complexes with DNA methyltransferase enzyme in a manner which is
independent of S-adenosylmethionine.

<>

<1>Szyf, M., Bigey, P.
<2>Oligonucleotides as inhibitors of DNA methyltransferase: novel antitumor drugs.
<3>Curr. Res. Mol. Ther.
<4>1
<5>93-101
<6>1998
<7>DNA MeTase is now emerging as an important new therapeutic target.  The recent understanding
of its critical role in development of cancer leads us to propose that modulation of this
enzyme will be of therapeutic value.  The hairpin oligonucleotide inhibitors discussed in this
review are, to our knowledge, the first representatives of novel DNA MeTase inhibitors that
are bona fide substrate analogs.  They also constitute a new example of the use of DNA as
antagonists of natural ligands of nuclear proteins.

<>

<1>Szyf, M., Bigey, P.
<2>Specific inhibitors of DNA methyltransferase.
<3>US Patent Office
<4>US 6268137 B
<5>
<6>2001
<7>The invention provides novel mechanism-based inhibitors of DNA methyltransferase enzyme, and
diagnostic and therapeutic uses for the same.  The novel inhibitors according to the invention
form stable, noncovalent complexes with DNA methyltransferase enzyme in a manner which is
independent of S-adenosylmethionine.

<>

<1>Szyf, M., Bigey, P.
<2>New inhibitors of DNA methyltransferase enzyme, useful for inhibiting tumorigenesis - DNA-methylase-inhibitor for use in tumor gene therapy.
<3>US Patent Office
<4>US 20010041337
<5>
<6>2001
<7>NOVELTY - Inhibitors of DNA methyltransferase enzyme are new. DETAILED DESCRIPTION -
Inhibitors of DNA methyltransferase (MTase)
enzyme of formula (I) or (II), (III) or (IV) is new. N = any nucleotide;
n, y = 0 - 20; C = 5-methylcytidine; G = guanidine; L = a linker; D = a
nucleotide that is complementary to an N such that Watson-Crick base
pairing takes place between the D and N such that the Nn-C-G-Ny and
Dn-G-B-Dy form a double helix; B = cytosine, inosine, uridine,
5-bromocytosine or 5-fluorocytosine, or a basic deoxyribose; and X = an
antisense oligonucleotide of about 10 - 50 nucleotides in length which is
complementary to a portion of an RNA encoding DNA MTase enzyme. The
linkage between B and G is a phosphorothioate or phosphorodithioate
linkage, dotted lines between nucleotides represent hydrogen bonding
between the nucleotides, and the total number of nucleotides ranges from
about 10 - 50. An INDEPENDENT CLAIM is also included for a diagnostic
method for determining whether a particular sample of cells is cancerous,
involving preparing an extract from the cells in the cell sample, adding
the labeled inhibitor, measuring the extent of formation of a complex
between the labeled inhibitor and DNA MTase enzyme, normalizing the level
of such complex formation to the number of cells represented in the sample
to obtain a normalized complex formation value, and comparing the
normalized complex formation value to a normalized complex formation value
for non-cancerous and/or cancerous cell samples. BIOTECHNOLOGY - Preferred
Inhibitor: The inhibitor of DNA MTase enzyme comprises at least one
internucleoside linkage selected from phosphodiester, phosphotriester,
phosphorothioate and/or phosphoramidate (preferably phosphorothioate
internucleoside linkage). Preferred Component: The extract is preferably a
nuclear extract. ACTIVITY - Cytostatic. MECHANISM OF ACTION -
Tumorigenesis inhibitor; Methyltransferase enzyme inhibitor. Nuclear
extracts were prepared from mid-log phase human H446 cells, human A549
cells or mouse Y1 cells (1x108). The cells were harvested and washed with
phosphate buffered saline (PBS), then the cell pellet was re-suspended in
0.5 ml buffer A (tris HCl pH 8.0 (10 mM), MgCl2 (1.5 mM), KCl2 (5 mM), DTT
(0.5 mM), PMSF (0.5 mM) and Nonidet P40 (0.5%)) to separate the nuclei
from other cell components. The nuclei were pelleted by centrifugation in
an eppendorf microfuge at 2000 revolution per minute (rpm) for 15 minutes
at 4degreesC. The nuclei were then washed once in buffer A and repelleted,
and resuspended in 0.5 ml buffer B (tris HCl pH 8.0 (20 mM), glycerol
(0.25%), MgCl2 (1.5 mM), PMSF (0.5 mM), EDTA (0.2 mM), DTT (0.2 mM) and
NaCl (0.4 mM)). The resuspended nuclei were incubated on ice for 15
minutes, and then spun at 15,000 rpm to pellet nuclear debris. The nuclear
extract in the supernatant was separated from the pellet and used for
assaying DNA MTase activity. For the assay, carried out in triplicate,
nuclear extract (3 mg) was added in a reaction mixture containing a
synthetic 33-base pair hemimethylated DNA molecule substrate (0.1 mg) with
S-(methyl-3H) adenosyl-L-methionine (0.5 approximatelyCi) as the methyl
donor in a buffer containing tris HCl (20 mM) pH 7.4, EDTA (10 mM),
glycerol (25%), PMSF (0.2 mM) and 2-mercaptoethanol (20 mM). The reaction
mixture was incubated for 1 hour at 37degreesC to measure the initial rate
of the DNA MTase. For the test assay, the assay was repeated in the
presence of DNA methyltransferase inhibitor (A) containing UG dinucleotide
and bromocytosine G or 5-fluorocytosine G dinucleotide opposite the
5-methyl CG nucleotide (1 - 1000 nM). The reaction was stopped by adding
10% TCA to precipitate the DNA, which were then counted in a beta
scintillation counter. The controls were DNA incubated in the reaction
mixture in the absence of nuclear extract, and the nuclear extract
incubated in the reaction mixture in the absence of DNA. The control
inhibitor had the same nucleotide sequence as the test

<>

<1>Szyf, M., Bigey, P., Ramchandani, S.
<2>DNA methyltransferase genomic sequences and antisense oligonucleotides.
<3>International Patent Office
<4>WO 9854313
<5>
<6>1998
<7>The invention provides recombinant nucleic acids comprising nucleic acid sequences from the
genomic DNA methyltransferase gene.  The invention further provides sequence information for
such nucleic acid sequences.  In addition, the invention provides antisense oligonucleotides
complementary to special regions of the genomic DNA methyltransferase gene or its RNA
transcript.  Finally, the invention provides methods for using such antisense oligonucleotides
as analytical and diagnostic tools, as potentiators of transgenic plant and animal studies and
gene therapy approaches, and as potential therapeutic agents.

<>

<1>Szyf, M., Bigey, P., Ramchandani, S.
<2>DNA methyltransferase genomic sequences and antisense oligonucleotides.
<3>US Patent Office
<4>US 6020318
<5>
<6>2000
<7>The invention provides recombinant nucleic acids comprising nucleic acid sequences from the
genomic DNA methyltransferase gene. The
invention further provides sequence information for such nucleic acid
sequences. In addition, the invention provides antisense
oligonucleotides complementary to special regions of the genomic DNA
methyltransferase gene or its RNA transcript. Finally, the invention
provides methods for using such antisense oligonucleotides as
analytical and diagnostic tools, as potentiators of transgenic plant
and animal studies and gene therapy approaches, and as potential
therapeutic agents.

<>

<1>Szyf, M., Bigey, P., Ramchandani, S.
<2>DNA methyltransferase genomic sequences and antisense oligonucleotides.
<3>US Patent Office
<4>US 6221849
<5>
<6>2001
<7>The invention provides recombinant nucleic acids comprising nucleic acid sequences from the
genomic DNA methyltransferase gene. The
invention further provides sequence information for such nucleic acid
sequences. In addition, the invention provides antisense
oligonucleotides complementary to special regions of the genomic DNA
methyltransferase gene or its RNA transcript. Finally, the invention
provides methods for using such antisense oligonucleotides as
analytical and diagnostic tools, as potentiators of transgenic plant
and animal studies and gene therapy approaches, and as potential
therapeutic agents.

<>

<1>Szyf, M., Bigey, P., Ramchandani, S.
<2>DNA methyltransferase genomic sequences and antisense.
<3>Japanese Patent Office
<4>JP 2002512508 A
<5>
<6>2002
<7>
<>

<1>Szyf, M., Gruenbaum, Y., Urieli-Shoval, S., Razin, A.
<2>Studies on the biological role of DNA methylation:  V. The pattern of E. coli DNA methylation.
<3>Nucleic Acids Res.
<4>10
<5>7247-7259
<6>1982
<7>The distribution of the methylatable sites GATC and CCA/TGG was studied by analyzing the
molecular average size of restriction fragments of E. coli DNA. Both sites were found to be
randomly distributed, reflecting a random pattern of methylation. The methylation pattern of
specific sequences such as the origin of replication and rRNA genes has been studied in wild
type E. coli and a methylation deficient (dam- dcm-) mutant. These sequences were found to be
methylated in wild type cells and unmethylated in the mutant indicating that there is no
effect of the state of methylation of these sequences on their expression. Analysis of the
state of methylation of GATC sites in newly replicating DNA using the restriction enzyme DpnI
(cleaves only when both strands are methylated) revealed no detectable hemimethylated DNA
suggesting that methylation occurs at the replication fork. Taking together the results
presented here and previously published data, we arrive at the conclusion that the most likely
function of E. coli DNA methylations is probably in preventing nuclease activity.

<>

<1>Szyf, M., Knox, D.J., Milutinovic, S., Slack, A.D., Araujo, F.D.
<2>How does DNA methyltransferase cause oncogenic transformation?
<3>Ann. NY Acad. Sci.
<4>910
<5>156-174
<6>2000
<7>Global hypomethylation of genes and repetitive sequences, as well as hypermethylation of
certain genes known to be involved in tumor suppression, are observed concurrently in cancer
cells. Aberrant expression of DNA methyltransferase 1 (dnmt1) is a downstream effector of
multiple tumorigenic pathways, and several data suggest that dnmt1 plays a causal role in
these pathways. These data raise two critical questions: Why does ectopic expression of dnmt1
transform cells? and How can global hypomethylation exist in a cell that bears high levels of
DNMT1 activity? It is proposed that DNMT1 induces cellular transformation by a mechanism that
does not involve DNA methylation and that the low level of methylation in cancer cells is a
result of induction of a DNA demethylase in these cells. Both DNMT1 and the demethylase play a
causal role in cellular transformation and are candidate anticancer targets.

<>

<1>Szyf, M., Meisels, E., Razin, A.
<2>Biological role of DNA methylation:  sequence-specific single-strand breaks associated with hypomethylation of GATC sites in Escherichia coli DNA.
<3>J. Bacteriol.
<4>168
<5>1487-1490
<6>1986
<7>The effect of methylation of GATC sites in Escherichia coli DNA on the formation of
single-strand breaks was studied with dam+, dam mutant, and Dam-overproducer strains.
Single-strand breaks have been observed in dam mutant cells predominantly at TpT and, to a
lesser extent, at CpC. In dam mutant cells harboring pTP166 (a plasmid containing the dam
gene), no such nicks were observed.

<>

<1>Szyf, M., Ramchandani, S., MacLeod, R., Croteau, S., von Hofe, E.
<2>Antisense oligonucleotides directed against DNA methyltransferase inhibit tumorigenesis.
<3>Proc. Amer. Assoc. Cancer Res.
<4>37
<5>353
<6>1996
<7>We have previously shown that DNA methyltransferase is a downstream
effector
of the Ras oncogenic signaling pathway.  To test the hypothesis that inhibition of DNA
MeTase
could reverse tumorigenesis and to test the possibility  that DNA MeTase could serve as an
anticancer target, we have treated a number of human and mouse cancer cell lines with
antisense
phosphorothioate modified oligonucleotides directed at the translation initiation region of
either
human or mouse DNA MeTase mRNA as well as control oligonucleotides.  The cell lines
tested
were Y1, a mouse adrenocortical cell carcinoma, NCI-H596, a human lung
adenosquamous
carcinoma, NCI-H496, a lung small cell carcinoma nd SF126 a human glioma.  The
antisense
oligonucleotides inhibited DNA MeTase mRNA, DNA MeTase activity as well as DNA
methylation in a dose dependent manner (0.1-20 uM).  Focus formation in soft agar was
dramatically inhibited by DNA MeTase antisense oligonucleotides.  Tumor formation in
nude mice
(the NCI-H446 line) was inhibited when the cells expressed an exogenously introduced
antisense
DNA MeTase RNA whereas control cells formed tumors.  Injection of DNA MeTase
antisense
oligonucleotides (ip) (0.55-30 mg/kg every 48 hours) delayed tumor formation by NCI-
H446 cells
in nude mice and by Y1 cells in syngeneic LAF-1 mice.  These studies support the
hypothesis that
inhibition of DNA MeTase could reverse tumorigenesis and that DNA MeTase is a target
for
anticancer therapy.

<>

<1>Szyf, M., Theberge, J.
<2>Mammalian cells contain a general (CpG) DNA demethylating activity.
<3>Mol. Biol. Cell
<4>4
<5>74a
<6>1993
<7>A hallmark of vertebrate DNA is the fact that a significant fraction of the cytosines in the
dinucleotide sequence CpG is methylated generating a pattern of methylation that is site and
tissue specific. The pattern of methylation is formed during development by de novo
methylation and demethylation processes. The biochemical mechanisms that are responsible for
demethylation are unclear. To address the question whether mammalian cells contain a general
activity that can remove methyl groups from CpG sequences, we transfected an in vitro
methylated SK plasmid (wih SS1 methylase) into a mouse embryonal teratocarcinoma cell line and
analyzed the state of methylation of the transfected demethylation between one and two days
after transfection as judged by a HpaII/MspI analysis of the transfected DNA. All HpaII sites
were demethylated suggesting that the demethylase activity exhibits limited specificity under
these conditions. However, demethylation of areas that contain a low density of CpG sequences
is inefficient relative to that of CpG dense regions. The demethylase activity is limiting in
embryonal cells as determined by a dose response curve. Demethylase activity is not restricted
to embryonal cells and demethylation of plasmid DNA was observed in 10T1/2 and C2 cell lines.
Previous publications have suggested that demethylation involves a base excision repair
process rather than an active removal of methyl groups. To address this question we developed
an in vitro assay for demethylation. Using this assay we show that demethylation of methylated
SK plasmid DNA by a P19 nuclear extract does not require the presence of dNTPs or dCTP. This
implies that demethylation involves an active removal of methyl groups from DNA. Our results
demonstrate that parallel to the general DNA methylase activity, mammalian cells contain a
general DNA demethylase activity. We suggest that the specificity of demethylation during the
developmental process is determined by factors that limit the accessibility of DNA to the DNA
demethylase machinery.

<>

<1>Szyf, M., von Hofe, E.
<2>DNA methyltransferase antisense oligonucleotides.
<3>US Patent Office
<4>US 5578716
<5>
<6>1996
<7>The invention encompasses tumorigenicity-inhibiting antisense oligonucleotide sequences
complementary to mRNA or double-stranded DNA that encodes mammalian DNA methyltransferase.  It
further encompasses methods for inhibiting tumorigenicity and pharmaceutical composition
comprises the tumorigenicity-inhibiting antisense nucleotide.

<>

<1>Tabata, M., Nakajima, K., Nyarko, E.
<2>Metalloporphyrin mediated DNA cleavage by a low concentration of HaeIII restriction enzyme.
<3>J. Inorg. Biochem.
<4>78
<5>383-389
<6>2000
<7>The plasmid DNA scission by the restriction enzyme HaeIII was investigated in the presence of
tetrakis(1-methylpyridinium-4-yl)porphyrin (H2TMPyP) and its manganese(III), iron(III),
nickel(II), cobalt(III) and zinc(II) derivatives. The effect of metalloporphyrins on plasmid
DNA cleavage was ascertained by gel electrophoresis. UV-Vis absorption spectroscopy and
circular dichroism (CD) spectroscopy. In the absence of the metalloporphyrins, plasmid DNA
scission did not occur in the presence of a low concentration of HaeIII (0.2 units microL(-1))
at 37 degrees C after 1 h incubation. However, DNA cleavage occurred in the presence of the
metalloporphyrins and HaeIII (0.2 units microL(-1)) at 37 degrees C after 1 h incubation. Gel
electrophoresis results indicate the catalytic effect of metalloporphyrins (Mn(III)-,
Fe(III)-, Co(III)- and Zn(II)TMPyP) by binding to both DNA and the enzyme through
electrostatic interaction, which was confirmed by the change in UV-Vis and CD spectra. A
mechanism for the enhanced DNA cleavage is proposed.

<>

<1>Tabata, M., Ohtsubo, Y., Ohhata, S., Tsuda, M., Nagata, Y.
<2>Complete Genome Sequence of the gamma-Hexachlorocyclohexane-Degrading Bacterium Sphingomonas sp. Strain MM-1.
<3>Genome Announcements
<4>1
<5>e00247-13
<6>2013
<7>gamma-Hexachlorocyclohexane (gamma-HCH) is a man-made chlorinated insecticide that has caused
serious environmental problems. Here, we report the complete
genome sequence of the gamma-HCH-degrading bacterium Sphingomonas sp. strain
MM-1, which consists of one chromosome and five plasmids. All the specific lin
genes that are almost identical to those of Sphingobium japonicum UT26 for the
conversion of gamma-HCH to beta-ketoadipate are dispersed on four out of the five
plasmids.

<>

<1>Tada, T., Kitao, T., Miyoshi-Akiyama, T., Kirikae, T.
<2>Genome Sequence of Multidrug-Resistant Pseudomonas aeruginosa NCGM1179.
<3>J. Bacteriol.
<4>193
<5>6397
<6>2011
<7>We report the annotated genome sequence of multidrug-resistant Pseudomonas aeruginosa strain
NCGM1179, which is highly resistant to carbapenems,
aminoglycosides, and fluoroquinolones and is emerging at medical
facilities in Japan.

<>

<1>Tae, H., Shallom, S., Settlage, R., Hawkins, G.N., Adams, L.G., Garner, H.R.
<2>Complete Genome Sequence of Brucella suis VBI22, Isolated from Bovine Milk.
<3>J. Bacteriol.
<4>194
<5>910
<6>2012
<7>Brucella suis is the causative agent of swine brucellosis and is known to be able to infect
several different hosts, including cattle, dogs, and
horses, without causing disease symptoms. Here we report the complete
genome sequence of Brucella suis VBI22, which was isolated from raw milk
from an infected cow.

<>

<1>Tae, H., Shallom, S., Settlage, R., Preston, D., Adams, L.G., Garner, H.R.
<2>Revised Genome Sequence of Brucella suis 1330.
<3>J. Bacteriol.
<4>193
<5>6410
<6>2011
<7>Brucella suis is a causative agent of porcine brucellosis. We report the resequencing of the
original sample upon which the published sequence of
Brucella suis 1330 is based and describe the differences between the
published assembly and our assembly at 12 loci.

<>

<1>Tagami, H., Tayama, K., Fukaya, M., Okumura, H., Masai, H.
<2>Restriction enzyme manufacture by culturing Acetobacter species bacterium.
<3>Japanese Patent Office
<4>JP 62011091
<5>
<6>1987
<7>Type II restriction enzyme, AaaI, producing bacterium which belongs to Acetobacter sp. is
cultured, and from the cultured cell Type II restriction enzyme is collected. For cultivation
of bacterium, the bacterium is inoculated on polypeptone, yeast extract, glucose containing
medium and spinner cultured at 25-35 deg.C for 24-48 hrs. aerobically. After culture, cells
are collected by centrifugation, enzyme is separated and purified.

<>

<1>Tagami, H., Tayama, K., Tohyama, T., Fukaya, M., Okumura, H., Kawamura, Y., Horinouchi, S., Beppu, T.
<2>Purification and properties of a site-specific restriction endonuclease AaaI from Acetobacter aceti subsp. aceti No. 1023.
<3>FEMS Microbiol. Lett.
<4>56
<5>161-166
<6>1988
<7>A type II restriction endonuclease, named AaaI, was purified from Acetobacter
aceti subsp. aceti No. 1023.  The optimum pH and temperature were determined to
be 8.5 and 37C, respectively.  The enzyme activity was stimulated by the
addition of either NaCl or KCl and their optimum concentrations were 100 mM for
both cations.  AaaI recognized the hexanucleotide sequence
5'-C^GGCCG-3'/GCCGG^C and cleaved it at the positions indicated by the arrows.
AaaI is an isoschizomer of XmaIII from Xanthomonas malvacearum and Eco52I from
Escherichia coli.

<>

<1>Taghavi, S., Izquierdo, J.A., van der Lelie, D.
<2>Complete Genome Sequence of Clostridium sp. Strain DL-VIII, a Novel Solventogenic Clostridium Species Isolated from Anaerobic Sludge.
<3>Genome Announcements
<4>1
<5>e00605-13
<6>2013
<7>We report the genome sequence of Clostridium sp. strain DL-VIII, a novel Gram-positive,
endospore-forming, solventogenic bacterium isolated from activated
anaerobic sludge of a wastewater treatment plant. Aside from a complete sol
operon, the 6,477,357-bp genome of DL-VIII reveals genes for several unique
enzymes with applications in lignocellulose degradation, including two phenolic
acid decarboxylases.

<>

<1>Tahrioui, A., Quesada, E., Llamas, I.
<2>Draft Genome Sequence of the Moderately Halophilic Gammaproteobacterium Halomonas anticariensis FP35T.
<3>Genome Announcements
<4>1
<5>e00497-13
<6>2013
<7>Halomonas anticariensis strain FP35(T) is a moderately halophilic bacterium isolated from a
soil sample taken from Fuente de Piedra, a saline wetland in the
province of Malaga (Spain), which produces an exopolysaccharide and
quorum-sensing signaling molecules of the type N-acylhomoserine lactone. We
report here the draft genome sequence of this gammaproteobacterium.

<>

<1>Taira, E., Iiyama, K., Mon, H., Mori, K., Akasaka, T., Tashiro, K., Yasunaga-Aoki, C., Lee, J.M., Kusakabe, T.
<2>Draft Genome Sequence of Entomopathogenic Serratia liquefaciens Strain FK01.
<3>Genome Announcements
<4>2
<5>e00609-14
<6>2014
<7>In the present study, we determined the draft genome sequence of the entomopathogenic
bacterium Serratia liquefaciens FK01, which is highly virulent
to the silkworm. The draft genome is ~5.28 Mb in size, and the G+C content is
55.8%.

<>

<1>Tajima, S.
<2>DNA methyltransferase and DNA methylation.
<3>Gendai Iryo
<4>35
<5>936-948
<6>2003
<7>
<>

<1>Tajima, S., Tsuda, H., Wakabayashi, N., Asano, A., Mizuno, S., Nishimori, K.
<2>Isolation and expression of a chicken DNA methyltransferase cDNA.
<3>J. Biochem. (Tokyo)
<4>117
<5>1050-1057
<6>1995
<7>A 0.5 kb fragment of chicken DNA methyltransferase cDNA was PCR-amplified using a set of
degenerate primers.  A clone harboring a 5 kb insert was isolated from a cDNA library by
screening with the PCR-amplified cDNA fragment as a probe.  The elucidated nucleotide sequence
gave a 4,614 nucleotide open reading frame, and the predicted protein was highly homologous to
the mouse and human DNA methyltransferases, especially in  the amino acid sequence of the
catalytic domain in the carboxyl-terminal region.  The cysteine-rich region and Lys-Gly repeat
first found in the mouse sequence were also conserved in chicken.  However, about 250 amino
acid residues in the amino-terminal portion of chicken DNA methyltransferase diverged from the
amino-terminus of the mouse or human sequence.  Northern blot analysis showed that the message
of chicken DNA methyltransferase was expressed at high levels in the testis, in the lung and
in Marek's virus-transformed chicken T-lymphoma cells.  Expression of the chicken DNA
methyltransferase is COS1 cells demonstrated that the enzyme is a so-called maintenance type
methylase.  When poly(dG-dC)-poly(dG-dC) was used as the methyl acceptor, to provide a measure
of de novo methylase activity, the Km value for S-adenosyl-L-methionine was about 5 microM,
which was 10 times higher than that when poly(dI-dC)-poly(dI-dC) was used.  The affinity of
DNA methyltransferase for S-adenosyl L-methionine in catalyzing de novo-type methylation
activity  was lower than that in catalyzing maintenance-type activity, though it was still
high enough for the enzyme to work as a de novo-type methylase under physiological conditions.

<>

<1>Tak, N. et al.
<2>Genome sequence of Ensifer sp. TW10; a Tephrosia wallichii (Biyani) microsymbiont native to the Indian Thar Desert.
<3>Standards in Genomic Sciences
<4>9
<5>304-314
<6>2013
<7>Ensifer sp. TW10 is a novel N2-fixing bacterium isolated from a root nodule of the perennial
legume Tephrosia wallichii Graham (known locally as Biyani) found
in the Great Indian (or Thar) desert, a large arid region in the northwestern
part of the Indian subcontinent. Strain TW10 is a Gram-negative, rod shaped,
aerobic, motile, non-spore forming, species of root nodule bacteria (RNB) that
promiscuously nodulates legumes in Thar Desert alkaline soil. It is fast growing,
acid-producing, and tolerates up to 2% NaCl and capable of growth at 40(o)C. In
this report we describe for the first time the primary features of this Thar
Desert soil saprophyte together with genome sequence information and annotation.
The 6,802,256 bp genome has a GC content of 62% and is arranged into 57 scaffolds
containing 6,470 protein-coding genes, 73 RNA genes and a single rRNA operon.
This genome is one of 100 RNB genomes sequenced as part of the DOE Joint Genome
Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria
(GEBA-RNB) project.

<>

<1>Takagi, M., Nishioka, M., Kakihara, H., Kitabayashi, M., Inoue, H., Kawakami, B., Oka, M., Imanaka, T.
<2>Characterization of DNA polymerase from Pyrococcus sp. strain KOD1 and its application to PCR.
<3>Appl. Environ. Microbiol.
<4>63
<5>4504-4510
<6>1997
<7>The DNA polymerase gene from the archaeon Pyrococcus sp. strain KOD1 (DOD DNA polymerase)
contains a long open reading frame of 5,013 bases that encodes 1,671 amino acid residues.
Similarity analysis revealed that the DNA polymerase contained a putative 3'-5' exonuclease
activity and two in-frame intervening sequences of 1,080 bp (360 amino acids; KOD pol
intein-1) and 1,611 bp (537 amino acids; KOD pol intein-2), which are located in the middle of
regions conserved among eukaryotic and archaeal a-like DNA polymerases.  The mature form of
the DNA polymerase gene was expressed in Escherichia coli, and the recombinant enzyme was
purified and characterized.  3'-5' exonuclease activity was confirmed, and although KOD DNA
polymerase's optimum temperature (75 C) and mutation frequency (3.5 x 10^-3) were similar to
those of a DNA polymerase from Pyrococcus furiosus (Pfu DNA polymerase), the KOD DNA
polymerase exhibited an extension rate (100 to 130 nucleotides/s) 5 times higher than those of
Pfu DNA polymerase.  These characteristics enabled the KOD DNA polymerase to perform a more
accurate PCR in a shorter reaction time.

<>

<1>Takahashi, H., Kojima, H., Saito, H.
<2>A new site-specific endonuclease, ScaI, from Streptomyces caespitosus.
<3>Biochem. J.
<4>231
<5>229-232
<6>1985
<7>A new site-specific endonuclease has been isolated from Streptomyces
caespitosus and named ScaI.  Based on analysis of sequences around the
restriction sites in pBR322 and pBR325, the recognition sequence of ScaI
endonuclease was deduced to be a new hexanucleotide 5'-AGTACT-3'.  The cleavage
site was determined by comparing the ScaI-cleaved roduct of a primer-extended
M13mp18-SCA DNA, which contains an AGTACT sequence, with dideoxy chain
terminator ladders of the same DNA.  ScaI was found to cleave the recognition
sequence between the internal T and A, leaving flush ends to the cleaved
fragments.

<>

<1>Takahashi, H., Saito, H., Ikeda, Y.
<2>Viable T4 bacteriophage containing cytosine-substituted DNA (T4dC phage).  I.  Behavior towards the restriction-modification systems of Escherichia coli and derivation of a new T4 phage strain (T4dC) having the complete T4 genome.
<3>J. Gen. Appl. Microbiol.
<4>24
<5>297-306
<6>1978
<7>A multiple mutant of bacteriophage T4, which had been proved to contain
cytosine-substituted DNA, was grown in a suppressor-containing or
non-containing (supo) strain of Escherichia coli B or K12, and the progeny
phage obtained was grown on test strains carrying various restriction systems.
As a result, the growth of this phage was found to be subject to the rK, rB,
and rP1 restriction systems.  Two T4 strains, named UNF-r1 and UNF-r+1,
sensitive to the rK, rB, and rP1 restriction systems, were newly derived by the
genetic cross and mutagenesis.  The former had the unf-39 mutation instead of
the alc-8 mutation, the latter had the rII+ region and the unf-39 mutation,
besides the amE51 (gene 56; dCTPase), denA (endonuclease II), and denB
(endonuclease IV) mutations.  It was demonstrated that UNF-r+1 DNA possessed a
SalI recognition site in the rII-denB region which was deleted in UNF-r1 DNA.

<>

<1>Takahashi, H., Shao, M., Furuya, N., Komano, T.
<2>The genome sequence of the incompatibility group Igamma plasmid R621a: Evolution of IncI plasmids.
<3>Plasmid
<4>66
<5>112-121
<6>2011
<7>We present the complete genome sequence of the tetracycline resistance plasmid R621a isolated
from Salmonella typhimurium, which belongs to the incompatibility group Igamma. In the
93,185bp circular double-stranded R621a genome, 96 complete ORFs are predicted. In addition,
one and six different kinds of proteins are produced by translational reinitiation and
shufflon multiple inversions, respectively. The genome consists of four regions: replication,
leading, transfer, and miscellaneous regions. The R621a genome is similar to those of IncI1
plasmids such as R64 and ColIb-P9 and particularly to those of pEK204 and pEC_Bactec. Three
major differences including inc, parAB, and excA regions were noted between R621a and
prototype IncI1 plasmids. Seven nucleotide replacements and one nucleotide deletion in the
putative Inc RNA sequence are found between R621a and IncI1 plasmids irrespective of close
similarity in the other parts of the rep system. The sequences of R621a parAB and excA genes
are significantly different from those of R64 and ColIb-P9, while those of R621a parAB and
excA genes exhibit close similarity to those of pEK204 and pEC_Bactec, respectively. The R621a
genome is suggested to be formed by acquiring parAB and excA genes from pEK204 and pEC_Bactec
genomes, respectively, and then novel inc function by the mutations. The insertions in the
R621a, pEK204, and pEC_Bactec genomes are flanked by direct repeats, suggesting that
insertions accompanied by long target duplications have also played an important role in the
evolution of IncI plasmids.

<>

<1>Takahashi, H., Shimizu, M., Saito, H., Ikeda, Y., Sugisaki, H.
<2>A new site-specific endonuclease from Streptomyces lavendulae (SlaI) .
<3>Gene
<4>5
<5>9-18
<6>1979
<7>A new restriction-like endonuclease, SlaI, was found and partially purified
from Streptomyces lavendulae ATCC8664.  This endonuclease cleaved bacteriophage
lambda DNA at only one site, and cytosine-substituted bacteriophage T4 DNA at
16 sites.  The recognition sequence was determined by using SlaI fragments of
cytosine-substituted bacteriophage T4 DNA.  The hexanucleotide recognized by
SlaI endonuclease was 5'-C^T-C-G-A-G-3' 3'-G-A-G-C-A^C-5' with the sites of
cleavage as indicated by the arrows.  Therefore, SlaI endonuclease was an
isoschizomer of XhoI endonuclease.

<>

<1>Takahashi, I., Hayano, D., Asayama, M., Masahiro, F., Watahiki, M., Shirai, M.
<2>Restriction barrier composed of an extracellular nuclease and restriction endonuclease in the unicellular cyanobacterium Microcystis sp.
<3>FEMS Microbiol. Lett.
<4>145
<5>107-111
<6>1996
<7>The unicellular cyanobacterium Microcystis aeruginosa K-81 has two types of restriction
barrier, an extracellular nuclease and sequence-specific endonucleases.  The nuclease was
detected in the culture supernatant and it was easily released from the cells by washing with
water or buffer containing Triton X-100.  This nuclease was identified as a polypeptide of
about 28 kDa that digested covalently closed circular and linear double-stranded DNAs,
including chromosomal DNA from M. aeruginosa K-81.  Among another 13 Microcystis strains
examined, 3 produced an extracellular nuclease.  Furthermore, M. aeruginosa K-81 contained two
sequence-specific endonucleases, MaeK81I and MaeK81II, which were isoschizomers of SplI and
Sau96I, respectively.

<>

<1>Takahashi, M., Kobayashi, M., Uchida, T.
<2>Molecular multiplicity of nuclease TT1 from Thermus thermophilus HB8.
<3>J. Biochem. (Tokyo)
<4>90
<5>1521-1527
<6>1981
<7>Purified nuclease TT1 from Thermus thermophilus HB8 has multimolecular weight
forms, each of which is composed of three different subunits, a (10.8 x 104), b
(7.8 x 104), and c (4.1 x 104).  The molecular weight of this enzyme were
estimated by gel filtration, polyacrylamide gel electrophoresis and equilibrium
sedimentation.  It was found that most of the enzyme has a molecular weight of
about 22 x 104 being a monomer having the subunit composition of abc.  The
remaining part of the enzyme has larger molecular weights and is considered to
be size-isomers of abc.  The alpha-helical content, 5.5-6.5%, and the
b-structure, about 28% were estimated from the CD spectrum at 4C.

<>

<1>Takahashi, N., Naito, Y., Handa, N., Kobayashi, I.
<2>A DNA methyltransferase can protect the genome from postdisturbance attack by a restriction-modification gene complex.
<3>J. Bacteriol.
<4>184
<5>6100-6108
<6>2002
<7>In prokaryotic genomes, some DNA methyltransferases form a restriction-modification gene
complex, but some others are present by themselves. Dcm gene product, one of these orphan
methyltransferases found in Escherichia coli and related bacteria, methylates DNA to generate
5'-CmCWGG just as some of its eukaryotic homologues do. Vsr mismatch repair function of an
adjacent gene prevents C-to-T mutagenesis enhanced by this methylation but promotes other
types of mutation and likely has affected genome evolution. The reason for the existence of
the dcm-vsr gene pair has been unclear. Earlier we found that several restriction-modification
gene complexes behave selfishly in that their loss from a cell leads to cell killing through
restriction attack on the genome. There is also increasing evidence for their potential
mobility. EcoRII restriction-modification gene complex recognizes the same sequence as Dcm,
and its methyltransferase is phylogenetically related to Dcm. In the present work, we found
that stabilization of maintenance of a plasmid by linkage of EcoRII gene complex, likely
through postsegregational cell killing, is diminished by dcm function. Disturbance of EcoRII
restriction-modification gene complex led to extensive chromosome degradation and severe loss
of cell viability. This cell killing was partially suppressed by chromosomal dcm and
completely abolished by dcm expressed from a plasmid. Dcm, therefore, can play the role of a
'molecular vaccine' by defending the genome against parasitism by a restriction-modification
gene complex.

<>

<1>Takahashi, N., Ohashi, S., Sadykov, M.R., Mizutani-Ui, Y., Kobayashi, I.
<2>IS-Linked Movement of a Restriction-Modification System.
<3>PLoS ONE
<4>6
<5>e16554
<6>2011
<7>Potential mobility of restriction-modification systems has been suggested by
evolutionary/bioinformatic analysis of prokaryotic
genomes. Here we demonstrate in vivo movement of a
restriction-modification system within a genome under a laboratory
condition. After blocking replication of a temperature-sensitive
plasmid carrying a PaeR7I restriction-modification system in
Escherichia coli cells, the plasmid was found integrated into the
chromosome of the surviving cells. Sequence analysis revealed that, in
the majority of products, the restriction-modification system was
linked to chromosomal insertion sequences (ISs). Three types of
products were: (I) apparent co-integration of the plasmid and the
chromosome at a chromosomal IS1 or IS5 copy (24/28 analyzed); (II) de
novo insertion of IS1 with the entire plasmid except for a 1-3 bp
terminal deletion (2/28); and (III) reciprocal crossing-over between
the plasmid and the chromosome involving 1-3 bp of sequence identity
(2/28). An R-negative mutation apparently decreased the efficiency of
successful integration by two orders of magnitude. Reconstruction
experiments demonstrated that the restriction-dependence was mainly due
to selection against cells without proper integration: their growth was
inhibited by the restriction enzyme action. These results demonstrate
collaboration of a mobile element and a restriction-modification system
for successful joint migration. This collaboration may have promoted
the spread and, therefore, the long-term persistence of these complexes
and restriction-modification systems in a wide range of prokaryotes.

<>

<1>Takahashi, S., Matsuno, H., Furusawa, H., Okahata, Y.
<2>Direct monitoring of EcoRII restriction enzyme reactions on a 27MHz quartz-crystal microbalance.
<3>Nucleic Acids Res. Suppl.
<4>2
<5>71-72
<6>2002
<7>EcoRII is a type IIE restriction endonuclease characterized by a highly cooperative reaction
mechanism that depends on simultaneous binding of the dimeric enzyme molecule to two copies of
its recognition site.  Gaining insights about reaction property of EcoRII binding to its
recognition site and effect of divalent metal ion on DNA cleavage process, a DNA-immobilized
quartz crystal microbalance, which enables real-time monitoring both the binding of enzyme and
the cleavage reaction on DNA strands as mass changes, was utilized.

<>

<1>Takahashi, S., Matsuno, H., Furusawa, H., Okahata, Y.
<2>Direct monitoring of allosteric recognition of type IIE restriction endonuclease EcoRII.
<3>J. Biol. Chem.
<4>283
<5>15023-15030
<6>2008
<7>EcoRII is a homodimer with two domains consisting of a DNA-binding N terminus and a catalytic
C terminus and recognizes two specific
sequences on DNA. It shows a relatively complicated cleavage reaction
in bulk solution. After binding to either recognition site, EcoRII
cleaves the other recognition site of the same DNA (cis-binding) strand
and/or the recognition site of the other DNA (trans-binding) strand.
Although it is difficult to separate these two reactions in bulk
solution, we could simply obtain the binding and cleavage kinetics of
only the cis-binding by following the frequency (mass) changes of a
DNA-immobilized quartz-crystal microbalance (QCM) responding to the
addition of EcoRII in aqueous solution. We obtained the maximum binding
amounts (Delta m(max)) the dissociation constants (K-d), the binding
and dissociation rate constants (k(on) and k(off)), and the catalytic
cleavage reaction rate constants (k(cat)) for wildtype EcoRII, the
N-terminal-truncated form (EcoRII N-domain), and the mutant derivatives
in its C-terminal domain (K263A and R330A). It was determined from the
kinetic analyses that the N-domain, which covers the catalytic C-domain
in the absence of DNA, preferentially binds to the one DNA recognition
site while transforming EcoRII into an active form allosterically, and
then the secondary C-domain binds to and cleaves the other recognition
site of the DNA strand.

<>

<1>Takai, D.
<2>Detection of DNA methylation using methylation sensitive restriction enzyme.
<3>Jikken Igaku Bessatsu
<4>JB12
<5>62-68
<6>2008
<7>
<>

<1>Takai, K., Horikoshi, K.
<2>Molecular phylogenetic analysis of archaeal intron-containing genes coding for rRNA obtained from a deep-subsurface geothermal water pool.
<3>Appl. Environ. Microbiol.
<4>65
<5>5586-5589
<6>1999
<7>Molecular phylogenetic analysis of a naturally occurring microbial community in a
deep-subsurface geothermal environment indicated that the phylogenetic diversity of the
microbial population in the environment was extremely limited and that only hyperthermophilic
archaeal members closely related to Pyrobaculum were present. All archaeal ribosomal DNA
sequences contained intron-like sequences, some of which had open reading frames with repeated
homing-endonuclease motifs. The sequence similarity analysis and the phylogenetic analysis of
these homing endonucleases suggested the possible phylogenetic relationship among archaeal
rRNA-encoded homing endonucleases.

<>

<1>Takami, H., Nakasone, K., Takaki, Y., Maeno, G., Sasaki, R., Masui, N., Fuji, F., Hirama, C., Nakamura, Y., Ogasawara, N., Kuhara, S., Horikoshi, K.
<2>Complete genome sequence of the alkaliphilic bacterium Bacillus halodurans and genomic sequence comparison with Bacillus subtilis.
<3>Nucleic Acids Res.
<4>28
<5>4317-4331
<6>2000
<7>The 4,202,353 bp genome of the alkaliphilic bacterium Bacillus halodurans C-125 contains 4066
predicted protein coding sequences (CDSs), 2141 (52.7%) of which have functional assignments,
1182 (29%) of which are conserved CDSs with unknown function and 743 (18. 3%) of which have no
match to any protein database.  Among the total CDSs, 8.8% match sequences of proteins found
only in Bacillus subtilis and 66.7% are widely conserved in comparison with the proteins of
various organisms, including B. subtilis. The B. halodurans genome contains 112 transposase
genes, indicating that transposases have played an important evolutionary role in horizontal
gene transfer and also in internal genetic rearrangement in the genome. Strain C-125 lacks
some of the necessary genes for competence, such as comS, srfA and rapC, supporting the fact
that competence has not been demonstrated experimentally in C-125. There is no paralog of
tupA, encoding teichuronopeptide, which contributes to alkaliphily, in the C-125 genome and an
ortholog of tupA cannot be found in the B. subtilis genome. Out of 11 sigma factors which
belong to the extracytoplasmic function family, 10 are unique to B. halodurans, suggesting
that they may have a role in the special mechanism of adaptation to an alkaline environment.

<>

<1>Takami, H., Takaki, Y., Chee, G.-J., Nishi, S., Shimamura, S., Suzuki, H., Matsui, S., Uchiyama, I.
<2>Thermoadaptation trait revealed by the genome sequence of thermophilic Geobacillus kaustophilus.
<3>Nucleic Acids Res.
<4>32
<5>6292-6303
<6>2004
<7>We present herein the first complete genome sequence of a thermophilic Bacillus-related
species, Geobacillus kaustophilus HTA426, which is composed of a 3.54 Mb chromosome and a 47.9
kb plasmid, along with a comparative analysis with five other mesophilic bacillar genomes.
Upon orthologous grouping of the six bacillar sequenced genomes, it was found that 1257 common
orthologous groups composed of 1308 genes (37%) are shared by all the bacilli, whereas 839
genes (24%) in the G.kaustophilus genome were found to be unique to that species. We were able
to find the first prokaryotic sperm protamine P1 homolog, polyamine synthase, polyamine ABC
transporter and RNA methylase in the 839 unique genes; these may contribute to thermophily by
stabilizing the nucleic acids. Contrasting results were obtained from the principal component
analysis (PCA) of the amino acid composition and synonymous codon usage for highlighting the
thermophilic signature of the G. kaustophilus genome. Only in the PCA of the amino acid
composition were the Bacillus-related species located near, but were distinguishable from, the
borderline distinguishing thermophiles from mesophiles on the second principal axis. Further
analysis revealed some asymmetric amino acid substitutions between the thermophiles and the
mesophiles, which are possibly associated with the thermoadaptation of the organism.

<>

<1>Takami, H., Takaki, Y., Uchiyama, I.
<2>Genome sequence of Oceanobacillus iheyensis isolated from the Iheya Ridge and its unexpected adaptive capabilities to extreme environments.
<3>Nucleic Acids Res.
<4>30
<5>3927-3935
<6>2002
<7>Oceanobacillus iheyensis HTE831 is an alkaliphilic and extremely halotolerant Bacillus-related
species isolated from deep-sea sediment. We present here the complete genome sequence of
HTE831 along with analyses of genes required for adaptation to highly alkaline and saline
environments. The genome consists of 3.6 Mb, encoding many proteins potentially associated
with roles in regulation of intracellular osmotic pressure and pH homeostasis. The candidate
genes involved in alkaliphily were determined based on comparative analysis with three
Bacillus species and two other Gram-positive species. Comparison with the genomes of other
major Gram-positive bacterial species suggests that the backbone of the genus Bacillus is
composed of approximately 350 genes. This second genome sequence of an alkaliphilic
Bacillus-related species will be useful in understanding life in highly alkaline environments
and microbial diversity within the ubiquitous bacilli.

<>

<1>Takamiya, M., Ozen, A., Rasmussen, M., Alter, T., Gilbert, T., Ussery, D.W., Knochel, S.
<2>Genome Sequences of Two Stress-Tolerant Campylobacter jejuni Poultry Strains, 305 and DFVF1099.
<3>J. Bacteriol.
<4>193
<5>5546-5547
<6>2011
<7>Campylobacter jejuni is a food-borne pathogen with a high prevalence in poultry meat, which in
fresh unfrozen condition is the major source of
campylobacteriosis. C. jejuni strains DFVF1099 and 305 are considered
tolerant to several environmental stresses (T. Birk et al., J. Food Prot.
73:258-265, 2010; S. L. On et al., Int. J. Med. Microbiol. 296:353-363,
2006). Here, we report the genome sequences of C. jejuni 305 and DFVF1099,
a turkey and a chicken isolate, respectively.

<>

<1>Takamiya, M., Ozen, A., Rasmussen, M., Alter, T., Gilbert, T., Ussery, D.W., Knochel, S.
<2>Genome Sequence of Campylobacter jejuni strain 327, a strain isolated from a turkey slaughterhouse.
<3>Standards in Genomic Sciences
<4>4
<5>113-122
<6>2011
<7>Campylobacter is one of the leading causes of food-borne gastroenteritis
and has a high prevalence in poultry. Campylobacter jejuni subsp. jejuni
327 is a subspecies of the genus Campylobacter of the family
Campylobacteraceae in the phylum Proteobacteria. The microaerophilic,
spiral shaped, catalase positive bacterium obtains energy from the
metabolism of amino acids and Krebs cycle intermediates. Strain 327 was
isolated from a turkey slaughter production line and is considered
environmentally sensitive to food processing (cold, heat, drying) and
storage conditions. The 327 whole genome shotgun sequence of 1,618,613 bp
long consists of 1,740 protein-coding genes, 46 tRNA genes and 3 rRNA
operons. A protein based BLAST analysis places the turkey isolate 327
close to the human clinical strain 81116 (NCTC 11828).

<>

<1>Takanami, M.
<2>Specific cleavage of coliphage fd DNA by five different restriction endonucleases from Haemophilus genus.
<3>FEBS Lett.
<4>34
<5>318-322
<6>1973
<7>None

<>

<1>Takanami, M.
<2>Restriction endonucleases AP, GA, and H-1 from three Haemophilus strains.
<3>Methods Mol. Biol.
<4>7
<5>113-133
<6>1974
<7>None

<>

<1>Takanami, M., Kojo, H.
<2>Cleavage site specificity of an endonuclease prepared from Haemophilus influenzae strain H-1.
<3>FEBS Lett.
<4>29
<5>267-270
<6>1973
<7>None

<>

<1>Takanami, M., Okamoto, T., Sugimoto, K., Sugisaki, H.
<2>Studies on Bacteriophage fd DNA I.  A Cleavage Map of the fd Genome.
<3>J. Mol. Biol.
<4>95
<5>21-31
<6>1975
<7>In order to construct a physical map of the bacteriophage fd genome, the doubly
closed replicative form (RFI) DNA of phage fd was cleaved into unique fragments
by four different restriction endonucleases (Hap, Hga, HinH and Hind) prepared
from Haemophilus strains H. aphirophilus, H. gallinarum, H. influenzae H-I and
H. influenzae Rd, respectively.  As Hind cleaved RFI DNA at a single site, this
site was used as a reference point for mapping.  HinH cleaved RFI DNA at three
sites, Hga at six sites and Hap at 13 sites, respectively.  The 5'-termini of
the fragments produced by either HinH or Hga were labelled with 32P in the
polynucleotide kinase reaction.  The labelled fragments were separated and
further cleaved by other enzymes.  The re-digestion products of partially
digested fragments were also analysed.  On the basis of these data and
estimates of the size of each fragment, a cleavage map of the phage fd genome
was constructed.

<>

<1>Takano, T., Ochi, A., Yamamoto, N.
<2>Restriction enzyme from Lactobacillus fermentum.
<3>FEMS Microbiol. Rev.
<4>7
<5>64
<6>1990
<7>Restriction-modification is important in phage resistance. However, little is known about
restriction enzymes of lactic acid bacteria. We have found that Lactobacillus fermentum CP-34
has apparent site-specific endonuclease activity, purified and studied the characteristics of
the enzyme.

<>

<1>Takano, T., Watanabe, T., Fukasawa, T.
<2>Specific inactivation of infectious lambda DNA by sonicates of restrictive bacteria with R factors.
<3>Biochem. Biophys. Res. Commun.
<4>25
<5>192-204
<6>1966
<7>In the studies on the host-controlled modification (Arber, 1965a) of
bacteriophage lambda, certain aspects of two fundamental functions of host
bacteria have been elucidated:  Modification is a function by which viral DNA
is subjected to some chemical alterations (Arber, 1965b).  Restriction is
another function which recognizes the modification pattern or the host
specificity and serves to restrict the phage mutliplication depending on the
host specificity (Arber and Dussoix, 1962).  Analogous experiments are now
under way in the system of P1-controlled restriction in our laboratories.

<>

<1>Takano, T., Watanabe, T., Fukasawa, T.
<2>Mechanism of host-controlled restriction of bacteriophage lambda by R factors in Escherichia coli K12.
<3>Virology
<4>34
<5>290-302
<6>1968
<7>In the host-controlled modification of phage lambda by fi- R factors, N-3 and
R-15, the infectivity of the unmodified lambda DNA was specifically destroyed
by the sonicated extracts from the restrictive, endonuclease I-less (endo
I-less) mutant of Escherichia coli K12 carrying N-3 or R-15, to a greater
extent than by the sonicates from the nonrestrictive, endo I-less bacteria.  In
the control experiments, the lambda DNA modified by N-3 was inactivated to much
lesser extents both by the sonicates from the restrictive, endo I-less bacteria
and by those from the nonrestrictive, endo I-less bacteria.  In all these
systems of reactions, the phage DNA was hardly degraded into acid-soluble
forms.  The host specifically inactivated, unmodified lambda DNA, however, was
split fragmentally as was shown by the patterns of zone sedimentation, but the
modified lambda DNA was not.  The activity which destroys the infectivity of
the unmodified lambda DNA resided totally in the spheroplasts of the
restrictive bacteria.

<>

<1>Takarada, H., Sekine, M., Kosugi, H., Matsuo, Y., Fujisawa, T., Omata, S., Kishi, E., Shimizu, A., Tsukatani, N., Tanikawa, S., Fujita, N., Harayama, S.
<2>Complete genome sequence of the soil actinomycete Kocuria rhizophila.
<3>J. Bacteriol.
<4>190
<5>4139-4146
<6>2008
<7>The soil actinomycete Kocuria rhizophila belongs to the suborder Micrococcineae, a divergent
bacterial group for which only a limited
amount of genomic information is currently available. K. rhizophila is
also important in industrial applications; e.g., it is commonly used as a
standard quality control strain for antimicrobial susceptibility testing.
Sequencing and annotation of the genome of K. rhizophila DC2201 (NBRC
103217) revealed a single circular chromosome (2,697,540 bp; G+C content
of 71.16%) containing 2,357 predicted protein-coding genes. Most of the
predicted proteins (87.7%) were orthologous to actinobacterial proteins,
and the genome showed fairly good conservation of synteny with
taxonomically related actinobacterial genomes. On the other hand, the
genome seems to encode much smaller numbers of proteins necessary for
secondary metabolism (one each of nonribosomal peptide synthetase and type
III polyketide synthase), transcriptional regulation, and lateral gene
transfer, reflecting the small genome size. The presence of probable
metabolic pathways for the transformation of phenolic compounds generated
from the decomposition of plant materials, and the presence of a large
number of genes associated with membrane transport, particularly amino
acid transporters and drug efflux pumps, may contribute to the organism's
utilization of root exudates, as well as the tolerance to various organic
compounds.

<>

<1>Takasaki, Y.
<2>Two forms of restriction enzyme HindIII.
<3>J. Biochem. (Tokyo)
<4>116
<5>1281-1286
<6>1994
<7>Restriction endonuclease HindIII was purified from Haemophilus influenzae Rd. Two active
fractions, P1 and P2, were obtained in phosphocellulose chromatography. HindIII could be
purified completely from the first fraction, P1, by subsequent DEAE-cellulose chromatography.
The second fraction, P2, showed HindIII activity higher than that of P1, though it was still
contaminated with some minor proteins. The HindIII in the P2 fraction showed differences in
stability, binding to substrate DNA, electrophoretic mobility, etc., from the HindIII in the
P1 fraction. It is likely that there are two forms of HindIII in the bacterial cell. The
endonuclease HindIII in the P2 fraction was finally purified by DNA-cellulose chromatography,
though considerable loss of enzymatic activity resulted. Upon infection of the cells with
phage T4, the P2 fraction in phosphocellulose chromatography almost disappeared. The presence
of two forms of HindIII may be related to bacterial defense against viral infection.

<>

<1>Takasaki, Y.
<2>Alternation of two forms of restriction endonuclease HindIII.
<3>Biosci. Biotechnol. Biochem.
<4>60
<5>396-400
<6>1996
<7>Two forms of the restriction enzyme HindIII were alternated with each other under
some physiological or biochemical conditions.  Addition of a low amount of phage T7 to the
culture of HindIII-producing Haemophilus influenzae Rd, resulted in appearance of some amounts
of the P2 fraction of HindIII, which was eluted with a high concentration of KCl from a
phosphocellulose column.  Higher amounts of T7 caused a decrease of the P2 fraction; finally
the
alternative P1 fraction of HindIII, which was eluted with a lower concentration of KCl,
remained
exclusively.  Addition of disaccharides such as maltose and trehalose to the bacterial
extract,
yielded more P2, although the disaccharides inhibited this enzyme.  Urea showed an interesting
distribution of these two forms of HindIII.  Phosphocellulose chromatography in the presence
of
2M urea generated a broad peak of HindIII activity.  Addition of 4M urea, on the contrary,
showed
only one active peak of this enzyme.  The HindIII could be purified by the following DEAE-
cellulose chromatography.  The results indicate the presence of only one kind of HindIII
molecule,
which was alternated between free and bound forms, and a certain kind of factor that would
equilibrate these two forms.

<>

<1>Takasu, Y., Sajwan, S., Daimon, T., Osanai-Futahashi, M., Uchino, K., Sezutsu, H., Tamura, T., Zurovec, M.
<2>Efficient TALEN construction for Bombyx mori gene targeting.
<3>PLoS ONE
<4>8
<5>E73458
<6>2013
<7>Engineered nucleases are artificial enzymes able to introduce double stranded
breaks at desired genomic locations. The double stranded breaks start the
error-prone repair process of non-homologous end-joining (NHEJ), which eventually
leads to the induction of mutations at target sites. We showed earlier that ZFNs
and TALENs are able to induce NHEJ mutations in the B. mori genome. In order to
optimize our mutagenesis protocol, we modified one of the reported truncated
TALEN scaffolds and optimized it for use in the B. mori embryo. We also
established a novel B. mori somatic cell assay suitable for the preselection of
highly efficient TALENs directly in the B. mori model system. We compared the
efficiency of several TALEN pairs based on three different frameworks using the
BmBLOS2 gene. The new active TALENs show one order of magnitude higher efficiency
than those we used previously. We confirmed the utility of our improved protocol
by mutagenesis of the autosomal gene, red egg (Bm-re) and showed that it allows
obtaining homozygous mutants in G1. Our procedure minimizes the chance of failure
in B. mori gene targeting experiments.

<>

<1>Takata, T., Aras, R., Tavakoli, D., Ando, T., Olivares, A.Z., Blaser, M.J.
<2>Phenotypic and genotypic variation in methylases involved in type II restriction-modification systems in Helicobacter pylori.
<3>Nucleic Acids Res.
<4>30
<5>2444-2452
<6>2002
<7>To determine relationships between Helicobacter pylori geographical origin and type II
methylase activity, we examined 122 strains from various locations around the world for
methylase expression. Most geographic regions possessed at least one strain resistant to
digestion by each of 14 restriction endonucleases studied. Across all of the strains studied,
the average number of active methylases was 8.2 +/- 1.9 with no significant variation between
the major geographic regions. Although seven pairs of isolates showed the same susceptibility
patterns, their cagA/vacA status differed, and the remaining 108 strains each possessed unique
patterns of susceptibility. From a single clonal group, 15 of 18 strains showed identical
patterns of resistance, but diverged with respect to M.MboII activity. All of the methylases
studied were present in all major human population groupings, suggesting that their horizontal
acquisition pre-dated the separation of these populations. For the hpyV and hpyAIV
restriction-modification systems, an in-depth analysis of genotype, indicating extensive
diversity of cassette size and chromosomal locations regardless of the susceptibility
phenotype, points toward substantial strain-specific selection involving these loci.

<>

<1>Takata, T., Wassenaar, T.M., Xu, Q., Blaser, M.J.
<2>The gene product of Campylobacter jejuni gene Cj0208 is a DNA methyltransferase with specificity for GAATTC.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>102
<5>164
<6>2002
<7>To understand the mechanisms of DNA methylation in Campylobacter jejuni, we examined the
genetic and phenotypic properties of a putative
adenine methyltransferase gene, Cj0208. This gene encodes a 364-amino
acid protein and was present in all 11 C. jejuni strains tested. A
BLAST search using the Cj208 amino acid sequence revealed that the
putative protein was most closely related to CATG-recognizing adenine
DNA methyltransferases, including M. HpyI of Helicobacter pylori.
However, phylogenetic analysis (using PAUP 4.0b2) revealed that Cj0208
was closely related to M. Sse9I, which recognizes AATT, and that it's
next closest relative is M. EcoRI, which recognizes GAATTC. DNA
isolated from C. jejuni strains was shown to be resistant to digestion
by EcoRI and susceptible to restriction enzymes recognizing AATT
(TSP509I), and CATG (NlaIII). After cloning Cj0208 from C. jejuni
strain 11168, overexpression in an Escherichia coli strain lacking
endogenous methyltransferases rendered DNA from these cells resistant
to EcoRI digestion. This suggests that Cj0208 is a methyltransferase
with specificity for GAATTC. A dot blot assay using antibodies that
react specifically with DNA containing m6A or m4C modifications
identified Cj0208 as an m6A methyltransferase. An isogenic Cj0208
mutant was susceptible to EcoRI, confirming the role of this methylase
in protecting GAATTC sites. We conclude that Cj0208 encodes a DNA
methyltransferase with m6A activity and specificity for GAATTC. The
resistance of C. jejuni strains to EcoRI digestion suggests that this
gene is well-conserved.

<>

<1>Takatani, N., Nakanishi, M., Meirelles, P., Mino, S., Suda, W., Oshima, K., Hattori, M., Ohkuma, M., Hosokawa, M., Miyashita, K., Thompson, F.L., Niwa, A., Sawabe, T., Sawabe, T.
<2>Draft Genome Sequences of Marine Flavobacterium Algibacter lectus Strains SS8 and NR4.
<3>Genome Announcements
<4>2
<5>e01168-14
<6>2014
<7>Here, we present the draft genome sequences of a zeaxanthin-producing flavobacterium,
Algibacter lectus strains SS8 and NR4, isolated from coastal
sediment and rock surfaces in Hakodate, Japan, respectively. This genomic
information represents the first Algibacter genome sequences, which will help us
to elucidate the biology and evolution of Flavobacteriaceae bacteria.

<>

<1>Takatani, N., Nakanishi, M., Meirelles, P., Mino, S., Suda, W., Oshima, K., Hattori, M., Ohkuma, M., Hosokawa, M., Miyashita, K., Thompson, F.L., Niwa, A., Sawabe, T., Sawabe, T.
<2>Draft Genome Sequence of Marine Flavobacterium Jejuia pallidilutea Strain 11shimoA1 and Pigmentation Mutants.
<3>Genome Announcements
<4>2
<5>e01236-14
<6>2014
<7>Here, we present the draft genome sequence of a novel carotenoid 2'-isopentenylsaproxanthin
producer, Jejuia pallidilutea strain 11shimoA1,
isolated from the surface of seaweed in Japan, and the ethyl
methanesulfonate-induced pigmentation mutants. This genomic information will help
to not only elucidate the 2'-isopentenylsaproxanthin biosynthetic pathway but
also understand the evolution of flavobacteria.

<>

<1>Takeno, A., Okamoto, A., Tori, K., Oshima, K., Hirakawa, H., Toh, H., Agata, N., Yamada, K., Ogasawara, N., Hayashi, T., Shimizu, T., Kuhara, S., Hattori, M., Ohta, M.
<2>Complete Genome Sequence of Bacillus cereus NC7401, Which Produces High Levels of the Emetic Toxin Cereulide.
<3>J. Bacteriol.
<4>194
<5>4767-4768
<6>2012
<7>We report the complete and annotated genome sequence of Bacillus cereus NC7401, a
representative of the strain group that causes emetic-type food poisoning. The
emetic toxin, cereulide, is produced by a nonribosomal protein synthesis (NRPS)
system that is encoded by a gene cluster on a large resident plasmid, pNCcld.

<>

<1>Takeshima, H., Suetake, I., Tajima, S.
<2>Mouse Dnmt3a Preferentially Methylates Linker DNA and Is Inhibited by Histone H1.
<3>J. Mol. Biol.
<4>383
<5>810-821
<6>2008
<7>In mammals, DNA methylation is crucial for embryonic development and germ cell
differentiation. The DNA methylation patterns are created by de novo-type DNA
methyltransferases (Dnmts) 3a and 3b. Dnmt3a is crucial for global methylation, including that
of imprinted genes in germ cells. In eukaryotic nuclei, genomic DNA is packaged into
multinucleosomes with linker histone H1, which binds to core nucleosomes, simultaneously
making contacts in the linker DNA that separates adjacent nucleosomes. In the present study,
we prepared oligonucleosomes from HeLa nuclei with or without linker histone H1 and used them
as a substrate for Dnmt3a. Removal of histone H1 enhanced the DNA methylation activity.
Furthermore, Dnmt3a preferentially methylated the linker between the two nucleosome core
regions of reconstituted dinucleosomes, and the binding of histone H1 inhibited the DNA
methylation activity of Dnmt3a towards the linker DNA. Since an identical amount of histone H1
did not inhibit the activity towards naked DNA, the inhibitory effect of histone H1 was not on
the Dnmt3a catalytic activity but on its preferential location in the linker DNA of the
dinucleosomes. The central globular domain and C-terminal tail of the histone H1 molecule were
indispensable for inhibition of the DNA methylation activity of Dnmt3a. We propose that the
binding and release of histone H1 from the linker portion of chromatin may regulate the local
DNA methylation of the genome by Dnmt3a, which is expressed ubiquitously in somatic cells in
vivo.

<>

<1>Takeshita, K., Shibata, T.F., Nikoh, N., Nishiyama, T., Hasebe, M., Fukatsu, T., Shigenobu, S., Kikuchi, Y.
<2>Whole-Genome Sequence of Burkholderia sp. Strain RPE67, a Bacterial Gut Symbiont  of the Bean Bug Riptortus pedestris.
<3>Genome Announcements
<4>2
<5>e00556-14
<6>2014
<7>Burkholderia sp. strain RPE67 is a bacterial symbiont isolated from a field-collected bean
bug, Riptortus pedestris. To understand the genetic basis of
the insect-microbe symbiosis, we performed whole-genome sequencing of the
Burkholderia strain, revealing an 8.69-Mb genome consisting of three chromosomes
and three plasmids.

<>

<1>Takeshita, K., Suetake, I., Yamashita, E., Suga, M., Narita, H., Nakagawa, A., Tajima, S.
<2>Structural insight into maintenance methylation by mouse DNA methyltransferase 1 (Dnmt1).
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>9055-9059
<6>2011
<7>Methylation of cytosine in DNA plays a crucial role in development through inheritable gene
silencing. The DNA methyltransferase Dnmt1 is responsible for the propagation of methylation
patterns to the next generation via its preferential methylation of hemimethylated CpG sites
in the genome; however, how Dnmt1 maintains methylation patterns is not fully understood. Here
we report the crystal structure of the large fragment (291-1620) of mouse Dnmt1 and its
complexes with cofactor S-adenosyl-L-methionine and its product S-adenosyl-L-homocystein.
Notably, in the absence of DNA, the N-terminal domain responsible for targeting Dnmt1 to
replication foci is inserted into the DNA-binding pocket, indicating that this domain must be
removed for methylation to occur. Upon binding of S-adenosyl-L-methionine, the catalytic
cysteine residue undergoes a conformation transition to a catalytically competent position.
For the recognition of hemimethylated DNA, Dnmt1 is expected to utilize a target recognition
domain that overhangs the putative DNA-binding pocket. Taking into considerations the recent
report of a shorter fragment structure of Dnmt1 that the CXXC motif positions itself in the
catalytic pocket and prevents aberrant de novo methylation, we propose that maintenance
methylation is a multistep process accompanied by structural changes.

<>

<1>Takeuchi, F., Watanabe, S., Baba, T., Yuzawa, H., Ito, T., Morimoto, Y., Kuroda, M., Cui, L., Takahashi, M., Ankai, A., Baba, S., Fukui, S., Lee, J.C., Hiramatsu, K.
<2>Whole-genome sequencing of Staphylococcus haemolyticus uncovers the extreme plasticity of its genome and the evolution of Human-colonizing   Staphylococcal species.
<3>J. Bacteriol.
<4>187
<5>7292-7308
<6>2005
<7>Staphylococcus haemolyticus is an opportunistic bacterial pathogen that colonizes human skin
and is remarkable for its highly antibiotic-resistant
phenotype. We determined the complete genome sequence of S.haemolyticus to
better understand its pathogenicity and evolutionary relatedness to the
other staphylococcal species. A large proportion of the open reading
frames in the genomes of S.haemolyticus, Staphylococcus aureus, and
Staphylococcus epidermidis were conserved in their sequence and order on
the chromosome. We identified a region of the bacterial chromosome just
downstream of the origin of replication that showed little homology among
the species but was conserved among strains within a species. This novel
region, designated the "oriC environ," likely contributes to the evolution
and differentiation of the staphylococcal species, since it was enriched
for species-specific nonessential genes that contribute to the biological
features of each staphylococcal species. A comparative analysis of the
genomes of S.haemolyticus, S.aureus, and S.epidermidis elucidated
differences in their biological and genetic characteristics and pathogenic
potentials. We identified as many as 82 insertion sequences in the
S.haemolyticus chromosome that probably mediated frequent genomic
rearrangements, resulting in phenotypic diversification of the strain.
Such rearrangements could have brought genomic plasticity to this species
and contributed to its acquisition of antibiotic resistance.

<>

<1>Takeuchi, H., Donahue, J.P., Krishna, U., Miller, G.G., Peek, R.M. Jr., Israel, D.A.
<2>Quantitative detection of expression of the Helicobacter pylori methyltransferase-encoding gene hpyIM in vivo and in vitro.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>102
<5>242
<6>2002
<7>Helicobacter pylori are highly diverse bacteria that persist in the human stomach and induce
chronic gastritis for the virtual lifetime of
their hosts, a process that increases risk for peptic ulceration,
gastric adenocarcinoma, and gastric lymphoma. Despite a high degree of
genetic diversity, particularly among restriction-modification systems,
hpyIM which encodes a DNA adenine methyltransferase is highly conserved
among all H. pylori strains. Because of this high level of
conservation, we sought to determine whether levels of hpyIM expression
within colonized human mucosa correlated with disease status or mucosal
injury and whether there was a relationship between levels of
expression in vivo and in vitro. PCR-light and slot blot hybridizations
were used to quantitate hypIM transcripts in gastric tissue or in
broth-grown cells and quantities were normalized to levels of H. pylori
16s rRNA. Among 41 clinical biopsies, levels of hpyIM transcripts
varied dramatically and were not related to disease outcome or severity
of inflammation. Four strains isolated from biopsies were selected for
more detailed in vitro studies, and for each strain, levels of hpyIM
were higher during log-phase growth compared with stationary phase. For
three of the four strains, levels of hypIM transcripts were lower in
vitro than in vivo. Since bacterial contact with host cells regulates
gene expression in other organisms, we next sought to determine if
adherence of H. pylori to gastric epithelial cells resulted in hpyIM
expression patterns that more closely reflected levels observed in
vivo. hpyIM transcript levels after two hours of adherence were not
related to levels identified within inflamed tissue, suggesting that
events independent from isolated bacterial:epithelial cell contact may
regulate hpyIM expression in vivo.

<>

<1>Takeuchi, H., Israel, D.A., Miller, G.G., Donahue, J.P., Krishna, U., Gaus, K., Peek, R.M.
<2>Characterization of expression of a functionally conserved Helicobacter pylori methyltransferase-encoding gene within inflamed mucosa and  during in vitro growth.
<3>J. Infect. Dis.
<4>186
<5>1186-1189
<6>2002
<7>Methylation of bacterial DNA can regulate microbial growth and virulence. Expression of hpyIM,
a conserved methyltransferase of the
gastric pathogen Helicobacter pylori, was quantitated in gastric biopsy
specimens from 41 H. pylori-infected patients and during growth in
vitro, by quantitative reverse transcriptase-polymerase chain reaction
and/or RNA slot-blot analysis, to determine whether levels of
transcription were associated with pathologic outcome, as based on both
severity of gastritis and inflammatory cytokine levels, or were
regulated by bacterial growth phase. The effects that hpyIM
inactivation has on bacterial morphology were determined by electron
microscopy. Expression of hpyIM varied dramatically within colonized
gastric tissue, and levels were not related to either colonization
density, severity of inflammation, mucosal IL-8 concentrations, or
clinical disease. In vitro, hpyIM expression was higher during
log-phase growth and was required for normal bacterial morphology,
suggesting that hpyIM expression may be growth-phase regulated within
the gastric niche.

<>

<1>Takeuchi, K., Noda, N., Someya, N.
<2>Complete Genome Sequence of the Biocontrol Strain Pseudomonas protegens Cab57 Discovered in Japan Reveals Strain-Specific Diversity of This Species.
<3>PLoS ONE
<4>9
<5>E93683
<6>2014
<7>The biocontrol strain Pseudomonas sp. Cab57 was isolated from the rhizosphere of
shepherd's purse growing in a field in Hokkaido by screening the antibiotic
producers. The whole genome sequence of this strain was obtained by paired-end
and whole-genome shotgun sequencing, and the gaps between the contigs were closed
using gap-spanning PCR products. The P. sp. Cab57 genome is organized into a
single circular chromosome with 6,827,892 bp, 63.3% G+C content, and 6,186
predicted protein-coding sequences. Based on 16S rRNA gene analysis and whole
genome analysis, strain Cab57 was identified as P. protegens. As reported in P.
protegens CHA0 and Pf-5, four gene clusters (phl, prn, plt, and hcn) encoding the
typical antibiotic metabolites and the reported genes associated with Gac/Rsm
signal transduction pathway of these strains are fully conserved in the Cab57
genome. Actually strain Cab57 exhibited typical Gac/Rsm activities and antibiotic
production, and these activities were enhanced by knocking out the retS gene (for
a sensor kinase acting as an antagonist of GacS). Two large segments (79 and 115
kb) lacking in the Cab57 genome, as compared with the Pf-5 genome, accounted for
the majority of the difference (247 kb) between these genomes. One of these
segments was the complete rhizoxin analog biosynthesis gene cluster (ca. 79 kb)
and another one was the 115-kb mobile genomic island. A whole genome comparison
of those relative strains revealed that each strain has unique gene clusters
involved in metabolism such as nitrite/nitrate assimilation, which was identified
in the Cab57 genome. These findings suggest that P. protegens is a ubiquitous
bacterium that controls its biocontrol traits while building up strain-specific
genomic repertoires for the biosynthesis of secondary metabolites and niche
adaptation.

<>

<1>Takeuchi, R., Certo, M., Caprara, M.G., Scharenberg, A.M., Stoddard, B.L.
<2>Optimization of in vivo activity of a bifunctional homing endonuclease and maturase reverses evolutionary degradation.
<3>Nucleic Acids Res.
<4>37
<5>877-890
<6>2009
<7>The LAGLIDADG homing endonuclease (LHE) I-AniI has adopted an extremely efficient secondary
RNA splicing activity that is beneficial to its host,
balanced against inefficient DNA cleavage. A selection experiment
identified point mutations in the enzyme that act synergistically to
improve endonuclease activity. The amino-acid substitutions increase
target affinity, alter the thermal cleavage profile and significantly
increase targeted recombination in transfected cells. The RNA splicing
activity is not affected by these mutations. The improvement in DNA
cleavage activity is largely focused on one of the enzyme's two active
sites, corresponding to a rearrangement of a lysine residue hypothesized
to act as a general base. Most of the constructs isolated in the screen
contain one or more mutations that revert an amino-acid identity to a
residue found in one or more close homologues of I-AniI. This implies that
mutations that have previously reduced the endonuclease activity of I-AniI
are identified and reversed, sometimes in combination with additional
'artificial' mutations, to optimize its in vivo activity.

<>

<1>Takeuchi, R., Lambert, A.R., Mak, A.N., Jacoby, K., Dickson, R.J., Gloor, G.B., Scharenberg, A.M., Edgell, D.R., Stoddard, B.L.
<2>Tapping natural reservoirs of homing endonucleases for targeted gene modification.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>13077-13082
<6>2011
<7>Homing endonucleases mobilize their own genes by generating double-strand breaks at individual
target sites within potential host DNA. Because of their high specificity, these proteins are
used for 'genome editing' in higher eukaryotes. However, alteration of homing endonuclease
specificity is quite challenging. Here we describe the identification and phylogenetic
analysis of over 200 naturally occurring LAGLIDADG homing endonucleases (LHEs). Biochemical
and structural characterization of endonucleases from one clade within the phylogenetic tree
demonstrates strong conservation of protein structure contrasted against highly diverged DNA
target sites and indicates that a significant fraction of these proteins are sufficiently
stable and active to serve as engineering scaffolds. This information was exploited to create
a targeting enzyme to disrupt the endogenous monoamine oxidase B gene in human cells. The
ubiquitous presence and diversity of LHEs described in this study may facilitate the creation
of many tailored nucleases for genome editing.

<>

<1>Takiguchi, M., Achanzar, W.E., Qu, W., Li, G., Waalkes, M.P.
<2>Effects of cadmium on DNA-(cytosine-5) methyltransferase activity and DNA methylation status during cadmium-induced cellular transformation.
<3>Exp. Cell Res.
<4>286
<5>355-365
<6>2003
<7>Cadmium is a human carcinogen that likely acts via epigenetic mechanisms. Since DNA
methylation alterations represent an important epigenetic event
linked to cancer, the effect of cadmium on DNA methyltransferase (MeTase)
activity was examined using in vitro (TRL1215 rat liver cells) and ex vivo
(M.SssI DNA MeTase) systems. Cadmium effectively inhibited DNA MeTases in
a manner that was noncompetitive with respect to substrate (DNA),
indicating an interaction with the DNA binding domain rather than the
active site. Based on these results, the effects of prolonged cadmium
exposure on DNA MeTase and genomic DNA methylation in TRL1215 cells were
studied. After 1 week of exposure to 0-2.5 microM cadmium, DNA MeTase
activity was reduced (up to 40%) in a concentration-dependent fashion,
while genomic DNA methylation showed slight but significant reductions at
the two highest concentrations. After 10 weeks of exposure, the cells
exhibited indications of transformation, including hyperproliferation,
increased invasiveness, and decreased serum dependence. Unexpectedly,
these cadmium-transformed cells exhibited significant increases in DNA
methylation and DNA MeTase activity. These results indicate that, while
cadmium is an effective inhibitor of DNA MeTase and initially induces DNA
hypomethylation, prolonged exposure results in DNA hypermethylation and
enhanced DNA MeTase activity.

<>

<1>Taktakishvili, M.O., Tabdzhun, A.
<2>Oligodeoxynucleotides, containing 6-O-alkylated guanine segments recognized by HaeIII restriction endonuclease.
<3>Bioorg. Khim.
<4>17
<5>806-808
<6>1991
<7>A series of self-complementary decadeoxynucleotides containing a native or modified HaeIII
site GGCC (with one or both guanine residues 6-0-cetylated) have been synthesized by the
phosphotriester approach.  The nonmodified decanucleotide is normally digested with snake
venom phosphodiesterase as well as with HaeIII and BspI restriction endonucleases, whereas the
bulky 6-O-alkyl substitutent strongly inhibits the VPDE hydrolysis and completely prevents
digestion with the endonucleases.

<>

<1>Taman-Chardonnens, A.A., Puzio, P., McKersie, B.
<2>Nucleic acid sequences encoding proteins associated with abiotic stress response and plant cells and plants with in-creased tolerance to environmental stress.
<3>European Patent Office
<4>EP 2145960 A
<5>
<6>2010
<7>This invention relates generally to nucleic acid sequences encoding proteins that are
associated with abiotic stress responses and abiotic stress tolerance in plants.  This
invention further relates to transformed plant cells with altered metabolic activity compared
to a corresponding non-transformed, wild-type plant cell, wherein the metabolic activity is
altered by transformation with a Stress-Related Protein coding nucleic acid and results in
increased tolerance and/or resistance to an environmental stress as compared to a
corresponding non-transformed, wild-type plant cell.

<>

<1>Tamaru, H., Selker, E.U.
<2>A histone H3 methyltransferase controls DNA methylation in Neurospora crassa.
<3>Nature
<4>414
<5>277-283
<6>2001
<7>DNA methylation is involved in epigenetic processes such as X-chromosome inactivation,
imprinting and silencing of transposons. We have demonstrated previously that dim-2 encodes a
DNA methyltransferase that is responsible for all known cytosine methylation in Neurospora
crassa. Here we report that another Neurospora gene, dim-5, is required for DNA methylation,
as well as for normal growth and full fertility. We mapped dim-5 and identified it by
transformation with a candidate gene. The mutant has a nonsense mutation in a SET domain of a
gene related to histone methyltransferases that are involved in heterochromatin formation in
other organisms.  Transformation of a wild-type strain with a segment of dim-5 reactivated a
silenced hph gene, apparently by 'quelling' of dim-5. We demonstrate that recombinant DIM-5
protein specifically methylates histone H3 and that replacement of lysine 9 in histone H3 with
either a leucine or an arginine phenocopies the dim-5 mutation. We conclude that DNA
methylation depends on histone methylation.

<>

<1>Tamaru, Y., Miyake, H., Kuroda, K., Nakanishi, A., Kawade, Y., Yamamoto, K., Uemura, M., Fujita, Y., Doi, R.H., Ueda, M.
<2>Genome Sequence of the Cellulosome-Producing Mesophilic Organism Clostridium cellulovorans 743B.
<3>J. Bacteriol.
<4>192
<5>901-902
<6>2010
<7>Clostridium cellulovorans 743B was isolated from a wood chip pile and is an anaerobic and
mesophilic spore-forming bacterium. This organism
degrades native substrates in soft biomass such as corn fiber and rice
straw efficiently by producing an extracellular enzyme complex called the
cellulosome. Here we report the genome sequence of C. cellulovorans 743B.

<>

<1>Tamas, I., Dedysh, S.N., Liesack, W., Stott, M.B., Alam, M., Murrell, J.C., Dunfield, P.F.
<2>Complete Genome Sequence of Beijerinckia indica subsp. indica.
<3>J. Bacteriol.
<4>192
<5>4532-4533
<6>2010
<7>Beijerinckia indica subsp. indica is an aerobic, acidophilic, exopolysaccharide-producing,
N2-fixing soil bacterium. It is a generalist
chemoorganotroph that is phylogenetically closely related to facultative
and obligate methanotrophs of the genera Methylocella and Methylocapsa.
Here we report the full genome sequence of this bacterium.

<>

<1>Tamas, I., Klasson, L., Canback, B., Naslund, A.K., Eriksson, A.-S., Wernegreen, J.J., Sandstrom, J.P., Moran, N.A., Andersson, S.G.E.
<2>50 million years of genomic stasis in endosymbiotic bacteria.
<3>Science
<4>296
<5>2376-2379
<6>2002
<7>Comparison of two fully sequenced genomes of Buchnera aphidicola, the obligate endosymbionts
of aphids, reveals the most extreme genome stability to date: no chromosome rearrangements or
gene acquisitions have occurred in the past 50 to 70 million years, despite substantial
sequence evolution and the inactivation and loss of individual genes. In contrast, the genomes
of their closest free-living relatives, Escherichia coli and Salmonella spp., are more than
2000-fold more labile in content and gene order. The genomic stasis of B. aphidicola, likely
attributable to the loss of phages, repeated sequences, and recA, indicates that B. aphidicola
is no longer a source of ecological innovation for its hosts.

<>

<1>Tamulaitiene, G., Grazulis, S., Janulaitis, A., Janowski, R., Bujacz, G., Jaskolski, M.
<2>Crystallization and preliminary crystallographic studies of a bifunctional restriction endonuclease Eco57I.
<3>Biochim. Biophys. Acta
<4>1698
<5>251-254
<6>2004
<7>Restriction endonuclease Eco57I from Escherichia coli recognizes asymmetric DNA sequence
5'-CTGAAG and has both restriction (DNA cleavage a short distance away from the recognition
site) and modification (methylation) activities residing in a single polypeptide chain. Single
crystals of wild-type Eco57I ternary complexes with double-stranded DNA and sinefungin, a
stimulator of endonuclease activity, were obtained by the vapor diffusion technique and
characterized crystallographically for different variants of the DNA component. The best data
for the complex with 25-mer DNA were collected to 4.2-A resolution at 100 K using synchrotron
radiation. The crystals are orthorhombic, space group P2(1)2(1)2, with a=164.3, b=293.0,
c=71.1 A, and contain two to four copies of the protein in the asymmetric unit.

<>

<1>Tamulaitiene, G., Jakubauskas, A., Urbanke, C., Huber, R., Grazulis, S., Siksnys, V.
<2>The crystal structure of the rare-cutting restriction enzyme SdaI reveals unexpected domain architecture.
<3>Structure
<4>14
<5>1389-1400
<6>2006
<7>Rare-cutting restriction enzymes are important tools in genome analysis.  We report here the
crystal structure of SdaI restriction endonuclease, which is specific for the 8 bp sequence
CCTGCA/GG ("/" designates the cleavage site).  Unlike orthodox Type IIP enzymes, which are
single domain proteins, the SdaI monomer is composed of two structural domains.  The N domain
contains a classical winged helix-turn-helix (wHTH) DNA binding motif, while the C domain
shows a typical restriction endonuclease fold.  The active site of SdaI is located within the
C domain and represents a variant of the canonical PD-(D/E)XK motif.  SdaI determinants of
sequence specificity are clustered on the recognition helix of the wHTH motif at the N domain.
The modular architecture of SdaI, wherein one domain mediates DNA binding while the other
domain is predicted to catalyze hydrolysis, distinguishes SdaI from previously characterized
restriction enzymes interacting with symmetric recognition sequences.

<>

<1>Tamulaitiene, G., Siksnys, V.
<2>NotI Is Not Boring.
<3>Structure
<4>16
<5>497-498
<6>2008
<7>Crystal structures of the restriction endonuclease NotI in free and DNA bound forms, presented
in this issue of Structure (Lambert et al., 2008), provide a unique insight into the
structural details of 8 base pair  sequence recognition by the restriction enzyme.

<>

<1>Tamulaitiene, G., Silanskas, A., Grazulis, S., Zaremba, M., Siksnys, V.
<2>Crystal structure of the R-protein of the multisubunit ATP-dependent restriction  endonuclease NgoAVII.
<3>Nucleic Acids Res.
<4>42
<5>14022-14030
<6>2014
<7>The restriction endonuclease (REase) NgoAVII is composed of two proteins, R.NgoAVII and
N.NgoAVII, and shares features of both Type II restriction enzymes and Type I/III
ATP-dependent restriction enzymes (see accompanying paper Zaremba et al., 2014). Here we
present crystal structures of the R.NgoAVII apo-protein and the R.NgoAVII C-terminal domain
bound to a specific DNA. R.NgoAVII is composed of two domains: an N-terminal nucleolytic PLD
domain; and a C-terminal B3-like DNA-binding domain identified previously in BfiI and EcoRII
REases, and in plant transcription factors. Structural comparison of the B3-like domains of
R.NgoAVII, EcoRII, BfiI and the plant transcription factors revealed a conserved DNA-binding
surface comprised of N- and C-arms that together grip the DNA. The C-arms of R.NgoAVII,
EcoRII, BfiI and plant B3 domains are similar in size, but the R.NgoAVII N-arm which makes the
majority of the contacts to the target site is much longer. The overall structures of
R.NgoAVII and BfiI are similar; however, whilst BfiI has stand-alone catalytic activity,
R.NgoAVII requires an auxiliary cognate N.NgoAVII protein and ATP hydrolysis in order to
cleave DNA at the target site. The structures we present will help formulate future
experiments to explore the molecular mechanisms of intersubunit crosstalk that control DNA
cleavage by R.NgoAVII and related endonucleases.

<>

<1>Tamulaitis, G., Lagunavicius, A.
<2>DNA bending induced by MunI restriction endonuclease.
<3>Biologija
<4>0
<5>11-15
<6>2000
<7>Bending of DNA induced by the specific binding of the restriction endonuclease MunI has been
investigated by the empirical method of Thompson and Landy and the method of calibrated
standards.  The R.MunI induced bending angles were found to be 42o +/- 4o at pH 6.5 and 56o
+/1 4o at pH 8.3 in the presence of Ca2+ ions.  Similar values of DNA bending angles were
obtained in the complexes of active site mutants D83A and E98A with specific DNA under the
same conditions.  Results of bending experiments support the assumption that elimination of
electrostatic repulsion between charged carboxylates and phosphate oxygens by protonation of
the active site carboxylate residue(s), their replacement to alanine or neutralization by Ca2+
binding yields similar DNA-protein complexes.

<>

<1>Tamulaitis, G., Mucke, M., Siksnys, V.
<2>Biochemical and mutational analysis of EcoRII functional domains reveals evolutionary links between restriction enzymes.
<3>FEBS Lett.
<4>580
<5>1665-1671
<6>2006
<7>The archetypal Type BE restriction endonuclease EcoRII is a dimer that has a modular
structure. DNA binding studies indicate that the isolated
C-terminal domain dimer has an interface that binds a single cognate
DNA molecule whereas the N-terminal domain is a monomer that also binds
a single copy of cognate DNA. Hence, the full-length EcoRII contains
three putative DNA binding interfaces: one at the C-terminal domain
dimer and two at each of the N-terminal domains. Mutational analysis
indicates that the C-terminal domain shares conserved active site
architecture and DNA binding elements with the tetrameric restriction
enzyme NgoMIV. Data provided here suggest possible evolutionary
relationships between different subfamilies of restriction enzymes.

<>

<1>Tamulaitis, G., Sasnauskas, G., Mucke, M., Siksnys, V.
<2>Simultaneous Binding of Three Recognition Sites is Necessary for a Concerted Plasmid DNA Cleavage by EcoRII Restriction Endonuclease.
<3>J. Mol. Biol.
<4>358
<5>406-419
<6>2006
<7>According to the current paradigm type IIE restriction endonucleases are homodimeric proteins
that simultaneously bind to two recognition sites but
cleave DNA at only one site per turnover: the other site acts as an
allosteric locus, activating the enzyme to cleave DNA at the first.
Structural and biochemical analysis of the archetypal type IIE restriction
enzyme EcoRII suggests that it has three possible DNA binding interfaces
enabling simultaneous binding of three recognition sites. To test if
putative synapsis of three binding sites has any functional significance,
we have studied EcoRII cleavage of plasmids containing a single, two and
three recognition sites under both single turnover and steady state
conditions. EcoRII displays distinct reaction patterns on different
substrates: (i) it shows virtually no activity on a single site plasmid;
(ii) it yields open-circular DNA form nicked at one strand as an
obligatory intermediate acting on a two-site plasmid; (iii) it cleaves
concertedly both DNA strands at a single site during a single turnover on
a three site plasmid to yield linear DNA. Cognate oligonucleotide added in
trans increases the reaction velocity and changes the reaction pattern for
the EcoRII cleavage of one and two-site plasmids but has little effect on
the three-site plasmid. Taken together the data indicate that EcoRII
requires simultaneous binding of three rather than two recognition sites
in cis to achieve concerted DNA cleavage at a single site. We show that
the orthodox type IIP enzyme PspGI which is an isoschisomer of EcoRII,
cleaves different plasmid substrates with equal rates. Data provided here
indicate that type IIE restriction enzymes EcoRII and NaeI follow
different mechanisms. We propose that other type IIE restriction enzymes
may employ the mechanism suggested here for EcoRII.

<>

<1>Tamulaitis, G., Solonin, A.S., Siksnys, V.
<2>Alternative arrangements of catalytic residues at the active sites of restriction enzymes.
<3>FEBS Lett.
<4>518
<5>17-22
<6>2002
<7>A catalytic sequence motif PDX10-30(E/D)XK is found in many restriction enzymes. On the basis
of sequence similarities and mapping of the conserved residues to the crystal structure of
NgoMIV we suggest that residues D160, K182, R186, R188 and E195 contribute to the
catalytic/DNA binding site of the Ecl18kI restriction endonuclease. Mutational analysis
confirms the functional significance of the conserved residues of Ecl18kI. Therefore, we
conclude that the active site motif 159VDX21KX12E of Ecl18kI differs from the canonical
PDX10-30(E/D)XK motif characteristic for most of the restriction enzymes. Moreover, we propose
that two subfamilies of endonucleases Ecl18kI/PspGI/EcoRII and Cfr10I/Bse634I/NgoMIV,
specific, respectively, for CCNGG/CCWGG and RCCGGY/GCCGGC sites, share conserved active site
architecture and DNA binding elements.

<>

<1>Tamulaitis, G., Zaremba, M., Szczepanowski, R.H., Bochtler, M., Siksnys, V.
<2>Nucleotide flipping by restriction enzymes analyzed by 2-aminopurine steady-state fluorescence.
<3>Nucleic Acids Res.
<4>35
<5>4792-4799
<6>2007
<7>Many DNA modification and repair enzymes require access to DNA bases and therefore flip
nucleotides. Restriction endonucleases (REases) hydrolyze the phosphodiester backbone within
or in the vicinity of the target recognition site and do not require base extrusion for the
sequence readout and catalysis. Therefore, the observation of extrahelical nucleotides in a
co-crystal of REase Ecl18kI with the cognate sequence, CCNGG, was unexpected. It turned out
that Ecl18kI reads directly only the CCGG sequence and skips the unspecified N nucleotides,
flipping them out from the helix. Sequence and structure conservation predict nucleotide
flipping also for the complexes of PspGI and EcoRII with their target DNAs (/CCWGG), but data
in solution are limited and indirect. Here, we demonstrate that Ecl18kI, the C-terminal domain
of EcoRII (EcoRII-C) and PspGI enhance the fluorescence of 2-aminopurines (2-AP) placed at the
centers of their recognition sequences. The fluorescence increase is largest for PspGI,
intermediate for EcoRII-C and smallest for Ecl18kI, probably reflecting the differences in the
hydrophobicity of the binding pockets within the protein. Omitting divalent metal cations and
mutation of the binding pocket tryptophan to alanine strongly increase the 2-AP signal in the
Ecl18kI-DNA complex. Together, our data provide the first direct evidence that Ecl18kI,
EcoRII-C and PspGI flip nucleotides in solution.

<>

<1>Tamulaitis, G., Zaremba, M., Szczepanowski, R.H., Bochtler, M., Siksnys, V.
<2>How PspGI, catalytic domain of EcoRII and Ecl18kI acquire specificities for different DNA targets.
<3>Nucleic Acids Res.
<4>36
<5>6101-6108
<6>2008
<7>Restriction endonucleases Ecl18kI and PspGI/catalytic domain of EcoRII recognize CCNGG and
CCWGG sequences (W stands for A or T), respectively.
The enzymes are structurally similar, interact identically with the
palindromic CC:GG parts of their recognition sequences and flip the
nucleotides at their centers. Specificity for the central nucleotides
could be influenced by the strength/stability of the base pair to be
disrupted and/or by direct interactions of the enzymes with the flipped
bases. Here, we address the importance of these contributions. We
demonstrate that wt Ecl18kI cleaves oligoduplexes containing canonical,
mismatched and abasic sites in the central position of its target sequence
CCNGG with equal efficiencies. In contrast, substitutions in the binding
pocket for the extrahelical base alter the Ecl18kI preference for the
target site: the W61Y mutant prefers only certain mismatched substrates,
and the W61A variant cuts exclusively at abasic sites, suggesting that
pocket interactions play a major role in base discrimination. PspGI and
catalytic domain of EcoRII probe the stability of the central base pair
and the identity of the flipped bases in the pockets. This 'double check'
mechanism explains their extraordinary specificity for an A/T pair in the
flipping position.

<>

<1>Tamura, T., Araki, Y., Yamaoka, S., Inagaki, K., Tanaka, H.
<2>Determination of methylation specificity of sequence-specific DNA methyltransferases using matrix assisted laser desorption/ionization time-of-flight mass spectrometry.
<3>Nucleic Acids Res.
<4>25
<5>4162-4164
<6>1997
<7>We describe here a sensitive and straightforward method for characterizing the methylation
specificity of type II DNA methyltransferases (Mtases) using matrix-assisted laser
desorption/ionization time-of-flight mass spectrometry.  DNA substrate, prepared by ligation
of a commercially available oligonucleotide, was modified by the subject MTase and was
derivatized to a mixture of single-stranded oligonucleotides through endonuclease treatment,
heat-denaturation and limited digestion by 3'-terminus-specific phosphodiesterase I.
MALDI-TOF mass spectrometry was used to determine the mass differences between the digestion
products, and the methylated nucleotide was explicitly identified by the mass increase in 14
Da due to the base modification.  The method was applicable to the three representative MTases
M.EcoRI, M.BamHI and M.HaeIII.

<>

<1>Tamura, T., Kataoka, A., Shu, L.Y., Ashida, A., Tanaka, H., Inagaki, K.
<2>An in vitro Screening Method for DNA Cytosine-C5-Methylase Inhibitor.
<3>Nat. Prod. Lett.
<4>16
<5>25-27
<6>2002
<7>A specific inhibitor of DNA cytosine C-5-methylases would be useful for studying genomic
imprinting, X-chromosome inactivation, carcinogenesis,
and regulation of tissue-specific gene expression, for these
physiological phenomena appears to be regulated through DNA methylation
in promoter sequences. This paper reports a novel convenient in vitro
assay method for screening DNA cytosine C-5-methylase inhibitor. Our
method uses a commercially available HaeIII methylase (cytosine C-5
methylase), its corresponding HaeIII endonuclease, and lambda DNA as
their substrate.

<>

<1>Tan, B., Charchuk, R., Li, C., Nesbo, C., Abu, L.N., Foght, J.
<2>Draft Genome Sequence of Uncultivated Firmicutes (Peptococcaceae SCADC) Single Cells Sorted from Methanogenic Alkane-Degrading Cultures.
<3>Genome Announcements
<4>2
<5>e00909-14
<6>2014
<7>The draft genome of an uncultivated bacterium affiliated with the Peptococcaceae  was
reconstructed by co-assembling Illumina MiSeq sequences from three single
cells sorted by microfluidics from two methanogenic alkane-degrading cultures.
Peptococcaceae SCADC (short-chain alkane-degrading culture) may be genetically
capable of anaerobic alkane activation by fumarate addition in the absence of
sulfate.

<>

<1>Tan, B., de Araujo, E.S.R., Rozycki, T., Nesbo, C., Foght, J.
<2>Draft Genome Sequences of Three Smithella spp. Obtained from a Methanogenic Alkane-Degrading Culture and Oil Field Produced Water.
<3>Genome Announcements
<4>2
<5>e01085-14
<6>2014
<7>Two draft genomes affiliated with Smithella spp. were obtained from a methanogenic
alkane-degrading enrichment culture by single-cell sorting and
metagenome contig binning, and a third was obtained by single-cell sorting of oil
field produced water. Two genomes contained putative assABC genes encoding
alkylsuccinate synthase, indicating genetic potential for fumarate activation of
alkanes.

<>

<1>Tan, B., Foght, J.
<2>Draft genome sequences of campylobacterales (epsilonproteobacteria) obtained from methanogenic oil sands tailings pond metagenomes.
<3>Genome Announcements
<4>2
<5>e01034-14
<6>2014
<7>Draft genome sequences of two Campylobacterales (Sulfurospirillum sp. strain SCADC and
Sulfuricurvum sp. strain MLSB [Mildred Lake Settling Basin]) were
obtained by taxonomic binning of metagenomes originating from an oil sands
tailings pond. Both genomes contain soxABXYZ genes involved in sulfur oxidation,
highlighting their potential roles in sulfur cycling in oil sands tailings ponds.

<>

<1>Tan, C.
<2>Virulence Gene and Protein, and Their Use.
<3>Japanese Patent Office
<4>JP 2003532404 A
<5>
<6>2003
<7>
<>

<1>Tan, C., Xu, Z., Zheng, H., Liu, W., Tang, X., Shou, J., Wu, B., Wang, S., Zhao, G.P., Chen, H.
<2>Genome Sequence of Porcine Extraintestinal Pathogenic Escherichia coli strain.
<3>J. Bacteriol.
<4>193
<5>5038
<6>2011
<7>Extraintestinal Pathogenic Escherichia coli is an important pathogen which can infect human
and animals and causing many diseases outside the
intestinal. Here, we report the first draft genome sequence of a porcine
ExPEC strain, PCN033, isolated from the pig with meningitis.

<>

<1>Tan, E., Chen, Y., Kuan, J., Lin, C., Jagoda, S.S., Lin, F., Tzou, W., Kinoshita, S., Watabe, S., Asakawa, S., Liu, S.
<2>Draft Genome Sequence of Thermoanaerobacterium saccharolyticum Strain NTOU1, a Thermophilic Bacterium Isolated from Marine Shallow Hydrothermal Vents.
<3>Genome Announcements
<4>2
<5>e01019-14
<6>2014
<7>Thermoanaerobacterium saccharolyticum strain NTOU1 has the ability to utilize several kinds of
sugars in lignocellulosic biomass to produce ethanol more
efficiently than other bacteria. Here, we report the draft genome sequence and
annotation of this strain, which may provide insights into the possible genes and
metabolic pathways related to ethanol production.

<>

<1>Tan, J.L., Ng, H.F., Wee, W.Y., Ang, M.Y., Wong, G.J., Ngeow, Y.F., Choo, S.W.
<2>First Whole-Genome Sequence of Mycobacterium iranicum, a Newly Reported Mycobacterial Species.
<3>Genome Announcements
<4>1
<5>e00732-13
<6>2013
<7>Mycobacterium iranicum is a new species of nontuberculous mycobacterium reported  in 2013.
Here, we describe the first whole-genome sequence of this species, that
of M. iranicum strain UM_TJL, isolated from a patient in Malaysia.

<>

<1>Tan, J.Y., Yin, W.F., Chan, K.G.
<2>Gene clusters of Hafnia alvei strain FB1 important in survival and pathogenesis: a draft genome perspective.
<3>Gut Pathog.
<4>6
<5>29
<6>2014
<7>BACKGROUND: Hafnia alvei is an opportunistic pathogen involved in various types of nosocomical
infections. The species has been found to inhabit food and mammalian guts. However, its status
as an enteropathogen, and whether the food-inhabiting strains could be a source of
gastrointestinal infection remains obscure. In this report we present a draft genome of H.
alvei strain FB1 isolated from fish paste meatball, a food popular among Malaysian and Chinese
populations.
The data was generated on the Illumina MiSeq platform. RESULTS: A comparative study was
carried out on FB1 against two other previously sequenced H. alvei genomes. Several gene
clusters putatively involved in survival and pathogenesis of H. alvei FB1 in food and gut
environment were characterised in this study.
These include the widespread colonisation island (WCI), the tad locus that is known to play an
essential role in biofilm formation, a eut operon that might contribute to advantage in
nutrient acquisition in gut environment, and genes responsible for siderophore production This
features enable the bacteria to successful colonise in the host gut environment. CONCLUSION:
With the whole genome data of H. alvei FB1 presented in this study, we hope to provide an
insight into future studies on this candidate of enteropathogen by looking into the possible
mechanisms employed to survive stresses and gain advantage in competitions, which eventually
leads to successful colonisation and pathogenesis.
This is to serve as the basis for more effective clinical diagnosis and treatment.

<>

<1>Tan, N.-W., Li, F.F.L.
<2>Interaction of oligonucleotides containing 6-O-methylguanine with human DNA (cytosine-5-)-methyltransferase.
<3>Biochemistry
<4>29
<5>9234-9240
<6>1990
<7>Thirty-base-pair synthetic oligonucleotide duplexes containing a single meG.C
(meG=6-O-methylguanine) or A.C base pair at the 16th position (i.e.,
5'-CCCGTTTAAATATACXTATACCCGGGTACC-3', where X=A or meG) were used to study de
novo methylation by the purified human DNA (cytosine-5)-methyltransferase
isolated from CEM cells.  Both duplexes containing meG.C and A.C base pairs
show enhanced methyl group acceptor properties.  Subsequent introduction of
hemimethylated sites at the 15th position of the top strand (the C residue next
to the abnormal base pair) and the 7th, 15th (which represents the C residue in
the 6meG.C and A.C base pairs), and 27th positions of the bottom strand were
used to study the maintenance methylation of the hemimethylated duplexes by the
methylase.  This revealed striking differences in the rate, amount, and sites
of methylation, which are dependent on the position of the hemimethylated site
in the duplex.  The possible mechanism of action of the methylase is discussed.
The data show that 6-O-methylguanine residues in DNA can have other genetic
effects apart from their miscoding behavior and that meG.C and A.C base pairs
exert different effects in terms of methylation.

<>

<1>Tan, W.G.H., Barkman, T.J., Gregory, C.V., Essani, K.
<2>Comparative genomic analyses of frog virus 3, type species of the genus Ranavirus (family Iridoviridae).
<3>Virology
<4>323
<5>70-84
<6>2004
<7>Frog virus 3 (FV3) is the type species member of the genus Ranavirus (family Iridoviridae). To
better understand the molecular mechanisms involved in the replication of FV3, including
transcription of its highly methylated DNA genome, we have determined the complete nucleotide
sequence of the FV3 genome. The FV3 genome is 105903 bp long excluding the terminal
redundancy. The G + C content of FV3 genome is 55% and it encodes 98 nonoverlapping potential
open reading frames (ORFs) containing 50-1293 amino acids. Eighty-four ORFs have significant
homology to known proteins of other iridoviruses, whereas twelve of these unique FV3 proteins
do not share homology to any known protein. A microsatellite containing a stretch of 34
tandemly repeated CA dinucleotide in a noncoding region was detected. To date, no such
sequence has been reported in any animal virus.

<>

<1>Tanaka, M.
<2>Recombinant vector for expression of fused protein, and method for degrading extranuclear gene of nonhuman organism using the recombinant vector.
<3>Japanese Patent Office
<4>JP 2002176988 A
<5>
<6>2002
<7>
<>

<1>Tanaka, T.
<2>Restriction of plasmid-mediated transformation in Bacillus subtilis 168.
<3>Mol. Gen. Genet.
<4>175
<5>235-237
<6>1979
<7>When plasmids carrying leucine genes of Bacillus subtilis 168 were isolated
from a restriction and modification deficient (r-m-) strain and used for
transformation of a restricting strain B. subtilis 168 leu recE4, the number of
transformants was greatly reduced.  Transformation of a rec+ strain
(transformation by integration of the donor DNA into the chromosome) with the
plasmids was not affected irrespective of whether the recipient carried the r+
or r- phenotype.  These results show that the plasmid-mediated transformation
is subject to the host controlled restriction and suggest that r-m- strains
should be used for construction of recombinant DNA molecules in B. subtilis
168.

<>

<1>Tandeau de Marsac, N., Houmard, J.
<2>Advances in cyanobacterial molecular genetics.
<3>Cyanobacteria, Elsevier, Fay, P., Van Baalen, C., 
<4>0
<5>251-302
<6>1987
<7>Until recently, the development of cyanobacteial genetics was limited by our
inability to transfer genetic material, both within a given species and between
different cyanobacterial species.  About 300 cyanobacterial strains have been
isolated as axenic cultures but less than 10, all unicellular species, have
been shown to be transformable by DNA.  Moreover, while specific
cyanobacteriophages have been described, gene transduction has not yet been
demonstrated.  Molecular genetics offers new possibilities for studying various
properties of cyanobacteria, such as oxygenic photosynthesis, aerobic and
anaerobic nitrogen fixation, light-regulated gene expression and cell
differentiation, properties for which biochemical and physiological data are
already available.  An increasing number of laboratories are now studying this
widespread and very large group of microorganisms using such genetic
approaches.  Recent reports that conjugation can occur in Anabaena/Nostoc
strains pave the way for the transfer of genetic material among filamentous
cyanobacteria and strengthen the work already initiated in order to understand,
at the molecular level, the important processes mentioned above.  We will
review current knowledge on cyanobacterial plasmids and restriction enzymes,
such data being of special interest for both molecular genetics and the
development of DNA transfer systems, and we will describe which cyanobacterial
genes have been cloned and characterized.  Finally, we will present some of the
features which have emerged from both sequence data and gene expression
experiments in homologous and/or heterologous hosts, as well as the
methodological approaches used.

<>

<1>Taneike, I., Otsuka, T., Dohmae, S., Saito, K., Ozaki, K., Takano, M., Higuchi, W., Takano, T., Yamamoto, T.
<2>Molecular nature of methicillin-resistant Staphylococcus aureus derived from explosive nosocomial outbreaks of the 1980s in Japan.
<3>FEBS Lett.
<4>580
<5>2323-2334
<6>2006
<7>Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA)
with Panton-Valentine leukocidin (PVL) genes is increasing worldwide.
Nosocomial outbreak-derived (hospital-acquired) MRSA (HA-MRSA) in Japan in
the 1980s was also largely PVL(+). PVL(+) HA-MRSA and CA-MRSA shared the
same multi-locus sequence type (ST30) and methicillin resistance cassette
(SCCmecIV), but were divergent in oxacillin resistance, spa typing, PFGE
analysis or clfA gene analysis. PVL(+) HA-MRSA, which probably originated
in PVL(+)S. aureus ST30, was highly adhesive (carrying cna and bbp genes),
highly-toxic (carrying luk(PV) and sea genes) and highly drug-resistant.
PVL(+) HA-MRSA was once replaced by other PVL(-) HA-MRSA (e.g., ST5), and
is re-emerging as CA-MRSA.

<>

<1>Tang, B., Wang, Q., Yang, M., Xie, F., Zhu, Y., Zhuo, Y., Wang, S., Gao, H., Ding, X., Zhang, L., Zhao, G., Zheng, H.
<2>ContigScape: a Cytoscape plugin facilitating microbial genome gap closing.
<3>BMC Genomics
<4>14
<5>289
<6>2013
<7>BACKGROUND: With the emergence of next-generation sequencing, the availability of
prokaryotic genome sequences is expanding rapidly. A total of 5,276 genomes have
been released since 2008, yet only 1,692 genomes were complete. The final phase
of microbial genome sequencing, particularly gap closing, is frequently the
rate-limiting step either because of complex genomic structures that cause
sequence bias even with high genomic coverage, or the presence of repeat
sequences that may cause gaps in assembly. RESULTS: We have developed a Cytoscape
plugin to facilitate gap closing for high-throughput sequencing data from
microbial genomes. This plugin is capable of interactively displaying the
relationships among genomic contigs derived from various sequencing formats. The
sequence contigs of plasmids and special repeats (IS elements, ribosomal RNAs,
terminal repeats, etc.) can be displayed as well. CONCLUSIONS: Displaying
relationships between contigs using graphs in Cytoscape rather than tables
provides a more straightforward visual representation. This will facilitate a
faster and more precise determination of the linkages among contigs and greatly
improve the efficiency of gap closing.

<>

<1>Tang, B., Zhao, W., Zheng, H., Zhuo, Y., Zhang, L., Zhao, G.P.
<2>Complete Genome Sequence of Amycolatopsis mediterranei S699 Based on De Novo Assembly via a Combinatorial Sequencing Strategy.
<3>J. Bacteriol.
<4>194
<5>5699-5700
<6>2012
<7>The genome of Amycolatopsis mediterranei S699 was resequenced and assembled de novo. By
comparing the sequences of S699 previously released and that of A.
mediterranei U32, about 10 kb of major indels was found to differ between the two
S699 genomes, and the differences are likely attributable to their different
assembly strategies.

<>

<1>Tang, C.
<2>Virulence genes, proteins, and their use.
<3>European Patent Office
<4>EP 1908839 A
<5>
<6>2008
<7>A series of genes from Neisseria meningitidis are shown to encode products which are
implicated in virulence.  The identification of these genes therefore allows attenuated
microorganisms to be produced.  Furthermore, the genes or their encoded products can be used
in the manufacture of vaccines for therapeutic application.

<>

<1>Tang, C.
<2>Virulence Gene and Protein, and Their Use.
<3>Korean Patent Office
<4>KR 1020027014876 A
<5>
<6>2002
<7>
<>

<1>Tang, C.U.
<2>Virulence genes, proteins, and their use.
<3>International Patent Office
<4>WO 0185772 A
<5>
<6>2001
<7>A series of genes from Neisseria meningitidis are shown to encode products which are
implicated in virulence.  The identification of these genes therefore allows attenuated
microorganisms to be produced.  Furthermore, the genes or their encoded products can be used
in the manufacture of vaccines for therapeutic application.

<>

<1>Tang, D., Ando, S., Takasaki, Y., Tadano, J.
<2>Mutational analyses of restriction endonuclease-HindIII mutant E86K with higher activity and altered specificity.
<3>Protein Eng.
<4>13
<5>283-289
<6>2000
<7>We have performed mutational analyses of restriction endonuclease HindIII in order to identify
the amino acid residues responsible for enzyme activity. Four of the seven HindIII mutants,
which had His-tag sequences at the N-termini, were expressed in Escherichia coli, and purified
to homogeneity. The His-tag sequence did not affect enzyme activity, whereas it hindered
binding of the DNA probe in gel retardation assays. A mutant E86K in which Lys was substituted
for Glu at residue 86 exhibited high endonuclease activity. Gel retardation assays showed high
affinity of this mutant to the DNA probe. Surprisingly, in the presence of a transition metal,
Mo(2+) or Mn(2+), the E86K mutant cleaved substrate DNA at a site other than HindIII.
Substitution of Glu for Val at residue 106 (V106E), and Asn for Lys at residue 125 (K125N)
resulted in a decrease in both endonucleolytic and DNA binding activities of the enzyme.
Furthermore, substitution of Leu for Asp at residue 108 (D108L) abolished both HindIII
endonuclease and DNA binding activities. CD spectra of the wild type and the two mutants, E86K
and D108L, were similar to each other, suggesting that there was little change in conformation
as a result of the mutations. These results account for the notion that Asp108 could be
directly involved in HindIII catalytic function, and that the substitution at residue 86 may
bring about new interactions between DNA and cations.

<>

<1>Tang, H., Yu, H., Li, Q., Wang, X., Gai, Z., Yin, G., Su, F., Tao, F., Ma, C., Xu, P.
<2>Genome Sequence of Pseudomonas putida Strain B6-2, a Superdegrader of Polycyclic Aromatic Hydrocarbons and Dioxin-Like Compounds.
<3>J. Bacteriol.
<4>193
<5>6789-6790
<6>2011
<7>Pseudomonas putida strain B6-2 can efficiently degrade environmental pollutants/toxicants,
such as polycyclic aromatic hydrocarbons and
dioxin-like compounds, and has unique tolerance to organic solvents. Here,
we present a 6.24-Mb draft genome sequence of B6-2, which could provide
further insights into the biodegradative mechanisms of a diverse range of
chemical compounds.

<>

<1>Tang, H., Yu, H., Tai, C., Huang, K., Liu, Y., Wang, L., Yao, Y., Wu, G., Xu, P.
<2>Genome Sequence of a Novel Nicotine-Degrading Strain, Pseudomonas geniculata N1.
<3>J. Bacteriol.
<4>194
<5>3553-3554
<6>2012
<7>A newly isolated bacterium, Pseudomonas geniculata N1, can efficiently degrade nicotine. Here
we present a 4.51-Mb assembly of its genome, which is the first
sequence of the P. geniculata group. The sequence contains the genes related to
nicotine catabolism and may provide insights into its molecular mechanism for
N-heterocyclic degradation.

<>

<1>Tang, J., Zhang, Y., Meng, H., Xue, Z., Ma, J.
<2>Complete Genome Sequence of Exiguobacterium sp. Strain MH3, Isolated from Rhizosphere of Lemna minor.
<3>Genome Announcements
<4>1
<5>e01059-13
<6>2013
<7>We report the complete genome sequence of Exiguobacterium sp. strain MH3, isolated from the
rhizosphere of duckweed. The genome assembly is 3.16 Mb, with a
G+C content of 47.24%, and it may provide useful information about plant-microbe
interactions and the genetic basis for the tolerance of the strain to various
environmental stresses.

<>

<1>Tang, K., Allman, S.L., Chen, C.H., Chang, L.Y., Schell, M.
<2>Matrix-assisted laser desorption/ionization of restriction enzyme-digested DNA.
<3>Rapid Commun. Mass. Spectrom.
<4>8
<5>183-186
<6>1994
<7>Matrix-assisted laser desorption/ionization, by a time-of-flight mass spectrometer, has been
successfully used for detection of restriction enzyme-digested DNA. However, the
oligonucleotide segments detected correspond to the molecular weights of single strands.

<>

<1>Tang, K., Liu, K., Jiao, N.
<2>Draft Genome Sequence of Oceaniovalibus guishaninsula JLT2003T.
<3>J. Bacteriol.
<4>194
<5>6683
<6>2012
<7>Oceaniovalibus guishaninsula, as a representative of a new genus within the family
Rhodobacteraceae, was isolated from surface seawater that was sulfidic.
Here, we present the draft genome sequence of the type strain, JLT2003(T).

<>

<1>Tang, K., Liu, K., Li, S., Jiao, N.
<2>Draft Genome Sequence of Strain JLT2015T, Belonging to the Family Sphingomonadaceae of the Alphaproteobacteria.
<3>Genome Announcements
<4>1
<5>e00226-13
<6>2013
<7>Strain JLT2015(T) was isolated from the southeastern Pacific, as a representative of a new
genus of the family Sphingomonadaceae of the Alphaproteobacteria. Here,
we present the draft genome sequence of strain JLT2015(T), which provides insight
into the oligotrophic strategy of this organism.

<>

<1>Tang, L.Y., Reddy, M.N., Rasheva, V., Lee, T.L., Lin, M.J., Hung, M.S., Shen, C.K.
<2>The eukaryotic DNMT2 genes encode a new class of cytosine-5 DNA methyltransferases.
<3>J. Biol. Chem.
<4>278
<5>33613-33616
<6>2003
<7>DNMT2 is a subgroup of the eukaryotic cytosine-5 DNA methyltransferase gene family. Unlike the
other family members, proteins encoded by DNMT2
genes were not known before to possess DNA methyltransferase activities.
Most recently, we have shown that the genome of Drosophila S2 cells stably
expressing an exogenous Drosophila dDNMT2 cDNA became anomalously
methylated at the 5'-positions of cytosines (Reddy, M. N., Tang, L. Y.,
Lee, T. L., and Shen, C.-K. J. (2003) Oncogene, in press). We present
evidence here that the genomes of transgenic flies overexpressing the
dDnmt2 protein also became hypermethylated at specific regions.
Furthermore, transient transfection studies in combination with sodium
bisulfite sequencing demonstrated that dDnmt2 as well as its mouse
ortholog, mDnmt2, are capable of methylating a cotransfected plasmid DNA.
These data provide solid evidence that the fly and mouse DNMT2 gene
products are genuine cytosine-5 DNA methyltransferases.

<>

<1>Tang, S., Edwards, E.A.
<2>Complete Genome Sequence of Bacteroidales Strain CF from a Chloroform-Dechlorinating Enrichment Culture.
<3>Genome Announcements
<4>1
<5>e01066-13
<6>2013
<7>Bacteroidales strain CF is the most abundant nondechlorinating organism in a
Dehalobacter-containing enrichment culture that consistently reductively
dechlorinates >50 mg/liter chloroform or 1,1,1-trichloroethane (methyl
chloroform). We assembled and closed the complete genome sequence of this
organism from the metagenomic sequencing data for enrichment cultures. This
organism is predicted to ferment l-lactate and ethanol.

<>

<1>Tang, S.L., Nuttall, S., Ngui, K., Fisher, C., Lopez, P., Dyall-Smith, M.
<2>HF2: a double-stranded DNA tailed haloarchaeal virus with a mosaic genome.
<3>Mol. Microbiol.
<4>44
<5>283-296
<6>2002
<7>HF2 is a haloarchaeal virus infecting two Halorubrum species (Family Halobacteriaceae). It is
lytic, has a head-and-tail morphology and belongs
to the Myoviridae (contractile tails). The linear double-stranded DNA
genome was sequenced and found to be 77 670 bp in length, with a mol% G+C
of 55.8. A total of 121 likely open reading frames (ORFs) were identified,
of which 37 overlapped at start and stop codons. The predicted proteins
were usually acidic (average pI of 4.8), and less than about 12% of them
had homologues in the sequence databases. Four complete tRNA-like
sequences (tRNA-Arg, -Asx, -Pro and -Tyr) and an incomplete tRNA-Thr were
detected. A transcription map showed that most of the genome was
transcribed and that the synthesis of transcripts occurred in a highly
organized and reproducible pattern over a 5 h infection cycle. Transcripts
often spanned multiple ORFs, suggesting that viral genes were organized
into operons. The predicted ORF and observed transcript directions matched
well and showed that transcription is mainly directed inwards from the
genome termini, meeting at about 45-48 kb, and this was also a turning
point in a cumulative GC-skew plot. The low point in cumulative GC-skew,
near the left end, was a region rich in short repeats and lacking ORFs,
which is likely to be an origin of replication. The HF2 genome is a mosaic
of components from widely different sources, demonstrating clearly that
viruses of haloarchaea, like their bacteriophage counterparts, are vectors
for the exchange and transmission of genetic material between wide
taxonomic distances, even across domains.

<>

<1>Tanizawa, Y., Fujisawa, T., Mochizuki, T., Kaminuma, E., Nakamura, Y., Tohno, M.
<2>Draft Genome Sequence of Lactobacillus oryzae Strain SG293T.
<3>Genome Announcements
<4>2
<5>e00861-14
<6>2014
<7>We report the 1.86-Mb draft genome and annotation of Lactobacillus oryzae SG293(T) isolated
from fermented rice grains. This genome information may provide
further insights into the mechanisms underlying the fermentation of rice grains.

<>

<1>Tanizawa, Y., Fujisawa, T., Mochizuki, T., Kaminuma, E., Suzuki, Y., Nakamura, Y., Tohno, M.
<2>Draft Genome Sequence of Weissella oryzae SG25T, Isolated from Fermented Rice Grains.
<3>Genome Announcements
<4>2
<5>e00667-14
<6>2014
<7>Weissella oryzae was originally isolated from fermented rice grains. Here we report the draft
genome sequence of the type strain of W. oryzae. This first
report on the genomic sequence of this species may help identify the mechanisms
underlying bacterial adaptation to the ecological niche of fermented rice grains.

<>

<1>Tanyashin, V.I., Li, L.I., Muijnieks, I.O., Baev, A.A.
<2>New endonuclease Hind GLU in Haemophilus influenzae Rd123.
<3>Dokl. Akad. Nauk.
<4>231
<5>226-228
<6>1976
<7>None

<>

<1>Tanyashin, V.I., Zimin, A.A., Shlyapnikov, M.G., Boronin, A.M.
<2>Transduction of plasmid antibiotic resistance determinants with pseudoT-even bacteriophages.
<3>Genetika
<4>39
<5>914-926
<6>2003
<7>Transduction of antibiotic resistance determinants of the plasmid pBR322 with pseudoT-even
bacteriophages RB42, RB43, and RB49 was
studied. It is established that antibiotic resistance determinants of
plasmid pBR322 from Escherichia coli recA(+) and recA(-) donor strains
do not differ significantly in respect to the efficiency of
transduction. Amber mutants RB43-21, RB43-33, and a double amber mutant
RB43am21am33 were obtained. These mutants facilitated transduction
experiments in some cases. Transduction of antibiotic resistance
markers of the vector plasmid pBR325 and recombinant plasmid pVT123,
containing a DNA fragment with hoc-segE-uvsW genes of phage T4, was
studied. The frequency of appearance of transductants resistant to
pseudoT-even bacteriophages used in transduction was determined, and
the sensitivity of resistant transductants to 32 RB bacteriophages and
also to phages lambda, T2, T4, T5, T6, T7, and BF23 was estimated. The
efficiency of plating pseudoT-even bacteriophages RB42 and RB43 on
strain E. coli 802 himA hip carrying mutations in genes that encode
subunits of the Integration Host Factor (IHF) was shown to be higher
than on isogenic strain E. coli 802. The growth of pseudoT-even
bacteriophages limited in vivo by modification - restriction systems of
chromosomal (EcoKI, EcoBI), phage ( EcoP1I), and plasmid (EcoRI,
EcoR124I, and EcoR124II) localization was analyzed. It was shown that
these phages were only slightly restricted by the type I modification -
restriction systems EcoBI, EcoR124I, and EcoR124II. Phage RB42 was
restricted by systems EcoKI, EcoP1I, and EcoRI; phage RB43, by
systems EcoKI and EcoRI; and phage RB49, by the Eco RI modification -
restriction system.

<>

<1>Tao, F., Shen, Y., Fan, Z., Tang, H., Xu, P.
<2>Genome Sequence of Pseudomonas putida S12, a Potential Platform Strain for Industrial Production of Valuable Chemicals.
<3>J. Bacteriol.
<4>194
<5>5985-5986
<6>2012
<7>Pseudomonas putida strain S12, a well-studied solvent-tolerant bacterium, is considered a
platform strain for the production of many chemicals. Here, we
present a 6.28-Mb assembly of its genome sequence. We have annotated 32 coding
sequences (CDSs) encoding efflux systems of organic compounds and 195 CDSs
responsible for the metabolism of aromatic compounds.

<>

<1>Tao, F., Tai, C., Liu, Z., Wang, A., Wang, Y., Li, L., Gao, C., Ma, C., Xu, P.
<2>Genome Sequence of Klebsiella pneumoniae LZ, a Potential Platform Strain for 1,3-Propanediol Production.
<3>J. Bacteriol.
<4>194
<5>4457-4458
<6>2012
<7>Klebsiella pneumoniae LZ is a bacterium isolated from soil which can produce 1,3-propanediol
from glycerol. Here we present a 5,431,750-bp assembly of its
genome sequence. We annotated 9 coding sequences (CDSs) responsible for glycerol
fermentation to 1,3-propanediol, 19 CDSs encoding glycerol utilization, and 134
CDSs related to its virulence and defense.

<>

<1>Tao, F., Tang, H., Gai, Z., Su, F., Wang, X., He, X., Xu, P.
<2>Genome Sequence of Pseudomonas putida Idaho, a Unique Organic-Solvent-Tolerant Bacterium.
<3>J. Bacteriol.
<4>193
<5>7011-7012
<6>2011
<7>Pseudomonas putida Idaho is an organic-solvent-tolerant strain which can degrade and adapt to
high concentrations of organic solvents. Here, we
announce its first draft genome sequence (6,363,067 bp). We annotated 192
coding sequences (CDSs) responsible for aromatic compound metabolism, 40
CDSs encoding phospholipid synthesis, and 212 CDSs related to stress
response.

<>

<1>Tao, F., Wang, X., Ma, C., Yang, C., Tang, H., Gai, Z., Xu, P.
<2>Genome Sequence of Xanthomonas campestris JX, an Industrially Productive Strain for Xanthan Gum.
<3>J. Bacteriol.
<4>194
<5>4755-4756
<6>2012
<7>Xanthomonas campestris JX, a soil bacterium, is an industrially productive strain for xanthan
gum. Here we present a 5.0-Mb assembly of its genome sequence. We
have annotated 12 coding sequences (CDSs) responsible for xanthan gum
biosynthesis, 346 CDSs encoding carbohydrate metabolism, and 69 CDSs related to
virulence, defense, and plant disease.

<>

<1>Tao, F., Zhao, P., Li, Q., Su, F., Yu, B., Ma, C., Tang, H., Tai, C., Wu, G., Xu, P.
<2>Genome Sequence of Rhodococcus erythropolis XP, a Biodesulfurizing Bacterium with Industrial Potential.
<3>J. Bacteriol.
<4>193
<5>6422-6423
<6>2011
<7>Rhodococcus erythropolis strains have shown excellent characteristics in petroleum oil
biodesulfurization. Here we present the first announcement
of the draft genome sequence of an efficient biodesulfurizing bacterium
named R. erythropolis XP (7,229,582 bp). The biodesulfurizing genes dszABC
are located on a plasmid, while the flavin reductase gene dszD is located
on the chromosome.

<>

<1>Tao, T.
<2>Characterization of the cloned PvuII type II restriction-modification system in Escherichia coli.
<3>Diss. Abstr.
<4>53
<5>115B
<6>1992
<7>The PvuII type II restriction-modification system (RMS2) cloned from Proteus vulgaris has been
sequenced in full and characterized in Escherichia coli. The sequence of pvuIIM showed that
this N4-methylcytosine (N4mC) generating DNA methyltransferase (MTase) is similar to
N6-methyladenine (N6mA) DNA MTases. Newly available data on other N4mC MTases support this
observation. Together with the known DNA structural data, this strongly suggests that these
two groups of DNA amino MTases are functionally and possibly evolutionarily related. It was
also found that M.PvuII contains an internal homology so far unique among type II DNA MTases.
Each homologous part includes its own DPPY and F/GXGXG motifs, which are sequences common to
all DNA amino MTases. Hypotheses for this homology were proposed. R.PvuII was shown to be a
homodimer. A segment of R.PvuII was found to be similar to R.BamHI at the amino acid sequence
level. The importance of this similarity is not known, but such similarities are very rare
among the sequenced endonuclease genes. A third open reading frame (ORF), 84 codons in length
with potential transcription and translation elements, was found to be associated with this
RMS2. Sequence comparison has revealed strong homology between this ORF and that of bacterial
and bacterial phage repressors. Genetic assays on this ORF showed that mutations introduced
into this ORF resulted in the reduction of R.PvuII production up to a 10/5-fold in vivo. This
effect could be complemented by providing the third ORF in trans. The third ORF was named
pvuIIC (for controller). Further experiments showed that a product of the expected size was
indeed made by pvuIIC both in vivo and in vitro. The pvuIIC product, C.PvuII, was shown to
bind to DNA, either alone or with other proteins. The binding site responsible for pvuIIR
regulation was narrowed down to a 70 bp fragment upstream of pvuIIR by deletion analysis. The
binding specificity has not yet been defined in vitro. A regulatory mechanism incorporating
these findings was hypothesized and further experiments to test the hypotheses discussed.

<>

<1>Tao, T., Blumenthal, R.M.
<2>Sequence and subunit structure of restriction endonuclease PvuII.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>175
<6>1991
<7>We have determined the DNA base sequence of the pvuIIR gene and the subunit
structure of its protein product.  The pvuIIR gene codes for a type II
restriction endonuclease, PvuII, which cleaves the sequence 5'-CAGCTG-3'
between the central two bases.  The pvuIIR open reading frame predicts a 157
amino acid, 18.4 kDa polypeptide.  The predicted size and amino acid sequence
are consistent with previous analyses of in vitro translation products and of
the N-terminal sequence of the purified protein.  The elution profile of active
PvuII endonuclease from a calibrated gel filtration column suggests a molecular
mass of about 40.5 kDa, with a Stokes radius of about 1.8nm, implying that the
enzyme is a homodimer.  The pvuIIR gene shows no obvious relationship to that
of pvuIIM, which codes for the PvuII methylase, but a region of similarity has
been found among the amino acid sequences of pvuIIR and those of four other
restriction endonucleases.  Combining the pvuIIR and pvuIIM sequence data, to
yield the full sequence of the divergently-transcribed restriction-modification
system, revealed an additional open reading frame between these two genes.  In
a separate study, this third open reading frame has been shown to regulate the
expression of pvuIIR.

<>

<1>Tao, T., Blumenthal, R.M.
<2>Sequence and characterization of pvuIIR, the PvuII endonuclease gene, and of pvuIIC, its regulatory gene.
<3>J. Bacteriol.
<4>174
<5>3395-3398
<6>1992
<7>An open reading frame partially overlaps pvuIIR, and genetic evidence implies that this open
reading frame, named pvuIIC, specifies a positive regulator of pvuIIR (T. Tao, J.C. Bourne,
and R.M. Blumenthal, J. Bacteriol. 173:1367-1375, 1991). Inducible constructs of pvuIIC
produced a protein of the expected size. The site of C.PvuII action appears to lie within
pvuIIC itself; thus, pvuIIC may be a self-contained regulatory cassette.

<>

<1>Tao, T., Bourne, J.C., Blumenthal, R.M.
<2>A family of regulatory genes associated with Type II restriction-modification systems.
<3>J. Bacteriol.
<4>173
<5>1367-1375
<6>1991
<7>Restriction-modification systems must be regulated to avoid autorestriction and
death of the host cell.  An open reading frame (ORF) in the PvuII
restriction-modification system appears to code for a regulatory protein from a
previously unrecognized family.  First, interruptions of this ORF result in a
nonrestricting phenotype.  Second, this ORF can restore restriction competence
to such interrupted mutants in trans.  Third, the predicted amino acid sequence
of this ORF resembles those of known DNA-binding proteins and includes a
probable helix-turn-helix motif.  A survey of unattributed ORFs in 15 other
type II restriction-modification systems revealed three that closely resemble
the PvuII ORF.  All four members of this putative regulatory gene family have a
common position relative to the endonuclease genes, suggesting a common
regulatory mechanism.

<>

<1>Tao, T., Walter, J., Brennan, K.J., Cotterman, M.M., Blumenthal, R.M.
<2>Sequence, internal homology and high-level expression of the gene for a DNA-(cytosine N4)-methyltransferase, M.Pvu II.
<3>Nucleic Acids Res.
<4>17
<5>4161-4175
<6>1989
<7>The base sequence of the pvuIIM gene has been determined.  This gene codes for
a DNA-(cytosine N4)-methyltransferase, M.Pvu II.  The base sequence contains a
single large open reading frame that predicts a 38.3kDa polypeptide, consistent
with experimental data.  The pvuIIM gene contains some sequences common to DNA
methyltransferases in general, but includes none of the sequences specifically
conserved among DNA-(cytosine 5)-methyltransferases.  The pvuIIM sequence also
reveals an internal homology at the amino acid level, each half of which spans
over 100 amino acids and is itself homologous to the sequences of some
DNA-(adenine N6)-methyltransferases.  A derivative of the pvuIIM plasmid was
constructed to allow high-level production of M.Pvu II.  Specifically, the
composite Ptac promoter was inserted 5' to pvuIIM, intervening DNA was deleted,
and the resulting construct was used to transform an mcrB lacIq strain of
Escherichia coli.  When this transformant was induced with
isopropyl-B-D-galactopyranoside (IPTG), growth rapidly ceased and M.Pvu II
accumulated to the point of comprising over 10% of the total soluble protein.

<>

<1>Tapia, P., Flores, F.M., Covarrubias, P.C., Acuna, L.G., Holmes, D.S., Quatrini, R.
<2>Complete Genome Sequence of Temperate Bacteriophage AcaML1 from the Extreme Acidophile Acidithiobacillus caldus ATCC 51756.
<3>J. Virol.
<4>86
<5>12452-12453
<6>2012
<7>Development of reproducible genetic tools in the industrially important
acidithiobacilli is urgently required. Inducible temperate phages which may be
modified in vitro, propagated in suitable hosts, and used to transduce relevant
genetic information to other strains and/or species are potentially valuable
tools in this field of research. In order to address these current limitations,
the genome sequence of an inducible temperate Myoviridae-like bacteriophage from
the Acidithiobacillus caldus type strain was annotated and analyzed
bioinformatically. Here, we announce the genome sequence of AcaML1 and report
major findings from its annotation.

<>

<1>Taranto, M.P., Villena, J., Salva, S., Alvarez, S., Savoy-de-Giori, G., Font-de-Valdez, G., Hebert, E.M.
<2>Draft Genome Sequence of Lactobacillus rhamnosus CRL1505, an Immunobiotic Strain  Used in Social Food Programs in Argentina.
<3>Genome Announcements
<4>1
<5>e00627-13
<6>2013
<7>We report the draft genome sequence of the probiotic Lactobacillus rhamnosus strain CRL1505.
This new probiotic strain has been included into official Nutritional Programs in Argentina.
The draft genome sequence is composed of 3,417,633 bp with 3,327 coding sequences.

<>

<1>Tarasova, G.V., Nayakshina, T.N., Degtyarev, S.K.
<2>Substrate specificity of new methyl-directed DNA endonuclease GlaI.
<3>BMC Mol. Biol.
<4>9
<5>7
<6>2008
<7>Recently, we have discovered site-specific endonucleases, which recognize and cleave only DNA
sequences with 5-methylcytosine. Two specificities of such endonucleases have been described.
Enzymes BisI, BlsI, and GluI are isoschizomers and hydrolyze the DNA sequence
5'-GCNGC-3'/3'-CGNCG-5', which is methylated in different ways. The enzyme GlaI cleaves
the DNA sequence 5'-GCGC-3'/3'-CGCG-5' if there are two, three or four 5-methylcytosines.
The goal of the present work is to study in detail the composition of recognition sequence and
effect of the methylated cytosines on the efficiency of DNA cleavage by the methyl-directed
DNA endonuclease GlaI. In a recent work we have studied the dependence of GlaI activity on the
quantity and location of 5-methylcytosines in the enzyme recognition sequence
5'-GCGC-3'/3'-CGCG-5'. A significant DNA cleavage has been observed for oligonucleotide
duplexes, which include either three or four 5-methylcytosines. In this work we have studied
dependence of the GlaI activity on quantity and location of methylated cytosines, as well as
on composition of the recognition sequence.The list of good substrates for GlaI includes a
fully methylated site 5'-CGCG-3'/3'-GCGC-5', sites with three cytosines of a general
structure 5'-PuMGM-3'/3'-PyGMG-5', and one recognition sequence with two methylated
cytosines 5'-AMGT-3'/3'-TGMA-5', where M is 5-methylcytosine. GlaI intermediate substrates
include sites with three methylated cytosines of a general structure
5'-GMPuM-3'/3'-MGPyG-5', as well as a site with two methylcytosines
5'-GMGT-3'/3'-CGMA-5'. The site 5'-GMGC-3'/3'-CGMG-5' may be considered a low activity
substrate.

<>

<1>Tarasova, M.V., Kuznetsov, V.V., Netesova, N.A., Gonchar, D.A., Degtyarev, S.C.
<2>Cloning and analysis of biochemical and catalytic properties of DNA methyltransferase M1.BspACI.
<3>Vestn. Mosk. Univ.
<4>0
<5>38-40
<6>2011
<7>Genes coding for the DNA methyltransferases of restriction-modification system BspACI from
Bacillus psychrodurans AC have been cloned in E. coli cells. The analysis of amino acid
sequences of the proteins showed that both these genes belong to C5 DNA methyltransferases.
Gene bspACIMI has been subcloned in pJW2 vector. High-purity recombinant enzyme has been
obtained with chromatography on different carriers. It has been shown that M1.BspACI modifies
the first cytosine residue in the sequence 5'-CCGC-3'. Kinetic parameters have been
determined. The catalytic constant appears to be 0,095 +/- 0,002 min(-1), K-m, (Delta HK phi
ara lambda) - 0,053 +/- 0,007 (MKM), K-m (SAM) - 5,1 +/- 0,3 (MKM).

<>

<1>Tarasova, M.V., Kuznetsov, V.V., Netesova, N.A., Gonchar, D.A., Degtyarev, S.K.
<2>Recombinant DNA-Methyltransferase M1.BspACI from Bacillus psychrodurans AC: Purification and Properties.
<3>Biochemistry
<4>75
<5>1484-1490
<6>2010
<7>A restriction-modification system from Bacillus psychrodurans AC ( recognition sequence
5'-CCGC-3') comprises two DNA methyltransferases:
M1.BspACI and M2.BspACI. The bspACIM1 gene was cloned in the pJW2
vector and expressed in Escherichia coli cells. High-purity M1.BspACI
preparation has been obtained by chromatography on different carriers.
M1.BspACI has a temperature optimum of 30 C and demonstrates maximum
activity at pH 8.0. M1.BspACI modifies the first cytosine in the
recognition sequence 5'-CCGC-3'. The kinetic parameters of M1.BspACI
DNA methylation are as follows: K-m for phage lambda DNA is 0.053 mu M
and K-m for S-adenosyl-L-methionine is 5.1 mu M. The catalytic constant
(k(cat)) is 0.095 min(-1).

<>

<1>Tarazona, D., Borda, V., Galarza, M., Agapito, J.C., Guio, H.
<2>Functional Analysis Using Whole-Genome Sequencing of a Drug-Sensitive Mycobacterium tuberculosis Strain from Peru.
<3>Genome Announcements
<4>2
<5>e00087-14
<6>2014
<7>We report the whole-genome sequence of a Latin American-Mediterranean (LAM) lineage
drug-sensitive Mycobacterium tuberculosis strain from Peru, INS-SEN. The
functional analysis revealed more mutations in secondary metabolite biosynthesis,
transport, and catabolism (clusters of orthologous groups [COG] category Q) than
for other LAM-sensitive strains. This study contributes to the understanding of
the genomic diversity of drug-sensitive M. tuberculosis.

<>

<1>Tarazona, D., Padilla, C., Caceres, O., Montenegro, J.D., Bailon, H., Ventura, G., Mendoza, G., Anaya, E., Guio, H.
<2>Whole Genome Sequencing and Comparative Analysis of Bartonella bacilliformis Strain INS, the Causative Agent of Carrion's Disease.
<3>Genome Announcements
<4>1
<5>e00053-12
<6>2013
<7>Bartonella bacilliformis is the etiological agent of human bartonellosis, which is highly
endemic to Peru. Here, we report the first genome that was sequenced
and analyzed from an isolate of B. bacilliformis strain INS, which originally was
isolated from the blood of an infected patient with an acute phase of Carrion's
disease from Jaen-Cajamarca, Peru.

<>

<1>Tardy-Planechaud, S., Fujimoto, J., Lin, S.S., Sowers, L.C.
<2>Solid phase synthesis and restriction endonuclease cleavage of oligodeoxynucleotides containing 5-(hydroxymethyl)-cytosine.
<3>Nucleic Acids Res.
<4>25
<5>553-558
<6>1997
<7>Emerging data suggest an important role for cytosine methylation in tumorigenesis.
Simultaneously, recent studies indicate a significant contribution of endogenous oxidative DNA
damage to the development of human disease.  Oxidation of the 5-methyl group of
5-methylcytosine (5mC) residues in DNA results in the formation of 5-(hydroxymethyl)cytosine
(hmC).  The biological consequences of hmC residues in vertebrate DNA are as yet unknown;
however, conversion of the hydrophobic methyl group to the hydrophilic hydroxymethyl group may
substantially alter the interaction of sequence-specific binding proteins with DNA.  Central
to both biophysical and biochemical studies on the potential consequences of specific DNA
damage products such as hmC are efficient methods for the synthesis of oligodeoxynucleotides
containing such modified bases at selected positions.  In this paper, we describe a method for
the placement of hmC residues in oligodeoxynucleotides using established phosphoramidite
chemistry.  In addition, we have examined the influence of specific hmC residues on enzymatic
cleavage of oligodeoxynucleotides by the methylation-sensitive restriction endonucleases MspI
and HpaII.

<>

<1>Taron, C.H., Van Cott, E.M., Wilson, G.G., Moran, L.S., Slatko, B.E., Hornstra, L.J., Benner, J.S., Kucera, R.B., Guthrie, E.P.
<2>Cloning, sequencing and overexpressing the NaeI restriction endonuclease gene from Nocardia aerocolonigenes.
<3>J. Cell Biochem. Suppl.
<4>17C
<5>175
<6>1993
<7>NaeI is a type II restriction-modification system from Nocardia aerocolonigenes. The
endonuclease recognizes the palindromic hexanucleotide sequence 5'GCC^GGC 3' and cleaves to
produce a blunt end. The NaeI methylase gene (naeIM) was cloned previously by Van Cott and
Wilson (Gene 74: 55-59, 1988). This clone exhibited methylase activity but no levels of
endonuclease activity were detected, suggesting the endonuclease gene (naeIR) was either not
present intact on the clone, or was present but not expressed. Subcloning the methylase clone
into Streptomyces lividans, an organism more closely related to Nocardia, confirmed the
absence of naeIR. To clone naeIR, a SacI genomic DNA library was made using the original
methylase clone in pBR322 as a cloning vector. This library was selected for naeIM by
digestion with NaeI, transformed into E. coli AP1-200 and positive clones appeared as blue
clonies on LB agar plates containing X-gal. Nucleotide sequencing of naeIM and naeIR yielded
two large open reading frames (ORFs). The naeIR ORF was identified by correlation of its
translated peptide sequence to the amino terminal peptide sequence of the NaeI endonuclease.
The endonuclease gene was amplified by PCR and cloned behind the strong promoter Ptac. This
clone was transformed into E. coli K802 containing the MspI methylase gene. The MspI
recognition sequence is 5'CCGG 3' (the internal tetramer of the NaeI recognition site) with
MspI modification fully protecting against NaeI digestion. This construct yielded
approximately 750 fold overexpression of NaeI.

<>

<1>Taron, C.H., Van Cott, E.M., Wilson, G.G., Moran, L.S., Slatko, B.E., Hornstra, L.J., Benner, J.S., Kucera, R.B., Guthrie, E.P.
<2>Cloning and expression of the NaeI restriction endonuclease-encoding gene and sequence analysis of the NaeI restriction-modification system.
<3>Gene
<4>155
<5>19-25
<6>1995
<7>NaeI, a type-II restriction-modification (R-M) system from the bacterium Nocardia
aerocolonigenes, recognizes the sequence 5'-GCCGGC. The NaeI DNA methyltransferase
(MTase)-encoding gene, naeIM, had been cloned previously in Escherichia coli. However, none of
these clones expressed detectable levels of the restriction endonuclease (ENase). The absence
of the intact ENase-encoding gene (naeIR) within the isolated MTase clones was confirmed by
recloning the MTase clones into Streptomyces lividans. The complete NaeI system was finally
cloned using E. coli AP1-200 and less stringent MTase-selection conditions. The naeIR gene was
expressed first by cloning into S. lividans, and later by cloning under control of a regulated
promoter in an E. coli strain preprotected by the heterologous MspI MTase (M.MspI). The DNA
sequence of the NaeI R-M system has been determined, analyzed and compared to previously
sequenced R-M systems.

<>

<1>Tasara, T., Weinmaier, T., Klumpp, J., Rattei, T., Stephan, R.
<2>Complete Genome Sequence of Listeria monocytogenes Lm60, a Strain with an Enhanced Cold Adaptation Capacity.
<3>Genome Announcements
<4>2
<5>e01248-14
<6>2014
<7>The complete genome sequence of Listeria monocytogenes Lm60, a fast cold-adapting serotype
1/2a human isolate, is presented.

<>

<1>Tasse, L., Bercovici, J., Pizzut-Serin, S., Robe, P., Tap, J., Klopp, C., Cantarel, B.L., Coutinho, P.M., Henrissat, B., Leclerc, M., Dore, J., Monsan, P., Remaud-Simeon, M., Potocki-Veronese, G.
<2>Functional metagenomics to mine the human gut microbiome for dietary fiber catabolic enzymes.
<3>Genome Res.
<4>20
<5>1605-1612
<6>2010
<7>The human gut microbiome is a complex ecosystem composed mainly of
uncultured bacteria. It plays an essential role in the catabolism of
dietary fibers, the part of plant material in our diet that is not
metabolized in the upper digestive tract, because the human genome does
not encode adequate carbohydrate active enzymes (CAZymes). We describe a
multi-step functionally based approach to guide the in-depth
pyrosequencing of specific regions of the human gut metagenome encoding
the CAZymes involved in dietary fiber breakdown. High-throughput
functional screens were first applied to a library covering 5.4 x 10(9) bp
of metagenomic DNA, allowing the isolation of 310 clones showing
beta-glucanase, hemicellulase, galactanase, amylase, or pectinase
activities. Based on the results of refined secondary screens, sequencing
efforts were reduced to 0.84 Mb of nonredundant metagenomic DNA,
corresponding to 26 clones that were particularly efficient for the
degradation of raw plant polysaccharides. Seventy-three CAZymes from 35
different families were discovered. This corresponds to a fivefold
target-gene enrichment compared to random sequencing of the human gut
metagenome. Thirty-three of these CAZy encoding genes are highly
homologous to prevalent genes found in the gut microbiome of at least 20
individuals for whose metagenomic data are available. Moreover, 18
multigenic clusters encoding complementary enzyme activities for plant
cell wall degradation were also identified. Gene taxonomic assignment is
consistent with horizontal gene transfer events in dominant gut species
and provides new insights into the human gut functional trophic chain.

<>

<1>Tasseron-de Jong, J.G., Aker, J., Giphart-Gassler, M.
<2>The ability of the restriction endonuclease EcoRI to digest hemi-methylated versus fully cytosine-methylated DNA of the herpes tk promoter region.
<3>Gene
<4>74
<5>147-149
<6>1988
<7>None

<>

<1>Tateishi, Y., Kitada, S., Miki, K., Maekura, R., Ogura, Y., Ozeki, Y., Nishiuchi, Y., Niki, M., Hayashi, T., Hirata, K., Kobayashi, K., Matsumoto, S.
<2>Whole-Genome Sequence of the Hypervirulent Clinical Strain Mycobacterium intracellulare M.i.198.
<3>J. Bacteriol.
<4>194
<5>6336
<6>2012
<7>We report herein the draft genome sequence of Mycobacterium intracellulare clinical strain
M.i.198, which consistently exhibits hypervirulence in human
patients, human macrophages in vitro, and immunocompetent mice.

<>

<1>Tateishi, Y., Tamaru, A., Ogura, Y., Niki, M., Wada, T., Yamamoto, T., Hirata, K., Hayashi, T., Matsumoto, S.
<2>Whole-Genome Sequence of the Potentially Hypertransmissible Multidrug-Resistant Mycobacterium tuberculosis Beijing Strain OM-V02_005.
<3>Genome Announcements
<4>1
<5>e00608-13
<6>2013
<7>We report the draft genome sequence of Mycobacterium tuberculosis Beijing strain  OM-V02_005,
which exhibits possible hypertransmissible characteristics among the
population of patients with multidrug-resistant tuberculosis in Osaka Prefecture,
the largest urban area in western Japan.

<>

<1>Tatti, K.M., Loparev, V.N., Ranganathanganakammal, S., Changayil, S., Frace, M., Weil, M.R., Sammons, S., Maccannell, D., Mayer, L.W., Tondella, M.L.
<2>Draft Genome Sequences of Bordetella holmesii Strains from Blood (F627) and Nasopharynx (H558).
<3>Genome Announcements
<4>1
<5>e0005613
<6>2013
<7>Bordetella holmesii, a human pathogen, can confound the diagnosis of respiratory  illness
caused by Bordetella pertussis. We present the draft genome sequences of
two B. holmesii isolates, one from blood, F627, and one from the nasopharynx,
H558. Interestingly, important virulence genes that are present in B. pertussis
are not found in B. holmesii.

<>

<1>Tatum, F.M., Briggs, R.E.
<2>Isolation, identification, and cloning of a non-palindromic type II DNA restriction endonuclease, PhaI, from Pasteurella haemolytica.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>93
<5>199
<6>1993
<7>Pasteurella haemolytica is an important cause of bovine pneumonic pasteurellosis. PhaI, an
isoschizomer of SfaNI, was isolated from P. haemolytica, strain #NADC-60, a plasmidless
isolate of serotype 1. PhaI recognizes the 5 base non-palindromic sequence 5'-GATGC-3'.
Analysis of the restriction products on sequencing gels showed that cleavage occurs 5 bases to
the right of the noted recognition site and 9 bases to the right on the opposite strand. A
clone encoding the PhaI restriction endonuclease and PhaI methyltransferase was isolated from
a P. haemolytica cosmid library and functional expression of both genes was obtained in
Escherichia coli. Preliminary results indicate that PhaI restriction activity is a barrier to
the introduction or establishment of exogenous DNA into this bacterium.

<>

<1>Tauch, A., Kaiser, O., Hain, T., Goesmann, A., Weisshaar, B., Albersmeier, A., Bekel, T., Bischoff, N., Brune, I., Chakraborty, T., Kalinowski, J., Meyer, F., Rupp, O., Schneiker, S., Viehoever, P., Puhler, A.
<2>Complete genome sequence and analysis of the multiresistant nosocomial pathogen Corynebacterium jeikeium K411, a lipid-requiring bacterium of the human skin flora.
<3>J. Bacteriol.
<4>187
<5>4671-4682
<6>2005
<7>Corynebacterium jeikeium is a "lipophilic" and multidrug-resistant bacterial species of the
human skin flora that has been recognized with increasing frequency as a serious nosocomial
pathogen. Here we report the genome sequence of the clinical isolate C. jeikeium K411, which
was initially recovered from the axilla of a bone marrow transplant patient. The genome of C.
jeikeium K411 consists of a circular chromosome of 2,462,499 bp and the 14,323-bp
bacteriocin-producing plasmid pKW4. The chromosome of C. jeikeium K411 contains 2,104
predicted coding sequences, 52% of which were considered to be orthologous with genes in the
Corynebacterium glutamicum, Corynebacterium efficiens, and Corynebacterium diphtheriae
genomes. These genes apparently represent the chromosomal backbone that is conserved between
the four corynebacteria. Among the genes that lack an ortholog in the known corynebacterial
genomes, many are located close to transposable elements or revealed an atypical G+C content,
indicating that horizontal gene transfer played an important role in the acquisition of genes
involved in iron and manganese homeostasis, in multidrug resistance, in bacterium-host
interaction, and in virulence. Metabolic analyses of the genome sequence indicated that the
"lipophilic" phenotype of C. jeikeium most likely originates from the absence of fatty acid
synthase and thus represents a fatty acid auxotrophy. Accordingly, both the complete gene
repertoire and the deduced lifestyle of C. jeikeium K411 largely reflect the strict dependence
of growth on the presence of exogenous fatty acids. The predicted virulence factors of C.
jeikeium K411 are apparently involved in ensuring the availability of exogenous fatty acids by
damaging the host tissue.

<>

<1>Tauch, A., Kirchner, O., Loffler, B., Gotker, S., Puhler, A., Kalinowski, J.
<2>Efficient electrotransformation of Corynebacterium diphtheriae with a mini-replicon derived from the Corynebacterium glutamicum plasmid pGA1.
<3>Curr. Microbiol.
<4>45
<5>362-367
<6>2002
<7>Efficient transformation of the human pathogen Corynebacterium
diphtheriae was achieved with novel cloning vectors consisting of a mini-replicon from the
cryptic C. glutamicum plasmid pGA1 as well as of the aph(3')-IIa or tetA(Z) antibiotic
resistance genes. Plasmid-containing transformants of C. diphtheriae were recovered at
frequencies ranging from 1.3 x 10^5 to 4.8 x 10^6 colony forming units (cfu)/microgram of
plasmid DNA. Vector DNA was directly transferred from Escherichia coli into C. diphtheriae
with frequencies up to 5.6 x 10^5 cfu/microgram of plasmid DNA. On the basis of the pGA1
mini-replicon, an expression vector system was established for C. diphtheriae by means of the
P-tac promoter and the green fluorescent reporter protein. In addition, other commonly used
vector systems from C. glutamicum, including the pBL1 and pHM1519 replicons, and the sacB
conditionally lethal selection market from Bacillus subtilis, were shown to be functional in
C. diphtheriae. Thus, the ability to apply the standard methods of C. glutamicum recombinant
DNA technology will greatly facilitate the functional analysis of the recently completed C.
diphtheriae genome sequence.

<>

<1>Tauch, A., Kirchner, O., Wehmeier, L., Kalinowski, J., Puhler, A.
<2>Corynebacterium glutamicum DNA is subjected to methylation-restriction in Escherichia coli.
<3>FEMS Microbiol. Lett.
<4>123
<5>343-347
<6>1994
<7>Efficient electroporation of Escherichia coli with plasmid DNA isolated from Corynebacterium
glutamicum depends on the use of Mcr-deficient E. coli strains. The transformation frequency
increased nearly 800-fold when the Mcr-deficient E. coli DH5a MCR was used instead of E. coli
DH5a. We used E. coli strains with different mutations in the methyl-specific restriction
systems to show that McrBC-deficiency is sufficient to generate this effect. The results imply
that C. glutamicum DNA contains methylcytosine in specific sequences recognized by the E. coli
McrBC system.

<>

<1>Tauch, A., Krieft, S., Kalinowski, J., Puhler, A.
<2>The 51,409-bp R-plasmid pTP10 from the multiresistant clinical isolate Corynebacterium striatum M82B is composed of DNA segments initially identified in soil bacteria and in plant, animal, and human pathogens.
<3>Mol. Gen. Genet.
<4>263
<5>1-11
<6>2000
<7>The 51,409-bp DNA sequence of the multiresistance plasmid pTP10 from the
gram-positive opportunistic human pathogen Corynebacterium striatum M82B
has been determined. Fully automated genome interpretation led to the
identification of 47 ORFs. Analysis of the genetic organization of pTP10
suggests that the plasmid is composed of eight DNA segments, the
boundaries of which are represented by transposons and insertion
sequences. The DNA segments of pTP10 are highly similar to (1) a
plasmid-encoded erythromycin resistance region from the human pathogen
Corynebacterium diphtheriae; (2) a chromosomal DNA region from
Mycobacterium tuberculosis; (3) a plasmid-encoded chloramphenicol
resistance region from the soil bacterium Corynebacterium glutamicum; (4)
transposable elements from phytopathogenic gram-negative Pseudomonas,
Xanthomonas and Erwinia species; and (5) a plasmid-encoded aminoglycoside
resistance region from the gram-negative fish pathogen Pasteurella
piscicida. The complete DNA sequence of pTP10 provides genetic information
regarding the mechanisms of resistance to 16 antimicrobial agents that
belong to six structural classes. In addition, the mosaic structure of
pTP10 represents the evolutionary consolidation into a single plasmid
molecule of antimicrobial resistances from microorganisms found in
different habitats by means of mobile elements, resulting in the
generation of a multiresistant bacterium that can infect humans.

<>

<1>Tauch, A., Trost, E., Tilker, A., Ludewig, U., Schneiker, S., Goesmann, A., Arnold, W., Bekel, T., Brinkrolf, K., Brune, I., Gotker, S., Kalinowski, J., Kamp, P.B., Lobo, F.P., Viehoever, P., Weisshaar, B., Soriano, F., Droge, M., Puhler, A.
<2>The lifestyle of Corynebacterium urealyticum derived from its complete genome sequence established by pyrosequencing.
<3>J. Biotechnol.
<4>136
<5>11-21
<6>2008
<7>Corynebacterium urealyticum is a lipid-requiring, urealytic bacterium of the human skin flora
that has been recognized as causative agent of
urinary tract infections. We report the analysis of the complete genome
sequence of C. urealyticum DSM7109, which was initially recovered from a
patient with alkaline-encrusted cystitis. The genome sequence was
determined by a combination of pyrosequencing and Sanger technology. The
chromosome of C. urealyticum DSM7109 has a size of 2,369,219bp and
contains 2024 predicted coding sequences, of which 78% were considered as
orthologous with genes in the Corynebacterium jeikeium K411 genome.
Metabolic analysis of the lipid-requiring phenotype revealed the absence
of a fatty acid synthase gene and the presence of a beta-oxidation pathway
along with a large repertoire of auxillary genes for the degradation of
exogenous fatty acids. A urease locus with the gene order ureABCEFGD may
play a pivotal role in virulence of C. urealyticum by the alkalinization
of human urine and the formation of struvite stones. Multidrug resistance
of C. urealyticum DSM7109 is mediated by transposable elements, conferring
resistances to macrolides, lincosamides, ketolides, aminoglycosides,
chloramphenicol, and tetracycline. The complete genome sequence of C.
urealyticum revealed a detailed picture of the lifestyle of this
opportunistic human pathogen.

<>

<1>Tautz, N., Kaluza, K., Frey, B., Jarsch, M., Schmitz, G.G., Kessler, C.
<2>SgrAI, a novel class-II restriction endonuclease from Streptomyces griseus recognizing the octanucleotide sequence 5'-CR/CCGGYG-3' .
<3>Nucleic Acids Res.
<4>18
<5>3087
<6>1990
<7>
<>

<1>Taveirne, M.E., Dunham, D.T., Miller, W.G., Parker, C.T., Huynh, S., DiRita, V.J.
<2>Complete Genome Sequence and Annotation of a Campylobacter jejuni Strain, MTVDSCj20, Isolated from a Naturally Colonized Farm-Raised Chicken.
<3>Genome Announcements
<4>2
<5>e00852-14
<6>2014
<7>Campylobacter jejuni is a major cause of human food-borne illness, with contaminated poultry
products serving as a main source of human infection. C.
jejuni strain MTVDSCj20 was isolated from the cecal contents of a farm-raised
chicken that was naturally colonized with Campylobacter. We present here the
complete annotated genome sequence of MTVDSCj20.

<>

<1>Tawfik, D.S., Griffiths, A.D.
<2>Man-made cell-like compartments for molecular evolution.
<3>Nat. Biotechnol.
<4>16
<5>652-656
<6>1998
<7>Cellular compartmentalization is vital for the evolution of all living organisms.  Cells keep
together the genes, the RNAs and proteins that they encode, and the products of their
activities, thus linking genotype to phenotype.  We have reproduced this linkage in the test
tube by transcribing and translating single genes in the aqueous compartments of water-in-oil
emulsions.  These compartments, with volumes close to those of bacteria, can be recruited to
select genes encoding catalysts.  A protein or RNA with a desired catalytic activity converts
a substrate attached to the gene that encodes it to product.  In other compartments,
substrates attached to genes that do not encode catalysts remain unmodified.  Subsequently,
genes encoding catalysts are selectively enriched by virtue of their linkage to the product.
We demonstrate the linkage of genotype to phenotype in man-made compartments using a model
system.  A selection for target-specific DNA methylation was based on the resistance of the
product (methylated DNA) to restriction digestion.  Genes encoding HaeIII methyltransferase
were selected from a 10^7-fold excess of genes encoding another enzyme.

<>

<1>Tay, A.P., Kaakoush, N.O., Deshpande, N.P., Chen, Z., Mitchell, H., Wilkins, M.R.
<2>Genome Sequence of Campylobacter showae UNSWCD, Isolated from a Patient with Crohn's Disease.
<3>Genome Announcements
<4>1
<5>e00193-12
<6>2013
<7>Campylobacter showae UNSWCD was isolated from a patient with Crohn's disease. Here we present
a 2.1 Mb draft assembly of its genome.

<>

<1>Tay, M., Roizman, D., Cohen, Y., Tolker-Nielsen, T., Givskov, M., Yang, L.
<2>Draft Genome Sequence of the Model Naphthalene-Utilizing Organism Pseudomonas putida OUS82.
<3>Genome Announcements
<4>2
<5>e01161-13
<6>2014
<7>Pseudomonas putida OUS82 was isolated from petrol- and oil-contaminated soil in 1992, and ever
since, it has been used as a model organism to study the microbial
assimilation of naphthalene and phenanthrene. Here, we report the 6.7-Mb draft
genome sequence of P. putida OUS82 and analyze its featured pathways for
biodegradation.

<>

<1>Tay, N.K., Blaschek, H.P.
<2>Purification and characterization of a restriction endonuclease (CpfI) from Clostridium perfringens 10543A.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>224
<6>1994
<7>Nucleases may play an important role in plasmid recovery and uptake in Clostridium
perfringens. A type II restriction endonuclease designated CpfI was isolated from C.
perfringens 10543A. CpfI was purified to homogeneity using a combination of heparin-based
affinity chromatography, DEAE-anion exchange chromatography, and gel permeation
chromatography. The molecular weight of the purified enzyme was determined by gel filtration
and sodium dodecyl sulfate polyacrylamide gel electrophoresis to be 40 and 44 kdal,
respectively. The isoelectric point of the endonuclease is approximately 3.7, and the activity
of the enzyme is inhibited above 45oC. Digestion of methylated and non-methylated pBR322 and
phage DNA substrates suggested CpfI is an isoschizomer of MboI (Sau3A) with a unique
sensitivity to methylation. Enzymes with this particular specificity may prove to be widely
distributed in the genus Clostridium.

<>

<1>Taylor, C., Ford, K., Connolly, B.A., Hornby, D.P.
<2>Determination of the order of substrate addition to MspI DNA methyltransferase using a novel mechanism-based inhibitor.
<3>Biochem. J.
<4>291
<5>493-504
<6>1993
<7>The cloning and overexpression of the MspI DNA methyltransferase as a functional fusion with
glutathione S-transferase is described. The fusion enzyme reatains full biological activity
and has been used to investigate the interaction of substrates and inhibitors with MspI DNA
methyltransferase. The fusion enzyme has been purifed to homogeneity in a single step on
GSH-agarose and is free from contaminating exonuclease activity. The enzyme can be
photolabelled with S-adenosyl-L-methionine and the level of incorporation of label is enhanced
by the presence of a non-specific DNA duplex. In the presence of a cognate
oligo-deoxynucleotide, no photolabelling was observed since methyl transfer occurs instead.
The inclusion of a mechanism-based inhibitor of C-5 deoxycytidine DNA methylation (an
oligodeoxynucleotide containing the base 2-pyrimidinone-1-beta-d-2'-deoxyribofuranoside in
the position of the deoxycytidine to which methyl addition occurs), which is thought to form a
covalent interaction with the reactive cysteine of such enzymes, led to an enhancement of
S-adenosyl-L-methionine photolabelling which suggests that, in contrast with results obtained
with EcoRII DNA methyltransferase [Som and Friedman (1991) J. Biol. Chem. 266, 2937-2945],
methylcysteine is not the photolabelled product. The implications of the results obtained with
this mechanism-based inhibitor are discussed with respect to other C-5-specific DNA
methyltransferases. Gel-retardation assays in the presence of cognate oligodeoxynucleotides
that contain the reactive pyrimidinone base in place of the deoxycytidine target base are
described. These demonstrate that most probably a stable covalent bond is formed between the
methyltransferase and this oligodeoxynucleotide. However, the alternative of extremely tight
non-covalent binding cannot be rigorously excluded. Furthermore, the results from these
experiments indicate that the reaction mechanism proceeds in a manner similar to that of HhaI
DNA methyltransferase with sequence-specific DNA binding being followed by addition of
S-adenosyl-L-methionine and concomitant isomerization of the ternary complex leading to methyl
transfer. S-Adenosyl-L-homocysteine appears to inhibit the reaction pathway was as a result of
either competition with the methyl donor and potentiation of a high affinity interaction
between the enzyme and DNA in an abortive ternary complex or through an allosteric
interaction.

<>

<1>Taylor, D.E., Eaton, M., Chang, N., Salama, S.M.
<2>Construction of a Helicobacter pylori genome map and demonstration of diversity at the genome level.
<3>J. Bacteriol.
<4>174
<5>6800-6806
<6>1992
<7>Genomic DNA from 30 strains of Helicobacter pylori was subjected to pulsed-field gel
electrophoresis after digestion with NotI and NruI.  The genome sizes of the strains ranged
from 1.6 to 1.73 Mb, with an average size of 1.67 Mb. By using NotI and NruI, a circular map
of H. pylori UA802 (1.7 Mb) which contained three copies of 16S and 23S rRNA genes was
constructed.  An unusual feature of the H. pylori genome was the separate location of at least
two copies of 16S and 23S rRNA genes.  Almost all strains had diferent PFGE patterns after
NotI and NruI digestion, suggesting that the H. pylori genome possesses a considerable degree
of genetic variability.  However, three strains from different sites (the fundus, antrum, and
body of the stomach) within the same patient gave identical PFGE patterns.  The genomic
pattern of individual isolates remained constant during multiple subcultures in vitro.  The
reason for the genetic diversity observed among H. pylori strains remains to be explained.

<>

<1>Taylor, G.K., Heiter, D.F., Pietrokovski, S., Stoddard, B.L.
<2>Activity, specificity and structure of I-Bth0305I: a representative of a new homing endonuclease family.
<3>Nucleic Acids Res.
<4>39
<5>9705-9719
<6>2011
<7>Novel family of putative homing endonuclease genes was recently discovered during analyses of
metagenomic and genomic sequence data. One such protein is encoded within a group I intron
that resides in the recA gene of the Bacillus thuringiensis 03058-36 bacteriophage. Named
I-Bth0305I, the endonuclease cleaves a DNA target in the uninterrupted recA gene at a position
immediately adjacent to the intron insertion site. The enzyme displays a multidomain,
homodimeric architecture and footprints a DNA region of  approximately 60 bp. Its highest
specificity corresponds to a 14-bp pseudopalindromic sequence that is directly centered across
the DNA cleavage site. Unlike many homing endonucleases, the specificity profile of the enzyme
is evenly distributed across much of its target site, such that few single base pair
substitutions cause a significant decrease in cleavage activity. A crystal structure of its
C-terminal domain confirms a nuclease fold that is homologous to very short patch repair (Vsr)
endonucleases. The domain architecture and DNA recognition profile displayed by I-Bth0305I,
which is the prototype of a homing lineage that we term the 'EDxHD' family, are distinct
from previously characterized homing endonucleases.

<>

<1>Taylor, G.K., Petrucci, L.H., Lambert, A.R., Baxter, S.K., Jarjour, J., Stoddard, B.L.
<2>LAHEDES: the LAGLIDADG homing endonuclease database and engineering server.
<3>Nucleic Acids Res.
<4>40
<5>W110-W116
<6>2012
<7>LAGLIDADG homing endonucleases (LHEs) are DNA cleaving enzymes, also termed 'meganucleases'
that are employed as gene-targeting reagents. This use of LHEs requires that their DNA
specificity be altered to match sequences in genomic targets. The choice of the most
appropriate LHE to target a particular gene is facilitated by the growing number of such
enzymes with well-characterized activities and structures. 'LAHEDES' (The LAGLIDADG Homing
Endonuclease Database and Engineering Server) provides both an online archive of LHEs with
validated DNA cleavage specificities and DNA-binding interactions, as well as a tool for the
identification of DNA sequences that might be targeted by various LHEs. Searches can be
performed using four separate scoring algorithms and user-defined choices of LHE scaffolds.
The webserver subsequently provides information regarding clusters of amino acids that should
be interrogated during engineering and selection experiments. The webserver is fully open
access and can be found at http://homingendonuclease.net.

<>

<1>Taylor, G.K., Stoddard, B.L.
<2>Structural, functional and evolutionary relationships between homing endonucleases and proteins from their host organisms.
<3>Nucleic Acids Res.
<4>40
<5>5189-5200
<6>2012
<7>Homing endonucleases (HEs) are highly specific DNA-cleaving enzymes that are encoded by
invasive DNA elements (usually mobile introns or inteins) within the genomes of phage,
bacteria, archea, protista and eukaryotic organelles. Six unique structural HE families, that
collectively span four distinct nuclease catalytic motifs, have been characterized to date.
Members of each family display structural homology and functional relationships to a wide
variety of proteins from various organisms. The biological functions of those proteins are
highly disparate and include non-specific DNA-degradation enzymes, restriction endonucleases,
DNA-repair enzymes, resolvases, intron splicing factors and transcription factors. These
relationships suggest that modern day HEs share common ancestors with proteins involved in
genome fidelity, maintenance and gene expression. This review summarizes the results of
structural studies of HEs and corresponding proteins from host organisms that have illustrated
the manner in which these factors are related.

<>

<1>Taylor, I., Patel, J., Firman, K., Kneale, G.
<2>Purification and biochemical characterisation of the EcoR124 type I modification methylase.
<3>Nucleic Acids Res.
<4>20
<5>179-186
<6>1992
<7>Large scale purification of the type I modification methylase EcoR124 has been
achieved from an overexpressing strain by a two step procedure using
ion-exchange and heparin chromatography.  Pure methylase is obtained at a yield
of 30mg per gm of cell paste.  Measurements of the molecular weight and subunit
stoichiometry show that the enzyme is a trimeric complex of 162 kDa consisting
of two subunits of HsdM (58 kDa) and one subunit of HsdS (46 kDa).  The
purified enzyme can methylate a DNA fragment bearing its cognate recognition
sequence.  Binding of the methylase to synthetic DNA fragments containing
either the EcoR124 recognition sequence GAAN6RT-CG, or the recognition sequence
GAAN7RTCG of the related enzyme EcoR124/3, was followed by fluorescence
competition assays and by gel retardation analysis.  The results show that the
methylase binds to its correct sequence with an affinity of the order 10/8 M-1
forming a 1:1 complex with the DNA.  The affinity for the incorrect sequence,
differing by an additional base pair in the non-specific spacer, is almost two
orders of magnitude lower.

<>

<1>Taylor, I., Watts, D., Kneale, G.
<2>Substrate recognition and selectivity in the type IC DNA modification methylase M.EcoR124I.
<3>Nucleic Acids Res.
<4>21
<5>4929-4935
<6>1993
<7>The type I DNA modification methylase M.EcoR124I binds sequence specifically to DNA and
protects a 25bp fragment containing its cognate recognition sequence from digestion by
exonuclease III. Using modified synthetic oligonucleotide duplexes we have investigated the
catalytic properties of the methylase, and have established that a specific adenine on each
strand of DNA is the site of methylation. We show that the rate of methylation of each adenine
is increased at least 100 fold by prior methylation at the other site. However this is
accompanied by a significant decrease in the affinity of the methylase for these substrates
according to competitive gel retardation assays. In contrast, methylation of an adenine in the
recognitiom site which is not a target for the enzyme results in only a small decrease in both
DNA binding affinity and rate of methylation by the enzyme.

<>

<1>Taylor, I.A.
<2>Biochemical characterisation of the EcoR124 type I DNA modification methylase.
<3>Diss. Abstr.
<4>53
<5>3942
<6>1993
<7>The hsdS and hsdM genes constituting the EcoR124I type I DNA modification methylase have been
expressed independently in Escherichia coli (E. coli) using the high level expression vectors
pUM120 and pJS491. Analysis of cell extracts demonstrated that the HsdM potein is found in the
E. coli soluble fraction whilst the HsdS protein is found only in the E. coli insoluble pellet
fraction. Protocols have been developed for the purification of both the HsdM and HsdS
proteins and a preliminary biochemical analysis undertaken involving an investigation of the
denaturation of both proteins by fluorescence spectroscopy. A more extensive analysis of the
HsdM protein has allowed estimations of its solution molecular mass and secondary structure to
be made. The hsdS and hsdM genes have also been co-expressed using the high level expression
vector pJS4M. The analysis of cell extracts in this case revealed that both polypeptides are
present in the E. coli soluble fraction. A protocol has been developed in which both
polypeptides co-purify as a complex which is capable of methylating the EcoR124I recognition
sequence. The enzyme complex has been shown to exist predominantly in solution as a trimer
consisting of two copies of the HsdM polypeptide and a single copy of the HsdS polypeptide.
Further experiments with the purified enzyme show that it can bind its recognition sequence
with a dissociation constant of around 5x10-9 and that it can also bind the recognition
sequence of the related type I DNA methylase EcoR124/3I with a lower affinity. A protein
nucleic acid complex consisting of methylase and a 30 base pair synthetic oligonucleotide
duplex has been investigated using 1H NMR spectroscopy and footprinting experiments. The
results suggest that contacts are made between bases within the recognition sequence and the
protein and also that the normal geometry of the nucleic acid may be distorted.

<>

<1>Taylor, I.A., Davis, K.G., Watts, D., Kneale, G.G.
<2>DNA binding induces a major structural transition in a type I methyltransferase.
<3>EMBO J.
<4>13
<5>5772-5778
<6>1994
<7>The type IC DNA methyltransferase M.EcoR124I is a complex multisubunit enzyme that recognizes
the nonpalindromic DNA sequence GAAN6RTCG. Small angle X-ray scattering has been used to
investigate the solution structure of the methyltransferase and of complexes of the enzyme
with unmethylated and hemimethylated 30 bp DNA duplexes containing the specific recognition
sequence. A major change in the quaternary structure of the enzyme is observed following DNA
binding, based on a decrease in the radius of gyration from 56 to 40 A and a reduction in the
maximum dimension of the enzyme from 180 to 112 A. The structural transition observed is
independent of the methylation state of the DNA. CD shows that there is no change in the
secondary structure of the protein subunits when DNA is bound. In contrast, there is a large
increase in the CD signal arising from the DNA, suggesting considerable structural distortion
which may allow access to the bases targeted for methylation. We propose that DNA binding
induces a large rotation of the two HsdM subunits towards the DNA, mediated by hinge bending
domains in the specificity subunit HsdS.

<>

<1>Taylor, I.A., Kneale, G.G.
<2>A competition assay for DNA binding using the fluorescent probe ANS.
<3>Methods Mol. Biol.
<4>543
<5>577-587
<6>2009
<7>Fluorescence spectroscopy is a technique frequently employed to stud), protein-nucleic acid
interactions. Often, the intrinsic fluorescence
emission spectrum of tryptophan residues in a nucleic-acid-binding
protein is strongly perturbed upon interaction with a target DNA or
RNA. These spectral changes can then be exploited in order to construct
binding isotherms and the extract equilibrium association constant
together with the stoichiometry of an interaction. However, when a
protein contains many tryptophan residues that are not located in the
proximity of the nucleic-acid-binding site, changes in the fluorescence
emission spectrum may not be apparent or the magnitude too small to be
useful. Here, we make use of an extrinsic fluorescence probe, the
environmentally sensitive fluorophore 1-anilinonaphthalene-8-sulphonic
acid (1,8-ANS). Displacement by DNA of 1,8-ANS Molecules from the
nucleic-acid-binding site of the Type I modification methylase EcoR124I
results in red shifting and an intensity decrease of the 1,8-ANS
fluorescence emission spectrum. These spectral changes have been used
to investigate the interaction of EcoR124I with DNA target recognition
sequences.

<>

<1>Taylor, I.A., Webb, M., Kneale, G.G.
<2>Surface labelling of the type I methyltransferase M.EcoR124I reveals lysine residues critical for DNA binding.
<3>J. Mol. Biol.
<4>258
<5>62-73
<6>1996
<7>The type IC methyltransferase M.EcoR124I consists of a specificity subunit
(HsdS) and two methylation subunits (HsdM).  Using chemical modification, we have investigated
the accessibility of lysine residues in the free enzyme and in the complex with its DNA
recognition
sequence.  A total of 41 of the 109 lysine residues in the enzyme are susceptible to
modification, of
which 19 are located in the HsdS subunit and 11 in each of the two HsdM subunits.  DNA binding
results in extensive protection of lysine residues in the HsdS subunit, while those in the
HsdM
subunit are only protected weakly.  The DNA binding activity of the methylase is abolished
when a
small fraction of the accessible lysine residues are modified.  Peptide mapping and N-terminal
sequencing has been used to locate the rapidly modified lysine residues in HsdS that are
critical for
DNA binding.  Highly modified residues (K297, K261 and K327) are found in the C-terminal
variable domain that is responsible for DNA recognition, but others (K196, K203 and K210) are
found in the conserved regions that had not previously been implicated in DNA binding.

<>

<1>Taylor, J., Moore, H., Beaujean, N., Gardner, J., Wilmut, I., Meehan, R., Young, L.
<2>Cloning and Expression of Sheep DNA Methyltransferase 1 and Its Development-Specific Isoform.
<3>Mol. Reprod. Dev.
<4>76
<5>501-513
<6>2009
<7>Unlike the mouse embryo, where loss of DNA methylation in the embryonic nucleus leaves
cleavage stage embryos globally hypomethylated, sheep
preimplantation embryos retain high levels of methylation until the
blastocyst stage. We have cloned and sequenced sheep Dnmt1 and found it
to be highly conserved with both the human and mouse homologues.
Furthermore, we observed that the transcript normally expressed in
adult somatic tissues is highly abundant in sheep oocytes. Throughout
sheep preimplantation development the protein is retained in the
cytoplasm whereas Dnmt1 transcript production declines after the
embryonic genome activation at the 8-16 cell stage. Attempts to clone
oocyte-specific 5' regions of Dnmt1, known to be present in the mouse
and human gene, were unsuccessful. However, a novel ovine Dnmt1 exon,
theoretically encoding 13 amino acids, was found to be expressed in
sheep oocytes, preimplantation embryos and early fetal lineages, but
not in the adult tissue. RNAi-mediated knockdown of this novel
transcript resulted in embryonic developmental arrest at the late
morula stage, suggesting an essential role for this isoform in sheep
blastocyst formation.

<>

<1>Taylor, J.D., Ackroyd, A.J., Halford, S.E.
<2>The gel shift assay for the analysis of DNA-protein interactions.
<3>The Nucleic Acid Protocols Handbook, Humana Press, Inc., Rapley, R., Totowa, NJ
<4>0
<5>745-756
<6>2000
<7>The gel shift assay is one of the most powerful methods for the analysis of DNA-protein
interactions.  The assay itself is simple.  DNA and protein are mixed together, the solution
subjected to electrophoresis through polyacrylamide, and the gel is then analyzed for DNA,
usually by autoradiography of radiolabeled DNA.  Binding of the protein to the DNA can result
in a complex that has a different electrophoretic mobility from the free DNA.  In general, the
mobility of the complex is retarded relative to the unbound DNA and thus the assay is often
called gel retardation.  However, with circular DNA substrates (typically, minicircles of
200-400 bp), the DNA-protein complex can migrate faster than the free DNA.  The separation of
the complex from the free DNA, and therefore the detection of the complex, is dependent on a
variety of factors.  These must be determined experimentally for each system.  However, the
ease with which the assay can be performed means that the optimal conditions can be discovered
quickly.  Factors that influence the electrophoretic mobility of DNA-protein complexes include
the molecular weight of the protein and the DNA, the ionic strength and the pH of the
electrophoresis buffer, the concentration of the gel matrix, and the temperature. Particularly
useful accounts of how modifications to the assay can affect the mobility of DNA-protein
complexes have been published.

<>

<1>Taylor, J.D., Ackroyd, A.J., Halford, S.E.
<2>The gel shift assay for the analysis of DNA-protein interactions.
<3>Methods Mol. Biol.
<4>30
<5>263-279
<6>1994
<7>The gel shift assay is one of the most powerful methods for the analysis of DNA-protein
interactions.  The assay itself is simple.  DNA and protein are mixed together, the solution
subjected to electrophoresis through polyacrylamide, and the gel is then analyzed for DNA,
usually by autoradiography of radiolabeled DNA.  Binding of the protein to the DNA can result
in a complex that has a different electrophoretic mobility from the free DNA.  In general, the
mobility of the complex is retarded relative to the unbound DNA and thus the assay is often
called gel retardation.  However, with circular DNA substrates (typically, minicircles of
200-400 bp), the DNA-protein complex can migrate faster than the free DNA.  The separation of
the complex from the free DNA, and therefore the detection of the complex, is dependent on a
variety of factors.  These must be determined experimentally for each system.  However, the
ease with which the assay can be performed means that the optimal conditions can be discovered
quickly.  Factors that influence the electrophoretic mobility of DNA-protein complexes include
the molecular weight of the protein and the DNA, the ionic strength and the pH of the
electrophoresis buffer, the concentration of the gel matrix, and the temperature.
Particularly useful accounts of how modifications to the assay can affect the mobility of
DNA-protein complexes have been published.

<>

<1>Taylor, J.D., Badcoe, I.G., Clarke, A.R., Halford, S.E.
<2>EcoRV restriction endonuclease binds all DNA sequences with equal affinity.
<3>Biochemistry
<4>30
<5>8743-8753
<6>1991
<7>In the presence of MgCl2, the EcoRV restriction endonuclease cleaves its
recognition sequence on DNA at least a million times more readily than any
other sequence.  In this study, the binding of the EcoRV restriction enzyme to
DNA was examined in the absence of Mg2+.  With each DNA fragment tested,
several DNA-protein complexes were detected by electrophoresis through
polyacrylamide.  No differences were observed between isogenic DNA molecules
that either contained or lacked the EcoRV recognition site.  The number of
complexes with each fragment varied with the length of the DNA.  Three
complexes were formed with a DNA molecule of 55 base pairs, corresponding to
the DNA bound to 1,2,3 molecules of the protein, while >15 complexes were
formed with a DNA of 381 base pairs.  A new method was developed to analyze the
binding of a protein to multiple sites on DNA.  The method showed that the
EcoRV enzyme binds to all DNA sequences, including the EcoRV recognition site,
with the same equilibrium constant, though two molecules of the protein bind
preferentially to adjacent sites on the DNA in a cooperative fashion.  All of
the complexes with a substrate that contained the EcoRV site dissociated upon
addition of competitor DNA, but when the competitor was mixed with MgCl2, a
fraction of the substrate was cleaved at the EcoRV site.  The fraction cleaved
was due mainly to the translocation of the enzyme from nonspecific sites on the
DNA to the specific site.

<>

<1>Taylor, J.D., Goodall, A.J., Vermote, C.L., Halford, S.E.
<2>Fidelity of DNA recognition by the EcoRV restriction/modification system in vivo.
<3>Biochemistry
<4>29
<5>10727-10733
<6>1990
<7>The EcoRV restriction/modification system consists of two enzymes that recognize the DNA
sequence GATATC. The EcoRV restriction endonuclease cleaves DNA at this site, but the DNA of
Escherichia coli carrying the EcoRV system is protected from this reaction by the EcoRV
methyltransferase. However, in vitro, the EcoRV nuclease also cleaves DNA at most sites that
differ from the recognition sequence by one base pair. Though the reaction of the nuclease at
these sites is much slower than that at the cognate site, it still appears to be fast enough
to cleave the chromosome of the cell into many fragments. The possibility that the EcoRV
methyltransferase also protects the noncognate sites on the chromosome was examined. The
modification enzyme methylated alternate sites in vivo, but these were not the same as the
alternate sites for the nuclease. The excess methylation was found at GATC sequences, which
are also the targets for the dam methyltransferase of E. coli, a protein that is homologous to
the EcoRV methyltransferase. Methylation at these sites gave virtually no protection against
the EcoRV nuclease: even when the EcoRV methyltransferase had been overproduced, the cellular
DNA remained sensitive to the EcoRV nuclease at its noncognate sites. The viability of E. coli
carrying the EcoRV restriction/modification R/M system was found instead to depend on the
activity of DNA ligase. Ligase appears to proofread the EcoRV R/M system in vivo: DNA, cut
initially in one strand at a noncognate site for the nuclease, is presumably repaired by
ligase before the scission of the second strand.

<>

<1>Taylor, J.D., Halford, S.E.
<2>The activity of the EcoRV restriction endonuclease is influenced by flanking DNA sequences both inside and outside the DNA-protein complex.
<3>Biochemistry
<4>31
<5>90-97
<6>1992
<7>The EcoRV restriction endonuclease cleaves DNA not only at its recognition
sequence but also at most other sequences that differ from the recognition site
by one base pair.  Compared to the reaction at the recognition site, the
reactions at noncognate sites are slow but 1 out of the 12 noncognate sites on
the plasmid pAT153 is cleaved more than 50 times faster than any other.  The
increase in the reaction rate at the preferred noncognate site, relative to
other sites, was caused by the DNA sequences in the 4 base pairs from either
side of the site.  For enhanced activity by EcoRV, particular bases were needed
immediately adjacent to the site, inside the DNA-protein complex.  At these
loci, the protein interacts with the phosphate groups in the DNA and the
flanking sequence may control the activity of the enzyme by determining the
conformation of the DNA, thus aligning the phosphate contacts.  But the
preferential cleavage also depended on sequences further away from the site, at
loci outside the complex.  At external positions, beyond the reach of the
protein, the EcoRV enzyme required flanking sequences that give rise to
flexibility in DNA conformation.  These may facilitate the distortion of the
DNA required for catalysis by EcoRV.

<>

<1>Taylor, J.D., Halford, S.E.
<2>Discrimination between DNA sequences by the EcoRV restriction endonuclease.
<3>Biochemistry
<4>28
<5>6198-6207
<6>1989
<7>The EcoRV restriction endonuclease cleaves not only its recognition sequence on
DNA, GATATC, but also, at vastly reduced rates, a number of alternative DNA
sequences.  The plasmid pAT153 contains 12 alternative sites, each of which
differs from the recognition sequence by one base pair.  The EcoRV nuclease
showed a marked preference for one particular site from among these
alternatives.  This noncognate site was located at the sequence GTTATC, and the
mechanism of action of EcoRV at this site was analyzed.  The mechanism differed
from that at the cognate site in three respects.  First, the affinity of the
enzyme for the noncognate site was lower than that for the cognate site, but,
by itself, this cannot account for the specificity of EcoRV as measured from
the values of Kcat/Km.  Second, the enzyme had a lower affinity for Mg2+ when
it was bound to the noncognate site than when it was bound to its cognate site:
this appears to be a key factor in limiting the rates of DNA cleavage at
alternative sites.  Third, the reaction pathway at the noncognate site differed
from that at the cognate site.  At the former, the EcoRV enzyme cleaved first
one strand of the DNA and then the other while at the latter, both strands were
cut in one concerted reaction.  The difference in reaction pathway allows DNA
ligase to proofread the activity of EcoRV by selective repair of single-strand
breaks at noncognate sites, as opposed to double-strand breaks at the cognate
site.  The addition of DNA ligase to reactions with EcoRV made no difference to
product formation at the cognate site, but products from reactions at
noncognate sites were no longer detected.

<>

<1>Taylor, J.E., Callow, P., Swiderska, A., Kneale, G.G.
<2>Structural and functional analysis of the engineered type I DNA methyltransferase EcoR124I(NT).
<3>J. Mol. Biol.
<4>398
<5>391-399
<6>2010
<7>The Type I R-M system EcoR124I is encoded by three genes. HsdM is responsible for modification
(DNA methylation), HsdS for DNA sequence
specificity and HsdR for restriction endonuclease activity. The trimeric
methyltransferase (M(2)S) recognises the asymmetric sequence
(GAAN(6)RTCG). An engineered R-M system, denoted EcoR124I(NT), has two
copies of the N-terminal domain of the HsdS subunit of EcoR124I, instead
of a single S subunit with two domains, and recognises the symmetrical
sequence GAAN(7)TTC. We investigate the methyltransferase activity of
EcoR124I(NT), characterise the enzyme and its subunits by analytical
ultracentrifugation and obtain low-resolution structural models from
small-angle neutron scattering experiments using contrast variation and
selective deuteration of subunits.

<>

<1>Taylor, J.E., Swiderska, A., Artero, J.-B., Callow, P., Kneale, G.
<2>Structural and Functional Analysis of the Symmetrical Type I Restriction Endonuclease R.EcoR124I(NT).
<3>PLoS ONE
<4>7
<5>e35263
<6>2012
<7>Type I restriction-modification (RM) systems are comprised of two multi-subunit enzymes, the
methyltransferase (similar to 160 kDa),
responsible for methylation of DNA, and the restriction endonuclease
(similar to 400 kDa), responsible for DNA cleavage. Both enzymes share
a number of subunits. An engineered RM system, EcoR124I(NT), based on
the N-terminal domain of the specificity subunit of EcoR124I was
constructed that recognises the symmetrical sequence GAAN(7)TTC and is
active as a methyltransferase. Here, we investigate the restriction
endonuclease activity of R.EcoR124I(NT) in vitro and the subunit
assembly of the multi-subunit enzyme. Finally, using small-angle
neutron scattering and selective deuteration, we present a
low-resolution structural model of the endonuclease and locate the
motor subunits within the multi-subunit enzyme. We show that the
covalent linkage between the two target recognition domains of the
specificity subunit is not required for subunit assembly or enzyme
activity, and discuss the implications for the evolution of Type I
enzymes.

<>

<1>Taylor, J.E., Swiderska, A., Kneale, G.G.
<2>A rapid purification procedure for the HsdM protein of EcoR124I and biophysical characterization of the purified protein.
<3>Protein Expr. Purif.
<4>87
<5>136-140
<6>2013
<7>Type I restriction-modification (R-M) systems are comprised of two multi-subunit enzymes with
complementary functions: the
methyltransferase (similar to 160 kDa), responsible for methylation of
DNA, and the restriction endonuclease (similar to 400 kDa), responsible
for DNA cleavage. Both enzymes share a number of subunits, including
HsdM. Characterisation of either enzyme first requires the expression
and purification of its constituent subunits, before reconstitution of
the multisubunit complex. Previously, purification of the HsdM protein
had proved problematic, due to the length of time required for the
purification and its susceptibility to degradation. A new protocol was
therefore developed to decrease the length of time required to purify
the HsdM protein and thus prevent degradation. Finally, we show that
the HsdM subunit exhibits a concentration dependent monomer-dimer
equilibrium.

<>

<1>Taylor, J.R., Fang, M.M., Nie, S.
<2>Probing specific sequences on single DNA molecules with bioconjugated fluorescent nanoparticles.
<3>Anal. Chem.
<4>72
<5>1979-1986
<6>2000
<7>Nanometer-sized fluorescent particles (latex nanobeads) have been covalently linked to DNA
binding proteins to probe specific sequences on stretched single DNA molecules.  In comparison
with single organic fluorophores, these nanoparticle probes are brighter, are more stable
against photobleaching, and do not suffer from intermittent on/off light emission (blinking).
Specifically, we demonstrate that the site-specific restriction enzyme EcoRI can be conjugated
to 20 nm fluorescent nanoparticles and that the resulting nanoconjugates display DNA binding
and cleavage activities of the native enzyme.  In the absence of cofactor magnesium ions, the
EcoRI conjugates bind to specific sequences on double-stranded DNA but do not initiate
enzymatic cutting.  For single DNA molecules that are stretched and immobilized on a solid
surface, nanoparticles bound at specific sites can be directly visualized by multicolor
fluorescence microscopy.  Direct observation os site-specific probes on single DNA molecules
opens new possibilities in optical gene mapping and in fundamental study of DNA-protein
interactions.

<>

<1>Taylor, J.W., Schmidt, W., Cosstick, R., Okruszek, A., Eckstein, F.
<2>The use of phosphorothioate-modified DNA in restriction enzyme reactions to prepare nicked DNA.
<3>Nucleic Acids Res.
<4>13
<5>8749-8763
<6>1985
<7>The RF IV form of M13 DNA was synthesized enzymatically in vitro, using the
viral (+)strand as template, to contain phosphorothioate-modified
internucleotidic linkages of the Rp configuration on the 5' side of every base
of a particular type in the newly-synthesized (-)strand.  Twenty nine
restriction enzymes were then tested for their reactions with the appropriate
modified DNA types having a phosphorothioate linkage placed exactly at the
cleavage site(s) of these enzymes in the (-)strand.  Eleven of the seventeen
restriction enzymes tested that had recognition sequences of five bases or more
could be used to convert the phosphorothioate DNA entirely into the nicked
form, either by simply allowing the reaction to go to completion with excess
enzyme (AvaI, AvaII, BanII, HindII, NciI, PstI or PvuI) or by stopping the
reaction at the appropriate time before the nicked DNA is linearized (BamHI,
BglI, EcoRI or HindIII).  Only modification of the exact cleavage site in the
(-)strand could block linearization by the first class of enzymes.  The results
presented imply that the restriction enzyme-directed nicking of
phosphorothioate M13 DNA occurs exclusively in the (+)strand.

<>

<1>Taylor, V.L., Titball, R.W., Oyston, P.C.F.
<2>Oral immunization with a dam mutant of Yersinia pseudotuberculosis protects against plague.
<3>Microbiology
<4>151
<5>1919-1926
<6>2005
<7>Inactivation of the gene encoding DNA adenine methylase (dam) has been shown to attenuate some
pathogens such as Salmonella enterica serovar
Typhimurium and is a lethal mutation in others such as Yersinia
pseudotuberculosis strain YPIII. In this study the dam methylase gene
in Yersinia pseudotuberculosis strain IP32953 was inactivated. Unlike
the wild-type, DNA isolated from the mutant could be digested with
Mbol, which is consistent with an altered pattern of DNA methylation.
The mutant was sensitive to bile salts but not to 2-aminopurine. The
effect of dam inactivation on gene expression was examined using a DNA
microarray. In BALB/c mice inoculated orally or intravenously with the
dam mutant, the median lethal dose (MLD) was at least 10(6)-fold higher
than the MLD of the wild-type. BALB/c mice inoculated with the mutant
were protected against a subcutaneous challenge with 100 MLDs of
Yersinia pestis strain GB and an intravenous challenge with 300 MLDs of
Y. pseudotuberculosis IP32953.

<>

<1>te Poele, E.M., Samborskyy, M., Oliynyk, M., Leadlay, P.F., Bolhuis, H., Dijkhuizen, L.
<2>Actinomycete integrative and conjugative pMEA-like elements of Amycolatopsis and Saccharopolyspora decoded.
<3>Plasmid
<4>59
<5>202-216
<6>2008
<7>Actinomycete integrative and conjugative elements (AICEs) are present in
diverse genera of the actinomycetes, the most important bacterial
producers of bioactive secondary metabolites. Comparison of pMEA100 of
Amycolatopsis mediterranei, pMEA300 of Amycolatopsis methanolica and
pSE211 of Saccharopolyspora erythraea, and other AICEs, revealed a highly
conserved structural organisation, consisting of four functional modules
(replication, excision/integration, regulation, and conjugative transfer).
Features conserved in all elements, or specific for a single element, are
discussed and analysed. This study also revealed two novel putative AICEs
(named pSE222 and pSE102) in the Sac. erythraea genome, related to the
previously described pSE211 and pSE101 elements. Interestingly, pSE102
encodes a putative aminoglycoside phosphotransferase which may confer
antibiotic resistance to the host. Furthermore, two of the six pSAM2-like
insertions in the Streptomyces coelicolor genome described by Bentley et
al. [Bentley, S.D., Chater, K.F., Cerdeno-Tarraga, A.M., et al., 2002.
Complete genome sequence of the model actinomycete Streptomyces coelicolor
A3(2). Nature 417, 141-147] could be functional AICEs. Homologues of
various AICE proteins were found in other actinomycetes, in Frankia
species and in the obligate marine genus Salinispora and may be part of
novel AICEs as well. The data presented provide a better understanding of
the origin and evolution of these elements, and their functional
properties. Several AICEs are able to mobilise chromosomal markers,
suggesting that they play an important role in horizontal gene transfer
and spread of antibiotic resistance, but also in evolution of genome
plasticity.

<>

<1>Teachman, A.M., Gumulak-Smith, J.J., Tu, A.T., Simecka, J.W., Lindsey, J.R., Dybvig, K.
<2>Variations in the surface proteins and in the restriction and modification systems of Mycoplasma pulmonis in the respiratory tract of  infected rats.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>101
<5>386
<6>2001
<7>Restriction and modification (R-M) systems are thought to afford bacteria protection from
invading foreign DNA. We hypothesize that the
phase-variable R-M systems found within the hsd loci, as well as the
variable surface antigens, of Mycoplasma pulmonis may additionally have
a critical role in survival within the rat respiratory tract.
Populations of M. pulmonis strain X1048 were isolated from
experimentally infected Fischer F344 rats and assayed by a combination
of Western blot, PCR and phage plaquing experiments to determine the
predominant variable surface antigen (Vsa) and Hsd proteins. Isolates
from the nose were found to produce VsaA and possessed no R-M activity,
while 38% of isolates from the throat at 14 days postinfection produced
a protein other than VsaA with 31% having active R-M systems. To assess
the stability of various isolates from the rat throat, selected
isolates were passaged 30 times in vitro and were found to revert to a
phenotype of VsaA with inactive R-M systems. Therefore, the data
suggest that different selection pressures within tissues influence the
cell population in regards to surface antigen variation and R-M
production, and subsequently may affect survival of the mycoplasma
within an animal.

<>

<1>Tediashivili, M.I., Goryan, T.V., Koberidse, T.D., Chanishvili, T.G., Nikolskaya, I.I.
<2>New systems of host specificity of DNA Pae610 and Pae603.
<3>Biull. Eksp. Biol. Med.
<4>112
<5>91-94
<6>1991
<7>In order to find new systems of host specificity, wide screening among different strains of P.
aeruginosa was carried out.  By means of cross-titration of phages BT and FP-series two new
systems of modification-restriction were identified: Pae610 and Pae603.  They differ from the
known system of host specificity PaeR7.

<>

<1>Tediashvili, M.I., Goryan, T.V., Koberidze, T.D., Chanishvili, T.G., Nikolskaya, I.I.
<2>New host DNA specificity systems Pae 610 and Pae 603.
<3>Bull. Exp. Biol. Med.
<4>112
<5>91-94
<6>1991
<7>To find new systems of host specificity (SHS), wide screening among different
strains of Pseudomonas aeruginosa was carried out.  By means of cross-titration
of phages BT and FP-series two new systems of restriction-modification were
identified:  Pae610 and Pae603.  They differ from the known system of host
specificity PaeR7.

<>

<1>Tediashvili, M.I., Uporova, T.M., Nikolskaya, I.I., Debov, S.S.
<2>Identification of host specific system in Shigella.
<3>Biull. Eksp. Biol. Med.
<4>90
<5>324-325
<6>1980
<7>The presence of a DNA host specific system in Shigella sonnei 47843 bacteria has been
demonstrated.  Phage DDIII grown on the cells of Shigella stutzeri 2, in Shigella sonnei 47843
cells is restricted by a factor of 10^5.  Phage T3 of EcoB phenotype as well as DDIII phage is
restricted in these cells.  This circumstance means that the restriction-modification system
of Shigella sonnei 47843 differs in specificity from the well known system E. coli.  The
results obtained are the second case of host specific system identification in Shigella.  The
biological properties of the strain (form of the colonies, colicinogenic activity, antibiotic
resistance, ability to ferment sugars, etc.) have been studied.

<>

<1>Teel, L.D., Melton-Celsa, A.R., Schmitt, C.K., O'Brien, A.D.
<2>One of Two Copies of the Gene for the Activatable Shiga Toxin Type 2d in Escherichia coli O91:H21 Strain B2F1 Is Associated with an Inducible Bacteriophage.
<3>Infect. Immun.
<4>70
<5>4282-4291
<6>2002
<7>Shiga toxin (Stx) types 1 and 2 are encoded within intact or defective temperate
bacteriophages in Stx-producing Escherichia coli (STEC), and expression of these toxins is
linked to bacteriophage induction. Among Stx2 variants, only stx(2e) from one human STEC
isolate has been reported to be carried within a toxin-converting phage. In this study, we
examined the O91:H21 STEC isolate B2F1, which carries two functional alleles for the potent
activatable Stx2 variant toxin, Stx2d, for the presence of Stx2d-converting bacteriophages. We
first constructed mutants of B2F1 that produced one or the other Stx2d toxin and found that
the mutant that produced only Stx2d1 made less toxin than the Stx2d2-producing mutant.
Consistent with that result, the Stx2d1-producing mutant was attenuated in a
streptomycin-treated mouse model of STEC infection. When the mutants were treated with
mitomycin C to promote bacteriophage induction, Vero cell cytotoxicity was elevated only in
extracts of the Stx2d1-producing mutant. Additionally, when mice were treated with
ciprofloxacin, an antibiotic that induces the O157:H7 Stx2-converting phage, the animals were
more susceptible to the Stx2d1-producing mutant. Moreover, an stx(2d1)-containing lysogen was
isolated from plaques on strain DH5alpha that had been exposed to lysates of the mutant that
produced Stx2d1 only, and supernatants from that lysogen transformed with a plasmid encoding
RecA were cytotoxic when the lysogen was induced with mitomycin C. Finally,
electron-microscopic examination of extracts from the Stx2d1-producing mutant showed hexagonal
particles that resemble the prototypic Stx2-converting phage 933W. Together these observations
provide strong evidence that expression of Stx2d1 is bacteriophage associated. We conclude
that despite the sequence similarity of the stx(2d1)- and stx(2d2)-flanking regions in B2F1,
Stx2d1 expression is repressed within the context of its toxin-converting phage while Stx2d2
expression is independent of phage induction.

<>

<1>Teerawanichpan, P., Chandrasekharan, M.B., Jiang, Y.M., Narangajavana, J., Hall, T.C.
<2>Characterization of two rice DNA methyltransferase genes and RNAi-mediated reactivation of a silenced transgene in rice callus.
<3>Planta
<4>218
<5>337-349
<6>2004
<7>Two genomic clones (OsMET1-1, AF462029 and OsMET1-2, TPA BK001405), each encoding a
cytosine-5 DNA methyltransferase (MTase), were isolated
from rice (Oryza sativa L.) BAC libraries. OsMET1-1 has an open reading
frame of 4,566 nucleotides with 12 exons and 11 introns while OsMET1-2
has an open reading frame of 4,491 nucleotides with 11 exons and 10
introns. Although OsMET1-1 and OsMET1-2 have high sequence similarity
overall, they share only 24% identity in exon 1, and intron 3 of
OsMET1-1 is absent from OsMET1-2. As for other eukaryotic DNA MTases of
the Dnmt1/MET l class, the derived amino acid sequences of OsMET1-1 and
OsMET1-2 suggest that they are comprised of two-thirds regulatory
domain and one-third catalytic domain. Most functional domains
identified for other MTases were present in the rice MET1 sequences.
Amino acid sequence comparison indicated high similarity (56-75%
identity) of rice MET1 proteins to other plant MET1 sequences but
limited similarity (approx. 24% identity) to animal Dnmt1 proteins.
Genomic blot and database analysis indicated the presence of a single
copy of OsMET1-1 (on chromosome 3) and single copy of OsMET1-2 (on
chromosome 7). Ribonuclease protection assays revealed expression of
both OsMET1-1 and OsMET1-2 in highly dividing cells, but the
steady-state level of OsMET1-2 was 7- to 12-fold higher than that for
OsMET1-1 in callus, root and inflorescence. The functional involvement
of the rice DNA MTases in gene silencing was investigated using an RNAi
strategy. Inverted repeat constructs of either the N- or C-terminal
regions of OsMET1-1 were supertransformed into calli derived from a
rice line bearing a silenced 35S-uidA-nos transgene. Restoration of
uidA expression in the bombarded calli was consistent with the
inactivation of maintenance methylation and with previous evidence for
the involvement of methylation in silencing of this line.

<>

<1>Teerawanichpan, P., Jiang, Y., Dong, J., Hall, T.C., Narangajavana, J.
<2>DNA methyltransferase and developmental processes in rice.
<3>Plant Biol.
<4>0
<5>155-156
<6>2001
<7>Expression of antisense constructs of Arabidopsis methyltransferase (METI) results in
decreased methylation and multiple morphological abnormalities. To explore the involvement of
DNA methylation in monocot plant development and gene silencing, the endogenous Dnmt1 of rice
(GenBank AF155874) was debilitated by either antisense RNA or dsRNA. N- and C- terminal
regions (N-Dnmt1 and C-Dnmt1) of rice Dnmt1 in pRM1 were separately cloned into a binary
vector (pPT2) in order to investigate the importance of each domain in developmental
processes. Each gene construct had a maize ubiquitin promoter, a nos terminator and included
either gfp or uidA as a reporter. Agrobacterium-mediated transformation was used to introduce
either antisense (as-N-Dnmt1 and as-C-Dnmt1) or inverted repeat (ir-N-Dnmt1 or ir-C-Dnmt1)
constructs of Dnmt1 into rice in order to silence expression of endogenous Dnmt1. Compared
with wt plants, transgenic plants PT41-1, PT41-2 and PT41-3 (carrying as-N-Dnmt1) showed
decreased fertility (80-100%), smaller leaves and reduced height (33%). Plants PT41-1 and -2
had 7-10 tillers while PT41-3 had multiple (20) tillers. Various defects in flower morphology
were noted, including stunted flowers with curled lemma and palea; stamen with short
filaments, and anthers with little pollen. Plants containing the dsRNA (ir) constructs are in
regeneration. Detailed molecular analyses of the methylation-deficient transformants are
underway and the relationship between the severity of phenotypic defects and DNA methylation
levels will be determined. The ability of the constructs to release silencing in other rice
lines is being evaluated using both in vitro and in vivo approaches.

<>

<1>Teerawanichpan, P., Krittanai, P., Chauvatcharin, N., Narangajavana, J.
<2>Purification and characterization of rice DNA methyltransferase.
<3>Plant Physiol. Biochem.
<4>47
<5>671-680
<6>2009
<7>Epigenetic modification is essential for normal development and plays important roles in gene
regulation in higher plants. Multiple factors
interact to regulate the establishment and maintenance of DNA
methylation in plant genome. We had previously cloned and characterized
DNA methyltransferase (DNA MTase) gene homologues (OsMET1) from rice.
In this present study, determination of DNA MTase activity in different
cellular compartments showed that DNA MTase was enriched in nuclei and
the activity was remarkably increased during imbibing dry seeds. We had
optimized the purification technique for DNA MTase enzyme from shoots
of 10-day-old rice seedlings using the three successive chromatographic
columns. The Econo-Pac Q the Hitrap-Heparin and the Superdex-200
columns yielded a protein fraction of a specific activity of 29, 298
and 800 purification folds, compared to the original nuclear extract,
respectively. The purified protein preferred hemi-methylated DNA
substrate, suggesting the maintenance activity of methylation. The
native rice DNA MTase was approximately 160-170 kDa and exhibited a
broad pH optimum in the range of 7.6 and 8.0. The enzyme kinetics and
inhibitory effects by methyl donor analogs, base analogs, cations, and
cationic amines on rice DNA MTase were examined. Global cytosine
methylation status of rice genome during development and in various
tissue culture systems were monitored and the results suggested that
the cytosine methylation level is not directly correlated with the DNA
MTase activity. The purification and characterization of rice DNA MTase
enzyme are expected to enhance our understanding of this enzyme
function and their possible contributions in Gramineae plant
development.

<>

<1>Teh, B.S., Abdul, R.A.Y., Saito, J.A., Hou, S., Alam, M.
<2>Complete Genome Sequence of the Thermophilic Bacterium Thermus sp. Strain CCB_US3_UF1.
<3>J. Bacteriol.
<4>194
<5>1240
<6>2012
<7>Thermus sp. strain CCB_US3_UF1, a thermophilic bacterium, has been isolated from  a hot spring
in Malaysia. Here, we present the complete genome sequence of
Thermus sp. CCB_US3_UF1.

<>

<1>Tekedar, H.C., Karsi, A., Gillaspy, A.F., Dyer, D.W., Benton, N.R., Zaitshik, J., Vamenta, S., Banes, M.M., Gulsoy, N., Aboko-Cole, M., Waldbieser, G.C., Lawrence, M.L.
<2>Genome Sequence of the Fish Pathogen Flavobacterium columnare ATCC 49512.
<3>J. Bacteriol.
<4>194
<5>2763-2764
<6>2012
<7>Flavobacterium columnare is a Gram-negative, rod-shaped, motile, and highly prevalent fish
pathogen causing columnaris disease in freshwater fish worldwide.
Here, we present the complete genome sequence of F. columnare strain ATCC 49512.

<>

<1>Telford, J., Masignani, V., Grandi, G., Fraser, C., Tettelin, H., Scarselli, M.
<2>Nucleic and proteins from streptococcus groups A.
<3>European Patent Office
<4>EP 2189473 A
<5>
<6>2010
<7>The invention provides proteins from group B
streptococcus (Streptococcus agalactiae) and group A
streptococcus (Streptococcus pyogenes), including amino
acid sequences and the corresponding nucleotide sequences.
Data are given to show that the proteins are
useful antigens for vaccines, immunogenic compositions,
and/or diagnostics. The proteins are also targets
for antibiotics.

<>

<1>Telford, J., Masignani, V., Margarit-y-Ros, I., Grandi, G., Fraser, C., Tettelin, H.
<2>Nucleic acids and proteins from Streptococcus groups A & B.
<3>International Patent Office
<4>WO 03093306 A
<5>
<6>2003
<7>The invention provides proteins from group B streptococcus (Streptococcus agalactiae) and
group A streptococcus (Streptococcus pyogenes), including amino acid sequences and the
corresponding nucleotide sequences.

<>

<1>Telford, J., Masignani, V., Margarit-y-Ros, I., Grandi, G., Fraser, C., Tettelin, H.
<2>Nucleic acids and proteins from streptococcus groups a.
<3>International Patent Office
<4>WO 0234771 A
<5>
<6>2002
<7>The invention provides proteins from group B streptococcus (Streptococcus agalactiae) and
group A streptococcus (Streptococcus pyogenes), including amino acid sequences and the
corresponding nucleotide sequences.

<>

<1>Telford, J., Masignani, V., Ross, I.M.I., Grandi, G., Fraser, C., Tetterin, H.
<2>NUCLEIC ACIDS AND PROTEINS FROM STREPTOCOCCUS GROUPS A and B.
<3>Japanese Patent Office
<4>JP 2005289966 A
<5>
<6>2005
<7>
<>

<1>Temperton, B., Thomas, S., Tait, K., Parry, H., Emery, M., Allen, M., Quinn, J., Macgrath, J., Gilbert, J.
<2>Permanent draft genome sequence of Vibrio tubiashii strain NCIMB 1337 (ATCC19106).
<3>Standards in Genomic Sciences
<4>4
<5>183-190
<6>2011
<7>Vibrio tubiashii NCIMB 1337 is a major and increasingly prevalent pathogen of bivalve
mollusks, and shares a close phylogenetic relationship with both V.
orientalis and V. coralliilyticus. It is a Gram-negative, curved rod-shaped
bacterium, originally isolated from a moribund juvenile oyster, and is both
oxidase and catalase positive. It is capable of growth under both aerobic and
anaerobic conditions. Here we describe the features of this organism, together
with the draft genome and annotation. The genome is 5,353,266 bp long, consisting
of two chromosomes, and contains 4,864 protein-coding and 86 RNA genes.

<>

<1>Tengs, T., LaFramboise, T., Den RB, H.D.N., Zhang, J., DebRoy, S., Gentleman, R.C., O'Neill, K., Birren, B., Meyerson, M.
<2>Genomic representations using concatenates of Type IIB restriction endonuclease digestion fragments.
<3>Nucleic Acids Res.
<4>32
<5>e121
<6>2004
<7>We have developed a method for genomic representation using Type IIB restriction
endonucleases. Representation by concatenation of restriction
digests, or RECORD, is an approach to sample the fragments generated by
cleavage with these enzymes. Here, we show that the RECORD libraries may
be used for digital karyotyping and for pathogen identification by
computational subtraction.

<>

<1>Tenover, F.C., Clark, V.L., Young, F.E.
<2>Presence of methyl adenine in the DNA of some strains of Neisseria Gonorrhoeae.
<3>Biochem. Biophys. Res. Commun.
<4>95
<5>1796-1800
<6>1980
<7>Sixteen strains of Neisseria gonorrhoeae were examined for the presence of
methyl adenine using the site-specific restriction endonucleases MboI and DpnI.
The DNA of four strains, all of which require arginine, hypoxanthine and
uracil for growth, contained methyl adenine.  Plasmid DNA from a
non-methylating strain transformed into methylating strains contained methyl
adenine when re-isolated.

<>

<1>Teo, J., Tan, S.Y., Tay, M., Ding, Y., Kjelleberg, S., Givskov, M., Lin, R.T., Yang, L.
<2>First case of E anophelis outbreak in an intensive-care unit.
<3>Lancet
<4>382
<5>855-856
<6>2013
<7>The hospital infection-control team at the National University Hospital of Singapore
identified three patients in the cardiothoracic intensive-care unit and two patients from the
surgical ICU that were colonized with Elizabethkingia during a 3 week period in 2012.  The
Elizabethkingia strains were identified as Elizabethkingia meningoseptica on the basis of
matrix-assisted laser desorption-ionization time-of-flight mass spectrometry analysis.  The
five patients, who were ventilated via tracheostomy and had central venous catheters in situ,
received multiple courses of brad-spectrum antibiotics.  Before isolation of Elizabethkingia,
three of the patients had underlying solid-organ malignancy, one patient had multiple
abdominal surgeries, two patients underwent thoracic surgery, and one patient was on
extracorporeal membrane oxygenation.  After isolation of the Elizabethkingia strain, all
patients were treated with intravenous piperacillin and taxobactam, cotrimoxazole, or
levofloxacin, either alone or in combination.  Three of the five patients died during their
intensive-care admission, with sepsis contributing to the death of two patients.  Isolates of
Elizabethkingia (designated as NUHP1, NUHP2, and NUHP3) were obtained from the three patients
who had been warded at the cardiothoracic ICU.  NUHP1 and NUHP3 isolates were recovered from
the sputum, whereas NUHP2 was isolated froma blood specimen.  Unfortunately, isolates from
patients who had been warded in the surgical ICU were no longer available for further
analysis.

<>

<1>Terabayashi, Y., Juan, A., Tamotsu, H., Ashimine, N., Nakano, K., Shimoji, M., Shiroma, A., Teruya, K., Satou, K., Hirano, T.
<2>First Complete Genome Sequence of Salmonella enterica subsp. enterica Serovar Typhimurium Strain ATCC 13311 (NCTC 74), a Reference Strain of Multidrug  Resistance, as Achieved by Use of PacBio Single-Molecule Real-Time Technology.
<3>Genome Announcements
<4>2
<5>e00986-14
<6>2014
<7>We report the first complete genomic sequence of Salmonella enterica subsp. enterica serovar
Typhimurium strain ATCC 13311, the leading food-borne pathogen
and a reference strain used in drug resistance studies. De novo assembly with
PacBio sequencing completed its chromosome and one plasmid. They will accelerate
the investigation into multidrug resistance in Salmonella Typhimurium.

<>

<1>Terfehr, D., Dahlmann, T.A., Specht, T., Zadra, I., Kurnsteiner, H., Kuck, U.
<2>Genome Sequence and Annotation of Acremonium chrysogenum, Producer of the beta-Lactam Antibiotic Cephalosporin C.
<3>Genome Announcements
<4>2
<5>e00948-14
<6>2014
<7>The filamentous fungus Acremonium chrysogenum is the industrial producer of the beta-lactam
antibiotic cephalosporin C. Here, we present the genome sequence of
strain ATCC 11550, which contains genes for 8,901 proteins, 127 tRNAs, and 22
rRNAs. Genome annotation led to the prediction of 42 gene clusters for secondary
metabolites.

<>

<1>Terpolilli, J., Garau, G., Hill, Y., Tian, R., Howieson, J., Brau, L., Goodwin, L., Han, J., Liolios, K., Huntemann, M., Pati, A., Woyke, T., Mavromatis, K., Markowitz, V., Ivanova, N., Kyrpides, N., Reeve, W.
<2>Genome sequence of Ensifer medicae strain WSM1369; an effective microsymbiont of  the annual legume Medicago sphaerocarpos.
<3>Standards in Genomic Sciences
<4>9
<5>420-430
<6>2013
<7>Ensifer medicae WSM1369 is an aerobic, motile, Gram-negative, non-spore-forming rod that can
exist as a soil saprophyte or as a legume microsymbiont of Medicago.
WSM1369 was isolated in 1993 from a nodule recovered from the roots of Medicago
sphaerocarpos growing at San Pietro di Rudas, near Aggius in Sardinia (Italy).
WSM1369 is an effective microsymbiont of the annual forage legumes M. polymorpha
and M. sphaerocarpos. Here we describe the features of E. medicae WSM1369,
together with genome sequence information and its annotation. The 6,402,557 bp
standard draft genome is arranged into 307 scaffolds of 307 contigs containing
6,656 protein-coding genes and 79 RNA-only encoding genes. This rhizobial genome
is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 Genomic
Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

<>

<1>Terpolilli, J., Hill, Y., Tian, R., Howieson, J., Brau, L., Goodwin, L., Han, J., Liolios, K., Huntemann, M., Pati, A., Woyke, T., Mavromatis, K., Markowitz, V., Ivanova, N., Kyrpides, N., Reeve, W.
<2>Genome sequence of Ensifer meliloti strain WSM1022; a highly effective microsymbiont of the model legume Medicago truncatula A17.
<3>Standards in Genomic Sciences
<4>9
<5>315-324
<6>2013
<7>Ensifer meliloti WSM1022 is an aerobic, motile, Gram-negative, non-spore-forming  rod that can
exist as a soil saprophyte or as a legume microsymbiont of Medicago.
WSM1022 was isolated in 1987 from a nodule recovered from the roots of the annual
Medicago orbicularis growing on the Cyclades Island of Naxos in Greece. WSM1022
is highly effective at fixing nitrogen with M. truncatula and other annual
species such as M. tornata and M. littoralis and is also highly effective with
the perennial M. sativa (alfalfa or lucerne). In common with other characterized
E. meliloti strains, WSM1022 will nodulate but fixes poorly with M. polymorpha
and M. sphaerocarpos and does not nodulate M. murex. Here we describe the
features of E. meliloti WSM1022, together with genome sequence information and
its annotation. The 6,649,661 bp high-quality-draft genome is arranged into 121
scaffolds of 125 contigs containing 6,323 protein-coding genes and 75 RNA-only
encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE
Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root
Nodule Bacteria (GEBA-RNB) project.

<>

<1>Terpolilli, J., Rui, T., Yates, R., Howieson, J., Poole, P., Munk, C., Tapia, R., Han, C., Markowitz, V., Tatiparthi, R., Mavrommatis, K., Ivanova, N., Pati, A., Goodwin, L., Woyke, T., Kyrpides, N., Reeve, W.
<2>Genome sequence of Rhizobium leguminosarum bv trifolii strain WSM1689, the microsymbiont of the one flowered clover Trifolium uniflorum.
<3>Standards in Genomic Sciences
<4>9
<5>527-539
<6>2014
<7>Rhizobium leguminosarum bv. trifolii is a soil-inhabiting bacterium that has the  capacity to
be an effective N2-fixing microsymbiont of Trifolium (clover)
species. R. leguminosarum bv. trifolii strain WSM1689 is an aerobic, motile,
Gram-negative, non-spore-forming rod that was isolated from a root nodule of
Trifolium uniflorum collected on the edge of a valley 6 km from Eggares on the
Greek Island of Naxos. Although WSM1689 is capable of highly effective
N2-fixation with T. uniflorum, it is either unable to nodulate or unable to fix
N2 with a wide range of both perennial and annual clovers originating from
Europe, North America and Africa. WSM1689 therefore possesses a very narrow host
range for effective N2 fixation and can thus play a valuable role in determining
the geographic and phenological barriers to symbiotic performance in the genus
Trifolium. Here we describe the features of R. leguminosarum bv. trifolii strain
WSM1689, together with the complete genome sequence and its annotation. The
6,903,379 bp genome contains 6,709 protein-coding genes and 89 RNA-only encoding
genes. This multipartite genome contains six distinct replicons; a chromosome of
size 4,854,518 bp and five plasmids of size 667,306, 518,052, 341,391, 262,704
and 259,408 bp. This rhizobial genome is one of 20 sequenced as part of a DOE
Joint Genome Institute 2010 Community Sequencing Program.

<>

<1>Terry, B.J., Jack, W.E., Modrich, P.
<2>Facilitated diffusion during catalysis by EcoRI endonuclease:  nonspecific interactions in EcoRI catalysis.
<3>J. Biol. Chem.
<4>260
<5>13130-13137
<6>1985
<7>The potential for processive EcoRI endonuclease hydrolysis has been examined on
several DNA substrates containing two EcoRI sites which were embedded in
identical sequence environments.  With a 388-base pair circular DNA, in which
the two recognition sites are separated by 51 base pairs (shorter distance) or
337 base pairs (longer distance), 77 and 34% of all events involved processive
hydrolysis at ionic strengths of 0.059 and 0.13, respectively.  However, the
frequency of processive action on linear substrates, in which the two sites
were separated by 51 base pairs, was only 42 and 17% at these ionic strengths,
values half those observed with the circular DNA.  Processive action not
detectable on circular or linear substrates at an ionic strength of 0.23.
These findings indicate that DNA search by the endonuclease occurs by
facilitated diffusion, a mechanism in which the protein locates and leaves it
recognition sequence by interacting with nonspecific DNA sites.  We suggest
that processivity on linear substrates is limited to values half that for small
circles due to partitioning of the enzyme between the two products generated by
cleavage of a linear molecule.  Given such topological effects, measured
processivity values imply that the endonuclease can diffuse within a DNA domain
to locate and recognize an EcoRI site 50 to 300 base pairs distant from an
initial binding site, with minimum search efficiencies being 80 and 30% at
ionic strengths of 0.059 and 0.13, respectively.  The high efficiency of
processive action indicates that a positionally correlated mode of search plays
a major role in facilitated diffusion in this system under such conditions.
Also consistent with this view was the identication of a striking position
effect when two closely spaced EcoRI sites were asymmetrically positioned near
the end of a linear DNA.  The endonuclease displays a substantial preference
for the more centrally located recognition sequence.  This preference does not
reflect differential sensitivity of the two sites to cleavage per se, but can
be simply explained by preferential entry of the enzyme via the larger
nonspecific target available to the more centrally positioned recognition
sequence.  These conclusions differ from those of a previous qualitative
analysis of endonuclease processivity over short distances.

<>

<1>Terry, B.J., Jack, W.E., Modrich, P.
<2>Mechanism of specific site location and DNA cleavage by EcoRI endonuclease.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>103-118
<6>1987
<7>The EcoRI restriction and modification system has become one of the most
convenient systems in which to study protein-DNA interactions.  Both EcoRI
endonuclease and methylase recognize a two-fold symmetrical DNA sequence.  The
endonuclease cleaves at sites indicated by arrows while the central adenines
are methylated on the 6-amino group by the methylase (*), with the latter
modification rendering the EcoRI sequence resistant to cleavage by the
endonuclease.  Like all sequence specific proteins, EcoRI endonuclease displays
finite affinity for nonspecific DNA sequences.  In this context, the problem of
specific recognition can be divided into two questions:  (i) what is the
mechanistic basis for discrimination of specific and nonspecific sites in
thermodynamic and molecular terms?  (ii) what role do nonspecific interactions
play in the kinetic path(s) utilized for specific site location?  In this paper
we will summarize work on the thermodynamics of specific and nonspecific
interactions and will discuss evidence indicating that nonspecific interactions
are involved in the pathways by which EcoRI endonuclease locates an EcoRI
sequence and leaves a cleaved site.  For discussion of molecular details of
endonuclease DNA interactions, see chapter by Rosenberg, et al., this volume.

<>

<1>Terry, B.J., Jack, W.E., Rubin, R.A., Modrich, P.
<2>Thermodynamic parameters governing interaction of EcoRI endonuclease with specific and nonspecific DNA sequences.
<3>J. Biol. Chem.
<4>258
<5>9820-9825
<6>1983
<7>Equilibrium binding of EcoRI endonuclease to DNA has been analyzed by
nitrocellulose filter and preferential DNA cleavage methods.  Association
constants for pBR322 and a 34-base pair molecule containing the EcoRI site of
this plasmid in a central position were determined to be 1.9 x 10(11) M-1 and
1.0 x 10(11) M-1 at 37C, respectively, with the stoichiometry of binding being
0.8-+ 0.1 mol of endonuclease dimer per mol of DNA.  In contrast, the affinity
of the enzyme for a pBR322 derivative from which the EcoRI site has been
deleted is 3.2 x 10(9) M-1 as judged by competitive binding experiments.  If it
is assumed that each base pair can define the beginning of a nonspecific
binding site, this value corresponds to an affinity for nonspecific sites of
7.4 x 10(5) M-1.  Furthermore, the affinity of the endonuclease for the
EcoRI-methylated sequence is at least three orders of magnitude less than that
for the unmodified recognition site.  The dependence on temperature and ionic
strength of the equilibrium constant governing specific interactions has also
been examined.  The temperature dependence of the reaction indicates that
entropy increase accounts for 70% of the free energy of specific binding at
37C.  Affinity of the endonuclease for the EcoRI site is highly dependent on
NaCl concentration.  Analysis of this dependence according to the theory of
Record and colleagues (Record, T.M., Jr., Lohman, T.M., and deHaseth, P. (1976)
J. Mol. Biol. 107, 145-158) has implicated 8 ion pairs in the stability of
specific complexes, a value identical with the number of phosphate contacts
determined by ethylation interference analysis (Lu, A.L., Jack, W.E., and
Modrich, P. (1981) J. Biol. Chem. 256, 13200-13206).  Extrapolation to 1 M NaCl
suggests that nonelectrostatic interactions account for 40% of the free energy
change associated with specific complex formation.

<>

<1>Terschuren, P.-A., Noyer-Weidner, M., Trautner, T.A.
<2>Recombinant derivatives of Bacillus subtilis phage Z containing the DNA methyltransferase genes of related methylation-proficient phages.
<3>J. Gen. Microbiol.
<4>198
<5>945-952
<6>1987
<7>The DNA methyltransferase (Mtase) genes of temperate Bacillus subtilis phages
SPR, Phi3T, SPbeta and alpha11 can be transferred by transfection and
recombination to the genome of the related nonmodifying phage Z.  Integration
of the Mtase genes occurs in phage Z DNA at a unique location which is
homologous with the flanking regions of the Mtase genes of the related phages.
In lysogenic cells carrying recombinant phages, expression of the Mtase genes
if repressed, irrespective of whether the Mtase genes were derived from phage
donors which were homo- or heteroimmune to phage Z.

<>

<1>Tesfazgi-Mebrhatu, M., Wywial, E., Ghosh, A., Michiels, C.W., Lindner, A.B., Taddei, F., Bujnicki, J.M., Van Melderen, L., Aertsen, A.
<2>Evidence for an evolutionary antagonism between Mrr and Type III modification systems.
<3>Nucleic Acids Res.
<4>39
<5>5991-6001
<6>2011
<7>The Mrr protein of Escherichia coli is a laterally acquired Type IV restriction endonuclease
with specificity for methylated DNA. While Mrr nuclease activity can be elicited by
high-pressure stress in E. coli MG1655, its (over)expression per se does not confer any
obvious toxicity. In this study, however, we discovered that Mrr of E. coli MG1655 causes
distinct genotoxicity when expressed in Salmonella typhimurium LT2. Genetic screening enabled
us to contribute this toxicity entirely to the presence of the endogenous Type III restriction
modification system (StyLTI) of S. typhimurium LT2. The StyLTI system consists of the Mod DNA
methyltransferase and the Res restriction endonuclease, and we revealed that expression of the
LT2 mod gene was sufficient to trigger Mrr activity in E. coli MG1655. Moreover, we could
demonstrate that horizontal acquisition of the MG1655 mrr locus can drive the loss of
endogenous Mod functionality present in S. typhimurium LT2 and E. coli ED1a, and observed a
strong anti-correlation between close homologues of MG1655 mrr and LT2 mod in the genome
database. This apparent evolutionary antagonism is further discussed in the light of a
possible role for Mrr as defense mechanism against the establishment of epigenetic regulation
by foreign DNA methyltransferases.

<>

<1>Tettelin, H. et al.
<2>Complete genome sequence of Neisseria meningitidis serogroup B strain MC58.
<3>Science
<4>287
<5>1809-1815
<6>2000
<7>The 2,272,351-base pair genome of Neisseria meningitidis strain MC58 (serogroup B), a
causative agent of meningitis and septicemia, contains 2158 predicted coding regions, 1158
(53.7%) of which were assigned a biological role. Three major islands of horizontal DNA
transfer were identified; two of these contain genes encoding proteins involved in
pathogenicity, and the third island contains coding sequences only for hypothetical proteins.
Insights into the commensal and virulence behavior of N. meningitidis can be gleaned from the
genome, in which sequences for structural proteins of the pilus are clustered and several
coding regions unique to serogroup B capsular polysaccharide synthesis can be identified.
Finally, N. meningitidis contains more genes that undergo phase variation than any pathogen
studied to date, a mechanism that controls their expression and contributes to the evasion of
the host immune system.

<>

<1>Tettelin, H. et al.
<2>Complete genome sequence of a virulent isolate of Streptococcus pneumoniae.
<3>Science
<4>293
<5>498-506
<6>2001
<7>The 2,160,837-base pair genome sequence of an isolate of Streptococcus pneumoniae, a
Gram-positive pathogen that causes pneumonia, bacteremia, meningitis, and otitis media,
contains 2236 predicted coding regions; of these, 1440 (64%) were assigned a biological role.
Approximately 5% of the genome is composed of insertion sequences that may contribute to
genome rearrangements through uptake of foreign DNA. Extracellular enzyme systems for the
metabolism of polysaccharides and hexosamines provide a substantial source of carbon and
nitrogen for S. pneumoniae and also damage host tissues and facilitate colonization. A motif
identified within the signal peptide of proteins is potentially involved in targeting these
proteins to the cell surface of low-guanine/cytosine (GC) Gram-positive species. Several
surface-exposed proteins that may serve as potential vaccine candidates were identified.
Comparative genome hybridization with DNA arrays revealed strain differences in S. pneumoniae
that could contribute to differences in virulence and antigenicity.

<>

<1>Tettelin, H. et al.
<2>Genome analysis of multiple pathogenic isolates of Streptococcus agalactiae: Implications for the microbial "pan-genome".
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>102
<5>13950
<6>2005
<7>The development of efficient and inexpensive genome sequencing methods has revolutionized the
study of human bacterial pathogens and improved vaccine design. Unfortunately, the sequence of
a single genome does not reflect how genetic variability drives pathogenesis within a
bacterial species and also limits genome-wide screens for vaccine candidates or for
antimicrobial targets. We have generated the genomic sequence of six strains representing the
five major disease-causing serotypes of Streptococcus agalactiae, the main cause of neonatal
infection in humans. Analysis of these genomes and those available in databases showed that
the S. agalactiae species can be described by a pan-genome consisting of a core genome shared
by all isolates, accounting for approximately 80% of any single genome, plus a dispensable
genome consisting of partially shared and strain-specific genes. Mathematical extrapolation of
the data suggests that the gene reservoir available for inclusion in the S. agalactiae
pan-genome is vast and that unique genes will continue to be identified even after sequencing
hundreds of genomes.

<>

<1>Tettelin, H., Sampaio, E.P., Daugherty, S.C., Hine, E., Riley, D.R., Sadzewicz, L., Sengamalay, N., Shefchek, K., Su, Q., Tallon, L.J., Conville, P., Olivier, K.N., Holland, S.M., Fraser, C.M., Zelazny, A.M.
<2>Genomic Insights into the Emerging Human Pathogen Mycobacterium massiliense.
<3>J. Bacteriol.
<4>194
<5>5450
<6>2012
<7>Mycobacterium massiliense (Mycobacterium abscessus group) is an emerging pathogen causing
pulmonary disease and skin and soft tissue infections. We report the
genome sequence of the type strain CCUG 48898.

<>

<1>Thanos, D., Scarpelis, G., Papamatheakis, J., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BseAI.
<3>Nucleic Acids Res.
<4>17
<5>8881
<6>1989
<7>None

<>

<1>The, A.G.I.
<2>Analysis of the genome sequence of the flowering plant Arabidopsis thaliana.
<3>Nature
<4>408
<5>796-815
<6>2000
<7>The flowering plant Arabidopsis thaliana is an important model system for identifying genes
and determining their functions.  Here we report the analysis of the genomic sequence of
Arabidopsis.  The sequenced regions cover 115.4 megabases of the 125-megabase genome and
extend into centromeric regions.  The evolution of Arabidopsis involved a whole-genome
duplication, followed by subsequent gene loss and extensive local gene duplications, giving
rise to a dynamic genome enriched by lateral gene transfer from a cyanobacterial-like ancestor
of the plastid.  The genome contains 25,498 genes encoding proteins from 11,000 families,
similar to the functional diversity of Drosophila and Caenorhabditis elegans-the other
sequenced multicellular eukaryotes.  Arabidopsis has many families of new proteins but also
lacks several common protein families, indicating that the sets of common proteins have
undergone differential expansion and contraction in the three multicellular eukaryotes.  This
is the first complete genome sequence of a plant and provides the foundations for more
comprehensive comparison of conserved processes in all eukaryotes, identifying a wide range of
plant-specific gene functions and establishing rapid systematic ways to identify genes for
crop improvement.

<>

<1>Thennarasu, S., Polireddy, D., Antony, A., Yada, M.R., Algarawi, S., Sivakumar, N.
<2>Draft Genome Sequence of a Highly Flagellated, Fast-Swimming Archaeon, Methanocaldococcus villosus Strain KIN24-T80 (DSM 22612).
<3>Genome Announcements
<4>1
<5>e00481-13
<6>2013
<7>We report the draft genome sequence of a hyperthermophilic Methanocaldococcus villosus strain,
KIN24-T80. The gene associated with its heavy flagellum
formation was annotated in the 1.2-Mb draft genome sequence, and this strain may
be a good model system to study the extensive functional role of flagella and
their fast motor activity.

<>

<1>Theriault, G., Roy, P.H.
<2>Cloning of Pseudomonas plasmid pMG7 and its restriction-modification system in Escherichia coli.
<3>Gene
<4>19
<5>355-359
<6>1982
<7>Plasmid pMG7 of Pseudomonas aeruginosa codes for a type II DNA
restriction-modification (r-m) system, PaeR7.  This plasmid has not been
observed to transfer to Escherichia coli either by conjugation or by
transformation.  We have cloned BglII linears (42 kb) and the BamHI large
fragment (37 kb) of pMG7 into cosmid pHC79 (6.4 kb) and introduced the
recombinant molecules into E. coli by in vitro packaging.  Several clones were
obtained which demonstrated in vivo restriction of phage u80.  One of these
clones, GT138, was further tested and showed in vivo modification of Phi80.
Extracts from two clones, GT138 and GT125, yielded a restriction endonuclease
activity which produced fragments of u80 DNA identical to those produced by
PaeR7.  Cosmid cloning should be useful for obtaining substantial yields of
large fragments of plasmids that are difficult to purify in their native
strains.

<>

<1>Theriault, G., Roy, P.H., Howard, K.A., Benner, J.S., Brooks, J.S., Waters, A.F., Gingeras, T.R.
<2>Nucleotide sequence of the PaeR7 restriction/modification system and partial characterization of its protein products.
<3>Nucleic Acids Res.
<4>13
<5>8441-8461
<6>1985
<7>Bal31 deletion experiments on clones of the PaeR7 restriction-modification
system from Pseudomonas aeruginosa demonstrate that it is arranged as an
operon, with the methylase gene preceding the endonuclease gene.  The DNA
sequence of this operon agrees with in vitro transcription-translation assays
which predict proteins of 532 amino acids, Mr = 59,260 daltons, and 246 amino
acids, Mr = 27,280 daltons, coincident with the methylase and endonuclease
genes, respectively.  These predicted values coincide with the measured
molecular weights of the purified, denatured PaeR7 endonuclease and methylase
proteins.  The first twenty amino acids from the amino-terminus of the purified
endonuclease exactly match those predicted from the DNA sequence.  Finally,
potential regulatory mechanisms for the expression of phage restriction are
described based on the properties of several PaeR7 subclones.

<>

<1>Thiel, T., Pratte, B.S., Zhong, J., Goodwin, L., Copeland, A., Lucas, S., Han, C., Pitluck, S., Land, M.L., Kyrpides, N.C., Woyke, T.
<2>Complete genome sequence of Anabaena variabilis ATCC 29413.
<3>Standards in Genomic Sciences
<4>9
<5>562-573
<6>2014
<7>Anabaena variabilis ATCC 29413 is a filamentous, heterocyst-forming cyanobacterium that has
served as a model organism, with an extensive literature
extending over 40 years. The strain has three distinct nitrogenases that function
under different environmental conditions and is capable of photoautotrophic
growth in the light and true heterotrophic growth in the dark using fructose as
both carbon and energy source. While this strain was first isolated in 1964 in
Mississippi and named Anabaena flos-aquae MSU A-37, it clusters phylogenetically
with cyanobacteria of the genus Nostoc. The strain is a moderate thermophile,
growing well at approximately 40( degrees ) C. Here we provide some additional
characteristics of the strain, and an analysis of the complete genome sequence.

<>

<1>Thiel, V., Hamilton, T.L., Tomsho, L.P., Burhans, R., Gay, S.E., Ramaley, R.F., Schuster, S.C., Steinke, L., Bryant, D.A.
<2>Draft Genome Sequence of the Moderately Thermophilic Bacterium Schleiferia thermophila Strain Yellowstone (Bacteroidetes).
<3>Genome Announcements
<4>2
<5>e00860-14
<6>2014
<7>The draft genome sequence of the moderately thermophilic bacterium Schleiferia thermophila
strain Yellowstone (Bacteroidetes), isolated from Octopus Spring
(Yellowstone National Park, WY, USA) was sequenced and comprises 2,617,694 bp in
35 contigs. The draft genome is predicted to encode 2,457 protein coding genes
and 37 tRNA encoding genes and two rRNA operons.

<>

<1>Thiel, V., Hamilton, T.L., Tomsho, L.P., Burhans, R., Gay, S.E., Schuster, S.C., Ward, D.M., Bryant, D.A.
<2>Draft Genome Sequence of a Sulfide-Oxidizing, Autotrophic Filamentous Anoxygenic  Phototrophic Bacterium, Chloroflexus sp. Strain MS-G (Chloroflexi).
<3>Genome Announcements
<4>2
<5>e00872-14
<6>2014
<7>The draft genome sequence of the thermophilic filamentous anoxygenic phototrophic bacterium
Chloroflexus sp. strain MS-G (Chloroflexi), isolated from Mushroom
Spring (Yellowstone National Park, WY, USA) was sequenced and comprises 4,784,183
bp in 251 contigs. The draft genome is predicted to encode 4,059 protein coding
genes, 49 tRNA encoding genes, and 3 rRNA operons.

<>

<1>Thielking, V.
<2>Oligodeoxynucleotides with basepair mismatches as a substrate for EcoRI and EcoRV.
<3>Ph.D. Thesis, Medizinische Hochscule Hannover, W. Germany
<4>
<5>1-84
<6>1988
<7>
<>

<1>Thielking, V., Alves, J., Fliess, A., Maass, G., Pingoud, A.
<2>Accuracy of the EcoRI restriction endonuclease:  Binding and cleavage studies with oligodeoxynucleotide substrates containing degenerate recognition sequences.
<3>Biochemistry
<4>29
<5>4682-4691
<6>1990
<7>We have synthesized a series of 18 non palindromic oligodeoxynucleotides that
carry all possible base changes within the recognition sequence of EcoRI.
These single strands can be combined with their complementary single strands to
obtain all possible EcoRI sequences (left), or they can be combined with a
single strand containing the canonical sequence to obtain double strands with
all possible mismatches within the recognition sequence (right):
GCGCAAATTCCGCG               GCGCAAATTCCGCG
CGCGTTTAAGGCGC               CGCGCTTAAGGCGC
The rate of phosphodiester bond cleavage of these oligodeoxynucleotides by
EcoRI was determined in single-turnover experiments under normal buffer
conditions in order to find out to what extent the canonical recognition site
can be distorted and yet serve as a substrate for EcoRI.  Our results show that
oligodeoxynucleotides containing mismatch base pairs are in general more
readily attacked by EcoRI than oligodeoxynucleotides containing EcoRI sites and
that the rates of cleavage of the two complementary strands of degenerate
oligodeoxynucleotides are quite different.  We have also determined the
affinities of these oligodeoxynucleotides to EcoRI.  They are higher for
oligodeoxynucleotides carrying a mismatch within the EcoRI recognition site
than for oligodeoxynucleotides containing an EcoRI site but otherwise do not
correlate with the rate with which these oligodeoxynucleotides are cleaved by
EcoRI.  Our results allow details to be given for the probability of EcoRI
making mistakes in cleaving DNA not only in its recognition sequence but also
in sequences closely related to it.  Due to the fact that the rates of cleavage
in the two strands of a degenerate sequence generally are widely different,
these mistakes are most likely not occurring in vivo since nicked intermediates
can be repaired by DNA ligase.

<>

<1>Thielking, V., Du Bois, S., Eritja, R., Guschlbauer, W.
<2>Dam methyltransferase from Escherichia coli: Kinetic studies using modified DNA oligomers: nonmethylated substrates.
<3>Biol. Chem.
<4>378
<5>407-415
<6>1997
<7>Steady-state kinetics of the N6-adenine Dam methyltransferase have been measured using as
substrates non-self-complementary tetradecanucleotide duplexes that contain the GATC target
sequence.  Modifications in the GATC target sequence of one or both of the strands included
substitution of guanine by hypoxanthine, thymine by uracil or 5-ethyl-uracil and adenine by
diamino-purine (2-amino-adenine).  Thermodynamic parameters for the 14-mer duplexes were also
determined.  DNA methylation of duplexes containing single dI for dG substitution of the Dam
recognition site was little perturbed compared with the canonical substrate.  Replacement of
dG residues by dI in both strands resulted in a decrease of the specificity constant.
Substitution in both strands appears to be cumulative.  Substitution of the methyl-accepting
adenine residues by 2-amino-adenine resulted in surprisingly little perturbation.  Dam
methyltransferase is rather tolerant to different substitutions.  The results show much less
spread than those for the analogous hemimethylated substrates studied previously.  The absence
of the methylation marker appears to be deleterious to the specificity of the transition state
of the active complex, while the binding of the DNA substrate to the enzyme appears to be
mostly determined by the thermodynamic stability of the DNA duplex.

<>

<1>Thielking, V., Selent, U., Kohler, E., Landgraf, Z., Wolfes, H., Alves, J., Pingoud, A.
<2>Mg2+ confers DNA binding specificity to the EcoRV restriction endonuclease.
<3>Biochemistry
<4>31
<5>3727-3732
<6>1992
<7>The EcoRV mutant D90A which carries an amino acid substitution in its active center does not
cleave DNA. Therefore, it is possible to perform DNA binding experiments with the EcoRV-D90A
mutant both in the absence and in the presence of Mg2+. Like wild-type EcoRV [Taylor et al.
(1991) Biochemistry 30, 8743-8753], it does not show a pronounced specificity for binding to
its recognition site in the absence of Mg2+ as judged by the appearance of multiple shifted
bands in an electrophoresis mobility shift assay with a 377-bp DNA fragment carrying a single
EcoRV recognition sequence. In the presence of Mg2+, however, only one band corresponding to a
1:1 complex appears even with a high excess of protein over DNA. This complex most likely is
the specific one because its formation is suppressed much more effectively by a 13-bp
oligodeoxynucleotide with an EcoRV site than by a corresponding oligodeoxynucleotide without
an EcoRV site. The preferential interaction of the EcoRV-D90A mutant with specific DNA in the
presence of Mg2+ was also demonstrated directly: a 20-bp oligodeoxynucleotide with an EcoRV
site is bound with Kass=4x10/8 M-1, while a corresponding oligodeoxynucleotide without an
EcoRV site is bound with Kass<or=1x10/5 M-1. From these data it appears that Mg2+ confers DNA
binding specificity to this mutant by lowering the affinity to nonspecific sites and raising
the affinity to specific sites as compared to binding in the absence of Mg2+. It is concluded
that this is also true for wild-type EcoRV.

<>

<1>Thielking, V., Selent, U., Kohler, E., Wolfes, H., Pieper, U., Geiger, R., Urbanke, C., Winkler, F.K., Pingoud, A.
<2>Site-directed mutagenesis studies with EcoRV restriction endonuclease to identify regions involved in recognition and catalysis.
<3>Biochemistry
<4>30
<5>6416-6422
<6>1991
<7>Guided by the X-ray structure analysis of a crystalline EcoRV-d(GGGATATCCC)
complex (Winkler, in preparation), we have begun to identify functionally
important amino acid residues of EcoRV.  We show here that Asn70, Asp74,
Ser183, Asn185, Thr186, and Asn188 are most likely involved in the binding
and/or cleavage of the DNA, because their conservative substitution leads to
mutants of no or strongly reduced activity.  In addition, C-terminal amino acid
residues of EcoRV seem to be important for its activity, since their deletion
inactivates the enzyme.  Following the identification of three functionally
important regions, we have inspected the sequences of other restriction and
modification enzymes for homologous regions.  It was found that two restriction
enzymes that recognize similar sequences as EcoRV (DpnII and HincII), as well
as two modification enzymes (M.DpnII and, in a less apparent form, M.EcoRV),
have the sequence motif -SerGlyXXXAsnIleXSer- in common, which in EcoRV
contains the essential Ser183 and Asn188 residues.  Furthermore, the C-terminal
region, shown to be essential for EcoRV, is highly homologous to a similar
region in the restriction endonuclease SmaI.  On the basis of these findings we
propose that these restriction enzymes and to a certain extent also some of
their corresponding modification enzymes interact with DNA in a similar manner.

<>

<1>Thieme, F. et al.
<2>Insights into genome plasticity and pathogenicity of the plant pathogenic bacterium Xanthomonas campestris pv. vesicatoria revealed by the complete   genome sequence.
<3>J. Bacteriol.
<4>187
<5>7254-7266
<6>2005
<7>The gram-negative plant-pathogenic bacterium Xanthomonas campestris pv. vesicatoria is the
causative agent of bacterial spot disease in pepper and
tomato plants, which leads to economically important yield losses. This
pathosystem has become a well-established model for studying bacterial
infection strategies. Here, we present the whole-genome sequence of the
pepper-pathogenic Xanthomonas campestris pv. vesicatoria strain 85-10,
which comprises a 5.17-Mb circular chromosome and four plasmids. The
genome has a high G+C content (64.75%) and signatures of extensive genome
plasticity. Whole-genome comparisons revealed a gene order similar to both
Xanthomonas axonopodis pv. citri and Xanthomonas campestris pv. campestris
and a structure completely different from Xanthomonas oryzae pv. oryzae. A
total of 548 coding sequences (12.2%) are unique to X. campestris pv.
vesicatoria. In addition to a type III secretion system, which is
essential for pathogenicity, the genome of strain 85-10 encodes all other
types of protein secretion systems described so far in gram-negative
bacteria. Remarkably, one of the putative type IV secretion systems
encoded on the largest plasmid is similar to the Icm/Dot systems of the
human pathogens Legionella pneumophila and Coxiella burnetii. Comparisons
with other completely sequenced plant pathogens predicted six novel type
III effector proteins and several other virulence factors, including
adhesins, cell wall-degrading enzymes, and extracellular polysaccharides.

<>

<1>Thierry, A., Perrin, A., Boyer, J., Fairhead, C., Dujon, B., Frey, B., Schmitz, G.
<2>Cleavage of yeast and bacteriophage T7 genomes at a single site using the rare cutter endonuclease I-SceI.
<3>Nucleic Acids Res.
<4>19
<5>189-190
<6>1991
<7>It is obvious that the availability of rare cutter restriction endonucleases would facilitate
large scale genome mapping, cloning and sequencing projects.  Restriction endonucleases of
bacterial origin have recognition sites of 8 bp long, at most, and even when used in
combination with methylases they can produce cleavage specificities of up to 12 bp only.
Artificial endonucleases have been made by chemical modifications of either DNA binding
proteins or synthetic oligodeoxynucleotides.  Although this last strategy can, in principle,
be generalized to many sequences, cleavage occurs at low efficiency.  Cleavage of yeast and E.
coli genomic DNAs at a single predetermined site using a frequent cutter bacterial
endonuclease has recently been achieved by the Achille's heel method, an elegant three step
procedure combining the action of the lac repressor on artificially inserted sites with that
of a methylase.  We present here the first evidence for cleavage of total yeast genomic DNA at
a single predetermined site in a one step procedure using a new endonuclease, I-SceI, encoded
by a mobile group I intron of yeast mitochondria.  We further show that, although I-SceI is
extremely specific, cleavable sites can also be found in natural DNA.

<>

<1>Thijs, S., Van Hamme, J., Gkorezis, P., Rineau, F., Weyens, N., Vangronsveld, J.
<2>Draft Genome Sequence of Raoultella ornithinolytica TNT, a Trinitrotoluene-Denitrating and Plant Growth-Promoting Strain Isolated from  Explosive-Contaminated Soil.
<3>Genome Announcements
<4>2
<5>e00491-14
<6>2014
<7>We report the draft genome of Raoultella ornithinolytica TNT, a Gram-negative bacterium of the
Enterobacteriaceae isolated from military soil in Belgium.
Strain TNT uses nitrite released from trinitrotoluene (TNT) for growth and is a
potent plant growth promoter. An analysis of its 5.6-Mb draft genome will bring
insights into TNT degradation-reinforcing bioremediation applications.

<>

<1>Thion, L., Laurine, E., Erard, M., Burlet-Schiltz, O., Monsarrat, B., Masson, J.M., Saves, I.
<2>The two-step cleavage activity of PI-TfuI intein endonuclease demonstrated by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.
<3>J. Biol. Chem.
<4>277
<5>45442-45450
<6>2002
<7>PI-TfuI, an intein spliced from the DNA polymerase of Thermococcus fumicolans, is a highly
specific endonuclease, whose cleavage efficiency and specificity depend on both the substrate
topology and the divalent cation used as cofactor. An open-circular intermediate was observed
during the cleavage of supercoiled DNA by PI-TfuI, suggesting a two-step cleavage of the DNA.
We characterised this nicked intermediate and, through the development of a method of analysis
of the cleavage reaction based on MALDI-tof mass spectrometry, we demonstrated that the
cleavage of DNA by PI-TfuI indeed results from two cleavage events. One step results in the
cleavage of the bottom strand, that is independent of the DNA conformation or choice of the
metal ion cofactor. A second step, which is slower, leads to the cleavage of the top strand
and governs the specific requirements of PI-TfuI concerning the essential cofactor and the DNA
topology. These two steps were shown to be independent in optimal conditions of cleavage.
These data give support to the existence of two distinct and independent active sites in the
endonuclease domain of the archaeal intein.

<>

<1>Thole, S., Kalhoefer, D., Voget, S., Berger, M., Engelhardt, T., Liesegang, H., Wollherr, A., Kjelleberg, S., Daniel, R., Simon, M., Thomas, T., Brinkhoff, T.
<2>Phaeobacter gallaeciensis genomes from globally opposite locations reveal high similarity of adaptation to surface life.
<3>ISME J.
<4>6
<5>2229-2244
<6>2012
<7>Phaeobacter gallaeciensis, a member of the abundant marine Roseobacter clade, is
known to be an effective colonizer of biotic and abiotic marine surfaces.
Production of the antibiotic tropodithietic acid (TDA) makes P. gallaeciensis a
strong antagonist of many bacteria, including fish and mollusc pathogens. In
addition to TDA, several other secondary metabolites are produced, allowing the
mutualistic bacterium to also act as an opportunistic pathogen. Here we provide
the manually annotated genome sequences of the P. gallaeciensis strains DSM 17395
and 2.10, isolated at the Atlantic coast of north western Spain and near Sydney,
Australia, respectively. Despite their isolation sites from the two different
hemispheres, the genome comparison demonstrated a surprisingly high level of
synteny (only 3% nucleotide dissimilarity and 88% and 93% shared genes). Minor
differences in the genomes result from horizontal gene transfer and phage
infection. Comparison of the P. gallaeciensis genomes with those of other
roseobacters revealed unique genomic traits, including the production of
iron-scavenging siderophores. Experiments supported the predicted capacity of
both strains to grow on various algal osmolytes. Transposon mutagenesis was used
to expand the current knowledge on the TDA biosynthesis pathway in strain DSM
17395. This first comparative genomic analysis of finished genomes of two closely
related strains belonging to one species of the Roseobacter clade revealed
features that provide competitive advantages and facilitate surface attachment
and interaction with eukaryotic hosts.The ISME Journal advance online
publication, 21 June 2012; doi:10.1038/ismej.2012.62.

<>

<1>Thomas, A.T., Brammar, W.J., Wilkins, B.M.
<2>Plasmid R16 ArdA protein preferentially targets restriction activity of the type I restriction-modification system EcoKI.
<3>J. Bacteriol.
<4>185
<5>2022-2025
<6>2003
<7>The ArdA antirestriction protein of the IncB plasmid R16 selectively inhibited the restriction
activity of EcoKI, leaving significant levels of
modification activity under conditions in which restriction was almost
completely prevented. The results are consistent with the hypothesis that
ArdA functions in bacterial conjugation to allow an unmodified plasmid to
evade restriction in the recipient bacterium and yet acquire cognate
modification.

<>

<1>Thomas, C.B., Gumport, R.I.
<2>Dimerization of the bacterial RsrI N6-adenine DNA methyltransferase.
<3>Nucleic Acids Res.
<4>34
<5>806-815
<6>2006
<7>Dimeric restriction endonucleases and monomeric modification methyltransferases were long
accepted as the structural paradigm for Type
II restriction systems. Recent studies, however, have revealed an
increasing number of apparently dimeric DNA methyltransferases. Our
initial characterization of RsrI methyltransferase (M.RsrI) was consistent
with the enzyme functioning as a monomer, but, subsequently, the enzyme
crystallized as a dimer with 1500 A2 of buried surface area. This result
led us to re-examine the biochemical properties of M.RsrI. Gel-shift
studies of M.RsrI binding to DNA suggested that binding cooperativity
targets hemimethylated DNA preferentially over unmethylated DNA.
Size-exclusion chromatography indicated that the M.RsrI-DNA complex had a
size and stoichiometry consistent with a dimeric enzyme binding to the
DNA. Kinetic measurements revealed a quadratic relationship between enzyme
velocity and concentration. Site-directed mutagenesis at the dimer
interface affected the kinetics and DNA-binding of the enzyme, providing
support for a model proposing an active enzyme dimer. We also identified a
conserved motif in the dimer interfaces of the beta-class
methyltransferases M.RsrI, M.MboIIA and M2.DpnII. Taken together, these
data suggest that M.RsrI may be part of a sub-class of MTases that
function as dimers.

<>

<1>Thomas, C.B., Scavetta, R., Churchill, M.E.A., Gumport, R.I.
<2>A tale of three tails, a mutant, and a dimer: Structural studies of RsrI methyltransferase.
<3>FASEB J.
<4>16
<5>A923-A924
<6>2002
<7>DNA methyltransferases are a ubiquitous class of enzymes first observed as component enzymes
of restriction-modification systems and implicated in many biological phenomena.  Recent
reports have shown them to be involved in genetic diseases such as fragile X-syndrome,
X-chromosome inactivation, gene regulation, cancer, and immunity.  RsrI DNA methyltransferase
methylates the exocyclic amino group of the central adenine of the double-stranded DNA
sequence GAATTC.  Using X-ray crystallography, we report co-crystal structures of the enzyme
with the methyl donor S-adenosylmethionine, the product S-adenosylhomocysteine, and the
inhibitor sinefungin.  The co-crystal structures reveal two ligand sites of varying affinity
and possibly by lipid raft-mediated clustering on the plasma membrane.

<>

<1>Thomas, C.B., Scavetta, R.D., Gumport, R.I., Churchill, M.E.A.
<2>Structures of liganded and unliganded RsrI N6-adenine DNA methyltransferase: a distinct orientation for active cofactor binding.
<3>J. Biol. Chem.
<4>278
<5>26094-26101
<6>2003
<7>The structures of RsrI DNA methyltransferase (M.RsrI) bound to the substrate
S-adenosyl-l-methionine (AdoMet), the product
S-adenosyl-l-homocysteine (AdoHcy), the inhibitor sinefungin, as well as a
mutant apo-enzyme have been determined by x-ray crystallography. Two
distinct binding configurations were observed for the three ligands. The
substrate AdoMet adopts a bent shape that directs the activated methyl
group toward the active site near the catalytic DPPY motif. The product
AdoHcy and the competitive inhibitor sinefungin bind with a straight
conformation in which the amino acid moiety occupies a position near the
activated methyl group in the AdoMet complex. Analysis of ligand binding
in comparison with other DNA methyltransferases reveals a small, common
subset of available conformations for the ligand. The structures of M.RsrI
with the non-substrate ligands contained a bound chloride ion in the
AdoMet carboxylate-binding pocket, explaining its inhibition by chloride
salts. The L72P mutant of M.RsrI is the first DNA methyltransferase
structure without bound ligand. With respect to the wild-type protein, it
had a larger ligand-binding pocket and displayed movement of a loop
(223-227) that is responsible for binding the ligand, which may account
for the weaker affinity of the L72P mutant for AdoMet. These studies show
the subtle changes in the tight specific interactions of substrate,
product, and an inhibitor with M.RsrI and help explain how each displays
its unique effect on the activity of the enzyme.

<>

<1>Thomas, D.K., Lone, A.G., Selinger, L.B., Taboada, E.N., Uwiera, R.R., Abbott, D.W., Inglis, G.D.
<2>Comparative Variation within the Genome of Campylobacter jejuni NCTC 11168 in Human and Murine Hosts.
<3>PLoS ONE
<4>9
<5>E88229
<6>2014
<7>Campylobacteriosis incited by C. jejuni is a significant enteric disease of human
beings. A person working with two reference strains of C. jejuni National
Collection of Type Cultures (NCTC) 11168 developed symptoms of severe enteritis
including bloody diarrhea. The worker was determined to be infected by C. jejuni.
In excess of 50 isolates were recovered from the worker's stool. All of the
recovered isolates and the two reference strains were indistinguishable from each
other based on comparative genomic fingerprint subtyping. Whole genome sequence
analysis indicated that the worker was infected with a C. jejuni NCTC 11168
obtained from the American Type Culture Collection; this strain (NCTC 11168-GSv)
is the genome sequence reference. After passage through the human host, major
genetic changes including indel mutations within twelve contingency loci
conferring phase variations were detected in the genome of C. jejuni. Specific
and robust single nucleotide polymorphism (SNP) changes in the human host were
also observed in two loci (Cj0144c, Cj1564). In mice inoculated with an isolate
of C. jejuni NCTC 11168-GSv from the infected person, the isolate underwent
further genetic variation. At nine loci, mutations specific to inoculated mice
including five SNP changes were observed. The two predominant SNPs observed in
the human host reverted in mice. Genetic variations occurring in the genome of C.
jejuni in mice corresponded to increased densities of C. jejuni cells associated
with cecal mucosa. In conclusion, C. jejuni NCTC 11168-GSv was found to be highly
virulent in a human being inciting severe enteritis. Host-specific mutations in
the person with enteritis occurred/were selected for in the genome of C. jejuni,
and many were not maintained in mice. Information obtained in the current study
provides new information on host-specific genetic adaptation by C. jejuni.

<>

<1>Thomas, G.A., Kubasek, W.L., Peticolas, W.L.
<2>Environmentally induced conformational changes in B-type DNA:  Comparison of the conformation of the oligonucleotide d(TCGCGAATTCGCG) in solution and in its crystalline complex with the restriction nuclease EcoRI.
<3>Biochemistry
<4>28
<5>2001-2009
<6>1989
<7>Raman spectroscopic analysis of the secondary structure of the crystalline
restriction endonuclease EcoRI, the oligonucleotide d(TCGCGAATTCGCG) in
solution, and the corresponding crystalline EcoRI-oligonucleotide complex
reveals structural differences between the complexed and uncomplexed protein
and oligonucleotide components that appear to be linked to complex formation.
Structural differences that are spectroscopically identified include (1) an
increase in the population of furanose rings adopting the C3'-endo conformation
and (2) spectroscopically observed changes in base stacking which are probably
associated with the crystallographically observed distortion of the phosphate
backbone about positions C(3)-G(4) and C(9)-G(10) and unwinding between the
symmetry-related segments GAA-TTC which make up the central recognition core
(McClarin et al., 1986).  Changes in base stacking due to distortions and
unwinding along the oligonucleotide result in differences in the base
vibrational region between spectra of the complex and the oligonucleotide in
solution.  The spectroscopic analysis indicates that the C2'-endo population is
similar for the oligonucleotide in solution and in the complex.  The additional
C3'-endo population in the complex appears to arise from the conversion of
rings adopting alternative conformations such as C1'-exo and O1'-endo.
Analysis of the vibrational bands derived from guanine indicates that the
population of guanine residues associated with furanose rings in a C2'-endo
conformation is similar for the oligonucleotide in solution and in the
crystalline complex.  This implies that the increase in C3'-endo population is
not associated with guanine residues.  Large conformational distortions such as
those observed in the crystal are not observed for the oligonucleotide in
solution.  Furthermore, Raman evidence is presented that these distortions are
not observed in either the crystal or the solution of the oligomer
d(CGCGAATTCGCG).  These data suggest that static distortions such as those
observed in the crystal are not employed for initial sequence recognition.
They appear either as a secondary response to interaction with the protein or
as transient fluctuations which exist at a very low level in solution.

<>

<1>Thomas, J.C., Godfrey, P.A., Feldgarden, M., Robinson, D.A.
<2>Draft Genome Sequences of Staphylococcus aureus Sequence Type 34 (ST34) and ST42  Hybrids.
<3>J. Bacteriol.
<4>194
<5>2740-2741
<6>2012
<7>Staphylococcus aureus is a major cause of antimicrobial-resistant infections of humans.
Hybrids of S. aureus, which originate from large-scale chromosomal
recombinations between parents of distinct genetic backgrounds, are of interest
from clinical and evolutionary perspectives. We present draft genome sequences of
two S. aureus hybrids of sequence type 34 (ST34) and ST42.

<>

<1>Thomas, J.C., Kong, Y., Sabharwal, V., Pelton, S.I., Pettigrew, M.M.
<2>Streptococcus pneumoniae serotype 6C: An Intra- and Interclonal Complex Comparison.
<3>J. Bacteriol.
<4>193
<5>3409-3410
<6>2011
<7>We report the annotated draft genome sequences of four serotype 6C Streptococcus pneumoniae
isolates of differing genetic backgrounds.
Serotype 6C isolates are increasing in prevalence and becoming
progressively more resistant to antibiotics. As a result these strains are
likely to become more important in the near future.

<>

<1>Thomas, M., Davis, R.W.
<2>Studies on the cleavage of bacteriophage lambda DNA with EcoRI restriction endonuclease.
<3>J. Mol. Biol.
<4>91
<5>315-328
<6>1975
<7>The five EcoRI restriction sites in bacteriophage lambda DNA have been mapped
at 0.445, 0.543, 0.656, 0.810, and 0.931 fractional lengths from the left end
of the DNA molecule.  These positions were determined electron-microscopically
by single-site cleavage of hydrogen-bonded circular lambda DNA molecules and by
cleavage of various DNA heteroduplexes between lambda DNA and DNA from well
defined lambdamutants.  The DNA lengths of the EcoRI fragments are in agreement
with their electrophoretic mobility on agarose gels but are not in agreement
with their mobilities on polyacrylamide gels.  These positions are different
from those previously published by Allet et al. (1973).  Partial cleavage of
pure lambda DNA by addition of small amounts of EcoRI endonuclease does not
lead to random cleavage between molecules.  Also, the first site cleaved is not
randomly distributed among the five sites within a molecule.  The site nearest
the right end is cleaved first about ten times more frequently than either of
the two center sites.

<>

<1>Thomas, M., Rey, M.
<2>Methods of improving the introduction of DNA into bacterial cells.
<3>International Patent Office
<4>WO 2008067423 A
<5>
<6>2008
<7>The present invention relates to methods of improving the introduction of DNA into bacterial
host cells. The genome of B. licheniformis strain SJ1904 was sequenced using 454 technology
and genes for M.Bli1904II and Bli1904II were identified.

<>

<1>Thomas, M.P., Brady, R.L., Halford, S.E., Sessions, R.B., Baldwin, G.S.
<2>Structural analysis of a mutational hot-spot in the EcoRV restriction endonuclease: a catalytic role for a main chain carbonyl group.
<3>Nucleic Acids Res.
<4>27
<5>3438-3445
<6>1999
<7>Following random mutagenesis of the Eco RV endonuclease, a high
proportion of the null mutants carry substitutions at Gln69. Such
mutants display reduced rates for the DNA cleavage step in the reaction
pathway, yet the crystal structures of wild-type Eco RV fail to explain
why Gln69 is crucial for activity. In this study, crystal structures
were determined for two mutants of Eco RV, with Leu or Glu at residue
69, bound to specific DNA. The structures of the mutants are similar to
the native protein and no function can be ascribed to the side chain of
the amino acid at this locus. Instead, the structures of the mutant
proteins suggest that the catalytic defect is due to the positioning of
the main chain carbonyl group. In the enzyme-substrate complex for Eco
RV, the main chain carbonyl of Gln69 makes no interactions with
catalytic functions but, in the enzyme-product complex, it coordinates
a metal ion bound to the newly liberated 5'-phosphate. This re-
positioning may be hindered in the mutant proteins. Molecular dynamics
calculations indicate that the metal on the phosphoryl oxygen interacts
with the carbonyl group upon forming the pentavalent intermediate
during phosphodiester hydrolysis. A main chain carbonyl may thus play a
role in catalysis by Eco RV.

<>

<1>Thomm, M., Frey, G., Bolton, B.J., Laue, F., Kessler, C., Stetter, K.O.
<2>MvnI: a restriction enzyme in the archaebacterium Methanococcus vannielii.
<3>FEMS Microbiol. Lett.
<4>52
<5>229-234
<6>1988
<7>The methanogenic archaebacerium Methanococcus vannielii contains a type II
restriction endonuclease.  The enzyme was purified by a simple three-step
procedure resulting in enzyme preparations free of contaminating unspecific
nucleases.  The restriction enzyme recognizes and cleaves the sequence
5'-CG^CG-3' (FnuDII and ThaI isoschizomer) and generates DNA fragments with
blunt ends.  Due to its purity and activity at moderate temperatures, MvnI
might be a useful alternative to FnuDII and ThaI active at 60C.

<>

<1>Thompson, A.J., Herrin, D.L.
<2>In vitro self-splicing reactions of the chloroplast group I intron Cr.LSU from Chlamydomonas reinhardtii and in vivo manipulation via gene-replacement.
<3>Nucleic Acids Res.
<4>19
<5>6611-6618
<6>1991
<7>The group I intron from the chloroplast rRNA large subunit of Chlamydomonas reinhardtii
(Cr.LSU) undergoes autocatalytic splicing in vitro. Cr.LSU displays a range of reactions
typical of other group I introns. Under optimal conditions, the 5' cleavage step proceeds
rapidly, but the exon-ligation step is relatively slow, and no pH dependent hydrolysis of the
3' splicing site occurs. A requirement for high temperature and high [Mg2+] suggests
involvement of additional splicing factors in vivo. The positions of three cyclization sites
of the free intron have been mapped; two of these sites represent reactions analogous to
5'-splice site cleavage, whereas the third is an example of G-exchange. Cr.LSU contains an
open reading frame (ORF) potentially encoding a 163 amino acid polypeptide. ORF function has
been investigated by using chloroplast gene replacement via particle bombardment. We have
shown that the ORF can be deleted from Cr.LSU without affecting splicing in vivo and it thus
does not encode an essential splicing factor.

<>

<1>Thompson, A.J., Yuan, X., Kudlicki, W., Herrin, D.L.
<2>Cleavage and recognition pattern of a double-strand-specific endonuclease (I-CreI) endcoded by the chloroplast 23S rRNA intron of Chlamydomonas reinhardtii.
<3>Gene
<4>119
<5>247-251
<6>1992
<7>Several group-I introns have been shown to specifically invade intron-minus alleles of the
genes that contain them. This type of intron mobility is referred to as "intron homing" and
depends on restriction endonucleases (ENases) encoded by the mobile introns. The ENase cleaves
the intron-minus allele near the site of intron insertion, thereby initiating gene conversion.
The 23S (LSU) rRNA-encoding gene (LSU) of the chloroplast genome of Chlamydomonas reinhardtii
contains a self-splicing group-I intron (CrLSU) that has a free-standing open reading frame
(ORF) of 163 codons. Translation of CrLSU intron RNA in cell-free systems produces a
polypeptide of approximately 18 kDa, the size expected for correct translation of the ORF. The
in vitro-synthesized 18-kDa protein cleaves plasmid DNA that contains a portion of LSU where
the intron normally resides, but lacking the intron itself. Cleavage by the intron-encoded
enzyme (I-CreI) occurs 5 bp and 1 bp 3' to the intron insertion site (in the 3'-exon) in the
top (/) and bottom (,) strands, respectively, resulting in 4-nt single-stranded overhangs with
3'-OH termini. We also show that the recognition sequence of I-CreI spans the cleavage site
and is 24 bp in length (5'-CAAAACGTC,GTGA/GACAGTTTGGT).

<>

<1>Thompson, C.C., Marin, M.A., Dias, G.M., Dutilh, B.E., Edwards, R.A., Iida, T., Thompson, F.L., Vicente, A.C.
<2>Genome Sequence of the Human Pathogen Vibrio cholerae Amazonia.
<3>J. Bacteriol.
<4>193
<5>5877-5878
<6>2011
<7>Vibrio cholerae O1 Amazonia is a pathogen that was isolated from cholera-like diarrhea cases
in at least two countries, Brazil and Ghana.
Based on multilocus sequence analysis, this lineage belongs to a distinct
profile compared to strains from El Tor and classical biotypes. The
genomic analysis revealed that it contains Vibrio pathogenicity island 2
and a set of genes related to pathogenesis and fitness, such as the type
VI secretion system, present in choleragenic V. cholerae strains.

<>

<1>Thompson, R., Hughes, S.G., Broda, P.
<2>Plasmid identification using specific endonucleases.
<3>Mol. Gen. Genet.
<4>133
<5>141-149
<6>1974
<7>Digestion with specific endonucleases followed by agarose gel electrophoresis
yields characteristic fragment patterns from different plasmid DNAs.  Since
even the very closely related R factors F100-1 and R6, for example, can be
distinguished this method provides a powerful test of the identity of two
plasmid DNAs.

<>

<1>Thompson, S.A., Hall, J.E., Baltzegar, A.D., Yates, B.D., Maani, E.V., Pajaniappan, M., Falkow, S., Gaynor, E.C., Mishra, A.K., Burns, C.M.
<2>Mutation of a predicted Campylobacter jejuni DNA methylase affects the expression of multiple genes.
<3>Int. J. Med. Microbiol.
<4>293
<5>21-22
<6>2003
<7>In the course of studies examining temperature regulated genes of Campylobacter jejuni 81-176,
we identified an ortholog of NCTC11168 CJ1461 as a gene that is more highly expressed at 37oC
than at 42oC.  CJ1461 encodes a predicted DNA methylase that is not associated with a cognate
restriction endonuclease, implying that its cellular role is not to protect DNA from
restriction endonucleases.  Since DNA methylases may have gene regulatory functions separate
from their housekeeping roles in protecting DNA from restriction enzymes or in DNA repair, we
inactivated the CJ1461 ortholog in strain 81-176, suggesting a regulatory role for CJ1461.
Some of these proteins were shown to be temperature regulated by other methods, suggesting
that a component of the putative CJ1461 regulon was mediated by growth temperature.
Importantly, one of the proteins that was over expressed in the CJ1461 mutant was CJ0355.
CJ0355 itself is the predicted response regulator of a two component regulatory system,
although CJ0355 is an "orphan" response regulator that lacks an associated histidine kinase
sensor protein.  These studies suggest that temperature regulation of multiple genes in C.
jejuni 81-176 may be complex and can be mediated in part by DNA methylation and by an orphan
response regulator.

<>

<1>Thoms, B., Wackernagel, W.
<2>UV-induced allevation of lambda restriction in Escherichia coli K-12:  Kinetics of induction and specificity of this SOS function.
<3>Mol. Gen. Genet.
<4>186
<5>111-117
<6>1982
<7>In UV-irradiated cells of Escherichia coli K-12 a partial release of the
restriction of non-modified phage lambda is observed when the cells are recA+
lexA+.  We show here that the induction of this restriction allevation (RA)
also depends on the recBC enzyme and that the expression of RA requires protein
synthesis.  Maximum expression was reached within 60 to 90 min after
irradiation.  Experiments are presented which show that upon UV-irradiation a
signal is created which triggers the development of RA when protein synthesis
is allowed.  This signal decayed with a half-life of only a few minutes in
cells treated with chloramphenicol.  The decay kinetics were similar in uvr+
and uvrA mutants.  RA appeared to be specific for EcoKI insofar as no
allevation of lambda restriction by EcoRI, EcoRII and EcoPI occurred.  During
maximum expression of RA no gross reduction of the activities of the recBC
enzyme (exonuclease V) and the restriction endonuclease EcoKI was observed and
no new DNA modifying activity appeared in the cells.  Since, in fully expressed
cells, up to 75% of the infecting lambda DNA was converted to acid-soluble
material within 20 min after infection we suggest that only a small specific
fraction of lambda infections may undergo RA.

<>

<1>Thoms, B., Wackernagel, W.
<2>Expression of ultraviolet-induced restriction alleviation in Escherichia coli K-12.
<3>Biochim. Biophys. Acta
<4>739
<5>42-47
<6>1983
<7>Ultraviolet-induced restriction alleviation is an SOS function which partially relieves the
K-12-specific DNA restriction in Escherichia coli.  Restriction alleviation is determined by
observing elevated survival of unmodified phage lambda in cells irradiated with ultraviolet
prior to infection.  We demonstrate that restriction of lambda is also relieved when log-phase
cells are irradiated as late as 50 min after adsorption of lambda.  At this time more than 60%
of the lambda DNA is already released as acid-soluble material from the cells.  Experiments
involving reextraction of lambda DNA from infected cells and a mild detergent treatment
removing adsorbed phages from the cellular surface showed that only a small specific fraction
of all lambda infections is destined to escape restriction due to restriction alleviation.
This fraction (10-20%) has a retarded mode of DNA injection (60 min or longer) after
adsorption which allows the expression of the restriction alleviation function before the
phage DNA is exposed to restriction endonucleases.  This behavior of a fraction of lambda
phages explains why the SOS function restriction alleviation could initially be discovered.
We show that the retarded mode of DNA injection is not required for another SOS function
acting on lambda DNA, the increased repair of ultraviolet-irradiated DNA (Weigle
reactivation).

<>

<1>Thomson, N.R. et al.
<2>The Complete Genome Sequence and Comparative Genome Analysis of the High Pathogenicity Yersinia enterocolitica Strain 8081.
<3>PLoS Genet.
<4>2
<5>E206
<6>2006
<7>The human enteropathogen, Yersinia enterocolitica, is a significant link
in the range of Yersinia pathologies extending from mild gastroenteritis
to bubonic plague. Comparison at the genomic level is a key step in our
understanding of the genetic basis for this pathogenicity spectrum. Here
we report the genome of Y. enterocolitica strain 8081 (serotype 0:8;
biotype 1B) and extensive microarray data relating to the genetic
diversity of the Y. enterocolitica species. Our analysis reveals that the
genome of Y. enterocolitica strain 8081 is a patchwork of horizontally
acquired genetic loci, including a plasticity zone of 199 kb containing an
extraordinarily high density of virulence genes. Microarray analysis has
provided insights into species-specific Y. enterocolitica gene functions
and the intraspecies differences between the high, low, and nonpathogenic
Y. enterocolitica biotypes. Through comparative genome sequence analysis
we provide new information on the evolution of the Yersinia. We identify
numerous loci that represent ancestral clusters of genes potentially
important in enteric survival and pathogenesis, which have been lost or
are in the process of being lost, in the other sequenced Yersinia
lineages. Our analysis also highlights large metabolic operons in Y.
enterocolitica that are absent in the related enteropathogen, Yersinia
pseudotuberculosis, indicating major differences in niche and nutrients
used within the mammalian gut. These include clusters directing, the
production of hydrogenases, tetrathionate respiration, cobalamin
synthesis, and propanediol utilisation. Along with ancestral gene
clusters, the genome of Y. enterocolitica has revealed species-specific
and enteropathogen-specific loci. This has provided important insights
into the pathology of this bacterium and, more broadly, into the evolution
of the genus. Moreover, wider investigations looking at the patterns of
gene loss and gain in the Yersinia have highlighted common themes in the
genome evolution of other human enteropathogens.

<>

<1>Thomson, N.R. et al.
<2>Comparative genome analysis of Salmonella Enteritidis PT4 and Salmonella Gallinarum 287/91 provides insights into evolutionary and host adaptation pathways.
<3>Genome Res.
<4>18
<5>1624-1637
<6>2008
<7>We have determined the complete genome sequences of a host-promiscuous Salmonella enterica
serovar Enteritidis PT4 isolate P125109 and a chicken-restricted Salmonella enterica serovar
Gallinarum isolate 287/91. Genome comparisons between these and other Salmonella isolates
indicate that S. Gallinarum 287/91 is a recently evolved descendent of S. Enteritidis.
Significantly, the genome of S. Gallinarum has undergone extensive degradation through
deletion and pseudogene formation. Comparison of the pseudogenes in S. Gallinarum with those
identified previously in other host-adapted bacteria reveals the loss of many common
functional traits and provides insights into possible mechanisms of host and tissue
adaptation. We propose that experimental analysis in chickens and mice of S.
Enteritidis-harboring mutations in functional homologs of the pseudogenes present in S.
Gallinarum could provide an experimentally tractable route toward unraveling the genetic basis
of host adaptation in S. enterica.

<>

<1>Thorogood, H., Grasby, J.A., Connolly, B.A.
<2>Influence of the phosphate backbone on the restriction and hydrolysis of DNA by the EcoRV restriction endonuclease.
<3>J. Biol. Chem.
<4>271
<5>8855-8862
<6>1996
<7>A set of phosphorothioate-containing oligonucleotides based on pGACGATATCGTC, a
self-complementary dodecamer that contains the EcoRV recognition sequence (GATATC), has been
prepared.  The phosphorothioate group has been individually introduced at the central nine
phosphate positions and the two diastereomers produced at each site separated and purified.
The Km and Vmax values found for each of these modified DNA molecules with the EcoRV
restriction endonuclease have been determined and compared with those seen for the unmodified
all-phosphate-containing dodecamer.  This has enabled an evaluation of the roles that both of
the non-esterified oxygen atoms in the individual phosphates play in DNA binding and
hydrolysis by the endonuclease.  The results have also been compared with crystal structures
of the EcoRV endonuclease, complexed with an oligodeoxynucleotide, to allow further definition
of phosphate group function during substrate binding and turnover.

<>

<1>Thorogood, H., Waters, T.R., Parker, A.W., Wharton, C., Connolly, B.A.
<2>Resonance raman spectroscopy of 4-thiothymidine and oligodeoxynucleotides containing this base both free in solution and bound to the restriction endonuclease EcoRV.
<3>Biochemistry
<4>35
<5>8723-8733
<6>1996
<7>The resonance Raman spectra of 4-thiothymidine [4ST] have been recorded (a) in
the free deoxynucleoside form, (b) when incorporated into the single stranded
oligodeoxynucleotide d(AG[4ST]-TC), and (c) within the double-stranded self-complementary
dodecamer d(GACGA[4sT]ATCGTC).  Vibrational mode assignments of almost all the major
Raman bands observed in each spectra have been made, mainly by comparison with the published
assignments of related nucleosides and nucleotides.  Differences between the spectra were
observed, particularly when [4sT] and d(AG[4ST]TC) were compared to
d(GACGA[4ST]ATCGTC).  This is explained in terms of the variations in structure between
single- and double-stranded DNA.  Good quality spectra were obtained at
nucleotide/oligonucleotide concentrations of between 100 and 500 uM and this coupled with an
apparatus that uses small volumes (100 uL) allowed measurement of the spectrum of
d(GACGA[4ST]ATCGTC) bound to the EcoRV endonuclease.  This well characterized nuclease,
for which crystal structures are available, recognizes d(GATAT) sequences.  When this is
replaced
with d(GA[4ST]ATC), a poor substrate results, but turnover can be prevented during data
accumulation by omission of the essential cation Mg2+.  Large shifts in several of the Raman
bands were observed, and these have been related to the environment of the [4ST] base in the
protein-bound oligonucleotide as deduced from the crystal structure.  The wavenumber for the
C=S stretch vibration in free d(GACGA[4ST]ATCGTC) has been used to calculate the strength of
the Watson-Crick hydrogen bond between the sulphur atom in [4ST] and the 6-NH2 group on its
partner dA.  On binding to the enzyme, the shift in the wavenumber of the C=S stretch
indicates
this Watson-Crick hydrogen bond is weakened, in good agreement with X-ray structures.  The
advantage of using [4ST] as a resonance Raman probe is that it absorbs at 340 nM, a wavelength
where other nucleic acid and protein absorbance is minimal.  Thus the spectra obtained are
very
simple and consist of signals that arise predominantly from the thiobase alone, and this
facilitates
data interpretation.

<>

<1>Thorpe, P.H.
<2>The DNA specificity of type I restriction and modification enzymes.
<3>Ph.D. Thesis, University of Edinburgh, , Edinburgh
<4>
<5>1-176
<6>1995
<7>The type I restriction and modification enzymes were the first R-M systems characterized and
provide a valuable system for studying protein-protein and protein-DNA interactions.  These
complex, multi-subunit enzymes are encoded for by three genes hsdR, M and S, although only one
of these, hsdS, is responsible for determining the specific DNA target sequence that the
enzyme recognizes.  Differences in the type I enzymes characterized have lead to their
subdivision into separate families.  The type I enzymes recognize a bipartite DNA target,
which has two, short, defined elements separated by a non-specific spacer.  For example, the
DNA target of EcoKI is 5'AACNNNNNNGTGC3'.  A series of observations and experiments have
lead to a limited understanding of how the HsdS subunit recognizes its DNA target.  Within
each family of type I enzymes HsdS subunits share regions of very similar amino acid sequence,
separating two larger regions of variable sequence.  Each variable region forms a domain that
specifies one half of the bipartite DNA target, and has been termed a Target Recognition
Domains (or TRD).  No structural information is available for the TRDs although recent crystal
structures of type II enzymes may give clues relevant to DNA interactions of type I enzymes.
A deletion analysis of the hsdS gene of EcoKI was initiated to provide information on the
roles of the conserved and variable regions.  The aim was to correlate phenotype with an
analysis of protein products, but attempts to overexpress the hsdS gene of EcoKI in a soluble
form were unsuccessful, and the HsdS subunit could not be purified.  Another approach to
studying the interaction of TRDs with their DNA targets is to compare the amino acid sequences
of those TRDs that specify the same DNA target.  Areas of sequence which are similar within
these TRDs may reflect a similarity of DNA recognition function.  From amongst the limited
number of TRDs available, there are several sequence alignments of TRDs which specify the same
DNA target.  A method based upon the Polymerase Chain Reaction was developed to amplify new
variable regions, which encode TRDs, from wild-type bacteria.  In a DNA hybridization screen
of members of the ECOR collection of wild-type Escherichia coli, Barcus et al. (1995) found
that almost half contained hsd genes.  The conserved regions of the hsd genes were used to
design primers that would amplify 5' variable regions of members of a given type I family.
Nine IA family and four IB family 5' variable regions were amplified and their DNA sequences
determined.  The information derived from these sequences illustrates both the evolutionary
diversity of the hsdS genes, and the flexible nature of the TRDs as independent target
recognizing domains.  The sequence of the N-terminal TRD from ECOR17 shares 28% identity with
those of EcoKI and StySPI.  A method dependent on the construction of hybrid hsdS genes, was
devised to find the DNA targets of these new TRD sequences.  Hybrid type I hsdS genes have
been shown to be functional for members of the IA and IC families.  Deletion derivatives of
the IA and IB family hsdS genes lacking the coding sequence for the amino-TRD were used to
insert coding sequences for alternative TRDs.  The four hybrid genes isolated all encoded
functional HsdS polypeptides conferring novel specificities.  The DNA targets of two hybrid
systems were determined, including that from ECOR17.  The sequence recognized was 5' ATR, and
not that expected (5' AAC) from its similarity to EcoKI.

<>

<1>Thorpe, P.H., Ternent, D., Murray, N.E.
<2>The specificity of StySKI, a type I restriction enzyme, implies a structure with rotational symmetry.
<3>Nucleic Acids Res.
<4>25
<5>1694-1700
<6>1997
<7>The type I restriction and modification (R-M) enzyme from Salmonella enterica serovar kaduna
(StySKI) recognizes the DNA sequence 5'-CGAT(N)7GTTA, an unusual target for a type I R-M
system in that it comprises two tetranucleotide components.  The amino target recognition
domain (TRD) of StySKI recognizes 5'-CGAT and shows 35% amino acid identity with the carboxy
TRD of EcoR124I which recognizes the complementary, but degenerate, sequence 5'-RTCG.
Current models predict that the amino and carboxy TRDs of the specificity subunit are in
inverted orientations within a structure with 2-fold rotational symmetry.  The complementary
target sequences recognized by the amino TRD of StySKI and the carboxy TRD of EcoR124I are
consistent with the predicted inverted positions of the TRDs.  Amino TRDs of similar amino
acid sequence have been shown to recognize the same nucleotide sequence.  The similarity
reported here, the first example of one between amino and carboxy TRDs, while consistent with
a conserved mechanism of target recognition, offers additional flexibility in the evolution of
sequence specificity by increasing the potential diversity of DNA targets for a given number
of TRDs.  StySKI identifies the first member of the IB family in Salmonella species.

<>

<1>Thorsted, P.B., Macartney, D.P., Akhtar, P., Haines, A.S., Ali, N., Davidson, P., Stafford, T., Pocklington, M.J., Pansegrau, W., Wilkins, B.M., Lanka, E., Thomas, C.M.
<2>Complete sequence of the IncPbeta plasmid R751: implications for evolution and organisation of the IncP backbone.
<3>J. Mol. Biol.
<4>282
<5>969-990
<6>1998
<7>The broad host range IncP plasmids are of particular interest because of
their ability to promote gene spread between diverse bacterial species. To
facilitate study of these plasmids we have compiled the complete sequence
of the IncPbeta plasmid R751. Comparison with the sequence of the
IncPalpha plasmids confirms the conservation of the IncP backbone of
replication, conjugative transfer and stable inheritance functions between
the two branches of this family. As in the IncPalpha genome the DNA of
this backbone appears to have been enriched for the GCCG/CGGC motifs
characteristic of the genome of organisms with a high G+C content, such as
P. aeruginosa, suggesting that IncPbeta plasmids have been subjected
during their evolution to similar mutational and selective forces as
IncPalpha plasmids and may have evolved in pseudomonad hosts. The IncP
genome is consistently interrupted by insertion of phenotypic markers
and/or transposable elements between oriV and trfA and between the tra and
trb operons. The R751 genome reveals a family of repeated sequences in
these regions which may form the basis of a hot spot for insertion of
foreign DNA. Sequence analysis of the cryptic transposon Tn4321 revealed
that it is not a member of the Tn21 family as we had proposed previously
from an inspection of its ends. Rather it is a composite transposon
defined by inverted repeats of a 1347 bp IS element belonging to a
recently discovered family which is distributed throughout the
prokaryotes. The central unique region of Tn4321 encodes two predicted
proteins, one of which is a regulatory protein while the other is
presumably responsible for an as yet unidentified phenotype. The most
striking feature of the IncPalpha plasmids, the global regulation of
replication and transfer by the KorA and KorB proteins encoded in the
central control operon, is conserved between the two plasmids although
there appear to be significant differences in the specificity of
repressor-operator interactions. The importance of these global regulatory
circuits is emphasised by the observation that the operator sequences for
KorB are highly conserved even in contexts where the surrounding region,
either a protein coding or intergenic sequence, has diverged considerably.
There appears to be no equivalent of the parABCDE region which in the
IncPalpha plasmids provides multimer resolution, lethality to plasmid-free
segregants and active partitioning functions. However, we found that the
continuous sector from co-ordinate 0 to 9100 bp, encoding the co-regulated
klc and kle operons as well as the central control region, could confer a
high degree of segregational stability on a low copy number test vector.
Thus R751 appears to exhibit very clearly what was first revealed by study
of the IncPalpha plasmids, namely a fully functional co-ordinately
regulated set of replication, transfer and stable inheritance functions.

<>

<1>Thrash, J.C., Cho, J.C., Bertagnolli, A.D., Ferriera, S., Johnson, J., Vergin, K.L., Giovannoni, S.J.
<2>Genome sequence of the marine Janibacter sp. strain HTCC2649.
<3>J. Bacteriol.
<4>193
<5>584-585
<6>2010
<7>Janibacter sp. strain HTCC2649 is a novel marine member of the Actinobacteria, family
Intrasporangiaceae, closely related to Janibacter melonis CM2104(T) and Knoellia sinensis HKI
0119(T). The organism was isolated from a sample collected at Hydrostation S south of Bermuda
using high throughput culturing techniques. Here we present the genome sequence of Janibacter
sp. strain HTCC2649.

<>

<1>Thrash, J.C., Cho, J.C., Ferriera, S., Johnson, J., Vergin, K.L., Giovannoni, S.J.
<2>Genome sequences of Pelagibaca bermudensis HTCC2601T and Maritimibacter alkaliphilus HTCC2654T, the type strains of two marine roseobacter genera.
<3>J. Bacteriol.
<4>192
<5>5552-5553
<6>2010
<7>Pelagibaca bermudensis HTCC2601(T) and Maritimibacter alkaliphilus HTCC2654(T) represent two
marine genera in the globally significant
Roseobacter clade of the Alphaproteobacteria. Here, we present the genome
sequences of these organisms, isolated from the Sargasso Sea using
dilution-to-extinction culturing, which offer insight into the genetic
basis for the metabolic and ecological diversity of this important group.

<>

<1>Thrash, J.C., Cho, J.C., Ferriera, S., Johnson, J., Vergin, K.L., Giovannoni, S.J.
<2>Genome sequences of strains HTCC2148 and HTCC2080, belonging to the OM60/NOR5 clade of the Gammaproteobacteria.
<3>J. Bacteriol.
<4>192
<5>3842-3843
<6>2010
<7>Organisms in the OM60/NOR5 clade of the Gammaproteobacteria are ubiquitous in the
world's oceans and can make up as much as 11% of bacterial cells in certain
areas. Isolated from coastal Oregon water, Gammaproteobacteria HTCC2148 and
HTCC2080 are two members of this important clade. Here we present the genome
sequences of the OM60 Gammaproteobacteria HTCC2148 and HTCC2080.

<>

<1>Thrash, J.C., Cho, J.C., Vergin, K.L., Giovannoni, S.J.
<2>Genome Sequences of Oceanicola granulosus HTCC2516T and Oceanicola batsensis HTCC2597T.
<3>J. Bacteriol.
<4>192
<5>3549-3550
<6>2010
<7>Genome sequences from the prolific Roseobacter clade in the Alphaproteobacteria are beginning
to reveal the genetic basis for the
diverse lifestyles of these organisms. Here we present the genome
sequences of Oceanicola granulosus HTCC2516(T) and Oceanicola batsensis
HTCC2597(T), two marine Roseobacter species isolated from the Sargasso Sea
using dilution-to-extinction culturing, whose genomes encode for
significant differences in metabolic potential.

<>

<1>Thrash, J.C., Cho, J.C., Vergin, K.L., Morris, R.M., Giovannoni, S.J.
<2>Genome sequence of Lentisphaera araneosa HTCC2155T, the type species of the order Lentisphaerales in the phylum Lentisphaerae.
<3>J. Bacteriol.
<4>192
<5>2938-2939
<6>2010
<7>Information on the genome content of deeply branching phyla with very few cultured members is
invaluable for expanding understanding of microbial evolution. Lentisphaera araneosa
HTCC2155(T) was isolated from the Oregon coast using dilution-to-extinction culturing. It is a
marine heterotroph found in surface and mesopelagic waters in both the Pacific and Atlantic
oceans and has the unusual property of producing a net-like matrix of secreted
exopolysaccharide. Here we present the genome sequence of L.
araneosa HTCC2155(T), importantly, one of only two sequenced members of
the phylum Lentisphaerae.

<>

<1>Thyme, S.B., Baker, D., Bradley, P.
<2>Improved Modeling of Side-Chain-Base Interactions and Plasticity in Protein-DNA Interface Design.
<3>J. Mol. Biol.
<4>419
<5>255-274
<6>2012
<7>Combinatorial sequence optimization for protein design requires libraries of discrete
side-chain conformations. The discreteness of
these libraries is problematic, particularly for long, polar side
chains, since favorable interactions can be missed. Previously, an
approach to loop remodeling where protein backbone movement is directed
by side-chain rotamers predicted to form interactions previously
observed in native complexes (termed 'motifs') was described. Here, we
show how such motif libraries can be incorporated into combinatorial
sequence optimization protocols and improve native complex
recapitulation. Guided by the motif rotamer searches, we made
improvements to the underlying energy function, increasing
recapitulation of native interactions. To further test the methods, we
carried out a comprehensive experimental scan of amino acid preferences
in the I-AniI protein-DNA interface and found that many positions
tolerated multiple amino acids. This sequence plasticity is not
observed in the computational results because of the fixed-backbone
approximation of the model. We improved modeling of this diversity by
introducing DNA flexibility and reducing the convergence of the
simulated annealing algorithm that drives the design process. In
addition to serving as a benchmark, this extensive experimental data
set provides insight into the types of interactions essential to
maintain the function of this potential gene therapy reagent.

<>

<1>Thyme, S.B., Boissel, S.J.S., Quadri, S., Arshiya, N., Tony, B., Dean, A., Park, R.U., Kusak, L., Ashworth, J., Baker, D.
<2>Reprogramming homing endonuclease specificity through computational design and directed evolution.
<3>Nucleic Acids Res.
<4>42
<5>2564-2576
<6>2014
<7>Homing endonucleases (HEs) can be used to induce targeted genome modification to reduce the
fitness of pathogen vectors such as the malaria-transmitting Anopheles gambiae and to correct
deleterious mutations in genetic diseases. We describe the creation of an extensive set of HE
variants with novel DNA cleavage specificities using an integrated experimental and
computational approach. Using computational modeling and an improved selection strategy, which
optimizes specificity in addition to activity, we engineered an endonuclease to cleave in a
gene associated with Anopheles sterility and another to cleave near a mutation that causes
pyruvate kinase deficiency. In the course of this work we observed unanticipated
context-dependence between bases which will need to be mechanistically understood for
reprogramming of specificity to succeed more generally.

<>

<1>Thyme, S.B., Jarjour, J., Takeuchi, R., Havranek, J.J., Ashworth, J., Scharenberg, A.M., Stoddard, B.L., Baker, D.
<2>Exploitation of binding energy for catalysis and design.
<3>Nature
<4>461
<5>1300-1304
<6>2009
<7>Enzymes use substrate-binding energy both to promote ground-state association and to stabilize
the reaction transition state
selectively1. The monomeric homing endonuclease I-AniI cleaves with
high sequence specificity in the centre of a 20-base-pair ( bp) DNA
target site, with the amino (N)-terminal domain of the enzyme making
extensive binding interactions with the left (-) side of the target
site and the similarly structured carboxy (C)-terminal domain
interacting with the right (+) side(2). Here we show that, despite the
approximate twofold symmetry of the enzyme-DNA complex, there is almost
complete segregation of interactions responsible for substrate binding
to the (-) side of the interface and interactions responsible for
transition-state stabilization to the (+) side. Although single
base-pair substitutions throughout the entire DNA target site reduce
catalytic efficiency, mutations in the (-) DNA half-site almost
exclusively increase the dissociation constant (K-D) and the Michaelis
constant under single-turnover conditions (K-M*), and those in the (+)
half-site primarily decrease the turnover number (k(cat)*). The
reduction of activity produced by mutations on the (-) side, but not
mutations on the (+) side, can be suppressed by tethering the substrate
to the endonuclease displayed on the surface of yeast. This dramatic
asymmetry in the use of enzyme-substrate binding energy for catalysis
has direct relevance to the redesign of endonucleases to cleave genomic
target sites for gene therapy and other applications. Computationally
redesigned enzymes that achieve new specificities on the (-) side do so
by modulating K-M*, whereas redesigns with altered specificities on the
(+) side modulate k(cat)*. Our results illustrate how classical
enzymology and modern protein design can each inform the other.

<>

<1>Thyme, S.B., Song, Y., Brunette, T.J., Szeto, M.D., Kusak, L., Bradley, P., Baker, D.
<2>Massively parallel determination and modeling of endonuclease substrate specificity.
<3>Nucleic Acids Res.
<4>42
<5>13839-13852
<6>2014
<7>We describe the identification and characterization of novel homing endonucleases using genome
database mining to identify putative target sites, followed by high throughput activity
screening in a bacterial selection system. We characterized the substrate specificity and
kinetics of these endonucleases by monitoring DNA cleavage events with deep sequencing. The
endonuclease specificities revealed by these experiments can be partially recapitulated using
3D structure-based computational models. Analysis of these models together with genome
sequence data provide insights into how alternative endonuclease specificities were generated
during natural evolution.

<>

<1>Tiago, I., Maranha, A., Mendes, V., Alarico, S., Moynihan, P.J., Clarke, A.J., Macedo-Ribeiro, S., Pereira, P.J., Empadinhas, N.
<2>Genome Sequence of Mycobacterium hassiacum DSM 44199, a Rare Source of Heat-Stable Mycobacterial Proteins.
<3>J. Bacteriol.
<4>194
<5>7010-7011
<6>2012
<7>Mycobacterium hassiacum is a rapidly growing mycobacterium isolated from human urine and so
far the most thermophilic among mycobacterial species. Its
thermotolerance and phylogenetic relationship to M. tuberculosis render its
proteins attractive tools for crystallization and structure-guided drug design.
We report the draft genome sequence of M. hassiacum DSM 44199.

<>

<1>Tian, G.-L., Michel, F., Macadre, C., Slonimski, P.P., Lazowska, J.
<2>Incipient mitochondrial evolution in yeasts.  II.  The complete sequence of the gene coding for cytochrome b in S. douglasii reveals the presence of both new and conserved introns and discloses major differences in the fixation of mutations in evolution.
<3>J. Mol. Biol.
<4>218
<5>747-760
<6>1991
<7>We have determined the complete sequence of the mitochondrial gene coding for cytochrome b in
Saccharomyces douglasii.  The gene is 6310 base-pairs long and is interrupted by four introns.
The first one (1311 base-pairs) belongs to the group ID of secondary structure, contains a
fragment open reading frame with a characteristic GIY . . . YIG motif, is absent from
Saccharomyces cerevisiae and is inserted in the same site in which introns 1 and 2 are
inserted in Neurospora crassa and Podospora anserina, respectively.  The next three S.
douglasii introns are homologous to the first three introns of S. cerevisiae, are inserted at
the same positions and display various degrees of similarity ranging from an almost complete
identity (intron 2 and 4) to a moderate one (intron 3).  We have compared secondary structures
of intron RNAs, and nucleotide and amino acid sequences of cytochrome b exons and intron open
reading grames in the two Saccharomyces species.  The rules that govern fixation of mutations
in exon and intron open reading frames are different:  the relative proportion of mutations
occurring in synonymous codons is low in some introns and high in exons.  The overall
frequency of mutations in cytochrome b exons is much smaller than in nuclear genes of yeasts,
contrary to what has been found in vertebrates, where mitochrondrial mutations are more
frequent.  The divergence of the cytochrome b gene is modular: various parts of the gene have
changed with a different mode and tempo of evolution.

<>

<1>Tian, H., Jing, C.
<2>Genome Sequence of the Aerobic Arsenate-Reducing Bacterium Pantoea sp. Strain IMH.
<3>Genome Announcements
<4>2
<5>e00267-14
<6>2014
<7>We here report the draft assembly for the genome of Pantoea sp. strain IMH, isolated from
arsenic-contaminated soil in Inner Mongolia, China, with the ability to aerobically reduce
arsenate to arsenite. The genome sequence will allow for the characterization of the molecular
mechanisms of arsenate reduction.

<>

<1>Tian, J., Xu, L., Zhang, S., Sun, W., Chu, X., Wu, N.
<2>Draft Genome Sequence of the Organophosphorus-Degrading Bacterium Pseudomonas sp. Strain 1-7, Isolated from Organophosphorus-Polluted Sludge.
<3>Genome Announcements
<4>2
<5>e00993-14
<6>2014
<7>Pseudomonas sp. strain 1-7, isolated from organophosphorus-polluted sludge, is able to degrade
many organophosphorus compounds. Here, we report the draft genome
sequence of Pseudomonas sp. strain 1-7.

<>

<1>Tian, S.-M., Gong, Z.-Z., Xie, W.-J., Zheng, L., Ruan, K.-C.
<2>Effect of high hydrostatic pressure on activity of restriction endonucleases.
<3>Acta Biochim. Biophys. Sin.
<4>31
<5>523-526
<6>1999
<7>The effect of high hydrostatic pressure on the activities of type II restriction enzymes
HindIII and XbaI in digesting plasmid pSPORT1 was studied.  The endonuclease activity of
HindIII and XbaI at 37 C were gradually inhibited by increasing pressure and completely
inhibited at 200 and 180 MPa, respectively.  No obvious irreversible effect was observed for
HindIII after suffering high pressure, while a considerable irreversible inactivation was
observed for XbaI.  The standard molar volume changes for HindIII and XbaI estimated from the
inhibition of endonuclease activity at different pressures were 213 and 103 ml/mol,
respectively.  It was also concluded that pressurization did not change the substrate sequence
specificity of both HindIII and XbaI.

<>

<1>Tice, H. et al.
<2>Complete genome sequence of Nakamurella multipartita type strain (Y-104).
<3>Standards in Genomic Sciences
<4>2
<5>168-175
<6>2010
<7>Nakamurella multipartita (Yoshimi et al. 1996) Tao et al. 2004 is the type species of the
monospecific genus Nakamurella in the actinobacterial suborder
Frankineae. The nonmotile, coccus-shaped strain was isolated from activated
sludge acclimated with sugar-containing synthetic wastewater, and is capable of
accumulating large amounts of polysaccharides in its cells. Here we describe the
features of the organism, together with the complete genome sequence and
annotation. This is the first complete genome sequence of a member of the family
Nakamurellaceae. The 6,060,298 bp long single replicon genome with its 5415
protein-coding and 56 RNA genes is part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Tidona, C.A., Schnitzler, P., Kehm, R., Darai, G.
<2>Identification of the gene encoding the DNA (cytosine-5) methyltransferase of lymphocystis disease virus.
<3>Virus Genes
<4>12
<5>219-229
<6>1996
<7>The gene encoding the DNA (cytosine-5) methyltransferase (m5C-MTase) of lymphocystis disease
virus (flounder isolate, LCDV-1) has been identified by polymerase chain reaction (PCR) using
oligonucleotide primers synthesized corresponding to different regions of the m5C-MTase gene
of frog virus 3 (FV3).  A DNA fragment of 487 bp was amplified using oligonucleotide primers
L3 and R4 which correspond to the nucleotide positions 87 to 109 and 530 to 550 of the
m5C-MTase gene of FV3, respectively.  The DNA nucleotide sequence of the PCR product was
determined by direct cycle sequencing.  The alignment of the deduced amino acid sequence
derived from the PCR product and the m5C-MTase protein of FV3 revealed a homology of 55.4%
identity and 29.1% similarity.  The amino acid sequence which was found to be significantly
homologous to the amino acid sequence deduced from the nucleotide sequence of the PCR product
was located at the amino acid position 37 to 175 of the m5C-MTase of FV3 indicating the
specificity of the amplified PCR product.  The DNA nucleotide sequence of the LCDV-1 genome
corresponding to the 5' and 3' termini of the m5C-MTase gene was determined by primer
walking.  The locus of the m5C-MTase gene of LCDV-1 was identified within the EcoRI DNA
fragment G of LCDV-1 (7.9 kbp; 0.947 to 0.034 map units).  The m5C-MTase gene of LCDV-1
comprises 684 nucleotides coding for a putative protein of 228 amino acid residues.  A high
degree of amino acid sequence homology (53.3% identity and 25.8% similarity) was detected
between the m5C-MTases of LCDV01 and FV3.

<>

<1>Tielen, P., Wibberg, D., Blom, J., Rosin, N., Meyer, A.K., Bunk, B., Schobert, M., Tupker, R., Schatschneider, S., Ruckert, C., Albersmeier, A., Goesmann, A., Vorholter, F.J., Jahn, D., Puhler, A.
<2>Genome Sequence of the Small-Colony Variant Pseudomonas aeruginosa MH27, Isolated from a Chronic Urethral Catheter Infection.
<3>Genome Announcements
<4>2
<5>e01174-13
<6>2014
<7>Pseudomonas aeruginosa is a notable nosocomial pathogen causing severe chronic infections.
Here we present the draft genome sequence of P. aeruginosa MH27,
isolated from a patient with a chronic hospital-acquired catheter-associated
urinary tract infection. The 7.1-Mb genome sequence organized in 24 scaffolds
contributes to the understanding of biofilm formation and antibiotic resistance.

<>

<1>Tikchonenko, T.I., Karamov, E.V., Zavizion, B.A., Naroditsky, B.S.
<2>EcoRI* activity:  Enzyme modification or activation of accompanying endonuclease?
<3>Gene
<4>4
<5>195-212
<6>1978
<7>A study has been made of the factors and mechanism leading to appearance of the
so-called EcoRI* activity described by Polisky et al. (1975) in the restrictase
EcoRI preparations.  The preparations of purified restrictase EcoRI,
precipitated at 0.9 ammonium sulphate saturation, as well as that obtained
using standard techniques have been found to contain an admixture of an
endonuclease which at neutral pH and high ionic strength multiply cleaves those
DNAs which normally have only one recognition site for EcoRI.  Under the
standard conditions for EcoRI digestion this activity is found only when large
amounts of freshly isolated enzyme are added to the incubation mixture and it
is sharply enhanced by replacement of Mg2+ with Mn2+.  The number and size of
DNa fragments produced under such conditions practically do not differ from
those found under the so-called EcoRI* conditions, that is for alkaline pH
values and low ionic strength.  The optimum incubation mixture for the EcoRI*
activity has been found to be 10 mM Tris-HCl buffer (pH 8.8) + 2 mM Mn2+.
Similar activity is induced also by addition to EcoRI solution of 40-50%
glycerol or a number of organic solvents (dimethylacetamide (DMA),
dimethylformamide (DMF), dimethylsulphoxide (DMSO), sulphalane (SP)) in
concentrations from 1 to 6%.  The EcoRI* activity induced by 50% glycerol or at
alkaline pH values and low ionic strength is suppressed or sharply inhibited by
2-3 mM parachloromercuribenzoate (PCMB), while EcoRI is not sensitive to this
agent.  The DNA fragments cleaved by EcoRI* have cohesive termini and can be
easily ligated.  It is suggested that the EcoRI* activity can be due not only
(or largely not) to modification of the "recognizing capacity" of the EcoRI
restrictase but to activation of a latent specific endonuclease which is
present in the restrictase preparation as an impurity.

<>

<1>Tikhonova, T.N., Malyuta, S.S., Nesterenko, V.F.
<2>DNA-methyltransferases from different Bacillus subtilis and Bacillus natto strains.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>11
<5>40-44
<6>1986
<7>DNA-methyltransferase activity has been detected in some Bacillus subtilis and Bacillus natto
strains.  Two strains of Bacillus subtilis exhibited DNA-cytosine methyltransferase activity,
and the strains of Bacillus natto exhibited DNA-adenine methyltransferase activity.  A
possible effect of DNA-methyltransferase specificity on transformation efficiency is
discussed.

<>

<1>Tilghman, S.M.
<2>DNA methylation: A phoenix rises.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>8761-8762
<6>1993
<7>A report in this issue of the Proceedings by Baylin and his colleagues describes a potential
new way in which DNA methylation affects the physiology of the mammalian cell. The authors
begin from the observation that transformed cells often display higher overall levels of DNA
methylation, as well as increased levels of the hemimethylase, DNA methyltransferase. The
preferred substrate for this enzyme is double-stranded DNA which is hemimethylated on the
cytosine residue of the dinucleotide CpG. When the investigators introduced extra copies of
the enzyme into nontumorigenic NIH 3T3 mouse fibroblasts by gene transfer, the cells acquired
tumorigenic properties, such as loss of contact inhibition, growth in soft agar, and the
ability to form tumors in nude mice.

<>

<1>Timar, E., Groma, G., Kiss, A., Venetianer, P.
<2>Changing the recognition specificity of a DNA-methyltransferase by in vitro evolution.
<3>Nucleic Acids Res.
<4>32
<5>3898-3903
<6>2004
<7>The gene coding for the SinI DNA-methyltransferase, a modification enzyme able to recognize
and methylate the internal cytosine of the GGWCC sequence, was subjected to in vitro
mutagenesis, DNA-shuffling and a strong selection for relaxed GGNCC recognition specificity.
As a result of this in vitro evolution experiment, a mutant gene with the required phenotype
was selected. The mutant SinI methyltransferase carried five amino acid substitutions. None of
these was found in the 'variable region' that were thought to be responsible for sequence
specificity. Three were located near the N-terminal end, preceding the first conserved
structural motif of the enzyme; two were found between conserved motifs VI and VII. A clone
engineered to carry out only the latter two replacements (L214S and Y229H) displays relaxed
recognition specificity similar to that of the parental mutant, whereas the clone carrying
only the N-terminal replacements showed a much weaker change in recognition specificity. The
enzyme with two internal mutations was purified and characterized. Its catalytic activity
(kcat/Km) was  5-fold lower towards GGWCC and 20-fold higher towards GGSCC than that of the
wild-type enzyme.

<>

<1>Timar, E., Venetianer, P., Kiss, A.
<2>In vivo DNA protection by relaxed-specificity SinI DNA methyltransferase variants.
<3>J. Bacteriol.
<4>190
<5>8003-8008
<6>2008
<7>The SinI DNA methyltransferase, a component of the SinI restriction-modification system,
recognizes the sequence GG(A/T)CC and
methylates the inner cytosine to produce 5-methylcytosine. Previously
isolated relaxed-specificity mutants of the enzyme also methylate, at a
lower rate, GG(G/C)CC sites. In this work we tested the capacity of the
mutant enzymes to function in vivo as the counterpart of a restriction
endonuclease, which can cleave either site. The viability of Escherichia
coli cells carrying recombinant plasmids with the mutant methyltransferase
genes and expressing the GGNCC-specific Sau96I restriction endonuclease
from a compatible plasmid was investigated. The sau96IR gene on the latter
plasmid was transcribed from the araBAD promoter, allowing tightly
controlled expression of the endonuclease. In the presence of low
concentrations of the inducer arabinose, cells synthesizing the N172S or
the V173L mutant enzyme displayed increased plating efficiency relative to
cells producing the wild-type methyltransferase, indicating enhanced
protection of the cell DNA against the Sau96I endonuclease. Nevertheless,
this protection was not sufficient to support long-term survival in the
presence of the inducer, which is consistent with incomplete methylation
of GG(G/C)CC sites in plasmid DNA purified from the N172S and V173L
mutants. Elevated DNA ligase activity was shown to further increase
viability of cells producing the V173L variant and Sau96I endonuclease.

<>

<1>Timinskas, A., Butkus, V., Janulaitis, A.
<2>Sequence motifs characteristic for DNA [cytosine-N4] and DNA [adenine-N6] methyltransferases.  Classification of all DNA methyltransferases.
<3>Gene
<4>157
<5>3-11
<6>1995
<7>Two additional conserved motifs (CM), CMIs and CMIII, have been found in addition to
well-known CMI and CMII within the primary amino acid sequences of almost all m6A- and
m4C-methyltransferases (MTases).  The boundaries of all four CM were defined and their
consensus sequences characteristic both for different classes, as well as for all N-MTases,
were derived.  Some regular deviations at fixed positions of the consensus sequences CMIs, CMI
and CMiI, typical for separate classes of N-MTases, were presumed to correlate.  A possible
structural basis for the supposed interregional correlations is discussed and experiments for
verification of the assumed interactions between CM are suggested.  A classification scheme
for all N-MTases is provided.

<>

<1>Timko, J., Horwitz, A.H., Zelinka, J., Wilcox, G.
<2>Characterization of a site-specific restriction endonuclease from Streptomyces aureofaciens.
<3>J. Bacteriol.
<4>145
<5>873-877
<6>1981
<7>A new type II sequence-specific restriction endonuclease, SauI, was isolated
from Streptomyces aureofaciens IKA18/4.  The purified enzyme was free of
contaminating exonuclease and phosphatase activities.  SauI cleaved lambda DNA
at two sites, but did not cleave pBR322, simian virus 40, or PhiX174 DNA.  SauI
recognized the septanucleotide sequence 5'-CC^TNAGG-3' and cleaved at the
position indicated by the arrow, producing a trinucleotide 5'-terminal
extension.

<>

<1>Timko, J., Matuskova, M., Zelinkova, E., Zelinka, J.
<2>Specific endonucleases in Streptomyces aureofaciens.
<3>Folia Microbiol. (Praha)
<4>23
<5>243-245
<6>1978
<7>Two strains of Streptomyces aureofaciens were found to contain restriction
endodeoxynucleases; S. aureofaciens TKA 18/4 contains SauI which splits lambda
DNA into three fragments, S. aureofaciens IKA 22201 contains SauI which splits
lambda DNA into more than 15 fragments.

<>

<1>Timko, J., Sisova, M., Ugorcakova, J., Zelinka, J.
<2>Restriction endonuclease SauI from Streptomyces aureofaciens - some physical and chemical properties.
<3>Biologia (Bratisl)
<4>43
<5>681-689
<6>1988
<7>Some physical and chemical properties of restriction endonuclease SauI, which
was isolated from Streptomyces aureofaciens IKA 18/4, were studied.  The
effects of divalent ions, monovalent ions, pH and temperature on enzyme
activity were determined.  For cleavage of substrate DNA by endonuclease SauI
the optimal reaction mixture was determined as: 10 mM Tris-HCl, pH 7.5, 10 mM
MgCl2, 75 mM NaCl at 37C.

<>

<1>Timko, J., Turna, J., Zelinka, J.
<2>Site-specific restriction endonuclease SauBMKI from Streptomyces aureofaciens.
<3>Metabolism and Enzymology of Nucleic Acids, Publishing House of the Slovak Akademy of Science, Zelinka, J., Balan, J., Bratislava
<4>6
<5>107-118
<6>1987
<7>Type II restriction endonucleases have been isolated from many bacteria
including Streptomycetes and their specificities have been characterized
(Roberts, 1985).  From Streptomyces aureofaciens restriction endonucleases SauI
(Timko et al., 1981) and Sau3239I (Gasperik et al., 1983; Simbochova et al.,
1986) have been isolated and characterized.  In this paper we describe a new
restriction endonuclease, SauBMKI from Streptomyces aureofaciens, strain BM-K,
producing about 2500 micrograms of CTC per ml.

<>

<1>Timko, J., Zelinka, J., Wilcox, G.
<2>Properties of the restriction endonuclease Saul.
<3>Metabolism and Enzymology of Nucleic Acids, Publishing House of the Slovak Akademy of Science., Zelinka, J., Balan, J., Bratislava
<4>4
<5>319-324
<6>1982
<7>None

<>

<1>Timme, R.E., Allard, M.W., Luo, Y., Strain, E., Pettengill, J., Wang, C., Li, C., Keys, C.E., Zheng, J., Stones, R., Wilson, M.R., Musser, S.M., Brown, E.W.
<2>Draft Genome Sequences of 21 Salmonella enterica Serovar Enteritidis Strains.
<3>J. Bacteriol.
<4>194
<5>5994-5995
<6>2012
<7>Salmonella enterica subsp. enterica serovar Enteritidis is a common food-borne pathogen, often
associated with shell eggs and poultry. Here, we report draft
genomes of 21 S. Enteritidis strains associated with or related to the U.S.-wide
2010 shell egg recall. Eleven of these genomes were from environmental isolates
associated with the egg outbreak, and 10 were reference isolates from previous
years, unrelated to the outbreak. The whole-genome sequence data for these 21
human pathogen strains are being released in conjunction with the newly formed
100K Genome Project.

<>

<1>Timme, R.E., Pettengill, J.B., Allard, M.W., Strain, E., Barrangou, R., Wehnes, C., Van Kessel, J.S., Karns, J.S., Musser, S.M., Brown, E.W.
<2>Phylogenetic diversity of the enteric pathogen Salmonella enterica subsp. enterica inferred from genome-wide reference-free SNP characters.
<3>Genome Biol. Evol.
<4>5
<5>2109-2123
<6>2013
<7>The enteric pathogen Salmonella enterica is one of the leading causes of
foodborne illness in the world. The species is extremely diverse, containing more
than 2,500 named serovars that are designated for their unique antigen characters
and pathogenicity profiles-some are known to be virulent pathogens, while others
are not. Questions regarding the evolution of pathogenicity, significance of
antigen characters, diversity of clustered regularly interspaced short
palindromic repeat (CRISPR) loci, among others, will remain elusive until a
strong evolutionary framework is established. We present the first large-scale S.
enterica subsp. enterica phylogeny inferred from a new reference-free k-mer
approach of gathering single nucleotide polymorphisms (SNPs) from whole genomes.
The phylogeny of 156 isolates representing 78 serovars (102 were newly sequenced)
reveals two major lineages, each with many strongly supported sublineages. One of
these lineages is the S. Typhi group; well nested within the phylogeny.
Lineage-through-time analyses suggest there have been two instances of
accelerated rates of diversification within the subspecies. We also found that
antigen characters and CRISPR loci reveal different evolutionary patterns than
that of the phylogeny, suggesting that a horizontal gene transfer or possibly a
shared environmental acquisition might have influenced the present character
distribution. Our study also shows the ability to extract reference-free SNPs
from a large set of genomes and then to use these SNPs for phylogenetic
reconstruction. This automated, annotation-free approach is an important step
forward for bacterial disease tracking and in efficiently elucidating the
evolutionary history of highly clonal organisms.

<>

<1>Timms, A.R., Cambray-Young, J., Scott, A.E., Petty, N.K., Connerton, P.L., Clarke, L., Seeger, K., Quail, M., Cummings, N., Maskell, D.J., Thomson, N.R., Connerton, I.F.
<2>Evidence for a lineage of virulent bacteriophages that target Campylobacter.
<3>BMC Genomics
<4>11
<5>214
<6>2010
<7>Background: Our understanding of the dynamics of genome stability versus gene flux within
bacteriophage lineages is limited. Recently,
there has been a renewed interest in the use of bacteriophages as
'therapeutic' agents; a prerequisite for their use in such therapies is
a thorough understanding of their genetic complement, genome stability
and their ecology to avoid the dissemination or mobilisation of phage
or bacterial virulence and toxin genes. Campylobacter, a food-borne
pathogen, is one of the organisms for which the use of bacteriophage is
being considered to reduce human exposure to this organism.
Results: Sequencing and genome analysis was performed for two
Campylobacter bacteriophages. The genomes were extremely similar at the
nucleotide level (>= 96%) with most differences accounted for by novel
insertion sequences, DNA methylases and an approximately 10 kb
contiguous region of metabolic genes that were dissimilar at the
sequence level but similar in gene function between the two phages.
Both bacteriophages contained a large number of radical
S-adenosylmethionine (SAM) genes, presumably involved in boosting host
metabolism during infection, as well as evidence that many genes had
been acquired from a wide range of bacterial species. Further
bacteriophages, from the UK Campylobacter typing set, were screened for
the presence of bacteriophage structural genes, DNA methylases, mobile
genetic elements and regulatory genes identified from the genome
sequences. The results indicate that many of these bacteriophages are
related, with 10 out of 15 showing some relationship to the sequenced
genomes.
Conclusions: Two large virulent Campylobacter bacteriophages were
found to show very high levels of sequence conservation despite
separation in time and place of isolation. The bacteriophages show
adaptations to their host and possess genes that may enhance
Campylobacter metabolism, potentially advantaging both the
bacteriophage and its host. Genetic conservation has been shown to
extend to other Campylobacter bacteriophages, forming a highly
conserved lineage of bacteriophages that predate upon campylobacters
and indicating that highly adapted bacteriophage genomes can be stable
over prolonged periods of time.

<>

<1>Tindall, B.J. et al.
<2>Complete genome sequence of Meiothermus ruber type strain (21T).
<3>Standards in Genomic Sciences
<4>3
<5>26-36
<6>2010
<7>Meiothermus ruber (Loginova et al. 1984) Nobre et al. 1996 is the type species of the genus
Meiothermus. This thermophilic genus is of special interest, as its members share relatively
low degrees of 16S rRNA gene sequence similarity and constitute a separate evolutionary
lineage from members of the genus Thermus, from which they can generally be distinguished by
their slightly lower temperature optima. The temperature related split is in accordance with
the chemotaxonomic feature of the polar lipids. M. ruber is a representative of the
low-temperature group. This is the first completed genome sequence of the genus Meiothermus
and only the third genome sequence to be published from a member of the family Thermaceae. The
3,097,457 bp long genome with its 3,052 protein-coding and 53 RNA genes is a part of the
Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Tindall, B.J. et al.
<2>Complete genome sequence of Halomicrobium mukohataei type strain (arg-2).
<3>Standards in Genomic Sciences
<4>1
<5>270-277
<6>2009
<7>Halomicrobium mukohataei (Ihara et al. 1997) Oren et al. 2002 is the type species of the genus
Halomicrobium. It is of phylogenetic interest because of its
isolated location within the large euryarchaeal family Halobacteriaceae. H.
mukohataei is an extreme halophile that grows essentially aerobically, but can
also grow anaerobically under a change of morphology and with nitrate as electron
acceptor. The strain, whose genome is described in this report, is a free-living,
motile, Gram-negative euryarchaeon, originally isolated from Salinas Grandes in
Jujuy, Andes highlands, Argentina. Its genome contains three genes for the 16S
rRNA that differ from each other by up to 9%. Here we describe the features of
this organism, together with the complete genome sequence and annotation. This is
the first completed genome sequence from the poorly populated genus
Halomicrobium, and the 3,332,349 bp long genome (chromosome and one plasmid) with
its 3416 protein-coding and 56 RNA genes is part of the Genomic Encyclopedia of
Bacteria and Archaea project.

<>

<1>Tippelt, A., Albersmeier, A., Brinkrolf, K., Ruckert, C., Fernandez-Natal, I., Soriano, F., Tauch, A.
<2>Complete Genome Sequence of Corynebacterium ureicelerivorans DSM 45051, a Lipophilic and Urea-Splitting Isolate from the Blood Culture of a Septicemia  Patient.
<3>Genome Announcements
<4>2
<5>e01211-14
<6>2014
<7>Corynebacterium ureicelerivorans is an opportunistic pathogen with a lipophilic lifestyle and
an exceptionally high urease activity. The genome sequence of the
type strain revealed that lipophilism is caused by the lack of a fatty acid
synthase gene. The ureABCEFGD genes are similar to the urease gene region of
Corynebacterium glucuronolyticum.

<>

<1>Tippelt, A., Mollmann, S., Albersmeier, A., Jaenicke, S., Ruckert, C., Tauch, A.
<2>Mycolic Acid Biosynthesis Genes in the Genome Sequence of Corynebacterium atypicum DSM 44849.
<3>Genome Announcements
<4>2
<5>e00845-14
<6>2014
<7>The complete chromosomal sequence of the type strain Corynebacterium atypicum DSM 44849
comprises 2,311,380 bp. A functional annotation revealed the presence of
genes involved in the synthesis and export of mycolic acids and in trehalose
corynomycolate biosynthesis, supporting the view that the cell envelope of C.
atypicum contains mycolic acids.

<>

<1>Tishchenko, E.N., Dubrovnaya, O.V.
<2>Methyltransferases of plants.
<3>Biopol. Kletka
<4>21
<5>463-472
<6>2005
<7>In the review the current data on the plant cytosine-C5-DNA-methyltransferases
(methyltransferase) are summarized.  Three different classes of these enzymes - Snmt1/MET1,
Dnmt3 and CMT are described. The proposed function, dealing with maintenance and de novo
methylation of cytosine residues in symmetric and asymmetric motifs of DNA, is under
discussion.

<>

<1>Titball, R.W., Mayers, C.N., Duffield, M.L., Miller, J., Rowe, S.C.
<2>Francisella tularensis immunogenic proteins and DNA encoding these.
<3>International Patent Office
<4>WO 2004003009 A
<5>
<6>2004
<7>Proteins and nucleic acids for use in producing a protective immune response in a mammal
against infection by Francisella tularensis, are described and claimed.  The proteins are
selected from the SEQ ID NO 1-100 as provided in the application, or protective variants
thereof, or fragments of either of these.

<>

<1>Titeeva, G.R., Vinogradov, S.V., Berlin, Y.A.
<2>Specific features of the restriction endonuclease BamHI interaction with oligodeoxynucleotides.
<3>Bioorg. Khim.
<4>12
<5>640-646
<6>1986
<7>Interaction of the restriction endonuclease BamHI with a series of synthetic
oligodeoxynucleotides containing the restriction site has been studied.  The
enzyme is shown to specifically cut the BamHI site in hexanucleotide (I) and in
non-selfcomplementary deca- and octanucleotides (II)-(IV).  The data obtained
led to the conclusion that BamHI reacts with duplex structures, while playing
an important role in their stabilization.  In 14-mer (V) BamHI cuts a
non-standard half-site GAA to yield the 5'-terminal tri- (rather than hepta-)
nucleotide.  Hypothetical mechanisms of the process are discussed basing on
conception of the role of higher DNA structures in the interaction with
restriction endonucleases.

<>

<1>Titeeva, G.R., Vinogradov, S.V., Berlin, Y.A.
<2>Complexes of the phage single-stranded DNA with synthetic oligodeoxynucleotides and their interaction with DNA polymerase and restriction endonucleases.
<3>Bioorg. Khim.
<4>12
<5>1484-1491
<6>1986
<7>Hybridization of synthetic oligodeoxynucleotides with single-stranded phage
M13mp2 DNA has been studied in terms of temperature, ionic strength,
oligonucleotide molar excess and chain length, and DNA secondary structure.
Combination of two decadeoxynucleotides corresponding to a nicked eicosamer
(composite primer) was found to be efficient in the template-directed DNA
polymerase-catalyzed chain elongation, where both decamers separately failed.
Circular SS DNA was specifically linearized by BamHI cleavage of a SS DNA -
tetradecadeoxynucleotide duplex.

<>

<1>Titheradge, A.J., King, J., Ryu, J., Murray, N.E.
<2>Families of restriction enzymes: an analysis prompted by molecular and genetic data for type ID restriction and modification systems.
<3>Nucleic Acids Res.
<4>29
<5>4195-4205
<6>2001
<7>Current genetic and molecular evidence places all the known type I restriction and
modification systems of Escherichia coli and Salmonella enterica into one of four discrete
families: type IA, IB, IC or ID. StySBLI is the founder member of the ID family. Similarities
of coding sequences have identified restriction systems in E.coli and Klebsiella pneumoniae as
probable members of the type ID family. We present complementation tests that confirm the
allocation of EcoR9I and KpnAI to the ID family. An alignment of the amino acid sequences of
the HsdS subunits of StySBLI and EcoR9I identify two variable regions, each predicted to be a
target recognition domain (TRD). Consistent with two TRDs, StySBLI was shown to recognise a
bipartite target sequence, but one in which the adenine residues that are the substrates for
methylation are separated by only 6 bp. Implications of family relationships are discussed and
evidence is presented that extends the family affiliations identified in enteric bacteria to a
wide range of other genera.

<>

<1>Titheradge, A.J.B., Ternent, D., Murray, N.E.
<2>A third family of allelic hsd genes in Salmonella enterica: sequence comparisons with related proteins identify conserved regions implicated in restriction of DNA.
<3>Mol. Microbiol.
<4>22
<5>437-447
<6>1996
<7>Salmonella enterica serovar blegdam has a restriction and modification system encoded by genes
linked to serB.  We have cloned these genes, putative alleles of the hsd locus of Escherichia
coli K-12, and confirmed by the sequence similarities of flanking DNA that the hsd genes of S.
enterica serovar blegdam have the same chromosomal location as those of E. coli K-12 and
Salmonella enterica serovar typhimurium LT2.  There is, however, no obvious similarity in
their nucleotide sequences, and while the gene order in S. enterica serovar blegdam is serB
hsdM, S and R, that in E. coli K-12 and S. enterica serovar typhimurium LT2 is serB hsdR, M
and S.  The hsd genes of S. enterica serovar blegdam identify a third family of serB-linked
hsd genes (type ID).  The polypeptide sequence predicted from the three hsd genes show some
similarities (18-50% identity) with the polypeptides of known and putative type I restriction
and modification systems; the highest levels of identity are with sequences of Haemophilus
influenzae Rd.  The HsdM polypeptide has the motifs characteristic of adenine
methyltransferases.  Comparisons of the HsdR sequence with those for three other families of
type I systems and three putative HsdR polypeptides identify two highly conserved regions in
addition to the seven proposed DEAD-box motifs.

<>

<1>Tiwari, P.K., Joshi, K., Rehman, R., Bhardwaj, V., Shamsudheen, K.V., Sivasubbu, S., Scaria, V.
<2>Draft Genome Sequence of Urease-Producing Sporosarcina pasteurii with Potential Application in Biocement Production.
<3>Genome Announcements
<4>2
<5>e01256-13
<6>2014
<7>We describe here the draft genome sequence of Sporosarcina pasteurii, a urease-producing
bacterium with potential applications in biocement production.

<>

<1>Tkachuk, J.Y., Tkachuk, L.V., Kvasnyuk, E.I., Zaitseva, G.V., Kalinichenko, E.M., Mikhailopulo, I.O., Matsuka, G.K.
<2>Functioning of EcoRI, BamHI and HindIII restriction enzymes as affected by (2',5') oligoadenylates.
<3>Dokl. Akad. Nauk UKR SSR Ser. B Geol. Khim. Biol. Nauk
<4>10
<5>78-81
<6>1988
<7>Peculiarities of the functioning of restriction enzymes EcoRI, BamHI and
HindIII in the presence of core trimers of (2'-5') oligoadenylic acid, viz.,
(2'-5') A3 and (2'-5')3'dA3, have been studied.  Depending on the enzyme, the
structure of the (2'-5') trimer, its concentration and method of its use
(preincubation of the enzyme and the trimer or successive mixing of the
reaction components), an inhibition of the restriction reaction, or
site-specific restriction of lambda DNA or non-specific hydrolysis of lambda
DNA are observed.  In the latter case, the restriction enzymes function as
nonspecific DNases.

<>

<1>Tkachuk, Z.Y., Tkachuk, L.V., Kvasyuk, E.I., Zaitseva, G.V., Kalinichenko, E.N., Mikhailopulo, I.A., Matsuka, G.K.
<2>Changes in functional properties of restriction enzymes under the influence of (2'-5') oligoadenylates in vitro.
<3>Biopol. Kletka
<4>5
<5>69-73
<6>1989
<7>It is found that core trimers of (2'-5') oligoadenylic acid. viz., (A2'p)2A and
(3'dA2'p)2/3'dA, exert an influence on functional properties of restriction
enzymes EcoRI, BamHI, and HindIII.  The presence of (2'-5') trimers in the
reaction mixture containing DNA and the restriction enzyme results in (i) an
inhibition of the restriction reaction, (ii) site-specific restriction of DNA
lambda, or (iii) nonspecific hydrolysis of DNA lambda.  In the last case
restriction enzymes function as nonspecific DNAses.

<>

<1>Tkaczuk, K.L., Obarska, A., Bujnicki, J.M.
<2>Molecular phylogenetics and comparative modeling of HEN1, a methyltransferase involved in plant microRNA biogenesis.
<3>BMC Evol. Biol.
<4>6
<5>6
<6>2006
<7>BACKGROUND: Recently, HEN1 protein from Arabidopsis thaliana was discovered as an essential
enzyme in plant microRNA (miRNA) biogenesis. HEN1 transfers a methyl group from
S-adenosylmethionine to the 2'-OH or 3'-OH group of the last nucleotide of miRNA/miRNA*
duplexes produced by the nuclease Dicer. Previously it was found that HEN1 possesses a
Rossmann-fold methyltransferase (RFM) domain and a long N-terminal extension including a
putative double-stranded RNA-binding motif (DSRM). However, little is known about the details
of the structure and the mechanism of action of this enzyme, and about its phylogenetic
origin. RESULTS: Extensive database searches were carried out to identify orthologs and close
paralogs of HEN1. Based on the multiple sequence alignment a phylogenetic tree of the HEN1
family was constructed. The fold-recognition approach was used to identify related
methyltransferases with experimentally solved structures and to guide the homology modeling of
the HEN1 catalytic domain. Additionally, we identified a La-like predicted RNA binding domain
located C-terminally to the DSRM domain and a domain with a peptide prolyl cis/trans isomerase
(PPIase) fold, but without the conserved PPIase active site, located N-terminally to the
catalytic domain. CONCLUSION: The bioinformatics analysis revealed that the catalytic domain
of HEN1 is not closely related to any known RNA:2'-OH methyltransferases (e.g. to the
RrmJ/fibrillarin superfamily), but rather to small-molecule methyltransferases. The structural
model was used as a platform to identify the putative active site and substrate-binding
residues of HEN and to propose its mechanism of action.

<>

<1>Tobes, R., Codoner, F.M., Lopez-Camacho, E., Salanueva, I.J., Manrique, M., Brozynska, M., Gomez-Gil, R., Martinez-Blanch, J.F., Alvarez-Tejado, M., Pareja, E., Mingorance, J.
<2>Genome Sequence of Klebsiella pneumoniae KpQ3, a DHA-1 beta-Lactamase-Producing Nosocomial Isolate.
<3>Genome Announcements
<4>1
<5>e00167-12
<6>2013
<7>KpQ3 is a multidrug-resistant isolate obtained from a blood culture of a patient  in a burn
unit in the Hospital Universitario La Paz (Madrid, Spain) in 2008. The
genome contains multiple antibiotic resistance genes, including a
plasmid-mediated DHA-1 cephalosporinase gene.

<>

<1>Tobes, R., Manrique, M., Brozynska, M., Stephan, R., Pareja, E., Johler, S.
<2>Noncontiguous Finished Genome Sequence of Staphylococcus aureus KLT6, a Staphylococcal Enterotoxin B-Positive Strain Involved in a Food Poisoning  Outbreak in Switzerland.
<3>Genome Announcements
<4>1
<5>e00214-13
<6>2013
<7>We present the first complete genome sequence of a Staphylococcus aureus strain assigned to
clonal complex 12. The strain was isolated in a food poisoning
outbreak due to contaminated potato salad in Switzerland in 2009, and it produces
staphylococcal enterotoxin B.

<>

<1>Tobias, N.J., Doig, K.D., Medema, M.H., Chen, H., Haring, V., Moore, R., Seemann, T., Stinear, T.P.
<2>Complete Genome Sequence of the Frog Pathogen Mycobacterium ulcerans ecovar Liflandii.
<3>J. Bacteriol.
<4>195
<5>556-564
<6>2012
<7>In 2004, a previously undiscovered mycobacterium resembling Mycobacterium ulcerans (the agent
of Buruli ulcer) was reported in an outbreak of a lethal mycobacteriosis in a laboratory
colony of the African clawed-frog Xenopus tropicalis. This mycobacterium makes mycolactone and
is one of several strains of M. ulcerans-like mycolactone-producing mycobacteria recovered
from ectotherms around the world. Here we describe the complete 6,399,543 bp genome of this
frog pathogen (previously unofficially named Mycobacterium 'liflandii') and we show that it
has undergone an intermediate degree of reductive evolution, between M. ulcerans Agy99 and the
fish pathogen Mycobacterium marinum M. Like M. ulcerans Agy99, it has the pMUM mycolactone
plasmid, over 200 chromosomal copies of the insertion sequence IS2404 and a high proportion of
pseudogenes. However, M. 'liflandii' has a larger genome that is closer in length, sequence
and architecture to M. marinum M than to M. ulcerans Agy99, suggesting that M. ulcerans Agy99
has undergone accelerated evolution. Scrutiny of genes specifically lost suggests M.
'liflandii' is a tryptophan, tyrosine and phenylalanine auxotroph. A once extensive M.
marinum-like secondary metabolome has also been diminished through reductive evolution. Our
analysis shows that M. 'liflandii', like M. ulcerans Agy99, has the characteristics of a
niche-adapted mycobacterium but with several distinctive features in important metabolic
pathways that suggests it is responding to different environmental pressures, supporting
earlier proposals that it could be considered an M. ulcerans ecotype, hence the name M.
ulcerans ecovar Liflandii.

<>

<1>Tock, M.R., Dryden, D.T.F.
<2>The biology of restriction and anti-restriction.
<3>Curr. Opin. Microbiol.
<4>8
<5>466-472
<6>2005
<7>The phenomena of prokaryotic restriction and modification, as well as anti-restriction, were
first discovered five decades ago but have
yielded only gradually to rigorous analysis. Work presented at the
5(th) New England Biolabs Meeting on Restriction-Modification
(available on REBASE, http://www.rebase.com) and several recently
published genetic, biochemical and biophysical analyses indicate that
these fields continue to contribute significantly to basic science.
Recently, there have been several studies that have shed light on the
still developing field of restriction-modification and on the newly
re-emerging field of anti-restriction.

<>

<1>Toh, H., Oshima, K., Nakano, A., Takahata, M., Murakami, M., Takaki, T., Nishiyama, H., Igimi, S., Hattori, M., Morita, H.
<2>Genomic adaptation of the Lactobacillus casei group.
<3>PLoS ONE
<4>8
<5>e75073
<6>2013
<7>Lactobacillus casei, L. paracasei, and L. rhamnosus form a closely related taxonomic group
(Lactobacillus casei group) within the facultatively heterofermentative lactobacilli. Here, we
report the complete genome sequences of L. paracasei JCM 8130 and L. casei ATCC 393, and the
draft genome sequence of L. paracasei COM0101, all of which were isolated
from daily products. Furthermore, we re-annotated the genome of L. rhamnosus ATCC 53103 (also
known as L. rhamnosus GG), which we have previously reported. We confirmed that ATCC 393 is
distinct from other strains previously described as L. paracasei. The core genome of 10
completely sequenced strains of the L. casei group comprised 1,682 protein-coding genes.
Although extensive genome-wide synteny was found among the L. casei group, the genomes of ATCC
53103, JCM 8130, and ATCC 393 contained genomic islands compared with L. paracasei ATCC 334.
Several genomic islands, including carbohydrate utilization gene clusters, were found at the
same loci in the chromosomes of the L. casei group. The spaCBA pilus gene cluster, which was
first identified in GG, was also found in other strains of the L. casei group, but several L.
paracasei strains including COM0101 contained truncated spaC gene. ATCC 53103 encoded a higher
number of proteins involved in carbohydrate utilization compared with intestinal
lactobacilli, and extracellular adhesion proteins, several of which are absent in other
strains of the L. casei group. In addition to previously fully sequenced L. rhamnosus and L.
paracasei strains, the complete genome sequences of L. casei will provide valuable insights
into the evolution of the L. casei group.

<>

<1>Toh, H., Oshima, K., Suzuki, T., Hattori, M., Morita, H.
<2>Complete Genome Sequence of the Equol-Producing Bacterium Adlercreutzia equolifaciens DSM 19450T.
<3>Genome Announcements
<4>1
<5>e00742-13
<6>2013
<7>Adlercreutzia equolifaciens DSM 19450(T) was isolated from human feces and is able to
metabolize daidzeins (soybean isoflavonoids) to equol. Here, we report
the finished and annotated genome sequence of this organism.

<>

<1>Toh, H., Oshima, K., Toyoda, A., Ogura, Y., Ooka, T., Sasamoto, H., Park, S.H., Iyoda, S., Kurokawa, K., Morita, H., Itoh, K., Taylor, T.D., Hayashi, T., Hattori, M.
<2>Complete genome sequence of the wild-type commensal Escherichia coli strain SE15 belonging to phylogenetic group B2.
<3>J. Bacteriol.
<4>192
<5>1165-1166
<6>2009
<7>Escherichia coli SE15 (O150:H5) is a human commensal bacterium recently isolated from feces of
a healthy adult and classified into E. coli
phylogenetic group B2 that includes the majority of extraintestinal
pathogenic E. coli (ExPEC). Here, we report the finished and annotated
genome sequence of this organism.

<>

<1>Toh, H., Sharma, V.K., Oshima, K., Kondo, S., Hattori, M., Ward, F.B., Free, A., Taylor, T.D.
<2>Complete Genome Sequences of Arcobacter butzleri ED-1 and Arcobacter sp. Strain L, Both Isolated from a Microbial Fuel Cell.
<3>J. Bacteriol.
<4>193
<5>6411-6412
<6>2011
<7>Arcobacter butzleri strain ED-1 is an exoelectrogenic epsilonproteobacterium isolated from the
anode biofilm of a microbial fuel
cell. Arcobacter sp. strain L dominates the liquid phase of the same fuel
cell. Here we report the finished and annotated genome sequences of these
organisms.

<>

<1>Toh, H., Weiss, B.L., Perkin, S.A., Yamashita, A., Oshima, K., Hattori, M., Aksoy, S.
<2>Massive genome erosion and functional adaptations provide insights into the symbiotic lifestyle of Sodalis glossinidius in the tsetse host.
<3>Genome Res.
<4>16
<5>149-156
<6>2006
<7>Sodalis glossinidius is a maternally transmitted endosymbiont of tsetse flies (Glossina spp.),
an insect of medical and veterinary significance. Analysis of the complete sequence of
Sodalis' chromosome (4,171,146 bp, encoding 2,432 protein coding sequences) indicates a
reduced coding capacity of 51%. Furthermore, the chromosome contains 972 pseudogenes, an
inordinately high number compared with that of other bacterial species. A high proportion of
these pseudogenes are homologs of known proteins that function either in defense or in the
transport and metabolism of carbohydrates and inorganic ions, suggesting Sodalis'
degenerative adaptations to the immunity and restricted nutritional status of the host.
Sodalis possesses three chromosomal symbiosis regions (SSR): SSR-1, SSR-2, and SSR-3, with
gene inventories similar to the Type-III secretion system (TTSS) ysa from Yersinia
enterolitica and SPI-1 and SPI-2 from Salmonella, respectively. While core components of the
needle structure have been conserved, some of the effectors and regulators typically
associated with these systems in pathogenic microbes are modified or eliminated in Sodalis.
Analysis of SSR-specific invA transcript abundance in Sodalis during host development
indicates that the individual symbiosis regions may exhibit different temporal expression
profiles. In addition, the Sodalis chromosome encodes a complete flagella structure, key
components of which are expressed in immature host developmental stages. These features may be
important for the transmission and establishment of symbiont infections in the intra-uterine
progeny. The data suggest that Sodalis represents an evolutionary intermediate transitioning
from a free-living to a mutualistic lifestyle.

<>

<1>Tokajian, S., Eisen, J.A., Jospin, G., Coil, D.A.
<2>Draft Genome Sequences of Streptococcus pyogenes Strains Associated with Throat and Skin Infections in Lebanon.
<3>Genome Announcements
<4>2
<5>e00358-14
<6>2014
<7>We present the draft genome sequences of nine clinical Streptococcus pyogenes isolates
recovered from patients suffering from sore throat and skin infections.
An average of 2,454,334 paired-end reads per sample were generated, which
assembled into 21 to 198 contigs, with a G+C content of 38.4 to 38.5%.

<>

<1>Tokajian, S., Eisen, J.A., Jospin, G., Farra, A., Coil, D.A.
<2>Draft Genome Sequences of Extended-Spectrum beta-Lactamase-Producing Escherchia coli Strains Isolated from Patients in Lebanon.
<3>Genome Announcements
<4>2
<5>e00123-14
<6>2014
<7>We present the draft genome sequences of nine extended-spectrum beta-lactamase
(ESBL)-producing Escherichia coli strains isolated from stool samples collected
from patients admitted for gastrointestinal and urological procedures/surgeries.
An average of 3,889,300 paired-end reads per sample were generated, which
assembled in 77 to 157 contigs.

<>

<1>Tokajian, S., Eisen, J.A., Jospin, G., Farra, A., Coil, D.A.
<2>Draft Genome Sequence of Extended-Spectrum beta-Lactamase-Producing Klebsiella pneumoniae Isolated from a Patient in Lebanon.
<3>Genome Announcements
<4>2
<5>e00121-14
<6>2014
<7>We present the draft genome sequence of extended-spectrum beta-lactamase (ESBL)-producing
Klebsiella pneumoniae isolated from a stool sample collected
from a patient admitted for a gastrointestinal procedure. The draft genome
sequence consists of 86 contigs, including a combined 5,632,663 bases with 57%
G+C content.

<>

<1>Toksoy, E., Onsan, Z.I., Kirdar, B.
<2>Effect of the co-expression of methyltransferase activity on extracellular production of TaqI restriction endonuclease in recombinant E. coli cells.
<3>Process. Biochem.
<4>37
<5>527-534
<6>2001
<7>TaqI methylase gene was cloned and expressed constitutively under the control of the
tetracycline resistance gene promoter.  The effect of TaqI methylase protection on the growth
kinetics, plasmid stability and production of MBP - TaqI fusion protein by recombinant E. coli
cells was investigated both in shake flasks and under controlled bioreactor conditions.  Shake
flask experiments indicated that the use of nutritionally richer medium formulations may
improve the growth characteristics, plasmid stability and also the total and extracellular
TaqI endonuclease recovery yields by the methylase protected cells.  Under controlled
bioreactor conditions, co-expression of the methylase gene led to higher plasmid stabilities
and improved the excretion levels considerably.

<>

<1>Toksoy, E., Onsan, Z.I., Kirdar, B.
<2>Expression of a thermostable restriction endonuclease in recombinant Escherichia coli cells and optimization of fermentation conditions.
<3>World J. Microbiol. Biotechnol.
<4>18
<5>23-27
<6>2002
<7>TaqI restriction endonuclease gene of the thermophilic eubacterium Thermus aquaticus YT-1
(ATCC 25104) was successfully cloned and expressed in recombinant Escherichia coli cells under
the control of the lac promoter/operator system.  Higher TaqI endonuclease specific activities
and biomass yields were obtained from E. coli ER2508(pUCTaq) cells when they were induced at
the late-exponential phase of their growth.  TaqI endonuclease expression was found to be
host-strain-dependent such that, among the three different strains examined, E. coli
XL1(pUCTaq) produced the highest specific TaqI endonuclease activities for longer induction
periods.  Decreasing the inducer concentration from 1 to 0.1 mM not only improved the specific
enzyme activity yields but also is more economical, considering the high cost of
isopropyl-beta-D-thiogalactopyranoside (PTG).  The optimum culture temperature was found to be
37 C.  TaqI endonuclease specific activity recovered from E. coli XL1(pUCTaq) cells was 935
U/mg under optimum conditions.

<>

<1>Toksoy, E., Onsan, Z.I., Kirdar, B.
<2>High-level production of TaqI restriction endonuclease by three different expression systems in Escherichia coli cells using the T7 phage promoter.
<3>Appl. Microbiol. Biotechnol.
<4>59
<5>239-245
<6>2002
<7>Three different expression systems were constructed for the high-level production of TaqI
restriction endonuclease in recombinant Escherichia coli cells. In system [R], the TaqI
endonuclease gene was cloned and expressed under the control of the strong T7 RNA polymerase
promoter. To protect cellular DNA, methylase protection was provided by constitutive
co-expression of TaqI methylase activity either by cloning the TaqI methylase gene on a second
plasmid (system [R,M]) or by constructing a recombinant plasmid harboring both the
endonuclease and methylase genes (system [R+M]). In batch shake flasks containing complex
media, co-expression of the methylase gene in systems [R,M] and [R+M] resulted in a 2- and
3-fold increase in volumetric productivity over system [R], yielding activities of 250x10^6 U
l^-1 and 350x10^6 U l^-1, which were 28 and 39 times higher than the data in the
literature, respectively. Under controlled bioreactor conditions in chemically defined medium,
co-expression of methylase activity greatly improved the yield and specific TaqI endonuclease
productivity of the recombinant cells, and reduced acetic acid excretion levels. System [R,M]
is preferable for high expression levels at longer operation periods, while system [R+M] is
well-suited for high expression levels in short-term bioreactor operation.

<>

<1>Tollefsbol, T.O., Hutchinson, C.A. III
<2>Analysis in Escherichia coli of the effects of in vivo CpG methylation catalyzed by the cloned murine maintenance methyltransferase.
<3>Biochem. Biophys. Res. Commun.
<4>245
<5>670-678
<6>1998
<7>Due in part to the complexity of mammalian systems, some of the proposed biological influences
of mammalian DNA methylation have not been fully established.  Escherichia coli cells, which
normally contain negligible CpG methylation, exhibited progressive slowing of replication and
lengthened generation times when expressing the murine DNA maintenance methyltransferase.
Genomic analysis indicated significant amounts of CpG methylation in expressing cells which
was absent from control cells.  Expressing cells exposed to the cytosine demethylating agent,
5-azacytidine, rapidly reverted to propagation levels of controls.  Substitution of cysteine
with alanine in the carboxyl-terminal region proline-cysteine dipeptide of the
methyltransferase completely inactivated methylating activity and cells expressing the
inactive enzyme replicated as well as controls.  These findings strongly implicate a role of
epigenetic de novo CpG methylation in modulating cellular propagation, demonstrate that the
maintenance methyltransferase can de novo methylate in vivo, and show that the
methyltransferase requires an active site cysteine for activity.

<>

<1>Tollefsbol, T.O., Hutchison, C.A. III
<2>Mammalian DNA (cytosine-5-)-methyltransferase expressed in Escherichia coli, purified and characterized.
<3>J. Biol. Chem.
<4>270
<5>18543-18550
<6>1995
<7>Besides modulating specific DNA-protein interactions, methylated cytosine, frequently referred
to as the fifth base of the genome, also influences DNA structure, recombination,
transposition, repair, transcription, imprinting, and mutagenesis.  DNA
(cytosine-5-)-methyltransferase catalyzes cytosine methylation in eukaryotes.  We have cloned
and expressed this enzyme in Escherichia coli, purified it to apparent homogeneity,
characterized its properties, and we have shown that it hemimethylates DNA.  The cDNA for
murine maintenance methyltransferase was reconstructed and cloned for direct expression in
native form.  Immunoblotting revealed a unique protein (Mr=190,000) not present in control
cells.  The mostly soluble overexpressed protein was purified by DEAE, Sephadex, and DNA
cellulose chromatography.  Peak methylating activity correlated with methyltransferase
immunoblots.  The purified enzyme preferentially transferred radioactive methyl moieties to
hemimethylated DNA in assays and on autoradiograms.  All of the examined properties of the
purified recombinant DNA methyltransferase are consistent with the enzyme purified from
mammalian cells.  Further characterization revealed enhanced in vitro methylation of
premethylated oligodeoxynucleotides.  The cloning of hemimethyltransferase in E. coli should
allow facilitated structure-function mutational analysis of this enzyme, studies of its
biological effects in prokayotes, and potential large scale methyltransferase production for
crystallography, and it may have broad applications in maintaining the native methylated state
of cloned DNA.

<>

<1>Tollefsbol, T.O., Hutchison, C.A. III
<2>Control of methylation spreading in synthetic DNA sequences by the murine DNA methyltransferase.
<3>J. Mol. Biol.
<4>269
<5>494-504
<6>1997
<7>Methylation spreading, which involves a propensity for the mammalian
DNA-(cytosine-5)-methyltransferase to de novo methylate cytosine-guanine dinucleotides (CpGs)
near pre-existing 5-methylcytosine bases, has been implicated in the control of numerous
biological processes.  We have assessed methylation spreading by the murine DNA
methyltransferase in vitro using synthetic copolymers and oligonucleotides which differ only
in their methylation state.  Double-stranded oligonucleotides were found to undergo higher
levels of de novo methylation overall than other identical single-stranded oligonucleotides.
This difference reflects the greater number of de novo methylatable cytosine bases in
double-stranded than single-stranded sequences.  All tested oligonucleotides containing
pre-existing 5-methyl-cytosine(s) were de novo methylated at several fold the rates of
non-methylated controls.  No mammalian proteins besides the DNA methyltransferase were
required for this observed enhancement of de novo methylation.  Studies using oligonucleotides
differing in patterns of pre-methylation showed that methylation spreading can be initiated by
hemimethylated or duplex methylated CpGs indicating that recognition of 5-methylcytosine by
the enzyme is sufficient to stimulate methylation spreading. Double and single-stranded
oligonucleotides with several bases between CpGs underwent considerably more de novo
methylation per CpG than sequences containing sequential uninterrupted methylatable sites.
Spacing preferences by the DNA methyltransferase were also observed in hemimethylated
oligonucleotides, suggesting that this is a general property of the enzyme.  Although
methylation spreading outside of CpG dinucleotides was relatively rare, single-stranded DNA
incurred higher levels of de novo methylation at sites other than CpG as compared to
double-stranded DNA.  This indicates less specificity of methylation spreading in
single-stranded sequences.  Finally, enhanced de novo methylation in the presence of fully
methylated CpG sites in double-stranded oligonucleotides was not as high as the rates of
methylation of hemimethylated CpGs in otherwise identical oligonucleotides.  These studies
provide further elucidation of the mechanisms and regulation of the methylation spreading
process and its potential role in the biological processes it influences.

<>

<1>Tolstoshev, C.M., Blakesley, R.W.
<2>RSITE: a computer program to predict the recognition sequence of a restriction enzyme.
<3>Nucleic Acids Res.
<4>10
<5>1-17
<6>1982
<7>A computer program (RSITE) was developed which predicts the recognition
sequence of a restriction endonuclease.  The sizes of fragments experimentally
determined on cleavage of a DNA of known sequence were input.  Possible
recognition sequences producing fragments of sizes matching those determined
empirically were printed out.  The program faithfully predicted the specificity
of restriction enzymes of known recognition sequence and also determined the
recognition sequence of a new restriction enzyme from Haemophilus influenzae GU
(HinGUII).

<>

<1>Tomaschewski, J., Ruger, W.
<2>Nucleotide sequence and primary structures of gene products coded for by the T4 genome between map positions 48.266 kb and 39.166 kb.
<3>Nucleic Acids Res.
<4>15
<5>3632-3633
<6>1987
<7>This sequence comprises 19 open reading frames (orfs) including T4 genes 49 and nrdC which
were identified. The nomenclature of the orfs within this sequence is in agreement with the T4
genomic map.

<>

<1>Tomassini, J., Roychoudhury, R., Wu, R., Roberts, R.J.
<2>Recognition sequence of restriction endonuclease KpnI from Klebsiella pneumoniae.
<3>Nucleic Acids Res.
<4>5
<5>4055-4064
<6>1978
<7>We have determined the recognition sequence of the restriction endonuclease
KpnI, previously isolated from Klebsiella pneumoniae.  The enzyme cleaves the
twofold rotationally symmetric sequence 5'-G-G-T-A-C-^C-3' 3'-C-^C-A-T-G-G-5'
at the positions indicated by the arrows, producing 3' protruding cohesive
ends, four nucleotides in length.  The specific cleavage site was unambiguously
deduced using both 3' and 5' end analysis of KpnI generated restriction
fragments of simian-virus 40 (SV40) DNA (1 site), adenovirus-2 (Ad-2) DNA (8
sites), and a plasmid (pCRI) DNA (2 sites).

<>

<1>Tomb, J.-F. et al.
<2>The complete genome sequence of the gastric pathogen Helicobacter pylori.
<3>Nature
<4>388
<5>539-547
<6>1997
<7>Helicobacter pylori, strain 26695, has a circular genome of 1,667,867 base pairs and 1,590
predicted coding sequences.  Sequence analysis indicates that H. pylori has well-developed
systems for motility, for scavenging iron, and for DNA restriction and modification.  Many
putative adhesins, lipoproteins and other outer membrane proteins were identified,
underscoring the potential complexity of host-pathogen interaction.  Based on the large number
of sequence-related genes encoding outer membrane proteins and the presence of homopolymeric
racts and dinucleotide repeats in coding sequences, H. pylori, like several other mucosal
pathogens, probably uses recombination and slipped-strand mispairing within repeats as
mechanisms for antigenic variation and adaptive evolution.  Consistent with its restricted
niche, H. pylori has a few regulatory networks, and a limited metabolic repertoire and
biosynthetic capacity.  Its survival in acid conditions depends, in part, on its ability to
establish a positive inside-membrane potential in low pH.

<>

<1>Tomilov, V.N., Chernukhin, V.A., Abdurashitov, M.A., Gonchar, D.A., Degtyarev, S.K.
<2>Cleavage of mammalian chromosomal DNA by restriction enzymes in silico.
<3>FEBS J.
<4>274
<5>73
<6>2007
<7>A theoretical method to simulate the digestion patterns of mammalian chromosomal DNA cleavage
by restriction endonucleases was proposed. New software for long mammalian DNAs analysis using
routine personal computers was developed. This computational technique includes short DNA
sequences searching, DNA cleavage simulation, data treatment and verification. Recently
published primary structures of mammalian genomes (Rattus norvegicus, Mus musculus and Homo
sapiens) were presented like databases and the analysis of short nucleotide sequences
distribution in the corresponding genomes was performed. Computational DNA cleavage of genomes
within the nucleotides sequences 5¢-GGCC-3¢, 5¢GATC-3¢, 5¢-CC(A/T)GG-3¢ and 5¢-CCGG-3¢, which
are the recognition sites of well known restriction endonucleases, was carried out and the
diagrams of chromosomal DNA fragments distribution were obtained. Experiments on the
chromosomal DNA digestion by corresponding restriction endonucleases were undertaken. The
comparison of
computational diagrams and results of chromosomal DNA cleavage was done and a high accordance
of theoretical and experimental data was shown.

<>

<1>Tomilova, J.E., Chernukhin, V.A., Degtyarev, S.K.
<2>Dependence of site-specific endonuclease GlaI activity on quantity and location of methylcytosines in the recognition sequence 5'-GCGC-3'.
<3>Ovchinnikov Bull. Biotechnol. Phys. Chem. Biol.
<4>2
<5>30-39
<6>2006
<7>The activity dependence of site-specific endonuclease GlaI that recognizes and hydrolyzes only
methylated DNA sequence 5'-GCGC-3' on the quantity and location of 5-methylcytosines in
enzyme's recognition sequence has been studied. A significant DNA cleavage has been observed
for oligonucleotides duplexes containing four and three 5-methylcytosines or two internal
modified bases. The cleavage efficiency is maximal for DNA duplex with four 5-methylcytosine
and decreases when a number of methylated bases are lower. GlaI hydrolyzes recognition
sequences with 5-methylcytosines but not with N4-methylcytosines.

<>

<1>Tomilova, J.E., Kuznetsov, V.V., Abdurashitov, M.A., Netesova, N.A., Degtyarev, S.K.
<2>Recombinant DNA-methyltransferase M1.Bst19I from Bacillus stearothermophilus 19: Purification, properties, and amino acid sequence analysis.
<3>Mol. Biol. (Mosk)
<4>44
<5>606-615
<6>2010
<7>The M1.Bst19I DNA-methyltransferase gene from restriction-modification system Bst19I
(recognition sequence 5'-GCATC-3') in Bacillus
stearothermophilus 19 has been cloned in the expressing vector pJW that
carries a tandem of thermo inducible promoters P (R) /P (L) from phage
lambda. Highly purified enzyme has been isolated by chromatography on
various resins from Escherichia coli cells where it is accumulated in a
soluble form. The study of M1.Bst19I properties has revealed that the
enzyme has a temperature optimum at 50A degrees C and demonstrates
maximal activity at pH 8.0. M1.Bst19I modifies adenine in sequence
5'-GCATC-3'. Kinetic parameters of M1.Bst19I DNA methylation reaction
have been determined as follows: Km for lambda DNA is 0.68 +/- 0.07 mu
M, Km for S-adenosyl-L-methionine is 2.02 +/- 0.31 mu M. Catalytical
constant (k (cat)) is 1.8 +/- 0.05 min(-1). Comparative analysis of
Target Recognition Domain amino acid sequences for M1.Bst19I and other
alpha-N6-DNA-methyltransferases has allowed us to suggest the presence
of two types of the enzymes containing ATG or ATC triplets in the
recognition sequence.

<>

<1>Tomilova, Y.E., Abdurashitov, M.A., Golikova, L.N., Netesova, N.A., Degtyarev, S.K.
<2>Cloning of the genes for DNA methyltransferases of the SfaNI and Bst19I restriction-modification systems and primary structure analysis of  protein products.
<3>Mol. Biol. (Mosk)
<4>38
<5>850-856
<6>2004
<7>The Streptococcus faecalis ND547 and Bacillus stearothermophilus 19 genes that code for DNA
methyltransferases (MTases, M.) of
restriction-modification (RM) systems with the same recognition
sequence, 5'-GCATC-3' were cloned and sequenced. The Bst19I RM system
includes two MTases, M1.Bst19I and M2.Bst19I. The SfaNI RM system has
only one MTase, M.SfaNI, whose N and C domains are homologous to
M2.Bst19I and M1.Bst19I, respectively. Both M1.Bst19I and M2.Bst19I and
the two domains of M.SfaNI contain conserved elements, which are
arranged in the order characteristic of class alpha N6-adenine MTases.
The enzymes of the SfaNI and Bst19I RM systems proved to be highly
homologous to their FokI and BstF5I counterparts, which was explained
by the presence of the common tetranucleotide 5'-GATG-3' in their
recognition sites. Based on sequence homology, the spatial arrangement
of highly conserved amino acid residues was determined using the known
three-dimensional model of M.DpnIIA, which belongs to the same MTase
class.

<>

<1>Tomilova, Y.E., Dedkov, V.S., Shinkarenko, N.M., Popichenko, D.V., Degtyarev, S.K.
<2>Restriction endonuclease Bst19I from Bacillus stearothermophilus recognizing the 5'-GCATC-3' DNA sequence.
<3>Biotekhnologiya
<4>6
<5>17-20
<6>2002
<7>A Bacillus stearothermophilus 19 strain, a producer of a new Bst19I restriction endonuclease
has been isolated. The indicated enzyme was
isolated and its characteristics including the substrate specificity
and site of DNA restriction were studied. The Bst19I restriction
endonuclease was identified as a heteroschizomer of SfaNI restrictase
who recognizes the 5'-GCATC-3' DNA sequence and cleaves DNA at the
distance of 4 and 6 nucleotides from the site of recognition in upper
and lower chains, respectively.

<>

<1>Tomita, M., Toyama, K., Hiramatsu, A., Sho, K., Takahashi, T.
<2>Bacteriophage-resistant lactic acid bacteria and bacteriophage DNA restriction enzyme.
<3>Japanese Patent Office
<4>JP 2000236872 A
<5>11
<6>2000
<7>
<>

<1>Tomkins, J.P., Wood, T.C., Stacey, M.G., Loh, J.T., Judd, A., Goicoechea, J.L., Stacey, G., Sadowsky, M.J., Wing, R.A.
<2>A marker-dense, sequence-ready map of the Bradyrhizobium japonicum genome.
<3>Genome Res.
<4>11
<5>1434-1440
<6>2001
<7>Bacterial artificial chromosome (BAC) clones are effective mapping and
sequencing reagents for use with a wide variety of small and large
genomes. This report describes the development of a physical framework for
the genome of Bradyrhizobium japonicum, the nitrogen-fixing symbiont of
soybean. A BAC library for B. japonicum was constructed that provides a
77-fold genome coverage based on an estimated genome size of 8.7 Mb. The
library contains 4608 clones with an average insert size of 146 kb. To
generate a physical map, the entire library was fingerprinted with
HindIII, and the fingerprinted clones were assembled into contigs using
the software (; Sanger Centre, UK). The analysis placed 3410 clones in six
large contigs. The ends of 1152 BAC inserts were sequenced to generate a
sequence-tagged connector (STC) framework. To join and orient the contigs,
high-density BAC colony filters were probed with 41 known gene probes and
17 end sequences from contig boundaries. STC sequences were searched
against the public databases using and algorithms. Query results allowed
the identification of 113 high probability matches with putative
functional identities that were placed on the physical map. Combined with
the hybridization data, a high-resolution physical map with 194 positioned
markers represented in two large contigs was developed, providing a marker
every 45 kb. Of these markers, 177 are known or putative B. japonicum
genes. Additionally, 1338 significant results (E < 10(-4)) were manually
sorted by function to produce a functionally categorized database of
relevant B. japonicum STC sequences that can also be traced to specific
locations in the physical map.

<>

<1>Tompa, R., McCallum, C.M., Delrow, J., Henikoff, J.G., van Steensel, B., Henikoff, S.
<2>Genome-Wide Profiling of DNA Methylation Reveals Transposon Targets of CHROMOMETHYLASE3.
<3>Curr. Biol.
<4>12
<5>65-68
<6>2002
<7>DNA methylation has been implicated in a variety of epigenetic processes, and abnormal
methylation patterns have been seen in tumors. Analysis of methylation patterns has
traditionally been conducted either by using Southern analysis after cleavage with
methyl-sensitive restriction endonucleases or by bisulfite sequencing. However, neither method
is practical for analyzing more than a few genes. Here, we describe a simple technique for
genome-wide mapping of DNA methylation patterns. Fragmentation by a methyl-sensitive
restriction endonuclease is followed by size fractionation and hybridization to microarrays.
We demonstrate the utility of this method by characterizing methylation patterns in
Arabidopsis methylation mutants. This analysis reveals that CHROMOMETHYLASE3 (CMT3), which was
previously shown to maintain CpXpG methylation, preferentially methylates transposons, even
when they are present as single copies within the genome. Methylation profiling has potential
applications in disease research and diagnostic screening.

<>

<1>Tompkins, T.A., Barreau, G., Broadbent, J.R.
<2>Complete Genome Sequence of Lactobacillus helveticus R0052, a Commercial Probiotic Strain.
<3>J. Bacteriol.
<4>194
<5>6349
<6>2012
<7>Lactobacillus helveticus R0052 is a commercially available strain that is widely  used in
probiotic preparations. The genome sequence consisted of 2,129,425 bases.
Comparative analysis showed that it was unique among L. helveticus strains in
that it contained genes encoding mucus-binding proteins similar to those found in
Lactobacillus acidophilus.

<>

<1>Tompkins, T.A., Barreau, G., de Carvalho, V.G.
<2>Draft Genome Sequence of Probiotic Strain Lactobacillus rhamnosus R0011.
<3>J. Bacteriol.
<4>194
<5>902
<6>2012
<7>Lactobacillus rhamnosus R0011 is a commercially available probiotic that is widely used in
human dietary supplements and pharmaceutical products.
We prepared a draft genome sequence consisting of 10 contigs totaling
2,900,620 bases and a G+C content of 46.7% for this strain.

<>

<1>Tong, H., Shang, N., Liu, L., Wang, X., Cai, J., Dong, X.
<2>Complete Genome Sequence of an Oral Commensal, Streptococcus oligofermentans Strain AS 1.3089.
<3>Genome Announcements
<4>1
<5>e00353-13
<6>2013
<7>Streptococcus oligofermentans, an oral commensal, inhibits the growth of the dental caries
pathogen Streptococcus mutans by producing large amounts of
hydrogen peroxide. Therefore, it can be a potential probiotic for oral health.
Here we report the complete genome sequence of S. oligofermentans strain AS
1.3089.

<>

<1>Tong, Y.J., Ji, X.J., Liu, L.G., Shen, M.Q., Huang, H.
<2>Genome Sequence of Paenibacillus polymyxa ATCC 12321, a Promising Strain for Optically Active (R,R)-2,3-Butanediol Production.
<3>Genome Announcements
<4>1
<5>e00572-13
<6>2013
<7>Paenibacillus polymyxa is a potential strain for (R,R)-2,3-butanediol production. Here, we
report an annotated draft genome sequence of P. polymyxa strain ATCC
12321, which contains 4,429 protein-coding genes and 49 structural RNAs. This
genome sequence provides a genetic basis for a better understanding of the
mechanism for the accumulation of highly optically active (R,R)-2,3-butanediol.

<>

<1>Too, P.H., Zhu, Z., Chan, S.H., Xu, S.Y.
<2>Engineering Nt.BtsCI and Nb.BtsCI nicking enzymes and applications in generating long overhangs.
<3>Nucleic Acids Res.
<4>38
<5>1294-1303
<6>2010
<7>Type IIS restriction endonuclease BtsCI (GGATG 2/0) is a neoschizomer of FokI (GGATG 9/13) and
cleaves closer to the recognition sequence. Although
M.BtsCI shows 62% amino acid sequence identity to M.FokI, BtsCI and FokI
restriction endonucleases do not share significant amino acid sequence
similarity. BtsCI belongs to a group of Type IIS restriction
endonucleases, BsmI, Mva1269I and BsrI, that carry two different catalytic
sites in a single polypeptide. By inactivating one of the catalytic sites
through mutagenesis, we have generated nicking variants of BtsCI that
specifically nick the bottom-strand or the top-strand of the target site.
By treating target DNA sequentially with the appropriate combinations of
FokI and BtsCI nicking variants, we are able to generate long overhangs
suitable for fluorescent labeling through end-filling or other techniques
based on annealing of complementary DNA sequences.

<>

<1>Toothman, P.
<2>Restriction alleviation by bacteriophages lambda and lambda reverse.
<3>J. Virol.
<4>38
<5>621-631
<6>1981
<7>Deletion analysis indicated that the phage lambda restriction alleviation
gene(s) ral resides between the cIII and N genes.  The Ral+ phenotype was expressed only
when lambda-ral+ carried a modification such that it was resistant to restriction by the host
specificity system.  Under these conditions, Ral function protected superinfecting
unmodified phages from restriction by EcoK or EcoB but not from restriction by EcoP1.
Ral-protected phage DNA was not concomitantly K and B modified, but rather received
only the modification specified by the system of the restricting host.  Possible mechanisms
for Ral action are discussed.  Of the other lambdoid phages tested, the hybrid phage
lambda-rev had Ral activity, whereas omega-80vir and one lambda-P22 hybrid did not.
The restriction alleviation activity of lambda-rev called Lar, may be the same as the activity
expressed in sbcA- strains of Escherichia coli, but it was functionally separable from
exonuclease VIII activity (the product of the recE gene), which is also expressed in sbcA-
strains.

<>

<1>Topal, M.D.
<2>Enzymes that cleave and ligate DNA.
<3>International Patent Office
<4>WO 9622365
<5>
<6>1996
<7>Enzymes useful for carrying out the cleavage and ligation of DNA are disclosed that: (a)
specifically bind to a DNA recognition site, (b) cleave the DNA on binding to the DNA
recognition site to form a cleaved DNA having a cleaved end segment, (c) form a covalent
intermediate with the cleaved DNA, (d) ligate the cleaved end segment with another end segment
to form a ligated DNA, and (e) release the DNA from the covalent intermediate upon the
formation of the ligated DNA.  Nucleotides encoding the enzymes and methods of using the
enzymes in vitro and in vivo are also disclosed.

<>

<1>Topal, M.D., Conrad, M.
<2>Changing endonuclease EcoRII Tyr308 to Phe abolishes cleavage but not recognition: possible homology with the Int-family of recombinases.
<3>Nucleic Acids Res.
<4>21
<5>2599-2603
<6>1993
<7>Endonuclease EcoRII is one of a group of type II restriction enzymes, including NaeI, NarI,
BspMI, HpaII, and SacII, that require binding of an enhancer sequence to cleave DNA.
Comparison of the EcoRII amino-acid sequence with the amino-acid consensus motifs that
differentiate between recombinase families uncovered similarity between a 29 amino-acid
sequence in the carboxyl end of EcoRII and the motif defining the integrase family of
recombinases. This similarity implied that EcoRII tyrosine 308 should be involved in
catalyzing hydrolysis of the scissile bond. Site-directed mutagenesis was used to mutate
Tyr308 to Phe. The phenylalanine-substituted enzyme could not cleave T5 DNA under conditions
in which wild-type enzyme completely cleaved this DNA. The Tyr308 to Phe mutation abolished
cleavage activity but not specific binding to DNA. No evidence was found for the existence
during the cleavage reaction of a covalent linkage between Tyr308 and DNA.

<>

<1>Topal, M.D., Conrad, M.J.
<2>Method of cleaving DNA.
<3>US Patent Office
<4>US 5248600
<5>
<6>1993
<7>A method of cleaving substrate DNA with a restriction enzyme, wherein the substrate DNA is
resistant to cleavage by the restriction enzyme, is disclosed. The method comprises
co-incubating the substrate DNA and the restriction enzyme with an activating DNA sequence.
The activating sequence comprises an oligonucleotide comprised of the restriction enzyme
recognition site and cleavage permissible flanking sequences joined directly to both the 5'
and 3' ends of the recognition site. Exemplary restriction enzymes which may be used in
practicing the present invention include NaeI, BspMI, HpaII, NarI and SacII.

<>

<1>Topal, M.D., Thresher, R.J., Conrad, M., Griffith, J.
<2>NaeI endonuclease binding to pBR322 DNA induces looping.
<3>Biochemistry
<4>30
<5>2006-2010
<6>1991
<7>Previous work has demonstrated the existence of both resistant and cleavable
NaeI sites.  Cleavable sites introduced on exogenous DNA can act in trans to
increase the catalysis of NaeI endonuclease cleavage at resistant sites without
affecting the apparent binding affinity of the enzyme for the resistant site
[Conrad, M. & Topal, M.D. (1988) Proc. Natl. Acad. Sci. 86, 9707-9711].  This
activation suggests allosteric regulation of NaeI cleavage by distant cis- and
trans-acting sites in DNAs containing both resistant and cleavable sites.
Plasmid pBR322 contains four NaeI sites, at least one of which is resistant to
cleavage.  Electron microscopy is used here to demonstrate that NaeI
endonuclease simultaneously binds to multiple recognition sites in pBR322 DNA
to form loops with NaeI protein bound at the loop's base.  The maximum number
of loops formed with a common base suggests four binding sites per enzyme
molecule.  Looping was inhibited by addition of enzyme-saturating amounts of
double-strand oligonucleotide without the NaeI site had no effect.  The number
of loops seen was not above background when double-stranded M13 DNA, which
contains only a single NaeI recognition site, was used as substrate.

<>

<1>Topin, A.N., Gritsenko, O.M., Brevnov, M.G., Gromova, E.S., Korshunova, G.A.
<2>Synthesis of a new photo-cross-linking nucleoside analogue containing an Aryl(trifluoromethyl)Diazirine group: Application for EcoRII and MvaI restriction-modification enzymes.
<3>Nucleosides and Nucleotides
<4>17
<5>1163-1175
<6>1998
<7>A new photo-cross-linking dU analog,
5-[4-(3-(trifluoromethyl)-3H-diazirin-3-yl)phenyl]-2'deoxyuridine, was synthesized and
incorporated into the recognition site of EcoRII and MvaI restriction-modification enzymes.
The resulting base-modified 14-mer substrate was tested for cross-linking to these enzymes.
Cross-linking is effected by irradiation of the enzyme-substrate complexes as 366 nm.

<>

<1>Toro, N., Martinez-Abarca, F., Nisa-Martinez, R.
<2>Complete Genome Sequence of the RmInt1 Group II Intronless Sinorhizobium meliloti Strain RMO17.
<3>Genome Announcements
<4>2
<5>e01001-14
<6>2014
<7>We report the complete genome sequence of the RmInt1 group II intronless Sinorhizobium
meliloti strain RMO17 isolated from Medicago orbicularis nodules
from Spanish soil. The genome consists of 6.73 Mb distributed between a single
chromosome and two megaplasmids (the chromid pSymB and pSymA).

<>

<1>Torosian, M.V., Shishkova, O.V., Rabinkova, E.V.
<2>Influence of recB and recA mutations on bacteriophage restriction by different restriction modification plasmid systems of E. coli.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>36-41
<6>1988
<7>The effect of recB and recA mutations on lambda vir and P1 vir restriction by
different restriction-modification plasmid systems of E. coli was studied.  It
was shown that the effect of RI plasmid coded restriction-modification in E.
coli K12 and E. coli B strains and pJA4620 plasmid coded restriction in E. coli
K12 is observed only in RecB+ strain.  Phenomenon of restriction-modification
determined by R124, R245 plasmids does not depend of recB mutation.  Effect of
recA mutation has not been found in cultures harbouring R1, R245, R124 pJA4620
plasmids.

<>

<1>Torosyan, M.V., Rainkova, E.V., Shishkova, L.A., Naumova, L.A., Fradkin, G.E.
<2>Phenomenon of restriction alleviation in Escherichia coli strains.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>10
<5>37-40
<6>1987
<7>The alleviation of foreign DNA restriction after treatment of cells by UV and gamma-rays or
ocr+-gene product of T3, T7 phages has been studied.  Both UV and gamma-radiation were shown
to induce restriction alleviation of unmodified phage.  The results of restriction alleviation
caused by T3 and T7 ocr+-gene products have been evaluated by F-lac+ transfer efficiencies in
heterologic crosses and plating of unmodified phage lambda.  The phenomenon of restriction
alleviation was established to depend on the strain used and to be expressed mostly in AB157
and its derivatives.

<>

<1>Torreblanca, J., Casadesus, J.
<2>DNA adenine methylase mutants of Salmonella typhimurium and a novel Dam-regulated locus.
<3>Genetics
<4>144
<5>15-26
<6>1996
<7>Mutants of Salmonella typhimurium lacking DNA adenine methylase were isolated; they include
insertion and deletion alleles.  The dam locus maps at 75 min between cysG and aroB, similar
to the Escherichia coli dam gene.  Dam- mutants of S. typhimurium resemble those of E. coli in
the following phenotypes: (1) increased spontaneous mutations, (2) moderate SOS induction, (3)
enhancement of duplication segregation, (4) inviability of dam recA and dam recB mutants, and
(5) suppression of the inviability of the dam recA and dam recB combinations by mutations that
eliminate mismatch repair.  However, differences between S. typhimurium and E. coli dam
mutants are also found: (1) S. typhimurium dam mutants do not show increased UV sensitivity,
suggesting that methyl-directed mismatch repair does not participate in the repair of
UV-induced DNA damage in Salmonella.  (2) S. typhimurium dam recJ mutants are viable,
suggesting that the Salmonella RecJ function does not participate in the repair of DNA strand
breaks formed in the absence of Dam methylation.  We also describe a genetic screen for
detecting novel genes regulated by Dam methylation and a locus repressed by Dam methylation in
the S. typhimurium virulence (or "cryptic") plasmid.

<>

<1>Torreblanca, J., Marques, S., Casadesus, J.
<2>Synthesis of FinP RNA by plasmids F and pSLT is regulated by DNA adenine methylation.
<3>Genetics
<4>151
<5>31-45
<6>1999
<7>DNA adenine methylase mutants of Salmonella typhimurium contain reduced amounts of FinP, an
antisense RNA encoded by the virulence plasmid pSLT.  Lowered FinP levels are detected in both
Dam- FinO+ and Dam- FinO- backgrounds, suggesting that Dam methylation regulates FinP
production rather than FinP half-life.  Reduced amounts of F-encoded FinP RNA are likewise
found in Dam- mutants of Escherichia coli.  A consequence of FinP RNA scarcity in the absence
of DNA adenine methylation is that Dam- mutants of both S. typhimurium and E. coli show
elevated levels of F plasmid transfer.  Inhibition of F fertility by the S. typhimurium
virulence plasmid is also impaired in a Dam- background.

<>

<1>Torres, B., Jaenecke, S., Timmis, K.N., Garcia, J.L., Diaz, E.
<2>A gene containment strategy based on a restriction-modification system.
<3>Environ. Microbiol.
<4>2
<5>555-563
<6>2000
<7>Engineering barriers to the spread of specific genes are of great interest both to increase
the predictability of recombinant microorganisms used for environmental applications and to
study the role of gene transfer in the adaptation of microbial communities to changing
environments. We report here a new gene containment circuit based on a toxin-antidote pair
that targets the cell DNA, i.e. the type II EcoRI restriction-modification system. The set-up
involved linkage of the ecoRIR lethal gene encoding the EcoRI endonuclease (toxin) to the
contained character in a plasmid and chromosomal insertion of the ecoRIM gene encoding the
cognate EcoRI methylase (antidote) that protects the target DNA from restriction. Transfer of
the contained character to a recipient cell lacking the antidote caused EcoRI-mediated
chromosomal breaks, leading to cell death, thereby preventing gene spread. Using
transformation and conjugation as mechanisms of DNA transfer and different environmentally
relevant bacteria as recipients, we have shown that the potentially universal EcoRI-based
containment system decreases gene transfer frequencies by more than four orders of magnitude.
Analyses of the survivors escaping killing revealed a number of possible inactivation
mechanisms.

<>

<1>Torres, B., Jaenecke, S., Timmis, K.N., Garcia, J.L., Diaz, E.
<2>A dual lethal system to enhance containment of recombinant micro-organisms.
<3>Microbiology
<4>149
<5>3595-3601
<6>2003
<7>Active containment systems based on the controlled expression of a lethal gene are designed to
increase containment of recombinant micro-organisms
used for environmental applications. A major drawback in containment is
the existence of mutations that generate surviving cells that cease to
respond to the toxic effect of the lethal function. In this work the
authors have developed for the first time a strategy to reduce the problem
of mutations and increase the efficiency of containment based on the
combination of two lethal functions acting on different cellular targets
of major concern in containment, DNA and RNA, and whose expression is
under control of different regulatory signals. To engineer the dual gene
containment circuit, two toxin-antitoxin pairs, i.e. the colicin
E3-immunity E3 and the EcoRI restriction-modification systems, were
combined. The genes encoding the immunity E3 and the EcoRI
methyltransferase proteins (antitoxins) were stably inserted into the
chromosome of the host cell, whereas the broad-host-range lethal genes
encoding the colicin E3 RNase and the EcoRI restriction endonuclease
(toxins) were flanking the contained trait in a plasmid. This dual lethal
cassette decreased gene transfer frequencies, through killing of the
recipient cells, by eight orders of magnitude, which provides experimental
evidence that the anticipated containment level due to the combination of
single containment systems is generally achieved. Survivors that escaped
killing were analysed and the mutational events involved were
characterized.

<>

<1>Torres, T.G., Lozano, L., Gonzalez, V., Bustos, P., Romero, D., Brom, S.
<2>Draft Genome Sequence of the Bean-Nodulating Sinorhizobium fredii Strain GR64.
<3>J. Bacteriol.
<4>194
<5>6978
<6>2012
<7>Sinorhizobium fredii GR64 is a peculiar strain that is able to effectively nodulate bean but
not soybean, the common host of S. fredii. Here we present the
draft genome of S. fredii GR64. This information will contribute to a better
understanding of the symbiotic rhizobium-plant interaction and of rhizobial
evolution.

<>

<1>Toth, A., Barna, T., Nagy, I., Horvath, B., Nagy, I., Tancsics, A., Kriszt, B., Baka, E., Fekete, C., Kukolya, J.
<2>Draft Genome Sequence of the Lignocellulose Decomposer Thermobifida fusca Strain  TM51.
<3>Genome Announcements
<4>1
<5>e00482-13
<6>2013
<7>Here, we present the complete genome sequence of Thermobifida fusca strain TM51,  which was
isolated from the hot upper layer of a compost pile in Hungary. T.
fusca TM51 is a thermotolerant, aerobic actinomycete with outstanding
lignocellulose-decomposing activity.

<>

<1>Toth, E., Huszar, K., Bencsura, P., Kulcsar, P.I., Vodicska, B., Nyeste, A., Welker, Z., Toth, S., Welker, E.
<2>Restriction enzyme body doubles and PCR cloning: on the general use of type IIs restriction enzymes for cloning.
<3>PLoS ONE
<4>9
<5>e90896
<6>2014
<7>The procedure described here allows the cloning of PCR fragments containing a recognition site
of the restriction endonuclease (Type IIP) used for cloning in
the sequence of the insert. A Type IIS endonuclease - a Body Double of the Type
IIP enzyme - is used to generate the same protruding palindrome. Thus, the insert
can be cloned to the Type IIP site of the vector without digesting the PCR
product with the same Type IIP enzyme. We achieve this by incorporating the
recognition site of a Type IIS restriction enzyme that cleaves the DNA outside of
its recognition site in the PCR primer in such a way that the cutting positions
straddle the desired overhang sequence. Digestion of the PCR product by the Body
Double generates the required overhang. Hitherto the use of Type IIS restriction
enzymes in cloning reactions has only been used for special applications, the
approach presented here makes Type IIS enzymes as useful as Type IIP enzymes for
general cloning purposes. To assist in finding Body Double enzymes, we summarised
the available Type IIS enzymes which are potentially useful for Body Double
cloning and created an online program
(http://group.szbk.u-szeged.hu/welkergr/body_double/index.html) for the selection
of suitable Body Double enzymes and the design of the appropriate primers.

<>

<1>Toth, J., van Aelst, K., Salmons, H., Szczelkun, M.D.
<2>Dissociation from DNA of Type III Restriction-Modification enzymes during helicase-dependent motion and following endonuclease activity.
<3>Nucleic Acids Res.
<4>40
<5>6752-6764
<6>2012
<7>DNA cleavage by the Type III Restriction-Modification (RM) enzymes requires the binding of a
pair of RM enzymes at two distant, inversely orientated recognition
sequences followed by helicase-catalysed ATP hydrolysis and long-range
communication. Here we addressed the dissociation from DNA of these enzymes at
two stages: during long-range communication and following DNA cleavage. First, we
demonstrated that a communicating species can be trapped in a DNA domain without
a recognition site, with a non-specific DNA association lifetime of approximately
200 s. If free DNA ends were present the lifetime became too short to measure,
confirming that ends accelerate dissociation. Secondly, we observed that Type III
RM enzymes can dissociate upon DNA cleavage and go on to cleave further DNA
molecules (they can 'turnover', albeit inefficiently). The relationship between
the observed cleavage rate and enzyme concentration indicated independent binding
of each site and a requirement for simultaneous interaction of at least two
enzymes per DNA to achieve cleavage. In light of various mechanisms for
helicase-driven motion on DNA, we suggest these results are most consistent with
a thermally driven random 1D search model (i.e. 'DNA sliding').

<>

<1>Tothova, T., Godany, A., Javorsky, P., Pristas, P.
<2>Production of SacI and SacII isoschizomers by soil streptomycetes.
<3>Biologia (Bratisl)
<4>62
<5>381-385
<6>2007
<7>Five fresh soil Streptomyces spp. strains were isolated, phylogenetically characterized on the
basis of 16S rDNA sequences and
analyzed for the presence of restriction modification systems. Three
type II site-specific endonucleases were detected and partially
purified. Two isolated enzymes were isoschizomers of SacI restriction
endonuclease recognizing 5'-GAGCTC-3' sequence; the third one
recognised 5'-CCGCGG-3' sequence and it was an isoschizomer of SacII.
SacII like modification was observed in other two isolates having no
detectable restriction activity. The lack of correlation between
restriction and modification phenotypes and phylogenetic classification
of the isolates indicates efficient gene transfer mechanism in the
Streptomyces genus.

<>

<1>Totsika, M., Beatson, S.A., Sarkar, S., Phan, M.D., Petty, N.K., Bachmann, N., Szubert, M., Sidjabat, H.E., Paterson, D.L., Upton, M., Schembri, M.A.
<2>Insights into a multidrug resistant Escherichia coli pathogen of the globally disseminated ST131 lineage: genome analysis and virulence mechanisms.
<3>PLoS ONE
<4>6
<5>E26578
<6>2011
<7>Escherichia coli strains causing urinary tract infection (UTI) are increasingly
recognized as belonging to specific clones. E. coli clone O25b:H4-ST131 has
recently emerged globally as a leading multi-drug resistant pathogen causing
urinary tract and bloodstream infections in hospitals and the community. While
most molecular studies to date examine the mechanisms conferring multi-drug
resistance in E. coli ST131, relatively little is known about their virulence
potential. Here we examined E. coli ST131 clinical isolates from two
geographically diverse collections, one representing the major pathogenic
lineages causing UTI across the United Kingdom and a second representing UTI
isolates from patients presenting at two large hospitals in Australia. We
determined a draft genome sequence for one representative isolate, E. coli EC958,
which produced CTX-M-15 extended-spectrum beta-lactamase, CMY-23 type AmpC
cephalosporinase and was resistant to ciprofloxacin. Comparative genome analysis
indicated that EC958 encodes virulence genes commonly associated with
uropathogenic E. coli (UPEC). The genome sequence of EC958 revealed a transposon
insertion in the fimB gene encoding the activator of type 1 fimbriae, an
important UPEC bladder colonization factor. We identified the same fimB
transposon insertion in 59% of the ST131 UK isolates, as well as 71% of ST131
isolates from Australia, suggesting this mutation is common among E. coli ST131
strains. Insertional inactivation of fimB resulted in a phenotype resembling a
slower off-to-on switching for type 1 fimbriae. Type 1 fimbriae expression could
still be induced in fimB-null isolates; this correlated strongly with adherence
to and invasion of human bladder cells and bladder colonisation in a mouse UTI
model. We conclude that E. coli ST131 is a geographically widespread, antibiotic
resistant clone that has the capacity to produce numerous virulence factors
associated with UTI.

<>

<1>Touchon, M. et al.
<2>Organised genome dynamics in the Escherichia coli species results in highly diverse adaptive paths.
<3>PLoS Genet.
<4>5
<5>e1000344
<6>2009
<7>The Escherichia coli species represents one of the best-studied model organisms,  but also
encompasses a variety of commensal and pathogenic strains that diversify
by high rates of genetic change. We uniformly (re-) annotated the genomes of 20
commensal and pathogenic E. coli strains and one strain of E. fergusonii (the
closest E. coli related species), including seven that we sequenced to
completion. Within the approximately 18,000 families of orthologous genes, we
found approximately 2,000 common to all strains. Although recombination rates are
much higher than mutation rates, we show, both theoretically and using
phylogenetic inference, that this does not obscure the phylogenetic signal, which
places the B2 phylogenetic group and one group D strain at the basal position.
Based on this phylogeny, we inferred past evolutionary events of gain and loss of
genes, identifying functional classes under opposite selection pressures. We
found an important adaptive role for metabolism diversification within group B2
and Shigella strains, but identified few or no extraintestinal virulence-specific
genes, which could render difficult the development of a vaccine against
extraintestinal infections. Genome flux in E. coli is confined to a small number
of conserved positions in the chromosome, which most often are not associated
with integrases or tRNA genes. Core genes flanking some of these regions show
higher rates of recombination, suggesting that a gene, once acquired by a strain,
spreads within the species by homologous recombination at the flanking genes.
Finally, the genome's long-scale structure of recombination indicates lower
recombination rates, but not higher mutation rates, at the terminus of
replication. The ensuing effect of background selection and biased gene
conversion may thus explain why this region is A+T-rich and shows high sequence
divergence but low sequence polymorphism. Overall, despite a very high gene flow,
genes co-exist in an organised genome.

<>

<1>Touchon, M., Barbier, P., Bernardet, J.F., Loux, V., Vacherie, B., Barbe, V., Rocha, E., Duchaud, E.
<2>Complete genome sequence of the fish pathogen Flavobacterium branchiophilum.
<3>Appl. Environ. Microbiol.
<4>77
<5>7656-7662
<6>2011
<7>Members of the genus Flavobacterium occur in a variety of ecological niches and represent an
interesting diversity of lifestyles. Flavobacterium branchiophilum is the main causative agent
of bacterial gill disease, a severe condition affecting various cultured freshwater fish
species worldwide, in particular salmonids in Canada and Japan. We report here the complete
genome sequence of strain FL-15 isolated from a diseased sheatfish (Silurus glanis) in
Hungary. The analysis of the F. branchiophilum genome revealed putative mechanisms of
pathogenicity strikingly different from those of the other, closely related fish pathogen
Flavobacterium psychrophilum, including the first cholera-like toxin in a non-Proteobacteria
and a wealth of adhesins. The comparison with available genomes of other Flavobacterium
species revealed a small genome size, large differences in chromosome organization, and fewer
rRNA and tRNA genes, in line with its more fastidious growth. In addition, horizontal gene
transfer shaped the evolution of F. branchiophilum, as evidenced by its virulence factors,
genomic islands, and CRISPR (clustered regularly interspaced short palindromic repeats)
systems. Further functional analysis should help in the understanding of host-pathogen
interactions and in the development of rational diagnostic tools and control strategies in
fish farms.

<>

<1>Tovkach, F.I.
<2>A study of Erwinia carotovora phage resistance with the use of temperate bacteriophage ZF40.
<3>Mikrobiologiia
<4>71
<5>82-88
<6>2002
<7>The causes of the unique phage resistance of the pectinolytic phytopathogenic strains of
Erwinia carotovora were studied with the use of temperate bacteriophage ZF40. It was shown
that, in these bacteria, the bacteriophage-cell interaction can be substantially blocked at
the adsorption level. An adequate indicator for studying the temperate bacteriophages of
Erwinias was developed on the basis of mutants resistant to macromolecular bacteriocins.
Various restriction-modification systems, which influence cell resistance to bacteriophages,
were revealed for the first time in E. carotovora. The phage resistance was shown to be
determined by the wide occurrence of homoimmune temperate viruses in pectinolytic Erwinias.

<>

<1>Tovkach, F.I., Chervatyuk, N.V.
<2>Phage system for studying the restriction-modification system of Erwinia carotovora.
<3>Mikrobiol. Zh.
<4>68
<5>27-35
<6>2006
<7>A model for revealing and type identifying the systems of restriction-modification of
phytopathogenic bacterium Ewinia carotovora with the help of two virulent polyvalent phages-FE
44 and T7 is proposed in the work.  The joint use of the phages permits to identify both the
limitations of phages development on the part of RM-systems of three major types, and on the
part of the competing episomes.  The proposed system also allows to reveal other kinds of
limitations, in particular those connected with the adsorption of the phage particles.

<>

<1>Townson, S.A., Samuelson, J.C., Bao, Y., Xu, S.Y., Aggarwal, A.K.
<2>BstYI Bound to Noncognate DNA Reveals a "Hemispecific" Complex: Implications for DNA Scanning.
<3>Structure
<4>15
<5>449-459
<6>2007
<7>DNA recognition by proteins is essential for specific expression of genes in a living
organism. En route to a target DNA site, a protein will often
sample noncognate DNA sites through nonspecific protein-DNA interactions,
resulting in a variety of conformationally different binding states. We
present here the crystal structure of endonuclease BstYI bound to a
noncognate DNA. Surprisingly, the structure reveals the enzyme in a
"hemispecific" binding state on the pathway between nonspecific and
specific recognition. A single base pair change in the DNA abolishes
binding of only one monomer, with the second monomer bound specifically.
We show that the enzyme binds essentially as a rigid body, and that one
end of the DNA is accommodated loosely in the binding cleft while the
other end is held tightly. Another intriguing feature of the structure is
Ser172, which has a dual role in establishing nonspecific and specific
contacts. Taken together, the structure provides a snapshot of an enzyme
in a "paused" intermediate state that may be part of a more general
mechanism of scanning DNA.

<>

<1>Townson, S.A., Samuelson, J.C., Vanamee, E.S., Edwards, T.A., Escalante, C.R., Xu, S.-Y., Aggarwal, A.K.
<2>Crystal structure of BstYI at 1.85 .ANG. resolution: A thermophilic restriction endonuclease with overlapping specificities to BamHI and  BglII.
<3>J. Mol. Biol.
<4>338
<5>725-733
<6>2004
<7>We report here the structure of BstYI, an "intermediate" type II restriction endonuclease with
overlapping sequence specificities to BamHI and BglII. BstYI, a thermophilic endonuclease,
recognizes and cleaves the degenerate hexanucleotide sequence 5'-RGATCY-3' (where R=A or G
and Y=C or T), cleaving DNA after the 5'-R on each strand to produce four-base (5')
staggered ends. The crystal structure of free BstYI was solved at 1.85A resolution by
multi-wavelength anomalous dispersion (MAD) phasing. Comparison with BamHI and BglII reveals a
strong structural consensus between all three enzymes mapping to the alpha/beta core domain
and residues involved in catalysis. Unexpectedly, BstYI also contains an additional "arm"
substructure outside of the core protein, which enables the enzyme to adopt a more compact,
intertwined dimer structure compared with BamHI and BglII. This arm substructure may underlie
the thermostability of BstYI. We identify putative DNA recognition residues and speculate as
to how this enzyme achieves a "relaxed" DNA specificity.

<>

<1>Townson, S.A., Samuelson, J.C., Xu, S.-Y., Aggarwal, A.K.
<2>Implications for switching restriction enzyme specificities from the structure of BstYI bound to a BglII DNA sequence.
<3>Structure
<4>13
<5>791-801
<6>2005
<7>The type II restriction endonuclease BstYI recognizes the degenerate sequence 5'-RGATCY-3'
(where R = A/G and Y = C/T), which overlaps with both BamHI (GGATCC) and BglII (AGATCT), and
thus raises the question of whether BstYI DNA recognition will be more BamHI-like or
BglII-like.  We present here the structure of BstYI bound to a cognate DNA sequence (AGATCT).
We find the complex to be more BglII-like with similarities mapping to DNA conformation,
domain organization, and residues involved in catalysis.  However, BstYI is unique in
containing an extended arm subdomain, and the mechanism of DNA capture has both BglII-like and
BamHI-like elements.  Further, DNA recognition is more minimal than BglII and BamHI, where
only two residues mediate recognition of the entire core sequence.  Taken together, the
structure reveals a mechanism of degenerate DNA recognition and offers insights into the
possibilities and limitations in altering specificities of closely related restriction
enzymes.

<>

<1>Toyobo, K.K.
<2>Recombinant plasmid replicable with Escherichia coli - incorporates chromosome DNA fragment containing BanIII restriction endonuclease gene from Bacillus aneurinolyticus.
<3>Japanese Patent Office
<4>JP 6416586
<5>
<6>1989
<7>Recombinant plasmid replicable with Escherichia coli, where in the plasmid is incorporated
chromosome DNA fragment containing BanIII restriction endonuclease gene origined from Bacillus
aneurinolyticus IAM 1077, is claimed. Escherichia genus of microorganism transformed with
recombinant plasmid incorporated chromosome DNA fragment containing BanIII restriction
endonuclease gene origined from Bacillus aneurinolyticus IAM 1077, is also claimed. BanIII
restriction endonuclease is produced by Escherichia genus of microorganism transformed with
recombinant plasmid incorporated with chromosome DNA fragment containing BanIII restriction
endonuclease gene origined from Bacillus aneurinolyticus IAM 1077, and taking out BanII
restriction endonuclease from culture mixture.

<>

<1>Toyoda, K., Nakamura, K., Kato, Y., Wada, A.
<2>Immobilization of DNA restriction endonuclease on nonporous carrier.
<3>Japanese Patent Office
<4>JP 01148186
<5>
<6>1989
<7>DNA restriction enzyme immobilized carrier is claimed whose character is that DNA restriction
enzyme is immobilized on the surface of a non-porous carrier.  Preferably, immobilized carrier
is an organic polymer, which has a functional group eg. polyvinyl alcohol,
polyhydroxy-methacrylate, cellulose, chitin, etc, silica gel, ceramic.  These carriers are
activated.  For activation, tosyl activation, cyanohalide activation, formyl activation, amino
activation, carboxyl activation are used.  As a restriction enzyme, AccI, BamHI, EcorI etc.
are used.  The enzyme is immobilized at 0-37 C.

<>

<1>Traglia, G., Vilacoba, E., Almuzara, M., Diana, L., Iriarte, A., Centron, D., Ramirez, M.S.
<2>Draft Genome Sequence of an Extensively Drug-Resistant Acinetobacter baumannii Indigo-Pigmented Strain.
<3>Genome Announcements
<4>2
<5>e01146-14
<6>2014
<7>Last year in 2013, we reported an outbreak due to indigo-pigmented Acinetobacter  baumannii
strains in a hospital from Buenos Aires, Argentina. Here, we present
the draft genome sequence of one of the strains (A. baumannii A33405) involved in
the outbreak. This isolate was categorized as extensively drug-resistant (XDR)
and harbors different genetic elements associated with horizontal genetic
transfer and multiple antibiotic resistances.

<>

<1>Tragut, V., Xiao, J., Bylina, E.J., Borthakur, D.
<2>Characterization of DNA restriction-modification systems in Spirulina platensis strain pacifica.
<3>J. Appl. Phycol.
<4>7
<5>561-564
<6>1995
<7>Four unique restriction enzymes were identified in the soluble protein fraction of Spirulina
platensis strain pacifica, a commercially important strain of marine cyanobacterium that is
used as a supplement in human diets.  These are SpaI, SpaII, SpaII and SpaIV, which are
isoschizomers of Tth111I, PvuI, PvuII and HindIII, respectively.  The recognition sites of
each of these four enzymes were identified by restriction digests of different plasmid DNAs of
known sequence and determining the cleavage sites by sequencing.  SpaI is the most predominant
restriction enzyme present in S. platensis strain pacifica.  It shows high activity at 37oC
compared to 65oC for its isoschizomer Tth111I.

<>

<1>Tran, P.H.
<2>X-ray crystal structure of N-6 adenine deoxyribose nucleic acid methyltransferase from Streptococcus pneumoniae.
<3>Diss. Abstr.
<4>61
<5>4801-B-4802-B
<6>2001
<7>X-ray diffraction by using resonant anomalous scattering has become a popular tool for solving
crystal structures in the last ten years with the expanded availability of tunable synchrotron
radiation for protein crystallography.  Mercury atoms were used for phasing.  The crystal
structure of N-6 deoxyribose nucleic acid methyltransferase from Streptococcus pneumoniae
(DpnM) was solved by using the Multiple Anomalous Diffraction technique.  The crystal
structure reveals the formation of mercaptide between the mercury ion and the thiol group on
the cysteine amino acid inn a hydrophobic environment.  The crystal structure contains the
bound ligand, S-adenosyl-L-methionine on the surface of the concave opening.  The direction of
the beta-strands on the beta sheets are identical to other solved methyltransferases.  The
highly conserved motifs, DPPY and the FxGxG, are found to be important in ligand binding and
possibly in methyl group transfer.  The structure has a concave cleft with an opening on the
order of 30 Angstroms that can accommodate a DNA duplex.  By molecular modeling coupled to
sequence alignment, two other highly conserved residues Arg21 and Gly19 are found to be
important in catalysis.

<>

<1>Tran, P.H., Korszun, Z.R., Cerritelli, S., Springhorn, S.S., Lacks, S.A.
<2>Crystal structure of the DpnM DNA adenine methyltransferase from the DpnII restriction system of Streptococcus pneumoniae bound to S-adenosylmethionine.
<3>Structure
<4>6
<5>1563-1575
<6>1998
<7>Methyltransferases catalyze the transfer of methyl groups from S-adenosylmethionine to a
variety of small molecular and macromolecular substrates.  These enzymes contain a
characteristic alpha/beta structural fold.  Four groups of DNA Mtases have been defined and
representative structures have been determined for three groups.  DpnM is a DNA Mtase that
acts on adenine N6 in the sequence GATC; the enzyme represents group alpha DNA Mtases, for
which no structures are known.

<>

<1>Tran-Betcke, A., Behrens, B., Noyer-Weidner, M., Trautner, T.A.
<2>DNA methyltransferase genes of Bacillus subtilis phages: comparison of their nucleotide sequences.
<3>Gene
<4>42
<5>89-96
<6>1986
<7>The Phi3T DNA methyltransferase (Mtase) and most of the SPbeta Mtase genes have
been sequenced.  With the exception of their promoters, no difference was found
between the Phi3T and SPbeta Mtase genes which code for an enzyme with a Mr, of
50507, consisting of 443 amino acids (aa).  Comparison of the deduced aa
sequence of the Phi3T/SPbeta type Mtase (target specificity: GGCC and GCNGC)
with that of the previously established sequence of the SPR Mtase (Buhk et al.,
1984) which has the target specificity GGCC and CCGG, reveals strong
similarities between these two types of enzymes.  There is, however, one
striking difference: both the Phi3T/SPbeta and the SPR enzymes contain at
different positions inserts of 33 aa, which have no homology to each other.  We
suggest that the methylation specificity unique to each of the two types of
Mtases (GCNGC in Phi3T/SPbeta ; CCGG in SPR) depends on these inserts, while
the GGCC-specific modification potential common to all Mtases is determined by
structures conserved in both types of enzymes.  A DNA fragment of non-modifying
phage Z, which shows homology to both flanks of the SPR Mtase gene, was also
sequenced.  This segment can be described as a derivative of SPR DNA, in which
the Mtase gene and sequences at its 5' end have been deleted, with the deletion
extending between two direct repeats of 25 bp.

<>

<1>Tran-Nguyen, L.T., Kube, M., Schneider, B., Reinhardt, R., Gibb, K.S.
<2>Comparative genome analysis of "Candidatus Phytoplasma australiense" (subgroup tuf-Australia I; rp-A) and "Ca. Phytoplasma asteris" Strains  OY-M and AY-WB.
<3>J. Bacteriol.
<4>190
<5>3979-3991
<6>2008
<7>The chromosome sequence of "Candidatus Phytoplasma australiense" (subgroup tuf-Australia I;
rp-A), associated with dieback in papaya, Australian
grapevine yellows in grapevine, and several other important plant
diseases, was determined. The circular chromosome is represented by
879,324 nucleotides, a GC content of 27%, and 839 protein-coding genes.
Five hundred two of these protein-coding genes were functionally assigned,
while 337 genes were hypothetical proteins with unknown function.
Potential mobile units (PMUs) containing clusters of DNA repeats comprised
12.1% of the genome. These PMUs encoded genes involved in DNA replication,
repair, and recombination; nucleotide transport and metabolism;
translation; and ribosomal structure. Elements with similarities to phage
integrases found in these mobile units were difficult to classify, as they
were similar to both insertion sequences and bacteriophages. Comparative
analysis of "Ca. Phytoplasma australiense" with "Ca. Phytoplasma asteris"
strains OY-M and AY-WB showed that the gene order was more conserved
between the closely related "Ca. Phytoplasma asteris" strains than to "Ca.
Phytoplasma australiense." Differences observed between "Ca. Phytoplasma
australiense" and "Ca. Phytoplasma asteris" strains included the
chromosome size (18,693 bp larger than OY-M), a larger number of genes
with assigned function, and hypothetical proteins with unknown function.

<>

<1>Trasler, J.M., Alcivar, A.A., Hake, L.E., Bestor, T., Hecht, N.B.
<2>DNA methyltransferase is developmentally expressed in replicating and non-replicating male germ cells.
<3>Nucleic Acids Res.
<4>20
<5>2541-2545
<6>1992
<7>Genomic methylation patterns are established during maturation of primordial germ cells and
during gametogenesis. While methylation is linked to DNA replication in somatic cells, active
de novo methylation and demethylation occur in post-replicative spermatocytes during meiotic
prophase. We have examined differentiating male germ cells for alternative forms of DNA
(cytosine-5)-methyltransfease (DNA MTase) and have found a 6.2 kb DNA MTase mRNA that is
present in appreciable quantitites only in testis; in post-replicative pachytene spermatocytes
it is the predominant form of DNA MTase mRNA. The 5.2 kb DNA MTase mRNA, characteristic of all
somatic cells, was detected in isolated type A and B spermatogonia and haploid round
spermatids. Immunoblot analysis detected a protein in spermatogenic cells with a relative mass
of 180,000-200,000, which is close to the known size of the somatic form of mammalian DNA
MTase. The demonstration of the differential developmental expression of DNA MTase in male
germ cells argues for a role for testicular DNA methylation events, not only during
replication in premeiotic cells, but also during meiotic prophase and postmeiotic development.

<>

<1>Trasler, J.M., Trasler, D.G., Beston, T.H., Li, E., Ghibu, F.
<2>DNA methyltransferase in normal and Dnmtn/Dnmtn mouse embryos.
<3>Dev. Dyn.
<4>206
<5>239-247
<6>1996
<7>The mouse genome experiences a large decrease in net 5-methylcytosine
between fertilization and implantation; de novo methylation brings 5-methylcytosine to
adult somatic cell levels between implantation and gastrulation.  Very little is known of the
regulation of demethylation or de novo methylation.  Levels of the one known form of
DNA methyltransferase are very high in early embryos, but the enzyme is localized to the
cytoplasm during most of preimplantation development.  We show here that DNA
methyltransferase is found exclusively in nuclei of the conceptus after implantation, and
that nuclei of proximal decidual cells are free of detectable DNA methyltransferase.  High
levels of DNA methyltransferase were seen in all tissues, including the developing nervous
system, of 9.5- to 12.5-day embryos.  The large maternal stores of DNA methyltransferase
become limiting prior to embryonic day 9.5, as shown by barely detectable immunostaining
in 9.5-day embryos homozygous for a loss-of-function mutation (Dnmtn) in the DNA
methyltransferase gene.  These mutant embryos failed to develop past the 25-somite stage
and showed evidence of developmental delay and some developmental asynchrony.
Normal embryonic and extraembryonic tissues contained similar levels of DNA
methyltransferase, even though severely reduced methylation levels and a loss of
imprinting have previously been observed in extraembryonic tissues.  These findings
suggest that methylation patterns are not a simple function of the concentration of DNA
methyltransferase, and that unidentified factors must be involved in the regulation of de
novo methylation during early development of the mouse.

<>

<1>Trautmann, D., Voss, B., Wilde, A., Al-Babili, S., Hess, W.R.
<2>Microevolution in cyanobacteria: re-sequencing a motile substrain of Synechocystis sp. PCC 6803.
<3>DNA Res.
<4>19
<5>435-448
<6>2012
<7>Synechocystis sp. PCC 6803 is a widely used model cyanobacterium for studying
photosynthesis, phototaxis, the production of biofuels and many other aspects.
Here we present a re-sequencing study of the genome and seven plasmids of one of
the most widely used Synechocystis sp. PCC 6803 substrains, the glucose tolerant
and motile Moscow or 'PCC-M' strain, revealing considerable evidence for recent
microevolution. Seven single nucleotide polymorphisms (SNPs) specifically shared
between 'PCC-M' and the 'PCC-N and PCC-P' substrains indicate that 'PCC-M'
belongs to the 'PCC' group of motile strains. The identified indels and SNPs in
'PCC-M' are likely to affect glucose tolerance, motility, phage resistance,
certain stress responses as well as functions in the primary metabolism,
potentially relevant for the synthesis of alkanes. Three SNPs in intergenic
regions could affect the promoter activities of two protein-coding genes and one
cis-antisense RNA. Two deletions in 'PCC-M' affect parts of clustered regularly
interspaced short palindrome repeats-associated spacer-repeat regions on plasmid
pSYSA, in one case by an unusual recombination between spacer sequences.

<>

<1>Trautner, T.A., Balganesh, T., Wilke, K., Noyer-Weidner, M., Rauhut, E., Lauster, R., Behrens, B., Pawlek, B.
<2>Organization of target-recognizing domains in the multispecific DNA (cytosine-5) methyltransferases of Bacillus subtilis phages SPR and Phi-3T.
<3>Gene
<4>74
<5>267
<6>1988
<7>Meeting Abstract

<>

<1>Trautner, T.A., Balganesh, T.S., Pawlek, B.
<2>Chimeric multispecific DNA methyltransferases with novel combinations of target recognition.
<3>Nucleic Acids Res.
<4>16
<5>6649-6658
<6>1988
<7>DNA target recognizing domains of different multispecific
DNA-cytosine-methyltransferases can be rearranged through engineering of the
corresponding genes to generate enzymes with novel combinations of target
recognition.

<>

<1>Trautner, T.A., Noyer-Weidner, M.
<2>Restriction/modification and methylation systems in Bacillus subtilis, related species, and their phages.
<3>Bacillus subtilis and other gram-positive bacteria: Biochemistry, Physiology, and Molecular Genetics, American Society for Microbiology, Hoch, J.A., Losick, R., Washington D.C.
<4>0
<5>539-552
<6>1993
<7>Several restriction/modification (R/M) systems have been identified in Bacillus subtilis and
related bacteria and will be described here. Accepting the view that R/M systems have evolved
to defend bacteria effectively against attack by bacterial viruses, we shall discuss the
question of what mechanisms bacteriophages have developed to overcome barriers provided by
host R/M systems. To what extent do R/M systems affect other inter- and intra-specific
transport of DNA? In this connection, we shall discuss the usefulness of restriction systems,
with their high substrate specificities for double-stranded DNA, in understanding the
processing of free or packaged DNA during uptake into B. subtilis cells or subcellular
structures. Recent progress in the characterization of genes encoding restriction
endonucleases (ENases) and modification methyltransferases (MTases) from a wide range of
organisms has made such systems per se interesting paradigms for the study of the evolution of
highly specific DNA-binding proteins. In particular, our interest is focused here on the
phylogenetic relationship among the various ENases and MTases of Bacillus and other bacterial
species. Furthermore, the requirement of coexistence of an ENase with an MTase represents an
interesting case of obligatory coevolution of two genes. The study of multispecific MTases,
discovered so far only in some temperate B. subtilis and Bacillus amyloliquefaciens phages,
has made significant contribution to our present understanding of the nature, function, and
evolution of R/M systems and particularly of their MTases. The interesting biochemical aspects
of the action of ENases and MTases will not be covered here. Such discussions are included in
a recent review.

<>

<1>Trautner, T.A., Pawlek, B., Behrens, B., Willert, J.
<2>Exact size and organization of DNA target-recognizing domains of multispecific DNA-(cytosine-C5)-methyltransferases.
<3>EMBO J.
<4>15
<5>1434-1442
<6>1996
<7>A large portion of the sequences of type II DNA-(cytosine-C5)-methyltransferases (C5-MTases)
represent highly conserved blocks of amino acids.  General steps in the methylation reaction
performed by C5-MTases have been found to be mediated by some of these domains.  C5-MTases
carry, in addition at the same relative location, a region variable in size and amino acid
composition, part of which is associated with the capacity of each C5-MTase to recognize its
characteristic target.  Individual target-recognizing domains (TRDs) for the targets CCGG (M),
CC(A/T)GG (E), GGCC (H), GCNGC (F) and G(G/A/T)GC(C/A/T)C (B) could be identified in the
C-terminal part of the variable region of multispecific C5-MTases.  With experiments reported
here, we have established the organization of the variable regions of the multispecific MTases
M.SPRI, M.Phi3TI, M.H2I and M.Rho11sI at the resolution of individual amino acids.  These
regions comprise 204, 175, 268 and 268 amino acids, respectively.  All variable regions are
bipartite.  They contain at their N-terminal side a very similar sequence of 71 amino acids.
The integrity of this sequence must be assured to provide enzyme activity.  Bracketed by 6-10
linker amino acids, they have, depending on the enzyme studied, towards their C-terminal end
ensembles of individual TRDs of 38 (M), 39 (E), 40 (H), 44 (F) and 54 (B) amino acids.  TRDs
of different enzymes with equal specificity have the same size.  TRDs do not overlap but are
either separated by linker amino acids or abut each other.

<>

<1>Trautner, T.A., Pawlek, B., Bron, S., Anagnostopoulos, C.
<2>Restriction and Modification in B. subtilis.
<3>Mol. Gen. Genet.
<4>131
<5>181-191
<6>1974
<7>Restriction and modification observed in Bacillus subtilis strain "R" affects
infection and transfection with phages SPP1, Phi105 and SPO2, but not with SP8,
Phi29, SP82, Sp50, H1 or PBS1.  It affects also PBS1 mediated transduction, but
not transformation with bacterial DNA.  The marker(s) determining
restriction/modification map between the origin of replication of the B.
subtilis chromosome and purA16.

<>

<1>Trautner, T.A., Pawlek, B., Gunthert, U., Canosi, U., Jentsch, S., Freund, M.
<2>Restriction and modification in Bacillus subtilis: Identification of a gene in the temperate phage SPbeta coding for a BsuR specific modification methyltransferase.
<3>Mol. Gen. Genet.
<4>180
<5>361-367
<6>1980
<7>A gene coding for a modifying DNA-methyltransferase which methylates the
central C in the BsuR recognition sequence 5'GGCC was identified in the genome
of the temperate Bacillus subtilis phage SPbeta.  This gene is expressed only
after induction of the prophage by either mitomycin C or UV.  The presence of
active methyltransferase in induced cells leads to modification of BsuR
recognition sites in SPbeta DNA as well as in heterologous DNA.

<>

<1>Travis, J.
<2>DNA flips out!  Enzymes repair and modify DNA in a surprising way.
<3>Sci. News
<4>148
<5>188-189
<6>1995
<7>According to recent research, enzymes have another way of tackling DNA.  Some apparently pry
apart a base pair, then rotate one of the freed nucleotides, bringing its base out of the
confines of the double helix and into the enzyme's active site, a pocket within the
protein's structure.  The enzyme can then remove this pocketed base from its nucleotide or
modify the base and sling it back into its proper position.

<>

<1>Travisany, D., Di Genova, A., Sepulveda, A., Bobadilla-Fazzini, R.A., Parada, P., Maass, A.
<2>Draft Genome Sequence of the Sulfobacillus thermosulfidooxidans Cutipay Strain, an Indigenous Bacterium Isolated from a Naturally Extreme Mining Environment in  Northern Chile.
<3>J. Bacteriol.
<4>194
<5>6327-6328
<6>2012
<7>Sulfobacillus thermosulfidooxidans strain Cutipay is a mixotrophic, acidophilic,  moderately
thermophilic bacterium isolated from mining environments of the north
of Chile, making it an interesting subject for studying the bioleaching of
copper. We introduce the draft genome sequence and annotation of this strain,
which provide insights into its mechanisms for heavy metal resistance.

<>

<1>Treangen, T.J., Maybank, R.A., Enke, S., Friss, M.B., Diviak, L.F., Karaolis, D.K., Koren, S., Ondov, B., Phillippy, A.M., Bergman, N.H., Rosovitz, M.J.
<2>Complete Genome Sequence of the Quality Control Strain Staphylococcus aureus subsp. aureus ATCC 25923.
<3>Genome Announcements
<4>2
<5>e01110-14
<6>2014
<7>Staphylococcus aureus subsp. aureus ATCC 25923 is commonly used as a control strain for
susceptibility testing to antibiotics and as a quality control strain
for commercial products. We present the completed genome sequence for the strain,
consisting of the chromosome and a 27.5-kb plasmid.

<>

<1>Trees, E., Strockbine, N., Changayil, S., Ranganathan, S., Zhao, K., Weil, R., MacCannell, D., Sabol, A., Schmidtke, A., Martin, H., Stripling, D., Ribot, E.M., Gerner-Smidt, P.
<2>Genome Sequences of 228 Shiga Toxin-Producing Escherichia coli Isolates and 12 Isolates Representing Other Diarrheagenic E. coli Pathotypes.
<3>Genome Announcements
<4>2
<5>e00718-14
<6>2014
<7>Shiga toxin-producing Escherichia coli (STEC) are a common cause for food-borne diarrheal
illness outbreaks and sporadic cases. Here, we report the availability
of the draft genome sequences of 228 STEC strains representing 32 serotypes with
known pulsed-field gel electrophoresis (PFGE) types and epidemiological
relationships, as well as 12 strains representing other diarrheagenic E. coli
pathotypes.

<>

<1>Trefault, N., De la Iglesia, R., Molina, A.M., Manzano, M., Ledger, T., Perez-Pantoja, D., Sanchez, M.A., Stuardo, M., Gonzalez, B.
<2>Genetic organization of the catabolic plasmid pJP4 from Ralstonia eutropha JMP134 (pJP4) reveals mechanisms of adaptation to chloroaromatic   pollutants and evolution of specialized chloroaromatic degradation   pathways.
<3>Environ. Microbiol.
<4>6
<5>655-668
<6>2004
<7>Ralstonia eutropha JMP134 (pJP4) is a useful model for the study of bacterial degradation of
substituted aromatic pollutants. Several key
degrading capabilities, encoded by tfd genes, are located in the 88 kb,
self-transmissible, IncP-1 beta plasmid pJP4. The complete sequence of the
87,688 nucleotides of pJP4, encoding 83 open reading frames (ORFs), is
reported. Most of the coding sequence corresponds to a well-conserved
IncP-1 beta backbone and the previously reported tfd genes. In addition,
we found hypothetical proteins putatively involved in the transport of
aromatic compounds and short-chain fatty acid oxidation. ORFs related to
mobile elements, including the Tn501-encoded mercury resistance
determinants, an IS1071-based composite transposon and a cryptic class II
transposon, are also present in pJP4. These mobile elements are
inefficient in transposition and are located in two regions of pJP4 that
are rich in remnants of lateral gene transfer events. pJP4 plasmid was
able to capture chromosomal genes and form hybrid plasmids with the IncP-1
alpha plasmid RP4. These observations are integrated into a model for the
evolution of pJP4, which reveals mechanisms of bacterial adaptation to
degrade pollutants.

<>

<1>Treml, S., Draveling, C., Huang, C., Heaster, J., Walker, D., DiFrancesco, R., Jolly, J.
<2>Ambient-temperature-stable restriction endonucleases for use in forensic DNA applications.
<3>Clin. Chem.
<4>40
<5>1092
<6>1994
<7>We have stabilized restriction endonucleases including HaeIII, HinfI, and PstI, which are
commonly used in forensic and paternity DNA profiling. Using a patented process (US patents
5098893 and 5250429) the enzyme and reaction buffer are stabilized as a glassy matrix of
carbohydrate polymers. These glassified reaction systems show excellent stability at ambient
temperatures for prolonged periods of time. The glass transition temperature (Tg) is a
physical property of the material and can be determined using a Differential Scanning
Calorimeter. At temperatures equal to or lower than the Tg the material remains a glass.
Stability studies show the recovery of enzyme activity immediately after processing is 75-100%
of activity prior to processing, and there is no further loss of activity over extended
periods of time provided the enzyme is stored at temperatures below the Tg. We have dried the
enzymes HinfI, HaeIII, and PstI by this process. The Gg's (oC) were 41.0, 45.2, and 47.4
respectively. After processing, the enzymes were stored at 25, 37, and 55oC for extended
periods of time. After 34 weeks HinfI retains 100% of initial activity at 25 and 37oC and 70%
of initial activity at 55oC. After 30 weeks HaeIII retains 100% initial activity at all three
storage temperatures. PstI after 36 weeks of storage retains 100% initial activity at all
three storage temperatures. PstI after 36 weeks of storage retains 100% of initial activity at
25 and 37oC and 75% of initial activity at 55oC. These pre-mixed single dose reactions also
offer convenience and reproducibility by reducing pipeting steps and possible cross
contamination. Since the enzymes are stable at ambient temperatures they offer the added
convenience of room temperature shipping and storage. The enzymes are readily rehydrated by
simply adding water and the DNA to be analyzed. The stabilizer used in this process eliminates
the need for glycerol in the enzyme preparation. The absence of glycerol reduces star activity
that is sometimes observed when digests are carried out in the presence of high glycerol
concentrations due to a high ratio of enzyme units to DNA. The stabilized enzymes have been
used in side by side studies with standard RFLP protocols. A PstI paternity analysis was done
using Lifecodes procedures with standard PstI and our stabilized PstI. The analysis done with
standard PstI resulted in an extra band due to partial digestions. The extra band was not
present in the analysis done with stabilized PstI. The stabilized restriction enzymes offer
quick, reliable and reproducible results for standard forensic and paternity profiling
protocols.

<>

<1>Treu, L., Vendramin, V., Bovo, B., Campanaro, S., Corich, V., Giacomini, A.
<2>Genome Sequences of Four Italian Streptococcus thermophilus Strains of Dairy Origin.
<3>Genome Announcements
<4>2
<5>e00126-14
<6>2014
<7>This report describes the genome sequences of four Streptococcus thermophilus strains, namely,
TH982, TH985, TH1477, and 1F8CT, isolated from different dairy
environments from the Campania and the Veneto regions in Italy. These data are
aimed at increasing the genomic information available on this species, which is
of paramount importance for the dairy industry.

<>

<1>Treu, L., Vendramin, V., Bovo, B., Campanaro, S., Corich, V., Giacomini, A.
<2>Whole-Genome Sequences of Streptococcus thermophilus Strains TH1435 and TH1436, Isolated from Raw Goat Milk.
<3>Genome Announcements
<4>2
<5>e01129-13
<6>2014
<7>We report the genome sequences of two Streptococcus thermophilus strains, TH1435  and TH1436,
isolated from raw goat milk devoted to the production of artisanal
cheese in the Friuli-Venezia Giulia region in Italy. The genome sequences of
these two quickly acidifying strains are the first available genome sequences of
S. thermophilus strains isolated in Italy.

<>

<1>Treu, L., Vendramin, V., Bovo, B., Campanaro, S., Corich, V., Giacomini, A.
<2>Genome Sequences of Streptococcus thermophilus Strains MTH17CL396 and M17PTZA496  from Fontina, an Italian PDO Cheese.
<3>Genome Announcements
<4>2
<5>e00067-14
<6>2014
<7>Here is presented the whole-genome sequences of Streptococcus thermophilus strains MTH17CL396
and M17PTZA496, isolated from fontina protected designation of
origin (PDO) cheese in the Valle d'Aosta Region (Italy). S. thermophilus is a
lactic acid bacterium widely present in dairy products, and these are the first
publicly available genome sequences of S. thermophilus strains isolated from
cheese.

<>

<1>Treu, L., Vendramin, V., Bovo, B., Giacomini, A., Corich, V., Campanaro, S.
<2>Genome Sequence of Lactobacillus fabifermentans Strain T30PCM01, Isolated from Fermenting Grape Marc.
<3>Genome Announcements
<4>2
<5>e00060-14
<6>2014
<7>Here, we report the draft genome assembly of Lactobacillus fabifermentans strain  T30PCM01
isolated from grape marc. Its genome is the largest (3.58 Mbp) among
Lactobacillus species and reveals an enormous potential for carbohydrate
utilization and transcriptional regulation.

<>

<1>Treven, P., Trmcic, A., Bogovic, M.B., Rogelj, I.
<2>Improved Draft Genome Sequence of Probiotic Strain Lactobacillus gasseri K7.
<3>Genome Announcements
<4>2
<5>e00725-14
<6>2014
<7>Lactobacillus gasseri K7 is an isolate from infant feces and has in vitro and in  vivo
established probiotic properties. Here, we report the improved version of
the draft genome sequence, which comprises 8 scaffolds (13 contigs), a total
length of 1.99 Mb, and 1,841 predicted protein-coding sequences.

<>

<1>Trevors, J.T.
<2>Molecular evolution in bacteria.
<3>Antonie Van Leeuwenhoek
<4>67
<5>315-324
<6>1995
<7>Recent advances in microbiology and molecular biology have a unifying influence on our
understanding of genetic diversity/similarity and evolutionary relationships in
microorganisms.  This article attempts to unify information from diverse areas such as
microbiology, molecular biology, microbial physiology, clay crystal genes, metals-microbe-clay
interactions and bacterial DNA restriction-modification systems (R-M) as they may apply to
molecular evolution of bacteria.  The possibility is discussed that the first informational
molecules may have been catalytic RNA (micro-assembler) not DNA (now the master copy) and
these first micro-assemblers may have been precursors of ribosomes.

<>

<1>Trevors, J.T.
<2>Review: Bacterial population genetics.
<3>World J. Microbiol. Biotechnol.
<4>14
<5>1-5
<6>1998
<7>Bacterial population genetics is the study of natural bacterial genetic diversity arising from
evolutionary processes.  The roles of molecular mistakes, restriction-modification, plasmids
and gene transfer in bacteria are also important components of population genetics.  These
aspects are of considerable scientific importance from a fundamental perspective, because of
the short generation times of bacteria, their microscopic cell size, the large population
sizes bacteria can achieve and their different mechanisms of gene transfer.

<>

<1>Trevors, J.T.
<2>Molecular evolution in bacteria: genome size, cell size, restriction-modification and recognition.
<3>Bull. Inst. Pasteur
<4>96
<5>25-33
<6>1998
<7>The ability of bacteria to alter their genome sizes and the order of their genes, yet maintain
a relatively constant genome, provides a mechanism for diversity and evolution in bacteria.
Moreover, bacteria may have evolved by increasing their genome sizes and rearranging gene
orders with the assistance of restriction endonucleases cleaving foreign DNA and providing a
diverse pool of DNA sequences for genetic recombination.  This review examines some of these
evolutionary aspects of bacteria including molecular recognition of biomolecules.

<>

<1>Tribelli, P.M., Raiger, I.L.J., Catone, M.V., Di Martino, C., Revale, S., Mendez, B.S., Lopez, N.I.
<2>Genome Sequence of the Polyhydroxybutyrate Producer Pseudomonas extremaustralis,  a Highly Stress-Resistant Antarctic Bacterium.
<3>J. Bacteriol.
<4>194
<5>2381-2382
<6>2012
<7>Pseudomonas extremaustralis 14-3b presents genes involved in the synthesis of different
polyhydroxyalkanoates, in tolerance and degradation of pollutants, and
in microaerobic metabolism. Several genomic islands were detected. Genetic
machinery could contribute to the adaptability to stressful conditions. This is
the first genome sequence reported from a Pseudomonas isolated from cold
environments.

<>

<1>Trieu-Cuot, P., Derlot, E., Courvalin, P.
<2>Enhanced conjugative transfer of plasmid DNA from Escherichia coli to Staphylococcus aureus and Listeria monocytogenes.
<3>FEMS Microbiol. Lett.
<4>109
<5>19-23
<6>1993
<7>Transfer of mobilizable shuttle cloning vectors by conjugation from Escherichia coli to
Staphylococcus aureus occurred at a very low frequency (10-9 transconjugants per donor
colony-forming unit after the mating period). It was observed that subinhibitory
concentrations of penicillins (oxacillin or penicillin G) in the mating medium resulted in
increased transfer frequency by conjugation of the shuttle vector pAT18 from E. coli SM10 to
S. aureus 80CR5 Str (54-fold) and to Listeria monocytogenes LO17RF (45-fold). These results
were interpreted as indicating that the cell wall of Gram-positive bacteria constitutes an
important barrier for conjugative transfer of genetic information delivered from E. coli. It
was also demonstrated that the presence of a restriction system(s) in S. aureus recipients
represented a major barrier to introduction of foreign DNA.

<>

<1>Trimble, W.L., Phung, L.T., Meyer, F., Gilbert, J.A., Silver, S.
<2>Draft Genome Sequence of Agrobacterium albertimagni Strain AOL15.
<3>J. Bacteriol.
<4>194
<5>6986-6987
<6>2012
<7>Agrobacterium albertimagni strain AOL15 is an alphaproteobacterium isolated from
arsenite-oxidizing biofilms whose draft genome contains 5.1 Mb in 55 contigs with
61.2% GC content and includes a 21-gene arsenic gene island. This is the first
available genome for this species and the second Agrobacterium arsenic gene
island.

<>

<1>Trimble, W.L., Phung, L.T., Meyer, F., Silver, S., Gilbert, J.A.
<2>Draft Genome Sequence of Achromobacter piechaudii Strain HLE.
<3>J. Bacteriol.
<4>194
<5>6355
<6>2012
<7>Achromobacter piechaudii strain HLE is a betaproteobacterium (previously known as Alcaligenes
faecalis) that was an early isolate with arsenite oxidase activity.
This draft genome of 6.89 Mb is the second available genome for this species in
the opportunistic pathogen Alcaligenaceae family.

<>

<1>Trotter, M., McAuliffe, O., Callanan, M., Edwards, R., Fitzgerald, G.F., Coffey, A., Ross, R.P.
<2>Genome analysis of the obligately lytic bacteriophage 4268 of Lactococcus lactic provides insight into its adaptable nature.
<3>Gene
<4>366
<5>189-199
<6>2006
<7>Analysis of the complete nucleotide sequence of the lactococcal phage 4268, which is lytic for
the cheese starter Lactococcus lactis DPC4268, is presented. Phage 4268 has a linear genome of
36,596 bp, which is modularly organised and encompasses 49 open reading frames. Putative
functions were assigned to approximately 45% of the predicted products of these open reading
frames based on sequence similarity with known proteins, N-terminal sequence analysis and
identification of conserved domains. Significantly, a segment of the genome has homology to
the recently sequenced lysogenic module in lactococcal phage phi 31 that contains a lytic
switch but no phage integrase or attachment site. This suggests that it is derived from a
prophage. A phage 4268-encoded and a host-encoded methylase were found to be highly similar,
having only two nucleotide mismatches, suggesting that the phage acquired the methylase gene
to protect it from a host endonuclease. Comparative genomic analysis revealed significant
homology between phage 4268 and the lactococcal phage BK5-T. The comparative analysis also
supported the classification of phage 4268 and other BK5-T-related phage as separate from the
proposed P335 species of lactococcal phage.

<>

<1>Trotter, M., Ross, R.P., Fitzgerald, G.F., Coffey, A.
<2>Lactococcus lactis DPC5598, a plasmid-free derivative of a commercial starter, provides a valuable alternative host for culture improvement studies.
<3>J. Appl. Microbiol.
<4>93
<5>134-143
<6>2002
<7>Aims: To generate a plasmid-free derivative of an extensively used industrial starter strain
Lactococcus lactis DPC4268, which could be used as a backbone strain for starter improvement
programmes. Methods and Results: DPC4268, containing four large plasmids, was subjected to
high temperature plasmid curing resulting in derivatives, each with a different plasmid
complement of one, two or three different plasmids in addition to a plasmid-free derivative.
Industrially relevant phenotypes were assigned to each plasmid on the basis of detailed
phenotypic and genetic analyses and these were (a) proteinase activity (Prt, 60 kb) (b)
lactose fermentation (Lac, 55 kb) (c) bacteriophage adsorption inhibition (Ads, 44 kb) and (d)
type I restriction/modification (R/M, 40 kb). The plasmid-free variant of DPC4268 was shown to
be transformable at frequencies comparable to the common laboratory strain L. lactis MG1614.
Furthermore its genome was demonstrated to be significantly different from the laboratory
strains L. lactis MG1614 and the recently sequenced L. lactis IL1403 genomes by pulsed-field
gel electrophoresis. Conclusions: This study produced an easily transformable plasmid-free
derivative which was genomically different from both MG1614 and IL1403. In addition, important
plasmid-borne industrial traits, including two phage-resistance mechanisms, were identified in
DPC4268. Significance and Impact of the Study: L. DPC4268 is a vitally important commercial
strain used in the manufacture of Cheddar cheese. The generation of a plasmid-free derivative
may provide an important backbone strain as a basis for future strain improvement purposes.

<>

<1>Trowbridge, J.J., Snow, J.W., Kim, J., Orkin, S.H.
<2>DNA Methyltransferase 1 Is Essential for and Uniquely Regulates Hematopoietic Stem and Progenitor Cells.
<3>Cell Stem Cell
<4>5
<5>442-449
<6>2009
<7>DNA methylation is essential for development and in diverse biological processes. The DNA
methyltransferase Dnmt1 maintains parental cell
methylation patterns on daughter DNA strands in mitotic cells; however,
the precise role of Dnmt1 in regulation of quiescent adult stem cells
is not known. To examine the role of Dnmt1 in adult hematopoietic stem
cells (HSCs), we conditionally disrupted Dnmt1 in the hematopoietic
system. Defects were observed in Dnmt1 deficient HSC self-renewal,
niche retention, and in the ability of Dnmt1-deficient HSCs to give
rise to multilineage hematopoiesis. Loss of Dnmt1 also had specific
impact on myeloid progenitor cells, causing enhanced cell cycling and
inappropriate expression of mature lineage genes. Dnmt1 regulates
distinct patterns of methylation and expression of discrete gene
families in long-term HSCs and multipotent and lineage-restricted
progenitors, suggesting that Dnmt1 differentially controls these
populations. These findings establish a unique and critical role for
Dnmt1 in the primitive hematopoietic compartment.

<>

<1>Trubitsyn, D., Geurink, C., Pikuta, E., Lefevre, C.T., McShan, W.M., Gillaspy, A.F., Bazylinski, D.A.
<2>Draft Genome Sequence of the Obligately Alkaliphilic Sulfate-Reducing Bacterium Desulfonatronum thiodismutans Strain MLF1.
<3>Genome Announcements
<4>2
<5>e00741-14
<6>2014
<7>Desulfonatronum thiodismutans strain MLF1, an alkaliphilic bacterium capable of sulfate
reduction, was isolated from Mono Lake, California. Here we report the
3.92-Mb draft genome sequence comprising 34 contigs and some results of its
automated annotation. These data will improve our knowledge of mechanisms by
which bacteria withstand extreme environments.

<>

<1>Trujillo, L.E., Brito, J.E., Reyes, G., Garcia, O.
<2>Purification of SmaI restriction enzyme.
<3>Biotecnol. Apl.
<4>7
<5>326-332
<6>1990
<7>SmaI restriction endonuclease produced by Serratia marcescens, has been
purified in our laboratory using PEG-6000 precipitation and ion exchange
chromatography in Phosphocellulose (P-11 Whatman).  We have obtained by this
way a final enzymatic preparation with a specific activity of 2972 U/mg of
total protein and 35% of global recovery of the process, that may be used in
cloning and other recombinant DNA techniques.

<>

<1>Trujillo, L.E., Pupo, E., Miranda, F., Perez, E., Gonzalez, E.
<2>Correspondence between radioactive and functional methods in the quality control of DNA restriction and modifying enzymes.
<3>Rev. Latinoam. Microbiol.
<4>38
<5>31-37
<6>1996
<7>We evaluated the use of two radiolabeled lambda DNA/HpaII substrates to detect 5'-3' single
and double stranded DNA dependent exonuclease and phosphatase activities found as contaminants
in restriction and modifying enzyme preparations.  Looking for the meaning of the radioactive
assays results in a real cloning experience, we performed a cloning simulation assay using the
same conditions established for the radioactive assay (enzyme units and pmols of DNA ends).
As a result, we found that for degradation percentages of the radioactive DNA substrate per
enzyme unit below 0.5, the false positives in the cloning simulation assay were less than 5%.
This conditions could ensure a good performance of the enzyme preparations for cloning
experiments.  Finally, we described the use of the radiolabeled [gamma 32P] ATP lambda HpaII
DNA substrate to detect 5'-3' single stranded DNA dependent exonuclease and phosphatase
contaminating activities in some critical steps of the purification process of the restriction
enzyme KpnI.

<>

<1>Trujillo, L.E., Reyes, G., Bayolo, E., Vazquez, M.M., Garcia, O.
<2>Purification of NciI restriction endonuclease.
<3>Biotecnol. Apl.
<4>7
<5>320-325
<6>1990
<7>NciI restriction endonuclease isolated from Neisseria cinerea, has been
purified in our laboratory using affinity and ion exchange chromatography.  We
have obtained an enzymatic preparation with a specific activity of 20,000 U/mg
of proteins and 67.2% of global recovery of the process, that may be used in
different genetic engineering techniques.

<>

<1>Tsai, S.P., Hartin, R.J., Ryu, J.-I.
<2>Transformation in restriction-deficient Salmonella typhimurium LT2.
<3>J. Gen. Microbiol.
<4>135
<5>2561-2567
<6>1989
<7>Stable restriction-deficient, modification-proficient galE (JR501) and F'galE+
(JR502) strains of Salmonella typhimurium were constructed and the effects of
restriction on transformation by plasmid pBR322 were tested.  Several factors
which affect transformation efficiency were systematically examined to
determine optimum transformation conditions and a simplified method is
presented.

<>

<1>Tsao, D.H.H., Maki, A.H.
<2>Optically detected magnetic resonance study of the interaction of an Arsenic(III) derivative of cacodylic acid with EcoRI methyl transferase.
<3>Biochemistry
<4>30
<5>4565-4572
<6>1991
<7>The interaction of the enzyme Escherichia coli RI methyl transferase
(methylase) with an arsenic(III) derivative of cacodylic acid has been
investigated by optical detection of triplet-state magnetic resonance (ODMR)
spectroscopy in zero applied magnet field.  The reactive derivative (CH3)2AsSR
is formed by the reduction of cacodylate by a thiol.  The As(III) derivative
binds to the enzyme by mercaptide exchange with a cysteine (Cys) residue
located close to a tryptophan (Trp) site.  The arsenical binding selectively
induces an external heavy-atom effect, perturbing the nearby Trp residue in the
enzyme.  Zero-field splittings (ZFS) and total decay rate constants of the
individual triplet-state sublevels of the Trp residue in the presence and
absence of perturbation by As(III) have been determined.  The perturbed Trp
shows a large reduction in the overall decay lifetime compared with unperturbed
Trp residue, exhibiting a high selectivity for the Tx sublevel.  This
selectivity suggests that the As atom lies in the xz plane of the principal
magnetic axis system of Trp, but not directly along the z (out-of-plane) axis.
The accessibility of this enzyme binding site to the arsenical is decreased
upon forming a ternary complex of methylase with sinefungin and a DNA oligomer,
d[GCGAA(BrU)(BrU)CGC], containing two 5-bromouracil (BrU) bases in place of
thymine within the hexadeoxynucleotide recognition sequence.  This result
indicates that the arsenical binding site in methylase which produces the Trp
heavy-atom effect is protected from this ligand by ternary complex formation or
the enzyme undergoes a conformation change, removing the Cys from the Trp site.
This protection is also observed in fluorescence quenching experiments.  The
As(III) reagent, upon binding to methylase, quenches the Trp fluorescence by
46%.  When the ternary complex is formed, the quenching of Trp fluorescence is
only 17%.  A binding constant for the arsenical to the high-affinity enzyme
site was obtained which is at least 27 times that of binding to a free
sulfhydryl residue.  The addition of a 1:1 molar ratio of the arsenical to
methylase did not affect the activity of the enzyme, but incubation with excess
arsenical quenches the activity, suggesting that the high-affinity Cys residue
is not involved in the DNA methylation process.  In the ternary complex
methylase-sinefungin-DNA, no heavy-atom perturbation of the two Trp residues in
the enzyme by BrU was observed, demonstrating that Trp residues are not
involved in close-range interactions with the two heavy-atom-derivatized
nucleic acid bases.  A similar result was observed previously with the
analogous E. coli RI endonuclease-decanucleotide complex [John, N.-I.,
Casas-Finet, J.R., Maki, A.H., & Modrich, P. (1988) Biochim. Biophys. Acta 949,
189-194].

<>

<1>Tsaplina, I.A., Bogdanova, T.I., Rechkunova, N.I., Degtyarev, S.K.
<2>Production of restriction enzyme Tru201I - using strain Thermus ruber as producer and conducting chromatographic purification of the enzyme in a specified regime.
<3>Soviet Patent Office
<4>SU 1703687 A
<5>
<6>1992
<7>The method of production of restriction enzyme Tru201I (restriction endonuclease, capable of
recognition and splitting the nucleotide sequence PuGATCPy) comprises cultivation of
microorganism-producer in a culture medium, separation of cells and their ultrasonic
destruction, followed by chromatographic purification on phosphocellulose and
heparin-sepharose sorbents. Cells of Thermus ruber BC-201 (used as the microorganism-producer)
are sown onto a solid medium containing 20% potato broth, 0.5% peptone and 0.02% yeast
extract, incubated for 1 day at 60 degrees C, transferred to 100 ml of a liquid medium of the
same composition and grown for 24 hrs. at 60 degrees C. The inoculate is again transferred
into 2 l of liquid medium of the same composition and grown at 60 degrees C for 14 hrs. at 60
rpm. After cooling, cells are collected by centrifuging for 10 min. at 6000 rpm. The yield of
biomass is 1 g per 1 l of the medium.  Collected cells are suspended in 0.02 M Tris-HCl
buffer, containing 10/-3 M beta-mercaptoethanol and 0.1% Triton X-100 and treated in an
ultrasonic disintegrator. The remaining cells are removed by centrifuging and the liquid phase
is subjected to stagewise chromatographic purification, using phosphocellulose and
heparin-sepharose as sorbents. The activity of the enzyme preparation is 2000 units/ml, the
yield is 1000 units of Tru201I per 1 g of the biomass. Tests show that the proposed method
increases the yield of produced restrictase by 8 and more times, compared to the known method.
The technology is simplified owing to reduced number of stages.

<>

<1>Tschoeke, D.A., Moreira, A.P., Chimetto, T.L.A., de Mesquita, M.M., Gregoracci, G.B., Gomez-Gil, B., Valle, R., Thompson, C.C., Thompson, F.L.
<2>Exploring the Genome of Cheese Starter Lactic Acid Bacterium Lactococcus lactis subsp. lactis CECT 4433.
<3>Genome Announcements
<4>2
<5>e01142-14
<6>2014
<7>Here, we present the draft genome sequences of Lactococcus lactis subsp. lactis CECT 4433, a
cheese fermentation starter strain. The genome provides further
insight into the genomic plasticity, biocomplexity (including gene strain
specifics), and evolution of these genera.

<>

<1>Tse, H., Tsoi, H.W., Leung, S.P., Lau, S.K., Woo, P.C., Yuen, K.Y.
<2>Complete Genome Sequence of Staphylococcus lugdunensis strain HKU09-01.
<3>J. Bacteriol.
<4>192
<5>1471-1472
<6>2010
<7>Staphylococcus lugdunensis is a member of the coagulase-negative staphylococci and commonly
found as part of the human skin flora. It is a significant cause of catheter-related
bacteremia, and also causes serious infections like native valve endocarditis in previously
healthy individuals. We report the complete genome sequence of this medically important
bacterium.

<>

<1>Tse, H., Tsoi, H.W., Leung, S.P., Urquhart, I.J., Lau, S.K., Woo, P.C., Yuen, K.Y.
<2>Complete genome sequence of the veterinary pathogen Staphylococcus pseudintermedius strain HKU10-03 isolated from a case of canine pyoderma.
<3>J. Bacteriol.
<4>193
<5>1783-1784
<6>2011
<7>Staphylococcus pseudintermedius is a member of the coagulase-positive staphylococci, and is
the commonest cause of canine pyoderma. We report the first genome sequence of S.
pseudintermedius, which showed the presence of numerous virulence factors akin to the related
human pathogen Staphylococcus aureus.

<>

<1>Tseng, S.P., Hsueh, P.R., Tsai, J.C., Teng, L.J.
<2>Tn6001, a Transposon-Like Element Containing the blaVIM-3-Harboring Integron In450.
<3>Antimicrob. Agents Chemother.
<4>51
<5>4187-4190
<6>2007
<7>We describe the structure of a transposon-like element named Tn6001, which
contains a bla(VIM-3)-harboring integron In450, which was derived from a
multidrug-resistant Pseudomonas aeruginosa clinical isolate in Taiwan. The
transposon backbone structure is most closely related to those of Tn1404*
and Tn1403. Tn6001 was inserted into the chromosome of the clinical
isolate.

<>

<1>Tsirigos, A., Rigoutsos, I.
<2>A new computational method for the detection of horizontal gene transfer events.
<3>Nucleic Acids Res.
<4>33
<5>922-933
<6>2005
<7>In recent years, the increase in the amounts of available genomic data has made it easier to
appreciate the extent by which organisms increase
their genetic diversity through horizontally transferred genetic
material. Such transfers have the potential to give rise to extremely
dynamic genomes where a significant proportion of their coding DNA has
been contributed by external sources. Because of the impact of these
horizontal transfers on the ecological and pathogenic character of the
recipient organisms, methods are continuously sought that are able to
computationally determine which of the genes of a given genome are
products of transfer events. In this paper, we introduce and discuss a
novel computational method for identifying horizontal transfers that
relies on a gene's nucleotide composition and obviates the need for
knowledge of codon boundaries. In addition to being applicable to
individual genes, the method can be easily extended to the case of
clusters of horizontally transferred genes. With the help of an
extensive and carefully designed set of experiments on 123 archaeal and
bacterial genomes, we demonstrate that the new method exhibits
significant improvement in sensitivity when compared to previously
published approaches. In fact, it achieves an average relative
improvement across genomes of between 11 and 41% compared to the Codon
Adaptation Index method in distinguishing native from foreign genes.
Our method's horizontal gene transfer predictions for 123 microbial
genomes are available online at http://cbcsrv.watson.ibm.com/HGT/.

<>

<1>Tsonos, J., Adriaenssens, E.M., Klumpp, J., Hernalsteens, J.P., Lavigne, R., De Greve, H.
<2>Complete Genome Sequence of the Novel Escherichia coli Phage phAPEC8.
<3>J. Virol.
<4>86
<5>13117-13118
<6>2012
<7>Bacteriophage phAPEC8 is an Escherichia coli-infecting myovirus, isolated on an
avian pathogenic Escherichia coli (APEC) strain. APEC strains cause
colibacillosis in poultry, resulting in high mortality levels and important
economic losses. Genomic analysis of the 147,737-bp double-stranded DNA phAPEC8
genome revealed that 53% of the 269 encoded proteins are unique to this phage.
Its closest relatives include the Salmonella phage PVP-SE1 and the coliphage rv5,
with 19% and 18% similar proteins, respectively. As such, phAPEC8 represents a
novel, phylogenetically distinct clade within the Myoviridae, with molecular
properties suitable for phage therapy applications.

<>

<1>Tsubouchi, T., Nishi, S., Usui, K., Shimane, Y., Takaki, Y., Maruyama, T., Hatada, Y.
<2>Draft Genome Sequence of the Dimorphic Prosthecate Bacterium Brevundimonas abyssalis TAR-001T.
<3>Genome Announcements
<4>1
<5>e00826-13
<6>2013
<7>We report the 3.0-Mb draft genome sequence of Brevundimonas abyssalis strain TAR-001(T),
isolated from deep-sea floor sediment. The draft genome sequence of
strain TAR-001(T) consists of 2,979,700 bp in 128 contigs, with a G+C content of
68.2%, 2,946 potential coding sequences (CDS), 3 rRNAs, and 41 tRNAs.

<>

<1>Tsuge, K., Matsui, K., Itaya, M.
<2>One step assembly of multiple DNA fragments with a designed order and orientation in Bacillus subtilis plasmid.
<3>Nucleic Acids Res.
<4>31
<5>e133
<6>2003
<7>A universal method to reconstitute sets of genes was developed. Owing to the intrinsic nature
of the plasmid establishment mechanism in Bacillus
subtilis, the assembly of five antibiotic resistance genes with a defined
order and orientation was achieved. These five fragments and the plasmid
have three-base protruding sequences at both ends. The protruding
sequences are designed so that each fragment is ligated once in a row
according to the pairing. Ligation by T4 DNA ligase in the presence of 150
mM NaCl and 10% polyethylene glycol at 37 degrees C yielded high molecular
tandem repeat linear form DNA. This multimeric form of DNA was
preferentially used for plasmid establishment in B.subtilis. The method,
referred to as Ordered Gene Assembly in B.subtilis (OGAB), allows for the
design of multiple fragments with very high efficiency and great fidelity.

<>

<1>Tsui, W.-C., Elgar, G., Merrill, C., Maunders, M.
<2>RspX I:  a new restriction endonuclease with a recognition sequence of 5'T^CATGA3'.
<3>Nucleic Acids Res.
<4>16
<5>4178
<6>1988
<7>A new TypeII restriction endonuclease, RspXI, was purified from a Rhodococcus
species isolated in our laboratory.  The Rhodococcus species grows well at 30C
in the following medium:  0.8% glucose, 0.8% yeast extract and 2% malt extract
at pH 7.2.  The yield of cell is 5 g/l.  RspXI recognizes the sequence
5'TCATGA3' and cleaves between T and C giving a four base 5' overhang.  An
enzyme BspH1 reported recently has the same cutting site.  (1) the enzyme was
purified by the following chromatographic steps:   1) phosphocellulose 2)
biorex-70 3) S-200.  It was judged to be essentially free of contaminating
nuclease activity.  After 10 fold overdigestion on lambda DNA greater than 95%
of the DNA fragments can be ligated and greater than 95% of the ligated DNA can
be recut by RspXI.  At 37C, the optimal conditions for RspXI activity are:  10
mM Tris, pH 7.5, 50 mM NaCl, 50 mM KCl and 10 mM MgCl2.  KCl is essential for
optimal activity.  The number of fragments generated by digestion with RspXI on
the following DNAs are:  lambda, 9; pBR322, 4; phiX174, 3; Adeno 2, 4 and
M13mp18, 1 (Fig. 1).  A computer search with Microgenie(TM) (Beckman Inst.
Inc.) suggested a recognition sequence of 5'TCATGA3'.  The cleavage site was
determined as follows:  pBR322 DNA was digested with RspXI.  DNA fragments were
then end-filled with DNA polymerase (Klenow fragment).  The resulting blunt-end
fragments were ligated into SmaI site of pUC18.  The recombinant plasmids were
transformed into E. Coli K803.  Ampicillin resistant and Lac- clones were
identified and plasmid DNAs were prepared from these clones.  The sequence of
the DNA fragment cloned into SmaI was determined by dideoxy Chain-termination
method (2).  The sequence over the cloning site is CATGAGCCC on one gel (not
shown) and is CATGAGCGT on the other gel.  This confirms that the cutting site
is between bases T and C.

<>

<1>Tsukahara, K., Kita, A., Nakashimada, Y., Hoshino, T., Murakami, K.
<2>Genome-guided analysis of transformation efficiency and carbon dioxide assimilation by Moorella thermoacetica Y72.
<3>Gene
<4>535
<5>150-155
<6>2014
<7>We determined a draft genome sequence for Moorella thermoacetica strain Y72, a
syngas-assimilating bacterium with high transformation efficiency. This strain was confirmed
to be M. thermoacetica because its overall genome sequence characteristics were similar to
those of M. thermoacetica strain ATCC39073. Y72 was confirmed to carry all the genes encoding
the enzymes in the reductive acetyl-CoA pathway, with very high similarities to those of
ATCC39073. In addition, it was confirmed to assimilate carbon dioxide using this pathway.
However, although both Y72 and ATCC39073 carried common genes encoding several enzymes related
to the reductive tricarboxylic acid (TCA) cycle, their gene sets were different. Our results
suggested that the reason for higher transformation efficiency in Y72 than that in ATC09073, a
reference strain of M. thermoacetica, may be that Y72 possesses only 2 sets of genes
considered to be involved in a restriction-modification system, which was half of those found
in ATCC39073.

<>

<1>Tsukahara, S., Kim, S.-G., Takaku, H.
<2>Inhibition of restriction endonuclease cleavage site via triple helix formation by homopyrimidine phosphorothioate oligonucleotides.
<3>Biochem. Biophys. Res. Commun.
<4>196
<5>990-996
<6>1993
<7>The ability of pyrimidine rich oligonucleotide phosphorothioate to form stable triple helical
structures with the sequence containing the recognition site for the class IIS restriction
enzyme Ksp632I was examined. First, we synthesized double strand oligonucleotides
corresponding to the SV40 sites and studied their interaction with homopyrimidine
oligodeoxyribonucleotides including replacement of the other chain either with PS group
(SO-ODNs) in the second nucleotide position (from 5'-terminus) and end capped with the PS
group at both 3'- and 5'-ends (S2O-ODNs). The resulting perfect DNA triplexes were detected
by gel-mobility shift. The phosphorothioate oligonucleotide analogues (SO-ODNs) and (S2O-ODNs)
were shown to inhibit enzymatic cleavage under conditions that allow for triple helix
formation. Inhibition is sequence-specific and occurs in the micromolar concentration range.
Of particular interest is the Sp-phosphorothioate analogue (Sp-SO-ODNs) which inhibited the
endonuclease more than the other phosphorothioate oligonucleotide analogues (Rp-SO-ODNs or
S2O-ODNs).

<>

<1>Tsukahara, S., Yamakawa, H., Takai, K., Takaku, H.
<2>Inhibition of restriction enzyme Ksp632I via triple helix formation by phosphorothioate oligonucleotides.
<3>Nucleosides and Nucleotides
<4>13
<5>1617-1626
<6>1994
<7>The ability of pyrimidine-rich oligonucleotide phosphorothioate to form stable triple helical
structures with the sequence containing the recognition site for the class II-S restriction
enzyme Ksp632I was examined. First, we prepared double strand oligonucleotides corresponding
to the major groove of SV40 DNA at 17 base pair homopurine-homopyrimidine sequences, and
studied their interaction with homopyrimidine oligodeoxyribonucleotides including replacement
of the PS group in the second nucleotide position from the 5'-terminus (SO-ODNs) and of the
PS group at both the 3'- and 5'ends (S2O-ODNs). The resulting perfect DNA triplexes were
detected by the gel-mobility shift. The phosphorothioate oligonucleotide analogues (SO-ODNs)
and (S2O-ODNs) were shown to inhibit enzymatic cleavage under conditions that allow for triple
helix formation. Inhibition is sequence-specific and occurs in the micromolar concentration
range. Of particular interest is the Sp-phosphorothioate analogue (Sp-SO-ODNs) which inhibited
endonuclease more than the other phosphorothioate oligonucleotide analogues (Rp-SO-ODNs or
S2O-ODNs).

<>

<1>Tsumura, A., Hayakawa, T., Kumaki, Y., Takebayashi, S., Sakaue, M., Matsuoka, C., Shimotohno, K., Ishikawa, F., Li, E., Ueda, H.R., Nakayama, J., Okano, M.
<2>Maintenance of self-renewal ability of mouse embryonic stem cells in the absence of DNA methyltransferases Dnmt1, Dnmt3a and Dnmt3b.
<3>Genes Cells
<4>11
<5>805-814
<6>2006
<7>DNA methyltransferases Dnmt1, Dnmt3a and Dnmt3b cooperatively regulate cytosine methylation in
CpG dinucleotides in mammalian genomes,
providing an epigenetic basis for gene silencing and maintenance of
genome integrity. Proper CpG methylation is required for the normal
growth of various somatic cell types, indicating its essential role in
the basic cellular function of mammalian cells. Previous studies using
Dnmt1(-/-) or Dnmt3a(-/-)Dnmt3b(-/-) ES cells, however, have shown that
undifferentiated embryonic stem (ES) cells can tolerate hypomethylation
for their proliferation. In an attempt to investigate the effects of
the complete loss of CpG DNA methyltransferase function, we established
mouse ES cells lacking all three of these enzymes by gene targeting.
Despite the absence of CpG methylation, as demonstrated by genome-wide
methylation analysis, these triple knockout (TKO) ES cells grew
robustly and maintained their undifferentiated characteristics. TKO ES
cells retained pericentromeric heterochromatin domains marked with
methylation at Lys9 of histone H3 and heterochromatin protein-1, and
maintained their normal chromosome numbers. Our results indicate that
ES cells can maintain stem cell properties and chromosomal stability in
the absence of CpG methylation and CpG DNA methyltransferases.

<>

<1>Tsuru, T., Kawai, M., Mizutani-Ui, Y., Uchiyama, I., Kobayashi, I.
<2>Evolution of paralogous genes: Reconstruction of genome rearrangements through comparison of multiple genomes within Staphylococcus aureus.
<3>Mol. Biol. Evol.
<4>23
<5>1269-1285
<6>2006
<7>Analysis of evolution of paralogous genes in a genome is central to our understanding of
genome evolution. Comparison of closely related
bacterial genomes, which has provided clues as to how genome sequences
evolve under natural conditions, would help in such an analysis. With
species Staphylococcus aureus, whole-genome sequences have been decoded
for seven strains. We compared their DNA sequences to detect large
genome polymorphisms and to deduce mechanisms of genome rearrangements
that have formed each of them. We first compared strains N315 and Mu50,
which make one of the most closely related strain pairs, at the
single-nucleotide resolution to catalogue all the middle-sized (more
than 10 bp) to large genome polymorphisms such as indels and
substitutions. These polymorphisms include two paralogous gene sets,
one in a tandem paralogue gene cluster for toxins in a genomic island
and the other in a ribosomal RNA operon. We also focused on two other
tandem paralogue gene clusters and type I restriction-modification (RM)
genes on the genomic islands. Then we reconstructed rearrangement
events responsible for these polymorphisms, in the paralogous genes and
the others, with reference to the other five genomes. For the tandem
paralogue gene clusters, we were able to infer sequences for homologous
recombination generating the change in the repeat number. These
sequences were conserved among the repeated paralogous units likely
because of their functional importance. The sequence specificity (S)
subunit of type I RM systems showed recombination, likely at the
homology of a conserved region, between the two variable regions for
sequence specificity. We also noticed novel alleles in the ribosomal
RNA operons and suggested a role for illegitimate recombination in
their formation. These results revealed importance of recombination
involving long conserved sequence in the evolution of paralogous genes
in the genome.

<>

<1>Tsurumaru, H., Kanesaki, Y., Hashimoto, S., Okizaki, K., Yoshikawa, H., Yamakawa, T.
<2>Draft Genome Sequence of Bradyrhizobium japonicum Is-34, Which Is Incompatible with Rj4 Genotype Soybeans.
<3>Genome Announcements
<4>2
<5>e01316-14
<6>2014
<7>We report here the draft genome sequence of Bradyrhizobium japonicum Is-34, which is
incompatible with Rj4 genotype soybeans. A candidate gene involved in this
incompatibility was found to be present in this genome.

<>

<1>Tsutakawa, S.E., Jingami, H., Morikawa, K.
<2>Recognition of a TG mismatch: The crystal structure of very short patch repair endonuclease in complex with a DNA duplex.
<3>Cell
<4>99
<5>615-623
<6>1999
<7>The crystal structure of very short patch repair (Vsr) endonuclease, in complex with Mg2+ and
with duplex DNA containing a TG mismatch, has been determined at 2.3 A resolution. In E. coli,
the enzyme recognizes a TG mismatched base pair, generated after spontaneous deamination of
methylated cytosines, and cleaves the phosphate backbone on the 5' side of the thymine.
Extensive interactions between the DNA and the protein characterize a novel recognition
mechanism, where three aromatic residues intercalate from the major groove into the DNA to
strikingly deform the base pair stacking. With the presence of a cleaved DNA intermediate in
the active center, the structure of the Vsr/DNA complex provides detailed insights into the
catalytic mechanism for endonuclease activity.

<>

<1>Tsutakawa, S.E., Morikawa, K.
<2>The structural basis of damaged DNA recognition and endonucleolytic cleavage for very short patch repair endonuclease.
<3>Nucleic Acids Res.
<4>29
<5>3775-3783
<6>2001
<7>Endonucleases in DNA repair must be able to recognize damaged DNA as well as cleave the
phosphodiester backbone. These functional prerequisites are manifested in very short patch
repair (Vsr) endonuclease through a common endonuclease topology that has been tailored for
recognition of TG mismatches. Structural and biochemical comparison with type II restriction
enzymes illustrates how Vsr resembles these endonucleases in overall topology but also how Vsr
diverges in terms of the detailed catalytic mechanism. A histidine and two metal-water
clusters catalyze the phosphodiester cleavage. The mode of DNA damage recognition is also
unique to Vsr. All other structurally characterized DNA damage-binding enzymes employ a
nucleotide flipping mechanism for substrate recognition and for catalysis. Vsr, on the other
hand, recognizes the TG mismatch as a wobble base pair and penetrates the DNA with three
aromatic residues on one side of the mismatch. Thus, Vsr endonuclease provides important
counterpoints in our understanding of endonucleolytic mechanisms and of damaged DNA
recognition.

<>

<1>Tsutakawa, S.E., Morikawa, K.
<2>New recognition mode for a TG mismatch: The atomic structure of a very short patch repair endonuclease-DNA complex.
<3>Cold Spring Harb. Symp. Quant. Biol., Cold Spring Harbor Laboratory Press, , Cold Spring Harbor
<4>65
<5>233-239
<6>2000
<7>The crystal structure determination of the T4 endo-V pyrimidine photodimer DNA glycosylase
provided the first direct view of DNA lesion recognition by a repair enzyme.  Similar damaged
DNA recognition modes, involving nucleotide flipping, were observed in various base excision
repair enzymes.  The implications of nucleotide flipping raised the new question of how DNA
endonucleases other than base excision repair enzymes recognize mismatched base pairs.  The
Escherichia coli very short patch repair endonuclease is a good target to address this
question in terms of three-dimensional structures.  The Vsr endonuclease is involved in the
initial reaction for the repair of mismatched TG base pairs generated through spontaneous
deamination of methylated cytosine.  This enzyme recognizes a TG mismatch within the duplex
5'CT(A/T)GG, where the second T forms the mismatch and all of the other bases are in standard
Watson-Crick base pairing.  It catalyzes the cleavage at the 5' side of the thymine, leaving
a 5' phosphate and a 3' hydroxyl at the termini.  The crystal structure of a truncated form
of this endonuclease was determined at 1.8 angstroms resolution.  The protein was found to
contain one structural zinc-binding module.  Unexpectedly, its overall topology resembles
members of the type II restriction endonuclease family, although the catalytic center with
critical histidines is distinct from those of restriction enzymes.  More recently, the crystal
structure of Vsr endonuclease in complex with DNA has been determined at 2.3 angstroms
resolution.  This endonuclease has been found to employ a novel mismatch base-pair recognition
scheme that does not involve base flipping-out.  Extensive interactions between the DNA and
the protein characterize the recognition mechanism, where three aromatic residues intercalate
from the major groove into the DNA to strikingly deform the base-pair stacking.  An
amino-terminal alpha-helix is accommodated into the expanded minor groove so that the amino
acid side chains make additional contacts with the DNA duplexes.  With the presence of a
cleaved DNA intermediate in the active center, the structure of the Vsr/DNA complex provides
detailed insights into the catalytic mechanism for endonuclease activity.

<>

<1>Tsutakawa, S.E., Muto, T., Kawate, T., Jingami, H., Kunishima, N., Ariyoshi, M., Kohda, D., Nakagawa, M., Morikawa, K.
<2>Crystallographic and functional studies of very short patch repair endonuclease.
<3>Mol. Cell
<4>3
<5>621-628
<6>1999
<7>Vsr endonuclease plays a crucial role in the repair of TG mismatched base pairs, which are
generated by the spontaneous degradation of methylated cytidines; Vsr recognizes the
mismatched base pair and cleaves the phosphate backbone 5' to the thymidine. We have
determined the crystal structure of a truncated form of this endonuclease at 1.8 A resolution.
The protein contains one structural zinc-binding module. Unexpectedly, its overall topology
resembles members of the type II restriction endonuclease family. Subsequent mutational and
biochemical analyses showed that certain elements in the catalytic site are also conserved.
However, the identification of a critical histidine and evidence of an active site
metal-binding coordination that is novel to endonucleases indicate a distinct catalytic
mechanism.

<>

<1>Tsvetkova, N.V.
<2>Development of rational schemes of the purification of restriction endonucleases.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>7
<5>77-81
<6>1987
<7>The work deals with the search for rational schemes of the purification of
endonucleases, suitable for the isolation of different enzymes.  The scheme
using the combination of Blue Sepharose and phosphocellulose has proved to be
most universal.  The expediency of starting the purification of Haemophilus
enzymes on Biogel A-0.5 m has been established.

<>

<1>Tsvetkova, N.V., Mileikovskaya, M.M., Gruber, I.M., Polyachenko, V.M., Butkus, V.V., Janulaitis, A., Sudzhyuvene, O.F., Tarasov, A.P.
<2>Purification and determination of the substrate specificity of restriction endonuclease BcmI.
<3>Mol. Gen. Mikrobiol. Virusol.
<4>0
<5>19-22
<6>1987
<7>The vigorous development of recombinant DNA and biotechnology is leading to
growing requirements for varied restriction enzymes; therefore the search for
new specific endonucleases remains urgent.  The restriction endonuclease BcmI
has been isolated from a Bacillus strain.  It was purified by chromatography on
blue Sepharose, phosphocellulose P-II, and heparin-Sepharose.  The new domestic
adsorbent, orange Sepharose, can also be used for the purification of the
restriction endonuclease.  The nucleotide sequence recognized by BcmI was
determined by a comparison of the nature of the digestion of DNA by known
enzymes and by the investigated enzyme.  The site of cleavage of the substrate
was determined by direct methods on pBR322 DNA.  The aggregate of data obtained
permits us to assert that the restriction endonuclease BcmI is a true
isoschizomer of the restriction endonuclease ClaI.

<>

<1>Tsymbal, N.V., Samoilenko, V.A., Tovkach, F.I.
<2>Site-specific endonuclease activity of filamentous cyanobacterium Plectonema boryanum.
<3>Dopov. Nats. Akad. Nauk Ukr.
<4>0
<5>146-149
<6>2011
<7>The site-specific endonuclease PboI was isolated from the filamentous cyanobacterium
Plectonema boryanum.  The purification included successive column chromatographies on
DEAE-Toyopearl, phosphocellulose, and blue sepharose.  Purified enzyme recognizes and cleaves
GGATCC-palindrome sequence is analogously to BamHI.  The enzyne belongs to type II restriction
endonucleases.  The enzyme activity is optimal at a temperature 45oC, pH 7.5, and ionic
stength 100mM.

<>

<1>Tu, C.-P.D., Jay, E., Bahl, C.P., Wu, R.
<2>A reliable mapping method for sequence determination of oligodeoxyribonucleotides by mobility shift analysis.
<3>Anal. Biochem.
<4>74
<5>73-93
<6>1976
<7>The method for sequence analysis of large oligodeoxyribonucleotides based on
the characteristic mobility shifts of their sequential partial degradation
products on two-dimensional homochromatography has been perfected using a large
number of synthetic oligodeoxyribonucleotides of defined sequences as
standards.  Flat bed electrophoresis with careful temperature control gave
entirely reproducible mobilities in the first dimension.  Using this
information, an accurate formula has been derived for calculating the relative
electrophoretic mobilities of oligodeoxyribonucleotides of any composition.
This formula is used to calculate the mobility shifts between two consecutive
oligodeoxyribonucleotides in a series of partial products of an unknown
oligomer distributed in the two-dimensional homochromatogram which differ by
one nucleotide in length.  This is compared with the observed mobility shift
value to identify the added nucleotide.  This provides a direct and rapid
method for obtaining the unambiguous sequence of an entire
oligodeoyribonucleotide up to 15 nucleotides in length.

<>

<1>Tu, C.-P.D., Roychoudhury, R., Wu, R.
<2>Nucleotide recognition sequence at the cleavage site of Haemophilus aegyptius II (HaeII) restriction endonuclease.
<3>Biochem. Biophys. Res. Commun.
<4>72
<5>355-362
<6>1976
<7>We have determined the nucleotide sequence recognized by the restriction
endonuclease HaeII from Haemophilus aegyptius which cleaves the simian virus 40
(SV40) DNA at a single specific site.  By using terminal radioactive labeling
of the cleavage site at both the 5' and 3'-ends we have deduced the recognition
sequence, 3'...A-G-C-G-C^pT...3' 3'..Tp^C-G-C-G--A...5' with elements of a
two-fold rotational symmetry.  The endonuclease produces staggered ends with
protruding 3'-terminated single-strands, four nucleotides in length.  In
plasmid RSF 2124 DNA, which contains multiple HaeII cleavage sites, it was
observed that the 5th nucleotide from the 3' terminus is either a pdA or a pdG,
indicating alternating recognition sequences.

<>

<1>Tucholski, J., Skowron, P.M., Podhajska, A.J.
<2>MmeI, a class-IIS restriction endonuclease: purification and characterization.
<3>Gene
<4>157
<5>87-92
<6>1995
<7>Two restriction endonucleases, MmeI and MmeII, from Methylophilus methylotrophus were purified
to homogeneity.  Both enzymes belong to the class-II restriction endonucleases (ENases) but
exhibit very different enzymatic and physical properties.  MmeII is a typical member of
class-II ENases.  It is a polymeric protein composed of 50-kDa subunits.  In contrast to
MmeII, MmeI is a monomeric protein of 101 kDa, cleaving a DNA molecule 20/18 nucleotides away
from the asymmetric recognition sequence (5'-TCCRAC-3'); therefore, it is classified as a
member of subclass-IIS.  MmeI has a pI of 7.85 and is active in the pH range 6.5 to 10 with
the optimum at 7 to 8.  Increasing salt concentration creates an inhibitory effect on MmeI: 40
mM KCl decreases activity by 50%, 100 mM completely inhibits DNA cleavage.  Tris.HCl (pH
7.5)  at a concentration exceeding 20 mM inhibits MmeI activity.  Mg2+ stimulates MmeI in the
range  of 0.2 to 35 mM, with the optimum between 0.5 and 10 mM.

<>

<1>Tucholski, J., Zmijewski, J.W., Podhajska, A.J.
<2>Two intertwined methylation activities of the MmeI restriction-modification class-IIS system from Methylophilus methylotrophus.
<3>Gene
<4>223
<5>293-302
<6>1998
<7>The class-IIS restriction endonuclease, R.MmeI, was isolated from Methylophilus
methylotrophus.  It was originally described as a monomeric enzyme, with the native Mr
105,000+/-7,000, which did not cleave DNA efficiently.  However, it was discovered that R.MmeI
endonucleolytic activity is enhanced by S-adenosyl-L-methionine and sinefungin, an analogue of
AdoMet.  Surprisingly, the purified R.MmeI endonuclease was found to have a second enzymatic
activity, namely methylation of the adenine residue to N6-methyladenine in the top strand of
the MmeI-recognition sequence, 5'-TCCR*AC-3' (*A = meA).  The R.MmeI methylating activity
requires AdoMet and is increased in the presence of several divalent cations, 20-fold by Mg2+
or Ca2+, and less by Mn2+, Zn2+ and Co2+; however, methylation is inhibited entirely by
sinefungin, at concentrations above 9 microM.  The latter observation shows that the enhancing
effect of AdoMet or sinefungin on the DNA cleavage was not related to the process of DNA
methylation.  Furthermore, a second component of the MmeI restriction-modification system, an
M.MmeI methyltransferase, was isolated and purified.  The M.MmeI protein was found to have an
Mr of 48,000 +/- 2,000 (under denaturing conditions) and to methylate both adenine residues
(*A) in the MmeI-recognition sequence 5'-TCCR*AC-3'/3'-*AGGYTG-5'.  Methylation of the top
strand does not inhibit the DNA cleavage by R.MmeI, whereas methylation of both DNA strands
blocks the cleavage process.

<>

<1>Tucker, K.L.
<2>A genetic investigation of the establishment of genomic imprinting.
<3>Ph.D. Thesis, MIT, USA
<4>
<5>1-80
<6>1997
<7>Much evidence indicates that in vertebrates the methylation of DNA at cytosine residues
correlates with gene inactivity.  A single methyltransferase activity, DNA
(cytosine-5)-methyltransferase, has been purified from mammalian cells and its gene (Dnmt)
cloned.  The crucial importance of DNA methylation in development was demonstrated by the
targeted mutation of Dnmt in murine embryonic stem cells.  Embryos with homozygous disruptions
of Dnmt die in mid-gestation.  In contrast, homozygous mutant ES cells proliferate normally
with their DNA highly demethylated, though they die upon differentiation.  The research
presented in this thesis focused on expressing the Dnmt cDNA in the backgroud of Dnmt mutant
ES cells.  Initial attempts using a cDNA and a corresponding promoter region were unsuccessful
in effecting adequate levels of functional Mtase expression.  A complete cDNA sequence was
obtained through the cloning of two new 5'-proximal exons, which extend the open reading
frame up to 171 codons upstream of the previously defined start site.  This showed the
previously-reported promoter sequence to lie in the second intron of the gene, with no
evidence that it functions in ES cells.  It was found that the extra coding sequence was
necessary for functional expression of the gene in ES cells.  Expression of the wild type Dnmt
cDNA in mutant ES cells caused an increase in methylation of bulk DNA to normal levels, but
did not restore the methylation of imprinted genes, whose expression is monoallelic and
dependent upon their parental derivation.  The expression of these genes was deregulated
because of this hypomethylation, as shown previously in Dnmt mutant embryos.  Full restoration
of monoallelic methylation and expression was imposed on imprinted genes upon germline
transmission.  These results are consistent with the presence of distanct de novo DNA
methyltransferase activities during oogenesis and spermatogenesis which specifically recognize
imprinted genes but which are absent in the post-implantation embryo and in ES cells.  Because
the "rescued" ES cells display biallelic hypomethylation and expression of imprinted genes,
their developmental capacities may be impaired, as is seen with cells derived from
parthenogenetic or androgenetic embryos, which also display biallelic expression of imprinted
genes.  Alternatively, the rescued ES cells may have normal developmental capacities, as
predicted by a model which explains genomic imprinting as a nonessential,
evolutionarily-recent addition to the basic mammalian developmental program.  Rescued cells
differentiated normally in vitro, formed teratomas with a wide variety of differentiated cell
types, and contributed substantially to coat color in adult chimeras.  Further analysis of
tissue-specific distribution of rescued ES cells in chimeras will allow a full assessment of
the developmental potential of a mouse lacking an imprinted genome.

<>

<1>Tucker, K.L., Beard, C., Dausman, J., Jackson-Grusby, L., Laird, P.W., Lei, H., Li, E., Jaenisch, R.
<2>Germ-line passage is required for establishment of methylation and expression patterns of imprinted but not of nonimprinted genes.
<3>Genes Dev.
<4>10
<5>1008-1020
<6>1996
<7>Embryonic stem cells homozygous for a disruption of the DNA (cytosine-5)-methyltransferase
gene (Dnmt) proliferate normally with their DNA highly demethylated but die upon
differentiation.  Expression of the wild-type Dnmt cDNA in mutant male ES cells caused an
increase in methylation of bulk DNA and of the Xist and Igf2 genes to normal levels, but did
not restore the methylation of the imprinted genes H19 and Igf2r.  These cells differentiated
normally in vitro and contributed substantially to adult chimeras.  While the Xist gene was
not expressed in the remethylated male ES cells, no restoration of the normal expression
profile was seen for H19, Igf2r, or Igf2.  This indicates that ES cells can faithfully
reestablish normal methylation and expression patterns of nonimprinted genes but lack the
ability to restore those of imprinted genes.  Full restoration of monoallelic methylation and
expression was imposed on H19, Igf2, and Igf2r upon germ-line transmission.  These results are
consistent with the presence of distinct de novo DNA methyltransferase activities during
oogenesis and spermatogenesis, which specifically recognize imprinted genes but are absent in
the postimplantation embryo and in ES cells.

<>

<1>Tucker, K.L., Talbot, D., Lee, M.A., Leonhardt, H., Jaenisch, R.
<2>Complementation of methylation deficiency in embryonic stem cells by a DNA methyltransferase minigene.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>93
<5>12920-12925
<6>1996
<7>Previous attempts to express functional DNA cytosine methyltransferase (EC 2.1.1.37) in cells
transfected with the available Dnmt cDNAs have met with little or no success.  We show that
the published Dnmt sequence encodes an amino terminal-truncated protein that is tolerated only
at very low levels when stably expressed in embryonic stem cells.  Normal expression levels
were, however, obtained with constructs containing a continuation of an ORF with a coding
capacity of up to 171 amino acids upstream of the previously defined start site.  The protein
encoded by these constructs comigrated in SDS/PAGE with the endogenous enzyme and restored
methylation activity in transfected cells.  This was shown by functional rescue of Dnmt mutant
embryonic stem cells that contain highly demethylated genomic DNA and fail to differentiate
normally.  When transfected with the minigene construct, the genomic DNA became remethylated
and the cells regained the capacity to form teratomas that displayed a wide variety of
differentiated cell types.  Our results define an amino-terminal domain of the mammalian Mtase
that is crucial for stable expression and function in vivo.

<>

<1>Tuffery, P., Dessen, P., Mugnier, C., Hazout, S.
<2>Restriction map construction using a 'complete sentences compatibility' algorithm.
<3>Comput. Appl. Biosci.
<4>4
<5>103-110
<6>1988
<7>We have developed a new algorithm 'Complete sentences compatibility' (CSC)
which uses single and double digestion fragments to rapidly determine
restriction maps of circular DNA.  From possible combinations of fragments of
each simple digestion, which we call "sentences of decomposition", we construct
a restriction map which combines the sentences while taking into account
compatibility rules.  The algorithm can also deal with experimental errors of
fragment weight and can suggest solutions that account for non-readable bands
(fragments of zero length or multiple bands) on the gel.  Because experiments
using pairs of restrictive enzymes often result in multiple solutions, a
complementary algorithm tries to reduce the number of proposed solutions by
establishing consensus maps.  The restriction map construction algorithm was
tested on real cases, some containing more than fifteen fragments.  Execution
times range from 1-10 s on an IBM PC compatible microcomputer.

<>

<1>Tukel, C., Sanlibaba, P., Ozden, B., Akcelik, M.
<2>Identification of adsorption inhibition, restriction/modification and abortive infection type phage resistance systems in Lactococcus lactis strains.
<3>Acta Biol. Hung.
<4>57
<5>377-385
<6>2006
<7>98 Lactococcus lactis strains were isolated from traditional fermented milk products in Turkey
tested against 60 lactococcal lytic phages to
determine their resistance levels. While 82 L. lactis strains were
sensitive against lactic phages at different levels, 16 L. lactis
strains showed resistance to all phages tested. Types of phage
resistance among 16 L. lactis strains were identified as phage
adsorption inhibition in eight strains, restriction/modification in six
strains and abortive infection (heat sensitive phage resistance) in two
strains, using three broad-spectrum phages Phi p11 98-32, Phi pld 67-42
and Phi pld 67-44.

<>

<1>Tumbula, D.L., Makula, R.A., Whitman, W.B.
<2>Transformation of Methanococcus maripaludis and identification of a PstI-like restriction system.
<3>FEMS Microbiol. Lett.
<4>121
<5>309-314
<6>1994
<7>An optimized polyethylene glycol (PEG) method of transformation was developed for
Methanococcus maripaludis using the pKAS102 integration vector. The frequency of
transformation with 0.8 ug of plasmid and 3x10/9 cells was 4.8x10-5 transformants cfu-1, or
1.8x10/5 transformants per ug, which was four orders of magnitude greater than with the
natural transformation method. A PstI restriction activity in M. maripaludis was also
identified. Methylation of the plasmid with PstI methylase increased the methanococcal
transformation frequency at least four-fold. Also, chromosomal DNA from M. maripaludis was
resistant to digestion by the PstI endonuclease.

<>

<1>Tuohimaa, A., Riipinen, K.A., Brandt, K., Alatossava, T.
<2>The genome of the virulent phage Lc-Nu of probiotic Lactobacillus rhamnosus, and comparative genomics with Lactobacillus casei phages.
<3>Arch. Virol.
<4>151
<5>947-965
<6>2006
<7>The complete 36,466-bp genome sequence of the virulent phage Lc-Nu of probiotic Lactobacillus
rhamnosus was determined. The linear dsDNA with
a GC-content of 44.2% contained 3' single-stranded cohesive ends of 12
nucleotides. A total of 51 putative open reading frames (orfs) were
predicted. Lc-Nu showed to be evolutionary closely related to the
temperate Lactobacillus casei phages phi AT3 and A2. High DNA homology
with phi AT3 was shared over the late transcribed genes, and the
highest homology with A2 was within the genetic switch region. The
truncated cI-like repressor was the only lysogeny related gene left,
which strongly suggested Lc-Nu to be recently evolved from a temperate
origin. Three putative methylases and endonucleases were detected from
the region of early-transcribed genes. The putative origin of
replication within the putative gene orf34 homologous to replisome
organizers resembled to that of lambdoid phages. The present study
suggested Lc-Nu to be a new candidate for the proposed Sfi21-like
species.

<>

<1>Turk, P.W., Laayoun, A., Smith, S.S., Weitzman, S.A.
<2>DNA adduct 8-hydroxyl-2'-deoxyguanosine (8-hydroxyguanine) affects function of human DNA methyltransferase.
<3>Carcinogenesis
<4>16
<5>1253-1255
<6>1995
<7>8-Hydroxyl-2'-deoxyguanosine (also referred to as 8-hydroxyguanine [8-OH-dG] or
7,8-dihydro-8-oxoguanine), a common DNA adduct resulting from injury to DNA via reactive
oxygen species, affects the in vitro methylation of nearby cytosine moieties by the human DNA
methyltransferase. The exact position of 8-OH-deoxyguanosine relative to a CpG dinucleotide
appears important to this effect. Our data indicate that 8-OH-deoxyguanosine diminishes the
ability of the methyltransferase to methylate a target cytosine when the 8-OH-deoxyguanosine
is one or two nucleotides 3' from the cytosine, on the same strand. On the other hand
8-OH-deoxyguanosine does not diminish the ability of the enzyme to respond to a methyl
director (5-methylcytosine) when the 8-OH-deoxyguanosine is on the same strand but one or two
nucleotides 3' from the methyl director. Differences in methylation rates as great as 13-fold
have been detected using various 8-OH-deoxyguanosine-containing oligonucleotides as substrates
in methylation assays. Our findings suggest that oxidative damage of parental strand guanines
would permit normal copying of methylation patterns through maintenance methylation, while
oxidative damage of guanines in the nascent strand DNA would inhibit such methylation.

<>

<1>Turk, P.W., Weitzman, S.A.
<2>Free radical DNA adduct 8-OH-deoxyguanosine affects activity of HpaII and MspI restriction endonuclease.
<3>Free Radic. Res.
<4>23
<5>255-258
<6>1995
<7>8-OH-deoxyguanosine can diminish the ability of the restriction endonucleases HpaII and MspI
to cleave DNA.  The exact position of the adduct within the recognition site appears to
determine the extent of the effect.

<>

<1>Turmel, M., Boulanger, J., Schnare, M.N., Gray, M.W., Lemieux, C.
<2>Six group I introns and three internal transcribed spacers in the chloroplast large subunit ribosomal RNA gene of the green alga Chlamydomonas eugametos.
<3>J. Mol. Biol.
<4>218
<5>293-311
<6>1991
<7>The chloroplast large subunit rRNA gene of Chlamydomonas eugametos and its 5' flanking region
encoding tRNA-Ile (GAU) and tRNA-Ala (UGC) have been sequenced. The DNA sequence data along
with the results of a detailed RNA analysis disclosed two unusual features of this green algal
large subunit rRNA gene: (1) the presence of six group I introns (CeLSU.1-CeLSU.6) whose
insertion positions have not been described previously, and (2) the presence of three short
internal transcribed spacers that are post-transcriptionally excised to yield four rRNA
species of 380, 52, 810 and 1720 nucleotides, positioned in this order (5' to 3') in the
primary transcript. Together, these RNA species can assume a secondary structure that is
almost identical to that proposed for the 23S rRNA of Escherichia coli. All three internal
transcribed spacers map to variable regions of primary sequence and/or potential secondary
structure, whereas all six introns lie within highly conserved regions. The first three
introns are inserted within the sequence encoding the 810 nucleotide rRNA species and map
within domain II of the large subunit rRNA structure; the remaining introns, found in the
sequence encoding the 1720 nucleotide rRNA species, lie within either domain IV or V, as is
the case for all other large subunit rDNA introns that have been documented to date. CeLSU.5
and CeLSU.6 each contain a long open reading frame (ORF) of more than 200 codons. While the
CeLSU.6 ORF is not related to any known ORFs, the CeLSU.5 ORF belongs to a fmaily of ORFs that
have been identified in Podospora and Neuospora mitochondrial group I introns. The finding
that a polymorphic marker showing unidirectional gene conversion during crosses between C.
eugametos and Chlamydomonas moewusii is located within the CeLSU.5 ORF makes it likely that
this intron is a mobile element and that its ORF encodes a site-specific endonuclease
promoting the transfer of the intron DNA sequence.

<>

<1>Turmel, M., Cote, V., Otis, C., Mercier, J.-P., Gray, M.W., Lonergan, K.M., Lemieux, C.
<2>Evolutionary transfer of ORF-containing group I introns between different subcellular compartments (chloroplast and mitochondrion).
<3>Mol. Biol. Evol.
<4>12
<5>533-545
<6>1995
<7>We describe here a case of homologous introns containing homologous open reading frames (ORFs)
that are inserted at the same site in the large subunit (LSU) rRNA gene of different
organelles in distantly related organisms.  We show that the chloroplast LSU rRNA gene of the
green alga Chlamydomonas pallidostigmatica contains a group I intron (CpLSU.2) encoding a
site-specific endonuclease (I-CpaI).  This intron is inserted at the identical site
(corresponding to positions 1931-1932 of the Escherichia coli 23S rRNA sequence) as a group I
intron (AcLSU.m1) in the mitochondrial LSU rRNA gene of the amoeboid protozoon Acanthamoeba
castellanii.  The CpLSU.2 intron displays a rmarkable degree of nucleotide similarity in both
primary sequence and secondary structure to the AcLSU.m1 intron; moreover, the Acanthamoeba
intron contains an ORF in the same location within its secondary structure as the CpLSU.2 ORF
and shares with it a strikingly high level of amino acid similarity (65%; 42% identity).  A
comprehensive survey of intron distribution at site 1931 of the chloroplast LSU rRNA gene
reveals a rather restricted occurrence within the polyphyletic genus Chlamydomonas, with no
evidence of this intron among a number of non-Chlamydomonad green algae surveyed, nor in land
plants.  A parallel survey of homologues of a previously described and similar intron/ORF pair
(C. reinhardtii chloroplast CrLSU/A. castellanii mitochondrial AcLSU.m3) also shows a
restricted occurrence of this intron (site 2593) among chloroplasts, although the intron
distribution is somewhat broader than that observed at site 1931, with site-2593 introns
appearing in several green algal branches outside of the Chlamydomonas lineage.  The available
data, while not definitive, are most consistent with a relatively recent horizontal transfer
of both site-1931 and site-2593 introns (and their contained ORFs) between the chloroplast of
a Chlamydomonas-type organism and the mitochondrial of an Acanthamoeba-like organism, probably
in the direction chlorplast to mitochondrion.  The data also suggest that both introns could
have been acquired in a single event.

<>

<1>Turmel, M., Gutell, R.R., Mercier, J.-P., Otis, C., Lemieux, C.
<2>Analysis of the chloroplast large subunit ribosomal RNA gene from 17 Chlamydomonas taxa.
<3>J. Mol. Biol.
<4>232
<5>446-467
<6>1993
<7>Previous reports on the chloroplast large subunit rRNA genes of the two distantly related
green algae Chlamydomonas eugametos and Chlamydomonas reinhardtii indicate differences in the
distribution of group I introns and suggest a different arrangement of internal transcribed
spacers. To provide insights into the origin of these two types of intervening sequences, we
have undertaken the sequencing of the chlorplast rrnL genes of 15 additional Chlamydomonas
taxa and have characterized the mature large subunit rRNA species they encode in addition to
those specified by the C. reinhardtii rrnL. These analyses disclosed the presence of three
internal transcribed spaces sharing the same positions in all of the 17 taxa as well as the
presence of a total of 39 group I introns representing 12 insertion sites. Of these insertion
sites, only one has been identified in non-Chlamydomonas taxa. The distribution of
Chlamydomonas introns is highly variable and, in many respects, is not consistent with the
phylogeny deduced from chloroplast rRNA sequence comparisons. This phylogeny features two main
lineages of Chlamydomonas taxa forming sister groups. Because earlier branching organisms in
the green algal/land plant lineage display no chloroplast rRNA introns, it appears that all of
the intron insertion positions in Chlamydomonas are of recent origins, with some of the
positions having arisen subsequent to the divergence of the two main Chlamydomonas lineages.
Remarkably, the rRNA regions corresponding to most of the group I intron insertion positions
in rRNA genes have been assigned functional roles suggesting that they lie in exposed regions
of the ribosome. On the basis of this striking correlation between exposed rRNA regions and
intron insertion sites, we speculate that the reversal of the self-splicing reaction has
played a major role in the creation of the multiple intron insertion positions found in rRNA
genes as well as in the proliferation of group I introns elsewhere in the Chlamydomonas
chloroplast genome.

<>

<1>Turmel, M., Mercier, J.-P., Cote, V., Otis, C., Lemieux, C.
<2>The site-specific DNA endonuclease encoded by a group I intron in the Chlamydomonas pallidostigmatica chloroplast small subunit rRNA gene introduces a single-strand break at low concentrations of Mg2+.
<3>Nucleic Acids Res.
<4>23
<5>2519-2525
<6>1995
<7>Two group I introns (CpSSU.1 and CpSSU.2) that each potentially encode a protein with two
copies of the LAGLI-DADG motif were identified in the Chlamydomonas pallidostigmatica
chloroplast small subunit rRNA gene. They both belong to subgroup IA3 and represent novel
insertion positions in this gene (sites 508 and 793 in the Escherichia coli 16S rRNA). The
proteins encoded by the two introns were synthesized in vitro and tested for their ability to
cleave the homing site of their respective introns. Only the CpSSU.1-encoded protein (I-CpaII)
was found to display specific DNA endonuclease activity. At 0.1 mM MgCl2, I-CpaII nicks only
the bottom (transcribed) DNA strand, but at concentrations ranging from 0.5 to 5.0 mM, it
cleaves both DNA strands (leaving a 4 nucleotide single-stranded extension with 3'-OH
overhangs) while preferentially nicking the bottom strand. The rate of cleavage of the top
strand increases with increasing concentration of MgCl2. The preliminary data derived from
these endonuclease assays suggest that the mode of DNA cleavage by I-CpaII is directed by the
availability of Mg2+ and the affinity of different binding sites for this cation.

<>

<1>Turmel, M., Otis, C., Cote, V., Lemieux, C.
<2>Evolutionarily conserved and functionally important residues in the I-CeuI homing endonuclease.
<3>Nucleic Acids Res.
<4>25
<5>2610-2619
<6>1997
<7>Two approaches were used to discern critical amino acid residues for the function of the
I-CeuI homing endonuclease: sequence comparison of subfamilies of homologous proteins and
genetic selection.  The first approach revealed residues potentially involved in catalysis and
DNA recognition.  Because I-CeuI is lethal in Escherichia coli, enzyme variants not perturbing
cell viability were readily selected from an expression library.  A collection of 49 variants
with single amino acid substitutions at 37 positions was assembled.  Most of these positions
are clustered within or around the LAGLI-DADG dodecapeptide and the TQH sequence, two motifs
found in all protein subfamilies examined.  The Km and kcat values of the wild-type and nine
variant enzymes synthesized in vitro were determined.  Three variants, including one showing a
substituion of the glutamine residue in the TQH motif, revealed no detectable endonuclease
activity; five others showed reduced activity compared to the wild-type enzyme; whereas the
remaining variant cleaved the top strand about three times more efficiently than the
wild-type.  Our results not only confirm recent reports indicating that amino acids in the
LAGLI-DADG dodecapeptide are functionally critical, but they also suggest that some residues
outside this motif directly participate in catalysis.

<>

<1>Turna, J., Mikulasova, D., Timko, J., Zelinka, J.
<2>Purification of restriction endonuclease SauBMKI.
<3>Biologia (Bratisl)
<4>45
<5>281-288
<6>1990
<7>The type II restriction endonuclease designated SauBMKI was isolated from Streptomyces
aureofaciens, strain BMK.  The enzyme was purified by chromatography using DEAE-cellulose,
ssDNA cellulose and phosphocellulose to the grade when no unspecific endo- and exonucleases
were detectable.  The optimal reaction conditions were established: glycine 20 mmol/l (pH
8.7), NaCl 20 mmol/l, MgCl2 20 mmol/l, 2-ME 7 mmol/l and BSA 100 micrograms/ml.

<>

<1>Turner, D.P., Connolly, B.A.
<2>Interaction of the E-coli DNA G : T-mismatch endonuclease (vsr protein) with oligonucleotides containing its target sequence.
<3>J. Mol. Biol.
<4>304
<5>765-778
<6>2000
<7>The Escherichia coli vsr endonuclease recognises G:T base-pair mismatches in double-stranded
DNA and initiates a repair pathway by
hydrolysing the phosphate group 5' to the incorrectly paired T. The
enzyme shows a preference for G:T mismatches within a particular
sequence context, derived from the recognition site of the E. coli dcm
DNA-methyltransferase (CC[A/T]GG). Thus, the preferred substrate for
the vsr protein is (CT[A/T]GG), where the underlined T is opposed by a
dG base. This paper provides quantitative data for the interaction of
the vsr protein with a number of oligonucleotides containing G:T
mismatches. Evaluation of specificity constant (k(st)/K-D; k(st) = rate
constant for single turnover, K-D = equilibrium dissociation constant)
confirms vsr's preference for a G:T mismatch within a hemi-methylated
dcm sequence, i.e. the best substrate is a duplex (both strands written
in the 5'-3' orientation) composed of CT[A/T]CG and C-5Me[T/A]GG.
Conversion of the mispaired T (underlined) to dU or the d(5Me)C to dC
gave poorer substrates. No interaction was observed with
oligonucleotides that lacked a G:T mismatch or did not possess a dcm
sequence. An analysis of the fraction of active protein, by
"reverse-titration" (i.e. adding increasing amounts of DNA to a fixed
amount of protein followed by gel-mobility shift analysis) showed that
less than 1% of the vsr endonuclease was able to bind to the substrate.
This was confirmed using "competitive titrations" (where competitor
oligonucleotides are used to displace a P-32-labelled nucleic acid from
the vsr protein) and burst kinetic analysis. This result is discussed
in the light of previous in vitro and in vivo data which indicate that
the MutL protein may be needed for full vsr activity.

<>

<1>Turner, P.C., Yomano, L.P., Jarboe, L.R., York, S.W., Baggett, C.L., Moritz, B.E., Zentz, E.B., Shanmugam, K.T., Ingram, L.O.
<2>Optical mapping and sequencing of the Escherichia coli KO11 genome reveal extensive chromosomal rearrangements, and multiple tandem copies of the Zymomonas mobilis pdc and adhB genes.
<3>J. Ind. Microbiol. Biotechnol.
<4>39
<5>629-639
<6>2012
<7>Escherichia coli KO11 (ATCC 55124) was engineered in 1990 to produce ethanol by
chromosomal insertion of the Zymomonas mobilis pdc and adhB genes into E. coli W
(ATCC 9637). KO11FL, our current laboratory version of KO11, and its parent E.
coli W were sequenced, and contigs assembled into genomic sequences using optical
NcoI restriction maps as templates. E. coli W contained plasmids pRK1 (102.5 kb)
and pRK2 (5.4 kb), but KO11FL only contained pRK2. KO11FL optical maps made with
AflII and with BamHI showed a tandem repeat region, consisting of at least 20
copies of a 10-kb unit. The repeat region was located at the insertion site for
the pdc, adhB, and chloramphenicol-resistance genes. Sequence coverage of these
genes was about 25-fold higher than average, consistent with amplification of the
foreign genes that were inserted as circularized DNA. Selection for higher levels
of chloramphenicol resistance originally produced strains with higher pdc and
adhB expression, and hence improved fermentation performance, by increasing the
gene copy number. Sequence data for an earlier version of KO11, ATCC 55124,
indicated that multiple copies of pdc adhB were present. Comparison of the W and
KO11FL genomes showed large inversions and deletions in KO11FL, mostly enabled by
IS10, which is absent from W but present at 30 sites in KO11FL. The early KO11
strain ATCC 55124 had no rearrangements, contained only one IS10, and lacked most
accumulated single nucleotide polymorphisms (SNPs) present in KO11FL. Despite
rearrangements and SNPs in KO11FL, fermentation performance was equal to that of
ATCC 55124.

<>

<1>Turroni, F. et al.
<2>Genome analysis of Bifidobacterium bifidum PRL2010 reveals metabolic pathways for host-derived glycan foraging.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>19514-19519
<6>2010
<7>The human intestine is densely populated by a microbial consortium whose
metabolic activities are influenced by, among others, bifidobacteria.
However, the genetic basis of adaptation of bifidobacteria to the human
gut is poorly understood. Analysis of the 2,214,650-bp genome of
Bifidobacterium bifidum PRL2010, a strain isolated from infant stool,
revealed a nutrient-acquisition strategy that targets host-derived
glycans, such as those present in mucin. Proteome and transcriptome
profiling revealed a set of chromosomal loci responsible for mucin
metabolism that appear to be under common transcriptional control and with
predicted functions that allow degradation of various O-linked glycans in
mucin. Conservation of the latter gene clusters in various B. bifidum
strains supports the notion that host-derived glycan catabolism is an
important colonization factor for B. bifidum with concomitant impact on
intestinal microbiota ecology.

<>

<1>Tuzikov, F.V., Tuzikova, N.A., Naumochkin, A.N., Zinovev, V.V., Malygin, E.G.
<2>Study of the equilibrium interation of T4 phage Dam-DNA-(N-adenine)-methyltransferase with substrates and ligands by fluorescence quenching method.
<3>Mol. Biol. (Mosk)
<4>31
<5>86-90
<6>1997
<7>
<>

<1>Tuzikov, F.V., Tuzikova, N.A., Naumochkin, A.N., Zinovev, V.V., Malygin, E.G.
<2>Fluorescence quenching study of equilibrium binding of phage T4 dam DNA-[N6-adenine]-methyltransferase with substrates and ligands.
<3>Mol. Biol. (Mosk)
<4>31
<5>73-76
<6>1997
<7>Selective fluorescence quenching by iodide ions was used to evaluate the surface accessibility
of tryptophan residues in T4 Dam DNA methyltransferase (T4 MTase).  Two of the three MTase
tryptophans proved fully accessible for iodide ions, suggesting that they are located near the
surface of the molecule.  Quenching of intrinsic fluorescence of T4 MTase tryptophans was used
to determine the parameters of enzyme binding with substrates (20-bp oligonucleotide duplex
with a GATC site, and S-adenosyl-L-methionine (SAM)) and substrate analogs (duplex without a
GATC site and S-adenosyl-L-homocysteine (SAH)).  Enzyme binding with small ligands (SAM and
SAH) conforms with the modified Sterm-Volmer equation applicable to 1:1 complexes.  The
dissociation constants for SAM and SAH were 0.24 and 2.3 microM, respectively.  In both cases,
about one third of the total Mtase fluorescence could be quenched by these ligands.
Saturating binding of oligonucleotide duplexes resulted in 12% (oligonucleotide with GATC) or
19% (without GATC) quenching.  In both cases, experimental quenching curves did not fit the
modified Sterm-Volmer equation, suggesting that the mechanism of oligonucleotide binding is
more complex than that for small ligands.

<>

<1>Tuzikov, F.V., Zinovev, V.V., Vavilin, V.I., Malygin, E.G., Buryanov, Y.I., Baev, A.A.
<2>Interaction between Ecodam methylase and double-stranded oligodeoxyribonucleotides.
<3>Dokl. Akad. Nauk.
<4>304
<5>231-234
<6>1989
<7>None

<>

<1>Tweedie, S., Ng, H.-H., Barlow, A.L., Turner, B.M., Hendrich, B., Bird, A.
<2>Vestiges of a DNA methylation system in Drosophila melanogaster?
<3>Nat. Genet.
<4>23
<5>389-390
<6>1999
<7>The insect Drosophila melanogaster belongs to an atypical group of animals with no detectable
genomic 5-methylcytosine.  We found, unexpectedly, that the Drosophila genome potentially
encodes two proteins that resemble a cytosine DNA methyltransferase and a mammalian
methyl-CpG-binding -domain protein, respectively.  The hypothetical DNA methyltransferase,
dDNMT, is closely related to the pmt1/DNMT2 family identified in fission yeast and mouse.

<>

<1>Twomey, D.P., Davis, R., Daly, C., Fitzgerald, G.F.
<2>Sequence of the gene encoding a second ScrFI m5C methyltransferase of Lactococcus lactis.
<3>Gene
<4>136
<5>205-209
<6>1993
<7>The nucleotide (nt) sequence of a second methyltransferase (MTase) encoding gene associated
with the ScrFI restriction/modification (R/M) system (5'CCNGG3') of Lactococcus lactis ssp.
cremoris UC503 has been determined. The coding region was 1080 nt in length and capable of
specifying a 41,847-Da protein of 360 amino acids (aa). The deduced aa sequence displayed
motifs characteristic of all prokaryotic 5-methylcytosine (m5C) MTases. Significant similarity
was observed with other m5C MTases, apart from the variable region which has been deemed
responsible for target sequence specificity. A larger than normal spacing between motif II and
motif III, comparable in length and bearing homology to a similar region in the SssI MTase,
was observed. A second open reading frame (ORF) of unknown function, which has the capacity to
encode a protein of 155 aa, was observed three nt upstream from the m5C MTase ORF.

<>

<1>Twomey, D.P., Davis, R., Daly, C., Fitzgerald, G.F.
<2>Molecular characterisation of the ScrFI restriction-modification system.
<3>Irish J. Agr. Food Res.
<4>32
<5>217
<6>1993
<7>The elucidation of the genetic determinants of bacteriophage resistance assists rational
approaches to designing and improving cheese-starter cultures. Lactococcus lactis subsp.
cremoris UC503 exhibits restriction-modification (R/M) activity, through the restriction
endonuclease, ScrFI, and the products of two methylase genes. Both of these methylases
independently confer resistance to ScrFI digestion in E. coli ED8739. These genes, designated
mscrFIA and mscrFIB, have been cloned from the chromosome of strain UC503 and their nucleotide
sequences determined. Both of the predicted amino acid sequences display 10 motifs with an
alignment and architecture characteristic of all procaryotic 5-methylcytosine methylases. The
so-called variable regions, located between motif VIII and motif IX, are responsible for the
target sequence specificity of these enzymes. However, the respective variable regions of
M.ScrFIA and M.ScrFIB display no significant similarity to each other, suggesting that the
enzymes have different recognition specificities. The variable region of M.ScrFIA displayed
substantial similarity to equivalent regions in M.EcoRII and dcm, both of which recognise 5'
CC(A/T)GG 3', a subset of the ScrFI endonuclease recognition site. In M.ScrFIA, a larger than
normal spacing between motifs II and III, comparable in length and bearing marginal homology
to a similar region in the SssI methylase, was observed. Recent evidence suggests that the
ScrFI restriction endonuclease gene resides in the 1 kb intervening region between mscrFIA and
mscrFIB. Furthermore, it is proposed that a gene (orfX) located three nucleotides upstream of
mscrFIB encodes a repair endonuclease which compensates for the inherent ability of
5-methylcytosine to deaminate spontaneously to thymine.

<>

<1>Twomey, D.P., De Urraza, P.J., McKay, L.L., O'Sullivan, D.J.
<2>Characterization of AbiR, a novel multicomponent abortive infection mechanism encoded by plasmid pKR223 of Lactococcus lactis subsp. lactis KR2.
<3>Appl. Environ. Microbiol.
<4>66
<5>2647-2651
<6>2000
<7>The native lactococcal plasmid pKR223 encodes two distinct phage resistance mechanisms, a
restriction and modification (R/M) system designated LlaKR2I and an abortive infection
mechanism (Abi) which affects prolate-headed-phage proliferation. The nucleotide sequence of a
16,174-bp segment of pKR223 encompassing both the R/M and Abi determinants has been
determined, and sequence analysis has validated the novelty of the Abi system, which has now
been designated AbiR. Analysis of deletion and insertion clones demonstrated that AbiR was
encoded by two genetic loci, separated by the LlaKR2I R/M genes. Mechanistic studies on the
AbiR phenotype indicated that it was heat sensitive and that it impeded phage DNA replication.
These data indicated that AbiR is a novel multicomponent, heat-sensitive, "early"-functioning
Abi system and is the first lactococcal Abi system described which is encoded by two separated
genetic loci.

<>

<1>Twomey, D.P., Gabillet, N., Daly, C., Fitzgerald, G.F.
<2>Molecular characterization of the restriction endonuclease gene (scrFIR) associated with the ScrFI restriction/modification system from Lactococcus lactis subsp. cremoris UC503.
<3>Microbiology
<4>143
<5>2277-2286
<6>1997
<7>The nucleotide sequence of the chromosomally encoded type II ScrFI restriction/modification
system from Lactococcus lactis subsp. cremoris UC503 was completed.  The ScrFI restriction
endonuclease has previously been shown to specifically recognize 5' CCNGG 3' sites, cleaving
after the second cytosine and the degenerate central base.  The Enase gene was located
between, and co-directionally transcribed with, two formerly characterized 5-methylcytosine
methyltransferase genes, which encode proteins that independently confer protection against
ScrFI digestion.  scrFIR codes for a protein of 272 amino acids with a predicted molecular
mass of 31470 Da, which agrees favorably with a previously estimated molecular mass of 34 kDa
for this enzyme.  The deduced sequence of this protein did not show any significant homology
with known protein sequences, including the isoschizomeric SsoII Enase from Shigella sonnei.
The Enase gene was cloned and expressed in Escherichia coli and lactococcus; however, no in
vivo restriction of phage was observed, suggesting that expression of the Enase gene may be
repressed, or that the appropriate expression signals may be absent in the cloned constructs.
The ability of ScrFi to cleave non-canonically modified 5'CCNGG3' sequences suggested that
some ScrFI sites may require complex modifications to fully impair digestion by this enzyme.

<>

<1>Twomey, D.P., McKay, L.L., O'Sullivan, D.J.
<2>Molecular characterization of the LlaKR2I restriction and modification system from Lactococcus lactis ssp. lactis KR2.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>97
<5>426
<6>1997
<7>Lactococcus lactis ssp. lactis KR2 contains at least two plasmid-encoded restriction and
modification (R/M) systems and the genetic determinants of one of these systems, designated
LlaKR2I, have been localized on the 36 kb plasmid, pKR223, and sequenced.  The LlaKR2I R/M
genes were found to be divergently transcribed with respect to each other, with a complete
copy of the IS-like element, IS982, located in the intergenic region.  The deduced protein
sequence of the modification component displayed greatest identity with M.Sau3AI throughout
the entire length of the protein.  Both methylases contained motifs characteristic of
5-methylcytosine methylases and shared extensive homology throughout their predicted target
recognition domains, regions responsible for the sequence specificity of these proteins.  In
pKR223, and various derivatives, M.LlaKR2I prevented Sau3AI from digesting at 5' GATC 3'
sites.  Furthermore, since the deduced protein sequence of the LlaKR2I endonuclease shared
significant homology with Sau3AI, the specificity of the LlaKR2I R/M system is believed to be
5' GATC 3', with the cytosine residue the target base for methylation.  The small
isometric-headed phage, osk11, was only modestly restricted on lactococcal cells harboring the
LlaKR2I R/M system and despite the presence of 5' GATC 3' sites in the genome of the
prolate-headed phage oc2, no detectable restriction occurred.  The poor in vivo restriction
may have resulted from either decreased expression of the endonuclease and/or enhanced
expression of the methylase and may be due, in part, to the presence of IS982 located between
the endonuclease and methylase genes, whose putative transposase gene is co-directionally
transcribed with the methylase gene.

<>

<1>Twomey, D.P., McKay, L.L., O'Sullivan, D.J.
<2>Molecular characterization of the Lactococcus lactis LlaKR2I restriction-modification system and effect of an IS982 element positioned between the restriction and modification genes.
<3>J. Bacteriol.
<4>180
<5>5844-5854
<6>1998
<7>The nucleotide sequence of the plasmid-encoded LlaKR2I restriction-modification system of
Lactococcus lactis subsp. lactis biovar diacetylactis KR2 was determined.  This R-M system
comprises divergently transcribed endonuclease (llaKR2IR) and methyltransferase llaKR2IM)
genes; located in the intergenic region is a copy of the insertion element IS982, whose
putative transposase gene is codirectionally transcribed with llaKR2IM.  The deduced sequence
of the LlaKR2I endonuclease shared homology with the type II endonuclease Sau3AI and with the
MutH mismatch repair protein, both of which recognize and cleave the sequence 5'GATC3'.  In
addition, M.LlaKR2I displayed homology with the 5-methycytosine methyltransferase family of
proteins, exhibiting greatest identity with M.Sau3AI.  Both of these proteins shared notable
homology throughout their putative target recognition domains.  Furthermore, subclones of the
native parental lactococcal plasmid pKR223, which encode M.LlaKR2I, all remained undigested
after treatment with Sau3AI despite the presence of multiple 5'GATC3' sites.  The
combination of these data suggested that the specificity of the LlaKR2I R-M system was likely
to be 5'GATC3', with the cytosine residue being modified to 5-methylcytosine.  The IS982
element located within the LlaKR2I R-M system contained at its extremities two 16-bp perfect
inverted repeats flanked by two 7-bp direct repeats.  A perfect extended promoter consensus,
which represented the likely original promoter of the llaKR21I gene, was shown to overlap the
direct repeat sequence on the other side of IS982.  Specific deletion of IS982 and one of
these direct repeats via a PCR strategy indicated that the LlaKR2I R-M determinants do not
rely on elements within IS982 for expression and that the efficiency of bacteriophage
restriction was not impaired.

<>

<1>Tyndall, C., Lehnherr, H., Sandmeier, U., Kulik, E., Bickle, T.A.
<2>The type IC hsd loci of the enterobacteria are flanked by DNA with high homology to the phage P1 genome: implications for the evolution and spread of DNA restriction systems.
<3>Mol. Microbiol.
<4>23
<5>729-736
<6>1997
<7>EcoR124I, EcoDXXI and EcoprrI are the known members of the type IC family of DNA restriction
and modification systems.  The first three are carried on large conjugative plasmids, while
EcoprrI is chromosomally encoded.  The enzymes are coded by three genes, hsdR, hsdM and hsdS.
Analysis of the DNA sequences upstream and downstream of the type IC hsd loci shows that all
are highly homologous to each other and also to sequences present in the bacteriophage P1
genome.  The upstream sequences include functional phd and doc genes, which encode an
addiction system that stabilizes the P1 prophage state, and extend to and beyond pac, the site
at which phage DNA packaging begins.  Downstream of the hsd loci, P1 DNA sequences begin at
exactly the same place for all of the systems.  For EcoDXXI and EcoprrI the P1 homology
extends for thousands of base pairs while for EcoR124I an IS1 insertion and an associated
deletion have removed most of the P1-homologous sequences.  The significance of these results
for the evolution of DNA restriction and modification systems is discussed.

<>

<1>Tyndall, C., Meister, J., Bickle, T.A.
<2>The Escherichia coli prr region encodes a functional type IC DNA restriction system closely integrated with an anticodon nuclease gene.
<3>J. Mol. Biol.
<4>237
<5>266-274
<6>1994
<7>The prr locus was originally described as coding a ribonuclease that is activated after phage
T4 infection to cut within the anticodon of a specific tRNA, inactivating protein synthesis
and thus blocking phage development. Wild-type T4 phage has two genes coding the enzymes
polynucleotide kinase and RNA ligase, whose only function seems to be to repair the damage
done by the anticodon nuclease. As the only apparent function of the prr ribonuclease is to
combat phage infection, it can be considered as an RNA-based restriction enzyme. In
non-infected cells, the prr enzyme is kept inactive in a complex with three other proteins
which were predicted on the basis of DNA homologies to be the subunits of a type IC DNA
restriction and modifiction system. Unlike other type IC systems so far characterized, prr is
chromosomally rather than plasmid coded. However, sequences upstream from prr also have
homology with sequences from the plasmid R124 and the prophage P1. We have now investigated
the prr system and shown that it is indeed a bona fide type IC system which we call EcoprrI,
and which is active both in vivo and in vitro. The system is fully functional even in the
absence of the anticodon nuclease and seems to be a typical type I enzyme. EcoprrI recognizes
the sequence CCA(N7)RTGC. One peculiarity is that, with low efficiency, EcoprrI will recognize
and methylate variants of its recognition sequence such as CCT(N7)ATGC, which is methylated in
one strand of the DNA only.

<>

<1>Tyson, G.W., Chapman, J., Hugenholtz, P., Allen, E.E., Ram, R.J., Richardson, P.M., Solovyev, V.V., Rubin, E.M., Rokhsar, D.S., Banfield, J.F.
<2>Community structure and metabolism through reconstruction of microbial genomes from the environment.
<3>Nature
<4>428
<5>37-43
<6>2004
<7>Microbial communities are vital in the functioning of all ecosystems; however, most
microorganisms are uncultivated, and their roles in natural
systems are unclear. Here, using random shotgun sequencing of DNA from a
natural acidophilic biofilm, we report reconstruction of near-complete
genomes of Leptospirillum group II and Ferroplasma type II, and partial
recovery of three other genomes. This was possible because the biofilm was
dominated by a small number of species populations and the frequency of
genomic rearrangements and gene insertions or deletions was relatively
low. Because each sequence read came from a different individual, we could
determine that single-nucleotide polymorphisms are the predominant form of
heterogeneity at the strain level. The Leptospirillum group II genome had
remarkably few nucleotide polymorphisms, despite the existence of
low-abundance variants. The Ferroplasma type II genome seems to be a
composite from three ancestral strains that have undergone homologous
recombination to form a large population of mosaic genomes. Analysis of
the gene complement for each organism revealed the pathways for carbon and
nitrogen fixation and energy generation, and provided insights into
survival strategies in an extreme environment.

<>

<1>Uchida, K., Pyle, A.M., Morii, T., Barton, J.K.
<2>High resolution footprinting of EcoRI and distamycin with Rh(phi)2(bpy)3+, a new photofootprinting reagent.
<3>Nucleic Acids Res.
<4>17
<5>10259-10279
<6>1989
<7>The complex bis(phenanthrenequinone diimine)(bipyridyl)rhodium(III),
Rh(phi)2(bpy)3+, cleaves DNA efficiently in a sequence-neutral fashion upon
photoactivation so as to provide a novel, high resolution, chemical
photofootprinting reagent.  Photofootprinting of two crystallographically
characterized DNA-binding agents, distamycin, a small natural product which
binds to DNA in the minor groove, and the endonuclease EcoRI, which binds in
the major groove, gave respectively a 5-7 base pair footprint for the drug at
its A6 binding site and a 10-12 base pair footprint for the enzyme centered at
its recognition site (5'-GAATTC-3').  Both footprints agree closely with the
crystallographic results.  The photocleavage reaction can be performed using
either a high intensity lamp or, conveniently, a simple transilluminator box,
and the photoreaction is not inhibited by moderate concentrations of reagents
which are sometimes required for examining interactions of molecules with DNA.
When compared with other popular footprinting agents, the rhodium complex shows
a number of distinct advantages:  sequence-neutrality, high resolution, ability
to footrpint major as well as minor groove-binding ligands, applicability in
the presence of additives such as Mg2+ or glycerol, ease of handling, and a
sharply footprinted pattern.  Light activated footprinting reactions
furthermore offer the possibility of examining DNA-binding interactions with
time resolution and within the cell.

<>

<1>Uchida, Y., Shinomiya, T., Sato, S., Fukuda, K., Kobayashi, M.
<2>Restriction endonuclease preparation by breaking Thermus thermophilus mycelium in buffer, and removing impurities by column chromatography techniques.
<3>Japanese Patent Office
<4>JP 5998687 A
<5>
<6>1984
<7>Process comprises (1) breaking mycelium of Thermus thermophilus in a buffer solution, (2)
removing impurities from the solution and (3) purifying the solution by successive use of
column chromatography with cellulose phosphate, column chromatography with heparin-crosslinked
agarose gel, and column chromatography with hydroxyl apatite. Preferably the mycelia of the
microorganism are suspended in a buffer of pH 6.8, and broken by ultrasonication. Addition of
lysozyme accelerates the breaking of the cell wall.

<>

<1>Ueda, K., Yamashita, A., Ishikawa, J., Shimada, M., Watsuji, T.-o., Morimura, K., Ikeda, H., Hattori, M., Beppu, T.
<2>Genome sequence of Symbiobacterium thermophilum, an uncultivable bacterium that depends on microbial commensalisms.
<3>Nucleic Acids Res.
<4>32
<5>4937-4944
<6>2004
<7>Symbiobacterium thermophilum is an uncultivable bacterium isolated from compost that depends
on microbial commensalism. The 16S ribosomal DNA-based phylogeny suggests that this bacterium
belongs to an unknown taxon in the Gram-positive bacterial cluster. Here, we describe the 3.57
Mb genome sequence of S.thermophilum. The genome consists of 3338 protein-coding sequences,
out of which 2082 have functional assignments. Despite the high G + C content (68.7%), the
genome is closest to that of Firmicutes, a phylum consisting of low G + C Gram-positive
bacteria. This provides evidence for the presence of an undefined category in the
Gram-positive bacterial group. The presence of both spo and related genes and microscopic
observation indicate that S.thermophilum is the first high G + C organism that forms
endospores. The S.thermophilum genome is also characterized by the widespread insertion of
class C group II introns, which are oriented in the same direction as chromosomal replication.
The genome has many membrane transporters, a number of which are involved in the uptake of
peptides and amino acids. The genes involved in primary metabolism are largely identified,
except those that code several biosynthetic enzymes and carbonic anhydrase. The organism also
has a variety of respiratory systems including Nap nitrate reductase, which has been found
only in Gram-negative bacteria. Overall, these features suggest that S.thermophilum is
adaptable to and thus lives in various environments, such that its growth requirement could be
a substance or a physiological condition that is generally available in the natural
environment rather than a highly specific substance that is present only in a limited niche.
The genomic information from S.thermophilum offers new insights into microbial diversity and
evolutionary sciences, and provides a framework for characterizing the molecular basis
underlying microbial commensalism.

<>

<1>Uehara, H., Iwami, T., Kawaguchi, N., Abe, K., Sugimoto, N., Keshi, A., Eda, S., Matsuhisa, A.
<2>Probe for Detecting Pseudomonas aeruginosa and Methods using them.
<3>Japanese Patent Office
<4>JP 2003325181 A
<5>
<6>2003
<7>
<>

<1>Ueno, H., Ishino, Y., Kato, I.
<2>Genes encoding enzymes of BalI restriction-modification system.
<3>US Patent Office
<4>US 5789226
<5>
<6>1998
<7>An isolated BalI restriction enzyme gene and an BalI isolated modification enzyme gene are
disclosed.  A process for preparing a large amount of BalI restriction enzyme which comprises
culturing a transformant containing said genes and collecting BalI restriction enzyme from the
culture broth is also disclosed.

<>

<1>Ueno, H., Kasatsu-Shi, S.-K.
<2>Genes encoding enzymes of BalI restriction-modification system.
<3>European Patent Office
<4>EP 0725138 A
<5>
<6>1996
<7>An isolated BalI restriction enzyme gene and a BalI isolated modification enzyme gene are
disclosed.  A process for preparing a large amount of BalI restriction enzyme which comprises
culturing a transformant containing said genes and collecting BalI restriction enzyme from the
culture broth is also disclosed.

<>

<1>Ueno, H., Kato, I., Ishino, Y.
<2>Cloning and expression of the BalI restriction-modification system.
<3>Nucleic Acids Res.
<4>24
<5>2268-2270
<6>1996
<7>BalI, a type II restriction-modification (R-M) system from the bacterium,
Brevibacterium albidum, recognizes the DNA sequence 5'-TGGCCA-3'.  We cloned the genes
encoding the BalI restriction endonuclease and methyltransferase and expressed them in
Escherichia coli.  The two genes were aligned tail-to-tail and their termination codons
overlapped.
BalI restriction endonuclease and methyltransferase comprise 260 and 280 amino acids,
respectively, and have molecular weights of 29,043 and 31,999 Da.  The amino acid sequence of
BalI methyltransferase is similar to that of other m6A Mtases, although it has been
categorized as
an m5C methyltransferase.  A high expression system for the BalI restriction endonuclease was
constructed in E.coli for the production of large quantities of enzyme.

<>

<1>Ueno, H., Kita, A., Miyahara, M., Ishiwata, N., Mise, K., Kato, I., Ishino, Y.
<2>Production of new restriction endonuclease VpaK11BI recognizing sequence 5'-GGWCC-3' in a Haemolysin-less mutant of Vibrio parahaemolyticus.
<3>Biosci. Biotechnol. Biochem.
<4>61
<5>2129-2130
<6>1997
<7>High restriction endonuclease activity was found in a haemolysinless mutant of the Vibrio
parahaemolyticus 1743-1 strain.  The endonuclease, named VpaK11BI, recognized the palindromic
pentanucleotide sequence of 5'-GGWCC-3' and cleaved double-stranded DNA after the first G,
which is exactly the same as the specificity of AvaII.  The haemolysin-less mutant of V.
parahaemolyticus is now available for producing the valuable restriction endonuclease on a
commercial scale.

<>

<1>Ueno, H., Ueno, T., Oshima, A., Kato, I.
<2>Cloning and expression of Nsp7524III restriction and methylase from Nostoc strain PCC7524.
<3>Japanese Patent Office
<4>JP 97191885 A
<5>
<6>1997
<7>
<>

<1>Ueno, T., Ito, H., Kimizuka, F., Kotani, H., Nakajima, K.
<2>Gene structure and expression of the MboI restriction-modification system.
<3>Nucleic Acids Res.
<4>21
<5>2309-2313
<6>1993
<7>The genes from Moraxella bovis encoding the MboI restriction-modification system were cloned
and expressed in Escherichia coli. Three open reading frames were found in the sequence
containing the genes. These genes, which we named mboA, mboB, and mboC, had the same
orientation in the genome. Genes mboA and mboC encoded MboI methyltransferases (named M.MboA
and M.MboC) with 294 and 273 amino acid residues, respectively. The mboB gene coded for MboI
restriction endonuclease (R.MboI) with 280 amino acid residues. Recombinant E. coli-MBOI,
which contained the whole MboI system, overproduced R.MboI. R.MboI activity from E.coli-MBOI
was 480-fold that of M.bovis. The amino acid sequences deduced from these genes were compared
with those of other restriction-modification systems. The protein sequences of the MboI system
had 38-49% homology with those of the DpnII system.

<>

<1>Ueno, T., Ito, H., Kotani, H.
<2>Nsp7524V restriction endonuclease production and gene - Escherichia coli transformation.
<3>Japanese Patent Office
<4>JP 5227956
<5>
<6>1993
<7>
<>

<1>Ueno, T., Ito, H., Kotani, H., Kimizuka, F., Nakajima, K.
<2>Cloning and expression of the NspV restriction modification genes of Nostoc sp. strain PCC7524.
<3>Nucleic Acids Res.
<4>21
<5>3899
<6>1993
<7>The NspV restriction-modification system of the filamentous cyanobacterium Nostoc sp. strain
PCC7524 consists of NspV restriction endonuclease (R.NspV) and NspV modification enzyme
(M.NspV). R.NspV recognizes and cleaves double-stranded DNA at the sequence 5'-TTCGAA-3',
and M.NspV recognizes and protects this sequence by methylation against R.NspV. We cloned the
genes in Escherichia coli and analyzed their structure.

<>

<1>Ueno, T., Ito, H., Kotani, H., Nakajima, K.
<2>Restriction endonuclease MboI and modification DNA-methylase gene cloning - from Moraxella bovis, and expression in Escherichia coli.
<3>Japanese Patent Office
<4>JP 5276943
<5>
<6>1993
<7>
<>

<1>Ueno, T., Ito, H., Kotani, H., Nakajima, K.
<2>Nsp7524V restriction-modification genes.
<3>US Patent Office
<4>US 5441881
<5>
<6>1995
<7>To provide Nsp7524V restriction-modification genes and a method for producing Nsp7524V
restriction enzyme and Nsp7524V modification enzyme by using a novel microorganism having,
introduced thereinto, plasmids containing said genes.  Nsp7524V restriction-modification
genes.  A method for producing Nsp7524V restriction enzyme and/or Nsp7524V modification enzyme
which comprises incubating a microorganism carrying a plasmid having, integrated thereinto,
Nsp7524V restriction-modification genes, and recovering the Nsp7524V restriction enzyme and/or
Nsp7524V modification enzyme thus produced from the culture.  It becomes possible to
efficiently produce Nsp7524V restriction enzyme and/or Nsp7524V modification enzyme which are
useful in the field of genetic engineering.

<>

<1>Ueno, T., Ito, I., Kotani, H., Nakajima, K.
<2>Nsp7524V restriction-modification genes.
<3>Biotech. Advances
<4>14
<5>465
<6>1996
<7>To provide Nsp7524V restriction-modification genes and a method for producing Nsp7524V
restriction enzyme and Nsp7524V modification enzyme by using a novel microorganism having,
introduced thereinto, plasmids containing said genes, Nsp7524V restriction-modification genes.
A method for producing Nsp7524V restriction enzyme and/or Nsp7524V modification enzyme which
comprises incubating a microorganism carrying a plasmid having, integrated thereinto, Nsp7524V
restriction-modification genes, and recovering the Nsp7524V restriction enzyme and/or Nsp7524V
modification enzyme thus produced from the culture.  It becomes possible to efficiently
produce Nsp7524V restriction enzyme and/or Nsp7524V modification enzyme which are useful in
the field of genetic engineering.

<>

<1>Uetake, H., Toyama, S., Hagiwara, S.
<2>On the mechanism of host-induced modification:  Multiplicity activation and thermolabile factor responsible for phage growth restriction.
<3>Virology
<4>22
<5>202-213
<6>1964
<7>Salmonella phage sigma15 grown undergoes host-induced modification (HIM).
Phage sigma15[A] (phage sigma15 grown on Salmonella anatum, called strain A)
plates with low EOP on Salmonella butantan (=I-1) and vice versa.  The
restricting properties of the host cells on phage growth can be overcome by
multiple infection (multiplicity activation) with restricted normal phage or
with its temperature-sensitive mutant, but not with ultraviolet
(UV)-inactivated phage.  The phage-growth restricting factor is inactivated by
a short exposure of cells to slight heat, and its synthesis is inhibited by
chloramphenicol, but not by mitomycin C.  Experiments with P32-labeled phages
shown that the DNA of the restricted phage is degraded rapidly after injection
of the nonpermissive host, and that the degradation of injected DNA is
prevented by heating the host cells prior to infection or by UV irradiation of
the restricted phage.

<>

<1>Ugalde, J.A., Narasingarao, P., Kuo, S., Podell, S., Allen, E.E.
<2>Draft Genome Sequence of 'Candidatus Halobonum tyrrellensis' Strain G22, Isolated from the Hypersaline Waters of Lake Tyrrell, Australia.
<3>Genome Announcements
<4>1
<5>e01001-13
<6>2013
<7>We report the draft 3.675-Mbp genome sequence of 'Candidatus Halobonum tyrrellensis' strain
G22, a novel halophilic archaeon isolated from the surface
hypersaline waters of Lake Tyrrell, Australia. The availability of the first
genome from the 'Candidatus Halobonum' genus provides a new genomic resource for
the comparative genomic analysis of halophilic Archaea.

<>

<1>Uhlemann, A.C., Porcella, S.F., Trivedi, S., Sullivan, S.B., Hafer, C., Kennedy, A.D., Barbian, K.D., McCarthy, A.J., Street, C., Hirschberg, D.L., Lipkin, W.I., Lindsay, J.A., DeLeo, F.R., Lowy, F.D.
<2>Identification of a highly transmissible animal-independent Staphylococcus aureus ST398 clone with distinct genomic and cell adhesion properties.
<3>MBio
<4>3
<5>e00027-12
<6>2012
<7>A methicillin-resistant Staphylococcus aureus (MRSA) clone known as ST398 has
emerged as a major cause of acute infections in individuals who have close
contact with livestock. More recently, the emergence of an animal-independent
ST398 methicillin-sensitive S. aureus (MSSA) clone has been documented in several
countries. However, the limited surveillance of MSSA has precluded an accurate
assessment of the global spread of ST398 and its clinical relevance. Here we
provide evidence that ST398 is a frequent source of MSSA infections in northern
Manhattan and is readily transmitted between individuals in households. This
contrasts with the limited transmissibility of livestock-associated ST398
(LA-ST398) MRSA strains between humans. Our whole-genome sequence analysis
revealed that the chromosome of the human-associated ST398 MSSA clone is smaller
than that of the LA-ST398 MRSA reference strain S0385, due mainly to fewer mobile
genetic elements (MGEs). In contrast, human ST398 MSSA isolates harbored the
prophage varphi3 and the human-specific immune evasion cluster (IEC) genes chp
and scn. While most of the core genome was conserved between the human ST398 MSSA
clone and S0385, these strains differed substantially in their repertoire and
composition of intact adhesion genes. These genetic changes were associated with
significantly enhanced adhesion of human ST398 MSSA isolates to human skin
keratinocytes and keratin. We propose that the human ST398 MSSA clone can spread
independent of animal contact using an optimized repertoire of MGEs and adhesion
molecules adapted to transmission among humans. IMPORTANCE: Staphylococcus aureus
strains have generally been considered to be species specific. However,
cross-species transfers of S. aureus clones, such as ST398 methicillin-resistant
S. aureus (MRSA), from swine to humans have been reported. Recently, we observed
the emergence of ST398 methicillin-susceptible S. aureus (MSSA) as a colonizing
strain of humans in northern Manhattan. Here we report that ST398 is a frequent
cause of MSSA infections in this urban setting. The ST398 MSSA clone was readily
transmitted within households, independent of animal contact. We discovered that
human ST398 MSSA genomes were smaller than that of the LA-ST398 strain S0385 due
to fewer mobile genetic elements. Human and LA-ST398 strains also differed in
their composition of adhesion genes and their ability to bind to human skin
keratinocytes, providing a potential mechanism of S. aureus host adaptation. Our
findings illustrate the importance of implementing molecular surveillance of MSSA
given the evidence for the rapid and clinically undetected spread of ST398 MSSA.

<>

<1>Ukanis, M., Sapranauskas, R., Lubys, A.
<2>Screening for catalytically active Type II restriction endonucleases using segregation-induced methylation deficiency.
<3>Nucleic Acids Res.
<4>40
<5>e149
<6>2012
<7>Type II restriction endonucleases (REases) are one of the basic tools of recombinant DNA
technology. They also serve as models for elucidation of
mechanisms for both site-specific DNA recognition and cleavage by proteins.
However, isolation of catalytically active mutants from their libraries is
challenging due to the toxicity of REases in the absence of protecting
methylation, and techniques explored so far had limited success. Here, we present
an improved SOS induction-based approach for in vivo screening of active REases,
which we used to isolate a set of active variants of the catalytic mutant,
Cfr10I(E204Q). Detailed characterization of plasmids from 64 colonies screened
from the library of approximately 200 000 transformants revealed 29 variants of
cfr10IR gene at the level of nucleotide sequence and 15 variants at the level of
amino acid sequence, all of which were able to induce SOS response. Specific
activity measurements of affinity-purified mutants revealed >200-fold variance
among them, ranging from 100% (wild-type isolates) to 0.5% (S188C mutant),
suggesting that the technique is equally suited for screening of mutants
possessing high or low activity and confirming that it may be applied for
identification of residues playing a role in catalysis.

<>

<1>Ulge, U.Y., Baker, D.A., Monnat, R.J. Jr.
<2>Comprehensive computational design of mCreI homing endonuclease cleavage specificity for genome engineering.
<3>Nucleic Acids Res.
<4>39
<5>4330-4339
<6>2011
<7>Homing endonucleases (HEs) cleave long ( approximately 20 bp) DNA target sites with high site
specificity to catalyze the lateral transfer of
parasitic DNA elements. In order to determine whether comprehensive
computational design could be used as a general strategy to engineer new
HE target site specificities, we used RosettaDesign (RD) to generate 3200
different variants of the mCreI LAGLIDADG HE towards 16 different base
pair positions in the 22 bp mCreI target site. Experimental verification
of a range of these designs demonstrated that over 2/3 (24 of 35 designs,
69%) had the intended new site specificity, and that 14 of the 15
attempted specificity shifts (93%) were achieved. These results
demonstrate the feasibility of using structure-based computational design
to engineer HE variants with novel target site specificities to facilitate
genome engineering.

<>

<1>Ulyanova, V., Shah, M.R., Dudkina, E., Vershinina, V., Ilinskaya, O.
<2>Draft Whole Genome Sequence of Bacillus pumilus Strain 3-19, a Chemical Mutant Overproducing Extracellular Ribonuclease.
<3>Genome Announcements
<4>2
<5>e00724-14
<6>2014
<7>Here, we present a draft genome sequence of Bacillus pumilus strain 3-19. It was  derived from
soil-isolated B. pumilus 7P using chemical mutagenesis and is
characterized by elevated production of extracellular ribonuclease which is known
to possess different biological activities with potential of applications in
experimental research, medicine, and biotechnology.

<>

<1>Um, J., Park, C., Lee, K.
<2>Alteration of specificity of restriction endonuclease EcoRI in organic solvent.
<3>Korean Biochem. J.
<4>27
<5>401-407
<6>1994
<7>In molecular biology, type-II restriction endonucleases, which specifically cleave DNA at a
limited number of sites, have been exploited as a means of characterizing DNA fragments, DNA
mapping and of modifying DNA for genetic engineering. Recently, many type-II restriction
endonucleases have been found to decrease their substrate specificity under modified
conditions such as extreme pH, low ionic strength, high enzyme concentration, substitution of
metallic cofactors, or addition of organic solvents. This study used restriction endonuclease
EcoRI which is used most frequently in genetic engineering. We investigated their specificity
change in buffer conditions including various organic solvents. The specificity of cleavage of
EcoRI is altered in the presence of hydrophobic reagents, such as ethanol, ethyleneglycol and
DMSO. The enzyme recognition site was not changed randomly but by preferential order by
increasing the concentration of organic solvent. When EcoRI reacted with substrate pGEM3
vector which has one canonical recognition site (GAATTC), EcoRI cleaved the noncanonical
TAATTC and GAGTTC subsequently in more than 10% DMSO solution. These changes of specificities
depended on the hydrophobicity of organic solvent (LogP: partition coefficient). As a result,
the recognition sequence site was changed in the presence of organic solvents whose LogP are
-2.0 to 0. The specificities were not easily changed in enzyme inactivating organic solvent
such as acetone, 2-methyl-2-propanol, 2-methyl-1-propanol. These results show that restriction
enzyme could be used to cleave at unusual site by changing the reaction conditions.

<>

<1>Underhill, P.A., Johnson, P.H.
<2>Site-specific mutagenesis of the EcoRI restriction endonuclease: substitution of cysteine-217 with glycine affects in vivo but not in vitro enzyme function.
<3>Fed. Proc.
<4>45
<5>1875
<6>1986
<7>Using oligonucleotide-directed mutagenesis, we have introduced a T - G
transversion mutation in the cloned EcoRI restriction endonuclease gene,
thereby substituting a glycine for the single cysteine-217.  Mutant clones were
identified by differential colony hybridization using the mutagenesis
oligonucleotide.  The structure of the mutant was confirmed by DNA sequence
analysis.  Cells producing the mutant endonuclease showed a four-log increase
in sensitivity to infection by bacteriophage lambda.  The mutant enzyme was
partially purified and characterized.  It did not show any changes in
recognition site-specificity or in kinetic behavior compared with that of the
wild-type enzyme either in standard assay buffer or under altered solvent
conditions that promoted its ability to cleave subcanonical sequences.  The
mutant and wild-type enzymes behaved identically during analytical gel
chromatography.  Reaction of the wild-type endonuclease with the sulfhydryl
inhibitor parahydroxymercuribenzoate resulted in a complete loss of enzyme
activity compared with less than tenfold inhibition for the mutant enzyme.We
conclude that cysteine-217 may play an important role in the in vivo function
of the EcoRI endonuclease but not in its in vitro enzymatic activity.

<>

<1>Unzaga, T.V., Diaz-Ricci, J.C., Rhee, J.I., Hernandez, M.R., Schugerl, K.
<2>Modeling of the controlled expression of a harmful protein by a three-plasmid harboring system.
<3>Biotechnol. Bioeng.
<4>80
<5>544-551
<6>2002
<7>A genetically structured mathematical model was developed and used to evaluate the influence
of molecular parameters involved in the expression
of a harmful recombinant protein (SPA::EcoRI). The system consists of the
controlled expression of the endonuclease EcoRI cloned in the plasmid
pMTC48. The control is exerted by the lambda Cl repressor expressed from
the plasmid pRK248clts. The deleterious effect of the activity of the
enzyme EcoRl on the host DNA is prevented by the action of the EcoRl
methylase that is expressed constitutively from a third plasmid, pEcoR4.
The model includes molecular mechanisms involved in the regulation of the
expression of these genes and is used to determine cultural conditions
that maximize the production of the recombinant protein.

<>

<1>Uozumi, T., Hoshino, T., Miwa, K., Horinouchi, S., Beppu, T., Arima, K.
<2>Restriction and modification in Bacillus species.
<3>Mol. Gen. Genet.
<4>152
<5>65-69
<6>1977
<7>Host specific restriction was detected in 13 Bacillus strains, when 63 strains of Bacillus
subtilis and 15 other Bacillus strains were tested with phage phi 105C. These 13 strains were
classified into 8 groups (M,H,C,N,E,F,G,P) by the type of restriction. M-type strains (B.
subtilis Marburg 168, its derivatives, and two other strains) showed relatively weak
restriction, restricting phi 105C from other groups of Bacillus by ratios of 10(-1) to 10(-3).
Strains of groups H,C,N,E,F,G, and P restricted phi 105C from other groups by ratios of 10(-2)
to 10(-8). It was confirmed with some of the strains that type-specific modification was
endowed only by the last host. Furthermore, we isolated one restriction deficient mutant of B.
subtilis Marburg 168-YS11, which had also lost its modification phenotype.

<>

<1>Uporova, T.M., Kartasheva, I.M., Skripkin, E.A., Lopareva, E.N., Nikolskaya, I.I., Debov, S.S.
<2>Restricting endonucleases from Shigella sonnei 47.
<3>Vopr. Med. Khim.
<4>31
<5>131-136
<6>1985
<7>Two restrictases SsoI and SsoII, belonging to the enzymes of restriction of the class II, were
isolated from a strain of dysenteric bacteria.  The structure of the site sensitive to SsoI
and SsoII was studied after fragmentation of testor DNA as well as by means of direct
determination of nucleotide sequency.  SsoI was shown to be an isoschizomer of the EcoRI
restrictase from E. coli. Restrictase SsoII proved to a new enzyme, which hydrolyzed the
sequence 5'...CCNGG..3' and was distinct from the known restrictases as shown by studies of
the type of DNA hydrolyzed.  A three-step procedure is developed for isolation of SsoII
restrictase involving the consecutive chromatography on blue sepharose, phosphocellulose P11
and phenyl-sepharose.  Restrictases SsoI and EcoRI were isolated by means of
isoelectrofocusing using ampholines.

<>

<1>Uporova, T.M., Nikolskaya, I.I., Rubtsova, E.N., Debov, S.S.
<2>Purification and identification of Eco CK restrictional endonuclease.
<3>Vestn. Akad. Med. Nauk SSSR
<4>2
<5>21-26
<6>1981
<7>A fundamentally new scheme for purifying the restrictional endonuclease from E.
coli CK cells has been developed, involving 3 consecutive stages:  gel
filtration (on biogel A-0.5 m), chromatography on single-stranded
DNA-cellulose, and fractionation on heparin-sepharose.  The proposed variant of
REco CK purification results in an enzyme preparation completely free from
nonspecific endonucleases and exonucleases which can be used in genetic
engineering and physical mapping work.  The fragmentation pattern of phage
lambda DNA has been defined more precisely:  as a result of complete
hydrolysis, 4 fragments with molecular masses of 13.25, 12.0, 5.15, and 0.7
megadaltons are formed.  It is shown that in as far as the fragmentation
pattern of the test phage lambda DNA is concerned, the REco CK enzyme has no
analogues among the enzymes described so far in the literature.

<>

<1>Urbanczyk, H., Ogura, Y., Hendry, T.A., Gould, A.L., Kiwaki, N., Atkinson, J.T., Hayashi, T., Dunlap, P.V.
<2>Genome Sequence of Photobacterium mandapamensis svers. 1.1., Bioluminescent Symbiont of the Cardinalfish Siphamia versicolor.
<3>J. Bacteriol.
<4>193
<5>3144-3145
<6>2011
<7>Photobacterium mandapamensis is one of three luminous Photobacterium species able to form
species-specific bioluminescent symbioses with marine
fishes. Here, we present the draft genome sequence of P. mandapamensis
svers.1.1, bioluminescent symbiont of the cardinalfish, Siphamia
versicolor, the first genome of a symbiotic, luminous Photobacterium
species to be sequenced. Analysis of the sequence provides insight into
differences between P. mandapamensis and other luminous and symbiotic
bacteria in genes involved in quorum sensing regulation of light
production and establishment of symbiosis.

<>

<1>Urbieta, M.S., Rascovan, N., Castro, C., Revale, S., Giaveno, M.A., Vazquez, M., Donati, E.R.
<2>Draft Genome Sequence of the Novel Thermoacidophilic Archaeon Acidianus copahuensis Strain ALE1, Isolated from the Copahue Volcanic Area in Neuquen,  Argentina.
<3>Genome Announcements
<4>2
<5>e00259-14
<6>2014
<7>Acidianus copahuensis is a recently characterized thermoacidophilic archaeon isolated from the
Copahue volcanic area in Argentina. Here, we present its draft
genome sequence, in which we found genes involved in key metabolic pathways for
developing under Copahue's extreme environmental conditions, such as sulfur and
iron oxidation, carbon fixation, and metal tolerance.

<>

<1>Urieli-Shoval, S., Gruenbaum, Y., Razin, A.
<2>Sequence and substrate specificity of isolated DNA methylases from Escherichia coli C.
<3>J. Bacteriol.
<4>153
<5>274-280
<6>1983
<7>Two DNA methylase activities of Escherichia coli C, the mec (designates DNA-cytosine-methylase
gene, which is also designated dcm) and dam gene products, were physically separated by
DEAE-cellulose column chromatography. The sequence and substrate specificity of the two
enzymes were studied in vitro. The experiments revealed that both enzymes show their expected
sequence specificity under in vitro conditions, methylating symmetrically on both DNA strands.
The mec enzyme methylates exclusively the internal cytosine residue of CC(A/T)GG sequences,
and the dam enzyme methylates adenine residues at GATC sites. Substrate specificity
experiments revealed that both enzyes methylate in vitro unmethylated duplex DNA as
efficiently as hemimethylated DNA. The results of these experiments suggest that the
methylation at a specific site takes place by two independent events. A methyl group in a site
on one strand of the DNA does not facilitate the methylation of the same site on the opposite
strand. With the dam methylase it was found that enzyme is incapable of methylating GATC sites
located at the ends of DNA molecules.

<>

<1>Urig, S., Gowher, H., Hermann, A., Beck, C., Fatemi, M., Humeny, A., Jeltsch, A.
<2>The Escherichia coli Dam DNA methyltransferase modifies DNA in a highly processive reaction.
<3>J. Mol. Biol.
<4>319
<5>1085-1096
<6>2002
<7>The Escherichia coli dam adenine-N6 methyltransferase modifies DNA at G A TC sequences. It is
involved in post-replicative mismatch repair, control of DNA replication and gene regulation.
We show that E. coli dam acts as a functional monomer and methylates only one strand of the
DNA in each binding event. The preferred way of ternary complex assembly is that the enzyme
first binds to DNA and then to S-adenosylmethionine. The enzyme methylates an oligonucleotide
containing two dam sites and an 879 bp PCR product with four sites in a fully processive
reaction. On -DNA comprising 48,502 bp and 116 dam sites, E. coli dam scans 3000 dam sites per
binding event in a random walk, that on average leads to a processive methylation of 55 sites.
Processive methylation of DNA considerably accelerates DNA methylation. The highly processive
mechanism of E. coli dam could explain why small amounts of E. coli dam are able to maintain
the methylation state of dam sites during DNA replication. Furthermore, our data support the
general rule that solitary DNA methyltransferase modify DNA processively whereas
methyltransferases belonging to a restriction-modification system show a distributive
mechanism, because processive methylation of DNA would interfere with the biological function
of restriction-modification systems.

<>

<1>Uritani, M., Takai, M., Yoshinaga, K.
<2>Protective effect of disaccharides on restriction endonucleases during drying under vacuum.
<3>J. Biochem. (Tokyo)
<4>117
<5>774-779
<6>1995
<7>Desiccation by vacuum-drying inactivates the restriction endonuclease HindIII completely.
However, when dried in the presence of a disaccharide such as trehalose, maltose, or sucrose,
the endonuclease retains its phage DNA-cleaving activity and produces the same digestive
fragments as does the intact enzyme. Thus, the disaccharides are effective in protecting the
restriction enzyme in terms of both recognition and accurate cleavage of the substrate. Among
the disaccharides, trehalose protects the enzyme most effectively; and it also stabilizes the
enzyme during dilution in aqueous solution. The restriction enzyme dried with trehalose
maintains its activity without detectable loss for at least 4 days at 37oC, but it shows
reduced activity after 30-day storage at either 4oC or room temperature. Trehalose also
protects other restriction endonucleases, EcoRI and BamHI, from inactivation during
vacuum-drying, whereas drying them alone leads to severe loss of their activity. The
restriction endonucleases dried with trehalose retain their activities for at least 20 days at
4oC and for 7 days at room temperature.

<>

<1>Uroz, S., Tech, J.J., Sawaya, N.A., Frey-Klett, P., Leveau, J.H.J.
<2>Structure and function of bacterial communities in ageing soils: insights from the Mendocino ecological staircase.
<3>Soil Biol. Biochem.
<4>69
<5>265-274
<6>2014
<7>
<>

<1>Urushibara, N., Kawaguchiya, M., Kobayashi, N.
<2>Two novel arginine catabolic mobile elements and staphylococcal chromosome cassette mec composite islands in community-acquired methicillin-resistant Staphylococcus aureus genotypes ST5-MRSA-V and ST5-MRSA-II.
<3>J. Antimicrob. Chemother.
<4>67
<5>1828-1834
<6>2012
<7>Objectives: The arginine catabolic mobile element (ACME) is a novel staphylococcal genetic
island. ACME is located downstream of the staphylococcal cassette chromosome mec (SCCmec),
forming the ACME-SCCmec composite island. Recently, ACME II (located upstream of SCCmec IV)
was described from a methicillin-resistant Staphylococcus aureus (MRSA) strain M1 in Denmark
(ST8-MRSA-IVa) and 15 MRSA isolates in Ireland (ST22-
MRSA-IVh). We report the novel genetic characteristics of the ACME-SCCmec composite islands
found in Japanese community-acquired MRSA (CA-MRSA) isolates.  Methods: ACME-SCCmec composite
islands from two ACME-arcA-positive CA-MRSA isolates with the genotypes ST5-MRSA-V (SR141) and
ST5-MRSA-II (SR388) were characterized using long-range PCR and nucleotide sequencing.
Results: Both isolates harboured a 12 kb DNA region primarily identified in ACME II in
Staphylococcus epidermidis ATCC 12228 upstream of each SCCmec. The arcA and its flanking
regions in SR141 and SR388 showed high sequence
identity (99.8% at the highest) to those in MRSA M1 and M08/0126 (the representative of 15
Irish ST22-MRSA-IVh isolates), suggesting that the ACMEs of these four isolates originated
from the same ancestral gene.
The ACME II-like element in SR141 included an insertion sequence IS1182 at a position close to
SCCmec, resulting in a new variant. SR388 contained 11.5 kb of the J1 region of type I SCCmec
(J1 SCCmecI) between orfX and ACME (orfX-J1 SCCmecI-ACME II), unlike the homologous region in
M08/0126 (orfX-ACME II-J1 SCCmecI).  Conclusions: This is the first report of the ACME II-like
element inserted upstream of SCCmec in CA-MRSA with the genotypes ST5-MRSA-V and ST5-MRSA-II.

<>

<1>Usami, S., Kurimura, H., Kino, K., Kamigaki, K., Kirimura, K.
<2>Mutated restriction enzyme.
<3>Japanese Patent Office
<4>JP 2003259876 A
<5>10
<6>2003
<7>
<>

<1>Ushakova, T.A., Puchkova, L.I., Gutorov, V.V., Kuvshinov, V.N., Repin, V.E.
<2>Restriction endonuclease Sst12I from a strain of Streptomyces - Recognizing the nucleotide sequence 5 '-CTGCAG-3 '.
<3>Prikl. Biokhim. Mikrobiol.
<4>38
<5>25-28
<6>2002
<7>A new restriction endonuclease Sst12I belonging to type II and recognizing the sequence
5'-CTGCAG-3' was isolated from the bacterial
strain Streptomyces sp. St-12. The enzyme hydrolyzes DNA between
adenine and guanine residues thus, it is a true isoschizomer of
restrictase PstI. In contrast to PstI, the restriction endonuclease
Sst12I hydrolyses DNA both at 37 and 55 C and remains active
after long-term storage.

<>

<1>Ushakova, T.A., Puchkova, L.I., Gutorov, V.V., Totmenina, O.D., Repin, V.E.
<2>Restriction endonuclease Asi256I recognizes and cuts the nucleotide sequence 5'-GATC-3'.
<3>Prikl. Biokhim. Mikrobiol.
<4>44
<5>28-31
<6>2008
<7>A strain producing a restriction endonuclease was isolated from soil samples and identified as
the Arthrobacter sp. strain Ck256. The enzyme
produced by this strain was termed Asi256I. The isolation procedure for
this enzyme was described, and the optimal conditions for its function
were determined. It was shown that the restriction endonuclease Asi256I
is a true isoschizomer of MboI, it has a temperature optimum of 6
degrees C, and can be used in molecular-biological and
genetic-engineering studies performed at low temperatures.

<>

<1>Ushakova, T.A., Puchkova, L.I., Polushkina, A.F., Kuvshinov, V.N., Repin, V.E.
<2>Isolation and chromatographic purification of restriction endonuclease Sst12I from Streptomyces.
<3>Russian Patent Office
<4>RU 2233877 C
<5>
<6>2004
<7>Invention relates to isolation and purification of site-specific endonuclease restriction
synthesized by microorganisms of genus Streptomyces, namely, to preparing site-specific
endonuclease restriction Sst12I that recognizes and cleaves the nucleotide sequence:
5'-CTGCAG-3'. Method involves disruption of cells by ultrasonic oscillation, fractional
thermal treatment (3 times for 3 min) at temperature 60 C, not above, centrifugation,
chromatography on phosphocellulose P-11 column and concentrating against 50% glycerol
solution. The yield of enzyme is 500000 U/10 g of biomass. This method provides preparing
enzyme without impurities of nonspecific nucleases and phosphatases.

<>

<1>Ushakova, T.A., Puchkova, L.I., Radionenko, Y.V., Gutorov, V.V., Kuvshinov, V.N., Repin, V.E.
<2>Restriction endonuclease SpmI recognizing the 5'-AT/CGAT-3' nucleotide sequence.
<3>Biotekhnologiya
<4>0
<5>38-42
<6>2004
<7>A strain producer of a restriction endonuclease (restrictase), that was identified as a
species of Sphingobacterium mizutae has been isolated from snow water, and its restrictase was
named SpmI.  The method of the enzymne isolation was described, and optimal conditions of its
action were determined.  It was shown that SpmI restrictase is a true isoschizomer of ClaI
restrictase.  The temperature optimum of SpmI restrictase is 6oC, and it can be used in
molecular biology and genetic engineering researches that need carrying out at low
temperatures.

<>

<1>Ushakova, T.A., Puchkova, L.I., Radionenko, Y.V., Kuvshinov, V.N., Repin, V.E.
<2>Restriction endonuclease Bst221 from Bacillus stearothermophilus 22 strain recognizing the 5'-CCNNNNN^NNGG-3' nucleotide sequence.
<3>Biotekhnologiya
<4>0
<5>11-15
<6>2003
<7>A strain producing the restriction endonuclease (restrictase) which was identified as a
species of Bacillus stearothermophilus 22 has been isolated by the authors, and its
restrictase was called Bst221.  The method of the enzyme isolation was described, and optimal
conditions of its action were determined.  It was shown that Bst221 restrictase is a true
isoschizomer of BsiYI restrictase.  Bst221 restrictase is active in a broad temperature range
and shows promise for molecular biology research.

<>

<1>Ushay, H.M., Tullius, T.D., Lippard, S.J.
<2>Inhibition of the BamHI cleavage and unwinding of pBR322 deoxyribonucleic acid by the antitumor drug cis-dichlorodiammineplatinum(II).
<3>Biochemistry
<4>20
<5>3744-3748
<6>1981
<7>The antitumor drug cis-dichlorodiammineplatinum(II) (cis-DDP) binds to pBR322 DNA and inhibits
the cleavage of this circular DNA into a linear form by the restriction endonuclease BamHI.
The binding of platinum to DNA was monitored by agarose gel electrophoresis, and the amount of
platinum bound per nucleotide (rb) was measured by carbon rod atomic absorption spectroscopy.
Electrophoretic mobility changes reflect a shortening and unwinding of the DNA duplex upon
platinum binding as observed previously for the reaction of cis- and trans-DDP with pSM2 DNA
[Cohen, G.L., Bauer, W.R., Barton, J.K., & Lippard, S.J. (1979) Science (Washington, DC) 203,
1014-1016].  The inhibition of BamHI nuclease activity occurs at very low binding levels and
is complete at rb = 0.045.  This value corresponds to the binding of one platinum atom within
+/- 3 base pairs of the recognition sequence of the enzyme shown below.  Treatment of the DNA
with 0.2 M sodium cyanide after BamHI cutting removes the platinum but does not alter the
point at which cis-DDP inhibits the formation of the linear form III DNA.  This result is in
contrast with a previous report claiming that BamHI could cut across a cis-DDP-induced GpG
cross-link in DNA which could be subsequently revealed by cyanide reversal of platinum
binding.  When the platinum is removed by cyanide treatment, the drug-induced mobility changes
are reversed and there is a pronounced sharpening of the bands in the gel. Quantitative study
of the cyanide reversal shows the presence of a small amount of unremovable platinum tightly
bound to the DNA at high ratios (~0.1) of bound platinum per nucleotide -G^GATCC- -CCTAG^G-.

<>

<1>Ushijima, B., Videau, P., Aeby, G.S., Callahan, S.M.
<2>Draft Genome Sequence of Vibrio coralliilyticus Strain OCN008, Isolated from Kane'ohe Bay, Hawai'i.
<3>Genome Announcements
<4>1
<5>e00786-13
<6>2013
<7>Vibrio coralliilyticus is a Gram-negative bacterium found in seawater and is associated with
diseased marine organisms. Strains of V. coralliilyticus have
been shown to infect coral from multiple genera. We report the draft genome
sequence of V. coralliilyticus strain OCN008, the third V. coralliilyticus genome
to be sequenced.

<>

<1>Ushijima, B., Videau, P., Poscablo, D., Vine, V., Salcedo, M., Aeby, G., Callahan, S.M.
<2>Complete Genome Sequence of Vibrio coralliilyticus Strain OCN014, Isolated from a Diseased Coral at Palmyra Atoll.
<3>Genome Announcements
<4>2
<5>e01318-14
<6>2014
<7>Vibrio coralliilyticus is a marine gammaproteobacterium that has been implicated  as an
etiological agent of disease for multiple coral genera on reefs worldwide.
We report the complete genome of V. coralliilyticus strain OCN014, isolated from
a diseased Acropora cytherea colony off the western reef terrace of Palmyra
Atoll.

<>

<1>Ushijima, K., Ishibashi, T., Tsukahara, S., Takai, K., Takaku, H.
<2>Inhibition of restriction endonuclease cleavage by triplex formation with oligo-2'-O-methyl-ribonucleotides containing 8-oxo-2'-O-methyladenosine in place of cytidine.
<3>Heterocycles
<4>52
<5>389-398
<6>2000
<7>The ability of homopyrimidine oligoribonucleotides and oligo-2'-O-methyl-ribonucleotides
containing 8-oxo-adenosine and 8-oxo-2'-O-methyl-adenosine to form stable, triple-helical
structures with sequences containing the recognition site for the class II-S restriction
enzyme, Ksp632I, was studied as a function of pH.  The AOH- and AmOH-substituted
oligoribonucleotides and oligo-2'-O-methyl-ribonucleotides were shown to bind within the
physiological pH range in a pH-independent fashion, without a compromise in specificity.  The
substitutions of three cytidine residues with AOH showed higher endonuclease inhibition than
the substitution of either one or two cytidine residues with AOH.  In particular, the
oligo-2'-O-methyl-ribonucleotide with only one cytidine substituted with AmOH showed higher
endonuclease inhibition.  Increased resistance to nucleases is observed with the introduction
of 2-O-methylnucleosides.  This stabilization should help us to design much more efficient
third strand homopyrimidine oligomer and antisense nucleic acid, which can be used as tools in
cellular biology and anti-viral therapy.

<>

<1>Ushijima, K., Ishibashi, T., Yamakawa, H., Tsukahara, S., Takai, K., Maruyama, T., Takaku, H.
<2>Inhibition of restriction endonuclease cleavage by triple helix formation with RNA and 2'-O-methyl RNA oligonucleotides containing 8-oxo-adenosine in place of cytidine.
<3>Biochemistry
<4>38
<5>6570-6575
<6>1999
<7>The ability of homopyrimidine oligoribonucleotides (RNA) and
oligo-2'-O-methyl-ribonucleotides (2'-O-methyl RNA) containing 8-oxo-adenosine (AOH) and
8-oxo-2'-O-methyl (AmOH) adenosine to form stable, triple-helical structures with sequences
containing the recognition site for the class II-S restriction enzyme, Ksp632-I, was studied
as a function of pH. The AOH- and AmOH-substituted RNA and 2'-O-methyl RNA oligonucleotides
were shown to bind within the physiological pH range in a pH-independent fashion, without a
compromise in specificity. The substitutions of three cytidine residues with AOH showed higher
endonuclease inhibition than the substitution of either one or two cytidine residues with AOH.
In particular, the 2'-O-methyl RNA oligonucleotide with only one cytidine substituted with
AmOH showed higher endonuclease inhibition than the homopyrimidine RNA and 2'-O-methyl RNA
oligonucleotides and the RNA oligonucleotides containing either one or two AOH moieties.
Furthermore, the AmOH-substituted 2'-O-methyl RNA oligonucleotides were stable (53%) after an
incubation in 10% fetal bovine serum for 8 h, whereas the RNA oligonucleotides were completely
degraded. Increased resistance to nucleases is observed with the introduction of
2'-O-methylnucleosides. This stabilization should help us to design much more efficient third
strand homopyrimidine oligomer and antisense nucleic acid-based antiviral therapies, which
could be used as tools in cellular biology.

<>

<1>Usuda, Y., Nishio, Y., Matsui, K., Sugimoto, S., Koseki, K.
<2>Polynucleotides encoding useful polypeptides in Corynebacterium glutamicum ssp. lactofermentum.
<3>US Patent Office
<4>US 7468262 B
<5>
<6>2008
<7>The present invention provides novel polypeptides and polynucleotides useful as
biotechnological tools, specifically identified in a coryneform bacterium Corynebacterium
glutamicum ssp. lactofermentum and methods of producing substances in organisms having
enhanced or attenuated expression of these polypeptides and/or polynucleotides.

<>

<1>Utter, B., Deutsch, D.R., Schuch, R., Winer, B.Y., Verratti, K., Bishop-Lilly, K., Sozhamannan, S., Fischetti, V.A.
<2>Beyond the Chromosome: The Prevalence of Unique Extra-Chromosomal Bacteriophages with Integrated Virulence Genes in Pathogenic Staphylococcus aureus.
<3>PLoS ONE
<4>9
<5>E100502
<6>2014
<7>In Staphylococcus aureus, the disease impact of chromosomally integrated
prophages on virulence is well described. However, the existence of
extra-chromosomal prophages, both plasmidial and episomal, remains obscure.
Despite the recent explosion in bacterial and bacteriophage genomic sequencing,
studies have failed to specifically focus on extra-chromosomal elements. We
selectively enriched and sequenced extra-chromosomal DNA from S. aureus isolates
using Roche-454 technology and uncovered evidence for the widespread distribution
of multiple extra-chromosomal prophages (ExPPhis) throughout both
antibiotic-sensitive and -resistant strains. We completely sequenced one such
element comprised of a 43.8 kbp, circular ExPPhi (designated capital EF,
CyrillicBU01) from a vancomycin-intermediate S. aureus (VISA) strain. Assembly
and annotation of capital EF, CyrillicBU01 revealed a number of putative
virulence determinants encoded within a bacteriophage immune evasion cluster
(IEC). Our identification of several potential ExPPhis and mobile genetic
elements (MGEs) also revealed numerous putative virulence factors and antibiotic
resistance genes. We describe here a previously unidentified level of genetic
diversity of stealth extra-chromosomal elements in S. aureus, including phages
with a larger presence outside the chromosome that likely play a prominent role
in pathogenesis and strain diversity driven by horizontal gene transfer (HGT).

<>

<1>Utturkar, S.M., Bollmann, A., Brzoska, R.M., Klingeman, D.M., Epstein, S.E., Palumbo, A.V., Brown, S.D.
<2>Draft Genome Sequence for Caulobacter sp. Strain OR37, a Bacterium Tolerant to Heavy Metals.
<3>Genome Announcements
<4>1
<5>e00322-13
<6>2013
<7>Caulobacter sp. strain OR37 belongs to the class Alphaproteobacteria and was isolated from
subsurface sediments in Oak Ridge, TN. Strain OR37 is noteworthy
due to its tolerance to high concentrations of heavy metals, such as uranium,
nickel, cobalt, and cadmium, and we present its draft genome sequence here.

<>

<1>Utturkar, S.M., Bollmann, A., Brzoska, R.M., Klingeman, D.M., Epstein, S.E., Palumbo, A.V., Brown, S.D.
<2>Draft Genome Sequence for Ralstonia sp. Strain OR214, a Bacterium with Potential  for Bioremediation.
<3>Genome Announcements
<4>1
<5>e00321-13
<6>2013
<7>Ralstonia sp. strain OR214 belongs to the class Betaproteobacteria and was isolated from
subsurface sediments in Oak Ridge, TN. A member of this genus has
been described as a potential bioremediation agent. Strain OR214 is tolerant to
various heavy metals, such as uranium, nickel, cobalt, and cadmium. We present
its draft genome sequence here.

<>

<1>Uyar, A., Kurkcuoglu, O., Nilsson, L., Doruker, P.
<2>The elastic network model reveals a consistent picture on intrinsic functional dynamics of type II restriction endonucleases.
<3>Phys. Biol.
<4>8
<5>056001
<6>2011
<7>The vibrational dynamics of various type II restriction endonucleases, in complex with
cognate/non-cognate DNA and in the apo form, are
investigated with the elastic network model in order to reveal common
functional mechanisms in this enzyme family. Scissor-like and tong-like
motions observed in the slowest modes of all enzymes and their
complexes point to common DNA recognition and cleavage mechanisms.
Normal mode analysis further points out that the scissor-like motion
has an important role in differentiating between cognate and
non-cognate sequences at the recognition site, thus implying its
catalytic relevance. Flexible regions observed around the DNA-binding
site of the enzyme usually concentrate on the highly conserved
beta-strands, especially after DNA binding. These beta-strands may have
a structurally stabilizing role in functional dynamics for target site
recognition and cleavage. In addition, hot spot residues based on
high-frequency modes reveal possible communication pathways between the
two distant cleavage sites in the enzyme family. Some of these hot
spots also exist on the shortest path between the catalytic sites and
are highly conserved.

<>

<1>Uyen, N.T., Nishi, K., Park, S.Y., Choi, J.W., Lee, H.J., Kim, J.S.
<2>Crystallization and preliminary X-ray diffraction analysis of the HsdR subunit of a putative type I restriction enzyme from Vibrio vulnificus YJ016.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>64
<5>926-928
<6>2008
<7>Type I restriction enzymes are multimeric proteins that consist of three subunits. The HsdS
and HsdM subunits form a complex protein that
shows methyltransferase activity, while the HsdR subunit functions as
an endonuclease as well as as a translocase. Of these three subunits,
no structural information on the HsdR subunit is yet available. The
putative HsdR gene from Vibrio vulnificus YJ016 (HsdR Vv) was cloned
and expressed and the expressed protein HsdR Vv was purified. HsdR Vv
was crystallized from 8%(w/v) polyethylene glycol 3350, 0.15 M ammonium
chloride, 0.1 M HEPES pH 7.5 and 2 mM beta-mercaptoethanol. Diffraction
data were collected to 2.60 angstrom resolution using synchrotron
radiation. The crystal belongs to the orthorhombic space group
P2(1)2(1)2(1), with unit-cell parameters a = 71.01, b = 89.04, c =
113.66 angstrom. With one HsdR Vv molecule in the asymmetric unit, the
Matthews coefficient was 2.14 angstrom(3) Da(-1) and the solvent
content was 42%.

<>

<1>Uyen, N.T., Park, S.Y., Choi, J.W., Lee, H.J., Nishi, K., Kim, J.S.
<2>The fragment structure of a putative HsdR subunit of a type I restriction enzyme from Vibrio vulnificus YJ016: implications for DNA restriction and  translocation activity.
<3>Nucleic Acids Res.
<4>37
<5>6960-6969
<6>2009
<7>Among four types of bacterial restriction enzymes that cleave a foreign DNA depending on its
methylation status, type I enzymes composed of three subunits are interesting because of their
unique DNA cleavage and translocation mechanisms performed by the restriction subunit (HsdR).
The elucidated N-terminal fragment structure of a putative HsdR subunit from Vibrio vulnificus
YJ016 reveals three globular domains. The nucleolytic core within an N-terminal nuclease
domain (NTD) is composed of one basic and three acidic residues, which include a metal-binding
site. An ATP hydrolase (ATPase) site at the interface of two RecA-like domains (RDs) is
located close to the probable DNA-binding site for translocation, which is far from the NTD
nucleolytic core. Comparison of relative domain arrangements with other functionally related
ATP and/or DNA complex structures suggests a possible translocation and restriction mechanism
of the HsdR subunit. Furthermore, careful analysis of its sequence and structure implies that
a linker helix connecting two RDs and an extended region within the nuclease domain may play a
central role in switching the DNA translocation into the restriction activity.

<>

<1>Uzelac, G., Bertani, I., Kojic, M., Paszkiewicz, K.H., Studholme, D.J., Passos-da-Silva, D., Venturi, V.
<2>Draft Genome Sequence of Beneficial Rice Rhizosphere Isolate Pseudomonas aeruginosa PUPa3.
<3>Genome Announcements
<4>2
<5>e00654-14
<6>2014
<7>Pseudomonas aeruginosa PUPa3 is a rhizosphere-colonizing and plant growth-promoting strain
isolated from the rhizosphere of rice. This strain has, however, been shown to be pathogenic
in two nonmammalian infection models. Here we report the draft genome sequence of P.
aeruginosa PUPa3.

<>

<1>Vader, A., Nielsen, H., Johansen, S.
<2>In vivo expression of the nucleolar group I intron-encoded I-DirI homing endonuclease involves the removal of a spliceosomal intron.
<3>EMBO J.
<4>18
<5>1003-1013
<6>1999
<7>The Didymium iridis DiSSU1 intron is located in the nuclear SSU rDNA and has an unusual
twin-ribozyme organization.  One of the ribozymes (DiGIR2) catalyses intron excision and exon
ligation.  The other ribozyme (DiGIR1), which along with the endonuclease-encoding I-DirI open
reading frame is inserted in DiGIR2, carries out hydrolysis at internal processing sites (IPS1
and IPS2) located at its 3' end.  Examination of the in vivo expression of DiSSU1 shows that
after excision, DiSSU1 is matured further into the I-DirI mRNA by internal DiGIR1-catalysed
cleavage upstream of the ORF 5' end, as well as truncation and polyadenylation downstream of
the ORF 3' end.  A spliceosomal intron, the first to be reported within a group I intron and
the rDNA, is removed before the I-DirI mRNA associates with the polysomes.  Taken together,
our results imply that DiSSU1 uses a unique combination of intron-supplied ribozyme activity
and adaptation to the general RNA polymerase II pathway of mRNA expression to allow a protein
to be produced from the RNA polymerase I-transcribed rDNA.

<>

<1>Vaid, R.K., Shanmugasundaram, K., Boora, A., Bera, B.C., Shukla, B.N., Anand, T., Singha, H., Riyesh, T., Virmani, N., Barua, S., Ahir, V.B., Koringa, P.G., Sajnani, M.R., Bhat, V.D., Rana, N., Singh, K.P., Malik, P., Singh, R.K., Joshi, C.G.
<2>Draft Genome Sequence of Pasteurella multocida subsp. multocida B:2 Strain VTCCBAA264 Isolated from Bubalus bubalis in North India.
<3>Genome Announcements
<4>2
<5>e00755-14
<6>2014
<7>The Pasteurella multocida subsp. multocida B:2 serotype causes hemorrhagic septicemia in
bubalines with high morbidity and mortality in the Indian
subcontinent. We report the draft genome sequence of Pasteurella multocida strain
VTCCBAA264 isolated from the small-intestine of a buffalo calf that died of high
fever.

<>

<1>Vainstein, A., Zuker, A.
<2>Plant viral expression vectors and use of same for generating genotypic variations in plant genomes.
<3>International Patent Office
<4>WO 2009130695 A
<5>
<6>2009
<7>A method of generating genotypic variation in a genome of a plant is disclosed.  The method
comprising introducing into the plant at least one viral expression vector encoding at least
one chimeric nuclease which comprises a DNA binding domain, a nuclease and a localization
signal to a DNA-containing organelle, wherein the DNA binding domain mediates specific
targeting of the nuclease to the genome of the plant, thereby generating genotypic variation
in the genome of the plant.

<>

<1>Vainstein, A., Zuker, A.
<2>Generating genotypic variations in plant genomes by gamete infection.
<3>International Patent Office
<4>WO 2011048600 A
<5>
<6>2011
<7>A method of generating genotypic variation in a genome of a plant is disclosed.  The method
comprising introducing into a gamete or a gamete producing tissue of the plant at least one
viral expression vector encoding at least one chimeric nuclease which comprises a DNA binding
domain, a nuclease and a localization signal to a DNA-containing organelle, wherein the DNA
binding domain mediates specific targeting of the nuclease to the genome of the plant, wherein
the introducing is performed such that the gamete or gamete producing tissue expresses the
chimeric nuclease but not all plant tissues express the chimeric nuclease, thereby generating
genotypic variation in the genome of the plant.

<>

<1>Vairapandi, M., Duker, N.J.
<2>Enzymic removal of 5-methylcytosine from DNA by a hyman DNA-glycosylase.
<3>Nucleic Acids Res.
<4>21
<5>5323-5327
<6>1993
<7>DNA 5-methylcytosine is a major factor in the silencing of mammalian genes; it is involved in
gene expression, differentiation, embryogenesis and neoplastic transformation. A decrease in
DNA 5-methylcytosine content is associated with activation of specific genes. There is much
evidence indicating this to be an enzymic process, with replacement of 5-methylcytosine by
cytosine. We demonstrate here enzymic release of 5-methylcytosines from DNA by a human
5-methylcytosine-DNA glycosylase activity, which affords a possible mechanism for such
replacement. This activity generates promutagenic apyrimidinic sites, which can be related to
the high frequency of mutations found at DNA 5-methylcytosine loci. The recovery of most
released pyrimidines as thymines indicates subsequent deamination of free 5-methylcytosines by
a 5-methylcytosine deaminase activity. This prevents possible recycling of 5-methylcytosine
into replicative DNA synthesis via a possible 5-methyl-dCTP intermediate synthesized through
the pyrimidine salvage pathway. Taken together, these findings indicate mechanisms for removal
of 5-methylcytosines from DNA, hypermutability of DNA 5-methylcytosine sites, and exclusion of
5-methylcytosines from DNA during replication.

<>

<1>Vaishampayan, P.A., Kuehl, J.V., Froula, J.L., Morgan, J.L., Ochman, H., Francino, M.P.
<2>Comparative metagenomics and population dynamics of the gut microbiota in mother and infant.
<3>Genome Biol. Evol.
<4>2
<5>53-66
<6>2010
<7>Colonization of the gastrointestinal tract (GIT) of human infants with a suitable microbial
community is essential for numerous aspects of health, but the progression of events by which
this microbiota becomes established is poorly understood. Here, we investigate two previously
unexplored areas of microbiota development in infants: the deployment of functional
capabilities at the community level and the population genetics of its most abundant genera.
To assess the progression of the infant microbiota toward an adult-like state and to evaluate
the contribution of maternal GIT bacteria to the infant gut, we compare the infant's
microbiota with that of the mother at 1 and 11 months after delivery. These comparisons reveal
that the infant's microbiota rapidly acquires and maintains the range of gene functions
present in the mother, without replicating the phylogenetic composition of her microbiota.
Microdiversity analyses for Bacteroides and Bifidobacterium, two of the main microbiota
constituents, reveal that by 11 months, the phylotypes detected in the infant are distinct
from those in the mother, although the maternal Bacteroides phylotypes were transiently
present at 1 month of age. The configuration of genetic variants within these genera reveals
populations far from equilibrium and likely to be undergoing rapid growth, consistent with
recent population turnovers. Such compositional turnovers and the associated loss of maternal
phylotypes should limit the potential for long-term coadaptation between specific bacterial
and host genotypes.

<>

<1>Vaisvila, R., Kucera, R.B., Raleigh, E.A., Morgan, R.D., Claus, T.E.
<2>Method for cloning and producing the MseI restriction endonuclease.
<3>European Patent Office
<4>EP 1197549 A
<5>
<6>2001
<7>A method for cloning restriction-modification system is provided whereby the target
modification methylase is produced and confers full protection during all growth phases in
which the cognate restriction enzyme is present.  The method is employed in the cloning of the
MseI restriction-modification system.

<>

<1>Vaisvila, R., Morgan, R.D., Kucera, R.B., Claus, T.E., Raleigh, E.A.
<2>Process for cloning MseI-restricted endonuclease and process for producing the same.
<3>Japanese Patent Office
<4>JP 2002306186 A
<5>
<6>2002
<7>
<>

<1>Vaisvila, R., Morgan, R.D., Kucera, R.B., Claus, T.E., Raleigh, E.A.
<2>Method for cloning and producing the Msel restriction endonuclease.
<3>US Patent Office
<4>US 6846658 A
<5>
<6>2005
<7>A method for cloning restriction-modification system is provided whereby the target
modification methylase is produced and confers full protection during all growth phases in
which the cognate restriction enzyme is present.  The method is employed in the cloning of the
MseI restriction-modification system.

<>

<1>Vaisvila, R., Rasmussen, L.J., Lobner-Olesen, A., von Freiesleben, U., Marinus, M.G.
<2>The LipB protein is a negative regulator of dam gene expression in Escherichia coli.
<3>Biochim. Biophys. Acta
<4>1494
<5>43-53
<6>2000
<7>Transcription initiation of the major promoter (P2) of the Escherichia coli dam gene increases
with growth rate.  The presence of three partially palindromic motifs, (TTCAGT(N(20))TGAG),
designated G (growth)-boxes, within the -52 to +31 region of the promoter, may be related to
growth rate control. Deletion of two of these repeats, downstream of the transcription
initiation point, result in constitutive high activity of the promoter. The unlinked
cde-4::miniTn10 insertion also results in several fold higher activity of the dam P2 promoter,
suggesting that this mutation resulted in the loss of a putative dam P2 repressor. The cde-4
mutation was mapped to the lipB (lipoic acid) gene, which we show encodes a 24 kDa protein
initiating at a TTG codon. LipB is a highly conserved protein in animal and plant species,
other bacteria, Archaea, and yeast. Plasmids expressing the native or hexahistidine-tagged
LipB complement the phenotype of the cde-4 mutant strain. The level of LipB in vivo was higher
in exponentially growing cells than those in the stationary phase. Three G-box motifs were
also found in the lipB region. Models for the regulation of expression of the two genes are
discussed.

<>

<1>Vaisvila, R., Sliesaraviciute, Z., Kulakauskas, S., Janulaitis, A.
<2>Cloning of the ppu21IM gene using a in vivo selection method.
<3>Gene
<4>157
<5>55-57
<6>1995
<7>A genetic system enabling the in vivo selection of genes encoding the DNA-modifying enzymes
was developed.  A gene library is transformed into a strain harboring the
restriction-modification (R-M) system which a recognition sequence is a subset of the target
sequence of the DNA methyltransferase (MTase) to be cloned.  If the residing MTase is
temperature sensitive, the inability of transformants to grow at 42oC provides a simple and
convenient procedure for the isolation of new MTase-encoding genes.  The feasibility of this
procedure has been demonstrated by the isolation of the ppu21IM gene from a Pseudomonas putida
RFL21 gene library.

<>

<1>Vaisvila, R., Vilkaitis, G., Janulaitis, A.
<2>Identification of a gene encoding a DNA invertase-like enzyme adjacent to the PaeR7I restriction-modification system.
<3>Gene
<4>157
<5>81-84
<6>1995
<7>A gene encoding a DNA invertase-like enzyme was identified adjacent to the PaeR7I
restriction-modification system (R-M), and was named paeR7IN (N for iNvertase).  Sequence
analysis revealed that this gene has the same polarity as the PaeR7IRM operon, and would
encode a polypeptide of 21,506 Da.  An amino-acid sequence similarity of 45-49% was found
between the deduced protein product and various DNA invertases.

<>

<1>Vaitkevicius, D., Naureckiene, S., Janulaitis, A.
<2>Advanced method for obtaining highly purified restriction enzyme preparations.
<3>Biotekhnologiya
<4>1
<5>35-41
<6>1991
<7>Using Eco130I restrictase isolated from cells of Escherichia coli RFL 130, a
possibility was investigated for selecting a scheme for restriction enzyme
purification under static conditions by analyzing the binding of protein to
various absorbents and evaluating the functional purity of the samples of
enzyme preparations.  The nature of restrictase and adsorbent interaction under
static conditions made it possible to define the restrictase's sorption
behavior in column chromatography and to select chromatography conditions.
Evaluation of concomitant nonspecific nucleases allowed to specify the
adsorbents that could be most effective from the point of view of functional
purification.  A highly effective scheme for Eco130I restrictase purification
has been developed.  Fractionation of cell-free extracts of E. coli RFL 130 on
heparin-sepharose, DEAE-cellulose and phosphocellulose yielded an enzyme
preparation of high functional purity.

<>

<1>Valdes, J., Ossandon, F., Quatrini, R., Dopson, M., Holmes, D.S.
<2>Draft Genome Sequence of the Extremely Acidophilic Biomining Bacterium Acidithiobacillus thiooxidans ATCC 19377 Provides Insights into the  Evolution of the Acidithiobacillus Genus.
<3>J. Bacteriol.
<4>193
<5>7003-7004
<6>2011
<7>Acidithiobacillus thiooxidans is a mesophilic, extremely acidophilic, chemolithoautotrophic
gammaproteobacterium that derives energy from the
oxidation of sulfur and inorganic sulfur compounds. Here we present the
draft genome sequence of A. thiooxidans ATCC 19377, which has allowed the
identification of genes for survival and colonization of extremely acidic
environments.

<>

<1>Valdes, J., Quatrini, R., Hallberg, K., Dopson, M., Valenzuela, P.D., Holmes, D.S.
<2>Draft genome sequence of the extremely acidophilic bacterium Acidithiobacillus caldus ATCC 51756 reveals metabolic versatility in the genus Acidithiobacillus.
<3>J. Bacteriol.
<4>191
<5>5877-5878
<6>2009
<7>Acidithiobacillus caldus is an extremely acidophilic, moderately thermophilic,
chemolithoautotrophic gammaproteobacterium that derives energy from the oxidation
of sulfur and reduced inorganic sulfur compounds. Here we present the draft
genome sequence of Acidithiobacillus caldus ATCC 51756 (the type strain of the
species), which has permitted the prediction of genes for survival in extremely
acidic environments, including genes for sulfur oxidation and nutrient
assimilation.

<>

<1>Valdes, N., Rivera-Araya, J., Bijman, J., Escudero, L., Demergasso, C., Fernandez, S., Ferrer, A., Chavez, R., Levican, G.
<2>Draft Genome Sequence of Nitrincola sp. Strain A-D6, an Arsenic-Resistant Gammaproteobacterium Isolated from a Salt Flat.
<3>Genome Announcements
<4>2
<5>e01144-14
<6>2014
<7>We report Nitrincola sp. strain A-D6, which was characterized as an arsenic-resistant
bacterium isolated from the Ascotan Salt Flat in northern
Chile. The size of the genome is 3,795,776 bp, with a G+C content of 49.96%.
Genes for the arsenic-resistant Ars system and arsenic oxidation have been
encoded.

<>

<1>Vale, A., Vitor, J.
<2>Helicobacter pylori genomic DNA methylation status: strain typing.
<3>Int. J. Med. Microbiol.
<4>293
<5>109
<6>2003
<7>The complete genome sequence of two different strains of H. pylori, revealed extreme genetic
diversity.  It also showed that both strains have an unusually high number of restriction and
modification systems (RM).  Based on the knowledge that in RM systems the restriction
endonuclease (ENase) impose high pressure on the expression of the companion methyltransferase
(MTase) it was hypothesised that genomic DNA methylation status could be used to develop a new
typing method of H. pylori.  To test this hypothesis 51 strains of H. pylori were studied, 28
of which were epidemiologically related.  Genomic DNA was isolated and digested with 33
different ENases.  After reisolation of DNA from seven strains we replicated 3.5% of the ENase
digests (59/1683) with 100% concordance.  The results were grouped as a data matrix, where "0"
means presence of unmethylated DNA and "1" presence of methylated DNA.  Dendrograms were
constructed with UPGMA method and Jaccard coefficient. After statistical cluster analysis
(NTSYSpc) and correspondence factorial analysis (PSS 11.0) ENases were gradually eliminated
until a final group of 16 ENases were selected as a tool to type H. pylori: Acil, BssHII,
BstUI, DdeI, DraI, FauI, HaeIII, Hpy181I, Hpy99I, HpyCH4III, HpyCH4IV, HpyCH4V, MspI, Sau96I,
ScrFI and TaqI.  The method has high discriminatory power (Simpson index of diversity=0,99,
strain group frequency nj/N=0.039), has adequate cophenetic correlation coefficient, r=0.782
and high typability.  That is adequate for typing H. pylori. A high number of MTases were
expressed in all strains (between 14 and 22) and 9 of which were common (M.ApaI, M.BseRI,
M.DpnI, M.EagI, M.HhaI, M.HinT1I, M.NaeI, M.NgoMIV and M.NIaIII).

<>

<1>Vale, F.F., Encarnacao, P., Vitor, J.M.B.
<2>A new algorithm for cluster analysis of genomic methylation: the Helicobacter pylori case.
<3>Bioinformatics
<4>24
<5>383-388
<6>2008
<7>Motivation: The genomic methylation analysis is useful to type bacteria that have a high
number of expressed type II methyltransferases.
Methyltransferases are usually committed to Restriction and
Modification (R-M) systems, in which the restriction endonuclease
imposes high pressure on the expression of the cognate
methyltransferase that hinder R-M system loss. Conventional cluster
methods do not reflect this tendency. An algorithm was developed for
dendrogram construction reflecting the propensity for conservation of
R-M Type II systems.
Results: The new algorithm was applied to 52 Helicobacter pylori
strains from different geographical regions and compared with
conventional clustering methods. The algorithm works by first grouping
strains that share a common minimum set of R-M systems and gradually
adds strains according to the number of the R-M systems acquired.
Dendrograms revealed a cluster of African strains, which suggest that
R-M systems are present in H.pylori genome since its human host
migrates from Africa.

<>

<1>Vale, F.F., Megraud, F., Vitor, J.M.
<2>Geographic distribution of methyltransferases of Helicobacter pylori: evidence of human host population isolation and migration.
<3>BMC Microbiol.
<4>9
<5>193
<6>2009
<7>ABSTRACT: BACKGROUND: Helicobacter pylori colonizes the human stomach and is associated with
gastritis, peptic ulcer, and gastric cancer. This
ubiquitous association between H. pylori and humans is thought to be
present since the origin of modern humans. The H. pylori genome encodes
for an exceptional number of restriction and modifications (R-M) systems.
To evaluate if R-M systems are an adequate tool to determine the
geographic distribution of H. pylori strains, we typed 221 strains from
Africa, America, Asia, and Europe, and evaluated the expression of 29
methyltransferases. RESULTS: Independence tests and logistic regression
models revealed that 10 R-M systems correlate with geographical
localization. The distribution pattern of these methyltransferases may
have been originated by co-divergence of regional H. pylori after its
human host migrated out of Africa. The expression of specific
methyltransferases in the H. pylori population may also reflect the
genetic and cultural background of its human host. Methyltransferases
common to all strains, M.HhaI and M.NaeI, are likely conserved in H.
pylori, and may have been present in the bacteria genome since the human
diaspora out of Africa. CONCLUSIONS: This study indicates that
methyltransferases are useful geomarkers, which allow discrimination of
bacterial populations, and that can be added to our tools to investigate
human migrations.

<>

<1>Vale, F.F., Vitor, J.M.
<2>Genomic Methylation: a Tool for Typing Helicobacter pylori Isolates.
<3>Appl. Environ. Microbiol.
<4>73
<5>4243-4249
<6>2007
<7>The genome sequences of three Helicobacter pylori strains revealed an abundant number of
putative restriction and modification (R-M) systems
within a small genome (1.60 to 1.67 Mb). Each R-M system includes an
endonuclease that cleaves a specific DNA sequence and a DNA
methyltransferase that methylates either adenosine or cytosine within the
same DNA sequence. These are believed to be a defense mechanism,
protecting bacteria from foreign DNA. They have been classified as selfish
genetic elements; in some instances it has been shown that they are not
easily lost from their host cell. Possibly because of this phenomenon, the
H. pylori genome is very rich in R-M systems, with considerable variation
in potential recognition sequences. For this reason the protective aspect
of the methyltransferase gene has been proposed as a tool for typing H.
pylori isolates. We studied the expression of H. pylori methyltransferases
by digesting the genomic DNAs of 50 strains with 31 restriction
endonucleases. We conclude that methyltransferase diversity is
sufficiently high to enable the use of the genomic methylation status as a
typing tool. The stability of methyltransferase expression was assessed by
comparing the methylation status of genomic DNAs from strains that were
isolated either from the same patient at different times or from different
stomach locations (antrum and corpus). We found a group of five
methyltransferases common to all tested strains. These five may be
characteristic of the genetic pool analyzed, and their biological role may
be important in the host/bacterium interaction.

<>

<1>Vale, F.F., Vitor, J.M.B.
<2>Genomic methylation status for discrimination among Helicobacter species: A bioinformatics approach.
<3>J. Proteomics Bioinformatics
<4>1
<5>258-266
<6>2008
<7>The genus Helicobacter comprises several species of both gastric and enterohepatic intestinal
bacteria. H. pylori, the type species of the genus, is associated with gastritis, peptic ulcer
and gastric cancer in humans. H. pylori genome has a high number of restriction and
modification (R-M) systems and their diversity is useful for strain typing. To analyse if such
a high number of expressed methyltransferases is a characteristic of the genus Helicobacter,
the genomic methylation of five non-pylori Helicobacter spp. (H. canadensis, H. canis, H.
felis, H.mustelae and H. pullorum) was determined. The results revealed that the number of R-M
systems among nonpylori Helicobacter spp. is smaller than those observed among a group of 221
H. pylori strains (p<0,001), but is greater than those observed for the mean of all bacteria
sequenced genomes (p=0,005). 16S ribosomal RNA analysis of H. pylori sequenced strains and
five non-pylori Helicobacter spp. clearly isolate H. pylori species.  Surprisingly, the
analysis of the genomic methylation status by MCRM algorithm performs similarly. This suggests
that R-M systems do not appear to be spread in a miscellaneous manner, once even that these
genes may be subjected to acquisition and loss; their expression still allows discriminating
among Helicobacter spp.

<>

<1>Valenzuela, C., Ugalde, J.A., Mora, G.C., Alvarez, S., Contreras, I., Santiviago, C.A.
<2>Draft Genome Sequence of Salmonella enterica Serovar Typhi Strain STH2370.
<3>Genome Announcements
<4>2
<5>e00104-14
<6>2014
<7>We report the draft genome sequence of Salmonella enterica serovar Typhi strain STH2370,
isolated from a typhoid fever patient in Santiago, Chile. This clinical
isolate has been used as the reference wild-type strain in numerous studies
conducted in our laboratories during the last 15 years.

<>

<1>Valinluck, B.
<2>Characterization of restriction-modification systems in Klebsiella pneumoniae.
<3>Diss. Abstr.
<4>53
<5>2701B
<6>1992
<7>Two restriction-modification (R-M) systems, KpnAI and KpnBI, found in Klebsiella pneumoniae
strains M5a1 and GM236, respectively, have been studied and confirmed to be different from
other R-M systems reported in K. pneumoniae. Mutant studies suggest that the KpnAI and KpnBI
systems may belong to either a type I or type III system, since approximately equal numbers of
r-m+ and r-m- mutants were obtained. However, a DNA hybridization study using representative
type I and type III probes from E. coli and S. typhimurium failed to show homologies to either
KpnAI or KpnBI. The restriction endonuclease KpnBI was found to be temperature-sensitive with
maximum restriction activity at 30oC and no restriction activity at 42oC. Further, the
activity of endonuclease KpnBI was found to be reduced to almost zero level by growing the
bacteria in the presence of 10% glycerol. Although the mechanism is not known, this is the
first time such a phenomenon has been observed in any of the reported R-M systems. These
studies also compared the efficiency of transformation in K. pneumoniae of three plasmid
transformation methods; CaCl2 heat-shock; freezing and thawing in the presence of CaCl2; and
electroporation. Electroporation was shown to be the most efficient method. Transformation
efficiency in both the r+KpnAI and r+KpnBI strains was 20- to 100-fold less than the
transformation efficiency of the r- strains, depending on plasmid size. Four different
approaches have been used to clone the hsd genes of the KpnBI system. Two clones were
obtained; these were named pKpnB1 and pKpnB2. The pKpnB1 and pKpnB2 clones were found to
complement the restriction activity of an r- KpnBIm+KpnBI K. pneumoniae mutant and were also
found to complement both the restriction and modification activities of an r-KpnBIm-KpnBI K.
pneumoniae mutant. A quick subcloning method which involves making subclones from a plasmid
clone in a single step was also developed. A preliminary analysis, based on complementation
studies, of the gene structure suggested that the KpnBI system may consist of three structural
genes, a characteristic of the type I R-M system.

<>

<1>Valinluck, B., Lee, N.S., Ryu, J.
<2>A new restriction-modification system, KpnBI, recognized in Klebsiella pneumoniae.
<3>Gene
<4>167
<5>59-62
<6>1995
<7>A unique DNA restriction-modification (R-M) system has been identified in the GM236 strain of
Klebsiella pneumoniae using the newly isolated phage, SBS.  The system was designated KpnBI.
The gene (hsdRKpnBI) complementing the restriction activity of the KpnBI system was cloned in
pBR322.  The nucleotide sequence of the cloned DNA revealed one open reading frame (ORF) of
3035 bp.  Analysis of the deduced amino-acid sequence shows seven helicase motifs which are
common to the restriction (R) subunit of both type-I and type-III R-M systems.  Computer
analysis (Dendrogram) of the R polypeptide of KpnBI suggests a closer relationship to
EcoR124/3I, a member of type-IC family, than to other representative type-I and type-III
systems.

<>

<1>Valinluck, B., Reyno, M., Ryu, J.
<2>Two new restriction-modification systems, KpnA and KpnB, in Klebsiella pneumoniae.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>89
<5>180
<6>1989
<7>Strains of K. pneumoniae, M5a1 and GM236, each possess a unique restriction-modification (R-M)
system. These two systems differ from KpnI, a type II system reported previously in K.
pneumoniae, since the KpnI endonuclease degrades chromosomal DNA from both M5a1 and GM236. No
plasmids or type II restriction endonuclease activity have so far been identified from these
two strains. Mutant analysis suggests that these two Klebsiella R-M systems may be classified
as type I since almost equal numbers of r- m+ and r- m- mutants were obtained from a single
mutagenic treatment. DNA hybridization revealed a weak but significant homology between an
hsdSB probe derived from S. typhimurium and GM236 chromosome. No homology was observed between
an hsdA probe derived from E. coli 15T and chromosomal DNA from either of the two Klebsiella
strains. A 7.2 kb EcoRI fragment of GM236 identified by its hybridization to the S.
typhimurium hsdSB probe was cloned in pBluescript (Stratagene). Only one PvuII fragment (1.7
kb) derived from it showed hybridization to hsdM region of hsdSB and hsdK (from E. coli K12)
clones. From the above observations, we concluded that M5a1 and GM236 have new R-M systems
which we designated provisionally KpnA and KpnB, respectively.

<>

<1>Valinluck, V., Sowers, L.C.
<2>Endogenous cytosine damage products alter the site selectivity of human DNA maintenance methyltransferase DNMT1.
<3>Cancer Res.
<4>67
<5>946-950
<6>2007
<7>Alterations in cytosine methylation patterns are usually observed in human tumors. The
consequences of altered cytosine methylation patterns
include both inappropriate activation of transforming genes and
silencing of tumor suppressor genes. Despite the biological effect of
methylation changes, little is known about how such changes are caused.
The heritability of cytosine methylation patterns from parent to
progeny cells is attributed to the fidelity of the
methylation-sensitive human maintenance methyltransferase DNMT1, which
methylates with high specificity the unmethylated strand of a
hemimethylated CpG sequence following DNA replication. We have been
studying DNA damage that might alter the specificity of DNMTl, either
inhibiting the methylation of hemimethylated sites or triggering the
inappropriate methylation of previously unmethylated sites. Here, we
show that known forms of endogenous DNA damage can cause either
hypermethylation or hypomethylation. Inflammation-induced 5-halogenated
cytosine damage products, including 5-chlorocytosine, mimic
5-methylcytosine and induce inappropriate DNMTl methylation within a
CpG sequence. In contrast, oxidation damage of the methyl group of
5-methylcytosine, with the formation of 5-hydroxymethylcytosine,
prevents DNMTI methylation of the target cytosine. We propose that
reduced DNMTI selectivity resulting from DNA damage could cause
heritable changes in cytosine methylation patterns, resulting in human
tumor formation. These data may provide a mechanistic link for the
associations documented between inflammation and cancer.

<>

<1>Valinluck, V., Wu, W., Liu, P.F., Neidigh, J.W., Sowers, L.C.
<2>Impact of cytosine 5-halogens on the interaction of DNA with restriction endonucleases and methyltransferase.
<3>Chem. Res. Toxicol.
<4>19
<5>556-562
<6>2006
<7>Growing evidence from both prokaryotes and eukaryotes indicates that pyrimidine 5-methyl
groups can have profound biological consequences
that are mediated by the affinity of DNA-protein interactions. The
presence of the 5-methyl group could potentially create a steric block
preventing the binding of some proteins whereas the affinity of many
other proteins is substantially increased by pyrimidine methylation. In
this paper, we have constructed a series of oligonucleotides containing
cytosine and a series of 5-substituted cytosine analogues including all
halogens. This set of oligonucleotides has been used to probe the
relationship between the size of the substituent and its capacity to
modulate cleavage by the methylation-sensitive restriction
endonucleases MspI and HpaII. Additionally, we have examined the impact
of the halogen substitution on the corresponding bacterial
methyltransferase (M.HpaII). We observed that MspI cleavage is only
subtly affected by substituted cytosine analogues at the inner position
of the CCGG recognition site. In contrast, HpaII cleaves
cytosine-containing oligonucleotides completely whereas
5-fluorocytosine-containing oligonucleotides are cleaved at a reduced
rate. The presence of the larger halogens Cl, Br, or I as well as a
methyl group completely prevents cleavage by HpaII. These data suggest
that the steric wall is encountered by HpaII slightly beyond the
fluorine substituent, at about 2.65 angstrom from the pyrimidine
C5-position. It is known that 5-fluorocytosine in an oligonucleotide
can form a covalent irreversible suicide complex with either
prokaryotic or eukaryotic methyltransferases. Kinetic data reported
here suggest that the 5-fluorocytosine-containing oligonucleotide can
also inhibit M.HpaII by formation of a reversible, noncovalent complex.
Our results indicate that although a 5-Cl substituent has electronic
properties similar to 5-F, 5-chlorocytosine duplexes neither form a
complex with M.HpaII nor inhibit enzymatic methylation. Emerging data
suggest that halogenation of cytosine can occur in DNA in vivo from
inflammation-mediated reactive molecules. The results reported here
suggest that the inadvertent halogenation of cytosine residues in DNA
could alter the affinity of sequence-specific DNA-binding proteins.

<>

<1>Vallenet, D. et al.
<2>Comparative analysis of Acinetobacters: three genomes for three lifestyles.
<3>PLoS ONE
<4>3
<5>e1805
<6>2008
<7>Acinetobacter baumannii is the source of numerous nosocomial infections in humans and
therefore deserves close attention as multidrug or even pandrug
resistant strains are increasingly being identified worldwide. Here we
report the comparison of two newly sequenced genomes of A. baumannii. The
human isolate A. baumannii AYE is multidrug resistant whereas strain SDF,
which was isolated from body lice, is antibiotic susceptible. As reference
for comparison in this analysis, the genome of the soil-living bacterium
A. baylyi strain ADP1 was used. The most interesting dissimilarities we
observed were that i) whereas strain AYE and A. baylyi genomes harbored
very few Insertion Sequence elements which could promote expression of
downstream genes, strain SDF sequence contains several hundred of them
that have played a crucial role in its genome reduction (gene disruptions
and simple DNA loss); ii) strain SDF has low catabolic capacities compared
to strain AYE. Interestingly, the latter has even higher catabolic
capacities than A. baylyi which has already been reported as a very
nutritionally versatile organism. This metabolic performance could explain
the persistence of A. baumannii nosocomial strains in environments where
nutrients are scarce; iii) several processes known to play a key role
during host infection (biofilm formation, iron uptake, quorum sensing,
virulence factors) were either different or absent, the best example of
which is iron uptake. Indeed, strain AYE and A. baylyi use
siderophore-based systems to scavenge iron from the environment whereas
strain SDF uses an alternate system similar to the Haem Acquisition System
(HAS). Taken together, all these observations suggest that the genome
contents of the 3 Acinetobacters compared are partly shaped by life in
distinct ecological niches: human (and more largely hospital environment),
louse, soil.

<>

<1>Vallone, P.M., Benight, A.S.
<2>Thermodynamic, spectroscopic, and equilibrium binding studies of DNA sequence context effects in four 40 base pair deoxyoligonucleotides.
<3>Biochemistry
<4>39
<5>7835-7846
<6>2000
<7>Effects of different end sequences on melting, circular dichroism spectra (CD), and enzyme
binding properties were investigated for four 40 base pair, non-self-complementary duplex DNA
oligomers. The center sequences of these oligoduplexes have either of two 22 base pair modules
flanked on both sides by sequences differing in AT content. Temperature-induced melting
transitions monitored by differential scanning calorimetry (DSC) and ultraviolet absorbance
were measured for the six duplexes in buffered 115 mM Na(+) solutions. Values of the melting
transition enthalpy, DeltaH(cal), and entropy, DeltaS(cal), were obtained directly from DSC
experiments. Melting transition parameters, DeltaH(vH) and DeltaS(vH), were also estimated
from a van't Hoff analysis of optical melting curves collected as a function of DNA
concentration, assuming that the melting transition is two-state. Melting free energies (20
degrees C) evaluated from DSC melting experiments on the four duplex DNAs ranged from -52.2 to
-77.5 kcal/mol. Free energies based on the van't Hoff analysis were -37.9 to -58.8 kcal/mol.
Although the values are different, trends in the melting free energies of the four duplex DNAs
as a function of sequence were identical in both DSC and optical analyses. Subject to several
assumptions, values for the initiation free energy were estimated for each duplex, defined as
DeltaG(int) = DeltaG(cal) - DeltaG(pred), where DeltaG(cal) is the experimental free energy at
20 degrees C determined from the experimentally measured values of the transition enthalpy,
DeltaH(cal), and entropy, DeltaS(cal). The predicted free energy of the sequence,
DeltaG(pred)(20 degrees C), is obtained using published nearest-neighbor sequence stability
values. For three of the four duplexes, values of DeltaG(int) are essentially nil. In
contrast, the duplex with 81.8% GC has a considerably higher estimate of DeltaG(int) = 7.1
kcal/mol. The CD spectra for the six duplexes collected over the wavelength range from 200 to
320 nm are also sequence-dependent. Factor analysis of the CD spectra by singular value
decomposition revealed that the experimental CD spectra could be reconstructed from linear
combinations of two minor and one major subspectra. Changes in the coefficients of the major
subspectrum for different sequences reflect incremental sequence-dependent variations of the
CD spectra. Equilibrium binding by BamHI restriction endonuclease to the 40 base pair DNAs
whose central eight base pairs contain the recognition sequence for BamHI restriction enzyme
bounded by A.T base pairs, 5'-A-GGATCC-A-3' was investigated. Binding assays were performed
by titering BamHI against a constant concentration of each of the duplex DNA substrates, in
the absence of Mg(2+), followed by analysis by gel retardation. Under the conditions employed,
the enzyme binds but does not cleave the DNAs. Results of the assays revealed two binding
modes with retarded gel mobilities. Binding isotherms for the fraction of bound DNA species
versus enzyme concentration for each binding mode were constructed and analyzed with a simple
two-step equilibrium binding model. This analysis provided semiquantitative estimates on the
equilibrium binding constants for BamHI to the four DNAs. Values obtained for the binding
constants varied only 7-fold and ranged from 6 x 10^-8 to 42 x 10^-8 M, with binding
free energies from -8.6 to -9.7 (+/- 0.2) kcal/mol depending on the sequence that flanks the
enzyme binding site. Unlike what was found earlier in binding studies of the 22 base pair
duplexes that constitute the core modules of the present 40-mers [Riccelli, P. V., Vallone, P.
M., Kashin, I., Faldasz, B. D., Lane, M. J., and Benight, A. S. (1999) Biochemistry 38,
11197-11208], no obvious relationship between binding and stability was found for these longer
DNAs. Apparently, effects of flanking sequence stability on restriction enzyme binding may
only be measurable in very short duplex deoxyoligonucleotides.

<>

<1>Valot, B., Rohmer, L., Jacobs, M.A., Miller, S.I., Bertrand, X., Hocquet, D.
<2>Comparative Genomic Analysis of Two Multidrug-Resistant Clinical Isolates of ST395 Epidemic Strain of Pseudomonas aeruginosa Obtained 12 Years Apart.
<3>Genome Announcements
<4>2
<5>e00515-14
<6>2014
<7>Pseudomonas aeruginosa can cause large and prolonged outbreaks in hospitals. We have sequenced
and annotated the genomes of two multidrug-resistant P. aeruginosa
isolates from the same strain obtained 12 years apart from different patients.
Genomic analysis provided insight on the genes acquired and lost by P. aeruginosa
during its spread.

<>

<1>Valton, J., Daboussi, F., Leduc, S., Molina, R., Redondo, P., Macmaster, R., Montoya, G., Duchateau, P.
<2>5 '-Cytosine-Phosphoguanine (CpG) Methylation Impacts the Activity of Natural and Engineered Meganucleases.
<3>J. Biol. Chem.
<4>287
<5>30139-30150
<6>2012
<7>In this study, we asked whether CpG methylation could influence the DNA binding affinity and
activity of meganucleases used for genome
engineering applications. A combination of biochemical and structural
approaches enabled us to demonstrate that CpG methylation decreases
I-CreI DNA binding affinity and inhibits its endonuclease activity in
vitro. This inhibition depends on the position of the methylated
cytosine within the DNA target and was almost total when it is located
inside the central tetrabase. Crystal structures of I-CreI bound to
methylated cognate target DNA suggested a molecular basis for such
inhibition, although the precise mechanism still has to be specified.
Finally, we demonstrated that the efficacy of engineered meganucleases
can be diminished by CpG methylation of the targeted endogenous site,
and we proposed a rational design of the meganuclease DNA binding
domain to alleviate such an effect. We conclude that although activity
and sequence specificity of engineered meganucleases are crucial
parameters, target DNA epigenetic modifications need to be considered
for successful gene editions.

<>

<1>van Aalten, D.M.F., Erlanson, D.A., Verdine, G.L., Joshua-Tor, L.
<2>A structural snapshot of base-pair opening in DNA.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>11809-11814
<6>1999
<7>The response of double-helical DNA to torsional stress may be a driving force for many
processes acting on DNA. The 1.55-A crystal structure of a duplex DNA oligonucleotide
d(CCAGGCCTGG)(2) with an engineered crosslink in the minor groove between the central guanine
bases depicts how the duplex can accommodate such torsional stress. We have captured in the
same crystal two rather different conformational states. One duplex contains a strained
crosslink that is stabilized by calcium ion binding in the major groove, directly opposite the
crosslink. For the other duplex, the strain in the crosslink is relieved through partial
rupture of a base pair and partial extrusion of a cytosine accompanied by helix bending. The
sequence used is the target sequence for the HaeIII methylase, and this partially flipped
cytosine is the same nucleotide targeted for extrusion by the enzyme. Molecular dynamics
simulations of these structures show an increased mobility for the partially flipped-out
cytosine.

<>

<1>van Aelst, K., Sisakova, E., Szczelkun, M.D.
<2>DNA cleavage by Type ISP Restriction-Modification enzymes is initially targeted to the 3'-5' strand.
<3>Nucleic Acids Res.
<4>41
<5>1081-1090
<6>2013
<7>The mechanism by which a double-stranded DNA break is produced following collision of two
translocating Type I Restriction-Modification enzymes is not
fully understood. Here, we demonstrate that the related Type ISP
Restriction-Modification enzymes LlaGI and LlaBIII can cooperate to cleave DNA
following convergent translocation and collision. When one of these enzymes is a
mutant protein that lacks endonuclease activity, DNA cleavage of the 3'-5' strand
relative to the wild-type enzyme still occurs, with the same kinetics and at the
same collision loci as for a reaction between two wild-type enzymes. The DNA
nicking activity of the wild-type enzyme is still activated by a protein variant
entirely lacking the Mrr nuclease domain and by a helicase mutant that cannot
translocate. However, the helicase mutant cannot cleave the DNA despite the
presence of an intact nuclease domain. Cleavage by the wild-type enzyme is not
activated by unrelated protein roadblocks. We suggest that the nuclease activity
of the Type ISP enzymes is activated following collision with another Type ISP
enzyme and requires adenosine triphosphate binding/hydrolysis but, surprisingly,
does not require interaction between the nuclease domains. Following the initial
rapid endonuclease activity, additional DNA cleavage events then occur more
slowly, leading to further processing of the initial double-stranded DNA break.

<>

<1>van Aelst, K., Toth, J., Ramanathan, S.P., Schwarz, F.W., Seidel, R., Szczelkun, M.D.
<2>Type III restriction enzymes cleave DNA by long-range interaction between sites in both head-to-head and tail-to-tail inverted repeat.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>9123-9128
<6>2010
<7>Cleavage of viral DNA by the bacterial Type III Restriction-Modification enzymes requires the
ATP-dependent long-range communication between a
distant pair of DNA recognition sequences. The classical view is that Type
III endonuclease activity is only activated by a pair of asymmetric sites
in a specific head-to-head inverted repeat. Based on this assumption and
due to the presence of helicase domains in Type III enzymes, various
motor-driven DNA translocation models for communication have been
suggested. Using both single-molecule and ensemble assays we demonstrate
that Type III enzymes can also cleave DNA with sites in tail-to-tail
repeat with high efficiency. The ability to distinguish both inverted
repeat substrates from direct repeat substrates in a manner independent of
DNA topology or accessory proteins can only be reconciled with an
alternative sliding mode of communication.

<>

<1>van Belkum, A., Jacobs, B., van Beek, E., Louwen, R., van Rijs, W., Debruyne, L., Gilbert, M., Li, J., Jansz, A., Megraud, F., Endtz, H.
<2>Can Campylobacter coli induce Guillain-Barre syndrome?
<3>Eur. J. Clin. Microbiol. Infect. Dis.
<4>28
<5>557-560
<6>2009
<7>Campylobacter jejuni enteritis is the most frequently identified infection preceding the
Guillain-Barre syndrome (GBS) and neural damage is thought to be induced through molecular
mimicry between C. jejuni lipo-oligosaccharide (LOS) and human gangliosides [1]. It has been
questioned whether or not other Campylobacter species, including C. curvus, C. upsaliensis and
C. coli, could be similarly involved [2-4]. This is relevant because it would imply that
bacterial factors considered important in the aetiology of GBS crossed species barriers. Two
prior reports have appeared where C. coli was putatively associated with a case of GBS.

<>

<1>van Bemmel, D.M., Brank, A.S., Eritja, R., Marquez, V.E., Christman, J.K.
<2>DNA (Cytosine-C5) methyltransferase inhibition by oligodeoxyribonucleotides containing 2-(1H)-pyrimidinone (zebularine aglycon) at the enzymatic target site.
<3>Biochem. Pharmacol.
<4>78
<5>633-641
<6>2009
<7>Aberrant cytosine methylation in promoter regions leads to gene silencing associated with
cancer progression. A number of DNA
methyltransferase inhibitors are known to reactivate silenced genes:
including 5-azacytidine and 2-(1H)-pyrimidinone riboside (zebularine).
Zebularine is a more stable, less cytotoxic inhibitor compared to
5-azacytidine. To determine the mechanistic basis for this difference,
we carried out a detailed comparisons of the interaction between
purified DNA methyltransferases and oligodeoxyribonucleotides (ODNs)
containing either 5-azacytosine or 2-(1H)-pyrimidinone in place of the
cytosine targeted for methylation. When incorporated into small ODNs,
the rate of C5 DNA methyltransferase inhibition by both nucleosides is
essentially identical. However, the stability and reversibility of the
enzyme complex in the absence and presence of cofactor differs.
5-Azacytosine ODNs form complexes with C5 DNA methyltransferases that
are irreversible when the 5-azacytosine ring is intact. ODNs containing
2-(1H)-pyrimidinone at the enzymatic target site are competitive
inhibitors of both prokaryotic and mammalian DNA C5 methyltransferases.
We determined that the ternary complexes between the enzymes,
2-(1H)-pyrimidinone inhibitor, and the cofactor S-adenosyl methionine
are maintained through the formation of a reversible covalent
interaction. The differing stability and reversibility of the covalent
bonds may partially account for the observed differences in
cytotoxicity between zebularine and 5-azacytidine inhibitors.

<>

<1>Van Bemmel, D.M., Marquez, V.E., Christman, J.K.
<2>Characterization of the cytidine analog zebularine as an inhibitor of mammalian DNA (cytosine C5)-methyltransferase.
<3>Proc. Amer. Assoc. Cancer Res.
<4>44
<5>430
<6>2003
<7>Aberrant methylation of the promoter regions of genes has been shown to result in gene
silencing associated with cancer progression.  Reactivation of silenced genes in cultured
cells by methylation inhibitors can be induced by 5-azacytidine (5-AzaC) and
5-aza-2-deoxycytidine (5-AzadC), which inhibit DNA methyltransferases when incorporated into
DNA.  Since both 5-AzaC and 5-AzadC are chemically unstable and toxic, these results have
stimulated the search for alternate inhibitors of the mammalian methyltransferase DNMT1.  We
analyzed the inhibitory capacity of the cytidine analog zebularine (2-H pyrimidione).
Zebularine is stable under both acidic and neutral conditions, and is minimally cytotoxic,
making it a promising therapeutic agent.  We show that, when incorporated in place of the
cytosine target in double stranded or looped oligodeoxyribonucleotides containing recognition
sites for the bacterial methyltransferase (M.HhaI) and DNMT1 respectively, zebularine is
equivalent to
5-AzaC as an inhibitor of DNA methylation.  Despite the effective inhibition of DNMT1 in vitro
by ODNs containing zebularine, no decrease in DNA methylation was observed in LNCaP cells
treated with zebularine.  GSTP1 transcription was detected by RT-PCR after 5-AzadC treatment
but not after zebularine treatment.  Analysis of the heavily methylated promoter region of the
GSTP1 gene by bisulfite sequencing of DNA showed that DNA methylation was inhibited by 5-AzadC
but not zebularine.  Interestingly, LNCaP cells grown in the presence of zebularine did
display a marked change in morphology.  These results suggest that zebularine may not be
efficiently incorporated into DNA in vivo, but may be able to alter cellular phenotype by
interfering with other processes including RNA methylation.

<>

<1>van Blokland, R., Ross, S., Corrado, G., Scollan, C., Meyer, P.
<2>Developmental abnormalities associated with deoxyadenosine methylation in transgenic tobacco.
<3>Plant J.
<4>15
<5>543-551
<6>1998
<7>As in other higher eukaryotes, DNA methylation in plants is predominantly found at
deoxycytosine residues, while deoxyadenosine residues are not methylated at significant
levels. 6mdA methylation has been successfully introduced into yeast and Drosophila via
expression of a heterologous methyltransferase, but similar attempts in tobacco had, up until
now, proved unsuccessful despite the correct expression of a methyltransferase construct. It
was unclear whether this result reflected the failure of heterologous methyltransferases to
enter the nucleus, or whether 6mdA methylation, which has been shown to interfere with
promoter activity, was toxic for plants. Here we show that 6mdA methylation can be
successfully introduced into transgenic tobacco plants via expression of the bacterial dam
enzyme. The efficiency of 6mdA methylation was directly proportional to expression levels of
the dam construct, and methylation of all GATC sites was observed in a highly expressing line.
Increasing expression levels of the enzyme in different plants correlated with increasingly
abnormal phenotypes affecting leaf pigmentation, apical dominance, and leaf and floral
structure. Whilst introduction of dam-specific methylation does not cause any developmental
abnormalities in yeast or Drosophila, our data suggest that methylation of deoxyadenine
residues in plants interferes with the expression of genes involved in leaf and floral
development.

<>

<1>van Boven, C.P.A.
<2>Restriction and modification of phages in Staphylococcal phage typing.
<3>Ann. NY Acad. Sci.
<4>236
<5>376-388
<6>1974
<7>Staphylococcus aureus is a remarkably versatile species, which has been very successful in
establishing itself as a commensal in various niches on the human and animal body.  No two
staphylococci are really alike, even if they have the same phage pattern or drug sensitivity.

<>

<1>Van Cleve, M., Gumport, R.I.
<2>Effect of flanking sequence length on EcoRI endonuclease reaction.
<3>Fed. Proc.
<4>46
<5>2041
<6>1987
<7>Michaelis-Menten parameters were determined for the cleavages by EcoRI
endonuclease of both scission points in a duplex composed of the pentadecamer
d(pATCGAATTCCGGCCA) and its complement.  The Km for this substrate is 38.6 +/-
2 nM, a value between those determined for plasmid DNA and an octameric
oligodeoxyribonucleotide.  Since the two scission points are flanked by
different lengths of sequence, comparison of turnover numbers for the two
cleavages may reflect interactions of the enzyme with nucleotides outside its
recognition sequence.  Preliminary results indicate that the scissile bond in
the above indicated oligomer is cleaved twice as fast as the hydrolyzed bond in
its complement.  The more rapidly cleaved bond is four residues from the 5'
terminus, whereas the more slowly hydrolyzed bond is seven base pairs from the
end of the duplex.

<>

<1>Van Cleve, M., Gumport, R.I.
<2>Effect of flanking sequence length on EcoRI endonuclease reaction.
<3>J. Cell Biol.
<4>107
<5>403a
<6>1988
<7>Michaelis-Menten parameters were determined for the cleavages by EcoRI
endonuclease at both scission points in a series of oligomeric DNA duplexes.
Each member of the series contains the EcoRI recognition sequence (GAATTC)
located with six base pairs flanking the 3' end and from zero to three base
pairs flanking the 5' end, e.g., ATCGAATTCCGGCCA.  Thus, cleavage at both
strands produces two different sized and 32P-labeled products, in the above
case a tetramer and heptamer.  Kinetic parameters were determined for both
strands independently.  In each duplex tested, the cleavage rates are different
for the two oligomers, with the rate for each strand varying inversely as the
length of its 5' flanking sequence from Kcat=11.5 min-1 with three base pairs,
to 14.0 min-1 with two, to 20.0 min-1 with one.  Km's also vary with length,
and are different for the two strands at each duplex.

<>

<1>Van Cleve, M.D., Gumport, R.I.
<2>Influence of enzyme-substrate contacts located outside the EcoRI recognition site on cleavage of duplex oligodeoxyribonucleotide substrates by EcoRI endonuclease.
<3>Biochemistry
<4>31
<5>334-339
<6>1992
<7>A complete understanding of the sequence-specific interaction between the EcoRI
restriction endonuclease and its DNA substrate requires identification of all
contacts between the enzyme and substrate, and evaluation of their
significance.  We have searched for possible contacts adjacent to the
recognition site, GAATTC, by using a series of substrates with differing
lengths of flanking sequence.  Each substrate is a duplex of
non-self-complementary oligodeoxyribonucleotides in which the recognition site
is flanked by six base pairs on one side and from zero to three base pairs on
the other.  Steady-state kinetic values were determined for the cleavage of
each strand of these duplexes.  A series of substrates in which the length of
flanking sequences was varied on both sides of the hexamer was also examined.
The enzyme cleaved both strands of each of the substrates.  Decreasing the
flanking sequence to fewer than three base pairs on one side of the recognition
site induced an asymmetry in the rates of cleavage of the two strands.  The
scissile bond nearest the shortening sequence was hydrolyzed with increasing
rapidity as base pairs were successively removed.  Taken together, the Km and
kcat values obtained may be interpreted to indicate the relative importance of
several likely enzyme-substrate contacts located outside the canonical
hexameric recognition site.

<>

<1>Van Cott, E., Wilson, G.
<2>Method for producing the FnuDI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0324248 A
<5>
<6>1987
<7>The present invention is directed to a method for cloning and producing the FnuDI restriction
endonuclease by 1) introducing the restriction endonuclease gene from F. nucleatum D into a
host whereby the restriction gene is expressed; 2) fermenting the host which contains the
vector encoding and expressing the FnuDI restriction endonuclease, and 3) purifying the FnuDI
restriction endonuclease from the fermented host which contains the vector encoding and
expressing the FnuDI restriction endonuclease activity.

<>

<1>Van Cott, E., Wilson, G.
<2>Method for producing the XbaI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0323081 B
<5>
<6>1995
<7>In accordance with the present invention there is provided a clone containing the genes for
the XbaI restriction endonuclease and modification methylase derived from Xanthomonas badrii -
(ATCC 11672), as well as related methods for the production of the enzymes. More specifically,
this invention relates to clones which express the restriction endonuclease XbaI, an enzyme
which recognizes the DNA sequence TCTAGA and cleaves between the T and C. XbaI restriction
endonuclease produced in accordance with the present invention is free of the contaminants
found in XbaI preparations made from Xanthomonas badrii. The preferred method for cloning this
enzyme comprises forming a library containing the DNA from X.badrii, isolating clones which
contain the XbaI modification methylase gene and screening among these to identify those that
also contain the XbaI restriction endonuclease gene.

<>

<1>Van Cott, E., Wilson, G.G.
<2>Method for the production of the XbaI restriction endonuclease and methylase.
<3>Japanese Patent Office
<4>JP 2746964
<5>
<6>1998
<7>
<>

<1>Van Cott, E., Wilson, G.G.
<2>Method for producing the XbaI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 4983542
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the XbaI restriction
endonuclease by (1) introducing the restriction endonuclease gene from Xanthomonas badrii into
a host whereby the restriction gene is expressed; (2) fermenting the host which contains the
vector encoding and expressing the XbaI restriction endonuclease, and (3) purifying the XbaI
restriction endonuclease from the fermented host which contains the vector encoding and
expressing the XbaI restriction endonuclease activity.

<>

<1>Van Cott, E.M.
<2>Method for cloning and producing the SfiI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5637476
<5>
<6>1997
<7>The present invention is directed to a method for cloning and producing the SfiI restriction
endonuclease by 1) introducing the restriction endonuclease gene from S. fimbriatus into a
host whereby the restriction gene is expressed; 2) fermenting the host which contains the
plasmid encoding and expressing the SfiI restriction endonuclease activity; and 3) purifying
the SfiI restriction endonuclease from the fermented host which contains the plasmid encoding
and expressing the SfiI restriction endonuclease activity.

<>

<1>Van Cott, E.M., Wilson, G.G.
<2>Method for producing the FnuDI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 4988620
<5>
<6>1991
<7>The present invention is directed to a method for cloning and producing the FnuDI restriction
endonuclease by (1) introducing the restriction endonuclease gene from F. nucleatum D into a
host whereby the restriction gene is expressed; (2) fermenting the host which contains the
vector encoding and expressing the FnuDI restriction endonuclease and (3) purifying the FnuDI
restriction endonuclease from the fermented host which contains the vector encoding and
expressing the FnuDI restriction endonuclease activity.

<>

<1>Van Cott, E.M., Wilson, G.G.
<2>Cloning the FnuDI, NaeI, NcoI and XbaI restriction-modification systems.
<3>Gene
<4>74
<5>55-59
<6>1988
<7>Methyltransferase genes from the FnuDI, NaeI, NcoI, and XbaI
restriction-modification systems have been isolated in Escherichia coli by
shot-gun cloning bacterial DNA fragments into plasmid vectors and selecting for
protectively modified molecules that resist digestion by the corresponding
endonuclease.

<>

<1>van de Guchte, M. et al.
<2>The complete genome sequence of Lactobacillus bulgaricus reveals extensive and ongoing reductive evolution.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>9274-9279
<6>2006
<7>Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) is a representative of the group of
lactic acid-producing bacteria, mainly known for its worldwide application in yogurt
production. The genome sequence of this bacterium has been determined and shows the signs of
ongoing specialization, with a substantial number of pseudogenes and incomplete metabolic
pathways and relatively few regulatory functions. Several unique features of the L. bulgaricus
genome support the hypothesis that the genome is in a phase of rapid evolution. (i)
Exceptionally high numbers of rRNA and tRNA genes with regard to genome size may indicate that
the L. bulgaricus genome has known a recent phase of important size reduction, in agreement
with the observed high frequency of gene inactivation and elimination; (ii) a much higher GC
content at codon position 3 than expected on the basis of the overall GC content suggests that
the composition of the genome is evolving toward a higher GC content; and (iii) the presence
of a 47.5-kbp inverted repeat in the replication termination region, an extremely rare feature
in bacterial genomes, may be interpreted as a transient stage in genome evolution. The results
indicate the adaptation of L. bulgaricus from a plant-associated habitat to the stable protein
and lactose-rich milk environment through the loss of superfluous functions and
protocooperation with Streptococcus thermophilus.

<>

<1>van den Bogert, B., Boekhorst, J., Smid, E.J., Zoetendal, E.G., Kleerebezem, M.
<2>Draft Genome Sequence of Enterococcus sp. Strain HSIEG1, Isolated from the Human  Small Intestine.
<3>Genome Announcements
<4>1
<5>e01013-13
<6>2013
<7>Enterococcus sp. strain HSIEG1 was isolated from the human small intestine. Its draft genome
predicts a broad carbohydrate fermentation capability, which matches
well with the observed physiological characteristics of this strain. This
metabolic flexibility is expected to be of importance for survival and growth in
the small intestinal habitat.

<>

<1>van den Bogert, B., Boekhorst, J., Smid, E.J., Zoetendal, E.G., Kleerebezem, M.
<2>Draft Genome Sequence of Veillonella parvula HSIVP1, Isolated from the Human Small Intestine.
<3>Genome Announcements
<4>1
<5>e00977-13
<6>2013
<7>Veillonella species are frequently encountered commensals in the human small intestine. Here,
we report the draft genome sequence of the first cultured
representative from this ecosystem, Veillonella parvula strain HSIVP1. The genome
is predicted to encode all the necessary enzymes required for the pathway
involved in the conversion of lactate to propanoate.

<>

<1>van den Broek, B., Noom, M.C., Wuike, G.J.L.
<2>DNA-tension dependence of restriction enzyme cleavage reveals details of the reaction pathway.
<3>Biophys. J.
<4>88
<5>184A
<6>2005
<7>Restriction endonucleases cleave recognition sequences on double-stranded DNA with
extraordinary specificity.  This capability arises from conformation changes in enzyme and/or
DNA upon binding to such a recognition site.  Here we present measurements of energy and rate
changes in the raction pathway as a function of tension on the DNA.  The experiments were
performed using single DNA molecules held between beads kept in laser tweezers while recording
the cleavage times after introduction of the restriction enzymes.  The data shows that EcoRV
cleavage rates drop with increasing tension, while BamHI activity does not decrease.  From the
force-rate curves of EcoRV, we are able to obtain values for the DNA bending angle (51o),
induced-fit rate (~100 s-1) and dsDNA hydrolysis (0.28 s-1).  BamHI, however, does not show
any sign of DNA bending.  For both enzymes a reduced association rate of ~5.106 M-1 s-1 is
found, presumably due to absence of intradomain dissociation-reassociation (jumping) between
non-specific sites on stretched DNA.  These measurements thus illustrate the relative
importance of jumping in the searching process.  Moreover, the tension dependence of the
induced-fit reaction reveals some of the underlying energetics of binding a specific site.
Finally, single-molecule detection of DNA cleavage allows for direct measurements of many of
the reaction steps and is generally applicable to other type II restriction enzymes.

<>

<1>van den Broek, B., Noom, M.C., Wuite, G.J.L.
<2>DNA-tension dependence of restriction enzyme activity reveals mechanochemical properties of the reaction pathway.
<3>Nucleic Acids Res.
<4>33
<5>2676-2684
<6>2005
<7>Type II restriction endonucleases protect bacteria against phage infections by cleaving
recognition sites on foreign double-stranded DNA (dsDNA) with extraordinary specificity. This
capability arises primarily from large conformational changes in enzyme and/or DNA upon target
sequence recognition. In order to elucidate the connection between the mechanics and the
chemistry of DNA recognition and cleavage, we used a single-molecule approach to measure rate
changes in the reaction pathway of EcoRV and BamHI as a function of DNA tension. We show that
the induced-fit rate of EcoRV is strongly reduced by such tension. In contrast, BamHI is found
to be insensitive, providing evidence that both substrate binding and hydrolysis are not
influenced by this force. Based on these results, we propose a mechanochemical model of
induced-fit reactions on DNA, allowing determination of induced-fit rates and DNA bend angles.
Finally, for both enzymes a strongly decreased association rate is obtained on stretched DNA,
presumably due to the absence of intradomain dissociation/re-association between non-specific
sites (jumping). The obtained results should apply to many other DNA-associated proteins.

<>

<1>van den Broek, B., Schmidt, C.F., Wuite, G.J.L.
<2>The effect of DNA tension on restriction-enzyme cutting rates studied with optical tweezers.
<3>Biophys. J.
<4>84
<5>141a
<6>2003
<7>Restriction enzymes recognize base pair sequences on DNA with high specificity.  These sites
trigger a conformational change in the enzyme and/or DNA, permitting the cleavage of both
strands.  Here we report single-molecule measurements of the effect of DNA tension on the
cleavage rate of restriction enzymes BamHI, EcoRV,

<>

<1>van den Broek, B., Vanzi, F., Normanno, D., Pavone, F.S., Wuite, G.J.L.
<2>Real-time observation of DNA looping dynamics of Type IIE restriction enzymes NaeI and NarI.
<3>Nucleic Acids Res.
<4>34
<5>167-174
<6>2006
<7>Many restriction enzymes require binding of two copies of a recognition sequence for DNA
cleavage, thereby introducing a loop in the DNA. We investigated looping dynamics of Type IIE
restriction enzymes NaeI and NarI by tracking the Brownian motion of single tethered DNA
molecules. DNA containing two endonuclease recognition sites spaced a few 100 bp apart connect
small polystyrene beads to a glass surface. The position of a bead is tracked through video
microscopy. Protein-mediated looping and unlooping is then observed as a sudden specific
change in Brownian motion of the bead. With this method we are able to directly follow DNA
looping kinetics of single protein-DNA complexes to obtain loop stability and loop formation
times. We show that, in the absence of divalent cations, NaeI induces DNA loops of specific
size. In contrast, under these conditions NarI mainly creates non-specific loops, resulting in
effective DNA compaction for higher enzyme concentrations. Addition of Ca2+ increases the
NaeI-DNA loop lifetime by two orders of magnitude and stimulates specific binding by NarI.
Finally, for both enzymes we observe exponentially distributed loop formation times,
indicating that looping is dominated by (re)binding the second recognition site.

<>

<1>van den Broek, B., Wuite, G.J.L.
<2>Single molecule study of DNA tension on restriction enzyme cleavage activity.
<3>Biophys. J.
<4>82
<5>51a
<6>2002
<7>Restriction enzymes recognizes dsDNA at highly specific base pair sequences.  These sites
trigger a large conformational change in the enzyme, permitting the cleavage of DNA.  During
this conformational change the DNA is often bent, although some enzymes do not deform the DNA
at all.  While the biochemistry and structure of many restriction enzymes are well studied,
these studies only superficially address the dynamics of enzyme-DNA interactions.  Here we
report single molecule measurements of the effect of DNA tension on the cleavage activity of
restriction enzymes in order to elucidate some of the enzyme dynamics.  DNA is held under
various tensions between two beads in optical tweezers while the time it takes for the enzymes
to cut is recorded; this allows us to obtain force vs. rate curves.  We present our first
results for the restriction enzyme NaeI, which kinks the DNA by 45 degrees before cutting it.
We expect to see several trends: DNA tension reduces the binding energy of the sugar backbone,
suggesting an increase in cutting rate.  At the same time, this tension opposes the DNA
bending by the enzyme.  Moreover, pulling on the DNA slightly deforms the base stacking,
degrading the recognition efficiency.  These effects should slow down the cutting.

<>

<1>van den Hondel, C.A.M.J.J., van Leen, R.W., van Arkel, G.A., Duyvesteyn, M., de Waard, A.
<2>Sequence-specific nucleases from the cyanobacterium Fremyella diplosiphon, and a peculiar resistance of its chromosomal DNA towards cleavage by other restriction enzymes.
<3>FEMS Microbiol. Lett.
<4>16
<5>7-12
<6>1983
<7>The filamentous cyanobacterium Fremyella diplosiphon PCC7601 is facultatively
heterotrophic.  Cells can grow in the dark at the expense of certain organic
substrates like sugars as an exogenous energy source, the utilization of which
probably depends on the presence in the cell of the appropriate permeases.  As
a first step in cloning the F. diplosiphon genetic information for these
permeases into the transformable strain R-2 of the unicellular, strictly
photoautotrophic cyanobacterium Anacystis nidulans, chromosomal DNA of F.
diplosiphon was digested with several bacterial sequence-specific
endodeoxyribonucleases.  As the DNA appeared to be resistant to an unexpectedly
large number of them, a restriction enzyme analysis with 31 different
endonucleases was made to elucidate the underlying modification pattern.  In
addition, F. diplosiphon cells were anlysed for the presence of restriction
endonucleases, and two were found:  FdiI and FdiII, which cleave the nucleotide
sequences 5'-G^G(A/T)CC and 5'-TGC^GCA, respectively.  EndoR.FdiI is an
isoschizomer of AvaII and endoR.FdiII is an isoschizomer of AosI.

<>

<1>Van den Wyngaert, I., Sprengel, J., Kass, S.U., Luyten, W.H.M.L.
<2>Cloning and analysis of a novel human putative DNA methyltransferase.
<3>FEBS Lett.
<4>426
<5>283-289
<6>1998
<7>DNA methylation is intricately involved in a variety of cellular processes, such as
differentiation, cell cycle progression, X-chromosome inactivation and genomic imprinting.
However, little is known about how specific DNA methylation patterns are established and
maintained.  Previously one mammalian DNA methyltransferase has been described, but there has
been considerable speculation about the presence of a second activity capable of methylation.
Here we report the identification and characterization of a novel human putative DNA
methyltransferase.  Using a bioinformatics screen we have identified several expressed
sequence tags which show high sequence similarity to the Schizosaccharomyces pombe gene pmt1+.
The cDNA for PuMet was cloned and the predicted amino acid sequence deduced.  The gene is
ubiquitously expressed, albeit at low levels.  Like several other DNA methyltransferases, the
bacterially overexpressed protein is not active in methylation assays.

<>

<1>Van der Auwera, G.A., Feldgarden, M., Kolter, R., Mahillon, J.
<2>Whole-Genome Sequences of 94 Environmental Isolates of Bacillus cereus Sensu Lato.
<3>Genome Announcements
<4>1
<5>e00380-13
<6>2013
<7>Bacillus cereus sensu lato is a species complex that includes the anthrax pathogen Bacillus
anthracis and other bacterial species of medical, industrial,
and ecological importance. Their phenotypes of interest are typically linked to
large plasmids that are closely related to the anthrax plasmids pXO1 and pXO2.
Here, we present the draft genome sequences of 94 isolates of B. cereus sensu
lato, which were chosen for their plasmid content and environmental origins.

<>

<1>van der Graaf-van, B.L., Miller, W.G., Yee, E., Bono, J.L., Rijnsburger, M., Campero, C., Wagenaar, J.A., Duim, B.
<2>First Closed Genome Sequence of Campylobacter fetus subsp. venerealis bv. intermedius.
<3>Genome Announcements
<4>2
<5>e01246-13
<6>2014
<7>Campylobacter fetus subsp. venerealis bv. intermedius is a variant of C. fetus subsp.
venerealis, the causative agent of bovine genital campylobacteriosis, a
venereal disease associated with abortion and infertility in cattle. We report
the first closed whole-genome sequence of this biovar.

<>

<1>van der Gun, B.T.F., Maluszynska-Hoffman, M., Kiss, A., Arendzen, A.J., Ruiters, M.H.J., McLaughlin, P.M.J., Weinhold, E., Rots, M.G.
<2>Targeted DNA Methylation by a DNA Methyltransferase Coupled to a Triple Helix Forming Oligonucleotide To Down-Regulate the Epithelial Cell Adhesion Molecule.
<3>Bioconjugate Chem.
<4>21
<5>1239-1245
<6>2010
<7>The epithelial cell adhesion molecule (EpCAM) is a membrane glycoprotein that has been
identified as a marker of cancer-initiating
cells. EpCAM is highly expressed on most carcinomas, and transient
silencing of EpCAM expression leads to reduced oncogenic potential. To
silence (he EpCAM gene in a persistent manner via targeted DNA
methylation, a low activity mutant (C141S) of the CpG-specific DNA
methyltransferase M.Sssl was coupled to a triple-helix-forming
oligonucleotide (TFO-C141S) specifically designed for the EpCAM gene.
Reporter plasmids encoding the green fluorescent protein under control
of different EpCAM promoter fragments were treated with the TFO-C141S
conjugate to determine the specificity of targeted DNA methylation in
the context of a functional EpCAM promoter. Treatment of the plasmids
with TFO-C141S resulted in efficient and specific methylation of the
targeted CpG located directly downstream of the triple helix forming
site (TFS). No background DNA methylation was observed neither in a 700
bp region of the EpCAM promoter nor in a 400 bp region of the reporter
gene downstream of the TFS. Methylation of the target CpG did not have
a detectable effect on promoter activity. This study shows that the
combination of a specific TFO and a reduced activity methyltransferase
variant can be used to target DNA methylation to predetermined sites
with high specificity, allowing determination of crucial CpGs for
promoter activity.

<>

<1>van der Mee-Marquet, N., Hernandez, D., Bertrand, X., Quentin, R., Corvaglia, A.R., Francois, P.
<2>Whole-Genome Sequence of the Ancestral Animal-Borne ST398 Staphylococcus aureus Strain S123.
<3>Genome Announcements
<4>1
<5>e00692-13
<6>2013
<7>Sequence type 398 (ST398) Staphylococcus aureus was originally reported in livestock. We
announce the complete genome sequence of an ST398
methicillin-susceptible S. aureus strain of animal origin, S123. The genome
sequence reveals a wild-type genome that probably corresponds to an ancestral
clone.

<>

<1>van der Ploeg, L.H.T., Flavell, R.A.
<2>DNA methylation in the human cdb-globin locus in Erythroid and Nonerythroid tissues.
<3>Cell
<4>19
<5>947-958
<6>1980
<7>We have analyzed DNA modification in the human cdb-globin gene region at 17
cleavage sites of restriction endonucleases which are unable to cleave DNA if
5-methylcytosine is present at certain positions in their respective cleavage
sites.  Using this criterion, all sites tested in the globin gene region are
fully modified in the germ line (sperm) DNA.  In somatic tissues, however,
methyl groups are absent at specific sites in the globin gene region.  In
tissues not expressing the genes, these losses range from one of these cleavage
sites in lymphocyte DNA to essentially all of these sites in the entire region
in placental DNA.  In the DNA of tissues expressing the globin genes, the
region surrounding and including the genes expressed shows a low level of
modification, whereas the neighboring DNA regions have a high level of
modification.  The data suggest that a low level of DNA methylation may be a
necessary, but not a sufficient, condition for gene expression in higher
eucaryotes.

<>

<1>van der Woerd, M.J., Pelletier, J.J., Xu, S.-Y., Friedman, A.M.
<2>Restriction enzyme BsoBI-DNA complex: A tunnel for recognition of degenerate DNA sequences and potential histidine catalysis.
<3>Structure
<4>9
<5>133-144
<6>2001
<7>Background: Restriction endonucleases form a diverse family of proteins with substantial
variation in sequence, structure, and interaction with recognition site DNA. BsoBI is a
thermophilic restriction endonuclease that exhibits both base-specific and degenerate
recognition within the sequence CPyCGPuG.  Results: The structure of BsoBI complexed to
cognate DNA has been determined to 1.7 Angstrom resolution, revealing several unprecedented
features. Each BsoBI monomer is formed by inserting a helical domain into an expanded
EcoRI-type catalytic domain. DNA is completely encircled by a BsoBI dimer. Recognition
sequence DNA lies within a 20 Angstrom long tunnel of protein that excludes bulk solvent.
Interactions with the specific bases are made in both grooves through direct and
water-mediated hydrogen bonding. Interaction with the degenerate position is mediated by a
purine-specific hydrogen bond to N7, ensuring specificity, and water-mediated H bonding to the
purine N6/O6 and pyrimidine N4/O4, allowing degeneracy. In addition to the conserved active
site residues of the DXn(E/D)ZK restriction enzyme motif, His253 is positioned to act as a
general base.  Conclusions: A catalytic mechanism employing His253 and two metal ions is
proposed. If confirmed, this would be the first example of histidine-mediated catalysis in a
restriction endonuclease. The structure also provides two novel examples of the role of water
in protein-DNA interaction. Degenerate recognition may be mediated by employing water as a
hydrogen bond donor or acceptor. The structure of DNA in the tunnel may also be influenced by
the absence of bulk solvent.

<>

<1>van der Woude, M., Hale, W.B., Low, D.A.
<2>Formation of DNA methylation patterns: Nonmethylated GATC sequences in gut and pap operons.
<3>J. Bacteriol.
<4>180
<5>5913-5920
<6>1998
<7>Most of the adenine residues in GATC sequences in the Escherichia coli chromosome are
methylated by the enzyme deoxyadenosine methyltransferase.  However, at least 20 GATC
sequences remain nonmethylated throughout the cell cycle.  Here we examined how the DNA
methylation patterns of GATC sequences within the regulatory regions of the
pyelonephritis-associated pilus operon and the glucitol utilization operon were formed.  The
results obtained with an in vitro methylation protection assay showed that the addition of the
leucine-responsive regulatory protein to pap DNA was sufficient to protect the two GATC
sequences in the pap regulatory region, GATC-I and GATC-II, from methylation by Dam.  This
finding was consistent with previously published data showing that Lrp was essential for
methylation protection of these DNA sites in vivo.  Methylation protection also occurred at a
GATC site (GATC-44.5) centered 44.5 bp upstream of the transcription start site of the gutABD
operon.  Two proteins, GutR and the catabolite gene activator protein, bound to DNA sites
overlapping the GATC-44.5-containing region of the gutABD operon.  GutR, an operon-specific
repressor, was essential for methylation protection in vivo, and binding of GutR protected
GATC-44.5 from methylation in vitro.  In contrast, binding of CAP at a site overlapping
GATC-44.5 did not protect this site from methylation.  Mutational anlayses indicated that
gutABD gene regulation was not controlled by methylation of GATC-44.5, in contrast to
regulation of Pap pilus expression, which is directly controlled by methylation of the pap
GATC-I and GATC-II sites.

<>

<1>van der Woude, M.W.
<2>Some types of bacterial phase variation are epigenetic.
<3>Microbe
<4>3
<5>21-26
<6>2008
<7>Individual bacterial cells can vary phenotypically, even within a clonal population.  Phase
variation provides a means for regulating specific genes between an expressed, or "on" state,
and a nonexpressed, or "off" state; these two expression states are heritable and reversible.
The mechanisms that lead to phase variation are either genetic or epigenetic; if epigenetic,
they entail no changes in DNA sequence.  More generally, phase variation apparently provides a
means for generating diversity and thereby increasing the changes of survival amid changing
environmental conditions.  Understanding phase variation may allow us to manipulate bacterial
populations to provide health benefits, including better diagnostic tests and vaccines.

<>

<1>van der Woude, M.W., Baumler, A.J.
<2>Phase and antigenic variation in bacteria.
<3>Clin. Microbiol. Rev.
<4>17
<5>581-611
<6>2004
<7>Phase and antigenic variation result in a heterogenic phenotype of a clonal bacterial
population, in which individual cells either express the phase-variable protein(s) or not, or
express one of multiple antigenic forms of the protein, respectively. This form of regulation
has been identified mainly, but by no means exclusively, for a wide variety of surface
structures in animal pathogens and is implicated as a virulence strategy. This review provides
an overview of the many bacterial proteins and structures that are under the control of phase
or antigenic variation. The context is mainly within the role of the proteins and variation
for pathogenesis, which reflects the main body of literature. The occurrence of phase
variation in expression of genes not readily recognizable as virulence factors is highlighted
as well, to illustrate that our current knowledge is incomplete. From recent genome sequence
analysis, it has become clear that phase variation may be more widespread than is currently
recognized, and a brief discussion is included to show how genome sequence analysis can
provide novel information, as well as its limitations. The current state of knowledge of the
molecular mechanisms leading to phase variation and antigenic variation are reviewed, and the
way in which these mechanisms form part of the general regulatory network of the cell is
addressed. Arguments both for and against a role of phase and antigenic variation in immune
evasion are presented and put into new perspective by distinguishing between a role in
bacterial persistence in a host and a role in facilitating evasion of cross-immunity. Finally,
examples are presented to illustrate that phase-variable gene expression should be taken into
account in the development of diagnostic assays and in the interpretation of experimental
results and epidemiological studies.

<>

<1>van der Woude, M.W., Braaten, B.A., Low, D.A.
<2>Evidence for global regulatory control of pilus expression in Escherichia coli by Lrp and DNA methylation: model building based on analysis of pap.
<3>Mol. Microbiol.
<4>6
<5>2429-2435
<6>1992
<7>Pyelonephritis-associated pilus (Pap) expression is regulated by a phase variation control
mechanism involving PapB, Papl, catabolite activator protein (CAP), leucine-responsive
regulatory protein (Lrp) and deoxyadenosine methylase (Dam). Lrp and Papl bind to a specific
non-methylated pap regulatory DNA region containing the sequence 'GATC' and facilitate the
formation of an active transcriptional complex.  Evidence indicates that binding of Lrp and
Papl to this region inhibits methylation of the GATC site by Dam.  However, if this GATC site
is first methylated by Dam, binding of Lrp and Papl is inhibited. These events lead to the
formation of two different pap methylation states characteristic of active (ON) and inactive
(OFF) pap transcription states. The fae (K88), daa (F1845) and sfa (S) pilus operons share
conserved 'GATC-box' domains with pap and may be subject to a similar regulatory control
mechanism involving Lrp and DNA methylation.

<>

<1>Van Dongen, W.M.A.M., Van Vlerken, M.M.A., De Graaf, F.K.
<2>Nucleotide sequence of a DNA fragment encoding a Vibrio cholerae haemagglutinin.
<3>Genetics (Life Sci. Adv.)
<4>6
<5>85-91
<6>1987
<7>
<>

<1>van Doorn, L.J., Wim, Q.
<2>Detection and typing of the iceA gene of Helicobacter pylori.
<3>Japanese Patent Office
<4>JP 2001521766 A
<5>
<6>2001
<7>
<>

<1>Van Emburgh, B.O., Robertson, K.D.
<2>DNA methyltransferases and methyl-CPG binding proteins as multifunctional regulators of chromatin structure and development in mammalian cells.
<3>EPIGENETICS, Caister Academic Press, Tost, J., 
<4>0
<5>23-61
<6>2008
<7>The epigenetic modification of DNA with 5-methylcytosine is an important regulatory event
involved in chromatin structure, genomic imprinting, inactivation of the X chromosome,
transcription, and retrotransposon silencing. This modification is catalysed and maintained by
the DNA methyltransferases and is interpreted by the methyl-CpG binding proteins. DNA
methyltransferases are not limited to catalysing DNA methylation, but also take part in the
regulation of gene expression through interactions with other proteins that repress
transcription and modify chromatin structure. The use of mouse models, as well as human
diseases resulting from deficiencies in the rnethylation machinery, have been integral parts
of understanding the role of these proteins in development and cellular homeostasis. More and
more studies are reporting additional roles within the cell beyond their DNA methyltransferase
and methyl-CpG binding properties. There is at this point, though, only limited understanding
of how these enzymes and proteins are targeted to specific genomic regions. The
methyltransferases that will be discussed in this review include DNMT1, DNMT2, and the DNMT3
family of enzymes as well as the methyl-CpG binding proteins MeCP2, MBD1, MBD2, MBD3, and
MBD4. The function of these enzymes, as well as their interactions with other cellular
proteins and each other, will be discussed along with the diseases attributed to aberrations
in the DNA methylation machinery.

<>

<1>Van Emburgh, B.O., Robertson, K.D.
<2>Modulation of Dnmt3b function in vitro by interactions with Dnmt3L, Dnmt3a and Dnmt3b splice variants.
<3>Nucleic Acids Res.
<4>39
<5>4984-5002
<6>2011
<7>DNA methylation, an essential regulator of transcription and chromatin structure, is
established and maintained by the coordinated action of
three DNA methyltransferases: DNMT1, DNMT3A and DNMT3B, and the inactive
accessory factor DNMT3L. Disruptions in DNMT3B function are linked to
carcinogenesis and genetic disease. DNMT3B is also highly alternatively
spliced in a tissue- and disease-specific manner. The impact of
intra-DNMT3 interactions and alternative splicing on the function of DNMT3
family members remains unclear. In the present work, we focused on DNMT3B.
Using a panel of in vitro assays, we examined the consequences of DNMT3B
splicing and mutations on its ability to bind DNA, interact with itself
and other DNMT3's, and methylate DNA. Our results show that, while the
C-terminal catalytic domain is critical for most DNMT3B functions, parts
of the N-terminal region, including the PWWP domain, are also important.
Alternative splicing and domain deletions also influence DNMT3B's cellular
localization. Furthermore, our data reveal the existence of extensive
DNMT3B self-interactions that differentially impact on its activity.
Finally, we show that catalytically inactive isoforms of DNMT3B are
capable of modulating the activity of DNMT3A-DNMT3L complexes. Our studies
therefore suggest that seemingly 'inactive' DNMT3B isoforms may influence
genomic methylation patterns in vivo.

<>

<1>Van Etten, J.L.
<2>DNA methyltransferases and DNA site-specific endonucleases encoded by Chlorella viruses.
<3>J. Phycol.
<4>27
<5>74
<6>1991
<7>Thirty seven large, polyhedral, dsDNA-containing (>300 kb in size),
plaque-forming viruses which infect a unicellular, eukaryotic, green alga,
Chlorella strain NC64A have been partially characterized.  Each of the viral
DNAs contains 5mC; the concentration of 5mC as a percentage of cytosine, ranges
from 0.1 to 47%.  In addition, 25 of the 37 viral DNAs also contain 6mA; the
concentration of 6mA, as a percentage of adenine, ranges from 1.45% to 37%.
The finding that methylation is sequence specific led to the discovery that at
least some of the viruses code for DNA methyltransferases and DNA site-specific
(restriction) endonucleases.  Some of the site-specific endonucleases recognize
and cleave the same base sequences as bacterial enzymes whereas, others have
unique specificities and cleavage sites.  Two additional site-specific
endonucleases are unusual because they cleave one strand of DNA at specific
sequences.  The presence of DNA methylating and site-specific endonucleases in
the virus infected cells leads to two questions. (i) What is the evolutionary
origin of these enzymes? (ii) What is the function of these enzymes.
Experiments designed to address these questions will be presented.

<>

<1>Van Etten, J.L., Meints, R.H.
<2>Giant viruses infecting algae.
<3>Annu. Rev. Microbiol.
<4>53
<5>447-494
<6>1999
<7>Paramecium bursaria chlorella virus (PBCV-1) is the prototype of a family of large,
icosahedral, plaque-forming, double-stranded-DNA-containing viruses that replicate in certain
unicellular, eukaryotic chlorella-like green algae. DNA sequence analysis of its 330, 742-bp
genome leads to the prediction that this phycodnavirus has 376 protein-encoding genes and 10
transfer RNA genes. The predicted gene products of approximately 40% of these genes resemble
proteins of known function. The chlorella viruses have other features that distinguish them
from most viruses, in addition to their large genome size. These features include the
following: (a) The viruses encode multiple DNA methyltransferases and DNA site-specific
endonucleases; (b) PBCV-1 encodes at least part, if not the entire machinery to glycosylate
its proteins; (c) PBCV-1 has at least two types of introns--a self-splicing intron in a
transcription factor-like gene and a splicesomal processed type of intron in its DNA
polymerase gene. Unlike the chlorella viruses, large double-stranded-DNA-containing viruses
that infect marine, filamentous brown algae have a circular genome and a lysogenic phase in
their life cycle.

<>

<1>Van Etten, J.L., Schuster, A.M., Girton, L., Burbank, D.E., Swinton, D., Hattman, S.
<2>DNA methylation of viruses infecting a eukaryotic Chlorella-like green alga.
<3>Nucleic Acids Res.
<4>13
<5>3471-3478
<6>1985
<7>The genomic DNAs of the eukaryotic Chlorella-like green alga, strain NC64A, and
eleven of its viruses all contain significant levels of 5-methyldeoxycytidine.
In addition, the host DNA as well as six of the viral DNAs also contain
N6-methyldeoxyadenosine.  At least some of the methylated bases in the host
reside in different base sequences than the methylated bases in the viruses as
shown by differential susceptibility to restriction endonuclease enzymes.  This
suggests that the viruses encode for DNA methyltransferases with sequence
specificities different from that of the host enzyme.

<>

<1>Van Etten, J.L., Xia, Y., Burbank, D.E.
<2>Viruses infecting a eukaryotic green alga are a new source of DNA methyltransferases and DNA site-specific endonucleases.
<3>J. Cell Biochem. Suppl.
<4>13D
<5>199
<6>1989
<7>We have isolated and partially characterized several large, complex, dsDNA
containing (ca. 330kbp), plaque forming viruses (NC64A viruses) which infect
the unicellular, eukaryotic, Chlorella-like green alga strain NC64A.  These
viruses can be distinguished on the basis of plaque size, reaction with
antibody, and the nature and abundance of methylated bases in their genomic
DNAs.  The concentration of methylated bases varies from viruses containing no
6-methyladenine (6mA) and 0.1% 5-methylcytosine (5mC) to one virus with 37% 6mA
and 45% 5mC.  Even though the host nuclear DNA contains 21% 5mC and 0.6% 6mA,
at least some of the methylated bases in the viruses reside in different DNA
sequences than those in the host.  This result led to the finding that virus
infection of the host results in the synthesis of DNA methyltransferases and
DNA site-specific (restriction) endonucleases.  Five different site-specific
endonucleases have been identified so far.  For example, virus NC-1A infected
Chlorella NC64A cells contain an endonuclease, named CviBI, which recognizes
the sequence GANTC and cleaves between G and A; CviBI does not cleave GmANTC
sequences.  NC-1A infected Chlorella cells contain the cognate
methyltransferase, M.CviBI, which specifically methylates adenine in GANTC
sequences and at least two other distinct methyltransferases; M.CviBII and
M.CviBIII methylate adenine in GATC and TCGA sequences, respectively.  The
M.CviBIII gene was cloned, sequenced and a single open reading frame of 1131 bp
identified.  A comparison of the predicted amino acid sequence of M.CviBIII
with M.TaqI, a bacterial isoschizomer, revealed 39% identity.  A second family
of viruses that infect another strain of Chlorella (Pbi) have recently been
isolated.  Like the NC64A viruses, the Pbi viruses contain large dsDNAs genomes
with various levels of methylated bases.  Infection of Chlorella Pbi with the
Pbi viruses also results in the synthesis of DNA methyltransferases and DNA
site-specific endonucleases.  Thus these Chlorella viruses are a new source of
both adenine and cytosine methyltransferases and DNA site-specific
endonucleases.

<>

<1>Van Etten, J.L., Xia, Y., Burbank, D.E., Narva, K.E.
<2>Chlorella viruses code for restriction and modification enzymes.
<3>Gene
<4>74
<5>113-115
<6>1988
<7>Meeting Abstract

<>

<1>Van Heuverswyn, H., Fiers, W.
<2>Recognition sequence for the restriction endonuclease BglI from Bacillus globigii.
<3>Gene
<4>9
<5>195-203
<6>1980
<7>Restriction endonuclease BglI recognizes the DNA sequence ...CCCNNNN^NGGC...3'
...CGGN^NNNNCCG...5' and cleaves each strand at the site indicated, thus
generating 3' protruding ends.  The recognition sequence was deduced by
correlating mapping data with nucleotide sequence information and the position
of cleavage was unambiguously determined by 32P labeling of 5' termini produced
by BglI digestion.

<>

<1>van Hijum, S.A., Szalowska, E., van der Maarel, M.J., Dijkhuizen, L.
<2>Biochemical and molecular characterization of a levansucrase from Lactobacillus reuteri.
<3>Microbiology
<4>150
<5>621-630
<6>2004
<7>Lactobacillus reuteri strain 121 employs a fructosyltransferase (FTF) to synthesize a fructose
polymer [a fructan of the levan type, with
beta(2-->6) linkages] from sucrose or raffinose. Purification of this FTF
(a levansucrase), and identification of peptide amino acid sequences,
allowed isolation of the first Lactobacillus levansucrase gene (lev),
encoding a protein (Lev) consisting of 804 amino acids. Lev showed highest
similarity with an inulosucrase of L. reuteri 121 [Inu; producing an
inulin polymer with beta(2-->1)-linked fructosyl units] and with FTFs from
streptococci. Expression of lev in Escherichia coli resulted in an active
FTF (Lev Delta 773His) that produced the same levan polymer [with only 2-3
% beta(2-->1-->6) branching points] as L. reuteri 121 cells grown on
raffinose. The low degree of branching of the L. reuteri levan is very
different from bacterial levans known up to now, such as that of
Streptococcus salivarius, having up to 30 % branches. Although Lev is
unusual in showing a higher hydrolysis than transferase activity,
significant amounts of levan polymer are produced both in vivo and in
vitro. Lev is strongly dependent on Ca(2+) ions for activity. Unique
properties of L. reuteri Lev together with Inu are: (i) the presence of a
C-terminal cell-wall-anchoring motif causing similar expression problems
in Escherichia coli, (ii) a relatively high optimum temperature for
activity for FTF enzymes, and (iii) at 50 degrees C, kinetics that are
best described by the Hill equation.

<>

<1>Van Hijum, S.A.F.T., Van Geel-Schutten, G.H., Dijkhuizen, L., Rahaoui, H.
<2>Fructosyltransferases.
<3>US Patent Office
<4>US 6730502 A
<5>
<6>2004
<7>The present invention describes two novel proteins having fructosyltransferase activity.  Both
enzymes are derived from lactobacilli, which are food-grade micro-organisms with the Generally
Recognized As Safe (GRAS) status.  One of these proteins produces an inulin and
fructo-oligosaccharides, while the other produces a levan and fructo-oligosaccharides.
According to the invention lactobacilli capable of producing an inulin and/or a levan and/or
fructo-oligosaccharides using one or both of the fructosyltransferases can be used as a
probiotic or a symbiotic.

<>

<1>Van Houdt, R., Van der Lelie, D., Izquierdo, J.A., Aertsen, A., Masschelein, J., Lavigne, R., Michiels, C.W., Taghavi, S.
<2>Genome Sequence of Serratia plymuthica RVH1, Isolated from a Raw Vegetable-Processing Line.
<3>Genome Announcements
<4>2
<5>e00021-14
<6>2014
<7>We announce the genome sequence of Serratia plymuthica strain RVH1, a psychroloterant strain
that was isolated from a raw vegetable-processing line and
that regulates the production of primary metabolites (acetoin and butanediol),
antibiotics, and extracellular enzymes through quorum sensing.

<>

<1>van Kranenburg, R., de Vos, W.M.
<2>Characterization of multiple regions involved in replication and mobilization of plasmid pNZ4000 coding for exopolysaccharide production in   Lactococcus lactis.
<3>J. Bacteriol.
<4>180
<5>5285-5290
<6>1998
<7>We characterized the regions involved in replication and mobilization of the 40-kb plasmid
pNZ4000, encoding exopolysaccharide (EPS) production in
Lactococcus lactis NIZO B40. The plasmid contains four highly conserved
replication regions with homologous rep genes (repB1, repB2, repB3, and
repB4) that belong to the lactococcal theta replicon family. Subcloning of
each replicon individually showed that all are functional and compatible
in L. lactis. Plasmid pNZ4000 and genetically labeled derivatives could be
transferred to different L. lactis strains by conjugation, and pNZ4000 was
shown to be a mobilization plasmid. Two regions involved in mobilization
were identified near two of the replicons; both included an oriT sequence
rich in inverted repeats. Conjugative mobilization of the nonmobilizable
plasmid pNZ124 was promoted by either one of these oriT sequences,
demonstrating their functionality. One oriT sequence was followed by a
mobA gene, coding for a trans-acting protein, which increased the
frequency of conjugative transfer 100-fold. The predicted MobA protein and
the oriT sequences show protein and nucleotide similarity, respectively,
with the relaxase and with the inverted repeat and nic site of the oriT
from the Escherichia coli plasmid R64. The presence on pNZ4000 of four
functional replicons, two oriT sequences, and several insertion
sequence-like elements strongly suggests that this EPS plasmid is a
naturally occurring cointegrate.

<>

<1>van Kranenburg, R., Kleerebezem, M., de Vos, W.M.
<2>Nucleotide sequence analysis of the lactococcal EPS plasmid pNZ4000.
<3>Plasmid
<4>43
<5>130-136
<6>2000
<7>The complete 42180-bp nucleotide sequence of the mobilization plasmid
pNZ4000, coding for exopolysaccharide (EPS) production in Lactococcus
lactis, was determined. This plasmid contains a region involved in EPS
biosynthesis, four functional replicons, a region containing mobilization
genes, and three origin of transfer (oriT) sequences. Sequences identical
to these oriT sequences were also found on two other lactococcal plasmids
and a plasmid from Lactobacillus helveticus. Several complete and partial
IS elements were identified on pNZ4000, including iso-ISS1, iso-IS946, and
iso-IS982 sequences. Furthermore, pNZ4000 contains a gene cluster that may
encode a cobalt transport system and a gene that encodes a CorA homologue
which may function as a magnesium transporter.

<>

<1>Van Laar, T.A., Chen, T., Childers, B.M., Chen, P., Abercrombie, J.J., Leung, K.P.
<2>Genome Sequence of a Multidrug-Resistant Strain of Klebsiella pneumoniae, BAMC 07-18, Isolated from a Combat Injury Wound.
<3>Genome Announcements
<4>2
<5>e01230-14
<6>2014
<7>Klebsiella pneumoniae is an important infectious agent of surgical sites and combat wounds.
Antibiotic resistance and tolerance are common impediments to the
healing of chronic infections. Here, we report the genome sequence of a highly
multidrug-resistant strain of K. pneumoniae, BAMC 07-18, isolated from a combat
wound of a soldier.

<>

<1>van Luijk, N., Stierli, M.P., Schwenninger, S.M., Herve, C., Dasen, G., Jore, J.P.M., Pouwels, P.H., van der Werf, M.J., Teuber, M., Meile, L.
<2>Genetics and molecular biology of propionibacteria.
<3>Lait
<4>82
<5>45-57
<6>2002
<7>Research in the genetics and molecular biology of propionibacteria is currently making much
progress. In order to develop efficient DNA
transfer systems for the genus Propionibacterium, dairy and
environmental propionibacteria were screened for the presence of
suitable plasmids as a first step. Following nucleotide sequence
analysis, potential replication functions were identified on several
Propionibacterium plasmids such as pLME 106/pRGO1, p545 and pLME108.
Furthermore, ppnA, the gene encoding the propionicin SM1, was detected
on pLME106. Three of these plasmids which had been fused with
antibiotic resistance selection markers (ermE, cml, hygB) originating
from bacteria with high G+C DNA content were recently successfully used
as Escherichia coli - Propionibacterium shuttle vectors. DNA
restriction/modification systems observed in propionibacteria have to
be taken into account since successful DNA transformation at high rates
(up to 10^8 Propionibacterium transformants/microgram of DNA) succeeds only
with plasmid DNA originating from propionibacteria with the same
restriction/modification system(s) as the strain to be transformed, and
not from E. coli hosts. The basis for an integrating vector has been
set up after identification of a potential attP site and an adjacent
integrase gene from a Propionibacterium phage/prophage system. Finally,
approximately 30 gene sequences with attributed coding functions from
propionibacteria are available on databases.

<>

<1>Van Ness, J.
<2>New LucNic and MetNic restriction endonucleases, useful in performing Strand Displacement Amplification or in recognizing relatively  non-complex substrate DNA sequence - recombinant enzyme protein  production for use in displacement amplification.
<3>International Patent Office
<4>WO 2003104417
<5>
<6>2003
<7>DERWENT ABSTRACT: NOVELTY - An isolated polynucleotide comprising a nucleotide sequence of
1875 or 1062 bp (SEQ ID NO: 1 and 8), encoding a
LucNic or MetNic restriction endonuclease polypeptide, respectively,
comprising a sequence of 624 or 353 amino acids (SEQ ID NO: 2 and 7) or
an amino acid sequence having at least 80% sequence identity to SEQ ID
NO: 2 or 7, is new. DETAILED DESCRIPTION - INDEPENDENT CLAIMS are also
included for: (1) a vector comprising the polynucleotide described
above; (2) a host cell transformed by the vector of (1); (3) producing
a LucNic or MetNic restriction endonuclease; and (4) performing Strand
Displacement Amplification (SDA) comprises using a LucNic or MetNic
restriction endonuclease polypeptide to nick template DNA, where the
polypeptide is capable of nicking template DNA. BIOTECHNOLOGY -
Preparation (claimed): Producing a LucNic or MetNic restriction
endonuclease comprises culturing the transformed host cell of (2) under
conditions and a time sufficient to express the LucNic or MetNic
restriction endonuclease polypeptide. USE - The LucNic or MetNic
restriction endonuclease are useful in performing SDA. The polypeptide
specifically recognizes relatively non-complex substrate DNA sequence
and does not require assembly into multi-subunit complexes to manifest
DNA nicking activity. EXAMPLE - Bacillus halodurans cells were obtained
and cultured supplemented with 100mM NaCl. Homogenate from the cultured
cells was obtained through centrifugation. The supernatant was then
loaded onto a fast flow Phosphocellulose column. The LucNic
endonuclease was then eluted from the column using buffer of increasing
salt concentration. Fractions were collected and assayed for LucNic
restriction activity. Active fractions were pooled and dialyzed against
100mM NaCl supplemented Buffer B. The dialyzed pool was diluted with
Buffer B to a final concentration of 50 NaCl. The precipitate formed
was spun out by centrifugation and clear supernatant was carried
forward. The active dialyzed solution was applied to a 15Q column,
washed and fractions were again collected and assayed for activity. The
washed column was also checked for activity. Active fractions were
pooled and diluted and loaded onto heparin column. The column was
washed and a salt gradient was run down into the column. Fractions were
collected and assayed for LucNic activity. Active fractions were then
pooled and diluted. Active fractions were then loaded onto the 15Q
column, washed and a salt gradient was then applied. Fractions
including flow through and wash were collected and assayed for the
activity. Active fractions were pooled and diluted. The most active
fraction was loaded onto an SDS-PAGE protein gel and subjected to
electrophoresis. The gel was stained and a prominent band with a 624
amino acid sequence was observed. (49 pages)

<>

<1>van Noort, J., van der Heijden, T., Dutta, F.C., Firman, K., Dekker, C.
<2>Initiation of translocation by Type I restriction-modification enzymes is associated with a short DNA extrusion.
<3>Nucleic Acids Res.
<4>32
<5>6540-6547
<6>2004
<7>Recognition of 'foreign' DNA by Type I restriction-modification (R-M) enzymes elicits an
ATP-dependent switch from methylase to endonuclease activity, which involves DNA translocation
by the restriction subunit HsdR. Type I R-M enzymes are composed of three (Hsd) subunits with
a stoichiometry of HsdR2:HsdM2:HsdS1 (R2-complex). However, the EcoR124I R-M enzyme can also
exist as a cleavage deficient, sub-assembly of HsdR1:HsdM2:HsdS1 (R1-complex). ATPS was used
to trap initial translocation complexes, which were visualized by Atomic Force Microscopy
(AFM). In the R1-complex, a small bulge, associated with a shortening in the contour-length of
the DNA of 8 nm, was observed. This bulge was found to be sensitive to single-strand DNA
nucleases, indicative of non-duplexed DNA. R2-complexes appeared larger in the AFM images and
the DNA contour length showed a shortening of approximately 11 nm, suggesting that two bulges
were formed. Disclosure of the structure of the first stage after the
recognition-translocation switch of Type I restriction enzymes forms an important first step
in resolving a detailed mechanistic picture of DNA translocation by SF-II DNA translocation
motors.

<>

<1>van Noort, J., van der Heijden, T., van der Scheer, C., Firman, K., Dekker, C.
<2>Single molecule characterization of DNA translocation by the type I restriction enzyme EcoR124I.
<3>Biophys. J.
<4>84
<5>304a
<6>2003
<7>In bacteria, viral DNA is degraded by dedicated type I restriction enzymes.  When DNA is not
methylated, it is recognized as foreign and the enzyme starts DNA translocation consuming ATP,
to form a loop.  Cleavage will occur when translocation is blocked.  In this study the motor
properties of EcoR124I are investigated with Atomic Force Microscopy and Magnetic Tweezers.
Energetic barriers due to DNA bending and torsion, can be expected when EcoR124I moves over
DNA.  In the initial DNA EcoR124I complex AFM images show DNA tightly wrapped around the
enzyme.  In circular DNA no supercoiling is observed in front and behind the complex after
translocation.  Magnetic tweezers data however, show that the translocation does impose a
twist on the DNA.  Time traces of the length of single DNA molecules being processed by
EcoR124I show a non-constant translocation velocity, and repeated hick-ups.  A discrepancy is
observed between the translocation distance and the expected imposed twist.  The observed
behavior is consistent with frequent slipping events, both in translation and in rotation.

<>

<1>van Ormondt, H., Gorter, J., Havelaar, K.J., de Waard, A.
<2>Specificity of a deoxyribonucleic acid transmethylase induced by bacteriophage T2.  I. Nucleotide sequences isolated from Micrococcus luteus DNA methylated in vitro.
<3>Nucleic Acids Res.
<4>2
<5>1391-1400
<6>1975
<7>Deoxyribonucleic acid from Micrococcus luteus was methylated in vitro in the presence of
S-adenosyl-(14C methyl)methionine with a DNA methyltransferase purified from extracts of E.
coli infected with bacteriophage T2. The labelled DNA was degraded by enzymatic and specific
chemical methods and the resulting short oligonucleotides were separated and characterized.
The analytical data permit the conclusion that the DNA transmethylase reacts specifically with
N-G-A-T-C-N sequences in which it converts adenine to a 6-methyl-aminopurine residue.

<>

<1>van Ormondt, H., Lautenberger, J.A., Linn, S., de Waard, A.
<2>Methylated oligonucleotides derived from bacteriophage fd RF-DNA modified in vitro by E. coli B modification methylase.
<3>FEBS Lett.
<4>33
<5>177-180
<6>1973
<7>E. coli B modification methylase converts four adenine residues (presumably two
per strand) to N(6)-methylaminopurine in wild-type unmodified fd RF-DNA.  When
RF-DNA of the one-site mutant fd sB01 sB2 (strain 101) is the substrate, the
DNA receives two methyl groups while fd sB01 sB02 RF-DNA has lost is
susceptibility to the methylase by mutation.  This methylation is concomitant
with an increase in infectivity on E. coli B spheroplasts.  These results
probably mean that wild-type RF-DNA has two recognizable and specific
nucleotide sequences ("sites"), fd sB01 sB2 one, and fd sB01 sB02 none, which
by becoming methylated confer resistance to the E. coli B restriction enzyme.
The present study was undertaken to elucidate the structure of those sites
which bear "E. coli B specificity".  To this end, unmodified RF-DNA of
wild-type fd was methylated in vitro in the presence of purified modification
methylase from E. coli B and methyl tritiated S-adenosylmethionine.  The
[3H-methyl]RF-DNA was degraded to small fragments, and the tritiated
oligonucleotides characterized as to their sequence. Although a definitive
sequence was not obtained by this approach, an analysis of the sequenced
oligonucleotides showed the 3'- and 5'-nearest neighbors of the methylated
adenine residues to be A or C, G or C, respectively.  This finding demonstrates
that the sequence surrounding the modified base does not contain a twofold
rotational symmetry.

<>

<1>van Passel, M.W. et al.
<2>Genome sequence of the Verrucomicrobium Opitutus terrae PB90-1, an abundant inhabitant of rice paddy soil ecosystems.
<3>J. Bacteriol.
<4>193
<5>2367-2368
<6>2011
<7>Bacteria of the deeply branching phylum Verrucomicrobia are rarely cultured, yet commonly
detected in metagenomic libraries from aquatic, terrestrial and intestinal environments. We
have sequenced the genome of Opitutus terrae PB90-1, a fermentative anaerobe within this
phylum, isolated from rice paddy soil and capable of propionate production from plant-derived
polysaccharides.

<>

<1>van Passel, M.W. et al.
<2>Genome sequence of Victivallis vadensis ATCC BAA-548, an anaerobic bacterium from the phylum Lentisphaerae, isolated from the human  gastro-intestinal tract.
<3>J. Bacteriol.
<4>193
<5>2373-2374
<6>2011
<7>Victivallis vadensis ATCC BAA-548 represents the first cultured representative from the novel
phylum Lentisphaerae, a deep-branching bacterial lineage. Few cultured bacteria from this
phylum are known, and V. vadensis therefore represents an important organism for evolutionary
studies. V. vadensis is a strictly anaerobic sugar-fermenting isolate from the human
gastro-intestinal tract.

<>

<1>Van Pel, A., Colson, C.
<2>DNA restriction and modification systems in Salmonella:  II. Genetic complementation between the K and B systems of Escherichia coli and the Salmonella typhimurium system SB, with the same chromosomal location.
<3>Mol. Gen. Genet.
<4>135
<5>51-60
<6>1974
<7>E. coli x S. typhimurium partially diploid hybrids were constructed to
investigate the possibility of genetic complementation between the SA and the
SB restriction and modification systems of S. typhimurium and the K and B
systems of E. coli.  An hsdR-K mutation was complemented in a stable hybrid in
which the hsd+SA-hsd+SB-serB+ portion of the S. typhimurium chromosome was
integrated at a non-homologous locus.  By isolating hsd- mutants in that
hybrid, it was shown that complementation occurred between K and SB, but not
between K and SA.  Similarly, in a set of F-prime merodiploids bearing the SA,
SB and B systems, complementation was observed between B and SB, but not
between B and SA.

<>

<1>Van Roey, P., Belfort, M., Derbyshire, V.
<2>Homing endonuclease I-TevI: An atypical zinc finger with a novel function.
<3>Zinc Finger Proteins: from Atomic Contact to Cellular Function; Molecular Biology Intelligence Unit, Landes Bioscience, Iuchi, S., Kuldell, N., 
<4>0
<5>35-38
<6>2005
<7>I-TevI is a site-specific, sequence-tolerant homing endonuclease encoded by the td intron of
bacteriophage T4.  The enzyme consists of an N-terminal catalytic domain and a C-terminal
DNA-binding domain that are connected by a long, flexible linker.  The crystal structure of
the DNA-binding domain of I-TevI, residues 130 to 245, complexed with the 20-bp primary
binding region of its DNA target, reveals the presence of a zinc finger, comprising residues
151 to 167, that makes backbone contacts with the DNA from the minor groove.  Biochemical data
have shown that the zinc finger does not contribute to the DNA-binding affinity or to the
specificity of the enzyme, but rather that it has a novel function and acts as a distance
determinant that controls the relative positions of the catalytic and DNA-binding domains.

<>

<1>Van Roey, P., Derbyshire, V.
<2>GIY-YIG homing endonucleases - beads on a string.
<3>Nucleic Acids Mol. Biol.
<4>16
<5>67-83
<6>2005
<7>Homing endonucleases are intron- and intein-encoded proteins that initiate the mobility of
their particular host elements.  They recognize an intron- or intein-less version of their
host gene and introduce a double-strand break in the DNA.  This break is then repaired by
copying the intron- or intein-plus allele, using the hosts cellular machinery.  This results
in a gene-conversion event whereby both alleles become intron- or intein-plus.  Homing
endonucleases can be classified into four distinct families, based on the presence of
conserved sequence elements: the LAGLIDADG, GIY-YIG, His-Cys box, and HNH families.  However,
the His-Cys box and HNH families have been hypothesized to constitute a single bba-Me family,
and recent structural data support this classification.

<>

<1>Van Roey, P., Meehan, L., Kowalski, J.C., Belfort, M., Derbyshire, V.
<2>Catalytic domain structure and hypothesis for function of GIY-YIG intron endonuclease I-TevI.
<3>Nat. Struct. Biol.
<4>9
<5>806-811
<6>2002
<7>I-TevI, a member of the GIY-YIG family of homing endonucleases, consists of an N-terminal
catalytic domain and a C-terminal DNA-binding domain joined by a flexible linker. The GIY-YIG
motif is in the N-terminal domain of I-TevI, which corresponds to a phylogenetically
widespread catalytic cartridge that is often associated with mobile genetic elements. The
crystal structure of the catalytic domain of I-TevI, the first of any GIY-YIG endonuclease,
reveals a novel alpha/beta-fold with a central three-stranded antiparallel beta-sheet flanked
by three helices. The most conserved and putative catalytic residues are located on a shallow,
concave surface and include a metal coordination site. Similarities in the three-dimensional
arrangement of the catalytically important residues and the cation-binding site with those of
the His-Cys box endonuclease I-PpoI suggest the possibility of mechanistic relationships among
these different families of homing endonucleases despite completely different folds.

<>

<1>Van Roey, P., Waddling, C.A., Fox, K.M., Belfort, M., Derbyshire, V.
<2>Intertwined structure of the DNA-binding domain of intron endonuclease I-TevI with its substrate.
<3>EMBO J.
<4>20
<5>3631-3637
<6>2001
<7>I-TevI is a site-specific, sequence-tolerant intron endonuclease. The crystal structure of the
DNA-binding domain of I-TevI complexed with the 20 bp primary binding region of its DNA target
reveals an unusually extended structure composed of three subdomains: a Zn finger, an
elongated segment containing a minor groove-binding alpha-helix, and a helix-turn-helix. The
protein wraps around the DNA, mostly following the minor groove, contacting the phosphate
backbone along the full length of the duplex. Surprisingly, while the minor groove-binding
helix and the helix-turn- helix subdomain make hydrophobic contacts, the few base-specific
hydrogen bonds occur in segments that lack secondary structure and flank the intron insertion
site. The multiple base-specific interactions over a long segment of the substrate are
consistent with the observed high site specificity in spite of sequence tolerance, while the
modular composition of the domain is pertinent to the evolution of homing endonucleases.

<>

<1>van Schaik, W., Top, J., Riley, D.R., Boekhorst, J., Vrijenhoek, J.E., Schapendonk, C.M., Hendrickx, A.P., Nijman, I.J., Bonten, M.J., Tettelin, H., Willems, R.J.
<2>Pyrosequencing-based comparative genome analysis of the nosocomial pathogen Enterococcus faecium and identification of a large transferable pathogenicity island.
<3>BMC Genomics
<4>11
<5>239
<6>2010
<7>BACKGROUND: The Gram-positive bacterium Enterococcus faecium is an
important cause of nosocomial infections in immunocompromized patients.
RESULTS: We present a pyrosequencing-based comparative genome analysis of
seven E. faecium strains that were isolated from various sources. In the
genomes of clinical isolates several antibiotic resistance genes were
identified, including the vanA transposon that confers resistance to
vancomycin in two strains. A functional comparison between E. faecium and
the related opportunistic pathogen E. faecalis based on differences in the
presence of protein families, revealed divergence in plant carbohydrate
metabolic pathways and oxidative stress defense mechanisms. The E. faecium
pan-genome was estimated to be essentially unlimited in size, indicating
that E. faecium can efficiently acquire and incorporate exogenous DNA in
its gene pool. One of the most prominent sources of genomic diversity
consists of bacteriophages that have integrated in the genome. The
CRISPR-Cas system, which contributes to immunity against bacteriophage
infection in prokaryotes, is not present in the sequenced strains. Three
sequenced isolates carry the esp gene, which is involved in urinary tract
infections and biofilm formation. The esp gene is located on a large
pathogenicity island (PAI), which is between 64 and 104 kb in size.
Conjugation experiments showed that the entire esp PAI can be transferred
horizontally and inserts in a site-specific manner. CONCLUSIONS: Genes
involved in environmental persistence, colonization and virulence can
easily be aquired by E. faecium. This will make the development of
successful treatment strategies targeted against this organism a challenge
for years to come.

<>

<1>Van Sluys, M.A. et al.
<2>Comparative analyses of the complete genome sequences of Pierce's disease and citrus variegated chlorosis strains of Xylella fastidiosa.
<3>J. Bacteriol.
<4>185
<5>1018-1026
<6>2003
<7>Xylella fastidiosa is a xylem-dwelling, insect-transmitted, gamma-proteobacterium that causes
diseases in many plants, including
grapevine, citrus, periwinkle, almond, oleander, and coffee. X. fastidiosa
has an unusually broad host range, has an extensive geographical
distribution throughout the American continent, and induces diverse
disease phenotypes. Previous molecular analyses indicated three distinct
groups of X. fastidiosa isolates that were expected to be genetically
divergent. Here we report the genome sequence of X. fastidiosa (Temecula
strain), isolated from a naturally infected grapevine with Pierce's
disease (PD) in a wine-grape-growing region of California. Comparative
analyses with a previously sequenced X. fastidiosa strain responsible for
citrus variegated chlorosis (CVC) revealed that 98% of the PD X.
fastidiosa Temecula genes are shared with the CVC X. fastidiosa strain
9a5c genes. Furthermore, the average amino acid identity of the open
reading frames in the strains is 95.7%. Genomic differences are limited to
phage-associated chromosomal rearrangements and deletions that also
account for the strain-specific genes present in each genome. Genomic
islands, one in each genome, were identified, and their presence in other
X. fastidiosa strains was analyzed. We conclude that these two organisms
have identical metabolic functions and are likely to use a common set of
genes in plant colonization and pathogenesis, permitting convergence of
functional genomic strategies.

<>

<1>van Soolingen, D., de Haas, P.E.W., Blumenthal, R.M., Kremer, K., Sluijter, M., Pijnenburg, J.E.M., Schouls, L.M., Thole, J.E.R., Desens-Kroon, M.W.G., van Embden, J.D.A., Hermans, P.W.M.
<2>Host-mediated modification of PvuII restriction in Mycobacterium tuberculosis.
<3>J. Bacteriol.
<4>178
<5>78-84
<6>1996
<7>Restriction endonuclease PvuII plays a central role in restriction fragment length
polymorphism analysis of Mycobacterium tuberculosis complex isolates with IS6110 as a genetic
marker.  We have investigated the basis for an apparent dichotomy in PvuII restriction
fragment patterns observed among strains of the M. tuberculosis complex.  The chromosomal
regions of two modified PvuII restriction sites, located upstream of the katG gene and
downstream of an IS108I insertion sequence, were studied in more detail.  An identical 10-bp
DNA sequence (CAGCTGGAGC) containing a PvuII site was found in both regions, and site-directed
mutagenesis analysis revealed that this sequence was a target for modification.
Strain-specific modification of PvuII sites was identified in DNA from over 80% of the nearly
800 isolates examined.  Furthermore, the proportion of modifying and nonmodifying strains
differs significantly from country to country.

<>

<1>van Steensel, B., Henikoff, S.
<2>Identification of in vivo DNA targets of chromatin proteins using tethered Dam methyltransferase.
<3>Nat. Biotechnol.
<4>18
<5>424-428
<6>2000
<7>We have developed a novel technique, named DamID, for the identification of DNA loci that
interact in vivo with specific nuclear proteins in eukaryotes. By tethering Escherichia coli
DNA adenine methyltransferase (Dam) to a chromatin protein, Dam can be targeted in vivo to
native binding sites of this protein, resulting in local DNA methylation. Sites of methylation
can subsequently be mapped using methylation-specific restriction enzymes or antibodies. We
demonstrate the successful application of DamID both in Drosophila cell cultures and in whole
flies. When Dam is tethered to the DNA-binding domain of GAL4, targeted methylation is limited
to a region of a few kilobases surrounding a GAL4 binding sequence. Using DamID, we identified
a number of expected and unexpected target loci for Drosophila heterochromatin protein 1.
DamID has potential for genome-wide mapping of in vivo targets of chromatin proteins in
various eukaryotes.

<>

<1>van Steensel, B., Henikoff, S.
<2>Epigenomic profiling using microarrays.
<3>Biotechniques
<4>35
<5>346-357
<6>2003
<7>Genes occupy only a minor fraction of genomes such as ours; however, histone and nonhistone
chromosomal proteins and methylated DNA bases are
distributed over both genic and nongenic regions. These widespread
"epigenomic" features can be mapped and characterized by alternative
applications of the same microarray technologies that have been used for
conventional transcriptional profiling. Here we describe diverse
microarray-based strategies for profiling patterns of DNA methylation, DNA
replication, DNA binding, and chromatin-associated proteins and histone
modifications. The rapid progress that is being made in developing and
applying epigenomic profiling methods and the increasing availability of
microarrays mean that epigenomic profiling is likely to become a standard
research tool for understanding chromatin structure and gene expression
during development.

<>

<1>van Zyl, L.J., Deane, S.M., Louw, L.A., Rawlings, D.E.
<2>Presence of a family of plasmids (29 to 65 kilobases) with a 26-kilobase common region in different strains of the sulfur-oxidizing bacterium Acidithiobacillus caldus.
<3>Appl. Environ. Microbiol.
<4>74
<5>4300-4308
<6>2008
<7>Three large cryptic plasmids from different isolates of Acidithiobacillus caldus were rescued
by using an in vitro
transposition system that delivers a kanamycin-selectable marker and an
Escherichia coli plasmid origin of replication. The largest of the
plasmids, the 65-kb plasmid pTcM1, was isolated from a South African A.
caldus strain, MNG. This plasmid was sequenced and compared to that of
pTcF1 (39 kb, from strain "f," South Africa) and pC-SH12 (29 kb, from
strain C-SH12, Australia). With the exception of a 2.7-kb insertion
sequence, pC-SH12 appears to represent the DNA common to all three
plasmids and includes a number of accessory genes plus the plasmid
"backbone" containing the replication region. The two larger plasmids
carry, in addition, a number of insertion sequences of the ISL3 family
and a composite transposon related to the Tn21 subfamily containing a
highly mosaic region within the borders of the inverted repeats. Genes
coding for arsenic resistance, plasmid mobilization, plasmid stability,
and a putative restriction-modification system occur within these
mosaic regions.

<>

<1>Vanamee, E.S., Aggarwal, A.K.
<2>Metal-dependent type II restriction endonucleases.
<3>Handbook of Metalloproteins
<4>3
<5>742-756
<6>2004
<7>Restriction endonucleases are phosphodiesterases that bind double-stranded DNA with high
specificity and cleave the DNA to yield 5'-phosphate and 3'-hydroxyl groups as products.
Restriction endonucleases with their corresponding methyltransferases form the
restriction-modification systems of bacteria.  R-M systems are ubiquitous in bacteria; only a
few are known not to contain any R-M gene.  The recently sequenced Helicobacter pylori strain
26695, for instance, reveals more than a dozen R-M systems, though less than 30% are
functional.

<>

<1>Vanamee, E.S., Berriman, J., Aggarwal, A.K.
<2>An EM View of the FokI Synaptic Complex by Single Particle Analysis.
<3>J. Mol. Biol.
<4>370
<5>207-212
<6>2007
<7>FokI is a type IIS restriction endonuclease that recognizes the 5'-GGATG-3' sequence and
cleaves non-specifically at 9 and 13 base-pairs away on the top and bottom strands,
respectively, to produce a 5' overhang. FokI is a bipartite endonuclease with separate
recognition and cleavage domains. Because of its bipartite nature, FokI has received
considerable interest in generating chimeric nucleases for use in biotechnology, and recently
as possible therapeutic agents in gene therapy by initiating homologous gene recombination and
repair. Here we show, using single-particle electron microscopic studies, that the FokI active
complex prefers a single conformation in which the subunits are arranged in a doughnut shape
complex with protein-protein and possibly protein-DNA interactions stabilizing the cleavage
complex. Our electron microscopy (EM) model provides new insights into the activation
mechanism of FokI and how non-specific cleavage is avoided.

<>

<1>Vanamee, E.S., Hsieh, P.C., Zhu, Z.Y., Yates, D., Garman, E., Xu, S.Y., Aggarwal, A.K.
<2>Glucocorticoid receptor-like Zn(Cys)(4) motifs in BslI restriction endonuclease.
<3>J. Mol. Biol.
<4>334
<5>595-603
<6>2003
<7>BslI restriction endonuclease cleaves the symmetric sequence CCN(7)GG (where N=A, C, G or T).
The enzyme is composed of two subunits, alpha and
beta, that form a heterotetramer (alpha(2)beta(2)) in solution. The alpha
subunit is believed to be responsible for DNA recognition, while the beta
subunit is thought to mediate cleavage. Here, for the first time, we
provide experimental evidence that BslI binds Zn(II). Specifically, using
X-ray absorption spectroscopic analysis we show that the alpha subunit of
BslI contains two Zn(Cys)(4)-type zinc motifs similar to those in the
DNA-binding domain of the glucocorticoid receptor. This conclusion is
supported by genetic analysis of the zinc-binding motifs, whereby amino
acid substitutions in the zinc finger motifs are demonstrated to abolish
or impair cleavage activity. An additional putative zinc-binding motif was
identified in the beta subunit, consistent with the X-ray absorption data.
These data were corroborated by proton induced X-ray emission measurements
showing that full BslI contains at least three fully occupied Zn sites per
alpha/beta heterodimer. On the basis of these data, we propose a role for
the BslI Zn motifs in protein-DNA as well as protein-protein interactions.

<>

<1>Vanamee, E.S., Santagata, S., Aggarwal, A.K.
<2>FokI requires two specific DNA sites for cleavage.
<3>J. Mol. Biol.
<4>309
<5>69-78
<6>2001
<7>FokI is a bipartite restriction endonuclease that recognizes a non-palindromic DNA sequence,
and then makes double-stranded cuts
outside of that sequence to leave a 5' overhang. Earlier kinetic and
crystallographic studies suggested that FokI might function as a dimer.
Here, we show, using dynamic light-scattering, gel-filtration and
analytical ultracentrifugation, that FokI dimerizes only in the
presence of divalent metal ions. Furthermore, analysis of the DNA-bound
complex reveals that two copies of the recognition sequence are
incorporated into the dimeric complex and that formation of this
complex is essential for full activation of cleavage. These results
have broad implications for the mechanism by which monomeric type II
endonucleases achieve high fidelity.

<>

<1>Vanamee, E.S., Viadiu, H., Chan, S.H., Ummat, A., Hartline, A.M., Xu, S.Y., Aggarwal, A.K.
<2>Asymmetric DNA recognition by the OkrAI endonuclease, an isoschizomer of BamHI.
<3>Nucleic Acids Res.
<4>39
<5>712-719
<6>2011
<7>Restriction enzymes share little or no sequence homology with the exception of isoschizomers,
or enzymes that recognize and cleave the same DNA sequence. We present here the structure of a
BamHI isoschizomer, OkrAI, bound to the same DNA sequence (TATGGATCCATA) as that
cocrystallized with BamHI. We show that OkrAI is a more minimal version of BamHI, lacking not
only the N- and C-terminal helices but also an internal 3(10) helix and containing ?-strands
that are shorter than those in BamHI. Despite these structural differences, OkrAI recognizes
the DNA in a remarkably similar manner to BamHI, including asymmetric contacts via C-terminal
'arms' that appear to 'compete' for the minor groove. However, the arms are shorter than
in BamHI. We observe similar DNA-binding affinities between OkrAI and BamHI but OkrAI has
higher star activity (at 37=B0C) compared to BamHI. Together, the OkrAI and BamHI structures
offer a rare opportunity to compare two restriction enzymes that work on exactly the same DNA
substrate.

<>

<1>Vanamee, E.S., Viadiu, H., Kucera, R., Dorner, L., Picone, S., Schildkraut, I., Aggarwal, A.K.
<2>A view of consecutive binding events from structures of tetrameric endonuclease SfiI bound to DNA.
<3>EMBO J.
<4>24
<5>4198-4208
<6>2005
<7>Many reactions in cells proceed via the sequestration of two DNA molecules in a synaptic
complex. SfiI is a member of a growing family of restriction enzymes that can bind and cleave
two DNA sites simultaneously. We present here the structures of tetrameric SfiI in complex
with cognate DNA. The structures reveal two different binding states of SfiI: one with both
DNA-binding sites fully occupied and the other with fully and partially occupied sites. These
two states provide details on how SfiI recognizes and cleaves its target DNA sites, and gives
insight into sequential binding events. The SfiI recognition sequence (GGCCNNNN[downward
arrow]NGGCC) is a subset of the recognition sequence of BglI (GCCNNNN[downward arrow]NGGC),
and both enzymes cleave their target DNAs to leave 3-base 3' overhangs. We show that even
though SfiI is a tetramer and BglI is a dimer, and there is little sequence similarity between
the two enzymes, their modes of DNA recognition are unusually similar.

<>

<1>Vanat, I., Pristas, P., Kutejova, E., Judova, J., Godany, A., Jovorsky, P.
<2>SbvI restriction endonuclease from Streptococcus bovis.
<3>Lett. Appl. Microbiol.
<4>17
<5>297-299
<6>1993
<7>Restriction endonuclease SbvI, an isoschizomer of HaeIII, has been isolated from the rumen
amylolytic bacterium Streptococcus bovis II/1. SbvI was purified from a cell extract by
phosphocellulose chromatography and heparin-Sepharose chromatography. The recognition sequence
of SbvI was identified by digestion of pBR322, pUC9 and lambda DNA and comparing the cleavage
patterns obtained with computer-derived data. SbvI recognizes the 4-bp palindrome,
5'-GGCC-3' and cleaves DNA after the second G in the sequence, producing blunt ends.

<>

<1>Vanat, I., Pristas, P., Rybosoval, E., Godany, A., Javorsky, P.
<2>SruI restriction endonuclease from Selenomonas ruminantium.
<3>FEMS Microbiol. Lett.
<4>113
<5>129-132
<6>1993
<7>SruI, specific restriction endonuclease, has been characterized from Selenomonas ruminantium
isolated from the rumen of fallow deer. Results from the study demonstrate that S. ruminantium
18D possesses a type II restriction endonuclease, which recognizes the sequence 5'TTTAAA 3'.
The recognition sequence of SruI was identified using digestions on pBR322, pBR328, pUC18,
M13mp18RF, pACYC184 and lambda DNA. The cleavage patterns obtained were compared with
computer-derived data. SruI recognizes the palindromic hexanucleotide sequence and cleaves DNA
after the third T in the sequence, producing blunt ends. The purification and characterization
of restriction endonuclease SruI presented here is the first described for Selenomonas
ruminantium spp. and demonstrates that this microorganism possesses a DNA-cleaving enzyme with
the same specificity as DraI or AhaIII.

<>

<1>Vancott, E.M.
<2>Method for cloning and producing the SfiI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0437100 A
<5>
<6>1995
<7>The present invention is directed to a method for cloning and producing the SfiI restriction
endonuclease by 1) introducing the restriction endonuclease gene from S. fimbriatus into a
host whereby the restriction gene is expressed; 2) fermenting the host which contains the
plasmid encoding and expressing the SfiI restriction endonuclease activity; and 3) purifying
the SfiI restriction endonuclease from the fermented host which contains the plasmid encoding
and expressing the SfiI restriction endonuclease activity.

<>

<1>VanCott, E.M.
<2>Method for cloning and producing the NcoI restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0483797 B
<5>
<6>1995
<7>The present invention relates to an expression vector which expresses recombinant NcoI
endonuclease.

<>

<1>VanCott, E.M.
<2>Method for cloning and producing the NcoI restriction endonuclease and methylase.
<3>US Patent Office
<4>US 5202248
<5>
<6>1993
<7>The present invention is directed to a method for cloning and producing the NcoI restriction
endonuclease by 1) introducing the restriction endonuclease gene from N. corallina into a host
whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid
encoding and expressing the NcoI restriction endonuclease activity, and 3) purifying the NcoI
restriction endonuclease from the fermented host which contains the plasmid encoding and
expressing the NcoI restriction endonuclease activity.

<>

<1>Vandecandelaere, I., Van Nieuwerburgh, F., Deforce, D., Nelis, H.J., Coenye, T.
<2>Draft Genome Sequence of Methicillin-Resistant Staphylococcus epidermidis Strain  ET-024, Isolated from an Endotracheal Tube Biofilm of a Mechanically Ventilated  Patient.
<3>Genome Announcements
<4>2
<5>e00527-14
<6>2014
<7>Staphylococcus epidermidis strain ET-024 was isolated from a biofilm on an endotracheal tube
of a mechanically ventilated patient. This strain is resistant
to methicillin, and the draft genome sequence shares some characteristics with
other nosocomial S. epidermidis strains (such as S. epidermidis RP62A).

<>

<1>Vandenbogaert, M., Makeev, V.
<2>Analysis of bacterial RM-systems through genome-scale analysis and related taxonomy issues.
<3>In Silico Biology
<4>3
<5>127-143
<6>2003
<7>Recognition sites for type II restriction and modification enzymes in genomes of several
bacteria are recognized as semi-palindromic motifs and
are avoided at a significant degree. The key idea of contrast word
analysis with respect to RMS recognition sites, is that under-represented
words are likely to be selected against. Starting from over- or
underrepresented words corresponding to RMS recognition sites in specific
clades, the specificity of unknown R-M systems can be highlighted. Among
the known restriction enzymes, that are described in the REBASE database
of restriction and modification systems, many of their recognition sites
are still uncharacterized. Eventually, this motivates studies aimed at
assessing horizontal transferring events of RMS in micro-organisms through
the analysis of word usage biases in well-determined genomic regions. A
probabilistic model is built on a first-order Markovian chain. Statistics
on the k-neighborhood of a word is carried out to assess the biological
significance of a genomic motif. Efficient word counting procedures have
been implemented and statistics are used for the assessment of the
significance of individual words in large sequences. On the basis of the
set of most avoided words, and in accordance to the IUPAC coding
standards, suggestions are made regarding potential recognition sequences.
In certain cases, a comparison of avoided palindromic words in
taxonomically related bacteria shows a pattern of relatedness of their R-M
systems. For strengthening this analysis, the primary protein structure of
all type II R-M systems known in REBASE have been blasted against the
nr-GENBANK database. The combination of these analyses has revealed some
interesting examples of possible horizontal transfer events of R-M
systems.

<>

<1>Vandersteegen, K., Kropinski, A.M., Nash, J.H., Noben, J.P., Hermans, K., Lavigne, R.
<2>Romulus and Remus, Two Phage Isolates Representing a Distinct Clade within the Twortlikevirus Genus, Display Suitable Properties for Phage Therapy Applications.
<3>J. Virol.
<4>87
<5>3237-3247
<6>2013
<7>The renewed interest in controlling Staphylococcus aureus infections using their
natural enemies, bacteriophages, has led to the isolation of a limited number of
virulent phages so far. These phages are all members of the Twortlikevirus,
displaying little variance. We present two novel closely related (95.9% DNA
homology) lytic myoviruses, Romulus and Remus, with double-stranded DNA (dsDNA)
genomes of 131,333 bp and 134,643 bp, respectively. Despite their relatedness to
Staphylococcus phages K, G1, ISP, and Twort and Listeria phages A511 and P100,
Romulus and Remus can be proposed as isolates of a new species within the
Twortlikevirus genus. A distinguishing feature for these phage genomes is the
unique distribution of group I introns compared to that in other staphylococcal
myoviruses. In addition, a hedgehog/intein domain was found within their DNA
polymerase genes, and an insertion sequence-encoded transposase exhibits splicing
behavior and produces a functional portal protein. From a phage therapy
application perspective, Romulus and Remus infected approximately 70% of the
tested S. aureus isolates and displayed promising lytic activity against these
isolates. Furthermore, both phages showed a rapid initial adsorption and
demonstrated biofilm-degrading capacity in a proof-of-concept experiment.

<>

<1>Vanderstraeten, C., D'Halluin, K., Ruite, R.
<2>Improved targeted DNA insertion in plants.
<3>Japanese Patent Office
<4>JP 2007511232 A
<5>
<6>2007
<7>
<>

<1>Vandzurova, A., Hraskova, I., Judova, J., Javorsky, P., Pristas, P.
<2>Antibiotic resistance and restriction endonucleases in fecal enterococci of chamois (Rupicapra rupicapra Linnaeus, 1758).
<3>Folia Microbiol. (Praha)
<4>57
<5>355-358
<6>2012
<7>Two hundred eighty-four isolates of enterococci from feces of wild living chamois from alpine
environments were tested for sensitivity to
three antibiotics. Low frequency of resistance was observed in studied
enterococcal populations (about 5 % for tetracycline and erythromycin
and 0 % for ampicillin). In six animals, the population of enterococci
lacked any detectable resistance. Our data indicated that enterococcal
population in feces of the majority of studied animals did not
encounter mobile genetic elements encoding antibiotic resistance
probably due to spatial separation and/or due to low exposure to the
antibiotics. Based on resistance profiles observed, three populations
were analyzed for the presence of restriction endonucleases. The
restriction enzymes from two isolates-31K and 1K-were further purified
and characterized. Restriction endonuclease Efa1KI recognizes CCWGG
sequence and is an isoschizomer of BstNI. Endonuclease Efc31KI, a BsmAI
isoschizomer, recognizes the sequence GTCTC and it is a first
restriction endonuclease identified in Enterococcus faecium. Our data
indicate that restriction-modification (R-M) systems do not represent
an efficient barrier for antibiotic resistance spreading; enterococcal
populations colonized by antibiotics resistance genes were also
colonized by the R-M systems.

<>

<1>Vanek, P.G.
<2>Cloning and characterization of the BamHI restriction-modification methylase gene from Bacillus amyloliquefaciens.
<3>Ph.D. Thesis, Georgetown University, USA
<4>
<5>1-195
<6>1991
<7>The gene encoding the restriction-modification methylase (M.BamHI) from Bacillus
amyloliquefaciens has been successfully cloned into Escherichia coli. The clone encodes a 423
amino acids methyltransferase with identical sequence specificity as the BamHI endonuclease.
Plasmid pBamM9.0 containing this methylase gene was isolated from a Bacillus amyloliquefaciens
genomic library in the pSP64 cloning vector. A subclone, pBamH2.0, contains a 1.86 kb
HindIII/XmnI fragment of pBamM9.0 ligated into plasmid pSP64. Plasmid and genomic DNA isolated
from Epicurean Coli SURE cells harboring plasmid pBamM2.0 was resistant to cleavage by BamHI
endonuclease. During the course of cloning the cellular methylase gene, a provirally encoded
methylase (M.BamH2) with identical sequence specificity as the cellular methylase was also
cloned. Restriction mapping and Southern analysis confirmed the identity of two separate
methylase genes. The sequence predicted from the cellular methylase DNA clone show two
distinct regions of sequence similarity with N6-methyladenine and N4-methycytosine
methyltransferases. These two regions were very highly conserved in the cellular and proviral
methylases at both the protein and DNA level. Amino terminal sequencing of the cellular
methylase protein produced in E. coli indicated that translation begins at a rare UUG
initiation site. Southern analysis using the cloned methylase gene as a probe identified
similar genes in Bacillus amyloliquefaciens strains F and H, but not in any other BamHI
isoschizomeric bearing strains. The DNA sequence contains a 1269 base pair open reading frame
(ORF) which encodes a 49.1 kDa protein. A cell free DNA directed transcription translation
assay using a subclone pBSBamM2.0 as a template produced a single protein with an apparent
molecular weight of 49,000, which is in agreement with the molecular weight predicted from the
cloned sequence and of the purified protein. The methylase protein was purified to apparent
homogeneity by a three step chromatographic procedure utilizing phosphocellulose,
phenyl-sepharose, and sephadex G-150 column matrices. Fourty-four grams of E. coli harboring
pBamM2.0 yielded 1.75 mg methylase protein, an approximate 100 fold increase in protein
production as compared to Bacillus amyloliquefaciens (Nardone et al. 1983).

<>

<1>Vanek, P.G., Connaughton, J.F., Kaloss, W.D., Chirikijian, G.
<2>Comparison of cellular and viral methyltransferase genes from Bacillus amyloliquefaciens.
<3>FASEB J.
<4>4
<5>A2295
<6>1990
<7>The restriction modification system of B. amyloliquefaciens consists of a
restriction endonuclease, BamHI, and a cognate methyltransferase, each of which
recognize the hexanucleotide palindrome GGATCC.  In addition to the 56 kd
cellular methyltransferase, B. amyloliquefaciens contains a 30 kd prophage
encoded methylase which exhibits identical sequences specificity.  Recombinant
clones of both methyltransferases have been characterized in our laboratory.
The two clones, pBamM2.5 (prophage methylase), and pBamM2.0 (cellular
methylase) have been shown to produce methyltransferases capable of protecting
host genomic DNA from BamHI endonuclease.  Further, a lysate from cells
harboring clone pBamM2.0 has been shown to be capable of protecting lamda DNA
from BamHI cleavage in an in vitro assay.  For sequence comparison restriction
fragments from both pBamM2.5 and pBamM2.0 have been subcloned into M13 and
sequenced by dideoxy sequence analysis.  The clones have also been tested as
probes for the identification of methyltransferase genes in each of the
following BamHI isoschizomer containing strains; B. amyloliquefaciens strain F,
B. amyloliquefaciens strain N, A. liquefaciens NCIB 9417, B.
stearothermophilus, G. industricus IFO3260, and G. oxydans sub. melonogenes.
Southern blot analysis using pBamM2.5 as a probe has identified prophage
methyltransferase genes in each of the isoschizomer bearing bacterial strains
identified above.  The characterization of the two clones should facilitate
further studies of the restriction modification system of B. amyloliquefaciens.

<>

<1>Vanek, P.G., Connaughton, J.F., Kaloss, W.D., Chirikjian, J.G.
<2>The complete sequence of the Bacillus amyloliquefaciens strain H, cellular BamHI methylase gene.
<3>Nucleic Acids Res.
<4>18
<5>6145
<6>1990
<7>None

<>

<1>Vannier, P., Marteinsson, V.T., Fridjonsson, O.H., Oger, P., Jebbar, M.
<2>Complete Genome Sequence of the Hyperthermophilic, Piezophilic, Heterotrophic, and Carboxydotrophic Archaeon Thermococcus barophilus MP.
<3>J. Bacteriol.
<4>193
<5>1481-1482
<6>2011
<7>Thermococcus barophilus is a hyperthermophilic, anaerobic, mixed heterotrophic, and
carboxydotrophic euryarchaeon isolated from the deep
sea hydrothermal vent Snakepit site on the mid-Atlantic ridge at a depth
of 3,550 m. T. barophilus is the first true piezophilic, hyperthermophilic
archaeon isolated, having an optimal growth at 40 MPa. Here we report the
complete genome sequence of strain MP, the type strain of T. barophilus.
The genome data reveal a close proximity with Thermococcus sibiricus,
another Thermococcus isolated from the deep biosphere and a possible
connection to life in the depths.

<>

<1>Vanyushin, B.F.
<2>Enzymatic DNA methylation is an epigenetic control for genetic functions of the cell.
<3>Biochemistry
<4>70
<5>598-611
<6>2005
<7>In eukaryotic cells nuclear DNA is subjected to enzymatic methylation resulting in formation
of 5-methylcytosine residues mainly in CG and
CNG sequences. In plants and animals, this DNA methylation is species-,
tissue-, and organelle-specific. It changes (diminishes) with age and
is regulated by hormones. On the other hand, genome methylation can
control hormonal signal. There are replicative and postreplicative DNA
methylations. They are served by multiple DNA-methyl-transferases with
different site specificity. Replication is accompanied by appearance of
hemimethylated sites in DNA; pronounced asymmetry of DNA chain
methylation disappears at the end of the cell cycled a model of
regulation of replication by DNA methylation is suggested. DNA
methylation controls all genetic processes in the cell (replication,
transcription, DNA repair, recombination, gene transposition) and it is
a mechanism of cell differentiation, gene discrimination, and
silencing. Prohibition of DNA methylation stops development
(embryogenesis), switches on apoptosis, and is usually lethal.
Distortions in DNA methylations result in cancerous cell
transformation, and the DNA methylation pattern is one of the safe
cancer diagnostics at early stages of carcinogenesis. The malignant
cell has a different DNA methylation pattern and a set of
DNA-methyltransferase activities expressed as compared with normal
cells. Inhibition of DNA methylation in plants is accompanied by
induction of genes of seed storage proteins and flowering. In
eukaryotes one and the same gene can be methylated both on cytosine and
adenine residues; thus, there are, at least, two different and probably
interdependent systems of DNA methylation in the cell. First higher
eukaryotic adenine DNA-methyltransferase was isolated from plants; this
enzyme methylates DNA with formation of N-6-methyladenine residues in
the sequence TGATCA -> TGm(6)ATCA. Plants have AdoMet-dependent
endonucleases sensitive to DNA methylation status, therefore, like
microorganisms, plants seem to have a restriction-modification (R-S)
system. Revelation of an essential role of DNA methylation in the
regulation of genetic processes has laid a foundation for and
materialized epigenetics and epigenomics.

<>

<1>Vanyushin, B.F.
<2>DNA methylation in plants.
<3>Curr. Top. Microbiol. Immunol., Springer-Verlag, Doerfler, W., Bohm, P., Berlin, Germany
<4>301
<5>67-122
<6>2006
<7>DNA in plants is highly methylated, containing 5-methylcytosine (m(5)C) and N-6-methyladenine
(m(6)A); m(5)C is located mainly in symmetrical CG and CNG sequences but it may occur also in
other non-symmetrical contexts. m(6)A but not m(5)C was found in plant mitochondrial DNA. DNA
methylation in plants is species-, tissue-, organelle- and age-specific. It is controlled by
phytohormones and changes on seed germination, flowering and under the influence of various
pathogens (viral, bacterial, fungal). DNA methylation controls plant growth and development,
with particular involvement in regulation of gene expression and DNA replication. DNA
replication is accompanied by the appearance of under-methylated, newly formed DNA strands
including Okazaki fragments; asymmetry of strand DNA methylation disappears until the end of
the cell cycle. A model for regulation of DNA replication by methylation is suggested.
Cytosine DNA methylation in plants is more rich and diverse compared with animals. It is
carried out by the families of specific enzymes that belong to at least three classes of DNA
methyltransferases. Open reading frames (ORF) for adenine DNA methyltransferases are found in
plant and animal genomes, and a first eukaryotic (plant) adenine DNA methyltransferase
(wadmtase) is described; the enzyme seems to be involved in regulation of the mitochondria
replication. Like in animals, DNA methylation in plants is closely associated with histone
modifications and it affects binding of specific proteins to DNA and formation of respective
transcription complexes in chromatin. The same gene (DRM2) in Arabidopsis thaliana is
methylated both at cytosine and adenine residues; thus, at least two different, and probably
interdependent, systems of DNA modification are present in plants. Plants seem to have a
restriction-modification (R-M) system. RNA-directed DNA methylation has been observed in
plants; it involves de novo methylation of almost all cytosine residues in a region of
siRNA-DNA sequence identity; therefore, it is mainly associated with CNG and non-symmetrical
methylations (rare in animals) in coding and promoter regions of silenced genes. Cytoplasmic
viral RNA can affect methylation of homologous nuclear sequences and it may be one of the
feedback mechanisms between the cytoplasm and the nucleus to control gene expression.

<>

<1>Vanyushin, B.F.
<2>A view of an elemental naturalist at the DNA world (base composition, sequences, methylation).
<3>Biochemistry
<4>72
<5>1289-1298
<6>2007
<7>The pioneering data on base composition and pyrimidine sequences in DNA of pro- and eukaryotes
are considered , and their significance for the origin of genosystematics is discussed. The
modern views on specificity and functional role of enzymatic DNA methylation in eukaryotes are
described. DNA methylation controls all genetic functions and is a mechanism of cellular
differentiation and gene silencing. A model of regulation of DNA replication by methylation is
suggested . Adenine DNA methylation in higher eukaryotes ( higher plants) was first observed,
and it was established that one and the same gene can be methylated at both cytosine and
adenine moieties. Thus, there are at least two different and seemingly interdependent DNA
methylation systems present in eukaryotic cells. The first eukaryotic adenine DNA-methyl-
transferase is isolated from wheat seedlings and described: the enzyme methylates DNA with
formation of N-6-methylade- nine in the sequence TGATCA -> TGm(6)ATCA. It is found that higher
plants have endonucleases that are dependent on S- adenosyl-L-methionine (SAM) and sensitive
to DNA methylation status. Therefore, as in bacteria, plants seem to have a
restriction-modification (R-M) system. A system of conjugated up- and down-regulation of
SAM-dependent endonucleases by SAM modulations is found in plants. Revelation of an essential
role of DNA methylation in regulation of genetic processes is a fundament of materialization
of epigenetics and epigenomics.

<>

<1>Vanyushin, B.F., Aleksandrushkina, N.I., Agarkova, O.A.
<2>Cytokinins do not markedly affect the methylation of DNA adenine residues in cell cultures of Escherichia coli B.
<3>Biokhimiia
<4>54
<5>1666-1672
<6>1989
<7>6-Benzylaminopurine (1mg/ml) does not influence the growth of E. coli B cell cultures or the
number of [8-14C] labeled N6-methyladenine residues in the total DNA [(100 m6A/(A + m6A)=
1.7].  The growth of bacterial cells in the presence of adenine or cytokinins (6-BAP, kinetin,
zeatin) (1 mg/ml) was unaccompanied by significant changes in the intracellular content of
plasmid pBR322.  The mode of restriction by endonuclease CfuI hydrolyzing the Gm6ATC site of
plasmid pBR322 from E. coli B cells grown in the presence of adenine or one of the
above-mentioned cytokinins is identical.  These plasmids also have identical restriction
products with MboI or Sau3AI.  Thus, the cytokinins under study do not markedly affect the
methylation of adenine residues in total DNA of E. coli B cell cultures and the GATC sequence
in plasmids pBR322 isolated from these cells.

<>

<1>Vanyushin, B.F., Buryanov, Y.I., Belozersky, A.N.
<2>Distribution of N6-methyladenine in DNA of T2 phage and its host Escherichia coli B.
<3>Nature New Biol.
<4>230
<5>25-27
<6>1971
<7>N6-methyladenine (6-MeAde) and 5-methylcytosine occur as minor bases in
bacterial and phage DNA and seem to result from the selective methylation of
adenine and cytosine residues by specific DNA methylases.  Methylation is the
final stage in DNA synthesis and is essential for the phenomenon of host
modification of phages; it is one of the mechanisms controlling DNA replication
in the cell.  A study of the distribution of minor bases in DNA is important
not only for the elucidation of the specificity and mechanism of action of DNA
methylases but also for an understanding of the purpose of this methylation.
Until recently there has been a lack of adequate methods for an analysis of the
distribution of purines, including 6-MeAde, in DNA.  We have developed a method
for the specific chemical degradation of DNA into purine sequences, based on
the hydrolysis of apyrimidinic DNA by aniline, which facilities a study of the
content and position of 6-MeAde residues in unmodified purine sequences.  By
means of this method we have hydrolysed E. coli B and T2 phage DNA to yield
purine sequences and have determined the frequency of different purine
isostichs (fragments with equal numbers of nucleotide residues) and the amounts
of 6-MeAde in each.  We have checked that 6-MeAde, and also the isolated purine
sequences, do not undergo marked degradation either in the course of DNA
hydrazinolysis or in the subsequent hydrolysis of the apyrimidinic acid by
aniline.  DNA was isolated from T2 phage and E. coli B cells collected at the
end of the logarithmic phage of growth, according to the procedure of Marmur,
with additional phenol deproteinziation.

<>

<1>Vanyushin, B.F., Danilevich, V.N.
<2>Location of 5-methylcytosine in Escherichia coli and phage DD7 DNA.
<3>Biokhimiia
<4>39
<5>1293-1301
<6>1974
<7>After the cultivation of E. coli C Met cells, infected and uninfected by phage
DD7, with (H3-methyl)-Met, the bacterial and phage DNA were isolated and the
distribution and location of 5-methylcytosine (MC) were studied in different
pyrimidine sequences.  Methylcytosine was not found in the monopyrimidine
fragments of phage DD7 and E. coli C MC DNA and was present mainly in the di-
and tripyrimidine sequences (about 75% of all the MC).  A small amount of MC
was found in long pyrimidine units (4-7 long).  The DNAs of different strains
of E. coli (C and K12) did not differ in frequency of occurrence of MC in
different isopliths and were very similar to the DNA of phage DD7.  The
specificity of the methylation of cytosine residues in phage DD7 DNA and its
host E. coli C was the same.  In phage DD7 and E. coli C-MC DNA MC is located
in the following sequences: ...Pu-C-MC-Pu...; ...Pu-C-MC-C-Pu...;
Pu-C-MC-T-Pu...

<>

<1>Vanyushin, B.F., Dobritsa, A.P.
<2>On the nature of the cytosine-methylated sequence in DNA of Bacillus brevis var. G.-B.
<3>Biochim. Biophys. Acta
<4>407
<5>61-72
<6>1975
<7>On growing the cells of Bacillus brevis S methionine-auxotroph mutant in the
presence of [Me-3 H] methionine, practically all the radioactivity incorporated
into DNA is found to exist in 5-methylcytosine and N6-methyladenine.  The
analysis of pyrimidine isopliths isolated from DNA shows that radioactivity
only exists in mono- and dinucleotides and the content of 5-methylcytosine in
R-m5 C-R and R-m5 C-T-R oligonucleotides is equal.  The analysis of
dinucleotides isolated from DNA by means of pancreatic DNAase hydrolysis allows
the nature of purine residues neighbouring 5-methylcytosine to be identified
and shows that 5-methylcytosine localizes in G-m5 C-A and G-m5 C-T-R fragments.
B. brevis S DNA methylase modifying cytosine residues recognizes the GC(A/T)GC
degenerate nucleotide sequence which is a part of the following complementary
structure with a two-fold rotational axis of symmetry: (5')
...N'-G-C*-T-G-C-N... (3') (3') ...N -C-G-A-C*-G-N'... (5') (Methylated
cytosine residues are asterisked).  Cytosine-modifying DNA methylase activity
is isolated from B. brevis cells; it is capable of methylating in vitro
homologous and heterologous DNA.  Hence DNA in bacterial cells can be
undermethylated.  This enzyme methylates cytosine residues in native and
denatured DNA in the same nucleotide sequences.  Specificity of methylation of
cytosine residues in vitro and in vivo does not depend on the nature of
substrate DNA.  DNA methylases of different variants of B. brevis (R, S, P+,
P-) methylate cytosine residues in the same nucleotide sequences.  It means
that specificity or methylation of DNA cytosine residues in the cells of
different variants of B. brevis is the same.

<>

<1>Vanyushin, B.F., Dobritsa, A.P.
<2>Specificity of the methylation of the cytosine residues in the DNA of Bacillus brevis var. G-B.
<3>Mol. Biol. (Mosk)
<4>9
<5>283-295
<6>1975
<7>After the growth of a methionine-auxotrophic mutant of Bacillus brevis S in the
prsence of [methyl-3H]-methionine, practically all the label incorporated into
DNA was detected in 5-methylcytosine and N6-methyladenine.  In an analysis of
the pyrimidine isopliths isolated from DNA, it was shown that all the
radioactivity of MC is uniformly contained in the oligonucleotides Pur-MC-Pur
and Pur-MC-T-Pur.  An analysis of the dinucleotides isolated from the DNA by
pancreatic DNAase made it possible to establish that MC is localized in the
fragments G-MC-A and G-MC-T-Pur.  DNA methylase of B. brevis S, which modifies
cytosine residues, recognizes the degenerate nucleotide sequence GC(T/A)GC,
which is included in the complementary structure with second-order rotational
symmetry:   (5') ...N'-G-MC-T-G-C-N...(3') (3') ....N-C-G-A-MC-G-N'...(5') an
extract possessing cytosine-modifying methylase activity and capable of
methylating homologous and heterologous DNA's in vitro was isolated from cells
of B. brevis.  This means that in the cells of the bacterium, DNA may be
partially undermethylated.  This enzyme methylates cytosine residues in native
and denatured DNA's in the same nucleotide sequence.  In comparison with the
native DNA, in the denatured DNA the level of methylation of the adenine
residues, but not of the cytosine residues, is decreased.  The specificity of
the methylation of the cytosine residues in vitro and in vivo is the same and
does not depend on the nature of the substrate DNA's (calf thymus, Pseudomonas
aeruginosa, etc.).  DNA methylases from different variants of B. brevis
(R,S,P+,P-) methylate the cytosine residues in the same nucleotide sequences.
Consequently, the specificity of the methylation of the cytosine residues in
the cells of different variants of B. brevis is the same.

<>

<1>Vanyushin, B.F., Poirier, L.A.
<2>Drosophila melanogaster genomic DNA sequence homologous to mammalian cytosine DNA-methyltransferase gene.
<3>Biochem. Mol. Biol. Int.
<4>39
<5>353-358
<6>1996
<7>Using the Southern blotting procedure we have shown that Drosophila genomic
DNA hybridizes with 4423-bp C-terminal fragment of murine cytosine DNA-methyltransferase
gene.  Thus, the Drosophila genome has a sequence homologous to the mammalian cytosine DNA-
methyltransferase gene.  We assume that DNA methylation most likely responsible for strong CpG
suppression in the Drosophila genome mainly was catalyzed by a cytosine DNA-methyltransferase
that has since been lost.

<>

<1>Varani, A.M., Lemos, M.V., Fernandes, C.C., Lemos, E.G., Alves, E.C., Desiderio, J.A.
<2>Draft Genome Sequence of Bacillus thuringiensis var. thuringiensis Strain T01-328, a Brazilian Isolate That Produces a Soluble Pesticide Protein, Cry1Ia.
<3>Genome Announcements
<4>1
<5>e00817-13
<6>2013
<7>Bacillus thuringiensis var. thuringiensis strain T01-328, isolated from Cubatao county (Sao
Paulo State, Brazil), produces a soluble pesticide protein, Cry1Ia,
during vegetative growth. Here, we report the 7.089-Mbp draft genome sequence,
composed of a 5.5-Mb chromosome and 14 plasmids, which is the largest B.
thuringiensis genome sequenced to date.

<>

<1>Vardimon, L., Gunthert, U., Doerfler, W.
<2>In vitro methylation of the BsuRI (5'-GGCC-3') sites in the E2a region of adenovirus type 2 DNA does not affect expression in Xenopus laevis oocytes.
<3>Mol. Cell. Biol.
<4>2
<5>1574-1580
<6>1982
<7>The early region 2a (E2a) of adenovirus type 2 (Ad2) DNA codes for a 72,000-dalton DNA-binding
protein and is expressed in the Ad2-transformed hamster cell line HE1 but not in cell lines
HE2 and HE3 (H. Esche, J. Virol. 41:1076-1082, 1982; K. Johansson et al., J. Virol.
27:628-639, 1978). An inverse correlation between DNA methylation at the 5'-CCGG-3' sites of
the E2a region and of gene expression in these cell lines has been observed (L. Vardimon et
al., Nucleic Acids Res. 8:2461-2473, 1980). When the cloned E2a region of Ad2 DNA is
methylated in vitro at the 5'-CCGG-3' sites, the gene is not transcribed after being
injected into the nuclei of Xenopus laevis oocytes, whereas unmethylated DNA is expressed (L.
Vardimon et al., Eur. J. Cell Biol. 25:13-15, 1981; L. Vardimon et al., Proc. Natl. Acad. Sci.
U.S.A. 79:1073-1077, 1982). These data demonstrate that DNA methylation is directly involved
in the shut-off of transcription. In the present communication we investigated in detail the
control region of the gene for the DNA-binding protein in Ad2-transformed cell lines and
showed that the first late control region (map coordinate 72 on the viral DNA) of the E2a
region is present in its entirety in cell lines HE1, HE2, and HE3. The HaeIII sites
(5'-GGCC-3') in the E2a region in all three cell lines were not methylated. When the DNA
methyltransferase BsuRI was used, all 5'-GGCC-3' sites in the cloned E2a region of Ad2 DNA
were methylated in vitro. It was shown that methylation of these sites did not inhibit the
expression of this viral gene in X. laevis oocytes. Thus, for methylation to affect gene
expression in the E2a region it has to occur at specific sites (e.g., 5'-CCGG-3') which may
be different for other genes.

<>

<1>Vardimon, L., Rich, A.
<2>In Z-DNA the sequence G-C-G-C is neither methylated by HhaI methyltransferase nor cleaved by HhaI restriction endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>81
<5>3268-3272
<6>1984
<7>Plasmids carrying 24- or 32-base-pair inserts of alternating (dG-dC) residues
were used to analyze the level of methylation of the G-C-G-C sites by HhaI DNA
methyltransferase and their cleavage by HhaI endonuclease in the B-DNA or Z-DNA
conformation.  In supercoiled plasmids in which the inserts formed Z-DNA, the
extent of methylation at the insert G-C-G-C sites was dramatically lower than
the level of methylation at the G-C-G-C sites located outside the insert in the
same plasmid.  Similarly, cleavage by HhaI endonuclease was sharply lowered
when the insert was in the Z-DNA form.  In the relaxed plasmid, all its G-C-G-C
sites were methylated to the same extent and the unmethylated sites were
readily cleaved.  After treatment with the methylase, the supercoiled plasmid
was linearized and then digested with Hha restriction endonuclease.  This
exposed unmethylated G-C-G-C sites from the insert that had been protected
against cleavage in the Z conformation.  A chemical reaction was used to study
the distribution of the unmethylated cytosine residues.  No accumulation of
unmethylated cytosine residues was found anywhere along the entire 32-base-pair
insert, which is consistent with a cooperative B-Z transition.

<>

<1>Varga, J.J., Losada, L., Zelazny, A.M., Brinkac, L., Harkins, D., Radune, D., Hostetler, J., Sampaio, E.P., Ronning, C.M., Nierman, W.C., Greenberg, D.E., Holland, S.M., Goldberg, J.B.
<2>Draft Genome Sequence Determination for Cystic Fibrosis and Chronic Granulomatous Disease Burkholderia multivorans Isolates.
<3>J. Bacteriol.
<4>194
<5>6356-6357
<6>2012
<7>Burkholderia multivorans is a Gram-negative bacterium and a member of the Burkholderia cepacia
complex, which is frequently associated with respiratory
infections in people with cystic fibrosis (CF) and chronic granulomatous disease
(CGD). We are reporting the genome sequences of 4 B. multivorans strains, 2 from
CF patients and 2 from CGD patients.

<>

<1>Varga, J.J., Losada, L., Zelazny, A.M., Kim, M., McCorrison, J., Brinkac, L., Sampaio, E.P., Greenberg, D.E., Singh, I., Heiner, C., Ashby, M., Nierman, W.C., Holland, S.M., Goldberg, J.B.
<2>Draft Genome Sequences of Burkholderia cenocepacia ET12 Lineage Strains K56-2 and BC7.
<3>Genome Announcements
<4>1
<5>e00841-13
<6>2013
<7>The Burkholderia cepacia complex (BCC) is a group of closely related bacteria that are
responsible for respiratory infections in immunocompromised humans, most
notably those with cystic fibrosis (CF). We report the genome sequences for
Burkholderia cenocepacia ET12 lineage CF isolates K56-2 and BC7.

<>

<1>Varshney, H., Iqbal, J., Seleemuddin, M.
<2>Immobilization of the restriction endonuclease EcoRI.  Usefulness of a polyclonal antibody support.
<3>Enzyme Microb. Technol.
<4>25
<5>172-176
<6>1999
<7>The restriction enzyme EcoRI has been immobilized by using an immunoaffinity-based procedure
on Sepharose 4B.  The antibody support, prepared by coupling the gamma-globulin fraction of
serum of immunized rabbits to CNBr-activated Sepharose-4B, was highly effective in binding
EcoRI from solution although only about half of the bound activity appeared to be expressed by
the immobilized preparations.  Restriction activity of EcoRI immobilized on the antibody
support was indistinguishable from that of soluble enzyme on the linear phage-DNA and
supercoiled plasmids pBR322 and pBHLUC P.  Binding to the antibody support markedly enhanced
the resistance of EcoRI to heat inactivation, and the preparation, unlike the native enzyme,
retained significant activity after exposure to a temperature of 65 C.  It was also possible
to immobilize EcoRI directly from the cell lysates of Escherichia coli pMB4, an EcoRI
overproducing strain.  The immobilized preparation did not possess nonspecific nuclease
activity that was prominent in the lysates, suggesting specificity in the binding of EcoRI to
the antibody support.

<>

<1>Varshney, H., Saleemuddin, M., Rhee, J.I., Schugerl, K.
<2>Immobilization of SpA::EcoRI on IgG support improves the thermal stability of restriction activity.
<3>Process. Biochem.
<4>37
<5>275-278
<6>2001
<7>A plasmid-harbouring E. coli JM109 (3P) strain was cultivated for the overproduction of the
genetically engineered fusion protein SpA::EcoRI. The fusion protein could be affinity bound
selectively and directly from the 25-50% ammonium sulphate fraction of the lysate of E. coli
JM109 on to IgG-Sepharose.  The preparation obtained thus exhibited high restriction activity
on lambda DNA and linearized the plasmids pBR322 and pUC19.  As compared to the native EcoRI
the activity of the immobilized preparation was more resistant to thermal inactivation.

<>

<1>Vasconcellos, R.L., Mendes, R., Taketani, R.G., Zucchi, T.D., Melo, I.S.
<2>Draft Genome Sequence of Pseudomonas sp. Strain CMAA 1215, a Plant Growth-Promoting Bacterium Isolated from a Brazilian Mangrove.
<3>Genome Announcements
<4>1
<5>e00995-13
<6>2013
<7>The aim of this study was to sequence the genome of the plant growth-promoting Pseudomonas sp.
strain CMAA 1215, an osmotolerant bacterium isolated from
mangrove soil.

<>

<1>Vasconcelos, A.T. et al.
<2>Swine and Poultry Pathogens: the Complete Genome Sequences of Two Strains of Mycoplasma hyopneumoniae and a Strain of Mycoplasma synoviae.
<3>J. Bacteriol.
<4>187
<5>5568-5577
<6>2005
<7>This work reports the results of analyses of three complete mycoplasma genomes, a pathogenic
(7448) and a nonpathogenic (J) strain of the swine
pathogen Mycoplasma hyopneumoniae and a strain of the avian pathogen
Mycoplasma synoviae; the genome sizes of the three strains were 920,079
bp, 897,405 bp, and 799,476 bp, respectively. These genomes were compared
with other sequenced mycoplasma genomes reported in the literature to
examine several aspects of mycoplasma evolution. Strain-specific regions,
including integrative and conjugal elements, and genome rearrangements and
alterations in adhesin sequences were observed in the M. hyopneumoniae
strains, and all of these were potentially related to pathogenicity.
Genomic comparisons revealed that reduction in genome size implied loss of
redundant metabolic pathways, with maintenance of alternative routes in
different species. Horizontal gene transfer was consistently observed
between M. synoviae and Mycoplasma gallisepticum. Our analyses indicated a
likely transfer event of hemagglutinin-coding DNA sequences from M.
gallisepticum to M. synoviae.

<>

<1>Vasiljeva, L.Y., Zheleznaya, L.A., Matvienko, N.I.
<2>A site-specific endonuclease BspR7I from Thermophilic strain.
<3>Biokhimiia
<4>61
<5>2147-2157
<6>1996
<7>A site-specific endonuclease BspR7I preparation has been isolated and purified to homogeneity
from the thermophilic strain Bacillus sp. R7.  DNA cleavage proceeds according to the scheme
5'-CC/TNAGC-3' 3'-GGANT/CC-5' and thus the enzyme belongs to the second class of
restriction endonucleases and is an isoschizomer of Bsu36I.  The isolated protein has a
molecular mass of 37 kD and is present in solution in the form of a monomer.  BspR7I is active
over a wide range of temperatures, from 37 to 48oC.  The enzyme is relatively stable.

<>

<1>Vasiljeva, L.Y., Zheleznaya, L.A., Matvienko, N.I.
<2>Site-specific endonuclease SscL1I from strain Staphylococcus species L1.
<3>Biokhimiia
<4>63
<5>212-218
<6>1998
<7>A site-specific endonuclease SscL1I preparation has been isolated and purified to near
homogeneity from the strain Staphylococcus sp. L1 without admixtures of other nuclease
activity.  DNA cleavage proceeds according to the scheme: 5'-G/ANTC-3' 3'-CTNA/G-5, and
thus the isolated enzyme is an isoschizomer of restriction endonuclease HinfI and belongs to
the second class of restriction endonucleases.  SscL1I works over a broad range of temperature
and pH.  The enzyme is characterized by high stability during storage.

<>

<1>Vasiljeva, L.Y., Zheleznaya, L.A., Matvienko, N.I.
<2>Cloning and expression of a new site-specific methyltransferase M.SscL1I from Staphylococcus sp. L1.
<3>Biokhimiia
<4>65
<5>565-570
<6>2000
<7>The gene of the new site-specific methyltransferase M.SscL1I belonging to the same
modification-restriction system as the previously described site-specific endonuclease
SscL1I has been cloned from the natural strain Staphylococcus sp. L1. A plasmid to express the
methylase gene under control of the T7 phage-specific promoter has been constructed.
Conditions were found to express the recombinant methylase M.SscL1I and to purify it to near
homogeneity. It is shown that the methylase modifies the adenine base in the recognition site
5'-GANTC-3'.

<>

<1>Vasquez, C.
<2>Isolation and partial characterization of BstVI, a thermostable isoschizomer of XhoI.
<3>Biochem. Int.
<4>10
<5>655-662
<6>1985
<7>A type II restriction endonuclease, which has been named BstVI, was isolated and partially
purified from a spore-forming, gram-positive thermophilic bacilli.  On the basis of its
digestion patterns on varous DNA's, it was concluded that this enzyme is an isoschizomer of
XhoI, isolated originally from Xanthomonas holcicola.  Besides being highly thermostable, the
enzyme is produced in very large amounts by this bacterial strain.  A single purification step
renders it free of unspecific nucleases and suitable for performing restriction analysis and
cloning experiments.

<>

<1>Vasquez, C.
<2>Structural and biochemical characterization of the modification-restriction system of Bacillus stearothermophilus V.
<3>Arch. Biol. Med. Exp. (Santiago)
<4>21
<5>r338
<6>1988
<7>None

<>

<1>Vasquez, C., Adasme, A., Gonzalez, E.
<2>Aislamiento Y caracterizacion de endonucleasas termoestables:  BstVI, un isosquizomero de XhoI (isolation and characterization of thermostable endonucleases:  BstVI, an isoschizomer of XhoI).
<3>Arch. Biol. Med. Exp. (Santiago)
<4>18
<5>r371
<6>1985
<7>Recientemente hemos purificado una endonucleasa de restriccion tipo II, la cual
fue aislade de un bacilo termofilico gram positivo.  De acuerdo a pruebas
microbiologicas estandares, la bacteria resulto ser del tipo Bacillus
stearothersophilus y la enzima fue denominada BstVI.  Sobre la base de los
patrones de digestion de los diversos DNAs utilizados como sustrato, hemos
concluido que BstVI es un isosquizomero de XhoI, aislada originalmente de
Xanthomonas holcicola.  Ademas de ser muy termoestable (tempertura optima de
75C), BstVI es producida en gran cantidad por esta cepa bacteriana.
Practicamente un solo paso de purificacion hace possible la eliminacion de
nucleasas inespecificas y port lo tanto, su uso con fines de analisis de
restriccion y de clonamiento molecular.  Se han determinado algunas de las
condiciones optimas para la activated enzimatica, ademas de probar la
estabilidad de la enzima frente a una serie de agentes desnaturantes de
proteinas.

<>

<1>Vasquez, C., Saavedra, C., Gonzalez, E.
<2>Cloning the BstVI restriction-modification system in Escherichia coli.
<3>Gene
<4>102
<5>83-85
<6>1991
<7>A standard DNA modification methyltransferase (MTase) selection protocol was
followed to clone the BstVI restriction and modification system from Bacillus
stearothermophilus in Escherichia coli.  Both genes were contained in a 4.4-kb
EcoRI fragment from B. stearothermophilus V chromosomal DNA.  The heterologous
expression of these genes did not depend on their orientation in the vector,
suggesting that the genes are expressed in E. coli under the control of
promoters located on the cloned fragment.  Subcloning experiments demonstrated
that the bstVIR gene was expressed in the absence of its cognate MTase.

<>

<1>Vasquez, C., Vicuna, R.
<2>The genus Thermus: restriction endonucleases and modification methylases.
<3>Arch. Biol. Med. Exp. (Santiago)
<4>15
<5>417-421
<6>1982
<7>This work reviews the present knowledge of the site-specific endonucleases and
methylases involved in the restriction-modification systems in bacteria
belonging to the genus Thermus.  In addition, we describe part of our work
concerning the purification and properties of these enzymes from Thermus
thermophilus HB8 and Thermus flavus AT-62.

<>

<1>Vasquez, C.C., Saavedra, C.P., Pichuantes, S.E.
<2>Nucleotide sequence of the gene encoding the BstLVI DNA methyltransferase: comparison with other amino-DNA methyltransferases.
<3>Curr. Microbiol.
<4>40
<5>114-118
<6>2000
<7>The nucleotide sequence of a 2837-base pairs (bp) EcoRI-PvuI fragment of Bacillus
stearothermophilus LV chromosomal DNA encoding the bstLVIM gene was determined. It revealed a
large open reading frame (ORF) of 1737 bp specifying a methylase of 579 amino acid (aa)
residues and Mr 66,831. This was in agreement with the size estimated for the M. BstLVI (
approximately 67 kDa) purified from Escherichia coli cells harboring a recombinant plasmid
containing the bstLVIM gene and with results of transcription-translation experiments
performed in vitro. Upstream the bstLVIM gene and in the opposite transcriptional orientation,
there is an 81-aa ORF that showed great homology with the regulatory C proteins identified in
other type II restriction and modification (R-M) systems. This 81-aa ORF precedes a truncated
ORF of 86 aa which in turn may represent the structural gene for the BstLVI restriction
endonuclease.

<>

<1>Vasu, K., Nagamalleswari, E., Nagaraja, V.
<2>Promiscuous restriction is a cellular defense strategy that confers fitness advantage to bacteria.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>109
<5>E1287-1293
<6>2012
<7>Most bacterial genomes harbor restriction-modification systems, encoding a REase  and its
cognate MTase. On attack by a foreign DNA, the REase recognizes it as
nonself and subjects it to restriction. Should REases be highly specific for
targeting the invading foreign DNA? It is often considered to be the case.
However, when bacteria harboring a promiscuous or high-fidelity variant of the
REase were challenged with bacteriophages, fitness was maximal under conditions
of catalytic promiscuity. We also delineate possible mechanisms by which the
REase recognizes the chromosome as self at the noncanonical sites, thereby
preventing lethal dsDNA breaks. This study provides a fundamental understanding
of how bacteria exploit an existing defense system to gain fitness advantage
during a host-parasite coevolutionary 'arms race.'

<>

<1>Vasu, K., Nagamalleswari, E., Zahran, M., Imhof, P., Xu, S.Y., Zhu, Z., Chan, S.H., Nagaraja, V.
<2>Increasing cleavage specificity and activity of restriction endonuclease KpnI.
<3>Nucleic Acids Res.
<4>41
<5>9812-9824
<6>2013
<7>Restriction enzyme KpnI is a HNH superfamily endonuclease requiring divalent metal ions for
DNA cleavage but not for binding. The active site of KpnI can accommodate metal ions of
different atomic radii for DNA cleavage. Although Mg2+ ion higher than 500 muM mediates
promiscuous activity, Ca2+ suppresses the promiscuity and induces high cleavage fidelity.
Here, we report that a conservative mutation of the metal-coordinating residue D148 to Glu
results in the elimination of the Ca2+-mediated cleavage but imparting high cleavage fidelity
with Mg2+. High cleavage fidelity of the mutant D148E is achieved through better
discrimination of the target site at the binding and cleavage steps. Biochemical experiments
and molecular dynamics simulations suggest that the mutation inhibits Ca2+-mediated cleavage
activity by altering the geometry of the Ca2+-bound HNH active site. Although the D148E mutant
reduces the specific activity of the enzyme, we identified a suppressor mutation that
increases the turnover rate to restore the specific activity of the high fidelity mutant to
the wild-type level. Our results show that active site plasticity in coordinating different
metal ions is related to KpnI promiscuous activity, and tinkering the metal ion coordination
is a plausible way to reduce promiscuous activity of metalloenzymes.

<>

<1>Vasu, K., Nagaraja, V.
<2>Diverse Functions of Restriction-Modification Systems in Addition to Cellular Defense.
<3>Microbiol. Mol. Biol. Rev.
<4>77
<5>53-72
<6>2013
<7>Restriction-modification (R-M) systems are ubiquitous and are often considered primitive
immune systems in bacteria. Their diversity and prevalence across the prokaryotic kingdom are
an indication of their success as a defense mechanism against invading genomes. However, their
cellular defense function does not adequately explain the basis for their immaculate
specificity in sequence recognition and nonuniform distribution, ranging from none to too
many, in diverse species. The present review deals with new developments which provide
insights into the roles of these enzymes in other aspects of cellular function. In this
review, emphasis is placed on novel hypotheses and various findings that have not yet been
dealt with in a critical review. Emerging studies indicate their role in various cellular
processes other than host defense, virulence, and even controlling the rate of evolution of
the organism. We also discuss how R-M systems could have successfully evolved and be involved
in additional cellular portfolios, thereby increasing the relative fitness of their hosts in
the population.

<>

<1>Vasu, K., Saravanan, M., Bujnicki, J.M., Nagaraja, V.
<2>Structural integrity of the Beta Beta Alpha-Metal finger motif is required for DNA binding and stable protein-DNA complex formation in R.KpnI.
<3>Biochim. Biophys. Acta
<4>1784
<5>269-275
<6>2008
<7>Restriction endonuclease (REase) R.KpnI from Klebsiella pneumoniae is a homodimeric enzyme,
which recognizes palindromic sequence GGTAC|C and
cleaves generating 4 base 3' end overhangs. R.KpnI belongs to the HNH
superfamily of nucleases, which are characterized by the presence of the
beta beta alpha-Me finger motif. Structurally, this motif consists of a
twisted beta-hairpin followed by an alpha-helix, and serves as a scaffold
for side chains of residues involved in co-ordination of a divalent metal
ion that is required for catalysis. Homology modeling studies of R.KpnI
suggested a crossover structure for the alpha-helix, which could possibly
form dimeric interface and/or structural scaffold for the active site. We
have evaluated the role of the residues present in this alpha-helix in
intersubunit interactions and/or stabilization of the active site. We show
here that mutations of residues in the alpha-helix lead to a loss of the
enzyme activity, but not dimerization ability. Intrinsic fluorescence and
circular dichroism studies revealed that the loss of function phenotype
was due to the structural perturbation of the beta beta alpha-Me finger
motif. The results of mutational analysis suggest that the alpha-helix of
the beta beta alpha-Me finger of R.KpnI plays an important role for the
stability of the protein-DNA complex.

<>

<1>Vasu, K., Saravanan, M., Rajendra, B.V.R.N., Nagaraja, V.
<2>Generation of a Manganese Specific Restriction Endonuclease with Nicking Activity.
<3>Biochemistry
<4>49
<5>8425-8433
<6>2010
<7>A typical feature of type II restriction endonucleases (REases) is their obligate sequence
specificity and requirement for Mg2+ during
catalysis. R.KpnI is an exception. Unlike most other type II REases,
the active site of this enzyme can accommodate Mg2+, Mn2+, Ca2+, or
Zn2+ and cleave DNA. The enzyme belongs to the HNH superfamily of
nucleases and is characterized by the presence of a beta beta alpha-Me
finger motif. Residues D148, H149, and Q175 together form the HNH
active site and are essential for Mg2+ binding and catalysis. The
unique ability of the enzyme to cleave DNA in the presence of different
metal ions is exploited to generate mutants that are specific to one
particular metal ion. We describe the generation of a Mn2+-dependent
sequence specific endonuclease, defective in DNA cleavage with Mg2+ and
other divalent metal ions. In the engineered mutant, only Mn2+ is
selectively bound at the active site, imparting Mn2+-mediated cleavage.
The mutant is impaired in concerted double-stranded DNA cleavage,
leading to accumulation of nicked intermediates. The nicking activity
of the mutant enzyme is further enhanced by altered reaction
conditions. The active site fluidity of R Eases allowing flexible
accommodation of catalytic cofactors thus forms a basis for engineering
selective metal ion-dependent REase additionally possessing nicking
activity.

<>

<1>Vasudev, V., Obe, G.
<2>Evidence for a receptor-mediated endocytosis of AluI in chinese hamster ovary cells.
<3>Mutat. Res.
<4>197
<5>109-116
<6>1988
<7>Pretreatment of Chinese hamster ovary cells with proteases or with NaN3 leads to less
chromosomal aberrations when the cells are posttreated with AluI compared to the treatment of
cells with AluI alone.  The same result is obtained when the cells are treated with AluI at
0oC instead of 37oC.  The cells recover from the protease treatment when they are kept in
medium before treatment with AluI.  These results are interpreted to mean that AluI is bound
by surface receptors and that the AluI-receptor complexes are internalized by an
energy-dependent endocytotic process.

<>

<1>Vaughn, J.C., Mason, M.T., Sper-Whitis, G.L., Kuhlman, P., Palmer, J.D.
<2>Fungal origin by horizontal transfer of a plant mitochondrial group I intron in the chimeric CoxI gene of Peperomia.
<3>J. Mol. Evol.
<4>41
<5>563-572
<6>1995
<7>We present phylogenetic evidence that a group I intron in an angiosperm mitochondrial gene
arose recently by horizontal transfer from a fungal donor species.  A 1,716-bp fragment of the
mitochondrial coxI gene from the angiosperm Peperomia polybotrya was amplified via the
polymerase chain reaction and sequenced.  Comparison to other coxI genes revealed a 966-bp
group I intron, which, based on homology with the related yeast coxI intron aI4, potentially
encodes a 279-amino-acid site-specific DNA endonuclease.  This intron, which is believed to
function as a ribozyme during its own splicing, is not present in any of 19 coxI genes
examined from other diverse vascular plant species.  Phylogenetic analysis of intron origin
was carried out using three different tree-generating algorithms, and on a variety of
nucleotide and amino acid data sets from the intron and its flanking exon sequences.  These
analyses show that the Peperomia coxI gene intron and exon sequences are of fundamentally
different evolutionary origin. The Peperomia intron is more closely related to several fungal
mitochondrial introns, two of which are located at identical positions in coxI, than to
identically located coxI introns from the land plant Marchantia and the green alga Prototheca.
Conversely, the exon sequence of this gene is, as expected, most closely related to other
angiosperm coxI genes.  These results, together with evidence suggestive of co-conversion of
exonic markers immediately flanking the intron insertion site, lead us to conclude that the
Peperomia coxI intron probably arose by horizontal transfer from a fungal donor, using the
double-strand-break repair pathway.  The donor species may have been one of the symbiotic
mycorrhizal fungi that live in close obligate association with most plants.

<>

<1>Vazquez, R., Brito, J., Guerra, M.
<2>Purification of the restriction endonuclease SacI, free of SacII and SacIII contaminants.
<3>Biotecnol. Apl.
<4>13
<5>21-24
<6>1996
<7>The restriction endonuclease SacI was isolated from Streptomyces achromogenes and was purified
to homogeneity, until no contaminant nuclease activities were detected.  On the basis of ion
exchange chromatography (Q-Sepharose and phosphocellulose P-11), SacI can be obtained with a
high level of purity and used in molecular cloning.  The practical utility of SacI enzyme is
given by a high stability, high yield, easy handling of producing cells, and the ability to
recognize new sequences, such as GAGCTC.  The molecular weight (MW) of this enzyme was
estimated by High Performance Liquid Chromatography and SDS polyacrylamide gel
electrophoresis, being of about 50 kDa approximately.  According to the results obtained from
the accelerated stability study, the enzyme preparation is stable for at least 20 months.

<>

<1>Vazquez, R., Martinez, D., Reyes, G., Marquez, G., Ryes, N., Diaz, Y., Luis, M., Gonzalez, B., Gonzalez, N.
<2>Purification of NcoI restriction endonuclease free of exonucleases and contaminant endonucleases.
<3>Biotecnol. Apl.
<4>13
<5>289-290
<6>1996
<7>
<>

<1>Vecherkovskaya, M.F., Tetz, G.V., Tetz, V.V.
<2>Complete Genome Sequence of the Streptococcus sp. Strain VT 162, Isolated from the Saliva of Pediatric Oncohematology Patients.
<3>Genome Announcements
<4>2
<5>e00647-14
<6>2014
<7>Streptococcus sp. strain VT 162 was isolated from the saliva of pediatric oncohematology
patients. Its full genome is 2,045,418 bp. The availability of
this genome will provide insights into the composition of microbial flora among
pediatric oncohematology patients and the host interaction and pathogenicity of
this species.

<>

<1>Vedantam, G., Hecht, D.W.
<2>Isolation and characterization of BTF-37: Chromosomal DNA captured from Bacteroides fragilis that confers self-transferability and expresses a pilus-like structure in Bacteroides spp. and Escherichia coli.
<3>J. Bacteriol.
<4>184
<5>728-738
<6>2002
<7>We report the isolation and preliminary characterization of BTF-37, a new 52-kb transfer
factor isolated from Bacteroides fragilis clinical isolate LV23. BTF-37 was obtained by the
capture of new DNA in the nonmobilizable Bacteroides-Escherichia coli shuttle vector
pGAT400DeltaBglII using a functional assay. BTF-37 is self-transferable within and from
Bacteroides and also self-transfers in E. coli. Partial DNA sequencing, colony hybridization,
and PCR revealed the presence of Tet element-specific sequences in BTF-37. In addition,
Tn5520, a small mobilizable transposon that we described previously (G. Vedantam, T. J.
Novicki, and D. W. Hecht, J. Bacteriol. 181:2564-2571, 1999), was also coisolated within
BTF-37. Scanning and transmission electron microscopy of Tet element-containing Bacteroides
spp. and BTF-37-harboring Bacteroides and E. coli strains revealed the presence of pilus-like
cell surface structures. These structures were visualized in Bacteroides spp. only when BTF-37
and Tet element strains were induced with subinhibitory concentrations of tetracycline and
resembled those encoded by E. coli broad-host-range plasmids. We conclude that we have
captured a new, self-transferable transfer factor from B. fragilis LV23 and that this new
factor encodes a tetracycline-inducible Bacteroides sp. conjugation apparatus.

<>

<1>Vedler, E., Vahter, M., Heinaru, A.
<2>The Completely Sequenced Plasmid pEST4011 Contains a Novel IncP1 Backbone and a Catabolic Transposon Harboring tfd Genes for 2,4-Dichlorophenoxyacetic Acid Degradation.
<3>J. Bacteriol.
<4>186
<5>7161-7174
<6>2004
<7>The herbicide 2,4-dichlorophenoxyacetic acid (2,4-D)-degrading bacterium
Achromobacter xylosoxidans subsp. denitrificans strain EST4002 contains
plasmid pEST4011. This plasmid ensures its host a stable 2,4-D(+)
phenotype. We determined the complete 76,958-bp nucleotide sequence of
pEST4011. This plasmid is a deletion and duplication derivative of pD2M4,
the 95-kb highly unstable laboratory ancestor of pEST4011, and was
self-generated during different laboratory manipulations performed to
increase the stability of the 2,4-D(+) phenotype of the original strain,
strain D2M4(pD2M4). The 47,935-bp catabolic region of pEST4011 forms a
transposon-like structure with identical copies of the hybrid insertion
element IS1071::IS1471 at the two ends. The catabolic regions of pEST4011
and pJP4, the best-studied 2,4-D-degradative plasmid, both contain
homologous, tfd-like genes for complete 2,4-D degradation, but they have
little sequence similarity other than that. The backbone genes of pEST4011
are most similar to the corresponding genes of broad-host-range
self-transmissible IncP1 plasmids. The backbones of the other three IncP1
catabolic plasmids that have been sequenced (the 2,4-D-degradative plasmid
pJP4, the haloacetate-catabolic plasmid pUO1, and the atrazine-catabolic
plasmid pADP-1) are nearly identical to the backbone of R751, the
archetype plasmid of the IncP1 beta subgroup. We show that despite the
overall similarity in plasmid organization, the pEST4011 backbone is
sufficiently different (51 to 86% amino acid sequence identity between
individual backbone genes) from the backbones of members of the three
IncP1 subgroups (the alpha, beta, and gamma subgroups) that it belongs to
a new IncP1subgroup, the delta subgroup. This conclusion was also
supported by a phylogenetic analysis of the trfA2, korA, and traG gene
products of different IncP1 plasmids.

<>

<1>Veiga, H., Pinho, M.G.
<2>Inactivation of the SauI Type I Restriction-Modification System Is Not Sufficient To Generate Staphylococcus aureus Strains Capable of Efficiently Accepting Foreign DNA.
<3>Appl. Environ. Microbiol.
<4>75
<5>3034-3038
<6>2009
<7>Genetic manipulation of Staphylococcus aureus is limited by the availability of only a single
strain, RN4220, that is capable of easily
accepting foreign DNA. Inactivation of the hsdR gene of the SauI type I
restriction-modification system was shown previously to be responsible
for the high transformation efficiency of RN4220 (D. E. Waldron and J.
A. Lindsay, J Bacteriol. 188:5578-5585, 2006). However, deletion of
this gene in three different S. aureus strains was not sufficient to
make them readily transformable, which would be remarkably useful for
genetic studies of this pathogenic organism. These results indicate
that another unknown factor(s) is required for the transformable
phenotype in S. aureus.

<>

<1>Veith, B., Herzberg, C., Steckel, S., Feesche, J., Maurer, K.H., Ehrenreich, P., Baumer, S., Henne, A., Liesegang, H., Merkl, R., Ehrenreich, A., Gottschalk, G.
<2>The complete genome sequence of Bacillus licheniformis DSM13, an organism with great industrial potential.
<3>J. Mol. Microbiol. Biotechnol.
<4>7
<5>204-211
<6>2004
<7>The genome of Bacillus licheniformis DSM13 consists of a single chromosome that has a size of
4,222,748 base pairs. The average G+C ratio is 46.2%. 4,286 open
reading frames, 72 tRNA genes, 7 rRNA operons and 20 transposase genes were
identified. The genome shows a marked co-linearity with Bacillus subtilis but
contains defined inserted regions that can be identified at the sequence as well
as at the functional level. B. licheniformis DSM13 has a well-conserved secretory
system, no polyketide biosynthesis, but is able to form the lipopeptide
lichenysin. From the further analysis of the genome sequence, we identified
conserved regulatory DNA motives, the occurrence of the glyoxylate bypass and the
presence of anaerobic ribonucleotide reductase explaining that B. licheniformis
is able to grow on acetate and 2,3-butanediol as well as anaerobically on
glucose. Many new genes of potential interest for biotechnological applications
were found in B. licheniformis; candidates include proteases, pectate lyases,
lipases and various polysaccharide degrading enzymes.

<>

<1>Veitinger, S., Schmitz, G.G., Kaluza, K., Jarsch, M., Braun, V., Kessler, C.
<2>SfuI, a novel AsuII isoschizomer from Streptomyces fulvissimus recognizing 5'-TT/CGAA-3'.
<3>Nucleic Acids Res.
<4>18
<5>3424
<6>1990
<7>None

<>

<1>Velkov, V.V.
<2>How environmental factors regulate mutagenesis and gene transfer in microorganisms.
<3>J. Biosci.
<4>24
<5>529-559
<6>1999
<7>This review is focused on the physiological and evolutionary strategies of the processes
occurring during the entry of microbial cells into
stationary phase and the subsequent period of stasis. The molecular
mechanisms adapting microorganisms from exponential growth to a static
state involve activation and complex regulation of the stationary
factor Sigma-S, which directs RNA polymerase to the specific promoters.
As a result the static cells acquire general resistance (simultaneous
tolerances) to different environmental stresses. In parallel with the
physiological adaptation to stasis, diverse genetical processes are
aimed towards resuming the growth of the static cells. Different types
of mutagenesis occur: (i) in cells entering stasis and (ii) in static
cells (adaptive mutagenesis). Cessation of growth induces the transient
hypermutator state resulting in the accumulation of random mutations in
the subpopulation of the static cells. If by chance, one or a few of
such mutations lead to resumption of division, the growing cell will
return to a normal mechanism of spontaneous mutagenesis. Another
mechanism for generating genetical variability in stressed cells
involves transposons and conjugative plasmids. Stresses can stimulate
the excision of some transposons, which, in turn, can generate
chromosomal mutations and activate intracellular mechanisms of
mutagenesis. Under stress some conjugative plasmids activate genes
encoding antirestriction proteins that repress restriction-modification
systems of the recipient cells. Moreover, under stress special cellular
mechanisms decrease (alleviate) the activity of
restriction-modification systems which, in turn, enhance the
probability of gene transfer into the stressed cells. Under stress, the
efficiency of inter-species genetical barriers also decreases. This,
stimulates inter-species gene transfer and may lead to a burst of
incipient speciation in the population of non-growing cells. After
resumption of growth the genetical barriers leading to isolation will
be restored. In general, the cessation of growth "switches on", and
resumption of growth "switches off", a set of special processes that
are responsible for generating bursts of genetical variability in
populations of microorganisms.

<>

<1>Vellarikkal, S.K., Singh, A.V., Singh, P.K., Garg, P., Katoch, V.M., Katoch, K., Chauhan, D.S., Sivasubbu, S., Scaria, V.
<2>Draft Genome Sequence of a Multidrug-Resistant Clinical Isolate of Mycobacterium  tuberculosis Belonging to a Novel Spoligotype.
<3>Genome Announcements
<4>1
<5>e00965-13
<6>2013
<7>We describe the genome sequencing and analysis of a multidrug-resistant (MDR) clinical isolate
of Mycobacterium tuberculosis, strain OSDD105 from India,
belonging to a novel spoligotype.

<>

<1>Vellarikkal, S.K., Singh, A.V., Singh, P.K., Garg, P., Katoch, V.M., Katoch, K., Chauhan, D.S., Sivasubbu, S., Scaria, V.
<2>Draft Genome Sequence of Multidrug-Resistant Mycobacterium tuberculosis Clinical  Isolate OSDD515, Belonging to the Uganda I Genotype.
<3>Genome Announcements
<4>1
<5>e00750-13
<6>2013
<7>We describe the genome sequencing and analysis of a clinical isolate of the
multidrug-resistant Mycobacterium tuberculosis Uganda I genotype (OSDD515) from
India.

<>

<1>Velly, H., Renault, P., Abraham, A.L., Loux, V., Delacroix-Buchet, A., Fonseca, F., Bouix, M.
<2>Genome Sequence of the Lactic Acid Bacterium Lactococcus lactis subsp. lactis TOMSC161, Isolated from a Nonscalded Curd Pressed Cheese.
<3>Genome Announcements
<4>2
<5>e01121-14
<6>2014
<7>Lactococcus lactis is a lactic acid bacterium used in the production of many fermented foods,
such as dairy products. Here, we report the genome sequence of
L. lactis subsp. lactis TOMSC161, isolated from nonscalded curd pressed cheese.
This genome sequence provides information in relation to dairy environment
adaptation.

<>

<1>Velusamy, N., Prakash, L., Sivakumar, N., Antony, A., Prajna, L., Mohankumar, V., Devarajan, B.
<2>Draft Genome Sequences of Staphylococcus aureus AMRF1 (ST22) and AMRF2 (ST672), Ocular Methicillin-Resistant Isolates.
<3>Genome Announcements
<4>2
<5>e00168-14
<6>2014
<7>Sequence type 22 (ST22) and ST672 are the two major emerging clones of community-acquired
methicillin-resistant Staphylococcus aureus in India. ST672
strains were found to cause severe ocular infections. We report the draft genome
sequences of two emerging strains of methicillin-resistant S. aureus, AMRF1
(ST22) and AMRF2 (ST672), isolated from patients with ocular infections.

<>

<1>Venclovas, C., Siksnys, V.
<2>Different enzymes with similar structures involved in Mg2+-mediated polynucleotidyl transfer.
<3>Nat. Struct. Biol.
<4>2
<5>838-841
<6>1995
<7>Comparison of x-ray structures of restriction endonucleases and polynucleotidyl transferase
superfamily enzymes reveals a structural resemblance.  Transfer of polynucleotidyl residues
plays a fundamental role in such critical cellular events as genetic recombination,
transposition and viral DNA integration.  A number of different enzymes is directly implicated
in this process.  Recently X-ray structures of RuvC protein, a Holliday junction resolvase
from Escherichia coli and HIV-1 integrase, involved in viral DNA integration, have been
solved.  Surprisingly, RuvC, HIV-integrase and ribonuclease H (RNase H) appear to have similar
three-dimensional structures, despite the lack of protein sequence similarities.  On the
basis of fold resemblance it was proposed that all these enzymes belong to the new superfamily
of polynucleotidyl transferases (PNT).

<>

<1>Venclovas, C., Timinskas, A., Siksnys, V.
<2>Five-stranded beta-sheet sandwiched with two a-helices: a structural link between restriction endonucleases EcoRI and EcoRV.
<3>Proteins
<4>20
<5>279-282
<6>1994
<7>Examination of crystal structures of restriction endonucleases EcoRI and EcoRV complexes with
their cognate DNA revealed a common structural element, which forms the core of both proteins.
This element consists of a five-stranded beta-sheet and two alpha-helices packed against it
and could be described as alpha-beta sandwich in which helices and beta-strands lie in two
stacked layers. While the spatial structure of this alpha-beta sandwich is conserved in both
enzymes, there are no detectable similarities between amino acid sequences except of a few
residues involved in active site formation. Probably, other restriction endonucleases which
have similar organization of the active site might possess similar structural element
regardless of DNA sequence recognized and recognition elements in the enzyme used.

<>

<1>Venditti, S., Wells, R.D.
<2>A DNA conformational alteration induced by a neighboring oligopurine tract on GAATTA enables nicking by EcoRI.
<3>J. Biol. Chem.
<4>266
<5>16786-16790
<6>1991
<7>The pseudo EcoRI site GAATTA in the U3 region of the long terminal repeat of
human immunodeficiency virus, which is flanked by a 26-base pair oligopurine
tract, is readily nicked by either EcoRI or RsrI.  The strand-specific nick
occurs predominantly between the G and A residues and is independent of
negative supercoiling.  Other GAATTA sites surrounded by random
(non-oligopurine) sequences are not nicked by these restriction endonucleases.
However, other types and lengths of oligopurine tracts are effective in
inducing the nicking in neighboring GAATTA sites.  Hence, we propose that the
flanking oligopurine tracts induce an altered DNA conformation on the GAATTA
target site which may be similar to the transition state induced by EcoRI when
binding to its canonical recognition site.  Gel retardation analyses on
restriction fragments containing the oligopurine-GAATTA-oligopurine sequences
suggest the presence of helical axis distortions which are consistent with this
interpretation.

<>

<1>Vendramin, V., Treu, L., Bovo, B., Campanaro, S., Corich, V., Giacomini, A.
<2>Whole-Genome Sequence of Streptococcus macedonicus Strain 33MO, Isolated from the Curd of Morlacco Cheese in the Veneto Region (Italy).
<3>Genome Announcements
<4>2
<5>e00746-14
<6>2014
<7>A genetic characterization of Streptococcus macedonicus is important to better understand the
characteristics of this lactic acid bacterium, frequently detected
in fermented food bacteria communities. This report presents the draft genome
sequence description of strain 33MO, the first publicly available genome sequence
of an Italian S. macedonicus isolate.

<>

<1>Venegas, A., Motles, M., Vasquez, C., Vicuna, R.
<2>Conditions affecting DNA cleavage by TthI at a TthI endonuclease-dam methylase overlapping sequence.
<3>FEBS Lett.
<4>130
<5>272-274
<6>1981
<7>The Escherichia coli dam gene codes for a site-specific DNA methylase which methylates adenine
in the sequence GATC at the N6 position of the purine ring. The effect of this modification on
the action of several restriction enzymes whose recognition sites are equal or include the
methylated sequence depends on the endonuclease itself. For example, the enzymes MboI, BclI
and DpnII do not cleave a sequence containing GAme^TC, while DpnI requires the presence of
6MeAde in GATC sites in order to cleave the DNA. On the other hand, isoschizomers Sau3AI,
FnuEI and PfaI actively hydrolyze sequences that have undergone adenine methylation; Sau3AI,
however, is inhibited by the methylation of cytosine of the indicated sequence.

<>

<1>Venegas, A., Vicuna, R., Alonso, A., Valdes, F., Yudelevich, A.
<2>A rapid procedure for purifying a restriction endonuclease from Thermus thermophilus (TthI).
<3>FEBS Lett.
<4>109
<5>156-158
<6>1980
<7>Restriction enzymes have proved to be a powerful tool for mapping genomes and
developing molecular cloning and DNA sequencing techniques.  A few restriction
enzymes have been isolated from thermophilic bacteria, showing a high
thermostability and resistance to protein denaturing agents.  These properties
could be useful to study DNA structure at higher temperatures.  Looking for a
stable enzyme with a new recognition sequence, we have isolated TthI, an enzyme
from the extreme thermophile Thermus thermophilus HB8. This thermostable enzyme
turned out to be an isoschizomer of TaqI, which recognizes the sequence
5'-TCGA-3'.  Thermus thermophilus is a very convenient source for purifying
this enzyme since this bacterium does not produce the pigmented 'slime'
described in Thermus aquaticus, which interferes with the Purification of TawI
and other enzymes.  Furthermore, TthI was readily released by osmotic shock,
which allowed us to develop a simplified, rapid two-step procedure to obtain an
enzyme preparation free of contaminating nucleases.  We report here the
purifiction method and some of the properties of TthI.

<>

<1>Venetianer, P.
<2>Purification and properties of the Bsp endonuclease.
<3>Methods Enzymol.
<4>65
<5>109-112
<6>1980
<7>None

<>

<1>Venetianer, P., Kiss, A.
<2>The restriction-modification enzymes of Bacillus sphaericus R.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>209-215
<6>1981
<7>*
  I. Biochemical characterization of the BspI restriction endonuclease
 II. Biochemical characterization of the BspI modification methylase
III. The genes of the Bsp restriction-modification system
 IV. Conclusion


<>

<1>Venetianer, P., Komocsin, M.
<2>Production of a new restriction endonuclease, BflI, from Bacillus cereus.
<3>Hungarian Patent Office
<4>HU 50492
<5>
<6>1987
<7>
<>

<1>Venetianer, P., Komocsin, M., Orosz, A.
<2>The method for producing BepI restriction endonuclease from Brevibacterium epidermidis.
<3>Hungarian Patent Office
<4>HU 29160
<5>
<6>1990
<7>
<>

<1>Venetianer, P., Orosz, A.
<2>BcefI, a new type IIS restriction endonuclease.
<3>Nucleic Acids Res.
<4>16
<5>3053-3060
<6>1988
<7>A new Type IIS restriction endonuclease was identified, partially purified and characterized
from a Bacillus cereus subsp. fluorescens strain.  The enzyme recognized the nonpalindromic
sequence ACGGC and cleaves at a distance from it.  The cleavage appears to occur with a +/-1
basepair uncertainity.  Thus the cleavage and recognition site is as shown below:
ACGGC(N) 11-13
TGCCG(N) 12-14.

<>

<1>Venetianer, P., Orosz, A.
<2>Isolation and characterization of two new restriction endonucleases (BepI and BcefI) .
<3>Gene
<4>74
<5>99
<6>1988
<7>Meeting Abstract

<>

<1>Venetianer, P., Orosz, A.
<2>BepI restriction endonuclease, a new isoschizomer of FnuDII.
<3>Nucleic Acids Res.
<4>16
<5>350
<6>1988
<7>A new TypeII restriction endonuclease was isolated and characterized from a bacterial strain
identified as Brevibacterium epidermidis by Dr. M. Konkoly-Thege (Budapest).  The strain grows
well in yeast-tryptone broth aerobically with a doubling time of 50 min at 25-30 C.  The
enzyme was purified by a conventional procedure, employing Bio-Gel A-0.5 m, DEAE-cellulose and
phosphocellulose chromatography.  The enzyme at this stage of purification was applicable for
all practical purposes, it was virtually free from contaminating nucleases.  The yield of the
enzyme was approximately 3-400 units/g wet weight of cell.  The optimal conditions for the
activity of the enzyme are as follows:  temperature 25-30 C., Mg2+ ions 10 mM, pH 7.4, no
salts.  The cleavage pattern of the enzyme tested with known DNA molecules (lambda phage,
pBR322) was identical with the pattern obtained with FnuDII.  This identity was confirmed by
parallel digestions with the new enzyme and commercial FnuDII, and double digestions with both
enzymes, that resulted in unaltered digestion patterns. The cleavage site was established by
running a Maxam-Gilbert sequence gel from a DNA fragment of known sequence, which contained a
FnuDII site.  The same end-labelled DNA fragment was then digested with the new enzyme, and it
was electrophoresed alongside the sequence ladder.  This experiment unambiguously proved that
the cleavage occured in the middle of the palindromic CGCG sequence, generating flush ends.
Thus the enzyme does not represent a new specificity, it is an isoschizomer of the well known
FnuDII enzyme. Nevertheless it offers some practical advantages.  The Fusobacterium nucleatum
strain, from which FnuDII is isolated, is an obligately anaerobic pathogenic microorganism,
therefore FnuDII is fairly expensive.  The only other commercially available isoschisomer,
ThaI is purified from an extremely thermophilic and acidophilic organism, the other described
isoschisomers (3,4) have not been characterized.  It is hoped that the BepI enzyme will be a
better substitute for these isoschisomers.

<>

<1>Venkatesan, M.M., Goldberg, M.B., Rose, D.J., Grotbeck, E.J., Burland, V., Blattner, F.R.
<2>Complete DNA sequence and analysis of the large virulence plasmid of Shigella flexneri.
<3>Infect. Immun.
<4>69
<5>3271-3285
<6>2001
<7>The complete sequence analysis of the 210-kb Shigella flexneri 5a
virulence plasmid was determined. Shigella spp. cause dysentery and
diarrhea by invasion and spread through the colonic mucosa. Most of the
known Shigella virulence determinants are encoded on a large plasmid that
is unique to virulent strains of Shigella and enteroinvasive Escherichia
coli; these known genes account for approximately 30 to 35% of the
virulence plasmid. In the complete sequence of the virulence plasmid, 286
open reading frames (ORFs) were identified. An astonishing 153 (53%) of
these were related to known and putative insertion sequence (IS) elements;
no known bacterial plasmid has previously been described with such a high
proportion of IS elements. Four new IS elements were identified. Fifty
putative proteins show no significant homology to proteins of known
function; of these, 18 have a G+C content of less than 40%, typical of
known virulence genes on the plasmid. These 18 constitute potentially
unknown virulence genes. Two alleles of shet2 and five alleles of ipaH
were also identified on the plasmid. Thus, the plasmid sequence suggests a
remarkable history of IS-mediated acquisition of DNA across bacterial
species. The complete sequence will permit targeted characterization of
potential new Shigella virulence determinants.

<>

<1>Venkatramanan, R., Prakash, O., Woyke, T., Chain, P., Goodwin, L.A., Watson, D., Brooks, S., Kostka, J.E., Green, S.J.
<2>Genome sequences for three denitrifying bacterial strains isolated from a uranium- and nitrate-contaminated subsurface environment.
<3>Genome Announcements
<4>1
<5>e00449-13
<6>2013
<7>Genome sequences for three strains of denitrifying bacteria (Alphaproteobacteria-Afipia sp.
strain 1NLS2 and Hyphomicrobium denitrificans
strain 1NES1; Firmicutes-Bacillus sp. strain 1NLA3E) isolated from the nitrate-
and uranium-contaminated subsurface of the Oak Ridge Integrated Field Research
Challenge (ORIFRC) site, Oak Ridge Reservation, TN, are reported.

<>

<1>Vente, A., Piepersberg, W., Wehmeier, U., Schmidt-Beissner, H., Aboshanab, K., Mohammed, A., Welzel, K.
<2>Novel genes from a biosynthesis gene cluster for aminoglycoside antibiotic synthesis and the thus obtained novel aminoglycoside antibiotics.
<3>International Patent Office
<4>WO 2005095591 A
<5>
<6>2005
<7>The invention relates to novel genes obtainable from aminoglycoside-producing microorganisms,
to novel gene clusters formed from the novel and/or known genes of said microorganisms, to the
expression products of the genes, the transport vehicles thereof or of the gene clusters and
to transformed cells containing said genes or gene clusters.  Said invention also relates to
methods for biologically producing aminoglycosides, methods for combinatory synthesis of
aminoglycoside derivatives, to novel aminoglycosides, to the use thereof and to pharmaceutical
compositions containing said aminoglycosides.

<>

<1>Venter, J.C. et al.
<2>Environmental Genome Shotgun Sequencing of the Sargasso Sea.
<3>Science
<4>304
<5>66-74
<6>2004
<7>We have applied "whole-genome shotgun sequencing" to microbial populations
collected en masse on tangential flow and impact filters from seawater
samples collected from the Sargasso Sea near Bermuda. A total of 1.045
billion base pairs of nonredundant sequence was generated, annotated, and
analyzed to elucidate the gene content, diversity, and relative abundance
of the organisms within these environmental samples. These data are
estimated to derive from at least 1800 genomic species based on sequence
relatedness, including 148 previously unknown bacterial phylotypes. We
have identified over 1.2 million previously unknown genes represented in
these samples, including more than 782 new rhodopsin-like photoreceptors.
Variation in species present and stoichiometry suggests substantial
oceanic microbial diversity.

<>

<1>Venter, J.C., Tetterin, H., Fraser, C., Grandi, G., Galeotti, C., Scarlato, E., Mora, M., Petersen, J., Pizza, M., Rappuoli, R., Ratti, G., Scarselli, M., Masignani, V., Hickey, E.
<2>Neisseria Meningitidis Antigens and Compositions.
<3>Japanese Patent Office
<4>JP 2004500801 A
<5>
<6>2004
<7>
<>

<1>Ventura, M. et al.
<2>The Bifidobacterium dentium Bd1 genome sequence reflects its genetic adaptation to the human oral cavity.
<3>PLoS Genet.
<4>5
<5>e1000785
<6>2009
<7>Bifidobacteria, one of the relatively dominant components of the human intestinal microbiota,
are considered one of the key groups of beneficial
intestinal bacteria (probiotic bacteria). However, in addition to
health-promoting taxa, the genus Bifidobacterium also includes
Bifidobacterium dentium, an opportunistic cariogenic pathogen. The genetic
basis for the ability of B. dentium to survive in the oral cavity and
contribute to caries development is not understood. The genome of B.
dentium Bd1, a strain isolated from dental caries, was sequenced to
completion to uncover a single circular 2,636,368 base pair chromosome
with 2,143 predicted open reading frames. Annotation of the genome
sequence revealed multiple ways in which B. dentium has adapted to the
oral environment through specialized nutrient acquisition, defences
against antimicrobials, and gene products that increase fitness and
competitiveness within the oral niche. B. dentium Bd1 was shown to
metabolize a wide variety of carbohydrates, consistent with genome-based
predictions, while colonization and persistence factors implicated in
tissue adhesion, acid tolerance, and the metabolism of human
saliva-derived compounds were also identified. Global transcriptome
analysis demonstrated that many of the genes encoding these predicted
traits are highly expressed under relevant physiological conditions. This
is the first report to identify, through various genomic approaches,
specific genetic adaptations of a Bifidobacterium taxon, Bifidobacterium
dentium Bd1, to a lifestyle as a cariogenic microorganism in the oral
cavity. In silico analysis and comparative genomic hybridization
experiments clearly reveal a high level of genome conservation among
various B. dentium strains. The data indicate that the genome of this
opportunistic cariogen has evolved through a very limited number of
horizontal gene acquisition events, highlighting the narrow boundaries
that separate commensals from opportunistic pathogens.

<>

<1>Venturini, C., Beatson, S.A., Djordjevic, S.P., Walker, M.J.
<2>Multiple antibiotic resistance gene recruitment onto the enterohemorrhagic Escherichia coli virulence plasmid.
<3>FASEB J.
<4>24
<5>1160-1166
<6>2010
<7>Enterohemorrhagic Escherichia coli (EHEC) strains are zoonotic pathogens
responsible for a range of severe human disease. The repertoire of
virulence determinants promoting EHEC disease is encoded on both the main
chromosome and virulence plasmid. We examined a multiply
antibiotic-resistant O26 EHEC strain for carriage of resistance genes on
the virulence plasmid. The EHEC virulence plasmid containing a complex
antibiotic-resistance gene locus, designated as pO26-CRL, was purified
from EHEC O26:H(-) (patient with hemorrhagic colitis) and subjected to
shotgun-sequencing and bioinformatic analysis. Determination of the
111,481-bp sequence of pO26-CRL revealed genes encoding a functional
enterohemolysin operon (ehxCABD), STEC-specific extracellular serine
protease (espP), putative EHEC adhesin (toxB), catalase/peroxidase (katP),
and myristoyl transferase (msbB) involved in lipid A synthesis. A
22,609-bp Tn21 derivative is inserted within the conjugal transfer gene
traC and encodes resistance to trimethoprim, streptomycin, sulfathiozole,
kanamycin, neomycin, beta-lactams, and mercuric chloride. Plasmid pO26-CRL
is nonconjugative but is mobilizable. This is the first report of an EHEC
virulence plasmid containing a complex antibiotic resistance locus, and
raises the concern that antibiotic use will coselect for virulence
determinants, leading to increased disease potential in both commensal and
pathogenic E. coli populations.-Venturini, C., Beatson, S. A., Djordjevic,
S. P., Walker, M. J. Multiple antibiotic resistance gene recruitment onto
the enterohemorrhagic Escherichia coli virulence plasmid.

<>

<1>Venturini, C., Hassan, K.A., Roy, C., P, P., I, T., Walker, M.J., Djordjevic, S.P.
<2>Sequences of Two Related Multiple Antibiotic Resistance Virulence Plasmids Sharing a Unique IS26-Related Molecular Signature Isolated from Different Escherichia coli Pathotypes from Different Hosts.
<3>PLoS ONE
<4>8
<5>E78862
<6>2013
<7>Enterohemorrhagic Escherichia coli (EHEC) and atypical enteropathogenic E. coli
(aEPEC) are important zoonotic pathogens that increasingly are becoming resistant
to multiple antibiotics. Here we describe two plasmids, pO26-CRL125 (125 kb) from
a human O26:H- EHEC, and pO111-CRL115 (115kb) from a bovine O111 aEPEC, that
impart resistance to ampicillin, kanamycin, neomycin, streptomycin,
sulfathiazole, trimethoprim and tetracycline and both contain atypical class 1
integrons with an identical IS26-mediated deletion in their 3 -conserved segment.
Complete sequence analysis showed that pO26-CRL125 and pO111-CRL115 are
essentially identical except for a 9.7 kb fragment, present in the backbone of
pO26-CRL125 but absent in pO111-CRL115, and several indels. The 9.7 kb fragment
encodes IncI-associated genes involved in plasmid stability during conjugation, a
putative transposase gene and three imperfect repeats. Contiguous sequence
identical to regions within these pO26-CRL125 imperfect repeats was identified in
pO111-CRL115 precisely where the 9.7 kb fragment is missing, suggesting it may be
mobile. Sequences shared between the plasmids include a complete IncZ replicon, a
unique toxin/antitoxin system, IncI stability and maintenance genes, a novel
putative serine protease autotransporter, and an IncI1 transfer system including
a unique shufflon. Both plasmids carry a derivate Tn21 transposon with an
atypical class 1 integron comprising a dfrA5 gene cassette encoding resistance to
trimethoprim, and 24 bp of the 3 -conserved segment followed by Tn6026, which
encodes resistance to ampicillin, kanymycin, neomycin, streptomycin and
sulfathiazole. The Tn21-derivative transposon is linked to a truncated Tn1721,
encoding resistance to tetracycline, via a region containing the IncP-1alpha
oriV. Absence of the 5 bp direct repeats flanking Tn3-family transposons,
indicates that homologous recombination events played a key role in the formation
of this complex antibiotic resistance gene locus. Comparative sequence analysis
of these closely related plasmids reveals aspects of plasmid evolution in
pathogenic E. coli from different hosts.

<>

<1>Venyaminov, S.Y., Kosykh, V.G., Kholodkov, O.A., Buryanov, Y.I.
<2>UV- and CD-spectra of restriction endonuclease EcoRII and DNA-methylase EcoRII.
<3>Bioorg. Khim.
<4>16
<5>47-51
<6>1990
<7>UV- and CD-spectra of homogeneous enzymes have been measured.  Extinction
coefficients estimated from the UV-spectra are 0.97 for restriction
endonuclease EcoRII at 279.5 nm and 1.17 for DNA-methylase EcoRII at 279 nm.
As it follows from the CD spectra, both enzymes have a well developed tertiary
structure and a highly ordered secondary structure, which consists of 22%
alpha-helices, 64% beta-structure and 9% bends for R.EcoRII and of 44%
alpha-helices, 48% beta-structure and 4% bends for M.EcoRII.  Restriction
endonuclease denatures at 50C, while DNA -methylase denatures at 45C, with
partial reversibility upon cooling.

<>

<1>Vera-Cabrera, L., Ortiz-Lopez, R., Elizondo-Gonzalez, R., Campos-Rivera, M.P., Gallardo-Rocha, A., Molina-Torres, C.A., Ocampo-Candiani, J.
<2>Draft Genome Sequence of Actinomadura madurae LIID-AJ290, Isolated from a Human Mycetoma Case.
<3>Genome Announcements
<4>2
<5>e00201-14
<6>2014
<7>Here we present the draft genome sequence of a member of the Thermomonosporaceae, Actinomadura
madurae LIID-AJ290, isolated from a human case of mycetoma. The
assembly contains 10,308,866 bp. This is to our knowledge the first reported
genome of a human-pathogenic Actinomadura species.

<>

<1>Vera-Cabrera, L., Ortiz-Lopez, R., Elizondo-Gonzalez, R., Perez-Maya, A.A., Ocampo-Candiani, J.
<2>Complete Genome Sequence of Nocardia brasiliensis HUJEG-1.
<3>J. Bacteriol.
<4>194
<5>2761-2762
<6>2012
<7>In Mexico, actinomycetoma is mainly caused by Nocardia brasiliensis, which is a soil
inhabitant actinobacterium. Here, we report for the first time the draft
genome of a strain isolated from a human case that has largely been found in in
vitro and experimental models of actinomycetoma, N. brasiliensis HUJEG-1.

<>

<1>Verdet, C., Gautier, V., Chachaty, E., Ronco, E., Hidri, N., Decre, D., Arlet, G.
<2>Genetic context of plasmid-carried blaCMY-2-like genes in Enterobacteriaceae.
<3>Antimicrob. Agents Chemother.
<4>53
<5>4002-4006
<6>2009
<7>Analysis of 15 European clinical Enterobacteriaceae isolates showed that
differences in the genetic context of blaCMY-2-like genes reflected the
replicon type, usually IncA/C or IncI1. These blaCMY-2 loci may originate
from the same ISEcp1-mediated mobilization from the Citrobacter freundii
chromosome as structures described in earlier studies.

<>

<1>Verdine, G.
<2>The flip side of DNA methylation.
<3>Cell
<4>76
<5>197-200
<6>1994
<7>DNA methylation has come of age. For more than 40 years, it has been known that eukaryotes tag
their DNA by the covalent addition of a methyl group to cytosine residues; however, until very
recently, the functional significance of this modification has remained on a precariously
speculative footing. A series of discoveries over the last few years has thrust DNA
methylation firmly into the mainstream of biology and medicine, thereby invigorating the field
with a firmly established sense of mission. Fragile X syndrome, the leading cause of inherited
mental retardation, has been traced to expansion and abnormal methylation of a triplet repeat,
through which transcription of the FMR-1 gene becomes silenced (Trottier et al., 1993).
Aberrant promoter methylation of tumor suppressor genes has now emerged as an epigenetic
inactivation pathway contributing to tumorigenesis; evidence increasingly supports the notion
that ectopic methylation may play a broad role in gene inactivation and mutation in mammals
(reviewed by Bestor and Coxon, 1993). Much excitement has revolved around the role of DNA
methylation in genomic imprinting, a phenomenon in which parental alleles of the same gene are
expressed at unequal dosages (Barlow, 193). Abnormal expression of imprinted loci is now
implicated in several human disorders (Ogawa et al., 1993; Rainier et al., 1993; Davies,
1992), and more instances seem likely to be discovered. The entire field of DNA methylation,
with its foundations resting largely on the strength of correlative data, took both a
collective breath of relief and a bold step forward with the direct demonstration of Li et al.
(1992) that the DNA MTase gene is essential for development in mice. A molecular mechanism for
spatiotemporal coupling of DNA replication and methylation has been suggested by the finding
that DNA MTase interacts directly with the replication apparatus (Leonhardt et al., 1992).

<>

<1>Verdine, G.L., Bruner, S.D.
<2>How do DNA repair proteins locate damaged bases in the genome?
<3>Chem. Biol.
<4>4
<5>329-334
<6>1997
<7>The genome is susceptible to the attack of reactive species that chemically modify the bases
of DNA.  If genetic information is to be transmitted faithfully to successive generations, it
is essential that the genome be repaired.  All organisms express proteins specifically
dedicated to this task.  But how do these proteins find the aberrant bases amongst the
enormous number of normal ones?

<>

<1>Vereijken, J.M., van Mansfeld, A.D.M., Baas, P.D., Jansz, H.S.
<2>Arthrobacter luteus Restriction endonuclease Cleavage Map of PhiX174 RF DNA.
<3>Virology
<4>68
<5>221-233
<6>1975
<7>Cleavage of PhiX174 RF DNA with the restriction endonuclease from Arthrobacter
luteus (AluI) produces 23 fragments of approximately 24-1100 base pairs in
length.  The order of most of these fragments has been established by digestion
of Haemophilus influenzae Rd (HindII) and Haemophilus aegyptius (HaeIII)
endonuclease fragments of PhiX RF with AluI and by reciprocal digestions of
AluI fragments with HindII and HaeIII.  In this way the Arthrobacter luteus map
could be aligned with the HindII and HaeIII cleavage maps of PhiX174 RF DNA of
A.S. Lee and R.L. Sinsheimer (1974) Proc. Natl. Acad. Sci. USA 71: 2882-2886).

<>

<1>Verhoef, J., van Boven, C.P.A., Holtrigter, B.
<2>Restriction and modification of phages in coagulase-negative staphylococci.
<3>Antonie Van Leeuwenhoek
<4>37
<5>256-267
<6>1971
<7>Phage typing of coagulase-negative staphylococci showed that certain
combination of phage reactions (phage patterns a-g) occurred frequently.  Six
of these could be arranged in three groups (M1, M2, M3) leaving the strains
with pattern g which are sensitive to all phages.  The differences in
susceptibility in patterns a to g were not due to resistance (as all strains
adsorbed all phages) and could not be explained by immunity.  A correlation was
observed between host range of the phage and the pattern of the propagating
strain.

<>

<1>Verhoef, J., van Boven, C.P.A., Holtrigter, B.
<2>Restriction and modification of phages in Staphylococcus epidermidis.
<3>Informative molecules in biological systems., North Holland., Ledoux, L., Amsterdam
<4>0
<5>213-220
<6>1971
<7>The present results demonstrate the occurrence of host-controlled modification
of phage DNA in coagulase-negative staphylococci.  Three modification and
restriction systems, which seem to determine the observed phage typing patterns
to a large extent, could be detected.  In these staphylococci, restrictive
cells appear to break down modified DNA.

<>

<1>Verhoef, J., van Boven, C.P.A., Holtrigter, B.
<2>Host-controlled modification and restriction of phages in coagulase-negative Staphylococci.
<3>J. Gen. Microbiol.
<4>71
<5>231-239
<6>1972
<7>Three host-controlled modification and restriction systems occurring among
coagulase-negative staphlococci belonging to the Staphylococcus subgroup II are
described.  The phage patterns, observed by typing the staphylococci with a
provisional typing set of eighteen phages, are mainly determined by these host
specificity systems.  A strain, which was not restrictive to the phages did not
become restrictive after lysogenization with any of the eighteen phages.
Infection of a restricting host with 3H-labelled phages was followed by a rapid
breakdown of phage DNA, as demonstrated by the appearance of radioactive label
in the cold-acid-soluble DNA fraction of extracted adsorption mixtures.

<>

<1>Verma, M., Kaur, J., Kumar, M., Kumari, K., Saxena, A., Anand, S., Nigam, A., Ravi, V., Raghuvanshi, S., Khurana, P., Tyagi, A.K., Khurana, J.P., Lal, R.
<2>Whole Genome Sequence of the Rifamycin B-Producing Strain Amycolatopsis mediterranei S699.
<3>J. Bacteriol.
<4>193
<5>5562-5563
<6>2011
<7>Amycolatopsis mediterranei S699 is an actinomycete that produces an important antibiotic,
rifamycin B. Semisynthetic derivatives of rifamycin
B are used for the treatment of tuberculosis, leprosy, and AIDS-related
mycobacterial infections. Here, we report the complete genome sequence
(10.2 Mb) of A. mediterranei S699, with 9,575 predicted coding sequences.

<>

<1>Vermersch, P.S., Bennett, G.N.
<2>The use of a selectable FokI cassette in DNA replacement mutagenesis of the R388 dihydrofolate reductase gene.
<3>Gene
<4>54
<5>229-238
<6>1987
<7>FokI, a class-IIS restriction endonuclease, cleaves double-stranded DNA to
produce a protruding 5' end consisting of four nucleotides, 10-13 residues 3'
from the nonpalindromic recognition sequence, GGATG.  Cassettes which utilize
this separation of cleavage and recognition site have been constructed for the
purpose of linker mutagenesis and DNA replacement experiments.  The cassettes
are flanked by FokI recognition sequences oriented such that the FokI cleavage
sites are several nucleotides beyond the cassette/vector fusion sites.  FokI
excises the cassette and several base pairs of the neighboring vector sequence.
The ends produced in the vector by FokI cleavage are generally
noncomplementary and suitable for the insertion of a segment of synthesized
double-stranded replacement DNA.  A cassette which contains a tyrosine tRNA
suppressor gene (supF) is selectable by the suppression of amber mutations in
the recipient host.  A vector containing a pBR322-derived origin of
replication, the Escherichia coli xanthine-guanine phosphoribosyl tranasferase
gene as a selectable marker, and no FokI sites has been constructed for use
with the FokI cassettes.  An experiment which utilized the FokI/supF cassette
to modify the N-terminal coding region of the R388 dihydrofolate reductase gene
is described.

<>

<1>Vermote, C.L.M., Halford, S.E.
<2>EcoRV restriction endonuclease:  communication between catalytic metal ions and DNA recognition.
<3>Biochemistry
<4>31
<5>6082-6089
<6>1992
<7>In the absence of magnesium ions, the EcoRV restriction endonuclease binds all DNA sequences
with equal affinity but cannot cleave DNA. In the presence of Mg2+, the EcoRV endonuclease
cleaves a DNA at one particular sequence, GATATC, at least a million times more readily than
any other sequence. To elucidate the role of the metal ion, the reactions of the EcoRV
restriction enzyme were studied in the presence of MnCl2 instead of MgCl2. The reaction at the
EcoRV recognition site was slower with Mn2+. This was caused partly by reduced rates for
phosphodiester hydrolysis but also by the translocation of the enzyme along the DNA after
cleaving it in one strand. In contrast, alternative sites that differ from the recognition
site by one base pair were cleaved faster in the presence of Mn2+ relative to Mg2+. When
located at an alternative site on the DNA, the EcoRV enzyme bound Mn2+ ions readily but had a
very low affinity for Mg2+. The EcoRV nuclease is thus restrained from cleaving DNA at
alternate sites in the presence of Mg2+, but the restraint fails to operate with Mn2+. A
discrimination factor, which measures the ratio of the activity of the EcoRV nuclease at its
recognition site over that at an alternative site, had values of 3 x 10/5 in MgCl2 and 6 in
MnCl2.

<>

<1>Vermote, C.L.M., Vipond, I.B., Halford, S.E.
<2>EcoRV restriction endonuclease:  communication between DNA recognition and catalysis.
<3>Biochemistry
<4>31
<5>6089-6097
<6>1992
<7>A genetic system was constructed for the mutagenesis of the EcoRV restriction endonuclease and
for the overproduction of mutant proteins. The system was used to make two mutants of EcoRV,
with Ala in place of either Asn185 or Asn188. In the crystal structure of the EcoRV-DNA
complex, both Asn185 and Asn188 contact the DNA within the EcoRV recognition sequence. But
neither mutation affected the ability of the protein to bind to DNA. In the absence of metal
ion cofactors, the mutants bound DNA with almost the same affinity as that of the wild-type
enzyme. In the presence of Mg2+, both mutants retained the ability to cleave DNA specifically
at the EcoRV recognition sequence, but their activities were severely depressed relative to
that of the wild-type. In contrast, with Mn2+ as the cofactor, the mutant enzymes cleaved the
EcoRV recognition site with activities that were close to that of the wild-type. When bound to
DNA at the EcoRV recognition site, the mutant proteins bound Mn2+ ions readily, but they had
much lower affinities for Mg2+ ions than the wild-type enzyme. This was the reason for their
low activities with Mg2+ as the cofactor. The arrangement of the DNA recognition functions, at
one location in the EcoRV restriction enzyme, are therefore responsible for organizing the
catalytic functions at a separate location in the protein.

<>

<1>Vertes, A.A., Inui, M., Kobayashi, M., Kurusu, Y., Yukawa, H.
<2>Presence of mrr- and mcr-like restriction systems in coryneform bacteria.
<3>Res. Microbiol.
<4>144
<5>181-185
<6>1993
<7>Efficient transformation of Brevibacterium flavum MJ233C and Corynebacterium glutamicum ATCC
31831 (up to 5.0 x 107 transformants/ug DNA) depends on the source of plasmid DNA. The
transformation efficiencies of B. flavum MJ233C and C. glutamicum ATCC 31831 increased nearly
103-fold when plasmid DNA was isolated from the recipient strain itself or from a damdcm
Escherichia coli mutant, as compared with DNA passed through a modification-proficient E. coli
strain. These results suggest the presence of a methyl-specific restriction system in certain
strains of coryneform bacteria. In addition, electroporation conditions were optimized.

<>

<1>Vertino, P.M.
<2>Eukaryotic DNA methyltransferases.
<3>S-Adenosylmethionine-dependent Methyltransferases: Structures and Functions., World Scientific Publishing, Cheng, X., Blumenthal, R.M., Singapore
<4>0
<5>341-372
<6>1999
<7>The post-replicative modification of cytosine is an important part of many
restriction-modification systems that defend prokaryotic organisms from invading parasitic
sequences.  DNA cytosine methylation may also play a role in genome defense in eukaryotes, but
has also evolved into a key determinant of epigenetic regulation (for an excellent
comprehensive volume on the topic of epigenetics, the reader is referred to ref 136) and is
involved in such diverse biological functions as gene repression, X-chromosome inactivation,
genome imprinting, and replication timing.  DNA methylation is essential for the normal
development of plant and animal species.  Moreover, somatic alterations in genome methylation
patterns contribute to the genesis of human cancers.  Therefore, just as preservation of the
primary genetic code is important, proper establishment and maintenance of DNA methylation
patterns is also critical to the well-being of the organism.  Until very recently, a single
species of DNA MTase was thought to be responsible for all cytosine methylation in eukaryotes.
Over the last two years, the identification of several new eukaryotic DNA MTases suggests that
the establishment of methylation patterns in early development and their maintenance in the
adult organism requires the activity of several enzymes with distinct cellular or
developmental roles.  This chapter will start with an overview of genome methylation patterns
in higher organisms and the current understanding of how these patterns are established and
maintained, before moving into a discussion of what is known about the structure and function
of the eukaryotic DNA MTases.  I will also discuss some of the evidence suggesting that DNA
MTases affect genome function not only through the passive maintenance of methylation patterns
but also by actively participating in the organization of chromatin and in the genesis of
human disease.

<>

<1>Vertino, P.M., Coll, J.M., Applegren, N., Malkas, L.H.
<2>Identification of DNA (cytosine-5)-methyltransferase as a component of a multiprotein DNA replication complex isolated from human cells.
<3>Proc. Amer. Assoc. Cancer Res.
<4>39
<5>92
<6>1998
<7>The aberrant methylation of normally unmethylated CpG island-containing gene promoters is a
somatic genetic alteration that has been implicated in the inactivation tumor suppressor and
other genes in human cancers.  The mechanisms underlying aberrant methylation are not well
understood, but recent studies showing that increased levels of the enzyme that normally
catalyzes DNA methylation in mammalian cells, DNA (cytosine-5)-methyltransferase (DNA MTase),
can promote aberrant CpG island methylation suggest that DNA MTase itself could play a role.
Little is known about the normal regulation of DNA MTase nor the precise molecular
determinants of its activity in vivo.  In this study, we show that DNA MTase is a component of
the DNA 'synthesome', a multiprotein complex isolated from whole cells that fully supports
origin-dependent DNA synthesis in a cell-free system.  DNA synthesome was purified from two
human cell lines through a series of cell fractionation and chromatography steps.  DNA MTase
protein and activity co-purified with DNA replication activity and were found only in the
replication-competent fractions.  DNA MTase in the replication complex displayed substrate
specificity similar to DNA MTase activity from whole cells.  Specific activity determinations
suggested that most of the cellular DNA MTase exists in a larger complex with the replication
machinery.  Other proteins that have been identified in the synthesome include DNA polymerases
a, b, and e, RP-A, RF-C, DNA primase, PCNA, and topoisomerases I and II.  The definition of
the molecular interactions between DNA MTase and other proteins in replication complex in
normal and neoplastic cells will provide further insight into the regulation of cellular DNA
methylation and the role of DNA/MTase in the establishment of altered DNA methylation patterns
during carcinogenesis.

<>

<1>Vertino, P.M., Yen, R.-W.C., Gao, J., Baylin, S.B.
<2>De Novo methylation of CpG island sequences in human fibroblasts overexpressing DNA (cytosine-5-)-methyltransferase.
<3>Mol. Cell. Biol.
<4>16
<5>4555-4565
<6>1996
<7>Recent studies showing a correlation between the levels of DNA (cytosine-5-)-methyltransferase
(DNA Mtase) enzyme activity and tumorigenicity have implicated this enzyme in the carcinogenic
process.  Moreover, hypermethylation of CpG island-containing promoters is associated with the
inactivation of genes important to tumor initiation and progression.  One proposed role for
DNA Mtase in tumorigenesis is therefore a direct role in the de novo methylation of these
otherwise unmethylated CpG islands.  In this study, we sought to determine whether increased
levels of DNA Mtase could directly affect CpG island methylation.  A full-length cDNA for
human DNA Mtase driven by the cytomegalovirus promoter was constitutively expressed in human
fibroblasts.  Individual clones derived from cells transfected with DNA Mtase (HMT) expressed
1- to 50-fold the level of DNA Mtase protein and enzyme activity of the parental cell line or
clones transfected with the control vector alone (neo).  To determine the effects of DNA Mtase
overexpression on CpG island methylation, we examined 12 endogenous CpG island loci in the HMT
clones.  HMT clones expressing greater-than or equal-to 9-fold the parental levels of DNA
Mtase activity was significantly hypermethylated relative to at least 11 Neo clones at five
CpG island loci.  In the HMT clones, methylation reached nearly 100% at susceptible CpG island
loci with time in culture.  In contrast, there was little change in the methylation status in
the Neo clones over the same time frame.  Taken together, the data indicate that
overexpression of DNA Mtase can drive the de novo methylation of susceptible CpG island loci,
thus providing support for the idea that DNA Mtase can contribute to tumor progression through
CpG island methylation-mediated gene inactivation.

<>

<1>Veselkov, A.G., Demidov, V.V., Nielsen, P.E., Frank-Kamenetskii, M.D.
<2>A new class of genome rare cutters.
<3>Nucleic Acids Res.
<4>24
<5>2483-2487
<6>1996
<7>Although significant efforts have been directed at developing efficient techniques
for rare and super rare genome cutting, only limited success has been achieved.  Here we
propose
a new approach to solve this problem.  We demonstrate that peptide nucleic acid 'clamps'
(bis-
PNAs) bind strongly and sequence specifically to short homopyrimidine sites on lambda and
yeast
genomic DNAs.  Such binding efficiently shields methylation/restriction sites which overlap
with
the bis-PNA binding sites from enzymatic methylation.  After removing the bis-PNA, the genomic
DNAs are quantitatively cleaved by restriction enzymes into a limited number of pieces of
lengths
from several hundred kbp to several Mbp.  By combining various bis-PNAs with different
methylation/restriction enzyme pairs, a huge new class of genome rare cutters can be created.
These cutters cover the range of recognition specificities where very few, if any, cutters are
now
available.

<>

<1>Vesely, Z., Muller, A., Schmitz, G.G., Kaluza, K., Jarsch, M., Kessler, C.
<2>RleAI: a novel class-IIS restriction endonuclease from Rhizobium leguminosarum recognizing 5'-CCCACA(N)12-3' and 3'-GGGTGT(N)9-5'.
<3>Gene
<4>95
<5>129-131
<6>1990
<7>The novel class-IIS restriction endonuclease, RleAI, has the
5'-CCCACA(N)12-3'
3'-GGGTGT(N)9-5' specificity.

<>

<1>Vesely, Z., Schmitz, G.G., Jarsch, M., Kessler, C.
<2>BbrPI, a novel PmaCI isoschizomer from Bacillus brevis recognizing 5'-CAC/GTG-3'.
<3>Nucleic Acids Res.
<4>18
<5>3423
<6>1990
<7>None

<>

<1>Veyrier, F.J., Ecobichon, C., Boneca, I.G.
<2>Draft Genome Sequence of Strain X47-2AL, a Feline Helicobacter pylori Isolate.
<3>Genome Announcements
<4>1
<5>e01095-13
<6>2013
<7>Helicobacter pylori is a human-specific pathogen that exclusively inhabits the human gastric
mucosa. However, occasionally, humans transmit H. pylori to
susceptible animal hosts bred in colonies. Here, we report the genome sequence of
strain X47-2AL, isolated from a domestic cat and used in anti-H. pylori
immunization studies.

<>

<1>Veyrier, F.J., Hong, E., Deghmane, A.E., Taha, M.K.
<2>Draft Genome Sequence of a Neisseria meningitidis Serogroup C Isolate of Sequence Type 11 Linked to an Outbreak among Men Who Have Sex with Men.
<3>Genome Announcements
<4>1
<5>e00795-13
<6>2013
<7>Meningococcal disease occurs as sporadic cases in developed countries, with the occasional
emergence of new clones of Neisseria meningitidis. Here, we report the
genome sequence of N. meningitidis strain LNP27256, an isolate of sequence type
11 linked to a recent outbreak among men who have sex with men in Europe.

<>

<1>Viadiu, H., Aggarwal, A.K.
<2>The role of metals in catalysis by the restriction endonuclease BamHI.
<3>Nat. Struct. Biol.
<4>5
<5>910-916
<6>1998
<7>Type II restriction enzymes are characterized by their remarkable specificity and simplicity.
They require only divalent metals (such as Mg2+ or Mn2+) as cofactors to catalyze the
hydrolysis of DNA. However, most of the structural work on endonucleases has been performed in
the absence of metals, leaving unanswered questions about their mechanisms of DNA cleavage.
Here we report structures of the endonuclease BamHI-DNA complex, determined in the presence of
Mn2+ and Ca2+, that describe the enzyme at different stages of catalysis. Overall, the results
support a two-metal mechanism of DNA cleavage for BamHI which is distinct from that of EcoRV.

<>

<1>Viadiu, H., Aggarwal, A.K.
<2>Structure of BamHI bound to nonspecific DNA: A model for DNA sliding.
<3>Mol. Cell
<4>5
<5>889-895
<6>2000
<7>The central problem faced by DNA binding proteins is how to select the correct DNA sequence
from the sea of nonspecific sequences in a cell.  The problem is particularly acute for
bacterial restriction enzymes because cleavage at an incorrect DNA site could be lethal.  To
understand the basis of this selectivity, we report here the crystal structure of endonuclease
BamHI bound to noncognate DNA.  We show that, despite only a single base pair change in the
recognition sequence, the enzyme adopts an open configuration that is on the pathway between
free and specifically bound forms of the enzyme.  Surprisingly, the DNA drops out of the
binding cleft with a total loss of base-specific and backbone contacts.  Taken together, the
structure provides a remarkable snapshot of an enzyme poised for linear diffusion (rather than
cleavage) along the DNA.

<>

<1>Viadiu, H., Kucera, R., Schildkraut, I., Aggarwal, A.K.
<2>Crystallization of Restriction Endonuclease BamHI with Nonspecific DNA.
<3>J. Struct. Biol.
<4>130
<5>81-85
<6>2000
<7>Restriction endonucleases show extraordinary specificity in distinguishing specific from
nonspecific DNA sequences. A single basepair change within the recognition sequence results in
over a million-fold loss in activity. To understand the basis of this sequence discrimination,
it is just as important to study the nonspecific complex as the specific complex. We describe
here the crystallization of restriction endonuclease BamHI with several nonspecific
oligonucleotides. The 11-mer, 5'-ATGAATCCATA-3', yielded cocrystals with BamHI, in the
presence of low salt, that diffracted to 1.9 A with synchrotron radiation. The cocrystals
belong to the space group P2(1)2(1)2(1) with unit cell dimensions of a = 114.8 A, b = 91.1 A,
c = 66.4 A, alpha = 90 degrees, beta = 90 degrees, gamma = 90 degrees. This success in the
cocrystallization of BamHI with a nonspecific DNA provides insights for future attempts at
crystallization of other nonspecific DNA-protein complexes. Copyright 2000 Academic Press.

<>

<1>Viadiu, H., Vanamee, E.S., Jacobson, E.M., Schildkraut, I., Aggarwal, A.K.
<2>Crystallization of restriction endonuclease SfiI in complex with DNA.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>59
<5>1493-1495
<6>2003
<7>The SfiI endonuclease from Streptomyces fimbriatus (EC 3.1.21.4) is a tetrameric enzyme that
binds simultaneously to two recognition sites and
cleaves both sites concertedly. It serves as a good model system for
studying both specificity and cooperative DNA binding. Crystals of the
enzyme were obtained by the hanging-drop vapor-diffusion method in complex
with a 21-mer oligonucleotide. The crystals are trigonal, with unit-cell
parameters a = b = 85.7, c = 202.6 A, and diffract to 2.6 A resolution on
a rotating-anode X-ray generator. Preliminary X-ray analysis reveals the
space group to be either P3(1)21 or P3(2)21. Interestingly, the crystals
change to space group P6(1)22, with unit-cell parameters a = b = 85.5, c =
419.6 A, when the selenomethionyl (SeMet) derivative of the enzyme is
co-crystallized with the same DNA. Phase information is currently being
derived from this SeMet SfiI-DNA complex.

<>

<1>Viadiu-Ilarraza, H.
<2>Structural study of specificity and catalysis by restriction endonuclease BamHI.
<3>Diss. Abstr.
<4>60
<5>5978
<6>2000
<7>The restriction endonuclease BamHI is a phosphoryl-transferase that recognizes the DNA
sequence 5'-GGATCC-3' and cleaves the phosphodiester bond between the first and the second
bases.  In this work we used X-ray crystallography to analyze the different steps in the
mechanism of DNA recognition and catalysis by BamHI.  BamHI, a polypeptide of 213 amino acids,
forms a dimer in solution and starts its catalytic mechanism binding DNA non-specifically.
Initially, the crystal structure of BamHI bound to a non-specific DNA (5'-GAATCC-3') at 2.0
A is presented.  The analysis of the non-specific DNA-BamHI structure shows striking
differences with the previously reported specific complex.  In the non-specific complex, BamHI
has very few contacts to the DNA which shows exceptionally high B-factors and its position in
the DNA cavity drops 7A with respect to the specific complex.  Although the protein
conformation is symmetrical and it closely resembles the one in the absence of DNA, the
quaternary arrangement results in a DNA cavity with an intermediate width between the ones
observed for the five protein and the specific complex.  In summary, the non-specific
DNA-BamHI complex explains the electrostatic character of the non-specific DNA-protein binding
and suggest that BamHI is trapped in a conformation suitable for sliding along the DNA.  Once
BamHI has found the specific site, it cleaves both DNA strands in a sequential manner.  In
this work, the crystal structures of BamHI before and after the reaction are presented, at 2.0
A and 1.8 A respectively.  These data shows evidence to support a two-metal mechanism for
BamHI catalysis.  The metals are coordinated by three acidic residues and they stabilize the
transition state.  The residue Glu113 was identified as a general base.  A water molecule
attacking the phosphate group, and a water donating a proton to the leaving group were also
located.  Furthermore, the comparison of BamHI, BamHI Glu113Lys mutant, and other restriction
endonucleases suggest a catalytic mechanism for those enzymes that contain three acidic
residues and a lysine in the active site.  Overall the work presented here represents the most
comprehensive study of different stages in the catalytic cycle of a restriction endonuclease.

<>

<1>Victoria, J.G., Kapoor, A., Li, L., Blinkova, O., Slikas, B., Wang, C., Naeem, A., Zaidi, S., Delwart, E.
<2>Metagenomic analyses of viruses in stool samples from children with acute flaccid paralysis.
<3>J. Virol.
<4>83
<5>4642-4651
<6>2009
<7>We analyzed viral nucleic acids in stool samples collected from 35 South
Asian children with nonpolio acute flaccid paralysis (AFP).
Sequence-independent reverse transcription and PCR amplification of
capsid-protected, nuclease-resistant viral nucleic acids were followed by
DNA sequencing and sequence similarity searches. Limited Sanger sequencing
(35 to 240 subclones per sample) identified an average of 1.4 distinct
eukaryotic viruses per sample, while pyrosequencing yielded 2.6 viruses
per sample. In addition to bacteriophage and plant viruses, we detected
known enteric viruses, including rotavirus, adenovirus, picobirnavirus,
and human enterovirus species A (HEV-A) to HEV-C, as well as numerous
other members of the Picornaviridae family, including parechovirus, Aichi
virus, rhinovirus, and human cardiovirus. The viruses with the most
divergent sequences relative to those of previously reported viruses
included members of a novel Picornaviridae genus and four new viral
species (members of the Dicistroviridae, Nodaviridae, and Circoviridae
families and the Bocavirus genus). Samples from six healthy contacts of
AFP patients were similarly analyzed and also contained numerous viruses,
particularly HEV-C, including a potentially novel Enterovirus genotype.
Determining the prevalences and pathogenicities of the novel genotypes,
species, genera, and potential new viral families identified in this study
in different demographic groups will require further studies with
different demographic and patient groups, now facilitated by knowledge of
these viral genomes.

<>

<1>Viedma, E., Juan, C., Otero, J.R., Oliver, A., Chaves, F.
<2>Draft Genome Sequence of VIM-2-Producing Multidrug-Resistant Pseudomonas aeruginosa ST175, an Epidemic High-Risk Clone.
<3>Genome Announcements
<4>1
<5>e00112-13
<6>2013
<7>The VIM-2-producing multidrug-resistant high-risk clone Pseudomonas aeruginosa sequence type
(ST) 175 was isolated in the setting of a large outbreak in
Hospital Universitario 12 de Octubre (Spain) from 2007 to 2010. This strain was
resistant to all beta-lactams, fluoroquinolones, and aminoglycosides, with the
exception of amikacin, and has become an endemic clone in our institution.

<>

<1>Viedma, E., Villa, J., Juan, C., Oliver, A., Chaves, F.
<2>Draft Genome Sequence of Colistin-Only-Susceptible Pseudomonas aeruginosa Strain  ST235, a Hypervirulent High-Risk Clone in Spain.
<3>Genome Announcements
<4>2
<5>e01097-14
<6>2014
<7>We report the genome of colistin-only-susceptible Pseudomonas aeruginosa strain ST235
(PA_ST235). This isolate was obtained in the setting of an outbreak in a
tertiary hospital in Spain. This clone was apparently associated with a
significantly higher mortality rate. The ST235 clone also appears to be
associated with greater virulence.

<>

<1>Vienozinskis, M.T., Nesterenko, V.F., Kanopkaite, S.I., Buryanov, Y.I.
<2>Effect of 5-azacytidine on E. coli cells with different DNA methylases.
<3>Biokhimiia
<4>50
<5>749-754
<6>1985
<7>A correlation was found between the bacteriocide effect of 5-aza-C and the amount of cytosine
DNA-methylases in E. coli cells.  5-Aza-C-DNA on cytosine DNA-methylases was due to the
formation of stable inactive complexes between the enzyme and the non-methylating cytosine
analog in the recognition sites.  Cytosine DNA-methylase EcoRII formed a relatively firm bond
with 5-aza-C-DNA, which could be disrupted by 1 M KCl; this disruption restores the
DNA-methylase activity and the inhibiting capacity of 5-aza-C-DNA.  Thus, the binding of
cytosine DNA-methylase to 5-aza-C in DNA is noncovalent; the inhibition of the enzyme by
5-aza-C-DNA is reversible.

<>

<1>Vijesurier, R.M.
<2>Regulation of the gene expression of the PvuII restriction-modification system.
<3>Diss. Abstr.
<4>57
<5>7379B-7380B
<6>1997
<7>The pvuIIC gene of the PvuII restriction-modification system of Proteus vulgaris encodes the
9.4 kDa C.PvuII protein.  This protein has a helix-turn-helix motif and is a DNA binding
protein.  The C.PvuII protein binds to the C-box sequence located within the pvuIIM gene;
C.PvuII binding to its target sequence would interfere with transcription of pvuIIM.
Therefore, C.PvuII may play the role of a repressor of methyltransferase expression.  Previous
studies have shown that the overexpression of the methyltransferase is lethal to the cell; as
such, the C.PvuII protein may be involved in preventing the overexpression of the
methyltransferase, thereby preventing the deleterious effects of hypermethylation.  This work
shows that the overexpression of C.PvuII is lethal to cells also containing the intact PvuII
restriction-modification system, perhaps due to the repression of pvuIIM, paving the way for
autorestriction.  The C.PvuII protein does not appear to regulate the transcription of either
the pvuIIC or pvuIIR genes, as seen in primer extension analyses.  In addition, the pvuIIC and
pvuIIR genes are transcribed as a polycistronic message, since the pvuIIR gene itself does not
appear to have a strong promoter; the polycistronic message originates from the relatively
stronger pvuIIC promoter.  The 'C-proteins', of which C.PvuII is a member, are encoded by
genes of several restriction-modification systems, and appear to enhance endonuclease
expression, while downregulating that of the methyltransferase.  This work suggests that
C.PvuII, like its counterparts of other restriction-modification systems, diminishes
methyltransferase expression.

<>

<1>Vijesurier, R.M., Carlock, L., Blumenthal, R.M., Dunbar, J.C.
<2>Role and mechanism of action of C.PvuII, a regulatory protein conserved among restriction-modification systems.
<3>J. Bacteriol.
<4>182
<5>477-487
<6>2000
<7>The PvuII restriction-modification system is a type II system, which means that its
restriction endonuclease and modification methyltransferase are independently active proteins.
The PvuII system is carried on a plasmid, and its movement into a new host cell is expected to
be followed initially by expression of the methyltransferase gene alone so that the new
host's DNA is protected before endonuclease activity appears.  Previous studies have
identified a regulatory gene (pvuIIC) between the divergently oriented genes for the
restriction endonuclease (pvuIIR) and modification methyltransferase (pvuIIM), with pvuIIC in
the same orientation as and partially overlapping pvuIIR.  This product of pvuIIC, C.PvuII,
was found to act in trans and to be required for expression of pvuIIR.  In this study we
demonstrate that premature expression of pvuIIC prevents establishment of the PvuII genes,
consistent with the model that requiring C.PvuII for pvuIIR expression provides a timing delay
essential for protection of the new host's DNA.  We find that the opposing pvuIIC and pvuIIM
transcripts overlap by over 60 nucleotides at their 5' ends, raising the possibility that
their hybridization might play a regulatory role.  We furthermore characterize the action of
C.PvuII, demonstrating that it is a sequence-specific DNA-binding protein that binds to the
pvuIIC promoter and stimulates transcription of both pvuIIC and pvuIIR into a polycistronic
mRNA.  The apparent location of C.PvuII binding, overlapping the -10 promoter hexamer and the
pvuIICR transcriptional starting points, is highly unusual for transcriptional activators.

<>

<1>Vikram, S., Kumar, S., Subramanian, S., Raghava, G.P.
<2>Draft Genome Sequence of the Nitrophenol-Degrading Actinomycete Rhodococcus imtechensis RKJ300.
<3>J. Bacteriol.
<4>194
<5>3543
<6>2012
<7>We report the 8.231-Mb genome sequence of Rhodococcus imtechensis RKJ300, isolated from
pesticide-contaminated soil in Punjab, India. The genome sequence
of the strain RKJ300 will be helpful in exploring the molecular pathways involved
in the degradation of nitrophenols.

<>

<1>Vikram, S., Kumar, S., Vaidya, B., Pinnaka, A.K., Raghava, G.P.
<2>Draft Genome Sequence of the 2-Chloro-4-Nitrophenol-Degrading Bacterium Arthrobacter sp. Strain SJCon.
<3>Genome Announcements
<4>1
<5>e0005813
<6>2013
<7>We report the 4.39-Mb draft genome sequence of the 2-chloro-4-nitrophenol-degrading bacterium
Arthrobacter sp. strain SJCon,
isolated from a pesticide-contaminated site. The draft genome sequence of strain
SJCon will be helpful in studying the genetic pathways involved in the
degradation of several aromatic compounds.

<>

<1>Vilacoba, E., Deraspe, M., Traglia, G.M., Roy, P.H., Ramirez, M.S., Centron, D.
<2>Draft Genome Sequence of an International Clonal Lineage 1 Acinetobacter baumannii Strain from Argentina.
<3>Genome Announcements
<4>2
<5>e01190-14
<6>2014
<7>In the last few years Acinetobacter baumannii has emerged worldwide as an important nosocomial
pathogen in medical institutions. Here, we present the draft
genome sequence of the international clonal lineage 1 (ICL1) A. baumannii strain
A144 that was isolated in a hospital in Buenos Aires City in the year 1997. The
strain is susceptible to carbapenems and resistant to trimethoprim and
gentamicin.

<>

<1>Vilain, A., Apiou, F., Dutrillaux, B., Malfoy, B.
<2>Assignment  of candidate DNA methyltransferase gene (DNMT2) to human chromosome band 10p15.1 by in situ hybridization.
<3>Cytogenet. Cell Genet.
<4>82
<5>120
<6>1998
<7>Methylation of deoxycytidine in mammalian DNA is thought to be involved in many biological
phenomena.  Only one DNA methyltransferase was known in human.  The gene (DNMT1) was localized
to 19p13.2.  Recently a putative new DNA methyltransferase (DNMT2) has been cloned and
characterized.  Using a panel of radiation hybrids, this gene has been mapped to 10p14-p12.

<>

<1>Vilkaitis, G., Dong, A., Weinhold, E., Cheng, X., Klimasauskas, S.
<2>Functional roles of the conserved threonine 250 in the target recognition domain of HhaI DNA methyltransferase.
<3>J. Biol. Chem.
<4>275
<5>38722-38730
<6>2000
<7>DNA cytosine-5-methyltransferase HhaI recognizes the GCGC sequence and flips the inner
cytosine out of the DNA helix and into the catalytic site for methylation. The 5'-phosphate
of the flipped out cytosine is in contact with the conserved Thr-250 from the target
recognition domain. We have produced 12 mutants of Thr-250 and examined their methylation
potential in vivo. Six active mutants were subjected to detailed biochemical and structural
studies. Mutants with similar or smaller side chains (Ser, Cys, and Gly) are very similar to
wild-type enzyme in terms of steady-state kinetic parameters k(cat), K(m)(DNA), K(m)(AdoMet).
In contrast, the mutants with bulkier side chains (Asn, Asp, and His) show increased K(m)
values for both substrates. Fluorescence titrations and stopped-flow kinetic analysis of
interactions with duplex oligonucleotides containing 2-aminopurine at the target base position
indicate that the T250G mutation leads to a more polar but less solvent-accessible position of
the flipped out target base. The x-ray structure of the ternary M.HhaI(T250G).DNA. AdoHcy
complex shows that the target cytosine is locked in the catalytic center of enzyme. The space
created by the mutation is filled by water molecules and the adjacent DNA backbone atoms
dislocate slightly toward the missing side chain. In aggregate, our results suggest that the
side chain of Thr-250 is involved in constraining the conformation the DNA backbone and the
target base during its rotation into the catalytic site of enzyme.

<>

<1>Vilkaitis, G., Klimasauskas, S.
<2>Bisulfite sequencing protocol displays both 5-methylcytosine and N4-methylcytosine.
<3>Anal. Biochem.
<4>271
<5>116-119
<6>1999
<7>DNA premethylated in vitro or in vivo with a DNA cytosine-N4 methyltransferase, M.MvaI, was
analyzed using a bisulfite DNA sequencing technique. We find that, under reaction conditions
reliably displaying 5-methylcytosine on the sequencing ladder, N4-methylcytosine residues are
displayed with efficiencies from 33 to 100%, depending on a sequence context. This finding
offers a convenient tool for determining sequence specificity of cytosine-N4
methyltransferases and demonstrates that the two types of biological cytosine methylation are
not reliably distinguished by this technique.

<>

<1>Vilkaitis, G., Lubys, A., Merkiene, E., Timinskas, A., Janulaitis, A., Klimasauskas, S.
<2>Circular permutation of DNA cytosine-N4 methyltransferases: in vivo coexistence in the BcnI system and in vitro probing by hybrid formation.
<3>Nucleic Acids Res.
<4>30
<5>1547-1557
<6>2002
<7>Sequence analysis of the BcnI restriction-modification system from Bacillus centrosporus
revealed four open reading frames (bcnIC, bcnIR, bcnIB and bcnIA) that are arranged as two
converging collinear pairs. One pair encodes a putative small regulatory protein, C.BcnI, and
the restriction endonuclease R.BcnI. The other two gene products are the DNA cytosine-N4
methyltransferases M.BcnIA and M.BcnIB, which differ by circular permutation of conserved
sequence motifs. The BcnI methyltransferases are isospecific on double-stranded DNA
[methylation specificity CC(C/G)GG], but M.BcnIA can also methylate the target sites in
single-stranded DNA. Functional analysis shows that bcnIA is dispensable (bcnIB is capable of
protecting the DNA against the in vivo activity of bcnIR); in contrast, no stable clones were
obtained if bcnIB alone was deleted from the system. By analogy with the DpnII system, the
second methylase M.BcnIA may play a role in the transformation proficiency of its
gram-positive host. The interchangeability of homologous elements in the beta class of
cytosine-N4 methylases was probed by hybrid formation between M.BcnIB and its closest homolog
M.Cfr9I (CCCGGG) employing a novel semi-random strategy combined with selection for catalytic
activity. The fusion points in the active hybrids mapped in a narrow region located between
sequence motifs X and I. Our data illustrate that recombination of two related sequences by
circular permutation may serve as an evolutionary mechanism for creating new specificities of
amino MTases.

<>

<1>Vilkaitis, G., Merkiene, E., Serva, S., Weinhold, E., Klimasauskas, S.
<2>The mechanism of DNA cytosine-5 methylation. Kinetic and mutational dissection of HhaI methyltransferase.
<3>J. Biol. Chem.
<4>276
<5>20924-20934
<6>2001
<7>Kinetic and binding studies involving a model DNA cytosine-5-methyltransferase, M.HhaI, and a
37-mer DNA duplex containing a single hemimethylated target site were applied to characterize
intermediates on the reaction pathway. Stopped-flow fluorescence studies reveal that cofactor
S-adenosyl-l-methionine (AdoMet) and product S-adenosyl-l-homocysteine (AdoHcy) form similar
rapidly reversible binary complexes with the enzyme in solution. The M.HhaI.AdoMet complex
(k(off) = 22 s^-1, KD = 6 microM) is partially converted into products during
isotope-partitioning experiments, suggesting that it is catalytically competent. Chemical
formation of the product M.HhaI.(Me)DNA.AdoHcy (k(chem) = 0.26 s^-1) is followed by a slower
decay step (k(off) = 0.045 s^-1), which is the rate-limiting step in the catalytic cycle
(k(cat) = 0.04 s^-1). Analysis of reaction products shows that the hemimethylated substrate
undergoes complete (>95%) conversion into fully methylated product during the initial burst
phase, indicating that M.HhaI exerts high binding selectivity toward the target strand. The
T250N, T250D, and T250H mutations, which introduce moderate perturbation in the catalytic
site, lead to substantially increased KD(DNA(ternary)), k(off)(DNA(ternary)),
KM(AdoMet(ternary)) values but small changes in KD(DNA(binary)), KD(AdoMet(binary)),
k(chem), and k(cat). When the target cytosine is replaced with 5-fluorocytosine, the chemistry
step leading to an irreversible covalent M.HhaI.DNA complex is inhibited 400-fold
(k(chem)(5FC) = 0.7 x 10^-3 s^-1), and the Thr-250 mutations confer further dramatic
decrease of the rate of the covalent methylation k(chem). We suggest that activation of the
pyrimidine ring via covalent addition at C-6 is a major contributor to the rate of the
chemistry step (k(chem)) in the case of cytosine but not 5-fluorocytosine. In contrast to
previous reports, our results imply a random substrate binding order mechanism for M.HhaI.

<>

<1>Vilkaitis, G., Suetake, I., Klimasauskas, S., Tajima, S.
<2>Processive methylation of hemimethylated CpG sites by mouse Dnmt1 DNA methyltransferase.
<3>J. Biol. Chem.
<4>280
<5>64-72
<6>2005
<7>DNA methyltransferase Dnmt1 ensures clonal transmission of lineage-specific DNA methylation
patterns in a mammalian genome during
replication. Dnmt1 is targeted to replication foci, interacts with
PCNA, and favors methylating the hemimethylated form of CpG sites. To
understand the underlying mechanism of its maintenance function, we
purified recombinant forms of full-length Dnmt1, a truncated form of
Dnmt1-(291-1620)lacking the binding sites for PCNA and DNA and examined
their processivity using a series of long unmethylated and
hemimethylated DNA substrates. Direct analysis of methylation patterns
using bisulfite-sequencing and hairpin-PCR techniques demonstrated that
full-length Dnmt1 methylates hemimethylated DNA with high processivity
and a fidelity of over 95%, but unmethylated DNA with much less
processivity. The truncated form of Dnmt1 showed identical properties
to full-length Dnmt1 indicating that the N-terminal 290-amino acid
residue region of Dnmt1 is not required for preferential activity
toward hemimethylated sites or for processivity of the enzyme.
Remarkably, our analyses also revealed that Dnmt1 methylates
hemimethylated CpG sites on one strand of double-stranded DNA during a
single processive run. Our findings suggest that these inherent
enzymatic properties of Dnmt1 play an essential role in the faithful
and efficient maintenance of methylation patterns in the mammalian
genome.

<>

<1>Villa, A.A., Kropinski, A.M., Abbasifar, R., Griffiths, M.W.
<2>Complete Genome Sequence of Vibrio parahaemolyticus Bacteriophage vB_VpaM_MAR.
<3>J. Virol.
<4>86
<5>13138-13139
<6>2012
<7>Vibrio parahaemolyticus is a major pathogen that is mainly associated with seafood and is a
global food safety issue. Our objective was to isolate and completely sequence a specific
phage against this bacterium. Phage vB_VpaM_MAR is able to lyse 76% of the V. parahaemolyticus
strains tested. MAR belongs to the Myoviridae family and has a genome comprised of
double-stranded DNA with a size of 41,351 bp, a G+C content of 51.3%, and 62 open reading
frames (ORFs). Bioinformatic analysis showed that phage MAR is closely related to Vibrio
phages VHML, VP58.5, and VP882 and Halomonas aquamarina phage PhiHAP-1.

<>

<1>Villa, J., Viedma, E., Branas, P., Mingorance, J., Chaves, F.
<2>Draft Whole-Genome Sequence of OXA-48-Producing Multidrug-Resistant Klebsiella pneumoniae KP_ST11_OXA-48.
<3>Genome Announcements
<4>2
<5>e00737-14
<6>2014
<7>We present the draft genome sequence of a blood culture isolate of OXA-48-producing Klebsiella
pneumoniae (sequence type 11 [ST11]) obtained in the
course of a hospital outbreak in Spain. Sequence analysis showed 121 genes
related to antibiotic and antiseptic resistance, including blaOXA-48, which was
located in an IncL/M plasmid.

<>

<1>Villa, J., Viedma, E., Otero, J.R., Chaves, F.
<2>Draft Whole-Genome Sequence of VIM-1-Producing Multidrug-Resistant Enterobacter cloacae EC_38VIM1.
<3>Genome Announcements
<4>1
<5>e00694-13
<6>2013
<7>The VIM-1-producing multidrug-resistant strain Enterobacter cloacae was isolated  from blood
culture. The strain showed multiple resistances to clinically used
antibiotics, including all beta-lactams, fluoroquinolones, aminoglycosides, and
sulfonamides. Sequence analysis showed the presence of 14 genes associated with
resistance to antibiotics, including the metallo-beta-lactamase VIM-1 gene, which
was located in a class 1 integron.

<>

<1>Villanueva, J., Switala, J., Ivancich, A., Loewen, P.C.
<2>Genome Sequence of Bacillus pumilus MTCC B6033.
<3>Genome Announcements
<4>2
<5>e00327-14
<6>2014
<7>Bacillus pumilus is a Gram-positive, rod-shaped, aerobic bacterium isolated from
the soil. B. pumilus strain B6033 was originally selected as a biocatalyst for
the stereospecific oxidation of beta-lactams. Here, we present a 3.8-Mb assembly
of its genome, which is the second fully assembled genome of a B. pumilus strain.

<>

<1>Villeneuve, C., Martineau, C., Mauffrey, F., Villemur, R.
<2>Complete Genome Sequences of Methylophaga sp. Strain JAM1 and Methylophaga sp. Strain JAM7.
<3>J. Bacteriol.
<4>194
<5>4126-4127
<6>2012
<7>Methylophaga sp. strains JAM1 and JAM7 have been isolated from a denitrification  system.
Strain JAM1 was the first Methylophaga strain reported to be able to grow
under denitrifying conditions. Here, we report the complete genome sequences of
the two strains, which allowed prediction of gene clusters involved in
denitrification in strain JAM1.

<>

<1>Villion, M., Chopin, M.C., Deveau, H., Ehrlich, S.D., Moineau, S., Chopin, A.
<2>P087, a lactococcal phage with a morphogenesis module similar to an Enterococcus faecalis prophage.
<3>Virology
<4>388
<5>49-56
<6>2009
<7>The virulent lactococcal phage P087 was isolated from a dairy environment
in 1978. This phage was then recognized as the reference member for one of
the ten phage groups currently known to infect Lactococcus lactis strains.
The double-stranded DNA genome of this Siphoviridae phage is composed of
60,074 bp and is circularly permuted. Five tRNA and 88 orfs were found
within an uncommon genome architecture. Eleven structural proteins were
also identified through SDS-PAGE and LC-MS/MS analyses. Of note, 11
translated orfs from the structural module of phage P087 have identities
to gene products found in a prophage located in the genome of Enterococcus
faecalis V583. The alignment of both genomic sequences suggests that DNA
exchanges could occur between these two phages which are infecting low G+C
bacteria found in similar ecological niches.

<>

<1>Vilo, C., Benedik, M.J., Ilori, M., Dong, Q.
<2>Draft Genome Sequence of Cupriavidus sp. Strain SK-4, a di-ortho-Substituted Biphenyl-Utilizing Bacterium Isolated from Polychlorinated Biphenyl-Contaminated   Sludge.
<3>Genome Announcements
<4>2
<5>e00474-14
<6>2014
<7>Cupriavidus sp. strain SK-4 is a bacterium capable of growing aerobically on
monochlorobiphenyls and dichlorobiphenyls as the sole carbon sources for growth.
Here, we report its draft genome sequence with the aim of facilitating an
understanding of polychlorinated biphenyl biodegradation mechanisms.

<>

<1>Vilo, C., Benedik, M.J., Ilori, M., Dong, Q.
<2>Draft Genome Sequence of Cupriavidus sp. Strain SK-3, a 4-Chlorobiphenyl- and 4-Clorobenzoic Acid-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e00664-14
<6>2014
<7>We report the draft genome sequence of Cupriavidus sp. strain SK-3, which can use
4-chlorobiphenyl and 4-clorobenzoic acid as the sole carbon source for growth.
The draft genome sequence allowed the study of the polychlorinated biphenyl
degradation mechanism and the recharacterization of the strain SK-3 as a
Cupriavidus species.

<>

<1>Vilo, C.A., Benedik, M.J., Kunz, D.A., Dong, Q.
<2>Draft Genome Sequence of the Cyanide-Utilizing Bacterium Pseudomonas fluorescens  Strain NCIMB 11764.
<3>J. Bacteriol.
<4>194
<5>6618-6619
<6>2012
<7>We report here the 6.97-Mb draft genome sequence of Pseudomonas fluorescens strain NCIMB
11764, which is capable of growth on cyanide as the sole nitrogen
source. The draft genome sequence allowed the discovery of several genes
implicated in enzymatic cyanide turnover and provided additional information
contributing to a better understanding of this organism's unique cyanotrophic
ability. This is the first sequenced genome of a cyanide-assimilating bacterium.

<>

<1>Vilpo, J.A., Vilpo, L.M.
<2>Restriction, methylation and ligation of 5-hydroxymethyluracil-containing DNA.
<3>Mutat. Res.
<4>316
<5>123-131
<6>1995
<7>Oxidation of DNA and its components can cause genetic mutations and chromosomal instability.
These changes have generally been implicated in aging. Oxidation of the methyl group of
thymidine residues in DNA is known to result in the formation of
5-hydroxymethyl-2'-deoxyuridine (5HmdUrd). We have utilized Bacillus subtilis phage SPO1 DNA
as a model of oxidatively damaged DNA. In this phage, all thymine (Thy) residues are replaced
by 5-hydroxymethyluracil (5HmUra), but the species is naturally devoid of other
oxidatively-induced DNA lesions. Particular attention was paid to the behavior of
5HmUra-containing DNA as a target for several enzymes employing DNA as substrate; restriction
endonucleases, dam DNA methylase and T4 DNA ligase. We noticed that susceptibility of SPO1 DNA
varied when different restriction endonucleases having 5HmUra in the restriction sites were
tested. Endonucleolytic cleavage brought about by Sau3AI proceeded as effectively with SPO1
DNA as with conventional DNA (lambda phage). The same was true when the ligation of Sau3AI
sites was performed with T7 DNA ligase. In contrast, both endonucleolytic cleavage and
ligation were slower in SPO1 DNA, compared with lambda phage, when TaqI and T4 DNA ligase were
used for restriction and ligation, respectively. We also noticed that SPO1 phage does not
naturally contain N6-methyladenine (N6MeAde) opposite 5HmUra, i.e., no hydrolysis of SPO1 DNA
was observed when assessed with methylation-dependent restriction endonuclease DpnI. Our
results show that the presence of 5HmUra in the respective site of DNA does not, per se,
prevent the activity of restriction endonucleases, ligases or DNA methylases. These data
support the view that oxidation of Thy to 5HmUra in target DNA does not necessarily result in
substantial deterioration in the functions of DNA processing enzymes.

<>

<1>Vincent, A.T., Trudel, M.V., Paquet, V.E., Boyle, B., Tanaka, K.H., Dallaire-Dufresne, S., Daher, R.K., Frenette, M., Derome, N., Charette, S.J.
<2>Detection of Variants of the pRAS3, pAB5S9, and pSN254 Plasmids in Aeromonas salmonicida subsp. salmonicida: Multidrug Resistance, Interspecies Exchanges, and Plasmid Reshaping.
<3>Antimicrob. Agents Chemother.
<4>58
<5>7367-7374
<6>2014
<7>The ubiquitous water-borne Gram-negative bacterium Aeromonas salmonicida subsp.
salmonicida is the causative agent of furunculosis, a worldwide disease in fish
farms. Plasmids carrying antibiotic resistance genes have already been described
for this bacterium. The aim of the present study was to identify and characterize
additional multidrug resistance plasmids in A. salmonicida subsp. salmonicida. We
sequenced the plasmids present in two multiple antibiotic-resistant isolates
using high-throughput technologies. We also investigated 19 other isolates with
various multidrug resistance profiles by genotyping PCR and assessed their
resistance to tetracycline. We identified variants of the pAB5S9 and pSN254
plasmids that carry several antibiotic resistance genes and that have been
previously reported in bacteria other than A. salmonicida subsp. salmonicida,
which suggests a high level of interspecies exchange. Genotyping analyses and the
antibiotic resistance profiles of the 19 other isolates support the idea that
multiple versions of pAB5S9 and pSN254 exist in A. salmonicida subsp.
salmonicida. We also identified variants of the pRAS3 plasmid. The present study
revealed that A. salmonicida subsp. salmonicida harbors a wide variety of
plasmids, which suggests that this ubiquitous bacterium may contribute to the
spread of antibiotic resistance genes in the environment.

<>

<1>Vincent, M., Xu, Y., Kong, H.
<2>Helicase Dependent Amplification of Nucleic Acids.
<3>Japanese Patent Office
<4>JP 2006500028 A
<5>
<6>2006
<7>
<>

<1>Vincze, T., Posfai, J., Roberts, R.J.
<2>NEBcutter: a program to cleave DNA with restriction enzymes.
<3>Nucleic Acids Res.
<4>31
<5>3688-3691
<6>2003
<7>NEBcutter, version 1.0, is a program available via a web server
(http://tools.neb.com/NEBcutter) that will accept an input DNA sequence
and produce a comprehensive report of the restriction enzymes that will
cleave the sequence. It produces a variety of outputs including
restriction enzyme maps, theoretical digests and links into the
restriction enzyme database, REBASE (http://www.neb.com/rebase).
Importantly, its table of recognition sites is updated daily from REBASE
and it marks all sites that are potentially affected by DNA methylation
(Dam, Dcm, etc.). Many options exist to choose the enzymes used for
digestion, including all known specificities, subsets of those that are
commercially available or sets of enzymes that produce compatible termini.

<>

<1>Vinella, D., Jaffe, A., D'Ari, R., Kohiyama, M., Hughes, P.
<2>Chromosome partitioning in Escherichia coli in the absence of Dam-directed methylation.
<3>J. Bacteriol.
<4>174
<5>2388-2390
<6>1992
<7>Escherichia coli dam mutants, lacking the GATC DNA methylase, do not produce anucleate cells
at high frequencies, suggesting that hemimethylation of the chromosome origin of replication,
oriC, is not essential for correct chromosome partitioning.

<>

<1>Vinogradova, M.N., Gromova, E.S., Gryaznova, O.I., Isagulyants, M.D., Kuznetsova, S.A., Kosych, V.G., Shabarova, Z.A.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments.  IX:  Cleavage of substrates with point modifications in the recognition site and flanking sequences.
<3>Bioorg. Khim.
<4>13
<5>1194-1204
<6>1987
<7>Ability of the EcoRII restriction endonuclease to cleave 14-base-pair DNA
duplexes with nucleotide substitutions in the recognition site CC(A/T)GG and in
the adjacent base pair has been studied.  Modifications leading to a local
change in the substrate conformation (rU residue in and outside the recognition
site, A.A- or A.C-pairs in the flanking sequences) reduce the rate of
hydrolysis, the effect being maximal when the modified base pair is outside the
recognition site.  No digestion occurs when the internal dC-residue of the
recognition site is 5-methylated in one or both strands.  Replacement of dT
residue in the EcoRII recognition site by dF5U residue results in a dramatic
inhibition of hydrolysis.  Km and Kcat for the cleavage of 14-base-pair DNA
duplex have been determined.  The cleavage rate of the dT-containing strand of
the recognition site is 1.5 fold higher comparing with the dA-containing
strand.  The cleavage of both strands of the substrate by EcoRII endonuclease
is confirmed to proceed in one enzyme-substrate complex.

<>

<1>Vinogradova, M.N., Gromova, E.S., Kosykh, V.G., Buryanov, Y.I., Shabarova, Z.A.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments.  XI. Determination of the EcoRII binding site.
<3>Mol. Biol. (Mosk)
<4>24
<5>847-850
<6>1990
<7>EcoRII restriction endonuclease binding site has been determined on the basis
of comparison of the binding parameters of the enzyme with synthetic
DNA-duplexes of concatemer type containing a different number of EcoRII
recognition sites.  It has been shown that it consists of 21 +/- 1 base pairs.

<>

<1>Vinogradova, M.N., Gromova, E.S., Purmal, A.A., Kosykh, V.G., Shabarova, Z.A.
<2>Interaction of restriction and modification enzymes EcoRII with synthetic fragments of DNA.
<3>Mol. Biol. (Mosk)
<4>20
<5>1329-1336
<6>1986
<7>It was shown by the method of binding on nitrocellulose filters that the
restriction endonuclease EcoRII in the absence of Mg2+ ions is specifically
bonded to synthetic DNA duplexes of the concatamer type of different lengths,
containing natural recognition sites for the enzyme CAA/TGG, as well as with
substrates possessing a noncomplementary AA or TT pair in the recognition site
of cleavage by the enzyme.  The degree of binding of substrates containing a
central AT, TT, or AA pair in the recognition site decreases in the series AT >
TT >> AA.  Replacement of the phosphodiester bond in the recognition site by a
pyrophosphate or phosphoamide bond has little effect on the stability of the
complexes.  The affinity of the enzyme for nonspecific DNA sequences is two
orders of magnitude lower than for the specific recognition sites of EcoRII.
The equilibrium constant of association for a substrate with one recognition
site is 3.9 x 10(8) M-1.  The introduction of Mg2+ ions into the incubation
mixture leads to destabilization of the complex of EcoRII endonuclease with the
DNA duplex containing pyrophosphate bonds.  The rate constants of dissociation
and the half-lives of the cocomplexes of the endonuclease EcoRII with synthetic
substrates were determined.

<>

<1>Vinogradova, M.N., Gromova, E.S., Uporova, T.M., Nikolskaya, I.I., Shabarova, Z.A.
<2>Restriction endonuclease SsoII:  interaction with modified substrates.
<3>Dokl. Akad. Nauk.
<4>295
<5>732-736
<6>1987
<7>None

<>

<1>Vipond, B., Halford, S.E.
<2>Structure-function correlation for the EcoRV restriction enzyme: from non-specific binding to specific DNA cleavage.
<3>Mol. Microbiol.
<4>9
<5>225-231
<6>1993
<7>The EcoRV restriction endonuclease cleaves DNA at its recognition sequence at least a million
times faster than at any other DNA sequence. The only cofactor it requires for activity is
Mg2+; but in binding to DNA in the absence of Mg2+, the EcoRV enzyme shows no specificity for
its recognition site. Instead, the reason why EcoRV cuts one DNA sequence faster than any
other is that the rate of cleavage is controlled by the binding of Mg2+ to EcoRV-DNA
complexes; the complex at the recognition site has a high affinity for Mg2+, while the
complexes at other DNA sequences have low affinities for Mg2+. The structures of the EcoRV
endonuclease, and of its complexes with either specific or non-specific DNA, have been solved
by X-ray crystallography. In the specific complex, the protein interacts with the bases in the
recognition sequence and the DNA takes up a highly distorted structure. In the non-specific
complex with an unrelated DNA sequence, there are virtually no interactions with the bases and
the DNA retains a B-like structure. Since the free energy changes for the formation of
specific and non-specific complexes are the same, the energy from the specific interactions
balances that required for the distortion of the DNA. The distortion inserts the phosphate at
the scissile bond into the active site of the enzyme, where it forms part of the binding site
for Mg2+. Without this distortion, the EcoRV-DNA complex would be unable to bind Mg2+ and thus
unable to cleave DNA. The specificity of the EcoRV restriction enzyme is therefore governed,
not by DNA binding as such, but by its ability to organize the structure of the DNA to which
it is bound.

<>

<1>Vipond, I.B., Baldwin, G.S., Halford, S.E.
<2>Divalent metal ions at the active sites of the EcoRV and EcoRI restriction endonucleases.
<3>Biochemistry
<4>34
<5>697-704
<6>1995
<7>Restriction enzymes cannot cleave DNA without a metal ion cofactor. The specificities of the
EcoRV and EcoRI endonucleases for metals were studied by measuring DNA cleavage rates with
several metal ions and with combinations of metal ions. Both EcoRV and EcoRI had optimal
activities with Mg2+, were less active with several other ions including Mn2+, and had
virtually no activity with Ca2+. But the activities of EcoRV and EcoRI with either Mg2+ or
Mn2+ were perturbed by Ca2+. For EcoRI, both Mg2+- and Mn2+-dependent activities, at both
cognate and noncognate sites, were all inhibited by Ca2+. The activity of EcoRV at its
recognition site with Mg2+ was also inhibited by Ca2+. But the Mn2+-dependent reaction at the
EcoRV recognition site was stimulated by Ca2+. EcoRV activities at noncognate sites with
either Mg2+ or Mn2+ displayed a biphasic response to Ca2+: stimulation at low concentrations
of Ca2+ and inhibition at higher concentrations. These observations, together with the known
structures of the proteins, indicate that EcoRI needs only one metal ion per active site and
is inactive when Mg2+ is displaced by Ca2+, while EcoRV needs two and that the displacement of
one by Ca2+ can enhance activity. We propose a mechanism for phosphodiester hydrolysis by
EcoRV that involves two metal ions.

<>

<1>Vipond, I.B., Baldwin, G.S., Oram, M., Erskine, S.G., Wentzell, L.M., Szczelkun, M.D., Nobbs, T.J., Halford, S.E.
<2>A general assay for restriction endonucleases and other DNA-modifying enzymes with plasmid substrates.
<3>Mol. Biotechnol.
<4>4
<5>259-268
<6>1995
<7>A procedure for measuring the activities of enzymes that alter the covalent structure of DNA
is described.  The assay utilizes covalently closed circles of DNA as the substrate and yields
quantitative data on the fraction of this DNA converted to both open-circle and linear forms.

<>

<1>Vipond, I.B., Halford, S.E.
<2>Specific DNA recognition by EcoRV restriction endonuclease induced by Calcium ions.
<3>Biochemistry
<4>34
<5>1113-1119
<6>1995
<7>In the presence of Mg2+, the EcoRV restriction endonuclease cleaves DNA specifically at its
recognition sequence, but in the absence of divalent metal ions, it binds DNA without any
specificity: gel-shift experiments had revealed multiple EcoRV-DNA complexes, due to the
binding of one, two, three, or more molecules of protein per molecule of DNA, with the same
equilibrium constant for each association. In this study, the binding of EcoRV to DNA was
measured by gel shift in the presence of Ca2+, an ion that perturbs the Mg2+-dependent
activity of EcoRV but that fails to support DNA cleavage. With Ca2+, and at a lower
concentration of EcoRV protein than that required for binding in the absence of divalent metal
ions, a single complex was observed with DNA containing the EcoRV recognition site. This
complex was not formed with DNA that had been methylated at the EcoRV site nor with an
isogenic DNA lacking the EcoRV recognition site. The single complex thus is due to the
specific binding of EcoRV to its recognition site on the DNA. From gel shifts with a permuted
set of DNA fragments, the degree of DNA bending by EcoRV at this recognition site was
estimated to be 53 degrees +/- 4 degrees. This angle is similar to that seen in the crystal
structure of the cognate DNA-protein complex. Calcium ions thus appear to mimic the role of
Mg2+ in generating a specific protein-metal-DNA complex, but in contrast to Mg2+, Ca2+ gives a
stable ternary complex in which the DNA-bound nuclease cannot cleave the DNA.

<>

<1>Vipond, I.B., Halford, S.E.
<2>Random mutagenesis targeted to the active site of the EcoRV restriction endonuclease.
<3>Biochemistry
<4>35
<5>1701-1711
<6>1996
<7>Two segments of the gene for the EcoRV restriction endonuclease, each encoding 10 amino acids
at the active site, were subjected to random mutagenesis with degenerate oligonucleotides.
Mutations that abolished the activity of the EcoRV endonuclease were selected by viability in
a strain of Escherichia coli that lacks the EcoRV methyltransferase, under conditions where
the gene for the wild-type endonuclease is lethal to the cell.  Sixty-five mutants were
isolated and analyzed by DNA sequencing to identify the mutations.  The collection of null
mutants contained 49 with single amino acid substitutions, 15 with double substitutions, and
one with a triple substitution.  The single substitutions were located at many different
positions within the two 10-amino acid segments, though several hot-spots gave rise to null
mutants at high frequencies.  Some hot-spots were readily explained by reference to the
crystal structure of EcoRV since they were at the amino acids immediately adjacent to the
scissile phosphodiester bond: for example, Asp90 and Lys92.  These residues may be directly
involved in the catalytic mechanism.  Other hot-spots, such as Gln69, Tyr72, and Ala88, were
at unexpected positions that appear to have no direct role in DNA binding or catalysis.  At
some of the unexpected hot-spots, the side chain of the amino acid lies distant from the DNA,
yet the enzyme was still inactivated by conservative substitutions at these positions.  The
sensitivity of the EcoRV endonuclease to conservative substitutions may be due to its
requirement to take up one particular conformation at the DNA--protein interface out of a
large number of alternative conformations.

<>

<1>Vipond, I.B., Moon, B.-J., Halford, S.E.
<2>An isoleucine to leucine mutation that switches the cofactor requirement of the EcoRV restriction endonuclease from magnesium to manganese.
<3>Biochemistry
<4>35
<5>1712-1721
<6>1996
<7>The EcoRV restriction endonuclease cleaves DNA at its recognition sequence more readily with
Mg2+ as the cofactor than with Mn2+ but, at noncognate sequences that differ from the EcoRV
site by one base pair, Mn2+ gives higher rates than Mg2+.  A mutant of EcoRV, in which an
isoleucine near the active site was replaced by leucine, showed the opposite behavior.  It had
low activity with Mg2+, but, in the presence of Mn2+ ions, it cleaved the recognition site
faster than wild-type EcoRV with either Mn2+ or Mg2+.  The mutant was also more specific for
the recognition sequence than the native enzyme: the noncognate DNA cleavages by wild-type
EcoRV and Mn2+ were not detected with the mutant.  Further mutagenesis showed that the protein
required the same acidic residues at its active site as wild-type EcoRV.  The Ile-Leu mutation
seems to perturb the configuration of the metal-binding ligands at the active site so that the
protein has virtually no affinity for Mg2+ yet it can still bind Mn2+ ions, though the latter
only occurs when the protein is at the recognition site.  This contrasts to wild-type EcoRV,
where Mn2+ ions bind readily to complexes with either cognate and noncognate DNA and only Mg2+
shows the discrimination between the complexes.  The structural perturbation is a specific
consequence of leucine in place of isoleucine, since mutants with valine or alanine were
similar to wild-type EcoRV.

<>

<1>Vipond, I.B., Moon, B.J., Halford, S.E.
<2>An Ile-to-Leu mutant of the EcoRV restriction endonuclease that requires Mn2+ as catalytic cofactor.
<3>Biochem. Soc. Trans.
<4>22
<5>301S
<6>1994
<7>The EcoRV restriction endonuclease recognizes and cleaves the sequence GAT^ATC with extreme
precision. The accuracy is such that alternate sites one bp different are cleaved at least a
million times slower. However, contrary to many DNA binding proteins, the basis of this
specificity is not due to preferred binding by EcoRV to its recognition sequence. In fact, the
EcoRV nuclease binds all DNA sequences with equal affinity. Instead, it is due to the affinity
of the enzyme-DNA complex for the divalent metal ion which serves as the catalytic cofactor.
Thus, EcoRV bound to GATATC has a high affinity for Mg2+, while the enzyme bound to any other
sequence has a low affinity for the metal ion.

<>

<1>Viprey, V., Rosenthal, A., Broughton, W.J., Perret, X.
<2>Genetic snapshots of the Rhizobium species NGR234 genome.
<3>Genome Biology
<4>1
<5>RESEARCH0014
<6>2000
<7>BACKGROUND: In nitrate-poor soils, many leguminous plants form
nitrogen-fixing symbioses with members of the bacterial family
Rhizobiaceae. We selected Rhizobium sp. NGR234 for its exceptionally broad
host range, which includes more than I 12 genera of legumes. Unlike the
genome of Bradyrhizobium japonicum, which is composed of a single 8.7 Mb
chromosome, that of NGR234 is partitioned into three replicons: a
chromosome of about 3.5 Mb, a megaplasmid of more than 2 Mb (pNGR234b) and
pNGR234a, a 536,165 bp plasmid that carries most of the genes required for
symbioses with legumes. Symbiotic loci represent only a small portion of
all the genes coded by rhizobial genomes, however. To rapidly characterize
the two largest replicons of NGR234, the genome of strain ANU265 (a
derivative strain cured of pNGR234a) was analyzed by shotgun sequencing.
RESULTS: Homology searches of public databases with 2,275 random sequences
of strain ANU265 resulted in the identification of 1,130 putative
protein-coding sequences, of which 922 (41%) could be classified into
functional groups. In contrast to the 18% of insertion-like sequences
(ISs) found on the symbiotic plasmid pNGR234a, only 2.2% of the shotgun
sequences represent known ISs, suggesting that pNGR234a is enriched in
such elements. Hybridization data also indicate that the density of known
transposable elements is higher in pNGR234b (the megaplasmid) than on the
chromosome. Rhizobium-specific intergenic mosaic elements (RIMEs) were
found in 35 shotgun sequences, 6 of which carry RIME2 repeats previously
thought to be present only in Rhizobium meliloti. As non-overlapping
shotgun sequences together represent approximately 10% of ANU265 genome,
the chromosome and megaplasmid may carry a total of over 200 RIMEs.
CONCLUSIONS: 'Skimming' the genome of Rhizobium sp. NGR234 sheds new light
on the fine structure and evolution of its replicons, as well as on the
integration of symbiotic functions in the genome of a soil bacterium.
Although most putative coding sequences could be distributed into
functional classes similar to those in Bacillus subtilis, functions
related to transposable elements were more abundant in NGR234. In contrast
to ISs that accumulated in pNGR234a and pNGR234b, the hundreds of RIME
elements seem mostly attributes of the chromosome.

<>

<1>Viratyosin, W., Kulawonganunchai, S., Smittipat, N., Juthayothin, T., Penpassakarn, P., Pasomsub, E., Chantratita, W., Chaiprasert, A., Palittapongarnpim, P.
<2>Draft Genome Sequence of the Mycobacterium tuberculosis Strain 43-16836, Belonging to the Indo-Oceanic Lineage, Isolated From Tuberculous Meningitis in  Thailand.
<3>Genome Announcements
<4>1
<5>e00801-13
<6>2013
<7>We present the draft genome sequence of Mycobacterium tuberculosis strain 43-16836, belonging
to the Indo-Oceanic lineage, isolated from a tuberculous
meningitis patient in Thailand. The genome is 4,381,942 bp long with 4,316
protein-coding genes and contains new single nucleotide polymorphisms (SNPs),
including SNPs of genes that may encode cell wall components and possibly
influence virulence.

<>

<1>Vishnivetskaya, T.A. et al.
<2>Draft genome sequences of 10 strains of the genus exiguobacterium.
<3>Genome Announcements
<4>2
<5>e01058-14
<6>2014
<7>High-quality draft genome sequences were determined for 10 Exiguobacterium strains in order to
provide insight into their evolutionary strategies for
speciation and environmental adaptation. The selected genomes include
psychrotrophic and thermophilic species from a range of habitats, which will
allow for a comparison of metabolic pathways and stress response genes.

<>

<1>Vissel, B., Choo, K.H.
<2>Altered activity of restriction endonuclease MnlI cleavage of mouse satellite DNA.
<3>Nucleic Acids Res.
<4>16
<5>4731
<6>1988
<7>MnlI is reported to recognise the sequence CCTCN7, cleaving after the N7
position to produce a blunt ended fragment.  However, we have observed that
following MnlI cleavage of mouse satellite DNA, the monomeric units that were
subsequently cloned uniformly lacked the N7 base.  Mouse satellite DNA is
composed of tandemly repeated 234bp monomeric units, most of which contain an
MnlI site.  As shown in Fig. 1, direct cloning and sequencing of the MnlI
monomeric units revealed that the N7 base was consistently absent from the end
of the clone with the MnlI site (Fig. 1C, D and G).  These results suggest that
cleavage of mouse satellite DNA by MnlI occurs after the N6, in addition to
after the N7 position.  It is not clear, however, if MnlI removes the N7 base
from both strands simultaneously, or, cleaves to leave a single-base 5' or 3'
extension that is subsequently lost during the cloning procedure.  This unusual
activity of MnlI may be related to the high AT content or methylated nature of
the mouse satellite DNA.

<>

<1>Visser, M. et al.
<2>Genome analyses of the carboxydotrophic sulfate-reducers Desulfotomaculum nigrificans and Desulfotomaculum carboxydivorans and reclassification of  Desulfotomaculum caboxydivorans as a later synonym of Desulfotomaculum  nigrificans.
<3>Standards in Genomic Sciences
<4>9
<5>655-675
<6>2014
<7>Desulfotomaculum nigrificans and D. carboxydivorans are moderately thermophilic members of the
polyphyletic spore-forming genus Desulfotomaculum in the family
Peptococcaceae. They are phylogenetically very closely related and belong to
'subgroup a' of the Desulfotomaculum cluster 1. D. nigrificans and D.
carboxydivorans have a similar growth substrate spectrum; they can grow with
glucose and fructose as electron donors in the presence of sulfate. Additionally,
both species are able to ferment fructose, although fermentation of glucose is
only reported for D. carboxydivorans. D. nigrificans is able to grow with 20%
carbon monoxide (CO) coupled to sulfate reduction, while D. carboxydivorans can
grow at 100% CO with and without sulfate. Hydrogen is produced during growth with
CO by D. carboxydivorans. Here we present a summary of the features of D.
nigrificans and D. carboxydivorans together with the description of the complete
genome sequencing and annotation of both strains. Moreover, we compared the
genomes of both strains to reveal their differences. This comparison led us to
propose a reclassification of D. carboxydivorans as a later heterotypic synonym
of D. nigrificans.

<>

<1>Visser, M. et al.
<2>Genome analysis of Desulfotomaculum kuznetsovii strain 17(T) reveals a physiological similarity with Pelotomaculum thermopropionicum strain SI(T).
<3>Standards in Genomic Sciences
<4>8
<5>69-87
<6>2013
<7>Desulfotomaculum kuznetsovii is a moderately thermophilic member of the polyphyletic
spore-forming genus Desulfotomaculum in the family Peptococcaceae.
This species is of interest because it originates from deep subsurface thermal
mineral water at a depth of about 3,000 m. D. kuznetsovii is a rather versatile
bacterium as it can grow with a large variety of organic substrates, including
short-chain and long-chain fatty acids, which are degraded completely to carbon
dioxide coupled to the reduction of sulfate. It can grow methylotrophically with
methanol and sulfate and autotrophically with H2 + CO2 and sulfate. For growth it
does not require any vitamins. Here, we describe the features of D. kuznetsovii
together with the genome sequence and annotation. The chromosome has 3,601,386 bp
organized in one contig. A total of 3,567 candidate protein-encoding genes and 58
RNA genes were identified. Genes of the acetyl-CoA pathway, possibly involved in
heterotrophic growth with acetate and methanol, and in CO2 fixation during
autotrophic growth are present. Genomic comparison revealed that D. kuznetsovii
shows a high similarity with Pelotomaculum thermopropionicum. Genes involved in
propionate metabolism of these two strains show a strong similarity. However,
main differences are found in genes involved in the electron acceptor metabolism.

<>

<1>Vitkute, J., Maneliene, Z., Janulaitis, A.
<2>AbeI, a restriction endonuclease from Azotobacter beijerinckii, which recognizes the asymmetric heptanucleotide sequence 5'-CCTCAGC-3'(-5/-2).
<3>Nucleic Acids Res.
<4>26
<5>4917-4918
<6>1998
<7>A new restriction endonuclease AbeI has been isolated from Azotobacter beijerinckii.  This
enzyme recognizes the asymmetric heptanucleotide sequence 5'-CCTCAGC-3' and cleaves within
it symmetrically at positions -5/-2 in the opposing strands, producing three base protruding
5'-ends.

<>

<1>Vitkute, J., Maneliene, Z., Janulaitis, A.
<2>Two new thermostable type II restriction endonucleases from Thermus aquaticus: TatI and TauI, which recognize the novel nucleotide sequences 5'-W^GTACW-3' and 5'-GCSG^C-3' respectively.
<3>FEMS Microbiol. Lett.
<4>204
<5>253-257
<6>2001
<7>One hundred and forty isolates of thermophilic bacteria from the genus Thermus were screened
for the presence of restriction endonuclease activity. Thermostable isoschizomers of
restriction endonucleases, such as AceIII, BbvI, BglI, BsePI, FnuDII, HgiAI, MaeII, MboI,
MseI, PvuII, StuI, TaqI, Tsp4CI, TspEI, XhoI and XmaIII, were isolated. Two restriction
enzymes, TatI and TauI, recognizing novel degenerate sequences 5'-W^GTACW-3' and
5'-GCSG^C-3' respectively were partially purified and the recognition and cleavage sites
were determined.

<>

<1>Vitkute, J., Maneliene, Z., Petrusyte, M., Janulaitis, A.
<2>BplI, a new BcgI-like restriction endonuclease, which recognizes a symmetric sequence.
<3>Nucleic Acids Res.
<4>25
<5>4444-4446
<6>1997
<7>BcgI and BcgI-like restriction endonucleases cleave double stranded DNA specifically on both
sides of their asymmetric recognition sequences which are interrupted by several ambiguous
base pairs.  Their heterosubunit structure, bifunctionality and stimulation by AdoMet make
them different from other classified restriction enzymes.  Here we report on a new BcgI-like
enzyme, which in contrast to all other BcgI-like enzymes, recognizes a symmetric interrupted
sequence, and which, like BcgI, cleaves double stranded DNA upstream and downstream of its
recognition sequence (8/13)GAGN5CTC(13/8).  Like BcgI, BplI is a bifunctional enzyme revealing
both DNA cleavage and methyltransferase activities.  There are two polypeptides in the
homogeneous preparation of BplI with molecular masses of ~74 and 37 kDa.  The sizes of the
BplI subunits are close to those of BcgI, but the proportion 1:1 in the final preparation is
different from that of 2:1 in BcgI.  Low activity observed with Mg2+ increases >100-fold in
the presence of AdoMet.  Even with AdoMet though, specific cleavage is incomplete.
S-adenosyl-homocysteine (AdoHcy) or sinefungin can replace AdoMet in the cleavage reaction.
AdoHcy activated BplI yields complete cleavage of DNA.

<>

<1>Vitkute, J., Maneliene, Z., Petrusyte, M., Janulaitis, A.
<2>BfiI, a restriction endonuclease from Bacillus firmus S8120, which recognizes the novel non-palindromic sequence 5'-ACTGGG(N)5/4-3'.
<3>Nucleic Acids Res.
<4>26
<5>3348-3349
<6>1998
<7>A new type IIS restriction endonuclease BfiI has been partially purified from Bacillus firmus
S8120.  BfiI recognizes the non-palindromic hexanucleotide sequence 5'-ACTGGG(N)5/4-3' and
makes a staggered cut at the fifth base pair downstream of the recognition sequence on the
upper strand, producing a single base 3' protruding end.

<>

<1>Vitkute, J., Stankevicius, K., Tamulaitiene, G., Maneliene, Z., Timinskas, A., Berg, D.E., Janulaitis, A.
<2>Specificities of eleven different DNA methyltransferases of Helicobacter pylori strain 26695.
<3>J. Bacteriol.
<4>183
<5>443-450
<6>2001
<7>Methyltransferases (MTases) of prokaryotes affect general cellular processes such as mismatch
repair, regulation of transcription, replication, and transposition, and in some cases may be
essential for viability. As components of restriction-modification systems, they contribute to
bacterial genetic diversity. The genome of Helicobacter pylori strain 26695 contains 25 open
reading frames encoding putative DNA MTases. To assess which MTase genes are active, strain
26695 genomic DNA was tested for cleavage by
147 restriction endonucleases; 24 were found that did not cleave this DNA. The specificities
of 11 expressed MTases and the genes encoding them were identified from this restriction data,
combined with the known sensitivities of restriction endonucleases to specific DNA
modification, homology searches, gene cloning and genomic mapping of the methylated bases m4C,
m5C, and m6A.

<>

<1>Vitkute, J., Tamulaitiene, G., Chalkauskas, H., Janulaitis, A.
<2>Potential of H. pylori to produce R-M enzymes and their possible biological role.
<3>Int. J. Med. Microbiol.
<4>291S
<5>95
<6>2001
<7>The unprecedented abundance of putative restriction-modification (R-M) genes found in H.
pylori genomes raises question on their role in life cycle of bacteria.  As a prerequisite to
such studies we screened 156 independent isolates for restriction endonucleases to assess
expression and specificity variability of H. pylori enzymes.  386 REs has been characterized
with 26 different specificities.  Every screened strain contained REs and their set was
specific for the majority of them.  In addition a set of seven selected strains was
investigated for DNA methylases, including J99 and 26695 strains.  We found 87 MTases with 22
different specificities.  Given that cognate MTase accompanies every RE, altogether 32 MTases
of different specificity has been identified.  Modeling indicates that this variability is
near the limit of potential specificity variability for R-M enzymes in H. pylori.  The
conclusion reinforces previous observations in Enterobacteriaceae on the phenomenon of
taxonomic specificity of R-M systems.  The character of taxonospecificity represents the first
nonexperimental evidence supporting hypothesis on "selfishness" of R-M genes.  The main role
of H. pylori R-M systems is likely to modulate involvement of H. pylori and non-H. pylori DNA
in genetic exchange of the bacteria.  Arguments are provided to support this hypothesis.
Other possible functions of R-M systems are also discussed.

<>

<1>Vitor, J.M.B.
<2>Restriction and modification systems in Campylobacter jejuni and Campylobacter coli.
<3>Ph.D. Thesis, University of Lisbon, , , Lisbon
<4>
<5>1-220
<6>1999
<7>The restriction endonucleases are one of the most widely used
tools in molecular biology and acronyms like EcoRI or BamHI are used
daily in any laboratory all over the world.  Although ubiquitous among
the prokaryotes there are only a few scientific groups studying these
systems.  The RE are a part of the restriction and modification systems
and it is believed that they are a defense mechanism of the host
bacteria against bacteriophages, although not all scientists agree with
this theory.  It is also believed that RM systems can act as intra and
inter-specific barriers to the exchange of genetic information and may
be involved in genetic recombination since they produce DNA fragments
that can be integrated in the host genome or in an extrachromosomal
element.

<>

<1>Vitor, J.M.B., Alves, P., Marques, M., Vital, J.
<2>Helicobacter pylori type IIG restriction and modification loci.
<3>Helicobacter, Blackwell Publishing Ltd., , 
<4>16
<5>98
<6>2011
<7>Helicobacter pylori complete sequenced genomes have a large number of putative restriction and
modification systems between 26 and 34.  The major consequence of RMS is keeping the bacterial
genomes free from alien DNA, acting as a speciation barrier.  RMS are classified into four
major types and several subtypes.  The H. pylori genomes annotated in REBASE were analyzed and
it was found that RMS are not randomly distributed over the genome.  Using H. pylori 26695 as
model, four type IIG RMS loci were found: 1st - glpC.horF; 2nd - nadC-HINFIM; 3rd - tmk-polA;
4th - res-nusA.  Primers were designed for the four loci and tested in 17 different H. pylori
strains, isolated from asymptomatic patients.  PCR products were obtained in all loci but
differences were observed among them.  All the PCR products were digested with HindIII to
evaluate the variability of the amplified genes.  Locus 1 and 4 are empty in a large
percentage of strains, 64.7 and 52.9 respectively.  Locus 2 has a high percentage of strains
with unspecific PCR products, 58.8%.  Different HindIII profiles were observed: 2 in locus 1,
5 in locus 2 and 4, and 6 in locus 3.  This might correspond to 18 different type IIG RMS in
17 strains, thus being one more example of H. pylori genetic diversity although their species
genomic organization might be conservative.

<>

<1>Vitor, J.M.B., Costa, L., Pires, I., Cabrita, J., Correia, R.V.
<2>Restriction enzymes in Campylobacter strains.
<3>Microb. Ecol. Health Dis.
<4>4
<5>S50
<6>1991
<7>Restriction enzymes (RE) are very important tools in Genetic Engineering.  They
are a part of the Restriction Modification (RM) systems, that are ubiquitous in
prokaryotes.  We have searched for RE activity in 100 enteropathogenic
Campylobacter strains (52 C. jejuni and 58 C. coli strains) from human and
animal origin.  The strains were isolated, subcultured and identified by
routine methods.  DNAse activity was searched for as described by Lior and the
RE screening by Whitehead and Brown's method, with modifications concerning the
time of lysis by lysozyme, the time and temperature of the detergent step and
the saline concentration of the digestion buffer.  The DNA used was from
bacteriophage lambda methyl free.

<>

<1>Vitor, J.M.B., Morgan, R.D.
<2>Two novel restriction endonucleases from Campylobacter jejuni.
<3>Gene
<4>157
<5>109-110
<6>1995
<7>We have discovered two unusual restriction endonucleases (ENases) in two Campylobacter jejuni
strains that recognize asymmetrical, interrupted sequences and cleave the DNA both before and
after their recognition sites.  Both enzymes require AdoMet as a cofactor for their ENase
activity.

<>

<1>Vitor, J.M.B., Polisson, C., Cabrita, J., Silva, R.V.C.
<2>Restriction enzymes from Campylobacter coli strains.
<3>Acta Gastroenterol. Belg.
<4>56
<5>41
<6>1993
<7>The type II restriction enzymes (RE) are ubiquitous among Prokaryotes. These enzymes belong to
restriction-modification systems, which include a methyltransferase. One function is thought
to protect the genomic information from other genetic pools. Also it is believed that when a
strain exhibits restriction activity there will be a methyltransferase. Eight Campylobacter
coli strains known to have RE activity from previous screening, isolated from human, pig and
chicken, were plated on BHI agar and incubated for 20h at 42C, in an anaerobic jar (5% O2, 10%
CO2). 1g of cells were lysed by sonication and centrifuged 25 min at 40,000g. The assay for RE
was made in NEB buffer no2, 1h at 37C with lambda and T7 DNAs. If necessary, the extract was
purified by affinity chromatography, with Heparin-Sepharose CL-6B. We have identified nine RE:
CcoP95I (GCGC) isoschizomer of HhaI; CcoP215I and CcoP216I (GCNGC) isoschizomers of Fnu4HI;
CcoP31I, CcoP84I and CcoP219I (GATC) isoschizomers of MboI; CcoP73I and CcoP76I (GTAC)
isoschizomers of RsaI. Although we have studied a small number of isolates, the results
suggest that the REs detected in Campylobacter coli recognize mainly GC-rich sequences. This
finding may explain the prevalence in those strains of AT-rich plasmid DNA as well as the low
chromosomal GC content (32-34%) of them.

<>

<1>Vitoriano, I., Vitor, J.M.B., Oleastro, M., Roxo-Rosa, M., Vale, F.F.
<2>Proteome variability among Helicobacter pylori isolates clustered according to genomic methylation.
<3>J. Appl. Microbiol.
<4>114
<5>1817-1832
<6>2013
<7>Aims To understand whether the variability found in the proteome of Helicobacter pylori
relates to the genomic methylation, virulence and
associated gastric disease. Methods and Results We applied the
Minimum-Common-Restriction-Modification (MCRM) algorithm to genomic
methylation data of 30 Portuguese H.pylori strains, obtained by genome
sensitivity to Type II restriction enzymes' digestion. All the
generated dendrograms presented three clusters with no association with
gastric disease. Comparative analysis of two-dimensional gel
electrophoresis (2DE) maps obtained for total protein extracts of 10 of
these strains, representative of the three main clusters, revealed that
among 70 matched protein spots (in a universe of 300), 16 were
differently abundant (P<0 center dot 05) among clusters. Of these, 13
proteins appear to be related to the cagA genotype or gastric disease.
The abundance of three protein species, DnaK, GlnA and HylB, appeared
to be dictated by the methylation status of their gene promoter.
Conclusions Variations in the proteome profile of strains with common
geographic origin appear to be related to differences in cagA genotype
or gastric disease, rather than to clusters organized according to
strain genomic methylation. Significance and Impact of the Study The
simultaneous study of the genomic methylation and proteome is important
to correlate epigenetic modifications with gene expression and pathogen
virulence.

<>

<1>Vlasova, T.I., Demidenko, Z.N., Kirnos, M., Vanyushin, B.F.
<2>In vitro DNA methylation by wheat nuclear cytosine DNA methyltransferase: effect of phytohormones.
<3>Gene
<4>157
<5>279-281
<6>1995
<7>Cytosine DNA methyltransferases (MTases) were isolated from nuclei of wheat seedlings and
germinating embryos.  The MTases isolated from both sources were able to perform de novo and
maintenance DNA methylations.  The most purified MTase fraction showed the presence of one
main 67-kDa protein (embryos) and of an 85-kDa protein (in seedlings) in SDS-PAGE.  Some plant
growth regulators (gibberellic acid A3, 6-benzylaminopurine and fusicoccin) elevate by 30-65%
the extent of in vitro DNA methylation by nuclear extracts with a maximal effect at 10 microM
phytohormone concentration.  The same phytohormones do not increase the extent of in vitro
DNA methylation by purified wheat MTase; rather they inhibit it at concentrations of 10/-4 to
10/-5 M.  Thus, DNA methylation in the plant nucleus is controlled by phytohormones.  The
phytohormone effect may be mediated by other proteins in nuclear extracts.

<>

<1>Vlasova, T.I., Kirnos, M.D., Vanyushin, B.F.
<2>Cytosine DNA-methyltransferase from wheat seedlings.
<3>Biokhimiia
<4>61
<5>774-780
<6>1996
<7>A cytosine DNA methyltransferase has been isolated from wheat seedling nuclei and purified by
chromatography on DEAE-cellulose, heparin-Sepharose, and Mono-Q FPLC.  The specific activity
of the enzyme was 236 units/mg protein; its molecular mass by SDS-PAGE was 85-kD.  The enzyme
catalyzes in vitro DNA methylation for a relatively long period, but it is very unstable in
solution in the absence of substrates.  The apparent Km value for S-adenosylmethionine is 6
microM.  The enzyme is active over a wide pH range (5.5-8.5); it is inhibited by NaCl and by
the reaction product S-adenosylhomocysteine with [I]50 values of 0.2M and 12uM, respectively.
The enzyme catalyzes both de novo and maintenance DNA methylation and displays a high rate of
de novo methylation.  The wheat seedling enzyme is similar to known plant cytosine DNA
methyltransferases.

<>

<1>Vlasova, T.I., Vanyushin, B.F.
<2>DNA methylation by wheat cytosine DNA methyltransferase: modulation by protease inhibitor E-64.
<3>Biochem. Mol. Biol. Int.
<4>45
<5>145-153
<6>1998
<7>Cytosine DNA methyltransferase isolated from wheat seedlings and purified in the presence of
metalloprotease and serine protease inhibitors has molecular mass and specific activity equal
to about 85 kDa and 250 units/mg protein, respectively.  Apparent Km for AdoMet and [I]50 for
AdoHcy values are about 6 microM and 12 microM, respectively.  The enzyme is active in a wide
pH range (pH 5.5 - 8.5) and is inhibited by NaCl.  The enzyme rapidly loses its
methyltransferase activity in the absence of substrates.  Using the cysteine protease
inhibitor E-64 it has been shown that rapid enzyme inactivation is caused by disappearance of
essential enzyme SH-groups but is not due to proteolytic enzyme cleavage.

<>

<1>Vlatakis, G., Bouriotis, V.
<2>Affinity partitioning of restriction endonucleases:  Application to the purification of EcoRI and EcoRV.
<3>J. Chromatogr.
<4>538
<5>311-321
<6>1991
<7>Partitioning of restriction endonucleases between two liquid aqueous phases can
be strongly influenced by group-specific ligands included in the two-phase
system.  Three restriction endonucleases, namely EcoRI, EcoRV and BamHI, were
partitioned within an aqueous dextran-polyethylene glycol (PEG) system.  The
enzymes could be extracted into the upper PEG phase by using either triazine
dyes or herring DNA as affinity ligands.  The influence of the endogenous
bacterial nucleic acids, concentration of polymer-bound dye and concentration
of sodium chloride on the system were examined.  A partial purification of
EcoRI (up to 52-fold) and EcoRV (up to 37-fold) was achieved using a
combination of affinity partitioning and ion-exchange chromatography, providing
an extremely fast and economical method for the isolation of restriction
endonucleases free from contaminating nuclease activities.

<>

<1>Vlatakis, G., Bouriotis, V.
<2>Sequence-specific DNA affinity chromatography:  Application to the purification of EcoRI and SphI.
<3>Anal. Biochem.
<4>195
<5>352-357
<6>1991
<7>Several rapid and effective methods have been described to obtain restriction
endonucleases suitable for commercial exploitation.  However, lengthy and
laborious protocols have been necessary to obtain homogeneous enzymes.  We now
report the use of sequence-specific DNA affinity chromatography to purify
restriction endonucleases to near homogeneity.  Restriction endonucleases EcoRI
and SphI from the microorganisms Escherichia coli RY 13 and Streptomyces
phaeochromogenes, respectively, were purified to near homogeneity employing a
two-step procedure which involves DNA-cellulose chromatography and
oligonucleotide-ligand affinity chromatography.

<>

<1>Vlatakis, G., Clark, D., Bouriotis, V.
<2>Isolation and identification of restriction endonuclease BshFI.
<3>Nucleic Acids Res.
<4>17
<5>8882
<6>1989
<7>None

<>

<1>Vlatakis, G., Skarpelis, G., Stratidaki, I., Bouriotis, V.
<2>Dye-ligand chromatography for the resolution and purification of restriction endonucleases.
<3>Appl. Biochem. Biotechnol.
<4>15
<5>201-212
<6>1987
<7>The resolution of restriction endonucleases from the same microorganism is
conventionally achieved by length fractionation protocols.  We now report
effective single-step procedures that exploit dye-ligand chromatography for the
resolution and purification of restriction enzymes.  After suitable initial
screening, we demonstrated that resolution of two restriction activities can be
achieved in one chromatographic step, and further purification can subsequently
be effected using selected dye-adsorbents.  Accordingly, we resolved in one
step, HpaI from HpaII, HindII from HindIII, and SacI from SacII.  Furthermore,
a three-step chromatographic procedure has been developed to purify EcoRV
suitable for commercial exploitation, as judged by the overdigestion and
cut-ligate-recut quality control tests.

<>

<1>Vockler, C.J., Greenfield, P., Tran-Dinh, N., Midgley, D.J.
<2>Draft Genome Sequence of Bacillus pumilus Fairview, an Isolate Recovered from a Microbial Methanogenic Enrichment of Coal Seam Gas Formation Water from Queensland, Australia.
<3>Genome Announcements
<4>2
<5>e00279-14
<6>2014
<7>Despite its global abundance, Bacillus pumilus is poorly studied. The Fairview strain was
obtained from a methanogenic anaerobic coal digester. The draft genome sequence was 3.8 Mbp
long and contained 3,890 protein-coding genes. Like the SAFR-032 strain, it includes B.
pumilus-specific proteins that likely confer enhanced resistance to environmental stresses.

<>

<1>Voelker, L.L., Dybvig, K.
<2>Gene transfer in Mycoplasma arthritidis: Transformation, conjugal transfer of Tn916, and evidence for a restriction system recognizing AGCT.
<3>J. Bacteriol.
<4>178
<5>6078-6081
<6>1996
<7>Mycoplasma arthritidis is a rat pathogen causing a severe polyarthritis.  The study of its
pathogenic mechanisms has been hampered by the lack of genetic systems for use with M.
arthritidis.  Described here are procedures for genetic transformation of M. arthritidis and
conjugal transfer of Tn916 from an enterococcal donor to M. arthritidis.  The location of
Tn916 insertion sites in the mycoplasmal chromosome was random, suggesting that Tn916 may be
useful as an insertional mutagen in this organism.  Additionally, a restriction and
modification system was identified which presented a strong barrier to gene transfer.  For
transformation, the restriction system was circumvented by using DNA that was modified in
vitro with the appropriate site-specific methylase (AluI).

<>

<1>Voelker, L.L., Dybvig, K.
<2>Sequence analysis of the Mycoplasma arthritidis bacteriophage MAV1 genome identifies the putative virulence factor.
<3>Gene
<4>233
<5>101-107
<6>1999
<7>The bacteriophage MAV1 is required for the development of arthritis in rats after infection
with its host Mycoplasma arthritidis.  To identify the phage-encoded virulence factor for this
arthritis, the complete nucleotide sequence of MAV1 was determined. The linear double-stranded
genome of MAV1 is 15644bp and contains 15 ORFs. Putative protein products from these ORFs were
identified by comparison of the deduced amino acid sequences to known proteins and comprise
DNA replication, restriction-modification, structural, regulatory, and integration/excision
proteins. Eight putative promoters were identified; four of these would produce polycistronic
transcripts. Translation of each ORF appears to be initiated independently, with each having
its own RBS. A single ORF, vir, was identified on the minus strand of the phage genome. The
putative protein product of vir contains a classic prokaryotic lipoprotein signal sequence and
is a strong candidate for the MAV1-encoded virulence determinant.

<>

<1>Voet, J.G., Voet, D.
<2>DNA structure and restriction/modification visualized with kinemages.
<3>FASEB J.
<4>11
<5>A843
<6>1997
<7>Teaching nucleic acid structure in more than a cursory way can be difficult.  What is it that
is different about the structures of the A and B conformations of DNA?  How do we identify the
Major and Minor grooves?  How do proteins interact with DNA in a specific manner?  How do
proteins cleave and modify DNA?  Students carry out restriction digestion of DNA and make
restriction maps in introductory biology courses.  But do they actually know what is
happening?  We have tried to remedy this problem by instituting the use of Kinemage
visualizations.  These accompany a set of laboratory experiments derived largely from Micklos,
D.A. and Freyer, G.A., DNA Science, a first course in recombinant DNA Technology, Cold Spring
Harbor Laboratory Press, 1990, on plasmid isolation, restriction mapping, and the effect of
methylation on restriction.  The kinemages we have made show the different conformations of
DNA and the complementary base pairs but, in addition, identify the functional groups of the
bases that line the major and minor grooves and how the different types of sugar puckers
associated with the different conformations.  We have also made kinemages showing the specific
interactions of EcoRI and EcoRV restriction endonucleases with DNA as well as the interactions
of HhaI methylase with DNA.  In this last kinemage we show the mechanism of methylation as
well as the dramatic conformational change in the DNA induced by the enzyme and the
protein-DNA interactions involved.  We will demonstrate these kinemages at the poster session.

<>

<1>Voeykova, T.A., Lomovskaya, N.D.
<2>Restriction and modification of Actinophage omega-C31.
<3>Genetika
<4>17
<5>1132-1135
<6>1981
<7>Actinophage omega-C31 was shown to be insensitive to a number of restriction and modification
systems.  Phage sensitivity to RM systems of those strains to which phage cannot adsorb, may
be tested using protoplast transfection.  For instance, the absence of omega-C13 transfection
in Streptomyces griseus Kr15 protoplasts, as compared to efficient transfection in protoplasts
of R- M+ mutant of this strain seems to imply the sensitivity of omega-C31 to the RM system of
S. griseus Kr15.  Restriction of mutant omega-C31 phage modified by S. albus G RM system has
been detected in S. coelicolor A3(2).  This effect being dependent on a previous host may
indicate that the mutant phage was rendered sensitive to an RM system of S. coelicolor A3(2).

<>

<1>Voeykova, T.A., Slavinskaya, E.V., Orechov, A.V., Lomovskaya, N.D.
<2>Identification of restriction and modification systems in Streptomyces strains.
<3>Genetika
<4>15
<5>1746-1754
<6>1979
<7>Host range of actinophage phi C31, VP5 and Pg81 in respect to 109 strains of Streptomyces
genus and hybrid strain Rcg2 from the cross S. coelicolor
A3(2)XS. griseus Kr was studied. The existence of RM-systems in strains S.
griseus Kr15, S. griseus Kr20, Rcg2, S. griseofovillus 43 was shown using
phage Pg81. Mutants of Pg81 were observed which to some extent lost
snesitivity to RM-system in the strain Rcg2. The presence of RM-system in
S. lividans 67 was demonstrated by the phage VP5.

<>

<1>Vogelsang-Wenke, H., Oesterhelt, D.
<2>Isolation of a halobacterial phage with a fully cytosine-methylated genome.
<3>Mol. Gen. Genet.
<4>211
<5>407-414
<6>1988
<7>A new bacteriophage from Halobacterium halobium has been isolated and partially characterized.
It is not homologous to the phage PhiH (Schnabel et al. 1982) which infects the same
bacterium, though it appeared spontaneously in a culture of PhiH adapted to H. halobium
NRL/JW. The size and morphology of PhiN are comparable to that of other known halophages. The
genome of PhiN consists of linear double-stranded DNA, 56 kb in size, whose dCMP is totally
replaced by 5-methyl-dCMP. This is the second case of a fully cytosine-methylated genome, the
bacteriophage XP12 from Xanthomonas oryzae being so far the only one reported. Like PhiH, the
PhiN genome seems to have terminal redundancy and circular permutation. PhiN is the first
halobacterial phage which survives prolonged exposure to low ionic strength environments.
After 48 h incubation in distilled water a loss in infectivity of less than 50% is observed.

<>

<1>Vogensen, F.K., Pedersen, B.M., Nielsen, E.W., Josephsen, J.
<2>Genetic and biological evidence for 5 different plasmid-encoded restriction and modification systems in Streptococcus cremoris strains.
<3>FEMS Microbiol. Rev.
<4>46
<5>44
<6>1987
<7>From plasmid curing data it was indicated that restriction- and modification systems
(R/M-systems) were plasmid-encoded in Streptococcus cremoris KH, V32.2, T29W5 and Tk5-56 (2
R/M-systems on different plasmids). 4 of the suspected R/M encoding plasmids were purified and
cotransformed together with pVS2 into Streptococcus lactis MG1614. Transformants containing
the individual suspected R/M encoding plasmids together with pVS2, showed restriction against
Phijj50 grown on S. lactis MG1614 containing pVS2, giving genetic evidence that the suspected
plasmids actually coded for R/M systems. The strength in the new host were in two cases lower
than that in the original host. Whether this is due to the host or the different phage is not
known. When Phijj50 were grown on transformants containing the different R/M encoding plasmids
it was in all cases restricted by the other R/M containing transformants, indicating 4
biological different R/M-systems were acting. The size of the 4 plasmids were in the range of
13-18 Kb. Additionally S. cremoris Tk5-56 contained a second plasmid encoded R/M-system. From
curing data and from transformation with total plasmid DNA from S. cremoris Tk5-56 into S.
lactis MG1614 it could be concluded that the second R/M-system is encoded either on
approximately 18 Kb or approximately 45 Kb plasmids. From biological data it can be concluded
that this system differs from the above 4 R/M systems mentioned.

<>

<1>Voget, S., Klippel, B., Daniel, R., Antranikian, G.
<2>Complete Genome Sequence of Carnobacterium sp. 17-4.
<3>J. Bacteriol.
<4>193
<5>3403-3404
<6>2011
<7>Members of the Carnobacteria have been extensively studied as probiotic cultures in
aquacultures and protective cultures in seafood, diary and
meat. We report on the finished genome sequence of Carnobacterium sp.
17-4, which has been isolated from permanently cold seawater. The genetic
information reveals a new circular bacteriocin biosynthesis cluster.

<>

<1>Voget, S., Wemheuer, B., Brinkhoff, T., Vollmers, J., Dietrich, S., Giebel, H.A., Beardsley, C., Sardemann, C., Bakenhus, I., Billerbeck, S., Daniel, R., Simon, M.
<2>Adaptation of an abundant Roseobacter RCA organism to pelagic systems revealed by genomic and transcriptomic analyses.
<3>ISME J.
<4>0
<5>0
<6>2014
<7>The RCA (Roseobacter clade affiliated) cluster, with an internal 16S rRNA gene
sequence similarity of >98%, is the largest cluster of the marine Roseobacter
clade and most abundant in temperate to (sub)polar oceans, constituting up to 35%
of total bacterioplankton. The genome analysis of the first described species of
the RCA cluster, Planktomarina temperata RCA23, revealed that this phylogenetic
lineage is deeply branching within the Roseobacter clade. It shares not >65.7% of
homologous genes with any other organism of this clade. The genome is the
smallest of all closed genomes of the Roseobacter clade, exhibits various
features of genome streamlining and encompasses genes for aerobic anoxygenic
photosynthesis (AAP) and CO oxidation. In order to assess the biogeochemical
significance of the RCA cluster we investigated a phytoplankton spring bloom in
the North Sea. This cluster constituted 5.1% of the total, but 10-31% (mean
18.5%) of the active bacterioplankton. A metatranscriptomic analysis showed that
the genome of P. temperata RCA23 was transcribed to 94% in the bloom with some
variations during day and night. The genome of P. temperata RCA23 was also
retrieved to 84% from metagenomic data sets from a Norwegian fjord and to 82%
from stations of the Global Ocean Sampling expedition in the northwestern
Atlantic. In this region, up to 6.5% of the total reads mapped on the genome of
P. temperata RCA23. This abundant taxon appears to be a major player in ocean
biogeochemistry.The ISME Journal advance online publication, 1 August 2014;
doi:10.1038/ismej.2014.134.

<>

<1>Voigt, J.M., Topal, M.D.
<2>O6 methylguanine in place of guanine causes asymmetric single-strand cleavage of DNA by some restriction enzymes.
<3>Biochemistry
<4>29
<5>1632-1637
<6>1990
<7>The interactions of restriction enzymes with their cognate DNA recognition sequences present a
model for protein-DNA interactions. We have investigated the effect of O6-methylguanine on
restriction enzyme cleavage of DNA; O6-methylguanine is a carcinogenic lesion and a structural
analogue of the biological restriction inhibitor N6-methyladenine. O6-methylguanine was
synthesized into oligonucleotides at unique positions. The oligonucleotides were purified and
analyzed by high-pressure liquid chromatography to assure that, within the limits of our
detection, O6-methylguanine was the only modified base present. These oligonucleotides were
annealed with their complement so that cytosine, and in one case thymine, opposed
O6-methylguanine. DNA cleavage by restriction enzymes that recognize a unique DNA sequence,
HpaII, HhaI, HinPI, NaeI, NarI, PvuII, and XhoI, was inhibited by a single O6-methylguanine in
place of guanine (adenine for PvuII) within the appropriate recognition sequences. However,
only the modified strand was nicked by HpaII, NaeI, and XhoI with O6-methylguanine at certain
positions, indicating asymmetric strand cleavage. For all the restriction enzymes studied but
AhaII, BanI, and NarI, lack of double- or single-strand cleavage correlated with inability of
the O6-methylguanine-containing recognition sequence to measurably bind enzyme. None of the
restriction enzymes studied were inhibited by O6-methylguanine outside their cognate
recognition sequences.

<>

<1>Volland, S., Rachinger, M., Strittmatter, A., Daniel, R., Gottschalk, G., Meyer, O.
<2>Complete genome sequences of the chemolithoautotrophic Oligotropha carboxidovorans strains OM4 and OM5.
<3>J. Bacteriol.
<4>193
<5>5043
<6>2011
<7>We report on genome sequencing of Oligotropha carboxidovorans strain OM4 and resequencing of
strain OM5. The genomes of both are composed of one
chromosome and two plasmids. The presence of two plasmids in the OM5
genome is inconsistent with the previously published sequence in which
only one plasmid was described (D. Paul, S. Bridges, S. Burgess, Y.
Dandass, and M. Lawrence. BMC Genomics 11:511, 2010).

<>

<1>Volna, P., Jarjour, J., Baxter, S., Roffler, S.R., Monnat, R.J. Jr., Stoddard, B.L., Scharenberg, A.M.
<2>Flow cytometric analysis of DNA binding and cleavage by cell surface-displayed homing endonucleases.
<3>Nucleic Acids Res.
<4>35
<5>2748-2758
<6>2007
<7>LAGLIDADG homing endonucleases (LHEs) cleave 18-24 bp DNA sequences and are promising enzymes
for applications requiring sequence-specific DNA cleavage amongst genome-sized DNA
backgrounds. Here, we report a method for cell surface display of LHEs, which facilitates
analysis of their DNA binding and cleavage properties by flow cytometry. Cells expressing
surface LHEs can be stained with fluorescently conjugated double-stranded oligonucleotides
(dsOligos) containing their respective target sequences. The signal is absolutely sequence
specific and undetectable with dsOligos carrying single base-pair substitutions. LHE-dsOligo
interactions facilitate rapid enrichment and viable recovery of rare LHE expressing cells by
both fluorescence-activated cell sorting (FACS) and magnetic cell sorting (MACS).
Additionally, dsOligos conjugated with unique fluorophores at opposite termini can be tethered
to the cell surface and used to detect DNA cleavage. Recapitulation of DNA binding and
cleavage by surface-displayed LHEs provides a high-throughput approach to library screening
that should facilitate rapid identification and analysis of enzymes with novel sequence
specificities.

<>

<1>von Gabain, A., Nagy, E., Meinke, A., Selak, S., Hanner, M., Smidt, M., Weissgram, M., Noiges, B., Seidel, S., Bacher, J., Satke, C., Schueler, W., Oleksiewicz, M.
<2>Nontypable haemophilus influenzae antigens.
<3>International Patent Office
<4>WO 2010092176 A
<5>
<6>2010
<7>The present invention relates to isolated nucleic acid molecules which encode an antigen from
a nontypable Haemophilus influenzae species, a vector which comprises such nucleic acid
molecule, and a host cell comprising such vector.  Furthermore, the invention provides
antigens from a nontypable Haemophilus influenzae species, as well as fragments and variants
thereof, a process for producing such antigens, and a process for producing a cell, which
expresses such antigen.  More specifically such antigens are produced by or associated with
bacterial infections caused by nontypable Haemophilus influenzae. Moreover, the present
invention provides antibodies binding to such antigen, a hybridoma cell producing such
antibodies, methods for producing such antibodies, a pharmaceutical composition comprising
such nucleic acid molecule, antigen, vector or antibody, the use of such nucleic acid
molecule, antigen, vector or antibody for the preparation of a pharmaceutical composition,
methods for identifying an antagonist capable of binding such antigen or of reducing or
inhibiting the interaction activity of such antigen, methods for diagnosing an infection with
nontypable Haemophilus influenzae and methods for the treatment or prevention of an infection
with nontypable Haemophilus influenzae.

<>

<1>von Jan, M. et al.
<2>Complete genome sequence of Archaeoglobus profundus type strain (AV18).
<3>Standards in Genomic Sciences
<4>2
<5>327-346
<6>2010
<7>Archaeoglobus profundus (Burggraf et al. 1990) is a hyperthermophilic archaeon in the
euryarchaeal class Archaeoglobi, which is currently represented by the single
family Archaeoglobaceae, containing six validly named species and two strains
ascribed to the genus 'Geoglobus' which is taxonomically challenged as the
corresponding type species has no validly published name. All members were
isolated from marine hydrothermal habitats and are obligate anaerobes. Here we
describe the features of the organism, together with the complete genome sequence
and annotation. This is the second completed genome sequence of a member of the
class Archaeoglobi. The 1,563,423 bp genome with its 1,858 protein-coding and 52
RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Vongs, A., Kakutani, T., Martienssen, R.A., Richards, E.J.
<2>Arabidopis thaliana DNA methylation mutants.
<3>Science
<4>260
<5>1926-1928
<6>1993
<7>Three DNA hypomethylation mutants of the flowering plant Arabidopsis thaliana were isolated by
screening mutagenized populations for plants containing centromeric repetitive DNA arrays
susceptible to digestion by restriction endonuclease that was sensitive to methylated
cytosines. The mutations are recessive, and at least two are alleles of a single locus,
designated DDM1 (for decrease in DNA methylation). Amounts of 5-methylcytosine were reduced
over 70 percent in ddm1 mutants. Despite this reduction in DNA methylation levels, ddm1
mutants developed normally and exhibited no striking morphological phenotypes. However, the
ddm1 mutations are associated with a segregation distortion phenotype. The ddm1 mutations were
used to demonstrate that de novo DNA methylation in vivo is slow.

<>

<1>Voo, K., Carlone, D., Jacobsen, B., Flodin, A., Skalnik, D.
<2>Cloning of a mammalian transcriptional activator that binds unmethylated CpG motifs and shares a CXXC domain with DNA methyltransferase, human trithorax, and methyl-CpG binding domain protein 1.
<3>Blood
<4>0
<5>469a
<6>1999
<7>Cytosine methylation of DNA at CpG motifs provides an important mechanism for controlling gene
expression.  Hypermethylation or hypomethylation of tumor suppressor genes and oncogenes,
respectively, has been associated with the progression of cancer.  Ligand screening was
utilized to isolate a human cDNA that encodes a novel CpG binding protein (hCGBP) that is
highly expressed in hematopoietic cell lines.  This factor contains three cysteine rich
domains, two of which exhibit homology to the plant homeodomain finger motif.  A third
cysteine rich domain conforms to the CXXC motif identified in DNA methyltransferase, human
thithorax, and methyl-CpG binding domain protein 1.  Interestingly, database searching
revealed that the hCGBP gene is located within 1 kilobase of the gene encoding methyl-CpG
binding domain protein 1, a factor that binds to methylated CpG motifs and functions as a
transcriptional repressor.  A fragment of hCGBP that contains the CXXC domain binds to an
oligonucleotide probe containing a single CpG site, and this complex is disrupted by distinct
oligonucleotide competitors that also contain CpG motif(s).  However, hCGBP fails to bind
oligonucleotides in which the CpG motif is either mutated or methylated.  Native hCGBP is
detected as an 88 kDa protein by Western analysis and is ubiquitously expressed.  The
DNA-binding activity of native hCGBP is apparent in electrophoretic mobility shift assays, and
hCGBP trans-activates promoters that contain CpG motifs, but not promoters in which the CpG is
ablated.  These data indicate that hCGBP is a transcriptional activator that recognizes
unmethylated CpG dinucleotides, suggesting a role in modulating the expression of genes
located within CpG islands.

<>

<1>Vorholt, J.A., Vaupel, M., Thauer, R.K.
<2>A selenium-dependent and a selenium-independent formylmethanofuran dehydrogenase and their transcriptional regulation in the hyperthermophilic Methanopyrus kandleri.
<3>Mol. Microbiol.
<4>23
<5>1033-1042
<6>1997
<7>The genome of Methanopyrus kandleri was found to harbour a gene, fwuB, predicted to encode the
catalytic subunit of a tungsten formylmethanofuran dehydrogenase with an active site
selenocysteine, and a second gene, fwcB, encoding a tungsten formylmethanofuran dehydrogenase
with an active site cysteine. Northern blot and primer-extension analysis revealed that both
genes were differentially transcribed. During growth of the methanogen on medium supplemented
with selenium only fwuB was transcribed, whereas transcription of both fwuB and fwcB was
observed on selenium-deprived medium. Growth of M. kandleri was stimulated by tungstate and
selenite but not by molybdate. The findings indicate that the hyperthermophilic archaeon
contains two tungsten isoenzymes of formylmethanofuran dehydrogenase, one of which is a novel
selenium enzyme. They also indicate that the hyperthermophilic methanogen probably does not
contain a molybdenum formylmethanofuran dehydrogenase which appears to be present only in
thermophilic and mesophilic methanogens.

<>

<1>Vorholter, F.J., Arnold, M., Wibberg, D., Blom, J., Winkler, A., Viehoever, P., Albersmeier, A., Goesmann, A., Zange, S., Heesemann, J., Puhler, A., Hogardt, M.
<2>Draft Genome Sequence of Pseudomonas aeruginosa Strain WS394, a Multidrug-Resistant and Highly Cytotoxic Wound Isolate from Chronic Ulcus Cruris.
<3>Genome Announcements
<4>2
<5>e01325-14
<6>2014
<7>Pseudomonas aeruginosa is a frequent human pathogen that increasingly causes chronic
infections of nonhealing wounds. Here we present the 6.8 Mb draft genome
of strain WS394, a multidrug-resistant chronic ulcer isolate that exhibited
outstanding high cell cytotoxicity despite defective secretion of exotoxin U,
suggesting a habitat-dependent adaptation process.

<>

<1>Vorholter, F.J., Schneiker, S., Goesmann, A., Krause, L., Bekel, T., Kaiser, O., Linke, B., Patschkowski, T., Ruckert, C., Schmid, J., Sidhu, V.K., Sieber, V., Tauch, A., Watt, S.A., Weisshaar, B., Becker, A., Niehaus, K., Puhler, A.
<2>The genome of Xanthomonas campestris pv. campestris B100 and its use for the reconstruction of metabolic pathways involved in xanthan biosynthesis.
<3>J. Biotechnol.
<4>134
<5>33-45
<6>2008
<7>The complete genome sequence of the Xanthomonas campestris pv. campestris strain B100 was
established. It consisted of a chromosome of 5,079,003bp,
with 4471 protein-coding genes and 62 RNA genes. Comparative genomics
showed that the genes required for the synthesis of xanthan and xanthan
precursors were highly conserved among three sequenced X. campestris pv.
campestris genomes, but differed noticeably when compared to the remaining
four Xanthomonas genomes available. For the xanthan biosynthesis genes
gumB and gumK earlier translational starts were proposed, while gumI and
gumL turned out to be unique with no homologues beyond the Xanthomonas
genomes sequenced. From the genomic data the biosynthesis pathways for the
production of the exopolysaccharide xanthan could be elucidated. The first
step of this process is the uptake of sugars serving as carbon and energy
sources wherefore genes for 15 carbohydrate import systems could be
identified. Metabolic pathways playing a role for xanthan biosynthesis
could be deduced from the annotated genome. These reconstructed pathways
concerned the storage and metabolization of the imported sugars. The
recognized sugar utilization pathways included the Entner-Doudoroff and
the pentose phosphate pathway as well as the Embden-Meyerhof pathway
(glycolysis). The reconstruction indicated that the nucleotide sugar
precursors for xanthan can be converted from intermediates of the pentose
phosphate pathway, some of which are also intermediates of glycolysis or
the Entner-Doudoroff pathway. Xanthan biosynthesis requires in particular
the nucleotide sugars UDP-glucose, UDP-glucuronate, and GDP-mannose, from
which xanthan repeat units are built under the control of the gum genes.
The updated genome annotation data allowed reconsidering and refining the
mechanistic model for xanthan biosynthesis.

<>

<1>Vorobeva, O.V., Karyagina, A.S., Volkov, E.M., Viryasov, M.B., Oretskaya, T.S., Kubareva, E.A.
<2>An analysis of methyltransferase SsoII-DNA contacts in the enzyme-substrate complex.
<3>Bioorg. Khim.
<4>28
<5>402-410
<6>2002
<7>The functional groups of the DNA methylation site that are involved in the DNA interaction
with methyltransferase SsoII at the recognition stage were identified. The contacts in the
enzyme-substrate complex were analyzed in the presence of S-adenosyl-L-homocysteine using the
interference footprinting assay with formic acid, hydrazine, dimethyl sulfate, or
N-ethyl-N-nitrosourea as a modifying reagent. It was shown that the replacement of the central
A.T by the G.C pair in the methylation site did not affect the enzyme-DNA interaction, whereas
the use of a substrate with one chain methylated (monomethylated substrate) instead of the
unmethylated substrate dramatically changes the DNA contacts. The binding constants of
unmethylated and monomethylated substrates with methyltransferase Ssoll in the presence of
S-adenosyl-L-homocysteine were calculated.

<>

<1>Vorobeva, O.V., Romanenkov, A.S., Metelev, V.G., Karyagina, A.S., Lavrova, N.V., Oretskaya, T.S., Kubareva, E.A.
<2>Crosslinking of Cys142 of Methyltransferase SsoII with DNA Duplexes Containing a Single Internucleotide Phosphoryldisulfide Link.
<3>Mol. Biol. (Mosk)
<4>37
<5>906-915
<6>2003
<7>DNA duplexes containing a single phosphoryldisulfide link in place of the natural
internucleotide phosphodiester bond were employed in affinity
modification of Cys142 in cytosine-C5 DNA methyltransferase SsoII
(M.SsoII). The possibility of duplex-M.SsoII conjugation as a result of
disulfide exchange was demonstrated. The crosslinking efficiency proved to
depend on the DNA primary structure, modification position, and the
presence of S-adenosyl-L-homocysteine, a nonreactive analog of the
methylation cofactor. The SH group of M.SsoII Cys142 was assumed to be
close to the DNA sugar-phosphate backbone in the DNA-enzyme complex.

<>

<1>Vorobeva, O.V., Vasilev, S.A., Karyagina, A.S., Oretskaya, T.S., Kubareva, E.A.
<2>Analysis of DNA-protein contacts in a complex between methyltransferase SsoII and a promoter region of the SsoII restriction-modification genes.
<3>Mol. Biol. (Mosk)
<4>34
<5>1074-1080
<6>2000
<7>.

<>

<1>Voros, A., Horvath, B., Hunyadkurti, J., McDowell, A., Barnard, E., Patrick, S., Nagy, I.
<2>Complete Genome Sequences of Three Propionibacterium acnes Isolates from the Type IA2 Cluster.
<3>J. Bacteriol.
<4>194
<5>1621-1622
<6>2012
<7>Propionibacterium acnes is an anaerobic Gram-positive bacterium that has been linked to a wide
range of opportunistic human infections and conditions, most
notably acne vulgaris (I. Kurokawa et al., Exp. Dermatol. 18:821-832, 2009). We
now present the whole-genome sequences of three P. acnes strains from the type
IA(2) cluster which were recovered from ophthalmic infections (A. McDowell et
al., Microbiology 157:1990-2003, 2011).

<>

<1>Vosberg, H.-P., Eckstein, F.
<2>Effect of deoxynucleoside phosphorothioates incorporated in DNA on cleavage by restriction enzymes.
<3>J. Biol. Chem.
<4>257
<5>6595-6599
<6>1982
<7>DNA synthesized in vitro using deoxynucleoside phosphorothioates as substrates is quite
similar to normal DNA in its biochemical properties. In order to investigate the effect of
phosphorothioate groups in DNA on the cleavage pattern of restriction endonucleases
phosphorothioate double-stranded, circular, replicative form of fd DNA was synthesized in
vitro with Escherichia coli DNA polymerase I using native single-stranded DNA as template and
mixtures of three normal nucleotides and one nucleoside phosphorothioate analogue as
substrates. The double-stranded products were hybrids with respect to their phosphorothioate
content. Restriction analysis of normal and phosphorothioate DNA with the restriction
endonucleases HaeIII, BamHI, HpaII, HindII, Alu I, and TaqI showed that the enzymes were
inhibited to different degrees depending on which of the nucleotides was replaced by the
phosphorothioate. Most significant, inhibition was seen throughout with those DNAs which
contained a phosphorothioate exactly at the cleavage site. Phosphorothioate substitutions at
other positions, but still within the recognition sequences, were, except for AluI, not or
weakly inhibitory. Phosphorothioate nucleotides not present in the recognition sequences did
not affect at all the fragment patterns. The results show that recognition sequences of
restriction endonucleases can be selectively protected against cleavage by base-specific
introduction of phosphorothioate groups into DNA.

<>

<1>Voss, B., Bolhuis, H., Fewer, D.P., Kopf, M., Moke, F., Haas, F., El-Shehawy, R., Hayes, P., Bergman, B., Sivonen, K., Dittmann, E., Scanlan, D.J., Hagemann, M., Stal, L.J., Hess, W.R.
<2>Insights into the physiology and ecology of the brackish-water-adapted Cyanobacterium Nodularia spumigena CCY9414 based on a genome-transcriptome analysis.
<3>PLoS ONE
<4>8
<5>E60224
<6>2013
<7>Nodularia spumigena is a filamentous diazotrophic cyanobacterium that dominates
the annual late summer cyanobacterial blooms in the Baltic Sea. But N. spumigena
also is common in brackish water bodies worldwide, suggesting special adaptation
allowing it to thrive at moderate salinities. A draft genome analysis of N.
spumigena sp. CCY9414 yielded a single scaffold of 5,462,271 nucleotides in
length on which genes for 5,294 proteins were annotated. A subsequent
strand-specific transcriptome analysis identified more than 6,000 putative
transcriptional start sites (TSS). Orphan TSSs located in intergenic regions led
us to predict 764 non-coding RNAs, among them 70 copies of a possible
retrotransposon and several potential RNA regulators, some of which are also
present in other N2-fixing cyanobacteria. Approximately 4% of the total coding
capacity is devoted to the production of secondary metabolites, among them the
potent hepatotoxin nodularin, the linear spumigin and the cyclic nodulapeptin.
The transcriptional complexity associated with genes involved in nitrogen
fixation and heterocyst differentiation is considerably smaller compared to other
Nostocales. In contrast, sophisticated systems exist for the uptake and
assimilation of iron and phosphorus compounds, for the synthesis of compatible
solutes, and for the formation of gas vesicles, required for the active control
of buoyancy. Hence, the annotation and interpretation of this sequence provides a
vast array of clues into the genomic underpinnings of the physiology of this
cyanobacterium and indicates in particular a competitive edge of N. spumigena in
nutrient-limited brackish water ecosystems.

<>

<1>Vovis, G.F., Horiuchi, K., Hartman, N., Zinder, N.D.
<2>Restriction endonuclease B and f1 heteroduplex DNA.
<3>Nature New Biol.
<4>246
<5>13-16
<6>1973
<7>In many strains of Escherichia coli, strain-specific restriction system
recognises phage and bacterial DNA synthesis in a different strain and
initiates its degradation.  The recognition and initial hydrolysis can
apparently occur at the same or different sites on the DNA molecule (resistance
transfer factor R1, (refs 1-3) and E. coli B4 restriction, respectively) and
are mediated by a strain-specific endonuclease.  Protection is afforded by
modification (glucosylation and methylation) and mutation of the DNA,
presumably at the recognition site(s).  A DNA duplex, only one of whose strands
is modified (by methylation) is resistant to restriction endonuclease K.
Similar conclusions about the sensitivity of hybrid molecules containing one
modified DNA strand were made from in vivo observations with T2 and lambda
phage with regard to restriction in E. coli B and P1 lysogens, respectively.

<>

<1>Vovis, G.F., Horiuchi, K., Zinder, N.D.
<2>Endonuclease R.EcoRII restriction of bacteriophage f1 DNA in vitro:  Ordering of genes V and VII, location of an RNA promotor for Gene VIII.
<3>J. Virol.
<4>16
<5>674-684
<6>1975
<7>Replicative form DNA of bacteriophage f1 was found to be sensitive in vitro to
restriction by endonuclease R.EcoRII if the DNA was isolated from an
Escherichia coli strain deficient in cytosine methylase activity.  A similar
observation was previously made with DNA from the closely related bacteriophage
fd (S. Schlagman, S. Hattman, M.S. May and L. Berger, submitted for
publication).  The two DNA fragments produced by the endo R.EcoRII digestion of
f1 DNA were localized on the f1 cleavage map and their genetic content was
determined.  The polypeptides synthesized in a "coupled"
transcription-translation system under the direction of each RII fragment were
examined.  The results of such experiments allow the ordering of genes V and
VII and indicate the location of a RNA promoter for gene VIII.

<>

<1>Vovis, G.F., Horiuchi, K., Zinder, N.D.
<2>Kinetics of methylation of DNA by a restriction endonuclease from Escherichia coli B.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>71
<5>3810-3813
<6>1974
<7>The restriction endonuclease from E. coli B is both an endonuclease and a DNA
methylase.  Both activities either require or are stimulated by Mg++, adenosine
triphosphate, and S-adenosyl-L-methionine.  The particular activity which the
enzyme exhibits depends upon the nature of the SB sites, the genetic sites that
identify substrate DNA.  Enzymatic treatment of DNA that has an unmodified,
wild-type SB site results in either rapid restriction of the DNA or very slow
methylation of the SB site.  On the other hand, a hybrid SB site (modified),
which protects the DNA molecule from restriction, results in rapid methylation
of that SB site.

<>

<1>Vovis, G.F., Lacks, S.
<2>Complementary action of restriction enzymes Endo R.DpnI and Endo R.DpnII on bacteriophage f1 DNA.
<3>J. Mol. Biol.
<4>115
<5>525-538
<6>1977
<7>Bacteriophage f1 duplex DNA was isolated from Escherichia coli strains containing different
DNA methylases and assayed for its sensitivity to endonucleolytic cleavage by the enzymes endo
R.DpnI and endo R.DpnII. the former enzyme is specific for methylated DNA, the latter for
unmethylated DNA (Lacks & Greenberg, 1975). The E. coli dam methylase was found to be
responsible for making f1 resistant to endo R.DpnII and sensitive to endo R.DpnI. Endo R.DpnI
cleaved f1 DNA from dam+ cells at four sites. Additional methylation by enzymes other than the
dam methylase gave no further cleavage. Endo R.DpnII cleaved f1 DNA from dam- cells also at
four sites to give restriction fragments identical to those obtained with endo R.DpnI
cleavage. Thus, the two enzymes are complementary in that they recognize and cleave within the
same DNA sequence, one if the DNA is methylated, the other if it is unmethylated. DNA duplexes
containing one methylated strand (dam+) and one unmethylated strand (dam-) were prepared in
vitro. These methylated hybrids were refractory to endonucleolytic cleavage by both endo
R.DpnI and endo R.DpnII. Neither enzyme, therefore, appears to make even a single strand break
at a methylated/unmethylated hybrid site.

<>

<1>Vovis, G.F., Zinder, N.D.
<2>Methylation of f1 DNA by a restriction endonuclease from Escherichia coli B.
<3>J. Mol. Biol.
<4>95
<5>557-568
<6>1975
<7>Bacteriophage f1 duplex DNA containing hybrid SB sites, the genetic sites which
confer upon DNA sensitivity to Escherichia coli B-specific restriction and
modification, were prepared in vitro.  The hybrid SB sites (modified and
mutant) were tested for their ability to be methylated in vitro by endonuclease
R.EcoB, the enzyme responsible for both B-specific restriction and modification
in vivo.  DNA containing hybrid (modified) SB sites can be methylated.  One
methyl group is added to the DNA per hybrid (modified) SB site.  On the other
hand, DNA containing hybrid (mutant) SB sites is refractory to modification.
The nature and the function of the SB site as well as the implications of these
observations for f1 recombination are discussed.

<>

<1>Voykov, Y.N., Repin, V.E., Degtyarev, S.K., Viesture, V.A., Stengrevitze, O.A.
<2>Manufacture of restriction endonuclease PstI from Providencia stuartii.
<3>Soviet Patent Office
<4>SU 1472493 A
<5>
<6>1989
<7>The present invention is directed to produce the restriction enzyme PstI with much greater
yield from Providencia stuartii. The yield of PstI is increased from 400-600 units/g cells to
40,000-45,000 units/g cells. Cells are very stable (can be stored for 5 years without
selection).

<>

<1>Vranken, C., Deen, J., Dirix, L., Stakenborg, T., Dehaen, W., Leen, V., Hofkens, J., Neely, R.K.
<2>Super-resolution optical DNA Mapping via DNA methyltransferase-directed click chemistry.
<3>Nucleic Acids Res.
<4>42
<5>e50
<6>2014
<7>We demonstrate an approach to optical DNA mapping, which enables near single-molecule
characterization of whole bacteriophage genomes. Our approach
uses a DNA methyltransferase enzyme to target labelling to specific sites and
copper-catalysed azide-alkyne cycloaddition to couple a fluorophore to the DNA.
We achieve a labelling efficiency of approximately 70% with an average labelling
density approaching one site every 500 bp. Such labelling density bridges the gap
between the output of a typical DNA sequencing experiment and the long-range
information derived from traditional optical DNA mapping. We lay the foundations
for a wider-scale adoption of DNA mapping by screening 11 methyltransferases for
their ability to direct sequence-specific DNA transalkylation; the first step of
the DNA labelling process and by optimizing reaction conditions for fluorophore
coupling via a click reaction. Three of 11 enzymes transalkylate DNA with the
cofactor we tested (a readily prepared s-adenosyl-l-methionine analogue).

<>

<1>Vuilleumier, S. et al.
<2>Methylobacterium genome sequences: a reference blueprint to investigate microbial metabolism of C1 compounds from natural and industrial sources.
<3>PLoS ONE
<4>4
<5>E5584
<6>2009
<7>BACKGROUND: Methylotrophy describes the ability of organisms to grow on
reduced organic compounds without carbon-carbon bonds. The genomes of two
pink-pigmented facultative methylotrophic bacteria of the
Alpha-proteobacterial genus Methylobacterium, the reference species
Methylobacterium extorquens strain AM1 and the dichloromethane-degrading
strain DM4, were compared. METHODOLOGY/PRINCIPAL FINDINGS: The 6.88 Mb
genome of strain AM1 comprises a 5.51 Mb chromosome, a 1.26 Mb megaplasmid
and three plasmids, while the 6.12 Mb genome of strain DM4 features a 5.94
Mb chromosome and two plasmids. The chromosomes are highly syntenic and
share a large majority of genes, while plasmids are mostly
strain-specific, with the exception of a 130 kb region of the strain AM1
megaplasmid which is syntenic to a chromosomal region of strain DM4. Both
genomes contain large sets of insertion elements, many of them
strain-specific, suggesting an important potential for genomic plasticity.
Most of the genomic determinants associated with methylotrophy are nearly
identical, with two exceptions that illustrate the metabolic and genomic
versatility of Methylobacterium. A 126 kb dichloromethane utilization
(dcm) gene cluster is essential for the ability of strain DM4 to use DCM
as the sole carbon and energy source for growth and is unique to strain
DM4. The methylamine utilization (mau) gene cluster is only found in
strain AM1, indicating that strain DM4 employs an alternative system for
growth with methylamine. The dcm and mau clusters represent two of the
chromosomal genomic islands (AM1: 28; DM4: 17) that were defined. The mau
cluster is flanked by mobile elements, but the dcm cluster disrupts a gene
annotated as chelatase and for which we propose the name "island
integration determinant" (iid). CONCLUSION/SIGNIFICANCE: These two genome
sequences provide a platform for intra- and interspecies genomic
comparisons in the genus Methylobacterium, and for investigations of the
adaptive mechanisms which allow bacterial lineages to acquire
methylotrophic lifestyles.

<>

<1>Vuilleumier, S. et al.
<2>Genome Sequence of the Haloalkaliphilic Methanotrophic Bacterium Methylomicrobium alcaliphilum 20Z.
<3>J. Bacteriol.
<4>194
<5>551-552
<6>2012
<7>Methylomicrobium strains are widespread in saline environments. Here, we report the complete
genome sequence of Methylomicrobium alcaliphilum 20Z,
a haloalkaliphilic methanotrophic bacterium, which will provide the basis
for detailed characterization of the core pathways of both single-carbon
metabolism and responses to osmotic and high-pH stresses. Final assembly
of the genome sequence revealed that this bacterium contains a 128-kb
plasmid, making M. alcaliphilum 20Z the first methanotrophic bacterium of
known genome sequence for which a plasmid has been reported.

<>

<1>Vuilleumier, S., Nadalig, T., Ul-Haque, M.F., Magdelenat, G., Lajus, A., Roselli, S., Muller, E.E., Gruffaz, C., Barbe, V., Medigue, C., Bringel, F.
<2>Complete Genome Sequence of the Chloromethane-Degrading Hyphomicrobium sp. Strain MC1.
<3>J. Bacteriol.
<4>193
<5>5035-5036
<6>2011
<7>Hyphomicrobium sp. strain MC1 is an aerobic methylotroph originally isolated from industrial
sewage. This prosthecate bacterium was the first
strain reported to grow with chloromethane as the sole carbon and energy
source. Its genome, consisting of a single 4.76-Mb chromosome, is the
first for a chloromethane-degrading bacterium to be formally reported.

<>

<1>Vyle, J.S., Connolly, B.A., Kemp, D., Cosstick, R.
<2>Sequence- and strand-specific cleavage in oligodeoxyribonucleotides and DNA containing 3'-thiothymidine.
<3>Biochemistry
<4>31
<5>3012-3018
<6>1992
<7>Oligonucleotides containing a 3'-thiothymidine residue (T3's) at the cleavage site for the
EcoRV restriction endonuclease (between the central T and A residues of the sequence GATATC)
have been prepared on an automated DNA synthesizer using 5'-O-monomethoxytritylthymidine
3'-S-(2-cyanoethyl N,N-di-isopropylphosphorothioamidite).  The self-complementary sequence
GACGAT3'sATCGTC was completely resistant to cleavage by EcoRV, while the heteroduplex
composed of 5'-TCTGAT3'sATCCTC and 5'-GAGGATATCAGA (duplex 4) was cleaved only in the
unmodified strand (5'-GAGGATATCAGA).  In contrast, strands containing a
3'-S-phosphorothiolate linkage could be chemically cleaved specifically at this site with
Ag+.  A T3's residue has also been incorporated in the (-) strand of double-stranded closed
circular (RF IV) M13mp18 DNA at the cleavage site of a unique EcoRV recognition sequence by
using 5'-pCGAGCTCGAT3'sATCGTAAT as a primer for polymerization on the template (+) strand of
M13mp18 DNA.  On treatment of this substrate with EcoRV, only one strand was cleaved to
produce the RFII or nicked DNA.  Taken in conjunction with the cleavage studies on the
oligonucleotides, this result demonstrates that the 3'-S-phosphorothiolate linkage is
resistant to scission by EcoRV.  Additionally, the phosphorothiolate-containing strand of the
M13mp18 DNA could be cleaved specifically at the point of modification using iodine in aqueous
pyridine.  The combination of enzymatic and chemical techniques provides, for the first time,
a demonstrated method for the sequence-specific cleavage of either the (+) or (-) strand.

<>

<1>Waalwijk, C., Flavell, R.A.
<2>MspI, an isochizomer of HpaII which cleaves both unmethylated and methylated HpaII sites.
<3>Nucleic Acids Res.
<4>5
<5>3231-3236
<6>1978
<7>The cleavage of DNA by restriction endonucleases HpaII and HapII is prevented
by the presence of a 5-methyl group at the internal C residue of its
recognition sequence CCGG.  MspI, an isoschizomer of HpaII available from New
England Biolabs, cleaves DNA irrespective of the presence of a methyl group at
this position.  This enzyme cleaves DNA from Haemophilus parainfluenzae and
Haemophilus aphrophilus readily while HpaII and HapII cannot degrade these
DNAs.  Practically all HpaII sites in mammalian sperm DNA are also protected by
methylation at the internal C position since HpaII and HapII barely cleave this
DNA (average molecular weight 40 kb).  MspI, however, cleaves the DNA to an
average size of about 5 kb.

<>

<1>Wachsman, J.T.
<2>DNA methylation and the association between genetic and epigenetic changes: relation to carcinogenesis.
<3>Mutat. Res.
<4>375
<5>1-8
<6>1997
<7>This paper examines the relationship between DNA mutagenic lesions, DNA methylation and the
involvement of these changes in the process of carcinogenesis.  Many types of DNA damage
(oxidative lesions, alkylation of bases, abasic sites, photodimers, etc.) interfere with the
ability of mammalian cell  DNA to be methylated at CpG dinucleotides by DNA-methyltransferases
(DNA-MTases).  This can result in altered patterns in the distribution of 5-methylcytosine
(5MeC) residues at CpG sites.  Methylation of DNA is an epigenetic change that by definition
is heritable, can result in changes in chromatin structure, and is often accompanied by
modified patterns of gene expression.  The presence of 5MeC in DNA, as well as oxidative
stress induced by the free radical nitric oxide, can interfere with the repair of alkylation
damage, thereby increasing the level fo potentially mutagenic lesions. CpG sites in DNA
represent mutational hotspots, with both the presence of 5MeC in DNA and the catalytic
activity of DNA-MTases being intrinsically mutagenic.  The process of carcinogenesis has
frequently been associated with an increased expression of DNA-MTase activity, accompanied by
either hypermethylation or hypomethylation of target cell (progenitor tumor cell) DNA.  In
addition, there is evidence that overexpression of DNA-MTase activity could result in
increased cytosine methylation at non-CpG sites.  A variety of chemicals can alter the extent
of DNA methylation in mammalian cells.  These include inhibitors of topisomerase II, as well
as inhibitors of DNA synthesis, microtubule formation, histone deacetylation,
transmethylation, etc.  Genetic and epigenetic changes in DNA have a profound influence on one
another and could play a major role in the process of carcinogenesis, by modulating both the
extent and the pattern of gene expression.

<>

<1>Wada, Y.
<2>Separate analysis of complementary strands of restriction enzyme-digested DNA.  An application of restriction fragment mass mapping by matrix-assisted laser desorption/ionization mass spectrometry.
<3>J. Mass Spectrom.
<4>33
<5>187-192
<6>1998
<7>Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry   MALDI/TOF-MS)
of a restriction endonuclease digest determines the molecular mass of PCR-amplified DNA more
easily than measurement of undigested DNA. With this method, a 664 bp region from the FAS gene
could be analyzed and a two-nucleotide deletion in the L1CAM gene was detected in a
restriction fragment of 105 nucleotides. Furthermore, the analysis of smaller fragments
allowed separate detection of single-stranded oligonucleotides comprising individual digested
fragments. This mixture analysis of restriction enzyme digests improves the resolution,
sensitivity and accuracy of MALDI/TOF-MS of DNA and is thus expected to facilitate its
application to genetic diagnosis.

<>

<1>Wada, Y.
<2>Physiological functions of plant DNA methyltransferases.
<3>Plant Biotechnol.
<4>22
<5>71-80
<6>2005
<7>Epigenetic regulation is defined as mechanisms that control gene expression without altering
base sequences.  Cytosine methylation, chromatin remodeling, and modifications at the
N-termini of core histones are key factors in this regard.  Epigenetic modifications are found
throughout the eukaryotes, suggesting that they developed at an early stage in biological
evolution, although actual molecular mechanisms show considerable variation among species.  In
particular, plants are unique in establishment and maintenance of epigenetic states, as
examplified by species-specific enzymes that catalyze DNA methylation.  Since the function and
diversity of DNA methyltransferases in individual species are not fully understood, I here
summarize recent findings in plant epigenetics, focusing on DNA methyltransferases classified
into three major groups.  Their possible biological functions are also discussed with
reference to histone modification and chromatin remodeling.

<>

<1>Wada, Y., Ohya, H., Yamaguchi, Y., Koizumi, N., Sano, H.
<2>Preferential de novo methylation of cytosine residues in non-CpG sequences by a domains rearranged DNA methyltransferase from tobacco plants.
<3>J. Biol. Chem.
<4>278
<5>42386-42393
<6>2003
<7>In plant DNA, cytosines in symmetric CpG and CpNpG (N is A, T, or C) are thought to be
methylated by DNA methyltransferases, MET1 and CMT3,
respectively. Cytosines in asymmetric CpNpN are also methylated, and
genetic analysis has suggested the responsible enzyme to be domains
rearranged methyltransferase (DRM). We cloned a tobacco cDNA, encoding a
novel protein consisting of 608 amino acids, that resembled DRMs found in
maize and Arabidopsis and designated this as NtDRM1. The protein could be
shown to be localized exclusively in the nucleus and exhibit methylation
activity toward unmethylated synthetic as well as native DNA samples upon
expression in Sf9 insect cells. It also methylated hemimethylated DNA, but
the activity was lower than that for unmethylated substrates. Methylation
mapping of a 962-bp DNA, treated with NtDRM1 in vitro, directly
demonstrated methylation of approximately 70% of the cytosines in
methylatable CpNpN and CpNpG sequences but only 10% in CpG. Further
analyses indicated that the enzyme apparently non-selectively methylates
any cytosines except in CpG, regardless of the adjacent nucleotide at both
5' and 3' ends. Transcripts of NtDRM1 ubiquitously accumulated in all
tissues and during the cell cycle in tobacco cultured BY2 cells. These
results indicate that NtDRM1 is a de novo cytosine methyltransferase,
which actively excludes CpG substrate.

<>

<1>Wagenfuhr, K., Pieper, S., Mackeldanz, P., Linscheid, M., Kruger, D.H., Reuter, M.
<2>Structural domains in the type III restriction endonuclease EcoP15I: Characterization by limited proteolysis, mass spectrometry and insertional mutagenesis.
<3>J. Mol. Biol.
<4>366
<5>93-102
<6>2007
<7>The Type III restriction endonuclease EcoP15I forms a hetero-oligomeric enzyme complex that
consists of two modification (Mod) subunits and two
restriction (Res) subunits. Structural data on Type III restriction
enzymes in general are lacking because of their remarkable size of more
than 400 kDa and the laborious and low-yield protein purification
procedures. We took advantage of the EcoP15I-overexpressing vector pQEP15
and affinity chromatography to generate a quantity of EcoP15I high enough
for comprehensive proteolytic digestion studies and analyses of the
proteolytic fragments by mass spectrometry. We show here that in the
presence of specific DNA the entire Mod subunit is protected from trypsin
digestion, whereas in the absence of DNA stable protein domains of the Mod
subunit were not detected. In contrast, the Res subunit is comprised of
two trypsin-resistant domains of approximately 77-79 kDa and 27-29 kDa,
respectively. The cofactor ATP and the presence of DNA, either specific or
unspecific, are important stabilizers of the Res subunit. The large
N-terminal domain of Res contains numerous functional motifs that are
predicted to be involved in ATP-binding and hydrolysis and/or DNA
translocation. The C-terminal small domain harbours the catalytic center.
Based on our data, we conclude that both structural Res domains are
connected by a flexible linker region that spans 23 amino acid residues.
To confirm this conclusion, we have investigated several EcoP15I enzyme
mutants obtained by insertion mutagenesis in and around the predicted
linker region within the Res subunit. All mutants tolerated the genetic
manipulation and did not display loss of function or alteration of the DNA
cleavage position.

<>

<1>Wagner, E., Schmitz, G.G., Kaluza, K., Jarsch, M., Gotz, F., Kessler, C.
<2>BmyI, a novel SduI isoschizomer from Bacillus mycoides recognizing 5'-GDGCH/C-3'.
<3>Nucleic Acids Res.
<4>18
<5>3088
<6>1990
<7>None

<>

<1>Wah, D.A.
<2>Structural study of the type IIs restriction endonuclease FokI.
<3>Ph.D. Thesis, Columbia University
<4>
<5>1-108
<6>1998
<7>This work presents the first three-dimensional structures of a type IIs restriction
endonuclease.  The type IIs restriction endonucleases are an unusual class of enzymes that
recognize a specific DNA sequence and cleave DNA non-specifically a short distance away from
that sequence.  FokI, from Flavobacterium okeanokoites, is a type IIs restriction endonuclease
that binds the cognate sequence 5'-GGATG-3' and cleaves DNA phosphodiester groups 9 base
pairs away on this strand and 13 base pairs away on the complementary strand.  Because of its
unusual bipartite nature, FokI has been used to create artificial enzymes with new DNA-binding
specificities.  The three-dimensional structures of the complete FokI enzyme in complex with
DNA and without DNA have been determined by X-ray crystallography at 2.8A and 2.3A resolution,
respectively.  As anticipated, the enzyme contains amino- and carboxy-terminal domains
corresponding to the DNA-recognition and cleavage functions.  The recognition domain is made
of three smaller subdomains that are evolutionarily related to the helix-turn-helix containing
DNA-binding domain of the catabolite gene activator protein CAP.  However, the CAP core has
been extensively embellished in the first two subdomains while in the third subdomain it has
been co-opted for protein-protein interactions.  Surprisingly, the cleavage domain resembles a
monomer of the type II restriction endonuclease BamHI and contains only a single catalytic
center.  Unexpectedly, the cleavage domain is sequestered in a "piggyback" fashion by the
recognition domain until its DNA cleavage activity is required.  The structure of FokI in the
absence of DNA reveals a dimer.  In corroboration with binding data, a model is proposed in
which DNA cleavage requires the dimerization of two FokI molecules, each recognizing an
independent FokI cognate sequence.  Taken together, the structures have implications for the
evolution of helix-turn-helix domains, suggest a novel mechanism of nuclease activation, and
provide a framework for the design of chimeric enzymes with altered specificities.

<>

<1>Wah, D.A.
<2>Structural study of the type IIs restriction endonuclease FokI.
<3>Diss. Abstr.
<4>58
<5>6439B
<6>1998
<7>This work represents the first three-dimensional structures of a type IIs restriction
endonuclease.  The type IIs restriction endonucleases are an unusual class of enzymes that
recognize a specific DNA sequence and cleave DNA non-specifically a short distance away from
that sequence.  FokI, from Flavobacterium okeanokoites, is a type IIs restriction endonuclease
that binds the cognate sequence 5'-GGATG-3' and cleaves DNA phosphodiester groups 9 base
pairs away on this strand and 13 base pairs away on the complementary strand.  Because of its
unusual bipartite nature, FokI has been used to create artificial enzymes with new DNA-binding
specificities.  The three-dimensional structures of the complete FokI enzyme in complex with
DNA and without DNA have been determined by X-ray crystallography at 2.8 and 2.3 angstroms
resolution, respectively.  As anticipated, the enzyme contains amino- and carboxy-terminal
domains corresponding to the DNA-recognition and cleavage functions.  The recognition domain
is made of three smaller subdomains that are evolutionarily related to the helix-turn-helix
containing DNA-binding domain of the catabolite gene activator protein CAP.  However, the CAP
core has been extensively embellished in the first two subdomains while in the third subdomain
it has been co-opted for protein-protein interactions.  Surprisingly, the cleavage domain
resembles a monomer of the type II restriction endonuclease BamHI and contains only a single
catalytic center.  Unexpectedly, the cleavage domain is sequestered in a "piggyback" fashion
by the recognition domain until its DNA cleavage activity is required.  The structure of FokI
in the absence of DNA reveals a dimer.  In corroboration with binding data, a model is
proposed in which DNA cleavage requires the dimerization of two FokI molecules, each
recognizing an independent FokI cognate sequence.  Taken together, the structures have
implications for the evolution of helix-turn-helix domains, suggest a novel mechanism of
nuclease activation, and provide a framework for the design of chimeric enzymes with altered
specificities.

<>

<1>Wah, D.A., Bitinaite, J., Schildkraut, I., Aggarwal, A.K.
<2>Structure of FokI has implications for DNA cleavage.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>95
<5>10564-10569
<6>1998
<7>FokI is a member of an unusual class of restriction enzymes that recognize a specific DNA
sequence and cleave nonspecifically a short distance away from that sequence.  FokI consists
of an N-terminal DNA recognition domain and a C-terminal cleavage domain.  The bipartite
nature of FokI has led to the development of artificial enzymes with novel specificities.  We
have solved the structure of FokI to 2.3 A resolution.  The structure reveals a dimer, in
which the dimerization interface is mediated by the cleavage domain.  Each monomer has an
overall conformation similar to that found in the FokI-DNA complex, with the cleavage domain
packing alongside the DNA recognition domain.  In corroboration with the cleavage data
presented in the accompanying paper in this issue of Proceedings, we propose a model for FokI
DNA cleavage that requires the dimerization of FokI on DNA to cleave both DNA strands.

<>

<1>Wah, D.A., Hirsch, J.A., Dorner, L.F., Schildkraut, I., Aggarwal, A.K.
<2>Structure of the multimodular endonuclease FokI bound to DNA.
<3>Nature
<4>388
<5>97-100
<6>1997
<7>FokI is a member of an unusual class of bipartite restriction enzymes that recognize a
specific DNA sequence and cleave DNA nonspecifically a short distance away from that sequence.
Because of its unusual bipartite nature, FokI has been used to create artificial enzymes with
new specificities.  We have determined the crystal structure at 2.8A resolution of the
complete FokI enzyme bound to DNA.  As anticipated, the enzyme contains amino- and
carboxy-terminal domains corresponding to the DNA-recognition and cleavage functions,
respectively.  The recognition domain is made of three smaller subdomains (D1, D2 and D3)
which are evolutionarily related to the helix-turn-helix-containing DNA-binding domain of the
catabolite gene activator protein CAP.  The CAP core has been extensively embellished in the
first two subdomains, whereas in the third subdomain it has been co-opted for protein-protein
interactions.  Surprisingly, the cleavage domain contains only a single catalytic center,
raising the question of how monomeric FokI manages to cleave both DNA strands.  Unexpectedly,
the cleavage domain is sequestered in a 'piggyback' fashion by the recognition domain.  The
structure suggests a new mechanism for nuclease activation and provides a framework for the
design of chimeric enzymes with altered specificities.

<>

<1>Wahab, T., Ferrari, S., Lindberg, M., Backman, S., Kaden, R.
<2>Draft Genome Sequences of Brucella suis Biovar 4 Strain NCTC 10385, Brucella ceti Strain NCTC 12891T, Brucella inopinata Strain CAMP 6436T, and Brucella neotomae  Strain ATCC 23459T.
<3>Genome Announcements
<4>2
<5>e00783-14
<6>2014
<7>With the aim of developing quantitative PCR methods for the detection and differentiation of
Brucella species, the genomes of Brucella ceti, Brucella
inopinata, Brucella netotomae, and Brucella suis biovar 4 were sequenced and
analyzed.

<>

<1>Wahhab, A., Besterman, J.M., Delorme, D., Isakovic, L., Llewellyn, D., Rahil, J., Saavedra, O., Deziel, R.
<2>Inhibitors of DNA methyltransferase.
<3>International Patent Office
<4>WO 200678752 A
<5>
<6>2006
<7>The invention relates to the inhibition of DNA methyltransferase isoforms DNMT1 and DNMT3b2.
The invention provides compounds and methods for inhibiting DNMT1 and DNMT3b2.

<>

<1>Wahhab, A., Besterman, J.M., Delorme, D., Ljubomir, I., Llewellyn, D., Rahil, J., Saavedra, O., Deziel, R.
<2>Preparation of nucleoside analogs as inhibitors of DNA methyltransferase isoforms DNMT1 and DNMT3b2.
<3>US Patent Office
<4>US 20080132525 A
<5>
<6>2008
<7>The invention relates to the inhibition of DNA methyltransferase isoforms DNMT1 and DNMT3b2.
The invention provides compounds and methods for inhibiting DNMT1 and DNMT3b2.

<>

<1>Wahnon, D.C., Shier, V.K., Benkovic, S.J.
<2>Mechanism-based inhibition of an essential bacterial adenine DNA methyltransferase: Rationally designed antibiotics.
<3>J. Am. Chem. Soc.
<4>123
<5>976-977
<6>2001
<7>The emergence of bacteria resistant to available antibiotics has caused great concern in the
medical community and has created a need for the discovery of novel antibiotic agents.  In the
past, new antibiotics have been discovered by the random screening of natural products and the
subsequent identification of the target protein.  In addition to this drug discovery method,
the recent advances in genome sequencing have made it possible to envision a complementary
drug discovery method in which a bacterial enzyme target is first identified and new
antibiotic compounds are discovered from the targeted inhibition of this enzyme.  Successful
antibiotics would be inhibitors of an essential bacterial enzyme that is unique to bacteria
and has no mammalian homologue.  Here we report on the progress toward the development of
small-molecule selective inhibitors of an essential bacterial N-6 adenine DNA
methyltransferase, using a mechanism-based multisubstrate adduct approach and demonstrate the
inhibition using CcrM (cell cycle regulated DNA MTase) from the pathogenic Brucella abortus.

<>

<1>Waite-Rees, P.A., Keating, C.J., Moran, L.S., Slatko, B.E., Hornstra, L.J., Benner, J.S.
<2>Characterization and expression of the Escherichia coli Mrr restriction system.
<3>J. Bacteriol.
<4>173
<5>5207-5219
<6>1991
<7>The mrr gene of Escherichia coli K-12 is involved in the acceptance of foreign DNA which is
modified.  The introduction of plasmids carrying the HincII, HpaI, and TaqI R and M genes is
severely restricted in E. coli strains that are Mrr+. A 2-kb EcoRI fragment from the plasmid
pBg3 (B. Sain and N.E. Murray, Mol. Gen. Genet. 180:35-46, 1980) was cloned.  The resulting
plasmid restores Mrr function to mrr strains of E. coli.  The boundaries of the mrr gene were
determined from an analysis of subclones, and plasmids with a functional mrr gene produce a
polypeptide of 33.5 kDa.  The nucleotide sequence of the entire fragment was determined; in
addition to mrr, it includes two open reading frames, one of which encodes part of the hsdR.
By using Southern blot analysis, E. coli RR1 and HB101 were found to lack the region
containing mrr. The acceptance of various cloned methylases in E. coli containing the cloned
mrr gene was tested.  Plasmid constructs containing the AccI, CviRI, HincII, HinfI (HhaII),
HpaI, NlaIII, PstI, and TaqI N6-adenine methylases and SssI and HhaI C5-cytosine methylases
were found to be restricted.  Plasmid constructs containing 16 other adenine methylases and 12
cytosine methylases were not restricted.  No simple consensus sequence causing restriction has
been determined.  The Mrr protein has been overproduced, an antibody has been prepared, and
the expression of mrr under various conditions has been examined. The use of mrr strains of E.
coli is suggested for the cloning of N6-adenine and C5-cytosine methyl-containing DNA.

<>

<1>Wajima, T., Sabui, S., Kano, S., Ramamurthy, T., Chatterjee, N.S., Hamabata, T.
<2>Entire sequence of the colonization factor coli surface antigen 6-encoding plasmid pCss165 from an enterotoxigenic Escherichia coli clinical isolate.
<3>Plasmid
<4>70
<5>343-352
<6>2013
<7>Coli surface antigen 6 (CS6) is one of the most prevalent colonization factors among
enterotoxigenic Escherichia coli (ETEC) isolated in developing countries. Although it is known
that CS6 is encoded by a plasmid, there are no reports on the sequence analysis of the
CS6-encoding plasmid or genes exhibiting similar behavior to CS6. Here, we report the
isolation of the CS6-encoding plasmid, pCss165Kan, from 4266 DcssB::kanamycin (Km) and its
complete nucleotide sequence. This plasmid consisted of 165,311 bp and 222 predicted coding
sequences. Remarkably, there were many insertion sequence (IS) elements, which comprised 24.4%
of the entire sequence. Virulence-associated genes such as heat-stable enterotoxin, homologues
of ATP-binding cassette transporter in enteroaggregative E. coli (EAEC), and ETEC
autotransporter A were also present, although the ETEC autotransporter A gene was disrupted by
the integration of IS629. We found that 2 transcriptional regulators belonging to the AraC
family were not involved in CS6 expression. Interestingly, pCss165 had conjugative transfer
genes, as well as 3 toxin-antitoxin
systems that potentially exclude other plasmid-free host bacteria. These genes might be
involved in the prevalence of CS6 among ETEC isolates

<>

<1>Wakefield, R.I.D., Smith, B.O., Nan, X., Free, A., Soteriou, A., Uhrin, D., Bird, A.P., Barlow, P.N.
<2>The solution structure of the domain from MeCP2 that binds to methylated DNA.
<3>J. Mol. Biol.
<4>291
<5>1055-1065
<6>1999
<7>MeCP2 is an abundant mammalian protein that binds methylated CpG (mCpG) sequences within
double-stranded DNA, represses transcription by recruiting histone deacetylases, and is
essential for embryonic development. It is one of a family of proteins which mediate the
biological consequences of DNA methylation. These proteins each possess a sequence motif of
about 70 residues which, in MeCP2, form a domain necessary and sufficient for binding to mCpG.
The solution structure of the mCpG-binding domain (MBD) from MeCP2 has been solved and the
DNA-binding surface of the domain mapped using NMR spectroscopy. Residues 95-162 of MeCP2
adopt a novel fold forming a wedge-shaped structure. An N-terminal four-stranded antiparallel
beta-sheet forms one face of the wedge, while the other face is formed mainly by a C-terminal
helical region. The thin end of the wedge is extended by a long loop between beta-strands B
and C containing many basic residues. The B-C loop together with residues in strands B, C and
D, and at the N terminus of the alpha-helix, appears to form an interface with methylated DNA.
Unstructured residues at the NH2 terminus of the domain are also involved in formation of the
complex. The presence of numerous arginine and lysine side-chains on the DNA-binding surface
of MBD is consistent with the requirement for the mCpG site to be flanked by non-specific
sequences of base-pairs. The absence of symmetry in the domain implies that recognition does
not exploit the symmetry of the binding site. A conserved hydrophobic pocket containing the
side-chains of Tyr123 and Ile125 on the positively charged beta-sheet face is a candidate for
the region of contact with the methyl-groups of the modified cytosine residues.

<>

<1>Walder, R., Walder, J., Donelson, J.
<2>The DNA Sequence and Organization of the PstI Restriction-Modification Genes.
<3>Fed. Proc.
<4>40
<5>1647
<6>1981
<7>We have determined the sequence of a 4000 base pair DNA fragment from P. stuartii 164 that
contains the genes for the PstI restriction-modification system.  The cloning and expression
of these genes in E. coli and their re-introduction into P. stuartii 164 was reported earlier
(Fed. proc. 39 (6) 1413, (198); Proc. Natl. Acad. Sci., in press).  Within the sequence, two
large open reading frames were identified; one corresponding to the restriction enzyme, the
other to the modification methylase as determined by site directed mutagenesis and subcloning
experiments.  The genes are encoded on opposite DNA strands, unlike the two other
restriction-modification systems that have been studied, HhaII and EcoRI, in which the two
genes are colinear on the same strand (Fed. Proc. 39 (6) 946, (1980); Fed. Proc. 39 (6) 1415,
91980)).  This rules out the possibility, at least in this system, that the two genes are
expressed on a polycistronic transcript.  The locations of the putative promoters for the two
genes corresponds well with the sites of initiation of transcription mapped in vitro.  The
4000 base pair fragment is now being used to study the control of expression of these two
genes.

<>

<1>Walder, R.Y.
<2>The cloning and sequence organization of the PstI restriction-modification system.
<3>Ph.D. Thesis, The University of Iowa, Department of Biology, , none
<4>
<5>1-228
<6>1984
<7>The genes for the type II restriction and modification enzymes of the bacterium Providencia
stuartii 164, were cloned and expressed in the Escherichia coli strain HB101.  The two genes
are closely linked and are located within a 4.0 kilobase DNA region.  The complete nucleotide
sequence of the 4.0 kilobase fragment was determined.  Two large open reading frames were
identified within the sequence and were ascribed to the restriction endonuclease (PstI) and
the modification (methylase) enzyme by the analysis of a series of deletion and insertion
mutants.  The two genes are encoded on opposite DNA strands, and hence must be transcribed
from separate promoters rather than as a polycistronic message.  The sequence of the first ten
amino acids of the restriction endonuclease was determined by sequential Edman degradation of
the purified protein, permitting the alignment of the polypeptide with the DNA sequence.  The
amino terminus of the modification enzyme was established by sequential Edman degradation of
the protein synthesized in bacterial minicells with different radiolabeled amino acids.  The
initiation codons of the two genes are separated by 130 base pairs.  The deduced amino acid
sequences indicate that the restriction endonuclease contains 326 amino acids with a
calculated molecular weight of 37,370; the modification enzyme is composed of 507 amino acids
with a calculated molecular weight of 56,830.  There is no significant homology between the
two proteins at the level of the primary structure.  Antibody raised against the purified
restriction endonuclease did not immunoprecipitate the modification enzyme.  The transcription
initiation sites were mapped using mung bean nuclease.  Both of the transcripts begin with
adenosine.  The initiation sites are separated by only 70 base pairs.  This close proximity
requires that the promoter sites for the two divergent genes overlap.  Dnase I protection
experiments show a higher affinity of E. coli RNA polymerase for the methylase promoter than
for the restriction enzyme promoter.  The expression of the PstI restriction and modification
genes was also examined in bacteriophage lambda and in yeast.

<>

<1>Walder, R.Y., Hartley, J.L., Donelson, J.E., Walder, J.A.
<2>Cloning of the PstI restriction-modification system.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>217-226
<6>1981
<7>I. Introduction II. Selection and characterization of clones carrying the PstI
restriction-modification system III. Transformation of P. stuartii 164:
Construction of a stable overproducing strain IV. Gene products of the PstI
restriction-modification system V. In vitro transcription of the PstI system
VI. Discussion

<>

<1>Walder, R.Y., Hartley, J.L., Donelson, J.E., Walder, J.A.
<2>Cloning and expression of the PstI restriction-modification system in Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>78
<5>1503-1507
<6>1981
<7>Here we report the cloning and preliminary characterization of the PstI
restriction-modification system of Providencia stuartii 164.  Transformants of
Escherichia coli carrying the PstI gene system inserted into the cloning vector
pBR322 were selected on the basis of acquired resistance to bacteriophage
lambda infection.  PstI endonuclease was detected in osmotic shock fluid from
each of the resistant clones.  Plasmid and chromosomal DNA from these clones
could not be digested by PstI, indicating that the gene for the corresponding
modification enzyme had also been cloned and was being expressed.  The smallest
recombinant plasmid encoding both activities, pPst201, contains an insert of
approximately 4000 base pairs.  In vitro transcription studies indicate that
this DNA fragment also contains the endogenous promoter(s) of the system.  When
pPst201 was introduced into a minicell-producing strain of E. coli, two new
proteins, 32,000 and 35,000 daltons, were synthesized.  We have assigned these
to the PstI modification (methylase) and restriction enzymes, respectively.
The active form of the restriction enzyme is a dimer, as determined by gel
filtration.  Constructed transformants of P. stuartii 164 that carry the PstI
system inserted into pBR322 produce approximately 10 times more PstI
endonuclease activity than does the native strain.

<>

<1>Walder, R.Y., Langtimm, C.J., Catterjee, R., Walder, J.A.
<2>Cloning of the MspI modification enzyme.
<3>J. Biol. Chem.
<4>258
<5>1235-1241
<6>1983
<7>The gene for the MspI modification enzyme from Moraxella was cloned in
Escherichia coli using the plasmid vector pBR322.  Selection of transformants
carrying the gene was based on the resistance of the modified plasmid encoding
the enzyme to cleavage by MspI.  Both chromosomal and plasmid DNA were modified
in the selected clones.  None of the clones obtained produced the cognate
restriction enzyme which suggests that in this system the genes for the
restriction enzyme and methylase are not closely linked.  Crude cell extracts
prepared from the recombinant strains, but not the host (E. coli HB101),
contain an S-adenosylmethionine-dependent methyltransferase specific for the
MspI recognition site, CCGG.  Production of the enzyme is 3-4-fold greater in
the transformants than in the original Moraxella strain.  5-Methylcytosine was
identified as the product of the reaction chromatographically.  The outer
cytosine of the recognition sequence, *CCGG, was shown to be the site of
methylation by DNA-sequencing methods.  This modification blocks cleavage by
both MspI and its isoschizomer HpaII.  HpaII, but not MspI, is able to cleave
the unmethylated strand of a hemimethylated substrate.  The relevance of these
results to the use of MspI and HpaII to analyze patterns of methylation in
genomic DNA is discussed.

<>

<1>Walder, R.Y., Langtimm, C.J., Chatterjee, R., Walder, J.A.
<2>Cloning of the MspI modification enzyme:  the site of modification and its effects on cleavage by MspI and HpaII.
<3>Fed. Proc.
<4>41
<5>1199
<6>1982
<7>The gene for the MspI modification enzyme from Moraxella was cloned in E. coli
using the plasmid vector pBR322.  Selection of transformants carrying the gene
was based on the resistance of the modified plasmid encoding the enzyme to
cleavage by MspI.  Both chromosomal and plasmid DNA were modified in these
clones.  None of the clones obtained produced the cognate restriction enzyme
indicating either that the genes for the restriction enzyme and methylase are
not closely linked or that simply the restriction enzyme is not expressed in E.
coli.  Crude extracts prepared from these clones, but not the host (E. coli
HB101), contain a methyl transferase specific for the MspI recognition
sequence, CCGG, which uses SAM as the methyl donor.  The level of this activity
is approximately 3 fold greater in the transformants than in the original
Moraxella strain.  5-methylcytidine was identified as the product of the
reaction by thin layer chromatography.  DNA sequencing showed that only the
external cytidine residue within the recognition sequence is methylated.  This
modification blocks cleavage by both MspI and its isoschizomer HpaII.  A
hemimethylated substance constructed in vitro was cleaved, although at a much
reduced rate, on the unmethylated strand by HpaII but not MspI.  The relevance
of these results to the use of MspI and HpaII to analyze patterns of
methylation in genomic DNA will be discussed.

<>

<1>Walder, R.Y., Walder, J.A.
<2>The organization and control of expression of the PstI restriction modification system.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>5
<5>209-226
<6>1987
<7>In Volume I of this series, we first reported the cloning and expression of the
PstI restriction-modification system in Escherichia coli.  The PstI restriction
enzyme and methylasae genes were isolated from a genomic library of Providencia
stuartii 164 DNA in E. coli HB101 on the basis of acquired resistance of the
host to infection by bacteriophage lambda.  Subcloning experiments localized
the two genes to a 4.0 kilobase HindIII fragment, the complete sequence of
which has been determined.  In this report we review the organization and
studies of the expression of the PstI genes, and report the cloning of the PstI
restriction enzyme and methylase genes in yeast.

<>

<1>Walder, R.Y., Walder, J.A., Donelson, J.E.
<2>The Organization and Complete Nucleotide Sequence of the PstI Restriction-Modification System.
<3>J. Biol. Chem.
<4>259
<5>8015-8026
<6>1984
<7>We have determined the nucleotide sequence of a 4.0-kilobase DNA fragment
containing the genes of the PstI restriction-modification system.  Two large
open reading frames were identified within the sequence and were ascribed to
the restriction enzyme and methylase by the analysis of a series of deletion
mutants.  The two genes are encoded on opposite DNA strands, and hence must be
transcribed from separate promoters rather than as a polycistronic message.
The sequence of the first 10 amino acids of the restriction endonuclease was
determined by sequential Edman degradation of the purified protein, permitting
the alignment of the polypeoptide with the DNA sequence.  The NH2 terminus of
the modification enzme was established by sequential Edman degradation of the
protein synthesized in bacterial minicells with different radiolabeled amino
acids.  The initiation codons of the two genes are separated by 130 base pairs.
The deduced amino acid sequences indicate that the restriction endonuclease
contains 326 amino acids with a calculated Mr = 37,370; the modification enzyme
is composed of 507 amino acids with a calculated Mr = 56,830.  There is no
significant homology between the two proteins at the level of the primary
structure.  Antibody raised against the purified restriction endonuclease did
not immunoprecipitate the modification enzyme.  The transcription initiation
sites were mapped using mung bean nuclease.  Both of the transcripts begin with
adenosine.  The initiation sites are separated by only 70 base pairs.  This
close proximity suggests that the promoters for the two divergent genes
overlap.  DNase I protection experiments show that Escherichia coli RNA
polymerase has a higher affinity for the methylase promoter than for the
restriction enzyme promoter.

<>

<1>Waldron, D.E., Lindsay, J.A.
<2>Sau1: a novel lineage-specific type I restriction-modification system that blocks horizontal gene transfer into Staphylococcus aureus and between S.  aureus isolates of different lineages.
<3>J. Bacteriol.
<4>188
<5>5578-5585
<6>2006
<7>The Sau1 type I restriction-modification system is found on the chromosome of all nine
sequenced strains of Staphylococcus aureus and includes a single hsdR (restriction) gene and
two copies of hsdM (modification) and sdS (sequence specificity) genes. The strain S. aureus
RN4220 is a vital intermediate for laboratory S. aureus manipulation, as it can accept plasmid
DNA from Escherichia coli. We show that it carries a mutation in the sau1hsdR gene and that
complementation restored a nontransformable phenotype. Sau1 was also responsible for reduced
conjugative transfer from enterococci, a model of vancomycin resistance transfer. This may
explain why only four vancomycin-resistant S. aureus strains have been identified despite
substantial selective pressure in the clinical setting. Using a multistrain S. aureus
microarray, we show that the two copies of sequence specificity genes (sau1hsdS1 and
sau1hsdS2) vary substantially between isolates and that the variation corresponds to the 10
dominant S. aureus lineages. Thus, RN4220 complemented with sau1hsdR was resistant to
bacteriophage lysis but only if the phage was grown on S. aureus of a different lineage.
Similarly, it could be transduced with DNA from its own lineage but not with the phage grown
on different S. aureus lineages.  Therefore, we propose that Sau1 is the major mechanism for
blocking transfer of resistance genes and other mobile genetic elements into S. aureus
isolates from other species, as well as for controlling the spread of resistance genes between
isolates of different S. aureus lineages.  Blocking Sau1 should also allow genetic
manipulation of clinical strains of S. aureus.

<>

<1>Waldron, D.E., Owen, P., Dorman, C.J.
<2>Competitive interaction of the OxyR DNA-binding protein and the Dam methylase at the antigen 43 gene regulatory region in Escherichia coli.
<3>Mol. Microbiol.
<4>44
<5>509-520
<6>2002
<7>The antigen 43 surface protein of Escherichia coli is expressed in a phase-variable manner by
a mechanism involving alternative activation
and repression of transcription of the agn43 gene. The repressor is the
OxyR DNA-binding protein, and its binding site was found to be located
downstream of the agn43 transcription start site in a region of DNA
that encompasses three 5' -GATC-3' sequences that are subject to
Dam-mediated DNA methylation. It has been suggested previously that the
phase-variable expression of antigen 43 results from a competition
between Dam methylase and the OxyR repressor for these sites. The 5'
-GATC-3' sequences were inactivated for methylation by site-directed
mutagenesis, and all possible combinations of inactive and active sites
were assessed for effects on phase-variable expression of the agn43
gene. Inactivation of any 5' -GATC-3' site individually had no effect;
at least two sites had to be inactivated to disrupt the normal pattern
of expression. Studies of OxyR interaction with agn43 DNA showed that
methylation of any two 5' -GATC-3' sites was necessary and sufficient
to block binding of the repressor. It was also found that the adenines
of the second and third 5' -GATC-3' sites are required for OxyR
binding, demonstrating that the sites for Dam methylation and for
repressor binding are intimately associated. This is consistent with a
competition model in which Dam and OxyR share a preference for specific
DNA sequences in the regulatory region of the agn43 gene.

<>

<1>Waleron, K., Nakonieczna, J., Waleron, M., Podhajska, A.J.
<2>50 years of studies of restriction - modification systems (Systemy restrykcyjno-modyfikacyjne - 50 lat badan).
<3>Biotechnologia (Poznan)
<4>2
<5>65-88
<6>2006
<7>The basic function of restriction endonucleases and methyltransferases is protection of the
host genome against foreign DNA. Their
recombination and transposition related functions are still being
discussed. Some authors postulate that R-M genes may act as selfish
genetic elements. Restriction endonucleases are indispensable tools in
molecular biology. As these enzymes recognize DNA sequence very
specifically, they serve as a model for protein-DNA interaction.
Restriction-modification genes have also played the same role as a
model for evolutionary studies as well as protein structure - function
relations. So far, there have been more than 3500 bacterial strains
studied which possessed R-M genes of more than 280 different
specificities towards recognition sequence and cleavage sites. They
became a very good commercial product for many biotechnological
companies. At present, in a genome sequencing era, R-M genes seem to be
much more common than it was thought before.

<>

<1>Waleron, K., Waleron, M., Osipiuk, J., Podhajska, A.J., Lojkowska, E.
<2>Identification of a DNA restriction-modification system in Pectobacterium carotovorum strains isolated from Poland.
<3>J. Appl. Microbiol.
<4>100
<5>343-351
<6>2006
<7>Aims: Polish isolates of pectinolytic bacteria from the species Pectobacterium carotovorum
were screened for the presence of a DNA
restriction-modification (R-M) system.
Methods and Results: Eighty-nine strains of P. carotovorum were
isolated from infected potato plants. Sixty-six strains belonged to P.
carotovorum ssp. atrosepticum and 23 to P. carotovorum ssp.
carotovorum. The presence of restriction enzyme Pca17AI, which is an
isoschizomer of EcoRII endonuclease, was observed in all isolates of P.
c. atrosepticum but not in P. c. carotovorum. The biochemical
properties, PCR amplification, and sequences of the Pca17AI restriction
endonuclease and methyltransferase genes were compared with the
prototype EcoRII R-M system genes. Only when DNA isolated from cells of
P. c. atrosepticum was used as a template, amplification of a 680 bp
homologous to the gene coding EcoRII endonuclease.
Conclusions: Endonuclease Pca17AI, having a relatively low
temperature optimum, was identified. PCR amplification revealed that
the nucleotide sequence of genes for EcoRII and Pca17AI R-M are
different. Dcm methylation was observed in all strains of
Pectobacterium and other Erwinia species tested. The sequence of a DNA
fragment coding Dcm methylase in P. carotovorum was different from that
of Escherichia coli.
Significance and Impact of the Study: Pca17AI is the first
psychrophilic isoschizomer of EcoRII endonuclease. The presence of
specific Dcm methylation in chromosomal DNA isolated from P.
carotovorum is described for the first time. A 680 bp PCR product,
unique for P. c. atrosepticum strains, could serve as a molecular
marker for detection of these bacteria in environmental samples.

<>

<1>Walker, C.A., Mannering, S.A., Shields, S., Blake, D.P., Brownlie, J.
<2>Complete Genome Sequence of Mycoplasma cynos Strain C142.
<3>Genome Announcements
<4>1
<5>e00196-12
<6>2013
<7>Here we report the de novo genome sequencing of Mycoplasma cynos strain C142, isolated from a
dog with canine infectious respiratory disease (CIRD) in the
United States.

<>

<1>Walker, C.B. et al.
<2>Nitrosopumilus maritimus genome reveals unique mechanisms for nitrification and autotrophy in globally distributed marine crenarchaea.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>8818-8823
<6>2010
<7>Ammonia-oxidizing archaea are ubiquitous in marine and terrestrial environments and now
thought to be significant contributors to carbon and
nitrogen cycling. The isolation of Candidatus 'Nitrosopumilus maritimus'
strain SCM1 provided the opportunity for linking its chemolithotrophic
physiology with a genomic inventory of the globally distributed archaea.
Here we report the 1,645,259-bp closed genome of strain SCM1, revealing
highly copper-dependent systems for ammonia oxidation and electron
transport that are distinctly different from known ammonia-oxidizing
bacteria. Consistent with in situ isotopic studies of marine archaea, the
genome sequence indicates N. maritimus grows autotrophically using a
variant of the 3-hydroxypropionate/4-hydroxybutryrate pathway for carbon
assimilation, while maintaining limited capacity for assimilation of
organic carbon. This unique instance of archaeal biosynthesis of the
osmoprotectant ectoine and an unprecedented enrichment of multicopper
oxidases, thioredoxin-like proteins, and transcriptional regulators points
to an organism responsive to environmental cues and adapted to handling
reactive copper and nitrogen species that likely derive from its
distinctive biochemistry. The conservation of N. maritimus gene content
and organization within marine metagenomes indicates that the unique
physiology of these specialized oligophiles may play a significant role in
the biogeochemical cycles of carbon and nitrogen.

<>

<1>Walker, G.T., Little, M.C., Nadeau, J.G., Shank, D.D.
<2>Isothermal in vitro amplification of DNA by a restriction enzyme/DNA polymerase system.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>89
<5>392-396
<6>1992
<7>An isothermal in vitro DNA amplification method was developed based upon the
following sequence of reaction events.  Restriction enzyme cleavage and
subsequent heat denaturation of a DNA sample generates two single-stranded
target DNA fragments (T1 and T2). Present in excess are two DNA amplification
primers (P1 and P2).  The 3' end of P1 binds to the 3' end of T1, forming a
duplex with 5' overhangs.  Likewise, P2 binds to T2.  The 5' overhangs of P1
and P2 contain a recognition sequence (5'-GTTGAC-3') for the restriction enzyme
HincII.  An exonuclease-deficient form of the large fragment of Escherichia
coli DNA polymerase I (exo- Klenow polymerase) [Derbyshire, V., Freemont, P.S.,
Sanderson, M.R., Beese, L., Friedman, J.M., Joyce, C.M. & Steitz, T.A. (1988)]
Science 240, 199-201] extends the 3' ends of the duplexes using dGTP, dCTP,
TTP, and deoxyadenosine 5'-[alpha-thio] triphosphate, which produces
hemiphosphorothioate recognition sites on P1T1 and P2T2.  HincII nicks the
unprotected primer strands of the hemiphosphorothioate recognition sites,
leaving intact the modified complementary strands.  The exo- Klenow polymerase
extends the 3' end at the nick on P1T1 and displaces the downstream strand that
is functionally equivalent to T2.  Likewise, extension at the nick on P2T2
results in displacement of a downstream strand functionally equivalent to T1.
Nicking and polymerization/displacement steps cycle continuously on P1T1 and
P2T2 because extension at a nick regenerates a nickable HincII recognition
site.  Target amplification is exponential because strands displaced from P1T1
serve as targets for P2 and strands displaced from P2T2 serve as targets for
P1.  A 10/6-fold amplification of a genomic sequence from Mycobacterium
tuberculosis or Mycobacterium bovis was achieved in 4 h at 37C.

<>

<1>Walker, J.N.B., Dean, P.D.G., Saunders, J.R.
<2>Identification and characterisation of PmaCI an endonuclease of novel specificity from Pseudomonas maltophila.
<3>Nucleic Acids Res.
<4>14
<5>1293-1301
<6>1986
<7>We report the use of MonoQ FPLC (Fast Protein Liquid Chromatography) for the
rapid purification of a novel Type II restriction endonuclease PmaCI, from
Pseudomonas maltophila, which recognises the sequence 5'-CAC^GTG-3'.  The
resulting enzyme is free of other nucleases to a level suitable for its
characterisation by multiple-substrate digestion and DNA sequencing techniques.
This method appears to be widely applicable and we have used it for the
isolation of restriction endonucleases of comparable purity from a range of
other organisms.  Also described is a rapid method for screening a library of
small inserted regions in recombinant M13 molecules for the presence and
subsequent screening of restriction sites of interest.

<>

<1>Walker, R., Watkin, E., Tian, R., Brau, L., O'Hara, G., Goodwin, L., Han, J., Lobos, E., Huntemann, M., Pati, A., Woyke, T., Mavromatis, K., Markowitz, V., Ivanova, N., Kyrpides, N., Reeve, W.
<2>Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2230 from Karijini National Park, Australia.
<3>Standards in Genomic Sciences
<4>9
<5>551-561
<6>2014
<7>Burkholderia sp. strain WSM2230 is an aerobic, motile, Gram-negative, non-spore-forming
acid-tolerant rod isolated from acidic soil collected in 2001
from Karijini National Park, Western Australia, using Kennedia coccinea (Coral
Vine) as a host. WSM2230 was initially effective in nitrogen-fixation with K.
coccinea, but subsequently lost symbiotic competence. Here we describe the
features of Burkholderia sp. strain WSM2230, together with genome sequence
information and its annotation. The 6,309,801 bp high-quality-draft genome is
arranged into 33 scaffolds of 33 contigs containing 5,590 protein-coding genes
and 63 RNA-only encoding genes. The genome sequence of WSM2230 failed to identify
nodulation genes and provides an explanation for the observed failure of the
laboratory grown strain to nodulate. The genome of this strain is one of 100
sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for
Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

<>

<1>Walker, R., Watkin, E., Tian, R., Brau, L., O'Hara, G., Goodwin, L., Han, J., Reddy, T., Huntemann, M., Pati, A., Woyke, T., Mavromatis, K., Markowitz, V., Ivanova, N., Kyrpides, N., Reeve, W.
<2>Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2232 from Karijini National Park, Australia.
<3>Standards in Genomic Sciences
<4>9
<5>1168-1180
<6>2014
<7>Burkholderia sp. strain WSM2232 is an aerobic, motile, Gram-negative, non-spore-forming
acid-tolerant rod that was trapped in 2001 from acidic soil
collected from Karijini National Park (Australia) using Gastrolobium capitatum as
a host. WSM2232 was effective in nitrogen fixation with G. capitatum but
subsequently lost symbiotic competence during long-term storage. Here we describe
the features of Burkholderia sp. strain WSM2232, together with genome sequence
information and its annotation. The 7,208,311 bp standard-draft genome is
arranged into 72 scaffolds of 72 contigs containing 6,322 protein-coding genes
and 61 RNA-only encoding genes. The loss of symbiotic capability can now be
attributed to the loss of nodulation and nitrogen fixation genes from the genome.
This rhizobial genome is one of 100 sequenced as part of the DOE Joint Genome
Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria
(GEBA-RNB) project.

<>

<1>Walker, S.A., Klaenhammer, T.R.
<2>The Genetics of Phage Resistance in Lactococcus lactis.
<3>Genetics of Phage Resistance, Kluwer Academic/Plenum Publishers, Wood, B.J.B. END  cut site unknown                           *>, 
<4>3
<5>291-315
<6>2003
<7>
<>

<1>Walkinshaw, M.D., Taylor, P., Sturrock, S.S., Atanasiu, C., Berge, T., Henderson, R.M., Edwardson, J.M., Dryden, D.T.
<2>Structure of Ocr from bacteriophage T7, a protein that mimics B-form DNA.
<3>Mol. Cell
<4>9
<5>187-194
<6>2002
<7>We have solved, by X-ray crystallography to a resolution of 1.8 A, the structure of a protein
capable of mimicking approximately 20 base pairs of
B-form DNA. This ocr protein, encoded by gene 0.3 of bacteriophage T7,
mimics the size and shape of a bent DNA molecule and the arrangement of
negative charges along the phosphate backbone of B-form DNA. We also
demonstrate that ocr is an efficient inhibitor in vivo of all known
families of the complex type I DNA restriction enzymes. Using atomic force
microscopy, we have also observed that type I enzymes induce a bend in DNA
of similar magnitude to the bend in the ocr molecule. This first structure
of an antirestriction protein demonstrates the construction of structural
mimetics of long segments of B-form DNA.

<>

<1>Wall, L.G. et al.
<2>Draft Genome Sequence of Frankia sp. Strain BCU110501, a Nitrogen-Fixing Actinobacterium Isolated from Nodules of Discaria trinevis.
<3>Genome Announcements
<4>1
<5>e00503-13
<6>2013
<7>Frankia forms a nitrogen-fixing symbiosis with actinorhizal plants. We report a draft genome
sequence for Frankia sp. strain BCU110501, a nitrogen-fixing
actinobacterium isolated from nodules of Discaria trinevis grown in the Patagonia
region of Argentina.

<>

<1>Wall, T., Bath, K., Britton, R.A., Jonsson, H., Versalovic, J., Roos, S.
<2>The Early Response to Acid Shock in Lactobacillus reuteri Involves the ClpL Chaperone and a Putative Cell Wall-Altering Esterase.
<3>Appl. Environ. Microbiol.
<4>73
<5>3924-3935
<6>2007
<7>To be able to function as a probiotic, bacteria have to survive the
passage through the gastrointestinal tract. We have examined survival and
gene expression of Lactobacillus reuteri ATCC 55730 after a sudden shift
in environmental acidity to a pH close to the conditions in the human
stomach. More than 80% of the L. reuteri cells survived at pH 2.7 for 1 h.
A genomewide expression analysis experiment using microarrays displayed 72
differentially expressed genes at this pH. The early response to severe
acid shock in L. reuteri differed from long-term acid adaptation to milder
acid stress studied in other lactic acid bacteria. The genes induced
included the following: clpL, genes putatively involved in alterations of
the cell membrane and the cell wall; genes encoding transcriptional
regulators; phage genes; and genes of unknown function. Two genes, clpL,
encoding an ATPase with chaperone activity, and lr1516, encoding a
putative esterase, were selected for mutation analyses. The mutants were
significantly more sensitive to acid than the wild type was. Thus, these
genes could contribute to the survival of L. reuteri in the
gastrointestinal tract.

<>

<1>Wallace, C.J.A.
<2>The curious case of protein splicing: mechanistic insights suggested by protein semisynthesis.
<3>Protein Sci.
<4>2
<5>697-705
<6>1993
<7>The gradual accumulation of examples of protein splicing, in which a
nested intervening sequence is spliced out of the interior of a polyprotein precursor,
suggests that this curious phenomenon might prove to have universal phylogenetic
distribution and biological significance.  The known examples are reviewed, with the aim
of establishing underlying patterns, and a generalized mechanism of autocatalytic protein
splicing is proposed.  The testable consequences of such a proposal and the possible
evolutionary origins of the phenomenon are discussed.

<>

<1>Wallace, J.G., Breaker, R.R.
<2>Improved genetic transformation methods for the model alkaliphile Bacillus halodurans C-125.
<3>Lett. Appl. Microbiol.
<4>52
<5>430-432
<6>2011
<7>Aims: Bacillus halodurans C-125 is a Gram-positive bacterium that was the
first alkaliphilic species to have its genome completely sequenced.
Despite its many years as a model for alkaliphily and source of
industrially important enzymes, genetic manipulation of B. halodurans
C-125 remains difficult, and therefore, we sought to develop a robust
method to allow routine transformation of this organism.
Methods and Results:
A plasmid artificial modification system (PAM system, <link
rid='b7'>Yasui et al. 2008) for B. halodurans C-125 was created that
increases transformation efficiency by 10- to 1000-fold. Also,
recovering transformed protoplasts on succinate nutrient agar (SNA)
yields faster, more robust colony recovery than on the traditional
recovery medium. Combining these two techniques often allows recovery
of transformants in as little as 48 h.
Conclusions:
Use of the B. halodurans C-125 PAM system and SNA greatly improves
the efficiency and speed of protoplast transformation of B. halodurans
C-125.
Significance and Impact of the Study:
These techniques allow routine genetic manipulation of B.
halodurans C-125, a model alkaliphilic bacterium with important
industrial properties.

<>

<1>Walsh, C.P., Bestor, T.H.
<2>Cytosine methylation and mammalian development.
<3>Genes Dev.
<4>13
<5>26-34
<6>1999
<7>Programmed methylation and demethylation of regulatory sequences has been proposed to play a
central role in vertebrate development.  We report here that the methylation status of the 5'
regions of a panel of tissue-specific genes could not be correlated with expression in tissues
of fetal and newborn mice.  Genes reported to be regulated by reversible methylation were not
expressed ectopically or precociously in Dnmt1-deficient mouse embryos under conditions where
demethylation caused biallelic expression of imprinted genes and activated transcription of
endogenous retroviruses of the IAP class.  These and other data suggest that the numerous
published expression-methylation correlations may have described not a cause but a consequence
of transcriptional activation.  A model is proposed under which cytosine methylation
represents a biochemical specialization of large genomes that participates in specialized
biological functions such as allele-specific gene expression and the heritable transcriptional
silencing of parasitic sequence elements, whereas cellular differentiation is controlled by
conserved regulatory networks that do not depend on covalent modification of the genome.

<>

<1>Walsh, C.P., Xu, G.L.
<2>Cytosine methylation and DNA repair.
<3>Curr. Top. Microbiol. Immunol., Springer-Verlag, Doerfler, W., Bohm, P., Berlin, Germany
<4>301
<5>283-315
<6>2006
<7>Cytosine methylation is a common form of post-replicative DNA modification seen in both
bacteria and eukaryotes. Modified cytosines have long been known to act as hotspots for
mutations due to the high rate of spontaneous deamination of this base to thymine, resulting
in a G/T mismatch. This will be fixed as a C -> T transition after replication if not repaired
by the base excision repair (BER) pathway or specific repair enzymes dedicated to this
purpose. This hypermutability has led to depletion of the target dinucleotide CpG outside of
special CpG islands in mammals, which are normally unmethylated. We review the importance of C
-> T transitions at non-island CpGs in human disease: When these occur in the germline, they
are a common cause of inherited diseases such as epidermolysis bullosa and
mucopolysaccharidosis, while in the soma they are frequently found in the genes for tumor
suppressors such as p53 and the retinoblastoma protein, causing cancer. We also examine the
specific repair enzymes involved, namely the endonuclease Vsr in Escherichia coli and two
members of the uracil DNA glycosylase (UDG) superfamily in mammals, TDG and MBD4. Repair
brings its own problems, since it will require remethylation of the replacement cytosine,
presumably coupling repair to methylation by either the maintenance methylase Dnmt1 or a de
novo enzyme such as Dnmt3a. Uncoupling of methylation from repair may be one way to remove
methylation from DNA. We also look at the possible role of specific cytosine deaminases such
as Aid and Apobec in accelerating deamination of methylcytosine and consequent DNA
demethylation.

<>

<1>Walsh, D.A., Zaikova, E., Howes, C.G., Song, Y.C., Wright, J.J., Tringe, S.G., Tortell, P.D., Hallam, S.J.
<2>Metagenome of a versatile chemolithoautotroph from expanding oceanic dead zones.
<3>Science
<4>326
<5>578-582
<6>2009
<7>Oxygen minimum zones, also known as oceanic "dead zones," are widespread
oceanographic features currently expanding because of global warming. Although
inhospitable to metazoan life, they support a cryptic microbiota whose metabolic
activities affect nutrient and trace gas cycling within the global ocean. Here,
we report metagenomic analyses of a ubiquitous and abundant but uncultivated
oxygen minimum zone microbe (SUP05) related to chemoautotrophic gill symbionts of
deep-sea clams and mussels. The SUP05 metagenome harbors a versatile repertoire
of genes mediating autotrophic carbon assimilation, sulfur oxidation, and nitrate
respiration responsive to a wide range of water-column redox states. Our analysis
provides a genomic foundation for understanding the ecological and biogeochemical
role of pelagic SUP05 in oxygen-deficient oceanic waters and its potential
sensitivity to environmental changes.

<>

<1>Walsh, D.A., Zaikova, E., Howes, C.L., Song, Y.C., Wright, J., Tringe, S.G., Tortell, P.D., Hallam, S.J.
<2>Metagenome of a versatile chemolithoautotroph from expanding oxygen minimum zones.
<3>Science
<4>326
<5>578-582
<6>2009
<7>Oxygen minimum zones, also known as oceanic "dead zones", are widesprad oceanographic features
currently expanding because of global warming.  Although inhospitable to metazoan life, they
support a cryptic microbiota whose metabolic activities affect nutrient and trace gas cycling
within the global ocean.  Here, we report metagenomic analyses of a ubiquitous and abundant
but uncultivated oxygen minimum zone microbe (SUPO5) related to chemoautotrophic gill
symbionts of deep-sea clams and mussels.  The SUP05 metagenome harbors a versatile repertoire
of genes mediating autotrophic carbon assimilation, sulfur oxidation, and nitrate respiration
responsive to a wide range of water-column redox states. Our analysis provides a genomic
foundation for understanding the ecological and biogeochemical role of pelagic SUP05 in
oxygen-deficient oceanic waters and its potential sensitivity to environmental changes.

<>

<1>Walter, F., Hartmann, M., Roth, M.
<2>Purification and characterization of a site specific endonuclease from Streptomyces hygroscopicus.
<3>Abstracts of 12th FEBS Symposium, Dresden.
<4>0
<5>648
<6>1978
<7>The isolation procedure of a new site specific endonuclease from Streptomyces
hygroscopicus J 0477 is described.  Using the Lysozyme treatment, we have
partially purified this enzyme (ShyI) by chromatography on Bio-Gel A0.5m, DEAE
Cellulose and Sepharose 4B coupled with 1,5-Diaminopentan ShyI require Mg2+ as
cofactor.  Salt concentrations higher than 0.2M NaCl inhibit the enzyme.  The
enzyme cleaves wild type lambda-DNA at two sites.

<>

<1>Walter, J., Noyer-Weidner, M., Trautner, T.A.
<2>The amino acid sequence of the CCGG recognizing DNA methyltransferase M.BsuFI: implications for the analysis of sequence recognition by cytosine DNA methyltransferases.
<3>EMBO J.
<4>9
<5>1007-1013
<6>1990
<7>The Bacillus subtilis FI DNA methyltransferase (M.BsuFI) modifies the outer
cytosine of the DNA sequence CCGG, causing resistance against R.BsuFI and
R.MspI restriction.  The M.BsuFI gene was cloned and expressed in B.subtilis
and Escherichia coli.  As derived from the nucleotide sequence, the M.BsuFI
protein has 409 amino acids, corresponding to a molecular mass of 46918
daltons.  Including these data we have compared the nucleotide and amino acid
sequences of different CCGG recognizing enzymes.  These analyses showed that
M.BsuFI is highly related to two other CCGG specific methyltransferases, M.MspI
and M.HpaII, which were isolated from Gram-negative bacteria.  Between M.BsuFI
and M.MspI the sequence similarity is particularly significant in a region,
which has been postulated to contain the target recognition domains (TRDs) of
cytosine-specific DNA methyltransferases.  Apparently M.BsuFI and M.MspI,
derived from phylogenetic distant organisms, use highly conserved structural
elements for the recognition of the CCGG target sequence.  In contrast the very
same region of M.HpaII is quite different from those of M.BsuFI and M.MspI.  We
attribute this difference to the different targeting of methylation within the
sequence CCGG, where M.HpaII methylates the inner, M.BsuFI/M.MspI the outer
cytosine.  Also the CCGG recognizing TRD of the multispecific B. subtilis phage
SPR Mtase is distinct from that of the host enzyme, possibly indicating
different requirements for TRDs operative in mono- and multispecific enzymes.

<>

<1>Walter, J., Trautner, T.A., Noyer-Weidner, M.
<2>High plasticity of multispecific DNA methyltransferases in the region carrying DNA target recognizing enzyme modules.
<3>EMBO J.
<4>11
<5>4445-4450
<6>1992
<7>Multispecific cytosine C5 DNA methyltransferases (MTases) methylate more than one specific DNA
target. This is due to the presence of several target recognizing domains (TRDs) in these
enzymes. Such TRDs form part of a variable center in the MTase primary sequence, which
separates conserved enzyme core sequences responsible for general steps in the methylation
reaction. By deleting, rearranging and exchanging several TRDs of multispecific MTases, we
demonstrate their modular character; they mediate target recognition independent of a
particular TRD or core sequence context. We show also that multipsecific MTases can
accommodate inert material of non-MTase origin within their variable region without losing
their activity. The remarkable plasticity with respect to the material that can be integrated
into this region suggests that the enzyme core sequences preceding or following it form
separable functional domains. In spite of the documented flexibitity multispecific MTass could
not be endowed with novel specificities by integration of putative TRDs of monospecific
MTases, pointing to differences between multi-and monospecific MTases in the way their core
and TRD sequences interact.

<>

<1>Wan, X., Qian, L., Hou, S., Drees, K.P., Foster, J.T., Douglas, J.T.
<2>Complete Genome Sequences of Beijing and Manila Family Strains of Mycobacterium tuberculosis.
<3>Genome Announcements
<4>2
<5>e01135-14
<6>2014
<7>The majority of isolates from tuberculosis patients in Hawaii arrive through the  immigration
of infected individuals from the western Pacific. We report here on
the annotated complete genomes of two strains of Mycobacterium tuberculosis from
the two main lineages/families in Hawaii, Beijing and Manila.

<>

<1>Wanchanthuek, P., Bellgard, M.I., La, T., Ryan, K., Moolhuijzen, P., Chapman, B., Black, M., Schibeci, D., Hunter, A., Barrero, R., Phillips, N.D., Hampson, D.J.
<2>The Complete Genome Sequence of the Pathogenic Intestinal Spirochete Brachyspira pilosicoli and Comparison with Other Brachyspira Genomes.
<3>PLoS ONE
<4>5
<5>E11455
<6>2010
<7>BACKGROUND: The anaerobic spirochete Brachyspira pilosicoli colonizes the
large intestine of various species of birds and mammals, including humans.
It causes "intestinal spirochetosis", a condition characterized by mild
colitis, diarrhea and reduced growth. This study aimed to sequence and
analyse the bacterial genome to investigate the genetic basis of its
specialized ecology and virulence. METHODOLOGY/PRINCIPAL FINDINGS: The
genome of B. pilosicoli 95/1000 was sequenced, assembled and compared with
that of the pathogenic Brachyspira hyodysenteriae and a near-complete
sequence of Brachyspira murdochii. The B. pilosicoli genome was circular,
composed of 2,586,443 bp with a 27.9 mol% G+C content, and encoded 2,338
genes. The three Brachyspira species shared 1,087 genes and showed
evidence of extensive genome rearrangements. Despite minor differences in
predicted protein functional groups, the species had many similar features
including core metabolic pathways. Genes distinguishing B. pilosicoli from
B. hyodysenteriae included those for a previously undescribed
bacteriophage that may be useful for genetic manipulation, for a glycine
reductase complex allowing use of glycine whilst protecting from oxidative
stress, and for aconitase and related enzymes in the incomplete TCA cycle,
allowing glutamate synthesis and function of the cycle during oxidative
stress. B. pilosicoli had substantially fewer methyl-accepting chemotaxis
genes than B. hyodysenteriae and hence these species are likely to have
different chemotactic responses that may help to explain their different
host range and colonization sites. B. pilosicoli lacked the gene for a new
putative hemolysin identified in B. hyodysenteriae WA1. Both B. pilosicoli
and B. murdochii lacked the rfbBADC gene cluster found on the B.
hyodysenteriae plasmid, and hence were predicted to have different
lipooligosaccharide structures. Overall, B. pilosicoli 95/1000 had a
variety of genes potentially contributing to virulence.
CONCLUSIONS/SIGNIFICANCE: The availability of the complete genome sequence
of B. pilosicoli 95/1000 will facilitate functional genomics studies aimed
at elucidating host-pathogen interactions and virulence.

<>

<1>Wang, A., Pattemore, J., Ash, G., Williams, A., Hane, J.
<2>Draft Genome Sequence of Bacillus thuringiensis Strain DAR 81934, Which Exhibits  Molluscicidal Activity.
<3>Genome Announcements
<4>1
<5>e0017512
<6>2013
<7>Bacillus thuringiensis has been widely used as a biopesticide for a long time. Its
molluscicidal activity, however, is rarely realized. Here, we report the
genome sequence of B. thuringiensis strain DAR 81934, a strain with molluscicidal
activity against the pest snail Cernuella virgata.

<>

<1>Wang, B., Jian, J., Lu, Y., Cai, S., Huang, Y., Tang, J., Wu, Z.
<2>Complete Genome Sequence of Streptococcus agalactiae ZQ0910, a Pathogen Causing Meningoencephalitis in the GIFT Strain of Nile Tilapia (Oreochromis niloticus).
<3>J. Bacteriol.
<4>194
<5>5132-5133
<6>2012
<7>Streptococcus agalactiae (group B streptococcus [GBS]) is a pathogen that causes
meningoencephalitis in Nile tilapia (Oreochromis niloticus). Here, we reported
the complete genome sequence of S. agalactiae strain ZQ0910, which was isolated
from the GIFT strain of Nile tilapia in Guangdong, China.

<>

<1>Wang, C., Chen, Q., Wang, R., Shi, C., Yan, X., He, J., Hong, Q., Li, S.
<2>A Novel Angular Dioxygenase Gene Cluster, Encoding 3-Phenoxybenzoate 1', 2' -Dioxygenase in Sphingobium wenxiniae JZ-1.
<3>Appl. Environ. Microbiol.
<4>80
<5>3811-3818
<6>2014
<7>Sphingobium wenxiniae JZ-1 utilizes a wide range of pyrethroids and their
metabolic product 3-phenoxybenzoate as source of carbon and energy. A mutant
MJZ-1 defective in the degradation of 3-phenoxybenzoate was obtained by
successive streaking on LB agar. Comparison of the draft genomes of strain JZ-1
and MJZ-1 revealed that a 29,371 bp DNA fragment containing a putative angular
dioxygenase gene cluster pbaA1A2B is missing in strain MJZ-1. PbaA1, PbaA2 and
PbaB share 65%, 52% and 10% identities with the corresponding alpha, beta
subunits and the ferredoxin component of dioxin dioxygenase from Sphingomonas
wittichii RW1, respectively. Complementation of pbaA1A2B in strain MJZ-1 resulted
in the active 3-phenoxybenzoate 1' , 2' -dioxygenase, but the enzyme activity in
Escherichia coli cells was achieved only through the co-expression of pbaA1A2B
and a GR (glutathione reductase)-type reductase gene pbaC, indicating that the
3-phenoxybenzoate 1' , 2' -dioxygenase belongs to Type IV Rieske non-heme iron
aromatic ring-hydroxylating oxygenase system consisting of an hetero-oligomeric
oxygenase, a [2Fe-2S]-type ferredoxin and a GR-type reductase. pbaC gene is not
located in the immediate vicinity of pbaA1A2B. 3-Phenoxybenzoate 1' , 2'
-dioxygenase catalyzes the hydroxylation in the 1' , 2' -positions of the phenol
moiety of 3-phenoxybenzoate, yielding 3-hydroxybenzoate and catechol. The
transcription of pbaA1A2B and pbaC were both induced by 3-phenoxybenzoate, but
the transcription level of pbaC was far low than that of pbaA1A2B, implying the
possibility that PbaC may be not the only reductase that can physiologically
transfer electrons to PbaA1A2B in strain JZ-1. Some GR-type reductases from other
sphingomonad strains could also transfer electrons to PbaA1A2B, suggesting that
PbaA1A2B has low specificity for reductase.

<>

<1>Wang, D., Han, C.S., Dichosa, A.E., Gleasner, C.D., Johnson, S.L., Daligault, H.E., Davenport, K.W., Li, P.E., Pierson, E.A., Pierson, L.S.I.I.I.
<2>Draft Genome Sequence of Enterobacter cloacae Strain S611.
<3>Genome Announcements
<4>2
<5>e00710-14
<6>2014
<7>We report draft genomes of Enterobacter cloacae strain S611, an endophytic bacterium isolated
from surface-sterilized germinating wheat seeds. We present
the assembly and annotation of its genome, which may provide insights into the
metabolic pathways involved in adaptation.

<>

<1>Wang, D., Han, C.S., Dichosa, A.E., Gleasner, C.D., Johnson, S.L., Daligault, H.E., Davenport, K.W., Li, P.E., Pierson, E.A., Pierson, L.S.I.I.I.
<2>Draft Genome Sequence of Pseudomonas putida Strain S610, a Seed-Borne Bacterium of Wheat.
<3>Genome Announcements
<4>1
<5>e01048-13
<6>2013
<7>We report the genome sequence of a seed-borne bacterium, Pseudomonas putida strain S610. The
size of the draft genome sequence is approximately 4.6 Mb, which
is the smallest among all P. putida strains sequenced to date.

<>

<1>Wang, F., Elmquist, C.E., Rizzo, C.J., Stone, M.P.
<2>Base-displaced intercalated structure of the food mutagen 2-amino-3methylimidazo[4,5-f]quinoline (IQ) in the recognition sequence  of the NarI restriction enzyme, a hotspot for-2 bp deletions.
<3>ACS Abstracts
<4>230
<5>U1858-U1859
<6>2005
<7>2-Amino-3-methylimidazo[4,5-f]quinoline (IQ) is a prominent member of heterocyclic mutagens
and carcinogens found during the cooking of meats. The C-G1-G2-C-G3-C-C recognition sequence
of the NarI restriction enzyme represents a hot spot for -2 frameshift mutations at G3. We
prepared and purified the oligodeoxynucleotide duplex containing the [IQ]dG adduct positioned
in the C-[IQ]G3-C NarI context, which was named NarIIQ3. NMR data revealed a base-displaced
intercalated conformation. Watson-Crick base pairing was perturbed at the adduct site. The
largest chemical shift perturbations were observed for the base aromatic protons in the
complementary strand, opposite to the adduct. Chemical shift perturbations were also observed
for the 31P resonances corresponding to the linkages between C6 and [IQ]G7, [IQ]G7 and C8, G17
and C18, and C18 and G19, the phosphodiesters around the adduct. There were twenty-one NOEs
observed between the IQ moiety and DNA protons. The [IQ]G3 aromatic protons had strong NOEs
with the sugar protons of the opposite C18, G17 and G19 in the complementary strand, and the
methyl protons of [IQ]G3 showed NOEs to C8 H6, G19 H8, G17 NH1 and G19 NH1. Deoxyribose
pseudorotational angles (P) were estimated by examining the 3JHH of sugar protons. J1'-2'
and J1'-2' were measured from ECOSY spectra, while the intensities of H2'-H3' and
H3'-H4' crosspeaks were determined from DQF-COSY spectra. Molecular dynamics calculations on
NarIIQ3, restrained by 1H NOEs and J couplings, yielded ensembles of intercalated structures
in which the modified guanine was in a syn alignment and displaced. NOE intensities calculated
using complete relaxation matrix theory were in agreement with experimental NOE intensities.
Structural differences between NarIIQ3 and NarIAF3 could provide insight into the greater
biological activity of IQ than that of AF. Funded by grant CA-55678.

<>

<1>Wang, F., Wang, J., Jian, H., Zhang, B., Li, S., Wang, F., Zeng, X., Gao, L., Bartlett, D.H., Yu, J., Hu, S., Xiao, X.
<2>Environmental adaptation: genomic analysis of the piezotolerant and psychrotolerant deep-sea iron reducing bacterium Shewanella piezotolerans WP3.
<3>PLoS ONE
<4>3
<5>E1937
<6>2008
<7>Shewanella species are widespread in various environments. Here, the
genome sequence of Shewanella piezotolerans WP3, a piezotolerant and
psychrotolerant iron reducing bacterium from deep-sea sediment was
determined with related functional analysis to study its environmental
adaptation mechanisms. The genome of WP3 consists of 5,396,476 base pairs
(bp) with 4,944 open reading frames (ORFs). It possesses numerous genes or
gene clusters which help it to cope with extreme living conditions such as
genes for two sets of flagellum systems, structural RNA modification,
eicosapentaenoic acid (EPA) biosynthesis and osmolyte transport and
synthesis. And WP3 contains 55 open reading frames encoding putative
c-type cytochromes which are substantial to its wide environmental
adaptation ability. The mtr-omc gene cluster involved in the insoluble
metal reduction in the Shewanella genus was identified and compared. The
two sets of flagellum systems were found to be differentially regulated
under low temperature and high pressure; the lateral flagellum system was
found essential for its motility and living at low temperature.

<>

<1>Wang, G.H.
<2>High-Efficiency Thermal Asymmetric Interlaced PCR (hiTAIL-PCR) for Determination of a Highly Degenerated Prophage WO Genome in a Wolbachia Strain Infecting a Fig Wasp Species.
<3>Appl. Environ. Microbiol.
<4>79
<5>7476-7481
<6>2013
<7>
<>

<1>Wang, G.L., Zhou, L.Y., Luo, H.Q., Li, N.B.
<2>Electrochemical strategy for sensing DNA methylation and DNA methyltransferase activity.
<3>Anal. Chim. Acta
<4>768
<5>76-81
<6>2013
<7>The present work demonstrates a novel signal-off electrochemical method for the determination
of DNA methylation and the assay of
methyltransferase activity using the electroactive complex
[Ru(NH3)(6)](3+) (RuHex) as a signal transducer. The assay exploits the
electrostatic interactions between RuHex and DNA strands. Thiolated
single strand DNA1 was firstly self-assembled on a gold electrode via
Au-S bonding, followed by hybridization with single strand DNA2 to form
double strand DNA containing specific recognition sequence of DNA
adenine methylation MTase and methylation-responsive restriction
endonuclease Dpn I. The double strand DNA may adsorb lots of
electrochemical species (Ru(NH3)(6)](3+)) via the electrostatic
interaction, thus resulting in a high electrochemical signal. In the
presence of DNA adenine methylation methyltransferase and
S-adenosyl-L-methionine, the formed double strand DNA was methylated by
DNA adenine methylation methyltransferase, then the double strand DNA
can be cleaved by methylation-responsive restriction endonuclease Dpn
I, leading to the dissociation of a large amount of signaling probes
from the electrode. As a result, the adsorption amount of RuHex
reduced, resulting in a decrease in electrochemical signal. Thus, a
sensitive electrochemical method for detection of DNA methylation is
proposed. The proposed method yielded a linear response to
concentration of Dam MTase ranging from 0.25 to IOU mL(-1) with a
detection limit of 0.18 U mL(-1) (S/N = 3), which might promise this
method as a good candidate for monitoring DNA methylation in the
future.

<>

<1>Wang, H., Guan, S., Quimby, A., Cohen-Karni, D., Pradhan, S., Wilson, G., Roberts, R.J., Zhu, Z., Zheng, Y.
<2>Comparative characterization of the PvuRts1I family of restriction enzymes and their application in mapping genomic 5-hydroxymethylcytosine.
<3>Nucleic Acids Res.
<4>39
<5>9294-9305
<6>2011
<7>PvuRts1I is a modification-dependent restriction endonuclease that recognizes
5-hydroxymethylcytosine (5hmC) as well as
5-glucosylhydroxymethylcytosine (5ghmC) in double-stranded DNA. Using
PvuRts1I as the founding member, we define a family of homologous proteins
with similar DNA modification-dependent recognition properties. At the
sequence level, these proteins share a few uniquely conserved features. We
show that these enzymes introduce a double-stranded cleavage at the
3'-side away from the recognized modified cytosine. The distances between
the cleavage sites and the modified cytosine are fixed within a narrow
range, with the majority being 11-13 nt away in the top strand and 9-10 nt
away in the bottom strand. The recognition sites of these enzymes
generally require two cytosines on opposite strand around the cleavage
sites, i.e. 5'-CN(11-13) downward arrowN(9-10)G-3'/3'-GN(9-10) downward
arrowN(11-13)C-5', with at least one cytosine being modified for efficient
cleavage. As one potential application for these enzymes is to provide
useful tools for selectively mapping 5hmC sites, we have compared the
relative selectivity of a few PvuRts1I family members towards different
forms of modified cytosines. Our results show that the inherently
different relative selectivity towards modified cytosines can have
practical implications for their application. By using AbaSDFI, a PvuRts1I
homolog with the highest relative selectivity towards 5ghmC, to analyze
rat brain DNA, we show it is feasible to map genomic 5hmC sites close to
base resolution. Our study offers unique tools for determining more
accurate hydroxymethylomes in mammalian cells.

<>

<1>Wang, H., Li, H., Shao, Z., Liao, S., Johnstone, L., Rensing, C., Wang, G.
<2>Genome Sequence of Deep-Sea Manganese-Oxidizing Bacterium Marinobacter manganoxydans MnI7-9.
<3>J. Bacteriol.
<4>194
<5>899-900
<6>2012
<7>Here we report the draft genome of Marinobacter manganoxydans MnI7-9, isolated from a deep-sea
hydrothermal vent in the Indian Ocean and capable
of oxidizing manganese even when there is a very high concentration of
Mn(2+). The strain also displayed high resistance and adsorption ability
toward many metal(loid)s.

<>

<1>Wang, H., Sivonen, K., Rouhiainen, L., Fewer, D.P., Lyra, C., Rantala-Ylinen, A., Vestola, J., Jokela, J., Rantasarkka, K., Li, Z., Liu, B.
<2>Genome-derived insights into the biology of the hepatotoxic bloom-forming cyanobacterium Anabaena sp. strain 90.
<3>BMC Genomics
<4>13
<5>613
<6>2012
<7>ABSTRACT: BACKGROUND: Cyanobacteria can form massive toxic blooms in fresh and
brackish bodies of water and are frequently responsible for the poisoning of
animals and pose a health risk for humans. Anabaena is a genus of filamentous
diazotrophic cyanobacteria commonly implicated as a toxin producer in blooms in
aquatic ecosystems throughout the world. The biology of bloom-forming
cyanobacteria is poorly understood at the genome level. RESULTS: Here, we report
the complete sequence and comprehensive annotation of the bloom-forming Anabaena
sp. strain 90 genome. It comprises two circular chromosomes and three plasmids
with a total size of 5.3 Mb, encoding a total of 4,738 genes. The genome is
replete with mobile genetic elements. Detailed manual annotation demonstrated
that almost 5% of the gene repertoire consists of pseudogenes. A further 5% of
the genome is dedicated to the synthesis of small peptides that are the products
of both ribosomal and nonribosomal biosynthetic pathways. Inactivation of the
hassallidin (an antifungal cyclic peptide) biosynthetic gene cluster through a
deletion event and a natural mutation of the buoyancy-permitting gvpG gas vesicle
gene were documented. The genome contains a large number of genes encoding
restriction-modification systems. Two novel excision elements were found in the
nifH gene that is required for nitrogen fixation. CONCLUSIONS: Genome analysis
demonstrated that this strain invests heavily in the production of bioactive
compounds and restriction-modification systems. This well-annotated genome
provides a platform for future studies on the ecology and biology of these
important bloom-forming cyanobacteria.

<>

<1>Wang, H., Wang, L., Ju, J., Yu, B., Ma, Y.
<2>Genome Sequence of Sporolactobacillus laevolacticus DSM442, an Efficient Polymer-Grade D-Lactate Producer from Agricultural Waste Cottonseed as a Nitrogen  Source.
<3>Genome Announcements
<4>1
<5>e01100-13
<6>2013
<7>Sporolactobacillus laevolacticus DSM442 is an efficient polymer-grade d-lactic acid producer
from low-cost agricultural waste cottonseed powder as the sole
nitrogen source. Here we present a 3.59-Mb assembly of its genome sequence, which
might provide useful information to further improve the strain for higher
production titers.

<>

<1>Wang, H.-L., Wang, H.-R., Zhang, W.-W., Sun, L.
<2>Cloning and analysis of the Vibrio harveyi dam gene.
<3>Wei Sheng Wu Xue Bao
<4>47
<5>855-859
<6>2007
<7>The DNA adenine methylase (dam) gene was cloned by degenerate PCR from Vibrio harveyi strain
T4. The gene was 840bp in length and encoded a
putative protein of 279 amino acids that shared relatively high
homology with the Dam of other Vibrias, especially with that of V.
parahaemolyticus (96% in identity). The V. harveyi dam gene was
subcloned into plasmid pBR322 and the resulting plasmid pBD was
introduced into the E. coli strain ER2925 in which the dam gene had
been knocked out. Dpn I, Dpn II, and Sau3A I restriction enzyme
analysis of the genomic DNA of ER2925 transformed with p13D indicated
that the cloned V. harveyi dam gene could functionally complement the
E. coli dam mutant and methylate E. coli chromosome at the GATC sites.
The 3251 bp upstream region of V. harveyi dam was obtained by genome
walking and analyzed at the sequence level. It was found that this 3251
bp region contained two complete open reading frames (ORF) : one was of
1101 bp in length and the other was of 1503 bp in length. The predicted
amino acid sequence of ORF1101 shared 91% identity with the
3-dehydroquinate synthase of V. parahaemolyticus. The amino acid
sequence of ORF1503 shared 80% identity with V. parahaemolylicus DamX.
A truncated ORF was found at the upstream of ORF1101, encoding 169
amino acids that shared 94% identity with the shikimate kinase of V.
parahaemolyticus. These three genes, together with dam, were arranged
in the order of shikimate kinase-3-dehydroquinate synthase-damX-dam.
The region immediate upstream of the V. harveyi dam structural gene was
cloned in three fragments of different length: 78bp, 112 bp and 477bp
(named P78, P112, and P477, respectively) and tested for promoter
activity. The results showed that, while all the three fragments had
detectable promoter activities, the activity of P78 appeared to be
higher than that of P-112 and P-477.

<>

<1>Wang, H.C., Ko, W.C., Shu, H.Y., Chen, P.L., Wang, Y.C., Wu, C.J.
<2>Genome Sequence of Aeromonas taiwanensis LMG 24683T, a Clinical Wound Isolate from Taiwan.
<3>Genome Announcements
<4>2
<5>e00579-14
<6>2014
<7>Aeromonas taiwanensis was first described in 2010 on the basis of one clinical wound isolate
(strain LMG 24683(T) = A2-50(T) = CECT 7403(T)) from Taiwan. We
present here the genome sequence of A. taiwanensis LMG 24683(T), which carries
several genes encoding virulence determinants and Ambler class C and D
beta-lactamases.

<>

<1>Wang, H.F., Muren, N.B., Ordinario, D., Gorodetsky, A.A., Barton, J.K., Nuckolls, C.
<2>Transducing methyltransferase activity into electrical signals in a carbon nanotube-DNA device.
<3>Chem. Sci.
<4>3
<5>62-65
<6>2012
<7>This study creates a device where the DNA is electronically integrated to serve as both the
biological target and electrical transducer in a CNT-DNA-CNT device. We detect DNA binding and
methylation by the methyltransferase M.SssI at the single molecule level. We demonstrate
sequence-specific, reversible binding of M.SssI and protein-catalyzed methylation that alters
the protein-binding affinity of the device. This device, which relies on the exquisite
electrical sensitivity of DNA, represents a unique route for the specific, single molecule
detection of enzymatic activity.

<>

<1>Wang, H.X., Wang, Y.S.
<2>6-Thioguanine Perturbs Cytosine Methylation at the CpG Dinucleotide Site by DNA Methyltransferases in Vitro and Acts as a DNA Demethylating Agent in Vivo.
<3>Biochemistry
<4>48
<5>2290-2299
<6>2009
<7>Thiopurines are among the most successful chemotherapeutic agents for treating a number of
human diseases including acute lymphoblastic
leukemia. The mechanisms through which the thiopurines elicit their
cytotoxic effects remain unclear. We postulate that the incorporation
of 6-thioguanine into the CpG site may perturb the
methyltransferase-mediated cytosine methylation at this site, thereby
interfering with the epigenetic pathways of gene regulation. To gain
biochemical evidence for this hypothesis, we assessed, by using a
restriction enzyme digestion coupled with LC-MS/MS method, the impact
of 6-thioguanine on cytosine methylation mediated by two DNA
methyltransferases, human DNMT1 and bacterial HpaII. Our results
revealed that the incorporation of 6-thioguanine into the CpG site
could affect the methylation of the cytosine residue by both
methyltransferases and the effect on cytosine methylation is dependent
on the position of 6-thioguanine with respect to the cytosine to be
methylated. The presence of 6-thioguanine at the methylated CpG site
enhanced the DNMT1-mediated methylation of the opposing cytosine in the
complementary strand, whereas the presence of 6-thioguanine at the
unmethylated CpG site abolished almost completely the methylation of
its 5' adjacent cytosine by both DNMT1 and HpaII. We further
demonstrated that the treatment of Jurkat T cells, which were derived
from acute lymphoblastic leukemia, with 6-thioguanine could result in
an appreciable drop in the level of global cytosine methylation. These
results showed that 6-thioguanine, after being incorporated into DNA,
may perturb the epigenetic pathway of gene regulation.

<>

<1>Wang, H.Z., Wong, M.M.L., O'Toole, D., Mak, M.M.H., Wu, R.S.S., Kong, R.Y.C.
<2>Identification of a DNA methyltransferase gene carried on a pathogenicity island-like element (VPAI) in Vibrio parahaemolyticus and  its prevalence among clinical and environmental isolates.
<3>Appl. Environ. Microbiol.
<4>72
<5>4455-4460
<6>2006
<7>In this study we identified a putative virulence-associated DNA methyltransferase (MTase) gene
carried on a novel 22.79-kb
pathogenicity island-like element (VPAI) in V. parahaemolyticus. The V.
parahaemolyticus MTase gene was shown by PCR to be prevalent (> 98%) in
pandemic thermostable direct hemolysin gene-positive isolates, which
suggests that VPAI may confer unique virulence traits to pandemic
strains of V. parahaemolyticus.

<>

<1>Wang, J., Hevi, S., Kurash, J.K., Lei, H., Gay, F., Bajko, J., Su, H., Sun, W., Chang, H., Xu, G., Gaudet, F., Li, E., Chen, T.
<2>The lysine demethylase LSD1 (KDM1) is required for maintenance of global DNA methylation.
<3>Nat. Genet.
<4>41
<5>125-129
<6>2009
<7>Histone methylation and DNA methylation cooperatively regulate chromatin structure and gene
activity. How these two systems coordinate with each
other remains unclear. Here we study the biological function of
lysine-specific demethylase 1 (LSD1, also known as KDM1 and AOF2), which
has been shown to demethylate histone H3 on lysine 4 (H3K4) and lysine 9
(H3K9). We show that LSD1 is required for gastrulation during mouse
embryogenesis. Notably, targeted deletion of the gene encoding LSD1
(namely, Aof2) in embryonic stem (ES) cells induces progressive loss of
DNA methylation. This loss correlates with a decrease in DNA
methyltransferase 1 (Dnmt1) protein, as a result of reduced Dnmt1
stability. Dnmt1 protein is methylated in vivo, and its methylation is
enhanced in the absence of LSD1. Furthermore, Dnmt1 can be methylated by
Set7/9 (also known as KMT7) and demethylated by LSD1 in vitro. Our
findings suggest that LSD1 demethylates and stabilizes Dnmt1, thus
providing a previously unknown mechanistic link between the histone and
DNA methylation systems.

<>

<1>Wang, J., Hu, W., Lux, R., He, X., Li, Y., Shi, W.
<2>Natural Transformation of Myxococcus xanthus.
<3>J. Bacteriol.
<4>193
<5>2122-2132
<6>2011
<7>Myxococcus xanthus belongs to the delta class of the proteobacteria and is notable for its
complex life-style with social behaviors and
relatively large genome. Although previous observations have suggested
the existence of horizontal gene transfer in M. xanthus, its ability to
take up exogenous DNA via natural transformation has not been
experimentally demonstrated. In this study, we achieved natural
transformation in M. xanthus using the autonomously replicating
myxobacterial plasmid pZJY41 as donor DNA. M. xanthus exopolysaccharide
(EPS) was shown to be an extracellular barrier for transformation.
Cells deficient in EPS production, e. g., mutant strains carrying Delta
difA or Delta epsA, became naturally transformable. Among the inner
barriers to transformation were restriction-modification systems in M.
xanthus, which could be partially overcome by methylating DNA in vitro
using cell extracts of M. xanthus prior to transformation. In addition,
the incubation time of DNA with cells and the presence of divalent
magnesium ion affected transformation frequency of M. xanthus.
Furthermore, we also observed a potential involvement of the type IV
pilus system in the DNA uptake machinery of M. xanthus. The natural
transformation was totally eliminated in the Delta pilQ/epsA and Delta
tgl/epsA mutants, and null mutation of pilB or pilC in an Delta epsA
background diminished the transformation rate. Our study, to the best
of our knowledge, provides the first example of a naturally
transformable species among deltaproteobacteria.

<>

<1>Wang, J., Jiang, Y., Vincent, M., Sun, Y., Yu, H., Wang, J., Bao, Q., Kong, H., Hu, S.
<2>Complete genome sequence of bacteriophage T5.
<3>Virology
<4>332
<5>45-65
<6>2005
<7>The 121,752-bp genome sequence of bacteriophage T5 was determined; the
linear, double-stranded DNA is nicked in one of the strands and has large
direct terminal repeats of 10,139 bp (8.3%) at both ends. The genome
structure is consistently arranged according to its lytic life cycle. Of
the 168 potential open reading frames (ORFs), 61 were annotated; these
annotated ORFs are mainly enzymes involved in phage DNA replication,
repair, and nucleotide metabolism. At least five endonucleases that
believed to help inducing nicks in T5 genomic DNA, and a DNA ligase gene
was found to be split into two separate ORFs. Analysis of T5 early
promoters suggests a probable motif AAA{3, 4 T}nTTGCTT{17, 18
n}TATAATA{12, 13 W}{10 R} for strong promoters that may strengthen the
step modification of host RNA polymerase, and thus control transcription
of phage DNA. The distinct protein domain profile and a mosaic genome
structure suggest an origin from the common genetic pool.

<>

<1>Wang, J., Kim, H.-H., Yuan, X., Herrin, D.L.
<2>Purification, biochemical characterization and protein-DNA interactions of the I-CreI endonuclease produced in Escherichia coli.
<3>Nucleic Acids Res.
<4>25
<5>3767-3776
<6>1997
<7>I-CreI is a member of the LAGLI-DADG family of homing nucleases; however, unlike most members
of this family it contains only a single copy of this signature motif.  I-CreI was
over-expressed in Escherichia coli, and a simple purification protocol developed that gave
reasonably pure protein in high yield.  Size-exclusion chromatography and chemical
cross-linking indicated that the protein is a dimer in solution.  DNA cleavage by I-CreI was
absolutely dependent on Mg2+ (or Mn2+), and was inhibited by monovalent cations.  I-CreI
displayed a surprisingly high temperature optimum (>50oC), with full activity occurring even
at 70oC.  Interestingly, SDS was needed for efficient release of the cleavage products from
the protein, indicating formation of very stable DNA-protein complexes.  In contrast to these
robust characteristics, purified I-CreI was unstable; however, it could be stabilized by the
addition of either target or non-target DNA.  Mobility shift assays revealed that I-CreI binds
to DNA in the absence of Mg2+.  Hydroxyl radical footprinting showed that I-CreI strongly
protected the backbone of a continuous stretch of at least 12 nt on each strand that were
shifted, relative to each other, by 2 bp in the 3' direction.  Methylation protection and
interference analyses were also performed, and together with the hydroxyl radical
footprinting, indicate that I-CreI binds in both the major and minor grooves of its target
DNA.

<>

<1>Wang, J., Kuenzel, S., Baines, J.F.
<2>Draft Genome Sequences of 11 Staphylococcus epidermidis Strains Isolated from Wild Mouse Species.
<3>Genome Announcements
<4>2
<5>e01148-13
<6>2014
<7>We report here the draft genome sequences of 11 strains of Staphylococcus epidermidis, a
common bacterium inhabiting the skin of humans and other animals.
These isolates, obtained from five mouse species, provide valuable information on
the native Staphylococcus spp. of this important model organism and form a basis
for studying host-bacterial interactions in their natural environment.

<>

<1>Wang, J., Li, Y., Guo, J., Zhang, X., Guo, Y., Chang, D., Li, T., Xu, G., Dai, W., Liu, C.
<2>Genome Sequence of Staphylococcus aureus Strain LCT-SA67, a Space Flight Strain with Altered Carbon Source Utilization Properties.
<3>Genome Announcements
<4>2
<5>e00095-14
<6>2014
<7>An increasing number of studies have confirmed that space flight environments can have a
significant effect on a variety of microbial properties. To explore the
effect of these environments on Staphylococcus aureus, we present the draft
genome sequence of an S. aureus strain, named LCT-SA67, which was isolated after
space flight.

<>

<1>Wang, J., Lin, T.
<2>New type II restriction endonuclease, useful for cutting and recognizing DNA found in bacteria, e.g. H. pylori, at a  particular sequence of the nucleotide-recombinant enzyme production via plasmid expression in host cell for use in DNA cleavage.
<3>US Patent Office
<4>US 20050202443
<5>
<6>2005
<7>NOVELTY - A type II restriction endonuclease comprising 538 amino acids (SEQ ID No. 3), given
in the specification, and recognizing and cutting DNA only at a particular sequence of
nucleotides, is new. DETAILED DESCRIPTION - INDEPENDENT CLAIMS are
included for: (1) an isolated nucleic acid encoding the type II
restriction endonuclease; (2) a vector comprising the nucleic acid; (3)
a transformed cell comprising the vector; and (4) a recombinant enzyme
specifically recognizing DNA at a particular sequence of nucleotides
and cleaves DNA downstream of the DNA sequence of nucleotides at the
fourth base in the upper strand and the fifth base in the lower strand,
and the recombinant enzyme composing the amino acid sequence of SEQ ID
No. 3. BIOTECHNOLOGY - Preferred Enzyme: The type II restriction
endonuclease recognizes and cuts the DNA is from bioorganism. The DNA
is manually synthesized. The particular sequence comprises the sequence
5'-CCATC-3' (SEQ ID No. 1). The type II restriction endonuclease
cleaves DNA downstream of said DNA sequence of nucleotides at the
fourth base in the upper strand and the fifth base in the lower strand
of SEQ ID No. 1. The type II restriction endonuclease is an enzyme
derived from microorganism. The microorganisms are Helicobacter pylori.
Preferred Nucleic Acid: The nucleic acid encoding the enzyme comprises
1617 base pairs (SEQ ID No. 2), given in the specification. The nucleic
acid is originated from Helicobacter pylori. USE - The type II
restriction endonuclease is useful for cutting and recognizing DNA
found in bacteria, e.g. Helicobacter pylori, at a particular sequence
of the nucleotide (claimed). ADVANTAGE - The invention provides a
DNA-cutting enzyme that can specifically recognize and cut a particular
nucleotide sequence in order to provide alternative choices for
cleaving DNA in the biotechnological manipulation of genetic
engineering and gene cloning, and to improve the cutting efficiency, as
compared to prior art. EXAMPLE - Experimental protocols are described
but no results are given.

<>

<1>Wang, J., Liu, Y., Wan, D., Fang, X., Li, T., Guo, Y., Chang, D., Su, L., Wang, Y., Zhao, J., Liu, C.
<2>Whole-Genome Sequence of Staphylococcus aureus Strain LCT-SA112.
<3>J. Bacteriol.
<4>194
<5>4124
<6>2012
<7>Staphylococcus aureus is a facultative anaerobic Gram-positive coccal bacterium.  S. aureus is
the most common species of Staphylococcus to cause staphylococcal
infections, which are very common in clinical medicine. Here we report the genome
sequence of S. aureus strain LCT-SA112, which was isolated from S. aureus subsp.
aureus CGMCC 1.230.

<>

<1>Wang, J., Stephan, R., Power, K., Yan, Q., Hachler, H., Fanning, S.
<2>Nucleotide sequences of 16 transmissible plasmids identified in nine multidrug-resistant Escherichia coli isolates expressing an ESBL phenotype isolated from food-producing animals and healthy humans.
<3>J. Antimicrob. Chemother.
<4>69
<5>2658-2668
<6>2014
<7>OBJECTIVES: Nine extended-spectrum beta-lactamase (ESBL)-producing Escherichia
coli isolated from healthy humans and food-producing animals were found to
transfer their cefotaxime resistance marker at high frequency in laboratory
conjugation experiments. The objective of this study was to completely
characterize 16 transmissible plasmids that were detected in these bacterial
isolates. METHODS: The nucleotide sequences of all 16 plasmids were determined
from transconjugants using next-generation sequencing technology. Open reading
frames were assigned using Rapid Annotation using Subsystem Technology and
analysed by BLASTn and BLASTp. The standard method was used for plasmid
multilocus sequence typing (pMLST) analysis. Plasmid structures were subsequently
confirmed by PCR amplification of selected regions. RESULTS: The complete
circularized nucleotide sequence of 14 plasmids was determined, along with that
of a further two plasmids that could not be confirmed as closed. These ranged in
size from 1.8 to 166.6 kb. Incompatibility groups and pMLSTs identified included
IncI1/ST3, IncI1/ST36, IncN/ST1, IncF and IncB/O, and those of the same Inc types
presented a similar backbone structure despite being isolated from different
sources. Eight plasmids contained bla(CTX-M-1) genes that were associated with
either ISEcp1 or IS26 insertion sequence elements. Six plasmids isolated from
humans and chickens were identical or closely related to the IncI1 reference
plasmid, R64. CONCLUSIONS: These data, based on comparative sequence analysis,
highlight the successful spread of blaESBL-harbouring plasmids of different Inc
types among isolates of human and food-producing animal origin and provide
further evidence for potential dissemination routes.

<>

<1>Wang, J.-T., Lin, T.-L.
<2>Purified HpyC1I and its use as a restriction endonuclease.
<3>US Patent Office
<4>US 7244603 A
<5>
<6>2007
<7>Disclosed is a novel type II restriction endonuclease.  Such enzyme recognizes a particular
non-palindromic sequence of 5 oligonucleotides and cleaves DNA downstream of the DNA
recognition sequence of nucleotides at the fourth base in the upper strand and the fifth base
in the lower strand, and forms a one-base protruding end in the 5'-end after cleavage.  The
recognition and cleavage site of HpyC1I is identical to the known restriction endonuclease
BccI respectively, but the nucleotide sequence and the amino acid sequence are different from
any other known restriction enzymes.

<>

<1>Wang, J.-T., Lin, T.-L.
<2>Type II restriction endonuclease.
<3>Taiwanese Patent Office
<4>TW 284675 B
<5>
<6>2007
<7>
<>

<1>Wang, K., Chen, J., Yao, H., Lu, C.
<2>Whole-Genome Sequence of Streptococcus suis Serotype 4 Reference Strain 6407.
<3>Genome Announcements
<4>2
<5>e00770-14
<6>2014
<7>We report here the second complete genome sequence of Streptococcus suis serotype 4 (strain
6407). The genome is 2,292,360 bp in length, covering 2,239 coding
sequences, 58 tRNAs, and 4 rRNA loci.

<>

<1>Wang, K., Chen, J., Yao, H., Lu, C.
<2>Whole-Genome Sequence of Streptococcus suis Serotype 3 Strain YB51.
<3>Genome Announcements
<4>1
<5>e00884-13
<6>2013
<7>We report here the second complete genome sequence of Streptococcus suis serotype 3 (strain
YB51). The genome is 2,043,655 bp in length, which is 14,840 bp longer
than the first reported genome of the same serotype, and it covers 2,012 coding
sequences, 56 tRNAs, and 4 rRNA loci.

<>

<1>Wang, K., Fan, W., Cai, L., Huang, B., Lu, C.
<2>Genetic analysis of the capsular polysaccharide synthesis locus in 15 Streptococcus suis serotypes.
<3>FEMS Microbiol. Lett.
<4>324
<5>117-124
<6>2011
<7>
<>

<1>Wang, K., Yao, H., Lu, C., Chen, J.
<2>Complete Genome Sequence of Streptococcus suis Serotype 16 Strain TL13.
<3>Genome Announcements
<4>1
<5>e00394-13
<6>2013
<7>We report here the first complete genome sequence of Streptococcus suis serotype  16, which
has been identified to be zoonotic. The sequenced strain TL13 was
isolated from a pig in China. The genome is 2,038,146 bp in length, covering
1,950 coding sequences, 53 tRNAs, and 4 rRNA loci.

<>

<1>Wang, K.-Y., Chen, C.-C., Shen, C.-Kun.J.
<2>Active DNA demethylation of the vertebrate genomes by DNA methyltransferases: deaminase, dehydroxymethylase or demethylase?
<3>Epigenomics
<4>6
<5>353-363
<6>2014
<7>Vertebrate DNA methyltransferases (DNMTs) have been thought to primarily function to
covalently add a methyl group to the 5-position of cytosine. However, recent discovery of the
DNA demethylation and dehydroxymethylation activities of DNMTs in vitro suggest new routes to
complete the dynamic cycle of DNA methylation-demethylation of the vertebrate genomes. The in
vitro reaction conditions suggest that vertebrate DNMTs can switch from DNA methylases to DNA
dehydroxymethylases under oxidative stress and to DNA demethylases in the presence of calcium
ion under nonreducing conditions. These environmental parameters provide clues regarding the
choices in vivo of DNMT activities utilized in different physiological systems. In particular,
the nature of these parameters suggest that the DNA demethylation and dehydroxymethylation
activities of the vertebrate DNMTs play essential roles in multiple biological processes
including early embryo development, regulation of neuronal plasticity, tumorigenesis and
hormone-regulated transcription.

<>

<1>Wang, L., Chen, S., Deng, Z.
<2>Phosphorothioation:  An unusual post-replicative modification on the DNA backbone.
<3>DNA Replication-Current Advances, InTech, Herve Seligmann, 
<4>
<5>57-74
<6>2011
<7>DNA molecules are polymers composed of basic repeating subunits of deoxyribonucleotides, which
consist of the deoxyribose sugar, phosphate groups, and a nitrogenous base.  They appear to
fulfill all requirements necessary to maintain the genetic function of DNA.  The five elements
of nitrogen, phosphorus, carbon, hydrogen, and oxygen had been regarded as the canonical
composition of DNA until the discovery of phosphorothioation, with a sixth element, sulfur,
identified as an additional naturally occurring constituent on the DNA backbone, as a
sequence-selective, stereospecific post-replicative modification governed by the dnd gene
cluster.  Unlike any other DNA or RNA modification system, DNA phosphorothioation is the
first-described physiological modification of the DNA sugar-phosphorothioation is the
first-described physiological modification of the DNA sugar-phosphate backbone.

<>

<1>Wang, L., Chen, S., Vergin, K.L., Giovannoni, S.J., Chan, S.W., Demott, M.S., Taghizadeh, K., Cordero, O.X., Cutler, M., Timberlake, S., Alm, E.J., Polz, M.F., Pinhassi, J., Deng, Z., Dedon, P.C.
<2>DNA phosphorothioation is widespread and quantized in bacterial genomes.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>2963-2968
<6>2011
<7>Phosphorothioate (PT) modification of DNA, with sulfur replacing a nonbridging phosphate
oxygen, was recently discovered as a product of the
dnd genes found in bacteria and archaea. Given our limited understanding
of the biological function of PT modifications, including sequence
context, genomic frequencies, and relationships to the diversity of dnd
gene clusters, we undertook a quantitative study of PT modifications in
prokaryotic genomes using a liquid chromatography-coupled tandem
quadrupole mass spectrometry approach. The results revealed a diversity of
unique PT sequence contexts and three discrete genomic frequencies in a
wide range of bacteria. Metagenomic analyses of PT modifications revealed
unique ecological distributions, and a phylogenetic comparison of dnd
genes and PT sequence contexts strongly supports the horizontal transfer
of dnd genes. These results are consistent with the involvement of PT
modifications in a type of restriction-modification system with wide
distribution in prokaryotes.

<>

<1>Wang, L., Chen, S., Xu, T., Taghizadeh, K., Wishnok, J.S., Zhou, X., You, D., Deng, Z., Dedon, P.C.
<2>Phosphorothioation of DNA in bacteria by dnd genes.
<3>Nat. Chem. Biol.
<4>3
<5>709-710
<6>2007
<7>Modifications of the canonical structures of DNA and RNA play critical roles in cell
physiology, DNA replication, transcription and translation in all organisms.
We now report that bacterial dnd gene clusters incorporate sulfur into the DNA
backbone as a sequence-selective, stereospecific phosphorothioate modification.
To our knowledge, unlike any other DNA or RNA modification systems, DNA
phosphorothioation by dnd gene clusters is the first physiological modification
described on the DNA backbone.

<>

<1>Wang, L., Hatem, A., Catalyurek, U.V., Morrison, M., Yu, Z.
<2>Metagenomic insights into the carbohydrate-active enzymes carried by the microorganisms adhering to solid digesta in the rumen of cows.
<3>PLoS ONE
<4>8
<5>E78507
<6>2013
<7>The ruminal microbial community is a unique source of enzymes that underpin the
conversion of cellulosic biomass. In this study, the microbial consortia adherent
on solid digesta in the rumen of Jersey cattle were subjected to an
activity-based metagenomic study to explore the genetic diversity of
carbohydrolytic enzymes in Jersey cows, with a particular focus on cellulases and
xylanases. Pyrosequencing and bioinformatic analyses of 120 carbohydrate-active
fosmids identified genes encoding 575 putative Carbohydrate-Active Enzymes
(CAZymes) and proteins putatively related to transcriptional regulation,
transporters, and signal transduction coupled with polysaccharide degradation and
metabolism. Most of these genes shared little similarity to sequences archived in
databases. Genes that were predicted to encode glycoside hydrolases (GH) involved
in xylan and cellulose hydrolysis (e.g., GH3, 5, 9, 10, 39 and 43) were well
represented. A new subfamily (S-8) of GH5 was identified from contigs assigned to
Firmicutes. These subfamilies of GH5 proteins also showed significant
phylum-dependent distribution. A number of polysaccharide utilization loci (PULs)
were found, and two of them contained genes encoding Sus-like proteins and
cellulases that have not been reported in previous metagenomic studies of samples
from the rumens of cows or other herbivores. Comparison with the large
metagenomic datasets previously reported of other ruminant species (or cattle
breeds) and wallabies showed that the rumen microbiome of Jersey cows might
contain differing CAZymes. Future studies are needed to further explore how host
genetics and diets affect the diversity and distribution of CAZymes and
utilization of plant cell wall materials.

<>

<1>Wang, L., Qiu, Y., Chen, Z., Xu, J., Wang, Z., Ke, Y., Li, T., Wang, D., Huang, L., Yu, Y., Zhen, Q.
<2>Draft Genome Sequence of Brucella abortus BCB027, a Strain Isolated from a Domestic Deer.
<3>Genome Announcements
<4>1
<5>e00130-12
<6>2013
<7>Many Brucella species are isolated from nonpreferred hosts, and these bacteria may show
genetic differences from isolates from the preferred hosts. Here, we report the draft genome
sequence of Brucella abortus BCB027, a novel strain isolated from a domestic deer.

<>

<1>Wang, L., Wang, S., He, Q., Yu, T., Li, Q., Hong, B.
<2>Draft Genome Sequence of Streptomyces globisporus C-1027, Which Produces an Antitumor Antibiotic Consisting of a Nine-Membered Enediyne with a Chromoprotein.
<3>J. Bacteriol.
<4>194
<5>4144
<6>2012
<7>Streptomyces globisporus C-1027 is the producer of antitumor antibiotic C-1027, a
nine-membered enediyne-containing compound. Here we present a draft genome
sequence of S. globisporus C-1027 containing the intact biosynthetic gene cluster
for this antibiotic. The genome also carries numerous sets of genes for the
biosynthesis of diverse secondary metabolites.

<>

<1>Wang, L., Xie, Y., Li, Q., He, N., Yao, E., Xu, H., Yu, Y., Chen, R., Hong, B.
<2>Draft Genome Sequence of Streptomyces sp. Strain SS, Which Produces a Series of Uridyl Peptide Antibiotic Sansanmycins.
<3>J. Bacteriol.
<4>194
<5>6988-6989
<6>2012
<7>Streptomyces sp. SS produces a series of uridyl peptide antibiotic sansanmycins.  Here, we
present a draft genome sequence of Streptomyces sp. SS containing the
biosynthetic gene cluster for the antibiotics. The identification of the
biosynthetic gene cluster of sansanmycins may provide further insight into
biosynthetic mechanisms for uridyl peptide antibiotics.

<>

<1>Wang, N., Ng, I.S., Chen, P.T., Li, Y., Chen, Y.C., Chen, B.Y., Lu, Y.
<2>Draft Genome Sequence of the Bioelectricity-Generating and Dye-Decolorizing Bacterium Proteus hauseri Strain ZMd44.
<3>Genome Announcements
<4>2
<5>e00992-13
<6>2014
<7>Proteus hauseri ZMd44 (CGMCC 6746), as a crucial biodecolorizing, bioelectricity-generating,
and copper-resistant bacterium, is distinguished from
the urinary pathogens Proteus penneri and Proteus mirabilis. To further
investigate the genetic functions of this strain, the genome sequence and
annotation of its open reading frames, which consist of 3,875,927 bp (G+C
content, 38.12%), are presented here.

<>

<1>Wang, P., Brank, A.S., Banavali, N.K., Nicklaus, M.C., Marquez, V.E., Christman, J.K., MacKerell, A.D. Jr.
<2>Use of oligodeoxyribonucleotides with conformationally constrained abasic sugar targets to probe the mechanism of base flipping by HhaI DNA (cytosine C5)-methyltransferase.
<3>J. Am. Chem. Soc.
<4>122
<5>12422-12434
<6>2000
<7>X-ray crystallographic studies of HhaI DNA (cytosine-C5)-methyltransferase covalently linked
to methylated 5-fluorocytosine in DNA provided the first direct evidence that the cytosine
residue targeted for methylation was "flipped" out of the helix during the transfer reaction.
Subsequent studies indicated that removal of the target cytosine base, i.e., introduction of
an abasic site, enhanced binding of M.HhaI to DNA and that the conformation of the
sugar-phosphate backbone at the abasic site in the resultant complexes was the same as that of
the sugar attached to a "flipped" cytosine.  In the present study, pseudorotationally
constrained sugar analogues, based on bicyclo[3.1.0]hexane templates, were placed in DNA
duplexes as abasic target sites in the M.HhaI recognition sequence.  Biochemical studies
demonstrate that binding affinity of M.HhaI for abasic sites increases when the abasic target
sugar analogue is constrained to the south conformation and decreases when it is constrained
to the north conformation.  In native gel-shift assays, M.HhaI exhibits a "closed"
conformation when bound to the abasic south or abasic furanose analogues, whereas an "open"
conformation predominates with the abasic north analogue.  A structural understanding of these
results was obtained via molecular dynamics simulations of the DNA duplex alone and in ternary
complex with M.HhaI and cofactor, along with quantum mechanical calculations on model
compounds representative of the abasic and modified sugars.  Binding affinities are shown to
be related to the ability of the abasic sugar analogues to spontaneously flip out of the DNA
duplex.  Enhanced binding of the abasic south analogue is suggested to be due to its increased
capacity for sampling the experimentally observed conformation of the DNA target site in the
M.HhaI ternary complex.  Decreased binding of the north analogue is due to decreased
flexibility of the phosphodiester backbone associated with a north pseudorotation angle,
thereby inhibiting spontaneous flipping of the sugar moiety out of the DNA duplex.
Spontaneous flipping of the sugar moiety out of the DNA duplex is also suggested to facilitate
formation of a "closed" complex between M.HhaI and DNA whereas partial or no flipping favor
the "open" conformation.  These results show that introduction of structural constraints into
DNA that induce enhanced sampling of protein-bound conformations facilitate DNA-protein
binding.  Implications of the present results with respect to the mechanism of vase flipping
in the M.HhaI catalytic cycle are discussed.

<>

<1>Wang, P., Harvey, S.S., Sims, P.F.G., Broda, P.
<2>The construction of Streptomyces cyaneus genomic libraries in Escherichia coli is dependent upon the use of Mcr-deficient strains.
<3>Gene
<4>119
<5>127-129
<6>1992
<7>Streptomyces cyaneus genomic DNA ligated into either lambda phage or plasmid vectors was very
inefficiently cloned into standard Escherichia coli host strains. However, the same material
could be efficiently cloned using Mcr-deficient E. coli strains. These results suggest that
the S. cyaneus genome contains 5-methylcytosine residues, some of which occur with the
recognition sequences of the E. coli Mcr restriction system.

<>

<1>Wang, P., Li, L., Chen, X., Jiang, N., Liu, G., Chen, L., Xu, J., Song, H., Chen, Z., Ma, Y.
<2>Draft Genome Sequence of Alicyclobacillus hesperidum Strain URH17-3-68.
<3>J. Bacteriol.
<4>194
<5>6348
<6>2012
<7>Alicyclobacillus hesperidum is a thermoacidophilic bacterium. We isolated strain  URH17-3-68
from hot spring sludge in Tengchong, Yunnan province, China. Its
extracellular products include heat- and acid-stable enzymes which are important
for industrial applications. Here we report the draft genome of this strain.

<>

<1>Wang, Q., Shao, Z., Wang, X., Gao, Y., Li, M., Xu, L., Xu, J., Wang, L.
<2>Genetic Study of Capsular Switching between Neisseria meningitidis Sequence Type 7 Serogroup A and C Strains.
<3>Infect. Immun.
<4>78
<5>3883-3888
<6>2010
<7>Neisseria meningitidis is a leading cause of septicemia and meningitis
worldwide. N. meningitidis capsular polysaccharides have been classified
into 13 distinct serogroups which are defined by antibody reactivity and
structural analysis, and the capsule plays an important role in virulence.
Serogroups A, B, C, W135, and Y have been reported to be clinically
important. Several newly identified serogroup C isolates belonging to the
unique sequence type 7 (ST-7) were identified in China. Since most ST-7
isolates from China belonged to serogroup A, the newly identified ST-7
serogroup C strains were proposed to have arisen from those belonging to
ST-7 serogroup A. In this study, six ST-7 serogroup C and three ST-7
serogroup A isolates were analyzed by pulsed-field gel electrophoresis to
confirm their sequence type. In order to clarify the genetic basis of
capsular switching between ST-7 serogroup A and C strains, the whole
capsular gene clusters and surrounding genes of the two representative
ST-7 strains belonging to serogroups A and C, respectively, were sequenced
and compared. Potential recombination sites were analyzed using the RDP3
beta software, and recombination-related regions in two other ST-7
serogroup A and five ST-7 serogroup C strains were also sequenced and
compared to the representative ST-7 serogroup A and C strain sequences.

<>

<1>Wang, Q., Tsukahara, S., Yamakawa, H., Takai, K., Takaku, H.
<2>pH-independent inhibition of restriction endonuclease cleavage via triple helix formation by oligonucleotides containing 8-oxo-2'-deoxyadenosine.
<3>FEBS Lett.
<4>355
<5>11-14
<6>1994
<7>The ability of homopyrimidine oliogonucleotides containing 8-oxo-2'-deoxyadenosine to form
stable, triple helical structures with the sequence containing the recognition site for the
class II-S restriction enzyme, Ksp632I, was studied as a function of pH. The
8-oxo-2'-deoxyadenosine-substituted oligomers were shown to inhibit enzymatic cleavage and to
bind within the physiological pH range in a pH-independent fashion without compromising
specificity.

<>

<1>Wang, Q., Yang, M., Xiao, J., Wu, H., Wang, X., Lv, Y., Xu, L., Zheng, H., Wang, S., Zhao, G., Liu, Q., Zhang, Y.
<2>Genome sequence of the versatile fish pathogen Edwardsiella tarda provides insights into its adaptation to broad host ranges and intracellular niches.
<3>PLoS ONE
<4>4
<5>E7646
<6>2009
<7>BACKGROUND: Edwardsiella tarda is the etiologic agent of edwardsiellosis,
a devastating fish disease prevailing in worldwide aquaculture industries.
Here we describe the complete genome of E. tarda, EIB202, a highly
virulent and multi-drug resistant isolate in China. METHODOLOGY/PRINCIPAL
FINDINGS: E. tarda EIB202 possesses a single chromosome of 3,760,463 base
pairs containing 3,486 predicted protein coding sequences, 8 ribosomal
rRNA operons, and 95 tRNA genes, and a 43,703 bp conjugative plasmid
harboring multi-drug resistant determinants and encoding type IV A
secretion system components. We identified a full spectrum of genetic
properties related to its genome plasticity such as repeated sequences,
insertion sequences, phage-like proteins, integrases, recombinases and
genomic islands. In addition, analysis also indicated that a substantial
proportion of the E. tarda genome might be devoted to the growth and
survival under diverse conditions including intracellular niches, with a
large number of aerobic or anaerobic respiration-associated proteins,
signal transduction proteins as well as proteins involved in various
stress adaptations. A pool of genes for secretion systems, pili formation,
nonfimbrial adhesions, invasions and hemagglutinins, chondroitinases,
hemolysins, iron scavenging systems as well as the incomplete flagellar
biogenesis might feature its surface structures and pathogenesis in a fish
body. CONCLUSION/SIGNIFICANCE: Genomic analysis of the bacterium offered
insights into the phylogeny, metabolism, drug-resistance, stress
adaptation, and virulence characteristics of this versatile pathogen,
which constitutes an important first step in understanding the
pathogenesis of E. tarda to facilitate construction of a practical
effective vaccine used for combating fish edwardsiellosis.

<>

<1>Wang, Q.Y., Xie, N.Z., Huang, Y.Y., Song, L.F., Du QS, Yu.B., Chen, D., Huang, R.B.
<2>Genome Sequence of Tumebacillus flagellatus GST4, the First Genome Sequence of a  Species in the Genus Tumebacillus.
<3>Genome Announcements
<4>2
<5>e01189-14
<6>2014
<7>We present here the first genome sequence of a species in the genus Tumebacillus. The draft
genome sequence of Tumebacillus flagellatus GST4 provides a genetic
basis for future studies addressing the origins, evolution, and ecological role
of Tumebacillus organisms, as well as a source of acid-resistant amylase-encoding
genes for further studies.

<>

<1>Wang, R., Jin, Y., Norris, D.
<2>Identification of a protein that binds to the HO endonuclease recognition sequence at the yeast mating type locus.
<3>Mol. Cell. Biol.
<4>17
<5>770-777
<6>1997
<7>Mating type switching in Saccharomyces cerevisiae initiates when HO endonuclease makes a
site-specific double-stranded break at MAT, the yeast mating type locus.  To identify other
proteins involved in this process, we examined whether extracts prepared from ho- mutants
contain additional factors that bind near the recognition sequence for HO.  Using an
electrophoretic mobility shift assay, we isolated a chromatographic fraction that contains an
activity, named YZbp, which binds to two sequences flanking the recognition sequence at
MATalpha and to one sequence overlapping it at MATa.  MAT plasmids carrying mutations in the
YZbp recognition sequence are cleaved by purified Ho at wild-type efficiencies in an in vitro
assay.  These same plasmids, however, are not cleaved by Ho inside cells, demonstrating that
YZbp acts as a positive activator of in vivo cleavage.  YZbp is present in all cell types,
even those not undergoing mating type switching, suggesting that it has additional cellular
functions.

<>

<1>Wang, R.Y.-H., Shedlarski, J.G., Farber, M.B., Kuebbing, D., Ehrlich, M.
<2>Two sequence-specific endonucleases from Xanthomonas oryzae.  Characterization and unusual properties.
<3>Biochim. Biophys. Acta
<4>606
<5>371-385
<6>1980
<7>XorI and XorII, two sequence-specific endonucleases, have been partially
purified from Xanthomas oryzae.  XorI and XorII were shown to be isoschizomers
of PstI and PvuI, respectively.  X. oryzae is a particularly good source of
this PvuI isoschizomer because of the high yield of XorII, its simple
purification scheme, and its relative stability.  Furthermore, XorII was shown
to cleave at different positions in its recognition sequence than do at least
two of its known isoschizomers; XorII cleaves between the C and the G at the
3'-end of its palindromic recognition sequence, 5'-CGATC^G-3'.  There is a
single XorII site in each of the plasmid-cloning vehicles pBR313 and pBR322.
Two unusual aspects of XorII digestion are discussed, namely, the kinetics of
digestion of pBR313 and pBR322 and the resistance of human DNA to XorII.

<>

<1>Wang, R.Y.-H., Shenoy, S., Ehrlich, M.
<2>DNA methylation inhibits the transfecting activity of replicative-form PhiX174 DNA.
<3>J. Virol.
<4>49
<5>674-679
<6>1984
<7>Replacement of virtually all the cytosine residues with 5-methylcytosine residues in the
complementary strand of the replicative form (RF) of PhiX174 DNA caused a 300- to 500-fold
loss in its transfecting activity.  Similar results were obtained with analogously methylated
M13 RF.  Transfection experiments with PhiX RF hemimethylated in only part of the molecule, as
assessed by analysis with restriction endonucleases, indicated that gene A of PhiX, which
needs to be nicked at a specific site by the gene A protein for RF replication, was not the
main target for this inhibition by DNA methylation.  We propose that the loss of transfecting
activity was due to hemimethylation of the PhiX RF interfering with the processively catalyzed
movement of the replication fork.

<>

<1>Wang, R.Y.-H., Zhang, X.-Y., Ehrlich, M.
<2>A human DNA-binding protein is methylation-specific and sequence-specific.
<3>Nucleic Acids Res.
<4>14
<5>1599-1614
<6>1986
<7>A nuclear protein isolated from human placenta, methylated DNA-binding protein, binds
selectively to DNA enriched in 5-methylcytosine.  We now demonstrate that MDBP is a
sequence-specific, as well as methylation-specific, DNA-binding protein.  From restriction
fragments of pBR322 DNA methylated with human DNA methyltransferase, one was bound to mDBP
very much more strongly than any of the others.  For this preferential binding to MDBP, the
DNA had to be methylated.  By a DNase I protection experiment (DNase I footprinting), a
22-base sequence within this methylated restriction fragment was shown to be specifically
protected by MDP.  The sequence-specificity of MDBP coupled with its dependence on DNA
methylation suggests that this is one of the proteins which modulates important functions of
human DNA methylation in vivo.

<>

<1>Wang, R.Y.-H., Zhang, X.-Y., Khan, R., Zhou, Y., Huang, L.-H., Ehrlich, M.
<2>Methylated DNA-binding protein from human placenta recognizes specific methylated sites on several prokaryotic DNAs.
<3>Nucleic Acids Res.
<4>14
<5>9843-9860
<6>1986
<7>Methylated DNA-binding protein (MDBP) from human placenta recognizes specific
DNA sequences containing 5-methylcytosine (m5C) residues.  Comparisons of
binding of various prokaryotic DNAs to MDBP indicate that m5CpG is present in
the recognition sites for this protein but is only part of the recognition
sequence.  Specific binding to MDBP was observed for bacteriophage XP12 DNA,
which naturally contains ~1/3 of its residues as m5C, and for Micrococcus
luteus DNA, M13mp8 replicative form (RF) DNA, and pBR322 when these three DNAs
were methylated at CpG sites by human DNA methyltransferase.  Five DNA regions
binding to MDBP have been localized by DNase I footprinting or restriction
mapping in methylated pBR322 and M13mp8 RF DNAs.  A comparison of their
sequences reveals a common 5'-m5CGRm5CG-3' element or closely related sequence
in which one of the m5C residues may be replaced by a T.  In addition to this
motif, one upstreaem and one downstream m5CpG as well as other common residues
over an ~20-bp long region may be recognized by MDBP.

<>

<1>Wang, S., Chng, K.R., Wilm, A., Zhao, S., Yang, K.L., Nagarajan, N., He, J.
<2>Genomic characterization of three unique Dehalococcoides that respire on persistent polychlorinated biphenyls.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>111
<5>12103-12108
<6>2014
<7>Fastidious anaerobic bacteria play critical roles in environmental bioremediation of
halogenated compounds. However, their characterization and application have
been largely impeded by difficulties in growing them in pure culture. Thus far,
no pure culture has been reported to respire on the notorious polychlorinated
biphenyls (PCBs), and functional genes responsible for PCB detoxification remain
unknown due to the extremely slow growth of PCB-respiring bacteria. Here we
report the successful isolation and characterization of three Dehalococcoides
mccartyi strains that respire on commercial PCBs. Using high-throughput
metagenomic analysis, combined with traditional culture techniques,
tetrachloroethene (PCE) was identified as a feasible alternative to PCBs to
isolate PCB-respiring Dehalococcoides from PCB-enriched cultures. With PCE as an
alternative electron acceptor, the PCB-respiring Dehalococcoides were boosted to
a higher cell density (1.2 x 10(8) to 1.3 x 10(8) cells per mL on PCE vs. 5.9 x
10(6) to 10.4 x 10(6) cells per mL on PCBs) with a shorter culturing time (30 d
on PCE vs. 150 d on PCBs). The transcriptomic profiles illustrated that the
distinct PCB dechlorination profile of each strain was predominantly mediated by
a single, novel reductive dehalogenase (RDase) catalyzing chlorine removal from
both PCBs and PCE. The transcription levels of PCB-RDase genes are 5-60 times
higher than the genome-wide average. The cultivation of PCB-respiring
Dehalococcoides in pure culture and the identification of PCB-RDase genes deepen
our understanding of organohalide respiration of PCBs and shed light on in situ
PCB bioremediation.

<>

<1>Wang, S., Chng, K.R., Wu, C., Wilm, A., Nagarajan, N., He, J.
<2>Draft Genome Sequence of Polychlorinated Biphenyl-Dechlorinating Dehalococcoides  mccartyi Strain SG1, Which Carries a Circular Putative Plasmid.
<3>Genome Announcements
<4>2
<5>e00901-14
<6>2014
<7>Dehalococcoides mccartyi strain SG1, isolated from digester sludge, dechlorinates
polychlorinated biphenyls (PCBs) to lower congeners. Here we report the draft
genome sequence of SG1, which carries a 22.65 kbp circular putative plasmid.

<>

<1>Wang, S., Hao, B., Li, J., Gu, H., Peng, J., Xie, F., Zhao, X., Frech, C., Chen, N., Ma, B., Li, Y.
<2>Whole-genome sequencing of Mesorhizobium huakuii 7653R provides molecular insights into host specificity and symbiosis island dynamics.
<3>BMC Genomics
<4>15
<5>440
<6>2014
<7>BACKGROUND: Evidence based on genomic sequences is urgently needed to confirm the
phylogenetic relationship between Mesorhizobium strain MAFF303099 and M. huakuii.
To define underlying causes for the rather striking difference in host
specificity between M. huakuii strain 7653R and MAFF303099, several probable
determinants also require comparison at the genomic level. An improved
understanding of mobile genetic elements that can be integrated into the main
chromosomes of Mesorhizobium to form genomic islands would enrich our knowledge
of how genome dynamics may contribute to Mesorhizobium evolution in general.
RESULTS: In this study, we sequenced the complete genome of 7653R and compared it
with five other Mesorhizobium genomes. Genomes of 7653R and MAFF303099 were found
to share a large set of orthologs and, most importantly, a conserved chromosomal
backbone and even larger perfectly conserved synteny blocks. We also identified
candidate molecular differences responsible for the different host specificities
of these two strains. Finally, we reconstructed an ancestral Mesorhizobium
genomic island that has evolved into diverse forms in different Mesorhizobium
species. CONCLUSIONS: Our ortholog and synteny analyses firmly establish
MAFF303099 as a strain of M. huakuii. Differences in nodulation factors and
secretion systems T3SS, T4SS, and T6SS may be responsible for the unique host
specificities of 7653R and MAFF303099 strains. The plasmids of 7653R may have
arisen by excision of the original genomic island from the 7653R chromosome.

<>

<1>Wang, S., Zhu, H., He, F., Luo, Y., Kang, Z., Lu, C., Feng, L., Lu, X., Xue, Y., Wang, H.
<2>Whole Genome Sequence of the Probiotic Strain Lactobacillus paracasei N1115, Isolated from Traditional Chinese Fermented Milk.
<3>Genome Announcements
<4>2
<5>e00059-14
<6>2014
<7>Lactobacillus paracasei N1115 is a new strain with probiotic properties isolated  from
traditional homemade dairy products in Inner Mongolia, China. Here, we
report the complete genome sequence of L. paracasei N1115, which shows high
similarity to the well-studied probiotic Lactobacillus rhamnosus GG, and 3
structures turned out to be inversions, according to the colinearity analysis of
the BLAST alignment.

<>

<1>Wang, T.-S.
<2>PflI, a restriction endonuclease from Pseudomonas fluorescens Migula.
<3>K'o Hsueh T'ung Pao
<4>26
<5>815-817
<6>1981
<7>Over two hundred kinds of restriction endonucleases from 39 genuses, 94
species, and 159 strains have been found up to now according to Roberts'
statistics.  Many of them are isoschizomers.  Nothing about restriction
endonucleases from Psuedomonas fluorescens Migula has been published.  This is
a report on restriction endonuclease extracted from Pseudomonas fluorescens
Migula.

<>

<1>Wang, W., Ma, T., Ren, Y., Li, G.
<2>Draft Genome Sequence of a Benzothiophene-Desulfurizing Bacterium, Gordona terrae Strain C-6.
<3>Genome Announcements
<4>1
<5>e00381-13
<6>2013
<7>Gordona terrae strain C-6 was isolated from oil-contaminated soil and is capable  of
desulfurizing benzothiophene (BT). Here we report the draft genome sequence of
G. terrae strain C-6, which may help to reveal the genetic basis of the BT
biodesulfurization pathway.

<>

<1>Wang, X., Gao, Z., Xu, X., Ruan, L.
<2>Complete Genome Sequence of Thermococcus sp. Strain 4557, a Hyperthermophilic Archaeon Isolated from a Deep-Sea Hydrothermal Vent  Area.
<3>J. Bacteriol.
<4>193
<5>5544-5545
<6>2011
<7>Thermococcus sp. strain 4557 is a hyperthermophilic anaerobic archaeon isolated from the
deep-sea hydrothermal vent Guaymas Basin site in the
Gulf of California at a depth of 2,000 m. Here, we present the complete
genome sequence of Thermococcus sp. 4557, which consists of a single
circular chromosome of 2,011,320 bp with a G+C content of 56.08%.

<>

<1>Wang, X., Greenfield, P., Li, D., Hendry, P., Volk, H., Sutherland, T.D.
<2>Complete Genome Sequence of a Nonculturable Methanococcus maripaludis Strain Extracted in a Metagenomic Survey of Petroleum Reservoir Fluids.
<3>J. Bacteriol.
<4>193
<5>5595
<6>2011
<7>Extraction of genome sequences from metagenomic data is crucial for reconstructing the
metabolism of microbial communities that cannot be
mimicked in the laboratory. A complete Methanococcus maripaludis genome
was generated from metagenomic data derived from a thermophilic subsurface
oil reservoir. M. maripaludis is a hydrogenotrophic methanogenic species
that is common in mesophilic saline environments. Comparison of the genome
from the thermophilic, subsurface environment with the genome of the type
species will provide insight into the adaptation of a methanogenic genome
to an oil reservoir environment.

<>

<1>Wang, X., Hao, H., Xu, Z., Zheng, H., Liu, C., Wei, L., Zhang, R., Bi, D., Chen, H., Tan, C.
<2>Plasmid-mediated multidrug resistance and virulence in an avian pathogenic Escherichia coli strain isolated in China.
<3>J. Glob. Antimicrob. Resist.
<4>2
<5>57-58
<6>2014
<7>Avian pathogenic Escherichia coli is the causative agent of avian colisepticaemia and has been
previously reported as a potential zoonotic risk.  APEC strains often display a multidrug
restance pattern including resistance to aminoglycosides, B-lactams, chloramphenicol,
fluoroquinolones, tetracycles and trimethoprim/sulfamethoxazole.  In this study, an APEC
strain (ACN001) isolated from the liver of a diseased chicken in China in 2010 was
characterised.  ACN001 was shown to belong to phylogenetic group B2 and was confirmed to be
highly virulent in chicken and mouse models using previously described methods.  Antimicrobial
susceptibility testing performed according to Clinical and Laboratory Standards Institute
standards revealed that the strain was multidrug-resistant, with high minimum inhibitory
concentrations to amikacin (>16 ug/mL), ampicillin (256 ug/mL), chlooramphenical (256 ug/mL),
ciprofloxacin (64 ug/mL), florfenicol (256 ug/mL), gentamicin (256 ug/mL), norfloxacin (64
ug/mL), streptomycin (256 ug/mL) and tetracycline (256 ug/mL).

<>

<1>Wang, X., Jin, D., Zhou, L., Wu, L., An, W., Chen, Y., Zhao, L.
<2>Draft Genome Sequence of Brevibacillus panacihumi Strain W25, a Halotolerant Hydrocarbon-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e01215-13
<6>2014
<7>Brevibacillus panacihumi strain W25 was isolated from hydrocarbon-contaminated saline soil.
Here, we report the 5.5-Mb draft genome sequence of this strain,
which may provide insights into the mechanism of microbial hydrocarbon
degradation in saline environments.

<>

<1>Wang, X., Jin, D., Zhou, L., Wu, L., An, W., Zhao, L.
<2>Draft Genome Sequence of Gordonia alkanivorans Strain CGMCC6845, a Halotolerant Hydrocarbon-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e01274-13
<6>2014
<7>Gordonia alkanivorans strain CGMCC6845 is a halotolerant hydrocarbon-degrading bacterium
isolated from petroleum-contaminated saline soil. Here we present the
5.0-Mb draft genome sequence of this strain, which will improve our understanding
of the diversity of G. alkanivorans and the mechanisms of microbial hydrocarbon
degradation in saline environment.

<>

<1>Wang, X., Jin, D., Zhou, L., Wu, L., An, W., Zhao, L.
<2>Draft Genome Sequence of Advenella kashmirensis Strain W13003, a Polycyclic Aromatic Hydrocarbon-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e00003-14
<6>2014
<7>Advenella kashmirensis strain W13003 is a polycyclic aromatic hydrocarbon (PAH)-degrading
bacterium isolated from PAH-contaminated marine sediments. Here,
we report the 4.8-Mb draft genome sequence of this strain, which will provide
insights into the diversity of A. kashmirensis and the mechanism of PAH
degradation in the marine environment.

<>

<1>Wang, X., Jin, D., Zhou, L., Wu, L., Qi, L., Li, C., An, W., Chen, Y.
<2>Draft Genome Sequence of Halotolerant Polycyclic Aromatic Hydrocarbon-Degrading Pseudomonas bauzanensis Strain W13Z2.
<3>Genome Announcements
<4>2
<5>e01049-14
<6>2014
<7>Pseudomonas bauzanensis W13Z2 is a halotolerant polycyclic aromatic hydrocarbon
(PAH)-degrading bacterium isolated from petroleum-contaminated drill cuttings in
the Bohai Sea. Here, we report the 8.6-Mb draft genome sequence of this strain,
which will provide insights into the diversity of Pseudomonas and the mechanism
of PAHs degradation in drill cuttings.

<>

<1>Wang, X., Li, Y., Jing, H., Ren, Y., Zhou, Z., Wang, S., Kan, B., Xu, J., Wang, L.
<2>Complete Genome Sequence of a Yersinia enterocolitica 'Old World' (3/O:9) Strain and Comparison with the 'New World' (1B/O:8) Strain.
<3>J. Clin. Microbiol.
<4>49
<5>1251-1259
<6>2011
<7>Yersinia enterocolitica is a heterogeneous bacterial species with a wide
range of animal reservoirs through which human intestinal illness can be
facilitated. In contrast to the epidemiological pattern observed in the
United States, infections in China present a pattern similar to those in
European countries and Japan, wherein "Old World" strains (biotypes 2-5)
are prevalent. To gain insights into the evolution of Y. enterocolitica
and pathogenic properties toward human hosts, we sequenced the genome of a
biotype 3 strain 105.5R(r) (O:9) obtained from a Chinese patient.
Comparative genome sequence analysis with strain 8081 (1B/O:8) revealed
new insights into Y. enterocolitica. Both strains have more than 14%
specific genes. In strain 105.5R(r), putative virulence factors were found
in strain-specific genomic pathogenicity islands that comprised a novel
type III secretion system and rtx-like genes. Many of the loci
representing ancestral clusters, which are believed to contribute to
enteric survival and pathogenesis, are present in strain 105.5R(r) but
lost in strain 8081. Insertion elements in 105.5R(r) have a distinct
pattern from those in strain 8081, and were exclusively located in a
strain-specific region. In summary, our comparative genome analysis
indicates that these two strains may have attained their pathogenicity by
completely separate evolutionary events, and the 105.5R(r) strain, a
representative of the "Old World" biogroup, lies in a branch of Y.
enterocolitica that is distinct from the "New World" 8081 strain.

<>

<1>Wang, X., Luo, C., Chen, Z.
<2>Genome Sequence of the Plant Growth-Promoting Rhizobacterium Bacillus sp. Strain  916.
<3>J. Bacteriol.
<4>194
<5>5467-5468
<6>2012
<7>Bacillus sp. strain 916, isolated from the soil, showed strong activity against Rhizoctonia
solani. Here, we present the high-quality draft genome sequence of
Bacillus sp. strain 916. Its 3.9-Mb genome reveals a number of genes whose
products are possibly involved in promotion of plant growth or antibiosis.

<>

<1>Wang, X., Tao, F., Gai, Z., Tang, H., Xu, P.
<2>Genome Sequence of the Welan Gum-Producing Strain Sphingomonas sp. ATCC 31555.
<3>J. Bacteriol.
<4>194
<5>5989-5990
<6>2012
<7>Sphingomonas sp. strain ATCC 31555 can produce an anionic heteropolysaccharide, welan gum,
which shows excellent stability and viscosity retention even at high
temperatures. Here we present a 4.0-Mb assembly of its genome sequence. We have
annotated 10 coding sequences (CDSs) responsible for the welan gum biosynthesis
and 55 CDSs related to monosaccharide metabolism.

<>

<1>Wang, X., Wang, Q., Zhang, W., Wang, Y., Li, L., Wen, T., Zhang, T., Zhang, Y., Xu, J., Hu, J., Li, S., Liu, L., Liu, J., Jiang, W., Tian, J., Li, Y., Schuler, D., Wang, L., Li, J.
<2>Complete Genome Sequence of Magnetospirillum gryphiswaldense MSR-1.
<3>Genome Announcements
<4>2
<5>e00171-14
<6>2014
<7>We report the complete genomic sequence of Magnetospirillum gryphiswaldense MSR-1 (DSM 6361),
a type strain of the genus Magnetospirillum belonging to the
Alphaproteobacteria. Compared to the reported draft sequence, extensive
rearrangements and differences were found, indicating high genomic flexibility
and 'domestication' by accelerated evolution of the strain upon repeated
passaging.

<>

<1>Wang, X., Zhang, Z., Hao, Q., Wu, J., Xiao, J., Jing, H.
<2>Complete Genome Sequence of Acinetobacter baumannii ZW85-1.
<3>Genome Announcements
<4>2
<5>e01083-13
<6>2014
<7>Acinetobacter baumannii is an aerobic, nonmotile Gram-negative bacterium that causes
nosocomial infections worldwide. Here, we report the complete genome
sequence of Acinetobacter baumannii strain ZW85-1 and its two plasmids. One of
the plasmids carries genes for NDM-1, which can hydrolyze a wide range of
antibiotics.

<>

<1>Wang, X., Zhang, Z., Jin, D., Zhou, L., Wu, L., Li, C., Zhao, L., An, W., Chen, Y.
<2>Draft Genome Sequence of Brachybacterium phenoliresistens Strain W13A50, a Halotolerant Hydrocarbon-Degrading Bacterium.
<3>Genome Announcements
<4>2
<5>e00899-14
<6>2014
<7>Brachybacterium phenoliresistens strain W13A50 was isolated from a petroleum-contaminated
saline site, which could degrade hydrocarbon under high
salinity conditions. Here, we present 4.2-Mb draft genome sequence of this
strain, which will provide insights into the diversity of Brachybacterium and the
mechanism of hydrocarbon degradation in saline environments.

<>

<1>Wang, X., Zhu, D., Wang, M., Cheng, A., Jia, R., Zhou, Y., Chen, Z., Luo, Q., Liu, F., Wang, Y., Chen, X.Y.
<2>Complete Genome Sequence of Riemerella anatipestifer Reference Strain.
<3>J. Bacteriol.
<4>194
<5>3270-3271
<6>2012
<7>Riemerella anatipestifer is an infectious pathogen causing serositis in ducks. We had the
genome of the R. anatipestifer reference strain ATCC 11845 sequenced. The
completed draft genome consists of one circular chromosome with 2,164,087 bp.
There are 2,101 genes in the draft, and its GC content is 35.01%.

<>

<1>Wang, X.J., Yan, Y.J., Zhang, B., An, J., Wang, J.J., Tian, J., Jiang, L., Chen, Y.H., Huang, S.X., Yin, M., Zhang, J., Gao, A.L., Liu, C.X., Zhu, Z.X., Xiang, W.S.
<2>Genome Sequence of the Milbemycins-producing Bacterium Streptomyces bingchenggensis.
<3>J. Bacteriol.
<4>192
<5>4526-4527
<6>2010
<7>Streptomyces bingchenggensis is a soil dwelling bacterium producing the commercially important
anthelmintic macrolide milbemycins. Besides
milbemycins, the insecticidal polyether antibiotic nanchangmycin and some
other antibiotics have also been isolated from this strain. Here we report
the complete genome sequence of S. bingchenggensis. The availability of
the genome sequence of S. bingchenggensis should enable us to understand
the biosynthesis of these structurally intricate antibiotics better and
facilitate rational improvement of this strain to increase their titers.

<>

<1>Wang, X.Q., Showmaker, K.C., Yu, X.Q., Bi, T., Hsu, C.Y., Baird, S.M., Peterson, D.G., Li, X.D., Lu, S.E.
<2>Draft Genome Sequence of Burkholderia pyrrocinia Lyc2, a Biological Control Strain That Can Suppress Multiple Plant Microbial Pathogens.
<3>Genome Announcements
<4>2
<5>e00991-14
<6>2014
<7>Burkholderia pyrrocinia strain Lyc2 was isolated from the tobacco rhizosphere in  China. This
bacterium exhibits a remarkable capacity to inhibit the growth of
multiple pathogens and shows strong suppression of cotton seedling damping-off.
Here, we present the draft genome sequence of Burkholderia pyrrocinia strain
Lyc2.

<>

<1>Wang, Y., Chen, C., Ai, L., Zhou, F., Zhou, Z., Wang, L., Zhang, H., Chen, W., Guo, B.
<2>Complete genome sequence of probiotic Lactobacillus plantarum ST-III.
<3>J. Bacteriol.
<4>193
<5>313-314
<6>2010
<7>Lactobacillus plantarum strain ST-III, a probiotic strain with several functions, was isolated
from kimchi. Here we report the complete genome sequence of ST-III and compared it with two
published L. plantarum genomes.

<>

<1>Wang, Y., Du Y, Y.Y., Guo, Y., Wang, J., Li, T., Chang, D., Liu, Y., Jiang, X., Dai, W., Liu, C.
<2>Draft Genome Sequence of Serratia marcescens Strain LCT-SM166, a Space Flight Strain with a Specific Carbon Source Utilization Pattern.
<3>Genome Announcements
<4>2
<5>e00069-14
<6>2014
<7>Serratia marcescens has been detected in space habitats. To explore the influence of the space
flight environment on this bacterium, we investigated the genome
sequence of LCT-SM166, which was isolated after space flight and has a specific
carbon source utilization pattern.

<>

<1>Wang, Y., Jorda, M., Jones, P.L., Maleszka, R., Ling, X., Robertson, H.M., Mizzen, C.A., Peinado, M.A., Robinson, G.E.
<2>Functional CpG methylation system in a social insect.
<3>Science
<4>314
<5>645-647
<6>2006
<7>DNA methylation systems are well characterized in vertebrates, but methylation in Drosophila
melanogaster and other invertebrates remains
controversial. Using the recently sequenced honey bee genome, we present a
bioinformatic, molecular, and biochemical characterization of a functional
DNA methylation system in an insect. We report on catalytically active
orthologs of the vertebrate DNA methyltransferases Dnmt1 and Dnmt3a and b,
two isoforms that contain a methyl-DNA binding domain, genomic
5-methyl-deoxycytosine, and CpG-methylated genes. The honey bee provides
an opportunity to study the roles of methylation in social contexts.

<>

<1>Wang, Y., Ke, Y., Gao, G., Zhen, Q., Yuan, X., Wang, Z., Xu, J., Li, T., Wang, D., Huang, L., Xu, X., Chen, Z.
<2>Complete Genome Sequence of Brucella abortus 134, a Biovar 1 Strain Isolated from Human.
<3>J. Bacteriol.
<4>194
<5>6658
<6>2012
<7>Brucella abortus is one of the common pathogens causing brucellosis in China. Here, we report
the genome sequence of B. abortus strain 134, a strain isolated
from a human patient and belonging to biovar 1, the most highly represented
biovar among B. abortus strains in China.

<>

<1>Wang, Y., Ke, Y., Zhen, Q., Yuan, X., Xu, J., Qiu, Y., Wang, Z., Li, T., Wang, D., Huang, L., Chen, Z.
<2>Complete Genome Sequence of Brucella canis BCB018, a Strain Isolated from a Human Patient.
<3>J. Bacteriol.
<4>194
<5>6697-6698
<6>2012
<7>Brucella canis is considered a rare cause of human brucellosis because of difficulties in
presumptive diagnosis and underestimation of the incidence. Here,
we report the draft genome sequence of a Brucella canis isolate, BCB018, isolated
from a human patient, providing precious resources for comparative genomics
analysis of Brucella field strains.

<>

<1>Wang, Y., Li, C., Gao, C., Ma, C., Xu, P.
<2>Genome Sequence of the Nonpathogenic Pseudomonas aeruginosa Strain ATCC 15442.
<3>Genome Announcements
<4>2
<5>e00421-14
<6>2014
<7>Pseudomonas aeruginosa ATCC 15442 is an environmental strain of the Pseudomonas genus. Here,
we present a 6.77-Mb assembly of its genome sequence. Besides giving
insights into characteristics associated with the pathogenicity of P. aeruginosa,
such as virulence, drug resistance, and biofilm formation, the genome sequence
may provide some information related to biotechnological utilization of the
strain.

<>

<1>Wang, Y., Ran, S., Yang, G.
<2>Single molecular investigation of DNA looping and aggregation by restriction endonuclease BspMI.
<3>Sci. Rep.
<4>4
<5>5897
<6>2014
<7>DNA looping and aggregation induced by restriction endonuclease BspMI are studied by atomic
force microscopy (AFM) and magnetic tweezers (MT). With Ca2+ substituted for the normal enzyme
cofactor Mg2+ and enzyme concentration below the critical concentration of 6 units/mL, AFM
images of DNA-BspMI complex show that the number of binding and looping events increases with
enzyme concentration. At the critical concentration 6 of units/mL, all the BspMI binding sites
are saturated. It is worth noting that nonspecific BspMI binding to DNA at saturation
concentration represents more than 8% of the total BspMI-DNA complexes directly observed in
AFM images. Furthermore, we used MT to prove that additional loops can form when enzyme
concentration is higher than its saturation valueand the complex is incubated for a long time
(>2 hrs). We ascribe this phenomenon to the aggregation of enzymes. The force spectroscopy of
the BspMI-DNA complex shows that the pulling force required to open the loop of the complex at
less than saturation concentration has a peak at about 3 pN, which is lower than the force
required to open additional loops due to enzyme aggregation at higher than saturation
concentration (>6 pN).

<>

<1>Wang, Y., Rocha, E.P.C., Leung, F.C.C., Danchin, A.
<2>Cytosine methylation is not the major factor inducing CpG dinucleotide deficiency in bacterial genomes.
<3>J. Mol. Evol.
<4>58
<5>692-700
<6>2004
<7>CpG dinucleotide deficiency has been found in viruses, mitochondria, prokaryotes, and
eukaryotes. The consensual explanation is that it is
due to deamination of methylated cytosines, as established for
vertebrate and plants. However, we still do not know whether C5
cytosine methylation is also the major cause of CpG deficiency in
bacteria. By combining annotation and experimental data identifying the
presence of C5 cytosine methyltransferases with analysis of CpG
relative abundance in 67 bacterial species, we found that CpG relative
abundance in most bacterial genomes that have cytosine C5
methyltransferases tends to be in the normal range (observed/expected
values between 0.82 and 1.21). In contrast, many bacterial species
likely to be lacking C5 cytosine methylation showed CpG deficiency.
Furthermore, when comparing genomes with one another, TpG and CpA
relative abundances were found to be independent from CpG relative
abundance. This contrasted with intragenome analyses, where C(3)pG(1)
relative abundance (the subscripts refer to position of a nucleotide in
a codon) was found to be generally positively correlated with T(3)pG(1)
relative abundances when plotted against GC content in protein coding
sequences (CDSs). This suggests the existence of alternative mechanisms
contributing to CpG deficiency in bacteria.

<>

<1>Wang, Y., Roos, K.P., Taylor, D.E.
<2>Transformation of Helicobacter pylori  by chromosomal metronidazole resistance and by a plasmid with a selectable chloramphenicol resistance marker.
<3>J. Gen. Microbiol.
<4>139
<5>2485-2493
<6>1993
<7>Most strains of Helicobacter pylori are naturally competent for uptake of chromosomal DNA.
Transformation frequencies for streptomycin resistance or rifampicin resistance markers ranged
from 1 x 10^-4 to 1 x 10^-3 per viable cell using a plate transformation procedure.
Transformation of a metronidazole resistance marker was demonstrated when either a
laboratory-derived mutant or an MtrR clinical isolate were used as the source of donor DNA.
MtrR was transformed at a frequency of 3 x 10^-5 per viable cell.  All H. pylori strains
tested produce large amounts of DNAase, which may reduce DNA available for transformation.
Four H. pylori plasmids were isolated.  DNA fragments from H. pylori plasmids were deleted or
rearranged when cloned in pUC19 and propagated in Escherichia coli DH5a.  An H. pylori
plasmid, pUOA26 which contained a chloramphenicol resistance determinant from Campylobacter
coli, was constructed in H. pylori.  This plasmid could be successfully introduced by natural
transformation only into H. pylori recipients which contained a homologous resident plasmid.
Transformation of pUOA26 into plasmid-free cells of H. pylori was achieved by electroporation.
Transformation frequencies were 1 x 10^-4 transformants per viable cell when plasmid DNA was
isolated from the same strain; however, introduction of pUOA26 DNA derived from H. pylori 8091
into a different H. pylori strain, NCTC 11639, resulted in transformation at much lower
frequencies (less than or equal to 1 x 10-7 per viable cell). Changes in plasmid restriction
patterns were also noted when pUOA26 was isolated from H. pylori NCTC 11639, suggesting the
presence of at least two different DNA restriction and modification systems in H. pylori which
may interfere with uptake of plasmid DNA.

<>

<1>Wang, Y., Tao, F., Li, C., Li, L., Xu, P.
<2>Genome Sequence of Klebsiella pneumoniae Strain ATCC 25955, an Oxygen-Insensitive Producer of 1,3-Propanediol.
<3>Genome Announcements
<4>1
<5>e00587-13
<6>2013
<7>Klebsiella pneumoniae strain ATCC 25955 is a 1,3-propanediol-producing bacterium  that is
insensitive to oxygen. Here, we present a 5.29-Mb assembly of its genome
sequence. We have annotated 10 coding sequences (CDSs) for 1,3-propanediol
fermentation and 18 CDSs for glycerol uptake. The CDSs related to virulence and
by-product formation were also annotated.

<>

<1>Wang, Y., Tao, F., Tang, H., Xu, P.
<2>Genome Sequence of Clostridium diolis Strain DSM 15410, a Promising Natural Producer of 1,3-Propanediol.
<3>Genome Announcements
<4>1
<5>e00542-13
<6>2013
<7>Clostridium diolis strain DSM 15410 is considered one of the best natural producers of
1,3-propanediol because of its appreciable substrate-tolerant
ability, yield, and productivity. Here, we present a 5.85-Mb assembly of its
genome sequence. We have annotated the coding sequences responsible for glycerol
utilization and 1,3-propanediol fermentation.

<>

<1>Wang, Y., Wang, J., Ahmed, Z., Bai, X., Wang, J.
<2>Complete Genome Sequence of Lactobacillus kefiranofaciens ZW3.
<3>J. Bacteriol.
<4>193
<5>4280-4281
<6>2011
<7>Lactobacillus kefiranofaciens ZW3 was isolated in Tibet, China, from kefir grain, a
traditional dairy product that is known to provide many health
benefits to humans. Here, we present the genome features of L.
kefiranofaciens ZW3 and the identification of a gene cluster related to
the synthesis of exopolysaccharide, an important constituent of the
Tibetan kefir.

<>

<1>Wang, Y., Wang, X., Greenfield, P., Jin, D., Bai, Z.
<2>Draft Genome Sequence of Bacillus amyloliquefaciens EBL11, a New Strain of Plant  Growth-Promoting Bacterium Isolated from Rice Rhizosphere.
<3>Genome Announcements
<4>2
<5>e00732-14
<6>2014
<7>Bacillus amyloliquefaciens strain EBL11 is a bacterium that can promote plant growth by
inhibiting the growth of fungi on plant surfaces and providing
nutrients as a nonchemical biofertilizer. The estimated genome of this strain is
4.05 Mb in size and harbors 3,683 coding genes (CDSs).

<>

<1>Wang, Y., Yuan, Y., Zhou, L., Su, Q., Fang, X., Li, T., Wang, J., Chang, D., Su, L., Xu, G., Guo, Y., Yang, R., Liu, C.
<2>Draft Genome Sequence of Serratia marcescens Strain LCT-SM213.
<3>J. Bacteriol.
<4>194
<5>4477-4478
<6>2012
<7>Serratia marcescens is a species of Gram-negative, rod-shaped bacterium of the family
Enterobacteriaceae. S. marcescens can cause nosocomial infections,
particularly catheter-associated bacteremia, urinary tract infections, and wound
infections. Here, we present the draft genome sequence of Serratia marcescens
strain LCT-SM213, which was isolated from CGMCC 1.1857.

<>

<1>Wang, Y., Zhang, R., Zheng, Q., Jiao, N.
<2>Draft Genome Sequences of Two Marine Phototrophic Bacteria, Erythrobacter longus  Strain DSM 6997 and Erythrobacter litoralis Strain DSM 8509.
<3>Genome Announcements
<4>2
<5>e00677-14
<6>2014
<7>Aerobic anoxygenic phototrophic bacteria (AAPB) are important functional groups and are widely
distributed in the global upper ocean. Here we report the draft
genomic sequences of two marine AAPB isolates belonging to the genus
Erythrobacter, Erythrobacter longus strain DSM 6997 and Erythrobacter litoralis
strain DSM 8509.

<>

<1>Wang, Y., Zheng, Y., Wang, M., Gao, Y., Xiao, Y., Peng, H.
<2>Non-contiguous finished genome sequence of Anoxybacillus flavithermus subsp. yunnanensis type strain (E13(T)), a strictly thermophilic and organic  solvent-tolerant bacterium.
<3>Standards in Genomic Sciences
<4>9
<5>735-743
<6>2014
<7>Anoxybacillus flavithermus subsp. yunnanensis is the only strictly thermophilic bacterium that
is able to tolerate a broad range of toxic solvents at its optimal
temperature of 55-60 degrees C. The type strain E13(T) was isolated from
water-sediment slurries collected from a hot spring. This study presents the
draft genome sequence of A. flavithermus subsp. yunnanensis E13(T) and its
annotation. The 2,838,393bp long genome (67 contigs) contains 3,035
protein-coding genes and 85 RNA genes, including 10 rRNA genes, and no plasmids.
The genome information has been used to compare with the genomes from A.
flavithermus subsp. flavithermus strains.

<>

<1>Wang, Y.H., He, X.X., Wang, K.M., Su, J., Chen, Z.F., Yan, G.P., Du, Y.D.
<2>A label-free electrochemical assay for methyltransferase activity detection based on the controllable assembly of single wall carbon nanotubes.
<3>Biosensors and Bioelectronics
<4>41
<5>238-243
<6>2013
<7>A sensitive label-free 'signal-on' electrochemical approach for detection of
methyltransferases (MTase) activity is developed based on
the signal transduction and amplification of single wall carbon
nanotubes (SWCNTs). In this method, the oligonucleotide I is first
self-assembled on the electrode via Au-S bonding. After hybridization
with its complement ssDNA (oligonucleotide II), duplex strand DNA
(dsDNA) probes containing specific recognition sequence of Dam MTase
and methylation-sensitive restriction endonuclease Dpn I is then formed
on the electrode. In the presence of Dam MTase and Dpn I, the dsDNA
probes are methylated and subsequently cleaved into two dsDNA
fragments. After heating, the remained dsDNA fragments on the electrode
melted into ssDNA fragments. Then the SWCNTs can be controllably
assembled on the ssDNA fragments remained on the electrode, mediating
efficient electron transfer between the electrode and electroactive
species. It generates measurable current signal (eT ON), which is
related to the concentration of the Dam MTase. The resulting change in
electron transfer efficiency is readily measured by differential pulse
voltammetry at Dam MTase concentrations as low as 0.04 U/mL. This
method does not need electroactive molecules labeling on the
methylation-responsive DNA probes. The linear response of the developed
facile signal-on electrochemical sensing system for Dam MTase is in the
range of 0.1-1.0 U/mL. In addition, such a SWCNTs based electrochemical
assay also has the ability to screen inhibitors for Dam MTase.

<>

<1>Wang, Y.P., Jarjour, J., Boissel, S., Thyme, S., Khan, I., Pangallo, J., Baker, D., Stoddard, B., Scharenberg, A.M., Rawlings, D.J.
<2>Engineering an XID-Site Specific I-AniI Homing Endonuclease through Combination of Rational Design and Directed Evolution.
<3>Mol. Ther.
<4>21
<5>S122-S123
<6>2013
<7>LAGLIDADG homing endonucleases (LHEs) are highly specific compact endonucleases with typical
20 base pair recognition sites, and thus are ideal scaffolds for engineering site-specific
enzymes for genome editing applications.  Here, we describe a progressive approach to LHE
engineering that combines rational design with directed evolution using yeast surface
display-based high throughput cleavage selection.  This approach was employed to alter the
binding and cleavage specificity of the I-Anil LHE to recognize a specific, unique mutation in
the mouse Bruton tyrosine kinase (Btk) gene - this mutation is known to cause mouse X-linked
immunodeficiency (XID) - a model of human X-linked agammaglobulimemia (XLA).  The required
retargeting of I-AniI involved resculpting of the DNA contact interface to accommodate nine
base differences from the native sequence.  As a first step, multiple site-specific variants
were engineered through randomization of amino acid residues contacting clusters of base pairs
altered in the partial targets.  As directly combining these "cluster target" LHE variants
failed to generate active enzymes cleaving the combined targets, variants with successful
re-specification of residue clusters proximal to the active site were subjected to a
progressive redesign strategy in which specificity and activity were extended outward across
the N-terminal and C-terminal XID half-site interfaces.  Successful "half-site" redesigns were
then combined to generate an active enzyme recognizing the full XID target site, and this
enzyme was refined by random mutatgenesis to create an enzyme with wildtype-level activity in
vitro and in cellulo reporter assays.  Finally, the in cellulo activity of this enzyme was
further increased by fusing to Transcription Activator-Like Effector (TALE) site-specific DNA
binding domain to copensate the partially reduced binding affinity of engineered XID enzyme.
This study not only provides a valuable roadmap for further LHE engineering by effectively
combining rationale and directed evolution strategies, but also demonstrates a novel means to
increase the binding affinity, thus the cleavage efficacy of engineered LHEs.

<>

<1>Wang, Z., Chang, X., Yang, X., Pan, L., Dai, J.
<2>Draft Genome Sequence of Polaromonas glacialis Strain R3-9, a Psychrotolerant Bacterium Isolated from Arctic Glacial Foreland.
<3>Genome Announcements
<4>2
<5>e00695-14
<6>2014
<7>Here we report the draft genome sequence of the psychrotolerant Polaromonas glacialis strain
R3-9, isolated from Midtre Lovenbreen glacial foreland near
Ny-Alesund, Svalbard Archipelago, Norway.

<>

<1>Wang, Z., Kadouri, D., Wu, M.
<2>Genomic Insights into An Obligate Epibiotic Bacterial Predator: Micavibrio aeruginosavorus ARL-13.
<3>BMC Genomics
<4>12
<5>453
<6>2011
<7>Background: Although bacterial predators play important roles in the dynamics of natural
microbial communities,
little is known about the molecular mechanism of bacterial predation and the evolution of
diverse predatory
lifestyles.
Results: We determined the complete genome sequence of Micavibrio aeruginosavorus ARL-13, an
obligate
bacterial predator that feeds by "leeching" externally to its prey. Despite being an obligate
predator depending on
prey for replication, M. aeruginosavorus encodes almost all major metabolic pathways. However,
our genome
analysis suggests that there are multiple amino acids that it can neither make nor import
directly from the
environment, thus providing a simple explanation for its strict dependence on prey.
Remarkably, despite apparent
genome reduction, there is a massive expansion of genomic islands of foreign origin. At least
nine genomic islands
encode many genes that are likely important for Micavibrio-prey interaction such as
hemolysin-related proteins.
RNA-Seq analysis shows substantial transcriptome differences between the attack phase, when M.
aeruginosavorus
seeks its prey, and the attachment phase, when it feeds and multiplies. Housekeeping genes as
well as genes
involved in protein secretion were all dramatically up-regulated in the attachment phase. In
contrast, genes
involved in chemotaxis and flagellum biosynthesis were highly expressed in the attack phase
but were shut down
in the attachment phase. Our transcriptomic analysis identified additional genes likely
important in Micavibrio
predation, including porins, pilins and many hypothetical genes.
Conclusions: The findings from our phylogenomic and transcriptomic analyses shed new light on
the biology and
evolution of the epibiotic predatory lifestyle of M. aeruginosavorus. The analysis reported
here and the availability of
the complete genome sequence should catalyze future studies of this organism.

<>

<1>Wang, Z., Lin, B., Hervey, W.J.I.V., Vora, G.J.
<2>Draft Genome Sequence of the Fast-Growing Marine Bacterium Vibrio natriegens Strain ATCC 14048.
<3>Genome Announcements
<4>1
<5>e00589-13
<6>2013
<7>Vibrio natriegens bacteria are Gram-negative aquatic microorganisms that are found primarily
in coastal seawater and sediments and are perhaps best known for
their high growth rates (generation time of <10 min). In this study, we report
the first sequenced genome of this species, that of the type strain Vibrio
natriegens ATCC 14048, a salt marsh mud isolate from Sapelo Island, GA.

<>

<1>Wang, Z., Wu, M.
<2>Complete Genome Sequence of the Endosymbiont of Acanthamoeba Strain UWC8, an Amoeba Endosymbiont Belonging to the 'Candidatus Midichloriaceae' Family in  Rickettsiales.
<3>Genome Announcements
<4>2
<5>e00791-14
<6>2014
<7>The endosymbiont of Acanthamoeba strain UWC8 is an obligate amoeba endosymbiont belonging to
the family of 'Candidatus Midichloriaceae' in Rickettsiales. We
report here the complete genome sequence of this bacterium, which should catalyze
future studies of amoeba-symbiont interactions.

<>

<1>Wang, Z.-G., Wu, J.-X.
<2>DNA methyltransferases: classification, functions and research progress.
<3>Yichuan
<4>31
<5>903-912
<6>2009
<7>DNA methylation is a postreplicative modification occurred in most prokaryotic and eukaryotic
genomes, which has a variety of important biological functions including regulation of gene
expression, gene imprinting, preservation of chromosomal integrity, and X-chromosome
inactivation. According to their structure and functions, DNA methyltransferases (Dnmts) are
divided into two major families in mammalian cells: maintenance methyltransferase (Dnmt1) and
de novo methyltransferases (Drmt3a, Dnmt3b, and Dnmt3L). In addition, Dnmt2 also displays weak
DNA methyltransferase catalytic activity, but newly founded function is to methylate cytosine
38 in the anti-codon loop of tRNA A,p. These Dnmts are crucial for mammalian growth and
development. Dnmts deficiency will lead to embryonic development defects, cancer, and other
diseases. Therefore, Dnmts could be important therapeutical targets. This article Summarizes
the classification, function, and recent research progress in DNA methyltransferases.

<>

<1>Wang, Z.G., Wu, Z., Xu, S.L., Zha, J.
<2>Genome Sequence of the Human-Pathogenetic Bacterium Vibrio vulnificus B2.
<3>J. Bacteriol.
<4>194
<5>7019
<6>2012
<7>Vibrio vulnificus, which can lead to rapidly expanding cellulitis or septicemia,  is present
in the marine environment. Here, we present the draft genome sequence
of strain B2, which was isolated from a septicemia patient in 2010.

<>

<1>Wani, A.A., Stephens, R.E., D'Ambrosio, S.M., Hart, R.W.
<2>A new sequence-specific endonuclease from Micrococcus radiodurans.
<3>Fed. Proc.
<4>40
<5>1848
<6>1981
<7>A new sequence specific endonuclease MraI has been purified from Micrococcus
radiodurans (ATCC 13939).  The procedure involves ammonium sulfate
precipitation of crude cell lysates followed by chromatography on Biogel-A,
DEAE-Biogel, Cibacron blue F3GA agarose, and phosphocellulose columns.  The
enzyme cleaves phage lambda DNA at three sites, and adenovirus type 2 DNA at
more than 12 sites.  It has no sites on SV40, PhiX174, PBR322 and PM2 DNA.  The
three sites on phage lambda DNA recognized by MraI are different from those
cleaved by SmaI, since the fragments obtained by the digestion of DNA with
these two enzymes are different and the double digestion generates new
fragments.  The sites of double digestion generates new fragments.  The sites
of cleavage on lambda DNA maps at 42.6, 44.9 and 83.4 percent genome length.
The enzyme requires Mg++ for its absolute activity but is active in the absence
of added 2-mercaptoethanol.  The enzyme shows activity at a broad range of
temperature and pH with optimum at 45C and pH 7.0.  Determination of the
nucleotide sequence that this enzyme recognizes is under investigation.

<>

<1>Wani, A.A., Stephens, R.E., D'Ambrosio, S.M., Hart, R.W.
<2>A sequence specific endonuclease from Micrococcus radiodurans.
<3>Biochim. Biophys. Acta
<4>697
<5>178-184
<6>1982
<7>A new sequence specific endonuclease, MraI has been purified from Micrococcus
radiodurans.  This enzyme cleaves bacteriophage lambda DNA at three sites,
adenovirus type 2 DNA at more than 12 sites and has a unique site on PhiX174
DNA.  It has no sites on SV40, PM2 and pBR322 DNA.  The three sites on phage
lambda DNA are different from those cleaved by SmaI, XmaI and XorII.  The sites
of cleavage are located at 0.424, 0.447 and 0.834 fractional lengths on the
physical map of lambda DNA.  MraI is shown to be an isoschizomer of SacII and
SstII recognizing the palindromic nucleotide sequence '5-CCGC^GG-3'.  The
enzyme shows an absolute requirement of Mg2+, but is active in the absence of
added 2-mercaptoethanol.  The enzyme shows activity at a broad range of
temperature and pH with an optimum at 45C and pH 7.0 MraI represents the first
restriction enzyme from a bacterium whose DNA lacks modified methylated bases.

<>

<1>Wank, H., SanFilippo, J., Singh, R.N., Matsuura, M., Lambowitz, A.M.
<2>A reverse transcriptase/maturase promotes splicing by binding at its own coding segment in a group II intron RNA.
<3>Mol. Cell
<4>4
<5>239-250
<6>1999
<7>Group II introns encode reverse transcriptases that promote RNA splicing (maturase activity)
and then with the excised intron form a DNA endonuclease that mediates intron mobility by
target DNA-primed reverse transcription (TPRT).  Here, we show that the primary binding site
for the maturase (LtrA) encoded by the Lactococcus lactis Ll.LtrB intron is within a region of
intron domain IV that includes the start codon of the LtrA ORF. This binding is enhanced by
other elements, particularly domain I and the EBS/IBS interactions, and helps position LtrA to
initiate cDNA synthesis in the 3' exon as occurs during TPRT. Our results suggest how the
maturase functions in RNA splicing and support the hypothesis that the reverse transcriptase
coding region was derived from an independent genetic element that was inserted into a
preexisting group II intron.

<>

<1>Wannemuehler, M.J., Overstreet, A.M., Ward, D.V., Phillips, G.J.
<2>Draft genome sequences of the altered schaedler flora, a defined bacterial community from gnotobiotic mice.
<3>Genome Announcements
<4>2
<5>e00287-14
<6>2014
<7>The altered Schaedler flora (ASF) is a bacterial community that supports normal growth and
development of gnotobiotic mice. We report here the draft genome sequences of the 8 bacteria
that comprise the ASF.

<>

<1>Ward, A.C., Davidson, B.E., Hillier, A.J., Powell, I.B.
<2>Conjugally transferable phage resistance activities from Lactococcus lactis DRC1.
<3>J. Dairy Sci.
<4>75
<5>683-691
<6>1992
<7>Lactococcus lactis ssp. lactis strain HID600 is a phage-resistant transconjugant produced by
mating HID113 (an antibiotic-resistant variant of plasmid-free strain LM0230) with L. lactis
ssp. lactis strain DRC1. Analysis of HID600 revealed that it had conjugally acquired phage
resistance, bacteriocin production, proteolytic activity, and a 131-kb plasmid. The
interaction of phages c2 and sk1 with HID113 and HID600 was studied. Plaque assays indicated
that only about 12.5% of c2 infections of HID600 produced infective centers and that c2
infections of HID600 had a longer latent period and smaller average burst size than did
infections of HID113. Less than .2% of sk1 infections of HID600 were apparently productive; no
burst was detected. The resistance to phage infection involved restriction-modification and
abortive infection effects. Such resistance might be useful in the construction of cheese
starter strains with reduced susceptibility to phage infection, although phage variants able
to infect HID600 with increased efficiency were observed.

<>

<1>Ward, B.
<2>Type IIS restriction enzyme footprint I.  Measurement of a triple helix dissociation constant with Eco57I at 25oC.
<3>Nucleic Acids Res.
<4>24
<5>2435-2440
<6>1996
<7>A method is described to measure triple helix dissociation constants by inhibiting
the cleavage of a plasmid constructed to contain a target sequence for the triplex forming
oligonucleotide (TFO) dT20 by the type IIS restriction enzyme Eco57I.  The method relies upon
the TFO's ability to block the cleavage reaction by occupying the enzyme's cleavage site but
not its
specific binding sequence.  Using this protocol, the dissociation constant for dT20 bound to
its
target was ~0.16 uM at 25oC.  The accuracy of this experiment was demonstrated by measuring
the Kd of an affinity cleavage TFO using Eco57I and Quantitative Affinity Cleavage Titration.
Type IIS restriction endonuclease footprinting should be useful for the qualitative and
quantitative
investigation of ligand-DNA interactions.

<>

<1>Ward, L.J.H., Heap, H.A., Kelly, W.J.
<2>Characterization of closely related lactococcal starter strains which show differing patterns of bacteriophage sensitivity.
<3>J. Appl. Microbiol.
<4>96
<5>144-148
<6>2004
<7>Aims: To characterize a group of closely related Lactococcus lactis subsp. lactis casein
starter strains used commercially, which differ in
their sensitivity to bacteriophages isolated from the same industrial
environment.
Methods and Results: Nine strains of L. lactis, six of which had
been used as starter cultures for lactic casein manufacture, were shown
to be closely related by pulsed-field gel electrophoresis and total DNA
profiles. Nineteen phages, which propagated on one or more of these
starter strains were isolated from industrial casein whey samples. The
phages were all small isometric-headed and could be divided into five
groups on the basis of host range on the nine strains. Most of the
phages did not give a PCR product with primers designed to detect the
two most common lactococcal small isometric phage species (936 and
P335). The hosts could be divided into six groups depending on their
phage sensitivity. Plasmids encoding genes for the cell envelope
associated PI-type proteinase, lactose metabolism and specificity
subunits of a type I restriction/modification system were identified.
Conclusions: This work demonstrates how isolates of the same
starter strain may come to be regarded as separate cultures because of
their different origins, and how these closely related strains may
differ in some of their industrially relevant characteristics.
Significance and Impact of the Study: This situation may be very
common among lactococci used as dairy starter cultures, and implies
that the dairy industry worldwide depends on a small number of
different strains.

<>

<1>Ward, N. et al.
<2>Genomic Insights into Methanotrophy: The Complete Genome Sequence of Methylococcus capsulatus (Bath).
<3>PLoS Biology
<4>2
<5>1616-1628
<6>2004
<7>Methanotrophs are ubiquitous bacteria that can use the greenhouse gas methane as a sole carbon
and energy source for growth, thus playing major roles in global carbon cycles, and in
particular, substantially reducing emissions of biologically generated methane to the
atmosphere. Despite their importance, and in contrast to organisms that play roles in other
major parts of the carbon cycle such as photosynthesis, no genome-level studies have been
published on the biology of methanotrophs. We report the first complete genome sequence to our
knowledge from an obligate methanotroph, Methylococcus capsulatus (Bath), obtained by the
shotgun sequencing approach. Analysis revealed a 3.3-Mb genome highly specialized for a
methanotrophic lifestyle, including redundant pathways predicted to be involved in
methanotrophy and duplicated genes for essential enzymes such as the methane monooxygenases.
We used phylogenomic analysis, gene order information, and comparative analysis with the
partially sequenced methylotroph Methylobacterium extorquens to detect genes of unknown
function likely to be involved in methanotrophy and methylotrophy. Genome analysis suggests
the ability of M. capsulatus to scavenge copper (including a previously unreported
nonribosomal peptide synthetase) and to use copper in regulation of methanotrophy, but the
exact regulatory mechanisms remain unclear. One of the most surprising outcomes of the project
is evidence suggesting the existence of previously unsuspected metabolic flexibility in M.
capsulatus, including an ability to grow on sugars, oxidize chemolithotrophic hydrogen and
sulfur, and live under reduced oxygen tension, all of which have implications for methanotroph
ecology. The availability of the complete genome of M. capsulatus (Bath) deepens our
understanding of methanotroph biology and its relationship to global carbon cycles. We have
gained evidence for greater metabolic flexibility than was previously known, and for genetic
components that may have biotechnological potential.

<>

<1>Ward, N.L. et al.
<2>Three genomes from the phylum Acidobacteria provide insight into the lifestyles of these microorganisms in soils.
<3>Appl. Environ. Microbiol.
<4>75
<5>2046-2056
<6>2009
<7>The complete genomes of three strains from the phylum Acidobacteria were compared.
Phylogenetic analysis placed them as a unique phylum. They share
genomic traits with members of the Proteobacteria, the Cyanobacteria, and
the Fungi. The three strains appear to be versatile heterotrophs. Genomic
and culture traits indicate the use of carbon sources that span simple
sugars to more complex substrates such as hemicellulose, cellulose, and
chitin. The genomes encode low-specificity major facilitator superfamily
transporters and high-affinity ABC transporters for sugars, suggesting
that they are best suited to low-nutrient conditions. They appear capable
of nitrate and nitrite reduction but not N(2) fixation or denitrification.
The genomes contained numerous genes that encode siderophore receptors,
but no evidence of siderophore production was found, suggesting that they
may obtain iron via interaction with other microorganisms. The presence of
cellulose synthesis genes and a large class of novel high-molecular-weight
excreted proteins suggests potential traits for desiccation resistance,
biofilm formation, and/or contribution to soil structure. Polyketide
synthase and macrolide glycosylation genes suggest the production of novel
antimicrobial compounds. Genes that encode a variety of novel proteins
were also identified. The abundance of acidobacteria in soils worldwide
and the breadth of potential carbon use by the sequenced strains suggest
significant and previously unrecognized contributions to the terrestrial
carbon cycle. Combining our genomic evidence with available culture
traits, we postulate that cells of these isolates are long-lived, divide
slowly, exhibit slow metabolic rates under low-nutrient conditions, and
are well equipped to tolerate fluctuations in soil hydration.

<>

<1>Waring, R.B., Davies, R.W., Scazzocchio, C., Brown, T.A.
<2>Internal structure of the mitochondrial intron of Aspergillus nidulans.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>79
<5>6332-6336
<6>1982
<7>The intron of the mitochondrial apocytochrome b gene, cobA, of Aspergillus nidulans has been
subjected to sequence analysis.  It contains an open reading frame of 957 base pairs
contiguous with the preceding exon.  Regions of the translated open reading frames of cobA and
the third intron of the cob gene in yeast show high amino acid homology.  Comparison of the
cobA intron with this and other yeast introns indicates that cobA codes for a maturase protein
that splices out the intron encoding it and possibly other mitochondrial introns.  Two very
similar decamer peptides are found in the protein sequences of the cobA intron, four
mitochondrial yeast introns, and the yeast mitochondrial sequence reading frame 1 and may be
diagnostic of one class of maturase-coding introns.  Four short DNA sequences, two of which
are in the region defined by box9 and box2 mutations in the cob gene of yeast, are conserved
in cobA and certain yeast introns.  Comparison with three yeast introns strongly suggests that
the first 200 base pairs of the open reading frame of the cobA intron do not code for any
amino acids present in the putative maturase protein but are required for splicing or the
control of splicing, or both.

<>

<1>Warncke, S., Gegout, A., Carell, T.
<2>Phosphorothioation of Oligonucleotides Strongly Influences the Inhibition of Bacterial (M.Hhal) and Human (Dnmt1) DNA Methyltransferases.
<3>Chembiochem
<4>10
<5>728-734
<6>2009
<7>The cytidine analogue 5-fluoro-2'-deoxycytidine (dC(F)) is a mechanism-based inhibitor of DNA
methyltransferases. We report the
synthesis of short 18-mer dsDNA oligomers containing a
triple-hemimethylated CpG motive as a recognition sequence for the
human methyltransferase Dnmt1. The DNA strands carry within these CpG
islands dC(F) building blocks that function as mechanism-based
inhibitors of the analyzed methyltransferases. In addition, we replaced
the phosphodiester backbones at defined positions by phosphorothioates.
These hypermodified DNA strands were investigated as inhibitors of the
DNA methyltransferases M.Hhal and Dnmtl in vitro. We could show that
both methylases behave substantially differently in respect to the
amount of DNA backbone modification.

<>

<1>Warnecke, F. et al.
<2>Metagenomic and functional analysis of hindgut microbiota of a wood-feeding higher termite.
<3>Nature
<4>450
<5>560-565
<6>2007
<7>From the standpoints of both basic research and biotechnology, there is
considerable interest in reaching a clearer understanding of the diversity
of biological mechanisms employed during lignocellulose degradation.
Globally, termites are an extremely successful group of wood-degrading
organisms and are therefore important both for their roles in carbon
turnover in the environment and as potential sources of biochemical
catalysts for efforts aimed at converting wood into biofuels. Only
recently have data supported any direct role for the symbiotic bacteria in
the gut of the termite in cellulose and xylan hydrolysis. Here we use a
metagenomic analysis of the bacterial community resident in the hindgut
paunch of a wood-feeding 'higher' Nasutitermes species (which do not
contain cellulose-fermenting protozoa) to show the presence of a large,
diverse set of bacterial genes for cellulose and xylan hydrolysis. Many of
these genes were expressed in vivo or had cellulase activity in vitro, and
further analyses implicate spirochete and fibrobacter species in gut
lignocellulose degradation. New insights into other important symbiotic
functions including H2 metabolism, CO2-reductive acetogenesis and N2
fixation are also provided by this first system-wide gene analysis of a
microbial community specialized towards plant lignocellulose degradation.
Our results underscore how complex even a 1-microl environment can be.

<>

<1>Warnecke, P.M., Bestor, T.H.
<2>Cytosine methylation and human cancer.
<3>Curr. Opin. Oncol.
<4>12
<5>68-73
<6>2000
<7>Abnormalities of genomic methylation patterns have been attributed a role in carcinogenesis
since the early 1980s, when large-scale demethylation of the genome
was thought be an early event in multistep colorectal carcinogenesis. In the 1990s, local de
novo methylation (with or without global demethylation) at tumor suppressor loci
was held to be involved in silencing of tumor suppressor genes. The mechanisms that might
mediate methylation and demethylation in carcinogenesis remain obscure, and
there are questions as to whether the methylation changes are a cause or consequence of
cellular transformation and clonal expansion. It is also important to derive a set of
defined criteria by which a tumor suppressor gene can be concluded to have been inactivated by
DNA methylation in a manner that contributes to carcinogenesis.

<>

<1>Warnecke, P.M., Biniszkiewicz, D., Jaenisch, R., Frommer, M., Clark, S.J.
<2>Sequence-specific methylation of the Mouse H19 gene in embryonic cells deficient in the Dnmt-1 gene.
<3>Dev. Genet.
<4>22
<5>111-121
<6>1998
<7>We have used Dnmtc/c ES cells that are homozygous for disruption of the DNA methyltransferase
gene to address how de novo methylation is propagated and whether it is directed to specific
sites in the early embryo.  We examined the imprinted H19 gene and the specific-sequence
region implicated as an "imprinting mark" to determine whether de novo methylation was
occurring at a restricted set of sites.  Since the "imprinting mark" was found to be
methylated differentially at all stages of development, we reasoned that the sequence may
still be a target for the de novo methylation activity found in the Dnmtc/c cells, even though
the loss of maintenance methylase activity renders the H19 promoter active.  We used bisulfite
genomic sequencing to determine the methylation state of the imprinted region of the H19 gene
and found a low level of DNA methylation at specific single CpG sites in the upstream region
of the imprinted H19 sequence in the Dnmtc/c mutant ES cells.  Moreover, these CpG sites
appeared to be favored targets for further de novo methylation of neighboring CpG sites in
rescued ES cells, which possess apparently normal maintenance activity.  Our data provide
further evidence for a separate methylating activity in ES cells and indicate that this
activity displays sequence specificity.

<>

<1>Warren, R.A.J.
<2>Modified Bases in Bacteriophage DNAs.
<3>Annu. Rev. Microbiol.
<4>34
<5>137-158
<6>1980
<7>This review considers bacteriophage DNAs in which all or a major proportion of
one of the four bases found commonly in DNA is replaced by a modified base.  It
does not consider the methylated bases that are found in small amounts in DNA
and that function usually to protect DNA against specific restriction
endonucleases.  The first modified base in a bacteriophage DNA,
5-hydroxymethylcytosine (hmCyt), was discovered 27 years ago.  It played a
crucial role in the development of the biochemistry of T-even phage-infected
Escherichia coli.  Since then, modified bases have been found in a variety of
other bacteriophages.  They are of interest from the point of view of their
biochemistry and, more recently, their effects on DNA conformation and
function.  The biosynthesis of some of these bases has been reviewed
extensively.  This review emphasizes the more recent developments in modified
base biochemistry and considers also some of their effects on DNA structure and
function.

<>

<1>Warren, W., Merion, M.
<2>Rapid HPLC purification of a restriction enzyme from Sphaerotilus sp.
<3>BioChromatography
<4>3
<5>225-230
<6>1988
<7>The rapid purification of the Type II restriction endonuclease SspI from
contaminating exo- and endonucleases contained in a cell lysate of the
bacterium Sphaerotilus sp. is described using a combination of anion and cation
exchange media.  Nucleic acids and a substantial portion of non-restriction
enzyme proteins were quickly removed from the initial cell lysate via solid
phase extraction using an anion exchange cartridge.  The SspI was subsequently
purified free from contaminating nucleases using cation exchange column
chromatography.  Compared to the initial cell lysate, the overall recovery of
SspI activity in the final product was greater than 75%.  In addition this
rapid 2-step procedure resulted in a 330-fold increase in enzyme specific
activity.

<>

<1>Waschkau, B., Waldeck, J., Wieland, S., Eichstadt, R., Meinhardt, F.
<2>Generation of readily transformable Bacillus licheniformis mutants.
<3>Appl. Microbiol. Biotechnol.
<4>78
<5>181-188
<6>2008
<7>A set of mutants was generated by targeted deletion of the hsdR loci of two type I restriction
modification systems ( RMS) identified in
Bacillus licheniformis DSM13. Single as well as double knock-outs
resulted in strains being readily transformable with plasmids isolated
from Bacilli. Introduction of shuttle plasmids isolated from
Escherichia coli was routinely possible when the double mutant B.
licheniformis MW3 (Delta hsdR1, Delta hsdR2) was used in transformation
experiments. Growth and secretion of extracellular enzymes were not
affected in any of the mutants. Thus, along with an optimized
transformation protocol, this study makes available an urgently needed
transformation system for this industrially exploited species.

<>

<1>Wassenegger, M.
<2>RNA-directed DNA methylation.
<3>Plant Mol. Biol.
<4>43
<5>203-220
<6>2000
<7>RNA-DNA interactions can serve as a signal that triggers de novo DNA methylation in plants. As
yet, this RNA-directed DNA methylation mechanism merely targets transgenes, but it appears
likely that methylation of some endogenous sequences is also directed by RNA. RNA-directed
methylation of cytosine residues specifically occurs along the DNA regions that are
complementary to the directing RNA pointing to the formation of a putative RNA-DNA duplex.
Dense methylation patterns and the methylation of cytosine residues at symmetric and
asymmetric sites are detectable on both DNA strands within these regions. Methylation
progressively decreases in the sequences adjacent to the putative RNA-DNA duplex. The extreme
sensitivity of RNA-directed DNA methylation was demonstrated by analyzing a short 30 bp DNA
region that was complementary to the targeting RNA. Association of RNA-directed DNA
methylation with homology-dependent gene silencing indicated that the methylation-directing
RNA molecules may be double-stranded or may contain double-stranded regions. Whereas the
function of DNA methylation in transcriptional gene silencing is nearly understood, its role
in post-transcriptional gene silencing is still under discussion. In mammals, X-chromosome
inactivation and genomic imprinting are associated with DNA methylation but how methylation is
initiated is unclear. The observation of a correlation between specific antisense RNAs and
transcriptional and post-transcriptional gene silencing may indicate that RNA-directed DNA
methylation is involved in epigenetic gene regulation throughout eukaryotes.

<>

<1>Watabe, H., Iino, T., Kaneko, T., Shibata, T., Ando, T.
<2>A new class of site-specific endodeoxyribonucleases.
<3>J. Biol. Chem.
<4>258
<5>4663-4665
<6>1983
<7>We had found that yeasts had intracellular endodeoxyribonucleases that cut
phage DNA into a set of double-stranded fragments with discrete chain lengths.
We purified one of them to apparent homogeneity from Saccharomyces cerevisiae
and designated it Endo.Sce.I.  Sequence analysis around 5 cleavage site in
plasmid DNA and phage DNA revealed that Endo.SceI cuts a defined phosphodiester
bond in each strand of double helix at the cleavage sites and produces free
cohesive ends consisting of 4 nucleotides protruding at 3'-termini.  However,
unlike in the case of prokaryotic type II-restriction endonucleases, (i)
Endo.SceI seems to consist of two nonidentical subunits, (ii) no common
palindrome or consensus sequence including more than 5 base pairs is detected
at or near these cleavage sites, and (iii) Endo.SceI can cut the DNA isolated
from the cells that produced Endo.SceI.  All of the 5 cleavage sites are
included in inverted repeats, but these inverted repeats are variable in size,
nucleotide sequence, and distance between repeating units.  An inverted repeat
itself is not a structure recognized by Endo.SceI.  This study shows that
Endo.SceI is the first example of eukaryotic site-specific endonuclease and has
properties, as described above, which distinguish it from prokaryotic
restriction endoncleases.

<>

<1>Watabe, H., Shibata, T., Ando, T.
<2>Site-specific endo-deoxyribonucleases in eukaryotes: endonucleases of yeasts, Saccharomyces and Pichia.
<3>J. Biochem. (Tokyo)
<4>90
<5>1623-1632
<6>1981
<7>A class of endo-deoxyribonucleases was found in cell-free extracts from yeasts;
Saccharomyces cerevisiae, S. uvarum and Pichia membranaefaciens.  These
endonucleases cleaved double-stranded DNA so that the treated DNA exhibited a
set of discrete bands on an agarose gel-electrophoregram.  The profiles
obtained with a certain DNA were unique to each endonuclease, suggesting that
these yeast endonucleases cleave DNA at well-defined sites specific to each
endonuclease.

<>

<1>Watabe, H.-O., Shibata, T., Iino, T., Ando, T.
<2>Purification of a eukaryotic site-specific endonuclease, Endo.SceI, from Saccharomyces cerevisiae and effectors on its specificity and activity.
<3>J. Biochem. (Tokyo)
<4>95
<5>1677-1690
<6>1984
<7>A site-specific endonuclease (Endo.SceI) which caused double-strand scission of DNA was highly
purified from a eukaryote, Saccharomyces cerevisiae IAM4274.  The molecular weight of the
active form of Endo.SceI was estimated to be 120,000 and 110,000 by sedimentation analysis on
a glycerol density gradient and gel filtration on Ultrogel AcA34, respectively.  Analysis of
the fractions from the last column chromatography by polyacrylamide gel electrophoresis in the
presence of sodium dodecyl sulfate and by an assay of the endonucleolytic activities suggested
that Endo.SceI consists of two non-identical subunits with molecular weights of 75,000 and
50,000.  Unlike restriction endonucleases, Endo.SceI was active on chromosomal DNA of the
cells which produced Endo.SceI.  Single-stranded DNA was not cleaved by Endo.SceI, but
inhibited the endonucleolytic activity of the enzyme on double-stranded DNA.  The
endonucleolytic activity of Endo.SceI required magnesium ions (Mg2+) as a sole cofactor; Mg2+
could not be replaced by Ca2+ or Zn2+.  When Mg2+ was replaced by manganese ions (Mn2+),
extensively purified Endo.SceI cleaved double-stranded DNA at many other sites in addition to
the sites at which DNA was cleaved in the presence of Mg2+.  Experiments indicated that this
is not the activation of contaminating endonuclease in the preparation of Endo.SceI, but the
result of relaxation in the site-specificity of cleavage.

<>

<1>Watahiki, M., Nagai, Y.
<2>Method of examining types of pathogenic bacteria, Enterohemorrhagic Escherichia coli and bacteria belonging to the genus Legionella.
<3>International Patent Office
<4>WO 2007097410 A
<5>
<6>2007
<7>To provide a method of examining types of pathogenic bacteria including EHEC and bacteria
belonging to the genus Legionella depending on polymorphisms in the base sequences that can be
stored as text data.  A method of examining types of pathogenic bacteria.  Single nucleotide
polymorphisms contained in a plural number of base sequence regions, which are highly
homologous with each other, in the genes of the pathogenic bacteria are identified and the
genotypes are determined based on the single nucleotide polymorphism data thus obtained.  The
above-described pathogenic bacteria belong to enterohemorrhagic Escherichia coli and the
highly homologous base sequence regions as described above occur in the prophage genomic
sequences of the E. coli bacteria.  The above-described pathogenic bacteria belong to the
genus Legionella.

<>

<1>Watahiki, M., Nagai, Y.
<2>Determination method of type of Pathogenic bacteria, enterohemorrhagic Escherichia coli (EHEC) and Legionella.
<3>Japanese Patent Office
<4>JP 2007252371 A
<5>
<6>2007
<7>
<>

<1>Watanabe, F., Yu, F., Kita, K.
<2>DNA and protein sequences of restriction enzyme Rrhj1I and its modification enzyme from Rhodococcus rhodochrous, and their use for genetic engineering.
<3>Japanese Patent Office
<4>JP 2007259853 A
<5>
<6>2007
<7>
<>

<1>Watanabe, K.I., Ehara, M., Inagaki, Y., Ohama, T.
<2>Distinctive origins of group I introns found in the COXI genes of three green algae.
<3>Gene
<4>213
<5>1-7
<6>1998
<7>Upon surveying the cytochrome c oxidase subunit I (COXI) gene of green algae, we found group I
introns in three species of algae, Chlorella vulgaris (Cv), Scenedesmus quadricauda (Sq) and
Protosiphon botryoides (Pb). The comparative analysis of these nucleotide sequences and their
secondary structures revealed that the introns of Cv, Sq, and Pb belong to groups IB1, ID, and
IB2, respectively. Each of the three introns contained an open reading frame (ORF) that showed
a similarity to the sequence of the LAGLIDADG endonuclease family. However, each of the
intronic ORFs in Sq and Pb had a discontinuity in the middle of the sequences coding for the
LAGLIDADG endonuclease. Either of the two ORFs could be restored to a sequence homologous to
the LAGLIDADG endonuclease by the insertion of a nucleotide in the appropriate position. In
Sq, a putative pseudo-knot structure was detected in the intronic ORF. This suggests the
occurrence of a ribosomal frameshift in the translation of the ORF, because such pseudo-knot
structures are common in viral ORFs employing a (-1) ribosomal frameshift. In the phylogenetic
tree that was inferred from the amino acid sequences of algal and non-algal intronic ORFs, the
three algal ORFs did not make a cluster, but were scattered throughout the tree. In addition.
Each of the three algal ORFs showed a close relationship to the ORFs of non-algal introns that
were inserted at the corresponding site of the COXI gene, suggesting distinctive origins of
the three algal introns via independent horizontal transfers.

<>

<1>Watanabe, M., Yuzawa, H., Handa, N., Kobayashi, I.
<2>Hyperthermophilic DNA methyltransferase M.Pabl from the archaeon Pyrococcus abyssi.
<3>Appl. Environ. Microbiol.
<4>72
<5>5367-5375
<6>2006
<7>Genome sequence comparisons among multiple species of Pyrococcus, a hyperthermophilic
archaeon, revealed a linkage between a putative
restriction-modification gene complex and several large genome
polymorphisms/rearrangements. From a region apparently inserted into
the Pyrococcus abyssi genome, a hyperthermoresistant restriction enzyme
[PabI; 5'-(GTA/C)] with a novel structure was discovered. In the
present work, the neighboring methyltransferase homologue, M.PabI, was
characterized. Its N-terminal half showed high similarities to the M
subunit of type I systems and a modification enzyme of an atypical type
II system, M.AhdI, while its C-terminal half showed high similarity to
the S subunit of type I systems. M.PabI expressed within Escherichia
coli protected PahI sites from RsaI, a PabI isoschizomer. M.PabI,
purified following overexpression, was shown to generate 5'-GTm6AC,
which provides protection against PabI digestion. M.PabI was found to
be highly thermophilic; it showed methylation at 95 degrees C and
retained at least half the activity after 9 min at 95 degrees C. This
hyperthermophilicity allowed us to obtain activation energy and other
thermodynamic parameters for the first time for any DNA
methyltransferases. We also determined the kinetic parameters of
k(cat), K-m,K- DNA, and K-m,K- AdoMet. The activity of M.PabI was
optimal at a slightly acidic pH and at an NaCl concentration of 200 to
500 mM and was inhibited by Zn2+ but not by Mg2+, Ca2+, or Mn2+. These
and previous results suggest that this unique methyltransferase and
PahI constitute a type II restriction-modification gene complex that
inserted into the P. abyssi genome relatively recently. As the most
thermophilic of all the characterized DNA methyltransferases, M.PabI
may help in the analysis of DNA methylation and its application to DNA
engineering.

<>

<1>Watanabe, N., Takasaki, Y., Sato, C., Ando, S., Tanaka, I.
<2>Structures of restriction endonuclease HindIII in complex with its cognate DNA and divalent cations.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>65
<5>1326-1333
<6>2009
<7>The three-dimensional crystal structures of HindIII bound to its cognate DNA with and without
divalent cations were solved at 2.17 and
2.00 angstrom resolution, respectively. HindIII forms a dimer. The
structures showed that HindIII belongs to the EcoRI-like (alpha-class)
subfamily of type II restriction endonucleases. The cognate DNA-complex
structures revealed the specific DNA-recognition mechanism of HindIII
by which it recognizes the palindromic sequence A/AGCTT. In the Mg2+
ion-soaked structure the DNA was cleaved and two ions were bound at
each active site, corresponding to the two-metal-ion mechanism.

<>

<1>Watanabe, S., Shiwa, Y., Itaya, M., Yoshikawa, H.
<2>Complete Sequence of the First Chimera Genome Constructed by Cloning the Whole Genome of Synechocystis Strain PCC6803 into the Bacillus subtilis 168 Genome.
<3>J. Bacteriol.
<4>194
<5>7007
<6>2012
<7>Genome synthesis of existing or designed genomes is made feasible by the first successful
cloning of a cyanobacterium, Synechocystis PCC6803, in Gram-positive,
endospore-forming Bacillus subtilis. Whole-genome sequence analysis of the
isolate and parental B. subtilis strains provides clues for identifying single
nucleotide polymorphisms (SNPs) in the 2 complete bacterial genomes in one cell.

<>

<1>Watanabe, T., Kojima, H., Fukui, M.
<2>Draft genome sequence of a psychrotolerant sulfur-oxidizing bacterium, Sulfuricella denitrificans skB26, and proteomic insights into cold adaptation.
<3>Appl. Environ. Microbiol.
<4>78
<5>6545-6549
<6>2012
<7>Except for several conspicuous cases, very little is known about sulfur oxidizers
living in natural freshwater environments. Sulfuricella denitrificans skB26 is a
psychrotolerant sulfur oxidizer recently isolated from a freshwater lake as a
representative of a new genus in the class Betaproteobacteria. In this study, an
approximately 3.2-Mb draft genome sequence of strain skB26 was obtained. In the
draft genome, consisting of 23 contigs, a single rRNA operon, 43 tRNA genes, and
3,133 coding sequences were identified. The identified genes include those
required for sulfur oxidation, denitrification, and carbon fixation. Comparative
proteomic analysis was conducted to assess cold adaptation mechanisms of this
organism. From cells grown at 22 degrees C and 5 degrees C, proteins were
extracted for analysis by nano-liquid chromatography-electrospray
ionization-tandem mass spectrometry. In the cells cultured at 5 degrees C,
relative abundances of ribosomal proteins, cold shock proteins, and DEAD/DEAH box
RNA helicases were increased in comparison to those at 22 degrees C. These
results suggest that maintenance of proper translation is critical for growth
under low-temperature conditions, similar to the case for other cold-adapted
prokaryotes.

<>

<1>Watanabe, T., Maruyama, F., Nozawa, T., Aoki, A., Okano, S., Shibata, Y., Oshima, K., Kurokawa, K., Hattori, M., Nakagawa, I., Abiko, Y.
<2>Complete Genome Sequence of the Bacterium Porphyromonas gingivalis TDC60, Which Causes Periodontal Disease.
<3>J. Bacteriol.
<4>193
<5>4259-4260
<6>2011
<7>Porphyromonas gingivalis is a black-pigmented asaccharolytic anaerobe and a major causative
agent of periodontitis. Here, we report the complete
genome sequence of P. gingivalis strain TDC60, which was recently isolated
from a severe periodontal lesion in a Japanese patient.

<>

<1>Watanabe, T., Nishida, H., Ogata, C., Arai, T., Sato, S.
<2>Episome-mediated transfer of drug resistance in enterobacteriaceae.  VII.  Two types of naturally occurring R factors.
<3>J. Bacteriol.
<4>88
<5>716-726
<6>1964
<7>Naturally occurring R factors are classified into two types, fi+ and fi-,
depending on their fi characters.  The term fi is an abbreviation of fertility
inhibition and fi+ and fi- mean, respectively, the presence and absence of
suppression of the functions of the sex factor F of Escherichia coli K-12.  It
was found that fi- R factors reduce the efficiency of plating of phages lambda
and T1 in K-12; fi+ R factors did not have this inhibitory action.  One of the
fi- R factors reduced the efficiency of plating of phage T7 as well.  Phages
lambda and T1 underwent host-induced modifications in the host carrying some
fi- R factors.  At least two types of fi- R factors were recognized by the
types of their restriction and host-induced modification of these phages.
CaCl2 exhibited antagonistic actions against the restrictions of phages lambda
and T1 by fi- R factors.  Transduction of the ability to ferment galactose with
HFT lysates of lambda was reduced by fi- R factors.  Ultraviolet induction of
lambda was not affected by any R factors.  Furthermore, adsorption of phages
lambda and T1 was not altered by the presence of any R factors.  From these
results, we concluded that the suppression of progeny formation of these phages
by fi- R factors is due to some step(s) after adsorption of the phages to the
bacteria.  Superinfection immunity and mutual exclusion were found between two
different fi+ R factors but not between fi+ and fi- R factors.  The two
different fi+ R factors were frequently genetically recombined, but fi+ and fi-
R factors were not genetically recombined, as indicated by findings of
independent transfer of these R factors by conjugation and by transduction from
the donors having these two R factors.  It was assumed from these findings that
fi+ and fi- R factors are considerably different episomes having different
resistance-transfer factors.

<>

<1>Watanabe, T., Takano, T., Arai, T., Nishida, H., Sato, S.
<2>Episome-mediated transfer of drug resistance in Enterobacteriaceae.   X.  Restriction and modification of phages by fi- R factors.
<3>J. Bacteriol.
<4>92
<5>477-486
<6>1966
<7>A fi- R factor, which restricts phages lambda, T1, and T7 without modifying
them, was found to restrict and not to modify an F- specific phage, W-31, in
Escherichia coli K-12, but not to restrict phage P-22 in Salmonella typhimurium
LT-2, whereas other fi- R factors restricted and modified P-22 but not W-31;
fi+ R factors did not restrict these phages.  Transduction and lysogenization
with phages lambda and P-22 were reduced by these fi- R factors in K-23 and
LT-2, respectively, and the transducing phages lambda and P-22 were modified by
these fi- R factors.  Spontaneous as well as ultraviolet-induced production of
phage P-22 and zygotic induction of phage lambda were not significantly
affected by any R factor.  Injection of the nucleic acids of phages T1 and
lambda was not affected by R factors, but the injected phage nucleic acids were
rapidly broken down in the bacteria carrying fi- R factors.  The nucleic acids
of the modified phages were not broken down in these bacteria.  It was assumed
from these results that the mechanism of restriction of phages by fi- R factors
is due to the breakdown of the injected phage nucleic acids by a
deoxyribonuclease(s), presumably located near the cell surface in the cells
carrying fi- R factors.  The deoxyribonuclease(s), formed in the cells carrying
the nonmodifying fi- R factor, is considered to be different from that
synthesized in the cells carrying the modifying fi- R factors.  It was further
shown that the average burst sizes of the unmodified as well as modified phages
are slightly reduced by the presence of the fi- R factors.

<>

<1>Waterbury, P.G., Rehfuss, R.P., Carroll, W.T., Smardon, A.M., Faldasz, B.D., Huckaby, C.S., Lane, M.J.
<2>Specific cleavage of the yeast genome at 5'-ATCGATCGAT-3'.
<3>Nucleic Acids Res.
<4>17
<5>9493
<6>1989
<7>M.ClaI followed by DpnI cleavage.  Conditions are described for complete
cleavage of yeast DNA.

<>

<1>Waterman, S.R., Hackett, J., Manning, P.A.
<2>Isolation of a restriction-less mutant and development of a shuttle vector for the genetic analysis of Campylobacter hyointestinalis.
<3>Gene
<4>125
<5>19-24
<6>1993
<7>A cosmid shuttle cloning vector, pCHI15, was constructed which could be mobilized from
Escherichia coli K-12 to a putative restriction-less mutant of Campylobacter hyointestinalis,
C. fetus subsp. fetus, and C. fetus subsp. venerealis at a frequency of 10-4 transconjugants
per donor. A previously described C. coli shuttle vector, pILL550, could not be mobilized into
the C. hyointestinalis restriction-less mutant, implying that the C. coli replicon was not
functional in a C. hyointestinalis host. The type strains of C. jejuni, C. coli, C. fetus
subsp. fetus, and C. hyointestinalis were analysed for their ability to be transformed by
plasmid DNA which had been modified by other Campylobacter species. Each Campylobacter species
was found to be most efficiently transformed by plasmid DNA that had been previously passaged
in the same species. pCHI15 could be mobilized from C. coli into C. fetus subsp. fetus and the
putative restriction-less mutant of C. hyointestinalis at a frequency of 3.0 x 10-4 and 2.5 x
10-3 transconjugants per donor, respectively.

<>

<1>Waters, E. et al.
<2>The genome of Nanoarchaeum equitans: Insights into early archaeal evolution and derived parasitism.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>100
<5>12984-12988
<6>2003
<7>The hyperthermophile Nanoarchaeum equitans is an obligate symbiont growing in coculture with
the crenarchaeon Ignicoccus. Ribosomal protein and
rRNA-based phylogenies place its branching point early in the archaeal
lineage, representing the new archaeal kingdom Nanoarchaeota. The N.
equitans genome (490,885 base pairs) encodes the machinery for information
processing and repair, but lacks genes for lipid, cofactor, amino acid, or
nucleotide biosyntheses. It is the smallest microbial genome sequenced to
date, and also one of the most compact, with 95% of the DNA predicted to
encode proteins or stable RNAs. Its limited biosynthetic and catabolic
capacity indicates that N. equitans' symbiotic relationship to Ignicoccus
is parasitic, making it the only known archaeal parasite. Unlike the small
genomes of bacterial parasites that are undergoing reductive evolution, N.
equitans has few pseudogenes or extensive regions of noncoding DNA. This
organism represents a basal archaeal lineage and has a highly reduced
genome.

<>

<1>Waters, T.R., Connolly, B.A.
<2>Continuous spectrophotometric assay for restriction endonucleases using synthetic oligodeoxynucleotides and based on the hyperchromic effect.
<3>Anal. Biochem.
<4>204
<5>204-209
<6>1992
<7>A continuous spectrophotometric assay for the EcoRV restriction endonuclease has been
developed. The synthetic self-complementary oligonucleotide d(GACGATATCGTC) (which is double
stranded under the assay conditions) is used as the substrate. The EcoRV endonuclease
recognizes d(GATATC)sequences cutting between the cental T and dA bases. Thus d(GACGATATCGTC)
is converted to d(GACGAT) and d(pATCGTC) during catalysis. Both of the hexameric products are
single stranded under the assay conditions. The conversion of the dodecameric substrate to the
two hexameric products and the concomitant change from double- to single-stranded DNA is
associated with an increase in absorbance at 254 nm due to the hyperchromic effect. This
change can be used to monitor column effluents for endonuclease activity and also for Km and
kcat determination under steady-state kinetic conditions.

<>

<1>Waters, T.R., Connolly, B.A.
<2>Interaction of the restriction endonuclease EcoRV with the deoxyguanosine and deoxycytidine bases in its recognition sequence.
<3>Biochemistry
<4>33
<5>1812-1819
<6>1994
<7>The interaction of the EcoRV restriction endonuclease with the dG and the dC bases in its
recognition sequence (GATATC) has been studied using base analogues. These modified dG and dC
bases each have a single potential protein contact removed. The analogues have been
incorporated into the self-complementary dodecamer d(pGACGATATCGTC) at the appropriate
positions (underlined). Many of the analogues caused no change in the Tm of the duplex or else
lowered the Tm by a small amount such that a duplex was still formed at temperatures suitable
for enzyme assay. However, the dG analogue 2-aminopurine-1-beta-D-2'-deoxyriboside
destabilized the duplex to such an extent that the 12'-mer could not be used for enzyme
assays. To overcome this, a longer self-complementary 18'-mer was used with this modified
base. The circular dichroism spectra of the modified base containing 12'-mers (and the
18'-mer in the case of 2-aminopurine) were very similar to the parent sequences lacking
modified bases. This demonstrates the formation of B-DNA structures in all cases and similar
overall conformations. The km and kcat values for the various modified oligomers have been
determined, and these data have been used to assess the roles that functional groups on the dG
and dC bases play in the recognition and hydrolysis of GATATC sequences by the endonuclease.
The results obtained here have been compared to the crystal structures of the EcoRV complexed
with a GATATC sequence, and this has allowed a critical evaluation of the base analogue
approach. Both methods indicate that the 6-keto oxygen and 7-ring nitrogen of dG exposed in
the major groove are vital for DNA recognition and hydrolysis.

<>

<1>Watrob, H., Liu, W., Chen, Y., Bartlett, S.G., Jen-Jacobson, L., Barkley, M.D.
<2>Solution conformation of EcoRI restriction endonuclease changes upon binding of cognate DNA and Mg2+ cofactor.
<3>Biochemistry
<4>40
<5>683-692
<6>2001
<7>EcoRI endonuclease has two tryptophans at positions 104 and 246 on the protein surface. A
single tryptophan mutant containing Trp246 and a
single cysteine labeling site at the N-terminus was used to determine
the position of the N-terminus in the protein structure. The N-termini
of EcoRI endonuclease are essential for tight binding and catalysis yet
are not resolved in any of the crystal structures. Resonance energy
transfer was used to measure the distance from the Trp246 donor to IAEDANS
or MIANS acceptors at Cys3. The distance is 36 angstroms in the apoenzyme,
decreasing to 26 angstroms in the DNA complex. Molecular modeling
suggests that the N-termini are located at the dimer interface formed
by the loops comprising residues 221-232. Protein conformational
changes upon binding of cognate DNA and cofactor Mg2+ were monitored by
tryptophan fluorescence of the single tryptophan mutant and wild-type
endonuclease. The fluorescence decay of Trp246 is a triple exponential
with lifetimes of 7, 3.5, and 0.7 ns. The decay-associated spectra of
the 7- and 3.5-ns components have emission maxima at approximately 345 and
approximately 338 nm in the apoenzyme, which shift to approximately 340 and
approximately 348 nm in the DNA complex. The fluorescence quantum yield of
the single tryptophan mutant drops 30% in the DNA complex, as compared
to 10% for the wild-type endonuclease. Fluorescence changes of Trp 104 upon
binding of DNA were inferred by comparison of the decay-associated
spectra of wild type and single tryptophan mutant. Fluorescence changes
are related to changes in proximity and orientation of quenching
functional groups in the tryptophan microenvironments, as seen in the
crystal structures.

<>

<1>Watson, M.A., Gowers, D.M., Halford, S.E.
<2>Alternative geometries of DNA looping: an analysis using the SfiI endonuclease.
<3>J. Mol. Biol.
<4>298
<5>461-475
<6>2000
<7>Many processes are governed by proteins that bind to separate sites in DNA and loop out the
intervening DNA, but the geometries of the loops have seldom been determined. The SfiI
endonuclease cleaves DNA after interacting with two recognition sites, and is a favourable
system for the analysis of DNA looping. A gel-shift assay was used here to examine the binding
of SfiI to a series of linear DNA molecules containing two SfiI sites separated by 109-170
base-pairs. The complexes in which SfiI trapped a loop by binding to two sites in the same DNA
were separated from the complexes containing SfiI bound to separate DNA molecules. Step-wise
changes in the inter-site spacing generated two forms of the looped complex with different
electrophoretic mobilities. The yields of each looped complex and the complexes from
intermolecular synapses all varied cyclically with the inter-site spacing, with similar
periodicities ( approximately 10.5 base-pairs) but with different phases. One looped complex
predominated whenever the DNA between the sites needed to be underwound in order to produce
the correct helical orientation of the binding sites. The other looped complex predominated
whenever the intervening DNA needed to be overwound. We conclude that the former has trapped a
right-handed loop with a negative node and the latter a left-handed loop with a positive node.

<>

<1>Watson, M.E., Jarisch, J., Smith, A.L.
<2>Inactivation of deoxyadenosine methyltransferase (dam) attenuates Haemophilus influenzae virulence.
<3>Mol. Microbiol.
<4>53
<5>651-664
<6>2004
<7>Mutants in deoxyadenosine methyltransferase (dam) from many Gram-negative pathogens suggest
multiple roles for Dam methylase:
directing post-replicative DNA mismatch repair to the correct strand,
guiding the temporal control of DNA replication and regulating the
expression of multiple genes (including virulence factors) by
differential promoter methylation. Dam methylase (HI0209) in strain Rd
KW20 was inactivated in Haemophilus influenzae strains Rd KW20, Strain
12 and INT-1; restriction with Dam methylation-sensitive enzymes DpnI
and DpnII confirmed the absence of Dam methylation, which was restored
by complementation with a single copy of dam ectopically expressed in
cis. Despite the lack of increased mutation frequency, the dam mutants
had a 2-aminopurine-susceptible phenotype that could be suppressed by
secondary mutations in mutS, suggesting a role for Dam in H. influenzae
DNA mismatch repair. Invasion of human brain microvascular endothelial
cells (HBMECs) and human respiratory epithelial cells (NCI-H292) by the
dam mutants was significantly attenuated in all strains, suggesting the
absence of a Dam-regulated event necessary for uptake or invasion of
host cells. Intracellular replication was inhibited only in the Strain
12 dam mutant, whereas in the infant rat model of infection, the INT-1
dam mutant was less virulent. Dam activity appears to be necessary for
both in vitro and in vivo virulence in a strain-dependent fashion and
may function as a regulator of gene expression including virulence
factors.

<>

<1>Watson, R., Zuker, M., Martin, S.M., Visentin, L.P.
<2>A new site-specific endonuclease from Neisseria cinerea.
<3>FEBS Lett.
<4>118
<5>47-50
<6>1980
<7>Sequence-specific deoxyribonucleases have greatly facilitated the analysis and
in vitro manipulation of DNA.  Most of the known type II restriction enzymes
recognize palindromic DNA sequences.  Although a large number of these enzymes
have been characterized, many of the possible palindromic DNA sequences are not
recognized by any known enzyme.  New restriction enzymes with unique
recognition sites are desirable, as they increase the flexibility of
recombinant DNA techniques.  We have screened a number of species of the genus
Neisseria for restriction endonucleases, and report here the isolation and
characterization of an enzyme from Neisseria cinerea which cleaves DNA at an
unreported recognition sequence 5' . . . CC(C/G)GG . .3'.  The identification
of this sequence was assisted by a computer compilation of the number of each
tetra-, penta- and hexa-palindromic base sequence in pBR322.  PhiX174 and SV40
DNAs, which is presented here as an aid to other investigators.

<>

<1>Watson, R.J., Schildkraut, I., Qiang, B.-Q., Martin, S.M., Visentin, L.P.
<2>NdeI: a restriction endonuclease from Neisseria denitrificans which cleaves DNA at 5'-CATATG-3' sequences.
<3>FEBS Lett.
<4>150
<5>114-116
<6>1982
<7>Type II restriction endonucleases recognize and cleave near specific DNA
sequences, usually 4-6 base pair palindromes.  They are of fundamental
importance as tools for the recombinant DNA technology as they provide the
selective cleavages required for the analysis and restructuring of DNA in
vitro.  The flexibility of these techniques is proportional to the number of
DNA sequences which can be cleaved by available enzymes.  Here, we describe the
isolation and characterization of a restriction enzyme from Neisseria
denitrificans with a new recognition sequence, 5'-CATATG-3'.

<>

<1>Waugh, D.S., Sauer, R.T.
<2>Single amino acid substitutions uncouple the DNA binding and strand scission activities of Fok I endonuclease.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>9596-9600
<6>1993
<7>Single alanine substitution mutations at Asp-450 or Asp-467 of the type IIS restriction enzyme
FokI have no effect on the ability of the enzyme to bind strongly and selectively to its
recognition site but completely eliminate its ability to cleave either strand of substrate
DNA. Since wild-type FokI shows no kinetic preference or required order of strand cleavage,
these results indicate that FokI, which evidently functions as a monomer, uses a single
catalytic center to cleave both strands of DNA. In this respect, FokI may resemble other
monomeric enzymes that cleave double-stranded DNA.

<>

<1>Waugh, D.S., Sauer, R.T.
<2>A novel class of FokI restriction endonuclease mutants that cleave hemi-methylated substrates.
<3>J. Biol. Chem.
<4>269
<5>12298-12303
<6>1994
<7>A genetic screen was used to identify amino acid substitutions that enable the FokI
restriction endonuclease to cleave DNA in cells that express the cognate methyltransferase
activity. Missense mutations that give rise to this phenotype were isolated at eight different
positions (G188K, P196S, T343I, S388N, S395F, E407K, E410K, D421N), clustered in two regions
of the polypeptide sequence of FokI. Two of the mutant endonucleases (P196S and D421N) were
purified to homogeneity and analyzed in detail. Both mutants cleave FokI target sites
(5'-GGATG-3') in a manner similar to the wild-type enzyme. Neither mutant cleaved
noncanonical sequences, but both efficiently cleaved DNA substrates containing hemi-methylated
FokI sites. This class of mutations has not been observed with other restriction enzymes.

<>

<1>Wawrik, C.B., Callaghan, A.V., Stamps, B.W., Wawrik, B.
<2>Genome Sequence of Youngiibacter fragilis, the Type Strain of the Genus Youngiibacter.
<3>Genome Announcements
<4>2
<5>e01183-13
<6>2014
<7>The genome of Youngiibacter fragilis, the type strain of the newly described genus
Youngiibacter, was sequenced. The genome consists of 3.996 Mb, with a G+C
content of 46.6 mol%. Y. fragilis originates from coal-bed methane-produced water
and may provide insight into the microbiological basis of biogas production in
coal beds.

<>

<1>Wayne, J., Holden, M., Xu, S.-Y.
<2>The Tsp45I restriction-modification system is plasmid-borne within its thermophilic host.
<3>Gene
<4>202
<5>83-88
<6>1997
<7>Thermus species YS45 harbors two small cryptic plasmids of 5.8 (pTsp45s) and approximately 12
kb (pTsp45I).  Plasmid pTsp45s has been entirely sequenced, revealing three significant ORFs.
In addition to a previously reported thermophilic plasmid-encoded replication protein, pTsp45s
contains two genes for the Tsp45I methyltransferase and restriction endonuclease (Tsp45I).
These two converging genes (tsp45IM and tsp45IR) overlap by 4 bp at their stop codons within
an XbaI site.  M.Tsp45I (413 aa, 47.0 kDa, recognizing 5'-GTSAC-3') is highly homologous to
other m6A-methyltransferases, especially M.EcaI (recognizing 5'-GGTNACC-3').  Tsp45I (332
aa, 37.4 kDa, cleaving 5'-/GTSAC-3') is not homologous to M.Tsp45I, or to other restriction
endonucleases.  Recombinant Tsp45I is stably produced in E. coli, and cleaves DNA at 65oC with
the same specificity as the native enzyme.  Therefore, the thermophilic Tsp45I
restriction-modification system is plasmid-borne within its native host.

<>

<1>Wayne, J., Xu, S.-Y.
<2>Method for cloning and producing the Tsp45I restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 5866422
<5>
<6>1999
<7>The present invention relates to recombinant DNA molecules encoding the Tsp45I gene (tsp45IR),
and the cognate M.Tsp45I gene (tsp45IM), from Thermus species YS45 introduced into E. coli as
well as expression of the recombinant Tsp45I restriction endonuclease in E. coli.

<>

<1>Wayne, J., Xu, S.-Y.
<2>Method for cloning and producing the Tsp45I restriction endonuclease in E. coli.
<3>International Patent Office
<4>WO 9921973
<5>
<6>1999
<7>Method for cloning and producing the Tsp45I restriction endonuclease in E. coli.

<>

<1>Webb, H., Kaszubska, W., Gumport, R.I.
<2>Overexpression of RsrI DNA methyltransferase in E. coli.
<3>FASEB J.
<4>4
<5>A1795
<6>1990
<7>RsrI DNA methyltransferase (M.RsrI) from Rhodobacter sphaeroides catalyzes
methylation of the same central adenine residue in the duplex recognition
sequence d(GAATTC) as does M.EcoRI.  M.RsrI has been purified to homogeneity
from Rhodobacter sphaeroides, and its gene cloned and sequenced (Kaszubska, W.
et al., Nucl. Acids Res., (1989) 17, 10403-101701.  M.RsrI has now been
overexpressed in E. coli and the purification improved.  A fragment of R.
sphaeroides chromosomal DNA, containing both the rsrIM and rsrIR genes was
inserted downstream of the T7 promoter in a pTZ18U vector.  M.RsrI was purified
to homogeneity from E. coli BL21(DE3)plysS cells with a 100-fold increase in
yield over that from R. sphaeroides.  The purification was simplified by the
substitution of a DNA-cellulose-sinefungin affinity column for the
hydroxylapatite and weak cation exchange chromatography steps.  Sinefungin, an
S-adenosylmethionine analogue, inhibits methyl group transfer by M.RsrI.  We
are presently further characterizing M.RsrI by determining its kinetic
constants and the number of methyl groups transferred per binding event.

<>

<1>Webb, J.L., King, G., Ternent, D., Titheradge, A.J.B., Murray, N.E.
<2>Restriction by EcoKI is enhanced by cooperative interactions between target sequences and is dependent on DEAD box motifs.
<3>EMBO J.
<4>15
<5>2003-2009
<6>1996
<7>One subunit of both type I and type III restriction and modification enzymes
contains motifs characteristic of DEAD box proteins, which implies that these enzymes may be
DNA helicases.  This subunit is essential for restriction, but not modification.  The current
model
for restriction by both types of enzyme postulates that DNA cutting is stimulated when two
enzyme
complexes bound to neighboring target sequences meet as the consequence of ATP-dependent
DNA translocation.  For type I enzymes, this model is supported by in vitro experiments, but
the
predicted cooperative interactions between targets have not been detected by assays that
monitor
restriction in vivo.  The experiments reported here clearly establish the required synergistic
effect
but, in contrast to earlier experiments, they use Escherichia coli K-12 strains deficient in
the
restriction alleviation function associated with the Rac prophage.  In bacteria with elevated
levels of
EcoKI the cooperative interactions are obscured, consistent with cooperation between free
enzyme
and that bound at target sites.  We have made changes in three of the motifs characteristic of
DEAD
box proteins, including motif III, which in RecG is implicated in the migration of Holliday
junctions.  Conservative changes in each of the three motifs impair restriction.

<>

<1>Webb, M., Taylor, I.A., Firman, K., Kneale, G.G.
<2>Probing the domain structure of the type IC DNA methyltransferase M.EcoR124I by limited proteolysis.
<3>J. Mol. Biol.
<4>250
<5>181-190
<6>1995
<7>Limited proteolysis has been used to probe the domain structure of the type I DNA
methyltransferase M.EcoR124I. Trypsin digestion of the methyltransferase generates two
fragments derived from the HsdS subunit, a 28 kDa N-terminal domain and a 19 kDa C-terminal
domain, leaving the HsdM subunit intact. Extensive digestion by chymotrypsin, however, removes
59 amino acid residues from the N terminus of the HsdM subunit to leave a 52 kDa C-terminal
domain. Binding of the cofactor S-adenosyl methionine has no appreciable effect on the rate of
cleavage, but binding of a 30 bp DNA duplex containing the cognate recognition sequence
confers almost total protection. Following trypsin cleavage of the methyltransferase, a stable
proteolytic product is produced which has been purified for biochemical characterization. The
trypsinized enzyme is shown to be a multimeric complex containing two intact HsdM subunits and
both fragments of the HsdS subunit, consistent with the circular model proposed for the
organization of domains in the specificity subunit in type IC methyltransferases. Gel
retardation studies show that the proteolysed enzyme still retains DNA binding activity, but
its specificity for the DNA recognition sequence is dramatically reduced.

<>

<1>Wegmann, U., O'Connell-Motherway, M., Zomer, A., Buist, G., Shearman, C., Canchaya, C., Ventura, M., Goesmann, A., Gasson, M.J., Kuipers, O.P., van Sinderen, D., Kok, J.
<2>Complete genome sequence of the prototype lactic acid bacterium Lactococcus lactis subsp. cremoris MG1363.
<3>J. Bacteriol.
<4>189
<5>3256-3270
<6>2007
<7>Lactococcus lactis is of great importance for the nutrition of hundreds of millions of people
worldwide. This paper describes the genome sequence of
Lactococcus lactis subsp. cremoris MG1363, the lactococcal strain most
intensively studied throughout the world. The 2,529,478-bp genome contains
81 pseudogenes and encodes 2,436 proteins. Of the 530 unique proteins, 47
belong to the COG (clusters of orthologous groups) functional category
"carbohydrate metabolism and transport," by far the largest category of
novel proteins in comparison with L. lactis subsp. lactis IL1403. Nearly
one-fifth of the 71 insertion elements are concentrated in a specific
56-kb region. This integration hot-spot region carries genes that are
typically associated with lactococcal plasmids and a repeat sequence
specifically found on plasmids and in the "lateral gene transfer hot spot"
in the genome of Streptococcus thermophilus. Although the parent of L.
lactis MG1363 was used to demonstrate lysogeny in Lactococcus, L. lactis
MG1363 carries four remnant/satellite phages and two apparently complete
prophages. The availability of the L. lactis MG1363 genome sequence will
reinforce its status as the prototype among lactic acid bacteria through
facilitation of further applied and fundamental research.

<>

<1>Wegmann, U., Overweg, K., Horn, N., Goesmann, A., Narbad, A., Gasson, M.J., Shearman, C.
<2>The complete genome sequence of Lactobacillus johnsonii FI9785, a competitive exclusion agent against pathogens in poultry.
<3>J. Bacteriol.
<4>191
<5>7142-7143
<6>2009
<7>Lactobacillus johnsonii is a member of the acidophilus group of lactobacilli. Because of their
probiotic properties including attachment
to epithelial cells, immunomodulation and competitive exclusion of
pathogens representatives of this group are being intensively studied.
Here we report the complete annotated genome sequence of Lactobacillus
johnsonii FI9785, a strain which prevents the colonisation of specific
pathogen-free chicks by Clostridium perfringens.

<>

<1>Wei, H., Therrien, C., Blanchard, A., Guan, S., Zhu, Z.
<2>The Fidelity Index provides a systematic quantitation of star activity of DNA restriction endonucleases.
<3>Nucleic Acids Res.
<4>36
<5>e50
<6>2008
<7>Restriction endonucleases are the basic tools of molecular biology. Many restriction
endonucleases show relaxed sequence recognition, called star
activity, as an inherent property under various digestion conditions
including the optimal ones. To quantify this property we propose the
concept of the Fidelity Index (FI), which is defined as the ratio of the
maximum enzyme amount showing no star activity to the minimum amount
needed for complete digestion at the cognate recognition site for any
particular restriction endonuclease. Fidelity indices for a large number
of restriction endonucleases are reported here. The effects of reaction
vessel, reaction volume, incubation mode, substrate differences, reaction
time, reaction temperature and additional glycerol, DMSO, ethanol and
Mn(2+) on the FI are also investigated. The FI provides a practical
guideline for the use of restriction endonucleases and defines a
fundamental property by which restriction endonucleases can be
characterized.

<>

<1>Wei, J., Goldberg, M.B., Burland, V., Venkatesan, M.M., Deng, W., Fournier, G., Mayhew, G.F., Plunkett, G. III, Rose, D.J., Darling, A., Mau, B., Perna, N.T., Payne, S.M., Runyen-Janecky, L.J., Zhou, S., Schwartz, D.C., Blattner, F.R.
<2>Complete genome sequence and comparative genomics of Shigella flexneri serotype 2a strain 2457T.
<3>Infect. Immun.
<4>71
<5>2775-2786
<6>2003
<7>We determined the complete genome sequence of Shigella flexneri serotype 2a strain 2457T
(4,599,354 bp). Shigella species cause >1 million deaths
per year from dysentery and diarrhea and have a lifestyle that is markedly
different from those of closely related bacteria, including Escherichia
coli. The genome exhibits the backbone and island mosaic structure of E.
coli pathogens, albeit with much less horizontally transferred DNA and
lacking 357 genes present in E. coli. The strain is distinctive in its
large complement of insertion sequences, with several genomic
rearrangements mediated by insertion sequences, 12 cryptic prophages, 372
pseudogenes, and 195 S. flexneri-specific genes. The 2457T genome was also
compared with that of a recently sequenced S. flexneri 2a strain, 301. Our
data are consistent with Shigella being phylogenetically indistinguishable
from E. coli. The S. flexneri-specific regions contain many genes that
could encode proteins with roles in virulence. Analysis of these will
reveal the genetic basis for aspects of this pathogenic organism's
distinctive lifestyle that have yet to be explained.

<>

<1>Wei, S., Guo, Z., Li, T., Zhang, T., Li, X., Zhou, Z., Li, Z., Liu, M., Luo, R., Bi, D., Chen, H., Zhou, R., Jin, H.
<2>Genome Sequence of Mycoplasma iowae Strain 695, an Unusual Pathogen Causing Deaths in Turkeys.
<3>J. Bacteriol.
<4>194
<5>547-548
<6>2012
<7>Mycoplasma iowae is associated mainly with reduced hatchability in turkeys and is well known
for the unusual ability of phenotypic variation in the
Mycoplasma surface components as well as a relative resistance to heat,
bile salts, and many antimicrobials. A subset of unique genes and a gene
cluster responsible for these characteristics could be identified from the
genome. Here, we report the first genome sequence of this species.

<>

<1>Wei, W., Gao, C., Xiong, Y., Zhang, Y., Liu, S., Pu, Y.
<2>A fluorescence method for detection of DNA and DNA methylation based on graphene oxide and restriction endonuclease HpaII.
<3>Talanta
<4>131
<5>342-347
<6>2015
<7>DNA methylation plays an important role in many biological events and is associated with
various diseases. Most traditional methods for detection of DNA methylation are based on the
complex and expensive bisulfite method. In this paper, we report a novel fluorescence method
to detect DNA and DNA methylation based on graphene oxide (GO) and restriction endonuclease
HpaII. The skillfully designed probe DNA labeled with 5-carboxyfluorescein (FAM) and optimized
GO concentration keep the probe/target DNA. still adsorbed on the GO. After the cleavage
action of HpaII the labeled FAM is released from the GO surface and its fluorescence recovers,
which could be used to detect DNA in the linear range of 50 pM-50 nM with a detection limit of
43 pM. DNA methylation induced by transmethylase (Mtase) or other chemical reagents prevents
HpaII from recognizing and cleaving the specific site; as a result, fluorescence cannot
recover. The fluorescence recovery efficiency is closely related to the DNA methylation level,
which can be used to detect DNA methylation by comparing it with the fluorescence in the
presence of intact target DNA. The method for detection of DNA and DNA methylation is simple,
reliable and accurate.

<>

<1>Wei, Y., Yang, Y., Zhou, L., Liu, Z., Wang, X., Yang, R., Su, Q., Zhou, Y., Zhao, J., Yang, J.
<2>Genome Sequence of Enterobacter cancerogenus YZ1.
<3>Genome Announcements
<4>1
<5>e00023-13
<6>2013
<7>is usually known as an opportunistic human pathogen. Recently, it has attracted great
attention for its capability to produce bioemulsifier, degrade xenobiotics,
and resist alkalis and antibiotics. Here we report the complete genome of YZ1,
isolated from a bran-feeding insect's frass.

<>

<1>Wei, Y., Zhou, H., Zhang, L., Zhang, J., Wang, Y., Wang, S., Zhou, Z., Yan, X.
<2>Draft Genome Sequence of Clostridium ultunense Strain BS (DSMZ 10521), Recovered  from a Mixed Culture.
<3>Genome Announcements
<4>2
<5>e01269-13
<6>2014
<7>Clostridium ultunense BS is the first isolated strain (type strain) of C. ultunense that was
identified as a mesophilic syntrophic acetate-oxidizing
bacterium (SAOB). Here, we report the draft genome sequence of this strain, which
will help us to elucidate the mechanism of syntrophic acetate oxidization.

<>

<1>Wei, Y.X., Zhang, Z.Y., Liu, C., Zhu, Y.Z., Zhu, Y.Q., Zheng, H.J., Zhao, G.P., Wang, S.Y., Guo, X.K.
<2>Complete Genome Sequence of Bifidobacterium longum JDM301.
<3>J. Bacteriol.
<4>192
<5>4076-4077
<6>2010
<7>Bifidobacteria, known as probiotic bacteria, are high G+C Gram-positive bacteria, which
naturally inhabit the human gastrointestinal tract (GIT) and vagina. Recently, we have a
Bifidobacterium longum JDM301 completely sequenced, which is a widely used Chinese commercial
strain with several probiotic properties.

<>

<1>Weidner, S., Baumgarth, B., Gottfert, M., Jaenicke, S., Puhler, A., Schneiker-Bekel, S., Serrania, J., Szczepanowski, R., Becker, A.
<2>Genome Sequence of Sinorhizobium meliloti Rm41.
<3>Genome Announcements
<4>1
<5>e00013-12
<6>2013
<7>Sinorhizobium meliloti Rm41 nodulates alfalfa plants, forming indeterminate type  nodules. It
is characterized by a strain-specific K-antigen able to replace exopolysaccharides in
promotion of nodule invasion. We present the Rm41 genome, composed of one chromosome, the
chromid pSymB, the megaplasmid pSymA, and the nonsymbiotic plasmid pRme41a.

<>

<1>Weidner, S., Becker, A., Bonilla, I., Jaenicke, S., Lloret, J., Margaret, I., Puhler, A., Ruiz-Sainz, J.E., Schneiker-Bekel, S., Szczepanowski, R., Vinardell, J.M., Zehner, S., Gottfert, M.
<2>Genome Sequence of the Soybean Symbiont Sinorhizobium fredii HH103.
<3>J. Bacteriol.
<4>194
<5>1617-1618
<6>2012
<7>Sinorhizobium fredii HH103 is a fast-growing rhizobial strain that is able to nodulate legumes
that develop determinate nodules, e.g., soybean, and legumes
that form nodules of the indeterminate type. Here we present the genome of HH103,
which consists of one chromosome and five plasmids with a total size of 7.22 Mb.

<>

<1>Weigand, M.R., Ryu, H., Bozcek, L., Konstantinidis, K.T., Santo, D.J.W.
<2>Draft Genome Sequence of Catellicoccus marimammalium, a Novel Species Commonly Found in Gull Feces.
<3>Genome Announcements
<4>1
<5>e00019-12
<6>2013
<7>Catellicoccus marimammalium is a relatively uncharacterized Gram-positive facultative anaerobe
with potential utility as an indicator of waterfowl fecal
contamination. Here, we report an annotated draft genome sequence that suggests
that this organism may be a symbiotic gut microbe.

<>

<1>Weil, M.D., McClelland, M.
<2>Enzymatic cleavage of a bacterial genome at a 10-base-pair recognition site.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>86
<5>51-55
<6>1989
<7>The circular genome of Staphylococcus aureus was cut into two fragments by a
simple enzymatic method that cleaves a 10-base-pair site.  The recognition
sequence, A-T-C-G-mA^T-C-G-mA-T, was created by the combined use of the
methylase M.ClaI (A-T-C-G-mA-T) and the restriction endonuclease DpnI
(G-mA^T-C).  This technique is insensitive to CpG methylation and in human DNA
is predicted to produce fragments that, on average, are greater than five
million base pairs.  The ability to create such long pieces of DNA should
facilitate mapping of large, complex chromosomes.

<>

<1>Weilharter, A., Mitter, B., Shin, M.V., Chain, P.S., Nowak, J., Sessitsch, A.
<2>Complete genome sequence of the plant growth-promoting endophyte Burkholderia phytofirmans strain PsJN.
<3>J. Bacteriol.
<4>193
<5>3383-3384
<6>2011
<7>Burkholderia phytofirmans PsJN(T) is able to efficiently colonize the rhizosphere, root and
above-ground plant tissues of a wide variety of
genetically unrelated plants such as potato, canola, maize and grapevine.
Strain PsJN shows strong plant growth-promoting effects and was reported
to enhance plant vigour and resistance to biotic and abiotic stresses.
Here, we report the genome sequence of this strain, which indicates the
presence of multiple traits relevant for endophytic colonization and plant
growth promotion.

<>

<1>Weiner, M., Costa, G.
<2>New methodologies for the directional and bidirectional cloning and expression of PCR-generated DNA fragments.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>94
<5>249
<6>1994
<7>We have optimized conditions for the efficient cloning of blunt-ended DNA fragments generated
by the polymerase chain reaction (PCR). In the bidirectional cloning procedure, the PCR
fragment is incubated with SrfI endonuclease-treated (recognition site: 5'GCCC^GGGC3')
plasmid DNA, SrfI endonuclease, and T4 DNA ligase. The ligation reaction is allowed to proceed
for 1 hour and transformed into E. coli. The presence of the endonuclease in the ligation
reaction reduces nonrecombinant background and increases the amount of linear vector available
for recombinant insertion. Directional PCR cloning is achieved by creating a
monophosphorylated vector and a monophosphorylated PCR fragment. Circularization of the cloned
product during subsequent incubation with SrfI and T4 DNA ligase occurs only when the
insertion is in the desired orientation. The plasmid vectors have been optimized for
expression using T7 RNA polymerase for transcriptional control.

<>

<1>Weinhold, E., Dalhoff, C., Klimasauskas, S., Lukinavicius, G.
<2>Preparation of S-adenosyl-L-methionine analogs with extended activated groups for transfer by DNA methyltransferases.
<3>International Patent Office
<4>WO 2006108678 A
<5>
<6>2006
<7>S-Adenosyl-L-methionine analogs of formula (I) wherein R comprises a carbon-carbon double
bond, carbon-oxygen double bond, carbon-sulfur double bond, carbon-nitrogen double bond, a
carbon-carbon triple bond, carbon-nitrogen triple bond or an aromatic carbocyclic or
heterocyclic system in b-position to the sulfonium center, X- is an organic or inorganic anion
carrying one or more negative charges, Z is -CR1R2-, -O-, -S- or -NR3- and R1, R3 are
independently selected from H, D and C1-C12 alkyl.

<>

<1>Weinhold, E., Dalhoff, C., Klimasauskas, S., Lukinavicius, G.
<2>S-adenosyl-L-methionine analogs with extended activated groups for transfer by methyltransferases.
<3>US Patent Office
<4>US 08008007
<5>
<6>2011
<7>S-Adenosyl-L-methionine analogs of formula (I): are disclosed wherein R comprises a
carbon-carbon double bond, carbon-sulfur double bond,
carbon-nitrogen double bond, -a carbon-carbon triple bond,
carbon-nitrogen triple bond or an aromatic carbocyclic or heterocyclic
system in beta-position to the sulfonium center, X{circle around (-)}
is an organic or inorganic anion carrying one or more negative charges,
Z is -(CRR2)-R-1-, -O-, -S- or -NR3- and R-1, R(2 )and R(3 )are
independently selected from H, D and C-1-C alkyl; as well as complexes
with a methyltransferase, pharmaceutical compositions, methods for
modifying a biomolecule, and methods for detecting sequences specific
methylation of biomolecules.

<>

<1>Weinhold, E., Meier, T., Duefel, H., Markert-Hahn, C., Schmuck, R.
<2>N-adenosylaziridine derivatives as cofactor of S-adenosyl-L-methionine-dependent methyltransferases for sequence-specific detection of nucleic acid or protein methylation.
<3>International Patent Office
<4>WO 2005121361 A
<5>
<6>2005
<7>The present invention relates to a method for detecting sequence specific methylation in a
biomolecule, comprising: (a) contacting a biomolecule with an
S-adenosyl-L-methionine-dependent methyltransferase in the presence of a detectable cofactor
of said methyltransferase; and (b) detecting whether the recognition sequence of said
methyltransferase has been modified with the cofactor or a derivative thereof, wherein
modification of the recognition sequence of said methyltransferase is indicative of an absence
of methylation at said recognition sequence.  The present invention also relates to a cofactor
which is specific for S-adenosyl-L-methionine-dependent methyltransferases, wherein said
cofactor is an N-adenosylaziridine derivative with a reporter group attached to the 6 or 7
position of the adenine ring or attached to the aziridine ring.  Moreover, the present
invention relates to a complex of the cofactor of the present invention and a
methyltransferase which normally uses S-adenosyl-L-methionine as a cofactor.  In addition, the
present invention relates to a diagnostic composition comprising the cofactor of the present
invention or the complex of the present invention. Finally, the present invention relates to
the use of the cofactor of the present invention or the complex of the present invention for
detecting sequence-specific methylation in DNA molecules.

<>

<1>Weinmaier, T., Riesing, M., Rattei, T., Bille, J., Arguedas-Villa, C., Stephan, R., Tasara, T.
<2>Complete Genome Sequence of Listeria monocytogenes LL195, a Serotype 4b Strain from the 1983-1987 Listeriosis Epidemic in Switzerland.
<3>Genome Announcements
<4>1
<5>e00152-12
<6>2013
<7>The complete genome sequence of Listeria monocytogenes LL195, a serotype 4b clinical strain
isolated during the 1983-1987 listeriosis epidemic in
Switzerland, is presented.

<>

<1>Weinstock, G.M., Robinson, G.E.
<2>Insights into social insects from the genome of the honeybee Apis mellifera.
<3>Nature
<4>443
<5>931-949
<6>2006
<7>Here we report the genome sequence of the honeybee Apis mellifera, a key model for social
behaviour and essential to global ecology through pollination.  Compared with other sequenced
insect genomes,the A. mellifera genome has a high A + T and CpG contents, lacks major
transposon families, evolves more slowly, and is more similar to vertebrates for circadian
rhythm, RNA interference and DNA methylation genes, among others.  Furthermore, A. mellifera
has fewer genes for innate immunity, detoxification enzymes, cuticle-forming proteins and
gustatory receptors, more genes for odorant receptors, and novel genes for nectar and pollen
utilization, consistent with its ecology and social organization.  Copmpared to Drosophila,
genes in early developmental pathways differ in Apis, whereas similarities exist for functions
that differ markedly, such as sex determination, brain function and behaviour.  Population
genetics suggests a novel African origin for the species A. mellifera nad insights into
whether Africanized bees spread throughout the New World via hybridization or displacement.

<>

<1>Weinstock, K.G., Deloughery, C., Bush, D.
<2>Nucleic acid and amino acid sequences relating to Enterobacter cloacae for diagnostics and therapeutics.
<3>US Patent Office
<4>US 7041814 B
<5>
<6>2006
<7>The invention provides isolated polypeptide and nucleic acid sequences derived from
Enterobacter cloacae that are useful in diagnosis and therapy of pathological conditions;
antibodies against the polypeptides; and methods for the production of the polypeptides.  The
invention also provides methods for the detection, prevention and treatment of pathological
conditions resulting from bacterial infection.

<>

<1>Weisenberger, D.J., Velicescu, M., Cheng, J.C., Gonzales, F.A., Liang, G.N., Jones, P.A.
<2>Role of the DNA methyltransferase variant DNMT3b3 in DNA methylation.
<3>Mol. Cancer Res.
<4>2
<5>62-72
<6>2004
<7>Several alternatively spliced variants of DNA methyltransferase (DNMT) 3b have been described.
Here, we identified new murine Dnmt3b mRNA
isoforms and found that mouse embryonic stem (ES) cells expressed only
Dnmt3b transcripts that contained exons 10 and 11, whereas the Dnmt3b
transcripts in somatic cells lacked these exons, suggesting that this
region is important for embryonic development. DNMT3b2 and 3b3 were the
major isoforms expressed in human cell lines and the mRNA levels of
these isoforms closely correlated with their protein levels. Although
DNMT3b3 may be catalytically inactive, it still may be biologically
important because D4Z4 and satellites 2 and 3 repeat sequences, all
known DNMT3b target sequences, were methylated in cells that
predominantly expressed DNMT3b3. Treatment of cells with the
mechanism-based inhibitor 5-aza-2'-deoxycytidine (5-Aza-CdR) caused a
complete depletion of DNMT1, 3a, 3b1, and 3b2 proteins. Human DNMT3b3
and the murine Dnmt3b3-like isoform, Dnmt3b6, were also depleted
although less efficiently, suggesting that DNMT3b3 also may be capable
of DNA binding. Moreover, de novo methylation of D4Z4 in T24 cancer
cells after 5-Aza-CdR treatment only occurred when DNMT3b3 was
expressed, reinforcing its role as a contributing factor of DNA
methylation. The expression of either DNMT3b2 or 3b3, however, was not
sufficient to explain the abnormal methylation of DNMT3b target
sequences in human cancers, which may therefore be dependent on factors
that affect DNMT3b targeting. Methylation analyses of immunodeficiency,
chromosomal instabilities, and facial abnormalities cells revealed that
an Alu repeat sequence was highly methylated, suggesting that Alu
sequences are not DNMT3b targets.

<>

<1>Weisenberger, D.J., Velicescu, M., Preciado-Lopez, M.A., Gonzales, F.A., Tsai, Y.C., Liang, G., Jones, P.A.
<2>Identification and characterization of alternatively spliced variants of DNA methyltransferase 3a in mammalian cells.
<3>Gene
<4>298
<5>91-99
<6>2002
<7>CpG methylation is mediated by the functions of at least three active DNA methyltransferases
(DNMTs). While DNMT1 is thought to perform maintenance methylation, the more recently
discovered DNMT3a and DNMT3b enzymes are thought to facilitate de novo methylation. Murine
Dnmt3a and 3b are developmentally regulated and a new Dnmt3a isoform, Dnmt3a2, has been
recently shown to be expressed preferentially in mouse embryonic stem (ES) cells. Here we have
characterized four alternatively spliced variants of human and mouse DNMT3a. These transcripts
included a novel exon 1 (1beta) that was spliced into the same exon 2 acceptor splice site
used by the original exon 1 (1alpha). Cloning and sequencing of the 5' region of the human
DNMT3a gene revealed that exon 1beta was situated upstream of exon 1alpha and that the entire
region was contained within a CpG island. We also identified other alternatively spliced
species containing intron 4 inclusions that were associated with either exon 1alpha or 1beta.
These were expressed at low levels in mouse and human cells. All transcripts were highly
conserved between human and mouse. The levels of Dnmt3a mRNA containing exon 1beta were
3-25-fold greater in mouse ES cells than in various somatic cells as determined by
semiquantitative reverse transcription-polymerase chain reaction analysis, while the levels of
exon 1alpha-containing transcripts were slightly higher in human and mouse somatic cells. The
preferential expression of the beta transcript in ES cells suggests that this transcript, in
addition to Dnmt3a2, may also be important for de novo methylation during development.

<>

<1>Weiserova, M., Dutta, C.F., Firman, K.
<2>A novel mutant of the type I restriction-modification enzyme EcoR124I is altered at a key stage of the subunit assembly pathway.
<3>J. Mol. Biol.
<4>304
<5>301-310
<6>2000
<7>The HsdS subunit of a type I restriction-modification (R-M) system plays an essential role in
the activity of both the modification methylase and the restriction endonuclease. This subunit
is responsible for DNA binding, but also contains conserved amino acid sequences responsible
for protein-protein interactions. The most important protein-protein interactions are those
between the HsdS subunit and the HsdM (methylation) subunit that result in assembly of an
independent methylase (MTase) of stoichiometry M(2)S(1). Here, we analysed the impact on the
restriction and modification activities of the change Trp(212)-->Arg in the distal border of
the central conserved region of the EcoR124I HsdS subunit. We demonstrate that this point
mutation significantly influences the ability of the mutant HsdS subunit to assemble with the
HsdM subunit to produce a functional MTase. As a consequence of this, the mutant MTase has
drastically reduced DNA binding, which is restored only when the HsdR (restriction) subunit
binds with the MTase. Therefore, HsdR acts as a chaperon allowing not only binding of the
enzyme to DNA, but also restoring the methylation activity and, at sufficiently high
concentrations in vitro of HsdR, restoring restriction activity.

<>

<1>Weiserova, M., Firman, K.
<2>Isolation of a non-classical mutant of the DNA recognition subunit of the type I restriction endonuclease R.EcoR124I.
<3>Biol. Chem.
<4>379
<5>585-589
<6>1998
<7>We have used deletion mutagenesis and PCR-based misincorporation mutagenesis to produce a
collection of mutations in the central conserved region of the DNA binding subunit of the type
IC restriction endonuclease EcoR124I.  It has been proposed that this domain is involved in
protein-protein interactions during the assembly of the endonuclease.  While a large
percentage of these mutations gave a classical Res- Mod- phenotype, one mutant was isolated
with a nonclassical Res-Mod+ phenotype.  The loss of restriction activity, but retention of
the ability to modify indicates that this mutation cannot affect DNA binding and must alter
the assembly of the endonuclease in such a way as to prevent DNA cleavage but allow
methylation.  This mutant resulted from a single amino acid change Trp212-Arg.  The location
of the single amino acid change is at the border of the central conserved region and the
second target recognition domain and suggests that this region is extremely important for the
assembly of the methylase with the HsdR subunit into a functional restriction endonuclease.

<>

<1>Weiserova, M., Janscak, P., Benada, O., Hubacek, J., Vitaly, E.A., Glover, S.W., Firman, K.
<2>Cloning, production and characterization of wild type and mutant forms of the R.EcoK endonucleases.
<3>Nucleic Acids Res.
<4>21
<5>373-379
<6>1993
<7>The hsdR, hsdM and hsdS genes coding for R.EcoK restriction endonucleases, both with and
without a temperature sensitive mutation (ts-1) in the hsdS gene, were cloned in pBR322
plasmid and introduced into E. coli C3-6. The presence of the hsdS ts-1 mutation has no effect
on the R-M phenotype of this contruct in bacteria grown at 42 degrees C. However, DNA
sequencing indicates that the mutation is still present on the pBR322-hsd ts1 operon. The
putative temperature-sensitive endonuclease was purified from bacteria carrying this plasmid
and the ability to cleave and methylate plasmid DNA was investigated. The mutant endonuclease
was found to show temperature-sensitivity for restriction. Modification was dramatically
reduced at both the permissive and non-permissive temperatures. The wild type enzyme was found
to cleave circular DNA in a manner which strongly suggests that only one endonuclease molecule
is required per cleavage event. Circular and linear DNA appear to be cleaved using different
mechanisms, and cleavage of linear DNA may require a second endonuclease molecule. The subunit
composition of the purified endonucleases was investigated and compared to the level of
subunit production in minicells. There is no evidence that HsdR is prevented from assembling
with HsdM and HsdS ts-1 to produce the mutant endonuclease. The data also suggests that the
level of HsdR subunit may be limiting within the cell. We suggest that an excess of HsdM and
HsdS may produce the methylase in vivo and that assembly of the endonuclease may be dependent
upon the prior production of this methylase.

<>

<1>Weiserova, M., Janscak, P., Zinkevich, V., Hubacek, J.
<2>Overproduction of the Hsd subunits leads to the loss of temperature-sensitive restriction and modification phenotype.
<3>Folia Microbiol. (Praha)
<4>39
<5>452-458
<6>1994
<7>The genes hsdM and hsdS for M.EcoKI modification methyltransferase and the complete set of
hsdR, hsdM and hsdS genes coding for R.EcoKI restriction endonuclease, both with and without a
temperature-sensitive (ts) mutation in hsdS gene, were cloned in pBR322 plasmid and introduced
into E. coli C (a strain without a natural restriction-modification (R-M) system).  The
strains producing only the methyltransferase, or together with the endonuclease, were thus
obtained.  The hsdSts-1 mutation, mapped previously in the distal variable region of the hsdS
gene with C1245-T transition has no effect on the R-M phenotype expressed from cloned genes in
bacteria grown at 42oC.  In clones transformed with the whole hsd region an alleviation of R-M
functions was observed immediately after the transformation, but after subculture the
transformants expressed the wild-type R-M phenotype irrespective of whether the wild-type or
the mutant hsdS allele was present in the hybrid plasmid.  Simultaneous overproduction of HsdS
and HsdM subunits impairs the ts effect of the hsdSts-1 mutation on restriction and
modification.

<>

<1>Weiserova, M., Ryu, J.C.
<2>Characterization of a restriction modification system from the commensal Escherichia coli strain A0 34/86 (O83 : K24 : H31).
<3>BMC Microbiol.
<4>8
<5>106
<6>2008
<7>Background: Type I restriction-modification (R-M) systems are the most complex restriction
enzymes discovered to date. Recent years have
witnessed a renaissance of interest in R-M enzymes Type I. The massive
ongoing sequencing programmes leading to discovery of, so far, more
than 1 000 putative enzymes in a broad range of microorganisms
including pathogenic bacteria, revealed that these enzymes are widely
represented in nature. The aim of this study was characterisation of a
putative R-M system EcoA0ORF42P identified in the commensal Escherichia
coli A0 34/86 (O83: K24: H31) strain, which is efficiently used at
Czech paediatric clinics for prophylaxis and treatment of nosocomial
infections and diarrhoea of preterm and newborn infants.
Results: We have characterised a restriction-modification system
EcoA0ORF42P of the commensal Escherichia coli strain A0 34/86 ( O83:
K24: H31). This system, designated as EcoAO831, is a new functional
member of the Type IB family, whose specificity differs from those of
known Type IB enzymes, as was demonstrated by an immunological
cross-reactivity and a complementation assay. Using the plasmid
transformation method and the RM search computer program, we identified
the DNA recognition sequence of the EcoAO83I as GGA(8N)ATGC. In
consistence with the amino acids alignment data, the 3' TRD component
of the recognition sequence is identical to the sequence recognized by
the EcoEI enzyme. The A-T ( modified adenine) distance is identical to
that in the EcoAI and EcoEI recognition sites, which also indicates
that this system is a Type IB member. Interestingly, the recognition
sequence we determined here is identical to the previously reported
prototype sequence for Eco377I and its isoschizomers.
Conclusion: Putative restriction-modification system EcoA0ORF42P in
the commensal Escherichia coli strain A0 34/86 ( O83: K24: H31) was
found to be a member of the Type IB family and was designated as
EcoAO83I. Combination of the classical biochemical and bacterial
genetics approaches with comparative genomics might contribute
effectively to further classification of many other putative Type-I
enzymes, especially in clinical samples.

<>

<1>Weiss, A., Keshet, I., Razin, A., Cedar, H.
<2>DNA demethylation in vitro: Involvement of RNA.
<3>Cell
<4>86
<5>709-718
<6>1996
<7>An in vitro system for studying DNA demethylation has been established using extracts from
tissue culture cells.  This reaction, which is unusually resistant to proteinase K, takes
place through the removal of a 5-methylcytosine nucleotide unit from the DNA substrate and its
conversion to an RNAse-sensitive form.  It is likely that this represents the in vivo
mechanism as well, since extracts from L8 myoblasts specifically demethylate an alpha-actin
gene, while extracts from F9 teratocarcinoma cells specifically demodify the Aprt CpG island.
After pretreatment with proteinase K, these extracts demethylate both genes equally,
suggesting that gene specificity may be controlled by protein factors.

<>

<1>Weiss, J.B.
<2>Neisseria gonorrhoeae-specific oligonucleotides.
<3>European Patent Office
<4>EP 0872561 A
<5>
<6>1997
<7>The present invention relates to oligonucleotides which hybridize specifically to the cytosine
DNA methyltransferase gene of Neisseria gonorrhoeae and distinguish the cytosine DNA
methyltransferase gene of N. gonorrhoeae from highly homologous sequences which have been
discovered in some strains of other species of the genus Neisseria.  The oligonucleotides are
useful as primers for the polymerase chain reaction amplification of a nucleic acid sequence
from the cytosine DNA methyltransferase gene of N. gonorrhoeae and in N. gonorrhoeae
amplification/detection assays.  The invention has applications in the detection of Neisseria
gonorrhoeae, the field of medical diagnostics generally, and the field of molecular biology.

<>

<1>Weiss, J.B.
<2>Neisseria gonorrhoeae-specific oligonucleotides.
<3>US Patent Office
<4>US 6090557
<5>
<6>2000
<7>The present invention relates to oligonucleotides which hybridize specifically to the cytosine
DNA methyltransferase gene of Neisseria gonorrhoeae and distinguish the cytosine DNA
methyltransferase gene of N. gonorrhoeae from highly homologous sequences which have been
discovered in some strains of other species of the genus Neisseria.  The oligonucleotides are
useful as primers for the polymerase chain reaction (PCR) amplification of a nucleic acid
sequence from the cytosine DNA methyltransferase gene of N. gonorrhoeae and in N. gonorrhoeae
amplification/detection assays.

<>

<1>Weiss, M., Kesberg, A.I., Labutti, K.M., Pitluck, S., Bruce, D., Hauser, L., Copeland, A., Woyke, T., Lowry, S., Lucas, S., Land, M., Goodwin, L., Kjelleberg, S., Cook, A.M., Buhmann, M., Thomas, T., Schleheck, D.
<2>Permanent draft genome sequence of Comamonas testosteroni KF-1.
<3>Standards in Genomic Sciences
<4>8
<5>239-254
<6>2013
<7>Comamonas testosteroni KF-1 is a model organism for the elucidation of the novel  biochemical
degradation pathways for xenobiotic 4-sulfophenylcarboxylates (SPC)
formed during biodegradation of synthetic 4-sulfophenylalkane surfactants (linear
alkylbenzenesulfonates, LAS) by bacterial communities. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. The 6,026,527 bp long chromosome (one sequencing gap) exhibits an
average G+C content of 61.79% and is predicted to encode 5,492 protein-coding
genes and 114 RNA genes.

<>

<1>Weiss, V.A., Faoro, H., Tadra-Sfeir, M.Z., Raittz, R.T., de Souza, E.M., Monteiro, R.A., Cardoso, R.L., Wassem, R., Chubatsu, L.S., Huergo, L.F., Muller-Santos, M., Steffens, M.B., Rigo, L.U., Pedrosa, F.O., Cruz, L.M.
<2>Draft Genome Sequence of Herbaspirillum lusitanum P6-12, an Endophyte Isolated from Root Nodules of Phaseolus vulgaris.
<3>J. Bacteriol.
<4>194
<5>4136-4137
<6>2012
<7>Herbaspirillum lusitanum strain P6-12 (DSM 17154) is, so far, the only species of
Herbaspirillum isolated from plant root nodules. Here we report a draft genome
sequence of this organism.

<>

<1>Weissgerber, T., Zigann, R., Bruce, D., Chang, Y.-j., Detter, J.C., Han, C., Hauser, L., Jeffries, C.D., Land, M., Munk, A.C., Tapia, R., Dahl, C.
<2>Complete genome sequence of Allochromatium vinosum DSM 180.
<3>Standards in Genomic Sciences
<4>5
<5>311-330
<6>2011
<7>Allochromatium vinosum formerly Chromatium vinosum is a mesophilic purple sulfur bacte-rium
belonging to the family Chromatiaceae in the bacterial class Gammaproteobacteria. The genus
Allochromatium contains currently five species. All members were isolated from fresh-water,
brackish water or marine habitats and are predominately obligate phototrophs. Here we describe
the features of the organism, together with the complete genome sequence and annotation. This
is the first completed genome sequence of a member of the Chromatiaceae within the purple
sulfur bacteria thriving in globally occurring habitats. The 3,669,074 bp ge-nome with its
3,302 protein-coding and 64 RNA genes was sequenced within the Joint Ge-nome Institute
Community Sequencing Program.

<>

<1>Welch, R.A. et al.
<2>Extensive mosaic structure revealed by the complete genome sequence of uropathogenic Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>99
<5>17020-17024
<6>2002
<7>We present the complete genome sequence of uropathogenic Escherichia coli, strain CFT073. A
three-way genome comparison of the CFT073, enterohemorrhagic E. coli EDL933, and laboratory
strain MG1655 reveals that, amazingly, only 39.2% of their combined (nonredundant) set of
proteins actually are common to all three strains. The pathogen genomes are as different from
each other as each pathogen is from the benign strain. The difference in disease potential
between O157:H7 and CFT073 is reflected in the absence of genes for type III secretion system
or phage- and plasmid-encoded toxins found in some classes of diarrheagenic E. coli. The
CFT073 genome is particularly rich in genes that encode potential fimbrial adhesins,
autotransporters, iron-sequestration systems, and phase-switch recombinases. Striking
differences exist between the large pathogenicity islands of CFT073 and two other well-studied
uropathogenic E. coli strains, J96 and 536. Comparisons indicate that extraintestinal
pathogenic E. coli arose independently from multiple clonal lineages. The different E. coli
pathotypes have maintained a remarkable synteny of common, vertically evolved genes, whereas
many islands interrupting this common backbone have been acquired by different horizontal
transfer events in each strain.

<>

<1>Welch, S.G., Al-Awadhi, S., Williams, R.A.D.
<2>Isoschizomers of the restriction endonuclease TaqI (T/CGA) requiring different metal ion concentrations and having a range of thermal stabilities from Thermus species from different continents.
<3>Microbiology
<4>144
<5>167-175
<6>1998
<7>One-hundred-and-fifty-two isolates of the genus Thermus, collected from hot springs on four
continents, were screened for evidence of the presence of the thermophilic Type II restriction
endonuclease TaqI (T/CGA).  The presence of isoschizomers of TaqI in 27 of the isolates,
originating from hot springs in New Zealand, Iceland, USA, Japan, mainland Portugal and the
island of Sao Miguel in the Azores, is reported.  Six of the TaqI-containing isolates from
diverse geographical locations, identified by means of DNA/DNA homology and 16S rRNA sequence
alignment as belonging to the Thermus species T. aquaticus, T. filiformis, T. themophilus, T.
scotoductus and T. brockianus, were selected for comparative studies.  The TaqI isoschizomers
from each of the six isolates were partially purified.  They differed in their magnesium ion
requirements, isoelectric points, subunit molecular masses and thermal stability.

<>

<1>Welch, S.G., Williams, R.A.D.
<2>Two thermostable type II restriction endonucleases from Icelandic strains of the genus Thermus: Tsp4C I (ACN/GT), a novel type II restriction endonuclease, and Tsp8E I, an isoschizomer of the mesophilic enzyme BglI (GCCNNNN/NGGC).
<3>Biochem. J.
<4>309
<5>595-599
<6>1995
<7>Sixteen isolates of thermophilic bacteria from the genus Thermus, isolated from neutral and
alkaline hot water springs in the southwest region of Iceland, were tested for the presence of
restriction endonucleases.  Extracts from five of the isolates showed evidence of the
presence of restriction endonuclease activity by producing discrete nucleotide fragments when
incubated at 65oC with lambda phage DNA.  Two of the isolates (Tsp4C and Tsp8E) were found to
have particularly high levels of restriction endonuclease activity, and the respective enzymes
from these two Thermus isolates were partially purified and characterized and their
recognition and cleavage sites were determined.  Enzyme Tsp4C I is a novel type II restriction
endonuclease recognizing the interrupted palindromic tetranucleotide sequence ACNGT, where N
can be any one of the four bases in DNA.  Tsp4CI, which retains full enzyme activity when
incubated for 10 min at temperatures up to 76oC, hydrolyses the phosphodiester bond in both
strands of a double-stranded DNA substrate between the third and fourth bases of the
recognition sequence (ACN/GT), generating fragments with a single base 3'-OH overhang.
Enzyme Tsp8EI is a thermostable isoschizomer of the mesophilic type II restriction
endonuclease BglI (GCCNNNN/NGGC), generating fragments with a three base 3'-OH overhang.
However, unlike BglI, Tsp8EI exhibits considerable thermal stability, retaining full enzyme
activity when incubated for 10 min at temperatures up to 78oC.  Both Tsp4CI and Tsp8EI
represent significant additions to the small but expanding list of the extremely thermostable
restriction endonucleases.

<>

<1>Welch, S.G., Williams, R.A.D.
<2>Taq52I, a novel and thermostable type II restriction endonuclease from the genus Thermus, recognising the pentanucleotide sequence GC(A or T)GC and cleaving DNA between the first and second bases of the recognition sequence: G/C(A or T)GC.
<3>Nucleic Acids Res.
<4>23
<5>4573-4575
<6>1995
<7>127 isolates of the genus Thermus, from neutral and alkaline hot water springs on four
continents, have been screened for the presence of restriction endonuclease activity.  An
isolate (YS52) from Yellowstone National Park, USA, showed a high level of restriction
endonuclease activity when a cell free extract was incubated with lambda phage DNA at 65oC.  A
type II restriction endonuclease (Taq52I) has been partially purified from this isolate and
the recognition and cleavage site determined.  Taq52I has a novel interrupted palindromic
tetranucleotide recognition sequence GCWGC, where W can be either adenine (A) or thymine (T).
It hydrolyses the phosphodiester bond in both strands of the substrate between the first and
second bases of the recognition sequence: 5'G/CWGC3', producing three-base 5'-OH overhangs
(sticky ends).  The enzyme has a pH optimum of 7.0, requires 8 mM MgCl2 for maximum activity
and is thermally stable, retaining full enzyme activity following incubation at 79oC for 10
min.  Taq52I not only represents a new addition to the type II restriction endonucleases, but
also increases the small list of thermostable restriction endonucleases.

<>

<1>Welch, S.G., Williams, R.A.D.
<2>Two different isoschizomers of the type-II restriction endonuclease TaqI (T/CGA) within the same Thermus isolate: Tsp32I, an enzyme with similar heat stability properties to the prototype enzyme TaqI, & Tsp32II, a hyperthermostable isoschizomer of TaqI.
<3>Biochem. J.
<4>312
<5>505-510
<6>1995
<7>We have recently screened 112 separate isolates of the genus Thermus, collected from neutral
and alkaline hot water springs on four continents, for the presence of the Type-II restriction
endonuclease TaqI (T/CGA).  One particular isolate from the Azores (strain 32) was found to
contain high levels of a restriction endonuclease with the same recognition and cleavage site
as TaqI.  Initial studies revealed that the partially purified enzyme from strain 32 was
considerably more resistant to heat inactivation than the prototype enzyme TaqI, being able to
withstand temperatures at least 10oC higher than TaqI, before showing evidence of heat
inactivation.  Subsequently it became clear that the partially purified extract from strain 32
contains two separate enzymes, both of which are isoschizomers of TaqI.  One of the enzymes,
Tsp32I, has similar thermal stability characteristics to TaqI, whereas the second TaqI
isoschizomer, Tsp32II, found in the same Thermus isolate as Tsp32I, is considerably more
thermostable than TaqI, retaining full enzyme activity up to a temperature of 85oC.  Tsp32I
and Tsp32II were further distinguished by virtue of their different requirements for magnesium
ions.

<>

<1>Welch, S.G., Williams, R.A.D.
<2>Tsp49I (ACGT^), a thermostable neoschizomer of the type II restriction endonuclease MaeII (A^CGT), discovered in isolates of the genus thermus from the Azores, Iceland and New Zealand.
<3>Nucleic Acids Res.
<4>24
<5>1799-1801
<6>1996
<7>One hundred and forty eight isolates of the genus Thermus, from neutral and
alkaline hot water springs on four continents, have been screened for the presence of
restriction
enodnuclease activity.  An isolate (SM49) from the island of Sao Miguel, in the Azores, showed
a
high level of restriction endonuclease activity when a cell-free extract was incubated with
lambda
phage DNA at 65oC.  A type II restriction endonuclease (Tsp49I) has been partially purified
from
this isolate and the recognition and cleavage site determined.  Tsp49I recognizes the four
base
sequence ACGT, which is the same as the recognition sequence of the mesophilic type II
restriction endonuclease MaeII.  However, unlike MaeII, which cleaves DNA between the first
and
second base of the recognition sequence (A^CGT), Tsp49I hydrolyses the phosphodiester bond in
both strands of the substrate after the last base of the recognition sequence 5'-ACGT^-3',
producing four base 3'-OH overhangs (sticky ends).  The enzyme has a pH optimum of 9.0,
requires 3 mM MgCl2 for maximum activity and retains full enzyme activity following incubation
for 10 min at temperatures up to 80oC.  Two further examples of the same restriction
endonuclease
specificity as Tsp49I were detected in Thermus isolates from Iceland (TspIDSI) and New Zealand
(TspWAM8AI).  The three MaeII neoschizomers, Tsp49I, TspIDSI and TspWAM8AI, exhibit
similar pH optima, heat stabilities and MgCl2 requirements, but differ in their requirements
for
NaCl and KCl.

<>

<1>Welch, T.J., Fricke, W.F., McDermott, P.F., White, D.G., Rosso, M.L., Rasko, D.A., Mammel, M.K., Eppinger, M., Rosovitz, M.J., Wagner, D., Rahalison, L., LeClerc, J.E., Hinshaw, J.M., Lindler, L.E., Cebula, T.A., Carniel, E., Ravel, J.
<2>Multiple Antimicrobial Resistance in Plague: An Emerging Public Health Risk.
<3>PLoS ONE
<4>2
<5>e309
<6>2007
<7>Antimicrobial resistance in Yersinia pestis is rare, yet constitutes a significant
international public health and biodefense threat.  In 1995, the first multidrug resistant
isolate of Y. pestis (strain IP275) was identified, and was shown to contain a
self-transmissible plasmid (pIP1202) that conferred resistance to many of the antimicrobials
recommended for plague treatment and prophylaxis.  Comparative analysis of the DNA sequence of
Y. pestis plasmid pIP1202 revealed a near identical IncA/C plasmid backbone that is shared by
MDR plasmids isolated from Salmonella enterica serotype Newport SL254 and the fish pathogen
Yersinia ruckeri YR71.  The high degree of sequence identity and gene synteny between the
plasmid backbones suggests recent acquisition of these plasmids from a common ancestor.  In
addition, the Y. pestis pIP1202-like plasmid backbone was detected in numerous MDR
enterobacterial pathogens iosolated from retail meat samples collected between 2002 and 2005
in the United States.  Plasmid-positive strains were isolated from beef, chicken, turkey and
pork, and were found in samples from the following states: California, Colorado, Connecticut,
Georgia, Maryland, Minnesota, New Mexico, New York and Oregon.  Our studies reveal that this
common plasmid backbone is broadly disseminated among MDR zoonotic pathogens associated with
agriculture.  This reservoir of mobile resistance determinants has the potential to
disseminate to Y. pestis and other human and zoonotic bacterial pathogens and therefore
represents a significant public health concern.

<>

<1>Weller, R.L., Rajski, S.R.
<2>DNA methyltransferase-moderated click chemistry.
<3>Org. Lett.
<4>7
<5>2141-2144
<6>2005
<7>Biological methylation plays a vital role in regulatory mechanisms of gene transcription.
Methylation of both promoter sequences within the
genome, as well as protein substrates, has a profound impact upon gene
transcription. Yet, few tools exist by which to identify sites of
biological methylation in complex biological mixtures. We have
generated a novel adenosine-derived N-mustard that serves as an
efficient synthetic cofactor and allows for subsequent "click"
chemistry involving the modified nucleic acid substrate.

<>

<1>Weller, R.L., Rajski, S.R.
<2>Design, synthesis, and preliminary biological evaluation of a DNA methyltransferase-directed alkylating agent.
<3>Chembiochem
<4>7
<5>243-245
<6>2006
<7>DNA-binding and -damaging agents often derive their sequence selectivity from a finite set of
specific contacts between the molecule of interest and select nucleotides.  Many such agents
are of great interest due to their medicinally useful properties, although for many of these
substances, it remains unclear that all relevant mechanisms of action have been fully
elucidated.  In the context of DNA damage however, it seems that damage at certain sites is
likely to have a greater impact than at others.  At the heart of DNA sequence selectivity is
the resounding influence of a limited set of interactions between target nucleotides and the
small molecule of interest.  We hypothesized that DNA alkylation selectivity can be drived
from the extensive contacts between a sequence-specific DNA-modifying enzyme and its
recognition site.  Indeed, precedence for this strategy has been noted for DNA
methyltransferases.  However, to date, the structures of such MTase-dependent DNa-alkylating
agents have lacked the functionalities expected to be necessary for optimum activity, by
analogy to the natural cofactor S-adenosyl-L-methionine.

<>

<1>Weller-Stuart, T., Chan, W.Y., Coutinho, T.A., Venter, S.N., Smits, T.H., Duffy, B., Goszczynska, T., Cowan, D.A., de Maayer, P.
<2>Draft Genome Sequences of the Onion Center Rot Pathogen Pantoea ananatis PA4 and  Maize Brown Stalk Rot Pathogen P. ananatis BD442.
<3>Genome Announcements
<4>2
<5>e00750-14
<6>2014
<7>Pantoea ananatis is an emerging phytopathogen that infects a broad spectrum of plant hosts.
Here, we present the genomes of two South African isolates, P.
ananatis PA4, which causes center rot of onion, and BD442, isolated from brown
stalk rot of maize.

<>

<1>Wells, R.D., Klein, R.D., Singleton, C.K.
<2>Type II restriction enzymes.
<3>The Enzymes, Academic Press, Boyer, P., 
<4>14
<5>157-191
<6>1981
<7>The current status of type II restriction endonuclease enzymology is quite
unusual due to the history of this segment of molecular biology.  More than 210
restriction endonucleases have been purified, at least partially, and their
recognition sites identified.  However, the genetic and biochemical
characterization of these enzymes is meager.  Indeed, they have usually been
isolated as site-specific DNA endonucleases without genetically determining if
they are involved in DNA restriction-modification.  Moreover, the vast majority
of the restriction endonucleases are relatively crude preparations, and little
or nothing is known about the properties of the enzymes.

<>

<1>Wells, R.D., Neuendorf, S.K.
<2>Cleavage of single-stranded viral DNAs by certain restriction endonucleases.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>101-111
<6>1981
<7>*
   I. Introduction
  II. Characterization of the HaeIII reaction on O/X174, M13 and f1 DNAs
 III. Features of DNA structure recognized by the endonucleases
  IV. Characteristics of other restriction endonucleases
   V. Types of DNA substrates cleaved
  VI. Some properties of isoschizomers
 VII. Application of the technique for cleaving other DNAs
VIII. Reaction of DNA methylases on single-stranded DNA
  IX. Prospects for the future


<>

<1>Welsh, A.J., Halford, S.E., Scott, D.J.
<2>Analysis of Type II restriction endonucleases that interact with two recognition sites.
<3>Nucleic Acids Mol. Biol., Springer-Verlag, Pingoud, A., Berlin Heidelberg
<4>14
<5>297-317
<6>2004
<7>Many students of molecular biology know for certain that the Type II restriction endonucleases
are dimeric proteins that recognize a palindromic DNA sequence, 4-8 bp (base pairs) long, and
cut this single sequence with exquisite specificity.  This idea is also propagated by many
text-books and commonly-used laboratory manuals in molecular biology.  Like most
generalizations, it is in fact wrong.  The Type II restriction enzymes encompass not only
dimeric proteins but also monomers, tetramers and higher assemblies.  Their recognition
sequences are not always unique palindromes but can instead be degenerate, discontinuous or
asymmetric sequences.  The sites at which they cleave the DNA are not necessarily within the
recognition sequence.  Many cleave the DNA at fixed positions several bp away from their
recognition sequence.  Most excitingly, many of the Type II enzymes have to interact with two
copies of their recognition sites before they can cleave DNA.  When these enzymes bind two
sites in the same DNA molecule, the intervening DNA is sequestered in a loop.  DNA-looping
interactions play central roles in many genetic processes, such as replication, recombination
and transcription, but these processes are often difficult to study.  In contrast, the
reactions of restriction enzymes can be monitored by a variety of simple techniques, so these
enzymes make good tools for analyzing the nature of long-range interactions between distant
DNA sites.  Indeed, the restriction enzymes that act at two DNA sites have become one of the
principal paradigms for studying such interactions.

<>

<1>Welsh, E.A., Liberton, M., Stockel, J., Loh, T., Elvitigala, T., Wang, C., Wollam, A., Fulton, R.S., Clifton, S.W., Jacobs, J.M., Aurora, R., Ghosh, B.K., Sherman, L.A., Smith, R.D., Wilson, R.K., Pakrasi, H.B.
<2>The genome of Cyanothece 51142, a unicellular diazotrophic cyanobacterium important in the marine nitrogen cycle.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>105
<5>15094-15099
<6>2008
<7>Unicellular cyanobacteria have recently been recognized for their contributions to nitrogen
fixation in marine environments, a function
previously thought to be filled mainly by filamentous cyanobacteria such
as Trichodesmium. To begin a systems level analysis of the physiology of
the unicellular N(2)-fixing microbes, we have sequenced to completion the
genome of Cyanothece sp. ATCC 51142, the first such organism. Cyanothece
51142 performs oxygenic photosynthesis and nitrogen fixation, separating
these two incompatible processes temporally within the same cell, while
concomitantly accumulating metabolic products in inclusion bodies that are
later mobilized as part of a robust diurnal cycle. The 5,460,377-bp
Cyanothece 51142 genome has a unique arrangement of one large circular
chromosome, four small plasmids, and one linear chromosome, the first
report of a linear element in the genome of a photosynthetic bacterium. On
the 429,701-bp linear chromosome is a cluster of genes for enzymes
involved in pyruvate metabolism, suggesting an important role for the
linear chromosome in fermentative processes. The annotation of the genome
was significantly aided by simultaneous global proteomic studies of this
organism. Compared with other nitrogen-fixing cyanobacteria, Cyanothece
51142 contains the largest intact contiguous cluster of nitrogen
fixation-related genes. We discuss the implications of such an
organization on the regulation of nitrogen fixation. The genome sequence
provides important information regarding the ability of Cyanothece 51142
to accomplish metabolic compartmentalization and energy storage, as well
as how a unicellular bacterium balances multiple, often incompatible,
processes in a single cell.

<>

<1>Wen, T.N., Tung, P.H., Chen, C.S.
<2>Purification and some properties of a restriction endonuclease Ccy I from Clostridium cylindrosporum.
<3>Bot. Bull. Acad. Sinica
<4>30
<5>91-98
<6>1989
<7>The restriction endonuclease CcyI has been purified to homogeneity from an
anaerobic bacterium, Clostridium cylindrosporum, by the procedures involving
streptomycin sulfate precipitation, ammonium sulfate fractionation and
chromatography on phosphocellulose, heparin-Sepharose 4 B and Ultrogel AcA 34.
This enzyme recognizes the sequence
5'-^GATC-3'
3' -CTAG-5^' and cleaves the phosphodiester bond at the 5' side of G as
indicated.  Several isoschizomers of CcyI have been found previously which
include Sau3AI and MboI.  Optimal NaCl concentration, pH and temperature for
the enzyme to digest lambda DNA was 100 mM, 7.0-8.0 and 30-35C, respectively.
CcyI is both acid-labile and heat-labile.

<>

<1>Wende, W., Grindl, W., Christ, F., Pingoud, A., Pingoud, V.
<2>Binding, bending and cleavage of DNA substrates by the homing endonuclease PI-SceI.
<3>Nucleic Acids Res.
<4>24
<5>4123-4132
<6>1996
<7>To characterize the interaction between the homing endonuclease PI-SceI and DNA, we prepared
different DNA substrates containing the natural recognition sequence or parts thereof.
Depending on the nature of the substrates, efficient cleavage is observed with a DNA
containing ~30 bp of the natural recognition sequence using supercoiled plasmids, ~40-50 bp
using linearized plasmids and >50 bp using synthetic double-stranded oligodeoxynucleotides.
Cleavage of supercoiled plasmids occurs without accumulation of the nicked intermediate.  In
the presence of Mn2+ DNA cleavage by PI-SceI is more efficient than with Mg2+ and already
occurs with substrates containing a shorter part of the recognition sequence.  The
requirements for strong binding are less stringent: a 35 bp oligodeoxynucleotide which is not
cleaved is bound as firmly as other longer oligodeoxynucleotides.  PI-SceI binds with high
affinity to one of its cleavage products, a finding which may explain why PI-SceI hardly shows
enzymatic turnover in vitro.  Upon binding, two complexes are formed, which differ in the
degree of bending (45o versus 75o).  According to a phasing analysis bending is directed into
the major groove.  Strong binding, not, however, cleavage is also observed with the
genetically engineered enzymatically inactive variant comprising amino acids 1-277.  Models
for binding and cleavage of DNA by PI-SceI are discussed based on these results.

<>

<1>Wende, W., Pingoud, A.
<2>Cloning, overexpression, purification and characterization of the VDE homing endonuclease.
<3>Biol. Chem. Hoppe Seyler
<4>375
<5>S110
<6>1994
<7>Homing endonucleases propagate their encoding sequences (introns) from a donor
(intron-containing) allele into an intronless recipient allele. Only the intronless allele
contains the specific target sequence. The homing endonuclease cleaves the DNA on both
strands, and the repair of the intronless allele results in the insertion of the sequence
encoding the homing endonuclease. The target sites are generally asymmetric and long, ranging
from 15 bp to 40 bp. The VDE (VMA1-derived endonuclease) cleaves intronless yeast genomic DNA
only at a single site. Thus these enzymes are extremely rare cutters and, therefore, important
tools for chromosomal mapping, sequencing and gene targeting. In order to study the
DNA/protein interactions of this interesting class of enzymes, we constructed a His6-tagged
variant of the VDE from Saccharomyces cervisiae and overexpressed it in E. coli. The affinity
tag allows for a rapid purification to near homogeneity. The protein sequence of our VDE
differs in 3 positions from the published sequence. Analysis of circular dichroism spectra
suggests that the protein consists of 43% alpha-helix and 48% beta-sheet. The specific
activity of the His6-tagged VDE variant is comparable with the native protein. Preliminary
studies show that the minimal recognition site is larger than the published one. We are
currently engaged in DNA binding and cleavage studies with the His6-VDE, the results of which
will be presented.

<>

<1>Wende, W., Schottler, S., Grindl, W., Christ, F., Steuer, S., Noel, A.J., Pingoud, V., Pingoud, A.
<2>A mutational analysis of DNA binding and cleavage by the homing endonuclease PI-SceI.
<3>Mol. Biol. (Mosk)
<4>34
<5>1054-1064
<6>2000
<7>We have carried out an extensive mutational analysis of PI-SceI, the best studied intein-like
homing endonuclease of the LAGLIDADG family,
to find out which amino acid residues are involved in substrate binding
and processing. Our analysis was focused on domain I, in which two
regions were shown to be in contact with DNA, and on domain II, in
which the amino acid residues making up catalytic centers I and II were
identified and their role in catalysis investigated. As a result of our
comprehensive mutational analysis a model is presented for DNA binding
and cleavage by PI-SceI.

<>

<1>Wende, W., Stahl, F., Pingoud, A.
<2>The production and characterization of artificial heterodimers of the restriction endonuclease EcoRV.
<3>Biol. Chem. Hoppe Seyler
<4>377
<5>625-632
<6>1996
<7>A novel approach to studying the inter- and intrasubunit communication required for the
activity of homodimeric proteins is described. It was developed for the restriction
endonuclease EcoRV, but should also be useful for other homodimeric enzymes.  Two ecorV genes
encoding different EcoRV mutants are coexpressed in the same Escherichia coli cell leading to
homo- and heterodimeric variants of the enzyme.  The two ecorV genes carry either a 5'
extension coding for the glutathione-S-transferase or a His6-tag.  The EcoRV heterodimer
produced in vivo is separated from the two EcoRV homodimers and purified to homogeneity by
affinity chromatography.  Purified EcoRV heterodimers are stable and are not subject to
reassortment of the subunits.  To investigate the interdependence of the two catalytic
centers, EcoRV heterodimers consisting of one subunit with wild type sequence and one subunit
with amino acid substitutions in the PD...(D/E)XK motif, characteristic for the active sites
of many restriction endonucleases, were produced.  While the homodimeric EcoRV active site
mutants are catalytically inactive, the heterodimeric EcoRV variants with one active and one
inactive catalytic center display a twofold reduced activity toward oligodeoxynucleotide
substrates compared to the wild type, and preferentially nick supercoiled plasmid DNA.  From
these results we conclude that in the wild type enzyme both catalytic centers function
independently of each other.

<>

<1>Wenner, J.R., Bloomfield, V.A.
<2>A novel kinetic mechanism for the cleavage of plasmid DNA by the restriction enzyme EcoRV accounts for nonspecific binding.
<3>J. Biophys.
<4>72
<5>A98
<6>1997
<7>The cleavage of pBR322 by the restriction enzyme EcoRV was assayed by quantifyingDNA bands on
agarose gels.  The nonlinear reaction kinetics were fit by the mechanism.  Good fits were
obtained only if nonspecific binding and slow product release were included.  A key parameter
is the fraction of total binding that results in specific binding.  EcoRV must cycle through
the nonspecific binding process a number of times before the specific site is located.  This
kinetic picture will be used to measure how macromolecular crowding with inert agents affects
the rate at which EcoRV finds its specific cleavage site.

<>

<1>Wenner, J.R., Bloomfield, V.A.
<2>Osmotic pressure effects on EcoRV cleavage and binding.
<3>J. Biomol. Struct. Dyn.
<4>17
<5>461-471
<6>1999
<7>Investigations of DNA binding proteins frequently measure pH and salt dependence, but
relatively few studies measure protein binding in high concentrations of small molecules often
found in vivo. We have measured kinetics of the restriction enzyme EcoRV in concentrated
solutions of three small cosolvents that produce osmotic pressures from 0.25 to 2.5 mol/kg (6
to 62 atm or water activity of 0.995 to 0.956). We have correlated DNA cleavage and binding
parameters with four solution parameters (dielectric constant, viscosity, water concentration,
and water activity). We found that the responses of maximum velocity (Vmax) and the
dissociation constant for nonspecific binding (Kd,ns) best correlate with water activity. The
Michaelis constant (Km) correlates with both water activity and solution viscosity, the latter
due to the highly dilute reactant concentrations, which make enzyme-substrate combination
diffusion limited. Dielectric constant does not influence any of the kinetic parameters, which
is consistent with a view that protein and DNA are preferentially hydrated, and excluded
solutes cannot affect the local dielectric constant.

<>

<1>Wenner, J.R., Bloomfield, V.A.
<2>Buffer effects on EcoRV kinetics as measured by fluorescent staining and digital imaging of plasmid cleavage.
<3>Anal. Biochem.
<4>268
<5>201-212
<6>1999
<7>We have developed a protocol to quantify polymer DNA cleavage which replaces the traditional
radiolabeling and scintillation counting with fluorescent staining and digital imaging. This
procedure offers high sensitivity, speed, and convenience, while avoiding waste and error
associated with traditional 32P radiolabeling. This protocol was used to measure cleavage of
pBR322 plasmid DNA by EcoRV, a type II restriction enzyme. EcoRV was found to exhibit an order
of magnitude difference in binding in two apparently similar buffers used in previous
investigations. To determine the origin of this effect, we measured reaction kinetics in
buffers of different chemical nature and concentration: Tris, bis-Tris propane, Tes, Hepes,
and cacodylate. We found that buffer concentration and identity had significant effects on
EcoRV reaction velocity through large changes in specific binding and nonspecific binding
(reflected in the Michaelis constant Km and the dissociation constant for nonspecific binding
Kns). There were only small changes in Vmax. The source of the buffer effect is the protonated
amines common to many pH buffers. These buffer cations likely act as counterions screening DNA
phosphates, where both the protonated buffer structure and concentration affect enzyme binding
strength. It appears that by choosing anionic buffers or zwitterionic buffers with a buried
positive charge, buffer influence on the protein binding to DNA can be largely eliminated.

<>

<1>Wenren, L.M., Sullivan, N.L., Cardarelli, L., Septer, A.N., Gibbs, K.A.
<2>Two Independent Pathways for Self-Recognition in Proteus mirabilis Are Linked by Type VI-Dependent Export.
<3>MBio
<4>4
<5>e00374-13
<6>2013
<7>ABSTRACT Swarming colonies of the bacterium Proteus mirabilis are capable of
self-recognition and territorial behavior. Swarms of independent P. mirabilis
isolates can recognize each other as foreign and establish a visible boundary
where they meet; in contrast, genetically identical swarms merge. The ids genes,
which encode self-identity proteins, are necessary but not sufficient for this
territorial behavior. Here we have identified two new gene clusters: one (idr)
encodes rhs-related products, and another (tss) encodes a putative type VI
secretion (T6S) apparatus. The Ids and Idr proteins function independently of
each other in extracellular transport and in territorial behaviors; however,
these self-recognition systems are linked via this type VI secretion system. The
T6S system is required for export of select Ids and Idr proteins. Our results
provide a mechanistic and physiological basis for the fundamental behaviors of
self-recognition and territoriality in a bacterial model system. IMPORTANCE Our
results support a model in which self-recognition in P. mirabilis is achieved by
the combined action of two independent pathways linked by a shared machinery for
export of encoded self-recognition elements. These proteins together form a
mechanistic network for self-recognition that can serve as a foundation for
examining the prevalent biological phenomena of territorial behaviors and
self-recognition in a simple, bacterial model system.

<>

<1>Wentzell, L.M., Halford, S.E.
<2>DNA looping by the SfiI restriction endonuclease.
<3>J. Mol. Biol.
<4>281
<5>433-444
<6>1998
<7>The SfiI endonuclease has to interact with two copies of its recognition sequence before it
can cleave DNA.  To demonstrate that the reaction of SfiI on a DNA with two sites involves the
formation of a DNA loop, and to characterize the looping interactions on supercoiled and
linear DNA, a series of plasmids was constructed with lengths of DNA between two SfiI sites
varying from 104 to 211 bp.  Both supercoiled and linear forms of each DNA were tested as
substrates for SfiI.  The reactions were monitored from the rates of DNA cleavage and from the
generation of partially cleaved products, the latter indicating loop disruption before
cleavage of both sites.  On both supercoiled and linear DNA, the stabilities of the complexes
spanning two SfiI sites varied in sinusoidal fashion with the distance between the sites, in
the manner characteristic of a process governed by the helical periodicity of DNA.  In all
cases, the looping interaction was stabilized by DNA supercoiling.  The sinusoidal variation
from SfiI reactions on supercoiled DNA at 50 C yielded a helical repeat of about 11.5
base-pairs per turn.

<>

<1>Wentzell, L.M., Nobbs, T.J., Halford, S.E.
<2>The SfiI restriction endonuclease makes a four-strand DNA break at two copies of its recognition sequence.
<3>J. Mol. Biol.
<4>248
<5>581-595
<6>1995
<7>The SfiI endonuclease cleaves DNA by a mechanism that differs from other restriction enzymes.
While most restriction enzymes are dimeric proteins that interact with a single DNA site, SfiI
exists in solution as a tetramer and it appears to interact with two copies of its recognition
sequence before it can cleave DNA. Its primary reaction is then to cut both strands at both
sites in a concerted process. The two sites can be on either the same or different DNA
molecules, so SfiI provides a test system for long-range interactions on DNA. On either
supercoiled or linear DNA with two sites separated by 1 kb, the bridging interaction between
the sites is an intramolecular event: the majority of the DNA is converted directly to
products cleaved at both sites, bypassing intermediates cut at one site. Sites on separate DNA
molecules, or two sites on linear DNA several kb apart, engage in an intermolecular
interaction prior to cleavage. The interaction between two DNA molecules with one site on each
is impeded by supercoiling in both partners but is permitted when one partner is linear: it
may require reptation of one DNA through another. SfiI reactions have marked similarities to
some of the reactions catalysed by site-specific recombination enzymes.

<>

<1>Wentzell, L.M., Oram, M., Halford, S.E.
<2>Purification and characterisation of the SfiI restriction endonuclease.
<3>Biochem. Soc. Trans.
<4>22
<5>302S
<6>1994
<7>Type II restriction endonucleases recognise specific DNA sequences and cleave at a defined
point within or close to that sequence. They require Mg2+ as a cofactor. In the presence of
Mg2+ they cut their recognition sites at least a million times faster than at any other DNA
sequence. At present over 2400 different type II restriction enzymes have been identified
representing over 188 different specificities. Most type II enzymes cleave DNA within
symmetrical sequences, termed palindromes. These are usually continuous sequences of 4 or 6
base pairs, or fairly rarely 8 base pairs. Some of these enzymes cleave at unique DNA
sequences as for EcoRV (GATATC), while others recognize degenerate sequences as for HindII
(GTYRAC). Other enzymes, SfiI being one of them, recognize discontinuous sequences in which
one or more unspecified bases interrupt the sequence of specified bases. A further group of
enzymes known as type IIs, recognize asymmetric sequences and cleave at a fixed point from
that sequence.

<>

<1>Wenz, C., Hahn, M., Pingoud, A.
<2>Engineering of variants of the restriction endonuclease EcoRV that depend in their cleavage activity on the flexibility of sequences flanking the recognition site.
<3>Biochemistry
<4>37
<5>2234-2242
<6>1998
<7>The present work describes mutants of the restriction enzyme EcoRV that discriminate very
efficiently between oligodeoxynucleotide substrates with an EcoRV recognition sequence in
different sequence contexts.  All of these EcoRV variants harbor substitutions at position
226, where in the cocrystal structure of the specific EcoRV/DNA complex an arginine contacts
the backbone of the DNA substrate upstream of the recognition sequence, and cleave an
oligodeoxynucleotide with an EcoRV site in a nonflexible sequence context (the recognition
site being flanked by runs of A and T) with much higher catalytic efficiency (kcat/Km) than an
oligodeoxynucleotide with an EcoRV site in a flexible sequence context (the recognition site
being flanked by runs of AT), in contrast to the wild-type enzyme, that cleaves both
substrates with the same catalytic efficiency.  Steady-state and single-turnover kinetics
indicate that the enhanced selectivity of the mutants is due to the catalytic step of the
reaction.  It is possible to enhance the discriminatory power of these EcoRV variants through
the choice of appropriate reaction conditions, in particular low salt concentration and low
reaction temperatures.  It must be emphasized that the enhanced selectivity of these EcoRV
variants toward EcoRV sites in a flexible and nonflexible sequence context, respectively, is
not only seen with oligodeoxynucleotides, but also with plasmid substrates.

<>

<1>Wenz, C., Jeltsch, A., Pingoud, A.
<2>Probing the indirect readout of the restriction enzyme EcoRV.
<3>J. Biol. Chem.
<4>271
<5>5565-5573
<6>1996
<7>According to the crystal structure of the specific EcoRV.DNA complex, not only the functional
groups of the nucleobases but also the phosphate groups of the DNA backbone are contacted by
the enzyme.  To examine the contribution of backbone contacts to substrate recognition and
catalysis by EcoRV, we exchanged 12 amino acids residues located close to phosphate groups by
site-directed mutagenesis.  We purified the resulting EcoRV mutants and characterized them
with respect to their DNA binding and cleavage activity.  According to our steady state
kinetic analysis, there are strong interactions between three basic amino acid residues
(Lys-119, Arg-140, and Arg-226) and the phosphate backbone that support specific binding
presumably by inducing and maintaining the kinked conformation of the DNA observed in the
specific EcoRV.DNA complex.  These contacts are important in both the ground state and the
transition state.  Other, uncharged residues (Thr-93 and Ser-112), which could be involved in
hydrogen bonds to the phosphate groups, are needed primarily to stabilize the transition
state.  An especially important amino acid residue is Thr-37, which seems to couple
recognition to catalysis by indirect readout.

<>

<1>Wenz, C., Pingoud, A.
<2>Probing the indirect readout of the restriction enzyme EcoRV; mutational analysis of contacts to the DNA-backbone.
<3>Biol. Chem. Hoppe Seyler
<4>376
<5>S168
<6>1995
<7>Specific recognition of DNA does not only include interactions between the protein and the
bases (direct readout), but also contacts to the phosphate groups of the DNA, which means that
the protein also recognizes a specific, sequence dependent conformation of the phosphodiester
backbone (indirect readout).  To examine contribution of phosphate contacts for the
recognition process we have carried out a mutational analysis: The amino acid residues which
are, according to the crystal structure of the specific EcoRV/DNA complex, in the vicinity of
phosphate groups of the DNA and bear functional groups suited for hydrogen bonds or ionic
interactions, were exchanged to Ala (Arg, Lys, Ser, Thr) or to Phe (Tyr) via PCR-mutagenesis.
All mutants displayed residual activity with plasmid-DNA and oligodeoxynucleotide substrates
and could, therefore, be analysed in terms of specific binding (with Ca2+ ions), steady state
and single turnover kinetics, linear diffusion and selectivity towards modified
oligodeoxynucleotides and various substrates with different sequences flanking the EcoRV-site
GAT/ATC- (the arrow denotes the position of phosphodiester bond cleavage).  Several mutants
displayed an altered selectivity for substrates with different flanking sequences, and/or a
different ability to bind specifically to DNA and to diffuse along the DNA.  These results
suggest that EcoRV makes use of phosphate contacts to locate EcoRV sites on macromolecular DNA
by linear diffusion and to attack different EcoRV sites more or less evenly.  Furthermore, our
results indicate that it should be possible to engineer mutants with an expanded specificity
by taking advantage of an altered selectivity towards substrates with different flanking
sequences.

<>

<1>Wenz, C., Pingoud, A.
<2>Protein engineering of the restriction endonuclease EcoRV.
<3>Biol. Chem. Hoppe Seyler
<4>375
<5>S110
<6>1994
<7>Specific recognition of DNA does not only include interactions between the protein and the
bases (direct readout), but also contacts to the phosphate groups of the DNA: the protein
recognizes a specific, sequence dependent conformation of the phosphodiester backbone
(indirect readout). Consequently, it should be possible to alter the specificity of a
restriction enzyme through manipulation of the phosphate contacts. Our approach to modulate
the specificity of EcoRV is to delete functional groups of amino acids which are in direct
contact with the phosphate groups of the DNA. Some of these EcoRV variants display a markedly
changed specificity, e.g. the mutant Arg140 -> Ala, which shows in comparison to the wild type
enzyme a strong preference for EcoRV sites with AT-rich flanking sequences. However, this is
accompanied with a decrease in specific activity. It remains to be seen, whether by other
and/or additinoal mutations the selectivity can be further increased without losing too much
of the activity of the wild type enzyme.

<>

<1>Wenz, C., Selent, U., Wende, W., Jeltsch, A., Wolfes, H., Pingoud, A.
<2>Protein engineering of the restriction endonuclease EcoRV: replacement of an amino acid residue in the DNA binding site leads to an altered selectivity towards unmodified and modified substrates.
<3>Biochim. Biophys. Acta
<4>1219
<5>73-80
<6>1994
<7>According to the crystal structure analysis of a specific EcoRV/DNA complex, the thymine
residues of the recognition sequence -GATATC- are not in direct contact with any amino acid
residue of the protein. However, several amino acid residues are sufficiently close that it
seemed worthwhile trying to create variants of EcoRV with altered specificity by site-directed
mutagenesis. Guided by molecular modelling we have replaced Asn-188 in the catalytic center of
EcoRV by Gln to produce a mutant with a relative preference (compared to wild type EcoRV) for
substrates in which one thymine of the recognition sequence is replaced by uracil. We have
purified and characterized the resulting N188Q mutant. The selectivity value for the
engineered enzyme (the ratio of the kcat/KM values for -GATAUC- versus -GATATC-) differs from
that of the wild type enzyme by a factor of more than 200.

<>

<1>Wenzel, C., Guschlbauer, W.
<2>Dam methyltransferase from Escherichia coli:  sequence of a peptide segment involved in S-adenosyl-methionine binding.
<3>Nucleic Acids Res.
<4>21
<5>4604-4609
<6>1993
<7>DNA adenine methyltransferase (Dam methylase) has been crosslinked with its cofactor
S-adenosyl methionine (AdoMet) by UV irradiation. About 3% of the enzyme was radioactively
labeled after the crosslinking reaction performed either with (methyl-3H)-AdoMet or with
(carboxy-14C)-AdoMet. Radiolabelled peptides were purified after trypsinolysis by high
performance liquid chromatography in two steps. They could not be sequenced due to radiolysis.
Therefore we performed the same experiment using non-radioactive AdoMet and were able to
identify the peptide modified by the crosslinking reaction by comparison of the separation
profiles obtained from two analytical control experiments performed with 3H-AdoMet and Dam
methylase without crosslink, respectively. This approach was possible due to the high
reproducibility of the chromatography profiles. In these three experiments only one
radioactively labelled peptide was present in the tryptic digestions of the crosslinked
enzyme. Its sequence was found to be XA-GGK, corresponding to amino acids 10 - 14 of Dam
methylase. The non-identified amino acid in the first sequence cycle should be a tryptophan,
which is presumably modified by the crosslinking reaction. The importance of this region near
the N-terminus for the structure and function of the enzyme was also demonstrated by
proteolysis and site-directed mutagenesis experiments.

<>

<1>Wenzel, C., Moulard, M., Lobner-Olesen, A., Guschlbauer, W.
<2>Crosslinking of Dam methyltransferase with S-adenosyl-methionine.
<3>FEBS J.
<4>280
<5>147-151
<6>1991
<7>Highly purified DNA-adenine methyltransferase was irradiated in the presence of different
concentrations of radiolabelled S-adenosyl-methionine (AdoMet) with a conventional Mineralight
UV-lamp from several minutes up to 1 h while incubating in ice. Incorporation of radioactivity
was monitored by electrophoresis of the crosslink between S-adenosyl-methionine and Dam
methylase on SDS-polyacrylamide gels followed by fluorography. Crosslinking reached a maximum
in presence of 10 uM S-adenosyl-methionine; it was inhibited in the presence of substances
which competitively inhibit methylation of DNA by DAM methylase, like sinefungin or
S-adenosyl-homocysteine, but not in the presence of non-inhibitors like ATP or
S-isobutyl-adenosine. The crosslink obtained was resistant against a wide range of even
drastic conditions commonly used in protein and peptide chemistry. Proteins, which do not bind
S-adenosyl-methionine, as well as heat inactivated Dam methylase were not photolabelled. After
limited proteolysis the radioactive label appeared only in certain of the peptides obtrained.
From Western blots carried out with polyclonal antibodies produced against a synthetic peptide
corresponding in its sequence to amino acids 92-106 of the Dam methylase, the crosslinking of
AdoMet could be tentatively mapped at a position after amino acid 106.

<>

<1>Wenzlau, J.M., Saldanha, R.J., Butow, R.A., Perlman, P.S.
<2>A latent intron-encoded maturase is also an endonuclease needed for intron mobility.
<3>Cell
<4>56
<5>421-430
<6>1989
<7>Some yeast mitochondrial introns encode proteins that promote either splicing (maturases) or
intron propagation via gene conversion (the fit1 endonuclease). We surveyed introns in the
cox1 gene for their ability to engage in gene conversion and found that the group I intron,
aI4alpha, was efficiently transmitted to genes lacking it. An endonucleolytic cleavage is
detectable in recipient DNA molecules near the site of intron insertion in vivo and in vitro.
Conversion is dependent on an intact aI4alpha open reading frame. This intron product is a
latent maturase, but these data show that it is also a potent endonuclease involved in
recombination. Dual function proteins that cleave DNA and facilitate RNA splicing may have
played a pivotal role in the propagation and tolerance of introns.

<>

<1>Werner, E., Wende, W., Pingoud, A., Heinemann, U.
<2>High resolution crystal structure of domain I of the Saccharomyces cerevisiae homing endonuclease PI-SceI.
<3>Nucleic Acids Res.
<4>30
<5>3962-3971
<6>2002
<7>The homing endonuclease PI-SceI from Saccharomyces cerevisiae consists of two domains. The
protein splicing domain I catalyzes the excision of the mature endonuclease (intein) from a
precursor protein and the religation of the flanking amino acid sequences (exteins) to a
functional protein. Furthermore, domain I is involved in binding and recognition of the
specific DNA substrate. Domain II of PI-SceI, the endonuclease domain, which is structurally
homologous to other homing endonucleases from the LAGLIDADG family, harbors the
endonucleolytic center of PI-SceI, which in vivo initiates the homing process by introducing a
double-strand cut in the 35 bp recognition sequence. At 1.35 angstrom resolution, the crystal
structure of PI-SceI domain I provides a detailed view of the part of the protein that is
responsible for tight and specific DNA binding. A geometry-based docking of the 75 degree bent
recognition sequence to the full-length protein implies a conformational change or hinge
movement of a subdomain of domain I, the tongs part, that is predicted to reach into the major
groove near base pairs +16 to +18.

<>

<1>Werner, E.A.R.
<2>Intracellular events following infection by bacteriophage P1:  development of host controlled modification and restriction.
<3>Diss. Abstr.
<4>29
<5>3420
<6>1968
<7>Investigations of the kinetics of development of host controlled restriction
and modification and of lysogenic immunology of P1-infected Shigella
dysenteriae were carried out.  The relationships between the three events are
of interest in order to elucidate their correlation to other events in the
intracellular life cycle of P1.

<>

<1>Werner, E.R., Christensen, J.R.
<2>Infection by bacteriophage P1 and development of host-controlled restriction and modification and of Lysogenic immunity.
<3>J. Virol.
<4>3
<5>363-368
<6>1969
<7>Shigella dysenteriae cells were infected with phage P1 or P1c1.  The outcome of
superinfection of these cells with phage T1.Sh or T1.Sh(P1) or P1c1 was studied
as a function of time after the initial infection.  Cells undergoing either a
lytic responses or a lysogenic response to the primary infection develop the
ability to specifically restrict T1.Sh between 30 and 45 min.  Between 15 and
30 min, the cells seem to develop the ability to produce T1.Sh(P1) after
infection by T1.Sh.  However, reasons are given for believing that this
apparent time difference is consistent with a simultaneous development of the
two capacities (restriction and modification) within the cell.  This
development occurs between 30 and 45 min.  Cells infected with P1c1 and
superinfected 45 or more min later with T1.Sh(P1) can yield both P1cl and T1.
Cells infected with P1 become resistant to infection by P1cl within 5 to 10
min.  It is argued that this early immunity is not necessarily different in
mechanism from true lysogenic immunity.

<>

<1>Wernette, C., Saldanha, R., Smith, D., Ming, D., Perlman, P.S., Butow, R.A.
<2>Complex recognition site for the group I intron-encoded endonuclease I-SceII.
<3>Mol. Cell. Biol.
<4>12
<5>716-723
<6>1992
<7>We have characterized features of the site recognized by a double-stranded DNA endonuclease,
I-SceII, encoded by intron 4a of the yeast mitochondrial COX1 gene.  We determined the effects
of 36 point mutations on the cleavage efficiency of natural and synthetic substrates
containing the Saccharomyces capensis I-SceII site.  Most mutations of the 18-bp I-SceII
recognition site are tolerated by the enzyme, and those mutant sites are cleaved between 42
and 100% as well as the wild-type substrate is.  Nine mutants blocked cleavage to less than or
equal to 33% of the wild-type, whereas only three point mutations, G-4-C, G-12-T, and G-15-C,
block cleavage completely.  Competition experiments indicate that these three substrates are
not cleaved, at least in part because of a marked reduction in the affinity of the enzyme for
those mutant DNAs.  About 90% of the DNAs derived from randomization of the nucleotide
sequence of the 4-bp staggered I-SceII cleavage site are not cleaved by the enzyme.  I-SceII
cleaves cloned DNA derived from human chromosome 3 about once every 110 kbp.  The I-SceII
recognition sites in four randomly chosen human DNA clones have 56 to 78% identity with the
18-bp site in yeast mitochondrial DNA; they are cleaved at least 50% as well as the wild-type
mitochondrial substrate despite the presence of some substitutions that individually
compromise

<>

<1>Wernette, C.M.
<2>Structure and activity of the mitochondrial intron-encoded endonuclease, I-SceIV.
<3>Biochem. Biophys. Res. Commun.
<4>248
<5>127-133
<6>1998
<7>Starting with crude yeast mitochondria, the intron homing endonuclease, I-SceIV, was purified
to near homogeneity.  This highly purified enzyme differs from some other well-characterized
yeast mitochondrial intron-encoded endonucleases in terms of its structure and DNA cleavage
specificity.  The enzyme is a heterodimer with a native molecular mass of 92 kDa.  A small
catalytic subunit (32 kDa) is probably encoded largely or entirely by intron 5a of the
cytochrome oxidase subunit I gene.  A larger polypeptide subunit (60 kDa) may be a nuclear
factor necessary for intron mobility.  I-SceIV exhibits a low DNA sequence specificity as it
cleaves a variety of DNA substrates.  Analysis of kinetic parameters shows that the purified
enzyme has a very high affinity for DNA and exhibits low turnover which may have implications
for subsequent steps in the intron homing process.

<>

<1>Wernette, C.M., Saldahna, R., Perlman, P.S., Butow, R.A.
<2>Purification of a site-specific endonuclease, I-Sce II, encoded by intron 4 alpha of the mitochondrial coxI gene of Saccharomyces cerevisiae.
<3>J. Biol. Chem.
<4>265
<5>18976-18982
<6>1990
<7>We have purified to near homogeneity a site-specific, double-stranded DNA
endonuclease (I-Sce II) encoded by intron 4 alpha (aI4 alpha) of the yeast
mitochondrial coxI gene.  Our purification starts with a high salt extract of
mitochondria isolated from a yeast strain that overproduces the enzyme because
of a block in splicing of aI4 alpha.  The final step of purification is an
affinity column consisting of covalently bound double-stranded DNA multimers of
a synthetic sequence, 5'-TTGGTCATCCAGAAGTAT-3', which contains the I-Sce II
cleavage/recognition site.  Typical yields of enzyme are 3-5% with a specific
activity of approximately 500,000 units/mg, where 1 unit of activity cleaves 50
ng of DNA substrate/h at 30C.  I-Sce II has a monomer molecular mass of 31 kDa
as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
Active enzyme purifies as a 55-kDa species, which we presume to be a homodimer.
I-Sce II monomer comigrates with an in vivo synthesized mitochondrial
translation product made in the strain that overproduces the enzyme.  We
conclude that I-Sce II is derived by proteolytic processing of a precursor
polypeptide, p62, encoded by an in-frame fusion of coxI exons 1-4 with the
downstream aI4 alpha reading frame.  I-Sce II is most active at pH 7.5 and at
20-30C.  Endonuclease activity is sensitive to salt and is dependent upon Mg2+
or Mn2+, but is unaffected by inclusion of ATP or GTP.  I-Sce II is the first
intron-encoded protein to be purified and characterized from yeast
mitochondria.

<>

<1>Wernick, D.G., Choi, K.Y., Tat, C.A., Lafontaine, R.J.G., Liao, J.C.
<2>Genome Sequence of the Extreme Obligate Alkaliphile Bacillus marmarensis Strain DSM 21297.
<3>Genome Announcements
<4>1
<5>e00967-13
<6>2013
<7>Bacillus marmarensis strain DSM 21297 is an extreme obligate alkaliphile able to  grow in
medium up to pH 12.5. A whole-shotgun strategy and de novo assembly led
to the generation of a 4-Mbp genome of this strain. The genome features
alkaliphilic adaptations and pathways for n-butanol and poly(3-hydroxybutyrate)
synthesis.

<>

<1>Wesselink, J.J., Lopez-Camacho, E., de la Pena, S., Ramos-Ruiz, R., Ruiz-Carrascoso, G., Lusa-Bernal, S., Fernandez-Soria, V.M., Gomez-Gil, R., Gomez-Puertas, P., Mingorance, J.
<2>Genome Sequence of OXA-48 Carbapenemase-Producing Klebsiella pneumoniae KpO3210.
<3>J. Bacteriol.
<4>194
<5>6981
<6>2012
<7>Klebsiella pneumoniae KpO3210 is a OXA-48 carbapenemase-producing isolate obtained from a
blood culture in the context of an outbreak in Hospital
Universitario La Paz (Madrid, Spain) in 2010. It belongs to the major clone
detected during the outbreak and is resistant to all beta-lactams and to several
other antibiotics.

<>

<1>Westberg, J., Persson, A., Holmberg, A., Goesmann, A., Lundeberg, J., Johansson, K.E., Pettersson, B., Uhlen, M.
<2>The Genome Sequence of Mycoplasma mycoides subsp. mycoides SC Type Strain PG1T, the Causative Agent of Contagious Bovine Pleuropneumonia (CBPP).
<3>Genome Res.
<4>14
<5>221-227
<6>2004
<7>Mycoplasma mycoides subsp. mycoidesSC (MmymySC)is the etiological agent of
contagious bovine pleuropneumonia (CBPP), a highly contagious respiratory
disease in cattle. The genome of Mmymy SC type strain PG1(T) has been
sequenced to map all the genes and to facilitate further studies regarding
the cell function of the organism and CBPP. The genome is characterized by
a single circular chromosome of 1211703 bp with the lowest G content (24
mole%)and the highest density of insertion sequences (13% of the genome
size)of all sequenced bacterial genomes. The genome contains 985 putative
genes, of which 72 are part of insertion sequences and encode
transposases. Anomalies in the GC-skew pattern and the presence of large
repetitive sequences indicate a high genomic plasticity. A variety of
potential virulence factors was identified, including genes encoding
putative variable surface proteins and enzymes and transport proteins
responsible for the production of hydrogen peroxide and the capsule, which
is believed to have toxic effects on the animal.

<>

<1>Whang, Y., Chae, K.S., Jang, W.H., Kim, K.T., Yoo, O.J.
<2>A new restriction endonuclease from Xanthomonas citri.
<3>Korean J. Microbiol.
<4>24
<5>406-410
<6>1986
<7>The isolation and characterization of a type II restriction endonuclease from
Xanthomonas citri IFO3835 were described.  This enzyme (XciI endonuclease) is
an isoschizomer of SalI endonuclease recognizing 5'-GTCGAC-3' and cleaving at
the site indicated by the arrow.  Unlike SalI endonuclease, XciI endonuclease
requires a NaCl concentration of 50 mM for its maximum activity.

<>

<1>Whiley, S.J., Lanser, J.A., Manning, P.A., Murray, C., Steele, T.W.
<2>Plasmid profile analysis of a Salmonellosis outbreak and identification of a restriction and modification system.
<3>Appl. Environ. Microbiol.
<4>54
<5>1591-1594
<6>1988
<7>After an outbreak of salmonellosis in humans caused by Salmonella typhimurium
bacteriophage type 135, 62 isolates from human, animal, and water sources were
retained for further analysis.  Most of the isolates (92%) could be placed in
one of five plasmmid pattern groups, with a majority containing a common
60-kilobase plasmid and a smaller 3.8-kilobase-pair plasmid.  This small
plasmid, pIMVS1, was labeled with [32P]phosphate and used as a probe in
subsequent colony and Southern hybridization studies.  We concluded that pIMVS1
from isolates obtained from humans was genetically different from plasmids of a
similar size found in isolates from chickens.  Studies to characterize pIMVS1
were undertaken to determine if it codes for known virulence factors.  It did
not appear to be associated with the formation of attachment pili or major
outer membrane proteins.  By using transposon mutagenesis techniques, Tn3(Apr)
was inserted into pIMVS1, and the existence of a restriction and modification
system was deduced.

<>

<1>Whitaker, R.D., Dorner, L.F., Schildkraut, I.
<2>A mutant of BamHI restriction endonuclease which requires N6-methyladenine for cleavage.
<3>J. Mol. Biol.
<4>285
<5>1525-1536
<6>1999
<7>Amino acid residues Asn116 and Ser118 of the restriction endonuclease BamHI make several
sequence-specific and water-bridged contacts to the DNA bases.  An in vivo selection was used
to isolate BamHI variants at position 116, 118 and 122 which maintained sequence specificity
to GGATCC sites.  Here, the variants N116H, N116H/S118G and S118G were purified and
characterized.  The variants N116H and N116H/S118G were found to have lost their ability to
cleave unmethylated GGATCC sequences by more than two orders of magnitude, while maintaining
nearly wild-type levels of activity on the N6-methyladenine-containing sequence, GGmATCC.  In
contrast, wild-type BamHI and variant S118G have only a three- to fourfold lower activity on
unmethylated GGATCC sequences compared with GGmATCC sequences.  The N116 to H116 mutation has
effectively altered the specificity of BamHI from an endonuclease which recognizes and cleaves
GGATCC and GGmATC, to an endonuclease which only cleaves GGmATCC.  The N116H change of
specificity is due to the lowered binding affinity for the unmethylated sequence because of
the loss of two asparagine-DNA hydrogen bonds and the introduction of a favorable van der
Waals contact between the imidazole group of histidine and the N6-methyl group of adenine.

<>

<1>White, J.R., Escobar-Paramo, P., Mongodin, E.F., Nelson, K.E., DiRuggiero, J.
<2>Extensive genome rearrangements and multiple horizontal gene transfers in a population of pyrococcus isolates from Vulcano Island, Italy.
<3>Appl. Environ. Microbiol.
<4>74
<5>6447-6451
<6>2008
<7>The extent of chromosome rearrangements in Pyrococcus isolates from marine
hydrothermal vents in Vulcano Island, Italy, was evaluated by
high-throughput genomic methods. The results illustrate the dynamic nature
of the genomes of the genus Pyrococcus and raise the possibility of a
connection between rapidly changing environmental conditions and adaptive
genomic properties.

<>

<1>White, O. et al.
<2>Genome sequence of the radioresistant bacterium Deinococcus radiodurans R1.
<3>Science
<4>286
<5>1571-1577
<6>1999
<7>The complete genome sequence of the radiation-resistant bacterium Deinococcus radiodurans R1
is composed of two chromosomes (2,648,638 and 412,348 base pairs), a megaplasmid (177,466 base
pairs), and a small plasmid (45,704 base pairs), yielding a total genome of 3,284,156 base
pairs.  Multiple components distributed on the chromosomes and megaplasmid that contribute to
the ability of D. radiodurans to survive under conditions of starvation, oxidative stress, and
high amounts of DNA damage were identified.  Deinococcus radiodurans represents an organism in
which all systems for DNA repair, DNA damage export, desiccation and starvation recovery, and
genetic redundancy are present in one cell.

<>

<1>White, O., Benta, J.C., Smith, H.O., Iii, C.A.H., Gocayne, J.D., Adams, M.D., Fraser, C.M.
<2>Nucleotide Sequence of the Mycoplasma Genitalium Genome, Fragments Thereof, and Uses Thereof.
<3>Japanese Patent Office
<4>JP 2004073185 A
<5>
<6>2004
<7>
<>

<1>White, R.A.I.I.I., Grassa, C.J., Suttle, C.A.
<2>First draft genome sequence from a member of the genus agrococcus, isolated from  modern microbialites.
<3>Genome Announcements
<4>1
<5>e00391-13
<6>2013
<7>We report the first draft genome sequence from a member of the genus Agrococcus,  isolated
from cold thrombolytic microbialites within Pavilion Lake, British
Columbia, Canada. The draft genome assembly for Agrococcus pavilionensis strain
RW-1 has a size of 2,878,403 bp with a G+C content of 72.56%.

<>

<1>White, R.A.I.I.I., Grassa, C.J., Suttle, C.A.
<2>Draft Genome Sequence of Exiguobacterium pavilionensis Strain RW-2, with Wide Thermal, Salinity, and pH Tolerance, Isolated from Modern Freshwater  Microbialites.
<3>Genome Announcements
<4>1
<5>e00597-13
<6>2013
<7>We report the draft genome sequence of Exiguobacterium pavilionensis strain RW-2, isolated
from a cold thrombolytic microbialite. The isolate grows at temperatures
from 4 to 50 degrees C, at pH levels from 5 to 11, and in media without added
NaCl or KCl or with 7% added NaCl.

<>

<1>White, R.A.I.I.I., Suttle, C.A.
<2>The Draft Genome Sequence of Sphingomonas paucimobilis Strain HER1398 (Proteobacteria), Host to the Giant PAU Phage, Indicates That It Is a Member of  the Genus Sphingobacterium (Bacteroidetes).
<3>Genome Announcements
<4>1
<5>e00598-13
<6>2013
<7>The draft genome sequence of Sphingomonas paucimobilis host index number (HER) 1398, host of
the giant PAU phage isolated from silk moths (Bombyx mori),
indicates that this isolate belongs within the genus Sphingobacterium. We suggest
that Sphingomonas paucimobilis strain HER1398 be reclassified as Sphingobacterium
paucimobilis strain HER1398.

<>

<1>Whitehead, E.P., Taddeo, B., Stampeggioni, E., Palitti, F., Carotti, D.
<2>Measurement of DNA methylase activity by tritium release from DNA cytosine.
<3>Cell Biophys.
<4>15
<5>145-147
<6>1989
<7>The advantages of assaying DNA methylase by measuring the transfer to water of
tritium from the 5-position of DNA cytosine, rather than the transfer to DNA of
labeled methyl groups are discussed.

<>

<1>Whitehead, P.R., Brown, N.L.
<2>A simple and rapid method for screening bacteria for type II restriction endonucleases:  enzymes in Aphanothece halophytica.
<3>Arch. Microbiol.
<4>141
<5>70-74
<6>1985
<7>A method is described which allows a large number of bacterial strains to be
rapidly and easily screened for the presence of site-specific endonucleases.
The method involves selective permeabilization of the bacterial cell and
analysis of the exuded material.  Type II restriction endonucleases from
cyanobacteria and Gram-negative eubacteria have been detected and new enzymes
have been found.  The method should be widely applicable and easy to modify for
use in genera other than those tested.  Three-site-specific endonuclease
activities, detected by this method in Apanothece halophytica PCC 7412, were
purified and their recognition and cleavage specificies were determined AhaI
and AhaII recognise and cleave the same DNA sequences as CauII and AcyI
respectively; the specificity of AhaIII (TTT^AAA) has been reported previously
(Whitehead and Brown, 1982, FEBS Lett. 143: 296-300).

<>

<1>Whitehead, P.R., Brown, N.L.
<2>EaeI: a restriction endonuclease from Enterobacter aerogenes.
<3>FEBS Lett.
<4>155
<5>97-101
<6>1983
<7>We describe the isolation and characterization of a type II restriction endonuclease from
Enterobacter aerogenes.  This recognises and cleaves the family of related sequences:
5'-Py^G-G-C-C--Pu-3' to generate DNA fragments with 5'-tetranucleotide extensions.  EaeI
may be useful in molecular cloning experiments, especially in conjunction with other enzymes
which generate the same terminal extensions.  Potential problems in the methods used to
determine the cleavage specificity are discussed.

<>

<1>Whitehead, P.R., Brown, N.L.
<2>Three restriction endonucleases from Anabaena flos-aquae.
<3>J. Gen. Microbiol.
<4>131
<5>951-958
<6>1985
<7>Three site-specific endonucleases, AflI, AflII and AflIII, have been partially
purified from the cyanobacterium Anabaena flos-aquae CCAP 1403/13f.  Their
recognition and cleavage specificities have been determined to be:AflI
5'-G-^G-(A/T)-C-C-3'AflII 5'-C-^T-T-A-A-G-3'AflIII 5'-A-^C-Pu-Py-G-T-3'AflII
and AflIII are new specificities and may be useful in molecular cloning, as
well as in the anlaysis of DNA.  The distribution of type II restriction
endonucleases in the cyanobacteria is briefly discussed.

<>

<1>Whitehead, P.R., Brown, N.L.
<2>AhaIII: A restriction endonuclease with a recognition sequence containing only A:T basepairs.
<3>FEBS Lett.
<4>143
<5>296-300
<6>1982
<7>None

<>

<1>Whitehead, P.R., Brown, N.L.
<2>The characterization of novel restriction endonuclease specificities.
<3>Biochem. Soc. Trans.
<4>9
<5>272P
<6>1981
<7>A very rapid method has been developed for DNA sequence analysis of the
recognition and cleavage sites of type II restriction endonucleases.  DNA
fragments containing the cleavage site for a novel restriction endonuclease are
cloned in vectors derived from bacteriophage M13, for sequence analysis by the
chain-termination method.  The phosphodiester bonds in both DNA strands cleaved
by the restriction enzyme are identified in parallel with the sequence
determination.  The recognition and cleavage specificity of the enzyme AspAI,
has been determined to be the symmetrical sequence 5'-G-^G-T-N-A-C-C-3', and
the recognition and cleavage specificities of other enzymes will be presented.

<>

<1>Whitehead, P.R., Jacobs, D., Brown, N.L.
<2>Restriction endonucleases from Herpetosiphon giganteus:  an example of the evolution of DNA recognition specificity.
<3>Nucleic Acids Res.
<4>14
<5>7031-7045
<6>1986
<7>We describe the partial purification and characterisation of five Type II
restriction endonucleases from two strains of Herpetosiphon giganteus.  One of
the activities, HgiJII, was the first enzyme found that cleaves DNA at the
family of related sequences 5'-G-R-G-C-Y/C-3'.  This enzyme may be related to
the enzyme HgiAI from a different strain of the same species, and which cleaves
at the sites 5'-G-W-G-C-W/C-3'.  We have shown that DNAs from the strains
producing HgiAI and HgiJII are resistant to both of these restriction
endonucleases.   The remaining four enzymes described here share recognition
and cleavage specificities with other restriction endonucleases.  The evolution
of Type II restriction-modification systems and their role in vivo are
discussed.

<>

<1>Whiteson, K.L., Hernandez, D., Lazarevic, V., Gaia, N., Farinelli, L., Francois, P., Pilo, P., Frey, J., Schrenzel, J.
<2>A genomic perspective on a new bacterial genus and species from the Alcaligenaceae family, Basilea psittacipulmonis.
<3>BMC Genomics
<4>15
<5>169
<6>2014
<7>BACKGROUND: A novel Gram-negative, non-haemolytic, non-motile, rod-shaped
bacterium was discovered in the lungs of a dead parakeet (Melopsittacus
undulatus) that was kept in captivity in a petshop in Basel, Switzerland. The
organism is described with a chemotaxonomic profile and the nearly complete
genome sequence obtained through the assembly of short sequence reads. RESULTS:
Genome sequence analysis and characterization of respiratory quinones, fatty
acids, polar lipids, and biochemical phenotype is presented here. Comparison of
gene sequences revealed that the most similar species is Pelistega europaea, with
BLAST identities of only 93% to the 16S rDNA gene, 76% identity to the rpoB gene,
and a similar GC content (~43%) as the organism isolated from the parakeet, DSM
24701 (40%). The closest full genome sequences are those of Bordetella spp. and
Taylorella spp. High-throughput sequencing reads from the Illumina-Solexa
platform were assembled with the Edena de novo assembler to form 195 contigs
comprising the ~2 Mb genome. Genome annotation with RAST, construction of
phylogenetic trees with the 16S rDNA (rrs) gene sequence and the rpoB gene, and
phylogenetic placement using other highly conserved marker genes with ML Tree all
suggest that the bacterial species belongs to the Alcaligenaceae family. Analysis
of samples from cages with healthy parakeets suggested that the newly discovered
bacterial species is not widespread in parakeet living quarters. CONCLUSIONS:
Classification of this organism in the current taxonomy system requires the
formation of a new genus and species. We designate the new genus Basilea and the
new species psittacipulmonis. The type strain of Basilea psittacipulmonis is DSM
24701 (= CIP 110308 T, 16S rDNA gene sequence Genbank accession number JX412111
and GI 406042063).

<>

<1>Wiatr, C.L., Witmer, H.J.
<2>Selective protection of 5'..GGCC..3' and 5'...GCNGC...3' sequences by the hypermodified oxopyrimidine in Bacillus subtilis bacteriophage SP10 DNA.
<3>J. Virol.
<4>52
<5>47-54
<6>1984
<7>The DNA of Bacillus subtilis bacteriophage SP10 is partially resistant to
cleavage and methylation in vitro by restriction enzyme R.BsuRI and its cognate
methylase even though >20 copies of the target sequence, 5'...GGCC...3', are
present on the phage genome.  YThy, a hypermodified oxopyrimidine that replaces
a fraction of the thymine residues in SP10 DNA, was responsible for this
protection, since YThy-free DNA was no longer resistant.  Sites that were
normally resistant could nevertheless be cleaved or methylated in vitro if the
salt concentration was reduced or dimethyl sulfoxide was added to the reaction
buffer.  Analysis of the termini produced by cleavage suggested that resistant
sites occurred in the sequence 5'...GGCC-YThy ...3', whereas sensitive sites,
of which there were only two per genome, occurred in the sequence
5'...GGCCG...3'.  These in vitro results provide an explanation for the in vivo
resistance of SP10 to restriction-modification by B. subtilis R.  They also
suggest ways in which the presence of the atypical base XThy in regions that
flank the target might upset critical DNA-enzyme interactions necessary to
locate and recognize the specific site of cleavage or methylation.  YThy also
strongly protected 5'...GCNGC...3' (R.Fnu4HI) sequences on SP10 DNA, but the
biological relevance of this protection is unclear.

<>

<1>Wibberg, D., Blom, J., Jaenicke, S., Kollin, F., Rupp, O., Scharf, B., Schneiker-Bekel, S., Sczcepanowski, R., Goesmann, A., Setubal, J.C., Schmitt, R., Puhler, A., Schluter, A.
<2>Complete Genome Sequencing of Agrobacterium sp H13-3, the former Rhizobium lupini H13-3, Reveals a Tripartite Genome Consisting of a Circular and a Linear Chromosome and an Accessory Plasmid but Lacking a Tumor-Inducing Ti-Plasmid.
<3>J. Biotechnol.
<4>155
<5>50-62
<6>2011
<7>Agrobacterium sp. H13-3, formerly known as Rhizobium lupini H13-3, is a soil bacterium that
was isolated from the rhizosphere of Lupinus luteus. The isolate has been established as a
model system for studying novel features of flagellum structure, motility and chemotaxis
within the family Rhizobiaceae. The complete genome sequence of Agrobacterium sp. H13-3 has
been established and the genome structure and phylogenetic assignment of the organism was
analysed. For de novo sequencing of the Agrobacterium sp. H13-3 genome, a combined strategy
comprising 454-pyrosequencing on the Genome Sequencer FLX platform and PCR-based amplicon
sequencing for gap closure was applied. The finished genome consists of three replicons and
comprises 5,573,770 bases. Based on phylogenetic analyses, the isolate could be assigned to
the genus Agrobacterium biovar I and represents a genomic species G1 strain within this
biovariety. The highly conserved circular chromosome (2.82Mb) of Agrobacterium sp. H13-3
mainly encodes housekeeping functions characteristic for an aerobic, heterotrophic bacterium.
Agrobacterium sp. H13-3 is a motile bacterium driven by the rotation of several complex
flagella. Its behaviour towards external stimuli is regulated by a large chemotaxis regulon
and a total of 17 chemoreceptors. Comparable to the genome of Agrobacterium tumefaciens C58,
Agrobacterium sp. H13-3 possesses a linear chromosome (2.15Mb) that is related to its
reference replicon and features chromosomal and plasmid-like properties. The accessory plasmid
pAspH13-3a (0.6Mb) is only distantly related to the plasmid pAtC58 of A. tumefaciens C58 and
shows a mosaic structure. A tumor-inducing Ti-plasmid is missing in the sequenced strain H13-3
indicating that it is a non-virulent isolate.

<>

<1>Wibberg, D., Tielen, P., Blom, J., Rosin, N., Schobert, M., Tupker, R., Schatschneider, S., Spilker, D., Albersmeier, A., Goesmann, A., Vorholter, F.J., Puhler, A., Jahn, D.
<2>Genome Sequence of the Acute Urethral Catheter Isolate Pseudomonas aeruginosa MH38.
<3>Genome Announcements
<4>2
<5>e00161-14
<6>2014
<7>Pseudomonas aeruginosa is a major nosocomial bacterial pathogen causing complicated
catheter-associated urinary tract infections (CAUTIs). Here, we
present the 6.9-Mb draft genome sequence of P. aeruginosa MH38 isolated from an
acute nosocomial CAUTI. It exhibits resistance to several antibiotics but
revealed low-level production of virulence factors.

<>

<1>Wiegand, S., Rabausch, U., Chow, J., Daniel, R., Streit, W.R., Liesegang, H.
<2>Complete Genome Sequence of Geobacillus sp. Strain GHH01, a Thermophilic Lipase-Secreting Bacterium.
<3>Genome Announcements
<4>1
<5>e00092-13
<6>2013
<7>Geobacillus sp. strain GHH01 was isolated during a screening for producers of extracellular
thermostable lipases. The completely sequenced and annotated 3.6-Mb
genome encodes 3,478 proteins. The strain is genetically equipped to utilize a
broad range of different substrates and might develop natural competence.

<>

<1>Wiens, G.D., LaPatra, S.E., Welch, T.J., Rexroad, C.I.I.I., Call, D.R., Cain, K.D., LaFrentz, B.R., Vaisvil, B., Schmitt, D.P., Kapatral, V.
<2>Complete Genome Sequence of Flavobacterium psychrophilum Strain CSF259-93, Used To Select Rainbow Trout for Increased Genetic Resistance against Bacterial Cold  Water Disease.
<3>Genome Announcements
<4>2
<5>e00889-14
<6>2014
<7>The genome sequence of Flavobacterium psychrophilum strain CSF259-93, isolated from rainbow
trout (Oncorhynchus mykiss), consists of a single circular genome of
2,900,735 bp and 2,701 predicted open reading frames (ORFs). Strain CSF259-93 has
been used to select a line of rainbow trout with increased genetic resistance
against bacterial cold water disease.

<>

<1>Wiesner, M., Calva, E., Fernandez-Mora, M., Cevallos, M.A., Campos, F., Zaidi, M.B., Silva, C.
<2>Salmonella Typhimurium ST213 is associated with two types of IncA/C plasmids carrying multiple resistance determinants.
<3>BMC Microbiol.
<4>11
<5>9
<6>2011
<7>BACKGROUND: Salmonella Typhimurium ST213 was first detected in the Mexican
Typhimurium population in 2001. It is associated with a multi-drug
resistance phenotype and a plasmid-borne blaCMY-2 gene conferring
resistance to extended-spectrum cephalosporins. The objective of the
current study was to examine the association between the ST213 genotype
and blaCMY-2 plasmids. RESULTS: The blaCMY-2 gene was carried by an IncA/C
plasmid. ST213 strains lacking the blaCMY-2 gene carried a different
IncA/C plasmid. PCR analysis of seven DNA regions distributed throughout
the plasmids showed that these IncA/C plasmids were related, but the
presence and absence of DNA stretches produced two divergent types I and
II. A class 1 integron (dfrA12, orfF and aadA2) was detected in most of
the type I plasmids. Type I contained all the plasmids carrying the
blaCMY-2 gene and a subset of plasmids lacking blaCMY-2. Type II included
all of the remaining blaCMY-2-negative plasmids. A sequence comparison of
the seven DNA regions showed that both types were closely related to
IncA/C plasmids found in Escherichia, Salmonella, Yersinia,
Photobacterium, Vibrio and Aeromonas. Analysis of our Typhimurium strains
showed that the region containing the blaCMY-2 gene is inserted between
traA and traC as a single copy, like in the E. coli plasmid pAR060302. The
floR allele was identical to that of Newport pSN254, suggesting a mosaic
pattern of ancestry with plasmids from other Salmonella serovars and E.
coli. Only one of the tested strains was able to conjugate the IncA/C
plasmid at very low frequencies (10-7 to 10-9). The lack of conjugation
ability of our IncA/C plasmids agrees with the clonal dissemination trend
suggested by the chromosomal backgrounds and plasmid pattern associations.
CONCLUSIONS: The ecological success of the newly emerging Typhimurium
ST213 genotype in Mexico may be related to the carriage of IncA/C
plasmids. We conclude that types I and II of IncA/C plasmids originated
from a common ancestor and that the insertion and deletion of DNA
stretches have shaped their evolutionary histories.

<>

<1>Wigler, M., Levy, D., Perucho, M.
<2>The somatic replication of DNA methylation.
<3>Cell
<4>24
<5>33-40
<6>1981
<7>We have tested the hypothesis that DNA methylation patterns are replicated in the somatic
cells of vertebrates. Using M.HpaII, the modification enzyme from Haemophilus parainfluenzae
which methylates the internal cytosine residues in the sequence 5'CCGG/3'GGCC, we methylated
bacteriophage PhiX174 RF DNA and the cloned chicken thymidine kinase (tk) gene in vitro and
then introduced these DNAs and unmethylated controls into tk- cultured mouse cells by
DNA-mediated transformation. Twenty-five cell generations later, the state of methylation of
transferred DNA was examined by restriction endonuclease analysis and blot hybridization. We
conclude that methylation at HpaII sites is replicated by these cultured cells but not with
100% fidelity. We have also noted that methylation of the cloned chicken tk gene decreases its
apparent transformation efficiency relative to unmethylated molecules.

<>

<1>Wijetunge, D.S., Karunathilake, K.H., Chaudhari, A., Katani, R., Dudley, E.G., Kapur, V., DebRoy, C., Kariyawasam, S.
<2>Complete nucleotide sequence of pRS218, a large virulence plasmid, that augments pathogenic potential of meningitis-associated Escherichia coli strain RS218.
<3>BMC Microbiol.
<4>14
<5>203
<6>2014
<7>BACKGROUND: Escherichia coli is the most predominant Gram-negative bacterial
pathogen associated with neonatal meningitis. Previous studies indicated that the
prototypic neonatal meningitis E. coli (NMEC) strain RS218 (O18:K1:H7) harbors
one large plasmid. Objectives of the present study were to analyze the complete
nucleotide sequence of this large plasmid (pRS218) and its contribution to NMEC
pathogenesis using in vitro and in vivo models of neonatal meningitis. RESULTS:
The plasmid is 114,231 bp in size, belongs to the incompatibility group FIB/IIA
(IncFIB/IIA), and contains a genetic load region that encodes several virulence
and fitness traits such as enterotoxicity, iron acquisition and copper tolerance.
The nucleotide sequence of pRS218 showed a 41- 46% similarity to other neonatal
meningitis-causing E. coli (NMEC) plasmids and remarkable nucleotide sequence
similarity (up to 100%) to large virulence plasmids of E. coli associated with
acute cystitis. Some genes located on pRS218 were overly represented by NMEC
strains compared to fecal E. coli isolated from healthy individuals. The
plasmid-cured strain was significantly attenuated relative to the RS218 wild-type
strain as determined in vitro by invasion potential to human cerebral
microvascular endothelial cells and in vivo by mortalities, histopathological
lesions in the brain tissue, and bacterial recovery from the cerebrospinal fluid
of infected rat pups. CONCLUSIONS: The pRS218 is an IncFIB/IIA plasmid which
shares a remarkable nucleotide sequence similarity to large plasmids of E. coli
associated with cystitis. Both in vitro and in vivo experiments indicated that
pRS218 plays an important role in NMEC pathogenesis.

<>

<1>Wijsman, E.M.
<2>Optimizing selection of restriction enzymes in the search for DNA variants.
<3>Nucleic Acids Res.
<4>12
<5>9209-9226
<6>1984
<7>A model is developed for predicting the relative efficiencies of different
enzymes for detecting DNA variants when such variants are the result of single
base-pair changes.  71 enzymes are analyzed for this ability in human DNA.
Their relative ranked efficiencies are influenced by the sizes of the probes
used, and the size of the smallest detectable fragment produced.

<>

<1>Wilke, K., Rauhut, E., Noyer-Weidner, M., Lauster, R., Pawlek, B., Behrens, B., Trautner, T.A.
<2>Sequential order of target-recognizing domains in multispecific DNA-methyltransferases.
<3>EMBO J.
<4>7
<5>2601-2609
<6>1988
<7>In the multispecific DNA (cytosine-5)-methyltransferases (Mtases) of Bacillus
subtilis phages SPR and Phi3T the domains responsible for recognition of DNA
methylation targets CC(A/T)GG, CCGG, GGCC (SPR) and GCNGC, GGCC (Phi3T)
represent contiguous sequences of approximately 50 amino acids each.  These
domains are tandemly arranged and do not overlap.  They are part of a variable
segment within the enzymes which is flanked by conserved amino acids, which are
very similar amongst bacterial monospecific and the multispecific Mtases
studied here.  These results follow from a mutational analysis of the SPR and
Phi3T Mtase genes.  They further support our concept of a modular enzyme
organization, according to which variability of type II Mtases with respect to
target recognition is achieved by a combination of the same enzyme core with a
variety of target-recognizing domains.

<>

<1>Wilkes, T., Darby, A.C., Choi, J., Colborne, J.K., Werren, J.H., Hurst, G.D.D.
<2>The draft genome sequence of Arsenophonus nasoniae, son-killer bacterium of Nasonia vitripennis, reveals genes associated with virulence and symbiosis.
<3>Insect Mol. Biol.
<4>19
<5>59-73
<6>2010
<7>Four percent of female Nasonia vitripennis carry the son-killer bacterium Arsenophonus
nasoniae, a microbe with notably different biology from other inherited parasites and
symbionts. In this paper, we examine a draft genome sequence of the bacterium for open reading
frames (ORFs), structures and pathways involved in interactions with its insect host. The
genome data suggest that A. nasoniae carries multiple type III secretion systems, and an array
of toxin and virulence genes found in Photorhabdus, Yersinia and other gammaproteobacteria. Of
particular note are ORFs similar to those known to affect host innate immune functioning in
other bacteria, and four ORFs related to pro-apoptotic exotoxins. The genome sequences for
both A. nasoniae and its Nasonia host are useful tools for examining functional genomic
interactions of microbial survival in hostile immune environments, and mechanisms of passage
through gut epithelia, in a whole organism context.

<>

<1>Wilkins, B.M.
<2>Plasmid promiscuity: meeting the challenge of DNA immigration control.
<3>Environ. Microbiol.
<4>4
<5>495-500
<6>2002
<7>Bacterial plasmids are ubiquitous components of the genomes of naturally occurring bacteria
and are typically transmissible between cells by the process of bacterial conjugation (see
reviews in Thomas, 2000).  A remarkable feature of conjugation is its promiscuity, allowing
DNA transfer between phylogenetically remote bacteria and even from bacteria to plant and
mammalian cells (Waters, 2001).  Conjugative promiscuity is biologically important in
dispersing the diverse cargoes of medically and environmentally significant genes carried on
plasmids, as well as contributing to the horizontal transfer of 'fitness islands',
consisting of clustered chromosomal loci that enable pathogens and symbionts to interact with
their eukaryotic hosts (Finan, 2002).

<>

<1>Wilkins, B.M., Chilley, P.M., Thomas, A.T., Pocklington, M.J.
<2>Distribution of restriction enzyme recognition sequences on broad host range plasmid RP4: molecular and evolutionary implications.
<3>J. Mol. Biol.
<4>258
<5>447-456
<6>1996
<7>IncPalpha plasmids, exemplified by RP4, are remarkable for their broad host range.  They
contain strikingly few cleavage sites for many commonly used type II restriction enzymes but
an overabundance of sites for certain enzymes that target G + C-rich sequences. To identify
factors responsible for these distributions, the recently compiled nucleotide sequence of RP4
was analyzed to determine the frequency of tetra- and hexanucleotide motifs in the 49 kb
plasmid backbone.  This is defined as the sectors encoding basic plasmid functions.  The
overabundant restriction targets in RP4 are concentrated in the backbone and contain
overlapping copies of CGGC/GCCG, identified as the most abundant tetranucleotide motif in the
plasmid. Motif frequencies in the RP4 backbone are shown to be similar to those in Pseudomonas
aeruginosa, a natural host of RP4, with the notable exception that a number of 6-bp
palindromes are underrepresented in the plasmid.  It is proposed that 6-bp palindromes were
counterselected as type II restriction enzyme recognition sequences.  Conjugative transfer of
RP4 and R751 (IncPbeta) is usually sensitive to restriction compared to enterobacterial
plasmids of the IncFII and IncI1 groups, implying that IncP plasmids experienced particularly
strong selection for loss of restriction targets.  Pseudomonas spp. of rRNA homology group I
specify many type II restriction enzymes that target 6-bp palindromes and are candidates for
the evolutionary hosts of IncPalpha plasmids.

<>

<1>Wilkinson, C.R.M., Bartlett, R., Nurse, P., Bird, A.P.
<2>The fission yeast gene pmt1+ encodes a DNA methyltransferase homologue.
<3>Nucleic Acids Res.
<4>23
<5>203-210
<6>1995
<7>DNA methylation of cytosine residues is a widespread phenomenon and has been implicated in a
number of biological processes in both prokaryotes and eukaryotes. This methylation occurs at
the 5-position of cytosine and is catalyzed by a distinct family of conserved enzymes, the
cytosine-5 methyltransferases (m5C-MTases). We have cloned a fission yeast gene pmt1+ (pombe
methyltransferase) which encodes a protein that shares significant homology with both
prokaryotic and eukaryotic m5C-MTases. All 10 conserved domains found in these enzymes are
present in the pmt1 protein. This is the first m5C-MTase homologue cloned from a fungal
species. Its presence is surprising, given the inability to detect DNA methylation in yeasts.
Haploid cells lacking the pmt1+ gene are viable, indicating that pmt1+ is not an essential
gene. Purified, bacterially produced pmt1 protein does not possess obvious methyltransferase
activity in vitro. Thus the biological significance of this m5C-MTase homologue in fission
yeast is currently unclear.

<>

<1>Wilkinson, P. et al.
<2>Comparative genomics of the emerging human pathogen Photorhabdus asymbiotica with the insect pathogen Photorhabdus luminescens.
<3>BMC Genomics
<4>10
<5>302
<6>2009
<7>BACKGROUND: The Gram-negative bacterium Photorhabdus asymbiotica (Pa) has
been recovered from human infections in both North America and Australia.
Recently, Pa has been shown to have a nematode vector that can also infect
insects, like its sister species the insect pathogen P. luminescens (Pl).
To understand the relationship between pathogenicity to insects and humans
in Photorhabdus we have sequenced the complete genome of Pa strain
ATCC43949 from North America. This strain (formerly referred to as
Xenorhabdus luminescens strain 2) was isolated in 1977 from the blood of
an 80 year old female patient with endocarditis, in Maryland, USA. Here we
compare the complete genome of Pa ATCC43949 with that of the previously
sequenced insect pathogen P. luminescens strain TT01 which was isolated
from its entomopathogenic nematode vector collected from soil in Trinidad
and Tobago. RESULTS: We found that the human pathogen Pa had a smaller
genome (5,064,808 bp) than that of the insect pathogen Pl (5,688,987 bp)
but that each pathogen carries approximately one megabase of DNA that is
unique to each strain. The reduced size of the Pa genome is associated
with a smaller diversity in insecticidal genes such as those encoding the
Toxin complexes (Tc's), Makes caterpillars floppy (Mcf) toxins and the
Photorhabdus Virulence Cassettes (PVCs). The Pa genome, however, also
shows the addition of a plasmid related to pMT1 from Yersinia pestis and
several novel pathogenicity islands including a novel Type Three Secretion
System (TTSS) encoding island. Together these data suggest that Pa may
show virulence against man via the acquisition of the pMT1-like plasmid
and specific effectors, such as SopB, that promote its persistence inside
human macrophages. Interestingly the loss of insecticidal genes in Pa is
not reflected by a loss of pathogenicity towards insects. CONCLUSION: Our
results suggest that North American isolates of Pa have acquired virulence
against man via the acquisition of a plasmid and specific virulence
factors with similarity to those shown to play roles in pathogenicity
against humans in other bacteria.

<>

<1>Willcock, D.F., Dryden, D.T.F., Murray, N.E.
<2>A mutational analysis of the two motifs common to adenine methyltransferases.
<3>EMBO J.
<4>13
<5>3902-3908
<6>1994
<7>All methyltransferases that use S-adenosyl methionine as the methyl group donor contain a
sequence similar to (D/E/S)XFXGXG which has been postulated to form part of the cofactor
binding site. In N6-adenine DNA methyltransferases there is a second motif, (D/N)PP(Y/F),
which has been proposed to play a role similar to the catalytically essential PC motif
conserved in all C5-cytosine DNA methyltransferases. We have made a series of amino acid
changes in these two motifs in the EcoKI N6-adenine DNA methyltransferase. The mutant enzymes
have been purified to homogeneity and characterized by physical biochemical methods. The first
G is the most conserved residue in motif I. Changing this G to D completely abolishes
S-adenosyl methionine binding, but left enzyme structure and DNA target recognition unaltered,
thus documenting the S-adenosyl methionine binding function of motif I in N6-adenine
methyltransferases. Substitution of the N with D, or F with either G or C, in motif II
abolished enzyme activity, but left S-adenosyl methionine and DNA binding unaltered. Changes
of F to Y or W resulted in partial enzyme activity, implying that an aromatic residue is
important for methylation. The substitution of W for F greatly enhanced UV-induced
cross-linking between the enzyme and S-adenosyl methionine, suggesting that the aromatic
residue is close in space to the methyl-group donor.

<>

<1>Willems, A., Tian, R., Brau, L., Goodwin, L., Han, J., Liolios, K., Huntemann, M., Pati, A., Woyke, T., Mavrommatis, K., Markowitz, V., Ivanova, N., Kyrpides, N., Reeve, W.
<2>Genome sequence of Burkholderia mimosarum strain LMG 23256(T), a Mimosa pigra microsymbiont from Anso, Taiwan.
<3>Standards in Genomic Sciences
<4>9
<5>484-494
<6>2014
<7>Burkholderia mimosarum strain LMG 23256(T) is an aerobic, motile, Gram-negative,
non-spore-forming rod that can exist as a soil saprophyte or as a legume
microsymbiont of Mimosa pigra (giant sensitive plant). LMG 23256(T) was isolated
from a nodule recovered from the roots of the M. pigra growing in Anso, Taiwan.
LMG 23256(T) is highly effective at fixing nitrogen with M. pigra. Here we
describe the features of B. mimosarum strain LMG 23256(T), together with genome
sequence information and its annotation. The 8,410,967 bp high-quality-draft
genome is arranged into 268 scaffolds of 270 contigs containing 7,800
protein-coding genes and 85 RNA-only encoding genes, and is one of 100 rhizobial
genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic
Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

<>

<1>Williams, B.J., Golomb, M., Phillips, T., Brownlee, J., Olson, M.V., Smith, A.L.
<2>Bacteriophage HP2 of Haemophilus influenzae.
<3>J. Bacteriol.
<4>184
<5>6893-6905
<6>2002
<7>Temperate bacteriophages effect chromosomal evolution of their bacterial hosts, mediating
rearrangements and the acquisition of novel genes from other taxa. Although the Haemophilus
influenzae genome shows evidence of past phage-mediated lateral transfer, the phages presumed
responsible have not been identified. To date, six different H. influenzae phages are known;
of these, only the HP1/S2 group, which lyosogenizes exclusively Rd strains (which were
originally encapsulated serotype d), is well characterized. Phages in this group are
genetically very similar, with a highly conserved set of genes. Because the majority of H.
influenzae strains are nonencapsulated (nontypeable), it is important to characterize phages
infecting this larger, genetically more diverse group of respiratory pathogens. We have
identified and sequenced HP2, a bacteriophage of nontypeable H. influenzae. Although related
to the fully sequenced HP1 (and even more so to the partially sequenced S2) and similar in
genetic organization, HP2 has a few novel genes and differs in host range; HP2 will not infect
or lysogenize Rd strains. Genomic comparisons between HP1/S2 and HP2 suggest recent
divergence, with new genes completely replacing old ones at certain loci. Sequence comparisons
suggest that H. influenzae phages evolve by recombinational exchange of genes with each other,
with cryptic prophages, and with the host chromosome.

<>

<1>Williams, D.M., Benseler, F., Eckstein, F.
<2>Properties of 2'-fluorothymidine-containing oligonucleotides:  Interaction with restriction endonuclease EcoRV.
<3>Biochemistry
<4>30
<5>4001-4009
<6>1991
<7>2'-Fluorothymidine (Tf) was synthesized via an improved procedure with
(diethylamino) sulfur trifluoride.  The compatibility of the analogue with DNA
synthesis via the phosphoramidite method was demonstrated after complete
enzymatic digestion of the oligonucleotides d(TfuT) and d(Tf3T), the sole
products of which were 2'-fluorothymidine and thymidine in the expected ratio.
The 2'-fluorothymidine was also incorporated into the EcoRV recognition
sequence, within the complementary oligonucleotide d(CAAACCGATATCGTTGTG) and
d(CACAACGATATCGGTTTG).  Thermal melting characteristics of these duplexes
showed a significant decrease in stability only when both of the thymidine
residues in one of the strands were replaced.  In contrast, when all of one
strand of a duplex contained 2'-fluorothymidine, as in d(TfuT)-d(A12), a
substantially higher Tm and cooperativity of melting was observed relative to
the unmodified structure.  EcoRV cleaved a duplex that contained a
2'-fluorothymidine at the scissile linkage in each strand at two-thirds of the
rate obtained for the unmodified structure.  A duplex containing two
2'-fluorothymidine residues in one strand and none in the other was cleaved at
one-third of the rate in its unsubstituted strand, whereas the cleavage rate
was reduced to 22% in its modified strand.

<>

<1>Williams, K., Savageau, M.A., Blumenthal, R.M.
<2>A bistable hysteretic switch in an activator-repressor regulated restriction-modification system.
<3>Nucleic Acids Res.
<4>41
<5>6045-6057
<6>2013
<7>Restriction-modification (RM) systems are extremely widespread among bacteria and archaea, and
are often specified by mobile genetic elements. In type II RM systems, where the restriction
endonuclease (REase) and protective DNA methyltransferase (MTase) are separate proteins, a
major regulatory challenge is delaying expression of the REase relative to the MTase after RM
genes enter a new host cell. Basic understanding of this regulation is available for few RM
systems, and detailed understanding for none. The PvuII RM system is one of a large subset in
which the central regulatory role is played by an activator-repressor protein (called C, for
controller). REase expression depends upon activation by C, whereas expression of the MTase
does not. Thus delay of REase expression depends on the rate of C-protein accumulation. This
is a nonlinear process, as C also activates transcription of its own gene. Mathematical
modeling of the PvuII system led to the unexpected predictions of responsiveness to a factor
not previously studied in RM system control-gene copy number-and of a hysteretic response. In
this study, those predictions have been confirmed experimentally. The results may apply to
many other C-regulated RM systems, and help explain their ability to spread so widely.

<>

<1>Williams, L.E., Detter, C., Barry, K., Lapidus, A., Summers, A.O.
<2>Facile Recovery of Individual High-Molecular-Weight, Low-Copy-Number Natural Plasmids for Genomic Sequencing.
<3>Appl. Environ. Microbiol.
<4>72
<5>4899-4906
<6>2006
<7>Sequencing of the large (>50 kb), low-copy-number (<5 per cell) plasmids that mediate
horizontal gene transfer has been hindered by the difficulty and expense of isolating DNA from
individual plasmids of this class. We report here that a kit method previously devised for
purification of bacterial artificial chromosomes (BACs) can be adapted for effective
preparation of individual plasmids up to 220 kb from wild gram-negative and gram-positive
bacteria. Individual plasmid DNA recovered from less than 10 ml of Escherichia coli,
Staphylococcus, and Corynebacterium cultures was of sufficient quantity and quality for
construction of high-coverage libraries, as shown by sequencing five native plasmids ranging
in size from 30 kb to 94 kb. We also report recommendations for vector screening to optimize
plasmid sequence assembly, preliminary annotation of novel plasmid genomes, and insights on
mobile genetic element biology derived from these sequences. Adaptation of this BAC method for
large plasmid isolation removes one major technical hurdle to expanding our knowledge of the
natural plasmid gene pool.

<>

<1>Williams, M.L., Gillaspy, A.F., Dyer, D.W., Thune, R.L., Waldbieser, G.C., Schuster, S.C., Gipson, J., Zaitshik, J., Landry, C., Banes, M.M., Lawrence, M.L.
<2>Genome Sequence of Edwardsiella ictaluri 93-146, a Strain Associated with a Natural Channel Catfish Outbreak of Enteric Septicemia of Catfish.
<3>J. Bacteriol.
<4>194
<5>740-741
<6>2012
<7>Edwardsiella ictaluri is the cause of extensive mortalities and economic losses to the channel
catfish industry of the southeast United States.
Here we report the complete genome of Edwardsiella ictaluri 93-146.
Whole-genome sequence analysis of E. ictaluri provides a tool for
understanding the genomic regions specific to the species and the
Edwardsiella genus.

<>

<1>Williams, R.J.
<2>Restriction endonucleases and their uses.
<3>Methods Mol. Biol.
<4>160
<5>409-429
<6>2001
<7>Restriction endonucleases, which cleave DNA in a site-specific manner, are a fundamental tool
of molecular biology.  The discovery of endonucleases began in the 1960s and led to commercial
availability in the early 1970s.  The number of characterized enzymes continues to grow, as
does the number of vendors and the size of their product lines.  Although many similarities
exist among endonucleases in terms of their structures, mechanisms, and uses, important
differences remain.  Now a staple of molecular biology, restriction endonucleases are an area
of active research as models of site-specific DNA recognition, cleavage mechanism, in vivo
function, and evolutionary origins.  New enzymes continue to be discovered or developed by
using protein engineering to modify the specificity of existing enzymes.

<>

<1>Williams, R.J.
<2>Isolation and characterization of an unknown restriction endonuclease.
<3>Methods Mol. Biol.
<4>160
<5>431-442
<6>2001
<7>Currently, there are approximately 3000 restriction endonucleases known, recognizing 235
different sequences.  Although primarily found in bacteria, they also exist in archaea,
viruses, and eukaryotes.  An estimated 25% of bacteria examined contain at least one
restriction endonuclease, and therefore the probability of encountering new ones is relatively
high.  Indeed, many new enzymes have been "discovered" in contaminated bacterial cultures.
The presence of three restriction activities in a single organism is not unusual.  Neisseria
strains appear to be particularly rich in restriction endonucleases and their corresponding
methyltransferases.  As many as seven different endonucleases from a single strain have been
identified through cloning.  The first enzyme discovered which recognizes a unique sequence,
although it may not be commercially available or commonly known, is designated the prototype.
Although few new prototypes have been discovered recently, two potential four-base palindromes
and seven potential six-base palindromes are not cleaved by any known restriction
endonucleases.  Most databases are arranged alphabetically by prototype, with isoschizomers
listed under the prototype heading.  A database of all known endonucleases, maintained by Dr.
Richard J. Roberts, is available at http://www.neb.com/rebase.  A number of formats are
available, and references are provided.  Detailed information on restriction endonuclease
biology, classification, structure, specificity, and catalytic mechanism is provided elsewhere
in this book.

<>

<1>Williams, R.J.
<2>Restriction endonucleases: Classification, properties, and applications.
<3>Mol. Biotechnol.
<4>23
<5>225-243
<6>2003
<7>Restriction endonucleases have become a fundamental tool of molecular biology with many
commercial vendors and extensive product lines. While a
significant amount has been learned about restriction enzyme diversity,
genomic organization, and mechanism, these continue to be active areas of
research and assist in classification efforts. More recently, one focus
has been their exquisite specificity for the proper recognition sequence
and the lack of homology among enzymes recognizing the same DNA sequence.
Some questions also remain regarding in vivo function. Site-directed
mutagenesis and fusion proteins based on known endonucleases show promise
for custom-designed cleavage. An understanding of the enzymes and their
properties can improve their productive application by maintaining
critical digest parameters and enhancing or avoiding alternative
activities.

<>

<1>Williams, S.A., Halford, S.E.
<2>SfiI endonuclease activity is strongly influenced by the non-specific sequence in the middle of its recognition site.
<3>Nucleic Acids Res.
<4>29
<5>1476-1483
<6>2001
<7>The SfiI endonuclease cleaves DNA at the sequence GGCCNNNNNGGCC, where N is any base and  is
the point of cleavage.  Proteins that recognise discontinuous sequences in DNA can be affected
by the unspecified sequence between the specified base pairs of the target site. To examine
whether this applies to SfiI, a series of DNA duplexes were made with identical sequences
apart from discrete variations in the 5 bp spacer. The rates at which SfiI cleaved each duplex
were measured under steady-state conditions: the steady-state rates were determined by the DNA
cleavage step in the reaction pathway. SfiI cleaved some of these substrates at faster rates
than other substrates. For example, the change in spacer sequence from AACAA to AAACA caused a
70-fold increase in reaction rate. In general, the extrapolated values for kcat and Km were
both higher on substrates with inflexible spacers than those with flexible structures. The
dinucleotide at the site of cleavage was largely immaterial. SfiI activity is thus highly
dependent on conformational variations in the spacer DNA.

<>

<1>Williams, S.A., Halford, S.E.
<2>Communications between catalytic sites in the protein-DNA synapse by the SfiI endonuclease.
<3>J. Mol. Biol.
<4>318
<5>387-394
<6>2002
<7>The SfiI endonuclease is a tetrameric protein with two DNA-binding clefts. It has to bind two
copies of its recognition sequence, one at each cleft, before it cleaves DNA. While SfiI binds
cooperatively to two cognate sites, it binds only one non-cognate DNA molecule at a time and
the resultant complex is precluded from binding cognate DNA at the vacant cleft. To examine
the communications between separate binding sites in a protein that synapses two segments of
DNA, SfiI was tested with oligonucleotide duplexes containing its recognition sequence but
with either R(p) or S(p) phosphorothioate linkages at the scissile bonds. Though SfiI has low
activity on the R(p) and none against the S(p) diastereoisomer, it bound these duplexes in the
same cooperative manner as oxyester duplexes, though with a reduced affinity for the S(p)
derivative. It also formed complexes with one phosphorothioate-duplex and one oxyester-duplex
but, when Mg(2+) was added to the hybrid complexes, the phosphorothioate moiety at one
DNA-binding cleft prevented the enzyme from cleaving the oxyester duplex at the other cleft.
SfiI is thus restrained from catalytic action until it recognises the correct nucleotide
sequence at two DNA loci and the correct phosphodiester functions at both loci.

<>

<1>Williamson, M.R., Doherty, J.P., Woodcock, D.M.
<2>Modified-cytosine restriction-system-induced recombinant cloning artefacts in Escherichia coli.
<3>Gene
<4>124
<5>37-44
<6>1993
<7>We have tested whether, and to what extent, recombinant clones from DNA segments with
5-methylation of cytosines recovered in methylation-restriction (mcr+) hosts contain
mutations. We constructed a model system in which the tetracycline-resistance-encoding gene
(tet) from pBR322 was cloned into the plasmid pGEM3Zf+. The central regon of tet was removed
from the construct, methylated in vitro and then religated back into the unmethylated
remainder of the construct. The central region of tet was either (1) methylated with a
combination of four bacterial methyltransferases (M.AluI, M.HaeIII, M.HpaII plus M.HhaI) or
(2) methylated with M.SssI which methylates at all CpG dinucleotides. These two protocols
generated theoretical levels of DNA methylation in the central fragment of 10.5% and 33%
respectively. The construct was transformed into a series of isogenic (recA+) bacterial
strains that were mcrA+ mcrB+C+, mcrA+, mcrB-C+, mcrA-, mcrB+C+, mcrA-mcrB-C+ or mcrA- mcrBC,
and also into a set of isogenic recA- derivatives of these strains. With the two methylation
protocols, there was an average 48- and 141-fold reduction, respectively, in the number of
transformants recovered from the recA+ mcr+ hosts compared with a methylation-tolerant host
(mcr-). Of the clones recovered in recA+ mcr+ hosts >20% of clones had an inactivation g
mutation in tet. The majority of such mutant clones contained deletions that frequently
extended into the unmethylated portion of tet and even into the plasmid sequences beyond the
end of the polylinker. With the recA- mcr+ hosts, effective restriction was much more
stringent, rendering the plasmid containing the methylated segment effectively unclonable. By
implication, any collection of clones derived from methylated genomic DNA using a recA+ mcr+
host may contain significant frequencies of sequence artefacts.

<>

<1>Willis, D.B., Goorha, R., Granoff, A.
<2>DNA methyltransferase induced by frog virus 3.
<3>J. Virol.
<4>49
<5>86-91
<6>1984
<7>Over 20% of the cytosine bases in frog virus 3 DNA are methylated at the 5-carbon position. To
determine whether this high degree of methylation is the result of a virus-specific enzyme, we
examined the kinetics of induction and the substrate specificity of a DNA methyltransferase
from frog virus 3-infected fathead minnow cells.  A novel DNA methyltransferase activity
appeared in the cytoplasm of infected cells at 3 h postinfection.  This activity was induced
in the absence of viral DNA replication and was therefore probably an early viral enzyme.  In
contrast to the methyltransferase activity extracted from uninfected cell nuclei, the
cytoplasmic enzyme showed a strong template preference for double-stranded over
single-stranded and for unmethylated over hemimethylated DNA.  The dinucleotide sequence dCpdG
was a necessary and sufficient exogenous substrate for methylation in vitro.  A mutant of frog
virus 3, isolated as resistant to 5-azacytidine and having unmethylated virion DNA, did not
induce cytoplasmic DNA methyltransferase, leading to the conclusion that this activity is
coded for by the virus.

<>

<1>Wilson, B.D., Strauss, M., Stickells, B.J., Hoal-van Helden, E.G., van Helden, P.D.
<2>An assay for O6-alkylguanine-DNA alkyltransferase based on restriction endonuclease inhibition and magnetic bead separation of products.
<3>Carcinogenesis
<4>15
<5>2143-2148
<6>1994
<7>We describe an improved, rapid and cost effective assay for measuring O6-alkylguanine-DNA
alkyltransferase (AGT) levels in large numbers of small biological samples. The assay is based
on the use of a synthetic O6-methylguanine oligonucleotide bound to magnetic beads via a
biotin-streptavidin linkage and bound to its complement. A 35S label is incorporated into the
free end of the duplex. The basis of the assay lies in the observation that the restriction
enzyme PvuII will not cleave the bead-bound duplex oligonucleotide having an O6-methylguanine
within the restriction sequence. However, on removal of the methyl group by AGT present in
cell extracts prior to incubation with PvuII, the 35S-labelled oligonucleotide is cleaved by
the restriction enzyme, releasing a 35S-labelled 8 bp fragment. Due to the stoichiometric
nature of the AGT reaction, quantitation of AGT is easily achieved by counting an aliquot of
the supernatant after pelleting the unrestricted bead complexes with a magnet. AGT levels
measured in certain cell lines and human lymphocytes by the reported assay are comparable to
other methods. The assay can be performed in basic laboratories and allows for the rapid
processing of many samples simultaneously, which could prove useful in clinical and
epidemiological studies.

<>

<1>Wilson, G.A., Williams, M.T., Baney, H.W., Young, F.E.
<2>Characterization of temperate bacteriophages of Bacillus subtilis by the restriction endonuclease EcoRI: Evidence for three different temperate bacteriophages.
<3>J. Virol.
<4>14
<5>1013-1016
<6>1974
<7>Temperate bacteriophages of Bacillus subtilis were characterized according to
host range and digestion of the bacteriophage genome by endonuclease EcoRI.
The three bacteriophages, Phi3T, SPO2, and Phi105, were all heteroimmune, and
the DNA digests showed dissimilar patterns by agarose-ethidium bromide gel
electrophoresis.

<>

<1>Wilson, G.A., Young, F.E.
<2>Restriction and modification in the Bacillus subtilis genospecies:  Isolation of an endonuclease in Bacillus amyloliquefaciens.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>75
<5>103
<6>1975
<7>Previous studies in this laboratory have demonstrated that deoxyribonucleic
acid (DNA) is capable of effecting DNA-mediated transformation of B. subtilis.
Inefficiency of heterospecific transformation prompted a search for restriction
in the genospecies.  Accordingly mutants of SP02 (SP01clh2 and SP02clh5) were
isolated that infect B. amyloliquefaciens H.  When propagated on B.
amyloliquefaciens H, these viruses infected B. amyloliquefaciens H, but were
markedly restricted by B. subtilis 168.  Restriction was overcome by passage of
virus on B. subtilis 168.  Transfection of B. subtilis occurred with DNA
isolated from SP02clh2 propagated on B. subtilis but not with DNA isolated from
SP02clh2 propagated on B. amyloliquefaciens.  In the latter case biologic
activity could be recovered by superinfection marker rescue or transfection of
restriction deficient mutants of B. subtilis.  An endonuclease (BamHI) was
purified from B. amyloliquefaciens H by (NH4)2S04 precipitation and
chromatography on DEAE cellulose and phosphocellulose.  Analysis of cleavage
patterns of standard viral DNA preparations (lambda, SV40, adeno and SP02) by
gel electrophoresis established that BamHI is different from previously
described nucleases.  Biochemical tests indicate that BamHI recognized unique
sites in DNA and may be responsible for restriction in B. amyloliquefaciens.

<>

<1>Wilson, G.A., Young, F.E.
<2>Isolation of a sequence-specific endonuclease (BamI) from Bacillus amyloliquefaciens H.
<3>J. Mol. Biol.
<4>97
<5>123-125
<6>1975
<7>A restriction endonuclease has been isolated from Bacillus amyloliquefaciens H
(strain RUB500).  The enzyme, BamI, cleaves adenovirus-2 DNA at three sites,
phage lambda DNA at five sites, lambda plac DNA at four sites, Phi80 pt DNA at
14 sites, and Phi3T+ DNA at four sites.  However, it does not cleave DNA from
bacteriophage SP02, Phi105 or Phi29.

<>

<1>Wilson, G.A., Young, F.E.
<2>Restriction and modification in the Bacillus subtilis genospecies.
<3>Microbiology-1976, American Society for Microbiology, Schlessinger, D., Washington
<4>0
<5>350-357
<6>1976
<7>Restriction and modification have not been as intensively studied in
gram-positive bacilli as in enterobacteriaceae.  This is particularly
surprising since the Bacillus subtilis genospecies appears to be one in which
restriction and modification can be studied quite readily.  Included among the
advantages is the high frequency of genetic exchange among closely related
members of the genospecies.  This high rate of transformation usually occurs
with a limited number of genetic loci.  In addition, recent emphasis on the
genetic organization of B. subtilis has resulted in a detailed map of the
chromosome (Anagnostopoulos and Trowsdale, p. 44).  The presence of the
plasmids in closely related strains affords the possibility of using these
elements in restriction and cloning studies.  The variety of methods for
genetic exchange and the ease of transformation afford one of the best
opportunities to study restriction and modification of the isolated DNA.
Restriction and modification enzymes have been demonstrated in this
genospecies, and a number of these appear to be unique in the sequence of
nucleotides they recognize, thus increasing the usefulness of these enzymes in
the field of recombinant molecule technology.  The bacilli also undergo a
carefully regulated series of morpho-genetic events leading to the production
of an endospore.  Finally, unlike the enterobacteriaceae, bacilli have been the
organisms of choice for the production of a large number of fermentation
products.  Because of the desire to study morphogenesis and to construct
strains of commercial importance, an examination of the factors contributing
toward (or hampering) efficient genetic exchange within the genospecies has
been intensified.

<>

<1>Wilson, G.A., Young, F.E.
<2>Purification and properties of the BamHI endonuclease.
<3>Methods Enzymol.
<4>65
<5>147-153
<6>1980
<7>Site-specific endonucleases have been isolated from a diverse variety of microorganisms and
used extensively in analyzing and manipulating complex genomes.  In fact, the utility of these
enzymes has resulted in an emphasis on their application and inadvertently hampered a thorough
examination of the properties and biological functions of these endonucleases.  For example,
the role of site-specific endonucleases in vivo has not been determined, although, restriction
and modification properties have been confirmed by genetic studies for EcoRI, EcoRII, BamHI,
BsuI, Bst1503, BglI, and BglII.  Although the methods of isolation and optimal reaction
conditions have not been systematically examined, most of the enzymes can be readily isolated
and purified from contaminating endonucleases and exonucleases.  BamHI proved to be a useful
enzyme because it produced a single-stranded end after cleaving between the guanine bases at
the site of 5'-GGATCC-3', thus providing a useful substrate for ligation.  In addition, a
number of cloning vectors existed or were developed that contained a single recognition site
for BamHI including vectors SV40, pMB9, pBR313, and pBR322.  These advantages contributed to
the extensive use of this enzyme and to the development of modifications of our original
purification procedure that were kindly communicated to us by numerous investigators.  This
report will summarize the methods of isolation of BamHI and related enzymes from the Bacillus
genospecies as well as indicate what progress has been made in characterizing these enzymes.

<>

<1>Wilson, G.G.
<2>Type II restriction-modification systems.
<3>Trends Genet.
<4>4
<5>314-318
<6>1988
<7>Restriction-modification (R-M) systems are enzymatic mechanisms that occur
primarily in bacteria.  Their principal function is defensive:  they enable
cells to resist infections by phage and plasmid DNA molecules that would
otherwise parasitize them, and they limit the spread of these molecules within
the population.  R-M systems are traditionally classified into three groups,
types I, II and III, according to their enzymology and cofactor requirements.
Type II systems are best understood, and are regarded as the simplest; they are
also the most familiar to molecular biologists since it is from these systems
that the restriction enzymes used in recombinant DNA research are purified.  A
summary of the biology of the type II systems is presented here; the type I and
type II systems have been reviewed elsewhere.

<>

<1>Wilson, G.G.
<2>Cloned restriction-modification systems - a review.
<3>Gene
<4>74
<5>281-289
<6>1988
<7>The genes for numerous restriction endonucleases and modification methylases
have been cloned into Escherichia coli.  A summary is given for the clones
isolated so far (115 entries) and of the procedures used to obtain them.

<>

<1>Wilson, G.G.
<2>Organization of restriction-modification systems.
<3>Nucleic Acids Res.
<4>19
<5>2539-2566
<6>1991
<7>The genes for over 100 restriction-modification systems have now been cloned,
and approximately one-half have been sequenced.  Despite their similar
function, they are exceedingly heterogeneous.  The heterogeneity is evident at
three levels: in the gene arrangements; in the enzyme compositions; and in the
protein sequences.  This paper summarizes the main features of the R-M systems
that have been cloned.

<>

<1>Wilson, G.G.
<2>Amino acid sequence arrangements of DNA-methyltranferases.
<3>Methods Enzymol.
<4>216
<5>259-279
<6>1992
<7>
<>

<1>Wilson, G.G.
<2>Cloning the HinfI restriction and modification genes.
<3>US Patent Office
<4>US 5215906
<5>
<6>1993
<7>Methods for cloning restriction enzymes and their corresponding modification enzymes by
selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent
screening to identify those clones containing the restriction gene.

<>

<1>Wilson, G.G.
<2>Cloning the TaqI restriction and modification genes.
<3>US Patent Office
<4>US 5208157
<5>
<6>1993
<7>Methods for cloning restriction enzymes and their corresponding modification enzymes by
selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent
screening to identify those clones containing the restriction gene.

<>

<1>Wilson, G.G.
<2>Cloning restriction and modification genes.
<3>US Patent Office
<4>US 5200333
<5>
<6>1993
<7>Methods for cloning restriction enzymes and their corresponding modification enzymes by
selecting clones resistant to digestion by the restriction enzyme and subsequent selection of
clones containing the restriction gene.

<>

<1>Wilson, G.G.
<2>Cloning the BanI restriction and modification genes.
<3>US Patent Office
<4>US 5198354
<5>
<6>1993
<7>Methods for cloning restriction enzymes and their corresponding modification enzymes by
selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent
screening to identify those clones containing the restriction gene.

<>

<1>Wilson, G.G.
<2>Cloning the HaeII restriction and modification genes.
<3>US Patent Office
<4>US 5196332
<5>
<6>1993
<7>Methods for cloning restriction enzymes and their corresponding modification enzymes by
selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent
screening to identify those clones containing the restriction gene.

<>

<1>Wilson, G.G., Meda, M.M.
<2>Heterospecific modification as a means to clone restriction genes.
<3>US Patent Office
<4>US 5179015
<5>
<6>1993
<7>The present invention provides a novel approach to the production of restriction enzymes. More
specifically, there is provided a novel method for cloning these enzymes, which comprises
preparing DNA libraries from the DNA of an organism that synthesizes the restriction enzyme of
interest, creating a suitable host containing a heterospecific methyltransferase gene able to
protectively modify DNA from digestion by the restriction enzyme of interest, introducing the
DNA libraries into the protectively modified host, and screening recombinant organisms to
identify those carrying the desired restriction enzyme gene. The application of this method to
the FspI and HaeIII restriction genes of Fischerella species and Haemophilius aegyptius,
respectively, is described in detail, together with the resulting clones that form the basis
of a new and useful process for purifying the FspI and HaeIII restriction enzymes.

<>

<1>Wilson, G.G., Meda, M.M.
<2>Heterospecific modification as a means to clone restriction genes.
<3>US Patent Office
<4>US 5246845
<5>
<6>1993
<7>The present invention provides a novel approach to the production of restriction enzymes. More
specifically, there is provided a novel method for cloning these enzymes, which comprises
preparing DNA libraries from the DNA of an organism that synthesizes the restriction enzyme of
interest, creating a suitable host containing a heterospecific methyltransferase gene able to
protectively modify DNA from digestion by the restriction enzyme of interest, introducing the
DNA libraries into the protectively modified host, and screening recombinant organisms to
identify those carrying the desired restriction enzyme gene. The application of this method to
the FspI and HaeIII restriction genes of Fischerella species and Haemophilus aegyptius,
respectively, is described in detail, together with the resulting clones that form the basis
of a new and useful process for purifying the FspI and HaeIII restriction enzymes.

<>

<1>Wilson, G.G., Murray, N.E.
<2>Restriction and Modification Systems.
<3>Annu. Rev. Genet.
<4>25
<5>585-627
<6>1991
<7>*
CONTENTS
INTRODUCTION
BIOLOGY OF RESTRICTION AND MODIFICATION
   Restriction and modification enzymes
   Occurrence of R-M systems
   Restriction and modification of viruses
   Cloning restriction and modification genes
CHARACTERISTICS OF R-M SYSTEMS
   Type I systems
   Type II systems
   Type IIs systems
   Type III systems
   Other system types
   Modification-requiring systems
   Regulation of expression
CONTRASTS AND COMPARISONS AMONG R-M SYSTEMS
   Type I systems
   Type II systems
   Type II endonucleases
   Methyltransferases
DISCUSSION


<>

<1>Wilson, G.G., Nwankwo, D.
<2>Cloning the MspI restriction and modification genes.
<3>US Patent Office
<4>US 5196331
<5>
<6>1993
<7>Methods for cloning restriction enzymes and their corresponding modification enzymes by
selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent
screening to identify those clones containing the restriction gene.

<>

<1>Wilson, G.G., Nwankwo, D.
<2>Cloning restriction and modification genes.
<3>US Patent Office
<4>US 5180673
<5>
<6>1993
<7>Methods for cloning restriction enzymes and their corresponding modification enzymes by
selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent
screening to identify those clones containing the restriction gene.

<>

<1>Wilson, G.G., Wang, H., Heiter, D.F., Lunnen, K.D.
<2>Restriction enzymes in microbiology, biotechnology and biochemistry.
<3>Encuentro
<4>93
<5>19-48
<6>2012
<7>Since their discovery in the nineteen-seventies, a collection of simple enzymes termed Type II
restriction endonucleases, made by microbes to ward off viral infections, have transformed
molecular biology, spawned the multi-billion dollar Biotechnology industry, and yielded
fundamental insights into the biochemistry of life, health and disease.  In this article we
describe how these enzymes were discovered, and we review their properties, organizations and
genetics.  We summarize current ideas about the mechanism underlying their remarkable ability
to recognize and bind to specific base pair sequences in DNA, and we discuss why these ideas
might not be correct.  We conclude by proposing an alternative explanation for
sequence-recognition that resolves certain inconsistencies and provides, in our view, a more
satisfactory account of the mechanism.

<>

<1>Wilson, G.W., Edgell, D.R.
<2>Phage T4 mobE promotes trans homing of the defunct homing endonuclease I-TevIII.
<3>Nucleic Acids Res.
<4>37
<5>7110-7123
<6>2009
<7>Homing endonucleases are site-specific DNA endonucleases that typically function as mobile
genetic elements by introducing a double-strand break
(DSB) in genomes that lack the endonuclease, resulting in a unidirectional
gene conversion event that mobilizes the homing endonuclease gene and
flanking DNA. Here, we characterize phage T4-encoded mobE, a predicted
free-standing HNH family homing endonuclease. We show that mobE is
promoterless and dependent on upstream transcription for expression, and
that an internal intrinsic terminator regulates mobE transcript levels.
Crucially, in vivo mapping experiments revealed a MobE-dependent,
strand-specific nick in the non-coding strand of the nrdB gene of phage
T2. An internal deletion of the predicted HNH catalytic motif of MobE
abolishes nicking, and reduces high-frequency inheritance of mobE.
Sequence polymorphisms of progeny phage that inherit mobE are consistent
with DSB repair pathways. Significantly, we found that mobility of the
neighboring I-TevIII, a defunct homing endonuclease encoded within a group
I intron interrupting the nrdB gene of phage T4, was dependent on an
intact mobE gene. Thus, our data indicate that the stagnant nrdB intron
and I-TevIII are mobilized in trans as a consequence of a MobE-dependent
gene conversion event, facilitating persistence of genetic elements that
have no inherent means of promoting their own mobility.

<>

<1>Wilson, J.G., French, W.T., Lipzen, A., Martin, J., Schackwitz, W., Woyke, T., Shapiro, N., Bullard, J.W., Champlin, F.R., Donaldson, J.R.
<2>Draft Genome Sequence of Enterobacter cloacae Strain JD6301.
<3>Genome Announcements
<4>2
<5>e00381-14
<6>2014
<7>Enterobacter cloacae strain JD6301 was isolated from a mixed culture with wastewater collected
from a municipal treatment facility and oleaginous
microorganisms. A draft genome sequence of this organism indicates that it has a
genome size of 4,772,910 bp, an average G+C content of 53%, and 4,509
protein-coding genes.

<>

<1>Wilson, R. et al.
<2>2.2 Mb of contiguous nucleotide sequence from chromosome III of C. elegans.
<3>Nature
<4>368
<5>32-38
<6>1994
<7>As part of our effort to sequence the 100-megabase (Mb) genome of the nematode Caenorhabditis
elegans, we have completed the nucleotide sequence of a contiguous 2,181,032 base pairs in the
central gene cluster of chromosome III. Analysis of the finished sequence has indicated an
average density of about one gene per five kilobases; comparison with the public sequence
databases reveals similarities to previously known genes for about one gene in three. In
addition, the genomic sequence contains several intriguing features, including putative gene
duplications and a variety of other repeats with potential evolutionary implications. [
Comment in: Nature 1994 Mar 3;368(6466):14-5 ]

<>

<1>Wilson, W.W., Hoffman, R.M.
<2>Methylation of intact chromosomes by bacterial methylases in agarose plugs suitable for pulsed-field electrophoresis.
<3>Anal. Biochem.
<4>191
<5>370-375
<6>1990
<7>Conditions were determined for the methylation of intact yeast chromosomes by
EcoRI, HhaI, and MspI bacterial methylases using an endonuclease protection
assay while the chromosomes were embedded in agarose plugs suitable for
transverse-field electrophoresis.  Parameters were also established for the
methylation of human chromosomes by EcoRI methylase.  Methylation of embedded
chromosomes by EcoRI methylase required prewashes with EDTA.  EcoRI, HhaI, and
MspI methylases showed optimal activity when nonacetylated bovine serum
albumin, high levels of S-adenosylmethionine, and high levels of methylase were
used.  The use of bacterial methylases for methylation of embedded chromosomes
will allow investigators to normalize variations in cellular DNA methylation
prior to restriction and create new and rare endonuclease recognition sites
which will facilitate the detection of chromosomal alterations and deletions.

<>

<1>Wilson, W.W., Mebane, E.W., Hoffman, R.M.
<2>Creation of ultra-rare restriction sites in intact eucaryotic chromosomes mediated by bacterial methylases: An approach to sequencing and analyzing tumor and normal genomes.
<3>Anticancer Res.
<4>13
<5>17-20
<6>1993
<7>The limited restriction of eucaryotic chromosomes would facilitate our understanding of the
aberrant genomes of genetic diseases and cancer. We have described methods for methylating
eucaryotic chromosomes embedded in agarose plugs (1). We now describe how ClaI methylase can
be utilized in a methylation-dependent restriction cleavage with DpnI to restrict eucaryotic
genomes into a limited number of fragments. We have restricted the genomes of Saccharomyces
cerevisiae at four sites and cleaved the three chromosomes of Schizosaccharomyces pombe into
eleven fragments. Unlike the recently published methodologies (2,3,4,5) using DNA sequences
inserted into procaryotic and eucaryotic genomes, the methodologies described here use
unaltered eucaryotic genomes for highly limited restriction. This methodology has applications
in speeding, simplifying, and reducing the cost of sequence the human genome.

<>

<1>Winckler, K., Bach, B., Obe, G.
<2>Survival of Saccharomyces cerevisiae after treament with the restriction endonuclease AluI.
<3>Int. J. Radiat. Biol.
<4>54
<5>563-566
<6>1988
<7>Treatment of yeast cells proficient in the repair of radiation damage
(Saccharomyces cervisiae) with the restriction endonuclease AluI leads to a
positive dose-effect relationship between inactivation level and enzyme
concentration.  The data suggest an uptake of the active restriction enzyme
into the cells and a relationship between induction of DNA double-strand breaks
and cell killing.

<>

<1>Windbichler, N., Schroeder, R.
<2>Double duty.
<3>Nat. Struct. Mol. Biol.
<4>11
<5>910-911
<6>2004
<7>A second high-affinity binding site for the I-TevI homing endonuclease has been discovered.
Surprisingly, the DNA sequence recognized is the
protein's own operator; at this site, the endonuclease represses its
own transcription instead of cleaving the DNA and inducing intron
homing.

<>

<1>Windolph, S., Alves, J.
<2>Influence of divalent cations on inner-arm mutants of restriction endonuclease EcoRI.
<3>Eur. J. Biochem.
<4>244
<5>134-139
<6>1997
<7>To understand the functional role of the inner arm region of EcoRI for the interaction with
its DNA substrate, we mutated each of the positively charged amino acid residues Lys130 and
Arg131 to Ala or Glu.  The resulting cleavage activities are only about tenfold reduced but
DNA binding in the absence of divalent cations is totally disrupted for three of the mutants
([Ala130]EcoRI, [Glu130]EcoRI and [Glu131]EcoRI).  The binding can be rescued by adding 1 mM
Ca2+.  This binding behavior resembles that of the blunt end cutter EcoRV.  Furthermore, the
cleavage activity of the [Glu130]EcoRI is stimulated by Ca2+.  Therefore, a second metal
binding site is involved in [Glu130]EcoRI catalyzed DNA cleavage.  Its location is postulated
to be in the inner arm region at positions 133 and 135 presenting an Asp-Xaa-Asp motif typical
for Ca2+ binding.  A new positive charge at the tip of the inner arm due to the basic amino
acids Lys130 and Arg131 in the wild-type enzyme or due to binding of a divalent cation in the
mutants is important for DNA recognition by EcoRI.  However, a direct phosphate contact
providing an indirect readout is not observed.  Implications of the binding and cleavage
behavior with regard to mechanistic differences between blunt end and sticky end cutters and a
general catalytic mechanism of restriction enzymes are discussed.

<>

<1>Windolph, S., Fritz, A., Oelgeschlager, T., Wolfes, H., Alves, J.
<2>Sequence context influencing cleavage activity of the K130E mutant of the restriction endonuclease EcoRI identified by a site selection assay.
<3>Biochemistry
<4>36
<5>9478-9485
<6>1997
<7>We have generated several EcoRI mutants which exhibit a decreased cleavage rate on one of the
five specific cleavage sites in bacteriophage lambda-DNA.  To study the influence of the
sequence context on the cleavage rate in more detail, we developed a site selection assay.
From a complete set of 4096 plasmid substrates, differing in three bases on both sides of a
recognition sequence, optimal (best cut) and unfavorable (worst cut) sequences were selected
by repeated limited digestion, separation, and in vivo amplification of cleaved and uncleaved
plasmids.  In order to compare the sequence preferences of the inner arm mutant K130E and the
wild type enzyme, the cleavage rates and sequences of individual plasmids from the resulting
pools were determined.  The inner arm mutant K130E selected pools with clearly defined
consensus sequences and a high amount of palindromic sequences.  The cleavage rates of the
selected sequences are specific for the K130E mutant as is shown by their cleavage with other
mutants.  In contrast, wild type EcoRI does not lead to a selection in this assay.  Pre-steady
state kinetics show that preferences for a certain sequence context are a result of
differences in the dissociation rates of the wild type enzyme.  EcoRI is evolved to
efficiently recognize and cleave each nonmethylated DNA invading the cell.  Therefore, a fast
dissociation after cleavage is not mandatory.

<>

<1>Winegar, R.A., Land, M.C., Morgan, W.F.
<2>Increased chromosomal radiosensitivity of a Chinese hamster ovary cell line that inducibly expresses the EcoRI restriction endonuclease.
<3>Biochem. Biophys. Res. Commun.
<4>160
<5>1079-1084
<6>1989
<7>We have transfected a Chinese hamster ovary cell line (CHO 6) with a plasmid
that inducibly expresses the EcoRI restriction endonuclease gene in the
presence of cadmium sulfate (CdSO4).  Expression of EcoRI results in DNA
double-strand breaks, which can lead to chromosome aberrations.  The new line,
designated CHO 10, also has a low level of constitutive expression of EcoRI in
the absence of CdSO4 without any cytogenetic effect.  This suggested that these
cells may be efficient at repairing low levels of DNA double-strand breaks.  To
test this, both cell lines were exposed to ionizing radiation, and aberration
yields were analyzed with or without induction of EcoRI.  CHO 10 cells showed
increased radiosensitivity after G1 irradiation, but after G2 exposure, only
doses > 0.4 Gy caused more damage in CHO 10 cells.  We conclude that CHO 10
cells can tolerate constitutive expression of EcoRI, but that when the cells
are subjected to additional stress, in this case ionizing radiation, they
become very sensitive to DNA double-strand breaks.

<>

<1>Winegar, R.A., Phillips, J.W., Youngblom, J.H., Morgan, W.F.
<2>Cell electroporation is a highly efficient method for introducing restriction endonucleases into cells.
<3>Mutat. Res.
<4>225
<5>49-53
<6>1989
<7>Restriction endonucleases that make either blunt- or cohesive-end DNA
double-strand breaks can induce chromosome aberrations.  We have used cell
electroporation with great success to permeabilize Chinese hamster ovary cells
for the introduction of restriction enzymes.  The introduction of restriction
enzymes by this method resulted in extremely high frequences (>90%) of aberrant
metaphase cells and also a dramatic decrease in cell survival, as measured by
subsequent colony formation.  Cell electroporation by itself caused no increase
in aberrant chromosomes and had only a slight effect on cell survival.

<>

<1>Wines, D.R., Talbert, P.B., Clark, D.V., Henikoff, S.
<2>Introduction of a DNA methyltransferase into Drosophila to probe chromatin structure in vivo.
<3>Chromosoma
<4>104
<5>332-340
<6>1996
<7>The dam DNA methyltransferase gene from Escherichia coli was introduced into Drosophila in
order to probe chromatin structure in vivo.  Expression of the gene caused no visible defects
or developmental delay even at high levels of active methylase.  About half of each target
site was found to be methylated in vivo, apparently reflecting a general property of chromatin
packaged in nucleosomes.  Although site-specific differences were detected, most euchromatic
and heterochromatic sites showed comparable degrees of methylation, at least at high methylase
levels.  Methylase accessibility of a lacZ reporter gene subject to position-effect
variegation throughout development was only slightly reduced, consistent with studies of
chromatin accessibility in vitro.  Silencing of lacZ during development differed from
silencing of an adjacent white eye pigment reporter gene in the adult, as though chromatin
structure can undergo dynamic alterations during development.

<>

<1>Winget, D.M., Wommack, K.E.
<2>Randomly amplified polymorphic DNA PCR as a tool for assessment of marine viral richness.
<3>Appl. Environ. Microbiol.
<4>74
<5>2612-2618
<6>2008
<7>Recent discoveries have uncovered considerable genetic diversity among
aquatic viruses and raised questions about the variability of this
diversity within and between environments. Studies of the temporal and
spatial dynamics of aquatic viral assemblages have been hindered by the
lack of a common genetic marker among viruses for rapid diversity
assessments. Randomly amplified polymorphic DNA (RAPD) PCR bypasses this
obstacle by sampling at the genetic level without requiring viral
isolation or previous sequence knowledge. In this study, the utility of
RAPD-PCR for assessing DNA viral richness within Chesapeake Bay water
samples was evaluated. RAPD-PCR using single 10-mer oligonucleotide
primers successfully produced amplicons from a variety of viral samples,
and banding patterns were highly reproducible, indicating that each band
likely represents a single amplicon originating from viral template DNA.
In agreement with observations from other community profiling techniques,
resulting RAPD-PCR banding patterns revealed more temporal than spatial
variability in Chesapeake Bay virioplankton assemblages. High-quality
hybridization probes and sequence information were also easily generated
from single RAPD-PCR products or whole reactions. Thus, the RAPD-PCR
technique appears to be practical and efficient for routine use in
high-resolution viral diversity studies by providing assemblage
comparisons through fingerprinting, probing, or sequence information.

<>

<1>Winkle, S.A., Aloyo, M.C., Morales, N., Zambrano, T.Y., Sheardy, R.D.
<2>Enhanced reactivity of a B-Z junction for cleavage by the restriction enzyme MboI.
<3>Biochemistry
<4>30
<5>10601-10606
<6>1991
<7>We have been investigating the structure, dynamics, and ligand-binding
properties of the interface that exists between a right-handed conformation and
a left-handed conformation (i.e., a B-Z junction) in synthetic DNA oligomers.
Since exo- and endonuclease activity is known to be sensitive to the
conformation of the template DNA, we have designed and synthesized a DNA
oligonucleotide of 20 base pairs (designated as BZ-III) with an MboI
recognition site (GATC) at the location of a potential B-Z junction.  The
activity of the MboI enzyme toward this molecule and DNA oligomers that contain
multiple MboI sites located at B-Z junctions was monitored in the absence and
presence of the Z-conformation-inducing reagent cobalt hexaammine.  In all
cases, the activity of the enzyme was enhanced in the presence of cobalt
hexaammine.  The activity of MboI toward BZ-III, in the presence and absence of
cobalt hexaammine, was also examined when the DNA oligomer is also in the
presence of the DNA binding drugs actinomycin D, ametantrone, or ethidium
bromide.  In all cases, the activity of the enzyme was inhibited in the
presence of drug.  The results suggest that B-Z junctions are structurally
unique and that this uniqueness may alter nuclease activity at sites in or near
the junction.

<>

<1>Winkle, S.A., Aloyo, M.C., Smoller, J.H., Herrera, J.E., Chaires, J.B., Sheardy, R.D.
<2>Junctions affect the sensitivity of DNAs to reactions with enzymes.
<3>FASEB J.
<4>6
<5>A219
<6>1992
<7>We have investigated the reactions of various enzymes with DNAs possessing junctions, for
example: BZI C*GC*GC*GC*GACTGACTG; BZIII ATC*GC*GC*GC*GATCAGTCAGT; BZIV TCGACGCGCGCGATCAGTCA
(where C* is 5-methylC and where potential junctions are underlined). DNase I and Exonuclease
III show inhibited cleavage at the junctions both under low salt B conditions and in the
presence of Z-inducing cobalt hexamine. Restriction enzymes attacking either the junction or,
when BZ IV is inserted into a plasmid, sequences flanking the inserted BZ IV sequence show
enhanced cleavage in the presence of cobalt hexamine or supercoiling. Dam methylase has
enhanced reactivity for the GATC at the junction of BZ IV in the presence of cobalt hexamine.
These results suggest that these junctions have a structural distinctiveness recognizable by
various enzyme types.

<>

<1>Winkle, S.A., Thomson, H., Aguilar, L.P., Sheardy, R.D.
<2>The effects of alternate DNA structures on restriction enzyme and polymerase binding and activities with DNAS.
<3>Biophys. J.
<4>82
<5>463a
<6>2002
<7>The variety of structural motifs which DNA molecules may assume, depending upon sequence,
affects the activities of proteins using DNA as a substrate.  We have examined the effects of
the (CG)3 segment and cruciform structures found in phiX174 RF DNA (in the linear and
supercoiled forms) on E. coli DNA polymerase I binding and activity.  Gel mobility shift
assays of DNA fragments containing these features suggest that DNA polymerase binds to these
structural features.  Polymerase activity studies, including studies under PCR conditions,
indicate that the structures alter the activity.  Cleavage studies at or near these structures
by restriction enzymes, e.g., BssHII, HhaI, also show anomalies.  Binding of restriction
enzymes and DNA polymerase I to small DNA hairpins was also investigated using gel mobility
shift assays.  The results of these studies suggest that local structures do affect the
workings of enzymes and thus these structures may help regulate enzyme activities.

<>

<1>Winkler, F.K.
<2>DNA totally flipped-out by methylase.
<3>Structure
<4>2
<5>79-83
<6>1994
<7>In research, surprises are always refreshing and stimulating. The latest such example in the
field of protein-DNA interactions is provided by Klimasauskas et al. reporting the crystal
structure determination of the enzyme HhaI methylase from Haemophilus haemolyticus, a DNA
cytosine-5-methyltransferase (m5C-MTase), covalently bound to a 13-mer DNA duplex (Fig.1). The
covalent complex represents a chemically trapped reaction intermediate and also contains
S-adenosyl-L-methionine (AdoMet). The most spectacular and unexpected feature of this
structure is that the target cytosine base is completely flipped out of the DNA duplex and in
its place the DNA is infiltrated by a glutamine and a serine side chain from the major and
minor groove side respectively. Just as we are becoming accustomed to drastic amounts of
bending, kinking, unwinding and base-pair unstacking, most notably by the TATA box binding
protein or the endonucleases EcoRI and EcoRV, this enzyme goes even further by forcing its
substrate DNA to flip out the base it is going to methylate.

<>

<1>Winkler, F.K.
<2>Restriction endonucleases, the ultimate in sequence specific DNA recognition.
<3>J. Mol. Recognit.
<4>6
<5>9
<6>1994
<7>X-ray crystallographic and NMR spectroscopic studies with a variety of proteins that interact
in a sequence-specific manner with DNA have revealed a wealth of detailed structural
information over the past few years. It has become clear that DNA sequence recognition or
discrimination can be achieved in many different ways and that conformational changes in
protein and DNA often accompany the binding. Among the systems studied, mostly DNA binding
proteins involved in transcriptional control, restriction endonucleases demonstrate the
highest discrimination between cognate and noncognate DNA sequences. The structures of two
type II restriction endonucleases, EcoRI and EcoRV recognizing the hexamer sequences GAATTC
and GATATC respectively, have thus far been determined as complexes with cognate DNA
fragments. The structural and functional data now available for these two enzymes have yielded
important insights into how they achieve their remarkable specificity. The two enzymes show no
structural similarity except in their active site region where the arrangement of two acidic
and one basic residue appears conserved. Both enzymes bind their cognate DNA fragments in
rather distorted conformations but the distortions are completely different in the two cases.
Recognition involves the formation of hydrogen bonds to the base pairs in the major groove and
the extended set of interactions appears highly cooperative. However, the distorted
conformation of productively bound DNA plays an important role too. It provides extra
discrimination against noncognate sequences and appears crucial in coupling proper recognition
to efficient catalysis.

<>

<1>Winkler, F.K.
<2>Structure and function of restriction endonucleases.
<3>Curr. Opin. Struct. Biol.
<4>2
<5>93-99
<6>1992
<7>New insight into how restriction endonucleases achieve their remarkable sequence specificity
has been gained by comparing the structures of EcoRI and EcoRV endonucleases complexed with
DNA fragments that contain their respective canonical sites. Despite the lack of overall
structural homology, essential features of the catalytic site, including the locations of two
acidic and one lysine residue, are structurally conserved. Mutagenesis studies, newly
initiated for the EcoRV enzyme, have confirmed the essential nature of these particular
residues but have also yielded interesting effects that were quite unforeseen.

<>

<1>Winkler, F.K., Banner, D.W., Oefner, C., Tsernoglou, D., Brown, R.S., Heathman, S.P., Bryan, R.K., Martin, P.D., Petratos, K., Wilson, K.S.
<2>The crystal structure of EcoRV endonuclease and of its complexes with cognate and non-cognate DNA fragments.
<3>EMBO J.
<4>12
<5>1781-1795
<6>1993
<7>The crystal structure of EcoRV endonuclease has been determined at 2.5 A resolution and that
of its complexes with the cognate DNA decamer GGGATATCC (recognition sequence underlined) and
the non-cognate DNA octamer CGAGCTCG at 3.0 A resolution. Two octamer duplexes of the
non-cognate DNA, stacked end-to-end, are bound to the dimeric enzyme in B-DNA like
conformations. The protein-DNA interactions of this complex are prototypic for non-specific
DNA binding. In contrast, only one cognate decamer duplex is bound and deviates considerably
from canonical B-form DNA. Most notably, a kink of approximately 50 degrees is observed at the
central TA step with a concomitant compression of the major groove. Base-specific hydrogen
bonds between the enzyme and the recognition base pairs occur exclusively in the major groove.
These interactions appear highly co-operative as they are all made through one short surface
loop comprising residues 182-186. Numerous contacts with the sugar phosphate backbone
extending beyond the recognition sequence are observed in both types of complex. However, the
total surface area buried on complex formation is >1800 A2 larger in the case of cognate DNA
binding. Two acidic side chains, Asp74 and Asp90, are close to the reactive phosphodiester
group in the cognate complex and most probably provide oxygen ligands for binding the
essential cofactor Mg2+. An important role is also indicated for Lys92, which together with
the two acidic functions appears to be conserved in the otherwise unrelated structure of EcoRI
endonuclease. The structural results give new insight into the physical basis of the
remarkable sequence specificity of this enzyme.

<>

<1>Winkler, F.K., Brown, R.S., Leonard, K., Berriman, J.
<2>Structural studies of EcoRV endonuclease and of its complexes with short DNA fragments.
<3>Nato Advanced Studies: Crystallography in Molecular Biology, Plenum Press, Moras, D., Drenth, J., Standberg, B., Suck, D., Wilson, K., New York
<4>0
<5>345-352
<6>1987
<7>None

<>

<1>Winkler, F.K., D'Arcy, A., Blocker, H., Frank, R., van Boom, J.H.
<2>Crystallization of complexes of EcoRV endonuclease with cognate and non-cognate DNA fragments.
<3>J. Mol. Biol.
<4>217
<5>235-238
<6>1991
<7>Complexes of the type II restriction endonuclease EcoRV with a variety of
short, self-complementary deoxyoligonucleotides have been crystallized.  The
best crytals diffract to about 2.7 angstrom resolution and consist of 1:1
complexes between endonuclease dimers and duplexes of the cognate decamer
GGGATATCCC containing the hexameric EcoRV recognition sequence GATATC.
Crystals with the non-cognate DNA octamer duplexes CGAGCTCG and CGAATTCG
diffract to 3.0 and 3.5 angstrom resolution, respectively, and contain two DNA
duplexes per enzyme dimer.

<>

<1>Winkler, F.K., Prota, A.E.
<2>Structure and function of EcoRV endonuclease.
<3>Nucleic Acids Mol. Biol.
<4>14
<5>179-214
<6>2004
<7>The homodimeric, orthodox Type II restriction endonuclease EcoRV cleaves double-stranded DNA
with high specificity at hexameric GAT/ATC sites in a blunt-ended fashion.  Like most of the
restriction enzymes used as tools in recombinant DNA technology it can be considered as a
simple hydrolytic enzyme of relatively small size with precisely defined substrate specificity
and the rather common requirement for Mg2+ ions.  Yet, these apparently simple enzymes become
fascinating objects for structural and functional studies, and their molecular simplicity
becomes an advantage once we start asking detailed mechanistic questions.  For example, it is
quite obvious that these enzymes have been engineered by evolution to deal with the problem of
efficiently locating a specific site on DNA, which is immersed in a huge molar excess of
other, partly very similar sites.

<>

<1>Winkler, K.-P.
<2>Isolation and characterization of a sequence specific restriction endonuclease from Rhizobium lupini.
<3>Ph.D. Thesis, Erlangen University, W. Germany
<4>
<5>1-135
<6>1980
<7>None

<>

<1>Winter, M.
<2>Investigation of de novo methylation activity in mutants of the EcoKI methyltransferase.
<3>Ph.D. Thesis, University of Edinburgh, Edinburgh, Scotland, , , Edinburgh
<4>
<5>1-216
<6>1997
<7>A group of mutants of the EcoKI R/M-system displaying de novo methylation activity have been
isolated.  The mutated genes were transferred into an overexpressing plasmid vector.  Two of
the over-expressed proteins were purified to near homogeneity from clones transformed with the
plasmids.  Cofactor binding activities of wild-type and the two mutant enzymes were compared
by 1,8-anilino-napthalene sulphonic acid fluorescence displacement experiments.  A DNA
methylation assay based upon the transfer of a tritiated methyl group from the cofactor AdoMet
to the substrate DNA was established and used to examine the dependency of the reaction on
cofactor, substrate, and enzyme concentration.  In addition the stability of the trimeric
enzyme at different protein concentrations was followed by HPLC gel filtration.  Sequence
alignments, secondary structure predictions, and tertiary structural modelling were used to
show the similarity of the Type I system EcoKI with methyltransferases from other classes
(especially Type II methyltransferases), thereby establishing a structural and suggesting an
evolutionary link between the different methyltransferase classes.  The information obtained
by these comparisons enabled the subsequent modelling of a more refined model of the EcoKI
structure.  A model is proposed to explain the different activities observed in wild-type and
mutant enzymes based on the biochemical and structural data obtained during these
investigations.

<>

<1>Wion, D., Casadesus, J.
<2>N-6-methyl-adenine: an epigenetic signal for DNA-protein interactions.
<3>Nat. Rev. Microbiol.
<4>4
<5>183-192
<6>2006
<7>N6-methyl-adenine is found in the genomes of bacteria, archaea, protists and fungi.  Most
bacterial DNA adenine methyltransferases are part of restriction-modification systems.
Certain groups of Proteobacteria also harbour solitary DNA adenine methyltransferases that
provide signals for DNA-protein interactions.  In gamma-proteobacteria, Dam methylation
regulates chromosome replication, nucleoid segregation, DNA repair, transposition of insertion
elements and transcription of specific genes.  In Salmonella, Haemophilus, Yersinia and Vibrio
species and in pathogenic Escherichia coli, Dam methylation is required for virulence.  In
alpha-proteobacteria, CcrM methylation regulates the cell cycle in Caulobacter, Rhizobium and
Agrobacterium, and has a role in Brucella abortus infection.

<>

<1>Wirth, R. et al.
<2>Complete genome sequence of Desulfurococcus mucosus type strain (O7/1).
<3>Standards in Genomic Sciences
<4>4
<5>173-182
<6>2011
<7>Desulfurococcus mucosus Zillig and Stetter 1983 is the type species of the genus
Desulfurococcus, which belongs to the crenarchaeal family Desulfurococcaceae. The
species is of interest because of its position in the tree of life, its ability
for sulfur respiration, and several biotechnologically relevant thermostable and
thermoactive extracellular enzymes. This is the third completed genome sequence
of a member of the genus Desulfurococcus and already the 8(th) sequence from a
member the family Desulfurococcaceae. The 1,314,639 bp long genome with its 1,371
protein-coding and 50 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Wirth, R. et al.
<2>Complete genome sequence of Thermocrinis albus type strain (HI 11/12).
<3>Standards in Genomic Sciences
<4>2
<5>194-202
<6>2010
<7>Thermocrinis albus Eder and Huber 2002 is one of three species in the genus Thermocrinis in
the family Aquificaceae. Members of this family have become of
significant interest because of their involvement in global biogeochemical cycles
in high-temperature ecosystems. This interest had already spurred several genome
sequencing projects for members of the family. We here report the first completed
genome sequence a member of the genus Thermocrinis and the first type strain
genome from a member of the family Aquificaceae. The 1,500,577 bp long genome
with its 1,603 protein-coding and 47 RNA genes is part of the Genomic
Encyclopedia of Bacteria and Archaea project.

<>

<1>Wise, K.S., Calcutt, M.J., Foecking, M.F., Madupu, R., Deboy, R.T., Roske, K., Hvinden, M.L., Martin, T.R., Durkin, A.S., Glass, J.I., Methe, B.A.
<2>Complete Genome Sequences of Mycoplasma leachii Strain PG50T and the Pathogenic Mycoplasma mycoides subsp. mycoides Small Colony Biotype Strain Gladysdale.
<3>J. Bacteriol.
<4>194
<5>4448-4449
<6>2012
<7>Mycoplasma mycoides subsp. mycoides small colony biotype (SC) is the high-consequence animal
pathogen causing contagious bovine pleuropneumonia. We
report the complete genome sequences of the pathogenic strain M. mycoides subsp.
mycoides SC Gladysdale and a close phylogenetic relative, Mycoplasma leachii
PG50(T), another bovine pathogen of the M. mycoides phylogenetic clade.

<>

<1>Wisniewski-Dye, F. et al.
<2>Azospirillum genomes reveal transition of bacteria from aquatic to terrestrial environments.
<3>PLoS Genet.
<4>7
<5>e1002430
<6>2011
<7>Fossil records indicate that life appeared in marine environments ,3.5 billion years ago (Gyr)
and transitioned to terrestrial
ecosystems nearly 2.5 Gyr. Sequence analysis suggests that 'hydrobacteria' and
'terrabacteria' might have diverged as early as 3 Gyr. Bacteria of the genus Azospirillum
are associated with roots of terrestrial plants; however, virtually all their close relatives
are aquatic. We obtained genome sequences of two Azospirillum species and analyzed their gene
origins.
While most Azospirillum house-keeping genes have orthologs in its close aquatic relatives,
this lineage has obtained nearly half of its genome from terrestrial organisms. The majority
of genes encoding functions critical for association with plants are among horizontally
transferred genes. Our results show that transition of some aquatic bacteria to terrestrial
habitats
occurred much later than the suggested initial divergence of hydro- and terrabacterial clades.
The birth of the genus Azospirillum approximately coincided with the emergence of vascular
plants on land.

<>

<1>Withers, B.E., Ambroso, L.A., Dunbar, J.C.
<2>Structure and evolution of the XcyI restriction-modification system.
<3>Nucleic Acids Res.
<4>20
<5>6267-6273
<6>1992
<7>The XcyI restriction-modificaton system from Xanthomonas cyanopsidis recognizes the sequence,
CCCGGG. The XcyI endonuclease and methylase genes have been cloned and sequenced and were
found to be aligned in a head to tail orientation with the methylase preceding and overlapping
the endonuclease by one base pair. The nucleotide sequence codes for an N4 cytosine
methyltransferase with a predicted molecular weight of 33,500 and an endonuclease comprised of
333 codons and a molecular weight of 36,600. Sequence comparisons revealed significant
similarity between the XcyI, CfrI and SmaI methylisomers. in contrast, no similarity was
detected between the primary structures of the XcyI and SmaI endonucleases. The XcyI
restriction-modification system is highly homologous to the XmaI genes, although the DNA
sequences flanking the genes rapidly diverge. The sequence of the XcyI endonuclease contains
two motifs which have recently been identified as essential to the activity of the EcoRV
endonuclease.

<>

<1>Withers, B.E., Dunbar, J.C.
<2>The endonuclease isoschizomers, SmaI and XmaI bend DNA in opposite orientations.
<3>Nucleic Acids Res.
<4>21
<5>2571-2577
<6>1993
<7>The SmaI and XmaI endonucleases are imperfect isoschizomers that recognize the sequence
CCCGGG. SmaI cleaves between the internal CpG to produce blunt end scissions whereas XmaI
cleaves between the external cytosines to produce a four base, five prime overhang. Each of
the endonucleases forms stable, specific complexes with DNA in the absence of magnesium.
Circular permutation analyses of the protein-DNA complexes revealed that each of the
endonucleases induces bending of the DNA. Phase sensitive detection analyses verified the
existence of the SmaI and XmaI induced bends. Furthermore bending of the helix axis by the
endonucleases appeared to be directed in opposite orientations. The orientation of the
SmaI-induced bend appeared to be towards the major groove and is reminiscent of the direction
of the bend induced by EcoRV which similarly induces blunt end scissions. Conversely, XmaI
appeared to bend the DNA towards the minor groove.

<>

<1>Withers, B.E., Dunbar, J.C.
<2>Sequence-specific DNA recognition by the SmaI endonuclease.
<3>J. Biol. Chem.
<4>270
<5>6496-6504
<6>1995
<7>SmaI endonuclease recognizes and cleaves the sequence CCC/GGG. The enzyme requires magnesium
for catalysis; however, equilibrium binding assays revealed that the enzyme binds specifically
to DNA in the absence of magnesium. A specific association constant of 0.9 x 10/8 M-1 was
determined for SmaI binding to a 22-base duplex oligonucleotide. Furthermore, the KA was a
function of the length of the DNA substrate and the enzyme exhibited an affinity of 1.2 x 10/9
M-1 for a 195-base pair fragment and which represented a 10/4-fold increase in affinity over
binding to nonspecific sequences. A Km of 17.5 nM was estimated from kinetic assays based on
cleavage of the 22-base oligonucleotide and is not significantly different from the KD
estimated from the thermodynamic analyses. Footprinting (dimethyl sulfate and missing
nucleoside) analyses revealed that SmaI interacts with each of the base pairs within the
recognition sequence. Ethylation interference assays suggested that the protein contacts three
adjacent phosphages on each strand of the recognition sequence. Significantly, a predicted
protein contact with the phosphate 3' of the scissile bond may have implications in the
mechanism of catalysis by SmaI.

<>

<1>Withers, B.E., Dunbar, J.C.
<2>DNA determinants in sequence-specific recognition by XmaI endonuclease.
<3>Nucleic Acids Res.
<4>23
<5>3571-3577
<6>1995
<7>The XmaI endonuclease recognizes and cleaves the sequence C/CCGGG.  Magnesium is required for
catalysis, however, the enzyme forms stable, specific complexes with DNA in the absence of
magnesium.  An association constant of 1.2 x 10/9 M was estimated for the affinity of the
enzyme for a specific 195 bp fragment.  Competition assays revealed that the site-specific
association constant represented an approximately 10/4-fold increase in affinity over that for
non-cognate sites.  Missing nucleoside analyses suggested an interaction of the enzyme with
each of the cytosines and guanines within the recognition site.  Recognition of each of the
guanines was also indicated by dimethylsulfate interference footprinting assays.  The
phosphates 5' to the guanines within the recognition site appeared to be the major sites of
interaction of XmaI with the sugar-phosphate backbone.  No significant interaction of the
protein was observed with phosphates flanking the recognition sequence.  Comparison of the
footprinting patterns of XmaI with those of the neoschizomer SmaI (CCC/GGG) revealed that the
two enzymes utilize the same DNA determinants in their specific interaction with the CCCGGG
recognition site.

<>

<1>Withers, T.R., Johnson, S.L., Yu, H.D.
<2>Draft Genome Sequence for Pseudomonas aeruginosa Strain PAO579, a Mucoid Derivative of PAO381.
<3>J. Bacteriol.
<4>194
<5>6617
<6>2012
<7>Pseudomonas aeruginosa is an opportunistic pathogen that establishes a chronic lung infection
in individuals afflicted with cystic fibrosis. Here, we announce
the draft genome of P. aeruginosa strain PAO579, an alginate-overproducing
derivative of strain PAO381.

<>

<1>Witmer, H., Franks, M.
<2>Restriction and modification of Bacteriophage SP10 DNA by Bacillus subtilis Marburg 168: Stabilization of SP10 DNA in restricting hosts preinfected with a heterologous phage, SP18.
<3>J. Virol.
<4>37
<5>148-155
<6>1981
<7>SP10 phage cannot propagate in Bacillus subtilis Marburg 168 containing the
wild-type allele of either gene nonA or gene nonB.  The latter gene codes for
the intrinsic cellular restriction activity.  SP10 DNA was degraded in nonB+
derivatives of Marburg 168.  The degree of degradation depended upon the
previous host in which SP10 was propagated.  In the case of SP10 grown in B.
subtilis W23 (a nonrestricting, nonmodifying bacterium), 90% of the phage DNA
was hydrolyzed to acid solubles, and the residual acid-precipitable material
was recovered as 0.5- to 1-megadalton fragments.  In contrast, if SP10 was
propagated in B. subtilis PS9W7 (a nonA nonB derivative of Marbury 168 that
retains modifying activity), 40 to 50% of the input DNA was degraded to acid
solubles, and most of the remainder was recovered as 15- to 20-megadalton
fragments.  In nonA+ nonB cells, SP10 DNA was conerved as unit-length molecules
(ca. 80 megadalton).  Prior infection of nonB+ cells with SP18 protected
superinfecting SP10 DNA, even when rifampin or chloramphenicol was added before
the primary infection.  The data are discussed in terms of the following
conclusions.  (i) The nonB gene product of B. subtilis Marburg 168 is required
for restriction of SP10 DNA.  (ii) Some sites on SP10 DNA are sensitive to both
the restricting and modifying activities, whereas other sites are nonmodifiable
even though they are sensitive to the restriction enzyme.  (iii)  In some
manner, SP18 antagonizes the action of the nonB gene product.

<>

<1>Wittmayer, P.K., McKenzie, J.L., Raines, R.T.
<2>Degenerate DNA recognition by I-PpoI endonuclease.
<3>Gene
<4>206
<5>11-21
<6>1998
<7>The I-PpoI endonuclease is encoded by a group I intron found in the slime mold Physarum
polycephalum.  To initiate homing of its encoding intron, I-PpoI catalyzes a specific
double-stranded break within a 15-bp recognition site.  The high substrate specificities of
I-PpoI and other homing endonucleases make these enzymes valuable tools for genomic mapping
and sequencing.  Here, we report on the ability of I-PpoI to cleave recognition sites that
contain a wide variety of mutations generated randomly or deliberately.  We find that much
degeneracy is tolerated within the recognition site of I-PpoI.  Few single substitutions
prevent cleavage completely.  In addition, many sites with multiple substitutions are cleaved
efficiently. In contrast, deletions or insertions within the I-PpoI recognition site are
detrimental to catalysis, indicating that proper registry between the protein and its
substrate is critical.  Finally, we find that the sequence of the flanking regions can
influence catalysis by I-PpoI.  Thus, I-PpoI has both the complex binding specificity of a
transcription factor and the catalytic ability of a restriction endonuclease.

<>

<1>Witzel, K., Gwinn-Giglio, M., Nadendla, S., Shefchek, K., Ruppel, S.
<2>Genome Sequence of Enterobacter radicincitans DSM16656T, a Plant Growth-Promoting Endophyte.
<3>J. Bacteriol.
<4>194
<5>5469
<6>2012
<7>Enterobacter radicincitans sp. nov. DSM16656(T) represents a new species of the genus
Enterobacter which is a biological nitrogen-fixing endophytic bacterium
with growth-promoting effects on a variety of crop and model plant species. The
presence of genes for nitrogen fixation, phosphorous mobilization, and
phytohormone production reflects this microbe's potential plant growth-promoting
activity.

<>

<1>Wohlrab, F., Wells, R.D.
<2>Enzymatic probes for left-handed Z-DNA.
<3>Gene Amplif. Anal., Elsevier, Chirikjian, J.G., New York
<4>5
<5>247-256
<6>1987
<7>DNA is a polymorphic molecule and can adopt a variety of conformations depending mainly on
base sequence and environmental conditions.  Recent interest in this polymorphism has been
prompted to a large part by the discovery of Z-DNA, a structure possessing a left-handed helix
sense as opposed to the canonical right-handed state (B-DNA).  Several findings imply a
biological function for Z-DNA or B-to-Z transitions.

<>

<1>Wojciechowski, M., Czapinska, H., Bochtler, M.
<2>CpG underrepresentation and the bacterial CpG-specific DNA methyltransferase M.MpeI.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>110
<5>105-110
<6>2013
<7>Cytosine methylation promotes deamination. In eukaryotes, CpG methylation is thought to
account for CpG underrepresentation. Whether scarcity of CpGs in
prokaryotic genomes is diagnostic for methylation is not clear. Here, we report
that Mycoplasms tend to be CpG depleted and to harbor a family of constitutively
expressed or phase variable CpG-specific DNA methyltransferases. The very CpG
poor Mycoplasma penetrans and its constitutively active CpG-specific
methyltransferase M.MpeI were chosen for further characterization. Genome-wide
sequencing of bisulfite-converted DNA indicated that M.MpeI methylated CpG target
sites both in vivo and in vitro in a locus-nonselective manner. A crystal
structure of M.MpeI with DNA at 2.15-A resolution showed that the substrate base
was flipped and that its place in the DNA stack was taken by a glutamine residue.
A phenylalanine residue was intercalated into the 'weak' CpG step of the
nonsubstrate strand, indicating mechanistic similarities in the recognition of
the short CpG target sequence by prokaryotic and eukaryotic DNA
methyltransferases.

<>

<1>Wolcke, J.
<2>The kinetic mechanism of the DNA methyltransferase from Thermus aquaticus and selection of a DNA binding peptide using phage display.
<3>Ph.D. Thesis, University of Dortmund, Germany
<4>
<5>1-164
<6>1998
<7>
<>

<1>Wolcke, J., Weinhold, E.
<2>Substrate specificity of the DNA-methyltransferase from Thermus aquaticus: influence of the 3'-neighbor base.
<3>Biol. Chem. Hoppe Seyler
<4>376
<5>S169
<6>1995
<7>The DNA-methyltransferase from Thermus aquaticus (M.TaqI) catalyzes the methyl group transfer
from S-adenosyl-L-methionine (SAM) to the N6-position of adenine within the double-stranded
DNA sequence 5' TCGA 3'.
 We have investigated the effect of changing the 3' neighboring base of
adenine which gets methylated.  We find that steady-state kinetic rates for the methylation of
adenine by M.TaqI depend on the nature of the 3'-neighboring base.  In general, double
stranded deoxyoligo-nucleotides containing the pyrimidine bases thymine or cytosine next to
adenine give higher rates than those containing the purine bases adenine or guanine.  This
observed kinetic effect correlates inversely with the lower base stacking energies between
purine and pyrimidine bases compared to those between two purine bases.  A contribution of the
energy needed to break the base stacking between the adenine and its 3'-neighbor to the rate
limiting step is therefore suggested.  Correlation of the influence of the neighbor base pair
on the rate with the stacking energy between base pair neighbors, which might indicate a
distortion of both strands, however would predict a different kinetic effect.  In light of the
large distance (15A) between the cofactor and the DNA-binding site observed in the crystal
structure of M.TaqI complexed with SAM, the influence of the 3' neighbor on the steady state
rate gives the first experimental evidence, that a base flipping out mechanism described for
(cytosine-5)-DNA-Methyltransferases might also operate in adenine specific
DNA-Methyltransferases.

<>

<1>Wolcke, J., Weinhold, E.
<2>A DNA-binding peptide from a phage display library.
<3>Nucleosides Nucleotides Nucleic Acids
<4>20
<5>1239-1241
<6>2001
<7>A DNA-binding peptide was selected from a random peptide phage display library. For
competitive elution using the DNA methyltransferase M.TaqI in the selection step, a
biotin-labeled duplex oligodeoxyribonucleotide containing the 5'-TCGA-3' recognition
sequence of M.TaqI was employed. Nine of ten phages selected were found to have the same
deduced amino acid sequence SVSVGMKPSPRP. The selected phage binds to DNA, as demonstrated in
an ELISA.

<>

<1>Wolf, Y.I., Rogozin, I.B., Kondrashov, A.S., Koonin, E.V.
<2>Genome alignment, evolution of prokaryotic genome organization, and prediction of gene function using genomic context.
<3>Genome Res.
<4>11
<5>356-372
<6>2001
<7>Gene order in prokaryotes is conserved to a much lesser extent than protein sequences. Only
several operons, primarily those that code for
physically interacting proteins, are conserved in all or most of the
bacterial and archaeal genomes. Nevertheless, even the limited
conservation of operon organization that is observed can provide
valuable evolutionary and functional clues through multiple genome
comparisons. A program for constructing gapped local alignments of
conserved gene strings in two genomes was developed. The statistical
significance of the local alignments was assessed using Monte Carlo
simulations. Sets of local alignments were generated for all pairs of
completely sequenced bacterial and archaeal genomes, and for each
genome a template-anchored multiple alignment was constructed. In most
pairwise genome comparisons, <10% of the genes in each genome belonged
to conserved gene strings. When closely related pairs of species (i.e.,
two mycoplasmas) are excluded, the total coverage of genomes by
conserved gene strings ranged from <5% for the cyanobacterium
Synechocystis sp. to 24% for the minimal genome of Mycoplasma
genitalium, and 23% in Thermotoga maritima. The coverage of the
archaeal genomes was only slightly lower than that of bacterial
genomes. The majority of the conserved gene strings are known operons,
with the ribosomal superoperon being the top-scoring string in most
genome comparisons. However, in some of the bacterial-archaeal pairs,
the superoperon is rearranged to the extent that other operons,
primarily those subject to horizontal transfer, show the greatest level
of conservation, such as the archaeal-type H+-ATPase operon or ABC-type
transport cassettes. The level of gene order conservation among
prokaryotic genomes was compared to the cooccurrence of genomes in
clusters of orthologous genes (COGs) and to the conservation of protein
sequences themselves. Only limited correlation was observed between
these evolutionary variables. Gene order conservation shows a much
lower variance than the cooccurrence of genomes in COGs, which
indicates that intragenome homogenization via recombination occurs in
evolution much faster than intergenome homogenization via horizontal
gene transfer and lineage-specific gene loss. The potential of using
template-anchored multiple-genome alignments for predicting functions
of uncharacterized genes was quantitatively assessed. Functions were
predicted or significantly clarified for approximately 90 COGs (approximately 4% of the total
of 2414 analyzed COGs). The most significant
predictions were obtained for the poorly characterized archaeal
genomes; these include a previously uncharacterized
restriction-modification system, a nuclease-helicase combination
implicated in DNA repair, and the probable archaeal counterpart of the
eukaryotic exosome. Multiple genome alignments are a resource for
studies on operon rearrangement and disruption, which is central to our
understanding of the evolution of prokaryotic genomes. Because of the
rapid evolution of the gene order, the potential of genome alignment
for prediction of gene functions is limited, but nevertheless, such
predictions information significantly complements the results obtained
through protein sequence and structure analysis.

<>

<1>Wolfes, H., Alves, J., Fliess, A., Geiger, R., Pingoud, A.
<2>Site directed mutagenesis experiments suggest that Glu 111, Glu 144 and Arg 145 are essential for endonucleolytic activity of EcoRI.
<3>Nucleic Acids Res.
<4>14
<5>9063-9080
<6>1986
<7>We have constructed a plasmid (pRIF 309+) carrying the EcoRI restriction
endonuclease gene and the f1 origin of replication.  Upon transformation of
this plasmid into E. coli and infection with bacteriophage f1 single stranded
plasmids are produced which can be used for sequencing and site directed
mutagenesis.  Using this single stranded DNA and synthetic
oligodeoxynucleotides we have introduced point mutations at defined positions
of the EcoRI gene.  Since in pRIF309+ the EcoRI gene is under the control of
the pL- promoter, high level expression of the mutated EcoRI gene could be
obtained upon induction.  Mutant EcoRI enzymes were purified to homogeneity and
characterized in structural and functional terms.  Our results demonstrate that
the Glu 111 -> Gln, Glu 144 -> Gln and Arg 145 -> Lys -mutants adopt a very
similar conformation as the wild type enzyme, but have by two orders of
magnitude smaller specific activities than the wild type enzyme, mainly due to
a reduction of the Vmax-value.

<>

<1>Wolfes, H., Fliess, A., Pingoud, A.
<2>A comparison of the structural requirements for DNA cleavage by the isoschizomers HaeIII, BspRI and BsuRI.
<3>Eur. J. Biochem.
<4>150
<5>105-110
<6>1985
<7>We have investigated the structural requirements for DNA cleavage by the
isoschizomers HaeIII, BspRI and BsuRI which recognize the sequence -d(GGCC)-.
For this purpose decadeoxynucleotides were synthesized by the solid-phase
phosphotriester method and purified by high-performance liquid chromatography.
The kinetics of cleavage of these oligodeoxynucleotides were determined for the
three isoschizomers with the following results.  1)  The sequence adjacent to
the recognition site strongly influences the rate of cleavage.  The preference
is qualitatively the same for all three enzymes:  AGGCCT > TGGCCA > GGGCCC ~
CGGCCG, and follows the thermal stability of the different decanucleotides.  2)
Substitutions within the recognition site, namely dI for dG and dU for dC,
affect the rate of cleavage differently for the three enzymes.  The results can
be rationalized in terms of an interaction of HaeIII with the major and minor
groove of the DNA of BspRI mainly with the minor groove and of BsuRI with the
major groove of DNA.  It is obvious from our data that the mechanism of
recognition of the same site is different for the three isoschizomers.

<>

<1>Wolfes, H., Fliess, A., Winkler, F., Pingoud, A.
<2>Cross-linking of bromodeoxyuridine-substituted oligonucleotides to the EcoRI and EcoRV restriction endonucleases.
<3>Eur. J. Biochem.
<4>159
<5>267-273
<6>1986
<7>We have synthesized several self-complementary oligodeoxynucleotides which
contain bromodeoxyuridine in various positions within and outside of te
recognition sequence for the EcoRI and EcoRV restriction endonucleases.  These
oligodeoxynucleotides are cleaved in the presence of Mg2+ by their respective
enzyme.  Upon irradiation by long-wavelength ultraviolet light and in the
absence of Mg2+ they are cross-linked in low yield to their enzymes, forming
1:1 and 1:2 (oligodeoxynucleotide: enzyme subunit) adducts.  Cross-linking
occurs with both specific and non-specific complexes.  With EcoRI the site of
cross-linking was determined to be at or close to Met-137, i.e. in a region of
the molecule implicated by other studies from our laboratory [Scholtissek et
al. (1986) J. Biol. Chem. 261,1118-1134] in the binding and cleavage of the
substrate.

<>

<1>Wolffe, A.P., Jones, P.L., Wade, P.A.
<2>DNA demethylation.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>5894-5896
<6>1999
<7>Cytosine 5' methylation of CpG dinucleotides within and around genes exerts a major influence
on transcription in many plants and animals. DNA methylation can be causal for transcriptional
silencing and targets the machinery necessary to assemble specialized chromatin enriched in
deacetylated histones. Once established in somatic cells, CpG methylation patterns within the
genome are very stable and provide an attractive mechanism for segregating a large fraction of
stably repressed chromatin.  In contrast, DNA methylation is remarkably dynamic during early
mammalian development and in certain tumor cells.  Alterations in the methylation status of
the entire genome, individual chromosomes, and specific genes are essential for normal
development and can promote tumorigenesis.  Understanding how these important transitions
might be regulated requires the biochemical definition of the enzymatic processes that both
methylate and demethylate the genome.

<>

<1>Wolk, C.P., Kraus, J.
<2>Two approaches to obtaining low, extracellular deoxyribonuclease activity in cultures of heterocyst-forming cyanobacteria.
<3>Arch. Microbiol.
<4>131
<5>302-307
<6>1982
<7>Six out of 158 axenic strains of heterocyst-forming cyanobacteria consistently
failed to produce circles of clearing in agar medium containing DNA-methyl
green.  When tested with [3H]DNA and coliphage lambda DNA, supernatant fluids
from cultures of two of these strains [University of Texas Culture Collection
(UTEX) strain 2014 and 19-6C-C] showed no detectable deoxyribonuclease
activity, and such fluids from another two of the six, and four others, showed
low but detectable deoxyribonuclease activity.  Covalently closed circular
(plasmid) DNA was not detectably degraded by supernatant fluids from UTEX 2014
and 19-6C-C and from four of the other strains.  When lambda DNA was incubated
with whole cells of certain strains, a series of fragments of discrete size was
produced, perhaps by cell-bound, periplasmic, restriction endonucleases.
Inclusion of one-tenth strength saline sodium citrate (SSC) in an eight-fold
dilution of the medium of Allen and Arnon had little effect on growth of
Anabaena variabilis American Type Culture Collection (ATCC) strain 29413 yet
prevented all but slight degradation of plasmid pBR322 or of lambda DNA.

<>

<1>Wong, C.F., Niu, H., Jiang, J., Li, J., Chan, C.M., Leung, D.Y., Leung, F.C.
<2>Genome Sequence of Pseudomonas mendocina DLHK, Isolated from a Biotrickling Reactor.
<3>J. Bacteriol.
<4>194
<5>6326
<6>2012
<7>Pseudomonas mendocina DLHK is an aerobic bacterium isolated from a biotrickling reactor which
can remove nitric oxide, a common air pollutant from combustion
exhaust gas. Here, we present the draft genome of Pseudomonas mendocina DLHK.

<>

<1>Wong, D.L., Pavlovich, J.G., Reich, N.O.
<2>Electrospray ionization mass spectrometric characterization of photocrosslinked DNA-EcoRI DNA methyltransferase complexes.
<3>Nucleic Acids Res.
<4>26
<5>645-649
<6>1998
<7>We describe a novel strategy combining photocrosslinking and HPLC-based electrospray
ionization mass spectrometry to identify UV crosslinked DNA-protein complexes.  EcoRI DNA
methyltransferase modifies the second adenine within the recognition sequence GAATTC.
Substitution of 5-iodouracil for the thymine adjacent to the target base (GAATTC) does not
detectably alter the DNA-protein complex.  Irradiation of the 5-iodouracil-substituted
DNA-protein complex at various wavelengths was optimized, with a crosslinking yield >60% at
313 nm after 1 min.  No protein degradation was observed under these conditions.  The
crosslinked DNA-protein complex was further anlayzed by electrospray ionization mass
spectrometry.  The total mass is consistent with irradiation-dependent covalent bond formation
between one strand of DNA and the protein.  These preliminary results support the possibility
of identifying picomole quantities of crosslinked peptides by similar strategies.

<>

<1>Wong, D.L., Reich, N.O.
<2>Identification of tyrosine 204 as the photo-cross-linking site in the DNA - EcoRI DNA methyltransferase complex by electrospray ionization mass spectrometry.
<3>Biochemistry
<4>39
<5>15410-15417
<6>2000
<7>We describe a highly sensitive strategy combining laser-induced photo-cross-linking and
HPLC-based electrospray ionization mass
spectrometry to identify amino acid residues involved in protein-DNA
recognition. The photoactivatible cross-linking thymine isostere,
5-iodouracil, was incorporated at a single site within the sequence
recognized by EcoRI DNA methyltransferase (GAATTC). UV irradiation of
the DNA-protein complex at 313 nm results in a >60% cross-linking
yield. SDS-polyacrylamide gel electrophoresis and mass spectrometry
were used to analyze the covalent cross-linked complex. The total mass
is consistent with covalent bond formation between one strand of DNA
and the protein with 1:1 stoichiometry. Protease digestion of the
cross-linked complex yields several peptide-DNA adducts that were
purified by anion-exchange column chromatography. A combination of mass
spectrometric analysis and amino acid sequencing revealed that tyrosine
204 was cross-linked to the DNA. Electrospray mass spectrometric
analysis of the peptide-nucleoside adduct confirmed this assignment.
Tyrosine 204 resides in a peptide motif previously thought to be
involved in AdoMet binding and methyl transfer. Thus, amino acids
within loop segments but outside of "DNA binding" motifs can be
critical to DNA recognition. Our method provides an accurate
characterization of picomole quantities of DNA-protein complexes.

<>

<1>Wong, D.M., Wetmur, J.G.
<2>Some class-IIS restriction endonucleases can cleave across a three-way DNA junction.
<3>Gene
<4>150
<5>63-66
<6>1994
<7>Three-way DNA junctions were synthesized with class-IIS restriction endonuclease (ENase)
recognition sites and potential cleavage sites located on separate arms. Cleavage was
investigated with junctions labeled in each of the three strands. BpmI and BsaI failed to
cleave either strand of either arm, whereas BsmAI cleaved one strand. FokI and HphI cleaved
both strands of both arms at the expected nucleotide positions. FokI cleavage was independent
of the spacing between the recognition site and the junction. This new activity of class-IIS
may be useful for investigating branched DNA structures.

<>

<1>Wong, K.K., McClelland, M.
<2>PCR with 5-methyl-dCTP replacing dCTP.
<3>Nucleic Acids Res.
<4>19
<5>1081-1085
<6>1991
<7>When dCTP is replaced by methyl-5-dCTP in the polymerase chain reaction some templates cannot
be efficiently amplified by Taq polymerase or Vent tm polymerase using standard cycling
parameters. However, this phenomenon can be overcome by increasing the temperature of the
denaturation steps to 100 C, or by adding dITP to destabilize the m5dC:dG base pairs. Once the
block to amplification of m5dC-substituted DNA was overcome, methylated DNA from the
superpolylinker of the plasmid pSL1180 was used as a substrate to check the methyl-sensitivity
of a variety of restriction endonucleases. The m5dC-substituted DNAs should also be valuable
substrates for defining the specificity of methyl-dependent endonucleases.

<>

<1>Wong, R.M.-Y., Wong, K.K., Benson, N.R., McClelland, M.
<2>Sample sequencing of a Salmonella typhimurium LT2 lambda library: comparison to the Escherichia coli K12 genome.
<3>FEMS Microbiol. Lett.
<4>173
<5>411-423
<6>1999
<7>As part of the ongoing sequencing of the complete Salmonella typhimurium
LT2 genome, a partly ordered set of 416 lambda clones has been developed,
representing over 90% of the genome. The average insert size is 17 kb.
Sequences were obtained from both ends of each clone in this set. A total
of over 600 kb of sequence has been deposited in the genome survey
sequence section of GenBank. This resource of clones is available from the
Salmonella Genome Stock Center. A preliminary comparison with the
Escherichia coli K12 genome indicates that there are likely to be many
hundred insertion deletion events, encompassing more than one gene, that
distinguish these genomes. Fully 30% of the S. typhimurium sequences have
no close homologs in the GenBank database.

<>

<1>Wong, Y.L., Choo, S.W., Tan, J.L., Ong, C.S., Ng, K.P., Ngeow, Y.F.
<2>Draft Genome Sequence of Mycobacterium bolletii Strain M24, a Rapidly Growing Mycobacterium of Contentious Taxonomic Status.
<3>J. Bacteriol.
<4>194
<5>4475
<6>2012
<7>The whole-genome sequence of Mycobacterium bolletii M24, isolated from the bronchoalveolar
lavage fluid of a Malaysian patient, is reported here. The
circular chromosome of 5,507,730 bp helped to clarify the taxonomic position of
this organism within the M. abscessus complex and revealed the presence of
proteins potentially important for pathogenicity in a human host.

<>

<1>Wong, Y.M., Juan, J.C., Gan, H.M., Austin, C.M.
<2>Draft Genome Sequence of Clostridium bifermentans Strain WYM, a Promising Biohydrogen Producer Isolated from Landfill Leachate Sludge.
<3>Genome Announcements
<4>2
<5>e00077-14
<6>2014
<7>Clostridium bifermentans strain WYM is an effective biohydrogen producer isolated from
landfill leachate sludge. Here, we present the assembly and annotation of
its genome, which may provide further insights into the metabolic pathways
involved in efficient biohydrogen production.

<>

<1>Wong, Y.M., Juan, J.C., Gan, H.M., Austin, C.M.
<2>Draft Genome Sequence of Clostridium perfringens Strain JJC, a Highly Efficient Hydrogen Producer Isolated from Landfill Leachate Sludge.
<3>Genome Announcements
<4>2
<5>e00064-14
<6>2014
<7>Clostridium perfringens strain JJC is an effective biohydrogen and biochemical producer that
was isolated from landfill leachate sludge. Here, we present the
assembly and annotation of its genome, which may provide further insights into
the gene interactions involved in efficient biohydrogen production.

<>

<1>Wong, Y.M., Juan, J.C., Ting, A., Wu, T.Y., Gan, H.M., Austin, C.M.
<2>Draft Genome Sequence of Clostridium sp. Strain Ade.TY, a New Biohydrogen- and Biochemical-Producing Bacterium Isolated from Landfill Leachate Sludge.
<3>Genome Announcements
<4>2
<5>e00078-14
<6>2014
<7>Clostridium sp. strain Ade.TY is potentially a new biohydrogen-producing species  isolated
from landfill leachate sludge. Here we present the assembly and
annotation of its genome, which may provide further insights into its gene
interactions for efficient biohydrogen production.

<>

<1>Woo, H.L., Utturkar, S., Klingeman, D., Simmons, B.A., DeAngelis, K.M., Brown, S.D., Hazen, T.C.
<2>Draft Genome Sequence of the Lignin-Degrading Burkholderia sp. Strain LIG30, Isolated from Wet Tropical Forest Soil.
<3>Genome Announcements
<4>2
<5>e00637-14
<6>2014
<7>Burkholderia species are common soil Betaproteobacteria capable of degrading recalcitrant
aromatic compounds and xenobiotics. Burkholderia sp. strain LIG30
was isolated from wet tropical forest soil and is capable of utilizing lignin as
a sole carbon source. Here we report the draft genome sequence of Burkholderia
sp. strain LIG30.

<>

<1>Woo, P.C., Lau, S.K., Tse, H., Teng, J.L., Curreem, S.O., Tsang, A.K., Fan, R.Y., Wong, G.K., Huang, Y., Loman, N.J., Snyder, L.A., Cai, J.J., Huang, J.D., Mak, W., Pallen, M.J., Lok, S., Yuen, K.Y.
<2>The complete genome and proteome of Laribacter hongkongensis reveal potential mechanisms for adaptations to different temperatures and habitats.
<3>PLoS Genet.
<4>5
<5>E1000416
<6>2009
<7>Laribacter hongkongensis is a newly discovered Gram-negative bacillus of
the Neisseriaceae family associated with freshwater fish-borne
gastroenteritis and traveler's diarrhea. The complete genome sequence of
L. hongkongensis HLHK9, recovered from an immunocompetent patient with
severe gastroenteritis, consists of a 3,169-kb chromosome with G+C content
of 62.35%. Genome analysis reveals different mechanisms potentially
important for its adaptation to diverse habitats of human and freshwater
fish intestines and freshwater environments. The gene contents support its
phenotypic properties and suggest that amino acids and fatty acids can be
used as carbon sources. The extensive variety of transporters, including
multidrug efflux and heavy metal transporters as well as genes involved in
chemotaxis, may enable L. hongkongensis to survive in different
environmental niches. Genes encoding urease, bile salts efflux pump,
adhesin, catalase, superoxide dismutase, and other putative virulence
factors-such as hemolysins, RTX toxins, patatin-like proteins,
phospholipase A1, and collagenases-are present. Proteomes of L.
hongkongensis HLHK9 cultured at 37 degrees C (human body temperature) and
20 degrees C (freshwater habitat temperature) showed differential gene
expression, including two homologous copies of argB, argB-20, and argB-37,
which encode two isoenzymes of N-acetyl-L-glutamate kinase (NAGK)-NAGK-20
and NAGK-37-in the arginine biosynthesis pathway. NAGK-20 showed higher
expression at 20 degrees C, whereas NAGK-37 showed higher expression at 37
degrees C. NAGK-20 also had a lower optimal temperature for enzymatic
activities and was inhibited by arginine probably as negative-feedback
control. Similar duplicated copies of argB are also observed in bacteria
from hot springs such as Thermus thermophilus, Deinococcus geothermalis,
Deinococcus radiodurans, and Roseiflexus castenholzii, suggesting that
similar mechanisms for temperature adaptation may be employed by other
bacteria. Genome and proteome analysis of L. hongkongensis revealed novel
mechanisms for adaptations to survival at different temperatures and
habitats.

<>

<1>Woo, Y.H., Rajagopalan, P.T.R., Benkovic, S.J.
<2>A nonradioactive DNA methyltransferase assay adaptable to high-throughput screening.
<3>Anal. Biochem.
<4>340
<5>336-340
<6>2005
<7>We have developed a nonradioactive assay method for DNA methyltransferases based on the
ability to protect substrate DNA from
restriction. DNA immobilized to a microplate well was treated
sequentially with methyltransferase and an appropriate endonuclease.
The amount of methylated DNA product is reflected by a proportional
decrease in endonuclease cleavage, which is in turn reflected by
increased retention of the end-labeled affinity probe. A single
universal substrate was designed to assay multiple methyltransferases
including those that do not have a cognate endonuclease. The
methodology developed is suited to screen a large number of compounds
for inhibitors of various methyltransferases.

<>

<1>Wood, D.W. et al.
<2>The genome of the natural genetic engineer Agrobacterium tumefaciens C58.
<3>Science
<4>294
<5>2317-2323
<6>2001
<7>The 5.67-megabase genome of the plant pathogen Agrobacterium tumefaciens C58 consists of a
circular chromosome, a linear chromosome, and two plasmids.  Extensive orthology and
nucleotide colinearity between the genomes of A. tumefaciens and the plant symbiont
Sinorhizobium meliloti suggest a recent evolutionary divergence.  Their similarities include
metabolic, transport, and regulatory systems that promote survival in the highly competitive
rhizosphere; differences are apparent in their genome structure and virulence gene complement.
Availability of the A. tumefaciens sequence will facilitate investigations into the molecular
basis of pathogenesis and the evolutionary divergence of pathogenic and symbiotic lifestyles.

<>

<1>Wood, K.M., Daniels, L.E., Halford, S.E.
<2>Long-range communications between DNA sites by the dimeric restriction endonuclease SgrAl.
<3>J. Mol. Biol.
<4>350
<5>240-253
<6>2005
<7>The SgrAI endonuclease displays its maximal activity on DNA with two copies of its recognition
sequence, cleaving both sites concertedly.
While most restriction enzymes that act concurrently at two sites are
tetramers, SgrAI is a dimer in solution. Its reaction at two cognate
sites involves the association of two DNA-bound dimers. SgrAI can also
bridge cognate and secondary sites, the latter being certain sequences
that differ from the cognate by one base-pair. The mechanisms for
cognate-cognate and cognate-secondary communications were examined for
sites in the following topological relationships: in cis, on plasmids
with two sites in a single DNA molecule; on catenanes containing two
interlinked rings of DNA with one site in each ring; and in trans, on
oligoduplexes carrying either a single site or the DNA termini
generated by SgrAI. Both cognate-cognate and cognate-secondary
interactions occur through 3-D space and not by 1-D tracking along the
DNA. Both sorts of communication arise more readily when the sites are
tethered to each other, either in cis on the same molecule of DNA or by
the interlinking of catenane rings, than when released from the tether.
However, the dimer bound to an oligoduplex carrying either a cognate or
a secondary site could be activated to cleave that duplex by
interacting with a second dimer bound to the recognition site, provided
both duplexes are at least 30 base-pairs long: the second dimer could
alternatively be bound to the two duplexes that correspond to the
products of DNA cleavage by SgrAI.

<>

<1>Wood, R.J., Maynard-Smith, M.D., Robinson, V.L., Oyston, P.C.F., Titball, R.W., Roach, P.L.
<2>Kinetic analysis of Yersinia pestis DNA adenine methyltransferase activity using a hemimethylated molecular break light oligonucleotide.
<3>PLoS ONE
<4>2
<5>0
<6>2007
<7>Background. DNA adenine methylation plays an important role in several critical bacterial
processes including mismatch
repair, the timing of DNA replication and the transcriptional control of gene expression. The
dependence of bacterial virulence
on DNA adenine methyltransferase (Dam) has led to the proposal that selective Dam inhibitors
might function as broad
spectrum antibiotics. Methodology/Principal Findings. Herein we report the expression and
purification of Yersinia pestis
Dam and the development of a continuous fluorescence based assay for DNA adenine
methyltransferase activity that is
suitable for determining the kinetic parameters of the enzyme and for high throughput
screening against potential Dam
inhibitors. The assay utilised a hemimethylated break light oligonucleotide substrate
containing a GATC methylation site.
When this substrate was fully methylated by Dam, it became a substrate for the restriction
enzyme DpnI, resulting in
separation of fluorophore (fluorescein) and quencher (dabcyl) and therefore an increase in
fluorescence. The assays were
monitored in real time using a fluorescence microplate reader in 96 well format and were used
for the kinetic characterisation
of Yersinia pestis Dam, its substrates and the known Dam inhibitor, S-adenosylhomocysteine.
The assay has been validated for
high throughput screening, giving a Z-factor of 0.7160.07 indicating that it is a sensitive
assay for the identification of
inhibitors. Conclusions/Significance. The assay is therefore suitable for high throughput
screening for inhibitors of DNA
adenine methyltransferases and the kinetic characterisation of the inhibition.

<>

<1>Wood, R.J., McKelvie, J.C., Maynard-Smith, M.D., Roach, P.L.
<2>A real-time assay for CpG-specific cytosine-C5 methyltransferase activity.
<3>Nucleic Acids Res.
<4>38
<5>e107
<6>2010
<7>A real-time assay for CpG-specific cytosine-C5 methyltransferase activity has been developed.
The assay applies a break light oligonucleotide in
which the methylation of an unmethylated 5'-CG-3' site is enzymatically
coupled to the development of a fluorescent signal. This sensitive assay
can measure rates of DNA methylation down to 0.34 +/- 0.06 fmol/s. The
assay is reproducible, with a coefficient of variation over six
independent measurements of 4.5%. Product concentration was accurately
measured from fluorescence signals using a linear calibration curve, which
achieved a goodness of fit (R(2)) above 0.98. The oligonucleotide
substrate contains three C5-methylated cytosine residues and one
unmethylated 5'-CG-3' site. Methylation yields an oligonucleotide
containing the optimal substrate for the restriction enzyme GlaI. Cleavage
of the fully methylated oligonucleotide leads to separation of fluorophore
from quencher, giving a proportional increase in fluorescence. This method
has been used to assay activity of DNMT1, the principle maintenance
methyltransferase in human cells, and for the kinetic characterization of
the bacterial cytosine-C5 methyltransferase M.SssI. The assay has been
shown to be suitable for the real-time monitoring of DNMT1 activity in a
high-throughput format, with low background signal and the ability to
obtain linear rates of methylation over long periods, making this a
promising method of high-throughput screening for inhibitors.

<>

<1>Wood, V. et al.
<2>The genome sequence of Schizosaccharomyces pombe.
<3>Nature
<4>415
<5>871-880
<6>2002
<7>We have sequenced and annotated the genome of fission yeast
(Schizosaccharomyces pombe), which contains the smallest number of
protein-coding genes yet recorded for a eukaryote: 4,824. The centromeres
are between 35 and 110 kilobases (kb) and contain related repeats
including a highly conserved 1.8-kb element. Regions upstream of genes are
longer than in budding yeast (Saccharomyces cerevisiae), possibly
reflecting more-extended control regions. Some 43% of the genes contain
introns, of which there are 4,730. Fifty genes have significant similarity
with human disease genes; half of these are cancer related. We identify
highly conserved genes important for eukaryotic cell organization
including those required for the cytoskeleton, compartmentation,
cell-cycle control, proteolysis, protein phosphorylation and RNA splicing.
These genes may have originated with the appearance of eukaryotic life.
Few similarly conserved genes that are important for multicellular
organization were identified, suggesting that the transition from
prokaryotes to eukaryotes required more new genes than did the transition
from unicellular to multicellular organization.

<>

<1>Wood, W.B.
<2>Host specificity of DNA produced by Escherichia coli:  bacterial mutations affecting the restriction and modification of DNA.
<3>J. Mol. Biol.
<4>16
<5>118-133
<6>1966
<7>Bacterial mutants lacking the ability to restrict lambda phage of foreign host
specificity have been isolated from Escherichia coli strains K12 and B using a
convenient selection technique.  About half of the mutants still impart host
specificity to lambda; the remainder are deficient in modification as well as
restriction.  In both K12 and B, mutations affecting restriction map close to
the thr locus on the side opposite to leu.  Behavior of the mutant strains in
conjugation experiments is consistent with the view that bacterial DNA is
restricted and modified by the same factors which act upon the DNA of phage
lambda, and that this restriction isi responsible for the abnormally low
frequency of recombinant formation observed in K12 X B crosses.  The properties
of the modification-restriction system in E. coli suggest that it serves as a
defense mechanism against incoming nucleic acid, preventing the expression or
integration of foreign DNA without hindering genetic exchange among cells of
the same strain.

<>

<1>Wood, W.B.
<2>Mutations in E. coli affecting the host-controlled modification bacteriophage lambda.
<3>Pathol. Microbiol. (Basel)
<4>28
<5>73-76
<6>1965
<7>This report describes the isolation and some properties of K12 and B strains
which have lost the restriction (r) and modification (m) functions as a result
of either spontaneous or ethyl-methane sulfonate (EMS)-induced mutations.
Glover et al. have recently described similar mutations carried by P1 prophage
which affect the host-controlled modification of lambda.K in lysogenic E. coli
K12 (P1) strains.

<>

<1>Woodbury, C.P. Jr., Downey, R.L., von Hippel, P.H.
<2>EcoRI Methylase: On its substrate specificity and use in radioactive labeling of DNA.
<3>Fed. Proc.
<4>37
<5>1415
<6>1978
<7>While investigating the mechanisms of the EcoRI endonuclease and methylase, we
have examined the effect of alterations in reaction conditions on the
methylation process.  We find that in 100 mM Tris, 2 mM EDTA, 50% glycerol (pH
9), the methylase transfers many more methyl groups to both ColE1 plasmid and
phage lambda DNA than can be accommodated in the known "canonical" substrate
sites of these DNAs.  We have confirmed that "non-canonical" sites are
methylated under these conditions by demonstrating the incorporation of
substantial amounts of label into phage PhiX174 DNA, which contains no
canonical sequences, as well as into in vivo-methylated E. coli strain RY13
(rRI+, mRI+) chromosomal DNA.  We have also incorporated label into both duplex
poly[d(A-T)] and poly(dA).poly(dT).  The rate of methylation of these
substrates decreases in the order: canonical sites > non-canonical sites
>duplex poly[d(A-T)] > poly(dA).poly(dT).  The implications of these findings
for mechanisms in the EcoRI system will be discussed.  In addition to its
mechanistic interest, this nonspecific methylation procedure provides a
convenient way to radioactively label DNA in vitro; we have achieved ~105
cpm/microgram with various DNAs, a level comparable to that obtainable by in
vivo 32P-labeling.  Examples of the use of this technique will be described.

<>

<1>Woodbury, C.P. Jr., Downey, R.L., von Hippel, P.H.
<2>DNA site recognition and overmethylation by the EcoRI methylase.
<3>J. Biol. Chem.
<4>255
<5>11526-11533
<6>1980
<7>The EcoRI modification methylase exhibits reduction of in vitro substrate
specificity under conditions of alkaline pH, low ionic strength, and igh
glycerol content.  Under these conditions the enzyme transfers many more methyl
groups to both plasmid ColE1 DNA and phage lambda DNA than can be accommodated
in the known "canonical" recognition sites of these DNAs.  Substantial
methylation of phage PhiX174 replicative form DNA, which contains no canonical
sites, has also been demonstrated.  This "noncanonical" methylation occurs at
sites recognized by the EcoRI restriction endonuclease under parallel
conditions of low ionic strength and alkaline pH, as evidenced by protection of
the overmethylated ColE1 or PhiX174 DNAs against attack by the nuclease
(Woodbury, C.P., Jr., Hagenbuchle, O., and von Hippel, P.H. (1980) J. Biol.
Chem. 255, 11534-11546).  The methylase also modifies, at a much lower rate,
the double-stranded form of the alternating copolymer, poly[d(A-T)], but does
not transfer appreciable numbers of methyl groups to the homoduplex,
poly(dA).poly(dT).  The substrates methylated, in order of decreasing specific
rate, are: canonical DNA sites > noncanonical DNA sites > duplex poly[d(A-T)] >
poly(dA).poly(dT).  In addition, this enzyme (under noncanonical methylating
conditions) modifies in vivo methylated Escherichia coli RY13 (r+ RI, M+RI)
chromosomal DNA, but at a much lower rate than the other DNAs tested.  This
suggests that the noncanonical (as well as the canonical) sites of this DNA
have been largely modified in vivo.  The high level of methyl group
incorporation attainable using the noncanonical activity of the EcoRI methylase
has also been employed as a tool to radioactively label DNA in vitro.
Double-stranded lambda plac DNA has been labeled easily to 130,000
cpm/microgram by this technique; this modified DNA has been shown to be fully
functional as operator in lac operator-repressor filter-binding assays.

<>

<1>Woodbury, C.P. Jr., Hagenbuchle, O., von Hippel, P.H.
<2>DNA site recognition and reduced specificity of the EcoRI endonuclease.
<3>J. Biol. Chem.
<4>255
<5>11534-11546
<6>1980
<7>It has been shown previously (Polisky, B., Green, P., Garfin, D.E., McCarthy,
B.J., Goodman, H.M. and Boyer, H.W. (1975) Proc. Natl. Acad. Sci. USA 72:
3310-3314; Hsu, M., and Berg, P. (1978) Biochemistry 17: 131-138) that the
cleavage sequence specificity of EcoRI endonuclease can be "relaxed" by various
means.  In this paper this phenomenon is explored in detail, in order to obtain
further insight into the nature and selectivity of sequence recognition
patterns between proteins and double-stranded nucleic acids.  Using conditions
of low ionic strength and alkaline pH, we have mapped the positions of
potentially cleavable sites in the (completely sequenced) replicative form of
the bacteriophage PhiX174 genome, and have deduced their sequence.  The time
course of digestion of PhiX174 DNA suggests that double-stranded sequences
reading GGATTT, AAATTT, GAATTT, and GAATTA (only "top" strands, written 5' -
3', are shown) are cleaved readily under these conditions, while sequences
reading CAATTN (N = A,T,G) resist attack.  Cleavages at (at least) the more
labile sites result in cohesive ends that are religatable.  End group analysis
of cleaved PhiX174 DNA fragments indicates the presence of a 5' - terminal
adenine residue on most of the fragments; some fragments may carry a
5'-terminal guanine residue, consistent with the cleavage site sequences
suggested above.  Addition of Mn2+ to cleavage reactions carried out at
moderate salt concentrations and near-neutral pH indices the same pattern of
cleavage seen at low ionic strength and alkaline pH.  These results are
combined with those from other studies, and are interpreted in terms of a model
for the site-specific interation of the EcoRI endonuclease with its substrate,
considering both the effects of changes in DNA sequence and of environmental
alterations.  The resulting model is compared with data developed on similar
grounds for EcoRI methylase (see Woodbury, C.P., Downey, R.L., and von Hippel,
Ph.H. (1980) J. Biol. Chem. 255: 11526-11533), and attempts are made to define
both common and differing molecular facets of the DNA recognition specificity
of these companion (but genetically distinct enzymes.

<>

<1>Woodbury, C.P. Jr., von Hippel, P.H.
<2>Relaxed sequence specificities of EcoRI endonuclease and methylase: Mechanisms, possible practical applications, and uses in defining protein-nucleic acid recognition mechanisms.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>181-207
<6>1981
<7>*
   I. Introduction
  II. Conditions for relaxation of enzymatic specificity
 III. Sequence specificity of noncanonical EcoRI activities
  IV. Is relaxed specificity due to an impurity
   V. The role of binding affinity in relaxed specificity
  VI. Recognition contacts for the endonuclease
 VII. Recognition contacts for the methylase
VIII. General features of the endonuclease recognition sequence
  IX. Free energy considerations in non-sequence-specific and sequence-specific binding
   X. Applications of the relaxed-specificity reactions of endonuclease and methylase
  XI. Appendix


<>

<1>Woodcock, D.M., Crowther, P.J., Diver, W.P., Graham, M., Bateman, C., Baker, D.J., Smith, S.S.
<2>RglB facilitated cloning of highly methylated eukaryotic DNA:  the human L1 transposon, plant DNA, and DNA methylated in vitro with human DNA methyltransferase.
<3>Nucleic Acids Res.
<4>16
<5>4465-4482
<6>1988
<7>In vitro methylation of Bluescribe plasmid DNA (pBS) with human placental DNA
methyltransferase to 6% 5-methylcytosine (mC) reduced transformation
efficiencies in rglB+ host strains C600 and DS410 by almost 2 orders of
magnitude.  By contrast, the rglB- derivative of DS410 showed no reduction in
transformation efficiency with methylation while the rglB- derivative of C600
was partially tolerant to methylation.  Further, we show that the 1.8 kilobase
(kb) and 1.2 kb KpnI fragments derived from the human L1 repeat have
respectively 18.3% and 2.3% mC in vivo.  Using these hyper- and hypo-methylated
genomic segments ligated into the pBS plasmid, transformants with the highly
methylated 1.8 kb L1 insert were recovered at 17 to 40 fold higher frequency
with the rglB- host strains than with the rglB+ hosts.  In addition,
recombinant phage (lambda 2001) containing inserts of plant genomic DNA with
26.7% mC (from Petunia hybrida) when plated on rglB- hosts gave titres up to
222 times higher than on rglB+ strains.

<>

<1>Woodcock, D.M., Lisenmeyer, M.E., Warren, W.D.
<2>DNA methylation in mouse A-repeats in DNA methyltransferase-knockout ES cells and in normal cells determined by bisulfite genomic sequencing.
<3>Gene
<4>206
<5>63-67
<6>1998
<7>Mouse ES cells with a null mutation of the known DNA methyltransferase retain some residual
DNA methylation and can methylate foreign sequences de novo.  We have used bisulfite genomic
sequencing to examine the sequence specificity and distributions of methylation of a
hypermethylated CG island sequence, mouse A-repeats.  There were 13 CG dinucleotides in the
region examined, 12 of which were methylated to variable extents in all DNAs.  We found that:
(1) there is considerable residual DNA methylation in ES cells lacking the known DNA
methyltransferase (29% of normal methylation in the complete knockout ES DNA); (2) this other
activity methylates at exactly the same CG sites as the major methyltransferase; and (3)
differences in the distribution of methylated sites between A-repeats in these DNAs are
consistent with this other activity methylating in a random de novo fashion.  Also, the lack
of any methylation in non-CG sites argues that, in other studies where non-CG methylation
sites have been found by bisulfite sequencing, detection of such sites of non-CG methylation
is not an inherent artifact in this methodology.

<>

<1>Woodford, N., Carattoli, A., Karisik, E., Underwood, A., Ellington, M.J., Livermore, D.M.
<2>Complete nucleotide sequences of plasmids pEK204, pEK499, and pEK516, encoding CTX-M enzymes in three major Escherichia coli lineages from the United Kingdom,  all belonging to the international O25:H4-ST131 clone.
<3>Antimicrob. Agents Chemother.
<4>53
<5>4472-4482
<6>2009
<7>We determined the complete nucleotide sequences of three plasmids that encode CTX-M
extended-spectrum beta-lactamases (ESBLs) in pulsed-field gel
electrophoresis-defined United Kingdom variants (strains A, C, and D) of the
internationally prevalent Escherichia coli O25:H4-ST131 clone. Plasmid pEK499
(strain A; 117,536 bp) was a fusion of type FII and FIA replicons and harbored
the following 10 antibiotic resistance genes conferring resistance to eight
antibiotic classes: bla(CTX-M-15), bla(OXA-1), bla(TEM-1,) aac6'-Ib-cr, mph(A),
catB4, tet(A), and the integron-borne dfrA7, aadA5, and sulI genes. pEK516
(strain D; 64,471 bp) belonged to incompatibility group IncFII and carried seven
antibiotic resistance genes: bla(CTX-M-15), bla(OXA-1), bla(TEM-1), aac6'-Ib-cr,
catB4, and tet(A), all as in pEK499. It also carried aac3-IIa, conferring
gentamicin resistance, and was highly related to pC15-1a, a plasmid encoding the
CTX-M-15 enzyme in Canada. By contrast, pEK204 (strain C; 93,732 bp) belonged to
incompatibility group IncI1 and carried only two resistance genes, bla(CTX-M-3)
and bla(TEM-1). It probably arose by the transposition of Tn3 and
ISEcp1-bla(CTX-M-3) elements into a pCOLIb-P9-like plasmid. We conclude that (i)
United Kingdom variants of the successful E. coli ST131 clone have acquired
different plasmids encoding CTX-M ESBLs on separate occasions, (ii) the
bla(CTX-M-3) and bla(CTX-M-15) genes on pEK204 and pEK499/pEK516 represent
separate escape events, and (iii) IncFII plasmids harboring bla(CTX-M-15) have
played a crucial role in the global spread of CTX-M-15 ESBLs in E. coli.

<>

<1>Woodhead, J.L., Bhave, N., Malcolm, D.B.
<2>Cation dependence of restriction endonuclease EcoRI activity.
<3>Eur. J. Biochem.
<4>115
<5>293-296
<6>1981
<7>Restriction endonuclease EcoRI cleaves the DNA sequence 5'd(-G-^A-A-T-T-C-)
under optimum digestion conditions.  A variation in pH and ionic strength can
result in EcoRI* activity when 5'd(-^A-A-T-T-) is cut. A divalent cation,
usually Mg2+, is required for enzyme activity, though Mn2+ can also be used.
Eight different cations with ionic radius/charge ratios similar to Mg2+ were
tested and Co2+ and Zn2+ were also found to act as cofactors for EcoRI.  A
comprehensive study has been made of the effect of NaCl and pH on the
EcoRI/EcoRI* transition in the presence of the above four cations.  Generally,
a decrease in NaCl and/or an increase in pH caused a decrease in enzyme
specificity.  The changeover depended on the cation.  They may be placed in
order of their ability to increase EcoRI specificity thus: Co2+ > Zn2+ > Mg2+ >
Mn2+.  The Km of EcoRI for ColE1 DNA, in the presence of Co2+, was found to be
0.4 nM, compared to 3 nM with Mg2+, whereas the turnover was only one
double-stranded scission/min with Co2+ compared to eight/min with Mg2+.  The
implications of all these findings on the enzyme's mechanism are discussed.

<>

<1>Woodhead, J.L., Malcolm, A.D.B.
<2>Non-specific binding of restriction endonuclease EcoRI to DNA.
<3>Nucleic Acids Res.
<4>8
<5>389-402
<6>1980
<7>Restriction endonuclease, EcoR1 cleaves the DNA sequence (5' - 3')G -^ A - A -
T - T - C (3' - 5')C - T - T - A - A -^ G at the points indicated.  Under
certain conditions, EcoRI activity is observed when (5' - 3')N1 -^ A - A - T -
T - N2 (3' - 5')N3 - T - T - A - A -^ N4 is cut.  Mg2+ is required for both
activities.  We find that in addition to binding to the above sites, EcoRI will
also bind, although less strongly, to DNA containing neither site.  Methyl
acetimidate, which reacts specifically with lysine residues, inactivates the
enzyme.  This specific effect can be prevented by SV40 DNA and lambda DNA which
contain EcoRI and EcoRI sites, by PhiX174 DNA, which has only EcoRI sites and
also by Polyd(AT) and polyd(GC) containing neither site.  Protection occurs in
the absence or presence of magnesium.  The significance of this non-specific
binding, both for the use and mechanism of EcoR1 will be discussed.

<>

<1>Woodhead, J.L., Malcolm, A.D.B.
<2>The essential carboxyl group in restriction endonuclease EcoRI.
<3>Eur. J. Biochem.
<4>120
<5>125-128
<6>1981
<7>We have carried out studies on type II restriction endonuclease R.EcoRI, wich
cleaves the DNA sequence 5'd(G^A-A-T-T-C-)3', as indicated.  The active form of
the enzyme consists of two subunits, each 31063 molecular weight.  A
water-soluble reagent, 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide
metho-p-sulphonate, which reacts with carboxyl groups and also with tyrosine
and cysteine residues, has been found to inactivate this enzyme.  Results are
presented which show the following. (1) This specific inactivation is not due
to modification of tyrosine or cysteine residues.  (2) There is one carboxyl
group per subunit which, when modified with carbodiimide, inactivates the
enzyme.  (3) PhiX174 DNA (which does not contain EcoRI sites) partially
protects the enzyme from the carbodiimide; protection is unaffected by the
additional presence of Mg2+, but significantly greater with Co2+ and
PhiX174DNA.

<>

<1>Woodhead, J.L., Malcolm, A.D.B.
<2>Restriction endonuclease EcoRI binds non-specifically to deoxyribonucleic acid.
<3>Biochem. Soc. Trans.
<4>8
<5>90-91
<6>1980
<7>EcoRI is a type-II restriction endonuclease.  These enzymes recognize and cleave specific DNA
sequences.  Despite the widespread use of these enzymes for genetic manipulation and DNA
mapping, little is known about the mechanism of any of them, or how they achieve their
specificity.  EcoRI recognizes and cleaves the DNA sequence 5'...G/A-A-T-T-C...3' at the
point indicated.  This activity occurs at around neutral pH and at salt concentrations above
50mM, but at lower salt concentrations and at pH values above 8, the enzyme exhibits EcoRI
activity when 5'...N/A-A-T-T-N...3' is cut.  Mg2+ is required for both activities.  We have
shown that EcoRI not only binds to DNA containing EcoRI or EcoRI sites but will also bind to
DNA containing neither site.

<>

<1>Woods, D.E., Jeddeloh, J.A., Fritz, D.L., DeShazer, D.
<2>Burkholderia thailandensis E125 harbors a temperate bacteriophage specific for Burkholderia mallei.
<3>J. Bacteriol.
<4>184
<5>4003-4017
<6>2002
<7>Burkholderia thailandensis is a nonpathogenic gram-negative bacillus that is closely related
to Burkholderia mallei and Burkholderia pseudomallei. We found that B. thailandensis E125
spontaneously produced a bacteriophage, termed phiE125, which formed turbid plaques in top
agar containing B. mallei ATCC 23344. We examined the host range of phiE125 and found that it
formed plaques on B. mallei but not on any other bacterial species tested, including B.
thailandensis and B. pseudomallei. Examination of the bacteriophage by transmission electron
microscopy revealed an isometric head and a long noncontractile tail. B. mallei NCTC 120 and
B. mallei DB110795 were resistant to infection with phiE125 and did not produce
lipopolysaccharide (LPS) O antigen due to IS407A insertions in wbiE and wbiG, respectively.
wbiE was provided in trans on a broad-host-range plasmid to B. mallei NCTC 120, and it
restored LPS O-antigen production and susceptibility to phiE125. The 53,373-bp phiE125 genome
contained 70 genes, an IS3 family insertion sequence (ISBt3), and an attachment site (attP)
encompassing the 3' end of a proline tRNA (UGG) gene. While the overall genetic organization
of the phiE125 genome was similar to lambda-like bacteriophages and prophages, it also
possessed a novel cluster of putative replication and lysogeny genes. The phiE125 genome
encoded an adenine and a cytosine methyltransferase, and purified bacteriophage DNA contained
both N6-methyladenine and N4-methylcytosine. The results presented here demonstrate that
phiE125 is a new member of the lambda supergroup of Siphoviridae that may be useful as a
diagnostic tool for B. mallei.

<>

<1>Worden, A.Z. et al.
<2>Green evolution and dynamic adaptations revealed by genomes of the marine picoeukaryotes Micromonas.
<3>Science
<4>324
<5>268-272
<6>2009
<7>Picoeukaryotes are a taxonomically diverse group of organisms less than 2
micrometers in diameter. Photosynthetic marine picoeukaryotes in the genus
Micromonas thrive in ecosystems ranging from tropical to polar and could serve as
sentinel organisms for biogeochemical fluxes of modern oceans during climate
change. These broadly distributed primary producers belong to an anciently
diverged sister clade to land plants. Although Micromonas isolates have high 18S
ribosomal RNA gene identity, we found that genomes from two isolates shared only
90% of their predicted genes. Their independent evolutionary paths were
emphasized by distinct riboswitch arrangements as well as the discovery of
intronic repeat elements in one isolate, and in metagenomic data, but not in
other genomes. Divergence appears to have been facilitated by selection and
acquisition processes that actively shape the repertoire of genes that are
mutually exclusive between the two isolates differently than the core genes.
Analyses of the Micromonas genomes offer valuable insights into ecological
differentiation and the dynamic nature of early plant evolution.

<>

<1>Workentine, M.L., Surette, M.G., Bernier, S.P.
<2>Draft Genome Sequence of Burkholderia dolosa PC543 Isolated from Cystic Fibrosis  Airways.
<3>Genome Announcements
<4>2
<5>e00043-14
<6>2014
<7>Burkholderia dolosa is a member of the Burkholderia cepacia complex, a group of opportunistic
bacterial pathogens often associated with fatal chronic infections
in the lungs of patients suffering from cystic fibrosis (CF). Here, we announce
the draft genome sequence of B. dolosa PC543 (LMG 19468), a CF airway isolate.

<>

<1>Woyke, T. et al.
<2>Complete genome sequence of the gliding freshwater bacterium Fluviicola taffensis type strain (RW262).
<3>Standards in Genomic Sciences
<4>5
<5>21-29
<6>2011
<7>Fluviicola taffensis O'Sullivan et al. 2005 belongs to the monotypic genus Fluviicola within
the family Cryomorphaceae. The species is of interest because
of its isolated phylogenetic location in the genome-sequenced fraction of the
tree of life. Strain RW262(T) forms a monophyletic lineage with uncultivated
bacteria represented in freshwater 16S rRNA gene libraries. A similar
phylogenetic differentiation occurs between freshwater and marine bacteria in the
family Flavobacteriaceae, a sister family to Cryomorphaceae. Most remarkable is
the inability of this freshwater bacterium to grow in the presence of Na(+) ions.
All other genera in the family Cryomorphaceae are from marine habitats and have
an absolute requirement for Na(+) ions or natural sea water. F. taffensis is the
first member of the family Cryomorphaceae with a completely sequenced and
publicly available genome. The 4,633,577 bp long genome with its 4,082
protein-coding and 49 RNA genes is a part of the Genomic Encyclopedia of Bacteria
and Archaea project.

<>

<1>Wright, D.J., Jack, W.E., Modrich, P.
<2>The kinetic mechanism of EcoRI endonuclease.
<3>J. Biol. Chem.
<4>274
<5>31896-31902
<6>1999
<7>Steady-state parameters governing cleavage of pBR322 DNA by EcoRI endonuclease are highly
sensitive to ionic environment, with K(m) and k(cat) increasing 1,000-fold and 15-fold,
respectively, when ionic strength is increased from 0.059 to 0.23 M.  By contrast,
pre-steady-state analysis has shown that recognition, as well as first and second strand
cleavage events that occur once the enzyme has arrived at the EcoRI site, are essentially
insensitive to ionic strength, and has demonstrated that the rate-limiting step for
endonuclease turnover occurs after double-strand cleavage under all conditions tested.
Furthermore, processive cleavage of a pBR322 variant bearing two closely spaced EcoRI sites is
governed by the same turnover number as hydrolysis of parental pBR322, which contains only a
single EcoRI sequence, ruling out slow release of the enzyme from the cleaved site or a slow
conformational change subsequent to double-strand cleavage. We attribute the effects of ionic
strength on steady-state parameters to nonspecific endonuclease.DNA interactions, reflecting
facilitated diffusion processes, that occur prior to EcoRI sequence recognition and subsequent
to DNA cleavage.

<>

<1>Wright, D.J., King, K., Modrich, P.
<2>The negative charge of Glu-111 is required to activate the cleavage center of EcoRI endonuclease.
<3>J. Biol. Chem.
<4>264
<5>11816-11821
<6>1989
<7>King et al. JBC (1989) 264, 11807-11815 have shown that Glu-111 is required for
DNA cleavage by EcoRI endonuclease and have suggested that this residue is
required for activation of the cleavage center upon specific recognition.  We
have substituted Gln or Asp for Glu-111 by oligonucleotide-directed
mutagenesis.  First and second strand cleavage rate constants are reduced by a
factor of more than 10/4 by the Gln-111 substitution.  However, these rate
constants are enhanced 9-fold when pH is increased from 7.6 to 8.5, which
enhances strand cleavage at EcoRI sites by wild type endonuclease to a similar
degree.  The specific affinity of Gln-111 endonuclease for EcoRI sites is 1000
times greater than that of wild type enzyme reflecting a decrease in the rate
constant governing specific complex dissociation.  In contrast to Gln-111
endonuclease, the equilibrium specific affinity of Asp-111 endonuclease for the
EcoRI sequence is similar to that of wild type enzyme, and first and second
strand cleavage rate constants are reduced only 100-fold relative to wild type
enzyme.  These results suggest that a negative charge on residue 111 is
required for strand cleavage and are consistent with participation of Glu-111
in activation of the DNA cleavage center, with energy associated with specific
sequence recognition driving this process.

<>

<1>Wright, R., Stephens, C., Shapiro, L.
<2>The CcrM DNA methyltransferase is widespread in the alpha subdivision of proteobacteria, and its essential functions are conserved in Rhizobium meliloti and Caulobacter crescentus.
<3>J. Bacteriol.
<4>179
<5>5869-5877
<6>1997
<7>the Caulobacter crescentus DNA methyltransferase CcrM (M.CcrMI) methylates the adenine residue
in the sequence GANTC.  The CcrM DNA methyltransferase is essential for viability, but it does
not appear to be part of a DNA restriction-modification system.  CcrM homologs are widespread
in the alpha subdivision of gram-negative bacteria.  We have amplified and sequenced a 258-bp
region of the ccrM gene from several of these bacteria, including Rhizobium meliloti, Brucella
abortus, Agrobacterium tumefaciens, and Rhodobacter capsulatus.  Alignment of the deduced
amino acid sequences revealed that these proteins constitute a highly conserved DNA
methyltransferase family.  Isolation of the full-length ccrM genes from the aquatic bacterium
C. crescentus, the soil bacterium R. meliloti, and the intracellular pathogen B. abortus
showed that this sequence conservation extends over the entire protein.  In at least two alpha
subdivision bacteria, R. meliloti and C. crescentus, CcrM-mediated methylation has important
cellular functions.  In both organisms, CcrM is essential for viability.  Over-expression of
CcrM in either bacterium results in the defects in cell division and cell morphology and in
the initiation of DNA replication.  Finally, the C. crescentus and R. meliloti ccrM genes are
functionally interchangeable, as the complemented strains are viable and the chromosomes are
methylated.  Thus, in both R. meliloti and C. crescentus, CcrM methylation is an integral
component of the cell cycle.  We speculate that CcrM-mediated DNA methylation is likely to
have similar roles among alpha subdivision bacteria.

<>

<1>Wright, R., Stephens, C., Zweiger, G., Shapiro, L., Alley, M.R.K.
<2>Caulobacter Lon protease has a critical role in cell-cyle control of DNA methylation.
<3>Genes Dev.
<4>10
<5>1532-1542
<6>1996
<7>CcrM an adenine DNA methyltransferase, is essential for viability in Caulobacter crescentus.
The CcrM protein is present only in the predivisional stage of the cell cycle, resulting in
cell-cycle-dependent variation of the DNA methylation state of the chromosome.  The
availability of CcrM is controlled in two ways: (1) the ccrM gene is transcribed only in the
predivisional cell, and (2) the CcrM protein is rapidly degraded prior to cell division.  We
demonstrate here that CcrM is an important target of the Lon protease pathway in C.
crescentus.  In a lon null mutant, ccrM transcription is still temporally regulated, but the
CcrM protein is present throughout the cell cycle because of a dramatic increase in its
stability that results in a fully methylated chromosome throughout the cell cycle.  Because
the Lon protease is present throughout the cell cycle, it is likely that the level of CcrM in
the cell is controlled by a dynamic balance between temporally varied transcription and
constitutive degradation.  We have shown previously that restriction of CcrM to the C.
crescentus predivisional cell is essential for normal morphogenesis and progression through
the cell cycle.  Comparison of the lon null mutant strain with a strain whose DNA remains
fully methylated as a result of constitutive expression of ccrM suggests that the effect of
Lon on DNA methylation contributes to several developmental defects observed in the lon
mutant.  These defects include a frequent failure to complete cell abnormalities exhibited by
the lon null mutant, such as the formation of abnormally long stalks, appear to be unrelated
to altered chromosome methylation state.  The Lon protease thus exhibits pleiotropic effects
in C. crescentus growth and development.

<>

<1>Wright, R.J., Stephens, C., Alley, D., Zweiger, G., Shapiro, L.
<2>Cell cycle turnover of a DNA methyltransferase in Caulobacter is dependent on the Lon protease.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>96
<5>527
<6>1996
<7>CcrM, an adenine DNA methyltransferase, is essential for viability in Caulobacter
crescentus.  CcrM is present only in the predivisional state of the Caulobacter cell cycle.
As a
consequence, the DNA methylation state of the chromosome is cell cycle regulated.  CcrM levels
are controlled in two ways (a) transcription of the ccrM gene is limited to the predivisional
cell and
(b) the CcrM protein is degraded prior to cell division.  This transient presence of CcrM is
important, as constitutive expression of ccrM causes developmental defects.  In our efforts to
identify the protease responsible for CcrM turnover, the Caulobacter Lon homolog was isolated.
In a Lon null mutant, CcrM is present throughout the cell cycle.  Thus, the Lon protease is
required
for turnover of CcrM, and is necessary for cell cycle regulation of DNA methylation.  We are
currently constructing mutant alleles of CcrM to examine the requirements of Lon-dependent
CcrM
proteolysis.  We have demonstrated that CcrM homologs are widespread in the alpha-subdivision
of purple gram negative bacteria and are currently cloning several of these homologs.

<>

<1>Wright, S., Wilson, S., Miller, W.G., Mandrell, R.E., Siletzky, R.M., Kathariou, S.
<2>Differences in Methylation at GATC Sites in Genomic DNA of Campylobacter coli from Turkeys and Swine.
<3>Appl. Environ. Microbiol.
<4>76
<5>7314-7317
<6>2010
<7>A significant fraction (46/108, 43%) of swine isolates of Campylobacter coli but none of 81
isolates of C. coli from turkeys had genomic DNA
that was resistant to digestion by MboI, suggesting methylation of
adenines at GATC sites. No consistent association was noted between
antimicrobial resistance and MboI resistance. Seven swine-associated
multilocus sequence typing-based sequence types (STs) were detected
among multiple isolates with MboI-resistant DNA. The data suggest
host-associated DNA modification system(s) specific for adenine at GATC
sites in C. coli from swine.

<>

<1>Wu, Bo., He, M., Feng, H., Zhang, Y., Hu, Q., Zhang, Y.
<2>Construction and Characterization of Restriction-Modification Deficient Mutants in Zymomonas mobilis ZM4.
<3>Chin. J. Appl. Environ. Biol.
<4>19
<5>189-197
<6>2013
<7>Low transformation efficiency is an obstacle to genetic or molecular manipulations in
ethanologen Zymomonas mobilis. In the present study, 5 defective strains were constructed in
Z. mobilis strain ZM4 by inactivating restriction-modification (R-M) system candidate genes.
Inactivation of ZMO0028 (mrr) and ZMO1933 significantly improved electroporation efficiency by
17 folds and 2 folds when ZM4 was transformed with the methylated plasmid DNA. Disruption of
ZMO0575 significantly decreased the transfor ation efficiency when transformed with both
methylated and unmethylated plasmid DNAs. In comparison with other mutants, Zmmrr and Zm1933
displayed high stability. Furthermore, fermentation results showed that R-M mutants did not
significantly alter the major bacterial traits such as growth, glucose utilization and ethanol
yield. In conclusion, R-M systems in Z. mobilis were investigated in this study, and the
characterization of those R-M genes contributed to creating engineering strains suitable for
genet ic and molecular manipulations.

<>

<1>Wu, C.-T., Li, W.-G., Gao, X.-S., Zhang, X.-F.
<2>Bioinformatics analysis of DNA methyltransferase from plants.
<3>Xinan Daxue Xuebao, Ziran Kexueban
<4>32
<5>83-89
<6>2010
<7>The nucleic acid sequences and amino acid sequences of DNA methyltransferase from oil palm,
Prunus, Pisum, Daucus and Arabidopsis, which were registerd in GenBank, were predicted and
analyzed, using bioinformatics tools according to the composition of nucleic acid sequences
and amino acid sequences, physical and chemical properties, homology alignments, signal
peptides, transmembrane topological structure, hydrophobicity and hydrophilicity, secondary
and tertiary structures of protein and molecular phylogenetic evolution.  The results
indicated that the full-length cDNA of DNMT contains 5'-untranslated region, an open reading
frame and 3'-untranslated region, that DNMT has no signal peptides and transmembrane peptides
and is a hydrophilic protein, including two BAH domains and a DNA methylase, and that alpha
helix and random coil are the main components of its secondary structure and all domains bind
in turn by means of electrostatic interaction.

<>

<1>Wu, C.H., Zhang, P., Morales, C.Q., Kiang, D.
<2>Draft Whole-Genome Sequences of Three Shiga Toxin-Producing Escherichia coli O91:H21 Isolates, Two from Hemolytic Uremic Syndrome Patients and One of Porcine   Origin.
<3>Genome Announcements
<4>2
<5>e01000-14
<6>2014
<7>This study presents three genomes of O91:H21 isolates, two from hemolytic uremic  syndrome
patients and one of porcine origin. Genome analyses reveal that one of
the human isolates contains both Shiga toxin-encoding genes (stx1 and stx2), and
all three isolates contain putative adhesin (iha and eaeH) and antibiotic
resistance (ampC) genes.

<>

<1>Wu, C.J., Wang, H.C., Chen, C.S., Shu, H.Y., Kao, A.W., Chen, P.L., Ko, W.C.
<2>Genome Sequence of a Novel Human Pathogen, Aeromonas aquariorum.
<3>J. Bacteriol.
<4>194
<5>4114-4115
<6>2012
<7>Aeromonas aquariorum, a recently described species, is associated with a variety  of human
diseases. We present here the first genome sequence of A. aquariorum
strain AAk1, which was isolated as the sole pathogen from the blood of a patient
with septicemia and necrotizing fasciitis.

<>

<1>Wu, D., Raymond, J., Wu, M., Chatterji, S., Ren, Q., Graham, J.E., Bryant, D.A., Robb, F., Colman, A., Tallon, L.J., Badger, J.H., Madupu, R., Ward, N.L., Eisen, J.A.
<2>Complete genome sequence of the aerobic CO-oxidizing thermophile Thermomicrobium roseum.
<3>PLoS ONE
<4>4
<5>E4207
<6>2009
<7>In order to enrich the phylogenetic diversity represented in the available
sequenced bacterial genomes and as part of an "Assembling the Tree of
Life" project, we determined the genome sequence of Thermomicrobium roseum
DSM 5159. T. roseum DSM 5159 is a red-pigmented, rod-shaped, Gram-negative
extreme thermophile isolated from a hot spring that possesses both an
atypical cell wall composition and an unusual cell membrane that is
composed entirely of long-chain 1,2-diols. Its genome is composed of two
circular DNA elements, one of 2,006,217 bp (referred to as the chromosome)
and one of 919,596 bp (referred to as the megaplasmid). Strikingly, though
few standard housekeeping genes are found on the megaplasmid, it does
encode a complete system for chemotaxis including both chemosensory
components and an entire flagellar apparatus. This is the first known
example of a complete flagellar system being encoded on a plasmid and
suggests a straightforward means for lateral transfer of flagellum-based
motility. Phylogenomic analyses support the recent rRNA-based analyses
that led to T. roseum being removed from the phylum Thermomicrobia and
assigned to the phylum Chloroflexi. Because T. roseum is a deep-branching
member of this phylum, analysis of its genome provides insights into the
evolution of the Chloroflexi. In addition, even though this species is not
photosynthetic, analysis of the genome provides some insight into the
origins of photosynthesis in the Chloroflexi. Metabolic pathway
reconstructions and experimental studies revealed new aspects of the
biology of this species. For example, we present evidence that T. roseum
oxidizes CO aerobically, making it the first thermophile known to do so.
In addition, we propose that glycosylation of its carotenoids plays a
crucial role in the adaptation of the cell membrane to this bacterium's
thermophilic lifestyle. Analyses of published metagenomic sequences from
two hot springs similar to the one from which this strain was isolated,
show that close relatives of T. roseum DSM 5159 are present but have some
key differences from the strain sequenced.

<>

<1>Wu, D.Q., Cheng, H., Wang, C., Zhang, C., Wang, Y., Shao, J., Duan, Q.
<2>Genome Sequence of Pseudomonas aeruginosa Strain AH16, Isolated from a Patient with Chronic Pneumonia in China.
<3>J. Bacteriol.
<4>194
<5>5976-5977
<6>2012
<7>Pseudomonas aeruginosa AH16 is a virulent strain isolated from a patient with chronic
pneumonia in China. Here, we present a 6.8-Mb (G+C content, 66.13%)
assembly of its genome with 6,332 putative coding sequences, which may provide
insights into the genomic basis of activity of the clinical P. aeruginosa strain
in China.

<>

<1>Wu, D.Q., Ye, J., Ou, H.Y., Wei, X., Huang, X., He, Y.W., Xu, Y.
<2>Genomic analysis and temperature-dependent transcriptome profiles of the rhizosphere originating strain Pseudomonas aeruginosa M18.
<3>BMC Genomics
<4>12
<5>438
<6>2011
<7>ABSTRACT: BACKGROUND: Our previously published reports have described an
effective biocontrol agent named Pseudomonas sp. M18 as its 16S rDNA
sequence and several regulator genes share homologous sequences with those
of P. aeruginosa, but there are several unusual phenotypic features. This
study aims to explore its strain specific genomic features and gene
expression patterns at different temperatures. RESULTS: The complete M18
genome is composed of a single chromosome of 6,327,754 base pairs
containing 5684 open reading frames. Seven genomic islands, including two
novel prophage clusters and five specific non-phage islands were
identified besides the conserved P. aeruginosa core genome. Each prophage
contains a putative chitinase coding gene, and the prophage II contains a
capB gene encoding a putative cold stress protein. The non-phage GIs
contain genes responsible for pyoluteorin biosynthesis, environmental
substance degradation and type I and III restriction-modification systems.
Compared with other P. aeruginosa strains, the fewest number (3) of
insertion sequences and the most number (3) of clustered regularly
interspaced short palindromic repeats in M18 genome may contribute to the
relative genome stability. Although the M18 genome is most closely related
to that of P. aeruginosa strain LESB58, the strain M18 is more susceptible
to several antimicrobial agents and easier to be erased in a mouse acute
lung infection model than the strain LESB58. The whole M18 transcriptomic
analysis indicated that 10.6% of the expressed genes are
temperature-dependent, with 22 genes up-regulated at 28 degrees Celsius in
three GIs and one prophage but none at 37 degrees Celsius. CONCLUSIONS:
The P. aeruginosa strain M18 has its evolved specific genomic structures
and temperature dependent expression patterns to meet the requirement of
its fitness and competitiveness under selective pressures imposed on the
strain in rhizosphere niche.

<>

<1>Wu, H., Lippmann, J.E., Oza, J.P., Zeng, M., Fives-Taylor, P., Reich, N.O.
<2>Inactivation of DNA adenine methyltransferase alters virulence factors in Actinobacillus actinomycetemcomitans.
<3>Oral Microbiol. Immunol.
<4>21
<5>238-244
<6>2006
<7>DNA adenine methyltransferase (DAM) plays critical roles in diverse biological pathways in
gram-negative bacteria, and specifically in
regulating the expression of virulence genes in several organisms.
Actinobacillus actinomycetemcomitans plays an important role in the
pathogenesis of juvenile and adult periodontal disease, yet little is
known about its mechanisms of gene regulation. DAM is shown here to
directly or indirectly affect well-known A. actinomycetemcomitans
virulence factors. A mutant A. actinomycetemcomitans strain lacking the
dam gene was created by homologous recombination and shows normal
growth phenotypes when grown exponentially. This mutant strain has four
sixfold increased levels of extracellular leukotoxin, altered cellular
levels of leukotoxin, and significant changes in bacterial invasion of
KB oral epithelial cells. These results provide a basis for further
characterization of regulatory mechanisms that control A.
actinomycetemcomitans virulence.

<>

<1>Wu, H., Qiao, J., Blom, J., Rueckert, C., Reva, O., Gao, X., Borriss, R.
<2>The Rhizobacterium Bacillus amyloliquefaciens subsp. plantarum NAU-B3 Contains a  Large Inversion within the Central Portion of the Genome.
<3>Genome Announcements
<4>1
<5>e00941-13
<6>2013
<7>The genome of rhizobacterium Bacillus amyloliquefaciens subsp. plantarum strain NAU-B3 is
4,196,170 bp in size and harbors 4,001 genes. Nine giant gene clusters
are dedicated to the nonribosomal synthesis of antimicrobial lipopeptides and
polyketides. Remarkably, NAU_B3 contains a large inversion within the central
portion of the genome.

<>

<1>Wu, H.N., Nakura, Y., Motooka, D., Nakamura, S., Nishiumi, F., Ishino, S., Kawai, Y., Tanaka, T., Takeuchi, M., Nakayama, M., Fujita, T., Yanagihara, I.
<2>Complete Genome Sequence of Ureaplasma parvum Serovar 3 Strain SV3F4, Isolated in Japan.
<3>Genome Announcements
<4>2
<5>e00256-14
<6>2014
<7>Here, we present the complete genome sequence of Ureaplasma parvum serovar 3, clinical strain
SV3F4, isolated from a Japanese patient with a history of an
infectious abortion.

<>

<1>Wu, J., Herman, J.G., Wilson, G., Lee, R.Y., Yen, R.-W.C., Mabry, M., de Bustros, A., Nelkin, B.D., Baylin, S.B.
<2>Expression of prokaryotic HhaI DNA methyltransferase is transforming and lethal to NIH 3T3 cells.
<3>Cancer Res.
<4>56
<5>616-622
<6>1996
<7>In neoplastic cells, levels of DNA methyltransferase activity are often increased, and
evidence is accruing to suggest an important role for this event in tumorigenesis.  To
evaluate this possibility further, and to investigate the contribution of increasing de novo,
as opposed to maintenance, DNA methylation in mammalian cells, we expressed the bacterial HhaI
methyltransferase in cultured murine fibroblasts.  This enzyme is a pure de novo DNA
methytransferase that methylates the internal C in the sequence GCGC.  We find that both
constitutive and induced expression of the wild-type HhaI results, primarily, in lethality to
the cells.  However, surviving cell clones that express low levels of M.HhaI demonstrate
increased tumorigenicity as assessed by soft agar cloning efficiency (8.6% for sense
HhaI-transduced PA 317 cells versus 0.4% for antisense controls; 1.7% for sense
HhaI-transfected NIH 3T3 cells versus 0% for a mutant HhaI control) and tumorigenicity in nude
mouse heterotransplants (75% for sense HhaI-transduced PA 317 cells versus 18.5% for antisense
controls).  DNA isolated from the clonogenic sense HhaI clones, versus clones expressing the
mutant HhaI gene, has no increase in overall CpG methylation but an average of 27% (range,
16.7-38.9) increase in methylcytosine content at GCGC sites.  These findings suggest that
eukaryotic cells tolerate a narrow window of increased de novo DNA methylating capacity, above
which cell death occurs and within which cell transformation results.  Our results further
emphasize the potential role of increased DNA methyltransferase activity in the evolution of
cancer.

<>

<1>Wu, J., Issa, J.P., Herman, J., Bassett, D.E., Nelkin, B.D., Baylin, S.B.
<2>Expression of an exogenous eukaryotic DNA methyltransferase gene induces transformation of NIH 3T3 cells.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>90
<5>8891-8895
<6>1993
<7>Abnormal regional increases in DNA methylation, which have potential for causing gene
inactivation and chromosomal instability, are consistently found in immortalized and
tumorigenic cells. Increased DNA methyltransferase activity, which is also a characteristic of
such cells, is a candidate to mediate these abnormal DNA methylation patterns. We now show
that in NIH 3T3 mouse fibroblasts constitutive overexpression of an exogenous mouse DNA
methyltransferase gene results in a marked increase in overall DNA methylation which is
accompanied by tumorigenic transformation. These transformation changes can also be elicited
by dexamethasone-inducible expression of an exogenous DNA methyltransferase gene. Our findings
provide strong evidence that the increase in DNA methyltransferase activity associated with
tumor progression could be a key step in carcinogenesis and provide a model system that can be
used to further study this possibility.

<>

<1>Wu, J.C., Santi, D.V.
<2>Kinetic and catalytic mechanism of HhaI methyltransferase.
<3>J. Biol. Chem.
<4>262
<5>4778-4786
<6>1987
<7>Kinetic and catalytic properties of the DNA (cytosine-5)-methyltransferase HhaI
are described.  With poly(dG-dC) as substrate, the reaction proceeds by an
equilibrium (or processive) ordered Bi-Bi mechanism in which DNA binds to the
enzyme first, followed by S-adenosylmethionine (AdoMet).  After methyl
transfer, S-adenosylhomocysteine (AdoHcy) dissociates followed by methylated
DNA.  AdoHcy is a potent competitive inhibitor with respect to AdoMet (K=2.0
microM) and its generation during reactions results in non-linear kinetics.
AdoMet and AdoHcy significantly interact with only the substrate enzyme-DNA
complex; they do not bind to free enzyme and bind poorly to the methylated
enzyme-DNA complex.  In the absence of AdoMet, HhaI methylase catalyzes
exchange of the 5-H of substrate cytosines for protons of water at about 7-fold
the rate of methylation.  The 5-H exchange reaction is inhibited by AdoMet or
AdoHcy.  In the enzyme-DNA-AdoHcy complex, AdoHcy also suppresses dissociation
of DNA and reassociation of the enzyme with other substrate sequences.  Our
studies reveal that the catalytic mechanism of DNA
(cytosine-5)-methyl-transferases involves attack of the C6 of substrate
cytosines by an enzyme nucleophile and formation of a transient covalent
adduct.  Based on precedents of other enzymes which catalyze similar reactions
and the susceptibilitiy of HhaI to inactivation by N-ethylmaleimide, we propose
that the sulfhydryl group of a cysteine residue is the nucleophilic catalyst.
Furthermore, we propose that Cys-81 is the active-site catalyst in HhaI.  This
residue is found in a Pro-Cys doublet which is conserved in all DNA
(cytosine-5)-methyltransferases whose sequences have been determined to date
and is found in related enzymes.  Finally, we discuss the possibility that
covalent adducts between C6 of pyrimidines and nucleophiles of proteins may be
important general components of protein-nucleic acid interactions.

<>

<1>Wu, J.C., Santi, D.V.
<2>High level expression and purification of HhaI methyltransferase.
<3>Nucleic Acids Res.
<4>16
<5>703-717
<6>1988
<7>A cloning system for the DNA- (cytosine-5) -methyltransferase MHhaI and high
level expression of the enzyme are described.  A parent plasmid was constructed
from fragments of the MHhaI gene and synthetic oligonucleotides.  The construct
permits introduction of various restriction sites for cloning at precise
positions near the initiation codon, and beyond the termination codon.  The
entire MHhaI coding sequence was introduced as a 1042 b.p. NdeI-XbaI fragment
into the vector pAR3040 which contains the T7 RNA polymerase promoter.  The
resultant plasmid pTNX3 (MHhaI-pAR3040) was introduced into McrB- E. coli
strains HB101 and GM2929, and expression of MHhaI was induced by infection with
the lambda phage CE6 carrying the T7 RNA polymerase gene.  In induced cells,
catalytically active MHhaI was produced at a level that corresponds to about 8%
of the total soluble protein; an insoluble form of the protein was also formed,
but could be readily removed.  The expressed soluble enzyme from HB101/pTNX3
was purified to apparent homogeneity in about 50% yield by a two-step
chromatographic procedure involving DEAE-cellulose and Heparin-Sepharose; a one
liter culture gave about 2.5 mg of pure enzyme.  The molecular weight and
kinetic properties of the expressed protein are identical to those reported for
the authentic MHhaI, and its amino terminal sequence agrees with that predicted
from the DNA sequence.

<>

<1>Wu, J.C., Santi, D.V.
<2>On the mechanism and inhibition of DNA cytosine methyltransferases.
<3>Biochemistry and Biology of DNA Methylation, Alan R. Liss, Cantoni, G.L., Razin, A., 
<4>0
<5>119-129
<6>1985
<7>Enzyme catalyzed methylation of the 5-position of pyrimidine nucleotides occurs in a number of
branches of nucleic acid biochemistry. Examples of this reaction include the formation of
thymidine 5'-monophosphate (TMP) from deoxyuridine 5'-monophosphate (dUMP) by thymidylate
synthetase (TS), post-transcriptional methylation of RNA molecules, and the methylation of DNA
cytosine residues by methyltransferases (DCMTases) found in eukaryotic cells and in bacteria.
While thymidylate synthetase uses 5,10-methylenetetrahydrofolate as the methyl group donor,
most of the other enzymes require S-adenosylmethionine (AdoMet) for their methyltransferase
activity.

<>

<1>Wu, J.J., Issa, J.P., Herman, J., Nelkin, B.D., Baylin, S.B.
<2>Expression of an exogenous eukaryotic DNA methyltransferase gene induces transformation of NIH 3T3 cells.
<3>Amer. Soc. Hum. Gen.
<4>4
<5>A74
<6>1992
<7>
<>

<1>Wu, K., Dasgupta, S., Jayaprakash, T.L., Cherian, S.
<2>Zea mays ribulose bisphosphate carboxylase activase promoter.
<3>US Patent Office
<4>US 7750207 B
<5>
<6>2010
<7>The present invention provides gene regulatory element polynucleotide molecules, including a
promoter and a leader, identified from the ribulose biphosphate carboxylase activase Zea mays
gene, useful for expressing transgenes in plants.  Th invention further discloses
compositions, polynucleotide constructs, transformed host cells, transgenic plants and seeds
comprising the Zea mays gene regulatory molecules, and methods for preparing and using the
same.

<>

<1>Wu, K.-Y., Liu, G.-R., Liu, W.-Q., Wang, A.Q., Zhan, S., Sanderson, K.E., Johnston, R.N., Liu, S.-L.
<2>The genome of Salmonella enterica serovar gallinarum: Distinct insertions/deletions and rare rearrangements.
<3>J. Bacteriol.
<4>187
<5>4720-4727
<6>2005
<7>Salmonella enterica serovar Gallinarum is a fowl-adapted pathogen, causing typhoid fever in
chickens. It has the same antigenic formula (1,9,12:-:-) as S. enterica serovar Pullorum,
which is also adapted to fowl but causes pullorum disease (diarrhea). The close relatedness
but distinct pathogeneses make this pair of fowl pathogens good models for studies of
bacterial genomic evolution and the way these organisms acquired pathogenicity. To locate and
characterize the genomic differences between serovar Gallinarum and other salmonellae, we
constructed a physical map of serovar Gallinarum strain SARB21 by using I-CeuI, XbaI, and
AvrII with pulsed-field gel electrophoresis techniques. In the 4,740-kb genome, we located two
insertions and six deletions relative to the genome of S. enterica serovar Typhimurium LT2,
which we used as a reference Salmonella genome. Four of the genomic regions with reduced
lengths corresponded to the four prophages in the genome of serovar Typhimurium LT2, and the
others contained several smaller deletions relative to serovar Typhimurium LT2, including
regions containing srfJ, std, and stj and gene clusters encoding a type I restriction system
in serovar Typhimurium LT2. The map also revealed some rare rearrangements, including two
inversions and several translocations. Further characterization of these insertions,
deletions, and rearrangements will provide new insights into the molecular basis for the
specific host-pathogen interactions and mechanisms of genomic evolution to create a new
pathogen.

<>

<1>Wu, K.M., Li, L.H., Yan, J.J., Tsao, N., Liao, T.L., Tsai, H.C., Fung, C.P., Chen, H.J., Liu, Y.M., Wang, J.T., Fang, C.T., Chang, S.C., Shu, H.Y., Liu, T.T., Chen, Y.T., Shiau, Y.R., Lauderdale, T.L., Su, I.J., Kirby, R., Tsai, S.F.
<2>Genome sequencing and comparative analysis of Klebsiella pneumoniae NTUH-K2044, a strain causing liver abscess and meningitis.
<3>J. Bacteriol.
<4>191
<5>4492-4501
<6>2009
<7>Nosocomial infections caused by antibiotic-resistant Klebsiella pneumoniae are emerging as a
major health problem worldwide, while community-acquired
K. pneumoniae infections present with a range of diverse clinical pictures
in different geographic areas. In particular, an invasive form of K.
pneumoniae that causes liver abscesses was first observed in Asia and then
was found worldwide. We are interested in how differences in gene content
of the same species result in different diseases. Thus, we sequenced the
whole genome of K. pneumoniae NTUH-K2044, which was isolated from a
patient with liver abscess and meningitis, and analyzed differences
compared to strain MGH 78578, which was isolated from a patient with
pneumonia. Six major types of differences were found in gene clusters that
included an integrative and conjugative element, clusters involved in
citrate fermentation, lipopolysaccharide synthesis, and capsular
polysaccharide synthesis, phage-related insertions, and a cluster
containing fimbria-related genes. We also conducted comparative genomic
hybridization with 15 K. pneumoniae isolates obtained from
community-acquired or nosocomial infections using tiling probes for the
NTUH-K2044 genome. Hierarchical clustering revealed three major groups of
genomic insertion-deletion patterns that correlate with the strains'
clinical features, antimicrobial susceptibilities, and virulence
phenotypes with mice. Here we report the whole-genome sequence of K.
pneumoniae NTUH-K2044 and describe evidence showing significant genomic
diversity and sequence acquisition among K. pneumoniae pathogenic strains.
Our findings support the hypothesis that these factors are responsible for
the changes that have occurred in the disease profile over time.

<>

<1>Wu, M. et al.
<2>Phylogenomics of the reproductive parasite Wolbachia pipientis wMeI: A streamlined genome overrun by mobile genetic elements.
<3>PLoS Biology
<4>2
<5>327-341
<6>2004
<7>The complete sequence of the 1,267,782 bp genome of Wolbachia pipientis wMel, an obligate
intracellular bacteria of Drosophila melanogaster, has been determined. Wolbachia, which are
found in a variety of invertebrate species, are of great interest due to their diverse
interactions with different hosts, which range from many forms of reproductive parasitism to
mutualistic symbioses. Analysis of the wMel genome, in particular phylogenomic comparisons
with other intracellular bacteria, has revealed many insights into the biology and evolution
of wMel and Wolbachia in general. For example, the wMel genome is unique among sequenced
obligate intracellular species in both being highly streamlined and containing very high
levels of repetitive DNA and mobile DNA elements. This observation, coupled with multiple
evolutionary reconstructions, suggests that natural selection is somewhat inefficient in wMel,
most likely owing to the occurrence of repeated population bottlenecks. Genome analysis
predicts many metabolic differences with the closely related Rickettsia species, including the
presence of intact glycolysis and purine synthesis, which may compensate for an inability to
obtain ATP directly from its host, as Rickettsia can. Other discoveries include the apparent
inability of wMel to synthesize lipopolysaccharide and the presence of the most genes encoding
proteins with ankyrin repeat domains of any prokaryotic genome yet sequenced. Despite the
ability of wMel to infect the germline of its host, we find no evidence for either recent
lateral gene transfer between wMel and D. melanogaster or older transfers between Wolbachia
and any host. Evolutionary analysis further supports the hypothesis that mitochondria share a
common ancestor with the alpha-Proteobacteria, but shows little support for the grouping of
mitochondria with species in the order Rickettsiales. With the availability of the complete
genomes of both species and excellent genetic tools for the host, the wMel-D. melanogaster
symbiosis is now an ideal system for studying the biology and evolution of Wolbachia
infections.

<>

<1>Wu, P., Qiu, C., Sohail, A., Zhang, X., Bhagwat, A.S., Cheng, X.
<2>Mismatch repair in methylated DNA. Structure and activity of the mismatch-specific thymine glycosylase domain of methyl-CpG-binding protein MBD4.
<3>J. Biol. Chem.
<4>278
<5>5285-5291
<6>2003
<7>MBD4 is a member of the methyl-CpG-binding protein family. It contains two DNA binding
domains, an amino-proximal methyl-CpG binding domain (MBD) and
a C-terminal mismatch-specific glycosylase domain. Limited in vitro
proteolysis of mouse MBD4 yields two stable fragments: a 139-residue
fragment including the MBD, and the other 155-residue fragment including
the glycosylase domain. Here we show that the latter fragment is active as
a glycosylase on a DNA duplex containing a G:T mismatch within a CpG
sequence context. The crystal structure confirmed the C-terminal domain is
a member of the helix-hairpin-helix DNA glycosylase superfamily. The MBD4
active site is situated in a cleft that likely orients and binds DNA.
Modeling studies suggest the mismatched target nucleotide will be flipped
out into the active site where candidate residues for catalysis and
substrate specificity are present.

<>

<1>Wu, Q., Liu, Z., Li, Y., Guan, G., Niu, Q., Chen, Z., Luo, J., Yin, H.
<2>Genome Sequence of Borrelia garinii Strain SZ, Isolated in China.
<3>Genome Announcements
<4>2
<5>e00010-14
<6>2014
<7>We announce the genome sequence of Borrelia garinii strain SZ, isolated from Dermacentor ticks
collected in northeastern China. B. garinii strain SZ carries
numerous plasmids, both 10 circular and 9 linear plasmids. The 902,487-bp linear
chromosome (28.2% GC content) contains 820 open reading frames, 33 tRNAs, and 4
complete rRNAs. The plasmid cp32-10 contains one clustered regularly interspaced
short palindromic repeat (CRISPR) with four repeats.

<>

<1>Wu, Q., Peng, S., Yu, Y., Li, Y., Xu, Y.
<2>Genome Sequence of Bacillus licheniformis CGMCC3963, a Stress-Resistant Strain Isolated in a Chinese Traditional Solid-State Liquor-Making Process.
<3>Genome Announcements
<4>1
<5>e00060-12
<6>2013
<7>Bacillus licheniformis CGMCC3963 is an important mao-tai flavor-producing strain. It was
isolated from the starter (Daqu) of a Chinese mao-tai-flavor liquor
fermentation process with solid-state fermentation. We report its genome of
4,525,096 bp here. Many potential insertion genes that are responsible for the
unique properties of B. licheniformis CGMCC3963 in mao-tai-flavor liquor
production were identified.

<>

<1>Wu, Q., Tun, H.M., Leung, F.C.C., Shah, N.P.
<2>Genomic insights into high exopolysaccharide-producing dairy starter bacterium Streptococcus thermophilus ASCC 1275.
<3>Sci. Rep.
<4>4
<5>4974
<6>2014
<7>
<>

<1>Wu, R., King, C.T., Jay, E.
<2>A new sequence-specific endonuclease from Streptococcus faecalis subsp. zymogenes.
<3>Gene
<4>4
<5>329-336
<6>1978
<7>A new sequence-specific endonuclease, SfaI, has been partially purified from
Streptococcus faecalis subsp. zymogenes.  SfaI recognizes the tetranucleotide
sequence.

<>

<1>Wu, R.S., Hurst-Calderone, S., Kohn, K.W.
<2>Measurement of O6-alkylguanine-DNA alkyltransferase activity in human cells and tumor tissues by restriction endonuclease inhibition.
<3>Cancer Res.
<4>47
<5>6229-6235
<6>1987
<7>A sensitive assay for O6-alkylguanine-DNA alkyltransferase activity in cell or
tumor extracts has been devised.  The theoretical basis of the new assay lies
in the observation that certain restriction enzymes will not cleave DNA
containing methylated bases.  Thus, if a synthetic oligodeoxynucleotide with a
restriction sequence containing O6-methylguanine wee incubated with the
restriction enzyme, this synthetic oligodeoxynucleotide should remain intact.
However, if the guanine-O6 methyl group were first removed by
O6-alkylguanine-DNA alkyltransferase present in certain cell or tissue extracts
the synthetic oligodeoxynucleotide would be cleaved by the restriction enzyme.
The parental oligodeoxynucleotide and its restriction products are separated
from each other and analyzed on denaturing polyacrylamide gels.  The extent of
cleavage by the restriction enzyme is a direct assay of the content of
O6-alkylguanine-DNA alkyltransferase in the cell/tumor extracts.  The assay has
been tested against cell culture and xenograft tumor systems and has performed
in a predictive manner, correctly predicting five Mer- and three Mer+
phenotypes.  Furthermore, the assay is quantitative and the number of molecules
of the O6-alkylguanine-DNA alkyltransferase per cell estimated using this assay
agrees with those that have been published.

<>

<1>Wu, S.Y., Lee, K.F., Kam, K.M., Shaw, P.C.
<2>Restriction enzyme BliHKI from a thermophilic Bacillus licheniformis strain.
<3>Biosci. Biotechnol. Biochem.
<4>57
<5>1193-1194
<6>1993
<7>Thermophilic Bacillus is a rich source of restriction enzymes. Using a screening method
described in ref. 1, we have isolated a thermophilic Bacillus licheniformis strain HK which
grows at 55 degrees C in L broth and contains a type II restriction enzyme, BliHKI.

<>

<1>Wu, T.-H., Grelland, E., Boye, E., Marinus, M.G.
<2>Identification of a weak promoter for the dam gene of Escherichia coli.
<3>Biochim. Biophys. Acta
<4>1131
<5>47-52
<6>1992
<7>We have used a combination of techniques to identify a weak promoter located about 70
nucleotides before the start site of translation of the Escherichia coli dam gene which
encodes a DNA methyltransferase.  The promoter activity was identified by the use of lacZ
fusions to fragments containing different lengths of upstream DNA.  In vitro run-off
transcription and primer extension determinations revealed transcription initiation sites at
either 69 or 73 nucleotides prior to the ARG of the dam coding sequence.  No ribosome binding
sequence was present close to the ATG codon suggesting that the transcript may be
inefficiently translated.

<>

<1>Wu, T.T.
<2>Locus determining P1 phage restriction in Escherichia coli.
<3>J. Bacteriol.
<4>98
<5>314
<6>1969
<7>The locus determing P1 phage restriction has been mapped at 89.3 min on the
Escherichia coli map, about 0.2 min away from the hsp marker.

<>

<1>Wu, T.T., Matsuda, A., Cavalieri, L.F.
<2>The restriction and modification enzymes of Escherichia coli.
<3>Fed. Proc.
<4>28
<5>465
<6>1969
<7>An E. coli endonuclease (MW ca. 78,000) which acts specifically on unmodified
lambda-DNA has been purified.  The endonuclease fraction, together with
S-adenosylmethionine (SAM) and another protein fraction, is required for the
methylation of DNA.  SAM can be eliminated from the methylation reaction, if it
is first allowed to react with the endonuclease fraction, forming a
methyl-donating complex.  The same enzyme fraction from a restrictionless
mutant has no endonucleaese activity but retains the property of reacting with
SAM.  The formation of this methyl-donating complex is partially inhibited by a
protein present only in modificationless mutants of E. coli.  This protein (MW
da. 52,000) has also been purified.  Recombinants having mostly E. coli B
chromosomes except for small segments which contain the restriction and
modification markers from E. coli K12 have been selected.  The DNA of these
recombinants contains both methylcytosine and methyladenine, whereas E. coli
and DNA contains only the later.

<>

<1>Wu, W., Wood, D.W., Belfort, G., Derbyshire, V., Belfort, M.
<2>Intein-mediated purification of cytotoxic endonuclease I-TevI by insertional inactivation and pH-controllable splicing.
<3>Nucleic Acids Res.
<4>30
<5>4864-4871
<6>2002
<7>An intein-mediated approach was developed for expression and affinity purification of a
protein that is lethal to Escherichia coli. The protein, I-TevI, is an intron-encoded
endonuclease. The approach involved the insertional inactivation of I-TevI with a controllable
mini-intein placed in front of a cysteine required for splicing (an I-TevI::intein fusion).
The purification was facilitated by a chitin-binding domain inserted into the mini-intein.
Affinity purification of the I-TevI::intein fusion precursor on a chitin column was followed
by pH-controllable splicing to restore the structure and function of I-TevI. To study the
impact of the insertion context on I-TevI inactivation, the chimeric intein was inserted
independently in front of seven cysteines of I-TevI. One of the seven intein integrants
yielded I-TevI of high activity. This technique is, in principle, generalizable to the
expression and purification of other cytotoxic proteins and is amenable to scale-up.

<>

<1>Wu, W., Zheng, H., Zhang, L., Wen, Z., Zhang, S., Pei, H., Yu, G., Zhu, Y., Cui, Z., Hu, Z., Wang, H., Li, Y.
<2>A genome-wide analysis of multidrug-resistant and extensively drug-resistant strains of Mycobacterium tuberculosis Beijing genotype.
<3>Mol. Genet. Genomics
<4>288
<5>425-436
<6>2013
<7>The Beijing genotype of Mycobacterium tuberculosis (MTB) is one of the most successful MTB
lineages that has disseminated in the world. In China, the rate of
multidrug-resistant (MDR) tuberculosis is significantly higher than the global
average rate, and the Beijing genotype strains take the largest share of MDR
strains. To study the genetic basis of the epidemiological findings that Beijing
genotype has often been associated with tuberculosis outbreaks and drug
resistance, we determined the genome sequences of four clinical isolates: two
extensively drug resistant (XDR1219, XDR1221) and two multidrug resistant (WX1,
WX3), using whole-genome sequencing. A large number of individual and shared SNPs
of the four Beijing strains were identified. Our isolates harbored almost all
classic drug resistance-associated mutations. The mutations responsible for drug
resistance in the two XDR strains were consistent with the clinical quantitative
drug resistance levels. COG analysis revealed that Beijing strains have
significantly higher abundances of the mutations responsible for cell
wall/membrane/envelope biogenesis (COG M), secondary metabolites biosynthesis,
transport and catabolism (COG Q), lipid transport and metabolism (COG I) and
defense mechanisms (COG V). The shared mutated genes of the four studied Beijing
strains were significantly overrepresented in three DNA repair pathways. Our
analyses promote the understanding of the genome polymorphism of the Beijing
family strains and provide the molecular genetic basis for their wide
dissemination capacity and drug resistance.

<>

<1>Wu, Y.R., Li, Y., Yang, K.L., He, J.
<2>Draft Genome Sequence of Butanol-Acetone-Producing Clostridium beijerinckii Strain G117.
<3>J. Bacteriol.
<4>194
<5>5470-5471
<6>2012
<7>A recently discovered wild-type strain, Clostridium beijerinckii G117, is unique  in producing
butanol and acetone but negligible amounts of ethanol, unlike
previously identified acetone-butanol-ethanol (ABE)-generating microbes. Here we
report the draft genome sequence of strain G117 (5,806,675 bp; GC content, 29.7%)
and the novel findings obtained from its genome annotations.

<>

<1>Wu, Y.R., Lin, B., Yu, Y.
<2>Draft Genome Sequence of a Xylanase-Producing Bacterial Strain, Cellvibrio mixtus J3-8.
<3>Genome Announcements
<4>2
<5>e01281-14
<6>2014
<7>The xylanase-producing bacterial strain Cellvibrio mixtus J3-8 was isolated from  grassland
giant snails. The draft genome of strain J3-8 comprises 5,171,890 bp in
152 contigs with a G+C content of 46.66%. This is the first genome report about
this bacterial species.

<>

<1>Wulff, N.A., Zhang, S., Setubal, J., Almeida, N.F., Martins, E.C., Harakava, R., Kumar, D., Rangel, L.T., Foissac, X., Bove, J.M., Gabriel, D.W.
<2>The complete genome sequence of Candidatus Liberibacter americanus, associated with citrus Huanglongbing.
<3>Mol. Plant Microbe Interact.
<4>27
<5>163-176
<6>2014
<7>Liberibacters form a Rhizobiaceae clade of phloem-limited pathogens of limited host range.
Two obligately parasitic species have been sequenced: Candidatus Liberibacter asiaticus Las),
which causes citrus Huanglongbing (HLB) world-wide, and Ca. L. solanacearum (Lso), which
causes potato "zebra chip" disease. A third species, Liberibacter crescens (Lcr),
was isolated from mountain papaya, grown in axenic culture and sequenced. In an effort to
identify common host determinants, the complete genomic DNA sequence of a second HLB species,
Ca. L. americanus (Lam) strain "São Paulo" was determined. The circular genome of 1,195,201 bp
had an average 31.12% GC content and 983 predicted protein encoding genes, 800 (81.4%) of
which had a predicted function. There were 658 genes common to all sequenced liberibacters and
only 8 genes common to Lam and Las but not found in Lso.   Surprisingly, most of the
lipopolysaccharide biosynthetic genes were missing from the Lam genome, as well OmpA and a key
regulator of flagellin, all indicating a Lam strategy of avoiding production of major
pathogen-associated molecular patterns (PAMPs) present in Las and Lso. As with Las, one of two
Lam prophages replicated as an excision plasmid and carried potential lysogenic conversion
genes that appeared fragmentary or degenerated in Lso.

<>

<1>Wyczechowska, D., Fabianowska-Majewska, K.
<2>Does 2-chlorodeoxyadenosine contribute to alteration of DNA methyltransferase activity?
<3>Purine and Pyrimidine Metabolism in Man IX, Plenum Press, Griesmacher, A., Chiba, P., Muller, M.M., New York
<4>
<5>595-598
<6>1998
<7>2-Chlorodeoxyadenosine (2CdA) is a new and effective drug for indolent lymphoid malignancies.
However, the mechanisms which link 2CdA action and tumor cell death (by apoptosis) are not
fully clarified.  2CdA is rapidly taken up by the target cells and phosphorylated by cytosolic
deoxycytidine kinase (dCK).  Its triphosphate is a potent inhibitor of human ribonucleotide
reductase and a good substrate for human DNA polymerases.  However, phosphorylation of 2CdA is
clearly not the only event responsible for its cytotoxic effect.  The 2CdA phosphorylation in
hairy cell leukemia was not higher than in chronic lymphocytic leukemia, despite a better
response to 2CdA therapy.  Using different cell lines, it was shown that  a wide range of cell
sensitivity to 2CdA could not be explained by different levels of 2CdA nucleotide.  2CdA was
also shown to inhibit the growth of myeloid progenitor cell in which the levels of dCK are
low.  Finally, we have recently shown that in vitro the inhibitory effect of 2CdA results from
complete suppression of deoxyadenosine phosphorylation by dCK in both human normal lymphocytes
and in lymphoma cells obtained from patients with a central nervous system involvement.  Also
an inhibition of adenosine deaminase activity in lysate of both types of cells was observed.
Moreover, we observed a large decrease in the activity of adenosine deaminase and
S-adenosylhomocysteine hydrolase, in the erythrocyte lysates of patients, after one week
treatment with 2CdA6.  We assumed that inhibitory effect of 2CdA on deoxyadenosine metabolism
could lead to inactivation of SAH-hydrolase with perturbation of methylation reactions.

<>

<1>Wynne, J.W., Seemann, T., Bulach, D.M., Coutts, S.A., Talaat, A.M., Michalski, W.P.
<2>Resequencing the Mycobacterium avium subsp. paratuberculosis K10 Genome: Improved Annotation and Revised Genome Sequence.
<3>J. Bacteriol.
<4>192
<5>6319-6320
<6>2010
<7>We report the resequencing and revised annotation of the Mycobacterium avium subsp.
paratuberculosis K10 genome. A total of 90 single-nucleotide
errors and a 51-bp indel in the original K10 genome were corrected, and
the whole genome annotation was revised. Correction of these sequencing
errors resulted in 28 frameshift alterations. The amended genome sequence
is accessible via the supplemental section of study SRR060191 in the NCBI
Sequence Read Archive and will serve as a valuable reference genome for
future studies.

<>

<1>Wyszomirski, K.H., Curth, U., Alves, J., Mackeldanz, P., Moncke-Buchner, E., Schutkowski, M., Kruger, D.H., Reuter, M.
<2>Type III restriction endonuclease EcoP15I is a heterotrimeric complex containing  one Res subunit with several DNA-binding regions and ATPase activity.
<3>Nucleic Acids Res.
<4>40
<5>3610-3622
<6>2012
<7>For efficient DNA cleavage, the Type III restriction endonuclease EcoP15I communicates with
two inversely oriented recognition sites in an ATP-dependent process. EcoP15I consists of
methylation (Mod) and restriction (Res) subunits forming a multifunctional enzyme complex able
to methylate or to cleave DNA. In this study, we determined by different analytical methods
that EcoP15I contains a single Res subunit in a Mod(2)Res stoichiometry. The Res subunit
comprises a translocase (Tr) domain carrying functional motifs of superfamily 2 helicases and
an endonuclease domain with a PD..D/EXK motif. We show that the isolated Tr domain retains
ATP-hydrolyzing activity and binds single- and double-stranded DNA in a sequence-independent
manner. To localize the regions of DNA binding, we screened peptide arrays representing the
entire Res sequence for their ability to interact with DNA. We discovered four DNA-binding
regions in the Tr domain and two DNA-binding regions in the endonuclease domain. Modelling of
the Tr domain shows that these multiple DNA-binding regions are located on the surface, free
to interact with DNA. Interestingly, the positions of the DNA-binding regions are conserved
among other Type III restriction endonucleases.

<>

<1>Wyszynski, M.W.
<2>Investigation of the molecular mechanism and mutagenic effect of C5-methyltransferases by mutational analysis.
<3>Diss. Abstr.
<4>55
<5>1426B-1427B
<6>1994
<7>It has been proposed that a cysteine conserved among all DNA (cytosine-5)-methyltransferases
initiates catalysis by attacking the C6 of cytosine. I have changed this cysteine to other
amino acids for the E. coli methylase M.EcoRII; which methylates the second cytosine in the
sequence 5'-CCWGG-3'. Replacement of the conserved cysteine with glycine, valine, tryptophan
or serine led to an apparent loss of methyl transferring ability. Unexpectedly, substitution
of the cysteine with glycine results in the inhibition of cell growth. In DNA binding studies
I show that mutants with either serine or glycine substitution bind tightly to substrate DNA
in a manner which resembles the wild-type enzyme. Hence the conserved cysteine is not
essential for the specific stable binding of the enzyme to its substrate. Further, I show that
a DNA substrate for M.EcoRII in which the target cytosine is replaced by 5-fluorocytosine is a
mechanism-based inhibitor of the enzyme. As expected, this modified substrate does not form
irreversible complexes with the mutants. Sites of cytosine methylation are hot-spots for
cytosine (C) to thymine (T) mutations in E. coli DNA. To study this phenomenon, I have
developed a genetic system based on the reversion of a mutation in a kanamycin-resistance gene
that allows direct selection of C to T mutations at a site of methylation. Using this system I
show that enzyme-catalyzed deaminations of cytosine do not play a major role in making Dcm
methylation sites hot-spots for mutations. Further, I have developed a genetic reversion assay
that quantitates the frequency of C to T mutations at Dcm sites and the reduction of such
mutations by DNA repair processes. Using this assay, the repair of U:G mismatches in DNA to
C:G have been studied. The results shown here demonstrate that the E. coli base mismatch
correction system called VSP repair is capable of correcting U:G mismatches.

<>

<1>Wyszynski, M.W., Gabbara, S., Bhagwat, A.S.
<2>Substitutions of a cysteine conserved among DNA cytosine methylases result in a variety of phenotypes.
<3>Nucleic Acids Res.
<4>20
<5>319-326
<6>1992
<7>The proposed mechanism for DNA (cytosine-5)-methyltransferases envisions a key
role for a cysteine residue.  It is expected to form a covalent link with
carbon 6 of the target cytosine, activating the normally inactive carbon 5 for
methyl transfer.  There is a single conserved cysteine among all DNA
(cytosine-5)-methyltransferases making it the candidate nucleophile.  We have
changed this cysteine to other amino acids for the EcoRII methylase; which
methylates the second cytosine in the sequence 5'-CCWGG-3'.  Mutants were
tested for their methyl transferring ability and for their ability to form
covalent complexes with DNA.  The latter property was tested indirectly with
the use of a genetic assay involving sensitivity of cells to 5-azacytidine.
Replacement of the conserved cysteine with glycine, valine, tryptophan or
serine led to an apparent loss of methyl transferring ability.  Interestingly,
cells carrying the mutant with serine did show sensitivity to 5-azacytidine,
suggesting the ability to link to DNA.  Unexpectedly, substitution of the
cysteine with glycine results in the inhibition of cell growth and the mutant
allele can be maintained in the cells only when it is poorly expressed.  These
results suggest that the conserved cysteine in the EcoRII methylase is
essential for methylase action and it may play more than one role in it.

<>

<1>Wyszynski, M.W., Gabbara, S., Kubareva, E.A., Romanova, E.A., Oretskaya, T.S., Gromova, E.S., Shabarova, Z.A., Bhagwat, A.S.
<2>The cysteine conserved among DNA cytosine methylases is required for methyl transfer, but not for specific DNA binding.
<3>Nucleic Acids Res.
<4>21
<5>295-301
<6>1993
<7>All DNA (cytosine-5)-methyltransferases contain a single conserved cysteine. It has been
proposed that this cysteine initiates catalysis by attacking the C6 of cytosine and thereby
activating the normally inert C5 position. We show here that substitutions of this cysteine in
the E. coli methylases M.EcoRII with either serine or trypotophan results in a complete loss
of ability to transfer methyl groups to DNA. Interestingly, mutants with either serine or
glycine substitution bind tightly to substrate DNA. These mutants resemble the wild-type
enzyme in that their binding to substrate is not eliminated by the presence of non-specific
DNA in the reaction, it is sensitive to methylation status of the substrate and is stimulated
by an analog of the methyl donor. Hence the conserved cysteine is not essential for the
specifc stable binding of the enzyme to its substrate. However, substitution of the cysteine
with the bulkier tryptophan does reduce DNA binding. We also report here a novel procedure for
the synthesis of DNA containing 5-fluorocytosine. Further, we show that a DNA substrate for
M.EcoRII in which the target cytosine is replaced by 5-fluorocytosine is a mechanism-based
inhibitor of the enzyme and that it forms an irreversible complex with the enzyme. As
expected, this modified substrate does not form irreversible complexes with the mutants.

<>

<1>Wyszynski, W., Garbara, S., Bhagwat, A.S.
<2>Cytosine deaminations catalyzed by DNA cytosine methyltransferases are unlikely to be the major cause of mutational hot spots at sites of cytosine methylation in Escherichia coli.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>91
<5>1574-1578
<6>1994
<7>Sites of cytosine methylation are hot spots for C to T mutations in Escherichia coli DNA. We
have developed a genetic reversion assay that allows direct selection of C to T mutations at a
site of methylation. Because the mutant gene is on a plasmid, this system can be used to study
mutational effects of biochemical agents in vitro as well as in vivo. Using this system we
show that in vitro an E. coli methyltransferase can cause C to U deaminations at a site of
methylation. Reaction conditions that are known to inhibit a side reaction of the
methyltransferase also suppress reversion frequency, suggesting that this side reaction is
required for deamination. Furthermore, a mutation in the enzyme that eliminates its catalytic
activity but not its ability to bind DNA eliminates the ability of the enzyme to cause C to U
deaminations. Despite this, in vivo experiments strongly suggest that enzyme-catalyzed
deaminations of cytosine do not play a major role in making methylation sites in E. coli hot
spots for mutations. For example, although uracil-DNA glycosylase (Ung) suppresses the
occurrence of mutations due to C to U deaminations, the frequency of C to T mutations at a
methylation site remains high in ung+ cells. Furthermore, the reversion frequencies in ung+
and ung- cells are quite similar.

<>

<1>Xavier, B.B., Vervoort, J., Stewardson, A., Adriaenssens, N., Coenen, S., Harbarth, S., Goossens, H., Malhotra-Kumar, S.
<2>Complete Genome Sequences of Nitrofurantoin-Sensitive and -Resistant Escherichia coli ST540 and ST2747 Strains.
<3>Genome Announcements
<4>2
<5>e00239-14
<6>2014
<7>Widespread multidrug resistance in Escherichia coli has necessitated the
reintroduction of older antibiotics, such as nitrofurantoin. However, mechanisms
by which resistance to nitrofurantoin emerges in E. coli are not well elucidated.
Toward this aim, we sequenced two nitrofurantoin-sensitive E. coli sequence types
(ST540 and ST2747) and their four nitrofurantoin-resistant derivatives generated
in vitro under aerobic and anaerobic growth conditions.

<>

<1>Xi, J., Sheng, X., He, L.
<2>Draft Genome Sequence of Rhizobium sp. H41, a Rock-Weathering Bacterium from a Weathered Rock Surface.
<3>Genome Announcements
<4>2
<5>e01127-14
<6>2014
<7>Rhizobium sp. H41 isolated from weathered tuff can weather tuff and release Fe, Si, and Al
from the rock under nutrient-poor conditions. Here, we report the
draft genome sequence of strain H41, which may facilitate a better understanding
of the molecular mechanism involved in rock weathering by the bacterium.

<>

<1>Xia, E., Khong, W.X., Marimuthu, K., Xu, W., Ong, R.T., Tan, E.L., Krishnan, P.U., Ang, B.S., Lye, D.C., Chow, A.L., Teo, Y.Y., Ng, O.T.
<2>Draft Genome Sequence of a Multidrug-Resistant New Delhi Metallo-beta-Lactamase-1 (NDM-1)-Producing Escherichia coli Isolate Obtained in Singapore.
<3>Genome Announcements
<4>1
<5>e01020-13
<6>2013
<7>We report the draft genome sequence of a New Delhi metallo-beta-lactamase-1 (NDM-1)-positive
Escherichia coli isolate obtained from a surgical patient. The
assembled data indicate the presence of 3 multidrug resistance plasmids, 1 of
which shares 100% identity with an NDM-1 plasmid isolated previously from a
nearby hospital, suggesting possible local transmission.

<>

<1>Xia, H., Wu, S.
<2>Construction of DNA transfer system of Streptomyces tenebrarius.
<3>Wei Sheng Wu Xue Bao
<4>42
<5>181-185
<6>2002
<7>To establish a gene transfer system in Streptomyces tenebrarius, several methods including
PEG-mediated transformation of protoplasts,
conjugal transfer were investigated. Many attempts were made to
introduce plasmid pIJ702 into Streptomyces tenebrarius. It was found
that plasmid pIJ702 isolated from S. lividans TK24 failed to transform
the protoplasts of Streptomyces tenebrarius. No transformant was
achieved even if the protoplast was inactivated by heat treatment or
dsDNA was converted ssDNA before transformation. All the results
suggested that Streptomyces tenebrarius exists a strong restriction and
modification system for the transformation of foreign DNA. A
recombinant E. coli ET12567 (pUZ8002, pHZ132) was obtained by
transforming E. coli ET12567 (pUZ8002) with oriT-containing E.
coli-Streptomyces shuttle plasmid pHZ132. In mating experiments, E. coli
ET12567 (pUZ8002, pHZ132) was the donor, and the recipient was
Streptomyces tenebrarius 9904 spores after pregerminating by heat shock.
Matings between donor and recipient were conducted. Plasmid pHZ132 was
introduced into Streptomyces tenebrarius 9904 by conjugation from E.
coli ET12567. The transfer system of Streptomyces tenebrarius was
established by conjugation. S. tenebrarius 9904 protoplasts were
transformed by plasmid DNA modified by the host itself, and the
transformation frequency was about 10^3/mug DNA (pHZ132).

<>

<1>Xia, Y., Burbank, D.E., Uher, L., Rabussay, D., Van Etten, J.L.
<2>Restriction endonuclease activity induced by PBCV-1 virus infection of a chlorella-like green alga.
<3>Mol. Cell. Biol.
<4>6
<5>1430-1439
<6>1986
<7>An enzyme was isolated from a eucaryotic, Chlorella-like green alga infected with the virus
PBCV-1 which exhibits type II restriction endonuclease activity. The enzyme recognized the
sequence GATC and cleaved DNA 5' to the G. Methylation of deoxyadenosine in the GATC sequence
inhibited enzyme activity. In vitro the enzyme cleaved host Chlorella nuclear DNA but not
viral DNA because host DNA contains GATC and PBCV-1 DNA contains GmATC sequences.  PBCV-1 DNA
is probably methylated in vivo by the PBCV-1-induced methyltransferase described elsewhere
(Y.Xia and J.L. Van Etten, Mol. Cell. Biol. 6: 1440-1445). Restriction endonuclease activity
was first detected 30 to 60 min after viral infection; the appearance of enzyme activity
required de novo protein synthesis, and the enzyme is probably virus encoded.  Appearance of
enzyme activity coincided with the onset of host DNA degradation after PBCV-1 infection.  We
propose that the PBCV-1-induced restriction endonuclease participates in host DNA degradation
and is part of a virus-induced restriction and modification system in PBCV-1-infected
Chlorella cells.

<>

<1>Xia, Y., Burbank, D.E., Uher, L., Rabussay, D., Van Etten, J.L.
<2>IL-3A virus infection of a Chlorella-like green alga induces a DNA restriction endonuclease with novel sequence specificity.
<3>Nucleic Acids Res.
<4>15
<5>6075-6090
<6>1987
<7>A type II restriction endonuclease, named CviJI, was isolated from a eukaryotic Chlorella-like
green alga infected with the dsDNA containing virus IL-3A. CviJI is the first restriction
endonuclease to recognize the sequence PuGCPy; CviJI cleaves DNA between the G and C.
Methylation of the cytosine in PuGCPy sequences prevents cleavage by CviJI. CviJI cleaved DNA
into smaller but defined fragments in the presence of ATP. This "star activity" was stimulated
by dithiothreitol and/or S-adenosylmethionine but did not occur under conditions which favor
"star activity" of other restriction endonuclease.

<>

<1>Xia, Y., Burbank, D.E., Van Etten, J.L.
<2>Restriction endonuclease activity induced by NC-1A virus infection of a Chlorella-like green alga.
<3>Nucleic Acids Res.
<4>14
<5>6017-6030
<6>1986
<7>A type II restriction endonuclease, CviBI, was isolated from a eukaryotic,
Chlorella-like green alga infected with the dsDNA containing virus NC-1A.  The
enzyme recognizes the sequence GANTC and cleaves DNA between the G and A.
Methylation of deoxyadenosine in the GANTC sequence probably inhibits enzyme
activity.  In vitro CviBI cleaves host nuclear DNA but not viral DNA.  A survey
of 18 other viruses which infect the same Chlorella sp. revealed that infection
with 5 of these viruses also induced a restriction endonuclease which cleaves
DNA into the same size fragments as CviBI.

<>

<1>Xia, Y., Morgan, R., Schildkraut, I., Van Etten, J.L.
<2>A site-specific single strand endonuclease activity induced by NYs-1 virus infection of a Chlorella-like green alga.
<3>Nucleic Acids Res.
<4>16
<5>9477-9487
<6>1988
<7>A site-specific endonuclease was isolated from a eukaryotic Chlorella-like
green alga infected with the dsDNA-containing virus NYs-1.  The enzyme
recognizes the sequence 5'-CC-3' and cleaves 5' to the first C.  It cleaves
5'-CmC-3' sequences but not 5'-mCC-3' sequences.  The enzyme creates breaks in
dsDNA whenever two 5'-CC-3' sequences on opposite strands are close enough for
the two strands to separate; when the 5'-CC-3' sequences on opposite strands
are further apart only a portion of the strands separate.  Consequently, NYs-1
endonuclease does not produce a completely stable DNA digestion pattern.  The
enzyme probably does not cleave ssDNA and definitely does not cleave ssRNA or
dsRNA.

<>

<1>Xia, Y., Narva, K.E., Van Etten, J.L.
<2>The cleavage site of the RsaI isoschizomer, CviII, is G^TAC.
<3>Nucleic Acids Res.
<4>15
<5>10063
<6>1987
<7>Infection of the green alga, Chlorella NC64A, with the dsDNA virus NY-2A
results in the synthesis of a restriction endonuclease CviII.  Chlorella cells
infected with NY-2A (m.o.i. of f10) were collected by centrifugation at 12 hr
p.i.  Note: this has been renamed CviQI.

<>

<1>Xia, Y., Van Etten, J.L.
<2>DNA methyltransferase induced by PBCV-1 virus infection of a chlorella-like green alga.
<3>Mol. Cell. Biol.
<4>6
<5>1440-1445
<6>1986
<7>A DNA methyltransferase was isolated from a eucaryotic, Chlorella-like green
alga infected with the virus PBCV-1.  The enzyme recognized the sequence GATC
and methylated deoxyadenosine solely in GATC sequences.  Host DNA, which
contains GATC sequences, but not PBCV-1 DNA, which contains GmATC sequences,
was a good substrate for the enzyme in vitro.  The DNA methyltransferase
activity was first detected about 1 h after viral infection; PBCV-1 DNA
synthesis and host DNA degradation also began at about this time.  The
appearance of the DNA methyltransferase activity required de novo protein
synthesis, and the enzyme was probably virus encoded.  Methylation of DNAs with
the PBCV-1-induced methyltransferase conferred resistance of the DNAs to a
PBCV-1-induced restriction endonuclease enzyme described previously (Y. Xia,
D.E. Burbank, L. Uher, D. Rabussay, and J.L. Van Etten, Mol. Cell Biol. 6:
1430-1439).  We propose that the PBCV-1-induced methyltransferase protects
viral DNA from the PBCV-1-induced restriction endonuclease and is part of a
virus-induced restriction and modification system in PBCV-1-infected Chlorella
cells.

<>

<1>Xia, Y., Van Etten, J.L., Dobos, P., Ling, Y.Y., Krell, P.J.
<2>Adenine DNA methyltransferase M.CviRI expression accelerates apoptosis in baculovirus-infected insect cells.
<3>Virology
<4>196
<5>817-824
<6>1993
<7>The adenine DNA methyltransferase M.CviRI (TGCmA) gene from chlorella virus XZ-6E was cloned
into the Autographa californica nuclear polyhedrosis virus (AcMNPV) genome and expressed in
Spodoptera frugiperda insect cells under the control of two tandemly arranged viral promoters,
the early ETL promoter and the late polyhedrin promoter. M.CviRI activity was first detected
at 10 hr p.i. and reached a maximum at 48 hr p.i. Viral DNA synthesized in insect cells
infected with M.CviRI expressing virus (AcMTRI) was methylated at all TGCA sites.
Unexpectedly, AcMTRI-infected cells lysed 48 hr earlier than wild-type AcMNPV-infected cells.
Moreover, cellular DNA, but not viral DNA, from AcM-TRI-infected cells was degraded to
fragment sizes characteristic of apoptosis. Thse results suggest that M.CviRI methylation
influences the onset of viral cytopathic effects and induces an apoptosis-like response.

<>

<1>Xia, Z.-G., Zhu, R.-F., Cao, X.-W., Zou, G.-L.
<2>Purification and characterization of restriction endonuclease Bsp78I.
<3>Acta Biochim. Biophys. Sin.
<4>19
<5>27-34
<6>1987
<7>Two kinds of direct single-step method using DNA-Sepharose and heparin-Sepharose affinity
chromatography to purify restriction endonuclease Bsp78I are reported. Chromatographic
conditions which may increase the purity and yield of the enzyme were studied. The purified
enzymes, obtained by the two methods, were found to be free of detectable contamination by
other DNases (exo and endo) caused by excess enzyme digestion of lambda DNA and by ligation
and recutting of enzymatic digests of pBR322 DNA. The specific activities and the yields of
the enzymes obtained by the two methods are both about 17,000 units per mg protein and 4,000
units per g wet bacteria, respectively. Some properties of the restriction endonuclease Bsp78I
are determined quantitatively. The optimal range of Mg++ concentration and optimal Tris-HCl
concentration are 20-30 m mol/L and 50 m mol/L respectively. The optimal pH is 7.5 and the
optimal temperature 40 C. 78I is very sensitive to PCMB.

<>

<1>Xiang, X.Y., Chen, L.M., Huang, X.X., Luo, Y.M., She, Q.X., Huang, L.
<2>Sulfolobus tengchongensis spindle-shaped virus STSV1: Virus-host interactions and genomic features.
<3>J. Virol.
<4>79
<5>8677-8686
<6>2005
<7>A virus infecting the hyperthermophilic archaeon Sulfolobus tengchongensis has been isolated
from a field sample from Tengchong,
China, and characterized. The virus, denoted STSV1 (Sulfolobus
tengchongensis spindle-shaped virus 1), has the morphology of a spindle
(230 by 107 nm) with a tail of variable length (68 nm on average) at
one end and is the largest of the known spindle-shaped viruses. After
infecting its host, the virus multiplied rapidly to high titers (>
10(10) PFU/ml). Replication of the virus retarded host growth but did
not cause lysis of the host cells. STSV1 did not integrate into the
host chromosome and existed in a carrier state. The STSV1 DNA was
modified in an unusual fashion, presumably by virally encoded
modification systems. STSV1 harbors a double-stranded DNA genome of
75,294 bp, which shares no significant sequence similarity to those of
fuselloviruses. The viral genome contains a total of 74 open reading
frames (ORFs), among which 14 have a putative function. Five ORFs
encode viral structural proteins, including a putative coat protein of
high abundance. The products of the other nine ORFs are probably
involved in polysaccharide biosynthesis, nucleotide metabolism, and DNA
modification. The viral genome divides into two nearly equal halves of
opposite gene orientation. This observation as well as a GC-skew
analysis point to the presence of a putative viral origin of
replication in the 1.4-kb intergenic region between ORF1 and ORF74.
Both morphological and genomic features identify STSV1 as a novel virus
infecting the genus Sulfolobus.

<>

<1>Xiao, J., Luo, Y., Xu, J.
<2>Genome Sequence of Serinicoccus profundi, a Novel Actinomycete Isolated from Deep-Sea Sediment.
<3>J. Bacteriol.
<4>193
<5>6413
<6>2011
<7>Serinicoccus profundi MCCC 1A05965(T) was isolated from deep-sea sediment collected from the
Indian Ocean. It was a Gram-positive, moderately
halophilic, aerobic bacterium. Here, we describe the 3.4-Mbp draft genome
sequence of S. profundi MCCC 1A05965(T).

<>

<1>Xiao, J.-P., Xu, S.-Y.
<2>Method for cloning the NspHI restriction-modification system in E. coli and producing the recombinant NspHI restriction endonuclease.
<3>US Patent Office
<4>US 6130078
<5>
<6>2000
<7>An isolated DNA coding for the NspHI restriction endonuclease and obtained from Nostoc spp.,
is claimed. Also claimed are: a recombinant
DNA vector into which a DNA segment encoding the NspHI restriction
endonuclease has been inserted; isolated DNA encoding the NspHI
restriction endonuclease and DNA-methylase obtained from ATCC 98989; a
cloning vector; a host cell transformed by the vector; and producing
NspHI restriction endonuclease comprising culturing a host cell
transformed with the vectors under conditions suitable for expression
of the endonuclease. The DNA encoding for the NspHI restriction
endonuclease and methylase are useful for creating recombinant
molecules in the laboratory and for producing large quantities of such
enzymes. The recombinant DNA is produced by methylase selection and DNA
sequencing of nspIM gene and the adjacent DNA.

<>

<1>Xie, B.B., Shu, Y.L., Qin, Q.L., Rong, J.C., Zhang, X.Y., Chen, X.L., Zhou, B.C., Zhang, Y.Z.
<2>Genome Sequence of the Cycloprodigiosin-Producing Bacterial Strain Pseudoalteromonas rubra ATCC 29570T.
<3>J. Bacteriol.
<4>194
<5>1637-1638
<6>2012
<7>The cycloprodigiosin biosynthetic gene cluster has not been reported. We sequenced the genome
of a cycloprodigiosin-producing bacterial strain,
Pseudoalteromonas rubra ATCC 29570(T). Analysis revealed a probable
cycloprodigiosin biosynthetic cluster, providing a good model for the study of
cycloprodigiosin synthesis and regulation.

<>

<1>Xie, F.
<2>Mechanistic studies of specific DNA cleavage by PvuII restriction endonuclease.
<3>Ph.D. Thesis, University of Missouri
<4>
<5>
<6>2008
<7>PvuII restriction endonuclease is a homodimeric protein which recognizes and cleaves the
palindromic sequence (CAGCTG) in the presence of Mg(II) ions. Starting with PvuII as a model
system, pKa calculations with crystallographically defined metal ligated water are applied to
PD…D/ExK motif metallonucleases in order to investigate the activation of nucleophile in metal
dependent DNA hydrolysis. These results establish the electrostatic contributions of the metal
ions and the conserved Lys in lowering water pKa. The calculated pKa values of metal ligands
have been used to simulate the pH dependence of Mg(II) binding to PvuII. The bell shaped
pH-rate profile is dissected into three ionizations. One is recognized as from the metal
ligands, and the other two have pKa's similar to calculated metal ligated
water pKa in the absence of DNA. The determined pH profiles agree well with
previous pH dependence studies on metallonucleases, and the correlation with pKa calculations
indicates the direct involvement of metal activated water in catalysis. iii The different
metal occupancies observed in crystal structures lead to controversy regarding the number and
function of metal ions involved in DNA hydrolysis by type II restriction endonucleases. Quench
flow experiments are used to monitor Mg(II) dependent single and multiple turnover DNA
cleavage reactions with PvuII. Several models which differ in order of binding and the number
of metal ions supporting catalysis are examined by global fits using DynaFit. The best fitted
model has a preference of binding order in the reaction scheme and supports one-metal ion
catalysis with 50 fold reduced activity compared with two-metal ion
catalysis. The same model is also found to account for multiple turnover data in fits and
simulations. A unique reaction scheme for PvuII is established to interpret the determined
Mg(II) dependence of kinetic data, which provides an insight into Mg(II) participation in
substrate binding, catalysis and product dissociation by restriction endonucleases.

<>

<1>Xie, G., Cui, Z., Tao, Z., Qiu, H., Liu, H., Ibrahim, M., Zhu, B., Jin, G., Sun, G., Almoneafy, A., Li, B.
<2>Genome Sequence of the Rice Pathogen Pseudomonas fuscovaginae CB98818.
<3>J. Bacteriol.
<4>194
<5>5479-5480
<6>2012
<7>Pseudomonas fuscovaginae is a phytopathogenic bacterium causing bacterial sheath  brown rot of
cereal crops. Here, we present the draft genome sequence of P.
fuscovaginae CB98818, originally isolated from a diseased rice plant in China.
The draft genome will aid in epidemiological studies, comparative genomics, and
quarantine of this broad-host-range pathogen.

<>

<1>Xie, G., Ramirez, M.S., Marshall, S.H., Hujer, K.M., Lo, C.C., Johnson, S., Li, P.E., Davenport, K., Endimiani, A., Bonomo, R.A., Tolmasky, M.E., Chain, P.S.
<2>Genome Sequences of Two Klebsiella pneumoniae Isolates from Different Geographical Regions, Argentina (Strain JHCK1) and the United States (Strain  VA360).
<3>Genome Announcements
<4>1
<5>e00168-13
<6>2013
<7>We report the sequences of two Klebsiella pneumoniae clinical isolates, strains JHCK1 and
VA360, from a newborn with meningitis in Buenos Aires, Argentina, and
from a tertiary care medical center in Cleveland, OH, respectively. Both isolates
contain one chromosome and at least five plasmids; isolate VA360 contains the
Klebsiella pneumoniae carbapenemase (KPC) gene.

<>

<1>Xie, G.L., Zhang, G.Q., Liu, H., Lou, M.M., Tian, W.X., Li, B., Zhou, X.P., Zhu, B., Jin, G.L.
<2>Genome sequence of the rice-pathogenic bacterium Acidovorax avenae subsp. avenae RS-1.
<3>J. Bacteriol.
<4>193
<5>5013-5014
<6>2011
<7>Acidovorax avenae subsp. avenae is a phytobacterium which is the causal agent of several
economic plant diseases. Here, we present the draft
genome sequence of strain RS-1, which was isolated from rice shoot in rice
field of China. This strain can cause bacterial stripe of rice.

<>

<1>Xie, J., Huang, J.F., Shi, X.F., Liu, C.Q.
<2>Analysis of the characteristic sequence of intein and revision of its motifs.
<3>Chinese Sci. Bull.
<4>46
<5>758-761
<6>2001
<7>Since the first intein (Sce VMA) was found in Saccharomyces cerevisiae ATPase gene in 1990,
more and more inteins were identified. It is necessary to analyze the new inteins to
understand the sequence characteristics of inteins. By searching protein and nucleic acid
database systematically, 101 inteins were found, of which 69 inteins contain homing
endonuclease moths. We only analyze the 69 inteins since most inteins are the classic inteins
with homing endonuclease motifs. We found that the distribution of these inteins is particular
among species and protein. By multiple sequence alignment, some new sequence characteristics
were found and the motifs described previously were revised.

<>

<1>Xie, J., Juliao, P.C., Gilsdorf, J.R., Ghosh, D., Patel, M., Marrs, C.F.
<2>Identification of New Genetic Regions More Prevalent in Nontypeable Haemophilus influenzae Otitis Media Strains than in Throat Strains.
<3>J. Clin. Microbiol.
<4>44
<5>4316-4325
<6>2006
<7>Nontypeable (NT) Haemophilus influenzae strains cause significant respiratory illness and are
isolated from up to half of middle ear aspirates from children with acute otitis media.
Previous studies have identified two genes, lic2B and hmwA, that are associated with NT H.
influenzae strains isolated from the middle ears of children with otitis media but that are
not associated with NT H. influenzae strains isolated from the throats of healthy children,
suggesting that they may play a role in virulence in otitis media. In this study, genomic
subtraction was used to identify additional genetic regions unique to middle ear strains. The
genome of NT H. influenzae middle ear strain G622 was subtracted from that of NT H. influenzae
throat strain 23221, and the resultant gene regions unique to the middle ear strain were
identified. Subsequently, the relative prevalence of the middle ear-specific gene regions
among a large panel of otitis media and throat strains was determined by dot blot
hybridization. By this approach, nine genetic regions were found to be significantly more
prevalent in otitis media strains. Classification tree analysis of lic2B, hmwA, and the nine
new potential otitis media virulence genes revealed two H. influenzae pathotypes associated
with otitis media.

<>

<1>Xie, J.B., Du, Z., Bai, L., Tian, C., Zhang, Y., Xie, J.Y., Wang, T., Liu, X., Chen, X., Cheng, Q., Chen, S., Li, J.
<2>Comparative Genomic Analysis of N2-Fixing and Non-N2-Fixing Paenibacillus spp.: Organization, Evolution and Expression of the Nitrogen Fixation Genes.
<3>PLoS Genet.
<4>10
<5>E1004231
<6>2014
<7>We provide here a comparative genome analysis of 31 strains within the genus
Paenibacillus including 11 new genomic sequences of N2-fixing strains. The
heterogeneity of the 31 genomes (15 N2-fixing and 16 non-N2-fixing Paenibacillus
strains) was reflected in the large size of the shell genome, which makes up
approximately 65.2% of the genes in pan genome. Large numbers of transposable
elements might be related to the heterogeneity. We discovered that a minimal and
compact nif cluster comprising nine genes nifB, nifH, nifD, nifK, nifE, nifN,
nifX, hesA and nifV encoding Mo-nitrogenase is conserved in the 15 N2-fixing
strains. The nif cluster is under control of a sigma(70)-depedent promoter and
possesses a GlnR/TnrA-binding site in the promoter. Suf system encoding [Fe-S]
cluster is highly conserved in N2-fixing and non-N2-fixing strains. Furthermore,
we demonstrate that the nif cluster enabled Escherichia coli JM109 to fix
nitrogen. Phylogeny of the concatenated NifHDK sequences indicates that
Paenibacillus and Frankia are sister groups. Phylogeny of the concatenated 275
single-copy core genes suggests that the ancestral Paenibacillus did not fix
nitrogen. The N2-fixing Paenibacillus strains were generated by acquiring the nif
cluster via horizontal gene transfer (HGT) from a source related to Frankia.
During the history of evolution, the nif cluster was lost, producing some
non-N2-fixing strains, and vnf encoding V-nitrogenase or anf encoding
Fe-nitrogenase was acquired, causing further diversification of some strains. In
addition, some N2-fixing strains have additional nif and nif-like genes which may
result from gene duplications. The evolution of nitrogen fixation in
Paenibacillus involves a mix of gain, loss, HGT and duplication of nif/anf/vnf
genes. This study not only reveals the organization and distribution of nitrogen
fixation genes in Paenibacillus, but also provides insight into the complex
evolutionary history of nitrogen fixation.

<>

<1>Xie, S., Wang, Z., Okano, M., Nogami, M., Li, Y., He, W.-W., Okumura, K., Li, E.
<2>Cloning, expression and chromosome locations of the human DNMT3 gene family.
<3>Gene
<4>236
<5>87-95
<6>1999
<7>DNA methylation plays an important role in animal development and gene regulation. In mammals,
several genes encoding DNA cytosine methyltransferases have been identified. DNMT1 is
constitutively expressed and is required for the maintenance of global methylation after DNA
replication. In contrast, the murine Dnmt3 family genes appear to be developmentally regulated
and behave like de novo DNA methyltransferases in vitro. In this study, we have cloned human
DNMT3A and DNMT3B that encode full-length DNMT3A and DNMT3B proteins with 98% and 94% amino
acid sequence identity to their murine homologues. The DNMT3A and DNMT3B show high homology in
the carboxy terminal catalytic domain and contain a conserved cysteine-rich region, which
shares homology with the X-linked ATRX gene of the SNF2/SWI family. We have mapped human
DNMT3A and DNMT3B to chromosomes 2p23 and 20q11.2 respectively, and determined the DNMT3B
genomic structure. We further show that DNMT3A expression is ubiquitous and can be readily
detected in most adult tissues, whereas DNMT3B is expressed at very low levels in most tissues
except testis, thyroid and bone marrow. Significantly, both DNMT3A and DNMT3B expression is
elevated in several tumor cell lines to levels comparable to DNMT1. The cloning of the human
DNMT3 genes will facilitate further biochemical and genetic studies of their functions in
establishment of DNA methylation patterns, regulation of gene expression and tumorigenesis.

<>

<1>Xie, X., Yu, Y., Liu, G., Yuan, Z., Song, J.
<2>Complexity and entropy analysis of DNA methyltransferase.
<3>J. Data Min. Genomics Proteomics
<4>1
<5>1000105
<6>2010
<7>The application of complexity information on DNA sequence and protein in biological processes
are well established in this study.  Available sequences for DNMT1 gene were thoroughly
explored in the informaion complexities.  DNMT1 gene is a maintenance methyltransferase
responsible for copy in DNA methylation patterns to the daughter strands during DNA
replication in different species.  We found that the entropy of DNMT1 gene in differnt species
is DNA base composition dependent, and its complexity in mammals is lower in introns than in
coding regions.  We also demonstrated the impacts of entropy on domains and non-domains(s) of
the DNMT1 gene.  The results from DNA and protein sequences indicated that DNA evolution has a
tendency toward complexity.  The most interest is that the methylation's changes of the gene
over aging in a unique chick model showed aging-driven entropy characteristics, which may give
an explanation of aging processes.  In summary, the information complexity of DNNT1 gene is
related to its genomic composition, which thereby associates to evolutionary and aging
processing, even though the intrinsic mechanism is not to be studied yet.

<>

<1>Xin, B., Tao, F., Wang, Y., Gao, C., Ma, C., Xu, P.
<2>Genome Sequence of Clostridium butyricum Strain DSM 10702, a Promising Producer of Biofuels and Biochemicals.
<3>Genome Announcements
<4>1
<5>e00563-13
<6>2013
<7>Clostridium butyricum strains have been considered promising producers of biofuels and
biochemicals, such as hydrogen, butanol, butyric acid, and
1,3-propanediol. Here, we present a 4.59-Mb assembly of the genome sequence of
DSM 10702 (VPI 3266), a type strain of C. butyricum.

<>

<1>Xin, D., El Karkouri, K., Robert, C., Raoult, D., Fournier, P.E.
<2>Genomic Comparison of Rickettsia honei Strain RBT and Other Rickettsia Species.
<3>J. Bacteriol.
<4>194
<5>4145
<6>2012
<7>Rickettsia honei strain RB(T) was isolated from a febrile patient on Flinders Island,
Australia, in 1991 and has been demonstrated to be the agent of Flinders
Island spotted fever, a disease transmitted to humans by ticks. The comparison of
this 1.27-Mb genome with other Rickettsia genomes provides additional insight
into the mechanisms of evolution in Rickettsia species.

<>

<1>Xiong, J., Li, D., Li, H., He, M., Miller, S.J., Yu, L., Rensing, C., Wang, G.
<2>Genome analysis and characterization of zinc efflux systems of a highly zinc-resistant bacterium, Comamonas testosteroni S44.
<3>Res. Microbiol.
<4>162
<5>671-679
<6>2011
<7>A novel and multiple metal(loid)-resistant strain Comamonas testosteroni S44 with
a high Zn(2+) resistance level (10 mM) was isolated. To understand the molecular
basis for the high zinc resistance, whole genome sequencing was performed and
revealed a large number of genes encoding putative metal(loid) resistance
proteins, mobile genetic elements (MGEs) and horizontal gene transfer (HGT)
events that may have occurred to adapt to a metal(loid)-contaminated environment.
In particular, 9 putative Zn(2+) transporters [4 znt operons encoding putative
Zn(2+)-translocating P-type ATPases and 5 czc operons encoding putative
RND-driven (resistance, nodulation, cell division protein family)] tripartite
protein complexes were identified. Real-time RT-PCR analysis revealed that the
four zntA-like genes were all induced by Zn(2+), while czcA genes were either
Zn(2+)-induced or downregulated by Zn(2+). Furthermore, a zntR1A1 operon encoding
a ZntR-type regulator and a P-type ATPase was studied in detail. The zntR1
deletion strain (S44DeltazntR1) displayed intermediate resistance to Zn(2+) (6
mM) and accumulated more intracellular Zn(2+). Reporter gene expression assays
indicated that ZntR1 responded to Zn(2+), Cd(2+) and Pb(2+), with Zn(2+) being
the best inducer. Gene transcription analysis indicated that ZntR1 was a
regulator for transcription of zntA1, while other putative ZntR-type regulators
may also regulate the transcription expression of zntA1.

<>

<1>Xiong, X.H., Han, S., Wang, J.H., Jiang, Z.H., Chen, W., Jia, N., Wei, H.L., Cheng, H., Yang, Y.X., Zhu, B., You, S., He, J.Y., Hou, W., Chen, M.X., Yu, C.J., Jiao, Y.H., Zhang, W.C.
<2>Complete genome sequence of the bacteria Ketogulonicigenium vulgare Y25.
<3>J. Bacteriol.
<4>193
<5>315-316
<6>2010
<7>Ketogulonicigenium vulgare is characterized by the efficient production of 2KGA from
L-sorbose. Ketogulonicigenium vulgare Y25 is known as a 2-keto-L-gulonic acid producing strain
in vitamin C industry. Here we report the finished, annotated genome sequence of
Ketogulonicigenium vulgare Y25.

<>

<1>Xiong, X.H., Zhi, J.J., Yang, L., Wang, J.H., Zhao, Y., Wang, X., Cui, Y.J., Dong, F., Li, M.X., Yang, Y.X., Wei, N., An, J.J., Du BH, L.L., Zhang, J.S., Zhou, W., Cheng, S.F., He, T., Wang, L., Chen, H.P., Liu, D.S., Zhang, W.C.
<2>Complete Genome Sequence of the Bacterium Methylovorus sp. Strain MP688, a High-Level Producer of Pyrroloquinolone Quinone.
<3>J. Bacteriol.
<4>193
<5>1012-1013
<6>2011
<7>Methylotrophic bacteria are widespread microbes which can use one carbon compound as their
only carbon and energy sources. Here we report the
finished, annotated genome sequence of the methylotrophic bacterium
Methylovorus sp. strain MP688, which was isolated from soil for high-level
production of pyrroloquinolone quinone (PQQ) in our lab.

<>

<1>Xiong, Y., Elckbush, T.H.
<2>Functional expression of a sequence-specific endonuclease encoded by the retrotransposon R2Bm.
<3>Cell
<4>55
<5>235-246
<6>1988
<7>A fraction of the 28S ribosomal genes in certain insect species is interrupted by the
insertion elements R1 and R2. These two elements from the silkworm Bombyx mori (R~1Bm and
R2Bm) are retrotransposons capable of transposing in a highly sequence-specific manner. We
report here the functional expression in E. coli of the entire single open reading frame of
R2Bm and show that it encodes a double-stranded endonuclease (integrase) that can specifically
cleave the 28S gene at the R2 insertion site. The resulting cleavage is a 4 bp staggered 5'
overhang. Deletion analysis of the 28S gene revealed that the DNA sequence required for
specific cleavage is asymmetric with respect to the actual insertion (cleavage) site, with
fewer than 10 bp required at one side and at least 24 bp at the other side of the site. A
model is proposed based on these and previous data to account for the sequence-specific
integration of the R2 retrotransposon.

<>

<1>Xiong, Y., Wang, P., Lan, R., Ye, C., Wang, H., Ren, J., Jing, H., Wang, Y., Zhou, Z., Bai, X., Cui, Z., Luo, X., Zhao, A., Wang, Y., Zhang, S., Sun, H., Wang, L., Xu, J.
<2>A Novel Escherichia coli O157:H7 Clone Causing a Major Hemolytic Uremic Syndrome Outbreak in China.
<3>PLoS ONE
<4>7
<5>E36144
<6>2012
<7>An Escherichia coli O157:H7 outbreak in China in 1999 caused 177 deaths due to
hemolytic uremic syndrome. Sixteen outbreak associated isolates were found to
belong to a new clone, sequence type 96 (ST96), based on multilocus sequence
typing of 15 housekeeping genes. Whole genome sequencing of an outbreak isolate,
Xuzhou21, showed that the isolate is phylogenetically closely related to the
Japan 1996 outbreak isolate Sakai, both of which share the most recent common
ancestor with the US outbreak isolate EDL933. The levels of IL-6 and IL-8 of
peripheral blood mononuclear cells induced by Xuzhou21 and Sakai were
significantly higher than that induced by EDL933. Xuzhou21 also induced a
significantly higher level of IL-8 than Sakai while both induced similar levels
of IL-6. The expression level of Shiga toxin 2 in Xuzhou21 induced by mitomycin C
was 68.6 times of that under non-inducing conditions, twice of that induced in
Sakai (32.7 times) and 15 times higher than that induced in EDL933 (4.5 times).
Our study shows that ST96 is a novel clone and provided significant new insights
into the evolution of virulence of E. coli O157:H7.

<>

<1>Xiong, Z., Jiang, Y., Qi, D., Lu, H., Yang, F., Yang, J., Chen, L., Sun, L., Xu, X., Xue, Y., Zhu, Y., Jin, Q.
<2>Complete genome sequence of the extremophilic Bacillus cereus strain Q1 with industrial applications.
<3>J. Bacteriol.
<4>191
<5>1120-1121
<6>2009
<7>Bacillus cereus strain Q1 was isolated from a deep-subsurface oil reservoir in the Daqing oil
field in northeastern China. This strain is
able to produce biosurfactants and to survive in extreme environments.
Here we report the finished and annotated genome sequence of this
organism.

<>

<1>Xiong, Z., Laird, P.W.
<2>COBRA: a sensitive and quantitative DNA methylation assay.
<3>Nucleic Acids Res.
<4>25
<5>2532-2534
<6>1997
<7>We report here on a quantitative technique called COBRA to determine DNA methylation levels at
specific gene loci in small amounts of genomic DNA.  Restriction enzyme digestion is used to
reveal methylation-dependent sequence differences in PCR products of sodium bisulfite-treated
DNA as described previously.  We show that methylation levels in the original DNA sample are
represented by the relative amounts of digested and undigested PCR product in a linearly
quantitative fashion across a wide spectrum of DNA methylation levels.  In addition, we show
that this technique can be reliably applied to DNA obtained from microdissected
paraffin-embedded tissue samples.  COBRA thus combines the powerful features of ease of use,
quantitative accuracy, and compatibility with paraffin sections.

<>

<1>Xiong, Z.Q., Wang, Y.
<2>Draft Genome Sequence of the Marine Streptomyces sp. Strain AA1529, Isolated from the Yellow Sea.
<3>J. Bacteriol.
<4>194
<5>5474-5475
<6>2012
<7>Here we report the draft genome sequence of a Streptomyces strain, AA1529, isolated from
marine sediment from the Yellow Sea. Its genome contains a subset
of unique genes and gene clusters that encode diverse secondary metabolites,
suggesting great potential as a source for the discovery of novel gene clusters
and bioactive compounds.

<>

<1>Xiong, Z.Q., Wang, Y.
<2>Draft Genome Sequence of Marine-Derived Streptomyces sp. Strain AA0539, Isolated  from the Yellow Sea, China.
<3>J. Bacteriol.
<4>194
<5>6622-6623
<6>2012
<7>Here, we report the draft genome sequence of Streptomyces sp. strain AA0539, isolated from
marine sediment of the Yellow Sea, China. Its small genome (
approximately 5.8 Mb) contains large, unique genes and gene clusters for diverse
secondary metabolites, suggesting great potential as a source for the discovery
of novel natural products.

<>

<1>Xu, C., Song, J., Ding, Y., Yu, F., Sun, L., Tang, L., Hu, X., Zhang, Z., He, J.
<2>Crystallization and preliminary X-ray analysis of Sau3AI/E64A mutant protein.
<3>Protein Pept. Lett.
<4>14
<5>505-506
<6>2007
<7>Sau3AI is a type II restriction endonuclease that recognizes the palindromic sequence
5'-GATC-3' and cleaves 5' to G residue on each
strand. The E64A mutant full length protein was cloned and expressed in
Escherichia coli. The purified (His) (6)-tagged protein has monomer and
dimer fraction and was crystallized by the hanging-drop vapor-diffusion
technique. The dimer protein crystals can diffract to 3.0A. resolution and
the monomer protein crystals can diffract to better than 2.8A. resolution.
One completed dataset has been collected and it shows that the monomer
orthorhombic Sau3AI/E64A crystal is in space group C2221 with unit cell
parameters (69.44, 197.60, 191.46, 90, 90, 90) and contains two molecules
in one asymmetric unit.

<>

<1>Xu, D., Cisar, J.O., Poly, F., Yang, J., Albanese, J., Dharmasena, M., Wai, T., Guerry, P., Kopecko, D.J.
<2>Genome Sequence of Salmonella enterica Serovar Typhi Oral Vaccine Strain Ty21a.
<3>Genome Announcements
<4>1
<5>e00650-13
<6>2013
<7>Attenuated Salmonella enterica serovar Typhi strain Ty21a is an important vaccine for
controlling typhoid fever and serves as an oral vector for delivering
heterologous antigens. The key attenuating features of this randomly mutated
strain remain in question. Genome sequencing has revealed 679 single nucleotide
polymorphisms (SNPs), and will help define alterations contributing to Ty21a
safety and immunogenicity.

<>

<1>Xu, F., Mao, C., Ding, Y., Rui, C., Wu, L., Shi, A., Zhang, H., Zhang, L., Xu, Z.
<2>Molecular and Enzymatic Profiles of Mammalian DNA Methyltransferases: Structures and Targets for Drugs.
<3>Curr. Med. Chem.
<4>17
<5>4052-4071
<6>2010
<7>DNA methylation is an epigenetic event involved in a variety array of processes that may be
the foundation of genetic phenomena and diseases.
DNA methyltransferase is a key enzyme for cytosine methylation in DNA,
and can be divided into two functional families (Dnmt1 and Dnmt3) in
mammals. All mammalian DNA methyltransferases are encoded by their own
single gene, and consisted of catalytic and regulatory regions (except
Dnmt2). Via interactions between functional domains in the regulatory
or catalytic regions and other adaptors or cofactors, DNA
methyltransferases can be localized at selective areas (specific
DNA/nucleotide sequence) and linked to specific chromosome status
(euchromatin/ heterochromatin, various histone modification status).
With assistance from UHRF1 and Dnmt3L or other factors in Dnmt1 and
Dnmt3a/Dnmt3b, mammalian DNA methyltransferases can be recruited, and
then specifically bind to hemimethylated and unmethylated
double-stranded DNA sequence to maintain and de novo setup patterns for
DNA methylation. Complicated enzymatic steps catalyzed by DNA
methyltransferases include methyl group transferred from cofactor
Ado-Met to C5 position of the flipped-out cytosine in targeted DNA
duplex. In the light of the fact that different DNA methyltransferases
are divergent in both structures and functions, and use unique
reprogrammed or distorted routines in development of diseases, design
of new drugs targeting specific mammalian DNA methyltransferases or
their adaptors in the control of key steps in either maintenance or de
novo DNA methylation processes will contribute to individually treating
diseases related to DNA methyltransferases.

<>

<1>Xu, F., Miao, D., Du Y, C.X., Zhang, P., Sun, H.
<2>Draft Genome Sequence of Avibacterium paragallinarum Strain 221.
<3>Genome Announcements
<4>1
<5>e00290-13
<6>2013
<7>Avibacterium paragallinarum is the causative agent of infectious coryza. Here we  report the
draft genome sequence of reference strain 221 of A. paragallinarum serovar A. The genome is
composed of 135 contigs for 2,685,568 bp with a 41% G+C content.

<>

<1>Xu, G., Davis, M., Glickman, F., Reich, N.
<2>Purification and initial characterization of the murine cytosine DNA methyltransferase.
<3>FASEB J.
<4>7
<5>A1175
<6>1993
<7>The murine DNA (cytosine-5-)-methyltransferase (MTase EC 2.1.1.37) transfers methyl groups
from S-adenosylmethionine to the 5-position of cytosine in the nucleotide pair dCpdG of DNA.
The methylation of cytosine residues on mammalian DNA has been implicated in the regulation of
gene expression. Our goal is to study the chemical mechanism of cytosine methylation and to
characterize the basis of the enzyme specificity. DNA MTase was purified to homogeneity from
nuclear extracts of Friend murine erythroleukemia cells (MEL cells). Chromatographic
purification of DNA MTase carried out on DEAE-Sepharose Fast Flow medium brought about a
6-fold increase in the specific activity of DNA MTase. Further purification of the enzyme was
achieved by precipitation by 30% saturated ammonium sulfate solution and chromatography on
Phenyl-Sepharose 6 Fast Flow medium. The purification step brought a 21-fold increase in
specific activity of the enzyme. After chromatographic separation of DNA MTase on
phenylsepharose, purification using a Hiload superdex 200 column provided DNA MTase with
80-fold higher specific activity. We are currently characterizing the enzyme-substrate
interaction.

<>

<1>Xu, G., Flynn, J., Glickman, J.F., Reich, N.O.
<2>Purification and stabilization of mouse DNA methyltransferase.
<3>Biochem. Biophys. Res. Commun.
<4>207
<5>544-551
<6>1995
<7>Cytosine methylation within DNA has been implicated in genetic imprinting, X-chromosome
inactivation, regulation of tissue-specific gene expression, aging, and cancer. Unfortunately,
DNA (cytosine-5)-methyltransferases (EC 2.1.1.37) from various mammalian sources have been
difficult to isolate and stabilize, precluding investigations of these critical enzymes. We
describe a novel FPLC purification of the 190,000 Mr DNA methyltransferase from mouse Friend
erythroleukemia cells. The homogeneous 190 kD Mr form of the enzyme is the only polypeptide
detected at various stages of cell growth and has not undergone detectable N-terminal
proteolysis.

<>

<1>Xu, G., Hu, J., Fang, X., Zhang, X., Wang, J., Guo, Y., Li, T., Chen, Z., Dai, W., Liu, C.
<2>Genome Sequence of Pseudomonas aeruginosa Strain LCT-PA220, Which Was Selected after Space Flight by Using Biolog's Powerful Carbon Source Utilization  Technology.
<3>Genome Announcements
<4>2
<5>e00169-14
<6>2014
<7>To explore the changes of Pseudomonas aeruginosa in space flight, we present the  draft genome
sequence of P. aeruginosa strain LCT-PA220, which originated from a
P. aeruginosa strain, ATCC 27853, that traveled on the Shenzhou-VIII spacecraft.

<>

<1>Xu, G., Willert, J., Kapfer, W., Trautner, T.A.
<2>BsuCI, a type-I restriction-modification system in Bacillus subtilis.
<3>Gene
<4>157
<5>59
<6>1995
<7>A type-I R-M system was identified in B. subtilis.  The genes comprising the system have
striking similarity to type-I R-M systems observed in Enterobacteriaceae.

<>

<1>Xu, G.-L., Bestor, T.H.
<2>Cytosine methylation targetted to pre-determined sequences.
<3>Nat. Genet.
<4>17
<5>376-378
<6>1997
<7>The incidence or severity of a number of human diseases is proportional to the activity of
individual viral transcription units or mutant endogenous genes.  Targetted methylation of
regulatory sites is proposed as a new method for selective gene inactivation that stimulates
an existing biological response.  Mammalian promoters are usually inactive when methylated at
CpG dinucleotides, and patterns of methylated CpG sites are replicated with the DNA during S
phase.  Pre-determined sequence specificities have now been conferred upon a DNA
methyltransferase by fusion to zinc-finger proteins.  The sequence specificity of zinc-finger
proteins can be modified to direct cytosine methylation to the promoters of target genes.

<>

<1>Xu, G.-L., Bestor, T.H., Bourchis, D., Hsieh, C.-L., Tommerup, N., Bugge, M., Hulten, M., Qu, X., Russo, J.J., Viegas-Pequignot, E.
<2>Chromosome instability and imunodeficiency syndrome caused by mutations in a DNA methyltransferase gene.
<3>Nature
<4>402
<5>187-191
<6>1999
<7>The recessive autosomal disorder known as ICF syndrome (for immunodeficiency, centromere
instability and facial anomalies; Mendelian Inheritance in Man number 242860) is characterized
by variable reductions in serum immunoglobulin levels which cause most ICF patients to succumb
to infectious diseases before adulthood.  Mild facial anomalies include hypertelorism, low-set
ears, epicanthal folds and macroglossia.  The cytogenetic abnormalities in lymphocytes are
exuberant: juxtacentromeric heterochromatin is greatly elongated and thread-like in metaphase
chromsomes, which is associated with the formation of complex multiradiate chromosomes.  The
same juxtacentromeric regions are subject to persistent interphase self-associations and are
extruded into nuclear blebs or micronuclei.  Abnormalities are largely confined to tracts of
classical satellites 2 and 3 at juxtacentromeric regions of chromosomes 1, 9 and 16.
Classical satellite DNA is normally heavily methylated at cytosine residues, but in ICF
syndrome it is almost completely unmethylated in all tissues.  ICF syndrome is the only
genetic disorder known to involve constitutive abnormalities of genomic methylation patterns.
Here we show that five unrelated ICF patients have mutations in both alleles of the gene that
encodes DNA methyltransferase 3B.  Cytosine methylation is essential for the organization and
stabilization of a specific type of heterochromatin, and this methylation appears to be
carried out by an enzyme specialized for the purpose.

<>

<1>Xu, G.L., Kapfer, W., Walter, J., Trautner, T.A.
<2>BsuBI-an isospecific restriction and modification system of PstI: characterization of the BsuBI genes and enzymes.
<3>Nucleic Acids Res.
<4>20
<5>6517-6523
<6>1992
<7>The enzymes of the Bacillus subtilis BsuBI restriction/modification (R/M) system recognize the
target sequence 5'CTGCAG. The genes of the BsuBI R/M system have been cloned and sequenced
and their products have been characterized following overexpression and purification. The gene
of the BsuBI DNA methyltransferase (M.BsuBI) consists of 1503 bp, encoding a protein of 501
amino acids with a calculated Mr of 57.2 kD. The gene of the restriction endonuclease
(R.BsuBI), comprising 948 bp, codes for a protein of 316 amino acids with a predicted Mr of
36.2 kD. M.BsuBI modifies the adenine (A) residue of the BsuBI target site, thus representing
the first A-N6_DNA methyltransferase identified in B. subtilis. Like R.PstI, R.BsuBI cleaves
between the A residue and the 3' terminal G of the target site. Both enzymes of the BsuBI R/M
system are, therefore, functionally identical with those of the PstI R/M system, encoded by
the Gram negative species Providencia stuartii. This functional equivalence coincides with a
pronounced similarity of the BsuBI/PstI DNA methyltransferases (41% amino acid identity) and
restriction endonucleases (46% amino acid identity). Since the genes are also very similar
(58% nucleotide identity), the BsuBI and PstI R/M systems apparently have a common
evolutionary origin. In spite of the sequence conservation the gene organization is strikingly
different in the two R/M systems. While the genes of the PstI R/M system are separated and
transcribed divergently, the genes of the BsuBI R/M system are transcribed in the same
direction with the 3' end of the M gene overlapping the 5' end of the R gene by 17 bp.

<>

<1>Xu, H., Han, D., Xu, Z.
<2>Overexpression of a lethal methylase, M.TneDI, in E-coli B1(DE3).
<3>Biotechnol. Lett.
<4>36
<5>1853-1859
<6>2014
<7>A pET-based vector pDH21 expressing the methylase, M.TneDI (recognizing CGCG) from Thermotoga
was constructed, and transformed into E. coli B1(DE3). Despite E. coli B1(DE3) being McrBC
positive, 30 transformants were isolated, which were suspected to be McrBC(-) mutants. The
overexpression of M.TneDI was verified by SDS-PAGE analysis. Compared to the previously
constructed pJC340 vector, a pACYC184 derivative expressing M.TneDI from a tet promotor, the
newly constructed pDH21 vector improved the expression of the methylase about fourfold,
allowing complete protection of DNA substrates. This study not only demonstrates a practical
approach to overexpressing potential lethal proteins in E. coli but also delivers a production
strain of M.TneDI that may be useful in various in vitro methylation applications.

<>

<1>Xu, J. et al.
<2>Evolution of Symbiotic Bacteria in the Distal Human Intestine.
<3>PLoS Biology
<4>5
<5>e156
<6>2007
<7>The adult human intestine contains trillions of bacteria, representing hundreds of species and
thousands of subspecies. Little is known about the
selective pressures that have shaped and are shaping this community's
component species, which are dominated by members of the Bacteroidetes and
Firmicutes divisions. To examine how the intestinal environment affects
microbial genome evolution, we have sequenced the genomes of two members
of the normal distal human gut microbiota, Bacteroides vulgatus and
Bacteroides distasonis, and by comparison with the few other sequenced gut
and non-gut Bacteroidetes, analyzed their niche and habitat adaptations.
The results show that lateral gene transfer, mobile elements, and gene
amplification have played important roles in affecting the ability of
gut-dwelling Bacteroidetes to vary their cell surface, sense their
environment, and harvest nutrient resources present in the distal
intestine. Our findings show that these processes have been a driving
force in the adaptation of Bacteroidetes to the distal gut environment,
and emphasize the importance of considering the evolution of humans from
an additional perspective, namely the evolution of our microbiomes.

<>

<1>Xu, J., Bjursell, M.K., Himrod, J., Deng, S., Carmichael, L.K., Chiang, H.C., Hooper, L.V., Gordon, J.I.
<2>A genomic view of the human-Bacteroides thetaiotaomicron symbiosis.
<3>Science
<4>299
<5>2074-2076
<6>2003
<7>The human gut is colonized with a vast community of indigenous microorganisms that help shape
our biology. Here, we present the complete
genome sequence of the Gram-negative anaerobe Bacteroides
thetaiotaomicron, a dominant member of our normal distal intestinal
microbiota. Its 4779-member proteome includes an elaborate apparatus for
acquiring and hydrolyzing otherwise indigestible dietary polysaccharides
and an associated environment-sensing system consisting of a large
repertoire of extracytoplasmic function sigma factors and one- and
two-component signal transduction systems. These and other expanded
paralogous groups shed light on the molecular mechanisms underlying
symbiotic host-bacterial relationships in our intestine.

<>

<1>Xu, J., Zheng, H.J., Liu, L., Pan, Z.C., Prior, P., Tang, B., Xu, J.S., Zhang, H., Tian, Q., Zhang, L.Q., Feng, J.
<2>Complete Genome Sequence of the Plant Pathogen Ralstonia solanacearum Strain Po82.
<3>J. Bacteriol.
<4>193
<5>4261-4262
<6>2011
<7>Ralstonia solanacearum strain Po82, a phylotype IIB/sequevar 4 strain, was found to be
pathogenic to both solanaceous plants and banana. Here, we
report the complete genome sequence of Po82 and its comparison with seven
published R. solanacearum genomes.

<>

<1>Xu, K., Su, F., Tao, F., Li, C., Ni, J., Xu, P.
<2>Genome Sequences of Two Morphologically Distinct and Thermophilic Bacillus coagulans Strains, H-1 and XZL9.
<3>Genome Announcements
<4>1
<5>e00254-13
<6>2013
<7>Two thermophilic Bacillus coagulans strains, H-1 and XZL9, both of which were isolated from
soils, have different morphological properties. Strain XZL9 but not
H-1 is an efficient pentose-utilizing producer of important platform compounds,
such as l-lactic acid and 2,3-butanediol. Here we announce the 2.86- and 3.43-Mb
sequences of their genomes.

<>

<1>Xu, M., Fang, Y., Liu, J., Chen, X., Sun, G., Guo, J., Hua, Z., Tu, Q., Wu, L., Zhou, J., Liu, X.
<2>Draft Genome Sequence of Shewanella decolorationis S12, a Dye-Degrading Bacterium Isolated from a Wastewater Treatment Plant.
<3>Genome Announcements
<4>1
<5>e00993-13
<6>2013
<7>Shewanella decolorationis is a valuable microorganism for degrading diverse synthetic textile
dyes. Here, we present an annotated draft genome sequence of S.
decolorationis S12, which contains 4,219 protein-coding genes and 86 structural
RNAs. This information regarding the genetic basis of this bacterium can greatly
advance our understanding of the physiology of this species.

<>

<1>Xu, M., Han, Q., Tan, Y.
<2>Modulators of methylation for control of bacterial virulence.
<3>International Patent Office
<4>WO 0074686 A
<5>
<6>2000
<7>Compositions and methods which ameliorate the virulence of bacterial infection are described
wherein the active ingredient modulates transmethylation reactions in bacterial cells.
Particularly useful compounds are inhibitors of S-adenosyl methionine synthetase (SAMS), of
S-adenosyl homocysteine hydrolase (SAHH) and of transmethylases.

<>

<1>Xu, M., Kladde, M.P., Van Etten, J.L., Simpson, R.T.
<2>Cloning, characterization and expression of the gene coding for a cytosine-5-DNA methyltransferase recognizing GpC.
<3>Nucleic Acids Res.
<4>26
<5>3961-3966
<6>1998
<7>A novel gene encoding a cytosine-5-DNA methyltransferase recognizing the dinucleotide GpC was
cloned from Chlorella virus NYs-1 and expressed in both Escherichia coli and Saccharomyces
cerevisiae.  The gene was sequenced and a predicted polypeptide of 362 amino acids with a
molecular weight of 41.903 kDa was identified.  The protein contains several amino acid motifs
with high similarity to those of other known 5-methylcytosine-forming methyltransferases.  In
addition, this enzyme, named M.CviPI, shares 66% identity and 76% similarity with M.CviJI, the
other only cytosine-5-DNA methyltransferase cloned from a Chlorella virus.  The short,
frequently occurring recognition sequence of the new methyltransferase will be very useful for
in vivo chromatin structure studies in both yeast and higher organisms.

<>

<1>Xu, M.-Q., Comb, D.G., Paulus, H., Noren, C.J., Shao, Y., Perler, F.B.
<2>Protein splicing: an analysis of the branched intermediate and its resolution by succinimide formation.
<3>EMBO J.
<4>13
<5>5517-5522
<6>1994
<7>Protein splicing involves the excision of an internal domain from a
precursor protein and the ligation of the external domains so as to generate two new
proteins.  Study of this process has recently been facilitated by the isolation of a precursor
and a branched intermediate from a thermophilic protein splicing element expressed in a
foreign protein context.  Two aspects of protein splicing are examined in this paper.  We
demonstrate a succinimide at the C-terminus of the spliced internal protein, implicating
cyclization of asparagine in resolution of the branched intermediate, and we identify an
alkali-labile bond in the branched intermediate.  A revised protein splicing model based on
these experimental results is presented.

<>

<1>Xu, M.-Q., Evans, T.C.
<2>Intein mediated peptide ligation.
<3>International Patent Office
<4>WO 0018881
<5>
<6>2000
<7>In accordance with the present invention, there is provided a method for producing a
semi-synthetic fusion protein in vitro, comprising the steps of producing a target protein
fused to a protein splicing element (an intein) and selectively cleaving the fusion and
ligating a synthetic protein or peptide at the C-terminal thioester of the target protein,
which overcome many of the disadvantages and problems noted above.  Specifically, the present
invention has higher yields due to better thiol-induced cleavage with thiol-reagents which
have been optimized for the ligation reaction, off-column ligation allows sample concentration
and allows the use of less peptide, MESNA is an odorless thiol-reagent for ligation, and Mxe
intein is from a bacterial source and often expresses better in bacterial cells.  Furthermore,
the present invention allows peptides to be directly ligated to the thioester bond formed
between an intein and the target protein.  The present invention also provides a method for
producing a cytotoxic protein, comprising the steps of producing a truncated, inactive form of
the protein in vivo which is fused to a protein splicing element, and selectively cleaving the
fusion and ligating a synthetic protein or peptide at a C-terminal thioester of the target
protein to restore the activity of the native cytotoxic protein.  Recombinant vectors for
producing such cleavable fusion proteins are also provided.

<>

<1>Xu, M.-Q., Perler, F.B.
<2>The mechanism of protein splicing and its modulation by mutation.
<3>EMBO J.
<4>15
<5>5146-5153
<6>1996
<7>Protein splicing results in the expression of two mature proteins from a single gene.  After
synthesis of a precursor protein, an internal segment (the intein) is excised and the external
domains are joined together.  A self-catalyzed mechanism for this cleavage-ligation reaction
is presented, based on mutagenesis data and analysis of splicing intermediates.  Mutations
were used to block various steps in the protein splicing pathway, allowing each isolated step
to be studied independently.  A linear ester intermediate was identified and functional roles
for the four conserved splice junction residues were determined.  Understanding the mechanism
of protein splicing provides a basis for protein engineering studies.  For example, inteins
can be constructed which fail to splice, but instead cleave the peptide bond at a chosen
splice junction.

<>

<1>Xu, M.-Q., Shub, D.A.
<2>The catalytic core of the sunY intron of bacteriophage T4.
<3>Gene
<4>82
<5>77-82
<6>1989
<7>The self-splicing sunY intron of bacteriophage T4 shares a common
secondary structure with
other group I introns.  A long open reading frame within the intron is entirely 3' of the
structural elements
conserved in all group I introns.  This catalytic core is the smallest yet described for a
self-
splicing intron.
An internal deletion of 728 nucleotides, leaving 196 nt at the 5' end and 109 nt at the 3'
end, allows normal
self-splicing.  Transcripts terminating 196 nt 3' of the 5' splice site retain catalytic
activity.

<>

<1>Xu, M.-Q., Southworth, M.W., Mersha, F.B., Hornstra, L.J., Perler, F.B.
<2>In vitro protein splicing of purified precursor and the identification of a branched intermediate.
<3>Cell
<4>75
<5>1371-1377
<6>1993
<7>Protein splicing is a posttranslational processing event in which an internal polypeptide is
excised from a protein precursor and the terminal polypeptides are then ligated together,
resulting in the production of two proteins. This report presents direct evidence for protein
splicing by demonstrating in vitro splicing of purified precursor that accumulated when the
protein splicing element from Pyrococcus DNA polymerase was cloned into a foreign gene. In
vitro splicing was temperature and pH dependent. A slowly migrating species exhibited kinetic
properties of a splicing intermediate and was shown to be a branched molecular by N-terminal
sequencing. The precursor and slowly migrating species were interconvertible in response to pH
shifts.

<>

<1>Xu, P., Alves, J.M., Kitten, T., Brown, A., Chen, Z., Ozaki, L.S., Manque, P., Ge, X., Serrano, M.G., Puiu, D., Hendricks, S., Wang, Y., Chaplin, M.D., Akan, D., Paik, S., Peterson, D.L., Macrina, F.L., Buck, G.A.
<2>Genome of the opportunistic pathogen Streptococcus sanguinis.
<3>J. Bacteriol.
<4>189
<5>3166-3175
<6>2007
<7>The genome of S. sanguinis is a circular DNA molecule of 2,388,435 base pairs, 177-590 kb
larger than the other 21 sequenced streptococcal genomes. The GC content of the S. sanguinis
genome is 43.4%, considerably higher than that of other streptococci. The genome encodes 2,274
predicted proteins, 61 tRNAs and 4 ribosomal RNA operons. A 70-kb region containing pathways
for vitamin B12 biosynthesis and degradation of ethanolamine and propanediol was apparently
acquired by horizontal gene transfer. The gene complement suggests new hypotheses for the
pathogenesis and virulence of S. sanguinis, and provides comparative contrasts with other
pathogenic and non-pathogenic streptococci. In particular, S. sanguinis possesses a remarkable
abundance of putative surface proteins, which may permit it to serve as a primary colonizer of
the oral cavity and agent of streptococcal endocarditis and infection in neutropenic patients.

<>

<1>Xu, P., Widmer, G., Wang, Y., Ozaki, L.S., Alves, J.M., Serrano, M.G., Puiu, D., Manque, P., Akiyoshi, D., Mackey, A.J., Pearson, W.R., Dear, P.H., Bankier, A.T., Peterson, D.L., Abrahamsen, M.S., Kapur, V., Tzipori, S., Buck, G.A.
<2>The genome of Cryptosporidium hominis.
<3>Nature
<4>431
<5>1107-1112
<6>2004
<7>Cryptosporidium species cause acute gastroenteritis and diarrhoea worldwide. They are members
of the Apicomplexa--protozoan pathogens that invade host cells by using a specialized apical
complex and are usually transmitted by an invertebrate vector or intermediate host. In
contrast to other Apicomplexans, Cryptosporidium is transmitted by ingestion of oocysts and
completes its life cycle in a single host. No therapy is available, and control focuses on
eliminating oocysts in water supplies. Two species, C. hominis and C. parvum, which differ in
host range, genotype and pathogenicity, are most relevant to humans. C. hominis is restricted
to humans, whereas C. parvum also infects other mammals. Here we describe the eight-chromosome
approximately 9.2-million-base genome of C. hominis. The complement of C. hominis
protein-coding genes shows a striking concordance with the requirements imposed by the
environmental niches the parasite inhabits. Energy metabolism is largely from glycolysis. Both
aerobic and anaerobic metabolisms are available, the former requiring an alternative electron
transport system in a simplified mitochondrion. Biosynthesis capabilities are limited,
explaining an extensive array of transporters. Evidence of an apicoplast is absent, but genes
associated with apical complex organelles are present. C. hominis and C. parvum exhibit very
similar gene complements, and phenotypic differences between these parasites must be due to
subtle sequence divergence.

<>

<1>Xu, P., Yu, H., Chakrabarty, A.M., Xun, L.
<2>Genome Sequence of the 2,4,5-Trichlorophenoxyacetate-Degrading Bacterium Burkholderia phenoliruptrix Strain AC1100.
<3>Genome Announcements
<4>1
<5>e00600-13
<6>2013
<7>Burkholderia phenoliruptrix strain AC1100 (ATCC 53867) degrades a variety of recalcitrant
xenobiotics, including 2,4,5-trichlorophenoxyacetate. The molecular
mechanism of 2,4,5-trichlorophenoxyacetate degradation has been extensively
studied. Here we present a 7.8-Mb assembly of the genome sequence of this
2,4,5-trichlorophenoxyacetate-degrading strain, which may provide useful
information related to the degradation of chlorinated aromatic compounds.

<>

<1>Xu, Q.
<2>Molecular analysis of restriction-modification systems in Helicobacter pylori.
<3>Ph.D. Thesis, Vanderbilt University, Nashville, Tennessee, , , Nashville
<4>
<5>1-156
<6>1999
<7>The phenomenon of restriction and modification in prokaryotes was first recognized more than
40 years ago when investigators worked on phage infections.  Certain bacterial strains of
bacteria were found to inhibit the propagation of phages grown previously on a different
bacterial strain.  The effect was later traced to a DNA sequence-specific endonuclease.  The
DNA of phages previously grown on the different strain was degraded by this certain strain's
endonuclease, because the phage DNA lacked specific methylation in the sequence recognized by
the endonuclease.  However, the few phages that did succeed in infecting the bacteria, were
modified so that they grew normally on the new host in a second cycle of infection.  By the
beginning of the 1970's, first several restriction endonucleases including HindII, HindIII,
and EcoRI, were purified.  Sequence-specific endonucleases were later found to be useful for
analyzing and rearranging DNA.  Subsequent applications not only led to a major breakthrough
in the field of molecular biology, but also led to the extensive search for new ones.

<>

<1>Xu, Q., Blaser, M.J.
<2>Promoters of the CATG-specific methyltransferase gene hpyIM differ between iceA1 and iceA2 Helicobacter pylori strains.
<3>J. Bacteriol.
<4>183
<5>3875-3884
<6>2001
<7>Helicobacter pylori strains can be divided into two groups, based on the presence of two
unrelated genes, iceA1 and iceA2, that occupy the same genomic locus. hpyIM, located
immediately downstream of either gene, encodes a functional CATG-specific methyltransferase.
Despite the strong conservation of the hpyIM open reading frame (ORF) among all H. pylori
strains, the sequences upstream of the ORF in iceA1 and iceA2 strains are substantially
different. To explore the roles of these upstream sequences in hpyIM regulation, promoter
analysis of hpyIM was performed. Both deletion mutation and primer extension analyses
demonstrate that the hpyIM promoters differ between H. pylori strains 60190 (iceA1) and J188
(iceA2). In strain 60190, hpyIM has two promoters, P(a) or P(I), which may function
independently, whereas only one hpyIM promoter, P(c), was found in strain J188. The XylE assay
showed that the hpyIM transcription level was much higher in strain 60190 than in strain J188,
indicating that regulation of hpyIM transcription differs between the H. pylori iceA1 strain
(60190) and iceA2 strains (J188). Since the iceA1 and iceA2 sequences are highly conserved
within iceA1 or iceA2 strains, we conclude that promoters of the CATG-specific methylase gene
hpyIM differ between iceA1 and iceA2 strains, which leads to differences in regulation of
hpyIM transcription.

<>

<1>Xu, Q., Morgan, R.D., Roberts, R.J., Blaser, M.J.
<2>Identification of type II restriction and modification systems in Helicobacter pylori reveals their substantial diversity among strains.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>97
<5>9671-9676
<6>2000
<7>A total of 22 type II restriction endonucleases with 18 distinct specificities have been
identified in six Helicobacter pylori strains. Among these 18 specificities are three
completely new endonucleases, Hpy178III, Hpy99I, and Hpy188I, that specifically cleave DNA at
TCNNGA, CGWCG, and TCNGA sites, respectively. The set of endonucleases identified in each
strain varies, but all have four- or five-base recognition sequences. Among 16 H. pylori
strains, examination of the DNA modification status at the recognition sites of 15 restriction
endonucleases reveals that each strain has a substantially different complement of type II
modification systems. We conclude that the type II restriction-modification systems in H.
pylori are highly diverse between strains, a unique characteristic of H. pylori. The diverse
methylation status of H. pylori chromosomal DNA may serve as a new typing system to
discriminate H. pylori isolates for epidemiological and clinical purposes. This study also
demonstrates that H. pylori is a rich source of type II restriction endonucleases.

<>

<1>Xu, Q., Morgan, R.D., Roberts, R.J., Xu, S.Y., van Doorn, L.J., Donahue, J.P., Miller, G.G., Blaser, M.J.
<2>Functional analysis of iceA1, a CATG-recognizing restriction endonuclease gene in Helicobacter pylori.
<3>Nucleic Acids Res.
<4>30
<5>3839-3847
<6>2002
<7>iceA1 in Helicobacter pylori is a homolog of nlaIIIR, which encodes the CATG-specific
restriction endonuclease NlaIII in Neisseria lactamica. Analysis of iceA1 sequences from 49
H.pylori strains shows that a full-length NlaIII-like ORF is present in 10 strains, including
CH4, but in other strains, including strain 60190, the ORFs are truncated due to a variety of
mutations. Our goal was to determine whether iceA1 can encode a NlaIII-like endonuclease.
Overexpression in Escherichia coli of iceA1 from CH4, but not from 60190, yielded NlaIII-like
activity, indicating that the full-length iceA1 is a functional endonuclease gene. Repair of
the iceA1 frameshift mutation in strain 60190 and its expression in E.coli yielded functional
NlaIII-like activity. We conclude that iceA1 in CH4 is a functional restriction endonuclease
gene, while iceA1 in 60190 is not, due to a frameshift mutation, but that its repair restores
its restriction endonuclease activity.

<>

<1>Xu, Q., Morgan, R.D., Xu, S.Y., van Doorn, L.J., Donahue, J.P., Miller, G.G., Blaser, M.J.
<2>H. pylori iceA1 is capable of encoding an NlaIII-like restriction endonuclease.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>100
<5>291
<6>2000
<7>H. pylori iceA1 has strong homology to nlaIIIR, which encodes NlaIII, a restriction
endonuclease in N. lactamica.  Our goal was to determine whether iceA1 encodes an NlaIII-like
RE.  Analysis of iceA1 sequences from 19 H. pylori strains was performed.  A full-length
NlaIII-like ORF is present in 4 strains including CH4, but iceA1 from 15 others, including
strain 60190, have various mutations.  Overexpression of iceA1 from CH4, but not from 60190,
in pAII17 vector in E. coli yielded NlaIII-like RE activity, indicating that it is a
functional gene.  To investigate the effect of mutations on iceA1 function, the frameshift
mutation of 60190 iceA1 was repaired.  The repaired 60190 iceA1 in pAII17 enabled expression
of a functional NlaIII-like RE in E. coli.  These results are consistent with biochemical
analysis of H. pylori, in which NlaIIII-like RE activity was detected in CH4, but not in 60190
cells.  We conclude that iceA1 in CH4 is a functional RE gene, and in 60190 is degenerate, but
that repair of its frameshift mutation restores its activity.

<>

<1>Xu, Q., O'Loughlin, T.J., Kucera, R.B., Schildkraut, I., Guo, H.-C.
<2>Structural study of restriction enzyme MspI/DNA complex by x-ray crystallography.
<3>Biophys. J.
<4>80
<5>296a
<6>2001
<7>The MspI restriction endonuclease is a type II restriction enzyme.  It recognizes the
palindromic sequence of four base pairs 5'-CCGG-3' and cleaves it between two cytosines,
leaving 5' two-base overhangs.  Owing to the nature of its cleavage pattern that is different
from all other restriction enzymes with known structure, it is likely that MspI could
represent a novel structural class of restriction endonucleases.  Solving the crystal
structure of MspI/DNA complex will allow us to examine the hypothesis that the cleavage
pattern is a result of intermediate organization of core architectures.  In addition, the
structure would provide us more information about the DNA recognition, specificity and
catalytic mechanisms of these enzymes.  Native crystals of the dimeric MspI restriction enzyme
bound to a duplex DNA molecule containing the specific recognition have been obtained in a P21
monoclinic unit cell with dimensions of a = 50.2A, b = 131.6A, c= 59.3A, beta = 109.7o.  The
crystals contain one dimeric complex in the asymmetric unit.  A complete native data set has
been collected to a resolution of 2.05A at beamline X12c at National Synchrotron Light Source
at Brookhaven National Laboratory.  Moreover, a complete MAD data set of Hg derivative and a
data set of Sm derivative have also been collected at X12c to the resolution of 3.2A and 2.6A,
respectively.  Phasing from these derivative data sets are currently underway.

<>

<1>Xu, Q., Peek, R., Miller, G., Blaser, M.
<2>Identification of an adenine methylase gene in Helicobacter pylori.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>97
<5>38
<6>1997
<7>DNA methylation has been demonstrated to be involved in several important cellular processes
such as host-specific defense mechanisms, DNA mismatch repair, regulation of initiation of
chromosomal replication, regulation of gene transcription, and DNA transposition.  To
understand mechanisms of DNA methylation in H. pylori, a human pathogen associated with peptic
ulcer disease and adenocarcinoma, we cloned a putative adenine methylase gene, hpyIM. This
gene contains a 990bp ORF encoding a 330 aa protein, M.HpyI.  Sequence analysis revealed that
M.HpyI was closely related to a CATG-recognizing adenine methylase, M.NlaIII in N. lactamica
(76% similarity).  Both Pcr and Southern blotting show that hpyIM is present in all 14 H.
pylori strains tested.  To investigate possible modification effects of M.HpyI, an isogenic
hpyIM mutant of H. pylori strain 88-23 was made by inserting a kanamycin resistance gene into
hpyIM.  SphI and NlaIII recognize DNA at sites containing CATG, and were used to test CATG
modification status.  DNA from wild type strains was resistant to digestion, whereas the hpyIM
mutant was susceptible, indicating lack of modification.  To further confirm that M.HpyI
modifies DNA at CATG sites, hpyIM was cloned into the expression vector pGEX-5X-1 to creat
pQX101.  DNA from BL21 was cleaved by NlaIII, as was the strain transformed with pGEX-5X-1.
However, DNA from BL21 with pQX101 encoding the GST-M.HpyI fusion protein was resistant,
confirming the role of hpyIM in modifying CATG sites.  We conclude that hpyIM encodes a
CATG-recognizing adenine methylase and is well-conserved among diverse H.pylori strains.

<>

<1>Xu, Q., Peek, R.M. Jr., Miller, G.G., Blaser, M.J.
<2>The Helicobacter pylori genome is modified at CATG by the product of hpyIM.
<3>J. Bacteriol.
<4>179
<5>6807-6815
<6>1997
<7>To understand mechanisms of DNA methylation in Helicobacter pylori, a human pathogen
associated with peptic ulcer disease and gastric adenocarcinoma, we cloned a putative DNA
methyltransferase gene, hpyIM.  This gene contains a 990-bp open reading frame encoding a
329-amino-acid protein, M.HpyI.  Sequence analysis revealed that M.HpyI was closely related to
CATG-recognizing adenine DNA methyltransferases, including M.NlaIII in N. lactamica.  hpyIM
was present in all H. pylori strains tested.  DNA from wild-type H. pylori strains was
resistant to digestion by SphI and NlaIII, which recognize DNA at sites containing CATG,
whereas their isogenic hpyIM mutants were susceptible, indicating lack of modification.
Overexpression of hpyIM in Escherichia coli rendered DNA from these cells resistant to NlaIII
digestion, confirming the role of hpyIM in modifying CATG sites.  We conclude that hpyIM
encodes a DNA methyltransferase, M.HpyI, that is well conserved among diverse H. pylori
strains and that modifies H. pylori genomes at CATG sites.

<>

<1>Xu, Q., Stickel, S., Roberts, R.J., Blaser, M.J., Morgan, R.D.
<2>Purification of the Novel Endonuclease, Hpy188I, and Cloning of Its Restriction-Modification Genes Reveal Evidence of Its Horizontal Transfer to the Helicobacter pylori Genome.
<3>J. Biol. Chem.
<4>275
<5>17086-17093
<6>2000
<7>We have isolated a novel restriction endonuclease, Hpy188I, from Helicobacter pylori strain
J188. Hpy188I recognizes the unique sequence, TCNGA, and cleaves the DNA between nucleotides N
and G in its recognition sequence to generate a one-base 3' overhang. Cloning and sequence
analysis of the Hpy188I modification gene in strain J188 reveal that hpy188IM has a 1299-base
pair (bp) open reading frame (ORF) encoding a 432-amino acid product. The predicted protein
sequence of M.Hpy188I contains conserved motifs typical of aminomethyltransferases, and
Western blotting indicates that it is an N-6 adenine methyltransferase. Downstream of hpy188IM
is a 513-bp ORF encoding a 170-amino acid product, that has a 41-bp overlap with hpy188IM. The
predicted protein sequence from this ORF matches the amino acid sequence obtained from
purified Hpy188I, indicating that it encodes the endonuclease. The Hpy188I R-M genes are not
present in either strain of H. pylori that has been completely sequenced but are found in two
of 11 H. pylori strains tested. The significantly lower G + C content of the Hpy188I R-M genes
implies that they have been introduced relatively recently during the evolution of the H.
pylori genome.

<>

<1>Xu, Q.S.
<2>Crystal structures of restriction endonuclease MspI-DNA complex and glycosylasparaginase.
<3>Ph.D. Thesis, Boston University, , , Boston
<4>
<5>1-161
<6>2003
<7>This dissertation is composed of two parts: (I) Chapter I, II, III and IV discuss the crystal
structures of restriction endonuclease MspI-DNA complex. (II) Chapter V depicts the
crystallographic studies of glycosylasparaginase.   Part I: MspI is a Type II restriction
endonuclease that recognizes and cleaves the palindromic tetranucleotide sequence C^CGG (the
arrowhead indicates the cleavage site) to produce two-base 5' overhanging ends. Since the
nature of this cleavage pattern is different from all other restriction enzymes with known
structures, MspI is anticipated to represent a new structural class of restriction
endonucleases.  This study reports two crystal structures of MspI bound to its cognate DNA.
One has been determined at 1.95 angstroms resolution in P21 space group, and the other at 2.7
angstroms resolution in P212121 space group. The two structures are essentially identical and
reveal several novel features. In the crystals, one MspI molecule is bound to a half-site of
its DNA recognition sequence. This is the first time that this binding mode has been observed
for Type IIP restriction endonucleases, suggesting a possibility that MspI may function as a
monomer although the dimer mechanism can't be ruled out at this point. DNA recognition is
accomplished mainly through a small three-stranded (-sheet region that is structurally
conserved in BglI, EcoRV as well as PvuII. While the architecture of the MspI active site is
similar to other restriction enzymes, an asparagine, instead of a conserved acidic residue, is
found in the catalytic sequence motif of Type II restriction enzymes. Meanwhile, although no
metal is found near the scissile phosphate, a cation can be seen at a position homologous to
the 74/45 metal site of EcoRV. Taken together, it appears that MspI is indeed a very unique
restriction enzyme.  Part II: Glycosylasparaginase (GA) is a member of a novel family of
N-terminal nucleophile hydrolases and is activated from a single chain precursor by
intramolecular autoproteolysis. The crystallographic studies of bacterial glycosylasparaginase
in mature form and its precursor reveal the catalytic mechanism of its hydrolase activity and
a novel mechanism of intramolecular autoproteolysis via an N-O or N-S acyl shift.

<>

<1>Xu, Q.S., Kucera, R.B., Roberts, R.J., Guo, H.-C.
<2>An asymmetric complex of restriction endonuclease MspI on its palindromic DNA recognition site.
<3>Structure
<4>12
<5>1741-1747
<6>2004
<7>Most well-known restriction endonucleases recognize palindromic DNA sequences and are
classified as Type IIP. Due to the recognition and cleavage symmetry, Type IIP enzymes are
usually found to act as homodimers in forming 2-fold symmetric enzyme-DNA complexes. Here we
report an asymmetric complex of the Type IIP restriction enzyme MspI in complex with its
cognate recognition sequence. Unlike any other Type IIP enzyme reported to date, an MspI
monomer and not a dimer binds to a palindromic DNA sequence. The enzyme makes specific
contacts with all 4 base pairs in the recognition sequence, by six direct and five
water-mediated hydrogen bonds and numerous van der Waal contacts. This MspI-DNA structure
represents the first example of asymmetric recognition of a palindromic DNA sequence by two
different structural motifs in one polypeptide. A few possible pathways are discussed for MspI
to cut both strands of DNA, either as a monomer or dimer.

<>

<1>Xu, Q.S., Roberts, R.J., Guo, H.C.
<2>Two crystal forms of the restriction enzyme MspI-DNA complex show the same novel structure.
<3>Protein Sci.
<4>14
<5>2590-2600
<6>2005
<7>The crystal structure of the Type IIP restriction endonuclease MspI bound to DNA containing
its cognate recognition sequence has been
determined in both monoclinic and orthorhombic space. groups.
Significantly, these two independent crystal forms present an identical
structure of a novel monomer-DNA complex, suggesting a functional role
for this novel enzyme-DNA complex. In both crystals, MspI interacts
with the CCGG DNA recognition sequence as a monomer, using an
asymmetric mode of recognition by two different structural motifs in a
single polypeptide. In the crystallographic asymmetric unit, the two
DNA molecules in the two MspI-DNA complexes appear to stack with each
other forming an end-to-end pseudo-continuous 19-mer duplex. They are
primarily B-form and no major bends or kinks are observed. For DNA
recognition, most of the specific contacts between the enzyme and the
DNA are preserved in the orthorhombic structure compared with the
monoclinic structure. A cation is observed near the catalytic center in
the monoclinic structure at a position homologous to the 74/45 metal
site of EcoRV, and the orthorhombic structure also shows signs of this
same cation. However, the coordination ligands of the metal are
somewhat different from those of the 74/45 metal site of EcoRV.
Combined with structural information from other solved structures of
Type II restriction enzymes, the possible relationship between the
structures of the enzymes and their cleavage behaviors is discussed.

<>

<1>Xu, S., Maunus, R., Benner, J.
<2>Method For Cloning And Expression of BseRI Restriction Endonuclease And BseRI Methylase In E. coli.
<3>Japanese Patent Office
<4>JP 2005522985 A
<5>
<6>2005
<7>
<>

<1>Xu, S., Schildkraut, I.
<2>Protecting recognition sequences on DNA by a cleavage-deficient restriction endonuclease.
<3>Biotechniques
<4>15
<5>310-315
<6>1993
<7>This report describes the use of a biochemical tool that has been developed to aid in the
manipulation of DNA. A DNA binding-proficient and cleavage-deficient BamHI mutant protein,
E113K, was used in vitro to protect its recognition sequence (5'-GGATCC-3') against the
catalytic action of site-specific endonuclease, exonuclease and methylase. In vitro conditions
are reported here in which the E113K protein protects BamHI sites (5'-GGATCC-3') from
cleavage by BamHI endonuclease or Sau3AI endonuclease (5'-GATC-3'); protects a neighboring
restriction site 5'-CCCGGG-3' from SmaI endonuclease digestion; blocks methylation of
5'-GGATCC-3' by Dam methylase (5'-GATC-3'); and blocks Bal31 exonuclease progression at a
BamHI site. The Bal31 procedure could be used to generate unidirectional deletions of a DNA
fragment. The use of mutant endonucleases that are binding-proficient and cleavage-deficient
to shield DNA from nuclease digestion or methylase modification expands the repertoire of
methods to manipulate DNA in vitro.

<>

<1>Xu, S., Shiyao, C.
<2>Cloning of BssHII restriction enzyme in Escherichia coli and production process.
<3>Japanese Patent Office
<4>JP 1998313883 A, JP 2000109482 A
<5>
<6>1998
<7>
<>

<1>Xu, S., Zhu, Z.
<2>Method for Cloning and Expression of StuI Restriction Endonuclease and StuI Methylase in E. coli.
<3>Japanese Patent Office
<4>JP 2009528839 A
<5>
<6>2009
<7>
<>

<1>Xu, S., Zhu, Z., Riggs, P.D., Hsieh, P.
<2>Recycled mutagenesis of restriction endonuclease toward enhanced catalytic activity.
<3>US Patent Office
<4>US 20050003420
<5>
<6>2005
<7>NOVELTY - Modifying the cleavage activity of a restriction endonuclease comprises creating a
pool of mutants of an
endonuclease gene coding for a target restriction endonuclease.
DETAILED DESCRIPTION - Modifying the cleavage activity of a restriction
endonuclease comprises: (a) creating a pool of mutants of an
endonuclease gene coding for a target restriction endonuclease; (b)
transforming a host cell with the mutants of step (a) under conditions
where the expressed unmutated gene coding for the target restriction
endonuclease generates little or no DNA damage within the host;(c)
screening the pool of transformed mutants within the host cell for an
increase in DNA damage; and (d) obtaining a mutant restriction
endonuclease with modified cleavage activity. INDEPENDENT CLAIMS are
also included for BsoBI variants, which cleave a subset of the native
BsoBI recognition/cleavage, sit and PvuII variants, which cleave a
subset of the native PvuII recognition/cleavage site. BIOTECHNOLOGY -
Preferred Method: Prior to creating the pool of mutants, a
predetermined mutation of the restriction endonuclease gene is
engineered to alter the specificity of the target restriction
endonuclease, where the altered specificity comprises an altered
binding or cleavage specificity of the target restriction endonuclease.
Altered specificity is screened for before and/or after the screening
for increased DNA damage. Preferred Variants: The BsoBI variants are
D246A, D246S, D246T, D246A/C317W, or
R38S/L62I/D112Y/N169T/L238F/D246A/A248S. The PvuII variants are
K93R/R105H, K93R/V50A and E18K/K93R/R105H. USE - The method is useful
for modifying the cleavage activity of a restriction endonuclease and
for increasing and enhancing catalytic activity. (23 pages)

<>

<1>Xu, S.-Y.
<2>Method for direct cloning and producing the BsoBI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 5492823
<5>
<6>1996
<7>The present invention discloses a novel method for the direct cloning of nuclease
genes such as restriction endonuclease genes in E. coli.  In addition, there is provided a
novel
strain which facilitates application of the method.  This method has been successfully
employed to
clone a number of genes coding for endonuclease including restriction endonuclease genes.

<>

<1>Xu, S.-Y.
<2>Cloning and expression of the ApaLI restriction endonuclease.
<3>US Patent Office
<4>US 5616484
<5>
<6>1997
<7>The present invention relates to a method of cloning ApaLI methylase gene (apaLIM) and ApaLI
endonuclease gene (apaLIR) from Acetobacter pasteurianus into E. coli by the methylase
selection method and inverse PCR.  The ApaLI methylase gene was cloned into pUC19 (3 ApaLI
sites inserted) by the methylase selection method.  Eight ApaLI-resistant clones were isolated
and found to contain apaLIM gene.  However, these clones are not stable such that sometimes
overnight cultures were lysed or plasmid DNA was lost in the unlysed culture.

<>

<1>Xu, S.-Y.
<2>Method for cloning and producing the NspI restriction endonuclease in E. coli and purification of the recombinant NspI restriction endonuclease.
<3>US Patent Office
<4>US 6027929
<5>
<6>2000
<7>The present invention relates to clones which express the recombinant NspI restriction
endonuclease using an NspI methylase premodified E. coli K strain RR1 (lambdaDE3) and to
methods for producing and purifying said enzyme.

<>

<1>Xu, S.-Y., Dore, A., Dalton, M., Benner, J.
<2>Method for cloning and expression of TspRI restriction endonuclease and TspRI methylase in E. coli.
<3>US Patent Office
<4>US 6589769
<5>
<6>2003
<7>The present invention relates to recombinant DNA that encodes the TspRI restriction
endonuclease as well as TspRI methylase, expression of
TspRI restriction endonuclease and TspRI methylase in E. coli cells
containing the recombinant DNA.

<>

<1>Xu, S.-Y., Dore, A., Dalton, M.A., Benner, J.
<2>Cloning and sequences of TspRI restriction endonuclease and TspRI methylase from Thermus and expression of TspRI in E. coli.
<3>International Patent Office
<4>WO 200335828 A
<5>68
<6>2003
<7>The present invention relates to recombinant DNA that encodes the TspRI restriction
endonuclease as well as TspRI, methylase, expression of TspRI restriction endonuclease and
TspRI methylase in E. coli cells containing the recombinant DNA.

<>

<1>Xu, S.-Y., Dore, A., Hume, A., Pelletier, J., Zhou, J.
<2>Method for cloning and expression of BsmBI restriction endonuclease and BsmBI methylase in E. coli and purification of BsmBI endonuclease.
<3>European Patent Office
<4>EP 1298212 A
<5>
<6>2003
<7>The present invention relates to recombinant DNA that encodes the BsmBI restriction
endonuclease as well as BsmBI methyltransferase, expression of BsmBI restriction endonuclease
and BsmBI methylase in E. coli cells containing the recombinant DNA.  It also relates to a
method for purification of the recombinant BsmBI restriction endonuclease.

<>

<1>Xu, S.-Y., Dore, A., Hume, A., Pelletier, J., Zhou, J.
<2>Method of cloning and expression of BsmBI restriction endonuclease and BsmBI methylase in E. coli and purification of BsmBI endonuclease.
<3>US Patent Office
<4>US 6764843
<5>
<6>2004
<7>The present invention relates to recombinant DNA that encodes the BsmBI restriction
endonuclease as well as BsmBI methyltransferase, expression
of BsmBI restriction endonuclease and BsmBI methylase in E. coli cells
containing the recombinant DNA. It also relates to a method for
purification of the recombinant BsmBI restriction endonuclease.

<>

<1>Xu, S.-Y., Fomenkov, A.
<2>Construction of pSC101 derivatives with Camr and Tetr for selection or LacZ' for blue/white screening.
<3>Biotechniques
<4>17
<5>57
<6>1994
<7>Methylase genes and a mutant bamhIR gene have been cloned in the vectors described.

<>

<1>Xu, S.-Y., Hsieh, P.-C.
<2>Method for cloning and producing the TfiI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 6133008
<5>
<6>2000
<7>A genomic DNA library of Thermus filiformis was constructed using pBR322 as a cloning vector.
The methylase selection method was used to clone the TfiI methylase gene (tfiIM).  A clone
carrying an active TfiI methylase was identified.  After sequencing the complete TfiI
methylase gene and its downstream DNA sequence, a recombinase homolog was found.  Because the
methylase and its cognate endonuclease gene are located in proximity to each other in a
particular restriction-modification system, efforts were made to clone the upstream DNA by
inverse PCR.  After two rounds of inverse PCR, one open reading frame (ORF1) was found
upstream of the TfiI methylase gene.  This ORF1, containing a Shine-Dalgarno sequence and a
TATA box on the upstream side, was cloned and expressed, and TfiI endonuclease activity was
detected in crude cell extracts.  It is concluded that ORF1 encodes TfiI restriction
endonuclease.

<>

<1>Xu, S.-Y., Kobbe, D., Zhu, Z., Samuelson, J.
<2>Methods for altering the cleavage specificity of a type IIG restriction endonuclease.
<3>US Patent Office
<4>US 20040219584 A
<5>34
<6>2004
<7>Methods are provided for altering the cleavage specificity of a Type IIG restriction
endonuclease, the Type IIG restriction endonuclease being characterized by a cleavage domain
adjacent to a methylase domain, the methylase domain located adjacent to a specificity domain.
The method includes ligating DNA or protein sequences to form a fusion DNA or fusion protein.
Where a fusion DNA is formed, the host cell is transformed with the fusion DNA to express a
Type IIG restriction endonuclease with altered cleavage specificity.

<>

<1>Xu, S.-Y., Maunus, R., Benner, J.
<2>Method for cloning and expression of BseRI restriction endonuclease and BseRI methylase of Bacillus in E. coli.
<3>International Patent Office
<4>WO 200331570 A
<5>74
<6>2003
<7>The present invention relates to recombinant DNA that encodes the BseRI restriction
endonuclease as well as M.BseRI, expression of BseRI restriction endonuclease and M.BseRI in
E. coli cells containing the recombinant DNA.

<>

<1>Xu, S.-y., Maunus, R., Benner, J.
<2>Method for cloning and expression of BseRI restriction endonuclease and BseRI methylase in E. coli.
<3>US Patent Office
<4>US 6593122 A
<5>
<6>2003
<7>The present invention relates to recombinant DNA that encodes the BseRI restriction
endonuclease as well as M.BseRI, expression of BseRI restriction endonuclease and M.BseRI in
E. coli cells containing the recombinant DNA.

<>

<1>Xu, S.-Y., Maunus, R., Stropnicky, K.
<2>Method for cloning and expression of MspA1l restriction endonuclease and MspA1l methylase in E. coli.
<3>US Patent Office
<4>US 6673588
<5>
<6>2004
<7>The present invention relates to recombinant DNA coding for the MspA1I restriction
endonuclease as well as MspA1I methylase, expression of
MspA1I restriction endonuclease and MspA1I methylase in E. coli cells
containing the recombinant DNA.

<>

<1>Xu, S.-Y., Maunus, R., Stropnicky, K.
<2>Cloning and expression of Moraxella MspA1I restriction endonuclease and MspA1I methylase in Escherichia coli.
<3>International Patent Office
<4>WO 200372702 A
<5>33
<6>2003
<7>The present invention relates to recombinant DNA coding for the MspA1I restriction
endonuclease as well as MspA1I methylase, expression of MspA1I restriction endonuclease and
MspA1I methylase in E. coli cells containing the recombinant DNA.

<>

<1>Xu, S.-Y., Maunus, R.E., Lunnen, K.D., Allen, R.
<2>Method for cloning and producing the AgeI restriction endonuclease in E. coli.
<3>European Patent Office
<4>EP 0959131 A
<5>
<6>1999
<7>The present invention relates to recombinant DNA which encodes the AgeI restriction
endonuclease as well as AgeI methyltransferase, and production of AgeI restriction
endonuclease from E. coli cells containing the recombinant DNA.

<>

<1>Xu, S.-Y., Maunus, R.E., Lunnen, K.D., Allen, R.
<2>Method for cloning and purifying AgeI restriction endonuclease in Escherichia coli.
<3>Japanese Patent Office
<4>JP 1999318476 A
<5>
<6>1999
<7>The present invention relates to recombinant DNA which encodes the AgeI restriction
endonuclease as well as AgeI methyltransferase, and production of AgeI restriction
endonuclease from E. coli cells containing the recombinant DNA.

<>

<1>Xu, S.-Y., Maunus, R.E., Lunnen, K.D., Allen, R.
<2>Method for cloning and producing AgeI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 5885818
<5>
<6>1999
<7>The present invention relates to recombinant DNA which encodes the AgeI restriction
endonuclease as well as AgeI methyltransferase, and production of AgeI restriction
endonuclease from E. coli cells containing the recombinant DNA.

<>

<1>Xu, S.-Y., Nwankwo, D.O.
<2>Method for cloning and producing the AatII restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0581436 A
<5>
<6>1992
<7>The present invention provides a novel approach to the production of restriction
endonucleases. More specifically, there is provided a novel method for the overexpression of
these enzymes, which comprises transforming a host cell with an expression vector containing
DNA coding for the restriction endonuclease of interest and a vector for the expression of a
methylase which is capable of protecting the host cell DNA against the restriction
endonuclease of interest. The methylase vector is compatible with the restriction endonuclease
expression vector. Preferably, the expression vector is a T7 expression vector and the cell is
transformed with a third compatible vector which regulates the expression vector. The cloning
and overexpression of the AatII restriction endonuclease and methylase is also described in
detail.

<>

<1>Xu, S.-Y., Nwankwo, D.O.
<2>Method for cloning and producing the AatII and AluI restriction endonuclease and methylase and related method for overexpressing restriction endonucleases.
<3>US Patent Office
<4>US 5405768
<5>
<6>1995
<7>The present invention provides a novel approach to the production of restriction
endonucleases. More specifically, there is provided a novel method for the overexpression of
these enzymes, which comprises transforming a host cell with an expression vector containing
DNA coding for the restriction endonuclease of interest and a vector for the expression of a
methylase which is capable of protecting the host cell DNA against the restriction
endonuclease of interest. The methylase vector is compatible with the restriction endonuclease
expression vector. Preferably, the expression vector is a T7 expression vector and the cell is
transformed with a third compatible vector which regulates the overexpression vector. Also
disclosed is the cloning and overexpression of the AatII restriction endonuclease and
methylase and the cloning and overexpression of the AluI restriction endonuclease and
methylase.

<>

<1>Xu, S.-Y., Nwankwo, D.O.
<2>Method for cloning and producing the AatII restriction endonuclease and methylase.
<3>European Patent Office
<4>EP 0581436 B
<5>
<6>2000
<7>The present invention relates to recombinant DNA which encodes the AatII restriction
endonuclease and modification methylase, and the production of these enzymes from the
recombinant DNA.

<>

<1>Xu, S.-Y., Samuelson, J.
<2>Method for engineering strand-specific nicking endonucleases from restriction endonucleases.
<3>US Patent Office
<4>US 20050158834 A
<5>
<6>2005
<7>
<>

<1>Xu, S.-Y., Samuelson, J.
<2>Method for engineering strand-specific nicking endonucleases from restriction endonucleases.
<3>US Patent Office
<4>US 07943303
<5>
<6>2011
<7>Methods are provided for engineering novel strand-specific nicking endonucleases by means of
an in vivo enrichment of a plasmid library
containing a randomly mutagenized restriction endonuclease gene. The
plasmids contain adjacent to the gene a cleavable or nickable sequence
for cleaving or nicking by the endonuclease product of the gene and a
second recognition site for a second endonuclease. The plasmid library
is used to transform unmodified host cells. Plasmids from the cultured
transformed cells may be analyzed by an in vitro assay for nicking and
the nicked plasmids pooled and used to transform host cells. The
product is then pooled and the single-stranded specificity of the
endonuclease is then determined. The product is either cloned after
amplification or identified by use of a selectable marker.

<>

<1>Xu, S.-Y., Samuelson, J.
<2>Method for engineering strand-specific nicking endonucleases from restriction endonucleases.
<3>US Patent Office
<4>US 7943303 B
<5>
<6>2011
<7>Methods are provided for engineering novel strand-specific nicking endonucleases by means of
an in vivo enrichment of a plasmid library containing a randomly mutagenized restriction
endonuclease gene.  The plasmids contain adjacent to the gene a cleavable or nickable sequence
for cleaving or nicking by the endonuclease product of the gene and a second recognition site
for a second endonuclease.  The plasmid library is used to transform unmodified host cells.
Plasmids from the cultured transformed cells may be analyzed by an in vitro assay for nicking
and the nicked plasmids pooled and used to transform host cells.  The product is then pooled
and the single-stranded specificity of the endonuclease is then determined.  The product is
either cloned after amplification or identified by use of a selectable marker.

<>

<1>Xu, S.-Y., Samuelson, J., Pelletier, J., Sibley, M., Wilson, G.G.
<2>Method for cloning, and expression in E. coli, and purification of BstYI restriction endonuclease and BstYI methylase from Bacillus stearothermophilus Y406.
<3>US Patent Office
<4>US 6403354 B
<5>18
<6>2002
<7>The present invention relates to recombinant DNA which encodes the BstYI restriction
endonuclease as well as BstYI methyltransferase, expression of BstYI restriction endonuclease
and M.BstYI in E. coli cells containing the recombinant DNA. It also relates to methods for
purification of the recombinant BstYI restriction endonuclease and BstYI methyltransferase.

<>

<1>Xu, S.-Y., Schildkraut, I.
<2>Isolation of BamHI variants with reduced cleavage activities.
<3>J. Biol. Chem.
<4>266
<5>4425-4429
<6>1991
<7>Derivation of the bamhIR sequence (Brooks, J.E., Nathan, P.D., Landry, D.,
Sznyter, L.A., Waite-Rees, P., Ives, C.C., Mazzola, L.M., Slatko, B.E., and
Benner, J.S. (1991) Nucleic Acids Res., in press), the gene coding for BamHI
endonuclease, has facilitated construction of an Escherichia coli strain that
overproduces BamHI endonuclease (W.E. Jack, L. Greenough, L.F. Dorner, S.Y. Xu,
T. Strezelecka, A.K. Aggarwal, and I. Schildkraut, submitted for publication).
As expected, low-level constitutive expression of the bamhIR gene in E. coli
from the Ptac promoter construct is lethal to the host unless the bamHIM gene,
which encodes the BamHI methylase, is also expressed within the cell.  We
identified four classes of BamHI endonuclease variants deficient in catalysis
by selecting for survival of a host deficient for bamHIM gene, transformed with
mutagenized copies of the bamhIR gene, and then screening the surviving cell
extracts for DNA cleavage and binding activities.  Class I variants (G56S,
G91S/T153I, G130R, E135K, T153I, T157I, G194D) displayed 0.1-1% of the
wild-type cleavage activity; class II variant (D94N) lacked cleavage activity
but retained wild-type DNA binding specificity; class III variants (E77K,
E113K) lacked cleavage activity but bound DNA more tightly; class IV variants
(G56D, G90D, G91S, R122H, R155H) lacked both binding and cleavage activities.
Variants with residual cleavage activities induced the E. coli SOS response and
thus are presumed to cleave chromosomal DNA in vivo.  We conclude that Glu77,
Asp94, and Glu113 residues are essential for BamHI catalytic function.

<>

<1>Xu, S.-Y., Schildkraut, I.
<2>Cofactor requirements of BamHI mutant endonuclease E77K and its suppressor mutants.
<3>J. Bacteriol.
<4>173
<5>5030-5035
<6>1991
<7>A mutant BamHI endonuclease, E77K, belongs to a class of catalytic mutants that
bind DNA efficiently but cleave DNA at a rate more than 10/3-fold lower than
that of the wild-type enzyme (S.Y. Xu and I. Schildkraut, J. Biol. Chem.
266:4425-4429, 1991).  The preferred cofactor for the wild-type BamHI is Mg2+.
BamHI is 10-fold less active with Mn2+ as the cofactor.  In contrast, the E77K
variant displays an increased activity when Mn2+ is substituted for Mg2+ in the
reaction buffer.  Mutations that partially suppress the E77K mutation were
isolated by using an Escherichia coli indicator strain containing the
dinD::lacZ fusion.  These pseudorevertant endonucleases induce E. coli SOS
response (as evidenced by blue colony formation) and thus presumably nick or
cleave chromosomal DNA in vivo.  Consistent with the in vivo result, the
pseudorevertant endonucleases in the crude cell extract display site-specific
partial DNA cleavage activity.  DNA sequencing revealed two unique suppressing
mutations that were located within two amino acid residues of the original
mutation.  Both pseudorevertant proteins were purified and shown to increase
specific activity at least 50-fold.  Like the wild-type enzyme, both
pseudorevertant endonucleases prefer Mg2+ as the cofactor.  Thus, the
second-site mutation not only restores partial cleavage activity but also
suppresses the metal preference as well.  These results suggest that the Glu-77
residue may play a role in metal ion binding or in enzyme activation
(allosteric transition) following sequence-specific recognition.

<>

<1>Xu, S.-Y., Sibley, M., Samuelson, J., Wilson, G., Pelletier, J.
<2>Method for cloning and expression of BstYI restriction endonuclease and BstYI methylase in E. coli and purification of BstYI and M.BstYI enzymes.
<3>European Patent Office
<4>EP 1225219 B
<5>
<6>2006
<7>The present invention relates to a method for cloning the BstYI restriction endonuclease from
Bacillus stearothermophilus into E. coli by methylase selection and inverse PCR amplification
of the adjacent DNA.  A methylase gene with high homology to amino-methyltransferases
(N4-cytosine methylases) was found in a DNA library after methylase selection.  This gene was
named BstYI methylase gene (bstYIM).

<>

<1>Xu, S.-y., Xiao, J.-i.
<2>Method for cloning and expression of the NheI restriction endonuclease in E. coli.
<3>European Patent Office
<4>EP 1096015 B
<5>
<6>2006
<7>
<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Method for cloning and producing the SacI restriction endonuclease.
<3>US Patent Office
<4>US 5532153
<5>
<6>1996
<7>The present invention relates to recombinant DNA which encodes the SacI restriction
endonuclease and modification methylase, and production of SacI restriction endonuclease from
the recombinant DNA.

<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Method for cloning and producing the ScaI restriction endonuclease in E. coli.
<3>European Patent Office
<4>EP 0778346 A
<5>
<6>1995
<7>The present invention relates to isolated DNA coding for the restriction endonuclease ScaI as
well as to a method for cloning methylase genes from Streptomyces into E. coli by a
modification of the methylase selection method.  At first, the standard methylase gene
selection method was tried to clone the ScaI methylase gene using a high-copy-number cloning
vector pUC19 during library construction.  The ScaI methylase gene was refractory to cloning
by using pUC19, presumably due to the poor expression of the ScaI methylase gene in E. coli.
If the ScaI methylase is not efficiently expressed in E. coli, the ScaI sites on the plasmid
will not be sufficiently modified by the methylase.  As a consequence, the plasmid will be
cleaved and lost in the plasmid library after ScaI endonuclease challenge.  Since the standard
methylase selection did not work, the "endo-blue method" was tried to clone the ScaI
endonuclease gene.  Nineteen blue colonies were identified, but none of them yielded any
detectable ScaI endonuclease activity.  The ScaI endonuclease gene was first cloned by inverse
PCR using primers that annealed to the end of the ScaI methylase gene.  In order to increase
the ScaI endonuclease expression in E. coli, an optimal ribosome binding site and spacing were
engineered in front of the ATG start codon and the gene was inserted into expression vector
pRRS.

<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Cloning and producing the BssHII restriction endonuclease and DNA methylase in Escherichia coli.
<3>US Patent Office
<4>US 5786195
<5>
<6>1998
<7>The present invention relates to cloning recombinant DNA molecules encoding a multi-specific
methylase gene (bssHIIM1), BssHII restriction endonuclease gene (bssHIIR), and the cognate
BssHII methylase gene (bssHIIM2) from Bacillus stearothermophilus H3 in E. coli.  The BssHII
multi-specific methylase gene was first cloned in a Sau3AI library using a modified pLITMUS28
vector (New England Biolabs, Inc., Beverly, Mass.) with two BssHII sites.  Expression of the
multi-specific BssHII methylase renders the two BssHII sites resistant to BssHII digestion.
Surprisingly, the cloned methylase also modifies some other sites in addition to the BssHII
site (5'GCGCGC3').  The methylase also modifies a BsrFI site (5'GCGCGC3') and a HaeII site
(5'RGCGCY3'); and partially modifies an EagI site (5'CGGCCG3') and an MluI site
(5'ACGCGT3').  The beginning of the bssHIIR gene was cloned by using two degenerate primers
based on the N-terminal amino acid sequence in PCR.  The rest of the bssHIIR gene was cloned
by inverse PCR.  The cognate bssHIIM2 gene was cloned by inverse PCR and PCR.  The BssHII
restriction endonuclease gene was expressed in E. coli host ER417 carrying three plasmids
pLysP, pLG-BssHIIM2, pET21AT-BssHIIR.

<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Method for cloning and producing the BslI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 5866398
<5>
<6>1999
<7>The methylase selection method was used to clone the BslI methylase gene (bslIM) from Bacillus
species.  A partially active BslI methylase lacking the 17 amino acid residues at the
N-terminus was cloned in E. coli using expression vector pRRS.  Inverse PCR was used to clone
the missing portion of the BslI methylase.  After cloning the complete BslI methylase gene and
its upstream DNA sequences, a RadC homolog was found upstream of the BslI methylase.  Because
methylase gene and restriction endonuclease gene are located in proximity to each other in a
particular restriction-modification system, efforts were made to clone the downstream DNA by
inverse PCR.  After two rounds of inverse PCR reactions two open reading frames were found
downstream of the BslI methylase gene.  Expression of the first ORF (ORF1) in a T7 expression
vector did not yield any active BslI endonuclease.  Expression of the second ORF (ORF2) in E.
coli and assay of the crude cell extract indicated that this gene product has DNA nicking
activity.  The gene product of ORF2 alone does not constitute BslI endonuclease activity.
Expression of ORF1 and ORF2 in the same E. coli cell produces BslI endonuclease activity.
BslI endonuclease activity can be reconstituted in vitro by mixing gene product of ORF1 and
ORF2 together.

<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Method for cloning and producing the ScaI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 5721126
<5>
<6>1998
<7>The present invention relates to isolated DNA coding for the restriction endonuclease ScaI as
well as to a  method for cloning methylase genes from Streptomyces into E. coli by a
modification of the methylase selection method.  At first, the standard methylase gene
selection method was tried to clone the ScaI methylase gene using a high-copy-number cloning
vector pUC19 during library construction.  The ScaI methylase gene was refractory to cloning
by using pUC19, presumably due to the poor expression of the ScaI methylase gene in E. coli.
If the ScaI methylase is not efficiently expressed in E. coli, the ScaI sites on the plasmid
will not be sufficiently modified by the methylase.  As a consequence, the plasmid will be
cleaved and lost in the plasmid library after ScaI endonuclease challenge.  Since the standard
methylase selection did not work, the "endo-blue method" was tried to clone the ScaI
endonuclease gene. Nineteen blue colonies were identified, but none of them yielded any
detectable ScaI endonuclease activity.  The ScaI endonuclease gene was first cloned by inverse
PCR using primers that annealed to the end of the ScaI methylase gene.  In order to increase
the ScaI endonuclease expression in E. coli, an optimal ribosome binding site and spacing were
engineered in front of the ATG start codon and the gene was inserted into expression vector
pRRS.

<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Method for cloning and expression of BsrFI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 6066487
<5>
<6>2000
<7>BsrFI restriction enzyme was purified from Bacillus stearothermophilus to near homogeneity.
The protein was sequenced to obtain its N-terminus amino acid sequence.  A set of degenerate
primers were synthesized based on the aa sequence.  The first 18 codons encoding BsrFI
restriction endonuclease (bsrFIR) was amplified by PCR and its coding sequence was obtained.
The methylase selection method was used to clone BsrFI methylase gene (bsrFIM).  Two clones
were found to be resistant to BsrFI digestion.  The entire insert in one clone was sequenced
and the insert encodes the BsrFI methylase (M.BsrFI).  In addition, a small truncated open
reading frame adjacent to the methylase gene has homology to Cfr10I restriction endonuclease
in a BlastX homology search in GenBank database.  BsrFI and Cfr10I are isoschizomers that
recognize and cleave 5'R/CCGGY3'.  Two primers were used to amplify the bsrFIR gene.  The
forward primer is a degenerate primer designed from the N-terminus aa sequence and the reverse
primer is the bona fide sequence derived from the BsrFI methylase + clone.  The bsrFIR gene
was amplified by PCR, ligated into a T7 expression vector pET21 at and the ligated DNA was
transformed into premodified cells ER2566 [pLG339-BsrFIM].  The final expression strain is
ER2566 [pLG339-BsrFIM, pET21at-BsrFIR].  Recombinant BsrFI activity was detected in E. coli
cell extract.  BsrFI is cloned from a thermophile Bacillus stearothermophilus.  Thus, BsrFI is
a thermostable enzyme and it is active at 37 C to 65 C.

<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Method for cloning and expression of the NheI restriction endonuclease in E. coli.
<3>European Patent Office
<4>EP 1096015 A
<5>
<6>2001
<7>The present invention relates to recombinant DNA which encodes the NheI restriction
endonuclease as well as NheI methyltransferase, expression of NheI restriction endonuclease
from E. coli cells containing the recombinant DNA.  An internal NdeI site in the nheIR gene
was eliminated by a silent mutation.  A new NdeI site was engineered at the start codon of
nheIR gene.  An NdeI-BamHI fragment containing the nheIR gene was cloned into a T7 expression
vector pAII17 and expressed in a premodified host ER2566 [pACYC-NheIM, pAII17-NheIR2].  The
recombinant clone produces approximately 10 million units of NheI per gram of wet cells.

<>

<1>Xu, S.-Y., Xiao, J.-P.
<2>Method for cloning and expression of NheI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 6387681
<5>
<6>2002
<7>The present invention relates to recombinant DNA which encodes the NheI restriction
endonuclease as well as NheI methyltransferase, expression of NheI restriction endonuclease
from E. coli cells containing the recombinant DNA. An internal NdeI site in the nheIR gene was
eliminated by a silent mutation. A new NdeI site was engineered at the start codon of nheIR
gene. An NdeI-BamHI fragment containing nheIR gene was cloned into a T7 expression vector
pAII17 and expressed in a premodified host ER2566 [pACYC-NheIM, pAII17-NheIR2]. The
recombinant clone produces approximately 10 million units of NheI per gram of wet cells.

<>

<1>Xu, S.-y., Xiao, J.-p.
<2>Cloning of BssHII restriction enzymes in Escherichia coli and its production.
<3>Japanese Patent Office
<4>JP 4165775
<5>
<6>2008
<7>
<>

<1>Xu, S.-Y., Xiao, J.-P., Ettwiller, L., Holden, M., Aliotta, J., Poh, C.L., Dalton, M., Robinson, D.P., Petronzio, T.R., Moran, L., Ganatra, M., Ware, J., Slatko, B., Benner, J.
<2>Cloning and expression of the ApaLI, NspI, NspHI, SacI, ScaI, and SapI restriction-modification systems in Escherichia coli.
<3>Mol. Gen. Genet.
<4>260
<5>226-231
<6>1998
<7>The genes encoding the ApaLI (5'-G^TGCAC-3'), NspI (5'-GCATG^Y-3'), NspHI
(5'-RCATG^Y-3'), SacI (5'-GAGCT^C-3'), SapI (5'-GCTCTTCN1^-3', 5'-^N4GAAGAGC-3') and
ScaI (5'-AGT^ACT-3') restriction-modification systems have been cloned in E. coli.  Amino
acid sequence comparison of M.ApaLI, M.NspI, M.NspHI, and M.SacI with known methylases
indicated that they contain the ten conserved motifs characteristic of C5 cytosine methylases.
NspI and NspHI restriction-modification systems are highly homologous in amino acid sequence.
The C-termini of the NspI and NlaIII (5'-CATG-3') restriction endonucleases share
significant similarity.  5mC modification of the internal C in a SacI site renders it
resistant to SacI digestion.  External 5mC modification of a SacI site has no effect on SacI
digestion.  N4mC modification of the second base in the sequence 5'-GCTCTTC-3' blocks SapI
digestion.  N4mC modification of the other cytosines in the SapI site does not affect SapI
digestion.  N4mC modification of ScaI site blocks ScaI digestion.  A DNA invertase homolog was
found adjacent to the ApaLI restriction-modification system.  A DNA transposase subunit
homolog was found upstream of the SapI restriction endonuclease gene.

<>

<1>Xu, S.-Y., Xiao, J.-P., Kucera, R.B., Samuelson, J.
<2>Method for cloning and expression of Oceanospirillum kriegii OkrAI restriction endonuclease and methyltransferase.
<3>US Patent Office
<4>US 20040175816 A
<5>22
<6>2004
<7>The present invention relates to recombinant DNA encoding the OkrAI restriction endonuclease
as well as OkrAI methylase, expression of OkrAI restriction endonuclease in E. coli cells
containing the recombinant DNA.

<>

<1>Xu, S.-Y., Xiao, J.-P., Maunus, R.E.
<2>Method for cloning and producing the SapI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 5663067
<5>
<6>1997
<7>The present invention relates to recombinant DNA which encodes the SapI restriction
endonuclease and modification methylase, and the production of SapI restriction endonuclease
from the recombinant DNA as well as to methods for cloning Actinomycetes genes into suitable
hosts such as E. coli.

<>

<1>Xu, S.-Y., Xiao, J.-P., Posfai, J., Maunus, R., Benner, J.
<2>Cloning of the BssHII restriction-modification system in Escherichia coli: BssHII methyltransferase contains circularly permuted cytosine-5 methyltransferase motifs.
<3>Nucleic Acids Res.
<4>25
<5>3991-3994
<6>1997
<7>BssHII restriction endonuclease cleaves 5'-GCGCGC-3' on double-stranded DNA between the
first and second bases to generate a four base 5' overhang.  BssHII restriction endonuclease
was purified from the native Bacillus stearothermophilus H3 cells and its N-terminal amino
acid sequence was determined.  Degenerate PCR primers were used to amplify the first 20 codons
of the BssHII restriction endonuclease gene.  The BssHII restriction endonuclease gene
(bssHIIR) and the cognate ssHII methyltransferase gene (bssHIIM) were cloned in Escherichia
coli by amplification of Bacillus stearothermophilus genomic DNA using PCR and inverse PCR.
BssHII methyltransferase (M.BssHII) contains all 10 conserved cytosine-5 methyltransferase
motifs, but motifs IX and X precede motifs I-VIII.  Thus, the conserved motifs of M.BssHII are
circularly permuted relative to the motif organizations of other cytosine-5
methyltransferases.  M.BssHII and the noncognate multi-specific PhiBssHII methyltransferase,
M.PhiBssHII share 34% identity in amino acid sequences from motifs I-VIII, and 40% identity in
motifs IX-X.  A conserved arginine is located upstream of a TV dipeptide in the N-terminus of
M.BssHII that may be responsible for the recognition of the guanine 5' of the target
cytosine.  The BssHII restriction enodnuclease gene was expressed in E.coli via a T7
expression vector.

<>

<1>Xu, S.-y., Xiao, J.-p., Zhu, Z.
<2>Method for cloning and expression of BpmI restriction endonuclease in E. coli.
<3>European Patent Office
<4>EP 1199365 A
<5>
<6>2002
<7>The present invention relates to recombinant DNA which encodes the BpmI restriction
endonuclease as well as BpmI methyltransferase, expression of BpmI restriction endonuclease
from E. coli cells containing the recombinant DNA.  BpmI endonuclease is a fusion of two
distinct elements with a possible structural domain of restriction-methylation specificity
(R-M-S).  This domain organization is analogous to the type I restriction-modification system
with three distinct subunits, restriction, methylation, and specificity (R, M, and S).
Because BpmI is quite distinct to other type IIS restriction enzymes, it is proposed that BpmI
belongs to a subgroup of type II restriction enzymes called type IIf (f stands for fusion of
restriction-modification-specificity domains).  The Type IIf group of restriction enzyme
includes Eco57I BpmI, GsuI, BseRI and some other restriction enzymes that cut downstream
sequences at long distance, 10-20 bp downstream of recognition sequence, such as MmeI
(N20/N18)).

<>

<1>Xu, S.-y., Xiao, J.-p., Zhu, Z.
<2>Method for cloning and expression of BpmI restriction endonuclease in E. coli.
<3>European Patent Office
<4>EP 1199365 B
<5>
<6>2005
<7>The present invention relates to a method for cloning the BpmI restriction endonuclease from
Bacillus pumilus into E. coli by methylase selection and inverse PCR amplification of the
adjacent DNA of the BpmI methylase gene.

<>

<1>Xu, S.-Y., Xiao, J.-P., Zhu, Z.
<2>Method for cloning and expression of BpmI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 6413758
<5>
<6>2002
<7>The present invention relates to recombinant DNA which encodes the BpmI restriction
endonuclease as well as BpmI methyltransferase, expression
of BpmI restriction endonuclease from E. coli cells containing the
recombinant DNA. BpmI endonuclease is a fusion of two distinct elements
with a possible structural domains of
restriction-methylation-specificity (R-M-S). This domain organization
is analogous to the type I restriction-modification system with three
distinct subunits, restriction, methylation, and specificity (R, M, and
S). Because BpmI is quite distinct to other type IIs restriction
enzymes, it is proposed that BpmI belongs to a subgroup of type II
restriction enzymes called type IIf (f stands for fusion of
restriction-modification-specificity domains). The Type IIf group of
restriction enzyme includes Eco57I, BpmI, GsuI, BseRI and some other
restriction enzymes that cut downstream sequences at long distance,
10-20 bp downstream of recognition sequence, such as MmeI (N20/N18)).

<>

<1>Xu, S.-Y., Zhou, J., Hume, A., Maunus, R.
<2>Method for cloning and expression of BsaWI restriction endonuclease and BsaWI methylase in E. coli.
<3>US Patent Office
<4>US 6586220
<5>
<6>2003
<7>The present invention relates to recombinant DNA encoding the BsaWI restriction endonuclease
as well as BsaWI methylase, expression of
BsaWI restriction endonuclease and BsaWI methylase in E. coli cells
containing the recombinant DNA.

<>

<1>Xu, S.-Y., Zhu, Z.
<2>Cloning and sequence of StuI restriction endonuclease and StuI methylase of Streptomyces tubercidicus, and recombinant expression of StuI in Escherichia coli.
<3>International Patent Office
<4>WO 2007103061 A
<5>
<6>2007
<7>The present invention relates to compositions including: (1) isolated DNA encoding the StuI
restriction endonuclease and isolated DNA encoding cognate and non-cognate methylase; (2)
vectors and cells containing the isolated DNA; and (3) methods for producing the StuI
restriction endonuclease.

<>

<1>Xu, S.-Y., Zhu, Z., Chan, S.-H., Xu, Y.
<2>DNA and protein sequences of DNA nicking endonuclease and associated methylase from Chlorella virus NC64A.
<3>International Patent Office
<4>WO 200647183 A
<5>
<6>2006
<7>Recombinant nicking endonucleases and associated methylases have been obtained and sequenced
and their specificity has been defined.  A mutant form of the nicking endonuclease has been
cloned where the mutation includes deletion of amino acid sequences at the C-terminal end of
the protein.  The nicking enzymes have been used for a number of purposes including:
amplifying DNA from as few cells as can be found in a single bacterial colony in the presence
of a strand displacing polymerase; and for removing genomic DNA in a biological preparation
where it is deemed to be a contaminant.

<>

<1>Xu, S.-Y., Zhu, Z., Meixsell, T.
<2>Preparation of strand-specific nicking endonucleases by site directed mutagenesis of restriction endonuclease.
<3>International Patent Office
<4>WO 200612593 A
<5>
<6>2006
<7>A nicking endonuclease is described which has an amino acid sequence with at least 70%
identity to SEQ ID NO: 6 and comprising a mutation at least one of an arginine or gutamic acid
corresponding to position 507 and position 546 respectively in SEQ ID No: 6.

<>

<1>Xu, S.-Y., Zhu, Z., Meixsell, T.
<2>Nicking endonuclease methods and compositions.
<3>US Patent Office
<4>US 7820424 B
<5>
<6>2010
<7>A nicking endonuclease is described which has an amino acid sequence with at least 70%
identity to SEQ ID NO: 6 and comprising a mutation at least one of an arginine or glutamic
acid corresponding to position 507 and position 546 respectively in SEQ ID NO: 6.

<>

<1>Xu, S.Y., Corvaglia, A.R., Chan, S.H., Zheng, Y., Linder, P.
<2>A type IV modification-dependent restriction enzyme SauUSI from Staphylococcus aureus subsp. aureus USA300.
<3>Nucleic Acids Res.
<4>39
<5>5597-5610
<6>2011
<7>A gene encoding a putative DNA helicase from Staphylococcus aureus USA300 was cloned and
expressed in Escherichia coli. The protein was purified to over 90% purity by chromatography.
The purified enzyme, SauUSI, predominantly cleaves modified DNA containing 5mC and
5-hydroxymethylcytosine. Cleavage of 5mC-modified plasmids indicated that the sites S5mCNGS (S
= C or G) are preferentially digested. The endonuclease activity requires the presence of
adenosine triphosphate (ATP) or dATP whereas the non-hydrolyzable gamma-S-ATP does not support
activity. SauUSI activity was inhibited by ethylenediaminetetraacetic acid. It is most active
in Mg(++) buffers. No companion methylase gene was found near the SauUSI restriction gene. The
absence of a cognate methylase and cleavage of modified DNA indicate that SauUSI belongs to
type IV restriction endonucleases, a group that includes EcoK McrBC and Mrr. SauUSI belongs to
a family of highly similar homologs found in other sequenced S. aureus, S. epidermidis and S.
carnosus genomes. More distant SauUSI orthologs can be found in over 150 sequenced
bacterial/archaea genomes. Finally, we demonstrated the biological function of the type IV
REase in restricting 5mC-modified plasmid DNA by transformation into clinical S. aureus strain
SA564, and in restricting phage lambda infection when the endonuclease is expressed in E.
coli.

<>

<1>Xu, S.Y., Nugent, R.L., Kasamkattil, J., Fomenkov, A., Gupta, Y., Aggarwal, A., Wang, X., Li, Z., Zheng, Y., Morgan, R.
<2>Characterization of Type II and III restriction-modification systems from Bacillus cereus strains ATCC10987 and ATCC14579.
<3>J. Bacteriol.
<4>194
<5>49-60
<6>2012
<7>The genomes of two Bacillus cereus strains (ATCC10987 and ATCC14579) have been sequenced. Here
we report the specificities of Type II/III restriction (R) and modification (M) enzymes. Found
in the ATCC10987 strain, BceSI is a restriction endonuclease (REase) with the recognition and
cut site CGAAG 24-25/27-28. BceSII is an isoschizomer of AvaII (G/GWCC). BceSIII cleaves at
ACGGC 12/14. The BceSIII C-terminus resembles the catalytic domains of AlwI, MlyI, and
Nt.BstNBI. BceSIV is composed of two subunits and cleaves on both sides of GCWGC. BceSIV
activity is strongly stimulated by the addition of cofactor ATP or GTP. The large subunit (R1)
of BceSIV contains conserved motifs of NTPases and DNA helicases. The R1 subunit has no
endonuclease activity by itself; it strongly stimulates REase activity when in complex with
the R2 subunit. BceSIV was demonstrated to hydrolyze GTP and ATP in vitro. BceSIV is similar
to CglI (GCSGC), and homologs of R1 are found in 11 sequenced bacterial genomes where they are
paired with specificity subunits. In addition, homologs of BceSIV R1-R2 fusion are found in
many sequenced microbial genomes. An orphan methylase M.BceSV was found to modify GCNGC, GGCC,
CCGG, GGNNCC, and GCGC sites. A ParB-methylase fusion protein appears to nick DNA
non-specifically. The ATCC14579 genome encodes an active enzyme Bce14579I (GCWGC). BceSIV and
Bce14579I belong to the phospholipase D (PLD) family of endonucleases that are widely
distributed among Bacteria and Archaea. A survey of Type II and III R-M genes is presented
from sequenced B. cereus, B. anthracis, and B. thuringiensis strains.

<>

<1>Xu, S.Y., Samuelson, J., Pelletier, J., Sibley, M., Wilson, G.G.
<2>Method for cloning and expression of BstYI restriction endonuclease and BstYI methylase in E. coli and method for purifying BstYI and M.BstYI enzymes.
<3>Japanese Patent Office
<4>JP 2002320493 A
<5>
<6>2002
<7>
<>

<1>Xu, S.Y., Sibley, M., Samuelson, J., Wilson, G., Pelletier, J.
<2>Method for cloning and expression of BstYI restriction endonuclease and BstYI methylase in E. coli and purification of BstYI and M.BstYI enzymes.
<3>European Patent Office
<4>EP 1225219 A
<5>
<6>2002
<7>The present invention relates to recombinant DNA which encodes the BstYI restriction
endonuclease as well as BstYI methyltransferase, expression of BstYI restriction endonuclease
and M.BstYI in E. coli cells containing the recombinant DNA.  It also relates to methods for
purification of the recombinant BstYI restriction endonuclease and BstYI methyltransferase.

<>

<1>Xu, S.Y., Xiao, J.-p.
<2>Method for cloning and producing the BslI restriction enodnuclease in E. coli.
<3>International Patent Office
<4>WO 9920744
<5>
<6>1999
<7>The methylase selection method was used to clone the BslI methylase gene bslIM from Bacillus
species.  A partially active BslI methylase lacking the 17 amino acid residues at the
N-terminus was cloned in E. coli using expression vector pRRS.  Inverse PCR was used to clone
the missing portion of the BslI methylase.  After cloning the complete BslI methylase gene and
its upstream DNA sequences, an RadC homolog was found upstream of the BslI methylase.  Because
methylase gene and restriction endonuclease gene are located in proximity to each other in a
particular restriction-modification system, efforts were made to clone the downstream DNA by
inverse PCR.  After two rounds of inverse PCR reactions two open reading frames (ORF) were
found downstream of the BslI methylase gene.  Expression of the first ORF (ORF1) in a T7
expression vector did not yield any active BslI endonuclease.  Expression of the second ORF
(ORF2) in E. coli and assay of the crude cell extract indicated that this gene product has DNA
nicking activity.  The gene product of ORF2 alone does not constitute BslI endonuclease
activity.  Expression of ORF1 and ORF2 in the same (E. coli) cell produces BslI endonuclease
activity.  BslI endonuclease activity can be reconstituted in vitro by mixing gene product of
ORF1 and ORF2 together.

<>

<1>Xu, S.Y., Xiao, J.I.
<2>Method for cloning and expression of the NheI restriction endonuclease in E. Coli.
<3>European Patent Office
<4>EP 1721984 A
<5>
<6>2006
<7>The present invention relates to recombinant DNA which encodes the NheI restriction
endonuclease as well as NheI methyltransferase; expression of NheI restriction endonuclease
from E. coli cells containing the recombinant DNA.  NheI restriction endonuclease is found in
the strain of Neisseria mucose heidelbergensis (ATCC 25999).  It cleaves double-stranded DNA
G/CTAGC to generate a 4-base 5' overhanging ends.

<>

<1>Xu, S.Y., Xiao, J.P.
<2>Method for cloning and expression of NheI restriction endonuclease in Escherichia coli.
<3>Japanese Patent Office
<4>JP 2001197889 A
<5>
<6>2001
<7>
<>

<1>Xu, S.Y., Xiao, J.P., Zhu, Z.
<2>Method For Cloning And Expression Of BpmI Restriction Endonuclease In E. coli.
<3>Japanese Patent Office
<4>JP 2002315587 A
<5>
<6>2002
<7>
<>

<1>Xu, S.Y., Zhou, J., Hume, A., Pelletier, J., Dore, A.
<2>Method For Cloning And Expression of BsmBI Restriction Endonuclese And BsmBI Methylase In E. coli And Purification Of BsmBI.
<3>Japanese Patent Office
<4>JP 2003230390 A
<5>
<6>2003
<7>
<>

<1>Xu, S.Y., Zhu, Z., Zhang, P., Chan, S.H., Samuelson, J.C., Xiao, J., Ingalls, D., Wilson, G.G.
<2>Discovery of natural nicking endonucleases Nb.BsrDI and Nb.BtsI and engineering of top-strand nicking variants from BsrDI and BtsI.
<3>Nucleic Acids Res.
<4>35
<5>4608-4618
<6>2007
<7>BsrDI and BtsI restriction endonucleases recognize and cleave double-strand DNA at the
sequences GCAATG (2/0) and GCAGTG (2/0),
respectively. We have purified and partially characterized these two
enzymes, and analyzed the genes that encode them. BsrDI and BtsI are
unusual in two respects: each cleaves DNA as a heterodimer of one large
subunit (B subunit) and one small subunit (A subunit); and, in the absence
of their small subunits, the large subunits behave as sequence-specific
DNA nicking enzymes and only nick the bottom strand of the sequences at
these respective positions: GCAATG (-/0) and GCAGTG (-/0). We refer to the
single subunit, the bottom-strand nicking forms as 'hemidimers'. Amino
acid sequence comparisons reveal that BsrDI and BtsI belong to a family of
restriction enzymes that possess two catalytic sites: a canonical
PD-X(n)-EXK and a second non-canonical PD-X(n)-E-X(12)-QR. Interestingly,
the other family members, which include BsrI (ACTGG 1/-1) and
BsmI/Mva1269I (GAATGC 1/-1) are single polypeptide chains, i.e. monomers,
rather than heterodimers. In BsrDI and BtsI, the two catalytic sites are
found in two separate subunits. Site-directed mutagenesis confirmed that
the canonical catalytic site located at the N-terminus of the large
subunit is responsible for the bottom-strand cleavage, whereas the
non-canonical catalytic site located in the small subunit is responsible
for hydrolysis of the top strand. Top-strand specific nicking variants,
Nt.BsrDI and Nt.BtsI, were successfully engineered by combining the
catalytic-deficient B subunit with wild-type A subunit.

<>

<1>Xu, T., Liang, J., Chen, S., Wang, L., He, X., You, D., Wang, Z., Li, A., Xu, Z., Zhou, X., Deng, Z.
<2>DNA phosphorothioation in Streptomyces lividans: mutational analysis of the dnd locus.
<3>BMC Microbiol.
<4>9
<5>41
<6>2009
<7>BACKGROUND: A novel DNA phosphorothioate modification (DNA sulfur modification),  in which one
of the non-bridging oxygen atoms in the phosphodiester bond linking
DNA nucleotides is exchanged by sulphur, was found to be genetically determined
by dnd or dnd-counterpart loci in a wide spectrum of bacteria from diverse
habitats. A detailed mutational analysis of the individual genes within the dnd
locus in Streptomyces lividans responsible for DNA phosphorothioation was
performed and is described here. It should be of great help for the mechanistic
study of this intriguing system. RESULTS: A 6,665-bp DNA region carrying just
five ORFs (dndA-E) was defined as the sole determinant for modification of the
DNA backbone in S. lividans to form phosphorothioate. This provides a
diagnostically reliable and easily assayable Dnd (DNA degradation) phenotype.
While dndA is clearly transcribed independently, dndB-E constitute an operon, as
revealed by RT-PCR analysis. An efficient mutation-integration-complementation
system was developed to allow for detailed functional analysis of these dnd
genes. The Dnd- phenotype caused by specific in-frame deletion of the dndA, C, D,
and E genes or the enhanced Dnd phenotype resulting from in-frame deletion of
dndB could be restored by expression vectors carrying the corresponding dnd
genes. Interestingly, overdosage of DndC or DndD, but not other Dnd proteins, in
vivo was found to be detrimental to cell viability. CONCLUSION: DNA
phosphorothioation is a multi-enzymatic and highly coordinated process controlled
by five dnd genes. Overexpression of some proteins in vivo prevented growth of
host strain, suggesting that expression of the gene cluster is strictly regulated
in the native host.

<>

<1>Xu, T., Yao, F., Zhou, X., Deng, Z., You, D.
<2>A novel host-specific restriction system associated with DNA backbone S-modification in Salmonella.
<3>Nucleic Acids Res.
<4>38
<5>7133-7141
<6>2010
<7>A novel, site-specific, DNA backbone S-modification (phosphorothioation) has been discovered,
but its in vivo function(s) have remained obscure.
Here, we report that the enteropathogenic Salmonella enterica serovar
Cerro 87, which possesses S-modified DNA, restricts DNA isolated from
Escherichia coli, while protecting its own DNA by site-specific
phosphorothioation. A cloned 15-kb gene cluster from S. enterica conferred
both host-specific restriction and DNA S-modification on E. coli.
Mutational analysis of the gene cluster proved unambiguously that the
S-modification prevented host-specific restriction specified by the same
gene cluster. Restriction activity required three genes in addition to at
least four contiguous genes necessary for DNA S-modification. This
functional overlap ensures that restriction of heterologous DNA occurs
only when the host DNA is protected by phosphorothioation. Meanwhile, this
novel type of host-specific restriction and modification system was
identified in many diverse bacteria. As in the case of
methylation-specific restriction systems, targeted inactivation of this
gene cluster should facilitate genetic manipulation of these bacteria, as
we demonstrate in Salmonella.

<>

<1>Xu, W.L., Muller, S.J.
<2>Exploring both sequence detection and restriction endonuclease cleavage kinetics by recognition site via single-molecule microfluidic trapping.
<3>Lab on a Chip, Royal Soc. Chemistry, , Cambridge
<4>11
<5>435-442
<6>2011
<7>We demonstrate the feasibility of a single-molecule microfluidic approach to both sequence
detection and obtaining kinetic information
for restriction endonucleases on dsDNA. In this method, a microfluidic
stagnation point flow is designed to trap, hold, and linearize
double-stranded (ds) genomic DNA to which a restriction endonuclease
has been pre-bound sequence-specifically. By introducing the cofactor
magnesium, we determine the binding location of the enzyme by the
cleavage process of dsDNA as in optical restriction mapping, however
here the DNA need not be immobilized on a surface. We note that no
special labeling of the enzyme is required, which makes it simpler than
our previous scheme using stagnation point flows for sequence
detection. Our accuracy in determining the location of the recognition
site is comparable to or better than other single molecule techniques
due to the fidelity with which we can control the linearization of the
DNA molecules. In addition, since the cleavage process can be followed
in real time, information about the cleavage kinetics, and subtle
differences in binding and cleavage frequencies among the recognition
sites, may also be obtained. Data for the five recognition sites for
the type II restriction endonuclease EcoRI on lambda-DNA are presented
as a model system. While the roles of the varying fluid velocity and
tension along the chain backbone on the measured kinetics remain to be
determined, we believe this new method holds promise for a broad range
of studies of DNA-protein interactions, including the kinetics of other
DNA cleavage processes, the dissociation of a restriction enzyme from
the cleaved substrate, and other macromolecular cleavage processes.

<>

<1>Xu, X., Jiang, L., Zhang, Z., Shi, Y., Huang, H.
<2>Genome Sequence of a Gamma- and UV-Ray-Resistant Strain, Deinococcus wulumuqiensis R12.
<3>Genome Announcements
<4>1
<5>e00206-13
<6>2013
<7>Deinococcus wulumuqiensis R12, isolated from radiation-polluted soil, is a red-pigmented
strain of the extremely radioresistant genus Deinococcus. It
contains a major carotenoid, namely, deinoxanthin. Here, we present a 3.39-Mb
assembly of its genome sequence, which might provide various kinds of useful
information related to Deinococcus, such as about the key enzymes of its
radioresistance mechanism and carotenoid biosynthetic pathways.

<>

<1>Xu, Y., Kersten, R.D., Nam, S.J., Lu, L., Al-Suwailem, A.M., Zheng, H., Fenical, W., Dorrestein, P.C., Moore, B.S., Qian, P.Y.
<2>Bacterial Biosynthesis and Maturation of the Didemnin Anti-cancer Agents.
<3>J. Am. Chem. Soc.
<4>134
<5>8625-8632
<6>2012
<7>The anti-neoplastic agent didemnin B from the Caribbean tunicate Trididemnum
solidum was the first marine drug to be clinically tested in humans. Because of
its limited supply and its complex cyclic depsipeptide structure, considerable
challenges were encountered during didemnin B's development that continue to
limit aplidine (dehydrodidemnin B), which is currently being evaluated in
numerous clinical trials. Herein we show that the didemnins are bacterial
products produced by the marine alpha-proteobacteria Tistrella mobilis and
Tistrella bauzanensis via a unique post-assembly line maturation process.
Complete genome sequence analysis of the 6,513,401 bp T. mobilis strain
KA081020-065 with its five circular replicons revealed the putative didemnin
biosynthetic gene cluster (did) on the 1,126,962 bp megaplasmid pTM3. The did
locus encodes a 13-module hybrid non-ribosomal peptide synthetase-polyketide
synthase enzyme complex organized in a collinear arrangement for the synthesis of
the fatty acylglutamine ester derivatives didemnins X and Y rather than didemnin
B as first anticipated. Imaging mass spectrometry of T. mobilis bacterial
colonies captured the time-dependent extracellular conversion of the didemnin X
and Y precursors to didemnin B, in support of an unusual post-synthetase
activation mechanism. Significantly, the discovery of the didemnin biosynthetic
gene cluster may provide a long-term solution to the supply problem that
presently hinders this group of marine natural products and pave the way for the
genetic engineering of new didemnin congeners.

<>

<1>Xu, Y., Liu, Y., Yao, S., Li, J., Cheng, C.
<2>Genome Sequence of Paenibacillus polymyxa Strain CICC 10580, Isolated from the Fruit of Noni (Morinda citrifolia L.) Grown in the Paracel Islands.
<3>Genome Announcements
<4>2
<5>e00854-14
<6>2014
<7>Noni is a plant reported to have nutritional and therapeutic properties. Paenibacillus
polymyxa CICC 10580 is a strain that was isolated from the fruit of
noni and showed comprehensive antagonistic activity against many pathogens. Its
genome was sequenced and assembled (6.10 Mb). The coding sequences (CDSs)
correlated with antagonistic activity were annotated.

<>

<1>Xu, Y., Lunnen, K.D., Kong, H.
<2>Engineering a nicking endonuclease N.AlwI by domain swapping.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>98
<5>12990-12995
<6>2001
<7>Changing enzymatic function through genetic engineering still presents a challenge to
molecular biologists.  Here we present an example in which changing the oligomerization state
of an enzyme changes its function.  Type IIs restriction endonucleases such as AlwI usually
fold into two separate domains: A DNA-binding domain and a catalytic/dimerization domain.  We
have swapped the putative dimerization domain of AlwI with a nonfunctional dimerization domain
from a nicking enzyme, N.BstNBI.  The resulting chimeric enzyme, N.AlwI, no longer forms a
dimer.  Interestingly, the monomeric N.AlwI still recognizes the same sequence as AlwI but
only cleaves the DNA strand containing the sequence 5'-GGATC-3' (top strand).  In contrast,
the wild-type AlwI exists as a dimmer in solution and cleaves two DNA strands; the top strand
is cleaved by an enzyme binding to that sequence, and its complementary bottom strand is
cleaved by the second enzyme dimerized with the first enzyme.  N.AlwI is unable to form a
dimer and therefore nicks DNA as a monomer.  In addition, the engineered nicking enzyme is at
least as active as the wild-type AlwI and is thus a useful enzyme.  To our knowledge, this is
the first report of creating a nicking enzyme by domain swapping.

<>

<1>Xu, Y., Wang, A., Tao, F., Su, F., Tang, H., Ma, C., Xu, P.
<2>Genome Sequence of Enterobacter cloacae subsp. dissolvens SDM, an Efficient Biomass-Utilizing Producer of Platform Chemical 2,3-Butanediol.
<3>J. Bacteriol.
<4>194
<5>897-898
<6>2012
<7>Enterobacter cloacae subsp. dissolvens SDM has an extraordinary characteristic of biomass
utilization for 2,3-butanediol production. Here
we present a 4.9-Mb assembly of its genome. The key genes for regulation
and metabolism of 2,3-butanediol production were annotated, which could
provide further insights into the molecular mechanism of high-yield
production of 2,3-butanediol.

<>

<1>Xu, Z. et al.
<2>Genome Biology of Actinobacillus pleuropneumoniae JL03, an Isolate of Serotype 3 Prevalent in China.
<3>PLoS ONE
<4>3
<5>e1450
<6>2008
<7>Actinobacillus pleuropneumoniae is the etiologic agent of porcine contagious pleuropneumonia,
a cause of considerable world wide economic
losses in the swine industry. We sequenced the complete genome of A.
pleuropneumoniae, JL03, an isolate of serotype 3 prevalent in China. Its
genome is a single chromosome of 2,242,062 base pairs containing 2,097
predicted protein-coding sequences, six ribosomal rRNA operons, and 63
tRNA genes. Preliminary analysis of the genomic sequence and the functions
of the encoded proteins not only confirmed the present physiological and
pathological knowledge but also offered new insights into the metabolic
and virulence characteristics of this important pathogen. We identified a
full spectrum of genes related to its characteristic chemoheterotrophic
catabolism of fermentation and respiration with an incomplete TCA system
for anabolism. In addition to confirming the lack of ApxI toxin,
identification of a nonsense mutation in apxIVA and a 5'-proximal
truncation of the flp operon deleting both its promoter and the
flp1flp2tadV genes have provided convincing scenarios for the low
virulence property of JL03. Comparative genomic analysis using the
available sequences of other serotypes, probable strain
(serotype)-specific genomic islands related to capsular polysaccharides
and lipopolysaccharide O-antigen biosyntheses were identified in JL03,
which provides a foundation for future research into the mechanisms of
serotypic diversity of A. pleuropneumoniae.

<>

<1>Xu, Z., Chen, X., Li, L., Li, T., Wang, S., Chen, H., Zhou, R.
<2>Comparative genomic characterization of Actinobacillus pleuropneumoniae.
<3>J. Bacteriol.
<4>192
<5>5625-5636
<6>2010
<7>The Gram-negative bacterium Actinobacillus pleuropneumoniae is the
etiologic agent of porcine contagious pleuropneumoniae, a lethal
respiratory infectious disease causing great economic losses in the swine
industry worldwide. In order to better interpret the genetic background of
serotypic diversity, nine genomes of A. pleuropneumoniae reference strains
of serovars 1, 2, 4, 6, 9, 10, 11, 12, and 13 were sequenced by using
rapid high-throughput approach. Based on 12 genomes of corresponding
serovar reference strains including three publicly available complete
genomes (serovars 3, 5b, and 7) of this bacterium, we performed a
comprehensive analysis of comparative genomics and first reported a global
genomic characterization for this pathogen. Clustering of 26,012 predicted
protein-coding genes showed that the pan genome of A. pleuropneumoniae
consists of 3,303 gene clusters, which contain 1,709 core genome genes,
822 distributed genes, and 772 strain-specific genes. The genome
components involved in the biogenesis of capsular polysaccharide and
lipopolysaccharide O antigen relative to serovar diversity were compared,
and their genetic diversity was depicted. Our findings shed more light on
genomic features associated with serovar diversity of A. pleuropneumoniae
and provide broader insight into both pathogenesis research and
clinical/epidemiological application against the severe disease caused by
this swine pathogen.

<>

<1>Xu, Z., Xia, J., Feng, X., Li, S., Xu, H., Bo, F., Sun, Z.
<2>Genome Sequence of Streptomyces albulus PD-1, a Productive Strain for Epsilon-Poly-L-Lysine and Poly-L-Diaminopropionic Acid.
<3>Genome Announcements
<4>2
<5>e00297-14
<6>2014
<7>Streptomyces albulus PD-1, a productive strain for epsilon-poly-l-lysine and
poly-l-diaminopropionic acid, was isolated from soils. We present the genome sequence of S.
albulus PD-1, which may provide abundant information regarding the production of
epsilon-poly-l-lysine and poly-l-diaminopropionic acid.

<>

<1>Xu, Z., Yin, H., Tian, Z., Zhou, Y., Ai, S.
<2>Electrochemical immunoassays for the detection the activity of DNA methyltransferase by using the rolling circle amplification technique.
<3>Microchimica Acta
<4>181
<5>471-477
<6>2014
<7>We report on an electrochemical method for the determination of the activity of the enzyme
methyltransferase (MTase). The methyl-binding domain-1 protein was applied to recognize
symmetrically methylated cytosine in CpG (-C-phosphate-G-) islands of ds-DNA which then
specifically bind to anti-His tag antibody. Hyperbranched rolling circle amplification (RCA)
was used to improve sensitivity. When the dsDNA was treated with M.Sss I methyltransferase,
the sequence 5'-CCGG-3' was methylated and recognize by the methyl binding protein. In turn,
the anti-His tag, biotinylated IgG, streptavidin and biotinylated oligonucleotide were
captured successively on the surface of an electrode. Subsequently, the RCA reaction was
initiated and streptavidin-labeled alkaline phosphatase immobilized on the surface of the
electrode. ALP was able to catalyze the hydrolysis of 1-naphthyl phosphate to form 1-naphthol
at pH 9.8. The oxidation peak current of 1-naphthol was used to monitor the methylation
process. The response  obtained by differential pulse voltammetry was linearly related to the
concentration of M.Sss I MTase in the range from 0.1 to 40 unit mL(-1), and the detection
limit was 0.03 unit mL(-1) (at an SNR of 3). The inhibitory action of paclitaxel on the
activity of M.Sss I MTase also was investigated.

<>

<1>Xu, Z.H., Han, D.M., Cao, J.J., Saini, U.
<2>Cloning and characterization of the TneDI restriction-modification system of Thermotoga neapolitana.
<3>Extremophiles
<4>15
<5>665-672
<6>2011
<7>A putative Type II restriction-modification system of Thermotoga neapolitana, TneDI, was
cloned into Escherichia coli XL1-Blue MRF' and
characterized. Gene CTN 0339 specifies the endonuclease R.TneDI, while
CTN 0340 encodes the cognate DNA methyltransferase M.TneDI. Both
enzymes were purified simply by heating the cell lysates of E. coli
followed by centrifugation. The enzymes were active over a broad range
of temperatures, from 42A degrees C to at least 77A degrees C, with the
highest activities observed at 77A degrees C. R.TneDI cleaved at the
center of the recognition sequence (CGa dagger'CG) and generated
blunt-end cuts. Overexpression of R.TneDI in BL21(DE3) was confirmed by
both SDS-PAGE and Western blotting.

<>

<1>Xue, C., Cao, L., Zhang, R., He, J., Li, S., Hong, Q.
<2>Draft Genome Sequence of Sphingobium sp. Strain BHC-A, Revealing Genes for the Degradation of Hexachlorocyclohexane.
<3>Genome Announcements
<4>2
<5>e00254-14
<6>2014
<7>Here, we report the draft genome sequence of Sphingobium sp. strain BHC-A, a lin  gene-based
hexachlorocyclohexane (HCH)-degrading strain, isolated from soil that suffered long-term HCH
contamination in an insecticide factory.

<>

<1>Xue, J., He, Z.
<2>Enzymology of BamHI DNA methylase.
<3>Shengwu Huaxue Zazhi
<4>7
<5>13-20
<6>1991
<7>The enzymology of BamHI DNA methylase has been studied.  The optimum pH is 7.8.
It is unstable in heat treatment and dependent on K+ or Na+.  The Km is 7.69 x
10-6 mol/L for SAM and the Ki is 7.33 x 10-4 mol/L in the presence of 1 mmol/L
SAH by Lineweaver-Burke plot.  The two lines intersected at the ordinate on L-B
plot.  This result shows that SAH is the competitive inhibitor for BamHI DNA
methylase.  The study of substrate specificity for different DNA shows that the
single-stranded M.L. DNA is the best substrate for this enzyme.  The enzyme
activity was assayed in the presence of different metabolites such as cytosine,
5-azacytosine, adenosine, 6-azacytosine, 5-azacytidine and 5-methylcytosine.
The enzyme activity could be inhibited by divalent cations such as Mn2+, Mg2+,
Zn2+, but was stimulated strongly by F-, WO3-/4, MoO2-/4.  Results from the
chemical modification by iodoacetamide and protection by DTT or
2-mercaptoethanol of this enzyme showed that the SH group is essential in
active site of this enzyme.  It has been proved by HPLC that this enzyme
transferred the methyl group from SAM to cytosine of DNA.  The methylation
level of Micrococcus luteus (M.L.) DNA is 2.39% when it was incubated with
BamHI DNA methylase for 30'.  It has been proved by electrophoresis on 1%
agarose that the BamHI DNA methylase modified DNA cannot be cleaved by BamHI
restriction endonuclease.

<>

<1>Xydas, S., Lange, C.S., Phil, D., Neimark, H.C.
<2>Effect of methylation on the electrophoretic mobility of chromosomal DNA in pulse-field agarose gels.
<3>Appl. Theor. Electrophor.
<4>6
<5>43-47
<6>1996
<7>Factors other than molecular weight are known to affect DNA electrophoretic mobility.  DNA
methylation has been found to affect the curvature of DNA, causing anomalous mobility in
polyacrylamide gels; the effect of methylation on the mobility of large DNA molecules in
agarose gels was unknown.  Chromosomal DNA from Mycoplasma capricolum, a wall-less prokaryote
which has a low intrinsic methylation rate, was methylated in agarose blocks by SssI
methylase, a de novo methylase with a CpG recognition sequence.  (A surprising finding was
that SssI methylase altered the structure of InCert, but not SeaKem Gold, agarose.)
Restriction enzyme analysis was used to estimate the extent of CpG methylation.  DNA
methylation was found to have no effect on the electrophoretic mobility of full-length
chromosomal DNA (1,120 kbp) in agarose gels.  Therefore, methylation is not a source of error
in PFGE-based size estimation for chromosomal DNA molecules less than 1.12 Mbp in agarose
gels.

<>

<1>Yagubi, A.I., Castle, A.J., Kropinski, A.M., Banks, T.W., Svircev, A.M.
<2>Complete Genome Sequence of Erwinia amylovora Bacteriophage vB_EamM_Ea35-70.
<3>Genome Announcements
<4>2
<5>e00413-14
<6>2014
<7>The complete genome of an Erwinia amylovora bacteriophage, vB_EamM_Ea35-70 (Ea35-70), is
271,084 bp, encodes 318 putative proteins, and contains one tRNA.
Comparative analysis with other Myoviridae genomes suggests that Ea35-70 is
related to the Phikzlikevirus genus within the family Myoviridae, since 26% of
Ea35-70 proteins share homology to proteins in Pseudomonas phage phiKZ.

<>

<1>Yahara, K., Furuta, Y., Oshima, K., Yoshida, M., Azuma, T., Hattori, M., Uchiyama, I., Kobayashi, I.
<2>Chromosome painting in silico in a bacterial species reveals fine population structure.
<3>Mol. Biol. Evol.
<4>30
<5>1454-1464
<6>2013
<7>Identifying population structure forms an important basis for genetic and evolutionary
studies. Most current methods to identify population structure have limitations in analyzing
haplotypes and recombination across the genome. Recently, a method of chromosome painting in
silico has been developed to overcome these shortcomings and has been applied to multiple
human genome sequences. This method detects the
genome-wide transfer of DNA sequence chunks through homologous recombination.
Here, we apply it to the frequently recombining bacterial species Helicobacter pylori, which
has infected Homo sapiens since their birth in Africa and shows wide phylogeographic
divergence. Multiple complete genome sequences were analyzed including sequences from Okinawa,
Japan, that we recently sequenced. The newer method revealed a finer population structure than
revealed by a previous method that examines only MLST housekeeping genes or a phylogenetic
network analysis of the core genome. Novel subgroups were found in Europe, Amerind and East
Asia groups.  Examination of genetic flux showed some singleton strains to be hybrids of
subgroups and revealed evident signs of population admixture in Africa, Europe, and parts of
Asia. We expect this approach to further our understanding of intraspecific bacterial
evolution by revealing population structure at a finer scale.

<>

<1>Yahara, K., Horie, R., Kobayashi, I., Sasaki, A.
<2>Evolution of DNA double-strand break repair by gene conversion: Coevolution between a phage and a restriction-modification system.
<3>Genetics
<4>176
<5>513-526
<6>2007
<7>The necessity to repair genome damage has been considered to be an immediate factor
responsible for the origin of sex. Indeed, attack by a
cellular restriction enzyme of invading DNA from several bacteriophages
initiates recombinational repair by gene conversion if there is
homologous DNA. In this work, we modeled the interaction between a
bacteriophage and a bacterium carrying a restriction enzyme as
antagonistic coevolution. We assume a locus on the bacteriophage genome
has either a restriction-sensitive or a restriction-resistant allele,
and another locus determines whether it is recombination/repair
proficient or defective. A restriction break can be repaired by a
co-infecting phage genome if one of them is recombination/repair
proficient. We define the fitness of phage (resistant/sensitive and
repair-positive/-negative) genotypes and bacterial
(restriction-positive/-negative) genotypes by assuming random encounter
of the genotypes, with given probabilities of single and double
infections, and the costs of resistance, repair, and restriction. Our
results show the evolution of the repair allele depends on b(1)/b(0),
the ratio of the burst size b, under damage to host cell physiology
induced by an unrepaired double-strand break to the default burst size
b(0). It was not until this effect was taken into account that the
evolutionary advantage of DNA repair became apparent.

<>

<1>Yahara, K., Kobayashi, I.
<2>Evolution of selfish homing endonuclease genes in the absence of horizontal transfer.
<3>Saibo Kogaku
<4>29
<5>190-195
<6>2010
<7>
<>

<1>Yaharaa, K., Fukuyo, M., Sasaki, A., Kobayashi, I.
<2>Evolutionary maintenance of selfish homing endonuclease genes in the absence of horizontal transfer.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>106
<5>18861-18866
<6>2009
<7>Homing endonuclease genes are ``selfish' mobile genetic elements whose endonuclease promotes
the spread of its own gene by creating a break at
a specific target site and using the host machinery to repair the break
by copying and inserting the gene at this site. Horizontal transfer
across the boundary of a species or population within which mating
takes place has been thought to be necessary for their evolutionary
persistence. This is based on the assumption that they will become
fixed in a host population, where opportunities of homing will
disappear, and become susceptible to degeneration. To test this
hypothesis, we modeled behavior of a homing endonuclease gene that
moves during meiosis through double-strand break repair. We
mathematically explored conditions for persistence of the homing
endonuclease gene and elucidated their parameter dependence as phase
diagrams. We found that, if the cost of the pseudogene is lower than
that of the homing endonuclease gene, the 2 forms can persist in a
population through autonomous periodic oscillation. If the cost of the
pseudogene is higher, 2 types of dynamics appear that enable
evolutionary persistence: bistability dependent on initial frequency or
fixation irrespective of initial frequency. The prediction of long
persistence in the absence of horizontal transfer was confirmed by
stochastic simulations in finite populations. The average time to
extinction of the endonuclease gene was found to be thousands of
meiotic generations or more based on realistic parameter values. These
results provide a solid theoretical basis for an understanding of these
and other extremely selfish elements. POPULATION BIOLOGY

<>

<1>Yamada, T., Chuchird, N., Kawasaki, T., Nishida, K., Hiramatsu, S.
<2>Chlorella viruses as a source of novel enzymes.
<3>J. Biosci. Bioeng.
<4>88
<5>353-361
<6>1999
<7>A special advantage has been conferred upon Chlorella cells as tools in biotechnology when
viruses (Phycodnaviridae) infecting Chlorella cells
were discovered and isolated. The viruses are large icosahedral
particles (150-200 nm in diameter), containing a giant, 330-380 kbp
long, linear dsDNA genome. Recently, the nucleotide sequence of the
330,740-bp genome of PBCV-1, the prototype virus of Phycodnaviridae,
was determined, and up to 702 open reading frames (ORFs) were
identified along the genome. The possible genes present include those
encoding a variety of enzymes involved in the modification of DNA, RNA,
protein and polysaccharides as well as those involved in the metabolism
of sugars, amino acids, lipids, nucleotides and nucleosides. Many of
these genes are actually expressed during viral infection, with
functional enzymes detected in the host cytoplasm or incorporated into
the virion. The successful utilization of these viral enzymes as
various DNA restriction and modification enzymes (Cvi enzymes) that are
now commercially available is well documented. Also noteworthy are
virion-associated chitinase and chitosanase activities that have
potentially important applications in the recycling of natural
resources. The virions of Chlorella viruses contain more than 50
different structural proteins, ranging in size from 10 to 200 kDa. Some
of these proteins may be replaced with useful foreign proteins using
recombinant DNA technology. The proteins of interest can be recovered
easily from the viral particles, and collected by centrifugation after
complete lysis of the host Chlorella cells.

<>

<1>Yamada, Y., Mizuno, H., Sato, H., Akagawa, M., Yamasato, K.
<2>A new restriction endonuclease from Agrobacterium gelatinovorum, a marine agrobacterium (AgeI).
<3>Agric. Biol. Chem.
<4>53
<5>1747-1749
<6>1989
<7>Type II restriction endonucleases, which are indispensable for gene analyses
and gene manipulation, have been recovered from prokaryotic cells such as those
of bacteria and cyanobacteria.  We have been studying acetic acid bacteria from
taxonomical, physiological and biochemical points of view.  The new restriction
endonuclease ApaLI was reported in Acetobacter pasteurianus IFO 13753.  During
the course of our screening of marine bacteria, we found a new restriction
endonuclease, designated as AgeI, in Agrobacterium gelatinovorum IAM 12617.
This paper describes its purification and properties and determination of its
recognition sequence and cleavage site.

<>

<1>Yamada, Y., Mizuno, H., Yamasato, K.
<2>New restriction enzyme and procedure for its isolation.
<3>German Patent Office
<4>DE 4010108 A
<5>
<6>1990
<7>*
A new restriction enzyme and a procedure for the production is given. The restriction enzyme
AgeI has the following properties:

a) Way of working and substrate specificity; Recognizes the base-sequence in the following
double stranded deoxyribonucleic acid molecule and cleaves at the positions which are marked
with arrows: A^CCGGT.

b) optimal pH: 7.5
c) stable between pH 5.0-8.0
d) optimal temp: 30oC
e) temperature stability: 45oC (when heating for at least 5 min)
f) stable salt concentration: 0-150 mM NaCl
g) molecular weight: 23000, as determined by gel filtration and 24000, determined by SDS-PAGE.


<>

<1>Yamada, Y., Murakami, M.
<2>New restriction enzyme and process for producing the same.
<3>US Patent Office
<4>US 4833082
<5>2594
<6>1989
<7>A restriction enzyme, ApaLI, which recognizes the base sequence in a double-stranded
deoxyribonucleic acid molecule as shown below, and cleaves it at the arrow-marked sites:
5'-G/TGCAC-3'
3'-CACGT/G-3'
wherein A, G, T and C represent adenosine, guanosine, thymidine and cytidine, respectively.
Also provided is a process for producing this enzyme, by growing a micro-organism belonging to
the genus Acetobacter.

<>

<1>Yamada, Y., Murakami, M.
<2>Restriction enzyme produced by Acetobactor.
<3>United Kingdom Patent Office
<4>GB 2184125
<5>
<6>1988
<7>The enzyme, designated ApaLI, recognizes the base sequence below and cleaves at the positions
marked with arrows: 5'-G/TGCAC-3' 3'-CACGT/G-5'. The optimum pH range for the enzyme is
7.5 to 8.5, the stable pH range is 7.0 to 9.0 and the optimum temperature is approximately
37oC. Its molecular weight is 26,000 +/- 8,000, as determined by gel filtration. The enzyme is
produced by growing a microorganism belonging to the genus Acetobactor and which is capable of
producing the enzyme ApaLI in a culture medium and subsequently collecting the enzyme thus
formed from the culture broth.

<>

<1>Yamada, Y., Murakami, M.
<2>New restriction enzyme and process for producing the same.
<3>German Patent Office
<4>DE 3631152
<5>2594
<6>1989
<7>A restriction enzyme, ApaLI, which recognizes the base sequence in a double-stranded
deoxyribonucleic acid molecule as shown below, and cleaves it at the arrow-marked sites:
5'-G/TGCAC-3'
3'-CACGT/G-3'
wherein A, G, T and C represent adenosine, guanosine, thymidine and cytidine, respectively.
Also provided is a process for producing this enzyme, by growing a micro-organism belonging to
the genus Acetobacter.


<>

<1>Yamada, Y., Murakami, M.
<2>A new restriction endonuclease from Acetobacter pasteurianus (ApaI).
<3>Agric. Biol. Chem.
<4>49
<5>3627-3629
<6>1985
<7>Type II restriction endonucleases indispensable for gene manipulation and gene
analyses have been recovered from procaryotic cells such as those of bacteria
and cyanobacteria.  We have been studying acetic acid bacteria from the
viewpoints of taxonomy, physiology, biochemistry and genetics.  During the
course of our studies, we found a new type II restriction endonuclease,
designated as ApaLI, in a strain of Acetobacter pasteurianus (Hansen 1879)
Beijerinck 1916.  This communication describes its purification and properties
and determination of its recognition sequence and cleavage sites in DNA
molecules.

<>

<1>Yamada, Y., Murakami, M., Murofushi, N., Narise, K.
<2>Type II restriction enzyme - obtained from culture of Gluconobacter genus.
<3>Japanese Patent Office
<4>JP 63068082
<5>
<6>1988
<7>(I)Type II restriction enzyme, GceinI, taken out from culture production is obtained by
culturing type II restriction enzyme Gcein I-producing bacteria of Gluconobacter genus. The
restriction enzyme recognises the base sequence in molecule of duplex deoxyribonucleic acid:
5'.G-G-A-T-C-C....3', 3'.C-C-T-A-G-G..5'.

<>

<1>Yamada, Y., Sasaki, J.
<2>Distribution of Restriction Enzymes in Strains of Acetobacter (Gluconoacetobacter) Liquefaciens.
<3>J. Gen. Appl. Microbiol.
<4>30
<5>309-312
<6>1984
<7>Acetobacter (Gluconoacetobacter) liquefaciens was first described by ASAI as a
species of the genus Gluconobacter. Three strains, G-1, AC-8, and U-4, were
reported.  Concerning the taxonomic position, these strains were designated as
pigment-producing strains of A. aceti because of their peritrichous
flagellation and ability to oxidize acetate to carbon dioxide and water.  In
contrast, Asai et al. stated their uniqueness in phenotypic features and
designated them as 'intermediate.'  Subsequently, Yamada et al. found that
these three strains and two other strains isolated by Komagata are
distinguishable from A. aceti strains by their coenzyme Q or ubiquinone system;
the five intermediate strains have Q-10[Q-9] whereas the A. aceti strains have
Q-9[Q-8].  In Bergey's Manual 8th Edition, however, these 'anomalous' strains
are classified as A. aceti subsp. liquefaciens.  The Approved Lists 1980
included A. aceti subsp. liquefaciens (Asai 1935) De Ley and Frateur 1974.
This subspecies was elevated to A. liquefaciens separate from A. aceti on the
basis of the ubiquinone system, numerical analyses of phenotypic features and
protein gel electrophoretic patterns of enzymes.  Recently, Yamada and Kondo
have set up a new subgenus, Gluconoacetobacter and classified these strains as
Acetobacter (Gluconoacetobacter) liquefaciens (Asai 1935) Gossele et al. 1983.
In previous papers, the authors have purified two restriction enzymes of A.
liquefaciens IAM 1834 and AJ 2881 to a homogeneous state and determined their
recognition sequences and cleavage sites on DNA molecules.  The enzymes have
been found to be isoschizomers of BamHI and PstI, respectively.  This paper
describes distribution of restriction enzymes in the five strains mentioned
above.

<>

<1>Yamada, Y., Sato, H.
<2>The manganese ion strongly activates restriction endonuclease AatII from Acetobacter aceti IFO 3281.
<3>Agric. Biol. Chem.
<4>53
<5>1745-1746
<6>1989
<7>Type II restriction endonucleases are indispensable for gene analyses and gene
manipulation, and their occurrence is widespread in the prokaryotic kingdom.
We have been studying restriction endonucleases of acetic acid bacteria.
During the course of our studies, restriction endonuclease ApaLI was found in
Acetobacter pasteurianus IFO 13753.  Restriction endonuclease AatII was
reported by Sugisaki et al. as a new enzyme from Acetobacter aceti IFO 3281.
However, the characteristics of the enzyme have remained unclarified as yet
except for its recognition sequence and cleavage site.  This paper reports that
the enzyme purified to a homogeneous state is strongly activated on the
addition of not Mg2+ but Mn2+.

<>

<1>Yamada, Y., Yoshioka, H., Sasaki, J., Tahara, Y.
<2>Purification, properties and recognition sequence of site-specific restriction endonuclease from "Acetobacter liquefaciens".
<3>J. Gen. Appl. Microbiol.
<4>29
<5>157-166
<6>1983
<7>A type II restriction endonuclease was purified from "Acetobacter liquefaciens"
IAM 1834 by consecutive column chromatography on heparin-Sepharose CL-6B,
DEAE-Sepharose CL-6B and Sephacryl S-400 superfine.  The purified enzyme was
homogeneous on polyacrylamide gel disc electrophoresis.  The enzyme preparation
was essentially free from other nuclease activity, as judged by constancy of a
lambda DNA-digest electrophoretic pattern after prolonged incubation for 24 hr.
The enzyme was optimally active at 37o at pH 7.5, and did not require NaCl,
which rather inhibited its activity.  The recognition sequence for the enzyme
was determined to be 5'-G-G-A-T-C-C-3', and the enzyme was found to cut between
G and G in the sequence, being an isoschizomer of the endonuclease from
"Bacillus amyloliquefaciens" H (Bam HI).

<>

<1>Yamaguchi, T., Nishifuji, K., Sasaki, M., Fudaba, Y., Aepfelbacher, M., Takata, T., Ohara, M., Komatsuzawa, H., Amagai, M., Sugai, M.
<2>Identification of the Staphylococcus aureus etd pathogenicity island which encodes a novel exfoliative toxin, ETD, and EDIN-B.
<3>Infect. Immun.
<4>70
<5>5835-5845
<6>2002
<7>We identified a novel pathogenicity island in Staphylococcus aureus which contains open
reading frames (ORFs) similar to the exfoliative
toxin (ET) gene, glutamyl endopeptidase gene, and edin-B gene in tandem
and the phage resistance gene, flanked by hsdM, hsdS (restriction and
modification system), and IS256. The protein encoded by the ET-like
gene showed 40, 59, and 68% amino acid sequence identities with
exfoliative toxin A (ETA), exfoliative toxin B (ETB), and
Staphylococcus hyicus ETB (ShETB), respectively. When injected into
neonatal mice, the recombinant protein derived from the ET-like gene
induced exfoliation of the skin with loss of cell-to-cell adhesion in
the upper part of the epidermis as observed in histological
examinations, just as was found in neonatal mice injected with ETA or
ETB. Western blot analysis indicated that the recombinant protein is
serologically distinct from ETA and ETB. Therefore, the product encoded
by this new ORF is a new ET member produced by S. aureus and is termed
ETD. ETD did not induce blisters in 1-day-old chickens. In the skins of
mice injected with ETD, cell surface staining of desmoglein 1 (Dsg1), a
cadherin type cell-to-cell adhesion molecule in desmosomes, was
abolished without affecting that of desmoglein 3 (Dsg3). Furthermore,
in vitro incubation of the recombinant extracellular domains of Dsg1
and Dsg3 with the recombinant protein demonstrated that both mouse and
human Dsg1, but not Dsg3, were directly cleaved in a dose-dependent
manner. These results demonstrate that ETD and ETA induce blister
formation by identical pathophysiological mechanisms. Clinical strains
positive for edin-B were suggested to be clonally associated, and all
edin-B-positive strains tested were positive for etd. Among 18
etd-positive strains, 12 produced ETD extracellularly. Interestingly,
these strains are mainly isolated from other sources of infections and
not from patients with bullous impetigo or staphylococcal scalded-skin
syndrome. This strongly suggests that ETD might play a pathogenic role
in a broader spectrum of bacterial infections than previously
considered.

<>

<1>Yamamoto, K., Sagawa, H., Kotani, H., Hiraoka, N., Nakamura, T.
<2>New restriction enzyme.
<3>German Patent Office
<4>DE 4112563 A
<5>
<6>1991
<7>The finding concerns a new restriction enzyme. A protocol for the industrial production and
isolation of a new restriction enzyme is given here. It requires growing the microorganism
Brevibacterium species, which is able to recognize and cleaves the nucleotide sequence at the
position marked with an arrow: C^CTAGG, where C,T,A, and G stand for Cytidine, Thymidine,
Adenosine, and Guanosine, and the isolation of this restriction enzyme out of the crude
extract.

<>

<1>Yamamoto, K., Sagawa, H., Kotani, H., Hiraoka, N., Nakamura, T.
<2>Restriction enzyme ex. Brevibacterium.
<3>United Kingdom Patent Office
<4>GB 2243153 A
<5>
<6>1990
<7>A process for producing a restriction enzyme is claimed, which comprises cultivating a
microorganism of the genus Brevibacterium which is capable of producing a restriction enzyme
(1) which cleaves the following nucleotide sequence C^CTAGG specifically at the sites marked
by the arrow. (1) is recovered from the culture broth. (1) has optimal temperature, pH, salt
concentration and MgCl2 concentrations at 37 degrees C, 7.0-8.5, 50-150 mM (NaCl) and 5-20 mM
respectively. The mol. wt. of (1) is 16,000 +/- 4,000 D from gel filtration with sephadex
G-100 and 42,000 +/- 2,000 D from SDS-PAGE. (1) is therefore dimeric, and the two subunits
have a mol. wt. of approx. 45,000 D.

<>

<1>Yamamoto, K., Sagawa, H., Kotani, H., Hiraoka, N., Nakamura, T.
<2>Restriction enzyme from Brevibacterium linens.
<3>US Patent Office
<4>US 5470732
<5>
<6>1995
<7>A restriction enzyme is obtained by cultivating a strain of Brevibacterium linens and
recovering the restriction enzyme.  The restriction enzyme, which has the same specificity as
AvrII, cleaves the following nucleotide sequence specifically at the arrow-marked sites.

<>

<1>Yamamoto, K., Tamaki, H., Cadillo-Quiroz, H., Imachi, H., Kyrpides, N., Woyke, T., Goodwin, L., Zinder, S.H., Kamagata, Y., Liu, W.T.
<2>Complete Genome Sequence of Methanolinea tarda NOBI-1T, a Hydrogenotrophic Methanogen Isolated from Methanogenic Digester Sludge.
<3>Genome Announcements
<4>2
<5>e00876-14
<6>2014
<7>Here, we report a 2.0-Mb complete genome sequence of Methanolinea tarda NOBI-1(T), a
methanogenic archaeon isolated from an anaerobic digested sludge.
This is the first genome report of the genus Methanolinea isolate belonging to
the family Methanoregulaceae, a recently proposed novel family within the order
Methanomicrobiales.

<>

<1>Yamamoto, K., Tamaki, H., Cadillo-Quiroz, H., Imachi, H., Kyrpides, N., Woyke, T., Goodwin, L., Zinder, S.H., Kamagata, Y., Liu, W.T.
<2>Complete Genome Sequence of Methanoregula formicica SMSPT, a Mesophilic Hydrogenotrophic Methanogen Isolated from a Methanogenic Upflow Anaerobic Sludge   Blanket Reactor.
<3>Genome Announcements
<4>2
<5>e00870-14
<6>2014
<7>Methanoregula formicica SMSP(T) is a mesophilic H2/formate-utilizing methanogenic archaeon and
a representative of the family Methanoregulaceae, a recently
proposed novel family within the order Methanomicrobiales. Here, we report a
2.8-Mb complete genome sequence of this methanogenic archaeon.

<>

<1>Yamamoto, Y. et al.
<2>Construction of a contiguous 874-kb sequence of the Escherichia coli -K12 genome corresponding to 50.0-68.8 min on the linkage map and analysis of its sequence features.
<3>DNA Res.
<4>4
<5>91-113
<6>1997
<7>The contiguous 874.423 base pair sequence corresponding to the 50.0-68.8 min region on the
genetic map of the Escherichia coli K-12 (W3110) was constructed by the determination of DNA
sequences in the 50.0-57.9 min region (360 kb) and two large (100 kb in all) and five short
gaps in the 57.9-68.8 min region whose sequences had been registered in the DNA databases. We
analyzed its sequence features and found that this region contained at least 894 potential
open reading frames (ORFs), of which 346 (38.7%) were previously reported, 158 (17.7%) were
homologous to other known genes, 232 (26.0%) were identical or similar to hypothetical genes
registered in databases, and the remaining 158 (17.7%) showed no significant similarity to any
other genes. A homology search of the ORFs also identified several new gene clusters. Those
include two clusters of fimbrial genes, a gene cluster of three genes encoding homologues of
the human long chain fatty acid degradation enzyme complex in the mitochondrial membrane, a
cluster of at least nine genes involved in the utilization of ethanolamine, a cluster of the
secondary set of 11 hyc genes participating in the formate hydrogenlyase reaction and a
cluster of five genes coding for the homologues of degradation enzymes for aromatic
hydrocarbons in Pseudomonas putida. We also noted a variety of novel genes, including two
ORFs, which were homologous to the putative genes encoding xanthine dehydrogenase in the fly
and a protein responsible for axonal guidance and outgrowth of the rat, mouse and nematode. An
isoleucine tRNA gene, designated ileY, was also newly identified at 60.0 min.

<>

<1>Yamamura, H., Ohnishi, Y., Ishikawa, J., Ichikawa, N., Ikeda, H., Sekine, M., Harada, T., Horinouchi, S., Otoguro, M., Tamura, T., Suzuki, K., Hoshino, Y., Arisawa, A., Nakagawa, Y., Fujita, N., Hayakawa, M.
<2>Complete genome sequence of the motile actinomycete Actinoplanes missouriensis 431(T) (= NBRC 102363(T)).
<3>Standards in Genomic Sciences
<4>7
<5>294-303
<6>2012
<7>Actinoplanes missouriensis Couch 1963 is a well-characterized member of the genus
Actinoplanes, which is of morphological interest because its members typically
produce sporangia containing motile spores. The sporangiospores are motile by
means of flagella and exhibit chemotactic properties. It is of further interest
that members of Actinoplanes are prolific sources of novel antibiotics, enzymes,
and other bioactive compounds. Here, we describe the features of A. missouriensis
431(T), together with the complete genome sequence and annotation. The 8,773,466
bp genome contains 8,125 protein-coding and 79 RNA genes.

<>

<1>Yamanaka, S.
<2>Genes with ES cell-specific expression.
<3>US Patent Office
<4>US 7250255 A
<5>
<6>2007
<7>The present invention relates to a probe for selecting ES cells, which characteristically
contains one of DNAs having base sequences depicted in SEQ ID Nos: 1, 2, 3, 4, 5, 6, 7 and 8,
or DNAs having base sequences depicted in SEQ ID Nos; 9, 11, 13, 15, 17, 19, 21, 23 and 41 and
a screening method of ES cell using this probe.  Preparation of a probe for selecting ES cells
becomes feasible by identifying plural gene with ES cell-specific expressions (ECAT genes) and
using the information of the base sequences of these gene groups.  Efficient selection of ES
cell enables supply of a large amount of ES cell expected to be applicable to regenerative
medicine.

<>

<1>Yamaoka, S., Tamura, T., Takenobu, H., Kojo, T., Tanaka, H., Inagaki, K.
<2>Cloning and nucleotide sequence of the genes encoding restriction-modification system from acidophilic bacterium Acidocella facilis 22M.
<3>Sci. Reports. Fac. Agric. Okayama Univ.
<4>92
<5>9-15
<6>2003
<7>The gene encoding the Afa22MI restriction-modification system recognizing the sequence
5'-CGATCG-3' was cloned from Acidocella facilis 22M and sequenced.  The cloned DNA fragment
contained three genes encoding the Afa22MI methylase (M.Afa22MI), the putative restriction
endonuclease Afa22MI (R.Afa22MI) and a very short patch repair endonuclease (Afa22MI vsr).  M.
Afa22MI gene has the conserved motifs of C5-cytosine methyltransferase.  Afa22MI vsr gene was
localized upstream of M.Afa22MI gene in opposite orientation, and an open reading frame of
R.Afa22MI gene was localized downstream of M.Afa22Mi gene in the same orientation.  M.Afa22MI
has about 63% sequence similarity to the entire amino acid sequence of M.XorII, and about 53%
sequence similarity to the amino acid sequence for the variable region of M.XorII.  Afa22MI
vsr has about 66% sequence similarity to the amino acid sequence of XorII vsr which was
associated with M.XorII.

<>

<1>Yamauchi, T., Moritoh, S., Johzuka-Hisatomi, Y., Ono, A., Terada, R., Nakamura, I., Iida, S.
<2>Alternative splicing of the rice OsMET1 genes encoding maintenance DNA methyltransferase.
<3>J. Plant Physiol.
<4>165
<5>1774-1782
<6>2008
<7>While the Arabidopsis genome carries one copy of the methyltransferase 1 (MET1) gene for DNA
methyltransferase, which is mainly responsible
for maintaining CpG methylation, the rice genome bears two copies of
the MET1 genes, OsMET1a and OsMET1b. The transcripts of OsMET1b
accumulate more abundantly than those of OsMET1a in all of the tissues
examined, and both genes actively transcribed at the callus, imbibed
embryo, root, meristem, young panicle, anther, pistil, and endosperm,
all of which contain actively dividing cells. The OsMET1a transcripts
contain two 5'-untranslated exons and alternatively spliced 3'-terminal
exons. The alternatively spliced transcripts consist of 14, 15, or 16
exons, and all of them encode a putative protein of 1527 amino acids.
While the 3'-terminal exon of OsMET1b is unique, alternative splicing
occurs in the T-terminal regions, which comprise either exons
containing 5'-untranslated regions or an exon bearing the initiation
codon. Depending upon alternative usage of 5' exons by alternative
splicing, the OsMET1b transcripts comprise 11, 12, 13, or 14 exons, and
the former two and the tatter two longer transcripts encode putative
proteins of 1486 and 1529 amino acids, respectively. Moreover, the 5'
splicing patterns of OsMET1b can vary in different tissues. These
findings are discussed with respect to the possible regulation of the
OsMET1 genes.

<>

<1>Yan, B.-X., Li, N., Wu, C.-X.
<2>The true recognizing sequence of the restriction enzyme whose recognizing sequence is nonpalindromic.
<3>Yichuan
<4>24
<5>420-422
<6>2002
<7>The recognizing sequence of restriction enzyme includes palindrome and nonpalindromic, and DNA
is double helix complementary strands. So the
recognizing sequences of palindrome enzyme in two strands of DNA were
identical, and can be considered of one sequence. But for
nonpalindromic restriction enzyme, the recognizing sequences of two
strands of DNA were not identical. Therefore the true recognizing
sequences are not only one. In this experiment, an enzyme cleavage
reaction was carried out which confirmed that the true recognizing
sites/sequences of non-palindromic enzyme are two instead of one.

<>

<1>Yan, J., Skoko, D., Marko, J.F.
<2>Near-field-magnetic-tweezer manipulation of single DNA molecules.
<3>Phys. Rev. E Stat. Nonlin. Soft Matter Phys.
<4>70
<5>011905
<6>2004
<7>We have developed an instrument for micromanipulation of single
DNA molecules end labeled with 3-microm-diameter paramagnetic particles. A
small, permanent magnet that can be moved as close as 10 microm to the
particle being manipulated can generate forces in excess of 200 pN,
significantly larger than obtained in other recent "magnetic-tweezer"
studies. Our instrument generates these forces in the focal plane of a
microscope objective, allowing straightforward real-time observation of
molecule extension with a position resolution of approximately 30 nm. We
show how our magnetic manipulation system can be combined with
manipulation and force measurement using glass micropipettes to allow
rapid switching between measurements in fixed-force and fixed-extension
ensembles. We demonstrate the use of our system to study formation of DNA
loops by an enzyme which strongly binds two copies of a specific
6-base-pair sequence.

<>

<1>Yan, P.-F., Ye, S.-Y., Wang, P.-Z., Li, Q.-L., Lu, Y.-Y., Zhou, B.
<2>The restriction endonucleases from Bacillus.
<3>Acta Biochim. Biophys. Sin.
<4>14
<5>151-158
<6>1982
<7>The activities of restriction endonucleases were found in 10 out of 66 strains
of Bacillus isolated and examined in our country.  These enzymes were partially
purified and their recognition sequences were characterized.  Bsp211I, Bsp226I
and Bce71I are the isoschizomers of HaeIII.  Bsp211I and HaeIII have the same
cleavage sites as indicated by the arrow in the sequence 5'-GG^CC-3'.  The
purification procedure of Bsp211I is very simple and its yield is rather high.
The recognition sequences of Bsp105I, Bsp67I, Bsp64I, Bsp74I and Bsp76I are the
same as that of MboI, which recognizes the sequence 5'-^GATC-3', the arrow
indicating the cleavage sites for Bsp 105I and Bsp67I.  While Bsp105I does not
digest modified DNA, Bsp67I digests DNA whether the adenine residue is
methylated or not.  Bsp63I and Bsp78I are isoschizomers of PstI, whose
recognition sequence is 5'-CTGCA^G-3'.  The cutting sites of Bsp 63I and PstI
are identical.

<>

<1>Yan, X.J., Xu, J., Gu, Z.H., Pan, C.M., Lu, G., Shen, Y., Shi, J.Y., Zhu, Y.M., Tang, L., Zhang, X.W., Liang, W.X., Mi, J.Q., Song, H.D., Li, K.Q., Chen, Z., Chen, S.J.
<2>Exome sequencing identifies somatic mutations of DNA methyltransferase gene DNMT3A in acute monocytic leukemia.
<3>Nat. Genet.
<4>43
<5>309-315
<6>2011
<7>Abnormal epigenetic regulation has been implicated in oncogenesis. We report here the
identification of somatic mutations by exome sequencing
in acute monocytic leukemia, the M5 subtype of acute myeloid leukemia
(AML-M5). We discovered mutations in DNMT3A (encoding DNA
methyltransferase 3A) in 23 of 112 (20.5%) cases. The DNMT3A mutants
showed reduced enzymatic activity or aberrant affinity to histone H3 in
vitro. Notably, there were alterations of DNA methylation patterns
and/or gene expression profiles (such as HOXB genes) in samples with
DNMT3A mutations as compared with those without such changes. Leukemias
with DNMT3A mutations constituted a group of poor prognosis with
elderly disease onset and of promonocytic as well as monocytic
predominance among AML-M5 individuals. Screening other leukemia
subtypes showed Arg882 alterations in 13.6% of acute myelomonocytic
leukemia (AML-M4) cases. Our work suggests a contribution of aberrant
DNA methyltransferase activity to the pathogenesis of acute monocytic
leukemia and provides a useful new biomarker for relevant cases.

<>

<1>Yan, Y., Zhao, C.W., Zhang, Y.Z., Zhang, Z.H., Pan, G.L., Liu, W.W., Ma, Q.Y., Hou, R., Tan, X.M.
<2>Draft Genome Sequence of Enterobacter cloacae subsp. cloacae Strain 08XA1, a Fecal Bacterium of Giant Pandas.
<3>J. Bacteriol.
<4>194
<5>6928-6929
<6>2012
<7>Enterobacter cloacae, a common pathogenic bacterium, is a Gram-negative bacillus. We analyzed
the draft genome of Enterobacter cloacae subsp. cloacae strain 08XA1
from the feces of a giant panda in China. Genes encoding a beta-lactamase and
efflux pumps, as well as other factors, have been found in the genome.

<>

<1>Yanagisawa, Y., Ito, E., Yuasa, Y., Maruyama, K.
<2>The human DNA methyltransferases DNMT3A and DNMT3B have two types of promoters with different CpG contents.
<3>Biochim. Biophys. Acta
<4>1577
<5>457
<6>2002
<7>DNA modification that is established by de novo methylation is involved in the epigenetic
control of genome functions. The DNMT3A and DNMT3B genes encode putative de novo
methyltransferases. In this paper, we investigated the transcriptional regulatory regions of
the human DNMT3A and DNMT3B genes. We found that the DNMT3A and DNMT3B genes have multiple
transcriptional start points (TSPs) that are separated on the chromosome and the expressions
of these genes are controlled by multiple promoters. The DNMT3A gene has at least four TSPs
and the expression is controlled by three different promoters. All three promoters lack
typical TATA sequences adjacent to the TSPs. Two of them bear CpG-rich promoters and the other
a CpG-poor promoter. The DNMT3B gene has at least two TSPs which exist in different exons and
the expression is controlled by different promoters. Both promoter regions of the DNMT3B gene
lack typical TATA sequences, where one promoter contains a CpG-rich area near the TSP, the
other promoter is CpG-poor. Together with the data that the human DNMT1 gene has both CpG-rich
and CpG-poor promoters, it is suggested that transcriptional regulation by two types of
promoters, CpG-rich and CpG-poor, might be common characteristics in the DNMT gene family.

<>

<1>Yang, A.S., Jones, P.A., Shibata, A.
<2>The mutational burden of 5-methylcytosine.
<3>Epigenetic Mechanisms of Gene Regulation, Cold Spring Harbor Laboratory Press, Russo, V.E.A., Martienssen, R.A., Riggs, A.D., Cold Spring Harbor
<4>0
<5>77-94
<6>1996
<7>The importance of the epigenetic modification of cytosine to 5-methylcytosine is discussed in
other chapters of this volume.  In this chapter, we address the heavy mutational burden
induced by the methylation of C at CpG dinucleotides, which is the price that must be paid for
having a m5C epigenetic system.  The mutability of m5C to thymine has presumably led to a
depletion of the CpG dinucleotide in the mammalian genome over the course of evolution.
Furthermore, m5C residues, which comprise only about 1% of the human genome, account for about
30% of the base substitution mutations found in genetic disease and 25% of the mutations found
in tumor suppressor genes.  The mutability of m5C was first demonstrated in Escherichia coli.
Cytosine bases that were methylated in the E. coli lacI gene were found to be hot spots for
spontaneous base substitution mutations, and the hot spots disappeared when the same sites
were unmethylated.  It was speculated that the reason for this increase was that whereas C
deaminates to uracil, m5C deaminates to T, which is a normal DNA base and therefore inherently
more difficult to repair.  This mechanism of DNA mutation is unique in that it is not induced
by exogenous chemicals and it occurs in nonreplicating DNA.  The deamination of m5C is a
first-order chemical process, which is consistent with the fact that deamination occurs on
both the transcribed and nontranscribed strand of DNA with equal probability.

<>

<1>Yang, A.S., Shen, J.-C., Zingg, J.-M., Mi, S., Jones, P.A.
<2>HhaI and HpaII DNA methyltransferases bind DNA mismatches, methylate uracil and block DNA repair.
<3>Nucleic Acids Res.
<4>23
<5>1380-1387
<6>1995
<7>The hydrolytic deamination of 5-methylcytosine (5-mC) to thymine (T) is believed to be
responsible for the high mutability of the CpG dinucleotide in DNA. We have shown a possible
alternate mechanism for mutagenesis at CpG in which HpaII DNA-(cytosine-5) methyltransferase
(M.HpaII) can enzymatically deaminate cytosine (C) to uracil (U) in DNA. Both the hydrolytic
deamination of 5-mC and enzymatic deamination of C create premutagenic DNA mismatches (G:U and
G:T) with the guanine (G) originally paired to the normal C. Surprisingly, we found that
DNA-(cytosine-5) methyltransferases have higher affinities for these DNA mismatches than for
their normal G:C targets and are capable of transferring a methyl group to the 5-position of
U, creating T at low efficiencies. This binding by methyltransferase to mismatches at the
recognition site prevented repair of G:U mismatches by uracil DNA glycosylase in vitro.

<>

<1>Yang, A.S., Shen, J.-C., Zingg, J.-M., Mi, S., Jones, P.A.
<2>DNA (cytosine-5) methyltransferase blocks DNA repair.
<3>Proc. Amer. Assoc. Cancer Res.
<4>36
<5>538
<6>1995
<7>the CpG dinucleotide is markedly underrepresented in the human genome, yet is the site of
about one third of all point mutations found in the germline and in cancer. The hydrolytic
deamination of 5-methylcytosine (5mC) to thymine (T) is believed to be responsible for the
increased mutability of the CpG dinucleotide. We have previously shown an alternate possible
mechanism in which HpaII DNA-(cytosine-5) methyltransferase (M.HpaII), a bacterial enzyme
which carries out methylation by a similar mechanism of action to human methyltransferase, can
enzymatically deaminate cytosine (C) to uracil (U) in DNA. Both the hydrolytic deamination of
5mC to T and enzymatic deamination of C to U create premutagenic DNA mismatches (G:U and G:T)
with the guanine (G) originally paired to the original C. Surprisingly, we found that
methyltransferase has a higher affinity for these mismatches than for the normal target G:C.
Methyltransferase was also shown to transfer a methyl group to the 5-position of U creating T,
presenting a new possible mechanism for C to T transition mutations. This binding by
methyltransferase prevented the repair of G:U mismatches by uracil DNA glycosylase in vitro.
Mutant forms of M.HhaI which bind to the target sequence but lack catalytic activity when
expressed in E. coli bestowed a mutator phenotype presumably due to the blockage of DNA
repair.

<>

<1>Yang, C., Zhang, W., Ren, M., Song, L., Li, T., Zhao, J.
<2>Whole-Genome Sequence of Microcystis aeruginosa TAIHU98, a Nontoxic Bloom-Forming Strain Isolated from Taihu Lake, China.
<3>Genome Announcements
<4>1
<5>e00333-13
<6>2013
<7>Microcystis aeruginosa is a dominant bloom-forming cyanobacterium in many freshwater lakes.
This report describes the first whole-genome sequence of the
nontoxic strain of M. aeruginosa TAIHU98, which was isolated from Taihu Lake in
eastern China.

<>

<1>Yang, C.C., Baxter, B.K., Topal, M.D.
<2>DNA cleavage by NaeI: protein purification, rate-limiting step, and accuracy.
<3>Biochemistry
<4>33
<5>14918-14925
<6>1994
<7>NaeI endonuclease must bind two DNA sites for cleavage to occur. NaeI was purified to apparent
homogeneity and used to determine the rate-limiting step for DNA cleavage and to measure
NaeI's specificity for its cognate recognition site. Steady-state cleavage by NaeI in the
presence of effector DNA (activated) gave values of 0.045s-1 and 10 nM for kcat and KM for M13
DNA substrate, respectively, but values of 0.4s-1 and 170 nM, respectively, for an M13 DNA
fragment substrate. Single-turnover cleavage of M13 DNA cleavage by NaeI showed an initial
burst of substrate cleavage that was proportional to NaeI concentration, implying that product
release is rate-limiting for turnover of NaeI. The NaeI effector and substrate binding sites
were found to prefer cognate over noncognate sequences by 1000-fold and at least 40-500-fold,
respectively. kcat for noncognate recognition sequence was at least 10/6-fold lower than that
for cognate. The specificity for activated NaeI, as measured by kcat/KM, for noncognate
recognition sequence was 10/8-fold lower than that for cognate, and over 10/11-fold lower when
the decreased affinity for noncognate sequence at the effector binding site was taken into
account. This specificity is approximately 10/4-fold larger than for any other restriction
enzyme measured.

<>

<1>Yang, C.C., Topal, M.D.
<2>Nonidentical DNA-binding sites of endonuclease NaeI recognize different families of sequences flanking the recognition site.
<3>Biochemistry
<4>31
<5>9657-9664
<6>1992
<7>NaeI endonuclease uses a two-site binding mechanism to cleave substrate DNA: reaction-rate
studies imply that occupancy of the second DNA site causes an allosteric change in the protein
that enables DNA cleavage at the first site [Conrad, M., & Topal, M.D. (1989) Proc. Natl.
Acad. Sci. U.S.A. 86, 9707-9711]. Measurements of relative binding affinities for 14-base-pair
DNA fragments containing the NaeI recognition sequence GCCGGC and various flanking sequencs
showed that the two DNA-binding sites are not identical. G-C-rich flanking sequences were
preferred by the activator binding site, whereas A-T-rich flanking sequences were preferred by
the substrate binding site: GGGTGCCGGCAGGG was preferred 8-fold more by the activator site but
14-fold less by the substrate site than TTTCGCCGGCGTTT. Substitution of pyrimidine or
7-deazapurine for purine immediately 3'to GCCGGC reduced DNA affinity for only the activator
site by up to 26-fold, implying that the activator DNA-binding site requires N-7 base contacts
immediately flanking GCCGGC. The implications of nonidentical DNA-binding sites, one of which
binds a specific DNA site to allosterically activate the other, are discussed.

<>

<1>Yang, F. et al.
<2>Genome dynamics and diversity of Shigella species, the etiologic agents of bacillary dysentery.
<3>Nucleic Acids Res.
<4>33
<5>6445-6458
<6>2005
<7>The Shigella bacteria cause bacillary dysentery, which remains a significant threat to public
health. The genus status and species
classification appear no longer valid, as compelling evidence indicates
that Shigella, as well as enteroinvasive Escherichia coli, are derived
from multiple origins of E.coli and form a single pathovar. Nevertheless,
Shigella dysenteriae serotype 1 causes deadly epidemics but Shigella
boydii is restricted to the Indian subcontinent, while Shigella flexneri
and Shigella sonnei are prevalent in developing and developed countries
respectively. To begin to explain these distinctive epidemiological and
pathological features at the genome level, we have carried out comparative
genomics on four representative strains. Each of the Shigella genomes
includes a virulence plasmid that encodes conserved primary virulence
determinants. The Shigella chromosomes share most of their genes with that
of E.coli K12 strain MG1655, but each has over 200 pseudogenes, 300
approximately 700 copies of insertion sequence (IS) elements, and numerous
deletions, insertions, translocations and inversions. There is extensive
diversity of putative virulence genes, mostly acquired via
bacteriophage-mediated lateral gene transfer. Hence, via convergent
evolution involving gain and loss of functions, through
bacteriophage-mediated gene acquisition, IS-mediated DNA rearrangements
and formation of pseudogenes, the Shigella spp. became highly specific
human pathogens with variable epidemiological and pathological features.

<>

<1>Yang, F., Tang, C., Wang, Y., Zhang, H., Yue, H.
<2>Genome Sequence of Mycoplasma ovipneumoniae Strain SC01.
<3>J. Bacteriol.
<4>193
<5>5018
<6>2011
<7>Mycoplasma ovipneumoniae is associated with chronic non-progressive pneumonia in both sheep
and goats. Studies concerning its molecular
pathogenesis, genetic analysis and vaccine development have been hindered
due to limited genomic information. Here, we announce the first complete
genome sequence of this organism.

<>

<1>Yang, H., Fitz-Gibbon, S., Marcotte, E.M., Tai, J.H., Hyman, E.C., Miller, J.H.
<2>Characterization of a thermostable DNA glycosylase specific for U/G and T/G mismatches from the hyperthermophilic archaeon Pyrobaculum aerophilum.
<3>J. Bacteriol.
<4>182
<5>1272-1279
<6>2000
<7>U/G and T/G mismatches commonly occur due to spontaneous deamination of cytosine and
5-methylcytosine in double-stranded DNA. This mutagenic
effect is particularly strong for extreme thermophiles, since the
spontaneous deamination reaction is much enhanced at high temperature.
Previously, a U/G and T/G mismatch-specific glycosylase (Mth-MIG) was
found on a cryptic plasmid of the archaeon Methanobacterium
thermoautotrophicum, a thermophile with an optimal growth temperature of
65 degrees C. We report characterization of a putative DNA glycosylase
from the hyperthermophilic archaeon Pyrobaculum aerophilum, whose optimal
growth temperature is 100 degrees C. The open reading frame was first
identified through a genome sequencing project in our laboratory. The
predicted product of 230 amino acids shares significant sequence homology
to [4Fe-4S]-containing Nth/MutY DNA glycosylases. The histidine-tagged
recombinant protein was expressed in Escherichia coli and purified. It is
thermostable and displays DNA glycosylase activities specific to U/G and
T/G mismatches with an uncoupled AP lyase activity. It also processes
U/7,8-dihydro-oxoguanine and T/7,8-dihydro-oxoguanine mismatches. We
designate it Pa-MIG. Using sequence comparisons among complete bacterial
and archaeal genomes, we have uncovered a putative MIG protein from
another hyperthermophilic archaeon, Aeropyrum pernix. The unique conserved
amino acid motifs of MIG proteins are proposed to distinguish MIG proteins
from the closely related Nth/MutY DNA glycosylases.

<>

<1>Yang, H., He, T., Wu, W., Zhu, W., Lu, B., Sun, W.
<2>Whole-Genome Shotgun Assembly and Analysis of the Genome of Streptomyces mobaraensis DSM 40847, a Strain for Industrial Production of Microbial  Transglutaminase.
<3>Genome Announcements
<4>1
<5>e0014313
<6>2013
<7>Here, we report the draft annotated genome sequence of Streptomyces mobaraensis strain DSM
40847, which is used in industry to produce microbial
transglutaminase. The genome sequence will allow for the characterization of the
molecular mechanisms underlying the beneficial properties of this organism.

<>

<1>Yang, H., Liao, Y., Wang, B., Lin, Y., Pan, L.
<2>Complete Genome Sequence of Bacillus amyloliquefaciens XH7, Which Exhibits Production of Purine Nucleosides.
<3>J. Bacteriol.
<4>193
<5>5593-5594
<6>2011
<7>Here, we report the complete annotated genome sequence of Bacillus amyloliquefaciens XH7,
which is used to produce purine nucleosides in
industry. The genome sequence will allow for the characterization of the
molecular mechanisms underlying its beneficial properties.

<>

<1>Yang, H., Liao, Y., Wang, B., Lin, Y., Pan, L.
<2>Draft Genome Sequence of Escherichia coli XH001, a Producer of L-Threonine in Industry.
<3>J. Bacteriol.
<4>193
<5>6406-6407
<6>2011
<7>l-Threonine has been widely used as a supplement in the food, pharmaceutical, and cosmetic
industries. Here, we present a high-quality
draft annotated genome sequence of Escherichia coli XH001, a producer of
l-threonine in industry. Its genome and plasmid sequence will provide
clues about the molecular mechanisms underlying its beneficial properties.

<>

<1>Yang, H., Liao, Y., Wang, B., Lin, Y., Pan, L.
<2>Genome Sequence of Escherichia coli XH140A, Which Produces L-Threonine.
<3>J. Bacteriol.
<4>193
<5>6090-6091
<6>2011
<7>Here we report the draft annotated genome sequence of Escherichia coli XH140A, which is used
to produce l-threonine in industry. The genome
sequence will allow the characterization of the molecular mechanisms
underlying its beneficial properties.

<>

<1>Yang, H., Zhang, Z., Yan, R., Wang, Y., Zhu, D.
<2>Draft Genome Sequence of Streptomyces sp. Strain PRh5, a Novel Endophytic Actinomycete Isolated from Dongxiang Wild Rice Root.
<3>Genome Announcements
<4>2
<5>e00012-14
<6>2014
<7>Here, we report the draft genome sequence of Streptomyces sp. strain PRh5 (China  Center for
Type Culture Collection [CCTCC] number 2013487), which is used to produce nigericin and
nocardamine. The genome sequence will allow for the characterization of the molecular
mechanisms underlying its beneficial properties.

<>

<1>Yang, J., Deng, C., Hemati, N., Hanash, S.M., Richardson, B.C.
<2>Effect of mitogenic stimulation and DNA methylation on human T cell DNA methyltransferase expression and activity.
<3>J. Immunol.
<4>159
<5>1303-1309
<6>1997
<7>DNA methylation, a mechanism modifying gene expression, is mediated in part by the enzyme DNA
methyltransferase.  Reduced levels of T cell DNA methyltransferase have been observed in
lupus-like diseases, and increased levels have been reported in malignancies.  Little is known
concerning the regulation of human DNA methyltransferase.  In this report we demonstrate that
mitogenic T cell stimulation causes an increase in DNA methyltransferase mRNA and enzyme
activity.  We also show that pharmacologic inhibition of T cell DNA methylation causes an
increase in the rate of DNA methyltransferase mRNA transcription and a corresponding increase
in mRNA levels and enzyme activity.  This suggests that DNA methyltransferase is itself
regulated in part by DNA methylation status, possibly representing a feedback mechanism.  DNA
methylation inhibition also resulted in an increase in Ha-ras and c-jun mRNA levels,
overexpression of which increases DNA methyltransferase in murine systems.  These results thus
identify two mechanisms regulating levels of human T cell DNA methyltransferase and raise the
possibility that abnormalities in either could contribute to disorders associated with altered
DNA methylation.

<>

<1>Yang, J., Dong, C.X., Huang, X., Zhao, J.D.
<2>Sulfonation of polyvinylidene difluoride resin and its application in extraction of restriction enzymes from DNA digestion solutions.
<3>Anal. Biochem.
<4>322
<5>99-103
<6>2003
<7>Sulfonation of polyvinylidene difluoride (PVDF) resin was achieved by incubation of the resin
with sulfuric acid at a moderately high
temperature. The sulfonated PVDF (SPVDF) resin was studied for its ability
to extract restriction enzymes from DNA digestion solutions. The SPVDF
resin was effective in adsorbing restriction enzymes such as EcoRI and
BamHI and the extraction procedure was easy and simple to perform. The
adsorption depended upon the amount of the resin added. We found that 1 mg
of the SPVDF resin could completely remove all restriction enzyme activity
routinely used in DNA digestion within 2 min after its addition. Treatment
of a digestion solution with the SPVDF resin did not change the reaction
solution and the same digestion buffer could be used for another digestion
of the same DNA with other enzymes. We also found that, in comparison with
normal PVDF, the SPVDF resin adsorbed less DNA, resulting in less loss of
DNA in the extraction step. The potential application of the SPVDF resin
in other procedures of molecular cloning and enzyme purification is
discussed.

<>

<1>Yang, J., Malik, H.S., Eickbush, T.H.
<2>Identification of the endonuclease domain encoded by R2 and other site-specific, non-long terminal repeat retrotransposable elements.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>96
<5>7847-7852
<6>1999
<7>The non-long terminal repeat retrotransposon, R2, encodes a sequence-specific endonuclease
responsible for its insertion at a unique site in the 28S rRNA genes of arthropods.  Although
most non-LTR retrotransposons encode an apurinic-like endonuclease upstream of a common
reverse transcriptase domain, R2 and many other site-specific non-LTR elements do not (CRE1
and 2, SLACS, CZAR, Dong, R4).  Sequence comparison of these site-specific elements has
revealed that the region downstream of their reverse transcriptase domain is conserved and
shares sequence features with various prokaryotic restriction endonucleases.  In particular,
these non-LTR elements have a Lys/Arg-Pro-Asp-X12-14aa-Asp/Glu motif known to lie near the
scissile phosphodiester bonds in the protein-DNA complexes of restriction enzymes.
Site-directed mutagenesis of the R2 protein was used to provide evidence that this motif is
also part of the active site of the endonuclease encoded by this element.  Mutations of this
motif eliminate both DNA-cleavage activities of the R2 protein: first-strand cleavage in which
the exposed 3' end is used to prime reverse transcription of the RNA template and
second-strand cleavage, which occurs after reverse transcription.  The general organization of
the R2 protein appears similar to the type IIS restriction enzyme, FokI, in which specific DNA
binding is controlled by a separate domain located amino terminal to the cleavage domain.
Previous phylogenetic analysis of their reverse transcriptase domains has indicated that the
non-LTR elements identified here as containing restriction-like endonucleases are the oldest
lineages of non-LTR elements, suggesting a scenario for the evolution of non-LTR elements.

<>

<1>Yang, J., Mohr, G., Perlman, P.S., Lambowitz, A.M.
<2>Group II intron mobility in yeast mitochondria: target DNA-primed reverse transcription activity of aI1 and reverse splicing into DNA transposition sites in vitro.
<3>J. Mol. Biol.
<4>282
<5>505-523
<6>1998
<7>The retrohoming of the yeast mtDNA intron aI1 occurs by a target DNA-primed reverse
transcription (TPRT) mechanism in which the intron RNA reverse splices directly into the
recipient DNA and is then copied by the intron-encoded reverse transcriptase. Here, we carried
out biochemical characterization of the intron-encoded reverse transcriptase and site-specific
DNA endonuclease activities required for this process. We show that the aI1 reverse
transcriptase has high TPRT activity in the presence of appropriate DNA target sites, but
differs from the closely related reverse transcriptase encoded by the yeast aI2 intron in
being unable to use artificial substrates efficiently. Characterization of TPRT products shows
that the fully reverse spliced intron RNA is an efficient template for cDNA synthesis, while
reverse transcription of partially reverse spliced intron RNA is impeded by the branch point.
Novel features of the aI1 reaction include a prominent open-circular product in which cDNAs
are incorporated at a nick at the antisense-strand cleavage site. The aI1 endonuclease
activity, which catalyzes the DNA cleavage and reverse splicing reactions, is associated with
ribonucleoprotein particles containing the intron-encoded protein and the excised intron RNA.
As shown for the aI2 endonuclease, both the RNA and protein components are used for DNA target
site recognition, but the aI1 protein has less stringent nucleotide sequence requirements for
the reverse splicing reaction. Finally, perhaps reflecting this relaxed target specificity, in
vitro experiments show that aI1 can reverse splice directly into ectopic mtDNA transposition
sites, consistent with the previously suggested possibility that this mechanism is used for
ectopic transposition of group II introns in vivo.

<>

<1>Yang, J., Zimmerly, S., Perlman, P.S., Lambowitz, A.M.
<2>Efficient integration of an intron RNA into double-stranded DNA by reverse splicing.
<3>Nature
<4>381
<5>332-335
<6>1996
<7>Some group II introns are mobile elements as well as catalytic RNAs.  Introns aI1 and aI2
found in the gene COX1 in yeast mitochondria encode reverse transcriptases which promote
site-specific insertion of the intron into intronless alleles ('homing').  For aI2 this
predominantly occurs by reverse transcription of unspliced precursor RNA at a break in
double-strand DNA made by an endonuclease encoded by the intron.  The aI2 endonuclease
involves both the excised intron RNA, which cleaves the DNA's sense strand by partial reverse
splicing; and the intron-encoded reverse transcriptase which cleaves the antisense strand.
Here we show that aI1 encodes an analogous endonuclease specific for a different target site
compatible with the different exon-binding sequences of the intron RNA.  Over half of aI1
undergoes complete reverse splicing in vitro, thus integrating linear intron RNA directly into
the DNA.  This unprecedented reaction has implications for both intron mobility and evolution,
and potential genetic engineering applications.

<>

<1>Yang, J.M., Deurraza, P.J., Matvienko, N., O'sullivan, D.J.
<2>Involvement of the LlaKR2I Methylase in Expression of the AbiR Bacteriophage Defense System in Lactococcus lactis subsp. lactis biovar   diacetylactis KR2.
<3>J. Bacteriol.
<4>188
<5>1920-1928
<6>2006
<7>The native lactococcal plasmid, pKR223, from Lactococcus lactis subsp. lactis biovar
diacetylactis KR2 encodes two distinct
bacteriophage-resistant mechanisms, the LlaKR2I restriction and
modification (R/M) system and the abortive infection (Abi) mechanism,
AbiR, that impedes bacteriophage DNA replication. This study completed the
characterization of AbiR, revealing that it is the first Abi system to be
encoded by three genes, abiRa, abiRb, and abiRc, arranged in an operon and
that it requires the methylase gene from the LlaKR2I R/M system. An
analysis of deletion and insertion clones demonstrated that the AbiR
operon was toxic in L. lactis without the presence of the LlaKR2I
methylase, which is required to protect L. lactis from AbiR toxicity. The
novelty of the AbiR system resides in its original gene organization and
the unusual protective role of the LlaKR2I methylase. Interestingly, the
AbiR genetic determinants are flanked by two IS982 elements generating a
likely transposable AbiR composite. This observation not only
substantiated the novel function of the LlaKR2I methylase in the AbiR
system but also illustrated the evolution of the LlaKR2I methylase toward
a new and separate cellular function. This unique structure of both the
LlaKR2I R/M system and the AbiR system may have contributed to the
evolution of the LlaKR2I methylase toward a novel role comparable to that
of the cell cycle-regulated methylases that include Dam and CcrM
methylases. This new role for the LlaKR2I methylase offers a unique
snapshot into the evolution of the cell cycle-regulated methylases from an
existing R/M system.

<>

<1>Yang, L.F., Som, S., Friedman, S.
<2>Effect of N-terminal deletions on the activity of EcoRII DNA methylase.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>A436
<6>1991
<7>DNA(cytosine-5)-methyltransferases (Mtases) contain highly conserved core
sequences and variable regions.  One of the variable regions is at the
N-terminus where about 100 amino acids present in the enzymes M.EcoRII or
M.MspI are almost totally absent in other Mtases.  In order to determine the
function of the N-terminal residues in M.EcoRII we prepared a series of
N-terminal deletion mutants in pUC19.  Analysis of the mutants showed that
almost the entire region N-terminal to the first conserved motif can be deleted
with retention of enzyme activity.  In some mutants translation starts with an
internal ATG codon of the gene.  Other active constructs are either fused in
frame with lac z or utilize an ATG codon in the cloning site of the vector.
Deletion of 96 residues resulted in marked decrease in activity.  Longer
deletions were inactive.  One such mutant protein deleted of 85 residues was
purified and characterized.  In vitro methylation studies showed that this
truncated mutant retained specificity.  However, the Km for AdoMet was 15 fold
higher than the Km found with the wild type enzyme.  The results indicate that
the N-terminal region of the protein is not vital for retention of catalytic
activity but increases the affinity of the enzyme for AdoMet.

<>

<1>Yang, M., Lv, Y., Xiao, J., Wu, H., Zheng, H., Liu, Q., Zhang, Y., Wang, Q.
<2>Edwardsiella comparative phylogenomics reveal the new intra/inter-species taxonomic relationships, virulence evolution and niche adaptation mechanisms.
<3>PLoS ONE
<4>7
<5>E36987
<6>2012
<7>Edwardsiella bacteria are leading fish pathogens causing huge losses to
aquaculture industries worldwide. E. tarda is a broad-host range pathogen that
infects more than 20 species of fish and other animals including humans while E.
ictaluri is host-adapted to channel catfish causing enteric septicemia of catfish
(ESC). Thus, these two species consist of a useful comparative system for
studying the intricacies of pathogen evolution. Here we present for the first
time the phylogenomic comparisons of 8 genomes of E. tarda and E. ictaluri
isolates. Genome-based phylogenetic analysis revealed that E. tarda could be
separate into two kinds of genotypes (genotype I, EdwGI and genotype II, EdwGII)
based on the sequence similarity. E. tarda strains of EdwGI were clustered
together with the E. ictaluri lineage and showed low sequence conservation to E.
tarda strains of EdwGII. Multilocus sequence analysis (MLSA) of 48 distinct
Edwardsiella strains also supports the new taxonomic relationship of the
lineages. We identified the type III and VI secretion systems (T3SS and T6SS) as
well as iron scavenging related genes that fulfilled the criteria of a key
evolutionary factor likely facilitating the virulence evolution and adaptation to
a broad range of hosts in EdwGI E. tarda. The surface structure-related genes may
underlie the adaptive evolution of E. ictaluri in the host specification
processes. Virulence and competition assays of the null mutants of the
representative genes experimentally confirmed their contributive roles in the
evolution/niche adaptive processes. We also reconstructed the hypothetical
evolutionary pathway to highlight the virulence evolution and niche adaptation
mechanisms of Edwardsiella. This study may facilitate the development of
diagnostics, vaccines, and therapeutics for this under-studied pathogen.

<>

<1>Yang, M.-K., Ser, S.-C., Lee, C.-H.
<2>Involvement of E. coli dcm methylase in Tn3 transposition.
<3>Proc. Natl. Sci. Counc. Repub. China B
<4>13
<5>276-283
<6>1989
<7>The effects of DNA methyltransferases on Tn3 transposition were investigated. The E. coli dam
(deoxyadenosine methylase) gene was found to have no effect on Tn3 transposition. In contrast,
Tn3 was found to transpose more frequently in dcm+ (deoxycytosine methylase) cells than in
dcm- mutants. When the EcoRII methylase gene was introduced into dcm- cells (E. coli strain
GM208), the frequency of Tn3 transposition in GM208 was dramatically increased. The EcoRII
methylase recognizes and methylates the same sequence as does the dcm methylase. These results
suggest tht deoxycytosine methylase modified DNA may be a preferred target for Tn3
transposition. Experiments were also performed to determine whether the Tn3 transposase was
involved in DNA modification. Plasmid DNA isolated from dcm- E. Coli containing the Tn3
transposase gene was susceptible to ApyI digestion but resistant to EcoRII digestion,
suggesting that Tn3 transposase modified the dcm recognition sequence. In addition,
restriction enzymes TaqI, AvaII, BglI and HpaII did not digest this DNA completely, suggesting
that the recognition sequences of TaqI, AvaII, BglI and HpaII were modified by Tn3 transposase
to a certain degree. The type(s), the extent and mechanism(s) of this modification remain to
be investigated.

<>

<1>Yang, S., Pappas, K.M., Hauser, L.J., Land, M.L., Chen, G.L., Hurst, G.B., Pan, C., Kouvelis, V.N., Typas, M.A., Pelletier, D.A., Klingeman, D.M., Chang, Y.J., Samatova, N.F., Brown, S.D.
<2>Improved genome annotation for Zymomonas mobilis.
<3>Nat. Biotechnol.
<4>27
<5>893-894
<6>2009
<7>The first genome sequence of the enthanologenic bacterium Zymomonas mobilis ZM4 was reported
in your journal five years ago.  Because of its productivity, high level of ethanol tolerance
and ability to be genetically manipulated, Z. mobilis is a promising industrial bacterium for
fermenting lignocellulosic biomass into enthanol, which is being advanced as an alternative to
petroleum-derived transportation fuels.  Strains of the bacterium have been developed to
ferment both hexoses and pantoses into ethanol, and the genome sequence will provide further
opportunities for strain developments as well as providing more fundamental insights.  We have
observed many differences between the primary annotation and one performed by the J. Craig
Venter Institute (JCVI) and have detected differential gene expression in genes predicted by
JCVI (http://cmr.jcvi.org/cgi-bin/CMR/GenomePage.cgi?org=ntzm01) that were absent from the
primary annotation in our previous work.  We present here an improved version of the ZM4
genome annotation based on the original primary gene sequence, new epxerimental data generated
by 454 pyrosequencing and mass spectrometry-based proteomics, a novel gene prediction
algorithm called Prodigal (prokaryotic dynamic programming genefinding algorithm) that is
incorporated into an annotation pipeline, and manual curation.

<>

<1>Yang, S.J., Kang, I., Cho, J.C.
<2>Genome Sequence of Strain IMCC14465, Isolated from the East Sea, Belonging to the PS1 Clade of Alphaproteobacteria.
<3>J. Bacteriol.
<4>194
<5>6952-6953
<6>2012
<7>Strain IMCC14465 was isolated from surface seawater of the East Sea using
dilution-to-extinction culturing. Phylogenetic analysis indicated that the strain
belongs to the PS1 clade, which is closely related to the OCS116 clade in the
Alphaproteobacteria. Here, we report the genome sequence of IMCC14465, the first
isolate of the PS1 clade.

<>

<1>Yang, W., Lee, J.Y., Nowotny, M.
<2>Making and breaking nucleic acids: Two-Mg2+-ion catalysis and substrate specificity.
<3>Mol. Cell
<4>22
<5>5-13
<6>2006
<7>DNA and a large proportion of RNA are antiparallel duplexes composed of an unvarying
phosphosugar backbone surrounding uniformly stacked and
highly similar base pairs. How do the myriad of enzymes (including
ribozymes) that perform catalysis on nucleic acids achieve exquisite
structure or sequence specificity? In all DNA and RNA polymerases and
many nucleases and transposases, two Mg2+ ions are jointly coordinated
by the nucleic acid substrate and catalytic residues of the enzyme.
Based on the exquisite sensitivity Of Mg2+ ions to the ligand geometry
and electrostatic environment, we propose that two-metal-ion catalysis
greatly enhances substrate recognition and catalytic specificity.

<>

<1>Yang, X. et al.
<2>A public genome-scale lentiviral expression library of human ORFs.
<3>Nat. Methods
<4>8
<5>659-661
<6>2011
<7>Functional characterization of the human genome requires tools for systematically
modulating gene expression in both loss-of-function and gain-of-function
experiments. We describe the production of a sequence-confirmed, clonal
collection of over 16,100 human open-reading frames (ORFs) encoded in a versatile
Gateway vector system. Using this ORFeome resource, we created a genome-scale
expression collection in a lentiviral vector, thereby enabling both targeted
experiments and high-throughput screens in diverse cell types.

<>

<1>Yang, X., Chen, C., Wang, D., Yang, S., Wu, R.
<2>Cloning and overexpression of the restriction gene of EcoRI.
<3>Acta Biochim. Biophys. Sin.
<4>22
<5>599-606
<6>1990
<7>Cloning and expression of the EcoRI genes are reported here.  A plasmid
containing the EcoRI (r/m) genes was extracted from the EcoRI producing strain
E. coli RI, and a 4.9 kb BamHI-HindIII fragment encoding the EcoRI genes was
cloned into pBR322, producing a hybrid plasmid pER101.  pER302, containing the
methylase gene of EcoRI, was obtained by inserting the 1.7 kb fragment from
pER101 into pACYC184.  pER304, in which the sequence coding for the C-terminal
of EcoRI was downstream from the lambda PL promoter, was constructed from pAS1
and pER101.  After cloning the 1.5 kb SalI-BglII fragment of pER101 into
pER304, an EcoRI overproducing plasmid pER306 was created using the strong
promoter PL as well as the endogenous promoter Pe to overexpress the EcoRI(r)
gene.  To delete the Pe promoter to control expression of EcoRI only by the PL
promoter, a 0.65 kb BamHI-HindIII fragment from pER401 was used to replace the
small BamHI-HindIII fragment downstream from PL promoter, resulting in another
overproducing plasmid pER307.  When transforming GI139 and GI139/pER302 the
relative survival efficiency of pER306 and pER307 was both 10/5, but when using
TG1/pRK248CIts and GI139/pER302 as hosts the efficiency was 10/5 and 1
respectively.  Therefore, in certain hosts, pER307 can express EcoRI in the
absence of its cognate methylase.  After heat induction, pER307
[TG1/pRK248CIts] produces EcoRI at a level of 10/5 units per gram of wet cell,
which is about 1-2 orders of magnitude lower than that of pER306/[GI139/pER302]
or pER307/[GI139/pER302].

<>

<1>Yang, X., Wang, Y., Huo, G.
<2>Complete Genome Sequence of Lactococcus lactis subsp. lactis KLDS4.0325.
<3>Genome Announcements
<4>1
<5>e00962-13
<6>2013
<7>We report the complete genome sequence of Lactococcus lactis subsp. lactis KLDS4.0325, a
probiotic bacterium isolated from homemade koumiss in Xinjiang,
China. We have determined the complete genome sequence of strain KLDS4.0325,
which consists of a chromosome and three plasmids and reveals genes that are
likely to be involved in dairy fermentation and that have probiotic qualities.

<>

<1>Yang, X., Xue, R., Shen, C., Li, S., Gao, C., Wang, Q., Zhao, X.
<2>Genome sequence of Rhodococcus sp. R04, a polychlorinated biphenyl biodegrader.
<3>J. Bacteriol.
<4>193
<5>5032-5033
<6>2011
<7>The genus Rhodococcus has proved to be a promising option for the clean-up of polluted sites
and application of a microbial biocatalyst. Rhodococcous
sp. strain R04, isolated from oil contaminated soil, could biodegrade
polychlorinated biphenyls. Here we report the draft genome sequence of
Rhodococcous sp. strain R04, which could be used to predict genes for
xenobiotic biodegradation and provide important insights into the
applications of this strain.

<>

<1>Yang, Y., Li, Q.
<2>Restriction enzyme SnaBI is sensitive to methylation of cytosine within its recognition sequence.
<3>Nucleic Acids Res.
<4>18
<5>3083
<6>1990
<7>None

<>

<1>Yang, Y., Yu, X., Zhang, R.
<2>Draft Genome Sequence of Ochrobactrum pseudogrignonense Strain CDB2, a Highly Efficient Arsenate-Resistant Soil Bacterium from Arsenic-Contaminated Cattle Dip   Sites.
<3>Genome Announcements
<4>1
<5>e00173-13
<6>2013
<7>We report the 4.97-Mb draft genome sequence of a highly efficient arsenate-resistant
bacterium, Ochrobactrum sp. strain CDB2. It contains a novel
arsenic resistance (ars) operon (arsR-arsC1-ACR3-arsC2-arsH-mfs) and two
non-operon-associated ars genes, arsC3 and arsB. The genome information will aid
in the understanding of the arsenic resistance mechanism of this and other
bacterial species.

<>

<1>Yang, Y.T., Chen, I.T., Lee, C.T., Chen, C.Y., Lin, S.S., Hor, L.I., Tseng, T.C., Huang, Y.T., Sritunyalucksana, K., Thitamadee, S., Wang, H.C., Lo, C.F.
<2>Draft Genome Sequences of Four Strains of Vibrio parahaemolyticus, Three of Which Cause Early Mortality Syndrome/Acute Hepatopancreatic Necrosis Disease in Shrimp   in China and Thailand.
<3>Genome Announcements
<4>2
<5>e00816-14
<6>2014
<7>We sequenced four Vibrio parahaemolyticus strains, three of which caused serious  acute
hepatopancreatic necrosis disease. Sequence analysis of the virulent
strains revealed not only genes related to cholera toxin and the type IV
pilus/type IV secretion system but also a unique, previously unreported, large
extrachromosomal plasmid that encodes a homolog to the insecticidal Photorhabdus
insect-related binary toxin PirAB.

<>

<1>Yang, Z., Horton, J.R., Maunus, R., Wilson, G.G., Roberts, R.J., Cheng, X.
<2>Structure of HinP1I endonuclease reveals a striking similarity to the monomeric restriction enzyme MspI.
<3>Nucleic Acids Res.
<4>33
<5>1892-1901
<6>2005
<7>HinP1I, a type II restriction endonuclease, recognizes and cleaves a palindromic
tetranucleotide sequence (G/CGC) in double-stranded DNA,
producing 2 nt 5' overhanging ends. Here, we report the structure of
HinP1I crystallized as one protein monomer in the crystallographic
asymmetric unit. HinP1I displays an elongated shape, with a conserved
catalytic core domain containing an active-site motif of SDX18QXK and a
putative DNA-binding domain. Without significant sequence homology, HinP1I
displays striking structural similarity to MspI, an endonuclease that
cleaves a similar palindromic DNA sequence (C/CGG) and binds to that
sequence crystallographically as a monomer. Almost all the structural
elements of MspI can be matched in HinP1I, including both the DNA
recognition and catalytic elements. Examining the protein-protein
interactions in the crystal lattice, HinP1I could be dimerized through two
helices located on the opposite side of the protein to the active site,
generating a molecule with two active sites and two DNA-binding surfaces
opposite one another on the outer surfaces of the dimer. A possible
functional link between this unusual dimerization mode and the tetrameric
restriction enzymes is discussed.

<>

<1>Yang, Z., Horton, J.R., Zhou, L., Zhang, X.J., Dong, A.P., Zhang, X., Schlagman, S.L., Kossykh, V., Hattman, S., Cheng, X.D.
<2>Structure of the bacteriophage T4 DNA adenine methyltransferase.
<3>Nat. Struct. Biol.
<4>10
<5>849-855
<6>2003
<7>DNA-adenine methylation at certain GATC sites plays a pivotal role in bacterial and phage gene
expression as well as bacterial virulence. We
report here the crystal structures of the bacteriophage T4Dam DNA adenine
methyltransferase (MTase) in a binary complex with the methyl-donor
product S-adenosyl-L-homocysteine (AdoHcy) and in a ternary complex with a
synthetic 12-bp DNA duplex and AdoHcy. T4Dam contains two domains: a
seven-stranded catalytic domain that harbors the binding site for AdoHcy
and a DNA binding domain consisting of a five-helix bundle and a
beta-hairpin that is conserved in the family of GATC-related MTase
orthologs. Unexpectedly, the sequence-specific T4Dam bound to DNA in a
nonspecific mode that contained two Dam monomers per synthetic duplex,
even though the DNA contains a single GATC site. The ternary structure
provides a rare snapshot of an enzyme poised for linear diffusion along
the DNA.

<>

<1>Yangtse, W., Zhou, Y., Lei, Y., Qiu, Y., Wei, X., Ji, Z., Qi, G., Yong, Y., Chen, L., Chen, S.
<2>Genome Sequence of Bacillus licheniformis WX-02.
<3>J. Bacteriol.
<4>194
<5>3561-3562
<6>2012
<7>Bacillus licheniformis is an important bacterium that has been used extensively for
large-scale industrial production of exoenzymes and peptide antibiotics. B.
licheniformis WX-02 produces poly-gamma-glutamate increasingly when fermented
under stress conditions. Here its genome sequence (4,270,104 bp, with G+C content
of 46.06%), which comprises a circular chromosome, is announced.

<>

<1>Yankovsky, N.K., Fonstein, M.Y., Lashina, S.Y., Bukanov, N.O., Yakubovich, N.V., Ermakova, L.M., Rebentish, B.A., Janulaitis, A., Debabov, V.G.
<2>Phasmids as effective and simple tools for construction and analysis of gene libraries.
<3>Gene
<4>81
<5>203-210
<6>1989
<7>Phasmid lambda-pMYF131, a hybrid of phage lambda vectors and plasmid pUC19, was
constructed.  The phasmid and its derivatives were shown to be efficient
vectors for construction and analysis of gene libraries in Escherichia coli
cells.  The lambda-pMYF131 DNA molecule contains all the genes and regions
essential for phage lytic development.  The plasmid cannot be packaged either
in the monomeric or the oligomeric form due to its specific length.  Elongation
of the DNA molecule by ligation with fragments of foreign DNA can make it
packageable and this is easily detected by plaque formation. Hence, the
procedures used to construct genomic libraries can be simplified by selection
of only recombinant DNA molecules just at the time and on the basis of their
packaging in vitro.  The output of recombinant clones per vector molecule was
several times higher for vector lambda-pMYF131, compared to phage vector
lambdaL47.1AB, and attained 3x10/6 clones per microgram DNA.  Vector and
recombinant phasmids can be obtained in large quantities in plasmid form.
Lambda-pMYF131 contains nine unique restriction sites which allow the cloning
of DNA fragments with blunt ends and of fragments with various types of
cohesive ends, obtained by digestion with 14 prototype restriction enzymes.
The maximal size of the cloned DNA fragments is approximately 20 kb for
lambda-pMYF131.  Phasmid vectors were used to construct libraries of bovine,
pig and quail genomes, and genomic libraries of 17 species of bacteria.
Application of suitable methods allowed the identification 13 individual genes
within these libraries.  Reports the cleavage sites for EcoICRI and Ecl136II.

<>

<1>Yanofsky, S., Boyer, H., Grable, J., McClarin, J., Rosenberg, J., Greene, P.
<2>Mutational analysis of the EcoRI endonuclease.
<3>J. Cell Biochem. Suppl.
<4>11C
<5>211
<6>1987
<7>Using in vitro mutagenesis with hydroxylamine a large number of null mutants of
the EcoRI endonuclease have been generated.  Fifty per cent of the 121 mutants
examined by western blot tested positive against anti-RI.  The entire
endonuclease gene from twenty-seven western positive mutants was sequenced by
using dideoxy sequencing directly from double stranded plasmid DNA.  Twenty of
these were single base change substitutions, with 10 of the mutants falling
within the region from ala 139 to glu 144.  Examination of the mutants with
respect to the structure of the EcoRI endonuclease-DNA co-crystal reveals that
all of the null mutants map at either the protein-DNA interface or at the
subunit-subunit interface.  Molecular weight determination of purified protein
on a BioSil TS250 HPLC column demonstrates that wild type EcoRI endonuclease
runs as a dimer of 60Kd.  Three mutants with alterations at the protein-DNA
interface (AT139, GS140 and RQ203) run as dimers while 3 mutants mapping at the
subunit-subunit interface (EK144, EK152 and GR210) run as monomers.

<>

<1>Yanofsky, S., Boyer, H., Greene, P.
<2>Positive control of EcoRI endonuclease expression by the EcoRI methylase.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>85
<5>106
<6>1985
<7>The presence of a wild type EcoRI endonuclease in cells lacking the cognate
methylase is lethal.  We have isolated a mutant endonuclease in which serine
replaces arginine at residue 187.  The Ser 187 enzyme retains RI specificity
but has reduced in vivo activity (as measured by ability to restrict growth of
lambda phage).  Cells carrying the Ser 187 endonuclease can survive when the
methylase is deleted.  Deletion of the methylase results in drastically reduced
endonuclease production but levels can be restored to near normal by the
presence of the methylase provided in trans on pSC101, a compatible low-copy
number plasmid.  We have constructed a number of strains in which the methylase
gene is truncated at different sites.  Analysis of these strains has allowed us
to define a domain within the methylase gene which activates endonuclease
production.  In some of the methylase-deficient strains we are able to increase
endonuclease production by induction of a linked lac promoter.  This induction
is lethal into the cell, and it is clear that survival depends upon both low
production of the Ser 187 endonuclease as well as its reduced in vivo activity.
This positive control of endonuclease production by methylase provides a
mechanism for establishment and maintenance of this system in a cell without
damaging cellular DNA.

<>

<1>Yanofsky, S.D., Love, R., McClarin, J.A., Rosenberg, J.M., Boyer, H.W., Greene, P.J.
<2>Clustering of null mutations in the EcoRI endonuclease.
<3>Proteins
<4>2
<5>273-282
<6>1987
<7>EcoRI endonuclease mutants were isolated in a methylase-deficient background
following in vitro hydroxylamine mutagenesis of plasmid pKG2 (Kuhn et al.: Gene
44:253-263, 1986).  Mutants which survived high-level endonuclease expression
(IPTG induction) were termed null mutants.  Sixty-two of 121 null mutants
tested by Western blot contained normal levels of endonuclease cross-reacting
protein.  The complete endonuclease gene was sequenced for 27 null mutants.
This group was found to consist of 20 single base-change missense mutantations,
6 double mutations, and 1 triple mutation.  Ten of the 20 single mutations were
clustered between residues 139 and 144.  When examined with respect to the
structure of the EcoRI-DNA complex (McClarin et al.:  Science 234:1526-1541,
1986), these alterations were found to fall predominantly into two classes:
substitutions at the protein-DNA interface or substitutions at the
protein-protein (dimer) interface.  Protein from several of the mutants was
purified and sized by using HPLC.  Wild-type EcoRI endonuclease and protein
from three of the DNA interface mutations (Ala139->Thr, Gly140->Ser,
Arg203->Gln) appeared to be dimeric, while protein from subunit interface
mutations (glu->Lys, Glu152->Lys, Gly210->Arg) migrated as monomers.

<>

<1>Yao, N., Ren, Y., Wang, W.
<2>Genome Sequence of a Thermophilic Bacillus, Geobacillus thermodenitrificans DSM465.
<3>Genome Announcements
<4>1
<5>e01046-13
<6>2013
<7>Geobacillus thermodenitrificans NG80-2 encodes a LadA-mediated alkane degradation pathway,
while G. thermodenitrificans DSM465 cannot utilize alkanes. Here, we
report the draft genome sequence of G. thermodenitrificans DSM465, which may help
reveal the genomic differences between these two strains in regards to the
biodegradation of alkanes.

<>

<1>Yao, Y., Imirzalioglu, C., Hain, T., Kaase, M., Gatermann, S., Exner, M., Mielke, M., Hauri, A., Dragneva, Y., Bill, R., Wendt, C., Wirtz, A., Domann, E., Chakraborty, T.
<2>Complete Nucleotide Sequence of a Citrobacter freundii Plasmid Carrying KPC-2 in  a Unique Genetic Environment.
<3>Genome Announcements
<4>2
<5>e01157-14
<6>2014
<7>The complete and annotated nucleotide sequence of a 54,036-bp plasmid harboring a blaKPC-2
gene that is clonally present in Citrobacter isolates from different
species is presented. The plasmid belongs to incompatibility group N (IncN) and
harbors the class A carbapenemase KPC-2 in a unique genetic environment.

<>

<1>Yao, Y., Tang, H., Ren, H., Yu, H., Wang, L., Xu, P.
<2>Genome sequence of a nicotine-degrading strain of arthrobacter.
<3>J. Bacteriol.
<4>194
<5>5714-5715
<6>2012
<7>We announce a 4.63-Mb genome assembly of an isolated bacterium that is the first  sequenced
nicotine-degrading Arthrobacter strain. Nicotine catabolism genes of
the nicotine-degrading plasmid pAO1 were predicted, but plasmid function genes
were not found. These results will help to better illustrate the molecular
mechanism of nicotine degradation by Arthrobacter.

<>

<1>Yap, H.Y., Ghazali, K., Wan, M.N.W.F., Mat, I.M.N., Zakaria, Z., Omar, A.R.
<2>Draft Genome Sequence of Pasteurella multocida subsp. multocida Strain PMTB, Isolated from a Buffalo.
<3>Genome Announcements
<4>1
<5>e00872-13
<6>2013
<7>Pasteurella multocida serotypes B:2 and E:2 are the main causative agents of ruminant
hemorrhagic septicemia in Asia and Africa, respectively. Pasteurella
multocida strain PMTB was isolated from a buffalo with hemorrhagic septicemia and
has been determined to be serotype B:2. Here we report the draft genome sequence
of strain PMTB.

<>

<1>Yap, K.P., Gan, H.M., Teh, C.S., Baddam, R., Chai, L.C., Kumar, N., Tiruvayipati, S.A., Ahmed, N., Thong, K.L.
<2>Genome Sequence and Comparative Pathogenomics Analysis of a Salmonella enterica Serovar Typhi Strain Associated with a Typhoid Carrier in Malaysia.
<3>J. Bacteriol.
<4>194
<5>5970-5971
<6>2012
<7>Salmonella enterica serovar Typhi is a human pathogen that causes typhoid fever predominantly
in developing countries. In this article, we describe the whole
genome sequence of the S. Typhi strain CR0044 isolated from a typhoid fever
carrier in Kelantan, Malaysia. These data will further enhance the understanding
of its host persistence and adaptive mechanism.

<>

<1>Yap, K.P., Teh, C.S., Baddam, R., Chai, L.C., Kumar, N., Avasthi, T.S., Ahmed, N., Thong, K.L.
<2>Insights from the Genome Sequence of a Salmonella enterica Serovar Typhi Strain Associated with a Sporadic Case of Typhoid Fever in Malaysia.
<3>J. Bacteriol.
<4>194
<5>5124-5125
<6>2012
<7>Salmonella enterica serovar Typhi is the causative agent of typhoid fever, which  causes
nearly 21.7 million illnesses and 217,000 deaths globally. Herein, we
describe the whole-genome sequence of the Salmonella Typhi strain ST0208,
isolated from a sporadic case of typhoid fever in Kuala Lumpur, Malaysia. The
whole-genome sequence and comparative genomics allow an in-depth understanding of
the genetic diversity, and its link to pathogenicity and evolutionary dynamics,
of this highly clonal pathogen that is endemic to Malaysia.

<>

<1>Yarmolinsky, M.B.
<2>Programmed cell death in bacterial populations.
<3>Science
<4>267
<5>836-837
<6>1995
<7>Multicellular organisms benefit not only from the death of competitors, but often, quite
dramatically, from the programmed death of specific subpopulations of their own cells. Popular
opinion to the contrary, programmed cell death is also alive and well in the microbial world.
A striking report by Naito et al. in this issue of Science is a case in point.

<>

<1>Yasar-Yildiz, S., Kambourova, M., Arga, K.Y., Toksoy, O.E.
<2>Draft Genome Sequence of Exopolysaccharide-Producing Thermophilic Bacterium Brevibacillus thermoruber Strain 423.
<3>Genome Announcements
<4>1
<5>e00774-13
<6>2013
<7>Brevibacillus thermoruber strain 423 is a Gram-positive, spore-forming, aerobic,  and
thermophilic bacterium that produces mannogalactoglucan exopolysaccharide (EPS). We report the
draft genome sequence of B. thermoruber 423, which will accelerate research on the cellular
organization of thermophilic bacteria, as well as the rational design and optimization of EPS
production.

<>

<1>Yasawong, M. et al.
<2>Complete genome sequence of Arcanobacterium haemolyticum type strain (11018).
<3>Standards in Genomic Sciences
<4>3
<5>126-135
<6>2010
<7>Arcanobacterium haemolyticum (ex MacLean et al. 1946) Collins et al. 1983 is the  type species
of the genus Arcanobacterium, which belongs to the family
Actinomycetaceae. The strain is of interest because it is an obligate parasite of
the pharynx of humans and farm animal; occasionally, it causes pharyngeal or skin
lesions. It is a Gram-positive, nonmotile and non-sporulating bacterium. The
strain described in this study was isolated from infections amongst American
soldiers of certain islands of the North and West Pacific. This is the first
completed sequence of a member of the genus Arcanobacterium and the ninth type
strain genome from the family Actinomycetaceae. The 1,986,154 bp long genome with
its 1,821 protein-coding and 64 RNA genes is a part of the Genomic Encyclopedia
of Bacteria and Archaea project.

<>

<1>Yasui, K., Kano, Y., Tanaka, K., Watanabe, K., Shimizu-Kadota, M., Yoshikawa, H., Suzuki, T.
<2>Improvement of bacterial transformation efficiency using plasmid artificial modification.
<3>Nucleic Acids Res.
<4>37
<5>e3
<6>2009
<7>We have developed a method to improve the transformation efficiency in genome-sequenced
bacteria, using 'Plasmid Artificial Modification' (PAM),
using the host's own restriction system. In this method, a shuttle vector
was pre-methylated in Escherichia coli cells, which carry all the putative
genes encoding the DNA modification enzymes of the target microorganism,
before electroporation was performed. In the case of Bifidobacterium
adolescentis ATCC15703 and pKKT427 (3.9 kb E. coli-Bifidobacterium shuttle
vector), introducing two Type II DNA methyltransferase genes lead to an
enhancement in the transformation efficiency by five orders of magnitude.
This concept was also applicable to a Type I restriction system. In the
case of Lactococcus lactis IO-1, by using PAM with a putative Type I
methyltransferase system, hsdMS1, the transformation efficiency was
improved by a factor of seven over that without PAM.

<>

<1>Yasuike, M., Nakamura, Y., Kai, W., Fujiwara, A., Fukui, Y., Satomi, M., Sano, M.
<2>Draft Genome Sequence of Agarivorans albus Strain MKT 106T, an Agarolytic Marine  Bacterium.
<3>Genome Announcements
<4>1
<5>e00367-13
<6>2013
<7>Agarivorans albus is a Gram-negative, strictly aerobic, and agar-hydrolyzing marine bacterium.
We present the draft genome sequence of the A. albus strain MKT
106(T), which is composed of 67 contigs (>500 bp) totaling 4,734,285 bp and
containing 4,397 coding DNA sequences (CDSs), four rRNAs, and 64 tRNA sequences.

<>

<1>Yau, E.K., Coward, J.K.
<2>The synthesis of complex 6'-alkynyl-6'-dethia analogs of s-adenosylhomocysteine as potential inhibitors of methyltransferases.
<3>ACS Abstracts
<4>198
<5>100
<6>1989
<7>A new series of multisubstrate adduct inhibitors was designed for
S-adenosylmethionine-dependent methyltransferases.  Three synthetic routes to
this class of compounds were investigated and
5'-(6-amino-6-carboxy-1-phenyl-1-hexynyl-3-)-5'-deoxyadenosine was synthesized
as a prototype.  The key step in the successful synthesis was the arylation of
N6-benzoyl-2',3'-isopropylidene-5'(5-tert-butyldiphenylsilyloxy-1-pentynyl-3)-5
'-deoxyadenosine, a terminal acetylene, with an aryl iodide in the presence of
Pd(II) and CuI catalysts.  This method should be generally useful for the
synthesis of specific inhibitors of S-adenosylmethionine-dependent
methyltransferases.  5'-(4-Amino-4-carboxybutyl-1-)-5'-deoxyadenosine, a
6'-carbon analog of S-adenosylhomocysteine, was also synthesized in the process
of investigating methods for constructing an amino acid moiety on the side
chain of this class of compounds.

<>

<1>Yau, S., Lauro, F.M., DeMaere, M.Z., Brown, M.V., Thomas, T., Raftery, M.J., Andrews-Pfannkoch, C., Lewis, M., Hoffman, J.M., Gibson, J.A., Cavicchioli, R.
<2>Virophage control of antarctic algal host-virus dynamics.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>108
<5>6163-6168
<6>2011
<7>Viruses are abundant ubiquitous members of microbial communities and in
the marine environment affect population structure and nutrient cycling by
infecting and lysing primary producers. Antarctic lakes are microbially
dominated ecosystems supporting truncated food webs in which viruses exert
a major influence on the microbial loop. Here we report the discovery of a
virophage (relative of the recently described Sputnik virophage) that
preys on phycodnaviruses that infect prasinophytes (phototrophic algae).
By performing metaproteogenomic analysis on samples from Organic Lake, a
hypersaline meromictic lake in Antarctica, complete virophage and
near-complete phycodnavirus genomes were obtained. By introducing the
virophage as an additional predator of a predator-prey dynamic model we
determined that the virophage stimulates secondary production through the
microbial loop by reducing overall mortality of the host and increasing
the frequency of blooms during polar summer light periods. Virophages
remained abundant in the lake 2 y later and were represented by
populations with a high level of major capsid protein sequence variation
(25-100% identity). Virophage signatures were also found in neighboring
Ace Lake (in abundance) and in two tropical lakes (hypersaline and fresh),
an estuary, and an ocean upwelling site. These findings indicate that
virophages regulate host-virus interactions, influence overall carbon flux
in Organic Lake, and play previously unrecognized roles in diverse aquatic
ecosystems.

<>

<1>Yazdanparast, S., Toma, E., Karska-Wysocki, B.
<2>Restriction enzyme activity for subgenomic discrimination of  Methicillin-resistant Staphylococcus aureus.
<3>Abstracts of IBC's 7th Annual World Congress, , , Boston, MA
<4>0
<5>abstract #100
<6>2002
<7>The dissemination of methicillin-resistant Staphylococcus aureus has important consequences in
health-care as it is the source of many outbreaks in hospital centers.  Presently, the primary
focus of most hospital infection control programs is to reduce the prevalence of MRSA
infection by rapidly detecting the source of these organisms and by interrupting their
transmission to patients.  We previously reported the development of a strain discrimination
procedure using fifty MRSA clinical isolates from a hospital center.  After evaluation of the
multidrug-resistance of MRSA strains, we determined their electrophoretic plasmid DNA profile.
We can differentiate fifty strains into eleven groups.  In addition, we have investigated the
Restriction Fragment Length Polymorphism of plasmidic and genomic DNA after digestion with
EcoRI, HindIII and the recently discovered LlaGI restriction enzyme.  In this study we
characterized the plasmid DNA fragments of MRSA strains digested by LlaGI.  This procedure was
followed by analysis of genomic DNA amplicons profiles generated by arbitrarily primed PCR.
We were able to subtype our representative MRSA isolates into 3 types which were clearly
different from each other.  The discriminatory power of genomic DNA AP-PCR fingerprinting can
be increased, after digestion of amplicons with restriction enzyme and analysis the generated
electrophoretic fragments profiles.  In order to discriminate more precisely the MRSA strains,
we digested AP-PCR amplicons by AluI.  After analysis of the electrophoretic digest patterns
of these amplicons, we were able to identify the individual strains.  To our knowledge, this
is the first report of MRSA strain identification with RFLP or AP-PCR-RFLP procedures, using
these two enzymes, LlaGI and AluI, respectively.  We conclude that the combination of
procedures, such as analysis of electrophoretic plasmid profiles, analysis of genomic DNA
using AP-PCR and AP-PCR-RFLP were most efficacious for identification of individual MRSA
strains.  Our development of a new typing schema demonstrates the novel restriction enzyme
activity application for subgenomic discrimination of MRSA strains.

<>

<1>Yazdanparast, S., Toma, E., Karska-Wysocki, B.
<2>Characterization of methicillin-resistant Staphylococcus aureus isolated at Hotel-Dieu Hospital, Montreal.
<3>Ottawa Life Sciences Internl Conf. and Exhibit., , , Ottawa, Ontario
<4>0
<5>Abstract #P301
<6>2001
<7>The first objective of this study was to characterize fifty methicillin resistant
Staphylococcus aureus (MRSZA) strains, isolated in 1999/2000 from Hotel-Dieu Hospital
patients, by using antibiogram and DNA analysis.  The second objective was to determine
whether our recently discovered restriction enzyme LlaGI shows a digestive activity towards
MRSA DNA.  The evaluation of antibiotic resistance (by MIC) to methicillin, oxacillin,
erythromycin and cephazolin, showed multi resistance for these drugs for all fifty MRSA
strains.  In contrast, all MRSA strains were sensitive to vancomycin.  We purified the DNA
(plasmidic

<>

<1>Ye, C., Zheng, H., Zhang, J., Jing, H., Wang, L., Xiong, Y., Wang, W., Zhou, Z., Sun, Q., Luo, X., Du, H., Gottschalk, M., Xu, J.
<2>Clinical, experimental, and genomic differences between intermediately pathogenic, highly pathogenic, and epidemic Streptococcus suis.
<3>J. Infect. Dis.
<4>199
<5>97-107
<6>2009
<7>BACKGROUND: Streptococcus suis emerged to cause an unusual outbreak of
streptococcal toxic-shock-like syndrome (STSLS) in 2005. The mechanisms
involved are unknown. METHODS: Clinical, laboratory, and epidemiologic
data on patients infected with culture-confirmed S. suis were analyzed.
The strain involved in the outbreak, "epidemic" strain ST7, was compared
with both a classical highly pathogenic strain, ST1, and an intermediately
pathogenic strain, ST25, to determine both its capacity to induce
cytokines in experimentally infected mice and its genomic difference.
RESULTS: Of 38 patients infected with culture-confirmed S. suis, 14
presented with STSLS. During the early phase of the disease, serum levels
of interleukin (IL)-1beta, IL-6, IL-8, IL-12p70, interferon-gamma, and
tumor necrosis factor-alpha were more elevated in patients with STSLS than
in those with meningitis only. Serum levels of proinflammatory cytokines
were significantly higher in mice infected with ST7 than in those infected
with either ST1 or ST25. Genomic comparisons with ST25 showed that ST1 had
acquired 132 genomic islands, including 5 pathogenicity islands, and that
ST7, the epidemic strain, had acquired an additional 5 genomic islands.
CONCLUSION: Intermediately pathogenic strain ST25 has evolved to become
highly pathogenic strain ST1, which, in turn, has more recently evolved to
become epidemic strain ST7. ST7 has the ability to stimulate the
production of massive amounts of proinflammatory cytokines, leading to
STSLS.

<>

<1>Ye, W., Ye, S., Liu, J., Chang, S., Chen, M., Zhu, B., Guo, L., An, Q.
<2>Genome Sequence of the Pathogenic Herbaspirillum seropedicae Strain Os34, Isolated from Rice Roots.
<3>J. Bacteriol.
<4>194
<5>6993-6994
<6>2012
<7>Most Herbaspirillum seropedicae strains are beneficial endophytes to plants. In contrast, H.
seropedicae strain Os34, isolated from rice roots, is pathogenic.
The draft genome sequence of strain Os34 presented here allows in-depth
comparative genome analyses to understand the specific mechanisms of beneficial
and pathogenic Herbaspirillum-plant interactions.

<>

<1>Ye, Y., Stivers, J.T.
<2>Fluorescence-based high-throughput assay for human DNA (cytosine-5)-methyltransferase 1.
<3>Anal. Biochem.
<4>401
<5>168-172
<6>2010
<7>We have developed the first economical and rapid nonradioactive assay method that is suitable
for high-throughput screening of the important
pharmacological target human DNA (cytosine-5)-methyltransferase 1
(DNMT1). The method combines three key innovations: the use of a
truncated form of the enzyme that is highly active on a 26-bp
hemimethylated DNA duplex substrate, the introduction of the
methylation site into the recognition sequence of a restriction
endonuclease, and the use of a fluorogenic read-out method. The extent
of DNMT1 methylation is reflected in the protection of the DNA
substrate from endonuclease cleavage that would otherwise result in a
large fluorescence increase. The assay has been validated in a
high-throughput format, and trivial changes in the substrate sequence
and endonuclease allow adaptation of the method to any bacterial or
human DNA methyltransferase.

<>

<1>Yebra, M.J., Bhagwat, A.S.
<2>A rapid and sensitive method to measure DNA endonuclease activity.
<3>Nucleic Acids Res.
<4>21
<5>5797-5798
<6>1993
<7>The high affinity of biotin for streptavidin has been used to isolate DNA-binding proteins
from cells. This is achieved by mixing DNA oligomers and containing biotin with cell extracts
and then passing the mixture over immobilized streptavidin. We describe below an adaptation of
this method to quantitate the activities of endonucleases. It should be useful for detecting
endonuclease activity during enzyme purification, for studying inhibitors and activators of
nucleases and for determining kinetic properties of endonucleases.

<>

<1>Yebra, M.J., Bhagwat, A.S.
<2>A cytosine methyltransferase converts 5-methylcytosine in DNA to thymine.
<3>Biochemistry
<4>34
<5>14752-14757
<6>1995
<7>Sites of cytosine methylation are known to be hot spots for C.G to T.A mutations in a number
of systems, including human cells.  Traditionally, spontaneous hydrolytic deamination of
5-methylcytosine to thymine has been invoked as the cause of this phenomenon.  We show here
that a bacterial cytosine methyltransferase can convert 5-methylcytosine in DNA to thymine and
that this reaction creates a mutational hot spot at a site of DNA methylation.  The reaction
is fairly insensitive to the methyl donor in the reaction, S-adenosylmethionine.  In many
cancers, the most frequent class of mutations is C to T changes within CG dinucleotides of the
tumor suppressor gene p53.  Because of the similarities of the reaction mechanisms of
mammalian and bacterial enzymes and the physiology of the cancer cells, this reaction is
expected to contribute to mutations at CG dinucleotides in precancerous cells.

<>

<1>Yebra, M.J., Bhagwat, A.S.
<2>A novel repetitive sequence lies near the gene encoding a cytosine methyltransferase in the cyanobacterium Dactylococcopsis salina.
<3>Gene
<4>164
<5>71-74
<6>1995
<7>An unusual cluster of tandemly repeated DNA sequences (TRS) was found downstream from the gene
encoding DsaV methyltransferase, the DNA modification enzyme in the DsaV
restriction-modification system found in a strain of Dactylococcopsis salina (Ds).  The repeat
unit is about 32-bp long and is present 13 times in the cluster.  Each repeat unit can be
divided into two distinct parts based on the level of sequence conservation and evolution.
Hybridization of Ds DNA with a probe specific for this cluster revealed that there were at
least two additional sites within the genome with similar TRS.  The TRS units are localized in
one region of the Ds genome.  They do not share significant sequence similarity with other TRS
found in prokaryotes.

<>

<1>Yebra, M.J., Novella, I.S., Barbes, C., Aparicio, J.F., Martin, C.G., Hardisson, C., Sanchez, J.
<2>Characterization of Rrh4273I, a restriction-modification system of Rhodococcus rhodochrous ATCC 4273 (Nocardia corallina) which recognizes the same sequence as the Streptomyces albus G SalI restriction-modification system.
<3>J. Gen. Microbiol.
<4>137
<5>1279-1284
<6>1991
<7>Rhodococcus rhodochrous ATCC 4273 (Nocardia corallina) has a
restriction-modification system with the same recognition sequence, methylation
site and cleavage site as the SalI restriction-modification system.  Both the
restriction endonuclease and the DNA-methyltransferase (DNA-MTase) have been
partially purified and characterized.  The nuclease has requirements for
activity similar to SalI, and a native Mr of about 46,000.  The DNA-MTase is a
protein with an Mr of about 67,000.  No DNA homology was detected between the
cloned SalI restriction-modification genes of Streptomyces albus and R.
rhodochrous chromosomal DNA.

<>

<1>Yebra, M.J., Sanchez, J., Martin, C.G., Hardisson, C., Barbes, C.
<2>The effect of sinefungin and synthetic analogues on RNA and DNA methyltransferase from Streptomyces.
<3>J. Antibiot. (Tokyo)
<4>44
<5>1141-1147
<6>1991
<7>Sinefungin is an antibiotic structurally related to S-adenosylmethionine.  It
has been described as an inhibitor of RNA transmethylation reactions in viruses
and eukaryotic organisms, but not in bacteria.  We show here that sinefungin
strongly inhibits RNA methyltransferase activity, but not the biosynthesis of
these enzymes in Streptomyces.  All the methylated bases found in Streptomyces
RNA (1-methyladenine, N6-methyladenine, N6,N6-dimethyladenine and
7-methylguanine) are inhibited by this antibiotic.  Experiments with sinefungin
analogues show that specific changes in the ornithine radical of the molecule
still preserve its inhibitory capability.  The substitution of the adenine
radical by uridine causes the loss of inhibitory effect.  These results and our
former studies on Streptomyces DNA methylation, suggest that nucleic acid
modification is the main target of sinefungin in Streptomyces.

<>

<1>Yee, L., Hosoyama, A., Ohji, S., Tsuchikane, K., Shimodaira, J., Yamazoe, A., Fujita, N., Suzuki-Minakuchi, C., Nojiri, H.
<2>Complete Genome Sequence of a Dimethyl Sulfide-Utilizing Bacterium, Acinetobacter guillouiae Strain 20B (NBRC 110550).
<3>Genome Announcements
<4>2
<5>e01048-14
<6>2014
<7>Acinetobacter guillouiae strain 20B can utilize dimethyl sulfide (DMS) as the sole sulfur
source and degrade chloroethylenes. We report here the complete
4,648,418-bp genome sequence for this strain, which contains 4,367 predicted
coding sequences (CDSs), including a well-characterized DMS degradative operon.

<>

<1>Yen, Ly.H.T., Park, S.-Y., Kim, J.-S.
<2>Cloning, crystallization and preliminary X-ray diffraction analysis of an intact DNA methyltransferase of a type I restriction-modification enzyme from Vibrio vulnificus.
<3>Acta Crystallogr. F Struct. Biol. Cryst. Commun.
<4>70
<5>489-492
<6>2014
<7>Independently of the restriction (HsdR) subunit, the specificity (HsdS) and methylation (HsdM)
subunits interact with each other, and function as a methyltransferase in type I
restriction-modification systems. A single gene that combines the HsdS and HsdM subunits in
Vibrio vulnificus YJ016 was expressed and purified. A crystal suitable for X-ray diffraction
was obtained from 25%(w/v) polyethylene glycol monomethylether 5000, 0.1 M HEPES pH 8.0, 0.2 M
ammonium sulfate at 291 K by hanging-drop vapour diffusion. Diffraction data were collected to
a resolution of 2.31 angstrom using synchrotron radiation. The crystal belonged to the
primitive monoclinic space group P2(1), with unit-cell parameters a = 93.25, b = 133.04, c =
121.49 angstrom, beta = 109.7 degrees. With four molecules in the asymmetric unit, the crystal
volume per unit protein weight was 2.61 angstrom(3) Da(-1), corresponding to a solvent content
of 53%.

<>

<1>Yen, R.-W.C., Vertino, P.M., Nelkin, B.D., Yu, J.J., El-Deiry, W., Cumaraswamy, A., Lennon, G.G., Trask, B.J., Celano, P., Baylin, S.B.
<2>Isolation and characterization of the cDNA encoding human DNA methyltransferase.
<3>Nucleic Acids Res.
<4>20
<5>2287-2291
<6>1992
<7>We have cloned a series of overlapping cDNA clones encoding a 5194 bp transcript for human DNA
methyltransferase (DNA MTase). This sequence potentially codes for a protein of 1495 amino
acids with a predicted molecular weight of 169 kDa. The human DNA MTase cDNA has eighty
percent homology at the nucleotide level, and the predicted protein has seventy-four percent
identity at the amino acid level, to the DNA MTase cDNA cloned from mouse cells. Like the
murine DNA MTase, the amino terminal two-thirds of the human protein contains a cysteine-rich
region suggestive of a metal-binding domain. The carboxy terminal one-third of the protein
shows considerable similarity to prokaryotic (cytosine-5)-methyltransferases. The arrangement
of multiple motifs conserved in the prokaryotic genes is preserved in the human DNA MTase,
including the relative position of a proline-cysteine dipeptide thought to be an essential
catalytic site in all (cytosine-5)-methyltransferases. A single 5.2 kb transcript was detected
in all human tissues tested, with the highest levels of expression observed in RNA from
placenta, brain, heart and lung. DNA MTase cDNA clones were used to screen a chromosome 19
genomic cosmid library. The DNA MTase-positive cosmids which are estimated to span a genomic
distance of 93 kb have been localized to 19p13.2-p13.3 by fluorescence in situ hybridization.
Isolation of the cDNA for human DNA MTase will further study the regulation of DNA MTase
expression, and of the role of this enzyme in establishing DNA methylation patterns in both
normal and neoplastic cells.

<>

<1>Yeo, C.C., Tham, J.M., Kwong, S.M., Poh, C.L.
<2>Characterization of the Pac25I restriction-modification genes isolated from the endogenous pRA2 plasmid of Pseudomonas alcaligenes NCIB 9867.
<3>Plasmid
<4>40
<5>203-213
<6>1998
<7>Genes for the class II Pseudomonas alcaligenes NCIB 9867 restriction-modification (R-M)
system, Pac25I, have been cloned from its 33-kb endogenous plasmid, pRA2. The Pac25I
endonuclease and methylase genes were found to be aligned in a head-to-tail orientation with
the methylase gene preceding and overlapping the endonuclease gene by 1 bp. The deduced amino
acid sequence of the Pac25I methylase revealed significant similarity with the XcyI, XmaI,
Cfr9I, and SmaI methylases. High sequence similarity was displayed between the Pac25I
endonuclease and the XcyI, XmaI, and Cfr9I endonucleases which cleave between the external
cytosines of the recognition sequence (i.e., 5'-C^CCGGG-3') and are thus perfect
isoschizomers. However, no sequence similarity was detected between the Pac25I endonuclease
and the SmaI endonuclease which cleaves between the internal CpG of the recognition sequence
(i.e., 5'-CCC^GGG-3'). Both the Pac25I methylase and endonuclease were expressed in
Escherichia coli. An open reading frame encoding a protein which shows significant similarity
to invertases and resolvases was located immediately upstream of the Pac25I R-M operon. In
addition, a transposon designated Tn5563 was located 1531 bp downstream of the R-M genes. The
location on a self-transmissible plasmid as well as the close association with genes involved
in DNA mobility suggests horizontal transfer as a possible mode of distribution of this family
of R-M genes in various bacteria.

<>

<1>Yeo, I.C., Lee, N.K., Hahm, Y.T.
<2>Genome Sequencing of Bacillus subtilis SC-8, Antagonistic to the Bacillus cereus Group, Isolated from Traditional Korean Fermented-Soybean Food.
<3>J. Bacteriol.
<4>194
<5>536-537
<6>2012
<7>Bacillus subtilis SC-8 is a Gram-positive bacterium displaying narrow antagonistic activity
for the Bacillus cereus group. B. subtilis SC-8 was
isolated from Korean traditional fermented-soybean food. Here we report
the draft genome sequence of B. subtilis SC-8, including biosynthetic
genes for antibiotics that may have beneficial effects for control of
food-borne pathogens.

<>

<1>Yeoman, C.J., Yildirim, S., Thomas, S.M., Durkin, A.S., Torralba, M., Sutton, G., Buhay, C.J., Ding, Y., Dugan-Rocha, S.P., Muzny, D.M., Qin, X., Gibbs, R.A., Leigh, S.R., Stumpf, R., White, B.A., Highlander, S.K., Nelson, K.E., Wilson, B.A.
<2>Comparative genomics of Gardnerella vaginalis strains reveals substantial differences in metabolic and virulence potential.
<3>PLoS ONE
<4>5
<5>E12411
<6>2010
<7>BACKGROUND: Gardnerella vaginalis is described as a common vaginal
bacterial species whose presence correlates strongly with bacterial
vaginosis (BV). Here we report the genome sequencing and comparative
analyses of three strains of G. vaginalis. Strains 317 (ATCC 14019) and
594 (ATCC 14018) were isolated from the vaginal tracts of women with
symptomatic BV, while Strain 409-05 was isolated from a healthy,
asymptomatic individual with a Nugent score of 9. PRINCIPAL FINDINGS:
Substantial genomic rearrangement and heterogeneity were observed that
appeared to have resulted from both mobile elements and substantial
lateral gene transfer. These genomic differences translated to differences
in metabolic potential. All strains are equipped with significant
virulence potential, including genes encoding the previously described
vaginolysin, pili for cytoadhesion, EPS biosynthetic genes for biofilm
formation, and antimicrobial resistance systems, We also observed systems
promoting multi-drug and lantibiotic extrusion. All G. vaginalis strains
possess a large number of genes that may enhance their ability to compete
with and exclude other vaginal colonists. These include up to six
toxin-antitoxin systems and up to nine additional antitoxins lacking
cognate toxins, several of which are clustered within each genome. All
strains encode bacteriocidal toxins, including two lysozyme-like toxins
produced uniquely by strain 409-05. Interestingly, the BV isolates encode
numerous proteins not found in strain 409-05 that likely increase their
pathogenic potential. These include enzymes enabling mucin degradation, a
trait previously described to strongly correlate with BV, although
commonly attributed to non-G. vaginalis species. CONCLUSIONS:
Collectively, our results indicate that all three strains are able to
thrive in vaginal environments, and therein the BV isolates are capable of
occupying a niche that is unique from 409-05. Each strain has significant
virulence potential, although genomic and metabolic differences, such as
the ability to degrade mucin, indicate that the detection of G. vaginalis
in the vaginal tract provides only partial information on the
physiological potential of the organism.

<>

<1>Yeon, S.M., Kim, Y.C.
<2>Complete sequence and organization of the Sphingobium chungbukense DJ77 pSY2 plasmid.
<3>J. Microbiol.
<4>49
<5>684-688
<6>2011
<7>Sphingobium chungbukense DJ77 is capable of metabolizing priority
chemicals of human health concern such as polycyclic aromatic hydrocarbons
(PAHs), extracellular polysaccharide (EPS), and antibiotics. Here, we
report the complete DNA and genetic organization of the plasmid pSY2 from
strain DJ77. A DNA sequence analysis revealed that pSY2 comprises 18,779
bp encoding 22 open reading frames (ORFs) with 59.5% G+C content. The ORFs
on pSY2 were classified into DNA replication, conjugative function,
transposition, plasmid stability/partition, and other functional groups
(transport, fatty acid biosynthesis, stress, and growth rate regulation).
Three ORFs on pSY2 were hypothetical proteins.

<>

<1>Yergeau, E., Lawrence, J.R., Waiser, M.J., Korber, D.R., Greer, C.W.
<2>Metatranscriptomic analysis of the response of river biofilms to pharmaceutical products, using anonymous DNA microarrays.
<3>Appl. Environ. Microbiol.
<4>76
<5>5432-5439
<6>2010
<7>Pharmaceutical products are released at low concentrations into aquatic
environments following domestic wastewater treatment. Such low
concentrations have been shown to induce transcriptional responses in
microorganisms, which could have consequences on aquatic ecosystem
dynamics. In order to test if these transcriptional responses could also
be observed in complex river microbial communities, biofilm reactors were
inoculated with water from two rivers of differing trophic statuses and
subsequently treated with environmentally relevant doses (ng/liter to
microg/liter range) of four pharmaceuticals (erythromycin [ER],
gemfibrozil [GM], sulfamethazine [SN], and sulfamethoxazole [SL]). To
monitor functional gene expression, we constructed a 9,600-feature
anonymous DNA microarray platform onto which cDNA from the biofilms was
hybridized. Pharmaceutical treatments induced both positive and negative
transcriptional responses from biofilm microorganisms. For instance, ER
induced the transcription of several stress, transcription, and
replication genes, while GM, a lipid regulator, induced transcriptional
responses from several genes involved in lipid metabolism. SN caused
shifts in genes involved in energy production and conversion, and SL
induced responses from a range of cell membrane and outer envelope genes,
which in turn could affect biofilm formation. The results presented here
demonstrate for the first time that low concentrations of small molecules
can induce transcriptional changes in a complex microbial community. The
relevance of these results also demonstrates the usefulness of anonymous
DNA microarrays for large-scale metatranscriptomic studies of communities
from differing aquatic ecosystems.

<>

<1>Yesufu, H.M.I., Hanley, A., Rinaldi, A., Adams, R.L.P.
<2>DNA methylase from Pisum sativum.
<3>Biochem. J.
<4>273
<5>469-475
<6>1991
<7>DNA methylase activity was detected in nuclei from pea shoots.  The enzyme can only be
extracted by low-salt treatment if the nuclei are pretreated with micrococcal nuclease.  Only
a single enzyme was detected, and it was purified to a specific activity of 1620 units/mg of
protien.  It has an Mr of 160,000 on gel filtration and SDS/PAGE.  Pea DNA methylase
methylates cytosine in all four dinucleotides, and this is interpreted to show that it acts on
CNG trinucleotides.  Although it shows a strong preference for hemi-methylated double-stranded
DNA, it is also capable of methylation de novo.  Homologous DNA is the best natural substrate.
In vitro the enzyme interacts with DNA to form a salt-resistant complex with DNA that is
stable for at least 4h.

<>

<1>Yew, S.M., Chan, C.L., Soo-Hoo, T.S., Na, S.L., Ong, S.S., Hassan, H., Ngeow, Y.F., Hoh, C.C., Lee, K.W., Yee, W.Y., Ng, K.P.
<2>Draft Genome Sequence of Dematiaceous Coelomycete Pyrenochaeta sp. Strain UM 256, Isolated from Skin Scraping.
<3>Genome Announcements
<4>1
<5>e00158-13
<6>2013
<7>Pyrenochaeta, classified under the order Pleosporales, is known to cause diseases in plants
and humans. Here, we report a draft genome sequence of a Pyrenochaeta
sp. isolated from a skin scraping, with an estimated genome size of 39.4 Mb.
Genes associated with the synthesis of proteases, toxins, plant cell wall
degradation, and multidrug resistance were found.

<>

<1>Yi, C.Q., Fong, C.C., Chen, W.W., Qi, S.J., Tzang, C.H., Lee, S.T., Yang, M.S.
<2>Interactions between carbon nanotubes and DNA polymerase and restriction endonucleases.
<3>Nanotechnol.
<4>18
<5>025102
<6>2007
<7>Effects of multi-walled carbon nanotubes (MWCNT) and single-walled carbon nanotubes (SWCNT)
functionalized with and without carboxylic
groups on polymerase chain reaction (PCR) and restriction digestion
reaction were investigated. The results showed that CNT can reduce and
even inhibit PCR and restriction digestion reaction, possibly due to
the decrease of respective enzyme activity. The inhibition effect on
double restriction digestion reaction and PCR was increased in the
order of CNT-COOH > pristine CNT and SWCNT > MWCNT. This study
demonstrated that CNT may significantly affect the efficiency of
biochemical reactions through different action mechanisms, which is
critical for understanding how nanomaterials impact biological systems.

<>

<1>Yi, H., Cho, Y.J., Hur, H.G., Chun, J.
<2>Genome sequence of Escherichia coli AA86, isolated from cow feces.
<3>J. Bacteriol.
<4>193
<5>3681
<6>2011
<7>Escherichia coli AA86 (= KACC 15541) is an enteric bacterium that was isolated from a sample
of healthy cow feces. Its genome sequence revealed
that it is most closely related to the human fecal strain E. coli SE15 and
could be classified under E. coli phylogenetic group B2. Here, we report
the genome sequence of E. coli AA86 consisting of 3 contigs and 2
plasmids.

<>

<1>Yi, H., Cho, Y.J., Yong, D., Chun, J.
<2>Genome Sequence of Escherichia coli J53, a Reference Strain for Genetic Studies.
<3>J. Bacteriol.
<4>194
<5>3742-3743
<6>2012
<7>Escherichia coli J53 (F(-) met pro Azi(r)) is a derivative of E. coli K-12 which  is resistant
to sodium azide. This strain has been widely used as a general
recipient strain for various conjugation experiments. Here, we report the genome
sequence of E. coli J53 (=KACC 16628).

<>

<1>Yilder, C., Onsan, Z.I., Kirdar, B.
<2>Optimization of starting time and period of induction and inducer concentration in the production of the restriction enzyme EcoRI from recombinant Escherichia coli 294.
<3>Turk. J. Chem.
<4>22
<5>221-226
<6>1998
<7>Induction parameters including inducer concentration, period of induction and the cell
concentration at which inducer is to be added to the fermentation broth were optimized in
order to increase the yield of the EcoRI restriction endonuclease isolated from recombinant E.
coli.  Bacterial cells harboring the plasmid pPG430 containing EcoRI endonuclease and the
methylase genes under the control of lac promoter were used in the experiments where induction
was accomplished by using lactose isopropyl-B-D-thiogalactoside.  An IPTG concentration of
0.1mM, the late exponential phase of growth (at an optical density of 1.2 at 595 nm) and an
induction period of 6 hours were determined to be the optimum conditions for induction.

<>

<1>Yildirim, S., Elhanafi, D., Lin, W., Hitchins, A.D., Siletzky, R.M., Kathariou, S.
<2>Conservation of Genomic Localization and Sequence Content of Sau3AI-Like Restriction-Modification Gene Cassettes among Listeria monocytogenes Epidemic Clone I and Selected Strains of Serotype 1/2a.
<3>Appl. Environ. Microbiol.
<4>76
<5>5577-5584
<6>2010
<7>Listeria monocytogenes is a food-borne pathogen with a clonal population structure and
apparently limited gene flow between strains
of different lineages. Strains of epidemic clone I (ECI) have been
responsible for numerous outbreaks and invariably have DNA that is
resistant to digestion by Sau3AI, suggesting methylation of cytosine at
GATC sites. A putative restriction-modification (RM) gene cassette has
been identified in the genome of the ECI strain F2365 and all other
tested ECI strains but is absent from other strains of the same
serotype (4b). Homologous RM cassettes have not been reported among L.
monocytogenes isolates of other serotypes. Furthermore, conclusive
evidence for the involvement of this RM cassette in the Sau3AI
resistance phenotype of ECI strains has been lacking. In this study, we
describe a highly conserved RM cassette in certain strains of serotypes
1/2a and 4a that have Sau3AI-resistant DNA. In these strains the RM
cassette was in the same genomic location as in the ECI reference
strain F2365. The cassette included a gene encoding a putative
recombinase, suggesting insertion via site-specific recombination.
Deletion of the RM cassette in the ECI strain F2365 and the serotype
1/2a strain A7 rendered the DNA of both strains susceptible to Sau3AI
digestion, providing conclusive evidence that the cassette includes a
gene required for methylation of cytosine at GATC sites in both
strains. The findings suggest that, in addition to its presence in ECI
strains, this RM cassette and the accompanying genomic DNA methylation
is also encountered among selected strains of other lineages.

<>

<1>Yildirim, S., Lin, W., Hitchins, A.D., Jaykus, L.A., Altermann, E., Klaenhammer, T.R., Kathariou, S.
<2>Epidemic clone I-specific genetic markers in strains of Listeria monocytogenes serotype 4b from foods.
<3>Appl. Environ. Microbiol.
<4>70
<5>4158-4164
<6>2004
<7>Listeria monocytogenes contamination of ready-to-eat foods has been implicated in numerous
outbreaks of food-borne listeriosis. However,
the health hazards posed by L. monocytogenes detected in foods may
vary, and speculations exist that strains actually implicated in
illness may constitute only a fraction of those that contaminate foods.
In this study, examination of 34 serogroup 4 (putative or confirmed
serotype 4b) isolates of L. monocytogenes obtained from various foods
and food-processing environments, without known implication in illness,
revealed that many of these strains had methylation of cytosines at
GATC sites in the genome, rendering their DNA resistant to digestion by
the restriction endonuclease Sau3AI. These strains also harbored a gene
cassette with putative restriction-modification system genes as well as
other, genomically unlinked genetic markers characteristic of the major
epidemic-associated lineage of L. monocytogenes (epidemic clone 1),
implicated in numerous outbreaks in Europe and North America. This may
reflect a relatively high fitness of strains with these genetic markers
in foods and food-related environments relative to other serotype 4b
strains and may partially account for the repeated involvement of such
strains in human food-borne listeriosis.

<>

<1>Yin, H., Zhang, X., Liang, Y., Xiao, Y., Niu, J., Liu, X.
<2>Draft Genome Sequence of the Extremophile Acidithiobacillus thiooxidans A01, Isolated from the Wastewater of a Coal Dump.
<3>Genome Announcements
<4>2
<5>e00222-14
<6>2014
<7>The draft genome of Acidithiobacillus thiooxidans A01 contains 3,820,158 bp, with a G+C
content of 53.08% and 3,660 predicted coding sequences (CDSs). The bacterium contains a series
of specific genes involved in the oxidation of elemental sulfur and reduced inorganic sulfur
compounds (RISCs).

<>

<1>Yin, H.S., Zhou, Y.L., Xu, Z.N., Chen, L.J., Zhang, D., Ai, S.Y.
<2>An electrochemical assay for DNA methylation, methyltransferase activity and inhibitor screening based on methyl binding domain protein.
<3>Biosensors and Bioelectronics
<4>41
<5>492-497
<6>2013
<7>DNA methylation is one of important epigenetics events, and responsible to transcription,
genomic imprinting and cellular differentiation.
Aberrant DNA methylation is always contacted with various diseases.
Methyl binding domain (MBD) proteins can specifically bind to the
methylated CpG dinucleotides. Conventional assay for DNA methylation
normally need bisulfide treatment, methylated nucleotide labeling or
PCR amplification. Here, we fabricated a novel electrochemical
biosensor for detection of DNA methylation, assay of DNA
methyltransferase (MTase) activity and screening of MTase inhibitor
based on MBD protein and coomassie brilliant blue G250 (CBB-G250),
where the electrochemical signal of CBB-G250 was used to monitor the
methylation event. After the hybrids of DNA S1 and DNA S2 were treated
with M. SssI MTase in the presence of S-adenosylmethionine, the MBD
proteins were specifically conjugated to the methylation site of CpG
dinucleotides, and then, the MBD proteins were stained with CBB-G250.
The electrochemical signal of CBB-G250 increased linearly with
increasing M. SssI MTase concentration in the range from 0.1 to 40
unit/mL. Furthermore, the inhibition investigation demonstrates that
fisetin and chlorogenic acid can inhibit the M. SssI MTase activity
with the IC50 value of 153.12 and 137.07 mu M, respectively. Therefore,
we think that this study may provide a sensitive platform for screening
of DNA MTase inhibitors.

<>

<1>Yin, J., Chen, J., Liu, G., Yu, Y., Song, L., Wang, X., Qu, X.
<2>Complete Genome Sequence of Glaciecola psychrophila Strain 170T.
<3>Genome Announcements
<4>1
<5>e00199-13
<6>2013
<7>Here, we report the complete genome sequence of Glaciecola psychrophila strain 170(T), a novel
species of the genus Glaciecola, isolated from sea ice at
high-latitude Arctic locations. The genome consists of a single chromosome
(5,413,691 bp) and 5,363 genes. The genomics information will facilitate the
study of the physiology, cold adaptation, and evolution of this genus.

<>

<1>Yin, Y., Withers, T.R., Govan, J.R., Johnson, S.L., Yu, H.D.
<2>Draft Genome Sequence of a Stable Mucoid Strain of Pseudomonas aeruginosa PAO581  with a mucA25 Mutation.
<3>Genome Announcements
<4>1
<5>e00834-13
<6>2013
<7>A mutation in the mucA gene, which encodes a negative regulator of alginate production in
Pseudomonas aeruginosa, is the main mechanism underlying the
conversion to mucoidy in clinical isolates from patients with cystic fibrosis
(CF). Here, we announce the draft genome sequence of the stable
alginate-overproducing mucoid strain P. aeruginosa PAO581 with a mucA25 mutation,
a derivative from the nonmucoid strains P. aeruginosa PAO381 and PAO1.

<>

<1>Yin, Y., Withers, T.R., Johnson, S.L., Yu, H.D.
<2>Draft Genome Sequence of a Mucoid Isolate of Pseudomonas aeruginosa Strain C7447m from a Patient with Cystic Fibrosis.
<3>Genome Announcements
<4>1
<5>e00837-13
<6>2013
<7>Alginate overproduction by Pseudomonas aeruginosa, or mucoidy, plays an important role in the
pathogenesis of chronic lung infections in cystic fibrosis (CF)
patients. Here we report the draft genome sequence of a clinical isolate of
mucoid P. aeruginosa strain C7447m from a CF patient with chronic lung infection.

<>

<1>Yin, Y., Withers, T.R., Niles, R.M., Johnson, S.L., Yu, H.D.
<2>Draft Genome Sequences of Two Alginate-Overproducing Variants of Pseudomonas aeruginosa, PAO1-VE2 and PAO1-VE13.
<3>Genome Announcements
<4>1
<5>e01031-13
<6>2013
<7>The small envelope protein MucE and the sensor kinase KinB are a positive and negative
alginate regulator, respectively. Here, we announce the draft genome
sequences of the alginate-overproducing variants Pseudomonas aeruginosa PAO1-VE2
(PAO1 with constitutive expression of mucE) and PAO1-VE13 (PAO1 with kinB
inactivated). Both mutants were generated from a transposon mutagenesis screen.

<>

<1>Yin, Y., Yue, G., Gao, Q., Wang, Z., Peng, F., Fang, C., Yang, X., Pan, L.
<2>Genome Sequence of Pedobacter arcticus sp. nov., a Sea Ice Bacterium Isolated from Tundra Soil.
<3>J. Bacteriol.
<4>194
<5>6688
<6>2012
<7>Pedobacter arcticus sp. nov. was originally isolated from tundra soil collected from
Ny-Alesund, in the Arctic region of Norway. It is a Gram-negative bacterium
which shows bleb-shaped appendages on the cell surface. Here, we report the draft
annotated genome sequence of Pedobacter arcticus sp. nov., which belongs to the
genus Pedobacter.

<>

<1>Yoder, J.A., Bestor, T.H.
<2>Genetic analysis of genomic methylation patterns in plants and mammals.
<3>Biol. Chem.
<4>377
<5>605-610
<6>1996
<7>While it is now accepted that methylation of cytosine residues plays a role in various
epigenetic phenomena in mammals and flowering plants, the involvement of methylation patterns
in the regulation of normal development has remained a controversial and essentially untested
issue in the 20 years since such a role was first proposed.  Antisense suppression of a DNA
methyltransferase in Arabidopsis and characterization of methylation-defective mutants of
Arabidopsis have shown that perturbations of methylation patterns disrupt the development of
plants, and targeted mutation of the murine gene that encodes the one known form of DNA
methyltransferase has shown that methylation is required for cellular differentiation, genomic
imprinting, and X chromosome inactivation in mammals.  Ectopic expression of homeotic genes
and homeotic transformations of floral organs in methylation-defective plants suggest that (in
plants and perhaps mammals) heritable methylation patterns reinforce and may have supplanted
heritable gene control mediated by chromosomal proteins of the Polycomb and trithorax groups.
It is also possible that the developmental abnormalities are the result of ectopic gene
expression caused by activation of transcription from nearby parasitic sequence elements that
are normally repressed by methylation.  Application of modern methods of genetic analysis
promises to give definite answers to long-standing questions as to the roles and significance
of genomic methylation patterns in normal development and genome defense.

<>

<1>Yoder, J.A., Bestor, T.H.
<2>A candidate mammalian DNA methyltransferase related to pmt1p of fission yeast.
<3>Hum. Mol. Genet.
<4>7
<5>279-284
<6>1998
<7>Trace levels of 5-methylcytosine persist in the DNA of mouse embryonic stem cells that are
homozygous for null mutations in Dnmt1, the gene for the one previously recognized metazoan
DNA methyltransferase.  This residual 5-methylcytosine may be the product of a candidate
second DNA methyltransferase, Dnmt2, that has now been identified in human and mouse.  Dnmt2
contains all the sequence motifs diagnostic of DNA (cytosine-5)-methyltransferases but appears
to lack the large N-terminal regulatory domain common to other eukaryotic methyltransferases.
Dnmt2 is more similar to a putative DNA methyltransferase of the fission yeast
Schizosaccharomyces pombe than to Dnmt1.  Dnmt2 produces multiple mRNA species that are
present at low levels in all tissues of human and mouse and is not restricted to those cell
types known to be active in de novo methylation.  The human DNMT2 gene was mapped to
chromosome 10p12-10p14 in a panel of radiation hybrids.  Dnmt2 is a candidate for the activity
that methylates newly integrated retroviral DNA and maintains trace levels of 5-methylcytosine
in the DNA of embryonic stem cells homozygous for null mutations in Dnmt1.

<>

<1>Yoder, J.A., Soman, N.S., Verdine, G.L., Bestor, T.H.
<2>DNA (cytosine-5)-methyltransferases in mouse cells and tissues.  Studies with a mechanism-based probe.
<3>J. Mol. Biol.
<4>270
<5>385-395
<6>1997
<7>The mechanisms that establish and maintain methylation patterns in the mammalian genome are
very poorly understood, even though perturbations of methylation patterns lead to a loss of
genomic imprinting, ectopic X chromosome inactivation, and death of mammalian embryos.  A
family of sequence-specific DNA methyltransferases has been proposed to be responsible for the
wave of de novo methylation that occurs in the early embryo, although no such enzyme has been
identified.  A universal mechanism-based probe for DNA (cytosine-5)-methyltransferases was
used to screen tissues and cell types known to be active in de novo methylation for new
species of DNA methyltransferase.  All identifiable de novo methyltransferase activity was
found to reside in Dnmt1.  As this enzyme is the predominant de novo methyltransferase at all
developmental stages inspected, it does not fit the definition of maintenance
methyltransferase or hemimethylase.  Recent genetic data indicate that de novo methylation of
retroviral DNA in embryonic stem cells is likely to involve one or more additional DNA
methyltransferases.  Such enzymes were not detected and are either present in very small
amounts or are very different from Dnmt1.  A new method was developed and used to determine
the sequence specificity of intact Dnmt1 in whole-cell lysates.  Specificity was found to be
confined to the sequence 5'-CpG-3'; there was little dependence on sequence context or
density of CpG dinucleotides.  These data suggest that any sequence-specific de novo
methylation mediated by Dnmt1 is either under the control of regulatory factors that interact
with Dnmt1, or is cued by alternative secondary structures in DNA.

<>

<1>Yoder, J.A., Walsh, C.P., Bestor, T.H.
<2>Cytosine methylation and the ecology of intragenomic parasites.
<3>Trends Genet.
<4>13
<5>335-340
<6>1997
<7>Most of the 5-methylcytosine in mammalian DNA resides in transposons, which are specialized
intragenomic parasites that represent at least 35% of the genome.  Transposon promoters are
inactive when methylated and, over time, C-T transition mutations at methylated sites destroy
many transposons.  Apart from that subset of genes subject to X inactivation and genomic
imprinting, no cellular gene in a non-expressing tissue has been proven to be methylated in a
pattern that prevents transcription.  It has become increasingly difficult to hold that
reversible promoter methylation is commonly involved in developmental gene control; instead,
suppression of parasitic sequence elements appears to be the primary function of cytosine
methylation, with crucial secondary roles in allele-specific gene expression as seen in X
inactivation and genomic imprinting.

<>

<1>Yoder, J.A., Yen, R.-W.C., Vertino, P.M., Bestor, T.H., Baylin, S.B.
<2>New 5' regions of the murine and human genes for DNA (cytosine-5)-methyltransferase.
<3>J. Biol. Chem.
<4>271
<5>31092-31097
<6>1996
<7>DNA (cytosine-5)-methyltransferases (EC 2.1.1.37)  maintain patterns of methylated cytosine
residues in the mammalian genome; faithful maintenance of methylation patterns is required for
normal development of mice, and aberrant methylation patterns are associated with certain
human tumors and developmental abnormalities.  The organization of coding sequences at the
5'-end of the murine and human DNA methyltransferase genes was investigated, and the DNA
methyltransferase open reading frame was found to be longer than previously suspected.
Expression of the complete open reading frame by in vitro transcription-translation and by
transfection of expression constructs into COS7 cells resulted in the production of an active
DNA methyltransferase of the same apparent mass as the endogenous protein, while translation
from the second inframe ATG codon produced a slightly smaller but fully active protein.
Characterization of mRNA 5' sequences and the intron-exon structure of the 5' region of the
murine and human genes indicated that a previously described promoter element actually lies in
an intron that is more than 5 kilobases downstream of the transcription start sites.

<>

<1>Yokochi, T., Robertson, K.D.
<2>Preferential methylation of unmethylated DNA by mammalian de novo DNA methyltransferase Dnmt3a.
<3>J. Biol. Chem.
<4>277
<5>11735-11745
<6>2002
<7>DNA methylation is an epigenetic modification of DNA. There are currently three catalytically
active mammalian DNA methyltransferases,
DNMT1, -3a, and -3b. DNMT1 has been shown to have a preference for
hemimethylated DNA and has therefore been termed the maintenance
methyltransferase. Although previous studies on DNMT3a and -3b revealed
that they act as functional enzymes during development, there is little
biochemical evidence about how new methylation patterns are established
and maintained. To study this mechanism we have cloned and expressed
Dnmt3a using a baculovirus expression system. The substrate specificity
of Dnmt3a and molecular mechanism of its methylation reaction were then
analyzed using a novel and highly reproducible assay. We report here
that Dnmt3a is a true de novo methyltransferase that prefers
unmethylated DNA substrates more than 3-fold to hemimethylated DNA.
Furthermore, Dnmt3a binds DNA nonspecifically, regardless of the
presence of CpG dinucleotides in the DNA substrate. Kinetic analysis
supports an Ordered Bi Bi mechanism for Dnmt3a, where DNA binds first,
followed by S-adenoSyl-L-methionine.

<>

<1>Yokochi, T., Robertson, K.D.
<2>Dimethyl sulfoxide stimulates the catalytic activity of de novo DNA methyltransferase 3a (Dnmt3a) in vitro.
<3>Bioorg. Chem.
<4>32
<5>234-243
<6>2004
<7>Mammalian DNA methyltransferase Dnmt3a is required for de novo methylation of CpG
dinucleotides in genomic DNA. While DNA
methyltransferase inhibitors have been extensively utilized both in
vitro and in vivo, no stimulator of catalytic activity has been
identified thus far. Here we show that the methyltransfer activity of
Dnmt3a is stimulated by the addition of dimethyl sulfoxide (DMSO) to
the reaction solution in vitro. Enzymatic analysis of initial reaction
velocity suggests that the DMSO stimulation effect depends on the
interaction between DMSO and the reaction substrates (DNA and AdoMet),
but not the enzyme itself.

<>

<1>Yokochi, T., Robertson, K.D.
<2>DMB (DNMT-magnetic beads) assay: Measuring DNA methyltransferase activity in vitro.
<3>Methods Mol. Biol.
<4>287
<5>285-296
<6>2004
<7>DNA methylation is an epigenetic modification of DNA that leads to heritable alterations in
transcriptional regulation and conformational changes in chromatin structure of higher
eukaryotes.  Mammalian DNA methyltransferases, which are the enzymes responsible for DNA
methylation, have attracted the attention of both basic and clinical researchers because they
appear to participate in embryogenesis and carcinogenesis via chromatin modification.  DNA
methyltransferase catalyzes the traansfer of a methyl group into DNA strands.  Since
traditional assays for DNA methyltransferase activity in vitro have insufficient
reproducibility, there is a need in the art for more sensitive and quantitative methods for
measuring enzymatic activity.  We report a novel assay system, in which the actiity of a DNA
methyltransferase is measured as the incorporation of tritium into biotinylated DNA
oligonucleotides.  The DNA is immobilized onto magnetic beads with streptavidin covalently
attached to the bead surface.  The radioactive DNA can easily be separated from the unreacted
radioactive substrate using a magnet.  The radioactivity is counted by the liquid
scintillation system.  This DMB assay is simple and easy, has very low background, and, most
importantly, is highly reproducible for the precise enzymatic analysis of any DNA
methyltransferase in vitro.

<>

<1>Yokoyama, K., Makino, K., Kubota, Y., Watanabe, M., Kimura, S., Yutsudo, C.H., Kurokawa, K., Ishii, K., Hattori, M., Tatsuno, I., Abe, H., Yoh, M., Iida, T., Ohnishi, M., Hayashi, T., Yasunaga, T., Honda, T., Sasakawa, C., Shinagawa, H.
<2>Complete nucleotide sequence of the prophage VT1-Sakai carrying the Shiga toxin 1 genes of the enterohemorrhagic Escherichia coli O157:H7 strain derived from the Sakai outbreak.
<3>Gene
<4>258
<5>127-139
<6>2000
<7>Shiga toxins 1 and 2 (Stx1 and Stx2) are encoded by prophages lysogenized in enterohemorrhagic
Escherichia coli (EHEC) O157:H7 strains. Lytic growth of the phage particles carrying the stx1
genes (stx1A and stx1B) of the EHEC O157:H7 strain RIMD 0509952, which was derived from the
Sakai outbreak in 1996 in Japan, was induced after treatment with mitomycin C, but the plaque
formation of the phage was not detected. We have determined the complete nucleotide sequence
of the prophage VT1-Sakai. The integration site of the prophage was identified within the yehV
gene at 47.7 min on the chromosome. The stx1 genes were downstream of the Q gene in the
prophage genome, suggesting that their expression was regulated by the Q protein, the
regulator of the late gene expression of the phage, which is similar to that of the stx1 or
stx2 genes carried by the lambdoid phages reported previously. The sequences of the N gene and
its recognition sites, nutL and nutR, were not homologous to those of the phages carrying the
stx genes thus far reported, but they were very similar to those of bacteriophage phi21. The
sequences of the repressor proteins, CI and Cro, that regulate expression of the early genes
had low similarities with those of the known repressors of other phages, and their operator
sequences were different from any sequence reported. These data suggest that multiple genetic
recombination among bacteriophages with different immunities took place to generate the
prophage VT1-Sakai. Comparison between the sequences of VT1-Sakai and lambda suggests that the
ancestor of VT1-Sakai was produced by illegitimate excision, like lambda gal and bio phages.
Full author list: Yokoyama, K., Makino, K., Kubota, Y., Watanabe, M., Kimura, S., Yutsudo,
C.H., Kurokawa, K., Ishii, K., Hattori, M., Tatsuno, I., Abe, H., Yoh, M., Iida, T., Ohnishi,
M., Hayashi, T., Yasunaga, T., Honda, T., Sasakawa, C., Shinagawa,H.

<>

<1>Yokoyama, S., Oshima, K., Nomura, I., Hattori, M., Suzuki, T.
<2>Complete Genomic Sequence of the O-Desmethylangolensin-Producing Bacterium Clostridium rRNA Cluster XIVa Strain SY8519, Isolated from Adult Human  Intestine.
<3>J. Bacteriol.
<4>193
<5>5568-5569
<6>2011
<7>The O-desmethylangolensin-producing Clostridium rRNA cluster XIVa strain SY8519 was isolated
from the intestinal flora of a healthy human as a key
isoflavonoid-metabolizing bacterium. Here, we report the finished and
annotated genomic sequence of this organism.

<>

<1>Yokoyama, S., Oshima, K., Nomura, I., Hattori, M., Suzuki, T.
<2>Complete Genomic Sequence of the Equol-Producing Bacterium Eggerthella sp. Strain YY7918, Isolated from Adult Human Intestine.
<3>J. Bacteriol.
<4>193
<5>5570-5571
<6>2011
<7>Eggerthella sp. strain YY7918 was isolated from the intestinal flora of a healthy human. It
metabolizes daidzein (a soybean isoflavonoid) and
produces S-equol, which has stronger estrogenic activities than daidzein.
Here, we report the finished and annotated genomic sequence of this
organism.

<>

<1>Yolov, A.A., Gromova, E.S., Potapov, V.K., Shabarova, Z.A.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments.  V.  Study of single-stranded cleavages.
<3>Nucleic Acids Res.
<4>13
<5>8969-8981
<6>1985
<7>Concatemer DNA duplexes which contain at the EcoRII restriction endonuclease
cleavage sites ...^C-C-A-G-G-......-G-G-T-C-C^... phosphodiester, phosphoamide
or pyrophosphate internucleotide bonds have been synthesized.  It has been
shown that this enzyme did not cleave the substrate at phosphoamide bond.
EcoRII endonuclease catalyzes single-strand cleavages both in dA- and
dT-containing strands of the recognition site if the cleavage of the other
strand has been blocked by modification of scissile bond or if the other strand
has been cleaved.  This enzyme interacts with both strands of the DNA
recognition site, each of them being cleaved independently of the cleavage of
another one.  Nucleotide sequences flanking the EcoRII site on both sides are
necessary for effective cleavage of the substrate.

<>

<1>Yolov, A.A., Gromova, E.S., Romanova, E.A., Oretskaya, T.S., Oganov, A.A., Buryanov, Y.I., Shabarova, Z.A.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments.
<3>FEBS Lett.
<4>167
<5>147-150
<6>1984
<7>Interaction of EcoRII restriction endonuclease with a set of synthetic
concatemer DNA duplexes with natural and modified sites for this enzyme has
been studied.  DNA duplexes with repeated natural sites are cleaved by EcoRII.
Substitution of central AT-pair in the recognition site for a non-complementary
TT- or AA-pair reduces the rate of cleaveage, this effect being much more
pronounced in the last case.  Absence of site flanking in one strand from the
5' -terminus also results in very slow cleavage.  The results obtained testify
to the interaction of EcoRII with both strands of the substrate.

<>

<1>Yolov, A.A., Gromova, E.S., Shabarova, Z.A.
<2>Processive cleavage of concatemer DNA duplexes by EcoRII restriction endonuclease.
<3>Mol. Biol. Rep.
<4>10
<5>173-176
<6>1985
<7>EcoRII restriction endonuclease cleaves synethetic DNA-duplexes in which the
recognition sites of this enzyme (5'...CC(A/T)GG...) are repeated every 9 base
pairs with the alternating orientation of the central AT pair.  It operates in
a processive mode, i.e. the bound enzyme molecule slides along the substrate
toward neighboring recognition sites.  Nona-nucleotides are the main products
of the cleavage.  The data obtained point to the capability of EcoRII
endonuclease to recognize and cleave the substrate under both possible
orientations of the central AT-pair of the recognition site with respect to the
bound enzyme molecule.  These data also show the close similarity of DNA
structures in a complex with the enzyme and without.

<>

<1>Yolov, A.A., Vinogradova, M.N., Gromova, E.S., Rosenthal, A., Cech, D., Veiko, V.P., Metelev, V.G., Kosykh, V.G., Buryanov, Y.I., Vayev, A.A., Shabarova, A.Z.
<2>Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments. VI.  The binding and cleavage of substrates containing nucleotide analogs.
<3>Nucleic Acids Res.
<4>13
<5>8983-8998
<6>1985
<7>The present study deals with the binding and cleavage by EcoRII endonuclease of
concatamer DNA duplexes containing EcoRII recognition sites (5'...^CCAGG) in
which dT is replaced by dU or 5-bromodeoxyuridine, or 5'-terminal dC in the
dT-containing strand is methylated at position 5.  The enzyme molecule is found
to interact with the methyl group of the dT residue of the DNA recognition site
and to be at least in proximity to the H5 atom of the 5'-terminal dC residue in
dT-containing strand of this site.  Modification of any of these positions
exerts an equal effects on the cleavage of both DNA strands.  Endonuclease
EcoRII was found to bind the substrate specifically.  At the same time
modification of the bases in recognized sequence may result in the formation of
unproductive,though stable, enzyme-substrate complexes.

<>

<1>Yonezawa, A., Sugiura, Y.
<2>DNA binding mode of class-IIS restriction endonuclease FokI revealed by DNA footprinting analysis.
<3>Biochim. Biophys. Acta
<4>1219
<5>369-379
<6>1994
<7>We investigate the interaction of FokI with its DNA recognition sequence by several
footprinting techniques. Methylation of three guanine bases in the recognition sequence
5'-GGATG-3' is strongly protected by FokI binding, whereas other guanine bases are not
masked from the modification. In footprinting using the methidiumpropyl-EDTA-Fe(II) complex,
binding of FokI strongly inhibits cleavage by the footprinting reagent at and near the
recognition sequence. In high-resolution footprinting techniques using hydroxyl radical and
the bleomycin-Fe(II) complex, all footprints in each binding site clearly face one side of the
DNA helix. Interference analysis with FokI digestion by preethylation of phosphate groups
suggests that essential phosphates for FokI digestion are located at and near the recognition
sequence and the cleavage site. Evidently, the results indicate that (i) the
sequence-recognition of FokI occurs in the major groove and that (ii) the enzyme interacts
with its target DNA from one side of the DNA helix.

<>

<1>Yong, B., Yang, B.Q., Zhao, C.W., Feng, H.
<2>Draft Genome Sequence of Bacillus subtilis Strain S1-4, Which Degrades Feathers Efficiently.
<3>Genome Announcements
<4>1
<5>e00766-13
<6>2013
<7>Bacillus subtilis strain S1-4, with the capacity to efficiently degrade feathers, was isolated
from chicken feathers. Sequencing showed that the genome of strain
S1-4 differs from that of other B. subtilis strains, with limited insertions and
deletions. The genome encodes multiple extracellular proteases and keratinases.

<>

<1>Yoo, H.Y., Noshari, J., Lapeyre, J.N.
<2>Subunit and functional size of human placental DNA methyltransferase involved in de novo and maintenance methylation.
<3>J. Biol. Chem.
<4>262
<5>8066-8070
<6>1987
<7>The subunit molecular size of human DNA methyltransferase isolated from nuclear extracts of
placenta was determined on the electroblotted
polypeptides after sodium dodecyl sulfate-polyacrylamide gel
electrophoresis and compared with the functional size by high performance
size exclusion chromatography on Superose 12 and gamma radiation
inactivation analysis. The sodium dodecyl sulfate-polyacrylamide gel
electrophoresis results indicated a subunit mass of 120 +/- 10 kDa, while
the functional size data indicates that the enzyme operates both in de
novo and maintenance modes as a dimer of molecular mass 220 +/- 15 kDa
with no evidence of monomers in solution of ionic strength between 0.1 and
0.8 M NaCl. The 220-kDa activity carried out the transmethylation of both
hemi- and unmethylated DNA substrates. There was no evidence for separate
functional catalytic sites on each monomer subunit acting independently
when engaged in methylation of hemimethylated or single-stranded DNA from
the invariance of radiation inactivation target size with these
substrates. The radiation inactivation target size was 230 +/- 15 kDa.

<>

<1>Yoo, J., Choi, S., Medina-Franco, J.L.
<2>Molecular Modeling Studies of the Novel Inhibitors of DNA Methyltransferases SGI-1027 and CBC12: Implications for the Mechanism of Inhibition of DNMTs.
<3>PLoS ONE
<4>8
<5>e62152
<6>2013
<7>DNA methylation is an epigenetic modification that regulates gene expression by DNA
methyltransferases (DNMTs). Inhibition of DNMTs is a
promising approach for cancer therapy. Recently, novel classes of the
quinolone-based compound, SGI-1027, and RG108-procainamide conjugates,
CBC12, have been identified as potent DNMT inhibitors. In this work, we
report comprehensive studies using induced-fit docking of SGI-1027 and
CBC12 with human DNMT1 and DNMT3A. The docking was performed in the
C-terminal MTase catalytic domain, which contains the substrate and
cofactor binding sites, in the presence and absence of other domains.
Induced-fit docking predicts possible binding modes of the ligands
through the appropriate structural changes in the receptor. This work
suggests a hypothesis of the inhibitory mechanisms of the new
inhibitors which is in agreement with the reported autoinhibitory
mechanism. The insights obtained in this work can be used to design
DNMT inhibitors with novel scaffolds.

<>

<1>Yoo, J., Kim, J.H., Robertson, K.D., Medina-Franco, J.L.
<2>MOLECULAR MODELING OF INHIBITORS OF HUMAN DNA METHYLTRANSFERASE WITH A CRYSTAL STRUCTURE: DISCOVERY OF A NOVEL DNMT1 INHIBITOR.
<3>Adv. Protein Chem. Struct. Biol.
<4>87
<5>219-247
<6>2012
<7>DNA methyltransferases (DNMTs) are promising epigenetic targets for the development of novel
anticancer drugs and other diseases. Molecular
modeling and experimental approaches are being used to identify and
develop inhibitors of human DNMTs. Most of the computational efforts
conducted so far with DNMT1 employ homology models of the enzyme.
Recently, a crystallographic structure of the methyltransferase domain
of human DNMT1 bound to unmethylated DNA was published. Following on
our previous computational and experimental studies with DNMTs, we
herein present molecular dynamics of the crystal structure of human
DNMT1. Docking studies of established DNMT1 inhibitors with the crystal
structure gave rise to a structure-based pharmacophore model that
suggests key interactions of the inhibitors with the catalytic binding
site. Results had a good agreement with the docking and pharmacophore
models previously developed using a homology model of the catalytic
domain of DNMT1. The docking protocol was able to distinguish active
DNMT1 inhibitors from, for example, experimentally known inactive DNMT1
inhibitors. As part of our efforts to identify novel inhibitors of
DNMT1, we conducted the experimental characterization of
aurintricarboxylic acid (ATA) that in preliminary docking studies
showed promising activity. ATA had a submicromolar inhibition
(IC50=0.68 mu M) against DNMT1. ATA was also evaluated for Dnmt3a
inhibition showing an IC50=1.4 mu M. This chapter illustrates the
synergy from integrating molecular modeling and experimental methods to
further advance the discovery of novel candidates for epigenetic
therapies.

<>

<1>Yoo, J., Medina-Franco, J.L.
<2>Inhibitors of DNA Methyltransferases: Insights from Computational Studies.
<3>Curr. Med. Chem.
<4>19
<5>3475-3487
<6>2012
<7>DNA methyltransferases (DNMTs) are a family of epigenetic enzymes for which inhibition is an
attractive strategy for the treatment of cancer
and other diseases. In synergy with experimental approaches,
computational methods are increasingly being used to identify and
optimize the activity of inhibitors of DNMTs as well as to rationalize
at the molecular level of the mechanism of established inhibitors.
Recently, a crystallographic structure of the methyltransferase domain
of human DNMT1 bound to unmethylated DNA was published encouraging the
application of structure-based approaches to design and optimize the
activity of currently known inhibitors. Herein, we review the progress
in the discovery and optimization of inhibitors of DNMTs using
computational approaches including homology modeling, docking,
pharmacophore modeling, molecular dynamics, and virtual screening.

<>

<1>Yoo, M., Kim, D., Choi, K.Y., Chae, J.C., Zylstra, G.J., Kim, E.
<2>Draft Genome Sequence and Comparative Analysis of the Superb Aromatic-Hydrocarbon Degrader Rhodococcus sp. Strain DK17.
<3>J. Bacteriol.
<4>194
<5>4440
<6>2012
<7>Rhodococcus sp. strain DK17 is capable of utilizing various derivatives of benzene and
bicyclics containing both aromatic and alicyclic moieties as sole
carbon and energy sources. Here, we present the 9,107,362-bp draft genome
sequence of DK17 and its genomic analysis in comparison with other members of the
genus Rhodococcus.

<>

<1>Yoo, O.J., Agarwal, K.L.
<2>Isolation and characterization of two proteins possessing HpaII methylase activity.
<3>J. Biol. Chem.
<4>255
<5>6445-6449
<6>1980
<7>Two proteins exhibiting HpaII methylase activity have been purified to
homogeneity from Haemophilus parainfluenzae and their physical and catalytic
properties have been studied.  Separation of the two HpaII methylase activities
was achieved by DEAE-Sephadex A-50 chromatography.  In subsequent steps, each
methylase was purified separately by chromatography on Sephacryl S-200,
phosphocellulose, and hydroxylapatite.  The proteins have molecular weights of
38,500 +/- 1,000 (HpaII) and 41,500 +/- 1,000 (HpaII') as judged by
polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate.
Sedimentation equilibrium analyses of the native proteins yield molecular
weights of 38,800 +/- 3,000 and 42,200 +/- 3,000 for HpaII and HpaII',
respectively, indicating that both enzymes are composed of a single subunit.
Furthermore, both methylases exhibit identical specificity in the methylation
of the nucleotide sequence dC-C-G-G in simian virus 40 (SV40) DNA and in a
short synthetic oligonucleotide duplex.  Although pH, temperature, and salt
optima are the same for both enzymes, homogenous HpaII' methylase is more
stable than HpaII methylase.  Preliminary peptide mapping indicates that the
two enzymes are structurally related, suggesting the possibility that HpaII'
methylase may represent a precursor form of HpaII methylase.

<>

<1>Yoo, O.J., Agarwal, K.L.
<2>Cleavage of single strand oligonucleotides and bacteriophage PhiX174 DNA by MspI endonuclease.
<3>J. Biol. Chem.
<4>255
<5>10559-10562
<6>1980
<7>Type II restriction endonucleases cleave duplex DNA at nucleotide sequences
displaying 2-fold symmetry.  Our data show that MspI cleaves single strand
oligonucleotides, d(GAACCGGAGA) and d(TCTCGGTT) at 4, 25, and 37C reaction
temperatures.  The rate of cleavage of d(GAACCGGAGA) is several-fold faster
than that of d(TCTCCGGTT).  Single strand PhiX174 DNA is also cleaved by MspI
endonuclease giving well defined fragments.  5'-nucleotide analysis of the
fragments generated from single strand and replicating form DNA suggest that
cleavage occurs at the recognition sequence d(CCGG).  The data show that MspI
endonuclease cleaves single strand oligonucleotides and prefers a recognition
sequence surrounded by purine nucleotides.  A general model for endonuclease
cleavage of single strand and duplex DNA is presented.

<>

<1>Yoo, O.J., Dwyer-Hallquist, P., Agarwal, K.L.
<2>Purification and properties of the HpaI methylase.
<3>Nucleic Acids Res.
<4>10
<5>6511-6519
<6>1982
<7>The purification and catalytic properties of the homogeneous HpaI methylase is
described.  The enzyme exists as a single polypeptide chain with a molecular
weight of 37,000 +/- 2,000 was shown by sedimentation equilibrium and
polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate.
The HpaI methylase transfers methyl groups of S-adenosylmethionine to adenine
present in the recognition sequence d(G-T-T-A-A*-C), A* is the N6 methyl
adenosine.  An average of 2.1 methyl groups per recognition site are
transferred by the HpaI methylase.

<>

<1>Yoon, H., Suh, H., Han, M.H., Yoo, O.J.
<2>Purification and characterization of AluI methylase.
<3>Korean Biochem. J.
<4>18
<5>82-87
<6>1985
<7>AluI methylase has been isolated from 300g (wet weight) cells of Arthrobacter
luteus.  After ammonium sulfate fractionation, the protein which has methylase
activity was purified through phosphocellulose, DEAE-cellulose, Heparin
agarose, and Hydroxy-lapatite column chromatography.  The methylated DNA by the
purified methylase was resistant against AluI endonuclease.  The purified AluI
methylase was essentially homogeneous as judged by 10% SDS-polyacrylamide gel
electrophoresis, and the apparent subunit molecular weight was 56,000+-1,000.
The specific activity of the enzyme was 132000 units per mg protein.

<>

<1>Yoon, H., Suh, H., Kim, K., Han, M.H., Yoo, O.J.
<2>The specificity and catalytic properties of AluI methylase.
<3>Korean Biochem. J.
<4>18
<5>88-93
<6>1985
<7>The specific methylation site for AluI methylase was the cytosine nucleotide in
AluI sequence.  The position of the methylated cytosine nucleotide was
determined by the chemical cleavage reactions of the Maxam-Gilbert DNA
sequencing procedure.  As expected, the methylated cytosine nucleotide bands
were disappeared on C+T and C lanes on 12% sequencing gels. AluI methylase was
maximally active at near pH 7.5 in the presence of 50 mM NaCl.  The methylase
did not require Mg++ for activity, and obeyed Michaelis-Menten Kinetics with
respect to both AdoMet and DNA.  At 37C, the Km for AdoMet was 0.44 lM, that
for the AluI site of pBR322 DNA was 4.03 nM, and the corresponding turnover
numbers were 1.83 methyl transfer per minute per monomer and 1.61 transfers per
minute per monomer, respectively.

<>

<1>Yoon, H.J., Kang, K.Y., Ahn, H.J., Shim, S.M., Ha, J.Y., Lee, S.K., Mikami, B., Suh, S.W.
<2>X-ray crystallographic studies of HemK from Thermotoga maritima, an N-5-glutamine methyltransferase.
<3>Mol. Cells
<4>16
<5>266-269
<6>2003
<7>The enzyme HemK (or PrmC) is one of the first identified methyltransferases that modify
glutamine. It methylates the highly
conserved GGQ motif in class I release factors (RF1 and RF2) in
Escherichia coli. HemK from Thermotoga maritima was over-expressed and
crystallized in the presence of S-adenosylmethionine at 296 K using
ammonium sulfate as the precipitant. X-ray diffraction data were collected
to 2.5 A resolution from a native crystal. The crystal is orthorhombic,
belonging to the space group I222 (or I2(1)2(1)2(1)), with unit-cell
parameters of a = 104.24, b = 118.73, and c = 146.62 A. Two (or three)
monomers of recombinant HemK are likely to be present in the
crystallographic asymmetric unit, giving a V(M) of 3.62 A3 Da(-1) (or 2.41
A3 Da(-1)), with a solvent content of 62.7% (or 44.0%).

<>

<1>Yoon, H.S., Kang, S.C., Yoo, O.J.
<2>Purification and characterization of HpaI endonuclease.
<3>Sanop Misaengmul Hakhoe Chi
<4>13
<5>87-91
<6>1985
<7>HpaI endonuclease from Haemophilus parainfluenzae has been purified to
homogeneity and its physical and enzymatic properties have been studied.  For
the purification of the enzyme, Heparin agarose, SP-sephadex C-25,
DEAE-sephadex A-50 and phosphocellulose chromatography columns were used.  The
denatured and reduced form of the enzyme is a monomer of molecular weight of
30,000 -+1,000 as judged by 10% polyacrylamide gel electrophoresis containing
0.1% sodium dodesyl sulfate.  HpaI endonuclease was maximally active at neutral
pH (7.0 to 7.5) in the presence of 50 mM NaCl.

<>

<1>Yoon, M.Y., Lee, K.M., Yoon, Y., Go, J., Park, Y., Cho, Y.J., Tannock, G.W., Yoon, S.S.
<2>Functional screening of a metagenomic library reveals operons responsible for enhanced intestinal colonization by gut commensal microbes.
<3>Appl. Environ. Microbiol.
<4>79
<5>3829-3838
<6>2013
<7>Evidence suggests that gut microbes colonize the mammalian intestine through
propagation as an adhesive microbial community. A bacterial artificial chromosome
(BAC) library of murine bowel microbiota DNA in the surrogate host Escherichia
coli DH10B was screened for enhanced adherence capability. Two out of 5,472 DH10B
clones, 10G6 and 25G1, exhibited enhanced capabilities to adhere to inanimate
surfaces in functional screens. DNA segments inserted into the 10G6 and 25G1
clones were 52 and 41 kb and included 47 and 41 protein-coding open reading
frames (ORFs), respectively. DNA sequence alignments, tetranucleotide frequency,
and codon usage analysis strongly suggest that these two DNA fragments are
derived from species belonging to the genus Bacteroides. Consistent with this
finding, a large portion of the predicted gene products were highly homologous to
those of Bacteroides spp. Transposon mutagenesis and subsequent experiments that
involved heterologous expression identified two operons associated with enhanced
adherence. E. coli strains transformed with the 10a or 25b operon adhered to the
surface of intestinal epithelium and colonized the mouse intestine more
vigorously than did the control strain. This study has revealed the genetic
determinants of unknown commensals (probably resembling Bacteroides species) that
enhance the ability of the bacteria to colonize the murine bowel.

<>

<1>Yooseph, S. et al.
<2>The Sorcerer II Global Ocean Sampling Expedition: Expanding the Universe of Protein Families.
<3>PLoS Biology
<4>5
<5>e16
<6>2007
<7>Metagenomics projects based on shotgun sequencing of populations of micro-organisms yield
insight into protein families. We used sequence
similarity clustering to explore proteins with a comprehensive dataset
consisting of sequences from available databases together with 6.12
million proteins predicted from an assembly of 7.7 million Global Ocean
Sampling (GOS) sequences. The GOS dataset covers nearly all known
prokaryotic protein families. A total of 3,995 medium- and large-sized
clusters consisting of only GOS sequences are identified, out of which
1,700 have no detectable homology to known families. The GOS-only clusters
contain a higher than expected proportion of sequences of viral origin,
thus reflecting a poor sampling of viral diversity until now. Protein
domain distributions in the GOS dataset and current protein databases show
distinct biases. Several protein domains that were previously categorized
as kingdom specific are shown to have GOS examples in other kingdoms.
About 6,000 sequences (ORFans) from the literature that heretofore lacked
similarity to known proteins have matches in the GOS data. The GOS dataset
is also used to improve remote homology detection. Overall, besides nearly
doubling the number of current proteins, the predicted GOS proteins also
add a great deal of diversity to known protein families and shed light on
their evolution. These observations are illustrated using several protein
families, including phosphatases, proteases, ultraviolet-irradiation DNA
damage repair enzymes, glutamine synthetase, and RuBisCO. The diversity
added by GOS data has implications for choosing targets for experimental
structure characterization as part of structural genomics efforts. Our
analysis indicates that new families are being discovered at a rate that
is linear or almost linear with the addition of new sequences, implying
that we are still far from discovering all protein families in nature.

<>

<1>Yoshida, Y., Mise, K.
<2>Occurrence of small Hsd plasmids in Salmonella typhi, Shigella boydii, and Escherichia coli.
<3>J. Bacteriol.
<4>165
<5>357-362
<6>1986
<7>The natural occurrence of small Hsd (host specificity for DNA) plasmids was
demonstrated in restriction endonuclease-producing strains of Salmonella typhi,
Shigella boydii, and Escherichia coli.  The five Hsd plasmids isolated were
between 5.0 and 12.2 kilobases long.  The copy number of all the Hsd plasmids
were high (more than 10 copies per cell).  Introduction of these small plasmids
into E. coli strain 0 drastically lowered the efficiency of plating of the k.0
phages (the efficiency of plating was less than 5 X 10-5 PFU-1).  High
restriction endonuclease activities were detected in the Hsd plasmid-positive
strains because of the elevated copy numbers of the hsdR+ gene.  The advantages
of using E. coli strains containing the small Hsd plasmids for purification of
type II restriction endonucleases are discussed.

<>

<1>Yoshii, A., Omatsu, T., Katayama, Y., Koyama, S., Mizutani, T., Moriyama, H., Fukuhara, T.
<2>Two types of genetic carriers, the IncP genomic island and the novel IncP-1beta plasmid, for the aac(2')-IIa gene that confers kasugamycin resistance in Acidovorax avenae subsp. avenae.
<3>Mol. Plant Pathol.
<4>0
<5>0
<6>2014
<7>A unique aminoglycoside antibiotic, kasugamycin (KSM), has been used to control
many plant bacterial and fungal diseases in several countries. The emergence of
KSM-resistant Acidovorax avenae ssp. avenae and Burkholderia glumae, which cause
rice bacterial brown stripe and rice bacterial grain and seedling rot,
respectively, is a serious threat for the effective control of these diseases.
Previously, we have identified the aac(2')-IIa gene, encoding a KSM
2'-N-acetyltransferase, from both KSM-resistant pathogens. Although all
KSM-resistant isolates from both species possess the aac(2')-IIa gene, only A.
avenae strain 83 showed higher resistance than other strains. In this research,
kinetic analysis indicates that an amino acid substitution from serine to
threonine at position 146 of AAC(2')-IIa in strain 83 is not involved in this
increased resistance. Whole draft genome analysis of A. avenae 83 shows that the
aac(2')-IIa gene is carried by the novel IncP-1beta plasmid pAAA83, whereas the
genetic carrier of other strains, the IncP genomic island, is inserted into their
chromosomes. The difference in the nucleotides of the promoter region of
aac(2')-IIa between strain 83 and other strains indicates an additional
transcription start site and results in the increased transcription of
aac(2')-IIa in strain 83. Moreover, biological characterization of pAAA83
demonstrates that it can be transferred by conjugation and maintained in the host
cells. These results demonstrate that acquisition of the aac(2')-IIa gene takes
place in at least two ways and that the gene module, which includes aac(2')-IIa
and the downstream gene, may be an important unit for the dissemination of
antibiotic resistance.

<>

<1>Yoshimori, R., Roulland-Dussoix, D., Boyer, H.W.
<2>R factor-controlled restriction and modification of deoxyribonucleic acids:  restriction mutants.
<3>J. Bacteriol.
<4>112
<5>1275-1279
<6>1972
<7>Restriction mutants of two different R factor-controlled host specificities (RI
and RII) were isolated.  All of the restriction mutants examined had a normal
modification phenotype.  No complementation was observed between the RI and RII
host specificities.  It is concluded that for each host specifcity no protein
subunit is shared by the restriction endonuclease and modification methylase.

<>

<1>Yoshimori, R.N.
<2>A genetic and biochemical analysis of the restriction and modification of DNA by resistance transfer factors.
<3>Ph.D. Thesis, University of California, San Francisco, USA
<4>
<5>1-73
<6>1971
<7>The DNA restriction and modification systems of RTF-1 and RTF-2, host
specificities RI and RII respectively, were studied genetically and
biochemically.  The presence of RTF-1 and RTF-2 plasmids in various E. coli
strains carrying RI and RII restriction and modification host specificities
restricted unmodified lambda with EOPS of 10-4 and 10-2 respectively.  The RTF
plasmids were mutagenized with NTG and only one mutant phenotype, r-m+, was
obtained.  This indicated that there were two genes, a restriction and a
modification gene, in the host specificity system of the RTFs and differed from
the three gene system of E. coli B and K.  Using mutant and wild-type RI and
RII host specificities in various combinations revealed that no complementation
occurred between the RI and RII host specificities and that they were mutually
exclusive.  The RI and RII restriction endonucleases were purified to enable
characterization of the enzymes.  RI and RII restriction endonucleases required
only Mg++ and unmodified DNA for their enzymatic activity in contrast to the
cofactor requirements (SAM, ATP, Mg++) of the E. coli B and K restriction
endonucleases.  The molecular weights of the RI and RII restriction
endonucleases were estimated to be 80,000 and 100,000 daltons, respectively,
and recent evidence indicates two 50,000 MW subunits for the RII restriction
endonuclease.  Optimal activity of RII restriction endonuclease was at 37o in
0.1M Tris or glycylglycine buffer at pH 7.5 in 5 mM Mg++.  Optimal conditions
for RI restriction endonuclease was between 26 to 37C in 0.1M Tris buffer at pH
7.5 or at pH 7.2 with either 0.1M phosphate or glycylglycine buffer using 10 mM
Mg++.  The average size of lambda DNA fragments produced by the RII
endonucleolytic scissions was estimated at 2 Md MW.  The RI restriction
endonuclease produced one fragment about 13 Md MW and four to six fragments of
3.8 Md MW.  The RI and RII host specificities are classified as a second type
of restriction and modification mechanism on the basis of the genetic and
biochemical data.  The mechanism is simple and two cistrons are involved in
restriction and modification.  Each enzyme is composed of two identical
cistronic subunits.  On the other hand the more complex mechanism, which is
represented by the K and B host specificities, is composed of large enzymes
with complex subunit structures.  Three cistrons are involved in this
restriction and modification mechanism and the cofactor requirements are SAM,
ATP and Mg++.  The RI modification methylase was found on polyacrylamide gel as
one of two protein bands, the other being the RI restriction endonuclease.
Preliminary data indicates that the modification methylase requires only SAM
and unmodified DNA for its activity.

<>

<1>Yoshimoto, H., Takahashi, Y., Hamada, N., Umemoto, T.
<2>Genetic transformation of Porphyromonas gingivalis by electroporation.
<3>Oral Microbiol. Immunol.
<4>8
<5>208-212
<6>1993
<7>Porphyromonas gingivalis was transformed by electroporation using the DNA of plasmid pE5-2, or
its derivative, pYT7. Prior to transformation, pE5-2 was transferred from Escherichia coli to
P. gingivalis strains by conjugation (mobilization with R751) and the plasmid DNA was purified
from the P. gingivalis transconjugants. Transformation occurred when the recipient strain and
the donor strain from which the plasmid DNA was purified were homologous. If they were
heterologous, transformation did not take place or did so at a very low frequency. This
suggested that a restriction-modification system is present in P. gingivalis strains. Plasmid
pYT7 was derived by removing an 8.0 kb AvaI fragment from pE5-2 that was purified from P.
gingivalis cells. It has several single-cutting restriction sites such as EcoRI, AvaI, and
ClaI usable for gene cloning, though it was not stable enough in P. gingivalis cells, probably
because the rep gene was derived from a relatively distant species, Bacteroides eggerthii.

<>

<1>Yoshino, T., Honda, T., Tanaka, M., Tanaka, T.
<2>Draft Genome Sequence of Marine Cyanobacterium Synechococcus sp. Strain NKBG15041c.
<3>Genome Announcements
<4>1
<5>e00954-13
<6>2013
<7>Synechococcus sp. strain NKBG15041c was isolated as a fast-growing marine cyanobacterium.
Genetic transformation techniques using this strain have been
well established for metabolic engineering. Here we report the draft genome
sequence for this strain, consisting of 44 contigs containing a total of
3,180,043 bp and 3,224 putative protein-coding genes.

<>

<1>Yoshioka, H., Nakamura, H., Sasaki, J., Tahara, Y., Yamada, Y.
<2>Purification, properties and recognition sequence of site-specific restriction endonuclease from "Acetobacter xylinus".
<3>Agric. Biol. Chem.
<4>47
<5>2871-2879
<6>1983
<7>A type II restriction endonuclease was purified from "Acetobacter xylinus" IFO
3288 by consecutive column chromatographies on heparin-Sepharose CL-6B,
DEAE-Sepharose CL-6B, DNA-cellulose and Sephacryl S-400 superfine.  The
purified enzyme was homogeneous on gel disc electrophoresis and free of other
endonuclease, exonuclease and phospatase activities.  The enzyme was optimally
active at 37C at pH 7.5 and required 50~-150 mM NaCl for the enzyme reaction.
The enzyme cleaved lambda and M13 mp7 RF DNAs at two and one site,
respectively, but did not cleave pBR322, SV40 and PhiX174 RF DNAs.  the
recognition sequence for the enzyme was determined to be 5'-C-C-T-N-A-G-G-3',
and the enzyme was found to cut between C and T in the sequence, being an
isoschizomer of endonuclease from a blue-green alga, Microcoleus species UTEX
LB2220 (MstII).

<>

<1>You, D.L., Indrakanti, R., Cao, B., Yao, F., Deng, Z.X., Dedon, P.C.
<2>Development of a mass spectrometry-based assay to define the biochemistry of phosphorothioate DNA modifications of DNA in bacterial restriction/modification systems.
<3>ACS Abstracts
<4>242
<5>0
<6>2011
<7>We recently discovered that a wide variety of microorganisms possess sequences-specific
phosphorothioate modifications (S-modifications) of their genomes, which are incorporated into
DNA by members of the Dnd protein family.  The S-modifications occur at two discrete levels
consistent with the 4- and 6-nucleotide consensus sequences of a restriction/modification
system.  Formal proof for this model arose in enteropathogenic Salmonella enterica serovar
Cerro 87, in which we discovered a host-specific restriciton system DNA S-modification, which
is more complicated than the well-known methylation-specific restriction/modification systems.
In an effort to develop an in vitro reconstituted phosphorothioation system, we have developed
a MALDI-TOF assay in which S-modificaiton of an oligonucleotide possessiong a consensus
sequence is monitored by a 16 amu mass increase upon insertion of sulfur into the DNA
backbone.  The assay is being used to define the biochemical mechanism involved in
phosphorothioation of DNA in this novel restriction/modification system in bacteria.

<>

<1>You, X.Y., Guo, X., Zheng, H.J., Zhang, M.J., Liu, L.J., Zhu, Y.Q., Zhu, B., Wang, S.Y., Zhao, G.P., Poetsch, A., Jiang, C.Y., Liu, S.J.
<2>Unraveling the Acidithiobacillus caldus complete genome and its central metabolisms for carbon assimilation.
<3>J. Genetics Genomics
<4>38
<5>243-252
<6>2011
<7>Acidithiobacillus caldus is one of the dominant sulfur-oxidizing bacteria
in bioleaching reactors. It plays the essential role in maintaining the
high acidity and oxidation of reduced inorganic sulfur compounds during
bioleaching process. In this report, the complete genome sequence of A.
caldus SM-1 is presented. The genome is composed of one chromosome
(2,932,225 bp) and four plasmids (pLAtc1, pLAtc2, pLAtc3, pLAtcm) and it
is rich in repetitive sequences (accounting for 11% of the total genome),
which are often associated with transposable genetic elements. In
particular, twelve copies of ISAtfe and thirty-seven copies of ISAtc1 have
been identified, suggesting that they are active transposons in the
genome. A. caldus SM-1 encodes all enzymes for the central metabolism and
the assimilation of carbon compounds, among which 29 proteins/enzymes were
identifiable with proteomic tools. The SM-1 fixes CO(2)via the classical
Calvin-Bassham-Benson (CBB) cycle, and can operate complete
Embden-Meyerhof pathway (EMP), pentose phosphate pathway (PPP), and
gluconeogenesis. It has an incomplete tricarboxylic acid cycle (TCA). Four
putative transporters involved in carbohydrate uptake were identified.
Taken together, the results suggested that SM-1 was able to assimilate
carbohydrates and this was subsequently confirmed experimentally because
addition of 1% glucose or sucrose in basic salt medium significantly
increased the growth of SM-1. It was concluded that the complete genome of
SM-1 provided fundamental data for further investigation of its physiology
and genetics, in addition to the carbon metabolism revealed in this study.

<>

<1>You, X.Y., Liu, C., Wang, S.Y., Jiang, C.Y., Shah, S.A., Prangishvili, D., Liu, S.J., Garrett, R.A.
<2>Genomic analyses of Acidianus hospitalis W1 a host for studying crenarchaeal virus and plasmid life cycles.
<3>Extremophiles
<4>15
<5>487-497
<6>2011
<7>The Acidianus hospitalis W1 genome consists of a minimally sized chromosome of about 2.13 Mb
and a conjugative plasmid pAH1 and it is a host for the model filamentous lipothrixvirus AFV1.
The chromosome carries three putative replication origins in conserved genomic regions and two
large regions where non-essential genes are clustered. Within these variable regions, a few
orphan orfB and other elements of the IS200/607/605 family are concentrated with a novel class
of MITE-like repeat elements.  There are also 26 highly diverse vapBC antitoxin-toxin gene
pairs proposed to facilitate maintenance of local chromosomal regions and to minimise the
impact of environmental stress. Complex and partially defective CRISPR/Cas/Cmr immune systems
are present and interspersed with five vapBC gene pairs. Remnants of integrated viral genomes
and plasmids are located at five intron-less tRNA genes and several non-coding RNA genes are
predicted that are conserved in other Sulfolobus genomes. The putative metabolic pathways for
sulphur metabolism show some significant differences from those proposed for other Acidianus
and Sulfolobus species. The small and relatively stable genome of A. hospitalis W1 renders it
a promising candidate for developing the first Acidianus genetic systems.

<>

<1>You, Y., He, L., Zhang, M., Fu, J., Gu, Y., Zhang, B., Tao, X., Zhang, J.
<2>Comparative Genomics of Helicobacter pylori Strains of China Associated with Different Clinical Outcome.
<3>PLoS ONE
<4>7
<5>e38528
<6>2012
<7>In this study, a whole-genome CombiMatrix Custom oligonucleotide tiling microarray with 90000
probes covering six sequenced Helicobacter pylori
(H. pylori) genomes was designed. This microarray was used to compare
the genomic profiles of eight unsequenced strains isolated from
patients with different gastroduodenal diseases in Heilongjiang
province of China. Since significant genomic variation was found among
these strains, an additional 76 H. pylori strains associated with
different clinical outcomes were isolated from various provinces of
China. These strains were tested by polymerase chain reaction to
demonstrate this distinction. We identified several highly variable
regions in strains associated with gastritis, gastric ulceration, and
gastric cancer. These regions are associated with genes involved in the
bacterial type I, type II, and type III R-M systems. They were also
associated with the virB gene, which lies on the well-studied cag
pathogenic island. While previous studies have reported on the diverse
genetic characterization of this pathogenic island, in this study, we
find that it is conserved in all strains tested by microarray.
Moreover, a number of genes involved in the type IV secretion system,
which is related to horizontal DNA transfer between H. pylori strains,
were identified in the comparative analysis of the strain-specific
genes. These findings may provide insight into new biomarkers for the
prediction of gastric diseases.

<>

<1>You, Y., Liu, L., Zhang, M., Han, X., He, L., Zhu, Y., Ni, P., Zhang, J.
<2>Genome Sequences of Three Helicobacter pylori Strains Isolated from Atrophic Gastritis and Gastric Ulcer Patients in China.
<3>J. Bacteriol.
<4>194
<5>6314-6315
<6>2012
<7>Helicobacter pylori is a bacterial pathogen which can lead to several human gastric diseases.
Here we describe the genome sequences of three strains isolated
from atrophic gastritis and gastric ulcers patients in China. The data will
permit genomic characterization of traits that may contribute to various gastric
diseases.

<>

<1>You, Y., Liu, L., Zhang, M., Zhu, Y., He, L., Li, D., Zhang, J.
<2>Genomic characterization of a Helicobacter pylori isolate from a patient with gastric cancer in China.
<3>Gut Pathog.
<4>6
<5>5
<6>2014
<7>Background: Helicobacter pylori is well known for its relationship with the occurrence of
several severe gastric diseases. The mechanisms of pathogenesis triggered by H. pylori are
less well known. In this study, we report the genome sequence and genomic characterizations of
H. pylori strain HLJ039 that was isolated from a patient with gastric cancer in the Chinese
province of Heilongjiang, where there is a high incidence of gastric cancer. To investigate
potential genomic features that may be involved in pathogenesis of carcinoma, the genome was
compared to three previously sequenced genomes in this area.Result: We obtained 42 contigs
with a total length of 1,611,192 bp and predicted 1,687 coding sequences. Compared to strains
isolated from gastritis and ulcers in this area, 10 different regions were identified as being
unique for HLJ039; they mainly encoded type II restriction-modification enzyme, type II m6A
methylase, DNA-cytosine methyltransferase, DNA methylase, and hypothetical proteins. A unique
547-bp fragment sharing 93% identity with a hypothetical protein of Helicobacter cinaedi ATCC
BAA-847 was not present in any other previous H. pylori strains. Phylogenetic analysis based
on core genome single nucleotide polymorphisms shows that HLJ039 is defined as hspEAsia
subgroup, which belongs to the hpEastAsia group.Conclusion: DNA methylations, variations of
the genomic regions involved in restriction and modification systems, are the 'hot' regions
that may be related to the mechanism of H. pylori-induced gastric cancer. The genome sequence
will provide useful information for the deep mining of potential mechanisms related to East
Asian gastric cancer.

<>

<1>You, Y., Yang, X., Song, Y., Yan, X., Yuan, Y., Li, D., Yan, Y., Wang, H., Tao, X., Li, L., Jiang, X., Zhou, H., Xiao, D., Jin, L., Feng, Z., Yang, R., Luo, F., Cui, Y., Zhang, J.
<2>Draft Genome Sequences of Two Streptococcus pyogenes Strains Involved in Abnormal Sharp Raised Scarlet Fever in China, 2011.
<3>J. Bacteriol.
<4>194
<5>5983-5984
<6>2012
<7>A scarlet fever outbreak caused by Streptococcus pyogenes occurred in China in 2011. To
determine the genomic features of the outbreak strains, we deciphered
genomes of two strains isolated from the regions with the highest incidence
rates. The sequences will provide valuable information for comprehensive study of
mechanisms related to this outbreak.

<>

<1>Youell, J., Firman, K.
<2>EcoR124I: from plasmid-encoded restriction-modification system to nanodevice.
<3>Microbiol. Mol. Biol. Rev.
<4>72
<5>365-377
<6>2008
<7>Plasmid R124 was first described in 1972 as being a new member of in compatibility group
IncFIV, yet early physical investigations of
plasmid DNA showed that this type of classification was more complex
than first imagined. Throughout the history of the study of this
plasmid, there have been many unexpected observations. Therefore, in
this review, we describe the history of our understanding of this
plasmid and the type I restriction-modification (R-M) system that it
encodes, which will allow an opportunity to correct errors, or
misunderstandings, that have arisen in the literature. We also describe
the characterization of the R-M enzyme EcoR124I and describe the
unusual properties of both type I R-M enzymes and EcoR124I in
particular. As we approached the 21st century, we began to see the
potential of the EcoR124I R-M enzyme as a useful molecular motor, and
this leads to a description of recent work that has shown that the R-M
enzyme can be used as a nanoactuator. Therefore, this is a history that
takes us from a plasmid isolated from (presumably) an infected source
to the potential use of the plasmid-encoded R-M enzyme in
bionanotechnology.

<>

<1>Youell, J., Firman, K.
<2>Mechanistic insight into Type I restriction endonucleases.
<3>Front. Biosci., Landmark Ed.
<4>17
<5>2122-2139
<6>2012
<7>Restriction and modification are two opposing activities that are used to protect bacteria
from cellular invasion by DNA (e.g. bacteriophage infection).  Restriction activity involves
cleavage of the DNA; while modification activity is the mechanism used to "mark" host DNA and
involves DNA methylation.  The study of Type I restriction enzymes has often been seen as an
esoteric exercise and this reflects some of their more unusual properties - non-stoichiometric
(non-catalytic) cleavage of the DNA substrate, random cleavage of DNA, a massive ATPase
activity, and the ability to both cleave DNA and methylate DNA.  Yet these enzymes have been
found in many bacteria and are very efficient as a means of protecting bacteria against
bacteriophage infection, indicating they are successful enzynmes.  In this review, we
summarise recent work on the mechanisms of action, describe switching of function and review
their mechanism of action.  We also discuss structural rearrangements and cellular
localisation, which provide powerful mechanisms for controlling the enzyme activity.  Finally,
we speculate as to their involvement in recombination and discuss their relationship to
helicase enzymes.

<>

<1>Youn, J.H., Moodley, A., Park, Y.H., Sugimoto, C.
<2>Genome Sequence of Methicillin-Resistant Staphylococcus pseudintermedius Sequence Type 233 (ST233) Strain K7, of Human Origin.
<3>Genome Announcements
<4>1
<5>e00310-13
<6>2013
<7>We report the genome sequence of the methicillin-resistant Staphylococcus pseudintermedius
strain K7, isolated from the nares of a veterinarian in Seoul,
South Korea.

<>

<1>Young, D.D., Govan, J.M., Lively, M.O., Deiters, A.
<2>Photochemical Regulation of Restriction Endonuclease Activity.
<3>Chembiochem
<4>10
<5>1612-1616
<6>2009
<7>To elucidate biological processes, precise control over these
processes is required. Light represents an ideal external control
element because it can be easily regulated in a spatial and
temporal fashion, and conveys spatiotemporal control of biological
activity to the system under study.[1] The photochemical
regulation of oligonucleotide function through the installation
of light-removable protecting groups (caging groups) on
either the phosphate or the nucleotide base has recently received
considerable attention.[2-4] Important applications of this
technology involve the transient disruption of DNA hybridization
to photochemically control DNAzyme activity, the polymerase
chain reaction, antisense activity, as well as inhibition
of transcription.[4, 5] In this context, we demonstrated that a
single caging group installed on one base of a typical oligonucleotide
20-mer still enables DNA-DNA and DNA-RNA hybridization,
but could disrupt processing of the oligomer by polymerases
and inactivate the catalytic ability of DNAzymes.[2] As
a result, we became interested in exploring other biologically
relevant processes with photocaged DNA that do not involve
perturbation of hybridization. Due to the prevalence of DNA-
protein interactions both in vivo and in vitro,[6] we hypothesized
that it might be feasible to photochemically control such
an interaction for restriction endonucleases by the incorporation
of our 6-nitropiperonyloxymethyl (NPOM)-caged thymidine
nucleotide (Scheme 1) into DNA. Very few studies have
been conducted on the effects of unnatural nucleotides on the
fidelity and functionality of restriction enzymes. Those that
have, primarily involve the effects of endogenous base mutations
(for example, methylation events) that do not drastically
affect hydrogen bonding and base pair recognition. In many of
these cases, the catalytic capabilities of the restriction endo-
ACHTUNGTRENUNGnucleases are dramatically decreased, if not abrogated.

<>

<1>Young, J.P. et al.
<2>The genome of Rhizobium leguminosarum has recognizable core and accessory components.
<3>Genome Biol.
<4>7
<5>R34
<6>2006
<7>ABSTRACT : BACKGROUND : Rhizobium leguminosarum is an alpha-proteobacterial N2-fixing symbiont
of legumes that has been the
subject of more than a thousand publications. Genes for the symbiotic
interaction with plants are well studied, but the adaptations that allow
survival and growth in the soil environment are poorly understood. We have
sequenced the genome of R. leguminosarum biovar viciae strain 3841.
RESULTS : The 7.75 Mb genome comprises a circular chromosome and six
circular plasmids, with 61% G+C overall. All three rRNA operons and 52
tRNA genes are on the chromosome; essential protein-encoding genes are
largely chromosomal, but most functional classes occur on plasmids as
well. Of the 7,263 protein-encoding genes, 2,056 had orthologs in each of
three related genomes (Agrobacterium tumefaciens, Sinorhizobium meliloti,
and Mesorhizobium loti), and these genes were over-represented in the
chromosome and had above average G+C. Most supported the rRNA-based
phylogeny, confirming A. tumefaciens to be the closest among these
relatives, but 347 genes were incompatible with this phylogeny; these were
scattered throughout the genome but were over-represented on the plasmids.
An unexpectedly large number of genes were shared by all three rhizobia
but were missing from A. tumefaciens. CONCLUSION : Overall, the genome can
be considered to have two main components: a 'core', which is higher in
G+C, is mostly chromosomal, is shared with related organisms, and has a
consistent phylogeny; and an 'accessory' component, which is sporadic in
distribution, lower in G+C, and located on the plasmids and chromosomal
islands. The accessory genome has a different nucleotide composition from
the core despite a long history of coexistence.

<>

<1>Young, M. et al.
<2>Genome Sequence of the Fleming Strain of Micrococcus luteus, a Simple Free-Living Actinobacterium.
<3>J. Bacteriol.
<4>192
<5>841-860
<6>2010
<7>Micrococcus luteus (NCTC2665, 'Fleming strain') has one of the smallest genomes of
free-living actinobacteria sequenced to date, comprising a
single circular chromosome of 2,501,097 bp (G+C content, 73%) predicted to
encode 2,403 proteins. The genome shows extensive synteny with that of the
closely related organism, Kocuria rhizophila, from which it was
taxonomically separated relatively recently. Despite its small size, the
genome harbors 73 insertion sequence (IS) elements, almost all of which
are closely related to elements found in other actinobacteria. An IS
element is inserted into the rrs gene of one of only two rrn operons found
in M. luteus. The genome encodes only four sigma factors and 14 response
regulators, a finding indicative of adaptation to a rather strict
ecological niche (mammalian skin). The high sensitivity of M. luteus to
beta-lactam antibiotics may result from the presence of a reduced set of
penicillin-binding proteins and the absence of a wblC gene, which plays an
important role in the antibiotic resistance in other actinobacteria.
Consistent with the restricted range of compounds it can use as a sole
source of carbon for energy and growth, M. luteus has a minimal complement
of genes concerned with carbohydrate transport and metabolism and its
inability to utilize glucose as a sole carbon source may be due to the
apparent absence of a gene encoding glucokinase. Uniquely among
characterized bacteria, M. luteus appears to be able to metabolize
glycogen only via trehalose and to make trehalose only via glycogen. It
has very few genes associated with secondary metabolism. In contrast to
most other actinobacteria, M. luteus encodes only one
resuscitation-promoting factor (Rpf) required for emergence from dormancy,
and its complement of other dormancy-related proteins is also much
reduced. M. luteus is capable of long-chain alkene biosynthesis, which is
of interest for advanced biofuel production; a three-gene cluster
essential for this metabolism has been identified in the genome.

<>

<1>Young, T.-S., Kim, S.-H., Modrich, P., Beth, A., Jay, E.
<2>Preliminary x-ray diffraction studies of EcoRI restriction endonuclease-DNA complex.
<3>J. Mol. Biol.
<4>145
<5>607-610
<6>1981
<7>A complex between EcoRI restriction endonuclease and cognate DNA fragment,
5'-G-A-A-T-T-C    C-T-T-A-A-T-5' has been crystallized.  The space group is
P4212 with a=b=183.2 angstroms, c=49.7 angstroms, a=b=k=90o.  The unit cell
contains four enzyme monomers plus two duplex DNA fragments in an asymmetric
unit.  High quality crystals of the enzyme alone have also been obtained.

<>

<1>Youngblood, B., Bonnist, E., Dryden, D.T., Jones, A.C., Reich, N.O.
<2>Differential stabilization of reaction intermediates: specificity checkpoints for M.EcoRI revealed by transient fluorescence and  fluorescence lifetime studies.
<3>Nucleic Acids Res.
<4>36
<5>2917-2925
<6>2008
<7>M.EcoRI, a bacterial sequence-specific S-adenosyl-L-methionine-dependent DNA
methyltransferase, relies on a complex conformational mechanism to
achieve its remarkable specificity, including DNA bending, base flipping
and intercalation into the DNA. Using transient fluorescence and
fluorescence lifetime studies with cognate and noncognate DNA, we have
characterized several reaction intermediates involving the WT enzyme.
Similar studies with a bending-impaired, enhanced-specificity M.EcoRI
mutant show minimal differences with the cognate DNA, but significant
differences with noncognate DNA. These results provide a plausible
explanation of the way in which destabilization of reaction intermediates
can lead to changes in substrate specificity.

<>

<1>Youngblood, B., Buller, F., Reich, N.O.
<2>Determinants of sequence-specific DNA methylation: target recognition and catalysis are coupled in M.HhaI.
<3>Biochemistry
<4>45
<5>15563-15572
<6>2006
<7>Sequence specificity studies of the wild-type bacterial DNA cytosine C5 methyltransferase HhaI
were carried out with cognate (5'GCGC3') and
noncognate DNA substrates containing single base pair changes at the first
and the fourth position (underlined). Specificity for noncognate site
methylation at the level of kcat/KDDNA is decreased 9000-80000-fold
relative to the cognate site, manifested through changes in methylation,
or a prior step, and changes in KDDNA. Analysis of a new high-resolution
enzyme-DNA cocrystal structure provides a partial mechanistic
understanding of this discrimination. To probe the significance of
conformational transitions occurring prior to catalysis in determining
specificity, we analyzed the double mutant (H127A/T132A). These amino acid
substitutions disrupt the interface between the flexible loop (residues
80-99), which interacts with the DNA minor groove, and the active site.
The mutant's methylation of the cognate site is essentially unchanged, yet
its methylation of noncognate sites is decreased up to 460-fold relative
to the wild-type enzyme. We suggest that a significant contribution to
M.HhaI's specificity involves the stabilization of reaction intermediates
prior to methyl transfer, mediated by DNA minor groove-protein flexible
loop interactions.

<>

<1>Youngblood, B., Reich, N.
<2>Conformational transitions as molecular determinants of specificity for the DNA methyltransferase EcoRI.
<3>FASEB J.
<4>20
<5>A40
<6>2006
<7>DNA modifying enzymes have evolved a delicate balance between sequence specificity and
efficient catalysis. Utilization of indirect readout of
a DNA sequence, such as DNA bending by an enzyme, provides further
discrimination beyond the initial binding event between the enzyme and
DNA. Changes in the DNA bending and base flipping transitions in a
previously characterized specificity-enhanced mutant M.EcoRI DNA
adenine methyltransferase suggest a close relationship between
precatalytic conformational transitions and specificity. We present
fluorescence resonance energy transfer (FRET) and 2-AminoPurine (2AP)
fluorescence studies with the cognate sequence (GAATTC) for WT M.EcoRI,
and reveal a temporal relationship between DNA bending, intercalation
of the DNA substrate by the enzyme, and base-flipping. The direct
measurement of kinetic rate constants for DNA bending, intercalation,
and base-flipping with cognate and non-cognate substrates (GAATTT,
GGATTC) by WT M.EcoRI, provide a molecular understanding of how these
conformational transitions impact on specificity. The 3500 and
23,000-fold decreases in sequence specificity of WT M.EcoRI for
noncognates GAATTT and GGATTC are accounted for largely by a similar to
2500 fold increase in the reverse rate constants for intercalation and
base flipping. The predominant contribution to specificity is a
partitioning of enzyme intermediates away from the Michaelis complex
prior to catalysis. Our results provide a basis for understanding
sequence-specific DNA modification.

<>

<1>Youngblood, B., Reich, N.O.
<2>Conformational transitions as determinants of specificity for the DNA methyltransferase EcoRI.
<3>J. Biol. Chem.
<4>281
<5>26821-26831
<6>2006
<7>Changes in DNA bending and base flipping in a previously characterized specificity-enhanced M.
EcoRI DNA adenine methyltransferase mutant
suggest a close relationship between precatalytic conformational
transitions and specificity (Allan, B. W., Garcia, R., Maegley, K.,
Mort, J., Wong, D., Lindstrom, W., Beechem, J. M., and Reich, N. O.
(1999) J. Biol. Chem. 274, 19269-19275). The direct measurement of the
kinetic rate constants for DNA bending, intercalation, and base
flipping with cognate and noncognate substrates (GAATTT, GGATTC) of
wild type M. EcoRI using fluorescence resonance energy transfer and
2-aminopurine fluorescence studies reveals that DNA bending precedes
both intercalation and base flipping, and base flipping precedes
intercalation. Destabilization of these intermediates provides a
molecular basis for understanding how conformational transitions
contribute to specificity. The 3500- and 23,000-fold decreases in
sequence specificity for noncognate sites GAATTT and GGATTC are
accounted for largely by an similar to 2500-fold increase in the
reverse rate constants for intercalation and base flipping,
respectively. Thus, a predominant contribution to specificity is a
partitioning of enzyme intermediates away from the Michaelis complex
prior to catalysis. Our results provide a basis for understanding
enzyme specificity and, in particular, sequence-specific DNA
modification. Because many DNA methyltransferases and DNA repair
enzymes induce similar DNA distortions, these results are likely to be
broadly relevant.

<>

<1>Youngblood, B., Shieh, F.K., Buller, F., Bullock, T., Reich, N.O.
<2>S-adenosyl-L-methionine-dependent methyl transfer: observable precatalytic intermediates during DNA cytosine methylation.
<3>Biochemistry
<4>46
<5>8766-8775
<6>2007
<7>S-adenosyl-L-methionine- (AdoMet-) dependent methyltransferases are widespread, play critical
roles in diverse biological pathways, and are
antibiotic and cancer drug targets. Presently missing from our
understanding of any AdoMet-dependent methyl-transfer reaction is a
high-resolution structure of a precatalytic enzyme/AdoMet/DNA complex. The
catalytic mechanism of DNA cytosine methylation was studied by
structurally and functionally characterizing several active site mutants
of the bacterial enzyme M.HhaI. The 2.64 A resolution protein/DNA/AdoMet
structure of the inactive C81A M.HhaI mutant suggests that active site
water, an approximately 13 degree tilt of the target base toward the
active site nucleophile, and the presence or absence of the cofactor
methylsulfonium are coupled via a hydrogen-bonding network involving
Tyr167. The active site in the mutant complex is assembled to optimally
align the pyrimidine for nucleophilic attack and subsequent methyl
transfer, consistent with previous molecular dynamics ab initio and
quantum mechanics/molecular mechanics calculations. The mutant/DNA/AdoHcy
structure (2.88 A resolution) provides a direct comparison to the
postcatalytic complex. A third C81A ternary structure (2.22 A resolution)
reveals hydrolysis of AdoMet to adenosine in the active site, further
validating the coupling between the methionine portion of AdoMet and
ultimately validating the structural observation of a
prechemistry/postchemistry water network. Disruption of this
hydrogen-bonding network by a Tyr167 to Phe167 mutation does not alter the
kinetics of nucleophilic attack or methyl transfer. However, the Y167F
mutant shows detectable changes in kcat, caused by the perturbed kinetics
of AdoHcy release. These results provide a basis for including an
extensive hydrogen-bonding network in controlling the rate-limiting
product release steps during cytosine methylation.

<>

<1>Youngblood, B., Shieh, F.K., De Los Rios, S., Perona, J.J., Reich, N.O.
<2>Engineered Extrahelical Base Destabilization Enhances Sequence Discrimination of DNA Methyltransferase M.HhaI.
<3>J. Mol. Biol.
<4>362
<5>334-346
<6>2006
<7>Improved sequence specificity of the DNA cytosine methyltransferase HhaI was achieved by
disrupting interactions at a hydrophobic interface between
the active site of the enzyme and a highly conserved flexible loop.
Transient fluorescence experiments show that mutations disrupting this
interface destabilize the positioning of the extrahelical, "flipped"
cytosine base within the active site. The ternary crystal structure of the
F124A M.HhaI bound to cognate DNA and the cofactor analogue
S-adenosyl-l-homocysteine shows an increase in cavity volume between the
flexible loop and the core of the enzyme. This cavity disrupts the
interface between the loop and the active site, thereby destabilizing the
extrahelical target base. The favored partitioning of the base-flipped
enzyme-DNA complex back to the base-stacked intermediate results in the
mutant enzyme discriminating better than the wild-type enzyme against
non-cognate sites. Building upon the concepts of kinetic proofreading and
our understanding of M.HhaI, we describe how a 16-fold specificity
enhancement achieved with a double mutation at the loop/active site
interface is acquired through destabilization of intermediates prior to
methyltransfer rather than disruption of direct interactions between the
enzyme and the substrate for M.HhaI.

<>

<1>Youngblood, B.A.
<2>Analysis of conformational transitions that facilitate DNA methyltransferase specificity and catalysis.
<3>Ph.D. Thesis, Univ. of California, Santa Barbara
<4>
<5>1-190
<6>2007
<7>DNA modifying enzymes have evolved a delicate balance between sequence specificity and
efficient catalysis. Utilization of indirect readout of a DNA sequence, such as DNA bending
and/or base flipping by an enzyme, can provide further discrimination between cognate and
noncognate substrates by the enzyme. Using fluorescence resonance energy transfer (FRET) and
2-Aminopurine (2AP) fluorescence studies with the cognate sequence (GAATTC) for WT M.EcoRI,
the temporal relationship between DNA bending, intercalation of the DNA substrate by the
enzyme, and base-flipping is elucidated. Destabilization of these intermediates provides a
molecular basis for understanding how conformational transitions contribute to specificity.
The 3500 and 23,000-fold decreases in sequence specificity for noncognate sites GAATTT and
GGATTC are accounted for largely by a ~2500 fold increase in the reverse rate constants for
intercalation and base flipping, respectively. The predominant contribution to specificity is
a partitioning of enzyme intermediates away from the Michaelis complex prior to catalysis.
Building upon the concepts that were developed with the M.EcoRI studies, an improvement in
sequence specificity of the DNA cytosine methyltransferase HhaI is achieved by disrupting
interactions at a hydrophobic interface between the active site of the enzyme and a highly
conserved flexible loop. Transient fluorescence experiments with 2AP show that mutations
disrupting this interface interfere with catalysis by destabilizing the extrahelical "flipped"
cytosine base in the active site. This destabilization, caused by a double mutant, results in
a ~500-fold specificity enhancement compared to the WT enzyme. To better understand the
contribution of specific active site atoms to the catalytic enhancement of M.HhaI and the
rearrangement of these atoms to facilitate the chemistry step, three crystal structures were
solved using the true cofactor S-adenosyl-L-methionine (pre-chemistry), and the cofactor
product S-adenosyl-L-homocysteine (post-chemistry). Much work remains in resolving how
discrimination occurs for eukaryotic DNA methyltransferases in comparison to prokaryotic DNA
methyltransferases mentioned above. To facilitate our advancement in understanding eukaryotic
DNA methyltransferases, a model system to explore substrate discrimination in cells was
developed. Using HIV-1, the mechanism for over-expression and targeting of the mammalian DNA
methyltransferase 1 (DNMT1) to specific sites in the genome has been probed.

<>

<1>Youngblood, B.A., Shieh, F.-K., Snider, S., Perona, J., Reich, N.
<2>Engineering a more specific DNA methyltransferase by inducing loop instability.
<3>FASEB J.
<4>19
<5>A846
<6>2005
<7>The disruption of aromatic-aromatic interactions between Phe84 within the flexible loop
(residues 80-99) and Phe124, which stabilizes important active sites residues in the bacterial
DNA cytosine C5 methyltransferase, M.HhaI, result in mutant enzymes that are significantly
increased in specificity.  The Phe124Ala M.HhaI-DNA cocrystal structure shows the flexibility
of this loop to be increased.  Loop destabilization results in a decreased ability to maintain
the extrahelical cytosine poised for catalysis; thus, this intermediate partitions toward
release rather than catalysis.  The net result is that the mutant is increased in specificity,
since more rapid release of non-cognate DNA leads to lower rates of methylation at such sites.
Double mutant cycles involving Phe84Ala, Phe124Ala, His127Ala, and Thr132Ala, show that these
effects involve structural accommodations over 2 angstroms.  Our results have implications for
the rational design of enzyme specificity, and for the induced fit mechanism occurring in the
base-flipping DNA modifying enzymes involved in diverse epigenetic transformations.

<>

<1>Youssoufian, H., Hammer, S.M., Hirsch, M.S., Mulder, C.
<2>Methylation of the viral genome in an in vitro model of herpes simplex virus latency.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>79
<5>2207-2210
<6>1982
<7>An in vitro model of latency of herpes simplex virus type 1 (HSV-1) in a
lymphoid cell line has been developed recently.  CEM cells persistently
infected with HSV-1 transiently ceased to produce virus for 24 days.  This
nonproductive state could either be reversed with phytohemagglutinin or
maintained with concanavalin A.  This system was used to study the relationship
between DNA methylation and HSV-1 latency.  DNA was probed for methylation by
comparing the cleavage pataterns generated by two pairs of restriction
endonucleases (SmaI vs. XmaI and HpaII vs. MspI); these enzymes show
differential activity reflecting methylation of the recognition sequences.
Viral DNA in the concanavalin A-treated cells (not producing virus) was found
to be extensively methylated.  By contrast, no methylated copies were detected
in viral DNA from producer cells.  About 800 days after the initial infection,
the productive culture once again became nonproductive.  Viral sequences in the
latter cells were also methylated.  Reconstitution experiments revealed 1-2
copies of viral DNA in cells from the latent stages and 40-80 copies in cells
from productive stages.  Most (if not all) of the viral genome is present in
cells from various productive and latent stages.  No differences in sequence
arrangement were detected (although a terminal fragment of intracellular HSV-1
DNA appeared to be under represented in latent cells).  These results suggest a
role for DNA methylation in the mechanism of HSV-1 latency in this system.

<>

<1>Youssoufian, H., Mulder, C.
<2>Detection of methylated sequences in eukaryotic DNA with the restriction endonucleases SmaI and XmaI.
<3>J. Mol. Biol.
<4>150
<5>133-136
<6>1981
<7>The restriction endonuclease XmaI cleaves eukaryotic DNA whether or not its
recognition sequence (C^-C-C-G-G-G) contains 5-methylcytosine in the CpG
dinucleotide.  This is in contrast to its isoschizomer, endo R SmaI, which does
not cleave DNA if the CpG of its recognition sequence (C-C-C^G-G-G) is
methylated.  Thus, this isoschizomer pair will be useful in the study of
eukaryotic DNA methylation.

<>

<1>Yousten, A.A.
<2>Bacteriophage typing of mosquito pathogenic strains of Bacillus sphaericus.
<3>J. Invertebr. Pathol.
<4>43
<5>124-125
<6>1984
<7>Certain strains of Bacillus sphaericus have been shown to be pathogenic for
mosquito larvae (S. Singer, Biotechnol. Bioeng. 2, 1335, 1980).  All of these
strains have been isolated directly from dead larvae or from habitats of
larvae.  In contrast, none of the 56 strains of B. sphaericus obtained from
various culture collections exhibited pathogenicity for larvae of Culex
quinquefasciatus (V. Krych, J., Johnson, and A. Yousten, Int. J. Syst.
Bacteriol. 30, 476-484, 1980).  In the same study, seven pathogens were shown
to be members of a single DNA homology group which showed only a low degree of
genetic relatedness to the type strain of the species.

<>

<1>Yu, B., Su, F., Wang, L., Xu, K., Zhao, B., Xu, P.
<2>Draft Genome Sequence of Sporolactobacillus inulinus Strain CASD, an Efficient D-Lactic Acid-Producing Bacterium with High-Concentration  Lactate Tolerance Capability.
<3>J. Bacteriol.
<4>193
<5>5864-5865
<6>2011
<7>Sporolactobacillus inulinus CASD is an efficient d-lactic acid producer with high optical
purity. Here we report for the first time the draft
genome sequence of S. inulinus (2,930,096 bp). The large number of
annotated two-component system genes makes it possible to explore the
mechanism of extraordinary lactate tolerance of S. inulinus CASD.

<>

<1>Yu, B., Su, F., Wang, L., Zhao, B., Qin, J., Ma, C., Xu, P., Ma, Y.
<2>Genome Sequence of Lactobacillus rhamnosus Strain CASL, an Efficient L-Lactic Acid Producer from Cheap Substrate Cassava.
<3>J. Bacteriol.
<4>193
<5>7013-7014
<6>2011
<7>Lactobacillus rhamnosus is a type of probiotic bacteria with industrial potential for l-lactic
acid production. We announce the draft genome
sequence of L. rhamnosus CASL (2,855,156 bp with a G+C content of 46.6%),
which is an efficient producer of l-lactic acid from cheap, nonfood
substrate cassava with a high production titer.

<>

<1>Yu, C.S., Yim, K.Y., Tsui, S.K., Chan, T.F.
<2>Complete Genome Sequence of Bacillus subtilis Strain QB928, a Strain Widely Used  in B. subtilis Genetic Studies.
<3>J. Bacteriol.
<4>194
<5>6308-6309
<6>2012
<7>The complete genome sequence of Bacillus subtilis strain QB928 was constructed to facilitate
studies in the evolution of the genetic code. With a widespread use of
the strain in Bacillus subtilis genetics studies, its complete genome sequence
would facilitate deeper understanding of Bacillus subtilis genetics.

<>

<1>Yu, D., Pi, B., Chen, Y., Wang, Y., Ruan, Z., Otto, M., Yu, Y.
<2>Characterization of the Staphylococcal Cassette Chromosome Composite Island of Staphylococcus haemolyticus SH32, a Methicillin-Resistant Clinical Isolate from China.
<3>PLoS ONE
<4>9
<5>6
<6>2014
<7>Staphylococcal cassette chromosome (SCC) elements contribute considerably to virulence and
resistance to antibiotic agents in staphylococci. SCC elements in coagulase-negative
staphylococci (CoNS) are highly diverse and there is evidence suggesting that they serve as a
reservoir for antibiotic resistance genes in methicillin-resistant Staphylococcus aureus
(MRSA). However, only a small number of SCC elements have been characterized in CoNS and their
exact roles in the emergence and evolution of MRSA remain to be demonstrated. Here, we
determined the structure of an SCC composite island (CISH32) found in the clinical
Staphylococcus haemolyticus isolate SH32 by whole-genome DNA sequencing. CISH32 was 48 kb in
length and mainly composed of two imperfect SCC elements, namely (i) a Psi SCCmec(SH32) part
containing a class C1 mec gene complex but lacking ccr genes and (ii) a SCCSH32 part with a
ccrA5B3 gene complex but lacking mec genes. In addition, CISH32 contained a type III
restriction-modification syst em and several resistance loci, for example genes conferring
resistance to cadmium and arsenic. Psi SCCmec(SH32) is almost entirely identical to a pseudo
SCCmec element found in S. haemolyticus WCH1 and shares pronounced sequence similarity to a
Psi SCCmec element of S. haemolyticus JCSC1435. However, staphylococci other than S.
haemolyticus, including S. aureus and S. epidermidis, contain homologs of SCCSH32 that are
more similar to SCCSH32 than those elements found in S. haemolyticus, suggesting that CIS H32
of S. haemolyticus SH32 was assembled in recent evolutionary events. Moreover, the composite
structure of CISH32 indicates that the detection of class C1 mec and ccrA5B3 gene complexes in
S. haemolyticus does not always indicate the existence of a UT9-type SCCmec element, which has
remained questionable.

<>

<1>Yu, D.S., Jeong, H., Lee, D.H., Kwon, S.K., Song, J.Y., Kim, B.K., Park, M.S., Ji, G.E., Oh, T.K., Kim, J.F.
<2>Complete Genome Sequence of the Probiotic Bacterium Bifidobacterium bifidum Strain BGN4.
<3>J. Bacteriol.
<4>194
<5>4757-4758
<6>2012
<7>Bifidobacterium bifidum, a common endosymbiotic inhabitant of the human gut, is considered a
prominent probiotic microorganism that may promote health. We
completely decrypted the 2.2-Mb genome sequence of B. bifidum BGN4, a strain that
had been isolated from the fecal sample of a healthy breast-fed infant, and
annotated 1,835 coding sequences.

<>

<1>Yu, F., Watanabe, F.
<2>A method of transforming bacteria belonging to Rhodococcus.
<3>Japanese Patent Office
<4>JP 2007228934 A
<5>
<6>2007
<7>
<>

<1>Yu, H., Li, Y., Tang, H., Xu, P.
<2>Genome Sequence of a Newly Isolated Nicotine-Degrading Bacterium, Ochrobactrum sp. SJY1.
<3>Genome Announcements
<4>2
<5>e00720-14
<6>2014
<7>Ochrobactrum sp. SJY1 uses nicotine as the sources of carbon, nitrogen, and energy. The genome
of SJY1 was sequenced in order to provide insights into its
mechanism of nicotine degradation. Physiological characteristics and genome
analysis indicate that strain SJY1 might have a different nicotine degradation
pathway from the pyridine or pyrrolidine pathway.

<>

<1>Yu, H., Liu, L., Chang, Z., Wang, S., Wen, B., Yin, P., Liu, D., Chen, B., Zhang, J.
<2>Genome Sequence of the Bacterium Bifidobacterium longum Strain CMCC P0001, a Probiotic Strain Used for Treating Gastrointestinal Disease.
<3>Genome Announcements
<4>1
<5>e00716-13
<6>2013
<7>Bifidobacterium longum subsp. longum CMCC P0001, a standard probiotic strain in China, has
been widely used in clinical medicine for more than 20 years. Here we
report the genome features of B. longum strain CMCC P0001.

<>

<1>Yu, H., Tang, H., Wang, L., Yao, Y., Wu, G., Xu, P.
<2>Complete Genome Sequence of the Nicotine-Degrading Pseudomonas putida Strain S16.
<3>J. Bacteriol.
<4>193
<5>5541-5542
<6>2011
<7>Pseudomonas putida S16 is an efficient degrader of nicotine. The complete genome of strain S16
(5,984,790 bp in length) includes genes related to
catabolism of aromatic and heterocyclic compounds. The genes of enzymes in
the core genome and a genomic island encode the proteins responsible for
nicotine catabolism.

<>

<1>Yu, H., Yuan, M., Lu, W., Yang, J., Dai, S., Li, Q., Yang, Z., Dong, J., Sun, L., Deng, Z., Zhang, W., Chen, M., Ping, S., Han, Y., Zhan, Y., Yan, Y., Jin, Q., Lin, M.
<2>Complete Genome Sequence of the Nitrogen-fixing and Rhizosphere-associated Bacterium Pseudomonas stutzeri Strain DSM4166.
<3>J. Bacteriol.
<4>193
<5>3422-3423
<6>2011
<7>We present here the analysis of the whole genome sequence of Pseudomonas stutzeri strain
DSM4166, a diazotrophic isolate from the rhizosphere of a
Sorghum nutans cultivar. To our knowledge, this is the second one to be
sequenced in P. stutzeri. The availability and analysis of the genome
provides insight into the evolution of the nitrogen fixation property, and
identification of rhizosphere competence traits required in interactions
with host plants.

<>

<1>Yu, H.B., Zhang, Y.L., Lau, Y.L., Yao, F., Vilches, S., Merino, S., Tomas, J.M., Howard, S.P., Leung, K.Y.
<2>Identification and characterization of putative virulence genes and gene clusters in Aeromonas hydrophila PPD134/91.
<3>Appl. Environ. Microbiol.
<4>71
<5>4469-4477
<6>2005
<7>Aeromonas hydrophila is a gram-negative opportunistic pathogen of animals
and humans. The pathogenesis of A. hydrophila is multifactorial. Genomic
subtraction and markers of genomic islands (GIs) were used to identify
putative virulence genes in A. hydrophila PPD134/91. Two rounds of genomic
subtraction led to the identification of 22 unique DNA fragments encoding
19 putative virulence factors and seven new open reading frames, which are
commonly present in the eight virulence strains examined. In addition,
four GIs were found, including O-antigen, capsule, phage-associated, and
type III secretion system (TTSS) gene clusters. These putative virulence
genes and gene clusters were positioned on a physical map of A. hydrophila
PPD134/91 to determine their genetic organization in this bacterium.
Further in vivo study of insertion and deletion mutants showed that the
TTSS may be one of the important virulence factors in A. hydrophila
pathogenesis. Furthermore, deletions of multiple virulence factors such as
S-layer, serine protease, and metalloprotease also increased the 50%
lethal dose to the same level as the TTSS mutation (about 1 log) in a blue
gourami infection model. This observation sheds light on the
multifactorial and concerted nature of pathogenicity in A. hydrophila. The
large number of putative virulence genes identified in this study will
form the basis for further investigation of this emerging pathogen and
help to develop effective vaccines, diagnostics, and novel therapeutics.

<>

<1>Yu, M., Ren, C., Qiu, J., Luo, P., Zhu, R., Zhao, Z., Hu, C.
<2>Draft Genome Sequence of the Opportunistic Marine Pathogen Vibrio harveyi Strain  E385.
<3>Genome Announcements
<4>1
<5>e00677-13
<6>2013
<7>Vibrio harveyi strain E385 was isolated from a diseased cage-cultured grouper in  Daya Bay,
China. Phylogenetic analysis based on the 16S rRNA gene sequence showed
similarity with V. harveyi strain BAA-1116. We sequenced the pathogenic strain V.
harveyi E385 and compared the genome with that of the nonpathogenic strain V.
harveyi BAA-1116.

<>

<1>Yu, M., Tang, K., Shi, X., Zhang, X.H.
<2>Genome Sequence of Pseudoalteromonas flavipulchra JG1, a Marine Antagonistic Bacterium with Abundant Antimicrobial Metabolites.
<3>J. Bacteriol.
<4>194
<5>3735
<6>2012
<7>The marine bacterium Pseudoalteromonas flavipulchra JG1 can synthesize various antibacterial
metabolites, including protein and small molecules. The draft
genome of JG1 is about 5.36 Mb and harbors approximate 4,913 genes, which will
provide further insight into the synthesis of antimicrobial agents and
antagonistic mechanisms of P. flavipulchra against pathogens.

<>

<1>Yu, Y.J., Yang, M.T.
<2>A novel restriction-modification system from Xanthomonas campestris pv. vesicatoria encodes a m4C-methyltransferase and a nonfunctional  restriction endonuclease.
<3>FEMS Microbiol. Lett.
<4>272
<5>83-90
<6>2007
<7>A novel restriction-modification (R-M) system, designated as xveIIRM, from chromosomal DNA of
the Xanthomonas campestris pv. vesicatoria.
strain 7-1 (Xcv7-1) was cloned and characterized. The xveIIRM genes
involved in this R-M system are aligned in a tail-to-tail orientation
and overlapped by 12 base pairs. XveII methyltransferase gene could
encode a 299-amino acid protein (M.XveII) with an estimated mass of
33.7 kDa and was classified to be a member of P-class of m4C-MTase.
M.XveII methylates the second cytosine of the 5'-CCCGGG-3' recognition
sequence. The predicted amino acid sequence of the intact Xvell
endonuclease shared 41.9% identity with SmaI. However, a premature TAA
translation termination codon was found in the open reading frame of
xveIIR and expected to encode an 18.3 kDa truncated protein. The
sequence data are consistent with observation of this study that no
SmaI-like restriction activity could be detected in the cell extract of
Xcv7-1.

<>

<1>Yuan, J., Li, L., Sun, M., Dong, J., Hu, Q.
<2>Genome Sequence of Avirulent Riemerella anatipestifer Strain RA-SG.
<3>Genome Announcements
<4>1
<5>e0021812
<6>2013
<7>Riemerella anatipestifer is a pathogenic bacterium that has spread all over the world and is
associated with epizootic infections in waterfowl and other avian
species. R. anatipestifer RA-SG is an avirulent strain, isolated from an infected
duck in Guangdong province, China. The genome sequence of this species is
presented herein.

<>

<1>Yuan, K.X., Adam, Z., Tambong, J., Levesque, C.A., Chen, W., Lewis, C.T., De Boer, S.H., Li, X.S.
<2>Draft Genome Sequence of Pectobacterium wasabiae Strain CFIA1002.
<3>Genome Announcements
<4>2
<5>e00214-14
<6>2014
<7>Pectobacterium wasabiae, originally causing soft rot disease in horseradish in Japan, was
recently found to cause blackleg-like symptoms on potato in the United
States, Canada, and Europe. A draft genome sequence of a Canadian potato isolate
of P. wasabiae CFIA1002 will enhance the characterization of its pathogenicity
and host specificity features.

<>

<1>Yuan, R.
<2>The reaction mechanism of type I restriction endonucleases.
<3>Gene Amplif. Anal., Elsevier North Holland, Chirikjian, J.G., 
<4>1
<5>45-72
<6>1981
<7>*
   I. Introduction
  II. Genetics
 III. Enzyme structure
  IV. Nature of DNA recognition and cleavage sites
   V. Mechanism of restriction
  VI. Mechanism of modification
 VII. Mechanism of antirestriction systems
VIII. Projections and speculations


<>

<1>Yuan, R.
<2>Structure and mechanism of multifunctional restriction endonucleases.
<3>Annu. Rev. Biochem.
<4>50
<5>285-315
<6>1981
<7>Restriction endonucleases are strain-specific enzymes, which enable bacteria to
recognize and rapidly destroy foreign DNA and which cause double-stranded
scissions at a limited number of sites on the DNA.  In addition to having a
restriction activity, each of these bacterial strains possesses a specific DNA
methylase that transfers methyl groups from S-adenosylmethionine (AdoMet) to
specific adenine or sytosine residues in the DNA.  In this way, the DNA of the
cell is protected against its own restriction enzyme.

<>

<1>Yuan, R., Bickle, T.A., Ebbers, W., Brack, C.
<2>Multiple steps in DNA recognition by restriction endonuclease from E. coli K.
<3>Nature
<4>256
<5>556-560
<6>1975
<7>The process of DNA recognition by the activated form of the restriction
endonuclease from E. coli K involves three enzyme-DNA complexes which can be
differentiated experimentally.  These are:  an initial complex formed at a
nonspecific site; a recognition complex involving the host specificity site;
and a cleavage complex dependent on the presence of ATP.

<>

<1>Yuan, R., Burckhardt, J., Weisemann, J., Hamilton, D.L.
<2>The mechanism of DNA methylation by the restriction endonuclease from E. coli K.
<3>Biochemistry of S-adenosylmethionine and related compounds, MacMillan, Usdin, E., Borchardt, R.T., Creveling, C.R., London
<4>0
<5>239-247
<6>1981
<7>Restriction endonucleases are strain-specific enzymes which enable bacteria to
recognize and destroy foreign DNA by making double-stranded scissions at a
limited number of sites.  These bacterial strains also possess DNA methylases
which modify the DNA in order to protect it from the homologous restriction
enzymes.  These restriction endonucleases have been classified into three
different types (Yuan, 1981).  The Type I enzymes, such as the one coded by E.
coli K, are multifunctional proteins composed of three different subunits.
EcoK can catalyze three different reactions:  DNA cleavage (which requires ATP
and AdoMet), ATP hydrolysis (which is coupled to DNA cleavage) and DNA
methylation (which requires AdoMet).  We have made a detailed study of the
reaction mechanism of EcoK in order to understand certain unusual features,
such as its ability to catalyze two opposing reactions - restriction and
modification - and the cleavage of DNA at sites distal from the original
binding site.

<>

<1>Yuan, R., Hamilton, D.L.
<2>Type I and Type III restriction-modification enzymes.
<3>DNA Methylation. Biochemistry and Biological Significance., Springer-Verlag, Razin, A., Cedar, H., Riggs, A.D., New York
<4>0
<5>11-38
<6>1984
<7>None

<>

<1>Yuan, R., Hamilton, D.L.
<2>Restriction and modification of DNA by a complex protein.
<3>Am. Sci.
<4>70
<5>61-69
<6>1982
<7>The interaction of certain proteins with specific nucleotide sequences forms the basis for
such diverse biological functions as genetic transformation, recombination, control of gene
expression, and DNA repair.  In many cases, this interaction involves both the recognition of
a particular DNA site and the catalysis of a biochemical reaction.  Host-controlled
restriction and modification is one biological system in which genetic and molecular
approaches have been combined to give us a better understanding of the complexity of these
protein-DNA interactions.  Host-controlled restriction is the process by which certain strains
of bacteria recognize and degrade foreign DNA.  The same strains are able to carry out
modification of their own DNA to protect it from restriction.

<>

<1>Yuan, R., Hamilton, D.L., Burckhardt, J.
<2>DNA translocation by the restriction enzyme from E. coli K.
<3>Cell
<4>20
<5>237-244
<6>1980
<7>The restriction endonuclease Eco K binds to a host specificity site and then
proceeds to cleave the DNA at sites that may be several thousand bases away.
It does this by translocating the DNA past the enzyme in an ATP-dependent
reaction that results in the formation of highly twisted loop intermediates.
DNA cleavage can occur on either side of the host specificity site.

<>

<1>Yuan, R., Hamilton, D.L., Hadi, S.M., Bickle, T.A.
<2>Role of ATP in the cleavage mechanism of the EcoP15 restriction endonuclease.
<3>J. Mol. Biol.
<4>144
<5>501-519
<6>1980
<7>The EcoP15 restriction endonuclease forms complexes at specific sites on
unmodified DNA both in the presence and in the absence of
S-adenosyl-L-methionine.  ATP acts as an allosteric effector of EcoP15 and
induces DNA cleavage followed by release of the enzyme from the DNA.  The
efficiency of endonucleolytic scission varies from site to site.  The
nucleotide sequences at sites that are cleaved at a high frequency were
compared.

<>

<1>Yuan, R., Heywood, J., Meselson, M.
<2>ATP hydrolysis by restriction endonuclease from E. coli K.
<3>Nature New Biol.
<4>240
<5>42-43
<6>1972
<7>We wish to report a puzzling ATPase activity associated with the DNA
restriction endonuclease from E. coli strain K.  This enzyme makes a limited
number of double chain breaks in DNA molecules lacking the host-controlled
modification imparted by strain K.  Unmodified DNA molecules from bacteriophage
lambda, which serve as a convenient substrate, are broken into fragments with a
weight average molecular weight of approximately 7 Md, about one-fifth the size
of the intact lambda chromosome.  The reaction requires Mg2+, ATP and
S-adenosylmethionine (SAM).

<>

<1>Yuan, R., Meselson, M.
<2>A specific complex between a restriction endonuclease and its DNA substrate.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>65
<5>357-362
<6>1970
<7>In the presence of Mg++, ATP, and S-adenosylmethionine, the DNA restriction
endonuclease R.K forms a specific complex with its DNA substrate.  The complex
can be detected by its retention on nitrocellulose membranes.

<>

<1>Yuan, R., Reiser, J.
<2>Steps in the reaction mechanism of the Escherichia coli plasmid P15-specific restriction endonuclease.
<3>J. Mol. Biol.
<4>122
<5>433-445
<6>1978
<7>The restriction endonuclease coded by the Escherichia coli plasmid P15 cleaves
unmodified DNA in the presence of ATP and magnesium ions.  This reaction is
stimulated by the addition of S-adenosylmethionine.  Both ATP and
S-adenosylmethionine behave as allosteric effectors.  The enzyme forms a
complex with unmodified DNA in the absence of S-adenosylmethionine and ATP.
Neither the rate of complex formation nor its stability is significantly
affected by S-adenosylmethionine.  The reaction of ATP with this complex is a
late step in the reaction sequence prior to DNA cleavage and is affected by the
presence of S-adenosylmethionine.

<>

<1>Yuan, R., Smith, H.O.
<2>The restriction and modification DNA methylases: An overview.
<3>DNA Methylation. Biochemistry and Biological Significance., Springer-Verlag, Razin, A., Cedar, H., Riggs, A.D., New York
<4>0
<5>73-80
<6>1984
<7>The preceding two chapters have presented in detail all that is known about the
genetic and enzymatic mechanisms of the modification DNA methylases and their
relationship to the homologous restriction endonucleases.  Although the number
of systems that have been characterized is limited, sufficient information is
available to allow a comparison of the three types of restriction-modification
systems from both biological and mechanistic viewpoints.

<>

<1>Yuichi, S., Norton JM, B.A., Klotz, M.G., Stein, L.Y., Laanbroek, H.J., Arp, D.J., Goodwin, L.A., Chertkov, O., Held, B., Bruce, D., Detter, J.C., Detter, J.C., Tapia, R., Han, C.S.
<2>Genome Sequence of Nitrosomonas sp. Strain AL212, an Ammonia-Oxidizing Bacterium Sensitive to High Levels of Ammonia.
<3>J. Bacteriol.
<4>193
<5>5047-5048
<6>2011
<7>Nitrosomonas sp. strain AL212 is an obligate chemolithotrophic ammonia-oxidizing bacterium
(AOB) that was originally isolated in 1997 by
Yuichi Suwa and colleagues. This organism belongs to Nitrosomonas cluster
6A, which is characterized by sensitivity to high ammonia concentrations,
higher substrate affinity (lower K(m)), and lower maximum growth rates
than strains in Nitrosomonas cluster 7, which includes Nitrosomonas
europaea and Nitrosomonas eutropha. Genome-informed studies of this
ammonia-sensitive cohort of AOB are needed, as these bacteria are found in
freshwater environments, drinking water supplies, wastewater treatment
systems, and soils worldwide.

<>

<1>Yukawa, H., Omumasaba, C.A., Nonaka, H., Kos, P., Okai, N., Suzuki, N., Suda, M., Tsuge, Y., Watanabe, J., Ikeda, Y., Vertes, A.A., Inui, M.
<2>Comparative analysis of the Corynebacterium glutamicum group and complete genome sequence of strain R.
<3>Microbiology
<4>153
<5>1042-1058
<6>2007
<7>The complete genome sequence of Corynebacterium glutamicum strain R was determined to allow
its comparative analysis with other corynebacteria.
The biology of corynebacteria was explored by refining the definition of
the subset of genes that constitutes the corynebacterial core as well as
those characteristic of saprophytic and pathogenic ecological niches. In
addition, the relative scarcity of corynebacterial sigma factors and the
plasticity of their two-component system machinery reflect their
relatively exacting nutritional requirements and reduced
membrane-associated and secreted proteins. The conservation of key genes
and pathways between corynebacteria, mycobacteria and Nocardia validates
the use of C. glutamicum to study fundamental processes that are conserved
in slow-growing mycobacteria, including pathogenesis-associated
mechanisms. The discovery of 39 novel genes in C. glutamicum R that have
not been previously reported in other corynebacteria supports the
rationale for sequencing additional corynebacterial genomes to better
define the corynebacterial pan-genome and identify previously undetected
metabolic pathways in these organisms.

<>

<1>Yuki, M., Oshima, K., Suda, W., Kitahara, M., Kitamura, K., Iida, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequences of Two Lactobacillus Strains, L. farraginis JCM 14108T and L. composti JCM 14202T, Isolated from Compost of Distilled Shochu Residue.
<3>Genome Announcements
<4>2
<5>e00257-14
<6>2014
<7>Here, we report the draft genome sequences of two type strains of Lactobacillus,
Lactobacillus farraginis JCM 14108(T) and Lactobacillus composti JCM 14202(T),
isolated from the compost of distilled shochu residue. Their genome information
will be useful for studies of ecological and physiological functions of these
Lactobacillus species.

<>

<1>Yuki, M., Oshima, K., Suda, W., Oshida, Y., Kitamura, K., Iida, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequence of Paenibacillus pini JCM 16418T, Isolated from the Rhizosphere of Pine Tree.
<3>Genome Announcements
<4>2
<5>e00210-14
<6>2014
<7>Paenibacillus pini strain JCM 16418(T) is a cellulolytic bacterium isolated from  the
rhizosphere of pine trees. Here, we report the draft genome sequence of this
strain. This genome information will be useful for studies of rhizosphere
bacteria.

<>

<1>Yuki, M., Oshima, K., Suda, W., Oshida, Y., Kitamura, K., Iida, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequences of Three Alkaliphilic Bacillus Strains, Bacillus wakoensis JCM 9140T, Bacillus akibai JCM 9157T, and Bacillus hemicellulosilyticus JCM  9152T.
<3>Genome Announcements
<4>2
<5>e01258-13
<6>2014
<7>Here, we report the draft genome sequences of the type strains of three cellulolytic or
hemicellulolytic alkaliphilic Bacillus species: Bacillus
wakoensis, Bacillus akibai, and Bacillus hemicellulosilyticus. The genome
information for these three strains will be useful for studies of alkaliphilic
Bacillus species, their evolution, and biotechnological applications for their
enzymes.

<>

<1>Yuki, M., Oshima, K., Suda, W., Sakamoto, M., Iida, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequence of Bacteroides reticulotermitis Strain JCM 10512T, Isolated from the Gut of a Termite.
<3>Genome Announcements
<4>2
<5>e00072-14
<6>2014
<7>Here we report the draft genome sequence of Bacteroides reticulotermitis strain JCM 10512(T),
a xylanolytic and cellulolytic bacterium isolated from the gut of a
wood-feeding termite. The genome information will facilitate the study of this
strain for biomass degradation and adaptation to the gut environment.

<>

<1>Yuki, M., Oshima, K., Suda, W., Sakamoto, M., Kitamura, K., Iida, T., Hattori, M., Ohkuma, M.
<2>Draft Genome Sequence of Clostridium straminisolvens Strain JCM 21531T, Isolated  from a Cellulose-Degrading Bacterial Community.
<3>Genome Announcements
<4>2
<5>e00110-14
<6>2014
<7>Here, we report the draft genome sequence of a fibrolytic bacterium, Clostridium
straminisolvens JCM 21531(T), isolated from a cellulose-degrading bacterial
community. The genome information of this strain will be useful for studies on
the degradation enzymes and functional interactions with other members in the
community.

<>

<1>Yun, J.H., Cho, Y.J., Chun, J., Hyun, D.W., Bae, J.W.
<2>Genome sequence of the chromate-resistant bacterium Leucobacter salsicius type strain M1-8(T.).
<3>Standards in Genomic Sciences
<4>9
<5>495-504
<6>2014
<7>Leucobacter salsicius M1-8(T) is a member of the Microbacteriaceae family within  the class
Actinomycetales. This strain is a Gram-positive, rod-shaped bacterium
and was previously isolated from a Korean fermented food. Most members of the
genus Leucobacter are chromate-resistant and this feature could be exploited in
biotechnological applications. However, the genus Leucobacter is poorly
characterized at the genome level, despite its potential importance. Thus, the
present study determined the features of Leucobacter salsicius M1-8(T), as well
as its genome sequence and annotation. The genome comprised 3,185,418 bp with a
G+C content of 64.5%, which included 2,865 protein-coding genes and 68 RNA genes.
This strain possessed two predicted genes associated with chromate resistance,
which might facilitate its growth in heavy metal-rich environments.

<>

<1>Yun, M.-S., Bae, M.
<2>Purification and characterization of a thermotolerable restriction endonuclease from Streptomyces violochromogenes D2-5.
<3>Proc. Mol. Biol. and Genet.
<4>8
<5>73-74
<6>1993
<7>A thermotolerable restriction endonuclease, SviI, found from Streptomyces violochromogenes
D2-5 was purified.  The purified enzyme was homogeneous and the molecular weight of the
enzymes estimated by polyacrylamide gel electrophoresis containing 0.1% SDS was about 32,000
daltons.  The recognition sequence and cleavage site (indicated by a slash) of this enzyme
were determined to be 5'-TT/CGAA-3', the same sequence of AsuII.

<>

<1>Yun, M.-S., Hwang, H., Bae, M.
<2>Purification and characterization of a thermotolerable restriction endonuclease from Streptomyces violochromogenes D2-5.
<3>J. Microbiol. Biotechnol.
<4>5
<5>269-273
<6>1995
<7>A thermotolerable restriction endonuclease, SviI, found in Streptomyces violochromogenes D2-5
was purified.  For the purification, streptomycin sulfate and ammonium sulfate precipitation
was used.  Phosphocellulose P-11, DEAE-Cellulose and Sephacry1-S200 HR column chromatography
were also performed.  The purified enzyme was found to be homogeneous and the molecular weight
of the enzyme estimated by polyacrylamide gel electrophoresis containing 0.1% SDS was about
32,000 daltons.  The recognition sequence and cleavage site of the enzyme were determined to
be 5M-U-TT/CGAA-3M-U which is the same sequence as that of AsuII.  Unlike AsuII, however, the
SviI shows high thermal stability.

<>

<1>Yung, P.Y., Burke, C., Lewis, M., Egan, S., Kjelleberg, S., Thomas, T.
<2>Phylogenetic screening of a bacterial, metagenomic library using homing endonuclease restriction and marker insertion.
<3>Nucleic Acids Res.
<4>37
<5>e144
<6>2009
<7>Metagenomics provides access to the uncultured majority of the microbial world. The approaches
employed in this field have, however, had limited success in linking functional genes to the
taxonomic or phylogenetic origin of the organism they belong to. Here we present an efficient
strategy to recover environmental DNA fragments that contain phylogenetic marker genes from
metagenomic libraries. Our method involves the cleavage of 23S ribsosmal RNA (rRNA) genes
within pooled library clones by the homing endonuclease I-CeuI followed by the insertion and
selection of an antibiotic resistance cassette. This approach was applied to screen a library
of 6500 fosmid clones derived from the microbial community associated with the sponge
Cymbastela concentrica. Several fosmid clones were recovered after the screen and detailed
phylogenetic and taxonomic assignment based on the rRNA gene showed that they belong to
previously unknown organisms. In addition, compositional features of these fosmid clones were
used to classify and taxonomically assign a dataset of environmental shotgun sequences. Our
approach represents a valuable tool for the analysis of rapidly increasing, environmental DNA
sequencing information.

<>

<1>Yunusova, A.K., Rogulin, E.A., Artyukh, R.I., Zheleznaya, L.A., Matvienko, N.I.
<2>Nickase and a protein encoded by an open reading frame downstream from the nickase BspD6I gene form a restriction endonuclease complex.
<3>Biokhimiia
<4>71
<5>1002-1008
<6>2006
<7>We are the first to have isolated a protein (186 amino acid residues) encoded by the open
reading frame adjacent to the end of the BspD6I nickase (N.BspD6I) gene. Cleavage of both DNA
strands near the sequence recognized by nickase (5'-GAGTC/5'-GACTC) occurs when this protein
is added to the reaction mixture containing N.BspD6I. The protein encoded by the open reading
frame and the nickase are suggested to be subunits of heterodimeric restriction endonuclease
R.BspD6I.

<>

<1>Yussifov, T.N., Zavilgelsky, G.B., Delver, E.P., Belogurov, A.A.
<2>Plasmid pKM101ard+-mediated alleviation of EcoK restriction.  II. Cloning of ard gene.
<3>Mol. Biol. (Mosk)
<4>21
<5>847-852
<6>1987
<7>Plasmid pKM101 affects the type I restriction by EcoK in E. coli.  The gene ard responsible
for alleviation of EcoK restriction was shown to be located within the BglIIB fragment of
pKM101.  Plasmid pD12 was constructed by introducing into pUC12 a 1.87 kb HindIII-Pst
fragment, carrying ard gene.  Tn5 and Tn1000 insertions were obtained in the ard gene region.

<>

<1>Zabaznaya, E.V., Nikiforev, V.V., Zheleznaya, L.A., Matvienko, N.I.
<2>Site-specific endonuclease AbaI from Azospirillum brasilense UQ 1796 is an isoschizomer of endonuclease BclI.
<3>Biokhimiia
<4>62
<5>403-410
<6>1997
<7>The site-specific endonuclease AbaI was isolated and purified to functional purity from the
soil nitrogen-fixing bacterium Azospirillum brasilense UQ 1796.  Purification included
successive chromatography on colums with phosphocellulose, heparin-Sepharose, and
hydroxyapatite.  The purified enzyme recognizes the palindromic DNA sequence 5'-T/GATCA-3'
and cleaves it as shown by the arrow.  The isolated enzyme belongs to class II restriction
endonucleases and is an isoschizomer of endonuclease BclI.  The enzyme AbaI is active at
26-56oC.  The optimal temperature is 48oC and the optimal buffer is LRB.

<>

<1>Zabaznaya, E.V., Zheleznaya, L.A., Matvienko, N.I.
<2>Site-specific endonucleases RspLKI and RspLKII from Rhodococcus species LK2 are isoschizomers of SphI and BamHI.
<3>Biokhimiia
<4>62
<5>1018-1028
<6>1997
<7>Two site-specific endonucleases, RspLKI and RspLKII, have been isolated and purified to
functional homogeneity from the soil bacterium Rhodococcus species LK2.  RspLKI recognizes the
5'-GCATG/C-3' DNA sequence and RspLKII recognizes the 5'-G/GATCC-3' sequence (arrows
indicate DNA cleavage sites).  The isolated enzymes are class II site specific endonucleases
and are isoschizomers of endonucleases SphI and BamHI, respectively.

<>

<1>Zabaznaya, E.V., Zheleznaya, L.A., Svadbina, I.V., Matvienko, N.I.
<2>Site-specific endonuclease NspLKI is an isoschizomer of Endonuclease HaeIII.
<3>Biokhimiia
<4>64
<5>234-238
<6>1999
<7>Site-specific endonuclease NspLKI has been isolated and purified to a functionally pure state
from soil bacterium Nocardia species LK by successive chromatography on columns with
phosphocellulose, HTP hydroxyapatite, and heparin-Sepharose.  The isolated enzyme recognizes
the 5'-GG/CC-3' sequence on DNA and cleaves it as indicated by the arrow, i.e., it is an
isoschizomer of HaeIII.  The final enzyme yield is 1.105 units per gram of wet biomass.  The
enzyme is active in the temperature range of 25-60 C with an optimum at 48-55 C; it does not
lose activity on storage for three days at room temperature.  An optimal buffer is HRB
containing 10 mM Tris-HCl, pH 7.4, 200 microgram/ml albumin, 10 mM MgCl2, and 100 mM NaCl.

<>

<1>Zabeau, M., Friedman, S., Van Montagu, M., Schell, J.
<2>The ral gene of phage lambda: I. Identification of a non-essential gene that modulates restriction and modification in E. coli.
<3>Mol. Gen. Genet.
<4>179
<5>63-73
<6>1980
<7>Host controlled restriction in Escherichia coli can be relieved by
pre-infecting restricting cells with modified lambda helper phages.  This
process, in which intact unmodified phage genomes are allowed to escape
restriction attack, is mediated by a newly identified lambda function called
ral.  The ral gene has been located by deletion mapping between cIII and N.
Efficient expression of the ral gene requires the product of the regulator gene
N.  Polyacrylamide gel analysis of the lambda proteins specified by the cIII-N
region failed to reveal the product of the ral gene, but demonstrated that
protein Ea10 is encoded by a gene located immediately to the left of ral.  From
these results the map order cIII-Ea10-ral-TL1-N was deduced.  Ral specifically
alleviates restriction in E. coli K and E. coli B, but does not affect
restriction systems EcoRI, EcoRII and EcoP1.  In addition, ral enhances the
modification activity of the EcoK and EcoB restriction enzymes:  we observed
that efficient modification of progeny phages obtained by propagating
unmodified lambda phages in r-m+ hosts, is dependent upon the presence of ral.
We thus conclude that the ral gene product acts by modulating the restriction
and modification activities of the type I restriction systems in E. coli, and
the possible mechanisms will be discussed.

<>

<1>Zabeau, M., Roberts, R.J.
<2>The role of restriction endonucleases in molecular genetics.
<3>Molecular Genetics, Academic Press, Inc., Taylor, J.H., New York
<4>3
<5>1-63
<6>1979
<7>This chapter will attempt to provide a summary of the general properties of
restriction enzymes and to describe their various applications in molecular
genetics.  Several earlier reviews have appeared (Arber, 1965, 1971, 1974;
Arber and Linn, 1969; Boyer, 1971; Meselson et al., 1972; Nathans and Smith,
1975; Roberts, 1976).

<>

<1>Zabeau, M., Schell, J., Van Montagu, M.
<2>The alleviation of host-controlled restriction of unmodified phages by functions of bacteriophage lambda.
<3>Arch. Int. Physiol. Biochim.
<4>81
<5>990
<6>1973
<7>The host-controlled restriction of unmodified lambda phage in Escherichia coli
K12 and B hosts can be overcome either by co-infecting these cells with a
modified lambda phage, or by inducing a resident lambda prophage.  The
following lambda functions involved in this rescue process have been
characterised:  (1) a new lambda function ral (for:  Restriction Alleviation)
located between CIII and N.  (2) The non-essential lambda functions red, int
and gam.  The rescue of unmodified phage can proceed by two different
mechanisms:  (1) Whole genome rescue.  The lambda ral function was shown to be
able to rescue both unmodified lambda and unmodified heterologous phages P2 and
Pi.  The ral function probably interferes directly with the restriction
process, since rescue by ral can only be observed when the ral function is
expressed prior to infection with unmodified phage.  Furthermore preliminary
results indicate that the E. coli DNA polymerase I is involved in this rescue
process.  These results suggest that ral interferes with restriction by
directing some repair of the restricted DNA with the result that double strand
scissions do not occur.  2) Using threefactor crosses it was shown that
fragments of restricted DNA can be taken up in helper phage genomes by a
recombination process specifically directed by the lambda red function.  Since
the lambda gam function specifically inhibits the E. coli exonuclease V, we
investigated the role of this nuclease in the restriction process:  it was
shown that exoV is responsible for the secondary degradation of the restriction
DNA fragments.  This results in a loss of biological activity of the fragments
as measured by marker rescue and by complementation.

<>

<1>Zaburannyi, N., Rabyk, M., Ostash, B., Fedorenko, V., Luzhetskyy, A.
<2>Insights into naturally minimised Streptomyces albus J1074 genome.
<3>BMC Genomics
<4>15
<5>11
<6>2014
<7>Background: The Streptomyces albus J1074 strain is one of the most widely used chassis for the
heterologous production of bioactive natural products. The fast growth and an efficient
genetic system make this strain an attractive model for expressing cryptic biosynthetic
pathways to aid drug discovery.Results: To improve its capabilities for the heterologous
expression of biosynthetic gene clusters, the complete genomic sequence of S. albus J1074 was
obtained. With a size of 6,841,649 bp, coding for 5,832 genes, its genome is the smallest
within the genus streptomycetes. Genome analysis revealed a strong tendency to reduce the
number of genetic duplicates. The whole transcriptomes were sequenced at different time points
to identify the early metabolic switch from the exponential to the stationary phase in S.
albus J1074.Conclusions: S. albus J1074 carries the smallest genome among the completely
sequenced species of the genus Streptomyces. The detailed genome and transcriptome analysis
discloses its capability to serve as a premium host for the heterologous production of natural
products. Moreover, the genome revealed 22 additional putative secondary metabolite gene
clusters that reinforce the strain's potential for natural product synthesis.

<>

<1>Zacharias, W., Larson, J.E., Kilpatrick, M.W., Wells, R.D.
<2>HhaI methylase and restriction endonuclease as probes for B to Z DNA conformational changes in d(GCGC) sequences.
<3>Nucleic Acids Res.
<4>12
<5>7677-7692
<6>1984
<7>The capacity of the modification methylase (MHhaI) and restriction endonuclease
(HhaI) from Haemophilus haemolyticus to methylate and cleave, respectively,
recognition sites which are in right-handed B or left-handed Z structures was
determined in vitro.  Plasmids containing tracts of (dC-dG) as well as numerous
individual d(GCGC) sites distributed around the vector were studied.  Negative
supercoiling was used to convert the (dC-dG) tracts (~ 30 bp in length) from a
right-handed to a left-handed conformation.  (Methyl-3H)-SAM was used to
localize and quantitate modified d(GCGC) recognition sites, whereas cleavage by
HhaI was used to detect unmethylated sites.  In the left-handed Z-form, the
(dC-dG) blocks were not methylated by MHhaI and not cleaved by Hha.  A
two-dimensional gel analysis of a family of 33 topoisomers treated with MHhaI
revealed that the lack of methylation in the (dC-dG) blocks was directly
correlated to the supercoil-induced B to Z transition in these segments.  These
results are significant with respect to enzyme-DNA interactions in general and
provide the basis for using HhaI and MHhaI as probes for different DNA
structures and conformational transitions under physiological conditions.

<>

<1>Zacharias, W., O'Connor, T.R., Larson, J.E.
<2>Methylation of cytosine in the 5-position alters the structural and energetic properties of the supercoil-induced Z-helix and B-Z junctions.
<3>Biochemistry
<4>27
<5>2970-2978
<6>1988
<7>The structural and energetic consequences of cytosine methylation in the
5-position on the supercoil-dependent B-Z equilibrium in alternating dC-dG
sequences cloned into recombinant plasmids were investigated.  The helical
parameters determined with the band shift method for right-handed [10.7 base
pairs (bp)/turn] and left-handed (12.8 bp/turn) 5MedC-dG inserts were different
from the helical repeat values for unmethylated dC-dG inserts (10.5 bp/turn in
the right-handed and 11.5 bp/turn in the left-handed form).  We analyzed the
thermodynamic parameters DeltaGBZ (free energy difference per base pair between
right-handed and left-handed helix structure), DeltaGjx (free energy for
formation of one B-Z junction), and b (helix unwinding at a junction region)
for varying lengths of dC-dG inserts by two-dimensional gel electrophoresis and
application of a statistical mechanics model.  A comparison of plasmids fully
methylated in vitro with HhaI methylase and their unmethylated counterparts
revealed that DeltaGjx is not significantly changed by cytosine methylation.
However, this base modification results in an approximate 3-fold decrease of
DeltaGBZ and an approximate 2-fold decrease of the unwinding b at B-Z junction
regions.  Analysis of a pair of related plasmids, each containing two dC-dG
blocks, revealed qualitatively different transition behaviors.  When the two
dC-dG blocks were separated by 95 bp of a mixed sequence, they underwent
independent B to Z transitions with separate nucleation events and junction
formations.  When the two blocks were separated by only a 4 bp GATC sequence,
only one nucleation event was necesary, and the Z-helix spread across the
nonalternating GATC region.  These structural and energetic alterations
demonstrate that methylation of cytosine in the 5-position may be an important
switch mechanism for influencing the B-Z equilibrium and DNA topology in
general, thus potentially affecting DNA-protein interactions and gene
regulation at physiological levels of DNA supercoiling.

<>

<1>Zahner, V., Priest, F.G.
<2>Distribution of restriction nucleases among some entomopathogenic strains of Bacillus sphaericus.
<3>Lett. Appl. Microbiol.
<4>24
<5>483-487
<6>1997
<7>The restriction enzyme BspTI, an isoschizomer of HaeIII (recognition site GGCC), has been
detected in eight strains of serotype 5a5b and two serotype 3 strains of the entomopathogenic
bacterium Bacillus sphaericus.  Strains from other serotypes contained the enzymes BspTII and
BspTIII, which digested pBR322 DNA into similar banding patterns after agarose gel
electrophoresis but differed in their susceptibility to methylation of the substrate.  Strains
from serotypes 9, 25 and 26a26b were lacking in restriction enzyme activity.  There was little
correlation between phage typing and restriction enzyme activity, suggesting that restriction
and modification are not responsible for phage specificity among entomopathogenic B.
sphaericus strains.

<>

<1>Zahran, M., Berezniak, T., Imhof, P., Smith, J.C.
<2>Role of magnesium ions in DNA recognition by the EcoRV restriction endonuclease.
<3>FEBS Lett.
<4>585
<5>2739-2743
<6>2011
<7>The restriction endonuclease EcoRV binds two magnesium ions. One of these ions, Mg-A(2+),
binds to the phosphate group where the cleavage
occurs and is required for catalysis, but the role of the other ion,
Mg-B(2+) is debated. Here, multiple independent molecular dynamics
simulations suggest that Mg-B(2+) is crucial for achieving a tightly
bound protein-DNA complex and stabilizing a conformation that allows
cleavage. In the absence of Mg-B(2+) in all simulations the protein-DNA
hydrogen bond network is significantly disrupted and the sharp kink at
the central base pair step of the DNA, which is observed in the
two-metal complex, is not present. Also, the active site residues
rearrange in such a way that the formation of a nucleophile, required
for DNA hydrolysis, is unlikely.

<>

<1>Zahran, M., Daidone, I., Smith, J.C., Imhof, P.
<2>Mechanism of DNA Recognition by the Restriction Enzyme EcoRV.
<3>J. Mol. Biol.
<4>401
<5>415-432
<6>2010
<7>EcoRV, a restriction enzyme in Escherichia coli, destroys invading foreign DNA by cleaving it
at the center step of a GATATC sequence. In the EcoRV-cognate DNA crystallographic complex, a
sharp kink of 50 degrees has been found at the center base-pair step (TA). Here, we examine
the interplay between the intrinsic propensity of the cognate sequence to kink and the
induction by the enzyme by performing all-atom molecular dynamics simulations of EcoRV unbound
and interacting with three DNA sequences: the cognate sequence, GATATC (TA); the non-cognate
sequence, GAATTC (AT); and with the cognate sequence methylated on the first adenine
GA(CH(3))TATC (TA-CH(3)). In the unbound EcoRV, the cleft between the two C-terminal
subdomains is found to be open. Binding to AT narrows the cleft and forms a partially bound
state. However, the intrinsic bending propensity of AT is insufficient to allow tight binding.
In contrast, the cognate TA sequence is easier to bend, allowing specific, high-occupancy
hydrogen bonds to form in the complex. The absence of cleavage for this methylated sequence is
found to arise from the loss of specific hydrogen bonds between the first adenine of the
recognition sequence and Asn185. On the basis of the results, we suggest a three-step
recognition mechanism. In the first step, EcoRV, in an open conformation, binds to the DNA at
a random sequence and slides along it. In the second step, when the two outer base pairs,
GAxxTC, are recognized, the R loops of the protein become more ordered, forming strong
hydrogen-bonding interactions, resulting in a partially bound EcoRV-DNA complex. In the third
step, the flexibility of the center base pair is probed, and in the case of the full cognate
sequence the DNA bends, the complex strengthens and the protein and DNA interact more closely,
allowing cleavage.

<>

<1>Zain, B.S., Roberts, R.J.
<2>A new specific endonuclease from Xanthomonas badrii.
<3>J. Mol. Biol.
<4>115
<5>249-255
<6>1977
<7>A restriction-like endonuclease, XbaI, has been partially purified from
Xanthomonas badrii.  This enzyme cleaves adenovirus-2 DNA at four sites,
bacteriophage lambda DNA at only one site, and does not cleave simian virus 40
DNA.  It recognizes the sequence 5'-T^-C-T-A-G-A-3' 3'-A-G-A-T-C^-T-5' and cuts
at the sites indicated by the arrows.

<>

<1>Zakharevich, N.V., Averina, O.V., Klimina, K.M., Kudryavtseva, A.V., Kasianov, A.S., Makeev, V.J., Danilenko, V.N.
<2>Complete Genome Sequence of Bifidobacterium longum GT15: Unique Genes for Russian Strains.
<3>Genome Announcements
<4>2
<5>e01348-14
<6>2014
<7>In this study, we report the first completely annotated genome sequence of the Russian-origin
Bifidobacterium longum subsp. longum strain GT15. We discovered 35
unique genes (UGs) which were detected from only the B. longum GT15 genome and
were absent from other B. longum strain genomes (not of Russian origin).

<>

<1>Zakharova, M., Minakhin, L., Solonin, A., Severinov, K.
<2>Regulation of RNA polymerase promoter selectivity by covalent modification of DNA.
<3>J. Mol. Biol.
<4>335
<5>103-111
<6>2004
<7>Expression of genes encoding type II restriction/modification (R/M) systems, which are widely
spread in eubacteria, must be tightly
regulated to ensure that host DNA is protected from restriction
endonucleases at all times. Examples of coordinated expression of R/M
genes that rely on the action of regulatory factors or the ability of
methyl transferases to repress their own synthesis by interacting with
the promoter DNA have been described. Here, we characterize the
molecular mechanism of factor-independent regulation in the CfrBI R/M
system. Regulation of the cfrBIM gene transcription occurs through CfrBIM-catalyzed
methylation of a cytosine residue in the cfrBIM
promoter. The covalent modification inhibits cfrBIM promoter complex
formation by interfering with the RNA polymerase sigma(70) subunit
region 4.2 recognition of the - 35 promoter element. The decrease in
the cfrBIM promoter complex formation leads to increase in the
activity of overlapping cfrBIR promoters. This elegant
factor-independent regulatory system ensures coordinated expression of
the cfrBI genes.

<>

<1>Zakharova, M.V., Beletskaya, I.V., Denjmukhametov, M.M., Yurkova, T.V., Semenova, L.M., Shlyapnikov, M.G., Solonin, A.S.
<2>Characterization of pECL18 and pKPN2: a proposed pathway for the evolution of two plasmids that carry identical genes for a Type II restriction-modification system.
<3>Mol. Genet. Genomics
<4>267
<5>171-178
<6>2002
<7>The primary structures of the plasmids pECL18 (5571 bp) and pKPN2 (4196 bp) from Escherichia
coli and Klebsiella pneumoniae, respectively, which carry genes for a Type II
restriction-modification system (RMS2) with the specificity 5'-CCNGG-3', were determined in
order to elucidate the structural relationship between them. The data suggest a possible role
for recombination events at bom (basis of mobility) regions and the sites of resolution of
multimer plasmid forms (so-called cer sequences) in the structural evolution of multicopy
plasmids. Analysis of the sequences of pECL18 and pKPN2 showed that the genes for RM(.)
Ecl18kI and RM(.) Kpn2kI, and the sequences of the rep (replication) regions in the two
plasmids, are almost identical. In both plasmids, these regions are localized between the bom
regions and the cer sites. The rest of the pECL18 sequence is almost identical to that of the
mob (mobilization) region of ColE1, and the corresponding segment of pKPN2 is almost identical
to part of pHS-2 from Shigella flexneri. The difference in primary structures results in
different mobilization properties of pECL18 and pKPN2. The complete sequences of pECL18, pKPN2
and the pairwise comparison of the sequences of pECL18, pKPN2, ColE1 and pHS-2 suggest that
plasmids may exchange DNA units via site-specific recombination events at bom and cer sites.
In the course of BLASTN database searches using the cer sites of pECL18 and pKPN2 as queries,
we found twenty cer sites of natural plasmids. Alignment of these sequences reveals that they
fall into two classes. The plasmids in each group possess related segments between their cer
and bom sites.

<>

<1>Zakharova, M.V., Beletskaya, I.V., Kravetz, A.N., Pertzev, A.V., Mayorov, S.G., Shlyapnikov, M.G., Solonin, A.S.
<2>Cloning and sequence anlaysis of the plasmid-borne genes encoding the Eco29kI restriction and modification enzymes.
<3>Gene
<4>208
<5>177-182
<6>1998
<7>The Eco29kI restriction-modification system has been found to be localized on the plasmid
pECO29 occurring naturally in the Escherichia coli strain 29k.  Eco29kI, a novel plasmid
encoded restriction endonuclease from Escherichia coli.  The genes coding for this RMS2, a
SacII isoschizomer recognizing the sequence CCGCGG have been cloned in Escherichia coli K802
and sequenced.  The DNA sequence predicts the restriction endonuclease of 214 amino acids
(24,556 Da) and the DNA-methyltransferase of 382 aa (43,007 Da) where the genes are separated
by 2 bp and arranged in tandem with eco29kIR preceding eco29kIM.  The recombinant plasmid with
eco29kIR produces a protein of expected size.  MEco29kI contains all the conserved aa sequence
motifs characteristic of m5C-MTases.  Remarkably, its variable region exhibits a significant
similarity to the part of the specific target-recognition domain from M.BssHII-multispecific
m5C-MTase, which recognizes five different sites on DNA (HaeII, MluI, Cfr10I, SacII and
BssHII), and the comparison of the nt sequences of its variable regions  allowed us to
determine the putative TRD of M.Eco29kI.

<>

<1>Zakharova, M.V., Kravets, A.N., Klimashina, N.V., Solonin, A.S.
<2>Properties of the replicator region of natural plasmid pLG13 carrying genes of the EcoRV restriction-modification system.
<3>Mol. Biol. (Mosk)
<4>25
<5>1615-1625
<6>1991
<7>The previously constructed plasmid pILRV8 that induces endonuclease EcoRV gene
overexpression kills cells of some E. coli strains under the induction of this
enzyme synthesis.  Cell transformation by natural plasmid pLG13 carrying genes
of the EcoRV restriction-modification system was found to appreciably enhance
cell viability (survival) under endonuclease overproduction.  A plasmid pLG13
region located in immediate proximity to the methylase gene was shown to be
responsible for the above effect.  This region was also capable of autonomous
replication.  The analysis of the DNA primary structure in the found replicator
region allowed to refer the pLG13 to ColE1 family plasmids.  Perturbations in
the region lead to loss of the survival effect and change of the plasmid
replicative properties.  A relationship between the replicon elements, the
EcoRV genes region and survival effect is discussed.  Based on the replicon
found multicopy vector molecules have been constructed.

<>

<1>Zakharova, M.V., Kravetz, A.N., Beletzkaja, I.V., Repyk, A.V., Solonin, A.S.
<2>Cloning and sequences of the genes encoding the CfrBI restriction-modification system from Citrobacter freundii.
<3>Gene
<4>129
<5>77-81
<6>1993
<7>The genes encoding the CfrBI restriction and modification (R-M) systems from Citrobacter
freundii and recognizing the sequences 5'-CCWWGG-3' (W=A or T) were cloned in Escherichia
coli McrBC- cells. The nucleotide (nt) sequences of the genes were determined. Two large open
reading frames were found. Deletion analysis showed that one of them [1128 nt coding for 376
amino acids (aa)] corresponds to a methyltransferase (MTase)-encoding gene and the other (1065
nt coding for 355 aa) to a restriction endonuclese-encoding gene. The genes are oriented
divergently and separated by 76 bp. A CfrBI site (5'-m4CCATGG) was found in the intergenic
region of the cfrBIRM genes. Analysis of the deduced aa sequence of M.CfrBI made it possible
to determine the typical features of a m4C-specific MTase. Limited homology between the
M.CfrBI and R.CfrBI proteins was also found.

<>

<1>Zakharova, M.V., Pertzev, A.V., Kravetz, A.N., Beletskaya, I.V., Shlyapnikov, M.G., Solonin, A.S.
<2>Complete nucleotide sequence of the Hsd plasmid pECO29 and identification of its functional regions.
<3>Biochim. Biophys. Acta
<4>1398
<5>106-112
<6>1998
<7>The complete nucleotide sequence of the Hsd plasmid pECO29 has been determined.  The plasmid
DNA consists of 3895 base pairs.  These include 4 genes and 5 sites.  Two genes encoding the
proteins (restriction endonuclease and DNA methyltransferase) have been fully characterized.
The pECO29 comprises a ColE1-type replication system coding for untranslated genes RNAI and
RNAII, the emr recombination site containing palindromic sequences and involved in stable
maintenance of the plasmid, two pseudo oriT sites homologous to the oriT site of R64 and F
plasmids, as well as the bom locus of a ColE1-like plasmid.  There are no genes involved in
the mobilization of pECO29 plasmid.

<>

<1>Zaleski, P., Piekarowicz, A.
<2>Characterization of a dam mutant of Haemophilus influenzae Rd.
<3>Microbiology
<4>150
<5>3773-3781
<6>2004
<7>The gene encoding Dam methyltransferase of Haemophilus influenzae was mutagenized by the
insertion of a chloramphenicol-resistance cassette into the middle of the Dam coding sequence.
This mutant construct was introduced into the H. influenzae chromosome by transformation and
selection for Cam(R) transformants. The authors have shown that several phenotypic properties,
resistance to antibiotics, dyes and detergent as well as efficiency of transformation, depend
on the Dam methylation state of the DNA. Although the major role of the methyl-directed
mismatch repair (MMR) system is to repair postreplicative errors, it seems that in H.
influenzae its effect is more apparent in repairing DNA damage caused by oxidative compounds.
In the dam mutant treated with hydrogen peroxide, MMR is not targeted to newly replicated DNA
strands and therefore mismatches are converted into single- and double-strand DNA breaks. This
is shown by the increased peroxide sensitivity of the dam mutant and the finding that the
sensitivity can be suppressed by a mutH mutation inactivating MMR. In the dam mutant treated
with nitrofurazone the resulting damage is not converted into DNA breaks but the high
sensitivity is also suppressed by a mutH mutation.

<>

<1>Zaleski, P., Wojciechowski, M., Piekarowicz, A.
<2>The role of Dam methylation in phase variation of Haemophilus influenzae genes involved in defence against phage infection.
<3>Microbiology
<4>151
<5>3361-3369
<6>2005
<7>Haemophilus influenzae uses phase variation (PV) to modulate the activity of its defence
systems against phage infection. The PV of the
restriction-modification (R-M) system Hindl, the main defence system
against phage infection and incoming chromosomal and phage DNA in H.
influenzae Rd, is driven by changes of the pentanucleoticle repeat
tract within the coding sequence of the hsdM gene and is influenced by
lack of Dam methylation. Phase-variable resistance/sensitivity to phage
infection correlates with changes in lipooligosaccharide (LOS)
structure and occurs by slippage of tetranucleotide repeats within the
gene lic2A, coding for a step in the biosynthesis of LOS. The lack of
Dam activity destabilizes the tetranuclotide (5'-CAAT) repeat tract and
increases the frequency of switching from sensitivity to resistance to
phage infection more than in the opposite direction. The PV of the lgtC
gene does not influence resistance or sensitivity to phage infection.
Insertional inactivation of lic2A, but not lgtC or lgtF, leads to
resistance to phage infection and to the same structure of the LOS as
observed among phase-variable phage-resistant variants. This indicates
that in the H. influenzae Rd LOS only the first two sugars (Glc-Gal)
extending from the third heptose are part of bacterial phage receptors.

<>

<1>Zan, J., Fricke, W.F., Fuqua, C., Ravel, J., Hill, R.
<2>Genome Sequence of Ruegeria sp. strain KLH11, an N-Acylhomoserine Lactone-Producing Bacterium Isolated from the Marine Sponge Mycale  laxissima.
<3>J. Bacteriol.
<4>193
<5>5011-5012
<6>2011
<7>Ruegeria sp. KLH1, isolated from marine sponge Mycale laxissima, produces a complex profile of
N-acylhomoserine lactone quorum sensing (QS)
molecules. The genome sequence provides insights into the genetic
potential of KLH11 to maintain complex QS systems and is the first genome
report of a cultivated symbiont from a marine sponge.

<>

<1>Zangi, R., Arrieta, A., Cossio, F.P.
<2>Mechanism of DNA Methylation: The Double Role of DNA as a Substrate and as a Cofactor.
<3>J. Mol. Biol.
<4>400
<5>632-644
<6>2010
<7>Methylation of cytosine residues in the DNA is one of the most important epigenetic marks
central to the control of differential expression of
genes. We perform quantum mechanical calculations to investigate the
catalytic mechanism of the bacterial HhaI DNA methyltransferase. We find
that the enzyme nucleophile, Cys81, can attack C6 of cytosine only after
it is deprotonated by the DNA phosphate group, a reaction facilitated by a
bridging water molecule. This finding, which indicates that the DNA acts
as both the substrate and the cofactor, can explain the total loss of
activity observed in an analogous enzyme, thymidylate synthase, when the
phosphate group of the substrate was removed. Furthermore, our results
displaying the inability of the phosphate group to deprotonate the side
chain of serine is in agreement with the total, or the large extent of,
inactivity observed for the C81S mutant. In contrast to results from
previous calculations, we find that the active site conserved residues,
Glu119, Arg163, and Arg165, are crucial for catalysis. In addition, the
enzyme-DNA adduct formation and the methyl transfer from the cofactor
S-adenosyl-l-methionine are not concerted but proceed via stepwise
mechanism. In many of the different steps of this methylation reaction,
the transfer of a proton is found to be necessary. To render these
processes possible, we find that several water molecules, found in the
crystal structure, play an important role, acting as a bridge between the
donating and accepting proton groups.

<>

<1>Zaremba, M., Owsicka, A., Tamulaitis, G., Sasnauskas, G., Shlyakhtenko, L.S., Lushnikov, A.Y., Lyubchenko, Y.L., Laurens, N., van den Broek, B., Wuite, G.J., Siksnys, V.
<2>DNA synapsis through transient tetramerization triggers cleavage by Ecl18kI restriction enzyme.
<3>Nucleic Acids Res.
<4>38
<5>7142-7154
<6>2010
<7>To cut DNA at their target sites, restriction enzymes assemble into different oligomeric
structures. The Ecl18kI endonuclease in the crystal
is arranged as a tetramer made of two dimers each bound to a DNA copy.
However, free in solution Ecl18kI is a dimer. To find out whether the
Ecl18kI dimer or tetramer represents the functionally important assembly,
we generated mutants aimed at disrupting the putative dimer-dimer
interface and analysed the functional properties of Ecl18kI and mutant
variants. We show by atomic force microscopy that on two-site DNA, Ecl18kI
loops out an intervening DNA fragment and forms a tetramer. Using the
tethered particle motion technique, we demonstrate that in solution DNA
looping is highly dynamic and involves a transient interaction between the
two DNA-bound dimers. Furthermore, we show that Ecl18kI cleaves DNA in the
synaptic complex much faster than when acting on a single recognition
site. Contrary to Ecl18kI, the tetramerization interface mutant R174A
binds DNA as a dimer, shows no DNA looping and is virtually inactive. We
conclude that Ecl18kI follows the association model for the synaptic
complex assembly in which it binds to the target site as a dimer and then
associates into a transient tetrameric form to accomplish the cleavage
reaction.

<>

<1>Zaremba, M., Sasnauskas, G., Siksnys, V.
<2>The link between restriction endonuclease fidelity and oligomeric state: A study with Bse634I.
<3>FEBS Lett.
<4>586
<5>3324-3329
<6>2012
<7>Type II restriction endonucleases (REases) exist in multiple oligomeric forms. The tetrameric
REases have two DNA binding interfaces and must
synapse two recognition sites to achieve cleavage. It was hypothesised
that binding of two recognition sites by tetrameric enzymes contributes
to their fidelity. Here, we experimentally determined the fidelity for
Bse634I REase in different oligomeric states. Surprisingly, we find
that tetramerisation does not increase REase fidelity in comparison to
the dimeric variant. Instead, an inherent ability to act concertedly at
two sites provides tetrameric REase with a safety-catch to prevent host
DNA cleavage if a single unmodified site becomes available.

<>

<1>Zaremba, M., Sasnauskas, G., Urbanke, C., Siksnys, V.
<2>Conversion of the tetrameric restriction endonuclease Bse634I into a dimer: Oligomeric structure-stability-function correlations.
<3>J. Mol. Biol.
<4>348
<5>459-478
<6>2005
<7>The Bse634I restriction endonuclease is a tetramer and belongs to the type IIF subtype of
restriction enzymes. It requires two recognition
sites for its optimal activity and cleaves plasmid DNA with two sites
much faster than a single-site DNA. We show that disruption of the
tetramerisation interface of Bse634I by site-directed mutagenesis
converts the tetrameric enzyme into a dimer. Dimeric W228A mutant
cleaves plasmid DNA containing one or two sites with the same
efficiency as the tetramer cleaves the two-site plasmid. Hence, the
catalytic activity of the Bse634I tetramer on a single-site DNA is
down-regulated due to the cross-talking interactions between the
individual dimers. The autoinhibition within the Bse634I tetramer is
relieved by bridging two DNA copies into the synaptic complex that
promotes fast and concerted cleavage at both sites. Cleavage analysis
of the oligonucleotide attached to the solid support revealed that
Bse634I is able to form catalytically competent synaptic complexes by
bridging two molecules of the cognate DNA, cognate DNA-miscognate DNA
and cognate DNA-product DNA. Taken together, our data demonstrate that
a single W228A mutation converts a tetrameric type IIF restriction
enzyme Bse634I into the orthodox dimeric type IIP restriction
endonuclease. However, the stability of the dimer towards chemical
denaturants, thermal inactivation and proteolytic degradation are
compromised.

<>

<1>Zaremba, M., Sasnauskas, G., Urbanke, C., Siksnys, V.
<2>Allosteric communication network in the tetrameric restriction endonuclease Bse634I.
<3>J. Mol. Biol.
<4>363
<5>500-512
<6>2006
<7>Restriction endonuclease Bse634I is a homotetramer arranged as a dimer of two primary dimers.
Bse634I displays its maximum catalytic efficiency upon
binding of two copies of cognate DNA, one per each primary dimer. The
catalytic activity of Bse634I on a single DNA copy is down-regulated due
to the cross-talking interactions between the primary dimers. The
mechanism of signal propagation between the individual active sites of
Bse634I remains unclear. To identify communication pathways involved in
the catalytic activity regulation of Bse634I tetramer we mutated a
selected set of amino acid residues at the dimer-dimer interface and
analysed the oligomeric state and catalytic properties of the mutant
proteins. We demonstrate that alanine replacement of N262 and V263
residues located in the loop at the tetramerisation interface did not
inhibit tetramer assembly but dramatically altered the catalytic
properties of Bse634I despite of the distal location from the active site.
Kinetic analysis using cognate hairpin oligonucleotide and one and
two-site plasmids as substrates allowed us to identify two types of
communication signals propagated through the dimer-dimer interface in the
Bse634I tetramer: the inhibitory, or "stopper" and the activating, or
"sync" signal. We suggest that the interplay between the two signals
determines the catalytic and regulatory properties of the Bse634I and
mutant proteins.

<>

<1>Zaremba, M., Siksnys, V.
<2>Molecular scissors under light control.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>107
<5>1259-1260
<6>2010
<7>Light plays a key role in the living world.  It serves as a source of energy through
photosynthesis, makes sight possible, and regulates circadian rhythms.  During evolution,
organisms from bacteria to higher mammals elaborate various mechanisms to sense and respond to
light, which manipulates their behavior.  The use of light as a trigger is particularly
attractive as a means of controlling biological processes at will, because light is
noninvasive and can be manipulated both temporally (from microseconds) and spatially
(microns).  In this issue of PNAS, Schierling et al. demonstrate the possibility of
controlling the enzymatic activity of the restriction endonuclease PvuII by light.

<>

<1>Zaremba, M., Toliusis, P., Grigaitis, R., Manakova, E., Silanskas, A., Tamulaitiene, G., Szczelkun, M.D., Siksnys, V.
<2>DNA cleavage by CgII and NgoAVII requires interaction between N- and R-proteins and extensive nucleotide hydrolysis.
<3>Nucleic Acids Res.
<4>42
<5>13887-13896
<6>2014
<7>The stress-sensitive restriction-modification (RM) system CglI from Corynebacterium glutamicum
and the homologous NgoAVII RM system from Neisseria gonorrhoeae FA1090 are composed of three
genes: a DNA methyltransferase (M.CglI and M.NgoAVII), a putative restriction endonuclease
(R.CglI and R.NgoAVII, or R-proteins) and a predicted DEAD-family helicase/ATPase (N.CglI and
N.NgoAVII or N-proteins). Here we report a biochemical characterization of the R- and
N-proteins. Size-exclusion chromatography and SAXS experiments reveal that the isolated
R.CglI, R.NgoAVII and N.CglI proteins form homodimers, while N.NgoAVII is a monomer in
solution. Moreover, the R.CglI and N.CglI proteins assemble in a complex with R2N2
stoichiometry. Next, we show that N-proteins have ATPase activity that is dependent on
double-stranded DNA and is stimulated by the R-proteins. Functional ATPase activity and
extensive ATP hydrolysis ( approximately 170 ATP/s/monomer) are required for site-specific DNA
cleavage by R-proteins. We show that ATP-dependent DNA cleavage by R-proteins occurs at fixed
positions (6-7 nucleotides) downstream of the asymmetric recognition sequence 5'-GCCGC-3'.
Despite similarities to both Type I and II restriction endonucleases, the CglI and NgoAVII
enzymes may employ a unique catalytic mechanism for DNA cleavage.

<>

<1>Zaremba, M., Urbanke, C., Halford, S.E., Siksnys, V.
<2>Generation of the BfiI restriction endonuclease from the fusion of a DNA recognition domain to a non-specific nuclease from the  phospholipase D superfamily.
<3>J. Mol. Biol.
<4>336
<5>81-92
<6>2004
<7>The Bfi I endonuclease cleaves DNA at fixed positions downstream of an asymmetric sequence.
Unlike other restriction enzymes, it functions
without metal ions. The N-terminal half of Bfi I is similar to Nuc, an
EDTA-resistant nuclease from Salmonella typhimurium that belongs to the
phosphoplipase D superfamily. Nuc is a dimer with one active site at
its subunit interface, as is Bfi I, but it cuts DNA non-specifically.
Bfi I was cleaved by thermolysin into an N-terminal domain, which forms
a dimer with non-specific nuclease activity, and a C-terminal domain,
which lacks catalytic activity but binds specifically to the
recognition sequence as a monomer. On denaturation with guanidinium,
Bfi I underwent two unfolding transitions: one at a relatively low
concentration of guanidinium, to a dimeric non-specific nuclease; a
second at a higher concentration, to an inactive monomer. The isolated
C-terminal domain unfolded at the first (relatively low) concentration,
the isolated N-terminal at the second. Hence, Bfi I consists of two
physically separate domains, with catalytic and dimerisation functions
in the N terminus and DNA recognition functions in the C terminus. It
is the first example of a restriction enzyme generated by the
evolutionary fusion of a DNA recognition domain to a phosphodiesterase
from the phospholipase D superfamily. Bfi I may consist of three
structural units: a stable central core with the active site, made from
two copies of the N-terminal domain, flanked by relatively unstable
C-terminal domains, that each bind a copy of the recognition sequence.
(C) 2003 Elsevier Ltd. All rights reserved.

<>

<1>Zarogiannis, S.G., Filippidis, A.S.
<2>Resenbase: A sophisticated and user-friendly, restriction endonuclease database compilation for handheld computers.
<3>Clin. Chim. Acta
<4>355
<5>S360
<6>2005
<7>A database of commonly used restriction enzymes was constructed and programmed into a PalmOs
handheld.

<>

<1>Zarrilli, R., Giannouli, M., Rocco, F., Loman, N.J., Haines, A.S., Constantinidou, C., Pallen, M.J., Triassi, M., Di Nocera, P.P.
<2>Genome sequences of three Acinetobacter baumannii strains assigned to ST2, ST25 and ST78 multilocus sequencing typing genotypes.
<3>J. Bacteriol.
<4>193
<5>2359-2360
<6>2011
<7>Acinetobacter baumannii is an emerging opportunistic gram-negative pathogen responsible for
hospital-acquired infections. A. baumannii epidemics described in Europe and worldwide were
caused by a limited number of genotypic clusters of multidrug-resistant strains. Here we
report the availability of draft genome sequences for three multidrug-resistant A. baumannii
strains assigned to ST2, ST25 and ST78 multilocus sequencing typing genotypes, that were more
frequently isolated during outbreaks occurred in Greece, Italy, Lebanon and Turkey.

<>

<1>Zatkovic, B., Molnarova, V., Kmet, V., Javorsky, P., Pristas, P.
<2>Diversity of DNA sequences among restriction endonucleases producing Selenomonas ruminantium isolates detected by enterobacterial repetitive intergenic consensus based polymerase chain reaction (ERIC-PCR).
<3>Anaerobe
<4>6
<5>299-304
<6>2000
<7>Enterobacterial repetitive intergenic consensus-based polymerase chain reaction (ERIC-PCR) was
found useful for discrimination of rumen
selenomonads. Simultaneous use of ERICIR and ERIC2 primers yielded
strain-specific banding patterns. The patterns were compared using Dice
similarity coefficients and a DNA relatedness dendrogram based on the
unweighted pair group method using arithmetic averages (UPGMA) was
constructed. Five clusters and four single strains were identified at a
similarity level of 50%. Very weak grouping was observed for
lactilytica and ruminantium subspecies of Selenomonas ruminantium,
indicating that lactate utilization has probably no taxonomic value.
Restriction and modification phenotypes are weakly reflected in the
dendrogram probably as the result of horizontal genetic transfer of
genes encoding these phenotypic traits. While diverse in ERIC-PCR
analysis, strains shown little variation in restriction fragment length
polymorphism of amplified 16S-rRNA genes. All but one strain produced
nearly identical profiles indicating that majority of DNA diversity
observed is due to epigenetic factors and not due to evolutionary
divergence.

<>

<1>Zavilgelskii, G.B., Bakalova, T.L., Duzhii, D.E., Kotova, V.Y.
<2>The ard gene encoding the type I restriction inhibitor is present in conjugative plasmids of FII, B/O, and K groups of incompatibility.
<3>Genetika
<4>30
<5>1582-1586
<6>1994
<7>The effect of conjugative plasmids of various incompatibility groups of the enterobacteria
family on the activity of the cell restriction-modification system of type I (EcoK) was
studied. Twenty-two conjugative plasmids of 15 incompatibility groups were tested. In addition
to plasmids of the incI1 and incN groups studied earlier, conjugative plasmids of the incFII,
incB/O, and incK groups were also shown to be able to weaken the action of type I restriction
enzymes upon nonmodified DNA (Ard phenotype). A hybridization analysis of all the plasmid DNAs
studied, using ard gene DNA sequences from the ColIb-P9 (incI1) plasmid as a probe, was
performed. The ard locus of the R100 (incFII) plasmid was cloned in the pBR322 and pACYC184
vectors. The ard gene was located 2.5 kb from the oriT site in the leading region on the R100
conjugative plasmid.

<>

<1>Zavilgelskii, G.B., Manukhov, I.V., Rastorguev, S.M.
<2>Attenuation of type I restriction in Escherichia coli: effect of the ard gene in UV-irradiated cells.
<3>Genetika
<4>32
<5>1013-1016
<6>1996
<7>The effect of conjugative plasmids ColIb-P9 (incI1) and pKM 101 (incN), containing the active
ard gene, on the efficiency of EcoK restriction of nonmodified phage lambda .0 in
UV-irradiated Escherichia coli cells was studied. ard-Dependent antirestriction enzyme
activity was shown to decrease in UV-irradiated cells. The efficiency of action of the ard
plasmid gene on lambda .0 was also shown not to depend on cell helicases RecBCD and UvrD, in
contrast to the UV-induced alleviation of EcoK restriction (SOS alleviation).
[Article in Russian]

<>

<1>Zavilgelsky, G.B.
<2>Antirestriction.
<3>Mol. Biol. (Mosk)
<4>34
<5>854-862
<6>2000
<7>Modern data on the molecular mechanisms of antirestriction are reviewed.  The systems of
inhibition are described for the restriction-modification enzymes of type I in phages and in
conjugative plasmids.  The phenomenon of alleviation of DNA restriction and its mechanism is
presented for bacterium Escherichia coli.  The principle of protein mimicry of nucleic acids
is discussed as a novel way to control biological processes in the cell with reference to
antirestriction proteins Ard.

<>

<1>Zavilgelsky, G.B., Kotova, V.Y.
<2>Antirestriction activity of the monomeric and dimeric forms of T7 Ocr.
<3>Mol. Biol. (Mosk)
<4>48
<5>150-157
<6>2014
<7>The Ocr antirestriction protein, which is encoded by bacteriophage T7 0.3 (ocr), specifically
inhibits type I restriction-modification enzymes. Ocr belongs to a family of DNA-mimicking
proteins. Native Ocr forms homodimers both in solution and in crystal. Ocr mutants with two
amino acid substitutions (Orc F53D A57E and Ocr F53R V77D) were constructed to occur as
monomers in solution. The dissociation constant K (d) for the Ocr complex with EcoKI (R2M2S)
proved to differ by three orders of magnitude between the (Ocr)(2) dimer and Ocr F53D A57E and
Ocr F53R V77D monomers (10(-10) M vs. 10(-7) M). Antimodification activity was substantially
lower in the Ocr monomers. The dimeric form found to be essential for high inhibitory activity
of Ocr.

<>

<1>Zavilgelsky, G.B., Kotova, V.Y., Melkina, O.E., Pustovoit, K.S.
<2>Antirestriction activity of the mercury resistance nonconjugative transposon Tn5053 is controlled by the protease ClpXP.
<3>Genetika
<4>50
<5>910-915
<6>2014
<7>When transformed into Escherichia coli K12 strains, the mercury resistance transposon Tn5053
exhibits high antirestriction activity against the EcoKI type I restriction and modification
system. The products of the genes merR and ardD contribute to the antirestriction activity of
Tn5053. The merR gene encodes the MerR protein, the transcription regulator of the mer operon
genes. The ardD gene is responsible for ArdD protein synthesis and is located within the tni
operon. In the following study, it was demonstrated that the antirestriction activity of the
transposon Tn5053 is absent in E. coli K12 strains with the mutant genes clpX, clpP, and recA.
The antirestriction effect of Tn5053 is not enhanced by 2-aminopurine. The Tn5053
antirestriction activity is not altered in E. coli K12 with the mutant dam gene; however, it
is decreased in the E. coli K12 mutD. It is assumed that the activities of the MerR and ArdD
proteins lead to the formation of a significant amount of unmodified DNA in the bacterial
cell, causing the SOS-dependent reduction of the EcoKI (R2M2S) enzyme activity associated with
ClpXP-induced proteolysis of the R-subunit.

<>

<1>Zavilgelsky, G.B., Kotova, V.Y., Rastorguev, S.M.
<2>Antirestriction and antimodification activities of T7 Ocr: Effects of amino acid substitutions in the interface.
<3>Mol. Biol. (Mosk)
<4>43
<5>93-100
<6>2009
<7>The T7 antirestriction protein Ocr, encoded by 0.3 (ocr), specifically inhibits ATP-dependent
type I restriction-modification systems. T7 0.3
(ocr) was cloned in pUC18. Ocr inhibited both restriction and
modification activities of the type I restriction-modification system
(EcoKI) in Escherichia coli K12. The Ocr F53D A57E mutant was obtained
and proved to inhibit only restriction activity of EcoKI. The 0.3 (ocr)
and Photorhabdus luminescens luxCDABE genes were cloned in pZ-series
vectors with the P (ltetO-1) promoter, strongly controlled by the TetR
repressor. The bioluminescence intensity and luciferase content varied
up to 5000-fold in E. coli K12 MG1655Z1 tetR(+) (pZE21-luxCDABE) cells,
depending on the environmental concentration of the inductor
anhydrotetracycline. The antirestriction activity of Ocr and Ocr F53D
A57E was studied as a function of their concentration in the cell. The
dissociation constant K (d), characterizing the binding with EcoKI,
differed 1000-fold between Ocr and Ocr F53D A57E (10(-10) M versus 10(-7) M).

<>

<1>Zavilgelsky, G.B., Kotova, V.Y., Rastorguev, S.M.
<2>Comparative analysis of anti-restriction activities of ArdA (ColIb-P9) and Ocr (T7) proteins.
<3>Biochemistry
<4>73
<5>906-911
<6>2008
<7>Anti-restriction proteins ArdA and Ocr are specific inhibitors of type I
restriction-modification enzymes. The IncI1 transmissible plasmid
ColIb-P9 ardA and bacteriophage T7 0.3(ocr) genes were cloned in pUC18
vector. Both ArdA (ColIb-P9) and Ocr (T7) proteins inhibit both
restriction and modification activities of the type I
restriction-modification enzyme (EcoKI) in Escherichia coli K12 cells.
ColIb-P9 ardA, T7 0.3(ocr), and the Photorhabdus luminescens luxCDABE
genes were cloned in pZ-series vectors with the PltetO-1 promoter,
which is tightly repressible by the TetR repressor. Controlling the
expression of the lux-genes encoding bacterial luciferase demonstrates
that the PltetO-1 promoter can be regulated over an up to 5000-fold
range by supplying anhydrotetracycline to the E. coli MG1655Z1 tetR(+)
cells. Effectiveness of the anti-restriction activity of the ArdA and
Ocr proteins depended on the intracellular concentration. It is shown
that the dissociation constants K (d) for ArdA and Ocr proteins with
EcoKI enzyme differ 1700-fold: K (d)(Ocr) = 10(-10) M, K (d)(ArdA) =
1.7 center dot 10(-7) M.

<>

<1>Zavilgelsky, G.B., Kotova, V.Y., Rastorguev, S.M.
<2>Antimodification activity of the ArdA and Ocr proteins.
<3>Genetika
<4>47
<5>159-167
<6>2011
<7>The ArdA and Ocr antirestriction proteins, whose genes are in transmissible plasmids (ardA)
and bacteriophage genomes (0.3 (ocr)), specifically inhibit type I restriction-modification
enzymes. The Ocr protein (T7 bacteriophage) was shown to inhibit both restriction
(endonuclease) and modification (methylase) activities of the EcoKI enzyme in a broad range of
intracellular concentrations (starting from 10-20 molecules per cell). In contrast to Ocr, the
ArdA protein (ColIb-P9 transmissible plasmid) inhibited both of the EcoKI activities only at
high intracellular concentrations (30000-40000 molecules per cell). When the ArdA
concentration was several fold lower, only endonuclease activity of EcoKI was inhibited. It
was assumed that a poorer ArdA ability to inhibit EcoKI modification activity is related to
the substantial difference in life cycle between transmissible plasmids (symbiosis with the
bacterial cell) and bacteriophages (infection and lysis of bacteria). The Ocr and ArdA mutants
that inhibited exclusively endonuclease activity of EcoKI were obtained. Antirestriction
proteins incapable of homodimerization were assumed to inhibit only endonuclease activity of
type I restriction-modification enzymes.

<>

<1>Zavilgelsky, G.B., Letuchaya, T.A., Rastorguev, S.M.
<2>Antirestriction and antimodification activities of the ArdA protein encoded by the IncI1 transmissive plasmids R-64 and ColIb-P9.
<3>Genetika
<4>42
<5>252-258
<6>2006
<7>Proteins of the Ard family are specific inhibitors of type I restriction-modification enzymes.
The ArdA of R64 is highly homologous
to ColIb-P9 ArdA, differing only by four amino acid residues of the
overall 166. However, unlike ColIb-P9 ArdA, which inhibits both the
endonuclease and the methylase activities of EcoKI, the R64 ArdA
protein inhibits only the endonuclease activity of this enzyme. The
mutant forms of R64 ArdA-A29T, S43A, and Y75W, capable of partially
reversing the protein to ColIb-P9 ArdA form-were produced by directed
mutagenesis. It was demonstrated that only Y75W mutation of these three
variants essentially influenced the functional activity of ArdA: the
antimodification activity was restored to approximately 90%. It is
assumed that R64 ArdA inhibits formation of the complex between
unmodified DNA and the R subunit of the type I restriction-modification
enzyme EcoKI (R2M2S), which translocates and cleaves DNA. ColIb-P9 ArdA
protein is capable of forming the DNA complex not only with the R
subunit, but also with the S subunit, which contacts sK site
(containing modified adenine residues) in DNA. ArdA bound to the
specific sK site inhibits concurrently the endonuclease and methylase
activities of EcoKI (R2M2S), while ArdA bound to the nonspecific site
in the R subunit blocks only its endonuclease activity.

<>

<1>Zavilgelsky, G.B., Letutchaja, T.A., Rastorguev, S.M.
<2>Antirestriction activity of ArdA encoded by the IncI1 transmissive plasmid R64.
<3>Mol. Biol. (Mosk)
<4>38
<5>901-906
<6>2004
<7>The transmissive plasmid R64 (IncI1) performs an antirestriction function, reducing the
efficiency of EcoKI-dependent restriction in
Escherichia coli K12 cells approximately fivefold. The R64 ardA gene
has been cloned and sequenced. The ArdA proteins specifically inhibit
type I restriction-modification enzymes. R64 ArdA is highly homologous
to ColIb-P9 ArdA: only 4 out of 166 amino acid residues differ. While
ColIb-P9 inhibits both endonuclease and methylase activities of the
type I restriction-modification enzyme EcoKI (R2M2S), R64 ArdA inhibits
only its endonuclease activity. It has been assumed that R64 ArdA
suppresses the binding of unmodified DNA with the R subunit, which is
responsible for DNA translocation and cleavage. ColIb-P9 ArdA
suppresses DNA binding not only with the R, but also with the S
subunit, which contacts the sK site containing target adenines. The
binding of ArdA with the specific site inhibits both endonuclease and
methylase activities; the binding of ArdA with the nonspecific site of
the R subunit inhibits only the endonuclease activity of EcoKI (R2M2S).

<>

<1>Zavilgelsky, G.B., Mershavka, V.Y., Jusifov, T.N., Belogurov, A.A.
<2>Plasmid pKM101 ard+-mediated alleviation of K-restriction of bacteriophage lambda.  I. General characteristics and genetic localization of the ard gene.
<3>Mol. Biol. (Mosk)
<4>18
<5>1590-1596
<6>1984
<7>The effectiveness of action of restriction endonuclease EcoK on DNA lambda.0 is considerably
alleviated (by 10 to 100 times approximately) if Escherichia  coli K12 cells contain plasmid
pKM101.  The region of plasmid pKM101 in which insertion sites of transposon Tn5, disturbing
the ability of the plasmid to alleviate EcoK restriction are located, was found to be in
fragment BglII-B at a distance of about 1.3-1.7 kilobases from the end of the fragment, and it
was named ard (alleviation of restriction of DNA).  Alleviation of K restriction of phage
lambda.0 determined by plasmid ard gene is independent of chromosomal genes lexA, recBC, and
umuC and plasmid gene muc.  The presence of plasmid pKM101 ard+ in the cell does not lead to
specific modification of DNA, nor does it intensify methylation of DNA by methylase K like the
action of the ral gene of phage lambda.  The action of the ard gene of plasmid pKM101 is
highly specific: Alleviation of DNA restriction is observed only in strains of E. coli K and
is virtually absent in strains of E. coli B, and also in strains of E. coli containing class
II restriction endonucleases (EcoRI and EcoRIII).

<>

<1>Zavilgelsky, G.B., Rastorguev, S.M.
<2>DNA mimicry by proteins as effective mechanism for regulation of activity of DNA-dependent enzymes.
<3>Biochemistry
<4>72
<5>913-919
<6>2007
<7>Modern concepts on mechanisms of DNA-dependent enzyme regulation involving specific
DNA-mimicking proteins are considered. There are proteins that share structural resemblance
with DNA duplexes. These include inhibitors of type I restriction-modification enzymes (Ocr
and ArdA), inhibitors of DNA gyrase MfpA and QnrABS, etc. We describe here structural features
of these proteins and mechanisms responsible for their interaction with DNA-dependent enzymes
and then discuss perspectives of use of DNA-mimicking proteins in analysis of replication,
repair, recombination, mechanisms underlying resistance to antibiotics, and also fields of
applied biotechnology.

<>

<1>Zavilgelsky, G.B., Rastorguev, S.M.
<2>Antirestriction proteins ArdA and Ocr as efficient inhibitors of type I restriction-modification enzymes.
<3>Mol. Biol. (Mosk)
<4>43
<5>241-248
<6>2009
<7>Genes encoding antirestriction proteins are found in transmissble plasmids (ardABC) and
bacteriophage genomes (ocr, darA).
Antirestriction proteins inhibit type I restriction-modification
enzymes and thus protect the unmodified plasmid or phage DNA from
degradation. Antirestriction proteins belong to the family of
DNA-mimicry proteins, whose spatial structure mimics the B-form of DNA.
Based on an analysis of the mutant forms of ArdA and Ocr obtained by
site-directed mutagenesis and the native form of ArdA that specifically
inhibit type I restriction enzymes but do not affect their methylase
activity, a model is proposed to describe the complex formation between
an antirestriction protein and a type I restriction-modification enzyme
(R2M2S): antirestriction proteins can displace a DNA strand from its
binding sites in the S subunit (which contacts a specific site on DNA)
and in the R subunit (which translocates the DNA strand and cleaves
it). Antirestriction and antimodification activities of ArdA and Ocr as
a function of ardA and ocr expression levels were studied by cloning
the genes under a strictly regulated promoter.

<>

<1>Zdziarski, J., Brzuszkiewicz, E.B., Wullt, B., Liesegang, H., Biran, D., Voigt, B., Gronberg-Hernandez, J., Ragnarsdottir, B., Hecker, M., Ron, E.Z., Daniel, R., Gottschalk, G., Hacker, J., Svanborg, C., Dobrindt, U.
<2>Host imprints on bacterial genomes--Rapid, divergent evolution in individual patients.
<3>PLoS Pathog.
<4>6
<5>e1001078
<6>2010
<7>Bacteria lose or gain genetic material and through selection, new variants become fixed in the
population.  Here we provide the first, genome-wide example of a single bacterial strain's
evolution in different deliberately colonized patients and the surprising insight that hosts
appear to personalize their microflora.  By first obtaining the complete genome sequence of
the prototype asymptomatic bacteriuria strain E. coli 83972 and then resequencing its
descendants after therapeutic bladder colonization of different patients, we identified 34
mutations, which affected metabolic and virulence-related genes.  Further transcriptome and
proteome analysis proved that these genome changes altered bacterial gene expression resulting
in unique adaptation patterns in each patient.  Our results provide evidence that, in addition
to stochastic events, adaptive bacterial evolution is driven by individual host environments.
Ongoing loss of gene function supports the hypothesis that evolution towards commensalism
rather than virulence is favored during asymptomatic bladder colonization.

<>

<1>Zebala, J., Choi, J., Barany, F.
<2>Recognition of base-analogue modified substrates by the TaqI restriction endonuclease.
<3>FASEB J.
<4>6
<5>A358
<6>1992
<7>It has been proposed that protein-DNA recognition is mediated via specific
hydrogen bond, hydrophobic and/or electrostatic interactions made between the
protein and DNA surface.  We have energetically quantitated these interactions
for the TaqI endonuclease (cleaves T^CGA) by constructing base-analogue
substituted substrates.  The base analogues N6-methyl-A, N7-deaza-A,
N7-deaza-G, inosine, N4-methyl-C, 5-methyl-C, uracil, 5-bromoU, alpha-thio-A,
alpha-thio-G, alpha-thio-T and alpha-thio-A were substituted for their
corresponding unmodified counterpart in one strand of the TCGA duplex.  The
effects of these analogues were monitored by measuring the steady-state
(Km,kcat) and single-turnover (kst) kinetic constants.  Only the N6-methyl-A
substituted DNA, which recreates in vivo methylation, was unreactive while the
remaining analogue substitutions exhibited Michaelis-Menten kinetics.  In
general, the Km was either unchanged or lowered by the analogue substitutions.
In contrast, many of the analogues severely reduced kcat, suggesting the
modified functional groups served mainly to destabilize the transition state.
Single-turnover measurements paralleled the kcat results, pointing to the N7
and N6 of A, the N7 of G and one of the nonbridging oxygens 3' to T as putative
contacts made in achieving the transition state.  The unmodified strand in ten
out of twelve hemi-substituted substrates had a normal kst value suggesting
that a particular cleavage center is controlled predominantly by recognition of
determinants on the same strand as the scissile bond.

<>

<1>Zebala, J., Choi, J., Barany, F.
<2>Characterization of steady state, single-turnover, and binding kinetics of the TaqI restriction endonuclease.
<3>J. Biol. Chem.
<4>267
<5>8097-8105
<6>1992
<7>The TaqI restriction endonuclease recognizes and cleaves the duplex DNA sequence T|CGA. Steady
state kinetic analysis with a small oligodeoxyribonucleotide substrate showed that the enzyme
obeyed Michaelis-Menten kinetics (Km=53 nM, kcat=1.3 min-1 at 50C and Km=0.5 nM, kcat=2.9
min-1 at 60C). At 0C, the enzyme was completely inactive, while at 15C, turnover produced
nicked substrate as the major product in excess of enzyme indicating dissociation between
nicking events. Above 37C, both strands in the duplex were cleaved prior to dissociation. In
contrast to the tight, temperature-dependent binding of substrate, binding of the Mg2+
cofactor was weak (Kd=2.5 mM) and the same at either 50C or 60C. Single-turnover experiments
using oligonucleotide substrate showed that hydrolysis of duplex DNA occurred via two
independent nicking events, each with a first order rate constant (kst) of 5.8 min-1 at 60C
and 3.5 min-1 at 50C. The pH dependence of Km (pKa=9) and kst (pKa=7) suggests Lys/Arg and
His, respectively, as possible amino acids influencing these constants. Moreover, although kst
increased significantly with pH, kcat did not, indicating that at least two steps can be
rate-controlling in the reaction pathway. Binding of protein to canonical DNA in the presence
of Mg2+ at 0C or in the absence of Mg2+ at 50C was weak (Kd=2.5 microM or 5,000 fold weaker
than the optimal measured Km) and equal to the binding of noncanonical DNA as judged by
retention on nitrocellulose. Similar results were seen in gel retardation assays. These
results suggest that both Mg2+ and high temperature are required to attain the correct protein
conformation to form the tight complex seen in the steady state analysis. In the accompanying
paper (Zebala, J.A., Choi, J., Trainor, G.L., and Barany, F. (1992) J. Biol. Chem. 267,
8106-8116), we report how these kinetic constants are altered using substrate analogues and
propose a model of functional groups involved in TaqI endonuclease recognition.

<>

<1>Zebala, J.A.
<2>DNA recognition of base-analogue and chemically modified substrates by the TaqI restriction endonuclease.
<3>Diss. Abstr.
<4>54
<5>1394-B
<6>1993
<7>The TaqI restriction endonuclease recognizes and cleaves the duplex DNA sequence T^CGA. It has
been proposed that protein-DNA recognition is mediated via specific hydrogen bond, hydrophobic
and/or electrostatic interactions between the protein and DNA surfaces. We have attempted to
map and quantitate the energies of these interactions for the TaqI endonuclease by
constructing substrates substituted with base or phosphate analogues that either remove or
sterically obstruct particular functional groups in the canonical TCGA sequence. The DNA
backbone was also modified using a chemical approach (phosphate ethylation) which identified
several phosphates in the recognition sequence essential for cleavage. The base analogues;
N6-methyl-A, N7-deaza-A, N7-deaza-G, inosine, N4-methyl-C, 5-methylC, uracil, 5-bromoU and the
phosphate analogues; a-thio-A, a-thioG, a-thio-T, a-thio-A were substituted for their
corresponding unmodified counterpart in one strand of the TCGA duplex. The effects of these
analogues were monitored by measuring the steady-state (Km, kcat) and single-turnover (kst)
kinetic constants. Only the N6-methyl-A substituted DNA, which mimics in vivo methylation, was
unreactive while the remaining analogue substitutions exhibited Michaelis-Menten kinetics. In
general, the Km was either unchanged or lowered by the analogue substitutions. In contrast,
many of the analogues severely reduced Kcat, suggesting the modified functional groups served
mainly to destabilize the transition state. Single-turnover measurements paralleled the kcat
results, pointing to the N7 and N6 of A, the N7 of G and one of the nonbridging oxygens 3' to
T (scissile bond) as putative contacts made in achieving the transition state. Substrates with
double substitutions displayed simple additivity of delta,deltaG' implying that these changes
behaved independently. The unmodified strand in ten out of twelve hemi-substituted substrates
had a normal kst value suggesting that a particular cleavage center is controlled
predominantly by recognition of determinants on the same strand as the scissile bond. These
results are discussed in relation to base analogue work from the EcoRI, RsrI and EcoRV
restriction endonucleases.

<>

<1>Zebala, J.A., Choi, J., Trainor, G.L., Barany, F.
<2>DNA recognition of base analogue and chemically modified substrates by the TaqI restriction endonuclease.
<3>J. Biol. Chem.
<4>267
<5>8106-8116
<6>1992
<7>It has been proposed that protein-DNA recognition is mediated via specific hydrogen bond,
hydrophobic, and/or electrostatic interactions between the protein and DNA surfaces. We have
attempted to map and quantitate the energies of these interactions for the TaqI endonuclease
by constructing substrates substituted with base or phosphate analogues that either remove or
sterically obstruct particular functional groups in the canonical TCGA sequence. The DNA
backbone was also modified using a chemical approach (phosphate ethylation) which identified
several phosphates in the recognition sequence essential for cleavage. The base analogues,
N6-methyl-A, N7-deaza-A, N7-deaza-G, inosine, N4-methyl-C, 5-methyl-C, uracil, 5-bromo-U, and
the phosphate analogues, alpha-thio-A, alpha-thio-G, alpha-thio-T, alpha-thio-A, were
substituted for their corresponding unmodified counterpart in one strand of the TCGA duplex.
The effects of these analogues were monitored by measuring the steady state (Km5 kcat) and
single-turnover (kst) kinetic constants. Only the N6-methyl-A-substituted DNA, which mimics in
vivo methylation, was unreactive while the remaining analogue substitutions exhibited
Michaelis-Menten kinetics. In general, the Km was either unchanged or lowered by the analogue
substitutions. In contrast, many of the analogues severely reduced kcat, suggesting the
modified functional groups served mainly to destabilize the transition state. Single-turnover
measurements paralleled the kcat results, pointing to the N7 and N6 of A, the N7 of G, and one
of the nonbridging oxygens 3' to T as putative contacts made in achieving the transition
state. Substrates with double substitutions displayed simple additivity of DeltaDeltaG
implying that these changes behaved independently. The unmodified strand in 10 out of 12
hemisubstituted substrates had a normal kst value suggesting that a particular cleavage center
is controlled predominantly by recognition of determinants on the same strand as the scissile
bond. These results are discussed in relation to base analogue work from the EcoRI, RsrI, and
EcoRV restriction endonucleases.

<>

<1>Zedelius, J., Rabus, R., Grundmann, O., Werner, I., Brodkorb, D., Schreiber, F., Ehrenreich, P., Behrends, A., Wilkes, H., Kube, M., Reinhardt, R., Widdel, F.
<2>Alkane degradation under anoxic conditions by a nitrate-reducing bacterium with possible involvement of the electron acceptor in substrate activation.
<3>Environ. Microbiol. Reports
<4>3
<5>125-135
<6>2011
<7>Microorganisms can degrade saturated hydrocarbons (alkanes) not only under oxic but also under
anoxic conditions. Three denitrifying isolates (strains HxN1, OcN1, HdN1) able to grow under
anoxic conditions by coupling alkane oxidation to CO2 with NO3 - reduction to N2 were compared
with respect to their alkane metabolism. Strains HxN1 and OcN1, which are both
Betaproteobacteria, utilized n-alkanes from C6 to C8 and C8 to C12 respectively. Both activate
alkanes anaerobically in a fumarate-dependent reaction yielding alkylsuccinates, as suggested
by present and previous metabolite and gene analyses. However, strain HdN1 was unique in
several respects. It belongs to the Gammaproteobacteria and was more versatile towards
alkanes, utilizing the range from C6 to C30. Neither analysis of metabolites nor analysis of
genes in the complete genome sequence of strain HdN1 hinted at fumarate-dependent alkane
activation. Moreover, whereas strains HxN1 and OcN1 grew with alkanes and NO3 -, NO2 - or N2O
added to the medium, strain HdN1 oxidized alkanes only with NO3 - or NO2 - but not with added
N2O; but N2O was readily used for growth with long-chain alcohols or fatty acids. Results
suggest that NO2 - or a subsequently formed nitrogen compound other than N2O is needed for
alkane activation in strain HdN1. From an energetic point of view, nitrogen-oxygen species are
generally rather strong oxidants. They may enable enzymatic mechanisms that are not possible
under conditions of sulfate reduction or methanogenesis and thus allow a special mode of
alkane activation.

<>

<1>Zehr, E.S., Tabatabai, L.B.
<2>Detection of a bacteriophage gene encoding a Mu-like portal protein in Haemophilus parasuis reference strains and field isolates by nested polymerase chain reaction.
<3>J. Vet. Diagn. Invest.
<4>23
<5>538-542
<6>2011
<7>A nested polymerase chain reaction assay was developed to determine the presence of a gene
encoding a bacteriophage Mu-like portal protein, gp29, in 15 reference strains and 31 field
isolates of Haemophilus parasuis.  Specific primers, based on the gene's sequence, were
utilized.  A majority of the virulent reference strains and field isolates tested harbored the
gene.  The results suggest that the nPCR technique described in the current report could serve
as a tool for epidemiological studies of H. parasuis.

<>

<1>Zehr, E.S., Tabatabai, L.B., Bayles, D.O.
<2>Genomic and proteomic characterization of SuMu, a Mu-like bacteriophage infecting Haemophilus parasuis.
<3>BMC Genomics
<4>13
<5>331
<6>2012
<7>ABSTRACT: BACKGROUND: Haemophilus parasuis, the causative agent of Glasser's
disease, is prevalent in swine herds and clinical signs associated with this
disease are meningitis, polyserositis, polyarthritis, and bacterial pneumonia.
Six to eight week old pigs in segregated early weaning herds are particularly
susceptible to the disease. Insufficient colostral antibody at weaning or the
mixing of pigs with heterologous virulent H. parasuis strains from other farm
sources in the nursery or grower-finisher stage are considered to be factors for
the outbreak of Glasser's disease. Previously, a Mu-like bacteriophage portal
gene was detected in a virulent swine isolate of H. parasuis by nested polymerase
chain reaction. Mu-like bacteriophages are related phyologenetically to
enterobacteriophage Mu and are thought to carry virulence genes or to induce host
expression of virulence genes. This study characterizes the Mu-like
bacteriophage, named SuMu, isolated from a virulent H. parasuis isolate. RESULTS:
Characterization was done by genomic comparison to enterobacteriophage Mu and
proteomic identification of various homologs by mass spectrometry. This is the
first report of isolation and characterization of this bacteriophage from the
Myoviridae family, a double-stranded DNA bacteriophage with a contractile tail,
from a virulent field isolate of H. parasuis. The genome size of bacteriophage
SuMu was 37,151 bp. DNA sequencing revealed fifty five open reading frames,
including twenty five homologs to Mu-like bacteriophage proteins: Nlp, phage
transposase-C-terminal, COG2842, Gam-like protein, gp16, Mor, peptidoglycan
recognition protein, gp29, gp30, gpG, gp32, gp34, gp36, gp37, gpL, phage tail
tube protein, DNA circulation protein, gpP, gp45, gp46, gp47, COG3778, tail fiber
protein gp37-C terminal, tail fiber assembly protein, and Com. The last open
reading frame was homologous to IS1414. The G + C content of bacteriophage SuMu
was 41.87% while its H. parasuis host genome's G + C content was 39.93%. Twenty
protein homologs to bacteriophage proteins, including 15 structural proteins, one
lysogeny-related and one lysis-related protein, and three DNA replication
proteins were identified by mass spectrometry. One of the tail proteins, gp36,
may be a virulence-related protein. CONCLUSION: Bacteriophage SuMu was
characterized by genomic and proteomic methods and compared to
enterobacteriophage Mu.

<>

<1>Zehr, J.P., Bench, S.R., Carter, B.J., Hewson, I., Niazi, F., Shi, T., Tripp, H.J., Affourtit, J.P.
<2>Globally distributed uncultivated oceanic N2-fixing cyanobacteria lack oxygenic photosystem II.
<3>Science
<4>322
<5>1110-1112
<6>2008
<7>Biological nitrogen (N2) fixation is important in controlling biological
productivity and carbon flux in the oceans. Unicellular N2-fixing
cyanobacteria have only recently been discovered and are widely
distributed in tropical and subtropical seas. Metagenomic analysis of flow
cytometry-sorted cells shows that unicellular N2-fixing cyanobacteria in
"group A" (UCYN-A) lack genes for the oxygen-evolving photosystem II and
for carbon fixation, which has implications for oceanic carbon and
nitrogen cycling and raises questions regarding the evolution of
photosynthesis and N2 fixation on Earth.

<>

<1>Zelby, L.S.
<2>Antisense RNA and ribozymes targeting DNA methyltransferase for in vivo studies.
<3>Diss. Abstr.
<4>57
<5>907
<6>1996
<7>The aim of this study was to make antisense RNA and ribozyme constructs
directed against mouse DNA methyltransferase (Mtase) and to use inducible expression to
perturb in vivo DNA methylation, both in tissue culture cells and in transgenic mice.  The
sheep metallothionein Ia promoter, known to be inducible by zinc in tissue culture cells and
mouse somatic tissues, and constitutive in mouse gonadal tissues, was used to drive in
vivo expression.  Several antisense RNA and ribozyme constructs were made and all were
shown to be expressed in a fibroblast-derived A9 hybrid cell line.  The ribozyme was
shown to cleave its target RNA efficiently in vitro, but the A9 hybrid cells were found to be
not suitable for determination of effects of DNA methyltransferase on methylation levels.
Therefore, another cell line, L61-M, was characterized.  L61-M cells were found to provide
a very sensitive and quantitative assay for methylation perturbations by measuring
reactivation of a methylation-silenced thymidine kinase (TK) gene.  Transfection
experiments using L61-M cells showed that the ribozyme caused a ten-fold increase over
background of stable TK+ revertants, and that the ribozyme was more effective than the
antisense RNA constructs, which gave results not significantly different from background.
Though the ribozyme clearly caused an effect, mRNA for the antisense RNAs or the
ribozyme could not be detected in transfected L61-M cells, so proof of mechanism of action
could not be obtained.  The present working hypothesis is that the ribozyme is quite
effective and deserving of further study, but a different promoter or expression vector
needs to be used in order to take full advantage of the excellent assay system provided by
the L61-M cells.

<>

<1>Zelder, O., Pompejus, M., Schroeder, H., Kroeger, B., Klopprogge, C., Haberhauer, G.
<2>Genes coding for DNA replication proteins and for proteins related to pathogenesis.
<3>International Patent Office
<4>WO 03040289 A
<5>
<6>2003
<7>The invention concerns novel nucleic acid molecules, the use of said molecules for producing
by recombination genetically improved micro-organisms, and a method for preparing fine
chemical products, in particular amino acids, using said genetically improved micro-organisms.

<>

<1>Zelder, O., Pompejus, M., Schroeder, H., Kroeger, B., Klopprogge, C., Haberhauer, G.
<2>Genes coding for DNA replication and pathogenesis proteins.
<3>Korean Patent Office
<4>KR 1020047006784 A
<5>
<6>2004
<7>
<>

<1>Zelder, O., Schroeder, H., Haberhauer, G., Kroeger, B., Pompejus, M.
<2>CORYNEBACTERIUM GLUTAMICUM GENES ENCODING PROTEINS INVOLVED IN HOMEOSTASIS AND ADAPTATION.
<3>Japanese Patent Office
<4>JP 2004512808 A
<5>
<6>2004
<7>
<>

<1>Zelinkova, E., Paulicek, M., Zelinka, J.
<2>Modification methylase M. Sau3239I from Streptomyces aureofaciens 3239.
<3>FEBS Lett.
<4>271
<5>147-148
<6>1990
<7>By chromatography on phosphocellulose and Heparin-Sepharose the modification
methylase M.Sau3239I was detected and partially purified from cells of
Streptomyces aureofaciens 3239.  Methylation by this enzyme protects DNA from
cleavage by the restriction endonuclease R.Sau3239I.  The enzyme catalyzes
methylation of adenine to N-6-methyladenine in the 5'-CTGGmAG-3' recognition
sequence.

<>

<1>Zelinskaya, N.V., Kovalevskaya, N.P., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>A site-specific endonuclease and methylase from the Thermophilic strain of Bacillus species IS4.
<3>Biokhimiia
<4>60
<5>1435-1448
<6>1995
<7>A site-specific endonuclease (R.BspIS4I) and methylase (M.BspIS4I) have been isolated and
purified to functional purity from the thermophilic strain of Bacillus species IS4.  R.BspIS4I
recognizes the sequence
5'-GAAGAC2N^-3'
3'-CTTCTG6N^-5' and cleaves it as shown by the arrows to form single-stranded
tetranucleotide 5'-protruding termini.  The enzyme is an isoschizomer of endonuclease BbvII.
M.BspIS4I is attributed to the adenine-specific methylases.

<>

<1>Zelinskaya, N.V., Kovalevskaya, N.P., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>A new site-specific endonuclease from Acinetobacter species (Strain M).
<3>Biokhimiia
<4>61
<5>1471-1482
<6>1996
<7>The site-specific endonuclease R.AspMI was isolated from Acinetobacter species (strain M) and
purified to functional purity.  The enzyme recognizes the symmetrical DNA sequence
5'-AGG/CCT-3' and cleaves it as shown by the arrow, forming blunt DNA ends.  The enzyme is
an isoschizomer of endonuclease StuI.  Cleavage of the DNA site is inhibited by
dcm-methylation.  AspM is in solution as an ~30-kD monomer.

<>

<1>Zelinskaya, N.V., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>A novel site-specific endonuclease from Bacillus species ST5.
<3>Biokhimiia
<4>60
<5>1999-2010
<6>1995
<7>A site-specific endonuclease, R.BspST5I, has been isolated in a functionally pure state from
the thermophilic strain Bacillus species ST5.  The enzyme recognizes the sequence 5'-
GCATC-3' on DNA and cleaves it at a distance of five nucleotides from the 3'-end of the
recognition site and at distances of nine or ten nucleotides along the complementary strand,
depending on the nucleotide surrounding the site on the DNA.  The enzyme is an isomer of
endonuclease SfaNI from Streptococcus faecalis ND547.

<>

<1>Zelinskaya, N.V., Matvienko, N.N., Zheleznaya, L.A., Matvienko, N.I.
<2>A new site-specific adenine DNA-methyltransferase from the strain of Bacillus species ST5.
<3>Biokhimiia
<4>61
<5>1006-1014
<6>1996
<7>The site-specific DNA-methylase M.BspST5I has been isolated from Bacillus species ST5 and
purified to functional purity.  M.BspST5I protects DNA from endonuclease R.BspST5I which
recognizes the nonpalindromic sequence
5'-GCATC-3'
3'-CGTAG-5'
on DNA.  M.BspST5I is an adenine-specific methylase.

<>

<1>Zell, R., Fritz, H.-J.
<2>DNA mismatch-repair in Escherichia coli counteracting the hydrolytic deamination of 5-methyl-cytosine residues.
<3>EMBO J.
<4>6
<5>1809-1815
<6>1987
<7>Derivatives of phage M13 were constructed and used for the in vitro
preparation of heteroduplex DNA molecules containing base/base mismatches that mimick
DNA lesions caused by hydrolytic deamination of 5-meC residues in Escherichia coli DNA
(i.e. they carry a T/G mismatch in the special sequence context provided by the recognition
site -CCA/TGG-of the Dcm-methyltransferase).  Upon introduction of these heteroduplex
DNAs into CaCl2-treated E. coli cells, the mismatches are efficiently repaired with high
bias in favour of the DNA strand containing the mismatched guanine residue.  This special
DNA mismatch-repair operates on fully dam-methylated DNA and is independent of gene
mutH.  It thus fulfills the salient requirements of a repair pathway responsible for
counteracting the spontaneous hydrolytic deamination of 5-meC in vivo.  The repair
efficiency is boosted by a 5-methyl group present on the cytosine residue at the next-nearest
position to the 5' side of the mismatched guanine.  The repair is severly impaired in host
strains carrying a mutation in any of the three loci dcm, mutL and mutS.

<>

<1>Zeng, Q.L., Bonocora, R.P., Shub, D.A.
<2>A Free-Standing Homing Endonuclease Targets an Intron Insertion Site in the psbA Gene of Cyanophages.
<3>Curr. Biol.
<4>19
<5>218-222
<6>2009
<7>Homing endonuclease genes are mobile elements that promote their duplication into cognate
sites that lack the endonuclease gene [1, 2].
The homing endonuclease initiates this event through site-specific DNA
cleavage. Copying of the endonuclease gene follows as a consequence of
DNA repair. A genome containing a homing endonuclease gene is subject
to self-cleavage. Protection is accomplished through DNA sequence
polymorphisms, as is the case in intronless homing of free-standing
endonuclease genes [3, 4], or by disruption of the recognition site by
a group I intron (or intein) into which the endonuclease ORF is
embedded. We describe here a novel free-standing homing endonuclease
from cyanobacteriophage S-PM2, which is similar to the DNA resolvase of
bacteriophage T4 and is encoded adjacent to an intron-containing psbA
gene [5, 6]. The endonuclease makes a specific double-strand cut near
the intron insertion site (IIS), its DNA recognition site spans the
IIS, and it is unable to cleave intron-containing psbA genes. This
interdependence of a free-standing endonuclease gene and a group I
intron, which we denote "collaborative homing," has not been reported
previously and gives support to a hypothesis of formation of composite
mobile introns by independent convergence of an intron and an
endonuclease gene on the same target sequence.

<>

<1>Zeng, W., Chen, G., Tang, Z., Wu, H., Shu, L., Liang, Z.
<2>Draft Genome Sequence of Bacillus subtilis GXA-28, a Thermophilic Strain with High Productivity of Poly-gamma-Glutamic Acid.
<3>Genome Announcements
<4>2
<5>e01259-14
<6>2014
<7>Bacillus subtilis GXA-28 is a thermophilic strain that can produce high yield and high
molecular weight of poly-gamma-glutamic acid under high temperature. Here,
we report the draft genome sequence of this strain, which may provide the genomic
basis for the high productivity of poly-gamma-glutamic acid.

<>

<1>Zeng, Y., Feng, F., Liu, Y., Li, Y., Koblizek, M.
<2>Genome Sequences and Photosynthesis Gene Cluster Composition of a Freshwater Aerobic Anoxygenic Phototroph, Sandarakinorhabdus sp. Strain AAP62, Isolated from  the Shahu Lake in Ningxia, China.
<3>Genome Announcements
<4>1
<5>e00034-13
<6>2013
<7>We report the first genome sequence from the recently established alpha-4 proteobacterial
genus . The genome of the sp. strain AAP62 contains a
photosynthesis gene cluster carrying major genes for bacterial reaction centers.
The presence of genes related to aerobic respiratory electron transport confirms
the lifestyle of this organism as an aerobic anoxygenic photoheterotroph.

<>

<1>Zeng, Y., Kasalicky, V., Simek, K., Koblizek, M.
<2>Genome sequences of two freshwater betaproteobacterial isolates, limnohabitans species strains rim28 and rim47, indicate their capabilities as both  photoautotrophs and ammonia oxidizers.
<3>J. Bacteriol.
<4>194
<5>6302-6303
<6>2012
<7>Betaproteobacterial genus Limnohabitans represents an important part of freshwater
bacterioplankton. Here, we report genome sequences of two
Limnohabitans isolates, Rim28 and Rim47. They contain a complete photosynthesis
gene cluster, RuBisCO, CO dehydrogenase, ammonia monooxygenase, and
sulfur-oxidizing genes, which indicates a great metabolic versatility of the
Limnohabitans species.

<>

<1>Zeng, Y., Koblizek, M., Feng, F., Liu, Y., Wu, Z., Jian, J.
<2>Whole-Genome Sequences of an Aerobic Anoxygenic Phototroph, Blastomonas sp. Strain AAP53, Isolated from a Freshwater Desert Lake in Inner Mongolia, China.
<3>Genome Announcements
<4>1
<5>e0007113
<6>2013
<7>Blastomonas is a strictly aerobic bacteriochlorophyll a-producing genus within the alpha-4
Proteobacteria. Here we report the first genome sequence from this
genus. The draft genome of Blastomonas sp. strain AAP53 contains a split
photosynthesis gene cluster and two gene clusters encoding a flagellar system.
Genes for the autotrophic CO2 fixation pathway are absent.

<>

<1>Zeng, Z., Dai, S., Xie, Y., Tian, X., Li, J., Wang, X.
<2>Genome sequences of two pseudoalteromonas strains isolated from the South china sea.
<3>Genome Announcements
<4>2
<5>e00305-14
<6>2014
<7>Two Pseudoalteromonas strains, SCSIO 04301 and SCSIO 11900, were isolated from the South China
Sea, and both strains form biofilms. Here we present the draft genome sequences of these two
strains, which will aid the study of marine microbes that are adapted to marine sediments or
are associated with eukaryotic hosts.

<>

<1>Zepeda, V., Dassa, B., Borovok, I., Lamed, R., Bayer, E.A., Cate, J.H.
<2>Draft Genome Sequence of the Cellulolytic Bacterium Clostridium papyrosolvens C7  (ATCC 700395).
<3>Genome Announcements
<4>1
<5>e00698-13
<6>2013
<7>We report the draft genome sequence of the cellulose-degrading bacterium Clostridium
papyrosolvens C7, originally isolated from mud collected below a
freshwater pond in Massachusetts. This Gram-positive bacterium grows in a
mesophilic anaerobic environment with filter paper as the only carbon source, and
it has a simple cellulosome system with multiple carbohydrate-degrading enzymes.

<>

<1>Zerillo, M.M., Van Sluys, M.A., Camargo, L.E., Monteiro-Vitorello, C.B.
<2>Characterization of new IS elements and studies of their dispersion in two subspecies of Leifsonia xyli.
<3>BMC Microbiol.
<4>8
<5>127
<6>2008
<7>BACKGROUND: Leifsonia xyli is a xylem-inhabiting bacterial species
comprised of two subspecies: L. xyli subsp. xyli (Lxx) and L. xyli subsp.
cynodontis (Lxc). Lxx is the causal agent of ratoon stunting disease in
sugarcane commercial fields and Lxc colonizes the xylem of several grasses
causing either mild or no symptoms of disease. The completely sequenced
genome of Lxx provided insights into its biology and pathogenicity. Since
IS elements are largely reported as an important source of bacterial
genome diversification and nothing is known about their role in chromosome
architecture of L. xyli, a comparative analysis of Lxc and Lxx elements
was performed. RESULTS: Sample sequencing of Lxc genome and comparative
analysis with Lxx complete DNA sequence revealed a variable number of IS
transposable elements acting upon genomic diversity. A detailed
characterization of Lxc IS elements and a comparative review with IS
elements of Lxx are presented. Each genome showed a unique set of elements
although related to same IS families when considering features such as
similarity among transposases, inverted and direct repeats, and element
size. Most of the Lxc and Lxx IS families assigned were reported to
maintain transposition at low levels using translation regulatory
mechanisms, consistent with our in silico analysis. Some of the IS
elements were found associated with rearrangements and specific regions of
each genome. Differences were also found in the effect of IS elements upon
insertion, although none of the elements were preferentially associated
with gene disruption. A survey of transposases among genomes of
Actinobacteria showed no correlation between phylogenetic relatedness and
distribution of IS families. By using Southern hybridization, we suggested
that diversification of Lxc isolates is also mediated by insertion
sequences in probably recent events. CONCLUSION: Collectively our data
indicate that transposable elements are involved in genome diversification
of Lxc and Lxx. The IS elements were probably acquired after the
divergence of the two subspecies and are associated with genome
organization and gene contents. In addition to enhancing understanding of
IS element dynamics in general, these data will contribute to our ongoing
comparative analyses aimed at understanding the biological differences of
the Lxc and Lxx.

<>

<1>Zernov, Y.P., Lebedev, L.R., Babkin, I.V., Chizhikov, V.E.
<2>Determination of substrate specificity of restriction endonuclease Kpn378I.
<3>Bioorg. Khim.
<4>16
<5>603-604
<6>1990
<7>The recognition sequence and cleavage site, CCGC^GG, of a new restriction
endonuclease Kpn378I has been determined.

<>

<1>Zeytun, A. et al.
<2>Complete genome sequence of Hydrogenobacter thermophilus type strain (TK-6).
<3>Standards in Genomic Sciences
<4>4
<5>131-143
<6>2011
<7>Hydrogenobacter thermophilus Kawasumi et al. 1984 is the type species of the genus
Hydrogenobacter. H. thermophilus was the first obligate autotrophic
organism reported among aerobic hydrogen-oxidizing bacteria. Strain TK-6(T) is of
interest because of the unusually efficient hydrogen-oxidizing ability of this
strain, which results in a faster generation time compared to other autotrophs.
It is also able to grow anaerobically using nitrate as an electron acceptor when
molecular hydrogen is used as the energy source, and able to aerobically fix
CO(2)via the reductive tricarboxylic acid cycle. This is the fifth completed
genome sequence in the family Aquificaceae, and the second genome sequence
determined from a strain derived from the original isolate. Here we describe the
features of this organism, together with the complete genome sequence and
annotation. The 1,742,932 bp long genome with its 1,899 protein-coding and 49 RNA
genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

<>

<1>Zeytun, A., Malfatti, S.A., Vergez, L.M., Shin, M., Garcia, E., Chain, P.S.
<2>Complete Genome Sequence of Francisella philomiragia ATCC 25017.
<3>J. Bacteriol.
<4>194
<5>3266
<6>2012
<7>Francisella philomiragia is a saprophytic gammaproteobacterium found only occasionally in
immunocompromised individuals and is the nearest neighbor to the
causative agent of tularemia and category A select agent Francisella tularensis.
To shed insight into the key genetic differences and the evolution of these two
distinct lineages, we sequenced the first complete genome of F. philomiragia
strain ATCC 25017, which was isolated as a free-living microorganism from water
in Bear River Refuge, Utah.

<>

<1>Zhalnina, K.V., Dias, R., Leonard, M.T., Dorr-de-Quadros, P., Camargo, F.A., Drew, J.C., Farmerie, W.G., Daroub, S.H., Triplett, E.W.
<2>Genome Sequence of Candidatus Nitrososphaera evergladensis from Group I.1b Enriched from Everglades Soil Reveals Novel Genomic Features of the  Ammonia-Oxidizing Archaea.
<3>PLoS ONE
<4>9
<5>e101648
<6>2014
<7>The activity of ammonia-oxidizing archaea (AOA) leads to the loss of nitrogen from soil,
pollution of water sources and elevated emissions of greenhouse gas.
To date, eight AOA genomes are available in the public databases, seven are from
the group I.1a of the Thaumarchaeota and only one is from the group I.1b,
isolated from hot springs. Many soils are dominated by AOA from the group I.1b,
but the genomes of soil representatives of this group have not been sequenced and
functionally characterized. The lack of knowledge of metabolic pathways of soil
AOA presents a critical gap in understanding their role in biogeochemical cycles.
Here, we describe the first complete genome of soil archaeon Candidatus
Nitrososphaera evergladensis, which has been reconstructed from metagenomic
sequencing of a highly enriched culture obtained from an agricultural soil. The
AOA enrichment was sequenced with the high throughput next generation sequencing
platforms from Pacific Biosciences and Ion Torrent. The de novo assembly of
sequences resulted in one 2.95 Mb contig. Annotation of the reconstructed genome
revealed many similarities of the basic metabolism with the rest of sequenced
AOA. Ca. N. evergladensis belongs to the group I.1b and shares only 40% of
whole-genome homology with the closest sequenced relative Ca. N. gargensis.
Detailed analysis of the genome revealed coding sequences that were completely
absent from the group I.1a. These unique sequences code for proteins involved in
control of DNA integrity, transporters, two-component systems and versatile
CRISPR defense system. Notably, genomes from the group I.1b have more gene
duplications compared to the genomes from the group I.1a. We suggest that the
presence of these unique genes and gene duplications may be associated with the
environmental versatility of this group.

<>

<1>Zhan, B., Angen, O., Hedegaard, J., Bendixen, C., Panitz, F.
<2>Draft Genome Sequences of Actinobacillus pleuropneumoniae Serotypes 2 and 6.
<3>J. Bacteriol.
<4>192
<5>5846-5847
<6>2010
<7>Actinobacillus pleuropneumoniae is a bacterial pathogen that causes highly contagious
respiratory infection in pigs and has a serious impact on the
production economy and animal welfare. As clear differences in virulence
between serotypes have been observed, the genetic basis should be
investigated at the genomic level. Here, we present the draft genome
sequences of the A. pleuropneumoniae serotypes 2 (strain 4226) and 6
(strain Femo).

<>

<1>Zhan, Y., Yan, Y., Zhang, W., Yu, H., Chen, M., Lu, W., Ping, S., Peng, Z., Yuan, M., Zhou, Z., Elmerich, C., Lin, M.
<2>Genome Sequence of Acinetobacter calcoaceticus PHEA-2, Isolated from Industry Wastewater.
<3>J. Bacteriol.
<4>193
<5>2672-2673
<6>2011
<7>Genome analysis of Acinetobacter calcoaceticus PHEA-2 was undertaken because of the importance
of this bacterium for bioremediation of phenol
polluted waster, and because of the close phylogenetic relationship of
this species with the human pathogen A. baumannii. To our knowledge, this
is the first strain of A. calcoaceticus whose genome has been sequenced.

<>

<1>Zhang, A., Yang, M., Hu, P., Wu, J., Chen, B., Hua, Y., Yu, J., Chen, H., Xiao, J., Jin, M.
<2>Comparative Genomic Analysis of Streptococcus suis reveals significant genomic diversity among different serotypes.
<3>BMC Genomics
<4>12
<5>523
<6>2011
<7>ABSTRACT:  BACKGROUND: Streptococcus suis (S. suis) is a major swine
pathogen and an emerging zoonotic agent. Serotypes 1, 2, 3, 7, 9, 14 and
1/2 are the most prevalent serotypes of this pathogen. However, almost all
studies were carried out on serotype 2 strains. Therefore,
characterization of genomic features of other serotypes will be required
to better understand their virulence potential and phylogenetic
relationships among different serotypes. RESULTS: Four Chinese S. suis
strains belonging to serotypes 1, 7, 9 and 1/2 were sequenced using a
rapid, high-throughput approach. Based on the 13 corresponding serotype
strains, including 9 previously completed genomes of this bacterium, a
full comparative genomic analysis was performed. The results provide
evidence that (i) the pan-genome of this species is open and the size
increases with addition of new sequenced genomes, (ii) strains of
serotypes 1, 3, 7 and 9 are phylogenetically distinct from serotype 2
strains, but all serotype 2 strains, plus the serotype 1/2 and 14 strains,
are very closely related. (iii) all these strains, except for the serotype
1 strain, could harbor a recombinant site for a pathogenic island (89K)
mediated by conjugal transfer, and may have the ability to gain the 89K
sequence. CONCLUSIONS: There is significant genomic diversity among
different strains in S. suis, and the gain and loss of large amount of
genes are involved in shaping their genomes. This is indicated by (i)
pairwise gene content comparisons between every pair of these strains,
(ii) the open pan-genome of this species, (iii) the observed indels,
invertions and rearrangements in the collinearity analysis. Phylogenetic
relationships may be associated with serotype, as serotype 2 strains are
closely related and distinct from other serotypes like 1, 3, 7 and 9, but
more strains need to be sequenced to confirm this.

<>

<1>Zhang, B., Li, L., Chen, F., Guan, Y.
<2>Initial investigation on restriction modification system of Streptomyces ahygroscopicus wuzhouensis neosubsp.
<3>Weishengwuxue Zazhi
<4>28
<5>65-68
<6>2008
<7>Adopting mycelium embedment in situ method and high voltage pulse electrophorsis two bands of
plasmid DNA from Streptomyces ahygroscopicus wuzhouensis neosubsp.  11371 (SAWNS) were
obtained, and they were linear molecules proved by bidirectionary electrophoresis and named as
pSAL1 and pSAL2 according to molecular weights from large to small successively.  Furthermore,
an initial investigation on SAWNS' restriction modification system was carried out, a copy of
plasmid pIJ702 from Streptomyces lividus TK54 was used to transfer plasma of SAWN and could
not obtain any transformant, however it succeeded to transform plasma of SAWN U-3 and obtained
a transformant.

<>

<1>Zhang, B., Tao, T., Wilson, G.G., Blumenthal, R.M.
<2>The M.AluI DNA-(cytosine C5)-methyltransferase has an unusually large, partially dispensable, variable region.
<3>Nucleic Acids Res.
<4>21
<5>905-911
<6>1993
<7>The DNA methyltransferase of the AluI restriction-modification system, from Arthrobacter
luteus, converts cytosine to 5-methylcytosine in the sequence AGCT. The gene for this
methyltransferase, aluIM, was cloned into Escherichia coli and sequenced. A 525-codon open
reading frame was found, consistent with deletion evidence, and the deduced amino acid
sequence revealed all ten conserved regions common to 5-methylcytosine methyltransferases. The
aluIM sequence predicts a protein of Mr 59.0K, in agreement with the observed Mr, making
M.AluI the largest known methyltransferase from a type II restriction-modification system.
M.AluI also contains the largest known variable region of any monospecific DNA
methyltransfese, larger than that of most multispecific methyltransferases. In other DNA
methyltransferases the variable region has been implicated as the sequence-specific target
recognition domain. An in-frame deletion that removes a third of this putative
target-recognition region leaves the AluI methyltransferase still fully active.

<>

<1>Zhang, C., Pang, B., Zhou, Z., Wang, H., Zhou, H., Lu, X., Du, P., Zhang, L., Li, J., Cui, Z., Chen, C., Stokes, H.W., Kan, B.
<2>The purifying trend in the chromosomal integron in Vibrio cholerae strains during the seventh pandemic.
<3>Infect. Genet. Evol.
<4>26C
<5>241-249
<6>2014
<7>Chromosomal integron (CI) arrays in Vibrio spp. are generally large and display great
variation. Here we determined the sequence of CI array in a toxigenic O139 Vibriocholerae
strain and compared it with the arrays from the genome of different O1 biotypes available in
GenBank. Then PCR scanning was used to determine the CI array variations in 83 epidemic O139
strains and subsequently these variations were compared with that found in toxigenic O1 El Tor
strains in our previous work. Few differences were observed in the cohort of toxigenic O139
strains compared to the toxigenic O1 El Tor strains. On the basis of CI arrays, the toxigenic
O1 El Tor and O139 strains isolated concurrently in recent years appear to be more similar to
each other than to the O1 strains isolated in previous decades, suggesting a closer
evolutionary relationship between them.
Comparison of CI arrays in toxigenic O1 El Tor and O139 V. cholerae strains isolated between
1961 and 2009 revealed a purifying trend in the CI arrays in the chronological order during
the seventh pandemic.

<>

<1>Zhang, D., Chang, D., Zhang, X., Yu, Y., Guo, Y., Wang, J., Li, T., Xu, G., Dai, W., Liu, C.
<2>Genome Sequence of Escherichia coli Strain LCT-EC52, Which Acquired Changes in Antibiotic Resistance Properties after the Shenzhou-VIII Mission.
<3>Genome Announcements
<4>2
<5>e00081-14
<6>2014
<7>Escherichia coli is a ubiquitous opportunistic pathogen that colonizes the lower  intestines
of humans and causes several diseases, such as septicemia, pneumonia,
and urinary tract infections. Here, we present the draft genome sequence of E.
coli strain LCT-EC52, which originated from E. coli strain CGMCC 1.2385 and
acquired changes in antibiotic resistance following travel on the Shenzhou-VIII
spacecraft.

<>

<1>Zhang, G., Deng, A., Xu, Q., Liang, Y., Chen, N., Wen, T.
<2>Complete genome sequence of Bacillus amyloliquefaciens TA208, a strain for industrial production of guanosine and ribavirin.
<3>J. Bacteriol.
<4>193
<5>3142-3143
<6>2011
<7>Here, we report the complete genome sequence of Bacillus amyloliquefaciens TA208, a strain for
industrial production of guanosine, and synthesis of
ribavirin by assimilation of formamide. Comparison of its genome sequence
with those of the strains DSM7 and FZB42 revealed horizontal gene transfer
represented by unique prophages and restriction-modification systems and
indicated significant accumulation of guanosine.

<>

<1>Zhang, G., Wang, W., Deng, A., Sun, Z., Zhang, Y., Liang, Y., Che, Y., Wen, T.
<2>A Mimicking-of-DNA-Methylation-Patterns Pipeline for Overcoming the Restriction Barrier of Bacteria.
<3>PLoS Genet.
<4>8
<5>e1002987
<6>2012
<7>Genetic transformation of bacteria harboring multiple Restriction-Modification (R-M) systems
is often difficult using
conventional methods. Here, we describe a
mimicking-of-DNA-methylation-patterns (MoDMP) pipeline to address this
problem in three difficult-to-transform bacterial strains. Twenty-four
putative DNA methyltransferases (MTases) from these
difficult-to-transform strains were cloned and expressed in an
Escherichia coli strain lacking all of the known R-M systems and orphan
MTases. Thirteen of these MTases exhibited DNA modification activity in
Southwestern dot blot or Liquid Chromatography-Mass Spectrometry
(LC-MS) assays. The active MTase genes were assembled into three
operons using the Saccharomyces cerevisiae DNA assembler and were
co-expressed in the E. coli strain lacking known R-M systems and orphan
MTases. Thereafter, results from the dot blot and restriction enzyme
digestion assays indicated that the DNA methylation patterns of the
difficult-to-transform strains are mimicked in these E. coli hosts. The
transformation of the Gram-positive Bacillus amyloliquefaciens TA208
and B. cereus ATCC 10987 strains with the shuttle plasmids prepared
from MoDMP hosts showed increased efficiencies (up to four orders of
magnitude) compared to those using the plasmids prepared from the E.
coli strain lacking known R-M systems and orphan MTases or its parental
strain. Additionally, the gene coding for uracil
phosphoribosyltransferase (upp) was directly inactivated using
non-replicative plasmids prepared from the MoDMP host in B.
amyloliquefaciens TA208. Moreover, the Gram-negative chemoautotrophic
Nitrobacter hamburgensis strain X14 was transformed and expressed Green
Fluorescent Protein (GFP). Finally, the sequence specificities of
active MTases were identified by restriction enzyme digestion, making
the MoDMP system potentially useful for other strains. The
effectiveness of the MoDMP pipeline in different bacterial groups
suggests a universal potential. This pipeline could facilitate the
functional genomics of the strains that are difficult to transform.

<>

<1>Zhang, H., Zhang, S., Peng, Y., Li, Y., Chen, Z., Zheng, W., Xu, H., Yu, Z., Zheng, T.
<2>Draft Genome Sequence of the Anti-Algal Marine Actinomycete Streptomyces sp. JS01.
<3>Genome Announcements
<4>2
<5>e01261-14
<6>2014
<7>Streptomyces sp. JS01 is the producer of an anti-algal compound that shows inhibitory activity
against a harmful algal species Phaeocystis globosa and can
also produce a red pigment. Its genome sequence will allow for the
characterization of the anti-algal compound and the molecular mechanisms
underlying its beneficial properties.

<>

<1>Zhang, H., Zhou, W., Zhuang, Y., Liang, X., Liu, T.
<2>Draft Genome Sequence of Streptomyces bottropensis ATCC 25435, a Bottromycin-Producing Actinomycete.
<3>Genome Announcements
<4>1
<5>e0001913
<6>2013
<7>A series of bottromycin antibiotics have been isolated and identified from Streptomyces
bottropensis strain ATCC 25435. Here, a draft genome sequence of S.
bottropensis ATCC 25435 is presented. The genome carries an intact biosynthetic
gene cluster for bottromycin antibiotics, which provides insight into the
combinatorial biosynthesis of bottromycin antibiotics.

<>

<1>Zhang, J., Cao, G., Xu, X., Jin, H., Yang, X., Allard, M., Brown, E., Meng, J.
<2>Draft Genome Sequences of Three Salmonella enterica Serotype Agona Strains from China.
<3>Genome Announcements
<4>1
<5>e00203-12
<6>2013
<7>Salmonellosis has been one of the major contributors to the global public health  burden.
serotype Agona has ranked among the top 10 and top 20 most frequent
serotypes isolated from human sources in China and the United States,
respectively. We report draft genomes of three Agona strains from China.

<>

<1>Zhang, J., Cao, G., Xu, X., Jin, H., Zhang, Q., Chen, J., Yang, X., Pan, H., Zhang, X., Allard, M., Brown, E., Meng, J.
<2>Whole-Genome Sequences of Four Salmonella enterica Serotype Newport Strains from  Humans.
<3>Genome Announcements
<4>1
<5>e00213-13
<6>2013
<7>Salmonellosis contributes significantly to the public health burden globally. Salmonella
enterica serotype Newport is among Salmonella serotypes most
associated with food-borne illness in the United States and China. It was thought
to be polyphyletic and to contain different lineages. We report draft genomes of
four S. Newport strains isolated from humans in China.

<>

<1>Zhang, J.X., Lin, B.R., Shen, H.F., Pu, X.M.
<2>Genome Sequence of the Banana Pathogen Dickeya zeae Strain MS1, Which Causes Bacterial Soft Rot.
<3>Genome Announcements
<4>1
<5>e00317-13
<6>2013
<7>We report a draft genome sequence of Dickeya zeae strain MS1, which is the causative agent of
banana soft rot in China, and we show several of its specific
properties compared with those of other D. zeae strains. Genome sequencing
provides a tool for understanding the genomic determination of the pathogenicity
and phylogeny placement of this pathogen.

<>

<1>Zhang, K., McClure, J.A., Elsayed, S., Conly, J.M.
<2>Novel Staphylococcal Cassette Chromosome mec Type, Tentatively Designated Type VIII, Harboring Class A mec and Type 4 ccr Gene Complexes in a Canadian Epidemic Strain of Methicillin-Resistant Staphylococcus aureus.
<3>Antimicrob. Agents Chemother.
<4>53
<5>531-540
<6>2009
<7>Staphylococcal cassette chromosome mec (SCCmec) is a mobile genetic
element characterized by flanking terminal direct and, in most cases,
inverted repeat sequences, the mec and ccr gene complexes, and their
surrounding DNA regions. Unique combinations of the mec and ccr gene
complexes generate various SCCmec types. Six SCCmec types have been
reported to date. We describe here a novel SCCmec type identified in a
Canadian methicillin-resistant Staphylococcus aureus (MRSA) epidemic
strain. MRSA clinical isolates were screened for known SCCmec types by
multiplex and conventional PCR methods. Three phenotypically and
genotypically identical MRSA clinical isolates with a pulsotype identical
to CMRSA9 were identified locally and found to be nontypeable by available
SCCmec typing schemes. Complete sequencing of the SCCmec element revealed
a nucleotide fragment of 32,168 bp integrated at an identical chromosomal
integration site (attBscc) at the 3' end of the orfX gene. The nucleotide
sequences at the chromosome-SCCmec junction regions were typical of other
SCCmec types, but the element harbored a unique combination of class A mec
and type 4 ccr gene complexes. Sequence recombination analysis suggested
that this unique SCCmec type may be derived from homologous recombination
between the previously described SCC(RP62A) of S. epidermidis strain RP62A
and SCC composite island of S. epidermidis ATCC 12228, respectively, or
via recombination of other staphylococcal strains that carry the same or
similar mobile cassettes. We identified a previously undescribed type of
SCCmec from isolate C10682, tentatively designated type VIII, and we
provide compelling evidence supporting the ability of SCC elements to
transfer horizontally or undergo recombination to generate new SCCmec
types.

<>

<1>Zhang, L., Li, X., Zhang, F., Wang, G.
<2>Genomic analysis of Agrobacterium radiobacter DSM 30147(T) and emended description of A. radiobacter (Beijerinck and van Delden 1902) Conn 1942  (Approved Lists 1980) emend. Sawada et al. 1993.
<3>Standards in Genomic Sciences
<4>9
<5>574-584
<6>2014
<7>Agrobacterium radiobacter is the only known non-phytopathogenic species in Agrobacterium
genus. In this study, the whole-genome sequence of A. radiobacter
type strain DSM 30147(T) was described and compared to the other available
Agrobacterium genomes. This bacterium has a genome size of 7,122,065 bp
distributed in 612 contigs, including 6,834 protein-coding genes and 41 RNA
genes. It harbors a circular chromosome and a linear chromosome but not a
tumor-inducing (Ti) plasmid. To the best of our knowledge, this is the first
report of a genome from the A. radiobacter species. In addition, an emended
description of A. radiobacter is described. This study reveals information that
enhances the current understanding of its non-phytopathogenicity and its
phylogenetic position within Agrobacterium genus.

<>

<1>Zhang, L., Morrison, M., O, C.P., Evans, P., Rickard, C.M.
<2>Genome Sequence of Staphylococcus epidermidis Strain AU12-03, Isolated from an Intravascular Catheter.
<3>J. Bacteriol.
<4>194
<5>6639
<6>2012
<7>In recent years, Staphylococcus epidermidis has become a major nosocomial pathogen and the
most common cause of intravascular catheter-related bacteremia,
which can increase morbidity and mortality and significantly affect patient
recovery. We report a draft genome sequence of Staphylococcus epidermidis
AU12-03, isolated from an intravascular catheter tip.

<>

<1>Zhang, L., Morrison, M., O, C.P., Evans, P., Rickard, C.M.
<2>Genome Sequence of Stenotrophomonas maltophilia Strain AU12-09, Isolated from an  Intravascular Catheter.
<3>Genome Announcements
<4>1
<5>e00195-13
<6>2013
<7>Stenotrophomonas maltophilia is an opportunistic nosocomial pathogen that is characterized by
its high-level intrinsic resistance to a variety of antibiotics
and its ability to form biofilms. Here, we report the draft genome sequence of
Stenotrophomonas maltophilia AU12-09, isolated from an intravascular catheter
tip.

<>

<1>Zhang, L., Morrison, M., Rickard, C.M.
<2>Draft Genome Sequence of Ralstonia pickettii AU12-08, Isolated from an Intravascular Catheter in Australia.
<3>Genome Announcements
<4>2
<5>e00027-14
<6>2014
<7>Ralstonia pickettii is a nonfermenting Gram-negative bacillus that creates a significant
problem in clinical settings, as it is a widespread cause of
nosocomial infections. Here, we report the draft genome sequence of R. pickettii
AU12-08, isolated from an intravascular catheter tip.

<>

<1>Zhang, L., Si, M., Zhu, L., Li, C., Wei, Y., Shen, X.
<2>Draft Genome Sequence of Nafulsella turpanensis ZLM-10T, a Novel Member of the Family Flammeovirgaceae.
<3>Genome Announcements
<4>2
<5>e00263-14
<6>2014
<7>Nafulsella turpanensis ZLM-10(T) is a slightly halophilic, Gram-negative, rod-shaped, gliding,
pale-pink-pigmented bacterium in the family Flammeovirgaceae, and it shows resistance to
gentamicin, kanamycin, neomycin, and streptomycin. Here, we report the genome sequence of N.
turpanensis strain ZLM-10(T), which has a 4.8-Mb genome and a G+C content of 45.67%.

<>

<1>Zhang, L., Skurnik, M.
<2>Isolation of an R- M+ mutant of Yersinia enterocolitica serotype O:8 and its application in construction of rough mutants utilizing mini-Tn5 derivatives and lipopolysaccharide-specific phage.
<3>J. Bacteriol.
<4>176
<5>1756-1760
<6>1994
<7>A generally applicable procedure was used to isolate a spontaneous restriction-deficient
mutant of Yersinia enterocolitica serotype O:8. Transposition frequency in the mutant strain
8081-res was approximately 6.7 x 10-6 per recipient, while it was practically zero in the
wild-type strain 8081-c. Mobilization frequency into 8081-res was 10/5 times higher than that
into the wild-type strain. The mutant had lost the ability to express the YenI restriction
endonuclease activity present in serotype 0:8 strains. This allowed the construction of a
transposon library in 8081-res. Insertion mutants with transposons in the genes of the rfa
region were selected from this library.

<>

<1>Zhang, Lu., Lu, D.-Yu., Ma, W.-Yun., Li, Y.
<2>Age-related changes in the localization of DNA methyltransferases during meiotic maturation in mouse oocytes.
<3>Fertil. Steril.
<4>95
<5>1531-1534
<6>2011
<7>The effects of maternal aging on the localization of DNA methyltransferases were evaluated
during mouse oocyte maturation using fluorescence staining. And we conclude that maternal
aging affects the cytoplasmic-to-nuclear trafficking of DNA methyltransferases in mouse
oocytes during the time from germinal vesicle breakdown to metaphase I.

<>

<1>Zhang, M., He, L., Li, Q., Sun, H., Gu, Y., You, Y., Meng, F., Zhang, J.
<2>Genomic characterization of the Guillain-Barre syndrome-associated Campylobacter jejuni ICDCCJ07001 Isolate.
<3>PLoS ONE
<4>5
<5>e15060
<6>2010
<7>Campylobacter jejuni ICDCCJ07001 (HS:41, ST2993) was isolated from a Guillain-Barre syndrome
(GBS) patient during a 36-case GBS outbreak
triggered by C. jejuni infections in north China in 2007. Sequence
analysis revealed that the ICDCCJ07001 genome consisted of 1,664,840 base
pairs (bp) and one tetracycline resistance plasmid of 44,084 bp. The GC
content was 59.29% and 1,579 and 37 CDSs were identified on the chromosome
and plasmid, respectively. The ICDCCJ07001 genome was compared to C.
jejuni subsp. jejuni strains 81-176, 81116, NCTC11168, RM1221 and C.
jejuni subsp. doylei 269.97. The length and organization of ICDCCJ07001
was similar to that of NCTC11168, 81-176 and 81-116 except that CMLP1 had
a reverse orientation in strain ICDCCJ07001. Comparative genomic analyses
were also carried out between GBS-associated C. jejuni strains. Thirteen
common genes were present in four GBS-associated strains and 9 genes
mapped to the LOS cluster and the ICDCCJ07001_pTet (44 kb) plasmid was
mosaic in structure. Thirty-seven predicted CDS in ICDCCJ07001_pTet were
homologous to genes present in three virulence-associated plasmids in
Campylobacter: 81-176_pTet, pCC31 and 81-176_pVir. Comparative analysis of
virulence loci and virulence-associated genes indicated that the LOS
biosynthesis loci of ICDCCJ07001 belonged to type A, previously reported
to be associated with cases of GBS. The polysaccharide capsular
biosynthesis (CPS) loci and the flagella modification (FM) loci of
ICDCCJ07001 were similar to corresponding sequences of strain 260.94 of
similar serotype as strain ICDCCJ07001. Other virulence-associated genes
including cadF, peb1, jlpA, cdt and ciaB were conserved between the C.
jejuni strains examined.

<>

<1>Zhang, M., Ito, T., Li, S., Misawa, S., Kondo, S., Miida, T., Ohsaka, A., Hiramatsu, K.
<2>Analysis of Staphylococcal cassette chromosome mec in BD GeneOhm MRSA assay-negative strains.
<3>Antimicrob. Agents Chemother.
<4>57
<5>2890-2891
<6>2013
<7>The BD GeneOhm MRSA assay could identify methicillin-resistant Staphylococcus
aureus (MRSA) strains at a high ratio (97.8%). Analysis of 11 assay-negative MRSA
strains suggested that insertion of non-mec staphylococcal cassette chromosome
elements (SCCs) downstream of orfX, and carriage of SCCmecs with a left extremity
that cannot be detected by the kit, might lead to their being given an incorrect
negative status.

<>

<1>Zhang, M., Li, Q., He, L., Meng, F., Gu, Y., Zheng, M., Gong, Y., Wang, P., Ruan, F., Zhou, L., Wu, J., Chen, L., Fitzgerald, C., Zhang, J.
<2>Association study between an outbreak of Guillain-Barre syndrome in Jilin, China, and preceding Campylobacter jejuni infection.
<3>Foodborne Pathog. Dis.
<4>7
<5>913-919
<6>2010
<7>From June to July 2007, 36 cases of Guillain-Barre syndrome (GBS) occurred
in a township in north China. Serological study and bacteria culture were
performed to investigate the association between preceding Campylobacter
jejuni infection and this GBS outbreak. Anti-C. jejuni antibodies were
found in significantly higher numbers of GBS patients (IgM 84%, IgG 87.5%)
than in healthy inspection cases (IgM 33%, IgG 27%). IgG anti-GM1 was the
dominant anti-ganglioside antibody among the GBS patients. Seven C. jejuni
isolates (four from human stool and three from poultry specimens taken
from the patients' houses) were obtained. Serotyping and molecular
analysis were used to investigate the genetic relatedness among these C.
jejuni isolates. The four human isolates, collected from residents of the
same district, were indistinguishable by both pulsed-field gel
electrophoresis and multilocus sequence typing, suggesting these patients
had a common source of infection. A new sequence type, sequence type-2993,
was assigned to the human C. jejuni isolates, three of which belonged to
Penner serotype heat-stable (HS):41. Both serotype and molecular subtype
of the human C. jejuni isolates were different from those of isolates
obtained from poultry specimens. Our results suggest that the antecedent
C. jejuni infection triggered this GBS outbreak in China.

<>

<1>Zhang, M., Yang, X., Liu, H., Liu, X., Huang, Y., He, L., Gu, Y., Zhang, J.
<2>Genome Sequences of the Guillain-Barre Syndrome Outbreak-Associated Campylobacter jejuni Strains ICDCCJ07002 and ICDCCJ07004.
<3>Genome Announcements
<4>1
<5>e00256-13
<6>2013
<7>The first world-known and largest outbreak of 36 cases of Guillain-Barre syndrome caused by a
preceding Campylobacter jejuni infection was reported previously in
China. During the outbreak, Campylobacter jejuni strain ICDCCJ07002 was isolated
from a patient with persistent diarrhea for 21 days, and C. jejuni strain
ICDCCJ07004 was from a healthy carrier without any clinical symptoms at the same
time. Here, we report the draft genome sequence of strain ICDCCJ07002 (1,698,407
bp, with a G+C content of 30.45%) and the genome resequencing result of strain
ICDCCJ07004 (1,701,584 bp, with a G+C content of 30.51%), and we compared these
with the completed genome of C. jejuni strain ICDCCJ07001.

<>

<1>Zhang, P., Bao, Y., Higgins, L., Xu, S.Y.
<2>Rational design of a chimeric endonuclease targeted to NotI recognition site.
<3>Protein Eng. Des. Sel.
<4>20
<5>497-504
<6>2007
<7>A cleavage-deficient variant of NotI restriction endonuclease (GCGGCCGC) was isolated by
random mutagenesis of the notIR gene. The NotI variant
D160N was shown to bind DNA and protect plasmid DNA from EagI (CGGCCG) and
NotI digestions. The EDTA-resistant BmrI restriction endonuclease cleaves
DNA sequence ACTGGG N(5)/N(4). The N-terminal cleavage domain of BmrI
(residues 1-198) with non-specific nuclease activity was fused to the NotI
variant D160N with a short linker. The engineered chimeric endonuclease
(CH-endonuclease) recognizes NotI sites specifically in the presence of
high salt (100-150 mM NaCl) and divalent cations Mg(++) or Ca(++). In
contrast to wild-type NotI, which cuts within its recognition sequence,
BmrI198-NotI (D160N) cleaves DNA outside of NotI sites, resulting in
deletion of the NotI site and the adjacent sequences. The fusion of the
BmrI cleavage domain to cleavage-deficient variants of Type II restriction
enzymes to generate novel cleavage sites will provide useful tools for DNA
manipulation.

<>

<1>Zhang, P.H., Too, P.H.M., Samuelson, J.C., Chan, S.H., Vincze, T., Doucette, S., Backstrom, S., Potamousis, K.D., Schramm, T.M., Forrest, D., Schwartz, D.C., Xu, S.Y.
<2>Engineering BspQI nicking enzymes and application of N.BspQI in DNA labeling and production of single-strand DNA.
<3>Protein Expr. Purif.
<4>69
<5>226-234
<6>2010
<7>BspQI is a thermostable Type IIS restriction endonuclease (REase) with the recognition
sequence 5'GCTCTTC N1/N4 3'. Here we report the cloning
and expression of the bspQIR gene for the BspQI restriction enzyme in
Escherichia coli. Alanine scanning of the BspQI charged residues
identified a number of DNA nicking variants. After sampling
combinations of different amino acid substitutions, an Nt.BspQI triple
mutant (E172A/E248A/E255K) was constructed with predominantly
top-strand DNA nicking activity. Furthermore, a triple mutant of BspQI
(Nb.BspQI, N235A/K331A/R428A) was engineered to create a bottom-strand
nicking enzyme. In addition, we demonstrated the application of
Nt.BspQI in optical mapping of single DNA molecules. Nt or
Nb.BspQI-nicked dsDNA can be further digested by E. coli exonuclease
III to create ssDNA for downstream applications. BspQI contains two
potential catalytic sites: a top-strand catalytic site (Ct) with a
D-H-N-K motif found in the HNH endonuclease family and a bottom-strand
catalytic site (Cb) with three scattered Glu residues. BlastP analysis
of proteins in GenBank indicated a putative restriction enzyme with
significant amino acid sequence identity to BspQI from the sequenced
bacterial genome Croceibacter atlanticus HTCC2559. This restriction
gene was amplified by PCR and cloned into a T7 expression vector.
Restriction mapping and run-off DNA sequencing of digested products
from the partially purified enzyme indicated that it is an Earl
isoschizomer with 6-bp recognition, which we named CatHI (CTCTTC N1/N4).

<>

<1>Zhang, Q., Xie, J., Wu, J., Lu, Z.
<2>RESP: a tool using for selecting a set of restriction enzymes.
<3>Prog. Post-Genome Technol., Southeast University Press, Nanjing, China, Zhou, G., Lu, Z., Takeyama, H., 
<4>0
<5>388-390
<6>2006
<7>The present restriction enzyme analysis tools, such as REBASE tool, REBsites etc., can provide
the functions of finding available enzymes to the given DNA sequence and supplying theoretical
digests.  In this paper, we report a novel restriction enzyme selection platform (RESP), which
can analyze nad optimize the length range and DNA fragment number after cleaving a given whole
genome by selecting a set of restriction enzymes.  The program runs under Microsoft Windows.
Compared to present programs, RESP has more excellent functions: it can give DNA digest
results with multiple enzymes, and simulate electrophoresis process for the digest results,
and accept the input of more large DNA sequences.  RESP has successfully been used to
demonstrate the selection of the enzyme set and their fragmentation process for the genome of
phage T7.

<>

<1>Zhang, Q., Ye, X., Ren, G., Zhang, N., Li, Lu., Li, D.
<2>A new method for the purification of restriction enzyme NotI.
<3>Zhongguo Shengwu Gongcheng Zazhi
<4>31
<5>102-109
<6>2011
<7>In order to study its specificities, the NotIR gene was cloned from Nocardia
otitidis-caviarum.  First, the genome DNA of Enterobacter agglomerans was extracted as
template, obtained EagI methylase gene by PCR and connected EagIM gene to pBR322 vector of
gain recombinant expression plasmid pBR322-EagIM.  Then transformed this plasmid ito E. coli
2555.  Secondly, extracted the genome DNA from Nocardia otitidis-caviarum as template and
obtained the restriction enzyme NHotIR gene by PCR.  After ligating the NotIR gene to
pACYC184-PT7, the pACYC184-PT7-NotIR plasmid was transformed into the ER2566 which was
protected through the methylation by pBR322-EagIM recombinant plasmid.  The engineered strain
ER2566 [pBR322-EagIM, pACYC184-PT7,-NotIR] could be induced to express restriction enzymes
NotI by IPTG and the induction conditions were optimized to make its expression mostly in
soluble form.  The enzyme was purified by AKTA purifier 100 protein purification system.
Through DEAE Sephrose FF, phenyl HP and Superdex 75 10/300 GL molecular sieve chromatography,
the NotI enzyme was purified 35-fold, the yield was 9.8 x 106 units/g wet cell which was up to
17.8% of the crude enzyme and the specific activity of the purified NotI was 1.37 x 106U/mg.
Digestion results showed that the enzyme was purified to homogeneity and was free of
detectable contamination by other DNase (exo and endo).  After optimization of the expression
and purification conditions, the yield and efficienty of NotI enzyme were greatly improved in
comparison with that previously reported.

<>

<1>Zhang, Q.-M., Sugiyama, H., Miyabe, I., Matsuda, S., Kino, K., Saito, I., Yonei, S.
<2>Replication in vitro and cleavage by restriction endonuclease of 5-formyluracil- and 5-hydroxymethyluracil-containing oligonucleotides.
<3>Int. J. Radiat. Biol.
<4>75
<5>59-65
<6>1999
<7>Purpose: To investigate the biological consequences of 5-formyluracil (5-foU) and
5-hydroxymethyluracil (5-hmU).  The authors constructed 22-mer oligonucleotides containing a
5-foU or 5-hmU residue at the same sites.  The effects of such modifications on the ability to
serve as a template for DNA polymerase and on the cleavage by sequence-specific restriction
endonuclease were examined.  The Klenow fragment of DNA polymerase I and Thermus thermophilus
DNA polymerase read through the sites of 5-foU and 5-hmU in the templates.  5-FoU directed the
incorporation of dCMP in addition to dAMP opposite the lesion during DNA synthesis.  The DNA
polymerases incorporated only dAMP opposite the 5-hmU.  The substitution of thymine by 5-foU
within the recognition site of the restriction endonucleases HincII and SalI inhibited or
prevented the cleavage by the enzymes, whereas the enzymes cleaved the 5-hmU-containing
oligonucleotides at the same rate as the T-containing oligonucleotides.  These results
indicated that the 5-foU-A base pair is less stable than the T-A base pair and that 5-foU-A
base pair is less stable than the T-A base pair and that 5-foU can form a base pair with C in
addition to A.  It was also demonstrated that the oxidation of thymine to 5-hmU does not
result in substantial deterioration.

<>

<1>Zhang, Q.Y., Xiao, F., Xie, J., Li, Z.Q., Gui, J.F.
<2>Complete Genome Sequence of Lymphocystis Disease Virus Isolated from China.
<3>J. Virol.
<4>78
<5>6982-6994
<6>2004
<7>Lymphocystis diseases in fish throughout the world have been extensively
described. Here we report the complete genome sequence of lymphocystis
disease virus isolated in China (LCDV-C), an LCDV isolated from cultured
flounder (Paralichthys olivaceus) with lymphocystis disease in China. The
LCDV-C genome is 186,250 bp, with a base composition of 27.25% G+C.
Computer-assisted analysis revealed 240 potential open reading frames
(ORFs) and 176 nonoverlapping putative viral genes, which encode
polypeptides ranging from 40 to 1,193 amino acids. The percent coding
density is 67%, and the average length of each ORF is 702 bp. A search of
the GenBank database using the 176 individual putative genes revealed 103
homologues to the corresponding ORFs of LCDV-1 and 73 potential genes that
were not found in LCDV-1 and other iridoviruses. Among the 73 genes, there
are 8 genes that contain conserved domains of cellular genes and 65 novel
genes that do not show any significant homology with the sequences in
public databases. Although a certain extent of similarity between putative
gene products of LCDV-C and corresponding proteins of LCDV-1 was revealed,
no colinearity was detected when their ORF arrangements and coding
strategies were compared to each other, suggesting that a high degree of
genetic rearrangements between them has occurred. And a large number of
tandem and overlapping repeated sequences were observed in the LCDV-C
genome. The deduced amino acid sequence of the major capsid protein (MCP)
presents the highest identity to those of LCDV-1 and other iridoviruses
among the LCDV-C gene products. Furthermore, a phylogenetic tree was
constructed based on the multiple alignments of nine MCP amino acid
sequences. Interestingly, LCDV-C and LCDV-1 were clustered together, but
their amino acid identity is much less than that in other clusters. The
unexpected levels of divergence between their genomes in size, gene
organization, and gene product identity suggest that LCDV-C and LCDV-1
shouldn't belong to a same species and that LCDV-C should be considered a
species different from LCDV-1.

<>

<1>Zhang, R., Xia, H., Xu, Q., Dang, F., Qin, Z.
<2>Recombinational cloning of the antibiotic biosynthetic gene clusters in linear plasmid SCP1 of Streptomyces coelicolor A3(2).
<3>FEMS Microbiol. Lett.
<4>345
<5>39-48
<6>2013
<7>The model organism Streptomyces coelicolor A3(2) harbors a 356-kb linear plasmid,
SCP1. We report here development of a recombinational cloning method for deleting
large segment from one telomere of SCP1 followed by replacing with the telomere
of pSLA2 and sequentially inserting with the overlapping cosmids in vivo. The
procedure depends on homologous recombination coupled with cleavage at telomere
termini by telomere terminal protein. Using this procedure, we cloned the 81-kb
avermectin and the 76-kb spinosad biosynthetic gene clusters into SCP1.
Heterologous expression of avermectin production in S. coelicolor was detected.
These results demonstrate the utility of SCP1 for cloning large DNA segments such
as antibiotic biosynthetic gene clusters.

<>

<1>Zhang, S., Flores-Cruz, Z., Kumar, D., Chakrabarty, P., Hopkins, D.L., Gabriel, D.W.
<2>The Xylella fastidiosa Biocontrol Strain EB92-1 Genome Is Very Similar and Syntenic to Pierce's Disease Strains.
<3>J. Bacteriol.
<4>193
<5>5576-5577
<6>2011
<7>Xylella fastidiosa infects a wide range of plant hosts and causes economically serious
diseases, including Pierce's disease (PD) of
grapevines. X. fastidiosa biocontrol strain EB92-1 is infectious to
grapevines but does not cause symptoms. The draft genome of EB92-1 reveals
that it may be missing 10 potential pathogenicity effectors.

<>

<1>Zhang, S., Luo, X., Cheng, J., Peng, J., Zhang, D., Liu, Y.
<2>Genome Sequence of Pyrethroid-Degrading Bacterium Rhodopseudomonas palustris Strain JSC-3b.
<3>Genome Announcements
<4>2
<5>e01228-13
<6>2014
<7>Rhodopseudomonas palustris strain JSC-3b is a facultative, thermophilic bacterium, which was
isolated from water in a canal adjacent to a vegetable
field. Strain JSC-3b biodegrades several varieties of pyrethroid residues
effectively through cometabolic pathways. Here, we present the genome sequence of
this biodegrader.

<>

<1>Zhang, S., Xu, Y., Zhou, Z., Wang, S., Yang, R., Wang, J., Wang, L.
<2>Complete genome sequence of Bordetella pertussis CS, a Chinese pertussis vaccine strain.
<3>J. Bacteriol.
<4>193
<5>4017-4018
<6>2011
<7>Bordetella pertussis is the causative agent of pertussis. Here, we report the genome sequence
of Bordetella pertussis strain CS, isolated from an
infant patient in Beijing and widely used as a vaccine strain for
production of acellular pertussis vaccine in China.

<>

<1>Zhang, S.D., Barbe, V., Garel, M., Zhang, W.J., Chen, H., Santini, C.L., Murat, D., Jing, H., Zhao, Y., Lajus, A., Martini, S., Pradel, N., Tamburini, C., Wu, L.F.
<2>Genome Sequence of Luminous Piezophile Photobacterium phosphoreum ANT-2200.
<3>Genome Announcements
<4>2
<5>e00096-14
<6>2014
<7>Bacteria of the genus Photobacterium thrive worldwide in oceans and show substantially varied
lifestyles, including free-living, commensal, pathogenic, symbiotic, and piezophilic. Here, we
present the genome sequence of a luminous, piezophilic Photobacterium phosphoreum strain,
ANT-2200, isolated from a water column at 2,200 m depth in the Mediterranean Sea. It is the
first genomic sequence of the P. phosphoreum group. An analysis of the sequence provides
insight into the adaptation of bacteria to the deep-sea habitat.

<>

<1>Zhang, T., Bao, M., Wang, Y., Su, H., Tan, T.
<2>Genome Sequence of Bacillus cereus Strain A1, an Efficient Starch-Utilizing Producer of Hydrogen.
<3>Genome Announcements
<4>2
<5>e00494-14
<6>2014
<7>Bacillus cereus strain A1 is a newly isolated hydrogen producer capable of utilizing
bioresources and biowaste, such as starch and starch wastewater. Here,
we present a 5.67-Mb assembly of the genome sequence of strain A1, which may
provide insights into the molecular mechanism of hydrogen production from
bioresources and biowaste.

<>

<1>Zhang, W., Yu, D., Sun, Z., Wu, R., Chen, X., Chen, W., Meng, H., Hu, S., Zhang, H.
<2>Complete genome sequence of Lactobacillus casei Zhang, a new probiotic strain isolated from traditional homemade koumiss in Inner Mongolia,  China.
<3>J. Bacteriol.
<4>192
<5>5268-5269
<6>2010
<7>Lactobacillus casei Zhang is a new probiotic bacterium isolated from koumiss collected in
Inner Mongolia, China. Here, we report the main
genome features of L. casei Zhang and the identification of several
predicted proteins implicated in interactions with the host.

<>

<1>Zhang, X., Bruice, T.C.
<2>The mechanism of M.HhaI DNA C5 cytosine methyltransferase enzyme: a quantum mechanics/molecular mechanics approach.
<3>Proc. Natl. Acad. Sci. U. S. A.
<4>103
<5>6148-6153
<6>2006
<7>The mechanism of DNA cytosine-5-methylation catalyzed by the bacterial M.HhaI enzyme has been
considered as a stepwise nucleophilic addition of Cys-81-S- to cytosine C6 followed by C5
nucleophilic replacement of the methyl of S-adenosyl-L-methionine to produce
5-methyl-6-Cys-81-S-5,6-dihydrocytosine. In this study, we show that the reaction is concerted
from a series of energy calculations by using the quantum mechanical/molecular mechanical
hybrid method. Deprotonation of 5-methyl-6-Cys-81-S-5,6-dihydrocytosine and expulsion of
Cys-81-S- provides the product DNA 5-methylcytosine. A required base catalyst for this
deprotonation is not available as a member of the active site structure. A water channel
between the active site and bulk water allows entrance of solvent to the active site.
Hydroxide at 10(-7) mole fraction (pH = 7) is shown to be sufficient for the required
catalysis. We also show that Glu-119-CO2H can divert the reaction by protonating cytosine N3
when Cys-81-S- attacks cytosine, to form the 6-Cys-81-S-3-hydrocytosine. The reactants and
6-Cys-81-S-3-hydrocytosine product are in rapid equilibrium, and this explains the observed
hydrogen exchange of cytosine with solvent.

<>

<1>Zhang, X., Hu, Q., Yuan, W., Shang, W., Cheng, H., Yuan, J., Zhu, J., Hu, Z., Li, S., Chen, W., Hu, X., Rao, X.
<2>First report of a sequence type 239 vancomycin-intermediate Staphylococcus aureus isolate in Mainland China.
<3>Diagn. Microbiol. Infect. Dis.
<4>77
<5>64-68
<6>2013
<7>Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen that
causes a wide range of both hospital- and community-acquired infections. The high
prevalence of MRSA and the extensive use of vancomycin in Mainland China may lead
to the emergence of vancomycin-intermediate S. aureus (VISA) isolates. In this
case, we report a VISA isolate from a 34-year-old male patient with steam burn.
The isolate was determined to be sequence type 239 staphylococcal cassette
chromosome mec type III, the most prevalent MRSA clone in Mainland China.

<>

<1>Zhang, X., Jacobsen, S.E.
<2>Genetic analyses of DNA methyltransferases in Arabidopsis thaliana.
<3>Cold Spring Harb. Symp. Quant. Biol.
<4>71
<5>439-447
<6>2006
<7>DNA methylation is a conserved epigenetic silencing mechanism that functions to suppress the
proliferation of transposons and regulate the
expression of endogenous genes. In plants, Mutations that cause severe
loss of DNA methylation result in reactivation of transposons as well
as developmental abnormalities. We use the flowering plant Arabidopsis
thaliana as a model system to study the establishment and maintenance
of DNA methylation as well as its role in regulating plant development.
The genetic evidence presented here suggests that methylation at CG and
non-CG sites function!; in a partially redundant and locus-specific
manner to regulate a wide range of developmental processes. Results
from recent studies also suggested that the dynamic nature of non-CG
methylation, which is critically important for its regulatory function,
is largely due to it, complicated interactions with other epigenetic
pathways such as RNAi and historic modifications. Finally, the use of
genomic approaches has significantly broadened our Understanding of the
patterning of DNA methylation on a genomewide sea e and has led to the
identification of hundreds of candidate genes that are controlled by
DNA methylation.

<>

<1>Zhang, X., Mathews, C.K.
<2>Effect of DNA cytosine methylation upon deamination-induced mutagenesis in a natural target sequence in duplex DNA.
<3>J. Biol. Chem.
<4>269
<5>7066-7069
<6>1994
<7>Are 5-methylcytosine residues in DNA hot spots for transition mutagenesis? Numerous studies
identify 1) structural changes induced by DNA methylation, 2) high percentages of human
mutations that result from GC to AT transition pathways, and 3) differences between G-C and
G-mC base pairs in susceptibility to nonenzymatic deamination. However, investigations of
chemical stability necessarily involve non-physiological conditions for chemical analysis of
deamination. Here we describe an experiment that compares rates of deamination-induced
mutagenesis between a G-C and G-mC base pair, when both are present in duplex DNA, incubated
at 37oC and pH 7.4, within identical sequence contexts, in a natural mutational target (the
Escherichia coli lacZ-alpha gene) that selects for mutagenesis at the specific site under
investigation. Under these conditions the rate of spontaneous deamination at G-mC exceeds that
at G-C by more than 21-fold. Our data implicate differences in chemical stability toward
deamination as a major causal factor relating DNA cytosine methylation to spontaneous
mutagenesis.

<>

<1>Zhang, X., Verdine, G.L.
<2>Mammalian DNA cytosine-5 methyltransferase interacts with p23 protein.
<3>FEBS Lett.
<4>392
<5>179-183
<6>1996
<7>In higher eukaryotic genomes, methylated cytosine residues (m5C) are distributed in heritable,
cell-type-specific patterns, which are believed to be involved in the control of gene
expression, developmental regulation and genomic imprinting.  These methylation patterns are
established and maintained by DNA cytosine-5 methyltransferase (Mtase), a ~1500 amino acid
enzyme containing a regulatory N-terminal domain and a catalytic C-terminal domain.  The
mechanism responsible for targeting Mtase to particular genes is poorly understood and might
possibly involve interactions with other proteins.  In an effort to identify proteins that
interact with the mammalian Mtase, we used the yeast two-hybrid system with several different
Mtase domains as baits.  Here we report an interaction between the C-terminal catalytic domain
of the Mtase and p23, a protein previously reported to associate with the progesterone
receptor (PR) complex.

<>

<1>Zhang, X., Wang, T., Liu, W., Guo, Y., Wang, J., Li, T., Fang, X., Zhang, X., Dai, W., Liu, C.
<2>Genome Sequence of Klebsiella pneumoniae Strain LCT-KP182, Which Acquired Hemolytic Properties after Space Flight.
<3>Genome Announcements
<4>2
<5>e01088-13
<6>2014
<7>The Klebsiella pneumoniae strain LCT-KP182 acquired hemolytic properties after space flight.
Here, we present the draft genome sequence of this strain.

<>

<1>Zhang, X., Wang, T., Su, L., Zhou, L., Li, T., Wang, J., Liu, Y., Jiang, X., Wu, C., Liu, C.
<2>Draft Genome Sequence of Bacillus cereus LCT-BC25, Isolated from Space Flight.
<3>Genome Announcements
<4>2
<5>e00667-13
<6>2014
<7>Bacillus cereus strain LCT-BC25, which was carried by the Shenzhou VIII spacecraft, traveled
in space for about 398 h. To investigate the response of B.
cereus to space environments, we determined the genome sequence of B. cereus
strain LCT-BC25, which was isolated after space flight.

<>

<1>Zhang, X., Xu, X., Yuan, W., Hu, Q., Shang, W., Hu, X., Tong, Y., Rao, X.
<2>Complete Genome Sequence of Staphylococcus aureus XN108, an ST239-MRSA-SCCmec III Strain with Intermediate Vancomycin Resistance Isolated in Mainland China.
<3>Genome Announcements
<4>2
<5>e00449-14
<6>2014
<7>ST239-MRSA-SCCmec III (ST239, sequence type 239; MRSA, methicillin-resistant Staphylococcus
aureus; SCCmec III, staphylococcal cassette chromosome mec type
III) is the most predominant clone of hospital-acquired methicillin-resistant S.
aureus in mainland China. We report here the complete genome sequence of XN108,
the first vancomycin-intermediate S. aureus strain isolated from a steam-burned
patient with a wound infection.

<>

<1>Zhang, X.S., Blaser, M.J.
<2>Natural Transformation of an Engineered Helicobacter pylori Strain Deficient in Type II Restriction Endonucleases.
<3>J. Bacteriol.
<4>194
<5>3407-3416
<6>2012
<7>Restriction-modification (RM) systems are important for bacteria to limit foreign DNA
invasion. The naturally competent bacterium Helicobacter pylori has highly
diverse strain-specific type II systems. To evaluate the roles of strain-specific
restriction in H. pylori natural transformation, a markerless type II restriction
endonuclease-deficient (REd) mutant was constructed. We deleted the genes
encoding all four active type II restriction endonucleases in H. pylori strain
26695 using sacB-mediated counterselection. Transformation by donor DNA with
exogenous cassettes methylated by Escherichia coli was substantially (1.7 and 2.0
log(10) for cat and aphA, respectively) increased in the REd strain. There also
was significantly increased transformation of the REd strain by donor DNA from
other H. pylori strains, to an extent corresponding to their shared type II R-M
system strain specificity with 26695. Comparison of the REd and wild-type strains
indicates that restriction did not affect the length of DNA fragment integration
during natural transformation. There also were no differentials in cell growth or
susceptibility to DNA damage. In total, the data indicate that the type II REd
mutant has enhanced competence with no loss of growth or repair facility compared
to the wild type, facilitating H. pylori mutant construction and other genetic
engineering.

<>

<1>Zhang, X.Y., Xie, B.B., Qin, Q.L., Liu, A., Chen, X.L., Zhou, B.C., Zhang, Y.Z.
<2>Draft genome sequence of strain p7-3-5, a new flavobacteriaceae bacterium isolated from intertidal sand.
<3>J. Bacteriol.
<4>194
<5>6632
<6>2012
<7>The Flavobacteriaceae bacterium strain P7-3-5 was isolated from intertidal sand of the Yellow
Sea, China. Analysis of the 16S rRNA gene sequences showed that
strain P7-3-5 formed a distinct phylogenetic lineage within the family
Flavobacteriaceae. The genome of strain P7-3-5 was sequenced to facilitate the
physiological, ecological, and evolutionary studies of the bacteria within the
family Flavobacteriaceae.

<>

<1>Zhang, X.Y., Zhang, Y.J., Qin, Q.L., Xie, B.B., Chen, X.L., Zhou, B.C., Zhang, Y.Z.
<2>Genome Sequence of the Protease-Producing Bacterium Rheinheimera nanhaiensis E407-8T, Isolated from Deep-Sea Sediment of the South China Sea.
<3>J. Bacteriol.
<4>194
<5>7001-7002
<6>2012
<7>The protease-producing bacterium E407-8(T) was isolated from deep-sea sediment of the South
China Sea and has been identified recently as representing a new
species, Rheinheimera nanhaiensis. The draft genome of R. nanhaiensis E407-8(T)
consists of 3,987,205 bp and contains 3,730 predicated protein-coding genes,
including 82 extracellular peptidase genes.

<>

<1>Zhang, Y., Chen, C., Liu, J., Deng, H., Pan, A., Zhang, L., Zhao, X., Huang, M., Lu, B., Dong, H., Du, P., Chen, W., Wan, K.
<2>Complete Genome Sequences of Mycobacterium tuberculosis Strains CCDC5079 and CCDC5080, Which Belong to the Beijing Family.
<3>J. Bacteriol.
<4>193
<5>5591-5592
<6>2011
<7>Mycobacterium tuberculosis is one of most prevalent pathogens in the world. Drug-resistant
strains of this pathogen caused by the excessive use
of antibiotics have long posed serious threats to public health worldwide.
A broader picture of drug resistance mechanisms at the genomic level can
be obtained only with large-scale comparative genomic methodology. Two
closely related Beijing family isolates, one resistant to four first-line
drugs (CCDC5180) and one sensitive to them (CCDC5079), were completely
sequenced. These sequences will serve as valuable references for further
drug resistance site identification studies and could be of great
importance for developing drugs targeting these sites.

<>

<1>Zhang, Y., Nelson, M., Nietfeldt, J., Xia, Y., Burbank, D., Ropp, S., Van Etten, J.L.
<2>Chlorella virus NY-2A encodes at least 12 DNA endonuclease/methyltransferase genes.
<3>Virology
<4>240
<5>366-375
<6>1998
<7>The 380-kb chlorella virus NY-2A genome is highly methylated; 45% of the cytosines are
5-methylcytosine and 37% of the adenine are N6-methyladenine.  Based on the
sensitivity/resistance of NY-2A DNA to 80 methylation-sensitive DNA restriction endonucleases,
the virus is predicted to encode at least 10 DNA methyltransferases: 7 6mA-specific
methyltransferases, M.CviQI (GTmAC), M.CvQII (RmAR), M.CviQIII (TCGmA), M.CviQV (TGCmA),
M.CviQVI (GmANTC), and M.CviQVII (CmATG); and 3 5mC-specific methyltransferases, M.CviQVIII
[RGmC(T/C/G)], M.CviQIX (mCC), and M.CviQX (mCGR).  Five of the 6mA methyltransferase genes,
M.CviQI, M.CviQIII, M.CviQV, M.CviQVI, and M.CviQVII, were cloned and sequenced.  In addition,
2 site-specific endonuclease activities, R.CviQI (G/TAC) and NY2A-nickase (R/AG), were
detected in cell-free extracts from NY-2A virus-infected chlorella.  Therefore, the NY-2A
genome contains at least 12 DNA methyltransferase and endonuclease genes which, altogether,
compose about 3-4% of the virus genome.

<>

<1>Zhang, Y., Nelson, M., Nietfeldt, J.W., Burbank, D.E., Van Etten, J.L.
<2>Characterization of Chlorella virus PBCV-1 CviAII restriction and modification system.
<3>Nucleic Acids Res.
<4>20
<5>5351-5356
<6>1992
<7>A second DNA site-specific (restriction) endonuclease (R.CviAII) and its cognate adenine DNA
methyltransferase (M.CviAII) were isolated from virus PBCV-1 infected Chlorella strain NC64A
cells. R.CviAII, a heteroschizomer of the bacterial restriction endonuclease NlaIII,
recognizes the sequence CATG, and does not cleave CmATG sequences. However, unlike NlaIII,
which cleaves after the G and does not cleave either CmATG or mCATG sequences, CviAII cleaves
between the C and A and is unaffected by mCATG methylation. The M.CviAII and R.CviAII genes
were cloned and their DNA sequences were determined. These genes are tandemly arranged
head-to-tail such that the TAA termination codon of the M.CviAII methyltransferase gene
overlaps the ATG translational start site of R.CviAII endonuclease. R.CviAII is the first
Chlorella virus site-specific endonuclease gene to be cloned and sequenced.

<>

<1>Zhang, Y., Nelson, M., Van Etten, J.L.
<2>A single amino acid change restores DNA cytosine methyltransferase activity in a cloned Chlorella virus pseudogene.
<3>Nucleic Acids Res.
<4>20
<5>1637-1642
<6>1992
<7>The Chlorella virus PBCV-1 contains an open reading frame, named P17-ORF4, which differs by
eight amino acids from a DNA cytosine methyltransferase, M.CviJI, encoded by a different
chlorella virus IL-3A. Whereas IL-3A expresses M.CviJI, which methylates the central cytosine
in (A/G)GC(T/C/G) sequences, P17-ORF4 is non-functional. Gene fusions between P17-ORF4 and
M.CviJI and site-directed point mutations revealed that changing Gln188 to Lys188 abolishes
M.CviJI methyltransferase activity. Conversely, changing Lys188 in P17-ORF4 to Gln188 results
in M.CviJI activity. The other altered seven amino acids do not appear to affect M.CviJI
activity.

<>

<1>Zhang, Y., Qin, F., Qiao, J., Li, G., Shen, C., Huang, T., Hu, Z.
<2>Draft Genome Sequence of Rhodococcus sp. Strain P14, a Biodegrader of High-Molecular-Weight Polycyclic Aromatic Hydrocarbons.
<3>J. Bacteriol.
<4>194
<5>3546
<6>2012
<7>The genus Rhodococcus is known for its ability to degrade various xenobiotic compounds.
Rhodococcus sp. strain P14 isolated from crude oil-contaminated
sediments can degrade mineral oil and polycyclic aromatic hydrocarbons (PAHs).
The draft genome sequence of Rhodococcus sp. P14 was obtained using Solexa
technology, which provided an invaluable genetic background for further
investigation of the ability of P14 to degrade xenobiotic compounds.

<>

<1>Zhang, Y., Yang, J., Xu, L., Zhu, Y., Liu, B., Shao, Z., Zhang, X., Jin, Q.
<2>Complete Genome Sequence of Neisseria meningitidis Serogroup A Strain NMA510612,  Isolated from a Patient with Bacterial Meningitis in China.
<3>Genome Announcements
<4>2
<5>e00360-14
<6>2014
<7>Serogroup A meningococcal strains have been involved in several pandemics and a series of
epidemics worldwide in the past. Determination of the genome sequence
of the prevalent genotype strain will help us understand the genetic background
of the evolutionary and epidemiological properties of these bacteria. We
sequenced the complete genome of Neisseria meningitidis NMA510612, a clinical
isolate from a patient with meningococcal meningitis.

<>

<1>Zhang, Y., Zhao, Z., Deng, W., Xie, X., Jiao, N.
<2>Draft Genome Sequence of Euryhalocaulis caribicus Strain JL2009T, a New Member of the Family Hyphomonadaceae Isolated from the Caribbean Sea.
<3>Genome Announcements
<4>1
<5>e00407-13
<6>2013
<7>Euryhalocaulis caribicus strain JL2009(T) is a novel genus and species of the family
Hyphomonadaceae and was first isolated from surface water in the Caribbean
Sea. Here, we report the first draft genome from this genus. Its genome contains
genes encoding proteins that are involved in organic acid metabolism and probable
low-affinity inorganic phosphate transporters, which suggests its competence in
oligotrophic oceans.

<>

<1>Zhang, Y.L., Ong, C.T., Leung, K.Y.
<2>Molecular analysis of genetic differences between virulent and avirulent strains of Aeromonas hydrophila isolated from diseased fish.
<3>Microbiology
<4>146
<5>999-1009
<6>2000
<7>Aeromonas hydrophila, a normal inhabitant of aquatic environments, is an
opportunistic pathogen of a variety of aquatic and terrestrial animals,
including humans. A. hydrophila PPD134/91 is defined as virulent whereas
PPD35/85 is defined as avirulent on the basis of their different LD50
values in fish. Suppression subtractive hybridization (SSH) was used to
identify genetic differences between these two strains. Sixty-nine genomic
regions of differences were absent in PPD35/85, and the DNA sequences of
these regions were determined. Sixteen ORFs encoded by 23 fragments showed
high homology to known proteins of other bacteria. ORFs encoded by the
remaining 46 fragments were identified as new proteins of A. hydrophila,
showing no significant homology to any known proteins. Among these
PPD134/91-specific genes, 22 DNA fragments (21 ORFs) were present in most
of the eight virulent strains studied but mostly absent in the seven
avirulent strains, suggesting that they are universal virulence genes in
A. hydrophila. The PPD134/91-specific genes included five known virulence
factors of A. hydrophila: haemolysin (hlyA), protease (oligopeptidase A),
outer-membrane protein (Omp), multidrug-resistance protein and
histone-like protein (HU-2). Another 47 DNA fragments (44 ORFs) were
mainly present in PPD134/91, indicating the heterogeneity among motile
aeromonads. Some of these fragments encoded virulence determinants. These
included genes for the synthesis of O-antigen and type II
restriction/modification system. The results indicated that SSH is
successful in identifying genetic differences and virulence genes among
different strains of A. hydrophila.

<>

<1>Zhang, Y.Y., Li, Q.G., Liang, J.X., Zhu, Y.B.
<2>Hairpin probes for real-time assay of restriction endonucleases.
<3>Acta Biochim. Biophys. Sin.
<4>34
<5>329-332
<6>2002
<7>New fluorogenic probes for restriction endonucleases based on molecular beacons were
developed. These hairpin probes were single-stranded
oligonucleotides that had stem-and-loop structure and carried
5'-fluorophor moiety and 3'-quencher moiety. Their stem sequences were
designed as recognition sites for restriction endonucleases and loop
sequences were unrelated nucleotides. Upon cleavage by endonuclease,
these probes became fluorescent and thus could trace the enzyme
activity continuously. Two probes were designed for BglII and NcoI,
respectively, and each was labeled with a fluorophor of different
color. The results showed that the two probes could specifically assay
for the corresponding enzymes either individually or simultaneously in
a real-time mode. Considering the simplicity, quantification and high
throughput, these probes could be extended to other applications such
as drug screening, protein-nucleic acid interaction study and searching
for small molecule DNA cleavage agents.

<>

<1>Zhang, Z., Chen, Y., Fu, Y., Jiao, N.
<2>Draft Genome Sequence of the Novel Exopolysaccharide-Producing Bacterium Altibacter lentus Strain JLT2010T, Isolated from Deep Seawater of the South China  Sea.
<3>Genome Announcements
<4>2
<5>e00954-14
<6>2014
<7>Altibacter lentus strain JLT2010(T) is the type strain of the recently identified novel genus
and species of the family Flavobacteriaceae and was first isolated
from deep seawater of the South China Sea. It can produce exopolysaccharide. Here
we report the first draft genome of JLT2010(T) (3,160,033 bp, with GC content of
42.12%) and major findings from its annotation. It is the first reported genome
in the genus Altibacter.

<>

<1>Zhang, Z., Hai, R., Song, Z., Xia, L., Liang, Y., Cai, H., Liang, Y., Shen, X., Zhang, E., Xu, J., Yu, D., Yu, X.J.
<2>Spatial variation of Yersinia pestis from Yunnan Province of China.
<3>Am. J. Trop. Med. Hyg.
<4>81
<5>714-717
<6>2009
<7>Yunnan Province of China is considered the site of origin for modern
plague. We analyzed the genotypes of eight Yersinia pestis strains
isolated from three counties in Yunnan Province by pulse field gel
electrophoresis (PFGE). PFGE showed that strains isolated from the same
site were identical regardless of hosts or year of isolation. However, Y.
pestis strains isolated from geographically distinct loci were genetically
divergent. Whole genome sequences of two strains from two foci in Yunnan
showed that the genetic variation of Y. pestis strains was caused by
genome rearrangement. We concluded that Y. pestis strains in each epidemic
focus in Yunnan were a clonal population and selected by host
environments. The genomic variability of the Y. pestis strains from
different foci were caused by genome rearrangement, which may provide a
positive selective advantage for Y. pestis to adapt to its host
environments.

<>

<1>Zhang, Z., Liu, C., Zhu, Y., Zhong, Y., Zhu, Y., Zheng, H., Zhao, G.P., Wang, S.Y., Guo, X.
<2>Complete genome sequence of Lactobacillus plantarum JDM1.
<3>J. Bacteriol.
<4>191
<5>5020-5021
<6>2009
<7>Lactobacillus plantarum is a commonly used lacic acid bacteria (LAB) species as probiotics. We
have sequenced the genome of Lactobacillus
plantarum JDM1, which is a Chinese commercial LAB with several probiotic
functions, with a GS 20 system. We recommend each commercial probiotics
strain should has the complete genome sequenced to insure the safety and
stability.

<>

<1>Zhang, Z.J., Chen, C.Q., Manev, H.
<2>Enzymatic regional methylation assay for determination of CpG methylation density.
<3>Anal. Chem.
<4>76
<5>6829-6832
<6>2004
<7>DNA methylation is a critical mechanism for epigenetic gene regulation. In mammalian cells,
CpG methylation density often influences the
transcription level of related genes. Here, we report a short and
accurate method to determine the CpG methylation density of any DNA
region by sequentially applying bisulfite PCR and SssI (CpG) methylase.
We introduced simple and efficient binding and washing steps that
greatly improve the previous methodology and considerably enhance the
accuracy of the assay. The accuracy of this method allows the detection
of differences at the level of a single CpG methylation site when
homogeneous PCR product was used as substrate. We validated our method
by bisulfite sequencing multiple clones of samples with variable levels
of CpG methylation. We envision that for its accuracy, simplicity,
low-cost, flexibility, minimum instrumentation requirements, and
high-throughput potential our method could greatly benefit research on
DNA methylation.

<>

<1>Zhang, Z.Y., Sun, Z.Q., Wang, Z.L., Wen, Z.L., Sun, Q.W., Zhu, Z.Q., Song, Y.Z., Zhao, J.W., Wang, H.H., Zhang, S.L., Guo, X.K.
<2>Complete Genome Sequence of a Novel Clinical Isolate, the Nontuberculous Mycobacterium Strain JDM601.
<3>J. Bacteriol.
<4>193
<5>4300-4301
<6>2011
<7>Mycobacteriosis is on the increase. Nontuberculous mycobacteria (NTM) are resistant to most
antituberculosis drugs naturally. We determined the
complete genome sequence of a novel NTM strain, JDM601, of the
Mycobacterium terrae complex, which was isolated from a patient with
tuberculosis-like disease and with various antibiotic resistances.

<>

<1>Zhao, B., Mesbah, N.M., Dalin, E., Goodwin, L., Nolan, M., Pitluck, S., Chertkov, O., Brettin, T.S., Han, J., Larimer, F.W., Land, M.L., Hauser, L., Kyrpides, N., Wiegel, J.
<2>Complete Genome Sequence of the Anaerobic, Halophilic Alkalithermophile Natranaerobius thermophilus JW/NM-WN-LFT.
<3>J. Bacteriol.
<4>193
<5>4023
<6>2011
<7>The genome of the anaerobic halophilic alkalithermophile Natranaerobius thermophiles consists
of one 3,165,557 bp chromosome and two plasmids
(17,207 bp, 8,689 bp). The present study is the first to report the
completely sequenced genome of an anaerobic polyextremophile and genes
associated with roles in regulation of intracellular osmotic pressure, pH
homeostasis, and growth at elevated temperatures.

<>

<1>Zhao, C., Qu, K.G., Song, Y.J., Ren, J.S., Qu, X.G.
<2>A Universal, Label-Free, and Sensitive Optical Enzyme-Sensing System for Nuclease and Methyltransferase Activity Based on Light Scattering of Carbon Nanotubes.
<3>Adv. Funct. Mater.
<4>21
<5>583-590
<6>2011
<7>A label-free, enzyme-responsive nanosystem that uses a DNA/single-walled carbon nanotube
(SWNT) assembly as the substrate is
demonstrated for the sensitive, universal detection of restriction and
nonrestriction endonucleases as well as methyltransferases in a
homogeneous solution on the basis of light scattering (LS) of carbon
nanotubes. This protocol is based on the different binding affinities
of SWNTs to single-and double-stranded DNA. This difference can lead to
different LS signals that can be used for the detection of nuclease
cleavage activity. The assay only requires a label-free oligonucleotide
probe, significantly reducing the typical cost. The LS technique and
the use of a nuclease-specific oligonucleotide probe impart
extraordinarily high sensitivity and selectivity. This light scattering
assay is universal and label-free with a detection limit of 5 x 10(-6)
U mu L-1 for S1 nuclease, 1 x 10(-4) U mu L-1 for EcoRI endonuclease,
and 1 x 10(-2) U mu L-1 for EcoRI methylase. In principle, this assay
can be used to detect any kind of nuclease by simply changing the DNA
sequences of the specific probe.

<>

<1>Zhao, C.W., Wang, H.Y., Zhang, Y.Z., Feng, H.
<2>Draft Genome Sequence of Bacillus pumilus BA06, a Producer of Alkaline Serine Protease with Leather-Dehairing Function.
<3>J. Bacteriol.
<4>194
<5>6668-6669
<6>2012
<7>Bacillus pumilus BA06 was isolated from the proteinaceous soil and produced an extracellular
alkaline protease with leather-dehairing function. The genome of
BA06 was sequenced. The comparative genome analysis indicated that strain BA06 is
different in genome from the other B. pumilus strains, with limited insertions,
deletions, and rearrangements.

<>

<1>Zhao, D.M., Jin, F., Huang, H.F.
<2>A review on DNA methyltransferases and early embryo development including DNA methyltransferases I (DnmT1) and DNA methyltransferases  III (DnmT3).
<3>Zhejiang Da Xue Xue Bao Yi Xue Ban
<4>34
<5>281-284
<6>2005
<7>
<>

<1>Zhao, F., Bai, J., Wu, J., Liu, J., Zhou, M., Xia, S., Wang, S., Yao, X., Yi, H., Lin, M., Gao, S., Zhou, T., Xu, Z., Niu, Y., Bao, Q.
<2>Sequencing and Genetic Variation of Multidrug Resistance Plasmids in Klebsiella pneumoniae.
<3>PLoS ONE
<4>5
<5>E10141
<6>2010
<7>Background: The development of multidrug resistance is a major problem in the treatment of
pathogenic microorganisms by distinct antimicrobial agents. Characterizing the genetic
variation among plasmids from different bacterial species or strains is a key step towards
understanding the mechanism of virulence and their evolution.
Results: We applied a deep sequencing approach to 206 clinical strains of Klebsiella
pneumoniae collected from 2002 to 2008 to understand the genetic variation of multidrug
resistance plasmids, and to reveal the dynamic change of drug resistance over time. First, we
sequenced three plasmids (70 Kb, 94 Kb, and 147 Kb) from a clonal strain of K. pneumoniae
using Sanger sequencing. Using the Illumina sequencing technology, we obtained more than 17
million of short reads from two pooled plasmid samples. We mapped these short reads to the
three reference plasmid sequences, and identified a large
number of single nucleotide polymorphisms (SNPs) in these pooled plasmids. Many of these SNPs
are present in drugresistance genes. We also found that a significant fraction of short reads
could not be mapped to the reference sequences, indicating a high degree of genetic variation
among the collection of K. pneumoniae isolates. Moreover, we identified that plasmid
conjugative transfer genes and antibiotic resistance genes are more likely to suffer from
positive selection, as indicated by the elevated rates of nonsynonymous substitution.
Conclusion: These data represent the first large-scale study of genetic variation in multidrug
resistance plasmids and provide insight into the mechanisms of plasmid diversification and the
genetic basis of antibiotic resistance.

<>

<1>Zhao, F.Q., Zhang, X.W., Liang, C.W., Wu, J.Y., Bao, Q.Y., Qin, S.
<2>Genome-wide analysis of restriction-modification system in unicellular and filamentous cyanobacteria.
<3>Physiol. Genomics
<4>24
<5>181-190
<6>2006
<7>Cyanobacteria are an ancient group of gram-negative bacteria with strong genome size variation
ranging from 1.6 to 9.1 Mb. Here, we first
retrieved all the putative restriction-modification (RM) genes in the
draft genome of Spirulina and then performed a range of comparative and
bioinformatic analyses on RM genes from unicellular and filamentous
cyanobacterial genomes. We have identified 6 gene clusters containing
putative Type I RMs and 11 putative Type II RMs or the solitary
methyltransferases (MTases). RT-PCR analysis reveals that 6 of 18
MTases are not expressed in Spirulina, whereas one hsdM gene, with a
mutated cognate hsdS, was detected to be expressed. Our results
indicate that the number of RM genes in filamentous cyanobacteria is
significantly higher than in unicellular species, and this expansion of
RM systems in filamentous cyanobacteria may be related to their wide
range of ecological tolerance. Furthermore, a coevolutionary pattern is
found between hsdM and hsdR, with a large number of site pairs
positively or negatively correlated, indicating the functional
importance of these pairing interactions between their tertiary
structures. No evidence for positive selection is found for the
majority of RMs, e. g., hsdM, hsdS, hsdR, and Type II restriction
endonuclease gene families, while a group of MTases exhibit a
remarkable signature of adaptive evolution. Sites and genes identified
here to have been under positive selection would provide targets for
further research on their structural and functional evaluations.

<>

<1>Zhao, G., Li, J., Tong, Z., Zhao, B., Mu, R., Guan, Y.
<2>Enzymatic Cleavage of Type II Restriction Endonucleases on the 2 '-O-Methyl Nucleotide and Phosphorothioate Substituted DNA.
<3>PLoS ONE
<4>8
<5>e79415
<6>2013
<7>The effects of nucleotide analogue substitution on the cleavage efficiencies of type II
restriction endonucleases have been investigated. Six restriction endonucleases (EcoRV, SpeI,
XbaI, XhoI, PstI and SphI) were investigated respectively regarding their cleavage when
substrates were substituted by 2'-O-methyl nucleotide (2'-OMeN) and phosphorothioate (PS).
Substitutions were made in the recognition sequence and the two nucleotides flanking the
recognition sequence for each endonuclease. The endonu lease cleavage efficiencies were
determined using FRET-based assay. Results demonstrated a position-dependent inhibitory effect
of substitution on the cleavage efficiency for all the six endonucleases. In general, the
2'-OMeN substitutions had greater impact than the PS substitutions on the enzymatic
activities. Nucleotides of optimal substitutions for protection against RE cleavage were
identified. Experimental results and conclusions in this study facilitate our insight into the
DNA-protein interactions  and the enzymatic cleavage mechanism, particularly for those whose
detailed structure information is not available. In addition, the information could benefit
the development of bioengineering and synthetic biology.

<>

<1>Zhao, L.
<2>Characterization of bacterial homing endonuclease I-Ssp6803I.
<3>Ph.D. Thesis, University of Washington, Seattle, USA
<4>
<5>
<6>2008
<7>The homing endonuclease I-Ssp6803I causes the insertion and persistence of a group I
self-splicing intron into the anticodon loop of the tRNAfMet gene in the cyanobacteria
Synechocystis PCC6803.  This enzyme and its host intron represent the only known combination
of a persistent intron and associated homing endonuclease within bacterial tRNA genes.  In
this study, we predicted that this enzyme, which is dissimilar to other known homing
endonucleases, might contain the PD-(D/E)XK nuclease catalytic motif -- a fold normally
associated with bacterial restriction endonucleases.  This prediction, combined with a
mutational screen for catalytically inactivating point mutations, allowed us to express,
solubilize and crystallize I-Ssp6803I in complex with its target DNA.  The 3.1A crystal
structure of the protein-DNA complex confirmed the presence of the PD-(D/E)XK motif and also
revealed the use of a unique tetrameric assembly to promote recognition of a single long
target site.  The binding specificity profile of I-Ssp6803I was then determined by measuring
the effect of every possible base substitution across its 23-basepair target site.  The
endonuclease displays relatively low binding specificity, corresponding to a profile reflects
sequence constraints on its host tRNAfMet gene.

<>

<1>Zhao, L., Bonocora, R.P., Shub, D.A., Stoddard, B.L.
<2>The restriction fold turns to the dark side: a bacterial homing endonuclease with a PD-(D/E)-XK motif.
<3>EMBO J.
<4>26
<5>2432-2442
<6>2007
<7>The homing endonuclease I-Ssp6803I causes the insertion of a group I intron into a bacterial
tRNA gene-the only example of an invasive mobile
intron within a bacterial genome. Using a computational fold prediction,
mutagenic screen and crystal structure determination, we demonstrate that
this protein is a tetrameric PD-(D/E)-XK endonuclease-a fold normally used
to protect a bacterial genome from invading DNA through the action of
restriction endonucleases. I-Ssp6803I uses its tetrameric assembly to
promote recognition of a single long target site, whereas restriction
endonuclease tetramers facilitate cooperative binding and cleavage of two
short sites. The limited use of the PD-(D/E)-XK nucleases by mobile
introns stands in contrast to their frequent use of LAGLIDADG and HNH
endonucleases-which in turn, are rarely incorporated into
restriction/modification systems.

<>

<1>Zhao, L., Pellenz, S., Stoddard, B.L.
<2>Activity and Specificity of the Bacterial PD-(D/E)XK Homing Endonuclease I-Ssp6803I.
<3>J. Mol. Biol.
<4>385
<5>1498-1510
<6>2009
<7>The restriction endonuclease fold [a three-layer alpha-beta sandwich containing variations of
the PD-(D/E)XK nuclease motif] has been greatly
diversified during evolution, facilitating its use for many biological
functions. Here we characterize DNA binding and cleavage by the PD-(D/E)XK
homing endonuclease I-Ssp6803I. Unlike most restriction endonucleases
harboring the same core fold, the specificity profile of this enzyme
extends over a long (17 bp) target site. The DNA binding and cleavage
specificity profiles of this enzyme were independently determined and
found to be highly correlated. However, the DNA target sequence contains
several positions where binding and cleavage activities are not tightly
coupled: individual DNA base-pair substitutions at those positions that
significantly decrease cleavage activity have minor effects on binding
affinity. These changes in the DNA target sequence appear to correspond to
substitutions that uniquely increase the free energy change between the
ground state and the transition state, rather than simply decreasing the
overall DNA binding affinity. The specificity of the enzyme reflects
constraints on its host gene and limitations imposed by the enzyme's
quaternary structure and illustrate the highly diverse repertoire of DNA
recognition specificities that can be adopted by the related folds
surrounding the PD-(D/E)XK nuclease motif.

<>

<1>Zhao, N., Bai, Y., Zhao, X.Q., Yang, Z.Y., Bai, F.W.
<2>Draft Genome Sequence of the Flocculating Zymomonas mobilis Strain ZM401 (ATCC 31822).
<3>J. Bacteriol.
<4>194
<5>7008-7009
<6>2012
<7>Zymomonas mobilis ZM401 is a flocculating strain which can be self-immobilized within
fermentors for a high-cell-density culture to improve ethanol
productivity, as well as high-gravity fermentation to increase ethanol titer, due
to its improved ethanol tolerance associated with the morphological change. Here,
we report its draft genome sequence.

<>

<1>Zhao, W., Chen, Y., Sun, Z., Wang, J., Zhou, Z., Sun, T., Wang, L., Chen, W., Zhang, H.
<2>Complete Genome Sequence of the Lactobacillus helveticus H10.
<3>J. Bacteriol.
<4>193
<5>2666-2667
<6>2011
<7>Lactobacillus helveticus strain H10 was isolated from the traditional fermented milk in Tibet,
China. We sequenced the whole genome of strain
H10 and compared it to the published genome sequence of Lactobacillus
helveticus DPC4571.

<>

<1>Zhao, W., Zeng, X., Xiao, X.
<2>Thermococcus eurythermalis sp. nov., a conditional piezophilic hyperthermophilic archaeon with a wide temperature range isolated from an oil-immersed chimney in the Guaymas Basin.
<3>Int. J. Syst. Evol. Microbiol.
<4>0
<5>0
<6>2014
<7>A conditional piezophilic hyperthermophilic archaeon with a wide temperature/ pH/
pressure range was isolated from an oil-immersed hydrothermal chimney at a depth
of 2006.9 m in the Guaymas Basin. The enrichment and isolation of strain A501T
were performed at 80 degrees C at 0.1 MPa. The isolate, A501T, is an irregular
motile coccus with a polar tuft of flagella and is generally 0.6-2.6 mum in
diameter. Growth was detected over the range of 50-100 degrees C (optimum growth
at 85 degrees C) at atmospheric pressure and was observed at 102 degrees C at a
pressure of 10 MPa. At 85 degrees C, growth was observed at a pressure of 0.1-70
MPa (optimum pressure 0.1 MPa-30 MPa), while at 95 degrees C, the pressure of
growth ranged from 0.1 MPa to 50 MPa (optimum pressure 10 MPa). Cells of strain
A501T grew at pH 4-9 (optimum pH 7.0) and a NaCl concentration of 1.0-5.0% (w/v)
(optimum concentration 2.5%) This isolate was an anaerobic
chemoorgano-heterotroph and was able to utilize yeast extract, peptone, tryptone
and starch as the single carbon source for growth. Elemental sulfur and cysteine
stimulated growth; however, these molecules were not necessary. The G+C content
of the complete genome was 53.47%. The results of the 16S rRNA gene analysis
indicated that strain A501T belongs to the genus Thermococcus. There was no
significant homology between strain A501T and the phylogenetically related
Thermococcus species based on complete genome sequence alignments and calculation
of the average nucleotide identity and the tetranucleotide signature frequency
correlation coefficient. These results indicate that strain A501T represents a
novel species, Thermococcus eurythermalis sp. nov. The type strain is A501T
(=CGMCC 7834T; =JCM 30233T).

<>

<1>Zhao, X., Geng, X., Chen, C., Chen, L., Jiao, W., Yang, C.
<2>Draft Genome Sequence of the Marine Actinomycete Streptomyces sulphureus L180, Isolated from Marine Sediment.
<3>J. Bacteriol.
<4>194
<5>4482
<6>2012
<7>Marine-derived actinobacteria are rich sources of valuable natural products and industrial
enzymes for biotechnology applications. The marine-derived
Streptomyces sulphureus strain L180 was isolated from the marine sediment in a
sea cucumber farm at a depth of about 10 m in Dalian, China, and its 16S rRNA
gene sequence was determined to have the highest identity to that of Streptomyces
sulphureus NRRL B-1627(T) (99.65%). Here, we report the draft genome sequence of
this strain.

<>

<1>Zhao, X., Yang, T.
<2>Draft Genome Sequence of the Marine Sediment-Derived Actinomycete Streptomyces xinghaiensis NRRL B24674T.
<3>J. Bacteriol.
<4>193
<5>5543
<6>2011
<7>Actinobacteria are a rich source of novel natural products. We recently characterized
Streptomyces xinghaiensis NRRL B24674(T) as a novel species
of marine origin. Here we report the draft genome sequence of this
species. This is the first validly published marine streptomycete for
which a genome sequence has been presented.

<>

<1>Zhao, Y., Caspers, M.P., Abee, T., Siezen, R.J., Kort, R.
<2>Complete Genome Sequence of Geobacillus thermoglucosidans TNO-09.020, a Thermophilic Sporeformer Associated with a Dairy-Processing Environment.
<3>J. Bacteriol.
<4>194
<5>4118
<6>2012
<7>Thermophilic spore-forming bacteria are a common cause of contamination in dairy  products. We
isolated the thermophilic strain Geobacillus thermoglucosidans
TNO-09.020 from a milk processing plant and report the complete genome of a dairy
plant isolate consisting of a single chromosome of 3.75 Mb.

<>

<1>Zhao, Y., Chen, M., Su, L., Wang, T., Liu, S., Yang, H.
<2>Molecular cloning and expression-profile analysis of sea cucumber DNA (Cytosine-5)-methyltransferase 1 and methyl-CpG binding domain type 2/3 genes during aestivation.
<3>Comp. Biochem. Physiol. B, Biochem. Mol. Biol.
<4>165
<5>26-35
<6>2013
<7>The sea cucumber Apostichopus japonicus Selenka survives high summer temperature by entering
aestivation, characterized by hypometabolism and global gene silencing. We investigated the
hypothesis that aestivation is associated with DNA methylation-dependent epigenetic mechanisms
by cloning, sequencing and measuring the transcript abundances of two genes dnmt1 and mbd2/3,
which comprise the DNA methylation system in A. japonicus Selenka. The deduced amino acid
sequences and characteristic motifs of sea cucumber DNMT1 and MBD2/3 showed high homology to
those of their mammalian counterparts. Quantitative real-time RT-PCR analysis showed that
dnmt1 and mbd2/3 genes were similarly expressed in all four tissues examined (intestine,
respiratory tree, muscle and body wall). dnmt1 expression in the intestine was up-regulated
during deep aestivation (P < 0.05), while mbd2/3 was over-expressed in both the intestine and
respiratory tree during the same period (P < 0.01). No differences in expression levels were
observed between other tissues. The results of this study suggest that DNA methylation may be
involved in transcriptional silencing, and that the intestine is the major site for epigenetic
regulation during aestivation in the sea cucumber.

<>

<1>Zhao, Z., Wang, L., Wang, B., Liang, H., Ye, Q., Zeng, M.
<2>Complete Genome Sequence of Cronobacter sakazakii Strain CMCC 45402.
<3>Genome Announcements
<4>2
<5>e01139-13
<6>2014
<7>Cronobacter sakazakii is considered to be an important pathogen involved in life-threatening
neonatal infections. Here, we report the annotated complete
genome sequence of C. sakazakii strain CMCC 45402, obtained from a milk sample in
China. The major findings from the genomic analysis provide a better
understanding of the isolates from China.

<>

<1>Zhao, Z.-H., Yung, M.-H., Cui, T., Wang, Y.-Z., Shaw, P.C.
<2>Restriction endonuclease from thermophilic bacterial species IV. Isolation and characterization of BpuB5I.
<3>Nucleic Acids Res.
<4>20
<5>1156
<6>1992
<7>BpuB5I is a type II restriction endonuclease from a Bacillus pumilus strain
isolated from soil.  To purify the enzyme, bacterial cells were grown on nine L
agar plates at 55C overnight.  Cell pellet (3 g wet weight) obtained was
resuspended in 10 ml extraction buffer (1) and disrupted by sonication.  The
extract after centrifugation was dialysed against extraction buffer at 4C
overnight and then passed through a heparin-affin gel column (7 ml bed volume).
Enzyme eluted at 0.2 - 0.25 M NaCl gradient was concentrated and dialysed and
then further purified by a Mono Q column with enzyme eluted at 0.3 M NaCl.  The
purified enzyme was used to digest lambda, PhiX-174 and Ad-2 DNA (Fig. 1).
Restriction patterns produced were identifical to those cleaved by mesophilic
enzyme SplI which recognizes 5'CGTACG3'(2).  The cleavage site of BpuB5I was
determined according to ref. 1 using PhiX-174 as the template and a 18 mer
oligonucleotide with sequence 5'TCCGACTACCCTCCCGAC3' as the primer.  The primer
was annealed to position 2691 - 2708 of the denatured PhiX-174 DNA and was
extended through the BpuB5I site at position 2794.  Figure 2 shows that the
cleaveage product of reaction I comigrates with the band corresponding to the
first C nucleotide in the sequence CGTACG and reaction II comigrates with the
band corresponding to the second C nucleotide.  Therefore, BpuB5I recognizes
and cleaves 5'C^GTACG3'.  The optimal reaction buffer of BpuB5I contained 5 mM
NaCl, 10 mM Tris.Cl, pH 8.0, 10 mM MgCl2, 1 mM dithiothreitol and optimal
temperature was at 55C.

<>

<1>Zharmukhamedova, T.Y., Privezencova, N.G., Shishova, O.V., Shiryaev, S.A., Zheleznaya, L.A., Matvienko, N.I.
<2>A site-specific endonuclease from the thermophilic strain Bacillus species F4.
<3>Biokhimiia
<4>64
<5>697-702
<6>1999
<7>A strain producing the site-specific endonuclease BspF4I was found during screening of
thermophilic bacteria isolated from soil.  The restriction endonuclease, free from contaminant
nonspecific nucleases, was purified using three steps of column chromatography--on
hydroxyapatite, blue agarose, and DEAE-Trisacryl. The enzyme is stable on storage and exhibits
maximal activity at 48-56 degrees C in the presence of albumin in buffer containing 10 mM
Tris-HCl (pH 7.5) and 10 mM MgCl2.  BspF4I recognizes the sequence 5;-GGNCC-3; on DNA and is
an isomer and not an isoschizomer of the endonuclease Sau96I.  Unlike the prototype, BspF4I
does not cleave the site in a defined way. A strand with purine in the center of the sequence
is cleaved after the first G, as in the case of the prototype, while the strand with
pyrimidine is cleaved either before or after the first G.

<>

<1>Zheleznaya, L., Shiryaev, S., Zheleznyakova, E., Matvienko, N., Matvienko, N.
<2>R.SspD5I is a neoschizomer of HphI producing blunt end DNA fragments.
<3>FEBS Lett.
<4>448
<5>38-40
<6>1999
<7>The strain Staphylococcus species D5 produces a restriction enzyme.  It is the neoschizomer of
HphI endonuclease, which cleaves DNA at a distance of eight nucleotides from the recognition
sequences producing blunt end DNA fragments: 5'-GGTGA8N/-3' and 3'-CCACT8N/-5'.

<>

<1>Zheleznaya, L.A., Kainov, D.E., Yunusova, A.K., Matvienko, N.I.
<2>Regulatory C protein of the EcoRV modification-restriction system.
<3>Biokhimiia
<4>68
<5>125-132
<6>2003
<7>The C gene product of the modification-restriction system PvuII binds to its own promoter (C
box) and stimulates transcription of both the C gene and the endonuclease gene. According to
our data the same regulatory mechanism is realized in the EcoRV system. It was found that
upstream of the EcoRV endonuclease gene two ATG codons give rise to two open reading frames
(ORF1 and ORF2) ending at the same point inside the endonuclease gene. Two DNA fragments
corresponding to ORF1 and ORF2 were cloned, and the homogenous products of proteins encoded by
them were found to be DNA-binding proteins. A specific DNA sequence (C box) recognized by the
proteins was determined with DNAse I footprinting. The C box CCCATTTTGGGTTATCCCATTTTGGG is
located inside ORF1 and, similar to the PvuII C box consisting of tandem repeats of 11
nucleotides, is divided by four nucleotides. In its turn each of the repeats contains inverted
repeats of four terminal nucleotides. The EcoRV C box sequence differs both from the PvuII C
box sequence and from the proposed consensus sequence of C boxes in other
modification-restriction systems.

<>

<1>Zheleznaya, L.A., Matvienko, N.N., Matvienko, N.I.
<2>Two site-specific endonucleases from the thermophilic strain of Bacillus species LU11.
<3>Biokhimiia
<4>58
<5>1315-1322
<6>1993
<7>Upon screening of natural strains of thermophilic bacteria, a strain has been found which
contains two restriction endonucleases. One of those, BspLU11II, is an isoschizomer of XbaI,
while the other one, BspLU11I, recognizes the new palindromic sequence 5'-A^CATGT-3' and
cleaves it as indicated by the arrow. Functionally pure enzymes were obtained by stepwise
chromatography with blue agarose, hydroxyapatite and heparin-Sepharose. The restriction
endonuclease BspLU11I produces sticky ends identical to those produced by the restriction
endonuclease NcoI; hence a combination of BspLU11I and NcoI can be used for enzymatic
selection of recombinant DNA. The recognition sequence of BspLU11I contains the ATG codon and
can be used to construct expression vectors for chemically synthesized genes. ies.

<>

<1>Zheleznaya, L.A., Perevyazova, T.A., Alzhanova, D.V., Matvienko, N.I.
<2>Site-specific nickase from Bacillus species strain D6.
<3>Biokhimiia
<4>66
<5>1215-1220
<6>2001
<7>Three site-specific endonucleases were found in thermophilic strain Bacillus species D6. One
of them, BspD6II, is an isoschizomer of
Eco57I. The second, BspD6III, is present in the strain in very small
amount; therefore, it has not been characterized. This paper is devoted
to the third, BspD6I, which recognizes the pentanucleotide site 5'-GAGTC-3'
and cleaves only one DNA strand at a distance of 4 nucleotides from the
site in the 3'-direction in the chain with the GAGTC sequence, i.e., it
behaves as a site-specific nickase. Nickase N.BspD6I cleaves one DNA
strand only in double-stranded DNA and does not cleave single-stranded
DNA. Site-specific methylase SscL1I (an isohypectomer of M.HinfI) that
methylates adenine in the sequence 5'-GANTC-3' prevents DNA hydrolysis
by nickase BspD6I.

<>

<1>Zheleznaya, L.A., Perevyazova, T.A., Zheleznyakova, E.N., Matvienko, N.I.
<2>Some properties of site-specific nickase BspD6I and the possibility of its use in hybridization analysis of DNA.
<3>Biokhimiia
<4>67
<5>595-600
<6>2002
<7>A new method for hybridization analysis of nucleic acids is proposed on the basis of the
ability of site-specific nickases to cleave only one DNA strand. The method is based on the
use of a labeled oligonucleotide with the recognition site of the nickase hybridized with the
target (DNA or RNA) at an optimal temperature of the enzyme (55 degrees C). The two shorter
oligonucleotides formed after the cleavage with the nickase do not complex with the target.
Thus, a multiple cleavage of the labeled oligonucleotide takes place on one target molecule.
The cleavage of the nucleotide is recorded either by polyacrylamide gel electrophoresis (when
a radioactive labeled oligonucleotide is used) or by fluorescence measurements (if the
oligonucleotide has the structure of a molecular beacon). The new method was tested on nickase
BspD6I and a radioactive oligonucleotide complementary to the polylinker region of the viral
DNA strand in bacteriophage M13mp19. Unfortunately, nickase BspD6I does not cleave DNA in the
RNA-DNA duplexes and therefore cannot be used for detection of RNA targets.

<>

<1>Zheleznyakova, E.N., Zheleznaya, L.A., Svadbina, I.V., Matvienko, N.I.
<2>A site-specific endonuclease from thermophilic strain Bacillus species ZE is a ClaI isoschizomer.
<3>Biokhimiia
<4>63
<5>1675-1681
<6>1998
<7>Strain Bacillus species ZE with a maximal yield of the ClaI isoschizomer was chosen from 12
natural thermophilic strains producing ClaI isomers.  The yield is 110 times higher than that
of the ClaI prototype endonuclease from Caryophanon latum L.  The enzyme is sensitive to DNA
dam-methylation and is highly stable under storage.

<>

<1>Zheng, B., Tomita, H., Inoue, T., Ike, Y.
<2>Isolation of VanB-type Enterococcus faecalis strains from nosocomial infections:  first report of the isolation and identification of the pheromone-responsive  plasmids pMG2200, Encoding VanB-type vancomycin resistance and a Bac41-type  bacteriocin, and.
<3>Antimicrob. Agents Chemother.
<4>53
<5>735-747
<6>2009
<7>Eighteen identical VanB-type Enterococcus faecalis isolates that were obtained from different
hospitalized patients were examined for their drug resistance and
plasmid DNAs. Of the 18 strains, 12 strains exhibited resistance to erythromycin
(Em), gentamicin (Gm), kanamycin (Km), tetracycline (Tc), and vancomycin (Van)
and produced cytolysin (Hly/Bac) and a bacteriocin (Bac) active against E.
faecalis strains. Another six of the strains exhibited resistance to Gm, Km, Tc,
and Van and produced a bacteriocin. Em and Van resistance was transferred
individually to E. faecalis FA2-2 strains at a frequency of about 10(-4) per
donor cell by broth mating. The Em-resistant transconjugants and the
Van-resistant transconjugants harbored a 65.7-kbp plasmid and a 106-kbp plasmid,
respectively. The 106-kbp and 65.7-kbp plasmids isolated from the representative
E. faecalis NKH15 strains were designated pMG2200 and pMG2201, respectively.
pMG2200 conferred vancomycin resistance and bacteriocin activity on the host
strain and responded to the synthetic pheromone cCF10 for pCF10, while pMG2201
conferred erythromycin resistance and cytolysin activity on its host strain and
responded to the synthetic pheromone cAD1 for pAD1. The complete DNA sequence of
pMG2200 (106,527 bp) showed that the plasmid carried a Tn1549-like element
encoding vanB2-type resistance and the Bac41-like bacteriocin genes of
pheromone-responsive plasmid pYI14. The plasmid contained the regulatory region
found in pheromone-responsive plasmids and encoded the genes prgX and prgQ, which
are the key negative regulatory elements for plasmid pCF10. pMG2200 also encoded
TraE1, a key positive regulator of plasmid pAD1, indicating that pMG2200 is a
naturally occurring chimeric plasmid that has a resulting prgX-prgQ-traE1 genetic
organization in the regulatory region of the pheromone response. The functional
oriT region and the putative relaxase gene of pMG2200 were identified and found
to differ from those of pCF10 and pAD1. The putative relaxase of pMG2200 was
classified as a member of the MOB(MG) family, which is found in
pheromone-independent plasmid pHTbeta of the pMG1-like plasmids. This is the
first report of the isolation and characterization of a pheromone-responsive
highly conjugative plasmid encoding vanB resistance.

<>

<1>Zheng, D., Koller, W.
<2>Characterization of the mitochondrial cytochrome b gene from Venturia inaequalis.
<3>Curr. Genet.
<4>32
<5>361-366
<6>1997
<7>A new class of agricultural fungicides derived from the group of antifungal strobilurins acts
as specific respiration inhibitors by binding to mitochondrial cytochrome b.  The cytochrome b
gene was cloned and sequenced from the mitochondrial genome of Venturia inaequalis, the causal
agent of apple scab.  The gene was 10.65 kbp in size and contained seven exons and six
introns.  The exons encoded a protein of 393 amino acids.  Comparison of the deduced
amino-acid sequence with cytochrome b proteins from other fungi revealed highest homologies to
the respective proteins of Aspergillis nidulans, Podospora anserina and Neurospora crassa.
All amino acids of the V. inaequalis cytochrome b at positions altered in mutants of
Saccharomyces cerevisiae resistant to strobilurins, and other fungi with reduced sensitivities
to strobilurins, were identical to wild-type isolates of several fungi.  The cloning and
characterization of the V. inaequalis cytochrome b gene is the initial step in the assessment
of resistance risks inherent to the strobilurin fungicides.

<>

<1>Zheng, H., Guo, L., Du N, L.J., Song, L., Liu, G., Chen, F.
<2>Draft Genome Sequences of Two Clinical Isolates of Streptococcus mutans.
<3>Genome Announcements
<4>2
<5>e00441-14
<6>2014
<7>We report the draft genome sequences of PKUSS-HG01 and PKUSS-LG01, two clinical isolates of
Streptococcus mutans from human dental plaque. The genomics
information will facilitate the study of the mechanisms of pathogenicity and
evolution of S. mutans.

<>

<1>Zheng, L., Braymer, H.D.
<2>Overproduction and purification of McrC protein from Escherichia coli K-12.
<3>J. Bacteriol.
<4>173
<5>3918-3920
<6>1991
<7>The McrC protein, encoded by one of the two genes involved in the McrB
restriction system, was produced in Escherichia coli cells by using a T7
expression system.  Following sequential DEAE-Sepharose and hydroxylapatite
column chromatography, the protein was purified to apparent homogeneity as
judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.  The
N-terminal amino acid sequence of the purified McrC protein agreed exactly with
the one deduced from the DNA sequence by Ross et al. (J. Bacteriol.
171:1974-1981, 1989).

<>

<1>Zheng, L., Simmons, L.A., Braymer, H.D.
<2>Determination of the recognition sequence for McrB restriction of E. coli K12.
<3>Abstr. Gen. Meet. Am. Soc. Microbiol.
<4>91
<5>175
<6>1991
<7>Restriction of methylated DNA by the McrB restriction system of E. coli
involves two genes designated mcrB and mcrC.  The mcrB gene is thought to code
for an endonuclease subunit and the mcrC and the mcrC subunit confers
specificity to the enzyme complex.  The restriction enzyme requires GTP for
activity and recognizes G5mC.  However, we know these two deoxynucleotides
alone are not sufficient for recognition since not every methylated AluI site
(AG5mCT) is recognized. To determine the nucleotide recognition sequence for
McrB restriction, a series of deletion plasmids were constructed from pBR322
and tested as substrates of McrB activity, after the plasmids were methylated
with AluI methylase.  Our study first narrowed the McrB recognition sequence
down to one of the four AluI sites in the region between two MspI sites at 1812
and 2121 bp of pBR322.  SI nuclease deletion and further sequencing analyses of
this region revealed that only one AluI site was involved in the recognition of
McrB restriction.  Site-specific mutagenesis will be used to define all base
composition required for McrB recognition.

<>

<1>Zheng, L., Wang, X., Braymer, H.D.
<2>Purification and N-terminal amino acid sequences of two polypeptides encoded by the mcrB gene from Escherichia coli K-12.
<3>Gene
<4>112
<5>97-100
<6>1992
<7>This report provides a purification method for the two proteins, 51 kDa and 33
kDa, both encoded by the same mcrB gene of the McrBC restriction system in
Escherichia coli K-12.  The two proteins were produced in large quantity using
a T7 expression system and copurified to near homogeneity by DEAE-Sepharose and
Affi-Gel blue column chromatography.  The N-terminal amino acid sequences of
these purified McrB proteins were the same as those predicted from the mcrB DNA
sequence by Ross et al. [J. Bacteriol. 171 (1989b) 1974-1981].  The 33 kDa
protein totally overlaps the C-terminal part of the 51-kDa protein.

<>

<1>Zheng, P.X., Chung, K.T., Chiang-Ni, C., Wang, S.Y., Tsai, P.J., Chuang, W.J., Lin, Y.S., Liu, C.C., Wu, J.J.
<2>Complete Genome Sequence of emm1 Streptococcus pyogenes A20, a Strain with an Intact Two-Component System, CovRS, Isolated from a Patient with Necrotizing  Fasciitis.
<3>Genome Announcements
<4>1
<5>e00149-12
<6>2013
<7>Here, we announce the complete sequence of Streptococcus pyogenes A20. This strain was
isolated from a patient with necrotizing fasciitis. Given that A20
harbors an intact two-component system, CovRS, the discovery of its genome
sequence provides more insight into the pathogenesis of a pandemic emm1 strain.

<>

<1>Zheng, Q., Zhang, R., Jiao, N.
<2>Genome Sequence of Citromicrobium Strain JLT1363, Isolated from the South China Sea.
<3>J. Bacteriol.
<4>193
<5>2074-2075
<6>2011
<7>Citromicrobium is a member of the alpha-4 subcluster in the Alphaproteobacteria and is
identified as a typical aerobic anoxygenic
phototrophic bacterium (AAPB). Here we report the draft genome sequence of
a non-AAPB strain, Citromicrobium sp. JLT1363. The genome sequence reveals
a multimechanism of horizontal gene transfer, as well.

<>

<1>Zheng, W.
<2>Isolation and purification of the restriction endonuclease Bsu1076I.
<3>Weishengwuxue Tongbao
<4>14
<5>105-108
<6>1987
<7>Restriction endonuclease Bsu1076I has been isolated from Bacillus subtilis and
purified by passing first through a phosphocellulose (Whatman P11) column, and
then through a heparin-Sepharose 4B column.  Bsu1076I was eluted at 0.83-0.87M
NaCl in a gradient from the phosphocellulose column and at 0.80-0.87M NaCl from
heparin-Sepharose 4B column.  As much as 3000 units of highly purified Bsu1076I
was obtained from 20g of wet cells by this procedure.  Digestion of lambda DNA,
SPP1 DNA, PhiX174 DNA, SV40 DNA and Ad-2 DNA with Bsu1076I formed DNA fragments
identical to those digested with its isoschizomer HaeIII (from Promega Biotec)
as assayed by agarose gel electrophoresis.  Over digestion of 1 microgram
lambda DNA with 50-150 units of Bsu1076I showed typical bands on agarose gel,
indicating the exclusion of contamination by other deoxyribonucleases.  After
digestion with Bsu1076I lambda DNA can be fairly ligated and recut.

<>

<1>Zheng, W., Kathariou, S.
<2>Host-mediated modification of Sau3AI restriction in Listeria monocytogenes: Prevalence in epidemic-associated strains.
<3>Appl. Environ. Microbiol.
<4>63
<5>3085-3089
<6>1997
<7>Most major food-related outbreaks of listeriosis have been traced to a cluster of genetically
related strains of serovar 4b (epidemic clone).  In spite of numerous searches, distinct
bacteriologic or virulence-related features unique to these strains have eluded
identification, although a restriction fragment length polymorphism characteristic of the
epidemic clone has previously been described.  We found that DNAs from 75 strains which were
derived from three separate outbreaks and which had the epidemic clone-specific RFLP were also
invariably resistant to digestion by Sau3AI and other restriction endonucleases sensitive to
cytosine methylation at 5' GATC 3' sites.  This modification of Sau3AI restriction was host
mediated, as it did not persist when DNA was cloned and propagated in Escherichia coli, and
was uncommon among other Listeria strains.  Epidemic-associated strains with this modification
were resistant to infection by phage propagated in a serotype 4b strain which was not known to
be involved in an epidemic and which lacked the epidemic clone-specific RFLP.  Screening for
susceptibility to MboI digestion revealed that these epidemic strains lacked methylation of
adenines at GATC sites.  This type of modification was rare among Listeria strains and was
found in only three (of eight screened) strains of serovar 1/2b, possibly representing one
clonal lineage.

<>

<1>Zheng, Y., Cohen-Karni, D., Xu, D., Chin, H.G., Wilson, G., Pradhan, S., Roberts, R.J.
<2>A unique family of Mrr-like modification-dependent restriction endonucleases.
<3>Nucleic Acids Res.
<4>38
<5>5527-5534
<6>2010
<7>Mrr superfamily of homologous genes in microbial genomes restricts modified DNA in vivo.
However, their biochemical properties in vitro
have remained obscure. Here, we report the experimental
characterization of MspJI, a remote homolog of Escherichia coli's Mrr
and show it is a DNA modification-dependent restriction endonuclease.
Our results suggest MspJI recognizes (CNNR)-C-m (R = G/A) sites and
cleaves DNA at fixed distances (N-12/N-16) away from the modified
cytosine at the 3' side (or N-9/N-13 from R). Besides 5-methylcytosine,
MspJI also recognizes 5-hydroxymethylcytosine but is blocked by
5-glucosylhydroxymethylcytosine. Several other close homologs of MspJI
show similar modification-dependent endonuclease activity and display
substrate preferences different from MspJI. A unique feature of these
modification-dependent enzymes is that they are able to extract small
DNA fragments containing modified sites on genomic DNA, for example
similar to 32 bp around symmetrically methylated CG sites and similar
to 31 bp around methylated CNG sites. The digested fragments can be
directly selected for high-throughput sequencing to map the location of
the modification on the genomic DNA. The MspJI enzyme family, with
their different recognition specificities and cleavage properties,
provides a basis on which many future methods can build to decode the
epigenomes of different organisms.

<>

<1>Zheng, Y., Posfai, J., Morgan, R.D., Vincze, T., Roberts, R.J.
<2>Using shotgun sequence data to find active restriction enzyme genes.
<3>Nucleic Acids Res.
<4>37
<5>e1-11
<6>2009
<7>Whole genome shotgun sequence analysis has become the standard method for beginning to
determine a genome sequence. The preparation of the shotgun sequence clones is, in fact, a
biological experiment. It determines which segments of the genome can be cloned into
Escherichia coli and which cannot. By analyzing the complete set of sequences from such an
experiment, it is possible to identify genes lethal to E. coli. Among this set are genes
encoding restriction enzymes which, when active in E. coli, lead to cell death by cleaving the
E. coli genome at the restriction enzyme recognition sites. By analyzing shotgun sequence data
sets we show that this is a reliable method to detect active restriction enzyme genes in newly
sequenced genomes, thereby facilitating functional annotation. Active restriction enzyme genes
have been identified, and their activity demonstrated biochemically, in the sequenced genomes
of Methanocaldococcus jannaschii, Bacillus cereus ATCC 10987 and Methylococcus capsulatus.

<>

<1>Zheng, Y., Roberts, R.J.
<2>Selection of restriction endonucleases using artificial cells.
<3>Nucleic Acids Res.
<4>35
<5>e83
<6>2007
<7>We describe in this article an in vitro system for the selection of restriction endonucleases
using artificial cells. The artificial cells are
generated in the form of a water-in-oil emulsion by in vitro
compartmentalization. Each aqueous compartment contains a reconstituted
transcription/translation mix along with the dispersed DNA templates. In
the compartments containing endonuclease genes, an endonuclease expressed
in vitro cleaves its own DNA template adjacent to the gene, leaving a
sticky end. The pooled DNA templates are then ligated to an adaptor with a
compatible end. The endonuclease genes are then enriched by
adaptor-specific PCR on the ligation mix. We demonstrate that the system
can achieve at least 100-fold enrichment in a single round of selection.
It is sensitive enough to enrich an active endonuclease gene from a
1:10(5) model library in 2-3 rounds of selection. Finally, we describe
experiments where we selected endonuclease genes directly from a bacterial
genomic DNA source in three rounds of selections: the known PstI gene from
Providencia stuartii and the new TspMI gene from Thermus sp. manalii. This
method provides a unique tool for cloning restriction endonuclease genes
and has many other potential applications.

<>

<1>Zheng, Y., Roberts, R.J.
<2>Selection and enrichment of proteins using in vitro compartmentalization.
<3>International Patent Office
<4>WO 2008103900 A
<5>
<6>2008
<7>Compositions and methods are provided for selection and enrichment of a target gene from a
library of polynucleotide sequences such as might be formed from a genome or by random
mutagenesis of a genetic sequence.  The selection and enrichment occurs in aqueous droplets
formed in an emulsion that compartmentalize individual polynucleotides from the library or a
plurality of polynucleotides that may include polynucleotides not derived from the library,
transcription and translation reagents and optionally additional chemical and enzyme reagents.
The selection and enrichment method utilizes a polynucleotide adaptor which when ligated to
the polynucleotide fragment enables amplification to occur in the presence of an adaptor
specific primer.

<>

<1>Zheng, Y., Roberts, R.J.
<2>Selection and enrichment of proteins using in vitro compartmentalization.
<3>US Patent Office
<4>US 20080206832
<5>
<6>2008
<7>Compositions and methods are provided for selection and enrichment of a target gene from a
library of polynucleotide sequences such as might be formed from a genome or by random
mutagenesis of a genetic sequence.  The selection and enrichment occurs in aqueous droplets
formed in an emulsion that compartmentalize individual polynucleotides from the library or a
plurality of polynucleotides that may include polynucleotides not derived from the library,
transcription and translation reagents and optionally additional chemical and enzyme reagents.
The selection and enrichment method utilizes a polynucleotide adaptor which when ligated to
the polynucleotide fragment enables amplification to occur in the presence of an adaptor
specific primer.

<>

<1>Zheng, Y., Roberts, R.J.
<2>Compositions, methods and related uses for cleaving modified DNA.
<3>International Patent Office
<4>WO 2010075375
<5>
<6>2010
<7>Compositions, methods and related uses are provided relating to cleaving modified DNA.  For
example, a set of DNA fragments obtainable by enzymatic cleavage of a large DNA is described
where at least 50% are similarly sized and have a centrally positioned modified nucleotide.
In addition, an enzyme preparation is provided that includes one or more enzymes that
recognize a modified nucleotide in a DNA and cleave the DNA at a site that is at a non-random
distance from the modified nucleotide.  The one or more enzymes are further characterized by
an N-terminal conserved domain with greater than 90% amino acid sequence homology to
WXD(X)10YXGD.  The related uses include creating a methylome, methods of purifying DNA
fragments containing a modified nucleotide and diagnostic applications.

<>

<1>Zheng, Y., Roberts, R.J.
<2>SELECTION AND ENRICHMENT OF PROTEINS USING IN VITRO.
<3>Japanese Patent Office
<4>JP 2010518863 A
<5>
<6>2010
<7>
<>

<1>Zheng, Y., Roberts, R.J.
<2>Selection and enrichment of proteins using in vitro compartmentalization.
<3>US Patent Office
<4>US 8153358 B
<5>
<6>2012
<7>Compositions and methods are provided for selection and enrichment of a target gene from a
library of polynucleotide sequences such as might be formed from a genome or by random
mutagenesis of a genetic sequence.  The selection and enrichment occurs in aqueous droplets
formed in an emulsion that compartmentalize individual polynucleotides from the library or a
plurality of polynucleotides that may include polynucleotides not derived from the library,
transcription and translation reagents and optionally additional chemical and enzyme reagents.
The selection and enrichment method utilizes a polynucleotide adaptor which when ligated to
the polynucleotide fragment enables amplification to occur in the presence of an adaptor
specific primer.

<>

<1>Zheng, Z., Deng, X., Chen, J.
<2>Whole-Genome Sequence of 'Candidatus Liberibacter asiaticus' from Guangdong, China.
<3>Genome Announcements
<4>2
<5>e00273-14
<6>2014
<7>The draft genome sequence of 'Candidatus Liberibacter asiaticus' strain A4, isolated from a
mandarin citrus in Guangdong, People's Republic of China, is reported. The A4 strain has a
genome size of 1,208,625 bp, G+C content of 36.4%, 1,107 predicted open reading frames, and 53
RNA genes.

<>

<1>Zheng, Z., Deng, X., Chen, J.
<2>Draft Genome Sequence of 'Candidatus Liberibacter asiaticus' from California.
<3>Genome Announcements
<4>2
<5>e00999-14
<6>2014
<7>We report here the draft genome sequence of 'Candidatus Liberibacter asiaticus' strain HHCA,
collected from a lemon tree in California. The HHCA strain has a
genome size of 1,150,620 bp, 36.5% G+C content, 1,119 predicted open reading
frames, and 51 RNA genes.

<>

<1>Zhilkina, O.A., Repin, V.E., Degtyarev, S.K.
<2>Effect of some cultivation parameters on the yield of restriction endonuclease SfaNI.
<3>Prikl. Biokhim. Mikrobiol.
<4>27
<5>486-490
<6>1991
<7>A simple technique is proposed for testing restriction endonuclease SfaNI activity in lysates
of Streptococcus faecalis cells.  The technique was used to study the effect of inorganic
phosphate and the growth phase on the enzyme yield.  Conditions were chosen that provide a
high yield of the SfaNI activity and a significantly reduced level of nucleases in the cells.

<>

<1>Zhong, X., Du J, H.C.J., Gallego-Bartolome, J., Feng, S., Vashisht, A.A., Chory, J., Wohlschlegel, J.A., Patel, D.J., Jacobsen, S.E.
<2>Molecular Mechanism of Action of Plant DRM De Novo DNA Methyltransferases.
<3>Cell
<4>157
<5>1050-1060
<6>2014
<7>DNA methylation is a conserved epigenetic gene-regulation mechanism. DOMAINS REARRANGED
METHYLTRANSFERASE (DRM) is a key de novo methyltransferase in plants,
but how DRM acts mechanistically is poorly understood. Here, we report the
crystal structure of the methyltransferase domain of tobacco DRM (NtDRM) and
reveal a molecular basis for its rearranged structure. NtDRM forms a functional
homodimer critical for catalytic activity. We also show that Arabidopsis DRM2
exists in complex with the small interfering RNA (siRNA) effector ARGONAUTE4
(AGO4) and preferentially methylates one DNA strand, likely the strand acting as
the template for RNA polymerase V-mediated noncoding RNA transcripts. This
strand-biased DNA methylation is also positively correlated with strand-biased
siRNA accumulation. These data suggest a model in which DRM2 is guided to target
loci by AGO4-siRNA and involves base-pairing of associated siRNAs with nascent
RNA transcripts.

<>

<1>Zhong, X., Tian, Y., Niu, G., Tan, H.
<2>Assembly and features of secondary metabolite biosynthetic gene clusters in Streptomyces ansochromogenes.
<3>Sci. China Life. Sci.
<4>56
<5>609-618
<6>2013
<7>A draft genome sequence of Streptomyces ansochromogenes 7100 was generated using
454 sequencing technology. In combination with local BLAST searches and gap filling
techniques, a comprehensive antiSMASH-based method was adopted to assemble the secondary
metabolite biosynthetic gene clusters in the draft genome of S. ansochromogenes. A total of at
least 35 putative gene clusters were identified and assembled. Transcriptional analysis showed
that 20 of the 35 gene clusters were expressed in either or all of the three different media
tested, whereas the other 15 gene clusters were silent in all three different media. This
study provides a comprehensive method to identify and assemble secondary metabolite
biosynthetic gene clusters in draft genomes of Streptomyces, and will significantly promote
functional studies of these secondary metabolite biosynthetic gene clusters.

<>

<1>Zhong, Y., Chang, X., Cao, X.J., Zhang, Y., Zheng, H., Zhu, Y., Cai, C., Cui, Z., Zhang, Y., Li, Y.Y., Jiang, X.G., Zhao, G.P., Wang, S., Li, Y., Zeng, R., Li, X., Guo, X.K.
<2>Comparative proteogenomic analysis of the Leptospira interrogans virulence-attenuated strain IPAV against the pathogenic strain 56601.
<3>Cell Res.
<4>21
<5>1210-1229
<6>2011
<7>The virulence-attenuated Leptospira interrogans serovar Lai strain IPAV
was derived by prolonged laboratory passage from a highly virulent
ancestral strain isolated in China. We studied the genetic variations of
IPAV that render it avirulent via comparative analysis against the
pathogenic L. interrogans serovar Lai strain 56601. The complete genome
sequence of the IPAV strain was determined and used to compare with, and
then rectify and reannotate the genome sequence of strain 56601. Aside
from their highly similar genomic structure and gene order, a total of 33
insertions, 53 deletions and 301 single-nucleotide variations (SNVs) were
detected throughout the genome of IPAV directly affecting 101 genes,
either in their 5' upstream region or within their coding region. Among
them, the majority of the 44 functional genes are involved in signal
transduction, stress response, transmembrane transport and nitrogen
metabolism. Comparative proteomic analysis based on quantitative liquid
chromatography (LC)-MS/MS data revealed that among 1 627 selected pairs of
orthologs, 174 genes in the IPAV strain were upregulated, with enrichment
mainly in classes of energy production and lipid metabolism. In contrast,
228 genes in strain 56601 were upregulated, with the majority enriched in
the categories of protein translation and DNA replication/repair. The
combination of genomic and proteomic approaches illustrated that altered
expression or mutations in critical genes, such as those encoding a
Ser/Thr kinase, carbon-starvation protein CstA, glutamine synthetase,
GTP-binding protein BipA, ribonucleotide-diphosphate reductase and
phosphate transporter, and alterations in the translational profile of
lipoproteins or outer membrane proteins are likely to account for the
virulence attenuation in strain IPAV.

<>

<1>Zhong, Z., Toukdarian, A., Helinski, D., Knauf, V., Sykes, S., Wilkinson, J.E., O'Bryne, C., Shea, T., DeLoughery, C., Caspi, R.
<2>Sequence analysis of a 101-kilobase plasmid required for agar degradation by a Microscilla isolate.
<3>Appl. Environ. Microbiol.
<4>67
<5>5771-5779
<6>2001
<7>An agar-degrading marine bacterium identified as a Microscilla species was
isolated from coastal California marine sediment. This organism harbored a
single 101-kb circular DNA plasmid designated pSD15. The complete
nucleotide sequence of pSD15 was obtained, and sequence analysis indicated
a number of genes putatively encoding a variety of enzymes involved in
polysaccharide utilization. The most striking feature was the occurrence
of five putative agarase genes. Loss of the plasmid, which occurred at a
surprisingly high frequency, was associated with loss of agarase activity,
supporting the sequence analysis results.

<>

<1>Zhong, Z., Wang, L., Chen, Y., Wang, Z., Wang, Y., Cui, M., Li, T., Ke, Y., Yuan, X., Wang, D., Chen, Z., Peng, G.
<2>Complete Genome Sequence of Brucella abortus Strain BCB034, a Strain of Biovar 2  Isolated from Human.
<3>J. Bacteriol.
<4>194
<5>6943
<6>2012
<7>Brucella abortus is divided into eight biovars, of which biovars 1 to 3 are the most
frequently represented biovars in strains isolated from humans. Here, we
report the genome sequence of B. abortus strain BCB034, a strain isolated from a
human patient and that belongs to biovar 2.

<>

<1>Zhou, B., Li, Q.
<2>Bacillus subtilis 36 a highly productive strain containing the isoschizomer of restriction enzyme MstII.
<3>Acta Biochim. Biophys. Sin.
<4>19
<5>537-540
<6>1987
<7>From Bacillus subtilis 36 we have isolated and partially purified a restriction
enzyme Bsu36I, which is the isoschizomer of MstII.  200000 units of Bsu36I can
be obtained from one liter of B. subtilis 36 culture.  It is a strain for
production of MstII.

<>

<1>Zhou, E.X., Reuter, M., Meehan, E.J., Chen, L.
<2>A new crystal form of restriction endonuclease EcoRII that diffracts to 2.8 A resolution.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>58
<5>1343-1345
<6>2002
<7>EcoRII, a type IIe restriction endonuclease, has been crystallized in space group P2(1), with
unit-cell parameters a = 58.3, b = 127.8, c = 59.9 A, beta = 91.4 degrees. There are two
monomers in the asymmetric unit and the solvent content is estimated to be 49% by volume. The
crystals diffract to 2.8 A resolution, which is much higher than that of the previously
reported cubic crystal form, which diffracted to 4 A resolution.

<>

<1>Zhou, H., Shatz, W., Purdy, M.M., Fera, N., Dahlquist, F.W., Reich, N.O.
<2>Long-range structural and dynamical changes induced by cofactor binding in DNA methyltransferase M.HhaI.
<3>Biochemistry
<4>46
<5>7261-7268
<6>2007
<7>The bacterial DNA cytosine methyltransferase M.HhaI sequence-specifically modifies DNA in an
S-adenosylmethionine dependent reaction. The enzyme stabilizes the target cytosine (GCGC) into
an extrahelical position, with a concomitant large movement of an active site loop involving
residues 80-99. We used multidimensional, transverse relaxation-optimized NMR experiments to
assign nearly 80% of all residues in the cofactor-bound enzyme form, providing a basis for
detailed structural and dynamical characterization. We examined details of the previously
unknown effects of the cofactor binding with M.HhaI in solution. Addition of the cofactor
results in numerous structural changes throughout the protein, including those decorating the
cofactor binding site, and distal residues more than 30 A away. The active site loop is
involved in motions both on a picosecond to nanosecond time scale and on a microsecond to
millisecond time scale and is not significantly affected by cofactor binding except for a few
N-terminal residues. The cofactor also affects residues near the DNA binding cleft, suggesting
a role for the cofactor in regulating DNA interactions. The allosteric properties we observed
appear to be closely related to the significant amount of dynamics and dynamical changes in
response to ligand binding detected in the protein.

<>

<1>Zhou, H., Wang, Y., Yu, Y., Bai, T., Chen, L., Liu, P., Guo, H., Zhu, C., Tao, M., Deng, Z.
<2>A non-restricting and non-methylating Escherichia coli strain for DNA cloning and high-throughput conjugation to Streptomyces coelicolor.
<3>Curr. Microbiol.
<4>64
<5>185-190
<6>2012
<7>Escherichia coli strains are used in secondary metabolism research for DNA cloning and
transferring plasmids by intergeneric conjugation. Non-restricting
strains are desirable for DNA cloning and non-methylating strains are beneficial
for transferring DNA to methyl-restricting hosts, like Streptomyces coelicolor.
We have constructed a non-methylating E. coli strain, JTU007, by deleting the DNA
methylation genes dcm and dam from the widely used non-restricting cloning host
DH10B. JTU007 was tested as donor for the conjugative transfer of a plasmid
containing the 39 kb actinorhodin biosynthesis gene cluster to S. lividans and S.
coelicolor. The Dcm Dam strain JTU007 transferred DNA into S. coelicolor A(3)2
derivatives at high frequency. To demonstrate the usefulness of E. coli JTU007
for gene cloning, we constructed a comprehensive S. toxytricini genomic cosmid
library, and transferred it using high-throughput conjugation to the
methyl-restricting S. coelicolor. One of the cosmid clones produced a brown
pigment, and the clone was revealed to carry a tyrosinase operon. JTU007 is more
useful than ET12567 because it does not restrict methylated DNA in primary
cloning, and gives higher transformation and cosmid infection frequencies.

<>

<1>Zhou, H., Zhang, T.W., Yu, D.L., Pi, B.R., Yang, Q., Zhou, J.Y., Hu, S.N., Yu, Y.S.
<2>Genomic Analysis of the Multidrug-Resistant Acinetobacter baumannii Strain MDR-ZJ06 Widely Spread in China.
<3>Antimicrob. Agents Chemother.
<4>55
<5>4506-4512
<6>2011
<7>We previously reported that the multidrug-resistant (MDR) Acinetobacter baumannii strain
MDR-ZJ06, belonging to European clone II, was widely
spread in China. In this study, we report the whole-genome sequence of
this clinically important strain. A 38.6-kb AbaR-type genomic
resistance island (AbaR22) was identified in MDR-ZJ06. AbaR22 has a
structure similar to those of the resistance islands found in A.
baumannii strains AYE and AB0057, but it contained only a few
antibiotic resistance genes. The region of resistant gene accumulation
as previously described was not found in AbaR22. In the chromosome of
the strain MDR-ZJ06, we identified the gene bla(oxa-23) in a composite
transposon (Tn2009). Tn2009 shared the backbone with other A. baumannii
transponsons that harbor bla(oxa-23), but it was bracketed by two
ISAba1 elements which were transcribed in the same orientation.
MDR-ZJ06 also expressed the armA gene on its plasmid pZJ06, and this
gene has the same genetic environment as the armA gene of the
Enterobacteriaceae. These results suggest variability of resistance
acquisition even in closely related A. baumannii strains.

<>

<1>Zhou, H.J., Purdy, M.M., Dahlquist, F.W., Reich, N.O.
<2>The Recognition Pathway for the DNA Cytosine Methyltransferase M.HhaI.
<3>Biochemistry
<4>48
<5>7807-7816
<6>2009
<7>Enzymatic sequence-specific DNA modification involves multiple poorly understood
intermediates. DNA methyltransferases like M.HhaI initially
bind nonspecific DNA and then selectively bind and modify a unique
sequence. High-resolution NMR was used to map conformational changes
occurring in M. HhaI upon binding nonspecific DNA, a one base pair
altered noncognate DNA sequence, and both hemimethylated and
unmethylated cognate DNA sequences. Comparisons with previous NMR
studies of the apoenzyme and enzyme-cofactor complex provide snapshots
of the pathway to sequence-specific complex formation. Dramatic
chemical shift perturbations reaching many distal sites within the
protein are detected with cognate DNA, while much smaller changes are
observed upon nonspecific and noncognate DNA binding. A cooperative
rather than stepwise transition from a nonspecific to a cognate complex
is revealed, Furthermore, switching from unmethylated to hemimethylated
cognate DNA involves delectable protein conformational changes 20-30
angstrom away from the methyl group, indicating high protein
sensitivity and plasticity to DNA modification.

<>

<1>Zhou, J., Sun, D., Childers, A., McDermott, T.R., Wang, Y., Liles, M.R.
<2>Three novel virophage genomes discovered from Yellowstone Lake metagenomes.
<3>J. Virol.
<4>0
<5>0
<6>2014
<7>Virophages are a unique group of circular double-stranded DNA viruses that are
considered parasites of giant DNA viruses, which in turn are known to infect
eukaryotic hosts. In this study the genomes of three novel virophages YSLV5,
YSLV6 and YSLV7 were identified from Yellowstone Lake through metagenomic
analyses. The relative abundance of these three novel virophages and previously
identified Yellowstone Lake virophages YSLVs 1-4 were determined in different
locations of the lake, revealing that most of the sampled locations in the lake,
including both mesophilic and thermophilic habitats, had multiple virophage
genotypes. This likely reflects the diverse habitats or diversity of the
eukaryotic hosts and their associated giant viruses that serve as putative hosts
for these virophages. YSLV5 has a 29,767 bp genome with 32 predicted ORFs, YSLV6
has a 24,837 bp genome with 29 predicted ORFs, and YSLV7 has a 23,193 bp genome
with 26 predicted ORFs. Based on multilocus phylogenetic analysis, YSLV6 shows a
close evolutionary relationship with YSLVs 1-4, whereas YSLV5 and YSLV7 are
distantly related to the others, and YSLV7 represents the fourth novel virophage
lineage. In addition, the genome of YSLV5 has a G+C content of 51.1% that is much
higher than all other known virophages, indicating a unique host range for YSLV5.
These results suggest that virophages are abundant and have diverse genotypes
that likely mirror diverse giant viral and eukaryotic hosts within the
Yellowstone Lake ecosystem. IMPORTANCE: This study discovered novel virophages
present within the Yellowstone Lake ecosystem using a conserved major capsid
protein as a phylogenetic anchor for assembly of sequence reads from Yellowstone
Lake metagenomic samples. The three novel virophage genomes (YSLV5-7) were
completed by identifying specific environmental samples containing these
respective virophages, and closing gaps by targeted PCR and sequencing. Most of
the YSLV genotypes were associated primarily with photic zone and
non-hydrothermal samples; however, YSLV5 had a unique distribution with an
occurrence in vent samples similar to that in photic zone samples, and with a
higher GC content that suggests a distinct host and habitat compared to other
YSLVs. In addition, genome content and phylogenetic analyses indicate that YSLV5
and YSLV7 are distinct from known virophages, and that additional
as-yet-uncharacterized virophages are likely present within the Yellowstone Lake
ecosystem.

<>

<1>Zhou, J., Xu, S.Y., Zhu, Z.
<2>Method of cloning and expressing BsmI restriction/modification system in E. coli.
<3>Japanese Patent Office
<4>JP 2002262889 A
<5>
<6>2002
<7>
<>

<1>Zhou, J., Zhu, Z., Xu, S.Y.
<2>Method for cloning and producing the BsmI restriction endonuclease in E. coli.
<3>US Patent Office
<4>US 6335190 B
<5>
<6>2002
<7>The present invention relates to recombinant DNA which encodes the BsmI restriction
endonuclease as well as BsmI methyltransferases, expression of BsmI restriction endonuclease
in E. coli cells containing the recombinant DNA by using a low copy number T7 expression
vector pACYC-T7ter, and purification of BsmI restriction endonuclease by heat treatment and
chromatography through heparin Sepharose column.

<>

<1>Zhou, L., Cheng, X., Connolly, B.A., Dickman, M.J., Hurd, P.J., Hornby, D.P.
<2>Zebularine: A novel DNA methylation inhibitor that forms a covalent complex with DNA methyltransferases.
<3>J. Mol. Biol.
<4>321
<5>591-599
<6>2002
<7>Mechanism-based inhibitors of enzymes, which mimic reactive intermediates in the reaction
pathway, have been deployed extensively in the analysis of metabolic pathways and as candidate
drugs. The inhibition of cytosine-[C5]-specific DNA methyltransferases (C5 MTases) by
oligodeoxynucleotides containing 5-azadeoxycytidine (AzadC) and 5-fluorodeoxycytidine (FdC)
provides a well-documented example of mechanism-based inhibition of enzymes central to nucleic
acid metabolism. Here, we describe the interaction between the C5 MTase from Haemophilus
haemolyticus (M.HhaI) and an oligodeoxynucleotide duplex containing 2-H pyrimidinone, an
analogue often referred to as zebularine and known to give rise to high-affinity complexes
with MTases. X-ray crystallography has demonstrated the formation of a covalent bond between
M.HhaI and the 2-H pyrimidinone-containing oligodeoxynucleotide. This observation enables a
comparison between the mechanisms of action of 2-H pyrimidinone with other mechanism-based
inhibitors such as FdC. This novel complex provides a molecular explanation for the mechanism
of action of the anti-cancer drug zebularine.

<>

<1>Zhou, L., Li, M., Yang, J., Wei, L., Ji, G.
<2>Draft Genome Sequence of Antagonistic Agent Lysobacter antibioticus 13-6.
<3>Genome Announcements
<4>2
<5>e00566-14
<6>2014
<7>Lysobacter antibioticus 13-6, isolated from the roots of Chinese cabbage, effectively controls
the pathogens Plasmodiophora brassicae, Xanthomonas oryzae
pv. oryzicola, X. oryzae pv. oryzae, Xanthomonas axonopodis pv. dieffenbachiae,
and Pseudomonas syringae pv. tabaci. We report the first draft genome sequence of
the L. antibioticus species in China.

<>

<1>Zhou, M., Chen, W., Chen, H., Wei, G.
<2>Draft Genome Sequence of Mesorhizobium alhagi CCNWXJ12-2T, a Novel Salt-Resistant Species Isolated from the Desert of Northwestern China.
<3>J. Bacteriol.
<4>194
<5>1261-1262
<6>2012
<7>Mesorhizobium alhagi strain CCNWXJ12-2(T) is a novel species of soil-dwelling, nitrogen-fixing
bacteria that can form symbiotic root nodules with Alhagi
sparsifolia. Moreover, the strain has high resistance to salt and alkali. Here we
report the draft genome sequence of Mesorhizobium alhagi strain CCNWXJ12-2(T). A
large number of osmotic regulation-related genes have been identified.

<>

<1>Zhou, S., Li, L., Wei, J., Qin, Q.
<2>Genome Sequence of Klebsiella pneumoniae HSL4, a New Strain Isolated from Mangrove Sediment for Biosynthesis of 1,3-Propanediol.
<3>Genome Announcements
<4>1
<5>e00177-13
<6>2013
<7>Klebsiella pneumoniae HSL4 is a 1,3-propanediol-producing bacterium strain isolated from
mangrove sediment. We present here a 5,221,448-bp assembly of its
genome sequence. Genome analysis revealed that it contains 10 coding sequences
(CDSs) responsible for glycerol fermentation to 1,3-propanediol, 19 CDSs encoding
glycerol utilization, and 140 CDSs related to its virulence.

<>

<1>Zhou, X., Deng, Z., Firmin, J.L., Hopwood, D.A., Kieser, T.
<2>Site-specific degradation of Streptomyces lividans DNA during electrophoresis in  buffers contaminated with ferrous iron.
<3>Nucleic Acids Res.
<4>16
<5>4341-4352
<6>1988
<7>Streptomyces lividans DNA contains a modification which makes it susceptible to double-strand
cleavage during electrophoresis in buffers contaminated with
ferrous iron (which may be present in some batches of EDTA). The cleavage of the
DNA is site-specific and the average fragment size resulting from limit digestion
of total S. lividans DNA is about 6kb. DNA from Streptomyces coelicolor A3(2) and
several other Streptomyces strains, and from E. coli, is not cleaved under the
same conditions. A S. lividans mutant has been isolated which lacks the DNA
modification. We suspect that many reports of 'poor' preparations of S. lividans
plasmids may be due to the above effect.

<>

<1>Zhou, X., He, X., Liang, J., Li, A., Xu, T., Kieser, T., Helmann, J.D., Deng, Z.
<2>A novel DNA modification by sulphur.
<3>Mol. Microbiol.
<4>57
<5>1428-1438
<6>2005
<7>Streptomyces lividans has a novel DNA modification, which sensitises its DNA to degradation
during electrophoresis (the Dnd phenotype). The entire
gene cluster (dnd) involved in this modification was localized on an 8 kb
DNA fragment and was expressed in a S. lividans deletion mutant (dnd) and
in several heterologous hosts. Disruption of the dnd locus abolishes the
Dnd phenotype, and gain of the dnd locus conferred the Dnd phenotype
respectively. Extensive analysis of the dnd gene cluster revealed five
open reading frames, whose hypothetic functions suggested an incorporation
of sulphur or a sulphur-containing substance into S. lividans genome, yet
in an unknown manner. The Dnd phenotype was also discovered to exist in
DNA of widespread bacterial species of variable origin and diverse
habitat. Similarly organized gene clusters were found in several bacterial
genomes representing different genera and in eDNA of marine organisms,
suggesting such modification as a widespread phenomenon. A coincidence
between the Dnd phenotype and DNA modification by sulphur was demonstrated
to occur in several representative bacterial genomes by the in
vivo(35)S-labelling experiments.

<>

<1>Zhou, X.E., Wang, Y., Reuter, M., Mackeldanz, P., Kruger, D.H., Meehan, E.J., Chen, L.Q.
<2>A single mutation of restriction endonuclease EcoRII led to a new crystal form that diffracts to 2.1 angstrom resolution.
<3>Acta Crystallogr. D Biol. Crystallogr.
<4>59
<5>910-912
<6>2003
<7>R88A, a mutant of the type IIE restriction endonuclease EcoRII, has been crystallized in space
group P2(1), with unit-cell parameters a = 58.7, b =
92.4, c = 88.3 A, beta = 108.1 degrees. There are two monomers in the
asymmetric unit and the solvent content is estimated to be 50% by volume.
The crystals diffract to 2.1 A resolution, which is much higher than that
of the wild type, which diffracted to 2.8 A resolution. The mutant
crystals have been used in the identification of an excellent heavy-atom
derivative.

<>

<1>Zhou, X.W., Li, J., Wang, K.M., Tan, W.H., Yang, X.H., Meng, X.X., Yan, H.F.
<2>An assay for the activity of DNA methylase based on a hairpin fluorescence molecular probe and its application in drug selection.
<3>Acta Chimi. Sin.
<4>64
<5>2096-2100
<6>2006
<7>A new activity assay for DNA methylase based on a hairpin molecular probe was proposed in this
paper. The recognition site of DNA methylase
and corresponding restriction endonuclease was designed in the stem
part of the probe, tetramethy1rhodamine (TAMRA) was attached at the 5'
terminus and its fluorescence was quenched by
4-(4'-dimethylaniinophenylazo)benzoic acid (DABCYL) linked at the 3'
terminus. The unmethylated probe can be cut in the recognition site by
restriction endonuclease, causing the restoration of the fluorescence
of TAMRA. Therefore, the activity of methylase would be analyzed
according to the degree of fluorescence restoration. Based on this
assay, the influence of anti-tumor drugs on the activity of methylase
was investigated by adding drugs in the methylation reaction. This
method can provide a potential in selecting proper drugs for tumors
caused by altered gene methylation pattern.

<>

<1>Zhou, X.Y.E., Wang, Y.J., Reuter, M., Mucke, M., Kruger, D.H., Meehan, E.J., Chen, L.Q.
<2>Crystal structure of Type IIE restriction endonuclease EcoRII reveals an autoinhibition mechanism by a novel effector-binding fold.
<3>J. Mol. Biol.
<4>335
<5>307-319
<6>2004
<7>EcoRII is a Type IIE restriction endonuclease that interacts with two copies of the DNA
recognition sequence 5'CCWGG, one being the actual
target of cleavage, the other serving as the allosteric effector. The
mode of enzyme activation by effector binding is unknown. To
investigate the molecular basis of activation and cleavage mechanisms
by EcoRII, the crystal structure of EcoRII mutant R88A has been
solved at 2.1 Angstrom resolution. The EcoRII monomer has two domains
linked through a hinge loop. The N-terminal effector-binding domain has
a novel DNA recognition fold with a prominent cleft. The C-terminal
catalytic domain has a restriction endonuclease-like fold.
Structure-based sequence alignment identified the putative catalytic
site of EcoRII that is spatially blocked by the N-terminal domain. The
structure together with the earlier characterized EcoRII enzyme
activity enhancement in the absence of its N-terminal domain reveal an
autoinhibition/activation mechanism of enzyme activity mediated by a
novel effector-binding fold. This is the first case of autoinhibition,
a mechanism described for many transcription factors and signal
transducing proteins, of a restriction endonuclease.

<>

<1>Zhou, Y., Li, R., Gao, X.Y., Lapidus, A., Han, J., Haynes, M., Lobos, E., Huntemann, M., Pati, A., Ivanova, N.N., Rohde, M., Mavromatis, K., Tindall, B.J., Markowitz, V., Woyke, T., Klenk, H.P., Kyrpides, N.C., Li, W.J.
<2>High quality draft genome sequence of the slightly halophilic bacterium Halomonas zhanjiangensis type strain JSM 078169(T) (DSM 21076(T)) from a sea urchin in  southern China.
<3>Standards in Genomic Sciences
<4>9
<5>1020-1030
<6>2014
<7>Halomonas zhanjiangensis Chen et al. 2009 is a member of the genus Halomonas, family
Halomonadaceae, class Gammaproteobacteria. Representatives of the genus
Halomonas are a group of halophilic bacteria often isolated from salty
environments. The type strain H. zhanjiangensis JSM 078169(T) was isolated from a
sea urchin (Hemicentrotus pulcherrimus) collected from the South China Sea. The
genome of strain JSM 078169(T) is the fourteenth sequenced genome in the genus
Halomonas and the fifteenth in the family Halomonadaceae. The other thirteen
genomes from the genus Halomonas are H. halocynthiae, H. venusta, H. alkaliphila,
H. lutea, H. anticariensis, H. jeotgali, H. titanicae, H. desiderata, H.
smyrnensis, H. salifodinae, H. boliviensis, H. elongata and H stevensii. Here, we
describe the features of strain JSM 078169(T), together with the complete genome
sequence and annotation from a culture of DSM 21076(T). The 4,060,520 bp long
draft genome consists of 17 scaffolds with the 3,659 protein-coding and 80 RNA
genes and is a part of Genomic Encyclopedia of Type Strains, Phase I: the one
thousand microbial genomes (KMG) project.

<>

<1>Zhou, Z., Li, X., Liu, B., Beutin, L., Xu, J., Ren, Y., Feng, L., Lan, R., Reeves, P.R., Wang, L.
<2>Derivation of Escherichia coli O157:H7 from its O55:H7 precursor.
<3>PLoS ONE
<4>5
<5>E8700
<6>2010
<7>There are 29 E. coli genome sequences available, mostly related to studies
of species diversity or mode of pathogenicity, including two genomes of
the well-known O157:H7 clone. However, there have been no genome studies
of closely related clones aimed at exposing the details of evolutionary
change. Here we sequenced the genome of an O55:H7 strain, closely related
to the major pathogenic O157:H7 clone, with published genome sequences,
and undertook comparative genomic and proteomic analysis. We were able to
allocate most differences between the genomes to individual mutations,
recombination events, or lateral gene transfer events, in specific
lineages. Major differences include a type II secretion system present
only in the O55:H7 chromosome, fewer type III secretion system effectors
in O55:H7, and 19 phage genomes or phagelike elements in O55:H7 compared
to 23 in O157:H7, with only three common to both. Many other changes were
found in both O55:H7 and O157:H7 lineages, but in general there has been
more change in the O157:H7 lineages. For example, we found 50% more
synonymous mutational substitutions in O157:H7 compared to O55:H7. The two
strains also diverged at the proteomic level. Mutational synonymous SNPs
were used to estimate a divergence time of 400 years using a new clock
rate, in contrast to 14,000 to 70,000 years using the traditional clock
rates. The same approaches were applied to three closely related
extraintestinal pathogenic E. coli genomes, and similar levels of mutation
and recombination were found. This study revealed for the first time the
full range of events involved in the evolution of the O157:H7 clone from
its O55:H7 ancestor, and suggested that O157:H7 arose quite recently. Our
findings also suggest that E. coli has a much lower frequency of
recombination relative to mutation than was observed in a comparable study
of a Vibrio cholerae lineage.

<>

<1>Zhou, Z., Peng, X., Xiao, Y., Wang, X., Guo, Z., Zhu, L., Liu, M., Jin, H., Bi, D., Li, Z., Sun, M.
<2>Genome Sequence of the poultry pathogen Riemerella anatipestifer Strain RA-YM.
<3>J. Bacteriol.
<4>193
<5>1284-1285
<6>2010
<7>Riemerella anatipestifer is a Gram-negative rod-shaped bacillus associated with epizootic
infections in poultry. R. anatipestifer strain RA-YM, belonging to the serotype 1 prevalent in
China, is a clinically isolated strain with high-level virulence. Here, we report the first
genome sequence of this species.

<>

<1>Zhu, B., Chen, M., Lin, L., Yang, L., Li, Y., An, Q.
<2>Genome sequence of Enterobacter sp. strain SP1, an endophytic nitrogen-fixing bacterium isolated from sugarcane.
<3>J. Bacteriol.
<4>194
<5>6963-6964
<6>2012
<7>Enterobacter sp. strain SP1 is an endophytic nitrogen-fixing bacterium isolated
from a sugarcane stem and can promote plant growth. The draft genome sequence of
strain SP1 presented here will promote comparative genomic studies to determine
the genetic background of interactions between endophytic enterobacteria and
plants.

<>

<1>Zhu, B., Liu, H., Tian, W.X., Fan, X.Y., Li, B., Zhou, X.P., Jin, G.L., Xie, G.L.
<2>Genome Sequence of Stenotrophomonas maltophilia RR-10, Isolated as an Endophyte from Rice Root.
<3>J. Bacteriol.
<4>194
<5>1280-1281
<6>2012
<7>Stenotrophomonas maltophilia is an endophyte which plays important roles in agricultural
production as a plant growth-promoting bacterium. Here, we present
the draft genome sequence of strain RR-10, which was isolated from a rice root in
a rice field of China.

<>

<1>Zhu, B., Ye, S., Chang, S., Chen, M., Sun, L., An, Q.
<2>Genome Sequence of the Pathogenic Herbaspirillum seropedicae Strain Os45, Isolated from Rice Roots.
<3>J. Bacteriol.
<4>194
<5>6995-6996
<6>2012
<7>Most Herbaspirillum seropedicae strains are beneficial to plants. In contrast, H. seropedicae
strain Os45, isolated from rice roots, is pathogenic. The draft
genome sequence of strain Os45 presented here allows an in-depth comparative
genome analysis to understand the subtle mechanisms of beneficial and pathogenic
Herbaspirillum-plant interactions.

<>

<1>Zhu, D., Li, P., Tanabe, S.H., Sun, J.
<2>Genome Sequence of the Alkaliphilic Bacterial Strain Bacillus ligninesis L1, a Novel Degrader of Lignin.
<3>Genome Announcements
<4>1
<5>e0004213
<6>2013
<7>Bacillus ligninesis strain L1, isolated from seafloor sediment, was able to grow  on medium
with lignin as its sole carbon source. Here, we report a 3.8-Mbp
high-quality genome sequence for this bacterium. The genes involving ectoine and
glycine betaine synthesis, as well as those involved in the degradation of
lignin, were identified.

<>

<1>Zhu, H., Wang, Q., Wen, L., Xu, J., Shao, Z., Chen, M., Chen, M., Reeves, P.R., Cao, B., Wang, L.
<2>Development of a Multiplex PCR Assay for Detection and Genogrouping of Neisseria meningitidis.
<3>J. Clin. Microbiol.
<4>50
<5>46-51
<6>2012
<7>Neisseria meningitidis is a leading pathogen of epidemic bacterial meningitis and
fulminant sepsis worldwide. Twelve different N. meningitidis serogroups have been
identified to date based on antigenic differences in the capsular polysaccharide.
However, more than 90% of human cases of N. meningitidis meningitis are the
result of infection with just five serogroups, A, B, C, W135, and Y. Efficient
methods of detection and genogrouping of N. meningitidis isolates are needed,
therefore, in order to monitor prevalent serogroups as a means of disease control
and prevention. The capsular gene complex regions have been sequenced from only
seven out of the 12 serogroups. In this study, the capsular gene complexes of the
remaining five serogroups were sequenced and analyzed. Primers were designed that
were specific for N. meningitidis species and for the 12 individual serogroups,
and a multiplex PCR assay using these specific primers was developed for N.
meningitidis detection and genogrouping. The assay was tested using 15 reference
strains covering all 12 serogroups, 143 clinical isolates, and 21 strains from
closely related species or from species that cause meningitis. The assay could
detect N. meningitidis serogroups and was shown to be specific, with a detection
sensitivity of 1 ng of genomic DNA (equivalent to approximately 4 x 10(5)
genomes) or 3 x 10(5) CFU/ml in noncultured mock cerebrospinal fluid (CSF)
specimens. This study, therefore, describes for the first time the development of
a molecular protocol for the detection of all N. meningitidis serogroups. This
multiplex PCR-based assay may have use for the clinical diagnosis and
epidemiological surveillance of N. meningitidis.

<>

<1>Zhu, J.
<2>Susceptibility of AatII and ApaI 3' protruding ends to exonuclease III degradation.
<3>Biotechniques
<4>11
<5>757-759
<6>1991
<7>None

<>

<1>Zhu, L., Yan, Z., Zhang, Z., Zhou, Q., Zhou, J., Wakeland, E.K., Fang, X., Xuan, Z., Shen, D., Li, Q.Z.
<2>Complete Genome Analysis of Three Acinetobacter baumannii Clinical Isolates in China for Insight into the Diversification of Drug Resistance Elements.
<3>PLoS ONE
<4>8
<5>E66584
<6>2013
<7>BACKGROUND: The emergence and rapid spreading of multidrug-resistant
Acinetobacter baumannii strains has become a major health threat worldwide. To
better understand the genetic recombination related with the acquisition of
drug-resistant elements during bacterial infection, we performed complete genome
analysis on three newly isolated multidrug-resistant A. baumannii strains from
Beijing using next-generation sequencing technology. METHODOLOGIES/PRINCIPAL
FINDINGS: Whole genome comparison revealed that all 3 strains share some common
drug resistant elements including carbapenem-resistant bla OXA-23 and
tetracycline (tet) resistance islands, but the genome structures are diversified
among strains. Various genomic islands intersperse on the genome with transposons
and insertions, reflecting the recombination flexibility during the acquisition
of the resistant elements. The blood-isolated BJAB07104 and ascites-isolated
BJAB0868 exhibit high similarity on their genome structure with most of the
global clone II strains, suggesting these two strains belong to the dominant
outbreak strains prevalent worldwide. A large resistance island (RI) of about
121-kb, carrying a cluster of resistance-related genes, was inserted into the
ATPase gene on BJAB07104 and BJAB0868 genomes. A 78-kb insertion element carrying
tra-locus and bla OXA-23 island, can be either inserted into one of the tniB gene
in the 121-kb RI on the chromosome, or transformed to conjugative plasmid in the
two BJAB strains. The third strains of this study, BJAB0715, which was isolated
from spinal fluid, exhibit much more divergence compared with above two strains.
It harbors multiple drug-resistance elements including a truncated AbaR-22-like
RI on its genome. One of the unique features of this strain is that it carries
both bla OXA-23 and bla OXA-58 genes on its genome. Besides, an Acinetobacter
lwoffii adeABC efflux element was found inserted into the ATPase position in
BJAB0715. CONCLUSIONS: Our comparative analysis on currently completed
Acinetobacter baumannii genomes revealed extensive and dynamic genome
organizations, which may facilitate the bacteria to acquire drug-resistance
elements into their genomes.

<>

<1>Zhu, P., Morelli, G., Achtman, M.
<2>The opcA and (psi)opcB regions in Neisseria: genes, pseudogenes, deletions, insertion elements and DNA islands.
<3>Mol. Microbiol.
<4>33
<5>635-650
<6>1999
<7>Previous data have indicated that the opc gene encoding an immunogenic invasin is specific to
Neisseria meningitidis (Nm) and is lacking in
Neisseria gonorrhoeae (Ng). The data presented here show that Nm and Ng
both contain two paralogous opc-like genes, opcA, corresponding to the
former opc gene, and (psi)opcB, a pseudogene. The predicted OpcA and OpcB
proteins possess transmembrane regions with conserved non-polar faces but
differ extensively in four of the five surface-exposed loops. Gonococcal
OpcA was expressed weakly under in vitro conditions, and it is unknown
whether these bacteria can express this protein at high levels. Analysis
of the sequences flanking opcA and (psi)opcB revealed a framework of
conserved housekeeping genes interspersed with DNA islands. These regions
also contained several pseudogenes, deletions and IS elements, attesting
to considerable genome plasticity. Both opcA and (psi)opcB are located on
DNA islands that have probably been imported from unrelated bacteria. A
third island encodes the dcmD/dcrD R/M genes in Ng versus a small open
reading frame in most strains of Nm. Rare strains of Nm were identified in
which the R/M island has been imported. DNA islands in Nm and Ng seem to
have been acquired by recombination via conserved flanking housekeeping
genes rather than by insertion of mobile genetic elements.

<>

<1>Zhu, S., Wang, H.L., Wang, C., Tang, L., Wang, X., Yu, K.J., Liu, S.L.
<2>Non-contiguous finished genome sequence and description of Salmonella enterica subsp. houtenae str. RKS3027.
<3>Standards in Genomic Sciences
<4>8
<5>198-205
<6>2013
<7>Salmonella enterica subsp. houtenae serovar 16:z4, z32:-- str. RKS3027 was isolated from a
human in Illinois, USA. S. enterica subsp. houtenae is a
facultative aerobic rod-shaped Gram-negative bacterium. Here we describe the
features of this organism, together with the draft genome sequence and
annotation. The 4,404,136 bp long genome (97 contigs) contains 4,335
protein-coding gene and 28 RNA genes.

<>

<1>Zhu, W., Huang, J., Li, M., Li, X., Wang, G.
<2>Genomic analysis of Skermanella stibiiresistens type strain SB22 (T.).
<3>Standards in Genomic Sciences
<4>9
<5>1211-1220
<6>2014
<7>Members of genus Skermanella were described as Gram-negative, motile, aerobic, rod-shaped,
obligate-heterotrophic bacteria and unable to fix nitrogen. In this
study, the genome sequence of Skermanella stibiiresistens SB22(T) is reported.
Phylogenetic analysis using core proteins confirmed the phylogenetic assignment
based on 16S rRNA gene sequences. Strain SB22(T) has all the proteins for
complete glycolysis, tricarboxylic acid cycle and pentose phosphate pathway. The
RuBisCO encoding genes cbbL1S1 and nitrogenase delta subunit gene anfG are
absent, consistent with its inability to fix carbon and nitrogen, respectively.
In addition, the genome possesses a series of flagellar assembly and chemotaxis
genes to ensure its motility.

<>

<1>Zhu, Y., Shang, H., Zhu, Q., Ji, F., Wang, P., Fu, J., Deng, Y., Xu, C., Ye, W., Zheng, J., Zhu, L., Sun, M.
<2>Complete Genome Sequence of Bacillus thuringiensis serovar. finitimus Strain YBT-020.
<3>J. Bacteriol.
<4>193
<5>2379-2380
<6>2011
<7>Bacillus thuringiensis is a Gram-positive, spore-forming bacterium that forms parasporal
crystals at the onset of the sporulation phase of its growth. Here we report the complete
genome sequence of B. thuringiensis serovar. finitimus strain YBT-020, whose parasporal
crystals consist of Cry26Aa and Cry28Aa crystal proteins and are located between the
exosporium and the spore coat and remain adhering to the spore after sporulation.

<>

<1>Zhu, Y., Zeng, H., Gao, X., Lu, Z.H.
<2>Quantitative analysis of EcoR1 methylase-DNA complex by atomic force microscopy.
<3>Scanning
<4>28
<5>15-19
<6>2006
<7>The EcoRI methylase specifically recognize 5'-GA*ATTC-3' in DNA duplex.  We directly applied
atomic force microscopy to investigate linear pBR322-EcoRI methylase complexes and
quantitatively analyzed the bend angles of linear pBR322-EcoRI methylse complexes and the
bound protein widths.  In this study, we made a novel observation that DNA-EcoRI methylase
complexes exhibited two populations of conformation at the recognition site: DNA was bent at
an acute
angle at the recognition site in the presence of one EcoRI methylase monomeric molecule, while
DNA was bent at an obtuse angle at the recognition site and the complementary site on duplex
DNAs in
the presence of EcoRI methylase dimer.  The data indicated that the obtuse angle state was
the result of unique interactions between EcoRI methylase and the recognition site and the
complementary site on duplex DNAs, and suggested that the acute angle conformation could be an
intermediate in the formation of the obtuse angle state.  Our works provide a detail insight
into the DNA structural variations involved in EcoRI methylase-binding processes and
demonstrate further the versatility of AFM as an imaging technique for studying the
interaction between large DNA fragment and protein.

<>

<1>Zhu, Z., Benner, J., Xu, S.-Y.
<2>Cloning, sequence and expression of NruI and Sbo13I restriction endonucleases and modification methylases from Nocardia rubra.
<3>International Patent Office
<4>WO 2007120541 A
<5>
<6>2007
<7>Recombinant DNA encoding NtruI- and SboI-like restriction endonucleases and methylases and
their amino acid sequences are provided as well as methods for expressing the enzymes in
transformed host cells and purifying the enzymes.

<>

<1>Zhu, Z., Benner, J.S., Xu, S.-Y.
<2>Polynucleotides encoding polypeptides and host cells therefor.
<3>US Patent Office
<4>US 08361773
<5>
<6>2013
<7>Recombinant DNA encoding NruI- and SboI-like restriction endonucleases and methylases and
their amino acid sequences are provided as well as
methods for expressing the enzymes in transformed host cells and
purifying the enzymes.

<>

<1>Zhu, Z., Blanchard, A., Xu, S.-Y., Guan, S., Wei, H., Zhang, P., Sun, D., Chan, S.-h.
<2>High fidelity restriction endonucleases.
<3>International Patent Office
<4>WO 2009009797
<5>
<6>2009
<7>Compositions and methods are provided for enzymes with altered properties that involve a
systematic approach to mutagenesis and a screening assay that permits selection of the desired
proteins.  Embodiments of the method are particularly suited for modifying specific properties
of restriction endonucleases such as star activity.  The compositions include restriction
endonucleases with reduced star activity as defined by an overall fidelity index improvement
factor.

<>

<1>Zhu, Z., Blanchard, A., Xu, S.-Y., Guan, S., Wei, H., Zhang, P., Sun, D., Chan, S.-H.
<2>High fidelity restriction endonucleases.
<3>US Patent Office
<4>US 08372619
<5>
<6>2013
<7>Compositions and methods are provided for enzymes with altered properties that involve a
systematic approach to mutagenesis and a
screening assay that permits selection of the desired proteins.
Embodiments of the method are particularly suited for modifying
specific properties of restriction endonucleases such as star activity.
The compositions include restriction endonucleases with reduced star
activity as defined by an overall fidelity index improvement factor.

<>

<1>Zhu, Z., Blanchard, A., Xu, S.-Y., Guan, S., Wei, H., Zhang, P., Sun, D., Chang, S.-H.
<2>Engineering of high-fidelity restriction endonucleases with reduced star activity.
<3>International Patent Office
<4>WO 2009009797 A
<5>
<6>2009
<7>Compositions and methods are provided for enzymes with altered properties that involve a
systematic approach to mutagenesis and a screening assay that permits selection of the desired
proteins.  Embodiments of the method are particularly suited for modifying specific properties
of restriction endonucleases such as star activity.  The compositions include restriction
endonucleases with reduced star activity as defined by an overall fidelity index improvement
factor.

<>

<1>Zhu, Z., Chan, S., Sun, D., Zhang, P., Wei, H., Guan, S., Blanchard, A., Xu, S.
<2>High Fidelity Restriction Endonucleases.
<3>Japanese Patent Office
<4>JP 2010532999 A
<5>
<6>2010
<7>
<>

<1>Zhu, Z., Guan, S., Blanchard, A.
<2>Engineering of a Bacillus stearothermophilus methylation-specific restriction endonuclease BstNI variant.
<3>International Patent Office
<4>WO 2010104737 A
<5>
<6>2010
<7>A restriction endonuclease is provided that has been engineered to have a cleavage specificity
for a DNA recognition sequence containing a modified nucleotide.  Methods for engineering
enzymes to cleave DNA containing modified nucleotides at specific sequences are provided.

<>

<1>Zhu, Z., Nagaraja, V., Saravanan, M.
<2>Engineering of KpnI restriction endonuclease variants with reduced star activity.
<3>International Patent Office
<4>WO 200727464 A
<5>
<6>2007
<7>Methods are provided for making restriction endonucleases with reduced star activity by one or
more targeted mutations to a catalytic site within the restriction endonuclease.  Examples of
modifications to restriction endonucleases with significant sequence identity with KpnI are
provided and reduced star activity demonstrated.

<>

<1>Zhu, Z., Pedamallu, C.S., Fomenkov, A., Benner, J., Xu, S.Y.
<2>Cloning of NruI and Sbo13I restriction and modification systems in E. coli and amino acid sequence comparison of M.NruI and M.Sbo13I with other amino-methyltransferases.
<3>BMC Res. Notes
<4>3
<5>139
<6>2010
<7>ABSTRACT: BACKGROUND: NruI and Sbo13I are restriction enzyme isoschizomers
with the same recognition sequence 5' TCG downward arrowCGA 3' (cleavage
as indicated downward arrow). Here we report the cloning of NruI and
Sbo13I restriction-modification (R-M) systems in E. coli. The NruI
restriction endonuclease gene (nruIR) was cloned by PCR and inverse PCR
using primers designed from the N-terminal amino acid sequence. The NruI
methylase gene (nruIM) was derived by inverse PCR walking. RESULTS: The
amino acid sequences of NruI endonuclease and methylase are very similar
to the Sbo13I R-M system which has been cloned and expressed in E. coli by
phage selection of a plasmid DNA library. Dot blot analysis using rabbit
polyclonal antibodies to N6mA- or N4mC-modified DNA indicated that M.NruI
is possibly a N6mA-type amino-methyltransferase that most likely modifies
the external A in the 5' TCGCGA 3' sequence. M.Sbo13I, however, is
implicated as a probable N4mC-type methylase since plasmid carrying
sbo13IM gene is not restricted by Mrr endonuclease and Sbo13I digestion is
not blocked by Dam methylation of the overlapping site. The amino acid
sequence of M.NruI and M.Sbo13I did not show significant sequence
similarity to many known amino-methyltransferases in the alpha, beta, and
gamma groups, except to a few putative methylases in sequenced microbial
genomes. CONCLUSIONS: The order of the conserved amino acid motifs
(blocks) in M.NruI/M.Sbo13I is similar to the gamma. group
amino-methyltranferases, but with two distinct features: In motif IV, the
sequence is DPPY instead of NPPY; there are two additional conserved
motifs, IVa and Xa as extension of motifs IV and X, in this family of
enzymes. We propose that M.NruI and M.Sbo13I form a subgroup in the gamma
group of amino-methyltransferases.

<>

<1>Zhu, Z., Quimby, A., Guan, S., Sun, D., Huang, Y., Lai, X., Chan, S.-H., Li, X., Xu, S.-Y., Zhang, C.
<2>High fidelity restriction endonucleases.
<3>US Patent Office
<4>US 20110151450 A
<5>
<6>2011
<7>Methods and compositions are provided for engineering mutant enzymes with reduced star
activity where the mutant enzymes have a fidelity index in a specified buffer that is greater
than the FI of the non-mutated enzyme in the same buffer.

<>

<1>Zhu, Z., Robinson, D., Benner, J., Xu, S.-Y.
<2>Method for cloning and expression of Tth111II restriction endonuclease-methylase in E. coli.
<3>US Patent Office
<4>US 20040132129 A
<5>31
<6>2004
<7>The present invention relates to recombinant DNA which encodes the Tth111II restriction
endonuclease-methylase fusion protein (Tth111IIRM), expression of Tth111II restriction
endonuclease-methylase fusion protein in E. coli cells containing the recombinant DNA, and
purification of Tth111II endonuclease-methylase fusion protein to near homogeneity.

<>

<1>Zhu, Z., Robinson, D., Benner, J., Xu, S.-y.
<2>Method for cloning and expression of Tth111II restriction endonuclease-methylase in E. coli.
<3>US Patent Office
<4>US 6919194 B
<5>
<6>2005
<7>The present invention relates to recombinant DNA which encodes the Tth111II restriction
endonuclease-methylase fusion protein (Tth111IIRM), expression of Tth111II restriction
endonuclease-methylase fusion protein in E. coli cells containing the recombinant DNA, and
purification of Tth111II endonuclease-methylase fusion protein to near homogeneity.

<>

<1>Zhu, Z., Samuelson, J.C., Zhou, J., Dore, A., Xu, S.-y.
<2>Engineering strand-specific DNA nicking enzymes from the type IIS restriction endonucleases BsaI, BsmBI, and BsmAI.
<3>J. Mol. Biol.
<4>337
<5>573-583
<6>2004
<7>More than 80 type IIA/IIS restriction endonucleases with different recognition specificities
are now known. In contrast, only a limited number of strand-specific nicking endonucleases are
currently available. To overcome this limitation, a novel genetic screening method was devised
to convert type IIS restriction endonucleases into strand-specific nicking endonucleases. The
genetic screen consisted of four steps: (1) random mutagenesis to create a plasmid library,
each bearing an inactivated endonuclease gene; (2) restriction digestion of plasmids
containing the wild-type and the mutagenized endonuclease gene; (3) back-crosses with the
wild-type gene by ligation to the wild-type N-terminal or C-terminal fragment; (4)
transformation of the ligated DNA into a pre-modified host and screening for nicking
endonuclease activity in total cell culture or cell extracts of the transformants. Nt.BsaI and
Nb.BsaI nicking endonucleases were isolated from BsaI using this genetic screen. In addition,
site-directed mutagenesis was carried out to isolate BsaI nicking variants with minimal
double-stranded DNA cleavage activity. The equivalent amino acid substitutions were introduced
into BsmBI and BsmAI restriction endonucleases with similar recognition sequence and
significant amino acid sequence identity and their nicking variants were successfully
isolated. This work provides strong evidence that some type IIS restriction endonucleases
carry two separate active sites. When one of the active sites is inactivated, the type IIS
restriction endonuclease may nick only one strand.

<>

<1>Zhu, Z., Xu, S.
<2>Engineering a strand-specific nicking endonuclease, comprises transforming a first host cell population lacking methylase protection, with plasmids containing a randomly mutagenized restriction endonuclease gene - recombinant endonuclease production.
<3>US Patent Office
<4>US 20050136462
<5>
<6>2005
<7>DERWENT ABSTRACT: NOVELTY - Engineering a strand-specific nicking endonuclease comprises
transforming a first host cell population
lacking methylase protection, with plasmids containing a randomly
mutagenized restriction endonuclease gene. DETAILED DESCRIPTION -
Engineering a strand-specific nicking endonuclease comprises: (a)
transforming a first host cell population lacking methylase protection,
with plasmids containing a randomly mutagenized restriction
endonuclease gene; (b) culturing the transformed host cell and
isolating the plasmids from it; (c) cleaving the mutagenized
restriction endonuclease gene and a corresponding wild-type restriction
endonuclease gene into fragments; (d) performing an in vitro backcross
between the wild-type and mutagenized restriction endonuclease
fragments and obtaining a ligated gene; (e) detecting a strand-specific
nicking activity of a protein expressed by the ligated gene; and (f)
identifying the engineered strand-specific nicking endonuclease.
INDEPENDENT CLAIMS are included for the following: (1) a nicking
endonuclease made according to the new method; (2) a nicking
endonuclease comprising a modified recombinant BsaI, BsmAI, or BsmBI;
(3) introducing one or more site-specific nicks into pre-selected
strands of a DNA duplex, comprising digesting the DNA duplex with the
nicking endonuclease made under conditions permitting nicking
activity;(4) amplifying a target sequence comprising: (a) providing a
single-stranded nucleic acid fragment containing the target sequence,
the fragment having a 5' end and a 3' end; (b) binding an amplification
primer for SDA to the 3' end of the fragment so that the primer forms a
5' single-stranded overhang, the amplification primer comprising a
recognition/cleavage site for the synthetic nicking endonuclease; (c)
extending the amplification primer on the fragment in the absence of a
derivatized or substituted deoxynucleoside triphosphate and in the
presence of a DNA polymerase having strand-displacing activity and
lacking 5'-3' exonuclease activity, and four deoxynucleoside
triphosphates; and (d) nicking the amplified double-stranded target
sequence with the nicking endonuclease extending from the nick using
the DNA polymerase, thus displacing the first newly synthesized strand
from the fragment and generating a second extension product comprising
a second newly synthesized strand; and (e) repeating the nicking,
extending and displacing steps so that the target sequence is
amplified; (5) engineering an enzyme with at least one of a modified
substrate specificity and activity, comprising: (a) forming a randomly
mutagenized DNA library where the library has one or more genes
encoding whole or part of a mutant enzyme, the mutant enzyme being
substantially inactive, the substantially inactive enzyme having an
N-terminal end and a C-terminal end, where the inactivation results
from a mutation in the N-terminal end or C-terminal end of a wild-type
enzyme; (b) cleaving the one or more genes expressing the inactive
endonuclease into at least a first fragment and a second fragment,
where the first fragment encodes the C-terminal end of the enzyme and
the second fragment encodes the N-terminal end of the enzyme; (c)
performing a ligation between fragments selected from the first
fragment and a third fragment encoding an N-terminal end of the
wild-type enzyme, the second fragment with a fourth fragment encoding
the C-terminal end of the wild-type enzyme, or both first and second
fragments to third and fourth fragments respectively; and (d)
expressing the ligated DNA in a host cell to obtain the enzyme having
modified substrate specificity and activity; (6) amplifying a target
nucleic acid, comprising: (a) nicking at least one strand of a
double-stranded target nucleic acid at several sites with the nicking
enzyme to form at least two new 3' termini; (b) extending one or more
of the at least two new 3' termini with a DNA polymerase; (c) nicking
the extension product; and (d) extending the nicking product.

<>

<1>Zhu, Z., Xu, S.-Y.
<2>Method for cloning and expression of AsiSI restriction endonuclease and AsiSI methylase in E. coli.
<3>US Patent Office
<4>US 6514737
<5>
<6>2003
<7>The present invention relates to recombinant DNA which encodes the AsiSI restriction
endonuclease as well as AsiSI methylase, expression of AsiSI
restriction endonuclease and AsiSI methylase in E. coli cells containing
the recombinant DNA.

<>

<1>Zhu, Z., Xu, S.-Y.
<2>A Method for cloning and expression of BsaI restriction endonuclease and BsaI methylase in E. coli.
<3>US Patent Office
<4>US 20030186363 A
<5>
<6>2003
<7>The present invention relates to recombinant DNA which encodes the BsaI restriction
endonuclease as well as BsaI methylase, expression of BsaI restriction endonuclease and BsaI
methylase in E. coli cells containing the recombinant DNA, and purification of BsaI
restriction endonuclease to near homogeneity.

<>

<1>Zhu, Z., Xu, S.-Y.
<2>A Method for cloning and expression of BsaI restriction endonuclease and BsaI methylase in E. coli.
<3>European Patent Office
<4>EP 1350849 A
<5>
<6>2003
<7>The present invention relates to recombinant DNA which encodes the BsaI restriction
endonuclease as well as BsaI methylase, expression of BsaI restriction endonuclease and BsaI
methylase in E. coli cells containing the recombinant DNA, and purification of BsaI
restriction endonuclease to near homogeneity.

<>

<1>Zhu, Z., Xu, S.-Y.
<2>Method for cloning and expression of BsaI restriction endonuclease and BsaI methylase in E. coli.
<3>US Patent Office
<4>US 6723546 B
<5>
<6>2004
<7>The present invention relates to recombinant DNA which encodes the BsaI restriction
endonuclease as well as BsaI methylase, expression of BsaI restriction endonuclease and BsaI
methylase in E. coli cells containing the recombinant DNA, and purification of BsaI
restriction endonuclease to near homogeneity.

<>

<1>Zhu, Z., Xu, S.-Y.
<2>Method for cloning and expression of AsiSI restriction endonuclease and AsiSI methylase in E. coli.
<3>International Patent Office
<4>WO 03016519 A
<5>
<6>2003
<7>The present invention relates to recombinant DNA which encodes the AsiSI restriction
endonuclease as well as AsiSI methylase, expression of AsiSI restriction endonuclease and
AsiSI methylase in E. coli cells containing the recombinant DNA.

<>

<1>Zhu, Z., Xu, S.-Y.
<2>Method for cloning and expression of StuI restriction endonuclease and StuI methylase in E. coli.
<3>US Patent Office
<4>US 08058029
<5>
<6>2011
<7>The present invention relates to compositions including: (1) isolated DNA encoding the StuI
restriction endonuclease and isolated DNA encoding cognate and non-cognate methylase; (2)
vectors and cells containing the isolated DNA; and (3) methods for producing the StuI
restriction endonuclease.

<>

<1>Zhu, Z., Xu, S.Y.
<2>Method for cloning and expression of BsaI restriction endonuclease and BsaI methylase in E. coli.
<3>Japanese Patent Office
<4>JP 2005034001 A
<5>
<6>2005
<7>
<>

<1>Zhu, Z., Zhou, J., Xu, S.-Y.
<2>Method for cloning and expression of BsmAI restriction endonuclease and BsmAI methylase in E. coli.
<3>US Patent Office
<4>US 6596524 B
<5>
<6>2003
<7>The present invention relates to recombinant DNA which encodes the BsmAI restriction
endonuclease as well as BsmAI methylase, expression of BsmAI restriction endonuclease and
BsmAI methylase in E. coli cells containing the recombinant DNA, and purification of BsmAI
endonuclease to near homogeneity.

<>

<1>Zhu, Z., Zhou, J., Xu, S.-Y.
<2>Cloning, sequencing and expression in E. coli of BsmAI restriction endonuclease and BsmAI methylase from Bacillus stearothermophilus.
<3>US Patent Office
<4>US 20030104388
<5>27
<6>2003
<7>The present invention relates to recombinant DNA which encodes the BsmAI restriction
endonuclease as well as BsmAI methylase, expression of BsmAI restriction endonuclease and
BsmAI methylase in E. coli cells containing the recombinant DNA, and purification of BsmAI
endonuclease to near homogeneity.

<>

<1>Zhu, Z.Y., Zhou, J., Friedman, A.M., Xu, S.Y.
<2>Isolation of BsoBI restriction endonuclease variants with altered substrate specificity.
<3>J. Mol. Biol.
<4>330
<5>359-372
<6>2003
<7>BsoBI is a thermophilic restriction endonuclease that cleaves the degenerate DNA sequence
C/PyCGPuG (where/=the cleavage site and Py=C or T,
Pu=A or G). In the BsoBI-DNA co-crystal structure the D246 residue makes a
water-mediated hydrogen bond to N6 of the degenerate base adenine and was
proposed to make a complementary bond to O6 of the alternative guanine
residue. To investigate the substrate specificity conferred by D246 and to
potentially alter BsoBI specificity, the D246 residue was changed to the
other 19 amino acids. Variants D246A, D246C, D246E, D246R, D246S, D246T,
and D246Y were purified and their cleavage activity determined. Variants
D246A, D246S, and D246T display 0.2% to 0.7% of the wild-type cleavage
activity. However, the substrate specificity of the three variants is
altered significantly. D246A, D246S, and D246T cleave CTCGAG sites poorly.
In filter binding assays using oligonucleotides, wild-type BsoBI shows
almost equal affinity for CTCGAG and CCCGGG sites. In contrast, the D246A
variant shows 70-fold greater binding affinity for the CCCGGG substrate.
Recycled mutagenesis was carried out on the D246A variant, and revertants
with enhanced activity were isolated by their dark blue phenotype on a
dinD Colon, two colons lacZ DNA damage indicator strain. Most of the amino
acid substitutions present within the revertants were located outside the
DNA-protein interface. This study demonstrates that endonuclease mutants
with altered specificity and non-lethal activity can be evolved towards
more active variants using a laboratory evolution strategy.

<>

<1>Zhuo, W., Lai, X., Zhang, L., Chan, S.-H., Li, F., Zhu, Z., Yang, M., Sun, D.
<2>Elimination of inter-domain interactions increases the cleavage fidelity of the restriction endonuclease DraIII.
<3>Protein Cell
<4>5
<5>357-368
<6>2014
<7>DraIII is a type IIP restriction endonucleases (REases) that recognizes and creates a double
strand break within the gapped palindromic sequence CACa dagger NNNa dagger'GTG of
double-stranded DNA (a dagger indicates nicking on the bottom strand; a dagger' indicates
nicking on the top strand). However, wild type DraIII shows significant star activity. In this
study, it was found that the prominent star site is CATa dagger GTTa dagger'GTG, consisting
of a star 5' half (CAT) and a canonical 3' half (GTG). DraIII nicks the 3' canonical half
site at a faster rate than the 5' star half site, in contrast to the similar rate with the
canonical full site. The crystal structure of the DraIII protein was solved. It indicated, as
supported by mutagenesis, that DraIII possesses a beta beta I +/--metal HNH active site. The
structure revealed extensive intra-molecular interactions between the N-terminal domain and
the C-terminal domain containing the HNH active site. Disruptions of these interactions
through site-directed mutagenesis drastically increased cleavage fidelity. The understanding
of fidelity mechanisms will enable generation of high fidelity REases.

<>

<1>Zhuo, Y., Liu, L., Wang, Q., Liu, X., Ren, B., Liu, M., Ni, P., Cheng, Y.Q., Zhang, L.
<2>Revised Genome Sequence of Burkholderia thailandensis MSMB43 with Improved Annotation.
<3>J. Bacteriol.
<4>194
<5>4749-4750
<6>2012
<7>There is growing interest in discovery of novel bioactive natural products from Burkholderia
thailandensis. Here we report a significantly improved genome
sequence and reannotation of Burkholderia thailandensis MSMB43, which will
facilitate the discovery of new natural products through genome mining and
studies of the metabolic versatility of this bacterium.

<>

<1>Zhuravleva, L.I., Oreshkin, E.N.
<2>The effect of cultivation conditions and ultrasonic destruction of Streptomyces achromogenes ATCC 12767 cells on the yield of restriction endonucleases.
<3>Prikl. Biokhim. Mikrobiol.
<4>21
<5>401-406
<6>1985
<7>The effect of the cultivation time, aeration rate and nutrient medium
composition on the yield of the restrictase activity of the submerged culture
of Streptomyces achromogenes ATCC 12767 was investigated.  A maximum value of
the specific restrictase activity was observed in the second part of the
exponential phase of growth (after 22-24 hours of cultivation).  An optimal
composition of the nutrient medium for cultivation of S. achromogenes was found
using mathematical methods of experiment planning.  The medium includes (g/l):
glucose-10.0; peptone-4.0; yeast extract-4.0; MgSO4.7H2O-0.5; K2HPO4-4.0;
KH2PO4-2.0.  An addition of detergents to the buffer during ultrasonic
destruction of S. achromogenes cells enabled the yield of the restrictase
activity to be increased more than twice.

<>

<1>Zhuravleva, L.I., Oreshkin, E.N., Bezborodov, A.M.
<2>Isolation and purification of restriction endonuclease SacI from Streptomyces achromogenes ATCC 12767.
<3>Prikl. Biokhim. Mikrobiol.
<4>23
<5>208-215
<6>1987
<7>A procedure for isolation and purification of restriction endonuclease SacI
from Streptomyces achromogenes ATCC 12767 is proposed.  It allows to obtain an
electrophoretically homogeneous enzyme yield by activity 3.7%.  The molecular
weight of SacI was found to be 52,000 +/-5000 D, and isoelectric point 6.2.
The enzyme consists of two subunits, which was found by polyacrylamide gel
electrophoresis under denaturing conditions.  Km and Vmax values were
determined for the enzymatic reaction; they are equal to 4.6x10-9 M and
9.19x10-10 M/min, respectively.

<>

<1>Zhurina, D., Dudnik, A., Waidmann, M.S., Grimm, V., Westermann, C., Breitinger, K.J., Yuan, J., van Sinderen, D., Riedel, C.U.
<2>High-Quality Draft Genome Sequence of Bifidobacterium longum E18, Isolated from a Healthy Adult.
<3>Genome Announcements
<4>1
<5>e01084-13
<6>2013
<7>Bifidobacteria are important gastrointestinal commensals of a number of animals,  including
humans, and various beneficial effects on host health have been
attributed to them. Here, we announce the noncontiguous finished genome sequence
of Bifidobacterium longum E18, isolated from a healthy adult, which reveals
traits involved in its interaction with the host.

<>

<1>Zhurina, D., Zomer, A., Gleinser, M., Brancaccio, V.F., Auchter, M., Waidmann, M.S., Westermann, C., van Sinderen, D., Riedel, C.U.
<2>Complete genome sequence of Bifidobacterium bifidum S17.
<3>J. Bacteriol.
<4>193
<5>301-302
<6>2010
<7>Here, we report on the first completely annotated genome sequence of a Bifidobacterium bifidum
strain. B. bifidum S17, isolated from faeces of a
breast-fed infant, was shown to strongly adhere to intestinal epithelial
cells and has potent anti-inflammatory activity in vitro and in vivo. The
genome sequence will provide new insights into the biology of this
potential probiotic organism and allow for the characterization of the
molecular mechanisms underlying its beneficial properties.

<>

<1>Zieger, M., Patillon, M., Roizes, G., Lerouge, T., Dupret, D., Jeltsch, J.M.
<2>Two restriction endonucleases from Bacillus sphaericus:  BspXI and BspXII.
<3>Nucleic Acids Res.
<4>15
<5>3919
<6>1987
<7>In screening wild strains of Bacillus sphaericus, one, which we call X -not totally
identifiable when compared to well established strains- exhibited a high yield of two
restriction endonucleases: BspXI and BspXII. A cleared sonic extract, obtained from 12 grams
of frozen cells yielded as much as 500,000 units of each enzyme by the following steps of
chromatography and dialysis [against PC buffer (10% glycerol, 10 mM KPO4-pH 7.4, 10 mM
B-mercaptoethanol, 0.1 mM EDTA)]. (I) A Biogel A-0.5 m filtration (II) Dialysis of active
fractions (III) DEAE Sephacel chromatography: a gradient elution from 0 to 1 M NaCl gave pool
I (0-0.5 M NaCl) and pool II (0.5-1 M NaCl). (IV-VI) Dialysed pool I was loaded on
phosphocellulose column and eluted with a 0 to 1 M KCl gradient; the fraction at 0.55 M KCl
was dialysed and the phosphocellulose chromatographic step repeated, yielding, at 0.55 M KCl,
a pure BspXI fraction. (VII) Dialysed pool II was applied on a Blue Trisacryl column and
eluted with a 0 to 0.5 M KCl gradient giving, at 0.3 M KCl, a pure BspXII fraction. The
digestion patterns of different DNAs (k, pBR322 and Ad2) with BspXI and BspXII were identical
to those obtained with ClaI and BclI, respectively, or with BspXI + ClaI and BspXII + BclI. It
was therefore concluded that BspXI and BspXII are isoschizomers of ClaI (5'-AT/CGAT-3') and
BclI (5'-T/GATCA-3') respectively (1,2). BspXI was further investigated using: [a] standard
procedures (3) to determine the cleavage site at the nucleotide level and [b] a recombinant
DNA (4) that contains a ClaI recognition sequence cleaved by the enzyme only when it is
produced in an E. coli dam- strain. Since BspXI behaves exactly like ClaI (sensitive to
N6-adenine methylation - generates 5' protruding dinucleotide CG), BspXI is a full
isoschizomer of ClaI with the property that this new enzyme is able to cleave DNA at 37C in
standard buffers containing from 0 to 200 mM NaCl without any noticeable loss of activity.

<>

<1>Zienkiewicz, M., Kern-Zdanowicz, I., Golebiewski, M., Zylinska, J., Mieczkowski, P., Gniadkowski, M., Bardowski, J., Ceglowski, P.
<2>Mosaic Structure of p1658/97, a 125-Kilobase Plasmid Harboring an Active Amplicon with the Extended-Spectrum -Lactamase Gene blaSHV-5.
<3>Antimicrob. Agents Chemother.
<4>51
<5>1164-1171
<6>2007
<7>Escherichia coli isolates recovered from patients during a clonal outbreak
in a Warsaw, Poland, hospital in 1997 produced different levels of an
extended-spectrum beta-lactamase (ESBL) of the SHV type. The
beta-lactamase hyperproduction correlated with the multiplication of ESBL
gene copies within a plasmid. Here, we present the complete nucleotide
sequence of plasmid p1658/97 carried by the isolates recovered during the
outbreak. The plasmid is 125,491 bp and shows a mosaic structure in which
all modules constituting the plasmid core are homologous to those found in
plasmids F and R100 and are separated by segments of homology to other
known regions (plasmid R64, Providencia rettgeri genomic island R391,
Vibrio cholerae STX transposon, Klebsiella pneumoniae or E. coli
chromosomes). Plasmid p1658/97 bears two replication systems, IncFII and
IncFIB; we demonstrated that both are active in E. coli. The presence of
an active partition system (sopABC locus) and two postsegregational
killing systems (pemIK and hok/sok) indicates that the plasmid should be
stably maintained in E. coli populations. The conjugative transfer is
ensured by the operons of the tra and trb genes. We also demonstrate that
the plasmidic segment undergoing amplification contains the blaSHV-5 gene
and is homologous to a 7.9-kb fragment of the K. pneumoniae chromosome.
The amplicon displays the structure of a composite transposon of type I.

<>

<1>Zimmerly, S., Guo, H., Eskes, R., Yang, J., Perlman, P.S., Lambowitz, A.M.
<2>A group II intron RNA is a catalytic component of a DNA endonuclease involved in intron mobility.
<3>Cell
<4>83
<5>529-538
<6>1995
<7>The mobility (homing) of the yeast mitochondrial DNA group II intron aI2 occurs via target
DNA-primed reverse transcription at a double-strand break in the recipient DNA.  Here, we show
that the site-specific DNA endonuclease that makes the double-strand break is a
ribonucleoprotein complex containing the aI2-encoded reverse transcriptase protein and excised
at aI2 RNA.  Remarkably, the aI2 RNA catalyzes cleavage of the sense strand of the recipient
DNA, while the AI2 protein appears to cleave the antisense strand.  The RNA-catalyzed sense
strand cleavage occurs via a partial reverse splicing reaction in which the protein component
stabilizes the active intron structure and appears to confer preference for DNA substrates.
Our results demonstrate a biologically relevant ribozyme reaction with a substrate other than
RNA.

<>

<1>Zimmerly, S., Guo, H., Perlman, P.S., Lambowitz, A.M.
<2>Group II intron mobility occurs by target DNA-primed reverse transcription.
<3>Cell
<4>82
<5>545-554
<6>1995
<7>Mobile group II introns encode reverse transcriptases and insert site specifically into
intronless alleles (homing).  Here, in vitro experiments show that homing of the yeast mtDNA
group II intron aI2 occurs by reverse transcription at a double-strand break in the recipient
DNA.  A site-specific endonuclease cleaves the antisense strand of recipient DNA at position
+10 of exon 3 and the sense strand at the intron insertion site.  Reverse transcription of
aI2-containing pre-mRNA is primed by the antisense strand cleaved in exon 3 and results in
cotransfer of the intron and flanking exon sequences.  Remarkably, the DNA endonuclease that
initiates homing requires both the AI2 reverse transcriptase protein and aI2 RNA.  Parallels
in their reverse transcription mechanisms raise the possibility that mobile group II introns
were ancestors of nuclear non-long terminal repeat retrotransposons and telomerases.

<>

<1>Zimmerly, S., Moran, J.V., Perlman, P.S., Lambowitz, A.M.
<2>Group II intron reverse transcriptase in yeast mitochondria.  Stabilization and regulation of reverse transcriptase activity by the intron RNA.
<3>J. Mol. Biol.
<4>289
<5>473-490
<6>1999
<7>Group II introns encode reverse transcriptases that function in both intron mobility and RNA
splicing. The proteins bind specifically to unspliced precursor RNA to promote splicing, and
then remain associated with the excised intron to form a DNA endonuclease that mediates intron
mobility by target DNA-primed reverse transcription. Here, immunoblotting and UV cross-linking
experiments show that the reverse transcriptase activity encoded by the yeast mtDNA group II
intron aI2 is associated with an intron-encoded protein of 62 kDa (p62). p62 is bound tightly
to endogenous RNAs in mitochondrial ribonucleoprotein particles, and the reverse transcriptase
activity is rapidly and irreversibly lost when the protein is released from the endogenous
RNAs by RNase digestion. Non-denaturing gel electrophoresis and activity assays show that the
aI2 reverse transcriptase is associated predominantly with the excised intron RNA, while a
smaller amount is associated with unspliced precursor RNA, as expected from the role of the
protein in RNA splicing. Although the reverse transcriptase in wild-type yeast strains is
bound tightly to endogenous RNAs, it is regulated so that it does not copy these RNAs unless a
suitable DNA oligonucleotide primer or DNA target site is provided. Certain mutations in the
intron-encoded protein or RNA circumvent this regulation and activate reverse transcription of
endogenous RNAs in the absence of added primer.  Although p62 is bound to unspliced precursor
RNA in position to initiate cDNA synthesis in the 3' exon, the major template for target
DNA-primed reverse transcription in vitro is the reverse-spliced intron RNA, as found
previously for aI1. Together, our results show that binding to intron-containing RNAs
stabilizes and regulates the activity of p62.

<>

<1>Zimmerman, W.J., Culley, D.E.
<2>Genetic variation at the apcAB, cpcAB, gvpA1, and nifH loci and in DNA methylation among N2-fixing cyanobacteria designated Nostoc punctiforme.
<3>Microb. Ecol.
<4>21
<5>199-209
<6>1991
<7>Genetic similarity among cyanobacteria of a morphological subgroup of Nostoc was evaluated
through a comparison of several specific genes and the extent of DNA methylation.  Four of six
cyanobacteria were originally cultured from facultative symbioses with higher plants (Gunnera
and Encephalartos); these and one free-living isolate had been identified or repute to be N.
punctiforme.  No consistent correlation to species or symbiotic history was found from DNA
hybridizations to genes coding for phycocyanin, allophycocyanin, gas vesicle protein, and
dinitrogenase reductase.  One gene (gvpC) was not present, and gvpA1 was a single-copy gene in
all strains.  The gas vesicle genes were concluded to be potentially useful for broadly
characterizing Nostoc or at least this subgroup.  Incubations of Nostoc genomic DNA with 22
restriction endonucleases indicated a higher degree of methylation and similarity of its
methylated DNA to that of other heterocystous cyanobacteria.  The genetic variation of the
Nostoc isolates was judged to reflect primarily different soil origins.

<>

<1>Zimmermann, C., Guhl, E., Graessmann, A.
<2>Mouse DNA methyltransferase (MTase) deletion mutants that retain the catalytic domain display neither de novo nor maintenance methylation activity in vivo.
<3>Biol. Chem.
<4>378
<5>393-405
<6>1997
<7>The mammalian genome encodes a DNA cytosine-5-methyltransferase (MTase) of about 170 kDa that
is apparently responsible for both de novo and maintenance methylation at CpG sites.  Both
methylation activities have to be regulated accurately to ensure correct developmental and
cell type-specific gene activity.  Distorted DNA methylation patterns have been associated
with cell aging and diseases such as cancer and fragile X syndrome.  Structural and functional
in vitro studies of the mouse MTase have indicated that the enzyme has both a regulatory and a
catalytic region located in the N-terminal and C-terminal parts of the protein, respectively.
The regulatory region includes the nuclear localization signal (NLS), the sequence for DNA
targeting and the Zn-binding domain.  The catalytic domain carries the ten consensus sequence
motifs specific for all known pro- and eukaryotic DNA cytosine-5-methyltransferases.  In an
attempt to separate regulatory and catalytic functions of the enzyme in vivo, we have tested
various deletion mutations by means of transient and stable cell transfection experiments.
Expression of the transgenes, all of which retained the C-terminal catalytic domain, was
monitored by immunofluorescence staining, Northern blot analysis and SDS gel electrophoresis.
Despite high levels of transgene expression, the truncated MTase molecules exhibited neither
de novo nor maintenance methylation activity.  These findings might indicate that in vivo, an
efficient control mechanism prevents the ectopic activity of the DNA Mtase that is
structurally compromised in its N-terminal regulatory region.

<>

<1>Zimmermann, K., Schogl, D., Mannhalter, J.W.
<2>Digestion of terminal restriction endonuclease recognition sites on PCR products.
<3>Biotechniques
<4>24
<5>582-584
<6>1998
<7>One of the common methods for cloning polymerase chain reaction products is overhanging-end
cloning (also known as sticky-end or directional cloning).  Frequently, it is not possible to
use restriction enzyme sites already present in the amplified product, and primers that encode
recognition sites of restriction endonucleases in addition to the specific sequence have to be
designed.  After amplification of the target sequence with these primers, the PCR products are
purified, digested with restriction enzymes and cloned into vectors treated with the same
enzymes.  However, it has been found that many restriction enzymes fail to cleave at the end
of PCR fragments.  To circumvent this problem, the addition of at least three more nucleotides
at the end of a restriction site was suggested.

<>

<1>Zingg, J.-M., Jones, P.A.
<2>Genetic and epigenetic aspects of DNA methylation on genome expression, evolution, mutation and carcinogenesis.
<3>Carcinogenesis
<4>18
<5>869-882
<6>1997
<7>DNA methylation has at least two important roles in tumorigenesis.  The target cytosines (C*)
of (cytosine-5)-DNA methyltransferase (Mtase) are mutated to thymine (T) in ~30% of inherited
diseases and cancer, and genome-wide alterations of DNA methylation patterns occur at early
stages of tumor development.  Insight into the normal function of DNA methylation will provide
the knowledge to understand the origins of these aberrations and their importance for disease
initiation and progression.  Originally the aberrations seen in tumors were attributed to the
higher spontaneous deamination rate of 5-methylcytosine (5-mC) as compared with C and to
misregulation of the Mtase gene.  Recently, it has become clear that the Mtase can actively
participate in mutagenesis by enzymatically increasing both the rate of genetic and epigenetic
alterations.  Proteins that recognize and repair these alterations determine the frequency of
their fixation as disease causing mutations.  In addition, alterations in the metabolism of
S-adenosylmethionine can disturb DNA methylation by depleting the cofactor
S-adenosylmethionine or by increasing the level of metabolites acting as inhibtors of DNA
methylation.  This review concentrates on the normal role of DNA methylation in mammals and on
aberrations of DNA methylation in inherited disease and cancer.

<>

<1>Zingg, J.-M., Shen, J.-C., Jones, P.A.
<2>Enzyme-mediated cytosine deamination by the bacterial methyltransferase M.MspI.
<3>Biochem. J.
<4>332
<5>223-230
<6>1998
<7>Most prokaryotic (cytosine-5)-DNA methyltransferases increase the frequency of deamination at
the cytosine targeted for methylation in vitro in the absence of the cofactor
S-adenosyl-methionine or the reaction product S-adenosyl-homocysteine.  We show here that,
under the same in vitro conditions, the prokaryotic methyltransferase, M.MspI (from Moraxella
sp.), causes very few cytosine deaminations, suggesting a mechanism in which M.MspI may avoid
enzyme-mediated cytosine deamination.  Two analogues of AdoMet, sinefungin and
5'-amino-5'-deoxyadenosine, greatly increased the frequency of cytosine deamination mediated
by M.MspI presumably by introducing a proton-donating amino group into the catalytic center,
thus facilitating the formation of an unstable enzyme-dihydrocytosine intermediate and
hydrolytic deamination.  Interestingly, two naturally occurring analogues, adenosine and
5'-methylthio-5'-deoxyadenosine, which do not contain a proton-donating amino group, also
weakly increased the deamination frequency by M.MspI, even in the presence of AdoMet or
AdoHcy. These analogues may trigger a conformational change in the enzyme without completely
inhibiting the access of solvent water to the catalytic center, thus allowing hydrolytic
deamination of the enzyme-dihydrocytosine intermediate.  Under normal physiological conditions
the enzymes M.HpaII (from Haemophilus parainfluenzae), M.HhaI (from Haemophilus haemolytica)
and M.MspI all increased the in vivo deamination frequency at the target cytosines with
comparable efficiency.

<>

<1>Zingg, J.-M., Shen, J.-C., Yang, A.S., Rapoport, H., Jones, P.A.
<2>Methylation inhibitors can increase the rate of cytosine deamination by (cytosine- 5)-DNA methyltransferase.
<3>Nucleic Acids Res.
<4>24
<5>3267-3275
<6>1996
<7>The target cytosines of (cytosine-5)-DNA methyltransferases in prokaryotic
and eukaryotic DNA show increased rates of C->T transition mutations compared to non-
target cytosines.  These mutations are induced either by the spontaneous deamination of 5-
mC->T generating inefficiently repaired G:T rather than G:U mismatches, or by the
enzyme-induced C->U deamination which occurs under conditions of reduced levels of S-
adenosylmethionine (AdoMet) and S-adenosyl-homocysteine (AdoHcy).  We tested
whether various inhibitors of (cytosine-5)-DNA methyltransferases analogous to AdoMet
and AdoHcy would affect the rate of enzyme-induced deamination of the target cytosine by
M.HpaII and M.SssI.  Interestingly, we found two compounds, sinefungin and 5'-amino-
5'-deoxyadenosine, that increased the rate of deamination 103-fold in the presence and
104-fold in the absence of AdoMet and AdoHcy.  We have therefore identified the first
mutagenic compounds specific for the target sites of (cytosine-5)-DNA methyltransferases.
A number of analogs of AdoMet and AdoHcy have been considered as possible antiviral,
anticancer, antifungal and anti-parasitic agents.  Our findings show that chemotherapeutic
agents with affinities to the cofactor binding pocket of (cytosine-5)-DNA methyltransferase
should be tested for their potential mutagenic effects.

<>

<1>Zinkevich, V., Heslop, P., Glover, S.W., Weiserova, M., Hubacek, J., Firman, K.
<2>Mutation in the specificity polypeptide of the Type I restriction endonuclease R EcoK that affects subunit assembly.
<3>J. Mol. Biol.
<4>227
<5>597-601
<6>1992
<7>We describe the isolation and characterization of a temperature-sensitive mutation within the
hsdS gene of the type I restriction and modification system EcoK. This mutation appears to
affect the ability of the HsdR subunit to interact with the HsdS subunit when forming an
active endonuclease. We discuss the possibility that this mutant, together with another
mutation described previously, may define a discontinuous domain, involved in protein-protein
interactions, within the HsdS polypeptide.

<>

<1>Zinkevich, V., Popova, L., Kryukov, V., Abadjieva, A., Bogdarina, I., Janscak, P., Firman, K.
<2>The HsdR subunit of R.EcoR124II: cloning and over-expression of the gene and unexpected properties of the subunit.
<3>Nucleic Acids Res.
<4>25
<5>503-510
<6>1997
<7>Type I restriction endonucleases are composed of three subunits, HsdR, HsdM and HsdS.  The
HsdR subunit is absolutely required for restriction activity; while an independent methylase
is composed of HsdM and HsdS subunits.  DNA cleavage is associated with a powerful ATPase
activity during which DNA is translocated by the enzyme prior to cleavage.  The presence of a
Walker type I ATP-binding site within the HsdR subunit suggested that the subunit may be
capable of independent enzymatic activity.  Therefore, we have, for the first time, cloned and
over-expressed the hsdR gene of the type IC restriction endonuclease EcoR124II.  The purified
HsdR subunit was found to be a soluble monomeric protein capable of DNA- and Mg2+-dependent
ATP hydrolysis.  The subunit was found to have a weak nuclease activity both in vivo and in
vitro, and to bind plasmid DNA; although was not capable of binding a DNA oligoduplex.  We
were also able to reconstitute the fully active endonuclease from purified M.EcoR124I and
HsdR.  This is the first clear demonstration that the HsdR subnit of a type I restriction
endonuclease is capable of independent enzyme activity, and suggests a mechanism for the
evolution of the endonuclease from the independent methylase.

<>

<1>Zinkevich, V.E., Alekseev, A.M., Tanyashin, V.I.
<2>Influence of bacteriophage lambda ral gene on the level of synthesis of the restriction endonuclease EcoK beta-subunit.
<3>Mol. Biol. (Mosk)
<4>20
<5>1638-1644
<6>1986
<7>E. coli hsd genes were subcloned from lambda 642 (ra1+) into the lambda
L47.1
vector (ra1- after replacement).  We investigated the influence of the bacteriophage lambda
ra1 gene
on the expression efficiency of hsdSk and hsdMk genes.  Its presence in vitro enhanced the
synthesis of the beta-subunit and hsdMk gene product.  Increased modification in vivo was
also
observed.  It is proposed that the increase in the modification rate of lambda-phage fully
unmodified DNA is connected with the appearance of E. coli DNA methylase consisting of
beta-
and gamma-subunits.

<>

<1>Zinkevich, V.E., Solonin, A.S., Bogdarina, I.G., Tanyashin, V.I.
<2>Cloning and restriction analysis of the BamHI-EcoRI fragment of DNA, containing genes of the hsd region of Escherichia coli.
<3>Dokl. Akad. Nauk.
<4>259
<5>216-218
<6>1981
<7>
<>

<1>Zinkevich, V.E., Solonin, A.S., Tanyashin, V.I., Bayev, A.A.
<2>Function of hsdS gene of E. coli K in recombinant plasmid containing the regulatory lambda phage region.
<3>Dokl. Akad. Nauk.
<4>263
<5>717-721
<6>1982
<7>None

<>

<1>Zinkevich, V.E., Weiserova, M., Kryukov, V.M., Hubacek, J.
<2>A mutation that converts serine 340 of the HsdSK polypeptide to phenylalanine and its effects on restriction and modification in Escherichia coli K-12.
<3>Gene
<4>90
<5>125-128
<6>1990
<7>A hybrid hsdS gene, encoding the HsdSts+d polypeptide, was constructed by joining the proximal
region of the wild-type (wt) hsdS sequence with the distal region of the hsdSts+d sequence, at
the hsdS BglII site. The hybrid hsdS-Sts+d gene exerts a trans-dominant effect on restriction
and modification, which points to the location of the temperature-sensitive (ts)
trans-dominant (+d) mutation in the gene hsdSts+d distal region. Sequencing of the region
downstream from the HindIII target in the Escherichia coli K-12 hsdSts+d mutant was carried
out. It is identical to the wt hsdS sequence (GenBank/EMBL accession number ECHSDK LV00288),
except for a single base-pair transition C1245 -> T. The results obtained support the idea
that the trans-dominant effect of the ts mutation described earlier is related to the single
base-pair transition in the nonhomologous region of the hsdSts+d sequence.

<>

<1>Zinkevich, V.E., Zograf, Y.N., Tanyashin, V.I.
<2>Expression of the EcoK DNA-methylase genes cloned.
<3>Dokl. Akad. Nauk.
<4>282
<5>993-995
<6>1985
<7>The present article gives results on in vivo and in vitro expression of genes
hsdSk and hsdMk, cloned in pBR322, and the possible involvement of the rho
factor in this process.

<>

<1>Zinkevich, V.E., Zograf, Y.N., Tanyashin, V.I.
<2>The genes of EcoK DNA methylase:  cloning and expression.
<3>Dokl. Akad. Nauk.
<4>279
<5>1493-1496
<6>1984
<7>None

<>

<1>Zinn, A.R., Butow, R.A.
<2>Nonreciprocal exchange between alleles of the yeast mitochondrial 21S rRNA gene:  kinetics and the involvement of a double-strand break.
<3>Cell
<4>40
<5>887-895
<6>1985
<7>A 1.1 kb intron containing an open reading frame (ORF) in one allele (Omega+) of the yeast
mitochondrial 21S rRNA gene is nearly quantitatively inserted in crosses into a 21S rRNA
allele lacking that intron (Omega-). We have determined that this nonreciprocal exchange
initiates soon after cells fuse to form zygotes and is complete by 10-16 hr after mating. We
have discovered a unique in vivo double-strand cut in Omega- mitochondrial DNA (mtDNA) at or
near the site of intron insertion that is implicated in the process. Markers flanking the
intron insertion site are coconverted with frequencies inversely proportional to their
distance from that site. There is no net conversion of Omega- to Omega+ in crosses between
petites retaining these alleles, nor do we observe the unique double-strand cut in the mtDNA
from zygotes of such crosses. The data suggest that a translation product of the intron ORF is
required for the double-strand cut and nonreciprocal recombination at Omega.

<>

<1>Zinovev, V.V., Kolesnikov, V.A., Beznedelnaya, N.L., Gilev, A.F., Gorbunov, Y.A., Popov, S.G., Malygin, E.G.
<2>Interaction of restrictase BamHI with synthetic substrates containing complete or partial recognition sites.
<3>Mol. Biol. (Mosk)
<4>18
<5>169-175
<6>1984
<7>The cleavage of self-complementary oligodeoxyribonucleotides by restrictase
BamHI was studied.  Oligonucleotides containing the complete recognition
sequence GGATCC were cleaved by restrictase BamHI in accordance with the
established specificity of the enzyme.  The oligonucleotide TCCAGATCTGGA
contains part of the recognition sequence (GATC) in the middle of the chain and
separate halves of the sequence at each end.  Hydrolysis of this
oligonucleotide yields products which indicate the interaction of restrictase
BamHI with the composite recognition site GGA...TCC formed by the ends of two
separate substrate molecules.  The oligonucleotide was not cleaved at the GATC
sequence.  The results obtained support a symmetrical model for restrictase
interaction with DNA.  According to this model, n/2 of the nucleotides in the
recognition site bind to the enzyme.

<>

<1>Zinovev, V.V., Kolesnikov, V.A., Beznedelnaya, N.L., Gorbunov, J.A., Popov, S.G., Malygin, E.G.
<2>Restriction endonuclease BamHI interaction with a synthetic duplex containing half-size recognition sequences.
<3>FEBS Lett.
<4>132
<5>98-100
<6>1981
<7>Restriction endonuclease BamHI is a site-specific deoxyribonuclease which
cleaves the phosphodiester bonds between the guanine residues within the duplex
DNA sequence 5'GGATCC   CCTAGG It is well known that restriction endonucleases
recognise and cleave relatively short synthetic duplexes containing appropriate
recognition sequence, therefore oligodeoxyribonucleotides called 'linkers' are
widely used for genetic manipulations.  On the other hand, the synthetic
oligonucleotides with defined sequences are useful tools to study mechanisms of
interaction of the enzymes with DNA.

<>

<1>Zinovev, V.V., Ovechkina, L.G., Malygin, E.G.
<2>Stoichiometry of phage T4 Dam DNA (Adenine-N6)-methyltransferase binding with oligonucleotide substrates.
<3>Mol. Biol. (Mosk)
<4>30
<5>1203-1208
<6>1996
<7>Phage T4 Dam DNA methyltransferase recognizes the GATC site and methylates the adenine.  The
stoichiometry of its complex with the substrate was assessed by gel filtration and sucrose
gradient ultracentrifugation.  The results obtained by both methods indicate that two enzyme
subunits bind with 32- and 20-bp DNA duplexes.

<>

<1>Zinovev, V.V., Rechkunova, N.I., Gorbunov, Y.A., Buryanov, Y.I., Malygin, E.G.
<2>The significance of internucleotide phosphates and noncomplementary substitutions in the oligonucleotide substrates on the interaction with the DNA methylase Ecodam.
<3>Biopol. Kletka
<4>5
<5>22-27
<6>1989
<7>Eco dam methylase was investigated for its interaction with different synthetic
oligonucleotide substrates containing some defects in the GATC sequence. The defects are as
follows: the absence of one or several nucleotide residues, the presence of 3'-phosphate
residue with CH3-S-group, noncomplementary substitution of A by G or of G by A in the
recognition site of methylase Eco dam. The presence of the 3'S-methyl thiophosphate residue
has no essential effect on the methylation of oligonucleotide complexes as compared to the
analogous complexes lacking internucleotide phosphate. The introduction of the
noncomplementary base pair in the recognition site results in the loss of substrate properties
of such imperfect duplexes.

<>

<1>Zinoviev, V.V., Evdokimov, A.A., Gorbunov, Y.A., Malygin, E.G., Kossykh, V.G., Hattman, S.
<2>Phage T4 DNA [N6-adenine] methyltransferase: Kinetic studies using oligonucleotides containing native or modified recognition sites.
<3>Biol. Chem.
<4>379
<5>481-488
<6>1998
<7>The DNA-[N6-adenine] methyltransferase of T4 phage catalyzes methyl group transfer from
S-adenosyl-L-methionine to the N6-position of adenine in the palindromic sequence, GATC.  We
have investigated the effect of eliminating different structural components of the recognition
site on the ability of a substrate to be bound and methylated by T4 Dam.  For this purpose,
steady state binding (by gel shift assays) and kinetic parameters of methylation (using the
methyl donor, [3H-CH3]-AdoMet, at 25 C) were studied using various synthetic duplex
oligonucleotides containing some defect in the DNA-target site; e.g., the absence of an
internucleotide phosphate or a nucleotide(s) within the recognition site, or a single stranded
region.  The salient results are summarized as follows: (1) Addition of T4 Dam to a complete
reaction mixture (with a 20-mer duplex as substrate) resulted in a 'burst' of 3H-methylated
product, followed by a constant rate of product formation that reflected establishment of
steady-state conditions.  This suggests that the rate-limiting step is release of product
methylated DNA from the enzyme [and not the transfer of the methyl group].  (2) A number of
the defects in duplex structure had only weak influence on the binding and Km values, but
strongly reduced the kcat. At the same time, several poorly bound duplexes retained good
substrate characteristics, especially duplexes having uninterrupted GAT-sequences in both
strands.  Whereas having only one half of the recognition site element intact was sufficient
for stable complex formation, the catalytic turnover process had a strict requirement for an
uninterrupted GAT-sequence on both strands.  (3) There was no correlation between Km and
binding capability; the apparent Kd for some duplexes was 5-70 times higher than Km.  This
indicates that the T4 Dam methylation reaction cannot be explained by a simple Michaelian
scheme.

<>

<1>Zinoviev, V.V., Evdokimov, A.A., Hattman, S., Malygin, E.G.
<2>Molecular enzymology of phage T4 Dam DNA methyltransferase.
<3>Mol. Biol. (Mosk)
<4>38
<5>737-751
<6>2004
<7>This review summarizes the results of a study of the molecular mechanisms of phage T4 DNA
adenine methyltransferase (T4Dam) action.
T4Dam [EC 2.1.1.72] catalyzes the transfer of a methyl group from
S-adenosyl-L-methionine (AdoMet) to N-6 of the adenine located in the
palindromic recognition site GATC. The subunit structure of T4Dam,
substrate-binding properties, and kinetic parameters of methylation of
a variety of native and modified oligonucleotide duplexes are
considered. A kinetic scheme of the reaction was proposed, assuming
that T4Dam is isomerized into a catalytically active form. The
mechanisms of DNA-induced dimerization of T4Dam, flipping of the target
base, reorientation of T4Dam on an asymmetrically methylated
recognition site, the effector action of substrates, and processive
methylation of extended DNA containing more than one specific site are
discussed. The results obtained for T4Dam may provide a better
understanding of the action mechanisms of other homologous enzymes
including, first and foremost, those of the vast Dam family.

<>

<1>Zinoviev, V.V., Evdokimov, A.A., Malygin, E.G.
<2>DNA-[N4-Cytosine]-Methyltransferase from Bacillus amyloliquefaciens: Mechanism of Action Derived from Steady-State Kinetics.
<3>Mol. Biol. (Mosk)
<4>37
<5>128-138
<6>2003
<7>Kinetic analysis of methyl group transfer from S-adenosyl-L-methionine (SAM) to the 5'-GGATCC
recognition site catalyzed by the
DNA-[N4-cytosine]-methyltransferase from Bacillus amyloliquefaciens [EC
2.1.1.113] has shown that the dependence of the rate of methylation of the
20-meric substrate duplex on SAM and DNA concentration are normally
hyperbolic, and the maximal rate is attained upon enzyme saturation with
both substrates. No substrate inhibition is observed even at
concentrations many times higher than the Km values (0.107 microM for DNA
and 1.45 microM for SAM), which means that no nonreactive enzyme-substrate
complexes are formed during the reaction. The overall pattern of product
inhibition corresponds to an ordered steady-state mechanism following the
sequence SAM decreases DNA decreases metDNA increases SAH increases
(S-adenosyl-L-homocysteine). However, more detailed numerical analysis of
the aggregate experimental data admits an alternative order of substrate
binding, DNA decreases SAM decreases, though this route is an order of
magnitude slower.

<>

<1>Zinoviev, V.V., Evdokimov, A.A., Malygin, E.G., Schlagman, S.L., Hattman, S.
<2>Bacteriophage T4 dam DNA-[N6-adenine] methyltransferase: Processivity and orientation to the methylation target.
<3>J. Biol. Chem.
<4>278
<5>7829-7833
<6>2002
<7>We carried out steady state and pre-steady state (burst) kinetic analyses of the bacteriophage
T4 Dam DNA-[N(6)-adenine] methyltransferase (MTase) mediated methyl group transfer from
S-adenosyl-L-methionine (AdoMet) to Ade in oligonucleotide duplexes containing one or two
specific GATC sites with different combinations of methylated and unmodified targets. We
compared the results for ligated 40-mer duplexes with those of mixtures of the two unligated
duplexes used to generate the 40-mers. The salient results are as follows: (i) T4 Dam MTase
modifies 40-mer duplexes in a processive fashion. (ii) During processive movement, T4 Dam
rapidly exchanges product S-adenosyl-L-homocysteine (AdoHcy) for substrate AdoMet without
dissociating from the DNA duplex. (iii) T4 Dam processivity is consistent with an ordered
bi-bi mechanism AdoMet{downward arrow}DNA{downward arrow}DNA(Me){upward arrow}AdoHcy{upward
arrow}. However, in contrast to the steady state, here DNA(Me){upward arrow} signifies
departure from a methylated site GMTC{upward arrow}without physically dissociating from the
DNA. (iv) Following methyl transfer at one site and linear diffusion to a hemi-methylated
site, a reconstituted T4 Dam-AdoMet complex rapidly reorients itself to the (productive)
unmethylated strand. T4 Dam-AdoHcy can not reorient at an enzymatically created GMTC site. (v)
The inhibition potential of fully methylated sites 5'-GMTC/5'-GMTC is much lower for a long
DNA molecule compared to short single-site duplexes.

<>

<1>Zinoviev, V.V., Evdokimov, A.A., Malygin, E.G., Sclavi, B., Buckle, M., Hattman, S.
<2>Differential methylation kinetics of individual target site strands by T4Dam DNA methyltransferase.
<3>Biol. Chem.
<4>388
<5>1199-1207
<6>2007
<7>Prokaryote DNA methyltransferases (MTases) of the Dam family (including those of
bacteriophages T2 and T4) catalyze methyl group transfer from
S-adenosyl-L-methionine (AdoMet), producing S-adenosyl-L-homocysteine
(AdoHcy) and methylated adenine residues in palindromic GATC sequences.
Dam DNA MTases, as all site-specific enzymes interacting with polymeric
DNA, require a mechanism of action that ensures a rapid search for
specific targets for catalytic action, during both the initial and
subsequent rounds of methylation. The results of presteady-state
(reaction burst) and steady-state methylation analyses of individual
targets permitted us to monitor the action of T4Dam, which has three
degrees of freedom: sliding, reorientation and adaptation to the
canonical GATC sequence. The salient results are as follows: (i) 40mer
substrate duplexes containing two canonical GATC sites showed
differential methylation of the potential targets, i.e., T4Dam
exhibited a preference for one site/target, which may present the
better 'kinetic trap' for the enzyme. (ii) Prior hemimethylation of the
two sites made both targets equally capable of being methylated during
the pre-steady-state reaction. (iii) Although capable of moving in
either direction along double-stranded DNA, there are some restrictions
on T4Dam reorientation/adaptation on 40mer duplexes.

<>

<1>Zinoviev, V.V., Gorbunov, J.A., Baclanov, M.M., Popov, S.G., Malygin, E.G.
<2>Structure subtraction as an approach to investigation of the mechanism of restriction enzyme action.
<3>FEBS Lett.
<4>154
<5>282-284
<6>1983
<7>Endonuclease BamHI cleaves the phosphodiester bonds between the guanine residues within the
duplex DNA sequence G/GATCC.  The substrate characteristics of oligonucleotides, containing
some defects in the sequence recognized by endonuclease (nick, absence of some internucleotide
phosphate or nucleotide, partially single-stranded form of the recognition site) were
investigated.  The results suggest that the specificity of synthetic oligonucleotide cleavage
is strongly dependent on the ribosophosphate backbone intactness inside the recognition site.
BamHI was found not to hydrolyse the phosphodiester bonds outside the double helix.  Also
BamHI forms a productive complex with the non-symmetrical substrate, having half the
recognition sites, of a single strand.

<>

<1>Zinoviev, V.V., Gorbunov, Y.A., Popov, S.G., Malygin, E.G., Buryanov, J.I., Nesterenko, V.F., Baev, A.A., Venoginskis, M.T.
<2>Interaction of EcoDam methylase with single stranded sequences and synthetic oligonucleotides.
<3>Mol. Biol. (Mosk)
<4>19
<5>947-954
<6>1985
<7>Interaction of the Ecodam methylase with different substrates was investigated among them
double- and single-stranded DNAs and synthetic oligonucleotides containing some defects in the
GATC sequence.  These defects were: nick, the absence of one internucleotide phosphate of
nucleotide; partially single-stranded form of the recognition site etc.  It was demonstrated
that the presence of both G.A-dinucleotides in the recognition site is necessary for
productive enzyme-substrate interaction.  The absence of T and/or C residues is less dramatic
for methylase activity.  The Ecodam methylase is able to modify single-stranded
oligonucleotides by forming the double-stranded structure in the symmetric recognition
sequences GATC.

<>

<1>Zinoviev, V.V., Yakishchik, S.I., Evdokimov, A.A., Malygin, E.G., Hattman, S.
<2>Symmetry elements in DNA structure important for recognition/methylation by DNA [amino]-methyltransferases.
<3>Nucleic Acids Res.
<4>32
<5>3930-3934
<6>2004
<7>The phage T4Dam and EcoDam DNA-[adenine-N6] methyltransferases (MTases) methylate GATC
palindromic sequences, while the BamHI DNA-[cytosine-N4] MTase methylates the GGATCC
palindrome (which contains GATC) at the internal cytosine residue. We compared the ability of
these enzymes to interact productively with defective duplexes in which individual elements
were deleted on one chain. A sharp decrease in kcat was observed for all three enzymes if a
particular element of structural symmetry was disrupted. For the BamHI MTase, integrity of the
ATCC was critical, while an intact GAT sequence was necessary for the activity of T4Dam, and
an intact GA was necessary for EcoDam. Theoretical alignment of the region of best contacts
between the protein and DNA showed that in the case of a palindromic interaction site, a zone
covering the 5'-symmetric residues is located in the major groove versus a zone of contact
covering the 3'-symmetric residues in the minor groove. Our data fit a simple rule of thumb
that the most important contacts are aligned around the methylation target base: if the target
base is in the 5' half of the palindrome, the interaction between the enzyme and the DNA
occurs mainly in the major groove; if it is in the 3' half, the interaction occurs mainly in
the minor groove.

<>

<1>Zischka, M., Kuenne, C., Blom, J., Dabrowski, P.W., Linke, B., Hain, T., Nitsche, A., Goesmann, A., Larsen, J., Jensen, L.B., Witte, W., Werner, G.
<2>Complete Genome Sequence of the Porcine Isolate Enterococcus faecalis D32.
<3>J. Bacteriol.
<4>194
<5>5490-5491
<6>2012
<7>The complete and annotated genome sequence of Enterococcus faecalis D32, a commensal strain
isolated from a Danish pig, suggests putative adaptation to the
porcine host and absence of distinct virulence-associated traits.

<>

<1>Zivanovic, Y., Armengaud, J., Lagorce, A., Leplat, C., Guerin, P., Dutertre, M., Anthouard, V., Forterre, P., Wincker, P., Confalonieri, F.
<2>Genome analysis and genome-wide proteomics of Thermococcus gammatolerans, the most radioresistant organism known amongst the Archaea.
<3>Genome Biol.
<4>10
<5>R70
<6>2009
<7>ABSTRACT: BACKGROUND: Thermococcus gammatolerans was isolated from samples collected from
hydrothermal chimneys. It is one of the most radioresistant
organisms known amongst the Archaea. We report the determination and
annotation of its complete genome sequence, its comparison with other
Thermococcales genomes, and a proteomic analysis. RESULTS: T.
gammatolerans has a circular chromosome of 2.045 Mbp without any
extra-chromosomal elements, coding for 2,157 proteins. A thorough
comparative genomics analysis revealed important but unsuspected genome
plasticity differences between sequenced Thermococcus and Pyrococcus
species which could not be attributed to the presence of specific mobile
elements. Two virus-related regions tgv1 and tgv2 are the only mobile
elements identified in this genome. A proteogenome analysis was performed
by a shotgun LC-MS/MS approach allowing the identification of 10,931
unique peptides corresponding to 951 proteins. This information
concurrently validates the accuracy of the genome annotation.
Semi-quantitation of proteins by spectral count was done on exponential-
and stationary-phase cells. Insights into general catabolism, hydrogenase
complexes, detoxification systems, and the DNA repair toolbox of this
archaeon are revealed through this genome and proteome analysis.
CONCLUSIONS: This work is the first archaeal proteome investigation done
at the stage of primary genome annotation. This archaeon is shown to use a
large variety of metabolic pathways even under a rich medium growth
condition. This proteogenomic study also indicates that the high
radiotolerance of T. gammatolerans is probably due to proteins that remain
to be characterized rather than a larger arsenal of known DNA repair
enzymes.

<>

<1>Zobanikova, M., Mikolka, P., Cejkova, D., Pospisilova, P., Chen, L., Strouhal, M., Qin, X., Weinstock, G.M., Smajs, D.
<2>Complete genome sequence of Treponema pallidum strain DAL-1.
<3>Standards in Genomic Sciences
<4>7
<5>12-21
<6>2012
<7>strain DAL-1 is a human uncultivable pathogen causing the sexually transmitted disease
syphilis. Strain DAL-1 was isolated from the amniotic fluid of a pregnant
woman in the secondary stage of syphilis. Here we describe the 1,139,971 bp long
genome of strain DAL-1 which was sequenced using two independent sequencing
methods (454 pyrosequencing and Illumina). In rabbits, strain DAL-1 replicated
better than the strain Nichols. The comparison of the complete DAL-1 genome
sequence with the Nichols sequence revealed a list of genetic differences that
are potentially responsible for the increased rabbit virulence of the DAL-1
strain.

<>

<1>Zobanikova, M., Strouhal, M., Mikalova, L., Cejkova, D., Ambrozova, L., Pospisilova, P., Fulton, L.L., Chen, L., Sodergren, E., Weinstock, G.M., Smajs, D.
<2>Whole genome sequence of the Treponema pallidum Fribourg-Blanc: Unspecified simian isolate is highly similar to the yaws subspecies.
<3>PLoS Neglected Trop. Dis.
<4>7
<5>E2172
<6>2013
<7>Background: Unclassified simian strain Treponema Fribourg-Blanc was isolated in 1966 from
baboons (Papio cynocephalus)in West Africa. This strain was morphologically indistinguishable
from T. pallidum ssp. pallidum or ssp. pertenue strains, and it was shown to cause human
infections.
Methodology/Principal Findings: To precisely define genetic differences between Treponema
Fribourg-Blanc (unclassified simian isolate, FB) and T. pallidum ssp. pertenue strains (TPE),
a high quality sequence of the whole Fribourg-Blanc genome was determined with
454-pyrosequencing and Illumina sequencing platforms. Combined average coverage of both
methods was greater than 5006. Restriction target sites (n = 1,773), identified in silico, of
selected restriction enzymes within the Fribourg-Blanc genome were verified experimentally and
no discrepancies were found. When compared to the other
three sequenced TPE genomes (Samoa D, CDC-2, Gauthier), no major genome rearrangements were
found. The Fribourg-
Blanc genome clustered with other TPE strains (especially with the TPE CDC-2 strain), while T.
pallidum ssp. pallidum strains clustered separately as well as the genome of T.
paraluiscuniculi strain Cuniculi A. Within coding regions, 6 deletions, 5 insertions and 117
substitutions differentiated Fribourg-Blanc from other TPE genomes.  Conclusions/Significance:
The Fribourg-Blanc genome showed similar genetic characteristics as other TPE strains.
Therefore, we propose to rename the unclassified simian isolate to Treponema pallidum ssp.
pertenue strain Fribourg-Blanc. Since the Fribourg-Blanc strain was shown to cause
experimental infection in human hosts, non-human primates could serve as possible reservoirs
of TPE strains. This could considerably complicate recent efforts to eradicate yaws. Genetic
differences specific for Fribourg-Blanc could then contribute for identification of cases of
animal-derived yaws infections.

<>

<1>Zomer, A., de Vries, S.P., Riesbeck, K., Meinke, A.L., Hermans, P.W., Bootsma, H.J.
<2>Genome Sequence of Moraxella catarrhalis RH4, an Isolate of Seroresistant Lineage.
<3>J. Bacteriol.
<4>194
<5>6969
<6>2012
<7>Here we report the annotated genome sequence of Moraxella catarrhalis strain RH4, a
seroresistant-lineage strain isolated from the blood of an infected patient.
This genome sequence will allow us to gain further insight into the genetic
diversity of clinical M. catarrhalis isolates and will facilitate study of M.
catarrhalis pathogenesis.

<>

<1>Zong, Z., Lu, X.
<2>Characterization of a New SCCmec Element in Staphylococcus cohnii.
<3>PLoS ONE
<4>5
<5>E14016
<6>2010
<7>BACKGROUND: Many SCCmec elements of coagulase-negative staphylococci
(CoNS) could not be typed using multiplex PCR. Such a 'non-typable' SCCmec
was encountered in a Staphylococcus cohnii isolate. METHODOLOGY/PRINCIPAL
FINDINGS: The SCCmec type of methicillin-resistant S. cohnii clinical
isolate WC28 could not be assigned using multiplex PCR. Newly-designed
primers were used to amplify ccrA and ccrB genes. The whole SCCmec was
obtained by three overlapping long-range PCR, targeting regions from
left-hand inverted repeat (IRL) to ccrA/B, from ccrA/B to mecA and from
mecA to orfX. The region abutting IRL was identified using inverse PCR
with self-ligated enzyme-restricted WC28 fragments as the template. WC28
SCCmec had a class A mec gene complex (mecI-mecR1-mecA). The ccrA and ccrB
genes were closest (89.7% identity) to ccrA(SHP) of Staphylococcus
haemolyticus strain H9 and to ccrB3 (90% identity) of Staphylococcus
pseudintermedius strain KM241, respectively. Two new genes potentially
encoding AAA-type ATPase were found in J1 region and a psiTn554 transposon
was present in J2 region, while J3 region was the same as many SCCmec of
Staphylococcus aureus. WC28 SCCmec abutted an incomplete SCC element with
a novel allotype of ccrC, which was closest (82% identity) to ccrC1 allele
9 in Staphylococcus saprophyticus strain ATCC 15305. Only two direct
target repeat sequences, one close to the 3'-end of orfX and the other
abutting the left end of WC28 SCCmec, could be detected.
CONCLUSIONS/SIGNIFICANCE: A new 35-kb SCCmec was characterized in a S.
cohnii isolate, carrying a class A mec gene complex, new variants of ccrA5
and ccrB3 and two novel genes in the J1 region. This element is flanked by
8-bp perfect inverted repeats and is similar to type III SCCmec in S.
aureus and a SCCmec in S. pseudintermedius but with different J1 and J3
regions. WC28 SCCmec was arranged in tandem with an additional SCC element
with ccrC, SCC(WC28), but the two elements might have integrated
independently rather than constituted a composite. This study adds new
evidence of the diversity of SCCmec in CoNS and highlights the need for
characterizing the 'non-typable' SCCmec to reveal the gene pool associated
with mecA.

<>

<1>Zoropogui, A. et al.
<2>Genome Sequence of the Human- and Animal-Pathogenic Strain Nocardia cyriacigeorgica GUH-2.
<3>J. Bacteriol.
<4>194
<5>2098-2099
<6>2012
<7>The pathogenic strain Nocardia cyriacigeorgica GUH-2 was isolated from a fatal human
nocardiosis case, and its genome was sequenced. The complete genomic
sequence of this strain contains 6,194,645 bp, an average G+C content of 68.37%,
and no plasmids. We also identified several protein-coding genes to which N.
cyriacigeorgica's virulence can potentially be attributed.

<>

<1>Zotchev, S.B., Schrempf, H., Hutchinson, C.R.
<2>Identification of a methyl-specific restriction system mediated by a conjugative element from Streptomyces bambergiensis.
<3>J. Bacteriol.
<4>177
<5>4809-4812
<6>1995
<7>pBL2 was identified genetically but not physically in Streptomyces lividans after its mating
with S. bambergiensis.  During conjugation, pBL2 was transferred at high frequency to S.
lividans and S. coelicolor, pBL2.1 DNA isolated from S. coelicolor exconjugants as a circular
plasmid was shown to derive from the genome of S. bambergiensis.  S. lividans carrying pBL2 or
pBL2.1 acquired a methyl-specific restriction (MsrA+) phenotype.  The corresponding enzyme
was partially purified and shown to resemble a class II endonuclease which cleaves
Dam-methylated DNA preferentially.

<>

<1>Zou, G.
<2>Methods for preparation and determination of the purity and activity of restriction endonucleases.
<3>Weishengwuxue Zazhi
<4>7
<5>63-66
<6>1987
<7>The main source of DNA restriction endonucleases (restriction enzymes) is from
bacteria.  They are widespread, and have many varieties.  The first restriction
enzyme was discovered in 1968, since then close to 500 of them have been
discovered, and most of them are type II restriction enzymes.  Type II
restriction enzymes are very important, they have been called the surgeon's
knife for molecular biology; they have served as an ideal model to study the
interaction between protein and DNA.  This article describes the general
methods for preparation and determination of reactivities of type II
restriction endonucleases.

<>

<1>Zou, G., Cao, X., Zhu, R.
<2>One step purification of restriction endonuclease PstI by sepharose affinity column separation.
<3>Shengwu Huaxue Yu Shengwu Wuli Jinzhan
<4>62
<5>64-66
<6>1985
<7>None

<>

<1>Zou, G., Gao, C., Pi, X.
<2>Kinetic studies on the irreversible inhibition of restriction endonuclease PstI by site-specific inhibitors.
<3>Wuhan Univ. J. Natural Sciences
<4>6
<5>859-863
<6>2001
<7>The irreversible modifying effects on PstI of several inhibitors have been studied with the
irreversible inhibition kinetic theory of single substrate reaction provided by Tsou, C.L.
Pyridoxal phosphate (PLP), p-chloromercuribenzoic acid (PCMB), diisopropyl fluorophosphates
(DFP), 2,3-diacetyl (DAC) and N-ethyl-5-phenylisoxazolium-3' sulfonate (Woodward's reagent
K, WRK), modify the lysine, cysteine, serine, arginine and carboxyl groups of the protein
molecule respectively.  These five inhibitors have been found to inhibit both the prime
activity and star activity of PstI.  Used with the irreversible inhibition theory, the
apparent inhibition rate constant, A, and the microcosmic inhibition rate constants, k+o and
k'+o of every inhibitor were calculated.  We also found that their inhibition effects belong
to the noncompetitive irreversible inhibition.  Results show that among the groups to be
modified, some have nothing to do with the combination with the substrate, and some may have,
but none of them is the only factor involved in the specific binding.  Despite all this, they
may take part in the catalysis of enzyme or have important effects on maintaining the active
structure of enzyme molecules.  Furthermore, serine and arginine residues are related to the
alteration of PstI conformation and then influence the ability of PstI recognizing the
incising DNA specifically.

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<1>Zou, G.-L., Cao, X.-W., Zhu, R.-F.
<2>The purification and some properties of restriction endonuclease PstI.
<3>Acta Biochim. Biophys. Sin.
<4>17
<5>268-274
<6>1985
<7>Restriction endonuclease PstI has been isolated and purified by chromatography
on heparin-Sepharose, DEAE-cellulose, phosphocellulose, Cibacron Blue
F3GA-Sepharose and hydroxylapatite.  The purified enzyme was found to be
homogeneous as judged by polyacrylamide gel disc electrophoresis.  The specific
activity of the enzyme is greater than 57,000 units per mg protein.  The enzyme
has a molecular weight of about 54,000 daltons by Sephadex G-100 gel
filtration, that of the enzyme subunit is about 17,500 daltons as assayed by
sodium dodecyl sulphate-polyacrylamide gel electrophoresis.  On electrophoresis
the subunit migrated as a single band.  The result shows that restriction
endonuclease PstI consists of two similar subunits.  The N-terminal amino acid
of the enzyme is proline as assayed by dansyl chloride.  Other properties of
restriction endonuclease PstI were also given preliminary study.

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<1>Zou, G.-L., Gao, C.-Z., Pi, X.-C., Hu, W.J.
<2>Studies on the star activity of restriction endonuclease PstI.
<3>Wuhan Daxue Xuebao
<4>45
<5>873-875
<6>1999
<7>The influence of factors such as temperature, enzyme concentration, reaction time and several
organic solvents on the star activity of PstI has been studied.  Under the reaction condition
beneficial for the star activity, the Km and Vmax of the star activity and prime activity
respectively are 1.46 x 10^-7 mol/L, 1.02 x 10^-8 mol/L and 2.86 x 10^-16 mol/s, 3.06 x 10^-15
mol/s.

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<1>Zou, G.-l., Gao, C.-Z., Pi, X.-C., Zhang, J.-J.
<2>Alteration of the specificity of PstI restriction endonuclease.
<3>Wuhan Univ. J. Nat. Sci.
<4>5
<5>361-365
<6>2000
<7>The influence of factors on the substrate-specificity of PstI restriction endonuclease has
been studied with the method of electrophoresis.  The results show that, the specificity of
PstI almost cannot be influenced by the single alteration of the concentration of Tris.HCl,
Mg2+ or Na+ in the reaction system, but it can be altered by the reduction of any two of them.
The specificity cannot be altered by the single alteration of pH or the replacement of Mg2+
with Mn2+.  The addition of glycerol or dimethylsulphoxide (DM-SO) to the reaction system
results in the relaxation of the substrate-specificity of PstI, but dimethyl-methylformide,
glycol and ethyl alcohol cannot bring about the alteration of PstI specificity.  Through the
method of cloning and sequencing, the nucleotides of No. 1 and 6 in the recognition sequence
of PstI have changed (1C-A or 6G-T).  Used with the enzyme analysis of an artificially
synthetic DNA segment containing a special sequence, the nucleotides of No. 1 and 6 have both
changed (1C-A and 6G-T).  The recognition sequence of PstI is speculated to be changed from
CTGCA/G to TGCA/.

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<1>Zou, S., Zhang, M., Hong, J., Ma, Y., Zhang, W.
<2>Comparison of the electro transformation of plasmids and plasmid stability between Zymomonas mobilis ZM4 and CP4.
<3>Afr. J. Microbiol. Res.
<4>5
<5>2026-2033
<6>2011
<7>Four different plasmids were electro transformed into Zymomonas mobilis ZM4 and CP4, two
important ethanol-producing strains. The results
showed that the best source strain for preparing plasmids was the
transformed host strain itself, and Escherichia coli JM110 as the
source strain could yield significantly higher transformation
efficiencies than Top10. The optimal recovery time of transformed ZM4
or CP4 cells to obtain maximum number of transformants and highest
transformation efficiency was 11 h for pZB21-mini, pZB21 and pZA22, but
24 or 20 h for pBBR1MCS-2. The optimal electric field strength for
pZB21-mini was 13.25 kV /cm in ZM4 and 14.0 kV /cm in CP4. But for
pZA22 and pBBR1MCS-2, it was 11.75 kV /cm in ZM4 and 12.5 kV /cm in
CP4; for pZB21, also 12.5 kV /cm in CP4. These plasmids were shown to
be more stable in ZM4 than in CP4 by serial transfer to antibiotic-free
medium and the 3 plasmids were more stable than pBBR1MCS-2. The results
will help to support the genetic and biotechnological research of Z.
mobilis by providing information about some of the most important
factors that influence the transformation of ZM4 and CP4, and also
providing insights into the similarities and differences in their
restriction-modification (R-M) systems.

<>

<1>Zubair, S., de Villiers, E.P., Fuxelius, H.H., Andersson, G., Johansson, K.E., Bishop, R.P., Bongcam-Rudloff, E.
<2>Genome Sequence of Streptococcus agalactiae Strain 09mas018883, Isolated from a Swedish Cow.
<3>Genome Announcements
<4>1
<5>e00456-13
<6>2013
<7>We announce the complete genome sequence of Streptococcus agalactiae strain 09mas018883,
isolated from the milk of a cow with clinical mastitis. The
availability of this genome may allow identification of candidate genes, leading
to discovery of antigens that might form the basis for development of a vaccine
as an alternative means of mastitis control.

<>

<1>Zubair, S., de Villiers, E.P., Younan, M., Andersson, G., Tettelin, H., Riley, D.R., Jores, J., Bongcam-Rudloff, E., Bishop, R.P.
<2>Genome Sequences of Two Pathogenic Streptococcus agalactiae Isolates from the One-Humped Camel Camelus dromedarius.
<3>Genome Announcements
<4>1
<5>e00515-13
<6>2013
<7>Streptococcus agalactiae causes a range of clinical syndromes in camels (Camelus
dromedarius). We report the genome sequences of two S. agalactiae isolates that
induce abscesses in Kenyan camels. These genomes provide novel data on the
composition of the S. agalactiae 'pan genome' and reveal the presence of multiple
genomic islands.

<>

<1>Zueva, V.S., Dmitrenko, O.A., Gladkova, K.K., Zueva, E.A.
<2>New collection of phages for typing methicillin-resistant Staphylococcus aureus.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>2
<5>20-23
<6>1994
<7>A new collection of phages for typing methicillin-resistant S. aureus is proposed.  The
collection includes phage 85 (modified by the MRSA strain), capable of selecting stains with
the similar specificity of the restriction-modification system, and 9 MRSA-induced phages.
The latter differentiate MRSA strains according to the specificity of prophages present in
bacterial cells.  The use of this phage collection has permitted the typing of MRSA strains
insensitive to the phages of the international collection.  Among these cultures on epidemic
strain has been detected and the source of its spread in the burn center has been established.

<>

<1>Zueva, V.S., Dmitrienko, O.A., Krupina, E.A., Belikov, N.G., Nesterenko, L.N.
<2>To the mechanism of the resistance of methicillin-resistant staphylococcus aureus to phages of the international collection.
<3>Zh. Mikrobiol. Epidemiol. Immunobiol.
<4>10
<5>11-15
<6>1990
<7>The resistance of methicillin-resistant staphylococci to phage 85 is due to the
presence of a certain system restriction modification in microbial cells.  The
loss of the capacity for restricting phage DNA by the cell as the consequence
of the loss of the mec determinant is not accompanied by the loss of its
capacity for modifying phage DNA.

<>

<1>Zulkifli, M.H., Teh, L.K., Lee, L.S., Zakaria, Z.A., Salleh, M.Z.
<2>Draft Genome Sequence of Klebsiella pneumoniae Isolate PR04.
<3>Genome Announcements
<4>1
<5>e00418-13
<6>2013
<7>Klebsiella pneumoniae PR04 was isolated from a patient hospitalized in Malaysia.  The draft
genome sequence of K. pneumoniae PR04 shows differences compared to the
reference sequences of K. pneumoniae strains MGH 78578 and NTUH-K2044 in terms of
their genomic structures.

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<1>Zuo, F., Feng, X., Sun, X., Du, C., Chen, S.
<2>Characterization of Plasmid pML21 of Enterococcus faecalis ML21 from Koumiss.
<3>Curr. Microbiol.
<4>66
<5>103-105
<6>2013
<7>
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<1>Zurawski, D.V., Thompson, M.G., McQueary, C.N., Matalka, M.N., Sahl, J.W., Craft, D.W., Rasko, D.A.
<2>Genome Sequences of Four Divergent Multidrug-Resistant Acinetobacter baumannii Strains Isolated from Patients with Sepsis or Osteomyelitis.
<3>J. Bacteriol.
<4>194
<5>1619-1620
<6>2012
<7>Acinetobacter baumannii is a Gram-negative bacterium that causes nosocomial infections
worldwide, with recent prevalence and higher frequency in wounded
military personnel. Four A. baumannii strains from the Walter Reed Army Medical
Center (WRAMC) isolated between 2008 and 2009 were sequenced, representing
diverse, multidrug-resistant isolates from osteomyelitis or septic patients.

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<1>Zweiger, G., Marczynski, G., Shapiro, L.
<2>A Caulobacter DNA methyltransferase that functions only in the predivisional cell.
<3>J. Mol. Biol.
<4>235
<5>472-474
<6>1994
<7>Caulobacter crescentus was found to have a DNA methyltransferase CcrM, that methylates the
adenine base of the HinfI recognition sequence, GANTC. The ccrM gene was cloned, and DNA
sequence analysis revealed that the predicted amino acid sequence has 49% identity with the
Haemophilus influenzae methyltransferase HinfM. Expression of the ccrM gene was found to be
restricted to the portion of the cell cycle immediately prior to cell division. At three
separate chromosomal sites the CcrM recognition sequence is fully methylated in swarmer cells,
becomes hemimethylated upon DNA replication in stalked cells, and does not become remethylated
until just prior to cell division. The time of methyltransferase expression coincides with the
time of methylation of these three chromosomal sites and of plasmid DNA in the predivisional
cell. When ccrM gene expression is placed under control of a constitutive promoter, these
chromosomal sites are fully methylated throughout the cell cycle. A high proportion of
morphologically aberrant cells, and cells that have undergone an additional chromosome
replication initiation, are found in this population. Thus, the temporal control of this
methyltransferase appears to contribute to the accurate cell-cycle control of DNA replication
and cellular morphology.

<>

<1>Zwick, M.E., Joseph, S.J., Didelot, X., Chen, P.E., Bishop-Lilly, K.A., Stewart, A.C., Willner, K., Nolan, N., Lentz, S., Thomason, M.K., Sozhamannan, S., Mateczun, A.J., Du, L., Read, T.D.
<2>Genomic characterization of the Bacillus cereus sensu lato species: Backdrop to the evolution of Bacillus anthracis.
<3>Genome Res.
<4>22
<5>1512-1524
<6>2012
<7>The key genes required for Bacillus anthracis to cause anthrax have been acquired
recently by horizontal gene transfer. To understand the genetic background for
the evolution of B. anthracis virulence, we obtained high-redundancy genome
sequences of 45 strains of the Bacillus cereus sensu lato (s.l.) species that
were chosen for their genetic diversity within the species based on the existing
multilocus sequence typing scheme. From the resulting data, we called more than
324,000 new genes representing more than 12,333 new gene families for this group.
The core genome size for the B. cereus s.l. group was approximately 1750 genes,
with another 2150 genes found in almost every genome constituting the extended
core. There was a paucity of genes specific and conserved in any clade. We found
no evidence of recent large-scale gene loss in B. anthracis or for unusual
accumulation of nonsynonymous DNA substitutions in the chromosome; however,
several B. cereus genomes isolated from soil and not previously associated with
human disease were degraded to various degrees. Although B. anthracis has
undergone an ecological shift within the species, its chromosome does not appear
to be exceptional on a macroscopic scale compared with close relatives.

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<1>Zylicz-Stachula, A., Bujnicki, J.M., Skowron, P.M.
<2>Cloning and analysis of a bifunctional methyltransferase/restriction endonuclease TspGWI, the prototype of a Thermus sp. enzyme family.
<3>BMC Mol. Biol.
<4>10
<5>52
<6>2009
<7>ABSTRACT: BACKGROUND: Restriction-modification systems are a diverse class of enzymes. They
are classified into four major types: I, II, III and IV.
We have previously proposed the existence of a Thermus sp. enzyme family,
which belongs to type II restriction endonucleases (REases), however, it
features also some characteristics of types I and III. Members include
related thermophilic endonucleases: TspGWI, TaqII, TspDTI, and Tth111II.
RESULTS: Here we describe cloning, mutagenesis and analysis of the
prototype TspGWI enzyme that recognises the 5'-ACGGA-3' site and cleaves
11/9 nt downstream. We cloned, expressed, and mutagenised the tspgwi gene
and investigated the properties of its product, the bifunctional TspGWI
restriction/modification enzyme. Since TspGWI does not cleave DNA
completely, a cloning method was devised, based on amino acid sequencing
of internal proteolytic fragments. The deduced amino acid sequence of the
enzyme shares significant sequence similarity with another representative
of the Thermus sp. family - TaqII. Interestingly, these enzymes recognise
similar, yet different sequences in the DNA. Both enzymes cleave DNA at
the same distance, but differ in their ability to cleave single sites and
in the requirement of S-adenosylmethionine as an allosteric activator for
cleavage. Both the restriction endonuclease (REase) and methyltransferase
(MTase) activities of wild type (wt) TspGWI (either recombinant or
isolated from Thermus sp.) are dependent on the presence of divalent
cations. CONCLUSIONS: TspGWI is a bifunctional protein comprising a tandem
arrangement of Type I-like domains; particularly noticeable is the central
HsdM-like module comprising a helical domain and a highly conserved
S-adenosylmethionine-binding/catalytic MTase domain, containing DPAVGTG
and NPPY motifs. TspGWI also possesses an N-terminal PD-(D/E)XK nuclease
domain related to the corresponding domains in HsdR subunits, but lacks
the ATP-dependent translocase module of the HsdR subunit and the
additional domains that are involved in subunit-subunit interactions in
Type I systems. The MTase and REase activities of TspGWI are autonomous
and can be uncoupled. Structurally and functionally, the TspGWI protomer
appears to be a streamlined 'half' of a Type I enzyme.

<>

<1>Zylicz-Stachula, A., Harasimowicz-Slowinska, R.I., Sobolewski, I., Skowron, P.M.
<2>TspGWI, a thermophilic class-IIS restriction endonuclease from Thermus sp., recognizes novel asymmetric sequence 5'-ACGGA(N11/9)-3'.
<3>Nucleic Acids Res.
<4>30
<5>e33
<6>2002
<7>A novel prototype class-IIS restriction endonuclease, TspGWI, was isolated from the
thermophilic bacterium Thermus sp. GW. The recognition sequence and cleavage positions have
been established: TspGWI recognizes the non-palindromic 5-bp sequence 5'-ACGGA-3' and
cleaves the DNA 11 and 9 nt downstream in the top and bottom strand, respectively. In
addition, an accompanying endonuclease, TspGWII, an isoschizomer of PstI, was found in Thermus
sp. GW cells.

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<1>Zylicz-Stachula, A., Jezewska-Frackowiak, J., Skowron, P.M.
<2>Cofactor analogue-induced chemical reactivation of endonuclease activity in a DNA cleavage/methylation deficient TspGWI N(473)A variant in the NPPY motif.
<3>Mol. Biol. Rep.
<4>41
<5>2313-2323
<6>2014
<7>We reported previously that TspGWI, a prototype enzyme of a new Thermus sp. family of
restriction endonucleases-methyltransferases (REases-MTases), undergoes the novel phenomenon
of sinefungin (SIN)-caused specificity transition. Here we investigated mutant TspGWI N(473)A,
containing a single amino acid (aa) substitution in the NPPY motif of the MTase. Even though
the aa substitution is located within the MTase polypeptide segment, DNA cleavage and
modification are almost completely abolished, indicating that the REase and MTase are
intertwined. Remarkably, the TspGWI N(473)A REase functionality can be completely
reconstituted by the addition of SIN. We hypothesize that SIN binds specifically to the enzyme
and restores the DNA cleavage-competent protein tertiary structure. This indicates the
significant role of allosteric effectors in DNA cleavage in Thermus sp. enzymes. This is the
first case of REase mutation suppression by an S-adenosylmethionine (SAM) cofactor analogue.
Moreover, the TspGWI N(473)A clone strongly affects E. coli division control, acting as a
'selfish gene'. The mutant lacks the competing MTase activity and therefore might be useful
for applications in DNA manipulation. Here we present a case study of a novel strategy for
REase activity/specificity alteration by a single aa substitution, based on the bioinformatic
analysis of active motif locations, combining (a) aa sequence engineering (b) the alteration
of protein enzymatic properties, and (c) the use of cofactor-analogue cleavage reconstitution
and stimulation.

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<1>Zylicz-Stachula, A., Zolnierkiewicz, O., Lubys, A., Ramanauskaite, D., Mitkaite, G., Bujnicki, J.M., Skowron, P.M.
<2>Related bifunctional restriction endonuclease-methyltransferase triplets: TspDTI, Tth111II/TthHB27I and TsoI with distinct specificities.
<3>BMC Mol. Biol.
<4>13
<5>13
<6>2012
<7>Background: We previously defined a family of restriction endonucleases (REases) from Thermus
sp., which share common biochemical and
biophysical features, such as the fusion of both the nuclease and
methyltransferase (MTase) activities in a single polypeptide, cleavage
at a distance from the recognition site, large molecular size,
modulation of activity by S-adenosylmethionine (SAM), and incomplete
cleavage of the substrate DNA. Members include related thermophilic
REases with five distinct specificities: TspGWI, TaqII,
Tth111II/TthHB27I, TspDTI and TsoI.
Results: TspDTI, TsoI and isoschizomers Tth111II/TthHB27I recognize
different, but related sequences: 5'-ATGAA-3', 5'-TARCCA-3' and
5'-CAARCA-3' respectively. Their amino acid sequences are similar,
which is unusual among REases of different specificity. To gain insight
into this group of REases, TspDTI, the prototype member of the Thermus
sp. enzyme family, was cloned and characterized using a recently
developed method for partially cleaving REases.
Conclusions: TspDTI, TsoI and isoschizomers Tth111II/TthHB27I are
closely related bifunctional enzymes. They comprise a tandem
arrangement of Type I-like domains, like other Type IIC enzymes (those
with a fusion of a REase and MTase domains), e. g. TspGWI, TaqII and
MmeI, but their sequences are only remotely similar to these previously
characterized enzymes. The characterization of TspDTI, a prototype
member of this group, extends our understanding of sequence-function
relationships among multifunctional restriction-modification enzymes.

<>

<1>Zylicz-Stachula, A., Zolnierkiewicz, O., Sliwinska, K., Jezewska-Frackowiak, J., Skowron, P.M.
<2>Bifunctional TaqII restriction endonuclease: redefining the prototype DNA recognition site and establishing the Fidelity Index for partial cleaving.
<3>BMC Biochem.
<4>12
<5>62
<6>2011
<7>ABSTRACT: BACKGROUND: The TaqII enzyme is a member of the Thermus sp. enzyme family that we
propounded previously within Type IIS restriction endonucleases,
containing related thermophilic bifunctional endonucleases-methyltransferases
from various Thermus sp.: TaqII, Tth111II, TthHB27I, TspGWI, TspDTI and TsoI.
These enzymes show significant nucleotide and amino acid sequence similarities, a
rare phenomenon among restriction endonucleases, along with similarities in
biochemical properties, molecular size, DNA recognition sequences and cleavage
sites. They also feature some characteristics of Types I and III. RESULTS: Barker
et al. reported the Type IIS/IIC restriction endonuclease TaqII as recognizing
two distinct cognate site variants (5'-GACCGA-3' and 5'-CACCCA-3') while cleaving
11/9 nucleotides downstream. We used four independent methods, namely, shotgun
cloning and sequencing, restriction pattern analysis, digestion of particular
custom substrates and GeneScan analysis, to demonstrate that the recombinant
enzyme recognizes only 5'-GACCGA-3' sites and cleaves 11/9 nucleotides
downstream. We did not observe any 5'-CACCCA-3' cleavage under a variety of
conditions and site arrangements tested. We also characterized the enzyme
biochemically and established new digestion conditions optimal for practical
enzyme applications. Finally, we developed and propose a new version of the
Fidelity Index - the Fidelity Index for Partial Cleavage (FI-PC). CONCLUSIONS:
The DNA recognition sequence of the bifunctional prototype TaqII
endonuclease-methyltransferase from Thermus aquaticus has been redefined as
recognizing only 5'-GACCGA-3' cognate sites. The reaction conditions (pH and salt
concentrations) were designed either to minimize (pH = 8.0 and 10 mM ammonium
sulphate) or to enhance star activity (pH = 6.0 and no salt). Redefinition of the
recognition site and reaction conditions makes this prototype endonuclease a
useful tool for DNA manipulation; as yet, this enzyme has no practical
applications. The extension of the Fidelity Index will be helpful for DNA
manipulation with enzymes only partially cleaving DNA.

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<1>Zylicz-Stachula, A., Zolnierkiewicz, O., Sliwinska, K., Jezewska-Frackowiak, J., Skowron, P.M.
<2>Modified 'one amino acid-one codon' engineering of high GC content TaqII-coding gene from thermophilic Thermus aquaticus results in radical expression increase.
<3>Microb. Cell Fact.
<4>13
<5>7
<6>2014
<7>BACKGROUND: An industrial approach to protein production demands maximization of
cloned gene expression, balanced with the recombinant host's viability.
Expression of toxic genes from thermophiles poses particular difficulties due to
high GC content, mRNA secondary structures, rare codon usage and impairing the
host's coding plasmid replication.TaqII belongs to a family of bifunctional
enzymes, which are a fusion of the restriction endonuclease (REase) and
methyltransferase (MTase) activities in a single polypeptide. The family contains
thermostable REases with distinct specificities: TspGWI, TaqII,
Tth111II/TthHB27I, TspDTI and TsoI and a few enzymes found in mesophiles. While
not being isoschizomers, the enzymes exhibit amino acid (aa) sequence homologies,
having molecular sizes of ~120 kDa share common modular architecture, resemble
Type-I enzymes, cleave DNA 11/9 nt from the recognition sites, their activity is
affected by S-adenosylmethionine (SAM). RESULTS: We describe the taqIIRM gene
design, cloning and expression of the prototype TaqII. The enzyme amount in
natural hosts is extremely low. To improve expression of the taqIIRM gene in
Escherichia coli (E. coli), we designed and cloned a fully synthetic, low GC
content, low mRNA secondary structure taqIIRM, codon-optimized gene under a
bacteriophage lambda (lambda) PR promoter. Codon usage based on a modified 'one
amino acid-one codon' strategy, weighted towards low GC content codons, resulted
in approximately 10-fold higher expression of the synthetic gene. 718 codons of
total 1105 were changed, comprising 65% of the taqIIRM gene. The reason for we
choose a less effective strategy rather than a resulting in high expression
yields 'codon randomization' strategy, was intentional, sub-optimal TaqII in vivo
production, in order to decrease the high 'toxicity' of the REase-MTase protein.
CONCLUSIONS: Recombinant wt and synthetic taqIIRM gene were cloned and expressed
in E. coli. The modified 'one amino acid-one codon' method tuned for
thermophile-coded genes was applied to obtain overexpression of the 'toxic'
taqIIRM gene. The method appears suited for industrial production of thermostable
'toxic' enzymes in E. coli. This novel variant of the method biased toward
increasing a gene's AT content may provide economic benefits for industrial
applications.

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